U.S. patent application number 14/365536 was filed with the patent office on 2015-10-29 for vaccines against hpv.
The applicant listed for this patent is Vaccibody AS. Invention is credited to Ali Areffard, Ole Henrik Brekke, Agnete Brunsvik Fredriksen, Mona Mari Lindeberg.
Application Number | 20150306217 14/365536 |
Document ID | / |
Family ID | 47471832 |
Filed Date | 2015-10-29 |
United States Patent
Application |
20150306217 |
Kind Code |
A9 |
Brekke; Ole Henrik ; et
al. |
October 29, 2015 |
VACCINES AGAINST HPV
Abstract
The present invention relates to therapeutic compounds, such as
vaccines against human papillomavirus (HPV) and in particular to
DNA vaccines against HPV16 or HPV18. The invention further relates
to protein construct encoding homodimeric peptides, which peptides
may be released from a DNA vaccine or used separately. Further
described are pharmaceutical formulations, host cells and methods
for producing the vaccines, as well as methods for the treatment of
various HPV induced diseases, such as cancers and infectious
diseases by application.
Inventors: |
Brekke; Ole Henrik; (Oslo,
NO) ; Fredriksen; Agnete Brunsvik; (Raelingen,
NO) ; Areffard; Ali; (Oslo, NO) ; Lindeberg;
Mona Mari; (Oslo, NO) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Vaccibody AS |
Oslo |
|
NO |
|
|
Prior
Publication: |
|
Document Identifier |
Publication Date |
|
US 20150056197 A1 |
February 26, 2015 |
|
|
Family ID: |
47471832 |
Appl. No.: |
14/365536 |
Filed: |
December 20, 2012 |
PCT Filed: |
December 20, 2012 |
PCT NO: |
PCT/EP2012/076404 PCKC 00 |
371 Date: |
June 13, 2014 |
Related U.S. Patent Documents
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|
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Application
Number |
Filing Date |
Patent Number |
|
|
61578542 |
Dec 21, 2011 |
|
|
|
Current U.S.
Class: |
424/134.1 ;
435/328; 435/69.3; 530/387.3; 536/23.4 |
Current CPC
Class: |
A61P 31/20 20180101;
C12N 2710/20022 20130101; A61K 2039/64 20130101; A61K 39/0011
20130101; C07K 2319/30 20130101; C12N 2710/20071 20130101; C07K
16/084 20130101; C07K 2319/00 20130101; C07K 2317/53 20130101; A61K
48/00 20130101; C07K 14/523 20130101; C12N 2710/20034 20130101;
A61K 2039/53 20130101; A61P 31/12 20180101; A61K 2039/505 20130101;
C12N 2800/22 20130101; A61K 39/39575 20130101; A61P 37/04 20180101;
A61K 39/12 20130101; C07K 2317/526 20130101; C07K 2319/02 20130101;
A61K 2039/585 20130101; A61K 2039/6056 20130101; A61P 35/00
20180101; C07K 14/005 20130101; C07K 2319/735 20130101; C12N 7/00
20130101; A61K 2039/892 20180801; A61K 2039/6031 20130101 |
International
Class: |
A61K 39/395 20060101
A61K039/395; C12N 7/00 20060101 C12N007/00; C07K 16/08 20060101
C07K016/08 |
Claims
1. A homodimeric protein of two identical amino acid chains, each
amino acid chain comprising (1) a signal peptide, (2) a targeting
unit, (3) a dimerization motif, and (4) an antigenic unit, said
targeting unit comprising an amino acid sequence having at least
80% sequence identity to the amino acid sequence 24-93 of SEQ ID
NO:1, and an antigenic unit comprising an amino acid sequence of
human papillomavirus (HPV), such as an antigenic unit comprising an
amino acid sequence of HPV 16 and/or HPV18, such as an antigenic
unit derived from early proteins E6 and/or E7 of HPV 16 and/or
HPV18.
2. (canceled)
3. The homodimeric protein according to claim 1, wherein said
signal peptide consists of an amino acid sequence having at least
80% sequence identity to the amino acid sequence 1-23 of SEQ ID
NO:l.
4. (canceled)
5. The homodimeric protein according to claim 1, wherein said
targeting unit consists of an amino acid sequence having at least
85%, such as at least 86%, such as at least 87%, such as at least
88%, such as at least 89%, such as at least 90%, such as at least
91%, such as at least 92%, such as at least 93%, such as at least
94%, such as at least 95%, such as at least 96%, such as at least
97%, such as at least 98%, such as at least 99% sequence identity
to the amino acid sequence 24-93 of SEQ ID NO:l.
6. The homodimeric protein according to claim 1, wherein the
dimerization motif comprises a hinge region and optionally another
domain that facilitate dimerization, such as an immunoglobulin
domain, optionally connected through a linker.
7. The homodimeric protein according to claim 6, wherein the hinge
region is Ig derived, such as derived from IgG3.
8-21. (canceled)
22. The homodimeric protein according to claim 1, wherein said
antigenic unit comprises an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to the amino acid
sequence 243-293 of SEQ ID NO:3.
23-39. (canceled)
40. The homodimeric protein according to claim 1, in its mature
form without any signal peptide sequence.
41. An amino acid chain comprising (1) a signal peptide, (2) a
targeting unit, (3) a dimerization motif, and (4) an antigenic
unit, said targeting unit comprising an amino acid sequence having
at least 80% sequence identity to the amino acid sequence 24-93 of
SEQ ID NO:1, and an antigenic unit comprising an amino acid
sequence of human papillomavirus (HPV), such as an antigenic unit
comprising an amino acid sequence of HPV 16 and/or HPV18, such as
an antigenic unit derived from early proteins E6 and/or E7 of HPV
16 and/or HPV18, which amino acid chain is able to form a
homodimeric protein according to claim 1.
42. A nucleic acid molecule, such as a DNA, encoding the amino acid
chain according to claim 41.
43. The nucleic acid molecule according to claim 42, which nucleic
acid molecule is human codon optimized.
44. A nucleic acid molecule comprising any one of nucleotide
sequences selected from the list consisting of SEQ ID NO:2, SEQ ID
NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID
NO:14, SEQ ID NO:16, SEQ ID NO:18, SEQ ID NO:20, SEQ ID NO:31 and
SEQ ID NO:33, or a variant thereof.
45. (canceled)
46. The nucleic acid molecule according to claim 42 formulated for
administration to a patient to induce production of the homodimeric
protein in said patients.
47. The homodimeric protein according to claim 1, for use as a
medicament,
48. A pharmaceutical composition comprising the homodimeric protein
according to claim 1.
49. A host cell comprising the nucleic acid molecule according to
claim 42.
50. A method for preparing a homodimeric protein, or an amino acid
chain, the method comprising a) transfecting the nucleic acid
molecule according to claim 42 into a cell population; b) culturing
the cell population; c) collecting and purifying the homodimeric
protein, or amino acid chain expressed from the cell
population.
51. A method for preparing a vaccine, such as a DNA vaccine,
comprising an immunologically effective amount of a nucleic acid
molecule, the method comprising a) preparing a nucleic acid
molecule according to claim 42; b) dissolving the nucleic acid
molecule obtained under step a) in a pharmaceutically acceptable
carrier, diluent, or buffer.
52. A vaccine against HPV comprising an immunologically effective
amount of a homodimeric protein according to claim 1, wherein said
vaccine is able to trigger both a T-cell- and B-cell immune
response.
53. (canceled)
54. A method of treating or preventing a HPV induced disease or
condition, such as a cancer or an infectious disease caused by HPV
in a patient, the method comprising administering to the patient in
need thereof, a homodimeric protein according to claim 1.
55. The method according to claim 54, wherein the method comprises
administering to the patient in need thereof of a nucleic acid
molecule, with a subsequent step of electroporation.
56. (canceled)
Description
FIELD OF THE INVENTION
[0001] The present invention relates to therapeutic compounds, such
as vaccines against human papillomavirus (HPV) and in particular to
DNA vaccines against HPV16 and/or HPV18. The invention further
relates to protein construct encoding homodimeric peptides, which
peptides may be released from a DNA vaccine or used separately.
Further described are pharmaceutical formulations, host cells and
methods for producing the vaccines, as well as methods for the
treatment of various HPV induced diseases, such as cancers and
infectious diseases by application.
BACKGROUND OF THE INVENTION
[0002] It is now well established that human papillomavirus (HPV)
is the cause of cervical cancer and other HPV-associated
malignancies such as anogenital (anus, vulvar, vaginal and penile)
cancers and a subset of head and neck cancers. In particular, HPV16
and HPV 18 are responsible for about 70% of all cervical cancers
worldwide.
[0003] To date, two prophylactic HPV vaccines are on the market
(Gardasil and Cervarix). The aim of the prophylactic vaccines is to
induce humoral immune responses by stimulating the production of
neutralizing antibodies specific for the HPV viral capsid proteins,
L1 and L2. Although the preventive vaccines are an important
milestone for the control of HPV induced cervical cancer and
possibly other HPV-associated malignancies, the effect of these
vaccines will not be significantly observed for 20-40 years (Ma B
et al., Current Cancer Therapy Reviews, 2010). Moreover, since the
coverage of mass vaccination for the prophylactic vaccines are to
date limited in addition to a substantial population worldwide that
already are HPV infected, HPV-associated malignancies will continue
to progress. Thus, it will be important to develop HPV-specific
therapeutic vaccines in order to reduce the mortality and morbidity
of HPV-associated malignancies and its precursor lesions (Ma B et
al., Current Cancer Therapy Reviews, 2010).
[0004] The development of various cancer vaccines and cancer
immunotherapy strategies has throughout the last two decades
expanded. Still, only one therapeutic cancer vaccine, called
Provenge (Dendreon INC) has so far been approved to be applied as
standard therapy for prostate cancer. Notably, due to ethical
reasons the majority of therapeutic cancer vaccines are tested on a
patient group bearing a late stage tumor. This patient group is
substantially immunosuppressed meaning that the tumor cells have
for long escaped the immune system and contributed to induce
immunological tolerance to the tumor along carcinogenesis. In
addition, the choice of antigens (tumor-specific vs.
tumor-associated) applied as vaccines are critical in order to
induce tumor-specific immune responses and avoid killing of healthy
cells in the patients which may lead to serious adverse events.
Thus, the major challenges in cancer immunotherapy are to break the
immunological tolerance and activate tumor-specific effector
functions to recognize and kill tumor cells. Although some case
reports show clinical response to therapeutic cancer vaccines in
late stage tumor patients, the most common primary endpoint is to
observe the impact on overall survival compared to conventional
therapy (surgery, chemo and radiation therapy). However, most
studies are either not conclusive or that they completely fail to
show this. One reason for the negative results lies in the patient
group carrying end-stage tumors that are challenging to treat in
the first place. A possible strategy could be to include patients
with early-stage tumors in therapeutic vaccine trials.
[0005] One strategy is to target pre-cancerous lesions. The
challenges for this strategy are mainly the lack of reliable
biomarkers that are specifically expressed by precancerous lesions
for many tissues and poor medical screening (either non-existing or
that the existing method suffers from lack of sensitivity).
Exceptionally, this is not the case for HPV-induced malignancies.
For instance, the majority of western countries have good screening
programs for cervical dysplasia and cervical cancer by performing
the papanicolaou test (Pap smear test). If there are unclear or
abnormal results from Pap smear test, colposcopy will be performed
(National Cervical Cancer Coalition). HPV-testing may also be
recommended for some patients to detect the presence of "high-risk"
HPV-type in the precancerous lesion. Thus, HPV represents a
potential biomarker for HPV-associated precancerous lesions, in
particular cervical intraepithelial dysplasia (CIN).
[0006] DNA vaccines have shown increasing promise for the treatment
of human diseases, in particular cancer. DNA vaccines induce strong
antigen-specific immune responses and can be repeatedly
administered to maintain the target-specific immune responses. Such
vaccines are considered to be safe and simple and cheap to produce
on a large scale compared to other cancer therapeutic formats.
Numerous immunotherapeutic interventions fail to induce
immunological memory. Exceptionally, DNA vaccination ensures
sustained release of the vaccine product in vivo which enhances
antigen-specific immunological memory. Direct delivery of antigens
to professional antigen-presenting cells (APCs) stimulates both
CD4+ and CD8+ T cell immune responses in vivo. Such strong cellular
immune responses have been demonstrated to specifically recognize
and kill antigen-positive malignant cells efficiently both in vitro
and in vivo.
[0007] There is still a need in the art for improved vaccines for
inducing strong and specific immune responses against HPV
responsible for both infectious diseases and cancers.
OBJECT OF THE INVENTION
[0008] It is an object of embodiments of the invention to provide
specific and highly effective therapeutic compounds, such as DNA
vaccines against diseases and conditions caused by HPV.
SUMMARY OF THE INVENTION
[0009] It has been found by the present inventors that by combining
the antigens of the early gene products E6 and E7 from HPV, such as
from HPV16 and/or HPV18 with the targeting module of hMIP-1.alpha.,
therapeutic vaccines are provided, wherein the strong immunogenic
epitopes of HPV gene products are presented with high efficiency to
APCs to induce a specific and strong immune response. The products
according to the present invention is primarily envisioned as
therapeutic nucleic acid vaccines, such as DNA vaccines, wherein a
nucleic acid construct encoding the vaccibody construct is used as
the therapeutic compound leading to in vivo production of the
protein product within the person receiving the vaccine. However,
as an alternative the protein product itself may be formulated and
used directly in the vaccine.
[0010] Accordingly, in a first aspect the present invention relates
to a homodimeric protein of two identical amino acid chains, each
amino acid chain comprising (1) a signal peptide, (2) a targeting
unit, (3) a dimerization motif, and (4) an antigenic unit, said
targeting unit comprising an amino acid sequence having at least
80% sequence identity to the amino acid sequence 24-93 of SEQ ID
NO:1, and an antigenic unit comprising an amino acid sequence of
human papillomavirus (HPV), such as an antigenic unit comprising an
amino acid sequence of HPV16 and/or HPV18, such as an antigenic
unit derived from early proteins E6 and/or E7 of HPV16 and/or
HPV18.
[0011] In a second aspect the present invention relates to an amino
acid chain comprising (1) a signal peptide, (2) a targeting unit,
(3) a dimerization motif, and (4) an antigenic unit, said targeting
unit comprising an amino acid sequence having at least 80% sequence
identity to the amino acid sequence 24-93 of SEQ ID NO:1, and an
antigenic unit comprising an amino acid sequence of human
papillomavirus (HPV), such as an antigenic unit comprising an amino
acid sequence of HPV16 and/or HPV18, such as an antigenic unit
derived from early proteins E6 and/or E7 of HPV16 and/or HPV18,
which amino acid chain is able to form a homodimeric protein
according to the invention.
[0012] In a third aspect the present invention relates to a nucleic
acid molecule, such as a DNA, encoding an amino acid chain
comprising (1) a signal peptide, (2) a targeting unit, (3) a
dimerization motif, and (4) an antigenic unit, said targeting unit
comprising an amino acid sequence having at least 80% sequence
identity to the amino acid sequence 24-93 of SEQ ID NO:1, and an
antigenic unit comprising an amino acid sequence of human
papillomavirus (HPV), such as an antigenic unit comprising an amino
acid sequence of HPV16 and/or HPV18, such as an antigenic unit
derived from early proteins E6 and/or E7 of HPV16 and/or HPV18,
which amino acid chain is able to form a homodimeric protein
according to the invention.
[0013] In a further aspect the present invention relates to a
homodimeric protein according to the invention, or an amino acid
chain according to the invention, or the nucleic acid molecule
according to the invention for use as a medicament.
[0014] In a further aspect the present invention relates to a
pharmaceutical composition comprising a homodimeric protein
according to the invention, or an amino acid chain according to the
invention, or the nucleic acid molecule according to the
invention.
[0015] In a further aspect the present invention relates to a host
cell comprising the nucleic acid molecule according to the
invention.
[0016] In a further aspect the present invention relates to a
method for preparing a homodimeric protein according to the
invention, or an amino acid chain of the invention, the method
comprising a) transfecting the nucleic acid molecule according to
the invention into a cell population; b) culturing the cell
population; c) collecting and purifying the homodimeric protein, or
amino acid chain expressed from the cell population.
[0017] In a further aspect the present invention relates to a
method for preparing a vaccine, such as a DNA vaccine, comprising
an immunologically effective amount of a nucleic acid molecule
according to the invention, the method comprising a) preparing a
nucleic acid molecule according to the invention; b) dissolving the
nucleic acid molecule obtained under step a) in a pharmaceutically
acceptable carrier, diluent, or buffer.
[0018] In a further aspect the present invention relates to a
vaccine against HPV comprising an immunologically effective amount
of a homodimeric protein according to the invention, or an amino
acid chain according to the invention, or nucleic acid molecule,
such as a DNA, according to the invention, wherein said vaccine is
able to trigger both a T-cell- and B-cell immune response.
[0019] In a further aspect the present invention relates to a
method of treating or preventing a HPV induced disease or
condition, such as a cancer or an infectious disease caused by HPV
in a patient, the method comprising administering to the patient in
need thereof, a homodimeric protein according to the invention, or
an amino acid chain according to the invention, or the nucleic acid
molecule, such as a DNA, according to the invention.
LEGENDS TO THE FIGURE
[0020] FIG. 1: The overall structure of vaccibody vaccines with
E7/E6 fusion antigen. Shown are both DNA and protein formats. The
vaccibody consist of three functional modules; the chemokine human
MIP-1.alpha. (LD78(3) in the targeting module, hinge and CH3
sequences from human IgG3 in the dimerization module and
full-length E7 and/or E6 fusion in the vaccine module.
[0021] FIG. 2: The suggested mode of action for a Vaccibody DNA
vaccine against HPV -induced malignancies. Naked DNA plasmid
encoding vaccibody is injected intradermal followed by
electroporation. The plasmid is taken up by local cells and
vaccibody proteins are produced and secreted. The chemotactic
targeting modules attract CCR1 and CCR5 expressing antigen
presenting cells (APC) and ensure binding and uptake into dendritic
cells (DC). The DC will present antigenic peptides to CD4+ and CD8+
T cells and the CD8+ T cells will kill HPV infected and transformed
cells in the cervix.
[0022] FIG. 3: ELISPOT results showing the number of E7 and E6
specific T cell responses as a function of different amounts of
vaccine administered. C57BL/6 mice were injected i.d. with naked
DNA plasmids encoding VB1009 and VB1016 and their corresponding
controls followed by electroporation (Cellectis, France) on day 0
and day 7. Splenocytes were harvested at day 21 and stimulated with
MHC class I-restricted E7 or E6 peptide for 24 h. The number of
IFNy secreting splenocytes was calculated by ELISPOT.
