U.S. patent application number 17/230353 was filed with the patent office on 2021-08-26 for cartilage-homing peptide conjugates and methods of use thereof.
The applicant listed for this patent is Blaze Bioscience, Inc.. Invention is credited to Claudia Jochheim, Dennis M. Miller, Natalie Winblade Nairn, Julia E. Novak, Scott Presnell, Mark Stroud.
Application Number | 20210260205 17/230353 |
Document ID | / |
Family ID | 1000005553327 |
Filed Date | 2021-08-26 |
United States Patent
Application |
20210260205 |
Kind Code |
A1 |
Novak; Julia E. ; et
al. |
August 26, 2021 |
CARTILAGE-HOMING PEPTIDE CONJUGATES AND METHODS OF USE THEREOF
Abstract
Peptides that home, target, migrate to, are directed to, are
retained by, or accumulate in and/or bind to the cartilage or
kidney of a subject are disclosed. Pharmaceutical compositions and
uses for peptides or peptide-active agent complexes comprising such
peptides are also disclosed. Such compositions can be formulated
for targeted delivery of an active agent to a target region,
tissue, structure or cell in the cartilage. Targeted compositions
of the disclosure can deliver peptide or peptide-active agent
complexes to target regions, tissues, structures, or cells targeted
by the peptide.
Inventors: |
Novak; Julia E.; (Sequim,
WA) ; Nairn; Natalie Winblade; (Seattle, WA) ;
Miller; Dennis M.; (Woodinville, WA) ; Presnell;
Scott; (Tacoma, WA) ; Jochheim; Claudia;
(Seattle, WA) ; Stroud; Mark; (Seattle,
WA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Blaze Bioscience, Inc. |
Seattle |
WA |
US |
|
|
Family ID: |
1000005553327 |
Appl. No.: |
17/230353 |
Filed: |
April 14, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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16492914 |
Sep 10, 2019 |
11013814 |
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PCT/US2018/023006 |
Mar 16, 2018 |
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17230353 |
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62472485 |
Mar 16, 2017 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 49/0058 20130101;
A61K 47/65 20170801; A61K 51/08 20130101; C07K 2319/33 20130101;
C07K 14/43518 20130101; A61B 6/4057 20130101; A61B 5/055 20130101;
A61K 47/6415 20170801; A61P 13/12 20180101; A61B 6/037 20130101;
A61P 19/02 20180101; A61B 8/481 20130101; C07K 2319/50 20130101;
A61B 6/481 20130101; C07K 16/244 20130101; C07K 2319/30 20130101;
C07K 16/248 20130101; A61K 49/0056 20130101; C07K 14/43522
20130101; A61B 5/0071 20130101; A61B 2505/05 20130101; C07K 14/4726
20130101 |
International
Class: |
A61K 47/64 20060101
A61K047/64; A61K 47/65 20060101 A61K047/65; A61P 19/02 20060101
A61P019/02; A61P 13/12 20060101 A61P013/12; A61B 5/00 20060101
A61B005/00; A61B 6/03 20060101 A61B006/03; A61B 6/00 20060101
A61B006/00; A61K 49/00 20060101 A61K049/00; A61K 51/08 20060101
A61K051/08; C07K 14/435 20060101 C07K014/435; C07K 14/47 20060101
C07K014/47; C07K 16/24 20060101 C07K016/24 |
Claims
1-115. (canceled)
116. A peptide active agent conjugate, the peptide active agent
conjugate comprising: a) a peptide comprising a sequence that has
at least 85% sequence identity to SEQ ID NO: 396 or a fragment
thereof comprising at least 18 amino acid residues that has at
least 85% sequence identity to SEQ ID NO: 396; and b) an active
agent, wherein the active agent is desciclesonide, kineret, a j
anus kinase (JAK) inhibitor, tofacitinib, dasatinib, flunisolide,
an interleukin-1 (IL-1) inhibitor, an interleukin-1 (IL-1) receptor
antagonist, an interleukin-12 (IL-12) antagonist, fibroblast growth
factor 18 (FGF-18), insulin-like growth factor 1 (IGF-1),
transforming growth factor beta (TFG- ), a cathepsin K inhibitor,
an NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3)
inflammasome drug, or MCC950.
117. The peptide active agent conjugate of claim 116, wherein the
peptide comprises 4 or more cysteine residues.
118. The peptide active agent conjugate of claim 116, wherein the
peptide or the fragment thereof comprises three or more disulfide
bridges formed between cysteine residues, wherein one of the
disulfide bridges passes through a loop formed by two other
disulfide bridges.
119. The peptide active agent conjugate of claim 116, wherein the
IL-1 receptor antagonist is canakinumab, rilonacept, or
gevokizumab.
120. The peptide active agent conjugate of claim 116, wherein the
peptide or the fragment thereof comprises 5 to 12 basic
residues.
121. The peptide active agent conjugate of claim 116, wherein the
peptide is SEQ ID NO: 396.
122. The peptide active agent conjugate of claim 116, wherein 1, 2,
3, 4, 5, 6, 7, 8, 9, or 10 active agents are linked to the
peptide.
123. The peptide active agent conjugate of claim 116, wherein the
peptide is linked to the active agent by fusion of the active agent
to the peptide at an N-terminus or a C-terminus of the peptide.
124. The peptide active agent conjugate of claim 116, wherein the
peptide is linked to the active agent at an N-terminus, at the
epsilon amine of an internal lysine residue, at the carboxylic acid
of an aspartic acid or glutamic acid residue, or a C-terminus of
the peptide by a linker.
125. The peptide active agent conjugate of claim 116, wherein the
peptide is linked to the active agent at a non-natural amino acid
present in the peptide, wherein the non-natural amino acid is an
insertion, appendage, or substitution for another amino acid.
126. The peptide active agent conjugate of claim 1, wherein the
peptide is linked to the active agent by a cleavable linker
comprising a cleavage site for matrix metalloproteinases, thrombin,
cathepsins, or beta-glucuronidase.
127. The peptide active agent conjugate of claim 116, wherein the
peptide is linked to the active agent by a linker comprising an
amide bond, an ester bond, a carbamate bond, a carbonate bond, a
hydrazone bond, an oxime bond, a disulfide bond, a thioester bond,
a thioether bond, a triazole, a carbon-carbon bond, or a
carbon-nitrogen bond.
128. The peptide active agent conjugate of claim 116, wherein the
peptide is linked to the active agent by a hydrolytically labile
linker.
129. The peptide active agent conjugate of claim 116, wherein the
peptide is linked to the active agent by a pH sensitive, reducible,
glutathione-sensitive, or protease cleavable linker.
130. The peptide active agent conjugate of claim 116, wherein the
peptide is linked to the active agent by a stable linker.
131. A pharmaceutical composition comprising the peptide active
agent conjugate of claim 116, or a salt thereof, and a
pharmaceutically acceptable carrier.
132. A method of imaging an organ or body region of a subject
comprising administering to the subject a peptide active agent
conjugate comprising: a) a peptide comprising a sequence that has
at least 85% sequence identity to SEQ ID NO: 396 or a fragment
thereof comprising at least 18 amino acid residues that has at
least 85% sequence identity to SEQ ID NO: 396; b) an active agent,
wherein the active agent is desciclesonide, kineret, a j anus
kinase (JAK) inhibitor, tofacitinib, dasatinib, flunisolide, an
interleukin-1 (IL-1) inhibitor, an interleukin-1 (IL-1) receptor
antagonist, an interleukin-12 (IL-12) antagonist, fibroblast growth
factor 18 (FGF-18), insulin-like growth factor 1 (IGF-1),
transforming growth factor beta (TFG- ), a cathepsin K inhibitor,
an NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3)
inflammasome drug, or MCC950; and c) a detectable agent.
133. The method of claim 132, wherein the detectable agent is a
fluorophore, a near-infrared dye, a contrast agent, a nanoparticle,
a metal-containing nanoparticle, a metal chelate, an X-ray contrast
agent, a PET agent, a radioisotope, or a radionuclide chelator.
134. The method of claim 132, further comprising detecting a cancer
or diseased region, tissue, structure, or cell.
135. The method of claim 132, further comprising performing surgery
on the subject and removing the cancer or diseased region, the
tissue, the structure, or the cell.
136. A method of treating a cartilage or kidney disorder in a
subject comprising administering to the subject a peptide active
agent conjugate comprising: a) a peptide comprising a sequence that
has at least 85% sequence identity to SEQ ID NO: 396 or a fragment
thereof comprising at least 18 amino acid residues that has at
least 85% sequence identity to SEQ ID NO: 396; and b) an active
agent, wherein the active agent is desciclesonide, kineret, a j
anus kinase (JAK) inhibitor, tofacitinib, dasatinib, flunisolide,
an interleukin-1 (IL-1) inhibitor, an interleukin-1 (IL-1) receptor
antagonist, an interleukin-12 (IL-12) antagonist, fibroblast growth
factor 18 (FGF-18), insulin-like growth factor 1 (IGF-1),
transforming growth factor beta (TFG- ), a cathepsin K inhibitor,
an NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3)
inflammasome drug, or MCC950.
137. The method of claim 136, wherein the disorder is lupus
nephritis, rheumatoid arthritis, gout, ankylosing spondylitis,
psoriatic arthritis, osteoarthritis, fibrosis, scleroderma, chronic
kidney disease, diabetic nephropathy, renal fibrosis, acute kidney
injury, or calcium pyrophosphate deposition disease.
138. The method of claim 136, wherein upon administration to the
subject the peptide active agent conjugate homes to a cartilage or
a kidney of the subject.
139. The method of claim 136, wherein upon administration to the
subject, the peptide active agent conjugate homes to proximal
tubules of a kidney.
140. The method of claim 136, wherein the administering comprises
administering by inhalation, intranasally, orally, topically,
parenterally, intravenously, subcutaneously, intra-articularly,
intramuscularly administration, intraperitoneally, dermally,
transdermally, or a combination thereof.
Description
CROSS REFERENCE
[0001] This application is a continuation of U.S. application Ser.
No. 16/492,914, filed Sep. 10, 2019, which is a U.S. National Phase
Application under 35 U.S.C. .sctn. 371 of International Application
No. PCT/US2018/023006, filed Mar. 16, 2018, which claims the
benefit of U.S. Provisional Patent Application No. 62/472,485,
filed Mar. 16, 2017, the entire disclosures of which are
incorporated by reference.
SEQUENCE LISTING
[0002] The instant application contains a Sequence Listing which
has been submitted electronically in ASCII format and is hereby
incorporated by reference in its entirety. Said ASCII copy, created
on Mar. 15, 2018, is named 45639-711_601 SL.txt and is 302,571
bytes in size.
BACKGROUND
[0003] Cartilage comprises chondrocytes, a specialized cell-type
which produces components of the extracellular matrix, mainly
including collagen, proteoglycans (e.g., aggrecan), and elastic
fibers. The extracellular matrix proteins provide support, cushion,
and durability to cartilage-rich portions of the body such as
joints, ears, nose, and windpipe. Cartilage is one of few tissues
in the body which does not contain blood vessels and is considered
an avascular tissue. Unlike many cells in the body which rely on a
combination of blood flow and diffusion, chondrocytes rely on
diffusion. Because it does not have a direct blood supply, compared
to other connective tissues, cartilage grows and repairs much more
slowly. As a result, cartilage disorders are particularly difficult
to treat.
SUMMARY
[0004] The present disclosure relates to compositions and methods
for treatment of cartilage disorders. Described herein are peptides
that home to, migrate to, accumulate in, bind to, are retained by,
or are directed to, and/or bind in cartilage following
administration in a subject. In some embodiments, the homing
peptides of the present disclosure are used to deliver a detection
agent to image and/or diagnose cartilage, injury, or disease. In
some embodiments, compositions and methods for treatment of kidney
disorders are described. In other embodiments, the homing peptides
of the present disclosure are used to treat or deliver an active
agent to a region, tissue, structure, or cell thereof.
[0005] In some aspects, a peptide active agent conjugate comprises:
a) a peptide, wherein the peptide comprises a sequence that has at
least 70% sequence identity with any one of SEQ ID NO: 24-SEQ ID
NO: 274 and upon administration to a subject the peptide homes,
targets, migrates to, accumulates in, binds to, is retained by, or
is directed to a cartilage of the subject, and an active agent
selected from an active agent class selected from TABLE 3 or TABLE
5; b) a peptide, wherein the peptide comprises a sequence that has
at least 70% sequence identity with any one of SEQ ID NO: 24-SEQ ID
NO: 274 and upon administration to a subject the peptide homes,
targets, migrates to, accumulates in, binds to, is retained by, or
is directed to a kidney of the subject, and an active agent
selected from an active agent class selected from TABLE 4 or TABLE
5; c) a peptide, wherein the peptide comprises a sequence that has
at least 70% sequence identity with any one of SEQ ID NO: 24-SEQ ID
NO: 274 and upon administration to a subject the peptide homes,
targets, migrates to, accumulates in, binds to, is retained by, or
is directed to a cartilage or kidney of the subject, and an active
agent selected from TABLE 3, TABLE 4, or TABLE 5; d) a peptide,
wherein the peptide comprises a sequence that has at least 70%
sequence identity with any one of SEQ ID NO: 314-SEQ ID NO: 564 and
upon administration to a subject the peptide homes, targets,
migrates to, accumulates in, binds to, is retained by, or is
directed to a cartilage of the subject, and an active agent
selected from TABLE 3 or TABLE 5; e) a peptide, wherein the peptide
comprises a sequence that has at least 70% sequence identity with
any one of SEQ ID NO: 314-SEQ ID NO: 564 and upon administration to
a subject the peptide homes, targets, migrates to, accumulates in,
binds to, is retained by, or is directed to a kidney of the
subject, and an active agent selected from an active agent class
selected from TABLE 4 or TABLE 5; f) a peptide, wherein the peptide
comprises a sequence that has at least 70% sequence identity with
any one of SEQ ID NO: 314-SEQ ID NO: 564 and upon administration to
a subject the peptide homes, targets, migrates to, accumulates in,
binds to, is retained by, or is directed to a cartilage or kidney
of the subject, and an active agent selected from an active agent
class selected from TABLE 3, TABLE 4, or TABLE 5; g) a peptide,
wherein the peptide comprises a sequence that has at least 70%
sequence identity with any one of SEQ ID NO: 260-SEQ ID NO: 274 and
upon administration to a subject the peptide homes, targets,
migrates to, accumulates in, binds to, is retained by, or is
directed to a cartilage of the subject, and an active agent
selected from TABLE 3, TABLE 5, or TABLE 6; h) a peptide, wherein
the peptide comprises a sequence that has at least 70% sequence
identity with any one of SEQ ID NO: 260-SEQ ID NO: 274 and upon
administration to a subject the peptide homes, targets, migrates
to, accumulates in, binds to, is retained by, or is directed to a
kidney of the subject, and an active agent selected from TABLE 4,
TABLE 5, or TABLE 6; i) a peptide, wherein the peptide comprises a
sequence that has at least 70% sequence identity with any one of
SEQ ID NO: 260-SEQ ID NO: 274 and upon administration to a subject
the peptide homes, targets, migrates to, accumulates in, binds to,
is retained by, or is directed to a cartilage or a kidney of the
subject, and an active agent selected from TABLE 3, TABLE 4, TABLE
5, or TABLE 6; j) a peptide, wherein the peptide comprises a
sequence that has at least 70% sequence identity with any one of
SEQ ID NO: 550-SEQ ID NO: 564 and upon administration to a subject
the peptide homes, targets, migrates to, accumulates in, binds to,
is retained by, or is directed to a cartilage of the subject, and
an active agent selected from TABLE 3, TABLE 5, or TABLE 6; k) a
peptide, wherein the peptide comprises a sequence that has at least
70% sequence identity with any one of SEQ ID NO: 550-SEQ ID NO: 564
and upon administration to a subject the peptide homes, targets,
migrates to, accumulates in, binds to, is retained by, or is
directed to a kidney of the subject, and an active agent selected
from TABLE 4, TABLE 5, or TABLE 6; or 1) a peptide, wherein the
peptide comprises a sequence that has at least 70% sequence
identity with any one of SEQ ID NO: 550-SEQ ID NO: 564 and upon
administration to a subject the peptide homes, targets, migrates
to, accumulates in, binds to, is retained by, or is directed to a
cartilage or a kidney of the subject, and an active agent selected
from TABLE 3, TABLE 4, TABLE 5, or TABLE 6. In some embodiments,
the peptide comprises: a) a sequence that has at least 75%, at
least 80%, at least 85%, at least 90%, at least 95%, at least 97%,
at least 99% or 100% sequence identity with any one of SEQ ID NO:
24-SEQ ID NO: 274 or a fragment thereof, b) a sequence that has at
least 75%, at least 80%, at least 85%, at least 90%, at least 95%,
at least 97%, at least 99% or 100% sequence identity with any one
of SEQ ID NO: 260-SEQ ID NO: 274 or a fragment thereof; c) a
sequence that has at least 75%, at least 80%, at least 85%, at
least 90%, at least 95%, at least 97%, at least 99% or 100%
sequence identity with any one of SEQ ID NO: 314-SEQ ID NO: 564 or
a fragment thereof; or d) a sequence that has at least 75%, at
least 80%, at least 85%, at least 90%, at least 95%, at least 97%,
at least 99% or 100% sequence identity with any one of SEQ ID NO:
550-SEQ ID NO: 564 or a fragment thereof. In some embodiments, the
peptide comprises: a) a sequence of any one of SEQ ID NO: 24-SEQ ID
NO: 274 or a fragment thereof, b) a sequence of any one of SEQ ID
NO: 260-SEQ ID NO: 274 or a fragment thereof, c) a sequence of any
one of SEQ ID NO: 314-SEQ ID NO: 564 or a fragment thereof, or d) a
sequence of any one of SEQ ID NO: 550-SEQ ID NO: 564 or a fragment
thereof.
[0006] In some aspects, a peptide comprises a sequence that has at
least 70%, at least 75%, at least 80%, at least 85%, at least 90%,
at least 95%, at least 97%, at least 99%, or 100% sequence identity
with any one of SEQ ID NO: 260-SEQ ID NO: 574 or at least 70%, at
least 75%, at least 80%, at least 85%, at least 90%, at least 95%,
at least 97%, at least 99%, or 100% sequence identity with any one
of SEQ ID NO: 550-SEQ ID NO: 564.
[0007] In some embodiments, the peptide comprises: a) a sequence of
any one of SEQ ID NO: 1-SEQ ID NO: 23 or a fragment thereof, b) a
sequence of any one of SEQ ID NO: 275-SEQ ID NO: 297 or a fragment
thereof; c) a sequence of any one of SEQ ID NO: 21-SEQ ID NO: 23 or
a fragment thereof, or d) a sequence of any one of SEQ ID NO:
295-SEQ ID NO: 297 or a fragment thereof. In some embodiments, the
peptide is at least 30%, at least 40%, at least 50%, at least 60%,
at least 70%, at least 80%, at least 90%, at least 95%, at least,
97%, at least 98%, or at least 99% identical to any one of SEQ ID
NO: 494-SEQ ID NO: 540 or at least 30%, at least 40%, at least 50%,
at least 60%, at least 70%, at least 80%, at least 90%, at least
95%, at least 97%, at least 98%, or at least 99% identical to any
one of SEQ ID NO: 204-SEQ ID NO: 250. In some embodiments, the
peptide is at least 70%, at least 80%, at least 90%, at least 95%,
at least 97%, or 100% identical to: a) SEQ ID NO: 111; b) SEQ ID
NO: 401; c) SEQ ID NO: 24; d) SEQ ID NO: 314; e) SEQ ID NO: 27; f)
SEQ ID NO: 317; g) SEQ ID NO: 185; h) SEQ ID NO: 475; i) SEQ ID NO:
30; j) SEQ ID NO: 320; k) SEQ ID NO: 108; 1) SEQ ID NO: 398; m) SEQ
ID NO: 36; n) SEQ ID NO: 326; o) SEQ ID NO: 199; p) SEQ ID NO: 478;
q) SEQ ID NO: 25; r) SEQ ID NO: 315; s) SEQ ID NO: 106; t) SEQ ID
NO: 396; u) SEQ ID NO: 26; v) SEQ ID NO: 316; w) SEQ ID NO: 187; x)
SEQ ID NO: 477; y) SEQ ID NO: 107; or z) SEQ ID NO: 397. In some
embodiments, the peptide is at least 70%, at least 80%, at least
90%, at least 95%, at least 97%, at least 99%, or 100% identical
to: a) SEQ ID NO: 550; b) SEQ ID NO: 551; c) SEQ ID NO: 552; d) SEQ
ID NO: 553; e) SEQ ID NO: 554; f) SEQ ID NO: 555; g) SEQ ID NO:
556; h) SEQ ID NO: 557; i) SEQ ID NO: 558; j) SEQ ID NO: 559; k)
SEQ ID NO: 560; 1) SEQ ID NO: 561; m) SEQ ID NO: 562; n) SEQ ID NO:
563; o) SEQ ID NO: 564; p) SEQ ID NO: 260; q) SEQ ID NO: 261; r)
SEQ ID NO: 262; s) SEQ ID NO: 263; t) SEQ ID NO: 264; u) SEQ ID NO:
265; v) SEQ ID NO: 266; w) SEQ ID NO: 267; x) SEQ ID NO: 268; y)
SEQ ID NO: 269; z) SEQ ID NO: 270; aa) SEQ ID NO: 271; bb) SEQ ID
NO: 272; cc) SEQ ID NO: 273; or dd) SEQ ID NO: 274. In some
embodiments, the peptide homes, targets, migrates to, accumulates
in, binds to, is retained by, or is directed to cartilage, to
kidney, or to cartilage and kidney. In some embodiments, the
peptide homes, targets, migrates to, accumulates in, binds to, is
retained by, or is directed to proximal tubules of the kidney. In
some embodiments, the peptide is covalently conjugated to the
active agent. In some embodiments, the peptide active agent
conjugate homes, targets, migrates to, accumulates in, binds to, is
retained by, or is directed to a cartilage or a kidney of the
subject.
[0008] In some embodiments, the peptide comprises 4 or more
cysteine residues. In some embodiments, the peptide comprises three
or more disulfide bridges formed between cysteine residues, wherein
one of the disulfide bridges passes through a loop formed by two
other disulfide bridges. In some embodiments, the peptide comprises
a plurality of disulfide bridges formed between cysteine residues.
In some embodiments, the peptide comprises a disulfide through a
disulfide knot. In some embodiments, at least one amino acid
residue of the peptide is in an L configuration or, wherein at
least one amino acid residue of the peptide is in a D
configuration.
[0009] In some embodiments, the sequence comprises at least 11, at
least 12, at least 13, at least 14, at least 15, at least 16, at
least 17, at least 18, at least 19, at least 20, at least 21, at
least 22, at least 23, at least 24, at least 25, at least 26, at
least 27, at least 28, at least 29, at least 30, at least 31, at
least 32, at least 33, at least 34, at least 35, at least 36, at
least 37, at least 38, at least 39, at least 40, at least 41, at
least 42, at least 43, at least 44, at least 45, at least 46, at
least 47, at least 48, at least 49, at least 50, at least 51, at
least 52, at least 53, at least 54, at least 55, at least 56, at
least 57, at least 58 residues, at least 59, at least 60, at least
61, at least 62, at least 63, at least 64, at least 65, at least
66, at least 67, at least 68, at least 69, at least 70, at least
71, at least 72, at least 73, at least 74, at least 75, at least
76, at least 77, at least 78, at least 79, at least 80, or at least
81 residues.
[0010] In some embodiments, any one or more K residues are replaced
by an R residue or wherein any one or more R residues are replaced
by for a K residue In some embodiments, any one or more M residues
are replaced by any one of the I, L, or V residues. In some
embodiments, any one or more L residues are replaced by any one of
the V, I, or M residues. In some embodiments, any one or more I
residues are replaced by any of the M, L, or V residues. In some
embodiments, any one or more V residues are replaced by any of the
M, I, or L residues. In some embodiments, any one or more G
residues are replaced by an A residue or wherein any one or more A
residues are replaced by a G residue. In some embodiments, any one
or more S residues are replaced by a T residue or wherein any one
or more T residues are replaced by for an S residue. In some
embodiments, any one or more Q residues are replaced by an N
residue or wherein any one or more N residues are replaced by a Q
residue. In some embodiments, any one or more D residues are
replaced by an E residue or wherein any one or more E residues are
replaced by a D residue.
[0011] In some embodiments, the peptide has a charge distribution
comprising an acidic region and a basic region. In some
embodiments, the acidic region is a nub. In some embodiments, the
basic region is a patch. In some embodiments, the peptide comprises
5-12 basic residues. In some embodiments, the peptide comprises 0-5
acidic residues. In some embodiments, the peptide comprises 6 or
more basic residues and 2 or fewer acidic residues. In some
embodiments, the peptide comprises a 4-19 amino acid residue
fragment containing at least 2 cysteine residues, and at least 2
positively charged amino acid residues. In some embodiments, the
peptide comprises a 20-70 amino acid residue fragment containing at
least 2 cysteine residues, no more than 2 basic residues and at
least 2 positively charged amino acid residues. In some
embodiments, the peptide comprises at least 3 positively charged
amino acid residues. In some embodiments, the positively charged
amino acid residues are selected from K, R, or a combination
thereof.
[0012] In some embodiments, the peptide has a charge greater than 2
at physiological pH. In some embodiments, the peptide has a charge
greater than 3.5 at physiological pH. In some embodiments, the
peptide has a charge greater than 4.5 at physiological pH. In some
embodiments, the peptide has a charge greater than 5.5 at
physiological pH. In some embodiments, the peptide has a charge
greater than 6.5 at physiological pH. In some embodiments, the
peptide has a charge greater than 7.5 at physiological pH. In some
embodiments, the peptide has a charge greater than 8.5 at
physiological pH. In some embodiments, the peptide has a charge
greater than 9.5 at physiological pH.
[0013] In some embodiments, the peptide is selected from a
potassium channel agonist, a potassium channel antagonist, a
portion of a potassium channel, a sodium channel agonist, a sodium
channel antagonist, a calcium channel agonist, a calcium channel
antagonist, a hadrucalcin, a theraphotoxin, a huwentoxin, a
kaliotoxin, a cobatoxin, or a lectin. In some embodiments, the
lectin is SHL-Ib2.
[0014] In some embodiments, the peptide is arranged in a multimeric
structure with at least one other peptide.
[0015] In some embodiments, at least one residue of the peptide
comprises a chemical modification. In some embodiments, the
chemical modification is blocking the N-terminus of the peptide. In
some embodiments, wherein the chemical modification is methylation,
acetylation, or acylation. In some embodiments, the chemical
modification is: methylation of one or more lysine residues or
analogue thereof methylation of the N-terminus; or methylation of
one or more lysine residue or analogue thereof and methylation of
the N-terminus. In some embodiments, the peptide is linked to an
acyl adduct.
[0016] In some embodiments, the peptide is linked to an active
agent. In some embodiments, the active agent is fused with the
peptide at an N-terminus or a C-terminus of the peptide. In some
embodiments, the active agent is another peptide. In some
embodiments, the active agent is an antibody. In some embodiments,
the active agent is an Fc domain, Fab domain, scFv, or Fv fragment.
In some embodiments, the peptide fused with an Fc domain comprises
a contiguous sequence. In some embodiments, 1, 2, 3, 4, 5, 6, 7, 8,
9, or 10 active agents are linked to the peptide. In some
embodiments, the peptide is linked to the active agent at an
N-terminus, at the epsilon amine of an internal lysine residue, at
the carboxylic acid of an aspartic acid or glutamic acid residue,
or a C-terminus of the peptide by a linker. In some embodiments,
the peptide is linked to the active agent via a cleavable linker.
In some embodiments, the peptide or peptide active agent conjugate
further comprises a non-natural amino acid, wherein the non-natural
amino acid is an insertion, appendage, or substitution for another
amino acid.
[0017] In some embodiments, the peptide is linked to the active
agent at the non-natural amino acid by a linker. In some
embodiments, the linker comprises an amide bond, an ester bond, a
carbamate bond, a carbonate bond, a hydrazone bond, an oxime bond,
a disulfide bond, a thioester bond, a thioether bond, a triazole, a
carbon-carbon bond, or a carbon-nitrogen bond. In some embodiments,
the cleavable linker comprises a cleavage site for matrix
metalloproteinases, thrombin, cathepsins, or beta-glucuronidase. In
some embodiments, the linker is a hydrolytically labile linker. In
some embodiments, the linker is pH sensitive, reducible,
glutathione-sensitive, or protease cleavable. In some embodiments,
the peptide is linked to the active agent via a stable linker. In
some embodiments, the peptide has an isoelectric point of about
9.
[0018] In some embodiments, the peptide is linked to a detectable
agent. In some embodiments, the detectable agent is fused with the
peptide at an N-terminus or a C-terminus of the peptide. In some
embodiments, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 detectable agents are
linked to the peptide. In some embodiments, the peptide is linked
to the detectable agent via a cleavable linker. In some
embodiments, the peptide is linked to the detectable agent at an
N-terminus, at the epsilon amine of an internal lysine residue, or
a C-terminus of the peptide by a linker. In some embodiments, the
peptide active agent conjugate or peptide further comprises a
non-natural amino acid, wherein the non-natural amino acid is an
insertion, appendage, or substitution for another amino acid.
[0019] In some embodiments, the peptide is linked to the detectable
agent at the non-natural amino acid by a linker In some
embodiments, the linker comprises an amide bond, an ester bond, a
carbamate bond, a hydrazone bond, an oxime bond, or a
carbon-nitrogen bond. In some embodiments, the cleavable linker
comprises a cleavage site for matrix metalloproteinases, thrombin,
cathepsins, or beta-glucuronidase. In some embodiments, the peptide
is linked to the detectable agent via a stable linker. In some
embodiments, the detectable agent is a fluorophore, a near-infrared
dye, a contrast agent, a nanoparticle, a metal-containing
nanoparticle, a metal chelate, an X-ray contrast agent, a PET
agent, a radioisotope, or a radionuclide chelator. In some
embodiments, the detectable agent is a fluorescent dye.
[0020] In some aspects, a pharmaceutical composition comprises the
peptide active agent conjugate of any embodiment as described
herein or a salt thereof, or the peptide of any embodiment as
described herein or a salt thereof, and a pharmaceutically
acceptable carrier. In some embodiments, the pharmaceutical
composition is formulated for administration to a subject. In some
embodiments, the pharmaceutical composition is formulated for
inhalation, intranasal administration, oral administration, topical
administration, parenteral administration, intravenous
administration, subcutaneous administration, intra-articular
administration, intramuscular administration, intraperitoneal
administration, dermal administration, transdermal administration,
or a combination thereof.
[0021] In some aspects, a method of treating a condition in a
subject in need thereof comprises administering to the subject the
peptide active agent conjugate of any of embodiment as described
herein, the peptide of any of any embodiment as described herein,
or a pharmaceutical composition of any embodiment as described
herein. In some embodiments, the peptide active agent conjugate,
peptide, or pharmaceutical composition is administered by
inhalation, intranasally, orally, topically, parenterally,
intravenously, subcutaneously, intra-articularly, intramuscularly
administration, intraperitoneally, dermally, transdermally, or a
combination thereof. In some embodiments, the peptide active agent
conjugate or the peptide homes, targets, or migrates to cartilage
of the subject following administration. In some embodiments, the
condition is associated with cartilage. In some embodiments, the
condition is associated with a joint. In some embodiments, the
condition is an inflammation, a cancer, a degradation, a growth
disturbance, genetic, a tear, an infection, a disease, or an
injury. In some embodiments, the condition is a chondrodystrophy.
In some embodiments, the condition is a traumatic rupture or
detachment. In some embodiments, the condition is a
costochondritis. In some embodiments, the condition is a
herniation. In some embodiments, the condition is a polychondritis.
In some embodiments, the condition is a chordoma. In some
embodiments, the condition is a type of arthritis. In some
embodiments, the type of arthritis is rheumatoid arthritis. In some
embodiments, the type of arthritis is osteoarthritis. In some
embodiments, the condition is achondroplasia. In some embodiments,
the condition is benign chondroma or malignant chondrosarcoma. In
some embodiments, the condition is bursitis, tendinitis, gout,
pseudogout, an arthropathy, psoriatic arthritis, ankylosing
spondylitis, or an infection. In some embodiments, the peptide
active agent conjugate, peptide, or pharmaceutical composition is
administered to treat the injury, to repair a tissue damaged by the
injury, or to treat a pain caused by the injury. In some
embodiments, the peptide active agent conjugate, peptide, or
pharmaceutical composition is administered to treat the tear or to
repair a tissue damaged by the tear. In some embodiments, the
peptide active agent conjugate, peptide, or pharmaceutical
composition homes, targets, or migrates to a kidney of the subject
following administration. In some embodiments, the condition is
associated with a kidney. In some embodiments, the condition is
lupus nephritis, acute kidney injury (AM), chronic kidney disease
(CKD), hypertensive kidney damage, diabetic nephropathy, or renal
fibrosis.
[0022] In some aspects, a method of imaging an organ or body region
of a subject comprises: administering to the subject the peptide
active agent conjugate of any embodiment as described herein, the
peptide of any embodiment as described herein, or the
pharmaceutical composition of any embodiment as described herein;
and imaging the subject. In some embodiments, the method further
comprises detecting a cancer or diseased region, tissue, structure,
or cell. In some embodiments, the method further comprises
performing surgery on the subject. In some embodiments, the method
further comprises treating the cancer. In some embodiments, the
surgery comprises removing the cancer or the diseased region,
tissue, structure or cell of the subject. In some embodiments, the
method further comprises imaging the cancer or diseased region,
tissue, structure, or cell of the subject after surgical removal.
In some embodiments, the peptide active agent conjugate is
expressed as a fusion protein.
INCORPORATION BY REFERENCE
[0023] All publications, patents, and patent applications
mentioned, disclosed or referenced in this specification are herein
incorporated by reference in their entirety and to the same extent
as if each individual publication, patent, or patent application
was specifically and individually indicated to be incorporated by
reference.
BRIEF DESCRIPTION OF THE FIGURES
[0024] The novel features of the invention are set forth with
particularity in the appended claims. A better understanding of the
features and advantages of the present disclosure will be obtained
by reference to the following detailed description that sets forth
illustrative embodiments, in which the principles of the disclosure
are utilized, and the accompanying drawings of which:
[0025] FIG. 1 illustrates the identification of the .sup.14C signal
in the joint and other cartilage of an animal treated with the
peptide of SEQ ID NO: 27.
[0026] FIG. 2 illustrates a three-dimensional structure and a line
structure of a peptide of SEQ ID NO: 31.
[0027] FIG. 3 illustrates an exemplary architecture of constructs
expressing sequences of SEQ ID NO: X, where X can be any one of
peptides of SEQ ID NO: 24-SEQ ID NO: 36.
[0028] FIG. 4 illustrates a schematic of a method of manufacturing
of a peptide of the disclosure.
[0029] FIG. 5 illustrates alignment of SEQ ID NO: 541 (SEQ ID NO:
541 is SEQ ID NO: 27, but without the first three amino acids "GSG"
and is also SEQ ID NO: 317, but without the first amino acid "G")
with SEQ ID NO: 316, SEQ ID NO: 541 with SEQ ID NO: 542 (SEQ ID NO:
542 is SEQ ID NO: 30, but without the first three amino acids "GSQ"
and is SEQ ID NO: 320, but without the first amino acid "Q"), and
SEQ ID NO: 541 with SEQ ID NO: 483. FIG. 5A illustrates the
alignment of the peptide of SEQ ID NO: 541 with the peptide of SEQ
ID NO: 316. Boxes delineate conserved positively charged residues.
FIG. 5B illustrates the alignment of the peptide of SEQ ID NO: 541
with the peptide of SEQ ID NO: 542. Boxes delineate conserved
positively charged residues. FIG. 5C illustrates the alignment of
the peptide of SEQ ID NO: 541 with the peptide of SEQ ID NO: 483.
Boxes delineate conserved positively charged residues.
[0030] FIG. 6 illustrates the alignment of the peptide of SEQ ID
NO: 320 with the peptide of SEQ ID NO: 484. Boxes delineate
conserved positively charged residues.
[0031] FIG. 7 illustrates alignment of peptides within the
pfam00451:toxin 2 structural class family of SEQ ID NO: 494-SEQ ID
NO: 540. Boxed and bolded residues indicate relative conservation
of sequence while non-boxed and non-bolded residues indicate areas
of higher sequence variability.
[0032] FIG. 8 illustrates alignment of a peptide of SEQ ID NO: 494
from the pfam00451:toxin 2 structural class family with a cartilage
homing peptide of this disclosure of SEQ ID NO: 27. Asterisks
indicate positions with a single, fully conserved residue, a colon
indicates conservation between groups of strongly similar
properties (scoring >0.5 in the Gonnet point accepted mutation
(PAM) 250 matrix), and a period indicates conservation between
groups of weakly similar properties (scoring .ltoreq.0.5 in the
Gonnet PAM 250 matrix).
[0033] FIG. 9 illustrates the .sup.14C signal in the cartilage of
an animal with intact kidneys 24 hours after treatment with a
peptide of SEQ ID NO: 27.
[0034] FIG. 10 shows white light images and corresponding whole
body fluorescence images of a mouse administered 10 nmol of a
peptide of SEQ ID NO: 108 conjugated to a Cy5. 5 fluorophore (SEQ
ID NO: 108A) at 24 hours post-administration. FIG. 10A illustrates
an image of a frozen section of a mouse, 24 hours after
administration of 10 nmol of a peptide of SEQ ID NO: 108 conjugated
to a Cy5. 5 fluorophore (SEQ ID NO: 108A). FIG. 10B illustrates the
fluorescence signal in the mouse, corresponding to the section
shown in FIG. 10A, 24 hours after administration of 10 nmol of a
peptide of SEQ ID NO: 108 conjugated to a Cy5. 5 fluorophore (SEQ
ID NO: 108A). FIG. 10C illustrates an image of a different frozen
section of the mouse, 24 hours after administration of 10 nmol of a
peptide of SEQ ID NO: 108 conjugated to a Cy5. 5 fluorophore (SEQ
ID NO: 108A). FIG. 10D illustrates the fluorescence signal in the
mouse, corresponding to the section shown in FIG. 10C, 24 hours
after administration of 10 nmol of a peptide of SEQ ID NO: 108
conjugated to a Cy5. 5 fluorophore (SEQ ID NO: 108A). FIG. 10E
illustrates an image of a different frozen section of the mouse, 24
hours after administration of 10 nmol of a peptide of SEQ ID NO:
108 conjugated to a Cy5. 5 fluorophore (SEQ ID NO: 108A).
[0035] FIG. 10F illustrates a fluorescence signal in the mouse,
corresponding to the section shown in FIG. 10E, 24 hours after
administration of 10 nmol of a peptide of SEQ ID NO: 108 conjugated
to a Cy5. 5 fluorophore (SEQ ID NO: 108A).
[0036] FIG. 11 illustrates a multiple sequence alignment of SEQ ID
NO: 316, SEQ ID NO: 317, SEQ ID NO: 321, SEQ ID NO: 333, SEQ ID NO:
337, SEQ ID NO: 338, SEQ ID NO: 340, SEQ ID NO: 398, SEQ ID NO:
474, SEQ ID NO: 483, SEQ ID NO: 486, and SEQ ID NO: 543-SEQ ID NO:
549 were used to predict enhanced peptide stability and
immunogenicity. SEQ ID NO: 295 is a consensus sequence.
[0037] FIG. 12 illustrates the identification of locations the
.sup.14C signal in the nasal, spinal, tracheal, and other cartilage
of an animal treated with the peptide of SEQ ID NO: 27.
[0038] FIG. 13 shows IVIS fluorescence imaging of an isolated hind
limb from a first mouse and an isolated hind limb from a second
mouse after administration of 10 nmol SEQ ID NO: 108 peptide
conjugated to a Cy5.5 fluorophore (SEQ ID NO: 108A). Areas of low
signal intensity are shown in a thin solid line, areas of medium
signal intensity are shown in a thick sold line, and areas of high
signal intensity are shown in a thin dotted line. FIG. 13A shows
the right hind limb with skin removed from a first mouse and from a
second mouse 3 hours after peptide administration. FIG. 13B shows
the right hind limb with muscle removed from a first mouse and from
a second mouse 3 hours after peptide administration of 10 nmol SEQ
ID NO: 108 peptide conjugated to a Cy5.5 fluorophore (SEQ ID NO:
108A). FIG. 13C shows the right hind limb with skin removed from a
first mouse and from a second mouse 24 hours after peptide
administration of 10 nmol SEQ ID NO: 108 peptide conjugated to a
Cy5.5 fluorophore (SEQ ID NO: 108A). FIG. 13D shows the right hind
limb with muscle removed from a first mouse and from a second mouse
24 hours after peptide administration of 10 nmol SEQ ID NO: 108
peptide conjugated to a Cy5.5 fluorophore (SEQ ID NO: 108A). FIG.
13E shows the right hind limb with skin removed from a first mouse
and from a second mouse 48 hours after peptide administration of 10
nmol SEQ ID NO: 108 peptide conjugated to a Cy5.5 fluorophore (SEQ
ID NO: 108A). FIG. 13F shows the right hind limb with muscle
removed from a first mouse and from a second mouse 48 hours after
peptide administration of 10 nmol SEQ ID NO: 108 peptide conjugated
to a Cy5.5 fluorophore (SEQ ID NO: 108A). FIG. 13G shows the right
hind limb with skin removed from a first mouse and from a second
mouse 72 hours after peptide administration of 10 nmol SEQ ID NO:
108 peptide conjugated to a Cy5.5 fluorophore (SEQ ID NO: 108A).
FIG. 13H shows the right hind limb with muscle removed from a first
mouse and from a second mouse 72 hours after administration of 10
nmol SEQ ID NO: 108 peptide conjugated to a Cy5.5 fluorophore (SEQ
ID NO: 108A).
[0039] FIG. 14 illustrates autoradiography images of frozen
sections from a mouse, 3 hours after administration of 100 nmol of
a radiolabeled peptide of SEQ ID NO: 108. FIG. 14A illustrates the
.sup.14C signal in a different frozen section of the mouse, 3 hours
after administration of 100 nmol of a radiolabeled peptide of SEQ
ID NO: 108. FIG. 14B illustrates the .sup.14C signal in a different
frozen section of a mouse, 3 hours after administration of 100 nmol
of a radiolabeled peptide of SEQ ID NO: 108
[0040] FIG. 15 illustrates autoradiography images of frozen
sections from a mouse, 3 hours after administration of 100 nmol of
a radiolabeled peptide of SEQ ID NO: 106. FIG. 15A illustrates the
.sup.14C signal in a frozen section of a mouse. FIG. 15B
illustrates the .sup.14C signal in a frozen section of the mouse, 3
hours after administration of 100 nmol of a radiolabeled peptide of
SEQ ID NO: 106.
[0041] FIG. 16 illustrates autoradiography images of frozen
sections from a mouse, 3 hours after administration of 100 nmol of
a radiolabeled peptide of SEQ ID NO: 187. FIG. 16A illustrates the
.sup.14C signal in a frozen section of the mouse, 3 hours after
administration of 100 nmol of a radiolabeled peptide of SEQ ID NO:
187. FIG. 16B illustrates the .sup.14C signal in a frozen section
of the mouse, 3 hours after administration of 100 nmol of a
radiolabeled peptide of SEQ ID NO: 187.
DETAILED DESCRIPTION
[0042] The present disclosure relates generally to compositions and
methods for cartilage therapy. In some embodiments, the
compositions and methods herein utilize peptides that home, target,
are directed to, are retained by, accumulate in, migrate to, and/or
bind to cartilage following administration to a subject. In some
embodiments, the cartilage homing peptides of the present
disclosure exert therapeutic effect in cartilage or tissue or cell
thereof. In some embodiments, the cartilage homing peptides of the
present disclosure are used to deliver an active agent to cartilage
or tissue or cell thereof. The active agent can exert a therapeutic
effect on cartilage or tissue or cell thereof. For example, in
certain embodiments, the peptide itself or the active agent allows
for localized delivery of an anti-inflammatory or other agent to
cartilage or tissue or cell thereof. As another example, the active
agent is a fluorophore that can be used for imaging of cartilage.
In certain embodiments, the peptide itself induces therapeutic
responses.
[0043] Cartilage disorders are particularly difficult to treat. A
direct route for active agent administration can be parenterally
(e.g., intravenously, subcutaneously, intramuscularly),
intra-articularly, by inhalation, dermally, topically, or orally.
However, cartilage can be avascular thus intravenous administration
of drugs can fail to reach the cartilage in significant amounts.
Drugs for cartilage diseases, such as osteoarthritis, can be
injected directly locally into the affected area, for example,
directly injected into the joint. Few drugs aimed at treating
cartilage disorders have proved therapeutically viable with lack of
access to target tissue being a primary reason for failure. The
lack of access to the target tissue can also lead to administration
of doses that are higher than would be necessary if a drug could
home, target, or be directed to, is retained by, and/or binds to a
target region, tissue, structure or cell. Thus, treatment of
cartilage conditions often requires the use of high concentrations
of non-specific drugs. In addition, a number of therapeutics are of
interest in treating joint disorders, but are problematic because
of the level of side effects caused by systemic administration of
the drug (Dancevic and McCulloch, Arthritis Res Ther. 16:429
(2014)).
