U.S. patent application number 10/422571 was filed with the patent office on 2004-04-29 for nucleic acids and corresponding proteins entitled 191p4d12(b) useful in treatment and detection of cancer.
Invention is credited to Challita-Eid, Pia M., Faris, Mary, Ge, Wangmao, Jakobovits, Aya, Raitano, Arthur B..
Application Number | 20040083497 10/422571 |
Document ID | / |
Family ID | 31891426 |
Filed Date | 2004-04-29 |
United States Patent
Application |
20040083497 |
Kind Code |
A1 |
Raitano, Arthur B. ; et
al. |
April 29, 2004 |
Nucleic acids and corresponding proteins entitled 191P4D12(b)
useful in treatment and detection of cancer
Abstract
A novel gene 191P4D12(b) and its encoded protein, and variants
thereof, are described wherein 191P4D12(b) exhibits tissue specific
expression in normal adult tissue, and is aberrantly expressed in
the cancers listed in Table I. Consequently, 191P4D12(b) provides a
diagnostic, prognostic, prophylactic and/or therapeutic target for
cancer. The 191P4D12(b) gene or fragment thereof, or its encoded
protein, or variants thereof, or a fragment thereof, can be used to
elicit a humoral or cellular immune response; antibodies or T cells
reactive with 191P4D12(b) can be used in active or passive
immunization.
Inventors: |
Raitano, Arthur B.; (Los
Angeles, CA) ; Challita-Eid, Pia M.; (Encino, CA)
; Jakobovits, Aya; (Beverly Hills, CA) ; Faris,
Mary; (Los Angeles, CA) ; Ge, Wangmao; (Culver
City, CA) |
Correspondence
Address: |
MORRISON & FOERSTER LLP
3811 VALLEY CENTRE DRIVE
SUITE 500
SAN DIEGO
CA
92130-2332
US
|
Family ID: |
31891426 |
Appl. No.: |
10/422571 |
Filed: |
April 23, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
60404306 |
Aug 16, 2002 |
|
|
|
60423290 |
Nov 1, 2002 |
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Current U.S.
Class: |
800/9 ;
424/155.1; 435/320.1; 435/325; 435/6.14; 435/69.1; 435/7.23;
514/44R; 530/350; 536/23.5 |
Current CPC
Class: |
A61P 37/04 20180101;
C07K 14/47 20130101; C07K 14/435 20130101; A61K 38/00 20130101;
A01K 2217/075 20130101; A61K 47/6851 20170801; A61P 35/00 20180101;
Y02A 50/30 20180101; A61K 2039/505 20130101; A61P 43/00 20180101;
C07K 2319/00 20130101; C12N 5/0693 20130101; A01K 2217/05 20130101;
Y02A 50/466 20180101; G01N 33/57484 20130101 |
Class at
Publication: |
800/009 ;
424/155.1; 435/006; 435/007.23; 435/069.1; 435/320.1; 435/325;
514/044; 536/023.5; 530/350 |
International
Class: |
C12Q 001/68; G01N
033/574; A01K 067/00; C12P 021/02; C12N 005/06; A61K 048/00; C07K
014/47 |
Claims
1. A composition that comprises, consists essentially of, or
consists of: a) a peptide of eight, nine, ten, or eleven contiguous
amino acids of a protein of FIG. 2; b) a peptide of Tables
VIII-XXI; c) a peptide of Tables XXII to XLV; or, d) a peptide of
Tables XLVI to XLIX.
2. A composition of claim 1 that comprises a protein related to a
protein of FIG. 2.
3. A protein of claim 2 that is at least 90, 91, 92, 93, 94, 95,
96, 97, 98, or 99% homologous to an entire amino acid sequence
shown in FIG. 2.
4. A composition of claim 1 wherein the substance comprises a CTL
polypeptide or an analog thereof, from the amino acid sequence of a
protein of FIG. 2.
5. A composition of claim 4 further limited by a proviso that the
epitope is not an entire amino acid sequence of FIG. 2.
6. A composition of claim 1 further limited by a proviso that the
polypeptide is not an entire amino acid sequence of a protein of
FIG. 2.
7. A composition of claim 1 that comprises an antibody polypeptide
epitope from an amino acid sequence of FIG. 2.
8. A composition of claim 7 further limited by a proviso that the
epitope is not an entire amino acid sequence of FIG. 2.
9. A composition of claim 7 wherein the antibody epitope comprises
a peptide region of at least 5 amino acids of FIG. 2 in any whole
number increment up to the end of said peptide, wherein the epitope
comprises an amino acid position selected from: a) an amino acid
position having a value greater than 0.5 in the Hydrophilicity
profile of FIG. 5, b) an amino acid position having a value less
than 0.5 in the Hydropathicity profile of FIG. 6; c) an amino acid
position having a value greater than 0.5 in the Percent Accessible
Residues profile of FIG. 7; d) an amino acid position having a
value greater than 0.5 in the Average Flexibility profile of FIG.
8; e) an amino acid position having a value greater than 0.5 in the
Beta-turn profile of FIG. 9; f) a combination of at least two of a)
through e); g) a combination of at least three of a) through e); h)
a combination of at least four of a) through e); or i) a
combination of five of a) through e).
10. A polynucleotide that encodes a protein of claim 1.
11. A polynucleotide of claim 10 that comprises a nucleic acid
molecule set forth in FIG. 2.
12. A polynucleotide of claim 10 further limited by a proviso that
the encoded protein is not an entire amino acid sequence of FIG.
2.
13. A composition of claim 11 wherein the substance comprises a
polynucleotide that comprises a coding sequence of a nucleic acid
sequence of FIG. 2.
14. A polynucleotide of claim 11 that further comprises an
additional nucleotide sequence that encodes an additional peptide
of claim 1.
15. A composition comprising a polynucleotide that is fully
complementary to a polynucleotide of claim 10.
16. A method of generating a mammalian immune response directed to
a protein of FIG. 2, the method comprising: exposing cells of the
mammal's immune system to a portion of a) a 191P4D12(b)-related
protein and/or b) a nucleotide sequence that encodes said protein,
whereby an immune response is generated to said protein.
17. A method of generating an immune response of claim 16, said
method comprising: providing a 191P4D12(b)-related protein that
comprises at least one T cell or at least one B cell epitope; and,
contacting the epitope with a mammalian immune system T cell or B
cell respectively, whereby the T cell or B cell is activated.
18. A method of claim 17 wherein the immune system cell is a B
cell, whereby the induced B cell generates antibodies that
specifically bind to the 191P4D12(b)-related protein.
19. A method of claim 17 wherein the immune system cell is a T cell
that is a cytotoxic T cell (CTL), whereby the activated CTL kills
an autologous cell that expresses the 191P4D12(b)-related
protein.
20. A method of claim 17 wherein the immune system cell is a T cell
that is a helper T cell (HTL), whereby the activated HTL secretes
cytokines that facilitate the cytotoxic activity of a cytotoxic T
cell (CTL) or the antibody-producing activity of a B cell.
21. Amethod for detecting, in a sample, the presence of a
191P4D12(b)-related protein or a 191P4D12(b)-related
polynucleotide, comprising steps of: contacting the sample with a
substance that specifically binds to the 191P4D12(b)-related
protein or to the 191P4D12(b)-related polynucleotide, respectively;
and, determining that there is a complex of the substance with the
191P4D12(b)-related protein or the substance with the
191P4D12(b)-related polynucleotide, respectively.
22. A method of claim 21 for detecting the presence of a
191P4D12(b)-related protein in a sample comprising steps of:
contacting the sample with an antibody or fragment thereof either
of which specifically bind to the 191P4D12(b)-related protein; and,
determining that there is a complex of the antibody or fragment
thereof and the 191P4D12(b)-related protein.
23. A method of claim 21 further comprising a step of: taking the
sample from a patient who has or who is suspected of having
cancer.
24. A method of claim 21 for detecting the presence of a protein of
FIG. 2 mRNA in a sample comprising: producing cDNA from the sample
by reverse transcription using at least one primer; amplifying the
cDNA so produced using 191P4D12(b) polynucleotides as sense and
antisense primers, wherein the 191P4D12(b) polynucleotides used as
the sense and antisense primers serve to amplify a 191P4D12(b)
cDNA; and, detecting the presence of the amplified 191P4D12(b)
cDNA.
25. A method of claim 21 for monitoring one or more 191P4D12(b)
gene products in a biological sample from a patient who has or who
is suspected of having cancer, the method comprising: determining
the status of one or more 191P4D12(b) gene products expressed by
cells in a tissue sample from an individual; comparing the status
so determined to the status of one or more 191P4D12(b) gene
products in a corresponding normal sample; and, identifying the
presence of one or more aberrant gene products of 191P4D12(b) in
the sample relative to the normal sample.
26. The method of claim 25 further comprising a step of determining
if there are one or more elevated gene products of a 191P4D12(b)
mRNA or a 191P4D12(b) protein, whereby the presence of one or more
elevated gene products in the test sample relative to the normal
tissue sample indicates the presence or status of a cancer.
27. A method of claim 26 wherein the cancer occurs in a tissue set
forth in Table I.
28. A composition comprising: a substance that a) modulates the
status of a protein of FIG. 2, or b) a molecule that is modulated
by a protein of FIG. 2, whereby the status of a cell that expresses
a protein of FIG. 2 is modulated.
29. A composition of claim 28, further comprising a physiologically
acceptable carrier.
30. A pharmaceutical composition that comprises the composition of
claim 28 in a human unit dose form.
31. A composition of claim 28 wherein the substance comprises an
antibody or fragment thereof that specifically binds to a protein
of FIG. 2.
32. An antibody or fragment thereof of claim 31, which is
monoclonal.
33. An antibody of claim 31, which is a human antibody, a humanized
antibody or a chimeric antibody.
34. A non-human transgenic animal that produces an antibody of
claim 31.
35. A hybridoma that produces an antibody of claim 32.
36. A method of delivering a cytotoxic agent or a diagnostic agent
to a cell that expresses a protein of FIG. 2, said method
comprising: providing the cytotoxic agent or the diagnostic agent
conjugated to an antibody or fragment thereof of claim 4; and,
exposing the cell to the antibody-agent or fragment-agent
conjugate.
37. A composition of claim 28 wherein the substance comprises a
polynucleotide that encodes an antibody or fragment thereof, either
of which immunospecifically bind to a protein of FIG. 2.
38. A composition of claim 28 wherein the substance comprises a) a
ribozyme that cleaves a polynucleotide having a 191P4D12(b) coding
sequence, or b) a nucleic acid molecule that encodes the ribozyme;
and, a physiologically acceptable carrier.
39. A composition of claim 28 wherein the substance comprises human
T cells, wherein said T cells specifically recognize a 191P4D12(b)
peptide subsequence in the context of a particular HLA
molecule.
40. A method of inhibiting growth of cancer cells that express a
protein of FIG. 2, the method comprising: administering to the
cells the composition of claim 28.
41. A method of claim 40 of inhibiting growth of cancer cells that
express a protein of FIG. 2, the method comprising steps of:
administering to said cells an antibody or fragment thereof, either
of which specifically bind to a 191P4D12(b)-related protein.
42. A method of claim 40 of inhibiting growth of cancer cells that
express a protein of FIG. 2, the method comprising steps of:
administering to said cells a 191P4D12(b)-related protein.
43. A method of claim 40 of inhibiting growth of cancer cells that
express a protein of FIG. 2, the method comprising steps of:
administering to said cells a polynucleotide comprising a coding
sequence for a 191P4D12(b)-related protein or comprising a
polynucleotide complementary to a coding sequence for a
191P4D12(b)-related protein.
44. A method of claim 40 of inhibiting growth of cancer cells that
express a protein of FIG. 2, the method comprising steps of:
administering to said cells a ribozyme that cleaves a
polynucleotide that encodes a protein of FIG. 2.
45. A method of claim 40 of inhibiting growth of cancer cells that
express a protein of FIG. 2 and a particular HLA molecule, the
method comprising steps of: administering human T cells to said
cancer cells, wherein said T cells specifically recognize a peptide
subsequence of a protein of FIG. 2 while the subsequence is in the
context of the particular HLA molecule.
46. A method of claim 40, the method comprising steps of:
administering a vector that delivers a nucleotide that encodes a
single chain monoclonal antibody, whereby the encoded single chain
antibody is expressed intracellularly within cancer cells that
express a protein of FIG. 2.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a non-provisional utility patent
application that claims priority from U.S. provisional patent
application U.S. S No. 60/404,306, filed 16 Aug. 2002 and this
application claims priority from U.S. provisional patent
application U.S. S No. 60/423,290, filed 01 Nov. 2002. The contents
of the applications listed in this paragraph are fully incorporated
by reference herein.
STATEMENT OF RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSORED
RESEARCH
[0002] Not applicable.
FIELD OF THE INVENTION
[0003] The invention described herein relates to genes and their
encoded proteins, termed 191P4D12(b), expressed in certain cancers,
and to diagnostic and therapeutic methods and compositions useful
in the management of cancers that express 191P4D12(b).
BACKGROUND OF THE INVENTION
[0004] Cancer is the second leading cause of human death next to
coronary disease. Worldwide, millions of people die from cancer
every year. In the United States alone, as reported by the American
Cancer Society, cancer causes the death of well over a half-million
people annually, with over 1.2 million new cases diagnosed per
year. While deaths from heart disease have been declining
significantly, those resulting from cancer generally are on the
rise. In the early part of the next century, cancer is predicted to
become the leading cause of death.
[0005] Worldwide, several cancers stand out as the leading killers.
In particular, carcinomas of the lung, prostate, breast, colon,
pancreas, and ovary represent the primary causes of cancer death.
These and virtually all other carcinomas share a common lethal
feature. With very few exceptions, metastatic disease from a
carcinoma is fatal. Moreover, even for those cancer patients who
initially survive their primary cancers, common experience has
shown that their lives are dramatically altered. Many cancer
patients experience strong anxieties driven by the awareness of the
potential for recurrence or treatment failure. Many cancer patients
experience physical debilitations following treatment. Furthermore,
many cancer patients experience a recurrence.
[0006] Worldwide, prostate cancer is the fourth most prevalent
cancer in men. In North America and Northern Europe, it is by far
the most common cancer in males and is the second leading cause of
cancer death in men. In the United States alone, well over 30,000
men die annually of this disease--second only to lung cancer.
Despite the magnitude of these figures, there is still no effective
treatment for metastatic prostate cancer. Surgical prostatectomy,
radiation therapy, hormone ablation therapy, surgical castration
and chemotherapy continue to be the main treatment modalities.
Unfortunately, these treatments are ineffective for many and are
often associated with undesirable consequences.
[0007] On the diagnostic front, the lack of a prostate tumor marker
that can accurately detect early-stage, localized tumors remains a
significant limitation in the diagnosis and management of this
disease. Although the serum prostate specific antigen (PSA) assay
has been a very useful tool, however its specificity and general
utility is widely regarded as lacking in several important
respects.
[0008] Progress in identifying additional specific markers for
prostate cancer has been improved by the generation of prostate
cancer xenografts that can recapitulate different stages of the
disease in mice. The LAPC (Los Angeles Prostate Cancer) xenografts
are prostate cancer xenografts that have survived passage in severe
combined immune deficient (SCID) mice and have exhibited the
capacity to mimic the transition from androgen dependence to
androgen independence (Klein et al., 1997, Nat. Med. 3:402). More
recently identified prostate cancer markers include PCTA-1 (Su et
al., 1996, Proc. Natl. Acad. Sci. USA 93: 7252), prostate-specific
membrane (PSM) antigen (Pinto et al., Clin Cancer Res 1996 Sep. 2
(9): 1445-51), STEAP (Hubert, et al., Proc Natl Acad Sci U S A.
1999 Dec. 7; 96(25): 14523-8) and prostate stem cell antigen (PSCA)
(Reiter et al., 1998, Proc. Natl. Acad. Sci. USA 95: 1735).
[0009] While previously identified markers such as PSA, PSM, PCTA
and PSCA have facilitated efforts to diagnose and treat prostate
cancer, there is need for the identification of additional markers
and therapeutic targets for prostate and related cancers in order
to further improve diagnosis and therapy.
[0010] Renal cell carcinoma (RCC) accounts for approximately 3
percent of adult malignancies. Once adenomas reach a diameter of 2
to 3 cm, malignant potential exists. In the adult, the two
principal malignant renal tumors are renal cell adenocarcinoma and
transitional cell carcinoma of the renal pelvis or ureter. The
incidence of renal cell adenocarcinoma is estimated at more than
29,000 cases in the United States, and more than 11,600 patients
died of this disease in 1998. Transitional cell carcinoma is less
frequent, with an incidence of approximately 500 cases per year in
the United States.
[0011] Surgery has been the primary therapy for renal cell
adenocarcinoma for many decades. Until recently, metastatic disease
has been refractory to any systemic therapy. With recent
developments in systemic therapies, particularly immunotherapies,
metastatic renal cell carcinoma may be approached aggressively in
appropriate patients with a possibility of durable responses.
Nevertheless, there is a remaining need for effective therapies for
these patients.
[0012] Of all new cases of cancer in the United States, bladder
cancer represents approximately 5 percent in men (fifth most common
neoplasm) and 3 percent in women (eighth most common neoplasm). The
incidence is increasing slowly, concurrent with an increasing older
population. In 1998, there was an estimated 54,500 cases, including
39,500 in men and 15,000 in women. The age-adjusted incidence in
the United States is 32 per 100,000 for men and eight per 100,000
in women. The historic male/female ratio of 3:1 may be decreasing
related to smoking patterns in women. There were an estimated
11,000 deaths from bladder cancer in 1998 (7,800 in men and 3,900
in women). Bladder cancer incidence and mortality strongly increase
with age and will be an increasing problem as the population
becomes more elderly.
[0013] Most bladder cancers recur in the bladder. Bladder cancer is
managed with a combination of transurethral resection of the
bladder (TUR) and intravesical chemotherapy or immunotherapy. The
multifocal and recurrent nature of bladder cancer points out the
limitations of TUR. Most muscle-invasive cancers are not cured by
TUR alone. Radical cystectomy and urinary diversion is the most
effective means to eliminate the cancer but carry an undeniable
impact on urinary and sexual function. There continues to be a
significant need for treatment modalities that are beneficial for
bladder cancer patients.
[0014] An estimated 130,200 cases of colorectal cancer occurred in
2000 in the United States, including 93,800 cases of colon cancer
and 36,400 of rectal cancer. Colorectal cancers are the third most
common cancers in men and women. Incidence rates declined
significantly during 1992-1996 (-2.1% per year). Research suggests
that these declines have been due to increased screening and polyp
removal, preventing progression of polyps to invasive cancers.
There were an estimated 56,300 deaths (47,700 from colon cancer,
8,600 from rectal cancer) in 2000, accounting for about 11% of all
U.S. cancer deaths.
[0015] At present, surgery is the most common form of therapy for
colorectal cancer, and for cancers that have not spread, it is
frequently curative. Chemotherapy, or chemotherapy plus radiation,
is given before or after surgery to most patients whose cancer has
deeply perforated the bowel wall or has spread to the lymph nodes.
A permanent colostomy (creation of an abdominal opening for
elimination of body wastes) is occasionally needed for colon cancer
and is infrequently required for rectal cancer. There continues to
be a need for effective diagnostic and treatment modalities for
colorectal cancer.
[0016] There were an estimated 164,100 new cases of lung and
bronchial cancer in 2000, accounting for 14% of all U.S. cancer
diagnoses. The incidence rate of lung and bronchial cancer is
declining significantly in men, from a high of 86.5 per 100,000 in
1984 to 70.0 in 1996. In the 1990s, the rate of increase among
women began to slow. In 1996, the incidence rate in women was 42.3
per 100,000.
[0017] Lung and bronchial cancer caused an estimated 156,900 deaths
in 2000, accounting for 28% of all cancer deaths. During 1992-1996,
mortality from lung cancer declined significantly among men (-1.7%
per year) while rates for women were still significantly increasing
(0.9% per year). Since 1987, more women have died each year of lung
cancer than breast cancer, which, for over 40 years, was the major
cause of cancer death in women. Decreasing lung cancer incidence
and mortality rates most likely resulted from decreased smoking
rates over the previous 30 years; however, decreasing smoking
patterns among women lag behind those of men. Of concern, although
the declines in adult tobacco use have slowed, tobacco use in youth
is increasing again.
[0018] Treatment options for lung and bronchial cancer are
determined by the type and stage of the cancer and include surgery,
radiation therapy, and chemotherapy. For many localized cancers,
surgery is usually the treatment of choice. Because the disease has
usually spread by the time it is discovered, radiation therapy and
chemotherapy are often needed in combination with surgery.
Chemotherapy alone or combined with radiation is the treatment of
choice for small cell lung cancer; on this regimen, a large
percentage of patients experience remission, which in some cases is
long lasting. There is however, an ongoing need for effective
treatment and diagnostic approaches for lung and bronchial
cancers.
[0019] An estimated 182,800 new invasive cases of breast cancer
were expected to occur among women in the United States during
2000. Additionally, about 1,400 new cases of breast cancer were
expected to be diagnosed in men in 2000. After increasing about 4%
per year in the 1980s, breast cancer incidence rates in women have
leveled off in the 1990s to about 110.6 cases per 100,000.
[0020] In the U.S. alone, there were an estimated 41,200 deaths
(40,800 women, 400 men) in 2000 due to breast cancer. Breast cancer
ranks second among cancer deaths in women. According to the most
recent data, mortality rates declined significantly during
1992-1996 with the largest decreases in younger women, both white
and black. These decreases were probably the result of earlier
detection and improved treatment.
[0021] Taking into account the medical circumstances and the
patient's preferences, treatment of breast cancer may involve
lumpectomy (local removal of the tumor) and removal of the lymph
nodes under the arm; mastectomy (surgical removal of the breast)
and removal of the lymph nodes under the arm; radiation therapy;
chemotherapy; or hormone therapy. Often, two or more methods are
used in combination. Numerous studies have shown that, for early
stage disease, long-term survival rates after lumpectomy plus
radiotherapy are similar to survival rates after modified radical
mastectomy. Significant advances in reconstruction techniques
provide several options for breast reconstruction after mastectomy.
Recently, such reconstruction has been done at the same time as the
mastectomy.
[0022] Local excision of ductal carcinoma in situ (DCIS) with
adequate amounts of surrounding normal breast tissue may prevent
the local recurrence of the DCIS. Radiation to the breast and/or
tamoxifen may reduce the chance of DCIS occurring in the remaining
breast tissue. This is important because DCIS, if left untreated,
may develop into invasive breast cancer. Nevertheless, there are
serious side effects or sequelae to these treatments. There is,
therefore, a need for efficacious breast cancer treatments.
[0023] There were an estimated 23,100 new cases of ovarian cancer
in the United States in 2000. It accounts for 4% of all cancers
among women and ranks second among gynecologic cancers. During
1992-1996, ovarian cancer incidence rates were significantly
declining. Consequent to ovarian cancer, there were an estimated
14,000 deaths in 2000. Ovarian cancer causes more deaths than any
other cancer of the female reproductive system.
[0024] Surgery, radiation therapy, and chemotherapy are treatment
options for ovarian cancer. Surgery usually includes the removal of
one or both ovaries, the fallopian tubes (salpingo-oophorectomy),
and the uterus (hysterectomy). In some very early tumors, only the
involved ovary will be removed, especially in young women who wish
to have children. In advanced disease, an attempt is made to remove
all intra-abdominal disease to enhance the effect of chemotherapy.
There continues to be an important need for effective treatment
options for ovarian cancer.
[0025] There were an estimated 28,300 new cases of pancreatic
cancer in the United States in 2000. Over the past 20 years, rates
of pancreatic cancer have declined in men. Rates among women have
remained approximately constant but may be beginning to decline.
Pancreatic cancer caused an estimated 28,200 deaths in 2000 in the
United States. Over the past 20 years, there has been a slight but
significant decrease in mortality rates among men (about --0.9% per
year) while rates have increased slightly among women.
[0026] Surgery, radiation therapy, and chemotherapy are treatment
options for pancreatic cancer. These treatment options can extend
survival and/or relieve symptoms in many patients but are not
likely to produce a cure for most. There is a significant need for
additional therapeutic and diagnostic options for pancreatic
cancer.
SUMMARY OF THE INVENTION
[0027] The present invention relates to a gene, designated
191P4D12(b), that has now been found to be over-expressed in the
cancer(s) listed in Table I. Northern blot expression analysis of
191P4D12(b) gene expression in normal tissues shows a restricted
expression pattern in adult tissues. The nucleotide (FIG. 2) and
amino acid (FIG. 2, and FIG. 3) sequences of 191P4D12(b) are
provided. The tissue-related profile of 191P4D12(b) in normal adult
tissues, combined with the over-expression observed in the tissues
listed in Table I, shows that 191P4D12(b) is aberrantly
over-expressed in at least some cancers, and thus serves as a
useful diagnostic, prophylactic, prognostic, and/or therapeutic
target for cancers of the tissue(s) such as those listed in Table
I.
[0028] The invention provides polynucleotides corresponding or
complementary to all or part of the 191P4D12(b) genes, mRNAs,
and/or coding sequences, preferably in isolated form, including
polynucleotides encoding 191P4D12(b)-related proteins and fragments
of 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, or more than 25 contiguous amino acids; at
least 30, 35, 40, 45, 50, 55, 60, 65, 70, 80, 85, 90, 95, 100 or
more than 100 contiguous amino acids of a 191P4D12(b)-related
protein, as well as the peptides/proteins themselves; DNA, RNA,
DNA/RNA hybrids, and related molecules, polynucleotides or
oligonucleotides complementary or having at least a 90% homology to
the 191P4D12(b) genes or mRNA sequences or parts thereof, and
polynucleotides or oligonucleotides that hybridize to the
191P4D12(b) genes, mRNAs, or to 191P4D12(b)-encoding
polynucleotides. Also provided are means for isolating cDNAs and
the genes encoding 191P4D12(b). Recombinant DNA molecules
containing 191P4D12(b) polynucleotides, cells transformed or
transduced with such molecules, and host-vector systems for the
expression of 191P4D12(b) gene products are also provided. The
invention further provides antibodies that bind to 191P4D12(b)
proteins and polypeptide fragments thereof, including polyclonal
and monoclonal antibodies, murine and other mammalian antibodies,
chimeric antibodies, humanized and fully human antibodies, and
antibodies labeled with a detectable marker or therapeutic agent.
In certain embodiments, there is a proviso that the entire nucleic
acid sequence of FIG. 2 is not encoded and/or the entire amino acid
sequence of FIG. 2 is not prepared. In certain embodiments, the
entire nucleic acid sequence of FIG. 2 is encoded and/or the entire
amino acid sequence of FIG. 2 is prepared, either of which are in
respective human unit dose forms.
[0029] The invention further provides methods for detecting the
presence and status of 191P4D12(b) polynucleotides and proteins in
various biological samples, as well as methods for identifying
cells that express 191P4D12(b). A typical embodiment of this
invention provides methods for monitoring 191P4D12(b) gene products
in a tissue or hematology sample having or suspected of having some
form of growth dysregulation such as cancer.
[0030] The invention further provides various immunogenic or
therapeutic compositions and strategies for treating cancers that
express 191P4D12(b) such as cancers of tissues listed in Table I,
including therapies aimed at inhibiting the transcription,
translation, processing or function of 191P4D12(b) as well as
cancer vaccines. In one aspect, the invention provides
compositions, and methods comprising them, for treating a cancer
that expresses 191P4D12(b) in a human subject wherein the
composition comprises a carrier suitable for human use and a human
unit dose of one or more than one agent that inhibits the
production or function of 191P4D12(b). Preferably, the carrier is a
uniquely human carrier. In another aspect of the invention, the
agent is a moiety that is immunoreactive with 191P4D12(b) protein.
Non-limiting examples of such moieties include, but are not limited
to, antibodies (such as single chain, monoclonal, polyclonal,
humanized, chimeric, or human antibodies), functional equivalents
thereof (whether naturally occurring or synthetic), and
combinations thereof. The antibodies can be conjugated to a
diagnostic or therapeutic moiety. In another aspect, the agent is a
small molecule as defined herein.
[0031] In another aspect, the agent comprises one or more than one
peptide which comprises a cytotoxic T lymphocyte (CTL) epitope that
binds an HLA class I molecule in a human to elicit a CTL response
to 191P4D12(b) and/or one or more than one peptide which comprises
a helper T lymphocyte (HTL) epitope which binds an HLA class II
molecule in a human to elicit an HTL response. The peptides of the
invention may be on the same or on one or more separate polypeptide
molecules. In a further aspect of the invention, the agent
comprises one or more than one nucleic acid molecule that expresses
one or more than one of the CTL or HTL response stimulating
peptides as described above. In yet another aspect of the
invention, the one or more than one nucleic acid molecule may
express a moiety that is immunologically reactive with 191P4D12(b)
as described above. The one or more than one nucleic acid molecule
may also be, or encodes, a molecule that inhibits production of
191P4D12(b). Non-limiting examples of such molecules include, but
are not limited to, those complementary to a nucleotide sequence
essential for production of 191P4D12(b) (e.g. antisense sequences
or molecules that form a triple helix with a nucleotide double
helix essential for 191P4D12(b) production) or a ribozyme effective
to lyse 191P4D12(b) mRNA.
[0032] Note that to determine the starting position of any peptide
set forth in Tables VIII-XXI and XXII to XLIX (collectively HLA
Peptide Tables) respective to its parental protein, e.g., variant
1, variant 2, etc., reference is made to three factors: the
particular variant, the length of the peptide in an HLA Peptide
Table, and the Search Peptides in Table VII. Generally, a unique
Search Peptide is used to obtain HLA peptides of a particular for a
particular variant. The position of each Search Peptide relative to
its respective parent molecule is listed in Table VII. Accordingly,
if a Search Peptide begins at position "X", one must add the value
"X-1" to each position in Tables VIII-XXI and XXII to XLIX to
obtain the actual position of the HLA peptides in their parental
molecule. For example, if a particular Search Peptide begins at
position 150 of its parental molecule, one must add 150-1, i.e.,
149 to each HLA peptide amino acid position to calculate the
position of that amino acid in the parent molecule.
[0033] One embodiment of the invention comprises an HLA peptide,
that occurs at least twice in Tables VIII-XXI and XXII to XLIX
collectively, or an oligonucleotide that encodes the HLA peptide.
Another embodiment of the invention comprises an HLA peptide that
occurs at least once in Tables VIII-XXI and at least once in tables
XXII to XLIX, or an oligonucleotide that encodes the HLA
peptide.
[0034] Another embodiment of the invention is antibody epitopes,
which comprise a peptide regions, or an oligonucleotide encoding
the peptide region, that has one two, three, four, or five of the
following characteristics:
[0035] i) a peptide region of at least 5 amino acids of a
particular peptide of FIG. 3, in any whole number increment up to
the full length of that protein in FIG. 3, that includes an amino
acid position having a value equal to or greater than 0.5, 0.6,
0.7, 0.8, 0.9, or having a value equal to 1.0, in the
Hydrophilicity profile of FIG. 5;
[0036] ii) a peptide region of at least 5 amino acids of a
particular peptide of FIG. 3, in any whole number increment up to
the full length of that protein in FIG. 3, that includes an amino
acid position having a value equal to or less than 0.5, 0.4, 0.3,
0.2, 0.1, or having a value equal to 0.0, in the Hydropathicity
profile of FIG. 6;
[0037] iii) a peptide region of at least 5 amino acids of a
particular peptide of FIG. 3, in any whole number increment up to
the full length of that protein in FIG. 3, that includes an amino
acid position having a value equal to or greater than 0.5, 0.6,
0.7, 0.8, 0.9, or having a value equal to 1.0, in the Percent
Accessible Residues profile of FIG. 7;
[0038] iv) a peptide region of at least 5 amino acids of a
particular peptide of FIG. 3, in any whole number increment up to
the full length of that protein in FIG. 3, that includes an amino
acid position having a value equal to or greater than 0.5, 0.6,
0.7, 0.8, 0.9, or having a value equal to 1.0, in the Average
Flexibility profile of FIG. 8; or
[0039] v) a peptide region of at least 5 amino acids of a
particular peptide of FIG. 3, in any whole number increment up to
the full length of that protein in FIG. 3, that includes an amino
acid position having a value equal to or greater than 0.5, 0.6,
0.7, 0.8, 0.9, or having a value equal to 1.0, in the Beta-turn
profile of FIG. 9.
BRIEF DESCRIPTION OF THE FIGURES
[0040] FIG. 1. The 191P4D12(b) SSH sequence of 223 nucleotides.
[0041] FIG. 2. A) The cDNA and amino acid sequence of 191P4D12(b)
variant 1 (also called "191P4D12(b) v.1" or "191P4D12(b) variant
1") is shown in FIG. 2A. The start methionine is underlined. The
open reading frame extends from nucleic acid 264-1796 including the
stop codon.
[0042] B) The cDNA and amino acid sequence of 191P4D12(b) variant 2
(also called "191P4D12(b) v.2") is shown in FIG. 2B. The codon for
the start methionine is underlined. The open reading frame extends
from nucleic acid 264-1796 including the stop codon.
[0043] C) The cDNA and amino acid sequence of 191P4D12(b) variant 3
(also called "191P4D12(b) v.3") is shown in FIG. 2C. The codon for
the start methionine is underlined. The open reading frame extends
from nucleic acid 264-1796 including the stop codon.
[0044] D) The cDNA and amino acid sequence of 191P4D12(b) variant 4
(also called "191P4D12(b) v.4") is shown in FIG. 2D. The codon for
the start methionine is underlined. The open reading frame extends
from nucleic acid 264-1796 including the stop codon.
[0045] E) The cDNA and amino acid sequence of 191P4D12(b) variant 5
(also called "191P4D12(b) v.5") is shown in FIG. 2E. The codon for
the start methionine is underlined. The open reading frame extends
from nucleic acid 264-1796 including the stop codon.
[0046] F) The cDNA and amino acid sequence of 191P4D12(b) variant 6
(also called "191P4D12(b) v.6") is shown in FIG. 2F. The codon for
the start methionine is underlined. The open reading frame extends
from nucleic acid 789-1676 including the stop codon.
[0047] G) The cDNA and amino acid sequence of 191P4D12(b) variant 7
(also called "191P4D12(b) v.7") is shown in FIG. 2G. The codon for
the start methionine is underlined. The open reading frame extends
from nucleic acid 264-1721 including the stop codon.
[0048] H) The cDNA and amino acid sequence of 191P4D12(b) variant 8
(also called "191P4D12(b) v.8") is shown in FIG. 2H. The codon for
the start methionine is underlined. The open reading frame extends
from nucleic acid 264-1796 including the stop codon.
[0049] I) The cDNA and amino acid sequence of 191P4D12(b) variant 9
(also called "191P4D12(b) v.9") is shown in FIG. 21. The codon for
the start methionine is underlined. The open reading frame extends
from nucleic acid 708-1121 including the stop codon.
[0050] J) The cDNA and amino acid sequence of 191P4D12(b) variant
10 (also called "191P4D12(b) v.10") is shown in FIG. 2J. The codon
for the start methionine is underlined. The open reading frame
extends from nucleic acid 264-1796 including the stop codon.
[0051] K) The cDNA and amino acid sequence of 191P4D12(b) variant
11 (also called "191P4D12(b) v.11") is shown in FIG. 2K. The codon
for the start methionine is underlined. The open reading frame
extends from nucleic acid 264-1796 including the stop codon.
[0052] L) The cDNA and amino acid sequence of 191P4D12(b) variant
12 (also called "191P4D12(b) v.12") is shown in FIG. 2L. The codon
for the start methionine is underlined. The open reading frame
extends from nucleic acid 264-1796 including the stop codon.
[0053] M) The cDNA and amino acid sequence of 191P4D12(b) variant
13 (also called "191P4D12(b) v.13") is shown in FIG. 2M. The codon
for the start methionine is underlined. The open reading frame
extends from nucleic acid 264-1799 including the stop codon.
[0054] N) The cDNA and amino acid sequence of 191P4D12(b) variant
14 (also called "191P4D12(b) v.14") is shown in FIG. 2N. The codon
for the start methionine is underlined. The open reading frame
extends from nucleic acid 708-1121 including the stop codon.
[0055] FIG. 3.
[0056] A) The amino acid sequence of 191P4D12(b) v.1 is shown in
FIG. 3A; it has 510 amino acids.
[0057] B) The amino acid sequence of 191P4D12(b) v.2 is shown in
FIG. 3B; it has 510 amino acids.
[0058] C) The amino acid sequence of 191P4D12(b) v.6 is shown in
FIG. 3C; it has 295 amino acids.
[0059] D) The amino acid sequence of 191P4D12(b) v.7 is shown in
FIG. 3D; it has 485 amino acids.
[0060] E) The amino acid sequence of 191P4D12(b) v.10 is shown in
FIG. 3E; it has 510 amino acids.
[0061] F) The amino acid sequence of 191P4D12(b) v.11 is shown in
FIG. 3F; it has 510 amino acids.
[0062] G) The amino acid sequence of 191P4D12(b) v.12 is shown in
FIG. 3G; it has 510 amino acids.
[0063] H) The amino acid sequence of 191P4D12(b) v.13 is shown in
FIG. 3H; it has 511 amino acids.
[0064] I) The amino acid sequence of 191P4D12(b) v.9 is shown in
FIG. 31; it has 137 amino acids.
[0065] J) The amino acid sequence of 191P4D12(b) v.14 is shown in
FIG. 3J; it has 137 amino acids.
[0066] As used herein, a reference to 191P4D12(b) includes all
variants thereof, including those shown in FIGS. 2, 3, 10, and 11,
unless the context clearly indicates otherwise.
[0067] FIG. 4. Alignment of 191P4D12(b) with known homologs. FIG.
4(A) Alignment of 191P4D12(b) with human Ig superfamily receptor
LNIR (gi 14714574). FIG. 4(B) Alignment of 191P4D12(b) with mouse
nectin 4 (gi 18874521).
[0068] FIG. 5. Hydrophilicity amino acid profile of 191P4D12(b)v.1,
v.7, and v.9 determined by computer algorithm sequence analysis
using the method of Hopp and Woods (Hopp T. P., Woods K. R., 1981.
Proc. Natl. Acad. Sci. U.S.A. 78:3824-3828) accessed on the
Protscale website located on the World Wide Web at
(expasy.ch/cgi-bin/protscale.pl) through the ExPasy molecular
biology server.
[0069] FIG. 6. Hydropathicity amino acid profile of 191P4D12(b)v.1,
v.7, and v.9 determined by computer algorithm sequence analysis
using the method of Kyte and Doolittle (Kyte J., Doolittle R. F.,
1982. J. Mol. Biol. 157:105-132) accessed on the ProtScale website
located on the World Wide Web at (.expasy.ch/cgi-bin/protscale.pl)
through the ExPasy molecular biology server.
[0070] FIG. 7. Percent accessible residues amino acid profile of
191P4D12(b)v.1, v.7, and v.9 determined by computer algorithm
sequence analysis using the method of Janin (Janin J., 1979 Nature
277:491-492) accessed on the ProtScale website located on the World
Wide Web at (.expasy.ch/cgi-bin/protscale.pl) through the ExPasy
molecular biology server.
[0071] FIG. 8. Average flexibility amino acid profile of
191P4D12(b)v.1, v.7, and v.9 determined by computer algorithm
sequence analysis using the method of Bhaskaran and Ponnuswamy
(Bhaskaran R., and Ponnuswamy P. K., 1988. Int. J. Pept. Protein
Res. 32:242-255) accessed on the ProtScale website located on the
World Wide Web at (.expasy.ch/cgi-bin/protscale.pl- ) through the
ExPasy molecular biology server.
[0072] FIG. 9. Beta-turn amino acid profile of 191P4D12(b)v.1, v.7,
and v.9 determined by computer algorithm sequence analysis using
the method of Deleage and Roux (Deleage, G., Roux B. 1987 Protein
Engineering 1:289-294) accessed on the ProtScale website located on
the World Wide Web at (.expasy.ch/cgi-bin/protscale.pl) through the
ExPasy molecular biology server.
[0073] FIG. 10. Schematic alignment of SNP variants of 191P4D12(b).
Variants 191P4D12(b) v.2 through v.5 and v.10 through v.12 are
variants with single nucleotide differences. Compared with v.1,
v.13 had an insertion of three bases (GCA) between 1262 and 1263
and added one amino acid "A" to the protein. Variant v.14 was a SNP
variant of transcript variant v.9, corresponding to the SNP at 2688
of v.1. Though these SNP variants were shown separately, they could
also occur in any combinations and in any transcript variants, as
shown in FIG. 12, that contained the base pairs. Numbers correspond
to those of 191P4D12(b) v.1. Black box shows the same sequence as
191P4D12(b) v.1. SNPs are indicated above the box.
[0074] FIG. 11. Schematic alignment of protein variants of
191P4D12(b). Protein variants correspond to nucleotide variants.
Nucleotide variants 191P4D12(b) v.3, v.4, v.5 and v.8 coded for the
same protein as v.1. Nucleotide variants 191P4D12(b) v.6, v.7, v.8
and v.9 were splice variants of v.1, as shown in FIG. 12. Variant
v.9 translated to a totally different protein than other variants,
with two isoforms that different from each other by one amino acid
at 64: A or D. Variant v.13 had an insertion of one amino acid "A"
at 334. Single amino acid differences were indicated above the
boxes. Black boxes represent the same sequence as 191P4D12(b) v.1.
Numbers underneath the box correspond to 191P4D12(b) v.1.
[0075] FIG. 12. Exon compositions of transcript variants of
191P4D12(b). Variant 191P4D12(b) v.6, v.7, v.8 and v.9 are
transcript variants of v.1. Variants v.6, v.7 and v.8 spliced out
202-321, 1497-1571 and 2951-3013 of v.1, respectively. Variant v.9
was part of the last exon of v.1. The order of the potential exons
on the human genome is shown at the bottom. Poly A tails were not
shown in the figure. Ends of exons are shown above the boxes.
Numbers in "()" underneath the boxes correspond to those of
191P4D12(b) v.1. Lengths of introns and exons are not
proportional.
[0076] FIG. 13. Secondary structure and transmembrane domains
prediction for 191P4D12(b) protein variants. The secondary
structure of 191P4D12(b) protein variants 1 (SEQ ID NO: 127), v6
(SEQ ID NO: 128), v7 (SEQ ID NO: 129), and v9 (SEQ ID NO: 130)
(FIGS. 13A-D respectively) were predicted using the
HNN--Hierarchical Neural Network method (Guermeur, 1997,
http://pbil.ibcp.fr/cgi-bin/npsa_automat.pl?page=npsa_nn.html),
accessed from the ExPasy molecular biology server located on the
World Wide Web at (.expasy.ch/tools/). This method predicts the
presence and location of alpha helices, extended strands, and
random coils from the primary protein sequence.
[0077] The percent of the protein in a given secondary structure is
also listed. FIGS. 13E, 13G, 13I, 13K: Schematic representations of
the probability of existence of transmembrane regions and
orientation of 191P4D12(b) variants 1, 6, 7, and 9, respectively,
based on the TMpred algorithm of Hofmann and Stoffel which utilizes
TMBASE (K. Hofmann, W. Stoffel. TMBASE--A database of membrane
spanning protein segments Biol. Chem. Hoppe-Seyler 374:166, 1993).
FIGS. 13F, 13H, 13J, 13L. Schematic representations of the
probability of the existence of transmembrane regions and the
extracellular and intracellular orientation of 191P4D12(b) variants
1, 6, 7, and 9, respectively, based on the TMHMM algorithm of
Sonnhammer, von Heijne, and Krogh (Erik L. L. Sonnhammer, Gunnar
von Heijne, and Anders Krogh: A hidden Markov model for predicting
transmembrane helices in protein sequences. In Proc. of Sixth Int.
Conf. on Intelligent Systems for Molecular Biology, p 175-182 Ed J.
Glasgow, T. Littlejohn, F. Major, R. Lathrop, D. Sankoff, and C.
Sensen Menlo Park, Calif.: AAAI Press, 1998). The TMpred and TMHMM
algorithms are accessed from the ExPasy molecular biology server
located on the World Wide Web at (.expasy.ch/tools/).
[0078] FIG. 14. 191P4D12(b) Expression by RT-PCR. First strand cDNA
was prepared from (A) vital pool 1 (liver, lung and kidney), vital
pool 2 (pancreas, colon and stomach), normal kidney, prostate
cancer pool, bladder cancer pool, colon cancer pool, lung cancer
pool, breast cancer pool and cancer metastasis pool; (B) prostate
cancer metastasis to lymph node, prostate cancer pool, bladder
cancer pool, kidney cancer pool, colon cancer pool, lung cancer
pool, ovary cancer pool, breast cancer pool, cancer metastasis
pool, pancreas cancer pool, and LAPC prostate xenograft pool.
Normalization was performed by PCR using primers to actin and
GAPDH. Semi-quantitative PCR, using primers to 191P4D12(b), was
performed at 26 and 30 cycles of amplification. In (A) results show
strong expression of 191P4D12(b) in bladder cancer pool. Expression
of 191P4D12(b) was also detected in prostate cancer pool, colon
cancer pool, lung cancer pool, breast cancer pool and cancer
metastasis pool but very weakly in vital pool 1 and vital pool 2.
In (B) results show strong expression of 191P4D12(b) in prostate,
bladder, kidney, colon, lung, ovary, breast, cancer metastasis, and
pancreas cancer specimens.
[0079] FIG. 15. Expression of 191P4D12(b) in normal tissues. Two
multiple tissue northern blots (Clontech) both with 2 ug of
mRNA/lane were probed with the 191P4D12(b) sequence. Size standards
in kilobases (kb) are indicated on the side. Results show
expression of an approximately 4 kb transcript in placenta and very
weakly in prostate but not in any other normal tissue tested. A
smaller 191P4D12(b) transcript of approximately 2.5 kb was detected
in heart and skeletal muscle.
[0080] FIG. 16. Expression of 191P4D12(b) in Patient Cancer
Specimens and Normal Tissues. RNA was extracted from a pool of 3
bladder cancer patient specimens, as well as from normal prostate
(NP), normal bladder (NB), normal kidney (NK), normal colon (NC),
normal lung (NL), normal breast (NBr), normal ovary (NO), and
normal pancreas (NPa). Northern blot with 10 ug of total RNA/lane
was probed with 191P4D12(b) SSH sequence. Size standards in
kilobases (kb) are indicated on the side. The 191P4D12(b)
transcript was detected in the bladder cancer specimens, but not in
the normal tissues tested.
[0081] FIG. 17. Expression of 191P4D12(b) in Bladder Cancer Patient
Specimens. RNA was extracted from bladder cancer cell lines (CL),
normal bladder (N), and bladder cancer patient tumors (T). Northern
blots with 10 ug of total RNA were probed with the 191P4D12(b) SSH
fragment. Size standards in kilobases are on the side. Results show
expression of the approximately 4 kb 191P4D12(b) transcript in the
bladder tumor tissues but not in normal bladder. A smaller
transcript was detected in the HT1197 cell line but not in the
other cancer cell lines tested.
[0082] FIG. 18. Expression of 191P4D12(b) in Prostate Cancer
Xenografts. RNA was extracted from normal prostate, and from the
prostate cancer xenografts LAPC-4AD, LAPC-4AI, LAPC-9AD, and
LAPC-9AI. Northern blots with 10 ug of total RNA were probed with
the 191P4D12(b) SSH fragment. Size standards in kilobases are on
the side. Results show expression of the approximately 4 kb
191P4D12(b) transcript in all the LAPC xenograft tissues but not in
normal prostate.
[0083] FIG. 19. Expression of 191P4D12(b) in Cervical Cancer
Patient Specimens. RNA was extracted from normal cervix, Hela
cancer cell line, and 3 cervix cancer patient tumors (T). Northern
blots with 10 ug of total RNA were probed with the 191P4D12(b) SSH
fragment. Size standards in kilobases are on the side. Results show
expression of the approximately 4 kb 191P4D12(b) transcript in 2
out of 3 cervix tumors but not in normal cervix nor in the Hela
cell line.
[0084] FIG. 20. Expression of 191P4D12(b) in Lung Cancer Patient
Specimens. RNA was extracted from lung cancer cell lines (CL),
normal lung (N), bladder cancer patient tumors (T), and normal
adjacent tissue (Nat). Northern blots with 10 ug of total RNA were
probed with the 191P4D12(b). Size standards in kilobases are on the
side. Results show expression of the approximately 4 kb 191P4D12(b)
transcript in the lung tumor tissues but not in normal lung nor in
the cell lines tested.
[0085] FIG. 21. FIG. 21A. 191P4D12(b) Expression in Lung Cancer.
First strand cDNA was prepared from a panel of lung cancer
specimens. Normalization was performed by PCR using primers to
actin. Semi-quantitative PCR, using primers to 191P4D12(b) SSH
fragment, was performed at 26 and 30 cycles of amplification.
Expression level was recorded as 0=no expression detected; 1=weak
expression, 2=moderate expression; 3=strong expression. Results
show expression of 191P4D12(b) in 97% of the 31 lung cancer patient
specimens tested. FIG. 21B. 191P4D12(b) Expression in Bladder
Cancer. First strand cDNA was prepared from a panel of bladder
cancer specimens. Normalization was performed by PCR using primers
to actin. Semi-quantitative PCR, using primers to 191P4D12(b) SSH
fragment, was performed at 26 and 30 cycles of amplification.
Expression level was recorded as 0=no expression detected, 1=weak
expression, 2=moderate expression; 3=strong expression. Results
show expression of 191P4D12(b) in 94% of the 18 bladder cancer
patient specimens tested. FIG. 21C. 191P4D12(b) Expression in
Prostate Cancer. First strand cDNA was prepared from a panel of
prostate cancer specimens, and four LAPC prostate cancer
xenografts. Normalization was performed by PCR using primers to
actin. Semi-quantitative PCR, using primers to 191P4D12(b) SSH
fragment, was performed at 26 and 30 cycles of amplification.
Expression level was recorded as 0=no expression detected; 1=weak
expression, 2=moderate expression, 3=strong expression. Results
show expression of 191P4D12(b) in 100% of the 20 prostate cancer
patient specimens tested, and in all 4 prostate cancer xenografts.
FIG. 21D. 191P4D12(b) Expression in Colon Cancer. First strand cDNA
was prepared from a panel of colon cancer specimens. Normalization
was performed by PCR using primers to actin. Semi-quantitative PCR,
using primers to 191P4D12(b) SSH fragment, was performed at 26 and
30 cycles of amplification. Expression level was recorded as 0=no
expression detected; 1=weak expression, 2=moderate expression;
3=strong expression. Results show expression of 191P4D12(b) in 100%
of the 22 colon cancer patient specimens tested. FIG. 21E.
191P4D12(b) Expression in Uterus Cancer. First strand cDNA was
prepared from a panel of uterus cancer specimens. Normalization was
performed by PCR using primers to actin. Semi-quantitative PCR,
using primers to 191P4D12(b) SSH fragment, was performed at 26 and
30 cycles of amplification. Expression level was recorded as 0=no
expression detected; 1=weak expression, 2=moderate expression;
3=strong expression. Results show expression of 191P4D12(b) in 100%
of the 12 uterus cancer patient specimens tested. FIG. 21F.
191P4D12(b) Expression in Cervical Cancer. First strand cDNA was
prepared from a panel of cervix cancer specimens. Normalization was
performed by PCR using primers to actin. Semi-quantitative PCR,
using primers to 191P4D12(b) SSH fragment, was performed at 26 and
30 cycles of amplification. Expression level was recorded as 0=no
expression detected; 1=weak expression, 2=moderate expression;
3=strong expression. Results show expression of 191P4D12(b) in 100%
of the 14 cervix cancer patient specimens tested.
[0086] FIG. 22. Transient Expression of 191P4D12(b) in Transfected
293T Cells. 293T cells were transfected with either
191P4D12(b).pTag5, 191P4D12(b).pcDNA3.1/mychis or pcDNA3.1/mychis
vector control. Forty hours later, cell lysates and supernatant
were collected. Samples were run on an SDS-PAGE acrylamide gel,
blotted and stained with anti-his antibody. The blot was developed
using the ECL chemiluminescence kit and visualized by
autoradiography. Results show expression from 191P4D12(b).pTag5
plasmid of 191P4D12(b) extracellular domain in the lysate (Lane 2)
and secretion in the culture supernatant (Lane 1). Also, expression
of 191P4D12(b) was detected from in the lysates of
191P4D12(b).pcDNA3.1/mychis transfected cells (Lane 3), but not
from the control pcDNA3.1/mychis (Lane 4).
[0087] FIG. 23. Expression of 191P4D12(b) in Transduced Cells
Following Retroviral Gene Transfer. 3T3 cells were transduced with
the pSRa retroviral vector encoding the 191P4D12(b) gene. Following
selection with neomycin, the cells were expanded and RNA was
extracted. Northern blot with 10 ug of total RNA/lane was probed
with the 191P4D12(b) SSH sequence. Size standards in kilobases (kb)
are indicated on the side. Results show expression of the
191P4D12(b) transcript driven from the retroviral LTR, which
migrates slower than the endogenous 4 kb 191P4D12(b) transcript
detected in the positive control LAPC-4AD.
DETAILED DESCRIPTION OF THE INVENTION
Outline of Sections
[0088] I.) Definitions
[0089] II.) 191P4D12(b) Polynucleotides
[0090] II.A.) Uses of 191P4D12(b) Polynucleotides
[0091] II.A.1.) Monitoring of Genetic Abnormalities
[0092] II.A.2.) Antisense Embodiments
[0093] II.A.3.) Primers and Primer Pairs
[0094] II.A.4.) Isolation of 191P4D12(b)-Encoding Nucleic Acid
Molecules
[0095] II.A.5.) Recombinant Nucleic Acid Molecules and Host-Vector
Systems
[0096] III.) 191P4D12(b)-related Proteins
[0097] III.A.) Motif-bearing Protein Embodiments
[0098] III.B.) Expression of 191P4D12(b)-related Proteins
[0099] III.C.) Modifications of 191P4D12(b)-related Proteins
[0100] III.D.) Uses of 191P4D12(b)-related Proteins
[0101] IV.) 191P4D12(b) Antibodies
[0102] V.) 191P4D12(b) Cellular Immune Responses
[0103] VI.) 191P4D12(b) Transgenic Animals
[0104] VIl.) Methods for the Detection of 191P4D12(b)
[0105] VIII.) Methods for Monitoring the Status of
191P4D12(b)-related Genes and Their Products
[0106] IX.) Identification of Molecules That Interact With
191P4D12(b)
[0107] X.) Therapeutic Methods and Compositions
[0108] X.A.) Anti-Cancer Vaccines
[0109] X.B.) 191P4D12(b) as a Target for Antibody-Based Therapy
[0110] X.C.) 191P4D12(b) as a Target for Cellular Immune
Responses
[0111] X.C.1. Minigene Vaccines
[0112] X.C.2. Combinations of CTL Peptides with Helper Peptides
[0113] X.C.3. Combinations of CTL Peptides with T Cell Priming
Agents
[0114] X.C.4. Vaccine Compositions Comprising DC Pulsed with CTL
and/or HTL Peptides
[0115] X.D.) Adoptive Immunotherapy
[0116] X.E.) Administration of Vaccines for Therapeutic or
Prophylactic Purposes
[0117] XI.) Diagnostic and Prognostic Embodiments of
191P4D12(b).
[0118] XII.) Inhibition of 191P4D12(b) Protein Function
[0119] XII.A.) Inhibition of 191P4D12(b) With Intracellular
Antibodies
[0120] XII.B.) Inhibition of 191P4D12(b) with Recombinant
Proteins
[0121] XII.C.) Inhibition of 191P4D12(b) Transcription or
Translation
[0122] XII.D.) General Considerations for Therapeutic
Strategies
[0123] XIII.) Identification, Characterization and Use of
Modulators of 191P4D12(b)
[0124] XIV.) KITS/Articles of Manufacture
[0125] I.) Definitions:
[0126] Unless otherwise defined, all terms of art, notations and
other scientific terms or terminology used herein are intended to
have the meanings commonly understood by those of skill in the art
to which this invention pertains. In some cases, terms with
commonly understood meanings are defined herein for clarity and/or
for ready reference, and the inclusion of such definitions herein
should not necessarily be construed to represent a substantial
difference over what is generally understood in the art. Many of
the techniques and procedures described or referenced herein are
well understood and commonly employed using conventional
methodology by those skilled in the art, such as, for example, the
widely utilized molecular cloning methodologies described in
Sambrook et al., Molecular Cloning: A Laboratory Manual 2nd.
edition (1989) Cold Spring Harbor Laboratory Press, Cold Spring
Harbor, N.Y. As appropriate, procedures involving the use of
commercially available kits and reagents are generally carried out
in accordance with manufacturer defined protocols and/or parameters
unless otherwise noted.
[0127] The terms "advanced prostate cancer", "locally advanced
prostate cancer", "advanced disease" and "locally advanced disease"
mean prostate cancers that have extended through the prostate
capsule, and are meant to include stage C disease under the
American Urological Association (AUA) system, stage C1-C2 disease
under the Whitmore-Jewett system, and stage T3-T4 and N+ disease
under the TNM (tumor, node, metastasis) system. In general, surgery
is not recommended for patients with locally advanced disease, and
these patients have substantially less favorable outcomes compared
to patients having clinically localized (organ-confined) prostate
cancer. Locally advanced disease is clinically identified by
palpable evidence of induration beyond the lateral border of the
prostate, or asymmetry or induration above the prostate base.
Locally advanced prostate cancer is presently diagnosed
pathologically following radical prostatectomy if the tumor invades
or penetrates the prostatic capsule, extends into the surgical
margin, or invades the seminal vesicles.
[0128] "Altering the native glycosylation pattern" is intended for
purposes herein to mean deleting one or more carbohydrate moieties
found in native sequence 191P4D12(b) (either by removing the
underlying glycosylation site or by deleting the glycosylation by
chemical and/or enzymatic means), and/or adding one or more
glycosylation sites that are not present in the native sequence
191P4D12(b). In addition, the phrase includes qualitative changes
in the glycosylation of the native proteins, involving a change in
the nature and proportions of the various carbohydrate moieties
present.
[0129] The term "analog" refers to a molecule which is structurally
similar or shares similar or corresponding attributes with another
molecule (e.g. a 191P4D12(b)-related protein). For example, an
analog of a 191P4D12(b) protein can be specifically bound by an
antibody or T cell that specifically binds to 191P4D12(b).
[0130] The term "antibody" is used in the broadest sense.
Therefore, an "antibody" can be naturally occurring or man-made
such as monoclonal antibodies produced by conventional hybridoma
technology. Anti-191P4D12(b) antibodies comprise monoclonal and
polyclonal antibodies as well as fragments containing the
antigen-binding domain and/or one or more complementarity
determining regions of these antibodies.
[0131] An "antibody fragment" is defined as at least a portion of
the variable region of the immunoglobulin molecule that binds to
its target, i.e., the antigen-binding region. In one embodiment it
specifically covers single anti-191P4D12(b) antibodies and clones
thereof (including agonist, antagonist and neutralizing antibodies)
and anti-191P4D12(b) antibody compositions with polyepitopic
specificity.
[0132] The term "codon optimized sequences" refers to nucleotide
sequences that have been optimized for a particular host species by
replacing any codons having a usage frequency of less than about
20%. Nucleotide sequences that have been optimized for expression
in a given host species by elimination of spurious polyadenylation
sequences, elimination of exon/intron splicing signals, elimination
of transposon-like repeats and/or optimization of GC content in
addition to codon optimization are referred to herein as an
"expression enhanced sequences."
[0133] A "combinatorial library" is a collection of diverse
chemical compounds generated by either chemical synthesis or
biological synthesis by combining a number of chemical "building
blocks" such as reagents. For example, a linear combinatorial
chemical library, such as a polypeptide (e.g., mutein) library, is
formed by combining a set of chemical building blocks called amino
acids in every possible way for a given compound length (i.e., the
number of amino acids in a polypeptide compound). Numerous chemical
compounds are synthesized through such combinatorial mixing of
chemical building blocks (Gallop et al., J. Med. Chem. 37(9):
1233-1251 (1994)).
[0134] Preparation and screening of combinatorial libraries is well
known to those of skill in the art. Such combinatorial chemical
libraries include, but are not limited to, peptide libraries (see,
e.g., U.S. Pat. No. 5,010,175, Furka, Pept. Prot. Res. 37:487-493
(1991), Houghton et al., Nature, 354:84-88 (1991)), peptoids (PCT
Publication No WO 91/19735), encoded peptides (PCT Publication WO
93/20242), random bio-oligomers (PCT Publication WO 92/00091),
benzodiazepines (U.S. Pat. No. 5,288,514), diversomers such as
hydantoins, benzodiazepines and dipeptides (Hobbs et al., Proc.
Nat. Acad. Sci. USA 90:6909-6913 (1993)), vinylogous polypeptides
(Hagihara et al., J. Amer. Chem. Soc. 114:6568 (1992)), nonpeptidal
peptidomimetics with a Beta-D-Glucose scaffolding (Hirschmann et
al., J. Amer. Chem. Soc. 114:9217-9218 (1992)), analogous organic
syntheses of small compound libraries (Chen et al., J. Amer. Chem.
Soc. 116:2661 (1994)), oligocarbarnates (Cho, et al., Science
261:1303 (1993)), and/or peptidyl phosphonates (Campbell et al., J.
Org. Chem. 59:658 (1994)). See, generally, Gordon et al., J. Med.
Chem. 37:1385 (1994), nucleic acid libraries (see, e.g.,
Stratagene, Corp.), peptide nucleic acid libraries (see, e.g., U.S.
Pat. No. 5,539,083), antibody libraries (see, e.g., Vaughn et al.,
Nature Biotechnology 14(3): 309-314 (1996), and PCT/US96/10287),
carbohydrate libraries (see, e.g., Liang et al., Science
274:1520-1522 (1996), and U.S. Pat. No. 5,593,853), and small
organic molecule libraries (see, e.g., benzodiazepines, Baum,
C&EN, Jan 18, page 33 (1993); isoprenoids, U.S. Pat. No.
5,569,588; thiazolidinones and metathiazanones, U.S. Pat. No.
5,549,974; pyrrolidines, U.S. Pat. Nos. 5,525,735 and 5,519,134;
morpholino compounds, U.S. Pat. No. 5,506,337; benzodiazepines,
U.S. Pat. No. 5,288,514; and the like).
[0135] Devices for the preparation of combinatorial libraries are
commercially available (see, e.g., 357 NIPS, 390 NIPS, Advanced
Chem Tech, Louisville Ky.; Symphony, Rainin, Woburn, Mass.; 433A,
Applied Biosystems, Foster City, Calif.; 9050, Plus, Millipore,
Bedford, NIA). A number of well-known robotic systems have also
been developed for solution phase chemistries. These systems
include automated workstations such as the automated synthesis
apparatus developed by Takeda Chemical Industries, LTD. (Osaka,
Japan) and many robotic systems utilizing robotic arms (Zymate H,
Zymark Corporation, Hopkinton, Mass.; Orca, Hewlett-Packard, Palo
Alto, Calif.), which mimic the manual synthetic operations
performed by a chemist. Any of the above devices are suitable for
use with the present invention. The nature and implementation of
modifications to these devices (if any) so that they can operate as
discussed herein will be apparent to persons skilled in the
relevant art. In addition, numerous combinatorial libraries are
themselves commercially available (see, e.g., ComGenex, Princeton,
N.J.; Asinex, Moscow, RU; Tripos, Inc., St. Louis, Mo.; ChemStar,
Ltd, Moscow, RU; 3D Pharmaceuticals, Exton, Pa.; Martek
Biosciences, Columbia, Md.; etc.).
[0136] The term "cytotoxic agent" refers to a substance that
inhibits or prevents the expression activity of cells, function of
cells and/or causes destruction of cells. The term is intended to
include radioactive isotopes chemotherapeutic agents, and toxins
such as small molecule toxins or enzymatically active toxins of
bacterial, fungal, plant or animal origin, including fragments
and/or variants thereof. Examples of cytotoxic agents include, but
are not limited to auristatins, auromycins, maytansinoids, yttrium,
bismuth, ricin, ricin A-chain, combrestatin, duocarmycins,
dolostatins, doxorubicin, daunorubicin, taxol, cisplatin, cc1065,
ethidium bromide, mitomycin, etoposide, tenoposide, vincristine,
vinblastine, colchicine, dihydroxy anthracin dione, actinomycin,
diphtheria toxin, Pseudomonas exotoxin (PE) A, PE40, abrin, abrin A
chain, modeccin A chain, alpha-sarcin, gelonin, mitogellin,
retstrictocin, phenomycin, enomycin, curicin, crotin,
calicheamicin, Sapaonaria officinalis inhibitor, and glucocorticoid
and other chemotherapeutic agents, as well as radioisotopes such as
At.sup.211, I.sup.131, I.sup.125,Y.sup.90, Re.sup.186, Re.sup.188,
Sm.sup.153, Bi212 or 213, P.sup.32 and radioactive isotopes of Lu
including Lu.sup.177. Antibodies may also be conjugated to an
anti-cancer pro-drug activating enzyme capable of converting the
pro-drug to its active form.
[0137] The "gene product" is sometimes referred to herein as a
protein or mRNA. For example, a "gene product of the invention" is
sometimes referred to herein as a "cancer amino acid sequence",
"cancer protein", "protein of a cancer listed in Table I", a
"cancer mRNA", "mRNA of a cancer listed in Table I", etc. In one
embodiment, the cancer protein is encoded by a nucleic acid of FIG.
2. The cancer protein can be a fragment, or alternatively, be the
full-length protein to the fragment encoded by the nucleic acids of
FIG. 2. In one embodiment, a cancer amino acid sequence is used to
determine sequence identity or similarity. In another embodiment,
the sequences are naturally occurring allelic variants of a protein
encoded by a nucleic acid of FIG. 2. In another embodiment, the
sequences are sequence variants as further described herein.
[0138] "High throughput screening" assays for the presence,
absence, quantification, or other properties of particular nucleic
acids or protein products are well known to those of skill in the
art. Similarly, binding assays and reporter gene assays are
similarly well known. Thus, e.g., U.S. Pat. No. 5,559,410 discloses
high throughput screening methods for proteins; U.S. Pat. No.
5,585,639 discloses high throughput screening methods for nucleic
acid binding (i.e., in arrays); while U.S. Pat. Nos. 5,576,220 and
5,541,061 disclose high throughput methods of screening for
ligand/antibody binding.
[0139] In addition, high throughput screening systems are
commercially available (see, e.g., Amersham Biosciences,
Piscataway, N.J.; Zymark Corp., Hopkinton, Mass.; Air Technical
Industries, Mentor, Ohio; Beckman Instruments, Inc. Fullerton,
Calif.; Precision Systems, Inc., Natick, Mass.; etc.). These
systems typically automate entire procedures, including all sample
and reagent pipetting, liquid dispensing, timed incubations, and
final readings of the microplate in detector(s) appropriate for the
assay. These configurable systems provide high throughput and rapid
start up as well as a high degree of flexibility and customization.
The manufacturers of such systems provide detailed protocols for
various high throughput systems. Thus, e.g., Zymark Corp. provides
technical bulletins describing screening systems for detecting the
modulation of gene transcription, ligand binding, and the like.
[0140] The term "homolog" refers to a molecule which exhibits
homology to another molecule, by for example, having sequences of
chemical residues that are the same or similar at corresponding
positions.
[0141] "Human Leukocyte Antigen" or "HLA" is a human class I or
class II Major Histocompatibility Complex (MHC) protein (see, e.g.,
Stites, et al., IMMUNOLOGY, 8.sup.TH ED., Lange Publishing, Los
Altos, Calif. (1994).
[0142] The terms "hybridize", "hybridizing", "hybridizes" and the
like, used in the context of polynucleotides, are meant to refer to
conventional hybridization conditions, preferably such as
hybridization in 50% formamide/6.times.SSC/0.1% SDS/100 .mu.g/ml
ssDNA, in which temperatures for hybridization are above 37 degrees
C. and temperatures for washing in 0.1.times.SSC/0.1% SDS are above
55 degrees C.
[0143] The phrases "isolated" or "biologically pure" refer to
material which is substantially or essentially free from components
which normally accompany the material as it is found in its native
state. Thus, isolated peptides in accordance with the invention
preferably do not contain materials normally associated with the
peptides in their in situ environment. For example, a
polynucleotide is said to be "isolated" when it is substantially
separated from contaminant polynucleotides that correspond or are
complementary to genes other than the 191P4D12(b) genes or that
encode polypeptides other than 191P4D12(b) gene product or
fragments thereof. A skilled artisan can readily employ nucleic
acid isolation procedures to obtain an isolated 191P4D12(b)
polynucleotide. A protein is said to be "isolated," for example,
when physical, mechanical or chemical methods are employed to
remove the 191P4D12(b) proteins from cellular constituents that are
normally associated with the protein. A skilled artisan can readily
employ standard purification methods to obtain an isolated
191P4D12(b) protein. Alternatively, an isolated protein can be
prepared by chemical means.
[0144] The term "mammal" refers to any organism classified as a
mammal, including mice, rats, rabbits, dogs, cats, cows, horses and
humans. In one embodiment of the invention, the mammal is a mouse.
In another embodiment of the invention, the mammal is a human.
[0145] The terms "metastatic prostate cancer" and "metastatic
disease" mean prostate cancers that have spread to regional lymph
nodes or to distant sites, and are meant to include stage D disease
under the AUA system and stage T.times.N.times.M+ under the TNM
system. As is the case with locally advanced prostate cancer,
surgery is generally not indicated for patients with metastatic
disease, and hormonal (androgen ablation) therapy is a preferred
treatment modality. Patients with metastatic prostate cancer
eventually develop an androgen-refractory state within 12 to 18
months of treatment initiation. Approximately half of these
androgen-refractory patients die within 6 months after developing
that status. The most common site for prostate cancer metastasis is
bone. Prostate cancer bone metastases are often osteoblastic rather
than osteolytic (i.e., resulting in net bone formation). Bone
metastases are found most frequently in the spine, followed by the
femur, pelvis, rib cage, skull and humerus. Other common sites for
metastasis include lymph nodes, lung, liver and brain. Metastatic
prostate cancer is typically diagnosed by open or laparoscopic
pelvic lymphadenectomy, whole body radionuclide scans, skeletal
radiography, and/or bone lesion biopsy.
[0146] The term "modulator" or "test compound" or "drug candidate"
or grammatical equivalents as used herein describe any molecule,
e.g., protein, oligopeptide, small organic molecule,
polysaccharide, polynucleotide, etc., to be tested for the capacity
to directly or indirectly alter the cancer phenotype or the
expression of a cancer sequence, e.g., a nucleic acid or protein
sequences, or effects of cancer sequences (e.g., signaling, gene
expression, protein interaction, etc.) In one aspect, a modulator
will neutralize the effect of a cancer protein of the invention. By
"neutralize" is meant that an activity of a protein is inhibited or
blocked, along with the consequent effect on the cell. In another
aspect, a modulator will neutralize the effect of a gene, and its
corresponding protein, of the invention by normalizing levels of
said protein. In preferred embodiments, modulators alter expression
profiles, or expression profile nucleic acids or proteins provided
herein, or downstream effector pathways. In one embodiment, the
modulator suppresses a cancer phenotype, e.g. to a normal tissue
fingerprint. In another embodiment, a modulator induced a cancer
phenotype. Generally, a plurality of assay mixtures is run in
parallel with different agent concentrations to obtain a
differential response to the various concentrations. Typically, one
of these concentrations serves as a negative control, i.e., at zero
concentration or below the level of detection.
[0147] Modulators, drug candidates or test compounds encompass
numerous chemical classes, though typically they are organic
molecules, preferably small organic compounds having a molecular
weight of more than 100 and less than about 2,500 Daltons.
Preferred small molecules are less than 2000, or less than 1500 or
less than 1000 or less than 500 D. Candidate agents comprise
functional groups necessary for structural interaction with
proteins, particularly hydrogen bonding, and typically include at
least an amine, carbonyl, hydroxyl or carboxyl group, preferably at
least two of the functional chemical groups. The candidate agents
often comprise cyclical carbon or heterocyclic structures and/or
aromatic or polyaromatic structures substituted with one or more of
the above functional groups. Modulators also comprise biomolecules
such as peptides, saccharides, fatty acids, steroids, purines,
pyrimidines, derivatives, structural analogs or combinations
thereof. Particularly preferred are peptides. One class of
modulators are peptides, for example of from about five to about 35
amino acids, with from about five to about 20 amino acids being
preferred, and from about 7 to about 15 being particularly
preferred. Preferably, the cancer modulatory protein is soluble,
includes a non-transmembrane region, and/or, has an N-terminal Cys
to aid in solubility. In one embodiment, the C-terminus of the
fragment is kept as a free acid and the N-terminus is a free amine
to aid in coupling, i.e., to cysteine. In one embodiment, a cancer
protein of the invention is conjugated to an immunogenic agent as
discussed herein. In one embodiment, the cancer protein is
conjugated to BSA. The peptides of the invention, e.g., of
preferred lengths, can be linked to each other or to other amino
acids to create a longer peptide/protein. The modulatory peptides
can be digests of naturally occurring proteins as is outlined
above, random peptides, or "biased" random peptides. In a preferred
embodiment, peptide/protein-based modulators are antibodies, and
fragments thereof, as defined herein.
[0148] Modulators of cancer can also be nucleic acids. Nucleic acid
modulating agents can be naturally occurring nucleic acids, random
nucleic acids, or "biased" random nucleic acids. For example,
digests of prokaryotic or eukaryotic genomes can be used in an
approach analogous to that outlined above for proteins.
[0149] The term "monoclonal antibody" refers to an antibody
obtained from a population of substantially homogeneous antibodies,
i.e., the antibodies comprising the population are identical except
for possible naturally occurring mutations that are present in
minor amounts.
[0150] A "motif", as in biological motif of a 191P4D12(b)-related
protein, refers to any pattern of amino acids forming part of the
primary sequence of a protein, that is associated with a particular
function (e.g. protein-protein interaction, protein-DNA
interaction, etc) or modification (e.g. that is phosphorylated,
glycosylated or amidated), or localization (e.g. secretory
sequence, nuclear localization sequence, etc.) or a sequence that
is correlated with being immunogenic, either humorally or
cellularly. A motif can be either contiguous or capable of being
aligned to certain positions that are generally correlated with a
certain function or property. In the context of HLA motifs, "motif"
refers to the pattern of residues in a peptide of defined length,
usually a peptide of from about 8 to about 13 amino acids for a
class I HLA motif and from about 6 to about 25 amino acids for a
class II HLA motif, which is recognized by a particular HLA
molecule. Peptide motifs for HLA binding are typically different
for each protein encoded by each human HLA allele and differ in the
pattern of the primary and secondary anchor residues.
[0151] A "pharmaceutical excipient" comprises a material such as an
adjuvant, a carrier, pH-adjusting and buffering agents, tonicity
adjusting agents, wetting agents, preservative, and the like.
[0152] "Pharmaceutically acceptable" refers to a non-toxic, inert,
and/or composition that is physiologically compatible with humans
or other mammals.
[0153] The term "polynucleotide" means a polymeric form of
nucleotides of at least 10 bases or base pairs in length, either
ribonucleotides or deoxynucleotides or a modified form of either
type of nucleotide, and is meant to include single and double
stranded forms of DNA and/or RNA. In the art, this term if often
used interchangeably with "oligonucleotide". A polynucleotide can
comprise a nucleotide sequence disclosed herein wherein thymidine
(T), as shown for example in FIG. 2, can also be uracil (U); this
definition pertains to the differences between the chemical
structures of DNA and RNA, in particular the observation that one
of the four major bases in RNA is uracil (U) instead of thymidine
(T).
[0154] The term "polypeptide" means a polymer of at least about 4,
5, 6, 7, or 8 amino acids. Throughout the specification, standard
three letter or single letter designations for amino acids are
used. In the art, this term is often used interchangeably with
"peptide" or "protein".
[0155] An HLA "primary anchor residue" is an amino acid at a
specific position along a peptide sequence which is understood to
provide a contact point between the immunogenic peptide and the HLA
molecule. One to three, usually two, primary anchor residues within
a peptide of defined length generally defines a "motif" for an
immunogenic peptide. These residues are understood to fit in close
contact with peptide binding groove of an HLA molecule, with their
side chains buried in specific pockets of the binding groove. In
one embodiment, for example, the primary anchor residues for an HLA
class I molecule are located at position 2 (from the amino terminal
position) and at the carboxyl terminal position of a 8, 9, 10, 11,
or 12 residue peptide epitope in accordance with the invention.
Alternatively, in another embodiment, the primary anchor residues
of a peptide binds an HLA class II molecule are spaced relative to
each other, rather than to the termini of a peptide, where the
peptide is generally of at least 9 amino acids in length. The
primary anchor positions for each motif and supermotif are set
forth in Table IV. For example, analog peptides can be created by
altering the presence or absence of particular residues in the
primary and/or secondary anchor positions shown in Table IV. Such
analogs are used to modulate the binding affinity and/or population
coverage of a peptide comprising a particular HLA motif or
supermotif.
[0156] "Radioisotopes" include, but are not limited to the
following (non-limiting exemplary uses are also set forth):
Examples of Medical Isotopes:
[0157]
1 Isotope Description of use Actinium-225 See Thorium-229 (Th-229)
(AC-225) Actinium-227 Parent of Radium-223 (Ra-223) which is an
alpha (AC-227) emitter used to treat metastases in the skeleton
resulting from cancer (i.e., breast and prostate cancers), and
cancer radioimmunotherapy Bismuth-212 See Thorium-228 (Th-228)
(Bi-212) Bismuth-213 See Thorium-229 (Th-229) (Bi-213) Cadmium-109
Cancer detection (Cd-109) Cobalt-60 Radiation source for
radiotherapy of cancer, for food (Co-60) irradiators, and for
sterilization of medical supplies Copper-64 A positron emitter used
for cancer therapy and (Cu-64) SPECT imaging Copper-67 Beta/gamma
emitter used in cancer radio- (Cu-67) immunotherapy and diagnostic
studies (i.e., breast and colon cancers, and lymphoma)
Dysprosium-166 Cancer radioimmunotherapy (Dy-166) Erbium-169
Rheumatoid arthritis treatment, particularly for (Er-169) the small
joints associated with fingers and toes Europium-152 Radiation
source for food irradiation and for (Eu-152) sterilization of
medical supplies Europium-154 Radiation source for food irradiation
and for (Eu-154) sterilization of medical supplies Gadolinium-153
Osteoporosis detection and nuclear (Gd-153) medical quality
assurance devices Gold-198 Implant and intracavity therapy of
ovarian, (Au-198) prostate, and brain cancers Holmium-166 Multiple
myeloma treatment in targeted skeletal (Ho-166) therapy, cancer
radioimmunotherapy, bone marrow ablation, and rheumatoid arthritis
treatment Iodine-125 Osteoporosis detection, diagnostic imaging,
tracer (1-125) drugs, brain cancer treatment, radiolabeling, tumor
imaging, mapping of receptors in the brain, interstitial radiation
therapy, brachytherapy for treatment of prostate cancer,
determination of glomerular filtration rate (GFR), determination of
plasma volume, detection of deep vein thrombosis of the legs
Iodine-131 Thyroid function evaluation, thyroid disease detection,
(1-131) treatment of thyroid cancer as well as other non- malignant
thyroid diseases (i.e., Graves disease, goiters, and
hyperthyroidism), treatment of leukemia, lymphoma, and other forms
of cancer (e.g., breast cancer) using radioimmunotherapy
Iridium-192 Brachytherapy, brain and spinal cord tumor treatment,
(Ir-192) treatment of blocked arteries (i.e., arteriosclerosis and
restenosis), and implants for breast and prostate tumors
Lutetium-177 Cancer radioimmunotherapy and treatment of blocked
(Lu-177) arteries (i.e., arteriosclerosis and restenosis)
Molybdenum-99 Parent of Technetium-99m (Tc-99m) which is used for
(Mo-99) imaging the brain, liver, lungs, heart, and other organs.
Currently, Tc-99m is the most widely used radioisotope used for
diagnostic imaging of various cancers and diseases involving the
brain, heart, liver, lungs; also used in detection of deep vein
thrombosis of the legs Osmium-194 Cancer radioimmunotherapy
(Os-194) Palladium-103 Prostate cancer treatment (Pd-103)
Platinum-195m Studies on biodistribution and metabolism of
(Pt-195m) cisplatin, a chemotherapeutic drug Phosphorus-32
Polycythemia rubra vera (blood cell disease) and (P-32) leukemia
treatment, bone cancer diagnosis/treatment; colon, pancreatic, and
liver cancer treatment; radiolabeling nucleic acids for in vitro
research, diagnosis of superficial tumors, treatment of blocked
arteries (i.e., arteriosclerosis and restenosis), and intracavity
therapy Phosphorus-33 Leukemia treatment, bone disease diagnosis/
(P-33) treatment, radiolabeling, and treatment of blocked arteries
(i.e., arteriosclerosis and restenosis) Radium-223 See Actinium-227
(Ac-227) (Ra-223) Rhenium-186 Bone cancer pain relief, rheumatoid
arthritis treatment, (Re-186) and diagnosis and treatment of
lymphoma and bone, breast, colon, and liver cancers using
radioimmunotherapy Rhenium-188 Cancer diagnosis and treatment using
radio- (Re-188) immunotherapy, bone cancer pain relief, treatment
of rheumatoid arthritis, and treatment of prostate cancer
Rhodium-105 Cancer radioimmunotherapy (Rh-105) Samarium-145 Ocular
cancer treatment (Sm-145) Samarium-153 Cancer radioimmunotherapy
and bone (Sm-153) cancer pain relief Scandium-47 Cancer
radioimmunotherapy and bone (Sc-47) cancer pain relief Selenium-75
Radiotracer used in brain studies, imaging of adrenal (Se-75)
cortex by gamma-scintigraphy, lateral locations of steroid
secreting tumors, pancreatic scanning, detection of hyperactive
parathyroid glands, measure rate of bile acid loss from the
endogenous pool Strontium-85 Bone cancer detection and brain scans
(Sr-85) Strontium-89 Bone cancer pain relief, multiple myeloma
(Sr-89) treatment, and osteoblastic therapy Technetium-99m See
Molybdenum-99 (Mo-99) (Tc-99m) Thorium-228 Parent of Bismuth-212
(Bi-212) which is an alpha (Th-228) emitter used in cancer
radioimmunotherapy Thorium-229 Parent of Actinium-225 (Ac-225) and
grandparent (Th-229) of Bismuth-213 (Bi-213) which are alpha
emitters used in cancer radioimmunotherapy Thulium-170 Gamma source
for blood irradiators, energy (Tm-170) source for implanted medical
devices Tin-117m Cancer immunotherapy and bone cancer pain relief
(Sn-117m) Tungsten-188 Parent for Rhenium-188 (Re-188) which is
used for (W-188) cancer diagnostics/treatment, bone cancer pain
relief, rheumatoid arthritis treatment, and treatment of blocked
arteries (i.e., arteriosclerosis and restenosis) Xenon-127
Neuroimaging of brain disorders, high resolution (Xe-127) SPECT
studies, pulmonary function tests, and cerebral blood flow studies
Ytterbium-175 Cancer radioimmunotherapy (Yb-175) Yttrium-90
Microseeds obtained from irradiating (Y-90) Yttrium-89 (Y-89) for
liver cancer treatment Yttrium-91 A gamma-emitting label for
Yttrium-90 (Y-90) which (Y-91) is used for cancer
radioimmunotherapy (i.e., cancers) breast, colon, kidney, lung,
ovarian, prostate, pancreatic, and inoperable liver cancers)
[0158] By "randomized" or grammatical equivalents as herein applied
to nucleic acids and proteins is meant that each nucleic acid and
peptide consists of essentially random nucleotides and amino acids,
respectively. These random peptides (or nucleic acids, discussed
herein) can incorporate any nucleotide or amino acid at any
position. The synthetic process can be designed to generate
randomized proteins or nucleic acids, to allow the formation of all
or most of the possible combinations over the length of the
sequence, thus forming a library of randomized candidate bioactive
proteinaceous agents.
[0159] In one embodiment, a library is "fully randomized," with no
sequence preferences or constants at any position. In another
embodiment, the library is a "biased random" library. That is, some
positions within the sequence either are held constant, or are
selected from a limited number of possibilities. For example, the
nucleotides or amino acid residues are randomized within a defined
class, e.g., of hydrophobic amino acids, hydrophilic residues,
sterically biased (either small or large) residues, towards the
creation of nucleic acid binding domains, the creation of
cysteines, for cross-linking, prolines for SH-3 domains, serines,
threonines, tyrosines or histidines for phosphorylation sites,
etc., or to purines, etc.
[0160] A "recombinant" DNA or RNA molecule is a DNA or RNA molecule
that has been subjected to molecular manipulation in vitro.
[0161] Non-limiting examples of small molecules include compounds
that bind or interact with 191P4D12(b), ligands including hormones,
neuropeptides, chemokines, odorants, phospholipids, and functional
equivalents thereof that bind and preferably inhibit 191P4D12(b)
protein function. Such non-limiting small molecules preferably have
a molecular weight of less than about 10 kDa, more preferably below
about 9, about 8, about 7, about 6, about 5 or about 4 kDa. In
certain embodiments, small molecules physically associate with, or
bind, 191P4D12(b) protein; are not found in naturally occurring
metabolic pathways; and/or are more soluble in aqueous than
non-aqueous solutions
[0162] "Stringency" of hybridization reactions is readily
determinable by one of ordinary skill in the art, and generally is
an empirical calculation dependent upon probe length, washing
temperature, and salt concentration. In general, longer probes
require higher temperatures for proper annealing, while shorter
probes need lower temperatures. Hybridization generally depends on
the ability of denatured nucleic acid sequences to reanneal when
complementary strands are present in an environment below their
melting temperature. The higher the degree of desired homology
between the probe and hybridizable sequence, the higher the
relative temperature that can be used. As a result, it follows that
higher relative temperatures would tend to make the reaction
conditions more stringent, while lower temperatures less so. For
additional details and explanation of stringency of hybridization
reactions, see Ausubel et al., Current Protocols in Molecular
Biology, Wiley Interscience Publishers, (1995).
[0163] "Stringent conditions" or "high stringency conditions", as
defined herein, are identified by, but not limited to, those that:
(1) employ low ionic strength and high temperature for washing, for
example 0.015 M sodium chloride/0.0015 M sodium citrate/0.1% sodium
dodecyl sulfate at 50.degree. C.; (2) employ during hybridization a
denaturing agent, such as formamide, for example, 50% (v/v)
formamide with 0.1% bovine serum albumin/0.1% Ficoll/0.1%
polyvinylpyrrolidone/50 mM sodium phosphate buffer at pH 6.5 with
750 mM sodium chloride, 75 mM sodium citrate at 42.degree. C.; or
(3) employ 50% formamide, 5.times.SSC (0.75 M NaCl, 0.075 M sodium
citrate), 50 mM sodium phosphate (pH 6.8), 0.1% sodium
pyrophosphate, 5.times. Denhardt's solution, sonicated salmon sperm
DNA (50 .mu.g/ml), 0.1% SDS, and 10% dextran sulfate at 42.degree.
C., with washes at 42.degree. C. in 0.2.times.SSC (sodium
chloride/sodium. citrate) and 50% formamide at 55.degree. C.,
followed by a high-stringency wash consisting of 0.1.times.SSC
containing EDTA at 55.degree. C. "Moderately stringent conditions"
are described by, but not limited to, those in Sambrook et al.,
Molecular Cloning: A Laboratory Manual, New York: Cold Spring
Harbor Press, 1989, and include the use of washing solution and
hybridization conditions (e.g., temperature, ionic strength and
%SDS) less stringent than those described above. An example of
moderately stringent conditions is overnight incubation at
37.degree. C. in a solution comprising: 20% formamide, 5.times.SSC
(150 mM NaCl, 15 mM trisodium citrate), 50 mM sodium phosphate (pH
7.6), 5.times. Denhardt's solution, 10% dextran sulfate, and 20
mg/mL denatured sheared salmon sperm DNA, followed by washing the
filters in 1.times.SSC at about 37-50.degree. C. The skilled
artisan will recognize how to adjust the temperature, ionic
strength, etc. as necessary to accommodate factors such as probe
length and the like.
[0164] An HLA "supermotif" is a peptide binding specificity shared
by HLA molecules encoded by two or more HLA alleles. Overall
phenotypic frequencies of HLA-supertypes in different ethnic
populations are set forth in Table IV (F). The non-limiting
constituents of various supetypes are as follows.
[0165] A2: A*0201, A*0202, A*0203, A*0204, A*0205, A*0206, A*6802,
A*6901, A*0207
[0166] A3: A3, A11, A31, A*3301, A*6801, A*0301, A*1101, A*3101
[0167] B7: B7, B*3501-03, B*51, B*5301, B*5401, B*5501, B*5502,
B*5601, B*6701, B*7801, B*0702, B*5101, B*5602
[0168] B44: B*3701, B*4402, B*4403, B*60 (B*4001), B61 (B*4006)
[0169] A1: A*0102, A*2604, A*3601, A*4301, A*8001
[0170] A24: A*24, A*30, A*2403, A*2404, A*3002, A*3003
[0171] B27: B*1401-02, B*1503, B*1509, B*1510, B*1518, B*3801-02,
B*3901, B*3902, B*3903-04, B*4801-02, B*7301, B*2701 -08
[0172] B58: B*1516, B*1517, B*5701, B*5702, B58
[0173] B62: B*4601, B52, B*1501 (B62), B*1502 (B75), B*1513
(B77)
[0174] Calculated population coverage afforded by different
HLA-supertype combinations are set forth in Table IV (G).
[0175] As used herein "to treat" or "therapeutic" and grammatically
related terms, refer to any improvement of any consequence of
disease, such as prolonged survival, less morbidity, and/or a
lessening of side effects which are the byproducts of an
alternative therapeutic modality; full eradication of disease is
not required.
[0176] A "transgenic animal" (e.g., a mouse or rat) is an animal
having cells that contain a transgene, which transgene was
introduced into the animal or an ancestor of the animal at a
prenatal, e.g., an embryonic stage. A "transgene" is a DNA that is
integrated into the genome of a cell from which a transgenic animal
develops.
[0177] As used herein, an HLA or cellular immune response "vaccine"
is a composition that contains or encodes one or more peptides of
the invention. There are numerous embodiments of such vaccines,
such as a cocktail of one or more individual peptides; one or more
peptides of the invention comprised by a polyepitopic peptide; or
nucleic acids that encode such individual peptides or polypeptides,
e.g., a minigene that encodes a polyepitopic peptide. The "one or
more peptides" can include any whole unit integer from 1-150 or
more, e.g., at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48,
49, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115,
120, 125, 130, 135, 140, 145, or 150 or more peptides of the
invention. The peptides or polypeptides can optionally be modified,
such as by lipidation, addition of targeting or other sequences.
HLA class I peptides of the invention can be admixed with, or
linked to, HLA class II peptides, to facilitate activation of both
cytotoxic T lymphocytes and helper T lymphocytes. HLA vaccines can
also comprise peptide-pulsed antigen presenting cells, e.g.,
dendritic cells.
[0178] The term "variant" refers to a molecule that exhibits a
variation from a described type or norm, such as a protein that has
one or more different amino acid residues in the corresponding
position(s) of a specifically described protein (e.g. the
191P4D12(b) protein shown in FIG. 2 or FIG. 3. An analog is an
example of a variant protein. Splice isoforms and single
nucleotides polymorphisms (SNPs) are further examples of
variants.
[0179] The "191P4D12(b)-related proteins" of the invention include
those specifically identified herein, as well as allelic variants,
conservative substitution variants, analogs and homologs that can
be isolated/generated and characterized without undue
experimentation following the methods outlined herein or readily
available in the art. Fusion proteins that combine parts of
different 191P4D12(b) proteins or fragments thereof, as well as
fusion proteins of a 191P4D12(b) protein and a heterologous
polypeptide are also included. Such 191P4D12(b) proteins are
collecfively referred to as the 191P4D12(b)-related proteins, the
proteins of the invention, or 191P4D12(b). The term
"191P4D12(b)-related protein" refers to a polypeptide fragment or a
191P4D12(b) protein sequence of 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,
14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, or more than 25
amino acids; or, at least 30, 35, 40, 45, 50, 55, 60, 65, 70, 80,
85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150,
155, 160, 165, 170, 175, 180, 185, 190, 195, 200, 225, 250, 275,
300, 325, 350, 375, 400, 425, 450, 475, 500, 525, 550, 575, or 576
or more amino acids.
[0180] II.) 191P4D12(b) Polynucleotides
[0181] One aspect of the invention provides polynucleotides
corresponding or complementary to all or part of a 191P4D12(b)
gene, mRNA, and/or coding sequence, preferably in isolated form,
including polynucleotides encoding a 191P4D12(b)-related protein
and fragments thereof, DNA, RNA, DNA/RNA hybrid, and related
molecules, polynucleotides or oligonucleotides complementary to a
191P4D12(b) gene or mRNA sequence or a part thereof, and
polynucleotides or oligonucleotides that hybridize to a 191P4D12(b)
gene, mRNA, or to a 191P4D12(b) encoding polynucleotide
(collectively, "191P4D12(b) polynucleotides"). In all instances
when referred to in this section, T can also be U in FIG. 2.
[0182] Embodiments of a 191P4D12(b) polynucleotide include: a
191P4D12(b) polynucleotide having the sequence shown in FIG. 2, the
nucleotide sequence of 191P4D12(b) as shown in FIG. 2 wherein T is
U; at least 10 contiguous nucleotides of a polynucleotide having
the sequence as shown in FIG. 2; or, at least 10 contiguous
nucleotides of a polynucleotide having the sequence as shown in
FIG. 2 where T is U. For example, embodiments of 191P4D12(b)
nucleotides comprise, without limitation:
[0183] (I) a polynucleotide comprising, consisting essentially of,
or consisting of a sequence as shown in FIG. 2, wherein T can also
be U;
[0184] (II) a polynucleotide comprising, consisting essentially of,
or consisting of the sequence as shown in FIG. 2A, from nucleotide
residue number 264 through nucleotide residue number 1796,
including the stop codon, wherein T can also be U;
[0185] (III) a polynucleotide comprising, consisting essentially
of, or consisting of the sequence as shown in FIG. 2B, from
nucleotide residue number 264 through nucleotide residue number
1796, including the stop codon, wherein T can also be U;
[0186] (IV) a polynucleotide comprising, consisting essentially of,
or consisting of the sequence as shown in FIG. 2C, from nucleotide
residue number 264 through nucleotide residue number 1796,
including the a stop codon, wherein T can also be U;
[0187] (V) a polynucleotide comprising, consisting essentially of,
or consisting of the sequence as shown in FIG. 2D, from nucleotide
residue number 264 through nucleotide residue number 1796,
including the stop codon, wherein T can also be U;.
[0188] (VI) a polynucleotide comprising, consisting essentially of,
or consisting of the sequence as shown in FIG. 2E, from nucleotide
residue number 264 through nucleotide residue number 1796,
including the stop codon, wherein T can also be U;,
[0189] (VII) a polynucleotide comprising, consisting essentially
of, or consisting of the sequence as shown in FIG. 2F, from
nucleotide residue number 789 through nucleotide residue number
1676, including the stop codon, wherein T can also be U;
[0190] (VIII) a polynucleotide comprising, consisting essentially
of, or consisting of the sequence as shown in FIG. 2G, from
nucleotide residue number 264 through nucleotide residue number
1721, including the stop codon, wherein T can also be U;
[0191] (IX) a polynucleotide comprising, consisting essentially of,
or consisting of the sequence as shown in FIG. 2H, from nucleotide
residue number 264 through nucleotide residue number 1796,
including the stop codon, wherein T can also be U;
[0192] (X) a polynucleotide comprising, consisting essentially of,
or consisting of the sequence as shown in FIG. 2I, from nucleotide
residue number 708 through nucleotide residue number 1121,
including the stop codon, wherein T can also be U;
[0193] (XI) a polynucleotide comprising, consisting essentially of,
or consisting of the sequence as shown in FIG. 2J, from nucleotide
residue number 264 through nucleotide residue number 1796,
including the stop codon, wherein T can also be U;
[0194] (XII) a polynucleotide comprising, consisting essentially
of, or consisting of the sequence as shown in FIG. 2K, from
nucleotide residue number 264 through nucleotide residue number
1796, including the stop codon, wherein T can also be U;
[0195] (XIII) a polynucleotide comprising, consisting essentially
of, or consisting of the sequence as shown in FIG. 2L, from
nucleotide residue number 264 through nucleotide residue number
1796, including the stop codon, wherein T can also be U;
[0196] (XIV) a polynucleotide comprising, consisting essentially
of, or consisting of the sequence as shown in FIG. 2M, from
nucleotide residue number 264 through nucleotide residue number
1799, including the stop codon, wherein T can also be U;
[0197] (XV) a polynucleotide comprising, consisting essentially of,
or consisting of the sequence as shown in FIG. 2N, from nucleotide
residue number 708 through nucleotide residue number 1121,
including the stop codon, wherein T can also be U;
[0198] (XVI) a polynucleotide that encodes a 191P4D12(b)-related
protein that is at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or
100% homologous to an entire amino acid sequence shown in FIGS.
2A-N;
[0199] (XVII) a polynucleotide that encodes a 191P4D12(b)-related
protein that is at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or
100% identical to an entire amino acid sequence shown in FIGS.
2A-N;
[0200] (XVIII) a polynucleotide that encodes at least one peptide
set forth in Tables VIII-XXI and XXII-XLIX;
[0201] (XIX) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3A-B and 3E-G in any whole number
increment up to 510 that includes at least 1, 2, 3, 4, 5, 6, 7, 8,
9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,
26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s)
having a value greater than 0.5 in the Hydrophilicity profile of
FIG. 5;
[0202] (XX) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3A-B and 3E-G in any whole number
increment up to 510 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a
value less than 0.5 in the Hydropathicity profile of FIG. 6;
[0203] (XXI) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3A-B and 3E-G in any whole number
increment up to 510 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a
value greater than 0.5 in the Percent Accessible Residues profile
of FIG. 7;
[0204] (XXII) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3A-B and 3E-G in any whole number
increment up to 510 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a
value greater than 0.5 in the Average Flexibility profile of FIG.
8;
[0205] (XXIII) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3A-B and 3E-G in any whole number
increment up to 510 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a
value greater than 0.5 in the Beta-turn profile of FIG. 9;
[0206] (XXIV) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3C in any whole number increment up to
295 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Hydrophilicity profile of FIG. 5;
[0207] (XXV) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3C in any whole number increment up to
295 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value less than 0.5
in the Hydropathicity profile of FIG. 6;
[0208] (XXVI) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3C in any whole number increment up to
295 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Percent Accessible Residues profile of FIG. 7;
[0209] (XXVII) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3C in any whole number increment up to
295 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Average Flexibility profile of FIG. 8;
[0210] (XXVIII) a polynucleotide that encodes a peptide region of
at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3C in any whole number increment up to
295 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Beta-turn profile of FIG. 9
[0211] (XXIX) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3D in any whole number increment up to
485 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Hydrophilicity profile of FIG. 5;
[0212] (XXX) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3D in any whole number increment up to
485 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value less than 0.5
in the Hydropathicity profile of FIG. 6;
[0213] (XXXI) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3D in any whole number increment up to
485 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Percent Accessible Residues profile of FIG. 7;
[0214] (XXXII) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3D in any whole number increment up to
485 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Average Flexibility profile of FIG. 8;
[0215] (XXXIII) a polynucleotide that encodes a peptide region of
at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3D in any whole number incrementup to
485 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Beta-turn profile of FIG. 9
[0216] (XXXIV) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3H in any whole number increment up to
511 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Hydrophilicity profile of FIG. 5;
[0217] (XXXV) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3H in any whole number increment up to
511 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value less than 0.5
in the Hydropathicity profile of FIG. 6;
[0218] (XXXVI) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3H in any whole number increment up to
511 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Percent Accessible Residues profile of FIG. 7;
[0219] (XXXVII) a polynucleotide that encodes a peptide region of
at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3H in any whole number increment up to
511 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Average Flexibility profile of FIG. 8;
[0220] (XXXVIII) a polynucleotide that encodes a peptide region of
at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIG. 3H in any whole number increment up to
511 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Beta-turn profile of FIG. 9
[0221] (XXXIX) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3I-J in any whole number increment up
to 137 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Hydrophilicity profile of FIG. 5;
[0222] (XL) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3I-J in any whole number increment up
to 137 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value less than 0.5
in the Hydropathicity profile of FIG. 6;
[0223] (XLI) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3I-J in any whole number increment up
to 137 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Percent Accessible Residues profile of FIG. 7;
[0224] (XLII) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3I-J in any whole number increment up
to 137 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Average Flexibility profile of FIG. 8;
[0225] (XLIII) a polynucleotide that encodes a peptide region of at
least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino
acids of a peptide of FIGS. 3I-J in any whole number increment up
to 137 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Beta-turn profile of FIG. 9
[0226] (XLIV) a polynucleotide that is fully complementary to a
polynucleotide of any one of (I)-(XLIII).
[0227] (XLV) a peptide that is encoded by any of (I) to (XLIV);
and
[0228] (XLVI) a composition comprising a polynucleotide of any of
(I)-(XLIII) or peptide of (XLV) together with a pharmaceutical
excipient and/or in a human unit dose form.
[0229] (XLVII) a method of using a polynucleotide of any (I)-(XLIV)
or peptide of (XLV) or a composition of (XLVI) in a method to
modulate a cell expressing 191P4D12(b),
[0230] (XLVIII) a method of using a polynucleotide of any
(I)-(XLIV) or peptide of (XLV) or a composition of (XLVI) in a
method to diagnose, prophylax, prognose, or treat an individual who
bears a cell expressing 191P4D12(b)
[0231] (XLIX) a method of using a polynucleotide of any (I)-(XLIV)
or peptide of (XLV) or a composition of (XLVI) in a method to
diagnose, prophylax, prognose, or treat an individual who bears a
cell expressing 191P4D12(b), said cell from a cancer of a tissue
listed in Table I;
[0232] (L) a method of using a polynucleotide of any (I)-(XLIV) or
peptide of (XLV) or a composition of (XLVI) in a method to
diagnose, prophylax, prognoses or treat a a cancer,
[0233] (LI) a method of using a polynucleotide of any (I)-(XLIV) or
peptide of (XLV) or a composition of (XLVI) in a method to
diagnose, prophylax, prognose, or treat a a cancer of a tissue
listed in Table I; and,
[0234] (LII) a method of using a polynucleotide of any (I)-(XLIV)
or peptide of (XLV) or a composition of (XLVI) in a method to
identify or characterize a modulator of a cell expressing
191P4D12(b).
[0235] As used herein, a range is understood to disclose
specifically all whole unit positions thereof.
[0236] Typical embodiments of the invention disclosed herein
include 191P4D12(b) polynucleotides that encode specific portions
of 191P4D12(b) mRNA sequences (and those which are complementary to
such sequences) such as those that encode the proteins and/or
fragments thereof, for example:
[0237] (a) 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,
19, 20, 21, 22, 23, 24, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75,
80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145,
150, 155, 160, 165, 170, 175, 180, 185, 190, 195, 200, 225, 250,
275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 505 or 510 more
contiguous amino acids of 191P4D12(b) variant 1; the maximal
lengths relevant for other variants are: variant 2, 510 amino
acids; variant 6, 295 amino acids, variant 7, 485 amino acids,
variant 10, 510 amino acids, variant 11, 510 amoni acids, variant
12, 510 amoni acids, variant 13, 511 amino acids, variant 9, 137
amino acids, and variant 14, 137 amino acids.
[0238] For example, representative embodiments of the invention
disclosed herein include: polynucleotides and their encoded
peptides themselves encoding about amino acid 1 to about amino acid
10 of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 10 to about amino acid 20
of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 20 to about amino acid 30
of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 30 to about amino acid 40
of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 40 to about amino acid 50
of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 50 to about amino acid 60
of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 60 to about amino acid 70
of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 70 to about amino acid 80
of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 80 to about amino acid 90
of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polynucleotides encoding about amino acid 90 to about amino acid
100 of the 191P4D12(b) protein shown in FIG. 2 or FIG. 3, in
increments of about 10 amino acids, ending at the carboxyl terminal
amino acid set forth in FIG. 2 or FIG. 3. Accordingly,
polynucleotides encoding portions of the amino acid sequence (of
about 10 amino acids), of amino acids, 100 through the carboxyl
terminal amino acid of the 191P4D12(b) protein are embodiments of
the invention. Wherein it is understood that each particular amino
acid position discloses that position plus or minus five amino acid
residues.
[0239] Polynucleotides encoding relatively long portions of a
191P4D12(b) protein are also within the scope of the invention. For
example, polynucleotides encoding from about amino acid 1 (or 20 or
30 or 40 etc.) to about amino acid 20, (or 30, or 40 or 50 etc.) of
the 191P4D12(b) protein "or variant" shown in FIG. 2 or FIG. 3 can
be generated by a variety of techniques well known in the art.
These polynucleotide fragments can include any portion of the
191P4D12(b) sequence as shown in FIG. 2.
[0240] Additional illustrative embodiments of the invention
disclosed herein include 191P4D12(b) polynucleotide fragments
encoding one or more of the biological motifs contained within a
191P4D12(b) protein "or variant" sequence, including one or more of
the motif-bearing subsequences of a 191P4D12(b) protein "or
variant" set forth in Tables VIII-XXI and XXII-XLIX. In another
embodiment, typical polynucleotide fragments of the invention
encode one or more of the regions of 191P4D12(b) protein or variant
that exhibit homology to a known molecule. In another embodiment of
the invention, typical polynucleotide fragments can encode one or
more of the 191P4D12(b) protein or variant N-glycosylation sites,
cAMP and cGMP-dependent protein kinase phosphorylation sites,
casein kinase II phosphorylation sites or N-myristoylation site and
amidation sites.
[0241] Note that to determine the starting position of any peptide
set forth in Tables VIII-XXI and Tables XXII to XLIX (collectively
HLA Peptide Tables) respective to its parental protein, e.g.,
variant 1, variant 2, etc., reference is made to three factors: the
particular variant, the length of the peptide in an HLA Peptide
Table, and the Search Peptides listed in Table VII. Generally, a
unique Search Peptide is used to obtain HLA peptides for a
particular variant. The position of each Search Peptide relative to
its respective parent molecule is listed in Table VII. Accordingly,
if a Search Peptide begins at position "X", one must add the value
"X minus 1" to each position in Tables VIII-XXI and Tables XXII-IL
to obtain the actual position of the HLA peptides in their parental
molecule. For example if a particular Search Peptide begins at
position 150 of its parental molecule, one must add 150-1, i.e.,
149 to each HLA peptide amino acid position to calculate the
position of that amino acid in the parent molecule.
[0242] II.A.) Uses of 191P4D12(b) Polynucleotides
[0243] II.A.1.) Monitoring of Genetic Abnormalities
[0244] The polynucleotides of the preceding paragraphs have a
number of different specific uses. The human 191P4D12(b) gene maps
to the chromosomal location set forth in the Example entitled
"Chromosomal Mapping of 191P4D12(b)." For example, because the
191P4D12(b) gene maps to this chromosome, polynucleotides that
encode different regions of the 191P4D12(b) proteins are used to
characterize cytogenetic abnormalities of this chromosomal locale,
such as abnormalities that are identified as being associated with
various cancers. In certain genes, a variety of chromosomal
abnormalities including rearrangements have been identified as
frequent cytogenetic abnormalities in a number of different cancers
(see e.g. Krajinovic et al., Mutat. Res. 382(3-4): 81-83 (1998);
Johansson et al., Blood 86(10): 3905-3914 (1995) and Finger et al.,
P.N.A.S. 85(23): 9158-9162 (1988)). Thus, polynucleotides encoding
specific regions of the 191P4D12(b) proteins provide new tools that
can be used to delineate, with greater precision than previously
possible, cytogenetic abnormalities in the chromosomal region that
encodes 191P4D12(b) that may contribute to the malignant phenotype.
In this context, these polynucleotides satisfy a need in the art
for expanding the sensitivity of chromosomal screening in order to
identify more subtle and less common chromosomal abnormalities (see
e.g. Evans et al., Am. J. Obstet. Gynecol 171(4): 1055-1057
(1994)).
[0245] Furthermore, as 191P4D12(b) was shown to be highly expressed
in prostate and other cancers, 191P4D12(b) polynucleotides are used
in methods assessing the status of 191P4D12(b) gene products in
normal versus cancerous tissues. Typically, polynucleotides that
encode specific regions of the 191P4D12(b) proteins are used to
assess the presence of perturbations (such as deletions,
insertions, point mutations, or alterations resulting in a loss of
an antigen etc.) in specific regions of the 191P4D12(b) gene, such
as regions containing one or more motifs. Exemplary assays include
both RT-PCR assays as well as single-strand conformation
polymorphism (SSCP) analysis (see, e.g., Marrogi et al., J. Cutan.
Pathol. 26(8): 369-378 (1999), both of which utilize
polynucleotides encoding specific regions of a protein to examine
these regions within the protein.
[0246] II.A.2.) Antisense Embodiments
[0247] Other specifically contemplated nucleic acid related
embodiments of the invention disclosed herein are genomic DNA,
cDNAs, ribozymes, and antisense molecules, as well as nucleic acid
molecules based on an alternative backbone, or including
alternative bases, whether derived from natural sources or
synthesized, and include molecules capable of inhibiting the RNA or
protein expression of 191P4D12(b). For example, antisense molecules
can be RNAs or other molecules, including peptide nucleic acids
(PNAs) or non-nucleic acid molecules such as phosphorothioate
derivatives that specifically bind DNA or RNA in a base
pair-dependent manner. A skilled artisan can readily obtain these
classes of nucleic acid molecules using the 191P4D12(b)
polynucleotides and polynucleotide sequences disclosed herein.
[0248] Antisense technology entails the administration of exogenous
oligonucleotides that bind to a target polynucleotide located
within the cells. The term "antisense" refers to the fact that such
oligonucleotides are complementary to their intracellular targets,
e.g., 191P4D12(b). See for example, Jack Cohen,
Oligodeoxynucleotides, Antisense Inhibitors of Gene Expression, CRC
Press, 1989; and Synthesis 1:1-5 (1988). The 191P4D12(b) antisense
oligonucleotides of the present invention include derivatives such
as S-oligonucleotides (phosphorothioate derivatives or S-oligos,
see, Jack Cohen, supra), which exhibit enhanced cancer cell growth
inhibitory action. S-oligos (nucleoside phosphorothioates) are
isoelectronic analogs of an oligonucleotide (O-oligo) in which a
non bridging oxygen atom of the phosphate group is replaced by a
sulfur atom. The S-oligos of the present invention can be prepared
by treatment of the corresponding O-oligos with
3H-1,2-benzodithiol-3-one-1,1-dioxide, which is a sulfur transfer
reagent. See, e.g., Iyer, R. P. et al., J. Org. Chem. 55:4693-4698
(1990); and Iyer, R. P. et al., J. Am. Chem. Soc. 112:1253-1254
(1990). Additional 191P4D12(b) antisense oligonucleotides of the
present invention include morpholino antisense oligonucleotides
known in the art (see; e.g., Partridge et al., 1996, Antisense
& Nucleic Acid Drug Development 6: 169-175).
[0249] The 191P4D12(b) antisense oligonucleotides of the present
invention typically can be RNA or DNA that is complementary to and
stably hybridizes with the first 100 5' codons or last 100 3'
codons of a 191P4D12(b) genomic sequence or the corresponding mRNA.
Absolute complementarity is not required, although high degrees of
complementarity are preferred. Use of an oligonucleotide
complementary to this region allows for the selective hybridization
to 191P4D12(b) mRNA and not to mRNA specifying other regulatory
subunits of protein kinase. In one embodiment, 191P4D12(b)
antisense oligonucleotides of the present invention are 15 to
30-mer fragments of the antisense DNA molecule that have a sequence
that hybridizes to 191P4D12(b) mRNA. Optionally, 191P4D12(b)
antisense oligonucleotide is a 30-mer oligonucleotide that is
complementary to a region in the first 10 5' codons or last 10 3'
codons of 191P4D12(b). Alternatively, the antisense molecules are
modified to employ ribozymes in the inhibition of 191P4D12(b)
expression, see, e.g., L. A. Couture & D. T. Stinchcomb; Trends
Genet 12: 510-515 (1996).
[0250] II.A.3.) Primers and Primer Pairs
[0251] Further specific embodiments of these nucleotides of the
invention include primers and primer pairs, which allow the
specific amplification of polynucleotides of the invention or of
any specific parts thereof, and probes that selectively or
specifically hybridize to nucleic acid molecules of the invention
or to any part thereof. Probes can be labeled with a detectable
marker, such as, for example, a radioisotope, fluorescent compound,
bioluminescent compound, a chemiluminescent compound, metal
chelator or enzyme. Such probes and primers are used to detect the
presence of a 191P4D12(b) polynucleotide in a sample and as a means
for detecting a cell expressing a 191P4D12(b) protein.
[0252] Examples of such probes include polypeptides comprising all
or part of the human 191P4D12(b) cDNA sequence shown in FIG. 2.
Examples of pnmer pairs capable of specifically amplifying
191P4D12(b) mRNAs are also described in the Examples. As will be
understood by the skilled artisan, a great many different primers
and probes can be prepared based on the sequences provided herein
and used effectively to amplify and/or detect a 191P4D12(b)
mRNA.
[0253] The 191P4D12(b) polynucleotides of the invention are useful
for a variety of purposes, including but not limited to their use
as probes and primers for the amplification and/or detection of the
191P4D12(b) gene(s), mRNA(s), or fragments thereof; as reagents for
the diagnosis and/or prognosis of prostate cancer and other
cancers; as coding sequences capable of directing the expression of
191P4D12(b) polypeptides; as tools for modulating or inhibiting the
expression of the 191P4D12(b) gene(s) and/or translation of the
191P4D12(b) transcript(s); and as therapeutic agents.
[0254] The present invention includes the use of any probe as
described herein to identify and isolate a 191P4D12(b) or
191P4D12(b) related nucleic acid sequence from a naturally
occurring source, such as humans or other mammals, as well as the
isolated nucleic acid sequence per se, which would comprise all or
most of the sequences found in the probe used.
[0255] II.A.4.) Isolation of 191P4D12(b)-Encoding Nucleic Acid
Molecules
[0256] The 191P4D12(b) cDNA sequences described herein enable the
isolation of other polynucleotides encoding 191P4D12(b) gene
product(s), as well as the isolation of polynucleotides encoding
191P4D12(b) gene product homologs, alternatively spliced isoforms,
allelic variants, and mutant forms of a 191P4D12(b) gene product as
well as polynucleotides that encode analogs of 191P4D12(b)-related
proteins. Various molecular cloning methods that can be employed to
isolate full length cDNAs encoding a 191P4D12(b) gene are well
known (see, for example, Sambrook, J. et al., Molecular Cloning: A
Laboratory Manual, 2d edition, Cold Spring Harbor Press, New York,
1989; Current Protocols in Molecular Biology. Ausubel et al., Eds.,
Wiley and Sons, 1995). For example, lambda phage cloning
methodologies can be conveniently employed, using commercially
available cloning systems (e.g., Lambda ZAP Express, Stratagene).
Phage clones containing 191P4D12(b) gene cDNAs can be identified by
probing with a labeled 191P4D12(b) cDNA or a fragment thereof. For
example, in one embodiment, a 191P4D12(b) cDNA (e.g., FIG. 2) or a
portion thereof can be synthesized and used as a probe to retrieve
overlapping and full-length cDNAs corresponding to a 191P4D12(b)
gene. A 191P4D12(b) gene itself can be isolated by screening
genomic DNA libraries, bacterial artificial chromosome libraries
(BACs), yeast artificial chromosome libraries (YACs), and the like,
with 191P4D12(b) DNA probes or primers.
[0257] II.A.5.) Recombinant Nucleic Acid Molecules and Host-Vector
Systems
[0258] The invention also provides recombinant DNA or RNA molecules
containing a 191P4D12(b) polynucleotide, a fragment, analog or
homologue thereof, including but not limited to phages, plasmids,
phagemids, cosmids, YACs, BACs, as well as various viral and
non-viral vectors well known in the art, and cells transformed or
transfected with such recombinant DNA or RNA molecules. Methods for
generating such molecules are well known (see, for example,
Sambrook et al., 1989, supra).
[0259] The invention further provides a host-vector system
comprising a recombinant DNA molecule containing a 191P4D12(b)
polynucleotide, fragment, analog or homologue thereof within a
suitable prokaryotic or eukaryotic host cell. Examples of suitable
eukaryotic host cells include a yeast cell, a plant cell, or an
animal cell, such as a mammalian cell or an insect cell (e.g., a
baculovirus-infectible cell such as an Sf9 or HighFive cell).
Examples of suitable mammalian cells include various prostate
cancer cell lines such as DU145 and TsuPr1, other transfectable or
transducible prostate cancer cell lines, primary cells (PrEC), as
well as a number of mammalian cells routinely used for the
expression of recombinant proteins (e.g., COS, CHO, 293, 293T
cells). More particularly, a polynucleotide comprising the coding
sequence of 191P4D12(b) or a fragment, analog or homolog thereof
can be used to generate 191P4D12(b) proteins or fragments thereof
using any number of host-vector systems routinely used and widely
known in the art.
[0260] A wide range of host-vector systems suitable for the
expression of 191P4D12(b) proteins or fragments thereof are
available, see for example, Sambrook et al., 1989, supra; Current
Protocols in Molecular Biology, 1995, supra). Preferred vectors for
mammalian expression include but are not limited to pcDNA 3.1
myc-Histag (Invitrogen) and the retroviral vector pSR.alpha.tkneo
(Muller et al., 1991, MCB 11:1785). Using these expression vectors,
191P4D12(b) can be expressed in several prostate cancer and
non-prostate cell lines, including for example 293, 293T, rat-1,
NIH 3T3 and TsuPr1. The host-vector systems of the invention are
useful for the production of a 191P4D12(b) protein or fragment
thereof. Such host-vector systems can be employed to study the
functional properties of 191P4D12(b) and 191P4D12(b) mutations or
analogs.
[0261] Recombinant human 191P4D12(b) protein or an analog or
homolog or fragment thereof can be produced by mammalian cells
transfected with a construct encoding a 191P4D12(b)-related
nucleotide. For example, 293T cells can be transfected with an
expression plasmid encoding 191P4D12(b) or fragment, analog or
homolog thereof, a 191P4D12(b)-related protein is expressed in the
293T cells, and the recombinant 191P4D12(b) protein is isolated
using standard purification methods (e.g., affinity purification
using anti-191P4D12(b) antibodies). In another embodiment, a
191P4D12(b) coding sequence is subcloned into the retroviral vector
pSR.alpha.MSVtkneo and used to infect various mammalian cell lines,
such as NIH 3T3, TsuPr1, 293 and rat-1 in order to establish
191P4D12(b) expressing cell lines. Various other expression systems
well known in the art can also be employed. Expression constructs
encoding a leader peptide joined in frame to a 191P4D12(b) coding
sequence can be used for the generation of a secreted form of
recombinant 191P4D12(b) protein.
[0262] As discussed herein, redundancy in the genetic code permits
variation in 191P4D12(b) gene sequences. In particular, it is known
in the art that specific host species often have specific codon
preferences, and thus one can adapt the disclosed sequence as
preferred for a desired host. For example, preferred analog codon
sequences typically have rare codons (i.e., codons having a usage
frequency of less than about 20% in known sequences of the desired
host) replaced with higher frequency codons. Codon preferences for
a specific species are calculated, for example, by utilizing codon
usage tables available on the INTERNET such as at URL
dna.affrc.go.jp/.about.nakamura/codon.html.
[0263] Additional sequence modifications are known to enhance
protein expression in a cellular host. These include elimination of
sequences encoding spurious polyadenylation signals, exon/intron
splice site signals, transposon-like repeats, and/or other such
well-characterized sequences that are deleterious to gene
expression. The GC content of the sequence is adjusted to levels
average for a given cellular host, as calculated by reference to
known genes expressed in the host cell. Where possible, the
sequence is modified to avoid predicted hairpin secondary mRNA
structures. Other useful modifications include the addition of a
translational initiation consensus sequence at the start of the
open reading frame, as described in Kozak, Mol. Cell Biol.,
9:5073-5080 (1989). Skilled artisans understand that the general
rule that eukaryotic ribosomes initiate translation exclusively at
the 5' proximal AUG codon is abrogated only under rare conditions
(see, e.g., Kozak PNAS 92(7): 2662-2666, (1995) and Kozak NAR
15(20): 8125-8148 (1987)).
[0264] III.) 191P4D12(b)-related Proteins
[0265] Another aspect of the present invention provides
191P4D12(b)-related proteins. Specific embodiments of 191P4D12(b)
proteins comprise a polypeptide having all or part of the amino
acid sequence of human 191P4D12(b) as shown in FIG. 2 or FIG. 3.
Alternatively, embodiments of 191P4D12(b) proteins comprise
variant, homolog or analog polypeptides that have alterations in
the amino acid sequence of 191P4D12(b) shown in FIG. 2 or FIG.
3.
[0266] Embodiments of a 191P4D12(b) polypeptide include: a
191P4D12(b) polypeptide having a sequence shown in FIG. 2, a
peptide sequence of a 191P4D12(b) as shown in FIG. 2 wherein T is
U; at least 10 contiguous nucleotides of a polypeptide having the
sequence as shown in FIG. 2; or, at least 10 contiguous peptides of
a polypeptide having the sequence as shown in FIG. 2 where T is U.
For example, embodiments of 191P4D12(b) peptides comprise, without
limitation:
[0267] (I) a protein comprising, consisting essentially of, or
consisting of an amino acid sequence as shown in FIGS. 2A-N or
FIGS. 3A-J;
[0268] (II) a 191P4D12(b)-related protein that is at least 90, 91,
92, 93, 94, 95, 96, 97, 98, 99 or 100% homologous to an entire
amino acid sequence shown in FIGS. 2A-N or 3A-J;
[0269] (III) a 191P4D12(b)-related protein that is at least 90, 91,
92, 93, 94, 95, 96, 97, 98, 99 or 100% identical to an entire amino
acid sequence shown in FIGS. 2A-N or 3A-J;
[0270] (IV) a protein that comprises at least one peptide set forth
in Tables VIII to XLIX, optionally with a proviso that it is not an
entire protein of FIG. 2;
[0271] (V) a protein that comprises at least one peptide set forth
in Tables VIII-XXI, collectively, which peptide is also set forth
in Tables XXII to XLIX, collectively, optionally with a proviso
that it is not an entire protein of FIG. 2;
[0272] (VI) a protein that comprises at least two peptides selected
from the peptides set forth in Tables VIII-XLIX, optionally with a
proviso that it is not an entire protein of FIG. 2;
[0273] (VIl) a protein that comprises at least two peptides
selected from the peptides set forth in Tables VIII to XLIX
collectively, with a proviso that the protein is not a contiguous
sequence from an amino acid sequence of FIG. 2;
[0274] (VIII) a protein that comprises at least one peptide
selected from the peptides set forth in Tables VIII-XXI; and at
least one peptide selected from the peptides set forth in Tables
XXII to XLIX, with a proviso that the protein is not a contiguous
sequence from an amino acid sequence of FIG. 2;
[0275] (IX) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIGS.
3A-B or 3E-G, in any whole number increment up to 510 respectively
that includes at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,
14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30,
31, 32, 33, 34, 35 amino acid position(s) having a value greater
than 0.5 in the Hydrophilicity profile of FIG. 5;
[0276] (X) a polypeptide comprising at least 5, 6, 7, 8, 9, 10, 11,
12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28,
29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIGS. 3A-B
or 3E-G, in any whole number increment up to 510 respectively that
includes at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,
13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,
30, 31, 32, 33, 34, 35 amino acid position(s) having a value less
than 0.5 in the Hydropathicity profile of FIG. 6;
[0277] (XI) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIGS.
3A-B or 3E-G, in any whole number increment up to 510 respectively
that includes at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,
12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28,
29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value
greater than 0.5 in the Percent Accessible Residues profile of FIG.
7;
[0278] (XII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIGS.
3A-B or 3E-G, in any whole number increment up to 510 respectively
that includes at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,
12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28,
29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value
greater than 0.5 in the Average Flexibility profile of FIG. 8;
[0279] (XIII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, amino acids of a protein of FIGS. 3A-B
or 3E-G in any whole number increment up to 510 respectively that
includes at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,
13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,
30, 31, 32, 33, 34, 35 amino acid position(s) having a value
greater than 0.5 in the Beta-turn profile of FIG. 9;
[0280] (XIV) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3C,
in any whole number increment up to 295 respectively that includes
at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,
18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34,
35 amino acid position(s) having a value greater than 0.5 in the
Hydrophilicity profile of FIG. 5;
[0281] (XV) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3C,
in any whole number increment up to 295 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value less than 0.5
in the Hydropathicity profile of FIG. 6;
[0282] (XVI) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3C,
in any whole number increment up to 295 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Percent Accessible Residues profile of FIG. 7;
[0283] (XVII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3C,
in any whole number increment up to 295 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Average Flexibility profile of FIG. 8;
[0284] (XVIII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, amino acids of a protein of FIG. 3C in
any whole number increment up to 295 respectively that includes at
least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15,
16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32,
33, 34, 35 amino acid position(s) having a value greater than 0.5
in the Beta-turn profile of FIG. 9;
[0285] (XIX) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3D,
in any whole number increment up to 485 respectively that includes
at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,
18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34,
35 amino acid position(s) having a value greater than 0.5 in the
Hydrophilicity profile of FIG. 5;
[0286] (XX) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3D,
in any whole number increment up to 485 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value less than 0.5
in the Hydropathicity profile of FIG. 6;
[0287] (XXI) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3D,
in any whole number increment up to 485 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Percent Accessible Residues profile of FIG. 7;
[0288] (XXII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3D,
in any whole number increment up to 485 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Average Flexibility profile of FIG. 8;
[0289] (XXIII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, amino acids of a protein of FIG. 3D in
any whole number increment up to 485 respectively that includes at
least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15,
16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32,
33, 34, 35 amino acid position(s) having a value greater than 0.5
in the Beta-turn profile of FIG. 9;
[0290] (XXIV) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3H,
in any whole number increment up to 511 respectively that includes
at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,
18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34,
35 amino acid position(s) having a value greater than 0.5 in the
Hydrophilicity profile of FIG. 5;
[0291] (XXV) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3H,
in any whole number increment up to 511 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value less than 0.5
in the Hydropathicity profile of FIG. 6;
[0292] (XXVI) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3H,
in any whole number increment up to 511 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Percent Accessible Residues profile of FIG. 7;
[0293] (XXVII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIG. 3H,
in any whole number increment up to 511 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Average Flexibility profile of FIG. 8;
[0294] (XXVIII) a polypeptide comprising at least 5, 6, 7, 8, 9,
10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26,
27, 28, 29, 30, 31, 32, 33, 34, amino acids of a protein of FIG. 3H
in any whole number increment up to 511 respectively that includes
at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Beta-turn profile of FIG. 9;
[0295] (XXIX) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIGS.
3I-J, in any whole number increment up to 137 respectively that
includes at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,
32, 33, 34, 35 amino acid position(s) having a value greater than
0.5 in the Hydrophilicity profile of FIG. 5;
[0296] (XXX) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIGS.
3I-J, in any whole number increment up to 137 respectively that
includes at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,
13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,
30, 31, 32, 33, 34, 35 amino acid position(s) having a value less
than 0.5 in the Hydropathicity profile of FIG. 6;
[0297] (XXXI) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIGS.
3I-J, in any whole number increment up to 137 respectively that
includes at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,
13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,
30, 31, 32, 33, 34, 35 amino acid position(s) having a value
greater than 0.5 in the Percent Accessible Residues profile of FIG.
7;
[0298] (XXXII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,
28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of FIGS.
3I-J, in any whole number increment up to 137 respectively that
includes at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,
13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,
30, 31, 32, 33, 34, 35 amino acid position(s) having a value
greater than 0.5 in the Average Flexibility profile of FIG. 8;
[0299] (XXXIII) a polypeptide comprising at least 5, 6, 7, 8, 9,
10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26,
27, 28, 29, 30, 31, 32, 33, 34, amino acids of a protein of FIGS.
3I-J in any whole number increment up to 137 respectively that
includes at least at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,
13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,
30, 31, 32, 33, 34, 35 amino acid position(s) having a value
greater than 0.5 in the Beta-turn profile of FIG. 9;
[0300] (XXXIV) a peptide that occurs at least twice in Tables
VIII-XXI and XXII to XLIX, collectively;
[0301] (XXXV) a peptide that occurs at least three times in Tables
VIII-XXI and XXII to XLIX, collectively;
[0302] (XXXVI) a peptide that occurs at least four times in Tables
VIII-XXI and XXII to XLIX, collectively;
[0303] (XXXVII) a peptide that occurs at least five times in Tables
VIII-XXI and XXII to XLIX, collectively;
[0304] (XXXVIII) a peptide that occurs at least once in Tables
VIII-XXI, and at least once in tables XXII to XLIX;
[0305] (XXXIX) a peptide that occurs at least once in Tables
VIII-XXI, and at least twice in tables XXII to XLIX;
[0306] (XL) a peptide that occurs at least twice in Tables
VIII-XXI, and at least once in tables XXII to XLIX;
[0307] (XLI) a peptide that occurs at least twice in Tables
VIII-XXI, and at least twice in tables XXII to XLIX;
[0308] (XLII) a peptide which comprises one two, three, four, or
five of the following characteristics, or an oligonucleotide
encoding such peptide:
[0309] i) a region of at least 5 amino acids of a particular
peptide of FIG. 3, in any whole number increment up to the full
length of that protein in FIG. 3, that includes an amino acid
position having a value equal to or greater than 0.5, 0.6, 0.7,
0.8, 0.9, or having a value equal to 1.0, in the Hydrophilicity
profile of FIG. 5;
[0310] ii) a region of at least 5 amino acids of a particular
peptide of FIG. 3, in any whole number increment up to the full
length of that protein in FIG. 3, that includes an amino acid
position having a value equal to or less than 0.5, 0.4, 0.3, 0.2,
0.1, or having a value equal to 0.0, in the Hydropathicity profile
of FIG. 6;
[0311] iii) a region of at least 5 amino acids of a particular
peptide of FIG. 3, in any whole number increment up to the full
length of that protein in FIG. 3, that includes an amino acid
position having a value equal to or greater than 0.5, 0.6, 0.7,
0.8, 0.9, or having a value equal to 1.0, in the Percent Accessible
Residues profile of FIG. 7;
[0312] iv) a region of at least 5 amino acids of a particular
peptide of FIG. 3, in any whole number increment up to the full
length of that protein in FIG. 3, that includes an amino acid
position having a value equal to or greater than 0.5, 0.6, 0.7,
0.8, 0.9, or having a value equal to 1.0, in the Average
Flexibility profile of FIG. 8; or,
[0313] v) a region of at least 5 amino acids of a particular
peptide of FIG. 3, in any whole number increment up to the full
length of that protein in FIG. 3, that includes an amino acid
position having a value equal to or greater than 0.5, 0.6, 0.7,
0.8, 0.9, or having a value equal to 1.0, in the Beta-turn profile
of FIG. 9;
[0314] (XLIII) a composition comprising a peptide of (I)-(XLII) or
an antibody or binding region thereof together with a
pharmaceutical excipient and/or in a human unit dose form.
[0315] (XLIV) a method of using a peptide of (I)-(XLII), or an
antibody or binding region thereof or a composition of (XLIII) in a
method to modulate a cell expressing 191P4D12(b),
[0316] (XLV) a method of using a peptide of (I)-(XLII) or an
antibody or binding region thereof or a composition of (XLIII) in a
method to diagnose, prophylax, prognose, or treat an individual who
bears a cell expressing 191P4D12(b)
[0317] (XLVI) a method of using a peptide of (I)-(XLII) or an
antibody or binding region thereof or a composition (XIIII) in a
method to diagnose, prophylax, prognose, or treat an individual who
bears a cell expressing 191P4D12(b), said cell from a cancer of a
tissue listed in Table I;
[0318] (XLVII) a method of using a peptide of (I)-(XLII) or an
antibody or binding region thereof or a composition of (XLIII) in a
method to diagnose, prophylax, prognose, or treat a a cancer;
[0319] (XLVIII) a method of using a peptide of (I)-(XLII) or an
antibody or binding region thereof or a composition of (XLIII) in a
method to diagnose, prophylax, prognose, or treat a a cancer of a
tissue listed in Table I; and,
[0320] (XLIX) a method of using a a peptide of (I)-(XLII) or an
antibody or binding region thereof or a composition (XLIII) in a
method to identify or characterize a modulator of a cell expressing
191P4D12(b).
[0321] As used herein, a range is understood to specifically
disclose all whole unit positions thereof.
[0322] Typical embodiments of the invention disclosed herein
include 191P4D12(b) polynucleotides that encode specific portions
of 191P4D12(b) mRNA sequences (and those which are complementary to
such sequences) such as those that encode the proteins and/or
fragments thereof, for example:
[0323] (a) 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,
19, 20, 21, 22, 23, 24, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75,
80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145,
150, 155, 160, 165, 170, 175, 180, 185, 190, 195, 200, 225, 250,
275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 505, or 510 or
more contiguous amino acids of 191P4D12(b) variant 1; the maximal
lengths relevant for other variants are: variant 2, 510 amino
acids; variant 6, 295 amino acids, variant 7, 485 amino acids,
variant 10, 510 amino acids, variant 11, 510 amino acids, variant
12, 510 amino acids, variant 13, 511 amino acids, variant 9, 137
amino acids, and variant 14, 137 amino acids.
[0324] In general, naturally occurring allelic variants of human
191P4D12(b) share a high degree of structural identity and homology
(e.g., 90% or more homology). Typically, allelic variants of a
191P4D12(b) protein contain conservative amino acid substitutions
within the 191P4D12(b) sequences described herein or contain a
substitution of an amino acid from a corresponding position in a
homologue of 191P4D12(b). One class of 191P4D12(b) allelic variants
are proteins that share a high degree of homology with at least a
small region of a particular 191P4D12(b) amino acid sequence, but
further contain a radical departure from the sequence, such as a
non-conservative substitution, truncation, insertion or frame
shift. In comparisons of protein sequences, the terms, similarity,
identity, and homology each have a distinct meaning as appreciated
in the field of genetics. Moreover, orthology and paralogy can be
important concepts describing the relationship of members of a
given protein family in one organism to the members of the same
family in other organisms.
[0325] Amino acid abbreviations are provided in Table II.
Conservative amino acid substitutions can frequently be made in a
protein without altering either the conformation or the function of
the protein. Proteins of the invention can comprise 1, 2, 3, 4, 5,
6, 7, 8, 9, 10, 11, 12, 13, 14, 15 conservative substitutions. Such
changes include substituting any of isoleucine (I), valine (V), and
leucine (L) for any other of these hydrophobic amino acids;
aspartic acid (D) for glutamic acid (E) and vice versa; glutamine
(Q) for asparagine (N) and vice versa; and serine (S) for threonine
(T) and vice versa. Other substitutions can also be considered
conservative, depending on the environment of the particular amino
acid and its role in the three-dimensional structure of the
protein. For example, glycine (G) and alanine (A) can frequently be
interchangeable, as can alanine (A) and valine (V). Methionine (M),
which is relatively hydrophobic, can frequently be interchanged
with leucine and isoleucine, and sometimes with valine. Lysine (K)
and arginine (R) are frequently interchangeable in locations in
which the significant feature of the amino acid residue is its
charge and the differing pK's of these two amino acid residues are
not significant. Still other changes can be considered
"conservative" in particular environments (see, e.g. Table III
herein; pages 13-15 "Biochemistry" 2.sup.nd ED. Lubert Stryer ed
(Stanford University); Henikoff et al., PNAS 1992 Vol 89
10915-10919; Lei et al., J Biol Chem 1995 May 19;
270(20):11882-6).
[0326] Embodiments of the invention disclosed herein include a wide
variety of art-accepted variants or analogs of 191P4D12(b) proteins
such as polypeptides having amino acid insertions, deletions and
substitutions. 191P4D12(b) variants can be made using methods known
in the art such as site-directed mutagenesis, alanine scanning, and
PCR mutagenesis. Site-directed mutagenesis (Carter et al., Nucl.
Acids Res., 13:4331 (1986); Zoller et al., Nucl. Acids Res.,
10:6487 (1987)), cassette mutagenesis (Wells et al., Gene, 34:315
(1985)), restriction selection mutagenesis (Wells et al., Philos.
Trans. R. Soc. London SerA, 317:415 (1986)) or other known
techniques can be performed on the cloned DNA to produce the
191P4D12(b) variant DNA.
[0327] Scanning amino acid analysis can also be employed to
identify one or more amino acids along a contiguous sequence that
is involved in a specific biological activity such as a
protein-protein interaction. Among the preferred scanning amino
acids are relatively small, neutral amino acids. Such amino acids
include alanine, glycine, serine, and cysteine. Alanine is
typically a preferred scanning amino acid among this group because
it eliminates the side-chain beyond the beta-carbon and is less
likely to alter the main-chain conformation of the variant. Alanine
is also typically preferred because it is the most common amino
acid. Further, it is frequently found in both buried and exposed
positions (Creighton, The Proteins, (W.H. Freeman & Co., N.Y.);
Chothia, J. Mol. Biol., 150:1 (1976)). If alanine substitution does
not yield adequate amounts of variant, an isosteric amino acid can
be used.
[0328] As defined herein, 191P4D12(b) variants, analogs or
homologs, have the distinguishing attribute of having at least one
epitope that is "cross reactive" with a 191P4D12(b) protein having
an amino acid sequence of FIG. 3. As used in this sentence, "cross
reactive" means that an antibody or T cell that specifically binds
to a 191P4D12(b) variant also specifically binds to a 191P4D12(b)
protein having an amino acid sequence set forth in FIG. 3. A
polypeptide ceases to be a variant of a protein shown in FIG. 3,
when it no longer contains any epitope capable of being recognized
by an antibody or T cell that specifically binds to the starting
191P4D12(b) protein. Those skilled in the art understand that
antibodies that recognize proteins bind to epitopes of varying
size, and a grouping of the order of about four or five amino
acids, contiguous or not, is regarded as a typical number of amino
acids in a minimal epitope. See, e.g., Nair et al., J. Immunol 2000
165(12): 6949-6955. Hebbes et al., Mol Immunol (1989) 26(9):865-73;
Schwartz et al., J Immunol (1985) 135(4);2598-608.
[0329] Other classes of 191P4D12(b)-related protein variants share
70%, 75%, 80%, 85% or 90% or more similarity with an amino acid
sequence of FIG. 3, or a fragment thereof. Another specific class
of 191P4D12(b) protein variants or analogs comprises one or more of
the 191P4D12(b) biological motifs described herein or presently
known in the art. Thus, encompassed by the present invention are
analogs of 191P4D12(b) fragments (nucleic or amino acid) that have
altered functional (e.g. immunogenic) properties relative to the
starting fragment. It is to be appreciated that motifs now or which
become part of the art are to be applied to the nucleic or amino
acid sequences of FIG. 2 or FIG. 3.
[0330] As discussed herein, embodiments of the claimed invention
include polypeptides containing less than the full amino acid
sequence of a 191P4D12(b) protein shown in FIG. 2 or FIG. 3. For
example, representative embodiments of the invention comprise
peptides/proteins having any 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15 or more contiguous amino acids of a 191P4D12(b) protein shown in
FIG. 2 or FIG. 3.
[0331] Moreover, representative embodiments of the invention
disclosed herein include polypeptides consisting of about amino
acid 1 to about amino acid 10 of a 191P4D12(b) protein shown in
FIG. 2 or FIG. 3, polypeptides consisting of about amino acid 10 to
about amino acid 20 of a 191P4D12(b) protein shown in FIG. 2 or
FIG. 3, polypeptides consisting of about amino acid 20 to about
amino acid 30 of a 191P4D12(b) protein shown in FIG. 2 or FIG. 3,
polypeptides consisting of about amino acid 30 to about amino acid
40 of a 191P4D12(b) protein shown in FIG. 2 or FIG. 3, polypeptides
consisting of about amino acid 40 to about amino acid 50 of a
191P4D12(b) protein shown in FIG. 2 or FIG. 3, polypeptides
consisting of about amino acid 50 to about amino acid 60 of a
191P4D12(b) protein shown in FIG. 2 or FIG. 3, polypeptides
consisting of about amino acid 60 to about amino acid 70 of a
191P4D12(b) protein shown in FIG. 2 or FIG. 3, polypeptides
consisting of about amino acid 70 to about amino acid 80 of a
191P4D12(b) protein shown in FIG. 2 or FIG. 3, polypeptides
consisting of about amino acid 80 to about amino acid 90 of a
191P4D12(b) protein shown in FIG. 2 or FIG. 3, polypeptides
consisting of about amino acid 90 to about amino acid 100 of a
191P4D12(b) protein shown in FIG. 2 or FIG. 3, etc. throughout the
entirety of a 191P4D12(b) amino acid sequence. Moreover,
polypeptides consisting of about amino acid 1 (or 20 or 30 or 40
etc.) to about amino acid 20, (or 130, or 140 or 150 etc.) of a
191P4D12(b) protein shown in FIG. 2 or FIG. 3 are embodiments of
the invention. It is to be appreciated that the starting and
stopping positions in this paragraph refer to the specified
position as well as that position plus or minus 5 residues.
[0332] 191P4D12(b)-related proteins are generated using standard
peptide synthesis technology or using chemical cleavage methods
well known in the art. Alternatvely, recombinant methods can be
used to generate nucleic acid molecules that encode a
191P4D12(b)-related protein. In one embodiment, nucleic acid
molecules provide a means to generate defined fragments of a
191P4D12(b) protein (or variants, homologs or analogs thereof).
[0333] III.A.) Motif-bearing Protein Embodiments
[0334] Additional illustrative embodiments of the invention
disclosed herein include 191P4D12(b) polypeptides comprising the
amino acid residues of one or more of the biological motifs
contained within a 191P4D12(b) polypeptide sequence set forth in
FIG. 2 or FIG. 3. Various motifs are known in the art, and a
protein can be evaluated for the presence of such motifs by a
number of publicly available Internet sites (see, e.g., URL
addresses: pfam.wustl.edu/; searchlauncher.bcm.tmc.edu/se-
q-search/struc-predict.html; psort.ims.u-tokyo.ac.jp/; cbs.dtu.dk/;
ebi.ac.uk/interpro/scan.html; expasy.ch/tools/scnpsit1.html;
Epimatrix.TM. and Epimer.TM., Brown University,
brown.edu/Research/TB-HIV- _Lab/epimatrix/epimatrix.html; and
BIMAS, bimas.dcrt.nih.gov/.).
[0335] Motif bearing subsequences of all 191P4D12(b) variant
proteins are set forth and identified in Tables VIII-XXI and
XXII-XLIX.
[0336] Table V sets forth several frequently occurring motifs based
on pfam searches (see URL address pfam.wustl.edu/). The columns of
Table V list (1) motif name abbreviation, (2) percent identity
found amongst the different member of the motif family, (3) motif
name or description and (4) most common function; location
information is included if the motif is relevant for location.
[0337] Polypeptides comprising one or more of the 191P4D12(b)
motifs discussed above are useful in elucidating the specific
characteristics of a malignant phenotype in view of the observation
that the 191P4D12(b) motifs discussed above are associated with
growth dysregulation and because 191P4D12(b) is overexpressed in
certain cancers (See, e.g., Table I). Casein kinase II, cAMP and
camp-dependent protein kinase, and Protein Kinase C, for example,
are enzymes known to be associated with the development of the
malignant phenotype (see e.g. Chen et al., Lab Invest., 78(2):
165-174 (1998); Gaiddon et al., Endocrinology 136(10): 4331-4338
(1995); Hall et al., Nucleic Acids Research 24(6): 1119-1126
(1996); Peterziel et al., Oncogene 18(46): 6322-6329 (1999) and
O'Brian, Oncol. Rep. 5(2): 305-309 (1998)). Moreover, both
glycosylation and myristoylation are protein modifications also
associated with cancer and cancer progression (see e.g. Dennis et
al., Biochem. Biophys. Acta 1473(1):21-34 (1999); Raju et aL, Exp.
Cell Res. 235(1): 145-154 (1997)). Amidation is another protein
modification also associated with cancer and cancer progression
(see e.g. Treston et al., J. Natl. Cancer Inst. Monogr. (13):
169-175 (1992)).
[0338] In another embodiment, proteins of the invention comprise
one or more of the immunoreactive epitopes identified in accordance
with art-accepted methods, such as the peptides set forth in Tables
VIII-XXI and XXII-XLIX. CTL epitopes can be determined using
specific algorithms to identify peptides within a 191P4D12(b)
protein that are capable of optimally binding to specified HLA
alleles (e.g., Table IV; Epimatrix.TM. and Epimer.TM., Brown
University, URL brown.edu/Research/TB-HIV_Lab/epima-
trix/epimatrix.html; and BIMAS, URL bimas.dcrt.nih.gov/.) Moreover,
processes for identifying peptides that have sufficient binding
affinity for HLA molecules and which are correlated with being
immunogenic epitopes, are well known in the art, and are carried
out without undue experimentation. In addition, processes for
identifying peptides that are immunogenic epitopes, are well known
in the art, and are carried out without undue experimentation
either in vitro or in vivo.
[0339] Also known in the art are principles for creating analogs of
such epitopes in order to modulate immunogenicity. For example, one
begins with an epitope that bears a CTL or HTL motif (see, e.g.,
the HLA Class I and HLA Class II motifs/supermotifs of Table IV).
The epitope is analoged by substituting out an amino acid at one of
the specified positions, and replacing it with another amino acid
specified for that position. For example, on the basis of residues
defined in Table IV, one can substitute out a deleterious residue
in favor of any other residue, such as a preferred residue;
substitute a less-preferred residue with a preferred residue; or
substitute an originally-occurring preferred residue with another
preferred residue. Substitutions can occur at primary anchor
positions or at other positions in a peptide; see, e.g., Table
IV.
[0340] A variety of references reflect the art regarding the
identification and generation of epitopes in a protein of interest
as well as analogs thereof. See, for example, WO 97/33602 to
Chesnut et al.; Sette, Immunogenetics 1999 50(3-4): 201-212; Sette
et al., J. Immunol. 2001 166(2): 1389-1397; Sidney et al., Hum.
Immunol. 1997 58(1): 12-20; Kondo et al., Immunogenetics 1997
45(4): 249-258; Sidney et al., J. Immunol. 1996 157(8): 3480-90;
and Falk et al., Nature 351: 290-6 (1991); Hunt et al., Science
255:1261-3 (1992); Parker et al., J. Immunol. 149:3580-7 (1992);
Parker et al., J. Immunol. 152:163-75 (1994)); Kast et al., 1994
152(8): 3904-12; Borras-Cuesta et al., Hum. Immunol. 2000 61(3):
266-278; Alexander et al., J. Immunol. 2000 164(3); 164(3):
1625-1633; Alexander et al., PMID: 7895164, UI: 95202582;
O'Sullivan et al., J. Immunol. 1991 147(8): 2663-2669; Alexander et
al., Immunity 1994 1(9): 751-761 and Alexander et al., Immunol.
Res. 1998 18(2): 79-92.
[0341] Related embodiments of the invention include polypeptides
comprising combinations of the different motifs set forth in Table
VI, and/or, one or more of the predicted CTL epitopes of Tables
VIII-XXI and XXII-XLIX, and/or, one or more of the predicted HTL
epitopes of Tables XLVI-XLIX, and/or, one or more of the T cell
binding motifs known in the art. Preferred embodiments contain no
insertions, deletions or substitutions either within the motifs or
within the intervening sequences of the polypeptides. In addition,
embodiments which include a number of either N-terminal and/or
C-terminal amino acid residues on either side of these motifs may
be desirable (to, for example, include a greater portion of the
polypeptide architecture in which the motif is located). Typically,
the number of N-terminal and/or C-terminal amino acid residues on
either side of a motif is between about 1 to about 100 amino acid
residues, preferably 5 to about 50 amino acid residues.
[0342] 191P4D12(b)-related proteins are embodied in many forms,
preferably in isolated form. A purified 191P4D12(b) protein
molecule will be substantially free of other proteins or molecules
that impair the binding of 191P4D12(b) to antibody, T cell or other
ligand. The nature and degree of isolation and purification will
depend on the intended use. Embodiments of a 191P4D12(b)-related
proteins include purified 191P4D12(b)-related proteins and
functional, soluble 191P4D12(b)-related proteins. In one
embodiment, a functional, soluble 191P4D12(b) protein or fragment
thereof retains the ability to be bound by antibody, T cell or
other ligand.
[0343] The invention also provides 191P4D12(b) proteins comprising
biologically active fragments of a 191P4D12(b) amino acid sequence
shown in FIG. 2 or FIG. 3. Such proteins exhibit properties of the
starting 191P4D12(b) protein, such as the ability to elicit the
generation of antibodies that specifically bind an epitope
associated with the starting 191P4D12(b) protein; to be bound by
such antibodies; to elicit the activation of HTL or CTL; and/or, to
be recognized by HTL or CTL that also specifically bind to the
starting protein.
[0344] 191P4D12(b)-related polypeptides that contain particularly
interesting structures can be predicted and/or identified using
various analytical techniques well known in the art, including, for
example, the methods of Chou-Fasman, Garnier-Robson,
Kyte-Doolittle, Eisenberg, Karplus-Schultz or Jameson-Wolf
analysis, or based on immunogenicity. Fragments that contain such
structures are particularly useful in generating subunit-specific
anti-191P4D12(b) antibodies or T cells or in identifying cellular
factors that bind to 191P4D12(b). For example, hydrophilicity
profiles can be generated, and immunogenic peptide fragments
identified, using the method of Hopp, T. P. and Woods, K. R., 1981,
Proc. Natl. Acad. Sci. U.S.A. 78:3824-3828. Hydropathicity profiles
can be generated, and immunogenic peptide fragments identified,
using the method of Kyte, J. and Doolittle, R. F., 1982, J. Mol.
Biol. 157:105-132. Percent (%) Accessible Residues profiles can be
generated, and immunogenic peptide fragments identified, using the
method of Janin J., 1979, Nature 277:491-492. Average Flexibility
profiles can be generated, and immunogenic peptide fragments
identified, using the method of Bhaskaran R., Ponnuswamy P. K.,
1988, Int. J. Pept. Protein Res. 32:242-255. Beta-turn profiles can
be generated, and immunogenic peptide fragments identified, using
the method of Deleage, G., Roux B., 1987, Protein Engineering
1:289-294.
[0345] CTL epitopes can be determined using specific algorithms to
identfy peptides within a 191P4D12(b) protein that are capable of
optimally binding to specified HLA alleles (e.g., by using the
SYFPEITHI site at World Wide Web URL syfpeithi.bmi-heidelberg.com/;
the listings in Table IV(A)-(E); Epimatrix.TM. and Epimer.TM.,
Brown University, URL
(brown.edu/Research/TB-HIV_Lab/epimatrix/epimatrix.html); and
BIMAS, URL bimas.dcrt.nih.gov/). Illustrating this, peptide
epitopes from 191P4D12(b) that are presented in the context of
human MHC Class I molecules, e.g., HLA-A1, A2, A3, A11, A24, B7 and
B35 were predicted (see, e.g., Tables VIII-XXI, XXII-XLIX).