(A)E7-specific responses after i.d. vaccination with 25 .mu.g of
VB1009, control 1 (antigen alone) and pUMVC4a (empty vector).(B)
E7-specific responses after i.d. vaccination with 12.5 and 1.4
.mu.g of VB1016, control 2 (antigen alone) and pUMVC4a (empty
vector). (C) E6-specific responses after i.d. vaccination with 12.5
and 1.4 .mu.g of VB1016, control 2 (antigen alone) and pUMVC4a
(empty vector).
[0023] FIG. 4.Therapeutic effect of VB1016 shown by measured tumor
volume. C57BL/6 mice were injected s.c. with 5.times.10.sup.5 TC-1
cells at day 0. At day 3 and day 10, the mice were injected i.d.
with 12.5 .mu.g naked DNA plasmids encoding VB1016, control 2 or
empty vector followed by electroporation (Cellectis, France). The
tumor sizes were measured by caliper two to three times a week and
tumor volume calculated.
[0024] FIG. 5.Therapeutic effect of VB1016 shown by measured tumor
volume. C57BL/6 mice were injected s.c. in the neck area with
5.times.10.sup.4 TC-1 cells at day 0. At day 3,7 and day 10, the
mice were injected i.d. with 20 .mu.g or 2 .mu.g naked DNA plasmids
encoding VB1016, control 2 or empty vector followed by
electroporation (Cellectis, France). The tumor sizes were measured
by caliper two to three times a week and tumor volume
calculated.
[0025] FIG. 6.Therapeutic effect of VB1020 and VB1021 shown by
measured tumor volume. C57BL/6 mice were injected s.c. in the thigh
with 5.times.10.sup.4 TC-1 cells at day 0. At day 3 and day 10, the
mice were injected i.d. with 10 .mu.g naked DNA plasmids encoding
VB1016, VB1020, VB1021 or empty vector followed by electroporation
(Cellectis, France). The tumor sizes were measured by caliper two
to three times a week and tumor volume calculated.
DETAILED DISCLOSURE OF THE INVENTION
[0026] The constructs and DNA vaccine technology described herein
by the inventors of the present invention (also referred to as
"vaccibody" molecules/vaccines/constructs) represents a novel
vaccine strategy to induce strong and specific immune responses for
both infectious diseases and cancer. The HPV E6/E7, such as HPV16
or HPV18 E6/E7 vaccine described herein may be administered as a
DNA vaccine by intradermal injection, preferably followed by
electroporation. This results in the uptake of the DNA-construct
encoding the vaccibody-HPV16 and/or HPV18 E6/E7 vaccine in cells at
the site of injection (dermis) including dendritic cells
(Langerhans cells), leading to in vivo production of the
vaccibody-E6/E7 molecule.
[0027] The early gene products E6 and E7 from "high-risk" HPV types
such as HPV16 and 18 may be responsible for transformation of the
basal-epithelium cells and induction of precancerous lesions. Both
proteins consist of highly immunogenic epitopes and are shown
herein to induce strong immune responses leading to specific
eradication of "high-risk" HPV positive tumor cells both in vitro
and in vivo.
[0028] The vaccibody molecule described herein is a homodimer
consisting of three modules; targeting module, dimerization module
and the vaccine module (FIG. 1). Genes encoding the three modules
are genetically engineered to be expressed as one gene. When
expressed in vivo, the vaccibody molecule targets antigen
presenting cells (APCs) which results in an enhanced vaccine
potency compared to identical, non-targeted antigens. In vivo
expression of the chemokine human macrophage inflammatory protein 1
alpha (hMIP-1.alpha./LD78.beta.) leads to attraction of DCs,
neutrophils and other immune cells carrying the CCR1 and CCR5
receptors to the site of expression. Thus, the vaccibody molecule
consisting of hMIP-1.alpha. as the targeting module, will not only
target the antigens to specific cells, but in addition give a
response-amplifying effect (adjuvant effect) by recruiting specific
immune cells to the injection site. This unique mechanism may be of
great importance in a clinical setting where patients can receive
the vaccine without any additional adjuvants since the vaccine
itself gives the adjuvant effect.
[0029] The inventors of the present invention describes herein
vaccine constructs where the antigenic module consist of the E7
full length genetic sequence in fusion to the E6 full length
sequence originating from the HPV16 or HPV18 subtype. The advantage
of this format is that both E6 and E7 will be present in one
construct and may thus be equally expressed in vivo. Consequently,
one vaccibody molecule consisting of a multi-antigenic unit may
represent equal levels of E6 and E7 for the immune system. The
HPV16 E6 and E7 gene products are oncogenic in their natural form.
To neutralize their oncogenic properties, mutations at specific
sites may be introduced in the E6 and E7 genetic sequence.
[0030] The mutations, including deletions, may be introduced at
specific sites, known to inhibit the oncogenic properties of E6 and
E7, such as any one described in any of Dalal S et al., J Virol,
1996; Munger K et al., EMBO, 1989; Nakagawa S et al., Virology,
1995; Crook T et al., Cell, 1991; Munger K et al., HPV Compendium
Online, 1997
(http://www.stdgen.lanl.gov/COMPENDIUM_PDF/97PDF/3/E7.pdf); Nguyen,
M et al., J Virol, 2002; Nomine Yet a., Molecular Cell, 2006; Moody
C et al., Nat Rev Cancer, 2010, Polakova I et al., Vaccine, 2010;
Xie Q, Virologica Sinica, 2011; Mesplede T et al., J Virol, 2012;
US 2008/0102084 and U.S. Pat. No. 6,306,397, which references are
hereby incorporated by reference. Accordingly, in some aspects of
the invention, the constructs according to the present invention
contain HPV16 E6, E7 or HPV16 E6/E7 chimeric constructs with one or
more mutations in either of HPV16 E6, E7 or both at a position
known to inhibit the oncogenic properties as described in Dalal S
et al., 3 Virol, 1996; Munger K et al., EMBO, 1989; Nakagawa S et
al., Virology, 1995; Crook T et al., Cell, 1991; Munger K et al.,
HPV Compendium Online, 1997
(http://www.stdgen.lanl.gov/COMPENDIUM_PDF/97PDF/3/E7.pdf); Nguyen,
M et al., J Virol, 2002; Nomine Y et a., Molecular Cell, 2006;
Moody C et al., Nat Rev Cancer, 2010, Polakova I et al., Vaccine,
2010; Xie Q, Virologica Sinica, 2011; Mesplede T et al., J Virol,
2012; US 2008/0102084 or U.S. Pat. No. 6,306,397. In other aspects
of the invention, the constructs according to the present invention
contain HPV18 E6, E7 or HPV18 E6/E7 chimeric constructs with one or
more mutations in either of HPV18 E6, E7 or both at a position
known to inhibit the oncogenic properties as described in Dalal S
et al., J Virol, 1996; Munger K et al., EMBO, 1989; Nakagawa S et
al., Virology, 1995; Crook T et al., Cell, 1991; Munger K et al.,
HPV Compendium Online, 1997
(http://www.stdgen.lanl.gov/COMPENDIUM_PDF/97PDF/3/E7.pdf); Moody C
et al., Nat Rev Cancer, 2010, US 2008/0102084 and U.S. Pat. No.
6,306,397.
[0031] There is a possibility that the vaccibody-moiety (targeting
and dimerization modules) may eradicate the oncogenic properties of
E6 and E7 wildtype proteins in the final fusion protein. Thus, in
yet another aspect of the invention is the utilization of the
wildtype full-length E6 and/or E7 sequences in the vaccibody
construction.
[0032] The invention describes several variant of Vaccibody HPV
therapeutic DNA vaccines all based on the overall format described
in FIG. 1, the therapeutic vaccibody-HPV DNA vaccines encodes genes
that are naturally expressed in humans; the targeting module genes
encode the chemokine hMIP-1.alpha., which binds to its cognate
receptors, CCR1 and CCR5 expressed on the cell surface of APCs. The
dimerization module genes may encode hinge regions and constant
heavy chain 3, such as from human IgG3 which connects two vaccibody
monomers generating a homodimer molecule. Genes encoding the
vaccine module for the current strategy consist of HPV, such as
HPV16 and/or HPV18 E7 and E6 antigens, such as the full length
HPV16 E7 and E6 antigens, optionally comprising one or more
mutation to inhibit the oncogenic properties. Once administered in
vivo by i.d. injection followed by electroporation, dermal cells
taking up the vaccine construct will express the vaccibody-HPV
molecule. The in vivo produced vaccibody vaccines target to CCR1
and CCR5 expressed on the surface of APCs in the skin, in
particular DCs. The binding of the vaccibody molecule to its
cognate receptors leads to internalization of the complex in the
APC, degradation of the proteins into small peptides that are
loaded onto MHC molecules and presented to CD4.sup.+ and CD8.sup.+
T cells to induce HPV16 E6 and E7 specific immune responses. Once
stimulated and with help from activated CD4.sup.+ T cells,
CD8.sup.+ T cells will target and kill HPV16 E6 and E7 expressing
cells (FIG. 2). Such enhanced immune responses to a vaccine with a
"built-in" adjuvant effect may potentially overcome tumor-escape
(tumor immune surveillance) by breaking immunological tolerance and
efficiently kill malignant cells. The hMIP-1.alpha. targeting unit
may be connected through a dimerization motif, such as a hinge
region, to an antigenic unit, wherein the later is in either the
COOH-terminal or the NH2-terminal end. The present invention not
only relates to a DNA sequence coding for this recombinant protein,
but also to expression vectors comprising these DNA sequences, cell
lines comprising said expression vectors, to treatment of mammals
preferentially by immunization by means of Vaccibody DNA, Vaccibody
RNA, or Vaccibody protein, and finally to pharmaceuticals and a kit
comprising the said molecules.
[0033] The dimerization motif in the proteins according to the
present invention may be constructed to include a hinge region and
an immunoglobulin domain (e.g. Cy3 domain), e.g. carboxyterminal C
domain (C.sub.H3 domain), or a sequence that is substantially
identical to said C domain. The hinge region may be Ig derived and
contributes to the dimerization through the formation of an
interchain covalent bond(s), e.g. disulfide bridge(s). In addition,
it functions as a flexible spacer between the domains allowing the
two targeting units to bind simultaneously to two target molecules
on APC expressed with variable distances. The immunoglobulin
domains contribute to homodimerization through non-covalent
interactions, e.g. hydrophobic interactions. In a preferred
embodiment the C.sub.H3 domain is derived from IgG. These
dimerization motifs may be exchanged with other multimerization
moieties (e.g. from other Ig isotypes/subclasses). Preferably the
dimerization motif is derived from native human proteins, such as
human IgG.
[0034] It is to be understood that the dimerization motif may have
any orientation with respect to antigenic unit and targeting unit.
In one embodiment the antigenic unit is in the COOH-terminal end of
the dimerization motif with the targeting unit in the N-terminal
end of the dimerization motif. In another embodiment the antigenic
unit is in the N-terminal end of the dimerization motif with the
targeting unit in the COOH-terminal end of the dimerization
motif.
[0035] International application WO 2004/076489, which is hereby
incorporated by reference discloses nucleic acid sequences and
vectors, which may be used according to the present invention.
[0036] The proteins according to the present invention include an
antigenic unit derived from HPV, such as HPV16 E7 and E6 antigens,
such as the full length HPV16 E7 and E6 antigens, as well as
immunogenic fragments or variants thereof. The antigenic sequence
should be of sufficient length. The minimal length of such
antigenic unit may be around 9 amino acids. Accordingly in some
embodiments, the antigenic unit derived from HPV comprises an amino
acid sequence of at least 9 amino acids corresponding to at least
about 27 nucleotides in a nucleic acids sequence encoding such
antigenic unit. Preferably the antigenic unit derived from HPV is
considerably longer, such as the full length HPV16 E7 and E6
antigens. Diversity arises within a given HPV genotype through
limited nucleotide changes in the coding (at a frequency of <2%)
and non-coding (at a frequency of <5%) regions (Bernard, HU et
al., Int 3 Cancer, 2006). Such variants phylogenetically segregate
based on their geographical origin and are therefore labeled
European, African, Asian, Asian-American and North American.
Insertion of such sequences in a Vaccibody format might lead to
activation of both arms of the immune response.
[0037] Immunization by means of Vaccibody protein, Vaccibody DNA,
or Vaccibody RNA, the latter two executed e.g. by intramuscular or
intradermal injection with or without a following electroporation,
are all feasible methods according to the present invention.
[0038] As discussed above, the present invention relates to a
vaccine composition against cancer or infectious diseases caused by
HPV, the vaccine composition comprising an immunologically
effective amount of the nucleic acid encoding the molecule of the
invention or degenerate variants thereof. The vaccine may be able
to trigger both a T-cell- and B-cell immune response. The present
invention also relates to a kit comprising Vaccibody DNA, RNA, or
protein for diagnostic, medical or scientific purposes.
[0039] The invention further relates to a method of preparing the
recombinant molecule of the invention comprising, transfecting the
vector comprising the molecule of the invention into a cell
population; culturing the cell population; collecting recombinant
protein expressed from the cell population; and purifying the
expressed protein.
[0040] The above described nucleotide sequences may be inserted
into a vector suited for gene therapy, e.g. under the control of a
specific promoter, and introduced into the cells. In some
embodiments the vector comprising said DNA sequence is a virus,
e.g. an adenovirus, vaccinia virus or an adeno-associated virus. In
some embodiments a retroviruses is used as vector. Examples of
suitable retroviruses are e.g. MoMuLV or HaMuSV. For the purpose of
gene therapy, the DNA/RNA sequences according to the invention can
also be transported to the target cells in the form of colloidal
dispersions. They comprise e.g. liposomes or lipoplexes.
[0041] The present invention encompasses the use of a targeting
unit as well as an antigenic unit having minimum degree of sequence
identity or sequence homology with amino acid sequence(s) defined
herein or with a polypeptide having the specific properties defined
herein. The present invention encompasses, in particular, the use
of peptide variants or peptide units to be used in the constructs
according to the present invention having a degree of sequence
identity with any one of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ
ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:15, SEQ
ID NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23,
SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID
NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:32, or SEQ ID NO:34.
Here, the term "variant" means an entity having a certain degree of
sequence identity with the subject amino acid sequences or the
subject nucleotide sequences, where the subject amino acid sequence
preferably is SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4,
SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9,
SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID
NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ
ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23,
SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID
NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ
ID NO:33, or SEQ ID NO:34.
[0042] In one aspect, the variant or fragment amino acid sequence
and/or nucleotide sequence should provide and/or encode a
polypeptide which retains the functional activity and/or enhances
the activity of a polypeptide of SEQ ID NO:1, SEQ ID NO:3, SEQ ID
NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID
NO:15, SEQ ID NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:22, SEQ
ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27,
SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:32, or SEQ ID
NO:34.
[0043] In the present context, a variant sequence is taken to
include an amino acid sequence which may be at least 80%, at least
85%, at least 90%, at least 95%, at least 96%, at least 97%, at
least 98% or at least 99%, identical to the subject sequence.
Typically, the variants used according to the present invention
will comprise the same active sites etc. as the subject amino acid
sequence. Although homology can also be considered in terms of
similarity (i.e. amino acid residues having similar chemical
properties/functions), in the context of the present invention it
is preferred to express homology in terms of sequence identity.
[0044] Sequence identity comparisons can be conducted by eye, or
more usually, with the aid of readily available sequence comparison
computer programs. These commercially available computer programs
use complex comparison algorithms to align two or more sequences
that best reflect the evolutionary events that might have led to
the difference(s) between the two or more sequences. Therefore,
these algorithms operate with a scoring system rewarding alignment
of identical or similar amino acids and penalising the insertion of
gaps, gap extensions and alignment of non-similar amino acids. The
scoring system of the comparison algorithms include: [0045] i)
assignment of a penalty score each time a gap is inserted (gap
penalty score), [0046] ii) assignment of a penalty score each time
an existing gap is extended with an extra position (extension
penalty score), [0047] iii) assignment of high scores upon
alignment of identical amino acids, and [0048] iv) assignment of
variable scores upon alignment of non-identical amino acids.
[0049] Most alignment programs allow the gap penalties to be
modified. However, it is preferred to use the default values when
using such software for sequence comparisons.
[0050] The scores given for alignment of non-identical amino acids
are assigned according to a scoring matrix also called a
substitution matrix. The scores provided in such substitution
matrices are reflecting the fact that the likelihood of one amino
acid being substituted with another during evolution varies and
depends on the physical/chemical nature of the amino acid to be
substituted. For example, the likelihood of a polar amino acid
being substituted with another polar amino acid is higher compared
to being substituted with a hydrophobic amino acid. Therefore, the
scoring matrix will assign the highest score for identical amino
acids, lower score for non-identical but similar amino acids and
even lower score for non-identical non-similar amino acids. The
most frequently used scoring matrices are the PAM matrices (Dayhoff
et al. (1978), Jones et al. (1992)), the BLOSUM matrices (Henikoff
and Henikoff (1992)) and the Gonnet matrix (Gonnet et al.
(1992)).
[0051] Suitable computer programs for carrying out such an
alignment include, but are not limited to, Vector NTI (Invitrogen
Corp.) and the ClustalV, ClustalW and ClustalW2 programs (Higgins D
G & Sharp P M (1988), Higgins et al. (1992), Thompson et al.
(1994), Larkin et al. (2007). A selection of different alignment
tools is available from the ExPASy Proteomics server at
www.expasy.org. Another example of software that can perform
sequence alignment is BLAST (Basic Local Alignment Search Tool),
which is available from the webpage of National Center for
Biotechnology Information which can currently be found at
http://www.ncbi.nlm.nih.gov/and which was firstly described in
Altschul et al. (1990) J. Mol. Biol. 215; 403-410.
[0052] Once the software has produced an alignment, it is possible
to calculate % similarity and .degree. A) sequence identity. The
software typically does this as part of the sequence comparison and
generates a numerical result.
[0053] In one embodiment, it is preferred to use the ClustalW
software for performing sequence alignments. Preferably, alignment
with ClustalW is performed with the following parameters for
pairwise alignment:
TABLE-US-00001 Substitution matrix: Gonnet 250 Gap open penalty: 20
Gap extension penalty: 0.2 Gap end penalty: None
[0054] ClustalW2 is for example made available on the internet by
the European Bioinformatics Institute at the EMBL-EBI webpage
www.ebi.ac.uk under tools--sequence analysis--ClustalW2. Currently,
the exact address of the ClustalW2 tool is
www.ebi.ac.uk/Tools/clustalw2.