[0044] Specific and potent drugs that are capable of contacting the
cartilage can counteract the non-specificity of many treatments by
selectively targeting and delivering compounds to specific regions,
tissues, cells and structures. Such drugs can also be useful to
modulate ion channels, protein-protein interactions, extracellular
matrix remodeling (i.e., protease inhibition), and the like. Such
targeted therapy can allow for lower dosing, reduced side effects,
improved patient compliance, and improvement in therapeutic
outcomes, which would be advantageous not only in acute disease of
the cartilage, but in chronic conditions as well.
[0045] The present disclosure provides peptides that can comprise
or can be derived from cystine-dense peptides. As used herein, the
term "cystine-dense peptide" can be interchangeable with the terms
"knotted peptide," "knottin," and "optide," and cystine-dense
peptides can also be abbreviated as "CDPs." Hitchins, amongst other
disulfide-containing peptides, can also be considered "knotted
peptides" or "cystine-dense peptides" for the purposes of this
disclosure. Knottins, for example, are a class of cystine-dense
peptides comprising from about 11 to about 80 amino acids in length
that are often folded into a compact structure. Knottins and other
cystine-dense peptides are typically assembled into a complex
tertiary structure that is characterized by a number of
intramolecular disulfide crosslinks and can contain beta strands,
an alpha helix, and other secondary structures. The presence of the
disulfide bonds can give cystine-dense peptides remarkable
environmental stability, allowing them to withstand extremes of
temperature and pH, to resist proteolytic enzymes in the blood
stream or digestive tract, and can provide specific
biodistribution, pharmacokinetic, binding interactions, cellular
processing, or other properties of physiologic and therapeutic
value. The peptides disclosed herein can be derived from certain
cystine-dense peptides. The present disclosure describes a class of
cystine-dense peptides that can effectively contact cartilage and
be used either directly or as carriers of active drugs, peptides,
or molecules to treat a cartilage condition. For instance,
osteoarthritis is a cartilage condition that is associated with the
thinning of cartilage covering the ends of bones resulting in bone
directly contacting bone within the joint. Over time, the ends of
the bones are subjected to increased levels of friction which
ultimately causes erosion of the end of the bone. Individuals
suffering from osteoarthritis experience reduced motion and
increased pain. A therapeutic peptide that could contact the
cartilage at the joint and ends of the bone to interact with the
chondrocytes and induce increased expression of extracellular
matrix proteins could be used in the treatment and prevention of
osteoarthritis by increasing expression of collagen through, for
example, the rate of production, amount of production, inhibition
of proteins which degrade collagen, promote expression of other
proteins which maintain the integrity of existing collagen
proteins, or other mechanism. A peptide could also affect nearby
tissues or cells such as the bone, ligaments, muscle, tendons,
bursa, connective tissue, blood vessels, peripheral nerves,
osteoclasts, osteoblasts, fibroblasts, synoviocytes,
monocytes/macrophages, lymphocytes, plasma cells, adipocytes,
endothelial cells, neurons, ligaments, muscle, tendons, and bursa.
The peptides of the disclosure can be used to treat the symptoms of
various conditions. The peptides of the disclosure can bind to,
home to, migrate to, accumulate in, be retained by, or be directed
to cartilage and its components, including chondrocytes,
extracellular matrix, collagen, hyaluranon, aggrecan (also known as
cartilage-specific proteoglycan core protein (CSPCP)), or other
components of the extracellular matrix and the joint, or to other
nearby components such as those described herein in joints and
cartilaginous tissues as listed above.
[0046] Also described herein are peptides that selectively home,
target, are directed to, migrate to, are retained by, or accumulate
in and/or bind to specific regions, tissues, structures or cells of
the cartilage that aid in managing, decreasing, ablating or
reducing pain (e.g., joint pain) due to chronic disease or
cartilage injury or other therapeutic indications as described
herein. A peptide that homes, targets, migrates to, is directed to,
is retained by, or accumulates in and/or binds to one or more
specific regions, tissues, structures or cells of the cartilage can
have fewer off-target and potentially negative effects, for
example, side effects that often limit use and efficacy of pain
drugs. In addition, such peptides can reduce dosage and increase
the efficacy of existing drugs by directly targeting them to a
specific region, tissue, structure or cell of the cartilage and
helping the contact the cartilage or increasing the local
concentration of agent. The peptide itself can modulate pain or it
can be conjugated to an agent that modulates pain. Such pain
modulation may operate by various mechanisms such as modulating
inflammation, autoimmune responses, direct or indirect action on
pain receptors, cell killing, or programmed cell death (whether via
an apoptotic and/or non-apoptotic pathway of diseased cells or
tissues, and the like (Tait et al., J Cell Sci 127(Pt 10):2135-44
(2014)).
[0047] Peptides of this disclosure that home, target, are directed
to, migrate to, are retained by, accumulate in, or bind to specific
regions, tissues, structures or cells of the cartilage can do so
with different degrees of efficiency. Peptides can have a higher
concentration in cartilage than in other locations, such as blood
or muscle. Peptides can be recorded as having a signal in cartilage
as a percentage of signal in blood. For example, a cartilage signal
of 200% indicates that the signal in cartilage is twice as high as
the signal in blood. In some embodiments, peptides that have
cartilage homing properties can have a cartilage signal of >170%
by radiographic densitometry measurements. In other embodiments,
peptides that are cartilage homers can have a cartilage signal of
>200% by radiographic densitometry measurements. In other
embodiments, peptides that are more efficient cartilage homers can
have a cartilage signal of >300% by radiographic densitometry
measurements. In other embodiments, peptides that are more
efficient cartilage homers can have a cartilage signal of >400%
by radiographic densitometry measurements. In other embodiments,
peptides that are strongest cartilage homers of highest interest
can have a cartilage signal of >500% by radiographic
densitometry measurements. In some embodiments, measurement of the
ratio of peptide concentration in blood, muscle, or other tissues
relative to the peptide concentration in cartilage can be performed
using various methods including measuring the densitometry signal
of peptides labeled with radioisotopes (as described above), or by
using other assays.
[0048] Peptides that selectively home, target, are directed to,
migrate to, are retained by, or accumulate in and/or bind to
specific regions, tissues, structures or cells of the cartilage can
occur after administration of the peptide to a subject. A subject
can be a human or a non-human animal.
[0049] The peptides disclosed herein can be used as active agents,
or conjugated to detection agents such a fluorophores,
iodide-containing X-ray contrast agents, lanthanide chelates (e.g.,
gadolinium for MRI imaging), perfluorocarbons (for ultrasound), or
PET tracers (e.g., 18F or 11C) for imaging and tracing the peptide,
or conjugated to agents such as anti-inflammatory active agents or
other active agents to the joint to treat inflammation or other
disease.
[0050] The peptides disclosed herein can be used to bind cartilage
explants ex vivo. Cartilage explants can be from any subject, such
as a human or an animal. Assessment of peptide binding to cartilage
explants can be used to screen peptides that may efficiently home
to cartilage in vivo
[0051] In some embodiments, peptides of this disclosure home,
target, are directed to, migrate to, are retained by, accumulate
in, or bind to specific regions, tissues, structures or cells of
the kidneys. For example, in some embodiments, peptides of this
disclosure home, target, are directed to, migrate to, are retained
by, accumulate in, or bind to the proximal tubules of the kidneys,
kidney nephrons, or podocytes. Peptides that selectively home,
target, are directed to, migrate to, are retained by, or accumulate
in and/or bind to specific regions, tissues, structures or cells of
the kidney can occur after administration of the peptide to a
subject. A subject can be a human or a non-human animal. The
peptides disclosed herein can be used as active agents, or
conjugated to detection agents such a fluorophores,
iodide-containing X-ray contrast agents, lanthanide chelates (e.g.,
gadolinium for MRI imaging), perfluorocarbons (for ultrasound), or
PET tracers (e.g., 18F or 11C) for imaging and tracing the peptide,
or conjugated to agents such as anti-inflammatory agents or other
agents to the kidney to treat renal cancer, chronic kidney failure
or other kidney disease.
[0052] One roadblock in the advancement and wide spread use of
peptides as a therapeutic is that peptides can be chemically and
physically unstable. During the process of manufacturing of
therapeutic peptides essential considerations can include storage
conditions, sustained biochemical function, and in vivo delivery.
Peptide degradation products can result in the formation of species
that alter the safety profile, potency, and immunogenicity of the
peptide. These peptide degradation products can form during
manufacture and storage, as well as in vivo after delivery to a
patient. Furthermore, peptide degradation may limit the shelf-life
and increase production cost due to unstable peptides requiring
refrigeration or shipment on dry ice. The latter can necessitate
continual monitoring and validation of peptides as degradation
products could have formed during the manufacturing process. Hence,
there is an urgent need for the rationale design and production of
therapeutic peptides that have enhanced stability, for example, in
the ambient environment, during the process of manufacturing, in
storage, and that prevent the likelihood of peptide degradation
under a variety of conditions.
[0053] In some embodiments, the peptides and peptide-drug
conjugates of the present disclosure have stability properties that
minimize peptide or peptide-drug conjugate degradation to enable
adequate storage. Long term, accelerated, and intermediate storage
conditions for the peptides and peptide-drug conjugates of the
present disclosure can include long term storage conditions of
25.degree. C..+-.2.degree. C./60% relative humidity (RH) .+-.5% RH,
or 30.degree. C..+-.2.degree. C./65% RH .+-.5% RH for at least 6
months, at least 12 months, and up to 1 year, up to 2 years, up to
3 years, up to 4 years, or longer than 4 years. In addition,
intermediate and short term storage conditions (e.g., during
transport, distribution, manufacturing, or handling), or long term
storage conditions for certain climates and infrastructures, can
include storage conditions of 30.degree. C..+-.2.degree. C./65% RH
.+-.5% RH or 40.degree. C..+-.2.degree. C./75% RH .+-.5% RH for up
to 1 hour, for up to 8 hours, for up to 1 day, for up to 3 days,
for up to 1 week, for up to 1 month, for up to 3 months, for up to
6 months or at least 6 months, up to 1 year, up to 2 years, up to 3
years, up to 4 years, or longer than 4 years). Moreover, the
peptides and peptide-drug conjugates of the present disclosure can
be refrigerated, for example between 5.degree. C..+-.3.degree. C.
for at least 6 months, at least 12 months, and up to 1 year, up to
2 years, up to 3 years, up to 4 years, or longer than 4 years. In
addition, intermediate and short term refrigeration conditions
(e.g., during transport, distribution, manufacturing, or handling)
can include 25.degree. C..+-.2.degree. C./60% RH .+-.5% RH for up
to 1 hour, for up to 8 hours, for up to 1 day, for up to 3 days,
for up to 1 week, for up to 1 month, for up to 3 months, for up to
6 months or at least 6 months, and potentially longer (at least 12
months and up to 1 year, up to 2 years, up to 3 years, up to 4
years, or longer than 4 years). Such conditions for storage,
whether based on ambient or refrigerated conditions can be adjusted
based upon the four zones in the world (e.g., the International
Council for Harmonisation of Technical Requirements for
Pharmaceuticals for Human Use (ICH) stability Zone I, II, III, or
IV) that are distinguished by their characteristic prevalent annual
climatic conditions. In addition, formulation components can be
principally chosen for their ability to preserve the native
conformation and chemical structure of the peptides and
peptide-drug conjugates of the present disclosure in storage by
preventing denaturation due to hydrophobic interactions and
aggregation, as well as by preventing chemical degradation,
including truncation, oxidation, deamidation, cleavage, hydrolysis,
isomerization, disulfide exchange, racemization, and beta
elimination (Cleland, et al., Crit Rev Ther Drug Carrier Syst
10(4): 307-377 (1993); Shire et al., J Pharm Sci 93(6): 1390-1402
(2004); Wakankar and Borchardt, J Pharm Sci 95(11): 2321-2336
(2006)).
[0054] In some embodiments, the peptides and peptide-drug
conjugates of the present disclosure have incorporated properties
that minimize immunogenicity of the peptides and peptide-drug
conjugates. Immunogenicity can be a major concern with the
development of therapeutic peptides and proteins, and there is an
urgent need for the rationale design and production of therapeutic
peptides that have reduced immunogenicity and that increase their
safety and efficacy. Immunogenicity can occur against a desired
peptide sequence or a peptide degradation product. Immunogenicity
can occur when a patient develops an immune response to the
therapeutic peptide, protein, conjugate, or other drug, such as by
producing antibodies that bind to and/or neutralize the therapeutic
peptide, protein, conjugate, or other drug. The likelihood of
immunogenicity can increase when drugs are administered more than
once or chronically. Immunogenicity can reduce patient exposure to
the drug, can reduce effectiveness of the drug, and can also result
in safety risks for the patient, such as generating an immune
response to self-proteins or other adverse responses related to
increased immunogenicity to the therapeutic peptide, protein,
conjugate, or other drug. Immunogenic responses can vary from
patient to patient and also amongst different groups of HLA
alleles, as well as over time. As such, minimizing risk of
immunogenicity with a therapeutic peptide or protein can be
important for developing a drug that can be effectively and safely
used for treatment. Various methods exist for assessment of
immunogenic potential, which can include in silico methods, in
vitro testing, preclinical in vivo testing, and assessment during
clinical dosing. Evaluation early in product design and development
of the therapeutic peptides and peptide-drug conjugates of the
present disclosure in the in vivo milieu in which they function
(e.g., in inflammatory environments or at physiologic pH) can
reveal susceptibilities to modifications (e.g., aggregation and
deamidation) that can result in loss of efficacy or induction of
immune responses. Such information can be used to facilitate
product engineering to enhance the stability of the product under
such in vivo conditions or reduce immunogenicity. Moreover, the
therapeutic peptides and peptide-drug conjugates of the present
disclosure can be designed to minimize protein aggregation.
Strategies to minimize aggregate formation can be used early in
drug development, for example, by using an appropriate cell
substrate, selecting manufacturing conditions that minimize
aggregate formation, employing a robust purification scheme that
removes aggregates to the greatest extent possible, and choosing a
formulation and container closure system that minimize aggregation
during storage.
[0055] Additional aspects and advantages of the present disclosure
will become apparent to those skilled in this art from the
following detailed description, wherein illustrative embodiments of
the present disclosure are shown and described. As will be
realized, the present disclosure is capable of other and different
embodiments, and its several details are capable of modifications
in various respects, all without departing from the disclosure.
Accordingly, the drawings and description are to be regarded as
illustrative in nature, and not as restrictive.
[0056] As used herein, the abbreviations for the natural
L-enantiomeric amino acids are conventional and are as follows:
alanine (A, Ala); arginine (R, Arg); asparagine (N, Asn); aspartic
acid (D, Asp); cysteine (C, Cys); glutamic acid (E, Glu); glutamine
(Q, Gln); glycine (G, Gly); histidine (H, His); isoleucine (I,
Ile); leucine (L, Leu); lysine (K, Lys); methionine (M, Met);
phenylalanine (F, Phe); proline (P, Pro); serine (S, Ser);
threonine (T, Thr); tryptophan (W, Trp); tyrosine (Y, Tyr); valine
(V, Val). Typically, Xaa can indicate any amino acid. In some
embodiments, X can be asparagine (N), glutamine (Q), histidine (H),
lysine (K), or arginine (R).
[0057] Some embodiments of the disclosure contemplate D-amino acid
residues of any standard or non-standard amino acid or analogue
thereof. When an amino acid sequence is represented as a series of
three-letter or one-letter amino acid abbreviations, the left-hand
direction is the amino terminal direction and the right-hand
direction is the carboxyl terminal direction, in accordance with
standard usage and convention.
Peptides
[0058] The cystine-dense peptides herein can bind targets with
antibody-like affinity. The cystine-dense peptides can modulate the
activity of a plurality of cartilage regions, tissues, structures
or cells. For example, in some embodiments, the cystine-dense
peptide conjugated to a bone-modifying drug homes to the cartilage
of a diseased joint and releases the drug, creating a higher local
concentration of drug in an area of eroded or damaged bone than
would be achieved without the cartilage targeting function of the
peptide. The cystine-dense peptide can be conjugated to a drug that
can affect nearby tissues or cells such as the ligaments, muscle,
tendons, bursa, connective tissue, blood vessels, peripheral
nerves, osteoclasts, osteoblasts, fibroblasts, synoviocytes,
monocytes/macrophages, lymphocytes, plasma cells, adipocytes,
endothelial cells, neurons, ligaments, muscle, tendons, and bursa.
The cystine-dense peptide conjugated to a drug can bind to, home
to, migrate to, accumulate in, be retained by, or be directed to
cartilage and its components, including chondrocytes, extracellular
matrix, collagen of any type, hyaluranon, aggrecan (also known as
cartilage-specific proteoglycan core protein (CSPCP)),
proteoglycans, glycoasminoglycans, glycoproteins, decorin,
biclycan, fibromodulin, or other components of the extracellular
matrix and the joint, or to other nearby components such as those
described herein in joints and cartilaginous tissues as listed
above. Some of the cartilage regions, tissues, and structures that
peptides and peptide-drug conjugates can target to treat a
cartilage-associated disorder include: (a) elastic cartilage; (b)
hyaline cartilage, such as articular cartilage and physeal
cartilage; (c) fibrocartilage; and (d) any cells or cell types in
(a)-(c) above. Some of the areas where the peptide and peptide-drug
conjugates can target to treat a cartilage-associated disorder
include: cartilage includes joints such as knees, hips, or digits,
nasal cartilage, spinal cartilage, tracheal cartilage, and rib
cartilage. In various aspects, cartilage components include
aggrecan and type II collagen. Additionally, in some embodiments,
cystine-dense peptides can penetrate into cells. In other
embodiments, cystine-dense peptides do not enter cells. In other
embodiments, cystine-dense peptides exhibit more rapid clearance
and cellular uptake compared to other types of molecules.
[0059] The peptides of the present disclosure can comprise cysteine
amino acid residues. In some cases, the peptide has at least 4
cysteine amino acid residues. In some cases, the peptide has at
least 6 cysteine amino acid residues. In other cases, the peptide
has at least 8 cysteine amino acid residues, at least 10 cysteine
amino acid residues, at least 12 cysteine amino acid residues, at
least 14 cysteine amino acid residues or at least 16 cysteine amino
acid residues.
[0060] A cystine-dense peptide can comprise disulfide bridges. A
cystine-dense can be a peptide wherein 5% or more of the residues
are cysteines forming intramolecular disulfide bonds as cystines. A
disulfide-linked peptide can be a drug scaffold. In some
embodiments, the disulfide bridges form an inhibitor knot. A
disulfide bridge can be formed between cysteine residues, for
example, between cysteines 1 and 4, 2 and 5, or, 3 and 6. In some
cases, one disulfide bridge passes through a loop formed by the
other two disulfide bridges, for example, to form the inhibitor
knot. In other cases, the disulfide bridges can be formed between
any two cysteine residues.
[0061] The present disclosure further includes peptide scaffolds
that, e.g., can be used as a starting point for generating
additional peptides that can target and home to cartilage. In some
embodiments, these scaffolds can be derived from a variety of
cystine-dense peptides. In certain embodiments, cystine-dense
peptides are assembled into a complex tertiary structure that is
characterized by a number of intramolecular disulfide crosslinks,
and optionally contain beta strands and other secondary structures
such as an alpha helix. For example, cystine-dense peptides
include, in some embodiments, small disulfide-rich proteins
characterized by a disulfide through disulfide knot. This knot can
be, e.g., obtained when one disulfide bridge crosses the macrocycle
formed by two other disulfides and the interconnecting backbone. In
some embodiments, the cystine-dense peptides can include growth
factor cysteine knots or inhibitor cysteine knots. Other possible
peptide structures can include peptide having two parallel helices
linked by two disulfide bridges without .beta.-sheets (e.g.,
hefutoxin).
[0062] A cystine-dense peptide can comprise at least one amino acid
residue in an L configuration. A cystine-dense peptide can comprise
at least one amino acid residue in a D configuration. In some
embodiments, a cystine-dense peptide is 15-40 amino acid residues
long. In other embodiments, a cystine-dense peptide is 11-57 amino
acid residues long. In further embodiments, a cystine-dense peptide
is at least 20 amino acid residues long.
[0063] In some embodiments, the peptides are members of the
pfam00451:toxin 2 family. The pfam00451:toxin 2 structural class
family can include a peptide of any one of SEQ ID NO: 494-SEQ ID
NO: 540. A cartilage homing peptide of this disclosure can be a
variant of any peptide members of the pfam00451:toxin 2 family. In
some embodiments, an exemplary cartilage homing peptide of this
disclosure that is a variant of the pfam00451:toxin 2 structural
class family is a peptide of SEQ ID NO: 27. In other embodiments,
an exemplary cartilage homing peptide of this disclosure that is a
variant of the pfam00451:toxin 2 structural class family is a
peptide of SEQ ID NO: 108. In other embodiments, the variant
peptides are at least 30% identical to a peptide of the structural
class pfam00451:toxin 2 family. In some embodiments, the variant
peptides are 30%, 40%, 50%, 60%, 80%, 90% or 95% identical to a
peptide of the structural class pfam00451:toxin 2 family. In some
embodiments, the variant peptides are at least 30%, at least 40%,
at least 50%, at least 60%, at least 80%, at least 90% or at least
95% identical to a peptide of the structural class pfam00451:toxin
2 family.
[0064] In some embodiments, cartilage homing peptides are family
members the sequences
TABLE-US-00001 (SEQ ID NO: 9)
GSXVXXXVKCXGSKQCXXPCKRXXGXRXGKCINKKXCKCYXXX or (SEQ ID NO: 283)
XVXXXVKCXGSKQCXXPCKRXXGXRXGKCINKKXCKCYXXX
wherein X can be any amino acid or amino acid analogue or null, in
which these sequences are based on the most common elements found
in the following sequences:
TABLE-US-00002 (SEQ ID NO: 27)
GSGVPINVKCRGSRDCLDPCKKA-GMRFGKCINSK-CHCTP--, (SEQ ID NO: 26)
GS-VRIPVSCKHSGQCLKPCKDA-GMRFGKCMNGK-CDCTPK-, (SEQ ID NO: 30)
GSQVQTNVKCQGGS-CASVCRREIGVAAGKCINGK-CVCYRN-, (SEQ ID NO: 29)
GS-----ISCTGSKQCYDPCKRKTGCPNAKCMNKS-CKCYGCG, (SEQ ID NO: 31)
GSEV---IRCSGSKQCYGPCKQQTGCTNSKCMNKV-CKCYGCG, (SEQ ID NO: 28)
GSAVCVYRT------CDKDCKRR-GYRSGKCINNA-CKCYPYG, (SEQ ID NO: 24)
GS----GIVC---KVCKIICGMQ-GKKVNICKAPIKCKCKKG-, and (SEQ ID NO: 33)
GSQIYTSKECNGSSECYSHCEGITGKRSGKCINKK-CYCYR--,
where the following residues may be independently interchanged in
the sequences: K and R; M, I, L, and V; G and A; S and T; Q and N;
and X can independently be any number of any amino acid or no amino
acid. The N-terminal GS sequence can be included or excluded
between the peptides of the present disclosure.
[0065] In some embodiments, cartilage homing peptides are family
members of the sequences
TABLE-US-00003 (SEQ ID NO. 21)
GSXVXIXVRCXGSXQCLXPCRXAXGXRXGKCMNGRCXCXPXX or (SEQ ID NO. 295)
XVXIXVRCXGSXQCLXPCRXAXGXRXGRCMNGRCXCXPXX,
wherein X can be any amino acid or amino acid analogue or null, in
which these sequences are based on the most common elements found
in the following sequences:
TABLE-US-00004 (SEQ ID NO: 316)
-VRIPVSCKHSGQCLKPCKDA-GMRFGKCMNGKCDCTPK-, (SEQ ID NO: 317)
GVPINVKCRGSRDCLDPCKKA-GMRFGKCINSKCHCTP--, (SEQ ID NO: 321)
---EVIRCSGSKQCYGPCKQQTGCTNSKCMNKVCKCYGCG, (SEQ ID NO: 333)
GVIINVKCKISRQCLEPCKKA-GMRFGKCMNGKCHCTPK-, (SEQ ID NO: 337)
GVPTDVKCRGSPQCIQPCKDA-GMRFGKCMNGKCHCTPK-, (SEQ ID NO: 338)
GVPINVSCTGSPQCIKPCKDA-GMRFGKCMNRKCHCTPK-, (SEQ ID NO: 340)
-VGINVKCKHSGQCLKPCKDA-GMRFGKCINGKCDCTPK-, (SEQ ID NO: 398)
GVPINVRCRGSRDCLDPCRRA-GMRFGRCINSRCHCTP--, (SEQ ID NO: 474)
QKILSNRCNNSSECIPHCIRIFGTRAAKCINRKCYCYP--, (SEQ ID NO: 483)
-VFINVKCRGSPECLPKCKEAIGKSAGKCMNGKCKCYP--, (SEQ ID NO: 486)
-VPTDVKCRGSPQCIQPCKDA-GMRFGKCMNGKCHCTP--, (SEQ ID NO: 543)
--AEIIRCSGTRECYAPCQKLTGCLNAKCMNKACKCYGCV, (SEQ ID NO: 544)
-RPTDIKCSASYQCFPVCKSRFGKTNGRCVNGLCDCF---, (SEQ ID NO: 545)
-QFTDVKCTGSKQCWPVCKQMFGKPNGKCMNGKCRCYS--, (SEQ ID NO: 546)
-VGINVKCKHSRQCLKPCKDA-GMRFGKCTNGKCHCTPK-, (SEQ ID NO: 547)
-VVIGQRCYRSPDCYSACKKLVGKATGKCTNGRCDC----, (SEQ ID NO: 548)
--NFKVEGACSKPCRKYCIDK-GARNGKCINGRCHCYY--, and (SEQ ID NO: 549)
QIDTNVKCSGSSKCVKICIDRYNTRGAKCINGRCTCYP--.
[0066] In some embodiments, the cartilage homing peptides are
family members of the
TABLE-US-00005 (SEQ ID NO: 22)
GSXVXIXVRCXGSXQCLXPCRXAXGXRXGRCMNGRCXCXPXX or (SEQ ID NO: 296)
XVXIXVRCXGSXQCLXPCRXAXGXRXGRCMNGRCXCXPXX
[0067] sequences wherein X can be any amino acid or amino acid
analogue or null, in which these sequences are based on the most
common elements found in the following sequences and with K
interchanged with R: SEQ ID NO: 316, SEQ ID NO: 317, SEQ ID NO:
321, SEQ ID NO: 333, SEQ ID NO: 337, SEQ ID NO: 338, SEQ ID NO:
340, SEQ ID NO: 398, SEQ ID NO: 483, SEQ ID NO: 486, or SEQ ID NO:
543-SEQ ID NO: 549.
[0068] In some embodiments, a peptide comprises the sequence
TABLE-US-00006 (SEQ ID NO: 23)
GSGVPIX.sup.1VRCRGSRDCX.sup.2X.sup.3PCRRAGX.sup.4RFGRCIX.sup.5X.sup.6RCX.-
sup.7CX.sup.8P or (SEQ ID NO: 297)
GVPIX.sup.1VRCRGSRDCX.sup.2X.sup.3PCRRAGX.sup.4RFGRCIX.sup.5X.sup.6RCX.su-
p.7CX.sup.8P,
where the following residues where X.sup.1 is selected from N, S,
or G, wherein X.sup.2 is selected from L or Y, wherein X.sup.3 is
selected from D or E, wherein X.sup.4 is selected from M or T,
wherein X.sup.5 is selected from N, Q, A, S, T, or L, wherein
X.sup.6 is selected from S, G, or R, wherein X.sup.7 is selected
from H or Y, and wherein X.sup.8 is selected from T or Y. In some
embodiments, zero or one or more of the R residues in SEQ ID NO: 23
or SEQ ID NO: 297 can be replaced with K residues. In some
embodiments, zero or one or more of the R residues in SEQ ID NO: 23
or SEQ ID NO: 297 can be replaced with A residues. In other
embodiments, zero or one or more R residues in SEQ ID NO: 23 or SEQ
ID NO: 297 can each be replaced with either a K or an A residue in
any combination. In other embodiments, peptides are family members
of the sequence
TABLE-US-00007 (SEQ ID NO: 10)
GSXXXGCVXXXXKCRPGXKXCCXPXKRCSRRFGXXXXKKCKXXXXXX or (SEQ ID NO: 284)
XXXGCVXXXXKCRPGXKXCCXPXKRCSRRFGXXXXKKCKXXXXXX,
in which the sequence is based on the most common elements found in
the following sequences:
TABLE-US-00008 (SEQ ID NO: 32)
GS---ACKGVFDACTPGKNECC-PNRVCSDK-H----KWCKWKL---, (SEQ ID NO: 34)
GS---GCLEFWWKCNPNDDKCCRPKLKCSKLF-----KLCNFSFG--, (SEQ ID NO: 25)
GSSEKDCIKHLQRCR-ENKDCC--SKKCSRR-GTNPEKRCR------, and (SEQ ID NO:
36) GS---GCFGY--KCDYY-KGCCSGYV-CSPTW-----KWCVRPGPGR,
where the following residues may be independently interchanged in
the sequences: K and R; M, I, L, and V; G and A; S and T; Q and N;
and X can independently be any number of any amino acid or no amino
acid. The N-terminal GS sequence can be included or excluded
between the peptides of the present disclosure.
[0069] In some embodiments, a peptide comprises the sequence
TABLE-US-00009 (SEQ ID NO: 1)
GSGVX.sup.1IX.sup.2X.sup.3KCX.sup.4GSKQCX.sup.5DPCKX.sup.6X.sup.7X.sup.8G-
X.sup.9RX.sup.10GKCX.sup.11NKKCK CX.sup.12X.sup.13X.sup.14X.sup.15
or (SEQ ID NO: 275)
GVX.sup.1IX.sup.2X.sup.3KCX.sup.4GSKQCX.sup.5DPCKX.sup.6X.sup.7X.sup.8GX.-
sup.9RX.sup.10GKCX.sup.11NKKCKC
X.sup.12X.sup.13X.sup.14X.sup.15,
wherein X.sup.1, X.sup.2, X.sup.3, X.sup.4, X.sup.5, X.sup.6,
X.sup.7, X.sup.8, X.sup.9, X.sup.10, X.sup.11, X.sup.12, X.sup.13,
X.sup.14 and X.sup.15 are each individually any amino acid or amino
acid analogue or null. In some cases, the peptide comprises the
sequence
TABLE-US-00010 (SEQ ID NO: 2)
GSGVX.sup.1IX.sup.2X.sup.3KCX.sup.4GSKQCX.sup.5DPCKX.sup.6X.sup.7X.sup.8G-
X.sup.9RX.sup.10GKCX.sup.11NKKCKC X.sup.12X.sup.13X.sup.14X.sup.15
or (SEQ ID NO: 276)
GVX.sup.1IX.sup.2X.sup.3KCX.sup.4GSKQCX.sup.5DPCKX.sup.6X.sup.7X.sup.8GX.-
sup.9RX.sup.10GKCX.sup.11NKKCKC
X.sup.12X.sup.13X.sup.14X.sup.15,
[0070] where X.sup.1 is selected from P or R, wherein X.sup.2 is
selected from P or N, wherein X.sup.3 is selected from V or I,
wherein X.sup.4 is selected from S, T, R or K, wherein X.sup.5 is
selected from Y or L, wherein X.sup.6 is selected from Q, R or K,
wherein X.sup.7 is selected from A, K or R, wherein X.sup.8 is
selected from T or A, wherein X.sup.9 is selected from C or M,
wherein X.sup.10 is selected from F or N, wherein X.sup.11 is
selected from M or I, wherein X.sup.12 is selected from Y or T,
wherein X.sup.13 is selected from G or P, wherein X.sup.14 is
selected from C or null, and wherein X.sup.15 is selected from G or
null.
[0071] In some embodiments, a peptide comprises the sequence
TABLE-US-00011 (SEQ ID NO: 3)
GSX.sup.1X.sup.2X.sup.3X.sup.4IX.sup.5CX.sup.6GSKQCYX.sup.7PCKX.sup.8X.su-
p.9TGCX.sup.10X.sup.11X.sup.12KCX.sup.13 X.sup.14KX.sup.15CKCYGCG
or (SEQ ID NO: 277)
X.sup.1X.sup.2X.sup.3X.sup.4IX.sup.5CX.sup.6GSKQCYX.sup.7PCKX.sup.8X.sup.-
9TGCX.sup.10X.sup.11X.sup.12KCX.sup.13X.sup.14
KX.sup.15CKCYGCG,
wherein X.sup.1, X.sup.2, X.sup.3, X.sup.4, X.sup.5, X.sup.6,
X.sup.7, X.sup.8, X.sup.9, X.sup.10, X.sup.11, X.sup.12, X.sup.13,
X.sup.14, and X.sup.15 are each individually any amino acid or
amino acid analogue or null. In some cases, the peptide comprises
the sequence
TABLE-US-00012 (SEQ ID NO: 4)
GSX.sup.1X.sup.2X.sup.3X.sup.4IX.sup.5CX.sup.6GSKQCYX.sup.7PCKX.sup.8X.su-
p.9TGCX.sup.10X.sup.11X.sup.12KCX.sup.13 X.sup.14KX.sup.15CKCYGCG
or (SEQ ID NO: 278)
X.sup.1X.sup.2X.sup.3X.sup.4IX.sup.5CX.sup.6GSKQCYX.sup.7PCKX.sup.8X.sup.-
9TGCX.sup.10X.sup.11X.sup.12KCX.sup.13X.sup.14
KX.sup.15CKCYGCG,
[0072] where X.sup.1 is selected from G or null, wherein X.sup.2 is
selected from S or null, wherein X.sup.3 is selected from E, G or
null, wherein X.sup.4 is selected from V, S, or null, wherein
X.sup.5 is selected from R or S, wherein X.sup.6 is selected from S
or T, wherein X.sup.7 is selected from G or D, wherein X.sup.8 is
selected from Q or R, wherein X.sup.9 is selected from Q or K,
wherein X.sup.10 is selected from T or P, wherein X.sup.11 is
selected from N or Q, wherein X.sup.12 is selected from S or A,
wherein X.sup.13 is selected from M or L, wherein X.sup.14 is
selected from N or Q, and wherein X.sup.15 is selected from V or
S.
[0073] In some embodiments, a peptide comprises the sequence
TABLE-US-00013 (SEQ ID NO: 5)
GSX.sup.1X.sup.2X.sup.3VX.sup.4IX.sup.5VX.sup.6CX.sup.7X.sup.8SX.sup.9X.s-
up.10CLX.sup.11PCKX.sup.12AGMRFGKCX.sup.13
NX.sup.14KCX.sup.15CTPX.sup.16 or (SEQ ID NO: 279)
X.sup.1X.sup.2X.sup.3VX.sup.4IX.sup.5VX.sup.6CX.sup.7X.sup.85X.sup.9X.sup-
.10CLX.sup.11PCKX.sup.12AGMRFGKCX.sup.13N
X.sup.14KCX.sup.15CTPX.sup.16,
wherein X.sup.1, X.sup.2, X.sup.3, X.sup.4, X.sup.5, X.sup.6,
X.sup.7, X.sup.8, X.sup.9, X.sup.10, X.sup.11, X.sup.12, X.sup.13,
X.sup.14, X.sup.15, X.sup.16 are each individually any amino acid
or amino acid analogue or null. In some cases, the peptide
comprises the sequence
TABLE-US-00014 (SEQ ID NO: 6)
GSX.sup.1X.sup.2X.sup.3VX.sup.4IX.sup.5VX.sup.6CX.sup.7X.sup.8SX.sup.9X.s-
up.10CLX.sup.11PCKX.sup.12AGMRFGKCX.sup.13
NX.sup.14KCX.sup.15CTPX.sup.16 or (SEQ ID NO: 280)
X.sup.1X.sup.2X.sup.3VX.sup.4IX.sup.5VX.sup.6CX.sup.7X.sup.8SX.sup.9X.sup-
.10CLX.sup.11PCKX.sup.12AGMRFGKCX.sup.13
NX.sup.14KCX.sup.15CTPX.sup.16,
where X.sup.1 is selected from G or null, wherein X.sup.2 is
selected from G, S or null, wherein X.sup.3 is selected from G, S
or null, wherein X.sup.4 is selected from P or R, wherein X.sup.5
is selected from N or P, wherein X.sup.6 is selected from K or S,
wherein X.sup.7 is selected from R or K, wherein X.sup.8 is
selected from G or H, wherein X.sup.9 is selected from R or G,
wherein X.sup.10 is selected from D or Q, wherein X.sup.11 is
selected from D or K, wherein X.sup.12 is selected from K or D,
wherein X.sup.13 is selected from I or M, wherein X.sup.14 is
selected from S or G, wherein X.sup.15 is selected from H or D, and
wherein X.sup.16 is selected from K or null.
[0074] In some embodiments, a peptide comprises the sequence
TABLE-US-00015 (SEQ ID NO: 7) GSXVXVKCXGSKQCXPCKRXGXRXGKCINKKXCKCYX
or (SEQ ID NO: 8) GSXGCVXKCRPGXKXCCXPXKRCSRRFGXKKCKX,
wherein each letter is each individually any amino acid or amino
acid analogue and where X is no amino acid or a 1-10 amino acid
long peptide fragment wherein each amino acid within such peptide
fragment can in each case be any amino acid or amino acid analogue.
In some embodiments, a peptide comprises
TABLE-US-00016 (SEQ ID NO: 281) XVXVKCXGSKQCXPCKRXGXRXGKCINKKXCKCYX
or (SEQ ID NO: 282) XGCVXKCRPGXKXCCXPXKRCSRRFGXKKCKX,
the sequence wherein each letter is each individually any amino
acid or amino acid analogue and where X is no amino acid or a 1-10
amino acid long peptide fragment wherein each amino acid within
such peptide fragment can in each case be any amino acid or amino
acid analogue.
[0075] In some embodiments, a peptide comprises the sequence
TABLE-US-00017 (SEQ ID NO: 11)
GSGVX.sup.1IX.sup.2X.sup.3RCX.sup.4GSRQCX.sup.5DPCRX.sup.6X.sup.7X.sup.8G-
X.sup.9RX.sup.10GRCX.sup.11 NRRCRCX.sup.12X.sup.13X.sup.14X.sup.15
or (SEQ ID NO: 285)
GVX.sup.1IX.sup.2X.sup.3RCX.sup.4GSRQCX.sup.5DPCRX.sup.6X.sup.7X.sup.8GX.-
sup.9RX.sup.10GRCX.sup.11
NRRCRCX.sup.12X.sup.13X.sup.14X.sup.15,
wherein X.sup.1, X.sup.2, X.sup.3, X.sup.4, X.sup.5, X.sup.6,
X.sup.7, X.sup.8, X.sup.9, X.sup.11, X.sup.12, X.sup.13, X.sup.14,
and X.sup.15 are each individually any amino acid or amino acid
analogue or null. In some cases, the peptide comprises the
sequence
TABLE-US-00018 (SEQ ID NO: 12)
GSGVX.sup.1IX.sup.2X.sup.3RCX.sup.4GSRQCX.sup.5DPCRX.sup.6X.sup.7X.sup.8G-
X.sup.9RX.sup.10GRCX.sup.11 NRRCRCX.sup.12X.sup.13X.sup.14X.sup.15
or (SEQ ID NO: 286)
GVX.sup.1IX.sup.2X.sup.3RCX.sup.4GSRQCX.sup.5DPCRX.sup.6X.sup.7X.sup.8GX.-
sup.9RX.sup.10GRCX.sup.11
NRRCRCX.sup.12X.sup.13X.sup.14X.sup.15,
[0076] where X.sup.1 is selected from P or R, wherein X.sup.2 is
selected from P or N, wherein X.sup.3 is selected from V or I,
wherein X.sup.4 is selected from S, T, R or K, wherein X.sup.5 is
selected from Y or L, wherein X.sup.6 is selected from Q, R or K,
wherein X.sup.7 is selected from A, K or R, wherein X.sup.8 is
selected from T or A, wherein X.sup.9 is selected from C or M,
wherein X.sup.10 is selected from F or N, wherein X.sup.11 is
selected from M or I, wherein X.sup.12 is selected from Y or T,
wherein X.sup.13 is selected from G or P, wherein X.sup.14 is
selected from C or null, and wherein X.sup.15 is selected from G or
null.
[0077] In some embodiments, a peptide comprises the sequence
TABLE-US-00019 (SEQ ID NO: 13)
GSX.sup.1X.sup.2X.sup.3X.sup.4IX.sup.5CX.sup.6GSRQCYX.sup.7PCRX.sup.8X.su-
p.9TGCX.sup.10X.sup.11X.sup.l2RCX.sup.13 X.sup.14RX.sup.15CRCYGCG
or (SEQ ID NO: 287)
X.sup.1X.sup.2X.sup.3X.sup.4IX.sup.5CX.sup.6GSRQCYX.sup.7PCRX.sup.8X.sup.-
9TGCX.sup.10X.sup.11X.sup.12RCX.sup.13X.sup.14
RX.sup.15CRCYGCG,
wherein X.sup.1, X.sup.2, X.sub.3, X.sub.4, X.sub.5, X.sub.6,
X.sub.7, X.sub.8, X.sub.9, X.sub.10, X.sub.11, X.sub.12, X.sub.13,
X.sub.14, and X.sup.15 are each individually any amino acid or
amino acid analogue or null. In some cases, the peptide comprises
the sequence
TABLE-US-00020 (SEQ ID NO: 14)
GSX.sup.1X.sup.2X.sup.3X.sup.4IX.sup.5CX.sup.6GSRQCYX.sup.7PCRX.sup.8X.su-
p.9TGCX.sup.10X.sup.11X.sup.12RCX.sup.13 X.sup.14RX.sup.15CRCYGCG,
or (SEQ ID NO: 288)
X.sup.1X.sup.2X.sup.3X.sup.4IX.sup.5CX.sup.6GSRQCYX.sup.7PCRX.sup.8X.sup.-
9TGCX.sup.10X.sup.11X.sup.12RCX.sup.13X.sup.14
RX.sup.15CRCYGCG,
[0078] where X.sup.1 is selected from G or null, wherein X.sup.2 is
selected from S or null, wherein X.sup.3 is selected from E, G or
null, wherein X.sup.4 is selected from V, S, or null, wherein
X.sup.5 is selected from R or S, wherein X.sup.6 is selected from S
or T, wherein X.sup.7 is selected from G or D, wherein X.sup.8 is
selected from Q or R, wherein X.sup.9 is selected from Q, R, or K,
wherein X.sup.10 is selected from T or P, wherein X.sup.11 is
selected from N or Q, wherein X.sup.12 is selected from S or A,
wherein X.sup.13 is selected from M or L, wherein X.sup.14 is
selected from N or Q, and wherein X.sup.15 is selected from V or
S.
[0079] In some embodiments, a peptide comprises the sequence
TABLE-US-00021 (SEQ ID NO: 15)
GSX.sup.1X.sup.2X.sup.3VX.sup.4IX.sup.5VX.sup.6CX.sup.7X.sup.8SX.sup.9X.s-
up.10CLX.sup.11PCRX.sup.12AGMRFGRCX.sup.13
NX.sup.14RCX.sup.15CTPX.sup.16 or (SEQ ID NO: 289)
X.sup.1X.sup.2X.sup.3VX.sup.4IX.sup.5VX.sup.6CX.sup.7X.sup.8SX.sup.9X.sup-
.10CLX.sup.11PCRX.sup.12AGMRFGRCX.sup.13
NX.sup.14RCX.sup.15CTPX.sup.16,
wherein X.sup.1, X.sup.2, X.sup.3, X.sup.4, X.sup.5, X.sup.6,
X.sup.7, X.sup.8, X.sup.9, X.sup.10, X.sup.11, X.sup.12, X.sup.13,
X.sup.14, X.sup.15, X.sup.16 are each individually any amino acid
or amino acid analogue or null. In some cases, the peptide
comprises the sequence
TABLE-US-00022 (SEQ ID NO: 16)
GSX.sup.1X.sup.2X.sup.3VX.sup.4IX.sup.5VX.sup.6CX.sup.7X.sup.8SX.sup.9X.s-
up.10CLX.sup.11PCRX.sup.12AGMRFGRCX.sup.13
NX.sup.14RCX.sup.15CTPX.sup.16 or (SEQ ID NO: 290)
X.sup.1X.sup.2X.sup.3VX.sup.4IX.sup.5VX.sup.6CX.sup.7X.sup.8SX.sup.9X.sup-
.10CLX.sup.11PCRX.sup.12AGMRFGRCX.sup.13
NX.sup.14RCX.sup.15CTPX.sup.16,
where X.sup.1 is selected from G or null, wherein X.sup.2 is
selected from G, S or null, wherein X.sup.3 is selected from G, S
or null, wherein X.sup.4 is selected from P or R, wherein X.sup.5
is selected from N or P, wherein X.sup.6 is selected from R, K or
S, wherein X.sup.7 is selected from R or K, wherein X.sup.8 is
selected from G or H, wherein X.sup.9 is selected from R or G,
wherein X.sup.10 is selected from D or Q, wherein X.sup.11 is
selected from D, R, or K, wherein X.sup.12 is selected from K, R,
or D, wherein X.sup.13 is selected from I or M, wherein X.sup.14 is
selected from S or G, wherein X.sup.15 is selected from H or D, and
wherein X.sup.16 is selected from K, R, or null.
[0080] In some embodiments, a peptide comprises the sequence
TABLE-US-00023 (SEQ ID NO: 17)
GSXVXVRCXGSRQCXPCRRXGXRXGRCINRRXCRCYX or (SEQ ID NO: 18)
GSXGCVXRCRPGXRXCCXPXRRCSRRFGXRRCRX,
wherein each letter is each individually any amino acid or amino
acid analogue and where X is no amino acid or a 1-10 amino acid
long peptide fragment wherein each amino acid within such peptide
fragment can in each case be any amino acid or amino acid analogue.