Specifically, the complete amino acid sequence of the 191P4D12(b)
protein and relevant portions of other variants, i.e., for HLA
Class I predictions 9 flanking residues on either side of a point
mutation or exon juction, and for HLA Class II predictions 14
flanking residues on either side of appoint mutation or exon
junction corresponding to that variant, were entered into the HLA
Peptide Motif Search algorithm found in the Bioinformatics and
Molecular Analysis Section (BIMAS) web site listed above; in
addition to the site SYFPEITHI, at URL
syfpeithi.bmi-heidelberg.com/.
[0346] The HLA peptide motif search algorithm was developed by Dr.
Ken Parker based on binding of specific peptide sequences in the
groove of HLA Class I molecules, in particular HLA-A2 (see, e.g.,
Falk et al., Nature 351: 290-6 (1991); Hunt et al., Science
255:1261-3 (1992); Parker et al., J. Immunol. 149:3580-7 (1992);
Parker et al., J. Immunol. 152:163-75 (1994)). This algorithm
allows location and ranking of 8-mer, 9-mer, and 10-mer peptides
from a complete protein sequence for predicted binding to HLA-A2 as
well as numerous other HLA Class I molecules. Many HLA class I
binding peptides are 8-, 9-, 10 or 11-mers. For example, for Class
I HLA-A2, the epitopes preferably contain a leucine (L) or
methionine (M) at position 2 and a valine (V) or leucine (L) at the
C-terminus (see, e.g., Parker et al., J. Immunol. 149:3580-7
(1992)). Selected results of 191P4D12(b) predicted binding peptides
are shown in Tables VIII-XXI and XXII-XLIX herein. In Tables
VIII-XXI and XXII-XLVII, selected candidates, 9-mers and 10-mers,
for each family member are shown along with their location, the
amino acid sequence of each specific peptide, and an estimated
binding score. In Tables XLVI-XLIX, selected candidates, 15-mers,
for each family member are shown along with their location, the
amino acid sequence of each specific peptide, and an estimated
binding score. The binding score corresponds to the estimated half
time of dissociation of complexes containing the peptide at
37.degree. C. at pH 6.5. Peptides with the highest binding score
are predicted to be the most tightly bound to HLA Class I on the
cell surface for the greatest period of time and thus represent the
best immunogenic targets for T-cell recognition.
[0347] Actual binding of peptides to an HLA allele can be evaluated
by stabilization of HLA expression on the antigen-processing
defective cell line T2 (see, e.g., Xue et al., Prostate 30:73-8
(1997) and Peshwa et aL, Prostate 36:129-38 (1998)). Immunogenicity
of specific peptides can be evaluated in vitro by stimulation of
CD8+ cytotoxic T lymphocytes (CTL) in the presence of antigen
presenting cells such as dendritic cells.
[0348] It is to be appreciated that every epitope predicted by the
BIMAS site, Epimer.TM. and Epimatrix.TM. sites, or specified by the
HLA class I or class II motifs available in the art or which become
part of the art such as set forth in Table IV (or determined using
World Wide Web site URL syfpeithi.bmi-heidelberg.com/, or BIMAS,
bimas.dcrt.nih.gov/) are to be "applied" to a 191P4D12(b) protein
in accordance with the invention. As used in this context "applied"
means that a 191P4D12(b) protein is evaluated, e.g., visually or by
computer-based patterns finding methods, as appreciated by those of
skill in the relevant art. Every subsequence of a 191P4D12(b)
protein of 8, 9, 10, or 11 amino acid residues that bears an HLA
Class I motif, or a subsequence of 9 or more amino acid residues
that bear an HLA Class II motif are within the scope of the
invention.
[0349] III.B.) Expression of 191P4D12(b)-related Proteins
[0350] In an embodiment described in the examples that follow,
191P4D12(b) can be conveniently expressed in cells (such as 293T
cells) transfected with a commercially available expression vector
such as a CMV-driven expression vector encoding 191P4D12(b) with a
C-terminal 6XHis and MYC tag (pcDNA3.1/mycHIS, Invitrogen or Tag5,
GenHunter Corporation, Nashville Tenn.). The Tag5 vector provides
an IgGK secretion signal that can be used to facilitate the
production of a secreted 191P4D12(b) protein in transfected cells.
The secreted HIS-tagged 191P4D12(b) in the culture media can be
purified, e.g., using a nickel column using standard
techniques.
[0351] III.C.) Modifications of 191P4D12(b)-related Proteins
[0352] Modifications of 191P4D12(b)-related proteins such as
covalent modifications are included within the scope of this
invention. One type of covalent modification includes reacting
targeted amino acid residues of a 191P4D12(b) polypeptide with an
organic derivatizing agent that is capable of reacting with
selected side chains or the N- or C- terminal residues of a
191P4D12(b) protein. Another type of covalent modification of a
191P4D12(b) polypeptide included within the scope of this invention
comprises altering the native glycosylation pattern of a protein of
the invention. Another type of covalent modification of 191P4D12(b)
comprises linking a 191P4D12(b) polypeptide to one of a variety of
nonproteinaceous polymers, e.g., polyethylene glycol (PEG),
polypropylene glycol, or polyoxyalkylenes, in the manner set forth
in U.S. Pat. Nos. 4,640,835; 4,496,689; 4,301,144; 4,670,417;
4,791,192 or 4,179,337.
[0353] The 191P4D12(b)-related proteins of the present invention
can also be modified to form a chimeric molecule comprising
191P4D12(b) fused to another, heterologous polypeptide or amino
acid sequence. Such a chimeric molecule can be synthesized
chemically or recombinantly. A chimeric molecule can have a protein
of the invention fused to another tumor-associated antigen or
fragment thereof. Alternatively, a protein in accordance with the
invention can comprise a fusion of fragments of a 191P4D12(b)
sequence (amino or nucleic acid) such that a molecule is created
that is not, through its length, directly homologous to the amino
or nucleic acid sequences shown in FIG. 2 or FIG. 3. Such a
chimeric molecule can comprise multiples of the same subsequence of
191P4D12(b). A chimeric molecule can comprise a fusion of a
191P4D12(b)-related protein with a polyhistidine epitope tag, which
provides an epitope to which immobilized nickel can selectively
bind, with cytokines or with growth factors. The epitope tag is
generally placed at the amino- or carboxyl-terminus of a
191P4D12(b) protein. In an alternative embodiment, the chimeric
molecule can comprise a fusion of a 191P4D12(b)-related protein
with an immunoglobulin or a particular region of an immunoglobulin.
For a bivalent form of the chimeric molecule (also referred to as
an "immunoadhesin"), such a fusion could be to the Fc region of an
IgG molecule. The Ig fusions preferably include the substitution of
a soluble (transmembrane domain deleted or inactivated) form of a
191P4D12(b) polypeptide in place of at least one variable region
within an Ig molecule. In a preferred embodiment, the
immunoglobulin fusion includes the hinge, CH2 and CH3, or the
hinge, CHI, CH2 and CH3 regions of an IgGl molecule. For the
production of immunoglobulin fusions see, e.g., U.S. Pat. No.
5,428,130 issued Jun. 27, 1995.
[0354] III.D.) Uses of 191P4D12(b)-related Proteins
[0355] The proteins of the invention have a number of different
specific uses. As 191P4D12(b) is highly expressed in prostate and
other cancers, 191P4D12(b)-related proteins are used in methods
that assess the status of 191P4D12(b) gene products in normal
versus cancerous tissues, thereby elucidating the malignant
phenotype. Typically, polypeptides from specific regions of a
191P4D12(b) protein are used to assess the presence of
perturbations (such as deletions, insertions, point mutations etc.)
in those regions (such as regions containing one or more motifs).
Exemplary assays utilize antibodies or T cells targeting
191P4D12(b)-related proteins comprising the amino acid residues of
one or more of the biological motifs contained within a 191P4D12(b)
polypeptide sequence in order to evaluate the characteristics of
this region in normal versus cancerous tissues or to elicit an
immune response to the epitope. Alternatively, 191P4D12(b)-related
proteins that contain the amino acid residues of one or more of the
biological motifs in a 191P4D12(b) protein are used to screen for
factors that interact with that region of 191P4D12(b).
[0356] 191P4D12(b) protein fragments/subsequences are particularly
useful in generating and characterizing domain-specific antibodies
(e.g., antibodies recognizing an extracellular or intracellular
epitope of a 191P4D12(b) protein), for identifying agents or
cellular factors that bind to 191P4D12(b) or a particular
structural domain thereof, and in various therapeutic and
diagnostic contexts, including but not limited to diagnostic
assays, cancer vaccines and methods of preparing such vaccines.
[0357] Proteins encoded by the 191P4D12(b) genes, or by analogs,
homologs or fragments thereof, have a variety of uses, including
but not limited to generating antibodies and in methods for
identifying ligands and other agents and cellular constituents that
bind to a 191P4D12(b) gene product. Antibodies raised against a
191P4D12(b) protein or fragment thereof are useful in diagnostic
and prognostic assays, and imaging methodologies in the management
of human cancers characterized by expression of 191P4D12(b)
protein, such as those listed in Table I. Such antibodies can be
expressed intracellularly and used in methods of treating patients
with such cancers. 191P4D12(b)-related nucleic acids or proteins
are also used in generating HTL or CTL responses.
[0358] Various immunological assays useful for the detection of
191P4D12(b) proteins are used, including but not limited to various
types of radioimmunoassays, enzyme-linked immunosorbent assays
(ELISA), enzyme-linked immunofluorescent assays (ELIFA),
immunocytochemical methods, and the like. Antibodies can be labeled
and used as immunological imaging reagents capable of detecting
191P4D12(b)-expressing cells (e.g., in radioscintigraphic imaging
methods). 191P4D12(b) proteins are also particularly useful in
generating cancer vaccines, as further described herein.
[0359] IV.) 191P4D12(b) Antibodies
[0360] Another aspect of the invention provides antibodies that
bind to 191P4D12(b)-related proteins. Preferred antibodies
specifically bind to a 191P4D12(b)-related protein and do not bind
(or bind weakly) to peptides or proteins that are not
191P4D12(b)-related proteins under physiological conditions. In
this context, examples of physiological conditions include: 1)
phosphate buffered saline; 2) Tris-buffered saline containing 25 mM
Tris and 150 mM NaCl; or normal saline (0.9% NaCl); 4) animal serum
such as human serum; or, 5) a combination of any of 1) through 4);
these reactions preferably taking place at pH 7.5, alternatively in
a range of pH 7.0 to 8.0, or alternatively in a range of pH 6.5 to
8.5; also, these reactions taking place at a temperature between
4.degree. C. to 37.degree. C. For example, antibodies that bind
191P4D12(b) can bind 191P4D12(b)-related proteins such as the
homologs or analogs thereof.
[0361] 191P4D12(b) antibodies of the invention are particularly
useful in cancer (see, e.g., Table I) diagnostic and prognostic
assays, and imaging methodologies. Similarly, such antibodies are
useful in the treatment, diagnosis, and/or prognosis of other
cancers, to the extent 191P4D12(b) is also expressed or
overexpressed in these other cancers. Moreover, intracellularly
expressed antibodies (e.g., single chain antibodies) are
therapeutically useful in treating cancers in which the expression
of 191P4D12(b) is involved, such as advanced or metastatic prostate
cancers.
[0362] The invention also provides various immunological assays
useful for the detection and quantification of 191P4D12(b) and
mutant 191P4D12(b)-related proteins. Such assays can comprise one
or more 191P4D12(b) antibodies capable of recognizing and binding a
191P4D12(b)-related protein, as appropriate. These assays are
performed within various immunological assay formats well known in
the art, including but not limited to various types of
radioimmunoassays, enzyme-linked immunosorbent assays (ELISA),
enzyme-linked immunofluorescent assays (ELIFA), and the like.
[0363] Immunological non-antibody assays of the invention also
comprise T cell immunogenicity assays (inhibitory or stimulatory)
as well as major histocompatibility complex (MHC) binding
assays.
[0364] In addition, immunological imaging methods capable of
detecting prostate cancer and other cancers expressing 191P4D12(b)
are also provided by the invention, including but not limited to
radioscintigraphic imaging methods using labeled 191P4D12(b)
antibodies. Such assays are clinically useful in the detection,
monitoring, and prognosis of 191P4D12(b) expressing cancers such as
prostate cancer.
[0365] 191P4D12(b) antibodies are also used in methods for
purifying a 191P4D12(b)-related protein and for isolating
191P4D12(b) homologues and related molecules. For example, a method
of purifying a 191P4D12(b)-related protein comprises incubating a
191P4D12(b) antibody, which has been coupled to a solid matrix,
with a lysate or other solution containing a 191P4D12(b)-related
protein under conditions that permit the 191P4D12(b) antibody to
bind to the 191P4D12(b)-related protein; washing the solid matrix
to eliminate impurities; and eluting the 191P4D12(b)-related
protein from the coupled antibody. Other uses of 191P4D12(b)
antibodies in accordance with the invention include generating
anti-idiotypic antibodies that mimic a 191P4D12(b) protein.
[0366] Various methods for the preparation of antibodies are well
known in the art. For example, antibodies can be prepared by
immunizing a suitable mammalian host using a 191P4D12(b)-related
protein, peptide, or fragment, in isolated or immunoconjugated form
(Antibodies: A Laboratory Manual, CSH Press, Eds., Harlow, and Lane
(1988); Harlow, Antibodies, Cold Spring Harbor Press, NY (1989)).
In addition, fusion proteins of 191P4D12(b) can also be used, such
as a 191P4D12(b) GST-fusion protein. In a particular embodiment, a
GST fusion protein comprising all or most of the amino acid
sequence of FIG. 2 or FIG. 3 is produced, then used as an immunogen
to generate appropriate antibodies. In another embodiment, a
191P4D12(b)-related protein is synthesized and used as an
immunogen.
[0367] In addition, naked DNA immunization techniques known in the
art are used (with or without purified 191P4D12(b)-related protein
or 191P4D12(b) expressing cells) to generate an immune response to
the encoded immunogen (for review, see Donnelly et al., 1997, Ann.
Rev. Immunol. 15: 617-648).
[0368] The amino acid sequence of a 191P4D12(b) protein as shown in
FIG. 2 or FIG. 3 can be analyzed to select specific regions of the
191P4D12(b) protein for generating antibodies. For example,
hydrophobicity and hydrophilicity analyses of a 191P4D12(b) amino
acid sequence are used to identify hydrophilic regions in the
191P4D12(b) structure. Regions of a 191P4D12(b) protein that show
immunogenic structure, as well as other regions and domains, can
readily be identified using various other methods known in the art,
such as Chou-Fasman, Garnier-Robson, Kyte-Doolittle, Eisenberg,
Karplus-Schultz or Jameson-Wolf analysis. Hydrophilicity profiles
can be generated using the method of Hopp, T. P. and Woods, K. R.,
1981, Proc. Natl. Acad. Sci. U.S.A. 78:3824-3828. Hydropathicity
profiles can be generated using the method of Kyte, J. and
Doolittle, R. F., 1982, J. Mol. Biol. 157:105-132. Percent (%)
Accessible Residues profiles can be generated using the method of
Janin J., 1979, Nature 277:491-492. Average Flexibility profiles
can be generated using the method of Bhaskaran R., Ponnuswamy P.
K., 1988, Int. J. Pept. Protein Res. 32:242-255. Beta-turn profiles
can be generated using the method of Deleage, G., Roux B., 1987,
Protein Engineering 1:289-294. Thus, each region identified by any
of these programs or methods is within the scope of the present
invention. Methods for the generation of 191P4D12(b) antibodies are
further illustrated by way of the examples provided herein. Methods
for preparing a protein or polypeptide for use as an immunogen are
well known in the art. Also well known in the art are methods for
preparing immunogenic conjugates of a protein with a carrier, such
as BSA, KLH or other carrier protein. In some circumstances, direct
conjugation using, for example, carbodiimide reagents are used; in
other instances linking reagents such as those supplied by Pierce
Chemical Co., Rockford, Ill., are effective. Administration of a
191P4D12(b) immunogen is often conducted by injection over a
suitable time period and with use of a suitable adjuvant, as is
understood in the art. During the immunization schedule, titers of
antibodies can be taken to determine adequacy of antibody
formation.
[0369] 191P4D12(b) monoclonal antbodies can be produced by various
means well known in the art. For example, immortalized cell lines
that secrete a desired monoclonal antibody are prepared using the
standard hybridoma technology of Kohler and Milstein or
modificabons that immortalize antibody-producing B cells, as is
generally known. Immortalized cell lines that secrete the desired
antibodies are screened by immunoassay in which the antigen is a
191P4D12(b)-related protein. When the appropriate immortalized cell
culture is identified, the cells can be expanded and antibodies
produced either from in vitro cultures or from ascites fluid.
[0370] The antibodies or fragments of the invention can also be
produced, by recombinant means. Regions that bind specifically to
the desired regions of a 191P4D12(b) protein can also be produced
in the context of chimeric or complementarity-determining region
(CDR) grafted antibodies of multiple species origin. Humanized or
human 191P4D12(b) antibodies can also be produced, and are
preferred for use in therapeutic contexts. Methods for humanizing
murine and other non-human antibodies, by substituting one or more
of the non-human antibody CDRs for corresponding human antibody
sequences, are well known (see for example, Jones et al., 1986,
Nature 321: 522-525; Riechmann et al., 1988, Nature 332: 323-327;
Verhoeyen et al., 1988, Science 239: 1534-1536). See also, Carter
et al., 1993, Proc. Natl. Acad. Sci. USA 89: 4285 and Sims et al,
1993, J. Immunol. 151: 2296.
[0371] Methods for producing fully human monoclonal antibodies
include phage display and transgenic methods (for review, see
Vaughan et al., 1998, Nature Biotechnology 16: 535-539). Fully
human 191P4D12(b) monoclonal antibodies can be generated using
cloning technologies employing large human Ig gene combinatorial
libraries (i.e., phage display) (Griffiths and Hoogenboom, Building
an in vitro immune system: human antibodies from phage display
libraries. In: Protein Engineering of Antibody Molecules for
Prophylactic and Therapeutic Applications in Man, Clark, M. (Ed.),
Nottingham Academic, pp 45-64 (1993); Burton and Barbas, Human
Antibodies from combinatorial libraries. Id., pp 65-82). Fully
human 191P4D12(b) monoclonal antibodies can also be produced using
transgenic mice engineered to contain human immunoglobulin gene
loci as described in PCT Patent Application WO98/24893,
Kucherlapati and Jakobovits et al., published Dec. 3, 1997 (see
also, Jakobovits, 1998, Exp. Opin. Invest. Drugs 7(4): 607-614;
U.S. Pat. Nos. 6,162,963 issued 19 Dec. 2000; 6,150,584 issued 12
Nov. 2000; and, 6,114,598 issued 5 Sep. 2000). This method avoids
the in vitro manipulation required with phage display technology
and efficiently produces high affinity authentic human
antibodies.
[0372] Reactivity of 191P4D12(b) antibodies with a
191P4D12(b)-related protein can be established by a number of well
known means, including Western blot, immunoprecipitation, ELISA,
and FACS analyses using, as appropriate, 191P4D12(b)-related
proteins, 191P4D12(b)-expressing cells or extracts thereof. A
191P4D12(b) antibody or fragment thereof can be labeled with a
detectable marker or conjugated to a second molecule. Suitable
detectable markers include, but are not limited to, a radioisotope,
a fluorescent compound, a bioluminescent compound, chemiluminescent
compound, a metal chelator or an enzyme. Further, bi-specific
antibodies specific for two or more 191P4D12(b) epitopes are
generated using methods generally known in the art. Homodimeric
antibodies can also be generated by cross-linking techniques known
in the art (e.g., Wolff et al., Cancer Res. 53: 2560-2565).
[0373] V.) 191P4D12(b) Cellular Immune Responses
[0374] The mechanism by which T cells recognize antigens has been
delineated. Efficacious peptide epitope vaccine compositions of the
invention induce a therapeutic or prophylactic immune responses in
very broad segments of the world-wide population. For an
understanding of the value and efficacy of compositions of the
invention that induce cellular immune responses, a brief review of
immunology-related technology is provided.
[0375] A complex of an HLA molecule and a peptidic antigen acts as
the ligand recognized by HLA-restricted T cells (Buus, S. et al.,
Cell 47:1071, 1986; Babbitt, B. P. et al., Nature 317:359, 1985;
Townsend, A. and Bodmer, H., Annu. Rev. Immunol. 7:601, 1989;
Germain, R. N., Annu. Rev. Immunol. 11:403, 1993). Through the
study of single amino acid substituted antigen analogs and the
sequencing of endogenously bound, naturally processed peptides,
critical residues that correspond to motifs required for specific
binding to HLA antigen molecules have been identified and are set
forth in Table IV (see also, e.g., Southwood, et al., J. Immunol.
160:3363, 1998; Rammensee, et al., Immunogenetics 41:178, 1995;
Rammensee et al., SYFPEITHI, access via World Wide Web at URL
(134.2.96.221/scripts.hlaserver.dll/home.htm); Sette, A. and
Sidney, J. Curr. Opin. Immunol. 10:478,1998; Engelhard, V. H.,
Curr. Opin. Immunol. 6:13, 1994; Sette, A. and Grey, H. M., Curr.
Opin. Immunol. 4:79, 1992; Sinigaglia, F. and Hammer, J. Curr.
Biol. 6:52, 1994; Ruppert et al., Cell 74:929-937, 1993; Kondo et
al., J. Immunol. 155:4307-4312, 1995; Sidney et al., J. Immunol.
157:3480-3490, 1996; Sidney et al., Human Immunol. 45:79-93, 1996;
Sette, A. and Sidney, J. Immunogenetics 1999 Nov; 50(3-4):201-12,
Review).
[0376] Furthermore, x-ray crystallographic analyses of HLA-peptide
complexes have revealed pockets within the peptide binding
cleft/groove of HLA molecules which accommodate, in an
allele-specific mode, residues borne by peptide ligands; these
residues in turn determine the HLA binding capacity of the peptides
in which they are present. (See, e.g., Madden, D. R. Annu. Rev.
Immunol. 13:587, 1995; Smith, et al., Immunity 4:203, 1996; Fremont
et al., Immunity 8:305, 1998; Stern et al., Structure 2:245, 1994;
Jones, E. Y. Curr. Opin. Immunol. 9:75, 1997; Brown, J. H. et al.,
Nature 364:33, 1993; Guo, H. C. et al., Proc. Natl. Acad. Sci. USA
90:8053, 1993; Guo, H. C. et al., Nature 360:364, 1992; Silver, M.
L. et al., Nature 360:367, 1992; Matsumura, M. et al., Science
257:927, 1992; Madden et al., Cell 70:1035, 1992; Fremont, D. H. et
al., Science 257:919,1992; Saper, M. A., Bjorkman, P. J. and Wiley,
D. C., J. Mol. Biol. 219:277,1991.)
[0377] Accordingly, the definition of class I and class II
allele-specific HLA binding motifs, or class I or class II
supermotifs allows identification of regions within a protein that
are correlated with binding to particular HLA antigen(s).
[0378] Thus, by a process of HLA motif identification, candidates
for epitope-based vaccines have been identified; such candidates
can be further evaluated by HLA-peptide binding assays to determine
binding affinity and/or the time period of association of the
epitope and its corresponding HLA molecule. Additional confirmatory
work can be performed to select, amongst these vaccine candidates,
epitopes with preferred characteristics in terms of population
coverage, and/or immunogenicity.
[0379] Various strategies can be utilized to evaluate cellular
immunogenicity, including:
[0380] 1) Evaluation of primary T cell cultures from normal
individuals (see, e.g., Wentworth, P. A. et al., Mol. Immunol.
32:603, 1995; Celis, E. et al., Proc. Natl. Acad. Sci. USA 91:2105,
1994; Tsai, V. et al., J. Immunol. 158:1796, 1997; Kawashima, I. et
al., Human Immunol. 59:1, 1998). This procedure involves the
stimulation of peripheral blood lymphocytes (PBL) from normal
subjects with a test peptide in the presence of antigen presenting
cells in vitro over a period of several weeks. T cells specific for
the peptide become activated during this time and are detected
using, e.g., a lymphokine- or .sup.51Cr-release assay involving
peptide sensitized target cells.
[0381] 2) Immunization of HLA transgenic mice (see, e.g.,
Wentworth, P. A. et al., J. Immunol. 26:97, 1996; Wentworth, P. A.
et al., Int. Immunol. 8:651, 1996; Alexander, J. et al., J.
Immunol. 159:4753, 19,97). For example, in such methods peptides in
incomplete Freund's adjuvant are administered subcutaneously to HLA
transgenic mice. Several weeks following immunization, splenocytes
are removed and cultured in vitro in the presence of test peptide
for approximately one week. Peptide-specific T cells are detected
using, e.g., a .sup.51Cr-release assay involving peptide sensitized
target cells and target cells expressing endogenously generated
antigen.
[0382] 3) Demonstration of recall T cell responses from immune
individuals who have been either effectively vaccinated and/or from
chronically ill patients (see, e.g., Rehermann, B. et al., J. Exp.
Med. 181:1047, 1995; Doolan, D. L. et al., Immunity 7:97, 1997;
Bertoni, R. et al., J. Clin. Invest. 100:503, 1997; Threlkeld, S.
C. et al., J. Immunol. 159:1648, 1997; Diepolder, H. M. et al., J.
Virol. 71:6011, 1997). Accordingly, recall responses are detected
by culturing PBL from subjects that have been exposed to the
antigen due to disease and thus have generated an immune response
"naturally", or from patients who were vaccinated against the
antigen. PBL from subjects are cultured in vitro for 1-2 weeks in
the presence of test peptide plus antigen presenting cells (APC) to
allow activation of "memory" T cells, as compared to "naive" T
cells. At the end of the culture period, T cell activity is
detected using assays including .sup.51Cr release involving
peptide-sensitized targets, T cell proliferation, or lymphokine
release.
[0383] VI.) 191P4D12(b) Transgenic Animals
[0384] Nucleic acids that encode a 191P4D12(b)-related protein can
also be used to generate either transgenic animals or "knock out"
animals that, in turn, are useful in the development and screening
of therapeutically useful reagents. In accordance with established
techniques, cDNA encoding 191P4D12(b) can be used to clone genomic
DNA that encodes 191P4D12(b). The cloned genomic sequences can then
be used to generate transgenic animals containing cells that
express DNA that encode 191P4D12(b). Methods for generating
transgenic animals, particularly animals such as mice or rats, have
become conventional in the art and are described, for example, in
U.S. Pat. Nos. 4,736,866 issued 12 Apr. 1988, and 4,870,009 issued
26 Sep. 1989. Typically, particular cells would be targeted for
191P4D12(b) transgene incorporation with tissue-specific
enhancers.
[0385] Transgenic animals that include a copy of a transgene
encoding 191P4D12(b) can be used to examine the effect of increased
expression of DNA that encodes 191P4D12(b). Such animals can be
used as tester animals for reagents thought to confer protection
from, for example, pathological conditions associated with its
overexpression. In accordance with this aspect of the invention, an
animal is treated with a reagent and a reduced incidence of a
pathological condition, compared to untreated animals that bear the
transgene, would indicate a potential therapeutic intervention for
the pathological condition.
[0386] Alternatively, non-human homologues of 191P4D12(b) can be
used to construct a 191P4D12(b) "knock out" animal that has a
defective or altered gene encoding 191P4D12(b) as a result of
homologous recombination between the endogenous gene encoding
191P4D12(b) and altered genomic DNA encoding 191P4D12(b) introduced
into an embryonic cell of the animal. For example, cDNA that
encodes 191P4D12(b) can be used to clone genomic DNA encoding
191P4D12(b) in accordance with established techniques. A portion of
the genomic DNA encoding 191P4D12(b) can be deleted or replaced
with another gene, such as a gene encoding a selectable marker that
can be used to monitor integration. Typically, several kilobases of
unaltered flanking DNA (both at the 5' and 3' ends) are included in
the vector (see, e.g., Thomas and Capecchi, Cell, 51:503 (1987) for
a description of homologous recombination vectors). The vector is
introduced into an embryonic stem cell line (e.g., by
electroporation) and cells in which the introduced DNA has
homologously recombined with the endogenous DNA are selected (see,
e.g., Li et al., Cell, 69:915 (1992)). The selected cells are then
injected into a blastocyst of an animal (e.g., a mouse or rat) to
form aggregation chimeras (see, e.g., Bradley, in Teratocarcinomas
and Embryonic Stem Cells: A Practical Approach, E. J. Robertson,
ed. (IRL, Oxford, 1987), pp. 113-152). A chimeric embryo can then
be implanted into a suitable pseudopregnant female foster animal,
and the embryo brought to term to create a "knock out" animal.
Progeny harboring the homologously recombined DNA in their germ
cells can be identified by standard techniques and used to breed
animals in which all cells of the animal contain the homologously
recombined DNA. Knock out animals can be characterized, for
example, for their ability to defend against certain pathological
conditions or for their development of pathological conditions due
to absence of a 191P4D12(b) polypeptide.
[0387] VII.) Methods for the Detection of 191P4D12(b)
[0388] Another aspect of the present invention relates to methods
for detecting 191P4D12(b) polynucleotides and 191P4D12(b)-related
proteins, as well as methods for identifying a cell that expresses
191P4D12(b). The expression profile of 191P4D12(b) makes it a
diagnostic marker for metastasized disease. Accordingly, the status
of 191P4D12(b) gene products provides information useful for
predicting a variety of factors including susceptibility to
advanced stage disease, rate of progression, and/or tumor
aggressiveness. As discussed in detail herein, the status of
191P4D12(b) gene products in patient samples can be analyzed by a
variety protocols that are well known in the art including
immunohistochemical analysis, the variety of Northern blotting
techniques including in situ hybridization, RT-PCR analysis (for
example on laser capture micro-dissected samples), Western blot
analysis and tissue array analysis.
[0389] More particularly, the invention provides assays for the
detection of 191P4D12(b) polynucleotides in a biological sample,
such as serum, bone, prostate, and other tissues, urine, semen,
cell preparations, and the like. Detectable 191P4D12(b)
polynucleotides include, for example, a 191P4D12(b) gene or
fragment thereof, 191P4D12(b) mRNA, alternative splice variant
191P4D12(b) mRNAs, and recombinant DNA or RNA molecules that
contain a 191P4D12(b) polynucleotide. A number of methods for
amplifying and/or detecting the presence of 191P4D12(b)
polynucleotides are well known in the art and can be employed in
the practice of this aspect of the invention.
[0390] In one embodiment, a method for detecting a 191P4D12(b) mRNA
in a biological sample comprises producing cDNA from the sample by
reverse transcription using at least one primer; amplifying the
cDNA so produced using a 191P4D12(b) polynucleotides as sense and
antisense primers to amplify 191P4D12(b) cDNAs therein; and
detecting the presence of the amplified 191P4D12(b) cDNA.
Optionally, the sequence of the amplified 191P4D12(b) cDNA can be
determined.
[0391] In another embodiment, a method of detecting a 191P4D12(b)
gene in a biological sample comprises first isolating genomic DNA
from the sample; amplifying the isolated genomic DNA using
191P4D12(b) polynucleotides as sense and antisense primers; and
detecting the presence of the amplified 191P4D12(b) gene. Any
number of appropriate sense and antisense probe combinations can be
designed from a 191P4D12(b) nucleotide sequence (see, e.g., FIG. 2)
and used for this purpose.
[0392] The invention also provides assays for detecting the
presence of a 191P4D12(b) protein in a tissue or other biological
sample such as serum, semen, bone, prostate, urine, cell
preparations, and the like. Methods for detecting a
191P4D12(b)-related protein are also well known and include, for
example, immunoprecipitation, immunohistochemical analysis, Western
blot analysis, molecular binding assays, ELISA, ELIFA and the like.
For example, a method of detecting the presence of a
191P4D12(b)-related protein in a biological sample comprises first
contacting the sample with a 191P4D12(b) antibody, a
191P4D12(b)-reactive fragment thereof, or a recombinant protein
containing an antigen-binding region of a 191P4D12(b) antibody; and
then detecting the binding of 191P4D12(b)-related protein in the
sample.
[0393] Methods for identifying a cell that expresses 191P4D12(b)
are also within the scope of the invention. In one embodiment, an
assay for identifying a cell that expresses a 191P4D12(b) gene
comprises detecting the presence of 191P4D12(b) mRNA in the cell.
Methods for the detection of particular mRNAs in cells are well
known and include, for example, hybridization assays using
complementary DNA probes (such as in situ hybridization using
labeled 191P4D12(b) riboprobes, Northern blot and related
techniques) and various nucleic acid amplification assays (such as
RT-PCR using complementary primers specific for 191P4D12(b), and
other amplification type detection methods, such as, for example,
branched DNA, SISBA, TMA and the like). Alternatively, an assay for
identifying a cell that expresses a 191P4D12(b) gene comprises
detecting the presence of 191P4D12(b)-related protein in the cell
or secreted by the cell. Various methods for the detection of
proteins are well known in the art and are employed for the
detection of 191P4D12(b)-related proteins and cells that express
191P4D12(b)-related proteins.
[0394] 191P4D12(b) expression analysis is also useful as a tool for
identifying and evaluating agents that modulate 191P4D12(b) gene
expression. For example, 191P4D12(b) expression is significantly
upregulated in prostate cancer, and is expressed in cancers of the
tissues listed in Table I. Identification of a molecule or
biological agent that inhibits 191P4D12(b) expression or
over-expression in cancer cells is of therapeutic value. For
example, such an agent can be identified by using a screen that
quantifies 191P4D12(b) expression by RT-PCR, nucleic acid
hybridization or antibody binding.
[0395] VIII.) Methods for Monitoring the Status of
191P4D12(b)-related Genes and Their Products
[0396] Oncogenesis is known to be a multistep process where
cellular growth becomes progressively dysregulated and cells
progress from a normal physiological state to precancerous and then
cancerous states (see, e.g., Alers et al., Lab Invest. 77(5):
437-438 (1997) and Isaacs et al., Cancer Surv. 23: 19-32 (1995)).
In this context, examining a biological sample for evidence of
dysregulated cell growth (such as aberrant 191P4D12(b) expression
in cancers) allows for early detection of such aberrant physiology,
before a pathologic state such as cancer has progressed to a stage
that therapeutic options are more limited and or the prognosis is
worse. In such examinations, the status of 191P4D12(b) in a
biological sample of interest can be compared, for example, to the
status of 191P4D12(b) in a corresponding normal sample (e.g. a
sample from that individual or alternatively another individual
that is not affected by a pathology). An alteration in the status
of 191P4D12(b) in the biological sample (as compared to the normal
sample) provides evidence of dysregulated cellular growth. In
addition to using a biological sample that is not affected by a
pathology as a normal sample, one can also use a predetermined
normative value such as a predetermined normal level of mRNA
expression (see, e.g., Grever et al., J. Comp. Neurol. 1996 Dec 9;
376(2): 306-14 and U.S. Pat. No. 5,837,501) to compare 191P4D12(b)
status in a sample.
[0397] The term "status" in this context is used according to its
art accepted meaning and refers to the condition or state of a gene
and its products. Typically, skilled artisans use a number of
parameters to evaluate the condition or state of a gene and its
products. These include, but are not limited to the location of
expressed gene products (including the location of 191P4D12(b)
expressing cells) as well as the level, and biological activity of
expressed gene products (such as 191P4D12(b) mRNA, polynucleotides
and polypeptides). Typically, an alteration in the status of
191P4D12(b) comprises a change in the location of 191P4D12(b)
and/or 191P4D12(b) expressing cells and/or an increase in
191P4D12(b) mRNA and/or protein expression.
[0398] 191P4D12(b) status in a sample can be analyzed by a number
of means well known in the art, including without limitation,
immunohistochemical analysis, in situ hybridization, RT-PCR
analysis on laser capture micro-dissected samples, Western blot
analysis, and tissue array analysis. Typical protocols for
evaluating the status of a 191P4D12(b) gene and gene products are
found, for example in Ausubel et al. eds., 1995, Current Protocols
In Molecular Biology, Units 2 (Northern Blotting), 4 (Southern
Blotting), 15 (Immunoblotting) and 18 (PCR Analysis). Thus, the
status of 191P4D12(b) in a biological sample is evaluated by
various methods utilized by skilled artisans including, but not
limited to genomic Southern analysis (to examine, for example
perturbations in a 191P4D12(b) gene), Northern analysis and/or PCR
analysis of 191P4D12(b) mRNA (to examine, for example alterations
in the polynucleotide sequences or expression levels of 191P4D12(b)
mRNAs), and, Western and/or immunohistochemical analysis (to
examine, for example alterations in polypeptide sequences,
alterations in polypeptide localization within a sample,
alterations in expression levels of 191P4D12(b) proteins and/or
associations of 191P4D12(b) proteins with polypeptide binding
partners). Detectable 191P4D12(b) polynucleotides include, for
example, a 191P4D12(b) gene or fragment thereof, 191P4D12(b) mRNA,
alternative splice vaniants, 191P4D12(b) mRNAs, and recombinant DNA
or RNA molecules containing a 191P4D12(b) polynucleotide.
[0399] The expression profile of 191P4D12(b) makes it a diagnostic
marker for local and/or metastasized disease, and provides
information on the growth or oncogenic potential of a biological
sample. In particular, the status of 191P4D12(b) provides
information useful for predicting susceptibility to particular
disease stages, progression, and/or tumor aggressiveness. The
invention provides methods and assays for determining 191P4D12(b)
status and diagnosing cancers that express 191P4D12(b), such as
cancers of the tissues listed in Table I. For example, because
191P4D12(b) mRNA is so highly expressed in prostate and other
cancers relative to normal prostate tissue, assays that evaluate
the levels of 191P4D12(b) mRNA transcripts or proteins in a
biological sample can be used to diagnose a disease associated with
191P4D12(b) dysregulation, and can provide prognostic information
useful in defining appropriate therapeutic options.
[0400] The expression status of 191P4D12(b) provides information
including the presence, stage and location of dysplastic,
precancerous and cancerous cells, predicting susceptibility to
various stages of disease, and/or for gauging tumor aggressiveness.
Moreover, the expression profile makes it useful as an imaging
reagent for metastasized disease. Consequently, an aspect of the
invention is directed to the various molecular prognostic and
diagnostic methods for examining the status of 191P4D12(b) in
biological samples such as those from individuals suffering from,
or suspected of suffering from a pathology characterized by
dysregulated cellular growth, such as cancer.
[0401] As described above, the status of 191P4D12(b) in a
biological sample can be examined by a number of well-known
procedures in the art. For example, the status of 191P4D12(b) in a
biological sample taken from a specific location in the body can be
examined by evaluating the sample for the presence or absence of
191P4D12(b) expressing cells (e.g. those that express 191P4D12(b)
mRNAs or proteins). This examination can provide evidence of
dysregulated cellular growth, for example, when
191P4D12(b)-expressing cells are found in a biological sample that
does not normally contain such cells (such as a lymph node),
because such alterations in the status of 191P4D12(b) in a
biological sample are often associated with dysregulated cellular
growth. Specifically, one indicator of dysregulated cellular growth
is the metastases of cancer cells from an organ of origin (such as
the prostate) to a different area of the body (such as a lymph
node). In this context, evidence of dysregulated cellular growth is
important for example because occult lymph node metastases can be
detected in a substantial proportion of patients with prostate
cancer, and such metastases are associated with known predictors of
disease progression (see, e.g., Murphy et al., Prostate 42(4):
315-317 (2000);Su et al., Semin. Surg. Oncol. 18(1): 17-28 (2000)
and Freeman et al., J Urol 1995 Aug 154(2 Pt 1):474-8).
[0402] In one aspect, the invention provides methods for monitoring
191P4D12(b) gene products by determining the status of 191P4D12(b)
gene products expressed by cells from an individual suspected of
having a disease associated with dysregulated cell growth (such as
hyperplasia or cancer) and then comparing the status so determined
to the status of 191P4D12(b) gene products in a corresponding
normal sample. The presence of aberrant 191P4D12(b) gene products
in the test sample relative to the normal sample provides an
indication of the presence of dysregulated cell growth within the
cells of the individual.
[0403] In another aspect, the invention provides assays useful in
determining the presence of cancer in an individual, comprising
detecting a significant increase in 191P4D12(b) mRNA or protein
expression in a test cell or tissue sample relative to expression
levels in the corresponding normal cell or tissue. The presence of
191P4D12(b) mRNA can, for example, be evaluated in tissues
including but not limited to those listed in Table I. The presence
of significant 191P4D12(b) expression in any of these tissues is
useful to indicate the emergence, presence and/or severity of a
cancer, since the corresponding normal tissues do not express
191P4D12(b) mRNA or express it at lower levels.
[0404] In a related embodiment, 191P4D12(b) status is determined at
the protein level rather than at the nucleic acid level. For
example, such a method comprises determining the level of
191P4D12(b) protein expressed by cells in a test tissue sample and
comparing the level so determined to the level of 191P4D12(b)
expressed in a corresponding normal sample. In one embodiment, the
presence of 191P4D12(b) protein is evaluated, for example, using
immunohistochemical methods. 191P4D12(b) antibodies or binding
partners capable of detecting 191P4D12(b) protein expression are
used in a variety of assay formats well known in the art for this
purpose.
[0405] In a further embodiment, one can evaluate the status of
191P4D12(b) nucleotide and amino acid sequences in a biological
sample in order to identify perturbations in the structure of these
molecules. These perturbations can include insertions, deletions,
substitutions and the like. Such evaluations are useful because
perturbations in the nucleotide and amino acid sequences are
observed in a large number of proteins associated with a growth
dysregulated phenotype (see, e.g., Marrogi et al., 1999, J. Cutan.
Pathol. 26(8):369-378). For example, a mutation in the sequence of
191P4D12(b) may be indicative of the presence or promotion of a
tumor. Such assays therefore have diagnostic and predictive value
where a mutation in 191P4D12(b) indicates a potential loss of
function or increase in tumor growth.
[0406] A wide variety of assays for observing perturbations in
nucleotide and amino acid sequences are well known in the art. For
example, the size and structure of nucleic acid or amino acid
sequences of 191P4D12(b) gene products are observed by the
Northern, Southern, Western, PCR and DNA sequencing protocols
discussed herein. In addition, other methods for observing
perturbations in nucleotide and amino acid sequences such as single
strand conformation polymorphism analysis are well known in the art
(see, e.g., U.S. Pat. Nos. 5,382,510 issued 7 Sep. 1999, and
5,952,170 issued 17 Jan. 1995).
[0407] Additionally, one can examine the methylation status of a
191P4D12(b) gene in a biological sample. Aberrant demethylation
and/or hypermethylation of CpG islands in gene 5' regulatory
regions frequently occurs in immortalized and transformed cells,
and can result in altered expression of various genes. For example,
promoter hypermethylation of the pi-class glutathione S-transferase
(a protein expressed in normal prostate but not expressed in
>90% of prostate carcinomas) appears to permanently silence
transcription of this gene and is the most frequently detected
genomic alteration in prostate carcinomas (De Marzo et al., Am. J.
Pathol. 155(6): 1985-1992 (1999)). In addition, this alteration is
present in at least 70% of cases of high-grade prostatic
intraepithelial neoplasia (PIN) (Brooks et al., Cancer Epidemiol.
Biomarkers Prev., 1998, 7:531-536). In another example, expression
of the LAGE-I tumor specific gene (which is not expressed in normal
prostate but is expressed in 25-50% of prostate cancers) is induced
by deoxy-azacytidine in lymphoblastoid cells, suggesting that
tumoral expression is due to demethylation (Lethe et al., Int. J.
Cancer 76(6): 903-908 (1998)). A variety of assays for examining
methylation status of a gene are well known in the art. For
example, one can utlize, in Southern hybridization approaches,
methylation-sensitive restriction enzymes that cannot cleave
sequences that contain methylated CpG sites to assess the
methylation status of CpG islands. In addition, MSP (methylation
specific PCR) can rapidly profile the methylation status of all the
CpG sites present in a CpG island of a given gene. This procedure
involves initial modification of DNA by sodium bisulfite (which
will convert all unmethylated cytosines to uracil) followed by
amplification using primers specific for methylated versus
unmethylated DNA. Protocols involving methylation interference can
also be found for example in Current Protocols In Molecular
Biology, Unit 12, Frederick M. Ausubel et al. eds., 1995.
[0408] Gene amplification is an additional method for assessing the
status of 191P4D12(b). Gene amplification is measured in a sample
directly, for example, by conventional Southern blotting or
Northern blotting to quantitate the transcription of mRNA (Thomas,
1980, Proc. Natl. Acad. Sci. USA, 77:5201-5205), dot blotting (DNA
analysis), or in situ hybridization, using an appropriately labeled
probe, based on the sequences provided herein. Alternatively,
antibodies are employed that recognize specific duplexes, including
DNA duplexes, RNA duplexes, and DNA-RNA hybrid duplexes or
DNA-protein duplexes. The antibodies in turn are labeled and the
assay carried out where the duplex is bound to a surface, so that
upon the formation of duplex on the surface, the presence of
antibody bound to the duplex can be detected.
[0409] Biopsied tissue or peripheral blood can be conveniently
assayed for the presence of cancer cells using for example,
Northern, dot blot or RT-PCR analysis to detect 191P4D12(b)
expression. The presence of RT-PCR amplifiable 191P4D12(b) mRNA
provides an indication of the presence of cancer. RT-PCR assays are
well known in the art. RT-PCR detection assays for tumor cells in
peripheral blood are currently being evaluated for use in the
diagnosis and management of a number of human solid tumors. In the
prostate cancer field, these include RT-PCR assays for the
detection of cells expressing PSA and PSM (Verkaik et al., 1997,
Urol. Res. 25:373-384; Ghossein et al., 1995, J. Clin. Oncol.
13:1195-2000; Heston et al., 1995, Clin. Chem. 41:1687-1688).
[0410] A further aspect of the invention is an assessment of the
susceptibility that an individual has for developing cancer. In one
embodiment, a method for predicting susceptibility to cancer
comprises detecting 191P4D12(b) mRNA or 191P4D12(b) protein in a
tissue sample, its presence indicating susceptibility to cancer,
wherein the degree of 191P4D12(b) mRNA expression correlates to the
degree of susceptibility. In a specific embodiment, the presence of
191P4D12(b) in prostate or other tissue is examined, with the
presence of 191P4D12(b) in the sample providing an indication of
prostate cancer susceptibility (or the emergence or existence of a
prostate tumor). Similarly, one can evaluate the integrity
191P4D12(b) nucleotide and amino acid sequences in a biological
sample, in order to identify perturbations in the structure of
these molecules such as insertions, deletions, substitutions and
the like. The presence of one or more perturbations in 191P4D12(b)
gene products in the sample is an indication of cancer
susceptibility (or the emergence or existence of a tumor).
[0411] The invention also comprises methods for gauging tumor
aggressiveness. In one embodiment, a method for gauging
aggressiveness of a tumor comprises determining the level of
191P4D12(b) mRNA or 191P4D12(b) protein expressed by tumor cells,
comparing the level so determined to the level of 191P4D12(b) mRNA
or 191P4D12(b) protein expressed in a corresponding normal tissue
taken from the same individual or a normal tissue reference sample,
wherein the degree of 191P4D12(b) mRNA or 191P4D12(b) protein
expression in the tumor sample relative to the normal sample
indicates the degree of aggressiveness. In a specific embodiment,
aggressiveness of a tumor is evaluated by determining the extent to
which 191P4D12(b) is expressed in the tumor cells, with higher
expression levels indicating more aggressive tumors. Another
embodiment is the evaluation of the integrity of 191P4D12(b)
nucleotide and amino acid sequences in a biological sample, in
order to identify perturbations in the structure of these molecules
such as insertions, deletions, substitutions and the like. The
presence of one or more perturbations indicates more aggressive
tumors.
[0412] Another embodiment of the invention is directed to methods
for observing the progression of a malignancy in an individual over
time. In one embodiment, methods for observing the progression of a
malignancy in an individual over time comprise determining the
level of 191P4D12(b) mRNA or 191P4D12(b) protein expressed by cells
in a sample of the tumor, comparing the level so determined to the
level of 191P4D12(b) mRNA or 191P4D12(b) protein expressed in an
equivalent tissue sample taken from the same individual at a
different time, wherein the degree of 191P4D12(b) mRNA or
191P4D12(b) protein expression in the tumor sample over time
provides information on the progression of the cancer. In a
specific embodiment, the progression of a cancer is evaluated by
determining 191P4D12(b) expression in the tumor cells over time,
where increased expression overtime indicates a progression of the
cancer. Also, one can evaluate the integrity 191P4D12(b) nucleotide
and amino acid sequences in a biological sample in order to
identify perturbations in the structure of these molecules such as
insertions, deletions, substitutions and the like, where the
presence of one or more perturbations indicates a progression of
the cancer.
[0413] The above diagnostic approaches can be combined with any one
of a wide variety of prognostic and diagnostic protocols known in
the art. For example, another embodiment of the invention is
directed to methods for observing a coincidence between the
expression of 191P4D12(b) gene and 191P4D12(b) gene products (or
perturbations in 191P4D12(b) gene and 191P4D12(b) gene products)
and a factor that is associated with malignancy, as a means for
diagnosing and prognosticating the status of a tissue sample. A
wide variety of factors associated with malignancy can be utilized,
such as the expression of genes associated with malignancy (e.g.
PSA, PSCA and PSM expression for prostate cancer etc.) as well as
gross cytological observations (see, e.g., Bocking et al, 1984,
Anal. Quant. Cytol. 6(2):74-88; Epstein, 1995, Hum. Pathol.
26(2):223-9; Thorson et al., 1998, Mod. Pathol. 11(6):543-51;
Baisden et al., 1999, Am. J. Surg. Pathol. 23(8):918-24). Methods
for observing a coincidence between the expression of 191P4D12(b)
gene and 191P4D12(b) gene products (or perturbations in 191P4D12(b)
gene and 191P4D12(b) gene products) and another factor that is
associated with malignancy are useful, for example, because the
presence of a set of specific factors that coincide with disease
provides information crucial for diagnosing and prognosticating the
status of a tissue sample.
[0414] In one embodiment, methods for observing a coincidence
between the expression of 191P4D12(b) gene and 191P4D12(b) gene
products (or perturbations in 191P4D12(b) gene and 191P4D12(b) gene
products) and another factor associated with malignancy entails
detecting the overexpression of 191P4D12(b) mRNA or protein in a
tissue sample, detecting the overexpression of PSA mRNA or protein
in a tissue sample (or PSCA or PSM expression), and observing a
coincidence of 191P4D12(b) mRNA or protein and PSA mRNA or protein
overexpression (or PSCA or PSM expression). In a specific
embodiment, the expression of 191P4D12(b) and PSA mRNA in prostate
tissue is examined, where the coincidence of 191P4D12(b) and PSA
mRNA overexpression in the sample indicates the existence of
prostate cancer, prostate cancer susceptibility or the emergence or
status of a prostate tumor.
[0415] Methods for detecting and quantifying the expression of
191P4D12(b) mRNA or protein are described herein, and standard
nucleic acid and protein detection and quantification technologies
are well known in the art. Standard methods for the detection and
quantification of 191P4D12(b) mRNA include in situ hybridization
using labeled 191P4D12(b) riboprobes, Northern blot and related
techniques using 191P4D12(b) polynucleotide probes, RT-PCR analysis
using primers specific for 191P4D12(b), and other amplification
type detection methods, such as, for example, branched DNA, SISBA,
TMA and the like. In a specific embodiment, semi-quantitative
RT-PCR is used to detect and quantify 191P4D12(b) mRNA expression.
Any number of primers capable of amplifying 191P4D12(b) can be used
for this purpose, including but not limited to the various primer
sets specifically described herein. In a specific embodiment,
polyclonal or monoclonal antibodies specifically reactive with the
wild-type 191P4D12(b) protein can be used in an immunohistochemical
assay of biopsied tissue.
[0416] IX.) Identification of Molecules That Interact With
191P4D12(b)
[0417] The 191P4D12(b) protein and nucleic acid sequences disclosed
herein allow a skilled artisan to identify proteins, small
molecules and other agents that interact with 191P4D12(b), as well
as pathways activated by 191P4D12(b) via any one of a variety of
art accepted protocols. For example, one can utilize one of the
so-called interaction trap systems (also referred to as the
"two-hybrid assay"). In such systems, molecules interact and
reconstitute a transcription factor which directs expression of a
reporter gene, whereupon the expression of the reporter gene is
assayed. Other systems identify protein-protein interactions in
vivo through reconstitution of a eukaryotic transcriptional
activator, see, e.g., U.S. Pat. Nos. 5,955,280 issued 21 Sep. 1999,
5,925,523 issued 20 Jul. 1999, 5,846,722 issued 8 Dec. 1998 and
6,004,746 issued 21 Dec. 1999. Algorithms are also available in the
art for genome-based predictions of protein function (see, e.g.,
Marcotte, et al, Nature 402: 4 Nov. 1999, 83-86).
[0418] Alternatively one can screen peptide libraries to identify
molecules that interact with 191P4D12(b) protein sequences. In such
methods, peptides that bind to 191P4D12(b) are identified by
screening libraries that encode a random or controlled collection
of amino acids. Peptides encoded by the libraries are expressed as
fusion proteins of bacteriophage coat proteins, the bacteriophage
particles are then screened against the 191P4D12(b) protein(s).
[0419] Accordingly, peptides having a wide variety of uses, such as
therapeutic, prognostic or diagnostic reagents, are thus identified
without any prior information on the structure of the expected
ligand or receptor molecule. Typical peptide libraries and
screening methods that can be used to identify molecules that
interact with 191P4D12(b) protein sequences are disclosed for
example in U.S. Pat. Nos. 5,723,286 issued 3 Mar. 1998 and
5,733,731 issued 31 Mar. 1998.
[0420] Alternatively, cell lines that express 191P4D12(b) are used
to identify protein-protein interactions mediated by 191P4D12(b).
Such interactions can be examined using immunoprecipitation
techniques (see, e.g., Hamilton B. J., et al. Biochem. Biophys.
Res. Commun. 1999, 261:646-51). 191P4D12(b) protein can be
immunoprecipitated from 191P4D12(b)-expressing cell lines using
anti-191P4D12(b) antibodies. Alternatively, antibodies against
His-tag can be used in a cell line engineered to express fusions of
191P4D12(b) and a His-tag (vectors mentioned above). The
immunoprecipitated complex can be examined for protein association
by procedures such as Western blotting, .sup.35S-methionine
labeling of proteins, protein microsequencing, silver staining and
two-dimensional gel electrophoresis.
[0421] Small molecules and ligands that interact with 191P4D12(b)
can be identified through related embodiments of such screening
assays. For example, small molecules can be identified that
interfere with protein function, including molecules that interfere
with 191P4D12(b)'s ability to mediate phosphoryation and
de-phosphorylation, interaction with DNA or RNA molecules as an
indication of regulation of cell cycles, second messenger signaling
or tumorigenesis. Similarly, small molecules that modulate
191P4D12(b)-related ion channel, protein pump, or cell
communication functions are identified and used to treat patients
that have a cancer that expresses 191P4D12(b) (see, e.g., Hille,
B., Ionic Channels of Excitable Membranes 2.sup.nd Ed., Sinauer
Assoc., Sunderland, Mass., 1992). Moreover, ligands that regulate
191P4D12(b) function can be identified based on their ability to
bind 191P4D12(b) and activate a reporter construct. Typical methods
are discussed for example in U.S. Pat. No. 5,928,868 issued 27 Jul.
1999, and include methods for forming hybrid ligands in which at
least one ligand is a small molecule. In an illustrative
embodiment, cells engineered to express a fusion protein of
191P4D12(b) and a DNA-binding protein are used to co-express a
fusion protein of a hybrid ligand/small molecule and a cDNA library
transcriptional activator protein. The cells further contain a
reporter gene, the expression of which is conditioned on the
proximity of the first and second fusion proteins to each other, an
event that occurs only if the hybrid ligand binds to target sites
on both hybrid proteins. Those cells that express the reporter gene
are selected and the unknown small molecule or the unknown ligand
is identified. This method provides a means of identifying
modulators, which activate or inhibit 191P4D12(b).
[0422] An embodiment of this invention comprises a method of
screening for a molecule that interacts with a 191P4D12(b) amino
acid sequence shown in FIG. 2 or FIG. 3, comprising the steps of
contacting a population of molecules with a 191P4D12(b) amino acid
sequence, allowing the population of molecules and the 191P4D12(b)
amino acid sequence to interact under conditions that facilitate an
interaction, determining the presence of a molecule that interacts
with the 191P4D12(b) amino acid sequence, and then separating
molecules that do not interact with the 191P4D12(b) amino acid
sequence from molecules that do. In a specific embodiment, the
method further comprises purifying, characterizing and identifying
a molecule that interacts with the 191P4D12(b) amino acid sequence.
The identified molecule can be used to modulate a function
performed by 191P4D12(b). In a preferred embodiment, the
191P4D12(b) amino acid sequence is contacted with a library of
peptides.
[0423] X. Therapeutic Methods and Compositions
[0424] The identification of 191P4D12(b) as a protein that is
normally expressed in a restricted set of tissues, but which is
also expressed in cancers such as those listed in Table I, opens a
number of therapeutic approaches to the treatment of such
cancers.
[0425] Of note, targeted antitumor therapies have been useful even
when the targeted protein is expressed on normal tissues, even
vital normal organ tissues. A vital organ is one that is necessary
to sustain life, such as the heart or colon. A non-vital organ is
one that can be removed whereupon the individual is still able to
survive. Examples of non-vital organs are ovary, breast, and
prostate.
[0426] For example, Herceptin.RTM. is an FDA approved
pharmaceutical that has as its active ingredient an antibody which
is immunoreactive with the protein variously known as HER2,
HER2/neu, and erb-b-2. It is marketed by Genentech and has been a
commercially successful antitumor agent. Herceptin sales reached
almost $400 million in 2002. Herceptin is a treatment for HER2
positive metastatic breast cancer. However, the expression of HER2
is not limited to such tumors. The same protein is expressed in a
number of normal tissues. In particular, it is known that HER2/neu
is present in normal kidney and heart, thus these tissues are
present in all human recipients of Herceptin. The presence of
HER2/neu in normal kidney is also confirmed by Latif, Z., et al.,
B.J.U. International (2002) 89:5-9. As shown in this article (which
evaluated whether renal cell carcinoma should be a preferred
indication for anti-HER2 antibodies such as Herceptin) both protein
and mRNA are produced in benign renal tissues. Notably, HER2/neu
protein was strongly overexpressed in benign renal tissue. Despite
the fact that HER2/neu is expressed in such vital tissues as heart
and kidney, Herceptin is a very useful, FDA approved, and
commercially successful drug. The effect of Herceptin on cardiac
tissue, i.e., "cardiotoxicity," has merely been a side effect to
treatment. When patients were treated with Herceptin alone,
significant cardiotoxicity occurred in a very low percentage of
patients.
[0427] Of particular note, although kidney tissue is indicated to
exhibit normal expression, possibly even higher expression than
cardiac tissue, kidney has no appreciable Herceptin side effect
whatsoever. Moreover, of the diverse array of normal tissues in
which HER2 is expressed, there is very little occurrence of any
side effect. Only cardiac tissue has manifested any appreciable
side effect at all. A tissue such as kidney, where HER2/neu
expression is especially notable, has not been the basis for any
side effect.
[0428] Furthermore, favorable therapeutic effects have been found
for anti tumor therapies that target epidermal growth factor
receptor (EGFR). EGFR is also expressed in numerous normal tissues.
There have been very limited side effects in normal tissues
following use of anti-EGFR therapeutics.
[0429] Thus, expression of a target protein in normal tissue, even
vital normal tissue, does not defeat the utility of a targeting
agent for the protein as a therapeutic for certain tumors in which
the protein is also overexpressed.
[0430] Accordingly, therapeutic approaches that inhibit the
activity of a 191P4D12(b) protein are useful for patients suffering
from a cancer that expresses 191P4D12(b). These therapeutic
approaches generally fall into two classes. One class comprises
various methods for inhibiting the binding or association of a
191P4D12(b) protein with its binding partner or with other
proteins. Another class comprises a variety of methods for
inhibiting the transcription of a 191P4D12(b) gene or translation
of 191P4D12(b) mRNA.
[0431] X.A.) Anti-Cancer Vaccines
[0432] The invention provides cancer vaccines comprising a
191P4D12(b)-related protein or 191P4D12(b)-related nucleic acid. In
view of the expression of 191P4D12(b), cancer vaccines prevent
and/or treat 191P4D12(b)-expressing cancers with minimal or no
effects on non-target tissues. The use of a tumor antigen in a
vaccine that generates humoral and/or cell-mediated immune
responses as anti-cancer therapy is well known in the art and has
been employed in prostate cancer using human PSMA and rodent PAP
immunogens (Hodge et al., 1995, Int. J. Cancer 63:231-237; Fong et
al., 1997, J. Immunol. 159:3113-3117).
[0433] Such methods can be readily practiced by employing a
191P4D12(b)-related protein, or a 191P4D12(b)-encoding nucleic acid
molecule and recombinant vectors capable of expressing and
presenting the 191P4D12(b) immunogen (which typically comprises a
number of antibody or T cell epitopes). Skilled artisans understand
that a wide variety of vaccine systems for delivery of
immunoreactive epitopes are known in the art (see, e.g., Heryln et
al., Ann Med 1999 Feb 31(1):66-78; Maruyama et al., Cancer Immunol
Immunother 2000 Jun 49(3):123-32) Briefly, such methods of
generating an immune response (e.g. humoral and/or cell-mediated)
in a mammal, comprise the steps of: exposing the mammal's immune
system to an immunoreactive epitope (e.g. an epitope present in a
191P4D12(b) protein shown in FIG. 3 or analog or homolog thereof)
so that the mammal generates an immune response that is specific
for that epitope (e.g. generates antibodies that specifically
recognize that epitope). In a preferred method, a 191P4D12(b)
immunogen contains a biological motif, see e.g., Tables VIII-XXI
and XXII-XLIX, or a peptide of a size range from 191P4D12(b)
indicated in FIG. 5, FIG. 6, FIG. 7, FIG. 8, and FIG. 9.
[0434] The entire 191P4D12(b) protein, immunogenic regions or
epitopes thereof can be combined and delivered by various means.
Such vaccine compositions can include, for example, lipopeptides
(e.g., Vitiello, A. et al., J. Clin. Invest. 95:341, 1995), peptide
compositions encapsulated in poly(DL-lactide-co-glycolide) ("PLG")
microspheres (see, e.g., Eldridge, et al., Molec. Immunol.
28:287-294, 1991: Alonso et al., Vaccine 12:299-306, 1994; Jones et
al., Vaccine 13:675-681, 1995), peptide compositions contained in
immune stimulating complexes (ISCOMS) (see, e.g., Takahashi et al.,
Nature 344:873-875,1990; Hu et al., Clin Exp Immunol.
113:235-243,1998), multiple antigen peptide systems (MAPs) (see
e.g., Tam, J. P., Proc. Natl. Acad. Sci. U.S.A. 85:5409-5413, 1988;
Tam, J. P., J. Immunol. Methods 196:17-32, 1996), peptides
formulated as multivalent peptides; peptides for use in ballistic
delivery systems, typically crystallized peptides, viral delivery
vectors (Perkus, M. E. et al., In: Concepts in vaccine development,
Kaufmann, S. H. E., ed., p. 379, 1996; Chakrabarti, S. et al.,
Nature 320:535, 1986; Hu, S. L. et al., Nature 320:537, 1986;
Kieny, M.-P. et al., AIDS Bio/Technology 4:790, 1986; Top, F. H. et
al., J. Infect. Dis. 124:148, 1971; Chanda, P. K. et al., Virology
175:535, 1990), particles of viral or synthetic origin (e.g.,
Kofler, N. et al., J. Immunol. Methods. 192:25, 1996; Eldridge, J.
H. et al., Sem. Hematol. 30:16, 1993; Falo, L. D., Jr. et al.,
Nature Med. 7:649,1995), adjuvants (Warren, H. S., Vogel, F. R.,
and Chedid, L. A. Annu. Rev. Immunol. 4:369,1986; Gupta, R. K. et
al., Vaccine 11:293, 1993), liposomes (Reddy, R. et al., J.
Immunol. 148:1585, 1992; Rock, K. L., Immunol. Today 17:131, 1996),
or, naked or particle absorbed cDNA (Ulmer, J. B. et al., Science
259:1745, 1993; Robinson, H. L., Hunt, L. A., and Webster, R. G.,
Vaccine 11:957, 1993; Shiver, J. W. et al., In: Concepts in vaccine
development, Kaufmann, S. H. E., ed., p. 423, 1996;, Cease, K. B.,
and Berzofsky, J. A., Annu. Rev. Immunol. 12:923, 1994 and
Eldridge, J. H. et al., Sem. Hematol. 30:16, 1993). Toxin-targeted
delivery technologies, also known as receptor mediated targeting,
such as those of Avant Immunotherapeutics, Inc. (Needham, Mass.)
may also be used.
[0435] In patients with 191P4D12(b)-associated cancer, the vaccine
compositions of the invention can also be used in conjunction with
other treatments used for cancer, e.g., surgery, chemotherapy, drug
therapies, radiation therapies, etc. including use in combination
with immune adjuvants such as IL-2, IL-12, GM-CSF, and the
like.
[0436] Cellular Vaccines:
[0437] CTL epitopes can be determined using specific algorithms to
identify peptides within 191P4D12(b) protein that bind
corresponding HLA alleles (see e.g., Table IV; Epimer.TM. and
Epimatrix.TM., Brown University (URL
brown.edu/Research/TB-HIV_Lab/epimatrix/epimatrix.html); and,
BIMAS, (URL bimas.dcrt.nih.gov/; SYFPEITHI at URL
syfpeithi.bmi-heidelberg.com/). In a preferred embodiment, a
191P4D12(b) immunogen contains one or more amino acid sequences
identified using techniques well known in the art, such as the
sequences shown in Tables VIII-XXI and XXII-XLIX or a peptide of 8,
9, 10 or 11 amino acids specified by an HLA Class I
motif/supermotif (e.g., Table IV (A), Table IV (D), or Table IV
(E)) and/or a peptide of at least 9 amino acids that comprises an
HLA Class II motif/supermotif (e.g., Table IV (B) or Table IV (C)).
As is appreciated in the art, the HLA Class I binding groove is
essentially closed ended so that peptides of only a particular size
range can fit into the groove and be bound, generally HLA Class I
epitopes are 8, 9, 10, or 11 amino acids long. In contrast, the HLA
Class II binding groove is essentially open ended; therefore a
peptide of about 9 or more amino acids can be bound by an HLA Class
II molecule. Due to the binding groove differences between HLA
Class I and II, HLA Class I motifs are length specific, i.e.,
position two of a Class I motif is the second amino acid in an
amino to carboxyl direction of the peptide. The amino acid
positions in a Class II motif are relative only to each other, not
the overall peptide, i.e., additional amino acids can be attached
to the amino and/or carboxyl termini of a motif-bearing sequence.
HLA Class II epitopes are often 9, 10, 11, 12, 13, 14, 15, 16, 17,
18, 19, 20, 21, 22, 23, 24, or 25 amino acids long, or longer than
25 amino acids.
[0438] Antibody-based Vaccines
[0439] A wide variety of methods for generating an immune response
in a mammal are known in the art (for example as the first step in
the generation of hybridomas). Methods of generating an immune
response in a mammal comprise exposing the mammal's immune system
to an immunogenic epitope on a protein (e.g. a 191P4D12(b) protein)
so that an immune response is generated. A typical embodiment
consists of a method for generating an immune response to
191P4D12(b) in a thereof; and at least one periodic interval
thereafter re-contacting the host with the 191P4D12(b) B cell or
cytotoxic T-cell epitope or analog thereof. A specific embodiment
consists of a method of generating an immune response against a
191P4D12(b)-related protein or a man-made multiepitopic peptide
comprising: administering 191P4D12(b) immunogen (e.g. a
191P.sup.4D12(b) protein or a peptide fragment thereof, a
191P4D12(b) fusion protein or analog etc.) in a vaccine preparation
to a human or another mammal. Typically, such vaccine preparations
further contain a suitable adjuvant (see, e.g., U.S. Pat. No.
6,146,635) or a universal helper epitope such as a PADRE.TM.
peptide (Epimmune Inc., San Diego, Calif.; see, e.g., Alexander et
al., J. Immunol. 2000 164(3); 164(3): 1625-1633; Alexander et al.,
Immunity 1994 1(9): 751-761 and Alexander et al., Immunol. Res.
1998 18(2): 79-92). An alternative method comprises generating an
immune response in an individual against a 191P4D12(b) immunogen
by: administering in vivo to muscle or skin of the individual's
body a DNA molecule that comprises a DNA sequence that encodes a
191P4D12(b) immunogen, the DNA sequence operatively linked to
regulatory sequences which control the expression of the DNA
sequence; wherein the DNA molecule is taken up by cells, the DNA
sequence is expressed in the cells and an immune response is
generated against the immunogen (see, e.g., U.S. Pat. No.
5,962,428). Optionally a genetic vaccine facilitator such as
anionic lipids; saponins; lectins; estrogenic compounds;
hydroxylated lower alkyls; dimethyl sulfoxide; and urea is also
administered. In addition, an antiidiotypic antibody can be
administered that mimics 191P4D12(b), in order to generate a
response to the target antigen.
[0440] Nucleic Acid Vaccines:
[0441] Vaccine compositions of the invention include nucleic
acid-mediated modalities. DNA or RNA that encode protein(s) of the
invention can be administered to a patient. Genetic immunization
methods can be employed to generate prophylactic or therapeutic
humoral and cellular immune responses directed against cancer cells
expressing 191P4D12(b). Constructs comprising DNA encoding a
191P4D12(b)-related protein/immunogen and appropriate regulatory
sequences can be injected directly into muscle or skin of an
individual, such that the cells of the muscle or skin take-up the
construct and express the encoded 191P4D12(b) protein/immunogen.
Alternatively, a vaccine comprises a 191P4D12(b)-related protein.
Expression of the 191P4D12(b)-related protein immunogen results in
the generation of prophylactic or therapeutic humoral and cellular
immunity against cells that bear a 191P4D12(b) protein. Various
prophylactic and therapeutic genetic immunization techniques known
in the art can be used (for review, see information and references
published at Internet address genweb.com). Nucleic acid-based
delivery is described, for instance, in Wolff et. al., Science
247:1465 (1990) as well as U.S. Pat. Nos. 5,580,859; 5,589,466;
5,804,566; 5,739,118; 5,736,524; 5,679,647; WO 98/04720. Examples
of DNA-based delivery technologies include "naked DNA", facilitated
(bupivicaine, polymers, peptide-mediated) delivery, cationic lipid
complexes, and particle-mediated ("gene gun") or pressure-mediated
delivery (see, e.g., U.S. Pat. No. 5,922,687).
[0442] For therapeutic or prophylactic immunization purposes,
proteins of the invention can be expressed via viral or bacterial
vectors. Various viral gene delivery systems that can be used in
the practice of the invention include, but are not limited to,
vaccinia, fowlpox, canarypox, adenovirus, influenza, poliovirus,
adeno-associated virus, lenfivirus, and sindbis virus (see, e.g.,
Restifo, 1996, Curr. Opin. Immunol. 8:658-663; Tsang et al. J.
Natl. Cancer Inst. 87:982-990 (1995)). Non-viral delivery systems
can also be employed by introducing naked DNA encoding a
191P4D12(b)-related protein into the patient (e.g., intramuscularly
or intradermally) to induce an anti-tumor response.
[0443] Vaccinia virus is used, for example, as a vector to express
nucleotide sequences that encode the peptides of the invention.
Upon introduction into a host, the recombinant vaccinia virus
expresses the protein immunogenic peptide, and thereby elicits a
host immune response. Vaccinia vectors and methods useful in
immunization protocols are described in, e.g., U.S. Pat. No.
4,722,848. Another vector is BCG (Bacille Calmette Guerin). BCG
vectors are described in Stover et al., Nature 351:456-460 (1991).
A wide variety of other vectors useful for therapeutic
administration or immunization of the peptides of the invention,
e.g. adeno and adeno-associated virus vectors, retroviral vectors,
Salmonella typhi vectors, detoxified anthrax toxin vectors, and the
like, will be apparent to those skilled in the art from the
description herein.
[0444] Thus, gene delivery systems are used to deliver a
191P4D12(b)-related nucleic acid molecule. In one embodiment, the
full-length human 191P4D12(b) cDNA is employed. In another
embodiment, 191P4D12(b) nucleic acid molecules encoding specific
cytotoxic T lymphocyte (CTL) and/or antibody epitopes are
employed.
[0445] Ex Vivo Vaccines
[0446] Various ex vivo strategies can also be employed to generate
an immune response. One approach involves the use of antigen
presenting cells (APCs) such as dendritic cells (DC) to present
191P4D12(b) antigen to a patient's immune system. Dendritic cells
express MHC class I and II molecules, B7 co-stimulator, and IL-12,
and are thus highly specialized antigen presenting cells. In
prostate cancer, autologous dendritic cells pulsed with peptides of
the prostate-specific membrane antigen (PSMA) are being used in a
Phase I clinical trial to stimulate prostate cancer patients'
immune systems (Tjoa et al., 1996, Prostate 28:65-69; Murphy et
al., 1996, Prostate 29:371-380). Thus, dendritic cells can be used
to present 191P4D12(b) peptides to T cells in the context of MHC
class I or II molecules. In one embodiment, autologous dendritic
cells are pulsed with 191P4D12(b) peptides capable of binding to
MHC class I and/or class II molecules. In another embodiment,
dendritic cells are pulsed with the complete 191P4D12(b) protein.
Yet another embodiment involves engineering the overexpression of a
191P4D12(b) gene in dendritic cells using various implementing
vectors known in the art, such as adenovirus (Arthur et al., 1997,
Cancer Gene Ther. 4:17-25), retrovirus (Henderson et al., 1996,
Cancer Res. 56:3763-3770), lentivirus, adeno-associated virus, DNA
transfection (Ribas et al., 1997, Cancer Res. 57:2865-2869), or
tumor-derived RNA transfection (Ashley et al., 1997, J. Exp. Med.
186:1177-1182). Cells that express 191P4D12(b) can also be
engineered to express immune modulators, such as GM-CSF, and used
as immunizing agents.
[0447] X.B.) 191P4D12(b) as a Target for Antibody-based Therapy
[0448] 191P4D12(b) is an attractive target for antibody-based
therapeutic strategies. A number of antibody strategies are known
in the art for targeting both extracellular and intracellular
molecules (see, e.g., complement and ADCC mediated killing as well
as the use of intrabodies). Because 191P4D12(b) is expressed by
cancer cells of various lineages relative to corresponding normal
cells, systemic administration of 191P4D12(b)-immunoreactive
compositions are prepared that exhibit excellent sensitivity
without toxic, non-specific and/or non-target effects caused by
binding of the immunoreactive composition to non-target organs and
tissues. Antibodies specifically reactive with domains of
191P4D12(b) are useful to treat 191P4D12(b)-expressing cancers
systemically, either as conjugates with a toxin or therapeutic
agent, or as naked antibodies capable of inhibiting cell
proliferation or function.
[0449] 191P4D12(b) antibodies can be introduced into a patient such
that the antibody binds to 191P4D12(b) and modulates a function,
such as an interaction with a binding partner, and consequently
mediates destruction of the tumor cells and/or inhibits the growth
of the tumor cells. Mechanisms by which such antibodies exert a
therapeutic effect can include complement-mediated cytolysis,
antibody-dependent cellular cytotoxicity, modulation of the
physiological function of 191P4D12(b), inhibition of ligand binding
or signal transduction pathways, modulation of tumor cell
differentiation, alteration of tumor angiogenesis factor profiles,
and/or apoptosis.
[0450] Those skilled in the art understand that antibodies can be
used to specifically target and bind immunogenic molecules such as
an immunogenic region of a 191P4D12(b) sequence shown in FIG. 2 or
FIG. 3. In addition, skilled artisans understand that it is routine
to conjugate antibodies to cytotoxic agents (see, e.g., Slevers et
al. Blood 93:11 3678-3684 (Jun. 1, 1999)). When cytotoxic and/or
therapeutic agents are delivered directly to cells, such as by
conjugating them to antibodies specific for a molecule expressed by
that cell (e.g. 191P4D12(b)), the cytotoxic agent will exert its
known biological effect (i.e. cytotoxicity) on those cells.
[0451] A wide variety of compositions and methods for using
antibody-cytotoxic agent conjugates to kill cells are known in the
art. In the context of cancers, typical methods entail
administering to an animal having a tumor a biologically effective
amount of a conjugate comprising a selected cytotoxic and/or
therapeutic agent linked to a targeting agent (e.g. an
anti-191P4D12(b) antibody) that binds to a marker (e.g.
191P4D12(b)) expressed, accessible to binding or localized on the
cell surfaces. A typical embodiment is a method of delivering a
cytotoxic and/or therapeutic agent to a cell expressing
191P4D12(b), comprising conjugating the cytotoxic agent to an
antibody that immunospecifically binds to a 191P4D12(b) epitope,
and, exposing the cell to the antibody-agent conjugate. Another
illustrative embodiment is a method of treating an individual
suspected of suffering from metastasized cancer, comprising a step
of administering parenterally to said individual a pharmaceutical
composition comprising a therapeutically effective amount of an
antibody conjugated to a cytotoxic and/or therapeutic agent.
[0452] Cancer immunotherapy using anti-191P4D12(b) antibodies can
be done in accordance with various approaches that have been
successfully employed in the treatment of other types of cancer,
including but not limited to colon cancer (Arlen et al., 1998,
Crit. Rev. Immunol. 18:133-138), multiple myeloma (Ozaki et al.,
1997, Blood 90:3179-3186, Tsunenari et al., 1997, Blood
90:2437-2444), gastric cancer (Kasprzyk et al., 1992, Cancer Res.
52:2771-2776), B-cell lymphoma (Funakoshi et al., 1996, J.
Immunother. Emphasis Tumor Immunol. 19:93-101), leukemia (Zhong et
al., 1996, Leuk. Res. 20:581-589), colorectal cancer (Moun et al.,
1994, Cancer Res. 54:6160-6166; Velders et al., 1995, Cancer Res.
55:4398-4403), and breast cancer (Shepard et al., 1991, J. Clin.
Immunol. 11:117-127). Some therapeutic approaches involve
conjugation of naked antibody to a toxin or radioisotope, such as
the conjugation of Y91 or I.sup.131 to anti-CD20 antibodies (e.g.,
Zevalin.TM., IDEC Pharmaceuticals Corp. or Bexxar.TM., Coulter
Pharmaceuticals), while others involve co-administration of
antibodies and other therapeutic agents, such as Herceptin.TM.
(trastuzumab) with paclitaxel (Genentech, Inc.). The antibodies can
be conjugated to a therapeutic agent. To treat prostate cancer, for
example, 191P4D12(b) antibodies can be administered in conjunction
with radiation, chemotherapy or hormone ablation. Also, antibodies
can be conjugated to a toxin such as calicheamicin (e.g.,
Mylotarg.TM., Wyeth-Ayerst, Madison, N.J., a recombinant humanized
lgG.sub.4 kappa antibody conjugated to antitumor antibiotic
calicheamicin) or a maytansinoid (e.g., taxane-based
Tumor-Activated Prodrug, TAP, platform, ImmunoGen, Cambridge,
Mass., also see e.g., U.S. Pat. No. 5,416,064).
[0453] Although 191P4D12(b) antibody therapy is useful for all
stages of cancer, antibody therapy can be particularly appropriate
in advanced or metastatic cancers. Treatment with the antibody
therapy of the invention is indicated for patients who have
received one or more rounds of chemotherapy. Alternatively,
antibody therapy of the invention is combined with a
chemotherapeutic or radiation regimen for patients who have not
received chemotherapeutic treatment. Additionally, antibody therapy
can enable the use of reduced dosages of concomitant chemotherapy,
particularly for patients who do not tolerate the toxicity of the
chemotherapeutic agent very well. Fan et al. (Cancer Res.
53:4637-4642, 1993), Prewett et al. (International J. of Onco.
9:217-224, 1996), and Hancock et al. (Cancer Res. 51:4575-4580,
1991) describe the use of various antibodies together with
chemotherapeutic agents.
[0454] Although 191P4D12(b) antibody therapy is useful for all
stages of cancer, antibody therapy can be particularly appropriate
in advanced or metastatic cancers. Treatment with the antibody
therapy of the invention is indicated for patients who have
received one or more rounds of chemotherapy. Alternatively,
antibody therapy of the invention is combined with a
chemotherapeutic or radiation regimen for patients who have not
received chemotherapeutic treatment. Additionally, antibody therapy
can enable the use of reduced dosages of concomitant chemotherapy,
particularly for patients who do not tolerate the toxicity of the
chemotherapeutic agent very well.
[0455] Cancer patients can be evaluated for the presence and level
of 191P4D12(b) expression, preferably using immunohistochemical
assessments of tumor tissue, quantitative 191P4D12(b) imaging, or
other techniques that reliably indicate the presence and degree of
191P4D12(b) expression. Immunohistochemical analysis of tumor
biopsies or surgical specimens is preferred for this purpose.
Methods for immunohistochemical analysis of tumor tissues are well
known in the art.
[0456] Anti-191P4D12(b) monoclonal antibodies that treat prostate
and other cancers include those that initiate a potent immune
response against the tumor or those that are directly cytotoxic. In
this regard, anti-191P4D12(b) monoclonal antibodies (mAbs) can
elicit tumor cell lysis by either complement-mediated or
antibody-dependent cell cytotoxicity (ADCC) mechanisms, both of
which require an intact Fc portion of the immunoglobulin molecule
for interaction with effector cell Fc receptor sites on complement
proteins. In addition, anti-191P4D12(b) mAbs that exert a direct
biological effect on tumor growth are useful to treat cancers that
express 191P4D12(b). Mechanisms by which directly cytotoxic mAbs
act include: inhibition of cell growth, modulation of cellular
differentiation, modulation of tumor angiogenesis factor profiles,
and the induction of apoptosis. The mechanism(s) by which a
particular anti-191P4D12(b) mAb exerts an anti-tumor effect is
evaluated using any number of in vitro assays that evaluate cell
death such as ADCC, ADMMC, complement-mediated cell lysis, and so
forth, as is generally known in the art.
[0457] In some patients, the use of murine or other non-human
monoclonal antibodies, or human/mouse chimeric mAbs can induce
moderate to strong immune responses against the non-human antibody.
This can result in clearance of the antibody from circulation and
reduced efficacy. In the most severe cases, such an immune response
can lead to the extensive formation of immune complexes which,
potentially, can cause renal failure. Accordingly, preferred
monoclonal antibodies used in the therapeutic methods of the
invention are those that are either fully human or humanized and
that bind specifically to the target 191P4D12(b) antigen with high
affinity but exhibit low or no antigenicity in the patient.
[0458] Therapeutic methods of the invention contemplate the
administration of single anti-191P4D12(b) mAbs as well as
combinations, or cocktails, of different mAbs. Such mAb cocktails
can have certain advantages inasmuch as they contain mAbs that
target different epitopes, exploit different effector mechanisms or
combine directly cytotoxic mAbs with mAbs that rely on immune
effector functionality. Such mAbs in combination can exhibit
synergistic therapeutic effects, In addition, anti-191P4D12(b) mAbs
can be administered concomitantly with other therapeutic
modalities, including but not limited to various chemotherapeutic
agents, androgen-blockers, immune modulators (e.g., IL-2, GM-CSF),
surgery or radiation. The anti-191P4D12(b) mAbs are administered in
their "naked" or unconjugated form, or can have a therapeutic
agent(s) conjugated to them.
[0459] Anti-191P4D12(b) antibody formulations are administered via
any route capable of delivering the antibodies to a tumor cell.
Routes of administration include, but are not limited to,
intravenous, intraperitoneal, intramuscular, intratumor,
intradermal, and the like. Treatment generally involves repeated
administration of the anti-191P4D12(b) antibody preparation, via an
acceptable route of administration such as intravenous injection
(IV), typically at a dose in the range of about 0.1, 0.2, 0.3, 0.4,
0.5, 0.6, 0.7, 0.8, 0.9., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or
25 mg/kg body weight. In general, doses in the range of 10-1000 mg
mAb per week are effective and well tolerated.
[0460] Based on clinical experience with the Herceptin.TM. mAb in
the treatment of metastatic breast cancer, an initial loading dose
of approximately 4 mg/kg patient body weight IV, followed by weekly
doses of about 2 mg/kg IV of the anti-191P4D12(b) mAb preparation
represents an acceptable dosing regimen. Preferably, the initial
loading dose is administered as a 90-minute or longer infusion. The
periodic maintenance dose is administered as a 30 minute or longer
infusion, provided the initial dose was well tolerated. As
appreciated by those of skill in the art, various factors can
influence the ideal dose regimen in a particular case. Such factors
include, for example, the binding affinity and half life of the Ab
or mAbs used, the degree of 191P4D12(b) expression in the patient,
the extent of circulating shed 191P4D12(b) antigen, the desired
steady-state antibody concentration level, frequency of treatment,
and the influence of chemotherapeutic or other agents used in
combination with the treatment method of the invention, as well as
the health status of a particular patient.
[0461] Optionally, patients should be evaluated for the levels of
191P4D12(b) in a given sample (e.g. the levels of circulating
191P4D12(b) antigen and/or 191P4D12(b) expressing cells) in order
to assist in the determination of the most effective dosing
regimen, etc. Such evaluations are also used for monitoring
purposes throughout therapy, and are useful to gauge therapeutic
success in combination with the evaluation of other parameters (for
example, urine cytology and/or ImmunoCyt levels in bladder cancer
therapy, or by analogy, serum PSA levels in prostate cancer
therapy).
[0462] Anti-idiotypic anti-191P4D12(b) antibodies can also be used
in anti-cancer therapy as a vaccine for inducing an immune response
to cells expressing a 191P4D12(b)-related protein. In particular,
the generation of anti-idiotypic antibodies is well known in the
art; this methodology can readily be adapted to generate
anti-idiotypic anti-191P4D12(b) antibodies that mimic an epitope on
a 191P4D12(b)-related protein (see, for example, Wagner et al.,
1997, Hybridomal16: 33-40; Foon et al., 1995, J. Clin. Invest.
96:334-342;, Herlyn et al., 1996, Cancer Immunol. Immunother.
43:65-76). Such an anti-idiotypic antibody can be used in cancer
vaccine strategies.
[0463] X.C.) 191P4D12(b) as a Target for Cellular Immune
Responses
[0464] Vaccines and methods of preparing vaccines that contain an
immunogenically effective amount of one or more HLA-binding
peptides as described herein are further embodiments of the
invention. Furthermore, vaccines in accordance with the invention
encompass compositions of one or more of the claimed peptides. A
peptide can be present in a vaccine individually. Alternatively,
the peptide can exist as a homopolymer comprising multiple copies
of the same peptide, or as a heteropolymer of various peptides.
Polymers have the advantage of increased immunological reaction
and, where different peptide epitopes are used to make up the
polymer, the additional ability to induce antibodies and/or CTLs
that react with different antigenic determinants of the pathogenic
organism or tumor-related peptide targeted for an immune response.
The composition can be a naturally occurring region of an antigen
or can be prepared, e.g., recombinantly or by chemical
synthesis.
[0465] Carriers that can be used with vaccines of the invention are
well known in the art, and include, e.g., thyroglobulin, albumins
such as human serum albumin, tetanus toxoid, polyamino acids such
as poly L-lysine, poly L-glutamic acid, influenza, hepatitis B
virus core protein, and the like. The vaccines can contain a
physiologically tolerable (i.e., acceptable) diluent such as water,
or saline, preferably phosphate buffered saline. The vaccines also
typically include an adjuvant. Adjuvants such as incomplete
Freund's adjuvant, aluminum phosphate, aluminum hydroxide, or alum
are examples of materials well known in the art. Additionally, as
disclosed herein, CTL responses can be primed by conjugating
peptides of the invention to lipids, such as
tripalmitoyl-S-glycerylcysteinlyseryl-serine (P.sub.3CSS).
Moreover, an adjuvant such as a synthetic
cytosine-phosphorothiolated-guanine-containi- ng (CpG)
oligonucleotides has been found to increase CTL responses 10- to
100-fold. (see, e.g. Davila and Celis, J. Immunol. 165:539-547
(2000))
[0466] Upon immunization with a peptide composition in accordance
with the invention, via injection, aerosol, oral, transdermal,
transmucosal, intrapleural, intrathecal, or other suitable routes,
the immune system of the host responds to the vaccine by producing
large amounts of CTLs and/or HTLs specific for the desired antigen.
Consequently, the host becomes at least partially immune to later
development of cells that express or overexpress 191P4D12(b)
antigen, or derives at least some therapeutic benefit when the
antigen was tumor-associated.
[0467] In some embodiments, it may be desirable to combine the
class I peptide components with components that induce or
facilitate neutralizing antibody and or helper T cell responses
directed to the target antigen. A preferred embodiment of such a
composition comprises class I and class II epitopes in accordance
with the invention. An alternative embodiment of such a composition
comprises a class I and/or class II epitope in accordance with the
invention, along with a cross reactive HTL epitope such as
PADRE.TM. (Epimmune, San Diego, Calif.) molecule (described e.g.,
in U.S. Pat. No. 5,736,142).
[0468] A vaccine of the invention can also include
antigen-presenting cells (APC), such as dendritic cells (DC), as a
vehicle to present peptides of the invention. Vaccine compositions
can be created in vitro, following dendritic cell mobilization and
harvesting, whereby loading of dendritic cells occurs in vitro. For
example, dendritic cells are transfected, e.g., with a minigene in
accordance with the invention, or are pulsed with peptides. The
dendritic cell can then be administered to a patient to elicit
immune responses in vivo. Vaccine compositions, either DNA- or
peptide-based, can also be administered in vivo in combination with
dendritic cell mobilization whereby loading of dendritic cells
occurs in vivo.
[0469] Preferably, the following principles are utilized when
selecting an array of epitopes for inclusion in a polyepitopic
composition for use in a vaccine, or for selecting discrete
epitopes to be included in a vaccine and/or to be encoded by
nucleic acids such as a minigene. It is preferred that each of the
following principles be balanced in order to make the selection.
The multiple epitopes to be incorporated in a given vaccine
composition may be, but need not be, contiguous in sequence in the
native antigen from which the epitopes are derived.
[0470] 1.) Epitopes are selected which, upon administration, mimic
immune responses that have been observed to be correlated with
tumor clearance. For HLA Class I this includes 3-4 epitopes that
come from at least one tumor associated antigen (TAA). For HLA
Class II a similar rationale is employed; again 3-4 epitopes are
selected from at least one TAA (see, e.g., Rosenberg et al.,
Science 278:1447-1450). Epitopes from one TAA may be used in
combination with epitopes from one or more additional TAAs to
produce a vaccine that targets tumors with varying expression
patterns of frequently-expressed TAAs.
[0471] 2.) Epitopes are selected that have the requisite binding
affinity established to be correlated with immunogenicity: for HLA
Class I an IC.sub.50 of 500 nM or less, often 200 nM or less; and
for Class II an IC.sub.50 of 1000 nM or less.
[0472] 3.) Sufficient supermotif bearing-peptides, or a sufficient
array of allele-specific motif-bearing peptides, are selected to
give broad population coverage. For example, it is preferable to
have at least 80% population coverage. A Monte Carlo analysis, a
statistical evaluation known in the art, can be employed to assess
the breadth, or redundancy of, population coverage.
[0473] 4.) When selecting epitopes from cancer-related antigens it
is often useful to select analogs because the patient may have
developed tolerance to the native epitope.
[0474] 5.) Of particular relevance are epitopes referred to as
"nested epitopes." Nested epitopes occur where at least two
epitopes overlap in a given peptide sequence. A nested peptide
sequence can comprise B cell, HLA class I and/or HLA class II
epitopes. When providing nested epitopes, a general objective is to
provide the greatest number of epitopes per sequence. Thus, an
aspect is to avoid providing a peptide that is any longer than the
amino terminus of the amino terminal epitope and the carboxyl
terminus of the carboxyl terminal epitope in the peptide. When
providing a multi-epitopic sequence, such as a sequence comprising
nested epitopes, it is generally important to screen the sequence
in order to insure that it does not have pathological or other
deleterious biological properties.
[0475] 6.) If a polyepitopic protein is created, or when creating a
minigene, an objective is to generate the smallest peptide that
encompasses the epitopes of interest. This principle is similar, if
not the same as that employed when selecting a peptide comprising
nested epitopes. However, with an artificial polyepitopic peptide,
the size minimization objective is balanced against the need to
integrate any spacer sequences between epitopes in the polyepitopic
protein. Spacer amino acid residues can, for example, be introduced
to avoid junctional epitopes (an epitope recognized by the immune
system, not present in the target antigen, and only created by the
man-made juxtaposition of epitopes), or to facilitate cleavage
between epitopes and thereby enhance epitope presentation.
Junctional epitopes are generally to be avoided because the
recipient may generate an immune response to that non-native
epitope. Of particular concern is a junctional epitope that is a
"dominant epitope." A dominant epitope may lead to such a zealous
response that immune responses to other epitopes are diminished or
suppressed.
[0476] 7.) Where the sequences of multiple variants of the same
target protein are present, potential peptide epitopes can also be
selected on the basis of their conservancy. For example, a
criterion for conservancy may define that the entire sequence of an
HLA class I binding peptide or the entire 9-mer core of a class II
binding peptide be conserved in a designated percentage of the
sequences evaluated for a specific protein antigen.
[0477] X.C.1. Minigene Vaccines
[0478] A number of different approaches are available which allow
simultaneous delivery of multiple epitopes. Nucleic acids encoding
the peptides of the invention are a particularly useful embodiment
of the invention. Epitopes for inclusion in a minigene are
preferably selected according to the guidelines set forth in the
previous section. A preferred means of administering nucleic acids
encoding the peptides of the invention uses minigene constructs
encoding a peptide comprising one or multiple epitopes of the
invention.
[0479] The use of multi-epitope minigenes is described below and
in, Ishioka et al., J. Immunol. 162:3915-3925, 1999; An, L. and
Whitton, J. L., J. Virol. 71:2292, 1997; Thomson, S. A. et al., J.
Immunol. 157:822, 1996; Whitton, J. L. et al., J. Virol. 67:348,
1993; Hanke, R. et al., Vaccine 16:426, 1998. For example, a
multi-epitope DNA plasmid encoding supermotif- and/or motif-bearing
epitopes derived 191P4D12(b), the PADRE.RTM. universal helper T
cell epitope or multiple HTL epitopes from 191P4D12(b) (see e.g.,
Tables VIII-XXI and XXII to XLIX), and an endoplasmic
reticulum-translocating signal sequence can be engineered. A
vaccine may also comprise epitopes that are derived from other
TAAs.
[0480] The immunogenicity of a multi-epitopic minigene can be
confirmed in transgenic mice to evaluate the magnitude of CTL
induction responses against the epitopes tested. Further, the
immunogenicity of DNA-encoded epitopes in vivo can be correlated
with the in vitro responses of specific CTL lines against target
cells transfected with the DNA plasmid. Thus, these experiments can
show that the minigene serves to both: 1.) generate a CTL response
and 2.) that the induced CTLs recognized cells expressing the
encoded epitopes.
[0481] For example, to create a DNA sequence encoding the selected
epitopes (minigene) for expression in human cells, the amino acid
sequences of the epitopes may be reverse translated. A human codon
usage table can be used to guide the codon choice for each amino
acid. These epitope-encoding DNA sequences may be directly
adjoined, so that when translated, a continuous polypeptide
sequence is created. To optimize expression and/or immunogenicity,
additional elements can be incorporated into the minigene design.
Examples of amino acid sequences that can be reverse translated and
included in the minigene sequence include: HLA class I epitopes,
HLA class II epitopes, antibody epitopes, a ubiquitination signal
sequence, and/or an endoplasmic reticulum targeting signal. In
addition, HLA presentation of CTL and HTL epitopes may be improved
by including synthetic (e.g. poly-alanine) or naturally-occurring
flanking sequences adjacent to the CTL or HTL epitopes; these
larger peptides comprising the epitope(s) are within the scope of
the invention.
[0482] The minigene sequence may be converted to DNA by assembling
oligonucleotides that encode the plus and minus strands of the
minigene. Overlapping oligonucleotides (30-100 bases long) may be
synthesized, phosphorylated, purified and annealed under
appropriate conditions using well known techniques. The ends of the
oligonucleotides can be joined, for example, using T4 DNA ligase.
This synthetic minigene, encoding the epitope polypeptide, can then
be cloned into a desired expression vector.
[0483] Standard regulatory sequences well known to those of skill
in the art are preferably included in the vector to ensure
expression in the target cells. Several vector elements are
desirable: a promoter with a down-stream cloning site for minigene
insertion; a polyadenylation signal for efficient transcription
termination; an E. coli origin of replication; and an E. coli
selectable marker (e.g. ampicillin or kanamycin resistance).
Numerous promoters can be used for this purpose, e.g., the human
cytomegalovirus (hCMV) promoter. See, e.g., U.S. Pat. Nos.
5,580,859 and 5,589,466 for other suitable promoter sequences.
[0484] Additional vector modifications may be desired to optimize
minigene expression and immunogenicity. In some cases, introns are
required for efficient gene expression, and one or more synthetic
or naturally-occurring introns could be incorporated into the
transcribed region of the minigene. The inclusion of mRNA
stabilization sequences and sequences for replication in mammalian
cells may also be considered for increasing minigene
expression.
[0485] Once an expression vector is selected, the minigene is
cloned into the polylinker region downstream of the promoter. This
plasmid is transformed into an appropriate E. coli strain, and DNA
is prepared using standard techniques. The orientation and DNA
sequence of the minigene, as well as all other elements included in
the vector, are confirmed using restriction mapping and DNA
sequence analysis. Bacterial cells harboring the correct plasmid
can be stored as a master cell bank and a working cell bank.
[0486] In addition, immunostimulatory sequences (ISSs or CpGs)
appear to play a role in the immunogenicity of DNA vaccines. These
sequences may be included in the vector, outside the minigene
coding sequence, if desired to enhance immunogenicity.
[0487] In some embodiments, a bi-cistronic expression vector which
allows production of both the minigene-encoded epitopes and a
second protein (included to enhance or decrease immunogenicity) can
be used. Examples of proteins or polypeptides that could
beneficially enhance the immune response if co-expressed include
cytokines (e.g., IL-2, IL-12, GM-CSF), cytokine-inducing molecules
(e.g., LeIF), costimulatory molecules, or for HTL responses, pan-DR
binding proteins (PADRE.TM., Epimmune, San Diego, Calif.). Helper
(HTL) epitopes can be joined to intracellular targeting signals and
expressed separately from expressed CTL epitopes; this allows
direction of the HTL epitopes to a cell compartment different than
that of the CTL epitopes. If required, this could facilitate more
efficient entry of HTL epitopes into the HLA class II pathway,
thereby improving HTL induction. In contrast to HTL or CTL
induction, specifically decreasing the immune response by
co-expression of immunosuppressive molecules (e.g. TGF-.beta.) may
be beneficial in certain diseases.
[0488] Therapeutic quantities of plasmid DNA can be produced for
example, by fermentation in E. coli, followed by purification.
Aliquots from the working cell bank are used to inoculate growth
medium, and grown to saturation in shaker flasks or a bioreactor
according to well-known techniques. Plasmid DNA can be purified
using standard bioseparation technologies such as solid phase
anion-exchange resins supplied by QIAGEN, Inc. (Valencia, Calif.).
If required, supercoiled DNA can be isolated from the open circular
and linear forms using gel electrophoresis or other methods.
[0489] Purified plasmid DNA can be prepared for injection using a
variety of formulations. The simplest of these is reconstitution of
lyophilized DNA in sterile phosphate-buffer saline (PBS). This
approach, known as "naked DNA," is currently being used for
intramuscular (IM) administration in clinical trials. To maximize
the immunotherapeutic effects of minigene DNA vaccines, an
alternative method for formulating purified plasmid DNA may be
desirable. A variety of methods have been described, and new
techniques may become available. Gationic lipids, glycolipids, and
fusogenic liposomes can also be used in the formulation (see, e.g.,
as described by WO 93/24640; Mannino & Gould-Fogerite, Bio
Techniques 6(7): 682 (1988); U.S. Pat No. 5,279,833; WO 91/06309;
and Feigner, et al., Proc. Nat'l Acad. Sci. USA 84:7413 (1987). In
addition, peptides and compounds referred to collectively as
protective, interactive, non-condensing compounds (PINC) could also
be complexed to purified plasmid DNA to influence variables such as
stability, intramuscular dispersion, or trafficking to specific
organs or cell types.
[0490] Target cell sensitization can be used as a functional assay
for expression and HLA class I presentation of minigene-encoded CTL
epitopes. For example, the plasmid DNA is introduced into a
mammalian cell line that is suitable as a target for standard CTL
chromium release assays. The transfection method used will be
dependent on the final formulation. Electroporation can be used for
"naked" DNA, whereas cationic lipids allow direct in vitro
transfection. A plasmid expressing green fluorescent protein (GFP)
can be co-transfected to allow enrichment of transfected cells
using fluorescence activated cell sorting (FACS). These cells are
then chromium-51 (.sup.51Cr) labeled and used as target cells for
epitope-specific CTL lines; cytolysis, detected by .sup.51Cr
release, indicates both production of, and HLA presentation of,
minigene-encoded CTL epitopes. Expression of HTL epitopes may be
evaluated in an analogous manner using assays to assess HTL
activity.
[0491] In vivo immunogenicity is a seconds approach for functional
testing of minigene DNA formulations. Transgenic mice expressing
appropriate human HLA proteins are immunized with the DNA product.
The dose and route of administration are formulation dependent
(e.g., IM for DNA in PBS, intraperitoneal (i.p.) for
lipid-complexed DNA). Twenty-one days after immunization,
splenocytes are harvested and restimulated for one week in the
presence of peptides encoding each epitope being tested.
Thereafter, for CTL effector cells, assays are conducted for
cytolysis of peptide-loaded, .sup.51Cr-labeled target cells using
standard techniques. Lysis of target cells that were sensitized by
HLA loaded with peptide epitopes, corresponding to minigene-encoded
epitopes, demonstrates DNA vaccine function for in vivo induction
of CTLs. Immunogenicity of HTL epitopes is confirmed in transgenic
mice in an analogous manner.
[0492] Alternatively, the nucleic acids can be administered using
ballistic delivery as described, for instance, in U.S. Pat. No.
5,204,253. Using this technique, particles comprised solely of DNA
are administered. In a further alternative embodiment, DNA can be
adhered to particles, such as gold particles.
[0493] Minigenes can also be delivered using other bacterial or
viral delivery systems well known in the art, e.g., an expression
construct encoding epitopes of the invention can be incorporated
into a viral vector such as vaccinia.
[0494] X.C.2. Combinations of CTL Peptides with Helper Peptides
[0495] Vaccine compositions comprising CTL peptides of the
invention can be modified, e.g., analoged, to provide desired
attributes, such as improved serum half life, broadened population
coverage or enhanced immunogenicity.
[0496] For instance, the ability of a peptide to induce CTL
activity can be enhanced by linking the peptide to a sequence which
contains at least one epitope that is capable of inducing a T
helper cell response. Although a CTL peptide can be directly linked
to a T helper peptide, often CTL epitope/HTL epitope conjugates are
linked by a spacer molecule. The spacer is typically comprised of
relatively small, neutral molecules, such as amino acids or amino
acid mimetics, which are substantially uncharged under
physiological conditions. The spacers are typically selected from,
e.g., Ala, Gly, or other neutral spacers of nonpolar amino acids or
neutral polar amino acids. It will be understood that the
optionally present spacer need not be comprised of the same
residues and thus may be a hetero- or homo-oligomer. When present,
the spacer will usually be at least one or two residues, more
usually three to six residues and sometimes 10 or more residues.
The CTL peptide epitope can be linked to the T helper peptide
epitope either directly or via a spacer either at the amino or
carboxy terminus of the CTL peptide. The amino terminus of either
the immunogenic peptide or the T helper peptide may be
acylated.
[0497] In certain embodiments, the T helper peptide is one that is
recognized by T helper cells present in a majority of a genetically
diverse population. This can be accomplished by selecting peptides
that bind to many, most, or all of the HLA class II molecules.
Examples of such amino acid bind many HLA Class II molecules
include sequences from antigens such as tetanus toxoid at positions
830-843 (QYIKANSKFIGITE; SEQ ID NO: 44), Plasmodium falciparum
circumsporozoite (CS) protein at positions 378-398
(DIEKKIAKMEKASSVFNVVNS; SEQ ID NO: 45), and Streptococcus 18 kD
protein at positions 116-131 (GAVDSILGGVATYGAA; SEQ ID NO. 46).
Other examples include peptides bearing a DR 1-4-7 supermotif, or
either of the DR3 motifs.
[0498] Alternatively, it is possible to prepare synthetic peptides
capable of stimulating T helper lymphocytes, in a loosely
HLA-restricted fashion, using amino acid sequences not found in
nature (see, e.g., PCT publication WO 95/07707). These synthetic
compounds called Pan-DR-binding epitopes (e.g., PADRE.TM.,
Epimmune, Inc., San Diego, Calif.) are designed, most preferably,
to bind most HLA-DR (human HLA class II) molecules. For instance, a
pan-DR-binding epitope peptide having the formula: XKXVAAWTLKAX
(SEQ ID NO: 47), where "X" is either cycldhexylalanine,
phenylalanine, or tyrosine, and a is either D-alanine or L-alanine,
has been found to bind to most HLA-DR alleles, and to stimulate the
response of T helper lymphocytes from most individuals, regardless
of their HLA type. An alternative of a pan-DR binding epitope
comprises all "L" natural amino acids and can be provided in the
form of nucleic acids that encode the epitope.
[0499] HTL peptide epitopes can also be modified to alter their
biological properties. For example, they can be modified to include
D-amino acids to increase their resistance to proteases and thus
extend their serum half life, or they can be conjugated to other
molecules such as lipids, proteins, carbohydrates, and the like to
increase their biological activity. For example, a T helper peptide
can be conjugated to one or more palmitic acid chains at either the
amino or carboxyl termini.
[0500] X.C.3. Combinations of CTL Peptides with T Cell Priming
Agents
[0501] In some embodiments it may be desirable to include in the
pharmaceutical compositions of the invention at least one component
which primes B lymphocytes or T lymphocytes. Lipids have been
identified as agents capable of priming CTL in vivo. For example,
palmitic acid residues can be attached to the .epsilon.- and
.alpha.-amino groups of a lysine residue and then linked, e.g., via
one or more linking residues such as Gly, Gly-Gly-, Ser, Ser-Ser,
or the like, to an immunogenic peptide. The lipidated peptide can
then be administered either directly in a micelle or particle,
incorporated into a liposome, or emulsified in an adjuvant, e.g.,
incomplete Freund's adjuvant. In a preferred embodiment, a
particularly effective immunogenic composition comprises palmitic
acid attached to .epsilon.- and .alpha.-amino groups of Lys, which
is attached via linkage, e.g., Ser-Ser, to the amino terminus of
the immunogenic peptide.
[0502] As another example of lipid priming of CTL responses, E.
coli lipoproteins, such as
tripalmitoyl-S-glycerylcysteinlyseryl-serine (P.sub.3CSS) can be
used to prime virus specific CTL when covalently attached to an
appropriate peptide (see, e.g., Deres, et al., Nature 342:561,
1989). Peptides of the invention can be coupled to P.sub.3CSS, for
example, and the lipopeptide administered to an individual to prime
specifically an immune response to the target antigen. Moreover,
because the induction of neutralizing antibodies can also be primed
with P.sub.3CSS-conjugated epitopes, two such compositions can be
combined to more effectively elicit both humoral and cell-mediated
responses.
[0503] X.C.4. Vaccine Compositions Comprising DC Pulsed with CTL
and/or HTL Peptides
[0504] An embodiment of a vaccine composition in accordance with
the invention comprises ex vivo administration of a cocktail of
epitope-bearing peptides to PBMC, or isolated DC therefrom, from
the patient's blood. A pharmaceutical to facilitate harvesting of
DC can be used, such as Progenipoietin.TM. (Pharmacia-Monsanto, St.
Louis, Mo.) or GM-CSF/IL-4. After pulsing the DC with peptides and
prior to reinfusion into patients, the DC are washed to remove
unbound peptides. In this embodiment, a vaccine comprises
peptide-pulsed DCs which present the pulsed peptide epitopes
complexed with HLA molecules on their surfaces.
[0505] The DC can be pulsed ex vivo with a cocktail of peptides,
some of which stimulate CTL responses to 191P4D12(b). Optionally, a
helper T cell (HTL) peptide, such as a natural or artificial
loosely restricted HLA Class II peptide, can be included to
facilitate the CTL response. Thus, a vaccine in accordance with the
invention is used to treat a cancer which expresses or
overexpresses 191P4D12(b).
[0506] X.D. Adoptive Immunotherapy
[0507] Antigenic 191P4D12(b)-related peptides are used to elicit a
CTL and/or HTL response ex vivo, as well. The resulting CTL or HTL
cells, can be used to treat tumors in patients that do not respond
to other conventional forms of therapy, or will not respond to a
therapeutic vaccine peptide or nucleic acid in accordance with the
invention. Ex vivo CTL or HTL responses to a particular antigen are
induced by incubating in tissue culture the patient's, or
genetically compatible, CTL or HTL precursor cells together with a
source of antigen-presenting cells (APC), such as dendritic cells,
and the appropriate immunogenic peptide. After an appropriate
incubation time (typically about 7-28 days), in which the precursor
cells are activated and expanded into effector cells, the cells are
infused back into the patient, where they will destroy (CTL) or
facilitate destruction (HTL) of their specific target cell (e.g., a
tumor cell). Transfected dendritic cells may also be used as
antigen presenting cells.
[0508] X.E. Administration of Vaccines for Therapeutic or
Prophylactic Purposes
[0509] Pharmaceutical and vaccine compositions of the invention are
typically used to treat and/or prevent a cancer that expresses or
overexpresses 191P4D12(b). In therapeutic applications, peptide
and/or nucleic acid compositions are administered to a patient in
an amount sufficient to elicit an effective B cell, CTL and/or HTL
response to the antigen and to cure or at least partially arrest or
slow symptoms and/or complications. An amount adequate to
accomplish this is defined as "therapeutically effective dose."
Amounts effective for this use will depend on, e.g., the particular
composition administered, the manner of administration, the stage
and severity of the disease being treated, the weight and general
state of health of the patient, and the judgment of the prescribing
physician.
[0510] For pharmaceutical compositions, the immunogenic peptides of
the invention, or DNA encoding them, are generally administered to
an individual already bearing a tumor that expresses 191P4D12(b).
The peptides or DNA encoding them can be administered individually
or as fusions of one or more peptide sequences. Patients can be
treated with the immunogenic peptides separately or in conjunction
with other treatments, such as surgery, as appropriate.
[0511] For therapeutic use, administration should generally begin
at the first diagnosis of 191P4D12(b)-associated cancer. This is
followed by boosting doses until at least symptoms are
substantially abated and for a period thereafter. The embodiment of
the vaccine composition (ie., including, but not limited to
embodiments such as peptide cocktails, polyepitopic polypeptides,
minigenes, or TAA-specific CTLs or pulsed dendritic cells)
delivered to the patient may vary according to the stage of the
disease or the patient's health status. For example, in a patient
with a tumor that expresses 191P4D12(b), a vaccine comprising
191P4D12(b)-specific CTL may be more efficacious in killing tumor
cells in patient with advanced disease than alternative
embodiments.
[0512] It is generally important to provide an amount of the
peptide epitope delivered by a mode of administration sufficient to
stimulate effectively a cytotoxic T cell response; compositions
which stimulate helper T cell responses can also be given in
accordance with this embodiment of the invention.
[0513] The dosage for an initial therapeutic immunization generally
occurs in a unit dosage range where the lower value is about 1, 5,
50, 500, or 1,000 .mu.g and the higher value is about 10,000;
20,000; 30,000; or 50,000 .mu.g. Dosage values for a human
typically range from about 500 .mu.g to about 50,000 .mu.g per 70
kilogram patient. Boosting dosages of between about 1.0 .mu.g to
about 50,000 .mu.g of peptide pursuant to a boosting regimen over
weeks to months may be administered depending upon the patient's
response and condition as determined by measuring the specific
activity of CTL and HTL obtained from the patient's blood.
Administration should continue until at least clinical symptoms or
laboratory tests indicate that the neoplasia, has been eliminated
or reduced and for a period thereafter. The dosages, routes of
administration, and dose schedules are adjusted in accordance with
methodologies known in the art.
[0514] In certain embodiments, the peptides and compositions of the
present invention are employed in serious disease states, that is,
life-threatening or potentially life threatening situations. In
such cases, as a result of the minimal amounts of extraneous
substances and the relative nontoxic nature of the peptides in
preferred compositions of the invention, it is possible and may be
felt desirable by the treating physician to administer substantial
excesses of these peptide compositions relative to these stated
dosage amounts.
[0515] The vaccine compositions of the invention can also be used
purely as prophylactic agents. Generally the dosage for an initial
prophylactic immunization generally occurs in a unit dosage range
where the lower value is about 1, 5, 50, 500, or 1000 .mu.g and the
higher value is about 10,000; 20,000; 30,000; or 50,000 .mu.g.
Dosage values for a human typically range from about 500 .mu.g to
about 50,000 .mu.g per 70 kilogram patient. This is followed by
boosting dosages of between about 1.0 .mu.g to about 50,000 .mu.g
of peptide administered at defined intervals from about four weeks
to six months after the initial administration of vaccine. The
immunogenicity of the vaccine can be assessed by measuring the
specific activity of CTL and HTL obtained from a sample of the
patient's blood.
[0516] The pharmaceutical compositions for therapeutic treatment
are intended for parenteral, topical, oral, nasal, intrathecal, or
local (e.g. as a cream or topical ointment) administration.
Preferably, the pharmaceutical compositions are administered
parentally, e.g., intravenously, subcutaneously, intradermally, or
intramuscularly. Thus, the invention provides compositions for
parenteral administration which comprise a solution of the
immunogenic peptides dissolved or suspended in an acceptable
carrier, preferably an aqueous carrier.
[0517] A variety of aqueous carriers may be used, e.g., water,
buffered water, 0.8% saline, 0.3% glycine, hyaluronic acid and the
like. These compositions may be sterilized by conventional,
well-known sterilization techniques, or may be sterile filtered.
The resulting aqueous solutions may be packaged for use as is, or
lyophilized, the lyophilized preparation being combined with a
sterile solution prior to administration.
[0518] The compositions may contain pharmaceutically acceptable
auxiliary substances as required to approximate physiological
conditions, such as pH-adjusting and buffering agents, tonicity
adjusting agents, wetting agents, preservatives, and the like, for
example, sodium acetate, sodium lactate, sodium chloride, potassium
chloride, calcium chloride, sorbitan monolaurate, triethanolamine
oleate, etc.
[0519] The concentration of peptides of the invention in the
pharmaceutical formulations can vary widely, i.e., from less than
about 0.1%, usually at or at least about 2% to as much as 20% to
50% or more by weight, and will be selected primarily by fluid
volumes, viscosities, etc., in accordance with the particular mode
of administration selected.
[0520] A human unit dose form of a composition is typically
included in a pharmaceutical composition that comprises a human
unit dose of an acceptable carrier, in one embodiment an aqueous
carrier, and is administered in a volume/quantity that is known by
those of skill in the art to be used for administration of suchf
compositions to humans (see, e.g., Remington's Pharmaceutical
Sciences, 17.sup.th Edition, A. Gennaro, Editor, Mack Publishing
Co., Easton, Pa., 1985). For example a peptide dose for initial
immunization can be from about 1 to about 50,000 .mu.g, generally
100-5,000 .mu.g, for a 70 kg patient. For example, for nucleic
acids an initial immunization may be performed using an expression
vector in the form of naked nucleic acid administered IM (or SC or
ID) in the amounts of 0.5-5 mg at multiple sites. The nucleic acid
(0.1 to 1000 .mu.g) can also be administered using a gene gun.
Following an incubation period of 3-4 weeks, a booster dose is then
administered. The booster can be recombinant fowlpox virus
administered at a dose of 5-10.sup.7 to 5.times.10.sup.9 pfu.
[0521] For antibodies, a treatment generally involves repeated
administration of the anti-191P4D12(b) antibody preparation, via an
acceptable route of administration such as intravenous injection
(IV), typically at a dose in the range of about 0.1 to about 10
mg/kg body weight. In general, doses in the range of 10-500 mg mAb
per week are effective and well tolerated. Moreover, an initial
loading dose of approximately 4 mg/kg patient body weight IV,
followed by weekly doses of about 2 mg/kg IV of the
anti-191P4D12(b) mAb preparation represents an acceptable dosing
regimen. As appreciated by those of skill in the art, various
factors can influence the ideal dose in a particular case. Such
factors include, for example, half life of a composition, the
binding affinity of an Ab, the immunogenicity of a substance, the
degree of 191P4D12(b) expression in the patient, the extent of
circulating shed 191P4D12(b) antigen, the desired steady-state
concentration level, frequency of treatment, and the influence of
chemotherapeutic or other agents used in combination with the
treatment method of the invention, as well as the health status of
a particular patient. Non-limiting preferred human unit doses are,
for example, 500 .mu.g-1 mg, 1 mg-50 mg, 50 mg-100 mg, 100 mg-200
mg, 200 mg-300 mg, 400 mg-500 mg, 500 mg-600 mg, 600 mg-700 mg, 700
mg-800 mg, 800 mg-900 mg, 900 mg-1 g, or 1 mg-700 mg. In certain
embodiments, the dose is in a range of 2-5 mg/kg body weight, e.g.,
with follow on weekly doses of 1-3 mg/kg; 0.5 mg, 1, 2, 3, 4, 5, 6,
7, 8, 9, 10 mg/kg body weight followed, e.g., in two, three or four
weeks by weekly doses; 0.5-10 mg/kg body weight, e.g., followed in
two, three or four weeks by weekly doses; 225, 250, 275, 300, 325,
350, 375, 400 mg m.sup.2 of body area weekly; 1-600 mg m.sup.2 of
body area weekly; 225-400 mg m.sup.2 of body area weekly; these
does can be followed by weekly doses for 2, 3, 4, 5, 6, 7, 8, 9,
19, 11, 12 or more weeks.
[0522] In one embodiment, human unit dose forms of polynucleotides
comprise a suitable dosage range or effective amount that provides
any therapeutic effect. As appreciated by one of ordinary skill in
the art a therapeutic effect depends on a number of factors,
including the sequence of the polynucleotide, molecular weight of
the polynucleotide and route of administration. Dosages are
generally selected by the physician or other health care
professional in accordance with a variety of parameters known in
the art, such as severity of symptoms, history of the patient and
the like. Generally, for a polynucleotide of about 20 bases, a
dosage range may be selected from, for example, an independently
selected lower limit such as about 0.1, 0.25, 0.5, 1, 2, 5, 10, 20,
30, 40, 50, 60, 70, 80, 90, 100, 200, 300, 400 or 500 mg/kg up to
an independently selected upper limit, greater than the lower
limit, of about 60, 80, 100, 200, 300, 400, 500, 750, 1000, 1500,
2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000 or 10,000 mg/kg. For
example, a dose may be about any of the following: 0.1 to 100
mg/kg, 0.1 to 50 mg/kg, 0.1 to 25 mg/kg, 0.1 to 10 mg/kg, 1 to 500
mg/kg, 100 to 400 mg/kg, 200 to 300 mg/kg, 1 to 100 mg/kg, 100 to
200 mg/kg, 300 to 400 mg/kg, 400 to 500 mg/kg, 500 to 1000 mg/kg,
500 to 5000 mg/kg, or 500 to 10,000 mg/kg. Generally, parenteral
routes of administration may require higher doses of polynucleotide
compared to more direct application to the nucleotide to diseased
tissue, as do polynucleotides of increasing length.
[0523] In one embodiment, human unit dose forms of T-cells comprise
a suitable dosage range or effective amount that provides any
therapeutic effect. As appreciated by one of ordinary skill in the
art, a therapeutic effect depends on a number of factors. Dosages
are generally selected by the physician or other health care
professional in accordance with a variety of parameters known in
the art, such as severity of symptoms, history of the patient and
the like. A dose may be about 10.sup.4 cells to about 10.sup.6
cells, about 10.sup.6 cells to about 10.sup.8 cells, about 10.sup.8
to about 10.sup.11 cells, or about 10.sup.8 to about
5.times.10.sup.10 cells. A dose may also about 10.sup.6
cells/m.sup.2 to about 10.sup.10 cells/m.sup.2, or about 10.sup.6
cells/m.sup.2 to about 10.sup.8 cells/m.sup.2 .
[0524] Proteins(s) of the invention, and/or nucleic acids encoding
the protein(s), can also be administered via liposomes, which may
also serve to: 1) target the proteins(s) to a particular tissue,
such as lymphoid tissue; 2) to target selectively to diseases
cells; or, 3) to increase the half-life of the peptide composition.
Liposomes include emulsions, foams, micelles, insoluble monolayers,
liquid crystals, phospholipid dispersions, lamellar layers and the
like. In these preparations, the peptide to be delivered is
incorporated as part of a liposome, alone or in conjunction with a
molecule which binds to a receptor prevalent among lymphoid cells,
such as monoclonal antibodies which bind to the CD45 antigen, or
with other therapeutic or immunogenic compositions. Thus, liposomes
either filled or decorated with a desired peptide of the invention
can be directed to the site of lymphoid cells, where the liposomes
then deliver the peptide compositions. Liposomes for use in
accordance with the invention are formed from standard
vesicle-forming lipids, which generally include neutral and
negatively charged phospholipids and a sterol, such as cholesterol.
The selection of lipids is generally guided by consideration of,
e.g., liposome size, acid lability and stability of the liposomes
in the blood stream. A variety of methods are available for
preparing liposomes, as described in, e.g., Szoka, et al., Ann.
Rev. Biophys. Bioeng. 9:467 (1980), and U.S. Pat. Nos. 4,235,871,
4,501,728, 4,837,028, and 5,019,369.
[0525] For targeting cells of the immune system, a ligand to be
incorporated into the liposome can include, e.g., antibodies or
fragments thereof specific for cell surface determinants of the
desired immune system cells. A liposome suspension containing a
peptide may be administered intravenously, locally, topically, etc.
in a dose which varies according to, inter alia, the manner of
administration, the peptide being delivered, and the stage of the
disease being treated.
[0526] For solid compositions, conventional nontoxic solid carriers
may be used which include, for example, pharmaceutical grades of
mannitol, lactose, starch, magnesium stearate, sodium saccharin,
talcum, cellulose, glucose, sucrose, magnesium carbonate, and the
like. For oral administration, a pharmaceutically acceptable
nontoxic composition is formed by incorporating any of the normally
employed excipients, such as those carriers previously listed, and
generally 10-95% of active ingredient, that is, one or more
peptides of the invention, and more preferably at a concentration
of 25%-75%.
[0527] For aerosol administration, immunogenic peptides are
preferably supplied in finely divided form along with a surfactant
and propellant. Typical percentages of peptides are about 0.01%-20%
by weight, preferably about 1%-10%. The surfactant must, of course,
be nontoxic, and preferably soluble in the propellant.
Representative of such agents are the esters or partial esters of
fatty acids containing from about 6 to 22 carbon atoms, such as
caproic, octanoic, lauric, palmitic, stearic, linoleic, linolenic,
olesteric and oleic acids with an aliphatic polyhydric alcohol or
its cyclic anhydride. Mixed esters, such as mixed or natural
glycerides may be employed. The surfactant may constitute about
0.1%-20% by weight of the composition, preferably about 0.25-5%.
The balance of the composition is ordinarily propellant. A carrier
can also be included, as desired, as with, e.g., lecithin for
intranasal delivery.
[0528] XI.) Diagnostic and Prognostic Embodiments of
191P4D12(b).
[0529] As disclosed herein, 191P4D12(b) polynucleotides,
polypeptides, reactive cytotoxic T cells (CTL), reactive helper T
cells (HTL) and anti-polypeptide antibodies are used in well known
diagnostic, prognostic and therapeutic assays that examine
conditions associated with dysregulated cell growth such as cancer,
in particular the cancers listed in Table I (see, e.g., both its
specific pattern of tissue expression as well as its overexpression
in certain cancers as described for example in the Example entitled
"Expression analysis of 191P4D12(b) in normal tissues, and patient
specimens").
[0530] 191P4D12(b) can be analogized to a prostate associated
antigen PSA, the archetypal marker that has been used by medical
practitioners for years to identify and monitor the presence of
prostate cancer (see, e.g., Merrill et al., J. Urol. 163(2):
503-5120 (2000); Polascik et al., J. Urol. Aug; 162(2):293-306
(1999) and Fortier et al., J. Nat. Cancer Inst. 91(19):
1635-1640(1999)). A variety of other diagnostic markers are also
used in similar contexts including p53 and K-ras (see, e.g.,
Tulchinsky et al., Int J Mol Med 1999 Jul 4(1):99-102 and Minimoto
et al., Cancer Detect Prev 2000;24(1):1-12). Therefore, this
disclosure of 191P4D12(b) polynucleotides and polypeptides (as well
as 191P4D12(b) polynucleotide probes and anti-191P4D12(b)
antibodies used to identify the presence of these molecules) and
their properties allows skilled artisans to utilize these molecules
in methods that are analogous to those used, for example, in a
variety of diagnostic assays directed to examining conditions
associated with cancer.
[0531] Typical embodiments of diagnostic methods which utilize the
191P4D12(b) polynucleotides, polypeptides, reactive T cells and
antibodies are analogous to those methods from well-established
diagnostic assays, which employ, e.g., PSA polynucleotides,
polypeptides, reactive T cells and antibodies. For example, just as
PSA polynucleotides are used as probes (for example in Northern
analysis, see, e.g., Sharief et al., Biochem. Mol. Biol. Int.
33(3):567-74(1994)) and primers (for example in PCR analysis, see,
e.g., Okegawa et al., J. Urol. 163(4): 1189-1190 (2000)) to observe
the presence and/or the level of PSA mRNAs in methods of monitoring
PSA overexpression or the metastasis of prostate cancers, the
191P4D12(b) polynucleotides described herein can be utilized in the
same way to detect 191P4D12(b) overexpression or the metastasis of
prostate and other cancers expressing this gene. Alternatively,
just as PSA polypeptides are used to generate antibodies specific
for PSA which can then be used to observe the presence and/or the
level of PSA proteins in methods to monitor PSA protein
overexpression (see, e.g., Stephan et al., Urology 55(4):560-3
(2000)) or the metastasis of prostate cells (see, e.g., Alanen et
al., Pathol. Res. Pract. 192(3):233-7 (1996)), the 191P4D12(b)
polypeptides described herein can be utilized to generate
antibodies for use in detecting 191P4D12(b) overexpression or the
metastasis of prostate cells and cells of other cancers expressing
this gene.
[0532] Specifically, because metastases involves the movement of
cancer cells from an organ of origin (such as the lung or prostate
gland etc.) to a different area of the body (such as a lymph node),
assays which examine a biological sample for the presence of cells
expressing 191P4D12(b) polynucleotides and/or polypeptides can be
used to provide evidence of metastasis. For example, when a
biological sample from tissue that does not normally contain
191P4D12(b)-expressing cells (lymph node) is found to contain
191P4D12(b)-expressing cells such as the 191P4D12(b) expression
seen in LAPC4 and LAPC9, xenografts isolated from lymph node and
bone metastasis, respectively, this finding is indicative of
metastasis.
[0533] Alternatively 191P4D12(b) polynucleotides and/or
polypeptides can be used to provide evidence of cancer, for
example, when cells in a biological sample that do not normally
express 191P4D12(b) or express 191P4D12(b) at a different level are
found to express 191P4D12(b) or have an increased expression of
191P4D12(b) (see, e.g., the 191P4D12(b) expression in the cancers
listed in Table I and in patient samples etc. shown in the
accompanying Figures). In such assays, artisans may further wish to
generate supplementary evidence of metastasis by testing the
biological sample for the presence of a second tissue restricted
marker (in addition to 191P4D12(b)) such as PSA, PSCA etc. (see,
e.g., Alanen et al., Pathol. Res. Pract. 192(3): 233-237
(1996)).
[0534] The use of immunohistochemistry to identify the presence of
a 191P4D12(b) polypeptide within a tissue section can indicate an
altered state of certain cells within that tissue. It is well
understood in the art that the ability of an antibody to localize
to a polypeptide that is expressed in cancer cells is a way of
diagnosing presence of disease, disease stage, progression and/or
tumor aggressiveness. Such an antibody can also detect an altered
distribution of the polypeptide within the cancer cells, as
compared to corresponding non-malignant tissue.
[0535] The 191P4D12(b) polypeptide and immunogenic compositions are
also useful in view of the phenomena of altered subcellular protein
localization in disease states. Alteration of cells from normal to
diseased state causes changes in cellular morphology and is often
associated with changes in subcellular protein
localization/distribution. For example, cell membrane proteins that
are expressed in a polarized manner in normal cells can be altered
in disease, resulting in distribution of the protein in a non-polar
manner over the whole cell surface.
[0536] The phenomenon of altered subcellular protein localization
in a disease state has been demonstrated with MUC1 and Her2 protein
expression by use of immunohistochemical means. Normal epithelial
cells have a typical apical distribution of MUC1, in addition to
some supranuclear localization of the glycoprotein, whereas
malignant lesions often demonstrate an apolar staining pattern
(Diaz et al, The Breast Journal, 7; 40-45 (2001); Zhang et al,
Clinical Cancer Research, 4; 2669-2676 (1998): Cao, et al, The
Journal of Histochemistry and Cytochemistry, 45: 1547-1557 (1997)).
In addition, normal breast epithelium is either negative for Her2
protein or exhibits only a basolateral distribution whereas
malignant cells can express the protein over the whole cell surface
(De Potter, et al, International Journal of Cancer, 44; 969-974
(1989): McCormick, et al, 117; 935-943 (2002)). Alternatively,
distribution of the protein may be altered from a surface only
localization to include diffuse cytoplasmic expression in the
diseased state. Such an example can be seen with MUC1 (Diaz, et al,
The Breast Journal, 7: 40-45 (2001)).
[0537] Alteration in the localization/distribution of a protein in
the cell, as detected by immunohistochemical methods, can also
provide valuable information concerning the favorability of certain
treatment modalities. This last point is illustrated by a situation
where a protein may be intracellular in normal tissue, but cell
surface in malignant cells; the cell surface location makes the
cells favorably amenable to antibody-based diagnostic and treatment
regimens. When such an alteration of protein localization occurs
for 191P4D12(b), the 191P4D12(b) protein and immune responses
related thereto are very useful. Accordingly, the ability to
determine whether alteration of subcellular protein localization
occurred for 24P4C12 make the 191P4D12(b) protein and immune
responses related thereto very useful. Use of the 191P4D12(b)
compositions allows those skilled in the art to make important
diagnostic and therapeutic decisions. Immunohistochemical reagents
specific to 191P4D12(b) are also useful to detect metastases of
tumors expressing 191P4D12(b) when the polypeptide appears in
tissues where 191P4D12(b) is not normally produced.
[0538] Thus, 191P4D12(b) polypeptides and antibodies resulting from
immune responses thereto are useful in a variety of important
contexts such as diagnostic, prognostic, preventative and/or
therapeutic purposes known to those skilled in the art.
[0539] Just as PSA polynucleotide fragments and polynucleotide
variants are employed by skilled artisans for use in methods of
monitoring PSA, 191P4D12(b) polynucleotide fragments and
polynucleotide variants are used in an analogous manner. In
particular, typical PSA polynucleotides used in methods of
monitoring PSA are probes or primers which consist of fragments of
the PSA cDNA sequence. Illustrating this, primers used to PCR
amplify a PSA polynucleotide must include less than the whole PSA
sequence to function in the polymerase chain reaction. In the
context of such PCR reactions, skilled artisans generally create a
variety of different polynucleotide fragments that can be used as
primers in order to amplify different portions of a polynucleotide
of interest or to optimize amplification reactions (see, e.g.,
Caetano-Anolles, G. Biotechniques 25(3): 472-476, 478-480 (1998);
Robertson et al., Methods Mol. Biol. 98:121-154 (1998)). An
additional illustration of the use of such fragments is provided in
the Example entitled "Expression analysis of 191P4D12(b) in normal
tissues, and patient specimens," where a 191P4D12(b) polynucleotide
fragment is used as a probe to show the expression of 191P4D12(b)
RNAs in cancer cells. In addition, variant polynucleotide sequences
are typically used as primers and probes for the corresponding
mRNAs in PCR and Northern analyses (see, e.g., Sawai et al., Fetal
Diagn. Ther. 1996 Nov-Dec 11(6):407-13 and Current Protocols In
Molecular Biology, Volume 2, Unit 2, Frederick M. Ausubel et al.
eds., 1995)). Polynucleotide fragments and variants are useful in
this context where they are capable of binding to a target
polynucleotide sequence (e.g., a 191P4D12(b) polynucleotide shown
in FIG. 2 or variant thereof) under conditions of high
stringency.
[0540] Furthermore, PSA polypeptides which contain an epitope that
can be recognized by an antibody or T cell that specifically binds
to that epitope are used in methods of monitoring PSA. 191P4D12(b)
polypeptide fragments and polypeptide analogs or variants can also
be used in an analogous manner. This practice of using polypeptide
fragments or polypeptide variants to generate antibodies (such as
anti-PSA antibodies or T cells) is typical in the art with a wide
variety of systems such as fusion proteins being used by
practitioners (see, e.g., Current Protocols In Molecular Biology,
Volume 2, Unit 16, Frederick M. Ausubel et al. eds., 1995). In this
context, each epitope(s) functions to provide the architecture with
which an antibody or T cell is reactive. Typically, skilled
artisans create a variety of different polypeptide fragments that
can be used in order to generate immune responses specific for
different portions of a polypeptide of interest (see, e.g., U.S.
Pat. No. 5,840,501 and U.S. Pat. No. 5,939,533). For example it may
be preferable to utilize a polypeptide comprising one of the
191P4D12(b) biological motifs discussed herein or a motif-bearing
subsequence which is readily identified by one of skill in the art
based on motifs available in the art. Polypeptide fragments,
variants or analogs are typically useful in this context as long as
they comprise an epitope capable of generating an antibody or T
cell specific for a target polypeptide sequence (e.g. a 191P4D12(b)
polypeptide shown in FIG. 3).
[0541] As shown herein, the 191P4D12(b) polynucleotides and
polypeptides (as well as the 191P4D12(b) polynucleotide probes and
anti-191P4D12(b) antibodies or T cells used to identify the
presence of these molecules) exhibit specific properties that make
them useful in diagnosing cancers such as those listed in Table I.
Diagnostic assays that measure the presence of 191P4D12(b) gene
products, in order to evaluate the presence or onset of a disease
condition described herein, such as prostate cancer, are used to
identify patients for preventive measures or further monitoring, as
has been done so successfully with PSA. Moreover, these materials
satisfy a need in the art for molecules having similar or
complementary characteristics to PSA in situations where, for
example, a definite diagnosis of metastasis of prostatic origin
cannot be made on the basis of a test for PSA alone (see, e.g.,
Alanen et al., Pathol. Res. Pract. 192(3): 233-237 (1996)), and
consequently, materials such as 191P4D12(b) polynucleotides and
polypeptides (as well as the 191P4D12(b) polynucleotide probes and
anti-191P4D12(b) antibodies used to identify the presence of these
molecules) need to be employed to confirm a metastases of prostatic
origin.
[0542] Finally, in addition to their use in diagnostic assays, the
191P4D12(b) polynucleotides disclosed herein have a number of other
utilities such as their use in the identification of oncogenetic
associated chromosomal abnormalities in the chromosomal region to
which the 191P4D12(b) gene maps (see the Example entitled
"Chromosomal Mapping of 191P4D12(b)" below). Moreover, in addition
to their use in diagnostic assays, the 191P4D12(b)-related proteins
and polynucleotides disclosed herein have other utilities such as
their use in the forensic analysis of tissues of unknown origin
(see, e.g., Takahama K Forensic Sci Int 1996 Jun 28;80(1-2):
63-9).
[0543] Additionally, 191P4D12(b)-related proteins or
polynucleotides of the invention can be used to treat a pathologic
condition characterized by the over-expression of 191P4D12(b). For
example, the amino acid or nucleic acid sequence of FIG. 2 or FIG.
3, or fragments of either, can be used to generate an immune
response to a 191P4D12(b) antigen. Antibodies or other molecules
that react with 191P4D12(b) can be used to modulate the function of
this molecule, and thereby provide a therapeutic benefit.
[0544] XII.) Inhibition of 191P4D12(b) Protein Function
[0545] The invention includes various methods and compositions for
inhibiting the binding of 191P4D12(b) to its binding partner or its
association with other protein(s) as well as methods for inhibiting
191P4D12(b) function.
[0546] XII.A.) Inhibition of 191P4D12(b) With Intracellular
Antibodies
[0547] In one approach, a recombinant vector that encodes single
chain antibodies that specifically bind to 191P4D12(b) are
introduced into 191P4D12(b) expressing cells via gene transfer
technologies. Accordingly, the encoded single chain
anti-191P4D12(b) antibody is expressed intracellularly, binds to
191P4D12(b) protein, and thereby inhibits its function. Methods for
engineering such intracellular single chain antibodies are well
known. Such intracellular antibodies, also known as "intrabodies",
are specifically targeted to a particular compartment within the
cell, providing control over where the inhibitory activity of the
treatment is focused. This technology has been successfully applied
in the art (for review, see Richardson and Marasco, 1995, TIBTECH
vol. 13). Intrabodies have been shown to virtually eliminate the
expression of otherwise abundant cell surface receptors (see, e.g.,
Richardson et al., 1995, Proc. Natl. Acad. Sci. USA 92: 3137-3141;
Beerli et al., 1994, J. Biol. Chem. 289: 23931-23936; Deshane et
al., 1994, Gene Ther. 1: 332-337).
[0548] Single chain antibodies comprise the variable domains of the
heavy and light chain joined by a flexible linker polypeptide, and
are expressed as a single polypeptide. Optionally, single chain
antibodies are expressed as a single chain variable region fragment
joined to the light chain constant region. Well-known intracellular
trafficking signals are engineered into recombinant polynucleotide
vectors encoding such single chain antibodies in order to target
precisely the intrabody to the desired intracellular compartment.
For example, intrabodies targeted to the endoplasmic reticulum (ER)
are engineered to incorporate a leader peptide and, optionally, a
C-terminal ER retention signal, such as the KDEL amino acid motif.
Intrabodies intended to exert activity in the nucleus are
engineered to include a nuclear localization signal. Lipid moieties
are joined to intrabodies in order to tether the intrabody to the
cytosolic side of the plasma membrane. Intrabodies can also be
targeted to exert function in the cytosol. For example, cytosolic
intrabodies are used to sequester factors within the cytosol,
thereby preventing them from being transported to their natural
cellular destination.
[0549] In one embodiment, intrabodies are used to capture
191P4D12(b) in the nucleus, thereby preventing its activity within
the nucleus. Nuclear targeting signals are engineered into such
191P4D12(b) intrabodies in order to achieve the desired targeting.
Such 191P4D12(b) intrabodies are designed to bind specifically to a
particular 191P4D12(b) domain. In another embodiment, cytosolic
intrabodies that specifically bind to a 191P4D12(b) protein are
used to prevent 191P4D12(b) from gaining access to the nucleus,
thereby preventing it from exerting any biological activity within
the nucleus (e.g., preventing 191P4D12(b) from forming
transcription complexes with other factors).
[0550] In order to specifically direct the expression of such
intrabodies to particular cells, the transcription of the intrabody
is placed under the regulatory control of an appropriate
tumor-specific promoter and/or enhancer. In order to target
intrabody expression specifically to prostate, for example, the PSA
promoter and/or promoter/enhancer can be utilized (See, for
example, U.S. Pat. No. 5,919,652 issued 6 Jul. 1999).
[0551] XII.B.) Inhibition of 191P4D12(b) with Recombinant
Proteins
[0552] In another approach, recombinant molecules bind to
191P4D12(b) and thereby inhibit 191P4D12(b) function. For example,
these recombinant molecules prevent or inhibit 191P4D12(b) from
accessing/binding to its binding partner(s) or associating with
other protein(s). Such recombinant molecules can, for example,
contain the reactive part(s) of a 191P4D12(b) specific antibody
molecule. In a particular embodiment, the 191P4D12(b) binding
domain of a 191P4D12(b) binding partner is engineered into a
dimeric fusion protein, whereby the fusion protein comprises two
191P4D12(b) ligand binding domains linked to the Fc portion of a
human IgG, such as human IgG1. Such IgG portion can contain, for
example, the C.sub.H2 and C.sub.H3 domains and the hinge region,
but not the C.sub.H1 domain. Such dimeric fusion proteins are
administered in soluble form to patients suffering from a cancer
associated with the expression of 191P4D12(b), whereby the dimeric
fusion protein specifically binds to 191P4D12(b) and blocks
191P4D12(b) interaction with a binding partner. Such dimeric fusion
proteins are further combined into multimeric proteins using known
antibody linking technologies.
[0553] XII.C.) Inhibition of 191P4D12(b) Transcription or
Translation
[0554] The present invention also comprises various methods and
compositions for inhibiting the transcription of the 191P4D12(b)
gene. Similarly, the invention also provides methods and
compositions for inhibiting the translation of 191P4D12(b) mRNA
into protein.
[0555] In one approach, a method of inhibiting the transcription of
the 191P4D12(b) gene comprises contacting the 191P4D12(b) gene with
a 191P4D12(b) antisense polynucleotide. In another approach, a
method of inhibiting 191P4D12(b) mRNA translation comprises
contacting a 191P4D12(b) mRNA with an antisense polynucleotide. In
another approach, a 191P4D12(b) specific ribozyme is used to cleave
a 191P4D12(b) message, thereby inhibiting translation. Such
antisense and ribozyme based methods can also be directed to the
regulatory regions of the 191P4D12(b) gene, such as 191P4D12(b)
promoter and/or enhancer elements. Similarly, proteins capable of
inhibiting a 191P4D12(b) gene transcription factor are used to
inhibit 191P4D12(b) mRNA transcription. The various polynucleotides
and compositions useful in the aforementioned methods have been
described above. The use of antisense and ribozyme molecules to
inhibit transcription and translation is well known in the art.
[0556] Other factors that inhibit the transcription of 191P4D12(b)
by interfering with 191P4D12(b) transcriptional activation are also
useful to treat cancers expressing 191P4D12(b). Similarly, factors
that interfere with 191P4D12(b) processing are useful to treat
cancers that express 191P4D12(b). Cancer treatment methods
utilizing such factors are also within the scope of the
invention.
[0557] XII.D.) General Considerations for Therapeutic
Strategies
[0558] Gene transfer and gene therapy technologies can be used to
deliver therapeutic polynucleotide molecules to tumor cells
synthesizing 191P4D12(b) (i.e., antisense, ribozyme,
polynucleotides encoding intrabodies and other 191P4D12(b)
inhibitory molecules). A number of gene therapy approaches are
known in the art. Recombinant vectors encoding 191P4D12(b)
antisense polynucleotides, ribozymes, factors capable of
interfering with 191P4D12(b) transcription, and so forth, can be
delivered to target tumor cells using such gene therapy
approaches.
[0559] The above therapeutic approaches can be combined with any
one of a wide variety of surgical, chemotherapy or radiation
therapy regimens. The therapeutic approaches of the invention can
enable the use of reduced dosages of chemotherapy (or other
therapies) and/or less frequent administration, an advantage for
all patients and particularly for those that do not tolerate the
toxicity of the chemotherapeutic agent well.
[0560] The anti-tumor activity of a particular compositon (e.g.,
antisense, ribozyme, intrabody), or a combination of such
compositions, can be evaluated using various in vitro and in vivo
assay systems. In vitro assays that evaluate therapeutic activity
include cell growth assays, soft agar assays and other assays
indicative of tumor promoting activity, binding assays capable of
determining the extent to which a therapeutic composition will
inhibit the binding of 191P4D12(b) to a binding partner, etc.
[0561] In vivo, the effect of a 191P4D12(b) therapeutic composition
can be evaluated in a suitable animal model. For example, xenogenic
prostate cancer models can be used, wherein human prostate cancer
explants or passaged xenograft issues are introduced into immune
compromised animals, such as nude or SCID mice (Klein et al., 1997,
Nature Medicine 3: 402-408). For example, PCT Patent Application
WO98/16628 and U.S. Pat. No. 6,107,540 describe various xenograft
models of human prostate cancer capable of recapitulating the
development of primary tumors, micrometastasis, and the formation
of osteoblastic metastases characteristic of late stage disease.
Efficacy can be predicted using assays that measure inhibition of
tumor formation, tumor regression or metastasis, and the like.
[0562] In vivo assays that evaluate the promotion of apoptosis are
useful in evaluating therapeutic compositions. In one embodiment,
xenografts from tumor bearing mice treated with the therapeutic
composition can be examined for the presence of apoptotic foci and
compared to untreated control xenograft-bearing mice. The extent to
which apoptotic foci are found in the tumors of the treated mice
provides an indication of the therapeutic efficacy of the
composition.
[0563] The therapeutic compositions used in the practice of the
foregoing methods can be formulated into pharmaceutical
compositions comprising a carrier suitable for the desired delivery
method. Suitable carriers include any material that when combined
with the therapeutic composition retains the anti-tumor function of
the therapeutic composition and is generally non-reactive with the
patient's immune system. Examples include, but are not limited to,
any of a number of standard pharmaceutical carriers such as sterile
phosphate buffered saline solutions, bacteriostatic water, and the
like (see, generally, Remington's Pharmaceutical Sciences 16.sup.th
Edition, A. Osal., Ed., 1980).
[0564] Therapeutic formulations can be solubilized and administered
via any route capable of delivering the therapeutic composition to
the tumor site. Potentially effective routes of administration
include, but are not limited to, intravenous, parenteral,
intraperitoneal, intramuscular, intratumor, intradermal,
intraorgan, orthotopic, and the like. A preferred formulation for
intravenous injection comprises the therapeutic composition in a
solution of preserved bacteriostatic water, sterile unpreserved
water, and/or diluted in polyvinylchloride or polyethylene bags
containing 0.9% sterile Sodium Chloride for Injection, USP.
Therapeutic protein preparations can be lyophilized and stored as
sterile powders, preferably under vacuum, and then reconstituted in
bacteriostatic water (containing for example, benzyl alcohol
preservative) or in sterile water prior to injection.
[0565] Dosages and administration protocols for the treatment of
cancers using the foregoing methods will vary with the method and
the target cancer, and will generally depend on a number of other
factors appreciated in the art.
[0566] XIII.) Identification, Characterization and Use of
Modulators of 191P4D12(b)
[0567] Methods to Identify and Use Modulators
[0568] In one embodiment, screening is performed to identify
modulators that induce or suppress a particular expression profile,
suppress or induce specific pathways, preferably generating the
associated phenotype thereby. In another embodiment, having
identified differentially expressed genes important in a particular
state; screens are performed to identify modulators that alter
expression of individual genes, either increase or decrease. In
another embodiment, screening is performed to identify modulators
that alter a biological function of the expression product of a
differentially expressed gene. Again, having identified the
importance of a gene in a particular state, screens are performed
to identify agents that bind and/or modulate the biological
activity of the gene product.
[0569] In addition, screens are done for genes that are induced in
response to a candidate agent. After identifying a modulator (one
that suppresses a cancer expression pattern leading to a normal
expression pattern, or a modulator of a cancer gene that leads to
expression of the gene as in normal tissue) a screen is performed
to identify genes that are specifically modulated in response to
the agent. Comparing expression profiles between normal tissue and
agent-treated cancer tissue reveals genes that are not expressed in
normal tissue or cancer tissue, but are expressed in agent treated
tissue, and vice versa. These agent-specific sequences are
identified and used by methods described herein for cancer genes or
proteins. In particular these sequences and the proteins they
encode are used in marking or identifying agent-treated cells. In
addition, antibodies are raised against the agent-induced proteins
and used to target novel therapeutics to the treated cancer tissue
sample.
[0570] Modulator-related Identification and Screening Assays:
[0571] Gene Expression-related Assays
[0572] Proteins, nucleic acids, and antibodies of the invention are
used in screening assays. The cancer-associated proteins,
antibodies, nucleic acids, modified proteins and cells containing
these sequences are used in screening assays, such as evaluating
the effect of drug candidates on a "gene expression profile,"
expression profile of polypeptides or alteration of biological
function. In one embodiment, the expression profiles are used,
preferably in conjunction with high throughput screening techniques
to allow monitoring for expression profile genes after treatment
with a candidate agent (e.g., Davis, G F, et al, J Biol Screen 7:69
(2002); Zlokarnik, et al., Science 279:84-8 (1998); Heid, Genome
Res 6:986-94,1996).
[0573] The cancer proteins, antibodies, nucleic acids, modified
proteins and cells containing the native or modified cancer
proteins or genes are used in screening assays. That is, the
present invention comprises methods for screening for compositions
which modulate the cancer phenotype or a physiological function of
a cancer protein of the invention. This is done on a gene itself or
by evaluating the effect of drug candidates on a "gene expression
profile" or biological function. In one embodiment, expression
profiles are used, preferably in conjunction with high throughput
screening techniques to allow monitoring after treatment with a
candidate agent, see Zlokamik, supra.
[0574] A variety of assays are executed directed to the genes and
proteins of the invention. Assays are run on an individual nucleic
acid or protein level. That is, having identified a particular gene
as up regulated in cancer, test compounds are screened for the
ability to modulate gene expression or for binding to the cancer
protein of the invention. "Modulation" in this context includes an
increase or a decrease in gene expression. The preferred amount of
modulation will depend on the original change of the gene
expression in normal versus tissue undergoing cancer, with changes
of at least 10%, preferably 50%, more preferably 100-300%, and in
some embodiments 300-1000% or greater. Thus, if a gene exhibits a
4-fold increase in cancer tissue compared to normal tissue, a
decrease of about four-fold is often desired; similarly, a 10-fold
decrease in cancer tissue compared to normal tissue a target value
of a 10-fold increase in expression by the test compound is often
desired. Modulators that exacerbate the type of gene expression
seen in cancer are also useful, e.g., as an upregulated target in
further analyses.
[0575] The amount of gene expression is monitored using nucleic
acid probes and the quantification of gene expression levels, or,
alternatively, a gene product itself is monitored, e.g., through
the use of antibodies to the cancer protein and standard
immunoassays. Proteomics and separation techniques also allow for
quantification of expression.
[0576] Expression Monitoring to Identify Compounds that Modify Gene
Expression
[0577] In one embodiment, gene expression monitoring, i.e., an
expression profile, is monitored simultaneously for a number of
entities. Such profiles will typically involve one or more of the
genes of FIG. 2. In this embodiment, e.g., cancer nucleic acid
probes are attached to biochips to detect and quantify cancer
sequences in a particular cell. Alternatively, PCR can be used.