[0055] In another embodiment, it is preferred to use the program
Align X in Vector NTI (Invitrogen) for performing sequence
alignments. In one embodiment, Exp10 has been may be used with
default settings: [0056] Gap opening penalty: 10 [0057] Gap
extension penalty: 0.05 [0058] Gapseparation penalty range: 8
[0059] Score matrix: blosum62mt2
[0060] Thus, the present invention also encompasses the use of
variants, fragments, and derivatives of any amino acid sequence of
a protein, polypeptide, motif or domain as defined herein,
particularly those of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID
NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:15, SEQ ID
NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ
ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28,
SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:32, or SEQ ID NO:34.
[0061] The sequences, particularly those of variants, fragments,
and derivatives of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID
NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:15, SEQ ID
NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ
ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28,
SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:32, or SEQ ID NO:34, may also
have deletions, insertions or substitutions of amino acid residues
which produce a silent change and result in a functionally
equivalent substance. Deliberate amino acid substitutions may be
made on the basis of similarity in polarity, charge, solubility,
hydrophobicity, hydrophilicity, and/or the amphipathic nature of
the residues as long as the secondary binding activity of the
substance is retained. For example, negatively charged amino acids
include aspartic acid and glutamic acid; positively charged amino
acids include lysine and arginine; and amino acids with uncharged
polar head groups having similar hydrophilicity values include
leucine, isoleucine, valine, glycine, alanine, asparagine,
glutamine, serine, threonine, phenylalanine, and tyrosine.
[0062] The present invention also encompasses conservative
substitution (substitution and replacement are both used herein to
mean the interchange of an existing amino acid residue, with an
alternative residue) that may occur i.e. like-for-like substitution
such as basic for basic, acidic for acidic, polar for polar etc.
Non-conservative substitution may also occur i.e. from one class of
residue to another or alternatively involving the inclusion of
unnatural amino acids such as ornithine (hereinafter referred to as
Z), diaminobutyric acid ornithine (hereinafter referred to as B),
norleucine ornithine (hereinafter referred to as 0), pyriylalanine,
thienylalanine, naphthylalanine and phenylglycine.
[0063] Conservative substitutions that may be made are, for example
within the groups of basic amino acids (Arginine, Lysine and
Histidine), acidic amino acids (glutamic acid and aspartic acid),
aliphatic amino acids (Alanine, Valine, Leucine, Isoleucine), polar
amino acids (Glutamine, Asparagine, Serine, Threonine), aromatic
amino acids (Phenylalanine, Tryptophan and Tyrosine), hydroxyl
amino acids (Serine, Threonine), large amino acids (Phenylalanine
and Tryptophan) and small amino acids (Glycine, Alanine).
[0064] Replacements may also be made by unnatural amino acids
include; alpha* and alpha-disubstituted* amino acids, N-alkyl amino
acids*, lactic acid*, halide derivatives of natural amino acids
such as trifluorotyrosine*, p-Cl-phenylalanine*,
p-Br-phenylalanine*, p-I-phenylalanine*, L-allyl-glycine*,
0-alanine*, L-.alpha.-amino butyric acid*, L-.gamma.-amino butyric
acid*, L-.alpha.-amino isobutyric acid*, L-.epsilon.-amino caproic
acid*, 7-amino heptanoic acid*, L-methionine sulfone*,
L-norleucine*, L-norvaline*, p-nitro-L-phenylalanine*,
L-hydroxyproline*, L-thioproline*, methyl derivatives of
phenylalanine (Phe) such as 4-methyl-Phe*, pentannethyl-Phe*, L-Phe
(4-annino)*, L-Tyr (methyl)*, L-Phe (4-isopropyl)*, L-Tic
(1,2,3,4-tetrahydroisoquinoline-3-carboxyl acid)*,
L-diaminopropionic acid # and L-Phe (4-benzyl)*. The notation * has
been utilised for the purpose of the discussion above (relating to
homologous or non-conservative substitution), to indicate the
hydrophobic nature of the derivative whereas # has been utilised to
indicate the hydrophilic nature of the derivative, #* indicates
amphipathic characteristics.
[0065] Variant amino acid sequences may include suitable spacer
groups that may be inserted between any two amino acid residues of
the sequence including alkyl groups such as methyl, ethyl or propyl
groups in addition to amino acid spacers such as glycine or
.beta.-alanine residues. A further form of variation, involves the
presence of one or more amino acid residues in peptoid form, will
be well understood by those skilled in the art. For the avoidance
of doubt, "the peptoid form" is used to refer to variant amino acid
residues wherein the a-carbon substituent group is on the residue's
nitrogen atom rather than the a-carbon. Processes for preparing
peptides in the peptoid form are known in the art, for example
Simon R J et al. (1992), Horwell D C. (1995).
[0066] In one embodiment, the variant targeting unit used in the
homodimeric protein according to the present invention is variant
having the sequence of amino acids at least 80%, at least 85%, at
least 90%, at least 95%, at least 96%, at least 97%, at least 98%
or at least 99% amino acid sequence identity therewith.
[0067] In one aspect, preferably the protein or sequence used in
the present invention is in a purified form. The term "purified"
means that a given component is present at a high level. The
component is desirably the predominant active component present in
a composition.
[0068] A "variant" or "variants" refers to proteins, polypeptides,
units, motifs, domains or nucleic acids. The term "variant" may be
used interchangeably with the term "mutant." Variants include
insertions, substitutions, transversions, truncations, and/or
inversions at one or more locations in the amino acid or nucleotide
sequence, respectively. The phrases "variant polypeptide",
"polypeptide", "variant" and "variant enzyme" mean a
polypeptide/protein that has an amino acid sequence that has been
modified from the amino acid sequence of SEQ ID NO: 1. The variant
polypeptides include a polypeptide having a certain percent, e.g.,
80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, of
sequence identity with the amino acid sequence of SEQ ID NO:1, SEQ
ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ
ID NO:13, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:19, SEQ ID NO:21,
SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID
NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ
ID NO:32, or SEQ ID NO:34.
[0069] "Variant nucleic acids" can include sequences that are
complementary to sequences that are capable of hybridizing to the
nucleotide sequences presented herein. For example, a variant
sequence is complementary to sequences capable of hybridizing under
stringent conditions, e.g., 50.degree. C. and 0.2.times. SSC
(1.times. SSC=0.15 M NaCl, 0.015 M sodium citrate, pH 7.0), to the
nucleotide sequences presented herein. More particularly, the term
variant encompasses sequences that are complementary to sequences
that are capable of hybridizing under highly stringent conditions,
e.g., 65.degree. C. and 0.1.times. SSC, to the nucleotide sequences
presented herein. The melting point (Tm) of a variant nucleic acid
may be about 1, 2, or 3.degree. C. lower than the Tm of the
wild-type nucleic acid. The variant nucleic acids include a
polynucleotide having a certain percent, e.g., 80%, 85%, 90%, 95%,
or 99%, of sequence identity with the nucleic acid encoding SEQ ID
NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID
NO:11, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:19, SEQ
ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25,
SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID
NO:30, SEQ ID NO:32, or SEQ ID NO:34, encoding the monomeric
protein which can form the homodimeric protein according to
invention.
[0070] A specific category of mutations are the mutations in E6 and
E7:
[0071] The E6 protein may be detoxified by rendering the p53
binding impossible. Five positions in the full length HPV16 E6
protein are sites for mutations for inactivation of E6
functionality, F47, L50, C63, C106 and 1128. Any amino acid
substitution in these positions may lead to inactivation of E6 and
induces tumor suppression. Substitutions in any one of these
positions with any one different amino acid may potentially be
utilized. Sites for potential mutations are shown in SEQ ID
NO:22.
[0072] In the E7 protein there are conserved regions associated
with oncogenic properties (see Phelps et al 3. Virol. April 1992,
vol. 66, no. 42418-242; Gulliver et al 3 Virol. 1997, August;
71(8)) including an N-terminal Rb (retinoblastoma binding protein)
binding-site motif (LXCXE) and two conserved regions 3 (upstream
and downstream) with a Zn-binding motif (CXXC). The preferred
mutation sites in the LXCXE-motif are C24 and E26. Preferred sites
in the two CXXC motifs are C58, C61, C91 and C94. However, any
mutations in these regions can be envisaged to be substituted for
the reduction of binding functions and thus abolish the oncogenic
effects of E7. Sites for potential mutations are shown in SEQ ID
NO:23.
[0073] Signal Peptide:
[0074] A signal peptide at the N-terminal end of the nascent
polypeptide directs the molecule into the ER before transport to
into the Golgi complex. The signal peptide is cleaved off by signal
peptidase once it has served its purpose of targeting and importing
the protein to the ER. These signal peptides are generally between
15 and 30 amino acids, but can have more than 50 residues
(Martoglio, B. et al., Trends in Cell Biology, 1998, Knappskog, S.
et al., J Biotechnol, 2007). The native signal peptide may be
replaced by signal peptides from any mammalian, prokaryotic or
marine origin. Commonly used signal peptides are e.g. humanlL-2 and
human albumin due to their natural ability to secrete large amounts
of protein. The choice of signal peptide can have a considerable
impact on the amount of synthesized and secreted protein.
[0075] In some embodiments, the signal peptide used in the protein
construct according to the present invention is derived from a
chemokine protein, such as the signal sequence of LD78beta.
[0076] In some embodiments the signal peptide is not derived from
pLNOH2 (B1-8 variable immunoglobulin leader) disclosed in the
international application with International Application No:
PCT/EP2011/060628.
[0077] In some embodiments the signal peptide is not derived from
an immunoglobulin gene.
[0078] The term "honnodinneric protein" as used herein refers to a
protein comprising two individual identical strands of amino acids,
or subunits held together as a single, dimeric protein by hydrogen
bonding, ionic (charged) interactions, actual covalent disulfide
bonding, or some combination of these interactions.
[0079] The term "dimerization motif", as used herein, refers to the
sequence of amino acids between the antigenic unit and the
targeting unit comprising the hinge region and the optional second
domain that may contribute to the dimerization. This second domain
may be an immunoglobulin domain, and optionally the hinge region
and the second domain are connected through a linker. Accordingly
the dimerization motif serves to connect the antigenic unit and the
targeting unit, but also contain the hinge region that facilitates
the dimerization of the two monomeric proteins into a homodimeric
protein according to the invention.
[0080] The term "targeting unit" as used herein refers to a unit
that delivers the protein with its antigen to mouse or human APC
for MHC class II-restricted presentation to CD4+ T cells or for
providing cross presentation to CD8+ T cells by MHC class I
restriction. The targeting unit used in the constructs according to
the present invention is derived from or identical to mature
LD78-beta.
[0081] The term "antigenic unit" as used herein refers to any
molecule, such as a peptide which is able to be specifically
recognized by an antibody or other component of the immune system,
such as a surface receptor on T-cells. Included within this
definition are also immunogens that are able to induce an immune
response. The terms "epitope" or "antigenic epitope" is used to
refer to a distinct molecular surface, such as a molecular surface
provided by a short peptide sequence within an antigenic unit. In
some embodiments the antigenic unit comprises two ore more
antigenic epitopes. The antigenic unit used in the constructs
according to the present invention is derived from or identical to
the early gene products E6 and E7 from HPV, such as from HPV16 or
HPV18.
[0082] The term "hinge region" refers to a peptide sequence of the
homodimeric protein that facilitates the dimerization, such as
through the formation of an interchain covalent bond(s), e.g.
disulfide bridge(s). The hinge region may be Ig derived, such as
hinge exons h1+h4 of an Ig, such as IgG3.
[0083] Specific Embodiments of the Invention:
[0084] As described above, the present invention relates to a
homodimeric protein of two identical amino acid chains, each amino
acid chain comprising (1) a signal peptide, (2) a targeting unit,
(3) a dimerization motif, and (4) an antigenic unit, said targeting
unit comprising an amino acid sequence having at least 80% sequence
identity to the amino acid sequence 24-93 of SEQ ID NO:1, and an
antigenic unit comprising an amino acid sequence of human
papillomavirus (HPV), such as an antigenic unit comprising an amino
acid sequence of HPV16 and/or HPV18, such as an antigenic unit
derived from early proteins E6 and/or E7 of HPV16 and/or HPV18. In
some embodiments according to the present invention, the targeting
unit, dimerization motif and antigenic unit in the amino acid chain
are in the N-terminal to C-terminal order of targeting unit,
dimerization motif and antigenic unit.
[0085] In some embodiments, the antigenic unit used in the
constructs according to the present invention is derived from
HPV16, such as from early proteins E6 and/or E7.
[0086] In some embodiments, the antigenic unit used in the
constructs according to the present invention is derived from E6 of
HPV16.
[0087] In some embodiments, the antigenic unit used in the
constructs according to the present invention is derived from E7 of
HPV16.
[0088] In some embodiments, the antigenic unit used in the
constructs according to the present invention is derived from
HPV18, such as from early proteins E6 and/or E7.
[0089] In some embodiments, the antigenic unit used in the
constructs according to the present invention is derived from E6 of
HPV18.
[0090] In some embodiments, the antigenic unit used in the
constructs according to the present invention is derived from E7 of
HPV18.
[0091] In some embodiments according to the present invention, the
signal peptide consists of an amino acid sequence having at least
80% sequence identity to the amino acid sequence 1-23 of SEQ ID
NO:1.
[0092] In some embodiments according to the present invention, the
signal peptide consists of an amino acid sequence having at least
85%, such as at least 86%, such as at least 87%, such as at least
88%, such as at least 89%, such as at least 90%, such as at least
91%, such as at least 92%, such as at least 93%, such as at least
94%, such as at least 95%, such as at least 96%, such as at least
97%, such as at least 98%, such as at least 99%, such as 100%
sequence identity to the amino acid sequence 1-23 of SEQ ID
NO:1.
[0093] In some embodiments according to the present invention, the
targeting unit consists of an amino acid sequence having at least
85%, such as at least 86%, such as at least 87%, such as at least
88%, such as at least 89%, such as at least 90%, such as at least
91%, such as at least 92%, such as at least 93%, such as at least
94%, such as at least 95%, such as at least 96%, such as at least
97%, such as at least 98%, such as at least 99% sequence identity
to the amino acid sequence 24-93 of SEQ ID NO:1.
[0094] In some embodiments according to the present invention, the
dimerization motif comprises a hinge region and optionally another
domain that facilitate dimerization, such as an immunoglobulin
domain, optionally connected through a linker.
[0095] In some embodiments according to the present invention, the
hinge region is Ig derived, such as derived from IgG3.
[0096] In some embodiments according to the present invention, the
hinge region has the ability to form one, two, or several covalent
bonds. In some embodiments according to the present invention, the
covalent bond is a disulphide bridge.
[0097] In some embodiments according to the present invention, the
immunoglobulin domain of the dimerization motif is a
carboxyterminal C domain, or a sequence that is substantially
identical to the C domain or a variant thereof.
[0098] In some embodiments according to the present invention, the
carboxyterminal C domain is derived from IgG.
[0099] In some embodiments according to the present invention, the
immunoglobulin domain of the dimerization motif has the ability to
homodimerize.
[0100] In some embodiments according to the present invention, the
immunoglobulin domain has the ability to homodimerize via
noncovalent interactions. In some embodiments according to the
present invention, the noncovalent interactions are hydrophobic
interactions.
[0101] In some embodiments according to the present invention, the
dimerization domain does not comprise the CH2 domain.
[0102] In some embodiments according to the present invention, the
dimerization motif consists of hinge exons h1 and h4 connected
through a linker to a C.sub.H3 domain of human IgG3.
[0103] In some embodiments according to the present invention, the
dimerization motif consist of an amino acid sequence having at
least 80% sequence identity to the amino acid sequence 94-237 of
SEQ ID NO:3.
[0104] In some embodiments according to the present invention, the
linker is a G.sub.3S.sub.2G.sub.3SG linker.
[0105] In some embodiments according to the present invention, the
antigenic unit and the dimerization motif is connected through a
linker, such as a GLGGL linker or a GLSGL linker.
[0106] In some embodiments according to the present invention, the
targeting unit consists of amino acids 24-93 of SEQ ID NO:1, or a
variant thereof.
[0107] In some embodiments according to the present invention, the
homodimeric protein have increased affinity for any one chemokine
receptor selected from CCR1, CCR3 and CCR5 as compared to the
affinity of the same homodimeric protein with the targeting unit
consisting of amino acids 24-93 of SEQ ID NO:1, or a variant
thereof.
[0108] In some embodiments according to the present invention, the
antigenic unit comprises an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to the amino acid
sequence 243-293 of SEQ ID NO:3.
[0109] In some embodiments according to the present invention, the
antigenic unit consists of an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to the amino acid
sequence 243-293 of SEQ ID NO:3.
[0110] In some embodiments according to the present invention, the
antigenic unit comprises one or more amino acid substitutions at a
position selected from the list consisting of F47, L50, C63, C106
and 1128 of SEQ ID NO:22, or a deletion involving one or more amino
acid selected from the list consisting of Y43-L50 of SEQ ID
NO:22.
[0111] In some embodiments according to the present invention, the
antigenic unit comprises not more than 1, 2, 3, 4, 5, 6, 7, 8, 9,
10, 12, 14, 16, 18 or 20 amino acid substitutions and/or deletions
relative to SEQ ID NO:22.
[0112] In some embodiments according to the present invention, the
antigenic unit comprises the amino acid sequence 243-293 of SEQ ID
NO:3, SEQ ID NO:5, SEQ ID NO:7, or SEQ ID NO:9, or a variant or
antigenic fragment thereof.
[0113] In some embodiments according to the present invention, the
antigenic unit consists of the amino acid sequence 243-293 of SEQ
ID NO:3, SEQ ID NO:5, SEQ ID NO:7, or SEQ ID NO:9, or a variant or
antigenic fragment thereof.
[0114] In some embodiments according to the present invention, the
antigenic unit comprises an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to the amino acid
sequence 243-340 of SEQ ID NO:11.
[0115] In some embodiments according to the present invention, the
antigenic unit consists of an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to the amino acid
sequence 243-340 of SEQ ID NO:11.
[0116] In some embodiments according to the present invention, the
antigenic unit comprises one or more amino acid substitutions at a
position selected from the list consisting of C24, E26, C58, C61,
C91, and C94 of SEQ ID NO:23, or a deletion involving one or more
amino acid selected from the list consisting of L22-E26 and/or
C58-C61 and/or C91-595 of SEQ ID NO:23.