In some embodiments, a peptide comprises the sequence
TABLE-US-00024 (SEQ ID NO: 291) XVXVRCXGSRQCXPCRRXGXRXGRCINRRXCRCYX
or (SEQ ID NO: 292) XGCVXRCRPGXRXCCXPXRRCSRRFGXRRCRX,
wherein each letter is each individually any amino acid or amino
acid analogue and where X is no amino acid or a 1-10 amino acid
long peptide fragment wherein each amino acid within such peptide
fragment can in each case be any amino acid or amino acid
analogue.
[0081] In some embodiments, a peptide comprises the sequence
TABLE-US-00025 (SEQ ID NO: 19)
GSXVXXXVRCXGSRQCXXPCRRXXGXRXGRCINRRXCRCYXXX, (SEQ ID NO: 293)
XVXXXVRCXGSRQCXXPCRRXXGXRXGRCINRRXCRCYXXX, (SEQ ID NO: 20)
GSXXXGCVXXXXRCRPGXRXCCXPXRRCSRRFGXXXXRRCRXXXXXX, or (SEQ ID NO:
294) XXXGCVXXXXRCRPGXRXCCXPXRRCSRRFGXXXXRRCRXXXXXX
wherein X is no amino acid or any amino acid analogue.
[0082] In some embodiments, a peptide comprises the one or more of
the following peptide fragments:
TABLE-US-00026 (SEQ ID NO: 312) GKCMNGKC; (SEQ ID NO: 313)
GRCMNGRC; (SEQ ID NO: 298) GKCINKKCKC; (SEQ ID NO: 299) KCIN; (SEQ
ID NO: 300) KKCK; (SEQ ID NO: 301) PCKR; (SEQ ID NO: 302) KRCSRR;
(SEQ ID NO: 303) KQC; (SEQ ID NO: 304) GRCINRRCRC; (SEQ ID NO: 305)
RCIN; (SEQ ID NO: 306) RRCR; (SEQ ID NO: 307) PCRR; (SEQ ID NO:
308) RRCSRR; (SEQ ID NO: 309) RQC; (SEQ ID NO: 310) PCKK, and (SEQ
ID NO: 311) KKCSKK.
[0083] TABLE 1 lists some exemplary peptides according to the
present disclosure.
TABLE-US-00027 TABLE 1 Exemplary Amino Acid Sequences SEQ ID NO:
Amino Acid Sequence SEQ ID NO: 24
GSGIVCKVCKIICGMQGKKVNICKAPIKCKCKKG SEQ ID NO: 25
GSSEKDCIKHLQRCRENKDCCSKKCSRRGTNPEKRCR SEQ ID NO: 26
GSVRIPVSCKHSGQCLKPCKDAGMRFGKCMNGKCDCTPK SEQ ID NO: 27
GSGVPINVKCRGSRDCLDPCKKAGMRFGKCINSKCHCTP SEQ ID NO: 28
GSAVCVYRTCDKDCKRRGYRSGKCINNACKCYPYG SEQ ID NO: 29
GSISCTGSKQCYDPCKRKTGCPNAKCMNKSCKCYGCG SEQ ID NO: 30
GSQVQTNVKCQGGSCASVCRREIGVAAGKCINGKCVCYRN SEQ ID NO: 31
GSEVIRCSGSKQCYGPCKQQTGCTNSKCMNKVCKCYGCG SEQ ID NO: 32
GSACKGVFDACTPGKNECCPNRVCSDKHKWCKWKL SEQ ID NO: 33
GSQIYTSKECNGSSECYSHCEGITGKRSGKCINKKCYCYR SEQ ID NO: 34
GSGCLEFWWKCNPNDDKCCRPKLKCSKLFKLCNFSFG SEQ ID NO: 35
GSDCVRFWGKCSQTSDCCPHLACKSKWPRNICVWDGSVG SEQ ID NO: 36
GSGCFGYKCDYYKGCCSGYVCSPTWKWCVRPGPGR SEQ ID NO: 37
GSMNAKFILLLVLTTMMLLPDTKGAEVIRCSGSKQCYGPCKQQTGCTNSKC MNKVCKCYGCG SEQ
ID NO: 38 GSMNAKLIYLLLVVTTMTLMFDTAQAVDIMCSGPKQCYGPCKKETGCPNA
KCMNRRCKCYGCV SEQ ID NO: 39
GSMNAKLIYLLLVVTTMMLTFDTTQAGDIKCSGTRQCWGPCKKQTTCTNS KCMNGKCKCYGCVG
SEQ ID NO: 40 GSMNTKFIFLLLVVTNTMMLFDTKPVEGISCTGSKQCYDPCKRKTGCPNAK
CMNKSCKCYGCG SEQ ID NO: 41 GSGVPINVKCSGSRDCLEPCKKAGMRFGKCINRKCHCTPK
SEQ ID NO: 42 GSGVPINVKCTGSPQCLKPCKDAGMRFGKCINGKCHCTPK SEQ ID NO:
43 GSGVIINVKCKISRQCLEPCKKAGMRFGKCMNGKCHCTPK SEQ ID NO: 44
GSGVPINVKCRGSPQCIQPCRDAGMRFGKCMNGKCHCTPQ SEQ ID NO: 45
GSGVEINVKCTGSHQCIKPCKDAGMRFGKCINRKCHCTPK SEQ ID NO: 46
GSGVEINVKCSGSPQCLKPCKDAGMRFGKCMNRKCHCTPK SEQ ID NO: 47
GSGVPTDVKCRGSPQCIQPCKDAGMRFGKCMNGKCHCTPK SEQ ID NO: 48
GSGVPINVSCTGSPQCIKPCKDAGMRFGKCMNRKCHCTPK SEQ ID NO: 49
GSGVPINVPCTGSPQCIKPCKDAGMRFGKCMNRKCHCTPK SEQ ID NO: 50
GSVGINVKCKHSGQCLKPCKDAGMRFGKCINGKCDCTPK SEQ ID NO: 51
GSVGINVKCKHSGQCLKPCKDAGMRFGKCMNGKCDCTPK SEQ ID NO: 52
GSVGIPVSCKHSGQCIKPCKDAGMRFGKCMNRKCDCTPK SEQ ID NO: 53
GSRKGCFKEGHSCPKTAPCCRPLVCKGPSPNTKKCTRP SEQ ID NO: 54
GSSFCIPFKPCKSDENCCKKFKCKTTGIVKLCRW SEQ ID NO: 55
GSLKGCLPRNRFCNALSGPRCCSGLRCKELSIWASKCL SEQ ID NO: 56
GSGNYCLRGRCLPGGRKCCNGRPCECFAKICSCKPK SEQ ID NO: 57
GSTVKCGGCNRKCCPGGCRSGKCINGKCQCY SEQ ID NO: 58
GSGCMKEYCAGQCRGKVSQDYCLKHCKCIPR SEQ ID NO: 59
GSACLGFGEKCNPSNDKCCKSSSLVCSQKHKWCKYG SEQ ID NO: 60
GSRGGCLPHNRFCNALSGPRCCSGLRCKELSIRDSRCLG SEQ ID NO: 61
GSRGGCLPRNKFCNPSSGPRCCSGLTCKELNIWASKCL SEQ ID NO: 62
GSQRSCAKPGDMCMGIKCCDGQCGCNRGTGRCFCK SEQ ID NO: 63
GSARGCADAYKSCNHPRTCCDGYNGYKRACICSGSNCKCKKS SEQ ID NO: 64
GSRGGCLPHNRFCNALSGPRCCSGLRCKELSIWDSRCLG SEQ ID NO: 65
GSRGGCLPHNRFCNALSGPRCCSGLKCKELSIYDSRCLG SEQ ID NO: 66
GSRGGCLPHNRFCNALSGPRCCSRLKCKELSIWDSRCLG SEQ ID NO: 67
GSRGGCLPHNRFCNALTGPRCCSRLRCKELSIWDSICLG SEQ ID NO: 68
GSSCADAYKSCDSLKCCNNRTCMCSMIGTNCTCRKK SEQ ID NO: 69
GSERRCLPAGKTCVRGPMRVPCCGSCSQNKCT SEQ ID NO: 70
GSLCSREGEFCYKLRKCCAGFYCKAFVLHCYRN SEQ ID NO: 71
GSACGSCRKKCKGSGKCINGRCKCY SEQ ID NO: 72 GSACGSCRKKCKGPGKCINGRCKCY
SEQ ID NO: 73 GSACQGYMRKCGRDKPPCCKKLECSKTWRWCVWN SEQ ID NO: 74
GSGRYCQKWMWTCDSKRACCEGLRCKLWCRKI SEQ ID NO: 75
GSNAKCRGSPECLPKCKEAIGKAAGKCMNGKCKCYP SEQ ID NO: 76
GSNVKCRGSKECLPACKAAVGKAAGKCMNGKCKCYP SEQ ID NO: 77
GSNVKCRGSPECLPKCKEAIGKSAGKCMNGKCKCYP SEQ ID NO: 78
GSNAKCRGSPECLPKCKQAIGKAAGKCMNGKCKCYP SEQ ID NO: 79
GSRGYCAEKGIKCHNIHCCSGLTCKCKGSSCVCRK SEQ ID NO: 80
GSERGCKLTFWKCKNKKECCGWNACALGICMPR SEQ ID NO: 81
GSKKKCIAKDYGRCKWGGTPCCRGRGCICSIMGTNCECKPR SEQ ID NO: 82
GSGCKLTFWKCKNKKECCGWNACALGICMPR SEQ ID NO: 83
GSACKGLFVTCTPGKDECCPNHVCSSKHKWCKYK SEQ ID NO: 84
GSIACAPRGLLCFRDKECCKGLTCKGRFVNTWPTFCLV SEQ ID NO: 85
GSACAGLYKKCGKGVNTCCENRPCKCDLAMGNCICKKK SEQ ID NO: 86
GSFTCAISCDIKVNGKPCKGSGEKKCSGGWSCKFNVCVKV SEQ ID NO: 87
GSGFCAQKGIKCHDIHCCTNLKCVREGSNRVCRKA SEQ ID NO: 88
GSCAKKRNWCGKNEDCCCPMKCIYAWYNQQGSCQSTITGLFKKC SEQ ID NO: 89
GSYCQKWMWTCDSARKCCEGLVCRLWCKKI SEQ ID NO: 90
GSRGGCLPHNKFCNALSGPRCCSGLKCKELTIWNTKCLE SEQ ID NO: 91
GSNVKCTGSKQCLPACKAAVGKAAGKCMNGKCKCYT SEQ ID NO: 92
GSQRSCAKPGEMCMRIKCCDGQCGCNRGTGRCFCK SEQ ID NO: 93
GSGCIPKHKRCTWSGPKCCNNISCHCNISGTLCKCRPG SEQ ID NO: 94
GSNYCVAKRCRPGGRQCCSGKPCACVGKVCKCPRD SEQ ID NO: 95
GSERGCSGAYKRCSSSQRCCEGRPCVCSAINSNCKCRKT SEQ ID NO: 96
GSRYCPRNPEACYNYCLRTGRPGGYCGGRSRITCFCFR SEQ ID NO: 97
GSQRSCAKPGEMCMGIKCCDGQCGCNRGTGRCFCK SEQ ID NO: 98
GSRRGCFKEGKWCPKSAPCCAPLKCKGPSIKQQKCVRE SEQ ID NO: 99
GSTVKCGGCNRKCCAGGCRSGKCINGKCQCYGR SEQ ID NO: 100
GSERRCEPSGKPCRPLMRIPCCGSCVRGKCA SEQ ID NO: 101
GSRGGCLPRNKFCNPSSGPRCCSGLTCKELNIWANKCL SEQ ID NO: 102
GSCAKKRNWCGKNEDCCCPMKCIYAWYNQQGSCQTTITGLFKKC SEQ ID NO: 103
GSVRIPVSCKHSGQCLKPCKDAGMRTGKCMNGKCDCTPK SEQ ID NO: 104
GSVKCTTSKDCWPPCKKVTGRA SEQ ID NO: 105
GSGIVCRVCRIICGMQGRRVNICRAPIRCRCRRG SEQ ID NO: 106
GSSERDCIRHLQRCRENRDCCSRRCSRRGTNPERRCR SEQ ID NO: 107
GSVRIPVSCRHSGQCLRPCRDAGMRFGRCMNGRCDCTPR SEQ ID NO: 108
GSGVPINVRCRGSRDCLDPCRRAGMRFGRCINSRCHCTP SEQ ID NO: 109
GSAVCVYRTCDRDCRRRGYRSGRCINNACRCYPYG SEQ ID NO: 110
GSISCTGSRQCYDPCRRRTGCPNARCMNRSCRCYGCG SEQ ID NO: 111
GSQVQTNVRCQGGSCASVCRREIGVAAGRCINGRCVCYRN SEQ ID NO: 112
GSEVIRCSGSRQCYGPCRQQTGCTNSRCMNRVCRCYGCG SEQ ID NO: 113
GSACRGVFDACTPGRNECCPNRVCSDRHRWCRWRL SEQ ID NO: 114
GSQIYTSRECNGSSECYSHCEGITGRRSGRCINRRCYCYR SEQ ID NO: 115
GSGCLEFWWRCNPNDDRCCRPRLRCSRLFRLCNFSFG SEQ ID NO: 116
GSDCVRFWGRCSQTSDCCPHLACRSRWPRNICVWDGSVG SEQ ID NO: 117
GSGCFGYRCDYYRGCCSGYVCSPTWRWCVRPGPGR SEQ ID NO: 118
GSMNARFILLLVLTTMMLLPDTRGAEVIRCSGSRQCYGPCRQQTGCTNSRC MNRVCRCYGCG SEQ
ID NO: 119 GSMNARLIYLLLVVTTMTLMFDTAQAVDIMCSGPRQCYGPCRRETGCPNA
RCMNRRCRCYGCV SEQ ID NO: 120
GSMNARLIYLLLVVTTMMLTFDTTQAGDIRCSGTRQCWGPCRRQTTCTNSR CMNGRCRCYGCVG
SEQ ID NO: 121 GSMNTRFIFLLLVVTNTMMLFDTRPVEGISCTGSRQCYDPCRRRTGCPNARC
MNRSCRCYGCG SEQ ID NO: 122 GSGVPINVRCSGSRDCLEPCRRAGMRFGRCINRRCHCTPR
SEQ ID NO: 123 GSGVPINVRCTGSPQCLRPCRDAGMRFGRCINGRCHCTPR SEQ ID NO:
124 GSGVIINVRCRISRQCLEPCRRAGMRFGRCMNGRCHCTPR SEQ ID NO: 125
GSGVPINVRCRGSPQCIQPCRDAGMRFGRCMNGRCHCTPQ SEQ ID NO: 126
GSGVEINVRCTGSHQCIRPCRDAGMRFGRCINRRCHCTPR SEQ ID NO: 127
GSGVEINVRCSGSPQCLRPCRDAGMRFGRCMNRRCHCTPR SEQ ID NO: 128
GSGVPTDVRCRGSPQCIQPCRDAGMRFGRCMNGRCHCTPR SEQ ID NO: 129
GSGVPINVSCTGSPQCIRPCRDAGMRFGRCMNRRCHCTPR SEQ ID NO: 130
GSGVPINVPCTGSPQCIRPCRDAGMRFGRCMNRRCHCTPR SEQ ID NO: 131
GSVGINVRCRHSGQCLRPCRDAGMRFGRCINGRCDCTPR SEQ ID NO: 132
GSVGINVRCRHSGQCLRPCRDAGMRFGRCMNGRCDCTPR SEQ ID NO: 133
GSVGIPVSCRHSGQCIRPCRDAGMRFGRCMNRRCDCTPR SEQ ID NO: 134
GSRRGCFREGHSCPRTAPCCRPLVCRGPSPNTRRCTRP SEQ ID NO: 135
GSSFCIPFRPCRSDENCCRRFRCRTTGIVRLCRW SEQ ID NO: 136
GSLRGCLPRNRFCNALSGPRCCSGLRCRELSIWASRCL SEQ ID NO: 137
GSGNYCLRGRCLPGGRRCCNGRPCECFARICSCRPR SEQ ID NO: 138
GSTVRCGGCNRRCCPGGCRSGRCINGRCQCY SEQ ID NO: 139
GSGCMREYCAGQCRGRVSQDYCLRHCRCIPR SEQ ID NO: 140
GSACLGFGERCNPSNDRCCRSSSLVCSQRHRWCRYG SEQ ID NO: 141
GSRGGCLPHNRFCNALSGPRCCSGLRCRELSIRDSRCLG SEQ ID NO: 142
GSRGGCLPRNRFCNPSSGPRCCSGLTCRELNIWASRCL
SEQ ID NO: 143 GSQRSCARPGDMCMGIRCCDGQCGCNRGTGRCFCR SEQ ID NO: 144
GSARGCADAYRSCNHPRTCCDGYNGYRRACICSGSNCRCRRS SEQ ID NO: 145
GSRGGCLPHNRFCNALSGPRCCSGLRCRELSIWDSRCLG SEQ ID NO: 146
GSRGGCLPHNRFCNALSGPRCCSGLRCRELSIYDSRCLG SEQ ID NO: 147
GSRGGCLPHNRFCNALSGPRCCSRLRCRELSIWDSRCLG SEQ ID NO: 148
GSRGGCLPHNRFCNALTGPRCCSRLRCRELSIWDSICLG SEQ ID NO: 149
GSSCADAYKSCDSLRCCNNRTCMCSMIGTNCTCRRR SEQ ID NO: 150
GSERRCLPAGRTCVRGPMRVPCCGSCSQNRCT SEQ ID NO: 151
GSLCSREGEFCYRLRRCCAGFYCRAFVLHCYRN SEQ ID NO: 152
GSACGSCRRRCRGSGRCINGRCRCY SEQ ID NO: 153 GSACGSCRRRCRGPGRCINGRCRCY
SEQ ID NO: 154 GSACQGYMRRCGRDRPPCCRRLECSRTWRWCVWN SEQ ID NO: 155
GSGRYCQRWMWTCDSRRACCEGLRCRLWCRRI SEQ ID NO: 156
GSNARCRGSPECLPRCREAIGRAAGRCMNGRCRCYP SEQ ID NO: 157
GSNVRCRGSRECLPACRAAVGRAAGRCMNGRCRCYP SEQ ID NO: 158
GSNVRCRGSPECLPRCREAIGRSAGRCMNGRCRCYP SEQ ID NO: 159
GSNARCRGSPECLPRCRQAIGRAAGRCMNGRCRCYP SEQ ID NO: 160
GSRGYCAERGIRCHNIHCCSGLTCRCRGSSCVCRR SEQ ID NO: 161
GSERGCRLTFWRCRNRRECCGWNACALGICMPR SEQ ID NO: 162
GSRRRCIARDYGRCRWGGTPCCRGRGCICSIMGTNCECRPR SEQ ID NO: 163
GSGCRLTFWRCRNRRECCGWNACALGICMPR SEQ ID NO: 164
GSACRGLFVTCTPGRDECCPNHVCSSRHRWCRYR SEQ ID NO: 165
GSIACAPRGLLCFRDRECCRGLTCRGRFVNTWPTFCLV SEQ ID NO: 166
GSACAGLYRRCGRGVNTCCENRPCRCDLAMGNCICRRR SEQ ID NO: 167
GSFTCAISCDIRVNGRPCRGSGERRCSGGWSCRFNVCVRV SEQ ID NO: 168
GSGFCAQRGIRCHDIHCCTNLRCVREGSNRVCRRA SEQ ID NO: 169
GSCARRRNWCGRNEDCCCPMRCIYAWYNQQGSCQSTITGLFRRC SEQ ID NO: 170
GSYCQRWMWTCDSARRCCEGLVCRLWCRRI SEQ ID NO: 171
GSRGGCLPHNRFCNALSGPRCCSGLRCRELTIWNTRCLE SEQ ID NO: 172
GSNVRCTGSRQCLPACRAAVGRAAGRCMNGRCRCYT SEQ ID NO: 173
GSQRSCARPGEMCMRIRCCDGQCGCNRGTGRCFCR SEQ ID NO: 174
GSGCIPRHRRCTWSGPRCCNNISCHCNISGTLCRCRPG SEQ ID NO: 175
GSNYCVARRCRPGGRQCCSGRPCACVGRVCRCPRD SEQ ID NO: 176
GSERGCSGAYRRCSSSQRCCEGRPCVCSAINSNCRCRRT SEQ ID NO: 177
GSQRSCARPGEMCMGIRCCDGQCGCNRGTGRCFCR SEQ ID NO: 178
GSRRGCFREGRWCPRSAPCCAPLRCRGPSIRQQRCVRE SEQ ID NO: 179
GSTVRCGGCNRRCCAGGCRSGRCINGRCQCYGR SEQ ID NO: 180
GSERRCEPSGRPCRPLMRIPCCGSCVRGRCA SEQ ID NO: 181
GSRGGCLPRNRFCNPSSGPRCCSGLTCRELNIWANRCL SEQ ID NO: 182
GSCARRRNWCGRNEDCCCPMRCIYAWYNQQGSCQTTITGLFRRC SEQ ID NO: 183
GSVRIPVSCRHSGQCLRPCRDAGMRTGRCMNGRCDCTPR SEQ ID NO: 184
GSQKILSNRCNNSSECIPHCIRIFGTRAAKCINRKCYCYP SEQ ID NO: 185
GSAVCNLKRCQLSCRSLGLLGKCIGDKCECVKHG SEQ ID NO: 186
GSISIGIRCSPSIDLCEGQCRIRRYFTGYCSGDTCHCSG SEQ ID NO: 187
GSGDCLPHLRRCRENNDCCSRRCRRRGANPERRCR SEQ ID NO: 188
GSSCEPGRTFRDRCNTCKCGADGRSAACTLRACPNQ SEQ ID NO: 189
GSGDCLPHLKRCKADNDCCGKKCKRRGTNAEKRCR SEQ ID NO: 190
GSGDCLPHLKRCKENNDCCSKKCKRRGTNPEKRCR SEQ ID NO: 191
GSKDCLKKLKLCKENKDCCSKSCKRRGTNIEKRCR SEQ ID NO: 192
GSGDCLPHLKRCKENNDCCSKKCKRRGANPEKRCR SEQ ID NO: 193
GSVFINVKCRGSPECLPKCKEAIGKSAGKCMNGKCKCYP SEQ ID NO: 194
GSVFINAKCRGSPECLPKCKEAIGKAAGKCMNGKCKCYP SEQ ID NO: 195
GSVIINVKCKISRQCLEPCKKAGMRFGKCMNGKCHCTP SEQ ID NO: 196
GSVPTDVKCRGSPQCIQPCKDAGMRFGKCMNGKCHCTP SEQ ID NO: 197
GSVRIPVSCKHSGQCLKPCKDAGMRFGKCMNGKCDCTP SEQ ID NO: 198
GSVRIPVSCRHSGQCLRPCRDAGMRFGRCMNGRCDCTP SEQ ID NO: 199
GSTNVSCTTSKECWSVCQRLHNTSRGKCMNKKCRC SEQ ID NO: 200
GSNVKCTGSKQCLPACKAAVGKAAGKCMNGKCKC SEQ ID NO: 201
GSGVPINVRCRGSRDCLDPCRGAGERHGRCGNSRCHCTP SEQ ID NO: 202
GSVRIPVSCRHSGQCLRPCRDAGERHGRCGGGRCDCTPR SEQ ID NO: 203
GSQVQTNVRCQGGSCGSVCRREGGGAGGGCGNGRCGCYRN SEQ ID NO: 204
GSIKCSESYQCFPVCKSRFGKTNGRCVNGFCDCF SEQ ID NO: 205
GSVKCSSPQQCLKPCKAAFGISAGGKCINGKCKCY SEQ ID NO: 206
GSVSCSASSQCWPVCKKLFGTYRGKCMNSKCRCY SEQ ID NO: 207
GSESCTASNQCWSICKRLHNTNRGKCMNKKCRCY SEQ ID NO: 208
GSVSCTTSKECWSVCEKLYNTSRGKCMNKKCRCY SEQ ID NO: 209
GSMRCKSSKECLVKCKQATGRPNGKCMNRKCKCY SEQ ID NO: 210
GSIKCTLSKDCYSPCKKETGCPRAKCINRNCKCY SEQ ID NO: 211
GSIRCSGSRDCYSPCMKQTGCPNAKCINKSCKCY SEQ ID NO: 212
GSIRCSGTRECYAPCQKLTGCLNAKCMNKACKCY SEQ ID NO: 213
GSISCTNPKQCYPHCKKETGYPNAKCMNRKCKCF SEQ ID NO: 214
GSASCRTPKDCADPCRKETGCPYGKCMNRKCKCN SEQ ID NO: 215
GSTSCISPKQCTEPCRAKGCKHGKCMNRKCHCM SEQ ID NO: 216
GSKECTGPQHCTNFCRKN-KCTHGKCMNRKCKCF SEQ ID NO: 217
GSIKCRTPKDCADPCRKQTGCPHAKCMNKTCRCH SEQ ID NO: 218
GSVKCTTSKECWPPCKAATGKAAGKCMNKKCKCQ SEQ ID NO: 219
GSLECGASRECYDPCFKAFGRAHGKCMNNKCRCY SEQ ID NO: 220
GSEKCFATSQCWTPCKKAIGSLQSKCMNGKCKCY SEQ ID NO: 221
GSVRCYASRECWEPCRRVTGSAQAKCQNNQCRCY SEQ ID NO: 222
GSVKCSASRECWVACKKVTGSGQGKCQNNQCRCY SEQ ID NO: 223
GSVKCISSQECWIACKKVTGRFEGKCQNRQCRCY SEQ ID NO: 224
GSVRCYDSRQCWIACKKVTGSTQGKCQNKQCRCY SEQ ID NO: 225
GSVDCTVSKECWAPCKAAFGVDRGKCMGKKCKCY SEQ ID NO: 226
GSAKCRGSPECLPKCKEAIGKAAGKCMNGKCKCY SEQ ID NO: 227
GSKKCQGGSCASVCRRVIGVAAGKCINGRCVCY SEQ ID NO: 228
GSKKCSNTSQCYKTCEKVVGVAAGKCMNGKCICY SEQ ID NO: 229
GSVKCSGSSKCVKICIDRYNTRGAKCINGRCTCY SEQ ID NO: 230
GSNRCNNSSECIPHCIRIFGTRAAKCINRKCYCY SEQ ID NO: 231
GSKECNGSSECYSHCEGITGKRSGKCINKKCYCY SEQ ID NO: 232
GSAFCNLRRCELSCRSLGLLGKCIGEECKCV SEQ ID NO: 233
GSAVCNLKRCQLSCRSLGLLGKCIGDKCECV SEQ ID NO: 234
GSAACYSS-DCRVKCVAMGFSSGKCINSKCKCY SEQ ID NO: 235
GSAICATDADCSRKCPGNPPCRNGFCACT SEQ ID NO: 236
GSTECQIKNDCQRYCQSVKECKYGKCYCN SEQ ID NO: 237
GSTQCQSVRDCQQYCLTPDRCSYGTCYCK SEQ ID NO: 238
GSVSCRYGSDCAEPCKRLKCLLPSKCINGKCTCY SEQ ID NO: 239
GSIKCRYPADCHIMCRKVTGRAEGKCMNGKCTCY SEQ ID NO: 240
GSIKCSSSSSCYEPCRGVTGRAHGKCMNGRCTCY SEQ ID NO: 241
GSVKCTGSKQCLPACKAAVGKAAGKCMNGKCKCY SEQ ID NO: 242
GSVSCKHSGQCIKPCKDA-GMRFGKCMNRKCDCT SEQ ID NO: 243
GSVKCRGSPQCIQPCRDA-GMRFGKCMNGKCHCT SEQ ID NO: 244
GSVKCTSPKQCLPPCKAQFGIRAGAKCMNGKCKCY SEQ ID NO: 245
GSVKCTSPKQCSKPCKELYGSSAGAKCMNGKCKCY SEQ ID NO: 246
GSVKCTSPKQCLPPCKEIYGRHAGAKCMNGKCHCS SEQ ID NO: 247
GSVKCTGSKQCWPVCKQMFGKPNGKCMNGKCRCY SEQ ID NO: 248
GSVKCRGSRDCLDPCKKAGMRFGKCINSKCHCT SEQ ID NO: 249
GSVRCVTDDDCFRKCPGNPSCKRGFCACK SEQ ID NO: 250
GSVPCNNSRPCVPVCIREVNNKNGKCSNGKCLCY SEQ ID NO: 251
GSVPINVKCRGSRDCLDPCKKAGMRFGKCINSKCHCTP SEQ ID NO: 252
GSVQTNVKCQGGSCASVCRREIGVAAGKCINGKCVCYRN SEQ ID NO: 253
GSAEIIRCSGTRECYAPCQKLTGCLNAKCMNKACKCYGCV SEQ ID NO: 254
GSRPTDIKCSASYQCFPVCKSRFGKTNGRCVNGLCDCF SEQ ID NO: 255
GSQFTDVKCTGSKQCWPVCKQMFGKPNGKCMNGKCRCYS SEQ ID NO: 256
GSVGINVKCKHSRQCLKPCKDAGMRFGKCTNGKCHCTPK SEQ ID NO: 257
GSVVIGQRCYRSPDCYSACKKLVGKATGKCTNGRCDC SEQ ID NO: 258
GSNFKVEGACSKPCRKYCIDKGARNGKCINGRCHCYY SEQ ID NO: 259
GSQIDTNVKCSGSSKCVKICIDRYNTRGAKCINGRCTCYP SEQ ID NO: 260
GSGVPISVRCRGSRDCLEPCRRAGTRFGRCINGRCHCTP SEQ ID NO: 261
GSGVPISVRCRGSRDCLEPCRRAGTRFGRCIQSRCHCTP SEQ ID NO: 262
GSGVPISVRCRGSRDCLEPCRRAGTRFGRCINRRCHCTP SEQ ID NO: 263
GSGVPINVRCRGSRDCLEPCRRAGTRFGRCINSRCHCTP SEQ ID NO: 264
GSGVPINVRCRGSRDCLEPCRRAGTRFGRCIQSRCHCTP SEQ ID NO: 265
GSGVPINVRCRGSRDCLEPCRRAGTRFGRCIQSRCHCYP SEQ ID NO: 266
GSGVPINVRCRGSRDCYEPCRRAGTRFGRCIQSRCHCTP SEQ ID NO: 267
GSGVPINVRCRGSRDCLEPCRRAGTRFGRCIQSRCYCTP
SEQ ID NO: 268 GSGVPISVRCRGSRDCLEPCRRAGTRFGRCIQSRCHCYP SEQ ID NO:
269 GSGVPISVRCRGSRDCYEPCRRAGTRFGRCIQSRCHCTP SEQ ID NO: 270
GSGVPISVRCRGSRDCLEPCRRAGTRFGRCIQSRCYCTP SEQ ID NO: 271
GSGVPINVRCRGSRDCLEPCRRAGTRFGRCIASRCHCYP SEQ ID NO: 272
GSGVPINVRCRGSRDCLEPCRRAGTRFGRCISSRCHCYP SEQ ID NO: 273
GSGVPINVRCRGSRDCLEPCRRAGTRFGRCITSRCHCYP SEQ ID NO: 274
GSGVPINVRCRGSRDCLEPCRRAGTRFGRCINSRCHCYP SEQ ID NO: 314
GIVCKVCKIICGMQGKKVNICKAPIKCKCKKG SEQ ID NO: 315
SEKDCIKHLQRCRENKDCCSKKCSRRGTNPEKRCR SEQ ID NO: 316
VRIPVSCKHSGQCLKPCKDAGMRFGKCMNGKCDCTPK SEQ ID NO: 317
GVPINVKCRGSRDCLDPCKKAGMRFGKCINSKCHCTP SEQ ID NO: 318
AVCVYRTCDKDCKRRGYRSGKCINNACKCYPYG SEQ ID NO: 319
ISCTGSKQCYDPCKRKTGCPNAKCMNKSCKCYGCG SEQ ID NO: 320
QVQTNVKCQGGSCASVCRREIGVAAGKCINGKCVCYRN SEQ ID NO: 321
EVIRCSGSKQCYGPCKQQTGCTNSKCMNKVCKCYGCG SEQ ID NO: 322
ACKGVFDACTPGKNECCPNRVCSDKHKWCKWKL SEQ ID NO: 323
QIYTSKECNGSSECYSHCEGITGKRSGKCINKKCYCYR SEQ ID NO: 324
GCLEFWWKCNPNDDKCCRPKLKCSKLFKLCNFSFG SEQ ID NO: 325
DCVRFWGKCSQTSDCCPHLACKSKWPRNICVWDGSVG SEQ ID NO: 326
GCFGYKCDYYKGCCSGYVCSPTWKWCVRPGPGR SEQ ID NO: 327
MNAKFILLLVLTTMMLLPDTKGAEVIRCSGSKQCYGPCKQQTGCTNSKCM NKVCKCYGCG SEQ
ID NO: 328 MNAKLIYLLLVVTTMTLMFDTAQAVDIMCSGPKQCYGPCKKETGCPNAKC
MNRRCKCYGCV SEQ ID NO: 329
MNAKLIYLLLVVTTMMLTFDTTQAGDIKCSGTRQCWGPCKKQTTCTNSKC MNGKCKCYGCVG SEQ
ID NO: 330 MNTKFIFLLLVVTNTMMLFDTKPVEGISCTGSKQCYDPCKRKTGCPNAKCM
NKSCKCYGCG SEQ ID NO: 331 GVPINVKCSGSRDCLEPCKKAGMRFGKCINRKCHCTPK
SEQ ID NO: 332 GVPINVKCTGSPQCLKPCKDAGMRFGKCINGKCHCTPK SEQ ID NO:
333 GVIINVKCKISRQCLEPCKKAGMRFGKCMNGKCHCTPK SEQ ID NO: 334
GVPINVKCRGSPQCIQPCRDAGMRFGKCMNGKCHCTPQ SEQ ID NO: 335
GVEINVKCTGSHQCIKPCKDAGMRFGKCINRKCHCTPK SEQ ID NO: 336
GVEINVKCSGSPQCLKPCKDAGMRFGKCMNRKCHCTPK SEQ ID NO: 337
GVPTDVKCRGSPQCIQPCKDAGMRFGKCMNGKCHCTPK SEQ ID NO: 338
GVPINVSCTGSPQCIKPCKDAGMRFGKCMNRKCHCTPK SEQ ID NO: 339
GVPINVPCTGSPQCIKPCKDAGMRFGKCMNRKCHCTPK SEQ ID NO: 340
VGINVKCKHSGQCLKPCKDAGMRFGKCINGKCDCTPK SEQ ID NO: 341
VGINVKCKHSGQCLKPCKDAGMRFGKCMNGKCDCTPK SEQ ID NO: 342
VGIPVSCKHSGQCIKPCKDAGMRFGKCMNRKCDCTPK SEQ ID NO: 343
RKGCFKEGHSCPKTAPCCRPLVCKGPSPNTKKCTRP SEQ ID NO: 344
SFCIPFKPCKSDENCCKKFKCKTTGIVKLCRW SEQ ID NO: 345
LKGCLPRNRFCNALSGPRCCSGLRCKELSIWASKCL SEQ ID NO: 346
GNYCLRGRCLPGGRKCCNGRPCECFAKICSCKPK SEQ ID NO: 347
TVKCGGCNRKCCPGGCRSGKCINGKCQCY SEQ ID NO: 348
GCMKEYCAGQCRGKVSQDYCLKHCKCIPR SEQ ID NO: 349
ACLGFGEKCNPSNDKCCKSSSLVCSQKHKWCKYG SEQ ID NO: 350
RGGCLPHNRFCNALSGPRCCSGLRCKELSIRDSRCLG SEQ ID NO: 351
RGGCLPRNKFCNPSSGPRCCSGLTCKELNIWASKCL SEQ ID NO: 352
QRSCAKPGDMCMGIKCCDGQCGCNRGTGRCFCK SEQ ID NO: 353
ARGCADAYKSCNHPRTCCDGYNGYKRACICSGSNCKCKKS SEQ ID NO: 354
RGGCLPHNRFCNALSGPRCCSGLRCKELSIWDSRCLG SEQ ID NO: 355
RGGCLPHNRFCNALSGPRCCSGLKCKELSIYDSRCLG SEQ ID NO: 356
RGGCLPHNRFCNALSGPRCCSRLKCKELSIWDSRCLG SEQ ID NO: 357
RGGCLPHNRFCNALTGPRCCSRLRCKELSIWDSICLG SEQ ID NO: 358
SCADAYKSCDSLKCCNNRTCMCSMIGTNCTCRKK SEQ ID NO: 359
ERRCLPAGKTCVRGPMRVPCCGSCSQNKCT SEQ ID NO: 360
LCSREGEFCYKLRKCCAGFYCKAFVLHCYRN SEQ ID NO: 361
ACGSCRKKCKGSGKCINGRCKCY SEQ ID NO: 362 ACGSCRKKCKGPGKCINGRCKCY SEQ
ID NO: 363 ACQGYMRKCGRDKPPCCKKLECSKTWRWCVWN SEQ ID NO: 364
GRYCQKWMWTCDSKRACCEGLRCKLWCRKI SEQ ID NO: 365
NAKCRGSPECLPKCKEAIGKAAGKCMNGKCKCYP SEQ ID NO: 366
NVKCRGSKECLPACKAAVGKAAGKCMNGKCKCYP SEQ ID NO: 367
NVKCRGSPECLPKCKEAIGKSAGKCMNGKCKCYP SEQ ID NO: 368
NAKCRGSPECLPKCKQAIGKAAGKCMNGKCKCYP SEQ ID NO: 369
RGYCAEKGIKCHNIHCCSGLTCKCKGSSCVCRK SEQ ID NO: 370
ERGCKLTFWKCKNKKECCGWNACALGICMPR SEQ ID NO: 371
KKKCIAKDYGRCKWGGTPCCRGRGCICSIMGTNCECKPR SEQ ID NO: 372
GCKLTFWKCKNKKECCGWNACALGICMPR SEQ ID NO: 373
ACKGLFVTCTPGKDECCPNHVCSSKHKWCKYK SEQ ID NO: 374
IACAPRGLLCFRDKECCKGLTCKGRFVNTWPTFCLV SEQ ID NO: 375
ACAGLYKKCGKGVNTCCENRPCKCDLAMGNCICKKK SEQ ID NO: 376
FTCAISCDIKVNGKPCKGSGEKKCSGGWSCKFNVCVKV SEQ ID NO: 377
GFCAQKGIKCHDIHCCTNLKCVREGSNRVCRKA SEQ ID NO: 378
CAKKRNWCGKNEDCCCPMKCIYAWYNQQGSCQSTITGLFKKC SEQ ID NO: 379
YCQKWMWTCDSARKCCEGLVCRLWCKKI SEQ ID NO: 380
RGGCLPHNKFCNALSGPRCCSGLKCKELTIWNTKCLE SEQ ID NO: 381
NVKCTGSKQCLPACKAAVGKAAGKCMNGKCKCYT SEQ ID NO: 382
QRSCAKPGEMCMRIKCCDGQCGCNRGTGRCFCK SEQ ID NO: 383
GCIPKHKRCTWSGPKCCNNISCHCNISGTLCKCRPG SEQ ID NO: 384
NYCVAKRCRPGGRQCCSGKPCACVGKVCKCPRD SEQ ID NO: 385
ERGCSGAYKRCSSSQRCCEGRPCVCSAINSNCKCRKT SEQ ID NO: 386
RYCPRNPEACYNYCLRTGRPGGYCGGRSRITCFCFR SEQ ID NO: 387
QRSCAKPGEMCMGIKCCDGQCGCNRGTGRCFCK SEQ ID NO: 388
RRGCFKEGKWCPKSAPCCAPLKCKGPSIKQQKCVRE SEQ ID NO: 389
TVKCGGCNRKCCAGGCRSGKCINGKCQCYGR SEQ ID NO: 390
ERRCEPSGKPCRPLMRIPCCGSCVRGKCA SEQ ID NO: 391
RGGCLPRNKFCNPSSGPRCCSGLTCKELNIWANKCL SEQ ID NO: 392
CAKKRNWCGKNEDCCCPMKCIYAWYNQQGSCQTTITGLFKKC SEQ ID NO: 393
VRIPVSCKHSGQCLKPCKDAGMRTGKCMNGKCDCTPK SEQ ID NO: 394
VKCTTSKDCWPPCKKVTGRA SEQ ID NO: 395
GIVCRVCRIICGMQGRRVNICRAPIRCRCRRG SEQ ID NO: 396
SERDCIRHLQRCRENRDCCSRRCSRRGTNPERRCR SEQ ID NO: 397
VRIPVSCRHSGQCLRPCRDAGMRFGRCMNGRCDCTPR SEQ ID NO: 398
GVPINVRCRGSRDCLDPCRRAGMRFGRCINSRCHCTP SEQ ID NO: 399
AVCVYRTCDRDCRRRGYRSGRCINNACRCYPYG SEQ ID NO: 400
ISCTGSRQCYDPCRRRTGCPNARCMNRSCRCYGCG SEQ ID NO: 401
QVQTNVRCQGGSCASVCRREIGVAAGRCINGRCVCYRN SEQ ID NO: 402
EVIRCSGSRQCYGPCRQQTGCTNSRCMNRVCRCYGCG SEQ ID NO: 403
ACRGVFDACTPGRNECCPNRVCSDRHRWCRWRL SEQ ID NO: 404
QIYTSRECNGSSECYSHCEGITGRRSGRCINRRCYCYR SEQ ID NO: 405
GCLEFWWRCNPNDDRCCRPRLRCSRLFRLCNFSFG SEQ ID NO: 406
DCVRFWGRCSQTSDCCPHLACRSRWPRNICVWDGSVG SEQ ID NO: 407
GCFGYRCDYYRGCCSGYVCSPTWRWCVRPGPGR SEQ ID NO: 408
MNARFILLLVLTTMMLLPDTRGAEVIRCSGSRQCYGPCRQQTGCTNSRCMN RVCRCYGCG SEQ
ID NO: 409 MNARLIYLLLVVTTMTLMFDTAQAVDIMCSGPRQCYGPCRRETGCPNARC
MNRRCRCYGCV SEQ ID NO: 410
MNARLIYLLLVVTTMMLTFDTTQAGDIRCSGTRQCWGPCRRQTTCTNSRC MNGRCRCYGCVG SEQ
ID NO: 411 MNTRFIFLLLVVTNTMMLFDTRPVEGISCTGSRQCYDPCRRRTGCPNARCM
NRSCRCYGCG SEQ ID NO: 412 GVPINVRCSGSRDCLEPCRRAGMRFGRCINRRCHCTPR
SEQ ID NO: 413 GVPINVRCTGSPQCLRPCRDAGMRFGRCINGRCHCTPR SEQ ID NO:
414 GVIINVRCRISRQCLEPCRRAGMRFGRCMNGRCHCTPR SEQ ID NO: 415
GVPINVRCRGSPQCIQPCRDAGMRFGRCMNGRCHCTPQ SEQ ID NO: 416
GVEINVRCTGSHQCIRPCRDAGMRFGRCINRRCHCTPR SEQ ID NO: 417
GVEINVRCSGSPQCLRPCRDAGMRFGRCMNRRCHCTPR SEQ ID NO: 418
GVPTDVRCRGSPQCIQPCRDAGMRFGRCMNGRCHCTPR SEQ ID NO: 419
GVPINVSCTGSPQCIRPCRDAGMRFGRCMNRRCHCTPR SEQ ID NO: 420
GVPINVPCTGSPQCIRPCRDAGMRFGRCMNRRCHCTPR SEQ ID NO: 421
VGINVRCRHSGQCLRPCRDAGMRFGRCINGRCDCTPR SEQ ID NO: 422
VGINVRCRHSGQCLRPCRDAGMRFGRCMNGRCDCTPR SEQ ID NO: 423
VGIPVSCRHSGQCIRPCRDAGMRFGRCMNRRCDCTPR SEQ ID NO: 424
RRGCFREGHSCPRTAPCCRPLVCRGPSPNTRRCTRP SEQ ID NO: 425
SFCIPFRPCRSDENCCRRFRCRTTGIVRLCRW SEQ ID NO: 426
LRGCLPRNRFCNALSGPRCCSGLRCRELSIWASRCL SEQ ID NO: 427
GNYCLRGRCLPGGRRCCNGRPCECFARICSCRPR SEQ ID NO: 428
TVRCGGCNRRCCPGGCRSGRCINGRCQCY
SEQ ID NO: 429 GCMREYCAGQCRGRVSQDYCLRHCRCIPR SEQ ID NO: 430
ACLGFGERCNPSNDRCCRSSSLVCSQRHRWCRYG SEQ ID NO: 431
RGGCLPHNRFCNALSGPRCCSGLRCRELSIRDSRCLG SEQ ID NO: 432
RGGCLPRNRFCNPSSGPRCCSGLTCRELNIWASRCL SEQ ID NO: 433
QRSCARPGDMCMGIRCCDGQCGCNRGTGRCFCR SEQ ID NO: 434
ARGCADAYRSCNHPRTCCDGYNGYRRACICSGSNCRCRRS SEQ ID NO: 435
RGGCLPHNRFCNALSGPRCCSGLRCRELSIWDSRCLG SEQ ID NO: 436
RGGCLPHNRFCNALSGPRCCSGLRCRELSIYDSRCLG SEQ ID NO: 437
RGGCLPHNRFCNALSGPRCCSRLRCRELSIWDSRCLG SEQ ID NO: 438
RGGCLPHNRFCNALTGPRCCSRLRCRELSIWDSICLG SEQ ID NO: 439
SCADAYKSCDSLRCCNNRTCMCSMIGTNCTCRRR SEQ ID NO: 440
ERRCLPAGRTCVRGPMRVPCCGSCSQNRCT SEQ ID NO: 441
LCSREGEFCYRLRRCCAGFYCRAFVLHCYRN SEQ ID NO: 442
ACGSCRRRCRGSGRCINGRCRCY SEQ ID NO: 443 ACGSCRRRCRGPGRCINGRCRCY SEQ
ID NO: 444 ACQGYMRRCGRDRPPCCRRLECSRTWRWCVWN SEQ ID NO: 445
GRYCQRWMWTCDSRRACCEGLRCRLWCRRI SEQ ID NO: 446
NARCRGSPECLPRCREAIGRAAGRCMNGRCRCYP SEQ ID NO: 447
NVRCRGSRECLPACRAAVGRAAGRCMNGRCRCYP SEQ ID NO: 448
NVRCRGSPECLPRCREAIGRSAGRCMNGRCRCYP SEQ ID NO: 449
NARCRGSPECLPRCRQAIGRAAGRCMNGRCRCYP SEQ ID NO: 450
RGYCAERGIRCHNIHCCSGLTCRCRGSSCVCRR SEQ ID NO: 451
ERGCRLTFWRCRNRRECCGWNACALGICMPR SEQ ID NO: 452
RRRCIARDYGRCRWGGTPCCRGRGCICSIMGTNCECRPR SEQ ID NO: 453
GCRLTFWRCRNRRECCGWNACALGICMPR SEQ ID NO: 454
ACRGLFVTCTPGRDECCPNHVCSSRHRWCRYR SEQ ID NO: 455
IACAPRGLLCFRDRECCRGLTCRGRFVNTWPTFCLV SEQ ID NO: 456
ACAGLYRRCGRGVNTCCENRPCRCDLAMGNCICRRR SEQ ID NO: 457
FTCAISCDIRVNGRPCRGSGERRCSGGWSCRFNVCVRV SEQ ID NO: 458
GFCAQRGIRCHDIHCCTNLRCVREGSNRVCRRA SEQ ID NO: 459
CARRRNWCGRNEDCCCPMRCIYAWYNQQGSCQSTITGLFRRC SEQ ID NO: 460
YCQRWMWTCDSARRCCEGLVCRLWCRRI SEQ ID NO: 461
RGGCLPHNRFCNALSGPRCCSGLRCRELTIWNTRCLE SEQ ID NO: 462
NVRCTGSRQCLPACRAAVGRAAGRCMNGRCRCYT SEQ ID NO: 463
QRSCARPGEMCMRIRCCDGQCGCNRGTGRCFCR SEQ ID NO: 464
GCIPRHRRCTWSGPRCCNNISCHCNISGTLCRCRPG SEQ ID NO: 465
NYCVARRCRPGGRQCCSGRPCACVGRVCRCPRD SEQ ID NO: 466
ERGCSGAYRRCSSSQRCCEGRPCVCSAINSNCRCRRT SEQ ID NO: 467
QRSCARPGEMCMGIRCCDGQCGCNRGTGRCFCR SEQ ID NO: 468
RRGCFREGRWCPRSAPCCAPLRCRGPSIRQQRCVRE SEQ ID NO: 469
TVRCGGCNRRCCAGGCRSGRCINGRCQCYGR SEQ ID NO: 470
ERRCEPSGRPCRPLMRIPCCGSCVRGRCA SEQ ID NO: 471
RGGCLPRNRFCNPSSGPRCCSGLTCRELNIWANRCL SEQ ID NO: 472
CARRRNWCGRNEDCCCPMRCIYAWYNQQGSCQTTITGLFRRC SEQ ID NO: 473
VRIPVSCRHSGQCLRPCRDAGMRTGRCMNGRCDCTPR SEQ ID NO: 474
QKILSNRCNNSSECIPHCIRIFGTRAAKCINRKCYCYP SEQ ID NO: 475
AVCNLKRCQLSCRSLGLLGKCIGDKCECVKHG SEQ ID NO: 476
ISIGIRCSPSIDLCEGQCRIRRYFTGYCSGDTCHCSG SEQ ID NO: 477
GDCLPHLRRCRENNDCCSRRCRRRGANPERRCR SEQ ID NO: 478
SCEPGRTFRDRCNTCKCGADGRSAACTLRACPNQ SEQ ID NO: 479
GDCLPHLKRCKADNDCCGKKCKRRGTNAEKRCR SEQ ID NO: 480
GDCLPHLKRCKENNDCCSKKCKRRGTNPEKRCR SEQ ID NO: 481
KDCLKKLKLCKENKDCCSKSCKRRGTNIEKRCR SEQ ID NO: 482
GDCLPHLKRCKENNDCCSKKCKRRGANPEKRCR SEQ ID NO: 483
VFINVKCRGSPECLPKCKEAIGKSAGKCMNGKCKCYP SEQ ID NO: 484
VFINAKCRGSPECLPKCKEAIGKAAGKCMNGKCKCYP SEQ ID NO: 485
VIINVKCKISRQCLEPCKKAGMRFGKCMNGKCHCTP SEQ ID NO: 486
VPTDVKCRGSPQCIQPCKDAGMRFGKCMNGKCHCTP SEQ ID NO: 487
VRIPVSCKHSGQCLKPCKDAGMRFGKCMNGKCDCTP SEQ ID NO: 488
VRIPVSCRHSGQCLRPCRDAGMRFGRCMNGRCDCTP SEQ ID NO: 489
TNVSCTTSKECWSVCQRLHNTSRGKCMNKKCRC SEQ ID NO: 490
NVKCTGSKQCLPACKAAVGKAAGKCMNGKCKC SEQ ID NO: 491
GVPINVRCRGSRDCLDPCRGAGERHGRCGNSRCHCTP SEQ ID NO: 492
VRIPVSCRHSGQCLRPCRDAGERHGRCGGGRCDCTPR SEQ ID NO: 493
QVQTNVRCQGGSCGSVCRREGGGAGGGCGNGRCGCYRN SEQ ID NO: 494
IKCSESYQCFPVCKSRFGKTNGRCVNGFCDCF SEQ ID NO: 495
VKCSSPQQCLKPCKAAFGISAGgKCINGKCKCY SEQ ID NO: 496
VSCSASSQCWPVCKKLFGTYRGKCMNSKCRCY SEQ ID NO: 497
ESCTASNQCWSICKRLHNTNRGKCMNKKCRCY SEQ ID NO: 498
VSCTTSKECWSVCEKLYNTSRGKCMNKKCRCY SEQ ID NO: 499
MRCKSSKECLVKCKQATGRPNGKCMNRKCKCY SEQ ID NO: 500
IKCTLSKDCYSPCKKETGCPRAKCINRNCKCY SEQ ID NO: 501
IRCSGSRDCYSPCMKQTGCPNAKCINKSCKCY SEQ ID NO: 502
IRCSGTRECYAPCQKLTGCLNAKCMNKACKCY SEQ ID NO: 503
ISCTNPKQCYPHCKKETGYPNAKCMNRKCKCF SEQ ID NO: 504
ASCRTPKDCADPCRKETGCPYGKCMNRKCKCN SEQ ID NO: 505
TSCISPKQCTEPCRAKGCKHGKCMNRKCHCM SEQ ID NO: 506
KECTGPQHCTNFCRKN-KCTHGKCMNRKCKCF SEQ ID NO: 507
IKCRTPKDCADPCRKQTGCPHAKCMNKTCRCH SEQ ID NO: 508
VKCTTSKECWPPCKAATGKAAGKCMNKKCKCQ SEQ ID NO: 509
LECGASRECYDPCFKAFGRAHGKCMNNKCRCY SEQ ID NO: 510
EKCFATSQCWTPCKKAIGSLQSKCMNGKCKCY SEQ ID NO: 511
VRCYASRECWEPCRRVTGSAQAKCQNNQCRCY SEQ ID NO: 512
VKCSASRECWVACKKVTGSGQGKCQNNQCRCY SEQ ID NO: 513
VKCISSQECWIACKKVTGRFEGKCQNRQCRCY SEQ ID NO: 514
VRCYDSRQCWIACKKVTGSTQGKCQNKQCRCY SEQ ID NO: 515
VDCTVSKECWAPCKAAFGVDRGKCMGKKCKCY SEQ ID NO: 516
AKCRGSPECLPKCKEAIGKAAGKCMNGKCKCY SEQ ID NO: 517
KKCQGGSCASVCRRVIGVAAGKCINGRCVCY SEQ ID NO: 518
KKCSNTSQCYKTCEKVVGVAAGKCMNGKCICY SEQ ID NO: 519
VKCSGSSKCVKICIDRYNTRGAKCINGRCTCY SEQ ID NO: 520
NRCNNSSECIPHCIRIFGTRAAKCINRKCYCY SEQ ID NO: 521
KECNGSSECYSHCEGITGKRSGKCINKKCYCY SEQ ID NO: 522
AFCNLRRCELSCRSLGLLGKCIGEECKCV SEQ ID NO: 523
AVCNLKRCQLSCRSLGLLGKCIGDKCECV SEQ ID NO: 524
AACYSS-DCRVKCVAMGFSSGKCINSKCKCY SEQ ID NO: 525
AICATDADCSRKCPGNPPCRNGFCACT SEQ ID NO: 526
TECQIKNDCQRYCQSVKECKYGKCYCN SEQ ID NO: 527
TQCQSVRDCQQYCLTPDRCSYGTCYCK SEQ ID NO: 528
VSCRYGSDCAEPCKRLKCLLPSKCINGKCTCY SEQ ID NO: 529
IKCRYPADCHIMCRKVTGRAEGKCMNGKCTCY SEQ ID NO: 530
IKCSSSSSCYEPCRGVTGRAHGKCMNGRCTCY SEQ ID NO: 531
VKCTGSKQCLPACKAAVGKAAGKCMNGKCKCY SEQ ID NO: 532
VSCKHSGQCIKPCKDA-GMRFGKCMNRKCDCT SEQ ID NO: 533
VKCRGSPQCIQPCRDA-GMRFGKCMNGKCHCT SEQ ID NO: 534
VKCTSPKQCLPPCKAQFGIRAGAKCMNGKCKCY SEQ ID NO: 535
VKCTSPKQCSKPCKELYGSSAGAKCMNGKCKCY SEQ ID NO: 536
VKCTSPKQCLPPCKEIYGRHAGAKCMNGKCHCS SEQ ID NO: 537
VKCTGSKQCWPVCKQMFGKPNGKCMNGKCRCY SEQ ID NO: 538
VKCRGSRDCLDPCKKAGMRFGKCINSKCHCT SEQ ID NO: 539
VRCVTDDDCFRKCPGNPSCKRGFCACK SEQ ID NO: 540
VPCNNSRPCVPVCIREVNNKNGKCSNGKCLCY SEQ ID NO: 541
VPINVKCRGSRDCLDPCKKAGMRFGKCINSKCHCTP SEQ ID NO: 542
VQTNVKCQGGSCASVCRREIGVAAGKCINGKCVCYRN SEQ ID NO: 543
AEIIRCSGTRECYAPCQKLTGCLNAKCMNKACKCYGCV SEQ ID NO: 544
RPTDIKCSASYQCFPVCKSRFGKTNGRCVNGLCDCF SEQ ID NO: 545
QFTDVKCTGSKQCWPVCKQMFGKPNGKCMNGKCRCYS SEQ ID NO: 546
VGINVKCKHSRQCLKPCKDAGMRFGKCTNGKCHCTPK SEQ ID NO: 547
VVIGQRCYRSPDCYSACKKLVGKATGKCTNGRCDC SEQ ID NO: 548
NFKVEGACSKPCRKYCIDKGARNGKCINGRCHCYY SEQ ID NO: 549
QIDTNVKCSGSSKCVKICIDRYNTRGAKCINGRCTCYP SEQ ID NO: 550
GVPISVRCRGSRDCLEPCRRAGTRFGRCINGRCHCTP SEQ ID NO: 551
GVPISVRCRGSRDCLEPCRRAGTRFGRCIQSRCHCTP SEQ ID NO: 552
GVPISVRCRGSRDCLEPCRRAGTRFGRCINRRCHCTP SEQ ID NO: 553
GVPINVRCRGSRDCLEPCRRAGTRFGRCINSRCHCTP
SEQ ID NO: 554 GVPINVRCRGSRDCLEPCRRAGTRFGRCIQSRCHCTP SEQ ID NO: 555
GVPINVRCRGSRDCLEPCRRAGTRFGRCIQSRCHCYP SEQ ID NO: 556
GVPINVRCRGSRDCYEPCRRAGTRFGRCIQSRCHCTP SEQ ID NO: 557
GVPINVRCRGSRDCLEPCRRAGTRFGRCIQSRCYCTP SEQ ID NO: 558
GVPISVRCRGSRDCLEPCRRAGTRFGRCIQSRCHCYP SEQ ID NO: 559
GVPISVRCRGSRDCYEPCRRAGTRFGRCIQSRCHCTP SEQ ID NO: 560
GVPISVRCRGSRDCLEPCRRAGTRFGRCIQSRCYCTP SEQ ID NO: 561
GVPINVRCRGSRDCLEPCRRAGTRFGRCIASRCHCYP SEQ ID NO: 562
GVPINVRCRGSRDCLEPCRRAGTRFGRCISSRCHCYP SEQ ID NO: 563
GVPINVRCRGSRDCLEPCRRAGTRFGRCITSRCHCYP SEQ ID NO: 564
GVPINVRCRGSRDCLEPCRRAGTRFGRCINSRCHCYP
[0084] In any of SEQ ID NO: 1-SEQ ID NO: 564 or fragment thereof,
any one or more K residues can be replaced by an R residue or an A
residue, any one or more R residues can be replaced by a K residue
or an A residue, any one or more A residues can be replaced by a K
residue or an R residue, all K residues can be replaced by R
residues or A residues, all but one K residue can be replaced by R
or A residues, all but two K residues can be replaced by R residues
or A residues, or in any combination thereof. In any of SEQ ID NO:
1-SEQ ID NO: 564 or any fragment thereof, any one or more M
residues can be replaced by any one of I, L, or V residues, any one
or more L residues can be replaced by any one of V, I, or M
residues, any one or more I residues can be replaced by any one of
M, L, or V residues, or any one or more V residues can be replaced
by any one of I, L, or M residues. In any embodiment, at least one
of the amino acids alone or in combination can be interchanged in
the peptides or peptide fragments as follows: K/R, M/I/L/V, G/A,
S/T, Q/N, and D/E wherein each letter is each individually any
amino acid or amino acid analogue. In some instances, the peptide
can contain only one lysine residue, or no lysine residue. In any
of SEQ ID NO: 1-SEQ ID NO: 564 or fragment thereof, any amino acid
can be replaced with citrulline. In any of SEQ ID NO: 1-SEQ ID NO:
564 or any fragment thereof, X can independently be any number of
any amino acid or no amino acid. In some cases, a peptide can
include the first two N-terminal amino acids GS, as with peptides
of SEQ ID NO: 1-SEQ ID NO: 274, or such N-terminal amino acids (GS)
can be substituted by any other one or two amino acids. In other
cases, a peptide does not include the first two N-terminal amino
acids GS, as with peptides of SEQ ID NO: 275-SEQ ID NO: 564. In
some cases, the N-terminus of the peptide is blocked, such as by an
acetyl group; in other instances the C-terminus of the peptide is
block, such as by an amide group.
[0085] In some instances, the peptide is any one of SEQ ID NO:
1-564 or a functional fragment thereof. In other embodiments, the
peptide of the disclosure further comprises a peptide with 100%,
99%, 97%, 95%, 90%, 85%, or 80% homology to any one of SEQ ID NO:
1-SEQ ID NO: 564. In further embodiments, the peptide fragment
comprises a contiguous fragment of any one of SEQ ID NO: 1-SEQ ID
NO: 564 that is at least 17, at least 18, at least 19, at least 20,
at least 21, at least 22, at least 23, at least 24, at least 25, at
least 26, at least 27, at least 28, at least 29, at least 30, at
least 31, at least 32, at least 33, at least 34, at least 35, at
least 36, at least 37, at least 38, at least 39, at least 40, at
least 41, at least 42, at least 43, at least 44, at least 45, at
least 46 residues long, wherein the peptide fragment is selected
from any portion of the peptide. In some embodiments, such peptide
fragments contact the cartilage and exhibit properties of those
described herein for peptide and peptide-active agent
conjugates.
[0086] The peptides of the present disclosure can further comprise
negative amino acid residues. In some cases, the peptide has 2 or
fewer negative amino acid residues. In other cases, the peptide has
4 or fewer negative amino acid residues, 3 or fewer negative amino
acid residues, or 1 or fewer negative amino acid residues. The
negative amino acid residues can be selected from any negative
charged amino acid residues. The negative amino acid residues can
selected from either E or D, or a combination of both E and D.
[0087] The peptides of the present disclosure can further comprise
basic amino acid residues. In some embodiments, basic residues are
added to the peptide sequence to increase the charge at
physiological pH. The added basic residues can be any basic amino
acid. The added basic residues can be selected from K or R, or a
combination of K or R.
[0088] In some embodiments, the peptide has a charge distribution
comprising an acidic region and a basic region. An acidic region
can be a nub. A nub is a portion of a peptide extending out of the
peptide's three-dimensional structure. A basic region can be a
patch. A patch is a portion of a peptide that does not designate
any specific topology characteristic of the peptide's
three-dimensional structure. In further embodiments, a
cystine-dense peptide can be 6 or more basic residues and 2 or
fewer acidic residues.
[0089] The peptides of the present disclosure can further comprise
positively charged amino acid residues. In some cases, the peptide