Thus, a series, e.g., wells of a microtiter plate, can be used with
dispensed primers in desired wells. A PCR reaction can then be
performed and analyzed for each well.
[0578] Expression monitoring is performed to identify compounds
that modify the expression of one or more cancer-associated
sequences, e.g., a polynucleotide sequence set out in FIG. 2.
Generally, a test modulator is added to the cells prior to
analysis. Moreover, screens are also provided to identify agents
that modulate cancer, modulate cancer proteins of the invention,
bind to a cancer protein of the invention, or interfere with the
binding of a cancer protein of the invention and an antibody or
other binding partner.
[0579] In one embodiment, high throughput screening methods involve
providing a library containing a large number of potential
therapeutic compounds (candidate compounds). Such "combinatorial
chemical libraries" are then screened in one or more assays to
identify those library members (particular chemical species or
subclasses) that display a desired characteristic activity. The
compounds thus identified can serve as conventional "lead
compounds," as compounds for screening, or as therapeutics.
[0580] In certain embodiments, combinatorial libraries of potential
modulators are screened for an ability to bind to a cancer
polypeptide or to modulate activity. Conventionally, new chemical
entities with useful properties are generated by identifying a
chemical compound (called a "lead compound") with some desirable
property or activity, e.g., inhibiting activity, creating variants
of the lead compound, and evaluating the property and activity of
those variant compounds. Often, high throughput screening (HTS)
methods are employed for such an analysis.
[0581] As noted above, gene expression monitoring is conveniently
used to test candidate modulators (e.g., protein, nucleic acid or
small molecule). After the candidate agent has been added and the
cells allowed to incubate for a period, the sample containing a
target sequence to be analyzed is, e.g., added to a biochip.
[0582] If required, the target sequence is prepared using known
techniques. For example, a sample is treated to lyse the cells,
using known lysis buffers, electroporation, etc., with purification
and/or amplification such as PCR performed as appropriate. For
example, an in vitro transcription with labels covalently attached
to the nucleotides is performed. Generally, the nucleic acids are
labeled with biotin-FITC or PE, or with cy3 or cy5.
[0583] The target sequence can be labeled with, e.g., a
fluorescent, a chemiluminescent, a chemical, or a radioactive
signal, to provide a means of detecting the target sequence's
specific binding to a probe. The label also can be an enzyme, such
as alkaline phosphatase or horseradish peroxidase, which when
provided with an appropriate substrate produces a product that is
detected. Alternatively, the label is a labeled compound or small
molecule, such as an enzyme inhibitor, that binds but is not
catalyzed or altered by the enzyme. The label also can be a moiety
or compound, such as, an epitope tag or biotin which specifically
binds to streptavidin. For the example of biotin, the streptavidin
is labeled as described above, thereby, providing a detectable
signal for the bound target sequence. Unbound labeled streptavidin
is typically removed prior to analysis.
[0584] As will be appreciated by those in the art, these assays can
be direct hybridization assays or can comprise "sandwich assays",
which include the use of multiple probes, as is generally outlined
in U.S. Pat. Nos. 5,681,702; 5,597,909; 5,545,730; 5,594,117;
5,591,584; 5,571,670; 5,580,731; 5,571,670; 5,591,584; 5,624,802;
5,635,352; 5,594,118; 5,359,100; 5,124,246; and 5,681,697. In this
embodiment, in general, the target nucleic acid is prepared as
outlined above, and then added to the biochip comprising a
plurality of nucleic acid probes, under conditions that allow the
formation of a hybridization complex.
[0585] A variety of hybridization conditions are used in the
present invention, including high, moderate and low stringency
conditions as outlined above. The assays are generally run under
stringency conditions which allow formation of the label probe
hybridization complex only in the presence of target. Stringency
can be controlled by altering a step parameter that is a
thermodynamic variable, including, but not limited to, temperature,
formamide concentration, salt concentration, chaotropic salt
concentration pH, organic solvent concentration, etc. These
parameters may also be used to control non-specific binding, as is
generally outlined in U.S. Pat. No. 5,681,697. Thus, it can be
desirable to perform certain steps at higher stringency conditions
to reduce non-specific binding.
[0586] The reactions outlined herein can be accomplished in a
variety of ways. Components of the reaction can be added
simultaneously, or sequentially, in different orders, with
preferred embodiments outlined below. In addition, the reaction may
include a variety of other reagents. These include salts, buffers,
neutral proteins, e.g. albumin, detergents, etc. which can be used
to facilitate optimal hybridization and detection, and/or reduce
nonspecific or background interactions. Reagents that otherwise
improve the efficiency of the assay, such as protease inhibitors,
nuclease inhibitors, anti-microbial agents, etc., may also be used
as appropriate, depending on the sample preparation methods and
purity of the target. The assay data are analyzed to determine the
expression levels of individual genes, and changes in expression
levels as between states, forming a gene expression profile.
[0587] Biological Activity-related Assays
[0588] The invention provides methods identify or screen for a
compound that modulates the activity of a cancer-related gene or
protein of the invention. The methods comprise adding a test
compound, as defined above, to a cell comprising a cancer protein
of the invention. The cells contain a recombinant nucleic acid that
encodes a cancer protein of the invention. In another embodiment, a
library of candidate agents is tested on a plurality of cells.
[0589] In one aspect, the assays are evaluated in the presence or
absence or previous or subsequent exposure of physiological
signals, e.g. hormones, antibodies, peptides, antigens, cytokines,
growth factors, action potentials, pharmacological agents including
chemotherapeutics, radiation, carcinogenics, or other cells (i.e.,
cell-cell contacts). In another example, the determinations are
made at different stages of the cell cycle process. In this way,
compounds that modulate genes or proteins of the invention are
identified. Compounds with pharmacological activity are able to
enhance or interfere with the activity of the cancer protein of the
invention. Once identified, similar structures are evaluated to
identify critical structural features of the compound.
[0590] In one embodiment, a method of modulating (e.g., inhibiting)
cancer cell division is provided; the method comprises
administration of a cancer modulator. In another embodiment, a
method of modulating (e.g., inhibiting) cancer is provided; the
method comprises administration of a cancer modulator. In a further
embodiment, methods of treating cells or individuals with cancer
are provided; the method comprises administration of a cancer
modulator.
[0591] In one embodiment, a method for modulating the status of a
cell that expresses a gene of the invention is provided. As used
herein status comprises such art-accepted parameters such as
growth, proliferation, survival, function, apoptosis, senescence,
location, enzymatic activity, signal transduction, etc. of a cell.
In one embodiment, a cancer inhibitor is an antibody as discussed
above. In another embodiment, the cancer inhibitor is an antisense
molecule. A variety of cell growth, proliferation, and metastasis
assays are known to those of skill in the art, as described
herein.
[0592] High Throughput Screening to Identify Modulators
[0593] The assays to identify suitable modulators are amenable to
high throughput screening. Preferred assays thus detect enhancement
or inhibition of cancer gene transcription, inhibition or
enhancement of polypeptide expression, and inhibition or
enhancement of polypeptide activity.
[0594] In one embodiment, modulators evaluated in high throughput
screening methods are proteins, often naturally occurring proteins
or fragments of naturally occurring proteins. Thus, e.g., cellular
extracts containing proteins, or random or directed digests of
proteinaceous cellular extracts, are used. In this way, libraries
of proteins are made for screening in the methods of the invention.
Particularly preferred in this embodiment are libraries of
bacterial, fungal, viral, and mammalian proteins, with the latter
being preferred, and human proteins being especially preferred.
Particularly useful test compound will be directed to the class of
proteins to which the target belongs, e.g., substrates for enzymes,
or ligands and receptors.
[0595] Use of Soft Agar Growth and Colony Formation to Identify and
Characterize Modulators
[0596] Normal cells require a solid substrate to attach and grow.
When cells are transformed, they lose this phenotype and grow
detached from the substrate. For example, transformed cells can
grow in stirred suspension culture or suspended in semi-solid
media, such as semi-solid or soft agar. The transformed cells, when
transfected with tumor suppressor genes, can regenerate normal
phenotype and once again require a solid substrate to attach to and
grow. Soft agar growth or colony formation in assays are used to
identify modulators of cancer sequences, which when expressed in
host cells, inhibit abnormal cellular proliferation and
transformation. A modulator reduces or eliminates the host cells'
ability to grow suspended in solid or semisolid media, such as
agar.
[0597] Techniques for soft agar growth or colony formation in
suspension assays are described in Freshney, Culture of Animal
Cells a Manual of Basic Technique (3rd ed., 1994). See also, the
methods section of Garkavtsev et al. (1996), supra.
[0598] Evaluation of Contact Inhibition and Growth Density
Limitation to Identify and Characterize Modulators
[0599] Normal cells typically grow in a flat and organized pattern
in cell culture until they touch other cells. When the cells touch
one another, they are contact inhibited and stop growing.
Transformed cells, however, are not contact inhibited and continue
to grow to high densities in disorganized foci. Thus, transformed
cells grow to a higher saturation density than corresponding normal
cells. This is detected morphologically by the formation of a
disoriented monolayer of cells or cells in foci. Alternatively,
labeling index with (.sup.3H)-thymidine at saturation density is
used to measure density limitation of growth, similarly an MTT or
Alamar blue assay will reveal proliferation capacity of cells and
the the ability of modulators to affect same. See Freshney (1994),
supra. Transformed cells, when transfected with tumor suppressor
genes, can regenerate a normal phenotype and become contact
inhibited and would grow to a lower density.
[0600] In this assay, labeling index with .sup.3H)-thymidine at
saturation density is a preferred method of measuring density
limitation of growth. Transformed host cells are transfected with a
cancer-associated sequence and are grown for 24 hours at saturation
density in non-limiting medium conditions. The percentage of cells
labeling with (.sup.3H)-thymidine is determined by incorporated
cpm.
[0601] Contact independent growth is used to identify modulators of
cancer sequences, which had led to abnormal cellular proliferation
and transformation. A modulator reduces or eliminates contact
independent growth, and returns the cells to a normal
phenotype.
[0602] Evaluation of Growth Factor or Serum Dependence to Identify
and Characterize Modulators
[0603] Transformed cells have lower serum dependence than their
normal counterparts (see, e.g., Temin, J. Natl. Cancer Inst.
37:167-175 (1966); Eagle et al., J. Exp. Med 131:836-879 (1970));
Freshney, supra. This is in part due to release of various growth
factors by the transformed cells. The degree of growth factor or
serum dependence of transformed host cells can be compared with
that of control. For example, growth factor or serum dependence of
a cell is monitored in methods to identify and characterize
compounds that modulate cancer-associated sequences of the
invention.
[0604] Use of Tumor-specific Marker Levels to Identify and
Characterize Modulators
[0605] Tumor cells release an increased amount of certain factors
(hereinafter "tumor specific markers") than their normal
counterparts. For example, plasminogen activator (PA) is released
from human glioma at a higher level than from normal brain cells
(see, e.g., Gullino, Angiogenesis, Tumor Vascularization, and
Potential Interference with Tumor Growth, in Biological Responses
in Cancer, pp. 178-184 (Mihich (ed.) 1985)). Similarly, Tumor
Angiogenesis Factor (TAF) is released at a higher level in tumor
cells than their normal counterparts. See, e.g., Folkman,
Angiogenesis and Cancer, Sem Cancer Biol. (1992)), while bFGF is
released from endothelial tumors (Ensoli, B et al).
[0606] Various techniques which measure the release of these
factors are described in Freshney (1994), supra. Also, see, Unkless
et al., J. Biol. Chem. 249:4295-4305 (1974); Strickland &
Beers, J. Biol. Chem. 251:5694-5702 (1976); Whur et al., Br. J.
Cancer 42:305 312 (1980);, Gullino, Angiogenesis, Tumor
Vascularization; and Potential Interference with Tumor Growth, in
Biological Responses in Cancer, pp. 178-184 (Mihich (ed.) 1985);
Freshney, Anticancer Res. 5:111-130 (1985). For example, tumor
specific marker levels are monitored in methods to identify and
characterize compounds that modulate cancer-associated sequences of
the invention.
[0607] Invasiveness into Matrigel to Identify and Characterize
Modulators
[0608] The degree of invasiveness into Matrigel or an extracellular
matrix constituent can be used as an assay to identify and
characterize compounds that modulate cancer associated sequences.
Tumor cells exhibit a positive correlation between malignancy and
invasiveness of cells into Matrigel or some other extracellular
matrix constituent. In this assay, tumorigenic cells are typically
used as host cells. Expression of a tumor suppressor gene in these
host cells would decrease invasiveness of the host cells.
Techniques described in Cancer Res. 1999; 59:6010; Freshney (1994),
supra, can be used. Briefly, the level of invasion of host cells is
measured by using filters coated with Matrigel or some other
extracellular matrix constituent. Penetration into the gel, or
through to the distal side of the filter, is rated as invasiveness,
and rated histologically by number of cells and distance moved, or
by prelabeling the cells with .sup.1251 and counting the
radioactivity on the distal side of the filter or bottom of the
dish. See, e.g., Freshney (1984), supra.
[0609] Evaluation of Tumor Growth In Vivo to Identify and
Characterize Modulators
[0610] Effects of cancer-associated sequences on cell growth are
tested in transgenic or immune-suppressed organisms. Transgenic
organisms are prepared in a variety of art-accepted ways. For
example, knock-out transgenic organisms, e.g., mammals such as
mice, are made, in which a cancer gene is disrupted or in which a
cancer gene is inserted. Knock-out transgenic mice are made by
insertion of a marker gene or other heterologous gene into the
endogenous cancer gene site in the mouse genome via homologous
recombination. Such mice can also be made by substituting the
endogenous cancer gene with a mutated version of the cancer gene,
or by mutating the endogenous cancer gene, e.g., by exposure to
carcinogens.
[0611] To prepare transgenic chimeric animals, e.g., mice, a DNA
construct is introduced into the nuclei of embryonic stem cells.
Cells containing the newly engineered genetic lesion are injected
into a host mouse embryo, which is re-implanted into a recipient
female. Some of these embryos develop into chimeric mice that
possess germ cells some of which are derived from the mutant cell
line. Therefore, by breeding the chimeric mice it is possible to
obtain a new line of mice containing the introduced genetic lesion
(see, e.g., Capecchi et al., Science 244:1288 (1989)). Chimeric
mice can be derived according to U.S. Pat. No. 6,365,797, issued 2
Apr. 2002; U.S. Pat. No. 6,107,540 issued 22 Aug. 2000; Hogan et
al., Manipulating the Mouse Embryo: A laboratory Manual, Cold
Spring Harbor Laboratory (1988) and Teratocarcinomas and Embryonic
Stem Cells: A Practical Approach, Robertson, ed., IRL Press,
Washington, D.C., (1987).
[0612] Alternatively, various immune-suppressed or immune-deficient
host animals can be used. For example, a genetically athymic "nude"
mouse (see, e.g., Giovanella et al., J. Natl. Cancer Inst. 52:921
(1974)), a SCID mouse, a thymectornized mouse, or an irradiated
mouse (see, e.g., Bradley et al., Br. J. Cancer 38:263 (1978);
Selby et al., Br. J. Cancer 41:52 (1980)) can be used as a host.
Transplantable tumor cells (typically about 10.sup.6 cells)
injected into isogenic hosts produce invasive tumors in a high
proportion of cases, while normal cells of similar origin will not.
In hosts which developed invasive tumors, cells expressing
cancer-associated sequences are injected subcutaneously or
orthotopically. Mice are then separated into groups, including
control groups and treated experimental groups) e.g. treated with a
modulator). After a suitable length of time, preferably 4-8 weeks,
tumor growth is measured (e.g., by volume or by its two largest
dimensions, or weight) and compared to the control. Tumors that
have statistically significant reduction (using, e.g., Student's T
test) are said to have inhibited growth,
[0613] In Vitro Assays to Identify and Characterize Modulators
[0614] Assays to identify compounds with modulating activity can be
performed in vitro. For example, a cancer polypeptide is first
contacted with a potential modulator and incubated for a suitable
amount of time, e.g., from 0.5 to 48 hours. In one embodiment, the
cancer polypeptide levels are determined in vitro by measuring the
level of protein or mRNA. The level of protein is measured using
immunoassays such as Western blotting, ELISA and the like with an
antibody that selectively binds to the cancer polypeptide or a
fragment thereof. For measurement of mRNA, amplification, e.g.,
using PCR, LCR, or hybridization assays, e. g., Northern
hybridization, RNAse protection, dot blotting, are preferred. The
level of protein or mRNA is detected using directly or indirectly
labeled detection agents, e.g., fluorescently or radioactively
labeled nucleic acids, radioactively or enzymatically labeled
antibodies, and the like, as described herein.
[0615] Alternatively, a reporter gene system can be devised using a
cancer protein promoter operably linked to a reporter gene such as
luciferase, green fluorescent protein, CAT, or P-gal. The reporter
construct is typically transfected into a cell. After treatment
with a potential modulator, the amount of reporter gene
transcription, translation, or activity is measured according to
standard techniques known to those of skill in the art (Davis G F,
supra; Gonzalez, J. & Negulescu, P. Curr. Opin. Biotechnol.
1998: 9:624).
[0616] As outlined above, in vitro screens are done on individual
genes and gene products. That is, having identified a particular
differentially expressed gene as important in a particular state,
screening of modulators of the expression of the gene or the gene
product itself is performed.
[0617] In one embodiment, screening for modulators of expression of
specific gene(s) is performed. Typically, the expression of only
one or a few genes is evaluated. In another embodiment, screens are
designed to first find compounds that bind to differentially
expressed proteins. These compounds are then evaluated for the
ability to modulate differentially expressed activity. Moreover,
once initial candidate compounds are identified, variants can be
further screened to better evaluate structure activity
relationships.
[0618] Binding Assays to Identify and Characterize Modulators
[0619] In binding assays in accordance with the invention, a
purified or isolated gene product of the invention is generally
used. For example, antibodies are generated to a protein of the
invention, and immunoassays are run to determine the amount and/or
location of protein. Alternatively, cells comprising the cancer
proteins are used in the assays.
[0620] Thus, the methods comprise combining a cancer protein of the
invention and a candidate compound such as a ligand, and
determining the binding of the compound to the cancer protein of
the invention. Preferred embodiments utilize the human cancer
protein; animal models of human disease of can also be developed
and used. Also, other analogous mammalian proteins also can be used
as appreciated by those of skill in the art. Moreover, in some
embodiments variant or derivative cancer proteins are used.
[0621] Generally, the cancer protein of the invention, or the
ligand, is non-diffusibly bound to an insoluble support. The
support can, e.g., be one having isolated sample receiving areas (a
microtiter plate, an array, etc.). The insoluble supports can be
made of any composition to which the compositions can be bound, is
readily separated from soluble material, and is otherwise
compatible with the overall method of screening. The surface of
such supports can be solid or porous and of any convenient
shape.
[0622] Examples of suitable insoluble supports include microtiter
plates, arrays, membranes and beads. These are typically made of
glass, plastic (e.g., polystyrene), polysaccharide, nylon,
nitrocellulose, or Teflon.TM., etc. Microtiter plates and arrays
are especially convenient because a large number of assays can be
carried out simultaneously, using small amounts of reagents and
samples. The particular manner of binding of the composition to the
support is not crucial so long as it is compatible with the
reagents and overall methods of the invention, maintains the
activity of the composition and is nondiffusable. Preferred methods
of binding include the use of antibodies which do not sterically
block either the ligand binding site or activation sequence when
attaching the protein to the support, direct binding to "sticky" or
ionic supports, chemical crosslinking, the synthesis of the protein
or agent on the surface, etc. Following binding of the protein or
ligand/binding agent to the support, excess unbound material is
removed by washing. The sample receiving areas may then be blocked
through incubation with bovine serum albumin (BSA), casein or other
innocuous protein or other moiety.
[0623] Once a cancer protein of the invention is bound to the
support, and a test compound is added to the assay. Alternatively,
the candidate binding agent is bound to the support and the cancer
protein of the invention is then added. Binding agents include
specific antibodies, non-natural binding agents identified in
screens of chemical libraries, peptide analogs, etc.
[0624] Of particular interest are assays to identify agents that
have a low toxicity for human cells. A wide variety of assays can
be used for this purpose, including proliferation assays, cAMP
assays, labeled in vitro protein-protein binding assays,
electrophoretic mobility shift assays, immunoassays for protein
binding, functional assays (phosphorylation assays, etc.) and the
like.
[0625] A determination of binding of the test compound (ligand,
binding agent, modulator, etc.) to a cancer protein of the
invention can be done in a number of ways. The test compound can be
labeled, and binding determined directly, e.g., by attaching all or
a portion of the cancer protein of the invention to a solid
support, adding a labeled candidate compound (e.g., a fluorescent
label), washing off excess reagent, and determining whether the
label is present on the solid support. Various blocking and washing
steps can be utilized as appropriate.
[0626] In certain embodiments, only one of the components is
labeled, e.g., a protein of the invention or ligands labeled.
Alternatively, more than one component is labeled with different
labels, e.g., I.sup.125, for the proteins and a fluorophor for the
compound. Proximity reagents, e.g., quenching or energy transfer
reagents are also useful.
[0627] Competitive Binding to Identify and Characterize
Modulators
[0628] In one embodiment, the binding of the "test compound" is
determined by competitive binding assay with a "competitor." The
competitor is a binding moiety that binds to the target molecule
(e.g., a cancer protein of the invention). Competitors include
compounds such as antibodies, peptides, binding partners, ligands,
etc. Under certain circumstances, the competitive binding between
the test compound and the competitor displaces the test compound.
In one embodiment, the test compound is labeled. Either the test
compound, the competitor, or both, is added to the protein for a
time sufficient to allow binding. Incubations are performed at a
temperature that facilitates optimal activity, typically between
four and 40.degree. C. Incubation periods are typically optimized,
e.g., to facilitate rapid high throughput screening; typically
between zero and one hour will be sufficient. Excess reagent is
generally removed or washed away. The second component is then
added, and the presence or absence of the labeled component is
followed, to indicate binding.
[0629] In one embodiment, the competitor is added first, followed
by the test compound. Displacement of the competitor is an
indication that the test compound is binding to the cancer protein
and thus is capable of binding to, and potentially modulating, the
activity of the cancer protein. In this embodiment, either
component can be labeled. Thus, e.g., if the competitor is labeled,
the presence of label in the post-test compound wash solution
indicates displacement by the test compound. Alternatively, if the
test compound is labeled, the presence of the label on the support
indicates displacement.
[0630] In an alternative embodiment, the test compound is added
first, with incubation and washing, followed by the competitor. The
absence of binding by the competitor indicates that the test
compound binds to the cancer protein with higher affinity than the
competitor. Thus, if the test compound is labeled, the presence of
the label on the support, coupled with a lack of competitor
binding, indicates that the test compound binds to and thus
potentially modulates the cancer protein of the invention.
[0631] Accordingly, the competitive binding methods comprise
differential screening to identity agents that are capable of
modulating the activity of the cancer proteins of the invention. In
this embodiment, the methods comprise combining a cancer protein
and a competitor in a first sample. A second sample comprises a
test compound, the cancer protein, and a competitor. The binding of
the competitor is determined for both samples, and a change, or
difference in binding between the two samples indicates the
presence of an agent capable of binding to the cancer protein and
potentially modulating its activity. That is, if the binding of the
competitor is different in the second sample relative to the first
sample, the agent is capable of binding to the cancer protein.
[0632] Alternatively, differential screening is used to identify
drug candidates that bind to the native cancer protein, but cannot
bind to modified cancer proteins. For example the structure of the
cancer protein is modeled and used in rational drug design to
synthesize agents that interact with that site, agents which
generally do not bind to site-modified proteins. Moreover, such
drug candidates that affect the activity of a native cancer protein
are also identified by screening drugs for the ability to either
enhance or reduce the activity of such proteins.
[0633] Positive controls and negative controls can be used in the
assays. Preferably control and test samples are performed in at
least triplicate to obtain statistically significant results.
Incubation of all samples occurs for a time sufficient to allow for
the binding of the agent to the protein. Following incubation,
samples are washed free of non-specifically bound material and the
amount of bound, generally labeled agent determined. For example,
where a radiolabel is employed, the samples can be counted in a
scintillation counter to determine the amount of bound
compound.
[0634] A variety of other reagents can be included in the screening
assays. These include reagents like salts, neutral proteins, e.g.
albumin, detergents, etc. which are used to facilitate optimal
protein-protein binding and/or reduce non-specific or background
interactions. Also reagents that otherwise improve the efficiency
of the assay, such as protease inhibitors, nuclease inhibitors,
anti-microbial agents, etc., can be used. The mixture of components
is added in an order that provides for the requisite binding.
[0635] Use of Polynucleotides to Down-regulate or Inhibit a Protein
of the Invention.
[0636] Polynucleotide modulators of cancer can be introduced into a
cell containing the target nucleotide sequence by formation of a
conjugate with a ligand-binding molecule, as described in WO
91/04753. Suitable ligand-binding molecules include, but are not
limited to, cell surface receptors, growth factors, other
cytokines, or other ligands that bind to cell surface receptors.
Preferably, conjugation of the ligand binding molecule does not
substantially interfere with the ability of the ligand binding
molecule to bind to its corresponding molecule or receptor, or
block entry of the sense or anti sense oligonucleotide or its
conjugated version into the cell. Alternatively, a polynucleotide
modulator of cancer can be introduced into a cell containing the
target nucleic acid sequence, e.g., by formation of a
polynucleotide-lipid complex, as described in WO 90/10448. It is
understood that the use of antisense molecules or knock out and
knock in models may also be used in screening assays as discussed
above, in addition to methods of treatment.
[0637] Inhibitory and Antisense Nucleotides
[0638] In certain embodiments, the activity of a cancer-associated
protein is down-regulated, or entirely inhibited, by the use of
antisense polynucleotide or inhibitory small nuclear RNA (snRNA),
i.e., a nucleic acid complementary to, and which can preferably
hybridize specifically to, a coding mRNA nucleic acid sequence,
e.g., a cancer protein of the invention, mRNA, or a subsequence
thereof. Binding of the antisense polynucleotide to the mRNA
reduces the translation and/or stability of the mRNA.
[0639] In the context of this invention, antisense polynucleotides
can comprise naturally occurring nucleotides, or synthetic species
formed from naturally occurring subunits or their close homologs.
Antisense polynucleotides may also have altered sugar moieties or
inter-sugar linkages. Exemplary among these are the
phosphorothioate and other sulfur containing species which are
known for use in the art. Analogs are comprised by this invention
so long as they function effectively to hybridize with nucleotides
of the invention. See, e.g., Isis Pharmaceuticals, Carlsbad,
Calif.; Sequitor, Inc., Natick, Mass.
[0640] Such antisense polynucleotides can readily be synthesized
using recombinant means, or can be synthesized in vitro. Equipment
for such synthesis is sold by several vendors, including Applied
Biosystems. The preparation of other oligonucleotides such as
phosphorothioates and alkylated derivatives is also well known to
those of skill in the art.
[0641] Antisense molecules as used herein include antisense or
sense oligonucleotides. Sense oligonucleotides can, e.g., be
employed to block transcription by binding to the anti-sense
strand. The antisense and sense oligonucleotide comprise a single
stranded nucleic acid sequence (either RNA or DNA) capable of
binding to target mRNA (sense) or DNA (antisense) sequences for
cancer molecules. Antisense or sense oligonucleotides, according to
the present invention, comprise a fragment generally at least about
12 nucleotides, preferably from about 12 to 30 nucleotides. The
ability to derive an antisense or a sense oligonucleotide, based
upon a cDNA sequence encoding a given protein is described in,
e.g., Stein & Cohen (Cancer Res. 48:2659 (1988 and van der Krol
et al. (BioTechniques 6:958 (1988)).
[0642] Ribozymes
[0643] In addition to antisense polynucleotides, ribozymes can be
used to target and inhibit transcription of cancer-associated
nucleotide sequences. A ribozyme is an RNA molecule that
catalytically cleaves other RNA molecules. Different kinds of
ribozymes have been described, including group I ribozymes,
hammerhead ribozymes, hairpin ribozymes, RNase P, and axhead
ribozymes (see, e.g., Castanotto et al., Adv. in Pharmacology 25:
289-317 (1994) for a general review of the properties of different
ribozymes).
[0644] The general features of hairpin ribozymes are described,
e.g., in Hampel et al., Nucl. Acids Res. 18:299-304 (1990);
European Patent Publication No. 0360257; U.S. Pat. No. 5,254,678.
Methods of preparing are well known to those of skill in the art
(see, e.g., WO 94/26877; Ojwang et al., Proc. Natl. Acad. Sci. USA
90:6340-6344 (1993); Yamada et al., Human Gene Therapy 1:39-45
(1994); Leavitt et al., Proc. Natl. Acad Sci. USA 92:699-703
(1995); Leavitt et al., Human Gene Therapy 5: 1151-120 (1994); and
Yamada et al., Virology 205: 121-126 (1994)).
[0645] Use of Modulators in Phenotypic Screening
[0646] In one embodiment, a test compound is administered to a
population of cancer cells, which have an associated cancer
expression profile. By "administration" or "contacting" herein is
meant that the modulator is added to the cells in such a manner as
to allow the modulator to act upon the cell, whether by uptake and
intracellular action, or by action at the cell surface. In some
embodiments, a nucleic acid encoding a proteinaceous agent (i.e., a
peptide) is put into a viral construct such as an adenoviral or
retroviral construct, and added to the cell, such that expression
of the peptide agent is accomplished, e.g., PCT US97/01019.
Regulatable gene therapy systems can also be used. Once the
modulator has been administered to the cells, the cells are washed
if desired and are allowed to incubate under preferably
physiological conditions for some period. The cells are then
harvested and a new gene expression profile is generated. Thus,
e.g., cancer tissue is screened for agents that modulate, e.g.,
induce or suppress, the cancer phenotype. A change in at least one
gene, preferably many, of the expression profile indicates that the
agent has an effect on cancer activity. Similarly, altering a
biological function or a signaling pathway is indicative of
modulator activity. By defining such a signature for the cancer
phenotype, screens for new drugs that alter the phenotype are
devised. With this approach, the drug target need not be known and
need not be represented in the original gene/protein expression
screening platform, nor does the level of transcript for the target
protein need to change. The modulator inhibiting function will
serve as a surrogate marker As outlined above, screens are done to
assess genes or gene products. That is, having identified a
particular differentially expressed gene as important in a
particular state, screening of modulators of either the expression
of the gene or the gene product itself is performed.
[0647] Use of Modulators to Affect Peptides of the Invention
[0648] Measurements of cancer polypeptide activity, or of the
cancer phenotype are performed using a variety of assays. For
example, the effects of modulators upon the function of a cancer
polypeptide(s) are measured by examining parameters described
above. A physiological change that affects activity is used to
assess the influence of a test compound on the polypeptides of this
invention. When the functional outcomes are determined using intact
cells or animals, a variety of effects can be assesses such as, in
the case of a cancer associated with solid tumors, tumor growth,
tumor metastasis, neovascularization, hormone release,
transcriptional changes to both known and uncharacterized genetic
markers (e.g., by Northern blots), changes in cell metabolism such
as cell growth or pH changes, and changes in intracellular second
messengers such as cGNIP.
[0649] Methods of Identifying Characterizing Cancer-associated
Sequences
[0650] Expression of various gene sequences is correlated with
cancer. Accordingly, disorders based on mutant or variant cancer
genes are determined. In one embodiment, the invention provides
methods for identifying cells containing variant cancer genes,
e.g., determining the presence of, all or part, the sequence of at
least one endogenous cancer gene in a cell. This is accomplished
using any number of sequencing techniques. The invention comprises
methods of identifying the cancer genotype of an individual, e.g.,
determining all or part of the sequence of at least one gene of the
invention in the individual. This is generally done in at least one
tissue of the individual, e.g., a tissue set forth in Table I, and
may include the evaluation of a number of tissues or different
samples of the same tissue. The method may include comparing the
sequence of the sequenced gene to a known cancer gene, i.e., a
wild-type gene to determine the presence of family members,
homologies, mutations or variants. The sequence of all or part of
the gene can then be compared to the sequence of a known cancer
gene to determine if any differences exist. This is done using any
number of known homology programs, such as BLAST, Bestfit, etc. The
presence of a difference in the sequence between the cancer gene of
the patient and the known cancer gene correlates with a disease
state or a propensity for a disease state, as outlined herein.
[0651] In a preferred embodiment, the cancer genes are used as
probes to determine the number of copies of the cancer gene in the
genome. The cancer genes are used as probes to determine the
chromosomal localization of the cancer genes. Information such as
chromosomal localization finds use in providing a diagnosis or
prognosis in particular when chromosomal abnormalities such as
translocations, and the like are identified in the cancer gene
locus.
[0652] XIV.) Kits/Articles of Manufacture
[0653] For use in the diagnostic and therapeutic applications
described herein, kits are also within the scope of the invention.
Such kits can comprise a carrier, package, or container that is
compartmentalized to receive one or more containers such as vials,
tubes, and the like, each of the container(s) comprising one of the
separate elements to be used in the method. For example, the
container(s) can comprise a probe that is or can be detectably
labeled. Such probe can be an antibody or polynucleotide specific
for a FIG. 2-related protein or a FIG. 2 gene or message,
respectively. Where the method utilizes nucleic acid hybridization
to detect the target nucleic acid, the kit can also have containers
containing nucleotide(s) for amplification of the target nucleic
acid sequence and/or a container comprising a reporter-means, such
as a biotin-binding protein, such as avidin or streptavidin, bound
to a reporter molecule, such as an enzymatic, florescent, or
radioisotope label. The kit can include all or part of the amino
acid sequences in FIG. 2 or FIG. 3 or analogs thereof, or a nucleic
acid molecules that encodes such amino acid sequences.
[0654] The kit of the invention will typically comprise the
container described above and one or more other containers
comprising materials desirable from a commercial and user
standpoint, including buffers, diluents, filters, needles,
syringes; carrier, package, container, vial and/or tube labels
listing contents and/or instructions for use, and package inserts
with instructions for use.
[0655] A label can be present on the container to indicate that the
composition is used for a specific therapy or non-therapeutic
application, such as a diagnostic or laboratory application, and
can also indicate directions for either in vivo or in vitro use,
such as those described herein. Directions and or other information
can also be included on an insert(s) or label(s) which is included
with or on the kit.
[0656] The terms "kit" and "article of manufacture" can be used as
synonyms.
[0657] In another embodiment of the invention, an article(s) of
manufacture containing compositions, such as amino acid
sequence(s), small molecule(s), nucleic acid sequence(s), and/or
antibody(s), e.g., materials useful for the diagnosis, prognosis,
prophylaxis and/or treatment of neoplasias of tissues such as those
set forth in Table I is provided. The article of manufacture
typically comprises at least one container and at least one label.
Suitable containers include, for example, bottles, vials, syringes,
and test tubes. The containers can be formed from a variety of
materials such as glass or plastic. The container can hold amino
acid sequence(s), small molecule(s), nucleic acid sequence(s),
and/or antibody(s), in one embodiment the container holds a
polynucleotide for use in examining the mRNA expression profile of
a cell, together with reagents used for this purpose.
[0658] The container can alternatively hold a composition which is
effective for treating, diagnosis, prognosing or prophylaxing a
condition and can have a sterile access port (for example the
container can be an intravenous solution bag or a vial having a
stopper pierceable by a hypodermic injection needle). The active
agents in the composition can be an antibody capable of
specifically binding 191P4D12(b) and modulating the function of
191P4D12(b).
[0659] The label can be on or associated with the container. A
label a can be on a container when letters, numbers or other
characters forming the label are molded or etched into the
container itself; a label can be associated with a container when
it is present within a receptacle or carrier that also holds the
container, e.g., as a package insert. The label can indicate that
the composition is used for diagnosing, treating, prophylaxing or
prognosing a condition, such as a neoplasia of a tissue set forth
in Table I. The article of manufacture can further comprise a
second container comprising a pharmaceutically-acceptable buffer,
such as phosphate-buffered saline, Ringer's solution and/ordextrose
solution. It can further include other materials desirable from a
commercial and user standpoint, including other buffers, diluents,
filters, stirrers, needles, syringes, and/or package inserts with
indications and/or instructions for use.
EXAMPLES
[0660] Various aspects of the invention are further described and
illustrated by way of the several examples that follow, none of
which are intended to limit the scope of the invention.
Example 1
SSH-Generated Isolation of cDNA Fragment of the 191P4D12(b)
Gene
[0661] To isolate genes that are over-expressed in prostate cancer
we used the Suppression Subtractive Hybridization (SSH) procedure
using cDNA derived from prostate cancer tissues. The 191P4D12(b)
SSH cDNA sequence was derived from bladder tumor minus cDNAs
derived from a pool of 9 normal tissues. The 191P4D12(b) cDNA was
identified as highly expressed in the bladder cancer.
Materials and Methods
[0662] Human Tissues:
[0663] The patient cancer and normal tissues were purchased from
different sources such as the NDRI (Philadelphia, Pa.). mRNA for
some normal tissues were purchased from Clontech, Palo Alto,
Calif.
[0664] RNA Isolation:
[0665] Tissues were homogenized in Trizol reagent (Life
Technologies, Gibco BRL) using 10 ml/g tissue isolate total RNA.
Poly A RNA was purified from total RNA using Qiagen's Oligotex mRNA
Mini and Midi kits. Total and mRNA were quantified by
spectrophotometric analysis (O.D. 260/280 nm) and analyzed by gel
electrophoresis.
[0666] Oligonucleotides:
[0667] The following HPLC purified oligonucleotides were used.
2 DPNGDN (cDNA synthesis primer): 5'TTTTGATCAAGCTT.sub.303- ' (SEQ
ID NO: 48) Adaptor 1: 5'CTMTACGACTCACTATAGGGCTCGAGCGGGCGCCCGGGCAG3'
(SEQ ID NO: 49) 3'GGCCCGTCCTAG5' (SEQ ID NO: 50) Adaptor 2:
5'GTAATACGACTCACTATAGGGCAGCGTGGTCGCGGCCGAG3' (SEQ ID NO: 51)
3'CGGCTCCTAG5' (SEQ ID NO: 52) POR primer 1:
5'CTAATACGACTCACTATAGGGC3' (SEQ ID NO: 53) Nested primer (NP)1:
5'TGGAGCGGCCGCCCGGGCAGGA3' (SEQ ID NO: 54) Nested primer (NP)2:
5'AGCGTGGTCGCGGGGGAGGA3' (SEQ ID NO: 55)
[0668] Suppression Subtractive Hybridization:
[0669] Suppression Subtractive Hybridization (SSH) was used to
identify cDNAs corresponding to genes that may be differentially
expressed in bladder cancer. The SSH reaction utilized cDNA from
bladder cancer and normal tissues.
[0670] The gene 191P4D12(b) sequence was derived from bladder
cancer minus normal tissue cDNA subtraction. The SSH DNA sequence
(FIG. 1) was identified.
[0671] The cDNA derived from of pool of normal tissues was used as
the source of the "driver" cDNA, while the cDNA from bladder cancer
was used as the source of the "tester" cDNA. Double stranded cDNAs
corresponding to tester and driver cDNAs were synthesized from 2
.mu.g of poly(A).sup.+ RNA isolated from the relevant xenograft
tissue, as described above, using CLONTECH's PCR-Select cDNA
Subtraction Kit and 1 ng of oligonucleotide DPNCDN as primer.
First- and second-strand synthesis were carried out as described in
the Kit's user manual protocol (CLONTECH Protocol No. PT1117-1,
Catalog No. K1804-1). The resulting cDNA was digested with Dpn II
for 3 hrs at 37.degree. C. Digested cDNA was extracted with
phenol/chloroform (1:1) and ethanol precipitated.
[0672] Driver cDNA was generated by combining in a 1:1 ratio Dpn II
digested cDNA from the relevant tissue source (see above) with a
mix of digested cDNAs derived from the nine normal tissues:
stomach, skeletal muscle, lung, brain, liver, kidney, pancreas,
small intestine, and heart.
[0673] Tester cDNA was generated by diluting 1 .mu.l of Dpn II
digested cDNA from the relevant tissue source (see above) (400 ng)
in 5 .mu.l of water. The diluted cDNA (2 .mu.l, 160 ng) was then
ligated to 2 .mu.l of Adaptor 1 and Adaptor 2 (10 .mu.M), in
separate ligation reactions, in a total volume of 10 .mu.l at
16.degree. C. overnight, using 400 u of T4 DNA ligase (CLONTECH).
Ligation was terminated with 1 .mu.l of 0.2 M EDTA and heating at
72.degree. C. for 5 min.
[0674] The first hybridization was performed by adding 1.5 .mu.l
(600 ng) of driver cDNA to each of two tubes containing 1.5 .mu.l
(20 ng) Adaptor 1- and Adaptor 2-ligated tester cDNA. In a final
volume of 4 .mu.l, the samples were overlaid with mineral oil,
denatured in an MJ Research thermal cycler at 98.degree. C. for 1.5
minutes, and then were allowed to hybridize for 8 hrs at 68.degree.
C. The two hybridizations were then mixed together with an
additional 1 .mu.l of fresh denatured driver cDNA and were allowed
to hybridize overnight at 68.degree. C. The second hybridization
was then diluted in 200 .mu.l of 20 mM Hepes, pH 8.3, 50 mM NaCl,
0.2 mM EDTA, heated at 70.degree. C. for 7 min. and stored at
-20.degree. C.
[0675] PCR Amplification, Cloning and Sequencing of Gene Fragments
Generated from SSH:
[0676] To amplify gene fragments resulting from SSH reactions, two
PCR amplifications were performed. In the primary PCR reaction 1
.mu.l of the diluted final hybridization mix was added to 1 .mu.l
of PCR primer 1 (10 .mu.M), 0.5 .mu.l dNTP mix (10 .mu.M), 2.5
.mu.l 10.times. reaction buffer (CLONTECH) and 0.5 .mu.l 50.times.
Advantage cDNA polymerase Mix (CLONTECH) in a final volume of 25
.mu.l. PCR 1 was conducted using the following conditions:
75.degree. C. for 5 min., 94.degree. C. for 25 sec., then 27 cycles
of 94.degree. C. for 10 sec, 66.degree. C. for 30 sec, 72.degree.
C. for 1.5 min. Five separate primary PCR reactions were performed
for each experiment. The products were pooled and diluted 1:10 with
water. For the secondary PCR reaction, 1 .mu.l from the pooled and
diluted primary PCR reaction was added to the same reaction mix as
used for PCR 1, except that primers NP1 and NP2 (10 .mu.M) were
used instead of PCR primer 1. PCR 2 was performed using 10-12
cycles of 94.degree. C. for 10 sec, 68.degree. C. for 30 sec, and
72.degree. C. for 1.5 minutes. The PCR products were analyzed using
2% agarose gel electrophoresis.
[0677] The PCR products were inserted into pCR2.1 using the T/A
vector cloning kit (Invitrogen). Transformed E. coli were subjected
to blue/white and ampicillin selection. White colonies were picked
and arrayed into 96 well plates and were grown in liquid culture
overnight. To identify inserts, PCR amplification was performed on
1 .mu.l of bacterial culture using the conditions of PCR1 and NP1
and NP2 as primers. PCR products were analyzed using 2% agarose gel
electrophoresis.
[0678] Bacterial clones were stored in 20% glycerol in a 96 well
format. Plasmid DNA was prepared, sequenced, and subjected to
nucleic acid homology searches of the GenBank, dBest, and NCI-CGAP
databases.
[0679] RT-PCR Expression Analysis:
[0680] First strand cDNAs can be generated from 1 .mu.g of mRNA
with oligo (dT)12-18 priming using the Gibco-BRL Superscript
Preamplification system. The manufacturer's protocol was used which
included an incubation for 50 min at 42.degree. C. with reverse
transcriptase followed by RNAse H treatment at 37.degree. C. for 20
min. After completing the reaction, the volume can be increased to
200 .mu.l with water prior to normalization. First strand cDNAs
from 16 different normal human tissues can be obtained from
Clontech.
[0681] Normalization of the first strand cDNAs from multiple
tissues was performed by using the primers
5'atatcgccgcgctcgtcgtcgacaa3' (SEQ ID NO: 56) and
5'agccacacgcagctcattgtagaagg 3' (SEQ ID NO: 57) to amplify
.beta.-actin. First strand cDNA (5 .mu.l) were amplified in a total
volume of 50 .mu.l containing 0.4 .mu.M primers, 0.2 .mu.M each
dNTPs, 1.times.PCR buffer (Clontech, 10 mM Tris-HCL, 1.5 mM
MgCl.sub.2, 50 mM KCl, pH8.3) and 1.times. Klentaq DNA polymerase
(Clontech). Five .mu.l of the POR reaction can be removed at 18,
20, and 22 cycles and used for agarose gel electrophoresis. PCR was
performed using an MJ Research thermal cycler under the following
conditions: Initial denaturation can be at 94.degree. C. for 15
sec, followed by a 18, 20, and 22 cycles of 94.degree. C. for 15,
65.degree. C. for 2 min, 72.degree. C. for 5 sec. A final extension
at 72.degree. C. was carried out for 2 min. After agarose gel
electrophoresis, the band intensities of the 283 b.p. .beta.-actin
bands from multiple tissues were compared by visual inspection.
Dilution factors for the first strand cDNAs were calculated to
result in equal .beta.-actin band intensities in all tissues after
22 cycles of PCR. Three rounds of normalization can be required to
achieve equal band intensities in all tissues after 22 cycles of
PCR.
[0682] To determine expression levels of the 191P4D12(b) gene, 5
.mu.l of normalized first strand cDNA were analyzed by PCR using
26, and 30 cycles of amplification. Semi-quantitative expression
analysis can be achieved by comparing the PCR products at cycle
numbers that give light band intensities. The primers used for
RT-PCR were designed using the 191P4D12(b) SSH sequence and are
listed below:
3 191P4012(b).1 5'-GGCTGGAGTTCAATGAGGTTTATTT-3' (SEQ ID NO: 58) 191
P4012(b)2 5'-TGCAGCAGATTTCAGACTAAGAAGA- -3' (SEQ ID NO: 59)
[0683] A typical RT-PCR expression analysis is shown in FIG. 14.
First strand cDNA was prepared from vital pool 1 (liver, lung and
kidney), vital pool 2 (pancreas, colon and stomach), normal kidney,
prostate cancer pool, bladder cancer pool, colon cancer pool, lung
cancer pool, breast cancer pool and cancer metastasis pool.
Normalization was performed by PCR using primers to actin and
GAPDH. Semi-quantitative PCR, using primers to 191P4D12(b), was
performed at 26 and 30 cycles of amplification. Results show strong
expression of 191P4D12(b) in bladder cancer pool. Expression of
191P4D12(b) was also detected in prostate cancer pool, colon cancer
pool, lung cancer pool, breast cancer pool and cancer metastasis
pool but very weakly in vital pool 1 and vital pool 2.
Example 2
Isolation of Full Length 191P4D12(b) Encoding cDNA
[0684] The 191P4D12(b) SSH cDNA sequence was derived from a
subtraction consisting of bladder cancer minus a mixture of 9
normal tissues: stomach, skeletal muscle, lung, brain, liver,
kidney, pancreas, small intestine and heart. The SSH cDNA sequence
of 223 bp (FIG. 1) was designated 191P4D12(b).
[0685] 191P4D12(b) v.1 (clone 1A1) of 3464 bp was cloned from
bladder cancer cDNA library, revealing an ORF of 510 amino acids
(FIG. 2 and FIG. 3). Other variants of 191P4D12(b) were also
identified and these are listed in FIGS. 2 and 3.
[0686] 191P4D12(b) v.1, v.2, v.10, v.11, and v.12 proteins are 510
amino acids in length and differ from each other by one amino acid
as shown in FIG. 11. 191P4D12(b) v.3, v.4, v.5, and v.8 code for
the same protein as 191P4D12(b) v.1. 191P4D12(b) v.6 and v.7 are
splice variants and code for proteins of 295 and 485 amino acids,
respectively. 191P4D12(b) v.13 clone 9C was cloned from bladder
cancer cDNA and has one amino acid insertion at position 334
compared to 191P4D12(b) v.1. 191P4D12(b) v.9 clone BCP1 is a splice
variant of 191P4D12(b) v.1 and was cloned from a bladder cancer
cDNA library. 191P4D12(b) v.14 is a SNP variant and differs from
191P4D12(b) v.9 by one amino acid as shown in FIG. 2.
[0687] 191P4D12(b) v.1 shows 99% identity over 2744 to the Ig
superfamily receptor LNIR (nectin-4), accession number
NM.sub.--030916. 191P4D12(b) v.9 protein is 100% identical to clone
AF218028 with function of inhibiting cancer cell growth.
Example 3
Chromosomal Mapping of 191P4D12(b)
[0688] Chromosomal localization can implicate genes in disease
pathogenesis. Several chromosome mapping approaches are available
including fluorescent in situ hybridization (FISH), human/hamster
radiation hybrid (RH) panels (Walter et al., 1994; Nature Genetics
7:22; Research Genetics, Huntsville Ala.), human-rodent somatic
cell hybrid panels such as is available from the Cornell Institute
(Camden, N.J.), and genomic viewers utilizing BLAST homologies to
sequenced and mapped genomic clones (NCBI, Bethesda, Md.).
[0689] 191P4D12(b) maps to chromosome 1q22-q23.2 using 191P4D12(b)
sequence and the NCBI BLAST tool located on the World Wide Web at
(.ncbi.nlm.nih.gov/genome/seq/page.cgi?F=HsBlast.html&&ORG=Hs).
Example 4
Expression Analysis of 191P4D12(b) in Normal Tissues and Patient
Specimens
[0690] Expression analysis by RT-PCR demonstrated that 191P4D12(b)
is strongly expressed in bladder cancer patient specimens (FIG.
14). First strand cDNA was prepared from (A) vital pool 1 (liver,
lung and kidney), vital pool 2 (pancreas, colon and stomach),
normal kidney, prostate cancer pool, bladder cancer pool, colon
cancer pool, lung cancer pool, breast cancer pool and cancer
metastasis pool; (B) prostate cancer metastasis to lymph node,
prostate cancer pool, bladder cancer pool, kidney cancer pool,
colon cancer pool, lung cancer pool, ovary cancer pool, breast
cancer pool, cancer metastasis pool, pancreas cancer pool, and LAPC
prostate xenograff pool. Normalization was performed by PCR using
primers to actin and GAPDH. Semi-quantitative PCR, using primers to
191P4D12(b), was performed at 26 and 30 cycles of amplification. In
(A), results show strong expression of 191P4D12(b) in bladder
cancer pool. Expression of 191P4D12(b) was also detected in
prostate cancer pool, colon cancer pool, lung cancer pool, breast
cancer pool and cancer metastasis pool but very weakly in vital
pool 1 and vital pool 2. In (B), results show strong expression of
191P4D12(b) in prostate, bladder, kidney, colon, lung, ovary,
breast, cancer metastasis, and pancreas cancer specimens.
[0691] Northern blot analysis of 251P5G2 is a technique known to
those skilled in the art to detect 251P5G2 protein production.
Northern blotting detects relative levels of mRNA expressed from a
251P5G2 gene. Specific mRNA is measured using a nucleic acid
hybridization technique and the signal is detected on an autoradiog
ram. The stronger the signal, the more abundant is the mRNA. For
251P5G2 genes that produce mRNA that contains an open reading frame
flanked by a good Kozak translation initiation site and a stop
codon, in the vast majority of cases the synthesized mRNA is
expressed as a protein.
[0692] The level of expression of the 251P5G2 gene is determined in
various normal tissues and in various tumor tissues and tumor cell
lines using the technique of Northern blotting, which detects
production of messenger RNA. It is well known in the art that the
production of messenger RNA, that encodes the protein, is a
necessary step in the production of the protein itself. Thus,
detection of high levels of messenger RNA by, for example, Northern
blot, is a way of determining that the protein itself is produced.
The Northern blot technique is used as a routine procedure because
it does not require the time delays (as compared to Western
blotting, immunoblotting or immunohistochemistry) involved in
isolating or synthesizing the protein, preparing an immunological
composition of the protein, eliciting a humoral immune response,
harvesting the antibodies, and verifying the specificity
thereof.
[0693] The Kozak consensus sequence for translation initiation
CCACCATGG, where the ATG start codon is noted, is the sequence with
the highest established probability of initiating translation. This
was confirmed by Peni and Pandey Trends in Genetics (2001) 17:
685-687. The conclusion is consistent with the general knowledge in
the art that, with rare exceptions, expression of an mRNA is
predictive of expression of its encoded protein. This is
particularly true for mRNA with an open reading frame and a Kozak
consensus sequence for translation initiation.
[0694] It is understood in the art that the absolute levels of
messenger RNA present and the amounts of protein produced do not
always provide a 1:1 correlation. In those instances where the
Northern blot has shown mRNA to be present, it is almost always
possible to detect the presence of the corresponding protein in the
tissue which provided a positive result in the Northern blot. The
levels of the protein compared to the levels of the mRNA may be
differential, but generally, cells that exhibit detectable mRNA
also exhibit detectable corresponding protein and vice versa. This
is particularly true where the mRNA has an open reading frame and a
good Kozak sequence (See, Peni and Pandey, supra.).
[0695] Occasionally those skilled in the art encounter a rare
occurrence where there is no detectable protein in the presence of
corresponding mRNA. (See, Fu, L., et al., Embo. Journal,
15:4392-4401 (1996)). In many cases, a reported lack of protein
expression is due to technical limitations of the protein detection
assay. These limitations are readily known to those skilled in the
art. These limitations include but are not limited to, available
antibodies that only detect denatured protein and not native
protein present in a cell and unstable proteins with very short
half-life. Short-lived proteins are still functional and have been
previously described to induce tumor formation. (See, e.g.,
Reinstein, et al., Oncogene, 19: 5944-5950). In such situations,
when more sensitive detection techniques are performed and/or other
antibodies are generated, protein expression is detected. When
studies fail to take these principles into account, they are likely
to report artifactually lowered correlations of mRNA to protein.
Outside of these rare exceptions the use of Northern blot analysis
is recognized to those skilled in the art to be predictive and
indicative of the detection of 251P5G2 protein production.
[0696] Extensive expression of 191P4D12(b) in normal tissues is
shown in FIG. 15. Two multiple tissue northern blots (Clontech)
both with 2 ug of mRNA/lane were probed with the 191P4D12(b)
sequence. Size standards in kilobases (kb) are indicated on the
side. Results show expression of an approximately 4 kb transcript
in placenta and very weakly in prostate but not in any other normal
tissue tested. A smaller 191P4D12(b) transcript of approximately
2.5 kb was detected in heart and skeletal muscle.
[0697] Expression of 191P4D12(b) in bladder cancer patient
specimens and human normal tissues is shown in FIG. 16. RNA was
extracted from a pool of 3 bladder cancer patient specimens, as
well as from normal prostate (NP), normal bladder (NB), normal
kidney (NK), normal colon (NC), normal lung (NL), normal breast
(NBr), normal ovary (NO), and normal pancreas (NPa). Northern blot
with 10 ug oftotal RNA/lane was probed with 191P4D12(b) SSH
sequence. Size standards in kilobases (kb) are indicated on the
side. The 191P4D12(b) transcript was detected in the bladder cancer
specimens, but not in the normal tissues tested.
[0698] Analysis of individual bladder cancer patient specimens is
depicted in FIG. 17. RNA was extracted from bladder cancer cell
lines (CL), normal bladder (N), and bladder cancer patient tumors
(T). Northern blots with 10 ug of total RNA were probed with the
191P4D12(b) SSH fragment. Size standards in kilobases are on the
side. Results show expression of the approximately 4 kb 191P4D12(b)
transcript in the bladder tumor tissues but not in normal bladder.
A smaller transcript was detected in the HT1197 cell line but not
in the other cancer cell lines tested.
[0699] Expression of 191P4D12(b) was also detected in prostate
cancer xenograft tissues (FIG. 18). RNA was extracted from normal
prostate, and from the prostate cancer xenograffs LAPC-4AD,
LAPC-4AI, LAPC-9AD, and LAPC-9AI. Northern blots with 10 ug of
total RNA were probed with the 191P4D12(b) SSH fragment. Size
standards in kilobases are on the side. Results show expression of
the approximately 4 kb 191P4D12(b) transcript in all the LAPC
xenograft tissues but not in normal prostate.
[0700] FIG. 19 shows expression of 191P4D12(b) in cervical cancer
patient specimens. RNA was extracted from normal cervix, Hela
cancer cell line, and 3 cervix cancer patient tumors (T). Northern
blots with 10 ug of total RNA were probed with the 191P4D12(b) SSH
fragment. Size standards in kilobases are on the side. Results show
expression of the approximately 4 kb 191P4D12(b) transcript in 2
out of 3 cervix tumors tested but not in normal cervix nor in the
Hela cell line.
[0701] 191P4D12(b) was also expressed in lung cancer patient
specimens (FIG. 20). RNA was extracted from lung cancer cell lines
(CL), normal lung (N), bladder cancer patient tumors (T), and
normal adjacent tissue (Nat). Northern blots with 10 ug of total
RNA were probed with the 191P4D12(b). Size standards in kilobases
are on the side. Results show expression of the approximately 4 kb
191P4D12(b) transcript in the lung tumor tissues but not in normal
lung nor in the cell lines tested.
[0702] 191P4D12(b) expression was tested in a panel of individual
patient cancer specimens (FIG. 21). First strand cDNA was prepared
from a panel of lung cancer specimens (A), bladder cancer specimens
(B), prostate cancer specimens (C), colon cancer specimens (D),
uterus cancer specimens (E), and cervix cancer specimens (F).
Normalization was performed by PCR using primers to actin.
Semi-quantitative PCR, using primers to 191P4D12(b) SSH fragment,
was performed at 26 and 30 cycles of amplification. Expression
level was recorded as 0=no expression detected; 1=weak expression,
2=moderate expression; 3=strong expression. Results show expression
of 191P4D12(b) in 97% of the 31 lung cancer patient specimens
tested, 94% of 18 bladder cancer patient specimens, 100% of 20
prostate cancer patient specimens, 100% of 22 colon cancer patient
specimens, 100% of 12 uterus cancer patient specimens, and 100% of
14 cervix cancer patient specimens tested.
[0703] The restricted expression of 191P4D12(b) in normal tissues
and the expression detected in cancer patient specimens suggest
that 191P4D12(b) is a potential therapeutic target and a diagnostic
marker for human cancers.
Example 5
Transcript Variants of 191P4D12(b)
[0704] Transcript variants are variants of mature mRNA from the
same gene which arise by alternative transcription or alternative
splicing. Alternative transcripts are transcripts from the same
gene but start transcription at different points. Splice variants
are mRNA variants spliced differently from the same transcript. In
eukaryotes, when a multi-exon gene is transcribed from genomic DNA,
the initial RNA is spliced to produce functional mRNA, which has
only exons and is used for translation into an amino acid sequence.
Accordingly, a given gene can have zero to many alternative
transcripts and each transcript can have zero to many splice
variants. Each transcript variant has a unique exon makeup, and can
have different coding and/or non-coding (5' or 3' end) portions,
from the original transcript. Transcript variants can code for
similar or different proteins with the same or a similar function
or can encode proteins with different functions, and can be
expressed in the same tissue at the same time, or in different
tissues at the same time, or in the same tissue at different times,
or in different tissues at different times. Proteins encoded by
transcript variants can have similar or different cellular or
extracellular localizations, e.g., secreted versus
intracellular.
[0705] Transcript variants are identified by a variety of
art-accepted methods. For example, alternative transcripts and
splice variants are identified by full-length cloning experiment,
or by use of full-length transcript and EST sequences. First, all
human ESTs were grouped into clusters which show direct or indirect
identity with each other. Second, ESTs in the same cluster were
further grouped into sub-clusters and assembled into a consensus
sequence. The original gene sequence is compared to the consensus
sequence(s) or other full-length sequences. Each consensus sequence
is a potential splice variant for that gene. Even when a variant is
identified that is not a full-length clone, that portion of the
variant is very useful for antigen generation and for further
cloning of the full-length splice variant, using techniques known
in the art.
[0706] Moreover, computer programs are available in the art that
identify transcript variants based on genomic sequences.
Genomic-based transcript variant identification programs include
FgenesH (A. Salamov and V. Solovyev, "Ab initio gene finding in
Drosophila genomic DNA," Genome Research. 2000 April;
10(4):516-22); Grail (URL compbio.ornl.gov/Grail-bi-
n/EmptyGrailForm) and GenScan (URL genes.mit.edu/GENSCAN.html). For
a general discussion of splice variant identification protocols
see., e.g., Southan, C., A genomic perspective on human proteases,
FEBS Lett. 2001 Jun 8; 498(2-3):214-8; de Souza, S. J., et al.,
Identification of human chromosome 22 transcribed sequences with
ORF expressed sequence tags, Proc. Natl Acad Sci U S A. 2000 Nov 7;
97(23):12690-3.
[0707] To further confirm the parameters of a transcript variant, a
variety of techniques are available in the art, such as full-length
cloning, proteomic validation, PCR-based validation, and 5' RACE
validation, etc. (see e.g., Proteomic Validation: Brennan, S. O.,
et al., Albumin banks peninsula: a new termination variant
characterized by electrospray mass spectrometry, Biochem Biophys
Acta. 1999 Aug 17;1433(1-2):321-6; Ferranti P, et al., Differential
splicing of pre-messenger RNA produces multiple forms of mature
caprine alpha(s1)-casein, Eur J Biochem. 1997 Oct 1;249(1):1-7. For
PCR-based Validation: Wellmann S, et al., Specific reverse
transcription-PCR quantification of vascular endothelial growth
factor (VEGF) splice variants by LightCycler technology, Clin Chem.
2001 Apr;47(4):654-60; Jia, H. P., et al., Discovery of new human
beta-defensins using a genomics-based approach, Gene. 2001 Jan 24;
263(1-2):211-8. For PCR-based and 5' RACE Validation: Brigle, K.
E., et al., Organization of the murine reduced folate carrier gene
and identification of variant splice forms, Biochem Biophys Acta.
1997 Aug 7;, 1353(2): 191-8).
[0708] It is known in the art that genomic regions are modulated in
cancers. When the genomic region to which a gene maps is modulated
in a particular cancer, the alternative transcripts or splice
variants of the gene are modulated as well. Disclosed herein is
that 191P4D12(b) has a particular expression profile related to
cancer. Alternative transcripts and splice variants of 191P4D12(b)
may also be involved in cancers in the same or different tissues,
thus serving as tumor-associated markers/antigens.
[0709] Using the full-length gene and EST sequences, four
additional transcript variants were identified, designated as
191P4D12(b) v.6, v.7, v.8 and v.9 as shown in FIG. 12. The
boundaries of exons in the original transcript, 191P4D12(b) v.1
were shown in Table LI. Compared with 191P4D12(b) v.1, variant v.6
spliced out 202-321 from the first exon of v.1 while variant v.8
spliced out 63 bases from the last exon of v.1. Variant v.7 spliced
out exon 8 of v.1. Variant 9 was part of the last exon of v.1.
Theoretically, each different combination of exons in spatial
order, e.g. exons 2, 3, 5, 7 and 9 of v.1, is a potential splice
variant. Tables LII (a)-(d) through LV (a)-(d) are set forth on a
variant-by-variant bases. Tables LII (a)-(d) shows nucleotide
sequence of the transcript variants. Tables LIII (a)-(d) shows the
alignment of the transcript variant with nucleic acid sequence of
191P4D12(b) v.1. Tables LIV (a)-(d) lays out amino acid translation
of the transcript variant for the identified reading frame
orientation. Tables LV (a)-(d) displays alignments of the amino
acid sequence encoded by the splice variant with that of
191P4D12(b) v.1.
Example 6
Single Nucleotide Polymorphisms of 191P4D12(b)
[0710] A Single Nucleotide Polymorphism (SNP) is a single base pair
variation in a nucleotide sequence at a specific location. At any
given point of the genome, there are four possible nucleotide base
pairs: A/T, C/G, G/C and T/A. Genotype refers to the specific base
pair sequence of one or more locations in the genome of an
individual. Haplotype refers to the base pair sequence of more than
one location on the same DNA molecule (or the same chromosome in
higher organisms), often in the context of one gene or in the
context of several tightly linked genes. SNP that occurs on a cDNA
is called cSNP. This cSNP may change amino acids of the protein
encoded by the gene and thus change the functions of the protein.
Some SNP cause inherited diseases; others contribute to
quantitative variations in phenotype and reactions to environmental
factors including diet and drugs among individuals. Therefore, SNP
and/or combinations of alleles (called haplotypes) have many
applications, including diagnosis of inherited diseases,
determination of drug reactions and dosage, identification of genes
responsible for diseases, and analysis of the genetic relationship
between individuals (P. Nowotny, J. M. Kwon and A. M. Goate, "SNP
analysis to dissect human traits," Curr. Opin. Neurobiol. 2001 Oct;
11(5):637-641; M. Pirmohamed and B. K. Park, "Genetic
susceptibility to adverse drug reactions," Trends Pharmacol. Sci.
2001 Jun; 22(6):298-305; J. H. Riley, C. J. Allan, E. Lai and A.
Roses, "The use of single nucleotide polymorphisms in the isolation
of common disease genes," Pharmacogenomics. 2000 Feb; 1(1):39-47;
R. Judson, J. C. Stephens and A. Windemuth, "The predictive power
of haplotypes in clinical response," Pharmacogenomics. 2000 feb;
1(1):15-26).
[0711] SNP are identified by a variety of art-accepted methods (P.
Bean, "The promising voyage of SNP target discovery," Am. Clin.
Lab. 2001 Oct-Nov; 20(9):18-20; K. M. Weiss, "In search of human
variation," Genome Res. 1998 Jul; 8(7):691-697; M. M. She,
"Enabling large-scale pharmacogenetic studies by high-throughput
mutation detection and genotyping technologies," Clin. Chem. 2001
Feb; 47(2):164-172). For example, SNP can be identified by
sequencing DNA fragments that show polymorphism by gel-based
methods such as restriction fragment length polymorphism (RFLP) and
denaturing gradient gel electrophoresis (DGGE). They can also be
discovered by direct sequencing of DNA samples pooled from
different individuals or by comparing sequences from different DNA
samples. With the rapid accumulation of sequence data in public and
private databases, one can discover SNP by comparing sequences
using computer programs (Z. Gu, L. Hillier and P. Y. Kwok, "Single
nucleotide polymorphism hunting in cyberspace," Hum. Mutat. 1998;
12(4):221-225). SNP can be verified and genotype or haplotype of an
individual can be determined by a variety of methods including
direct sequencing and high throughput microarrays (P. Y. Kwok,
"Methods for genotyping single nucleotide polymorphisms," Annu.
Rev. Genomics Hum. Genet. 2001; 2:235-258; M. Kokoris, K. Dix, K.
Moynihan, J. Mathis, B. Erwin, P. Grass, B. Hines and A.
Duesterhoeft, "High-throughput SNP genotyping with the Masscode
system," Mol. Diagn. 2000 Dec; 5(4):329-340). Using the methods
described above, seven SNP and one insertion/deletion of three
bases were identified in the original transcript, 191P4D12(b) v.1,
at positions 420 (T/C), 2184 (G/T), 2341 (G/A), 2688 (C/A), 367
(A/G), 699 (C/A), 1590 (C/T), and insertion of GCA in between 1262
and 1263I. The transcripts or proteins with alternative allele were
designated as variant 191P4D12(b) v.2 through v.5 and v.10 through
v.13, as shown in FIG. 10. FIG. 11 shows the schematic alignment of
protein variants, corresponding to nucleotide variants. Nucleotide
variants that code for the same amino acid sequence as v.1 are not
shown in FIG. 11. These alleles of the SNP, though shown separately
here, can occur in different combinations (haplotypes) and in any
one of the transcript variants (such as 191P4D12(b) v.9) that
contains the site of the SNP. The SNP at 2688 of v.1 occurs also in
transcript variant v.9 and contributed to one codon change of v.9
at amino acid 64 from Ala to Asp (FIG. 11).
Example 7
Production of Recombinant 191P4D12(b) in Prokaryotic Systems
[0712] To express recombinant 191P4D12(b) and 191P4D12(b) variants
in prokaryotic cells, the full or partial length 191P4D12(b) and
191P4D12(b) variant cDNA sequences are cloned into any one of a
variety of expression vectors known in the art. One or more of the
following regions of 191P4D12(b) variants are expressed: the full
length sequence presented in FIGS. 2 and 3, or any 8, 9, 10, 11,
12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28,
29, 30 or more contiguous amino acids from 191P4D12(b), variants,
or analogs thereof.
[0713] A. In vitro Transcription and Translation Constructs:
[0714] pCII: To generate 191P4D12(b) sense and anti-sense RNA
probes for RNA in situ investigations, pCRII constructs
(Invitrogen, Carlsbad Calif.) are generated encoding either all or
fragments of the 191P4D12(b) cDNA. The pCRII vector has Sp6 and T7
promoters flanking the insert to drive the transcription of
191P4D12(b) RNA for use as probes in RNA in situ hybridization
experiments. These probes are used to analyze the cell and tissue
expression of 191P4D12(b) at the RNA level. Transcribed 191P4D12(b)
RNA representing the cDNA amino acid coding region of the
191P4D12(b) gene is used in in vitro translation systems such as
the TnT.TM. Coupled Reticulolysate System (Promega, Corp., Madison,
Wis.) to synthesize 191P4D12(b) protein.
[0715] B. Bacterial Constructs:
[0716] pGEX Constructs: To generate recombinant 191P4D12(b)
proteins in bacteria that are fused to the Glutathione
S-transferase (GST) protein, all or parts of the 191P4D12(b) cDNA
protein coding sequence are cloned into the pGEX family of
GST-fusion vectors (Amersham Pharmacia Biotech, Piscataway, N.J.).
These constructs allow controlled expression of recombinant
191P4D12(b) protein sequences with GST fused at the amino-terminus
and a six histidine epitope (6X His) at the carboxyl-terminus. The
GST and 6X His tags permit purification of the recombinant fusion
protein from induced bacteria with the appropriate affinity matrix
and allow recognition of the fusion protein with anti-GST and
anti-His antibodies. The 6X His tag is generated by adding 6
histidine codons to the cloning primer at the 3' end, e.g., of the
open reading frame (ORF). A proteolytic cleavage site, such as the
PreScission.TM. recognition site in pGEX-6P-1, may be employed such
that it permits cleavage of the GST tag from 191P4D12(b)-related
protein. The ampicillin resistance gene and pBR322 origin permits
selection and maintenance of the pGEX plasmids in E. coli.
[0717] pMAL Constructs: To generate, in bacteria, recombinant
191P4D12(b) proteins that are fused to maltose-binding protein
(MBP), all or parts of the 191P4D12(b) cDNA protein coding sequence
are fused to the MBP gene by cloning into the pMAL-c2X and pMAL-p2X
vectors (New England Biolabs, Beverly, Mass.). These constructs
allow controlled expression of recombinant 191P4D12(b) protein
sequences with MBP fused at the amino-terminus and a 6X His epitope
tag at the carboxyl-terminus. The MBP and 6X His tags permit
purification of the recombinant protein from induced bacteria with
the appropriate affinity matrix and allow recognition of the fusion
protein with anti-MBP and anti-His antibodies. The 6X His epitope
tag is generated by adding 6 histidine codons to the 3' cloning
primer. A Factor Xa recognition site permits cleavage of the pMAL
tag from 191P4D12(b). The pMAL-c2X and pMAL-p2X vectors are
optimized to express the recombinant protein in the cytoplasm or
periplasm respectively. Periplasm expression enhances folding of
proteins with disulfide bonds.
[0718] pET Constructs: To express 191P4D12(b) in bacterial cells,
all or parts of the 191P4D12(b) cDNA protein coding sequence are
cloned into the pET family of vectors (Novagen, Madison, Wis.).
These vectors allow tightly controlled expression of recombinant
191P4D12(b) protein in bacteria with and without fusion to proteins
that enhance solubility, such as NusA and thioredoxin (Trx), and
epitope tags, such as 6X His and S-Tag.TM. that aid purification
and detection of the recombinant protein. For example, constructs
are made utilizing pET NusA fusion system 43.1 such that regions of
the 191P4D12(b) protein are expressed as amino-terminal fusions to
NusA.
[0719] C. Yeast Constructs:
[0720] pESC Constructs. To express 191P4D12(b) in the yeast species
Saccharomyces cerevisiae for generation of recombinant protein and
functional studies, all or parts of the 191P4D12(b) cDNA protein
coding sequence are cloned into the pESC family of vectors each of
which contain 1 of 4 selectable markers, HIS3, TRP1, LEU2, and URA3
(Stratagene, La Jolla, Calif.). These vectors allow controlled
expression from the same plasmid of up to 2 different genes or
cloned sequences containing either Flag.TM. or Myc epitope tags in
the same yeast cell. This system is useful to confirm
protein-protein interactions of 191P4D12(b). In addition,
expression in yeast yields similar post-translational
modifications, such as glycosylations and phosphorylations, that
are found when expressed in eukaryotic cells.
[0721] pESP Constructs: To express 191P4D12(b) in the yeast species
Saccharomyces pombe, all or parts of the 191P4D12(b) cDNA protein
coding sequence are cloned into the pESP family of vectors. These
vectors allow controlled high level of expression of a 191P4D12(b)
protein sequence that is fused at either the amino terminus or at
the carboxyl terminus to GST which aids purification of the
recombinant protein. A Flag.TM. epitope tag allows detection of the
recombinant protein with anti-Flag.TM. antibody.
Example 8
Production of Recombinant 191P4D12(b) in Higher Eukaryotic
Systems
[0722] A. Mammalian Constructs:
[0723] To express recombinant 191P4D12(b) in eukaryotic cells, the
full or partial length 191P4D12(b) cDNA sequences can be cloned
into any one of a variety of expression vectors known in the art.
One or more of the following regions of 191P4D12(b) are expressed
in these constructs, amino acids 1 to 510, or any 8, 9, 10, 11, 12,
13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,
30 or more contiguous amino acids from 191P4D12(b) v.1, v.2, v.10,
v.11, v.12; amino acids 1 to 511, or any 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30 or
more contiguous amino acids from 191P4D12(b) v.13, variants, or
analogs thereof.
[0724] The constructs can be transfected into any one of a wide
variety of mammalian cells such as 293T cells. Transfected 293T
cell lysates can be probed with the anti-191P4D12(b) polyclonal
serum, described herein.
[0725] pcDNA4/HisMax Constructs: To express 191P4D12(b) in
mammalian cells, a 191P4D12(b) ORF, or portions thereof, of
191P4D12(b) were cloned into pcDNA4/HisMax Version A (Invitrogen,
Carlsbad, Calif.). Protein expression is driven from the
cytomegalovirus (CMV) promoter and the SP16 translational enhancer.
The recombinant protein has Xpress.TM. and six histidine (6X His)
epitopes fused to the amino-terminus. The pcDNA4/HisMax vector also
contains the bovine growth hormone (BGH) polyadenylation signal and
transcription termination sequence to enhance mRNA stability along
with the SV40 origin for episomal replication and simple vector
rescue in cell lines expressing the large T antigen. The Zeocin
resistance gene allows for selection of mammalian cells expressing
the protein and the ampicillin resistance gene and ColE1 origin
permits selection and maintenance of the plasmid in E. coli.
[0726] pcDNA3.11/MycHis Constructs: To express 191P4D12(b) in
mammalian cells, a 191P4D12(b) ORF, or portions thereof, of
191P4D12(b) with a consensus Kozak translation initiation site was
cloned into pcDNA3.1/MycHis Version A (Invitrogen, Carlsbad,
Calif.). Protein expression is driven from the cytomegalovirus
(CMV) promoter. The recombinant proteins have the myc epitope and
6X His epitope fused to the carboxyl-terminus. The pcDNA3.1/MycHis
vector also contains the bovine growth hormone (BGH)
polyadenylation signal and transcription termination sequence to
enhance mRNA stability, along with the SV40 origin for episomal
replication and simple vector rescue in cell lines expressing the
large T antigen. The Neomycin resistance gene can be used, as it
allows for selection of mammalian cells expressing the protein and
the ampicillin resistance gene and ColE1 origin permits selection
and maintenance of the plasmid in E. coli. FIG. 22 shows expression
of 191P4D12(b).pcDNA3.1/MycHis following vector transfection into
293T cells. 293T cells were transfected with either
191P4D12(b).pcDNA3.1/mychi- s or pcDNA3.1/mychis vector control.
Forty hours later cell lysates were collected. Samples were run on
an SDS-PAGE acrylamide gel, blotted and stained with anti-his
antibody. The blot was developed using the ECL chemiluminescence
kit and visualized by autoradiography. Results show expression of
191P4D12(b) in the lysates of 191P4D12(b).pcDNA3.1/mychis
transfected cells (Lane 3), but not from the control
pcDNA3.1/mychis (Lane 4).
[0727] pcDNA3.1/CT-GFP-TOPO Construct: To express 191P4D12(b) in
mammalian cells and to allow detection of the recombinant proteins
using fluorescence, a 191P4D12(b) ORF, or portions thereof, with a
consensus Kozak translation initiation site are cloned into
pcDNA3.1/CT-GFP-TOPO (Invitrogen, Calif.). Protein expression is
driven from the cytomegalovirus (CMV) promoter. The recombinant
proteins have the Green Fluorescent Protein (GFP) fused to the
carboxyl-terminus facilitating non-invasive, in vivo detection and
cell biology studies. The pcDNA3.1CT-GFP-TOPO vector also contains
the bovine growth hormone (BGH) polyadenylation signal and
transcription termination sequence to enhance mRNA stability along
with the SV40 origin for episomal replication and simple vector
rescue in cell lines expressing the large T antigen. The Neomycin
resistance gene allows for selection of mammalian cells that
express the protein, and the ampicillin resistance gene and ColE1
origin permits selection and maintenance of the plasmid in E. coli.
Additional constructs with an amino-terminal GFP fusion are made in
pcDNA3.1/NT-GFP-TOPO spanning the entire length of a 191P4D12(b)
protein.
[0728] PAPtag: A 191P4D12(b) ORF, or portions thereof, is cloned
into pAPtag-5 (GenHunter Corp. Nashville, Tenn.). This construct
generates an alkaline phosphatase fusion at the carboxyl-terminus
of a 191P4D12(b) protein while fusing the IgG.kappa. signal
sequence to the amino-terminus. Constructs are also generated in
which alkaline phosphatase with an amino-terminal IgG.kappa. signal
sequence is fused to the amino-terminus of a 191P4D12(b) protein.
The resulting recombinant 191P4D12(b) proteins are optimized for
secretion into the media of transfected mammalian cells and can be
used to identify proteins such as ligands or receptors that
interact with 191P4D12(b) proteins. Protein expression is driven
from the CMV promoter and the recombinant proteins also contain myc
and 6X His epitopes fused at the carboxyl-terminus that facilitates
detection and purification. The Zeocin resistance gene present in
the vector allows for selection of mammalian cells expressing the
recombinant protein and the ampicillin resistance gene permits
selection of the plasmid in E. coli.
[0729] pTaq5: A 191P4D12(b) v.1 extracellular domain was cloned
into pTag-5 plasmid. This vector is similar to pAPtag but without
the alkaline phosphatase fusion. This construct generates
191P4D12(b) protein with an amino-terminal IgG.kappa. signal
sequence and myc and 6X His epitope tags at the carboxyl-terminus
that facilitate detection and affinity purification. The resulting
recombinant 191P4D12(b) protein is optimized for secretion into the
media of transfected mammalian cells, and is used as immunogen or
ligand to identify proteins such as ligands or receptors that
interact with the 191P4D12(b) proteins. Protein expression is
driven from the CMV promoter. The Zeocin resistance gene present in
the vector allows for selection of mammalian cells expressing the
protein, and the ampicillin resistance gene permits selection of
the plasmid in E. coli. FIG. 22 shows expression and secretion of
the extracellular domain of 191P4D12(b) following 191P4D12(b).pTag5
vector transfection into 293T cells. 293T cells were transfected
with 191P4D12(b).pTag5. Forty hours later, cell lysate and
supernatant were collected. Samples were run on an SDS-PAGE
acrylamide gel, blotted and stained with anti-his antibody. The
blot was developed using the ECL chemiluminescence kit and
visualized by autoradiography. Results show expression from
191P4D12(b).pTag5 plasmid of 191P4D12(b) extracellular domain in
the lysate (Lane 2) and secretion in the culture supernatant (Lane
1).
[0730] 191P4D12(b) ORF, or portions thereof, is also cloned into
pTag-5 plasmid.
[0731] PsecFc: A 191P4D12(b) ORF, or portions thereof, is also
cloned into psecFc. The psecFc vector was assembled by cloning the
human immunoglobulin G1 (IgG) Fc (hinge, CH2, CH3 regions) into
pSecTag2 (Invitrogen, Calif.). This construct generates an IgG1 Fc
fusion at the carboxyl-terminus of the 191P4D12(b) proteins, while
fusing the IgGK signal sequence to N-terminus. 191P4D12(b) fusions
utilizing the murine IgG1 Fc region are also used. The resulting
recombinant 191P4D12(b) proteins are optimized for secretion into
the media of transfected mammalian cells, and can be used as
immunogens or to identify proteins such as ligands or receptors
that interact with 191P4D12(b) protein. Protein expression is
driven from the GMV promoter. The hygromycin resistance gene
present in the vector allows for selection of mammalian cells that
express the recombinant protein, and the ampicillin resistance gene
permits selection of the plasmid in E. coli.
[0732] pSR.alpha. Constructs: To generate mammalian cell lines that
express 191P4D12(b) constitutively, 191P4D12(b) ORF, or portions
thereof, of 191P4D12(b) were cloned into pSR.alpha. constructs.
Amphotropic and ecotropic retroviruses were generated by
transfection of pSR.alpha. constructs into the 293T-10A1 packaging
line or co-transfection of pSR.alpha. and a helper plasmid
(containing deleted packaging sequences) into the 293 cells,
respectively. The retrovirus is used to infect a variety of
mammalian cell lines, resulting in the integration of the cloned
gene, 191P4D12(b), into the host cell-lines. Protein expression is
driven from a long terminal repeat (LTR). The Neomycin resistance
gene present in the vector allows for selection of mammalian cells
that express the protein, and the ampicillin resistance gene and
ColE1 origin permit selection and maintenance of the plasmid in E.
coli. The retroviral vectors can thereafter be used for infection
and generation of various cell lines using, for example, PC3, NIH
3T3, TsuPr1, 293 or rat-1 cells.
[0733] FIG. 23 shows stable expression of 191P4D12(b) following
191P4D12(b).pSRa transduction into 3T3 cells. 3T3 cells were
transduced with the pSRa retroviral vector encoding the 191P4D12(b)
gene. Following selection with neomycin, the cells were expanded
and RNA was extracted. Northern blot with 10 ug of total RNA/lane
was probed with the 191P4D12(b) SSH sequence. Size standards in
kilobases (kb) are indicated on the side. Results show expression
of the 191P4D12(b) transcript driven from the retroviral LTR, which
migrates slower than the endogenous 4kb 191P4D12(b) transcript
detected in the positive control LAPC-4AD.
[0734] Additional pSR.alpha. constructs are made that fuse an
epitope tag such as the FLAG.TM. tag to the carboxyl-terminus of
191P4D12(b) sequences to allow detection using anti-Flag
antibodies. For example, the FLAG.TM. sequence 5' gat tac aag gat
gac gac gat aag 3' (SEQ ID NO: 60) is added to cloning primer at
the 3' end of the ORF. Additional pSR.alpha. constructs are made to
produce both amino-terminal and carboxyl-terminal GFP and myc/6X
His fusion proteins of the full-length 191P4D12(b) proteins.
[0735] Additional Viral Vectors: Additional constructs are made for
viral-mediated delivery and expression of 191P4D12(b). High virus
titer leading to high level expression of 191P4D12(b) is achieved
in viral delivery systems such as adenoviral vectors and herpes
amplicon vectors. A 191P4D12(b) coding sequences or fragments
thereof are amplified by PCR and subcloned into the AdEasy shuttle
vector (Stratagene). Recombination and virus packaging are
performed according to the manufacturer's instructions to generate
adenoviral vectors. Alternatively, 191P4D12(b) coding sequences or
fragments thereof are cloned into the HSV-1 vector (Imgenex) to
generate herpes viral vectors. The viral vectors are thereafter
used for infection of various cell lines such as PC3, NIH 3T3, 293
or rat-1 cells.
[0736] Regulated Expression Systems: To control expression of
191P4D12(b) in mammalian cells, coding sequences of 191P4D12(b), or
portions thereof, are cloned into regulated mammalian expression
systems such as the T-Rex System (Invitrogen), the GeneSwitch
System (Invitrogen) and the tightly-regulated Ecdysone System
(Sratagene). These systems allow the study of the temporal and
concentration dependent effects of recombinant 191P4D12(b). These
vectors are thereafter used to control expression of 191P4D12(b) in
various cell lines such as PC3, NIH 3T3, 293 or rat-1 cells.
[0737] B. Baculovirus Expression Systems
[0738] To generate recombinant 191P4D12(b) proteins in a
baculovirus expression system, 191P4D12(b) ORF, or portions
thereof, are cloned into the baculovirus transfer vector pBlueBac
4.5 (Invitrogen), which provides a His-tag at the N-terminus.
Specifically, pBlueBac-191P4D12(b) is co-transfected with helper
plasmid pBac-N-Blue (Invitrogen) into SF9 (Spodoptera frugiperda)
insect cells to generate recombinant baculovirus (see Invitrogen
instruction manual for details). Baculovirus is then collected from
cell supernatant and purified by plaque assay.
[0739] Recombinant 191P4D12(b) protein is then generated by
infection of HighFive insect cells (Invitrogen) with purified
baculovirus. Recombinant 191P4D12(b) protein can be detected using
anti-191P4D12(b) or anti-His-tag antibody. 191P4D12(b) protein can
be purified and used in various cell-based assays or as immunogen
to generate polyclonal and monoclonal antibodies specific for
191P4D12(b).
Example 9
Antigenicity Profiles and Secondary Structure
[0740] FIGS. 5(A-C), FIGS. 6(A-C), FIGS. 7(A-E), FIGS. 8(A-C), and
FIGS. 9(A-C) depict graphically five amino acid profiles of
191P4D12(b) variants 1, 7, and 9, each assessment available by
accessing the ProtScale website located on the World Wide Web at
(.expasy.ch/cgi-bin/protscale.pl) on the ExPasy molecular biology
server.
[0741] These profiles: FIG. 5, Hydrophilicity, (Hopp T. P., Woods
K. R., 1981. Proc. Natl. Acad. Sci. U.S.A. 78:3824-3828); FIG. 6,
Hydropathicity, (Kyte J., Doolittle R. F., 1982. J. Mol. Biol.
157:105-132); FIG. 7, Percentage Accessible Residues (Janin J.,
1979 Nature 277:491-492); FIG. 8, Average Flexibility, (Bhaskaran
R., and Ponnuswamy P. K., 1988. Int. J. Pept. Protein Res.
32:242-255); FIG. 9, Beta-turn (Deleage, G., Roux B. 1987 Protein
Engineering 1:289-294); and optionally others available in the art,
such as on the ProtScale website, were used to identify antigenic
regions of each of the 191P4D12(b) variant proteins. Each of the
above amino acid profiles of 191P4D12(b) variants were generated
using the following ProtScale parameters for analysis: 1) A window
size of 9; 2) 100% weight of the window edges compared to the
window center; and, 3) amino acid profile values normalized to lie
between 0 and 1.
[0742] Hydrophilicity (FIG. 5), Hydropathicity (FIG. 6) and
Percentage Accessible Residues (FIG. 7) profiles were used to
determine stretches of hydrophilic amino acids (i.e., values
greater than 0.5 on the Hydrophilicity and Percentage Accessible
Residues profile, and values less than 0.5 on the Hydropathicity
profile). Such regions are likely to be exposed to the aqueous
environment, be present on the surface of the protein, and thus
available for immune recognition, such as by antibodies.
[0743] Average Flexibility (FIG. 8) and Beta-turn (FIG. 9) profiles
determine stretches of amino acids (i.e., values greater than 0.5
on the Beta-turn profile and the Average Flexibility profile) that
are not constrained in secondary structures such as beta sheets and
alpha helices. Such regions are also more likely to be exposed on
the protein and thus accessible to immune recognition, such as by
antibodies.
[0744] Antigenic sequences of the 191P4D12(b) variant proteins
indicated, e.g., by the profiles set forth in FIGS. 5(A-C), FIGS.
6(A-C), FIGS. 7(A-C), FIGS. 8(A-C), and/or FIGS. 9(A-C) are used to
prepare immunogens, either peptides or nucleic acids that encode
them, to generate therapeutic and diagnostic anti-191P4D12(b)
antibodies. The immunogen can be any 5, 6, 7, 8, 9, 10, 11, 12, 13,
14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 30, 35, 40, 45, 50
or more than 50 contiguous amino acids, or the corresponding
nucleic acids that encode them, from the 191P4D12(b) protein
variants listed in FIGS. 2 and 3, of which the amino acid profiles
are shown in FIG. 9, or are identical to the variant sequences that
are the same as a variant depicted in FIG. 9. In particular,
peptide immunogens of the invention can comprise, a peptide region
of at least 5 amino acids of FIGS. 2 and 3 in any whole number
increment that includes an amino acid position having a value
greater than 0.5 in the Hydrophilicity profiles of FIG. 5; a
peptide region of at least 5 amino acids of FIGS. 2 and 3 in any
whole number increment that includes an amino acid position having
a value less than 0.5 in the Hydropathicity profile of FIG. 6; a
peptide region of at least 5 amino acids of FIGS. 2 and 3 in any
whole number increment that includes an amino acid position having
a value greater than 0.5 in the Percent Accessible Residues
profiles of FIG. 7; a peptide region of at least 5 amino acids of
FIGS. 2 and 3 in any whole number increment that includes an amino
acid position having a value greater than 0.5 in the Average
Flexibility profiles on FIG. 8 and, a peptide region of at least 5
amino acids of FIGS. 2 and 3 in any whole number increment that
includes an amino acid position having a value greater than 0.5 in
the Beta-turn profile of FIG. 9. Peptide immunogens of the
invention can also comprise nucleic acids that encode any of the
forgoing.
[0745] All immunogens of the invention, peptide or nucleic acid,
can be embodied in human unit dose form, or comprised by a
composition that includes a pharmaceutical excipient compatible
with human physiology.
[0746] The secondary structure of 191P4D12(b) protein variants 1,
7, and 9, namely the predicted presence and location of alpha
helices, extended strands, and random coils, is predicted from the
primary amino acid sequence using the HNN--Hierarchical Neural
Network method (Guermeur, 1997,
http://pbil.ibcp.fr/cgi-bin/npsa_automat.pl?page=npsa_nn.html),
accessed from the ExPasy molecular biology server located on the
World Wide Web at (.expasy.ch/tools/). The analysis indicates that
191P4D12(b) variant 1 is composed of 24.90% alpha helix, 18.63%
extended strand, and 56.47% random coil (FIG. 13A). Variant 6 is
composed of 28.47% alpha helix, 19.32% extended strand, and 52.20%
random coil (FIG. 13B). Variant 7 is composed of 26.19% alpha
helix, 18.76% extended strand, and 55.05% random coil (FIG. 13C).
Variant 7 is composed of 56.20% alpha helix, 8.76% extended strand,
and 35.04% random coil (FIG. 13D).
[0747] Analysis for the potential presence of transmembrane domains
in the 191P4D12(b) variant proteins was carried out using a variety
of transmembrane prediction algorithms accessed from the ExPasy
molecular biology server located on the World Wide Web at
(.expasy.ch/tools/). Shown graphically in FIGS. 13E and 13F are the
results of analysis of variant 1 depicting the presence and
location of 1 transmembrane domain using the TMpred program (FIG.
13E) and 1 transmembrane domain using the TMHMM program (FIG. 13F).
Shown graphically in FIGS. 13G and 13H are the results of analysis
of variant 6 depicting the presence and location of 1 transmembrane
domains using the TMpred program (FIG. 13G) and 1 transmembrane
domain using the TMHMM program (FIG. 13H). Shown graphically in
FIG. 13I and 13J are the results of analysis of variant 7 depicting
the presence and location of 1 transmembrane domain using the
TMpred program (FIG. 13I) and 1 transmembrane domain using the
TMHMM program (FIG. 13J). Shown graphically in FIG. 13K and 13L are
the results of analysis of variant 9 depicting the presence and
location of 2 transmembrane domains using the TMpred program (FIG.
1K) and 1 transmembrane domain using the TMHMM program (FIG. 13L).
The results of each program, namely the amino acids encoding the
transmembrane domains are summarized in Table VI and Table L.
Example 10
Generation of 191P4D12(b) Polyclonal Antibodies
[0748] Polyclonal antibodies can be raised in a mammal, for
example, by one or more injections of an immunizing agent and, if
desired, an adjuvant. Typically, the immunizing agent and/or
adjuvant will be injected in the mammal by multiple subcutaneous or
intraperitoneal injections. In addition to immunizing with a full
length 191P4D12(b) protein variant, computer algorithms are
employed in design of immunogens that, based on amino acid sequence
analysis contain characteristics of being antigenic and available
for recognition by the immune system of the immunized host (see the
Example entitled "Antigenicity Profiles and Secondary Structures").
Such regions would be predicted to be hydrophilic, flexible, in
beta-turn conformations, and be exposed on the surface of the
protein (see, e.g., FIGS. 5(A-C), FIGS. 6(A & C), FIGS. 7(A-C),
FIGS. 8(A-C), or FIGS. 9(A-C) for amino acid profiles that indicate
such regions of 191P4D12(b) protein variants).
[0749] For example, recombinant bacterial fusion proteins or
peptides containing hydrophilic, flexible, beta-turn regions of
191P4D12(b) protein variants are used as antigens to generate
polyclonal antibodies in New Zealand White rabbits or monoclonal
antibodies as described in Example 11. For example, in 191P4D12(b)
variant 1, such regions include, but are not limited to, amino
acids 27-39, amino acids 93-109, and amino acids 182-204. In
sequence unique to variant 7, such regions include, but are not
limited to, amino acids 400-420. In sequence specific for variant
9, such regions include, but are not limited to, amino acids 80-94.
It is useful to conjugate the immunizing agent to a protein known
to be immunogenic in the mammal being immunized. Examples of such
immunogenic proteins include, but are not limited to, keyhole
limpet hemocyanin (KLH), serum albumin, bovine thyroglobulin, and
soybean trypsin inhibitor. In one embodiment, a peptide encoding
amino acids 52-63 of 191P4D12(b) variant 1 and amino acids 179-197
were each conjugated to KLH and used to immunize separate rabbits.
Alternatively the immunizing agent may include all or portions of
the 191P4D12(b) variant proteins, analogs or fusion proteins
thereof. For example, the 191P4D12(b) variant 1 amino acid sequence
can be fused using recombinant DNA techniques to any one of a
variety of fusion protein partners that are well known in the art,
such as glutathione-S-transferase (GST) and HIS tagged fusion
proteins. In another embodiment, amino acids 2-349 of 191P4D12(b)
variant 1 was fused to GST using recombinant techniques and the
pGEX expression vector, expressed, purified and used to immunize a
rabbit. Such fusion proteins are purified from induced bacteria
using the appropriate affinity matrix.
[0750] Other recombinant bacterial fusion proteins that may be
employed include maltose binding protein, LacZ, thioredoxin, NusA,
or an immunoglobulin constant region (see the section entitled
"Production of 191P4D12(b) in Prokaryotic Systems" and Current
Protocols In Molecular Biology, Volume 2, Unit 16, Frederick M.
Ausubul et al. eds., 1995; Linsley, P. S., Brady, W., Urnes, M.,
Grosmaire, L., Damle, N., and Ledbetter, L.(1991) J.Exp. Med. 174,
561-566).
[0751] In addition to bacterial derived fusion proteins, mammalian
expressed protein antigens are also used. These antigens are
expressed from mammalian expression vectors such as the Tag5 and
Fc-fusion vectors (see the section entitled "Production of
Recombinant 191P4D12(b) in Eukaryotic Systems"), and retain
post-translational modifications such as glycosylations found in
native protein. In one embodiment, amino acids 31-347 of variant 1,
encoding the extracellular domain, was cloned into the Tag5
mammalian secretion vector, and expressed in 293T cells resulting
in a soluble secreted protein (FIG. 22). The recombinant protein is
purified by metal chelate chromatography from tissue culture
supernatants of 293T cells stably expressing the recombinant
vector. The purified Tag5 191 P4D12(b) protein is then used as
immunogen.
[0752] During the immunization protocol, it is useful to mix or
emulsify the antigen in adjuvants that enhance the immune response
of the host animal. Examples of adjuvants include, but are not
limited to, complete Freund's adjuvant (CFA) and MPL-TDM adjuvant
(monophosphory Lipid A, synthetic trehalose dicorynomycolate).
[0753] In a typical protocol, rabbits are initially immunized
subcutaneously with up to 200 .mu.g, typically 100-200 .mu.g, of
fusion protein or peptide conjugated to KLH mixed in complete
Freund's adjuvant (CFA). Rabbits are then injected subcutaneously
every two weeks with up to 200 .mu.g, typically 100-200 .mu.g, of
the immunogen in incomplete Freund's adjuvant (IFA). Test bleeds
are taken approximately 7-10 days following each immunization and
used to monitor the titer of the antiserum by ELISA.
[0754] To test reactivity and specificity of immune serum, such as
the rabbit serum derived from immunization with the Tag5
-191P4D12(b) variant 1 protein, the full-length 191P4D12(b) variant
1 cDNA is cloned into pCDNA 3.1 myc-his expression vector
(Invitrogen, see the Example entitled "Production of Recombinant
191P4D12(b) in Eukaryotic Systems"). After transfection of the
constructs into 293T cells, cell lysates are probed with the
anti-191P4D12(b) serum and with anti-His antibody (Santa Cruz
Biotechnologies, Santa Cruz, Calif.) to determine specific
reactivity to denatured 191P4D12(b) protein using the Western blot
technique. In addition, the immune serum is tested by fluorescence
microscopy, flow cytometry and immunoprecipitation against 293T
(FIG. 22) and other recombinant 191P4D12(b)-expressing cells to
determine specific recognition of native protein. Western blot,
immunoprecipitation, fluorescent microscopy, and flow cytometric
techniques using cells that endogenously express 191P4D12(b) are
also carried out to test reactivity and specificity. Anti-serum
from rabbits immunized with 191P4D12(b) variant fusion proteins,
such as GST and MBP fusion proteins, are purified by depletion of
antibodies reactive to the fusion partner sequence by passage over
an affinity column containing the fusion partner either alone or in
the context of an irrelevant fusion protein. For example, antiserum
derived from a GST-191P4D12(b) variant 1 fusion protein is first
purified by passage over a column of GST protein covalently coupled
to AffiGel matrix (BioRad, Hercules, Calif.). The antiserum is then
affinity purified by passage over a column composed of a
MBP-191P4D12(b) fusion protein covalently coupled to Affigel
matrix. The serum is then further purified by protein G affinity
chromatography to isolate the IgG fraction. Sera from other
His-tagged antigens and peptide immunized rabbits as well as fusion
partner depleted sera are affinity purified by passage over a
column matrix composed of the original protein immunogen or free
peptide.
Example 11
Generation of 191P4D12(b) Monoclonal Antibodies (mAbs)
[0755] In one embodiment, therapeutic mAbs to 191P4D12(b) variants
comprise those that react with epitopes specific for each variant
protein or specific to sequences in common between the variants
that would disrupt or modulate the biological function of the
191P4D12(b) variants, for example those that would disrupt the
interaction with ligands and binding partners. Immunogens for
generation of such mAbs include those designed to encode or contain
the entire 191P4D12(b) protein variant sequence, regions of the
191P4D12(b) protein variants predicted to be antigenic from
computer analysis of the amino acid sequence (see, e.g., FIGS.
5(A-C), FIGS. 6(A-C), FIGS. 7(A-C), FIGS. 8(A-C), or FIGS. 9(A-C),
and the Example entitled "Antigenicity Profiles"). Immunogens
include peptides, recombinant bacterial proteins, and mammalian
expressed Tag 5 proteins and human and murine IgG FC fusion
proteins. In addition, cells engineered to express high levels of a
respective 191P4D12(b) variant, such as 293T-191P4D12(b) variant 1
or 300.19-191P4D12(b) variant 1murine Pre-B cells, are used to
immunize mice.
[0756] To generate mAbs to a 191P4D12(b) variant, mice are first
immunized intraperitoneally (IP) with, typically, 10-50 .mu.g of
protein immunogen or 10.sup.7 191P4D12(b)-expressing cells mixed in
complete Freund's adjuvant. Mice are then subsequently immunized IP
every 2-4 weeks with, typically, 10-50 .mu.g of protein immunogen
or 10.sup.7 cells mixed in incomplete Freund's adjuvant.
Alternatively, MPL-TDM adjuvant is used in immunizations. In
addition to the above protein and cell-based immunization
strategies, a DNA-based immunization protocol is employed in which
a mammalian expression vector encoding a 191P4D12(b) variant
sequence is used to immunize mice by direct injection of the
plasmid DNA. For example, amino acids 31-347 was cloned into the
Tag5 mammalian secretion vector and the recombinant vector will
then be used as immunogen. In another example the same amino acids
are cloned into an Fc-fusion secretion vector in which the
191P4D12(b) variant 1 sequence is fused at the amino-terminus to an
IgK leader sequence and at the carboxyl-terminus to the coding
sequence of the human or murine IgG Fc region. This recombinant
vector is then used as immunogen. The plasmid immunization
protocols are used in combination with purified proteins expressed
from the same vector and with cells expressing the respective
191P4D12(b) variant.
[0757] During the immunization protocol, test bleeds are taken 7-10
days following an injection to monitor titer and specificity of the
immune response. Once appropriate reactivity and specificity is
obtained as determined by ELISA, Western blotting,
immunoprecipitation, fluorescence microscopy, and flow cytometric
analyses, fusion and hybridoma generation is then carried out with
established procedures well known in the art (see, e.g., Harlow and
Lane, 1988).
[0758] In one embodiment for generating 191P4D12(b) monoclonal
antibodies, a Tag5-191P4D12(b) variant 1 antigen encoding amino
acids 31-347, was expressed (FIG. 22) and then purified from stably
transfected 293T cells. Balb C mice are initially immunized
intraperitoneally with 25 .mu.g of the Tag5-191P4D12(b) variant 1
protein mixed in complete Freund's adjuvant. Mice are subsequently
immunized every two weeks with 25 .mu.g of the antigen mixed in
incomplete Freund's adjuvant for a total of three immunizations.
ELISA using the Tag5 antigen determines the titer of serum from
immunized mice. Reactivity and specificity of serum to full length
191P4D12(b) variant 1 protein is monitored by Western blotting,
immunoprecipitation and flow cytometry using 293T cells transfected
with an expression vector encoding the 191P4D12(b) variant 1 cDNA
(see e.g., the Example entitled "Production of Recombinant
191P4D12(b) (a) & (b) in Eukaryotic Systems" and FIG. 22).
Other recombinant 191P4D12(b) variant 1-expressing cells or cells
endogenously expressing 191P4D12(b) variant 1 are also used. Mice
showing the strongest reactivity are rested and given a final
injection of Tag5 antigen in PBS and then sacrificed four days
later. The spleens of the sacrificed mice are harvested and fused
to SPO/2 myeloma cells using standard procedures (Harlow and Lane,
1988). Supernatants from HAT selected growth wells are screened by
ELISA, Western blot, immunoprecipitation, fluorescent microscopy,
and flow cytometry to identify 191P4D12(b) specific
antibody-producing clones.
[0759] To generate monoclonal antibodies that are specific for each
191P4D12(b) variant protein, immunogens are designed to encode
sequences unique for each variant. In one embodiment, a GST-fusion
antigen encoding the full sequence of 191P4D12(b) variant 9 (AA
1-137) is produced, purified, and used as immunogen to derive
monoclonal antibodies specific to 191P4D12(b) variant 2. In another
embodiment, an antigenic peptide composed of amino acids 400-420 of
191P4D12(b) variant 7 is coupled to KLH and used as immunogen.
Hybridoma supernatants are then screened on the respective antigen
and then further screened on cells expressing the specific variant
and cross-screened on cells expressing the other variants to derive
variant-specific monoclonal antibodies.
[0760] The binding affinity of a 191P4D12(b) variant monoclonal
antibody is determined using standard technologies. Affinity
measurements quantify the strength of antibody to epitope binding
and are used to help define which 191P4D12(b) variant monoclonal
antibodies preferred for diagnostic or therapeutic use, as
appreciated by one of skill in the art. The BIAcore system
(Uppsala, Sweden) is a preferred method for determining binding
affinity. The BIAcore system uses surface plasmon resonance (SPR,
Welford K. 1991, Opt. Quant. Elect. 23:1; Morton and Myszka, 1998,
Methods in Enzymology 295: 268) to monitor biomolecular
interactions in real time. BIAcore analysis conveniently generates
association rate constants, dissociation rate constants,
equilibrium dissociation constants, and affinity constants.
Example 12
HLA Class I and Class II Binding Assays
[0761] HLA class I and class II binding assays using purified HLA
molecules are performed in accordance with disclosed protocols
(e.g., PCT publications WO 94/20127 and WO 94/03205; Sidney et al.,
Current Protocols in Immunology 18.3.1 (1998); Sidney, et al., J.
Immunol. 154:247 (1995); Sette, et al., Mol. Immunol. 31:813
(1994)). Briefly, purified MHC molecules (5 to 500 nM) are
incubated with various unlabeled peptide inhibitors and 1-10 nM
.sup.125I-radiolabeled probe peptides as described. Following
incubation, MHC-peptide complexes are separated from free peptide
by gel filtration and the fraction of peptide bound is determined.
Typically, in preliminary experiments, each MHC preparation is
titered in the presence of fixed amounts of radiolabeled peptides
to determine the concentration of HLA molecules necessary to bind
10-20% of the total radioactivity. All subsequent inhibition and
direct binding assays are performed using these HLA
concentrations.
[0762] Since under these conditions [label]<[HLA] and
IC.sub.50.gtoreq.[HLA], the measured IC.sub.50 values are
reasonable approximations of the true K.sub.D values. Peptide
inhibitors are typically tested at concentrations ranging from 120
.mu.g/ml to 1.2 ng/ml, and are tested in two to four completely
independent experiments. To allow comparison of the data obtained
in different experiments, a relative binding figure is calculated
for each peptide by dividing the IC.sub.50 of a positive control
for inhibition by the IC.sub.50 for each tested peptide (typically
unlabeled versiors of the radiolabeled probe peptide). For database
purposes, and inter-experiment comparisons, relative binding values
are compiled. These values can subsequently be converted back into
IC.sub.50 nM values by dividing the IC.sub.50 nM of the positive
controls for inhibition by the relative binding of the peptide of
interest. This method of data compilation is accurate and
consistent for comparing peptides that have been tested on
different days, or with different lots of purified MHC.
[0763] Binding assays as outlined above may be used to analyze HLA
supermotif and/or HLA motif-bearing peptides (see Table IV).
Example 13
Identification of HLA Supermotif- and Motif-Bearing CTL Candidate
Epitopes
[0764] HLA vaccine compositions of the invention can include
multiple epitopes. The multiple epitopes can comprise multiple HLA
supermotifs or motifs to achieve broad population coverage. This
example illustrates the identification and confirmation of
supermotif- and motif-bearing epitopes for the inclusion in such a
vaccine composition. Calculation of population coverage is
performed using the strategy described below.
[0765] Computer Searches and Algorithms for Identification of
Supermotif and/or Motif-bearing Epitomes
[0766] The searches performed to identify the motif-bearing peptide
sequences in the Example entitled "Antigenicity Profiles" and
Tables VIII-XXI and XXII-XLIX employ the protein sequence data from
the gene product of 191P4D12(b) set forth in FIGS. 2 and 3, the
specific search peptides used to generate the tables are listed in
Table VII.
[0767] Computer searches for epitopes bearing HLA Class I or Class
II supermotifs or motifs are performed as follows. All translated
191P4D12(b) protein sequences are analyzed using a text string
search software program to identify potential peptide sequences
containing appropriate HLA binding motifs; such programs are
readily produced in accordance with information in the art in view
of known motif/supermotif disclosures. Furthermore, such
calculations can be made mentally.
[0768] Identified A2-, A3-, and DR-supermotif sequences are scored
using polynomial algorithms to predict their capacity to bind to
specific HLA-Class I or Class II molecules. These polynomial
algorithms account for the impact of different amino acids at
different positions, and are essentially based on the premise that
the overall affinity (or AG) of peptide-HLA molecule interactions
can be approximated as a linear polynomial function of the
type:
".DELTA.G"=a.sub.1i.times.a.sub.2i.times.a.sub.3i . . .
.times.a.sub.ni
[0769] where a.sub.ji is a coefficient which represents the effect
of the presence of a given amino acid (j) at a given position (i)
along the sequence of a peptide of n amino acids. The crucial
assumption of this method is that the effects at each position are
essentially independent of each other (i.e., independent binding of
individual side-chains). When residue j occurs at position i in the
peptide, it is assumed to contribute a constant amount j.sub.i to
the free energy of binding of the peptide irrespective of the
sequence of the rest of the peptide.
[0770] The method of derivation of specific algorithm coefficients
has been described in Gulukota et al., J. Mol. Biol. 267:1258-126,
1997; (see also Sidney et al., Human Immunol. 45:79-93, 1996; and
Southwood et al., J. Immunol. 160:3363-3373, 1998). Briefly, for
all i positions, anchor and non-anchor alike, the geometric mean of
the average relative binding (ARB) of all peptides carrying j is
calculated relative to the remainder of the group, and used as the
estimate of j.sub.i. For Class II peptides, if multiple alignments
are possible, only the highest scoring alignment is utilized,
following an iterative procedure. To calculate an algorithm score
of a given peptide in a test set, the ARB values corresponding to
the sequence of the peptide are multiplied. If this product exceeds
a chosen threshold, the peptide is predicted to bind. Appropriate
thresholds are chosen as a function of the degree of stringency of
prediction desired.
[0771] Selection of HLA-A2 Supertype Cross-reactive Peptides
[0772] Protein sequences from 191P4D12(b) are scanned utilizing
motif identification software, to identify 8-, 9-10- and 11-mer
sequences containing the HLA-A2-supermotif main anchor specificity.
Typically, these sequences are then scored using the protocol
described above and the peptides corresponding to the
positive-scoring sequences are synthesized and tested for their
capacity to bind purified HLA-A*0201 molecules in vitro (HLA-A*0201
is considered a prototype A2 supertype molecule).
[0773] These peptides are then tested for the capacity to bind to
additional A2-supertype molecules (A*0202, A*0203, A*0206, and
A*6802). Peptides that bind to at least three of the five
A2-supertype alleles tested are typically deemed A2- supertype
cross-reactive binders. Preferred peptides bind at an affinity
equal to or less than 500 nM to three or more HLA-A2 supertype
molecules.
[0774] Selection of HLA-A3 Supermotif-beaning Epitopes
[0775] The 191P4D12(b) protein sequence(s) scanned above is also
examined for the presence of peptides with the HLA-A3-supermotif
primary anchors. Peptides corresponding to the HLA A3
supermotif-bearing sequences are then synthesized and tested for
binding to HLA-A*0301 and HLA-A*L1101 molecules, the molecules
encoded by the two most prevalent A3-supertype alleles. The
peptides that bind at least one of the two alleles with binding
affinities of .ltoreq.500 nM, often .ltoreq.200 nM, are then tested
for binding cross-reactivity to the other common A3-supertype
alleles (e.g., A*3101, A*3301, and A*6801) to identify those that
can bind at least three of the five HLA-A3-supertype molecules
tested.
[0776] Selection of HLA-B7 Supermotif Bearing Epitopes
[0777] The 191P4D12(b) protein(s) scanned above is also analyzed
for the presence of 8-, 9-10-, or 11-mer peptides with the
HLA-B7-supermotif. Corresponding peptides are synthesized and
tested for binding to HLA-B*0702, the molecule encoded by the most
common B7-supertype allele (i.e., the prototype B7 supertype
allele). Peptides binding B*0702 with IC.sub.50 of .ltoreq.500 nM
are identified using standard methods. These peptides are then
tested for binding to other common B7-supertype molecules (e.g.,
B*3501, B*5101, B*5301, and B*5401). Peptides capable of binding to
three or more of the five B7-supertype alleles tested are thereby
identified.
[0778] Selection of A1 and A24 Motif-bearing Epitopes
[0779] To further increase population coverage, HLA-A1 and -A24
epitopes can also be incorporated into vaccine compositions. An
analysis of the 191P4D12(b) protein can also be performed to
identify HLA-A1- and A24-motif-containing sequences.
[0780] High affinity and/or cross-reactive binding epitopes that
bear other motif and/or supermotifs are identified using analogous
methodology.
Example 14
Confirmation of Immunogenicity
[0781] Cross-reactive candidate CTL A2-supermotif-bearing peptides
that are identified as described herein are selected to confirm in
vitro immunogenicity. Confirmation is performed using the following
methodology:
[0782] Target Cell Lines for Cellular Screening:
[0783] The 0.221A2.1 cell line, produced by transferring the
HLA-A2.1 gene into the HLA-A, -B, -C null mutant human
B-lymphoblastoid cell line 721.221, is used as the peptide-loaded
target to measure activity of HLA-A2.1-restricted CTL. This cell
line is grown in RPMI-1640 medium supplemented with antibiotics,
sodium pyruvate, nonessential amino acids and 10% (v/v) heat
inactivated FCS. Cells that express an antigen of interest, or
transfectants comprising the gene encoding the antigen of interest,
can be used as target cells to confirm the ability of
peptide-specific CTLs to recognize endogenous antigen.
[0784] Primary CTL Induction Cultures:
[0785] Generation of Dendritic Cells (DC): PBMCs are thawed in RPMI
with 30 .mu.g/ml DNAse, washed twice and resuspended in complete
medium (RPMI-1640 plus 5% AB human serum, non-essential amino
acids, sodium pyruvate, L-glutamine and penicillin/streptomycin).
The monocytes are purified by plating 10.times.10.sup.6 PBMC/well
in a 6-well plate. After 2 hours at 37.degree. C., the non-adherent
cells are removed by gently shaking the plates and aspirating the
supernatants. The wells are washed a total of three times with 3 ml
RPMI to remove most of the non-adherent and loosely adherent cells.
Three ml of complete medium containing 50 ng/ml of GM-CSF and 1,000
U/ml of IL-4 are then added to each well. TNF.alpha. is added to
the DCs on day 6 at 75 ng/ml and the cells are used for CTL
induction cultures on day 7.
[0786] Induction of CTL with DC and Peptide: CD8+ T-cells are
isolated by positive selection with Dynal immunomagnetic beads
(Dynabeads.RTM. M-450) and the detacha-bead.RTM. reagent. Typically
about 200-250.times.10.sup.6 PBMC are processed to obtain
24.times.10.sup.6 CD8.sup.+ T-cells (enough for a 48-well plate
culture). Briefly, the PBMCs are thawed in RPMI with 30 .mu.g/ml
DNAse, washed once with PBS containing 1% human AB serum and
resuspended in PBS/1% AB serum at a concentration of
20.times.10.sup.6 cells/ml. The magnetic beads are washed 3 times
with PBS/AB serum, added to the cells (140 .mu.l
beads/20.times.10.sup.6 cells) and incubated for 1 hour at
4.degree. C. with continuous mixing. The beads and cells are washed
4.times. with PBS/AB serum to remove the nonadherent cells and
resuspended at 100.times.10.sup.6 cells/ml (based on the original
cell number) in PBS/AB serum containing 100 .mu.l/ml
detacha-bead.RTM. reagent and 30 .mu.g/ml DNAse. The mixture is
incubated for 1 hour at room temperature with continuous mixing.
The beads are washed again with PBS/AB/DNAse to collect the CD8+
T-cells. The DC are collected and centrifuged at 1300 rpm for 5-7
minutes, washed once with PBS with 1% BSA, counted and pulsed with
40 .mu.g/ml of peptide at a cell concentration of
1-2.times.10.sup.6/ml in the presence of 3 .mu.g/ml
.beta..sub.2-microglobulin for 4 hours at 20.degree. C. The DC are
then irradiated (4,200 rads), washed 1 time with medium and counted
again.
[0787] Setting up induction cultures: 0.25 ml cytokine-generated DC
(at 1.times.10.sup.5 cells/ml) are co-cultured with 0.25 ml of CD8+
T-cells (at 2.times.10.sup.6 cell/ml) in each well of a 48-well
plate in the presence of 10 ng/ml of IL-7. Recombinant human IL-10
is added the next day at afinal concentration of 10 ng/ml and
rhuman IL-2 is added 48 hours later at 10 U/ml.
[0788] Restimulation of the induction cultures with peptide-pulsed
adherent cells: Seven and fourteen days after the primary
induction, the cells are restimulated with peptide-pulsed adherent
cells. The PBMCs are thawed and washed twice with RPMI and DNAse.
The cells are resuspended at 5.times.10.sup.6 cells/ml and
irradiated at -4200 rads. The PBMCs are plated at 2.times.10.sup.6
in 0.5 ml complete medium per well and incubated for 2 hours at
37.degree. C. The plates are washed twice with RPMI by tapping the
plate gently to remove the nonadherent cells and the adherent cells
pulsed with 10 .mu.g/ml of peptide in the presence of 3 .mu.g/ml
.beta..sub.2 microglobulin in 0.25 ml RPMI/5%AB per well for 2
hours at 37.degree. C. Peptide solution from each well is aspirated
and the wells are washed once with RPMI. Most of the media is
aspirated from the induction cultures (CD8+ cells) and brought to
0.5 ml with fresh media. The cells are then transferred to the
wells containing the peptide-pulsed adherent cells. Twenty four
hours later recombinant human IL-10 is added at a final
concentration of 10 ng/ml and recombinant human IL2 is added the
next day and again 2-3 days later at 501 U/ml (Tsai et al.,
Critical Reviews in Immunology 18(1-2):65-75, 1998). Seven days
later, the cultures are assayed for CTL activity in a .sup.51Cr
release assay. In some experiments the cultures are assayed for
peptide-specific recognition in the in situ IFN.gamma. ELISA at the
time of the second restimulation followed by assay of endogenous
recognition 7 days later. After expansion, activity is measured in
both assays for a side-by-side comparison.
[0789] Measurement of CTL Lytic Activity by .sup.51Cr Release.
[0790] Seven days after the second restimulation, cytotoxicity is
determined in a standard (5 hr) .sup.51Cr release assay by assaying
individual wells at a single E:T. Peptide-pulsed targets are
prepared by incubating the cells with 10 .mu.g/ml peptide overnight
at 37.degree. C.
[0791] Adherent target cells are removed from culture flasks with
trypsin-EDTA. Target cells are labeled with 200 .mu.Ci of .sup.51Cr
sodium chromate (Dupont, Wilmington, Del.) for 1 hour at 37.degree.
C. Labeled target cells are resuspended at 10.sup.6 per ml and
diluted 1:10 with K562 cells at a concentration of
3.3.times.10.sup.6/ml (an NK-sensitive erythroblastoma cell line
used to reduce non-specific lysis). Target cells (100 .mu.l) and
effectors (100 .mu.l) are plated in 96 well round-bottom plates and
incubated for 5 hours at 37.degree. C. At that time, 100 .mu.l of
supernatant are collected from each well and percent lysis is
determined according to the formula:
[(cpm of the test sample-cpm of the spontaneous .sup.51Cr release
sample)/(cpm of the maximal .sup.51Cr release sample-cpm of the
spontaneous .sup.51Cr release sample)].times.100.
[0792] Maximum and spontaneous release are determined by incubating
the labeled targets with 1% Triton X-100 and media alone,
respectively. A positive culture is defined as one in which the
specific lysis (sample-background) is 10% or higher in the case of
individual wells and is 15% or more at the two highest E:T ratios
when expanded cultures are assayed.
[0793] In situ Measurement of Human IFN.gamma. Production as an
Indicator of Peptide-specific and Endogenous Recognition
[0794] Immulon 2 plates are coated with mouse anti-human IFN.gamma.
monoclonal antibody (4 .mu.g/ml 0.1M NaHCO.sub.3, pH8.2) overnight
at 4.degree. C. The plates are washed with Ca.sup.2+,
Mg.sup.2+-free PBS/0.05% Tween 20 and blocked with PBS/10% FCS for
two hours, after which the CTLs (100 .mu.l/well) and targets (100
.mu.l/well) are added to each well, leaving empty wells for the
standards and blanks (which received media only). The target cells,
either peptide-pulsed or endogenous targets, are used at a
concentration of 1.times.10.sup.6 cells/ml. The plates are
incubated for 48 hours at 37.degree. C. with 5% CO.sub.2.
[0795] Recombinant human IFN-gamma is added to the standard wells
starting at 400 pg or 1200 pg/100 microliter/well and the plate
incubated for two hours at 37.degree. C. The plates are washed and
100 .rho.l of biotinylated mouse anti-human IFN-gamma monoclonal
antibody (2 microgram/ml in PBS/3%FCS/0.05% Tween 20) are added and
incubated for 2 hours at room temperature. After washing again, 100
microliter HRP-streptavidin (1:4000) are added and the plates
incubated for one hour at room temperature. The plates are then
washed 6.times. with wash buffer, 100 microliter/well developing
solution (TMB 1:1) are added, and the plates allowed to develop for
5-15 minutes. The reaction is stopped with 50 microliter/well 1M
H.sub.3PO.sub.4 and read at OD450. A culture is considered positive
if it measured at least 50 pg of IFN-gamma/well above background
and is twice the background level of expression.
[0796] CTL Expansion.
[0797] Those cultures that demonstrate specific lytic activity
against peptide-pulsed targets and/or tumor targets are expanded
over a two week period with anti-CD3. Briefly, 5.times.10.sup.4
CD8+ cells are added to a T25 flask containing the following:
1.times.10.sup.6 irradiated (4,200 rad) PBMC (autologous or
allogeneic) per ml, 2.times.10.sup.5 irradiated (8,000 rad)
EBV-transformed cells per ml, and OKT3 (anti-CD3) at 30 ng per ml
in RPMI-1640 containing 10% (v/v) human AB serum, non-essential
amino acids, sodium pyruvate, 25 .mu.M 2-mercaptoethanol,
L-glutamine and penicillin/streptomycin. Recombinanthuman IL2 is
added 24 hours later at a final concentration of 2001 U/ml and
every three days thereafter with fresh media at 501 U/ml. The cells
are split if the cell concentration exceeds 1.times.10.sup.6/ml and
the cultures are assayed between days 13 and 15 at E:T ratios of
30, 10, 3 and 1:1 in the 51Cr release assay or at
1.times.10.sup.6/ml in the in situ IFN.gamma. assay using the same
targets as before the expansion.
[0798] Cultures are expanded in the absence of anti-CD3+ as
follows. Those cultures that demonstrate specific lytic activity
against peptide and endogenous targets are selected and
5.times.10.sup.4 CD8+ cells are added to a T25 flask containing the
following: 1.times.10.sup.6 autologous PBMC per ml which have been
peptide-pulsed with 10 .mu.g/ml peptide for two hours at 37.degree.
C. and irradiated (4,200 rad); 2.times.10.sup.5 irradiated (8,000
rad) EBV-transformed cells per ml RPMI-1640 containing 10%(v/v)
human AB serum, non-essential AA, sodium pyruvate, 25 mM 2-ME,
L-glutamine and gentamicin.
[0799] Immunogenicity of A2 Supermotif-bearing Peptides
[0800] A2-supermotif cross-reactive binding peptides are tested in
the cellular assay for the ability to induce peptide-specific CTL
in normal individuals. In this analysis, a peptide is typically
considered to be an epitope if it induces peptide-specific CTLs in
at least individuals, and preferably, also recognizes the
endogenously expressed peptide.
[0801] Immunogenicity can also be confirmed using PBMCs isolated
from patients bearing a tumor that expresses 191P4D12(b). Briefly,
PBMCs are isolated from patients, re-stimulated with peptide-pulsed
monocytes and assayed for the ability to recognize peptide-pulsed
target cells as well as transfected cells endogenously expressing
the antigen.
[0802] Evaluation of A*03/A11 Immunogenicity
[0803] HLA-A3 supermotif-bearing cross-reactive binding peptides
are also evaluated for immunogenicity using methodology analogous
for that used to evaluate the immunogenicity of the HLA-A2
supermotif peptides.
[0804] Evaluation of B7 Immunogenicity
[0805] Immunogenicity screening of the B7-supertype cross-reactive
binding peptides identified as set forth herein are confirmed in a
manner analogous to the confirmation of A2-and
A3-supermotif-bearing peptides.
[0806] Peptides bearing other supermotifs/motifs, e.g., HLA-A1,
HLA-A24 etc. are also confirmed using similar methodology
Example 15
Implementation of the Extended Supermotif to Improve the Binding
Capacity of Native Epitopes by Creating Analogs
[0807] HLA motifs and supermotifs (comprising primary and/or
secondary residues) are useful in the identification and
preparation of highly cross-reactive native peptides, as
demonstrated herein. Moreover, the definition of HLA motifs and
supermotifs also allows one to engineer highly cross-reactive
epitopes by identifying residues within a native peptide sequence
which can be analoged to confer upon the peptide certain
characteristics, e.g. greater cross-reactivity within the group of
HLA molecules that comprise a supertype, and/or greater binding
affinity for some or all of those HLA molecules. Examples of
analoging peptides to exhibit modulated binding affinity are set
forth in this example.
[0808] Analoging at Primary Anchor Residues
[0809] Peptide engineering strategies are implemented to further
increase the cross-reactivity of the epitopes. For example, the
main anchors of A2-supermotif-bearing peptides are altered, for
example, to introduce a preferred L, I, V, or M at position 2, and
I or V at the C-terminus.
[0810] To analyze the cross-reactivity of the analog peptides, each
engineered analog is initially tested for binding to the prototype
A2 supertype allele A*0201, then, if A*0201 binding capacity is
maintained, for A2-supertype cross-reactivity.
[0811] Alternatively, a peptide is confirmed as binding one or all
supertype members and then analoged to modulate binding affinity to
any one (or more) of the supertype members to add population
coverage.
[0812] The selection of analogs for immunogenicity in a cellular
screening analysis is typically further restricted by the capacity
of the parent wild type (WT) peptide to bind at least weakly, i.e.,
bind at an IC.sub.50 of 5000 nM or less, to three of more A2
supertype alleles. The rationale for this requirement is that the
WT peptides must be present endogenously in sufficient quantity to
be biologically relevant. Analoged peptides have been shown to have
increased immunogenicity and cross-reactivity by T cells specific
for the parent epitope (see, e.g., Parkhurst et al., J. Immunol.
157:2539, 1996; and Pogue et al., Proc. Natl. Acad. Sci. USA
92:8166, 1995).
[0813] In the cellular screening of these peptide analogs, it is
important to confirm that analog-specific CTLs are also able to
recognize the wild-type peptide and, when possible, target cells
that endogenously express the epitope.
[0814] Analoging of HLA-A3 and B7-Supermotif-bearing Peptides
[0815] Analogs of HLA-A3 supermotif-bearing epitopes are generated
using strategies similar to those employed in analoging HLA-A2
supermotif-bearing peptides. For example, peptides binding to 3/5
of the A3-supertype molecules are engineered at primary anchor
residues to possess a preferred residue (V, S, M, or A) at position
2.
[0816] The analog peptides are then tested for the ability to bind
A*03 and A*11 (prototype A3 supertype alleles). Those peptides that
demonstrate .ltoreq.500 nM binding capacity are then confirmed as
having A3-supertype cross-reactivity.
[0817] Similarly to the A2- and A3-motif bearing peptides, peptides
binding 3 or more B7-supertype alleles can be improved, where
possible, to achieve increased cross-reactive binding or greater
binding affinity or binding half life. B7 supermotif-bearing
peptides are, for example, engineered to possess a preferred
residue (V, I, L, or F) at the C-terminal primary anchor position,
as demonstrated by Sidney et al. (J. Immunol. 157:3480-3490,
1996).
[0818] Analoging at primary anchor residues of other motif and/or
supermotif-bearing epitopes is performed in a like manner.
[0819] The analog peptides are then be confirmed for
immunogenicity, typically in a cellular screening assay. Again, it
is generally important to demonstrate that analog-specific CTLs are
also able to recognize the wild-type peptide and, when possible,
targets that endogenously express the epitope.
[0820] Analoging at Secondary Anchor Residues
[0821] Moreover, HLA supermotifs are of value in engineering highly
cross-reactive peptides and/or peptides that bind HLA molecules
with increased affinity by identifying particular residues at
secondary anchor positions that are associated with such
properties. For example, the binding capacity of a B7
supermotif-bearing peptide with an F residue at position 1 is
analyzed. The peptide is then analoged to, for example, substitute
L for F at position 1. The analoged peptide is evaluated for
increased binding affinity, binding half life and/or increased
cross-reactivity. Such a procedure identifies analoged peptides
with enhanced properties.
[0822] Engineered analogs with sufficiently improved binding
capacity or cross-reactivity can also be tested for immunogenicity
in HLA-B7-transgenic mice, following for example, IFA immunization
or lipopeptide immunization. Analoged peptides are additionally
tested for the ability to stimulate a recall response using PBMC
from patients with 191P4D12(b)-expressing tumors.
[0823] Other Analoging Strategies
[0824] Another form of peptide analoging, unrelated to anchor
positions, involves the substitution of a cysteine with
.alpha.-amino butyric acid. Due to its chemical nature, cysteine
has the propensity to form disulfide bridges and sufficiently alter
the peptide structurally so as to reduce binding capacity.
Substitution of .alpha.-amino butyric acid for cysteine not only
alleviates this problem, but has been shown to improve binding and
crossbinding capabilities in some instances (see, e.g., the review
by Sette et al., In: Persistent Viral Infections, Eds. R. Ahmed and
I. Chen, John Wiley & Sons, England, 1999).
[0825] Thus, by the use of single amino acid substitutions, the
binding properties and/or cross-reactivity of peptide ligands for
HLA supertype molecules can be modulated.
Example 16
Identification and Confirmation of 191P4D12(b)-derived Sequences
with HLA-DR Binding Motifs
[0826] Peptide epitopes bearing an HLA class II supermotif or motif
are identified and confirmed as outlined below using methodology
similar to that described for HLA Class I peptides.
[0827] Selection of HLA-DR-supermotif-bearing Epitopes.
[0828] To identify 191P4D12(b)-derived, HLA class II HTL epitopes,
a 191P4D12(b) antigen is analyzed for the presence of sequences
bearing an HLA-DR-motif or supermotif. Specifically, 15-mer
sequences are selected comprising a DR-supermotif, comprising a
9-mer core, and three-residue N- and C-terminal flanking regions
(15 amino acids total).
[0829] Protocols for predicting peptide binding to DR molecules
have been developed (Southwood et al., J. Immunol. 160:3363-3373,
1998). These protocols, specific for individual DR molecules, allow
the scoring, and ranking, of 9-mer core regions. Each protocol not
only scores peptide sequences for the presence of DR-supermotif
primary anchors (i.e., at position 1 and position 6) within a 9-mer
core, but additionally evaluates sequences for the presence of
secondary anchors. Using allele-specific selection tables (see,
e.g., Southwood et al., ibid.), it has been found that these
protocols efficiently select peptide sequences with a high
probability of binding a particular DR molecule. Additionally, it
has been found that performing these protocols in tandem,
specifically those for DR1, DR4w4, and DR7, can efficiently select
DR cross-reactive peptides.
[0830] The 191P4D12(b)-derived peptides identified above are tested
for their binding capacity for various common HLA-DR molecules. All
peptides are initially tested for binding to the DR molecules in
the primary panel: DR1, DR4w4, and DR7. Peptides binding at least
two of these three DR molecules are then tested for binding to
DR2w2 .beta.1, DR2w2 .beta.2, DR6w19, and DR9 molecules in
secondary assays. Finally, peptides binding at least two of the
four secondary panel DR molecules, and thus cumulatively at least
four of seven different DR molecules, are screened for binding to
DR4w15, DR5w11, and DR8w2 molecules in tertiary assays. Peptides
binding at least seven of the ten DR molecules comprising the
primary, secondary, and tertiary screening assays are considered
cross-reactive DR binders. 191P4D12(b)-derived peptides found to
bind common HLA-DR alleles are of particular interest.
[0831] Selection of DR3 Motif Peptides
[0832] Because HLA-DR3 is an allele that is prevalent in Caucasian,
Black, and Hispanic populations, DR3 binding capacity is a relevant
criterion in the selection of HTL epitopes. Thus, peptides shown to
be candidates may also be assayed for their DR3 binding capacity.
However, in view of the binding specificity of the DR3 motif,
peptides binding only to DR3 can also be considered as candidates
for inclusion in a vaccine formulation.
[0833] To efficiently identify peptides that bind DR3, target
191P4D12(b) antigens are analyzed for sequences carrying one of the
two DR3-specific binding motifs reported by Geluk et al. (J.
Immunol. 152:5742-5748, 1994). The corresponding peptides are then
synthesized and confirmed as having the ability to bind DR3 with an
affinity of 1 .mu.M or better, i.e., less than 1 .mu.M. Peptides
are found that meet this binding criterion and qualify as HLA class
II high affinity binders.
[0834] DR3 binding epitopes identified in this manner are included
in vaccine compositions with DR supermotif-bearing peptide
epitopes.
[0835] Similarly to the case of HLA class I motif-bearing peptides,
the class II motif-bearing peptides are analoged to improve
affinity or cross-reactivity. For example, aspartic acid at
position 4 of the 9-mer core sequence is an optimal residue for DR3
binding, and substitution for that residue often improves DR 3
binding.
Example 17
Immunogenicity of 191P4D12(b)-derived HTL Epitopes
[0836] This example determines immunogenic DR supermotif- and DR3
motif-bearing epitopes among those identified using the methodology
set forth herein.
[0837] Immunogenicity of HTL epitopes are confirmed in a manner
analogous to the determination of immunogenicity of CTL epitopes,
by assessing the ability to stimulate HTL responses and/or by using
appropriate transgenic mouse models. Immunogenicity is determined
by screening for: 1.) in vitro primary induction using normal PBMC
or 2.) recall responses from patients who have
191P4D12(b)-expressing tumors.
Example 18
Calculation of Phenotypic Frequencies of HLA-supertypes in Various
Ethnic Backgrounds to Determine Breadth of Population Coverage
[0838] This example illustrates the assessment of the breadth of
population coverage of a vaccine composition comprised of multiple
epitopes comprising multiple supermotifs and/or motifs.
[0839] In order to analyze population coverage, gene frequencies of
HLA alleles are determined. Gene frequencies for each HLA allele
are calculated from antigen or allele frequencies utilizing the
binomial distribution formulae gf=1-(SQRT(1-af)) (see, e.g., Sidney
et al., Human Immunol. 45:79-93, 1996). To obtain overall
phenotypic frequencies, cumulative gene frequencies are calculated,
and the cumulative antigen frequencies derived by the use of the
inverse formula [af=1-(1-Cgf).sup.2].
[0840] Where frequency data is not available at the level of DNA
typing, correspondence to the serologically defined antigen
frequencies is assumed. To obtain total potential supertype
population coverage no linkage disequilibrium is assumed, and only
alleles confirmed to belong to each of the supertypes are included
(minimal estimates). Estimates of total potential coverage achieved
by inter-loci combinations are made by adding to the A coverage the
proportion of the non-A covered population that could be expected
to be covered by the B alleles considered (e.g., total=A+B*(1-A)).
Confirmed members of the A3-like supertype are A3, A11, A31,
A*3301, and A*6801. Although the A3-like supertype may also include
A34, A66, and A*7401, these alleles were not included in overall
frequency calculations. Likewise, confirmed members of the A2-like
supertype family are A*0201, A*0202, A*0203, A*0204, A*0205,
A*0206, A*0207, A*6802, and A*6901. Finally, the B7-like
supertype-confirmed alleles are: B7, B*3501-03, B51, B*5301,
B*5401, B*5501-2, B*5601, B*6701, and B*7801 (potentially also
B*1401, B*3504-06, B*4201, and B*5602).
[0841] Population coverage achieved by combining the A2-, A3- and
B7-supertypes is approximately 86% in five major ethnic groups.
Coverage may be extended by including peptides bearing the A1 and
A24 motifs. On average, A1 is present in 12% and A24 in 29% of the
population across five different major ethnic groups (Caucasian,
North American Black, Chinese, Japanese, and Hispanic). Together,
these alleles are represented with an average frequency of 39% in
these same ethnic populations. The total coverage across the major
ethnicities when A1 and A24 are combined with the coverage of the
A2-, A3- and B7-supertype alleles is >95%, see, e.g., Table IV
(G). An analogous approach can be used to estimate population
coverage achieved with combinations of class II motif-bearing
epitopes.
[0842] Immunogenicity studies in humans (e.g., Bertoni et al., J.
Clin. Invest. 100:503, 1997; Doolan et al., Immunity 7:97, 1997;
and Threlkeld et al., J. Immunol. 159:1648, 1997) have shown that
highly cross-reactive binding peptides are almost always recognized
as epitopes. The use of highly cross-reactive binding peptides is
an important selection criterion in identifying candidate epitopes
for inclusion in a vaccine that is immunogenic in a diverse
population.
[0843] With a sufficient number of epitopes (as disclosed herein
and from the art), an average population coverage is predicted to
be greater than 95% in each of five major ethnic populations. The
game theory Monte Carlo simulation analysis, which is known in the
art (see e.g., Osborne, M. J. and Rubinstein, A. "A course in game
theory" MIT Press, 1994), can be used to estimate what percentage
of the individuals in a population comprised of the Caucasian,
North American Black, Japanese, Chinese, and Hispanic ethnic groups
would recognize the vaccine epitopes described herein. A preferred
percentage is 90%. A more preferred percentage is 95%.
Example 19
CTL Recognition Of Endogenously Processed Antigens After
Priming
[0844] This example confirms that CTL induced by native or analoged
peptide epitopes identified and selected as described herein
recognize endogenously synthesized, i.e., native antigens.
[0845] Effector cells isolated from transgenic mice that are
immunized with peptide epitopes, for example HLA-A2
supermotif-bearing epitopes, are re-stimulated in vitro using
peptide-coated stimulator cells. Six days later, effector cells are
assayed for cytotoxicity and the cell lines that contain
peptide-specific cytotoxic activity are further re-stimulated. An
additional six days later, these cell lines are tested for
cytotoxic activity on .sup.51Cr labeled Jurkat-A2.1/K.sup.b target
cells in the absence or presence of peptide, and also tested on
.sup.51Cr labeled target cells bearing the endogenously synthesized
antigen, i.e. cells that are stably transfected with 191P4D12(b)
expression vectors.
[0846] The results demonstrate that CTL lines obtained from animals
primed with peptide epitope recognize endogenously synthesized
191P4D12(b) antigen. The choice of transgenic mouse model to be
used for such an analysis depends upon the epitope(s) that are
being evaluated. In addition to HLA-A*0201/K.sup.b transgenic mice,
several other transgenic mouse models including mice with human
A11, which may also be used to evaluate A3 epitopes, and B7 alleles
have been characterized and others (e.g., transgenic mice for
HLA-A1 and A24) are being developed. HLA-DR1 and HLA-DR3 mouse
models have also been developed, which may be used to evaluate HTL
epitopes.
Example 20
Activity Of CTL-HTL Coniugated Epitopes In Transgenic Mice
[0847] This example illustrates the induction of CTLs and HTLs in
transgenic mice, by use of a 191P4D12(b)-derived CTL and HTL
peptide vaccine compositions. The vaccine composition used herein
comprise peptides to be administered to a patient with a
191P4D12(b)-expressing tumor. The peptide composition can comprise
multiple CTL and/or HTL epitopes. The epitopes are identified using
methodology as described herein. This example also illustrates that
enhanced immunogenicity can be achieved by inclusion of one or more
HTL epitopes in a CTL vaccine composition; such a peptide
composition can comprise an HTL epitope conjugated to a CTL
epitope. The CTL epitope can be one that binds to multiple HLA
family members at an affinity of 500 nM or less, or analogs of that
epitope. The peptides may be lipidated, if desired.
[0848] Immunization procedures: Immunization of transgenic mice is
performed as described (Alexander et al., J. Immunol.
159:4753-4761, 1997). For example, A2/K.sup.b mice, which are
transgenic for the human HLA A2.1 allele and are used to confirm
the immunogenicity of HLA-A*0201 motif- or HLA-A2
supermotif-bearing epitopes, and are primed subcutaneously (base of
the tail) with a 0.1 ml of peptide in Incomplete Freund's Adjuvant,
or if the peptide composition is a lipidated CTL/HTL conjugate, in
DMSO/saline, or if the peptide composition is a polypeptide, in PBS
or incomplete Freund's Adjuvant. Seven days after priming,
splenocytes obtained from these animals are restimulated with
syngenic irradiated LPS-activated lymphoblasts coated with
peptide.
[0849] Cell lines: Target cells for peptide-specific cytotoxicity
assays are Jurkat cells transfected with the HLA-A2.1/K.sup.b
chimeric gene (e.g., Vitiello et al., J. Exp. Med. 173:1007,
1991)
[0850] In vitro CTL activation: One week after priming, spleen
cells (30.times.10.sup.6 cells/flask) are co-cultured at 37.degree.
C. with syngeneic, irradiated (3000 rads), peptide coated
lymphoblasts (10.times.10.sup.6 cells/flask) in 10 ml of culture
medium/T25 flask. After six days, effector cells are harvested and
assayed for cytotoxic activity.
[0851] Assay for cytotoxic activty: Target cells (1.0 to
1.5.times.10.sup.6) are incubated at 37.degree.60 C. in the
presence of 200 .mu.l of .sup.51Cr. After 60 minutes, cells are
washed three times and resuspended in R10 medium. Peptide is added
where required at a concentration of 1 .mu.g/ml. For the assay,
10.sup.4 51Cr-labeled target cells are added to different
concentrations of effector cells (final volume of 200 .mu.l) in
U-bottom 96-well plates. After a six hour incubation period at
37.degree. C., a 0.1 ml aliquot of supernatant is removed from each
well and radioactivity is determined in a Micromedic automatic
gamma counter. The percent specific lysis is determined by the
formula: percent specific release=100.times.(experimental
release-spontaneous release)/(maximum release-spontaneous release).
To facilitate comparison between separate CTL assays run under the
same conditions, % .sup.51Cr release data is expressed as lytic
units/10.sup.6 cells. One lytic unit is arbitrarily defined as the
number of effector cells required to achieve 30% lysis of 10,000
target cells in a six hour .sup.51Cr release assay. To obtain
specific lytic units/10.sup.6, the lytic units/10.sup.6 obtained in
the absence of peptide is subtracted from the lytic units/10.sup.6
obtained in the presence of peptide. For example, if 30% .sup.51Cr
release is obtained at the effector (E): target (T) ratio of 50:1
(i.e., 5.times.10.sup.5 effector cells for 10,000 targets) in the
absence of peptide and 5:1 (i.e., 5.times.10.sup.4 effector cells
for 10,000 targets) in the presence of peptide, the specific lytic
units would be: [(1/50,000)-(1/500,000)].times.10.sup.6=18 LU.
[0852] The results are analyzed to assess the magnitude of the CTL
responses of animals injected with the immunogenic CTL/HTL
conjugate vaccine preparation and are compared to the magnitude of
the CTL response achieved using, for example, CTL epitopes as
outlined above in the Example entitled "Confirmation of
Immunogenicity." Analyses similar to this may be performed to
confirm the immunogenicity of peptide conjugates containing
multiple CTL epitopes and/or multiple HTL epitopes. In accordance
with these procedures, it is found that a CTL response is induced,
and concomitantly that an HTL response is induced upon
administration of such compositions.
Example 21
Selection of CTL and HTL Epitopes for Inclusion in a
191P4D12(b)-Specific Vaccine
[0853] This example illustrates a procedure for selecting peptide
epitopes for vaccine compositions of the invention. The peptides in
the composition can be in the form of a nucleic acid sequence,
either single or one or more sequences (i.e., minigene) that
encodes peptide(s), or can be single and/or polyepitopic
peptides.
[0854] The following principles are utilized when selecting a
plurality of epitopes for inclusion in a vaccine composition. Each
of the following principles is balanced in order to make the
selection.
[0855] Epitopes are selected which, upon administration, mimic
immune responses that are correlated with 191P4D12(b) clearance.
The number of epitopes used depends on observations of patients who
spontaneously clear 191P4D12(b). For example, if it has been
observed that patients who spontaneously clear
191P4D12(b)-expressing cells generate an immune response to at
least three (3) epitopes from 191P4D12(b) antigen, then at least
three epitopes should be included for HLA class I. A similar
rationale is used to determine HLA class II epitopes.
[0856] Epitopes are often selected that have a binding affinity of
an IC.sub.50 of 500 nM or less for an HLA class I molecule, or for
class II, an IC.sub.50 of 1000 nM or less, or HLA Class I peptides
with high binding scores from the BIMAS web site, at URL
bimas.dcrt.nih.gov/.
[0857] In order to achieve broad coverage of the vaccine through
out a diverse population, sufficient supermotif bearing peptides,
or a sufficient array of allele-specific motif bearing peptides,
are selected to give broad population coverage. In one embodiment,
epitopes are selected to provide at least 80% population coverage.
A Monte Carlo analysis, a statistical evaluation known in the art,
can be employed to assess breadth, or redundancy, of population
coverage.
[0858] When creating polyepitopic compositions, or a minigene that
encodes same, it is typically desirable to generate the smallest
peptide possible that encompasses the epitopes of interest. The
principles employed are similar, if not the same, as those employed
when selecting a peptide comprising nested epitopes. For example, a
protein sequence for the vaccine composition is selected because it
has maximal number of epitopes contained within the sequence, i.e.,
it has a high concentration of epitopes. Epitopes may be nested or
overlapping (i.e., frame shifted relative to one another). For
example, with overlapping epitopes, two 9-mer epitopes and one
10-mer epitope can be present in a 10 amino acid peptide. Each
epitope can be exposed and bound by an HLA molecule upon
administration of such a peptide. A multi-epitopic, peptide can be
generated synthetically, recombinantly, or via cleavage from the
native source. Alternatively, an analog can be made of this native
sequence, whereby one or more of the epitopes comprise
substitutions that alter the cross-reactivity and/or binding
affinity properties of the polyepitopic peptide. Such a vaccine
composition is administered for therapeutic or prophylactic
purposes. This embodiment provides for the possibility that an as
yet undiscovered aspect of immune system processing will apply to
the native nested sequence and thereby facilitate the production of
therapeutic or prophylactic immune response-inducing vaccine
compositions. Additionally such an embodiment provides for the
possibility of motif-bearing epitopes for an HLA makeup that is
presently unknown. Furthermore, this embodiment (absent the
creating of any analogs) directs the immune response to multiple
peptide sequences that are actually present in 191P4D12(b), thus
avoiding the need to evaluate any junctional epitopes. Lastly, the
embodiment provides an economy of scale when producing nucleic acid
vaccine compositions. Related to this embodiment, computer programs
can be derived in accordance with principles in the art, which
identify in a target sequence, the greatest number of epitopes per
sequence length.
[0859] A vaccine composition comprised of selected peptides, when
administered, is safe, efficacious, and elicits an immune response
similar in magnitude to an immune response that controls or clears
cells that bear or overexpress 191P4D12(b).
Example 22
Construction of "Minigene" Multi-Epitope DNA Plasmids
[0860] This example discusses the construction of a minigene
expression plasmid. Minigene plasmids may, of course, contain
various configurations of B cell, CTL and/or HTL epitopes or
epitope analogs as described herein.
[0861] A minigene expression plasmid typically includes multiple
CTL and HTL peptide epitopes. In the present example, HLA-A2, -A3,
-B7 supermotif-bearing peptide epitopes and HLA-A1 and -A24
motif-bearing peptide epitopes are used in conjunction with DR
supermotif-bearing epitopes and/or DR3 epitopes. HLA class I
supermotif or motif-bearing peptide epitopes derived 191P4D12(b),
are selected such that multiple supermotifs/motifs are represented
to ensure broad population coverage. Similarly, HLA class II
epitopes are selected from 191P4D12(b) to provide broad population
coverage, i.e. both HLA DR-1-4-7 supermotif-bearing epitopes and
HLA DR-3 motif-bearing epitopes are selected for inclusion in the
minigene construct. The selected CTL and HTL epitopes are then
incorporated into a minigene for expression in an expression
vector.
[0862] Such a construct may additionally include sequences that
direct the HTL epitopes to the endoplasmic reticulum. For example,
the Ii protein may be fused to one or more HTL epitopes as
described in the art, wherein the CLIP sequence of the Ii protein
is removed and replaced with an HLA class II epitope sequence so
that HLA class II epitope is directed to the endoplasmic reticulum,
where the epitope binds to an HLA class II molecules.
[0863] This example illustrates the methods to be used for
construction of a minigene-bearing expression plasmid. Other
expression vectors that may be used for minigene compositions are
available and known to those of skill in the art.
[0864] The minigene DNA plasmid of this example contains a
consensus Kozak sequence and a consensus murine kappa Ig-light
chain signal sequence followed by CTL and/or HTL epitopes selected
in accordance with principles disclosed herein. The sequence
encodes an open reading frame fused to the Myc and His antibody
epitope tag coded for by the pcDNA 3.1 Myc-His vector.
[0865] Overlapping oligonucleotides that can, for example, average
about 70 nucleotides in length with 15 nucleotide overlaps, are
synthesized and HPLC-purified. The oligonucleotides encode the
selected peptide epitopes as well as appropriate linker
nucleotides, Kozak sequence, and signal sequence. The final
multiepitope minigene is assembled by extending the overlapping
oligonucleotides in three sets of reactions using PCR. A
Perkin/Elmer 9600 PCR machine is used and a total of 30 cycles are
performed using the following conditions: 95.degree. C. for 15 sec,
annealing temperature (5.degree. below the lowest calculated Tm of
each primer pair) for 30 sec, and 72.degree. C. for 1 min.
[0866] For example, a minigene is prepared as follows. For a first
PCR reaction, 5 .mu.g of each of two oligonucleotides are annealed
and extended: In an example using eight oligonucleotides, i.e.,
four pairs of primers, oligonucleotides 1+2, 3+4, 5+6, and 7+8 are
combined in 100 .mu.l reactions containing Pfu polymerase buffer
(1x=10 mM KCL, 10 mM (NH4).sub.2SO.sub.4, 20 mM Tris-chloride, pH
8.75, 2 mM MgSO.sub.4, 0.1% Triton X-100, 100 .mu.g/ml BSA), 0.25
mM each dNTP, and 2.5 U of Pfu polymerase. The full-length dimer
products are gel-purified, and two reactions containing the product
of 1+2 and 3+4, and the product of 5+6 and 7+8 are mixed, annealed,
and extended for 10 cycles. Half of the two reactions are then
mixed, and 5 cycles of annealing and extension carried out before
flanking primers are added to amplify the full length product. The
full-length product is gel-purified and cloned into pCR-blunt
(Invitrogen) and individual clones are screened by sequencing.