[0117] In some embodiments according to the present invention, the
antigenic unit comprises not more than 1, 2, 3, 4, 5, 6, 7, 8, 9,
10, 12, 14, 16, 18 or 20 amino acid substitutions and/or deletions
relative to SEQ ID NO:23.
[0118] In some embodiments according to the present invention, the
antigenic unit comprises the amino acid sequence 243-340 of SEQ ID
NO:11, SEQ ID NO:13, SEQ ID NO:15, or SEQ ID NO:17, or a variant or
antigenic fragment thereof.
[0119] In some embodiments according to the present invention, the
antigenic unit consists of the amino acid sequence 243-340 of SEQ
ID NO:11, SEQ ID NO:13, SEQ ID NO:15, or SEQ ID NO:17, or a variant
or antigenic fragment thereof.
[0120] In some embodiments according to the present invention, the
antigenic unit comprises an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to the amino acid
sequence 243-501 of SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:32, or
SEQ ID NO:34.
[0121] In some embodiments according to the present invention, the
antigenic unit consists of an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to the amino acid
sequence 243-501 of SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:32, or
SEQ ID NO:34.
[0122] In some embodiments according to the present invention, the
antigenic unit comprising an amino acid sequence of human
papillomavirus 16 (HPV16) derived from both early proteins E6 and
E7.
[0123] In some embodiments according to the present invention, the
antigenic unit comprising an amino acid sequence of human
papillomavirus 18 (HPV18) derived from both early proteins E6 and
E7.
[0124] In some embodiments according to the present invention, the
antigenic unit comprises one or more amino acid substitutions at a
position selected from the list consisting of F47, L50G, C63, C106,
I128T of SEQ ID NO:22 and C24, E26, C58, C61, C91, C94 of SEQ ID
NO:23.
[0125] In some embodiments according to the present invention, the
antigenic unit comprises not more than 1, 2, 3, 4, 5, 6, 7, 8, 9,
10, 12, 14, 16, 18 or 20 amino acid substitutions and/or deletions
relative to SEQ ID NO:22 and SEQ ID NO:23.
[0126] In some embodiments according to the present invention, the
antigenic unit consists of the amino acid sequence 243-501 of SEQ
ID NO:19, SEQ ID NO:21, SEQ ID NO:32, or SEQ ID NO:34, or a variant
or antigenic fragment thereof.
[0127] In some embodiments according to the present invention, the
amino acid chain consists of an amino acid sequence selected from
the list consisting of SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ
ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17,
SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:32, and SEQ ID NO:34, or a
variant or antigenic fragment thereof.
[0128] In some embodiments according to the present invention, the
antigenic unit comprises an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to any one amino acid
sequence selected from SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24,
and SEQ ID NO:25.
[0129] In some embodiments according to the present invention, the
antigenic unit consist of an amino acid sequence having at least
80%, such as at least 81%, such as at least 82%, such as at least
83%, such as at least 84%, such as at least 85%, such as at least
86%, such as at least 87%, such as at least 88%, such as at least
89%, such as at least 90%, such as at least 91%, such as at least
92%, such as at least 93%, such as at least 94%, such as at least
95%, such as at least 96%, such as at least 97%, such as at least
98%, such as at least 99% sequence identity to any one amino acid
sequence selected from SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24,
and SEQ ID NO:25.
[0130] In some embodiments the homodimeric protein according to the
present invention, is in its mature form without any signal peptide
sequence.
[0131] In some embodiments the nucleic acid molecule according to
the present invention is human codon optimized.
[0132] It is to be understood that a human codon optimized nucleic
acid molecule according to the present invention comprises one or
more nucleic acid substitution as compared to the wild type
sequence, which substitution provides for a codon with higher
frequency of usage in human coding regions. Frequency of codon
usage in homo sapiens can be found at
http://biowiki.edu-wiki.org/en/codon_table
[0133] In some embodiments the nucleic acid molecule according to
the present invention is comprising any one of nucleotide sequences
selected from the list consisting of SEQ ID NO:2, SEQ ID NO:4, SEQ
ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ
ID NO:16, SEQ ID NO:18, SEQ ID NO:20, SEQ ID NO:31 and SEQ ID
NO:33, or a variant thereof.
[0134] In some embodiments the nucleic acid molecule according to
the present invention is comprised by a vector.
[0135] In some embodiments the nucleic acid molecule according to
the present invention is formulated for administration to a patient
to induce production of the homodimeric protein in said
patient.
[0136] In some embodiments the vaccine according to the present
invention further comprises a pharmaceutically acceptable carrier
and/or adjuvant.
[0137] In some embodiments, the method of treating or preventing a
HPV induced disease or condition, such as a cancer or an infectious
disease caused by HPV in a patient according to the present
invention comprises administering to the patient in need thereof of
a nucleic acid molecule, such as a DNA, according to the present
invention with a subsequent step of electroporation. In some
embodiments the administration is performed intra dermal or intra
muscular.
EXAMPLE 1
[0138] Construction and Expression of the Vaccines.
[0139] Gene sequences were designed according to the following
structure: 1: native leader sequence for human LD78 b, 2: full
length LD78b sequence. 3: Human hinge-region 1 from IgG3. 4: Human
hinge region 4 from IgG3. 5: Glycine-Serine linker. 6: Human CH3
domain from IgG3. 7: Glycine-Leucine linker. 8: wildtype and mutant
Human papilloma virus oncogenes E6, E7 and fusion proteins of both
E6 and E7 divided by a Glycine-Serine linker. The constructs are
designated according to their E6 and or E7 composition as
follows:
[0140] VB1001: Vaccibody-E6 wild type;
[0141] VB1005: Vaccibody-E7 wild type;
[0142] The mutants are designated according to the amino acid
position in the corresponding native E6 or E7 sequence.
[0143] VB1002: Vaccibody-E6 C63R;
[0144] VB1003: Vaccibody-E6 C106R;
[0145] VB1004: Vaccibody-E6 F47R, C63R, C106R;
[0146] VB1006: Vaccibody-E7 C24G, E26G;
[0147] VB1007: Vaccibody-E7 C24G, E26G, C58G, C61G;
[0148] VB1008: Vaccibody-E7 C24G, E26G, C91G, C94G;
[0149] VB1009: Vaccibody-E7 C24G, E26G/E6 F47R, C63R, C106R;
[0150] VB1016: Vaccibody-E7 C24G, E26G/E6 C63R, C106R;
[0151] VB1020: Vaccibody-E7 C24G, E26G/E6 F47R, C63R, C106R human
codon optimized
[0152] VB1021: Vaccibody-E7 C24G, E26G/E6 F47R, L50G, C106R, I128T
human codon optimized
[0153] Control vaccines composed of only the antigens were
included:
[0154] Control 1: E7 C24G, E26G/E6 F47R, C63R, C106R; Control 2: E7
C24G, E26G/E6 C63R, C106R
[0155] All gene sequences were ordered from Aldevron (Fargo ND,
USA) or Eurofins MWG GmbH and cloned into the expression vector
pUMVC4a.
[0156] All constructs were transfected in to 293E cells and
verified expression of intact vaccibody proteins were performed by
dot blot and ELISA (data not shown). All amino acid sequences
except for Controls 1 and 2 are shown as SEQ IDs.
EXAMPLE 2
[0157] Immune Response Studies
[0158] VB 1009,VB1016, VB1020 and VB1021 were selected as vaccine
candidates with their corresponding controls 1 and 2 respectively.
As a negative control empty pUMVC4a vector was utilized.
[0159] 25, 12.5 and 1.4 .mu.g plasmid DNA of each candidate was
injected intradermal in the lower back of C57B1/6 mice followed by
electroporation, Dermavax, Cellectis (Paris, France). 7 days later
the mice were boosted with similar amounts of vaccines and control
plasmids. At day 21 the mice were killed and spleens were
harvested.
[0160] The T cell responses were calculated by ELISPOT. (FIGS. 3a,
b and c)
EXAMPLE 3.
[0161] Therapeutic Effect
[0162] VB1016, VB1020 and VB1021 with the corresponding controls 1
and 2 were selected as the vaccine candidate for therapeutic
vaccine studies.
[0163] 5.times.10.sup.4 or 5.times.10.sup.5 TC-1 cells (Johns
Hopkins University, Baltimore, USA, Lin K Y et al., Cancer Res,
1996) were injected in the neck or thigh region of C57B1/6 mice.
After days 3 and 10 or day 3,7 and 10, the mice were vaccinated
with 2 .mu.g, 10 .mu.g, 12.5 .mu.g or 20 .mu.g of plasmid DNA
followed by electroporation, Dermavax, Cellectis France. Tumor size
were measured two to three times a week up until day 49 after TC-1
cell injection (FIGS. 4, 5 and 6)
EXAMPLE 4
[0164] A therapeutic DNA vaccine to be used may be prepared by GMP
manufacturing of the plasmid vaccine according to regulatory
authorities' guidelines, including GMP cell banking, GMP
manufacturing of drug substance and drug product, ICH stability
studies and Fill & Finish of the DNA vaccine. The DNA vaccine
may be formulated by dissolving in a saline solution, such as 10nM
Tris, 1mM EDTA at a concentration of 2-5 mg/ml. The vaccine may be
administered either intra-dermal or intra-muscular with or without
following electroporation.
[0165] SEQUENCES:
[0166] C-C motif chemokine 3-like 1 precursor including signal
peptide (aa 1-23 in bold) and mature peptide (LD78-beta), aa 24-93
(SEQ ID NO:1):
TABLE-US-00002 MQVSTAALAVLLCTMALCNQVLSAPLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSA
[0167] The specific DNA and corresponding amino acid sequences of
vaccibody HPV constructs:
[0168] E6 or E7 single constructs:
[0169] For the purpose of illustration only, the different domains
of the constructs are separated by an "|" with the domains in the
following order: Signal peptide human MIP-1.alpha.|Hinge h1|Hinge
h4|Gly-Ser Linker or Gly-Leu linkers|hCH3 IgG3|Gly-Ser Linker or
Gly-Leu linkers|wildtype or mutant full length E6 or E7. Amino
acids or nucleotides in bold illustrates sites of mutations.
TABLE-US-00003 DNA sequence of VB1001 (SEQ ID NO: 2):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGTTTCAGGACCCACAGGAGCGACCCAGAAAGTTACCACAGTTATGCACAG-
AGCTG
CAAACAACTATACATGATATAATATTAGAATGTGTGTACTGCAAGCAACAGTTACTGCGACGTGAGGTATATGA-
CTTTG
CTTTTCGGGATTTATGCATAGTATATAGAGATGGGAATCCATATGCTGTATGTGATAAATGTTTAAAGTTTTAT-
TCTAA
AATTAGTGAGTATAGACATTATTGTTATAGTTTGTATGGAACAACATTAGAACAGCAATACAACAAACCGTTGT-
GTGAT
TTGTTAATTAGGTGTATTAACTGTCAAAAGCCACTGTGTCCTGAAGAAAAGCAAAGACATCTGGACAAAAAGCA-
AAGAT
TCCATAATATAAGGGGTCGGTGGACCGGTCGATGTATGTCTTGTTGCAGATCATCAAGAACACGTAGAGAAACC-
CAGCT GTAA Protein sequence of VB1001 (Homodimeric construct
according to the invention with E6, SEQ ID NO: 3): Amino acid
sequence 393 amino acids.
MQVSTAALAVLLCTMALCNQVLS|APLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSA|ELKTPLG
DTTHT|EPKSCDTPPPCPRCP|GGGSSGGGSG|GQPREPQVYTLPPSREEM
TKNQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSK
LTVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGK|GLGGL|MFQDPQ
ERPRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFAFRDLCIVYRDG
NPYAVCDKCLKFYSKISEYRHYCYSLYGTTLEQQYNKPLCDLLIRCINCQK
PLCPEEKQRHLDKKQRFHNIRGRWTGRCMSCCRSSRTRRETQL* DNA sequence of VB1002
(SEQ ID NO: 4):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGTTTCAGGACCCACAGGAGCGACCCAGAAAGTTACCACAGTTATGCACAG-
AGCTG
CAAACAACTATACATGATATAATATTAGAATGTGTGTACTGCAAGCAACAGTTACTGCGACGTGAGGTATATGA-
CTTTG
CTTTTCGGGATTTATGCATAGTATATAGAGATGGGAATCCATATGCTGTACGAGATAAATGTTTAAAGTTTTAT-
TCTAA
AATTAGTGAGTATAGACATTATTGTTATAGTTTGTATGGAACAACATTAGAACAGCAATACAACAAACCGTTGT-
GTGAT
TTGTTAATTAGGTGTATTAACTGTCAAAAGCCACTGTGTCCTGAAGAAAAGCAAAGACATCTGGACAAAAAGCA-
AAGAT
TCCATAATATAAGGGGTCGGTGGACCGGTCGATGTATGTCTTGTTGCAGATCATCAAGAACACGTAGAGAAACC-
CAGCT GTAA Protein sequence of VB1002 (Homodimeric construct
according to the invention, SEQ ID NO: 5): Amino acid sequence, 393
amino acids. MQVSTAALAVLLCTMALCNQVLS|APLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSA|ELKTPLG
DTTHT|EPKSCDTPPPCPRCP|GGGSSGGGSG|GQPREPQVYTLPPSREEM
TKNQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSK
LTVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGK|GLGGL|MFQDPQ
ERPRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFAFRDLCIVYRDG
NPYAVRDKCLKFYSKISEYRHYCYSLYGTTLEQQYNKPLCDLLIRCINCQK
PLCPEEKQRHLDKKQRFHNIRGRWTGRCMSCCRSSRTRRETQL* DNA sequence of VB
1003 (SEQ ID NO: 6):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGTTTCAGGACCCACAGGAGCGACCCAGAAAGTTACCACAGTTATGCACAG-
AGCTG
CAAACAACTATACATGATATAATATTAGAATGTGTGTACTGCAAGCAACAGTTACTGCGACGTGAGGTATATGA-
CTTTG
CTTTTCGGGATTTATGCATAGTATATAGAGATGGGAATCCATATGCTGTATGTGATAAATGTTTAAAGTTTTAT-
TCTAA
AATTAGTGAGTATAGACATTATTGTTATAGTTTGTATGGAACAACATTAGAACAGCAATACAACAAACCGTTGT-
GTGAT
TTGTTAATTAGGTGTATTAACCGACAAAAGCCACTGTGTCCTGAAGAAAAGCAAAGACATCTGGACAAAAAGCA-
AAGAT
TCCATAATATAAGGGGTCGGTGGACCGGTCGATGTATGTCTTGTTGCAGATCATCAAGAACACGTAGAGAAACC-
CAGCT GTAA Protein sequence of VB1003 (Homodimeric construct
according to the invention, SEQ ID NO: 7): Amino acid sequence, 393
amino acids. MQVSTAALAVLLCTMALCNQVLS|APLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSA|ELKTPLG
DTTHT|EPKSCDTPPPCPRCP|GGGSSGGGSG|GQPREPQVYTLPPSREEM
TKNQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSK
LTVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGK|GLGGL|MFQDPQ
ERPRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFAFRDLCIVYRDG
NPYAVCDKCLKFYSKISEYRHYCYSLYGTTLEQQYNKPLCDLLIRCINRQK
PLCPEEKQRHLDKKQRFHNIRGRWTGRCMSCCRSSRTRRETQL* DNA sequence of VB1004
(SEQ ID NO: 8):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGTTTCAGGACCCACAGGAGCGACCCAGAAAGTTACCACAGTTATGCACAG-
AGCTG
CAAACAACTATACATGATATAATATTAGAATGTGTGTACTGCAAGCAACAGTTACTGCGACGTGAGGTATATGA-
CTTTG
CTCGACGGGATTTATGCATAGTATATAGAGATGGGAATCCATATGCTGTACGAGATAAATGTTTAAAGTTTTAT-
TCTAA
AATTAGTGAGTATAGACATTATTGTTATAGTTTGTATGGAACAACATTAGAACAGCAATACAACAAACCGTTGT-
GTGAT
TTGTTAATTAGGTGTATTAACCGACAAAAGCCACTGTGTCCTGAAGAAAAGCAAAGACATCTGGACAAAAAGCA-
AAGAT
TCCATAATATAAGGGGTCGGTGGACCGGTCGATGTATGTCTTGTTGCAGATCATCAAGAACACGTAGAGAAACC-
CAGCT GTAA Protein sequence of VB1004 (Homodimeric construct
according to the invention, SEQ ID NO: 9): Amino acid sequence, 393
amino acids. MQVSTAALAVLLCTMALCNQVLSAPLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSAELKTPLG
DTTHTEPKSCDTPPPCPRCPGGGSSGGGSGGQPREPQVYTLPPSREEMTK
NQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSKL
TVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGKGLGGLMFQDPQER
PRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFARRDLCIVYRDGN
PYAVRDKCLKFYSKISEYRHYCYSLYGTTLEQQYNKPLCDLLIRCINRQK
PLCPEEKQRHLDKKQRFHNIRGRWTGRCMSCCRSSRTRRETQL* DNA sequence of VB1005
(SEQ ID NO: 10):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGCATGGAGATACACCTACATTGCATGAATATATGTTAGATTTGCAACCAG-
AGACA
ACTGATCTCTACTGTTATGAGCAATTAAATGACAGCTCAGAGGAGGAGGATGAAATAGATGGTCCAGCTGGACA-
AGCAG
AACCGGACAGAGCCCATTACAATATTGTAACCTTTTGTTGCAAGTGTGACTCTACGCTTCGGTTGTGCGTACAA-
AGCAC
ACACGTAGACATTCGTACTTTGGAAGACCTGTTAATGGGCACACTAGGAATTGTGTGCCCCATCTGTTCTCAGA-
AACCA TAA Protein sequence of VB1005 (Homodimeric construct
according to the invention with E7, SEQ ID NO: 11): Amino acid
sequence, 340 amino acids.