has at least 2 positively charged residues. In other cases, the
peptide has at least 3 positively charged residues, at least 4
positively charged residues, at least 5 positively charged
residues, at least 6 positively charged residues, at least 7
positively charged residues, at least 8 positively charged residues
or at least 9 positively charged residues. The positively charged
residues can be selected from any positively charged amino acid
residues. The positively charged residues can be selected from
either K or R, or a combination of K and R.
[0090] In addition, the peptides herein can comprise a 4-19 amino
acid residue fragment of any of the above sequences containing at
least 2 cysteine residues, and at least 2 or 3 positively charged
amino acid residues (for example, arginine, lysine or histidine, or
any combination of arginine, lysine or histidine). In other
embodiments, the peptides herein is a 20-70 amino acid residue
fragment of any of the above sequences containing at least 2
cysteine residues, no more than 2 basic residues, and at least 2 or
3 positively charged amino acid residues (for example, arginine,
lysine or histidine, or any combination of arginine, lysine or
histidine). In some embodiments, such peptide fragments contact the
cartilage and exhibit properties of those described herein for
peptide and peptide-active agent conjugates.
[0091] In some embodiments, the peptide contains one or more
disulfide bonds and has a positive net charge at neutral pH. At
physiological pH, peptides can have a net charge, for example, of
-5, -4, -3, -2, -1, 0, +1, +2, +3, +4, or +5. When the net charge
is zero, the peptide can be uncharged or zwitterionic. In some
instances, the peptide can have a positive charge at physiological
pH. In some instances, the peptide can have a charge .gtoreq.+2 at
physiological pH, .gtoreq.+3.5 at physiological pH, .gtoreq.+4.5 at
physiological pH. In some embodiments, the peptide contains one or
more disulfide bonds and has a positive net charge at neutral pH
where the net charge can be +0.5 or less than +0.5, +1 or less than
+1, +1.5 or less than +1.5, +2 or less than +2, +2.5 or less than
+2.5, +3 or less than +3, +3.5 or less than +3.5, +4 or less than
+4, +4.5 or less than +4.5, +5 or less than +5, +5.5 or less than
+5.5, +6 or less than +6, +6.5 or less than +6.5, +7 or less than
+7, +7.5 or less than +7.5, +8 or less than +8, +8.5 or less than
+8.5, +9 or less than +9.5, +10 or less than +10. In some
embodiments, the peptide has a negative net charge at physiological
pH where the net charge can be -0.5 or less than -0.5, -1 or less
than -1, -1.5 or less than -1.5, -2 or less than -2, -2.5 or less
than -2.5, -3 or less than -3, -3.5 or less than -3.5, -4 or less
than -4, -4.5 or less than -4.5, -5 or less than -5, -5.5 or less
than -5.5, -6 or less than -6, -6.5 or less than -6.5, -7 or less
than -7, -7.5 or less than -7.5, -8 or less than -8, -8.5 or less
than -8.5, -9 or less than -9.5, -10 or less than -10. In some
cases, the engineering of one or more mutations within a peptide
yields a peptide with an altered isoelectric point, charge, surface
charge, or rheology at physiological pH. Such engineering of a
mutation to a peptide derived from a scorpion or spider can change
the net charge of the complex, for example, by decreasing the net
charge by 1, 2, 3, 4, or 5, or by increasing the net charge by 1,
2, 3, 4, or 5. In such cases, the engineered mutation may
facilitate the ability of the peptide to contact the cartilage.
Suitable amino acid modifications for improving the rheology and
potency of a peptide can include conservative or non-conservative
mutations. A peptide can comprises at most 1 amino acid mutation,
at most 2 amino acid mutations, at most 3 amino acid mutations, at
most 4 amino acid mutations, at most 5 amino acid mutations, at
most 6 amino acid mutations, at most 7 amino acid mutations, at
most 8 amino acid mutations, at most 9 amino acid mutations, at
most 10 amino acid mutations, or another suitable number as
compared to the sequence of the venom or toxin that the peptide is
derived from. In other cases, a peptide, or a functional fragment
thereof, comprises at least 1 amino acid mutation, at least 2 amino
acid mutations, at least 3 amino acid mutations, at least 4 amino
acid mutations, at least 5 amino acid mutations, at least 6 amino
acid mutations, at least 7 amino acid mutations, at least 8 amino
acid mutations, at least 9 amino acid mutations, at least 10 amino
acid mutations, or another suitable number as compared to the
sequence of the venom or toxin that the peptide is derived from. In
some embodiments, mutations can be engineered within a peptide to
provide a peptide that has a desired charge or stability at
physiological pH.
[0092] Peptides can be mutated to add function or remove function.
For example, peptides and peptide-conjugates of the present
disclosure can be mutated to retain, remove, or add the ability to
bind to ion channels, or to promote agonizing or antagonizing ion
channels, such as potassium channel binding that may occur with the
peptide or peptide-conjugates (e.g., the potassium channel hERG).
In some instances, it can be advantageous to remove potassium
channel binding from a peptide used for delivery of an active
agent. Mutations can include one or more N to S, D to E, M to T, N
to Q, N to A, N to S, N to T, N to L, S to G, and S to R amino acid
substitutions, or one or more L to Y, H to Y, and T to Y amino acid
substitutions, or any combination of thereof, depending on whether
the variant is designed to retain function or to remove function of
binding to the ion channel. In some embodiments the peptides and
peptide-drug conjugates of the present disclosure are mutated to
minimize ion channel binding in order to minimize side effects or
enhance the safety either in the target tissue or systemically.
[0093] In some embodiments, charge can play a role in cartilage
homing. The interaction of a peptide of this disclosure in solution
and in vivo can be influenced by the isoelectric point (pI) of the
peptide and/or the pH of the solution or the local environment it
is in. The charge of a peptide in solution can impact the
solubility of the protein as well as parameters such as
biodistribution, bioavailability, and overall pharmacokinetics.
Additionally, positively charged molecules can interact with
negatively charged molecules. Positively charged molecules such as
the peptides disclosed herein can interact and bind with negatively
charged molecules such as the negatively charged extracellular
matrix molecules in the cartilage including hyaluranon and
aggrecan. Positively charged residues can also interact with
specific regions of other proteins and molecules, such as
negatively charged residues of receptors or electronegative regions
of an ion channel pore on cell surfaces. As such, the pI of a
peptide can influence whether a peptide of this disclosure can
efficiently home to cartilage. Identifying a correlation between pI
and cartilage homing can be an important strategy in identifying
lead peptide candidates of the present disclosure. The pI of a
peptide can be calculated using a number of different methods
including the Expasy pI calculator and the Sillero method. The
Expasy pI can be determined by calculating pKa values of amino
acids as described in Bjellqvist et al., which were defined by
examining polypeptide migration between pH 4.5 to pH 7.3 in an
immobilized pH gradient gel environment with 9.2M and 9.8M urea at
15.degree. C. or 25.degree. C. (Bjellqvist et al. Electrophoresis.
14(10):1023-31 (1993)). The Sillero method of calculating pI can
involve the solution of a polynomial equation and the individual
pKas of each amino acid. This method does not use denaturing
conditions (urea) (Sillero et al. 179(2): 319-35 (1989)) Using
these pI calculation methods and quantifying the cartilage to blood
ratio of peptide signal after administration to a subject can be a
strategy for identifying a trend or correlation in charge and
cartilage homing. In some embodiments, a peptide with a pI above
biological pH (.about.pH 7.4) can exhibit efficient homing to
cartilage. In some embodiments, a peptide with a pI of at least 8,
at least 9, at least 10, or at least 11 can efficiently home to
cartilage. In other embodiments, a peptide with a pI of 11-12 can
home most efficiently to cartilage. In certain embodiments, a
peptide can have a pI of about 9. In other embodiments, a peptide
can have a pI of 8-10. In some embodiments, more basic peptides can
home more efficiently to cartilage. In other embodiments, a high pI
alone may not be sufficient to cause cartilage homing of a
peptide.
[0094] In some embodiments, the tertiary structure and
electrostatics of a peptide of the disclosure can impact cartilage
homing. Structural analysis or analysis of charge distribution can
be a strategy to predict residues important in biological function,
such as cartilage homing. For example, several peptides of this
disclosure that home to cartilage can be grouped into a structural
class defined herein as "hitchins," and can share the properties of
disulfide linkages between C1-C4, C2-C5, and C3-C6. The folding
topologies of peptides linked through three disulfide linkages
(C1-C4, C2-C5, and C3-C6), can be broken down into structural
families based on the three-dimensional arrangement of the
disulfides. Some cystine-dense peptides have the C3-C6 disulfide
linkage passing through the macrocycle formed by the C1-C4 and
C2-C5 disulfide linkages, hitchins have the C2-C5 disulfide linkage
passing through the macrocycle formed by the C1-C4 and C3-C6
disulfide linkages, and yet other structural families have the
C1-C4 disulfide linkage passing through the macrocycle formed by
the C2-C5 and C3-C6 disulfide linkages. Variants of "hitchin" class
peptides with preserved disulfide linkages at these cysteine
residues, primary sequence identity, and/or structural homology can
be a method of identifying or predicting other potential peptide
candidates that can home to cartilage. Additionally, members and
related members of the calcin family of peptides can also home to
cartilage, despite having a distinct tertiary structure from the
"hitchin" class of peptides. Calcin peptides are structurally a
subset of the cystine-dense peptides, with cystine-dense disulfide
connectivity and topology, but are further classified on the basis
of functioning to bind and activate ryanodine receptors (RyRs).
These receptors are calcium channels that act to regulate the
influx and efflux of calcium in muscle (Schwartz et al. Br J
Pharmacol 157(3):392-403. (2009)). Variants of the calcin family of
peptides with preserved key residues can be one way to predict
promising candidates that can home to cartilage. In some
embodiments, structural analysis of a peptide of this disclosure
can be determined by evaluating peptides for resistance to
degradation in buffers with various proteases or reducing agents.
Structural analysis of the distribution of charge density on the
surface of a peptide can also be a strategy for predicting
promising candidates that can home to cartilage. Peptides with
large patches of positive surface charge (when at pH 7.5) can home
to cartilage.
[0095] The NMR solution structures, x-ray crystallography, or
crystal structures of related structural homologs can be used to
inform mutational strategies that can improve the folding,
stability, and manufacturability, while maintaining the ability of
a peptide to home to cartilage. They can be used to predict the 3D
pharmacophore of a group of structurally homologous scaffolds, as
well as to predict possible graft regions of related proteins to
create chimeras with improved properties. For example, this
strategy can be used to identify critical amino acid positions and
loops that can be used to design drugs with improved properties or
to correct deleterious mutations that complicate folding and
manufacturability for the peptides. These key amino acid positions
and loops can be retained while other residues in the peptide
sequences can be mutated to improve, change, remove, or otherwise
modify function, homing, and activity of the peptide.
[0096] Additionally, the comparison of the primary sequences and
the tertiary sequences of two or more peptides can be used to
reveal sequence and 3D folding patterns that can be leveraged to
improve the peptides and parse out the biological activity of these
peptides. For example, comparing two different peptide scaffolds
that home to cartilage can lead to the identification of conserved
pharmacophores that can guide engineering strategies, such as
designing variants with improved folding properties. Important
pharmacophore, for example, can comprise aromatic residues or basic
residues, which can be important for binding.
[0097] Improved peptides can also be engineered based upon
immunogenicity information, such as immunogenicity information
predicted by TEPITOPE and TEPITOPEpan. TEPITOPE is a computational
approach which uses position specific scoring matrix to provide
prediction rules for whether a peptide will bind to 51 different
HLA-DR alleles, and TEPITOPEpan is method that uses TEPITOPE to
extrapolate from HLA-DR molecules with known binding specificities
to HLA-DR molecules with unknown binding specificities based on
pocket similarity. For example, TEPITOPE and TEPITOPEpan can be
used to determine immunogenicity of peptides that home to
cartilage. Immunogenicity information can also be predicted using
the program NetMHCII version 2. 3, which can determine the
likelihood that a sequence might be presented as an immunogenic
peptide via the major histocompatibility complex (MHC) presentation
system of antigen presenting cells (APCs). (Nielson, M et al. BMC
Bioinformatics, 8: 238 (2007); Nielsen, M. et al. BMC
Bioinformatics, 10: 296 (2009)). This program can create an
immunogenicity score by predicting the binding of a peptide to MHC
alleles. Strong binding alleles and weak binding alleles in each
major MHC allele group (DR, DQ, and DP) can be tallied separately.
The number of peptides of a specific length within the sequence
(e.g., a `core` peptide that can be nine residues long) that are
immunogenic can also be tallied. Comparison of peptides or `core`
peptides with high immunogenicity to peptides or `core` peptides
with low immunogenicity can guide engineering strategies for
designing variants with decreased immunogenicity. Stronger binding
peptides can be more likely to generate an immune response in
patient carrying that given MHC alleles. Mutating stronger binding
amino acids or peptides out of a peptide sequence can reduce the
immunogenicity of the entire peptide. Another aspect of
immunogenicity, in addition to whether a peptide binds to a
patient's MHC allele, can be whether the patient's immune cells,
such as a professional antigen presenting cells such as a
macrophage, a B cell, or a dendritic cell, can process the peptide.
A dendritic cell can take up a protein or peptide, and then can
process a peptide, such as by cleaving to form a nine residue long
peptide, which then can bind to the MHC and can be presented on the
surface of the dendritic cell to the immune system's various T
cells, including helper T cells and cytotoxic T cells, and thus can
stimulate an immune response. The processing can involve peptide
bond cleavage by enzymes and disulfide bond reduction, and thus a
peptide or protein that is resistant to enzymatic cleavage and/or
reduction can be resistant to processing and subsequent MHC
presentation to the immune system. Therefore, having a peptide or
protein that is resistant to enzymatic cleavage and/or reduction
can reduce its immunogenic potential.
[0098] Furthermore, multiple sequence alignment can also be used to
inform mutational strategies using previously identified sequences,
and thus providing a guide to making changes that would eliminate
labile residues and immunogenic regions of a peptide sequence.
Peptides can be evaluated for residues of potential biochemical
instability and regions of potential immunogenicity. Then, a
residue that can allow for greater peptide stability at a certain
location in a peptide can be identified from a multiple sequence
alignment. For example, a specific residue can be identified from a
multiple sequence alignment as providing greater stability for a
peptide at position previously identified as a possible risk for a
significant rate of deamidation, cleavage, degradation, oxidation,
hydrolysis, isomerization, disulfide exchange, racemization, beta
elimination, or aggregation. This information can then be used to
create peptides with greater stability or reduced
immunogenicity.
[0099] In addition to utilizing co-crystal x-ray structures, NMR
solution structures, and mutagenesis studies, a multiple alignment
of peptide sequences can be used to identify specific amino acids
or regions of high conservation that indicate an important
interaction with a target or receptor (e.g., binding to a potassium
channel protein) or are important for folding and structure or
other properties. Once the conserved amino acid or region is
identified, then amino acids replacements can be determined that
maintain the important properties of the peptide, such as
maintenance of the structure, reduction in immunogenicity,
reduction in binding to an ion channel protein, increased
stability, or any combination of thereof.
[0100] The multiple sequence alignment can also identify possible
locations to add a tyrosine or tryptophan residue for
spectrophotometric reporting. Incorporation of aromatic amino acids
such as Tyrosine or Tryptophan into a peptide such as SEQ ID NO:
108, which otherwise contains only amino acids of low UV absorbance
at 280 nm, can be analytically advantageous. Tyrosine and
Tryptophan amino acids contain aromatic ring structures. These
residues have distinct absorption and emission wavelengths and good
quantum yields, as shown in TABLE 2, not present in other amino
acids. Both Tyrosine and Tryptophan can provide a good `handle` for
analytical detection of a peptide in solution since UV absorbance
in the 250-300 nm range and peptide fluorescence is specific for
these aromatic molecules. While detection of a peptide such as SEQ
ID NO: 108 relies on the absorbance of the peptide bond at 220 nm,
where many other materials including minor impurities in solvents
also often contribute to signal, the absorbance and fluorescence
properties of Tryptophan and Tyrosine containing peptides can
provide for a significantly more selective and sensitive detection.
Thus incorporating an aromatic amino acid can create peptides
better suited for concentration and purity measurements, which can
be useful during analytics, process development, manufacturing, and
other drug development and drug manufacturing activities.
Incorporation can be achieved either through substitutions of one
or more amino acids in the peptide to Tyr and/or Trp, insertion of
Tyr and/or Trp into the peptide, or via addition of Tyr and/or Trp
to the N-terminus or C-terminus of the peptide.
TABLE-US-00028 TABLE 2 Absorbance and Fluorescence Characteristics
of Tryptophan and Tyrosine. Absorbance Fluorescence Wavelength
Absorbtivity Wavelength Quantum Amino Acid (nm) (M * cm).sup.-1
(nm) Yield Tryptophan 280 5,600 348 0.20 Tyrosine 274 1,400 303
0.14
[0101] A peptide of this disclosure can bind to chloride,
potassium, or sodium channels. The peptide can also bind to calcium
channels. The peptide can block potassium channels and/or sodium
channels. The peptide can block calcium channels. In some
embodiments, the peptide can activate any one or more of such
channels. In some embodiments, the peptide can block any one or
more of such channels. In some embodiments, the peptide cannot
interact with any of such channels or can be mutated to reduce or
remove binding to any such channels. In still other embodiments,
the peptide can be a potassium channel agonist, a potassium channel
antagonist, a portion of a potassium channel, a sodium channel
agonist, a sodium channel antagonist, a chloride channel agonist, a
chloride channel antagonist, a calcium channel agonist, a calcium
channel antagonist, a hadrucalcin, a theraphotoxin, a huwentoxin, a
kaliotoxin, a cobatoxin or a lectin. In some embodiments, the
lectin can be SHL-Ib2. In some embodiments, the peptide can
interact with, binds, inhibits, inactivates, or alters expression
of ion channels or chloride channels. In some embodiments, the
peptide can interact with an Nav1.7 ion channel. In some
embodiments, the peptide can interact with a Kv 1.3 ion channel. In
still other embodiments, the peptide interacts with proteases,
matrix metalloproteinase, inhibits cancer cell migration or
metastases, has antimicrobial activity, or has antitumor activity.
In addition to acting on matrix metalloproteinases, the peptide can
interact with other possible proteases (e.g., elastases). In some
embodiments, a peptide of this disclosure can bind to multidrug
resistance transporters. Peptide and peptide drug conjugate binding
to and blocking multidrug resistance transporters can be used to
treat bacterial infections or cancers of the joint and/or bone.
[0102] In some embodiments, the peptide has other therapeutic
effects on the cartilage or structures thereof or nearby. Beta
defensin expression in articular cartilage can be correlated with
immunomodulatory functions as we well as osteoarthritis, autoimmune
rheumatic disorders such as systemic lupus erythematosus and
rheumatoid arthritis (Vordenbaumen and Schneider 2011, Varoga 2004
and Varoga 2005). In some embodiments, the peptides or their
mutants inhibit beta defensins, supplement beta defensins, are
competitive inhibitors of beta defensins, active or block
activation of beta defensin targets, and are used as immune
modulators, or to treat autoimmune, arthritis, infections, and
other articular disorders.
[0103] The present disclosure can also encompass multimers of the
various peptides described herein. Examples of multimers include
dimers, trimers, tetramers, pentamers, hexamers, heptamers, and so
on. A multimer can be a homomer formed from a plurality of
identical subunits or a heteromer formed from a plurality of
different subunits. In some embodiments, a peptide of the present
disclosure is arranged in a multimeric structure with at least one
other peptide, or two, three, four, five, six, seven, eight, nine,
ten, or more other peptides. In certain embodiments, the peptides
of a multimeric structure each have the same sequence. In
alternative embodiments, some or all of the peptides of a
multimeric structure have different sequences.
[0104] The present disclosure further includes peptide scaffolds
that, e.g., can be used as a starting point for generating
additional peptides. In some embodiments, these scaffolds can be
derived from a variety of cystine-dense peptides. Some suitable
peptides for scaffolds can include, but are not limited to,
chlorotoxin, brazzein, circulin, stecrisp, hanatoxin, midkine,
hefutoxin, potato carboxypeptidase inhibitor, bubble protein,
attractin, .alpha.-GI, .alpha.-GID, .mu.-PIIA, .omega.-MVIIA,
.omega.-CVID, .chi.-MrIA, .rho.-TIA, conantokin G, contulakin G,
GsMTx4, margatoxin, shK, toxin K, chymotrypsin inhibitor (CTI), and
EGF epiregulin core.
[0105] In some embodiments, the peptide sequences of the disclosure
are flanked by additional amino acids. One or more additional amino
acids can, for example, confer a desired in vivo charge,
isoelectric point, chemical conjugation site, stability, or
physiologic property to a peptide.
[0106] Identifying sequence homology can be important for
determining key residues that preserve cartilage homing function.
For example, in some embodiments identification of conserved
positively charged residues can be important in preserving
cartilage homing in any homologous variants that are made. In other
embodiments, identification of basic or aromatic dyads, can be
important in preserving interaction and activity with Kv ion
channels in homologous variants.
[0107] Two or more peptides can share a degree of homology and
share similar properties in vivo. For instance, a peptide can share
a degree of homology with a peptide of the present disclosure. In
some cases, a peptide of the disclosure can have up to about 20%
pairwise homology, up to about 25% pairwise homology, up to about
30% pairwise homology, up to about 35% pairwise homology, up to
about 40% pairwise homology, up to about 45% pairwise homology, up
to about 50% pairwise homology, up to about 55% pairwise homology,
up to about 60% pairwise homology, up to about 65% pairwise
homology, up to about 70% pairwise homology, up to about 75%
pairwise homology, up to about 80% pairwise homology, up to about
85% pairwise homology, up to about 90% pairwise homology, up to
about 95% pairwise homology, up to about 96% pairwise homology, up
to about 97% pairwise homology, up to about 98% pairwise homology,
up to about 99% pairwise homology, up to about 99.5% pairwise
homology, or up to about 99.9% pairwise homology with a second
peptide. In some cases, a peptide of the disclosure can have at
least about 20% pairwise homology, at least about 25% pairwise
homology, at least about 30% pairwise homology, at least about 35%
pairwise homology, at least about 40% pairwise homology, at least
about 45% pairwise homology, at least about 50% pairwise homology,
at least about 55% pairwise homology, at least about 60% pairwise
homology, at least about 65% pairwise homology, at least about 70%
pairwise homology, at least about 75% pairwise homology, at least
about 80% pairwise homology, at least about 85% pairwise homology,
at least about 90% pairwise homology, at least about 95% pairwise
homology, at least about 96% pairwise homology, at least about 97%
pairwise homology, at least about 98% pairwise homology, at least
about 99% pairwise homology, at least about 99.5% pairwise
homology, at least about 99.9% pairwise homology with a second
peptide. Various methods and software programs can be used to
determine the homology between two or more peptides, such as NCBI
BLAST, Clustal W, MAFFT, Clustal Omega, AlignMe, Praline, or
another suitable method or algorithm.
[0108] In still other instances, the variant nucleic acid molecules
of a peptide of any one of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID
NO: 314-SEQ ID NO: 564 can be identified by either a determination
of the sequence identity or homology of the encoded peptide amino
acid sequence with the amino acid sequence of any one of SEQ ID NO:
24-SEQ ID NO: 274, SEQ ID NO: 314-SEQ ID NO: 564, or by a nucleic
acid hybridization assay. Such peptide variants can include nucleic
acid molecules (1) that remain hybridized with a nucleic acid
molecule having the nucleotide sequence of any one of SEQ ID NO:
24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564 (or any
complement of the previous sequences) under stringent washing
conditions, in which the wash stringency is equivalent to
0.5.times.-2.times.SSC with 0.1% SDS at 55-65.degree. C., and (2)
that encode a peptide having at least 70%, at least 80%, at least
90%, at least 95% or greater than 95% sequence identity or homology
to the amino acid sequence of any one SEQ ID NO: 24-SEQ ID NO: 274
or SEQ ID NO: 314-SEQ ID NO: 564. Alternatively, peptide variants
of any one SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID
NO: 564 can be characterized as nucleic acid molecules (1) that
remain hybridized with a nucleic acid molecule having the
nucleotide sequence of any one SEQ ID NO: 24-SEQ ID NO: 274 or SEQ
ID NO: 314-SEQ ID NO: 564 (or any complement of the previous
sequences) under highly stringent washing conditions, in which the
wash stringency is equivalent to 0.1.times.-0.2.times.SSC with 0.1%
SDS at 50-65.degree. C., and (2) that encode a peptide having at
least 70%, at least 80%, at least 90%, at least 95% or greater than
95% sequence identity or homology to the amino acid sequence of any
one of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO:
564.
[0109] Percent sequence identity or homology can be determined by
conventional methods. See, for example, Altschul et al., Bull.
Math. Bio. 48:603 (1986), and Henikoff and Henikoff, Proc. Natl.
Acad. Sci. USA 89:10915 (1992). Briefly, two amino acid sequences
are aligned to optimize the alignment scores using a gap opening
penalty of 10, a gap extension penalty of 1, and the "BLOSUM62"
scoring matrix of Henikoff and Henikoff (Id.). The sequence
identity or homology is then calculated as: ([Total number of
identical matches]/[length of the longer sequence plus the number
of gaps introduced into the longer sequence in order to align the
two sequences])(100).
[0110] Additionally, there are many established algorithms
available to align two amino acid sequences. For example, the
"FASTA" similarity search algorithm of Pearson and Lipman is a
suitable protein alignment method for examining the level of
sequence identity or homology shared by an amino acid sequence of a
peptide disclosed herein and the amino acid sequence of a peptide
variant. The FASTA algorithm is described by Pearson and Lipman,
Proc. Nat'l Acad. Sci. USA 85:2444 (1988), and by Pearson, Meth.
Enzymol. 183:63 (1990). Briefly, FASTA first characterizes sequence
similarity by identifying regions shared by the query sequence
(e.g., SEQ ID NO: 1) and a test sequence that has either the
highest density of identities (if the ktup variable is 1) or pairs
of identities (if ktup=2), without considering conservative amino
acid substitutions, insertions, or deletions. The ten regions with
the highest density of identities are then rescored by comparing
the similarity of all paired amino acids using an amino acid
substitution matrix, and the ends of the regions are "trimmed" to
include only those residues that contribute to the highest score.
If there are several regions with scores greater than the "cutoff"
value (calculated by a predetermined formula based upon the length
of the sequence and the ktup value), then the trimmed initial
regions are examined to determine whether the regions can be joined
to form an approximate alignment with gaps. Finally, the highest
scoring regions of the two amino acid sequences are aligned using a
modification of the Needleman-Wunsch-Sellers algorithm (Needleman
and Wunsch, J. Mol. Biol. 48:444 (1970); Sellers, Siam J. Appl.
Math. 26:787 (1974)), which allows for amino acid insertions and
deletions. Illustrative parameters for FASTA analysis are: ktup=1,
gap opening penalty=10, gap extension penalty=1, and substitution
matrix=BLOSUM62. These parameters can be introduced into a FASTA
program by modifying the scoring matrix file ("SMATRIX"), as
explained in Appendix 2 of Pearson, Meth. Enzymol. 183:63
(1990).
[0111] FASTA can also be used to determine the sequence identity or
homology of nucleic acid molecules using a ratio as disclosed
above. For nucleotide sequence comparisons, the ktup value can
range between one to six, preferably from three to six, most
preferably three, with other parameters set as described above.
[0112] Some examples of common amino acids that are a "conservative
amino acid substitution" are illustrated by a substitution among
amino acids within each of the following groups: (1) glycine,
alanine, valine, leucine, and isoleucine, (2) phenylalanine,
tyrosine, and tryptophan, (3) serine and threonine, (4) aspartate
and glutamate, (5) glutamine and asparagine, and (6) lysine,
arginine and histidine. The BLOSUM62 table is an amino acid
substitution matrix derived from about 2,000 local multiple
alignments of protein sequence segments, representing highly
conserved regions of more than 500 groups of related proteins
(Henikoff and Henikoff, Proc. Nat'l Acad. Sci. USA 89:10915
(1992)). Accordingly, the BLOSUM62 substitution frequencies can be
used to define conservative amino acid substitutions that may be
introduced into the amino acid sequences of the present invention.