Example 23
The Plasmid Construct and the Degree to Which It Induces
Immunogenicity
[0867] The degree to which a plasmid construct, for example a
plasmid constructed in accordance with the previous Example, is
able to induce immunogenicity is confirmed in vitro by determining
epitope presentation by APC following transduction or transfection
of the APC with an epitope-expressing nucleic acid construct. Such
a study determines "antigenicity" and allows the use of human APC.
The assay determines the ability of the epitope to be presented by
the APC in a context that is recognized by a T cell by quantifying
the density of epitope-HLA class I complexes on the cell surface.
Quantitation can be performed by directly measuring the amount of
peptide eluted from the APC (see, e.g., Sijts et al., J. Immunol.
156:683-692, 1996; Demotz et al., Nature 342:682-684, 1989); or the
number of peptide-HLA class I complexes can be estimated by
measuring the amount of lysis or lymphokine release induced by
diseased or transfected target cells, and then determining the
concentration of peptide necessary to obtain equivalent levels of
lysis or lymphokine release (see, e.g., Kageyama et al., J.
Immunol. 154:567-576, 1995).
[0868] Alternatively, immunogenicity is confirmed through in vivo
injections into mice and subsequent in vitro assessment of CTL and
HTL activity, which are analyzed using cytotoxicity and
proliferation assays, respectively, as detailed e.g., in Alexander
et al., Immunity 1:751-761, 1994.
[0869] For example, to confirm the capacity of a DNA minigene
construct containing at least one HLA-A2 supermotif peptide to
induce CTLs in vivo, HLA-A2.1/K.sup.b transgenic mice, for example,
are immunized intramuscularly with 100 .mu.g of naked cDNA. As a
means of comparing the level of CTLs induced by cDNA immunization,
a control group of animals is also immunized with an actual peptide
composition that comprises multiple epitopes synthesized as a
single polypeptide as they would be encoded by the minigene.
[0870] Splenocytes from immunized animals are stimulated twice with
each of the respective compositions (peptide epitopes encoded in
the minigene or the polyepitopic peptide), then assayed for
peptide-specific cytotoxic activity in a .sup.51Cr release assay.
The results indicate the magnitude of the CTL response directed
against the A2-restricted epitope, thus indicating the in vivo
immunogenicity of the minigene vaccine and polyepitopic
vaccine.
[0871] It is, therefore, found that the minigene elicits immune
responses directed toward the HLA-A2 supermotif peptide epitopes as
does the polyepitopic peptide vaccine. A similar analysis is also
performed using other HLA-A3 and HLA-B7 transgenic mouse models to
assess CTL induction by HLA-A3 and HLA-B7 motif or supermotif
epitopes, whereby it is also found that the minigene elicits
appropriate immune responses directed toward the provided
epitopes.
[0872] To confirm the capacity of a class II epitope-encoding
minigene to induce HTLs in vivo, DR transgenic mice, or for those
epitopes that cross react with the appropriate mouse MHC molecule,
I-A.sup.b-restricted mice, for example, are immunized
intramuscularly with 100 .mu.g of plasmid DNA. As a means of
comparing the level of HTLs induced by DNA immunization, a group of
control animals is also immunized with an actual peptide
composition emulsified in complete Freund's adjuvant. CD4+ T cells,
i.e. HTLs, are purified from splenocytes of immunized animals and
stimulated with each of the respective compositions (peptides
encoded in the minigene). The HTL response is measured using a
.sup.3H-thymidine incorporation proliferation assay, (see, e.g.,
Alexander et al. Immunity 1:751-761, 1994). The results indicate
the magnitude of the HTL response, thus demonstrating the in vivo
immunogenicity of the minigene.
[0873] DNA minigenes, constructed as described in the previous
Example, can also be confirmed as a vaccine in combination with a
boosting agent using a prime boost protocol. The boosting agent can
consist of recombinant protein (e.g., Barnett et al., Aids Res. and
Human Retroviruses 14, Supplement 3:S299-S309, 1998) or recombinant
vaccinia, for example, expressing a minigene or DNA encoding the
complete protein of interest (see, e.g., Hanke et al., Vaccine
16:439-445,1998; Sedegah et al., Proc. Natl. Acad. Sci USA
95:7648-53, 1998; Hanke and McMichael, Immunol. Letters 66:177-181,
1999; and Robinson et al., Nature Med. 5:526-34, 1999).
[0874] For example, the efficacy of the DNA minigene used in a
prime boost protocol is initially evaluated in transgenic mice. In
this example, A2.1/K.sup.b transgenic mice are immunized IM with
100 .mu.g of a DNA minigene encoding the immunogenic peptides
including at least one HLA-A2 supermotif-bearing peptide. After an
incubation period (ranging from 3-9 weeks), the mice are boosted IP
with 10.sup.7 pfu/mouse of a recombinant vaccinia virus expressing
the same sequence encoded by the DNA minigene. Control mice are
immunized with 100 .mu.g of DNA or recombinant vaccinia without the
minigene sequence, or with DNA encoding the minigene, but without
the vaccinia boost. After an additional incubation period of two
weeks, splenocytes from the mice are immediately assayed for
peptide-specific activity in an ELISPOT assay. Additionally,
splenocytes are stimulated in vitro with the A2- restricted peptide
epitopes encoded in the minigene and recombinant vaccinia, then
assayed for peptide-specific activity in an alpha, beta and/or
gamma IFN ELISA.
[0875] It is found that the minigene utilized in a prime-boost
protocol elicits greater immune responses toward the HLA-A2
supermotif peptides than with DNA alone. Such an analysis can also
be performed using HLA-A11 or HLA-B7 transgenic mouse models to
assess CTL induction by HLA-A3 or HLA-B7 motif or supermotif
epitopes. The use of prime boost protocols in humans is described
below in the Example entitled "Induction of CTL Responses Using a
Prime Boost Protocol."
Example 24
Peptide Compositions for Prophylactic Uses
[0876] Vaccine compositions of the present invention can be used to
prevent 191P4D12(b) expression in persons who are at risk for
tumors that bear this antigen. For example, a polyepitopic peptide
epitope composition (or a nucleic acid comprising the same)
containing multiple CTL and HTL epitopes such as those selected in
the above Examples, which are also selected to target greater than
80% of the population, is administered to individuals at risk for a
191P4D12(b)-associated tumor.
[0877] For example, a peptide-based composition is provided as a
single polypeptide that encompasses multiple epitopes. The vaccine
is typically administered in a physiological solution that
comprises an adjuvant, such as Incomplete Freunds Adjuvant. The
dose of peptide for the initial immunization is from about 1 to
about 50,000 .mu.g, generally 100-5,000 .mu.g, for a 70 kg patient.
The initial administration of vaccine is followed by booster
dosages at 4 weeks followed by evaluation of the magnitude of the
immune response in the patient, by techniques that determine the
presence of epitope-specific CTL populations in a PBMC sample.
Additional booster doses are administered as required. The
composition is found to be both safe and efficacious as a
prophylaxis against 191P4D12(b)-associated disease.
[0878] Alternatively, a composition typically comprising
transfecting agents is used for the administration of a nucleic
acid-based vaccine in accordance with methodologies known in the
art and disclosed herein.
Example 25
Polyepitopic Vaccine Compositions Derived from Native 191P4D12(b)
Sequences
[0879] A native 191P4D12(b) polyprotein sequence is analyzed,
preferably using computer algorithms defined for each class I
and/or class II supermotif or motif, to identify "relatively short"
regions of the polyprotein that comprise multiple epitopes. The
"relatively short" regions are preferably less in length than an
entire native antigen. This relatively short sequence that contains
multiple distinct or overlapping, "nested" epitopes can be used to
generate a minigene construct. The construct is engineered to
express the peptide, which corresponds to the native protein
sequence. The "relatively short" peptide is generally less than 250
amino acids in length, often less than 100 amino acids in length,
preferably less than 75 amino acids in length, and more preferably
less than 50 amino acids in length. The protein sequence of the
vaccine composition is selected because it has maximal number of
epitopes contained within the sequence, i.e., it has a high
concentration of epitopes. As noted herein, epitope motifs may be
nested or overlapping (i.e., frame shifted relative to one
another). For example, with overlapping epitopes, two 9-mer
epitopes and one 10-mer epitope can be present in a 10 amino acid
peptide. Such a vaccine composition is administered for therapeutic
or prophylactic purposes.
[0880] The vaccine composition will include, for example, multiple
CTL epitopes from 191P4D12(b) antigen and at least one HTL epitope.
This polyepitopic native sequence is administered either as a
peptide or as a nucleic acid sequence which encodes the peptide.
Alternatively, an analog can be made of this native sequence,
whereby one or more of the epitopes comprise substitutions that
alter the cross-reactivity and/or binding affinity properties of
the polyepitopic peptide.
[0881] The embodiment of this example provides for the possibility
that an as yet undiscovered aspect of immune system processing will
apply to the native nested sequence and thereby facilitate the
production of therapeutic or prophylactic immune response-inducing
vaccine compositions. Additionally, such an embodiment provides for
the possibility of motif-bearing epitopes for an HLA makeup(s) that
is presently unknown. Furthermore, this embodiment (excluding an
analoged embodiment) directs the immune response to multiple
peptide sequences that are actually present in native 191P4D12(b),
thus avoiding the need to evaluate any junctional epitopes. Lastly,
the embodiment provides an economy of scale when producing peptide
or nucleic acid vaccine compositions.
[0882] Related to this embodiment, computer programs are available
in the art which can be used to identify in a target sequence, the
greatest number of epitopes per sequence length.
Example 26
Polyepitopic Vaccine Compositions from Multiple Antigens
[0883] The 191P4D12(b) peptide epitopes of the present invention
are used in conjunction with epitopes from other target
tumor-associated antigens, to create a vaccine composition that is
useful for the prevention or treatment of cancer that expresses
191P4D12(b) and such other antigens. For example, a vaccine
composition can be provided as a single polypeptide that
incorporates multiple epitopes from 191P4D12(b) as well as
tumor-associated antigens that are often expressed with a target
cancer associated with 191P4D12(b) expression, or can be
administered as a composition comprising a cocktail of one or more
discrete epitopes. Alternatively, the vaccine can be administered
as a minigene construct or as dendritic cells which have been
loaded with the peptide epitopes in vitro.
Example 27
Use of Peptides to Evaluate an Immune Response
[0884] Peptides of the invention may be used to analyze an immune
response for the presence of specific antibodies, CTL or HTL
directed to 191P4D12(b). Such an analysis can be performed in a
manner described by Ogg et al., Science 279:2103-2106, 1998. In
this Example, peptides in accordance with the invention are used as
a reagent for diagnostic or prognostic purposes, not as an
immunogen.
[0885] In this example highly sensitive human leukocyte antigen
tetrameric complexes ("tetramers") are used for a cross-sectional
analysis of, for example, 191P4D12(b) HLA-A*0201-specific CTL
frequencies from HLA A*0201 -positive individuals at different
stages of disease or following immunization comprising a
191P4D12(b) peptide containing an A*0201 motif. Tetrameric
complexes are synthesized as described (Musey et al., N. Engl. J.
Med. 337:1267, 1997). Briefly, purified HLA heavy chain (A*0201 in
this example) and .beta.2-microglobulin are synthesized by means of
a prokaryotic expression system. The heavy chain is modified by
deletion of the transmembrane-cytosolic tail and COOH-terminal
addition of a sequence containing a BirA enzymatic biotinylation
site. The heavy chain, .beta.2-microglobulin, and peptide are
refolded by dilution. The 45-kD refolded product is isolated by
fast protein liquid chromatography and then biotinylated by BirA in
the presence of biotin (Sigma, St. Louis, Mo.), adenosine 5'
triphosphate and magnesium. Streptavidin-phycoerythrin conjugate is
added in a 1:4 molar ratio, and the tetrameric product is
concentrated to 1 mg/ml. The resulting product is referred to as
tetramer-phycoerythrin.
[0886] For the analysis of patient blood samples, approximately one
million PBMCs are centrifuged at 300 g for 5 minutes and
resuspended in 50 .mu.l of cold phosphate-buffered saline.
Tri-color analysis is performed with the tetramer-phycoerythrin,
along with anti-CD8-Tricolor, and anti-CD38. The PBMCs are
incubated with tetramer and antibodies on ice for 30 to 60 min and
then washed twice before formaldehyde fixation. Gates are applied
to contain >99.98% of control samples. Controls for the
tetramers include both A*0201-negative individuals and
A*0201-positive non-diseased donors. The percentage of cells
stained with the tetramer is then determined by flow cytometry. The
results indicate the number of cells in the PBMC sample that
contain epitope-restricted CTLs, thereby readily indicating the
extent of immune response to the 191P4D12(b) epitope, and thus the
status of exposure to 191P4D12(b), or exposure to a vaccine that
elicits a protective or therapeutic response.
Example 28
Use of Peptide Epitopes to Evaluate Recall Responses
[0887] The peptide epitopes of the invention are used as reagents
to evaluate T cell responses, such as acute or recall responses, in
patients. Such an analysis may be performed on patients who have
recovered from 191P4D12(b)-associated disease or who have been
vaccinated with a 191P4D12(b) vaccine.
[0888] For example, the class I restricted CTL response of persons
who have been vaccinated may be analyzed. The vaccine may be any
191P4D12(b) vaccine. PBMC are collected from vaccinated individuals
and HLA typed. Appropriate peptide epitopes of the invention that,
optimally, bear supermotifs to provide cross-reactivity with
multiple HLA supertype family members, are then used for analysis
of samples derived from individuals who bear that HLA type.
[0889] PBMC from vaccinated individuals are separated on
Ficoll-Histopaque density gradients (Sigma Chemical Co., St. Louis,
Mo.), washed three times in HBSS (GIBCO Laboratories), resuspended
in RPMI-1640 (GIBCO Laboratories) supplemented with L-glutamine
(2mM), penicillin (50 U/ml), streptomycin (50 .mu.g/ml), and Hepes
(10 mM) containing 10% heat-inactivated human AB serum (complete
RPMI) and plated using microculture formats. A synthetic peptide
comprising an epitope of the invention is added at 10 .mu.g/ml to
each well and HBV core 128-140 epitope is added at 1 .mu.g/ml to
each well as a source of T cell help during the first week of
stimulation.
[0890] In the microculture format, 4.times.10.sup.5 PBMC are
stimulated with peptide in 8 replicate cultures in 96-well round
bottom plate in 100 .mu.l/well of complete RPMI. On days 3 and 10,
100 .mu.l of complete RPMI and 20 U/ml final concentration of rlL-2
are added to each well. On day 7 the cultures are transferred into
a 96-well flat-bottom plate and restimulated with peptide, rlL-2
and 10.sup.5 irradiated (3,000 rad) autologous feeder cells. The
cultures are tested for cytotoxic activity on day 14. A positive
CTL response requires two or more of the eight replicate cultures
to display greater than 10% specific .sup.51Cr release, based on
comparison with non-diseased control subjects as previously
described (Rehermann, et al., Nature Med. 2:1104,1108, 1996;
Rehermann et al., J. Clin. Invest. 97:1655-1665, 1996; and
Rehermann et al. J. Clin. Invest. 98:1432-1440, 1996).
[0891] Target cell lines are autologous and allogeneic
EBV-transformed B-LCL that are either purchased from the American
Society for Histocompatibility and Immunogenetics (ASHI, Boston,
Mass.) or established from the pool of patients as described
(Guilhot, et al. J. Virol. 66:2670-2678, 1992).
[0892] Cytotoxicity assays are performed in the following manner.
Target cells consist of either allogeneic HLA-matched or autologous
EBV-transformed B lymphoblastoid cell line that are incubated
overnight with the synthetic peptide epitope of the invention at 10
.mu.M, and labeled with 100 .mu.Ci of .sup.51Cr (Amersham Corp.,
Arlington Heights, Ill.) for 1 hour after which they are washed
four times with HBSS.
[0893] Cytolytic activity is determined in a standard 4-h, split
well .sup.51Cr release assay using U-bottomed 96 well plates
containing 3,000 targets/well. Stimulated PBMC are tested at
effector/target (E/T) ratios of 20-50:1 on day 14. Percent
cytotoxicity is determined from the formula:
100.times.[(experimental release-spontaneous release)/maximum
release-spontaneous release)]. Maximum release is determined by
lysis of targets by detergent (2% Triton X-100; Sigma Chemical Co.,
St. Louis, Mo.). Spontaneous release is <25% of maximum release
for all experiments.
[0894] The results of such an analysis indicate the extent to which
HLA-restricted CTL populations have been stimulated by previous
exposure to 191P4D12(b) or a 191P4D12(b) vaccine.
[0895] Similarly, Class II restricted HTL responses may also be
analyzed. Purified PBMC are cultured in a 96-well flat bottom plate
at a density of 1.5.times.10.sup.5 cells/well and are stimulated
with 10 .mu.g/ml synthetic peptide of the invention, whole
191P4D12(b) antigen, or PHA. Cells are routinely plated in
replicates of 4-6 wells for each condition. After seven days of
culture, the medium is removed and replaced with fresh medium
containing 10 U/ml IL-2. Two days later, 1 .mu.Ci .sup.3H-thymidine
is added to each well and incubation is continued for an additional
18 hours. Cellular DNA is then harvested on glass fiber mats and
analyzed for .sup.3H-thymidine incorporation. Antigen-specific T
cell proliferation is calculated as the ratio of .sup.3H-thymidine
incorporation in the presence of antigen divided by the
.sup.3H-thymidine incorporation in the absence of antigen.
Example 29
Induction of Specific CTL Response in Humans
[0896] A human clinical trial for an immunogenic composition
comprising CTL and HTL epitopes of the invention is set up as an
IND Phase I, dose escalation study and carried out as a randomized,
double-blind, placebo-controlled trial. Such a trial is designed,
for example, as follows:
[0897] A total of about 27 individuals are enrolled and divided
into 3 groups:
[0898] Group I: 3 subjects are injected with placebo and 6 subjects
are injected with 5 .mu.g of peptide composition;
[0899] Group II: 3 subjects are injected with placebo and 6
subjects are injected with 50 .mu.g peptide composition;
[0900] Group III: 3 subjects are injected with placebo and 6
subjects are injected with 500 .mu.g of peptide composition.
[0901] After 4 weeks following the first injection, all subjects
receive a booster inoculation at the same dosage.
[0902] The endpoints measured in this study relate to the safety
and tolerability of the peptide composition as well as its
immunogenicity. Cellular immune responses to the peptide
composition are an index of the intrinsic activity of this the
peptide composition, and can therefore be viewed as a measure of
biological efficacy. The following summarize the clinical and
laboratory data that relate to safety and efficacy endpoints.
[0903] Safety: The incidence of adverse events is monitored in the
placebo and drug treatment group and assessed in terms of degree
and reversibility.
[0904] Evaluation of Vaccine Efficacy: For evaluation of vaccine
efficacy, subjects are bled before and after injection. Peripheral
blood mononuclear cells are isolated from fresh heparinized blood
by Ficoll-Hypaque density gradient centrifugation, aliquoted in
freezing media and stored frozen. Samples are assayed for CTL and
HTL activity.
[0905] The vaccine is found to be both safe and efficacious.
Example 30
Phase II Trials in Patients Expressing 191P4D12(b)
[0906] Phase II trials are performed to study the effect of
administering the CTL-HTL peptide compositions to patients having
cancer that expresses 191P4D12(b). The main objectives of the trial
are to determine an effective dose and regimen for inducing CTLs in
cancer patients that express 191P4D12(b), to establish the safety
of inducing a CTL and HTL response in these patients, and to see to
what extent activation of CTLs improves the clinical picture of
these patients, as manifested, e.g., by the reduction and/or
shrinking of lesions. Such a study is designed, for example, as
follows:
[0907] The studies are performed in multiple centers. The trial
design is an open-label, uncontrolled, dose escalation protocol
wherein the peptide composition is administered as a single dose
followed six weeks later by a single booster shot of the same dose.
The dosages are 50, 500 and 5,000 micrograms per injection.
Drug-associated adverse effects (severity and reversibility) are
recorded.
[0908] There are three patient groupings. The first group is
injected with 50 micrograms of the peptide composition and the
second and third groups with 500 and 5,000 micrograms of peptide
composition, respectively. The patients within each group range in
age from 21-65 and represent diverse ethnic backgrounds. All of
them have a tumor that expresses 191P4D12(b).
[0909] Clinical manifestations or antigen-specific T-cell responses
are monitored to assess the effects of administering the peptide
compositions. The vaccine composition is found to be both safe and
efficacious in the treatment of 191P4D12(b)-associated disease.
Example 31
Induction of CTL Responses Using a Prime Boost Protocol
[0910] A prime boost protocol similar in its underlying principle
to that used to confirm the efficacy of a DNA vaccine in transgenic
mice, such as described above in the Example entitled "The Plasmid
Construct and the Degree to Which It Induces Immunogenicity," can
also be used for the administration of the vaccine to humans. Such
a vaccine regimen can include an initial administration of, for
example, naked DNA followed by a boost using recombinant virus
encoding the vaccine, or recombinant protein/polypeptide or a
peptide mixture administered in an adjuvant.
[0911] For example, the initial immunization may be performed using
an expression vector, such as that constructed in the Example
entitled "Construction of "Minigene" Multi-Epitope DNA Plasmids" in
the form of naked nucleic acid administered IM (or SC or ID) in the
amounts of 0.5-5 mg at multiple sites. The nucleic acid (0.1 to
1000 .mu.g) can also be administered using a gene gun. Following an
incubation period of 3-4 weeks, a booster dose is then
administered. The booster can be recombinant fowlpox virus
administered at a dose of 5-10.sup.7 to 5.times.10.sup.9 pfu. An
alternative recombinant virus, such as an MVA, canarypox,
adenovirus, or adeno-associated virus, can also be used for the
booster, or the polyepitopic protein or a mixture of the peptides
can be administered. For evaluation of vaccine efficacy, patient
blood samples are obtained before immunization as well as at
intervals following administration of the initial vaccine and
booster doses of the vaccine. Peripheral blood mononuclear cells
are isolated from fresh heparinized blood by Ficoll-Hypaque density
gradient centrifugation, aliquoted in freezing media and stored
frozen. Samples are assayed for CTL and HTL activity.
[0912] Analysis of the results indicates that a magnitude of
response sufficient to achieve a therapeutic or protective immunity
against 191P4D12(b) is generated.
Example 32
Administration of Vaccine Compositions Using Dendritic Cells
(DC)
[0913] Vaccines comprising peptide epitopes of the invention can be
administered using APCs, or "professional" APCs such as DC. In this
example, peptide-pulsed DC are administered to a patient to
stimulate a CTL response in vivo. In this method, dendritic cells
are isolated, expanded, and pulsed with a vaccine comprising
peptide CTL and HTL epitopes of the invention. The dendritic cells
are infused back into the patient to elicit CTL and HTL responses
in vivo. The induced CTL and HTL then destroy or facilitate
destruction, respectively, of the target cells that bear the
191P4D12(b) protein from which the epitopes in the vaccine are
derived.
[0914] For example, a cocktail of epitope-comprising peptides is
administered ex vivo to PBMC, or isolated DC therefrom. A
pharmaceutical to facilitate harvesting of DC can be used, such as
Progenipoietin.TM. (Monsanto, St. Louis, Mo.) or GM-CSF/IL-4. After
pulsing the DC with peptides, and prior to reinfusion into
patients, the DC are washed to remove unbound peptides.
[0915] As appreciated clinically, and readily determined by one of
skill based on clinical outcomes, the number of DC reinfused into
the patient can vary (see, e.g., Nature Med. 4:328, 1998; Nature
Med. 2:52, 1996 and Prostate 32:272, 1997). Although
2-50.times.10.sup.6 DC per patient are typically administered,
larger number of DC, such as 10.sup.7 or 10.sup.8 can also be
provided. Such cell populations typically contain between 50-90%
DC.
[0916] In some embodiments, peptide-loaded PBMC are injected into
patients without purification of the DC. For example, PBMC
generated after treatment with an agent such as Progenipoietin.TM.
are injected into patients without purification of the DC. The
total number of PBMC that are administered often ranges from
10.sup.8 to 10.sup.10. Generally, the cell doses injected into
patients is based on the percentage of DC in the blood of each
patient, as determined, for example, by immunofluorescence analysis
with specific anti-DC antibodies. Thus, for example, if
Progenipoietin.TM. mobilizes 2% DC in the peripheral blood of a
given patient, and that patient is to receive 5.times.10.sup.6 DC,
then the patient will be injected with a total of
2.5.times.10.sup.8 peptide-loaded PBMC. The percent DC mobilized by
an agent such as Progenipoietin.TM. is typically estimated to be
between 2-10%, but can vary as appreciated by one of skill in the
art.
[0917] Ex vivo Activation of CTL/HTL Responses
[0918] Alternatively, ex vivo CTL or HTL responses to 191P4D12(b)
antigens can be induced by incubating, in tissue culture, the
patient's, or genetically compatible, CTL or HTL precursor cells
together with a source of APC, such as DC, and immunogenic
peptides. After an appropriate incubation time (typically about
7-28 days), in which the precursor cells are activated and expanded
into effector cells, the cells are infused into the patient, where
they will destroy (CTL) or facilitate destruction (HTL) of their
specific target cells, ie., tumor cells.
Example 33
An Alternative Method of Identifying and Confirming Motif-Bearing
Peptides
[0919] Another method of identifying and confirming motif-bearing
peptides is to elute them from cells bearing defined MHC molecules.
For example, EBV transformed B cell lines used for tissue typing
have been extensively characterized to determine which HLA
molecules they express. In certain cases these cells express only a
single type of HLA molecule. These cells can be transfected with
nucleic acids that express the antigen of interest, e.g.
191P4D12(b). Peptides produced by endogenous antigen processing of
peptides produced as a result of transfection will then bind to HLA
molecules within the cell and be transported and displayed on the
cell's surface. Peptides are then eluted from the HLA molecules by
exposure to mild acid conditions and their amino acid sequence
determined, e.g., by mass spectral analysis (e.g., Kubo et al., J.
Immunol. 152:3913, 1994). Because the majority of peptides that
bind a particular HLA molecule are motif-bearing, this is an
alternative modality for obtaining the motif-bearing peptides
correlated with the particular HLA molecule expressed on the
cell.
[0920] Alternatively, cell lines that do not express endogenous HLA
molecules can be transfected with an expression construct encoding
a single HLA allele. These cells can then be used as described,
i.e., they can then be transfected with nucleic acids that encode
191P4D12(b) to isolate peptides corresponding to 191P4D12(b) that
have been presented on the cell surface. Peptides obtained from
such an analysis will bear motif(s) that correspond to binding to
the single HLA allele that is expressed in the cell.
[0921] As appreciated by one in the art, one can perform a similar
analysis on a cell bearing more than one HLA allele and
subsequently determine peptides specific for each HLA allele
expressed. Moreover, one of skill would also recognize that means
other than transfection, such as loading with a protein antigen,
can be used to provide a source of antigen to the cell.
Example 34
Complementary Polynucleotides
[0922] Sequences complementary to the 191P4D12(b)-encoding
sequences, or any parts thereof, are used to detect, decrease, or
inhibit expression of naturally occurring 191P4D12(b). Although use
of oligonucleotides comprising from about 15 to 30 base pairs is
described, essentially the same procedure is used with smaller or
with larger sequence fragments. Appropriate oligonucleotides are
designed using, e.g., OLIGO 4.06 software (National Biosciences)
and the coding sequence of 191P4D12(b). To inhibit transcription, a
complementary oligonucleotide is designed from the most unique 5'
sequence and used to prevent promoter binding to the coding
sequence. To inhibit translation, a complementary oligonucleotide
is designed to prevent ribosomal binding to a 191P4D12(b)-encoding
transcript.
Example 35
Purification of Naturally-occurring or Recombinant 191P4D12(b)
Using 191P4D12(b)-Specific Antibodies
[0923] Naturally occurring or recombinant 191P4D12(b) is
substantially purified by immunoaffinity chromatography using
antibodies specific for 191P4D12(b). An immunoaffinity column is
constructed by covalently coupling anti-191P4D12(b) antibody to an
activated chromatographic resin, such as CNBr-activated SEPHAROSE
(Amersham Pharmacia Biotech). After the coupling, the resin is
blocked and washed according to the manufacturer's
instructions.
[0924] Media containing 191P4D12(b) are passed over the
immunoaffinity column, and the column is washed under conditions
that allow the preferential absorbance of 191P4D12(b) (e.g., high
ionic strength buffers in the presence of detergent). The column is
eluted under conditions that disrupt antibody/191P4D12(b) binding
(e.g., a buffer of pH 2 to pH 3, or a high concentration of a
chaotrope, such as urea or thiocyanate ion), and GCR.P is
collected.
Example 36
Identification of Molecules Which Interact with 191P4D12(b)
[0925] 191P4D12(b), or biologically active fragments thereof, are
labeled with 121 1 Bolton-Hunter reagent. (See, e.g., Bolton et al.
(1973) Biochem. J. 133:529.) Candidate molecules previously arrayed
in the wells of a multi-well plate are incubated with the labeled
191P4D12(b), washed, and any wells with labeled 191P4D12(b) complex
are assayed. Data obtained using different concentrations of
191P4D12(b) are used to calculate values for the number, affinity,
and association of 191P4D12(b) with the candidate molecules.
Example 37
In Vivo Assay for 191P4D12(b) Tumor Growth Promotion
[0926] The effect of the 191P4D12(b) protein on tumor cell growth
is evaluated in vivo by evaluating tumor development and growth of
cells expressing or lacking 191P4D12(b). For example, SCID mice are
injected subcutaneously on each flank with 1.times.10.sup.6 of
either 3T3, prostate (e.g. PC3 cells), bladder (e.g. UM-UC3 cells),
kidney (e.g. CaKi cells), or lung (e.g. A427 cells) cancer cell
lines containing tkNeo empty vector or 191P4D12(b). At least two
strategies may be used: (1) Constitutive 191P4D12(b) expression
under regulation of a promoter such as a constitutive promoter
obtained from the genomes of viruses such as polyoma virus, fowlpox
virus (UK 2,211,504 published 5 Jul. 1989), adenovirus (such as
Adenovirus 2), bovine papilloma virus, avian sarcoma virus,
cytomegalovirus, a retrovirus, hepatitis-B virus and Simian Virus
40 (SV40), or from heterologous mammalian promoters, e.g., the
actin promoter or an immunoglobulin promoter, provided such
promoters are compatible with the host cell systems, and (2)
Regulated expression under control of an inducible vector system,
such as ecdysone, tetracycline, etc., provided such promoters are
compatible with the host cell systems. Tumor volume is then
monitored by caliper measurement at the appearance of palpable
tumors and followed over time to determine if
191P4D12(b)-expressing cells grow at a faster rate and whether
tumors produced by 191P4D12(b)-expressing cells demonstrate
characteristics of altered aggressiveness (e.g. enhanced
metastasis, vascularization, reduced responsiveness to
chemotherapeutic drugs).
[0927] Additionally, mice can be implanted with 1.times.10.sup.5 of
the same cells orthotopically to determine if 191P4D12(b) has an
effect on local growth in the prostate, and whether 191P4D12(b)
affects the ability of the cells to metastasize, specifically to
lymph nodes, and bone (Miki T et al, Oncol Res. 2001;12:209; Fu X
et al, Int J Cancer. 1991, 49:938). The effect of 191P4D12(b) on
bone tumor formation and growth may be assessed by injecting tumor
cells intratibially.
[0928] The assay is also useful to determine the 191P4D12(b)
inhibitory effect of candidate therapeutic compositions, such as
for example, 191P4D12(b) intrabodies, 191P4D12(b) antisense
molecules and ribozymes.
Example 38
191P4D12(b) Monoclonal Antibody-mediated Inhibition of Tumors In
Vivo
[0929] The significant expression of 191P4D12(b) in cancer tissues
and surface localization, together with its restrictive expression
in normal tissues makes 191P4D12(b) a good target for antibody
therapy. Similarly, 191P4D12(b) is a target for T cell-based
immunotherapy. Thus, the therapeutic efficacy of anti-191P4D12(b)
mAbs in human cancer xenograft mouse models, including prostate,
lung, bladder, kidney and other -191P4D12(b)cancers listed in table
1, is evaluated by using recombinant cell lines such as
PC3-191P4D12(b), UM-UC3-191P4D12(b), CaKi--191P4D12(b),
A427-191P4D12(b) and 3T3-191P4D12(b) (see, e.g., Kaighn, M. E., et
al., Invest Urol, 1979. 17(1): 16-23), as well as human prostate,
kidney and bladder xenograft models such as LAPC 9AD, AGS-K3 and
AGS-B1 (Saffran et al PNAS 1999, 10:1073-1078).
[0930] Antibody efficacy on tumor growth and metastasis formation
is studied, e.g., in a mouse orthotopic prostate, kidney, bladder,
and lung cancer xenograft models. The antibodies can be
unconjugated, as discussed in this Example, or can be conjugated to
a therapeutic modality, as appreciated in the art. Anti-191P4D12(b)
mAbs inhibit formation of tumors in prostate kidney, bladder and
lung xenografts. Anti-191P4D12(b) mAbs also retard the growth of
established orthotopic tumors and prolonged survival of
tumor-bearing mice. These results indicate the utility of
anti-191P4D12(b) mAbs in the treatment of local and advanced stages
several solid tumors. (See, e.g., Saffran, D., et al., PNAS
10:1073-1078 or world wide web URL
pnas.org/cgi/doi/10.1073/pnas.051624698).
[0931] Administration of the anti-191P4D12(b) mAbs led to
retardation of established orthotopic tumor growth and inhibition
of metastasis to distant sites, resulting in a significant
prolongation in the survival of tumor-bearing mice. These studies
indicate that 191P4D12(b) as an attractive target for immunotherapy
and demonstrate the therapeutic potential of anti-191P4D12(b) mAbs
for the treatment of local and metastatic prostate cancer. This
example indicates that unconjugated 191P4D12(b) monoclonal
antibodies are effective to inhibit the growth of human prostate,
kidney, bladder and lung tumor xenografts grown in SCID mice;
accordingly a combination of such efficacious monoclonal antibodies
is also effective.
[0932] Tumor Inhibition Using Multiple Unconjugated 191P4D12(b)
mAbs
[0933] Materials and Methods
[0934] 191P4D12(b) Monoclonal Antibodies:
[0935] Monoclonal antibodies are raised against 191P4D12(b) as
described in the Example entitled "Generation of 191P4D12(b)
Monoclonal Antibodies (mAbs)." The antibodies are characterized by
ELISA, Western blot, FACS, and immunoprecipitation for their
capacity to bind 191P4D12(b). Epitope mapping data for the
anti-191P4D12(b) mAbs, as determined by ELISA and Western analysis,
recognize epitopes on the 191P4D12(b) protein. Immunohistochemical
analysis of prostate, kidney, bladder and lung cancer tissues and
cells with these antibodies is performed.
[0936] The monoclonal antibodies are purified from ascites or
hybridoma tissue culture supernatants by Protein-G Sepharose
chromatography, dialyzed against PBS, filter sterilized, and stored
at -20.degree. C. Protein determinations are performed by a
Bradford assay (Bio-Rad, Hercules, Calif.). A therapeutic
monoclonal antibody or a cocktail comprising a mixture of
individual monoclonal antibodies is prepared and used for the
treatment of mice receiving subcutaneous or orthotopic injections
of PC3, UM-UC3, CaKi and A427 tumor xenografts.
[0937] Cell Lines and Xenografts
[0938] The cancer cell lines, PC3, UM-UC3, CaKi, and A427 cell line
as well as the fibroblast line NIH 3T3 (American Type Culture
Collection) are maintained in RPMI (PC3) and DMEM (UM-UC3, CaKi,
and A427, 3T3) respectively, supplemented with L-glutamine and 10%
FBS.
[0939] PC3-191P4D12(b), UM-UC3-191P4D12(b), CaKi-191P4D12(b),
A427-191P4D12(b) and 3T3-191P4D12(b) cell populations are generated
by retroviral gene transfer as described in Hubert, R. S., et al.,
Proc Natl Acad Sci U S A, 1999. 96(25): 14523. The LAPC-9
xenograft, which expresses a wild-type androgen receptor and
produces prostate-specific antigen (PSA), is passaged in 6- to
8-week-old male ICR-severe combined immunodeficient (SCID) mice
(Taconic Farms) by s.c. trocar implant (Craft, N., et al., Nat Med.
1999, 5:280). Single-cell suspensions of LAPC-9 tumor cells are
prepared as described in Craft, et al. Similarly, kidney (AGS-K3)
and bladder (AGS-B1) patient-derived xenografts are passaged in 6-
to 8-week-old male ICR-SCID mice.
[0940] Xenograft Mouse Models.
[0941] Subcutaneous (s.c.) tumors are generated by injection of
2.times.10.sup.6 cancer cells mixed at a 1:1 dilution with Matrigel
(Collaborative Research) in the right flank of male SCID mice. To
test antibody efficacy on tumor formation, i.e. antibody injections
are started on the same day as tumor-cell injections. As a control,
mice are injected with either purified mouse IgG (ICN) or PBS; or a
purified monoclonal antibody that recognizes an irrelevant antigen
not expressed in human cells. In preliminary studies, no difference
is found between mouse IgG or PBS on tumor growth. Tumor sizes are
determined by caliper measurements, and the tumor volume is
calculated as length.times.width.times.height. Mice with
Subcutaneous tumors greater than 1.5 cm in diameter are
sacrificed.
[0942] Orthotopic injections are performed under anesthesia by
using ketamine/xylazine. For prostate orthotopic studies, an
incision is made through the abdomen to expose the prostate and
LAPC or PC3 tumor cells (5.times.10.sup.5) mixed with Matrigel are
injected into the prostate capsule in a 10-.mu.l volume. To monitor
tumor growth, mice are palpated and blood is collected on a weekly
basis to measure PSA levels. For kidney orthotopic models, an
incision is made through the abdominal muscles to expose the
kidney. AGS-K3 cells mixed with Matrigel are injected under the
kidney capsule. The mice are segregated into groups for the
appropriate treatments, with anti-191P4D12(b) or control mAbs being
injected i.p.
[0943] Anti-191P4D12(b) mAbs Inhibit Growth of
191P4D12(b)-Expressinq Xenograft-Cancer Tumors
[0944] The effect of anti-191P4D12(b) mAbs on tumor formation is
tested by using cell line (e.g. PC3, UM-UC3, CaKi, A427, and 3T3)
and patient-derived tumor (e.g. LAPC9, AGS-K3, AGS-B1) orthotopic
models. As compared with the s.c. tumor model, the orthotopic
model, which requires injection of tumor cells directly in the
mouse organ, such as prostate, bladder, kidney or lung, results in
a local tumor growth, development of metastasis in distal sites,
deterioration of mouse health, and subsequent death (Saffran, D.,
et al., PNAS supra). The features make the orthotopic model more
representative of human disease progression and allowed us to
follow the therapeutic effect of mAbs on clinically relevant end
points. For example, tumor cells are injected into the mouse
prostate, and 2 days later, the mice are segregated into two groups
and treated with either: a) 200-500 .mu.g, of anti-191P4D12(b) Ab,
or b) PBS three times per week for two to five weeks.
[0945] A major advantage of the orthotopic cancer models is the
ability to study the development of metastases. Formation of
metastasis in mice bearing established orthotopic tumors is studies
by IHC analysis on lung sections using an antibody against a
tumor-specific cell-surface protein such as anti-CK20 for prostate
cancer (Lin S et al, Cancer Detect Prev. 2001;25:202).
[0946] Another advantage of xenograft cancer models is the ability
to study neovascularization and angiogenesis. Tumor growth is
partly dependent on new blood vessel development. Although the
capillary system and developing blood network is of host origin,
the initiation and architecture of the neovascular is regulated by
the xenograft tumor (Davidoff A M et al, Clin Cancer Res.
2001;7:2870; Solesvik O et al,, Eur J Cancer Clin Oncol. 1984,
20:1295). The effect of antibody and small molecule on
neovascularization is studied in accordance with procedures known
in the art, such as by IHC analysis of tumor tissues and their
surrounding microenvironment.
[0947] Mice bearing established orthotopic tumors are administered
1000 .mu.g injections of either anti-191P4D12(b) mAb or PBS over a
4-week period. Mice in both groups are allowed to establish a high
tumor burden, to ensure a high frequency of metastasis formation in
mouse lungs. Mice then are killed and their bladders, livers, bone
and lungs are analyzed for the presence of tumor cells by IHC
analysis. These studies demonstrate a broad anti-tumor efficacy of
anti-191P4D12(b) antibodies on initiation and progression of
prostate cancer in xenograft mouse models. Anti-191P4D12(b)
antibodies inhibit tumor formation of tumors as well as retarding
the growth of already established tumors and prolong the survival
of treated mice. Moreover, anti-191P4D12(b) mAbs demonstrate a
dramatic inhibitory effect on the spread of local prostate tumor to
distal sites, even in the presence of a large tumor burden. Thus,
anti-191P4D12(b) mAbs are efficacious on major clinically relevant
end points (tumor growth), prolongation of survival, and
health.
Example 39
Therapeutic and Diagnostic use of Anti-191P4D12(b) Antibodies in
Humans
[0948] Anti-191 P4D12(b) monoclonal antibodies are safely and
effectively used for diagnostic, prophylactic, prognostic and/or
therapeutic purposes in humans. Western blot and
immunohistochemical analysis of cancer tissues and cancer
xenografts with anti-191P4D12(b) mAb show strong extensive staining
in carcinoma but significantly lower or undetectable levels in
normal tissues. Detection of 191P4D12(b) in carcinoma and in
metastatic disease demonstrates the usefulness of the mAb as a
diagnostic and/or prognostic indicator. Anti-191P4D12(b) antibodies
are therefore used in diagnostic applications such as
immunohistochemistry of kidney biopsy specimens to detect cancer
from suspect patients.
[0949] As determined by flow cytometry, anti-191P4D12(b) mAb
specifically binds to carcinoma cells. Thus, anti-191P4D12(b)
antibodies are used in diagnostic whole body imaging applications,
such as radioimmunoscintigraphy and radioimmunotherapy, (see, e.g.,
Potamianos S., et. al. Anticancer Res 20(2A):925-948 (2000)) for
the detection of localized and metastatic cancers that exhibit
expression of 191P4D12(b). Shedding or release of an extracellular
domain of 191P4D12(b) into the extracellular milieu, such as that
seen for alkaline phosphodiesterase B10 (Meerson, N. R., Hepatology
27:563-568 (1998)), allows diagnostic detection of 191P4D12(b) by
anti-191P4D12(b) antibodies in serum and/or urine samples from
suspect patients.
[0950] Anti-191P4D12(b) antibodies that specifically bind
191P4D12(b) are used in therapeutic applications for the treatment
of cancers that express 191P4D12(b). Anti-191P4D12(b) antibodies
are used as an unconjugated modality and as conjugated form in
which the antibodies are attached to one of various therapeutic or
imaging modalities well known in the art, such as a prodrugs,
enzymes or radioisotopes. In preclinical studies, unconjugated and
conjugated anti-191P4D12(b) antibodies are tested for efficacy of
tumor prevention and growth inhibition in the SCID mouse cancer
xenograft models, e.g., kidney cancer models AGS-K3 and AGS-K6,
(see, e.g., the Example entitled "191P4D12(b) Monoclonal
Antibody-mediated Inhibition of Bladder and Lung Tumors In Vivo").
Either conjugated and unconjugated anti-191P4D12(b) antibodies are
used as a therapeutic modality in human clinical trials either
alone or in combination with other treatments as described in
following Examples.
Example 40
Human Clinical Trials for the Treatment and Diagnosis of Human
Carcinomas through use of Human Anti-191P4D12(b) Antibodies In
vivo
[0951] Antibodies are used in accordance with the present invention
which recognize an epitope on 191P4D12(b), and are used in the
treatment of certain tumors such as those listed in Table I. Based
upon a number of factors, including 191P4D12(b) expression levels,
tumors such as those listed in Table I are presently preferred
indications. In connection with each of these indications, three
clinical approaches are successfully pursued.
[0952] I.) Adjunctive therapy: In adjunctive therapy, patients are
treated with anti-191P4D12(b) antibodies in combination with a
chemotherapeutic or antineoplastic agent and/or radiation therapy.
Primary cancer targets, such as those listed in Table I, are
treated under standard protocols by the addition anti-191P4D12(b)
antibodies to standard first and second line therapy. Protocol
designs address effectiveness as assessed by reduction in tumor
mass as well as the ability to reduce usual doses of standard
chemotherapy. These dosage reductions allow additional and/or
prolonged therapy by reducing dose-related toxicity of the
chemotherapeutic agent. Anti-191P4D12(b) antibodies are utilized in
several adjunctive clinical trials in combination with the
chemotherapeutic or antineoplastic agents adriamycin (advanced
prostrate carcinoma), cisplatin (advanced head and neck and lung
carcinomas), taxol (breast cancer), and doxorubicin
(preclinical).
[0953] II.) Monotherapy: In connection with the use of the
anti-191P4D12(b) antibodies in monotherapy of tumors, the
antibodies are administered to patients without a chemotherapeutic
or antineoplastic agent. In one embodiment, monotherapy is
conducted clinically in end stage cancer patients with extensive
metastatic disease. Patients show some disease stabilization.
Trials demonstrate an effect in refractory patients with cancerous
tumors.
[0954] III.) Imaging Agent: Through binding a radionuclide (e.g.,
iodine or yttrium (I.sup.131, Y.sup.90) to anti-191P4D12(b)
antibodies, the radiolabeled antibodies are utilized as a
diagnostic and/or imaging agent. In such a role, the labeled
antibodies localize to both solid tumors, as well as, metastatic
lesions of cells expressing 191P4D12(b). In connection with the use
of the anti-191P4D12(b) antibodies as imaging agents, the
antibodies are used as an adjunct to surgical treatment of solid
tumors, as both a pre-surgical screen as well as a post-operative
follow-up to determine what tumor remains and/or returns. In one
embodiment, a (.sup.111In)-191P4D12(b) antibody is used as an
imaging agent in a Phase I human clinical trial in patients having
a carcinoma that expresses 191P4D12(b) (by analogy see, e.g., Divgi
et al. J. Natl. Cancer Inst. 83:97-104 (1991)). Patients are
followed with standard anterior and posterior gamma camera. The
results indicate that primary lesions and metastatic lesions are
identified.
[0955] Dose and Route of Administration
[0956] As appreciated by those of ordinary skill in the art, dosing
considerations can be determined through comparison with the
analogous products that are in the clinic. Thus, anti-191P4D12(b)
antibodies can be administered with doses in the range of 5 to 400
mg/m.sup.2, with the lower doses used, e.g., in connection with
safety studies. The affinity of anti-191P4D12(b) antibodies
relative to the affinity of a known antibody for its target is one
parameter used by those of skill in the art for determining
analogous dose regimens. Further, anti-191P4D12(b) antibodies that
are fully human antibodies, as compared to the chimeric antibody,
have slower clearance; accordingly, dosing in patients with such
fully human anti-191P4D12(b) antibodies can be lower, perhaps in
the range of 50 to 300 mg/m.sup.2, and still remain efficacious.
Dosing in mg/m.sup.2, as opposed to the conventional measurement of
dose in mg/kg, is a measurement based on surface area and is a
convenient dosing measurement that is designed to include patients
of all sizes from infants to adults.
[0957] Three distinct delivery approaches are useful for delivery
of anti-191P4D12(b) antibodies. Conventional intravenous delivery
is one standard delivery technique for many tumors. However, in
connection with tumors in the peritoneal cavity, such as tumors of
the ovaries, biliary duct, other ducts, and the like,
intraperitoneal administration may prove favorable for obtaining
high dose of antibody at the tumor and to also minimize antibody
clearance. In a similar manner, certain solid tumors possess
vasculature that is appropriate for regional perfusion. Regional
perfusion allows for a high dose of antibody at the site of a tumor
and minimizes short term clearance of the antibody.
[0958] Clinical Development Plan (CDP)
[0959] Overview: The CDP follows and develops treatments of
anti-191P4D12(b) antibodies in connection with adjunctive therapy,
monotherapy, and as an imaging agent. Trials initially demonstrate
safety and thereafter confirm efficacy in repeat doses. Trails are
open label comparing standard chemotherapy with standard therapy
plus anti-191P4D12(b) antibodies. As will be appreciated, one
criteria that can be utilized in connection with enrollment of
patients is 191P4D12(b) expression levels in their tumors as
determined by biopsy.
[0960] As with any protein or antibody infusion-based therapeutic,
safety concerns are related primarily to (i) cytokine release
syndrome, i.e., hypotension, fever, shaking, chills; (ii) the
development of an immunogenic response to the material (i.e.,
development of human antibodies by the patient to the antibody
therapeutic, or HAHA response); and, (iii) toxicity to normal cells
that express 191P4D12(b). Standard tests and follow-up are utilized
to monitor each of these safety concerns. Anti-191P4D12(b)
antibodies are found to be safe upon human administration.
Example 41
Human Clinical Trial Adjunctive Therapy with Human Anti-191P4D12(b)
Antibody and Chemotherapeutic Agent
[0961] A phase I human clinical trial is initiated to assess the
safety of six intravenous doses of a human anti-191P4D12(b)
antibody in connection with the treatment of a solid tumor, e.g., a
cancer of a tissue listed in Table I. In the study, the safety of
single doses of anti-191P4D12(b) antibodies when utilized as an
adjunctive therapy to an antineoplastic or chemotherapeutic agent
as defined herein, such as, without limitation: cisplatin,
topotecan, doxorubicin, adriamycin, taxol, or the like, is
assessed. The trial design includes delivery of six single doses of
an anti-191P4D12(b) antibody with dosage of antibody escalating
from approximately about 25 mg/m.sup.2 to about 275 mg/m.sup.2 over
the course of the treatment in accordance with the following
schedule:
4 Day 0 Day 7 Day 14 Day 21 Day 28 Day 35 mAb Dose 25 75 125 175
225 275 mg/m.sup.2 mg/m.sup.2 mg/m.sup.2 mg/m.sup.2 mg/m.sup.2
mg/m.sup.2 Chemotherapy + + + + + + (standard dose)
[0962] Patients are closely followed for one-week following each
administration of antibody and chemotherapy. In particular,
patients are assessed for the safety concerns mentioned above: (i)
cytokine release syndrome, i.e., hypotension, fever, shaking,
chills; (ii) the development of an immunogenic response to the
material (i.e., development of human antibodies by the patient to
the human antibody therapeutic, or HAHA response); and, (iii)
toxicity to normal cells that express 191P4D12(b). Standards tests
and follow-up are utilized to monitor each of these safety
concerns. Patients are also assessed for clinical outcome, and
particularly reduction in tumor mass as evidenced by MRI or other
imaging.
[0963] The anti-191P4D12(b) antibodies are demonstrated to be safe
and efficacious, Phase II trials confirm the efficacy and refine
optimum dosing.
Example 42
Human Clinical Trial: Monotherapy with Human Anti-191P4D12(b)
Antibody
[0964] Anti-191P4D12(b) antibodies are safe in connection with the
above-discussed adjunctive trial, a Phase II human clinical trial
confirms the efficacy and optimum dosing for monotherapy. Such
trial is accomplished, and entails the same safety and outcome
analyses, to the above-described adjunctive trial with the
exception being that patients do not receive chemotherapy
concurrently with the receipt of doses of anti-191P4D12(b)
antibodies.
Example 43
Human Clinical Trial: Diagnostic Imaging with Anti-191P4D12(b)
Antibody
[0965] Once again, as the adjunctive therapy discussed above is
safe within the safety criteria discussed above, a human clinical
trial is conducted concerning the use of anti-191P4D12(b)
antibodies as a diagnostic imaging agent. The protocol is designed
in a substantially similar manner to those described in the art,
such as in Divgi et al. J. Natl. Cancer Inst. 83:97-104 (1991). The
antibodies are found to be both safe and efficacious when used as a
diagnostic modality.
Example 44
Homology Comparison of 191P4D12(b) to Known Sequences
[0966] The human 191P4D12(b) protein exhibit a high degree of
homology to a known human protein, namely Ig superfamily receptor
LNIR (gi 14714574), also known as human nectin 4 (gi 16506807).
Human LNIR shows 100% identity to 191P4D12(b) at the protein level.
The mouse homolog of 191P4D12(b) has been identified as murine
nectin 4 (gi 18874521). It shows strong homology to 191P4D12(b),
exhibiting 92% identity and 95% homology to 191P4D12(b). (See, FIG.
4).
[0967] The prototype member of the 191P4D12(b) family,
191P4D12(b)v.1, is a 510 amino acids protein, with the N-terminus
located extracellulary and intracellular C-terminus. Initial
bioinformatics analysis using topology prediction programs
suggested that 191P2D14 may contain 2 transmembranes based on
hydrophobicity profile. However, the first hydrophobic domain was
identified as a signal sequence, rendering 191P2D12 a type I
membrane protein, with an extracellular N-terminus.
[0968] The 191P4D12(b) gene has several variants, including one SNP
represented in 191P4D12(b) v.2, an N-terminal deletion variant
represented in 191P4D12(b) v.6 and 191P4D12(b) v.7 which lacks 25
amino acids between amino acids 411 and 412 of 191P4D12(b) v.1.
[0969] Motif analysis revealed the presence of several protein
functional motifs in the 191P4D12(b) protein (Table L). Two
immunoglobulin domains have been identified at positions 45-129 and
263-317. In addition, 191P4D12(b) contains a cadherin signature
which includes and RGD sequence. Immunoglobulin domains are found
in numerous proteins and participate in protein-protein such
including protein-ligand interactions (Weismann et al, J Mol Med
2000, 78:247). In addition, Ig-domains function in cell adhesion,
allowing the interaction of leukocytes and blood-born cells with
the endothelium (Wang and Springer, Immunol Rev 1998, 163:197).
Cadherins are single transmembrane proteins containing
immunoglobulin like domains, and are involved in cell adhesion and
sorting (Shan et al, Biophys Chem 1999, 82:157). They mediate
tissue-specific cell adhesion, such as adhesion of lymphocytes to
the surface of epithelial cells. Finally, the closest homolog to
191P4D12(b) is Nectin4, a known adhesion molecule that regulates
epithelial and endothelial junctions, strongly suggesting that
191P4D12(b) participates in cell adhesion (Reymond N et al, J Biol
Chem 2001, 276:43205).
[0970] The motifs found in 191P4D12(b) can participate in tumor
growth and progression by enhancing the initial stages of
tumorigenesis, such as tumor take or establishment of a tumor, by
allowing adhesion to basement membranes and surrounding cells, by
mediating cell communication and survival.
[0971] Accordingly, when 191P4D12(b) functions as a regulator of
tumor establishment, tumor formation, tumor growth, cell signaling
or as a modulator of transcription involved in activating genes
associated with survival, invasion, tumorigenesis or proliferation,
191P4D12(b) is used for therapeutic, diagnostic, prognostic and/or
preventative purposes. In addition, when a molecule, such as a
variant or SNP of 191P4D12(b) is expressed in cancerous tissues,
such as those listed in Table I, they are used for therapeutic,
diagnostic, prognostic and/or preventative purposes.
Example 45
Regulation of Transcription
[0972] The cell surface localization of 191P4D12(b) coupled to the
presence of Ig-domains within its sequence indicate that
191P4D12(b) modulates signal transduction and the transcriptional
regulation of eukaryotic genes. Regulation of gene expression is
confirmed, e.g., by studying gene expression in cells expressing or
lacking 191P4D12(b). For this purpose, two types of experiments are
performed.
[0973] In the first set of experiments, RNA from parental and
191P4D12(b)-expressing cells are extracted and hybridized to
commercially available gene arrays (Clontech) (Smid-Koopman E et
al. Br J Cancer. 2000. 83:246). Resting cells as well as cells
treated with FBS, androgen or growth factors are compared.
Differentially expressed genes are identified in accordance with
procedures known in the art. The differentially expressed genes are
then mapped to biological pathways (Chen K et al. Thyroid. 2001.
11:41.).
[0974] In the second set of experiments, specific transcriptional
pathway activation is evaluated using commercially available
(Stratagene) luciferase reporter constructs including: NFkB-luc,
SRE-luc, ELK1-luc, ARE-luc, p53-luc, and CRE-luc. These
transcriptional reporters contain consensus binding sites for known
transcription factors that lie downstream of well-characterized
signal transduction pathways, and represent a good tool to
ascertain pathway activation and screen for positive, and negative
modulators of pathway activation.
[0975] Thus, 191P4D12(b) plays a role in gene regulation, and it is
used as a target for diagnostic, prognostic, preventative and/or
therapeutic purposes.
Example 46
Identification and Confirmation of Potential Signal Transduction
Pathways
[0976] Many mammalian proteins have been reported to interact with
signaling molecules and to participate in regulating signaling
pathways. (J Neurochem. 2001; 76:217-223). Immunoglobulin-like
molecules in particular has been associated with several tyrpsine
kinases including Lyc, Blk, syk(), the MAPK signaling cascade that
control cell mitogenesis and calcium flux (Vilen J et al, J Immunol
1997, 159:231; Jiang F, Jia Y, Cohen I. Blood. 2002, 99:3579). In
addition, the 191P4D12(b) protein contains several phosphorylation
sites (see Table VI) indicating an association with specific
signaling cascades. Using immunoprecipitation and Western blotting
techniques, proteins are identified that associate with 191P4D12(b)
and mediate signaling events. Several pathways known to play a role
in cancer biology can be regulated by 191P4D12(b), including
phospholipid pathways such as PI3K, AKT, etc, adhesion and
migration pathways, including FAK, Rho, Rac-1, catenin, etc, as
well as mitogenic/survival cascades such as ERK, p38, etc (Cell
Growth Differ. 2000,11:279; J Biol Chem. 1999, 274:801; Oncogene.
2000, 19:3003, J. Cell Biol. 1997, 138:913.).). In order to
determine whether expression of 191P4D12(b) is sufficient to
regulate specific signaling pathways not otherwise active in
resting PC3 cells, the effect of these genes on the activation of
the p38 MAPK cascade was investigated in the prostate cancer cell
line PC3 (FIGS. 21A-B). Activation of the p38 kinase is dependent
on its phosphorylation on tyrosine and serine residues.
Phosphorylated p38 can be distinguished from the non-phosphorylated
state by a Phospho-p38 mAb. This phospho-specific Ab was used to
study the phosphorylation state of p38 in engineered PC3 cell
lines.
[0977] PC3 cells stably expressing 191P4D12(b) neo were grown
overnight in either 1% or 10% FBS. Whole cell lysates were analyzed
by western blotting. PC3 cells treated with the known p38
activators, NaSal or TNF, were used as a positive control. The
results show that while expression of the control neo gene has no
effect on p38 phosphorylation, expression of 191P4D12(b) in PC3
cells is sufficient to induce the activation of the p38 pathway
(FIG. 21A). The results were verified using western blotting with
an anti-p38 Ab, which shows equal protein loading on the gels (FIG.
21B). In another set of experiments, the sufficiency of expression
of 191P4D12(b) in the prostate cancer cell line PC3 to activate the
mitogenic MAPK pathway, namely the ERK cascade, was examined (FIGS.
22A-B). Activation of ERK is dependent on its phosphorylation on
tyrosine and serine residues. Phosphorylated ERK can be
distinguished from the non-phosphorylated state by a Phospho-ERK
mAb. This phospho-specific Ab was used to study the phosphorylation
state of ERK in engineered PC3 cell lines. PC3 cells, expressing an
activated form of Ras, were used as a positive control. The results
show that while expression of the control neo gene has no effect on
ERK phosphorylation, expression of 191P4D12(b) in PC3 cells is
sufficient to induce an increase in ERK phosphorylation (FIG. 22A).
These results were verified using anti-ERK western blotting (FIG.
22B) and confirm the activation of the ERK pathway by 191P4D12(b)
and STEAP-2.
[0978] Since FBS contains several components that may contribute to
receptor-mediated ERK activation, we examined the effect of
191P4D12(b) in low and optimal levels of FBS. PC3 cells expressing
neo or 191P4D12(b) were grown in either 0.1% or 10% FBS overnight.
The cells were analyzed by anti-Phospho-ERK western blotting. This
experiment shows that 191P4D12(b) induces the phosphorylation of
ERK in 0.1% FBS, and confirms that expression of 191P4D12(b) is
sufficient to induce activation of the ERK signaling cascade in the
absence of additional stimuli.
[0979] To confirm that 191P4D12(b) directly or indirectly activates
known signal transduction pathways in cells, luciferase (luc) based
transcriptional reporter assays are carried out in cells expressing
individual genes. These transcriptional reporters contain
consensus-binding sites for known transcription factors that lie
downstream of well-characterized signal transduction pathways. The
reporters and examples of these associated transcription factors,
signal transduction pathways, and activation stimuli are listed
below.
[0980] 1. NFkB-luc, NFkB/Rel; Ik-kinase/SAPK;
growth/apoptosis/stress
[0981] 2. SRE-luc, SRF/TCF/ELK1; MAPK/SAPK;
growth/differentiation
[0982] 3. AP-1-luc, FOS/JUN; MAPK/SAPK/PKC;
growth/apoptosis/stress
[0983] 4. ARE-luc, androgen receptor; steroids/MAPK;
growth/differentiation/apoptosis
[0984] 5. p53-luc, p53; SAPK; growth/differentiation/apoptosis
[0985] 6. CRE-luc, CREB/ATF2; PKA/p38; growth/apoptosis/stress
[0986] 7. TCF-luc, TCF/Lef; -catenin, Adhesion/invasion
[0987] Gene-mediated effects can be assayed in cells showing mRNA
expression. Luciferase reporter plasmids can be introduced by
lipid-mediated transfection (TFX-50, Promega). Luciferase activity,
an indicator of relative transcriptional activity, is measured by
incubation of cell extracts with luciferin substrate and
luminescence of the reaction is monitored in a luminometer.
[0988] Signaling pathways activated by 191P4D12(b) are mapped and
used for the identification and validation of therapeutic targets.
When 191P4D12(b) is involved in cell signaling, it is used as
target for diagnostic, prognostic, preventative and/or therapeutic
purposes.
Example 47
Involvement in Tumor Progression
[0989] Based on the role of Ig-domains and cadherin motifs in cell
growth and signal transduction, the 191P4D12(b) gene can contribute
to the growth, invasion and transformation of cancer cells. The
role of 191P4D12(b) in tumor growth is confirmed in a variety of
primary and transfected cell lines including prostate cell lines,
as well as NIH 3T3 cells engineered to stably express 191P4D12(b).
Parental cells lacking 191P4D12(b) and cells expressing 191P4D12(b)
are evaluated for cell growth using a well-documented proliferation
assay (Fraser S P, Grimes J A, Djamgoz M B. Prostate. 2000;44:61,
Johnson D E, Ochieng J, Evans S L. Anticancer Drugs. 1996,
7:288).
[0990] To confirm the role of 191P4D12(b) in the transformation
process, its effect in colony forming assays is investigated.
Parental NIH-3T3 cells lacking 191P4D12(b) are compared to NIH-3T3
cells expressing 191P4D12(b), using a soft agar assay under
stringent and more permissive conditions (Song Z. et al. Cancer
Res. 2000;60:6730).
[0991] To confirm the role of 191P4D12(b) in invasion and
metastasis of cancer cells, a well-established assay is used, e.g.,
a Transwell Insert System assay (Becton Dickinson) (Cancer Res.
1999; 59:6010). Control cells, including prostate, breast and
kidney cell lines lacking 191P4D12(b) are compared to cells
expressing 191P4D12(b). Cells are loaded with the fluorescent dye,
calcein, and plated in the top well of the Transwell insert coated
with a basement membrane analog. Invasion is determined by
fluorescence of cells in the lower chamber relative to the
fluorescence of the entire cell population.
[0992] 191P4D12(b) can also play a role in cell cycle and
apoptosis. Parental cells and cells expressing 191P4D12(b) are
compared for differences in cell cycle regulation using a
well-established BrdU assay (Abdel-Malek Z A. J Cell Physiol. 1988,
136:247). In short, cells are grown under both optimal (full serum)
and limiting (low serum) conditions are labeled with BrdU and
stained with anti-BrdU Ab and propidium iodide. Cells are analyzed
for entry into the G1, S, and G2M phases of the cell cycle.
Alternatively, the effect of stress on apoptosis is evaluated in
control parental cells and cells expressing 191P4D12(b), including
normal and tumor prostate cells. Engineered and parental cells are
treated with various chemotherapeutic agents, such as etoposide,
taxol, etc, and protein synthesis inhibitors, such as
cycloheximide. Cells are stained with annexin V-FITC and cell death
is measured by FACS analysis. The modulation of cell death by
191P4D12(b) can play a critical role in regulating tumor
progression and tumor load.
[0993] When 191P4D12(b) plays a role in cell growth,
transformation, invasion or apoptosis, it is used as a target for
diagnostic, prognostic, preventative and/or therapeutic
purposes.
Example 48
Involvement in Angiogenesis
[0994] Angiogenesis or new capillary blood vessel formation is
necessary for tumor growth (Hanahan D, Folkman J. Cell. 1996,
86:353; Folkman J. Endocrinology. 1998 139:441). Based on the
effect of cadherins on tumor cell adhesion and their interaction
with endothelial cells, 191P4D12(b) plays a role in angiogenesis
(Mareel and Leroy: Physiol Rev, 83:337; DeFouw L et al, Microvasc
Res 2001, 62:263). Several assays have been developed to measure
angiogenesis in vitro and in vivo, such as the tissue culture
assays endothelial cell tube formation and endothelial cell
proliferation. Using these assays as well as in vitro
neovasculanization, the role of 191P4D12(b) in angiogenesis,
enhancement or inhibition, is confirmed.
[0995] For example, endothelial cells engineered to express
191P4D12(b) are evaluated using tube formation and proliferation
assays. The effect of 191P4D12(b) is also confirmed in animal
models in vivo. For example, cells either expressing or lacking
191P4D12(b) are implanted subcutaneously in immunocompromised mice.
Endothelial cell migration and angiogenesis are evaluated 5-15 days
later using immunohistochemistry techniques. 191P4D12(b) affects
angiogenesis, and it is used as a target for diagnostic,
prognostic, preventative and/or therapeutic purposes.
Example 49
Involvement in Protein-Protein Interactions
[0996] Ig-domains and cadherin motifs have been shown to mediate
interaction with other proteins, including cell surface protein.
Using immunoprecipitation techniques as well as two yeast hybrid
systems, proteins are identified that associate with 191P4D12(b).
Immunoprecipitates from cells expressing 191P4D12(b) and cells
lacking 191P4D12(b) are compared for specific protein-protein
associations.
[0997] Studies are performed to confirm the extent of association
of 191P4D12(b) with effector molecules, such as nuclear proteins,
transcription factors, kinases, phosphates etc. Studies comparing
191P4D12(b) positive and 191P4D12(b) negative cells as well as
studies comparing unstimulated/resting cells and cells treated with
epithelial cell activators, such as cytokines, growth factors,
androgen and anti-integrin Ab reveal unique interactions.
[0998] In addition, protein-protein interactions are confirmed
using two yeast hybrid methodology (Curr. Opin. Chem Biol. 1999,
3:64). A vector carrying a library of proteins fused to the
activation domain of a transcription factor is introduced into
yeast expressing a 191P4D12(b)-DNA-binding domain fusion protein
and a reporter construct. Protein-protein interaction is detected
by colorimetric reporter activity. Specific association with
effector molecules and transcription factors directs one of skill
to the mode of action of 191P4D12(b), and thus identifies
therapeutic, prognostic, preventative and/or diagnostic targets for
cancer. This and similar assays are also used to identify and
screen for small molecules that interact with 191P4D12(b). Thus it
is found that 191P4D12(b) associates with proteins and small
molecules. Accordingly, 191P4D12(b) and these proteins and small
molecules are used for diagnostic, prognostic, preventative and/or
therapeutic purposes.
Example 50
Involvement of 191P4D12(b) in Cell-Cell Communication
[0999] Cell-cell communication is essential in maintaining organ
integrity and homeostasis, both of which become deregulated during
tumor formation and progression. Based on the presence of a
cadherin motif in 191P4D12(b), a motif known to be involved in cell
interaction and cell-cell adhesion, 191P4D12(b) can regulate cell
communication. Intercellular communications can be measured using
two types of assays (J. Biol. Chem. 2000, 275:25207). In the first
assay, cells loaded with a fluorescent dye are incubated in the
presence of unlabeled recipient cells and the cell populations are
examined under fluorescent microscopy. This qualitative assay
measures the exchange of dye between adjacent cells. In the second
assay system, donor and recipient cell populations are treated as
above and quantitative measurements of the recipient cell
population are performed by FACS analysis. Using these two assay
systems, cells expressing 191P4D12(b) are compared to controls that
do not express 191P4D12(b), and it is found that 191P4D12(b)
enhances cell communications. FIG. 19 and FIG. 20 demonstrate that
191P4D12(b) mediates the transfer of the small molecule calcein
between adjacent cells, and thereby regulates cell-cell
communication in prostate cancer cells. In this experiment,
recipient PC3 cells were labeled with dextran-Texas Red and donor
PC3 cells were labeled with calcein AM (green). The donor (green)
and recipient (red) cells were co-cultured at 37.degree. C. and
analyzed by microscopy for the co-localization of Texas red and
calcein. The results demonstrated that while PC3 control cells (no
detectable 191P4D12(b) protein expression) exhibit little calcein
transfer, the expression of 191P4D12(b) allows the transfer of
small molecules between cells (FIG. 19), whereby the initially red
recipient cells take on a brownish color, and co-localize the red
and green molecules. Small molecules and/or antibodies that
modulate cell-cell communication mediated by 191P4D12(b) are used
as therapeutics for cancers that express 191P4D12(b). When
191P4D12(b) functions in cell-cell communication and small molecule
transport, it is used as a target or marker for diagnostic,
prognostic, preventative and/or therapeutic purposes.
Example 51
Modulation of 191P4D12(b) Function
[1000] Knock Down of 191P4D12(b) Expression
[1001] Several techniques can be used to knock down or knock out
191P4D12(b) expression in vitro and in-vivo, including RNA
interference (RNAi) and other anti-sense technologies. RNAi makes
use of sequence specific double stranded RNA to prevent gene
expression. Small interfering RNA (siRNA) are transfected into
mammalian cells and thereby mediate sequence specific mRNA
degradation. (Elbashir, et al, Nature, 2001; vol. 411: 494-498).
Using this approach, 191P4D12(b)-specific RNAi is introduced in
191P4D12(b)-expressing cells by transfection. The effect of
knocking down the expression of 191P4D12(b) protein is evaluated
using the biological assays mentioned in examples 44 to 50
above.
[1002] Reduction of 191P4D12(b) Protein expression is detected
24-48 hours after transfection by immunostaining and flow
cytometry. The introduction of 191P4D12(b) specific RNAi reduced
the expression of 191P4D12(b) positive cells and reduce the
biological effect of 191P4D12(b) on tumor growth and
progression.
[1003] Accordingly, the RNA oligonucleotide sequences are used in
therapeutic and prophylactic applications. Moreover, the RNA
oligonucleotide sequences are used to assess how modulating the
expression of a 191P4D12(b) gene affects function of cancer cells
and/or tissues.
[1004] Inhibition Using Small Molecule and Antibodies
[1005] Using control cell lines and cell lines expressing
191P4D12(b), inhibitors of 191P4D12(b) function are identified. For
example, PC3 and PC3-191P4D12(b) cells can be incubated in the
presence and absence of mAb or small molecule inhibitors. The
effect of these mAb or small molecule inhibitors are investigated
using the cell communication, proliferation and signaling assays
described above.
[1006] Signal transduction and biological output mediated by
cadherins can be modulated through various mechanisms, including
inhibition of receptor binding, prevention of protein interactions,
or affecting the expression of co-receptors and binding partners
(Kamei et al, Oncogene 1999, 18:6776). Using control cell lines and
cell lines expressing 191P4D12(b), modulators (inhibitors or
enhancers) of 191P4D12(b) function are identified. For example, PC3
and PC3-191P4D12(b) cells are incubated in the presence and absence
of mAb or small molecule modulators. When mAb and small molecules
modulate, e.g., inhibit, the transport and tumorigenic function of
191P4D12(b), they are used for preventative, prognostic, diagnostic
and/or therapeutic purposes.
[1007] Throughout this application, various website data content,
publications, patent applications and patents are referenced.
(Websites are referenced by their Uniform Resource Locator, or URL,
addresses on the World Wide Web.) The disclosures of each of these
references are hereby incorporated by reference herein in their
entireties.
[1008] The present invention is not to be limited in scope by the
embodiments disclosed herein, which are intended as single
illustrations of individual aspects of the invention, and any that
are functionally equivalent are within the scope of the invention.
Various modifications to the models and methods of the invention,
in addition to those described herein, will become apparent to
those skilled in the art from the foregoing description and
teachings, and are similarly intended to fall within the scope of
the invention. Such modifications or other embodiments can be
practiced without departing from the true scope and spirit of the
invention.
TABLES
[1009]
5TABLE I Tissues that Express 191P4D12(b): a Malignant Tissues
Prostate Bladder Kidney Colon Lung Pancreas Ovary Breast Uterus
Cervix
[1010]
6TABLE II Amino Acid Abbreviations SINGLE THREE FULL LETTER LETTER
NAME F Phe phenylalanine L Leu leucine S Ser serine Y Tyr tyrosine
C Cys cysteine W Trp tryptophan P Pro proline H His histidine Q Gln
glutamine R Arg arginine I Ile isoleucine M Met methionine T Thr
threonine N Asn asparagine K Lys lysine V Val valine A Ala alanine
D Asp aspartic acid E Glu glutamic acid G Gly glycine
[1011]
7TABLE III Amino Acid Substitution Matrix A C D E F G H I K L M N P
Q R S T V W Y . 4 0 -2 -1 -2 0 -2 -1 -1 -1 -1 -2 -1 -1 -1 1 0 0 -3
-2 A 9 -3 -4 -2 -3 -3 -1 -3 -1 -1 -3 -3 -3 -3 -1 -1 -1 -2 -2 C 6 2
-3 -1 -1 -3 -1 -4 -3 1 -1 0 -2 0 -1 -3 -4 -3 D 5 -3 -2 0 -3 1 -3 -2
0 -1 2 0 0 -1 -2 -3 -2 E 6 -3 -1 0 -3 0 0 -3 -4 -3 -3 -2 -2 -1 1 3
F 6 -2 -4 -2 -4 -3 0 -2 -2 -2 0 -2 -3 -2 -3 G 8 -3 -1 -3 -2 1 -2 0
0 -1 -2 -3 -2 2 H 4 -3 2 1 -3 -3 -3 -3 -2 -1 3 -3 -1 I 5 -2 -1 0 -1
1 2 0 -1 -2 -3 -2 K 4 2 -3 -3 -2 -2 -2 -1 1 -2 -1 L 5 -2 -2 0 -1 -1
-1 1 -1 -1 M 6 -2 0 0 1 0 -3 -4 -2 N 7 -1 -2 -1 -1 -2 -4 -3 P 5 1 0
-1 -2 -2 -1 Q 5 -1 -1 -3 -3 -2 R 4 1 -2 -3 -2 S 5 0 -2 -2 T 4 -3 -1
V 11 2 W 7 Y Adapted from the GCG Software 9.0 BLOSUM62 amino acid
substitution matrix (block substitution matrix). The higher the
value, the more likely a substitution is found in related, natural
proteins. (See world wide web URL
ikp.unibe.ch/manual/blosum62.html)
Table IV
HLA Class I/II Motifs/Supermotifs
[1012]
8TABLE IV (A) HLA Class I Supermotifs/Motifs POSITION POSITION
POSITION C Terminus 2 (Primary 3 (Primary (Primary Anchor) Anchor)
Anchor) SUPERMOTIF A1 TILVMS FWY A2 LIVMATQ IVMATL A3 VSMATLI RK
A24 YFWIVLMT FIYWLM B7 P VILFMWYA B27 RHK FYLWMIVA B44 ED FWYLIMVA
B58 ATS FWYLIVMA B62 QLIVMP FWYMIVLA MOTIFS A1 TSM Y A1 DEAS Y A2 1
LMVQIAT VLIMAT A3 LMVISATFCGD KYRHFA A11 VTMLISAGNCDF KRYH A24 YFWM
FLIW A*3101 MVTALIS RK A*3301 MVALFIST RK A*6801 AVTMSLI RK B*0702
P LMFWYAIV B*3501 P LMFWYIVA B51 P LIVFWYAM B*5301 P IMFWYALV
B*5401 P ATIVLMFWY Bolded residues are preferred, italicized
residues are less preferred A peptide is considered motif-bearing
if it has primary anchors at each primary anchor position for a
motif or supermotif as specified in the above table
[1013]
9TABLE IV (B) HLA Class II Supermotif 1 6 9 W, F, Y, V, I, L A, V,
I, L, P, C, S, T A, V, I, L, C, S, T, M, Y
[1014]
10TABLE IV (C) HLA Class II Motifs MOTIFS 1.degree. anchor 1 2 3 4
5 1.degree. anchor 6 7 8 9 DR4 preferred FMYLIVW M T I VSTCPALIM MH
MH deleterious W R WDE DR1 preferred MFLIVWY PAMQ VMATSPLIC M AVM
deleterious C CH FD CWD GDE D DR7 preferred MFLIVWY M W A IVMSACTPL
M IV deleterious C G GRD N G DR3 MOTIFS 1.degree. anchor 1 2 3
1.degree. anchor 4 5 1.degree. anchor 6 Motif a preferred LIVMFY D
Motif b preferred LIVMFAY DNQEST KRH DR Supermotif MFLIVWY
VMSTACPLI Italicized residues indicate less preferred or
"tolerated" residues
[1015]
11TABLE IV (D) HLA Class I Supermotifs POSITION: SUPER- MOTIFS 1 2
3 4 5 6 7 8 C-terminus A1 1.degree. Anchor 1.degree. Anchor TILVMS
FWY A2 1.degree. Anchor 1.degree. Anchor {overscore (LIVMATQ)}
LIVMAT A3 Preferred 1.degree. Anchor YFW YFW YFW P 1.degree. Anchor
{overscore (VSMATLI)} (4/5) (3/5) (4/5) (4/5) RK deleterious DE
(3/5); DE P (5/5) (4/5) A24 1.degree. Anchor 1.degree. Anchor
{overscore (YFWIVLMT)} FIYWLM B7 Preferred FWY (5/5) 1.degree.
Anchor FWY FWY 1.degree. Anchor LIVM (3/5) P (4/5) (3/5) {overscore
(VILFMWYA)} deleterious DE (3/5); DE G QN DE P(5/5); (3/5) (4/5)
(4/5) (4/5) G(4/5); A(3/5); QN(3/5) B27 1.degree. Anchor 1.degree.
Anchor RHK {overscore (FYLWMIVA)} B44 1.degree. Anchor 1.degree.
Anchor ED {overscore (FWYLIMVA)} B58 1.degree. Anchor 1.degree.
Anchor ATS {overscore (FWYLIVMA)} B62 1.degree. Anchor 1.degree.
Anchor QLIVMP {overscore (FWYMIVLA)} Italicized residues indicate
less preferred or "tolerated" residues
[1016]
12TABLE IV (E) HLA Class I Motifs POSITION 1 2 3 4 5 6 A1 preferred
GFYW 1.degree. Anchor DEA YFW P 9-mer STM deleterious DE RHKLIVMP A
G A A1 preferred GRHK ASTCLIVM 1.degree. Anchor GSTC ASTC 9-mer
DEAS deleterious A RHKDEPYFW DE PQN RHK A1 preferred YFW 1.degree.
Anchor DEAQN A YFWQN 10-mer STM deleterious GP RHKGLIVM DE RHK QNA
A1 preferred YFW STCLIVM 1.degree. Anchor A YFW 10-mer DEAS
deleterious RHK RHKDEPYFW P G A2.1 preferred YFW 1.degree. Anchor
YFW STC YFW 9-mer {overscore (LMIVQAT)} deleterious DEP DERKH RKH
POSITION 9 or 7 8 C-terminus C-terminus A1 preferred DEQN YFW
1.degree. Anchor 9-mer Y deleterious A1 preferred LIVM DE 1.degree.
Anchor 9-mer Y deleterious PG GP A1 preferred PASTC GDE P 1.degree.
Anchor 10-mer Y deleterious RHKYFW RHK A A1 preferred PG G YFW
1.degree. Anchor 10-mer Y deleterious PRHK QN A2.1 preferred A P
1.degree. Anchor 9-mer VLIMAT deleterious DERKH POSITION: 1 2 3 4 5
6 A2.1 preferred AYFW 1.degree. Anchor LVIM G G 10-mer {overscore
(LMIVQAT)} deleterious DEP DE RKHA P A3 preferred RHK 1.degree.
Anchor YFW PRHKYF A YFW {overscore (LMVISATFCGD)} W deleterious DEP
DE A11 preferred A 1.degree. Anchor YFW YFW A YFW {overscore
(VTLMISAGNCDF)} deleterious DEP A24 preferred YFWRHK 1.degree.
Anchor STC 9-mer YFWM deleterious DEG DE G QNP DERHK A24 Preferred
1.degree. Anchor P YFWP 10-mer YFWM Deleterious GDE QN RHK DE A3101
Preferred RHK 1.degree. Anchor YFW P YFW {overscore (MVTALIS)}
Deleterious DEP DE ADE DE A3301 Preferred 1.degree. Anchor YFW
{overscore (MVALFIST)} Deleterious GP DE A6801 Preferred YFWSTC
1.degree. Anchor YFWLIV {overscore (AVTMSLI)} M deleterious GP DEG
RHK B0702 Preferred RHKFWY 1.degree. Anchor RHK RHK RHK P
deleterious DEQNP DEP DE DE GDE B3501 Preferred FWYLIVM 1.degree.
Anchor FWY P POSITION: C- 7 8 9 Terminus A2.1 preferred FYWL
1.degree. Anchor 10-mer VIM VLIMAT deleterious RKH DERKH RKH A3
preferred P 1.degree. Anchor {overscore (KYRHFA)} deleterious A11
preferred YFW P 1.degree. Anchor KRYH deleterious A G A24 preferred
YFW YFW 1.degree. Anchor 9-mer FLIW deleterious G AQN A24 Preferred
P 1.degree. Anchor 10-mer FLIW Deleterious A QN DEA A3101 Preferred
YFW AP 1.degree. Anchor RK Deleterious DE DE A3301 Preferred AYFW
1.degree. Anchor RK Deleterious A6801 Preferred YFW P 1.degree.
Anchor RK deleterious A B0702 Preferred RHK PA 1.degree. Anchor
{overscore (LMFWYAIV)} deleterious QN DE B3501 Preferred FWY
1.degree. Anchor {overscore (LMFWYIVA)} POSITION 1 2 3 4 5 6 A1
preferred GFYW 1.degree. Anchor DEA YFW P 9-mer STM deleterious DE
RHKLIVMP A G A A1 preferred GRHK ASTCLIVM 1.degree. Anchor GSTC
ASTC 9-mer DEAS deleterious A RHKDEPYFW DE PQN RHK deleterious AGP
G G B51 Preferred LIVMFWY 1.degree. Anchor FWY STC FWY P
deleterious AGPDER DE G HKSTC B5301 preferred LIVMFWY 1.degree.
Anchor FWY STC FWY P deleterious AGPQN G B5401 preferred FWY
1.degree. Anchor FWYLIVM LIVM P deleterious GPQNDE GDESTC RHKDE DE
POSITION 9 or 7 8 C-terminus C-terminus A1 preferred DEQN YFW
1.degree. Anchor 9-mer Y deleterious A1 preferred LIVM DE 1.degree.
Anchor 9-mer Y deleterious PG GP deleterious B51 Preferred G FWY
1.degree. Anchor {overscore (LIVFWYAM)} deleterious DEQN GDE B5301
preferred LIVMFWY FWY 1.degree. Anchor {overscore (IMFWYALV)}
deleterious RHKQN DE B5401 preferred ALIVM FWYA 1.degree. Anchor P
{overscore (ATIVLMFWY)} deleterious QNDGE DE
[1017]
13TABLE IV (F) Summary of HLA-supertypes Overall phenotypic
frequencies of HLA-supertypes in different ethnic populations
Specificity Phenotypic frequency Supertype Position 2 C-Terminus
Caucasian N.A. Black Japanese Chinese Hispanic Average B7 P
AILMVFWY 43.2 55.1 57.1 43.0 49.3 49.5 A3 AILMVST RK 37.5 42.1 45.8
52.7 43.1 44.2 A2 AILMVT AILMVT 45.8 39.0 42.4 45.9 43.0 42.2 A24
YF (WIVLMT) FI (YWLM) 23.9 38.9 58.6 40.1 38.3 40.0 B44 E (D)
FWYLIMVA 43.0 21.2 42.9 39.1 39.0 37.0 A1 TI (LVMS) FWY 47.1 16.1
21.8 14.7 26.3 25.2 B27 RHK FYL (WMI) 28.4 26.1 13.3 13.9 35.3 23.4
B62 QL (IVMP) FWY (MIV) 12.6 4.8 36.5 25.4 11.1 18.1 B58 ATS FWY
(LIV) 10.0 25.1 1.6 9.0 5.9 10.3
[1018]
14TABLE IV (G) Calculated population coverage afforded by different
HLA-supertype combinations Phenotypic frequency HLA-supertypes
Caucasian N.A Blacks Japanese Chinese Hispanic Average A2, A3 and
B7 83.0 86.1 87.5 88.4 86.3 86.2 A2, A3, B7, A24, 99.5 98.1 100.0
99.5 99.4 99.3 B44 and A1 99.9 99.6 100.0 99.8 99.9 99.8 A2, A3,
B7, A24, B44, A1, B27, B62, and B58 Motifs indicate the residues
defining supertype specificites. The motifs incorporate residues
determined on the basis of published data to be recognized by
multiple alleles within the supertype. Residues within brackets are
additional residues also predicted to be tolerated by multiple
alleles within the supertype.
[1019]
15TABLE V Frequently Occurring Motifs avrg. % Name identity
Description Potential Function zf-C2H2 34% Zinc finger, C2H2 type
Nucleic acid-binding protein functions as transcription factor,
nuclear location probable cytochrome_b_N 68% Cytochrome b(N-
membrane bound oxidase, generate terminal)/b6/petB superoxide lg
19% Immunoglobulin domain domains are one hundred amino acids long
and include a conserved intradomain disulfide bond. WD40 18% WD
domain, G-beta repeat tandem repeats of about 40 residues, each
containing a Trp-Asp motif. Function in signal transduction and
protein interaction PDZ 23% PDZ domain may function in targeting
signaling molecules to sub-membranous sites LRR 28% Leucine Rich
Repeat short sequence motifs involved in protein-protein
interactions Pkinase 23% Protein kinase domain conserved catalytic
core common to both serine/threonine and tyrosine protein kinases
containing an ATP bindmg site and a catalytic site PH 16% PH domain
pleckstrin homology involved in intracellular signaling or as
constituents of the cytoskeleton EGF 34% EGF-like domain 30-40
amino acid long found in the extracellular domain of membrane-
bound proteins or in secreted proteins Rvt 49% Reverse
transcriptase (RNA-dependent DNA polymerase) Ank 25% Ank repeat
Cytoplasmic protein, associates integral membrane proteins to the
cytoskeleton Oxidored_q1 32% NADH- membrane associated Involved in
Ubiquinone/plastoquinone proton translation across the (complex I),
various chains membrane Efhand 24% EF hand calcium-binding domain,
consists of a12 residue loop flanked on both sides by a 12 residue
alpha-helical domain Rvp 79% Retroviral aspartyl Aspartyl or acid
proteases, centered on protease a catalytic aspartyl residue
Collagen 42% Collagen triple helix repeat extracellular structural
proteins involved (20 copies) in formation of connective tissue The
sequence consists of the G-X-Y and the polypeptide chains forms a
triple helix Fn3 20% Fibronectin type III domain Located in the
extracellular ligand- binding region of receptors and is about 200
amino acid residues long with two pairs of cysteines involved in
disulfide bonds 7tm_1 19% 7 transmembrane receptor seven
hydrophobic transmembrane (rhodopsin family) regions, with the
N-terminus located extracellularly while the C-terminus is
cytoplasmic Signal through G proteins
Table VI
Motifs and Post-Translational Modifications of 191P4D12(b)
[1020]
16TABLE VI Post-translational modifications of 191P4D12(b)
N-glycosylation site 1 281-284 NWTR (SEQ ID NO: 61) 430-433 NSSC
(SEQ ID NO: 62) 489-492 NGTL (SEQ ID NO: 63) Tyrosine sulfation
site 118-132 VQADEGEYECRVSTF (SEQ ID NO: 64) Protein kinase C
phosphorylation site 26-28 TGR 192-194 SSR 195-197 SFK 249-251 SVR
322-324 SSR 339-341 SGK 383-385 TQK 397-399 SIR 426-428 SLK 450-452
TVR 465-467 SGR 491-493 TLR Casein kinase II phosphorylation site
283-286 TRLD (SEQ ID NO: 65) 322-325 SSRD (SEQ ID NO: 66) 410-413
SQPE (SEQ ID NO: 67) 426-429 SLKD (SEQ ID NO: 68) 450-453 TVRE (SEQ
ID NO: 69) 456-459 TQTE (SEQ ID NO: 70) N-myristoylation site
135-140 GSFQAR (SEQ ID NO: 71) 162-167 GQGLTL (SEQ ID NO: 72)
164-169 GLTLAA (SEQ ID NO: 73) 189-194 GTTSSR (SEQ ID NO: 74)
218-223 GQPLTC (SEQ ID NO: 75) 311-316 GIYVCH (SEQ ID NO: 76)
354-359 GVIAAL (SEQ ID NO: 77) 464-469 GSGRAE (SEQ ID NO: 78)
477-482 GIKQAM (SEQ ID NO: 79) 490-495 GTLRAK (SEQ ID NO: 80)
500-505 GIYING (SEQ ID NO: 81) RGD Cell attachment sequence 55-57
RGD
[1021]
17TABLE VII Search Peptides 191P4D12(b) v.1 aa1-510 9-mers, 10-mers
and 15-mers MPLSLGAEMW GPEAWLLLLL LLASFTGRCP AGELETSDVV TVVLGQDAKL
PCFYRGDSGE (SEQ ID NO: 82) QVGQVAWARV DAGEGAQELA LLHSKYGLHV
SPAYEGRVEQ PPPPRN PLDG SVLLRNAVQA DEGEYECRVS TFPAGSFQAR LRLRVLVPPL
PSLNPGPALE EGQGLTLAAS CTAEGSPAPS VTWDTEVKGT TSSRSFKHSR SMVTSEFHL
VPSRSMNGQP LTCWSHPGL LQDQRITHIL HVSFLAEASV RGLEDQNLWH IGREGAMLKC
LSEGQPPPSY NWTRLDGPLP SGVRVDGDTL GFPPLTTEHS GIYVCHVSNE FSSRDSQVTV
DVLDPQEDSG KQVDLVSASV VVVGVIAALL FCLLWVVVL MSRYHRRKAQ QMTQKYEEEL
TLTRENSIRR LHSHHTDPRS QPEESVGLRA EGHPDSLKDN SSCSVMSEEP EGRSYSTLTT
VREIETQTEL LSPGSGRAEE EEDQDEGIKQ AMNHFVQENG TLRAKPTGNG IYINGRGHLV
v.2 aa1-510 9-mers 45-61 GQDAKLPCLYRGDSGEQ (SEQ ID NO: 83) 10-mers
44-62 LGQDAKLPCLYRGDSGEQV (SEQ ID NO: 84) 15-mers 39-67
VVTWLGQDAKLPCLYRGDSGEQVGQVAW (SEQ ID NO: 85) v.7 ORF: 264.1721
Frame + 3 9-mers 403-418 SHHTDPRSQSEEPEGR (SEQ ID NO: 86) 10-mers
402-419 HSHHTDPRSQSEEPEGRS (SEQ ID NO: 87) 15-mers 397-424
SIRRLHSHHTDPRSQSEEPEGRSYSTLT (SEQ ID NO: 88) V.9: AA 1-137; 9-mers,
10-mers, 15-mers MRRELLAGIL LRITFNFFLF FFLPFPLVVF FIYFYFYFEL
EMESHYVAQA GLELLGSSNP (SEQ ID NO: 89) PASASLVAGT LSVHHCACFE
SFTKRKKKLK KAFRFIQCLL LGLLKVRPLQ HQGVNSCDCE RGYFQGIFMQ AAPWEGT v.10
SNP variant 9-mers 27-43 GRCPAGELGTSDVVTVV (SEQ ID NO: 90) 10-mers
26-44 TGRCPAGELGTSDVVTVVL (SEQ ID NO: 91) 15-mers 21-49
LLASFTGRCPAGELGTSDVVTWLGQDAK (SEQ ID NO: 92) v.11 SNP variant
9-mers 138-154 QARLRLRVMVPPLPSLN (SEQ ID NO: 93) 10-mers 137-155
FQARLRLRVMVPPLPSLNP (SEQ ID NO: 94) 15-mers 132-160
FPAGSFQARLRLRVMVPPLPSLNPGPALE (SEQ ID NO: 95) v.12 SNP variant
9-mers 435-451 VMSEEPEGCSYSTLTTV (SEQ ID NO: 96) 10-mers 434-452
SVMSEEPEGGSYSTLTTVRE (SEQ ID NO: 97) 15-mers 429-457
DNSSCSVMSEEPEGCSYSTLTTVREIETQ (SEQ ID NO: 98) v.13 insertion of one
AA at 333-4 9-mers 426-442 SQVTVDVLADPQEDSGK (SEQ ID NO: 99)
10-mers 425-443 DSQVTVDVLADPQEDSGKQ (SEQ ID NO: 100) 15-mers
420-448 EFSSRDSQVTVDVLADPQEDSGKQVDLVS (SEQ ID NO: 101) 191 P4012(b)
v.14: AA56-72; 9-mers GSSNPPASASLVAGTLS (SEQ ID NO: 102)
191P4012(b)v.14: AA55-73; 10-mers LGSSNPPASASLVAGTLSV (SEQ ID NO:
103) 191P4D12(b) v.14: AA50.78; 15-mers
AGLELLGSSNPPASASLVAGTLSVHHCAC (SEQ ID NO: 104)
[1022]
18TABLE VIII Start Subsequence Score V1-HLA-A1-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 294
RVDGDTLGF 25.000 437 SEEPEGRSY 22.500 97 RVEQPPPPR 18.000 306
TTEHSGIYV 11.250 332 VLDPQEDSG 5.000 252 GLEDQNLWH 4.500 457
QTELLSPGS 4.500 271 LSEGQPPPS 2.700 205 TSEFHLVPS 2.700 107
PLDGSVLLR 2.500 386 YEEELTLTR 2.250 411 QPEESVGLR 2.250 184
DTEVKGTTS 2.250 172 TAEGSPAPS 1.800 6 GAEMWGPEA 1.800 33 ELETSDVVT
1.800 36 TSDVVTVVL 1.500 45 GQDAKLPCF 1.500 436 MSEEPEGRS 1.350 305
LTTEHSGIY 1.250 405 HTDPRSQPE 1.250 11 GPEAWLLLL 1.125 119
QADEGEYEC 1.000 89 HVSPAYEGR 1.000 284 RLDGPLPSG 1.000 342
QVDLVSASV 1.000 158 ALEEGQGLT 0.900 245 LAEASVRGL 0.900 419
RAEGHPDSL 0.900 453 EIETQTELL 0.900 486 VQENGTLRA 0.675 76
AQELALLHS 0.675 117 AVQADEGEY 0.500 471 EEDQDEGIK 0.500 236
ITHILHVSF 0.500 365 VVVVVLMSR 0.500 366 VVVVLMSRY 0.500 189
GTTSSRSFK 0.500 78 ELALLHSKY 0.500 69 RVDAGEGAQ 0.500 378 KAQQMTQKY
0.500 124 EYECRVSTF 0.450 120 ADEGEYECR 0.450 439 EPEGRSYST 0.450
130 STFPAGSFQ 0.250 86 YGLHVSPAY 0.250 318 SNEFSSRDS 0.225 72
AGEGAQELA 0.225 122 EGEYECRVS 0.225 159 LEEGQGLTL 0.225 262
GREGAMLKC 0.225 58 SGEQVGQVA 0.225 31 AGELETSDV 0.225 145 VLVPPLPSL
0.200 180 SVTWDTEVK 0.200 368 WLMSRYHR 0.200 41 TVVLGQDAK 0.200 17
LLLLLLASF 0.200 409 RSQPEESVG 0.150 129 VSTFPAGSF 0.150 200
RSAAVTSEF 0.150 423 HPDSLKDNS 0.125 392 LTRENSIRR 0.125 448
LTTVREIET 0.125 55 RGDSGEQVG 0.125 190 TTSSRSFKH 0.125 353
VGVIAALLF 0.125 146 LVPPLPSLN 0.100 369 VLMSRYHRR 0.100 313
YVCHVSNEF 0.100 61 QVGQVAWAR 0.100 459 ELLSPGSGR 0.100 329
TVDVLDPQE 0.100 20 LLLASFTGR 0.100 316 HVSNEFSSR 0.100 209
HLVPSRSMN 0.100 460 LLSPGSGRA 0.100 485 FVQENGTLR 0.100 467
RAEEEEDQD 0.090 3 LSLGAEMWG 0.075 225 VSHPGLLQD 0.075 255 DQNLWHIGR
0.075 135 GSFQARLRL 0.075 231 LQDQRITHI 0.075 473 DQDEGIKQA 0.075
296 DGDTLGFPP 0.062 364 LVVVVVLMS 0.050 354 GVIAALLFC 0.050 224
VVSHPGLLQ 0.050 202 AAVTSEFHL 0.050 210 LVPSRSMNG 0.050 19
LLLLASFTG 0.050 355 VIAALLFCL 0.050 299 TLGFPPLTT 0.050 15
WLLLLLLLA 0.050 298 DTLGFPPLT 0.050 287 GPLPSGVRV 0.050 28
RCPAGELET 0.050 435 VMSEEPEGR 0.050 357 AALLFCLLV 0.050
V2-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
5; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 1 GQDAKLPCL 0.150 3 DAKLPCLYR 0.050 4
AKLPCLYRG 0.010 2 QDAKLPCLY 0.003 6 LPCLYRGDS 0.003 7 PCLYRGDSG
0.001 5 KLPCLYRGD 0.001 8 CLYRGDSGE 0.000 9 LYRGDSGEQ 0.000
V7-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
15; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 3 HTDPRSQSE 1.250 7 RSQSEEPEG 0.030 8
SQSEEPEGR 0.015 1 SHHTDPRSQ 0.001 2 HHTDPRSQS 0.001 5 DPRSQSEEP
0.000 4 TDPRSQSEE 0.000 6 PRSQSEEPE 0.000 V9-HLA-A1-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 19; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 116
SCDCERGYF 5.000 13 ITFNFFLFF 1.250 76 CACFESFTK 1.000 27 LVVFFIYFY
1.000 97 QCLLLGLLK 1.000 39 FLEMESHYV 0.900 41 EMESHYVAQ 0.900 78
CFESFTKRK 0.900 51 GLELLGSSN 0.900 115 NSCDCERGY 0.750 25 FPLVVFFIY
0.625 23 LPFPLVVFF 0.500 4 ELLAGILLR 0.500 12 RITFNFFLF 0.500 28
VVFFIYFYF 0.500 118 DCERGYFQG 0.450 71 LSVHHCACF 0.300 80 ESFTKRKKK
0.300 22 FLPFPLVVF 0.200 31 FIYFYFYFF 0.200 57 SSNPPASAS 0.150 7
AGILLRITF 0.125 99 LLLGLLKVR 0.100 113 GVNSCDCER 0.100 77 ACFESFTKR
0.100 95 FIQCLLLGL 0.050 9 ILLRITFNF 0.050 98 CLLLGLLKV 0.050 5
LLAGILLRI 0.050 26 PLVVFFIYF 0.050 46 YVAQAGLEL 0.050 49 QAGLELLGS
0.050 29 VFFIYFYFY 0.050 58 SNPPASASL 0.050 65 SLVAGTLSV 0.050 2
RRELLAGIL 0.045 56 GSSNPPASA 0.030 62 ASASLVAGT 0.030 14 TFNFFLFFF
0.025 69 GTLSVHHCA 0.025 30 FFIYFYFYF 0.025 21 FFLPFPLVV 0.025 17
FFLFFFLPF 0.025 38 FFLEMESHY 0.025 67 VAGTLSVHH 0.020 126 GIFMQAAPW
0.020 54 LLGSSNPPA 0.020 43 ESHYVAQAG 0.015 64 ASLVAGTLS 0.015 15
FNFFLFFFL 0.013 121 RGYFQGIFM 0.013 79 FESFTKRKK 0.010 70 TLSVHHCAC
0.010 105 KVRPLQHQG 0.010 66 LVAGTLSVH 0.010 63 SASLVAGTL 0.010 6
LAGILLRIT 0.010 47 VAQAGLELL 0.010 10 LLRITFNFF 0.010 75 HCACFESFT
0.010 8 GILLRITFN 0.010 48 AQAGLELLG 0.007 103 LLKVRPLQH 0.005 128
FMQAAPWEG 0.005 55 LGSSNPPAS 0.005 120 ERGYFQGIF 0.005 74 HHCACFESF
0.005 82 FTKRKKKLK 0.005 87 KKLKKAFRF 0.003 90 KKAFRFIQC 0.003 11
LRITFNFFL 0.003 59 NPPASASLV 0.003 101 LGLLKVRPL 0.003 123
YFQGIFMQA 0.003 36 FYFFLEMES 0.003 34 FYFYFFLEM 0.003 19 LFFFLPFPL
0.003 68 AGTLSVHHC 0.003 93 FRFIQCLLL 0.003 114 VNSCDCERG 0.003 122
GYFQGIFMQ 0.003 50 AGLELLGSS 0.003 32 IYFYFYFFL 0.003 3 RELLAGILL
0.003 107 RPLQHQGVN 0.003 73 VHHCACFES 0.003 94 RFIQCLLLG 0.003 18
FLFFFLPFP 0.002 102 GLLKVRPLQ 0.002 100 LLGLLKVRP 0.002 108
PLQHQGVNS 0.002 61 PASASLVAG 0.002 96 IQCLLLGLL 0.002 111 HQGVNSCDC
0.002 109 LQHQGVNSC 0.002 124 FQGIFMQAA 0.002 129 MQAAPWEGT 0.002
60 PPASASLVA 0.001 86 KKKLKKAFR 0.001 20 FFFLPFPLV 0.001 V10
HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO: 21;
each start position is specified the length of peptide is 9 ammo
acids, and the end position for each peptide is the start position
plus eight 5 AGELGTSDV 0.225 2 RCPAGELGT 0.050 9 GTSDVVTVV 0.025 7
ELGTSDVVT 0.020 1 GRCPAGELG 0.005 8 LGTSDVVTV 0.005 3 CPAGELGTS
0.003 6 GELGTSDVV 0.001 4 PAGELGTSD 0.000
V11-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
23; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 9 MVPPLPSLN 0.100 8 VMVPPLPSL 0.100 7
RVMVPPLPS 0.050 5 RLRVMVPPL 0.002 1 QARLRLRVM 0.001 3 RLRLRVMVP
0.001 6 LRVMVPPLP 0.000 2 ARLRLRVMV 0.000 4 LRLRVMVPP 0.000
V12-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 3 SEEPEGCSY 22.500 2 MSEEPEGCS 1.350 5
EPEGCSYST 0.450 8 GCSYSTLTT 0.050 9 CSYSTLTTV 0.015 1 VMSEEPEGC
0.005 7 EGCSYSTLT 0.003 4 EEPEGCSYS 0.001 6 PEGCSYSTL 0.000
V13-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
27; each start position is specified, the length peptide is 9 amino
acids, and the end position for each peptide is the start position
plus eight. 8 LADPQEDSG 5.000 4 TVDVLADPQ 0.500 9 ADPQEDSGK 0.010 7
VLADPQEDS 0.010 3 VTVDVLADP 0.005 2 QVTVDVLAD 0.005 1 SQVTVDVLA
0.003 6 DVLADPQED 0.001 5 VDVLADPQE 0.000
V14-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
29; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 2 SSNPPASAS 0.150 3 SNPPASASL 0.050 1
GSSNPPASA 0.030 7 ASASLVAGT 0.030 9 ASLVAGTLS 0.015 8 SASLVAGTL
0.010 4 NPPASASLV 0.003 6 PASASLVAG 0.002 5 PPASASLVA 0.001
[1023]
19TABLE IX Start Subsequence Score V1-HLA-A1-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 271
LSEGQPPPSY 135.000 332 VLDPQEDSGK 100.000 436 MSEEPEGRSY 67.500 205
TSEFHLVPSR 27.000 419 RAEGHPDSLK 18.000 119 QADEGEYECR 5.000 453
EIETQTELLS 4.500 306 TTEHSGIYVC 4.500 158 ALEEGQGLTL 4.500 45
GQDAKLPCFY 3.750 486 VQENGTLRAK 2.700 76 AQELALLHSK 2.700 405
HTDPRSQPEE 2.500 385 KYEEELTLTR 2.250 457 QTELLSPGSG 2.250 184
DTEVKGTTSS 2.250 33 ELETSDVVTV 1.800 97 RVEQPPPPRN 1.800 172
TAEGSPAPSV 1.800 36 TSDVVTVVLG 1.500 130 STFPAGSFQA 1.250 411
QPEESVGLRA 1.125 11 GPEAWLLLLL 1.125 72 AGEGAQELAL 1.125 470
EEEDQDEGIK 0.900 252 GLEDQNLWHI 0.900 6 GAEMWGPEAW 0.900 116
NAVQADEGEY 0.500 40 VTVVLGQDAK 0.500 493 RAKPTGNGIY 0.500 365
VVVVVLMSRY 0.500 352 VVGVIAALLF 0.500 342 QVDLVSASVV 0.500 209
HLVPSRSMNG 0.500 364 LVVVVVLMSR 0.500 284 RLDGPLPSGV 0.500 122
EGEYECRVST 0.450 437 SEEPEGRSYS 0.450 58 SGEQVGQVAW 0.450 409
RSQPEESVGL 0.300 296 DGDTLGFPPL 0.250 107 PLDGSVLLRN 0.250 390
LTLTRENSIR 0.250 275 QPPPSYNWTR 0.250 55 RGDSGEQVGQ 0.250 318
SNEFSSRDSQ 0.225 31 AGELETSDVV 0.225 439 EPEGRSYSTL 0.225 235
RITHILHVSF 0.200 16 LLLLLLLASF 0.200 367 VVVLMSRYHR 0.200 369
VLMSRYHRRK 0.200 242 VSFLAEASVR 0.150 225 VSHPGLLQDQ 0.150 135
GSFQARLRLR 0.150 443 RSYSTLTTVR 0.150 298 DTLGFPPLTT 0.125 189
GTTSSRSFKH 0.125 423 HPDSLKDNSS 0.125 106 NPLDGSVLLR 0.125 305
LTTEHSGIYV 0.125 471 EEDQDEGIKQ 0.125 400 RLHSHHTDPR 0.100 69
RVDAGEGAQE 0.100 145 VLVPPLPSLN 0.100 434 SVMSEEPEGR 0.100 260
HIGREGAMLK 0.100 89 HVSPAYEGRV 0.100 368 VVLMSRYHRR 0.100 128
RVSTFPAGSF 0.100 19 LLLLASFTGR 0.100 474 QDEGIKQAMN 0.090 467
RAEEEEDQDE 0.090 245 LAEASVRGLE 0.090 473 DQDEGIKQAM 0.075 214
RSMNGQPLTC 0.075 231 LQDQRITHIL 0.075 357 AALLFCLLVV 0.050 43
VLGQDAKLPC 0.050 188 KGTTSSRSFK 0.050 44 LGQDAKLPCF 0.050 217
NGQPLTCVVS 0.050 201 SAAVTSEFHL 0.050 294 RVDGDTLGFP 0.050 18
LLLLLASFTG 0.050 35 ETSDVVTVVL 0.050 171 CTAEGSPAPS 0.050 447
TLTTVREIET 0.050 221 LTCVVSHPGL 0.050 354 GVIAALLFCL 0.050 81
LLHSKYGLHV 0.050 323 SRDSQVTVDV 0.050 329 TVDVLDPQED 0.050 304
PLTTEHSGIY 0.050 273 EGQPPPSYNW 0.050 15 WLLLLLLLAS 0.050 363
LLVVVVVLMS 0.050 85 KYGLHVSPAY 0.050 146 LVPPLPSLNP 0.050 485
FVQENGTLRA 0.050 V2-HLA-A1-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 2 GQDAKLPCLY 3.750 6
KLPCLYRGDS 0.010 1 LGQDAKLPCL 0.005 3 QDAKLPCLYR 0.003 7 LPCLYRGDSG
0.003 4 DAKLPCLYRG 0.002 9 CLYRGDSGEQ 0.001 5 AKLPCLYRGD 0.001 8
PCLYRGDSGE 0.000 10 LYRGDSGEQV 0.000 V7-HLA-A1-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 4
HTDPRSQSEE 1.250 8 RSQSEEPEGR 0.150 1 HSHHTDPRSQ 0.015 9 SQSEEPEGRS
0.002 2 SHHTDPRSQS 0.001 7 PRSQSEEPEG 0.000 3 HHTDPRSQSE 0.000 6
DPRSQSEEPE 0.000 5 TDPRSQSEEP 0.000 V9-HLA-A1-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 19; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 39
FLEMESHYVA 1.800 13 ITFNFFLFFF 1.250 28 VVFFIYFYFY 1.000 116
SCDCERGYFQ 1.000 75 HCACFESFTK 1.000 78 CFESFTKRKK 0.900 41
EMESHYVAQA 0.900 12 RITFNFFLFF 0.500 27 LVVFFIYFYF 0.500 8
GILLRITFNF 0.500 6 LAGILLRITF 0.500 57 SSNPPASASL 0.300 2
RRELLAGILL 0.225 22 FLPFPLVVFF 0.200 70 TLSVHHCACF 0.200 77
ACFESFTKRK 0.200 96 IQCLLLGLLK 0.150 115 NSCDCERGYF 0.150 114
VNSCDCERGY 0.125 23 LPFPLVVFFI 0.125 25 FPLVVFFIYF 0.125 76
CACFESFTKR 0.100 26 PLVVFFIYFY 0.100 21 FFLPFPLVVF 0.100 98
CLLLGLLKVR 0.100 118 DCERGYFQGI 0.090 51 GLELLGSSNP 0.090 64
ASLVAGTLSV 0.075 31 FIYFYFYFFL 0.050 47 VAQAGLELLG 0.050 72
SVHHCACFES 0.050 4 ELLAGILLRI 0.050 97 QCLLLGLLKV 0.050 18
FLFFFLPFPL 0.050 43 ESHYVAQAGL 0.030 58 SNPPASASLV 0.025 3
RELLAGILLR 0.025 112 QGVNSCDCER 0.025 69 GTLSVHHCAC 0.025 11
LRITFNFFLF 0.025 82 FTKRKKKLKK 0.025 29 VFFIYFYFYF 0.025 16
NFFLFFFLPF 0.025 37 YFFLEMESHY 0.025 66 LVAGTLSVHH 0.020 54
LLGSSNPPAS 0.020 53 ELLGSSNPPA 0.020 56 GSSNPPASAS 0.015 62
ASASLVAGTL 0.015 80 ESFTKRKKKL 0.015 24 PFPLVVFFIY 0.013 59
NPPASASLVA 0.013 121 RGYFQGIFMQ 0.013 67 VAGTLSVHHC 0.010 105
KVRPLQHQGV 0.010 9 ILLRITFNFF 0.010 79 FESFTKRKKK 0.010 49
QAGLELLGSS 0.010 46 YVAQAGLELL 0.010 63 SASLVAGTLS 0.010 113
GVNSCDCERG 0.010 95 FIQCLLLGLL 0.010 30 FFIYFYFYFF 0.010 5
LLAGILLRIT 0.010 65 SLVAGTLSVH 0.010 100 LLGLLKVRPL 0.010 48
AQAGLELLGS 0.007 102 GLLKVRPLQH 0.005 55 LGSSNPPASA 0.005 101
LGLLKVRPLQ 0.005 73 VHHCACFESF 0.005 125 QGIFMQAAPW 0.005 10
LLRITFNFFL 0.005 107 RPLQHQGVNS 0.005 128 FMQAAPWEGT 0.005 86
KKKLKKAFRF 0.003 117 CDCERGYFQG 0.003 93 FRFIQCLLLG 0.003 14
TFNFFLFFFL 0.003 33 YFYFYFFLEM 0.003 120 ERGYFQGIFM 0.003 122
GYFQGIFMQA 0.003 35 YFYFFLEMES 0.003 68 AGTLSVHHCA 0.003 45
HYVAQAGLEL 0.003 50 AGLELLGSSN 0.003 7 AGILLRITFN 0.003 20
FFFLPFPLVV 0.003 94 RFIQCLLLGL 0.003 126 GIFMQAAPWE 0.002 99
LLLGLLKVRP 0.002 61 PASASLVAGT 0.002 71 LSVHHCACFE 0.002 15
FNFFLFFFLP 0.001 81 SFTKRKKKLK 0.001 103 LLKVRPLQHQ 0.001 108
PLQHQGVNSC 0.001 40 LEMESHYVAQ 0.001 91 KAFRFIQCLL 0.001 19
LFFFLPFPLV 0.001 V10-HLA-A1-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 21; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 6 AGELGTSDVV 0.225 10
GTSDVVTVVL 0.050 2 GRCPAGELGT 0.025 8 ELGTSDVVTV 0.020 3 RCPAGELGTS
0.010 9 LGTSDVVTVV 0.003 7 GELGTSDVVT 0.001 5 PAGELGTSDV 0.001 4
CPAGELGTSD 0.000 1 TGRCPAGELG 0.000 V11-HLA-A1-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 23; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 9
VMVPPLPSLN 0.050 10 MVPPLPSLNP 0.050 8 RVMVPPLPSL 0.020 7
LRVMVPPLPS 0.003 2 QARLRLRVMV 0.002 6 RLRVMVPPLP 0.000 4 RLRLRVMVPP
0.000 1 FQARLRLRVM 0.000 5 LRLRVMVPPL 0.000 3 ARLRLRVMVP 0.000
V12-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 3 MSEEPEGCSY 67.500 4 SEEPEGCSYS 0.450 6
EPEGCSYSTL 0.225 10 CSYSTLTTVR 0.150 8 EGCSYSTLTT 0.013 9
GCSYSTLTTV 0.010 1 SVMSEEPEGC 0.010 2 VMSEEPEGCS 0.005 5 EEPEGCSYST
0.001 11 SYSTLTTVRE 0.000 7 PEGCSYSTLT 0.000
V13-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
27; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 9 LADPQEDSGK 100.000 5 TVDVLADPQE 0.100 1
DSQVTVDVLA 0.030 4 VTVDVLADPQ 0.025 8 VLADPQEDSG 0.010 7 DVLADPQEDS
0.010 3 QVTVDVLADP 0.002 2 SQVTVDVLAD 0.001 10 ADPQEDSGKQ 0.001 6
VDVLADPQED 0.000 V14-HLA-A1-10mers-191P4- D12B Each peptide is a
portion of SEQ ID NO: 29; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 3 SSNPPASASL 0.300 10
ASLVAGTLSV 0.075 4 SNPPASASLV 0.025 8 ASASLVAGTL 0.015 2 GSSNPPASAS
0.015 5 NPPASASLVA 0.013 9 SASLVAGTLS 0.010 1 LGSSNPPASA 0.005 7
PASASLVAGT 0.002 6 PPASASLVAG 0.001
[1024]
20TABLE X Start Subsequence Score V1-HLA-A201-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eiqht. 359
LLFCLLVVV 412.546 18 LLLLLASFT 257.802 358 ALLFCLLVV 242.674 15
WLLLLLLLA 194.477 145 VLVPPLPSL 83.527 80 ALLHSKYGL 79.041 362
CLLVVVVVL 74.536 355 VIAALLFCL 66.613 8 EMWGPEAWL 52.823 502
YINGRGHLV 43.992 137 FQARLRLRV 32.438 112 VLLRNAVQA 31.249 363
LLVVVVVLM 19.425 357 AALLFCLLV 13.582 42 VVLGQDAKL 11.757 203
AVTSEFHLV 11.563 345 LVSASVVVV 9.756 410 SQPEESVGL 8.880 299
TLGFPPLTT 7.452 164 GLTLAASCT 7.452 351 VVVGVIAAL 7.309 361
FCLLVVVVV 7.287 354 GVIAALLFC 5.499 34 LETSDVVTV 5.288 10 WGPEAWLLL
4.471 21 LLASFTGRC 4.172 32 GELETSDVV 4.122 142 RLRVLVPPL 3.734 215
SMNGQPLTC 3.588 443 RSYSTLTTV 3.342 352 VVGVIAALL 3.178 242
VSFLAEASV 2.856 19 LLLLASFTG 2.719 342 QVDLVSASV 2.434 253
LEDQNLWHI 2.380 229 GLLQDQRIT 2.261 347 SASVVVVGV 2.222 344
DLVSASVVV 2.139 106 NPLDGSVLL 2.115 123 GEYECRVST 1.901 216
MNGQPLTCV 1.775 202 AAVTSEFHL 1.721 452 REIETQTEL 1.703 350
VVVVGVIAA 1.700 287 GPLPSGVRV 1.680 231 LQDQRITHI 1.654 244
FLAEASVRG 1.405 173 AEGSPAPSV 1.352 62 VGQVAWARV 1.312 495
KPTGNGIYI 1.311 460 LLSPGSGRA 1.098 17 LLLLLLASF 1.078 16 LLLLLLLAS
1.078 356 IAALLFCLL 0.958 263 REGAMLKCL 0.955
V1-HLA-A201-9mers-191P4- D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 390 LTLTRENSI 0.911 478 IKQAMNHFV 0.903 230
LLQDQRITH 0.519 135 GSFQARLRL 0.516 238 HILHVSFLA 0.498 60
EQVGQVAWA 0.478 481 AMNHFVQEN 0.470 266 AMLKCLSEG 0.458 110
GSVLLRNAV 0.454 196 FKHSRSAAV 0.444 64 QVAWARVDA 0.435 165
LTLAASCTA 0.434 13 EAWLLLLLL 0.425 121 DEGEYECRV 0.416 73 GEGAQELAL
0.415 275 QPPPSYNWT 0.401 384 QKYEEELTL 0.389 306 TTEHSGIYV 0.340
35 ETSDVVTVV 0.280 4 SLGAEMWGP 0.257 158 ALEEGQGLT 0.254 341
KQVDLVSAS 0.249 343 VDLVSASVV 0.249 382 MTQKYEEEL 0.247 446
STLTTVREI 0.247 223 CVVSHPGLL 0.243 304 PLTTEHSGI 0.230 44
LGQDAKLPC 0.226 1 MPLSLGAEM 0.204 450 TVREIETQT 0.203 237 THILHVSFL
0.188 217 NGQPLTCVV 0.186 214 RSMNGQPLT 0.180 349 SVVVVGVIA 0.178
20 LLLASFTGR 0.178 448 LTTVREIET 0.176 285 LDGPLPSGV 0.164 473
DQDEGIKQA 0.142 322 SSRDSQVTV 0.141 369 VLMSRYHRR 0.141 100
QPPPPRNPL 0.139 222 TCVVSHPGL 0.139 257 NLWHIGREG 0.124 163
QGLTLAASC 0.120 23 ASFTGRCPA 0.120 V2-HLA-A201-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 5; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 1
GQDAKLPCL 1.993 8 CLYRGDSGE 0.048 5 KLPCLYRGD 0.016 4 AKLPCLYRG
0.001 6 LPCLYRGDS 0.000 2 QDAKLPCLY 0.000 7 PCLYRGDSG 0.000 3
DAKLPCLYR 0.000 9 LYRGDSGEQ 0.000 V7-HLA-A201-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 8
SQSEEPEGR 0.003 7 RSQSEEPEG 0.000 4 TDPRSQSEE 0.000 2 HHTDPRSQS
0.000 3 HTDPRSQSE 0.000 1 SHHTDPRSQ 0.000 5 DPRSQSEEP 0.000 6
PRSQSEEPE 0.000 V9-HLA-A201-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 19; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 98 CLLLGLLKV 591.888 15
FNFFLFFFL 143.853 39 FLEMESHYV 112.619 65 SLVAGTLSV 69.552 5
LLAGILLRI 40.792 91 KAFRFIQCL 33.581 95 FIQCLLLGL 31.077 124
FQGIFMQAA 20.251 18 FLFFFLPFP 12.194 46 YVAQAGLEL 8.598 54
LLGSSNPPA 8.446 70 TLSVHHCAC 4.968 32 IYFYFYFFL 3.393 9 ILLRITFNF
2.719 88 KLKKAFRFI 2.671 109 LQHQGVNSC 1.969 28 VVFFIYFYF 1.963 128
FMQAAPWEG 1.857 31 FIYFYFYFF 1.576 20 FFFLPFPLV 1.562 3 RELLAGILL
1.537 21 FFLPFPLVV 1.281 96 IQCLLLGLL 1.101 129 MQAAPWEGT 1.070 40
LEMESHYVA 1.021 11 LRITFNFFL 0.611 121 RGYFQGIFM 0.571 47 VAQAGLELL
0.568 19 LFFFLPFPL 0.541 27 LVVFFIYFY 0.533 8 GILLRITFN 0.480 59
NPPASASLV 0.454 101 LGLLKVRPL 0.403 42 MESHYVAQA 0.378 22 FLPFPLVVF
0.323 13 ITFNFFLFF 0.259 69 GTLSVHHCA 0.255 58 SNPPASASL 0.139 12
RITFNFFLF 0.113 62 ASASLVAGT 0.112 10 LLRITFNFF 0.101 99 LLLGLLKVR
0.088 34 FYFYFFLEM 0.085 68 AGTLSVHHC 0.075 26 PLVVFFIYF 0.065 102
GLLKVRPLQ 0.055 93 FRFIQCLLL 0.050 44 SHYVAQAGL 0.047 90 KKAFRFIQC
0.046 30 FFIYFYFYF 0.043 23 LPFPLVVFF 0.039 63 SASLVAGTL 0.039 126
GIFMQAAPW 0.038 25 FPLVVFFIY 0.037 75 HCACFESFT 0.035 6 LAGILLRIT
0.033 56 GSSNPPASA 0.032 123 YFQGIFMQA 0.030 119 CERGYFQGI 0.029
100 LLGLLKVRP 0.025 111 HQGVNSCDC 0.017 106 VRPLQHQGV 0.016 81
SFTKRKKKL 0.015 14 TFNFFLFFF 0.014 24 PFPLVVFFI 0.012 66 LVAGTLSVH
0.010 4 ELLAGILLR 0.010 87 KKLKKAFRF 0.008 48 AQAGLELLG 0.008 72
SVHHCACFE 0.007 17 FFLFFFLPF 0.006 51 GLELLGSSN 0.005 103 LLKVRPLQH
0.004 53 ELLGSSNPP 0.004 38 FFLEMESHY 0.004 29 VFFIYFYFY 0.003 77
ACFESFTKR 0.003 49 QAGLELLGS 0.002 50 AGLELLGSS 0.002 52 LELLGSSNP
0.002 64 ASLVAGTLS 0.002 1 MRRELLAGI 0.002 67 VAGTLSVHH 0.002 105
KVRPLQHQG 0.002 33 YFYFYFFLE 0.002 108 PLQHQGVNS 0.002 16 NFFLFFFLP
0.002 113 GVNSCDCER 0.001 76 CACFESFTK 0.001 92 AFRFIQCLL 0.001 37
YFFLEMESH 0.001 71 LSVHHCACF 0.001 55 LGSSNPPAS 0.001 35 YFYFFLEME
0.001 73 VHHCACFES 0.001 7 AGILLRITF 0.000 57 SSNPPASAS 0.000 117
CDCERGYFQ 0.000 114 VNSCDCERG 0.000 115 NSCDCERGY 0.000
V10-HLA-A201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 8 VMVPPLPSL 60.325 5 RLRVMVPPL 3.734 2
ARLRLRVMV 0.036 7 RVMVPPLPS 0.024 9 MVPPLPSLN 0.011 3 RLRLRVMVP
0.001 1 QARLRLRVM 0.001 4 LRLRVMVPP 0.000 6 LRVMVPPLP 0.000
V11-HLA-A201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 23; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 9 GTSDVVTVV 3.735 8 LGTSDVVTV 1.775 6
GELGTSDVV 1.005 7 ELGTSDVVT 0.229 2 RCPAGELGT 0.049 5 AGELGTSDV
0.029 3 CPAGELGTS 0.000 4 PAGELGTSD 0.000 1 GRCPAGELG 0.000
V12-HLA-A201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 25; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 1 VMSEEPEGC 12.254 9 CSYSTLTTV 3.342 8
GCSYSTLTT 0.049 6 PEGCSYSTL 0.014 7 EGCSYSTLT 0.004 4 EEPEGCSYS
0.002 5 EPEGCSYST 0.000 3 SEEPEGCSY 0.000 2 MSEEPEGCS 0.000
V13-HLA-A201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 1 SQVTVDVLA 0.504 7 VLADPQEDS 0.255 3
VTVDVLADP 0.003 2 QVTVDVLAD 0.003 6 DVLADPQED 0.000 8 LADPQEDSG
0.000 4 TVDVLADPQ 0.000 5 VDVLADPQE 0.000 9 ADPQEDSGK 0.000
V14-HLA-A201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 4 NPPASASLV 0.454 3 SNPPASASL 0.139 7
ASASLVAGT 0.112 8 SASLVAGTL 0.039 1 GSSNPPASA 0.032 9 ASLVAGTLS
0.002 2 SSNPPASAS 0.000 5 PPASASLVA 0.000 6 PASASLVAG 0.000
[1025]
21TABLE XI Start Subsequence Score V1-HLA-A201-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 359
LLFCLLVVVV 412.546 17 LLLLLLASFT 257.802 358 ALLFCLLVVV 242.674 244
FLAEASVRGL 185.332 230 LLQDQRITHI 167.248 81 LLHSKYGLHV 118.238 215
SMNGQPLTCV 115.534 341 KQVDLVSASV 101.193 239 ILHVSFLAEA 73.815 8
EMWGPEAWLL 72.031 252 GLEDQNLWHI 47.223 362 CLLVVVVVLM 42.278 305
LTTEHSGIYV 37.032 284 RLDGPLPSGV 27.821 354 GVIAALLFCL 24.935 257
NLWHIGREGA 20.205 144 RVLVPPLPSL 15.907 20 LLLASFTGRC 15.437 181
VTWDTEVKGT 13.771 61 QVGQVAWARV 10.346 426 SLKDNSSCSV 9.981 355
VIAALLFCLL 9.488 7 AEMWGPEAWL 8.453 43 VLGQDAKLPC 8.446 485
FVQENGTLRA 8.198 381 QMTQKYEEEL 7.560 447 TLTTVREIET 7.452 350
VVVVGVIAAL 7.309 236 ITHILHVSFL 6.381 356 IAALLFCLLV 6.240 274
GQPPPSYNWT 6.233 10 WGPEAWLLLL 6.049 158 ALEEGQGLTL 5.605 319
NEFSSRDSQV 5.004 164 GLTLAASCTA 4.968 344 DLVSASVVVV 4.919 118
VQADEGEYEC 3.511 357 AALLFCLLVV 3.370 351 VVVGVIAALL 3.178 15
WLLLLLLLAS 2.917 18 LLLLLASFTG 2.719 125 YECRVSTFPA 2.577 132
FPAGSFQARL 2.438 361 FCLLVVVVVL 2.238 34 LETSDVVTVV 2.168 321
FSSRDSQVTV 2.088 137 FQARLRLRVL 1.879 41 TVVLGQDAKL 1.869 79
LALLHSKYGL 1.866 477 GIKQAMNHFV 1.841 202 AAVTSEFHLV 1.835 346
VSASVVVVGV 1.775 201 SAAVTSEFHL 1.721 111 SVLLRNAVQA 1.608 130
STFPAGSFQA 1.481 59 GEQVGQVAWA 1.222 500 GIYINGRGHL 1.222 370
LMSRYHRRKA 1.220 16 LLLLLLLASF 1.078 349 SVVVVGVIAA 1.000 342
QVDLVSASVV 0.998 73 GEGAQELALL 0.955 32 GELETSDVVT 0.901 452
REIETQTELL 0.834 389 ELTLTRENSI 0.782 33 ELETSDVVTV 0.768 39
VVTVVLGQDA 0.739 353 VGVIAALLFC 0.697 280 YNWTRLDGPL 0.692 231
LQDQRITHIL 0.604 221 LTCVVSHPGL 0.504 63 GQVAWARVDA 0.504 162
GQGLTLAASC 0.504 178 APSVTWDTEV 0.454 13 EAWLLLLLLL 0.425 176
SPAPSVTWDT 0.365 216 MNGQPLTCVV 0.316 384 QKYEEELTLT 0.312 270
CLSEGQPPPS 0.306 363 LLVVVVVLMS 0.291 229 GLLQDQRITH 0.276 343
VDLVSASVVV 0.249 150 LPSLNPGPAL 0.237 5 LGAEMWGPEA 0.226 112
VLLRNAVQAD 0.216 241 HVSFLAEASV 0.207 163 QGLTLAASCT 0.180 459
ELLSPGSGRA 0.179 19 LLLLASFTGR 0.178 25 FTGRCPAGEL 0.177 336
QEDSGKQVDL 0.166 99 EQPPPPRNPL 0.162 445 YSTLTTVREI 0.144 249
SVRGLEDQNL 0.142 334 DPQEDSGKQV 0.140 105 RNPLDGSVLL 0.139 409
RSQPEESVGL 0.139 134 AGSFQARLRL 0.139 156 GPALEEGQGL 0.139 145
VLVPPLPSLN 0.127 V2-HLA-A201-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 1 LGQDAKLPCL 2.236 6
KLPCLYRGDS 0.034 9 CLYRGDSGEQ 0.006 2 GQDAKLPCLY 0.003 10
LYRGDSGEQV 0.001 7 LPCLYRGDSG 0.000 3 QDAKLPCLYR 0.000 5 AKLPCLYRGD
0.000 8 PCLYRGDSGE 0.000 4 DAKLPCLYRG 0.000
V7-HLA-A201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 9 SQSEEPEGRS 0.004 2 SHHTDPRSQS 0.000 8
RSQSEEPEGR 0.000 5 TDPRSQSEEP 0.000 4 HTDPRSQSEE 0.000 3 HHTDPRSQSE
0.000 1 HSHHTDPRSQ 0.000 6 DPRSQSEEPE 0.000 7 PRSQSEEPEG 0.000
V9-HLA-A201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 31 FIYFYFYFFL 7861.874 18 FLFFFLPFPL 2108.811
10 LLRITFNFFL 334.570 23 LPFPLVVFFI 31.429 128 FMQAAPWEGT 20.623 38
FFLEMESHYV 18.538 100 LLGLLKVRPL 16.705 46 YVAQAGLELL 9.690 4
ELLAGILLRI 6.659 9 ILLRITFNFF 4.898 22 FLPFPLVVFF 4.336 95
FIQCLLLGLL 4.040 97 QCLLLGLLKV 3.864 91 KAFRFIQCLL 3.842 5
LLAGILLRIT 2.389 13 ITFNFFLFFF 1.815 64 ASLVAGTLSV 1.680 105
KVRPLQHQGV 1.619 53 ELLGSSNPPA 1.379 20 FFFLPFPLVV 1.281 90
KKAFRFIQCL 0.908 14 TFNFFLFFFL 0.899 39 FLEMESHYVA 0.600 19
LFFFLPFPLV 0.577 27 LVVFFIYFYF 0.530 58 SNPPASASLV 0.454 28
VVFFIYFYFY 0.429 12 RITFNFFLFF 0.407 87 KKLKKAFRFI 0.392 33
YFYFYFFLEM 0.367 25 FPLVVFFIYF 0.329 102 GLLKVRPLQH 0.276 67
VAGTLSVHHC 0.270 69 GTLSVHHCAC 0.255 108 PLQHQGVNSC 0.251 8
GILLRITFNF 0.220 57 SSNPPASASL 0.139 123 YFQGIFMQAA 0.139 54
LLGSSNPPAS 0.127 99 LLLGLLKVRP 0.094 26 PLVVFFIYFY 0.079 70
TLSVHHCACF 0.075 65 SLVAGTLSVH 0.070 15 FNFFLFFFLP 0.069 29
VFFIYFYFYF 0.059 55 LGSSNPPASA 0.055 98 CLLLGLLKVR 0.052 126
GIFMQAAPWE 0.042 41 EMESHYVAQA 0.040 80 ESFTKRKKKL 0.039 72
SVHHCACFES 0.038 94 RFIQCLLLGL 0.034 68 AGTLSVHHCA 0.032 62
ASASLVAGTL 0.018 48 AQAGLELLGS 0.017 88 KLKKAFRFIQ 0.016 59
NPPASASLVA 0.013 40 LEMESHYVAQ 0.011 66 LVAGTLSVHH 0.011 43
ESHYVAQAGL 0.010 17 FFLFFFLPFP 0.008 50 AGLELLGSSN 0.007 124
FQGIFMQAAP 0.007 7 AGILLRITFN 0.006 77 ACFESFTKRK 0.006 61
PASASLVAGT 0.005 122 GYFQGIFMQA 0.005 121 RGYFQGIFMQ 0.004 117
CDCERGYFQG 0.004 74 HHCACFESFT 0.004 110 QHQGVNSCDC 0.003 113
GVNSCDCERG 0.003 96 IQCLLLGLLK 0.003 109 LQHQGVNSCD 0.003 30
FFIYFYFYFF 0.002 3 RELLAGILLR 0.002 42 MESHYVAQAG 0.002 127
IFMQAAPWEG 0.002 103 LLKVRPLQHQ 0.002 52 LELLGSSNPP 0.002 107
RPLQHQGVNS 0.002 6 LAGILLRITF 0.002 47 VAQAGLELLG 0.002 115
NSCDCERGYF 0.001 16 NFFLFFFLPF 0.001 79 FESFTKRKKK 0.001 83
TKRKKKLKKA 0.001 92 AFRFIQCLLL 0.001 63 SASLVAGTLS 0.001 51
GLELLGSSNP 0.001 71 LSVHHCACFE 0.001 37 YFFLEMESHY 0.001 21
FFLPFPLVVF 0.001 89 LKKAFRFIQC 0.001 35 YFYFFLEMES 0.001 118
DCERGYFQGI 0.001 101 LGLLKVRPLQ 0.001 125 QGIFMQAAPW 0.000 56
GSSNPPASAS 0.000 93 FRFIQCLLLG 0.000 V10-HLA-A201-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 21; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 8
ELGTSDVVTV 11.998 9 LGTSDVVTVV 0.728 10 GTSDVVTVVL 0.499 7
GELGTSDVVT 0.220 5 PAGELGTSDV 0.087 6 AGELGTSDVV 0.006 2 GRCPAGELGT
0.001 3 RCPAGELGTS 0.000 4 CPAGELGTSD 0.000 1 TGRCPAGELG 0.000
V11-HLA-A201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 23; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 RVMVPPLPSL 15.907 1 FQARLRLRVM 0.437 9
VMVPPLPSLN 0.091 2 QARLRLRVMV 0.073 5 LRLRVMVPPL 0.043 4 RLRLRVMVPP
0.003 10 MVPPLPSLNP 0.002 6 RLRVMVPPLP 0.001 7 LRVMVPPLPS 0.000 3
ARLRLRVMVP 0.000 V12-HLA-A201-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 9 GCSYSTLTTV 1.044 1
SVMSEEPEGC 0.788 2 VMSEEPEGCS 0.049 0 EEPEGCSYST 0.045 8 EGCSYSTLTT
0.004 7 PEGCSYSTLT 0.003 6 EPEGCSYSTL 0.001 4 SEEPEGCSYS 0.001 3
MSEEPEGCSY 0.000 10 CSYSTLTTVR 0.000 11 SYSTLTTVRE 0.000
V13-HLA-A201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 VLADPQEDSG 0.255 2 SQVTVDVLAD 0.003 3
QVTVDVLADP 0.003 1 DSQVTVDVLA 0.002 7 DVLADPQEDS 0.001 4 VTVDVLADPQ
0.001 5 TVDVLADPQE 0.001 9 LADPQEDSGK 0.000 6 VDVLADPQED 0.000 10
ADPQEDSGKQ 0.000 V14-HLA-A201-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 29; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 10 ASLVAGTLSV 1.680 4
SNPPASASLV 0.454 3 SSNPPASASL 0.139 1 LGSSNPPASA 0.055 8 ASASLVAGTL
0.018 5 NPPASASLVA 0.013 7 PASASLVAGT 0.005 9 SASLVAGTLS 0.001 2
GSSNPPASAS 0.000 6 PPASASLVAG 0.000
[1026]
22TABLE XII Start Subsequence Score V1-HLA-A3-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 20
LLLASFTGR 18.000 435 VMSEEPEGR 6.000 369 VLMSRYHRR 6.000 370
LMSRYHRRK 6.000 17 LLLLLLASF 4.500 362 CLLVVVVVL 4.050 391
TLTRENSIR 4.000 107 PLDGSVLLR 3.600 145 VLVPPLPSL 3.038 189
GTTSSRSFK 3.000 41 TVVLGQDAK 3.000 80 ALLHSKYGL 2.700 365 VVVVVLMSR
2.700 459 ELLSPGSGR 2.700 8 EMWGPEAWL 2.025 180 SVTWDTEVK 2.000 61
QVGQVAWAR 1.800 368 VVLMSRYHR 1.800 142 RLRVLVPPL 1.800 359
LLFCLLVVV 1.500 363 LLVVVVVLM 1.350 316 HVSNEFSSR 1.200 252
GLEDQNLWH 1.200 78 ELALLHSKY 1.200 366 VVVVLMSRY 0.900 358
ALLFCLLVV 0.900 477 GIKQAMNHF 0.900 15 WLLLLLLLA 0.900 89 HVSPAYEGR
0.600 294 RVDGDTLGF 0.600 485 FVQENGTLR 0.600 97 RVEQPPPPR 0.600
215 SMNGQPLTC 0.600 392 LTRENSIRR 0.600 230 LLQDQRITH 0.400 351
VVVGVIAAL 0.304 313 YVCHVSNEF 0.300 112 VLLRNAVQA 0.300 299
TLGFPPLTT 0.300 164 GLTLAASCT 0.300 354 GVIAALLFC 0.270 45
GQDAKLPCF 0.270 355 VIAALLFCL 0.270 255 DQNLWHIGR 0.216 132
FPAGSFQAR 0.180 350 VVVVGVIAA 0.180 16 LLLLLLLAS 0.180 186
EVKGTTSSR 0.180 292 GVRVDGDTL 0.180 206 SEFHLVPSR 0.180 481
AMNHFVQEN 0.180 21 LLASFTGRC 0.180 11 GPEAWLLLL 0.162 18 LLLLLASFT
0.150 77 QELALLHSK 0.135 42 VVLGQDAKL 0.135 238 HILHVSFLA 0.135 274
GQPPPSYNW 0.121 378 KAQQMTQKY 0.120 239 ILHVSFLAE 0.120 117
AVQADEGEY 0.120 140 RLRLRVLVP 0.120 498 GNGIYINGR 0.108 236
ITHILHVSF 0.100 352 VVGVIAALL 0.090 19 LLLLASFTG 0.090 135
GSFQARLRL 0.090 4 SLGAEMWGP 0.090 344 DLVSASVVV 0.090 305 LTTEHSGIY
0.090 460 LLSPGSGRA 0.090 382 MTQKYEEEL 0.090 420 AEGHPDSLK 0.090
284 RLDGPLPSG 0.068 261 IGREGAMLK 0.060 417 GLRAEGHPD 0.060 81
LLHSKYGLH 0.060 203 AVTSEFHLV 0.060 192 SSRSFKHSR 0.060 260
HIGREGAML 0.060 304 PLTTEHSGI 0.060 113 LLRNAVQAD 0.060 87
GLHVSPAYE 0.060 345 LVSASVVVV 0.060 364 LVVVVVLMS 0.054 495
KPTGNGIYI 0.054 47 DAKLPCFYR 0.054 411 QPEESVGLR 0.054 209
HLVPSRSMN 0.045 229 GLLQDQRIT 0.045 349 SVVVVGVIA 0.045 390
LTLTRENSI 0.045 158 ALEEGQGLT 0.045 266 AMLKCLSEG 0.045 227
HPGLLQDQR 0.040 426 SLKDNSSCS 0.040 276 PPPSYNWTR 0.036 386
YEEELTLTR 0.036 377 RKAQQMTQK 0.030 244 FLAEASVRG 0.030
V2-HLA-A3-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
5; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight 8 CLYRGDSGE 0.100 1 GQDAKLPCL 0.081 3 DAKLPCLYR
0.036 5 KLPCLYRGD 0.006 2 QDAKLPCLY 0.004 6 LPCLYRGDS 0.000 4
AKLPCLYRG 0.000 7 PCLYRGDSG 0.000 9 LYRGDSGEQ 0.000
V7-HLA-A3-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
15; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight 8 SQSEEPEGR 0.180 3 HTDPRSQSE 0.002 7 RSQSEEPEG
0.000 2 HHTDPRSQS 0.000 5 DPRSQSEEP 0.000 4 TDPRSQSEE 0.000 6
PRSQSEEPE 0.000 1 SHHTDPRSQ 0.000 V9-HLA-A3-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 19; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 31
FIYFYFYFF 27.000 9 ILLRITFNF 13.500 13 ITFNFFLFF 9.000 27 LVVFFIYFY
8.100 99 LLLGLLKVR 6.750 10 LLRITFNFF 6.000 26 PLVVFFIYF 5.400 4
ELLAGILLR 5.400 28 VVFFIYFYF 4.500 22 FLPFPLVVF 4.500 5 LLAGILLRI
4.050 12 RITFNFFLF 1.800 113 GVNSCDCER 1.200 98 CLLLGLLKV 0.900 77
ACFESFTKR 0.900 25 FPLVVFFIY 0.810 76 CACFESFTK 0.600 65 SLVAGTLSV
0.600 97 QCLLLGLLK 0.600 88 KLKKAFRFI 0.540 29 VFFIYFYFY 0.540 82
FTKRKKKLK 0.500 23 LPFPLVVFF 0.450 18 FLFFFLPFP 0.450 91 KAFRFIQCL
0.405 103 LLKVRPLQH 0.400 126 GIFMQAAPW 0.300 70 TLSVHHCAC 0.200 54
LLGSSNPPA 0.200 39 FLEMESHYV 0.200 95 FIQCLLLGL 0.180 102 GLLKVRPLQ
0.135 46 YVAQAGLEL 0.120 80 ESFTKRKKK 0.075 69 GTLSVHHCA 0.068 128
FMQAAPWEG 0.060 51 GLELLGSSN 0.060 15 FNFFLFFFL 0.054 17 FFLFFFLPF