MQVSTAALAVLLCTMALCNQVLS|APLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSA|ELKTPLG
DTTHT|EPKSCDTPPPCPRCP|GGGSSGGGSG|GQPREPQVYTLPPSREEM
TKNQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSK
LTVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGK|GLGGL|MHGDTP
TLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTF
CCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKP* DNA sequence of VB1006
(SEQ ID NO: 12):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGCATGGAGATACACCTACATTGCATGAATATATGTTAGATTTGCAACCAG-
AGACA
ACTGATCTCTACGGATATGGACAATTAAATGACAGCTCAGAGGAGGAGGATGAAATAGATGGTCCAGCTGGACA-
AGCAG
AACCGGACAGAGCCCATTACAATATTGTAACCTTTTGTTGCAAGTGTGACTCTACGCTTCGGTTGTGCGTACAA-
AGCAC
ACACGTAGACATTCGTACTTTGGAAGACCTGTTAATGGGCACACTAGGAATTGTGTGCCCCATCTGTTCTCAGA-
AACCA TAA Protein sequence of VB1006 (Homodimeric construct
according to the invention, SEQ ID NO: 13): Amino acid sequence,
340 amino acids. MQVSTAALAVLLCTMALCNQVLSAPLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSAELKTPLG
DTTHTEPKSCDTPPPCPRCPGGGSSGGGSGGQPREPQVYTLPPSREEMTK
NQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSKL
TVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGKGLGGLMHGDTPTL
HEYMLDLQPETTDLYGYGQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFC
CKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKP* DNA sequence of VB1007
(SEQ ID NO: 14):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGCATGGAGATACACCTACATTGCATGAATATATGTTAGATTTGCAACCAG-
AGACA
ACTGATCTCTACGGATATGGACAATTAAATGACAGCTCAGAGGAGGAGGATGAAATAGATGGTCCAGCTGGACA-
AGCAG
AACCGGACAGAGCCCATTACAATATTGTAACCTTTGGATGCAAGGGAGACTCTACGCTTCGGTTGTGCGTACAA-
AGCAC
ACACGTAGACATTCGTACTTTGGAAGACCTGTTAATGGGCACACTAGGAATTGTGTGCCCCATCTGTTCTCAGA-
AACCA TAA Protein sequence of VB1007 (Homodimeric construct
according to the invention, SEQ ID NO: 15): Amino acid sequence,
340 amino acids. MQVSTAALAVLLCTMALCNQVLSAPLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSAELKTPLG
DTTHTEPKSCDTPPPCPRCPGGGSSGGGSGGQPREPQVYTLPPSREEMTK
NQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSKL
TVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGKGLGGLMHGDTPTL
HEYMLDLQPETTDLYGYGQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFG
CKGDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKP* DNA sequence of VB1008
(SEQ ID NO: 16):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGCATGGAGATACACCTACATTGCATGAATATATGTTAGATTTGCAACCAG-
AGACA
ACTGATCTCTACGGATATGGACAATTAAATGACAGCTCAGAGGAGGAGGATGAAATAGATGGTCCAGCTGGACA-
AGCAG
AACCGGACAGAGCCCATTACAATATTGTAACCTTTTGTTGCAAGTGTGACTCTACGCTTCGGTTGTGCGTACAA-
AGCAC
ACACGTAGACATTCGTACTTTGGAAGACCTGTTAATGGGCACACTAGGAATTGTGGGACCCATCGGATCTCAGA-
AACCA TAA Protein sequence of VB1008 (Homodimeric construct
according to the invention, SEQ ID NO: 17): Amino acid sequence,
340 amino acids. MQVSTAALAVLLCTMALCNQVLSAPLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSAELKTPLG
DTTHTEPKSCDTPPPCPRCPGGGSSGGGSGGQPREPQVYTLPPSREEMTK
NQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSKL
TVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGKGLGGLMHGDTPTL
HEYMLDLQPETTDLYGYGQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFC
CKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVGPIGSQKP*
[0170] Constructs with E6 and E7:
[0171] For the purpose of illustration only, the different domains
of the constructs are separated by an "|" with the domains in the
following order: Signal peptide|human MIP-1.alpha.|Hinge h1|Hinge
h4|Gly-Ser Linker or Gly-Leu linker|hCH3 IgG3|Gly-Ser Linker or
Gly-Leu linker|E7 mutant|Gly-Ser Linker or Gly-Leu linker|E6
mutant. Amino acids or nucleotides in bold illustrates sites of
mutations.
TABLE-US-00004 DNA sequence of VB1009 (SEQ ID NO: 18):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGCATGGAGATACACCTACATTGCATGAATATATGTTAGATTTGCAACCAG-
AGACA
ACTGATCTCTACGGATATGGACAATTAAATGACAGCTCAGAGGAGGAGGATGAAATAGATGGTCCAGCTGGACA-
AGCAG
AACCGGACAGAGCCCATTACAATATTGTAACCTTTTGTTGCAAGTGTGACTCTACGCTTCGGTTGTGCGTACAA-
AGCAC
ACACGTAGACATTCGTACTTTGGAAGACCTGTTAATGGGCACACTAGGAATTGTGTGCCCCATCTGTTCTCAGA-
AACCA
|GGCGGTGGAAGCAGCGGAGGTGGAAGTGGA|ATGTTTCAGGACCCACAGGAGCGACCCAGAAAGTTACCACAG-
TTATG
CACAGAGCTGCAAACAACTATACATGATATAATATTAGAATGTGTGTACTGCAAGCAACAGTTACTGCGACGTG-
AGGTA
TATGACTTTGCTCGACGGGATTTATGCATAGTATATAGAGATGGGAATCCATATGCTGTACGAGATAAATGTTT-
AAAGT
TTTATTCTAAAATTAGTGAGTATAGACATTATTGTTATAGTTTGTATGGAACAACATTAGAACAGCAATACAAC-
AAACC
GTTGTGTGATTTGTTAATTAGGTGTATTAACCGACAAAAGCCACTGTGTCCTGAAGAAAAGCAAAGACATCTGG-
ACAAA
AAGCAAAGATTCCATAATATAAGGGGTCGGTGGACCGGTCGATGTATGTCTTGTTGCAGATCATCAAGAACACG-
TAGAG AAACCCAGCTGTAA Protein sequence of VB1009 (Homodimeric
construct according to the invention, SEQ ID NO: 19): Amino acid
sequence, 501 amino acids.
MQVSTAALAVLLCTMALCNQVLS|APLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSA|ELKTPLG
DTTHT|EPKSCDTPPPCPRCP|GGGSSGGGSG|GQPREPQVYTLPPSREEM
TKNQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSK
LTVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGK|GLGGL|MHGDTP
TLHEYMLDLQPETTDLYGYGQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTF
CCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKP|GGGSSGGGS
G|MFQDPQERPRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFARRD
LCIVYRDGNPYAVRDKCLKFYSKISEYRHYCYSLYGTTLEQQYNKPLCDLL
IRCINRQKPLCPEEKQRHLDKKQRFHNIRGRWTGRCMSCCRSSRTRRETQL* DNA sequence
of VB1016 (SEQ ID NO: 20):
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG|ATGCATGGAGATACACCTACATTGCATGAATATATGTTAGATTTGCAACCAG-
AGACA
ACTGATCTCTACGGATATGGACAATTAAATGACAGCTCAGAGGAGGAGGATGAAATAGATGGTCCAGCTGGACA-
AGCAG
AACCGGACAGAGCCCATTACAATATTGTAACCTTTTGTTGCAAGTGTGACTCTACGCTTCGGTTGTGCGTACAA-
AGCAC
ACACGTAGACATTCGTACTTTGGAAGACCTGTTAATGGGCACACTAGGAATTGTGTGCCCCATCTGTTCTCAGA-
AACCA
1GGCGGTGGAAGCAGCGGAGGTGGAAGTGGA|ATGTTTCAGGACCCACAGGAGCGACCCAGAAAGTTACCACAG-
TTATG
CACAGAGCTGCAAACAACTATACATGATATAATATTAGAATGTGTGTACTGCAAGCAACAGTTACTGCGACGTG-
AGGTA
TATGACTTTGCTTTTCGGGATTTATGCATAGTATATAGAGATGGGAATCCATATGCTGTACGAGATAAATGTTT-
AAAGT
TTTATTCTAAAATTAGTGAGTATAGACATTATTGTTATAGTTTGTATGGAACAACATTAGAACAGCAATACAAC-
AAACC
GTTGTGTGATTTGTTAATTAGGTGTATTAACCGACAAAAGCCACTGTGTCCTGAAGAAAAGCAAAGACATCTGG-
ACAAA
AAGCAAAGATTCCATAATATAAGGGGTCGGTGGACCGGTCGATGTATGTCTTGTTGCAGATCATCAAGAACACG-
TAGAG AAACCCAGCTGTAA Protein sequence of VB1016 (Homodimeric
construct according to the invention, SEQ ID NO: 21): Amino acid
sequence, 501 amino acids
MQVSTAALAVLLCTMALCNQVLSAPLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSAELKTPLG
DTTHTEPKSCDTPPPCPRCPGGGSSGGGSGGQPREPQVYTLPPSREEMTK
NQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSKL
TVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGKGLGGLMHGDTPTL
HEYMLDLQPETTDLYGYGQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFC
CKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKPGGGSSGGGSG
MFQDPQERPRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFAFRDL
CIVYRDGNPYAVRDKCLKFYSKISEYRHYCYSLYGTTLEQQYNKPLCDLL
IRCINRQKPLCPEEKQRHLDKKQRFHNIRGRWTGRCMSCCRSSRTRRETQ L* SEQ ID NO:
22: >tr|Q778I6|Q778I6_HPV16 E6 protein OS = Human papillomavirus
type 16 GN = E6 PE = 4 SV = 1; (Underlined amino acids denotes
amino acids that may be deleted; Potential amino acids that may be
mutated are highlighted)
MFQDPQERPRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFAFRDLCIVYRDGNPYAVCDKCLKFY
SKISEYRHYCYSLYGTTLEQQYNKPLCDLLIRCINCQKPLCPEEKQRHLDKKQRFHNIRGRWTGRCMSCC
RSSRTRRETQL SEQ ID NO: 23: >sp|P03129|VE7_HPV16 Protein E7 OS =
Human papillomavirus type 16 GN = E7 PE = 1 SV = 1; (Underlined
amino acids denotes amino acids that may be deleted; Potential
amino acids that may be mutated are highlighted)
MHGDTPTLHEYMLDLQPETTDLYCYEQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTFCCKCDSTLRLCVQ
STHVDIRTLEDLLMGTLGIVCPICSQKP SEQ ID NO: 24: >sp|P06463|VE6_HPV18
Protein E6 OS = Human papillomavirus type 18 GN = E6 PE = 1 SV = 1
MARFEDPTRRPYKLPDLCTELNTSLQDIEITCVYCKTVLELTEVFEFAFKDLFVVYRDSI
PHAACHKCIDFYSRIRELRHYSDSVYGDTLEKLTNTGLYNLLIRCLRCQKPLNPAEKLRH
LNEKRRFHNIAGHYRGQCHSCCNRARQERLQRRRETQV SEQ ID NO: 25:
>sp|P06788|VE7_HPV18 Protein E7 OS = Human papillomavirus type
18 GN = E7 PE = 3 SV = 2
MHGPKATLQDIVLHLEPQNEIPVDLLCHEQLSDSEEENDEIDGVNHQHLPARRAEPQRHT
MLCMCCKCEARIKLVVESSADDLRAFQQLFLNTLSFVCPWCASQQ SEQ ID NO: 26: Hinge
regions (IgG3 UH hinge), 12 amino acids: ELKTPLGDTTHT SEQ ID NO:
27: Hinge region (IgG3, MH hinge, 15 amino acids): EPKSCDTPPPCPRCP
SEQ ID NO: 28: Gly-Ser Linker: GGGSSGGGSG SEQ ID NO: 29: hCH3 IgG3:
GQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSKL
TVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGK SEQ ID NO: 30: Linker: GLGGL
SEQ ID NO: 31: DNA sequence of VB1020:
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
AIGAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCAIGGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA|GGCCTCGGTGGCCTG/ATGCATGGCGATACCCCAACACTCCATGAGTACATGCTGGACCTTCAGCCCG-
AGAC
TACGGATCTGTATGGCTATGGGCAGTTGAATGACTCATCTGAGGAGGAGGACGAAATAGACGGCCCAGCTGGTC-
AAGCC
GAACCGGATAGAGCCCACTACAACATTGTGACCTTTTGCTGTAAGTGTGACAGCACTCTGAGACTGTGTGTTCA-
GTCCA
CTCATGTCGACATACGCACATTGGAGGATCTCCTGATGGGAACACTGGGAATTGTGTGTCCCATCTGTTCCCAA-
AAGCC
T/GGAGGTGGAAGCAGTGGAGGCGGTTCAGGC/ATGTTCCAAGATCCTCAAGAACGTCCTCGTAAGCTGCCACA-
GCTGT
GTACCGAGCTTCAGACCACCATTCACGACATCATCCTGGAGTGCGTCTATTGCAAACAGCAGCTCCTTAGAAGG-
GAAGT
GTACGATTTTGCACGGAGGGACCTCTGCATCGTGTATCGGGACGGCAATCCCTATGCGGTACGGGATAAATGCC-
TGAAG
TTCTACAGCAAAATCTCCGAGTACCGGCACTACTGCTACTCTCTCTATGGGACGACTCTGGAACAGCAGTACAA-
CAAGC
CCTTGTGCGATCTGCTGATTCGCTGCATTAATCGCCAGAAACCTCTGTGCCCAGAAGAGAAGCAAAGACACCTG-
GACAA
GAAACAGCGATTCCACAACATCCGAGGGAGATGGACAGGGAGGTGTATGAGCTGCTGTCGGAGTTCTAGGACAA-
GGCGC GAAACCCAGCTTTGA SEQ ID NO: 32: Protein sequence of VB1020
(Homodimeric construct according to the invention Amino acid
sequence, 501 amino acids:
MQVSTAALAVLLCTMALCNQVLS|APLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSA|ELKTPLG
DTTHT|EPKSCDTPPPCPRCP|GGGSSGGGSG|GQPREPQVYTLPPSREEM
TKNQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSK
LTVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGK|GLGGL|MHGDTP
TLHEYMLDLQPETTDLYGYGQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTF
CCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKP|GGGSSGGGS
G|MFQDPQERPRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFARRD
LCIVYRDGNPYAVRDKCLKFYSKISEYRHYCYSLYGTTLEQQYNKPLCDLL
IRCINRQKPLCPEEKQRHLDKKQRFHNIRGRWTGRCMSCCRSSRTRRETQL* SEQ ID NO: 33:
DNA sequence of VB1021:
ATGCAGGTCTCCACTGCTGCCCTTGCCGTCCTCCTCTGCACCATGGCTCTCTGCAACCAGGTCCTCTCT|GCAC-
CACTT
GCTGCTGACACGCCGACCGCCTGCTGCTTCAGCTACACCTCCCGACAGATTCCACAGAATTTCATAGCTGACTA-
CTTTG
AGACGAGCAGCCAGTGCTCCAAGCCCAGTGTCATCTTCCTAACCAAGAGAGGCCGGCAGGTCTGTGCTGACCCC-
AGTGA
GGAGTGGGTCCAGAAATACGTCAGTGACCTGGAGCTGAGTGCC|GAGCTCAAAACCCCACTTGGTGACACAACT-
CACAC
A|GAGCCCAAATCTTGTGACACACCTCCCCCGTGCCCAAGGTGCCCA|GGCGGTGGAAGCAGCGGAGGTGGAAG-
TGGA|
GGACAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCT-
GACCT
GCCTGGTCAAAGGCTTCTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAGCGGGCAGCCGGAGAACAACTAC-
AACAC
CACGCCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGC-
AGCAG
GGGAACATCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCGCTTCACGCAGAAGAGCCTCTCCCTGTC-
TCCGG
GTAAA/GGCCTCGGTGGCCTG/ATGCATGGTGACACACCAACCCTGCACGAATACATGCTCGATCTGCAGCCAG-
AGACT
ACCGACCTTTACGGCTATGGGCAGTTGAACGACAGCTCTGAGGAGGAGGACGAGATCGATGGTCCTGCTGGACA-
AGCAG
AACCAGACAGAGCCCACTACAACATCGTAACCTTTTGCTGCAAGTGTGACAGTACCCTTCGTTTGTGCGTTCAG-
AGCAC
GCATGTCGACATTCGGACACTGGAGGATCTGCTCATGGGGACTCTGGGGATTGTGTGTCCTATTTGCAGCCAGA-
AACCA
/GGCGGAGGATCTTCAGGAGGCGGGAGTGGC/ATGTTCCAAGACCCTCAGGAACGCCCTCGGAAACTGCCCCAA-
TTGTG
TACTGAGCTCCAGACAACGATACACGACATAATCCTGGAGTGCGTGTATTGCAAGCAGCAGCTTCTGAGGAGGG-
AAGTG
TACGATTTTGCCAGGAGAGATGGCTGCATTGTCTACCGAGATGGCAATCCCTATGCGGTGTGTGATAAGTGTCT-
GAAGT
TCTATTCCAAAATCAGCGAATATCGGCATTATTGCTACTCACTGTACGGAACTACCCTCGAACAGCAGTACAAC-
AAACC
GCTCTGTGATCTGCTGATCAGATGCATCAATCGGCAGAAACCCCTTTGTCCCGAAGAGAAGCAAAGACACCTGG-
ACAAG
AAGCAGAGGTTCCACAATACCCGAGGTCGTTGGACTGGGCGCTGCATGTCCTGTTGTCGCTCCTCTCGCACAAG-
GAGAG AGACACAACTGTGA SEQ ID NO: 34: Protein sequence of VB1021
(Homodimeric construct according to the invention. Amino acid
sequence, 501 amino acids:
MQVSTAALAVLLCTMALCNQVLS|APLAADTPTACCFSYTSRQIPQNFIAD
YFETSSQCSKPSVIFLTKRGRQVCADPSEEWVQKYVSDLELSA|ELKTPLG
DTTHT|EPKSCDTPPPCPRCP|GGGSSGGGSG|GQPREPQVYTLPPSREEM
TKNQVSLTCLVKGFYPSDIAVEWESSGQPENNYNTTPPMLDSDGSFFLYSK
LTVDKSRWQQGNIFSCSVMHEALHNRFTQKSLSLSPGK|GLGGL|MHGDTP
TLHEYMLDLQPETTDLYGYGQLNDSSEEEDEIDGPAGQAEPDRAHYNIVTF
CCKCDSTLRLCVQSTHVDIRTLEDLLMGTLGIVCPICSQKP|GGGSSGGGS
G|MFQDPQERPRKLPQLCTELQTTIHDIILECVYCKQQLLRREVYDFARRD
GCIVYRDGNPYAVCDKCLKFYSKISEYRHYCYSLYGTTLEQQYNKPLCDLL
IRCINRQKPLCPEEKQRHLDKKQRFHNTRGRWTGRCMSCCRSSRTRRETQL*
Sequence CWU 1
1
34193PRTArtificialSynthetic 1Met Gln Val Ser Thr Ala Ala Leu Ala
Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu Ser
Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe Ser
Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp Tyr
Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60 Phe
Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65 70
75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala 85 90
21182DNAArtificialSynthetic 2atgcaggtct ccactgctgc ccttgccgtc
ctcctctgca ccatggctct ctgcaaccag 60gtcctctctg caccacttgc tgctgacacg
ccgaccgcct gctgcttcag ctacacctcc 120cgacagattc cacagaattt
catagctgac tactttgaga cgagcagcca gtgctccaag 180cccagtgtca
tcttcctaac caagagaggc cggcaggtct gtgctgaccc cagtgaggag
240tgggtccaga aatacgtcag tgacctggag ctgagtgccg agctcaaaac
cccacttggt 300gacacaactc acacagagcc caaatcttgt gacacacctc
ccccgtgccc aaggtgccca 360ggcggtggaa gcagcggagg tggaagtgga
ggacagcccc gagaaccaca ggtgtacacc 420ctgcccccat cccgggagga
gatgaccaag aaccaggtca gcctgacctg cctggtcaaa 480ggcttctacc
ccagcgacat cgccgtggag tgggagagca gcgggcagcc ggagaacaac
540tacaacacca cgcctcccat gctggactcc gacggctcct tcttcctcta
cagcaagctc 600accgtggaca agagcaggtg gcagcagggg aacatcttct
catgctccgt gatgcatgag 660gctctgcaca accgcttcac gcagaagagc
ctctccctgt ctccgggtaa aggcctcggt 720ggcctgatgt ttcaggaccc
acaggagcga cccagaaagt taccacagtt atgcacagag 780ctgcaaacaa
ctatacatga tataatatta gaatgtgtgt actgcaagca acagttactg
840cgacgtgagg tatatgactt tgcttttcgg gatttatgca tagtatatag
agatgggaat 900ccatatgctg tatgtgataa atgtttaaag ttttattcta
aaattagtga gtatagacat 960tattgttata gtttgtatgg aacaacatta
gaacagcaat acaacaaacc gttgtgtgat 1020ttgttaatta ggtgtattaa