Although it is possible to design amino acid substitutions based
solely upon chemical properties (as discussed above), the language
"conservative amino acid substitution" preferably refers to a
substitution represented by a BLOSUM62 value of greater than -1.
For example, an amino acid substitution is conservative if the
substitution is characterized by a BLOSUM62 value of 0, 1, 2, or 3.
According to this system, preferred conservative amino acid
substitutions are characterized by a BLOSUM62 value of at least 1
(e.g., 1, 2 or 3), while more preferred conservative amino acid
substitutions are characterized by a BLOSUM62 value of at least 2
(e.g., 2 or 3).
[0113] Determination of amino acid residues that are within regions
or domains that are critical to maintaining structural integrity
can be determined. Within these regions one can determine specific
residues that can be more or less tolerant of change and maintain
the overall tertiary structure of the molecule. Methods for
analyzing sequence structure include, but are not limited to,
alignment of multiple sequences with high amino acid or nucleotide
identity or homology and computer analysis using available software
(e.g., the Insight II..RTM. viewer and homology modeling tools;
MSI, San Diego, Calif.), secondary structure propensities, binary
patterns, complementary packing and buried polar interactions
(Barton, G. J., Current Opin. Struct. Biol. 5:372-6 (1995) and
Cordes, M. H. et al., Current Opin. Struct. Biol. 6:3-10 (1996)).
In general, when designing modifications to molecules or
identifying specific fragments determination of structure can
typically be accompanied by evaluating activity of modified
molecules.
[0114] Pairwise sequence alignment is used to identify regions of
similarity that may indicate functional, structural and/or
evolutionary relationships between two biological sequences
(protein or nucleic acid). By contrast, multiple sequence alignment
(MSA) is the alignment of three or more biological sequences. From
the output of MSA applications, homology can be inferred and the
evolutionary relationship between the sequences assessed. One of
skill in the art would recognize as used herein, "sequence
homology" and "sequence identity" and "percent (%) sequence
identity" and "percent (%) sequence homology" have been used
interchangeably to mean the sequence relatedness or variation, as
appropriate, to a reference polynucleotide or amino acid
sequence.
Chemical Modifications
[0115] A peptide can be chemically modified one or more of a
variety of ways. In some embodiments, the peptide can be mutated to
add function, delete function, or modify the in vivo behavior. One
or more loops between the disulfide linkages can be modified or
replaced to include active elements from other peptides (such as
described in Moore and Cochran, Methods in Enzymology, 503, p.
223-251, 2012). Amino acids can also be mutated, such as to
increase half-life or bioavailability, modify, add or delete
binding behavior in vivo, add new targeting function, modify
surface charge and hydrophobicity, or allow conjugation sites.
N-methylation is one example of methylation that can occur in a
peptide of the disclosure. In some embodiments, the peptide can be
modified by methylation on free amines. For example, full
methylation can be accomplished through the use of reductive
methylation with formaldehyde and sodium cyanoborohydride.
[0116] A chemical modification can, for instance, extend the
terminal half-life, the absorption half-life, the distribution
half-life of a peptide, change the biodistribution or
pharmacokinetic profile, or the modification itself can be useful
to provide viscosupplementation to a joint. A chemical modification
can comprise a polymer, a polyether, polyethylene glycol, a
biopolymer, a polyamino acid, a fatty acid, a dendrimer, an Fc
region, a simple saturated carbon chain such as palmitate or
myristolate, sugars, hyaluronic acid, or albumin. The chemical
modification of a peptide with an Fc region can be a fusion
Fc-peptide. A polyamino acid can include, for example, a polyamino
acid sequence with repeated single amino acids (e.g., polyglycine),
and a polyamino acid sequence with mixed polyamino acid sequences
(e.g., gly-ala-gly-ala (SEQ ID NO: 568)) that can or cannot follow
a pattern, or any combination of the foregoing.
[0117] In some embodiments, the peptides of the present disclosure
may be modified such that the modification increases the stability
and/or the half-life of the peptides. In some embodiments, the
attachment of a hydrophobic moiety, such as to the N-terminus, the
C-terminus, or an internal amino acid, can be used to extend
half-life of a peptide of the present disclosure. In other
embodiments, the peptide of the present disclosure can include
post-translational modifications (e.g., methylation and/or
amidation), which can affect, e.g., serum half-life. In some
embodiments, simple carbon chains (e.g., by myristoylation and/or
palmitylation) can be conjugated to the peptides. In some
embodiments, for example, the simple carbon chains may render
conjugated peptides easily separable from unconjugated material.
For example, methods that may be used to separate the desired
peptides of the invention from unconjugated material include, but
are not limited to, solvent extraction and reverse phase
chromatography. In some embodiments, lipophilic moieties can be
conjugated to the peptide and can extend half-life through
reversible binding to serum albumin. Moreover, the conjugated
moieties can be lipophilic moieties that extend half-life of the
peptides through reversible binding to serum albumin. In some
embodiments, the lipophilic moiety can be cholesterol or a
cholesterol derivative including cholestenes, cholestanes,
cholestadienes and oxysterols. In some embodiments, the peptides
can be conjugated to myristic acid (tetradecanoic acid) or a
derivative thereof. In other embodiments, the peptides of the
present disclosure are coupled (e.g., conjugated) to a half-life
modifying agent. Examples of half-life modifying agents include but
are not limited to: a polymer, a polyethylene glycol (PEG), a
hydroxyethyl starch, polyvinyl alcohol, a water soluble polymer, a
zwitterionic water soluble polymer, a water soluble poly(amino
acid), a water soluble polymer of proline, alanine and serine, a
water soluble polymer containing glycine, glutamic acid, and
serine, an Fc region, a fatty acid, palmitic acid, antibodies, or a
molecule that binds to albumin.
[0118] In some embodiments, the first two N-terminal amino acids
(GS) of SEQ ID NO: 1-SEQ ID NO: 274 can serve as a spacer or linker
in order to facilitate conjugation or fusion to another molecule,
as well as to facilitate cleavage of the peptide from such
conjugated or fused molecules. In some embodiments, the peptides of
the present disclosure can be conjugated to other moieties that can
modify or effect changes to the properties of the peptides.
Active Agent Conjugates
[0119] Peptides according to the present disclosure can be
conjugated or fused to a peptide biological agent or other agent
comprising amino acids (e.g., an antibody or antibody fragment,
receptor or receptor fragment, ligand or ligand fragment, hormone
or hormone fragment, growth factors and growth factor fragments,
biological toxins and fragments thereof, or other active portion of
a peptide), a protein, a peptide, or to a small molecule, RNA, DNA,
or other active agent molecular structure for use in the treatment
of cartilage diseases, disorders, or injuries. A peptide active
agent conjugate can be a peptide conjugated to an active agent by
any mechanism described herein. For example, a peptide can be
covalently conjugated to an active agent to form a peptide active
agent conjugate. A peptide can be chemically conjugated to an
active agent to form a peptide active agent conjugate. A peptide
and active agent can be expressed as a fusion protein to form a
peptide active agent conjugate. For example, an antibody or
fragment thereof and a peptide can be expressed as a fusion protein
to form a peptide active agent conjugate. For example, in certain
embodiments, a peptide as described herein can be fused to another
molecule, such as an active agent that provides a functional
capability. A peptide can be conjugated with an active agent
through expression of a vector containing the sequence of the
peptide with the sequence of the active agent. In various
embodiments, the sequence of the peptide and the sequence of the
active agent are expressed from the same Open Reading Frame (ORF).
In various embodiments, the sequence of the peptide and the
sequence of the active agent can comprise a contiguous sequence.
Various vectors and recombinant systems known in the art can be
employed to make such fusion peptides. The peptide and the active
agent can each retain similar functional capabilities in the fusion
peptide compared with their functional capabilities when expressed
separately.
[0120] Furthermore, for example, in certain embodiments, the
peptides described herein are attached to another molecule, such as
an active agent that provides a functional capability. In some
embodiments, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 active agents can be
linked to a peptide. Multiple active agents can be attached by
methods such as conjugating to multiple lysine residues and/or the
N-terminus, or by linking the multiple active agents to a scaffold,
such as a polymer or dendrimer and then attaching that
agent-scaffold to the peptide (such as described in Yurkovetskiy,
A. V., Cancer Res 75(16): 3365-72 (2015).
[0121] Described herein are active agents that can be conjugated to
the peptides of the present invention for use in either cartilage
disorders or kidney disorders, or both. In some embodiments,
certain compounds or drugs are appropriate for use in either
cartilage or kidney disorders, certain drug classes may be
preferred for specific treatment depending on the indication or
disorder. As described herein, it is understood that certain active
agents are described in a non-limiting exemplary manner for use in
treatments of cartilage and/or kidney indications. One or more of
such active agents can be conjugated to a peptide of the present
invention alone or in combination with one or more detectable
agents described herein. In some embodiments, active agents that
can be conjugated to any peptide of this disclosure can be
classified by mechanism. For example, active agents can belong to
the class of anti-inflammatory drugs, immunosuppressive (immune
suppression) drugs, analgesics/pain relief drugs, disease modifying
osteoarthritic drugs (DMOADs), cell depleting agents/apoptosis
modifiers, bone resorptive agents and viscosupplementing agents,
and tissue normalization (disease modifying) drugs.
[0122] Anti-inflammatory active agents can include, but are not
limited to, corticosteroids, glucocorticoids, nonsteroidal
anti-inflammatory drugs (NSAIDs), biologics, and other small
molecules. Examples of corticosteroid active agents that can be
conjugated to any peptide of this disclosure for delivery to the
joints and kidneys include triamcinolone dexamethasone, budesonide,
and triamcinolone acetonide. Examples of NSAID active agents that
can be conjugated to any peptide of this disclosure for delivery to
the joints and kidneys include naproxen and ibuprofen. Other active
agents include acetylsalicylic acid and acetaminophen. NSAID active
agents can be further classified into COX2 inhibitors. An example
of a COX2 inhibitor active agent directed to a prostaglandin
pathway that can be conjugated to any peptide of this disclosure
for delivery to the joint includes celecoxib. An example of a COX2
inhibitor active agent with anti-leukotriene receptor antagonist
that can be conjugated to any peptide of this disclosure for
delivery to the joint includes montelukast. An example of a COX2
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the kidneys includes iguratimod.
Biologic active agents can be further classified into active agents
that are IL-1 family inhibitors, IL-17 or IL-23 pathway inhibitors,
IL-6 family inhibitors, interferon receptor inhibitors, tumor
necrosis factor (TNF) inhibitors, RANK pathway inhibitors, B cell
inhibitors, anti-IgE active agents, and co-stimulation inhibitors.
An example of an IL-1 family inhibitor active agent that can be
conjugated to any peptide of this disclosure for delivery to the
joints includes anakinra. An example of an IL-17/IL-23 pathway
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the joints includes secukinumab. An
example of an IL-6 family inhibitor active agent that can be
conjugated to any peptide of this disclosure for delivery to the
kidneys includes sirukumab. An example of an interferon receptor
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the kidneys includes anifrolumab.
An example of a TNF inhibitor active agent that can be conjugated
to any peptide of this disclosure for delivery to the joints
includes infliximab or etanercept. An example of a RANK pathway
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the joints includes denosumab. An
example of a B cell inhibitor active agent that can be conjugated
to any peptide of this disclosure for delivery to the joints and
kidneys includes rituximab. An example of an anti-IgE active agent
that can be conjugated to any peptide of this disclosure for
delivery to the kidneys includes omalizumab. An example of a
co-stimulation inhibitor active agent that can be conjugated to any
peptide of this disclosure for delivery to the joints includes
abatacept.
[0123] Pain relief active agents can include, but are not limited
to analgesics, counter-irritants, and pain receptor blocking drugs.
Analgesics can be further classified into non-narcotic agents and
narcotic agents. An example of a non-narcotic active agent that can
be conjugated to any peptide of this disclosure for delivery to the
joints includes acetaminophen. An example of a narcotic active
agent that can be conjugated to any peptide of this disclosure for
delivery to joints includes oxycodone. Counter-irritant active
agents can be further classified as natural products. An example of
a natural product that can be conjugated to any peptide of this
disclosure for delivery to the joints includes capsaicin. Pain
receptor blocking active agents can be further classified as TRPV4
inhibitors. An example of a TRPV4 inhibitor active agent that can
be conjugated to any peptide of this disclosure for delivery to the
joints includes GSK2193874.
[0124] Apoptosis modifier active agents can include, but are not
limited to, biologics and small molecules. Biologic apoptosis
modifier active agents can be further classified as Fas/FasL
inhibitors, TNF/TNFR inhibitors, TRAIL/TRAILR inhibitors,
TWEAK/Fn14 inhibitors, IL-1 inhibitors, IL-1 receptor antagonists,
growth factors, and sclerostin inhibitors. An example of a TNF/TNFR
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the joints includes infliximab. An
example of a TRAIL/TRAILR inhibitor active agent that can be
conjugated to any peptide of this disclosure for delivery to the
joints includes osteoprotegrin. An example of a TWEAK/Fn14
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the kidneys includes BIIB023. An
example of an IL-1 receptor antagonist that can be conjugated to
any peptide of this disclosure for delivery to the joints includes
anakinra. An example of a growth factor active agent that can be
conjugated to any peptide of this disclosure for delivery to the
joints includes IGF-1. An example of a growth factor active agent
that can be conjugated to any peptide of this disclosure for
delivery to the kidneys includes EGF. An example of a sclerostin
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the joints includes romosozumab.
Small molecule apoptosis modifier active agents can be further
classified as caspase inhibitors, iNOS inhibitors, surfactants, and
bisphosphonates. An example of a caspase inhibitor active agent
that can be conjugated to any peptide of this disclosure for
delivery to the joints includes ZVAD-fmk. An example of an iNOS
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the joints include
S-methylisothiourea. An example of a surfactant active agent that
can be conjugated to any peptide of this disclosure for delivery to
the joints include P188. An example of a bisphosphonate active
agent that can be conjugated to any peptide of this disclosure for
delivery to the joints includes alendronate. Moreover, the known
class of drugs called senotherapeutics, also referred to as
senolytics or senolytic drugs or senolytic compounds, refers to
small molecules that can selectively induce death of senescent
cells and for example by directly or indirectly inducing apoptosis
in senescent cells. In addition, senolytics may also act via
non-apoptotic mechanisms of cell death including by necroptis,
autophagic cell death, pyroptis and caspase-independent cell death
(Journal of Cell Science 127; 2135-2144 (2014)). Such drugs can
attenuate age-related deterioration of tissues or organs. Examples
of drugs that can be conjugated to any peptide of this disclosure
to induce apoptosis or induce cell death via non-apoptotic
mechanisms include quercetin, dasatinib, bortezomib, carfilzomib,
and navitoclax amongst other compounds disclosed herein. Additional
active agents are described in the following references: Zhu, Y et
al., Aging Cell 14(4):644-58 (2015); Kirkland, J L, Exp Gerontol.
48(1): 1-5 (2013); Kirkland J and Tchkonia T, Exp Gereontol. 68:
19-25 (2015) Tchkonia, T et al., J Clin Invest., 123(3): 966-72
(2013); WO2016118859; Sugumar, D et al., Pharmagenomics Pers Med.
8: 23-33 (2015); Jiafa, R et al., Sci Rep. 6: 23968 (2016);
Swanson, C D et al., Nat Rev Rheumatol., 5(6): 317-324 (2009); Oh,
C J et al., PLoS One, 7(10):e45870 (2012); and Adebajo, A and
Boehncke, W, Psoriatic Arthritis and Psoriasis: Pathology and
Clinical Aspects, Springer (2016).
[0125] Tissue normalization (disease modifying) active agents can
include, but are not limited to, biologics and small molecules.
Biologic active agents can be further classified as chemokines
(e.g., for stem cell recruitment) and growth factors. An example of
a tissue normalization chemokine active agent that can be
conjugated to any peptide of this disclosure for delivery to the
joints includes MIP-3a. An example of a tissue normalization growth
factor active agent that can be conjugated to any peptide of this
disclosure for delivery to the joints includes BMP-2. Small
molecule active agents can be further classified as flavonoids, ACE
inhibitors, and anti-proliferative active agents. An example of a
tissue normalization flavonoid active agent that can be conjugated
to any peptide of this disclosure for delivery to the joints
includes icariin. An example of a tissue normalization ACE
inhibitor active agent that can be conjugated to any peptide of
this disclosure for delivery to the kidneys includes captopril. An
example of a tissue normalization anti-proliferative active agent
that can be conjugated to any peptide of this disclosure for
delivery to the joints includes methotrexate.
[0126] TABLE 3 describes active agents for treatment of a cartilage
disorder that can be conjugated to any peptide of the present
disclosure to form peptide-drug conjugates.
TABLE-US-00029 TABLE 3 Exemplary Active Agents for Cartilage
Disorders Active Agent Class Active Agent Gold compound Gold Gold
compound Auranofin Gold compound Gold Sodium Thiomalate Gold
compound Gold Thioglucose Gold compound Thiomalic Acid Gold
compound Gold Thiosulphate Analgesics Tramadol (e.g., Ultram,
Ultracet) and derivatives Analgesics Oxycodone (e.g., Percocet,
Oxycontin) and derivatives Analgesics Hydrocodone (e.g., Norco,
Vicoprofen) Analgesics Morphine Analgesics Fentanyl Analgesics
Oxymorphone Analgesics Hydromorphone Analgesics Meperidine
Analgesics Buprenorphine Analgesics Methadone Bisphosphonate
Alendronate Bisphosphonate Ibandronate Bisphosphonate Risedronate
Bisphosphonate Pamidronate Bisphosphonate Zoledronate Non-Nitrogen
Containing First Clodronate Generation Bisphosphonate Non-Nitrogen
Containing First Etidronate Generation Bisphosphonate Non-Nitrogen
Containing First Tiludronate Generation Bisphosphonate Apoptosis
Inhibitors Osteoprotegerin (OPG) Sclerostin Antagonist Apoptosis
AMG785 (Romosozumab) Inhibitors Caspase-1 ICE Inhibitors VX-740
(Pralnacasan) Counter-irritants Menthol Counter-irritants Capsaicin
RANKL Targeting Agents Denosumab Cathepsin K Targeting Agents
Odanacatib TNF-.alpha. Antagonists CDP571 TNF-.alpha. Antagonists
ISIS 104838 Anti-Pain Drugs Duloxetine Polymers Low Molecular
Weight Chitosan Matrix Drugs Chondroitin sulfate glucosamine
Cytokines/Growth Factors TGF-beta Matrix Laminin Matrix Fibronectin
Matrix Lubricin Matrix Hyaluronic acid injections Matrix
Glucosamine Immunosuppressants Rapamycin HIF-1.alpha. Modulators
HIF-2.alpha. Modulators Corticosteroid Tixocortol pivalate
Glucocorticoid Corticosteroid Hydrocortisone Acetate Glucocorticoid
Corticosteroid Hydrocortisone t-Butyl Acetate Glucocorticoid
Corticosteroid Prednisolone Acetate Glucocorticoid Corticosteroid
Prednisolone t-Butyl Acetate Corticosteroid Dexamethasone Acetate
Corticosteroid Dexamethasone t-Butyl Acetate Glucocorticoid
Corticosteroid Triamcinolone Diacetate
[0127] TABLE 4 describes active agents for treatment of a kidney
disorder that can be conjugated to any peptide of the present
disclosure to form peptide-drug conjugates.
TABLE-US-00030 TABLE 4 Exemplary Active Agents for Kidney Disorders
Active Agent Class Active Agent ACE Inhibitors Captopril
Angiotensin receptor Angiotensin receptor blocker losartan blockers
(Cozaar) Hormones Adrenocorticotropic hormone Hormones
corticotropin-releasing hormone amphotericin B digitalis glycosides
potassium-depleting diuretics Coumarine anticoagulants NLRP3
Inflammosome MCC950 Targeted Drugs NLRP3 Inflammosome BHB Targeted
Drugs NLRP3 Inflammosome Type I interferon Targeted Drugs NLRP3
Inflammosome IFN-beta Targeted Drugs NLRP3 Inflammosome Resveratrol
Targeted Drugs NLRP3 Inflammosome Arglabin Targeted Drugs NLRP3
Inflammosome CB2R agonist Targeted Drugs NLRP3 Inflammosome
MicroRNA-223 Targeted Drugs
[0128] TABLE 5 describes active agents for treatment of a cartilage
disorder and a kidney disorder that can be conjugated to any
peptide of the present disclosure to form peptide-drug
conjugates.
TABLE-US-00031 TABLE 5 Exemplary Active Agents for Cartilage
Disorders and Kidney Disorders Active Agent Class Active Agent IL-6
Receptor Modulators Tocilizumab IL-6 Receptor Modulators Sarilumab
IL-6 Receptor Modulators ALX-0061 IL-6 Receptor Modulators
Sirukumab IL-6 Receptor Modulators Clazakizumab IL-6 Receptor
Modulators Olokizumab IL-6 Receptor Modulators MEDI5117 IL-17
Antagonists Secukinumab IL-17 Antagonists Brodalumab IL-17
Antagonists Ixekizumab Antagonists of p40 Subunit of IL-
Ustekinumab 12/IL-23 Antagonists of p40 Subunit of IL- Briakinumab
12/IL-23 Antagonists of p19 Subunit of IL-23 Tildrakizumab
Antagonists of p19 Subunit of IL-23 Guselkumab IL-23 Antagonists
Soluble IL-23 (or cytokine-binding homology region of soluble
IL-23) IL-1 Antagonists Canakinumab IL-1 Antagonists Rilonacept
IL-1 Antagonists Gevokizumab IL-1 Antagonists LY2189102 IL-1
Antagonists Lentiviral-mediated RNAi IL-12 Antagonists IL-1
Receptor Antagonists Anakinra IL-1 Receptor Antagonists MEDI-8968
IL-1 Receptor Antagonists AMG-108 IL-1 Receptor Kineret
Interleukins/Pro-Inflammatory Pro-inflammatory IL-1.alpha. or
IL-1.beta. Cytokines Interleukins IL-8 Interleukins IL-15
Interleukins IL-18 Interleukins IL-4 Interleukins IL-10
Interleukins IL-13 Interleukins IL-22 Interleukins IL-17 p38
Inhibitors VX-745 p38 Inhibitors BIRB 796 p38 Inhibitors SCIO-469
p38 Inhibitors VX-702 p38 Inhibitors Pamapimod p38 Inhibitors
ARRY-797 Corticosteroids 17-monopropionate Corticosteroids
Desciclesonide Corticosteroids Flunisolide Corticosteroids
Mometasone furoate Corticosteroids 22-hydroxy intermediate
budesonide derivative Corticosteroids 6.beta.-hydroxy budesonide
derivative Corticosteroids .DELTA.6-budesonide derivative
Corticosteroids 23-hydroxy budesonide derivative Corticosteroids
16.alpha.-butryloxyprednisolone budesonide derivative
Corticosteroids 16.alpha.-hydroxyprednisolone budesonide derivative
Corticosteroid (Beclomethasone) QVAR inhalation Corticosteroid
(Budesonide) pulmicort respules Corticosteroid Flovent HFA 44
Corticosteroid (Mometasone) Asmanex HFA Corticosteroid (Mometasone)
Budesonide symbicort Corticosteroid Dexamethasone sodium phosphate
Corticosteroid Tixocortol pivalate Corticosteroid Ciclesonide
Glucocorticoids 21-nortriamcincolone acetonide Glucocorticoids
.DELTA.6-triamcinolone Glucocorticoids 6b-hydroxy triamcinolone
acetonide Glucocorticoids 21-carboxy triamcinolone acetonide
Glucocorticoids 6b-OH, 21-COOH triamcinolone acetonide
Glucocorticoids 6.alpha. fluorocortisol Glucocorticoids 9.alpha.
fluorocortisol Glucocorticoids .DELTA.1-dehydro configuration in
prednisolone Glucocorticoids 16-methylene dexamethasone derivative
Glucocorticoids 16.alpha.-methyl dexamethasone derivative
Glucocorticoids 16.beta.-methyl betamethasone derivative
Glucocorticoids Cyclophosphamide Glucocorticoids Mycophenolate
Glucocorticoids/Mineralocorticoids Cortisol
Glucocorticoids/Mineralocorticoids Hydrocortisone
Glucocorticoids/Mineralocorticoids Prednisolone
Glucocorticoids/Mineralocorticoids Betamethasone Glucocorticoid
Fluticasone Glucocorticoid Fluticasone propionate Steroid
(flunisolide) Aerobid Steroid (flunisolide) Aerobid-M Steroid
(flunisolide) Aerospan Steroid (Flunisolide) Fluticasone Furoate
Steroid (Fluticasone) Flovent HFA 110 Steroid (Fluticasone) Flovent
HFA 220 Steroid (Fluticasone) Flovent Diskus 50 Steroid
(Fluticasone) Asmanex Steroid Betamethasone acetate Steroid
Betamethasone sodium phosphate Steroid Betamethasone valerate
Steroid Beclomethasone dipropionate Local Anesthetic procaine
hydrochloride Local Anesthetic Novacain Anesthetic bupivacaine
hydrochloride Anesthetic lidocaine hydrochloride Local Anesthetic
ropivacaine hydrochloride Analgesics Morphine Analgesics Fentanyl
Quinazolines Feitinib/Iressa Quinazolines Sorafenib/Nexavar
Quinazolines Lapatinib ditosylate/Tykerb/Tyverb Quinazolines
Sunitinib/Sutent Quinazolines Bortezomib/Velcade/Cytomib
Quinazolines Everolimus/Temsirolimus Quinazolines Inhibitors of
IAPS Quinazolines Activators of caspase pathway Quinazolines
Activators of AKT pathway Quinazolines Propylpeptidase inhibitors
Quinazolines Activators of p53 Quinazolines Inhibitors of
anti-apoptotic protein inhibitors Prolyl Hydroxylase (PHD)
Desferrioxamine Inhibitors Prolyl Hydroxylase (PHD)
Dimethyloxalylglycine (DMOG) Inhibitors Prolyl Hydroxylase (PHD)
L-mimosine (L-mim) Inhibitors Aptamers Peptide aptamers Aptamers
RNA aptamer A-p50 Aptamers Peptide A aptamer TrxLef1D Aptamers
Aptamer E07 Aptamers Aptamer gemcitabine polymers Aptamers RAGE
Aptamers Pegaptanib Proteosome Inhibitors Bortezomib Proteosome
Inhibitors Carfilzomib Second Generation Proteosome Ixazomib
Inhibitors Second Generation Proteosome Delanzomib Inhibitors
Second Generation Proteosome Oprozomib Inhibitors Second Generation
Proteosome Marizomib Inhibitors Apoptosis Inhibitors FLIP agonist
Apoptosis Inhibitors nitric oxide synthase inhibitors Apoptosis
Inhibitors caspase-3 inhibitors (Z-DEVD-fmk (SEQ ID NO: 569))
Apoptosis Inhibitors caspase-9 inhibitors (Z-LEHD-fmk (SEQ ID NO:
570)) Apoptosis Inhibitors Sclerostin antagonists Apoptosis
Inhibitors/Growth Factor IGF-1 BCL-2 Agonist Apoptosis Inhibitors
Oblimersen BCL-2 Agonist Apoptosis Inhibitors Obatoclax BCL-2
Agonist Apoptosis Inhibitors Navitoclax BCL-2 Agonist Apoptosis
Inhibitors Venetoclax (ABT-199) BCL-2 Agonist Apoptosis Inhibitors
Navotoclax (ABT-263) BCL-2 Agonist Apoptosis Inhibitors GX01 series
of compounds BCL-2 Agonist Apoptosis Inhibitors BCL-2 small
molecule antagonists BCL-2 Agonist Apoptosis Inhibitors
Tetraocarcin-A derivatives BCL-2 Agonist Apoptosis Inhibitors
Chelerythrine BCL-2 Agonist Apoptosis Inhibitors Antimycin A
derivatives BCL-2 Agonist Apoptosis Inhibitors HA14-1 BCL-2 Agonist
Apoptosis Inhibitors Synthetic compound antagonist of BH3 BCL-2
Agonist Apoptosis Inhibitors Genasense BCL-2 Agonist Apoptosis
Inhibitors ISIS 22783 BCL-2/BCL-XL Agonist Apoptosis Bispecific
Antisense Inhibitors Proapoptotic BCL-2 Targeting Bax, Bak, Bid,
Bad-derived BH3 Peptides Drugs Proapoptotic BCL-2 Targeting SAHBs
Drugs Proapoptotic BCL-2 Targeting BH3Is Drugs BCL-2/BCL-XL Agonist
Apoptosis ABT-737 Inhibitors BCL-X Inhibitors Apoptosis Modifiers
Caspase-1 Inhibitors Apoptosis Modifiers Caspase-8 Inhibitors
Pan-caspase Caspase Inhibitor IDN-6556 Pan-caspase Caspase
Inhibitor IDN-6734 Pan-caspase Caspase Inhibitor VX-799 Pan-caspase
Inhibitor MX1013 Pan-caspase Caspase Inhibitor M-920 Pan-caspase
Caspase Activator MX-2060 derivatives Pan-caspase Caspase
Activators Small-molecule compounds Pan-caspase Caspase Activators
RGD peptides Pan-caspase inhibitors ZVAD-fmk Caspase-1 ICE
Inhibitors IDN-11104 Caspase-1 ICE Inhibitors VX-756 Caspase-3
Inhibitors M-826 Caspase-3 Inhibitors M-791 Caspase-3 Inhibitors
Immunocasp-3 Caspase-3 Inhibitors Ad-G/iCasp3 Caspase-3 Inhibitors
PEF-F8-CP3 Caspase-6 Inhibitors Immunocasp-6 Caspase-9 Inhibitors
FKBP12/caspase-9 fusion protein IAP Antagonists BIR3 antagonists
XIAP Antagonists Capped tripeptide XIAP Antagonists XIAP
Antagonists Smac-mimetic compounds XIAP Antagonists
AEG35156/GEM.RTM.640 XIAP Inhibitors Embelin XIAP Inhibitors XIAP
antisense and RNA constructs XIAP/cIAP-1/cIAP-2 Inhibitors Small
molecule SMAC mimetics IAP/Caspase Inhibitors
HIV-Tat/polyarginine-conjugated SMAC peptides BIR2/Caspase-3
Inhibitors TWX024 BIR2 Inhibitors Polyphenylurea derivatives
Survivin Targeting Drugs LY2181308 Survivin Targeting Drugs
Ad-Survivin T34A Anti-TWEAK Apoptosis Modifiers BIIB023 Xanthine
Oxidase Inhibitors Allopurinol Xanthine Oxidase Inhibitors
Febuxostat Xanthine Oxidase Inhibitors Zyloprin Growth Factor bFGF
Growth Factor IGF Growth Factor TFG-beta Growth Factor BMP-2 Growth
Factor BMP-9 Growth Factor BMP-13 Growth Factor BMP-7 Growth Factor
BMP-3 inhibitors Growth Factor TFG-.beta.1 Growth Factor OP-1
Growth Factor PDGF Growth Factor PTH Growth Factor PTHrP Growth
Factor MIP-3.alpha. Growth Factor EPO Growth Factor FGF Growth
Factor FGF-2 Growth Factor FGF-18 Growth Factor TGF-.beta.3 Growth
Factor VEGF Growth Factor Wnt proteins Growth Factor EGF Growth
Factor GM-CSF Flavonoid Icariin Flavonoid Quercetin Tyrosine Kinase
Inhibitor (Lck/Btk Dasatinib Inhibitor) TRPV4 Activators
GSK1016790A TRPV4 Activators 4alpha-PDD TRPV4 Inhibitors HC-067047
TRPV4 Inhibitors GSK2193874 NSAID Ampion NSAID Phenylbutazone NSAID
Naproxen lysozyme conjugate NSAID Acetal salicylic acid DMARDs
Sulfasalazine DMARDs Leflunomide DMARDs Hydroxychloroquine
(Plaquenil) Disease-Modifying Osteoarthritis FGF-18 Drugs (DMOADs)
Uricosurics Sulfinpyrazone MSC Matrix Collagen MSC Matrix
Fibrin
MSC Matrix Polylactatous Surfactant P188 and other surfactants
Molecules for Bone Marrow Niches Angiopoetin Molecules for Bone
Marrow Niches Bone morphogenitic proteins Molecules for Bone Marrow
Niches Epinephrine Molecules for Bone Marrow Niches Norepinephrine
Molecules for Bone Marrow Niches GDF5 Molecules for Bone Marrow
Niches ICAN1 Molecules for Bone Marrow Niches Jagged1 Molecules for
Bone Marrow Niches Osteopontin Molecules for Bone Marrow Niches
parathyoid hormone Molecules for Bone Marrow Niches Calcitonin
Molecules for Bone Marrow Niches steel factor Molecules for Bone
Marrow Niches Thrombopoetin Molecules for Bone Marrow Niches
vascular cell adhesion molecule 1 Chemokine Molecules for Bone
CXCL12 Marrow Niches B Cell Targeting Agents Rituximab B Cell
Targeting Agents BLys B Cell Targeting Agents TACI T Cell
Co-stimulation Antagonists Abatacept JAK Targeting Agents
Tofacitinib Calcineurin Inhibitors Tacrolimus Calcineurin
Inhibitors Cyclosporin Calcineurin Inhibitors Voclosporin COX-2
Inhibitors Iguratimod COX-2 Inhibitors Montelukast COX-2 Inhibitors
Rofecoxib COX-2 Inhibitors Valdecoxib Interferon Receptor
Inhibitors Anifrolumab IFN-.alpha. Inhibitors Sifalimumab Anti-IgE
Agents Omalizumab iNOS Inhibitors S-methylisothiourea CD20
Antagonists/B Cell Inhibitors Ocrelizumab BAFF Antagonists/B Cell
Inhibitors Belimumab TNF Superfamily BAFF and APRIL Atacicept
Antagonists/B cell Inhibitors TNF-.alpha. Antagonists Thalidomide
TNF-.alpha. Antagonists Lenalidomide TNF-.alpha. Antagonists
Pomalidomide TNF-.alpha. Antagonists Pentocifylline TNF-.alpha.
Antagonists Bupropion TNF Antagonists Lentiviral-mediated RNAi TNF
Agonists Recombinant TNF-.alpha. TRAIL Receptor Agonists HGS-ETR1
TRAIL Receptor Agonists HGS-ETR2 TRAIL Receptor Agonists HGS-TR2J
TRAIL Receptor Agonists PRO1762 TRAIL Receptor Agonists TRA-8
CD95/Fas Agonists CD95-Fc Marine Bioactive Compounds
TRAIL-Resistance Overcoming Marine Bioactive Compounds Marine
Bioactive Compounds mazamine A Marine Bioactive Compounds
marine-derived chroomycins Marine Bioactive Compounds Carotenoids
Marine Bioactive Compounds Aplysin Marine Bioactive Compounds
Aplidin Marine Bioactive Compounds Siphonaxanthin Marine Bioactive
Compounds pectinotoxin-2 Anti-Complement Drugs Eculizumab PAR-2
Modulators Pepducin P2pal-18 miR-2013 Blockers Anti-sense
oligonucleotides Nrf2 Activator Dimethyl fumarate p53 Targeting
Drugs INGN201 p53 Targeting Drugs SCH58500 p53 Targeting Drugs
ONYX-015 p53 Targeting Drugs C-terminal p53 peptides p53 Targeting
Drugs CDB3 p53 Targeting Drugs CP31398 p53 Targeting Drugs Prima-1
p53 Targeting Drugs HPV E6-binding peptide aptamers p53 Targeting
Drugs Nutlins p53 Targeting Drugs Chalcones p53 Targeting Drugs
Small peptides p53 Targeting Drugs Pifithrin-.alpha. p53 Targeting
Drugs/Apoptosis QP1-1002 Modifiers (T cells) Apaf-1 Targeting
Drugs/Apoptosis QM56 Modifiers (T cells) Apaf-1 Targeting
Drugs/Apoptosis SVT016426 Modifiers (T cells) Ferrostatin 16/86
BASP1 Targeting Drugs/Apoptosis BASP siRNA Modifiers (T cells)
Anti-Inflammatory Drugs CCX140 Anti-Inflammatory Drugs CXA-10
Anti-Inflammatory Drugs/Anti- Alkaline phosphatase Fibrotic Drugs
Anti-Fibrotic Drugs Dnmt1 inhibitors Anti-Inflammatory
Drugs/Apoptosis THR-184 Modifiers (T cells) Immunosuppressants
Lithium .beta.2-Adrenergic Agonists Formoterol Anti-Inflammatory
Drugs CRMD-001 Endothelin-1 Targeting Drugs Astrasentan Vasopressin
Receptor Antagonists Tolvaptan Vasopressin Receptor Antagonists
RWJ-676070 Immunosuppressants Azathioprine Immunosuppressants
Mycophenolic acid Immunosuppressants Cyclosporine Immune Modulators
Laquinimod Slow-acting antirheumatic drugs (SAARDs) Colcrys
Hormones parathyroid hormone Hormones growth hormone 11-beta
hydroxysteroid dehydrogenases Mineralocorticoid Proopiomelanocortin
fludrocortisonesoxycorticosterone acetate vaccines from live
attenuated viruses Aspirin Insulin Isonizaid Oral hypoglycemic
agents Antacids Carbamazepine Cholestyramine Colestipol Ephedrine
Erythromycin Mitotane oral contraceptives Phenobarbital Phenytoin
Rifampin Troleandomycin Non-selective caspase inhibitor okadaic
acid Camptothetic Staurosporine HFA Alvesco inhalation Breo Ellipta
Advair Mometasone Dulera Umeclidinium Anoro Reactive Oxygen Species
Targeting Drugs Cytokines/Growth Factors TGF-beta NOD-like receptor
protein 3- dependent caspase 1 Targeting Drugs NSAID Etoricoxib
Apoptosis Modifiers MCL1 inhibitors Teriparatide BH3 mimetics AZD
4320 Carrier Proteins Low molecular weight human serum albumin
Ceramide Targeting Drugs DMARDs Penicillamine Chondrogenic factors
Anti-oxidative factors A(1)AR agonist S1P(2)R antagonist
Antimalarials BAX/BAK activating drugs Selective GR Activators
(SEGRAs) Rap1 Targeted Drugs Senolytic Ephrin Ligand (EFN) B1
blockers Senolytic Cyclin-dependent kinase inhibitor 1A (p21)
phosphatidylinositol-4,5-bishophate 3-kinase delta catlyatic
subunit (PI3KCD) blockers Senolytic Plasminogen-activated
inhibitor-2 (PAI-2) blockers Senesce-associated secretory phenotype
(SASP) inhibitors Hormone Tetracosactide
[0129] TABLE 6 describes additional active agents for treatment of
a cartilage disorder and a kidney disorder that can be conjugated
to any peptide of the present disclosure to form peptide-drug
conjugates.