0.054 66 LVAGTLSVH 0.045 83 TKRKKKLKK 0.040 78 CFESFTKRK 0.030 30
FFIYFYFYF 0.027 14 TFNFFLFFF 0.027 32 IYFYFYFFL 0.027 124 FQGIFMQAA
0.027 81 KKLKKAFRF 0.027 119 CERGYFQGI 0.024 100 LLGLLKVRP 0.020
109 LQHQGVNSC 0.018 34 FYFYFFLEM 0.018 71 LSVHHCACF 0.015 53
ELLGSSNPP 0.013 8 GILLRITFN 0.013 86 KKKLKKAFR 0.012 38 FFLEMESHY
0.009 47 VAQAGLELL 0.009 105 KVRPLQHQG 0.009 19 LFFFLPFPL 0.009 11
LRITFNFFL 0.008 96 IQCLLLGLL 0.008 74 HHCACFESF 0.006 7 AGILLRITF
0.006 41 EMESHYVAQ 0.006 116 SCDCERGYF 0.006 111 HQGVNSCDC 0.006 93
FRFIQCLLL 0.006 79 FESFTKRKK 0.006 3 RELLAGILL 0.005 42 MESHYVAQA
0.005 56 GSSNPPASA 0.005 20 FFFLPFPLV 0.005 129 MQAAPWEGT 0.005 40
LEMESHYVA 0.004 108 PLQHQGVNS 0.004 90 KKAFRFIQC 0.004 44 SHYVAQAGL
0.003 75 HCACFESFT 0.003 123 YFQGIFMQA 0.003 16 NFFLFFFLP 0.003 21
FFLPFPLVV 0.003 33 YFYFYFFLE 0.003 63 SASLVAGTL 0.003 72 SVHHCACFE
0.002 115 NSCDCERGY 0.002 67 VAGTLSVHH 0.002 121 RGYFQGIFM 0.002 59
NPPASASLV 0.002 58 SNPPASASL 0.002 48 AQAGLELLG 0.002 37 YFFLEMESH
0.002 62 ASASLVAGT 0.002 122 GYFQGIFMQ 0.001 1 MRRELLAGI 0.001 49
QAGLELLGS 0.001 85 RKKKLKKAF 0.001 92 AFRFIQCLL 0.001 24 PFPLVVFFI
0.001 68 AGTLSVHHC 0.001 120 ERGYFQGIF 0.001
V10-HLA-A3-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
21; each start position is specified the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 9 GTSDVVTVV 0.135 7 ELGTSDVVT 0.030 6
GELGTSDVV 0.004 2 RCPAGELGT 0.002 8 LGTSDVVTV 0.001 3 CPAGELGTS
0.000 5 AGELGTSDV 0.000 1 GRCPAGELG 0.000 4 PAGELGTSD 0.000
V11-HLA-A3-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
23; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 8 VMVPPLPSL 3.038 5 RLRVMVPPL 1.800 3
RLRLRVMVP 0.120 7 RVMVPPLPS 0.018 9 MVPPLPSLN 0.003 1 QARLRLRVM
0.000 2 ARLRLRVMV 0.000 4 LRLRVMVPP 0.000 6 LRVMVPPLP 0.000
V13-HLA-A3-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
27; each start position is specified the length of peptide is 9
amino acids and the end position for each peptide is the start
position plus eight. 7 VLADPQEDS 0.060 9 ADPQEDSGK 0.020 1
SQVTVDVLA 0.013 2 QVTVDVLAD 0.012 3 VTVDVLADP 0.003 4 TVDVLADPQ
0.002 6 DVLADPQED 0.001 8 LADPQEDSG 0.000 5 VDVLADPQE 0.000
V14-HLA-A39-mers-191P4D12B Each peptide is a portion of SEQ ID NO:
29; each start position is specified the length of peptide is 9
amino acids and the end position for each peptide is the start
position plus eight 1 GSSNPPASA 0.005 8 SASLVAGTL 0.003 4 NPPASASLV
0.002 3 SNPPASASL 0.002 7 ASASLVAGT 0.002 2 SSNPPASAS 0.000 5
PPASASLVA 0.000 9 ASLVAGTLS 0.000 6 PASASLVAG 0.000
[1027]
23TABLE XIII Start Subsequence Score V1-HLA-A3-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 332
VLDPQEDSGK 30.000 19 LLLLASFTGR 18.000 369 VLMSRYHRRK 9.000 252
GLEDQNLWHI 8.100 391 TLTRENSIRR 8.000 16 LLLLLLLASF 4.500 8
EMWGPEAWLL 4.050 400 RLHSHHTDPR 4.000 260 HIGREGAMLK 4.000 359
LLFCLLVVVV 3.000 364 LVVVVVLMSR 2.700 381 QMTQKYEEEL 1.800 158
ALEEGQGLTL 1.800 229 GLLQDQRITH 1.800 367 VVVLMSRYHR 1.800 40
VTVVLGQDAK 1.500 362 CLLVVVVVLM 1.350 354 GVIAALLFCL 1.215 81
LLHSKYGLHV 1.200 257 NLWHIGREGA 1.000 76 AQELALLHSK 0.900 365
VVVVVLMSRY 0.900 239 ILHVSFLAEA 0.900 230 LLQDQRITHI 0.900 215
SMNGQPLTCV 0.675 434 SVMSEEPEGR 0.600 164 GLTLAASCTA 0.600 368
VVLMSRYHRR 0.600 363 LLVVVVVLMS 0.540 275 QPPPSYNWTR 0.540 419
RAEGHPDSLK 0.450 358 ALLFCLLVVV 0.450 123 GEYECRVSTF 0.405 43
VLGQDAKLPC 0.400 352 VVGVIAALLF 0.400 60 EQVGQVAWAR 0.364 106
NPLDGSVLLR 0.360 45 GQDAKLPCFY 0.360 390 LTLTRENSIR 0.300 284
RLDGPLPSGV 0.300 244 FLAEASVRGL 0.270 500 GIYINGRGHL 0.270 87
GLHVSPAYEG 0.270 344 DLVSASVVVV 0.270 20 LLLASFTGRC 0.270 130
STFPAGSFQA 0.225 144 RVLVPPLPSL 0.203 351 VVVGVIAALL 0.203 350
VVVVGVIAAL 0.203 426 SLKDNSSCSV 0.200 447 TLTTVREIET 0.200 235
RITHILHVSF 0.200 15 WLLLLLLLAS 0.180 33 ELETSDVVTV 0.180 355
VIAALLFCLL 0.180 349 SVVVVGVIAA 0.180 389 ELTLTRENSI 0.180 410
SQPEESVGLR 0.162 17 LLLLLLASFT 0.150 304 PLTTEHSGIY 0.120 417
GLRAEGHPDS 0.120 49 KLPCFYRGDS 0.108 443 RSYSTLTTVR 0.100 242
VSFLAEASVR 0.100 18 LLLLLASFTG 0.090 249 SVRGLEDQNL 0.090 209
HLVPSRSMNG 0.090 41 TVVLGQDAKL 0.090 80 ALLHSKYGLH 0.090 189
GTTSSRSFKH 0.090 486 VQENGTLRAK 0.090 152 SLNPGPALEE 0.090 112
VLLRNAVQAD 0.090 311 GIYVCHVSNE 0.090 236 ITHILHVSFL 0.090 128
RVSTFPAGSF 0.090 188 KGTTSSRSFK 0.060 270 CLSEGQPPPS 0.060 477
GIKQAMNHFV 0.060 485 FVQENGTLRA 0.060 191 TSSRSFKHSR 0.060 205
TSEFHLVPSR 0.060 119 QADEGEYECR 0.060 11 GPEAWLLLLL 0.054 218
GQPLTCVVSH 0.054 140 RLRLRVLVPP 0.045 299 TLGFPPLTTE 0.045 271
LSEGQPPPSY 0.045 135 GSFQARLRLR 0.045 145 VLVPPLPSLN 0.045 306
TTEHSGIYVC 0.045 96 GRVEQPPPPR 0.041 361 FCLLVVVVVL 0.041 341
KQVDLVSASV 0.041 181 VTWDTEVKGT 0.037 385 KYEEELTLTR 0.036 383
TQKYEEELTL 0.036 376 RRKAQQMTQK 0.030 305 LTTEHSGIYV 0.030 221
LTCVVSHPGL 0.030 V2-HLA-A3-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 2 GQDAKLPCLY 0.360 6
KLPCLYRGDS 0.108 9 CLYRGDSGEQ 0.030 3 QDAKLPCLYR 0.012 1 LGQDAKLPCL
0.001 10 LYRGDSGEQV 0.000 4 DAKLPCLYRG 0.000 7 LPCLYRGDSG 0.000 8
PCLYRGDSGE 0.000 5 AKLPCLYRGD 0.000 V7-HLA-A3-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 8
RSQSEEPEGR 0.020 4 HTDPRSQSEE 0.002 9 SQSEEPEGRS 0.001 2 SHHTDPRSQS
0.000 6 DPRSQSEEPE 0.000 5 TDPRSQSEEP 0.000 3 HHTDPRSQSE 0.000 1
HSHHTDPRSQ 0.000 7 PRSQSEEPEG 0.000 V9-HLA-A3-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 19; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 28
VVFFIYFYFY 54.000 18 FLFFFLPFPL 9.000 9 ILLRITFNFF 9.000 26
PLVVFFIYFY 8.100 13 ITFNFFLFFF 6.750 22 FLPFPLVVFF 6.000 10
LLRITFNFFL 5.400 98 CLLLGLLKVR 4.500 8 GILLRITFNF 4.050 12
RITFNFFLFF 3.600 31 FIYFYFYFFL 2.700 77 ACFESFTKRK 2.250 82
FTKRKKKLKK 2.000 70 TLSVHHCACF 2.000 102 GLLKVRPLQH 1.800 27
LVVFFIYFYF 1.350 4 ELLAGILLRI 1.215 96 IQCLLLGLLK 1.200 23
LPFPLVVFFI 0.608 75 HCACFESFTK 0.600 39 FLEMESHYVA 0.600 25
FPLVVFFIYF 0.540 88 KLKKAFRFIQ 0.540 41 EMESHYVAQA 0.540 65
SLVAGTLSVH 0.450 100 LLGLLKVRPL 0.180 16 NFFLFFFLPF 0.180 128
FMQAAPWEGT 0.150 53 ELLGSSNPPA 0.135 91 KAFRFIQCLL 0.135 76
CACFESFTKR 0.120 105 KVRPLQHQGV 0.090 46 YVAQAGLELL 0.090 29
VFFIYFYFYF 0.090 30 FFIYFYFYFF 0.081 51 GLELLGSSNP 0.060 108
PLQHQGVNSC 0.060 3 RELLAGILLR 0.054 69 GTLSVHHCAC 0.045 99
LLLGLLKVRP 0.045 103 LLKVRPLQHQ 0.045 54 LLGSSNPPAS 0.040 6
LAGILLRITF 0.040 66 LVAGTLSVHH 0.030 79 FESFTKRKKK 0.030 126
GIFMQAAPWE 0.030 122 GYFQGIFMQA 0.027 11 LRITFNFFLF 0.027 95
FIQCLLLGLL 0.027 5 LLAGILLRIT 0.022 37 YFFLEMESHY 0.020 86
KKKLKKAFRF 0.018 33 YFYFYFFLEM 0.018 118 DCERGYFQGI 0.016 72
SVHHCACFES 0.012 21 FFLPFPLVVF 0.010 81 SFTKRKKKLK 0.010 97
QCLLLGLLKV 0.009 90 KKAFRFIQCL 0.008 119 CERGYFQGIF 0.008 112
QGVNSCDCER 0.006 73 VHHCACFESF 0.006 67 VAGTLSVHHC 0.006 113
GVNSCDCERG 0.006 20 FFFLPFPLW 0.006 24 PFPLVVFFIY 0.005 15
FNFFLFFFLP 0.005 48 AQAGLELLGS 0.005 14 TFNFFLFFFL 0.005 19
LFFFLPFPLV 0.005 57 SSNPPASASL 0.005 85 RKKKLKKAFR 0.004 59
NPPASASLVA 0.004 84 KRKKKLKKAF 0.003 64 ASLVAGTLSV 0.003 115
NSCDCERGYF 0.003 94 RFIQCLLLGL 0.003 32 IYFYFYFFLE 0.003 80
ESFTKRKKKL 0.002 78 CFESFTKRKK 0.002 45 HYVAQAGLEL 0.002 36
FYFFLEMESH 0.002 123 YFQGIFMQAA 0.001 62 ASASLVAGTL 0.001 2
RRELLAGILL 0.001 89 LKKAFRFIQC 0.001 92 AFRFIQCLLL 0.001 109
LQHQGVNSCD 0.001 56 GSSNPPASAS 0.001 43 ESHYVAQAGL 0.001 87
KKLKKAFRFI 0.001 114 VNSCDCERGY 0.001 116 SCDCERGYFQ 0.001 111
HQGVNSCDCE 0.001 58 SNPPASASLV 0.001 107 RPLQHQGVNS 0.001 124
FQGIFMQAAP 0.001 38 FFLEMESHYV 0.000 34 FYFYFFLEME 0.000 121
RGYFQGIFMQ 0.000 V10-HLA-A3-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 21; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 8 ELGTSDVVTV 0.180 10
GTSDVVTVVL 0.135 7 GELGTSDVVT 0.002 2 GRCPAGELGT 0.001 9 LGTSDVVTVV
0.001 5 PAGELGTSDV 0.000 4 CPAGELGTSD 0.000 6 AGELGTSDVV 0.000 3
RCPAGELGTS 0.000 1 TGRCPAGELG 0.000 V11-HLA-A3-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 23; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 8
RVMVPPLPSL 0.203 9 VMVPPLPSLN 0.045 4 RLRLRVMVPP 0.045 6 RLRVMVPPLP
0.030 10 MVPPLPSLNP 0.009 5 LRLRVMVPPL 0.003 2 QARLRLRVMV 0.002 1
FQARLRLRVM 0.001 7 LRVMVPPLPS 0.000 3 ARLRLRVMVP 0.000
V12-HLA-A3-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 10 CSYSTLTTVR 0.100 1 SVMSEEPEGC 0.030 3
MSEEPEGCSY 0.030 2 VMSEEPEGCS 0.027 9 GCSYSTLTTV 0.009 6 EPEGCSYSTL
0.003 5 EEPEGCSYST 0.000 4 SEEPEGCSYS 0.000 7 PEGCSYSTLT 0.000 8
EGCSYSTLTT 0.000 11 SYSTLTTVRE 0.000 V13-HLA-A3-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 27; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 9
LADPQEDSGK 0.300 8 VLADPQEDSG 0.026 2 SQVTVDVLAD 0.005 3 QVTVDVLADP
0.005 7 DVLADPQEDS 0.003 5 TVDVLADPQE 0.002 4 VTVDVLADPQ 0.002 1
DSQVTVDVLA 0.000 6 VDVLADPQED 0.000 10 ADPQEDSGKQ 0.000
V14-HLA-A3-10mers-191P4D12B Each peptide s a portion of SEQ ID NO:
29; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 3 SSNPPASASL 0.005 5 NPPASASLVA 0.004 10
ASLVAGTLSV 0.003 8 ASASLVAGTL 0.001 2 GSSNPPASAS 0.001 4 SNPPASASLV
0.001 9 SASLVAGTLS 0.000 1 LGSSNPPASA 0.000 7 PASASLVAGT 0.000 6
PPASASLVAG 0.000
[1028]
24TABLE XIV Start Subsequence Score V1-HLA-A1101-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 41
TVVLGQDAK 3.000 189 GTTSSRSFK 3.000 180 SVTWDTEVK 2.000 365
VVVVVLMSR 1.200 97 RVEQPPPPR 1.200 368 VVLMSRYHR 1.200 61 QVGQVAWAR
0.800 485 FVQENGTLR 0.400 392 LTRENSIRR 0.400 89 HVSPAYEGR 0.400
316 HVSNEFSSR 0.400 369 VLMSRYHRR 0.160 186 EVKGTTSSR 0.120 294
RVDGDTLGF 0.120 20 LLLASFTGR 0.120 77 QELALLHSK 0.090 391 TLTRENSIR
0.080 444 SYSTLTTVR 0.080 435 VMSEEPEGR 0.080 255 DQNLWHIGR 0.072
377 RKAQQMTQK 0.060 292 GVRVDGDTL 0.060 350 VVVVGVIAA 0.060 420
AEGHPDSLK 0.060 243 SFLAEASVR 0.060 370 LMSRYHRRK 0.040 411
QPEESVGLR 0.040 261 IGREGAMLK 0.040 227 HPGLLQDQR 0.040 132
FPAGSFQAR 0.040 459 ELLSPGSGR 0.036 47 DAKLPCFYR 0.036 274
GQPPPSYNW 0.036 42 VVLGQDAKL 0.030 349 SVVVVGVIA 0.030 190
TTSSRSFKH 0.030 366 VVVVLMSRY 0.030 351 VVVGVIAAL 0.030 223
CVVSHPGLL 0.030 498 GNGIYINGR 0.024 386 YEEELTLTR 0.024 206
SEFHLVPSR 0.024 252 GLEDQNLWH 0.024 117 AVQADEGEY 0.020 342
QVDLVSASV 0.020 352 VVGVIAALL 0.020 333 LDPQEDSGK 0.020 306
TTEHSGIYV 0.020 345 LVSASVVVV 0.020 313 YVCHVSNEF 0.020 203
AVTSEFHLV 0.020 415 SVGLRAEGH 0.020 64 QVAWARVDA 0.020 238
HILHVSFLA 0.018 144 RVLVPPLPS 0.018 354 GVIAALLFC 0.018 471
EEDQDEGIK 0.018 45 GQDAKLPCF 0.018 107 PLDGSVLLR 0.016 40 VTVVLGQDA
0.015 390 LTLTRENSI 0.015 165 LTLAASCTA 0.015 75 GAQELALLH 0.012 85
KYGLHVSPA 0.012 358 ALLFCLLVV 0.012 11 GPEAWLLLL 0.012 495
KPTGNGIYI 0.012 486 VQENGTLRA 0.012 15 WLLLLLLLA 0.012 142
RLRVLVPPL 0.012 80 ALLHSKYGL 0.012 477 GIKQAMNHF 0.012 137
FQARLRLRV 0.012 355 VIAALLFCL 0.012 236 ITHILHVSF 0.010 382
MTQKYEEEL 0.010 305 LTTEHSGIY 0.010 287 GPLPSGVRV 0.009 202
AAVTSEFHL 0.009 230 LLQDQRITH 0.008 359 LLFCLLVVV 0.008 276
PPPSYNWTR 0.008 363 LLVVVVVLM 0.006 231 LQDQRITHI 0.006 112
VLLRNAVQA 0.006 410 SQPEESVGL 0.006 419 RAEGHPDSL 0.006 128
RVSTFPAGS 0.006 364 LVVVVVLMS 0.006 378 KAQQMTQKY 0.006 501
IYINGRGHL 0.006 69 RVDAGEGAQ 0.006 362 CLLVVVVVL 0.006 6 GAEMWGPEA
0.006 131 TFPAGSFQA 0.006 357 AALLFCLLV 0.006 17 LLLLLLASF 0.006
493 RAKPTGNGI 0.006 487 QENGTLRAK 0.006 301 GFPPLTTEH 0.006
V2-HLA-A1101-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 3 DAKLPCLYR 0.024 1 GQDAKLPCL 0.018 8
CLYRGDSGE 0.001 9 LYRGDSGEQ 0.000 6 LPCLYRGDS 0.000 2 QDAKLPCLY
0.000 5 KLPCLYRGD 0.000 4 AKLPCLYRG 0.000 7 PCLYRGDSG 0.000
V7-HLA-A1101-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 8 SQSEEPEGR 0.120 3 HTDPRSQSE 0.001 7
RSQSEEPEG 0.000 5 DPRSQSEEP 0.000 4 TDPRSQSEE 0.000 2 HHTDPRSQS
0.000 6 PRSQSEEPE 0.000 1 SHHTDPRSQ 0.000
V9-HLA-A1101-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 113 GVNSCDCER 1.200 76 CACFESFTK 0.600 97
QCLLLGLLK 0.600 82 FTKRKKKLK 0.500 28 VVFFIYFYF 0.120 78 CFESFTKRK
0.100 77 ACFESFTKR 0.080 4 ELLAGILLR 0.072 27 LVVFFIYFY 0.060 99
LLLGLLKVR 0.060 69 GTLSVHHCA 0.045 83 TKRKKKLKK 0.040 13 ITFNFFLFF
0.040 46 YVAQAGLEL 0.040 12 RITFNFFLF 0.036 126 GIFMQAAPW 0.024 32
IYFYFYFFL 0.024 66 LVAGTLSVH 0.020 9 ILLRITFNF 0.018 34 FYFYFFLEM
0.016 31 FIYFYFYFF 0.016 86 KKKLKKAFR 0.012 19 LFFFLPFPL 0.012 98
CLLLGLLKV 0.012 91 KAFRFIQCL 0.012 65 SLVAGTLSV 0.012 30 FFIYFYFYF
0.009 25 FPLVVFFIY 0.009 103 LLKVRPLQH 0.008 5 LLAGILLRI 0.008 95
FIQCLLLGL 0.008 29 VFFIYFYFY 0.008 122 GYFQGIFMQ 0.007 21 FFLPFPLVV
0.006 14 TFNFFLFFF 0.006 96 IQCLLLGLL 0.006 80 ESFTKRKKK 0.006 17
FFLFFFLPF 0.006 124 FQGIFMQAA 0.006 79 FESFTKRKK 0.006 105
KVRPLQHQG 0.006 3 RELLAGILL 0.005 37 YFFLEMESH 0.004 123 YFQGIFMQA
0.004 39 FLEMESHYV 0.004 10 LLRITFNFF 0.004 23 LPFPLVVFF 0.004 20
FFFLPFPLV 0.004 54 LLGSSNPPA 0.004 22 FLPFPLVVF 0.004 38 FFLEMESHY
0.003 87 KKLKKAFRF 0.003 15 FNFFLFFFL 0.002 121 RGYFQGIFM 0.002 40
LEMESHYVA 0.002 47 VAQAGLELL 0.002 92 AFRFIQCLL 0.002 116 SCDCERGYF
0.002 67 VAGTLSVHH 0.002 72 SVHHCACFE 0.002 59 NPPASASLV 0.002 63
SASLVAGTL 0.002 102 GLLKVRPLQ 0.002 94 RFIQCLLLG 0.002 8 GILLRITFN
0.002 36 FYFFLEMES 0.002 26 PLVVFFIYF 0.001 33 YFYFYFFLE 0.001 48
AQAGLELLG 0.001 88 KLKKAFRFI 0.001 16 NFFLFFFLP 0.001 51 GLELLGSSN
0.001 81 SFTKRKKKL 0.001 11 LRITFNFFL 0.001 107 RPLQHQGVN 0.001 128
FMQAAPWEG 0.001 18 FLFFFLPFP 0.001 93 FRFIQCLLL 0.001 2 RRELLAGIL
0.001 24 PFPLVVFFI 0.001 109 LQHQGVNSC 0.001 129 MQAAPWEGT 0.001
111 HQGVNSCDC 0.001 7 AGILLRITF 0.001 56 GSSNPPASA 0.001 45
HYVAQAGLE 0.001 119 CERGYFQGI 0.001 42 MESHYVAQA 0.001 44 SHYVAQAGL
0.000 100 LLGLLKVRP 0.000 70 TLSVHHCAC 0.000 35 YFYFFLEME 0.000 49
QAGLELLGS 0.000 127 IFMQAAPWE 0.000 58 SNPPASASL 0.000 60 PPASASLVA
0.000 71 LSVHHCACF 0.000 85 RKKKLKKAF 0.000 84 KRKKKLKKA 0.000 1
MRRELLAGI 0.000 V10-A1101-9mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 21; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 9 GTSDVVTVV 0.030 6 GELGTSDVV 0.003
2 RCPAGELGT 0.001 8 LGTSDVVTV 0.000 5 AGELGTSDV 0.000 3 CPAGELGTS
0.000 7 ELGTSDVVT 0.000 1 GRCPAGELG 0.000 4 PAGELGTSD 0.000
V11-A1101-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
23; each start position is specified,the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 7 RVMVPPLPS 0.024 5 RLRVMVPPL 0.012 8
VMVPPLPSL 0.006 3 RLRLRVMVP 0.002 9 MVPPLPSLN 0.002 2 ARLRLRVMV
0.000 1 QARLRLRVM 0.000 4 LRLRVMVPP 0.000 6 LRVMVPPLP 0.000
V12-A1101-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified,the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 8 GCSYSTLTT 0.001 3 SEEPEGCSY 0.001 9
CSYSTLTTV 0.000 1 VMSEEPEGC 0.000 5 EPEGCSYST 0.000 6 PEGCSYSTL
0.000 2 MSEEPEGCS 0.000 4 EEPEGCSYS 0.000 7 EGCSYSTLT 0.000
V13-A1101-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
27; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 9 ADPQEDSGK 0.020 1 SQVTVDVLA 0.009 2
QVTVDVLAD 0.004 4 TVDVLADPQ 0.002 3 VTVDVLADP 0.002 6 DVLADPQED
0.001 7 VLADPQEDS 0.000 8 LADPQEDSG 0.000 5 VDVLADPQE 0.000
V14-A1101-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
29; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 8 SASLVAGTL 0.002 4 NPPASASLV 0.002 1
GSSNPPASA 0.001 5 PPASASLVA 0.000 3 SNPPASASL 0.000 9 ASLVAGTLS
0.000 7 ASASLVAGT 0.000 2 SSNPPASAS 0.000 6 PASASLVAG 0.000
[1029]
25TABLE XV Start Subsequence Score V1-HLA-A1101-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 40
VTVVLGQDAK 1.500 364 LVVVVVLMSR 1.200 367 VVVLMSRYHR 1.200 260
HIGREGAMLK 0.800 434 SVMSEEPEGR 0.800 76 AQELALLHSK 0.600 419
RAEGHPDSLK 0.600 368 VVLMSRYHRR 0.600 385 KYEEELTLTR 0.480 332
VLDPQEDSGK 0.400 390 LTLTRENSIR 0.300 354 GVIAALLFCL 0.270 400
RLHSHHTDPR 0.240 391 TLTRENSIRR 0.160 19 LLLLASFTGR 0.120 106
NPLDGSVLLR 0.120 410 SQPEESVGLR 0.120 60 EQVGQVAWAR 0.108 189
GTTSSRSFKH 0.090 144 RVLVPPLPSL 0.090 369 VLMSRYHRRK 0.080 275
QPPPSYNWTR 0.080 486 VQENGTLRAK 0.060 188 KGTTSSRSFK 0.060 376
RRKAQQMTQK 0.060 349 SVVVVGVIAA 0.060 128 RVSTFPAGSF 0.060 484
HFVQENGTLR 0.060 130 STFPAGSFQA 0.060 119 QADEGEYECR 0.040 352
VVGVIAALLF 0.040 485 FVQENGTLRA 0.040 131 TFPAGSFQAR 0.040 229
GLLQDQRITH 0.036 41 TVVLGQDAKL 0.030 365 VVVVVLMSRY 0.030 350
VVVVGVIAAL 0.030 111 SVLLRNAVQA 0.030 351 VVVGVIAALL 0.030 63
GQVAWARVDA 0.027 341 KQVDLVSASV 0.027 443 RSYSTLTTVR 0.024 500
GIYINGRGHL 0.024 252 GLEDQNLWHI 0.024 342 QVDLVSASVV 0.020 61
QVGQVAWARV 0.020 249 SVRGLEDQNL 0.020 305 LTTEHSGIYV 0.020 241
HVSFLAEASV 0.020 89 HVSPAYEGRV 0.020 39 VVTVVLGQDA 0.020 96
GRVEQPPPPR 0.018 470 EEEDQDEGIK 0.018 185 TEVKGTTSSR 0.018 218
GQPLTCVVSH 0.018 458 TELLSPGSGR 0.018 45 GQDAKLPCFY 0.018 46
QDAKLPCFYR 0.012 11 GPEAWLLLLL 0.012 477 GIKQAMNHFV 0.012 235
RITHILHVSF 0.012 164 GLTLAASCTA 0.012 85 KYGLHVSPAY 0.012 383
TQKYEEELTL 0.012 284 RLDGPLPSGV 0.012 373 RYHRRKAQQM 0.012 25
FTGRCPAGEL 0.010 221 LTCVVSHPGL 0.010 236 ITHILHVSFL 0.010 359
LLFCLLVVVV 0.008 242 VSFLAEASVR 0.008 158 ALEEGQGLTL 0.008 257
NLWHIGREGA 0.008 81 LLHSKYGLHV 0.008 315 CHVSNEFSSR 0.006 88
LHVSPAYEGR 0.006 156 GPALEEGQGL 0.006 358 ALLFCLLVVV 0.006 501
IYINGRGHLV 0.006 201 SAAVTSEFHL 0.006 79 LALLHSKYGL 0.006 80
ALLHSKYGLH 0.006 231 LQDQRITHIL 0.006 493 RAKPTGNGIY 0.006 357
AALLFCLLVV 0.006 97 RVEQPPPPRN 0.006 362 CLLVVVVVLM 0.006 294
RVDGDTLGFP 0.006 16 LLLLLLLASF 0.006 312 IYVCHVSNEF 0.006 69
RVDAGEGAQE 0.006 6 GAEMWGPEAW 0.006 292 GVRVDGDTLG 0.006 223
CVVSHPGLLQ 0.006 8 EMWGPEAWLL 0.005 490 GTLRAKPTGN 0.005 239
ILHVSFLAEA 0.004 426 SLKDNSSCSV 0.004 411 QPEESVGLRA 0.004 146
LVPPLPSLNP 0.004 V2-HLA-A1101-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 2 GQDAKLPCLY 0.018 3
QDAKLPCLYR 0.008 10 LYRGDSGEQV 0.004 6 KLPCLYRGDS 0.001 9
CLYRGDSGEQ 0.001 7 LPCLYRGDSG 0.000 1 LGQDAKLPCL 0.000 4 DAKLPCLYRG
0.000 8 PCLYRGDSGE 0.000 5 AKLPCLYRGD 0.000
V7-HLA-A1101-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 RSQSEEPEGR 0.012 4 HTDPRSQSEE 0.001 9
SQSEEPEGRS 0.001 6 DPRSQSEEPE 0.000 5 TDPRSQSEEP 0.000 3 HHTDPRSQSE
0.000 2 SHHTDPRSQS 0.000 7 PRSQSEEPEG 0.000 1 HSHHTDPRSQ 0.000
V9-HLA-A1101- 10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 82 FTKRKKKLKK 2.000 96 IQCLLLGLLK 1.200 75
HCACFESFTK 0.600 77 ACFESFTKRK 0.200 3 RELLAGILLR 0.108 81
SFTKRKKKLK 0.100 27 LVVFFIYFYF 0.090 28 VVFFIYFYFY 0.080 98
CLLLGLLKVR 0.060 105 KVRPLQHQGV 0.060 13 ITFNFFLFFF 0.060 8
GILLRITFNF 0.054 122 GYFQGIFMQA 0.048 76 CACFESFTKR 0.040 102
GLLKVRPLQH 0.036 79 FESFTKRKKK 0.030 12 RITFNFFLFF 0.024 31
FIYFYFYFFL 0.024 18 FLFFFLPFPL 0.024 46 YVAQAGLELL 0.020 78
CFESFTKRKK 0.020 66 LVAGTLSVHH 0.020 94 RFIQCLLLGL 0.018 85
RKKKLKKAFR 0.012 91 KAFRFIQCLL 0.012 29 VFFIYFYFYF 0.012 10
LLRITFNFFL 0.012 45 HYVAQAGLEL 0.012 23 LPFPLVVFFI 0.012 20
FFFLPFPLVV 0.008 16 NFFLFFFLPF 0.008 33 YFYFYFFLEM 0.008 36
FYFFLEMESH 0.008 39 FLEMESHYVA 0.008 112 QGVNSCDCER 0.006 9
LLRITFNFF 0.006 72 SVHHCACFES 0.006 65 SLVAGTLSVH 0.006 25
FPLVVFFIYF 0.006 113 GVNSCDCERG 0.006 30 FFIYFYFYFF 0.006 97
QCLLLGLLKV 0.006 14 TFNFFLFFFL 0.006 69 GTLSVHHCAC 0.005 6
LAGILLRITF 0.004 37 YFFLEMESHY 0.004 59 NPPASASLVA 0.004 22
FLPFPLVVFF 0.004 19 LFFFLPFPLV 0.004 70 TLSVHHCACF 0.004 92
AFRFIQCLLL 0.004 95 FIQCLLLGLL 0.004 88 KLKKAFRFIQ 0.004 4
ELLAGILLRI 0.004 21 FFLPFPLVVF 0.003 38 FFLEMESHYV 0.003 32
IYFYFYFFLE 0.002 126 GIFMQAAPWE 0.002 123 YFQGIFMQAA 0.002 86
KKKLKKAFRF 0.002 53 ELLGSSNPPA 0.002 51 GLELLGSSNP 0.001 2
RRELLAGILL 0.001 48 AQAGLELLGS 0.001 26 PLVVFFIYFY 0.001 41
EMESHYVAQA 0.001 11 LRITFNFFLF 0.001 107 RPLQHQGVNS 0.001 34
FYFYFFLEME 0.001 127 IFMQAAPWEG 0.001 35 YFYFFLEMES 0.001 24
PFPLVVFFIY 0.001 64 ASLVAGTLSV 0.001 99 LLLGLLKVRP 0.001 90
KKAFRFIQCL 0.001 111 HQGVNSCDCE 0.001 124 FQGIFMQAAP 0.001 109
LQHQGVNSCD 0.001 119 CERGYFQGIF 0.001 118 DCERGYFQGI 0.001 128
FMQAAPWEGT 0.000 116 SCDCERGYFQ 0.000 47 VAQAGLELLG 0.000 54
LLGSSNPPAS 0.000 100 LLGLLKVRPL 0.000 58 SNPPASASLV 0.000 103
LLKVRPLQHQ 0.000 121 RGYFQGIFMQ 0.000 125 QGIFMQAAPW 0.000 84
KRKKKLKKAF 0.000 17 FFLFFFLPFP 0.000 15 FNFFLFFFLP 0.000 115
NSCDCERGYF 0.000 63 SASLVAGTLS 0.000 68 AGTLSVHHCA 0.000 73
VHHCACFESF 0.000 49 QAGLELLGSS 0.000 1 MRRELLAGIL 0.000 67
VAGTLSVHHC 0.000 62 ASASLVAGTL 0.000 V10-HLA-A1101-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 21; each start position is
specified,the length of peptide is 10 amino acids and the end
position for each peptide is the start position plus nine. 10
GTSDVVTVVL 0.030 8 ELGTSDVVTV 0.001 3 RCPAGELGTS 0.001 7 GELGTSDVVT
0.000 9 LGTSDVVTVV 0.000 6 AGELGTSDVV 0.000 4 CPAGELGTSD 0.000 5
PAGELGTSDV 0.000 2 GRCPAGELGT 0.000 1 TGRCPAGELG 0.000
V11-HLA-A1101-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 23; each start position is specified,the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 RVMVPPLPSL 0.120 10 MVPPLPSLNP 0.004 2
QARLRLRVMV 0.002 6 RLRVMVPPLP 0.001 4 RLRLRVMVPP 0.001 9 VMVPPLPSLN
0.001 1 FQARLRLRVM 0.001 5 LRLRVMVPPL 0.000 3 ARLRLRVMVP 0.000 7
LRVMVPPLPS 0.000 V12-HLA-A1101-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 10 amino acids and the end position for each
peptide is the start position plus nine. 10 CSYSTLTTVR 0.008 9
GCSYSTLTTV 0.006 1 SVMSEEPEGC 0.004 6 EPEGCSYSTL 0.001 11
SYSTLTTVRE 0.000 2 VMSEEPEGCS 0.000 3 MSEEPEGCSY 0.000 4 SEEPEGCSYS
0.000 5 EEPEGCSYST 0.000 8 EGCSYSTLTT 0.000 7 PEGCSYSTLT 0.000
V13-HLA-A1101-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 9 LADPQEDSGK 0.200 5 TVDVLADPQE 0.002 3
QVTVDVLADP 0.002 2 SQVTVDVLAD 0.002 4 VTVDVLADPQ 0.002 7 DVLADPQEDS
0.001 8 VLADPQEDSG 0.000 1 DSQVTVDVLA 0.000 6 VDVLADPQED 0.000 10
ADPQEDSGKQ 0.000 V14-HLA-A1101-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 29; each start position is specified, the
length of peptide is 10 amino acids and the end position for each
peptide is the start position plus nine. 5 NPPASASLVA 0.004 10
ASLVAGTLSV 0.001 4 SNPPASASLV 0.000 8 ASASLVAGTL 0.000 9 SASLVAGTLS
0.000 1 LGSSNPPASA 0.000 3 SSNPPASASL 0.000 2 GSSNPPASAS 0.000 6
PPASASLVAG 0.000 7 PASASLVAGT 0.000
[1030]
26TABLE XVI Start Subsequence Score V1-HLA-A24-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids and the end
position for each peptide is the start position plus eight. 501
IYINGRGHL 300.000 124 EYECRVSTF 150.000 484 HFVQENGTL 30.000 385
KYEEELTLT 18.000 105 RNPLDGSVL 12.000 419 RAEGHPDSL 12.000 85
KYGLHVSPA 10.000 142 RLRVLVPPL 9.600 100 QPPPPRNPL 8.640 362
CLLVVVVVL 8.400 351 VVVGVIAAL 8.400 14 AWLLLLLLL 7.200 410
SQPEESVGL 7.200 145 VLVPPLPSL 7.200 106 NPLDGSVLL 7.200 10
WGPEAWLLL 7.200 42 VVLGQDAKL 6.600 382 MTQKYEEEL 6.600 71 DAGEGAQEL
6.336 200 RSAAVTSEF 6.160 222 TCVVSHPGL 6.000 223 CVVSHPGLL 6.000
325 DSQVTVDVL 6.000 453 EIETQTELL 6.000 80 ALLHSKYGL 6.000 202
AAVTSEFHL 6.000 11 GPEAWLLLL 6.000 245 LAEASVRGL 6.000 356
IAALLFCLL 5.760 352 VVGVIAALL 5.600 36 TSDVVTVVL 5.600 281
NWTRLDGPL 4.800 13 EAWLLLLLL 4.800 355 VIAALLFCL 4.800 9 MWGPEAWLL
4.800 26 TGRCPAGEL 4.400 8 EMWGPEAWL 4.000 294 RVDGDTLGF 4.000 135
GSFQARLRL 4.000 138 QARLRLRVL 4.000 292 GVRVDGDTL 4.000 260
HIGREGAML 4.000 74 EGAQELALL 4.000 188 KGTTSSRSF 4.000 313
YVCHVSNEF 3.696 17 LLLLLLASF 3.600 353 VGVIAALLF 3.000 493
RAKPTGNGI 2.880 236 ITHILHVSF 2.400 477 GIKQAMNHF 2.400 348
ASVVVVGVI 2.100 45 GQDAKLPCF 2.000 129 VSTFPAGSF 2.000 495
KPTGNGIYI 2.000 390 LTLTRENSI 1.800 446 STLTTVREI 1.650 452
REIETQTEL 1.584 363 LLVVVVVLM 1.050 231 LQDQRITHI 1.000 373
RYHRRKAQQ 1.000 1 MPLSLGAEM 0.990 157 PALEEGQGL 0.864 232 QDQRITHIL
0.840 263 REGAMLKCL 0.800 93 AYEGRVEQP 0.750 312 IYVCHVSNE 0.750
279 SYNWTRLDG 0.750 131 TFPAGSFQA 0.750 207 EFHLVPSRS 0.700 360
LFCLLVVVV 0.600 151 PSLNPGPAL 0.600 444 SYSTLTTVR 0.600 393
TRENSIRRL 0.600 159 LEEGQGLTL 0.600 237 THILHVSFL 0.600 53
FYRGDSGEQ 0.550 320 EFSSRDSQV 0.500 195 SFKHSRSAA 0.500 213
SRSMNGQPL 0.480 297 GDTLGFPPL 0.480 250 VRGLEDQNL 0.480 384
QKYEEELTL 0.480 251 RGLEDQNLW 0.432 341 KQVDLVSAS 0.432 73
GEGAQELAL 0.400 277 PPSYNWTRL 0.400 337 EDSGKQVDL 0.400 133
PAGSFQARL 0.400 378 KAQQMTQKY 0.396 28 RCPAGELET 0.330 144
RVLVPPLPS 0.300 214 RSMNGQPLT 0.300 235 RITHILHVS 0.280 58
SGEQVGQVA 0.252 146 LVPPLPSLN 0.216 110 GSVLLRNAV 0.216 217
NGQPLTCVV 0.216 275 QPPPSYNWT 0.216 40 VTVVLGQDA 0.216 349
SVVVVGVIA 0.210 V2-HLA-A24-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 1 GQDAKLPCL 4.000 9
LYRGDSGEQ 0.550 6 LPCLYRGDS 0.100 5 KLPCLYRGD 0.036 2 QDAKLPCLY
0.012 8 CLYRGDSGE 0.010 3 DAKLPCLYR 0.010 4 AKLPCLYRG 0.002 7
PCLYRGDSG 0.002 V7-HLA-A24-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 15; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 7 RSQSEEPEG 0.033 3
HTDPRSQSE 0.014 8 SQSEEPEGR 0.012 2 HHTDPRSQS 0.012 5 DPRSQSEEP
0.011 4 TDPRSQSEE 0.002 1 SHHTDPRSQ 0.001 6 PRSQSEEPE 0.000
V9-HLA-A24-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
19; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 32 IYFYFYFFL 200.000 34 FYFYFFLEM 33.000 92
AFRFIQCLL 28.000 19 LFFFLPFPL 24.000 81 SFTKRKKKL 22.000 17
FFLFFFLPF 18.000 30 FFIYFYFYF 15.000 14 TFNFFLFFF 15.000 91
KAFRFIQCL 9.600 95 FIQCLLLGL 7.200 58 SNPPASASL 7.200 36 FYFFLEMES
6.600 47 VAQAGLELL 6.000 101 LGLLKVRPL 6.000 15 FNFFLFFFL 5.760 63
SASLVAGTL 5.600 96 IQCLLLGLL 4.800 12 RITFNFFLF 4.800 46 YVAQAGLEL
4.400 9 ILLRITFNF 4.200 7 AGILLRITF 3.600 22 FLPFPLVVF 3.000 71
LSVHHCACF 3.000 10 LLRITFNFF 2.880 23 LPFPLVVFF 2.880 28 VVFFIYFYF
2.800 31 FIYFYFYFF 2.400 13 ITFNFFLFF 2.400 88 KLKKAFRFI 2.400 116
SCDCERGYF 2.000 2 RRELLAGIL 1.440 5 LLAGILLRI 1.400 123 YFQGIFMQA
1.260 3 RELLAGILL 1.200 24 PFPLVVFFI 1.050 121 RGYFQGIFM 1.000 38
FFLEMESHY 0.900 21 FFLPFPLVV 0.900 45 HYVAQAGLE 0.750 11 LRITFNFFL
0.600 20 FFFLPFPLV 0.600 29 VFFIYFYFY 0.600 87 KKLKKAFRF 0.600 122
GYFQGIFMQ 0.500 85 RKKKLKKAF 0.480 44 SHYVAQAGL 0.400 93 FRFIQCLLL
0.400 26 PLVVFFIYF 0.360 107 RPLQHQGVN 0.300 25 FPLVVFFIY 0.252 74
HHCACFESF 0.240 50 AGLELLGSS 0.216 69 GTLSVHHCA 0.210 120 ERGYFQGIF
0.200 51 GLELLGSSN 0.180 57 SSNPPASAS 0.180 98 CLLLGLLKV 0.165 94
RFIQCLLLG 0.150 39 FLEMESHYV 0.150 59 NPPASASLV 0.150 64 ASLVAGTLS
0.150 65 SLVAGTLSV 0.150 27 LVVFFIYFY 0.150 8 GILLRITFN 0.150 119
CERGYFQGI 0.144 1 MRRELLAGI 0.144 62 ASASLVAGT 0.120 124 FQGIFMQAA
0.120 6 LAGILLRIT 0.120 109 LQHQGVNSC 0.120 115 NSCDCERGY 0.120 56
GSSNPPASA 0.100 55 LGSSNPPAS 0.100 49 QAGLELLGS 0.100 129 MQAAPWEGT
0.100 111 HQGVNSCDC 0.100 126 GIFMQAAPW 0.100 68 AGTLSVHHC 0.100 75
HCACFESFT 0.100 70 TLSVHHCAC 0.100 54 LLGSSNPPA 0.100 127 IFMQAAPWE
0.075 78 CFESFTKRK 0.075 33 YFYFYFFLE 0.060 16 NFFLFFFLP 0.060 37
YFFLEMESH 0.050 35 YFYFFLEME 0.050 105 KVRPLQHQG 0.029 90 KKAFRFIQC
0.024 84 KRKKKLKKA 0.022 102 GLLKVRPLQ 0.021 106 VRPLQHQGV 0.018 40
LEMESHYVA 0.018 99 LLLGLLKVR 0.018 97 QCLLLGLLK 0.018 53 ELLGSSNPP
0.018 43 ESHYVAQAG 0.017 128 FMQAAPWEG 0.017 113 GVNSCDCER 0.017 77
ACFESFTKR 0.016 V10-HLA-A24-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 21; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 2 RCPAGELGT 0.300 9
GTSDVVTVV 0.168 5 AGELGTSDV 0.150 7 ELGTSDVVT 0.100 8 LGTSDVVTV
0.100 3 CPAGELGTS 0.100 6 GELGTSDVV 0.015 4 PAGELGTSD 0.001 1
GRCPAGELG 0.001 V11-HLA-A24-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 23; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 5 RLRVMVPPL 8.000 8
VMVPPLPSL 7.200 1 QARLRLRVM 0.500 7 RVMVPPLPS 0.300 9 MVPPLPSLN
0.216 3 RLRLRVMVP 0.020 2 ARLRLRVMV 0.018 6 LRVMVPPLP 0.002 4
LRLRVMVPP 0.002 V12-HLA-A24-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 2 MSEEPEGCS 0.180 5
EPEGCSYST 0.150 1 VMSEEPEGC 0.120 9 CSYSTLTTV 0.100 7 EGCSYSTLT
0.100 8 GCSYSTLTT 0.100 6 PEGCSYSTL 0.040 3 SEEPEGCSY 0.018 4
EEPEGCSYS 0.018 V13-HLA-A24-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 27; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 1 SQVTVDVLA 0.210 7
VLADPQEDS 0.120 3 VTVDVLADP 0.025 6 DVLADPQED 0.020 8 LADPQEDSG
0.012 4 TVDVLADPQ 0.012 2 QVTVDVLAD 0.010 9 ADPQEDSGK 0.002 5
VDVLADPQE 0.002 V14-HLA-A24-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 29; each start position is specified; the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 3 SNPPASASL 7.200 8
SASLVAGTL 5.600 2 SSNPPASAS 0.180 9 ASLVAGTLS 0.150 4 NPPASASLV
0.150 7 ASASLVAGT 0.120 1 GSSNPPASA 0.100 5 PPASASLVA 0.010 6
PASASLVAG 0.001
[1031]
27TABLE XVII Start Subsequence Score V1-HLA-A24-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 312
IYVCHVSNEF 277.200 373 RYHRRKAQQM 60.000 409 RSQPEESVGL 14.400 85
KYGLHVSPAY 14.000 144 RVLVPPLPSL 12.000 105 RNPLDGSVLL 12.000 99
EQPPPPRNPL 8.640 351 VVVGVIAALL 8.400 361 FCLLVVVVVL 8.400 350
VVVVGVIAAL 8.400 501 IYINGRGHLV 7.500 158 ALEEGQGLTL 7.200 11
GPEAWLLLLL 7.200 10 WGPEAWLLLL 7.200 354 GVIAALLFCL 7.200 35
ETSDVVTVVL 6.720 41 TVVLGQDAKL 6.600 291 SGVRVDGDTL 6.000 79
LALLHSKYGL 6.000 439 EPEGRSYSTL 6.000 72 AGEGAQELAL 6.000 222
TCVVSHPGLL 6.000 355 VIAALLFCLL 5.760 231 LQDQRITHIL 5.600 53
FYRGDSGEQV 5.000 249 SVRGLEDQNL 4.800 244 FLAEASVRGL 4.800 13
EAWLLLLLLL 4.800 392 LTRENSIRRL 4.800 280 YNWTRLDGPL 4.800 235
RITHILHVSF 4.800 9 MWGPEAWLLL 4.800 296 DGDTLGFPPL 4.800 156
GPALEEGQGL 4.800 25 FTGRCPAGEL 4.400 381 QMTQKYEEEL 4.400 132
FPAGSFQARL 4.000 236 ITHILHVSFL 4.000 221 LTCVVSHPGL 4.000 128
RVSTFPAGSF 4.000 137 FQARLRLRVL 4.000 201 SAAVTSEFHL 4.000 134
AGSFQARLRL 4.000 500 GIYINGRGHL 4.000 8 EMWGPEAWLL 4.000 383
TQKYEEELTL 4.000 150 LPSLNPGPAL 4.000 16 LLLLLLLASF 3.600 44
LGQDAKLPCF 3.600 476 EGIKQAMNHF 3.600 207 EFHLVPSRSM 2.500 385
KYEEELTLTR 2.160 352 VVGVIAALLF 2.000 252 GLEDQNLWHI 1.800 230
LLQDQRITHI 1.800 452 REIETQTELL 1.440 347 SASVVVVGVI 1.400 93
AYEGRVEQPP 1.260 389 ELTLTRENSI 1.200 227 HPGLLQDQRI 1.200 445
YSTLTTVREI 1.100 124 EYECRVSTFP 1.050 362 CLLVVVVVLM 1.050 473
DQDEGIKQAM 1.008 301 GFPPLTTEHS 0.900 136 SFQARLRLRV 0.900 324
RDSQVTVDVL 0.800 279 SYNWTRLDGP 0.750 141 LRLRVLVPPL 0.720 360
LFCLLVVVVV 0.700 451 VREIETQTEL 0.660 262 GREGAMLKCL 0.600 259
WHIGREGAML 0.600 320 EFSSRDSQVT 0.600 276 PPPSYNWTRL 0.600 7
AEMWGPEAWL 0.600 70 VDAGEGAQEL 0.528 341 KQVDLVSASV 0.504 258
LWHIGREGAM 0.500 195 SFKHSRSAAV 0.500 444 SYSTLTTVRE 0.500 418
LRAEGHPDSL 0.480 212 PSRSMNGQPL 0.480 336 QEDSGKQVDL 0.400 483
NHFVQENGTL 0.400 73 GEGAQELALL 0.400 293 VRVDGDTLGF 0.360 199
SRSAAVTSEF 0.308 97 RVEQPPPPRN 0.300 214 RSMNGQPLTC 0.300 28
RCPAGELETS 0.300 49 KLPCFYRGDS 0.300 411 QPEESVGLRA 0.252 284
RLDGPLPSGV 0.240 493 RAKPTGNGIY 0.240 123 GEYECRVSTF 0.240 145
VLVPPLPSLN 0.216 274 GQPPPSYNWT 0.216 363 LLVVVVVLMS 0.210 348
ASVVVVGVIA 0.210 V2-HLA-A24-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 1 LGQDAKLPCL 7.200 10
LYRGDSGEQV 5.000 6 KLPCLYRGDS 0.300 2 GQDAKLPCLY 0.120 9 CLYRGDSGEQ
0.011 7 LPCLYRGDSG 0.010 4 DAKLPCLYRG 0.010 5 AKLPCLYRGD 0.002 8
PCLYRGDSGE 0.002 3 QDAKLPCLYR 0.001 V7-HLA-A24-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 9
SQSEEPEGRS 0.120 8 RSQSEEPEGR 0.030 4 HTDPRSQSEE 0.013 6 DPRSQSEEPE
0.010 1 HSHHTDPRSQ 0.010 2 SHHTDPRSQS 0.010 5 TDPRSQSEEP 0.002 3
HHTDPRSQSE 0.001 7 PRSQSEEPEG 0.000 V9-HLA-A24-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 19; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 45
HYVAQAGLEL 330.000 94 RFIQCLLLGL 72.000 14 TFNFFLFFFL 43.200 92
AFRFIQCLLL 20.000 30 FFIYFYFYFF 18.000 21 FFLPFPLVVF 18.000 16
NFFLFFFLPF 12.000 91 KAFRFIQCLL 11.200 29 VFFIYFYFYF 10.000 122
GYFQGIFMQA 8.400 57 SSNPPASASL 7.200 95 FIQCLLLGLL 7.200 62
ASASLVAGTL 5.600 12 RITFNFFLFF 4.800 18 FLFFFLPFPL 4.800 80
ESFTKRKKKL 4.400 9 ILLRITFNFF 4.320 8 GILLRITFNF 4.200 27
LVVFFIYFYF 4.200 31 FIYFYFYFFL 4.000 10 LLRITFNFFL 4.000 46
YVAQAGLELL 4.000 100 LLGLLKVRPL 4.000 43 ESHYVAQAGL 4.000 25
FPLVVFFIYF 3.600 22 FLPFPLVVFF 3.600 33 YFYFYFFLEM 3.300 115
NSCDCERGYF 2.400 6 LAGILLRITF 2.400 118 DCERGYFQGI 2.160 4
ELLAGILLRI 2.100 13 ITFNFFLFFF 2.000 70 TLSVHHCACF 2.000 23
LPFPLVVFFI 1.680 2 RRELLAGILL 1.200 90 KKAFRFIQCL 0.960 123
YFQGIFMQAA 0.900 38 FFLEMESHYV 0.900 35 YFYFFLEMES 0.660 32
IYFYFYFFLE 0.600 19 LFFFLPFPLV 0.600 1 MRRELLAGIL 0.576 34
FYFYFFLEME 0.500 37 YFFLEMESHY 0.500 20 FFFLPFPLVV 0.500 36
FYFFLEMESH 0.500 84 KRKKKLKKAF 0.480 86 KKKLKKAFRF 0.400 11
LRITFNFFLF 0.360 87 KKLKKAFRFI 0.360 107 RPLQHQGVNS 0.300 105
KVRPLQHQGV 0.288 73 VHHCACFESF 0.240 50 AGLELLGSSN 0.216 119
CERGYFQGIF 0.200 58 SNPPASASLV 0.180 97 QCLLLGLLKV 0.165 53
ELLGSSNPPA 0.150 64 ASLVAGTLSV 0.150 39 FLEMESHYVA 0.150 128
FMQAAPWEGT 0.150 125 QGIFMQAAPW 0.150 59 NPPASASLVA 0.150 69
GTLSVHHCAC 0.150 7 AGILLRITFN 0.150 41 EMESHYVAQA 0.150 68
AGTLSVHHCA 0.140 24 PFPLVVFFIY 0.126 28 VVFFIYFYFY 0.120 49
QAGLELLGSS 0.120 5 LLAGILLRIT 0.120 72 SVHHCACFES 0.110 55
LGSSNPPASA 0.100 114 VNSCDCERGY 0.100 54 LLGSSNPPAS 0.100 48
AQAGLELLGS 0.100 56 GSSNPPASAS 0.100 63 SASLVAGTLS 0.100 67
VAGTLSVHHC 0.100 78 CFESFTKRKK 0.083 127 IFMQAAPWEG 0.083 17
FFLFFFLPFP 0.075 120 ERGYFQGIFM 0.050 81 SFTKRKKKLK 0.050 101
LGLLKVRPLQ 0.021 121 RGYFQGIFMQ 0.020 88 KLKKAFRFIQ 0.020 108
PLQHQGVNSC 0.018 99 LLLGLLKVRP 0.018 98 CLLLGLLKVR 0.018 47
VAQAGLELLG 0.018 112 QGVNSCDCER 0.017 51 GLELLGSSNP 0.015 110
QHQGVNSCDC 0.015 26 PLVVFFIYFY 0.015 102 GLLKVRPLQH 0.015 71
LSVHHCACFE 0.015 106 VRPLQHQGVN 0.015 65 SLVAGTLSVH 0.015 113
GVNSCDCERG 0.015 V10-HLA-A24-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 21; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 10 GTSDVVTVVL 6.720 3
RCPAGELGTS 0.300 6 AGELGTSDVV 0.150 9 LGTSDVVTVV 0.140 8 ELGTSDVVTV
0.100 7 GELGTSDVVT 0.015 4 CPAGELGTSD 0.012 5 PAGELGTSDV 0.012 2
GRCPAGELGT 0.012 1 TGRCPAGELG 0.010 V11-HLA-A24-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 23; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 8
RVMVPPLPSL 12.000 5 LRLRVMVPPL 0.600 1 FQARLRLRVM 0.500 9
VMVPPLPSLN 0.216 2 QARLRLRVMV 0.120 6 RLRVMVPPLP 0.028 4 RLRLRVMVPP
0.028 10 MVPPLPSLNP 0.018 7 LRVMVPPLPS 0.015 3 ARLRLRVMVP 0.002
V12-HLA-A24-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 25; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 6 EPEGCSYSTL 6.000 11 SYSTLTTVRE 0.500 3
MSEEPEGCSY 0.180 1 SVMSEEPEGC 0.150 2 VMSEEPEGCS 0.120 8 EGCSYSTLTT
0.100 9 GCSYSTLTTV 0.100 5 EEPEGCSYST 0.018 4 SEEPEGCSYS 0.018 10
CSYSTLTTVR 0.012 7 PEGCSYSTLT 0.001 V13-HLA-A24-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 27; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 1
DSQVTVDVLA 0.210 7 DVLADPQEDS 0.150 4 VTVDVLADPQ 0.022 2 SQVTVDVLAD
0.015 3 QVTVDVLADP 0.014 8 VLADPQEDSG 0.012 9 LADPQEDSGK 0.012 5
TVDVLADPQE 0.010 6 VDVLADPQED 0.002 10 ADPQEDSGKQ 0.002
V14-HLA-A24-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 3 SSNPPASASL 7.200 8 ASASLVAGTL 5.600 4
SNPPASASLV 0.180 10 ASLVAGTLSV 0.150 5 NPPASASLVA 0.150 9
SASLVAGTLS 0.100 1 LGSSNPPASA 0.100 2 GSSNPPASAS 0.100 7 PASASLVAGT
0.012 6 PPASASLVAG 0.001
[1032]
28TABLE XVIII Start Subsequence Score V1-HLA-B7-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 292
GVRVDGDTL 200.000 100 QPPPPRNPL 180.000 138 QARLRLRVL 120.000 106
NPLDGSVLL 80.000 26 TGRCPAGEL 60.000 142 RLRVLVPPL 40.000 202
AAVTSEFHL 36.000 11 GPEAWLLLL 24.000 42 VVLGQDAKL 20.000 1
MPLSLGAEM 20.000 351 VVVGVIAAL 20.000 352 VVGVIAALL 20.000 223
CVVSHPGLL 20.000 13 EAWLLLLLL 12.000 71 DAGEGAQEL 12.000 80
ALLHSKYGL 12.000 356 IAALLFCLL 12.000 277 PPSYNWTRL 8.000 495
KPTGNGIYI 8.000 135 GSFQARLRL 6.000 8 EMWGPEAWL 6.000 145 VLVPPLPSL
6.000 450 TVREIETQT 5.000 222 TCVVSHPGL 4.000 325 DSQVTVDVL 4.000
287 GPLPSGVRV 4.000 362 CLLVVVVVL 4.000 10 WGPEAWLLL 4.000 260
HIGREGAML 4.000 410 SQPEESVGL 4.000 355 VIAALLFCL 4.000 105
RNPLDGSVL 4.000 74 EGAQELALL 4.000 382 MTQKYEEEL 4.000 407
DPRSQPEES 4.000 419 RAEGHPDSL 3.600 245 LAEASVRGL 3.600 203
AVTSEFHLV 3.000 275 QPPPSYNWT 2.000 322 SSRDSQVTV 2.000 150
LPSLNPGPA 2.000 357 AALLFCLLV 1.800 371 MSRYHRRKA 1.500 133
PAGSFQARL 1.200 493 RAKPTGNGI 1.200 14 AWLLLLLLL 1.200 36 TSDVVTVVL
1.200 453 EIETQTELL 1.200 157 PALEEGQGL 1.200 348 ASVVVVGVI 1.200
249 SVRGLEDQN 1.000 374 YHRRKAQQM 1.000 441 EGRSYSTLT 1.000 363
LLVVVVVLM 1.000 345 LVSASVVVV 1.000 126 ECRVSTFPA 1.000 64
QVAWARVDA 0.750 103 PPRNPLDGS 0.600 358 ALLFCLLVV 0.600 178
APSVTWDTE 0.600 501 IYINGRGHL 0.600 151 PSLNPGPAL 0.600 50
LPCFYRGDS 0.600 439 EPEGRSYST 0.600 347 SASVVVVGV 0.600 349
SVVVVGVIA 0.500 350 VVVVGVIAA 0.500 354 GVIAALLFC 0.500 23
ASFTGRCPA 0.450 29 CPAGELETS 0.400 446 STLTTVREI 0.400 297
GDTLGFPPL 0.400 232 QDQRITHIL 0.400 263 REGAMLKCL 0.400 281
NWTRLDGPL 0.400 390 LTLTRENSI 0.400 484 HFVQENGTL 0.400 452
REIETQTEL 0.400 384 QKYEEELTL 0.400 302 FPPLTTEHS 0.400 237
THILHVSFL 0.400 250 VRGLEDQNL 0.400 73 GEGAQELAL 0.400 9 MWGPEAWLL
0.400 213 SRSMNGQPL 0.400 337 EDSGKQVDL 0.400 289 LPSGVRVDG 0.300
110 GSVLLRNAV 0.300 117 AVQADEGEY 0.300 216 MNGQPLTCV 0.300 147
VPPLPSLNP 0.300 137 FQARLRLRV 0.300 67 WARVDAGEG 0.300 342
QVDLVSASV 0.300 462 SPGSGRAEE 0.300 214 RSMNGQPLT 0.300 211
VPSRSMNGQ 0.200 217 NGQPLTCVV 0.200 35 ETSDVVTVV 0.200 154
NPGPALEEG 0.200 V2-HLA-B7-9mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 5; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 1 GQDAKLPCL 1.200 6 LPCLYRGDS 0.600
3 DAKLPCLYR 0.045 8 CLYRGDSGE 0.010 9 LYRGDSGEQ 0.010 5 KLPCLYRGD
0.010 4 AKLPCLYRG 0.003 2 QDAKLPCLY 0.002 7 PCLYRGDSG 0.001
V7-HLA-B7-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
15; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 5 DPRSQSEEP 2.000 7 RSQSEEPEG 0.010 8
SQSEEPEGR 0.010 2 HHTDPRSQS 0.005 3 HTDPRSQSE 0.003 4 TDPRSQSEE
0.001 1 SHHTDPRSQ 0.001 6 PRSQSEEPE 0.000 V9 HLA-B7-9mers-J91P4D12B
Each peptide is a portion of SEQ ID NO: 19; each start position is
specified,the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 46
YVAQAGLEL 20.000 92 AFRFIQCLL 12.000 91 KAFRFIQCL 12.000 63
SASLVAGTL 12.000 47 VAQAGLELL 12.000 59 NPPASASLV 4.000 95
FIQCLLLGL 4.000 96 IQCLLLGLL 4.000 15 FNFFLFFFL 4.000 101 LGLLKVRPL
4.000 58 SNPPASASL 4.000 121 RGYFQGIFM 1.000 105 KVRPLQHQG 0.500 5
LLAGILLRI 0.400 107 RPLQHQGVN 0.400 23 LPFPLVVFF 0.400 88 KLKKAFRFI
0.400 44 SHYVAQAGL 0.400 19 LFFFLPFPL 0.400 81 SFTKRKKKL 0.400 25
FPLVVFFIY 0.400 32 IYFYFYFFL 0.400 3 RELLAGILL 0.400 119 CERGYFQGI
0.400 93 FRFIQCLLL 0.400 1 MRRELLAGI 0.400 11 LRITFNFFL 0.400 6
LAGILLRIT 0.300 62 ASASLVAGT 0.300 68 AGTLSVHHC 0.300 60 PPASASLVA
0.200 10 LLRITFNFF 0.200 98 CLLLGLLKV 0.200 65 SLVAGTLSV 0.200 56
GSSNPPASA 0.150 129 MQAAPWEGT 0.150 2 RRELLAGIL 0.120 70 TLSVHHCAC
0.100 109 LQHQGVNSC 0.100 69 GTLSVHHCA 0.100 28 VVFFIYFYF 0.100 34
FYFYFFLEM 0.100 54 LLGSSNPPA 0.100 27 LVVFFIYFY 0.100 124 FQGIFMQAA
0.100 75 HCACFESFT 0.100 111 HQGVNSCDC 0.100 7 AGILLRITF 0.090 49
QAGLELLGS 0.060 64 ASLVAGTLS 0.060 50 AGLELLGSS 0.060 39 FLEMESHYV
0.060 66 LVAGTLSVH 0.050 72 SVHHCACFE 0.050 113 GVNSCDCER 0.050 48
AQAGLELLG 0.030 40 LEMESHYVA 0.030 77 ACFESFTKR 0.030 67 VAGTLSVHH
0.030 22 FLPFPLVVF 0.030 76 CACFESFTK 0.030 20 FFFLPFPLV 0.030 57
SSNPPASAS 0.030 71 LSVHHCACF 0.020 55 LGSSNPPAS 0.020 106 VRPLQHQGV
0.020 21 FFLPFPLVV 0.020 12 RITFNFFLF 0.020 9 ILLRITFNF 0.020 115
NSCDCERGY 0.020 13 ITFNFFLFF 0.020 126 GIFMQAAPW 0.020 8 GILLRITFN
0.020 31 FIYFYFYFF 0.020 102 GLLKVRPLQ 0.015 80 ESFTKRKKK 0.015 125
QGIFMQAAP 0.010 128 FMQAAPWEG 0.010 18 FLFFFLPFP 0.010 97 QCLLLGLLK
0.010 100 LLGLLKVRP 0.010 1231 YFQGIFMQA 0.010 103 LLKVRPLQH 0.010
83 TKRKKKLKK 0.010 90 KKAFRFIQC 0.010 112 QGVNSCDCE 0.010 42
MESHYVAQA 0.010 4 ELLAGILLR 0.010 82 FTKRKKKLK 0.010 43 ESHYVAQAG
0.010 84 KRKKKLKKA 0.010 99 LLLGLLKVR 0.010 53 ELLGSSNPP 0.010 114
VNSCDCERG 0.010 116 SCDCERGYF 0.009 51 GLELLGSSN 0.006 24 PFPLVVFFI
0.004 127 IFMQAAPWE 0.003 61 PASASLVAG 0.003 118 DCERGYFQG 0.003
V11-HLA-B7-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
23; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 5 RLRVMVPPL 40.000 1 QARLRLRVM 30.000 8
VMVPPLPSL 6.000 7 RVMVPPLPS 0.450 9 MVPPLPSLN 0.100 3 RLRLRVMVP
0.100 2 ARLRLRVMV 0.090 6 LRVMVPPLP 0.001 4 LRLRVMVPP 0.001
V12-HLA-B7-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 5 EPEGCSYST 0.600 9 CSYSTLTTV 0.200 7
EGCSYSTLT 0.100 1 VMSEEPEGC 0.100 8 GCSYSTLTT 0.100 6 PEGCSYSTL
0.040 2 MSEEPEGCS 0.009 4 EEPEGCSYS 0.002 3 SEEPEGCSY 0.001
V13-HLA-B7-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
27; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 1 SQVTVDVLA 0.100 6 DVLADPQED 0.050 2
QVTVDVLAD 0.050 7 VLADPQEDS 0.030 4 TVDVLADPQ 0.015 3 VTVDVLADP
0.010 8 LADPQEDSG 0.009 9 ADPQEDSGK 0.003 5 VDVLADPQE 0.001
V14-HLA-B7-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
29; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 8 SASLVAGTL 12.000 4 NPPASASLV 4.000 3
SNPPASASL 4.000 7 ASASLVAGT 0.300 5 PPASASLVA 0.200 1 GSSNPPASA
0.150 9 ASLVAGTLS 0.060 2 SSNPPASAS 0.030 6 PASASLVAG 0.003
[1033]
29TABLE XIX Start Sequence Score V1-HLA-B7-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 249
SVRGLEDQNL 200.000 150 LPSLNPGPAL 120.000 156 GPALEEGQGL 80.000 132
FPAGSFQARL 80.000 407 DPRSQPEESV 60.000 392 LTRENSIRRL 40.000 144
RVLVPPLPSL 30.000 11 GPEAWLLLLL 24.000 439 EPEGRSYSTL 24.000 350
VVVVGVIAAL 20.000 351 VVVGVIAALL 20.000 354 GVIAALLFCL 20.000 41
TVVLGQDAKL 20.000 134 AGSFQARLRL 18.000 178 APSVTWDTEV 12.000 13
EAWLLLLLLL 12.000 201 SAAVTSEFHL 12.000 79 LALLHSKYGL 12.000 99
EQPPPPRNPL 9.000 138 QARLRLRVLV 9.000 276 PPPSYNWTRL 8.000 227
HPGLLQDQRI 8.000 500 GIYINGRGHL 6.000 25 FTGRCPAGEL 6.000 7
AEMWGPEAWL 5.400 409 RSQPEESVGL 4.000 103 PPRNPLDGSV 4.000 244
FLAEASVRGL 4.000 8 EMWGPEAWLL 4.000 383 TQKYEEELTL 4.000 137
FQARLRLRVL 4.000 236 ITHILHVSFL 4.000 291 SGVRVDGDTL 4.000 334
DPQEDSGKQV 4.000 10 WGPEAWLLLL 4.000 222 TCVVSHPGLL 4.000 212
PSRSMNGQPL 4.000 280 YNWTRLDGPL 4.000 221 LTCVVSHPGL 4.000 355
VIAALLFCLL 4.000 381 QMTQKYEEEL 4.000 35 ETSDVVTVVL 4.000 361
FCLLVVVVVL 4.000 105 RNPLDGSVLL 4.000 158 ALEEGQGLTL 3.600 72
AGEGAQELAL 3.600 67 WARVDAGEGA 3.000 176 SPAPSVTWDT 2.000 233
DQRITHILHV 2.000 202 AAVTSEFHLV 1.800 357 AALLFCLLVV 1.800 231
LQDQRITHIL 1.200 347 SASVVVVGVI 1.200 296 DGDTLGFPPL 1.200 261
IGREGAMLKC 1.000 397 SIRRLHSHHT 1.000 61 QVGQVAWARV 1.000 441
EGRSYSTLTT 1.000 89 HVSPAYEGRV 1.000 362 CLLVVVVVLM 1.000 241
HVSFLAEASV 1.000 303 PPLTTEHSGI 0.800 411 QPEESVGLRA 0.600 356
IAALLFCLLV 0.600 358 ALLFCLLVVV 0.600 349 SVVVVGVIAA 0.500 485
FVQENGTLRA 0.500 450 TVREIETQTE 0.500 292 GVRVDGDTLG 0.500 39
VVTVVLGQDA 0.500 111 SVLLRNAVQA 0.500 22 LASFTGRCPA 0.450 452
REIETQTELL 0.400 324 RDSQVTVDVL 0.400 70 VDAGEGAQEL 0.400 1
MPLSLGAEMW 0.400 389 ELTLTRENSI 0.400 259 WHIGREGAML 0.400 73
GEGAQELALL 0.400 495 KPTGNGIYIN 0.400 418 LRAEGHPDSL 0.400 9
MWGPEAWLLL 0.400 483 NHFVQENGTL 0.400 230 LLQDQRITHI 0.400 141
LRLRVLVPPL 0.400 445 YSTLTTVREI 0.400 342 QVDLVSASVV 0.300 215
SMNGQPLTCV 0.300 71 DAGEGAQELA 0.300 214 RSMNGQPLTC 0.300 348
ASVVVVGVIA 0.300 109 DGSVLLRNAV 0.300 169 ASCTAEGSPA 0.300 91
SPAYEGRVEQ 0.300 473 DQDEGIKQAM 0.300 172 TAEGSPAPSV 0.270 289
LPSGVRVDGD 0.200 81 LLHSKYGLHV 0.200 417 GLRAEGHPDS 0.200 321
FSSRDSQVTV 0.200 V2-HLA-B7-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 1 LGQDAKLPCL 4.000 7
LPCLYRGDSG 0.200 10 LYRGDSGEQV 0.200 4 DAKLPCLYRG 0.030 6
KLPCLYRGDS 0.030 9 CLYRGDSGEQ 0.010 2 GQDAKLPCLY 0.006 5 AKLPCLYRGD
0.003 3 QDAKLPCLYR 0.002 8 PCLYRGDSGE 0.001
V7-HLA-B7-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
15; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 6 DPRSQSEEPE 2.000 9 SQSEEPEGRS 0.030 8
RSQSEEPEGR 0.010 1 HSHHTDPRSQ 0.010 2 SHHTDPRSQS 0.005 4 HTDPRSQSEE
0.003 3 HHTDPRSQSE 0.001 5 TDPRSQSEEP 0.001 7 PRSQSEEPEG 0.000
V9-HLA-B7-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
19; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 10 LLRITFNFFL 40.000 46 YVAQAGLELL 20.000 92
AFRFIQCLLL 12.000 91 KAFRFIQCLL 12.000 62 ASASLVAGTL 12.000 105
KVRPLQHQGV 10.000 23 LPFPLVVFFI 8.000 100 LLGLLKVRPL 4.000 31
FIYFYFYFFL 4.000 1 MRRELLAGIL 4.000 95 FIQCLLLGLL 4.000 57
SSNPPASASL 4.000 80 ESFTKRKKKL 4.000 18 FLFFFLPFPL 4.000 43
ESHYVAQAGL 4.000 59 NPPASASLVA 2.000 64 ASLVAGTLSV 0.600 4
ELLAGILLRI 0.400 107 RPLQHQGVNS 0.400 14 TFNFFLFFFL 0.400 25
FPLVVFIYF 0.400 94 RFIQCLLLGL 0.400 45 HYVAQAGLEL 0.400 90
KKAFRFIQCL 0.400 67 VAGTLSVHHC 0.300 68 AGTLSVHHCA 0.300 97
QCLLLGLLKV 0.200 58 SNPPASASLV 0.200 128 FMQMPWEGT 0.150 55
LGSSNPPASA 0.150 2 RRELLAGILL 0.120 118 DCERGYFQGI 0.120 33
YFYFYFFLEM 0.100 28 VVFFIYFYFY 0.100 53 ELLGSSNPPA 0.100 72
SVHHCACFES 0.100 83 TKRKKKLKKA 0.100 5 LLAGILLRIT 0.100 69
GTLSVHHCAC 0.100 27 LVVFFIYFYF 0.100 120 ERGYFQGIFM 0.100 6
LAGILLRITF 0.090 63 SASLVAGTLS 0.060 48 AQAGLELLGS 0.060 7
AGILLRITFN 0.060 50 AGLELLGSSN 0.060 49 QAGLELLGSS 0.060 113
GVNSCDCERG 0.050 66 LVAGTLSVHH 0.050 87 KKLKKAFRFI 0.040 115
NSCDCERGYF 0.030 47 VAQAGLELLG 0.030 61 PASASLVAGT 0.030 76
CACFESFTKR 0.030 56 GSSNPPASAS 0.030 19 LFFFLPFPLV 0.030 77
ACFESFTKRK 0.030 39 FLEMESHYVA 0.030 41 EMESHYVAQA 0.030 38
FFLEMESHYV 0.020 22 FLPFPLVVFF 0.020 119 CERGYFQGIF 0.020 9
ILLRITFNFF 0.020 70 TLSVHHCACF 0.020 125 QGIFMQAAPW 0.020 60
PPASASLVAG 0.020 8 GILLRITFNF 0.020 12 RITFNFFLFF 0.020 114
VNSCDCERGY 0.020 54 LLGSSNPPAS 0.020 13 ITFNFFLFFF 0.020 20
FFFLPFPLVV 0.020 101 LGLLKVRPLQ 0.015 103 LLKVRPLQHQ 0.015 88
KLKKAFRFIQ 0.015 108 PLQHQGVNSC 0.010 96 IQCLLLGLLK 0.010 89
LKKAFRFIQC 0.010 75 HCACFESFTK 0.010 82 FTKRKKKLKK 0.010 102
GLLKVRPLQH 0.010 121 RGYFQGIFMQ 0.010 15 FNFFLFFFLP 0.010 65
SLVAGTLSVH 0.010 98 CLLLGLLKVR 0.010 109 LQHQGVNSCD 0.010 110
QHQGVNSCDC 0.010 122 GYFQGIFMQA 0.010 111 HQGVNSCDCE 0.010 71
LSVHHCACFE 0.010 126 GIFMQAAPWE 0.010 99 LLLGLLKVRP 0.010 123
YFQGIFMQAA 0.010 124 FQGIFMQAAP 0.010 74 HHCACFESFT 0.010 112
QGVNSCDCER 0.010 21 FFLPFPLVVF 0.003 127 IFMQAAPWEG 0.003 40
LEMESHYVAQ 0.003 116 SCDCERGYFQ 0.003 V10-HLA-B7-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 21; each start positions
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 10
GTSDVVTVVL 4.000 9 LGTSDVVTVV 0.200 8 ELGTSDVVTV 0.200 4 CPAGELGTSD
0.200 6 AGELGTSDVV 0.180 1 TGRCPAGELG 0.100 5 PAGELGTSDV 0.060 3
RCPAGELGTS 0.020 2 GRCPAGELGT 0.010 7 GELGTSDVVT 0.010
V11-HLA-B7-10mers-191P4D12B Each peptide is a portion of SEQD NO:
23; each start positions specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 8 RVMVPPLPSL 90.000 2 QARLRLRVMV 9.000 1
FQARLRLRVM 1.000 5 LRLRVMVPPL 0.400 6 RLRVMVPPLP 0.100 4 RLRLRVMVPP
0.100 10 MVPPLPSLNP 0.075 9 VMVPPLPSLN 0.020 7 LRVMVPPLPS 0.003 3
ARLRLRVMVP 0.003 V12-HLA-B7-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 6 EPEGCSYSTL 24.000 1
SVMSEEPEGC 1.500 9 GCSYSTLTTV 0.200 8 EGCSYSTLTT 0.100 2 VMSEEPEGCS
0.030 5 EEPEGCSYST 0.010 10 CSYSTLTTVR 0.010 3 MSEEPEGCSY 0.006 11
SYSTLTTVRE 0.001 7 PEGCSYSTLT 0.001 4 SEEPEGCSYS 0.001
V13-HLA-B7-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
27; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 7 DVLADPQEDS 0.150 1 DSQVTVDVLA 0.100 3
QVTVDVLADP 0.050 5 TVDVLADPQE 0.015 4 VTVDVLADPQ 0.010 2 SQVTVDVLAD
0.010 8 VLADPQEDSG 0.010 9 LADPQEDSGK 0.009 10 ADPQEDSGKQ 0.003 6
VDVLADPQED 0.001 V14-HLA-B7-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 29; each start position is specified, the
length; of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 8 ASASLVAGTL 12.000 3
SSNPPASASL 4.000 5 NPPASASLVA 2.000 10 ASLVAGTLSV 0.600 4
SNPPASASLV 0.200 1 LGSSNPPASA 0.150 9 SASLVAGTLS 0.060 7 PASASLVAGT
0.030 2 GSSNPPASAS 0.030 6 PPASASLVAG 0.020
[1034]
30TABLE XX Start Subsequence Score V1-HLA-B3501-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 1
MPLSLGAEM 40.000 106 NPLDGSVLL 40.000 100 QPPPPRNPL 20.000 495
KPTGNGIYI 16.000 378 KAQQMTQKY 12.000 200 RSAAVTSEF 10.000 138
QARLRLRVL 9.000 493 RAKPTGNGI 7.200 322 SSRDSQVTV 6.000 407
DPRSQPEES 6.000 142 RLRVLVPPL 6.000 11 GPEAWLLLL 6.000 71 DAGEGAQEL
6.000 129 VSTFPAGSF 5.000 325 DSQVTVDVL 5.000 135 GSFQARLRL 5.000
292 GVRVDGDTL 4.500 305 LTTEHSGIY 4.000 287 GPLPSGVRV 4.000 117
AVQADEGEY 3.000 26 TGRCPAGEL 3.000 202 AAVTSEFHL 3.000 251
RGLEDQNLW 3.000 29 CPAGELETS 3.000 105 RNPLDGSVL 3.000 13 EAWLLLLLL
3.000 356 IAALLFCLL 3.000 410 SQPEESVGL 3.000 477 GIKQAMNHF 3.000
175 GSPAPSVTW 2.500 366 VVVVLMSRY 2.000 275 QPPPSYNWT 2.000 50
LPCFYRGDS 2.000 150 LPSLNPGPA 2.000 78 ELALLHSKY 2.000 348
ASVVVVGVI 2.000 363 LLVVVVVLM 2.000 57 DSGEQVGQV 2.000 86 YGLHVSPAY
2.000 10 WGPEAWLLL 2.000 188 KGTTSSRSF 2.000 302 FPPLTTEHS 2.000
277 PPSYNWTRL 2.000 443 RSYSTLTTV 2.000 419 RAEGHPDSL 1.800 74
EGAQELALL 1.500 260 HIGREGAML 1.500 36 TSDVVTVVL 1.500 83 HSKYGLHVS
1.500 198 HSRSAAVTS 1.500 371 MSRYHRRKA 1.500 8 EMWGPEAWL 1.000 222
TCVVSHPGL 1.000 17 LLLLLLASF 1.000 80 ALLHSKYGL 1.000 355 VIAALLFCL
1.000 42 VVLGQDAKL 1.000 242 VSFLAEASV 1.000 214 RSMNGQPLT 1.000
351 VVVGVIAAL 1.000 382 MTQKYEEEL 1.000 313 YVCHVSNEF 1.000 309
HSGIYVCHV 1.000 353 VGVIAALLF 1.000 352 VVGVIAALL 1.000 362
CLLVVVVVL 1.000 90 VSPAYEGRV 1.000 194 RSFKHSRSA 1.000 145
VLVPPLPSL 1.000 223 CVVSHPGLL 1.000 338 DSGKQVDLV 1.000 110
GSVLLRNAV 1.000 236 ITHILHVSF 1.000 157 PALEEGQGL 0.900 294
RVDGDTLGF 0.900 245 LAEASVRGL 0.900 321 FSSRDSQVT 0.750 425
DSLKDNSSC 0.750 347 SASVVVVGV 0.600 357 AALLFCLLV 0.600 439
EPEGRSYST 0.600 450 TVREIETQT 0.600 334 DPQEDSGKQ 0.600 423
HPDSLKDNS 0.600 103 PPRNPLDGS 0.600 426 SLKDNSSCS 0.600 374
YHRRKAQQM 0.600 23 ASFTGRCPA 0.500 274 GQPPPSYNW 0.500 191
TSSRSFKHS 0.500 151 PSLNPGPAL 0.500 402 HSHHTDPRS 0.500 383
TQKYEEELT 0.450 428 KDNSSCSVM 0.400 446 STLTTVREI 0.400 390
LTLTRENSI 0.400 35 ETSDVVTVV 0.400 341 KQVDLVSAS 0.400 452
REIETQTEL 0.400 491 TLRAKPTGN 0.300 V2-HLA-B3501-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 5; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 6
LPCLYRGDS 2.000 1 GQDAKLPCL 0.300 2 QDAKLPCLY 0.200 3 DAKLPCLYR
0.090 5 KLPCLYRGD 0.020 8 CLYRGDSGE 0.010 9 LYRGDSGEQ 0.005 4
AKLPCLYRG 0.001 7 PCLYRGDSG 0.001 V7-HLA-B3501-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 5
DPRSQSEEP 0.600 7 RSQSEEPEG 0.150 8 SQSEEPEGR 0.030 2 HHTDPRSQS
0.020 3 HTDPRSQSE 0.003 1 SHHTDPRSQ 0.002 4 TDPRSQSEE 0.001 6
PRSQSEEPE 0.000 V9-HLA-B3501-9mers-191P- 4D12B Each peptide is a
portion of SEQ ID NO: 19; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 25 FPLVVFFIY 40.000 23
LPFPLVVFF 20.000 115 NSCDCERGY 20.000 91 KAFRFIQCL 6.000 71
LSVHHCACF 5.000 107 RPLQHQGVN 4.000 59 NPPASASLV 4.000 121
RGYFQGIFM 4.000 10 LLRITFNFF 3.000 47 VAQAGLELL 3.000 63 SASLVAGTL
3.000 88 KLKKAFRFI 2.400 27 LVVFFIYFY 2.000 12 RITFNFFLF 2.000 46
YVAQAGLEL 1.000 15 FNFFLFFFL 1.000 7 AGILLRITF 1.000 22 FLPFPLVVF
1.000 95 FIQCLLLGL 1.000 101 LGLLKVRPL 1.000 31 FIYFYFYFF 1.000 58
SNPPASASL 1.000 28 VVFFIYFYF 1.000 9 ILLRITFNF 1.000 13 ITFNFFLFF
1.000 96 IQCLLLGLL 1.000 85 RKKKLKKAF 0.600 126 GIFMQAAPW 0.500 57
SSNPPASAS 0.500 62 ASASLVAGT 0.500 64 ASLVAGTLS 0.500 56 GSSNPPASA
0.500 116 SCDCERGYF 0.450 49 QAGLELLGS 0.450 5 LLAGILLRI 0.400 38
FFLEMESHY 0.400 92 AFRFIQCLL 0.300 6 LAGILLRIT 0.300 1 MRRELLAGI
0.240 87 KKLKKAFRF 0.200 60 PPASASLVA 0.200 3 RELLAGILL 0.200 98
CLLLGLLKV 0.200 34 FYFYFFLEM 0.200 65 SLVAGTLSV 0.200 50 AGLELLGSS
0.200 29 VFFIYFYFY 0.200 119 CERGYFQGI 0.120 93 FRFIQCLLL 0.100 70
TLSVHHCAC 0.100 19 LFFFLPFPL 0.100 111 HQGVNSCDC 0.100 30 FFIYFYFYF
0.100 11 LRITFNFFL 0.100 55 LGSSNPPAS 0.100 32 IYFYFYFFL 0.100 54
LLGSSNPPA 0.100 26 PLVVFFIYF 0.100 14 TFNFFLFFF 0.100 44 SHYVAQAGL
0.100 69 GTLSVHHCA 0.100 109 LQHQGVNSC 0.100 17 FFLFFFLPF 0.100 81
SFTKRKKKL 0.100 124 FQGIFMQAA 0.100 74 HHCACFESF 0.100 75 HCACFESFT
0.100 120 ERGYFQGIF 0.100 68 AGTLSVHHC 0.100 129 MQAAPWEGT 0.100 8
GILLRITFN 0.100 39 FLEMESHYV 0.090 84 KRKKKLKKA 0.060 105 KVRPLQHQG
0.060 2 RRELLAGIL 0.060 80 ESFTKRKKK 0.050 43 ESHYVAQAG 0.050 76
CACFESFTK 0.045 67 VAGTLSVHH 0.030 82 FTKRKKKLK 0.030 103 LLKVRPLQH
0.030 51 GLELLGSSN 0.030 90 KKAFRFIQC 0.020 20 FFFLPFPLV 0.020 40
LEMESHYVA 0.020 77 ACFESFTKR 0.020 106 VRPLQHQGV 0.020 21 FFLPFPLVV
0.020 114 VNSCDCERG 0.015 42 MESHYVAQA 0.010 66 LVAGTLSVH 0.010 72
SVHHCACFE 0.010 100 LLGLLKVRP 0.010 18 FLFFFLPFP 0.010 125
QGIFMQAAP 0.010 97 QCLLLGLLK 0.010 99 LLLGLLKVR 0.010 48 AQAGLELLG
0.010 102 GLLKVRPLQ 0.010 73 VHHCACFES 0.010
V10-HLA-B3501-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 3 CPAGELGTS 3.000 9 GTSDVVTVV 0.400 8
LGTSDVVTV 0.300 2 RCPAGELGT 0.200 7 ELGTSDVVT 0.100 5 AGELGTSDV
0.060 6 GELGTSDVV 0.020 4 PAGELGTSD 0.006 1 GRCPAGELG 0.001
V11-HLA-B3501-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 23; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 1 QARLRLRVM 18.000 5 RLRVMVPPL 6.000 8
VMVPPLPSL 1.000 7 RVMVPPLPS 0.200 9 MVPPLPSLN 0.100 3 RLRLRVMVP
0.060 2 ARLRLRVMV 0.020 6 LRVMVPPLP 0.001 4 LRLRVMVPP 0.001
V12-HLA-B3501-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 25; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 9 CSYSTLTTV 1.000 5 EPEGCSYST 0.600 1
VMSEEPEGC 0.300 2 MSEEPEGCS 0.300 8 GCSYSTLTT 0.100 7 EGCSYSTLT
0.100 3 SEEPEGCSY 0.090 4 EEPEGCSYS 0.020 6 PEGCSYSTL 0.010
V13-HLA-B3501-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 7 VLADPQEDS 0.200 1 SQVTVDVLA 0.100 3
VTVDVLADP 0.020 2 QVTVDVLAD 0.015 6 DVLADPQED 0.015 8 LADPQEDSG
0.009 4 TVDVLADPQ 0.003 9 ADPQEDSGK 0.002 5 VDVLADPQE 0.001
V14-HLA-B3501-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 4 NPPASASLV 4.000 8 SASLVAGTL 3.000 3
SNPPASASL 1.000 9 ASLVAGTLS 0.500 7 ASASLVAGT 0.500 1 GSSNPPASA
0.500 2 SSNPPASAS 0.500 5 PPASASLVA 0.200 6 PASASLVAG 0.003
[1035]
31TABLE XXI Start Subsequence Score V1-HLA-B3501-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 493
RAKPTGNGIY 36.000 156 GPALEEGQGL 30.000 150 LPSLNPGPAL 20.000 132
FPAGSFQARL 20.000 409 RSQPEESVGL 15.000 407 DPRSQPEESV 12.000 1
MPLSLGAEMW 10.000 116 NAVQADEGEY 9.000 436 MSEEPEGRSY 9.000 334
DPQEDSGKQV 8.000 227 HPGLLQDQRI 8.000 11 GPEAWLLLLL 6.000 392
LTRENSIRRL 6.000 439 EPEGRSYSTL 6.000 383 TQKYEEELTL 4.500 249
SVRGLEDQNL 4.500 178 APSVTWDTEV 4.000 495 KPTGNGIYIN 4.000 271
LSEGQPPPSY 3.000 79 LALLHSKYGL 3.000 13 EAWLLLLLLL 3.000 201
SAAVTSEFHL 3.000 365 VVVVVLMSRY 2.000 276 PPPSYNWTRL 2.000 128
RVSTFPAGSF 2.000 35 ETSDVVTVVL 2.000 362 CLLVVVVVLM 2.000 235
RITHILHVSF 2.000 44 LGQDAKLPCF 2.000 144 RVLVPPLPSL 2.000 445
YSTLTTVREI 2.000 10 WGPEAWLLLL 2.000 176 SPAPSVTWDT 2.000 105
RNPLDGSVLL 2.000 244 FLAEASVRGL 2.000 138 QARLRLRVLV 1.800 291
SGVRVDGDTL 1.500 192 SSRSFKHSRS 1.500 212 PSRSMNGQPL 1.500 8
EMWGPEAWLL 1.500 426 SLKDNSSCSV 1.200 411 QPEESVGLRA 1.200 103
PPRNPLDGSV 1.200 303 PPLTTEHSGI 1.200 347 SASVVVVGVI 1.200 473
DQDEGIKQAM 1.200 361 FCLLVVVVVL 1.000 236 ITHILHVSFL 1.000 221
LTCVVSHPGL 1.000 222 TCVVSHPGLL 1.000 25 FTGRCPAGEL 1.000 346
VSASVVVVGV 1.000 354 GVIAALLFCL 1.000 57 DSGEQVGQVA 1.000 194
RSFKHSRSAA 1.000 214 RSMNGQPLTC 1.000 381 QMTQKYEEEL 1.000 137
FQARLRLRVL 1.000 355 VIAALLFCLL 1.000 350 VVVVGVIAAL 1.000 352
VVGVIAALLF 1.000 351 VVVGVIAALL 1.000 317 VSNEFSSRDS 1.000 500
GIYINGRGHL 1.000 16 LLLLLLLASF 1.000 99 EQPPPPRNPL 1.000 41
TVVLGQDAKL 1.000 280 YNWTRLDGPL 1.000 134 AGSFQARLRL 1.000 476
EGIKQAMNHF 1.000 321 FSSRDSQVTV 1.000 202 AAVTSEFHLV 0.900 67
WARVDAGEGA 0.900 341 KQVDLVSASV 0.800 230 LLQDQRITHI 0.800 169
ASCTAEGSPA 0.750 71 DAGEGAQELA 0.600 233 DQRITHILHV 0.600 158
ALEEGQGLTL 0.600 45 GQDAKLPCFY 0.600 477 GIKQAMNHFV 0.600 75
GAQELALLHS 0.600 357 AALLFCLLVV 0.600 261 IGREGAMLKC 0.600 356
IAALLFCLLV 0.600 423 HPDSLKDNSS 0.600 309 HSGIYVCHVS 0.500 248
ASVRGLEDQN 0.500 348 ASVVVVGVIA 0.500 174 EGSPAPSVTW 0.500 425
DSLKDNSSCS 0.500 338 DSGKQVDLVS 0.500 273 EGQPPPSYNW 0.500 6
GAEMWGPEAW 0.450 339 SGKQVDLVSA 0.450 106 NPLDGSVLLR 0.400 377
RKAQQMTQKY 0.400 452 REIETQTELL 0.400 389 ELTLTRENSI 0.400 305
LTTEHSGIYV 0.400 V2-HLA-B3501-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 1 LGQDAKLPCL 2.000 2
GQDAKLPCLY 0.600 7 LPCLYRGDSG 0.200 6 KLPCLYRGDS 0.200 4 DAKLPCLYRG
0.090 10 LYRGDSGEQV 0.060 9 CLYRGDSGEQ 0.015 3 QDAKLPCLYR 0.001 8
PCLYRGDSGE 0.001 5 AKLPCLYRGD 0.001 V7-HLA-B3501-10Mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 6
DPRSQSEEPE 0.600 9 SQSEEPEGRS 0.200 8 RSQSEEPEGR 0.150 1 HSHHTDPRSQ
0.075 2 SHHTDPRSQS 0.010 4 HTDPRSQSEE 0.003 3 HHTDPRSQSE 0.002 5
TDPRSQSEEP 0.001 7 PRSQSEEPEG 0.000 V9-HLA-B3501-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 19; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 25
FPLVVFFIYF 20.000 115 NSCDCERGYF 15.000 23 LPFPLVVFFI 8.000 91
KAFRFIQCLL 6.000 57 SSNPPASASL 5.000 80 ESFTKRKKKL 5.000 43
ESHYVAQAGL 5.000 62 ASASLVAGTL 5.000 107 RPLQHQGVNS 4.000 6
LAGILLRITF 3.000 10 LLRITFNFFL 3.000 59 NPPASASLVA 2.000 28
VVFFIYFYFY 2.000 114 VNSCDCERGY 2.000 12 RITFNFFLFF 2.000 105
KVRPLQHQGV 1.200 64 ASLVAGTLSV 1.000 70 TLSVHHCACF 1.000 13
ITFNFFLFFF 1.000 18 FLFFFLPFPL 1.000 100 LLGLLKVRPL 1.000 95
FIQCLLLGLL 1.000 8 GILLRITFNF 1.000 9 ILLRITFNFF 1.000 46
YVAQAGLELL 1.000 31 FIYFYFYFFL 1.000 27 LVVFFIYFYF 1.000 22
FLPFPLVVFF 1.000 86 KKKLKKAFRF 0.600 84 KRKKKLKKAF 0.600 1
MRRELLAGIL 0.600 56 GSSNPPASAS 0.500 125 QGIFMQAAPW 0.500 4
ELLAGILLRI 0.400 119 CERGYFQGIF 0.300 63 SASLVAGTLS 0.300 67
VAGTLSVHHC 0.300 92 AFRFIQCLLL 0.300 49 QAGLELLGSS 0.300 120
ERGYFQGIFM 0.200 58 SNPPASASLV 0.200 90 KKAFRFIQCL 0.200 33
YFYFYFFLEM 0.200 50 AGLELLGSSN 0.200 97 QCLLLGLLKV 0.200 26
PLVVFFIYFY 0.200 37 YFFLEMESHY 0.200 94 RFIQCLLLGL 0.200 48
AQAGLELLGS 0.150 118 DCERGYFQGI 0.120 21 FFLPFPLVVF 0.100 14
TFNFFLFFFL 0.100 30 FFIYFYFYFF 0.100 72 SVHHCACFES 0.100 55
LGSSNPPASA 0.100 69 GTLSVHHCAC 0.100 45 HYVAQAGLEL 0.100 53
ELLGSSNPPA 0.100 16 NFFLFFFLPF 0.100 128 FMQAAPWEGT 0.100 11
LRITFNFFLF 0.100 68 AGTLSVHHCA 0.100 7 AGILLRITFN 0.100 54
LLGSSNPPAS 0.100 73 VHHCACFESF 0.100 29 VFFIYFYFYF 0.100 5
LLAGILLRIT 0.100 87 KKLKKAFRFI 0.080 38 FFLEMESHYV 0.060 88
KLKKAFRFIQ 0.060 2 RRELLAGILL 0.060 71 LSVHHCACFE 0.050 83
TKRKKKLKKA 0.030 47 VAQAGLELLG 0.030 103 LLKVRPLQHQ 0.030 61
PASASLVAGT 0.030 76 CACFESFTKR 0.030 82 FTKRKKKLKK 0.030 89
LKKAFRFIQC 0.030 41 EMESHYVAQA 0.030 39 FLEMESHYVA 0.030 121
RGYFQGIFMQ 0.020 24 PFPLVVFFIY 0.020 60 PPASASLVAG 0.020 77
ACFESFTKRK 0.020 19 LFFFLPFPLV 0.020 20 FFFLPFPLVV 0.020 75
HCACFESFTK 0.015 113 GVNSCDCERG 0.015 108 PLQHQGVNSC 0.010 98
CLLLGLLKVR 0.010 110 QHQGVNSCDC 0.010 15 FNFFLFFFLP 0.010 99
LLLGLLKVRP 0.010 65 SLVAGTLSVH 0.010 101 LGLLKVRPLQ 0.010 111
HQGVNSCDCE 0.010 126 GIFMQAAPWE 0.010 96 IQCLLLGLLK 0.010 102
GLLKVRPLQH 0.010 V10-HLA-B3501-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 21; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 10 GTSDVVTVVL 2.000 8
ELGTSDVVTV 0.300 3 RCPAGELGTS 0.300 4 CPAGELGTSD 0.200 9 LGTSDVVTVV
0.200 5 PAGELGTSDV 0.120 6 AGELGTSDVV 0.060 1 TGRCPAGELG 0.030 2
GRCPAGELGT 0.010 7 GELGTSDVVT 0.010 V11-HLA-B3501-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 23; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 8
RVMVPPLPSL 2.000 1 FQARLRLRVM 2.000 2 QARLRLRVMV 1.800 9 VMVPPLPSLN
0.100 5 LRLRVMVPPL 0.100 4 RLRLRVMVPP 0.060 6 RLRVMVPPLP 0.060 10
MVPPLPSLNP 0.010 7 LRVMVPPLPS 0.010 3 ARLRLRVMVP 0.001
V12-HLA-B3501-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 25; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 3 MSEEPEGCSY 9.000 6 EPEGCSYSTL 6.000 2
VMSEEPEGCS 0.200 9 GCSYSTLTTV 0.200 1 SVMSEEPEGC 0.150 8 EGCSYSTLTT
0.100 10 CSYSTLTTVR 0.050 5 EEPEGCSYST 0.020 4 SEEPEGCSYS 0.003 7
PEGCSYSTLT 0.001 11 SYSTLTTVRE 0.001 V13-HLA-B3501-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 27; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 1
DSQVTVDVLA 0.500 7 DVLADPQEDS 0.100 8 VLADPQEDSG 0.020 4 VTVDVLADPQ
0.020 2 SQVTVDVLAD 0.015 9 LADPQEDSGK 0.013 3 QVTVDVLADP 0.010 5
TVDVLADPQE 0.003 10 ADPQEDSGKQ 0.002 6 VDVLADPQED 0.002
V14-HLA-B3501-10mers-19- 1P4D12B Each peptide is a portion of SEQ
ID NO: 29; each start position is specified, the length of peptide
is 10 amino acids, and the end position for each peptide is the
start position plus nine. 8 ASASLVAGTL 5.000 3 SSNPPASASL 5.000 5
NPPASASLVA 2.000 10 ASLVAGTLSV 1.000 2 GSSNPPASAS 0.500 9
SASLVAGTLS 0.300 4 SNPPASASLV 0.200 1 LGSSNPPASA 0.100 7 PASASLVAGT
0.030 6 PPASASLVAG 0.020
[1036]
32TABLE XXII Pos 1 2 3 4 5 6 7 8 9 score V1-HLA-A1-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 437 S E
E P E G R S Y 32 107 P L D G S V L L R 21 305 L T T E H S G I Y 21
306 T T E H S G I Y V 21 159 L E E G Q G L T L 20 252 G L E D Q N L
W H 20 405 H T D P R S Q P E 20 86 Y G L H V S P A Y 19 262 G R E G
A M L K C 19 412 P E E S V G L R A 19 486 V Q E N G T L R A 19 494
A K P T G N G I Y 19 11 G P E A W L L L L 18 78 E L A L L H S K Y
18 272 S E G Q P P P S Y 18 332 V L D P Q E D S G 18 386 Y E E E L
T L T R 18 36 T S D V V T V V L 17 76 A Q E L A L L H S 17 184 D T
E V K G T T S 17 225 V S H P G L L Q D 17 271 L S E G Q P P P S 17
294 R V D G D T L G F 17 378 K A Q Q M T Q K Y 17 58 S G E Q V G Q
V A 16 117 A V Q A D E G E Y 16 158 A L E E G Q G L T 16 323 S R D
S Q V T V D 16 366 V V V V L M S R Y 16 457 Q T E L L S P G S 16 46
Q D A K L P C F Y 15 436 M S E E P E G R S 15
V2-HLA-A1-9mers-191P4D12 Each peptide is a portion of SEQ ID NO: 5;
each start position is specified, the length of peptide is 9 amino
acids, and the end position for each peptide is the start position
plus eight. 2 Q D A K L P C L Y 17 1 G Q D A K L P C L 10
V7-HLA-A1-9mers-191P4D12 Each peptide is a portion of SEQ ID NO:
15; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 3 H T D P R S Q S E 20
V9-HLA-A1-9mers-191P4D12 Each peptide is a portion of SEQ ID NO:
19; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 25 F P L V V F F I Y 21 29 V F F I Y F Y F Y
20 115 N S C D C E R G Y 19 38 F F L E M E S H Y 16 13 I T F N F F
L F F 15 27 L V V F F I Y F Y 15 116 S C D C E R G Y F 13 21 F F L
P F P L V V 12 39 F L E M E S H Y V 12 51 G L E L L G S S N 12 118
D C E R G Y F Q G 12 4 E L L A G I L L R 11 57 S S N P P A S A S 11
65 S L V A G T L S V 11 93 F R F I Q C L L L 11 98 C L L L G L L K
V 11 2 R R E L L A G I L 10 17 F F L F F F L P F 10 34 F Y F Y F F
L E M 10 41 E M E S H Y V A Q 10 48 A Q A G L E L L G 10 78 C F E S
F T K R K 10 V10-HLA-A1-9mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 21; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 5 A G E L G T S D V 13 9 G T S D V V
T V V 10 2 R C P A G E L G T 8 1 G R C P A G E L G 7
V11-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
23; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 7 R V M V P P L P S 7 8 V M V P P L P S L 6 9
M V P P L P S L N 6 6 L R V M V P P L P 4 2 A R L R L R V M V 3 3 R
L R L R V M V P 3 V12-HLA-A1-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 3 S E E P E G C S Y 32
V13-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
27; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 8 L A D P Q E D S G 16 4 T V D V L A D P Q 10
3 V T V D V L A D P 9 2 Q V T V D V L A D 7
V14-HLA-A1-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
29; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 2 S S N P P A S A S 11 9 A S L V A G T L S 8 5
P P A S A S L V A 7 3 S N P P A S A S L 6 7 A S A S L V A G T 6 1 G
S S N P P A S A 5
[1037]
33TABLE XXIII Pos 1 2 3 4 5 6 7 8 9 Score
V1-HLA-A0201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 145 V L V P P L P S L 31 359 L L F C L L V V V
30 358 A L L F C L L V V 28 362 C L L V V V V V L 28 80 A L L H S K
Y G L 26 142 R L R V L V P P L 26 355 V I A A L L F C L 26 351 V V
V G V I A A L 24 502 Y I N G R G H L V 24 17 L L L L L L A S F 23
42 V V L G Q D A K L 23 347 S A S V V V V G V 23 15 W L L L L L L L
A 22 345 L V S A S V V V V 22 363 L L V V V V V L M 22 446 S T L T
T V R E I 22 8 E M W G P E A W L 21 16 L L L L L L L A S 21 344 D L
V S A S V V V 21 14 A W L L L L L L L 20 245 L A E A S V R G L 20
260 H I G R E G A M L 20 284 R L D G P L P S G 20 357 A A L L F C L
L V 20 460 L L S P G S G R A 20 18 L L L L L A S F T 19 34 L E T S
D V V T V 19 71 D A G E G A Q E L 19 112 V L L R N A V Q A 19 152 S
L N P G P A L E 19 158 A L E E G Q G L T 19 356 I A A L L F C L L
19 360 L F C L L V V V V 19 361 F C L L V V V V V 19 390 L T L T R
E N S I 19 13 E A W L L L L L L 18 138 Q A R L R L R V L 18 266 A M
L K C L S E G 18 342 Q V D L V S A S V 18 481 A M N H F V Q E N 18
21 L L A S F T G R C 17 106 N P L D G S V L L 17 113 L L R N A V Q
A D 17 139 A R L R L R V L V 17 229 G L L Q D Q R I T 17 234 Q R I
T H I L H V 17 244 F L A E A S V R G 17 287 G P L P S G V R V 17
292 G V R V D G D T L 17 299 T L G F P P L T T 17 322 S S R D S Q V
T V 17 352 V V G V I A A L L 17 382 M T Q K Y E E E L 17 410 S Q P
E E S V G L 17 419 R A E G H P D S L 17 443 R S Y S T L T T V 17 19
L L L L A S F T G 16 35 E T S D V V T V V 16 157 P A L E E G Q G L
16 159 L E E G Q G L T L 16 173 A E G S P A P S V 16 202 A A V T S
E F H L 16 203 A V T S E F H L V 16 215 S M N G Q P L T C 16 237 T
H I L H V S F L 16 242 V S F L A E A S V 16 285 L D G P L P S G V
16 350 V V V V G V I A A 16 384 Q K Y E E E L T L 16 452 R E I E T
Q T E L 16 453 E I E T Q T E L L 16 501 I Y I N G R G H L 16 11 G P
E A W L L L L 15 12 P E A W L L L L L 15 20 L L L A S F T G R 15 32
G E L E T S D V V 15 57 D S G E Q V G Q V 15 74 E G A Q E L A L L
15 137 F Q A R L R L R V 15 140 R L R L R V L V P 15 216 M N G Q P
L T C V 15 217 N G Q P L T C V V 15 230 L L Q D Q R I T H 15 240 L
H V S F L A E A 15 270 C L S E G Q P P P 15 304 P L T T E H S G I
15 309 H S G I Y V C H V 15 332 V L D P Q E D S G 15 493 R A K P T
G N G I 15 V2-HLA-A0201-9mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 5; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 1 G Q D A K L P C L 17 8 C L Y R G D
S G E 14 5 K L P C L Y R G D 13 4 A K L P C L Y R G 11
V7-HLA-A0201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 3 H T D P R S Q S E 8 8 S Q S E E P E G R 5 1
S H H T D P R S Q 4 7 R S Q S E E P E G 3
V9-HLA-A0201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 98 C L L L G L L K V 31 5 L L A G I L L R I 29
65 S L V A G T L S V 29 95 F I Q C L L L G L 26 39 F L E M E S H Y
V 21 46 Y V A Q A G L E L 21 47 V A Q A G L E L L 21 91 K A F R F I
Q C L 21 99 L L L G L L K V R 20 101 L G L L K V R P L 19 1 M R R E
L L A G I 18 58 S N P P A S A S L 18 63 S A S L V A G T L 18 88 K L
K K A F R F I 18 18 F L F F F L P F P 17 21 F F L P F P L V V 17 22
F L P F P L V V F 17 54 L L G S S N P P A 17 96 I Q C L L L G L L
17 4 E L L A G I L L R 16 9 I L L R I T F N F 16 44 S H Y V A Q A G
L 16 62 A S A S L V A G T 16 6 L A G I L L R I T 15 8 G I L L R I T
F N 15 11 L R I T F N F F L 15 100 L L G L L K V R P 15
V10-HLA-A0201-9mers-191P4D1- 2B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 9 G T S D V V T V V 20 8 L G T S D V V T V 19
5 A G E L G T S D V 15 6 G E L G T S D V V 15 7 E L G T S D V V T
13 3 C P A G E L G T S 10 V11-HLA-A0201-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 23; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 8 V M V
P P L P S L 29 5 R L R V M V P P L 25 2 A R L R L R V M V 17 3 R L
R L R V M V P 14 V12-HLA-A0201-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 9 C S Y S T L T T V 17 1
V M S E E P E G C 12 6 P E G C S Y S T L 9 8 G C S Y S T L T T 9
V13-HLA-A0201-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 7 V L A D P Q E D S 15 3 V T V D V L A D P 12
8 L A D P Q E D S G 10 2 Q V T V D V L A D 9 1 S Q V T V D V L A 8
6 D V L A D P Q E D 7 V14-HLA-A0201-9mers-191P4D12B Each peptide is
a portion of SEQ ID NO: 29; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 3 S N P P A S A S L 18 8
S A S L V A G T L 18 7 A S A S L V A G T 16 1 G S S N P P A S A 10
4 N P P A S A S L V 10 6 P A S A S L V A G 8
[1038]
34TABLE XXIV Pos 1 2 3 4 5 6 7 8 9 score
V1-HLA-A0203-9mers-191P4D12B No Results Found.
V2-HLA-A0203-9mers-191P4D12B No Results Found.
V7-HLA-A0203-9mers-191P4D12B No Results Found.
V9-HLA-A0203-9mers-191P4D12B No Results Found.
V10-HLA-A0203-9mers-191P4D12B No Results Found.
V11-HLA-A0203-9mers-191P4D12B No Results Found.
V12-HLA-A0203-9mers-191P4D12B No Results Found.
V13-HLA-A0203-9mers-191P4D12B No Results Found.
V14-HLA-A0203-9mers-191P4D12B No Results Found.
[1039]
35TABLE XXV Pos 1 2 3 4 5 6 7 8 9 score V1-HLA-A03-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 140 R L
R L R V L V P 27 112 V L L R N A V Q A 25 180 S V T W D T E V K 25
41 T V V L G Q D A K 24 111 S V L L R N A V Q 23 294 R V D G D T L
G F 23 17 L L L L L L A S F 22 117 A V Q A D E G E Y 22 186 E V K G
T T S S R 22 261 I G R E G A M L K 22 358 A L L F C L L V V 22 397
S I R R L H S H H 22 459 E L L S P G S G R 22 61 Q V G Q V A W A R
21 78 E L A L L H S K Y 21 362 C L L V V V V V L 21 415 S V G L R A
E G H 21 69 R V D A G E G A Q 20 144 R V L V P P L P S 20 152 S L N
P G P A L E 20 230 L L Q D Q R I T H 20 292 G V R V D G D T L 20
316 H V S N E F S S R 20 345 L V S A S V V V V 20 391 T L T R E N S
I R 20 500 G I Y I N G R G H 20 18 L L L L L A S F T 19 20 L L L A
S F T G R 19 97 R V E Q P P P P R 19 107 P L D G S V L L R 19 243 S
F L A E A S V R 19 249 S V R G L E D Q N 19 252 G L E D Q N L W H
19 342 Q V D L V S A S V 19 349 S V V V V G V I A 19 366 V V V V L
M S R Y 19 377 R K A Q Q M T Q K 19 485 F V Q E N G T L R 19 33 E L
E T S D V V T 18 64 Q V A W A R V D A 18 77 Q E L A L L H S K 18
128 R V S T F P A G S 18 209 H L V P S R S M N 18 260 H I G R E G A
M L 18 284 R L D G P L P S G 18 299 T L G F P P L T T 18 311 G I Y
V C H V S N 18 344 D L V S A S V V V 18 354 G V I A A L L F C 18
359 L L F C L L V V V 18 365 V V V V V L M S R 18 417 G L R A E G H
P D 18 450 T V R E I E T Q T 18 491 T L R A K P T G N 18 2 P L S L
G A E M W 17 16 L L L L L L L A S 17 19 L L L L A S F T G 17 42 V V
L G Q D A K L 17 89 H V S P A Y E G R 17 142 R L R V L V P P L 17
146 L V P P L P S L N 17 158 A L E E G Q G L T 17 164 G L T L A A S
C T 17 351 V V V G V I A A L 17 368 V V L M S R Y H R 17 15 W L L L
L L L L A 16 81 L L H S K Y G L H 16 197 K H S R S A A V T 16 224 V
V S H P G L L Q 16 235 R I T H I L H V S 16 239 I L H V S F L A E
16 244 F L A E A S V R G 16 288 P L P S G V R V D 16 352 V V G V I
A A L L 16 369 V L M S R Y H R R 16 420 A E G H P D S L K 16 426 S
L K D N S S C S 16 460 L L S P G S G R A 16 39 V V T V V L G Q D 15
80 A L L H S K Y G L 15 105 R N P L D G S V L 15 113 L L R N A V Q
A D 15 145 V L V P P L P S L 15 166 T L A A S C T A E 15 200 R S A
A V T S E F 15 313 Y V C H V S N E F 15 327 Q V T V D V L D P 15
332 V L D P Q E D S G 15 363 L L V V V V V L M 15 364 L V V V V V L
M S 15 367 V V V L M S R Y H 15 373 R Y H R R K A Q Q 15 400 R L H
S H H T D P 15 437 S E E P E G R S Y 15 487 Q E N G T L R A K 15
502 Y I N G R G H L V 15 38 D V V T V V L G Q 14 87 G L H V S P A Y
E 14 189 G T T S S R S F K 14 198 H S R S A A V T S 14 21 Q P L T C
V V S H 14 220 P L T C V V S H P 14 241 H V S F L A E A S 14 384 Q
K Y E E E L T L 14 396 N S I R R L H S H 14 409 R S Q P E E S V G
14 4 S L G A E M W G P 13 43 V L G Q D A K L P 13 49 K L P C F Y R
G D 13 84 S K Y G L H V S P 13 124 E Y E C R V S T F 13 139 A R L R
L R V L V 13 203 A V T S E F H L V 13 210 L V P S R S M N G 13 236
I T H I L H V S F 13 257 N L W H I G R E G 13 270 C L S E G Q P P P
13 304 P L T T E H S G I 13 322 S S R D S Q V T V 13 329 T V D V L
D P Q E 13 331 D V L D P Q E D S 13 333 L D P Q E D S G K 13 350 V
V V V G V I A A 13 370 L M S R Y H R R K 13 374 Y H R R K A Q Q M
13 443 R S Y S T L T T V 13 477 G I K Q A M N H F 13
V2-HLA-A03-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
5; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 8 C L Y R G D S G E 22 5 K L P C L Y R G D 13
2 Q D A K L P C L Y 10 V7-HLA-A03-9mers-191P4D- 12B Each peptide is
a portion of SEQ ID NO: 15; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 2 H H T D P R S Q S 8 3 H
T D P R S Q S E 7 8 S Q S E E P E G R 7 4 T D P R S Q S E E 6 1 S H
H T D P R S Q 4 7 R S Q S E E P E G 4 5 D P R S Q S E E P 3
V9-HLA-A03-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
19; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 66 L V A G T L S V H 24 103 L L K V R P L Q H
24 4 E L L A G I L L R 23 22 F L P F P L V V F 22 99 L L L G L L K
V R 22 105 K V R P L Q H Q G 22 9 I L L R I T F N F 21 97 Q C L L L
G L L K 21 65 S L V A G T L S V 20 51 G L E L L G S S N 19 10 L L R
I T F N F F 18 98 C L L L G L L K V 18 46 Y V A Q A G L E L 17 83 T
K R K K K L K K 17 108 P L Q H Q G V N S 17 5 L L A G I L L R I 16
7 A G I L L R I T F 16 12 R I T F N F F L F 16 27 L V V F F I Y F Y
16 31 F I Y F Y F Y F F 16 82 F T K R K K K L K 15 100 L L G L L K
V R P 15 8 G I L L R I T F N 14 26 P L V V F F I Y F 14 28 V V F F
I Y F Y F 14 53 E L L G S S N P P 14 72 S V H H C A C F E 14 76 C A
C F E S F T K 14 88 K L K K A F R F I 14 102 G L L K V R P L Q 14
113 G V N S C D C E R 14 126 G I F M Q A A P W 14 21 F F L P F P L
V V 13 86 K K K L K K A F R 13 87 K K L K K A F R F 13 38 F F L E M
E S H Y 12 80 E S F T K R K K K 12 23 L P F P L V V F F 11 57 S S N
P P A S A S 11 63 S A S L V A G T L 11 70 T L S V H H C A C 11 95 F
I Q C L L L G L 11 107 R P L Q H Q G V N 11 121 R G Y F Q G I F M
11 V10-HLA-A03-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 7 E L G T S D V V T 18 2 R C P A G E L G T 11
5 A G E L G T S D V 9 3 C P A G E L G T S 8 6 G E L G T S D V V 8 8
L G T S D V V T V 8 V11-HLA-A03-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 23; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 3 R L R L R V M V P 25 7
R V M V P P L P S 18 5 R L R V M V P P L 17 9 M V P P L P S L N 17
2 A R L R L R V M V 14 1 Q A R L R L R V M 12
V12-HLA-A03-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 3 S E E P E G C S Y 15 9 C S Y S T L T T V 9 6
P E G C S Y S T L 7 8 G C S Y S T L T T 7
V13-HLA-A03-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
27; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 2 Q V T V D V L A D 16 9 A D P Q E D S G K 16
6 D V L A D P Q E D 15 4 T V D V L A D P Q 13 7 V L A D P Q E D S
12 V14-HLA-A03-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 2 S S N P P A S A S 11 8 S A S L V A G T L 11
3 S N P P A S A S L 9 9 A S L V A G T L S 9 4 N P P A S A S L V 8 5
P P A S A S L V A 8 1 G S S N P P A S A 7 6 P A S A S L V A G 7 7 A
S A S L V A G T 7
[1040]
36TABLE XXVI Pos 1 2 3 4 5 6 7 8 9 score V1-HLA-A26-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 38 D V
V T V V L G Q 27 351 V V V G V I A A L 27 366 V V V V L M S R Y 26
13 E A W L L L L L L 24 124 E Y E C R V S T F 24 223 C V V S H P G
L L 24 455 E T Q T E L L S P 24 35 E T S D V V T V V 23 78 E L A L
L H S K Y 23 74 E G A Q E L A L L 22 186 E V K G T T S S R 22 305 L
T T E H S G I Y 22 453 E I E T Q T E L L 22 117 A V Q A D E G E Y
21 292 G V R V D G D T L 20 325 D S Q V T V D V L 20 350 V V V V G
V I A A 20 352 V V G V I A A L L 20 364 L V V V V V L M S 20 42 V V
L G Q D A K L 19 184 D T E V K G T T S 19 294 R V D G D T L G F 19
331 D V L D P Q E D S 19 337 E D S G K Q V D L 19 354 G V I A A L L
F C 19 365 V V V V V L M S R 19 8 E M W G P E A W L 18 60 E Q V G Q
V A W A 18 71 D A G E G A Q E L 18 145 V L V P P L P S L 18 236 I T
H I L H V S F 18 237 T H I L H V S F L 18 313 Y V C H V S N E F 18
449 T T V R E I E T Q 18 39 V V T V V L G Q D 17 328 V T V D V L D
P Q 17 355 V I A A L L F C L 17 41 T V V L G Q D A K 16 57 D S G E
Q V G Q V 16 130 S T F P A G S F Q 16 298 D T L G F P P L T 16 327
Q V T V D V L D P 16 349 S V V V V G V I A 16 382 M T Q K Y E E E L
16 450 T V R E I E T Q T 16 413 E E S V G L R A E 15 414 E S V G L
R A E G 15 473 D Q D E G I K Q A 15 12 P E A W L L L L L 14 14 A W
L L L L L L L 14 17 L L L L L L A S F 14 40 V T V V L G Q D A 14
160 E E G Q G L T L A 14 260 H I G R E G A M L 14 345 L V S A S V V
V V 14 367 V V V L M S R Y H 14 387 E E E L T L T R E 14 437 S E E
P E G R S Y 14 452 R E I E T Q T E L 14 472 E D Q D E G I K Q 14
476 E G I K Q A M N H 14 484 H F V Q E N G T L 14 485 F V Q E N G T
L R 14 11 G P E A W L L L L 13 45 G Q D A K L P C F 13 109 D G S V
L L R N A 13 135 G S F Q A R L R L 13 142 R L R V L V P P L 13 146
L V P P L P S L N 13 161 E G Q G L T L A A 13 222 T C V V S H P G L
13 249 S V R G L E D Q N 13 320 E F S S R D S Q V 13 329 T V D V L
D P Q E 13 344 D L V S A S V V V 13 353 V G V I A A L L F 13 393 T
R E N S I R R L 13 421 E G H P D S L K D 13 438 E E P E G R S Y S
13 446 S T L T T V R E I 13 459 E L L S P G S G R 13 501 I Y I N G
R G H L 13 V2-HLA-A26-9mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 5; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 1 G Q D A K L P C L 13 2 Q D A K L P
C L Y 11 3 D A K L P C L Y R 9 V7-HLA-A26-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 3 H T D
P R S Q S E 10 5 D P R S Q S E E P 9 2 H H T D P R S Q S 4
V9-HLA-A26-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
19; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 27 L V V F F I Y F Y 28 28 V V F F I Y F Y F
24 13 I T F N F F L F F 21 46 Y V A Q A G L E L 20 120 E R G Y F Q
G I F 19 23 L P F P L V V F F 18 95 F I Q C L L L G L 18 80 E S F T
K R K K K 16 91 K A F R F I Q C L 16 4 E L L A G I L L R 15 7 A G I
L L R I T F 15 66 L V A G T L S V H 15 12 R I T F N F F L F 14 29 V
F F I Y F Y F Y 14 96 I Q C L L L G L L 14 14 T F N F F L F F F 13
15 F N F F L F F F L 13 19 L F F F L P F P L 13 26 P L V V F F I Y
F 13 38 F F L E M E S H Y 13 93 F R F I Q C L L L 13 101 L G L L K
V R P L 13 105 K V R P L Q H Q G 13 V10-HLA-A26-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 21; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 9 G T S
D V V T V V 13 7 E L G T S D V V T 10 8 L G T S D V V T V 7 3 C P A
G E L G T S 6 V11-HLA-A26-9mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 23; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 8 V M V P P L P S L 18 9 M V P P L P
S L N 13 5 R L R V M V P P L 12 7 R V M V P P L P S 11
V12-HLA-A26-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 3 S E E P E G C S Y 14 4 E E P E G C S Y S 13
5 E P E G C S Y S T 11 7 E G C S Y S T L T 11 6 P E G C S Y S T L
10 9 C S Y S T L T T V 6 V13-HLA- A26-9mers-191P4D12B Each peptide
is a portion of SEQ ID NO: 27; each start position is specified,
the length of peptide is 9 amino acids, and the end position for
each peptide is the start position plus eight. 6 D V L A D P Q E D
18 2 Q V T V D V L A D 17 3 V T V D V L A D P 17 4 T V D V L A D P
Q 12 V14-HLA- A26-9mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 29; each start position is specified, the length of peptide
is 9 amino acids, and the end position for each peptide is the
start position plus eight. 3 S N P P A S A S L 11 8 S A S L V A G T
L 11 7 A S A S L V A T G 6 6 P A S A S L V A G 5
[1041]
37TABLE XXVII Pos 1 2 3 4 5 6 7 8 9 score
V1-HLA-B0702-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 100 Q P P P P R N P L 26 11 G P E A W L L L L
23 277 P P S Y N W T R L 23 106 N P L D G S V L L 22 287 G P L P S
G V R V 20 495 K P T G N G I Y I 20 150 L P S L N P G P A 19 439 E
P E G R S Y S T 19 1 M P L S L G A E M 18 8 E M W G P E A W L 17
275 Q P P P S Y N W T 17 289 L P S G V R V D G 17 337 E D S G K Q V
D L 17 142 R L R V L V P P L 16 151 P S L N P G P A L 16 26 T G R C
P A G E L 15 36 T S D V V T V V L 15 73 G E G A Q E L A L 15 103 P
P R N P L D G S 15 132 F P A G S F Q A R 15 145 V L V P P L P S L
15 147 V P P L P S L N P 15 159 L E E G Q G L T L 15 14 A W L L L L
L L L 14 176 S P A P S V T W D 14 178 A P S V T W D T E 14 213 S R
S M N G Q P L 14 351 V V V G V I A A L 14 362 C L L V V V V V L 14
12 P E A W L L L L L 13 13 E A W L L L L L L 13 29 C P A G E L E T
S 13 42 V V L G Q D A K L 13 74 E G A Q E L A L L 13 91 S P A Y E G
R V E 13 105 R N P L D G S V L 13 135 G S F Q A R L R L 13 138 Q A
R L R L R V L 13 161 E G Q G L T L A A 13 173 A E G S P A P S V 13
219 Q P L T C V V S H 13 260 H I G R E G A M L 13 263 R E G A M L K
C L 13 292 G V R V D G D T L 13 294 R V D G D T L G F 13 297 G D T
L G F P P L 13 345 L V S A S V V V V 13 356 I A A L L F C L L 13
419 R A E G H P D S L 13 462 S P G S G R A E E 13 9 M W G P E A W L
L 12 10 W G P E A W L L L 12 35 E T S D V V T V V 12 80 A L L H S K
Y G L 12 82 L H S K Y G L H V 12 101 P P P P R N P L D 12 102 P P P
R N P L D G 12 133 P A G S F Q A R L 12 148 P P L P S L N P G 12
154 N P G P A L E E G 12 202 A A V T S E F H L 12 211 V P S R S M N
G Q 12 237 T H I L H V S F L 12 245 L A E A S V R G L 12 299 T L G
F P P L T T 12 324 R D S Q V T V D V 12 325 D S Q V T V D V L 12
352 V V G V I A A L L 12 355 V I A A L L F C L 12 384 Q K Y E E E L
T L 12 407 D P R S Q P E E S 12 410 S Q P E E S V G L 12 452 R E I
E T Q T E L 12 453 E I E T Q T E L L 12 501 I Y I N G R G H L 12
V2-HLA-B0702-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 1 G Q D A K L P C L 13 6 L P C L Y R G D S 11
V7-HLA-B0702-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 5 D P R S Q S E E P 12
V9-HLA-B0702-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 23 L P F P L V V F F 21 60 P P A S A S L V A
20 59 N P P A S A S L V 17 46 Y V A Q A G L E L 14 92 A F R F I Q C
L L 14 3 R E L L A G I L L 12 15 F N F F L F F F L 12 22 F L P F P
L V V F 12 32 I Y F Y F Y F F L 12 56 G S S N P P A S A 12 58 S N P
P A S A S L 12 63 S A S L V A G T L 12 93 F R F I Q C L L L 12 95 F
I Q C L L L G L 12 101 L G L L K V R P L 12 107 R P L Q H Q G V N
12 2 R R E L L A G I L 11 5 L L A G I L L R I 11 11 L R I T F N F F
L 11 13 I T F N F F L F F 11 19 L F F F L P F P L 11 20 F F F L P F
P L V 11 25 F P L V V F F I Y 11 44 S H Y V A Q A G L 11 4 V A Q A
G L E L L 11 62 A S A S L V A G T 11 81 S F T K R K K K L 11 91 K A
F R F I Q C L 11 96 I Q C L L L G L L 11 119 C E R G Y F Q G I 11
129 M Q A A P W E G T 11 10 L L R I T F N F F 10 17 F F L F F F L P
F 10 21 F F L P F P L V V 10 42 M E S H Y V A Q A 10 65 S L V A G T
L S V 10 88 K L K K A F R F I 10 V10-HLA-B0702-9mers-191P4D12- B
Each peptide is a portion of SEQ ID NO: 21; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 3 C P A
G E L G T S 13 7 E L G T S D V V T 11 9 G T S D V V T V V 11 2 R C
P A G E L G T 10 5 A G E L G T S D V 9 6 G E L G T S D V V 9 8 L G
T S D V V T V 9 V11-HLA-B0702-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 23; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 5 R L R V M V P P L 16 8
V M V P P L P S L 15 2 A R L R L R V M V 11 1 Q A R L R L R V M 9 7
R V M V P P L P S 8 V12-HLA-B0702-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 5 E P E G C S Y S T 19 6
P E G C S Y S T L 11 8 G C S Y S T L T T 11
V13-HLA-B0702-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 1 S Q V T V D V L A 8 2 Q V T V D V L A D 4 7
V L A D P Q E D S 4 V14-HLA-B0702-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 29; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 5 P P A S A S L V A 20 4
N P P A S A S L V 17 1 G S S N P P A S A 12 3 S N P P A S A S L 12
8 S A S L V A G T L 12 7 A S A S L V A G T 11
[1042]
38TABLE XXVIII Pos 1 2 3 4 5 6 7 8 9 score
V1-HLA-B08-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
3; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 138 Q A R L R L R V L 29 142 R L R V L V P P L
24 337 E D S G K Q V D L 23 140 R L R L R V L V P 22 491 T L R A K
P T G N 22 477 G I K Q A M N H F 21 493 R A K P T G N G I 20 362 C
L L V V V V V L 19 292 G V R V D G D T L 18 426 S L K D N S S C S
18 11 G P E A W L L L L 17 13 E A W L L L L L L 17 26 T G R C P A G
E L 17 45 G Q D A K L P C F 17 71 D A G E G A Q E L 17 106 N P L D
G S V L L 17 124 E Y E C R V S T F 17 145 V L V P P L P S L 17 27 P
P S Y N W T R L 17 80 A L L H S K Y G L 16 81 L L H S K Y G L H 16
100 Q P P P P R N P L 16 157 P A L E E G Q G L 16 247 E A S V R G L
E D 16 265 G A M L K C L S E 16 267 M L K C L S E G Q 16 356 I A A
L L F C L L 16 374 Y H R R K A Q Q M 16 439 E P E G R S Y S T 16
453 E I E T Q T E L L 16 47 D A K L P C F Y R 15 65 V A W A R V D A
G 15 101 P P P P R N P L D 15 231 L Q D Q R I T H I 15 245 L A E A
S V R G L 15 260 H I G R E G A M L 15 355 V I A A L L F C L 15 369
V L M S R Y H R R 15 410 S Q P E E S V G L 15 113 L L R N A V Q A D
14 133 P A G S F Q A R L 14 202 A A V T S E F H L 14 390 L T L T R
E N S I 14 419 R A E G H P D S L 14 V2-HLA-B08-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 5; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 1 G Q D
A K L P C L 21 3 D A K L P C L Y R 15 V7-HLA-B08-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 5 D P R
S Q S E E P 13 3 H T D P R S Q S E 9 V9-HLA-B08-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 19; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 103 L L
K V R P L Q H 25 82 F T K R K K K L K 22 88 K L K K A F R F I 22
101 L G L L K V R P L 22 811 S F T K R K K K L 21 84 K R K K K L K
K A 21 86 K K K L K K A F R 21 10 L L R I T F N F F 18 85 R K K K L
K K A F 18 63 S A S L V A G T L 17 83 T K R K K K L K K 16 87 K K L
K K A F R F 16 92 A F R F I Q C L L 16 8 G I L L R I T F N 15 47 V
A Q A G L E L L 15 91 K A F R F I Q C L 15
V9-HLA-B08-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
19; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 95 F I Q C L L L G L 15 1 M R R E L L A G I 14
22 F L P F P L V V F 14 23 L P F P L V V F F 14 9 I L L R I T F N F
13 26 P L V V F F I Y F 13 44 S H Y V A Q A G L 13 80 E S F T K R K
K K 13 5 L L A G I L L R I 12 32 I Y F Y F Y F F L 12 58 S N P P A
S A S L 12 96 I Q C L L L G L L 12 119 C E R G Y F Q G I 12
V10-HLA-B08-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
21; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 7 E L G T S D V V T 9 3 C P A G E L G T S 6 4
P A G E L G T S D 6 V11-HLA-B08-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 23; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 5 R L R V M V P P L 24 3
R L R L R V M V P 22 1 Q A R L R L R V M 19 8 V M V P P L P S L 11
V12-HLA-B08-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 6 P E G C S Y S T L 10 5 E P E G C S Y S T 8 4
E E P E G C S Y S 4 V13-HLA-B08-9mers-191P4D1- 2B Each peptide is a
portion of SEQ ID NO: 27; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 7 V L A D P Q E D S 7 8 L
A D P Q E D S G 4 1 S Q V T V D V L A 3 2 Q V T V D V L A D 3
V14-HLA-B08-9mers-191P4D12B Each peptide is a portion of SEQ ID NO:
29; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 8 S A S L V A G T L 17 3 S N P P A S A S L
12
[1043]
39TABLE XXIX Pos 1 2 3 4 5 6 7 8 9 score
V1-HLA-B1510-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 237 T H I L H V S F L 22 208 F H L V P S R S M
20 259 W H I G R E G A M 18 374 Y H R R K A Q Q M 17 393 T R E N S
I R R L 17 36 T S D V V T V V L 16 362 C L L V V V V V L 16 135 G S
F Q A R L R L 15 308 E H S G I Y V C H 15 337 E D S G K Q V D L 15
100 Q P P P P R N P L 14 106 N P L D G S V L L 14 138 Q A R L R L R
V L 14 145 V L V P P L P S L 14 245 L A E A S V R G L 14 277 P P S
Y N W T R L 14 325 D S Q V T V D V L 14 501 I Y I N G R G H L 14 8
E M W G P E A W L 13 26 T G R C P A G E L 13 71 D A G E G A Q E L
13 74 E G A Q E L A L L 13 142 R L R V L V P P L 13 151 P S L N P G
P A L 13 159 L E E G Q G L T L 13 197 K H S R S A A V T 13 222 T C
V V S H P G L 13 292 G V R V D G D T L 13 297 G D T L G F P P L 13
351 V V V G V I A A L 13 356 I A A L L F C L L 13 403 S H H T D P R
S Q 13 404 H H T D P R S Q P 13 410 S Q P E E S V G L 13 419 R A E
G H P D S L 13 9 M W G P E A W L L 12 11 G P E A W L L L L 12 73 G
E G A Q E L A L 12 82 L H S K Y G L H V 12 88 L H V S P A Y E G 12
105 R N P L D G S V L 12 133 P A G S F Q A R L 12 213 S R S M N G Q
P L 12 382 M T Q K Y E E E L 12 384 Q K Y E E E L T L 12 422 G H P
D S L K D N 12 452 R E I E T Q T E L 12 453 E I E T Q T E L L 12
484 H F V Q E N G T L 12 10 W G P E A W L L L 11 12 P E A W L L L L
L 11 13 E A W L L L L L L 11 42 V V L G Q D A K L 11 80 A L L H S K
Y G L 11 157 P A L E E G Q G L 11 223 C V V S H P G L L 11 226 S H
P G L L Q D Q 11 240 L H V S F L A E A 11 315 C H V S N E F S S 11
352 V V G V I A A L L 11 355 V I A A L L F C L 11 401 L H S H H T D
P R 11 440 P E G R S Y S T L 11 483 N H F V Q E N G T 11 14 A W L L
L L L L L 10 124 E Y E C R V S T F 10 202 A A V T S E F H L 10 232
Q D Q R I T H I L 10 236 I T H I L H V S F 10 250 V R G L E D Q N L
10 260 H I G R E G A M L 10 263 R E G A M L K C L 10 281 N W T R L
D G P L 10 363 L L V V V V V L M 10 474 Q D E G I K Q A M 10
V2-HLA-B1510-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 1 G Q D A K L P C L 12
V7-HLA-B1510-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 1 S H H T D P R S Q 13 2 H H T D P R S Q S 13
V9-HLA-B1510-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 44 S H Y V A Q A G L 21 74 H H C A C F E S F
16 46 Y V A Q A G L E L 14 101 L G L L K V R P L 13 32 I Y F Y F Y
F F L 12 58 S N P P A S A S L 12 63 S A S L V A G T L 12 81 S F T K
R K K K L 12 96 I Q C L L L G L L 12 2 R R E L L A G I L 11 19 L F
F F L P F P L 11 22 F L P F P L V V F 11 23 L P F P L V V F F 11 47
V A Q A G L E L L 11 73 V H H C A C F E S 11 91 K A F R F I Q C L
11 110 Q H Q G V N S C D 11 3 R E L L A G I L L 10 11 L R I T F N F
F L 10 15 F N F F L F F F L 10 92 A F R F I Q C L L 10 93 F R F I Q
C L L L 10 95 F I Q C L L L G L 10 V10-HLA-B1510-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 21; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 9 G T S
D V V T V V 6 7 E L G T S D V V T 5 6 G E L G T S D V V 4 8 L G T S
D V V T V 4 1 G R C P A G E L G 3 3 C P A G E L G T S 3 5 A G E L G
T S D V 2 V11-HLA-B1510-9mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 23; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 8 V M V P P L P S L 14 5 R L R V M V
P P L 13 1 Q A R L R L R V M 10 V12-HLA-B1510-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 25; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 6 P E G
C S Y S T L 11 V13-HLA-B1510-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 27; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 2 Q V T V D V L A D 3 7 V
L A D P Q E D S 3 1 S Q V T V D V L A 2 4 T V D V L A D P Q 2 6 D V
L A D P Q E D 2 8 L A D P Q E D S G 2 3 V T V D V L A D P 1 5 V D V
L A D P Q E 1 9 A D P Q E D S G K 1 V14-HLA-B1510-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 29; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 3 S N P
P A S A S L 12 8 S A S L V A G T L 12
[1044]
40TABLE XXX Pos 1 2 3 4 5 6 7 8 9 score
V1-HLA-B2705-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight 393 T R E N S I R R L 26 250 V R G L E D Q N L
25 452 R E I E T Q T E L 22 135 G S F Q A R L R L 21 213 S R S M N
G Q P L 20 377 R K A Q Q M T Q K 19 42 V V L G Q D A K L 18 97 R V
E Q P P P P R 18 262 G R E G A M L K C 18 351 V V V G V I A A L 18
376 R R K A Q Q M T Q 18 399 R R L H S H H T D 18 14 A W L L L L L
L L 17 17 L L L L L L A S F 17 105 R N P L D G S V L 17 142 R L R V
L V P P L 17 200 R S A A V T S E F 17 206 S E F H L V P S R 17 294
R V D G D T L G F 17 297 G D T L G F P P L 17 419 R A E G H P D S L
17 498 G N G I Y I N G R 17 41 T V V L G Q D A K 16 45 G Q D A K L
P C F 16 80 A L L H S K Y G L 16 96 G R V E Q P P P P 16 106 N P L
D G S V L L 16 145 V L V P P L P S L 16 234 Q R I T H I L H V 16
243 S F L A E A S V R 16 261 I G R E G A M L K 16 293 V R V D G D T
L G 16 301 G F P P L T T E H 16 337 E D S G K Q V D L 16 362 C L L
V V V V V L 16 384 Q K Y E E E L T L 16 442 G R S Y S T L T T 16
476 E G I K Q A M N H 16 477 G I K Q A M N H F 16 484 H F V Q E N G
T L 16 11 G P E A W L L L L 15 20 L L L A S F T G R 15 61 Q V G Q V
A W A R 15 71 D A G E G A Q E L 15 74 E G A Q E L A L L 15 75 G A Q
E L A L L H 15 77 Q E L A L L H S K 15 10 P L D G S V L L R 15 133
P A G S F Q A R L 15 139 A R L R L R V L V 15 141 L R L R V L V P P
15 188 K G T T S S R S F 15 189 G T T S S R S F K 15 227 H P G L L
Q D Q R 15 237 T H I L H V S F L 15 263 R E G A M L K C L 15 283 T
R L D G P L P S 15 333 L D P Q E D S G K 15 365 V V V V V L M S R
15 392 L T R E N S I R R 15 466 G R A E E E E D Q 15 492 L R A K P
T G N G 15 501 I Y I N G R G H L 15 8 E M W G P E A W L 14 9 M W G
P E A W L L 14 13 E A W L L L L L L 14 27 G R C P A G E L E 14 73 G
E G A Q E L A L 14 104 P R N P L D G S V 14 114 L R N A V Q A D E
14 120 A D E G E Y E C R 14 143 L R V L V P P L P 14 151 P S L N P
G P A L 14 157 P A L E E G Q G L 14 159 L E E G Q G L T L 14 186 E
V K G T T S S R 14 193 S R S F K H S R S 14 199 S R S A A V T S E
14 236 I T H I L H V S F 14 277 P P S Y N W T R L 14 286 D G P L P
S G V R 14 292 G V R V D G D T L 14 313 Y V C H V S N E F 14 323 S
R D S Q V T V D 14 368 V V L M S R Y H R 14 375 H R R K A Q Q M T
14 378 K A Q Q M T Q K Y 14 386 Y E E E L T L T R 14 408 P R S Q P
E E S V 14 410 S Q P E E S V G L 14 418 L R A E G H P D S 14 420 A
E G H P D S L K 14 444 S Y S T L T T V R 14 459 E L L S P G S G R
14 1 M P L S L G A E M 13 12 P E A W L L L L L 13 26 T G R C P A G
E L 13 36 T S D V V T V V L 13 78 E L A L L H S K Y 13 86 Y G L H V
S P A Y 13 100 Q P P P P R N P L 13 124 E Y E C R V S T F 13 129 V
S T F P A G S F 13 132 F P A G S F Q A R 13 138 Q A R L R L R V L
13 202 A A V T S E F H L 13 208 F H L V P S R S M 13 219 Q P L T C
V V S H 13 222 T C V V S H P G L 13 231 L Q D Q R I T H I 13 252 G
L E D Q N L W H 13 272 S E G Q P P P S Y 13 276 P P P S Y N W T R
13 316 H V S N E F S S R 13 352 V V G V I A A L L 13 353 V G V I A
A L L F 13 356 I A A L L F C L L 13 366 V V V V L M S R Y 13 382 M
T Q K Y E E E L 13 391 T L T R E N S I R 13 394 R E N S I R R L H
13 398 I R R L H S H H T 13 411 Q P E E S V G L R 13 428 K D N S S
C S V M 13 440 P E G R S Y S T L 13 485 F V Q E N G T L R 13 487 Q
E N G T L R A K 13 500 G I Y I N G R G H 13 10 W G P E A W L L L 12
47 D A K L P C F Y R 12 54 Y R G D S G E Q V 12 68 A R V D A G E G
A 12 127 C R V S T F P A G 12 134 A G S F Q A R L R 12 192 S S R S
F K H S R 12 228 P G L L Q D Q R I 12 245 L A E A S V R G L 12 255
D Q N L W H I G R 12 259 W H I G R E G A M 12 260 H I G R E G A M L
12 281 N W T R L D G P L 12 308 E H S G I Y V C H 12 325 D S Q V T
V D V L 12 355 V I A A L L F C L 12 363 L L V V V V V L M 12 369 V
L M S R Y H R R 12 370 L M S R Y H R R K 12 372 S R Y H R R K A Q
12 396 N S I R R L H S H 12 435 V M S E E P E G R 12 451 V R E I E
T Q T E 12 471 E E D Q D E G I K 12 474 Q D E G I K Q A M 12 493 R
A K P T G N G I 12 494 A K P T G N G I Y 12
V2-HLA-B2705-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 1 G Q D A K L P C L 16 3 D A K L P C L Y R 13
2 Q D A K L P C L Y 11 4 A K L P C L Y R G 8
V7-HLA-B2705-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 6 P R S Q S E E P E 13 8 S Q S E E P E G R 12
7 R S Q S E E P E G 7 V9-HLA-B2705-9mers-191P4D12B Each peptide is
a portion of SEQ ID NO: 19; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 2 R R E L L A G I L 27 93
F R F I Q C L L L 24 11 L R I T F N F F L 23 120 E R G Y F Q G I F
22 1 M R R E L L A G I 20 77 A C F E S F T K R 20 87 K K L K K A F
R F 20 3 R E L L A G I L L 18 1 E L L A G I L L R 18 84 K R K K K L
K K A 18 85 R K K K L K K A F 18 91 K A F R F I Q C L 18 7 A G I L
L R I T F 17 23 L P F P L V V F F 17 83 T K R K K K L K K 17 99 L L
L G L L K V R 17 9 I L L R I T F N F 16 80 E S F T K R K K K 16 86
K K K L K K A F R 16 13 I T F N F F L F F 15 44 S H Y V A Q A G L
15 81 S F T K R K K K L 15 97 Q C L L L G L L K 15 101 L G L L K V
R P L 15 113 G V N S C D C E R 15 121 R G Y F Q G I F M 15 12 R I T
F N F F L F 14 15 F N F F L F F F L 14 19 L F F F L P F P L 14 22 F
L P F P L V V F 14 28 V V F F I Y F Y F 14 32 I Y F Y F Y F F L 14
37 Y F F L E M E S H 14 46 Y V A Q A G L E L 14 58 S N P P A S A S
L 14 63 S A S L V A G T L 14 92 A F R F I Q C L L 14 96 I Q C L L L
G L L 14 5 L L A G I L L R I 13 17 F F L F F F L P F 13 27 L V V F
F I Y F Y 13 31 F I Y F Y F Y F F 13 34 F Y F Y F F L E M 13 47 V A
Q A G L E L L 13 66 L V A G T L S V H 13 76 C A C F E S F T K 13 79
F E S F T K R K K 13 95 F I Q C L L L G L 13 122 G Y F Q G I F M Q
13 V10-HLA-B2705-9mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 21; each start position is specified, the length of peptide
is 9 amino acids, and the end position for each peptide is the
start position plus eight. 1 G R C P A G E L G 14 6 G E L G T S D V
V 9 9 G T S D V V T V V 8 2 R C P A G E L G T 7 3 C P A G E L G T S
5 4 P A G E L G T S D 5 5 A G E L G T S D V 5
V11-HLA-B2705-9mers-191- P4D12B Each peptide is a portion of SEQ ID
NO: 23; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 5 R L R V M V P P L 16 8 V M V P P L P S L 16
2 A R L R L R V M V 15 4 L R L R V M V P P 14 6 L R V M V P P L P
13 1 Q A R L R L R V M 11 3 R L R L R V M V P 8
V12-HLA-B2705-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 25; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 6 P E G C S Y S T L 13 3 S E E P E G C S Y 11
8 G C S Y S T L T T 6 9 C S Y S T L T T V 6
V13-HLA-B2705-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 9 A D P Q E D S G K 16
V14-HLA-B2705-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 3 S N P P A S A S L 14 8 S A S L V A G T L 14
1 G S S N P P A S A 6
[1045]
41TABLE XXXI Start Subsequence Score V1-HLA-B2709-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 3; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 139
ARLRLRVLV 22 250 VRGLEDQNL 21 393 TRENSIRRL 21 213 SRSMNGQPL 20 234
QRITHILHV 20 54 YRGDSGEQV 19 104 PRNPLDGSV 19 408 PRSQPEESV 18 135
GSFQARLRL 17 142 RLRVLVPPL 16 287 GPLPSGVRV 16 399 RRLHSHHTD 16 96
GRVEQPPPP 15 105 RNPLDGSVL 15 297 GDTLGFPPL 15 443 RSYSTLTTV 15 452
REIETQTEL 15 11 GPEAWLLLL 14 14 AWLLLLLLL 14 27 GRCPAGELE 14 73
GEGAQELAL 14 80 ALLHSKYGL 14 262 GREGAMLKC 14 263 REGAMLKCL 14 292
GVRVDGDTL 14 294 RVDGDTLGF 14 362 CLLVVVVVL 14 376 RRKAQQMTQ 14 419
RAEGHPDSL 14 442 GRSYSTLTT 14 32 GELETSDVV 13 34 LETSDVVTV 13 106
NPLDGSVLL 13 127 CRVSTFPAG 13 141 LRLRVLVPP 13 145 VLVPPLPSL 13 151
PSLNPGPAL 13 283 TRLDGPLPS 13 324 RDSQVTVDV 13 384 QKYEEELTL 13 466
GRAEEEEDQ 13 493 RAKPTGNGI 13 9 MWGPEAWLL 12 42 VVLGQDAKL 12 45
GQDAKLPCF 12 68 ARVDAGEGA 12 110 GSVLLRNAV 12 133 PAGSFQARL 12 143
LRVLVPPLP 12 157 PALEEGQGL 12 173 AEGSPAPSV 12 200 RSAAVTSEF 12 202
AAVTSEFHL 12 222 TCVVSHPGL 12 223 CVVSHPGLL 12 237 THILHVSFL 12 323
SRDSQVTVD 12 352 VVGVIAALL 12 357 AALLFCLLV 12 358 ALLFCLLVV 12 361
FCLLVVVVV 12 372 SRYHRRKAQ 12 501 IYINGRGHL 12 1 MPLSLGAEM 11 10
WGPEAWLLL 11 12 PEAWLLLLL 11 13 EAWLLLLLL 11 26 TGRCPAGEL 11 36
TSDVVTVVL 11 71 DAGEGAQEL 11 100 QPPPPRNPL 11 159 LEEGQGLTL 11 188
KGTTSSRSF 11 193 SRSFKHSRS 11 199 SRSAAVTSE 11 203 AVTSEFHLV 11 228
PGLLQDQRI 11 232 QDQRITHIL 11 245 LAEASVRGL 11 277 PPSYNWTRL 11 281
NWTRLDGPL 11 293 VRVDGDTLG 11 325 DSQVTVDVL 11 337 EDSGKQVDL 11 343
VDLVSASVV 11 344 DLVSASVVV 11 348 ASVVVVGVI 11 351 VVVGVIAAL 11 353
VGVIAALLF 11 356 IAALLFCLL 11 359 LLFCLLVVV 11 363 LLVVVVVLM 11 398
IRRLHSHHT 11 410 SQPEESVGL 11 418 LRAEGHPDS 11 428 KDNSSCSVM 11 446
STLTTVREI 11 477 GIKQAMNHF 11 484 HFVQENGTL 11 492 LRAKPTGNG 11 495
KPTGNGIYI 11 8 EMWGPEAWL 10 17 LLLLLLASF 10 57 DSGEQVGQV 10 74
EGAQELALL 10 114 LRNAVQADE 10 129 VSTFPAGSF 10 137 FQARLRLRV 10 138
QARLRLRVL 10 208 FHLVPSRSM 10 236 ITHILHVSF 10 242 VSFLAEASV 10 260
HIGREGAML 10 320 EFSSRDSQV 10 345 LVSASVVVV 10 347 SASVVVVGV 10 355
VIAALLFCL 10 360 LFCLLVVVV 10 374 YHRRKAQQM 10 375 HRRKAQQMT 10 382
MTQKYEEEL 10 390 LTLTRENSI 10 440 PEGRSYSTL 10 451 VREIETQTE 10 453
EIETQTELL 10 V2-HLA-B2709-9mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 5; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 1 GQDAKLPCL 14 4 AKLPCLYRG 6
V7-HLA-B2709-9mers- 191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 6 PRSQSEEPE 10 7 RSQSEEPEG 6
V9-HLA-B2709-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 2 RRELLAGIL 25 93 FRFIQCLLL 23 11 LRITFNFFL 21
1 MRRELLAGI 18 106 VRPLQHQGV 18 120 ERGYFQGIF 18 3 RELLAGILL 16 87
KKLKKAFRF 14 91 KAFRFIQCL 14 121 RGYFQGIFM 14 9 LLRITFNF 13 12
RITFNFFLF 13 23 LPFPLVVFF 13 32 IYFYFYFFL 13 101 LGLLKVRPL 13 13
ITFNFFLFF 12 15 FNFFLFFFL 12 19 LFFFLPFPL 12 21 FFLPFPLVV 12 44
SHYVAQAGL 12 84 KRKKKLKKA 12 85 RKKKLKKAF 12 92 AFRFIQCLL 12
V10-HLA-B2709-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 1 GRCPAGELG 14 6 GELGTSDVV 13 8 LGTSDVVTV 13
V10-HLA-B2709-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 9 GTSDVVTVV 12 5 AGELGTSDV 9
V11-HLA-B2709-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 23; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 2 ARLRLRVMV 22 5 RLRVMVPPL 16 4 LRLRVMVPP 13 8
VMVPPLPSL 13 6 LRVMVPPLP 12 V12-HLA-B2709-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 25; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 9
CSYSTLTTV 11 6 PEGCSYSTL 10 V13-HLA-B2709-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 27; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 2
QVTVDVLAD 4 5 VDVLADPQE 3 6 DVLADPQED 3 1 SQVTVDVLA 2 3 VTVDVLADP 1
4 TVDVLADPQ 1 8 LADPQEDSG 1 9 ADPQEDSGK 1
V14-HLA-B2709-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 3 SNPPASASL 11 8 SASLVAGTL 11 4 NPPASASLV
9
[1046]
42TABLE XXXII Pos 1 2 3 4 5 6 7 8 9 score
V1-HLA-B44Q2-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 7 A E M W G P E A W 27 437 S E E P E G R S Y
25 12 P E A W L L L L L 23 59 G E Q V G Q V A W 23 73 G E G A Q E L
A L 23 159 L E E G Q G L T L 23 263 R E G A M L K C L 23 452 R E I
E T Q T E L 23 272 S E G Q P P P S Y 22 440 P E G R S Y S T L 22
253 L E D Q N L W H I 21 470 E E E D Q D E G I 21 14 A W L L L L L
L L 18 413 E E S V G L R A E 17 13 E A W L L L L L L 16 100 Q P P P
P R N P L 16 351 V V V G V I A A L 16 388 E E L T L T R E N 16 9 M
W G P E A W L L 15 106 N P L D G S V L L 15 124 E Y E C R V S T F
15 138 Q A R L R L R V L 15 237 T H I L H V S F L 15 246 A E A S V
R G L E 15 337 E D S G K Q V D L 15 393 T R E N S I R R L 15 453 E
I E T Q T E L L 15 487 Q E N G T L R A K 15 494 A K P T G N G I Y
15 501 I Y I N G R G H L 15 36 T S D V V T V V L 14 74 E G A Q E L
A L L 14 78 E L A L L H S K Y 14 80 A L L H S K Y G L 14 98 V E Q P
P P P R N 14 135 G S F Q A R L R L 14 145 V L V P P L P S L 14 151
P S L N P G P A L 14 160 E E G Q G L T L A 14 173 A E G S P A P S V
14 202 A A V T S E F H L 14 206 S E F H L V P S R 14 232 Q D Q R I
T H I L 14 274 G Q P P P S Y N W 14 294 R V D G D T L G F 14 307 T
E H S G I Y V C 14 319 N E F S S R D S Q 14 362 C L L V V V V V L
14 387 E E E L T L T R E 14 394 R E N S I R R L H 14 420 A E G H P
D S L K 14 438 E E P E G R S Y S 14 2 P L S L G A E M W 13 8 E M W
G P E A W L 13 10 W G P E A W L L L 13 11 G P E A W L L L L 13 17 L
L L L L L A S F 13 34 L E T S D V V T V 13 42 V V L G Q D A K L 13
77 Q E L A L L H S K 13 86 Y G L H V S P A Y 13 105 R N P L D G S V
L 13 117 A V Q A D E G E Y 13 175 G S P A P S V T W 13 188 K G T T
S S R S F 13 213 S R S M N G Q P L 13 231 L Q D Q R I T H I 13 251
R G L E D Q N L W 13 348 A S V V V V G V I 13 352 V V G V I A A L L
13 353 V G V I A A L L F 13 356 I A A L L F C L L 13 378 K A Q Q M
T Q K Y 13 386 Y E E E L T L T R 13 410 S Q P E E S V G L 13 446 S
T L T T V R E I 13 458 T E L L S P G S G 13 468 A E E E E D Q D E
13 471 E E D Q D E G I K 13 V2-HLA-B4402-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 5; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 1 G Q D
A K L P C L 12 2 Q D A K L P C L Y 12 4 A K L P C L Y R G 8
V7-HLA-B4402-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 3 H T D P R S Q S E 5 1 S H H T D P R S Q 4 2
H H T D P R S Q S 3 8 S Q S E E P E G R 3 4 T D P R S Q S E E 2
V9-HLA-B4402-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 3 R E L L A G I L L 24 7 A G I L L R I T F 20
119 C E R G Y F Q G I 20 23 L P F P L V V F F 17 91 K A F R F I Q C
L 17 13 I T F N F F L F F 15 58 S N P P A S A S L 15 63 S A S L V A
G T L 15 81 S F T K R K K K L 15 92 A F R F I Q C L L 15 9 I L L R
I T F N F 14 11 L R I T F N F F L 14 22 F L P F P L V V F 14 85 R K
K K L K K A F 14 93 F R F I Q C L L L 14 101 L G L L K V R P L 14
12 R I T F N F F L F 13 15 F N F F L F F F L 13 17 F F L F F F L P
F 13 19 L F F F L P F P L 13 27 L V V F F I Y F Y 13 28 V V F F I Y
F Y F 13 29 V F F I Y F Y F Y 13 30 F F I Y F Y F Y F 13 42 M E S H
Y V A Q A 13 79 F E S F T K R K K 13 87 K K L K K A F R F 13 96 I Q
C L L L G L L 13 115 N S C D C E R G Y 13 116 S C D C E R G Y F 13
126 G I F M Q A A P W 13 2 R R E L L A G I L 12 5 L L A G I L L R I
12 10 L L R I T F N F F 12 25 F P L V V F F I Y 12 26 P L V V F F I
Y F 12 32 I Y F Y F Y F F L 12 40 L E M E S H Y V A 12 47 V A Q A G
L E L L 12 52 L E L L G S S N P 12 95 F I Q C L L L G L 12 120 E R
G Y F Q G I F 12 14 T F N F F L F F F 11 24 P F P L V V F F I 11 31
F I Y F Y F Y F F 11 38 F F L E M E S H Y 11 44 S H Y V A Q A G L
11 46 Y V A Q A G L E L 11 74 H H C A C F E S F 11 88 K L K K A F R
F I 11 V10-HLA-B4402-9mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 21; each start position is specified, the length of
peptide is 9 amino acids, and the end position for each peptide is
the start position plus eight. 6 G E L G T S D V V 13
V11-HLA-B4402-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 23; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 8 V M V P P L P S L 14 5 R L R V M V P P L 11
2 A R L R L R V M V 7 9 M V P P L P S L N 6
V12-HLA-B4402-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 25; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 3 S E E P E G C S Y 24 6 P E G C S Y S T L 21
4 E E P E G C S Y S 13 V13-HLA-B4402-9mers-191- P4D12B Each peptide
is a portion of SEQ ID NO: 27; each start position is specified,
the length of peptide is 9 amino acids, and the end position for
each peptide is the start position plus eight. 1 S Q V T V D V L A
4 2 Q V T V D V L A D 4 8 L A D P Q E D S G 4 9 A D P Q E D S G K 4
3 V T V D V L A D P 4 T V D V L A D P Q 2 5 V D V L A D P Q E 2 6 D
V L A D P Q E D 2 7 V L A D P Q E D S 1
V14-HLA-B4402-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 3 S N P P A S A S L 15 8 S A S L V A G T L 15
2 S S N P P A S A S 7
[1047]
43TABLE XXXIIII Pos 1 2 3 4 5 6 7 8 9 score
V1-HLA-B5101-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 71 D A G E G A Q E L 23 245 L A E A S V R G L
23 287 G P L P S G V R V 23 347 S A S V V V V G V 23 493 R A K P T
G N G I 22 495 K P T G N G I Y I 22 106 N P L D G S V L L 21 138 Q
A R L R L R V L 21 357 A A L L F C L L V 21 157 P A L E E G Q G L
20 11 G P E A W L L L L 19 13 E A W L L L L L L 19 202 A A V T S E
F H L 19 228 P G L L Q D Q R I 19 356 I A A L L F C L L 19 361 F C
L L V V V V V 19 100 Q P P P P R N P L 18 217 N G Q P L T C V V 18
277 P P S Y N W T R L 18 334 D P Q E D S G K Q 18 345 L V S A S V V
V V 18 419 R A E G H P D S L 18 35 E T S D V V T V V 17 92 P A Y E
G R V E Q 17 133 P A G S F Q A R L 17 348 A S V V V V G V I 17 443
R S Y S T L T T V 17 446 S T L T T V R E I 17 10 W G P E A W L L L
16 32 G E L E T S D V V 16 57 D S G E Q V G Q V 16 62 V G Q V A W A
R V 16 121 D E G E Y E C R V 16 219 Q P L T C V V S H 16 289 L P S
G V R V D G 16 325 D S Q V T V D V L 16 343 V D L V S A S V V 16
344 D L V S A S V V V 16 359 L L F C L L V V V 16 360 L F C L L V V
V V 16 362 C L L V V V V V L 16 390 L T L T R E N S I 16 34 L E T S
D V V T V 15 65 V A W A R V D A G 15 79 L A L L H S K Y G 15 148 P
P L P S L N P G 15 231 L Q D Q R I T H I 15 276 P P P S Y N W T R
15 338 D S G K Q V D L V 15 358 A L L F C L L V V 15 384 Q K Y E E
E L T L 15 407 D P R S Q P E E S 15 411 Q P E E S V G L R 15 22 L A
S F T G R C P 14 26 T G R C P A G E L 14 29 C P A G E L E T S 14 31
A G E L E T S D V 14 47 D A K L P C F Y R 14 75 G A Q E L A L L H
14 82 L H S K Y G L H V 14 91 S P A Y E G R V E 14 132 F P A G S F
Q A R 14 172 T A E G S P A P S 14 176 S P A P S V T W D 14 253 L E
D Q N L W H I 14 286 D G P L P S G V R 14 302 F P P L T T E H S 14
303 P P L T T E H S G 14 1 M P L S L G A E M 13 30 P A G E L E T S
D 13 36 T S D V V T V V L 13 50 L P C F Y R G D S 13 74 E G A Q E L
A L L 13 90 V S P A Y E G R V 13 102 P P P R N P L D G 13 147 V P P
L P S L N P 13 150 L P S L N P G P A 13 177 P A P S V T W D T 13
178 A P S V T W D T E 13 211 V P S R S M N G Q 13 275 Q P P P S Y N
W T 13 300 L G F P P L T T E 13 322 S S R D S Q V T V 13 378 K A Q
Q M T Q K Y 13 478 I K Q A M N H F V 13 42 V V L G Q D A K L 12 54
Y R G D S G E Q V 12 86 Y G L H V S P A Y 12 101 P P P P R N P L D
12 1091 D G S V L L R N A 12 119 Q A D E G E Y E C 12 154 N P G P A
L E E G 12 159 L E E G Q G L T L 12 167 L A A S C T A E G 12 168 A
A S C T A E G S 12 234 Q R I T H I L H V 12 265 G A M L K C L S E
12 309 H S G I Y V C H V 12 339 S G K Q V D L V S 12 467 R A E E E
E D Q D 12 480 Q A M N H F V Q E 12 5 L G A E M W G P E 11 58 S G E
Q V G Q V A 11 67 W A R V D A G E G 11 103 P P R N P L D G S 11 116
N A V Q A D E G E 11 137 F Q A R L R L R V 11 139 A R L R L R V L V
11 201 S A A V T S E F H 11 216 M N G Q P L T C V 11 247 E A S V R
G L E D 11 251 R G L E D Q N L W 11 261 I G R E G A M L K 11 285 L
D G P L P S G V 11 296 D G D T L G F P P 11 304 P L T T E H S G I
11 306 T T E H S G I Y V 11 310 S G I Y V C H V S 11 324 R D S Q V
T V D V 11 335 P Q E D S G K Q V 11 351 V V V G V I A A L 11 393 T
R E N S I R R L 11 427 L K D N S S C S V 11 439 E P E G R S Y S T
11 470 E E E D Q D E G I 11 502 Y I N G R G H L V 11
V2-HLA-B5101-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is 9
amino acids, and the end position for each peptide is the start
position plus eight. 3 D A K L P C L Y R 1 6 L P C L Y R G D S 13
11 IL G Q D A K L P C 9 V7-HLA-B5101-9mers-191P4D12B Each peptide
is a portion of SEQ ID NO: 15; each start position is specified,
the length of peptide is 9 amino acids, and the end position for
each peptide is the start position plus eight. 5 D P R S Q S E E P
14 V9-HLA-B5101-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 59 N P P A S A S L V 23 63 S A S L V A G T L
21 101 L G L L K V R P L 20 47 V A Q A G L E L L 19 911 K A F R F I
Q C L 18 5 L L A G I L L R I 16 21 F F L P F P L V V 16 23 L P F P
L V V F F 16 25 F P L V V F F I Y 16 24 P F P L V V F F I 15 107 R
P L Q H Q G V N 15 1 M R R E L L A G I 14 6 L A G I L L R I T 14 60
P P A S A S L V A 14 61 P A S A S L V A G 14 67 V A G T L S V H H
14 98 C L L L G L L K V 14 88 K L K K A F R F I 13 119 C E R G Y F
Q G I 13 49 Q A G L E L L G S 12 76 C A C F E S F T K 12 20 F F F L
P F P L V 11 50 A G L E L L G S S 11 121 R G Y F Q G I F M 11
V10-HLA-B5101-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 8 L G T S D V V T V 21 9 G T S D V V T V V 17
6 G E L G T S D V V 15 3 C P A G E L G T S 14 5 A G E L G T S D V
14 4 P A G E L G T S D 13 V11-HLA-B5101-9mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 23; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 1 Q A R
L R L R V M 15 2 A R L R L R V M V 11 5 R L R V M V P P L 9 8 V M V
P P L P S L 8 4 L R L R V M V P P 7 V12-HLA-B5101-9mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 25; each start position is
specified, the length of peptide is 9 amino acids, and the end
position for each peptide is the start position plus eight. 9 C S Y
S T L T T V 17 5 E P E G C S Y S T 11 6 P E G C S Y S T L 9 7 E G C
S Y S T L T 8 V13-HLA-B5101-9mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 27; each start position is specified, the
length of peptide is 9 amino acids, and the end position for each
peptide is the start position plus eight. 8 L A D P Q E D S G 12 6
D V L A D P Q E D 8 3 V T V D V L A D P 5
V14-HLA-B5101-9mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
9 amino acids, and the end position for each peptide is the start
position plus eight. 4 N P P A S A S L V 23 8 S A S L V A G T L 21
5 P P A S A S L V A 14 6 P A S A S L V A G 14
[1048]
44TABLE XXXIV Pos 1 2 3 4 5 6 7 8 9 0 score
V1-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
3; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 271 L S E G Q P P P S Y 30 436 M S E E P E G R
S Y 30 45 G Q D A K L P C F Y 25 405 H T D P R S Q P E E 20 493 R A
K P T G N G I Y 20 158 A L E E G Q G L T L 19 11 G P E A W L L L L
L 18 72 A G E G A Q E L A L 18 107 P L D G S V L L R N 18 453 E I E
T Q T E L L S 18 36 T S D V V T V V L G 17 77 Q E L A L L H S K Y
17 306 T T E H S G I Y V C 17 377 R K A Q Q M T Q K Y 17 411 Q P E
E S V G L R A 17 437 S E E P E G R S Y S 17 471 E E D Q D E G I K Q
17 184 D T E V K G T T S S 16 304 P L T T E H S G I Y 16 332 V L D
P Q E D S G K 16 365 V V V V V L M S R Y 16 385 K Y E E E L T L T R
16 457 Q T E L L S P G S G 16 85 K Y G L H V S P A Y 15 116 N A V Q
A D E G E Y 15 205 T S E F H L V P S R 15
V2-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
5; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 2 G Q D A K L P C L Y 27
V7-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
15; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 4 H T D P R S Q S E E 20
V9-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
19; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 28 V V F F I Y F Y F Y 19 24 P F P L V V F F I
Y 18 2 R R E L L A G I L L 17 37 Y F F L E M E S H Y 17 26 P L V V
F F I Y F Y 16 114 V N S C D C E R G Y 16 82 F T K R K K K L K K 15
39 F L E M E S H Y V A 13 116 S C D C E R G Y F Q 13 118 D C E R G
Y F Q G I 13 78 C F E S F T K R K K 12 33 Y F Y F Y F F L E M 11 41
E M E S H Y V A Q A 11 51 G L E L L G S S N P 11 64 A S L V A G T L
S V 11 57 S S N P P A S A S L 10 12 R I T F N F F L F F 9 16 N F F
L F F F L P F 9 47 V A Q A G L E L L G 9 92 A F R F I Q C L L L 9
93 F R F I Q C L L L G 9 96 I Q C L L L G L L K
V10-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
21; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 6 A G E L G T S D V V 12 2 G R C P A G E L G T
10 10 G T S D V V T V V L 7 V11-HLA-A1-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 23; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 10 M V P
P L P S L N P 10 9 V M V P P L P S L N 7 7 L R V M V P P L P S 6
V12-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
25; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 3 M S E E P E G C S Y 30 4 S E E P E G C S Y S
16 V13-HLA-A1-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 9 L A D P Q E D S G K 14 5 T V D V L A D P Q E
10 2 S Q V T V D V L A D 9 4 V T V D V L A D P Q 7 1 D S Q V T V D
V L A 6 V14-HLA-A1-10mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 29; each start position is specified, the length of
peptide is 10 amino acids, and the end position for each peptide is
the start position plus nine. 10 A S L V A G T L S V 11 3 S S N P P
A S A S L 10 4 S N P P A S A S L V 8 5 N P P A S A S L V A 7 8 A S
A S L V A G T L 5
[1049]
45TABLE XXXV Pos 1 2 3 4 5 6 7 8 9 0 score
V1-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 244 F L A E A S V R G L 3 358 A L L F C L L V V
V 29 359 L L F C L L V V V V 29 215 S M N G Q P L T C V 27 158 A L
E E G Q G L T L 26 230 L L Q D Q R I T H I 25 344 D L V S A S V V V
V 25 33 E L E T S D V V T V 24 239 I L H V S F L A E A 24 426 S L K
D N S S C S V 24 81 L L H S K Y G L H V 23 144 R V L V P P L P S L
23 252 G L E D Q N L W H I 23 284 R L D G P L P S G V 23 357 A A L
L F C L L V V 23 16 L L L L L L L A S F 22 350 V V V V G V I A A L
22 362 C L L V V V V V L M 22 392 L T R E N S I R R L 22 354 G V I
A A L L F C L 21 355 V I A A L L F C L L 21 79 L A L L H S K Y G L
20 236 I T H I L H V S F L 20 346 V S A S V V V V G V 20 500 G I Y
I N G R G H L 20 141 L R L R V L V P P L 19 351 V V V G V I A A L L
19 356 I A A L L F C L L V 19 361 F C L L V V V V V L 19 381 Q M T
Q K Y E E E L 19 477 G I K Q A M N H F V 19 8 E M W G P E A W L L
18 15 W L L L L L L L A S 18 17 L L L L L L A S F T 18 41 T V V L G
Q D A K L 18 112 V L L R N A V Q A D 18 152 S L N P G P A L E E 18
172 T A E G S P A P S V 18 201 S A A V T S E F H L 18 221 L T C V V
S H P G L 18 249 S V R G L E D Q N L 18 347 S A S V V V V G V I 18
360 L F C L L V V V V V 18 418 L R A E G H P D S L 18 10 W G P E A
W L L L L 17 13 E A W L L L L L L L 17 25 F T G R C P A G E L 17 56
G D S G E Q V G Q V 17 70 V D A G E G A Q E L 17 73 G E G A Q E L A
L L 17 132 F P A G S F Q A R L 17 137 F Q A R L R L R V L 17 202 A
A V T S E F H L V 17 241 H V S F L A E A S V 17 305 L T T E H S G I
Y V 17 363 L L V V V V V L M S 17 389 E L T L T R E N S I 17 18 L L
L L L A S F T G 16 61 Q V G Q V A W A R V 16 89 H V S P A Y E G R V
16 138 Q A R L R L R V L V 16 140 R L R L R V L V P P 16 164 G L T
L A A S C T A 16 166 T L A A S C T A E G 16 257 N I W H I G R E G A
16 259 W H I G R E G A M L 16 341 K Q V D L V S A S V 16 370 L M S
R Y H R R K A 16 442 G R S Y S I L T T V 16 7 A E M W G P E A W L
15 11 G P E A W L L L L L 15 19 L L L L A S F T G R 15 34 L E T S D
V V T V V 15 72 A G E G A Q E L A L 15 181 V T W D T E V K G T 15
229 G L L Q D Q R I T H 15 262 G R E G A M L K C L 15 299 T L G F P
P L T T E 15 321 F S S R D S Q V T V 15 343 V D L V S A S V V V 15
349 S V V V V G V I A A 15 397 S I R R L H S H H T 15 409 R S Q P E
E S V G L 15 445 Y S T L T V R E 15 447 T L T T V R E I E T 15 460
L L S P G S G R A E 15 501 I Y I N G R G H L V 15 12 P E A W L L L
L L L 14 20 L L L A S F T G R C 14 21 L L A S F T G R C P 14 35 E T
S D V V T V V L 14 80 A L L H S K Y G L H 14 87 G L H V S P A Y E G
14 107 P L D G S V L L R N 14 111 S V L L R N A V Q A 14 113 L L R
N A V Q A D E 14 150 L P S L N P G P A L 14 156 G P A L E E G Q G L
14 178 A P S V T W D T E V 14 195 S F K H S R S A A V 14 233 D Q R
I T H I L H V 14 291 S G V R V D G D T L 14 298 D T L G F P P L T T
14 311 G I Y V C H V S N E 14 323 S R D S Q V T V D V 14 324 R D S
Q V T V D V L 14 332 V L D P Q E D S G K 14 342 Q V D L V S A S V V
14 452 R E I E T Q T E L L 14 492 L R A K P T G N G I 14
V2-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 1 L G Q D A K L P C L 18 10 L Y R G D S G E Q V
14 9 C L Y R G D S G E Q 13 6 K L P C L Y R G D S 11
V7-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 15; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 4 H T D P R S Q S E E 8 9 S Q S E E P E G R S 4
V9-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 100 L L G L L K V R P L 26 5 L L A G I L L R I
T 24 95 F I Q C L L L G L L 23 4 E L L A G I L L R I 22 10 L L R I
T F N F F L 22 46 Y V A Q A G L E L L 22 18 F L F F F L P F P L 21
31 F I Y F Y F Y F F L 19 57 S S N P P A S A S L 19 97 Q C L L L G
L L K V 19 94 R F I Q C L L L G L 18 99 L L L G L L K V R P 18 105
K V R P L Q H Q G V 18 23 L P F P L V V F F I 17 64 A S L V A G T L
S V 17 22 F L P F P L V V F F 16 38 F F L E M E S H Y V 16 53 E L L
G S S N P P A 16 62 A S A S L V A G T L 16 65 S L V A G I L S V H
16 90 K K A F R F I Q C L 16 91 K A F R F I Q C L L 16 9 I L L R I
I F N F F 15 39 F L E M E S H Y V A 15 98 C L L L G L L K V R 15
103 L L K V R P L Q H Q 15 41 E M E S H Y V A Q A 14 54 L L G S S N
P P A S 14 58 S N P P A S A S L V 14 102 G L L K V R P L Q H 14 108
P L Q H Q G V N S C 14 128 F M Q A A P W E G T 14 19 L F F F L P F
P L V 13 20 F F F L P F P L V V 13 45 H Y V A Q A G L E L 13 1 M R
R E L L A G I L 12 26 P L V V F F I Y F Y 12 48 A Q A G L E L L G S
12 61 P A S A S L V A G T 12 66 L V A G T L S V H H 12 70 T L S V H
H C A C F 12 92 A F R F I Q C L L L 12
V10-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 E L G T S D V V T V 25 10 G T S D V V T V V L
18 9 L G T S D V V T V V 15 5 P A G E L G T S D V 13
V11-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 23; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 R V M V P P L P S L 22 5 L R L R V M V P P L
19 2 Q A R L R L R V M V 16 4 R L R L R V M V P P 12 1 F Q A R L R
L R V M 11 6 R L R V M V P P L P 11 9 V M V P P L P S L N 11
V12-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 25; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 9 G C S Y S I L T T V 16 2 V M S E E P E G C S
11 6 E P E G C S Y S T L 10 1 S V M S E E P E G C 8
V13-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 V L A D P Q E D S G 16 3 Q V T V D V L A D P
9 9 L A D P Q E D S G K 9 2 S Q V T V D V L A D 8
V14-HLA-A0201-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 3 S S N P P A S A S L 19 10 A S L V A G T L S V
17 8 A S A S L V A G T L 16 4 S N P P A S A S L V 14 7 P A S A S L
V A G T 12 1 L G S S N P P A S A 10
[1050]
46TABLE XXXVI Pos 1 2 3 4 5 6 7 8 9 0 score V1-HLA-
A02Q3-10mers-191P4D12B Each peptide is a portion of SEQ ID NO: 3;
each start position is specified, the length of peptide is 10 amino
acids, and the end position for each peptide is the start position
plus nine. 160 E E G Q G L T L A A 19 194 R S F K H S R S A A 19
349 S V V V V G V I A A 19 59 G E Q V G Q V A W A 18 239 I L H V S
F L A E A 18 161 E G Q G L T L A A S 17 195 S F K H S R S A A V 17
350 V V V V G V I A A L 17 5 L G A E M W G P E A 10 14 A W L L L L
L L L A 10 22 L A S F T G R C P A 10 39 V V T V V L G Q D A 10 57 D
S G E Q V G Q V A 10 63 G Q V A W A R V D A 10 67 W A R V D A G E G
A 10 71 D A G E G A Q E L A 10 84 S K Y G L H V S P A 10 108 L D G
S V L L R N A 10 111 S V L L R N A V Q A 10 125 Y E C R V S T F P A
10 130 S T F P A G S F Q A 10 149 P L P S L N P G P A 10 159 L E E
G Q G L T L A 10 164 G L T L A A S C T A 10 169 A S C T A E G S P A
10 193 S R S F K H S R S A 10 237 T H I L H V S F L A 10 257 N L W
H I G R E G A 10 339 S G K Q V D L V S A 10 348 A S V V V V G V I A
10 370 L M S R Y H R R K A 10 411 Q P E E S V G L R A 10 459 E L L
S P G S G R A 10 472 E D Q D E G I K Q A 10 485 F V Q E N G I L R A
10 6 G A E M W G P E A W 9 15 W L L L L L L L A S 9 23 A S F T G R
C P A G 9 40 V T V V L G Q D A K 9 58 S G E Q V G Q V A W 9 60 E Q
V G Q V A W A R 9 64 Q V A W A R V D A G 9 68 A R V D A G E G A Q 9
72 A G E G A Q E L A L 9 85 K Y G L H V S P A Y 9 109 D G S V L L R
N A V 9 112 V L L R N A V Q A D 9 126 E C R V S T F P A G 9 131 T F
P A G S F Q A R 9 150 L P S L N P G P A L 9 165 L T L A A S C T A E
9 170 S C T A E G S P A P 9 238 H I L H V S F L A E 9 240 L H V S F
L A E A S 9 258 L W H I G R E G A M 9 340 G K Q V D L V S A S 9 371
M S R Y H R R K A Q 9 412 P E E S V G L R A E 9 460 L L S P G S G R
A E 9 473 D Q D E G I K Q A M 9 486 V Q E N G T L R A K 9
V2-HLA-A0203-10mers-191P4D1- 2B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 160 E E G Q G L I L A A 19
V7-HLA-A0203-10mers-191P4D12B No Results Found.