ctgtcaaaag ccactgtgtc ctgaagaaaa gcaaagacat 1080ctggacaaaa
agcaaagatt ccataatata aggggtcggt ggaccggtcg atgtatgtct
1140tgttgcagat catcaagaac acgtagagaa acccagctgt aa
11823393PRTArtificialSynthetic 3Met Gln Val Ser Thr Ala Ala Leu Ala
Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu Ser
Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe Ser
Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp Tyr
Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60 Phe
Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65 70
75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu Leu
Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys Ser
Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly Gly
Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro Gln
Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys Asn
Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr Pro
Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro Glu
Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185 190
Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln 195
200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu His
Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys
Gly Leu Gly 225 230 235 240 Gly Leu Met Phe Gln Asp Pro Gln Glu Arg
Pro Arg Lys Leu Pro Gln 245 250 255 Leu Cys Thr Glu Leu Gln Thr Thr
Ile His Asp Ile Ile Leu Glu Cys 260 265 270 Val Tyr Cys Lys Gln Gln
Leu Leu Arg Arg Glu Val Tyr Asp Phe Ala 275 280 285 Phe Arg Asp Leu
Cys Ile Val Tyr Arg Asp Gly Asn Pro Tyr Ala Val 290 295 300 Cys Asp
Lys Cys Leu Lys Phe Tyr Ser Lys Ile Ser Glu Tyr Arg His 305 310 315
320 Tyr Cys Tyr Ser Leu Tyr Gly Thr Thr Leu Glu Gln Gln Tyr Asn Lys
325 330 335 Pro Leu Cys Asp Leu Leu Ile Arg Cys Ile Asn Cys Gln Lys
Pro Leu 340 345 350 Cys Pro Glu Glu Lys Gln Arg His Leu Asp Lys Lys
Gln Arg Phe His 355 360 365 Asn Ile Arg Gly Arg Trp Thr Gly Arg Cys
Met Ser Cys Cys Arg Ser 370 375 380 Ser Arg Thr Arg Arg Glu Thr Gln
Leu 385 390 41182DNAArtificialSynthetic 4atgcaggtct ccactgctgc
ccttgccgtc ctcctctgca ccatggctct ctgcaaccag 60gtcctctctg caccacttgc
tgctgacacg ccgaccgcct gctgcttcag ctacacctcc 120cgacagattc
cacagaattt catagctgac tactttgaga cgagcagcca gtgctccaag
180cccagtgtca tcttcctaac caagagaggc cggcaggtct gtgctgaccc
cagtgaggag 240tgggtccaga aatacgtcag tgacctggag ctgagtgccg
agctcaaaac cccacttggt 300gacacaactc acacagagcc caaatcttgt
gacacacctc ccccgtgccc aaggtgccca 360ggcggtggaa gcagcggagg
tggaagtgga ggacagcccc gagaaccaca ggtgtacacc 420ctgcccccat
cccgggagga gatgaccaag aaccaggtca gcctgacctg cctggtcaaa
480ggcttctacc ccagcgacat cgccgtggag tgggagagca gcgggcagcc
ggagaacaac 540tacaacacca cgcctcccat gctggactcc gacggctcct
tcttcctcta cagcaagctc 600accgtggaca agagcaggtg gcagcagggg
aacatcttct catgctccgt gatgcatgag 660gctctgcaca accgcttcac
gcagaagagc ctctccctgt ctccgggtaa aggcctcggt 720ggcctgatgt
ttcaggaccc acaggagcga cccagaaagt taccacagtt atgcacagag
780ctgcaaacaa ctatacatga tataatatta gaatgtgtgt actgcaagca
acagttactg 840cgacgtgagg tatatgactt tgcttttcgg gatttatgca
tagtatatag agatgggaat 900ccatatgctg tacgagataa atgtttaaag
ttttattcta aaattagtga gtatagacat 960tattgttata gtttgtatgg
aacaacatta gaacagcaat acaacaaacc gttgtgtgat 1020ttgttaatta
ggtgtattaa ctgtcaaaag ccactgtgtc ctgaagaaaa gcaaagacat
1080ctggacaaaa agcaaagatt ccataatata aggggtcggt ggaccggtcg
atgtatgtct 1140tgttgcagat catcaagaac acgtagagaa acccagctgt aa
11825393PRTArtificialSynthetic 5Met Gln Val Ser Thr Ala Ala Leu Ala
Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu Ser
Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe Ser
Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp Tyr
Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60 Phe
Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65 70
75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu Leu
Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys Ser
Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly Gly
Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro Gln
Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys Asn
Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr Pro
Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro Glu
Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185 190
Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln 195
200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu His
Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys
Gly Leu Gly 225 230 235 240 Gly Leu Met Phe Gln Asp Pro Gln Glu Arg
Pro Arg Lys Leu Pro Gln 245 250 255 Leu Cys Thr Glu Leu Gln Thr Thr
Ile His Asp Ile Ile Leu Glu Cys 260 265 270 Val Tyr Cys Lys Gln Gln
Leu Leu Arg Arg Glu Val Tyr Asp Phe Ala 275 280 285 Phe Arg Asp Leu
Cys Ile Val Tyr Arg Asp Gly Asn Pro Tyr Ala Val 290 295 300 Arg Asp
Lys Cys Leu Lys Phe Tyr Ser Lys Ile Ser Glu Tyr Arg His 305 310 315
320 Tyr Cys Tyr Ser Leu Tyr Gly Thr Thr Leu Glu Gln Gln Tyr Asn Lys
325 330 335 Pro Leu Cys Asp Leu Leu Ile Arg Cys Ile Asn Cys Gln Lys
Pro Leu 340 345 350 Cys Pro Glu Glu Lys Gln Arg His Leu Asp Lys Lys
Gln Arg Phe His 355 360 365 Asn Ile Arg Gly Arg Trp Thr Gly Arg Cys
Met Ser Cys Cys Arg Ser 370 375 380 Ser Arg Thr Arg Arg Glu Thr Gln
Leu 385 390 61182DNAArtificialSynthetic 6atgcaggtct ccactgctgc
ccttgccgtc ctcctctgca ccatggctct ctgcaaccag 60gtcctctctg caccacttgc
tgctgacacg ccgaccgcct gctgcttcag ctacacctcc 120cgacagattc
cacagaattt catagctgac tactttgaga cgagcagcca gtgctccaag
180cccagtgtca tcttcctaac caagagaggc cggcaggtct gtgctgaccc
cagtgaggag 240tgggtccaga aatacgtcag tgacctggag ctgagtgccg
agctcaaaac cccacttggt 300gacacaactc acacagagcc caaatcttgt
gacacacctc ccccgtgccc aaggtgccca 360ggcggtggaa gcagcggagg
tggaagtgga ggacagcccc gagaaccaca ggtgtacacc 420ctgcccccat
cccgggagga gatgaccaag aaccaggtca gcctgacctg cctggtcaaa
480ggcttctacc ccagcgacat cgccgtggag tgggagagca gcgggcagcc
ggagaacaac 540tacaacacca cgcctcccat gctggactcc gacggctcct
tcttcctcta cagcaagctc 600accgtggaca agagcaggtg gcagcagggg
aacatcttct catgctccgt gatgcatgag 660gctctgcaca accgcttcac
gcagaagagc ctctccctgt ctccgggtaa aggcctcggt 720ggcctgatgt
ttcaggaccc acaggagcga cccagaaagt taccacagtt atgcacagag
780ctgcaaacaa ctatacatga tataatatta gaatgtgtgt actgcaagca
acagttactg 840cgacgtgagg tatatgactt tgcttttcgg gatttatgca
tagtatatag agatgggaat 900ccatatgctg tatgtgataa atgtttaaag
ttttattcta aaattagtga gtatagacat 960tattgttata gtttgtatgg
aacaacatta gaacagcaat acaacaaacc gttgtgtgat 1020ttgttaatta
ggtgtattaa ccgacaaaag ccactgtgtc ctgaagaaaa gcaaagacat
1080ctggacaaaa agcaaagatt ccataatata aggggtcggt ggaccggtcg
atgtatgtct 1140tgttgcagat catcaagaac acgtagagaa acccagctgt aa
11827393PRTArtificialSynthetic 7Met Gln Val Ser Thr Ala Ala Leu Ala
Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu Ser
Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe Ser
Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp Tyr
Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60 Phe
Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65 70
75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu Leu
Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys Ser
Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly Gly
Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro Gln
Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys Asn
Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr Pro
Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro Glu
Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185 190
Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln 195
200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu His
Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys
Gly Leu Gly 225 230 235 240 Gly Leu Met Phe Gln Asp Pro Gln Glu Arg
Pro Arg Lys Leu Pro Gln 245 250 255 Leu Cys Thr Glu Leu Gln Thr Thr
Ile His Asp Ile Ile Leu Glu Cys 260 265 270 Val Tyr Cys Lys Gln Gln
Leu Leu Arg Arg Glu Val Tyr Asp Phe Ala 275 280 285 Phe Arg Asp Leu
Cys Ile Val Tyr Arg Asp Gly Asn Pro Tyr Ala Val 290 295 300 Cys Asp
Lys Cys Leu Lys Phe Tyr Ser Lys Ile Ser Glu Tyr Arg His 305 310 315
320 Tyr Cys Tyr Ser Leu Tyr Gly Thr Thr Leu Glu Gln Gln Tyr Asn Lys
325 330 335 Pro Leu Cys Asp Leu Leu Ile Arg Cys Ile Asn Arg Gln Lys
Pro Leu 340 345 350 Cys Pro Glu Glu Lys Gln Arg His Leu Asp Lys Lys
Gln Arg Phe His 355 360 365 Asn Ile Arg Gly Arg Trp Thr Gly Arg Cys
Met Ser Cys Cys Arg Ser 370 375 380 Ser Arg Thr Arg Arg Glu Thr Gln
Leu 385 390 81182DNAArtificialSynthetic 8atgcaggtct ccactgctgc
ccttgccgtc ctcctctgca ccatggctct ctgcaaccag 60gtcctctctg caccacttgc
tgctgacacg ccgaccgcct gctgcttcag ctacacctcc 120cgacagattc
cacagaattt catagctgac tactttgaga cgagcagcca gtgctccaag
180cccagtgtca tcttcctaac caagagaggc cggcaggtct gtgctgaccc
cagtgaggag 240tgggtccaga aatacgtcag tgacctggag ctgagtgccg
agctcaaaac cccacttggt 300gacacaactc acacagagcc caaatcttgt
gacacacctc ccccgtgccc aaggtgccca 360ggcggtggaa gcagcggagg
tggaagtgga ggacagcccc gagaaccaca ggtgtacacc 420ctgcccccat
cccgggagga gatgaccaag aaccaggtca gcctgacctg cctggtcaaa
480ggcttctacc ccagcgacat cgccgtggag tgggagagca gcgggcagcc
ggagaacaac 540tacaacacca cgcctcccat gctggactcc gacggctcct
tcttcctcta cagcaagctc 600accgtggaca agagcaggtg gcagcagggg
aacatcttct catgctccgt gatgcatgag 660gctctgcaca accgcttcac
gcagaagagc ctctccctgt ctccgggtaa aggcctcggt 720ggcctgatgt
ttcaggaccc acaggagcga cccagaaagt taccacagtt atgcacagag
780ctgcaaacaa ctatacatga tataatatta gaatgtgtgt actgcaagca
acagttactg 840cgacgtgagg tatatgactt tgctcgacgg gatttatgca
tagtatatag agatgggaat 900ccatatgctg tacgagataa atgtttaaag
ttttattcta aaattagtga gtatagacat 960tattgttata gtttgtatgg
aacaacatta gaacagcaat acaacaaacc gttgtgtgat 1020ttgttaatta
ggtgtattaa ccgacaaaag ccactgtgtc ctgaagaaaa gcaaagacat
1080ctggacaaaa agcaaagatt ccataatata aggggtcggt ggaccggtcg
atgtatgtct 1140tgttgcagat catcaagaac acgtagagaa acccagctgt aa
11829393PRTArtificialSynthetic 9Met Gln Val Ser Thr Ala Ala Leu Ala
Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu Ser
Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe Ser
Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp Tyr
Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60 Phe
Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65 70
75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu Leu
Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys Ser
Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly Gly
Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro Gln
Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys Asn
Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr Pro
Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro Glu
Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185 190
Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln 195
200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu His
Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys
Gly Leu Gly 225 230 235 240 Gly Leu Met Phe Gln Asp Pro Gln Glu Arg
Pro Arg Lys Leu Pro Gln 245 250
255 Leu Cys Thr Glu Leu Gln Thr Thr Ile His Asp Ile Ile Leu Glu Cys
260 265 270 Val Tyr Cys Lys Gln Gln Leu Leu Arg Arg Glu Val Tyr Asp
Phe Ala 275 280 285 Arg Arg Asp Leu Cys Ile Val Tyr Arg Asp Gly Asn
Pro Tyr Ala Val 290 295 300 Arg Asp Lys Cys Leu Lys Phe Tyr Ser Lys
Ile Ser Glu Tyr Arg His 305 310 315 320 Tyr Cys Tyr Ser Leu Tyr Gly
Thr Thr Leu Glu Gln Gln Tyr Asn Lys 325 330 335 Pro Leu Cys Asp Leu
Leu Ile Arg Cys Ile Asn Arg Gln Lys Pro Leu 340 345 350 Cys Pro Glu
Glu Lys Gln Arg His Leu Asp Lys Lys Gln Arg Phe His 355 360 365 Asn
Ile Arg Gly Arg Trp Thr Gly Arg Cys Met Ser Cys Cys Arg Ser 370 375
380 Ser Arg Thr Arg Arg Glu Thr Gln Leu 385 390
101023DNAArtificialSynthetic 10atgcaggtct ccactgctgc ccttgccgtc
ctcctctgca ccatggctct ctgcaaccag 60gtcctctctg caccacttgc tgctgacacg
ccgaccgcct gctgcttcag ctacacctcc 120cgacagattc cacagaattt
catagctgac tactttgaga cgagcagcca gtgctccaag 180cccagtgtca
tcttcctaac caagagaggc cggcaggtct gtgctgaccc cagtgaggag
240tgggtccaga aatacgtcag tgacctggag ctgagtgccg agctcaaaac
cccacttggt 300gacacaactc acacagagcc caaatcttgt gacacacctc
ccccgtgccc aaggtgccca 360ggcggtggaa gcagcggagg tggaagtgga
ggacagcccc gagaaccaca ggtgtacacc 420ctgcccccat cccgggagga
gatgaccaag aaccaggtca gcctgacctg cctggtcaaa 480ggcttctacc
ccagcgacat cgccgtggag tgggagagca gcgggcagcc ggagaacaac
540tacaacacca cgcctcccat gctggactcc gacggctcct tcttcctcta
cagcaagctc 600accgtggaca agagcaggtg gcagcagggg aacatcttct
catgctccgt gatgcatgag 660gctctgcaca accgcttcac gcagaagagc
ctctccctgt ctccgggtaa aggcctcggt 720ggcctgatgc atggagatac
acctacattg catgaatata tgttagattt gcaaccagag 780acaactgatc
tctactgtta tgagcaatta aatgacagct cagaggagga ggatgaaata
840gatggtccag ctggacaagc agaaccggac agagcccatt acaatattgt
aaccttttgt 900tgcaagtgtg actctacgct tcggttgtgc gtacaaagca
cacacgtaga cattcgtact 960ttggaagacc tgttaatggg cacactagga
attgtgtgcc ccatctgttc tcagaaacca 1020taa
102311340PRTArtificialSynthetic 11Met Gln Val Ser Thr Ala Ala Leu
Ala Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu
Ser Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe
Ser Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp
Tyr Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60
Phe Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65
70 75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu
Leu Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys
Ser Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly
Gly Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro
Gln Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys
Asn Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr
Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro
Glu Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185
190 Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln
195 200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu
His Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly
Lys Gly Leu Gly 225 230 235 240 Gly Leu Met His Gly Asp Thr Pro Thr
Leu His Glu Tyr Met Leu Asp 245 250 255 Leu Gln Pro Glu Thr Thr Asp
Leu Tyr Cys Tyr Glu Gln Leu Asn Asp 260 265 270 Ser Ser Glu Glu Glu
Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu 275 280 285 Pro Asp Arg
Ala His Tyr Asn Ile Val Thr Phe Cys Cys Lys Cys Asp 290 295 300 Ser
Thr Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile Arg Thr 305 310
315 320 Leu Glu Asp Leu Leu Met Gly Thr Leu Gly Ile Val Cys Pro Ile
Cys 325 330 335 Ser Gln Lys Pro 340 121023DNAArtificialSynthetic