TABLE-US-00032 TABLE 6 Exemplary Active Agents for Cartilage
Disorders and Kidney Disorders Active Agent Class Active Agent
Peptide Oligopeptide Peptide Polypeptide Peptide Peptidomimetic
Nucleic Acid Polynucleotide Nucleic Acid Polyribonucleotide Nucleic
Acid Oligonucleotide Nucleic Acid DNA Nucleic Acid cDNA Nucleic
Acid ssDNA Nucleic Acid RNA Nucleic Acid dsRNA Nucleic Acid micro
RNA Nucleic Acid Interfering RNA Nucleic Acid Aptamer Antibody
single chain variable Fragment (scFv) Antibody Antibody Fragment
Antibody Aptamer Antibody Fc domains Antibody Fc regions Antibody
Fc active fragments or modifications thereof Cytokine Cytokine
antagonists Mavrilimumab Cytokine antagonists Ixekizumab Cytokine
antagonists Tocilizumab Cytokine antagonists Anakinra Cytokine
antagonists Ustekinumab Cytokine antagonists Secukinumab Interferon
Hormone Enzymes Growth Factor Checkpoint Inhibitor CD Antigen
Chemokines Neurotransmitters Ion Channel Inhibitors G-protein
coupled receptor inhibitors G-protein coupled receptor activators
Tumor necrosis factor inhibitors Chemical Agents Radiosensitizers
Radioprotectants Radionuclide Therapeutic Small Molecules Steroids
Corticosteroids Anti-inflammatory Agents Immune Modulators
Abatacept Immune Modulators Rituximab Complement Fixing Peptides or
Proteins Tumor Necrosis Factor Family Tumor Necrosis Factor (TNF)
soluble receptor or Inhibitors antibody Tumor Necrosis Factor
Family Activators Tumor Necrosis Factor (TNF) soluble receptor or
antibody Caspase protease inhibitors or activators NF-kB, RIPK1
and/or RIPK3 Inhibitors NF-kB, RIPK1 and/or RIPK3 Activators
Death-receptor ligand activator or inhibitor Tumor Necrosis Factor
Family TNFR1 Agonists Tumor Necrosis Factor Family TNFR2 Agonists
Tumor Necrosis Factor Family CD27/TNFRSF7 Agonists Tumor Necrosis
Factor Family CD30/TNFRSF8 Agonists Tumor Necrosis Factor Family
OX40/TNFRSF4 Agonists Tumor Necrosis Factor Family CD40/TNFRSF5
Agonists Tumor Necrosis Factor Family 4-1BB/TNFRSF9 Agonists Tumor
Necrosis Factor Family RANK (receptor activator of NF-kappa
Agonists B/TNFRSF11A) Tumor Necrosis Factor Family TWEAK
receptor/TNFRSF12A Agonists Tumor Necrosis Factor Family
TAC1/TNFRSF13B Agonists Tumor Necrosis Factor Family BAFF-R (BAFF
receptor/TNFRSF13C) Agonists Tumor Necrosis Factor Family HVEM
(herpes virus entry mediator/TNFRSF14) Agonists Tumor Necrosis
Factor Family RELT/TNFRSF19L Agonists Tumor Necrosis Factor Family
ectodysplasin A2 isoform receptor/TNFRS27 Agonists Tumor Necrosis
Factor Family ectodysplasin A1 Agonists TNF Family Member
Anhidrotic Receptor Tumor Necrosis Factor Family Decoy Receptor
3/TNFRSF6B Antagonists Tumor Necrosis Factor Family Decoy Receptor
1/TNFRSF10C Antagonists Tumor Necrosis Factor Family Decoy Receptor
2/TNFRSF10D Antagonists Tumor Necrosis Factor Family DR3 (death
receptor 3/TNFRSF25) Antagonists Tumor Necrosis Factor Family DR4
(death receptor 4/TNFRSF10A) Antagonists Tumor Necrosis Factor
Family DR5 (death receptor 5/TNFRSF10B) Antagonists Tumor Necrosis
Factor Family DR6 (death receptor 6/TNFRSF21) Antagonists Tumor
Necrosis Factor Family Fas/TNFRSF6 Antagonists Tumor Necrosis
Factor Family Lymphotoxin b receptor/TNFRS3 Antagonists Tumor
Necrosis Factor Family OPG (osteoprotegerin/TNFRSF11B) Antagonists
Tumor Necrosis Factor Family Nerve Growth Factor Receptor/TNFRSF16
Antagonists Tumor Necrosis Factor Family BCMA (B Cell Maturation
Antigen/TNFRSF17) Antagonists Tumor Necrosis Factor Family GITR
(Glucocorticoid-Induced TNF Receptor/ Antagonists TNFRSF18) Tumor
Necrosis Factor Family TAJ (Toxicity and JNK Inducer/TNFRSF19)
Antagonists Tumor Necrosis Factor Family TNFRSF22 Antagonists Tumor
Necrosis Factor Family TNFRSF23 Antagonists TNF Receptor
Superfamily Ligands TNF alpha TNF Receptor Superfamily Ligands
Lymphotoxin-a TNF Receptor Superfamily Ligands Tumor Necrosis
Factor Membrane Form TNF Receptor Superfamily Ligands Tumor
Necrosis Factor Shed Form TNF Receptor Superfamily Ligands LIGHT
TNF Receptor Superfamily Ligands Lymphotoxin b2a1 heterotrimer TNF
Receptor Superfamily Ligands OX-40 Ligand TNF Receptor Superfamily
Ligands Compound 1 [PMID: 24930776] TNF Receptor Superfamily
Ligands CD40 Ligand TNF Receptor Superfamily Ligands Fas Ligand TNF
Receptor Superfamily Ligands TL1A TNF Receptor Superfamily Ligands
CD70 TNF Receptor Superfamily Ligands CD30 Ligand TNF Receptor
Superfamily Ligands TRAF1 TNF Receptor Superfamily Ligands TRAF2
TNF Receptor Superfamily Ligands TRAF3 TNF Receptor Superfamily
Ligands TRAIL TNF Receptor Superfamily Ligands RANK Ligand TNF
Receptor Superfamily Ligands APRIL TNF Receptor Superfamily Ligands
BAFF TNF Receptor Superfamily Ligands B and T lymphocyte
Attenuators TNF Receptor Superfamily Ligands NGF TNF Receptor
Superfamily Ligands BDNF TNF Receptor Superfamily Ligands
Neurotrophin-3 TNF Receptor Superfamily Ligands Neurotrophin-4 TNF
Receptor Superfamily Ligands TL6 TNF Receptor Superfamily Ligands
Ectodysplasin A2 TNF Receptor Superfamily Ligands Ectodysplasin A1
TNF blockers Remicade (infliximab) TNF blockers Enbrel (etanercept)
TNF blockers Humira (adalimumab) TNF blockers Cimzia (certolizumab
pegol) TNF blockers Simponi (golimumab) Tumor Necrosis Factor
Receptor Family Agonists Toll Like Receptors Agonist TIMP-3
Inhibitors BCL-2 Family Inhibitors IAP Disruptors Protease
Inhibitors Amino Sugars Chemotherapeutic Cytotoxic chemical Toxins
Tyrosine Kinase inhibitors Imatinib Mesylate Protons Antivascular
Agents Bevacizumab EGFR Inhibitors Erlotinib Anti-Infective Agents
Antibiotics Anti-Viral Agents Anti-Fungal Agents Aminoglycoside
Statins Nanoparticles Liposomes Polymers Biopolymers Polysaccharide
Proteoglycan Glycosaminoglycans Polyethylene glycol Lipids
Dendrimers Fatty Acids Glucocorticoid Corticosteroid Collagenase
Inhibitor Matrix Metalloprotease Inhibitors MMP-13 inhibitor
Vitamins Vitamin D Antibiotics Antiviral Antifungal Statins Immune
Modulators Radioisotopes Toxins Enzymes Sensitizing drugs
Anti-Angiogenic Agents Cisplatin Anti-Angiogenic Agents
Anti-Metabolites Anti-Angiogenic Agents Mitotic Inhibitors
Anti-Angiogenic Agents Growth Factor Inhibitors Chemotherapeutic
Agent Paclitaxel Chemotherapeutic Agent Temozolomide
Chemotherapeutic Agent Topotecan Chemotherapeutic Agent
Fluorouracil Chemotherapeutic Agent Vincristine Chemotherapeutic
Agent Vinblastine Chemotherapeutic Agent Procarbazine
Chemotherapeutic Agent Decarbazine Chemotherapeutic Agent
Altretamine Chemotherapeutic Agent Methotrexate Chemotherapeutic
Agent Mercaptopurine Chemotherapeutic Agent Thioguanine
Chemotherapeutic Agent Fludarabine Phosphate Chemotherapeutic Agent
Cladribine Chemotherapeutic Agent Pentostatin Chemotherapeutic
Agent Cytarabine Chemotherapeutic Agent Azacitidine
Chemotherapeutic Agent Etoposide Chemotherapeutic Agent Teniposide
Chemotherapeutic Agent Irinotecan Chemotherapeutic Agent Docetaxel
Chemotherapeutic Agent Doxorubicin Chemotherapeutic Agent
Daunorubicin Chemotherapeutic Agent Dactinomycin Chemotherapeutic
Agent Idarubicin Chemotherapeutic Agent Plicamycin Chemotherapeutic
Agent Mitomycin Chemotherapeutic Agent Bleomycin Chemotherapeutic
Agent Tamoxifen Chemotherapeutic Agent Flutamide Chemotherapeutic
Agent Leuprolide Chemotherapeutic Agent Goserelin
Chemotherapeutic Agent Aminogluthimide Chemotherapeutic Agent
Anastrozole Chemotherapeutic Agent Amsacrine Chemotherapeutic Agent
Asparaginase Chemotherapeutic Agent Mitoxantrone Chemotherapeutic
Agent Mitotane Chemotherapeutic Agent Amifostine Apoptotic Agents
Cell Death or Cell Killing Agents Caspases Apoptosis Activators
Apoptosis Inhibitors XBP-1 Apoptosis Inhibitors Bcl-2 Apoptosis
Inhibitors Bcl-Xl Apoptosis Inhibitors Bcl-w Nonsteroidal
Anti-Inflammatory COX-2 Inhibitors Drugs (NSAID) Nonsteroidal
Anti-Inflammatory Ketorolac Drugs (NSAID) Nonsteroidal
Anti-Inflammatory Indomethacin Drugs (NSAID) Nonsteroidal
Anti-Inflammatory Etodolac Drugs (NSAID) Nonsteroidal
Anti-Inflammatory Tolemetin Drugs (NSAID) Nonsteroidal
Anti-Inflammatory Naproxen Drugs (NSAID) Nonsteroidal
Anti-Inflammatory Enolic Acid Derivatives Drugs (NSAID)
Nonsteroidal Anti-Inflammatory Anthranilic Acid Derivatives Drugs
(NSAID) Nonsteroidal Anti-Inflammatory Celecoxib Drugs (NSAID)
Nonsteroidal Anti-Inflammatory Sulfonanilides Drugs (NSAID)
Nonsteroidal Anti-Inflammatory Salicylates Drugs (NSAID)
Nonsteroidal Anti-Inflammatory Aceclofenac Drugs (NSAID)
Nonsteroidal Anti-Inflammatory Nabumetone Drugs (NSAID)
Nonsteroidal Anti-Inflammatory Sulindac Drugs (NSAID) Nonsteroidal
Anti-Inflammatory Diclofenac Drugs (NSAID) Nonsteroidal
Anti-Inflammatory Ibuprofen Drugs (NSAID) Steroids Dexamethasone
Steroids Budesonide Steroids Triamcinolone Steroids Triamcinolone
acetonide Steroids Cortisone Steroids Prednisone Steroids
Prednisolone Steroids Triamcinolone Hexacetonide Steroids
Methylprednisolone Pain Reliever Acetaminophen Opioids Local
Anesthetics Anti-Depressants Glutamate Receptor Antagonists
Adenosine Neuropeptides Uricase Elastase
[0130] Further examples of active agents include but are not
limited to: a peptide, an oligopeptide, a polypeptide, a
peptidomimetic, a polynucleotide, a polyribonucleotide, a DNA, a
cDNA, a ssDNA, a RNA, a dsRNA, a micro RNA, an RNAi, an
oligonucleotide, an antibody, a single chain variable fragment
(scFv), an antibody fragment, an aptamer, a cytokine, an
interferon, a hormone, an enzyme, a growth factor, a checkpoint
inhibitor, a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA4 inhibitor,
a CD antigen, aa chemokine, a neurotransmitter, an ion channel
inhibitor, a G-protein coupled receptor inhibitor, a G-protein
coupled receptor activator, a chemical agent, a radiosensitizer, a
radioprotectant, a radionuclide, a therapeutic small molecule, a
steroid, a corticosteroid, an anti-inflammatory agent, an immune
modulator, a complement fixing peptide or protein, a tumor necrosis
factor inhibitor, a tumor necrosis factor activator, a tumor
necrosis factor receptor family agonist, a tumor necrosis receptor
antagonist, a tumor necrosis factor (TNF) soluble receptor or
antibody, caspase protease activator or inhibitor, an NF-.kappa.B a
RIPK1 and/or RIPK3 inhibitor or activator (e.g., through Toll-like
receptors (TLRs) TLR-3 and/or TLR-4, or T-cell receptor (TCR) and
the like), a death-receptor ligand (e.g., Fas ligand) activator or
inhibitor, TNF receptor family (e.g., TNFR1, TNFR2, lymphotoxin
.beta. receptor/TNFRS3, OX40/TNFRSF4, CD40/TNFRSF5, Fas/TNFRSF6,
decoy receptor 3/TNFRSF6B, CD27/TNFRSF7, CD30/TNFRSF8,
4-1BB/TNFRSF9, DR4 (death receptor 4/TNFRS10A), DR5 (death receptor
5/TNFRSF10B), decoy receptor 1/TNFRSF10C, decoy receptor
2/TNFRSF10D, RANK (receptor activator of NF-kappa B/TNFRSF11A), OPG
(osteoprotegerin/TNFRSF11B), DR3 (death receptor 3/TNFRSF25), TWEAK
receptor/TNFRSF12A, TAC1/TNFRSF13B, BAFF-R (BAFF
receptor/TNFRSF13C), HVEM (herpes virus entry mediator/TNFRSF14),
nerve growth factor receptor/TNFRSF16, BCMA (B cell maturation
antigen/TNFRSF17), GITR (glucocorticoid-induced TNF
receptor/TNFRSF18), TAJ (toxicity and JNK inducer/TNFRSF19),
RELT/TNFRSF19L, DR6 (death receptor 6/TNFRSF21), TNFRSF22,
TNFRSF23, ectodysplasin A2 isoform receptor/TNFRS27, ectodysplasin
1, and anhidrotic receptor, a TNF receptor superfamily ligand
including--TNF alpha, lymphotoxin-.alpha., tumor necrosis factor
membrane form, tumor necrosis factor shed form, LIGHT, lymphotoxin
.beta..sub.2.alpha..sub.1 heterotrimer, OX-40 ligand, compound 1
[PMID: 24930776], CD40 ligand, Fas ligand, TL1A, CD70, CD30 ligand,
TRAF1, TRAF2, TRAF3, TRAIL, RANK ligand, APRIL, BAFF, B and T
lymphocyte attenuator, NGF, BDNF, neurotrophin-3, neurotrophin-4,
TL6, ectodysplasin A2, ectodysplasin A1-a TIMP-3 inhibitor, a BCL-2
family inhibitor, navitoclax (Aging Cell. 15(3): 428-435. (2016))
an IAP disruptor, a protease inhibitor, an amino sugar, a
chemotherapeutic (whether acting through an apoptotic or
non-apoptotic pathway) (Ricci et al. Oncologist 11(4):342-57
(2006)), a cytotoxic chemical, a toxin, a tyrosine kinase inhibitor
(e.g., imatinib mesylate), protons, bevacuzimab (antivascular
agent), erlotinib (EGFR inhibitor), an anti-infective agent, an
antibiotic, an anti-viral agent, an anti-fungal agent, an
aminoglycoside, a nonsteroidal anti-inflammatory drug (NSAID), a
statin, a nanoparticle, a liposome, a polymer, a biopolymer, a
polysaccharide, a proteoglycan, a glycosaminoglycan, polyethylene
glycol, a lipid, a dendrimer, a fatty acid, or an Fc domain or an
Fc region, or an active fragment or a modification thereof. Any
combination of the above active agents can be co-delivered with
peptides or peptide conjugates of this disclosure. Additionally, in
some embodiments, other co-therapies such as proton therapy or
ablative radiotherapy can be administered to a subject in need
thereof along with peptides or peptide conjugates of this
disclosure. In some embodiments, the peptide is covalently or
non-covalently linked to an active agent, e.g., directly or via a
linker. TNF blockers suppress the immune system by blocking the
activity of TNF, a substance in the body that can cause
inflammation and lead to immune-system diseases, such as Crohn's
disease, ulcerative colitis, rheumatoid arthritis, ankylosing
spondylitis, psoriatic arthritis and plaque psoriasis. The drugs in
this class include Remicade (infliximab), Enbrel (etanercept),
Humira (adalimumab), Cimzia (certolizumab pegol) and Simponi
(golimumab). The peptide disclosed herein can be used to home,
distribute to, target, directed to, is retained by, accumulate in,
migrate to, and/or bind to cartilage, and thus also be used for
localizing the attached or fused active agent. Furthermore,
cystine-dense chlorotoxin peptide can be internalized in cells
(Wiranowska, M., Cancer Cell Int, 11: 27 (2011)). Therefore,
cellular internalization, subcellular localization, and
intracellular trafficking after internalization of the peptide
itself, or an active agent peptide conjugate or fusion peptide can
be important factors in the efficacy of an active agent conjugate
or fusion. (Ducry, L., Antibody Drug Conjugates (2013); and Singh,
S. K., Pharm Res., 32(11): 3541-3571 (2015)). Exemplary linkers
suitable for use with the embodiments herein are discussed in
further detail below.
[0131] The peptides or peptide-active agent fusions of the present
disclosure can also be conjugated to other moieties that can serve
other roles, such as providing an affinity handle (e.g., biotin)
for retrieval of the peptides from tissues or fluids. For example,
peptides or peptide-active agent fusions of the present disclosure
can also be conjugated to biotin. In addition to extension of
half-life, biotin could also act as an affinity handle for
retrieval of peptides or peptide-active agent fusions from tissues
or other locations. In some embodiments, fluorescent biotin
conjugates that can act both as a detectable label and an affinity
handle can be used. Non limiting examples of commercially available
fluorescent biotin conjugates include Atto 425-Biotin, Atto
488-Biotin, Atto 520-Biotin, Atto-550 Biotin, Atto 565-Biotin, Atto
590-Biotin, Atto 610-Biotin, Atto 620-Biotin, Atto 655-Biotin, Atto
680-Biotin, Atto 700-Biotin, Atto 725-Biotin, Atto 740-Biotin,
fluorescein biotin, biotin-4-fluorescein, biotin-(5-fluorescein)
conjugate, and biotin-B-phycoerythrin, Alexa fluor 488 biocytin,
Alexa flour 546, Alexa Fluor 549, lucifer yellow cadaverine
biotin-X, Lucifer yellow biocytin, Oregon green 488 biocytin,
biotin-rhodamine and tetramethylrhodamine biocytin. In some other
examples, the conjugates could include chemiluminescent compounds,
colloidal metals, luminescent compounds, enzymes, radioisotopes,
and paramagnetic labels. In some embodiments, the peptide-active
agent fusions described herein can be attached to another molecule.
For example, the peptide sequence also can be attached to another
active agent (e.g., small molecule, peptide, polypeptide,
polynucleotide, antibody, aptamer, cytokine, growth factor,
neurotransmitter, an active fragment or modification of any of the
preceding, fluorophore, radioisotope, radionuclide chelator, acyl
adduct, chemical linker, or sugar, etc.). In some embodiments, the
peptide can be fused with, or covalently or non-covalently linked
to an active agent.
[0132] Additionally, more than one peptide sequence can be present
on or fused with a particular peptide. A peptide can be
incorporated into a biomolecule by various techniques, for example
by a chemical transformation, such as the formation of a covalent
bond, such as an amide bond, or by solid phase or solution phase
peptide synthesis, or by preparing a nucleic acid sequence encoding
the biomolecule, wherein the nucleic acid sequence includes a
subsequence that encodes the peptide. The subsequence can be in
addition to the sequence that encodes the biomolecule, or can
substitute for a subsequence of the sequence that encodes the
biomolecule.
Detectable Agent Conjugates
[0133] Described herein are agents that can be conjugated to the
peptides of the present invention for use in detection and tracing
either cartilage disorders or kidney disorders, or both. As
described herein, it is understood that certain active agents are
described in a non-limiting exemplary manner for use in
diagnostics, aiding surgery and treatment, prognosis and tracking
of progress or remission of cartilage and/or kidney disorders,
diseases or injury. One or more of such detectable agents can be
conjugated to a peptide of the present invention alone or in
combination with one or more active agents described herein.
Moreover some detectable agents (e.g., radionuclides,
radioisotopes, radiosensitizers and photosensitizers amongst
others) may also exert therapeutic activity as well. A peptide can
be conjugated to an agent used in imaging, research, therapeutics,
theranostics, pharmaceuticals, chemotherapy, chelation therapy,
targeted drug delivery, and radiotherapy. The agent can be a
detectable agent. In some embodiments, a peptide of the present
invention is conjugated to detectable agents, such as a metal, a
radioisotope, a dye, fluorophore, or another suitable material that
can be used in imaging. Non-limiting examples of radioisotopes
include alpha emitters, beta emitters, positron emitters, and gamma
emitters. In some embodiments, the metal or radioisotope is
selected from the group consisting of actinium, americium, bismuth,
cadmium, cesium, cobalt, europium, gadolinium, iridium, lead,
lutetium, manganese, palladium, polonium, radium, ruthenium,
samarium, strontium, technetium, thallium, and yttrium. In some
embodiments, the metal is actinium, bismuth, lead, radium,
strontium, samarium, or yttrium. In some embodiments, the
radioisotope is actinium-225 or lead-212. In some embodiments, the
fluorophore is a fluorescent agent emitting electromagnetic
radiation at a wavelength between 650 nm and 4000 nm, such
emissions being used to detect such agent. In some embodiments the
fluorophore is a fluorescent agent is selected from the group
consisting of non-limiting examples of fluorescent dyes that could
be used as a conjugating molecule (or as applied to each class of
molecules) in the present disclosure include DyLight-680,
DyLight-750, VivoTag-750, DyLight-800, IRDye-800, VivoTag-680, Cy5.
5, or indocyanine green (ICG class of dyes). In some embodiments,
near infrared dyes include cyanine dyes. Additional non-limiting
examples of fluorescent dyes for use as a conjugating molecule in
the present disclosure include acradine orange or yellow, Alexa
Fluors and any derivative thereof, 7-actinomycin D,
8-anilinonaphthalene-1-sulfonic acid, ATTO dye and any derivative
thereof, auramine-rhodamine stain and any derivative thereof,
bensantrhone, bimane, 9-10-bis(phenylethynyl)anthracene,
5,12-bis(phenylethynyl)naththacene, bisbenzimide, brainbow,
calcein, carbodyfluorescein and any derivative thereof,
1-chloro-9,10-bis(phenylethynyl)anthracene and any derivative
thereof, DAPI, DiOC6, DyLight Fluors and any derivative thereof,
epicocconone, ethidium bromide, FlAsH-EDT2, Fluo dye and any
derivative thereof, FluoProbe and any derivative thereof,
Fluorescein and any derivative thereof, Fura and any derivative
thereof, GelGreen and any derivative thereof, GelRed and any
derivative thereof, fluorescent proteins and any derivative
thereof, m isoform proteins and any derivative thereof such as for
example mCherry, hetamethine dye and any derivative thereof,
hoeschst stain, iminocoumarin, indian yellow, indo-1 and any
derivative thereof, laurdan, lucifer yellow and any derivative
thereof, luciferin and any derivative thereof, luciferase and any
derivative thereof, mercocyanine and any derivative thereof, nile
dyes and any derivative thereof, perylene, phloxine, phyco dye and
any derivative thereof, propium iodide, pyranine, rhodamine and any
derivative thereof, ribogreen, RoGFP, rubrene, stilbene and any
derivative thereof, sulforhodamine and any derivative thereof, SYBR
and any derivative thereof, synapto-pHluorin, tetraphenyl
butadiene, tetrasodium tris, Texas Red, Titan Yellow, TSQ,
umbelliferone, violanthrone, yellow fluorescent protein and YOYO-1.
Other Suitable fluorescent dyes include, but are not limited to,
fluorescein and fluorescein dyes (e.g., fluorescein isothiocyanine
or FITC, naphthofluorescein,
4',5'-dichloro-2',7'-dimethoxyfluorescein, 6-carboxyfluorescein or
FAM, etc.), carbocyanine, merocyanine, styryl dyes, oxonol dyes,
phycoerythrin, erythrosin, eosin, rhodamine dyes (e.g.,
carboxytetramethyl-rhodamine or TAMRA, carboxyrhodamine 6G,
carboxy-X-rhodamine (ROX), lissamine rhodamine B, rhodamine 6G,
rhodamine Green, rhodamine Red, tetramethylrhodamine (TMR), etc.),
coumarin and coumarin dyes (e.g., methoxycoumarin,
dialkylaminocoumarin, hydroxycoumarin, aminomethylcoumarin (AMCA),
etc.), Oregon Green Dyes (e.g., Oregon Green 488, Oregon Green 500,
Oregon Green 514, etc.), Texas Red, Texas Red-X, SPECTRUM RED,
SPECTRUM GREEN, cyanine dyes (e.g., CY-3, Cy-5, CY-3. 5, CY-5. 5,
etc.), ALEXA FLUOR dyes (e.g., ALEXA FLUOR 350, ALEXA FLUOR 488,
ALEXA FLUOR 532, ALEXA FLUOR 546, ALEXA FLUOR 568, ALEXA FLUOR 594,
ALEXA FLUOR 633, ALEXA FLUOR 660, ALEXA FLUOR 680, etc.), BODIPY
dyes (e.g., BODIPY FL, BODIPY R6G, BODIPY TMR, BODIPY TR, BODIPY
530/550, BODIPY 558/568, BODIPY 564/570, BODIPY 576/589, BODIPY
581/591, BODIPY 630/650, BODIPY 650/665, etc.), IRDyes (e.g.,
IRD40, IRD 700, IRD 800, etc.), indocyanine green dyes and the
like. For each of the above listed fluorescent dyes various
activated forms can be used for conjugation. Additional suitable
detectable agents are described in PCT/US14/56177. Non-limiting
examples of radioisotopes include alpha emitters, beta emitters,
positron emitters, and gamma emitters. In some embodiments, the
metal or radioisotope is selected from the group consisting of
actinium, americium, bismuth, cadmium, cesium, cobalt, europium,
gadolinium, iridium, lead, lutetium, manganese, palladium,
polonium, radium, ruthenium, samarium, strontium, technetium,
thallium, and yttrium. In some embodiments, the metal is actinium,
bismuth, lead, radium, strontium, samarium, or yttrium. In some
embodiments, the radioisotope is actinium-225 or lead-212.
[0134] Other embodiments of the present disclosure provide peptides
conjugated to a radiosensitizer or photosensitizer. Examples of
radiosensitizers include but are not limited to: ABT-263, ABT-199,
WEHI-539, paclitaxel, carboplatin, cisplatin, oxaliplatin,
gemcitabine, etanidazole, misonidazole, tirapazamine, and nucleic
acid base derivatives (e.g., halogenated purines or pyrimidines,
such as 5-fluorodeoxyuridine). Examples of photosensitizers include
but are not limited to: fluorescent molecules or beads that
generate heat when illuminated, porphyrins and porphyrin
derivatives (e.g., chlorins, bacteriochlorins, isobacteriochlorins,
phthalocyanines, and naphthalocyanines), metalloporphyrins,
metallophthalocyanines, angelicins, chalcogenapyrrillium dyes,
chlorophylls, coumarins, flavins and related compounds such as
alloxazine and riboflavin, fullerenes, pheophorbides,
pyropheophorbides, cyanines (e.g., merocyanine 540), pheophytins,
sapphyrins, texaphyrins, purpurins, porphycenes, phenothiaziniums,
methylene blue derivatives, naphthalimides, nile blue derivatives,
quinones, perylenequinones (e.g., hypericins, hypocrellins, and
cercosporins), psoralens, quinones, retinoids, rhodamines,
thiophenes, verdins, xanthene dyes (e.g., eosins, erythrosins, rose
bengals), dimeric and oligomeric forms of porphyrins, and prodrugs
such as 5-aminolevulinic acid. Advantageously, this approach allows
for highly specific targeting of diseased cells (e.g., cancer
cells) using both a therapeutic agent (e.g., drug) and
electromagnetic energy (e.g., radiation or light) concurrently. In
some embodiments, the peptide is covalently or non-covalently
linked to the agent, e.g., directly or via a linker. Exemplary
linkers suitable for use with the embodiments herein are discussed
in further detail below.
Linkers
[0135] Peptides according to the present disclosure that home,
target, migrate to, are retained by, accumulate in, and/or bind to,
or are directed to the cartilage can be attached to another moiety
(e.g., an active agent), such as a small molecule, a second
peptide, a protein, an antibody, an antibody fragment, an aptamer,
polypeptide, polynucleotide, a fluorophore, a radioisotope, a
radionuclide chelator, a polymer, a biopolymer, a fatty acid, an
acyl adduct, a chemical linker, or sugar or other active agent
described herein through a linker, or directly in the absence of a
linker.
[0136] A peptide can be directly attached to another molecule by a
covalent attachment. For example, the peptide is attached to a
terminus of the amino acid sequence of a larger polypeptide or
peptide molecule, or is attached to a side chain, such as the side
chain of a lysine, serine, threonine, cysteine, tyrosine, aspartic
acid, a non-natural amino acid residue, or glutamic acid residue.
The attachment can be via an amide bond, an ester bond, an ether
bond, a carbamate bond, a carbon-nitrogen bond, a triazole, a
macrocycle, an oxime bond, a hydrazone bond, a carbon-carbon single
double or triple bond, a disulfide bond, or a thioether bond. In
some embodiments, similar regions of the disclosed peptide(s)
itself (such as a terminus of the amino acid sequence, an amino
acid side chain, such as the side chain of a lysine, serine,
threonine, cysteine, tyrosine, aspartic acid, a non-natural amino
acid residue, or glutamic acid residue, via an amide bond, an ester
bond, an ether bond, a carbamate bond, a carbon-nitrogen bond, a
triazole, a macrocycle, an oxime bond, a hydrazone bond, a
carbon-carbon single double or triple bond, a disulfide bond, or a
thioether bond, or linker as described herein) can be used to link
other molecules.
[0137] Attachment via a linker can involve incorporation of a
linker moiety between the other molecule and the peptide. The
peptide and the other molecule can both be covalently attached to
the linker. The linker can be cleavable, labile, non-cleavable,
stable self-immolating, hydrophilic, or hydrophobic. As used
herein, the term "non-cleavable" (such as used in association with
an amide, cyclic, or carbamate linker or as otherwise as described
herein) is often used by a skilled artisan to distinguish a
relatively stable structure from one that is more labile or
"cleavable" (e.g., as used in association with cleavable linkers
that may be dissociated or cleaved structurally by enzymes,
proteases, self-immolation, pH, reduction, hydrolysis, certain
physiologic conditions, or as otherwise described herein). It is
understood that "non-cleavable" linkers offer stability against
cleavage or other dissociation as compared to "cleavable" linkers,
and the term is not intended to be considered an absolute
non-cleavable or non-dissociative structure under any conditions.
Consequently, as used herein, a "non-cleavable" linker is also
referred to as a "stable" linker. The linker can have at least two
functional groups with one bonded to the peptide, the other bonded
to the other molecule, and a linking portion between the two
functional groups.
[0138] Non-limiting examples of the functional groups for
attachment can include functional groups capable of forming an
amide bond, an ester bond, an ether bond, a carbonate bond, a
carbamate bond, or a thioether bond. Non-limiting examples of
functional groups capable of forming such bonds can include amino
groups; carboxyl groups; hydroxyl groups; aldehyde groups; azide
groups; alkyne and alkene groups; ketones; hydrazides; acid halides
such as acid fluorides, chlorides, bromides, and iodides; acid
anhydrides, including symmetrical, mixed, and cyclic anhydrides;
carbonates; carbonyl functionalities bonded to leaving groups such
as cyano, succinimidyl, and N-hydroxysuccinimidyl; hydroxyl groups;
sulfhydryl groups; and molecules possessing, for example, alkyl,
alkenyl, alkynyl, allylic, or benzylic leaving groups, such as
halides, mesylates, tosylates, triflates, epoxides, phosphate
esters, sulfate esters, and besylates.
[0139] Non-limiting examples of the linking portion can include
alkylene, alkenylene, alkynylene, polyether, such as polyethylene
glycol (PEG), hydroxy carboxylic acids, polyester, polyamide,
polyamino acids, polypeptides, cleavable peptides,
valine-citrulline, aminobenzylcarbamates, D-amino acids, and
polyamine, any of which being unsubstituted or substituted with any
number of substituents, such as halogens, hydroxyl groups,
sulfhydryl groups, amino groups, nitro groups, nitroso groups,
cyano groups, azido groups, sulfoxide groups, sulfone groups,
sulfonamide groups, carboxyl groups, carboxaldehyde groups, imine
groups, alkyl groups, halo-alkyl groups, alkenyl groups,
halo-alkenyl groups, alkynyl groups, halo-alkynyl groups, alkoxy
groups, aryl groups, aryloxy groups, aralkyl groups, arylalkoxy
groups, heterocyclyl groups, acyl groups, acyloxy groups, carbamate
groups, amide groups, urethane groups, epoxides, and ester
groups.
[0140] A peptide and drug conjugated via a linker is described with
the formula Peptide-A-B-C-Drug, wherein the linker is A-B-C. A can
be a stable amide link, is an amine on the peptide and the linker
and can be achieved via a tetrafluorophenyl (TFP) ester or an NHS
ester. B can be (--CH2-).sub.x- or a short PEG (--CH2CH2O--).sub.x
(x is 1-10), and C can be the ester bond to the hydroxyl or
carboxylic acid on the drug. In some embodiments, C can refer to
the "cleavable" or "stable" part of the linker. In other
embodiments, A can also be the "cleavable" part. In some
embodiments, A can be amide, carbamate, thioether via maleimide or
bromoacetamide, triazole, oxime, or oxacarboline. The cleaved
active agent or drug can retain the chemical structure of the
active agent before cleavage, or can be modified as a result of
cleavage. Moreover, depending on the desired therapeutic properties
of the peptide-drug conjugate, such active agent can be active
while linked to the peptide, remain active after cleavage or become
inactivated, be inactive while linked to the peptide, or it can be
activated upon cleavage.
[0141] In some embodiments, peptide conjugates have stable linkers.
A peptide of the disclosure can be expressed recombinantly or
chemically synthesized. The peptide can be conjugated to a
detectable agent or an active agent via a stable linker, such as an
amide linkage or a carbamate linkage. The peptide can be conjugated
to a detectable agent or an active agent via a stable linker, such
as an amide bond using standard
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) or
dicylcohexylcarbodiimide (DCC) based chemistry or thionyl chloride
or phosphorous chloride-based bioconjugation chemistries. A stable
linker may or may not be cleaved in buffer over extended periods of
time (e.g., hours, days, or weeks). A stable linker may or may not
be cleaved in body fluids such as plasma or synovial fluid over
extended periods of time (e.g., hours, days, or weeks). A stable
linker, may or may not be cleaved after exposure to enzymes,
reactive oxygen species, other chemicals or enzymes that can be
present in cells (e.g., macrophages), cellular compartments (e.g.,
endosomes and lysosomes), inflamed areas of the body (e.g.,
inflamed joints), tissues or body compartments. A stable linker may
be cleaved by unknown mechanisms. A stable linker may or may not be
cleaved in vivo but remains an active agent after peptide
conjugation.
[0142] A peptide and drug conjugated via a linker can be described
with the formula Peptide-A-B-C-Drug, wherein the linker is A-B-C. A
can be a stable amide link such as that formed by reacting an amine
on the peptide with a linker containing a tetrafluorophenyl (TFP)
ester or an NHS ester. A can also be a stable carbamate linker such
as that formed by reacting an amine on the peptide with an
imidazole carbamate active intermediate formed by reaction of CDI
with a hydroxyl on the linker. A can also be a stable secondary
amine linkage such as that formed by reductive alkylation of the
amine on the peptide with an aldehyde or ketone group on the
linker. A can also be a stable thioether linker formed using a
maleimide or bromoacetamide in the linker with a thiol in the
peptide, a triazole linker, a stable oxime linker, or a
oxacarboline linker. B can be (--CH2-).sub.x- or a short PEG
(--CH2CH2O--).sub.x (x is 0-20) or other spacers or no spacer. C
can be an amide bond formed with an amine or a carboxylic acid on
the drug, a thioether formed between a maleimide on the linker and
a sulfhydroyl on the drug, a secondary or tertiary amine, a
carbamate, or other stable bonds. Any linker chemistry described in
"Current ADC Linker Chemistry," Jain et al., Pharm Res, 2015 DOI
10. 1007/s11095-015-1657-7 can be used.
[0143] The resulting peptide conjugates can be administered to a
human or animal subcutaneously, intravenously, orally, or injected
directly into a joint to treat disease. The peptide is not
specifically cleaved from the detectable agent or active agent via
a targeted mechanism. The peptide can be degraded by mechanisms
such as catabolism, releasing a drug that is modified or not
modified form its native form (Antibody-Drug Conjugates: Design,
Formulation, and Physicochemical Stability, Singh, Luisi, and Pak.
Pharm Res (2015) 32:3541-3571). The peptide drug conjugate exerts
its pharmacological activity while still intact, or while partially
or fully degraded, metabolized, or catabolized.
[0144] In some embodiments, peptide conjugates can have cleavable
linkers. In some embodiments, a peptide and drug can be conjugated
via a linker and can be described with the formula
Peptide-A-B-C-Drug, wherein the linker is A-B-C. In some
embodiments, A can be a stable amide link such as that formed by
reacting an amine on the peptide with a linker containing a
tetrafluorophenyl (TFP) ester or an NHS ester. In certain
embodiments, A can also be a stable carbamate linker that is formed
by an amine reaction on the peptide with an imidazole carbamate
active intermediate formed by reaction of CDI with a hydroxyl on
the linker. In other embodiments, A can also be a stable secondary
amine linkage such as that formed by reductive alkylation of the
amine on the peptide with an aldehyde or ketone group on the
linker. In some embodiments, A can also be a stable thioether
linker formed using a maleimide or bromoacetamide in the linker
with a thiol in the peptide, a triazole linker, a stable oxime
linker, or an oxacarboline linker. B can be (--CH2-).sub.x- or a
short PEG (--CH2CH2O--).sub.x (x is 0-20) or other spacers or no
spacer. C can be an ester bond to the hydroxyl or carboxylic acid
on the drug, or a carbonate, hydrazone, or acylhydrazone, designed
for hydrolytic cleavage. The hydrolytic rate of cleavage can be
varied by varying the local environment around the bond, including
carbon length (--CH2-).sub.x, steric hindrance (including adjacent
side groups such as methyl, ethyl, cyclic), hydrophilicity or
hydrophobicity. In some embodiments, peptide conjugates can have a
linear or cyclic ester linkage, which can include or do not include
side chains such as methyl or ethyl groups. A linear ester linkage
can be more susceptible to cleavage (such as by hydrolysis, an
enzyme such as esterase, or other chemical reaction) than a cyclic
ester due to steric hindrance or hydrophobicity/hydrophilicity
effects. Likewise, side chains such as methyl or ethyl groups on
the linear ester linkage can optionally make the linkage less
susceptible to cleavage than without the side chains. In some
embodiments, hydrolysis rate can be affected by local pH, such as
lower pH in certain compartments of the body or of the cell such as
endosomes and lysosomes or diseased tissues. In some embodiments, C
can also be a pH sensitive group such as a hydrazone or oxime
linkage. In other embodiments, C can be a disulfide bond designed
to be released by reduction, such as by glutathione. In other
embodiments, (or A-B-C) can be a peptidic linkage design for
cleavable by enzymes. Optionally, a self-immolating group such as
pABC can be included to cause release of a free unmodified drug
upon cleavage (Antibody-Drug Conjugates: Design, Formulation, and
Physicochemical Stability, Singh, Luisi, and Pak. Pharm Res (2015)
32:3541-3571). The linker can be cleaved by enzymes such as
esterases, matrix metalloproteinases, cathepsins such as cathepsin
B, glucuronidases, a protease, or thrombin. Alternatively, the bond
designed for cleavage can be at A, rather than C, and C can be a
stable bond or a cleavable bond. An alternative design can be to
have stable linkers (such as amide or carbamate) at A and C and
have a cleavable linker in B, such as a disulfide bond. The rate of
reduction can be modulated by local effects such as steric
hindrance from methyl or ethyl groups or modulating
hydrophobicity/hydrophilicity. In some embodiments, peptide
conjugates can have an ester carbonyl linkage, a long hydrocarbon
linker, or carbamate linker, each of which can include hydrophilic
groups, such as alcohols, acids, or ethers, or include a
hydrocarbon side chain or other moiety that tunes the rate of
cleavage. For example, the rate of hydrolysis can be faster with
hydrophilic groups, such as alcohols, acids, or ethers, near an
ester carbonyl. In another example, hydrophobic groups present as
side chains or as a longer hydrocarbon linker can slow the cleavage
rate of the ester. Likewise, cleavage of a carbamate group can also
be tuned by hindrance, hydrophobicity, and the like. In another
example, using a less labile linking group, such as a carbamate
rather than an ester, can slow the cleavage rate of the linker.
[0145] Non-limiting examples of linkers include:
##STR00001##
wherein each n is independently 0 to about 1,000; 1 to about 1,000;
0 to about 500; 1 to about 500; 0 to about 250; 1 to about 250; 0
to about 200; 1 to about 200; 0 to about 150; 1 to about 150; 0 to
about 100; 1 to about 100; 0 to about 50; 1 to about 50; 0 to about
40; 1 to about 40; 0 to about 30; 1 to about 30; 0 to about 25; 1
to about 25; 0 to about 20; 1 to about 20; 0 to about 15; 1 to
about 15; 0 to about 10; 1 to about 10; 0 to about 5; or 1 to about
5. In some embodiments, each n is independently 0, about 1, about
2, about 3, about 4, about 5, about 6, about 7, about 8, about 9,
about 10, about 11, about 12, about 13, about 14, about 15, about
16, about 17, about 18, about 19, about 20, about 21, about 22,
about 23, about 24, about 25, about 26, about 27, about 28, about
29, about 30, about 31, about 32, about 33, about 34, about 35,
about 36, about 37, about 38, about 39, about 40, about 41, about
42, about 43, about 44, about 45, about 46, about 47, about 48,
about 49, or about 50. In some embodiments, m is 1 to about 1,000;
1 to about 500; 1 to about 250; 1 to about 200; 1 to about 150; 1
to about 100; 1 to about 50; 1 to about 40; 1 to about 30; 1 to
about 25; 1 to about 20; 1 to about 15; 1 to about 10; or 1 to
about 5. In some embodiments, m is 0, about 1, about 2, about 3,
about 4, about 5, about 6, about 7, about 8, about 9, about 10,
about 11, about 12, about 13, about 14, about 15, about 16, about
17, about 18, about 19, about 20, about 21, about 22, about 23,
about 24, about 25, about 26, about 27, about 28, about 29, about
30, about 31, about 32, about 33, about 34, about 35, about 36,
about 37, about 38, about 39, about 40, about 41, about 42, about
43, about 44, about 45, about 46, about 47, about 48, about 49, or
about 50.
[0146] In some cases a linker can be a succinic linker, and a drug
can be attached to a peptide via an ester bond or an amide bond
with two methylene carbons in between. In other cases, a linker can
be any linker with both a hydroxyl group and a carboxylic acid,
such as hydroxy hexanoic acid or lactic acid.
[0147] The linker can be a cleavable or a stable linker. The use of
a cleavable linker permits release of the conjugated moiety (e.g.,
a therapeutic agent) from the peptide, e.g., after targeting to the
cartilage. In some cases the linker is enzyme cleavable, e.g., a
valine-citrulline linker. In some embodiments, the linker contains
a self-immolating portion. In other embodiments, the linker
includes one or more cleavage sites for a specific protease, such
as a cleavage site for matrix metalloproteases (MMPs), thrombin, or
cathepsin. Alternatively or in combination, the linker is cleavable
by other mechanisms, such as via pH, reduction, or hydrolysis. A
hydrolytically labile linker, (amongst other cleavable linkers
described herein) can be advantageous in terms of releasing active
agents from the peptide. For example, an active agent in a
conjugate form with the peptide may not be active, but upon release
from the conjugate after targeting to the cartilage, the active
agent is active.
[0148] The rate of hydrolysis of the linker can be tuned. For
example, the rate of hydrolysis of linkers with unhindered esters
is faster compared to the hydrolysis of linkers with bulky groups
next an ester carbonyl. A bulky group can be a methyl group, an
ethyl group, a phenyl group, a ring, or an isopropyl group, or any
group that provides steric bulk. In some cases, the steric bulk can
be provided by the drug itself, such as by ketorolac when
conjugated via its carboxylic acid. The rate of hydrolysis of the
linker can be tuned according to the residency time of the
conjugate in the cartilage. For example, when a peptide is cleared
from the cartilage relatively quickly, the linker can be tuned to
rapidly hydrolyze. In contrast, for example, when a peptide has a
longer residence time in the cartilage, a slower hydrolysis rate
can allow for extended delivery of an active agent. This can be
important when the peptide is used to deliver a drug to the
cartilage. "Programmed hydrolysis in designing paclitaxel prodrug
for nanocarrier assembly" Sci Rep 2015, 5, 12023 Fu et al.,
provides an example of modified hydrolysis rates.
[0149] Peptide Stability
[0150] A peptide of the present disclosure can be stable in various
biological conditions, as well as during manufacturing, handling,
storage, and other conditions in either a liquid or a dried state.
Additionally, a peptide of the present disclosure can be resistant
to enzymatic cleavage needed for peptide processing by the immune
system. For example, any peptide of SEQ ID NO: 1-SEQ ID NO: 564 can
exhibit resistance to reducing agents, proteases, oxidative
conditions, or acidic conditions.
[0151] In some cases, biologic molecules (such as peptides and
proteins) can provide therapeutic functions, but such therapeutic
functions are decreased or impeded by instability caused by the in
vivo environment. (Moroz et al., Adv Drug Deliv Rev 101:108-21
(2016), Mitragotri et al., Nat Rev Drug Discov 13(9):655-72 (2014),
Bruno et al., Ther Deliv (11):1443-67 (2013), Sinha et al., Crit
Rev Ther Drug Carrier Syst. 24(1):63-92 (2007), Hamman et al.,
BioDrugs 19(3):165-77 (2005)). Peptide degradation can be a result
of a number of processes involving hydrolytic pathways, peptide
oxidation such as oxidation of methionine (Met) residues,
deamidation of asparagine (Asn) and glutamine (Gln) residues, and
isomerization and hydrolysis of an adjacent asparagine (Asp)
residue. (Manning et al., Pharmaceutical Research, Vol. 27 No. 4
(2010)). The amino acid immediately following the Asn or Gln
residue can also affect the rate of deamidation, whereas: Asn-Gly,
Asn-Ser, Asn-His, and Gln-Gly can be more likely to undergo
deamidation. Additionally, the peptide bond adjacent to amino acids
such as Asp can undergo hydrolysis with amino acid pairings such as
Asp-Gly, Asp-Ser, Asp-Tyr, and Asp-Pro, which can be more likely to
undergo hydrolysis. Oxidation of amino acid residues such as Met
can form a sulfoxide species. The specific degradation reactions
rates can vary for any given peptide or protein sequence.
[0152] Furthermore, the microenvironment within the molecular
structure of the peptide, solvent accessibility, and conformational
stability of each residue can impact the likelihood of peptide
degradation. Therefore, by modifying a peptide sequence to reduce
occurrence of such degradation events, a the modified peptide or
peptide-conjugate can have increased beneficial properties over
unmodified peptides or peptide-drug conjugates, such as improved
therapeutic efficacy, an increased safety profile, and can be less
expensive to manufacture and develop. Key formulaic considerations
that can prevent peptide decay can include the use of excipients,
formulation at a desired pH, and storage under specific conditions
(e.g., temperature, oxygen, light exposure, solid or liquid state,
and container excipient materials). To circumvent degradation,
peptide residues can be substituted with amino acids that increase
stability, which can result in more efficacious and durable
therapeutic peptides.
[0153] With respect to in vivo stability, the GI tract can contain
a region of low pH (e.g., pH .about.1), a reducing environment, or
a protease-rich environment that can degrade peptides and proteins.
Proteolytic activity in other areas of the body, such as the mouth,
eye, lung, intranasal cavity, joint, skin, vaginal tract, mucous
membranes, and serum, can also be an obstacle to the delivery of
functionally active peptides and polypeptides. Additionally, the
half-life of peptides in serum can be very short, in part due to
proteases, such that the peptide can be degraded too quickly to
have a lasting therapeutic effect when administering a therapeutic
and safe dosing regimen. Likewise, proteolytic activity in cellular
compartments, such as lysosomes, and reduction activity in
lysosomes and the cytosol can degrade peptides and proteins such
that they may be unable to provide a therapeutic function on
intracellular targets. Therefore, peptides that are resistant to
reducing agents, proteases, and low pH may be able to provide
enhanced therapeutic effects or enhance the therapeutic efficacy of
co-formulated or conjugated active agents in vivo.
[0154] Additionally, oral delivery of drugs can be desirable in
order to target certain areas of the body (e.g., disease in the GI
tract such as colon cancer, irritable bowel disorder, infections,
metabolic disorders, and constipation) despite the obstacles to the
delivery of functionally active peptides and polypeptides presented
by this method of administration. For example, oral delivery of
drugs can increase compliance by providing a dosage form that is
more convenient for patients to take as compared to parenteral
delivery. Oral delivery can be useful in treatment regimens that
have a large therapeutic window. Therefore, peptides that are
resistant to reducing agents, proteases, and low pH can allow for
oral delivery of peptides without nullifying their therapeutic
function.
[0155] Peptide Resistance to Reducing Agents. Peptides of this
disclosure can contain one or more cysteines, which can participate
in disulfide bridges that can be integral to preserving the folded
state of the peptide. Exposure of peptides to biological
environments with reducing agents can result in unfolding of the
peptide and loss of functionality and bioactivity. For example,
glutathione (GSH) is a reducing agent that can be present in many
areas of the body and in cells, and can reduce disulfide bonds. As
another example, a peptide can become reduced upon cellular
internalization during trafficking of a peptide across the
gastrointestinal epithelium after oral administration a peptide can
become reduced upon exposure to various parts of the GI tract. The
GI tract can be a reducing environment, which can inhibit the
ability of therapeutic molecules with disulfide bonds to have
optimal therapeutic efficacy, due to reduction of the disulfide
bonds. A peptide can also be reduced upon entry into a cell, such
as after internalization by endosomes or lysosomes or into the
cytosol, or other cellular compartments. Reduction of the disulfide
bonds and unfolding of the peptide can lead to loss of
functionality or affect key pharmacokinetic parameters such as
bioavailability, peak plasma concentration, bioactivity, and
half-life. Reduction of the disulfide bonds can also lead to
increased susceptibility of the peptide to subsequent degradation
by proteases, resulting in rapid loss of intact peptide after
administration. In some embodiments, a peptide that is resistant to
reduction can remain intact and can impart a functional activity
for a longer period of time in various compartments of the body and
in cells, as compared to a peptide that is more readily
reduced.