V9-HLA-A0203-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 123 Y F Q G I F M Q A A 19 41 E M E S H Y V A Q
A 18 55 L G S S N P P A S A 18 124 F Q G I F M Q A A P 17 39 F L E
M E S H Y V A 10 53 E L L G S S N P P A 10 59 N P P A S A S L V A
10 68 A G T L S V H H C A 10 83 T K R K K K L K K A 10 122 G Y F Q
G I F M Q A 10 40 L E M E S H Y V A Q 9 42 M E S H Y V A Q A G 9 54
L L G S S N P P A S 9 56 G S S N P P A S A S 9 60 P P A S A S L V A
G 9 69 G T L S V H H C A C 9 84 K R K K K L K K A F 9
V10-HLA-A0203-10mers-191P4D12B No Results Found.
V11-HLA-A0203-10mers-191P4D12B No Results Found.
V12-HLA-A0203-10mers-191P4D12B No Results Found.
V13-HLA-A0203-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 1 D S Q V T V D V L A 10 2 S Q V T V D V L A D
9 3 Q V T V D V L A D P 8 V14-HLA-A0203-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 29; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 1 L G S
S N P P A S A 18 5 N P P A S A S L V A 10 2 G S S N P P A S A S 9 6
P P A S A S L V A G 9 3 S S N P P A S A S L 8 7 P A S A S L V A G T
8
[1051]
47TABLE XXXVII Pos 1 2 3 4 5 6 7 8 9 0 score
V1-HLA-A03-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
3; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 332 V L D P Q E D S G K 26 69 R V D A G E G A Q
E 25 260 H I G R E G A M L K 25 111 S V L L R N A V Q A 24 128 R V
S T F P A G S F 24 158 A L E E G Q G L T L 24 342 Q V D L V S A S V
V 23 358 A L L F C L L V V V 23 16 L L L L L L L A S F 22 140 R L R
L R V L V P P 22 235 R I T H I L H V S F 22 229 G L L Q D Q R I T H
21 376 R R K A Q Q M T Q K 21 80 A L L H S K Y G L H 20 152 S L N P
G P A L E E 20 203 A V J S E F H L V P 20 284 R L D G P L P S G V
20 345 L V S A S V V V V G 20 352 V V G V I A A L L F 20 369 V L M
S R Y H R R K 20 17 L L L L L L A S F T 19 365 V V V V V L M S R Y
19 419 R A E G H P D S L K 19 19 L L L L A S F T G R 18 33 E L E T
S D V V T V 18 117 A V Q A D E G E Y E 18 142 R L R V L V P P L P
18 144 R V L V P P L P S L 18 344 D L V S A S V V V V 18 351 V V V
G V I A A L L 18 359 L L F C L L V V V V 18 400 R L H S H H I D P R
18 450 T V R E I E T Q T E 18 15 W L L L L L L L A S 17 18 L L L L
L A S F T G 17 42 V V L G Q D A K L P 17 113 L L R N A V Q A D E 17
145 V L V P P L P S L N 17 188 K G I T S S R S F K 17 197 K H S R S
A A V T S 17 294 R V D G D I L G F P 17 304 P L I T E H S G I Y 17
364 L V V V V V L M S R 17 391 T L T R E N S I R R 17 443 R S Y S T
L I T V R 17 460 L L S P G S G R A E 17 76 A Q E L A L L H S K 16
81 L L H S K Y G L H V 16 112 V L L R N A V Q A D 16 123 G E Y E C
R V S T F 16 146 L V P P L P S L N P 16 166 T L A A S C T A E G 16
186 E V K G T T S S R S 16 223 C V V S H P G L L Q 16 224 V V S H P
G L L Q D 16 249 S V R G L E D Q N L 16 362 C L L V V V Y V L M 16
367 V V V L M S R Y H R 16 368 V V L M S R Y H R R 16 434 S V M S E
E P E G R 16 491 T L R A K P T G N G 16 20 L L L A S F I G R C 15
49 K L P C F Y R G D S 15 61 Q V G Q V A W A R V 15 77 Q E L A L L
H S K Y 15 97 R V E Q P P P P R N 15 107 P L D G S V L L R N 15 139
A R L R L R V L V P 15 164 G I T L A A S C T A 15 180 S V J W D I E
V K G 15 239 I L H V S F L A E A 15 241 H V S F L A E A S V 15 242
V S F L A E A S V R 15 251 R G L E D Q N L W H 15 267 M L K C L S E
G Q P 15 288 P L P S G V R V D G 15 299 T L G F P P L T T E 15 311
G I Y V C H V S N E 15 331 D V L D P Q E D S G 15 354 G V I A A L L
F C L 15 385 K Y E E E L T L T R 15 397 S I R R L H S H H T 15 417
G L R A E G H P D S 15 426 S L K D N S S C S V 15 493 R A K P T G N
G I Y 15 500 G I Y J N G R G H L 15 4 S L G A E M W G P E 14 21 L L
A S F T C R C P 14 38 D V V T V V L G Q D 14 41 T V V L G Q D A K L
14 64 Q V A W A R V D A G 14 89 H V S P A Y E G R V 14 179 P S V T
W D T E V K 14 209 H L V P S R S M N G 14 238 H I L H V S F L A E
14 292 G V R V D G D T L G 14 316 H V S N E F S S R D 14 350 V V V
V G V I A A L 14 363 L L V V V V V L M S 14 366 V V V V L M S R Y H
14 485 F V Q E N G T L R A 14 2 P L S L G A E M W G 13 39 V V T V V
L G Q D A 13 43 V L G Q D A K L P C 13 87 G L H V S P A Y E G 13
104 P R N P L D G S V L 13 214 R S M N G Q P L T C 13 275 Q P P P S
Y N W T R 13 357 A A L L F C L L V V 13 373 R Y H R R K A Q Q M 13
389 E L I L T R E N S I 13 396 N S I R R L H S H H 13 415 S V G L R
A E G H P 13 458 T E L L S P G S G R 13 459 E L L S P G S G R A 13
78 E L A L L H S K Y G 12 149 P L P S L N P G P A 12 230 L L Q D Q
R I T H I 12 244 F L A E A S V R G L 12 259 W H I G R E G A M L 12
270 C L S E G Q P P P S 12 285 L D G P L P S G V R 12 298 D T L G F
P P L T T 12 327 Q V T V D V L D P Q 12 349 S V V V V G V I A A 12
436 M S E E P E G R S Y 12 470 E E E D Q D E G I K 12 486 V Q E N G
T L R A K 12 V2-HLA-A03-10mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 5; each start position is specified, the length of
peptide is 10 amino acids, and the end position for each peptide is
the start position plus nine. 9 C L Y R G D S G E Q 18 6 K L P C L
Y R G D S 15 10; L Y R G D S G E Q V 11 3 Q D A K L P C L Y R 10 2
G Q D A K L P C L Y 9 5 A K L P C L Y R G D 8
V7-HLA-A03-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
15; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 8 R S Q S E E P E G R 9 2 S H H T D P R S Q S 8
4 H T D P R S Q S E E 6 V9-HLA-A03-10mers-191P4D12B Each peptide is
a portion of SEQ ID NO: 19; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 65 S L V A G T L S V H 24
102 G L L K V R P L Q H 23 9 I L L R I T F N F F 21 66 L V A G T L
S V H H 21 98 C L L L G L L K V R 21 12 R I T F N F F L F F 19 96 I
Q C L L L G L L K 19 105 K V R P L Q H Q G V 19 22 F L P F P L V V
F F 18 99; L L L G L L K V R P 18 4 E L L A G J L L R I 17 21 F F L
P F P L V V F 17 70 T L S V H H C A C F 17 82 F T K R K K K L K K
17 26 P L V V F F I Y F Y 16 28 V V F F I Y F Y F Y 16 8 G I L L R
J I F N F 15 75 H C A C F E S F T K 15 88 K L K K A F R F I Q 15 3
R E L L A G I L L R 14 10 L L R I T F N F F L 14 27 L V V F F I Y F
Y F 14 39 F L E M E S H Y V A 14 50 A G L E L L G S S N 14 51 G L E
L L G S S N P 14 53 E L L G S S N P P A 14 77 A C F E S F T K R K
14 5 L L A G I L L R I T 13 107 R P L Q H Q G V N S 13 31 F I Y F Y
F Y F F L 12 54 L L G S S N P P A S 12 62 A S A S L V A G T L 12 85
R K K K L K K A F R 12 86 K K K L K K A F R F 12 108 P L Q H Q G V
N S C 12 126 G I F M Q A A P W E 12 18 F L F F F L P F P L 11 46 Y
V A Q A G L E L L 11 72 S V H H C A C F E S 11 79 F E S F T K R K K
K 11 81 S F T K R K K K L K 11 100 L L G L L K V R P L 11 103 L L K
V R P L Q H Q 11 125 Q G I F M Q A A P W 11
V10-HLA-A03-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 E L G T S D V V T V 18 7 G E L G T S D V V T
12 3 R C P A G E L G T S 11 4 C P A G E L G T S D 9 10 G T S D V V
T V V L 9 6 A G E L G T S D V V 8 V11-HLA-A03-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 23; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 4 R L R
L R V M V P P 22 6 R L R V M V P P L P 18 8 R V M V P P L P S L 16
10 M V P P L P S L N P 16 3 A R L R L R V M V P 13 2 Q A R L R L R
V M V 12 V12-HLA-A03-10mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 25; each start position is specified, the length of
peptide is 10 amino acids, and the end position for each peptide is
the start position plus nine. 10 C S Y S T L T T V R 13 1 S V M S E
E P E G C 12 3 M S E E P E G C S Y 12 6 E P E G C S Y S T L 9 4 S E
E P E G C S Y S 7 8 E G C S Y S J L T T 7
V13-HLA-A03-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 9 L A D P Q E D S G K 16 3 Q V T V D V L A D P
15 7 D V L A D P Q E D S 14 8 V L A D P Q E D S G 14 5 T V D V L A
D P Q E 13 V14-HLA-A03-10mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 29; each start position is specified, the length of
peptide is 10 amino acids, and the end position for each peptide is
the start position plus nine. 8 A S A S L V A G T L 12 4 S N P P A
S A S L V 10 10 A S L V A G T L S V 10 3 S S N P P A S A S L 9 5 N
P P A S A S L V A 9 2 G S S N P P A S A S 8 1 L G S S N P P A S A 6
6 P P A S A S L V A G 6 9 S A S L V A G T L S 6 7 P A S A S L V A G
T 5
[1052]
48TABLE XXXVIII Pos 1 2 3 4 5 6 7 8 9 0 score
V1-HLA-A26-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
3; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 38 D V V T V V L G Q D 28 35 E T S D V V T V V
L 27 350 V V V V G V I A A L 27 354 G V I A A L L F C L 26 365 V V
V V V L M S R Y 25 41 T V V L G Q D A K L 24 13 E A W L L L L L L L
23 144 R V L V P P L P S L 23 455 E T Q T E L L S P G 23 351 V V V
G V I A A L L 22 392 L T R E N S I R R L 22 476 E G I K Q A M N H F
22 186 E V K G T T S S R S 21 236 I T H I L H V S F L 21 349 S V V
V V G V I A A 21 128 R V S T F P A G S F 20 331 D V L D P Q E D S G
20 439 E P E G R S Y S T L 20 99 E Q P P P P R N P L 19 249 S V R G
L E D Q N L 19 352 V V G V I A A L L F 19 364 L V V V V V L M S R
19 8 E M W G P E A W L L 18 298 D T L G F P P L T T 18 25 F T G R C
P A G E L 17 184 D T E V K G T T S S 17 223 C V V S H P G L L Q 17
344 D L V S A S V V V V 17 123 G E Y E C R V S T F 16 221 L T C V V
S H P G L 16 224 V V S H P G L L Q D 16 296 D G D T L G F P P L 16
472 E D Q D E G I K Q A 16 10 W G P E A W L L L L 15 33 E L E T S D
V V T V 15 60 E Q V G Q V A W A R 15 64 Q V A W A R V D A G 15 116
N A V Q A D E G E Y 15 130 S T F P A G S F Q A 15 161 E G Q G L T L
A A S 15 291 S G V R V D G D T L 15 294 R V D G D T L G F P 15 327
Q V T V D V L D P Q 15 395 E N S I R R L H S H 15 421 E G H P D S L
K D N 15 453 E I E T Q T E L L S 15 204 V T S E F H L V P S 14 222
T C V V S H P G L L 14 235 R I T H I L H V S F 14 244 F L A E A S V
R G L 14 247 E A S V R G L E D Q 14 259 W H I G R E G A M L 14 293
V R V D G D T L G F 14 308 E H S G I Y V C H V 14 328 V T V D V L D
P Q E 14 337 E D S G K Q V D L V 14 345 L V S A S V V V V G 14 366
V V V V L M S R Y H 14 367 V V V L M S R Y H R 14 414 E S V G L R A
E G H 14 429 D N S S C S V M S E 14 436 M S E E P E G R S Y 14 448
L T T V R E I E T Q 14 449 T T V R E I E T Q T 14 450 T V R E I E T
Q T E 14 452 R E I E T Q T E L L 14 483 N H F V Q E N G T L 14 11 G
P E A W L L L L L 13 12 P E A W L L L L L L 13 16 L L L L L L L A S
F 13 40 V T V V L G Q D A K 13 44 L G Q D A K L P C F 13 158 A L E
E G Q G L T L 13 180 S V T W D T E V K G 13 181 V T W D T E V K G T
13 203 A V T S E F H L V P 13 233 D Q R I T H I L H V 13 255 D Q N
L W H I G R E 13 305 L T T E H S G I Y V 13 306 T T E H S G I Y V C
13 438 E E P E G R S Y S T 13 441 E G R S Y S T L T T 13 471 E E D
Q D E G I K Q 13 485 F V Q E N G T L R A 13 500 G I Y I N G R G H L
13 V2-HLA-A26-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 1 L G Q D A K L P C L 13 4 D A K L P C L Y R G
12 2 G Q D A K L P C L Y 10 V7-HLA-A26-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 4 H T D
P R S Q S E E 10 6 D P R S Q S E E P E 9 9 S Q S E E P E G R S 4
V9-HLA-A26-10mers-191P4D12B Each peptide is a portion of SEQ ID NO:
19; each start position is specified, the length of peptide is 10
amino acids, and the end position for each peptide is the start
position plus nine. 13 I T F N F F L F F F 24 28 V V F F I Y F Y F
Y 24 80 E S F T K R K K K L 23 27 L V V F F I Y F Y F 22 46 Y V A Q
A G L E L L 22 26 P L V V F F I Y F Y 18 43 E S H Y V A Q A G L 18
94 R F I Q C L L L G L 17 95; F I Q C L L L G L L 17 41 E M E S H Y
V A Q A 16 4 E L L A G I L L R I 15 37 Y F F L E M E S H Y 15 12 R
I T F N F F L F F 14 45 H Y V A Q A G L E L 14 16 N F F L F F F L P
F 13 21 F F L P F P L V V F 13 8 G I L L R I T F N F 12 11 L R I T
F N F F L F 12 18 F L F F F L P F P L 12 22 F L P F P L V V F F 12
29 V F F I Y F Y F Y F 12 30 F F I Y F Y F Y F F 12 31 F I Y F Y F
Y F F L 12 90 K K A F R F I Q C L 12 91 K A F R F I Q C L L 12 100
L L G L L K V R P L 12 120 E R G Y F Q G I F M 12 1 M R R E L L A G
I L 11 57 S S N P P A S A S L 11 62 A S A S L V A G T L 11 72 S V H
H C A C F E S 11 105 K V R P L Q H Q G V 11 113 G V N S C D C E R G
11 V10-HLA-A26-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 10 G T S D V V T V V L 17 8 E L G T S D V V T V
15 V11-HLA-A26-10mers-191P4D12- B Each peptide is a portion of SEQ
ID NO: 23; each start position is specified, the length of peptide
is 10 amino acids, and the end position for each peptide is the
start position plus nine. 8 R V M V P P L P S L 23 5 L R L R V M V
P P L 12 10 M V P P L P S L N P 12 V12-HLA-A26-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 25; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 6 E P E
G C S Y S T L 20 3 M S E E P E G C S Y 14 5 E E P E G C S Y S T 13
8 E G C S Y S T L T T 13 1 S V M S E E P E G C 12
V13-HLA-A26-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 7 D V L A D P Q E D S 18 3 Q V T V D V L A D P
15 4 V T V D V L A D P Q 13 5 T V D V L A D P Q E 12 2 S Q V T V D
V L A D 11 1 D S Q V T V D V L A 8 V14-HLA-A26-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 29; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 3 S S N
P P A S A S L 11 8 A S A S L V A G T L 11 6 P P A S A S L V A G
6
[1053]
49TABLE XXXIX Pos 1 2 3 4 5 6 7 8 9 0 score
V1-HLA-B0702-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 132 F P A G S F Q A R L 24 150 L P S L N P G P
A L 24 11 G P E A W L L L L L 23 439 E P E G R S Y S T L 23 156 G P
A L E E G Q G L 21 178 A P S V T W D T E V 21 276 P P P S Y N W T R
L 21 176 S P A P S V T W D T 19 103 P P R N P L D G S V 18 407 D P
R S Q P E E S V 18 411 Q P E E S V G L R A 18 35 E T S D V V T V V
L 17 72 A G E G A Q E L A L 17 134 A G S F Q A R L R L 17 227 H P G
L L Q D Q R I 17 303 P P L T T E H S G I 16 334 D P Q E D S G K Q V
16 289 L P S G V R V D G D 15 324 R D S Q V T V D V L 15 7 A E M W
G P E A W L 14 9 M W G P E A W L L L 14 29 C P A G E L E T S D 14
91 S P A Y E G R V E Q 14 99 E Q P P P P R N P L 14 158 A L E E G Q
G L T L 14 249 S V R G L E D Q N L 14 296 D G D T L G F P P L 14
361 F C L L V V V V V L 14 409 R S Q P E E S V G L 14 8 E M W G P E
A W L L 13 12 P E A W L L L L L L 13 13 E A W L L L L L L L 13 70 V
D A G E G A Q E L 13 73 G E G A Q E L A L L 13 101 P P P P R N P L
D G 13 105 R N P L D G S V L L 13 106 N P L D G S V L L R 13 141 L
R L R V L V P P L 13 212 P S R S M N G Q P L 13 236 I T H I L H V S
F L 13 259 W H I G R E G A M L 13 277 P P S Y N W T R L D 13 287 G
P L P S G V R V D 13 336 Q E D S G K Q V D L 13 351 V V V G V I A A
L L 13 355 V I A A L L F C L L 13 495 K P T G N G I Y I N 13 10 W G
P E A W L L L L 12 100 Q P P P P R N P L D 12 104 P R N P L D G S V
L 12 137 F Q A R L R L R V L 12 144 R V L V P P L P S L 12 148 P P
L P S L N P G P 12 154 N P G P A L E E G Q 12 160 E E G Q G L T L A
A 12 211 V P S R S M N G Q P 12 231 L Q D Q R I T H I L 12 244 F L
A E A S V R G L 12 262 G R E G A M L K C L 12 308 E H S G I Y V C H
V 12 337 E D S G K Q V D L V 12 350 V V V V G V I A A L 12 383 T Q
K Y E E E L T L 12 392 L T R E N S I R R L 12 441 E G R S Y S T L T
T 12 452 R E I E T Q T E L L 12 25 F T G R C P A G E L 11 41 T V V
L G Q D A K L 11 56 G D S G E Q V G Q V 11 138 Q A R L R L R V L V
11 147 V P P L P S L N P G 11 201 S A A V T S E F H L 11 219 Q P L
T C V V S H P 11 221 L T C V V S H P G L 11 275 Q P P P S Y N W T R
11 280 Y N W T R L D G P L 11 354 G V I A A L L F C L 11 357 A A L
L F C L L V V 11 358 A L L F C L L V V V 11 418 L R A E G H P D S L
11 423 H P D S L K D N S S 11 451 V R E I E T Q T E L 11 462 S P G
S G R A E E E 11 500 G I Y I N G R G H L 11
V2-HLA-B0702-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 5; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 1 L G Q D A K L P C L 11 7 L P C L Y R G D S G
10 10 L Y R G D S G E Q V 10 V7-HLA-B0702-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 6 D P R
S Q S E E P E 13 V9-HLA-B0702-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 19; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 59 N P P A S A S L V A 20
23 L P F P L V V F F I 19 25 F P L V V F F I Y F 17 92 A F R F I Q
C L L L 16 60 P P A S A S L V A G 14 10 L L R I T F N F F L 13 45 H
Y V A Q A G L E L 13 62 A S A S L V A G T L 13 94 R F I Q C L L L G
L 13 100 L L G L L K V R P L 13 107 R P L Q H Q G V N S 13 1 M R R
E L L A G I L 12 14 T F N F F L F F F L 12 43 E S H Y V A Q A G L
12 57 S S N P P A S A S L 12 90 K K A F R F I Q C L 12 2 R R E L L
A G I L L 11 12 R I T F N F F L F F 11 18 F L F F F L P F P L 11 31
F I Y F Y F Y F F L 11 46 Y V A Q A G L E L L 11 53 E L L G S S N P
P A 11 61 P A S A S L V A G T 11 64 A S L V A G T L S V 11 80 E S F
T K R K K K L 11 91 K A F R F I Q C L L 11 4 E L L A G I L L R I 10
16 N F F L F F F L P F 10 21 F F L P F P L V V F 10 22 F L P F P L
V V F F 10 87 K K L K K A F R F I 10 95 F I Q C L L L G L L 10 105
K V R P L Q H Q G V 10 119 C E R G Y F Q G I F 10 5 L L A G I L L R
I T 9 9 I L L R I T F N F F 9 20 F F F L P F P L V V 9 33 Y F Y F Y
F F L E M 9 41 E M E S H Y V A Q A 9 55 L G S S N P P A S A 9 70 T
L S V H H C A C F 9 83 T K R K K K L K K A 9 84 K R K K K L K K A F
9 120 E R G Y F Q G I F M 9 123 Y F Q G I F M Q A A 9
V10-HLA-B0702-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 10 G T S D V V T V V L 16 4 C P A G E L G T S D
14 7 G E L G T S D V V T 11 8 E L G T S D V V T V 11 2 G R C P A G
E L G T 9 6 A G E L G T S D V V 9 9 L G T S D V V T V V 9 5 P A G E
L G T S D V 8 V11-HLA-B0702-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 23; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 5 L R L R V M V P P L 13 8
R V M V P P L P S L 13 2 Q A R L R L R V M V 11 1 F Q A R L R L R V
M 8 4 R L R L R V M V P P 6 V12-HLA-B0702-10mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 25; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 6 E P E
G C S Y S T L 23 V13-HLA-B0702-10mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 27; each start position is specified, the
length of peptide is 10 amino acids, and the end position for each
peptide is the start position plus nine. 1 D S Q V T V D V L A 8 2
S Q V T V D V L A D 4 V14-HLA-B0702-10mers-191P4D12B Each peptide
is a portion of SEQ ID NO: 29; each start position is specified,
the length of peptide is 10 amino acids, and the end position for
each peptide is the start position plus nine. 5 N P P A S A S L V A
20 6 P P A S A S L V A G 14 8 A S A S L V A G T L 13 3 S S N P P A
S A S L 12 7 P A S A S L V A G T 11 10 A S L V A G T L S V 11 1 L G
S S N P P A S A 9
[1054]
50TABLE XL Pos 1234567890 score V1-HLA-B08-10mers-191P4D12B No
Results Found.
[1055]
51TABLE XXIV Pos 123456789 score V7-HLA-A0203-9mers-191P4D12B No
Results Found. V9-HLA-A0203-9mers-191P4D12B No Results Found.
V10-HLA-A0203-9mers-191P4D12B No Results Found.
V11-HLA-A0203-9mers-191P4D12B No Results Found.
V12-HLA-A0203-9mers-191P4D12B No Results Found.
V13-HLA-A0203-9mers-191P4D12B No Results Found.
V14-HLA-A0203-9mers-191P4D12B No Results Found.
[1056]
52TABLE XLI Pos 1234567890 score V1-HLA-B1510-10mers-191P4D12B No
Results Found. V2-HLA-B1510-10mers-191P4D12B No Results Found.
V7-HLA-B1510-10mers-191P4D12B No Results Found.
V9-HLA-B1510-10mers-191P4D12B No Results Found.
V10-HLA-B1510-10mers-191P4D12B No Results Found.
V11-HLA-B1510-10mers-191P4D12B No Results Found.
V12-HLA-B1510-10mers-191P4D12B No Results Found.
V13-HLA-B1510-10mers-191P4D12B No Results Found.
V14-HLA-B1510-10mers-191P4D12B No Results Found.
[1057]
53TABLE XLII Pos 1234567890 score V1-HLA-B2705-10mers-191P4D12B No
Results Found. V2-HLA-B2705-10mers-191P4D12B No Results Found.
V7-HLA-B2705-10mers-191P4D12B No Results Found.
V9-HLA-B2705-10mers-191P4D12B No Results Found.
V10-HLA-B2705-10mers-191P4D12B No Results Found.
V11-HLA-B2705-10mers-191P4D12B No Results Found.
V12-HLA-B2705-10mers-191P4D12B No Results Found.
V13-HLA-B2705-10mers-191P4D12B No Results Found.
V14-HLA-B2705-10mers-191P4D12B No Results Found.
[1058]
54TABLE XLIII Pos 1234567890 score V1-HLA-B2709-10mers-191P4D12B No
Results Found. V2-HLA-B2709-10mers-191P4D12B No Results Found.
V7-HLA-B2709-10mers-191P4D12B No Results Found.
V9-HLA-B2709-10mers-191P4D12B No Results Found.
V10-HLA-B2709-10mers-191P4D12B No Results Found.
V11-HLA-B2709-10mers-191P4D12B No Results Found.
V12-HLA-B2709-10mers-191P4D12B No Results Found.
V13-HLA-B2709-10mers-191P4D12B No Results Found.
V14-HLA-B2709-10mers-191P4D12B No Results Found.
[1059]
55TABLE XLIV Pos 1 2 3 4 5 6 7 8 9 0 score
V1-HLA-B4402-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 3; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 452 R E I E T Q T E L L 25 7 A E M W G P E A W
L 24 12 P E A W L L L L L L 23 73 G E G A Q E L A L L 22 77 Q E L A
L L H S K Y 22 123 G E Y E C R V S T F 22 336 Q E D S G K Q V D L
22 469 E E E E D Q D E G I 20 99 E Q P P P P R N P L 18 174 E G S P
A P S V T W 18 35 E T S D V V T V V L 17 72 A G E G A Q E L A L 17
13 E A W L L L L L L L 16 134 A G S F Q A R L R L 16 160 E E G Q G
L T L A A 16 476 E G I K Q A M N H F 16 8 E M W G P E A W L L 15 9
M W G P E A W L L L 15 98 V E Q P P P P R N P 15 158 A L E E G Q G
L T L 15 173 A E G S P A P S V T 15 273 E G Q P P P S Y N W 15 350
V V V V G V I A A L 15 361 F C L L V V V V V L 15 387 E E E L T L T
R E N 15 388 E E L T L T R E N S 15 420 A E G H P D S L K D 15 437
S E E P E G R S Y S 15 471 E E D Q D E G I K Q 15 10 W G P E A W L
L L L 14 58 S G E Q V G Q V A W 14 85 K Y G L H V S P A Y 14 104 P
R N P L D G S V L 14 105 R N P L D G S V L L 14 137 F Q A R L R L R
V L 14 150 L P S L N P G P A L 14 206 S E F H L V P S R S 14 246 A
E A S V R G L E D 14 259 W H I G R E G A M L 14 262 G R E G A M L K
C L 14 319 N E F S S R D S Q V 14 354 G V I A A L L F C L 14 392 L
T R E N S I R R L 14 409 R S Q P E E S V G L 14 412 P E E S V G L R
A E 14 413 E E S V G L R A E G 14 439 E P E G R S Y S T L 14 483 N
H F V Q E N G T L 14 494 A K P T G N G I Y I 14 6 G A E M W G P E A
W 13 11 G P E A W L L L L L 13 16 L L L L L L L A S F 13 32 G E L E
T S D V V T 13 128 R V S T F P A G S F 13 141 L R L R V L V P P L
13 159 L E E G Q G L T L A 13 199 S R S A A V T S E F 13 231 L Q D
Q R I T H I L 13 250 V R G L E D Q N L W 13 291 S G V R V D G D T L
13 293 V R V D G D T L G F 13 296 D G D T L G F P P L 13 324 R D S
Q V T V D V L 13 351 V V V G V I A A L L 13 352 V V G V I A A L L F
13 438 E E P E G R S Y S T 13 468 A E E E E D Q D E G 13 470 E E E
D Q D E G I K 13 487 Q E N G T L R A K P 13 493 R A K P T G N G I Y
13 1 M P L S L G A E M W 12 25 F T G R C P A G E L 12 34 L E T S D
V V T V V 12 41 T V V L G Q D A K L 12 44 L G Q D A K L P C F 12 45
G Q D A K L P C F Y 12 70 V D A G E G A Q E L 12 79 L A L L H S K Y
G L 12 121 D E G E Y E C R V S 12 125 Y E C R V S T F P A 12 144 R
V L V P P L P S L 12 187 V K G T T S S R S F 12 222 T C V V S H P G
L L 12 230 L L Q D Q R I T H I 12 244 F L A E A S V R G L 12 249 S
V R G L E D Q N L 12 253 L E D Q N L W H I G 12 271 L S E G Q P P P
S Y 12 272 S E G Q P P P S Y N 12 347 S A S V V V V G V I 12 355 V
I A A L L F C L L 12 377 R K A Q Q M T Q K Y 12 383 T Q K Y E E E L
T L 12 389 E L T L T R E N S I 12 394 R E N S I R R L H S 12 440 P
E G R S Y S T L T 12 454 I E T Q T E L L S P 12 458 T E L L S P G S
G R 12 V2-HLA-B4402-10mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 5; each start position is specified, the length of
peptide is 10 amino acids, and the end position for each peptide is
the start position plus nine. 2 G Q D A K L P C L Y 13 1 L G Q D A
K L P C L 12 5 A K L P C L Y R G D 8 V7-HLA-B4402-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 2 S H H
T D P R S Q S 4 4 H T D P R S Q S E E 4 1 H S H H T D P R S Q 2 5 T
D P R S Q S E E P 2 9 S Q S E E P E G R S 2 3 H H T D P R S Q S E 1
7 P R S Q S E E P E G 1 8 R S Q S E E P E G R 1
V9-HLA-B4402-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 19; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 119 C E R G Y F Q G I F 21 80 E S F T K R K K K
L 18 3 R E L L A G I L L R 17 21 F F L P F P L V V F 17 11 L R I T
F N F F L F 16 16 N F F L F F F L P F 16 62 A S A S L V A G T L 15
79 F E S F T K R K K K 15 84 K R K K K L K K A F 15 91 K A F R F I
Q C L L 15 92 A F R F I Q C L L L 15 94 R F I Q C L L L G L 15 9 I
L L R I T F N F F 14 13 I T F N F F L F F F 14 23 L P F P L V V F F
I 14 30 F F I Y F Y F Y F F 14 40 L E M E S H Y V A Q 14 42 M E S H
Y V A Q A G 14 57 S S N P P A S A S L 14 90 K K A F R F I Q C L 14
125 Q G I F M Q A A P W 14 2 R R E L L A G I L L 13 4 E L L A G I L
L R I 13 6 L A G I L L R I T F 13 8 G I L L R I T F N F 13 18 F L F
F F L P F P L 13 22 F L P F P L V V F F 13 24 P F P L V V F F I Y
13 25 F P L V V F F I Y F 13 26 P L V V F F I Y F Y 13 28 V V F F I
Y F Y F Y 13 37 Y F F L E M E S H Y 13 52 L E L L G S S N P P 13 86
K K K L K K A F R F 13 100 L L G L L K V R P L 13 115 N S C D C E R
G Y F 13 12 R I T F N F F L F F 12 29 V F F I Y F Y F Y F 12 43 E S
H Y V A Q A G L 12 46 Y V A Q A G L E L L 12 87 K K L K K A F R F I
12 95 F I Q C L L L G L L 12 114 V N S C D C E R G Y 12 1 M R R E L
L A G I L 11 14 T F N F F L F F F L 11 45 H Y V A Q A G L E L 11 70
T L S V H H C A C F 11 73 V H H C A C F E S F 11 7 A G I L L R I T
F N 10 10 L L R I T F N F F L 10 27 L V V F F I Y F Y F 10 31 F I Y
F Y F Y F F L 10 118 D C E R G Y F Q G I 10
V10-HLA-B4402-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 21; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 10 G T S D V V T V V L 15 7 G E L G T S D V V T
14 V11-HLA-B4402-10mers-191P4D- 12B Each peptide is a portion of
SEQ ID NO: 23; each start position is specified, the length of
peptide is 10 amino acids, and the end position for each peptide is
the start position plus nine. 5 L R L R V M V P P L 13 8 R V M V P
P L P S L 12 3 A R L R L R V M V P 7 V12-HLA-B4402-10mers-191P4D12B
Each peptide is a portion of SEQ ID NO: 25; each start position is
specified, the length of peptide is 10 amino acids, and the end
position for each peptide is the start position plus nine. 4 S E E
P E G C S Y S 14 6 E P E G C S Y S T L 14 5 E E P E G C S Y S T 13
7 P E G C S Y S T L T 11 3 M S E E P E G C S Y 10
V13-HLA-B4402-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 27; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 2 S Q V T V D V L A D 6 10 A D P Q E D S G K Q
5 9 L A D P Q E D S G K 4 1 D S Q V T V D V L A 2 4 V T V D V L A D
P Q 2 5 T V D V L A D P Q E 2 6 V D V L A D P Q E D 2
V14-HLA-B4402-10mers-191P4D12B Each peptide is a portion of SEQ ID
NO: 29; each start position is specified, the length of peptide is
10 amino acids, and the end position for each peptide is the start
position plus nine. 8 A S A S L V A G T L 15 3 S S N P P A S A S L
14 4 S N P P A S A S L V 7
[1060]
56TABLE XLV Pos 1234567890 score V1-HLA-B5101-10mers-191P4D12B No
Results Found. V2-HLA-B5101-10mers-191P4D12B No Results Found.
V7-HLA-B5101-10mers-191P4D12B No Results Found.
V9-HLA-B5101-10mers-191P4D12B No Results Found.
V10-HLA-B5101-10mers-191P4D12B No Results Found.
V11-HLA-B5101-10mers-191P4D12B No Results Found.
V12-HLA-B5101-10mers-191P4D12B No Results Found.
V13-HLA-B5101-10mers-191P4D12B No Results Found.
V14-HLA-B5101-10mers-191P4D12B No Results Found.
[1061]
57TABLE XLVI Pos 1 2 3 4 5 6 7 8 9 0 1 2 3 4 5 score V1-HLA-DRB1
-0101-15mers-191P4D12B Each peptide is a portion of SEQ ID NO: 3;
each start position is specified, the length of peptide is 15 amino
acids, and the end position for each peptide is the start position
plus fourteen. 279 S Y N W T R L D G P L P S G V 35 140 R L R L R V
L V P P L P S L N 32 205 T S E F H L V P S R S M N G Q 32 299 T L G
F P P L T T E H S G I Y 32 37 S D V V T V V L G Q D A K L P 31 40 V
T V V L G Q D A K L P C F Y 31 340 G K Q V D L V S A S V V V V G 31
349 S V V V V G V I A A L L F C L 31 144 R V L V P P L P S L N P G
P A 30 147 V P P L P S L N P G P A L E E 30 350 V V V V G V I A A L
L F C L L 30 51 P C F Y R G D S G E Q V G Q V 28 12 P E A W L L L L
L L L A S F T 27 247 E A S V R G L E D Q N L W H I 27 358 A L L F C
L L V V V V V L M S 27 371 M S R Y H R R K A Q Q M T Q K 26 6 G A E
M W G P E A W L L L L L 25 13 E A W L L L L L L L A S F T G 25 14 A
W L L L L L L L A S F T G R 25 15 W L L L L L L L A S F T G R C 25
19 L L L L A S F T G R C P A G E 25 102 P P P R N P L D G S V L L R
N 25 109 D G S V L L R N A V Q A D E G 25 122 E G E Y E C R V S T F
P A G S 25 193 S R S F K H S R S A A V T S E 25 239 I L H V S F L A
E A S V R G L 25 255 D Q N L W H I G R E G A M L K 25 265 G A M L K
C L S E G Q P P P S 25 310 S G I Y V C H V S N E F S S R 25 454 I E
T Q T E L L S P G S G R A 25 64 Q V A W A R V D A G E G A Q E 24 76
A Q E L A L L H S K Y G L H V 24 79 L A L L H S K Y G L H V S P A
24 126 E C R V S T F P A G S F Q A R 24 156 G P A L E E G Q G L T L
A A S 24 162 G Q G L T L A A S C T A E G S 24 181 V T W D T E V K G
T T S S R S 24 210 L V P S R S M N G Q P L T C V 24 213 S R S M N G
Q P L T C V V S H 24 282 W T R L D G P L P S G V R V D 24 347 S A S
V V V V G V I A A L L F 24 353 V G V I A A L L F C L L V V V 24 357
A A L L F C L L V V V V V L M 24 364 L V V V V V L M S R Y H R R K
24 395 E N S I R R L H S H H T D P R 24 442 G R S Y S T L T T V R E
I E T 24 16 L L L L L L L A S F T G R C P 23 28 R C P A G E L E T S
D V V T V 23 184 D T E V K G T T S S R S F K H 23 228 P G L L Q D Q
R I T H I L H V 23 233 D Q R I T H I L H V S F L A E 23 289 L P S G
V R V D G D T L G F P 23 339 S G K Q V D L V S A S V V V V 23 346 V
S A S V V V V G V I A A L L 23 361 F C L L V V V V V L M S R Y H 23
424 P D S L K D N S S C S V M S E 23 448 L T T V R E I E T Q T E L
L S 23 457 Q T E L L S P G S G R A E E E 23 483 N H F V Q E N G T L
R A K P T 23 3 L S L G A E M W G P E A W L L 22 55 R G D S G E Q V
G Q V A W A R 22 59 G E Q V G Q V A W A R V D A G 22 141 L R L R V
L V P P L P S L N P 22 204 V T S E F H L V P S R S M N G 22 250 V R
G L E D Q N L W H I G R E 22 268 L K C L S E G Q P P P S Y N W 22
311 G I Y V C H V S N E F S S R D 22 327 Q V T V D V L D P Q E D S
G K 22 360 L F C L L V V V V V L M S R Y 22 451 V R E I E T Q T E L
L S P G S 22 218 G Q P L T C V V S H P G L L Q 21 256 Q N L W H I G
R E G A M L K C 21 277 P P S Y N W T R L D G P L P S 21 33 E L E T
S D V V T V V L G Q D 20 65 V A W A R V D A G E G A Q E L 20 123 G
E Y E C R V S T F P A G S F 20 154 N P G P A L E E G Q G L T L A 20
321 F S S R D S Q V T V D V L D P 20 429 D N S S C S V M S E E P E
G R 20 482 M N H F V Q E N G T L R A K P 20 490 G T L R A K P T G N
G I Y I N 20 22 L A S F T G R C P A G E L E T 19 39 V V T V V L G Q
D A K L P C F 19 138 Q A R L R L R V L V P P L P S 19 234 Q R I T H
I L H V S F L A E A 19 242 V S F L A E A S V R G L E D Q 19 412 P E
E S V G L R A E G H P D S 19 415 S V G L R A E G H P D S L K D 19 7
A E M W G P E A W L L L L L L 18 91 S P A Y E G R V E Q P P P P R
18 134 A G S F Q A R L R L R V L V P 18 165 L T L A A S C T A E G S
P A P 18 264 E G A M L K C L S E G Q P P P 18 266 A M L K C L S E G
Q P P P S Y 18 280 Y N W T R L D G P L P S G V R 18 368 V V L M S R
Y H R R K A Q Q M 18 387 E E E L T L T R E N S I R R L 18 11 G P E
A W L L L L L L L A S F 17 67 W A R V D A G E G A Q E L A L 17 68 A
R V D A G E G A Q E L A L L 17 83 H S K Y G L H V S P A Y E G R 17
115 R N A V Q A D E G E Y E C R V 17 125 Y E C R V S T F P A G S F
Q A 17 135 G S F Q A R L R L R V L V P P 17 148 P P L P S L N P G P
A L E E G 17 150; L P S L N P G P A L E E G Q G 17 167 L A A S C T
A E G S P A P S V 17 201 S A A V T S E F H L V P S R S 17 221 L T C
V V S H P G L L Q D Q R 17 225 V S H P G L L Q D Q R I T H I 17 238
H I L H V S F L A E A S V R G 17 257 N L W H I G R E G A M L K C L
17 258 L W H I G R E G A M L K C L S 17 284 R L D G P L P S G V R V
D G D 17 291 S G V R V D G D T L G F P P L 17 294 R V D G D T L G F
P P L T T E 17 303 P P L T T E H S G I Y V C H V 17 330 V D V L D P
Q E D S G K Q V D 17 332 V L D P Q E D S G K Q V D L V 17 342 Q V D
L V S A S V V V V G V I 17 348 A S V V V V G V I A A L L F C 17 354
G V I A A L L F C L L V V V V 17 356 I A A L L F C L L V V V V V L
17 379 A Q Q M T Q K Y E E E L T L T 17 407 D P R S Q P E E S V G L
R A E 17 413 E E S V G L R A E G H P D S L 17 432 S C S V M S E E P
E G R S Y S 17 458 T E L L S P G S G R A E E E E 17 475 D E G I K Q
A M N H F V Q E N 17 486 V Q E N G T L R A K P T G N G 17
V2-HLA-DRB1-0101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 5; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 2 V T V V L G Q D A K L P C L Y 31 13
P C L Y R G D S G E Q V G Q V 28 9 D A K L P C L Y R G D S G E Q 24
1 V V T V V L G Q D A K L P C L 19
V7-HLA-DRB1-0101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 15; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 2 I R R L H S H H T D P R S Q S 14 8
H H T D P R S Q S E E P E G R 14 13 R S Q S E E P E G R S Y S T L
10 1 S I R R L H S H H T D P R S Q 9 11 D P R S Q S E E P E G R S Y
S 9 14 S Q S E E P E G R S Y S T L T 9 3 R R L H S H H T D P R S Q
S E 8 5 L H S H H T D P R S Q S E E P 8 9 H T D P R S Q S E E P E G
R S 8 12 P R S Q S E E P E G R S Y S T 8 4 R L H S H H T D P R S Q
S E E 7 6 H S H H T D P R S Q S E E P E 7
V9-HLA-DRB1-0101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 19; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 43 E S H Y V A Q A G L E L L G S 33
49 Q A G L E L L G S S N P P A S 32 36 F Y F F L E M E S H Y V A Q
A 31 103 L L K V R P L Q H Q G V N S C 28 17 F F L F F F L P F P L
V V F F 27 90 K K A F R F I Q C L L L G L L 27 98 C L L L G L L K V
R P L Q H Q 26 18 F L F F F L P F P L V V F F I 25 60 P P A S A S L
V A G T L S V H 24 61 P A S A S L V A G T L S V H H 24 93 F R F I Q
C L L L G L L K V R 24 97 Q C L L L G L L K V R P L Q H 24 121 R G
Y F Q G I F M Q A A P W E 24 6 L A G I L L R I T F N F F L F 23 16
N F F L F F F L P F P L V V F 23 7 A G I L L R I T F N F F L F F 22
52 L E L L G S S N P P A S A S L 22 100 L L G L L K V R P L Q H Q G
V 22 8 G I L L R I T F N F F L F F F 21 27 L V V F F I Y F Y F Y F
F L E 21 12 R I T F N F F L F F F L P F P 20 34 F Y F Y F F L E M E
S H Y V A 20 92 A F R F I Q C L L L G L L K V 20 4 E L L A G I L L
R I T F N F F 19 14 T F N F F L F F F L P F P L V 19 15 F N F F L F
F F L P F P L V V 19 31 F I Y F Y F Y F F L E M E S H 19 33 Y F Y F
Y F F L E M E S H Y V 19 46 Y V A Q A G L E L L G S S N P 19 95 F I
Q C L L L G L L K V R P L 19 10 L L R I T F N F F L F F F L P 18 19
L F F F L P F P L V V F F I Y 18 25 F P L V V F F I Y F Y F Y F F
18 28 V V F F I Y F Y F Y F F L E M 18 84 K R K K K L K K A F R F I
Q C 18 120 E R G Y F Q G I F M Q A A P W 18 13 I T F N F F L F F F
L P F P L 17 20 F F F L P F P L V V F F I Y F 17 22 F L P F P L V V
F F I Y F Y F 17 29 V F F I Y F Y F Y F F L E M E 17 37 Y F F L E M
E S H Y V A Q A G 17 44 S H Y V A Q A G L E L L G S S 17 94 R F I Q
C L L L G L L K V R P 17 2 R R E L L A G I L L R I T F N 16 21 F F
L P F P L V V F F I Y F Y 16 39 F L E M E S H Y V A Q A G L E 16 41
E M E S H Y V A Q A G L E L L 16 48 A Q A G L E L L G S S N P P A
16 51 G L E L L G S S N P P A S A S 16 54 L L G S S N P P A S A S L
V A 16 56 G S S N P P A S A S L V A G T 16 68 A G T L S V H H C A C
F E S F 16 70 T L S V H H C A C F E S F T K 16 105 K V R P L Q H Q
G V N S C D C 16 118 D C E R G Y F Q G I F M Q A A 16
V10-HLA-DRB1-0101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 21; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 8 R C P A G E L G T S D V V T V 23 13
E L G T S D V V T V V L G Q D 20 2 L A S F T G R C P A G E L G T 19
3 A S F T G R C P A G E L G T S 16 11 A G E L G T S D V V T V V L G
16 9 C P A G E L G T S D V V T V V 15
V11-HLA-DRB1-0101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 23; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 9 R L R L R V M V P P L P S L N 30 13
R V M V P P L P S L N P G P A 30 10 L R L R V M V P P L P S L N P
22 7 Q A R L R L R V M V P P L P S 19 3 A G S F Q A R L R L R V M V
P 18 4 G S F Q A R L R L R V M V P P 17 6 F Q A R L R L R V M V P P
L P 16 11 R L R V M V P P L P S L N P G 16 1 F P A G S F Q A R L R
L R V M 15 12 L R V M V P P L P S L N P G P 15 8 A R L R L R V M V
P P L P S L 14 V12-HLA-DRB1-0101-15mers-191P4- D12B Each peptide is
a portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 15 amino acids, and the end position for each
peptide is the start position plus fourteen. 14 G C S Y S T L T T V
R E I E T 24 1 D N S S C S V M S E E P E G C 20 4 S C S V M S E E P
E G C S Y S 17 5 C S V M S E E P E G C S Y S T 16 15 C S Y S T L T
T V R E I E T Q 11 V13-HLA-DRB1-0101-15mers-191P4D12B Each peptide
is a portion of SEQ ID NO: 27; each start position is specified,
the length of peptide is 15 amino acids, and the end position for
each peptide is the start position plus fourteen. 2 F S S R D S Q V
T V D V L A D 20 6 D S Q V T V D V L A D P Q E D 17 14 L A D P Q E
D S G K Q V D L V 17 8 Q V T V D V L A D P Q E D S G 16 10 T V D V
L A D P Q E D S G K Q 16 7 S Q V T V D V L A D P Q E D S 15 3 S S R
D S Q V T V D V L A D P 14 12 D V L A D P Q E D S G K Q V D 9
V14-HLA-DRB1-0101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 29; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 11 P P A S A S L V A G T L S V H 24
12 P A S A S L V A G T L S V H H 24 3 L E L L G S S N P P A S A S L
22 2 G L E L L G S S N P P A S A S 16 5 L L G S S N P P A S A S L V
A 16 7 G S S N P P A S A S L V A G T 16 1 A G L E L L G S S N P P A
S A 15 6 L G S S N P P A S A S L V A G 15 13 A S A S L V A G T L S
V H H C 15 4 E L L G S S N P P A S A S L V 14 8 S S N P P A S A S L
V A G T L 14 15 A S L V A G T L S V H H C A C 14
[1062]
58TABLE XLVII Pos 1 2 3 4 5 6 7 8 9 0 1 2 3 4 5 score
V1-HLA-DRB1-0301-5mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 3; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 178 A P S V T W D T E V K G T T S 29
227 H P G L L Q D Q R I T H I L H 28 41 T V V L G Q D A K L P C F Y
R 27 379 A Q Q M T Q K Y E E E L T L T 25 14 A W L L L L L L L A S
F T G R 23 290 P S G V R V D G D T L G F P P 23 39 V V T V V L G Q
D A K L P C F 22 103 P P R N P L D G S V L L R N A 22 247 E A S V R
G L E D Q N L W H I 22 115 R N A V Q A D E G E Y E C R V 21 142 R L
R V L V P P L P S L N P G 21 233 D Q R I T H I L H V S F L A E 21
325 D S Q V T V D V L D P Q E D S 21 348 A S V V V V G V I A A L L
F C 21 349 S V V V V G V I A A L L F C L 21 6 G A E M W G P E A W L
L L L L 20 156 G P A L E E G Q G L T L A A S 20 242 V S F L A E A S
V R G L E D Q 20 249 S V R G L E D Q N L W H I G R 20 292 G V R V D
G D T L G F P P L T 20 350 V V V V G V I A A L L F C L L 20 352 V V
G V I A A L L F C L L V V 20 353 V G V I A A L L F C L L V V V 20
363 L L V V V V V L M S R Y H R R 20 126 E C R V S T F P A G S F Q
A R 19 302 F P P L T T E H S G I Y V C H 19 328 V T V D V L D P Q E
D S G K Q 19 365 V V V V V L M S R Y H R R K A 19 387 E E E L T L T
R E N S I R R L 19 77 Q E L A L L H S K Y G L H V S 18 111 S V L L
R N A V Q A D E G E Y 18 265 G A M L K C L S E G Q P P P S 18 286 D
G P L P S G V R V D G D T L 18 319 N E F S S R D S Q V T V D V L 18
329 T V D V L D P Q E D S G K Q V 18 433 C S V M S E E P E G R S Y
S T 18 451 V R E I E T Q T E L L S P G S 18 87 G L H V S P A Y E G
R V E Q P 17 97 R V E Q P P P P R N P L D G S 17 239 I L H V S F L
A E A S V R G L 17 255 D Q N L W H I G R E G A M L K 17 311 G I Y V
C H V S N E F S S R D 17 334 D P Q E D S G K Q V D L V S A 17 368 V
V L M S R Y H R R K A Q Q M 17 381 Q M T Q K Y E E E L T L T R E 17
401 L H S H H T D P R S Q P E E S 17 413 E E S V G L R A E G H P D
S L 17 445 Y S T L T T V R E I E T Q T E 17 475 D E G I K Q A M N H
F V Q E N 17 479 K Q A M N H F V Q E N G T L R 17 491 T L R A K P T
G N G I Y I N G 17 5 L G A E M W G P E A W L L L L 16 13 E A W L L
L L L L L A S F T G 16 47 D A K L P C F Y R G D S G E Q 16 70 V D A
G E G A Q E L A L L H S 16 134 A G S F Q A R L R L R V L V P 16 114
L R N A V Q A D E G E Y E C R 15 130 S T F P A G S F Q A R L R L R
15 132 F P A G S F Q A R L R L R V L 15 199 S R S A A V T S E F H L
V P S 15 221 L T C V V S H P G L L Q D Q R 15 236 I T H I L H V S F
L A E A S V 15 481 A M N H F V Q E N G T L R A K 15 15 W L L L L L
L L A S F T G R C 14 17 L L L L L L A S F T G R C P A 14 78 E L A L
L H S K Y G L H V S P 14 109 D G S V L L R N A V Q A D E G 14 110 G
S V L L R N A V Q A D E G E 14 143 L R V L V P P L P S L N P G P 14
144 R V L V P P L P S L N P G P A 14 280 Y N W T R L D G P L P S G
V R 14 342 Q V D L V S A S V V V V G V I 14 356 I A A L L F C L L V
V V V V L 14 360 L F C L L V V V V V L M S R Y 14 448 L T T V R E I
E T Q T E L L S 14 449 T T V R E I E T Q T E L L S P 14 457 Q T E L
L S P G S G R A E E E 14 V2-HLA-DRB1-0301-15mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 5; each start position is
specified, the length of peptide is 15 amino acids, and the end
position for each peptide is the start position plus fourteen. 3 T
V V L G Q D A K L P C L Y R 27 1 V V T V V L G Q D A K L P C L 22 9
D A K L P C L Y R G D S G E Q 16 2 V T V V L G Q D A K L P C L Y 13
V7-HLA-DRB1-0301-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 15; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 5 L H S H H T D P R S Q S E E P 17 2
I R R L H S H H T D P R S Q S 11 13 R S Q S E E P E G R S Y S T L
10 9 H T D P R S Q S E E P E G R S 9 7 S H H T D P R S Q S E E P E
G 8 12 P R S Q S E E P E G R S Y S T 8 14 S Q S E E P E G R S Y S T
L T 8 V9-HLA-DRB1-0301-15mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 19; each start position is specified, the length of
peptide is 15 amino acids, and the end position for each peptide is
the start position plus fourteen. 8 G I L L R I T F N F F L F F F
25 112 Q G V N S C D C E R G Y F Q G 24 35 Y F Y F F L E M E S H Y
V A Q 23 6 L A G I L L R I T F N F F L F 22 7 A G I L L R I T F N F
F L F F 21 19 L F F F L P F P L V V F F I Y 21 10 L L R I T F N F F
L F F F L P 20 20 F F F L P F P L V V F F I Y F 20 44 S H Y V A Q A
G L E L L G S S 20 93 F R F I Q C L L L G L L K V R 20 97 Q C L L L
G L L K V R P L Q H 20 98 C L L L G L L K V R P L Q H Q 20 16 N F F
L F F F L P F P L V V F 19 24 P F P L V V F F I Y F Y F Y F 19 25 F
P L V V F F I Y F Y F Y F F 19 51 G L E L L G S S N P P A S A S 19
68 A G T L S V H H C A C F E S F 19 90 K K A F R F I Q C L L L G L
L 19 92 A F R F I Q C L L L G L L K V 19 14 T F N F F L F F F L P F
P L V 18 26 P L V V F F I Y F Y F Y F F L 18 29 V F F I Y F Y F Y F
F L E M E 18 12 R I T F N F F L F F F L P F P 17 22 F L P F P L V V
F F I Y F Y F 17 28 V V F F I Y F Y F Y F F L E M 17 79 F E S F T K
R K K K L K K A F 17 82 F T K R K K K L K K A F R F I 17 86 K K K L
K K A F R F I Q C L L 17 27 L V V F F I Y F Y F Y F F L E 16 76 C A
C F E S F T K R K K K L K 16 4 E L L A G I L L R I T F N F F 15 33
Y F Y F Y F F L E M E S H Y V 15 41 E M E S H Y V A Q A G L E L L
15 78 C F E S F T K R K K K L K K A 15 89 L K K A F R F I Q C L L L
G L 15 113 G V N S C D C E R G Y F Q G I 15 117 C D C E R G Y F Q G
I F M Q A 15 96 I Q C L L L G L L K V R P L Q 14 2 R R E L L A G I
L L R I T F N 13 49 Q A G L E L L G S S N P P A S 13 100 L L G L L
K V R P L Q H Q G V 13 101 L G L L K V R P L Q H Q G V N 13 103 L L
K V R P L Q H Q G V N S C 13 36 F Y F F L E M E S H Y V A Q A 12 37
Y F F L E M E S H Y V A Q A G 12 39 F L E M E S H Y V A Q A G L E
12 52 L E L L G S S N P P A S A S L 12 64 A S L V A G T L S V H H C
A C 12 106 V R P L Q H Q G V N S C D C E 12
V10-HLA-DRB1-0301-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 21; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 12 G E L G T S D V V T V V L G Q 12
11 A G E L G T S D V V T V V L G 11 2 L A S F T G R C P A G E L G T
10 3 A S F T G R C P A G E L G T S 9 5 F T G R C P A G E L G T S D
V 9 13 E L G T S D V V T V V L G Q D 9
V11-HLA-DRB1-0301-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 23; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 11 R L R V M V P P L P S L N P G 19 3
A G S F Q A R L R L R V M V P 16 1 F P A G S F Q A R L R L R V M 15
12 L R V M V P P L P S L N P G P 14 13 R V M V P P L P S L N P G P
A 14 7 Q A R L R L R V M V P P L P S 13 9 R L R L R V M V P P L P S
L N 12 5 S F Q A R L R L R V M V P P L 10 8 A R L R L R V M V P P L
P S L 10 15 M V P P L P S L N P G P A L E 10
V12-HLA-DRB1-0301-5mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 25; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 5 C S V M S E E P E G C S Y S T 18 4
S C S V M S E E P E G C S Y S 12 6 S V M S E E P E G C S Y S T L 10
3 S S C S V M S E E P E G C S Y 9 9 S E E P E G C S Y S T L T T V 9
V13-HLA-DRB1-0301-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 27; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 10 T V D V L A D P Q E D S G K Q 29 6
D S Q V T V D V L A D P Q E D 22 11 V D V L A D P Q E D S G K Q V
16 V14-HLA-DRB1-0301-15mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 29; each start position is specified, the length of
peptide is 15 amino acids, and the end position for each peptide is
the start position plus fourteen. 2 G L E L L G S S N P P A S A S
19 3 L E L L G S S N P P A S A S L 12 15 A S L V A G T L S V H H C
A C 12 14 S A S L V A G T L S V H H C A 11 6 L G S S N P P A S A S
L V A G 10 11 P P A S A S L V A G T L S V H 9
[1063]
59TABLE XLVIII Pos 1 2 3 4 5 6 7 8 9 0 1 2 3 4 5 score
V1-HLA-DRB1-0401-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 3; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 205 T S E F H L V P S R S M N G Q 28
299 T L G F P P L T T E H S G I Y 28 47 D A K L P C F Y R G D S G E
Q 26 162 G Q G L T L A A S C T A E G S 26 255 D Q N L W H I G R E G
A M L K 26 311 G I Y V C H V S N E F S S R D 26 395 E N S I R R L H
S H H T D P R 26 415 S V G L R A E G H P D S L K D 26 475 D E G I K
Q A M N H F V Q E N 26 7 A E M W G P E A W L L L L L L 22 12 P E A
W L L L L L L L A S F T 22 50 L P C F Y R G D S G E Q V G Q 22 51 P
C F Y R G D S G E Q V G Q V 22 180 S V T W D T E V K G T T S S R 22
193 S R S F K H S R S A A V T S E 22 241 H V S F L A E A S V R G L
E D 22 358 A L L F C L L V V V V V L M S 22 383 T Q K Y E E E L T L
T R E N S 22 442 G R S Y S T L T T V R E I E T 22 13 E A W L L L L
L L L A S F T G 20 15 W L L L L L L L A S F T G R C 20 16 L L L L L
L L A S F T G R C P 20 37 S D V V T V V L G Q D A K L P 20 59 G E Q
V G Q V A W A R V D A G 20 76 A Q E L A L L H S K Y G L H V 20 87 G
L H V S P A Y E G R V E Q P 20 111 S V L L R N A V Q A D E G E Y 20
144 R V L V P P L P S L N P G P A 20 147 V P P L P S L N P G P A L
E E 20 184 D T E V K G T T S S R S F K H 20 201 S A A V T S E F H L
V P S R S 20 218 G Q P L T C V V S H P G L L Q 20 227 H P G L L Q D
Q R I T H I L H 20 233 D Q R I T H I L H V S F L A E 20 239 I L H V
S F L A E A S V R G L 20 242 V S F L A E A S V R G L E D Q 20 247 E
A S V R G L E D Q N L W H I 20 258 L W H I G R E G A M L K C L S 20
264 E G A M L K C L S E G Q P P P 20 302 F P P L T T E H S G I Y V
C H 20 314 V C H V S N E F S S R D S Q V 20 325 D S Q V T V D V L D
P Q E D S 20 340 G K Q V D L V S A S V V V V G 20 342 Q V D L V S A
S V V V V G V I 20 347 S A S V V V V G V I A A L L F 20 349 S V V V
V G V I A A L L F C L 20 352 V V G V I A A L L F C L L V V 20 353 V
G V I A A L L F C L L V V V 20 357 A A L L F C L L V V V V V L M 20
360 L F C L L V V V V V L M S R Y 20 361 F C L L V V V V V L M S R
Y H 20 364 L V V V V V L M S R Y H R R K 20 368 V V L M S R Y H R R
K A Q Q M 20 389 E L T L T R E N S I R R L H S 20 424 P D S L K D N
S S C S V M S E 20 433 C S V M S E E P E G R S Y S T 20 445 Y S T L
T T V R E I E T Q T E 20 448 L T T V R E I E T Q T E L L S 20 457 Q
T E L L S P G S G R A E E E 20 479 K Q A M N H F V Q E N G T L R 20
483 N H F V Q E N G T L R A K P T 20 28 R C P A G E L E T S D V V T
V 18 29 C P A G E L E T S D V V T V V 18 33 E L E T S D V V T V V L
G Q D 18 38 D V V T V V L G Q D A K L P C 18 89 H V S P A Y E G R V
E Q P P P 18 103 P P R N P L D G S V L L R N A 18 107 P L D G S V L
L R N A V Q A D 18 108 L D G S V L L R N A V Q A D E 18 120 A D E G
E Y E C R V S T F P A 18 123 G E Y E C R V S T F P A G S F 18 128 R
V S T F P A G S F Q A R L R 18 155 P G P A L E E G Q G L T L A A 18
190 T T S S R S F K H S R S A A V 18 219 Q P L T C V V S H P G L L
Q D 18 308 E H S G I Y V C H V S N E F S 18 315 C H V S N E F S S R
D S Q V T 18 319 N E F S S R D S Q V T V D V L 18 328 V T V D V L D
P Q E D S G K Q 18 331 D V L D P Q E D S G K Q V D L 18 339 S G K Q
V D L V S A S V V V V 18 373 R Y H R R K A Q Q M T Q K Y E 18 386 Y
E E E L T L T R E N S I R R 18 392 L T R E N S I R R L H S H H T 18
407 D P R S Q P E E S V G L R A E 18 423 H P D S L K D N S S C S V
M S 18 435 V M S E E P E G R S Y S T L T 18 449 T T V R E I E T Q T
E L L S P 18 454 I E T Q T E L L S P G S G R A 18 472 E D Q D E G I
K Q A M N H F V 18 134 A G S F Q A R L R L R V L V P 17 318 S N E F
S S R D S Q V T V D V 17 64 Q V A W A R V D A G E G A Q E 16 83 H S
K Y G L H V S P A Y E G R 16 256 Q N L W H I G R E G A M L K C 16
279 S Y N W T R L D G P L P S G V 16 310 S G I Y V C H V S N E F S
S R 16 482 M N H F V Q E N G T L R A K P 16 367 V V V L M S R Y H R
R K A Q Q 15 2 P L S L G A E M W G P E A W L 14 6 G A E M W G P E A
W L L L L L 14 14 A W L L L L L L L A S F T G R 14 17 L L L L L L A
S F T G R C P A 14 18 L L L L L A S F T G R C P A G 14 19 L L L L A
S F T G R C P A G E 14 31 A G E L E T S D V V T V V L G 14 36 T S D
V V T V V L G Q D A K L 14 39 V V T V V L G Q D A K L P C F 14 41 T
V V L G Q D A K L P C F Y R 14 62 V G Q V A W A R V D A G E G A 14
95 E G R V E Q P P P P R N P L D 14 105 R N P L D G S V L L R N A V
Q 14 115 R N A V Q A D E G E Y E C R V 14 126 E C R V S T F P A G S
F Q A R 14 140 R L R L R V L V P P L P S L N 14 142 R L R V L V P P
L P S L N P G 14 143 L R V L V P P L P S L N P G P 14 156 G P A L E
E G Q G L T L A A S 14 164 G L T L A A S C T A E G S P A 14 178 A P
S V T W D T E V K G T T S 14 207 E F H L V P S R S M N G Q P L 14
213 S R S M N G Q P L T C V V S H 14 221 L T C V V S H P G L L Q D
Q R 14 228 P G L L Q D Q R I T H I L H V 14 236 I T H I L H V S F L
A E A S V 14 237 T H I L H V S F L A E A S V R 14 250 V R G L E D Q
N L W H I G R E 14 265 G A M L K C L S E G Q P P P S 14 268 L K C L
S E G Q P P P S Y N W 14 282 W T R L D G P L P S G V R V D 14 286 D
G P L P S G V R V D G D T L 14 290 P S G V R V D G D T L G F P P 14
292 G V R V D G D T L G F P P L T 14 327 Q V T V D V L D P Q E D S
G K 14 330 V D V L D P Q E D S G K Q V D 14 348 A S V V V V G V I A
A L L F C 14 350 V V V V G V I A A L L F C L L 14 356 I A A L L F C
L L V V V V V L 14 362 C L L V V V V V L M S R Y H R 14 363 L L V V
V V V L M S R Y H R R 14 365 V V V V V L M S R Y H R R K A 14 387 E
E E L T L T R E N S I R R L 14 398 I R R L H S H H T D P R S Q P 14
432 S C S V M S E E P E G R S Y S 14 451 V R E I E T Q T E L L S P
G S 14 V2-HLA-DRB1-0401-15mers-191P4- D12B Each peptide is a
portion of SEQ ID NO: 5; each start position is specified, the
length of peptide is 15 amino acids, and the end position for each
peptide is the start position plus fourteen. 9 D A K L P C L Y R G
D S G E Q 26 13 P C L Y R G D S G E Q V G Q V 22 12 L P C L Y R G D
S G E Q V G Q 20 1 V V T V V L G Q D A K L P C L 14 3 T V V L G Q D
A K L P C L Y R 14 4 V V L G Q D A K L P C L Y R G 12 15 L Y R G D
S G E Q V G Q V A W 12 V7-HLA-DRB1-0401-15mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 15; each start position is
specified, the length of peptide is 15 amino acids, and the end
position for each peptide is the start position plus fourteen. 5 L
H S H H T D P R S Q S E E P 18 14 S Q S E E P E G R S Y S T L T 18
2 I R R L H S H H T D P R S Q S 14 12 P R S Q S E E P E G R S Y S T
12 V9-HLA-DRB1-0401-15mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 19; each start position is specified, the length of
peptide is 15 amino acids, and the end position for each peptide is
the start position plus fourteen. 37 Y F F L E M E S H Y V A Q A G
26 86 K K K L K K A F R F I Q C L L 26 103 L L K V R P L Q H Q G V
N S C 26 12 R I T F N F F L F F F L P F P 22 17 F F L F F F L P F P
L V V F F 22 33 Y F Y F Y F F L E M E S H Y V 22 36 F Y F F L E M E
S H Y V A Q A 22 76 C A C F E S F T K R K K K L K 22 90 K K A F R F
I Q C L L L G L L 22 121 R G Y F Q G I F M Q A A P W E 22 3 R E L L
A G I L L R I T F N F 20 8 G I L L R I T F N F F L F F F 20 16 N F
F L F F F L P F P L V V F 20 44 S H Y V A Q A G L E L L G S S 20 49
Q A G L E L L G S S N P P A S 20 51 G L E L L G S S N P P A S A S
20 93 F R F I Q C L L L G L L K V R 20 98 C L L L G L L K V R P L Q
H Q 20 41 E M E S H Y V A Q A G L E L L 18 62 A S A S L V A G T L S
V H H C 18 73 V H H C A C F E S F T K R K K 18 89 L K K A F R F I Q
C L L L G L 18 14 T F N F F L F F F L P F P L V 16 15 F N F F L F F
F L P F P L V V 16 18 F L F F F L P F P L V V F F I 16 19 L F F F L
P F P L V V F F I Y 16 22 F L P F P L V V F F I Y F Y F 16 28 V V F
F I Y F Y F Y F F L E M 16 30 F F I Y F Y F Y F F L E M E S 16 31 F
I Y F Y F Y F F L E M E S H 16 32 I Y F Y F Y F F L E M E S H Y 16
34 F Y F Y F F L E M E S H Y V A 16 35 Y F Y F F L E M E S H Y V A
Q 16 43 E S H Y V A Q A G L E L L G S 16 92 A F R F I Q C L L L G L
L K V 16 120 E R G Y F Q G F F M Q A A P W 16 2 R R E L L A G I L L
R I T F N 14 7 A G I L L R I T F N F F L F F 14 24 P F P L V V F F
I Y F Y F Y F 14 25 F P L V V F F I Y F Y F Y F F 14 26 P L V V F F
I Y F Y F Y F F L 14 29 V F F I Y F Y F Y F F L E M E 14 39 F L E M
E S H Y V A Q A G L E 14 52 L E L L G S S N P P A S A S L 14 64 A S
L V A G T L S V H H C A C 14 70 T L S V H H C A C F E S F T K 14 97
Q C L L L G L L K V R P L Q H 14 100 L L G L L K V R P L Q H Q G V
14 4 E L L A G I L L R I T F N F F 12 5 L L A G I L L R I T F N F F
L 12 21 F F L P F P L V V F F I Y F Y 12 46 Y V A Q A G L E L L G S
S N P 12 47 V A Q A G L E L L G S S N P P 12 48 A Q A G L E L L G S
S N P P A 12 55 L G S S N P P A S A S L V A G 12 56 G S S N P P A S
A S L V A G T 12 57 S S N P P A S A S L V A G T L 12 60 P P A S A S
L V A G T L S V H 12 61 P A S A S L V A G T L S V H H 12 66 L V A G
T L S V H H C A C F E 12 67 V A G T L S V H H C A C F E S 12 75 H C
A C F E S F T K R K K K L 12 77 A C F E S F T K R K K K L K K 12 94
R F I Q C L L L G L L K V R P 12 95 F I Q C L L L G L L K V R P L
12 104 L K V R P L Q H Q G V N S C D 12 108 P L Q H Q G V N S C D C
E R G 12 114 V N S C D C E R G Y F Q G I F 12 118 D C E R G Y F Q G
I F M Q A A 12 122 G Y F Q G I F M Q A A P W E G 12
V10-HLA-DRB1-0401-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 21; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 8 R C P A G E L G T S D V V T V 18 13
E L G T S D V V T V V L G Q D 18 11 A G E L G T S D V V T V V L G
14 5 F T G R C P A G E L G T S D V 12 9 C P A G E L G T S D V V T V
V 12 12 G E L G T S D V V T V V L G Q 12
V11-HLA-DRB1-0401-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 23; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 13 R V M V P P L P S L N P G P A 20 3
A G S F Q A R L R L R V M V P 17 11 R L R V M V P P L P S L N P G
14 12 L R V M V P P L P S L N P G P 14 1 F P A G S F Q A R L R L R
V M 12 4 G S F Q A R L R L R V M V P P 12 8 A R L R L R V M V P P L
P S L 12 10 L R L R V M V P P L P S L N P 12
V12-HLA-DRB1-0401-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 25; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 14 G C S Y S T L T T V R E I E T 22 5
C S V M S E E P E G C S Y S T 20 4 S C S V M S E E P E G C S Y S 14
1 D N S S C S V M S E E P E G C 12 7 V M S E E P E G C S Y S T L T
12 8 M S E E P E G C S Y S T L T T 12 10 E E P E G C S Y S T L T T
V R 12 11 E P E G C S Y S T L T T V R E 12
V13-HLA-DRB1-0401-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 27; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 10 T V D V L A D P Q E D S G K Q 26
13 V L A D P Q E D S G K Q V D L 18 6 D S Q V T V D V L A D P Q E D
14 8 Q V T V D V L A D P Q E D S G 14 2 F S S R D S Q V T V D V L A
D 12 3 S S R D S Q V T V D V L A D P 12 7 S Q V T V D V L A D P Q E
D S 12 14 L A D P Q E D S G K Q V D L V 12
V14-HLA-DRB1-0401-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 29; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 2 G L E L L G S S N P P A S A S 20 13
A S A S L V A G T L S V H H C 18 3 L E L L G S S N P P A S A S L 14
15 A S L V A G T L S V H H C A C 14 6 L G S S N P P A S A S L V A G
12 7 G S S N P P A S A S L V A G T 12 8 S S N P P A S A S L V A G T
L 12 11 P P A S A S L V A G T L S V H 12 12 P A S A S L V A G T L S
V H H 12
[1064]
60TABLE XLIX Pos 1 2 3 4 5 6 7 8 9 0 1 2 3 4 5 score
V1-HLA-DRB1-1101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 3; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 255 D Q N L W H I G R E G A M L K 26
279 S Y N W T R L D G P L P S G V 25 12 P E A W L L L L L L L A S F
T 23 201 S A A V T S E F H L V P S R S 23 64 Q V A W A R V D A G E
G A Q E 22 140 R L R L R V L V P P L P S L N 22 218 G Q P L T C V V
S H P G L L Q 22 233 D Q R I T H I L H V S F L A E 22 286 D G P L P
S G V R V D G D T L 22 299 T L G F P P L T T E H S G I Y 22 368 V V
L M S R Y H R R K A Q Q M 22 37 S D V V T V V L G Q D A K L P 21
261 I G R E G A M L K C L S E G Q 21 361 F C L L V V V V V L M S R
Y H 21 47 D A K L P C F Y R G D S G E Q 20 134 A G S F Q A R L R L
R V L V P 20 180 S V T W D T E V K G T T S S R 20 365 V V V V V L M
S R Y H R R K A 20 386 Y E E E L T L T R E N S I R R 20 392 L T R E
N S I R R L H S H H T 20 415 S V G L R A E G H P D S L K D 20 347 S
A S V V V V G V I A A L L F 19 358 A L L F C L L V V V V V L M S 19
13 E A W L L L L L L L A S F T G 18 16 L L L L L L L A S F T G R C
P 18 76 A Q E L A L L H S K Y G L H V 18 91 S P A Y E G R V E Q P P
P P R 18 122 E G E Y E C R V S T F P A G S 18 144 R V L V P P L P S
L N P G P A 18 147 V P P L P S L N P G P A L E E 18 241 H V S F L A
E A S V R G L E D 18 265 G A M L K C L S E G Q P P P S 18 311 G I Y
V C H V S N E F S S R D 18 442 G R S Y S T L T T V R E I E T 18 204
V T S E F H L V P S R S M N G 17 205 T S E F H L V P S R S M N G Q
17 367 V V V L M S R Y H R R K A Q Q 17 190 T T S S R S F K H S R S
A A V 16 277 P P S Y N W T R L D G P L P S 16 346 V S A S V V V V G
V I A A L L 16 360 L F C L L V V V V V L M S R Y 16 487 Q E N G T L
R A K P T G N G I 16 75 G A Q E L A L L H S K Y G L H 15 107 P L D
G S V L L R N A V Q A D 15 178 A P S V T W D T E V K G T T S 15 192
S S R S F K H S R S A A V T S 15 219 Q P L T C V V S H P G L L Q D
15 230 L L Q D Q R I T H I L H V S F 15 343 V D L V S A S V V V V G
V I A 15 362 C L L V V V V V L M S R Y H R 15 363 L L V V V V V L M
S R Y H R R 15 411 Q P E E S V G L R A E G H P D 15 476 E G I K Q A
M N H F V Q E N G 15 485 F V Q E N G T L R A K P T G N 15 20 L L L
A S F T G R C P A G E L 14 34 L E T S D V V T V V L G Q D A 14 36 T
S D V V T V V L G Q D A K L 14 41 T V V L G Q D A K L P C F Y R 14
59 G E Q V G Q V A W A R V D A G 14 61 Q V G Q V A W A R V D A G E
G 14 81 L L H S K Y G L H V S P A Y E 14 138 Q A R L R L R V L V P
P L P S 14 162 G Q G L T L A A S C T A E G S 14 181 V T W D T E V K
G T T S S R S 14 184 D T E V K G T T S S R S F K H 14 227 H P G L L
Q D Q R I T H I L H 14 252 G L E D Q N L W H I G R E G A 14 276 P P
P S Y N W T R L D G P L P 14 290 P S G V R V D G D T L G F P P 14
308 E H S G I Y V C H V S N E F S 14 350 V V V V G V I A A L L F C
L L 14 357 A A L L F C L L V V V V V L M 14 364 L V V V V V L M S R
Y H R R K 14 397 S I R R L H S H H T D P R S Q 14 401 L H S H H T D
P R S Q P E E S 14 420 A E G H P D S L K D N S S C S 14 433 C S V M
S E E P E G R S Y S T 14 435 V M S E E P E G R S Y S T L T 14 445 Y
S T L T T V R E I E T Q T E 14 454 I E T Q T E L L S P G S G R A 14
457 Q T E L L S P G S G R A E E E 14 479 K Q A M N H F V Q E N G T
L R 14 483 N H F V Q E N G T L R A K P T 14 19 L L L L A S F T G R
C P A G E 13 40 V T V V L G Q D A K L P C F Y 13 85 K Y G L H V S P
A Y E G R V E 13 106 N P L D G S V L L R N A V Q A 13 137 F Q A R L
R L R V L V P P L P 13 215 S M N G Q P L T C V V S H P G 13 237 T H
I L H V S F L A E A S V R 13 327 Q V T V D V L D P Q E D S G K 13
340 G K Q V D L V S A S V V V V G 13 349 S V V V V G V I A A L L F
C L 13 353 V G V I A A L L F C L L V V V 13 451 V R E I E T Q T E L
L S P G S 13 3 L S L G A E M W G P E A W L L 12 14 A W L L L L L L
L A S F T G R 12 15 W L L L L L L L A S F T G R C 12 22 L A S F T G
R C P A G E L E T 12 62 V G Q V A W A R V D A G E G A 12 73 G E G A
Q E L A L L H S K Y G 12 82 L H S K Y G L H V S P A Y E G 12 83 H S
K Y G L H V S P A Y E G R 12 92 P A Y E G R V E Q P P P P R N 12
109 D G S V L L R N A V Q A D E G 12 112 V L L R N A V Q A D E G E
Y E 12 123 G E Y E C R V S T F P A G S F 12 141 L R L R V L V P P L
P S L N P 12 153 L N P G P A L E E G Q G L T L 12 159 L E E G Q G L
T L A A S C T A 12 164 G L T L A A S C T A E G S P A 12 207 E F H L
V P S R S M N G Q P L 12 236 I T H I L H V S F L A E A S V 12 239 I
L H V S F L A E A S V R G L 12 247 E A S V R G L E D Q N L W H I 12
268 L K C L S E G Q P P P S Y N W 12 292 G V R V D G D T L G F P P
L T 12 310 S G I Y V C H V S N E F S S R 12 324 R D S Q V T V D V L
D P Q E D 12 329 T V D V L D P Q E D S G K Q V 12 337 E D S G K Q V
D L V S A S V V 12 395 E N S I R R L H S H H T D P R 12 413 E E S V
G L R A E G H P D S L 12 421 E G H P D S L K D N S S C S V 12 429 D
N S S C S V M S E E P E G R 12 448 L T T V R E I E T Q T E L L S 12
455 E T Q T E L L S P G S G R A E 12 489 N G T L R A K P T G N G I
Y I 12 V2-HLA-DRB1-1101-15mers-191P4D12B Each peptide is a portion
of SEQ ID NO: 5; each start position is specified, the length of
peptide is 15 amino acids, and the end position for each peptide is
the start position plus fourteen. 9 D A K L P C L Y R G D S G E Q
26 3 T V V L G Q D A K L P C L Y R 14 2 V T V V L G Q D A K L P C L
Y 13 V7-HLA-DRB1-1101-15mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 15; each start position is specified, the length of
peptide is 15 amino acids, and the end position for each peptide is
the start position plus fourteen. 1 S I R R L H S H H T D P R S Q
14 5 L H S H H T D P R S Q S E E P 14 14 S Q S E E P E G R S Y S T
L T 14 3 R R L H S H H T D P R S Q S E 8 12 P R S Q S E E P E G R S
Y S T 8 2 I R R L H S H H T D P R S Q S 6 8 H H T D P R S Q S E E P
E G R 6 10 T D P R S Q S E E P E G R S Y 6
V9-HLA-DRB1-1101-15mers-191P4- D12B Each peptide is a portion of
SEQ ID NO: 19; each start position is specified, the length of
peptide is 15 amino acids, and the end position for each peptide is
the start position plus fourteen. 97 Q C L L L G L L K V R P L Q H
28 121 R G Y F Q G I F M Q A A P W E 22 37 Y F F L E M E S H Y V A
Q A G 21 79 F E S F T K R K K K L K K A F 21 76 C A C F E S F T K R
K K K L K 20 103 L L K V R P L Q H Q G V N S C 20 22 F L P F P L V
V F F I Y F Y F 19 17 F F L F F F L P F P L V V F F 18 49 Q A G L E
L L G S S N P P A S 18 66 L V A G T L S V H H C A C F E 18 34 F Y F
Y F F L E M E S H Y V A 17 90 K K A F R F I Q C L L L G L L 17 120
E R G Y F Q G I F M Q A A P W 17 15 F N F F L F F F L P F P L V V
16 33 Y F Y F Y F F L E M E S H Y V 16 36 F Y F F L E M E S H Y V A
Q A 16 86 K K K L K K A F R F I Q C L L 15 3 R E L L A G I L L R I
T F N F 14 4 E L L A G I L L R I T F N F F 14 13 I T F N F F L F F
F L P F P L 14 V9-HLA-DRB1-1101-15mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 19; each start position is specified, the
length of peptide is 15 amino acids, and the end position for each
peptide is the start position plus fourteen. 67 V A G T L S V H H C
A C F E S 14 83 T K R K K K L K K A F R F I Q 14 111 H Q G V N S C
D C E R G Y F Q 14 26 P L V V F F I Y F Y F Y F F L 13 61 P A S A S
L V A G T L S V H H 13 93 F R F I Q C L L L G L L K V R 13 98 C L L
L G L L K V R P L Q H Q 13 V10-HLA-DRB1-1101-15mers-191P4D12B Each
peptide is a portion of SEQ ID NO: 21; each start position is
specified, the length of peptide is 15 amino acids, and the end
position for each peptide is the start position plus fourteen. 14 L
G T S D V V T V V L G Q D A 14 2 L A S F T G R C P A G E L G T 12
13 E L G T S D V V T V V L G Q D 9 1 L L A S F T G R C P A G E L G
7 4 S F T G R C P A G E L G T S D 7 6 T G R C P A G E L G T S D V V
6 8 R C P A G E L G T S D V V T V 6 11 A G E L G T S D V V T V V L
G 6 V11-HLA-DRB1-1101-15mers-191P4D12B Each peptide is a portion of
SEQ ID NO: 23; each start position is specified, the length of
peptide is 15 amino acids, and the end position for each peptide is
the start position plus fourteen. 9 R L R L R V M V P P L P S L N
22 3 A G S F Q A R L R L R V M V P 20 13 R V M V P P L P S L N P G
P A 18 7 Q A R L R L R V M V P P L P S 14 6 F P A R L R L R V M V P
P L P 13 10 L R L R V M V P P L P S L N P 12 1 F P A G S F Q A R L
R L R V M 10 V12-HLA-DRB1-1101-15mers-191P4D12B Each peptide is a
portion of SEQ ID NO: 25; each start position is specified, the
length of peptide is 15 amino acids, and the end position for each
peptide is the start position plus fourteen. 14 G C S Y S T L T T V
R E I E T 18 1 D N S S C S V M S E E P E G C 12 5 C S V M S E E P E
G C S Y S T 12 2 N S S C S V M S E E P E G C S 7
V13-HLA-DRB1-1101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 27; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 6 D S Q V T V D V L A D P Q E D 17 8
Q V T V D V L A D P Q E D S G 13 10 T V D V L A D P Q E D S G K Q
12 11 V D V L A D P Q E D S G K Q V 12 4 S R D S Q V T V D V L A D
P Q 10 15 A D P Q E D S G K Q V D L V S 9
V14-HLA-DRB1-1101-15mers-191P4D12B Each peptide is a portion of SEQ
ID NO: 3; each start position is specified, the length of peptide
is 15 amino acids, and the end position for each peptide is the
start position plus fourteen. 12 P A S A S L V A G T L S V H H 13 2
G L E L L G S S N P P A S A S 12 31 L E L L G S S N P P A S A S L
12 11 P P A S A S L V A G T L S V H 8 8 S S N P P A S A S L V A G T
L 7 14 S A S L V A G T L S V H H C A 7 1 A G L E L L G S S N P P A
S A 6 4 E L L G S S N P P A S A S L V 6 5 L L G S S N P P A S A S L
V A 6 9 S N P P A S A S L V A G T L S 6 15 A S L V A G T L S V H H
C A C 6
[1065]
61TABLE L Properties of 191P4D12(b) Bioinformatic Program URL
Outcome 191P4D12(b)B v.1 ORF ORF finder 264-1796 Protein length 510
aa Transmembrane region TM Pred http://www.ch.embnet.org/ 2 TM, aa
14-30, 351-370 HMMTop http://www.enzim.hu/hmmtop/ 1 TM, aa 347-371
Sosui http://www.genome.ad.jp/SOSui/ 2 TM, aa 14-31, 347-369 TMHMM
http://www.cbs.dtu.dk/services/TMHMM 1 TM, aa 350-372 Signal
Peptide Signal P http://www.cbs.dtu.dk/services/SignalP/ yes,
cleaved aa 31-32 pI pI/MW tool http://www.expasy.ch/tools/ pI 5.27
Molecular weight pI/MW tool http://www.expasy.ch/tools/ 55.4 kDa
Localization PSORT http://psort.nibb.ac.jp/ 46% plasma membrane
39.1% cytoplasmic, 21% PSORT II http://psort.nibb.ac.jp/ nuclear
Motifs Pfam http://www.sanger.ac.uk/Pfam/ Immunoglobulin domain
Prints http://www.biochem.ucl.ac.uk/ Cadherin signature Blocks
http://www.blocks.fhcrc.org/ Ig domain, Herpesvirus glycoprotein D
v.6 ORF ORF finder Protein length 295 aa Transmembrane region TM
Pred http://www.ch.embnet.org/ 1 TM, aa 135-156 HMMTop
http://www.enzim.hu/hmmtop/ 1 TM, aa 132-156 Sosui
http://www.genome.ad.jp/SOSui/ 1 TM, aa 132-154 TMHMM
http://www.cbs.dtu.dk/services/TMHMM 1 TM, aa 135-157 Signal
Peptide Signal P http://www.cbs.dtu.dk/services/SignalP/ none pI
pI/MW tool http://www.expasy.ch/tools/ pI 5.28 Molecular weight
pI/MW tool http://www.expasy.ch/tools/ 32.6 kDa Localization PSORT
http://psort.nibb.ac.jp/ 70% plasma membrane, 20% endoplasmic
reticulum 39% cytoplasmic, 21% PSORT II http://psort.nibb.ac.jp/
nuclear Motifs Pfam http://www.sanger.ac.uk/Pfam/ Immunoglobulin
domain Prints http://www.biochem.ucl.ac.uk/ none Blocks
http://www.blocks.fhcrc- .org/ Herpesvirus glycoprotein D
[1066]
62TABLE LI Exon boundaries of transcript 191P4D12(b) v.1 Exon
Number Start End Length 1 2 342 341 2 343 702 360 3 703 993 291 4
994 1114 121 5 1115 1263 149 6 1264 1420 157 7 1421 1496 76 8 1497
1571 75 9 1572 3459 1888
[1067]
63TABLE LII(a) Nucleotide sequence of transcript variant
191P4012(b) v.6 (SEQ ID NO: 105) ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg ctactgctgg catcatttac aggccggtgc cccgcgggtg
240 agctggagac ctcagacgtg gtaactgtgg tgctgggcca ggacgcaaaa
ctgccctgct 300 tctaccgagg ggactccggc gagcaagtgg ggcaagtggc
atgggctcgg gtggacgcgg 360 gcgaaggcgc ccaggaacta gcgctactgc
actccaaata cgggcttcat gtgagcccgg 420 cttacgaggg ccgcgtggag
cagccgccgc ccccacgcaa ccccctggac ggctcagtgc 480 tcctgcgcaa
cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc agcaccttcc 540
ccgccggcag cttccaggcg cggctgcggc tccgagtgct ggtgcctccc ctgccctcac
600 tgaatcctgg tccagcacta gaagagggcc agggcctgac cctggcagcc
tcctgcacag 660 ctgagggcag cccagccccc agcgtgacct gggacacgga
ggtcaaaggc acaacgtcca 720 gccgttcctt caagcactcc cgctctgctg
ccgtcacctc agagttccac ttggtgccta 780 gccgcagcat gaatgggcag
ccactgactt gtgtggtgtc ccatcctggc ctgctccagg 840 accaaaggat
cacccacatc ctccacgtgt ccttccttgc tgaggcctct gtgaggggcc 900
ttgaagacca aaatctgtgg cacattggca gagaaggagc tatgctcaag tgcctgagtg
960 aagggcagcc ccctccctca tacaactgga cacggctgga tgggcctctg
cccagtgggg 1020 tacgagtgga tggggacact ttgggctttc ccccactgac
cactgagcac agcggcatct 1080 acgtctgcca tgtcagcaat gagttctcct
caagggattc tcaggtcact gtggatgttc 1140 ttgaccccca ggaagactct
gggaagcagg tggacctagt gtcagcctcg gtggtggtgg 1200 tgggtgtgat
cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg ctcatgtccc 1260
gataccatcg gcgcaaggcc cagcagatga cccagaaata tgaggaggag ctgaccctga
1320 ccagggagaa ctccatccgg aggctgcatt cccatcacac ggaccccagg
agccagccgg 1380 aggagagtgt agggctgaga gccgagggcc accctgatag
tctcaaggac aacagtagct 1440 gctctgtgat gagtgaagag cccgagggcc
gcagttactc cacgctgacc acggtgaggg 1500 agatagaaac acagactgaa
ctgctgtctc caggctctgg gcgggccgag gaggaggaag 1560 atcaggatga
aggcatcaaa caggccatga accattttgt tcaggagaat gggaccctac 1620
gggccaagcc cacgggcaat ggcatctaca tcaatgggcg gggacacctg gtctgaccca
1680 ggcctgcctc ccttccctag gcctggctcc ttctgttgac atgggagatt
ttagctcatc 1740 ttgggggcct ccttaaacac ccccatttct tgcggaagat
gctccccatc ccactgactg 1800 cttgaccttt acctccaacc cttctgttca
tcgggagggc tccaccaatt gagtctctcc 1860 caccatgcat gcaggtcact
gtgtgtgtgc atgtgtgcct gtgtgagtgt tgactgactg 1920 tgtgtgtgtg
gaggggtgac tgtccgtgga ggggtgactg tgtccgtggt gtgtattatg 1980
ctgtcatatc agagtcaagt gaactgtggt gtatgtgcca cgggatttga gtggttgcgt
2040 gggcaacact gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc
tctgcctgaa 2100 aaagcaggta ttttctcaga ccccagagca gtattaatga
tgcagaggtt ggaggagaga 2160 ggtggagact gtggctcaga cccaggtgtg
cgggcatagc tggagctgga atctgcctcc 2220 ggtgtgaggg aacctgtctc
ctaccacttc ggagccatgg gggcaagtgt gaagcagcca 2280 gtccctgggt
cagccagagg cttgaactgt tacagaagcc ctctgccctc tggtggcctc 2340
tgggcctgct gcatgtacat attttctgta aatatacatg cgccgggagc ttcttgcagg
2400 aatactgctc cgaatcactt ttaatttttt tctttttttt ttcttgccct
ttccattagt 2460 tgtatttttt atttattttt atttttattt ttttttagag
atggagtctc actatgttgc 2520 tcaggctggc cttgaactcc tgggctcaag
caatcctcct gcctcagcct ccctagtagc 2580 tgggacttta agtgtacacc
actgtgcctg ctttgaatcc tttacgaaga gaaaaaaaaa 2640 attaaagaaa
gcctttagat ttatccaatg tttactactg ggattgctta aagtgaggcc 2700
cctccaacac cagggggtta attcctgtga ttgtgaaagg ggctacttcc aaggcatctt
2760 catgcaggca gccccttggg agggcacctg agagctggta gagtctgaaa
ttagggatgt 2820 gagcctcgtg yttactgagt aaggtaaaat tgcatccacc
attgtttgtg ataccttagg 2880 gaattgcttg gacctggtga caagggctcc
tgttcaatag tggtgttggg gagagagaga 2940 gcagtgatta tagaccgaga
gagtaggagt tgaggtgagg tgaaggaggt gctgggggtg 3000 agaatgtcgc
ctttccccct gggttttgga tcactaattc aaggctcttc tggatgtttc 3060
tctgggttgg ggctggagtt caatgaggtt tatttttagc tggcccaccc agatacactc
3120 agccagaata cctagattta gtacccaaac tcttcttagt ctgaaatctg
ctggatttct 3180 ggcctaaggg agaggctccc atccttcgtt ccccagccag
cctaggactt cgaatgtgga 3240 gcctgaagat ctaagatcct aacatgtaca
ttttatgtaa atatgtgcaa atttgtacat 3300 aaaatgatat tctgttttta
aataaacaga caaaacttga aaaa 3344
[1068]
64TABLE LIII(a) Nucleotide sequence alignment of 191P4D12(b) v.1
(SEQ ID NO: 106) and 191 P4012(b) v.6 (SEQ ID NO: 107). V.1 1
gGCCGTCGTTGTTGGCCACAGCGTGGGAAGCAGCTCTG- GGGGAGCTCGGA 50
.vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline. V.6 1 ggccgtcgttgttqgccacagcqtgggaagcagctctggqggagctcgga
50 V.1 51 GCTCCCGATCACGGCTTCTTGGOGGTAGCTACGGCTGGGTGTGTAGA- ACG 100
.vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.6 51 gctcccgatcacggcttcttgggqgtagctacggctgggtgtgtagaacg 100 V.1
101 GGGCCGGGGCTGGGGCTGGGTCCCCTAGTGGAGACCCAAGTGCGAGAGGC 150
.vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline.
V.6 101 gggccggggctggggctgggtcccctagtggagacccaagtgcgagaggc 150 V.1
151 AAGAACTCTGCAGCTTCCTGCCTTCTGGGTCAGTTCCTTATTCAAGTCTG 200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline. V.6 151
aagaactctgcaqcttcctqccttctgg- gtcagttccttattcaagt--- 197 V.1 201
CAGCCGCCTCCCAGGGAGATCTC- GGTGGAACTTCAGAAACGCTGGGCAGT 250 V.6 198
-------------------------------------------------- 197 V.1 251
CTCCCTTTCAACCATGCCCCTGTCCCTGGGAGCCGAGATGTGGGGGCCTG 300 V.6 198
-------------------------------------------------- 197 V.1 301
AGGCCTGGCTGCTGCTGCTGCTACTGCTGGCATCATTTACAGGCCGGTGC 350
.vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline. V.6
198 -----------------ctgctactgctgqcatcatttacagqccggtgc 230 V.1 351
CCCGCGGGTGACCTGGAGACCTCAGACGTGGTAACTGTGGTGCTGGGCCA 400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 231
cccgcgqgtgagctggagacctcaqacgtggtaactgtggtgctgggcca 280 V.1 401
GGACGCAAAACTGCCCTGCTTCTACCGAGGGGACTCCGGCGAGCAAGTGG 450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 281
ggacgcaaaactgccctgcttctaccgaggggactccggcgagcaagtgg 330 V.1 451
GGCAAGTGGCATGGGCTCGGGTGGACGCGGGCGAAGGCGCCCAGGAACTA 500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 331
ggcaagtggcatgggctcgggtggacgcgggcgaaqgcgcccaggaacta 380 V.1 501
GCGCTACTGCACTCCAAATACGGGCTTCATGTGAGCCCGGCTTACGAGGG 550
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 381
gcgctactgcactccaaatacgggcttcatgtgagcccggcttacgaggg 430 V.1 551
CCGCGTGGAGCAGCCGCCGCCCCCACGCAACCCCCTGGACGGCTCAGTGC 600
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 431
ccgcgtggagcagccgccgcccccacgcaaccccctggacggctcagtgc 480 V.1 601
TCCTGCGCAACGCAGTGCAGGCGGATGAGGGCGAGTACGAGTGCCGGGTC 650
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 481
tcctgcgcaacgcagtgcaggcygatgagggcgagtacgagtgccgggtc 530 V.1 651
AGCACCTTCCCCGCCGGCAGCTTCCAGGCGCGGCTGCGGCTCCGAGTGCT 700
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 531
agcaccttccccgccggcagcttccaggcgcggctgcggctccgagtgct 580 V.1 701
GCTGCCTCCCCTGCCCTCACTGAATCCTGGTCCAGCACTAGAAGAGGGCC 750
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 581
ggtgcctcccctgccctcactgaatcctggtccagcactagaagagggcc 630 V.1 751
AGGGCCTGACCCTGGCAGCCTCCTGCACAGCTGAGGGCAGCCCAGCCCCC 800
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 631
agggcctgaccctggcagcctcctgcacagctgagggcagcccagccccc 680 V.1 801
AGCGTGACCTGGGACACGGAGGTCAAAGGCACAACGTCCAGCCGTTCCTT 850
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 681
agcgtgacctgggacacggaggtcaaaggcacaacgtccagccgttcctt 730 V.1 851
CAAGCACTCCCGCTCTGCTGCCGTCACCTCAGAGTTCCACTTGGTGCCTA 900
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 731
caagcactcccgctctgctgccgtcacctcagagttccacttggtgccta 780 V.1 901
GCCGCAGCATGAATGGGCAGCCACTGACTTGTGTGGTGTCCCATCCTGGC 950
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 781
gccgcagcatgaatgggcagccactgacttgtgtggtgtcccatcctggc 830 V.1 951
CTGCTCCAGGACCAAAGGATCACCCACATCCTCCACGTGTCCTTCCTTGC 1000
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 831
ctgctccaggaccaaaggatcacccacatcctccacgtgtccttccttgc 880 V.1 1001
TGAGGCCTCTGTGAGGGGCCTTGAAGACCAAAATCTGTGGCACATTGGCA 1050
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 881
tgaggcctctgtgaggggccttgaagaccaaaatctgtggcacattggca 930 V.1 1051
GAGAACGAGCTATGCTCAAGTGCCTGAGTGAAGGGCAGCCCCCTCCCTCA 1100
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 931
gagaaggagctatgctcaagtgcctgagtgaagggcagccccctccctca 980 V.1 1101
TACAACTGGACACGGCTGGATGGGCCTCTGCCCAGTGGGGTACGAGTGGA 1150
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 981
tacaactggacacggctggatgggcctctgcccagtggggtacgagtgga 1030 V.1 1151
TGGGGACACTTTGGGCTTTCCCCCACTGACCACTGAGCACAGCGGCATCT 1200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1031
tggggacactttgggctttcccccactgaccactgagcacagcggcatct 1080 V.1 1201
ACGTCTGCCATGTCAOCAATGAGTTCTCCTCAAGGGATTCTCAGGTCACT 1250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1081
acgtctgccatgtcagcaatgagttctcctcaagggattctcaggtcact 1130 V.1 1251
GTGGATCTTCTTGACCCCCAGGAAGACTCTGGGAAGCAGGTGGACCTAGT 1300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1131
gtggatgttcttgacccccaggaagactctgggaagcaggtggacctagt 1180 V.1 1301
GTCAGCCTCGGTGGTGGTGGTGGGTGTGATCGCCGCACTCTTGTTCTGCC 1350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1181
gtcagcctcggtggtggtggtgggtgtgatcgccgcactcttgttctgcc 1230 V.1 1351
TTCTGGTCGTGGTGGTGGTGCTCATGTCCCGATACCATCGGCGCAAGGCC 1400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1231
ttctggtggtggtggtggtgctcatgtcccgataccatcggcgcaaggcc 1280 V.1 1401
CAGCAGATGACCCAGAAATATGAGGAGGAGCTGACCCTGACCAGGGAGAA 1450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1281
cagcagatgacccagaaatatgaggaggagctgaccctgaccagggaaaa 1330 V.1 1451
CTCCATCCGGAGGCTGCATTCCCATCACACGGACCCCAGGAGCCAGCCGG 1500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1331
ctccatccggaggctgcattcccatcacacggaccccaggagccagccgg 1380 V.1 1501
AGGAGAGTGTAGGGCTGAGAGCCGAGGGCCACCCTGATAGTCTCAAGGAC 1550
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1381
aggagagtgtagqgctgagagccgagggccaccctgatagtctcaaggac 1430 V.1 1551
AACAGTAGCTGCTCTGTGATGAGTGAAGAGCCCGAGGGCCGCAGTTACTC 1600
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1431
aacagtagctgctctgtgatgagtgaagagcccgagggccgcagttactc 1480 V.1 1601
CACGCTGACCACGGTGAGGGAGATAGAAACACAGACTGAACTGCTGTCTC 1650
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1481
cacgctgaccacggtgagggagatagaaacacagactgaactgctgtctc 1530 V.1 1651
CAGGCTCTGGGCGGGCCGAGGAGGAGGAAGATCAGGATGAAGGCATCAAA 1700
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1531
caggctctgggcgggccgaggaggaggaagatcaggatgaaggcatcaaa 1580 V.1 1701
CAGGCCATGAACCATTTTGTTCAGGAGAATGCGACCCTACGGGCCAAGCC 1750
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1581
caggccatgaaccattttgttcaggagaatgggaccctacgggccaagcc 1630 V.1 1751
CACGGGCAATGGCATCTACATCAATGGGCGGGGACACCTGGTCTGACCCA 1800
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1631
cacgggcaatggcatctacatcaatgggcggqgacacctggtctgaccca 1680 V.1 1801
GGCCTGCCTCCCTTCCCTAGGCCTGGCTCCTTCTGTTGACATGGGAGATT 1850
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1681
ggcctgcctcccttccctaggcctggctccttctgttgacatgggagatt 1730 V.1 1851
TTAGCTCATCTTGGGGGCCTCCTTAAACACCCCCATTTCTTGCGGAAGAT 1900
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1731
ttagctcatcttgggggcctccttaaacacccccatttcttgcggaagat 1780 V.1 1901
GCTCCCCATCCCACTGACTGCTTGACCTTTACCTCCAACCCTTCTGTTCA 1950
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1781
gctccccatcccactgactgcttgacctttacctccaacccttctgttca 1830 V.1 1951
TCGGGAGGGCTCCACCAATTGAGTCTCTCCCACCATGCATGCAGGTCACT 2000
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1831
tcgggagggctccaccaattgagtctctcccaccatgcatgcaggtcact 1880 V.1 2001
GTGTGTGTGCATGTGTGCCTGTGTGAGTGTTGACTGACTGTGTGTGTGTG 2050
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1881
qtgtgtgtgcatgtgtgcctgtgtgagtgttgactgactgtgtgtgtgtg 1930 V.1 2051
GAGGGGTGACTGTCCGTGGAGGGGTGACTGTGTCCGTGGTGTGTATTATG 2100
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1931
gaggggtgactgtccgtggaggggtgactgtgtccgtggtgtgtattatg 1980 V.1 2101
CTGTCATATCAGAGTCAAGTGAACTGTGGTGTATCTGCCACGGGATTTGA 2150
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 1981
ctgtcatatcagagtcaagtgaactgtggtgtatgtgccacgggatttga 2030 V.1 2151
GTGGTTGCGTGGGCAACACTGTCAGGGTTTGGCGTGTGTGTCATGTGGCT 2200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2031
gtggttgcgtgggcaacactgtcagggtttggcgtgtgtgtcatgtggct 2080 V.1 2201
GTGTGTGACCTCTGCCTGAAAAAGCAGGTATTTTCTCAGACCCCAGAGCA 2250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2081
gtgtgtgacctctgcctgaaaaagcaggtattttctcagaccccagagca 2130 V.1 2251
GTATTAATGATGCAGAGGTTGGAGGAGAGAGGTGGAGACTGTGGCTCAGA 2300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2131
gtattaatgatgcagaggttggaggagagagqtggagactgtggctcaga 2180 V.1 2301
CCCAGGTGTGCGGGCATAGCTGGAGCTGGAATCTGCCTCCGGTGTGAGGG 2350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2181
cccaggtgtgcgggcatagctggagctggaatctgcctccggtgtgaggg 2230 V.1 2351
AACCTGTCTCCTACCACTTCGGAGCCATGGGGGCAAGTGTGAAGCAGCCA 2400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2231
aacctgtctcctaccacttcggagccatgggggcaagtgtgaagcagcca 2280 V.1 2401
GTCCCTGGGTCAGCCAGAGGCTTGAACTGTTACAGAAGCCCTCTGCCCTC 2450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2281
gtccctgggtcagccagaggcttgaactgttacagaagccctctgccctc 2330 V.1 2451
TGGTGGCCTCTGGGCCTGCTGCATGTACATATTTTCTGTAAATATACATG 2500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2331
tggtqgcctctgggcctgctgcatgtacatattttctgtaaatatacatg 2380 V.1 2501
CGCCGGGAGCTTCTTGCAGGAATACTGCTCCGAATCACTTTTAATTTTTT 2550
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2381
cgccgggagcttcttgcaggaatactgctccgaatcacttttaatttttt 2430 V.1 2551
TCTTTTTTTTTTCTTGCCCTTTCCATTAGTTGTATTTTTTATTTATTTTT 2600
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2431
tcttttttttttcttgccctttccattagttgtattttttatttattttt 2480 V.1 2601
ATTTTTATTTTTTTTTAGAGATGGAGTCTCACTATGTTGCTCAGGCTGGC 2650
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2481
atttttatttttttttagaqatggagtctcactatgttgctcaggctggc 2530 V.1 2651
CTTGAACTCCTGGGCTCAAGCAATCCTCCTGCCTCAGCCTCCCTAGTAGC 2700
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2531
cttgaactcctgggctcaagcaatcctcctgcctcagcctccctagtagc 2580 V.1 2701
TGGGACTTTAAGTGTACACCACTGTGCCTGCTTTGAATCCTTTACGAACA 2750
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2581
tgggactttaagtgtacaccactgtgcctgctttgaatcctttacgaaca 2630 V.1 2751
GAAAAAAAAAATTAAAGAAAGCCTTTAGATTTATCCAATGTTTACTACTG 2800
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2631
gaaaaaaaaaattaaagaaagcctttagatttatccaatgtttactactg 2680 V.1 2801
GGATTGCTTAAAGTGAGGCCCCTCCAACACCAGGGGGTTAATTCCTGTGA 2850
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2681
ggattgcttaaagtgaggcccctccaacaccagggggttaattcctgtga 2730 V.1 2851
TTGTGAAAGGGGCTACTTCCAAGGCATCTTCATGCAGGCAGCCCCTTGGG 2900
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2731
ttgtgaaaggggctacttccaaggcatcttcatgcaggcagccccttggg 2780 V.1 2901
AGGGCACCTGAGAGCTGGTAGAGTCTGAAATTAGGGATGTGAGCCTCGTG 2950
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2781
agggcacctgagagctggtagagtctgaaattagggatgtgagcctcgtg 2830 V.1 2951
GTTACTGAGTAAGGTAAAATTGCATCCACCATTGTTTGTGATACCTTAGG 3000
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2831
gttactgagtaaggtaaaattggatggaggattgtttgtgataggttagg 2880 V.1 3001
GAATTGCTTGGACCTGGTGACAAGGGCTCCTGTTCAATAGTGGTGTTGGG 3050
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2881
gaattgcttggacctggtgacaagggctcctgttcaatagtggtgttggg 2930 V.1 3051
GAGAGAGAGAGCAGTGATTATAGACCGAGAGAGTAGGAGTTGAGGTGAGG 3100
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2931
gagagagagagcagtgattatagaccgagagagtaggagttgaqgtgagg 2980 V.1 3101
TGAAGGAGGTGCTGGGGGTGAGAATGTCGCCTTTCCCCCTGGGTTTTGGA 3150
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 2981
tgaaggaggtgctggqggtgagaatgtcgcctttccccctgggttttgga 3030 V.1 3151
TCACTAATTCAAGGCTCTTCTGGATGTTTCTCTGGGTTGGGGCTGGAGTT 3200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 3031
tcactaattcaaggctcttctggatgtttctctgggttggggctggagtt 3080 V.1 3201
CAATGAGGTTTATTTTTAGCTGGCCCACCCAGATACACTCAGCCAGAATA 3250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 3081
caatgaggtttatttttagctggcccacccagatacactcagccagaata 3130 V.1 3251
CCTAGATTTAGTACCCAAACTCTTCTTAGTCTGAAATCTGCTGGATTTCT 3300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 3131
cctagatttagtacccaaactcttcttagtctgaaatctgctggatttct 3180 V.1 3301
GGCCTAAGGGAGAGGCTCCCATCCTTCGTTCCCCAGCCAGCCTAGGACTT 3350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 3181
ggcctaagggagaggctcccatccttcgttccccagccagcctaggactt 3230 V.1 3351
CGAATGTGGAGCCTGAAGATCTAAGATCCTAACATGTACATTTTATGTAA 3400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 3231
cgaatgtggagcctgaagatctaagatcctaacatgtacattttatgtaa 3280 V.1 3401
ATATGTGCATATTTGTACATAAAATGATATTCTGTTTTTAAATAAACAGA 3450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 3281
atatgtgcatatttgtacataaaatgatattctgtttttaaataaacaga 3330 V.1 3451
CAAAACTTGaaaaa 3464 .vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline. V.6 3331 caaaacttgaaaaa 3344
[1069]
65TABLE LIV(a) Peptide seguences of protein coded by 191P4D12(b)
v.6 (SEQ ID NO: 108) MNGQPLTCVV SHPGLLQDQR ITHILHVSFL AEASVRGLED
QNLWHIGREG AMLKCLSBGQ 60 PPPSYNWTRL DGPLPSGVRV DGDTLGFPPL
TTEHSGIYVC HVSNEFSSRD SQVTVDVLDP 120 QEDSGKQVDL VSASVVVVGV
IAALLFCLLV VVVVLMSRYH RRKAQQMTQK YEEELTLTRE 180 NSIRRLHSHH
TDPRSQPEES VGLPAEGHPD SLKDNSSCSV MSEEPEGRSY STLTTVREIE 240
TQTELLSPGS GRAEEEEDQD EGIKQAMNHF VQENGTLRAK PTGNGIYING RGHLV
295
[1070]
66TABLE LV(a) Amino acid sequence alignment of 191P4D12(b) v.1 (SEQ
ID NO: 109) and 191P4D12(b) v.6 (SEQ ID NO: 110) V.1 216
MNGQPLTCVVSHPGLLQDQRITHILHVSFLAEASVRG- LEDQNLWHIGREG 265
.vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline. V.6 1 MNGQPLTCVVSHPGLLQDQRITHILHVSFLAEASVRGLEDQNLWHIGREG
50 V.1 266 AMLKCLSEGQPPPSYNWTRLDGPLPSGVRVDGDTLGFPPLTTEHSG- IYVC 315
.vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.6 51 AMLKCLSEGQPPPSYNWTRLDGPLPSGVRVDGDTLGFPPLTTEHSGIYVC 100 V.1
316 HVSNEPSSRDSQVTVDVLDPQEDSGKQVDLVSASVVVVGVIAALLFCLLV 365
.vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline.
V.6 101 HVSNEFSSRDSQVTVDVLDPQEDSGKQVDLVSASVVVVGVIAALLFCLLV 150 V.1
366 VVVVLMSRYHRRKAQQMTQKYEEELTLTRENSIRRLHSHHTDPRSQPEES 415
.vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline. V.6
151 VVVVLMSRYHRRKAQQMTQKYEEELTLTRENSIRRLHSHHTDPRSQPEES 200 V.1 416
VGLRAEGHPDSLKDNSSCSVMSEEPEGRSYSTLTTVREIETQTELLSPGS 465
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 201
VGLRAEGHPDSLKDNSSCSVMSEEPEGRSYSTLTTVREIETQTELLSPGS 250 V.1 466
GRAEEEEDQDEGIKQAMNHFVQENGTLRAKPTGNGIYINGRGHLV 510
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.6 251
GRAEEEEDQDEGIKQAMNHFVQENGTLRAKPTGNGIYINGRGHLV 295
[1071]
67TABLE LII(b) Nucleotide seguence of transcript variant
191P4D12(b) v.7 (SEQ ID NO: 111) ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac
240 gctgggcagt ctgcctttca accatgcccc tgtccctggg agccgagatg
tgggggcctg 300 aggcctggct gctgctgctg ctactgctgg catcatttac
aggccggtgc cccgcgggtg 360 agctggagac ctcagacgtg gtaactgtgg
tgctgggcca ggacgcaaaa ctgccctgct 420 tctaccgagg ggactccggc
gagcaagtgg ggcaagtggc atgggctcgg gtggacgcgg 480 gcgaaggcgc
ccaggaacta gcgctactgc actccaaata cgggcttcat gtgagcccgg 540
cttacgaggg ccgcgtggag cagccgccgc ccccacgcaa ccccctggac ggctcagtgc
600 tcctgcgcaa cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc
agcaccttcc 660 ccgccggcag cttccaggcg cggctgcggc tccgagtgct
ggtgcctccc ctgccctcac 720 tgaatcctgg tccagcacta gaagagggcc
agggcctgac cctggcagcc tcctgcacag 780 ctgagggcag cccagccccc
agcgtgacct gggacacgga ggtcaaaggc acaacgtcca 840 gccgttcctt
caagcactcc cgctctgctg ccgtcacctc agagttccac ttggtgccta 900
gccgcagcat gaatgggcag ccactgactt gtgtggtgtc ccatcctggc ctgctccagg
960 accaaaggat cacccacatc ctccacgtgt ccttccttgc tgaggcctct
gtgaggggcc 1020 ttgaagacca aaatctgtgg cacattggca gagaaggagc
tatgctcaag tgcctgagtg 1080 aagggcagcc ccctccctca tacaactgga
cacggctgga tgggcctctg cccagtgggg 1140 tacgagtgga tggggacact
ttgggctttc ccccactgac cactgagcac agcggcatct 1200 acgtctgcca
tgtcagcaat gagttctcct caagggattc tcaggtcact gtggatgttc 1260
ttgaccccca ggaagactct gggaagcagg tggacctagt gtcagcctcg gtggtggtgg
1320 tgggtgtgat cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg
ctcatgtccc 1380 gataccatcg gcgcaaggcc cagcagatga cccagaaata
tgaggaggag ctgaccctga 1440 ccagggagaa ctccatccgg aggctgcatt
cccatcacac ggaccccagg agccagagtg 1500 aagagcccga gggccgcagt
tactccacgc tgaccacggt gagggagata gaaacacaga 1560 ctgaactgct
gtctccaggc tctgggcggg ccgaggagga ggaagatcag gatgaaggca 1620
tcaaacaggc catgaaccat tttgttcagg agaatgggac cctacgggcc aagcccacgg
1680 gcaatggcat ctacatcaat gggcggggac acctggtctg acccaggcct
gcctcccttc 1740 cctaggcctg gctccttctg ttgacatggg agattttagc
tcatcttggg ggcctcctta 1800 aacaccccca tttcttgcgg aagatgctcc
ccatcccact gactgcttga cctttacctc 1860 caacccttct gttcatcggg
agggctccac caattgagtc tctcccacca tgcatgcagg 1920 tcactgtgtg
tgtgcatgtg tgcctgtgtg agtgttgact gactgtgtgt gtgtggaggg 1980
gtgactgtcc gtggaggggt gactgtgtcc gtggtgtgta ttatgctgtc atatcagagt
2040 caagtgaact gtggtgtatg tgccacggga tttgagtggt tgcgtgggca
acactgtcag 2100 ggtttggcgt gtgtgtcatg tggctgtgtg tgacctctgc
ctgaaaaagc aggtattttc 2160 tcagacccca gagcagtatt aatgatgcag
aggttggagg agagaggtgg agactgtggc 2220 tcagacccag gtgtgcgggc
atagctggag ctggaatctg cctccggtgt gagggaacct 2280 gtctcctacc
acttcggagc catgggggca agtgtgaagc agccagtccc tgggtcagcc 2340
agaggcttga actgttacag aagccctctg ccctctggtg gcctctgggc ctgctgcatg
2400 tacatatttt ctgtaaatat acatgcgccg ggagcttctt gcaggaatac
tgctccgaat 2460 cacttttaat ttttttcttt tttttttctt gccctttcca
ttagttgtat tttttattta 2520 tttttatttt tatttttttt tagagatgga
gtctcactat gttgctcagg ctggccttga 2580 actcctgggc tcaagcaatc
ctcctgcctc agcctcccta gtagctggga ctttaagtgt 2640 acaccactgt
gcctgctttg aatcctttac gaagagaaaa aaaaaattaa agaaagcctt 2700
tagatttatc caatgtttac tactgggatt gcttaaagtg aggcccctcc aacaccaggg
2760 ggttaattcc tgtgattgtg aaaggggcta cttccaaggc atcttcatgc
aggcagcccc 2820 ttgggagggc acctgagagc tggtagagtc tgaaattagg
gatgtgagcc tcgtggttac 2880 tgagtaaggt aaaattgcat ccaccattgt
ttgtgatacc ttagggaatt gcttggacct 2940 ggtgacaagg gctcctgttc
aatagtggtg ttggggagag agagagcagt gattatagac 3000 cgagagagta
ggagttgagg tgaggtgaag gaggtgctgg gggtgagaat gtcgcctttc 3060
cccctgggtt ttggatcact aattcaaggc tcttctggat gtttctctgg gttggggctg
3120 gagttcaatg aggtttattt ttagctggcc cacccagata cactcagcca
gaatacctag 3180 atttagtacc caaactcttc ttagtctgaa atctgctgga
tttctggcct aagggagagg 3240 ctcccatcct tcgttcccca gccagcctag
gacttcgaat gtggagcctg aagatctaag 3300 atcctaacat gtacatttta
tgtaaatatg tgcatatttg tacataaaat gatattctgt 3360 ttttaaataa
acagacaaaa cttgaaaaa 3389
[1072]
68TABLE LIII(b) Nucleotide sequence alignment of 191P4D12(b) v.1
(SEQ ID NO: 112) and 191P4D12(b) v.7 (SEQ ID NO: 113) V.1 1
gGCCGTCGTTGTTGGCCACAGCGTGGGAAGCAGCTCTGG- GGGAGCTCGGA 50
.vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline. V.7 1 ggccgtcgttgttggccacagcgtgggaagcagctctqggggagctcgga 50
V.1 51 GCTCCCGATCACGGCTTCTTGGGGGTAGCTACGGCTGGGTGTGTAGAACA 100
.vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.7 51 gctcccgatcacggcttcttgggggtagctacggctgggtqtgtagaacg 100 V.1
101 GGGCCGGGGCTGGGGCTGGGTCCCCTAGTGGAGACCCAAGTGCGAGAGGC 150
.vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline. V.7
101 gggccggggctggggctgggtcccctagtggagacccaagtgcgagaggc 150 V.1 151
AAGAACTCTGCAGCTTCCTGCCTTCTGGGTCAGTTCCTTATTCAAGTCTG 200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 151
aagaactctgcagcttcctgccttctgggtcagttccttattcaagtctg 200 V.1 201
CAGCCGGCTCCCAGGGAGATCTCGGTGGAACTTCAGAAACGCTGGGCAGT 250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 201
cagccggctcccagggagatctcggtggaacttcagaaacgctgggcagt 250 V.1 251
CTGCCTTTCAACCATGCCCCTGTCCCTGGGAGCCGAGATGTGGGGGCCTG 300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 251
ctgcctttcaaccatgcccctgtccctgggagccgagatgtgggggcctg 300 V.1 301
AGGCCTGGCTGCTGCTGCTGCTACTGCTGGCATCATTTACAGGCCGGTGC 350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 301
aggcctggctgctgctgctgctactgctggcatcatttacaggccggtgc 350 V.1 351
CCCGCGGGTGAGCTGGAGACCTCAGACGTGGTAACTGTGGTGCTGGGCCA 400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 351
cccgcgggtgagctggagacctcagacgtggtaactgtggtgctgggcca 400 V.1 401
GGACCCAAAACTGCCCTGCTTCTACCGAGGGGACTCCGGCGAGCAAGTGG 450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 401
ggacccaaaactgccctgcttctaccgaggggactccggcgagcaagtgg 450 V.1 451
GGCAAGTGGCATGGGCTCGGGTGGACCCGCGCGAAGGCGCCCAGGAACTA 500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 451
ggcaagtggcatgggctcgggtqgacgcgggcgaaggcgcccaggaacta 500 V.1 501
GCGCTACTGCACTCCAAATACGGGCTTCATGTGAGCCCGGCTTACGAGGG 550
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 501
gcgctactgcactccaaatacgggcttcatgtgagcccggcttacgaggg 550 V.1 551
CCGCGTGGAGCAGCCGCCGCCCCCACGCAACCCCCTGGACCCCTCAGTGC 600
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 551
ccgcgtggagcagccgccgcccccacgcaaccccctggacggctcagtgc 600 V.1 601
TCCTGCGCAACGCAGTGCAGGCGGATGAGGGCGAGTACGAGTGCCGGGTC 650
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 601
tcctgcgcaacgcagtgcaggcggatgagggcgagtacgagtgccgggtc 650 V.1 651
AGCACCTTCCCCGCCGGCAGCTTCCAGGCGCGGCTGCGGCTCCGAGTGCT 700
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 651
agcaccttccccgccggcagcttccaggcgcggctgcggctccgagtgct 700 V.1 701
GGTGCCTCCCCTGCCCTCACTGAATCCTGGTCCAGCACTAGAAGAGGGCC 750
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 701
ggtgcctcccctgccctcactgaatcctggtccagcactagaagagggcc 750 V.1 751
AGGGCCTGACCCTGGCAGCCTCCTGCACAGCTGAGGGCAGCCCAGCCCCC 800
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 751
agggcctgaccctggcagcctcctgcacagctgagggcagcccagccccc 800 V.1 801
AGCGTGACCTGGGACACGGAGGTCAAAGGCACAACGTCCAGCCGTTCCTT 850
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 801
agcgtgacctgggacacggaggtcaaagqcacaacgtccagccgttcctt 850 V.1 851
CAAGCACTCCCGCTCTGCTGCCGTCACCTCAGAGTTCCACTTGGTGCCTA 900
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 851
caagcactcccgctctgctgccgtcacctcagagttccacttggtgccta 900 V.1 901
GCCGCAGCATGAATGGGCAGCCACTGACTTGTGTGGTGTCCCATCCTGGC 950
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 901
gccgcagcatgaatgggcagccactgacttgtgtggtgtcccatcctggc 950 V.1 951
CTGCTCCAGGACCAAAGGATCACCCACATCCTCCACGTGTCCTTCCTTGC 1000
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 951
ctgctccaggaccaaaggatcacccacatcctccacgtgtccttccttgc 1000 V.1 1001
TGAGGCCTCTGTGAGGGGCCTTGAAGACCAAAATCTGTGGCACATTGGCA 1050
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1001
tgaggcctctgtgaggggccttgaagaccaaaatctgtggcacattggca 1050 V.1 1051
GAGAAGGAGCTATGCTCAAGTGCCTGAGTGAAGGGCAGCCCCCTCCCTCA 1100
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1051
gagaaggagctatgctcaagtgcctgagtgaagggcagccccctccctca 1100 V.1 1101
TACAACTGGACACGGCTGGATGGGCCTCTGCCCAGTGGGGTACGAGTGGA 1150
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1101
tacaactggacacggctggatgggcctctgcccagtggggtacgagtgga 1150 V.1 1151
TGGGGACACTTTGGGCTTTCCCCCACTGACCACTGAGCACAGCGGCATCT 1200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1151
tggqgacactttgggctttcccccactgaccactgagcacagcggcatct 1200 V.1 1201
ACGTCTCCCATGTCAGCAATGAGTTCTCCTCAAGGGATTCTCAGGTCACT 1250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1201
acgtctgccatgtcagcaatgagttctcctcaagggattctcaggtcact 1250 V.1 1251
GTCGATGTTCTTGACCCCCAGGAAGACTCTGGGAAGCAGGTGGACCTAGT 1300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1251
gtggatgttcttgacccccaggaagactctgggaagcaggtggacctagt 1300 V.1 1301
GTCAGCCTCGGTGGTGGTGGTGGGTGTGATCGCCGCACTCTTGTTCTGCC 1350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1301
gtcagcctcggtggtggtggtgggtgtgatcgccgcactcttgttctgcc 1350 V.1 1351
TTCTGGTGGTGGTGGTGGTGCTCATGTCCCGATACCATCGGCGCAACGCC 1400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1351
ttctggtggtggtggtggtgctcatgtcccgataccatcggcgcaaggcc 1400 V.1 1401
CAGCAGATGACCCAGAAATATGAGGAGGAGCTGACCCTGACCAGGGAGAA 1450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1401
cagcagatgacccagaaatatgaggaggagctgaccctgaccagggagaa 1450 V.1 1451
CTCCATCCGGAGGCTGCATTCCCATCACACGGACCCCAGGAGCCAGCCGG 1500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 1451
ctccatccggaggctgcattcccatcacacggaccccaggagcca----- 1495 V.1 1501
AGGAGAGTGTAGGGCTGAGAGCCGAGGGCCACCCTGATAGTCTCAAGGAC 1550 V.7 1496
-------------------------------------------------- 1495 V.1 1551
AACAGTAGCTGCTCTGTGATGAGTGAAGAGCCCGAGGGCCGCAGTTACTC 1600 V.7 1496
--------------------gagtgaagagcccgagggccgca- gttactc 1525 V.1 1601
CACGCTCACCACGGTGAGGGAGATAGAAACACAGA- CTGAACTGCTCTCTC 1650
.vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline. V.7 1526
cacgctgaccacggtgagggagatagaaacacagactgaactgct- gtctc 1575 V.1 1651
CAGGCTCTGGGCGGGCCGAGGAGGAGGAAGATCAGGAT- GAAGGCATCAAA 1700
.vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline. V.7 1576
caggctctgggcgggccgaggaggaggaagatcaggatgaaggcatca- aa 1625 V.1 1701
CAGGCCATGAACCATTTTGTTCAGGAGAATGGGACCCTACG- GGCCAAGCC 1750
.vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline. V.7 1626 caggccatgaaccattttgttcaggagaatgggaccctacgggccaagcc
1675 V.1 1751 CACGGGCAATGGCATCTACATCAATGGGCGGGGACACCTGGTC- TGACCCA
1800 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 1676 cacgggcaatggcatctacatcaatgggcggggacacctggtctgaccca
1725 V.1 1801 GGCCTGCCTCCCTTCCCTAGGCCTGGCTCCTTCTGTTGACATG- GGAGATT
1850 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 1726 ggcctgcctcccttccctaggcctggctccttctgttgacatgggagatt
1775 V.1 1851 TTAGCTCATCTTGGGGGCCTCCTTAAACACCCCCATTTCTTGC- GGAAGAT
1900 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 1776 ttagctcatcttgggggcctccttaaacacccccatttcttgcggaagat
1825 V.1 1901 GCTCCCCATCCCACTGACTGCTTGACCTTTACCTCCAACCCTT- CTGTTCA
1950 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 1826 gctccccatcccactgactgcttgacctttacctccaacccttctgttca
1875 V.1 1951 TCCGGAGGGCTCCACCAATTGAGTCTCTCCCACCATGCATGCA- GGTCACT
2000 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 1876 tcgggagggctccaccaattgagtctctcccaccatgcatgcaggtcact
1925 V.1 2001 GTGTGTGTGCATGTGTGCCTGTGTGAGTGTTGACTGACTGTGT- GTGTGTG
2050 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 1926 gtgtgtgtgcatgtgtgcctgtgtgagtgttgactgactgtgtgtgtgtg
1975 V.1 2051 GAGGGGTGACTGTCCGTGGAGGGGTGACTGTGTCCGTGGTGTG- TATTATG
2100 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 1976 gaggggtgactgtccgtggaggggtgactgtgtccgtggtgtgtattatg
2025 V.1 2101 CTGTCATATCAGAGTCAAGTGAACTGTGGTGTATGTGCCACGG- GATTTGA
2150 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2026 ctgtcatatcagagtcaagtgaactgtggtgtatgtgccacgggatttga
2075 V.1 2151 GTGGTTGCGTGGGCAACACTGTCAGGGTTTGGCGTGTGTGTCA- TGTGGCT
2200 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2076 gtggttgcgtgggcaacactgtcagggtttggcgtgtgtgtcatgtggct
2125 V.1 2201 GTGTGTGACCTCTGCCTGAAAAAGCAGGTATTTTCTCACACCC- CAGAGCA
2250 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2126 gtgtgtgacctctgcctgaaaaagcaggtattttctcagaccccagagca
2175 V.1 2251 GTATTAATGATGCAGACGTTGGAGGAGAGAGGTGGAGACTGTG- GCTCAGA
2300 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2176 gtattaatgatgcagaggttggaggagagaggtggagactgtggccCaga
2225 V.1 2301 CCCAGGTGTGCGGGCATAGCTGGAGCTGGAATCTGCCTCCGGT- GTGAGGG
2350 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2226 cccaggtgtgcgggcatagctggagctggaatctgcctccggtgtgaggg
2275 V.1 2351 AACCTGTCTCCTACCACTTCGGAGCCATGGGGGCAAGTGTGAA- GCAGCCA
2400 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2276 aacctgtctcctaccacttcggagccatgggggcaagtgtgaagcagcca
2325 V.1 2401 GTCCCTGGGTCAGCCAGAGGCTTGAACTGTTACAGAAGCCCTC- TGCCCTC
2450 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2326 gtccctgggtcagccagaggcttgaactgttacagaagccctctgccctc
2375 V.1 2451 TGGTGGCCTCTGGGCCTGCTGCATGTACATATTTTCTGTAAAT- ATACATG
2500 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2376 tggtggcctctgggcctgctgcatgtacatattttctgtaaatatacatg
2425 V.1 2501 CGCCGGGAGCTTCTTGCAGGAATACTGCTCCGAATCACTTTTA- ATTTTTT
2550 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2426 cgccgggagcttcttgcaggaatactgctccgaatcacttttaatttttt
2475 V.1 2551 TCTTTTTTTTTTCTTGCCCTTTCCATTAGTTGTATTTTTTATT- TATTTTT
2600 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2476 tcttttttttttcttgccctttccattagttgtattttttatttattttt
2525 V.1 2601 ATTTTTATTTTTTTTTAGAGATGGAGTCTCACTATGTTGCTCA- GGCTGGC
2650 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2526 atttttatttttttttagagatggagtctcactatgttgctcaggctggc
2575 V.1 2651 CTTGAACTCCTGGGCTCAAGCAATCCTCCTGCCTCAGCCTCCC- TAGTAGC
2700 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2576 cttgaactcctgggctcaagcaatcctcctgcctcagcctccctagtagc
2625 V.1 2701 TGGGACTTTAAGTGTACACCACTGTGCCTGCTTTGAATCCTTT- ACGAACA
2750 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2626 tgggactttaagtgtacaccactgtgcctgctttgaatcctttacgaaga
2675 V.1 2751 GAAAAAAAAAATTAAAGAAAGCCTTTAGATTTATCCAATGTTT- ACTACTG
2800 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2676 gaaaaaaaaaattaaagaaagcctttagatttatccaatgtttactactg
2725 V.1 2801 GGATTGCTTAAAGTGAGGCCCCTCCAACACCAGGGGGTTAATT- CCTGTGA
2850 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2726 ggattgcttaaagtgaggcccctccaacaccagggggttaattcctgtga
2775 V.1 2851 TTGTGAAAGGGGCTACTTCCAAGGCATCTTCATGCAGGCAGCC-
CCTTGGG 2900 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2776 ttgtgaaaggggctacttccaaggcatcttcatgcaggcagccccttggg
2825 V.1 2901 AGGGCACCTGAGAGCTGGTAGACTCTGAAATTAGGGATGTGAG- CCTCGTG
2950 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2826 agggcacctgagagctggtagagtctgaaattagggatgtgagcctcgtg
2875 V.1 2951 GTTACTGAGTAAGGTAAAATTGCATCCACCATTGTTTGTGATA- CCTTAGG
3000 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2876 gttactgagtaaggtaaaattgcatccaccattgtttgtgataccttagg
2925 V.1 3001 GAATTGCTTGGACCTGGTGACAAGGGcTCCTGTTCAATAGTGG- TGTTGGG
3050 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2926 gaattgcttggacctggtgacaagggctcctgttcaatagtggtgttggg
2975 V.1 3051 GAGAGAGAGAGCAGTGATTATAGACCGAGAGAGTAGGAGTTGA- GGTGAGG
3100 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 2976 gagagagagagcagtgattatagaccgagagagtaggagttgaggtgagg
3025 V.1 3101 TGAAGGAGGTGCTGGGGGTGAGAATGTCGCCTTTCCCCCTGGG- TTTTGGA
3150 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 3026 tgaaggaggtgctgggggtgagaatgtcgcctttccccctgggttttgga
3075 V.1 3151 TCACTAATTCAAGCCTCTTCTGGATGTTTCTCTGGGTTGGGGC- TGGAGTT
3200 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 3076 tcactaattcaaggctcttctggatgtttctctgggttggggctggagtt
3125 V.1 3201 CAATGAGGTTTATTTTTAGCTGGCCCACCCAGATACACTCAGC- CAGAATA
3250 .vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. V.7 3126
caatgaggtttatttttagctggcccacccagatacactcagccagaata317- 5 V.1 3251
CCTAGATTTAGTACCCAAACTCTTCTTAGTCTGAAATCTGCTGGATT- TCT 3300
.vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.7 3176 cctagatttagtacccaaactcttcttagtctgaaatctgctggatttct 3225
V.1 3301 GGCCTAAGGGAGAGGCTCCCATCCTTCGTTCCCCAGCCAGCCTAGGACTT 3350
.vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.7 3226 ggcctaagggagaggctcccatccttcgttccccagccagcctaggactt 3275
V.1 3351 CGAATGTGGAGCCTGAAGATCTAAGATCCTAACATGTACATTTTATGTAA 3400
.vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.7 3276 cgaatgtggagcctgaagatctaagatcctaacatgtacattttatgtaa 3325
V.1 3401 ATATGTGCATATTTGTACATAAAATCATATTCTGTTTTTAAATAAACAGA 3450
.vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.7 3326 atatgtgcatatttgtacataaaatgatattctgtttttaaataaacaga 3375
V.1 3451 CAAAACTTGaaaaa 3464 .vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline. V.7 3376 caaaacttgaaaaa
3389
[1073]
69TABLE LIV(b) Peptide sequences of protein coded by 191P4D12(b)
v.7 (SEQ ID NO: 114) MPLSLGAEMW GPEAWLLLLL LLASFTGRCP AGELETSDVV
TVVLGQDAKL PCFYRGDSGE 60 QVGQVAWARV DAGEGAQELA LLHSKYGLHV
SPAYEGRVEQ PPPPRNPLDG SVLLRNAVQA 120 DEGEYECRVS TFPAGSFQAR
LRLRVLVPPL PSLNPGPALE EGQGLTLAAS CTAEGSPAPS 180 VTWDTEVKGT
TSSRSFKHSR SAAVTSEFHL VPSRSMNGQP LTCVVSHPGL LQDQRITHIL 240
HVSFLAEASV RGLEDQNLWH IGREGAMLKC LSEGQPPPSY NWTRLDGPLP SGVRVDGDTL
300 GFPPLTTEHS GIYVCHVSNE FSSRDSQVTV DVLDPQEDSG KQVDLVSASV
VVVGVIAALL 360 FCLLVVVVVL MSRYHRRKAQ QMTQKYEEEL TLTRENSIRR
LHSHHTDPRS QSEEPEGRSY 420 STLTTVREIE TQTELLSPGS GRAEEEEDQD
EGIKQAMNHF VQENGTLRAK PTGNGIYING 480 RGHLV 485
[1074]
70TABLE LV(b) Amino acid sequence alignment of 191P4D12(b) v.1 (SEQ
ID NO: 115) and 191P4D12(b) v.7 (SEQ ID NO: 116). V.1 1
MPLSLGAEMWGPEAWLLLLLLLASFTGRCPAGELETSD- VVTVVLGQDAKL 50
.vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline. V.7 1 MPLSLGAEMWGPEAWLLLLLLLASFTGRCPAGELETSDVVTVVLGQDAKL
50 V.1 51 PCFYRGDSGEQVGQVAWARVDAGEGAQELALLHSKYGLHVSPAYEGR- VEQ 100
.vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.7 51 PCFYRGDSGEQVGQVAWARVDAGEGAQELALLHSKYGLHVSPAYEGRVEQ 100 V.1
101 PPPPRNPLDGSVLLRNAVQADEGEYECRVSTFPAGSFQARLRLRVLVPPL 150
.vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline.