12atgcaggtct ccactgctgc ccttgccgtc ctcctctgca ccatggctct ctgcaaccag
60gtcctctctg caccacttgc tgctgacacg ccgaccgcct gctgcttcag ctacacctcc
120cgacagattc cacagaattt catagctgac tactttgaga cgagcagcca
gtgctccaag 180cccagtgtca tcttcctaac caagagaggc cggcaggtct
gtgctgaccc cagtgaggag 240tgggtccaga aatacgtcag tgacctggag
ctgagtgccg agctcaaaac cccacttggt 300gacacaactc acacagagcc
caaatcttgt gacacacctc ccccgtgccc aaggtgccca 360ggcggtggaa
gcagcggagg tggaagtgga ggacagcccc gagaaccaca ggtgtacacc
420ctgcccccat cccgggagga gatgaccaag aaccaggtca gcctgacctg
cctggtcaaa 480ggcttctacc ccagcgacat cgccgtggag tgggagagca
gcgggcagcc ggagaacaac 540tacaacacca cgcctcccat gctggactcc
gacggctcct tcttcctcta cagcaagctc 600accgtggaca agagcaggtg
gcagcagggg aacatcttct catgctccgt gatgcatgag 660gctctgcaca
accgcttcac gcagaagagc ctctccctgt ctccgggtaa aggcctcggt
720ggcctgatgc atggagatac acctacattg catgaatata tgttagattt
gcaaccagag 780acaactgatc tctacggata tggacaatta aatgacagct
cagaggagga ggatgaaata 840gatggtccag ctggacaagc agaaccggac
agagcccatt acaatattgt aaccttttgt 900tgcaagtgtg actctacgct
tcggttgtgc gtacaaagca cacacgtaga cattcgtact 960ttggaagacc
tgttaatggg cacactagga attgtgtgcc ccatctgttc tcagaaacca 1020taa
102313340PRTArtificialSynthetic 13Met Gln Val Ser Thr Ala Ala Leu
Ala Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu
Ser Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe
Ser Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp
Tyr Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60
Phe Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65
70 75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu
Leu Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys
Ser Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly
Gly Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro
Gln Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys
Asn Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr
Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro
Glu Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185
190 Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln
195 200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu
His Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly
Lys Gly Leu Gly 225 230 235 240 Gly Leu Met His Gly Asp Thr Pro Thr
Leu His Glu Tyr Met Leu Asp 245 250 255 Leu Gln Pro Glu Thr Thr Asp
Leu Tyr Gly Tyr Gly Gln Leu Asn Asp 260 265 270 Ser Ser Glu Glu Glu
Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu 275 280 285 Pro Asp Arg
Ala His Tyr Asn Ile Val Thr Phe Cys Cys Lys Cys Asp 290 295 300 Ser
Thr Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile Arg Thr 305 310
315 320 Leu Glu Asp Leu Leu Met Gly Thr Leu Gly Ile Val Cys Pro Ile
Cys 325 330 335 Ser Gln Lys Pro 340 141023DNAArtificialSynthetic
14atgcaggtct ccactgctgc ccttgccgtc ctcctctgca ccatggctct ctgcaaccag
60gtcctctctg caccacttgc tgctgacacg ccgaccgcct gctgcttcag ctacacctcc
120cgacagattc cacagaattt catagctgac tactttgaga cgagcagcca
gtgctccaag 180cccagtgtca tcttcctaac caagagaggc cggcaggtct
gtgctgaccc cagtgaggag 240tgggtccaga aatacgtcag tgacctggag
ctgagtgccg agctcaaaac cccacttggt 300gacacaactc acacagagcc
caaatcttgt gacacacctc ccccgtgccc aaggtgccca 360ggcggtggaa
gcagcggagg tggaagtgga ggacagcccc gagaaccaca ggtgtacacc
420ctgcccccat cccgggagga gatgaccaag aaccaggtca gcctgacctg
cctggtcaaa 480ggcttctacc ccagcgacat cgccgtggag tgggagagca
gcgggcagcc ggagaacaac 540tacaacacca cgcctcccat gctggactcc
gacggctcct tcttcctcta cagcaagctc 600accgtggaca agagcaggtg
gcagcagggg aacatcttct catgctccgt gatgcatgag 660gctctgcaca
accgcttcac gcagaagagc ctctccctgt ctccgggtaa aggcctcggt
720ggcctgatgc atggagatac acctacattg catgaatata tgttagattt
gcaaccagag 780acaactgatc tctacggata tggacaatta aatgacagct
cagaggagga ggatgaaata 840gatggtccag ctggacaagc agaaccggac
agagcccatt acaatattgt aacctttgga 900tgcaagggag actctacgct
tcggttgtgc gtacaaagca cacacgtaga cattcgtact 960ttggaagacc
tgttaatggg cacactagga attgtgtgcc ccatctgttc tcagaaacca 1020taa
102315340PRTArtificialSynthetic 15Met Gln Val Ser Thr Ala Ala Leu
Ala Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu
Ser Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe
Ser Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp
Tyr Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60
Phe Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65
70 75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu
Leu Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys
Ser Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly
Gly Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro
Gln Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys
Asn Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr
Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro
Glu Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185
190 Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln
195 200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu
His Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly
Lys Gly Leu Gly 225 230 235 240 Gly Leu Met His Gly Asp Thr Pro Thr
Leu His Glu Tyr Met Leu Asp 245 250 255 Leu Gln Pro Glu Thr Thr Asp
Leu Tyr Gly Tyr Gly Gln Leu Asn Asp 260 265 270 Ser Ser Glu Glu Glu
Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu 275 280 285 Pro Asp Arg
Ala His Tyr Asn Ile Val Thr Phe Gly Cys Lys Gly Asp 290 295 300 Ser
Thr Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile Arg Thr 305 310
315 320 Leu Glu Asp Leu Leu Met Gly Thr Leu Gly Ile Val Cys Pro Ile
Cys 325 330 335 Ser Gln Lys Pro 340 161023DNAArtificialSynthetic
16atgcaggtct ccactgctgc ccttgccgtc ctcctctgca ccatggctct ctgcaaccag
60gtcctctctg caccacttgc tgctgacacg ccgaccgcct gctgcttcag ctacacctcc
120cgacagattc cacagaattt catagctgac tactttgaga cgagcagcca
gtgctccaag 180cccagtgtca tcttcctaac caagagaggc cggcaggtct
gtgctgaccc cagtgaggag 240tgggtccaga aatacgtcag tgacctggag
ctgagtgccg agctcaaaac cccacttggt 300gacacaactc acacagagcc
caaatcttgt gacacacctc ccccgtgccc aaggtgccca 360ggcggtggaa
gcagcggagg tggaagtgga ggacagcccc gagaaccaca ggtgtacacc
420ctgcccccat cccgggagga gatgaccaag aaccaggtca gcctgacctg
cctggtcaaa 480ggcttctacc ccagcgacat cgccgtggag tgggagagca
gcgggcagcc ggagaacaac 540tacaacacca cgcctcccat gctggactcc
gacggctcct tcttcctcta cagcaagctc 600accgtggaca agagcaggtg
gcagcagggg aacatcttct catgctccgt gatgcatgag 660gctctgcaca
accgcttcac gcagaagagc ctctccctgt ctccgggtaa aggcctcggt
720ggcctgatgc atggagatac acctacattg catgaatata tgttagattt
gcaaccagag 780acaactgatc tctacggata tggacaatta aatgacagct
cagaggagga ggatgaaata 840gatggtccag ctggacaagc agaaccggac
agagcccatt acaatattgt aaccttttgt 900tgcaagtgtg actctacgct
tcggttgtgc gtacaaagca cacacgtaga cattcgtact 960ttggaagacc
tgttaatggg cacactagga attgtgggac ccatcggatc tcagaaacca 1020taa
102317340PRTArtificialSynthetic 17Met Gln Val Ser Thr Ala Ala Leu
Ala Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu
Ser Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe
Ser Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp
Tyr Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60
Phe Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65
70 75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu
Leu Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys
Ser Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly
Gly Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro
Gln Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys
Asn Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr
Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro
Glu Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185
190 Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln
195 200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu
His Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly
Lys Gly Leu Gly 225 230 235 240 Gly Leu Met His Gly Asp Thr Pro Thr
Leu His Glu Tyr Met Leu Asp 245 250 255 Leu Gln Pro Glu Thr Thr Asp
Leu Tyr Gly Tyr Gly Gln Leu Asn Asp 260 265 270 Ser Ser Glu Glu Glu
Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu 275 280 285 Pro Asp Arg
Ala His Tyr Asn Ile Val Thr Phe Cys Cys Lys Cys Asp 290 295 300 Ser
Thr Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile Arg Thr 305 310
315 320 Leu Glu Asp Leu Leu Met Gly Thr Leu Gly Ile Val Gly Pro Ile
Gly 325 330 335 Ser Gln Lys Pro 340 181506DNAArtificialSynthetic
18atgcaggtct ccactgctgc ccttgccgtc ctcctctgca ccatggctct ctgcaaccag
60gtcctctctg caccacttgc tgctgacacg ccgaccgcct gctgcttcag ctacacctcc
120cgacagattc cacagaattt catagctgac tactttgaga cgagcagcca
gtgctccaag 180cccagtgtca tcttcctaac caagagaggc cggcaggtct
gtgctgaccc cagtgaggag 240tgggtccaga aatacgtcag tgacctggag
ctgagtgccg agctcaaaac cccacttggt 300gacacaactc acacagagcc
caaatcttgt gacacacctc ccccgtgccc aaggtgccca 360ggcggtggaa
gcagcggagg tggaagtgga ggacagcccc gagaaccaca ggtgtacacc
420ctgcccccat cccgggagga gatgaccaag aaccaggtca gcctgacctg
cctggtcaaa 480ggcttctacc ccagcgacat cgccgtggag tgggagagca
gcgggcagcc ggagaacaac 540tacaacacca cgcctcccat gctggactcc
gacggctcct tcttcctcta cagcaagctc 600accgtggaca agagcaggtg
gcagcagggg aacatcttct catgctccgt gatgcatgag
660gctctgcaca accgcttcac gcagaagagc ctctccctgt ctccgggtaa
aggcctcggt 720ggcctgatgc atggagatac acctacattg catgaatata
tgttagattt gcaaccagag 780acaactgatc tctacggata tggacaatta
aatgacagct cagaggagga ggatgaaata 840gatggtccag ctggacaagc
agaaccggac agagcccatt acaatattgt aaccttttgt 900tgcaagtgtg
actctacgct tcggttgtgc gtacaaagca cacacgtaga cattcgtact
960ttggaagacc tgttaatggg cacactagga attgtgtgcc ccatctgttc
tcagaaacca 1020ggcggtggaa gcagcggagg tggaagtgga atgtttcagg
acccacagga gcgacccaga 1080aagttaccac agttatgcac agagctgcaa
acaactatac atgatataat attagaatgt 1140gtgtactgca agcaacagtt
actgcgacgt gaggtatatg actttgctcg acgggattta 1200tgcatagtat
atagagatgg gaatccatat gctgtacgag ataaatgttt aaagttttat
1260tctaaaatta gtgagtatag acattattgt tatagtttgt atggaacaac
attagaacag 1320caatacaaca aaccgttgtg tgatttgtta attaggtgta
ttaaccgaca aaagccactg 1380tgtcctgaag aaaagcaaag acatctggac
aaaaagcaaa gattccataa tataaggggt 1440cggtggaccg gtcgatgtat
gtcttgttgc agatcatcaa gaacacgtag agaaacccag 1500ctgtaa
150619501PRTArtificialSynthetic 19Met Gln Val Ser Thr Ala Ala Leu
Ala Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu
Ser Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe
Ser Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp
Tyr Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60
Phe Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65
70 75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu
Leu Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys
Ser Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly
Gly Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro
Gln Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys
Asn Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr
Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro
Glu Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185
190 Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln
195 200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu
His Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly
Lys Gly Leu Gly 225 230 235 240 Gly Leu Met His Gly Asp Thr Pro Thr
Leu His Glu Tyr Met Leu Asp 245 250 255 Leu Gln Pro Glu Thr Thr Asp
Leu Tyr Gly Tyr Gly Gln Leu Asn Asp 260 265 270 Ser Ser Glu Glu Glu
Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu 275 280 285 Pro Asp Arg
Ala His Tyr Asn Ile Val Thr Phe Cys Cys Lys Cys Asp 290 295 300 Ser
Thr Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile Arg Thr 305 310
315 320 Leu Glu Asp Leu Leu Met Gly Thr Leu Gly Ile Val Cys Pro Ile
Cys 325 330 335 Ser Gln Lys Pro Gly Gly Gly Ser Ser Gly Gly Gly Ser
Gly Met Phe 340 345 350 Gln Asp Pro Gln Glu Arg Pro Arg Lys Leu Pro
Gln Leu Cys Thr Glu 355 360 365 Leu Gln Thr Thr Ile His Asp Ile Ile
Leu Glu Cys Val Tyr Cys Lys 370 375 380 Gln Gln Leu Leu Arg Arg Glu
Val Tyr Asp Phe Ala Arg Arg Asp Leu 385 390 395 400 Cys Ile Val Tyr
Arg Asp Gly Asn Pro Tyr Ala Val Arg Asp Lys Cys 405 410 415 Leu Lys
Phe Tyr Ser Lys Ile Ser Glu Tyr Arg His Tyr Cys Tyr Ser 420 425 430
Leu Tyr Gly Thr Thr Leu Glu Gln Gln Tyr Asn Lys Pro Leu Cys Asp 435
440 445 Leu Leu Ile Arg Cys Ile Asn Arg Gln Lys Pro Leu Cys Pro Glu
Glu 450 455 460 Lys Gln Arg His Leu Asp Lys Lys Gln Arg Phe His Asn
Ile Arg Gly 465 470 475 480 Arg Trp Thr Gly Arg Cys Met Ser Cys Cys
Arg Ser Ser Arg Thr Arg 485 490 495 Arg Glu Thr Gln Leu 500
201506DNAArtificialSynthetic 20atgcaggtct ccactgctgc ccttgccgtc
ctcctctgca ccatggctct ctgcaaccag 60gtcctctctg caccacttgc tgctgacacg
ccgaccgcct gctgcttcag ctacacctcc 120cgacagattc cacagaattt
catagctgac tactttgaga cgagcagcca gtgctccaag 180cccagtgtca
tcttcctaac caagagaggc cggcaggtct gtgctgaccc cagtgaggag
240tgggtccaga aatacgtcag tgacctggag ctgagtgccg agctcaaaac
cccacttggt 300gacacaactc acacagagcc caaatcttgt gacacacctc
ccccgtgccc aaggtgccca 360ggcggtggaa gcagcggagg tggaagtgga
ggacagcccc gagaaccaca ggtgtacacc 420ctgcccccat cccgggagga
gatgaccaag aaccaggtca gcctgacctg cctggtcaaa 480ggcttctacc
ccagcgacat cgccgtggag tgggagagca gcgggcagcc ggagaacaac
540tacaacacca cgcctcccat gctggactcc gacggctcct tcttcctcta
cagcaagctc 600accgtggaca agagcaggtg gcagcagggg aacatcttct
catgctccgt gatgcatgag 660gctctgcaca accgcttcac gcagaagagc
ctctccctgt ctccgggtaa aggcctcggt 720ggcctgatgc atggagatac
acctacattg catgaatata tgttagattt gcaaccagag 780acaactgatc
tctacggata tggacaatta aatgacagct cagaggagga ggatgaaata
840gatggtccag ctggacaagc agaaccggac agagcccatt acaatattgt
aaccttttgt 900tgcaagtgtg actctacgct tcggttgtgc gtacaaagca
cacacgtaga cattcgtact 960ttggaagacc tgttaatggg cacactagga
attgtgtgcc ccatctgttc tcagaaacca 1020ggcggtggaa gcagcggagg
tggaagtgga atgtttcagg acccacagga gcgacccaga 1080aagttaccac
agttatgcac agagctgcaa acaactatac atgatataat attagaatgt
1140gtgtactgca agcaacagtt actgcgacgt gaggtatatg actttgcttt
tcgggattta 1200tgcatagtat atagagatgg gaatccatat gctgtacgag
ataaatgttt aaagttttat 1260tctaaaatta gtgagtatag acattattgt
tatagtttgt atggaacaac attagaacag 1320caatacaaca aaccgttgtg
tgatttgtta attaggtgta ttaaccgaca aaagccactg 1380tgtcctgaag
aaaagcaaag acatctggac aaaaagcaaa gattccataa tataaggggt
1440cggtggaccg gtcgatgtat gtcttgttgc agatcatcaa gaacacgtag
agaaacccag 1500ctgtaa 150621501PRTArtificialSynthetic 21Met Gln Val
Ser Thr Ala Ala Leu Ala Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu
Cys Asn Gln Val Leu Ser Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25