[0156] In certain embodiments, the peptides of this disclosure can
be analyzed for the characteristic of resistance to reducing agents
to identify stable peptides. In some embodiments, the peptides of
this disclosure can remain intact after being exposed to different
molarities of reducing agents such as 0.00001M-0.0001M,
0.0001M-0.001M, 0.001M-0.01M, 0.01 M-0.05 M, 0.05 M-0.1 M, for
greater 15 minutes or more. In some embodiments, the reducing agent
used to determine peptide stability can be dithiothreitol (DTT),
Tris (2-carboxyethyl) phosphine HCl (TCEP), 2-Mercaptoethanol,
(reduced) glutathione (GSH), or any combination thereof. In some
embodiments, at least 5%-10%, at least 10%-20%, at least 20%-30%,
at least 30%-40%, at least 40%-50%, at least 50%-60%, at least
60%-70%, at least 70%-80%, at least 80%-90%, or at least 90%-100%
of the peptide remains intact after exposure to a reducing
agent.
[0157] Peptide Resistance to Proteases. The stability of peptides
of this disclosure can be determined by resistance to degradation
by proteases. Proteases, also referred to as peptidases or
proteinases, can be enzymes that can degrade peptides and proteins
by breaking bonds between adjacent amino acids. Families of
proteases with specificity for targeting specific amino acids can
include serine proteases, cysteine proteases, threonine proteases,
aspartic proteases, glutamic proteases, esterases, serum proteases,
and asparagine proteases. Additionally, metalloproteases, matrix
metalloproteases, elastase, carboxypeptidases, Cytochrome P450
enzymes, and cathepsins can also digest peptides and proteins.
Proteases can be present at high concentration in blood, in mucous
membranes, lungs, skin, the GI tract, the mouth, nose, eye, and in
compartments of the cell. Misregulation of proteases can also be
present in various diseases such as rheumatoid arthritis and other
immune disorders. Degradation by proteases can reduce
bioavailability, biodistribution, half-life, and bioactivity of
therapeutic molecules such that they are unable to perform their
therapeutic function. In some embodiments, peptides that are
resistant to proteases can better provide therapeutic activity at
reasonably tolerated concentrations in vivo.
[0158] In some embodiments, peptides of this disclosure can resist
degradation by any class of protease. In certain embodiments,
peptides of this disclosure resist degradation by pepsin (which can
be found in the stomach), trypsin (which can be found in the
duodenum), serum proteases, or any combination thereof. In certain
embodiments, peptides of this disclosure can resist degradation by
lung proteases (e.g., serine, cysteinyl, and aspartyl proteases,
metalloproteases, neutrophil elastase, alpha-1 antitrypsin,
secretory leucoprotease inhibitor, elafin), or any combination
thereof. In some embodiments, the proteases used to determine
peptide stability can be pepsin, trypsin, chymotrypsin, or any
combination thereof. In some embodiments, at least 5%-10%, at least
10%-20%, at least 20%-30%, at least 30%-40%, at least 40%-50%, at
least 50%-60%, at least 60%-70%, at least 70%-80%, at least
80%-90%, or at least 90%-100% of the peptide remains intact after
exposure to a protease. Peptides of SEQ ID NO: 199, SEQ ID NO: 27,
and SEQ ID NO: 108 can have particular structural qualities, which
make them more resistant to protease degradation. For example,
peptide of SEQ ID NO: 27 and SEQ ID NO: 109 exhibit the "hitchin"
topology as described previously, which can be associated with
resistance to protease and chemical degradation.
[0159] Peptide Stability in Acidic Conditions. Peptides of this
disclosure can be administered in biological environments that are
acidic. For example, after oral administration, peptides can
experience acidic environmental conditions in the gastric fluids of
the stomach and gastrointestinal (GI) tract. The pH of the stomach
can range from .about.1-4 and the pH of the GI tract ranges from
acidic to normal physiological pH descending from the upper GI
tract to the colon. In addition, the vagina, late endosomes, and
lysosomes can also have acidic pH values, such as less than pH 7.
These acidic conditions can lead to denaturation of peptides and
proteins into unfolded states. Unfolding of peptides and proteins
can lead to increased susceptibility to subsequent digestion by
other enzymes as well as loss of biological activity of the
peptide.
[0160] In certain embodiments, the peptides of this disclosure can
resist denaturation and degradation in acidic conditions and in
buffers, which simulate acidic conditions. In certain embodiments,
peptides of this disclosure can resist denaturation or degradation
in buffer with a pH less than 1, a pH less than 2, a pH less than
3, a pH less than 4, a pH less than 5, a pH less than 6, a pH less
than 7, or a pH less than 8. In some embodiments, peptides of this
disclosure remain intact at a pH of 1-3. In certain embodiments, at
least 5%-10%, at least 10%-20%, at least 20%-30%, at least 30%-40%,
at least 40%-50%, at least 50%-60%, at least 60%-70%, at least
70%-80%, at least 80%-90%, or at least 90%-100% of the peptide
remains intact after exposure to a buffer with a pH less than 1, a
pH less than 2, a pH less than 3, a pH less than 4, a pH less than
5, a pH less than 6, a pH less than 7, or a pH less than 8. In
other embodiments, at least 5%-10%, at least 10%-20%, at least
20%-30%, at least 30%-40%, at least 40%-50%, at least 50%-60%, at
least 60%-70%, at least 70%-80%, at least 80%-90%, or at least
90%-100% of the peptide remains intact after exposure to a buffer
with a pH of 1-3. In other embodiments, the peptides of this
disclosure can be resistant to denaturation or degradation in
simulated gastric fluid (pH 1-2). In some embodiments, at least
5-10%, at least 10%-20%, at least 20%-30%, at least 30%-40%, at
least 40%-50%, at least 50%-60%, at least 60%-70%, at least
70%-80%, at least 80%-90%, or at least 90-100% of the peptide
remains intact after exposure to simulated gastric fluid. In some
embodiments, low pH solutions such as simulated gastric fluid or
citrate buffers can be used to determine peptide stability.
[0161] Peptide Stability at High Temperatures. Peptides of this
disclosure can be administered in biological environments with high
temperatures. For example, after oral administration, peptides can
experience high temperatures in the body. Body temperature can
range from 36.degree. C. to 40.degree. C. High temperatures can
lead to denaturation of peptides and proteins into unfolded states.
Unfolding of peptides and proteins can lead to increased
susceptibility to subsequent digestion by other enzymes as well as
loss of biological activity of the peptide. In some embodiments, a
peptide of this disclosure can remain intact at temperatures from
25.degree. C. to 100.degree. C. High temperatures can lead to
faster degradation of peptides. Stability at a higher temperature
can allow for storage of the peptide in tropical environments or
areas where access to refrigeration is limited. In certain
embodiments, 5%-100% of the peptide can remain intact after
exposure to 25.degree. C. for 6 months to 5 years. 5%-100% of a
peptide can remain intact after exposure to 70.degree. C. for 15
minutes to 1 hour. 5%-100% of a peptide can remain intact after
exposure to 100.degree. C. for 15 minutes to 1 hour. In other
embodiments, at least 5%-10%, at least 10%-20%, at least 20%-30%,
at least 30%-40%, at least 40%-50%, at least 50%-60%, at least
60%-70%, at least 70%-80%, at least 80%-90%, or at least 90%-100%
of the peptide remains intact after exposure to 25.degree. C. for 6
months to 5 years. In other embodiments, at least 5%-10%, at least
10%-20%, at least 20%-30%, at least 30%-40%, at least 40%-50%, at
least 50%-60%, at least 60%-70%, at least 70%-80%, at least
80%-90%, or at least 90%-100% of the peptide remains intact after
exposure to 70.degree. C. for 15 minutes to 1 hour. In other
embodiments, at least 5%-10%, at least 10%-20%, at least 20%-30%,
at least 30%-40%, at least 40%-50%, at least 50%-60%, at least
60%-70%, at least 70%-80%, at least 80%-90%, or at least 90%-100%
of the peptide remains intact after exposure to 100.degree. C. for
15 minutes to 1 hour.
[0162] In some embodiments, the peptide of the peptide active agent
conjugate comprises a sequence that has at least 75%, at least 80%,
at least 85%, at least 90%, at least 95%, at least 97%, at least
99% or 100% sequence identity with any one of SEQ ID NO: 24-SEQ ID
NO: 274 or a fragment thereof. In some embodiments, the peptide of
the peptide active agent conjugate comprises a sequence that has at
least 75%, at least 80%, at least 85%, at least 90%, at least 95%,
at least 97%, at least 99% or 100% sequence identity with any one
of SEQ ID NO: 260-SEQ ID NO: 274 or a fragment thereof. In some
embodiments, the peptide of the peptide active agent conjugate
comprises a sequence that has at least 75%, at least 80%, at least
85%, at least 90%, at least 95%, at least 97%, at least 99% or 100%
sequence identity with any one of SEQ ID NO: 314-SEQ ID NO: 564 or
a fragment thereof. In some embodiments, the peptide of the peptide
active agent conjugate comprises a sequence that has at least 75%,
at least 80%, at least 85%, at least 90%, at least 95%, at least
97%, at least 99% or 100% sequence identity with any one of SEQ ID
NO: 550-SEQ ID NO: 564 or a fragment thereof. In some embodiments,
the peptide of the peptide active agent conjugate comprises a
sequence of any one of SEQ ID NO: 24-SEQ ID NO: 274 or a fragment
thereof. In some embodiments, the peptide of the peptide active
agent conjugate comprises a sequence of any one of SEQ ID NO:
260-SEQ ID NO: 274 or a fragment thereof. In some embodiments, the
peptide of the peptide active agent conjugate comprises a sequence
of any one of SEQ ID NO: 314-SEQ ID NO: 564 or a fragment thereof.
In some embodiments, the peptide of the peptide active agent
conjugate comprises a sequence of any one of SEQ ID NO: 550-SEQ ID
NO: 564 or a fragment thereof. In some embodiments, the peptide
comprises a sequence that has at least 70%, at least 75%, at least
80%, at least 85%, at least 90%, at least 95%, at least 97%, at
least 99%, or 100% sequence identity with any one of SEQ ID NO:
260-SEQ ID NO: 574. In some embodiments, the peptide comprises at
least 70%, at least 75%, at least 80%, at least 85%, at least 90%,
at least 95%, at least 97%, at least 99%, or 100% sequence identity
with any one of SEQ ID NO: 550-SEQ ID NO: 564. In some embodiments,
the peptide active agent conjugate or the peptide comprises a
sequence of any one of SEQ ID NO: 1-SEQ ID NO: 23 or a fragment
thereof. In some embodiments, the peptide active agent conjugate or
the peptide comprises a sequence of any one of SEQ ID NO: 275-SEQ
ID NO: 297 or a fragment thereof. In some embodiments, the peptide
active agent conjugate or the peptide comprises a sequence of any
one of SEQ ID NO: 21-SEQ ID NO: 23 or a fragment thereof. In some
embodiments, the peptide active agent conjugate or the peptide
comprises a sequence of any one of SEQ ID NO: 295-SEQ ID NO: 297 or
a fragment thereof. In some embodiments, the peptide active agent
conjugate or the peptide comprises a peptide with at least 30%, at
least 40%, at least 50%, at least 60%, at least 70%, at least 80%,
at least 90%, at least 95%, at least, 97%, at least 98%, or at
least 99% identical to any one of SEQ ID NO: 494-SEQ ID NO: 540 or
at least 30%, at least 40%, at least 50%, at least 60%, at least
70%, at least 80%, at least 90%, at least 95%, at least 97%, at
least 98%, or at least 99% identical to any one of SEQ ID NO:
204-SEQ ID NO: 250.
Pharmacokinetics of Peptides
[0163] The pharmacokinetics of any of the peptides of this
disclosure can be determined after administration of the peptide
via different routes of administration. For example, the
pharmacokinetic parameters of a peptide of this disclosure can be
quantified after intravenous, subcutaneous, intramuscular, rectal,
aerosol, parenteral, ophthalmic, pulmonary, transdermal, vaginal,
optic, nasal, oral, sublingual, inhalation, dermal, intrathecal,
intranasal, intra-articular, peritoneal, buccal, synovial, or
topical administration. Peptides of the present disclosure can be
analyzed by using tracking agents such as radiolabels or
fluorophores. For example, a radiolabeled peptide of this
disclosure can be administered via various routes of
administration. Peptide concentration or dose recovery in various
biological samples such as plasma, urine, feces, any organ, skin,
muscle, and other tissues can be determined using a range of
methods including HPLC, fluorescence detection techniques (TECAN
quantification, flow cytometry, iVIS), or liquid scintillation
counting.
[0164] The methods and compositions described herein can relate to
pharmacokinetics of peptide administration via any route to a
subject. Pharmacokinetics can be described using methods and
models, for example, compartmental models or noncompartmental
methods. Compartmental models include but are not limited to
monocompartmental model, the two compartmental model, the
multicompartmental model or the like. Models can be divided into
different compartments and can be described by the corresponding
scheme. For example, one scheme is the absorption, distribution,
metabolism and excretion (ADME) scheme. For another example,
another scheme is the liberation, absorption, distribution,
metabolism and excretion (LADME) scheme. In some aspects,
metabolism and excretion can be grouped into one compartment
referred to as the elimination compartment. For example, liberation
can include liberation of the active portion of the composition
from the delivery system, absorption includes absorption of the
active portion of the composition by the subject, distribution
includes distribution of the composition through the blood plasma
and to different tissues, metabolism, which includes metabolism or
inactivation of the composition and finally excretion, which
includes excretion or elimination of the composition or the
products of metabolism of the composition. Compositions
administered intravenously to a subject can be subject to
multiphasic pharmacokinetic profiles, which can include but are not
limited to aspects of tissue distribution and metabolism/excretion.
As such, the decrease in plasma or serum concentration of the
composition is often biphasic, including, for example an alpha
phase and a beta phase, occasionally a gamma, delta or other phase
is observed
[0165] Pharmacokinetics includes determining at least one parameter
associated with administration of a peptide to a subject. In some
aspects, parameters include at least the dose (D), dosing interval
(.tau.), area under curve (AUC), maximum concentration (C.sub.max),
minimum concentration reached before a subsequent dose is
administered (C.sub.min), minimum time (T.sub.min), maximum time to
reach C.sub.max (T.sub.max), volume of distribution (V.sub.d),
steady-state volume of distribution (V.sub.ss), back-extrapolated
concentration at time 0 (C.sub.0), steady state concentration
(C.sub.ss), elimination rate constant (k.sub.e), infusion rate
(k.sub.in), clearance (CL), bioavailability (f), fluctuation (%
PTF) and elimination half-life (t.sub.1/2).
[0166] In certain embodiments, the peptides of any of SEQ ID NO:
1-SEQ ID NO 564 exhibit optimal pharmacokinetic parameters after
oral administration. In other embodiments, the peptides of any of
SEQ ID NO: 1-SEQ ID NO: 564 exhibit optimal pharmacokinetic
parameters after any route of administration, such as oral
administration, inhalation, intranasal administration, topical
administration, parenteral administration, intravenous
administration, subcutaneous administration, intra-articular
administration, intramuscular administration, intraperitoneal
administration, transdermal administration, dermal administration,
or any combination thereof.
[0167] In some embodiments any peptide of SEQ ID NO: 1-SEQ ID NO:
564 exhibits an average T.sub.max of 0.5-12 hours, or 1-48 hours at
which the C.sub.max is reached, an average bioavailability in serum
of 0.1%-10% in the subject after administering the peptide to the
subject by an oral route, an average bioavailability in serum of
less than 0.1% after oral administration to a subject for delivery
to the GI tract, an average bioavailability in serum of 10-100%
after parenteral administration, an average t.sub.1/2 of 0.1
hours-168 hours, or 0.25 hours-48 hours in a subject after
administering the peptide to the subject, an average clearance (CL)
of 0.5-100 L/hour or 0.5-50 L/hour of the peptide after
administering the peptide to a subject, an average volume of
distribution (V.sub.d) of 200-20,000 mL in the subject after
systemically administering the peptide to the subject, or
optionally no systemic uptake, any combination thereof.
Methods of Manufacture
[0168] Various expression vector/host systems can be utilized for
the production of the recombinant expression of peptides described
herein. Non-limiting examples of such systems include
microorganisms such as bacteria transformed with recombinant
bacteriophage DNA, plasmid DNA or cosmid DNA expression vectors
containing a nucleic acid sequence encoding peptides or peptide
fusion proteins/chimeric proteins described herein, yeast
transformed with recombinant yeast expression vectors containing
the aforementioned nucleic acid sequence, insect cell systems
infected with recombinant virus expression vectors (e.g.,
baculovirus) containing the aforementioned nucleic acid sequence,
plant cell systems infected with recombinant virus expression
vectors (e.g., cauliflower mosaic virus (CaMV), tobacco mosaic
virus (TMV) or transformed with recombinant plasmid expression
vectors (e.g., Ti plasmid) containing the aforementioned nucleic
acid sequence, or animal cell systems infected with recombinant
virus expression vectors (e.g., adenovirus, vaccinia virus)
including cell lines engineered to contain multiple copies of the
aforementioned nucleic acid sequence, either stably amplified
(e.g., CHO/dhfr, CHO/glutamine synthetase) or unstably amplified in
double-minute chromosomes (e.g., murine cell lines). Disulfide bond
formation and folding of the peptide could occur during expression
or after expression or both.
[0169] A host cell can be adapted to express one or more peptides
described herein. The host cells can be prokaryotic, eukaryotic, or
insect cells. In some cases, host cells are capable of modulating
the expression of the inserted sequences, or modifying and
processing the gene or protein product in the specific fashion
desired. For example, expression from certain promoters can be
elevated in the presence of certain inducers (e.g., zinc and
cadmium ions for metallothionine promoters). In some cases,
modifications (e.g., phosphorylation) and processing (e.g.,
cleavage) of peptide products can be important for the function of
the peptide. Host cells can have characteristic and specific
mechanisms for the post-translational processing and modification
of a peptide. In some cases, the host cells used to express the
peptides secretes minimal amounts of proteolytic enzymes.
[0170] In the case of cell- or viral-based samples, organisms can
be treated prior to purification to preserve and/or release a
target polypeptide. In some embodiments, the cells are fixed using
a fixing agent. In some embodiments, the cells are lysed. The
cellular material can be treated in a manner that does not disrupt
a significant proportion of cells, but which removes proteins from
the surface of the cellular material, and/or from the interstices
between cells. For example, cellular material can be soaked in a
liquid buffer or, in the case of plant material, can be subjected
to a vacuum, in order to remove proteins located in the
intercellular spaces and/or in the plant cell wall. If the cellular
material is a microorganism, proteins can be extracted from the
microorganism culture medium. Alternatively, the peptides can be
packed in inclusion bodies. The inclusion bodies can further be
separated from the cellular components in the medium. In some
embodiments, the cells are not disrupted. A cellular or viral
peptide that is presented by a cell or virus can be used for the
attachment and/or purification of intact cells or viral particles.
In addition to recombinant systems, Peptides can also be
synthesized in a cell-free system using a variety of known
techniques employed in protein and peptide synthesis.
[0171] In some cases, a host cell produces a peptide that has an
attachment point for a drug. An attachment point could comprise a
lysine residue, an N-terminus, a cysteine residue, a cysteine
disulfide bond, or a non-natural amino acid. The peptide could also
be produced synthetically, such as by solid-phase peptide
synthesis, or solution-phase peptide synthesis. The peptide could
be folded (formation of disulfide bonds) during synthesis or after
synthesis or both. Peptide fragments could be produced
synthetically or recombinantly and then joined together
synthetically, recombinantly, or via an enzyme.
[0172] FIG. 4 illustrates a schematic of a method of manufacturing
a construct that expresses a peptide of the disclosure, such as the
constructs illustrated in FIG. 3 and as described throughout the
disclosure and in SEQ ID NO: 1-SEQ ID NO: 564 provided herein.
[0173] In other aspects, the peptides of the present disclosure can
be prepared by conventional solid phase chemical synthesis
techniques, for example according to the Fmoc solid phase peptide
synthesis method ("Fmoc solid phase peptide synthesis, a practical
approach," edited by W. C. Chan and P. D. White, Oxford University
Press, 2000), Boc solid phase peptide synthesis, or solution phase
peptide synthesis. The disulfide bonds can be formed after cleavage
from the resin, such as by air oxidation or a buffer system with a
set pH range such as from 7-10 and can contain a redox system such
as glutathione/oxidized glutathione or cysteine/cystine. The
disulfide bonds can also be formed by selective protection and
deprotection of specific cysteine residues followed by oxidation,
or on the resin. The peptide can be purified, such as by
reversed-phase chromatography at any one or more steps during the
production process. The peptide can be isolated by lyophilization
and can be in various salt forms, such as TFA salt or ammonium and
acetate salt.
Pharmaceutical Compositions of Peptides
[0174] A pharmaceutical composition of the disclosure can be a
combination of any peptide described herein with other chemical
components, such as carriers, stabilizers, diluents, dispersing
agents, suspending agents, thickening agents, antioxidants,
solubilizers, buffers, osmolytes, salts, surfactants, amino acids,
encapsulating agents, bulking agents, cryoprotectants, and/or
excipients. The pharmaceutical composition facilitates
administration of a peptide described herein to an organism.
Pharmaceutical compositions can be administered in
therapeutically-effective amounts as pharmaceutical compositions by
various forms and routes including, for example, intravenous,
subcutaneous, intramuscular, rectal, aerosol, parenteral,
ophthalmic, pulmonary, transdermal, vaginal, optic, nasal, oral,
sublingual, inhalation, dermal, intrathecal, intranasal,
intra-articular, and topical administration. A pharmaceutical
composition can be administered in a local or systemic manner, for
example, via injection of the peptide described herein directly
into an organ, optionally in a depot.
[0175] Parenteral injections can be formulated for bolus injection
or continuous infusion. The pharmaceutical compositions can be in a
form suitable for parenteral injection as a sterile suspension,
solution or emulsion in oily or aqueous vehicles, and can contain
formulatory agents such as suspending, stabilizing and/or
dispersing agents. Pharmaceutical formulations for parenteral
administration include aqueous solutions of a peptide described
herein in water soluble form. Suspensions of peptides described
herein can be prepared as oily injection suspensions. Suitable
lipophilic solvents or vehicles include fatty oils such as sesame
oil, or synthetic fatty acid esters, such as ethyl oleate or
triglycerides, or liposomes. Aqueous injection suspensions can
contain substances which increase the viscosity of the suspension,
such as sodium carboxymethyl cellulose, sorbitol, or dextran. The
suspension can also contain suitable stabilizers or agents which
increase the solubility and/or reduce the aggregation of such
peptides described herein to allow for the preparation of highly
concentrated solutions. Alternatively, the peptides described
herein can be lyophilized or in powder form for re-constitution
with a suitable vehicle, e.g., sterile pyrogen-free water, before
use. In some embodiments, a purified peptide is administered
intravenously.
[0176] A peptide of the disclosure can be applied directly to an
organ, or an organ tissue or cells, such as brain or brain tissue
or cancer cells, during a surgical procedure. The recombinant
peptides described herein can be administered topically and can be
formulated into a variety of topically administrable compositions,
such as solutions, suspensions, lotions, gels, pastes, medicated
sticks, balms, creams, and ointments. Such pharmaceutical
compositions can contain solubilizers, stabilizers, tonicity
enhancing agents, buffers and preservatives.
[0177] In practicing the methods of treatment or use provided
herein, therapeutically-effective amounts of the peptide described
herein described herein can be administered in pharmaceutical
compositions to a subject suffering from a condition that affects
the immune system. In some embodiments, the subject is a mammal
such as a human. A therapeutically-effective amount can vary widely
depending on the severity of the disease, the age and relative
health of the subject, the potency of the compounds used, and other
factors.
[0178] Pharmaceutical compositions can be formulated using one or
more physiologically-acceptable carriers comprising excipients and
auxiliaries, which facilitate processing of the active compounds
into preparations that can be used pharmaceutically. Formulation
can be modified depending upon the route of administration chosen.
Pharmaceutical compositions comprising a peptide described herein
can be manufactured, for example, by expressing the peptide in a
recombinant system, purifying the peptide, lyophilizing the
peptide, mixing, dissolving, granulating, dragee-making,
levigating, emulsifying, encapsulating, entrapping, or compression
processes. The pharmaceutical compositions can include at least one
pharmaceutically acceptable carrier, diluent, or excipient and
compounds described herein as free-base or
pharmaceutically-acceptable salt form.
[0179] Methods for the preparation of peptides described herein
comprising the compounds described herein include formulating the
peptide described herein with one or more inert,
pharmaceutically-acceptable excipients or carriers to form a solid,
semi-solid, or liquid composition. Solid compositions include, for
example, powders, tablets, dispersible granules, capsules, cachets,
and suppositories. These compositions can also contain minor
amounts of nontoxic, auxiliary substances, such as wetting or
emulsifying agents, pH buffering agents, and other
pharmaceutically-acceptable additives.
[0180] Non-limiting examples of pharmaceutically-acceptable
excipients can be found, for example, in Remington: The Science and
Practice of Pharmacy, Nineteenth Ed (Easton, Pa.: Mack Publishing
Company, 1995); Hoover, John E., Remington's Pharmaceutical
Sciences, Mack Publishing Co., Easton, Pa. 1975; Liberman, H. A.
and Lachman, L., Eds., Pharmaceutical Dosage Forms, Marcel Decker,
New York, N.Y., 1980; and Pharmaceutical Dosage Forms and Drug
Delivery Systems, Seventh Ed. (Lippincott Williams & Wilkins
1999), each of which is incorporated by reference in its
entirety.
Administration of Pharmaceutical Compositions
[0181] A pharmaceutical composition of the disclosure can be a
combination of any peptide described herein with other chemical
components, such as carriers, stabilizers, diluents, dispersing
agents, suspending agents, thickening agents, and/or excipients.
The pharmaceutical composition facilitates administration of a
peptide described herein to an organism. Pharmaceutical
compositions can be administered in therapeutically-effective
amounts as pharmaceutical compositions by various forms and routes
including, for example, intravenous, subcutaneous, intramuscular,
rectal, aerosol, parenteral, ophthalmic, pulmonary, transdermal,
vaginal, optic, nasal, oral, inhalation, dermal, intra-articular,
intrathecal, intranasal, and topical administration. A
pharmaceutical composition can be administered in a local or
systemic manner, for example, via injection of the peptide
described herein directly into an organ, optionally in a depot.
[0182] Parenteral injections can be formulated for bolus injection
or continuous infusion. The pharmaceutical compositions can be in a
form suitable for parenteral injection as a sterile suspension,
solution or emulsion in oily or aqueous vehicles, and can contain
formulatory agents such as suspending, stabilizing and/or
dispersing agents. Pharmaceutical formulations for parenteral
administration include aqueous solutions of a peptide described
herein in water-soluble form. Suspensions of peptides described
herein can be prepared as oily injection suspensions. Suitable
lipophilic solvents or vehicles include fatty oils such as sesame
oil, or synthetic fatty acid esters, such as ethyl oleate or
triglycerides, or liposomes. Aqueous injection suspensions can
contain substances which increase the viscosity of the suspension,
such as sodium carboxymethyl cellulose, sorbitol, or dextran. The
suspension can also contain suitable stabilizers or agents which
increase the solubility and/or reduce the aggregation of such
peptides described herein to allow for the preparation of highly
concentrated solutions. Alternatively, the peptides described
herein can be lyophilized or in powder form for re-constitution
with a suitable vehicle, e.g., sterile pyrogen-free water, before
use. In some embodiments, a purified peptide is administered
intravenously. A peptide described herein can be administered to a
subject, home, target, migrates to, is retained by, and/or binds
to, or be directed to an organ, e.g., the cartilage.
[0183] A peptide of the disclosure can be applied directly to an
organ, or an organ tissue or cells, such as cartilage or cartilage
tissue or cells, during a surgical procedure. The recombinant
peptides described herein can be administered topically and can be
formulated into a variety of topically administrable compositions,
such as solutions, suspensions, lotions, gels, pastes, medicated
sticks, balms, creams, and ointments. Such pharmaceutical
compositions can contain solubilizers, stabilizers, tonicity
enhancing agents, buffers and preservatives.
[0184] In practicing the methods of treatment or use provided
herein, therapeutically-effective amounts of the peptide described
herein described herein are administered in pharmaceutical
compositions to a subject suffering from a condition. In some
instances the pharmaceutical composition will affect the physiology
of the animal, such as the immune system, inflammatory response, or
other physiologic affect. In some embodiments, the subject is a
mammal such as a human. A therapeutically-effective amount can vary
widely depending on the severity of the disease, the age and
relative health of the subject, the potency of the compounds used,
and other factors.
[0185] Pharmaceutical compositions can be formulated using one or
more physiologically-acceptable carriers comprising excipients and
auxiliaries, which facilitate processing of the active compounds
into preparations that can be used pharmaceutically. Formulation
can be modified depending upon the route of administration chosen.
Pharmaceutical compositions comprising a peptide described herein
can be manufactured, for example, by expressing the peptide in a
recombinant system, purifying the peptide, lyophilizing the
peptide, mixing, dissolving, granulating, dragee-making,
levigating, emulsifying, encapsulating, entrapping, or compression
processes. The pharmaceutical compositions can include at least one
pharmaceutically acceptable carrier, diluent, or excipient and
compounds described herein as free-base or
pharmaceutically-acceptable salt form.
[0186] Methods for the preparation of peptides described herein
comprising the compounds described herein include formulating the
peptide described herein with one or more inert,
pharmaceutically-acceptable excipients or carriers to form a solid,
semi-solid, or liquid composition. Solid compositions include, for
example, powders, tablets, dispersible granules, capsules, cachets,
and suppositories. These compositions can also contain minor
amounts of nontoxic, auxiliary substances, such as wetting or
emulsifying agents, pH buffering agents, and other
pharmaceutically-acceptable additives.
[0187] Non-limiting examples of pharmaceutically-acceptable
excipients can be found, for example, in Remington: The Science and
Practice of Pharmacy, Nineteenth Ed (Easton, Pa.: Mack Publishing
Company, 1995); Hoover, John E., Remington's Pharmaceutical
Sciences, Mack Publishing Co., Easton, Pa. 1975; Liberman, H. A.
and Lachman, L., Eds., Pharmaceutical Dosage Forms, Marcel Decker,
New York, N.Y., 1980; and Pharmaceutical Dosage Forms and Drug
Delivery Systems, Seventh Ed. (Lippincott Williams & Wilkins
1999), each of which is incorporated by reference in its
entirety.
Use of Peptide in Imaging and Surgical Methods
[0188] The present disclosure generally relates to peptides that
home, target, migrate to, are retained by, accumulate in, and/or
bind to, or are directed to specific regions, tissues, structures,
or cells within the body and methods of using such peptides. These
peptides have the ability to contact the cartilage, which makes
them useful for a variety of applications. In particular, the
peptides can have applications in site-specific modulation of
biomolecules to which the peptides are directed to. End uses of
such peptides can include, for example, imaging, research,
therapeutics, theranostics, pharmaceuticals, chemotherapy,
chelation therapy, targeted drug delivery, and radiotherapy. Some
uses can include targeted drug delivery and imaging.
[0189] In some embodiments, the present disclosure provides a
method for detecting a cancer, cancerous tissue, or tumor tissue,
the method comprising the steps of contacting a tissue of interest
with a peptide of the present disclosure, wherein the peptide is
conjugated to a detectable agent and measuring the level of binding
of the peptide, wherein an elevated level of binding, relative to
normal tissue, is indicative that the tissue is a cancer, cancerous
tissue or tumor tissue.
[0190] In some embodiments, the disclosure provides a method of
imaging an organ or body region or region, tissue or structure of a
subject, the method comprising administrating to the subject the
peptide or a pharmaceutical composition disclosed herein and
imaging the subject. In some embodiments such imaging is used to
detect a condition associated with cartilage, or a function of the
cartilage. In some cases the condition is an inflammation, a
cancer, a degradation, a growth disturbance, genetic, a tear or an
injury, or another suitable condition. In some cases the condition
is a chondrodystrophy, a traumatic rupture or detachment, pain
following surgery in regions of the body containing cartilage,
costochondritis, herniation, polychondritis, arthritis,
osteoarthritis, rheumatoid arthritis, ankylosing spondylitis (AS),
Systemic Lupus Erythematosus (SLE or "Lupus"), Psoriatic Arthritis
(PsA), gout, achondroplasia, or another suitable condition. In some
case the condition is associated with a cancer or tumor of the
cartilage. In some cases the condition is a type of chondroma or
chondrosarcoma, whether metastatic or not, or another suitable
condition. In some embodiments, such as those associated with
cancers, the imaging may be associated with surgical removal of the
diseased region, tissue, structure or cell of a subject.
[0191] Furthermore, the present disclosure provides methods for
intraoperative imaging and resection of a diseased or inflamed
tissue, cancer, cancerous tissue, or tumor tissue using a peptide
of the present disclosure conjugated with a detectable agent. In
some embodiments, the diseased or inflamed tissue, cancer,
cancerous tissue, or tumor tissue is detectable by fluorescence
imaging that allows for intraoperative visualization of the cancer,
cancerous tissue, or tumor tissue using a peptide of the present
disclosure. In some embodiments, the peptide of the present
disclosure is conjugated to one or more detectable agents. In a
further embodiment, the detectable agent comprises a fluorescent
moiety coupled to the peptide. In another embodiment, the
detectable agent comprises a radionuclide. In some embodiments,
imaging is achieved during open surgery. In further embodiments,
imaging is accomplished using endoscopy or other non-invasive
surgical techniques.
Treatment of Cartilage Disorders
[0192] The term "effective amount," as used herein, can refer to a
sufficient amount of an agent or a compound being administered
which will relieve to some extent one or more of the symptoms of
the disease or condition being treated. The result can be reduction
and/or alleviation of the signs, symptoms, or causes of a disease,
or any other desired alteration of a biological system.
Compositions containing such agents or compounds can be
administered for prophylactic, enhancing, and/or therapeutic
treatments. An appropriate "effective" amount in any individual
case can be determined using techniques, such as a dose escalation
study.
[0193] The methods, compositions, and kits of this disclosure can
comprise a method to prevent, treat, arrest, reverse, or ameliorate
the symptoms of a condition. The treatment can comprise treating a
subject (e.g., an individual, a domestic animal, a wild animal or a
lab animal afflicted with a disease or condition) with a peptide of
the disclosure. In treating a disease, the peptide can contact the
cartilage of a subject. The subject can be a human. A subject can
be a human; a non-human primate such as a chimpanzee, or other ape
or monkey species; a farm animal such as a cattle, horse, sheep,
goat, swine; a domestic animal such as a rabbit, dog, and cat; a
laboratory animal including a rodent, such as a rat, mouse and
guinea pig, or the like. A subject can be of any age. A subject can
be, for example, an elderly adult, adult, adolescent,
pre-adolescent, child, toddler, infant, or fetus in utero.
[0194] Treatment can be provided to the subject before clinical
onset of disease. Treatment can be provided to the subject after
clinical onset of disease. Treatment can be provided to the subject
after 1 day, 1 week, 6 months, 12 months, or 2 years or more after
clinical onset of the disease. Treatment may be provided to the
subject for more than 1 day, 1 week, 1 month, 6 months, 12 months,
2 years or more after clinical onset of disease. Treatment may be
provided to the subject for less than 1 day, 1 week, 1 month, 6
months, 12 months, or 2 years after clinical onset of the disease.
Treatment can also include treating a human in a clinical trial. A
treatment can comprise administering to a subject a pharmaceutical
composition, such as one or more of the pharmaceutical compositions
described throughout the disclosure. A treatment can comprise a
once daily dosing. A treatment can comprise delivering a peptide of
the disclosure to a subject, either parenterally, intravenously,
subcutaneously, intramuscularly, by inhalation, dermally,
intra-articular injection, orally, intrathecally, transdermally,
intranasally, via a peritoneal route, or directly onto or into a
joint, e.g., via topical, intra-articular injection route or
injection route of application. A treatment can comprise
administering a peptide-active agent complex to a subject, either
parenterally, intravenously, subcutaneously, intramuscularly, by
inhalation, dermally, intra-articular injection, orally,
intrathecally, transdermally, intranasally, via a peritoneal route,
or directly onto or into a joint or directly onto, near or into the
cartilage, e.g., via topical, intra-articular injection route or
injection route of application.
[0195] Types of cartilage diseases or conditions that can be
treated with a peptide of the disclosure can include inflammation,
pain management, anti-infective, pain relief, anti-cytokine,
cancer, injury, degradation, genetic basis, remodeling,
hyperplasia, surgical injury/trauma, or the like. Diseases or
conditions of bone adjacent to cartilage can also be treated with a
peptide of the disclosure. Examples of cartilage diseases or
conditions that can be treated with a peptide of the disclosure
include Costochondritis, Spinal disc herniation, Relapsing
polychondritis, Injury to the articular cartilage, any manner of
rheumatic disease (e.g., Rheumatoid Arthritis (RA), ankylosing
spondylitis (AS), Systemic Lupus Erythematosus (SLE or "Lupus"),
Psoriatic Arthritis (PsA), Osteoarthritis, Gout, and the like),
Herniation, Achondroplasia, Benign or non-cancerous chondroma,
Malignant or cancerous chondrosarcoma, Chondriodystrophies,
Chondromalacia patella, Costochondritis, Halus rigidus, Hip labral
tear, Osteochondritis dssecans, Osteochondrodysplasias, Torn
meniscus, Pectus carinatum, Pectus excavatum, Chondropathy,
Chondromalacia, Polychondritis, Relapsing Polychondritis, Slipped
epiphysis, Osteochondritis Dissecans, Chondrodysplasia,
Costochondritis, Perichondritis, Osteochondroma, Knee
osteoarthritis, Finger osteoarthritis, Wrist osteoarthritis, Hip
osteoarthritis, Spine osteoarthritis, Chondromalacia,
Osteoarthritis Susceptibility, Ankle Osteoarthritis, Spondylosis,
Secondary chondrosarcoma, Small and unstable nodules as seen in
osteoarthritis, Osteochondroses, Primary chondrosarcoma, Cartilage
disorders, scleroderma, collagen disorders, Chondrodysplasia,
Tietze syndrome, Dermochondrocomeal dystrophy of Francois,
Epiphyseal dysplasia multiple 1, Epiphyseal dysplasia multiple 2,
Epiphyseal dysplasia multiple 3, Epiphyseal dysplasia multiple 4,
Epiphyseal dysplasia multiple 5, Ossified Ear cartilages with
Mental deficiency, Muscle Wasting and Bony Changes, Periosteal
chondrosarcoma, Carpotarsal osteochondromatosis, Achondroplasia,
Genochondromatosis II, Genochondromatosis,
Chondrodysplasia--disorder of sex development, Chondroma, Chordoma,
Atelosteogenesis, type 1, Atelosteogenesis Type III,
Atelosteogenesis, type 2, Pyknoachondrogenesis, Osteoarthropathy of
fingers familial, Dyschondrosteosis--nephritis, Coloboma of
Alar-nasal cartilages with telecanthus, Alar cartilages
hypoplasia--coloboma--telecanthus, Pierre Robin syndrome--fetal
chondrodysplasia, Dysspondyloenchondromatosis, Achondroplasia
regional--dysplasia abdominal muscle, Osteochondritis Dissecans,
Familial Articular Chondrocalcinosis, Tracheobronchomalacia,
Chondritis, Dyschondrosteosis, Jequier-Kozlowski-skeletal
dysplasia, Chondrodystrophy, Cranio osteoarthropathy, Tietze's
syndrome, Hip dysplasia--ecchondromata, Bessel-Hagen disease,
Chondromatosis (benign), Enchondromatosis (benign),
Chondrocalcinosis due to apatite crystal deposition,
Meyenburg-Altherr-Uehlinger syndrome,
Enchondromatosis-dwarfism-deafness, premature growth plate closure
(e.g., due to dwarfism, injury, therapy such as retinoid therapy
for adolescent acne, or ACL repair), Astley-Kendall syndrome,
Synovial osteochondromatosis, Severe achondroplasia with
developmental delay and acanthosis nigricans, Chondrocalcinosis,
Stanescu syndrome, Familial osteochondritis dissecans,
Achondrogenesis type 1A, Achondrogenesis type 2, Achondrogenesis,
Langer-Saldino Type, Achondrogenesis type 1B, Achondrogenesis type
1A and 1B, Type II Achondrogenesis-Hypochondrogenesis,
Achondrogenesis, Achondrogenesis type 3, Achondrogenesis type 4,
Chondrocalcinosis 1, Chondrocalcinosis 2, Chondrocalcinosis
familial articular, Diastrophic dysplasia, Fibrochondrogenesis,
Hypochondroplasia, Keutel syndrome, Maffucci Syndrome,
Osteoarthritis Susceptibility 6, Osteoarthritis Susceptibility 5,
Osteoarthritis Susceptibility 4, Osteoarthritis Susceptibility 3,
Osteoarthritis Susceptibility 2, Osteoarthritis Susceptibility 1,
Pseudoachondroplasia, Cauliflower ear, Costochondritis, Growth
plate fractures, Pectus excavatum, septic arthritis, gout,
pseudogout (calcium pyrophosphate deposition disease or CPPD),
gouty arthritis, bacterial, viral, or fungal infections in or near
the joint, bursitis, tendinitis, arthropathies, or a joint disease
condition. Examples of bone diseases or conditions that can be
treated with a peptide of the disclosure include osteopenia,
post-menopausal bone loss, bone maintenance, bone fracture,
arthroplasty recovery, osteoporosis, bone loss due to metastatic
cancer, fractures due to bone loss (e.g., hip fractures in patients
with osteoporosis), pathological fracture, or atypical
fracture.
[0196] In some embodiments, a peptide or peptide conjugate of this
disclosure can be administered to a subject in order to target, an
arthritic joint. In other embodiments, a peptide or peptide
conjugate of this disclosure can be administered to a subject in
order to treat an arthritic joint.
[0197] In some embodiments, the present disclosure provides a
method for treating a cancer, the method comprising administering
to a subject in need thereof an effective amount of a peptide of
the present disclosure.
[0198] In some embodiments, the present disclosure provides a
method for treating a cancer, the method comprising administering
to a patient in need thereof an effective amount of a
pharmaceutical composition comprising a peptide of the present
disclosure and a pharmaceutically acceptable carrier.
[0199] In some embodiments, the peptides of the present disclosure
can be used to treat chondrosarcoma. Chondrosarcoma is a cancer of
cartilage producing cells and is often found in bones and joints.
It falls within the family of bone and soft-tissue sarcomas. In
certain embodiments, administration of a peptide or peptide
conjugate of the present disclosure can be used to image and
diagnose or target and treat a subject with chondrosarcoma. The
administration of a peptide or peptide conjugate of the present
disclosure can be used in combination with ablative radiotherapy or
proton therapy to treat chondrosarcoma. The subject can be a human
or an animal.
[0200] In some embodiments, a peptide or peptide conjugate of this
disclosure can be used to treat Chordoma. In certain embodiments,
administration of a peptide or peptide conjugate of the present
disclosure can be used to image and diagnose or target and treat a
subject with chordoma. The administration of a peptide or peptide
conjugate of the present disclosure can be used in combination with
a tyrosine kinase inhibitor, such as imatinib mesylate, and
ablative radiotherapy or proton therapy to treat chordoma. The
administration of a peptide or peptide conjugate of the present
disclosure can be used in combination with an antivascular agent
such as bevacizumab and an epidermal growth factor receptor
inhibitor such as erlotinib to treat chordoma. The subject can be a
human or an animal.
[0201] In some embodiments, the present disclosure provides a
method for inhibiting invasive activity of cells, the method
comprising administering an effective amount of a peptide of the
present disclosure to a subject.
[0202] In some embodiments, the peptides of the present disclosure
are conjugated to one or more therapeutic agents. In further
embodiments, the therapeutic agent is a chemotherapeutic,
anti-cancer drug, or anti-cancer agent selected from, but are not
limited to: anti-inflammatories, such as for example a
glucocorticoid, a corticosteroid, a protease inhibitor, such as for
example collagenase inhibitor or a matrix metalloprotease inhibitor
(i.e., MMP-13 inhibitor), an amino sugar, vitamin (e.g., Vitamin
D), and antibiotics, antiviral, or antifungal, a statin, an immune
modulator, radioisotopes, toxins, enzymes, sensitizing drugs,
nucleic acids, including interfering RNAs, antibodies,
anti-angiogenic agents, cisplatin, anti-metabolites, mitotic
inhibitors, growth factor inhibitors, paclitaxel, temozolomide,
topotecan, fluorouracil, vincristine, vinblastine, procarbazine,
decarbazine, altretamine, methotrexate, mercaptopurine,
thioguanine, fludarabine phosphate, cladribine, pentostatin,
cytarabine, azacitidine, etoposide, teniposide, irinotecan,
docetaxel, doxorubicin, daunorubicin, dactinomycin, idarubicin,
plicamycin, mitomycin, bleomycin, tamoxifen, flutamide, leuprolide,
goserelin, aminogluthimide, anastrozole, amsacrine, asparaginase,
mitoxantrone, mitotane and amifostine, and their equivalents, as
well as photo-ablation. Some of these active agents induce
programmed cell death such as apoptosis in target cells and thereby
improve symptoms or ameliorate disease. Apoptosis can be induced by
many active agents, including, for example, chemotherapeutics,
anti-inflammatories, corticosteroids, NSAIDS, tumor necrosis factor
alpha (TNF-.alpha.) modulators, tumor necrosis factor receptor
(TNFR) family modulators. In some embodiments, peptides of this
disclosure can be used to target active agents to pathways of cell
death or cell killing, such as caspases, apoptosis activators and
inhibitors, XBP-1, Bcl-2, Bcl-X1, Bcl-w, and other disclosed
herein. In other embodiments, the therapeutic agent is any
nonsteroidal anti-inflammatory drug (NSAID). The NSAID can be any
heterocyclic acetic acid derivatives such as ketorolac,
indomethacin, etodolac, or tolemetin, any propionic acid
derivatives such as naproxen, any enolic acid derivatives, any
anthranilic acid derivatives, any selective COX-2 inhibitors such
as celecoxib, any sulfonanilides, any salicylates, aceclofenac,
nabumetone, sulindac, diclofenac, or ibuprofen. In other
embodiments, the therapeutic agent is any steroid, such as
dexamethasone, budesonide, beclomethasone monopropionate,
desciclesonide, triamcinolone, cortisone, prednisone, prednisolone,
triamcinolone hexacetonide, or methylprednisolone. In other
embodiments, the therapeutic agent is a pain reliever, such as
acetaminophen, opioids, local anesthetics, anti-depressants,
glutamate receptor antagonists, adenosine, or neuropeptides. In
some embodiments, a treatment consists of administering a
combination of any of the above therapeutic agents and a peptide
conjugate, such as a treatment in which both a
dexamethasone-peptide conjugate and an NSAID are administered to a
patient. Peptides of the current disclosure that target the
cartilage can be used to treat the diseases conditions as described
herein, for example, any diseases or conditions including tears,
injuries (i.e., sports injuries), genetic factors, degradation,
thinning, inflammation, cancer or any other disease or condition of
the cartilage or to target therapeutically-active substances to
treat these diseases amongst others. In other cases, a peptide of
the disclosure can be used to treat traumatic rupture, detachment,
chostochondritis, spinal disc herniation, relapsing and
non-relapsing polychondritis, injury to the articular cartilage,
osteoarthritis, arthritis or achondroplasia. In some cases, the
peptide or peptide-active agent can be used to target cancer in the
cartilage, for example benign chondroma or malignant
chondrosarcoma, by contacting the cartilage by diffusion into
chondrocytes and then having antitumor function, targeted toxicity,
inhibiting metastases, etc. As well, such peptide or peptide-active
agent can be used to label, detect, or image such cartilage
lesions, including tumors and metastases amongst other lesions,
which may be removed through various surgical techniques or by
targeting with peptide-active agents that induce programmed cell
death or kill cells.