V.7 101 PPPPRNPLDGSVLLRNAVQADEGEYECRVSTFPAGSFQARLRLRVLVPPL 150 V.1
151 PSLNPGPALEEGQGLTLAASCTAEGSPAPSVTWDTEVKGTTSSRSFKHSR 200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline. V.7
151 PSLNPGPALEEGQGLTLAASCTAEGSPAPSVTWDTEVKGTTSSRSFKHSR 200 V.1 201
SAAVTSEFHLVPSRSMNGQPLTCVVSHPGLLQDQRITHILHVSFLAEASV 250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 201
SAAVTSEFHLVPSRSMNGQPLTCVVSHPGLLQDQRITHILHVSFLAEASV 250 V.1 251
RGLEDQNLWHIGREGAMLKCLSEGQPPPSYNWTRLDGPLPSGVRVDGDTL 300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 251
RGLEDQNLWHIGREGAMLKCLSEGQPPPSYNWTRLDGPLPSGVRVDGDTL 300 V.1 301
GFPPLTTEHSGIYVCHVSNEFSSRDSQVTVDVLDPQEDSGKQVDLVSASV 350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 301
GFPPLTTEHSGIYVCHVSNEFSSRDSQVTVDVLDPQEDSGKQVDLVSASV 350 V.1 351
VVVGVIAALLFCLLVVVVVLMSRYHRRKAQQMTQKYEEELTLTRENSIRR 400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.7 351
VVVGVIAALLFCLLVVVVVLMSRYHRRKAQQMTQKYEEELTLTRENSIRR 400 V.1 401
LHSHHTDPRSQPEESVGLRAEGHPDSLKDNSSCSVMSEEPEGRSYSTLTT 450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline.
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline.
V.7 401 LHSHHTDPRSQ-------------------------SEEPEGRSYSTLTT 425 V.1
451 VREIETQTELLSPGSGRAEEEEDQDEGIKQAMNHFVQENGTLRAKPTGNG 500
.vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline. V.7
426 VREIETQTELLSPGSGRAEEEEDQDEGIKQANNHFVQENGTLRAKPTGNG 475 V.1 501
IYINGRGHLV 510 .vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline. V.7
476 IYINGRGHLV 485
[1075]
71TABLE LII(c) Nucleotide sequence of transcript variant
191P4D12(b) v.8 (SEQ ID NO:117) ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac
240 gctgggcagt ctgcctttca accatgcccc tgtccctggg agccgagatg
tgggggcctg 300 aggcctggct gctgctgctg ctactgctgg catcatttac
aggccggtgc cccgcgggtg 360 agctggagac ctcagacgtg gtaactgtgg
tgctgggcca ggacgcaaaa ctgccctgct 420 tctaccgagg ggactccggc
gagcaagtgg ggcaagtggc atgggctcgg gtggacgcgg 480 gcgaaggcgc
ccaggaacta gcgctactgc actccaaata cgggcttcat gtgagcccgg 540
cttacgaggg ccgcgtggag cagccgccgc ccccacgcaa ccccctggac ggctcagtgc
600 tcctgcgcaa cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc
agcaccttcc 660 ccgccggcag cttccaggcg cggctgcggc tccgagtgct
ggtgcctccc ctgccctcac 720 tgaatcctgg tccagcacta gaagagggcc
agggcctgac cctggcagcc tcctgcacag 780 ctgagggcag cccagccccc
agcgtgacct gggacacgga ggtcaaaggc acaacgtcca 840 gccgttcctt
caagcactcc cgctctgctg ccgtcacctc agagttccac ttggtgccta 900
gccgcagcat gaatgggcag ccactgactt gtgtggtgtc ccatcctggc ctgctccagg
960 accaaaggat cacccacatc ctccacgtgt ccttccttgc tgaggcctct
gtgaggggcc 1020 ttgaagacca aaatctgtgg cacattggca gagaaggagc
tatgctcaag tgcctgagtg 1080 aagggcagcc ccctccctca tacaactgga
cacggctgga tgggcctctg cccagtgggg 1140 tacgagtgga tggggacact
ttgggctttc ccccactgac cactgagcac agcggcatct 1200 acgtctgcca
tgtcagcaat gagttctcct caagggattc tcaggtcact gtggatgttc 1260
ttgaccccca ggaagactct gggaagcagg tggacctagt gtcagcctcg gtggtggtgg
1320 tgggtgtgat cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg
ctcatgtccc 1380 gataccatcg gcgcaaggcc cagcagatga cccagaaata
tgaggaggag ctgaccctga 1440 ccagggagaa ctccatccgg aggctgcatt
cccatcacac ggaccccagg agccagccgg 1500 aggagagtgt agggctgaga
gccgagggcc accctgatag tctcaaggac aacagtagct 1560 gctctgtgat
gagtgaagag cccgagggcc gcagttactc cacgctgacc acggtgaggg 1620
agatagaaac acagactgaa ctgctgtctc caggctctgg gcgggccgag gaggaggaag
1680 atcaggatga aggcatcaaa caggccatga accattttgt tcaggagaat
gggaccctac 1740 gggccaagcc cacgggcaat ggcatctaca tcaatgggcg
gggacacctg gtctgaccca 1800 ggcctgcctc ccttccctag gcctggctcc
ttctgttgac atgggagatt ttagctcatc 1860 ttgggggcct ccttaaacac
ccccatttct tgcggaagat gctccccatc ccactgactg 1920 cttgaccttt
acctccaacc cttctgttca tcgggagggc tccaccaatt gagtctctcc 1980
caccatgcat gcaggtcact gtgtgtgtgc atgtgtgcct gtgtgagtgt tgactgactg
2040 tgtgtgtgtg gaggggtgac tgtccgtgga ggggtgactg tgtccgtggt
gtgtattatg 2100 ctgtcatatc agagtcaagt gaactgtggt gtatgtgcca
cgggatttga gtggttgcgt 2160 gggcaacact gtcagggttt ggcgtgtgtg
tcatgtggct gtgtgtgacc tctgcctgaa 2220 aaagcaggta ttttctcaga
ccccagagca gtattaatga tgcagaggtt ggaggagaga 2280 ggtggagact
gtggctcaga cccaggtgtg cgggcatagc tggagctgga atctgcctcc 2340
ggtgtgaggg aacctgtctc ctaccacttc ggagccatgg gggcaagtgt gaagcagcca
2400 gtccctgggt cagccagagg cttgaactgt tacagaagcc ctctgccctc
tggtggcctc 2460 tgggcctgct gcatgtacat attttctgta aatatacatg
cgccgggagc ttcttgcagg 2520 aatactgctc cgaatcactt ttaatttttt
tctttttttt ttcttgccct ttccattagt 2580 tgtatttttt atttattttt
atttttattt ttttttagag atggagtctc actatgttgc 2640 tcaggctggc
cttgaactcc tgggctcaag caatcctcct gcctcagcct ccctagtagc 2700
tgggacttta agtgtacacc actgtgcctg ctttgaatcc tttacgaaga gaaaaaaaaa
2760 attaaagaaa gcctttagat ttatccaatg tttactactg ggattgctta
aagtgaggcc 2820 cctccaacac cagggggtta attcctgtga ttgtgaaagg
ggctacttcc aaggcatctt 2880 catgcaggca gccccttggg agggcacctg
agagctggta gagtctgaaa ttagggatgt 2940 gagcctcgtg ctggtgacaa
gggctcctgt tcaatagtgg tgttggggag agagagagca 3000 gtgattatag
accgagagag taggagttga ggtgaggtga aggaggtgct gggggtgaga 3060
atgtcgcctt tccccctggg ttttggatca ctaattcaag gctcttctgg atgtttctct
3120 gggttggggc tggagttcaa tgaggtttat ttttagctgg cccacccaga
tacactcagc 3180 cagaatacct agatttagta cccaaactct tcttagtctg
aaatctgctg gatttctggc 3240 ctaagggaga ggctcccatc cttcgttccc
cagccagcct aggacttcga atgtggagcc 3300 tgaagatcta agatcctaac
atgtacattt tatgtaaata tgtgcatatt tgtacataaa 3360 atgatattct
gtttttaaat aaacagacaa aacttgaaaa a 3401
[1076]
72TABLE LIII(c) Nucleotide seguence alignment of 191P4D12(b) v.1
(SEQ ID NO: 118) and 191P4D12(b) v.8 (SEQ ID NO: 119) V.1 1
gGCCGTCGTTGTTGGCCACAGCGTGGGAAGCAGCTCTGG- GGGAGCTCGGA 50
.vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline. V.8 1 ggccgtcgttgttggccacagcgtgggaagcagctctgggggagctcgga 50
V.1 51 GCTCCCGATCACGGCTTCTTGGGGGTAGCTACGGCTGGGTGTGTAGAACG 100
.vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.8 51 gctcccgatcacggcttcttgggggtagctacggctgggtgtgtagaacg 100 V.1
101 GGGCCGGGGCTGGGGCTGGGTCCCCTAGTGGAGACCCAAGTGCGAGAGGC 150
.vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline. V.8
101 gggccggggctggggctgggtcccctagtggagacccaagtgcgagaggc 150 V.1 151
AAGAACTCTGCAGCTTCCTGCCTTCTGGGTCAGTTCCTTATTCAAGTCTG 200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 151
aagaactctgcagcttcctgccttctgggtcagttccttattcaagtctg 200 V.1 201
CAGCCGGCTCCCAGGGAGATCTCGGTGGAACTTCAGAAACGCTGGGCAGT 250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 201
cagccggctcccagggagatctcggtggaacttcagaaacgctgggcagt 250 V.1 251
CTGCCTTTCAACCATGCCCCTGTCCCTGGGAGCCGAGATGTGGGGGCCTG 300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 251
ctgcctttcaaccatgcccctgtccctgggagccgagatgtgggggcctg 300 V.1 301
AGGCCTGGCTGCTGCTGCTGCTACTGCTGGCATCATTTACAGGCCGGTGC 350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 301
aggcctggctgctgctgctgctactgctggcatcatttacaggccggtgc 350 V.1 351
CCCGCGGGTGAGCTGGAGACCTCAGACGTGGTAACTGTGGTGCTGGGCCA 400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 351
cccgcgggtgagctggagacctcagacgtggtaactgtggtgctgggcca 400 V.1 401
GGACGCAAAACTGCCCTGCTTCTACCGAGGGGACTCCGGCGAGCAAGTGG 450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 401
ggacgcaaaactgccctgcttctaccgaggggactccggcgagcaagtgg 450 V.1 451
GGCAAGTGGCATGGGCTCGGGTGGACGCGGGCGAAGGCGCCCAGGAACTA 500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 451
ggcaagtggcatgggctcgggtggacgcgggcgaaggcgcccaggaacta 500 V.1 501
GCGCTACTGCACTCCAAATACGGGCTTCATGTGAGCCCGGCTTACGAGGG 550
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 501
gcgctactgcactccaaatacggccttcatgtgagcccggcttacgaggg 550 V.1 551
CCGCGTGGAGCAGCCGCCGCCCCCACGCAACCCCCTGGACGGCTCAGTGC 600
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 551
ccgcgtggagcagccgccgcccccacgcaaccccctggacggctcagtgc 600 V.1 601
TCCTGCGCAACGCAGTGCAGGCGGATGAGGGCGAGTACGAGTGCCGGGTC 650
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 601
tcctgcgcaacgcagtgcaggcggatgagggcgagtacgagtgccgggtc 650 V.1 651
AGCACCTTCCCCGCCGGCAGCTTCCAGGCGCGGCTGCGGCTCCGAGTGCT 700
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 651
agcaccttccccgccggcagcttccaggcgcggctgcggctccgagtgct 700 V.1 701
GGTGCCTCCCCTGCCCTCACTGAATCCTGGTCCAGCACTAGAAGAGGGCC 750
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 701
ggtgcctcccctgccctcactgaatcctggtccagcactagaagagggcc 750 V.1 751
AGGGCCTGACCCTGGCAGCCTCCTGCACAGCTGAGGGCAGCCCAGCCCCC 800
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 751
agggcctgaccctggcagcctcctgcacagctgagggcagcccagccccc 800 V.1 801
AGCGTGACCTGGGACACGGAGGTCAAAGGCACAACGTCCAGCCGTTCCTT 850
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 801
agcgtgacctgggacacggaggtcaaaggcacaacgtccagccgttcctt 850 V.1 851
CAAGCACTCCCGCTCTGCTGCCGTCACCTCAGAGTTCCACTTCGTGCCTA 900
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 851
caagcactcccgctctgctgccgtcacctcagagttccacttggtgccta 900 V.1 901
GCCGCACCATGAATGGGCAGCCACTGACTTGTGTGGTGTCCCATCCTGGC 950
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 901
gccgcagcatgaatgggcagccactgacttgtgtggtgtcccatcctggc 950 V.1 951
CTGCTCCAGGACCAAACGATCACCCACATCCTCCACGTGTCCTTCCTTGC 1000
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 951
ctgctccaggaccaaaggatcacccacatcctccacgtgtccttccttgc 1000 V.1 1001
TGAGGCCTCTGTGAGGGGCCTTGAAGACCAAAATCTGTGGCACATTGGCA 1050
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1001
tgaggcctctgtgaggggccttgaagaccaaaatctgtggcacattggca 1050 V.1 1051
GAGAAGGAGCTATGCTCAAGTGCCTGAGTGAAGGGCAGCCCCCTCCCTCA 1100
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1051
gagaaggagctatgctcaagtgcctgagtgaagggcagccccctccctca 1100 V.1 1101
TACAACTGGACACGGCTGGATGCCCCTCTGCCCAGTGGGGTACGAGTGGA 1150
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1101
tacaactggacacggctggatgggcctctgcccagtggggtacgagtgga 1150 V.1 1151
TGGGGACACTTTGGGCTTTCCCCCACTGACCACTGAGCACAGCGGCATCT 1200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1151
tggggacactttgggctttcccccactgaccactgagcacagcggcatct 1200 V.1 1201
ACGTCTGCCATGTCAGCAATGAGTTCTCCTCAAGGGATTCTCAGGTCACT 1250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1201
acgtctgccatgtcagcaatgagttctcctcaagggattctcaggtcact 1250 V.1 1251
GTGGATGTTCTTGACCCCCAGGAAGACTCTGGGAAGCAGGTGGACCTAGT 1300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1251
gtggatgttcttgacccccaggaagactctgggaagcaggtggacctagt 1300 V.1 1301
GTCAGCCTCGGTGGTGGTGGTGGGTGTGATCGCCGCACTCTTGTTCTGCC 1350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1301
gtcagcctcggtggtggtggtgggtgtgatcgccgcactcttgttctgcc 1350 V.1 1351
TTCTGGTGGTGGTCGTGGTGCTCATGTCCCGATACCATCGGCGCAAGGCC 1400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1351
ttctggtggtggtggtggtgctcatgtcccgataccatcggcgcaaggcc 1400 V.1 1401
CAGCAGATGACCCAGAAATATGAGGAGGAGCTGACCCTGACCAGGGAGAA 1450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1401
cagcagatgacccagaaatatgaggaggagctgaccctgaccagggagaa 1450 V.1 1451
CTCCATCCGGAGGCTGCATTCCCATCACACGGACCCCAGGAGCCAGCCGG 1500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1451
ctccatccggaggctgcattcccatcacacggaccccaggagccagccgg 1500 V.1 1501
AGGAGAGTGTAGGGCTGAGAGCCCAGGGCCACCCTGATAGTCTCAAGGAC 1550
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1501
aggagagtgtagggctgagagccgagggccaccctgatagtctcaaggac 1550 V.1 1551
AACAGTAGCTGCTCTGTGATGAGTGAAGAGCCCGAGGGCCGCAGTTACTC 1600
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1551
aacagtagctgctctgtgatgagtgaagagcccgagggccgcagttactc 1600 V.1 1601
CACGCTGACCACGGTGAGGGAGATAGAAACACAGACTGAACTGCTGTCTC 1650
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1601
cacgctgaccacggtgagggagatagaaacacagactgaactgctgtctc 1650 V.1 1651
CAGGCTCTGGGCGGGCCGAGGAGGAGGAAGATCAGGATGAAGGCATCAAA 1700
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1651
caggctctgggcgggccgaggaggaggaagatcaggatgaaggcatcaaa 1700 V.1 1701
CAGGCCATGAACCATTTTGTTCAGGAGAATGGGACCCTACGGGCCAAGCC 1750
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1701
caggccatgaaccattttgttcaggagaatgggaccctacgggccaagcc 1750 V.1 1751
CACGGGCAATGGCATCTACATCAATGGGCGGGGACACCTGGTCTGACCCA 1800
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1751
cacgggcaatggcatctacatcaatgggcggggacacctggtctgaccca 1800 V.1 1801
GGCCTGCCTCCCTTCCCTAGGCCTGGCTCCTTCTGTTGACATGGGAGATT 1850
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1801
ggcctgcctcccttccctaggcctggctccttctgttgacatgggagatt 1850 V.1 1851
TTAGCTCATCTTGGGGGCCTCCTTAAACACCCCCATTTCTTGCGGAAGAT 1900
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1851
ttagctcatcttgggggcctccttaaacacccccatttcttgcggaagat 1900 V.1 1901
GCTCCCCATCCCACTGACTGCTTGACCTTTACCTCCAACCCTTCTGTTCA 1950
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1901
gctccccatcccactgactgcttgacctttacctccaacccttctgttca 1950 V.1 1951
TCGGGAGGGCTCCACCAATTGAGTCTCTCCCACCATGCATGCACGTCACT 2000
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 1951
tcgggagggctccaccaattgagtctctcccaccatgcatgcaggtcact 2000 V.1 2001
GTGTGTGTGCATGTGTGCCTGTGTGAGTGTTGACTGACTGTGTGTGTGTG 2050
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2001
gtgtgtgtgcatgtgtgcctgtgtgagtgttgactgactgtgtgtgtgtg 2050 V.1 2051
GAGGGGTGACTGTCCGTGGAGGGGTGACTGTGTCCGTGGTGTGTATTATG 2100
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2051
gaggggtgactgtccgtggaggggtgactgtgtccgtggtgtgtattatg 2100 V.1 2101
CTGTCATATCAGAGTCAAGTGAACTGTGGTGTATGTGCCACGGGATTTGA 2150
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2101
ctgtcatatcagagtcaagtgaactgtggtgtatgtgccacgggatttga 2150 V.1 2151
GTGGTTGCGTGGGCAACACTGTCAGGGTTTGGCGTGTGTGTCATGTGGCT 2200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2151
gtggttgcgtgggcaacactgtcagggtttggcgtgtgtgtcatgtggct 2200 V.1 2201
GTGTGTGACCTCTGCCTCACAAAGCAGGTATTTTCTCAGACCCCAGAGCA 2250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2201
gtgtgtgacctctgcctgaaaaagcaggtattttctcagaccccagagca 2250 V.1 2251
GTATTAATGATGCAGAGGTTGGAGGAGAGAGGTGGAGACTGTGGCTCAGA 2300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2251
gtattaatgatgcagaggttggaggagagaggtggagactgtggctcaga 2300 V.1 2301
CCCAGGTGTGCGGGCATACCTGGAGCTGGAATCTGCCTCCGGTGTGAGGG 2350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2301
cccaggtgtgcgggcatagctggagctggaatctgcctccggtgtgaggg 2350 V.1 2351
AACCTGTCTCCTACCACTTCGGAGCCATGGGGGCAAGTGTGAAGCAGCCA 2400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2351
aacctgtctcctiaccacttcggagccatgggggcaagtgtgaagcagcca 2400 V.1 2401
GTCCCTGGGTCAGCCAGAGGCTTGAACTGTTACAGGAAGCCCTCTGCCCTC 2450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline. V.8 2401
gtccctgggtcagccagaggcttgaactgttacagaagccctctgccctc 2450 V.1 2451
TGGTGGCCTCTGCGCCTGCTGCATGTACATATTTTCTGTAAATATACATG 2500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2451
tggtggcctctgggcctgctgcatgtacatattttctgtaaatatacatg 2500 V.1 2501
CGCCGGGAGCTTCTTGCAGGAATACTGCTCCGAATCACTTTTAATTTTTT 2550
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2501
cgccgggagcttcttgcaggaatactgctccgaatcacttttaatttttt 2550 V.1 2551
TCTTTTTTTTTTCTTGCCCTTTCCATTAGTTGTATTTTTTATTTATTTTT 2600
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2551
tcttttttttttcttgccctttccattagttgtattttttatttattttt 2600 V.1 2601
ATTTTTATTTTTTTTTAGAGATGGAGTCTCACTATGTTGCTCAGGCTGGC 2650
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2601
atttttatttttttttagagatggagtctcactatgttgctcaggctggc 2650 V.1 2651
CTTGAACTCCTGGGCTCAAGCAATCCTCCTGCCTCAGCCTCCCTAGTAGC 2700
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2651
cttgaactcctgggctcaagcaatcctcctgcctcagcctccctagtagc 2700 V.1 2701
TGGGACTTTAAGTGTACACCACTGTGCCTGCTTTGAATCCTTTACGAAGA 2750
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2701
tgggactttaagtgtacaccactgtgcctgctttgaatcctttacgaaga 2750 V.1 2751
GAAAAAAAAAATTAAAGAAAGCCTTTAGATTTATCCAATGTTTACTACTG 2800
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2751
gaaaaaaaaaattaaagaaagcctttagatttatccaatgtttactactg 2800 V.1 2801
GGATTGCTTAAAGTGAGGCCCCTCCAACACCAGGGGGTTAATTCCTGTGA 2850
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2801
ggattgcttaaagtgaggcccctccaacaccagggggttaattcctgtga 2850 V.1 2851
TTGTGAAAGGGGCTACTTCCAAGGCATCTTCATGCAGGCAGCCCCTTGGG 2900
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2851
ttgtgaaaggggctacttccaaggcatcttcatgcaggcagccccttggg 2900 V.1 2901
AGGGCACCTGAGAGCTGGTAGAGTCTGAAATTAGGGATGTGAGCCTCGTG 2950
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2901
agggcacctgagagctggtagagtctgaaattagggatgtgagcctcgtg 2950 V.1 2951
GTTACTGAGTAACGTAAAATTGCATCCACCATTGTTTGTGATACCTTAGG 3000 V.8 2951
-------------------------------------------------- 2950 V.1 3001
GAATTGCTTGGACCTGGTGACAAGGGCTCCTGTTCAATAGTGGTGTTGGG 3050
.vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline. V.8 2951
-------------ctggtgacaagggctc- ctgttcaatagtggtgttggg 2987 V.1 3051
GAGAGAGAGAGCAGTGATTATAGACCGAGAGAGTAGGAGTTGAGGTGAGG 3100
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 2988
gagagagagagcagtgattatagaccgagagagtaggagttgaggtgagg 3037 V.1 3101
TGAAGCAGGTGCTGGGGGTGAGAATGTCGCCTTTCCCCCTGGGTTTTGGA 3150
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 3038
tgaaggaggtgctgggggtgagaatgtcgcctttccccctgggttttgga 3087 V.1 3151
TCACTAATTCAAGGCTCTTCTGGATGTTTCTCTGGGTTGGGGCTGGAGTT 3200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 3088
tcactaattcaaggctcttctggatgtttctctgggttggygctggagtt 3137 V.1 3201
CAATGAGGTTTATTTTTAGCTGGCCCACCCAGATACACTCAGCCAGAATA 3250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 3138
caatgaggtttatttttagctggcccacccagatacactcagccagaata 3187 V.1 3251
CCTAGATTTAGTACCCAAACTCTTCTTAGTCTGAAATCTGCTGGATTTCT 3300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 3188
cctagatttagtacccaaactcttcttagtctgaaatctgctggatttct 3237 V.1 3301
GGCCTAAGGGAGAGGCTCCCATCCTTCGTTCCCCAGCCAGCCTAGGACTT 3350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 3238
ggcctaagggagaggctcccatccttcgttccccagccagcctaggactt 3287 V.1 3351
CGAATGTGGAGCCTGAAGATCTAAGATCCTAACATGTACATTTTATGTAA 3400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 3288
cgaatgtggagcctgaagatctaagatcctaacatgtacattttatgtaa 3337 V.1 3401
ATATGTGCATATTTGTACATAAAATGATATTCTGTTTTTAAATAAACAGA 3450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 3338
atatgtgcatatttgtacataaaatgatattctgtttttaaataaacaga 3387 V.1 3451
CAAAACTTGaaaaa 3464 .vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline. V.8 3388 caaaacttgaaaaa 3401
[1077]
73TABLE LIV(c) Peptide seguences of protein coded by 191P4D12(b)
v.8 (SEQ ID NO:120) MPLSLGAEMW GPEAWLLLLL LLASFTGRCP AGELETSDVV
TVVLGQDAKL PCFYRGDSGE 60 QVGQVAWARV DAGEGAQELA LLHSKYGLHV
SPAYEGRVEQ PPPPRNPLDG SVLLRNAVQA 120 DEGEYECRVS TFPAGSFQAR
LRLRVLVPPL PSLNPGPALE EGQGLTLAAS CTAEGSPAPS 180 VTWDTEVKGT
TSSRSFKHSR SAAVTSEFHL VPSRSMNGQP LTCVVSHPGL LQDQRITHIL 240
HVSFLAEASV RGLEDQNLWH IGREGAMLKC LSEGQPPPSY NWTRLDGPLP SGVRVDGDTL
300 GFPPLTTEHS GIYVCHVSNE FSSRDSQVTV DVLDPQEDSG KQVDLVSASV
VVVGVIAALL 360 FCLLVVVVVL MSRYHRRKAQ QMTQKYEEEL TLTRENSIRR
LHSHHTDPRS QPEESVGLRA 420 EGHPDSLKDN SSCSVMSEEP EGRSYSTLTT
VREIETQTEL LSPGSGRAEE EEDQDEGIKQ 480 AMNHFVQENG TLRAKPTGNG
IYINGRGHLV 510
[1078]
74TABLE LV(c) Amino acid seguence alignment of 191P4012(b) v.1 (SEQ
ID NO: 121) and 191P4D12(b) v.8 (SEQ ID NO: 122) V.1 1
MPLSLGAEMWGPEAWLLLLLLLASFTGRCPAGELETSDV- VTVVLGQDAKL 50
.vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline. V.8 1 MPLSLGAEMWGPEAWLLLLLLLASFTGRCPAGELETSDVVTVVLGQDAKL 50
V.1 51 PCFYRGDSGEQVGQVAWARVDAGEGAQELALLHSKYGLHVSPAYEGRVEQ 100
.vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline.
V.8 51 PCFYRGDSGEQVGQVAWARVDAGEGAQELALLHSKYGLHVSPAYEGRVEQ 100 V.1
101 PPPPRNPLDGSVLLRNAVQADEGEYECRVSTFPAGSFQARLRLRVLVPPL 150
.vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline. V.8
101 PPPPRNPLDGSVLLRNAVQADEGEYECRVSTFPAGSFQARLRLRVLVPPL 150 V.1 151
PSLNPGPALEEGQGLTLAASCTAEGSPAPSVTWDTEVKGTTSSRSFKHSR 200
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 151
PSLNPGPALEEGQGLTLAASCTAEGSPAPSVTWDTEVKGTTSSRSFKHSR 200 V.1 201
SAAVTSEFHLVPSRSMNGQPLTCVVSHPGLLQDQRITHILHVSFLAEASV 250
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 201
SAAVTSEFHLVPSRSMNGQPLTCVVSHPGLLQDQRITHILHVSFLAEASV 250 V.1 251
RGLEDQNLWHIGREGAMLKCLSEGQPPPSYNWTRLDGPLPSGVRVDGDTL 300
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 251
RGLEDQNLWHIGREGAMLKCLSEGQPPPSYNWTRLDGPLPSGVRVDGDTL 300 V.1 301
GFPPLTTEHSGIYVCHVSNEFSSRDSQVTVDVLDPQEDSGKQVDLVSASV 350
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 301
GFPPLTTEHSGIYVCHVSNEFSSRDSQVTVDVLDPQEDSGKQVDLVSASV 350 V.1 351
VVVGVIAALLFCLLVVVVVLMSRYHRRKAQQMTQKYEEELTLTRENSIRR 400
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 351
VVVGVIAALLFCLLVVVVVLMSRYHRRKAQQMTQKYEEELTLTRENSIRR 400 V.1 401
LHSHHTDPRSQPEESVGLRAEGHPDSLKDNSSCSVMSEEPEGRSYSTLTT 450
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 401
LHSHHTDPRSQPEESVGLRAEGHPDSLKDNSSCSVMSEEPEGRSYSTLTT 450 V.1 451
VREIETQTELLSPGSGRAEEEEDQDEGIKQAMNHFVQENGTLRAKPTGNG 500
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. V.8 451
VREIETQTELLSPGSGRAEEEEDQDEGIKQAMNHFVQENGTLRAKPTGNG 500 V.1 501
IYINGRGHLV 510 .vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline. V.8 501
IYINGRGHLV 510
[1079]
75TABLE LII(d) Nucleotide seguence of transcript variant
191P4012(b) v.9 (SEQ ID NO: 123) gtctgaccca ggcctgcctc ccttccctag
gcctggctcc ttctgttgac atgggagatt 60 ttagctcatc ttgggggcct
ccttaaacac ccccatttct tgcggaagat gctccccatc 120 ccactgactg
cttgaccttt acctccaacc cttctgttca tcgggagggc tccaccaatt 180
gagtctctcc caccatgcat gcaggtcact gtgtgtgtgc atgtgtgcct gtgtgagtgt
240 tgactgactg tgtgtgtgtg gaggggtgac tgtccgtgga ggggtgactg
tgtccgtggt 300 gtgtattatg ctgtcatatc agagtcaagt gaactgtggt
gtatgtgcca cgggatttga 360 gtggttgcgt gggcaacact gtcagggttt
ggcgtgtgtg tcatgtggct gtgtgtgacc 420 tctgcctgaa aaagcaggta
ttttctcaga ccccagagca gtattaatga tgcagaggtt 480 ggaggagaga
ggtggagact gtggctcaga cccaggtgtg cgggcatagc tggagctgga 540
atctgcctcc ggtgtgaggg aacctgtctc ctaccacttc ggagccatgg gggcaagtgt
600 gaagcagcca gtccctgggt cagccagagg cttgaactgt tacagaagcc
ctctgccctc 660 tggtggcctc tgggcctgct gcatgtacat attttctgta
aatatacatg cgccgggagc 720 ttcttgcagg aatactgctc cgaatcactt
ttaatttttt tctttttttt ttcttgccct 780 ttccattagt tgtatttttt
atttattttt atttttattt ttttttagag atggagtctc 840 actatgttgc
tcaggctggc cttgaactcc tgggctcaag caatcctcct gcctcagcct 900
ccctagtagc tgggacttta agtgtacacc actgtgcctg ctttgaatcc tttacgaaga
960 gaaaaaaaaa attaaagaaa gcctttagat ttatccaatg tttactactg
ggattgctta 1020 aagtgaggcc cctccaacac cagggggtta attcctgtga
ttgtgaaagg ggctacttcc 1080 aaggcatctt catgcaggca gccccttggg
agggcacctg agagctggta gagtctgaaa 1140 ttagggatgt gagcctcgtg
gttactgagt aaggtaaaat tgcatccacc attgtttgtg 1200 ataccttagg
gaattgcttg gacctggtga caagggctcc tgttcaatag tggtgttggg 1260
gagagagaga gcagtgatta tagaccgaga gagtaggagt tgaggtgagg tgaaggaggt
1320 gctgggggtg agaatgtcgc ctttccccct gggttttgga tcactaattc
aaggctcttc 1380 tggatgtttc tctgggttgg ggctggagtt caatgaggtt
tatttttagc tggcccaccc 1440 agatacactc agccagaata cctagattta
gtacccaaac tcttcttagt ctgaaatctg 1500 ctggatttct ggcctaaggg
agaggctccc atccttcgtt ccccagccag cctaggactt 1560 cgaatgtgga
gcctgaagat ctaagatcct aacatgtaca ttttatgtaa atatgtgcat 1620
atttgtacat aaaatgatat tctgttttta aataaacaga caaaacttg 1669
[1080]
76TABLE LIII(d) Nucleotide seguence alignment of 191P4D12(b) v.1
(SEQ ID NO: 124) and 191P4D12(b) v.9 (SEQ ID NO: 125) v.1 1791
GTCTGACCCAGGCCTGCCTCCCTTCCCTAGGCCTGG- CTCCTTCTGTTGAC 1840
.vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline. v.9 1
gtctgacccaggcctgcctcccttccctaggcctggctccttctgttg- ac 50 v.1 1841
ATGGGAGATTTTAGCTCATCTTGGGGGCCTCCTTAAACACCCC- CATTTCT 1890
.vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine. v.9 51 atgggagattttagctcatcttgggggcctccttaaacacccccatttct 100
v.1 1891 TGCGGAAGATGCTCCCCATCCCACTGACTGCTTGACCTTTACCTCCAA- CC 1940
.vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
v.9 101 tgcggaagatgctccccatcccactgactgcttgacctttacctccaacc 150 v.1
1941 CTTCTGTTCATCGGGAGGGCTCCACCAATTGAGTCTCTCCCACCATGCAT 1990
.vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline.
v.9 151 cttctgttcatcgggagggctccaccaattgagtctctcccaccatgcat 200 v.1
1991 GCAGCTCACTGTGTGTGTGCATGTGTGCCTGTGTGAGTGTTGACTGACTG 2040
.vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline.
v.9 201 gcaggtcactgtgtgtgtgcatgtgtgcctgtgtgagtgttgactgactg 250 v.1
2041 TGTGTGTGTGGAGGGGTGACTGTCCGTGGAGGGGTGACTGTGTCCGTGGT 2090
.vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline. v.9
251 tgtgtgtgtggaggggtgactgtccgtggaggggtgactgtgtccgtggt 300 v.1 2091
GTGTATTATGCTGTCATATCACAGTCAAGTGAACTGTGGTGTATGTGCCA 2140
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline. v.9 301
gtgtattatgctgtcatatcagagtcaagtgaactgtggtgtatgtgcca 350 v.1 2141
CGGGATTTGAGTGGTTGCGTGGGCAACACTGTCAGGGTTTGGCGTGTCTG 2190
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 351
cgggatttgagtggttgcgtgggcaacactgtcagggtttggcgtgtgtg 400 v.1 2191
TCATGTGGCTGTGTGTGACCTCTGCCTGAAAAAGCAGGTATTTTCTCAGA 2240
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 401
tcatgtggctgtgtgtgacctctgcctgaaaaagcaggtattttctcaga 450 v.1 2241
CCCCAGAGCAGTATTAATGATGCAGAGGTTGGAGGAGAGAGGTGGAGACT 2290
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 451
ccccagagcagtattaatgatgcagaggttggaggagagaggtggagact 500 v.1 2291
GTGGCTCAGACCCAGGTGTGCGGGCATAGCTGGAGCTGGAATCTGCCTCC 2340
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 501
gtggctcagacccaggtgtgcgggcatagctggagctggaatctgcctcc 550 v.1 2341
GGTCTGAGGGAACCTGTCTCCTACCACTTCGGAGCCATGGGGGCAAAGTGT 2390
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline..vertline..vertl-
ine..vertline..vertline..vertline..vertline..vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline..vertline..vertline..vertline..vertline..vertline..vertline..v-
ertline..vertline..vertline..vertline..vertline..vertline. v.9 551
ggtgtgagggaacctgtctcctaccacttcggagccatgggggcaagtgt 600 v.1 2391
GAAGCAGCCAGTCCCTGGGTCAGCCAGAGGCTTGAACTGTTACAGAAGCC 2440
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 601
gaagcagccagtccctgggtcagccagaggcttgaactgttacagaagcc 650 v.1 2441
CTCTGCCCTCTGGTGGCCTCTGCGCCTGCTGCATGTACATATTTTCTGTA 2490
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 651
ctctgccctctggtggcctctgggcctgctgcatgtacatattttctgta 700 v.1 2491
AATATACATCCGCCGGGAGCTTCTTGCAGGAATACTGCTCCGPATCACTT 2540
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 701
aatatacatgcgccgggagcttcttgcaggaatactgctccgaatcactt 750 v.1 2541
TTAATTTTTTTCTTTTTTTTTTCTTGCCCTTTCCATTAGTTGTATTTTTT 2590
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 751
ttaatttttttcttttttttttcttgccctttccattagttgtatttttt 800 v.1 2591
ATTTATTTTTATTTTTATTTTTTTTTAGAGATGGAGTCTCACTATGTTGC 2640
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 801
atttatttttatttttatttttttttagagatggagtctcactatgttgc 850 v.1 2641
TCAGGCTGGCCTTGAACTCCTGGCCTCAAGCAATCCTCCTGCCTCAGCCT 2690
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 851
tcaggctggccttgaactcctgggctcaagcaatcctcctgcctcagcct 900 v.1 2691
CCCTAGTAGCTGGGACTTTAAGTGTACACCACTGTGCCTGCTTTGAATCC 2740
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 901
ccctagtagctgggactttaagtgtacaccactgtgcctgctttgaatcc 950 v.1 2741
TTTACGAAGAGAAAAAAAAAATTAAAGAAAGCCTTTAGATTTATCCAATG 2790
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 951
tttacgaagagaaaaaaaaaattaaagaaagcctttagatttatccaatg 1000 v.1 2791
TTTACTACTGGGATTGCTTAAAGTGAGGCCCCTCCAACACCAGGGGGTTA 2840
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1001
tttactactgggattgcttaaagtgaggcccctccaacaccagggggtta 1050 v.1 2841
ATTCCTGTGATTGTGAAAGGGGCTACTTCCAAGGCATCTTCATGCAGGCA 2890
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1051
attcctgtgattgtgaaaggggctacttccaaggcatcttcatgcaggca 1100 v.1 2891
GCCCCTTGGGAGGGCACCTGAGAGCTGGTAGAGTCTGAAATTAGGGATGT 2940
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1101
gccccttgggagggcacctgagagctggtagagtctgaaattagggatgt 1150 v.1 2941
GAGCCTCGTGGTTACTGAGTAAGGTAAAATTGCATCCACCATTGTTTGTG 2990
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1151
gagcctcgtggttactgagtaaggtaaaattgcatccaccattgtttgtg 1200 v.1 2991
ATACCTTAGGGAATTGCTTGGACCTGGTGACAAGGGCTCCTGTTCAATAG 3040
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1201
ataccttagggaattgcttggacctggtgacaagggctcctgttcaatag 1250 v.1 3041
TGGTGTTGGGGAGAGAGAGAGCAGTGATTATAGACCGAGAGAGTAGGAGT 3090
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1251
tggtgttggggagagagagagcagtgattatagaccgagagagtaggagt 1300 v.1 3091
TGAGGTGAGGTGAAGGAGGTGCTGGGGGTGAGAATGTCGCCTTTCCCCCT 3140
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1301
tgaggtgaggtgaaggaggtgctgggggtgagaatgtcgcctttccccct 1350 v.1 3141
GGGTTTTGGATCACTAATTCAAGGCTCTTCTGGATGTTTCTCTGGGTTGG 3190
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1351
gggttttggatcactaattcaaggctcttctggatgtttctctgggttgg 1400 v.1 3191
CGCTGGAGTTCAATGAGGTTTATTTTTAGCTGGCCCACCCAGATACACTC 3240
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1401
ggctggagttcaatgaggtttatttttagctggcccacccagatacactc 1450 v.1 3241
AGCCAGAATACCTAGATTTAGTACCCAAACTCTTCTTAGTCTGAAATCTG 3290
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1451
agccagaatacctagatttagtacccaaactcttcttagtctgaaatctg 1500 v.1 3291
CTGGATTTCTGGCCTAAGGGACAGGCTCCCATCCTTCGTTCCCCAGCCAG 3340
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1501
ctggatttctggcctaagggagaggctcccatccttcgttccccagccag 1550 v.1 3341
CCTAGGACTTCGAATGTGGAGCCTGAAGATCTAAGATCCTAACATGTACA 3390
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1551
cctaggacttcgaatgtggagcctgaagatctaagatcctaacatgtaca 1600 v.1 3391
TTTTATGTAAATATGTGCATATTTGTACATAAAATGATATTCTGTTTTTA 3440
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline..vertline..vertli-
ne..vertline..vertline..vertline..vertline..vertline..vertline..vertline..-
vertline..vertline..vertline..vertline..vertline..vertline..vertline..vert-
line..vertline..vertline..vertline..vertline..vertline..vertline..vertline-
..vertline..vertline..vertline..vertline..vertline..vertline..vertline..ve-
rtline..vertline..vertline..vertline..vertline..vertline. v.9 1601
ttttatgtaaatatgtgcatatttgtacataaaatgatattctgttttta 1650 v.1 3441
AATAAACAGACAAAACTTG 3459 .vertline..vertline..vertline.-
.vertline..vertline..vertline..vertline..vertline..vertline..vertline..ver-
tline..vertline..vertline..vertline..vertline..vertline..vertline..vertlin-
e..vertline. v.9 1651 aataaacagacaaaacttg 1669
[1081]
77TABLE LIV(d) Peptide seguences of protein coded by 191P4D12(b)
v.9 (SEQ ID NO: 126) MRRELLAGIL LRITFNFFLF FFLPFPLVVF FIYFYFYFFL
EMESHYVAQA GLELLGSSNP 60 PASASLVAGT LSVHHCACFE SFTKRKKKLK
KAFRFIQCLL LGLLKVRPLQ HQGVNSCDCE 120 RGYFQGIFMQ AAPWEGT 137
[1082]
78TABLE LV(d) Amino acid sequence alignment of 191P4D12(b) v.1 and
191P4D12(b) v.9 (NO SIGNIFICANT MATCH)
[1083]
Sequence CWU 1
1
130 1 223 DNA Homo sapiens 1 gatcactaat tcaaggctct tctggatgtt
tctctgggtt ggggctggag ttcaatgagg 60 tttattttta gctggcccac
ccagatacac tcagccagaa tacctagatt tagtacccaa 120 actcttctta
gtctgaaatc tgctggattt ctggcctaag ggagaggctc ccatccttcg 180
ttccccagcc agcctaggac ttcgaatgtg gagcctgaag atc 223 2 3464 DNA Homo
sapiens CDS (264)...(1796) 2 ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac
240 gctgggcagt ctgcctttca acc atg ccc ctg tcc ctg gga gcc gag atg
tgg 293 Met Pro Leu Ser Leu Gly Ala Glu Met Trp 1 5 10 ggg cct gag
gcc tgg ctg ctg ctg ctg cta ctg ctg gca tca ttt aca 341 Gly Pro Glu
Ala Trp Leu Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr 15 20 25 ggc
cgg tgc ccc gcg ggt gag ctg gag acc tca gac gtg gta act gtg 389 Gly
Arg Cys Pro Ala Gly Glu Leu Glu Thr Ser Asp Val Val Thr Val 30 35
40 gtg ctg ggc cag gac gca aaa ctg ccc tgc ttc tac cga ggg gac tcc
437 Val Leu Gly Gln Asp Ala Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser
45 50 55 ggc gag caa gtg ggg caa gtg gca tgg gct cgg gtg gac gcg
ggc gaa 485 Gly Glu Gln Val Gly Gln Val Ala Trp Ala Arg Val Asp Ala
Gly Glu 60 65 70 ggc gcc cag gaa cta gcg cta ctg cac tcc aaa tac
ggg ctt cat gtg 533 Gly Ala Gln Glu Leu Ala Leu Leu His Ser Lys Tyr
Gly Leu His Val 75 80 85 90 agc ccg gct tac gag ggc cgc gtg gag cag
ccg ccg ccc cca cgc aac 581 Ser Pro Ala Tyr Glu Gly Arg Val Glu Gln
Pro Pro Pro Pro Arg Asn 95 100 105 ccc ctg gac ggc tca gtg ctc ctg
cgc aac gca gtg cag gcg gat gag 629 Pro Leu Asp Gly Ser Val Leu Leu
Arg Asn Ala Val Gln Ala Asp Glu 110 115 120 ggc gag tac gag tgc cgg
gtc agc acc ttc ccc gcc ggc agc ttc cag 677 Gly Glu Tyr Glu Cys Arg
Val Ser Thr Phe Pro Ala Gly Ser Phe Gln 125 130 135 gcg cgg ctg cgg
ctc cga gtg ctg gtg cct ccc ctg ccc tca ctg aat 725 Ala Arg Leu Arg
Leu Arg Val Leu Val Pro Pro Leu Pro Ser Leu Asn 140 145 150 cct ggt
cca gca cta gaa gag ggc cag ggc ctg acc ctg gca gcc tcc 773 Pro Gly
Pro Ala Leu Glu Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser 155 160 165
170 tgc aca gct gag ggc agc cca gcc ccc agc gtg acc tgg gac acg gag
821 Cys Thr Ala Glu Gly Ser Pro Ala Pro Ser Val Thr Trp Asp Thr Glu
175 180 185 gtc aaa ggc aca acg tcc agc cgt tcc ttc aag cac tcc cgc
tct gct 869 Val Lys Gly Thr Thr Ser Ser Arg Ser Phe Lys His Ser Arg
Ser Ala 190 195 200 gcc gtc acc tca gag ttc cac ttg gtg cct agc cgc
agc atg aat ggg 917 Ala Val Thr Ser Glu Phe His Leu Val Pro Ser Arg
Ser Met Asn Gly 205 210 215 cag cca ctg act tgt gtg gtg tcc cat cct
ggc ctg ctc cag gac caa 965 Gln Pro Leu Thr Cys Val Val Ser His Pro
Gly Leu Leu Gln Asp Gln 220 225 230 agg atc acc cac atc ctc cac gtg
tcc ttc ctt gct gag gcc tct gtg 1013 Arg Ile Thr His Ile Leu His
Val Ser Phe Leu Ala Glu Ala Ser Val 235 240 245 250 agg ggc ctt gaa
gac caa aat ctg tgg cac att ggc aga gaa gga gct 1061 Arg Gly Leu
Glu Asp Gln Asn Leu Trp His Ile Gly Arg Glu Gly Ala 255 260 265 atg
ctc aag tgc ctg agt gaa ggg cag ccc cct ccc tca tac aac tgg 1109
Met Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp 270
275 280 aca cgg ctg gat ggg cct ctg ccc agt ggg gta cga gtg gat ggg
gac 1157 Thr Arg Leu Asp Gly Pro Leu Pro Ser Gly Val Arg Val Asp
Gly Asp 285 290 295 act ttg ggc ttt ccc cca ctg acc act gag cac agc
ggc atc tac gtc 1205 Thr Leu Gly Phe Pro Pro Leu Thr Thr Glu His
Ser Gly Ile Tyr Val 300 305 310 tgc cat gtc agc aat gag ttc tcc tca
agg gat tct cag gtc act gtg 1253 Cys His Val Ser Asn Glu Phe Ser
Ser Arg Asp Ser Gln Val Thr Val 315 320 325 330 gat gtt ctt gac ccc
cag gaa gac tct ggg aag cag gtg gac cta gtg 1301 Asp Val Leu Asp
Pro Gln Glu Asp Ser Gly Lys Gln Val Asp Leu Val 335 340 345 tca gcc
tcg gtg gtg gtg gtg ggt gtg atc gcc gca ctc ttg ttc tgc 1349 Ser
Ala Ser Val Val Val Val Gly Val Ile Ala Ala Leu Leu Phe Cys 350 355
360 ctt ctg gtg gtg gtg gtg gtg ctc atg tcc cga tac cat cgg cgc aag
1397 Leu Leu Val Val Val Val Val Leu Met Ser Arg Tyr His Arg Arg
Lys 365 370 375 gcc cag cag atg acc cag aaa tat gag gag gag ctg acc
ctg acc agg 1445 Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu
Thr Leu Thr Arg 380 385 390 gag aac tcc atc cgg agg ctg cat tcc cat
cac acg gac ccc agg agc 1493 Glu Asn Ser Ile Arg Arg Leu His Ser
His His Thr Asp Pro Arg Ser 395 400 405 410 cag ccg gag gag agt gta
ggg ctg aga gcc gag ggc cac cct gat agt 1541 Gln Pro Glu Glu Ser
Val Gly Leu Arg Ala Glu Gly His Pro Asp Ser 415 420 425 ctc aag gac
aac agt agc tgc tct gtg atg agt gaa gag ccc gag ggc 1589 Leu Lys
Asp Asn Ser Ser Cys Ser Val Met Ser Glu Glu Pro Glu Gly 430 435 440
cgc agt tac tcc acg ctg acc acg gtg agg gag ata gaa aca cag act
1637 Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr Gln
Thr 445 450 455 gaa ctg ctg tct cca ggc tct ggg cgg gcc gag gag gag
gaa gat cag 1685 Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu
Glu Glu Asp Gln 460 465 470 gat gaa ggc atc aaa cag gcc atg aac cat
ttt gtt cag gag aat ggg 1733 Asp Glu Gly Ile Lys Gln Ala Met Asn
His Phe Val Gln Glu Asn Gly 475 480 485 490 acc cta cgg gcc aag ccc
acg ggc aat ggc atc tac atc aat ggg cgg 1781 Thr Leu Arg Ala Lys
Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg 495 500 505 gga cac ctg
gtc tga cccaggcctg cctcccttcc ctaggcctgg ctccttctgt 1836 Gly His
Leu Val * 510 tgacatggga gattttagct catcttgggg gcctccttaa
acacccccat ttcttgcgga 1896 agatgctccc catcccactg actgcttgac
ctttacctcc aacccttctg ttcatcggga 1956 gggctccacc aattgagtct
ctcccaccat gcatgcaggt cactgtgtgt gtgcatgtgt 2016 gcctgtgtga
gtgttgactg actgtgtgtg tgtggagggg tgactgtccg tggaggggtg 2076
actgtgtccg tggtgtgtat tatgctgtca tatcagagtc aagtgaactg tggtgtatgt
2136 gccacgggat ttgagtggtt gcgtgggcaa cactgtcagg gtttggcgtg
tgtgtcatgt 2196 ggctgtgtgt gacctctgcc tgaaaaagca ggtattttct
cagaccccag agcagtatta 2256 atgatgcaga ggttggagga gagaggtgga
gactgtggct cagacccagg tgtgcgggca 2316 tagctggagc tggaatctgc
ctccggtgtg agggaacctg tctcctacca cttcggagcc 2376 atgggggcaa
gtgtgaagca gccagtccct gggtcagcca gaggcttgaa ctgttacaga 2436
agccctctgc cctctggtgg cctctgggcc tgctgcatgt acatattttc tgtaaatata
2496 catgcgccgg gagcttcttg caggaatact gctccgaatc acttttaatt
tttttctttt 2556 ttttttcttg ccctttccat tagttgtatt ttttatttat
ttttattttt attttttttt 2616 agagatggag tctcactatg ttgctcaggc
tggccttgaa ctcctgggct caagcaatcc 2676 tcctgcctca gcctccctag
tagctgggac tttaagtgta caccactgtg cctgctttga 2736 atcctttacg
aagagaaaaa aaaaattaaa gaaagccttt agatttatcc aatgtttact 2796
actgggattg cttaaagtga ggcccctcca acaccagggg gttaattcct gtgattgtga
2856 aaggggctac ttccaaggca tcttcatgca ggcagcccct tgggagggca
cctgagagct 2916 ggtagagtct gaaattaggg atgtgagcct cgtggttact
gagtaaggta aaattgcatc 2976 caccattgtt tgtgatacct tagggaattg
cttggacctg gtgacaaggg ctcctgttca 3036 atagtggtgt tggggagaga
gagagcagtg attatagacc gagagagtag gagttgaggt 3096 gaggtgaagg
aggtgctggg ggtgagaatg tcgcctttcc ccctgggttt tggatcacta 3156
attcaaggct cttctggatg tttctctggg ttggggctgg agttcaatga ggtttatttt
3216 tagctggccc acccagatac actcagccag aatacctaga tttagtaccc
aaactcttct 3276 tagtctgaaa tctgctggat ttctggccta agggagaggc
tcccatcctt cgttccccag 3336 ccagcctagg acttcgaatg tggagcctga
agatctaaga tcctaacatg tacattttat 3396 gtaaatatgt gcatatttgt
acataaaatg atattctgtt tttaaataaa cagacaaaac 3456 ttgaaaaa 3464 3
510 PRT Homo sapiens 3 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly
Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe
Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val
Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe
Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp
Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu
Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90
95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val
100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu
Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg
Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn
Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu
Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val
Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser
Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His
Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215
220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu
225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu
Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met
Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn
Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val
Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His
Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser
Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335
Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340
345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val
Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln
Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu
Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp Pro
Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly
His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met
Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr
Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460
Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465
470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala
Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His
Leu Val 500 505 510 4 3464 DNA Homo sapiens CDS (264)...(1796) 4
ggccgtcgtt gttggccaca gcgtgggaag cagctctggg ggagctcgga gctcccgatc
60 acggcttctt gggggtagct acggctgggt gtgtagaacg gggccggggc
tggggctggg 120 tcccctagtg gagacccaag tgcgagaggc aagaactctg
cagcttcctg ccttctgggt 180 cagttcctta ttcaagtctg cagccggctc
ccagggagat ctcggtggaa cttcagaaac 240 gctgggcagt ctgcctttca acc atg
ccc ctg tcc ctg gga gcc gag atg tgg 293 Met Pro Leu Ser Leu Gly Ala
Glu Met Trp 1 5 10 ggg cct gag gcc tgg ctg ctg ctg ctg cta ctg ctg
gca tca ttt aca 341 Gly Pro Glu Ala Trp Leu Leu Leu Leu Leu Leu Leu
Ala Ser Phe Thr 15 20 25 ggc cgg tgc ccc gcg ggt gag ctg gag acc
tca gac gtg gta act gtg 389 Gly Arg Cys Pro Ala Gly Glu Leu Glu Thr
Ser Asp Val Val Thr Val 30 35 40 gtg ctg ggc cag gac gca aaa ctg
ccc tgc ctc tac cga ggg gac tcc 437 Val Leu Gly Gln Asp Ala Lys Leu
Pro Cys Leu Tyr Arg Gly Asp Ser 45 50 55 ggc gag caa gtg ggg caa
gtg gca tgg gct cgg gtg gac gcg ggc gaa 485 Gly Glu Gln Val Gly Gln
Val Ala Trp Ala Arg Val Asp Ala Gly Glu 60 65 70 ggc gcc cag gaa
cta gcg cta ctg cac tcc aaa tac ggg ctt cat gtg 533 Gly Ala Gln Glu
Leu Ala Leu Leu His Ser Lys Tyr Gly Leu His Val 75 80 85 90 agc ccg
gct tac gag ggc cgc gtg gag cag ccg ccg ccc cca cgc aac 581 Ser Pro
Ala Tyr Glu Gly Arg Val Glu Gln Pro Pro Pro Pro Arg Asn 95 100 105
ccc ctg gac ggc tca gtg ctc ctg cgc aac gca gtg cag gcg gat gag 629
Pro Leu Asp Gly Ser Val Leu Leu Arg Asn Ala Val Gln Ala Asp Glu 110
115 120 ggc gag tac gag tgc cgg gtc agc acc ttc ccc gcc ggc agc ttc
cag 677 Gly Glu Tyr Glu Cys Arg Val Ser Thr Phe Pro Ala Gly Ser Phe
Gln 125 130 135 gcg cgg ctg cgg ctc cga gtg ctg gtg cct ccc ctg ccc
tca ctg aat 725 Ala Arg Leu Arg Leu Arg Val Leu Val Pro Pro Leu Pro
Ser Leu Asn 140 145 150 cct ggt cca gca cta gaa gag ggc cag ggc ctg
acc ctg gca gcc tcc 773 Pro Gly Pro Ala Leu Glu Glu Gly Gln Gly Leu
Thr Leu Ala Ala Ser 155 160 165 170 tgc aca gct gag ggc agc cca gcc
ccc agc gtg acc tgg gac acg gag 821 Cys Thr Ala Glu Gly Ser Pro Ala
Pro Ser Val Thr Trp Asp Thr Glu 175 180 185 gtc aaa ggc aca acg tcc
agc cgt tcc ttc aag cac tcc cgc tct gct 869 Val Lys Gly Thr Thr Ser
Ser Arg Ser Phe Lys His Ser Arg Ser Ala 190 195 200 gcc gtc acc tca
gag ttc cac ttg gtg cct agc cgc agc atg aat ggg 917 Ala Val Thr Ser
Glu Phe His Leu Val Pro Ser Arg Ser Met Asn Gly 205 210 215 cag cca
ctg act tgt gtg gtg tcc cat cct ggc ctg ctc cag gac caa 965 Gln Pro
Leu Thr Cys Val Val Ser His Pro Gly Leu Leu Gln Asp Gln 220 225 230
agg atc acc cac atc ctc cac gtg tcc ttc ctt gct gag gcc tct gtg
1013 Arg Ile Thr His Ile Leu His Val Ser Phe Leu Ala Glu Ala Ser
Val 235 240 245 250 agg ggc ctt gaa gac caa aat ctg tgg cac att ggc
aga gaa gga gct 1061 Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile
Gly Arg Glu Gly Ala 255 260 265 atg ctc aag tgc ctg agt gaa ggg cag
ccc cct ccc tca tac aac tgg 1109 Met Leu Lys Cys Leu Ser Glu Gly
Gln Pro Pro Pro Ser Tyr Asn Trp 270 275 280 aca cgg ctg gat ggg cct
ctg ccc agt ggg gta cga gtg gat ggg gac 1157 Thr Arg Leu Asp Gly
Pro Leu Pro Ser Gly Val Arg Val Asp Gly Asp 285 290 295 act ttg ggc
ttt ccc cca ctg acc act gag cac agc ggc atc tac gtc 1205 Thr Leu
Gly Phe Pro Pro Leu Thr Thr Glu His Ser Gly Ile Tyr Val 300 305 310
tgc cat gtc agc aat gag ttc tcc tca agg gat tct cag gtc act gtg
1253 Cys His Val Ser Asn Glu Phe Ser Ser Arg Asp Ser Gln Val Thr
Val 315 320 325 330 gat gtt ctt gac ccc cag gaa gac tct ggg aag cag
gtg gac cta gtg 1301 Asp Val Leu Asp Pro Gln Glu Asp Ser Gly Lys
Gln Val Asp Leu Val 335 340 345 tca gcc tcg gtg gtg gtg gtg ggt gtg
atc gcc gca ctc ttg ttc tgc 1349 Ser Ala Ser Val Val Val Val Gly
Val Ile Ala Ala Leu Leu Phe Cys 350 355 360 ctt ctg gtg gtg gtg gtg
gtg ctc atg tcc cga tac cat cgg cgc aag 1397 Leu Leu Val Val Val
Val Val Leu Met Ser Arg Tyr His Arg Arg Lys 365 370 375 gcc cag cag
atg acc cag aaa tat gag gag gag ctg acc ctg acc agg 1445 Ala Gln
Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg 380 385 390
gag aac tcc atc cgg agg ctg cat tcc cat cac acg gac ccc agg agc
1493 Glu Asn Ser Ile Arg Arg Leu His Ser His His Thr Asp Pro Arg
Ser 395 400 405 410 cag ccg gag gag agt gta ggg ctg aga gcc gag ggc
cac cct gat agt 1541 Gln Pro Glu Glu Ser Val Gly
Leu Arg Ala Glu Gly His Pro Asp Ser 415 420 425 ctc aag gac aac agt
agc tgc tct gtg atg agt gaa gag ccc gag ggc 1589 Leu Lys Asp Asn
Ser Ser Cys Ser Val Met Ser Glu Glu Pro Glu Gly 430 435 440 cgc agt
tac tcc acg ctg acc acg gtg agg gag ata gaa aca cag act 1637 Arg
Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr Gln Thr 445 450
455 gaa ctg ctg tct cca ggc tct ggg cgg gcc gag gag gag gaa gat cag
1685 Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu Glu Asp
Gln 460 465 470 gat gaa ggc atc aaa cag gcc atg aac cat ttt gtt cag
gag aat ggg 1733 Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val
Gln Glu Asn Gly 475 480 485 490 acc cta cgg gcc aag ccc acg ggc aat
ggc atc tac atc aat ggg cgg 1781 Thr Leu Arg Ala Lys Pro Thr Gly
Asn Gly Ile Tyr Ile Asn Gly Arg 495 500 505 gga cac ctg gtc tga
cccaggcctg cctcccttcc ctaggcctgg ctccttctgt 1836 Gly His Leu Val *
510 tgacatggga gattttagct catcttgggg gcctccttaa acacccccat
ttcttgcgga 1896 agatgctccc catcccactg actgcttgac ctttacctcc
aacccttctg ttcatcggga 1956 gggctccacc aattgagtct ctcccaccat
gcatgcaggt cactgtgtgt gtgcatgtgt 2016 gcctgtgtga gtgttgactg
actgtgtgtg tgtggagggg tgactgtccg tggaggggtg 2076 actgtgtccg
tggtgtgtat tatgctgtca tatcagagtc aagtgaactg tggtgtatgt 2136
gccacgggat ttgagtggtt gcgtgggcaa cactgtcagg gtttggcgtg tgtgtcatgt
2196 ggctgtgtgt gacctctgcc tgaaaaagca ggtattttct cagaccccag
agcagtatta 2256 atgatgcaga ggttggagga gagaggtgga gactgtggct
cagacccagg tgtgcgggca 2316 tagctggagc tggaatctgc ctccggtgtg
agggaacctg tctcctacca cttcggagcc 2376 atgggggcaa gtgtgaagca
gccagtccct gggtcagcca gaggcttgaa ctgttacaga 2436 agccctctgc
cctctggtgg cctctgggcc tgctgcatgt acatattttc tgtaaatata 2496
catgcgccgg gagcttcttg caggaatact gctccgaatc acttttaatt tttttctttt
2556 ttttttcttg ccctttccat tagttgtatt ttttatttat ttttattttt
attttttttt 2616 agagatggag tctcactatg ttgctcaggc tggccttgaa
ctcctgggct caagcaatcc 2676 tcctgcctca gcctccctag tagctgggac
tttaagtgta caccactgtg cctgctttga 2736 atcctttacg aagagaaaaa
aaaaattaaa gaaagccttt agatttatcc aatgtttact 2796 actgggattg
cttaaagtga ggcccctcca acaccagggg gttaattcct gtgattgtga 2856
aaggggctac ttccaaggca tcttcatgca ggcagcccct tgggagggca cctgagagct
2916 ggtagagtct gaaattaggg atgtgagcct cgtggttact gagtaaggta
aaattgcatc 2976 caccattgtt tgtgatacct tagggaattg cttggacctg
gtgacaaggg ctcctgttca 3036 atagtggtgt tggggagaga gagagcagtg
attatagacc gagagagtag gagttgaggt 3096 gaggtgaagg aggtgctggg
ggtgagaatg tcgcctttcc ccctgggttt tggatcacta 3156 attcaaggct
cttctggatg tttctctggg ttggggctgg agttcaatga ggtttatttt 3216
tagctggccc acccagatac actcagccag aatacctaga tttagtaccc aaactcttct
3276 tagtctgaaa tctgctggat ttctggccta agggagaggc tcccatcctt
cgttccccag 3336 ccagcctagg acttcgaatg tggagcctga agatctaaga
tcctaacatg tacattttat 3396 gtaaatatgt gcatatttgt acataaaatg
atattctgtt tttaaataaa cagacaaaac 3456 ttgaaaaa 3464 5 510 PRT Homo
sapiens 5 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala
Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg
Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val
Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Leu Tyr Arg Gly
Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val
Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser
Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val
Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110
Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115
120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu
Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro
Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser
Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp
Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His
Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro
Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser
His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235
240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln
245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys
Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg
Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp
Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile
Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp
Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser
Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val
Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360
365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln
370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile
Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln
Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp
Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu
Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu
Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg
Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480
Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485
490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500
505 510 6 3464 DNA Homo sapiens CDS (264)...(1796) 6 ggccgtcgtt
gttggccaca gcgtgggaag cagctctggg ggagctcgga gctcccgatc 60
acggcttctt gggggtagct acggctgggt gtgtagaacg gggccggggc tggggctggg
120 tcccctagtg gagacccaag tgcgagaggc aagaactctg cagcttcctg
ccttctgggt 180 cagttcctta ttcaagtctg cagccggctc ccagggagat
ctcggtggaa cttcagaaac 240 gctgggcagt ctgcctttca acc atg ccc ctg tcc
ctg gga gcc gag atg tgg 293 Met Pro Leu Ser Leu Gly Ala Glu Met Trp
1 5 10 ggg cct gag gcc tgg ctg ctg ctg ctg cta ctg ctg gca tca ttt
aca 341 Gly Pro Glu Ala Trp Leu Leu Leu Leu Leu Leu Leu Ala Ser Phe
Thr 15 20 25 ggc cgg tgc ccc gcg ggt gag ctg gag acc tca gac gtg
gta act gtg 389 Gly Arg Cys Pro Ala Gly Glu Leu Glu Thr Ser Asp Val
Val Thr Val 30 35 40 gtg ctg ggc cag gac gca aaa ctg ccc tgc ttc
tac cga ggg gac tcc 437 Val Leu Gly Gln Asp Ala Lys Leu Pro Cys Phe
Tyr Arg Gly Asp Ser 45 50 55 ggc gag caa gtg ggg caa gtg gca tgg
gct cgg gtg gac gcg ggc gaa 485 Gly Glu Gln Val Gly Gln Val Ala Trp
Ala Arg Val Asp Ala Gly Glu 60 65 70 ggc gcc cag gaa cta gcg cta
ctg cac tcc aaa tac ggg ctt cat gtg 533 Gly Ala Gln Glu Leu Ala Leu
Leu His Ser Lys Tyr Gly Leu His Val 75 80 85 90 agc ccg gct tac gag
ggc cgc gtg gag cag ccg ccg ccc cca cgc aac 581 Ser Pro Ala Tyr Glu
Gly Arg Val Glu Gln Pro Pro Pro Pro Arg Asn 95 100 105 ccc ctg gac
ggc tca gtg ctc ctg cgc aac gca gtg cag gcg gat gag 629 Pro Leu Asp
Gly Ser Val Leu Leu Arg Asn Ala Val Gln Ala Asp Glu 110 115 120 ggc
gag tac gag tgc cgg gtc agc acc ttc ccc gcc ggc agc ttc cag 677 Gly
Glu Tyr Glu Cys Arg Val Ser Thr Phe Pro Ala Gly Ser Phe Gln 125 130
135 gcg cgg ctg cgg ctc cga gtg ctg gtg cct ccc ctg ccc tca ctg aat
725 Ala Arg Leu Arg Leu Arg Val Leu Val Pro Pro Leu Pro Ser Leu Asn
140 145 150 cct ggt cca gca cta gaa gag ggc cag ggc ctg acc ctg gca
gcc tcc 773 Pro Gly Pro Ala Leu Glu Glu Gly Gln Gly Leu Thr Leu Ala
Ala Ser 155 160 165 170 tgc aca gct gag ggc agc cca gcc ccc agc gtg
acc tgg gac acg gag 821 Cys Thr Ala Glu Gly Ser Pro Ala Pro Ser Val
Thr Trp Asp Thr Glu 175 180 185 gtc aaa ggc aca acg tcc agc cgt tcc
ttc aag cac tcc cgc tct gct 869 Val Lys Gly Thr Thr Ser Ser Arg Ser
Phe Lys His Ser Arg Ser Ala 190 195 200 gcc gtc acc tca gag ttc cac
ttg gtg cct agc cgc agc atg aat ggg 917 Ala Val Thr Ser Glu Phe His
Leu Val Pro Ser Arg Ser Met Asn Gly 205 210 215 cag cca ctg act tgt
gtg gtg tcc cat cct ggc ctg ctc cag gac caa 965 Gln Pro Leu Thr Cys
Val Val Ser His Pro Gly Leu Leu Gln Asp Gln 220 225 230 agg atc acc
cac atc ctc cac gtg tcc ttc ctt gct gag gcc tct gtg 1013 Arg Ile
Thr His Ile Leu His Val Ser Phe Leu Ala Glu Ala Ser Val 235 240 245
250 agg ggc ctt gaa gac caa aat ctg tgg cac att ggc aga gaa gga gct
1061 Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile Gly Arg Glu Gly
Ala 255 260 265 atg ctc aag tgc ctg agt gaa ggg cag ccc cct ccc tca
tac aac tgg 1109 Met Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro Pro
Ser Tyr Asn Trp 270 275 280 aca cgg ctg gat ggg cct ctg ccc agt ggg
gta cga gtg gat ggg gac 1157 Thr Arg Leu Asp Gly Pro Leu Pro Ser
Gly Val Arg Val Asp Gly Asp 285 290 295 act ttg ggc ttt ccc cca ctg
acc act gag cac agc ggc atc tac gtc 1205 Thr Leu Gly Phe Pro Pro
Leu Thr Thr Glu His Ser Gly Ile Tyr Val 300 305 310 tgc cat gtc agc
aat gag ttc tcc tca agg gat tct cag gtc act gtg 1253 Cys His Val
Ser Asn Glu Phe Ser Ser Arg Asp Ser Gln Val Thr Val 315 320 325 330
gat gtt ctt gac ccc cag gaa gac tct ggg aag cag gtg gac cta gtg
1301 Asp Val Leu Asp Pro Gln Glu Asp Ser Gly Lys Gln Val Asp Leu
Val 335 340 345 tca gcc tcg gtg gtg gtg gtg ggt gtg atc gcc gca ctc
ttg ttc tgc 1349 Ser Ala Ser Val Val Val Val Gly Val Ile Ala Ala
Leu Leu Phe Cys 350 355 360 ctt ctg gtg gtg gtg gtg gtg ctc atg tcc
cga tac cat cgg cgc aag 1397 Leu Leu Val Val Val Val Val Leu Met
Ser Arg Tyr His Arg Arg Lys 365 370 375 gcc cag cag atg acc cag aaa
tat gag gag gag ctg acc ctg acc agg 1445 Ala Gln Gln Met Thr Gln
Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg 380 385 390 gag aac tcc atc
cgg agg ctg cat tcc cat cac acg gac ccc agg agc 1493 Glu Asn Ser
Ile Arg Arg Leu His Ser His His Thr Asp Pro Arg Ser 395 400 405 410
cag ccg gag gag agt gta ggg ctg aga gcc gag ggc cac cct gat agt
1541 Gln Pro Glu Glu Ser Val Gly Leu Arg Ala Glu Gly His Pro Asp
Ser 415 420 425 ctc aag gac aac agt agc tgc tct gtg atg agt gaa gag
ccc gag ggc 1589 Leu Lys Asp Asn Ser Ser Cys Ser Val Met Ser Glu
Glu Pro Glu Gly 430 435 440 cgc agt tac tcc acg ctg acc acg gtg agg
gag ata gaa aca cag act 1637 Arg Ser Tyr Ser Thr Leu Thr Thr Val
Arg Glu Ile Glu Thr Gln Thr 445 450 455 gaa ctg ctg tct cca ggc tct
ggg cgg gcc gag gag gag gaa gat cag 1685 Glu Leu Leu Ser Pro Gly
Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln 460 465 470 gat gaa ggc atc
aaa cag gcc atg aac cat ttt gtt cag gag aat ggg 1733 Asp Glu Gly
Ile Lys Gln Ala Met Asn His Phe Val Gln Glu Asn Gly 475 480 485 490
acc cta cgg gcc aag ccc acg ggc aat ggc atc tac atc aat ggg cgg
1781 Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly
Arg 495 500 505 gga cac ctg gtc tga cccaggcctg cctcccttcc
ctaggcctgg ctccttctgt 1836 Gly His Leu Val * 510 tgacatggga
gattttagct catcttgggg gcctccttaa acacccccat ttcttgcgga 1896
agatgctccc catcccactg actgcttgac ctttacctcc aacccttctg ttcatcggga
1956 gggctccacc aattgagtct ctcccaccat gcatgcaggt cactgtgtgt
gtgcatgtgt 2016 gcctgtgtga gtgttgactg actgtgtgtg tgtggagggg
tgactgtccg tggaggggtg 2076 actgtgtccg tggtgtgtat tatgctgtca
tatcagagtc aagtgaactg tggtgtatgt 2136 gccacgggat ttgagtggtt
gcgtgggcaa cactgtcagg gtttggcttg tgtgtcatgt 2196 ggctgtgtgt
gacctctgcc tgaaaaagca ggtattttct cagaccccag agcagtatta 2256
atgatgcaga ggttggagga gagaggtgga gactgtggct cagacccagg tgtgcgggca
2316 tagctggagc tggaatctgc ctccggtgtg agggaacctg tctcctacca
cttcggagcc 2376 atgggggcaa gtgtgaagca gccagtccct gggtcagcca
gaggcttgaa ctgttacaga 2436 agccctctgc cctctggtgg cctctgggcc
tgctgcatgt acatattttc tgtaaatata 2496 catgcgccgg gagcttcttg
caggaatact gctccgaatc acttttaatt tttttctttt 2556 ttttttcttg
ccctttccat tagttgtatt ttttatttat ttttattttt attttttttt 2616
agagatggag tctcactatg ttgctcaggc tggccttgaa ctcctgggct caagcaatcc
2676 tcctgcctca gcctccctag tagctgggac tttaagtgta caccactgtg
cctgctttga 2736 atcctttacg aagagaaaaa aaaaattaaa gaaagccttt
agatttatcc aatgtttact 2796 actgggattg cttaaagtga ggcccctcca
acaccagggg gttaattcct gtgattgtga 2856 aaggggctac ttccaaggca
tcttcatgca ggcagcccct tgggagggca cctgagagct 2916 ggtagagtct
gaaattaggg atgtgagcct cgtggttact gagtaaggta aaattgcatc 2976
caccattgtt tgtgatacct tagggaattg cttggacctg gtgacaaggg ctcctgttca
3036 atagtggtgt tggggagaga gagagcagtg attatagacc gagagagtag
gagttgaggt 3096 gaggtgaagg aggtgctggg ggtgagaatg tcgcctttcc
ccctgggttt tggatcacta 3156 attcaaggct cttctggatg tttctctggg
ttggggctgg agttcaatga ggtttatttt 3216 tagctggccc acccagatac
actcagccag aatacctaga tttagtaccc aaactcttct 3276 tagtctgaaa
tctgctggat ttctggccta agggagaggc tcccatcctt cgttccccag 3336
ccagcctagg acttcgaatg tggagcctga agatctaaga tcctaacatg tacattttat
3396 gtaaatatgt gcatatttgt acataaaatg atattctgtt tttaaataaa
cagacaaaac 3456 ttgaaaaa 3464 7 510 PRT Homo sapiens 7 Met Pro Leu
Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu
Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25
30 Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala
35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val
Gly Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala
Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val
Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro
Arg Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val
Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe
Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu
Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155
160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser
165 170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr
Thr Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val
Thr Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly
Gln Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln
Asp Gln Arg Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu
Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp
His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu
Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280
285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro
290 295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser
Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp
Val Leu Asp Pro Gln 325 330
335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val
340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val
Val Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln
Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg
Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp
Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu
Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val
Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr
Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455
460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln
465 470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg
Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly
His Leu Val 500 505 510 8 3464 DNA Homo sapiens CDS (264)...(1796)
8 ggccgtcgtt gttggccaca gcgtgggaag cagctctggg ggagctcgga gctcccgatc
60 acggcttctt gggggtagct acggctgggt gtgtagaacg gggccggggc
tggggctggg 120 tcccctagtg gagacccaag tgcgagaggc aagaactctg
cagcttcctg ccttctgggt 180 cagttcctta ttcaagtctg cagccggctc
ccagggagat ctcggtggaa cttcagaaac 240 gctgggcagt ctgcctttca acc atg
ccc ctg tcc ctg gga gcc gag atg tgg 293 Met Pro Leu Ser Leu Gly Ala
Glu Met Trp 1 5 10 ggg cct gag gcc tgg ctg ctg ctg ctg cta ctg ctg
gca tca ttt aca 341 Gly Pro Glu Ala Trp Leu Leu Leu Leu Leu Leu Leu
Ala Ser Phe Thr 15 20 25 ggc cgg tgc ccc gcg ggt gag ctg gag acc
tca gac gtg gta act gtg 389 Gly Arg Cys Pro Ala Gly Glu Leu Glu Thr
Ser Asp Val Val Thr Val 30 35 40 gtg ctg ggc cag gac gca aaa ctg
ccc tgc ttc tac cga ggg gac tcc 437 Val Leu Gly Gln Asp Ala Lys Leu
Pro Cys Phe Tyr Arg Gly Asp Ser 45 50 55 ggc gag caa gtg ggg caa
gtg gca tgg gct cgg gtg gac gcg ggc gaa 485 Gly Glu Gln Val Gly Gln
Val Ala Trp Ala Arg Val Asp Ala Gly Glu 60 65 70 ggc gcc cag gaa
cta gcg cta ctg cac tcc aaa tac ggg ctt cat gtg 533 Gly Ala Gln Glu
Leu Ala Leu Leu His Ser Lys Tyr Gly Leu His Val 75 80 85 90 agc ccg
gct tac gag ggc cgc gtg gag cag ccg ccg ccc cca cgc aac 581 Ser Pro
Ala Tyr Glu Gly Arg Val Glu Gln Pro Pro Pro Pro Arg Asn 95 100 105
ccc ctg gac ggc tca gtg ctc ctg cgc aac gca gtg cag gcg gat gag 629
Pro Leu Asp Gly Ser Val Leu Leu Arg Asn Ala Val Gln Ala Asp Glu 110
115 120 ggc gag tac gag tgc cgg gtc agc acc ttc ccc gcc ggc agc ttc
cag 677 Gly Glu Tyr Glu Cys Arg Val Ser Thr Phe Pro Ala Gly Ser Phe
Gln 125 130 135 gcg cgg ctg cgg ctc cga gtg ctg gtg cct ccc ctg ccc
tca ctg aat 725 Ala Arg Leu Arg Leu Arg Val Leu Val Pro Pro Leu Pro
Ser Leu Asn 140 145 150 cct ggt cca gca cta gaa gag ggc cag ggc ctg
acc ctg gca gcc tcc 773 Pro Gly Pro Ala Leu Glu Glu Gly Gln Gly Leu
Thr Leu Ala Ala Ser 155 160 165 170 tgc aca gct gag ggc agc cca gcc
ccc agc gtg acc tgg gac acg gag 821 Cys Thr Ala Glu Gly Ser Pro Ala
Pro Ser Val Thr Trp Asp Thr Glu 175 180 185 gtc aaa ggc aca acg tcc
agc cgt tcc ttc aag cac tcc cgc tct gct 869 Val Lys Gly Thr Thr Ser
Ser Arg Ser Phe Lys His Ser Arg Ser Ala 190 195 200 gcc gtc acc tca
gag ttc cac ttg gtg cct agc cgc agc atg aat ggg 917 Ala Val Thr Ser
Glu Phe His Leu Val Pro Ser Arg Ser Met Asn Gly 205 210 215 cag cca
ctg act tgt gtg gtg tcc cat cct ggc ctg ctc cag gac caa 965 Gln Pro
Leu Thr Cys Val Val Ser His Pro Gly Leu Leu Gln Asp Gln 220 225 230
agg atc acc cac atc ctc cac gtg tcc ttc ctt gct gag gcc tct gtg
1013 Arg Ile Thr His Ile Leu His Val Ser Phe Leu Ala Glu Ala Ser
Val 235 240 245 250 agg ggc ctt gaa gac caa aat ctg tgg cac att ggc
aga gaa gga gct 1061 Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile
Gly Arg Glu Gly Ala 255 260 265 atg ctc aag tgc ctg agt gaa ggg cag
ccc cct ccc tca tac aac tgg 1109 Met Leu Lys Cys Leu Ser Glu Gly
Gln Pro Pro Pro Ser Tyr Asn Trp 270 275 280 aca cgg ctg gat ggg cct
ctg ccc agt ggg gta cga gtg gat ggg gac 1157 Thr Arg Leu Asp Gly
Pro Leu Pro Ser Gly Val Arg Val Asp Gly Asp 285 290 295 act ttg ggc
ttt ccc cca ctg acc act gag cac agc ggc atc tac gtc 1205 Thr Leu
Gly Phe Pro Pro Leu Thr Thr Glu His Ser Gly Ile Tyr Val 300 305 310
tgc cat gtc agc aat gag ttc tcc tca agg gat tct cag gtc act gtg
1253 Cys His Val Ser Asn Glu Phe Ser Ser Arg Asp Ser Gln Val Thr
Val 315 320 325 330 gat gtt ctt gac ccc cag gaa gac tct ggg aag cag
gtg gac cta gtg 1301 Asp Val Leu Asp Pro Gln Glu Asp Ser Gly Lys
Gln Val Asp Leu Val 335 340 345 tca gcc tcg gtg gtg gtg gtg ggt gtg
atc gcc gca ctc ttg ttc tgc 1349 Ser Ala Ser Val Val Val Val Gly
Val Ile Ala Ala Leu Leu Phe Cys 350 355 360 ctt ctg gtg gtg gtg gtg
gtg ctc atg tcc cga tac cat cgg cgc aag 1397 Leu Leu Val Val Val
Val Val Leu Met Ser Arg Tyr His Arg Arg Lys 365 370 375 gcc cag cag
atg acc cag aaa tat gag gag gag ctg acc ctg acc agg 1445 Ala Gln
Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg 380 385 390
gag aac tcc atc cgg agg ctg cat tcc cat cac acg gac ccc agg agc
1493 Glu Asn Ser Ile Arg Arg Leu His Ser His His Thr Asp Pro Arg
Ser 395 400 405 410 cag ccg gag gag agt gta ggg ctg aga gcc gag ggc
cac cct gat agt 1541 Gln Pro Glu Glu Ser Val Gly Leu Arg Ala Glu
Gly His Pro Asp Ser 415 420 425 ctc aag gac aac agt agc tgc tct gtg
atg agt gaa gag ccc gag ggc 1589 Leu Lys Asp Asn Ser Ser Cys Ser
Val Met Ser Glu Glu Pro Glu Gly 430 435 440 cgc agt tac tcc acg ctg
acc acg gtg agg gag ata gaa aca cag act 1637 Arg Ser Tyr Ser Thr
Leu Thr Thr Val Arg Glu Ile Glu Thr Gln Thr 445 450 455 gaa ctg ctg
tct cca ggc tct ggg cgg gcc gag gag gag gaa gat cag 1685 Glu Leu
Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln 460 465 470
gat gaa ggc atc aaa cag gcc atg aac cat ttt gtt cag gag aat ggg
1733 Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val Gln Glu Asn
Gly 475 480 485 490 acc cta cgg gcc aag ccc acg ggc aat ggc atc tac
atc aat ggg cgg 1781 Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile
Tyr Ile Asn Gly Arg 495 500 505 gga cac ctg gtc tga cccaggcctg
cctcccttcc ctaggcctgg ctccttctgt 1836 Gly His Leu Val * 510
tgacatggga gattttagct catcttgggg gcctccttaa acacccccat ttcttgcgga
1896 agatgctccc catcccactg actgcttgac ctttacctcc aacccttctg
ttcatcggga 1956 gggctccacc aattgagtct ctcccaccat gcatgcaggt
cactgtgtgt gtgcatgtgt 2016 gcctgtgtga gtgttgactg actgtgtgtg
tgtggagggg tgactgtccg tggaggggtg 2076 actgtgtccg tggtgtgtat
tatgctgtca tatcagagtc aagtgaactg tggtgtatgt 2136 gccacgggat
ttgagtggtt gcgtgggcaa cactgtcagg gtttggcgtg tgtgtcatgt 2196
ggctgtgtgt gacctctgcc tgaaaaagca ggtattttct cagaccccag agcagtatta
2256 atgatgcaga ggttggagga gagaggtgga gactgtggct cagacccagg
tgtgcgggca 2316 tagctggagc tggaatctgc ctccagtgtg agggaacctg
tctcctacca cttcggagcc 2376 atgggggcaa gtgtgaagca gccagtccct
gggtcagcca gaggcttgaa ctgttacaga 2436 agccctctgc cctctggtgg
cctctgggcc tgctgcatgt acatattttc tgtaaatata 2496 catgcgccgg
gagcttcttg caggaatact gctccgaatc acttttaatt tttttctttt 2556
ttttttcttg ccctttccat tagttgtatt ttttatttat ttttattttt attttttttt
2616 agagatggag tctcactatg ttgctcaggc tggccttgaa ctcctgggct
caagcaatcc 2676 tcctgcctca gcctccctag tagctgggac tttaagtgta
caccactgtg cctgctttga 2736 atcctttacg aagagaaaaa aaaaattaaa
gaaagccttt agatttatcc aatgtttact 2796 actgggattg cttaaagtga
ggcccctcca acaccagggg gttaattcct gtgattgtga 2856 aaggggctac
ttccaaggca tcttcatgca ggcagcccct tgggagggca cctgagagct 2916
ggtagagtct gaaattaggg atgtgagcct cgtggttact gagtaaggta aaattgcatc
2976 caccattgtt tgtgatacct tagggaattg cttggacctg gtgacaaggg
ctcctgttca 3036 atagtggtgt tggggagaga gagagcagtg attatagacc
gagagagtag gagttgaggt 3096 gaggtgaagg aggtgctggg ggtgagaatg
tcgcctttcc ccctgggttt tggatcacta 3156 attcaaggct cttctggatg
tttctctggg ttggggctgg agttcaatga ggtttatttt 3216 tagctggccc
acccagatac actcagccag aatacctaga tttagtaccc aaactcttct 3276
tagtctgaaa tctgctggat ttctggccta agggagaggc tcccatcctt cgttccccag
3336 ccagcctagg acttcgaatg tggagcctga agatctaaga tcctaacatg
tacattttat 3396 gtaaatatgt gcatatttgt acataaaatg atattctgtt
tttaaataaa cagacaaaac 3456 ttgaaaaa 3464 9 510 PRT Homo sapiens 9
Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5
10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala
Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly
Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly
Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly
Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly
Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro
Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg
Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val
Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135
140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu
145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala
Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val
Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser
Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser
Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly
Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235 240 His Val
Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255
Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260
265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly
Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly
Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys
His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val
Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln
Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val Gly Val Ile
Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360 365 Val Leu
Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380
Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385
390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln Pro Glu Glu
Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp Ser Leu Lys
Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu Pro Glu Gly
Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu Ile Glu Thr
Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg Ala Glu Glu
Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480 Ala Met Asn
His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485 490 495 Thr
Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500 505 510 10
3464 DNA Homo sapiens CDS (264)...(1796) 10 ggccgtcgtt gttggccaca
gcgtgggaag cagctctggg ggagctcgga gctcccgatc 60 acggcttctt
gggggtagct acggctgggt gtgtagaacg gggccggggc tggggctggg 120
tcccctagtg gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt
180 cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa
cttcagaaac 240 gctgggcagt ctgcctttca acc atg ccc ctg tcc ctg gga
gcc gag atg tgg 293 Met Pro Leu Ser Leu Gly Ala Glu Met Trp 1 5 10
ggg cct gag gcc tgg ctg ctg ctg ctg cta ctg ctg gca tca ttt aca 341
Gly Pro Glu Ala Trp Leu Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr 15
20 25 ggc cgg tgc ccc gcg ggt gag ctg gag acc tca gac gtg gta act
gtg 389 Gly Arg Cys Pro Ala Gly Glu Leu Glu Thr Ser Asp Val Val Thr
Val 30 35 40 gtg ctg ggc cag gac gca aaa ctg ccc tgc ttc tac cga
ggg gac tcc 437 Val Leu Gly Gln Asp Ala Lys Leu Pro Cys Phe Tyr Arg
Gly Asp Ser 45 50 55 ggc gag caa gtg ggg caa gtg gca tgg gct cgg
gtg gac gcg ggc gaa 485 Gly Glu Gln Val Gly Gln Val Ala Trp Ala Arg
Val Asp Ala Gly Glu 60 65 70 ggc gcc cag gaa cta gcg cta ctg cac
tcc aaa tac ggg ctt cat gtg 533 Gly Ala Gln Glu Leu Ala Leu Leu His
Ser Lys Tyr Gly Leu His Val 75 80 85 90 agc ccg gct tac gag ggc cgc
gtg gag cag ccg ccg ccc cca cgc aac 581 Ser Pro Ala Tyr Glu Gly Arg
Val Glu Gln Pro Pro Pro Pro Arg Asn 95 100 105 ccc ctg gac ggc tca
gtg ctc ctg cgc aac gca gtg cag gcg gat gag 629 Pro Leu Asp Gly Ser
Val Leu Leu Arg Asn Ala Val Gln Ala Asp Glu 110 115 120 ggc gag tac
gag tgc cgg gtc agc acc ttc ccc gcc ggc agc ttc cag 677 Gly Glu Tyr
Glu Cys Arg Val Ser Thr Phe Pro Ala Gly Ser Phe Gln 125 130 135 gcg
cgg ctg cgg ctc cga gtg ctg gtg cct ccc ctg ccc tca ctg aat 725 Ala
Arg Leu Arg Leu Arg Val Leu Val Pro Pro Leu Pro Ser Leu Asn 140 145
150 cct ggt cca gca cta gaa gag ggc cag ggc ctg acc ctg gca gcc tcc
773 Pro Gly Pro Ala Leu Glu Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser
155 160 165 170 tgc aca gct gag ggc agc cca gcc ccc agc gtg acc tgg
gac acg gag 821 Cys Thr Ala Glu Gly Ser Pro Ala Pro Ser Val Thr Trp
Asp Thr Glu 175 180 185 gtc aaa ggc aca acg tcc agc cgt tcc ttc aag
cac tcc cgc tct gct 869 Val Lys Gly Thr Thr Ser Ser Arg Ser Phe Lys
His Ser Arg Ser Ala 190 195 200 gcc gtc acc tca gag ttc cac ttg gtg
cct agc cgc agc atg aat ggg 917 Ala Val Thr Ser Glu Phe His Leu Val
Pro Ser Arg Ser Met Asn Gly 205 210 215 cag cca ctg act tgt gtg gtg
tcc cat cct ggc ctg ctc cag gac caa 965 Gln Pro Leu Thr Cys Val Val
Ser His Pro Gly Leu Leu Gln Asp Gln 220 225 230 agg atc acc cac atc
ctc cac gtg tcc ttc ctt gct gag gcc tct gtg 1013 Arg Ile Thr His
Ile Leu His Val Ser Phe Leu Ala Glu Ala Ser Val 235 240 245 250 agg
ggc ctt gaa gac caa aat ctg tgg cac att ggc aga gaa gga gct 1061
Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile Gly Arg Glu Gly Ala 255
260 265 atg ctc aag tgc ctg agt gaa ggg cag ccc cct ccc tca tac aac
tgg 1109 Met Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro Pro Ser Tyr
Asn Trp 270 275 280 aca cgg ctg gat ggg cct ctg ccc agt ggg gta cga
gtg gat ggg gac 1157 Thr Arg Leu Asp Gly Pro Leu Pro Ser Gly Val
Arg Val Asp Gly Asp 285 290 295 act ttg ggc ttt ccc cca ctg acc act
gag cac agc ggc atc tac gtc 1205 Thr Leu Gly Phe Pro Pro Leu Thr
Thr Glu His Ser Gly Ile Tyr Val 300 305 310 tgc cat gtc agc aat gag
ttc tcc tca agg gat tct cag gtc act gtg 1253 Cys His Val Ser Asn
Glu Phe Ser Ser Arg Asp Ser Gln Val Thr Val 315 320 325 330 gat gtt
ctt gac ccc cag gaa gac tct ggg
aag cag gtg gac cta gtg 1301 Asp Val Leu Asp Pro Gln Glu Asp Ser
Gly Lys Gln Val Asp Leu Val 335 340 345 tca gcc tcg gtg gtg gtg gtg
ggt gtg atc gcc gca ctc ttg ttc tgc 1349 Ser Ala Ser Val Val Val
Val Gly Val Ile Ala Ala Leu Leu Phe Cys 350 355 360 ctt ctg gtg gtg
gtg gtg gtg ctc atg tcc cga tac cat cgg cgc aag 1397 Leu Leu Val
Val Val Val Val Leu Met Ser Arg Tyr His Arg Arg Lys 365 370 375 gcc
cag cag atg acc cag aaa tat gag gag gag ctg acc ctg acc agg 1445
Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg 380
385 390 gag aac tcc atc cgg agg ctg cat tcc cat cac acg gac ccc agg
agc 1493 Glu Asn Ser Ile Arg Arg Leu His Ser His His Thr Asp Pro
Arg Ser 395 400 405 410 cag ccg gag gag agt gta ggg ctg aga gcc gag
ggc cac cct gat agt 1541 Gln Pro Glu Glu Ser Val Gly Leu Arg Ala
Glu Gly His Pro Asp Ser 415 420 425 ctc aag gac aac agt agc tgc tct
gtg atg agt gaa gag ccc gag ggc 1589 Leu Lys Asp Asn Ser Ser Cys
Ser Val Met Ser Glu Glu Pro Glu Gly 430 435 440 cgc agt tac tcc acg
ctg acc acg gtg agg gag ata gaa aca cag act 1637 Arg Ser Tyr Ser
Thr Leu Thr Thr Val Arg Glu Ile Glu Thr Gln Thr 445 450 455 gaa ctg
ctg tct cca ggc tct ggg cgg gcc gag gag gag gaa gat cag 1685 Glu
Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln 460 465
470 gat gaa ggc atc aaa cag gcc atg aac cat ttt gtt cag gag aat ggg
1733 Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val Gln Glu Asn
Gly 475 480 485 490 acc cta cgg gcc aag ccc acg ggc aat ggc atc tac
atc aat ggg cgg 1781 Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile
Tyr Ile Asn Gly Arg 495 500 505 gga cac ctg gtc tga cccaggcctg
cctcccttcc ctaggcctgg ctccttctgt 1836 Gly His Leu Val * 510
tgacatggga gattttagct catcttgggg gcctccttaa acacccccat ttcttgcgga
1896 agatgctccc catcccactg actgcttgac ctttacctcc aacccttctg
ttcatcggga 1956 gggctccacc aattgagtct ctcccaccat gcatgcaggt
cactgtgtgt gtgcatgtgt 2016 gcctgtgtga gtgttgactg actgtgtgtg
tgtggagggg tgactgtccg tggaggggtg 2076 actgtgtccg tggtgtgtat
tatgctgtca tatcagagtc aagtgaactg tggtgtatgt 2136 gccacgggat
ttgagtggtt gcgtgggcaa cactgtcagg gtttggcgtg tgtgtcatgt 2196
ggctgtgtgt gacctctgcc tgaaaaagca ggtattttct cagaccccag agcagtatta
2256 atgatgcaga ggttggagga gagaggtgga gactgtggct cagacccagg
tgtgcgggca 2316 tagctggagc tggaatctgc ctccggtgtg agggaacctg
tctcctacca cttcggagcc 2376 atgggggcaa gtgtgaagca gccagtccct
gggtcagcca gaggcttgaa ctgttacaga 2436 agccctctgc cctctggtgg
cctctgggcc tgctgcatgt acatattttc tgtaaatata 2496 catgcgccgg
gagcttcttg caggaatact gctccgaatc acttttaatt tttttctttt 2556
ttttttcttg ccctttccat tagttgtatt ttttatttat ttttattttt attttttttt
2616 agagatggag tctcactatg ttgctcaggc tggccttgaa ctcctgggct
caagcaatcc 2676 tcctgcctca gactccctag tagctgggac tttaagtgta
caccactgtg cctgctttga 2736 atcctttacg aagagaaaaa aaaaattaaa
gaaagccttt agatttatcc aatgtttact 2796 actgggattg cttaaagtga
ggcccctcca acaccagggg gttaattcct gtgattgtga 2856 aaggggctac
ttccaaggca tcttcatgca ggcagcccct tgggagggca cctgagagct 2916
ggtagagtct gaaattaggg atgtgagcct cgtggttact gagtaaggta aaattgcatc
2976 caccattgtt tgtgatacct tagggaattg cttggacctg gtgacaaggg
ctcctgttca 3036 atagtggtgt tggggagaga gagagcagtg attatagacc
gagagagtag gagttgaggt 3096 gaggtgaagg aggtgctggg ggtgagaatg
tcgcctttcc ccctgggttt tggatcacta 3156 attcaaggct cttctggatg
tttctctggg ttggggctgg agttcaatga ggtttatttt 3216 tagctggccc
acccagatac actcagccag aatacctaga tttagtaccc aaactcttct 3276
tagtctgaaa tctgctggat ttctggccta agggagaggc tcccatcctt cgttccccag
3336 ccagcctagg acttcgaatg tggagcctga agatctaaga tcctaacatg
tacattttat 3396 gtaaatatgt gcatatttgt acataaaatg atattctgtt
tttaaataaa cagacaaaac 3456 ttgaaaaa 3464 11 510 PRT Homo sapiens 11
Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5
10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala
Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly
Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly
Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly
Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly
Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro
Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg
Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val
Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135
140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu
145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala
Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val
Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser
Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser
Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly
Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235 240 His Val
Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255
Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260
265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly
Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly
Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys
His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val
Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln
Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val Gly Val Ile
Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360 365 Val Leu
Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380
Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385
390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln Pro Glu Glu
Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp Ser Leu Lys
Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu Pro Glu Gly
Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu Ile Glu Thr
Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg Ala Glu Glu
Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480 Ala Met Asn
His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485 490 495 Thr
Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500 505 510 12
3344 DNA Homo sapiens CDS (789)...(1676) 12 ggccgtcgtt gttggccaca
gcgtgggaag cagctctggg ggagctcgga gctcccgatc 60 acggcttctt
gggggtagct acggctgggt gtgtagaacg gggccggggc tggggctggg 120
tcccctagtg gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt
180 cagttcctta ttcaagtctg ctactgctgg catcatttac aggccggtgc
cccgcgggtg 240 agctggagac ctcagacgtg gtaactgtgg tgctgggcca
ggacgcaaaa ctgccctgct 300 tctaccgagg ggactccggc gagcaagtgg
ggcaagtggc atgggctcgg gtggacgcgg 360 gcgaaggcgc ccaggaacta
gcgctactgc actccaaata cgggcttcat gtgagcccgg 420 cttacgaggg
ccgcgtggag cagccgccgc ccccacgcaa ccccctggac ggctcagtgc 480
tcctgcgcaa cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc agcaccttcc
540 ccgccggcag cttccaggcg cggctgcggc tccgagtgct ggtgcctccc
ctgccctcac 600 tgaatcctgg tccagcacta gaagagggcc agggcctgac
cctggcagcc tcctgcacag 660 ctgagggcag cccagccccc agcgtgacct
gggacacgga ggtcaaaggc acaacgtcca 720 gccgttcctt caagcactcc
cgctctgctg ccgtcacctc agagttccac ttggtgccta 780 gccgcagc atg aat
ggg cag cca ctg act tgt gtg gtg tcc cat cct ggc 830 Met Asn Gly Gln
Pro Leu Thr Cys Val Val Ser His Pro Gly 1 5 10 ctg ctc cag gac caa
agg atc acc cac atc ctc cac gtg tcc ttc ctt 878 Leu Leu Gln Asp Gln
Arg Ile Thr His Ile Leu His Val Ser Phe Leu 15 20 25 30 gct gag gcc
tct gtg agg ggc ctt gaa gac caa aat ctg tgg cac att 926 Ala Glu Ala
Ser Val Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile 35 40 45 ggc
aga gaa gga gct atg ctc aag tgc ctg agt gaa ggg cag ccc cct 974 Gly
Arg Glu Gly Ala Met Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro 50 55
60 ccc tca tac aac tgg aca cgg ctg gat ggg cct ctg ccc agt ggg gta
1022 Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro Leu Pro Ser Gly
Val 65 70 75 cga gtg gat ggg gac act ttg ggc ttt ccc cca ctg acc
act gag cac 1070 Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro Leu
Thr Thr Glu His 80 85 90 agc ggc atc tac gtc tgc cat gtc agc aat
gag ttc tcc tca agg gat 1118 Ser Gly Ile Tyr Val Cys His Val Ser
Asn Glu Phe Ser Ser Arg Asp 95 100 105 110 tct cag gtc act gtg gat
gtt ctt gac ccc cag gaa gac tct ggg aag 1166 Ser Gln Val Thr Val
Asp Val Leu Asp Pro Gln Glu Asp Ser Gly Lys 115 120 125 cag gtg gac
cta gtg tca gcc tcg gtg gtg gtg gtg ggt gtg atc gcc 1214 Gln Val
Asp Leu Val Ser Ala Ser Val Val Val Val Gly Val Ile Ala 130 135 140
gca ctc ttg ttc tgc ctt ctg gtg gtg gtg gtg gtg ctc atg tcc cga
1262 Ala Leu Leu Phe Cys Leu Leu Val Val Val Val Val Leu Met Ser
Arg 145 150 155 tac cat cgg cgc aag gcc cag cag atg acc cag aaa tat
gag gag gag 1310 Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln Lys
Tyr Glu Glu Glu 160 165 170 ctg acc ctg acc agg gag aac tcc atc cgg
agg ctg cat tcc cat cac 1358 Leu Thr Leu Thr Arg Glu Asn Ser Ile
Arg Arg Leu His Ser His His 175 180 185 190 acg gac ccc agg agc cag
ccg gag gag agt gta ggg ctg aga gcc gag 1406 Thr Asp Pro Arg Ser
Gln Pro Glu Glu Ser Val Gly Leu Arg Ala Glu 195 200 205 ggc cac cct
gat agt ctc aag gac aac agt agc tgc tct gtg atg agt 1454 Gly His
Pro Asp Ser Leu Lys Asp Asn Ser Ser Cys Ser Val Met Ser 210 215 220
gaa gag ccc gag ggc cgc agt tac tcc acg ctg acc acg gtg agg gag
1502 Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg
Glu 225 230 235 ata gaa aca cag act gaa ctg ctg tct cca ggc tct ggg
cgg gcc gag 1550 Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly Ser
Gly Arg Ala Glu 240 245 250 gag gag gaa gat cag gat gaa ggc atc aaa
cag gcc atg aac cat ttt 1598 Glu Glu Glu Asp Gln Asp Glu Gly Ile
Lys Gln Ala Met Asn His Phe 255 260 265 270 gtt cag gag aat ggg acc
cta cgg gcc aag ccc acg ggc aat ggc atc 1646 Val Gln Glu Asn Gly
Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile 275 280 285 tac atc aat
ggg cgg gga cac ctg gtc tga cccaggcctg cctcccttcc 1696 Tyr Ile Asn
Gly Arg Gly His Leu Val * 290 295 ctaggcctgg ctccttctgt tgacatggga
gattttagct catcttgggg gcctccttaa 1756 acacccccat ttcttgcgga
agatgctccc catcccactg actgcttgac ctttacctcc 1816 aacccttctg
ttcatcggga gggctccacc aattgagtct ctcccaccat gcatgcaggt 1876
cactgtgtgt gtgcatgtgt gcctgtgtga gtgttgactg actgtgtgtg tgtggagggg
1936 tgactgtccg tggaggggtg actgtgtccg tggtgtgtat tatgctgtca
tatcagagtc 1996 aagtgaactg tggtgtatgt gccacgggat ttgagtggtt
gcgtgggcaa cactgtcagg 2056 gtttggcgtg tgtgtcatgt ggctgtgtgt
gacctctgcc tgaaaaagca ggtattttct 2116 cagaccccag agcagtatta
atgatgcaga ggttggagga gagaggtgga gactgtggct 2176 cagacccagg
tgtgcgggca tagctggagc tggaatctgc ctccggtgtg agggaacctg 2236
tctcctacca cttcggagcc atgggggcaa gtgtgaagca gccagtccct gggtcagcca
2296 gaggcttgaa ctgttacaga agccctctgc cctctggtgg cctctgggcc
tgctgcatgt 2356 acatattttc tgtaaatata catgcgccgg gagcttcttg
caggaatact gctccgaatc 2416 acttttaatt tttttctttt ttttttcttg
ccctttccat tagttgtatt ttttatttat 2476 ttttattttt attttttttt
agagatggag tctcactatg ttgctcaggc tggccttgaa 2536 ctcctgggct
caagcaatcc tcctgcctca gcctccctag tagctgggac tttaagtgta 2596
caccactgtg cctgctttga atcctttacg aagagaaaaa aaaaattaaa gaaagccttt
2656 agatttatcc aatgtttact actgggattg cttaaagtga ggcccctcca
acaccagggg 2716 gttaattcct gtgattgtga aaggggctac ttccaaggca
tcttcatgca ggcagcccct 2776 tgggagggca cctgagagct ggtagagtct
gaaattaggg atgtgagcct cgtggttact 2836 gagtaaggta aaattgcatc
caccattgtt tgtgatacct tagggaattg cttggacctg 2896 gtgacaaggg
ctcctgttca atagtggtgt tggggagaga gagagcagtg attatagacc 2956
gagagagtag gagttgaggt gaggtgaagg aggtgctggg ggtgagaatg tcgcctttcc
3016 ccctgggttt tggatcacta attcaaggct cttctggatg tttctctggg
ttggggctgg 3076 agttcaatga ggtttatttt tagctggccc acccagatac
actcagccag aatacctaga 3136 tttagtaccc aaactcttct tagtctgaaa
tctgctggat ttctggccta agggagaggc 3196 tcccatcctt cgttccccag
ccagcctagg acttcgaatg tggagcctga agatctaaga 3256 tcctaacatg
tacattttat gtaaatatgt gcatatttgt acataaaatg atattctgtt 3316
tttaaataaa cagacaaaac ttgaaaaa 3344 13 295 PRT Homo sapiens 13 Met
Asn Gly Gln Pro Leu Thr Cys Val Val Ser His Pro Gly Leu Leu 1 5 10
15 Gln Asp Gln Arg Ile Thr His Ile Leu His Val Ser Phe Leu Ala Glu
20 25 30 Ala Ser Val Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile
Gly Arg 35 40 45 Glu Gly Ala Met Leu Lys Cys Leu Ser Glu Gly Gln
Pro Pro Pro Ser 50 55 60 Tyr Asn Trp Thr Arg Leu Asp Gly Pro Leu
Pro Ser Gly Val Arg Val 65 70 75 80 Asp Gly Asp Thr Leu Gly Phe Pro
Pro Leu Thr Thr Glu His Ser Gly 85 90 95 Ile Tyr Val Cys His Val
Ser Asn Glu Phe Ser Ser Arg Asp Ser Gln 100 105 110 Val Thr Val Asp
Val Leu Asp Pro Gln Glu Asp Ser Gly Lys Gln Val 115 120 125 Asp Leu
Val Ser Ala Ser Val Val Val Val Gly Val Ile Ala Ala Leu 130 135 140
Leu Phe Cys Leu Leu Val Val Val Val Val Leu Met Ser Arg Tyr His 145
150 155 160 Arg Arg Lys Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu
Leu Thr 165 170 175 Leu Thr Arg Glu Asn Ser Ile Arg Arg Leu His Ser
His His Thr Asp 180 185 190 Pro Arg Ser Gln Pro Glu Glu Ser Val Gly
Leu Arg Ala Glu Gly His 195 200 205 Pro Asp Ser Leu Lys Asp Asn Ser
Ser Cys Ser Val Met Ser Glu Glu 210 215 220 Pro Glu Gly Arg Ser Tyr
Ser Thr Leu Thr Thr Val Arg Glu Ile Glu 225 230 235 240 Thr Gln Thr
Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu 245 250 255 Glu
Asp Gln Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val Gln 260 265
270 Glu Asn Gly Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile Tyr Ile
275 280 285 Asn Gly Arg Gly His Leu Val 290 295 14 3389 DNA Homo
sapiens CDS (264)...(1721) 14 ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac
240 gctgggcagt ctgcctttca acc atg ccc ctg tcc ctg gga gcc gag atg
tgg 293 Met Pro Leu Ser Leu Gly Ala Glu Met Trp 1 5 10 ggg cct gag
gcc tgg ctg ctg ctg ctg cta ctg ctg gca tca ttt aca 341 Gly Pro Glu
Ala Trp Leu Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr 15 20 25 ggc
cgg tgc ccc gcg ggt gag ctg gag acc tca gac gtg gta act gtg 389 Gly
Arg Cys Pro Ala Gly Glu Leu Glu Thr Ser Asp Val Val Thr Val 30 35
40 gtg ctg ggc cag gac gca aaa ctg ccc tgc ttc tac cga ggg gac tcc
437 Val Leu Gly Gln Asp Ala Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser
45 50 55 ggc gag caa gtg ggg caa gtg gca tgg gct cgg gtg gac gcg
ggc gaa 485 Gly Glu Gln Val Gly Gln Val Ala Trp Ala Arg Val Asp Ala
Gly Glu 60 65
70 ggc gcc cag gaa cta gcg cta ctg cac tcc aaa tac ggg ctt cat gtg
533 Gly Ala Gln Glu Leu Ala Leu Leu His Ser Lys Tyr Gly Leu His Val
75 80 85 90 agc ccg gct tac gag ggc cgc gtg gag cag ccg ccg ccc cca
cgc aac 581 Ser Pro Ala Tyr Glu Gly Arg Val Glu Gln Pro Pro Pro Pro
Arg Asn 95 100 105 ccc ctg gac ggc tca gtg ctc ctg cgc aac gca gtg
cag gcg gat gag 629 Pro Leu Asp Gly Ser Val Leu Leu Arg Asn Ala Val
Gln Ala Asp Glu 110 115 120 ggc gag tac gag tgc cgg gtc agc acc ttc
ccc gcc ggc agc ttc cag 677 Gly Glu Tyr Glu Cys Arg Val Ser Thr Phe
Pro Ala Gly Ser Phe Gln 125 130 135 gcg cgg ctg cgg ctc cga gtg ctg
gtg cct ccc ctg ccc tca ctg aat 725 Ala Arg Leu Arg Leu Arg Val Leu
Val Pro Pro Leu Pro Ser Leu Asn 140 145 150 cct ggt cca gca cta gaa
gag ggc cag ggc ctg acc ctg gca gcc tcc 773 Pro Gly Pro Ala Leu Glu
Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser 155 160 165 170 tgc aca gct
gag ggc agc cca gcc ccc agc gtg acc tgg gac acg gag 821 Cys Thr Ala
Glu Gly Ser Pro Ala Pro Ser Val Thr Trp Asp Thr Glu 175 180 185 gtc
aaa ggc aca acg tcc agc cgt tcc ttc aag cac tcc cgc tct gct 869 Val
Lys Gly Thr Thr Ser Ser Arg Ser Phe Lys His Ser Arg Ser Ala 190 195
200 gcc gtc acc tca gag ttc cac ttg gtg cct agc cgc agc atg aat ggg
917 Ala Val Thr Ser Glu Phe His Leu Val Pro Ser Arg Ser Met Asn Gly
205 210 215 cag cca ctg act tgt gtg gtg tcc cat cct ggc ctg ctc cag
gac caa 965 Gln Pro Leu Thr Cys Val Val Ser His Pro Gly Leu Leu Gln
Asp Gln 220 225 230 agg atc acc cac atc ctc cac gtg tcc ttc ctt gct
gag gcc tct gtg 1013 Arg Ile Thr His Ile Leu His Val Ser Phe Leu
Ala Glu Ala Ser Val 235 240 245 250 agg ggc ctt gaa gac caa aat ctg
tgg cac att ggc aga gaa gga gct 1061 Arg Gly Leu Glu Asp Gln Asn
Leu Trp His Ile Gly Arg Glu Gly Ala 255 260 265 atg ctc aag tgc ctg
agt gaa ggg cag ccc cct ccc tca tac aac tgg 1109 Met Leu Lys Cys
Leu Ser Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp 270 275 280 aca cgg
ctg gat ggg cct ctg ccc agt ggg gta cga gtg gat ggg gac 1157 Thr
Arg Leu Asp Gly Pro Leu Pro Ser Gly Val Arg Val Asp Gly Asp 285 290
295 act ttg ggc ttt ccc cca ctg acc act gag cac agc ggc atc tac gtc
1205 Thr Leu Gly Phe Pro Pro Leu Thr Thr Glu His Ser Gly Ile Tyr
Val 300 305 310 tgc cat gtc agc aat gag ttc tcc tca agg gat tct cag
gtc act gtg 1253 Cys His Val Ser Asn Glu Phe Ser Ser Arg Asp Ser
Gln Val Thr Val 315 320 325 330 gat gtt ctt gac ccc cag gaa gac tct
ggg aag cag gtg gac cta gtg 1301 Asp Val Leu Asp Pro Gln Glu Asp
Ser Gly Lys Gln Val Asp Leu Val 335 340 345 tca gcc tcg gtg gtg gtg
gtg ggt gtg atc gcc gca ctc ttg ttc tgc 1349 Ser Ala Ser Val Val
Val Val Gly Val Ile Ala Ala Leu Leu Phe Cys 350 355 360 ctt ctg gtg
gtg gtg gtg gtg ctc atg tcc cga tac cat cgg cgc aag 1397 Leu Leu
Val Val Val Val Val Leu Met Ser Arg Tyr His Arg Arg Lys 365 370 375
gcc cag cag atg acc cag aaa tat gag gag gag ctg acc ctg acc agg
1445 Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu Thr Leu Thr
Arg 380 385 390 gag aac tcc atc cgg agg ctg cat tcc cat cac acg gac
ccc agg agc 1493 Glu Asn Ser Ile Arg Arg Leu His Ser His His Thr
Asp Pro Arg Ser 395 400 405 410 cag agt gaa gag ccc gag ggc cgc agt
tac tcc acg ctg acc acg gtg 1541 Gln Ser Glu Glu Pro Glu Gly Arg
Ser Tyr Ser Thr Leu Thr Thr Val 415 420 425 agg gag ata gaa aca cag
act gaa ctg ctg tct cca ggc tct ggg cgg 1589 Arg Glu Ile Glu Thr
Gln Thr Glu Leu Leu Ser Pro Gly Ser Gly Arg 430 435 440 gcc gag gag
gag gaa gat cag gat gaa ggc atc aaa cag gcc atg aac 1637 Ala Glu
Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln Ala Met Asn 445 450 455
cat ttt gtt cag gag aat ggg acc cta cgg gcc aag ccc acg ggc aat
1685 His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro Thr Gly
Asn 460 465 470 ggc atc tac atc aat ggg cgg gga cac ctg gtc tga
cccaggcctg 1731 Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val * 475
480 485 cctcccttcc ctaggcctgg ctccttctgt tgacatggga gattttagct
catcttgggg 1791 gcctccttaa acacccccat ttcttgcgga agatgctccc
catcccactg actgcttgac 1851 ctttacctcc aacccttctg ttcatcggga
gggctccacc aattgagtct ctcccaccat 1911 gcatgcaggt cactgtgtgt
gtgcatgtgt gcctgtgtga gtgttgactg actgtgtgtg 1971 tgtggagggg
tgactgtccg tggaggggtg actgtgtccg tggtgtgtat tatgctgtca 2031
tatcagagtc aagtgaactg tggtgtatgt gccacgggat ttgagtggtt gcgtgggcaa
2091 cactgtcagg gtttggcgtg tgtgtcatgt ggctgtgtgt gacctctgcc
tgaaaaagca 2151 ggtattttct cagaccccag agcagtatta atgatgcaga
ggttggagga gagaggtgga 2211 gactgtggct cagacccagg tgtgcgggca
tagctggagc tggaatctgc ctccggtgtg 2271 agggaacctg tctcctacca
cttcggagcc atgggggcaa gtgtgaagca gccagtccct 2331 gggtcagcca
gaggcttgaa ctgttacaga agccctctgc cctctggtgg cctctgggcc 2391
tgctgcatgt acatattttc tgtaaatata catgcgccgg gagcttcttg caggaatact
2451 gctccgaatc acttttaatt tttttctttt ttttttcttg ccctttccat
tagttgtatt 2511 ttttatttat ttttattttt attttttttt agagatggag
tctcactatg ttgctcaggc 2571 tggccttgaa ctcctgggct caagcaatcc
tcctgcctca gcctccctag tagctgggac 2631 tttaagtgta caccactgtg
cctgctttga atcctttacg aagagaaaaa aaaaattaaa 2691 gaaagccttt
agatttatcc aatgtttact actgggattg cttaaagtga ggcccctcca 2751
acaccagggg gttaattcct gtgattgtga aaggggctac ttccaaggca tcttcatgca
2811 ggcagcccct tgggagggca cctgagagct ggtagagtct gaaattaggg
atgtgagcct 2871 cgtggttact gagtaaggta aaattgcatc caccattgtt
tgtgatacct tagggaattg 2931 cttggacctg gtgacaaggg ctcctgttca
atagtggtgt tggggagaga gagagcagtg 2991 attatagacc gagagagtag
gagttgaggt gaggtgaagg aggtgctggg ggtgagaatg 3051 tcgcctttcc
ccctgggttt tggatcacta attcaaggct cttctggatg tttctctggg 3111
ttggggctgg agttcaatga ggtttatttt tagctggccc acccagatac actcagccag
3171 aatacctaga tttagtaccc aaactcttct tagtctgaaa tctgctggat
ttctggccta 3231 agggagaggc tcccatcctt cgttccccag ccagcctagg
acttcgaatg tggagcctga 3291 agatctaaga tcctaacatg tacattttat
gtaaatatgt gcatatttgt acataaaatg 3351 atattctgtt tttaaataaa
cagacaaaac ttgaaaaa 3389 15 485 PRT Homo sapiens 15 Met Pro Leu Ser
Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu
Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30
Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35
40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly
Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln
Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser
Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg
Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln
Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro
Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val
Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160
Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165
170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr
Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr
Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln
Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp
Gln Arg Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala
Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His
Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly
Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285
Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290
295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn
Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val
Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val
Ser Ala Ser Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu
Phe Cys Leu Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr
His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu
Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu
His Ser His His Thr Asp Pro Arg Ser Gln Ser Glu Glu Pro Glu 405 410
415 Gly Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr Gln
420 425 430 Thr Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu
Glu Asp 435 440 445 Gln Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe
Val Gln Glu Asn 450 455 460 Gly Thr Leu Arg Ala Lys Pro Thr Gly Asn
Gly Ile Tyr Ile Asn Gly 465 470 475 480 Arg Gly His Leu Val 485 16
3401 DNA Homo sapiens CDS (264)...(1796) 16 ggccgtcgtt gttggccaca
gcgtgggaag cagctctggg ggagctcgga gctcccgatc 60 acggcttctt
gggggtagct acggctgggt gtgtagaacg gggccggggc tggggctggg 120
tcccctagtg gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt
180 cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa
cttcagaaac 240 gctgggcagt ctgcctttca acc atg ccc ctg tcc ctg gga
gcc gag atg tgg 293 Met Pro Leu Ser Leu Gly Ala Glu Met Trp 1 5 10
ggg cct gag gcc tgg ctg ctg ctg ctg cta ctg ctg gca tca ttt aca 341
Gly Pro Glu Ala Trp Leu Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr 15
20 25 ggc cgg tgc ccc gcg ggt gag ctg gag acc tca gac gtg gta act
gtg 389 Gly Arg Cys Pro Ala Gly Glu Leu Glu Thr Ser Asp Val Val Thr
Val 30 35 40 gtg ctg ggc cag gac gca aaa ctg ccc tgc ttc tac cga
ggg gac tcc 437 Val Leu Gly Gln Asp Ala Lys Leu Pro Cys Phe Tyr Arg
Gly Asp Ser 45 50 55 ggc gag caa gtg ggg caa gtg gca tgg gct cgg
gtg gac gcg ggc gaa 485 Gly Glu Gln Val Gly Gln Val Ala Trp Ala Arg
Val Asp Ala Gly Glu 60 65 70 ggc gcc cag gaa cta gcg cta ctg cac
tcc aaa tac ggg ctt cat gtg 533 Gly Ala Gln Glu Leu Ala Leu Leu His
Ser Lys Tyr Gly Leu His Val 75 80 85 90 agc ccg gct tac gag ggc cgc
gtg gag cag ccg ccg ccc cca cgc aac 581 Ser Pro Ala Tyr Glu Gly Arg
Val Glu Gln Pro Pro Pro Pro Arg Asn 95 100 105 ccc ctg gac ggc tca
gtg ctc ctg cgc aac gca gtg cag gcg gat gag 629 Pro Leu Asp Gly Ser
Val Leu Leu Arg Asn Ala Val Gln Ala Asp Glu 110 115 120 ggc gag tac
gag tgc cgg gtc agc acc ttc ccc gcc ggc agc ttc cag 677 Gly Glu Tyr
Glu Cys Arg Val Ser Thr Phe Pro Ala Gly Ser Phe Gln 125 130 135 gcg
cgg ctg cgg ctc cga gtg ctg gtg cct ccc ctg ccc tca ctg aat 725 Ala
Arg Leu Arg Leu Arg Val Leu Val Pro Pro Leu Pro Ser Leu Asn 140 145
150 cct ggt cca gca cta gaa gag ggc cag ggc ctg acc ctg gca gcc tcc
773 Pro Gly Pro Ala Leu Glu Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser
155 160 165 170 tgc aca gct gag ggc agc cca gcc ccc agc gtg acc tgg
gac acg gag 821 Cys Thr Ala Glu Gly Ser Pro Ala Pro Ser Val Thr Trp
Asp Thr Glu 175 180 185 gtc aaa ggc aca acg tcc agc cgt tcc ttc aag
cac tcc cgc tct gct 869 Val Lys Gly Thr Thr Ser Ser Arg Ser Phe Lys
His Ser Arg Ser Ala 190 195 200 gcc gtc acc tca gag ttc cac ttg gtg
cct agc cgc agc atg aat ggg 917 Ala Val Thr Ser Glu Phe His Leu Val
Pro Ser Arg Ser Met Asn Gly 205 210 215 cag cca ctg act tgt gtg gtg
tcc cat cct ggc ctg ctc cag gac caa 965 Gln Pro Leu Thr Cys Val Val
Ser His Pro Gly Leu Leu Gln Asp Gln 220 225 230 agg atc acc cac atc
ctc cac gtg tcc ttc ctt gct gag gcc tct gtg 1013 Arg Ile Thr His
Ile Leu His Val Ser Phe Leu Ala Glu Ala Ser Val 235 240 245 250 agg
ggc ctt gaa gac caa aat ctg tgg cac att ggc aga gaa gga gct 1061
Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile Gly Arg Glu Gly Ala 255
260 265 atg ctc aag tgc ctg agt gaa ggg cag ccc cct ccc tca tac aac
tgg 1109 Met Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro Pro Ser Tyr
Asn Trp 270 275 280 aca cgg ctg gat ggg cct ctg ccc agt ggg gta cga
gtg gat ggg gac 1157 Thr Arg Leu Asp Gly Pro Leu Pro Ser Gly Val
Arg Val Asp Gly Asp 285 290 295 act ttg ggc ttt ccc cca ctg acc act
gag cac agc ggc atc tac gtc 1205 Thr Leu Gly Phe Pro Pro Leu Thr
Thr Glu His Ser Gly Ile Tyr Val 300 305 310 tgc cat gtc agc aat gag
ttc tcc tca agg gat tct cag gtc act gtg 1253 Cys His Val Ser Asn
Glu Phe Ser Ser Arg Asp Ser Gln Val Thr Val 315 320 325 330 gat gtt
ctt gac ccc cag gaa gac tct ggg aag cag gtg gac cta gtg 1301 Asp
Val Leu Asp Pro Gln Glu Asp Ser Gly Lys Gln Val Asp Leu Val 335 340
345 tca gcc tcg gtg gtg gtg gtg ggt gtg atc gcc gca ctc ttg ttc tgc
1349 Ser Ala Ser Val Val Val Val Gly Val Ile Ala Ala Leu Leu Phe
Cys 350 355 360 ctt ctg gtg gtg gtg gtg gtg ctc atg tcc cga tac cat
cgg cgc aag 1397 Leu Leu Val Val Val Val Val Leu Met Ser Arg Tyr
His Arg Arg Lys 365 370 375 gcc cag cag atg acc cag aaa tat gag gag
gag ctg acc ctg acc agg 1445 Ala Gln Gln Met Thr Gln Lys Tyr Glu
Glu Glu Leu Thr Leu Thr Arg 380 385 390 gag aac tcc atc cgg agg ctg
cat tcc cat cac acg gac ccc agg agc 1493 Glu Asn Ser Ile Arg Arg
Leu His Ser His His Thr Asp Pro Arg Ser 395 400 405 410 cag ccg gag
gag agt gta ggg ctg aga gcc gag ggc cac cct gat agt 1541 Gln Pro
Glu Glu Ser Val Gly Leu Arg Ala Glu Gly His Pro Asp Ser 415 420 425
ctc aag gac aac agt agc tgc tct gtg atg agt gaa gag ccc gag ggc
1589 Leu Lys Asp Asn Ser Ser Cys Ser Val Met Ser Glu Glu Pro Glu
Gly 430 435 440 cgc agt tac tcc acg ctg acc acg gtg agg gag ata gaa
aca cag act 1637 Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile
Glu Thr Gln Thr 445 450 455 gaa ctg ctg tct cca ggc tct ggg cgg gcc
gag gag gag gaa gat cag 1685 Glu Leu Leu Ser Pro Gly Ser Gly Arg
Ala Glu Glu Glu Glu Asp Gln 460 465 470 gat gaa ggc atc aaa cag gcc
atg aac cat ttt gtt cag gag aat ggg 1733 Asp Glu Gly Ile Lys Gln
Ala Met Asn His Phe Val Gln Glu Asn Gly 475 480 485 490 acc cta cgg
gcc aag ccc acg ggc aat ggc atc tac atc aat ggg cgg 1781 Thr Leu
Arg Ala Lys Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg 495 500 505
gga cac ctg gtc tga cccaggcctg cctcccttcc ctaggcctgg ctccttctgt
1836 Gly His Leu Val * 510 tgacatggga gattttagct catcttgggg
gcctccttaa acacccccat ttcttgcgga 1896 agatgctccc catcccactg
actgcttgac ctttacctcc aacccttctg ttcatcggga 1956 gggctccacc
aattgagtct ctcccaccat gcatgcaggt cactgtgtgt gtgcatgtgt 2016
gcctgtgtga gtgttgactg actgtgtgtg tgtggagggg tgactgtccg tggaggggtg
2076 actgtgtccg tggtgtgtat tatgctgtca tatcagagtc aagtgaactg
tggtgtatgt 2136 gccacgggat ttgagtggtt gcgtgggcaa cactgtcagg
gtttggcgtg tgtgtcatgt 2196 ggctgtgtgt gacctctgcc tgaaaaagca
ggtattttct cagaccccag agcagtatta 2256 atgatgcaga ggttggagga
gagaggtgga gactgtggct cagacccagg tgtgcgggca 2316 tagctggagc
tggaatctgc ctccggtgtg agggaacctg tctcctacca cttcggagcc 2376
atgggggcaa gtgtgaagca gccagtccct gggtcagcca gaggcttgaa ctgttacaga
2436 agccctctgc cctctggtgg cctctgggcc tgctgcatgt acatattttc
tgtaaatata 2496 catgcgccgg gagcttcttg caggaatact gctccgaatc
acttttaatt tttttctttt 2556 ttttttcttg ccctttccat tagttgtatt
ttttatttat ttttattttt attttttttt 2616 agagatggag tctcactatg
ttgctcaggc tggccttgaa ctcctgggct
caagcaatcc 2676 tcctgcctca gcctccctag tagctgggac tttaagtgta
caccactgtg cctgctttga 2736 atcctttacg aagagaaaaa aaaaattaaa
gaaagccttt agatttatcc aatgtttact 2796 actgggattg cttaaagtga
ggcccctcca acaccagggg gttaattcct gtgattgtga 2856 aaggggctac
ttccaaggca tcttcatgca ggcagcccct tgggagggca cctgagagct 2916
ggtagagtct gaaattaggg atgtgagcct cgtgctggtg acaagggctc ctgttcaata
2976 gtggtgttgg ggagagagag agcagtgatt atagaccgag agagtaggag
ttgaggtgag 3036 gtgaaggagg tgctgggggt gagaatgtcg cctttccccc
tgggttttgg atcactaatt 3096 caaggctctt ctggatgttt ctctgggttg
gggctggagt tcaatgaggt ttatttttag 3156 ctggcccacc cagatacact
cagccagaat acctagattt agtacccaaa ctcttcttag 3216 tctgaaatct
gctggatttc tggcctaagg gagaggctcc catccttcgt tccccagcca 3276
gcctaggact tcgaatgtgg agcctgaaga tctaagatcc taacatgtac attttatgta
3336 aatatgtgca tatttgtaca taaaatgata ttctgttttt aaataaacag
acaaaacttg 3396 aaaaa 3401 17 510 PRT Homo sapiens 17 Met Pro Leu
Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu
Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25
30 Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala
35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val
Gly Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala
Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val
Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro
Arg Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val
Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe
Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu
Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155
160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser
165 170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr
Thr Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val
Thr Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly
Gln Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln
Asp Gln Arg Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu
Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp
His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu
Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280
285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro
290 295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser
Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp
Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu
Val Ser Ala Ser Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu
Leu Phe Cys Leu Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg
Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu
Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400
Leu His Ser His His Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405
410 415 Gly Leu Arg Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser
Ser 420 425 430 Cys Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr
Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu
Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp
Gln Asp Glu Gly Ile Lys Gln 465 470 475 480 Ala Met Asn His Phe Val
Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly
Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500 505 510 18 1669 DNA
Homo sapiens CDS (708)...(1121) 18 gtctgaccca ggcctgcctc ccttccctag
gcctggctcc ttctgttgac atgggagatt 60 ttagctcatc ttgggggcct
ccttaaacac ccccatttct tgcggaagat gctccccatc 120 ccactgactg
cttgaccttt acctccaacc cttctgttca tcgggagggc tccaccaatt 180
gagtctctcc caccatgcat gcaggtcact gtgtgtgtgc atgtgtgcct gtgtgagtgt
240 tgactgactg tgtgtgtgtg gaggggtgac tgtccgtgga ggggtgactg
tgtccgtggt 300 gtgtattatg ctgtcatatc agagtcaagt gaactgtggt
gtatgtgcca cgggatttga 360 gtggttgcgt gggcaacact gtcagggttt
ggcgtgtgtg tcatgtggct gtgtgtgacc 420 tctgcctgaa aaagcaggta
ttttctcaga ccccagagca gtattaatga tgcagaggtt 480 ggaggagaga
ggtggagact gtggctcaga cccaggtgtg cgggcatagc tggagctgga 540
atctgcctcc ggtgtgaggg aacctgtctc ctaccacttc ggagccatgg gggcaagtgt
600 gaagcagcca gtccctgggt cagccagagg cttgaactgt tacagaagcc
ctctgccctc 660 tggtggcctc tgggcctgct gcatgtacat attttctgta aatatac
atg cgc cgg 716 Met Arg Arg 1 gag ctt ctt gca gga ata ctg ctc cga
atc act ttt aat ttt ttt ctt 764 Glu Leu Leu Ala Gly Ile Leu Leu Arg
Ile Thr Phe Asn Phe Phe Leu 5 10 15 ttt ttt ttc ttg ccc ttt cca tta
gtt gta ttt ttt att tat ttt tat 812 Phe Phe Phe Leu Pro Phe Pro Leu
Val Val Phe Phe Ile Tyr Phe Tyr 20 25 30 35 ttt tat ttt ttt tta gag
atg gag tct cac tat gtt gct cag gct ggc 860 Phe Tyr Phe Phe Leu Glu
Met Glu Ser His Tyr Val Ala Gln Ala Gly 40 45 50 ctt gaa ctc ctg
ggc tca agc aat cct cct gcc tca gcc tcc cta gta 908 Leu Glu Leu Leu
Gly Ser Ser Asn Pro Pro Ala Ser Ala Ser Leu Val 55 60 65 gct ggg
act tta agt gta cac cac tgt gcc tgc ttt gaa tcc ttt acg 956 Ala Gly
Thr Leu Ser Val His His Cys Ala Cys Phe Glu Ser Phe Thr 70 75 80
aag aga aaa aaa aaa tta aag aaa gcc ttt aga ttt atc caa tgt tta
1004 Lys Arg Lys Lys Lys Leu Lys Lys Ala Phe Arg Phe Ile Gln Cys
Leu 85 90 95 cta ctg gga ttg ctt aaa gtg agg ccc ctc caa cac cag
ggg gtt aat 1052 Leu Leu Gly Leu Leu Lys Val Arg Pro Leu Gln His
Gln Gly Val Asn 100 105 110 115 tcc tgt gat tgt gaa agg ggc tac ttc
caa ggc atc ttc atg cag gca 1100 Ser Cys Asp Cys Glu Arg Gly Tyr
Phe Gln Gly Ile Phe Met Gln Ala 120 125 130 gcc cct tgg gag ggc acc
tga gagctggtag agtctgaaat tagggatgtg 1151 Ala Pro Trp Glu Gly Thr *
135 agcctcgtgg ttactgagta aggtaaaatt gcatccacca ttgtttgtga
taccttaggg 1211 aattgcttgg acctggtgac aagggctcct gttcaatagt
ggtgttgggg agagagagag 1271 cagtgattat agaccgagag agtaggagtt
gaggtgaggt gaaggaggtg ctgggggtga 1331 gaatgtcgcc tttccccctg
ggttttggat cactaattca aggctcttct ggatgtttct 1391 ctgggttggg
gctggagttc aatgaggttt atttttagct ggcccaccca gatacactca 1451
gccagaatac ctagatttag tacccaaact cttcttagtc tgaaatctgc tggatttctg
1511 gcctaaggga gaggctccca tccttcgttc cccagccagc ctaggacttc
gaatgtggag 1571 cctgaagatc taagatccta acatgtacat tttatgtaaa
tatgtgcata tttgtacata 1631 aaatgatatt ctgtttttaa ataaacagac
aaaacttg 1669 19 137 PRT Homo sapiens 19 Met Arg Arg Glu Leu Leu
Ala Gly Ile Leu Leu Arg Ile Thr Phe Asn 1 5 10 15 Phe Phe Leu Phe
Phe Phe Leu Pro Phe Pro Leu Val Val Phe Phe Ile 20 25 30 Tyr Phe
Tyr Phe Tyr Phe Phe Leu Glu Met Glu Ser His Tyr Val Ala 35 40 45
Gln Ala Gly Leu Glu Leu Leu Gly Ser Ser Asn Pro Pro Ala Ser Ala 50
55 60 Ser Leu Val Ala Gly Thr Leu Ser Val His His Cys Ala Cys Phe
Glu 65 70 75 80 Ser Phe Thr Lys Arg Lys Lys Lys Leu Lys Lys Ala Phe
Arg Phe Ile 85 90 95 Gln Cys Leu Leu Leu Gly Leu Leu Lys Val Arg
Pro Leu Gln His Gln 100 105 110 Gly Val Asn Ser Cys Asp Cys Glu Arg
Gly Tyr Phe Gln Gly Ile Phe 115 120 125 Met Gln Ala Ala Pro Trp Glu
Gly Thr 130 135 20 3464 DNA Homo sapiens CDS (264)...(1796) 20
ggccgtcgtt gttggccaca gcgtgggaag cagctctggg ggagctcgga gctcccgatc
60 acggcttctt gggggtagct acggctgggt gtgtagaacg gggccggggc
tggggctggg 120 tcccctagtg gagacccaag tgcgagaggc aagaactctg
cagcttcctg ccttctgggt 180 cagttcctta ttcaagtctg cagccggctc
ccagggagat ctcggtggaa cttcagaaac 240 gctgggcagt ctgcctttca acc atg
ccc ctg tcc ctg gga gcc gag atg tgg 293 Met Pro Leu Ser Leu Gly Ala
Glu Met Trp 1 5 10 ggg cct gag gcc tgg ctg ctg ctg ctg cta ctg ctg
gca tca ttt aca 341 Gly Pro Glu Ala Trp Leu Leu Leu Leu Leu Leu Leu
Ala Ser Phe Thr 15 20 25 ggc cgg tgc ccc gcg ggt gag ctg ggg acc
tca gac gtg gta act gtg 389 Gly Arg Cys Pro Ala Gly Glu Leu Gly Thr
Ser Asp Val Val Thr Val 30 35 40 gtg ctg ggc cag gac gca aaa ctg
ccc tgc ttc tac cga ggg gac tcc 437 Val Leu Gly Gln Asp Ala Lys Leu
Pro Cys Phe Tyr Arg Gly Asp Ser 45 50 55 ggc gag caa gtg ggg caa
gtg gca tgg gct cgg gtg gac gcg ggc gaa 485 Gly Glu Gln Val Gly Gln
Val Ala Trp Ala Arg Val Asp Ala Gly Glu 60 65 70 ggc gcc cag gaa
cta gcg cta ctg cac tcc aaa tac ggg ctt cat gtg 533 Gly Ala Gln Glu
Leu Ala Leu Leu His Ser Lys Tyr Gly Leu His Val 75 80 85 90 agc ccg
gct tac gag ggc cgc gtg gag cag ccg ccg ccc cca cgc aac 581 Ser Pro
Ala Tyr Glu Gly Arg Val Glu Gln Pro Pro Pro Pro Arg Asn 95 100 105
ccc ctg gac ggc tca gtg ctc ctg cgc aac gca gtg cag gcg gat gag 629
Pro Leu Asp Gly Ser Val Leu Leu Arg Asn Ala Val Gln Ala Asp Glu 110
115 120 ggc gag tac gag tgc cgg gtc agc acc ttc ccc gcc ggc agc ttc
cag 677 Gly Glu Tyr Glu Cys Arg Val Ser Thr Phe Pro Ala Gly Ser Phe
Gln 125 130 135 gcg cgg ctg cgg ctc cga gtg ctg gtg cct ccc ctg ccc
tca ctg aat 725 Ala Arg Leu Arg Leu Arg Val Leu Val Pro Pro Leu Pro
Ser Leu Asn 140 145 150 cct ggt cca gca cta gaa gag ggc cag ggc ctg
acc ctg gca gcc tcc 773 Pro Gly Pro Ala Leu Glu Glu Gly Gln Gly Leu
Thr Leu Ala Ala Ser 155 160 165 170 tgc aca gct gag ggc agc cca gcc
ccc agc gtg acc tgg gac acg gag 821 Cys Thr Ala Glu Gly Ser Pro Ala
Pro Ser Val Thr Trp Asp Thr Glu 175 180 185 gtc aaa ggc aca acg tcc
agc cgt tcc ttc aag cac tcc cgc tct gct 869 Val Lys Gly Thr Thr Ser
Ser Arg Ser Phe Lys His Ser Arg Ser Ala 190 195 200 gcc gtc acc tca
gag ttc cac ttg gtg cct agc cgc agc atg aat ggg 917 Ala Val Thr Ser
Glu Phe His Leu Val Pro Ser Arg Ser Met Asn Gly 205 210 215 cag cca
ctg act tgt gtg gtg tcc cat cct ggc ctg ctc cag gac caa 965 Gln Pro
Leu Thr Cys Val Val Ser His Pro Gly Leu Leu Gln Asp Gln 220 225 230
agg atc acc cac atc ctc cac gtg tcc ttc ctt gct gag gcc tct gtg
1013 Arg Ile Thr His Ile Leu His Val Ser Phe Leu Ala Glu Ala Ser
Val 235 240 245 250 agg ggc ctt gaa gac caa aat ctg tgg cac att ggc
aga gaa gga gct 1061 Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile
Gly Arg Glu Gly Ala 255 260 265 atg ctc aag tgc ctg agt gaa ggg cag
ccc cct ccc tca tac aac tgg 1109 Met Leu Lys Cys Leu Ser Glu Gly
Gln Pro Pro Pro Ser Tyr Asn Trp 270 275 280 aca cgg ctg gat ggg cct
ctg ccc agt ggg gta cga gtg gat ggg gac 1157 Thr Arg Leu Asp Gly
Pro Leu Pro Ser Gly Val Arg Val Asp Gly Asp 285 290 295 act ttg ggc
ttt ccc cca ctg acc act gag cac agc ggc atc tac gtc 1205 Thr Leu
Gly Phe Pro Pro Leu Thr Thr Glu His Ser Gly Ile Tyr Val 300 305 310
tgc cat gtc agc aat gag ttc tcc tca agg gat tct cag gtc act gtg
1253 Cys His Val Ser Asn Glu Phe Ser Ser Arg Asp Ser Gln Val Thr
Val 315 320 325 330 gat gtt ctt gac ccc cag gaa gac tct ggg aag cag
gtg gac cta gtg 1301 Asp Val Leu Asp Pro Gln Glu Asp Ser Gly Lys
Gln Val Asp Leu Val 335 340 345 tca gcc tcg gtg gtg gtg gtg ggt gtg
atc gcc gca ctc ttg ttc tgc 1349 Ser Ala Ser Val Val Val Val Gly
Val Ile Ala Ala Leu Leu Phe Cys 350 355 360 ctt ctg gtg gtg gtg gtg
gtg ctc atg tcc cga tac cat cgg cgc aag 1397 Leu Leu Val Val Val
Val Val Leu Met Ser Arg Tyr His Arg Arg Lys 365 370 375 gcc cag cag
atg acc cag aaa tat gag gag gag ctg acc ctg acc agg 1445 Ala Gln
Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg 380 385 390
gag aac tcc atc cgg agg ctg cat tcc cat cac acg gac ccc agg agc
1493 Glu Asn Ser Ile Arg Arg Leu His Ser His His Thr Asp Pro Arg
Ser 395 400 405 410 cag ccg gag gag agt gta ggg ctg aga gcc gag ggc
cac cct gat agt 1541 Gln Pro Glu Glu Ser Val Gly Leu Arg Ala Glu
Gly His Pro Asp Ser 415 420 425 ctc aag gac aac agt agc tgc tct gtg
atg agt gaa gag ccc gag ggc 1589 Leu Lys Asp Asn Ser Ser Cys Ser
Val Met Ser Glu Glu Pro Glu Gly 430 435 440 cgc agt tac tcc acg ctg
acc acg gtg agg gag ata gaa aca cag act 1637 Arg Ser Tyr Ser Thr
Leu Thr Thr Val Arg Glu Ile Glu Thr Gln Thr 445 450 455 gaa ctg ctg
tct cca ggc tct ggg cgg gcc gag gag gag gaa gat cag 1685 Glu Leu
Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln 460 465 470
gat gaa ggc atc aaa cag gcc atg aac cat ttt gtt cag gag aat ggg
1733 Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val Gln Glu Asn
Gly 475 480 485 490 acc cta cgg gcc aag ccc acg ggc aat ggc atc tac
atc aat ggg cgg 1781 Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile
Tyr Ile Asn Gly Arg 495 500 505 gga cac ctg gtc tga cccaggcctg
cctcccttcc ctaggcctgg ctccttctgt 1836 Gly His Leu Val * 510
tgacatggga gattttagct catcttgggg gcctccttaa acacccccat ttcttgcgga
1896 agatgctccc catcccactg actgcttgac ctttacctcc aacccttctg
ttcatcggga 1956 gggctccacc aattgagtct ctcccaccat gcatgcaggt
cactgtgtgt gtgcatgtgt 2016 gcctgtgtga gtgttgactg actgtgtgtg
tgtggagggg tgactgtccg tggaggggtg 2076 actgtgtccg tggtgtgtat
tatgctgtca tatcagagtc aagtgaactg tggtgtatgt 2136 gccacgggat
ttgagtggtt gcgtgggcaa cactgtcagg gtttggcgtg tgtgtcatgt 2196
ggctgtgtgt gacctctgcc tgaaaaagca ggtattttct cagaccccag agcagtatta
2256 atgatgcaga ggttggagga gagaggtgga gactgtggct cagacccagg
tgtgcgggca 2316 tagctggagc tggaatctgc ctccggtgtg agggaacctg
tctcctacca cttcggagcc 2376 atgggggcaa gtgtgaagca gccagtccct
gggtcagcca gaggcttgaa ctgttacaga 2436 agccctctgc cctctggtgg
cctctgggcc tgctgcatgt acatattttc tgtaaatata 2496 catgcgccgg
gagcttcttg caggaatact gctccgaatc acttttaatt tttttctttt 2556
ttttttcttg ccctttccat tagttgtatt ttttatttat ttttattttt attttttttt
2616 agagatggag tctcactatg ttgctcaggc tggccttgaa ctcctgggct
caagcaatcc 2676 tcctgcctca gcctccctag tagctgggac tttaagtgta
caccactgtg cctgctttga 2736 atcctttacg aagagaaaaa aaaaattaaa
gaaagccttt agatttatcc aatgtttact 2796 actgggattg cttaaagtga
ggcccctcca acaccagggg gttaattcct gtgattgtga 2856 aaggggctac
ttccaaggca tcttcatgca ggcagcccct tgggagggca cctgagagct 2916
ggtagagtct gaaattaggg atgtgagcct cgtggttact gagtaaggta aaattgcatc
2976 caccattgtt tgtgatacct tagggaattg cttggacctg gtgacaaggg
ctcctgttca 3036 atagtggtgt tggggagaga gagagcagtg attatagacc
gagagagtag gagttgaggt 3096 gaggtgaagg aggtgctggg ggtgagaatg
tcgcctttcc ccctgggttt tggatcacta 3156 attcaaggct cttctggatg
tttctctggg ttggggctgg agttcaatga ggtttatttt 3216 tagctggccc
acccagatac actcagccag aatacctaga tttagtaccc aaactcttct 3276
tagtctgaaa tctgctggat ttctggccta agggagaggc tcccatcctt cgttccccag
3336 ccagcctagg acttcgaatg tggagcctga agatctaaga tcctaacatg
tacattttat 3396 gtaaatatgt gcatatttgt acataaaatg atattctgtt
tttaaataaa cagacaaaac 3456 ttgaaaaa 3464 21 510 PRT Homo sapiens 21
Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5
10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala
Gly 20 25 30 Glu Leu Gly Thr Ser Asp Val Val Thr Val Val Leu Gly
Gln Asp Ala 35 40 45 Lys
Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55
60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala
65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr
Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu
Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu
Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser
Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu
Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln
Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro
Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185
190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe
195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr
Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile
Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser
Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg
Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro
Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser
Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu
Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310
315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro
Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser
Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu
Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg
Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr
Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His
His Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu
Arg Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430
Cys Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435
440 445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro
Gly 450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly
Ile Lys Gln 465 470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly
Thr Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn
Gly Arg Gly His Leu Val 500 505 510 22 3464 DNA Homo sapiens CDS
(264)...(1796) 22 ggccgtcgtt gttggccaca gcgtgggaag cagctctggg
ggagctcgga gctcccgatc 60 acggcttctt gggggtagct acggctgggt
gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg gagacccaag
tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180 cagttcctta
ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac 240
gctgggcagt ctgcctttca acc atg ccc ctg tcc ctg gga gcc gag atg tgg
293 Met Pro Leu Ser Leu Gly Ala Glu Met Trp 1 5 10 ggg cct gag gcc
tgg ctg ctg ctg ctg cta ctg ctg gca tca ttt aca 341 Gly Pro Glu Ala
Trp Leu Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr 15 20 25 ggc cgg
tgc ccc gcg ggt gag ctg gag acc tca gac gtg gta act gtg 389 Gly Arg
Cys Pro Ala Gly Glu Leu Glu Thr Ser Asp Val Val Thr Val 30 35 40
gtg ctg ggc cag gac gca aaa ctg ccc tgc ttc tac cga ggg gac tcc 437
Val Leu Gly Gln Asp Ala Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser 45
50 55 ggc gag caa gtg ggg caa gtg gca tgg gct cgg gtg gac gcg ggc
gaa 485 Gly Glu Gln Val Gly Gln Val Ala Trp Ala Arg Val Asp Ala Gly
Glu 60 65 70 ggc gcc cag gaa cta gcg cta ctg cac tcc aaa tac ggg
ctt cat gtg 533 Gly Ala Gln Glu Leu Ala Leu Leu His Ser Lys Tyr Gly
Leu His Val 75 80 85 90 agc ccg gct tac gag ggc cgc gtg gag cag ccg
ccg ccc cca cgc aac 581 Ser Pro Ala Tyr Glu Gly Arg Val Glu Gln Pro
Pro Pro Pro Arg Asn 95 100 105 ccc ctg gac ggc tca gtg ctc ctg cgc
aac gca gtg cag gcg gat gag 629 Pro Leu Asp Gly Ser Val Leu Leu Arg
Asn Ala Val Gln Ala Asp Glu 110 115 120 ggc gag tac gag tgc cgg gtc
agc acc ttc ccc gcc ggc agc ttc cag 677 Gly Glu Tyr Glu Cys Arg Val
Ser Thr Phe Pro Ala Gly Ser Phe Gln 125 130 135 gcg cgg ctg cgg ctc
cga gtg atg gtg cct ccc ctg ccc tca ctg aat 725 Ala Arg Leu Arg Leu
Arg Val Met Val Pro Pro Leu Pro Ser Leu Asn 140 145 150 cct ggt cca
gca cta gaa gag ggc cag ggc ctg acc ctg gca gcc tcc 773 Pro Gly Pro
Ala Leu Glu Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser 155 160 165 170
tgc aca gct gag ggc agc cca gcc ccc agc gtg acc tgg gac acg gag 821
Cys Thr Ala Glu Gly Ser Pro Ala Pro Ser Val Thr Trp Asp Thr Glu 175
180 185 gtc aaa ggc aca acg tcc agc cgt tcc ttc aag cac tcc cgc tct
gct 869 Val Lys Gly Thr Thr Ser Ser Arg Ser Phe Lys His Ser Arg Ser
Ala 190 195 200 gcc gtc acc tca gag ttc cac ttg gtg cct agc cgc agc
atg aat ggg 917 Ala Val Thr Ser Glu Phe His Leu Val Pro Ser Arg Ser
Met Asn Gly 205 210 215 cag cca ctg act tgt gtg gtg tcc cat cct ggc
ctg ctc cag gac caa 965 Gln Pro Leu Thr Cys Val Val Ser His Pro Gly
Leu Leu Gln Asp Gln 220 225 230 agg atc acc cac atc ctc cac gtg tcc
ttc ctt gct gag gcc tct gtg 1013 Arg Ile Thr His Ile Leu His Val
Ser Phe Leu Ala Glu Ala Ser Val 235 240 245 250 agg ggc ctt gaa gac
caa aat ctg tgg cac att ggc aga gaa gga gct 1061 Arg Gly Leu Glu
Asp Gln Asn Leu Trp His Ile Gly Arg Glu Gly Ala 255 260 265 atg ctc
aag tgc ctg agt gaa ggg cag ccc cct ccc tca tac aac tgg 1109 Met
Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp 270 275
280 aca cgg ctg gat ggg cct ctg ccc agt ggg gta cga gtg gat ggg gac
1157 Thr Arg Leu Asp Gly Pro Leu Pro Ser Gly Val Arg Val Asp Gly
Asp 285 290 295 act ttg ggc ttt ccc cca ctg acc act gag cac agc ggc
atc tac gtc 1205 Thr Leu Gly Phe Pro Pro Leu Thr Thr Glu His Ser
Gly Ile Tyr Val 300 305 310 tgc cat gtc agc aat gag ttc tcc tca agg
gat tct cag gtc act gtg 1253 Cys His Val Ser Asn Glu Phe Ser Ser
Arg Asp Ser Gln Val Thr Val 315 320 325 330 gat gtt ctt gac ccc cag
gaa gac tct ggg aag cag gtg gac cta gtg 1301 Asp Val Leu Asp Pro
Gln Glu Asp Ser Gly Lys Gln Val Asp Leu Val 335 340 345 tca gcc tcg
gtg gtg gtg gtg ggt gtg atc gcc gca ctc ttg ttc tgc 1349 Ser Ala
Ser Val Val Val Val Gly Val Ile Ala Ala Leu Leu Phe Cys 350 355 360
ctt ctg gtg gtg gtg gtg gtg ctc atg tcc cga tac cat cgg cgc aag
1397 Leu Leu Val Val Val Val Val Leu Met Ser Arg Tyr His Arg Arg
Lys 365 370 375 gcc cag cag atg acc cag aaa tat gag gag gag ctg acc
ctg acc agg 1445 Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu
Thr Leu Thr Arg 380 385 390 gag aac tcc atc cgg agg ctg cat tcc cat
cac acg gac ccc agg agc 1493 Glu Asn Ser Ile Arg Arg Leu His Ser
His His Thr Asp Pro Arg Ser 395 400 405 410 cag ccg gag gag agt gta
ggg ctg aga gcc gag ggc cac cct gat agt 1541 Gln Pro Glu Glu Ser
Val Gly Leu Arg Ala Glu Gly His Pro Asp Ser 415 420 425 ctc aag gac
aac agt agc tgc tct gtg atg agt gaa gag ccc gag ggc 1589 Leu Lys
Asp Asn Ser Ser Cys Ser Val Met Ser Glu Glu Pro Glu Gly 430 435 440
cgc agt tac tcc acg ctg acc acg gtg agg gag ata gaa aca cag act
1637 Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr Gln
Thr 445 450 455 gaa ctg ctg tct cca ggc tct ggg cgg gcc gag gag gag
gaa gat cag 1685 Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu
Glu Glu Asp Gln 460 465 470 gat gaa ggc atc aaa cag gcc atg aac cat
ttt gtt cag gag aat ggg 1733 Asp Glu Gly Ile Lys Gln Ala Met Asn
His Phe Val Gln Glu Asn Gly 475 480 485 490 acc cta cgg gcc aag ccc
acg ggc aat ggc atc tac atc aat ggg cgg 1781 Thr Leu Arg Ala Lys
Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg 495 500 505 gga cac ctg
gtc tga cccaggcctg cctcccttcc ctaggcctgg ctccttctgt 1836 Gly His
Leu Val * 510 tgacatggga gattttagct catcttgggg gcctccttaa
acacccccat ttcttgcgga 1896 agatgctccc catcccactg actgcttgac
ctttacctcc aacccttctg ttcatcggga 1956 gggctccacc aattgagtct
ctcccaccat gcatgcaggt cactgtgtgt gtgcatgtgt 2016 gcctgtgtga
gtgttgactg actgtgtgtg tgtggagggg tgactgtccg tggaggggtg 2076
actgtgtccg tggtgtgtat tatgctgtca tatcagagtc aagtgaactg tggtgtatgt
2136 gccacgggat ttgagtggtt gcgtgggcaa cactgtcagg gtttggcgtg
tgtgtcatgt 2196 ggctgtgtgt gacctctgcc tgaaaaagca ggtattttct
cagaccccag agcagtatta 2256 atgatgcaga ggttggagga gagaggtgga
gactgtggct cagacccagg tgtgcgggca 2316 tagctggagc tggaatctgc
ctccggtgtg agggaacctg tctcctacca cttcggagcc 2376 atgggggcaa
gtgtgaagca gccagtccct gggtcagcca gaggcttgaa ctgttacaga 2436
agccctctgc cctctggtgg cctctgggcc tgctgcatgt acatattttc tgtaaatata
2496 catgcgccgg gagcttcttg caggaatact gctccgaatc acttttaatt
tttttctttt 2556 ttttttcttg ccctttccat tagttgtatt ttttatttat
ttttattttt attttttttt 2616 agagatggag tctcactatg ttgctcaggc
tggccttgaa ctcctgggct caagcaatcc 2676 tcctgcctca gcctccctag
tagctgggac tttaagtgta caccactgtg cctgctttga 2736 atcctttacg
aagagaaaaa aaaaattaaa gaaagccttt agatttatcc aatgtttact 2796
actgggattg cttaaagtga ggcccctcca acaccagggg gttaattcct gtgattgtga
2856 aaggggctac ttccaaggca tcttcatgca ggcagcccct tgggagggca
cctgagagct 2916 ggtagagtct gaaattaggg atgtgagcct cgtggttact
gagtaaggta aaattgcatc 2976 caccattgtt tgtgatacct tagggaattg
cttggacctg gtgacaaggg ctcctgttca 3036 atagtggtgt tggggagaga
gagagcagtg attatagacc gagagagtag gagttgaggt 3096 gaggtgaagg
aggtgctggg ggtgagaatg tcgcctttcc ccctgggttt tggatcacta 3156
attcaaggct cttctggatg tttctctggg ttggggctgg agttcaatga ggtttatttt
3216 tagctggccc acccagatac actcagccag aatacctaga tttagtaccc
aaactcttct 3276 tagtctgaaa tctgctggat ttctggccta agggagaggc
tcccatcctt cgttccccag 3336 ccagcctagg acttcgaatg tggagcctga
agatctaaga tcctaacatg tacattttat 3396 gtaaatatgt gcatatttgt
acataaaatg atattctgtt tttaaataaa cagacaaaac 3456 ttgaaaaa 3464 23
510 PRT Homo sapiens 23 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly
Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe
Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val
Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe
Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp
Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu
Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90
95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val
100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu
Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg
Leu Arg Leu Arg 130 135 140 Val Met Val Pro Pro Leu Pro Ser Leu Asn
Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu
Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val
Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser
Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His
Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215
220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu
225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu
Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met
Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn
Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val
Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His
Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser
Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335
Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340
345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val
Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln
Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu
Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp Pro
Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly
His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met
Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr
Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460
Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465
470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala
Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His
Leu Val 500 505 510 24 3464 DNA Homo sapiens CDS (264)...(1796) 24
ggccgtcgtt gttggccaca gcgtgggaag cagctctggg ggagctcgga gctcccgatc
60 acggcttctt gggggtagct acggctgggt gtgtagaacg gggccggggc
tggggctggg 120 tcccctagtg gagacccaag tgcgagaggc aagaactctg
cagcttcctg ccttctgggt 180 cagttcctta ttcaagtctg cagccggctc