30 Ala Cys Cys Phe Ser Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile
35 40 45 Ala Asp Tyr Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser
Val Ile 50 55 60 Phe Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp
Pro Ser Glu Glu 65 70 75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu
Leu Ser Ala Glu Leu Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His
Thr Glu Pro Lys Ser Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg
Cys Pro Gly Gly Gly Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln
Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu
Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155
160 Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln
165 170 175 Pro Glu Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser
Asp Gly 180 185 190 Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys
Ser Arg Trp Gln 195 200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met
His Glu Ala Leu His Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser
Leu Ser Pro Gly Lys Gly Leu Gly 225 230 235 240 Gly Leu Met His Gly
Asp Thr Pro Thr Leu His Glu Tyr Met Leu Asp 245 250 255 Leu Gln Pro
Glu Thr Thr Asp Leu Tyr Gly Tyr Gly Gln Leu Asn Asp 260 265 270 Ser
Ser Glu Glu Glu Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu 275 280
285 Pro Asp Arg Ala His Tyr Asn Ile Val Thr Phe Cys Cys Lys Cys Asp
290 295 300 Ser Thr Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile
Arg Thr 305 310 315 320 Leu Glu Asp Leu Leu Met Gly Thr Leu Gly Ile
Val Cys Pro Ile Cys 325 330 335 Ser Gln Lys Pro Gly Gly Gly Ser Ser
Gly Gly Gly Ser Gly Met Phe 340 345 350 Gln Asp Pro Gln Glu Arg Pro
Arg Lys Leu Pro Gln Leu Cys Thr Glu 355 360 365 Leu Gln Thr Thr Ile
His Asp Ile Ile Leu Glu Cys Val Tyr Cys Lys 370 375 380 Gln Gln Leu
Leu Arg Arg Glu Val Tyr Asp Phe Ala Phe Arg Asp Leu 385 390 395 400
Cys Ile Val Tyr Arg Asp Gly Asn Pro Tyr Ala Val Arg Asp Lys Cys 405
410 415 Leu Lys Phe Tyr Ser Lys Ile Ser Glu Tyr Arg His Tyr Cys Tyr
Ser 420 425 430 Leu Tyr Gly Thr Thr Leu Glu Gln Gln Tyr Asn Lys Pro
Leu Cys Asp 435 440 445 Leu Leu Ile Arg Cys Ile Asn Arg Gln Lys Pro
Leu Cys Pro Glu Glu 450 455 460 Lys Gln Arg His Leu Asp Lys Lys Gln
Arg Phe His Asn Ile Arg Gly 465 470 475 480 Arg Trp Thr Gly Arg Cys
Met Ser Cys Cys Arg Ser Ser Arg Thr Arg 485 490 495 Arg Glu Thr Gln
Leu 500 22151PRTHuman papillomavirus 22Met Phe Gln Asp Pro Gln Glu
Arg Pro Arg Lys Leu Pro Gln Leu Cys 1 5 10 15 Thr Glu Leu Gln Thr
Thr Ile His Asp Ile Ile Leu Glu Cys Val Tyr 20 25 30 Cys Lys Gln
Gln Leu Leu Arg Arg Glu Val Tyr Asp Phe Ala Phe Arg 35 40 45 Asp
Leu Cys Ile Val Tyr Arg Asp Gly Asn Pro Tyr Ala Val Cys Asp 50 55
60 Lys Cys Leu Lys Phe Tyr Ser Lys Ile Ser Glu Tyr Arg His Tyr Cys
65 70 75 80 Tyr Ser Leu Tyr Gly Thr Thr Leu Glu Gln Gln Tyr Asn Lys
Pro Leu 85 90 95 Cys Asp Leu Leu Ile Arg Cys Ile Asn Cys Gln Lys
Pro Leu Cys Pro 100 105 110 Glu Glu Lys Gln Arg His Leu Asp Lys Lys
Gln Arg Phe His Asn Ile 115 120 125 Arg Gly Arg Trp Thr Gly Arg Cys
Met Ser Cys Cys Arg Ser Ser Arg 130 135 140 Thr Arg Arg Glu Thr Gln
Leu 145 150 2398PRTHuman papillomavirus 23Met His Gly Asp Thr Pro
Thr Leu His Glu Tyr Met Leu Asp Leu Gln 1 5 10 15 Pro Glu Thr Thr
Asp Leu Tyr Cys Tyr Glu Gln Leu Asn Asp Ser Ser 20 25 30 Glu Glu
Glu Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu Pro Asp 35 40 45
Arg Ala His Tyr Asn Ile Val Thr Phe Cys Cys Lys Cys Asp Ser Thr 50
55 60 Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile Arg Thr Leu
Glu 65 70 75 80 Asp Leu Leu Met Gly Thr Leu Gly Ile Val Cys Pro Ile
Cys Ser Gln 85 90 95 Lys Pro 24158PRTHuman papillomavirus 24Met Ala
Arg Phe Glu Asp Pro Thr Arg Arg Pro Tyr Lys Leu Pro Asp 1 5 10 15
Leu Cys Thr Glu Leu Asn Thr Ser Leu Gln Asp Ile Glu Ile Thr Cys 20
25 30 Val Tyr Cys Lys Thr Val Leu Glu Leu Thr Glu Val Phe Glu Phe
Ala 35 40 45 Phe Lys Asp Leu Phe Val Val Tyr Arg Asp Ser Ile Pro
His Ala Ala 50 55 60 Cys His Lys Cys Ile Asp Phe Tyr Ser Arg Ile
Arg Glu Leu Arg His 65 70 75 80 Tyr Ser Asp Ser Val Tyr Gly Asp Thr
Leu Glu Lys Leu Thr Asn Thr 85 90 95 Gly Leu Tyr Asn Leu Leu Ile
Arg Cys Leu Arg Cys Gln Lys Pro Leu 100 105 110 Asn Pro Ala Glu Lys
Leu Arg His Leu Asn Glu Lys Arg Arg Phe His 115 120 125 Asn Ile Ala
Gly His Tyr Arg Gly Gln Cys His Ser Cys Cys Asn Arg 130 135 140 Ala
Arg Gln Glu Arg Leu Gln Arg Arg Arg Glu Thr Gln Val 145 150 155
25105PRTHuman papillomavirus 25Met His Gly Pro Lys Ala Thr Leu Gln
Asp Ile Val Leu His Leu Glu 1 5 10 15 Pro Gln Asn Glu Ile Pro Val
Asp Leu Leu Cys His Glu Gln Leu Ser 20 25 30 Asp Ser Glu Glu Glu
Asn Asp Glu Ile Asp Gly Val Asn His Gln His 35 40 45 Leu Pro Ala
Arg Arg Ala Glu Pro Gln Arg His Thr Met Leu Cys Met 50 55 60 Cys
Cys Lys Cys Glu Ala Arg Ile Lys Leu Val Val Glu Ser Ser Ala 65 70
75 80 Asp Asp Leu Arg Ala Phe Gln Gln Leu Phe Leu Asn Thr Leu Ser
Phe 85 90 95 Val Cys Pro Trp Cys Ala Ser Gln Gln 100 105
2612PRTHomo sapiens 26Glu Leu Lys Thr Pro Leu Gly Asp Thr Thr His
Thr 1 5 10 2715PRTHomo sapiens 27Glu Pro Lys Ser Cys Asp Thr Pro
Pro Pro Cys Pro Arg Cys Pro 1 5 10 15 2810PRTArtificialSynthetic
28Gly Gly Gly Ser Ser Gly Gly Gly Ser Gly 1 5 10 29107PRTHomo
sapiens 29Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser
Arg Glu 1 5 10 15 Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu
Val Lys Gly Phe 20 25 30 Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu
Ser Ser Gly Gln Pro Glu 35 40 45 Asn Asn Tyr Asn Thr Thr Pro Pro
Met Leu Asp Ser Asp Gly Ser Phe 50 55 60 Phe Leu Tyr Ser Lys Leu
Thr Val Asp Lys Ser Arg Trp Gln Gln Gly 65 70 75 80 Asn Ile Phe Ser
Cys Ser Val Met His Glu Ala Leu His Asn Arg Phe 85 90 95 Thr Gln
Lys Ser Leu Ser Leu Ser Pro Gly Lys 100 105
305PRTArtificialSynthetic 30Gly Leu Gly Gly Leu 1 5
311506DNAArtificialSynthetic 31atgcaggtct ccactgctgc ccttgccgtc
ctcctctgca ccatggctct ctgcaaccag 60gtcctctctg caccacttgc tgctgacacg
ccgaccgcct gctgcttcag ctacacctcc 120cgacagattc cacagaattt
catagctgac tactttgaga cgagcagcca gtgctccaag 180cccagtgtca
tcttcctaac caagagaggc cggcaggtct gtgctgaccc cagtgaggag
240tgggtccaga aatacgtcag tgacctggag ctgagtgccg agctcaaaac
cccacttggt 300gacacaactc acacagagcc caaatcttgt gacacacctc
ccccgtgccc aaggtgccca 360ggcggtggaa gcagcggagg tggaagtgga
ggacagcccc gagaaccaca ggtgtacacc 420ctgcccccat cccgggagga
gatgaccaag aaccaggtca gcctgacctg cctggtcaaa 480ggcttctacc
ccagcgacat cgccgtggag tgggagagca gcgggcagcc ggagaacaac
540tacaacacca cgcctcccat gctggactcc gacggctcct tcttcctcta
cagcaagctc 600accgtggaca agagcaggtg gcagcagggg aacatcttct
catgctccgt gatgcatgag 660gctctgcaca accgcttcac gcagaagagc
ctctccctgt ctccgggtaa aggcctcggt 720ggcctgatgc atggcgatac
cccaacactc catgagtaca tgctggacct tcagcccgag 780actacggatc
tgtatggcta tgggcagttg aatgactcat ctgaggagga ggacgaaata
840gacggcccag ctggtcaagc cgaaccggat agagcccact acaacattgt
gaccttttgc 900tgtaagtgtg acagcactct gagactgtgt gttcagtcca
ctcatgtcga catacgcaca 960ttggaggatc tcctgatggg aacactggga
attgtgtgtc ccatctgttc ccaaaagcct 1020ggaggtggaa gcagtggagg
cggttcaggc atgttccaag atcctcaaga acgtcctcgt 1080aagctgccac
agctgtgtac cgagcttcag accaccattc acgacatcat cctggagtgc
1140gtctattgca aacagcagct ccttagaagg gaagtgtacg attttgcacg
gagggacctc 1200tgcatcgtgt atcgggacgg caatccctat gcggtacggg
ataaatgcct gaagttctac 1260agcaaaatct ccgagtaccg gcactactgc
tactctctct atgggacgac tctggaacag 1320cagtacaaca agcccttgtg
cgatctgctg attcgctgca ttaatcgcca gaaacctctg 1380tgcccagaag
agaagcaaag acacctggac
aagaaacagc gattccacaa catccgaggg 1440agatggacag ggaggtgtat
gagctgctgt cggagttcta ggacaaggcg cgaaacccag 1500ctttga
150632501PRTArtificialSynthetic 32Met Gln Val Ser Thr Ala Ala Leu
Ala Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu Cys Asn Gln Val Leu
Ser Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25 30 Ala Cys Cys Phe
Ser Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile 35 40 45 Ala Asp
Tyr Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser Val Ile 50 55 60
Phe Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp Pro Ser Glu Glu 65
70 75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu Leu Ser Ala Glu
Leu Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His Thr Glu Pro Lys
Ser Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg Cys Pro Gly Gly
Gly Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln Pro Arg Glu Pro
Gln Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu Glu Met Thr Lys
Asn Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155 160 Gly Phe Tyr
Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln 165 170 175 Pro
Glu Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser Asp Gly 180 185
190 Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln
195 200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met His Glu Ala Leu
His Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly
Lys Gly Leu Gly 225 230 235 240 Gly Leu Met His Gly Asp Thr Pro Thr
Leu His Glu Tyr Met Leu Asp 245 250 255 Leu Gln Pro Glu Thr Thr Asp
Leu Tyr Gly Tyr Gly Gln Leu Asn Asp 260 265 270 Ser Ser Glu Glu Glu
Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu 275 280 285 Pro Asp Arg
Ala His Tyr Asn Ile Val Thr Phe Cys Cys Lys Cys Asp 290 295 300 Ser
Thr Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile Arg Thr 305 310
315 320 Leu Glu Asp Leu Leu Met Gly Thr Leu Gly Ile Val Cys Pro Ile
Cys 325 330 335 Ser Gln Lys Pro Gly Gly Gly Ser Ser Gly Gly Gly Ser
Gly Met Phe 340 345 350 Gln Asp Pro Gln Glu Arg Pro Arg Lys Leu Pro
Gln Leu Cys Thr Glu 355 360 365 Leu Gln Thr Thr Ile His Asp Ile Ile
Leu Glu Cys Val Tyr Cys Lys 370 375 380 Gln Gln Leu Leu Arg Arg Glu
Val Tyr Asp Phe Ala Arg Arg Asp Leu 385 390 395 400 Cys Ile Val Tyr
Arg Asp Gly Asn Pro Tyr Ala Val Arg Asp Lys Cys 405 410 415 Leu Lys
Phe Tyr Ser Lys Ile Ser Glu Tyr Arg His Tyr Cys Tyr Ser 420 425 430
Leu Tyr Gly Thr Thr Leu Glu Gln Gln Tyr Asn Lys Pro Leu Cys Asp 435
440 445 Leu Leu Ile Arg Cys Ile Asn Arg Gln Lys Pro Leu Cys Pro Glu
Glu 450 455 460 Lys Gln Arg His Leu Asp Lys Lys Gln Arg Phe His Asn
Ile Arg Gly 465 470 475 480 Arg Trp Thr Gly Arg Cys Met Ser Cys Cys
Arg Ser Ser Arg Thr Arg 485 490 495 Arg Glu Thr Gln Leu 500
331506DNAArtificialSynthetic 33atgcaggtct ccactgctgc ccttgccgtc
ctcctctgca ccatggctct ctgcaaccag 60gtcctctctg caccacttgc tgctgacacg
ccgaccgcct gctgcttcag ctacacctcc 120cgacagattc cacagaattt
catagctgac tactttgaga cgagcagcca gtgctccaag 180cccagtgtca
tcttcctaac caagagaggc cggcaggtct gtgctgaccc cagtgaggag
240tgggtccaga aatacgtcag tgacctggag ctgagtgccg agctcaaaac
cccacttggt 300gacacaactc acacagagcc caaatcttgt gacacacctc
ccccgtgccc aaggtgccca 360ggcggtggaa gcagcggagg tggaagtgga
ggacagcccc gagaaccaca ggtgtacacc 420ctgcccccat cccgggagga
gatgaccaag aaccaggtca gcctgacctg cctggtcaaa 480ggcttctacc
ccagcgacat cgccgtggag tgggagagca gcgggcagcc ggagaacaac
540tacaacacca cgcctcccat gctggactcc gacggctcct tcttcctcta
cagcaagctc 600accgtggaca agagcaggtg gcagcagggg aacatcttct
catgctccgt gatgcatgag 660gctctgcaca accgcttcac gcagaagagc
ctctccctgt ctccgggtaa aggcctcggt 720ggcctgatgc atggtgacac
accaaccctg cacgaataca tgctcgatct gcagccagag 780actaccgacc
tttacggcta tgggcagttg aacgacagct ctgaggagga ggacgagatc
840gatggtcctg ctggacaagc agaaccagac agagcccact acaacatcgt
aaccttttgc 900tgcaagtgtg acagtaccct tcgtttgtgc gttcagagca
cgcatgtcga cattcggaca 960ctggaggatc tgctcatggg gactctgggg
attgtgtgtc ctatttgcag ccagaaacca 1020ggcggaggat cttcaggagg
cgggagtggc atgttccaag accctcagga acgccctcgg 1080aaactgcccc
aattgtgtac tgagctccag acaacgatac acgacataat cctggagtgc
1140gtgtattgca agcagcagct tctgaggagg gaagtgtacg attttgccag
gagagatggc 1200tgcattgtct accgagatgg caatccctat gcggtgtgtg
ataagtgtct gaagttctat 1260tccaaaatca gcgaatatcg gcattattgc
tactcactgt acggaactac cctcgaacag 1320cagtacaaca aaccgctctg
tgatctgctg atcagatgca tcaatcggca gaaacccctt 1380tgtcccgaag
agaagcaaag acacctggac aagaagcaga ggttccacaa tacccgaggt
1440cgttggactg ggcgctgcat gtcctgttgt cgctcctctc gcacaaggag
agagacacaa 1500ctgtga 150634501PRTArtificialSynthetic 34Met Gln Val
Ser Thr Ala Ala Leu Ala Val Leu Leu Cys Thr Met Ala 1 5 10 15 Leu
Cys Asn Gln Val Leu Ser Ala Pro Leu Ala Ala Asp Thr Pro Thr 20 25
30 Ala Cys Cys Phe Ser Tyr Thr Ser Arg Gln Ile Pro Gln Asn Phe Ile
35 40 45 Ala Asp Tyr Phe Glu Thr Ser Ser Gln Cys Ser Lys Pro Ser
Val Ile 50 55 60 Phe Leu Thr Lys Arg Gly Arg Gln Val Cys Ala Asp
Pro Ser Glu Glu 65 70 75 80 Trp Val Gln Lys Tyr Val Ser Asp Leu Glu
Leu Ser Ala Glu Leu Lys 85 90 95 Thr Pro Leu Gly Asp Thr Thr His
Thr Glu Pro Lys Ser Cys Asp Thr 100 105 110 Pro Pro Pro Cys Pro Arg
Cys Pro Gly Gly Gly Ser Ser Gly Gly Gly 115 120 125 Ser Gly Gly Gln
Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser 130 135 140 Arg Glu
Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys 145 150 155
160 Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Ser Gly Gln
165 170 175 Pro Glu Asn Asn Tyr Asn Thr Thr Pro Pro Met Leu Asp Ser
Asp Gly 180 185 190 Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys
Ser Arg Trp Gln 195 200 205 Gln Gly Asn Ile Phe Ser Cys Ser Val Met
His Glu Ala Leu His Asn 210 215 220 Arg Phe Thr Gln Lys Ser Leu Ser
Leu Ser Pro Gly Lys Gly Leu Gly 225 230 235 240 Gly Leu Met His Gly
Asp Thr Pro Thr Leu His Glu Tyr Met Leu Asp 245 250 255 Leu Gln Pro
Glu Thr Thr Asp Leu Tyr Gly Tyr Gly Gln Leu Asn Asp 260 265 270 Ser
Ser Glu Glu Glu Asp Glu Ile Asp Gly Pro Ala Gly Gln Ala Glu 275 280
285 Pro Asp Arg Ala His Tyr Asn Ile Val Thr Phe Cys Cys Lys Cys Asp
290 295 300 Ser Thr Leu Arg Leu Cys Val Gln Ser Thr His Val Asp Ile
Arg Thr 305 310 315 320 Leu Glu Asp Leu Leu Met Gly Thr Leu Gly Ile
Val Cys Pro Ile Cys 325 330 335 Ser Gln Lys Pro Gly Gly Gly Ser Ser
Gly Gly Gly Ser Gly Met Phe 340 345 350 Gln Asp Pro Gln Glu Arg Pro
Arg Lys Leu Pro Gln Leu Cys Thr Glu 355 360 365 Leu Gln Thr Thr Ile
His Asp Ile Ile Leu Glu Cys Val Tyr Cys Lys 370 375 380 Gln Gln Leu
Leu Arg Arg Glu Val Tyr Asp Phe Ala Arg Arg Asp Gly 385 390 395 400
Cys Ile Val Tyr Arg Asp Gly Asn Pro Tyr Ala Val Cys Asp Lys Cys 405
410 415 Leu Lys Phe Tyr Ser Lys Ile Ser Glu Tyr Arg His Tyr Cys Tyr
Ser 420 425 430 Leu Tyr Gly Thr Thr Leu Glu Gln Gln Tyr Asn Lys Pro
Leu Cys Asp 435 440 445 Leu Leu Ile Arg Cys Ile Asn Arg Gln Lys Pro
Leu Cys Pro Glu Glu 450 455 460 Lys Gln Arg His Leu Asp Lys Lys Gln
Arg Phe His Asn Thr Arg Gly 465 470 475 480 Arg Trp Thr Gly Arg Cys
Met Ser Cys Cys Arg Ser Ser Arg Thr Arg 485 490 495 Arg Glu Thr Gln
Leu 500
* * * * *
References