ccagggagat ctcggtggaa cttcagaaac 240 gctgggcagt ctgcctttca acc atg
ccc ctg tcc ctg gga gcc gag atg tgg 293 Met Pro Leu Ser Leu Gly Ala
Glu Met Trp 1 5 10 ggg cct gag gcc tgg ctg ctg ctg ctg cta ctg ctg
gca tca ttt aca 341 Gly Pro Glu Ala Trp Leu Leu Leu Leu Leu Leu Leu
Ala Ser Phe Thr 15 20 25 ggc cgg tgc ccc gcg ggt gag ctg gag acc
tca gac gtg gta act gtg 389 Gly Arg Cys Pro Ala Gly Glu Leu Glu Thr
Ser Asp Val Val Thr Val 30 35 40 gtg ctg ggc cag gac gca aaa ctg
ccc tgc ttc tac cga ggg gac tcc 437 Val Leu Gly Gln Asp Ala Lys Leu
Pro Cys Phe Tyr Arg Gly Asp Ser 45 50 55 ggc gag caa gtg ggg caa
gtg gca tgg gct cgg gtg gac gcg ggc gaa 485 Gly Glu Gln Val Gly Gln
Val Ala Trp Ala Arg Val Asp Ala Gly Glu 60 65 70 ggc gcc cag gaa
cta gcg cta ctg cac tcc aaa tac ggg ctt cat gtg 533 Gly Ala Gln Glu
Leu Ala Leu Leu His Ser Lys Tyr Gly Leu His Val 75 80 85 90 agc ccg
gct tac gag ggc cgc gtg gag cag ccg ccg ccc cca cgc aac 581 Ser Pro
Ala Tyr Glu Gly Arg Val Glu Gln Pro Pro Pro Pro Arg Asn 95 100 105
ccc ctg gac ggc tca gtg ctc ctg cgc aac gca gtg cag gcg gat gag 629
Pro Leu Asp Gly Ser Val Leu Leu Arg Asn Ala Val Gln Ala Asp Glu 110
115 120 ggc gag tac gag tgc cgg gtc agc acc ttc ccc gcc ggc agc ttc
cag 677 Gly Glu Tyr Glu Cys Arg Val Ser Thr Phe Pro Ala Gly Ser Phe
Gln 125 130 135 gcg cgg ctg cgg ctc cga gtg ctg gtg cct ccc ctg ccc
tca ctg aat 725 Ala Arg Leu Arg Leu Arg Val Leu Val Pro Pro Leu Pro
Ser Leu Asn 140 145 150 cct ggt cca gca cta gaa gag ggc cag ggc ctg
acc
ctg gca gcc tcc 773 Pro Gly Pro Ala Leu Glu Glu Gly Gln Gly Leu Thr
Leu Ala Ala Ser 155 160 165 170 tgc aca gct gag ggc agc cca gcc ccc
agc gtg acc tgg gac acg gag 821 Cys Thr Ala Glu Gly Ser Pro Ala Pro
Ser Val Thr Trp Asp Thr Glu 175 180 185 gtc aaa ggc aca acg tcc agc
cgt tcc ttc aag cac tcc cgc tct gct 869 Val Lys Gly Thr Thr Ser Ser
Arg Ser Phe Lys His Ser Arg Ser Ala 190 195 200 gcc gtc acc tca gag
ttc cac ttg gtg cct agc cgc agc atg aat ggg 917 Ala Val Thr Ser Glu
Phe His Leu Val Pro Ser Arg Ser Met Asn Gly 205 210 215 cag cca ctg
act tgt gtg gtg tcc cat cct ggc ctg ctc cag gac caa 965 Gln Pro Leu
Thr Cys Val Val Ser His Pro Gly Leu Leu Gln Asp Gln 220 225 230 agg
atc acc cac atc ctc cac gtg tcc ttc ctt gct gag gcc tct gtg 1013
Arg Ile Thr His Ile Leu His Val Ser Phe Leu Ala Glu Ala Ser Val 235
240 245 250 agg ggc ctt gaa gac caa aat ctg tgg cac att ggc aga gaa
gga gct 1061 Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile Gly Arg
Glu Gly Ala 255 260 265 atg ctc aag tgc ctg agt gaa ggg cag ccc cct
ccc tca tac aac tgg 1109 Met Leu Lys Cys Leu Ser Glu Gly Gln Pro
Pro Pro Ser Tyr Asn Trp 270 275 280 aca cgg ctg gat ggg cct ctg ccc
agt ggg gta cga gtg gat ggg gac 1157 Thr Arg Leu Asp Gly Pro Leu
Pro Ser Gly Val Arg Val Asp Gly Asp 285 290 295 act ttg ggc ttt ccc
cca ctg acc act gag cac agc ggc atc tac gtc 1205 Thr Leu Gly Phe
Pro Pro Leu Thr Thr Glu His Ser Gly Ile Tyr Val 300 305 310 tgc cat
gtc agc aat gag ttc tcc tca agg gat tct cag gtc act gtg 1253 Cys
His Val Ser Asn Glu Phe Ser Ser Arg Asp Ser Gln Val Thr Val 315 320
325 330 gat gtt ctt gac ccc cag gaa gac tct ggg aag cag gtg gac cta
gtg 1301 Asp Val Leu Asp Pro Gln Glu Asp Ser Gly Lys Gln Val Asp
Leu Val 335 340 345 tca gcc tcg gtg gtg gtg gtg ggt gtg atc gcc gca
ctc ttg ttc tgc 1349 Ser Ala Ser Val Val Val Val Gly Val Ile Ala
Ala Leu Leu Phe Cys 350 355 360 ctt ctg gtg gtg gtg gtg gtg ctc atg
tcc cga tac cat cgg cgc aag 1397 Leu Leu Val Val Val Val Val Leu
Met Ser Arg Tyr His Arg Arg Lys 365 370 375 gcc cag cag atg acc cag
aaa tat gag gag gag ctg acc ctg acc agg 1445 Ala Gln Gln Met Thr
Gln Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg 380 385 390 gag aac tcc
atc cgg agg ctg cat tcc cat cac acg gac ccc agg agc 1493 Glu Asn
Ser Ile Arg Arg Leu His Ser His His Thr Asp Pro Arg Ser 395 400 405
410 cag ccg gag gag agt gta ggg ctg aga gcc gag ggc cac cct gat agt
1541 Gln Pro Glu Glu Ser Val Gly Leu Arg Ala Glu Gly His Pro Asp
Ser 415 420 425 ctc aag gac aac agt agc tgc tct gtg atg agt gaa gag
ccc gag ggc 1589 Leu Lys Asp Asn Ser Ser Cys Ser Val Met Ser Glu
Glu Pro Glu Gly 430 435 440 tgc agt tac tcc acg ctg acc acg gtg agg
gag ata gaa aca cag act 1637 Cys Ser Tyr Ser Thr Leu Thr Thr Val
Arg Glu Ile Glu Thr Gln Thr 445 450 455 gaa ctg ctg tct cca ggc tct
ggg cgg gcc gag gag gag gaa gat cag 1685 Glu Leu Leu Ser Pro Gly
Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln 460 465 470 gat gaa ggc atc
aaa cag gcc atg aac cat ttt gtt cag gag aat ggg 1733 Asp Glu Gly
Ile Lys Gln Ala Met Asn His Phe Val Gln Glu Asn Gly 475 480 485 490
acc cta cgg gcc aag ccc acg ggc aat ggc atc tac atc aat ggg cgg
1781 Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly
Arg 495 500 505 gga cac ctg gtc tga cccaggcctg cctcccttcc
ctaggcctgg ctccttctgt 1836 Gly His Leu Val * 510 tgacatggga
gattttagct catcttgggg gcctccttaa acacccccat ttcttgcgga 1896
agatgctccc catcccactg actgcttgac ctttacctcc aacccttctg ttcatcggga
1956 gggctccacc aattgagtct ctcccaccat gcatgcaggt cactgtgtgt
gtgcatgtgt 2016 gcctgtgtga gtgttgactg actgtgtgtg tgtggagggg
tgactgtccg tggaggggtg 2076 actgtgtccg tggtgtgtat tatgctgtca
tatcagagtc aagtgaactg tggtgtatgt 2136 gccacgggat ttgagtggtt
gcgtgggcaa cactgtcagg gtttggcgtg tgtgtcatgt 2196 ggctgtgtgt
gacctctgcc tgaaaaagca ggtattttct cagaccccag agcagtatta 2256
atgatgcaga ggttggagga gagaggtgga gactgtggct cagacccagg tgtgcgggca
2316 tagctggagc tggaatctgc ctccggtgtg agggaacctg tctcctacca
cttcggagcc 2376 atgggggcaa gtgtgaagca gccagtccct gggtcagcca
gaggcttgaa ctgttacaga 2436 agccctctgc cctctggtgg cctctgggcc
tgctgcatgt acatattttc tgtaaatata 2496 catgcgccgg gagcttcttg
caggaatact gctccgaatc acttttaatt tttttctttt 2556 ttttttcttg
ccctttccat tagttgtatt ttttatttat ttttattttt attttttttt 2616
agagatggag tctcactatg ttgctcaggc tggccttgaa ctcctgggct caagcaatcc
2676 tcctgcctca gcctccctag tagctgggac tttaagtgta caccactgtg
cctgctttga 2736 atcctttacg aagagaaaaa aaaaattaaa gaaagccttt
agatttatcc aatgtttact 2796 actgggattg cttaaagtga ggcccctcca
acaccagggg gttaattcct gtgattgtga 2856 aaggggctac ttccaaggca
tcttcatgca ggcagcccct tgggagggca cctgagagct 2916 ggtagagtct
gaaattaggg atgtgagcct cgtggttact gagtaaggta aaattgcatc 2976
caccattgtt tgtgatacct tagggaattg cttggacctg gtgacaaggg ctcctgttca
3036 atagtggtgt tggggagaga gagagcagtg attatagacc gagagagtag
gagttgaggt 3096 gaggtgaagg aggtgctggg ggtgagaatg tcgcctttcc
ccctgggttt tggatcacta 3156 attcaaggct cttctggatg tttctctggg
ttggggctgg agttcaatga ggtttatttt 3216 tagctggccc acccagatac
actcagccag aatacctaga tttagtaccc aaactcttct 3276 tagtctgaaa
tctgctggat ttctggccta agggagaggc tcccatcctt cgttccccag 3336
ccagcctagg acttcgaatg tggagcctga agatctaaga tcctaacatg tacattttat
3396 gtaaatatgt gcatatttgt acataaaatg atattctgtt tttaaataaa
cagacaaaac 3456 ttgaaaaa 3464 25 510 PRT Homo sapiens 25 Met Pro
Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15
Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20
25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp
Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln
Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly
Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His
Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro
Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala
Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr
Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val
Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150
155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly
Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly
Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala
Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn
Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu
Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe
Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu
Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270
Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275
280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro
Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val
Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val
Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp
Leu Val Ser Ala Ser Val Val Val 340 345 350 Val Gly Val Ile Ala Ala
Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360 365 Val Leu Met Ser
Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr
Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395
400 Leu His Ser His His Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val
405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn
Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu Pro Glu Gly Cys Ser
Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr
Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu
Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480 Ala Met Asn His Phe
Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn
Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500 505 510 26 3467 DNA
Homo sapiens CDS (264)...(1799) 26 ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac
240 gctgggcagt ctgcctttca acc atg ccc ctg tcc ctg gga gcc gag atg
tgg 293 Met Pro Leu Ser Leu Gly Ala Glu Met Trp 1 5 10 ggg cct gag
gcc tgg ctg ctg ctg ctg cta ctg ctg gca tca ttt aca 341 Gly Pro Glu
Ala Trp Leu Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr 15 20 25 ggc
cgg tgc ccc gcg ggt gag ctg gag acc tca gac gtg gta act gtg 389 Gly
Arg Cys Pro Ala Gly Glu Leu Glu Thr Ser Asp Val Val Thr Val 30 35
40 gtg ctg ggc cag gac gca aaa ctg ccc tgc ttc tac cga ggg gac tcc
437 Val Leu Gly Gln Asp Ala Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser
45 50 55 ggc gag caa gtg ggg caa gtg gca tgg gct cgg gtg gac gcg
ggc gaa 485 Gly Glu Gln Val Gly Gln Val Ala Trp Ala Arg Val Asp Ala
Gly Glu 60 65 70 ggc gcc cag gaa cta gcg cta ctg cac tcc aaa tac
ggg ctt cat gtg 533 Gly Ala Gln Glu Leu Ala Leu Leu His Ser Lys Tyr
Gly Leu His Val 75 80 85 90 agc ccg gct tac gag ggc cgc gtg gag cag
ccg ccg ccc cca cgc aac 581 Ser Pro Ala Tyr Glu Gly Arg Val Glu Gln
Pro Pro Pro Pro Arg Asn 95 100 105 ccc ctg gac ggc tca gtg ctc ctg
cgc aac gca gtg cag gcg gat gag 629 Pro Leu Asp Gly Ser Val Leu Leu
Arg Asn Ala Val Gln Ala Asp Glu 110 115 120 ggc gag tac gag tgc cgg
gtc agc acc ttc ccc gcc ggc agc ttc cag 677 Gly Glu Tyr Glu Cys Arg
Val Ser Thr Phe Pro Ala Gly Ser Phe Gln 125 130 135 gcg cgg ctg cgg
ctc cga gtg ctg gtg cct ccc ctg ccc tca ctg aat 725 Ala Arg Leu Arg
Leu Arg Val Leu Val Pro Pro Leu Pro Ser Leu Asn 140 145 150 cct ggt
cca gca cta gaa gag ggc cag ggc ctg acc ctg gca gcc tcc 773 Pro Gly
Pro Ala Leu Glu Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser 155 160 165
170 tgc aca gct gag ggc agc cca gcc ccc agc gtg acc tgg gac acg gag
821 Cys Thr Ala Glu Gly Ser Pro Ala Pro Ser Val Thr Trp Asp Thr Glu
175 180 185 gtc aaa ggc aca acg tcc agc cgt tcc ttc aag cac tcc cgc
tct gct 869 Val Lys Gly Thr Thr Ser Ser Arg Ser Phe Lys His Ser Arg
Ser Ala 190 195 200 gcc gtc acc tca gag ttc cac ttg gtg cct agc cgc
agc atg aat ggg 917 Ala Val Thr Ser Glu Phe His Leu Val Pro Ser Arg
Ser Met Asn Gly 205 210 215 cag cca ctg act tgt gtg gtg tcc cat cct
ggc ctg ctc cag gac caa 965 Gln Pro Leu Thr Cys Val Val Ser His Pro
Gly Leu Leu Gln Asp Gln 220 225 230 agg atc acc cac atc ctc cac gtg
tcc ttc ctt gct gag gcc tct gtg 1013 Arg Ile Thr His Ile Leu His
Val Ser Phe Leu Ala Glu Ala Ser Val 235 240 245 250 agg ggc ctt gaa
gac caa aat ctg tgg cac att ggc aga gaa gga gct 1061 Arg Gly Leu
Glu Asp Gln Asn Leu Trp His Ile Gly Arg Glu Gly Ala 255 260 265 atg
ctc aag tgc ctg agt gaa ggg cag ccc cct ccc tca tac aac tgg 1109
Met Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp 270
275 280 aca cgg ctg gat ggg cct ctg ccc agt ggg gta cga gtg gat ggg
gac 1157 Thr Arg Leu Asp Gly Pro Leu Pro Ser Gly Val Arg Val Asp
Gly Asp 285 290 295 act ttg ggc ttt ccc cca ctg acc act gag cac agc
ggc atc tac gtc 1205 Thr Leu Gly Phe Pro Pro Leu Thr Thr Glu His
Ser Gly Ile Tyr Val 300 305 310 tgc cat gtc agc aat gag ttc tcc tca
agg gat tct cag gtc act gtg 1253 Cys His Val Ser Asn Glu Phe Ser
Ser Arg Asp Ser Gln Val Thr Val 315 320 325 330 gat gtt ctt gca gac
ccc cag gaa gac tct ggg aag cag gtg gac cta 1301 Asp Val Leu Ala
Asp Pro Gln Glu Asp Ser Gly Lys Gln Val Asp Leu 335 340 345 gtg tca
gcc tcg gtg gtg gtg gtg ggt gtg atc gcc gca ctc ttg ttc 1349 Val
Ser Ala Ser Val Val Val Val Gly Val Ile Ala Ala Leu Leu Phe 350 355
360 tgc ctt ctg gtg gtg gtg gtg gtg ctc atg tcc cga tac cat cgg cgc
1397 Cys Leu Leu Val Val Val Val Val Leu Met Ser Arg Tyr His Arg
Arg 365 370 375 aag gcc cag cag atg acc cag aaa tat gag gag gag ctg
acc ctg acc 1445 Lys Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu
Leu Thr Leu Thr 380 385 390 agg gag aac tcc atc cgg agg ctg cat tcc
cat cac acg gac ccc agg 1493 Arg Glu Asn Ser Ile Arg Arg Leu His
Ser His His Thr Asp Pro Arg 395 400 405 410 agc cag ccg gag gag agt
gta ggg ctg aga gcc gag ggc cac cct gat 1541 Ser Gln Pro Glu Glu
Ser Val Gly Leu Arg Ala Glu Gly His Pro Asp 415 420 425 agt ctc aag
gac aac agt agc tgc tct gtg atg agt gaa gag ccc gag 1589 Ser Leu
Lys Asp Asn Ser Ser Cys Ser Val Met Ser Glu Glu Pro Glu 430 435 440
ggc cgc agt tac tcc acg ctg acc acg gtg agg gag ata gaa aca cag
1637 Gly Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr
Gln 445 450 455 act gaa ctg ctg tct cca ggc tct ggg cgg gcc gag gag
gag gaa gat 1685 Thr Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu
Glu Glu Glu Asp 460 465 470 cag gat gaa ggc atc aaa cag gcc atg aac
cat ttt gtt cag gag aat 1733 Gln Asp Glu Gly Ile Lys Gln Ala Met
Asn His Phe Val Gln Glu Asn 475 480 485 490 ggg acc cta cgg gcc aag
ccc acg ggc aat ggc atc tac atc aat ggg 1781 Gly Thr Leu Arg Ala
Lys Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly 495 500 505 cgg gga cac
ctg gtc tga cccaggcctg cctcccttcc ctaggcctgg 1829 Arg Gly His Leu
Val * 510 ctccttctgt tgacatggga gattttagct catcttgggg gcctccttaa
acacccccat 1889 ttcttgcgga agatgctccc catcccactg actgcttgac
ctttacctcc aacccttctg 1949 ttcatcggga gggctccacc aattgagtct
ctcccaccat gcatgcaggt cactgtgtgt 2009 gtgcatgtgt gcctgtgtga
gtgttgactg actgtgtgtg tgtggagggg tgactgtccg 2069 tggaggggtg
actgtgtccg tggtgtgtat tatgctgtca tatcagagtc aagtgaactg 2129
tggtgtatgt gccacgggat ttgagtggtt gcgtgggcaa cactgtcagg gtttggcgtg
2189 tgtgtcatgt ggctgtgtgt gacctctgcc tgaaaaagca ggtattttct
cagaccccag 2249 agcagtatta atgatgcaga ggttggagga gagaggtgga
gactgtggct cagacccagg 2309 tgtgcgggca tagctggagc tggaatctgc
ctccggtgtg agggaacctg tctcctacca 2369 cttcggagcc atgggggcaa
gtgtgaagca gccagtccct gggtcagcca gaggcttgaa 2429 ctgttacaga
agccctctgc cctctggtgg cctctgggcc tgctgcatgt acatattttc 2489
tgtaaatata catgcgccgg gagcttcttg caggaatact gctccgaatc acttttaatt
2549 tttttctttt ttttttcttg ccctttccat tagttgtatt ttttatttat
ttttattttt 2609 attttttttt agagatggag tctcactatg ttgctcaggc
tggccttgaa ctcctgggct 2669 caagcaatcc tcctgcctca gcctccctag
tagctgggac tttaagtgta caccactgtg 2729 cctgctttga atcctttacg
aagagaaaaa aaaaattaaa gaaagccttt agatttatcc 2789 aatgtttact
actgggattg cttaaagtga ggcccctcca acaccagggg gttaattcct 2849
gtgattgtga aaggggctac ttccaaggca tcttcatgca ggcagcccct tgggagggca
2909 cctgagagct ggtagagtct gaaattaggg atgtgagcct cgtggttact
gagtaaggta 2969 aaattgcatc caccattgtt tgtgatacct tagggaattg
cttggacctg
gtgacaaggg 3029 ctcctgttca atagtggtgt tggggagaga gagagcagtg
attatagacc gagagagtag 3089 gagttgaggt gaggtgaagg aggtgctggg
ggtgagaatg tcgcctttcc ccctgggttt 3149 tggatcacta attcaaggct
cttctggatg tttctctggg ttggggctgg agttcaatga 3209 ggtttatttt
tagctggccc acccagatac actcagccag aatacctaga tttagtaccc 3269
aaactcttct tagtctgaaa tctgctggat ttctggccta agggagaggc tcccatcctt
3329 cgttccccag ccagcctagg acttcgaatg tggagcctga agatctaaga
tcctaacatg 3389 tacattttat gtaaatatgt gcatatttgt acataaaatg
atattctgtt tttaaataaa 3449 cagacaaaac ttgaaaaa 3467 27 511 PRT Homo
sapiens 27 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala
Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg
Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val
Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly
Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val
Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser
Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val
Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110
Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115
120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu
Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro
Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser
Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp
Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His
Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro
Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser
His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235
240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln
245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys
Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg
Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp
Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile
Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp
Ser Gln Val Thr Val Asp Val Leu Ala Asp Pro 325 330 335 Gln Glu Asp
Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val 340 345 350 Val
Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val 355 360
365 Val Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr
370 375 380 Gln Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser
Ile Arg 385 390 395 400 Arg Leu His Ser His His Thr Asp Pro Arg Ser
Gln Pro Glu Glu Ser 405 410 415 Val Gly Leu Arg Ala Glu Gly His Pro
Asp Ser Leu Lys Asp Asn Ser 420 425 430 Ser Cys Ser Val Met Ser Glu
Glu Pro Glu Gly Arg Ser Tyr Ser Thr 435 440 445 Leu Thr Thr Val Arg
Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro 450 455 460 Gly Ser Gly
Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys 465 470 475 480
Gln Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys 485
490 495 Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val
500 505 510 28 1669 DNA Homo sapiens CDS (708)...(1121) 28
gtctgaccca ggcctgcctc ccttccctag gcctggctcc ttctgttgac atgggagatt
60 ttagctcatc ttgggggcct ccttaaacac ccccatttct tgcggaagat
gctccccatc 120 ccactgactg cttgaccttt acctccaacc cttctgttca
tcgggagggc tccaccaatt 180 gagtctctcc caccatgcat gcaggtcact
gtgtgtgtgc atgtgtgcct gtgtgagtgt 240 tgactgactg tgtgtgtgtg
gaggggtgac tgtccgtgga ggggtgactg tgtccgtggt 300 gtgtattatg
ctgtcatatc agagtcaagt gaactgtggt gtatgtgcca cgggatttga 360
gtggttgcgt gggcaacact gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc
420 tctgcctgaa aaagcaggta ttttctcaga ccccagagca gtattaatga
tgcagaggtt 480 ggaggagaga ggtggagact gtggctcaga cccaggtgtg
cgggcatagc tggagctgga 540 atctgcctcc ggtgtgaggg aacctgtctc
ctaccacttc ggagccatgg gggcaagtgt 600 gaagcagcca gtccctgggt
cagccagagg cttgaactgt tacagaagcc ctctgccctc 660 tggtggcctc
tgggcctgct gcatgtacat attttctgta aatatac atg cgc cgg 716 Met Arg
Arg 1 gag ctt ctt gca gga ata ctg ctc cga atc act ttt aat ttt ttt
ctt 764 Glu Leu Leu Ala Gly Ile Leu Leu Arg Ile Thr Phe Asn Phe Phe
Leu 5 10 15 ttt ttt ttc ttg ccc ttt cca tta gtt gta ttt ttt att tat
ttt tat 812 Phe Phe Phe Leu Pro Phe Pro Leu Val Val Phe Phe Ile Tyr
Phe Tyr 20 25 30 35 ttt tat ttt ttt tta gag atg gag tct cac tat gtt
gct cag gct ggc 860 Phe Tyr Phe Phe Leu Glu Met Glu Ser His Tyr Val
Ala Gln Ala Gly 40 45 50 ctt gaa ctc ctg ggc tca agc aat cct cct
gcc tca gac tcc cta gta 908 Leu Glu Leu Leu Gly Ser Ser Asn Pro Pro
Ala Ser Asp Ser Leu Val 55 60 65 gct ggg act tta agt gta cac cac
tgt gcc tgc ttt gaa tcc ttt acg 956 Ala Gly Thr Leu Ser Val His His
Cys Ala Cys Phe Glu Ser Phe Thr 70 75 80 aag aga aaa aaa aaa tta
aag aaa gcc ttt aga ttt atc caa tgt tta 1004 Lys Arg Lys Lys Lys
Leu Lys Lys Ala Phe Arg Phe Ile Gln Cys Leu 85 90 95 cta ctg gga
ttg ctt aaa gtg agg ccc ctc caa cac cag ggg gtt aat 1052 Leu Leu
Gly Leu Leu Lys Val Arg Pro Leu Gln His Gln Gly Val Asn 100 105 110
115 tcc tgt gat tgt gaa agg ggc tac ttc caa ggc atc ttc atg cag gca
1100 Ser Cys Asp Cys Glu Arg Gly Tyr Phe Gln Gly Ile Phe Met Gln
Ala 120 125 130 gcc cct tgg gag ggc acc tga gagctggtag agtctgaaat
tagggatgtg 1151 Ala Pro Trp Glu Gly Thr * 135 agcctcgtgg ttactgagta
aggtaaaatt gcatccacca ttgtttgtga taccttaggg 1211 aattgcttgg
acctggtgac aagggctcct gttcaatagt ggtgttgggg agagagagag 1271
cagtgattat agaccgagag agtaggagtt gaggtgaggt gaaggaggtg ctgggggtga
1331 gaatgtcgcc tttccccctg ggttttggat cactaattca aggctcttct
ggatgtttct 1391 ctgggttggg gctggagttc aatgaggttt atttttagct
ggcccaccca gatacactca 1451 gccagaatac ctagatttag tacccaaact
cttcttagtc tgaaatctgc tggatttctg 1511 gcctaaggga gaggctccca
tccttcgttc cccagccagc ctaggacttc gaatgtggag 1571 cctgaagatc
taagatccta acatgtacat tttatgtaaa tatgtgcata tttgtacata 1631
aaatgatatt ctgtttttaa ataaacagac aaaacttg 1669 29 137 PRT Homo
sapiens 29 Met Arg Arg Glu Leu Leu Ala Gly Ile Leu Leu Arg Ile Thr
Phe Asn 1 5 10 15 Phe Phe Leu Phe Phe Phe Leu Pro Phe Pro Leu Val
Val Phe Phe Ile 20 25 30 Tyr Phe Tyr Phe Tyr Phe Phe Leu Glu Met
Glu Ser His Tyr Val Ala 35 40 45 Gln Ala Gly Leu Glu Leu Leu Gly
Ser Ser Asn Pro Pro Ala Ser Asp 50 55 60 Ser Leu Val Ala Gly Thr
Leu Ser Val His His Cys Ala Cys Phe Glu 65 70 75 80 Ser Phe Thr Lys
Arg Lys Lys Lys Leu Lys Lys Ala Phe Arg Phe Ile 85 90 95 Gln Cys
Leu Leu Leu Gly Leu Leu Lys Val Arg Pro Leu Gln His Gln 100 105 110
Gly Val Asn Ser Cys Asp Cys Glu Arg Gly Tyr Phe Gln Gly Ile Phe 115
120 125 Met Gln Ala Ala Pro Trp Glu Gly Thr 130 135 30 510 PRT Homo
sapiens 30 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala
Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg
Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val
Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly
Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val
Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser
Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val
Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110
Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115
120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu
Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro
Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser
Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp
Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His
Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro
Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser
His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235
240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln
245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys
Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg
Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp
Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile
Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp
Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser
Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val
Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360
365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln
370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile
Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln
Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp
Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu
Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu
Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg
Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480
Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485
490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500
505 510 31 510 PRT Homo sapiens 31 Met Pro Leu Ser Leu Gly Ala Glu
Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu
Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr
Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu
Pro Cys Leu Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60
Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65
70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr
Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu
Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu
Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser
Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu
Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln
Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro
Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185
190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe
195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr
Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile
Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser
Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg
Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro
Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser
Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu
Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310
315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro
Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser
Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu
Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg
Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr
Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His
His Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu
Arg Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430
Cys Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435
440 445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro
Gly 450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly
Ile Lys Gln 465 470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly
Thr Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn
Gly Arg Gly His Leu Val 500 505 510 32 295 PRT Homo sapiens 32 Met
Asn Gly Gln Pro Leu Thr Cys Val Val Ser His Pro Gly Leu Leu 1 5 10
15 Gln Asp Gln Arg Ile Thr His Ile Leu His Val Ser Phe Leu Ala Glu
20 25 30 Ala Ser Val Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile
Gly Arg 35 40 45 Glu Gly Ala Met Leu Lys Cys Leu Ser Glu Gly Gln
Pro Pro Pro Ser 50 55 60 Tyr Asn Trp Thr Arg Leu Asp Gly Pro Leu
Pro Ser Gly Val Arg Val 65 70 75 80 Asp Gly Asp Thr Leu Gly Phe Pro
Pro Leu Thr Thr Glu His Ser Gly 85 90 95 Ile Tyr Val Cys His Val
Ser Asn Glu Phe Ser Ser Arg Asp Ser Gln 100 105 110 Val Thr Val Asp
Val Leu Asp Pro Gln Glu Asp Ser Gly Lys Gln Val 115 120 125 Asp Leu
Val Ser Ala Ser Val Val Val Val Gly Val Ile Ala Ala Leu 130 135 140
Leu Phe Cys Leu Leu Val Val Val Val Val Leu Met Ser Arg Tyr His 145
150 155 160 Arg Arg Lys Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu
Leu Thr 165 170 175 Leu Thr Arg Glu Asn Ser Ile Arg Arg Leu His Ser
His His Thr Asp 180 185 190 Pro Arg Ser Gln Pro Glu Glu Ser Val Gly
Leu Arg Ala Glu Gly His 195 200 205 Pro Asp Ser Leu Lys Asp Asn Ser
Ser Cys Ser Val Met Ser Glu Glu 210 215 220 Pro Glu Gly Arg Ser Tyr
Ser Thr Leu Thr Thr Val
Arg Glu Ile Glu 225 230 235 240 Thr Gln Thr Glu Leu Leu Ser Pro Gly
Ser Gly Arg Ala Glu Glu Glu 245 250 255 Glu Asp Gln Asp Glu Gly Ile
Lys Gln Ala Met Asn His Phe Val Gln 260 265 270 Glu Asn Gly Thr Leu
Arg Ala Lys Pro Thr Gly Asn Gly Ile Tyr Ile 275 280 285 Asn Gly Arg
Gly His Leu Val 290 295 33 485 PRT Homo sapiens 33 Met Pro Leu Ser
Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu
Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30
Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35
40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly
Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln
Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser
Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg
Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln
Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro
Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val
Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160
Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165
170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr
Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr
Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln
Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp
Gln Arg Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala
Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His
Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly
Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285
Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290
295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn
Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val
Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val
Ser Ala Ser Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu
Phe Cys Leu Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr
His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu
Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu
His Ser His His Thr Asp Pro Arg Ser Gln Ser Glu Glu Pro Glu 405 410
415 Gly Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr Gln
420 425 430 Thr Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu
Glu Asp 435 440 445 Gln Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe
Val Gln Glu Asn 450 455 460 Gly Thr Leu Arg Ala Lys Pro Thr Gly Asn
Gly Ile Tyr Ile Asn Gly 465 470 475 480 Arg Gly His Leu Val 485 34
510 PRT Homo sapiens 34 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly
Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe
Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Gly Thr Ser Asp Val
Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe
Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp
Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu
Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90
95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val
100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu
Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg
Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn
Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu
Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val
Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser
Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His
Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215
220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu
225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu
Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met
Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn
Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val
Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His
Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser
Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335
Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340
345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val
Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln
Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu
Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp Pro
Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly
His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met
Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr
Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460
Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465
470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala
Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His
Leu Val 500 505 510 35 510 PRT Homo sapiens 35 Met Pro Leu Ser Leu
Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu
Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu
Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40
45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln
50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu
Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro
Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn
Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala
Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala
Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Met Val Pro
Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu
Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170
175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser
180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser
Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro
Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln
Arg Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu
Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile
Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln
Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu
Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295
300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu
305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu
Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser
Ala Ser Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe
Cys Leu Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His
Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu
Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His
Ser His His Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410 415
Gly Leu Arg Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420
425 430 Cys Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr
Leu 435 440 445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu
Ser Pro Gly 450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp
Glu Gly Ile Lys Gln 465 470 475 480 Ala Met Asn His Phe Val Gln Glu
Asn Gly Thr Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr
Ile Asn Gly Arg Gly His Leu Val 500 505 510 36 510 PRT Homo sapiens
36 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu
1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro
Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu
Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser
Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala
Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr
Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln
Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu
Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125
Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130
135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu
Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr
Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu
Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg
Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg
Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro
Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235 240 His
Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250
255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser
260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp
Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu
Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val
Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln
Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys
Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val Gly Val
Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360 365 Val
Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375
380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg
385 390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln Pro Glu
Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp Ser Leu
Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu Pro Glu
Gly Cys Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu Ile Glu
Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg Ala Glu
Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480 Ala Met
Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485 490 495
Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500 505 510
37 511 PRT Homo sapiens 37 Met Pro Leu Ser Leu Gly Ala Glu Met Trp
Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser
Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp
Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys
Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala
Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80
Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85
90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser
Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr
Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala
Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu
Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr
Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser
Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg
Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205
His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210
215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile
Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly
Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala
Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr
Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg
Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu
His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe
Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Ala Asp Pro 325 330
335 Gln Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val
340
345 350 Val Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val
Val 355 360 365 Val Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln
Gln Met Thr 370 375 380 Gln Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg
Glu Asn Ser Ile Arg 385 390 395 400 Arg Leu His Ser His His Thr Asp
Pro Arg Ser Gln Pro Glu Glu Ser 405 410 415 Val Gly Leu Arg Ala Glu
Gly His Pro Asp Ser Leu Lys Asp Asn Ser 420 425 430 Ser Cys Ser Val
Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr 435 440 445 Leu Thr
Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro 450 455 460
Gly Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys 465
470 475 480 Gln Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg
Ala Lys 485 490 495 Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly
His Leu Val 500 505 510 38 137 PRT Homo sapiens 38 Met Arg Arg Glu
Leu Leu Ala Gly Ile Leu Leu Arg Ile Thr Phe Asn 1 5 10 15 Phe Phe
Leu Phe Phe Phe Leu Pro Phe Pro Leu Val Val Phe Phe Ile 20 25 30
Tyr Phe Tyr Phe Tyr Phe Phe Leu Glu Met Glu Ser His Tyr Val Ala 35
40 45 Gln Ala Gly Leu Glu Leu Leu Gly Ser Ser Asn Pro Pro Ala Ser
Ala 50 55 60 Ser Leu Val Ala Gly Thr Leu Ser Val His His Cys Ala
Cys Phe Glu 65 70 75 80 Ser Phe Thr Lys Arg Lys Lys Lys Leu Lys Lys
Ala Phe Arg Phe Ile 85 90 95 Gln Cys Leu Leu Leu Gly Leu Leu Lys
Val Arg Pro Leu Gln His Gln 100 105 110 Gly Val Asn Ser Cys Asp Cys
Glu Arg Gly Tyr Phe Gln Gly Ile Phe 115 120 125 Met Gln Ala Ala Pro
Trp Glu Gly Thr 130 135 39 137 PRT Homo sapiens 39 Met Arg Arg Glu
Leu Leu Ala Gly Ile Leu Leu Arg Ile Thr Phe Asn 1 5 10 15 Phe Phe
Leu Phe Phe Phe Leu Pro Phe Pro Leu Val Val Phe Phe Ile 20 25 30
Tyr Phe Tyr Phe Tyr Phe Phe Leu Glu Met Glu Ser His Tyr Val Ala 35
40 45 Gln Ala Gly Leu Glu Leu Leu Gly Ser Ser Asn Pro Pro Ala Ser
Asp 50 55 60 Ser Leu Val Ala Gly Thr Leu Ser Val His His Cys Ala
Cys Phe Glu 65 70 75 80 Ser Phe Thr Lys Arg Lys Lys Lys Leu Lys Lys
Ala Phe Arg Phe Ile 85 90 95 Gln Cys Leu Leu Leu Gly Leu Leu Lys
Val Arg Pro Leu Gln His Gln 100 105 110 Gly Val Asn Ser Cys Asp Cys
Glu Arg Gly Tyr Phe Gln Gly Ile Phe 115 120 125 Met Gln Ala Ala Pro
Trp Glu Gly Thr 130 135 40 510 PRT Homo sapiens 40 Met Pro Leu Ser
Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu
Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30
Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35
40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly
Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln
Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser
Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg
Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln
Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro
Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val
Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160
Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165
170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr
Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr
Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln
Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp
Gln Arg Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala
Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His
Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly
Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285
Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290
295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn
Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val
Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val
Ser Ala Ser Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu
Phe Cys Leu Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr
His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu
Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu
His Ser His His Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410
415 Gly Leu Arg Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser
420 425 430 Cys Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser
Thr Leu 435 440 445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu
Leu Ser Pro Gly 450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln
Asp Glu Gly Ile Lys Gln 465 470 475 480 Ala Met Asn His Phe Val Gln
Glu Asn Gly Thr Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile
Tyr Ile Asn Gly Arg Gly His Leu Val 500 505 510 41 510 PRT Homo
sapiens 41 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala
Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg
Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val
Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly
Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val
Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser
Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val
Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110
Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115
120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu
Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro
Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser
Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp
Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His
Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro
Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser
His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235
240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln
245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys
Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg
Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp
Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile
Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp
Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser
Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val
Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360
365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln
370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile
Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln
Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp
Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu
Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu
Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg
Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480
Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485
490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500
505 510 42 510 PRT Homo sapiens 42 Met Pro Leu Ser Leu Gly Ala Glu
Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu
Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr
Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu
Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60
Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65
70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr
Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu
Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu
Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser
Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu
Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln
Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro
Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185
190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe
195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr
Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile
Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser
Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg
Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro
Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser
Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu
Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310
315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro
Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser
Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu
Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg
Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr
Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His
His Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu
Arg Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430
Cys Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435
440 445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro
Gly 450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly
Ile Lys Gln 465 470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly
Thr Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn
Gly Arg Gly His Leu Val 500 505 510 43 508 PRT Homo sapiens 43 Met
Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10
15 Arg Leu Leu Phe Leu Ala Ser Phe Thr Gly Gln Tyr Ser Ala Gly Glu
20 25 30 Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp
Ala Lys 35 40 45 Leu Pro Cys Phe Tyr Arg Gly Asp Pro Asp Glu Gln
Val Gly Gln Val 50 55 60 Ala Trp Ala Arg Val Asp Pro Asn Glu Gly
Ile Arg Glu Leu Ala Leu 65 70 75 80 Leu His Ser Lys Tyr Gly Leu His
Val Asn Pro Ala Tyr Glu Asp Arg 85 90 95 Val Glu Gln Pro Pro Pro
Pro Arg Asp Pro Leu Asp Gly Ser Val Leu 100 105 110 Leu Arg Asn Ala
Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg Val 115 120 125 Ser Thr
Phe Pro Ala Gly Ser Phe Gln Ala Arg Met Arg Leu Arg Val 130 135 140
Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Pro Leu Glu Glu 145
150 155 160 Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly
Ser Pro 165 170 175 Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly
Thr Gln Ser Ser 180 185 190 Arg Ser Phe Thr His Pro Arg Ser Ala Ala
Val Thr Ser Glu Phe His 195 200 205 Leu Val Pro Ser Arg Ser Met Asn
Gly Gln Pro Leu Thr Cys Val Val 210 215 220 Ser His Pro Gly Leu Leu
Gln Asp Arg Arg Ile Thr His Thr Leu Gln 225 230 235 240 Val Ala Phe
Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln Asn 245 250 255 Leu
Trp Gln Val Gly Arg Glu Gly Ala Thr Leu Lys Cys Leu Ser Glu 260 265
270 Gly Gln Pro Pro Pro Lys Tyr Asn Trp Thr Arg Leu Asp Gly Pro Leu
275 280 285 Pro Ser Gly Val Arg Val Lys Gly Asp Thr Leu Gly Phe Pro
Pro Leu 290 295 300 Thr Thr Glu His Ser Gly Val Tyr Val Cys His Val
Ser Asn Glu Leu 305 310 315 320 Ser Ser Arg Asp Ser Gln Val Thr Val
Glu Val Leu Asp Pro Glu Asp 325 330 335 Pro Gly Lys Gln Val Asp Leu
Val Ser Ala Ser Val Ile Ile Val Gly 340 345 350 Val Ile Ala Ala Leu
Leu Phe Cys Leu Leu Val Val Val Val Val Leu 355 360 365 Met Ser Arg
Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln Lys Tyr 370 375 380 Glu
Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg Leu His 385 390
395 400 Ser His His Ser Asp Pro Arg Ser Gln Pro Glu Glu Ser Val Gly
Leu 405 410 415 Arg Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser
Ser Cys Ser 420 425 430 Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr
Ser Thr Leu Thr Thr 435 440 445 Val Arg Glu Ile Glu Thr Gln Thr Glu
Leu Leu Ser Pro Gly Ser Gly 450
455 460 Arg Thr Glu Glu Asp Asp Asp Gln Asp Glu Gly Ile Lys Gln Ala
Met 465 470 475 480 Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala
Lys Pro Thr Gly 485 490 495 Asn Gly Ile Tyr Ile Asn Gly Arg Gly His
Leu Val 500 505 44 14 PRT Tetanus toxoid 44 Gln Tyr Ile Lys Ala Asn
Ser Lys Phe Ile Gly Ile Thr Glu 1 5 10 45 21 PRT Plasmodium
falciparum 45 Asp Ile Glu Lys Lys Ile Ala Lys Met Glu Lys Ala Ser
Ser Val Phe 1 5 10 15 Asn Val Val Asn Ser 20 46 16 PRT
Streptococcus 46 Gly Ala Val Asp Ser Ile Leu Gly Gly Val Ala Thr
Tyr Gly Ala Ala 1 5 10 15 47 13 PRT Artificial Sequence VARIANT 3
Xaa = cyclohexylalanine, phenylalanine, or tyrosine. 47 Xaa Lys Xaa
Val Ala Ala Trp Thr Leu Lys Ala Ala Xaa 1 5 10 48 14 DNA Artificial
Sequence Primer 48 ttttgatcaa gctt 14 49 42 DNA Artificial Sequence
Primer 49 ctaatacgac tcactatagg gctcgagcgg ccgcccgggc ag 42 50 12
DNA Artificial Sequence Primer 50 gatcctgccc gg 12 51 40 DNA
Artificial Sequence Primer 51 gtaatacgac tcactatagg gcagcgtggt
cgcggccgag 40 52 10 DNA Artificial Sequence Primer 52 gatcctcggc 10
53 22 DNA Artificial Sequence Primer 53 ctaatacgac tcactatagg gc 22
54 22 DNA Artificial Sequence Primer 54 tcgagcggcc gcccgggcag ga 22
55 20 DNA Artificial Sequence Primer 55 agcgtggtcg cggccgagga 20 56
25 DNA Artificial Sequence Primer 56 atatcgccgc gctcgtcgtc gacaa 25
57 26 DNA Artificial Sequence Primer 57 agccacacgc agctcattgt
agaagg 26 58 25 DNA Artificial Sequence Primer 58 ggctggagtt
caatgaggtt tattt 25 59 25 DNA Artificial Sequence Primer 59
tccagcagat ttcagactaa gaaga 25 60 24 DNA Artificial Sequence
Epitope Tag 60 gattacaagg atgacgacga taag 24 61 4 PRT Homo sapiens
61 Asn Trp Thr Arg 1 62 4 PRT Homo sapiens 62 Asn Ser Ser Cys 1 63
4 PRT Homo sapiens 63 Asn Gly Thr Leu 1 64 15 PRT Homo sapiens 64
Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg Val Ser Thr Phe 1 5 10
15 65 4 PRT Homo sapiens 65 Thr Arg Leu Asp 1 66 4 PRT Homo sapiens
66 Ser Ser Arg Asp 1 67 4 PRT Homo sapiens 67 Ser Gln Pro Glu 1 68
4 PRT Homo sapiens 68 Ser Leu Lys Asp 1 69 4 PRT Homo sapiens 69
Thr Val Arg Glu 1 70 4 PRT Homo sapiens 70 Thr Gln Thr Glu 1 71 6
PRT Homo sapiens 71 Gly Ser Phe Gln Ala Arg 1 5 72 6 PRT Homo
sapiens 72 Gly Gln Gly Leu Thr Leu 1 5 73 6 PRT Homo sapiens 73 Gly
Leu Thr Leu Ala Ala 1 5 74 6 PRT Homo sapiens 74 Gly Thr Thr Ser
Ser Arg 1 5 75 6 PRT Homo sapiens 75 Gly Gln Pro Leu Thr Cys 1 5 76
6 PRT Homo sapiens 76 Gly Ile Tyr Val Cys His 1 5 77 6 PRT Homo
sapiens 77 Gly Val Ile Ala Ala Leu 1 5 78 6 PRT Homo sapiens 78 Gly
Ser Gly Arg Ala Glu 1 5 79 6 PRT Homo sapiens 79 Gly Ile Lys Gln
Ala Met 1 5 80 6 PRT Homo sapiens 80 Gly Thr Leu Arg Ala Lys 1 5 81
6 PRT Homo sapiens 81 Gly Ile Tyr Ile Asn Gly 1 5 82 510 PRT Homo
sapiens 82 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala
Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg
Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val
Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly
Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val
Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser
Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val
Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110
Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115
120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu
Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro
Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser
Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp
Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His
Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro
Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser
His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235
240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln
245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys
Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg
Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp
Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile
Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp
Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser
Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val
Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360
365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln
370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile
Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln
Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp
Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu
Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu
Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg
Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480
Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485
490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500
505 510 83 17 PRT Homo sapiens 83 Gly Gln Asp Ala Lys Leu Pro Cys
Leu Tyr Arg Gly Asp Ser Gly Glu 1 5 10 15 Gln 84 19 PRT Homo
sapiens 84 Leu Gly Gln Asp Ala Lys Leu Pro Cys Leu Tyr Arg Gly Asp
Ser Gly 1 5 10 15 Glu Gln Val 85 29 PRT Homo sapiens 85 Val Val Thr
Val Val Leu Gly Gln Asp Ala Lys Leu Pro Cys Leu Tyr 1 5 10 15 Arg
Gly Asp Ser Gly Glu Gln Val Gly Gln Val Ala Trp 20 25 86 16 PRT
Homo sapiens 86 Ser His His Thr Asp Pro Arg Ser Gln Ser Glu Glu Pro
Glu Gly Arg 1 5 10 15 87 18 PRT Homo sapiens 87 His Ser His His Thr
Asp Pro Arg Ser Gln Ser Glu Glu Pro Glu Gly 1 5 10 15 Arg Ser 88 28
PRT Homo sapiens 88 Ser Ile Arg Arg Leu His Ser His His Thr Asp Pro
Arg Ser Gln Ser 1 5 10 15 Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr
Leu Thr 20 25 89 137 PRT Homo sapiens 89 Met Arg Arg Glu Leu Leu
Ala Gly Ile Leu Leu Arg Ile Thr Phe Asn 1 5 10 15 Phe Phe Leu Phe
Phe Phe Leu Pro Phe Pro Leu Val Val Phe Phe Ile 20 25 30 Tyr Phe
Tyr Phe Tyr Phe Phe Leu Glu Met Glu Ser His Tyr Val Ala 35 40 45
Gln Ala Gly Leu Glu Leu Leu Gly Ser Ser Asn Pro Pro Ala Ser Ala 50
55 60 Ser Leu Val Ala Gly Thr Leu Ser Val His His Cys Ala Cys Phe
Glu 65 70 75 80 Ser Phe Thr Lys Arg Lys Lys Lys Leu Lys Lys Ala Phe
Arg Phe Ile 85 90 95 Gln Cys Leu Leu Leu Gly Leu Leu Lys Val Arg
Pro Leu Gln His Gln 100 105 110 Gly Val Asn Ser Cys Asp Cys Glu Arg
Gly Tyr Phe Gln Gly Ile Phe 115 120 125 Met Gln Ala Ala Pro Trp Glu
Gly Thr 130 135 90 17 PRT Homo sapiens 90 Gly Arg Cys Pro Ala Gly
Glu Leu Gly Thr Ser Asp Val Val Thr Val 1 5 10 15 Val 91 19 PRT
Homo sapiens 91 Thr Gly Arg Cys Pro Ala Gly Glu Leu Gly Thr Ser Asp
Val Val Thr 1 5 10 15 Val Val Leu 92 29 PRT Homo sapiens 92 Leu Leu
Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly Glu Leu Gly Thr 1 5 10 15
Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala Lys 20 25 93 17 PRT
Homo sapiens 93 Gln Ala Arg Leu Arg Leu Arg Val Met Val Pro Pro Leu
Pro Ser Leu 1 5 10 15 Asn 94 19 PRT Homo sapiens 94 Phe Gln Ala Arg
Leu Arg Leu Arg Val Met Val Pro Pro Leu Pro Ser 1 5 10 15 Leu Asn
Pro 95 29 PRT Homo sapiens 95 Phe Pro Ala Gly Ser Phe Gln Ala Arg
Leu Arg Leu Arg Val Met Val 1 5 10 15 Pro Pro Leu Pro Ser Leu Asn
Pro Gly Pro Ala Leu Glu 20 25 96 17 PRT Homo sapiens 96 Val Met Ser
Glu Glu Pro Glu Gly Cys Ser Tyr Ser Thr Leu Thr Thr 1 5 10 15 Val
97 20 PRT Homo sapiens 97 Ser Val Met Ser Glu Glu Pro Glu Gly Cys
Ser Tyr Ser Thr Leu Thr 1 5 10 15 Thr Val Arg Glu 20 98 29 PRT Homo
sapiens 98 Asp Asn Ser Ser Cys Ser Val Met Ser Glu Glu Pro Glu Gly
Cys Ser 1 5 10 15 Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr
Gln 20 25 99 17 PRT Homo sapiens 99 Ser Gln Val Thr Val Asp Val Leu
Ala Asp Pro Gln Glu Asp Ser Gly 1 5 10 15 Lys 100 19 PRT Homo
sapiens 100 Asp Ser Gln Val Thr Val Asp Val Leu Ala Asp Pro Gln Glu
Asp Ser 1 5 10 15 Gly Lys Gln 101 29 PRT Homo sapiens 101 Glu Phe
Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Ala Asp 1 5 10 15
Pro Gln Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser 20 25 102 17
PRT Homo sapiens 102 Gly Ser Ser Asn Pro Pro Ala Ser Ala Ser Leu
Val Ala Gly Thr Leu 1 5 10 15 Ser 103 19 PRT Homo sapiens 103 Leu
Gly Ser Ser Asn Pro Pro Ala Ser Ala Ser Leu Val Ala Gly Thr 1 5 10
15 Leu Ser Val 104 29 PRT Homo sapiens 104 Ala Gly Leu Glu Leu Leu
Gly Ser Ser Asn Pro Pro Ala Ser Ala Ser 1 5 10 15 Leu Val Ala Gly
Thr Leu Ser Val His His Cys Ala Cys 20 25 105 3344 DNA Homo sapiens
105 ggccgtcgtt gttggccaca gcgtgggaag cagctctggg ggagctcgga
gctcccgatc 60 acggcttctt gggggtagct acggctgggt gtgtagaacg
gggccggggc tggggctggg 120 tcccctagtg gagacccaag tgcgagaggc
aagaactctg cagcttcctg ccttctgggt 180 cagttcctta ttcaagtctg
ctactgctgg catcatttac aggccggtgc cccgcgggtg 240 agctggagac
ctcagacgtg gtaactgtgg tgctgggcca ggacgcaaaa ctgccctgct 300
tctaccgagg ggactccggc gagcaagtgg ggcaagtggc atgggctcgg gtggacgcgg
360 gcgaaggcgc ccaggaacta gcgctactgc actccaaata cgggcttcat
gtgagcccgg 420 cttacgaggg ccgcgtggag cagccgccgc ccccacgcaa
ccccctggac ggctcagtgc 480 tcctgcgcaa cgcagtgcag gcggatgagg
gcgagtacga gtgccgggtc agcaccttcc 540 ccgccggcag cttccaggcg
cggctgcggc tccgagtgct ggtgcctccc ctgccctcac 600 tgaatcctgg
tccagcacta gaagagggcc agggcctgac cctggcagcc tcctgcacag 660
ctgagggcag cccagccccc agcgtgacct gggacacgga ggtcaaaggc acaacgtcca
720 gccgttcctt caagcactcc cgctctgctg ccgtcacctc agagttccac
ttggtgccta 780 gccgcagcat gaatgggcag ccactgactt gtgtggtgtc
ccatcctggc ctgctccagg 840 accaaaggat cacccacatc ctccacgtgt
ccttccttgc tgaggcctct gtgaggggcc 900 ttgaagacca aaatctgtgg
cacattggca gagaaggagc tatgctcaag tgcctgagtg 960 aagggcagcc
ccctccctca tacaactgga cacggctgga tgggcctctg cccagtgggg 1020
tacgagtgga tggggacact ttgggctttc ccccactgac cactgagcac agcggcatct
1080 acgtctgcca tgtcagcaat gagttctcct caagggattc tcaggtcact
gtggatgttc 1140 ttgaccccca ggaagactct gggaagcagg tggacctagt
gtcagcctcg gtggtggtgg 1200 tgggtgtgat cgccgcactc ttgttctgcc
ttctggtggt ggtggtggtg ctcatgtccc 1260 gataccatcg gcgcaaggcc
cagcagatga cccagaaata tgaggaggag ctgaccctga 1320 ccagggagaa
ctccatccgg aggctgcatt cccatcacac ggaccccagg agccagccgg 1380
aggagagtgt agggctgaga gccgagggcc accctgatag tctcaaggac aacagtagct
1440 gctctgtgat gagtgaagag cccgagggcc gcagttactc cacgctgacc
acggtgaggg 1500 agatagaaac acagactgaa ctgctgtctc caggctctgg
gcgggccgag gaggaggaag 1560 atcaggatga aggcatcaaa caggccatga
accattttgt tcaggagaat gggaccctac 1620 gggccaagcc cacgggcaat
ggcatctaca tcaatgggcg gggacacctg gtctgaccca 1680 ggcctgcctc
ccttccctag gcctggctcc ttctgttgac atgggagatt ttagctcatc 1740
ttgggggcct ccttaaacac ccccatttct tgcggaagat gctccccatc ccactgactg
1800 cttgaccttt acctccaacc cttctgttca tcgggagggc tccaccaatt
gagtctctcc 1860 caccatgcat gcaggtcact gtgtgtgtgc atgtgtgcct
gtgtgagtgt tgactgactg 1920 tgtgtgtgtg gaggggtgac tgtccgtgga
ggggtgactg tgtccgtggt gtgtattatg 1980 ctgtcatatc agagtcaagt
gaactgtggt gtatgtgcca cgggatttga gtggttgcgt 2040 gggcaacact
gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc tctgcctgaa 2100
aaagcaggta ttttctcaga ccccagagca gtattaatga tgcagaggtt ggaggagaga
2160 ggtggagact gtggctcaga cccaggtgtg cgggcatagc tggagctgga
atctgcctcc 2220 ggtgtgaggg aacctgtctc ctaccacttc ggagccatgg
gggcaagtgt gaagcagcca 2280 gtccctgggt cagccagagg cttgaactgt
tacagaagcc ctctgccctc tggtggcctc 2340 tgggcctgct gcatgtacat
attttctgta aatatacatg cgccgggagc ttcttgcagg 2400 aatactgctc
cgaatcactt ttaatttttt tctttttttt ttcttgccct ttccattagt 2460
tgtatttttt atttattttt atttttattt ttttttagag atggagtctc actatgttgc
2520 tcaggctggc cttgaactcc tgggctcaag caatcctcct gcctcagcct
ccctagtagc 2580 tgggacttta agtgtacacc actgtgcctg ctttgaatcc
tttacgaaga gaaaaaaaaa 2640 attaaagaaa gcctttagat ttatccaatg
tttactactg ggattgctta aagtgaggcc 2700 cctccaacac cagggggtta
attcctgtga ttgtgaaagg ggctacttcc aaggcatctt 2760 catgcaggca
gccccttggg agggcacctg agagctggta gagtctgaaa ttagggatgt 2820
gagcctcgtg gttactgagt aaggtaaaat tgcatccacc attgtttgtg ataccttagg
2880 gaattgcttg gacctggtga caagggctcc tgttcaatag tggtgttggg
gagagagaga 2940 gcagtgatta tagaccgaga gagtaggagt tgaggtgagg
tgaaggaggt gctgggggtg 3000 agaatgtcgc ctttccccct gggttttgga
tcactaattc aaggctcttc tggatgtttc 3060 tctgggttgg ggctggagtt
caatgaggtt tatttttagc tggcccaccc agatacactc 3120 agccagaata
cctagattta gtacccaaac tcttcttagt ctgaaatctg ctggatttct 3180
ggcctaaggg agaggctccc atccttcgtt ccccagccag cctaggactt cgaatgtgga
3240 gcctgaagat ctaagatcct aacatgtaca ttttatgtaa atatgtgcat
atttgtacat 3300 aaaatgatat tctgttttta aataaacaga caaaacttga aaaa
3344 106 3464 DNA Homo sapiens 106 ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac
240 gctgggcagt ctgcctttca accatgcccc tgtccctggg agccgagatg
tgggggcctg 300 aggcctggct gctgctgctg ctactgctgg catcatttac
aggccggtgc cccgcgggtg 360 agctggagac ctcagacgtg gtaactgtgg
tgctgggcca ggacgcaaaa ctgccctgct 420 tctaccgagg ggactccggc
gagcaagtgg ggcaagtggc atgggctcgg gtggacgcgg 480 gcgaaggcgc
ccaggaacta gcgctactgc actccaaata cgggcttcat gtgagcccgg 540
cttacgaggg ccgcgtggag cagccgccgc ccccacgcaa ccccctggac ggctcagtgc
600 tcctgcgcaa cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc
agcaccttcc 660 ccgccggcag cttccaggcg cggctgcggc tccgagtgct
ggtgcctccc ctgccctcac 720 tgaatcctgg tccagcacta gaagagggcc
agggcctgac cctggcagcc tcctgcacag 780 ctgagggcag cccagccccc
agcgtgacct gggacacgga ggtcaaaggc acaacgtcca 840 gccgttcctt
caagcactcc cgctctgctg ccgtcacctc agagttccac ttggtgccta 900
gccgcagcat gaatgggcag ccactgactt gtgtggtgtc ccatcctggc ctgctccagg
960 accaaaggat cacccacatc ctccacgtgt ccttccttgc tgaggcctct
gtgaggggcc 1020 ttgaagacca aaatctgtgg cacattggca gagaaggagc
tatgctcaag tgcctgagtg 1080 aagggcagcc ccctccctca tacaactgga
cacggctgga tgggcctctg cccagtgggg 1140 tacgagtgga tggggacact
ttgggctttc ccccactgac cactgagcac agcggcatct 1200 acgtctgcca
tgtcagcaat gagttctcct caagggattc
tcaggtcact gtggatgttc 1260 ttgaccccca ggaagactct gggaagcagg
tggacctagt gtcagcctcg gtggtggtgg 1320 tgggtgtgat cgccgcactc
ttgttctgcc ttctggtggt ggtggtggtg ctcatgtccc 1380 gataccatcg
gcgcaaggcc cagcagatga cccagaaata tgaggaggag ctgaccctga 1440
ccagggagaa ctccatccgg aggctgcatt cccatcacac ggaccccagg agccagccgg
1500 aggagagtgt agggctgaga gccgagggcc accctgatag tctcaaggac
aacagtagct 1560 gctctgtgat gagtgaagag cccgagggcc gcagttactc
cacgctgacc acggtgaggg 1620 agatagaaac acagactgaa ctgctgtctc
caggctctgg gcgggccgag gaggaggaag 1680 atcaggatga aggcatcaaa
caggccatga accattttgt tcaggagaat gggaccctac 1740 gggccaagcc
cacgggcaat ggcatctaca tcaatgggcg gggacacctg gtctgaccca 1800
ggcctgcctc ccttccctag gcctggctcc ttctgttgac atgggagatt ttagctcatc
1860 ttgggggcct ccttaaacac ccccatttct tgcggaagat gctccccatc
ccactgactg 1920 cttgaccttt acctccaacc cttctgttca tcgggagggc
tccaccaatt gagtctctcc 1980 caccatgcat gcaggtcact gtgtgtgtgc
atgtgtgcct gtgtgagtgt tgactgactg 2040 tgtgtgtgtg gaggggtgac
tgtccgtgga ggggtgactg tgtccgtggt gtgtattatg 2100 ctgtcatatc
agagtcaagt gaactgtggt gtatgtgcca cgggatttga gtggttgcgt 2160
gggcaacact gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc tctgcctgaa
2220 aaagcaggta ttttctcaga ccccagagca gtattaatga tgcagaggtt
ggaggagaga 2280 ggtggagact gtggctcaga cccaggtgtg cgggcatagc
tggagctgga atctgcctcc 2340 ggtgtgaggg aacctgtctc ctaccacttc
ggagccatgg gggcaagtgt gaagcagcca 2400 gtccctgggt cagccagagg
cttgaactgt tacagaagcc ctctgccctc tggtggcctc 2460 tgggcctgct
gcatgtacat attttctgta aatatacatg cgccgggagc ttcttgcagg 2520
aatactgctc cgaatcactt ttaatttttt tctttttttt ttcttgccct ttccattagt
2580 tgtatttttt atttattttt atttttattt ttttttagag atggagtctc
actatgttgc 2640 tcaggctggc cttgaactcc tgggctcaag caatcctcct
gcctcagcct ccctagtagc 2700 tgggacttta agtgtacacc actgtgcctg
ctttgaatcc tttacgaaga gaaaaaaaaa 2760 attaaagaaa gcctttagat
ttatccaatg tttactactg ggattgctta aagtgaggcc 2820 cctccaacac
cagggggtta attcctgtga ttgtgaaagg ggctacttcc aaggcatctt 2880
catgcaggca gccccttggg agggcacctg agagctggta gagtctgaaa ttagggatgt
2940 gagcctcgtg gttactgagt aaggtaaaat tgcatccacc attgtttgtg
ataccttagg 3000 gaattgcttg gacctggtga caagggctcc tgttcaatag
tggtgttggg gagagagaga 3060 gcagtgatta tagaccgaga gagtaggagt
tgaggtgagg tgaaggaggt gctgggggtg 3120 agaatgtcgc ctttccccct
gggttttgga tcactaattc aaggctcttc tggatgtttc 3180 tctgggttgg
ggctggagtt caatgaggtt tatttttagc tggcccaccc agatacactc 3240
agccagaata cctagattta gtacccaaac tcttcttagt ctgaaatctg ctggatttct
3300 ggcctaaggg agaggctccc atccttcgtt ccccagccag cctaggactt
cgaatgtgga 3360 gcctgaagat ctaagatcct aacatgtaca ttttatgtaa
atatgtgcat atttgtacat 3420 aaaatgatat tctgttttta aataaacaga
caaaacttga aaaa 3464 107 3344 DNA Homo sapiens 107 ggccgtcgtt
gttggccaca gcgtgggaag cagctctggg ggagctcgga gctcccgatc 60
acggcttctt gggggtagct acggctgggt gtgtagaacg gggccggggc tggggctggg
120 tcccctagtg gagacccaag tgcgagaggc aagaactctg cagcttcctg
ccttctgggt 180 cagttcctta ttcaagtctg ctactgctgg catcatttac
aggccggtgc cccgcgggtg 240 agctggagac ctcagacgtg gtaactgtgg
tgctgggcca ggacgcaaaa ctgccctgct 300 tctaccgagg ggactccggc
gagcaagtgg ggcaagtggc atgggctcgg gtggacgcgg 360 gcgaaggcgc
ccaggaacta gcgctactgc actccaaata cgggcttcat gtgagcccgg 420
cttacgaggg ccgcgtggag cagccgccgc ccccacgcaa ccccctggac ggctcagtgc
480 tcctgcgcaa cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc
agcaccttcc 540 ccgccggcag cttccaggcg cggctgcggc tccgagtgct
ggtgcctccc ctgccctcac 600 tgaatcctgg tccagcacta gaagagggcc
agggcctgac cctggcagcc tcctgcacag 660 ctgagggcag cccagccccc
agcgtgacct gggacacgga ggtcaaaggc acaacgtcca 720 gccgttcctt
caagcactcc cgctctgctg ccgtcacctc agagttccac ttggtgccta 780
gccgcagcat gaatgggcag ccactgactt gtgtggtgtc ccatcctggc ctgctccagg
840 accaaaggat cacccacatc ctccacgtgt ccttccttgc tgaggcctct
gtgaggggcc 900 ttgaagacca aaatctgtgg cacattggca gagaaggagc
tatgctcaag tgcctgagtg 960 aagggcagcc ccctccctca tacaactgga
cacggctgga tgggcctctg cccagtgggg 1020 tacgagtgga tggggacact
ttgggctttc ccccactgac cactgagcac agcggcatct 1080 acgtctgcca
tgtcagcaat gagttctcct caagggattc tcaggtcact gtggatgttc 1140
ttgaccccca ggaagactct gggaagcagg tggacctagt gtcagcctcg gtggtggtgg
1200 tgggtgtgat cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg
ctcatgtccc 1260 gataccatcg gcgcaaggcc cagcagatga cccagaaata
tgaggaggag ctgaccctga 1320 ccagggagaa ctccatccgg aggctgcatt
cccatcacac ggaccccagg agccagccgg 1380 aggagagtgt agggctgaga
gccgagggcc accctgatag tctcaaggac aacagtagct 1440 gctctgtgat
gagtgaagag cccgagggcc gcagttactc cacgctgacc acggtgaggg 1500
agatagaaac acagactgaa ctgctgtctc caggctctgg gcgggccgag gaggaggaag
1560 atcaggatga aggcatcaaa caggccatga accattttgt tcaggagaat
gggaccctac 1620 gggccaagcc cacgggcaat ggcatctaca tcaatgggcg
gggacacctg gtctgaccca 1680 ggcctgcctc ccttccctag gcctggctcc
ttctgttgac atgggagatt ttagctcatc 1740 ttgggggcct ccttaaacac
ccccatttct tgcggaagat gctccccatc ccactgactg 1800 cttgaccttt
acctccaacc cttctgttca tcgggagggc tccaccaatt gagtctctcc 1860
caccatgcat gcaggtcact gtgtgtgtgc atgtgtgcct gtgtgagtgt tgactgactg
1920 tgtgtgtgtg gaggggtgac tgtccgtgga ggggtgactg tgtccgtggt
gtgtattatg 1980 ctgtcatatc agagtcaagt gaactgtggt gtatgtgcca
cgggatttga gtggttgcgt 2040 gggcaacact gtcagggttt ggcgtgtgtg
tcatgtggct gtgtgtgacc tctgcctgaa 2100 aaagcaggta ttttctcaga
ccccagagca gtattaatga tgcagaggtt ggaggagaga 2160 ggtggagact
gtggctcaga cccaggtgtg cgggcatagc tggagctgga atctgcctcc 2220
ggtgtgaggg aacctgtctc ctaccacttc ggagccatgg gggcaagtgt gaagcagcca
2280 gtccctgggt cagccagagg cttgaactgt tacagaagcc ctctgccctc
tggtggcctc 2340 tgggcctgct gcatgtacat attttctgta aatatacatg
cgccgggagc ttcttgcagg 2400 aatactgctc cgaatcactt ttaatttttt
tctttttttt ttcttgccct ttccattagt 2460 tgtatttttt atttattttt
atttttattt ttttttagag atggagtctc actatgttgc 2520 tcaggctggc
cttgaactcc tgggctcaag caatcctcct gcctcagcct ccctagtagc 2580
tgggacttta agtgtacacc actgtgcctg ctttgaatcc tttacgaaga gaaaaaaaaa
2640 attaaagaaa gcctttagat ttatccaatg tttactactg ggattgctta
aagtgaggcc 2700 cctccaacac cagggggtta attcctgtga ttgtgaaagg
ggctacttcc aaggcatctt 2760 catgcaggca gccccttggg agggcacctg
agagctggta gagtctgaaa ttagggatgt 2820 gagcctcgtg gttactgagt
aaggtaaaat tgcatccacc attgtttgtg ataccttagg 2880 gaattgcttg
gacctggtga caagggctcc tgttcaatag tggtgttggg gagagagaga 2940
gcagtgatta tagaccgaga gagtaggagt tgaggtgagg tgaaggaggt gctgggggtg
3000 agaatgtcgc ctttccccct gggttttgga tcactaattc aaggctcttc
tggatgtttc 3060 tctgggttgg ggctggagtt caatgaggtt tatttttagc
tggcccaccc agatacactc 3120 agccagaata cctagattta gtacccaaac
tcttcttagt ctgaaatctg ctggatttct 3180 ggcctaaggg agaggctccc
atccttcgtt ccccagccag cctaggactt cgaatgtgga 3240 gcctgaagat
ctaagatcct aacatgtaca ttttatgtaa atatgtgcat atttgtacat 3300
aaaatgatat tctgttttta aataaacaga caaaacttga aaaa 3344 108 295 PRT
Homo sapiens 108 Met Asn Gly Gln Pro Leu Thr Cys Val Val Ser His
Pro Gly Leu Leu 1 5 10 15 Gln Asp Gln Arg Ile Thr His Ile Leu His
Val Ser Phe Leu Ala Glu 20 25 30 Ala Ser Val Arg Gly Leu Glu Asp
Gln Asn Leu Trp His Ile Gly Arg 35 40 45 Glu Gly Ala Met Leu Lys
Cys Leu Ser Glu Gly Gln Pro Pro Pro Ser 50 55 60 Tyr Asn Trp Thr
Arg Leu Asp Gly Pro Leu Pro Ser Gly Val Arg Val 65 70 75 80 Asp Gly
Asp Thr Leu Gly Phe Pro Pro Leu Thr Thr Glu His Ser Gly 85 90 95
Ile Tyr Val Cys His Val Ser Asn Glu Phe Ser Ser Arg Asp Ser Gln 100
105 110 Val Thr Val Asp Val Leu Asp Pro Gln Glu Asp Ser Gly Lys Gln
Val 115 120 125 Asp Leu Val Ser Ala Ser Val Val Val Val Gly Val Ile
Ala Ala Leu 130 135 140 Leu Phe Cys Leu Leu Val Val Val Val Val Leu
Met Ser Arg Tyr His 145 150 155 160 Arg Arg Lys Ala Gln Gln Met Thr
Gln Lys Tyr Glu Glu Glu Leu Thr 165 170 175 Leu Thr Arg Glu Asn Ser
Ile Arg Arg Leu His Ser His His Thr Asp 180 185 190 Pro Arg Ser Gln
Pro Glu Glu Ser Val Gly Leu Arg Ala Glu Gly His 195 200 205 Pro Asp
Ser Leu Lys Asp Asn Ser Ser Cys Ser Val Met Ser Glu Glu 210 215 220
Pro Glu Gly Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu 225
230 235 240 Thr Gln Thr Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu
Glu Glu 245 250 255 Glu Asp Gln Asp Glu Gly Ile Lys Gln Ala Met Asn
His Phe Val Gln 260 265 270 Glu Asn Gly Thr Leu Arg Ala Lys Pro Thr
Gly Asn Gly Ile Tyr Ile 275 280 285 Asn Gly Arg Gly His Leu Val 290
295 109 295 PRT Homo sapiens 109 Met Asn Gly Gln Pro Leu Thr Cys
Val Val Ser His Pro Gly Leu Leu 1 5 10 15 Gln Asp Gln Arg Ile Thr
His Ile Leu His Val Ser Phe Leu Ala Glu 20 25 30 Ala Ser Val Arg
Gly Leu Glu Asp Gln Asn Leu Trp His Ile Gly Arg 35 40 45 Glu Gly
Ala Met Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro Pro Ser 50 55 60
Tyr Asn Trp Thr Arg Leu Asp Gly Pro Leu Pro Ser Gly Val Arg Val 65
70 75 80 Asp Gly Asp Thr Leu Gly Phe Pro Pro Leu Thr Thr Glu His
Ser Gly 85 90 95 Ile Tyr Val Cys His Val Ser Asn Glu Phe Ser Ser
Arg Asp Ser Gln 100 105 110 Val Thr Val Asp Val Leu Asp Pro Gln Glu
Asp Ser Gly Lys Gln Val 115 120 125 Asp Leu Val Ser Ala Ser Val Val
Val Val Gly Val Ile Ala Ala Leu 130 135 140 Leu Phe Cys Leu Leu Val
Val Val Val Val Leu Met Ser Arg Tyr His 145 150 155 160 Arg Arg Lys
Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu Thr 165 170 175 Leu
Thr Arg Glu Asn Ser Ile Arg Arg Leu His Ser His His Thr Asp 180 185
190 Pro Arg Ser Gln Pro Glu Glu Ser Val Gly Leu Arg Ala Glu Gly His
195 200 205 Pro Asp Ser Leu Lys Asp Asn Ser Ser Cys Ser Val Met Ser
Glu Glu 210 215 220 Pro Glu Gly Arg Ser Tyr Ser Thr Leu Thr Thr Val
Arg Glu Ile Glu 225 230 235 240 Thr Gln Thr Glu Leu Leu Ser Pro Gly
Ser Gly Arg Ala Glu Glu Glu 245 250 255 Glu Asp Gln Asp Glu Gly Ile
Lys Gln Ala Met Asn His Phe Val Gln 260 265 270 Glu Asn Gly Thr Leu
Arg Ala Lys Pro Thr Gly Asn Gly Ile Tyr Ile 275 280 285 Asn Gly Arg
Gly His Leu Val 290 295 110 295 PRT Homo sapiens 110 Met Asn Gly
Gln Pro Leu Thr Cys Val Val Ser His Pro Gly Leu Leu 1 5 10 15 Gln
Asp Gln Arg Ile Thr His Ile Leu His Val Ser Phe Leu Ala Glu 20 25
30 Ala Ser Val Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile Gly Arg
35 40 45 Glu Gly Ala Met Leu Lys Cys Leu Ser Glu Gly Gln Pro Pro
Pro Ser 50 55 60 Tyr Asn Trp Thr Arg Leu Asp Gly Pro Leu Pro Ser
Gly Val Arg Val 65 70 75 80 Asp Gly Asp Thr Leu Gly Phe Pro Pro Leu
Thr Thr Glu His Ser Gly 85 90 95 Ile Tyr Val Cys His Val Ser Asn
Glu Phe Ser Ser Arg Asp Ser Gln 100 105 110 Val Thr Val Asp Val Leu
Asp Pro Gln Glu Asp Ser Gly Lys Gln Val 115 120 125 Asp Leu Val Ser
Ala Ser Val Val Val Val Gly Val Ile Ala Ala Leu 130 135 140 Leu Phe
Cys Leu Leu Val Val Val Val Val Leu Met Ser Arg Tyr His 145 150 155
160 Arg Arg Lys Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu Thr
165 170 175 Leu Thr Arg Glu Asn Ser Ile Arg Arg Leu His Ser His His
Thr Asp 180 185 190 Pro Arg Ser Gln Pro Glu Glu Ser Val Gly Leu Arg
Ala Glu Gly His 195 200 205 Pro Asp Ser Leu Lys Asp Asn Ser Ser Cys
Ser Val Met Ser Glu Glu 210 215 220 Pro Glu Gly Arg Ser Tyr Ser Thr
Leu Thr Thr Val Arg Glu Ile Glu 225 230 235 240 Thr Gln Thr Glu Leu
Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu 245 250 255 Glu Asp Gln
Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val Gln 260 265 270 Glu
Asn Gly Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile Tyr Ile 275 280
285 Asn Gly Arg Gly His Leu Val 290 295 111 3389 DNA Homo sapiens
111 ggccgtcgtt gttggccaca gcgtgggaag cagctctggg ggagctcgga
gctcccgatc 60 acggcttctt gggggtagct acggctgggt gtgtagaacg
gggccggggc tggggctggg 120 tcccctagtg gagacccaag tgcgagaggc
aagaactctg cagcttcctg ccttctgggt 180 cagttcctta ttcaagtctg
cagccggctc ccagggagat ctcggtggaa cttcagaaac 240 gctgggcagt
ctgcctttca accatgcccc tgtccctggg agccgagatg tgggggcctg 300
aggcctggct gctgctgctg ctactgctgg catcatttac aggccggtgc cccgcgggtg
360 agctggagac ctcagacgtg gtaactgtgg tgctgggcca ggacgcaaaa
ctgccctgct 420 tctaccgagg ggactccggc gagcaagtgg ggcaagtggc
atgggctcgg gtggacgcgg 480 gcgaaggcgc ccaggaacta gcgctactgc
actccaaata cgggcttcat gtgagcccgg 540 cttacgaggg ccgcgtggag
cagccgccgc ccccacgcaa ccccctggac ggctcagtgc 600 tcctgcgcaa
cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc agcaccttcc 660
ccgccggcag cttccaggcg cggctgcggc tccgagtgct ggtgcctccc ctgccctcac
720 tgaatcctgg tccagcacta gaagagggcc agggcctgac cctggcagcc
tcctgcacag 780 ctgagggcag cccagccccc agcgtgacct gggacacgga
ggtcaaaggc acaacgtcca 840 gccgttcctt caagcactcc cgctctgctg
ccgtcacctc agagttccac ttggtgccta 900 gccgcagcat gaatgggcag
ccactgactt gtgtggtgtc ccatcctggc ctgctccagg 960 accaaaggat
cacccacatc ctccacgtgt ccttccttgc tgaggcctct gtgaggggcc 1020
ttgaagacca aaatctgtgg cacattggca gagaaggagc tatgctcaag tgcctgagtg
1080 aagggcagcc ccctccctca tacaactgga cacggctgga tgggcctctg
cccagtgggg 1140 tacgagtgga tggggacact ttgggctttc ccccactgac
cactgagcac agcggcatct 1200 acgtctgcca tgtcagcaat gagttctcct
caagggattc tcaggtcact gtggatgttc 1260 ttgaccccca ggaagactct
gggaagcagg tggacctagt gtcagcctcg gtggtggtgg 1320 tgggtgtgat
cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg ctcatgtccc 1380
gataccatcg gcgcaaggcc cagcagatga cccagaaata tgaggaggag ctgaccctga
1440 ccagggagaa ctccatccgg aggctgcatt cccatcacac ggaccccagg
agccagagtg 1500 aagagcccga gggccgcagt tactccacgc tgaccacggt
gagggagata gaaacacaga 1560 ctgaactgct gtctccaggc tctgggcggg
ccgaggagga ggaagatcag gatgaaggca 1620 tcaaacaggc catgaaccat
tttgttcagg agaatgggac cctacgggcc aagcccacgg 1680 gcaatggcat
ctacatcaat gggcggggac acctggtctg acccaggcct gcctcccttc 1740
cctaggcctg gctccttctg ttgacatggg agattttagc tcatcttggg ggcctcctta
1800 aacaccccca tttcttgcgg aagatgctcc ccatcccact gactgcttga
cctttacctc 1860 caacccttct gttcatcggg agggctccac caattgagtc
tctcccacca tgcatgcagg 1920 tcactgtgtg tgtgcatgtg tgcctgtgtg
agtgttgact gactgtgtgt gtgtggaggg 1980 gtgactgtcc gtggaggggt
gactgtgtcc gtggtgtgta ttatgctgtc atatcagagt 2040 caagtgaact
gtggtgtatg tgccacggga tttgagtggt tgcgtgggca acactgtcag 2100
ggtttggcgt gtgtgtcatg tggctgtgtg tgacctctgc ctgaaaaagc aggtattttc
2160 tcagacccca gagcagtatt aatgatgcag aggttggagg agagaggtgg
agactgtggc 2220 tcagacccag gtgtgcgggc atagctggag ctggaatctg
cctccggtgt gagggaacct 2280 gtctcctacc acttcggagc catgggggca
agtgtgaagc agccagtccc tgggtcagcc 2340 agaggcttga actgttacag
aagccctctg ccctctggtg gcctctgggc ctgctgcatg 2400 tacatatttt
ctgtaaatat acatgcgccg ggagcttctt gcaggaatac tgctccgaat 2460
cacttttaat ttttttcttt tttttttctt gccctttcca ttagttgtat tttttattta
2520 tttttatttt tatttttttt tagagatgga gtctcactat gttgctcagg
ctggccttga 2580 actcctgggc tcaagcaatc ctcctgcctc agcctcccta
gtagctggga ctttaagtgt 2640 acaccactgt gcctgctttg aatcctttac
gaagagaaaa aaaaaattaa agaaagcctt 2700 tagatttatc caatgtttac
tactgggatt gcttaaagtg aggcccctcc aacaccaggg 2760 ggttaattcc
tgtgattgtg aaaggggcta cttccaaggc atcttcatgc aggcagcccc 2820
ttgggagggc acctgagagc tggtagagtc tgaaattagg gatgtgagcc tcgtggttac
2880 tgagtaaggt aaaattgcat ccaccattgt ttgtgatacc ttagggaatt
gcttggacct 2940 ggtgacaagg gctcctgttc aatagtggtg ttggggagag
agagagcagt gattatagac 3000 cgagagagta ggagttgagg tgaggtgaag
gaggtgctgg gggtgagaat gtcgcctttc 3060 cccctgggtt ttggatcact
aattcaaggc tcttctggat gtttctctgg gttggggctg 3120 gagttcaatg
aggtttattt ttagctggcc cacccagata cactcagcca gaatacctag 3180
atttagtacc caaactcttc ttagtctgaa atctgctgga tttctggcct aagggagagg
3240 ctcccatcct tcgttcccca gccagcctag gacttcgaat gtggagcctg
aagatctaag 3300 atcctaacat gtacatttta tgtaaatatg tgcatatttg
tacataaaat gatattctgt 3360 ttttaaataa acagacaaaa cttgaaaaa 3389 112
3464 DNA Homo sapiens 112 ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac
240 gctgggcagt ctgcctttca accatgcccc tgtccctggg agccgagatg
tgggggcctg 300 aggcctggct gctgctgctg ctactgctgg catcatttac
aggccggtgc cccgcgggtg 360 agctggagac ctcagacgtg gtaactgtgg
tgctgggcca ggacgcaaaa ctgccctgct 420 tctaccgagg ggactccggc
gagcaagtgg ggcaagtggc atgggctcgg gtggacgcgg 480 gcgaaggcgc
ccaggaacta gcgctactgc actccaaata cgggcttcat gtgagcccgg 540
cttacgaggg ccgcgtggag cagccgccgc ccccacgcaa ccccctggac ggctcagtgc
600 tcctgcgcaa cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc
agcaccttcc 660 ccgccggcag cttccaggcg cggctgcggc tccgagtgct
ggtgcctccc ctgccctcac 720 tgaatcctgg tccagcacta gaagagggcc
agggcctgac cctggcagcc tcctgcacag 780 ctgagggcag cccagccccc
agcgtgacct gggacacgga ggtcaaaggc acaacgtcca 840 gccgttcctt
caagcactcc cgctctgctg ccgtcacctc agagttccac ttggtgccta 900
gccgcagcat gaatgggcag ccactgactt gtgtggtgtc ccatcctggc ctgctccagg
960 accaaaggat cacccacatc ctccacgtgt ccttccttgc tgaggcctct
gtgaggggcc 1020 ttgaagacca aaatctgtgg cacattggca gagaaggagc
tatgctcaag tgcctgagtg 1080 aagggcagcc ccctccctca tacaactgga
cacggctgga tgggcctctg cccagtgggg 1140 tacgagtgga tggggacact
ttgggctttc ccccactgac cactgagcac agcggcatct 1200 acgtctgcca
tgtcagcaat gagttctcct caagggattc tcaggtcact gtggatgttc 1260
ttgaccccca ggaagactct gggaagcagg tggacctagt gtcagcctcg gtggtggtgg
1320 tgggtgtgat cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg
ctcatgtccc 1380 gataccatcg gcgcaaggcc cagcagatga cccagaaata
tgaggaggag ctgaccctga 1440 ccagggagaa ctccatccgg aggctgcatt
cccatcacac ggaccccagg agccagccgg 1500 aggagagtgt agggctgaga
gccgagggcc accctgatag tctcaaggac aacagtagct 1560 gctctgtgat
gagtgaagag cccgagggcc gcagttactc cacgctgacc acggtgaggg 1620
agatagaaac acagactgaa ctgctgtctc caggctctgg gcgggccgag gaggaggaag
1680 atcaggatga aggcatcaaa caggccatga accattttgt tcaggagaat
gggaccctac 1740 gggccaagcc cacgggcaat ggcatctaca tcaatgggcg
gggacacctg gtctgaccca 1800 ggcctgcctc ccttccctag gcctggctcc
ttctgttgac atgggagatt ttagctcatc 1860 ttgggggcct ccttaaacac
ccccatttct tgcggaagat gctccccatc ccactgactg 1920 cttgaccttt
acctccaacc cttctgttca tcgggagggc tccaccaatt gagtctctcc 1980
caccatgcat gcaggtcact gtgtgtgtgc atgtgtgcct gtgtgagtgt tgactgactg
2040 tgtgtgtgtg gaggggtgac tgtccgtgga ggggtgactg tgtccgtggt
gtgtattatg 2100 ctgtcatatc agagtcaagt gaactgtggt gtatgtgcca
cgggatttga gtggttgcgt 2160 gggcaacact gtcagggttt ggcgtgtgtg
tcatgtggct gtgtgtgacc tctgcctgaa 2220 aaagcaggta ttttctcaga
ccccagagca gtattaatga tgcagaggtt ggaggagaga 2280 ggtggagact
gtggctcaga cccaggtgtg cgggcatagc tggagctgga atctgcctcc 2340
ggtgtgaggg aacctgtctc ctaccacttc ggagccatgg gggcaagtgt gaagcagcca
2400 gtccctgggt cagccagagg cttgaactgt tacagaagcc ctctgccctc
tggtggcctc 2460 tgggcctgct gcatgtacat attttctgta aatatacatg
cgccgggagc ttcttgcagg 2520 aatactgctc cgaatcactt ttaatttttt
tctttttttt ttcttgccct ttccattagt 2580 tgtatttttt atttattttt
atttttattt ttttttagag atggagtctc actatgttgc 2640 tcaggctggc
cttgaactcc tgggctcaag caatcctcct gcctcagcct ccctagtagc 2700
tgggacttta agtgtacacc actgtgcctg ctttgaatcc tttacgaaga gaaaaaaaaa
2760 attaaagaaa gcctttagat ttatccaatg tttactactg ggattgctta
aagtgaggcc 2820 cctccaacac cagggggtta attcctgtga ttgtgaaagg
ggctacttcc aaggcatctt 2880 catgcaggca gccccttggg agggcacctg
agagctggta gagtctgaaa ttagggatgt 2940 gagcctcgtg gttactgagt
aaggtaaaat tgcatccacc attgtttgtg ataccttagg 3000 gaattgcttg
gacctggtga caagggctcc tgttcaatag tggtgttggg gagagagaga 3060
gcagtgatta tagaccgaga gagtaggagt tgaggtgagg tgaaggaggt gctgggggtg
3120 agaatgtcgc ctttccccct gggttttgga tcactaattc aaggctcttc
tggatgtttc 3180 tctgggttgg ggctggagtt caatgaggtt tatttttagc
tggcccaccc agatacactc 3240 agccagaata cctagattta gtacccaaac
tcttcttagt ctgaaatctg ctggatttct 3300 ggcctaaggg agaggctccc
atccttcgtt ccccagccag cctaggactt cgaatgtgga 3360 gcctgaagat
ctaagatcct aacatgtaca ttttatgtaa atatgtgcat atttgtacat 3420
aaaatgatat tctgttttta aataaacaga caaaacttga aaaa 3464 113 3389 DNA
Homo sapiens 113 ggccgtcgtt gttggccaca gcgtgggaag cagctctggg
ggagctcgga gctcccgatc 60 acggcttctt gggggtagct acggctgggt
gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg gagacccaag
tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180 cagttcctta
ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac 240
gctgggcagt ctgcctttca accatgcccc tgtccctggg agccgagatg tgggggcctg
300 aggcctggct gctgctgctg ctactgctgg catcatttac aggccggtgc
cccgcgggtg 360 agctggagac ctcagacgtg gtaactgtgg tgctgggcca
ggacgcaaaa ctgccctgct 420 tctaccgagg ggactccggc gagcaagtgg
ggcaagtggc atgggctcgg gtggacgcgg 480 gcgaaggcgc ccaggaacta
gcgctactgc actccaaata cgggcttcat gtgagcccgg 540 cttacgaggg
ccgcgtggag cagccgccgc ccccacgcaa ccccctggac ggctcagtgc 600
tcctgcgcaa cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc agcaccttcc
660 ccgccggcag cttccaggcg cggctgcggc tccgagtgct ggtgcctccc
ctgccctcac 720 tgaatcctgg tccagcacta gaagagggcc agggcctgac
cctggcagcc tcctgcacag 780 ctgagggcag cccagccccc agcgtgacct
gggacacgga ggtcaaaggc acaacgtcca 840 gccgttcctt caagcactcc
cgctctgctg ccgtcacctc agagttccac ttggtgccta 900 gccgcagcat
gaatgggcag ccactgactt gtgtggtgtc ccatcctggc ctgctccagg 960
accaaaggat cacccacatc ctccacgtgt ccttccttgc tgaggcctct gtgaggggcc
1020 ttgaagacca aaatctgtgg cacattggca gagaaggagc tatgctcaag
tgcctgagtg 1080 aagggcagcc ccctccctca tacaactgga cacggctgga
tgggcctctg cccagtgggg 1140 tacgagtgga tggggacact ttgggctttc
ccccactgac cactgagcac agcggcatct 1200 acgtctgcca tgtcagcaat
gagttctcct caagggattc tcaggtcact gtggatgttc 1260 ttgaccccca
ggaagactct gggaagcagg tggacctagt gtcagcctcg gtggtggtgg 1320
tgggtgtgat cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg ctcatgtccc
1380 gataccatcg gcgcaaggcc cagcagatga cccagaaata tgaggaggag
ctgaccctga 1440 ccagggagaa ctccatccgg aggctgcatt cccatcacac
ggaccccagg agccagagtg 1500 aagagcccga gggccgcagt tactccacgc
tgaccacggt gagggagata gaaacacaga 1560 ctgaactgct gtctccaggc
tctgggcggg ccgaggagga ggaagatcag gatgaaggca 1620 tcaaacaggc
catgaaccat tttgttcagg agaatgggac cctacgggcc aagcccacgg 1680
gcaatggcat ctacatcaat gggcggggac acctggtctg acccaggcct gcctcccttc
1740 cctaggcctg gctccttctg ttgacatggg agattttagc tcatcttggg
ggcctcctta 1800 aacaccccca tttcttgcgg aagatgctcc ccatcccact
gactgcttga cctttacctc 1860 caacccttct gttcatcggg agggctccac
caattgagtc tctcccacca tgcatgcagg 1920 tcactgtgtg tgtgcatgtg
tgcctgtgtg agtgttgact gactgtgtgt gtgtggaggg 1980 gtgactgtcc
gtggaggggt gactgtgtcc gtggtgtgta ttatgctgtc atatcagagt 2040
caagtgaact gtggtgtatg tgccacggga tttgagtggt tgcgtgggca acactgtcag
2100 ggtttggcgt gtgtgtcatg tggctgtgtg tgacctctgc ctgaaaaagc
aggtattttc 2160 tcagacccca gagcagtatt aatgatgcag aggttggagg
agagaggtgg agactgtggc 2220 tcagacccag gtgtgcgggc atagctggag
ctggaatctg cctccggtgt gagggaacct 2280 gtctcctacc acttcggagc
catgggggca agtgtgaagc agccagtccc tgggtcagcc 2340 agaggcttga
actgttacag aagccctctg ccctctggtg gcctctgggc ctgctgcatg 2400
tacatatttt ctgtaaatat acatgcgccg ggagcttctt gcaggaatac tgctccgaat
2460 cacttttaat ttttttcttt tttttttctt gccctttcca ttagttgtat
tttttattta 2520 tttttatttt tatttttttt tagagatgga gtctcactat
gttgctcagg ctggccttga 2580 actcctgggc tcaagcaatc ctcctgcctc
agcctcccta gtagctggga ctttaagtgt 2640 acaccactgt gcctgctttg
aatcctttac gaagagaaaa aaaaaattaa agaaagcctt 2700 tagatttatc
caatgtttac tactgggatt gcttaaagtg aggcccctcc aacaccaggg 2760
ggttaattcc tgtgattgtg aaaggggcta cttccaaggc atcttcatgc aggcagcccc
2820 ttgggagggc acctgagagc tggtagagtc tgaaattagg gatgtgagcc
tcgtggttac 2880 tgagtaaggt aaaattgcat ccaccattgt ttgtgatacc
ttagggaatt gcttggacct 2940 ggtgacaagg gctcctgttc aatagtggtg
ttggggagag agagagcagt gattatagac 3000 cgagagagta ggagttgagg
tgaggtgaag gaggtgctgg gggtgagaat gtcgcctttc 3060 cccctgggtt
ttggatcact aattcaaggc tcttctggat gtttctctgg gttggggctg 3120
gagttcaatg aggtttattt ttagctggcc cacccagata cactcagcca gaatacctag
3180 atttagtacc caaactcttc ttagtctgaa atctgctgga tttctggcct
aagggagagg 3240 ctcccatcct tcgttcccca gccagcctag gacttcgaat
gtggagcctg aagatctaag 3300 atcctaacat gtacatttta tgtaaatatg
tgcatatttg tacataaaat gatattctgt 3360 ttttaaataa acagacaaaa
cttgaaaaa 3389 114 485 PRT Homo sapiens 114 Met Pro Leu Ser Leu Gly
Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu
Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu
Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45
Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50
55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu
Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala
Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro
Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp
Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly
Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro
Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly
Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175
Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180
185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu
Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu
Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg
Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu Ala
Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly
Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro
Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro
Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300
Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305
310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp
Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala
Ser Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys
Leu Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg
Arg Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu
Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser
His His Thr Asp Pro Arg Ser Gln Ser Glu Glu Pro Glu 405 410 415 Gly
Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr Gln 420 425
430 Thr Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu Glu Asp
435 440 445 Gln Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val Gln
Glu Asn 450 455 460 Gly Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile
Tyr Ile Asn Gly 465 470 475 480 Arg Gly His Leu Val 485 115 510 PRT
Homo sapiens 115 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro
Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr
Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val
Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr
Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala
Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu
His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95
Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100
105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys
Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu
Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro
Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala
Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr
Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe
Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu
Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220
Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225
230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu
Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu
Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp
Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp
Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser
Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser
Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu
Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340 345
350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val
355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met
Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn
Ser Ile Arg Arg 385 390 395 400 Leu His Ser His His Thr Asp Pro Arg
Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His
Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser
Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val
Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser
Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470
475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys
Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu
Val 500 505 510 116 485 PRT Homo sapiens 116 Met Pro Leu Ser Leu
Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu
Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu
Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40
45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln
50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu
Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro
Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn
Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala
Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala
Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro
Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu
Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170
175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser
180 185 190 Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser
Glu Phe 195 200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro
Leu Thr Cys Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln
Arg Ile Thr His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu
Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile
Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln
Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu
Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295
300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu
305 310 315 320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu
Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser
Ala Ser Val Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe
Cys Leu Leu Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His
Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu
Glu
Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His
Ser His His Thr Asp Pro Arg Ser Gln Ser Glu Glu Pro Glu 405 410 415
Gly Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg Glu Ile Glu Thr Gln 420
425 430 Thr Glu Leu Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu Glu
Asp 435 440 445 Gln Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val
Gln Glu Asn 450 455 460 Gly Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly
Ile Tyr Ile Asn Gly 465 470 475 480 Arg Gly His Leu Val 485 117
3401 DNA Homo sapiens 117 ggccgtcgtt gttggccaca gcgtgggaag
cagctctggg ggagctcgga gctcccgatc 60 acggcttctt gggggtagct
acggctgggt gtgtagaacg gggccggggc tggggctggg 120 tcccctagtg
gagacccaag tgcgagaggc aagaactctg cagcttcctg ccttctgggt 180
cagttcctta ttcaagtctg cagccggctc ccagggagat ctcggtggaa cttcagaaac
240 gctgggcagt ctgcctttca accatgcccc tgtccctggg agccgagatg
tgggggcctg 300 aggcctggct gctgctgctg ctactgctgg catcatttac
aggccggtgc cccgcgggtg 360 agctggagac ctcagacgtg gtaactgtgg
tgctgggcca ggacgcaaaa ctgccctgct 420 tctaccgagg ggactccggc
gagcaagtgg ggcaagtggc atgggctcgg gtggacgcgg 480 gcgaaggcgc
ccaggaacta gcgctactgc actccaaata cgggcttcat gtgagcccgg 540
cttacgaggg ccgcgtggag cagccgccgc ccccacgcaa ccccctggac ggctcagtgc
600 tcctgcgcaa cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc
agcaccttcc 660 ccgccggcag cttccaggcg cggctgcggc tccgagtgct
ggtgcctccc ctgccctcac 720 tgaatcctgg tccagcacta gaagagggcc
agggcctgac cctggcagcc tcctgcacag 780 ctgagggcag cccagccccc
agcgtgacct gggacacgga ggtcaaaggc acaacgtcca 840 gccgttcctt
caagcactcc cgctctgctg ccgtcacctc agagttccac ttggtgccta 900
gccgcagcat gaatgggcag ccactgactt gtgtggtgtc ccatcctggc ctgctccagg
960 accaaaggat cacccacatc ctccacgtgt ccttccttgc tgaggcctct
gtgaggggcc 1020 ttgaagacca aaatctgtgg cacattggca gagaaggagc
tatgctcaag tgcctgagtg 1080 aagggcagcc ccctccctca tacaactgga
cacggctgga tgggcctctg cccagtgggg 1140 tacgagtgga tggggacact
ttgggctttc ccccactgac cactgagcac agcggcatct 1200 acgtctgcca
tgtcagcaat gagttctcct caagggattc tcaggtcact gtggatgttc 1260
ttgaccccca ggaagactct gggaagcagg tggacctagt gtcagcctcg gtggtggtgg
1320 tgggtgtgat cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg
ctcatgtccc 1380 gataccatcg gcgcaaggcc cagcagatga cccagaaata
tgaggaggag ctgaccctga 1440 ccagggagaa ctccatccgg aggctgcatt
cccatcacac ggaccccagg agccagccgg 1500 aggagagtgt agggctgaga
gccgagggcc accctgatag tctcaaggac aacagtagct 1560 gctctgtgat
gagtgaagag cccgagggcc gcagttactc cacgctgacc acggtgaggg 1620
agatagaaac acagactgaa ctgctgtctc caggctctgg gcgggccgag gaggaggaag
1680 atcaggatga aggcatcaaa caggccatga accattttgt tcaggagaat
gggaccctac 1740 gggccaagcc cacgggcaat ggcatctaca tcaatgggcg
gggacacctg gtctgaccca 1800 ggcctgcctc ccttccctag gcctggctcc
ttctgttgac atgggagatt ttagctcatc 1860 ttgggggcct ccttaaacac
ccccatttct tgcggaagat gctccccatc ccactgactg 1920 cttgaccttt
acctccaacc cttctgttca tcgggagggc tccaccaatt gagtctctcc 1980
caccatgcat gcaggtcact gtgtgtgtgc atgtgtgcct gtgtgagtgt tgactgactg
2040 tgtgtgtgtg gaggggtgac tgtccgtgga ggggtgactg tgtccgtggt
gtgtattatg 2100 ctgtcatatc agagtcaagt gaactgtggt gtatgtgcca
cgggatttga gtggttgcgt 2160 gggcaacact gtcagggttt ggcgtgtgtg
tcatgtggct gtgtgtgacc tctgcctgaa 2220 aaagcaggta ttttctcaga
ccccagagca gtattaatga tgcagaggtt ggaggagaga 2280 ggtggagact
gtggctcaga cccaggtgtg cgggcatagc tggagctgga atctgcctcc 2340
ggtgtgaggg aacctgtctc ctaccacttc ggagccatgg gggcaagtgt gaagcagcca
2400 gtccctgggt cagccagagg cttgaactgt tacagaagcc ctctgccctc
tggtggcctc 2460 tgggcctgct gcatgtacat attttctgta aatatacatg
cgccgggagc ttcttgcagg 2520 aatactgctc cgaatcactt ttaatttttt
tctttttttt ttcttgccct ttccattagt 2580 tgtatttttt atttattttt
atttttattt ttttttagag atggagtctc actatgttgc 2640 tcaggctggc
cttgaactcc tgggctcaag caatcctcct gcctcagcct ccctagtagc 2700
tgggacttta agtgtacacc actgtgcctg ctttgaatcc tttacgaaga gaaaaaaaaa
2760 attaaagaaa gcctttagat ttatccaatg tttactactg ggattgctta
aagtgaggcc 2820 cctccaacac cagggggtta attcctgtga ttgtgaaagg
ggctacttcc aaggcatctt 2880 catgcaggca gccccttggg agggcacctg
agagctggta gagtctgaaa ttagggatgt 2940 gagcctcgtg ctggtgacaa
gggctcctgt tcaatagtgg tgttggggag agagagagca 3000 gtgattatag
accgagagag taggagttga ggtgaggtga aggaggtgct gggggtgaga 3060
atgtcgcctt tccccctggg ttttggatca ctaattcaag gctcttctgg atgtttctct
3120 gggttggggc tggagttcaa tgaggtttat ttttagctgg cccacccaga
tacactcagc 3180 cagaatacct agatttagta cccaaactct tcttagtctg
aaatctgctg gatttctggc 3240 ctaagggaga ggctcccatc cttcgttccc
cagccagcct aggacttcga atgtggagcc 3300 tgaagatcta agatcctaac
atgtacattt tatgtaaata tgtgcatatt tgtacataaa 3360 atgatattct
gtttttaaat aaacagacaa aacttgaaaa a 3401 118 3464 DNA Homo sapiens
118 ggccgtcgtt gttggccaca gcgtgggaag cagctctggg ggagctcgga
gctcccgatc 60 acggcttctt gggggtagct acggctgggt gtgtagaacg
gggccggggc tggggctggg 120 tcccctagtg gagacccaag tgcgagaggc
aagaactctg cagcttcctg ccttctgggt 180 cagttcctta ttcaagtctg
cagccggctc ccagggagat ctcggtggaa cttcagaaac 240 gctgggcagt
ctgcctttca accatgcccc tgtccctggg agccgagatg tgggggcctg 300
aggcctggct gctgctgctg ctactgctgg catcatttac aggccggtgc cccgcgggtg
360 agctggagac ctcagacgtg gtaactgtgg tgctgggcca ggacgcaaaa
ctgccctgct 420 tctaccgagg ggactccggc gagcaagtgg ggcaagtggc
atgggctcgg gtggacgcgg 480 gcgaaggcgc ccaggaacta gcgctactgc
actccaaata cgggcttcat gtgagcccgg 540 cttacgaggg ccgcgtggag
cagccgccgc ccccacgcaa ccccctggac ggctcagtgc 600 tcctgcgcaa
cgcagtgcag gcggatgagg gcgagtacga gtgccgggtc agcaccttcc 660
ccgccggcag cttccaggcg cggctgcggc tccgagtgct ggtgcctccc ctgccctcac
720 tgaatcctgg tccagcacta gaagagggcc agggcctgac cctggcagcc
tcctgcacag 780 ctgagggcag cccagccccc agcgtgacct gggacacgga
ggtcaaaggc acaacgtcca 840 gccgttcctt caagcactcc cgctctgctg
ccgtcacctc agagttccac ttggtgccta 900 gccgcagcat gaatgggcag
ccactgactt gtgtggtgtc ccatcctggc ctgctccagg 960 accaaaggat
cacccacatc ctccacgtgt ccttccttgc tgaggcctct gtgaggggcc 1020
ttgaagacca aaatctgtgg cacattggca gagaaggagc tatgctcaag tgcctgagtg
1080 aagggcagcc ccctccctca tacaactgga cacggctgga tgggcctctg
cccagtgggg 1140 tacgagtgga tggggacact ttgggctttc ccccactgac
cactgagcac agcggcatct 1200 acgtctgcca tgtcagcaat gagttctcct
caagggattc tcaggtcact gtggatgttc 1260 ttgaccccca ggaagactct
gggaagcagg tggacctagt gtcagcctcg gtggtggtgg 1320 tgggtgtgat
cgccgcactc ttgttctgcc ttctggtggt ggtggtggtg ctcatgtccc 1380
gataccatcg gcgcaaggcc cagcagatga cccagaaata tgaggaggag ctgaccctga
1440 ccagggagaa ctccatccgg aggctgcatt cccatcacac ggaccccagg
agccagccgg 1500 aggagagtgt agggctgaga gccgagggcc accctgatag
tctcaaggac aacagtagct 1560 gctctgtgat gagtgaagag cccgagggcc
gcagttactc cacgctgacc acggtgaggg 1620 agatagaaac acagactgaa
ctgctgtctc caggctctgg gcgggccgag gaggaggaag 1680 atcaggatga
aggcatcaaa caggccatga accattttgt tcaggagaat gggaccctac 1740
gggccaagcc cacgggcaat ggcatctaca tcaatgggcg gggacacctg gtctgaccca
1800 ggcctgcctc ccttccctag gcctggctcc ttctgttgac atgggagatt
ttagctcatc 1860 ttgggggcct ccttaaacac ccccatttct tgcggaagat
gctccccatc ccactgactg 1920 cttgaccttt acctccaacc cttctgttca
tcgggagggc tccaccaatt gagtctctcc 1980 caccatgcat gcaggtcact
gtgtgtgtgc atgtgtgcct gtgtgagtgt tgactgactg 2040 tgtgtgtgtg
gaggggtgac tgtccgtgga ggggtgactg tgtccgtggt gtgtattatg 2100
ctgtcatatc agagtcaagt gaactgtggt gtatgtgcca cgggatttga gtggttgcgt
2160 gggcaacact gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc
tctgcctgaa 2220 aaagcaggta ttttctcaga ccccagagca gtattaatga
tgcagaggtt ggaggagaga 2280 ggtggagact gtggctcaga cccaggtgtg
cgggcatagc tggagctgga atctgcctcc 2340 ggtgtgaggg aacctgtctc
ctaccacttc ggagccatgg gggcaagtgt gaagcagcca 2400 gtccctgggt
cagccagagg cttgaactgt tacagaagcc ctctgccctc tggtggcctc 2460
tgggcctgct gcatgtacat attttctgta aatatacatg cgccgggagc ttcttgcagg
2520 aatactgctc cgaatcactt ttaatttttt tctttttttt ttcttgccct
ttccattagt 2580 tgtatttttt atttattttt atttttattt ttttttagag
atggagtctc actatgttgc 2640 tcaggctggc cttgaactcc tgggctcaag
caatcctcct gcctcagcct ccctagtagc 2700 tgggacttta agtgtacacc
actgtgcctg ctttgaatcc tttacgaaga gaaaaaaaaa 2760 attaaagaaa
gcctttagat ttatccaatg tttactactg ggattgctta aagtgaggcc 2820
cctccaacac cagggggtta attcctgtga ttgtgaaagg ggctacttcc aaggcatctt
2880 catgcaggca gccccttggg agggcacctg agagctggta gagtctgaaa
ttagggatgt 2940 gagcctcgtg gttactgagt aaggtaaaat tgcatccacc
attgtttgtg ataccttagg 3000 gaattgcttg gacctggtga caagggctcc
tgttcaatag tggtgttggg gagagagaga 3060 gcagtgatta tagaccgaga
gagtaggagt tgaggtgagg tgaaggaggt gctgggggtg 3120 agaatgtcgc
ctttccccct gggttttgga tcactaattc aaggctcttc tggatgtttc 3180
tctgggttgg ggctggagtt caatgaggtt tatttttagc tggcccaccc agatacactc
3240 agccagaata cctagattta gtacccaaac tcttcttagt ctgaaatctg
ctggatttct 3300 ggcctaaggg agaggctccc atccttcgtt ccccagccag
cctaggactt cgaatgtgga 3360 gcctgaagat ctaagatcct aacatgtaca
ttttatgtaa atatgtgcat atttgtacat 3420 aaaatgatat tctgttttta
aataaacaga caaaacttga aaaa 3464 119 3401 DNA Homo sapiens 119
ggccgtcgtt gttggccaca gcgtgggaag cagctctggg ggagctcgga gctcccgatc
60 acggcttctt gggggtagct acggctgggt gtgtagaacg gggccggggc
tggggctggg 120 tcccctagtg gagacccaag tgcgagaggc aagaactctg
cagcttcctg ccttctgggt 180 cagttcctta ttcaagtctg cagccggctc
ccagggagat ctcggtggaa cttcagaaac 240 gctgggcagt ctgcctttca
accatgcccc tgtccctggg agccgagatg tgggggcctg 300 aggcctggct
gctgctgctg ctactgctgg catcatttac aggccggtgc cccgcgggtg 360
agctggagac ctcagacgtg gtaactgtgg tgctgggcca ggacgcaaaa ctgccctgct
420 tctaccgagg ggactccggc gagcaagtgg ggcaagtggc atgggctcgg
gtggacgcgg 480 gcgaaggcgc ccaggaacta gcgctactgc actccaaata
cgggcttcat gtgagcccgg 540 cttacgaggg ccgcgtggag cagccgccgc
ccccacgcaa ccccctggac ggctcagtgc 600 tcctgcgcaa cgcagtgcag
gcggatgagg gcgagtacga gtgccgggtc agcaccttcc 660 ccgccggcag
cttccaggcg cggctgcggc tccgagtgct ggtgcctccc ctgccctcac 720
tgaatcctgg tccagcacta gaagagggcc agggcctgac cctggcagcc tcctgcacag
780 ctgagggcag cccagccccc agcgtgacct gggacacgga ggtcaaaggc
acaacgtcca 840 gccgttcctt caagcactcc cgctctgctg ccgtcacctc
agagttccac ttggtgccta 900 gccgcagcat gaatgggcag ccactgactt
gtgtggtgtc ccatcctggc ctgctccagg 960 accaaaggat cacccacatc
ctccacgtgt ccttccttgc tgaggcctct gtgaggggcc 1020 ttgaagacca
aaatctgtgg cacattggca gagaaggagc tatgctcaag tgcctgagtg 1080
aagggcagcc ccctccctca tacaactgga cacggctgga tgggcctctg cccagtgggg
1140 tacgagtgga tggggacact ttgggctttc ccccactgac cactgagcac
agcggcatct 1200 acgtctgcca tgtcagcaat gagttctcct caagggattc
tcaggtcact gtggatgttc 1260 ttgaccccca ggaagactct gggaagcagg
tggacctagt gtcagcctcg gtggtggtgg 1320 tgggtgtgat cgccgcactc
ttgttctgcc ttctggtggt ggtggtggtg ctcatgtccc 1380 gataccatcg
gcgcaaggcc cagcagatga cccagaaata tgaggaggag ctgaccctga 1440
ccagggagaa ctccatccgg aggctgcatt cccatcacac ggaccccagg agccagccgg
1500 aggagagtgt agggctgaga gccgagggcc accctgatag tctcaaggac
aacagtagct 1560 gctctgtgat gagtgaagag cccgagggcc gcagttactc
cacgctgacc acggtgaggg 1620 agatagaaac acagactgaa ctgctgtctc
caggctctgg gcgggccgag gaggaggaag 1680 atcaggatga aggcatcaaa
caggccatga accattttgt tcaggagaat gggaccctac 1740 gggccaagcc
cacgggcaat ggcatctaca tcaatgggcg gggacacctg gtctgaccca 1800
ggcctgcctc ccttccctag gcctggctcc ttctgttgac atgggagatt ttagctcatc
1860 ttgggggcct ccttaaacac ccccatttct tgcggaagat gctccccatc
ccactgactg 1920 cttgaccttt acctccaacc cttctgttca tcgggagggc
tccaccaatt gagtctctcc 1980 caccatgcat gcaggtcact gtgtgtgtgc
atgtgtgcct gtgtgagtgt tgactgactg 2040 tgtgtgtgtg gaggggtgac
tgtccgtgga ggggtgactg tgtccgtggt gtgtattatg 2100 ctgtcatatc
agagtcaagt gaactgtggt gtatgtgcca cgggatttga gtggttgcgt 2160
gggcaacact gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc tctgcctgaa
2220 aaagcaggta ttttctcaga ccccagagca gtattaatga tgcagaggtt
ggaggagaga 2280 ggtggagact gtggctcaga cccaggtgtg cgggcatagc
tggagctgga atctgcctcc 2340 ggtgtgaggg aacctgtctc ctaccacttc
ggagccatgg gggcaagtgt gaagcagcca 2400 gtccctgggt cagccagagg
cttgaactgt tacagaagcc ctctgccctc tggtggcctc 2460 tgggcctgct
gcatgtacat attttctgta aatatacatg cgccgggagc ttcttgcagg 2520
aatactgctc cgaatcactt ttaatttttt tctttttttt ttcttgccct ttccattagt
2580 tgtatttttt atttattttt atttttattt ttttttagag atggagtctc
actatgttgc 2640 tcaggctggc cttgaactcc tgggctcaag caatcctcct
gcctcagcct ccctagtagc 2700 tgggacttta agtgtacacc actgtgcctg
ctttgaatcc tttacgaaga gaaaaaaaaa 2760 attaaagaaa gcctttagat
ttatccaatg tttactactg ggattgctta aagtgaggcc 2820 cctccaacac
cagggggtta attcctgtga ttgtgaaagg ggctacttcc aaggcatctt 2880
catgcaggca gccccttggg agggcacctg agagctggta gagtctgaaa ttagggatgt
2940 gagcctcgtg ctggtgacaa gggctcctgt tcaatagtgg tgttggggag
agagagagca 3000 gtgattatag accgagagag taggagttga ggtgaggtga
aggaggtgct gggggtgaga 3060 atgtcgcctt tccccctggg ttttggatca
ctaattcaag gctcttctgg atgtttctct 3120 gggttggggc tggagttcaa
tgaggtttat ttttagctgg cccacccaga tacactcagc 3180 cagaatacct
agatttagta cccaaactct tcttagtctg aaatctgctg gatttctggc 3240
ctaagggaga ggctcccatc cttcgttccc cagccagcct aggacttcga atgtggagcc
3300 tgaagatcta agatcctaac atgtacattt tatgtaaata tgtgcatatt
tgtacataaa 3360 atgatattct gtttttaaat aaacagacaa aacttgaaaa a 3401
120 510 PRT Homo sapiens 120 Met Pro Leu Ser Leu Gly Ala Glu Met
Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala
Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser
Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro
Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val
Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70
75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu
Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp
Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly
Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe
Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu Pro
Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly
Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala
Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185 190
Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195
200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys
Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr
His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser Val
Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg Glu
Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro
Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly
Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr
Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310 315
320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln
325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val
Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu
Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg Lys
Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu
Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His His
Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg
Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys
Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440
445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly
450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile
Lys Gln 465 470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly Thr
Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly
Arg Gly His Leu Val 500 505 510 121 510 PRT Homo sapiens 121 Met
Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu 1 5 10
15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro Ala Gly
20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu Gly Gln
Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser Gly Glu
Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala Gly Glu
Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr Gly Leu
His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln Pro Pro
Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu Arg Asn
Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125 Val Ser
Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130
135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu
Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr
Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu
Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg
Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg
Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro
Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235 240 His
Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250
255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser
260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp
Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu
Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val
Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln
Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys
Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val Gly Val
Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360 365 Val
Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375
380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg
385 390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln Pro Glu
Glu Ser Val 405 410 415 Gly Leu Arg Ala Glu Gly His Pro Asp Ser Leu
Lys Asp Asn Ser Ser 420 425 430 Cys Ser Val Met Ser Glu Glu Pro Glu
Gly Arg Ser Tyr Ser Thr Leu 435 440 445 Thr Thr Val Arg Glu Ile Glu
Thr Gln Thr Glu Leu Leu Ser Pro Gly 450 455 460 Ser Gly Arg Ala Glu
Glu Glu Glu Asp Gln Asp Glu Gly Ile Lys Gln 465 470 475 480 Ala Met
Asn His Phe Val Gln Glu Asn Gly Thr Leu Arg Ala Lys Pro 485 490 495
Thr Gly Asn Gly Ile Tyr Ile Asn Gly Arg Gly His Leu Val 500 505 510
122 510 PRT Homo sapiens 122 Met Pro Leu Ser Leu Gly Ala Glu Met
Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala
Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser
Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro
Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val
Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70
75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu
Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp
Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly
Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe
Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu Pro
Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly
Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala
Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185 190
Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195
200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys
Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr
His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser Val
Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg Glu
Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro
Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly
Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr
Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310 315
320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln
325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val
Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu
Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg Lys
Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu
Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His His
Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg
Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys
Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440
445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly
450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile
Lys Gln 465 470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly Thr
Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly
Arg Gly His Leu Val 500 505 510 123 1669 DNA Homo sapiens 123
gtctgaccca ggcctgcctc ccttccctag gcctggctcc ttctgttgac atgggagatt
60 ttagctcatc ttgggggcct ccttaaacac ccccatttct tgcggaagat
gctccccatc 120 ccactgactg cttgaccttt acctccaacc cttctgttca
tcgggagggc tccaccaatt 180 gagtctctcc caccatgcat gcaggtcact
gtgtgtgtgc atgtgtgcct gtgtgagtgt 240 tgactgactg tgtgtgtgtg
gaggggtgac tgtccgtgga ggggtgactg tgtccgtggt 300 gtgtattatg
ctgtcatatc agagtcaagt gaactgtggt gtatgtgcca cgggatttga 360
gtggttgcgt gggcaacact gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc
420 tctgcctgaa aaagcaggta ttttctcaga ccccagagca gtattaatga
tgcagaggtt 480 ggaggagaga ggtggagact gtggctcaga cccaggtgtg
cgggcatagc tggagctgga 540 atctgcctcc ggtgtgaggg aacctgtctc
ctaccacttc ggagccatgg gggcaagtgt 600 gaagcagcca gtccctgggt
cagccagagg cttgaactgt tacagaagcc ctctgccctc 660 tggtggcctc
tgggcctgct gcatgtacat attttctgta aatatacatg cgccgggagc 720
ttcttgcagg aatactgctc cgaatcactt ttaatttttt tctttttttt ttcttgccct
780 ttccattagt tgtatttttt atttattttt atttttattt ttttttagag
atggagtctc 840 actatgttgc tcaggctggc cttgaactcc tgggctcaag
caatcctcct gcctcagcct 900 ccctagtagc tgggacttta agtgtacacc
actgtgcctg ctttgaatcc tttacgaaga 960 gaaaaaaaaa attaaagaaa
gcctttagat ttatccaatg tttactactg ggattgctta 1020 aagtgaggcc
cctccaacac cagggggtta attcctgtga ttgtgaaagg ggctacttcc 1080
aaggcatctt catgcaggca gccccttggg agggcacctg agagctggta gagtctgaaa
1140 ttagggatgt gagcctcgtg gttactgagt aaggtaaaat tgcatccacc
attgtttgtg 1200 ataccttagg gaattgcttg gacctggtga caagggctcc
tgttcaatag tggtgttggg 1260 gagagagaga gcagtgatta tagaccgaga
gagtaggagt tgaggtgagg tgaaggaggt 1320 gctgggggtg agaatgtcgc
ctttccccct gggttttgga tcactaattc aaggctcttc 1380 tggatgtttc
tctgggttgg ggctggagtt caatgaggtt tatttttagc tggcccaccc 1440
agatacactc agccagaata cctagattta gtacccaaac tcttcttagt ctgaaatctg
1500 ctggatttct ggcctaaggg agaggctccc atccttcgtt ccccagccag
cctaggactt 1560 cgaatgtgga gcctgaagat ctaagatcct aacatgtaca
ttttatgtaa atatgtgcat 1620 atttgtacat aaaatgatat tctgttttta
aataaacaga caaaacttg 1669 124 1669 DNA Homo sapiens 124 gtctgaccca
ggcctgcctc ccttccctag gcctggctcc ttctgttgac atgggagatt 60
ttagctcatc ttgggggcct ccttaaacac ccccatttct tgcggaagat gctccccatc
120 ccactgactg cttgaccttt acctccaacc cttctgttca tcgggagggc
tccaccaatt 180 gagtctctcc caccatgcat gcaggtcact gtgtgtgtgc
atgtgtgcct gtgtgagtgt 240 tgactgactg tgtgtgtgtg gaggggtgac
tgtccgtgga ggggtgactg tgtccgtggt 300 gtgtattatg ctgtcatatc
agagtcaagt gaactgtggt gtatgtgcca cgggatttga 360 gtggttgcgt
gggcaacact gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc 420
tctgcctgaa aaagcaggta ttttctcaga ccccagagca gtattaatga tgcagaggtt
480 ggaggagaga ggtggagact gtggctcaga cccaggtgtg cgggcatagc
tggagctgga 540 atctgcctcc ggtgtgaggg aacctgtctc ctaccacttc
ggagccatgg gggcaagtgt 600 gaagcagcca gtccctgggt cagccagagg
cttgaactgt tacagaagcc ctctgccctc 660 tggtggcctc tgggcctgct
gcatgtacat attttctgta aatatacatg cgccgggagc 720 ttcttgcagg
aatactgctc cgaatcactt ttaatttttt tctttttttt ttcttgccct 780
ttccattagt tgtatttttt atttattttt atttttattt ttttttagag atggagtctc
840 actatgttgc tcaggctggc cttgaactcc tgggctcaag caatcctcct
gcctcagcct 900 ccctagtagc tgggacttta agtgtacacc actgtgcctg
ctttgaatcc tttacgaaga 960 gaaaaaaaaa attaaagaaa gcctttagat
ttatccaatg tttactactg ggattgctta 1020 aagtgaggcc cctccaacac
cagggggtta attcctgtga ttgtgaaagg ggctacttcc 1080 aaggcatctt
catgcaggca gccccttggg agggcacctg agagctggta gagtctgaaa 1140
ttagggatgt gagcctcgtg gttactgagt aaggtaaaat tgcatccacc attgtttgtg
1200 ataccttagg gaattgcttg gacctggtga caagggctcc tgttcaatag
tggtgttggg 1260 gagagagaga gcagtgatta tagaccgaga gagtaggagt
tgaggtgagg tgaaggaggt 1320 gctgggggtg agaatgtcgc ctttccccct
gggttttgga tcactaattc aaggctcttc 1380 tggatgtttc tctgggttgg
ggctggagtt caatgaggtt tatttttagc tggcccaccc 1440 agatacactc
agccagaata cctagattta gtacccaaac tcttcttagt ctgaaatctg 1500
ctggatttct ggcctaaggg agaggctccc atccttcgtt ccccagccag cctaggactt
1560 cgaatgtgga gcctgaagat ctaagatcct aacatgtaca ttttatgtaa
atatgtgcat 1620 atttgtacat aaaatgatat tctgttttta aataaacaga
caaaacttg 1669 125 1669 DNA Homo sapiens 125 gtctgaccca ggcctgcctc
ccttccctag gcctggctcc ttctgttgac atgggagatt 60 ttagctcatc
ttgggggcct ccttaaacac ccccatttct tgcggaagat gctccccatc 120
ccactgactg cttgaccttt acctccaacc cttctgttca tcgggagggc tccaccaatt
180 gagtctctcc caccatgcat gcaggtcact gtgtgtgtgc atgtgtgcct
gtgtgagtgt 240 tgactgactg tgtgtgtgtg gaggggtgac tgtccgtgga
ggggtgactg tgtccgtggt 300 gtgtattatg ctgtcatatc agagtcaagt
gaactgtggt gtatgtgcca cgggatttga 360 gtggttgcgt gggcaacact
gtcagggttt ggcgtgtgtg tcatgtggct gtgtgtgacc 420 tctgcctgaa
aaagcaggta ttttctcaga ccccagagca gtattaatga tgcagaggtt 480
ggaggagaga ggtggagact gtggctcaga cccaggtgtg cgggcatagc tggagctgga
540 atctgcctcc ggtgtgaggg aacctgtctc ctaccacttc ggagccatgg
gggcaagtgt 600 gaagcagcca gtccctgggt cagccagagg cttgaactgt
tacagaagcc ctctgccctc 660 tggtggcctc tgggcctgct gcatgtacat
attttctgta aatatacatg cgccgggagc 720 ttcttgcagg aatactgctc
cgaatcactt ttaatttttt tctttttttt ttcttgccct 780 ttccattagt
tgtatttttt atttattttt atttttattt ttttttagag atggagtctc 840
actatgttgc tcaggctggc cttgaactcc tgggctcaag caatcctcct gcctcagcct
900 ccctagtagc tgggacttta agtgtacacc actgtgcctg ctttgaatcc
tttacgaaga 960 gaaaaaaaaa attaaagaaa gcctttagat ttatccaatg
tttactactg ggattgctta 1020 aagtgaggcc cctccaacac cagggggtta
attcctgtga ttgtgaaagg ggctacttcc 1080 aaggcatctt catgcaggca
gccccttggg agggcacctg agagctggta gagtctgaaa 1140 ttagggatgt
gagcctcgtg gttactgagt aaggtaaaat tgcatccacc attgtttgtg 1200
ataccttagg gaattgcttg gacctggtga caagggctcc tgttcaatag tggtgttggg
1260 gagagagaga gcagtgatta tagaccgaga gagtaggagt tgaggtgagg
tgaaggaggt 1320 gctgggggtg agaatgtcgc ctttccccct gggttttgga
tcactaattc aaggctcttc 1380 tggatgtttc tctgggttgg ggctggagtt
caatgaggtt tatttttagc tggcccaccc 1440 agatacactc agccagaata
cctagattta gtacccaaac tcttcttagt ctgaaatctg 1500 ctggatttct
ggcctaaggg agaggctccc atccttcgtt ccccagccag cctaggactt 1560
cgaatgtgga gcctgaagat ctaagatcct aacatgtaca ttttatgtaa atatgtgcat
1620 atttgtacat aaaatgatat tctgttttta aataaacaga caaaacttg 1669 126
137 PRT Homo sapiens 126 Met Arg Arg Glu Leu Leu Ala Gly Ile Leu
Leu Arg Ile Thr Phe Asn 1 5 10 15 Phe Phe Leu Phe Phe Phe Leu Pro
Phe Pro Leu Val Val Phe Phe Ile 20 25 30 Tyr Phe Tyr Phe Tyr Phe
Phe Leu Glu Met Glu Ser His Tyr Val Ala 35 40 45 Gln Ala Gly Leu
Glu Leu Leu Gly Ser Ser Asn Pro Pro Ala Ser Ala 50 55 60 Ser Leu
Val Ala Gly Thr Leu Ser Val His His Cys Ala Cys Phe Glu 65 70 75 80
Ser Phe Thr Lys Arg Lys Lys Lys Leu Lys Lys Ala Phe Arg Phe Ile 85
90 95 Gln Cys Leu Leu Leu Gly Leu Leu Lys Val Arg Pro Leu Gln His
Gln 100 105 110 Gly Val Asn Ser Cys Asp Cys Glu Arg Gly Tyr Phe Gln
Gly Ile Phe 115 120 125 Met Gln Ala Ala Pro Trp Glu Gly Thr 130 135
127 510 PRT Homo sapiens 127 Met Pro Leu Ser Leu Gly Ala Glu Met
Trp Gly Pro Glu Ala Trp Leu 1 5 10 15 Leu Leu Leu Leu Leu Leu Ala
Ser Phe Thr Gly Arg Cys Pro Ala Gly 20 25 30 Glu Leu Glu Thr Ser
Asp Val Val Thr Val Val Leu Gly Gln Asp Ala 35 40 45 Lys Leu Pro
Cys Phe Tyr Arg Gly Asp Ser Gly Glu Gln Val Gly Gln 50 55 60 Val
Ala Trp Ala Arg Val Asp Ala Gly Glu Gly Ala Gln Glu Leu Ala 65 70
75 80 Leu Leu His Ser Lys Tyr Gly Leu His Val Ser Pro Ala Tyr Glu
Gly 85 90 95 Arg Val Glu Gln Pro Pro Pro Pro Arg Asn Pro Leu Asp
Gly Ser Val 100 105 110 Leu Leu Arg Asn Ala Val Gln Ala Asp Glu Gly
Glu Tyr Glu Cys Arg 115 120 125 Val Ser Thr Phe Pro Ala Gly Ser Phe
Gln Ala Arg Leu Arg Leu Arg 130 135 140 Val Leu Val Pro Pro Leu Pro
Ser Leu Asn Pro Gly Pro Ala Leu Glu 145 150 155 160 Glu Gly Gln Gly
Leu Thr Leu Ala Ala Ser Cys Thr Ala Glu Gly Ser 165 170 175 Pro Ala
Pro Ser Val Thr Trp Asp Thr Glu Val Lys Gly Thr Thr Ser 180 185 190
Ser Arg Ser Phe Lys His Ser Arg Ser Ala Ala Val Thr Ser Glu Phe 195
200 205 His Leu Val Pro Ser Arg Ser Met Asn Gly Gln Pro Leu Thr Cys
Val 210 215 220 Val Ser His Pro Gly Leu Leu Gln Asp Gln Arg Ile Thr
His Ile Leu 225 230 235 240 His Val Ser Phe Leu Ala Glu Ala Ser Val
Arg Gly Leu Glu Asp Gln 245 250 255 Asn Leu Trp His Ile Gly Arg Glu
Gly Ala Met Leu Lys Cys Leu Ser 260 265 270 Glu Gly Gln Pro Pro Pro
Ser Tyr Asn Trp Thr Arg Leu Asp Gly Pro 275 280 285 Leu Pro Ser Gly
Val Arg Val Asp Gly Asp Thr Leu Gly Phe Pro Pro 290 295 300 Leu Thr
Thr Glu His Ser Gly Ile Tyr Val Cys His Val Ser Asn Glu 305 310 315
320 Phe Ser Ser Arg Asp Ser Gln Val Thr Val Asp Val Leu Asp Pro Gln
325 330 335 Glu Asp Ser Gly Lys Gln Val Asp Leu Val Ser Ala Ser Val
Val Val 340 345 350 Val Gly Val Ile Ala Ala Leu Leu Phe Cys Leu Leu
Val Val Val Val 355 360 365 Val Leu Met Ser Arg Tyr His Arg Arg Lys
Ala Gln Gln Met Thr Gln 370 375 380 Lys Tyr Glu Glu Glu Leu Thr Leu
Thr Arg Glu Asn Ser Ile Arg Arg 385 390 395 400 Leu His Ser His His
Thr Asp Pro Arg Ser Gln Pro Glu Glu Ser Val 405 410 415 Gly Leu Arg
Ala Glu Gly His Pro Asp Ser Leu Lys Asp Asn Ser Ser 420 425 430 Cys
Ser Val Met Ser Glu Glu Pro Glu Gly Arg Ser Tyr Ser Thr Leu 435 440
445 Thr Thr Val Arg Glu Ile Glu Thr Gln Thr Glu Leu Leu Ser Pro Gly
450 455 460 Ser Gly Arg Ala Glu Glu Glu Glu Asp Gln Asp Glu Gly Ile
Lys Gln 465 470 475 480 Ala Met Asn His Phe Val Gln Glu Asn Gly Thr
Leu Arg Ala Lys Pro 485 490 495 Thr Gly Asn Gly Ile Tyr Ile Asn Gly
Arg Gly His Leu Val 500 505 510 128 295 PRT Homo sapiens 128 Met
Asn Gly Gln Pro Leu Thr Cys Val Val Ser His Pro Gly Leu Leu 1 5 10
15 Gln Asp Gln Arg Ile Thr His Ile Leu His Val Ser Phe Leu Ala Glu
20 25 30 Ala Ser Val Arg Gly Leu Glu Asp Gln Asn Leu Trp His Ile
Gly Arg 35 40 45 Glu Gly Ala Met Leu Lys Cys Leu Ser Glu Gly Gln
Pro Pro Pro Ser 50 55 60 Tyr Asn Trp Thr Arg Leu Asp Gly Pro Leu
Pro Ser
Gly Val Arg Val 65 70 75 80 Asp Gly Asp Thr Leu Gly Phe Pro Pro Leu
Thr Thr Glu His Ser Gly 85 90 95 Ile Tyr Val Cys His Val Ser Asn
Glu Phe Ser Ser Arg Asp Ser Gln 100 105 110 Val Thr Val Asp Val Leu
Asp Pro Gln Glu Asp Ser Gly Lys Gln Val 115 120 125 Asp Leu Val Ser
Ala Ser Val Val Val Val Gly Val Ile Ala Ala Leu 130 135 140 Leu Phe
Cys Leu Leu Val Val Val Val Val Leu Met Ser Arg Tyr His 145 150 155
160 Arg Arg Lys Ala Gln Gln Met Thr Gln Lys Tyr Glu Glu Glu Leu Thr
165 170 175 Leu Thr Arg Glu Asn Ser Ile Arg Arg Leu His Ser His His
Thr Asp 180 185 190 Pro Arg Ser Gln Pro Glu Glu Ser Val Gly Leu Arg
Ala Glu Gly His 195 200 205 Pro Asp Ser Leu Lys Asp Asn Ser Ser Cys
Ser Val Met Ser Glu Glu 210 215 220 Pro Glu Gly Arg Ser Tyr Ser Thr
Leu Thr Thr Val Arg Glu Ile Glu 225 230 235 240 Thr Gln Thr Glu Leu
Leu Ser Pro Gly Ser Gly Arg Ala Glu Glu Glu 245 250 255 Glu Asp Gln
Asp Glu Gly Ile Lys Gln Ala Met Asn His Phe Val Gln 260 265 270 Glu
Asn Gly Thr Leu Arg Ala Lys Pro Thr Gly Asn Gly Ile Tyr Ile 275 280
285 Asn Gly Arg Gly His Leu Val 290 295 129 485 PRT Homo sapiens
129 Met Pro Leu Ser Leu Gly Ala Glu Met Trp Gly Pro Glu Ala Trp Leu
1 5 10 15 Leu Leu Leu Leu Leu Leu Ala Ser Phe Thr Gly Arg Cys Pro
Ala Gly 20 25 30 Glu Leu Glu Thr Ser Asp Val Val Thr Val Val Leu
Gly Gln Asp Ala 35 40 45 Lys Leu Pro Cys Phe Tyr Arg Gly Asp Ser
Gly Glu Gln Val Gly Gln 50 55 60 Val Ala Trp Ala Arg Val Asp Ala
Gly Glu Gly Ala Gln Glu Leu Ala 65 70 75 80 Leu Leu His Ser Lys Tyr
Gly Leu His Val Ser Pro Ala Tyr Glu Gly 85 90 95 Arg Val Glu Gln
Pro Pro Pro Pro Arg Asn Pro Leu Asp Gly Ser Val 100 105 110 Leu Leu
Arg Asn Ala Val Gln Ala Asp Glu Gly Glu Tyr Glu Cys Arg 115 120 125
Val Ser Thr Phe Pro Ala Gly Ser Phe Gln Ala Arg Leu Arg Leu Arg 130
135 140 Val Leu Val Pro Pro Leu Pro Ser Leu Asn Pro Gly Pro Ala Leu
Glu 145 150 155 160 Glu Gly Gln Gly Leu Thr Leu Ala Ala Ser Cys Thr
Ala Glu Gly Ser 165 170 175 Pro Ala Pro Ser Val Thr Trp Asp Thr Glu
Val Lys Gly Thr Thr Ser 180 185 190 Ser Arg Ser Phe Lys His Ser Arg
Ser Ala Ala Val Thr Ser Glu Phe 195 200 205 His Leu Val Pro Ser Arg
Ser Met Asn Gly Gln Pro Leu Thr Cys Val 210 215 220 Val Ser His Pro
Gly Leu Leu Gln Asp Gln Arg Ile Thr His Ile Leu 225 230 235 240 His
Val Ser Phe Leu Ala Glu Ala Ser Val Arg Gly Leu Glu Asp Gln 245 250
255 Asn Leu Trp His Ile Gly Arg Glu Gly Ala Met Leu Lys Cys Leu Ser
260 265 270 Glu Gly Gln Pro Pro Pro Ser Tyr Asn Trp Thr Arg Leu Asp
Gly Pro 275 280 285 Leu Pro Ser Gly Val Arg Val Asp Gly Asp Thr Leu
Gly Phe Pro Pro 290 295 300 Leu Thr Thr Glu His Ser Gly Ile Tyr Val
Cys His Val Ser Asn Glu 305 310 315 320 Phe Ser Ser Arg Asp Ser Gln
Val Thr Val Asp Val Leu Asp Pro Gln 325 330 335 Glu Asp Ser Gly Lys
Gln Val Asp Leu Val Ser Ala Ser Val Val Val 340 345 350 Val Gly Val
Ile Ala Ala Leu Leu Phe Cys Leu Leu Val Val Val Val 355 360 365 Val
Leu Met Ser Arg Tyr His Arg Arg Lys Ala Gln Gln Met Thr Gln 370 375
380 Lys Tyr Glu Glu Glu Leu Thr Leu Thr Arg Glu Asn Ser Ile Arg Arg
385 390 395 400 Leu His Ser His His Thr Asp Pro Arg Ser Gln Ser Glu
Glu Pro Glu 405 410 415 Gly Arg Ser Tyr Ser Thr Leu Thr Thr Val Arg
Glu Ile Glu Thr Gln 420 425 430 Thr Glu Leu Leu Ser Pro Gly Ser Gly
Arg Ala Glu Glu Glu Glu Asp 435 440 445 Gln Asp Glu Gly Ile Lys Gln
Ala Met Asn His Phe Val Gln Glu Asn 450 455 460 Gly Thr Leu Arg Ala
Lys Pro Thr Gly Asn Gly Ile Tyr Ile Asn Gly 465 470 475 480 Arg Gly
His Leu Val 485 130 137 PRT Homo sapiens 130 Met Arg Arg Glu Leu
Leu Ala Gly Ile Leu Leu Arg Ile Thr Phe Asn 1 5 10 15 Phe Phe Leu
Phe Phe Phe Leu Pro Phe Pro Leu Val Val Phe Phe Ile 20 25 30 Tyr
Phe Tyr Phe Tyr Phe Phe Leu Glu Met Glu Ser His Tyr Val Ala 35 40
45 Gln Ala Gly Leu Glu Leu Leu Gly Ser Ser Asn Pro Pro Ala Ser Ala
50 55 60 Ser Leu Val Ala Gly Thr Leu Ser Val His His Cys Ala Cys
Phe Glu 65 70 75 80 Ser Phe Thr Lys Arg Lys Lys Lys Leu Lys Lys Ala
Phe Arg Phe Ile 85 90 95 Gln Cys Leu Leu Leu Gly Leu Leu Lys Val
Arg Pro Leu Gln His Gln 100 105 110 Gly Val Asn Ser Cys Asp Cys Glu
Arg Gly Tyr Phe Gln Gly Ile Phe 115 120 125 Met Gln Ala Ala Pro Trp
Glu Gly Thr 130 135
* * * * *
References