[0203] Venom or toxin derived peptide(s), peptides, modified
peptides, labeled peptides, peptide-active agent conjugates and
pharmaceutical compositions described herein can be administered
for prophylactic and/or therapeutic treatments. In therapeutic
applications, the composition can be administered to a subject
already suffering from a disease or condition, in an amount
sufficient to cure or at least partially arrest the symptoms of the
disease or condition, or to cure, heal, improve, or ameliorate the
condition. Such peptides described herein can also be administered
to prevent (either in whole or in part), lessen a likelihood of
developing, contracting, or worsening a condition. Amounts
effective for this use can vary based on the severity and course of
the disease or condition, previous therapy, the subject's health
status, weight, response to the drugs, and the judgment of the
treating physician. Venom or toxin derived peptide(s), peptides,
modified peptides, labeled peptides, peptide-active agent
conjugates and pharmaceutical compositions described herein can
allow for targeted homing of the peptide and local delivery of any
conjugate. For example, a peptide conjugated to a steroid allows
for local delivery of the steroid, which is significantly more
effective and less toxic than traditional systemic steroids. A
peptide conjugated to an NSAID is another example. In this case,
the peptide conjugated to an NSAID allows for local delivery of the
NSAID, which allows for administration of a lower NSAID dose and is
subsequently less toxic. By delivering an active agent to the
joint, pain relief can be more rapid, may be more long lasting, and
can be obtained with a lower systemic dose and off-site undesired
effects than with systemic dosing without targeting.
[0204] Peptides of the current disclosure that target the cartilage
can be used to treat or manage pain associated with a cartilage
injury or disorder, or any other cartilage or joint condition as
described herein. The peptides can be used either directly or as
carriers of active drugs, peptides, or molecules. For example,
since ion channels can be associated with pain and can be activated
in disease states such as arthritis, peptides that interact with
ion channels can be used directly to reduce pain. In another
embodiment, the peptide is conjugated to an active agent with
anti-inflammatory activity, in which the peptide acts as a carrier
for the local delivery of the active agent to reduce pain.
[0205] In some embodiments, the peptides described herein provide a
method of treating a cartilage condition of a subject, the method
comprising administering to the subject a therapeutically-effective
amount of a peptide comprising the sequence SEQ ID NO: 1 or
fragment thereof. In some embodiments, the peptides described
herein provide a method of treating a cartilage condition of a
subject, the method comprising administering to the subject a
peptide of any one of SEQ ID NO: 2-SEQ ID NO: 564 or fragment
thereof.
Treatment of Kidney Disorders
[0206] In some embodiments, peptides of this disclosure that home,
target, are directed to, migrate to, are retained by, accumulate
in, or bind to specific regions, tissues, structures or cells of
the kidneys can be used to treat a kidney disorder. In other
embodiments, peptides are used in peptide conjugates of the present
disclosure to deliver an active agent for treatment of a kidney
disorder.
[0207] In some embodiments, the peptides and peptide-conjugates of
the present disclosure are used to treat a condition of the kidney,
or a region, tissue, structure, or cell thereof. In certain
embodiments, the condition is associated with kidney, or a function
of a subject's kidneys. The present disclosure encompasses various
acute and chronic renal diseases, including glomerular,
tubule-interstitial, and microvascular diseases. Examples of
conditions applicable to the present disclosure include but are not
limited to: hypertensive kidney damage, acute kidney diseases and
disorders (AKD), acute kidney injury (AM) due to
ischemia-reperfusion injury, drug treatment such as chemotherapy,
cardiovascular surgery, surgery, medical interventions or
treatment, radiocontrast nephropathy, or induced by cisplatin or
carboplatin, which can be treated prophylactically, established AM
including ischemic renal injury, endotoxemia-induced AM,
endotoxemia/sepsis syndrome, or established nephrotoxic AM (e.g.,
rhabdomyolysis, radiocontrast nephropathy, cisplatin/carboplatin
AM, aminoglycoside nephrotoxicity), end stage renal disease, acute
and rapidly progressive glomerulonephritis, acute presentations of
nephrotic syndrome, acute pyelonephritis, acute renal failure,
chronic glomerulonephritis, chronic heart failure, chronic
interstitial nephritis, graft versus host disease after renal
transplant, chronic kidney disease (CKD) such as diabetic
nephropathy, hypertensive nephrosclerosis, idiopathic chronic
glomerulonephritis (e.g., focal glomerular sclerosis, membranous
nephropathy, membranoproliferative glomerulonephritis, minimal
change disease transition to chronic disease, anti-GBM disease,
rapidly progressive cresentic glomerulonephritis, IgA nephropathy),
secondary chronic glomerulonephritis (e.g., systemic lupus,
polyarteritis nodosa, scleroderma, amyloidosis, endocarditis),
hereditary nephropathy (e.g., polycystic kidney disease, Alport's
syndrome), interstitial nephritis induced by drugs (e.g., Chinese
herbs, NSAIDs), multiple myeloma or sarcoid, or renal
transplantation such as donor kidney prophylaxis (treatment of
donor kidney prior to transplantation), treatment post
transplantation to treat delayed graft function, acute rejection,
or chronic rejection, chronic liver disease, chronic
pyelonephritis, diabetes, diabetic kidney disease, fibrosis, focal
segmental glomerulosclerosis, Goodpasture's disease, hypertensive
nephrosclerosis, IgG4-related renal disease, interstitial
inflammation, lupus nephritis, nephritic syndrome, partial
obstruction of the urinary tract, polycystic kidney disease,
progressive renal disease, renal cell carcinoma, renal fibrosis,
and vasculitis. For example, in certain embodiments, the peptides
and peptide-conjugates of the present disclosure are used to reduce
acute kidney injury in order to prevent it from progressing to
chronic kidney disease.
[0208] Alternatively or in combination, in some embodiments, the
peptide and peptide-conjugates of the present disclosure are used
to elicit a protective response such as ischemic preconditioning
and/or acquired cytoresistance in a kidney of the subject. In some
embodiments, ischemic preconditioning and/or acquired
cytoresistance is induced by administering an agent (e.g., a
peptide or peptide-conjugate of the present disclosure) that
upregulates the expression of protective stress proteins, such as
antioxidants, anti-inflammatory proteins, or protease inhibitors.
In certain embodiments, the induced response protects the kidney by
preserving kidney function in whole or in part and/or by reducing
injury to renal tissues and cells, e.g., relative to the situation
where no protective response is induced. The peptides and
peptide-conjugates of the present disclosure can provide certain
benefits compared to other agents for inducing ischemic
preconditioning and/or acquired cytoresistance, such as a
well-defined chemical structure and avoidance of low pH
precipitation.
[0209] In some embodiments, the protective response is induced in
order to protect the kidney or tissues or cells thereof from an
injury or insult that is predicted to occur (e.g., associated with
a planned event such as a medical procedure, is likely to occur due
to a condition in the subject) or has already occurred. In certain
embodiments, the induced response prevents or reduces the extent of
damage to the kidney or tissues or cells thereof caused by the
injury or insult. For instance, in certain embodiments, the
peptides and peptide-conjugates induce acquired cytoresistance by
activating protective pathways and/or upregulating expression of
protective stress proteins. Optionally, the peptides and
peptide-conjugates are capable of inducing such protective
responses while causing minimal or no injury to the kidney.
[0210] In various embodiments, the injury or insult is associated
with one or more of: surgery, radiocontrast imaging,
cardiopulmonary bypass, balloon angioplasty, induced cardiac or
cerebral ischemic-reperfusion injury, organ transplantation,
sepsis, shock, low blood pressure, high blood pressure, kidney
hypoperfusion, chemotherapy, drug administration, nephrotoxic drug
administration, blunt force trauma, puncture, poison, or smoking.
For instance, in certain embodiments, the injury or insult is
associated with a medical procedure that has been or will be
performed on the subject, such as one or more of: surgery,
radiocontrast imaging, cardiopulmonary bypass, balloon angioplasty,
induced cardiac or cerebral ischemic-reperfusion injury, organ
transplantation, chemotherapy, drug administration, or nephrotoxic
drug administration.
[0211] In some embodiments, the peptide itself exhibits a renal
therapeutic effect. For example, in certain embodiments, the
cystine-dense peptide interacts with a renal ion channel, inhibits
a protease, has antimicrobial activity, has anticancer activity,
has anti-inflammatory activity, induces ischemic preconditioning or
acquired cytoresistance, or produces a protective or therapeutic
effect on a kidney of the subject, or a combination thereof.
Optionally, the renal therapeutic effect exhibited by the peptide
is a renal protective effect or renal prophylactic effect (e.g.,
ischemic preconditioning or acquired cytoresistance) that protects
the kidney or a tissue or cell thereof from an upcoming injury or
insult.
[0212] For example, in certain embodiments, a peptide of the
present disclosure activates protective pathways and/or upregulates
expression of protective stress proteins in the kidney or tissues
or cells thereof. As another example, in certain embodiments, a
peptide of the present disclosure accesses and suppresses
intracellular injury pathways. In yet another example, in certain
embodiments, a peptide of the present disclosure inhibits
interstitial inflammation and prevents renal fibrosis. As a further
example, in certain embodiments, a peptide of the present
disclosure is administered prior to or currently with the
administration of a nephrotoxic agent (e.g., aminoglycoside
antibiotics such as gentamicin and minocycline, chemotherapeutics
such as cisplatin, immunoglobulins or fragments thereof, mannitol,
NSAIDs such as ketorolac or ibuprofen, cyclosporin,
cyclophosphamide, radiocontrast dyes) in order to minimize its
damaging effects, e.g., by blocking megalin-cubulin binding sites
so that the nephrotoxic agent passes through the kidneys.
[0213] In some embodiments, the present disclosure provides that
any peptide of the disclosure including SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564 can as a peptide conjugate
with an active agent for treatment of a kidney disorder. For
example, a peptide of SEQ ID NO: 27, SEQ ID NO: 108, or SEQ ID NO:
199 can be conjugated to an active agent and administered to a
subject in need thereof to treat a kidney disorder.
[0214] In some embodiments, homing of a peptide of this disclosure
to cartilage or the kidneys can be assessed in an animal model such
as those described in Alves et al. (Clin Rev Allergy Immunol. 2016
August; 51(1):27-47. doi: 10. 1007/s12016-015-8522-7), Kuyinu et
al. (J Orthop Surg Res. 2016 Feb. 2; 11:19. doi: 10.
1186/s13018-016-0346-5), Li et al. (Exp Biol Med (Maywood). 2015
August; 240(8):1029-38. doi: 10. 1177/1535370215594583), and
Mullins et al. (Dis Model Mech. 2016 Dec. 1; 9(12):1419-1433), all
of which are incorporated herein by reference.
[0215] Multiple peptides described herein can be administered in
any order or simultaneously. In some cases, multiple functional
fragments of peptides derived from toxins or venom can be
administered in any order or simultaneously. If simultaneously, the
multiple peptides described herein can be provided in a single,
unified form, such as an intravenous injection, or in multiple
forms, such as subsequent intravenous dosages.
[0216] Peptides can be packaged as a kit. In some embodiments, a
kit includes written instructions on the use or administration of
the peptides.
EXAMPLES
[0217] The following examples are included to further describe some
embodiments of the present disclosure, and should not be used to
limit the scope of the disclosure.
Example 1
Manufacture of Peptides
[0218] The peptide sequence was reverse-translated into DNA,
synthesized, and cloned in-frame with siderocalin using standard
molecular biology techniques. (M. R. Green, Joseph Sambrook.
Molecular Cloning. 2012 Cold Spring Harbor Press.). The resulting
construct was packaged into a lentivirus, transfected into HEK293
cells, expanded, isolated by immobilized metal affinity
chromatography (IMAC), cleaved with tobacco etch virus protease,
and purified to homogeneity by reverse-phase chromatography.
Following purification, each peptide was lyophilized and stored
frozen.
Example 2
Radiolabeling of Peptide
[0219] This example describes radiolabeling of peptides with
standard techniques. See J Biol Chem. 254(11):4359-65 (1979). The
sequences were engineered to have the amino acids, "G" and "S" at
the N terminus. See Methods in Enzymology V91:1983 p. 570 and
Journal of Biological Chemistry 254(11):1979 p. 4359. An excess of
formaldehyde was used to ensure complete methylation (dimethylation
of every free amine). The labeled peptides were isolated via
solid-phase extraction on Strata-X columns (Phenomenex
8B-S100-AAK), rinsed with water with 5% methanol, and recovered in
methanol with 2% formic acid. Solvent was subsequently removed in a
blowdown evaporator with gentle heat and a stream of nitrogen
gas.
Example 3
Peptide Detectable Agent Conjugates
[0220] This example describes the dye labeling of peptides. A
peptide of the disclosure is expressed recombinantly or chemically
synthesized, and then the N-terminus of the peptide is conjugated
to a detectable agent via an NHS ester using DCC or EDC to produce
a peptide-detectable agent conjugate. The detectable agent is the
fluorophore dye is a cyanine dye, such as Cy5.5 or an Alexa
fluorophore, such as Alexa647.
[0221] The peptide detectable agent conjugates are administered to
a subject. The subject can be a human or a non-human animal. After
administration, the peptide detectable agent conjugates home to
cartilage. The subject, or a biopsy from the subject, can be imaged
to visualize localization of the peptide detectable agent
conjugates to cartilage. In some aspects, visualization of the
peptide detectable agent conjugates in cartilage after
administration results in diagnosis of arthritis, cartilage damage,
or any cartilage disorder.
Example 4
Method to Determine Improved Peptide Variants
[0222] This example shows a method for determining ways to improve
peptide variants by comparing and analyzing the primary sequences
and tertiary structures of scaffold peptides. FIG. 5A-FIG. 5C show
sequences of SEQ ID NO: 541 aligned with SEQ ID NO: 316, SEQ ID NO:
541 aligned with SEQ ID NO: 542, and SEQ ID NO: 541 aligned with
SEQ ID NO: 483. The sequence alignment of the two scaffolds was
used to identify conserved positively charged residues (shown in
boxes) that may be important for cartilage homing. A peptide of SEQ
ID NO: 483 homes to cartilage and other peptides with positively
charged residues in similar positions, or cysteines in similar
positions, or other residues that are in similar positions are also
predicted to home to cartilage.
[0223] FIG. 6 shows sequences of SEQ ID NO: 320 aligned with SEQ ID
NO: 484. The sequence alignment of the two scaffolds was used to
identify the basic/aromatic dyad that may be involved in the
interaction with the Kv ion channel (K27 and Y36 of SEQ ID NO:
484). The mutation of K27 to alanine, arginine, or glutamic acid
destroyed activity against the squid Kv1A ion channel. K27 and Y36
may be desirable to maintain or add to a cartilage homing peptide
of this disclosure to maintain or improve homing, to maintain or
improve residence time in cartilage, or to maintain or improve
modulation of an ion channel such as Kv. In contrast, K27 and Y36
may be desirable to mutate out of a cartilage homing peptide to
reduce interaction with an ion channel such as Kv. Disruption of
either the basic or aromatic residue eliminates ion channel
activity. In another example, D amino acids are expected to reduce
or eliminate binding.
Example 5
Sequence Alignment to pFam00451:Toxin_2 Family to Identify
Cartilage Homing Peptides
[0224] This example describes a method for identifying new
cartilage homing peptides by sequence alignment to the
pFam00451:toxin 2 structural class family. The pFam00451:toxin 2
structural class is a family of peptides related by similarities in
sequence identity. FIG. 7 illustrates alignment of peptides within
the pfam00451:toxin_2 structural class family of SEQ ID NO: 494-SEQ
ID NO: 540. Boxed and bolded residues indicate relative
conservation of sequence while non-boxed and non-bolded residues
indicate areas of higher sequence variability. SEQ ID NO: 494 was
identified as a cartilage homing candidate peptide based on its
structural similarities with the pFam00451:toxin_structural class
family. FIG. 8 illustrates the sequence alignment of a peptide of
SEQ ID NO: 494 from the pfam00451:toxin_2 structural class family
with the sequence of SEQ ID NO: 27. Asterisks indicate positions
with a single, fully conserved residue, a colon indicates
conservation between groups of strongly similar properties (scoring
>0.5 in the Gonnet point accepted mutation (PAM) 250 matrix),
and a period indicates conservation between groups of weakly
similar properties (scoring .ltoreq.0.5 in the Gonnet PAM 250
matrix). SEQ ID NO: 108 was also identified as a cartilage homing
candidate based on its structural similarities with the
pfam00451:toxin_2 structural class family of peptides.
[0225] The pFam00451:toxin_2 structural class family is used as a
scaffold to identify variant peptides that have cartilage homing
properties. Any member of the pFam00451:toxin_2 structural class
family is used to predict new cartilage homing peptides based on
homology, preserved residues, or a preserved cysteine residue.
Example 6
Dosing of Peptide with Kidney Ligation
[0226] This example describes a dosing scheme for administering
peptides to mice in conjunction with kidney ligation. Different
dosages of the peptides were administered to Female Harlan athymic
nude mice, weighing 20 g-25 g, via tail vein injection (n=2 mice
per peptide). The sequence of thirteen cartilage homing peptides of
SEQ ID NO: 24-SEQ ID NO: 36 are shown in TABLE 1. The experiment
was done in duplicates. The kidneys were ligated to prevent renal
filtration of the peptides. Each peptide was radiolabeled by
methylating lysines and the N-terminus, so the actual binding agent
may contain methyl or dimethyl lysine(s) and a methylated or
dimethylated amino terminus.
[0227] A target dosage of 50-100 nmol of each peptide carrying
10-25 uCi of .sup.14C was administered to Female Harlan athymic
nude mice while anesthetized. Each peptide was allowed to freely
circulate within the animal before the animals were euthanized and
sectioned.
Example 7
Peptide Homing with Kidney Ligation
[0228] This example illustrates peptide homing to cartilage of mice
with kidneys that were ligated prior to peptide administration. At
the end of the dosing period in EXAMPLE 6, mice were frozen in a
hexane/dry ice bath and then frozen in a block of
carboxymethylcellulose. Whole animal sagittal slices were prepared
that resulted in thin frozen sections being available for imaging.
Thin, frozen sections of animal including imaging of tissues such
as brain, tumor, liver, kidney, lung, heart, spleen, pancreas,
muscle, adipose, gall bladder, upper gastrointestinal tract, lower
gastrointestinal tract, bone, bone marrow, reproductive track, eye,
cartilage, stomach, skin, spinal cord, bladder, salivary gland, and
other types of tissues were obtained with a microtome, allowed to
desiccate in a freezer, and exposed to phosphoimager plates for
about ten days.
[0229] These plates were developed, and the signal (densitometry)
from each organ was normalized to the signal found in the heart
blood of each animal. A signal in tissue darker than the signal
expected from blood in that tissue indicates peptide accumulation
in a region, tissue, structure or cell. For instance, the cartilage
is avascular and contains minute amounts of blood. A ratio of at
least 170% signal in the cartilage versus heart ventricle was
chosen as a reference level for significant targeting to cartilage,
which also correlated with clear accumulation in cartilaginous
tissues in the images of the slices. FIG. 1 identifies the
locations of the SEQ ID NO: 27 peptide distribution in joint and
other cartilage. FIG. 12 identifies the locations of the SEQ ID NO:
27 peptide distribution in nasal, spinal, tracheal, and other
cartilage, including to hyaline cartilage such as articular
cartilage and physeal cartilage, as well as fibrocartilage.
[0230] Additionally, the peptide can be retained in cartilage for
hours after treatment. The SEQ ID NO: 27 peptide was radiolabeled
as in EXAMPLE 6 and 100 nmol of peptide was injected into a mouse
with intact kidneys. FIG. 9 illustrates the retention of and the
tissue distribution in the cartilage of a peptide of SEQ ID NO: 27,
24 hours after administration.
Example 8
Dosing of Peptide without Kidney Ligation
[0231] This example describes a dosing scheme for administering
peptides to mice without kidney ligation. The peptide administered
had the sequence of SEQ ID NO: 27 as shown in TABLE 1. The peptide
was radiolabeled by methylating lysines and the N-terminus, so the
actual binding agent may contain methyl or dimethyl lysine(s) and a
methylated or dimethylated amino terminus.
[0232] A target dosage of 100 nmol of each peptide carrying 10-25
.mu.Ci of .sup.14C was administered to Female Harlan athymic nude
mice by a tail vein injection. Each peptide was allowed to freely
circulate within the animal for either 4 hours or 24 hours before
the animals were euthanized and sectioned.
Example 9
Peptide Homing with Intact Kidneys
[0233] This example illustrates peptide homing to cartilage in
animals with intact kidneys. At the end of the 4 hour or 24 hour
dosing periods in EXAMPLE 8, mice were frozen in a hexane/dry ice
bath and then frozen in a block of carboxymethylcellulose. Whole
animal sagittal slices were prepared that resulted in thin frozen
sections being available for imaging. Thin, frozen sections of
animal including imaging of tissues such as brain, tumor, liver,
kidney, lung, heart, spleen, pancreas, muscle, adipose, gall
bladder, upper gastrointestinal track, lower gastrointestinal
track, bone, bone marrow, reproductive track, eye, cartilage,
stomach, skin, spinal cord, bladder, salivary gland, and other
types of tissues were obtained with a microtome, allowed to
desiccate in a freezer, and exposed to phosphoimager plates for
about ten days.
[0234] These plates were developed. A signal in tissue darker than
the signal expected from blood in that tissue indicates peptide
accumulation in a region, tissue, structure or cell. For instance,
the cartilage is avascular and contains minute amounts of blood.
High signal in the kidneys indicates presence and accumulation of
the peptide in the kidneys. FIG. 1 identifies the locations of the
SEQ ID NO: 27 peptide distribution in joint and other cartilage as
well as kidneys.
Example 10
Peptide Homing with Therapeutic Agents
[0235] This example describes certain exemplary therapeutic agents
that are conjugated to a peptide. A peptide of the disclosure is
expressed recombinantly or chemically synthesized and then is
conjugated to an exemplary drug, such as paclitaxel or
triamcinolone acetonide or budesonide using techniques known in the
art, such as those described in Bioconjugate Techniques by Greg
Hermanson (Elsevier Inc., 3.sup.rd Edition, 2013). One or more
drugs is conjugated per peptide, or an average of less than one
drug is conjugated per peptide.
[0236] Coupling of these drugs to a peptide of any of SEQ ID NO:
24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564 targets the drug
to the cartilage of the subject. One or more drug-peptide
conjugates are administered to a human or animal.
Example 11
Peptide Homing to an Arthritic Joint
[0237] This example illustrates peptide homing to cartilage in
humans or animals with arthritis. A peptide of the present
disclosure is expressed recombinantly or chemically synthesized and
is used directly, after radiolabeling, or after conjugation to a
fluorophore or therapeutic compound. A peptide is selected from any
one of the peptides of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO:
314-SEQ ID NO: 564. The peptide or peptide conjugate is
administered to a human or animal subcutaneously, intravenously, or
orally, or is injected directly into a joint intraarticularly. The
peptide or peptide conjugate homes to cartilage.
Example 12
Peptide Homing to Cartilage in Non-Human Animals
[0238] This example illustrates a peptide or peptide conjugate of
this disclosure homing to cartilage in non-human animals. Non-human
animals include but are not limited to guinea pigs, rabbits, dog,
cats, horses, rats, mice, cows, pigs, non-human primates, and other
non-human animals. A peptide of the present disclosure is
recombinantly expressed or chemically synthesized and are used
directly, after radiolabeling, or after conjugation to a
fluorophore or therapeutic compound. The peptide is selected from
any one of the peptides of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID
NO: 314-SEQ ID NO: 564. The resulting peptide or peptide conjugate
is administered to a non-human animal subcutaneously,
intravenously, or orally, or is injected directly into a joint
intra-articularly. Biodistribution is assessed by LC/MS,
autoradiography, positron emission tomography (PET), or
fluorescence imaging. A peptide or peptide conjugate is homed to
cartilage in non-human animals.
Example 13
Whole Body Fluorescence and Isolated Limb Fluorescence of Homing
Peptides
[0239] This example illustrates whole body fluorescence and
isolated limb fluorescence of peptide homers of this disclosure.
Any peptide of the present disclosure is chemically conjugated to
one molecule of a near infrared fluorophore, at the N-terminus of
the peptide via an active NHS ester on the dye. A dose of 10 nmol
of each peptide conjugated to a fluorophore is administered to
Female Harlan athymic nude mice, weighing 20-25 g, and is
administered via tail vein injection. Each experiment is done at
least in duplicate (n=2 mice per group). The peptide fluorophore
conjugate is allowed to freely circulate for the described time
period before the mice were euthanized at various time points. Mice
are evaluated for peptide distribution of the peptide fluorescence
in whole body imaging and in isolated hind limb imaging.
[0240] For Whole body fluorescence (WBF), at the end of the dosing
period, mice are frozen in a hexane/dry ice bath and then embedded
in a frozen block of carboxymethylcellulose. Whole animal sagittal
slices are prepared that resulted in thin frozen sections for
imaging. Thin frozen sections are obtained using a microtome and
allowed visualization of tissues. Sections are allowed to dessicate
in a freezer prior to imaging. WBF is performed on fluorescent
sections, which are scanned on a Li-Cor Odyssey scanner at a
setting of 169 .mu.m resolution, medium quality, 700 channel, L-2.0
intensity.
[0241] For isolated hind limb fluorescence studies, mice are
euthanized by CO.sub.2 asphyxiation at the end of the dosing
period. The right hind limb is removed at the hip joint and imaged
on a Sepctrum IVIS imager (ex/em: 675 nm. 720 nm) with a 1 second
exposure length and a focal height of 0.5 cm. Limbs are imaged with
skin removed and with muscle removed.
Example 14
Whole Body Autoradiography of Homing Peptides
[0242] This example illustrates whole body autoradiography of
peptide homers of this disclosure. Peptides are radiolabeled by
methylating lysines at the N-terminus as described in EXAMPLE 2. As
such, the peptide may contain methyl or dimethyl lysines and a
methylated or demethylated amino terminus. A dose of 100 nmol
radiolabeled peptide is administered via tail vein injection in
Female Harlan athymic nude mice, weighing 20-25 g. The experiment
is done in at least duplicate (n=2 animals per group). In some
animals, kidneys are ligated to prevent renal filtration of the
radiolabled peptides and extend plasma half-life. Each radiolabeled
peptide is allowed to freely circulate within the animal for the
described time period before the animals were euthanized and
sectioned.
[0243] Whole body autoradiography (WBA) sagittal sectioning is
performed as follows. At the end of the dosing period, mice are
frozen in a hexane/dry ice bath and then embedded in a frozen block
of carboxymethylcellulose. Whole animal sagittal slices are
prepared that resulted in thin frozen sections for imaging. Thin
frozen sections are obtained using a microtome and allowed
visualization of tissues such as brain, tumor, liver, kidney, lung,
heart, spleen, pancreas, muscle, adipose, gall bladder, upper
gastrointestinal tract, lower gastrointestinal tract, bone, bone
marrow, reproductive tract, eye, cartilage, stomach, skin, spinal
cord, bladder, salivary gland, and more. Sections are allowed to
dessicate in a freezer prior to imaging.
[0244] For the autoradiography imaging, tape mounted thin sections
are freeze dried and radioactive samples were exposed to
phosphoimager plates for 7 days. These plates are developed and the
signal (densitometry) from each organ was normalized to the signal
found in the cardiac blood of each animal. A signal in tissue
darker than the signal expected from blood in that tissue indicates
accumulation in a region, tissue, structure, or cell.
Example 15
Peptide Localization in Chondrocytes
[0245] This example illustrates binding of peptides of this
disclosure to chondrocytes within cartilage in animals with intact
kidneys. In one embodiment, animals are dosed and are processed as
described in EXAMPLE 13 and EXAMPLE 14. At the end of the dosing
period, animals are euthanized and cartilage is optionally removed
for use in staining and imaging procedures. Whole animal sagittal
slices are prepared that result in thin frozen sections being
available for staining and imaging. One or more of the following
cartilage components are identified in thin frozen sections or live
cartilage explants using standard staining techniques: collagen
fibrils, glycosaminoglycans, or chondrocytes. A peptide of this
disclosure is found to localize to chondrocytes in cartilage,
localized intracellularly or extracellularly bound or both.
Localization is visualized and confirmed by microscopy.
[0246] In another embodiment, peptides or peptide-drug conjugates
of this disclosure are administered in humans and are localized on
or in chondrocytes in cartilage.
Example 16
Peptide Localization in Cartilage Extracellular Matrix
[0247] This example illustrates localization of peptides of this
disclosure in cartilage extracellular matrix. In one embodiment,
animals are dosed and are processed as described in EXAMPLE 13 and
EXAMPLE 14 in animals with intact kidneys. At the end of the dosing
period, animals are euthanized and cartilage is optionally removed
for use in staining and imaging procedures. Whole animal sagittal
slices are prepared that result in thin frozen sections being
available for staining and imaging. Thin frozen sections or live
cartilage explants are acquired, stained, and visualized as
described in EXAMPLE 15. A peptide of the present disclosure is
found to localize to the extracellular matrix in cartilage. The
peptide may be bound to one or more components of the extracellular
matrix, such as proteoglycans, glycosaminoglycans, aggrecan,
decorin, or collagen. Localization is visualized and confirmed by
microscopy.
[0248] In another embodiment, peptides or peptide-drug conjugates
of this disclosure are administered in humans and are localized in
cartilage extracellular matrix.
Example 17
Peptide Binding to Cartilage Explants
[0249] This example illustrates a peptide or peptide conjugation of
this disclosure homing, targeting, being directed to, migrating to,
being retained by, accumulating in, or binding to human and animal
cartilage explants in culture. A peptide is selected from any one
of the peptides of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO:
314-SEQ ID NO: 564. Peptides are recombinantly expressed or
chemically synthesized and are used directly, after radiolabeling,
or after conjugation to a fluorophore or therapeutic compound. A
peptide of peptide conjugate of this disclosure is incubated with
cartilage explants derived from humans or animals. Peptides of
peptide conjugate are found to bind to cartilage explants. The
interaction with cartilage is confirmed using various methods that
include but are not limited to liquid scintillation counting,
confocal microscopy, immunohistochemistry, HPLC, or LC/MS. The
peptide shows a higher level of signal than a control peptide that
is administered that is not a cartilage binding peptide.
Example 18
Effects of Peptide on Ion Channels
[0250] This example describes the interaction between peptides of
the present disclosure and ion channels. Ion channels can be
associated with pain and can be activated in disease states such as
arthritis. A peptide of the disclosure is expressed and
administered in a pharmaceutical composition to a patient to treat
a joint condition or disease associated with an ion channel and
treatable by binding, blocking, or interacting with the ion
channel. Ion channels, such as Nav 1. 7, are inhibited by peptides
of the present disclosure. A given peptide is expressed
recombinantly or chemically synthesized, wherein the peptide
selected from SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID
NO: 564. Following expression or synthesis, the peptide is used
directly or conjugated to a therapeutic compound, such as those
described herein. A peptide of the present disclosure selectively
interacts with ion channels, or is mutated in order to interact
with ion channels. For example, a peptide of this disclosure is
bound to Nav 1. 7 or Nav 1. 7 is blocked by a peptide of this
disclosure. When the peptide is administered to a human subject,
Nav 1.7 signaling is reduced in the tissues in proximity to the
joints, and pain relief is thereby provided.
Example 19
Peptide-Fc Protein Fusions
[0251] This example illustrates making and using peptide-Fc protein
fusions. A peptide of SEQ ID NO: 108 was recombinantly expressed
with the sequence for the human IgG1 Fc protein in HEK293 cells to
yield a sequence of
TABLE-US-00033 SEQ ID NO: 565
(METDTLLLWVLLLWVPGSTGGSGVPINVRCRGSRDCLDPCRRAGMRFGR
CINSRCHCTPGGSGGSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMIS
RTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS
VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPS
RDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF
FLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK).
[0252] The sequence of any peptide of this disclosure is expressed
as a fusion protein with either murine or human Fc by adding a
secretion signal sequence to the N-terminus and an Fc sequence to
the C-terminus. This creates a bivalent molecule with improved
secretion properties. The larger peptide-Fc fusion is expressed in
different mammalian or insect cell lines and is useful as a
research reagent and a therapeutic.
[0253] Fc fusion to a peptide of SEQ ID NO: 108 to yield a sequence
of SEQ ID NO: 565 extends half-life and improves biodistribution of
the peptide to cartilage. Any peptide of this disclosure is
co-expressed with Fc protein to yield Fc-fusion peptides with
longer half-life and improved homing to cartilage. In SEQ ID NO:
565, the secretion signal sequence METDTLLLWVLLLWVPGSTG (SEQ ID NO:
566) is followed by the peptide of SEQ ID NO: 108, and is followed
by the sequence for Fc protein. Cleaving can be imprecise,
resulting in cleavage at position 20 or position 21 of SEQ ID NO:
565.
Example 20
Peptide Conjugate Hydrolysis
[0254] This example describes preparation of peptide conjugates
having tunable hydrolysis rates. The peptide-drug conjugates
described below are synthesized with the modification that instead
of using succinic anhydride, other molecules are used to provide
steric hindrance to hydrolysis or an altered local environment at
the carbon adjacent to the final hydrolyzable ester. In one
exemplary conjugate, the peptide-drug conjugate is synthesized with
tetramethyl succinic anhydride to generate hindered esters, which
causes a decreased rate of hydrolysis. In another exemplary
conjugate, one methyl group is present at the adjacent carbon. In
another exemplary conjugate, two methyl groups are present at the
adjacent carbon. In another exemplary conjugate, one ethyl group is
present at the adjacent carbon. In another exemplary conjugate, two
ethyl groups are present at the adjacent carbon. In another
exemplary conjugate, the carbon linker length is increased such as
by using glutaric anhydride instead of succinic anhydride,
increasing the local hydrophobicity and lowering the hydrolysis
rate. In another exemplary conjugate, a hydroxyl group is located
on the adjacent carbon, increasing the local hydrophilicity and
increasing the hydrolysis rate. The rate of hydrolysis in these
exemplary conjugates is therefore adjusted, preventing premature
cleavage and ensuring that the majority of peptide-dexamethasone
conjugates accumulate in cartilage prior to release of the drug by
hydrolysis but that the dexamethasone is also released in the
cartilage in a timely manner.
[0255] The resulting peptide conjugates are administered to a human
or animal subcutaneously, intravenously, orally, or injected
directly into a joint to treat disease.
Example 21
Peptide Conjugates with Stable Linkers
[0256] This example describes preparation of peptide conjugates
with stable linkers. A peptide of the disclosure is expressed
recombinantly or is chemically synthesized. The peptide is
conjugated to a detectable agent or an active agent via a stable
linker, such as an amide linkage or a carbamate linkage. The
peptide is conjugated to a detectable agent or an active agent via
a stable linker, such as an amide bond using standard
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) or
dicylcohexylcarbodiimide (DCC) based chemistry or thionyl chloride
or phosphorous chloride-based bioconjugation chemistries.
[0257] A peptide and drug conjugated via a linker are described
with the formula Peptide-A-B-C-Drug, wherein the linker is A-B-C. A
can be a stable amide link that is formed by reacting with an amine
on the peptide with a linker containing a tetrafluorophenyl (TFP)
ester or an NHS ester. A can also be a stable carbamate linker that
is formed by reacting with an amine on the peptide imidazole
carbamate active intermediate formed by the reaction of CDI with a
hydroxyl on the linker. A can also be a stable secondary amine
linkage that is formed by reductive alkylation of the amine on the
peptide with an aldehyde or ketone group on the linker. A can also
be a stable thioether linker formed using a maleimide or
bromoacetamide in the linker with a thiol in the peptide, a
triazole linker, a stable oxime linker, or an oxacarboline linker.
B is (--CH2-).sub.x-, a short PEG (--CH.sub.2CH.sub.2O--).sub.x (x
is 0-20). Alternatively, spacers within the linker is optional and
can be included or not at all. C is an amide bond formed with an
amine or a carboxylic acid on the drug, a thioether formed between
a maleimide on the linker and a sulfhydroyl on the drug, a
secondary or tertiary amine, a carbamate, or other stable bonds.
Any linker chemistry described in "Current ADC Linker Chemistry,"
Jain et al., Pharm Res, 2015 DOI 10. 1007/s11095-015-1657-7 can be
used.
[0258] The resulting peptide conjugates are administered to a human
or animal subcutaneously, intravenously, orally, or injected
directly into a joint to treat disease. The peptide is not
specifically cleaved from the detectable agent or active agent via
a targeted mechanism. The peptide can be degraded by mechanisms
such as catabolism, releasing a drug that is modified or not
modified form its native form (Singh, Luisi, and Pak, Pharm Res
32:3541-3571 (2015)). The peptide drug conjugate exerts its
pharmacological activity while still intact, or while partially or
fully degraded, metabolized, or catabolized.
Example 22
Peptide Conjugates with Cleavable Linkers
[0259] This example describes preparation of peptide conjugates
having cleavable linkers. A peptide of the disclosure is expressed
recombinantly or chemically synthesized. A peptide and drug are
conjugated via a linker and is described with the formula
Peptide-A-B-C-Drug, wherein the linker is A-B-C. A is a stable
amide link such as that formed by reacting an amine on the peptide
with a linker containing a tetrafluorophenyl (TFP) ester or an NHS
ester. A can also be a stable carbamate linker such as that formed
by reacting an amine on the peptide with an imidazole carbamate
active intermediate formed by reaction of CDI with a hydroxyl on
the linker. A can also be a stable secondary amine linkage such as
that formed by reductive alkylation of the amine on the peptide
with an aldehyde or ketone group on the linker. A can also be a
stable thioether linker formed using a maleimide or bromoacetamide
in the linker with a thiol in the peptide, a triazole linker, a
stable oxime linker, or a oxacarboline linker. B is (--CH2-).sub.x-
or a short PEG (--CH2CH2O--).sub.x (x is 0-20) or other spacers or
no spacer. C is an ester bond to the hydroxyl or carboxylic acid on
the drug, or a carbonate, hydrazone, or acylhydrazone, designed for
hydrolytic cleavage. The hydrolytic rate of cleavage is varied by
varying the local environment around the ester, including carbon
length (--CH2-).sub.x, steric hindrance (including adjacent side
groups such as methyl, ethyl, cyclic), hydrophilicity or
hydrophobicity. Hydrolysis rate is affected by local pH, such as
lower pH in certain compartments of the body or of the cell such as
endosomes and lysosomes or diseased tissues. C is a pH sensitive
group such as a hydrazone or oxime linkage. Alternatively C is a
disulfide bond designed to be released by reduction, such as by
glutathione. Alternatively C (or A-B-C) is a peptidic linkage
design for cleavable by enzymes. Optionally, a self-immolating
group such as pABC is included to cause release of a free
unmodified drug upon cleavage (Antibody-Drug Conjugates: Design,
Formulation, and Physicochemical Stability, Singh, Luisi, and Pak.
Pharm Res (2015) 32:3541-3571). The linker is cleaved by enzymes
such as esterases, matrix metalloproteinases, cathepsins such as
cathepsin B, glucuronidases, a protease, or thrombin.
Alternatively, the bond designed for cleavage is at A, rather than
C, and C could be a stable bond or a cleavable bond. An alternative
design is to have stable linkers (such as amide or carbamate) at A
and C and have a cleavable linker in B, such as a disulfide bond.
The rate of reduction is modulated by local effects such as steric
hindrance from methyl or ethyl groups or modulating
hydrophobicity/hydrophilicity.
[0260] The resulting peptide conjugates are administered to a human
or animal subcutaneously, intravenously, orally, or injected
directly into a joint to treat disease.
Example 23
Acetylsalicylic Acid Peptide Conjugate
[0261] This example describes the conjugation of acetylsalicylic
acid to a peptide using a lactic acid linker. A conjugate is
produced from a mixture of (R,S)-acetylsalicylic acid, lactic acid,
and a peptide:
[0262] The acetylsalicylic acid-lactic acid linker conjugate
depicted above is then reacted with a lysine or the N-terminus of a
cystine-dense peptide to create an acetylsalicylic acid-lactic
acid-peptide conjugate. The cystine-dense peptide is selected from
the peptides of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ
ID NO: 564.
[0263] Acetylsalicylic acid is currently dosed as an enantiomeric
mixture, in which enantiomers with a single racemic stereocenter
are very difficult to separate. As in the reaction scheme (I), a
diastereomer with two chiral centers is created by the addition of
a chiral linker such as L-lactic acid. Since diastereomers are
easily separated, the active enantiomer of acetylsalicylic acid
conjugated to the lactic acid linker can be purified prior to
conjugation to a cystine-dense peptide. The chemical synthesis can
use any conjugation techniques known in the art, such as described
in Bioconjugate Techniques by Greg Hermanson and in
"Ketorolac-dextran conjugates: synthesis, in vitro, and in vivo
evaluation:" Acta Pharm. 57 (2007) 441-450, Vyas, Trivedi, and
Chaturvedi. The conjugate can display anti-inflammatory activity,
or free acetylsalicylic acid is released from the conjugate to
provide anti-inflammatory activity. The free acetylsalicylic acid
can result from hydrolysis that occurs after administration, such
as hydrolysis at the ester bond. By dosing the conjugate containing
the cartilage homing peptide, a higher AUC of acetylsalicylic acid
delivery to the joint may be achieved than would be achieved by
systemic dosing of acetylsalicylic acid alone.
[0264] Such peptide-drug conjugates can be made using either a
cleavable or stable linker as described herein (e. g., EXAMPLES 21
and 22).
Example 24
Acetylsalicylic Acid Peptide Conjugate
[0265] This example describes the conjugation of acetylsalicylic
acid to a peptide using a PEG linker. A conjugate is produced using
acetylsalicylic acid and a PEG linker, which forms an ester bond
that can hydrolyze as described in "In vitro and in vivo study of
poly(ethylene glycol) conjugated ibuprofen to extend the duration
of action," Scientia Pharmaceutica, 2011, 79:359-373, Nayak and
Jain. Fischer esterification is used to conjugate ibuprofen with a
short PEG, e.g., with triethylene glycol, to yield
ibuprofen-ester-PEG-OH.
[0266] Following preparation of the PEG-ibuprofen conjugate as
shown above, the hydroxyl moiety of PEG is activated with
N,N'-disuccinimidyl carbonate (DSC) to form
ibuprofen-ester-PEG-succinimidyl carbonate, which is then reacted
with a lysine or the N-terminus of a cystine-dense peptide to form
an ibuprofen-ester-PEG-peptide conjugate. The cystine-dense peptide
is selected from any one of the peptides of sequence SEQ ID NO:
24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. The conjugate
can display anti-inflammatory activity, or free ibuprofen is
released from the conjugate to provide anti-inflammatory activity.
The free ibuprofen can result from hydrolysis that occurs after
administration, such as hydrolysis at the ester bond.
[0267] Ibuprofen-peptide conjugates are administered to a subject
in need thereof. The subject can be a human or a non-human
animal.
[0268] Such peptide-drug conjugates can be made using either a
cleavable or stable linker as described herein (e.g., EXAMPLES 21
and 22).
Example 25
Dexamethasone Peptide Conjugate
[0269] This example describes different methods of conjugating
dexamethasone with a peptide of this disclosure. A peptide of SEQ
ID NO: 108 was recombinantly expressed. Dexamethasone was readily
conjugated to a peptide of this disclosure using a dicarboxylic
acid linker. The peptide-dexamethasone conjugate was made by first
converting dexamethasone to a hemisuccinate by reacting it with
succinic anhydride. The hemisuccinate was then converted to a
succinate carboxylic acid containing an active ester, using
dicyclohexyl carbodiimide (DCC) or
1-ethyl-3-(3-dimethylamninopropyl)carbodiimide (EDC) in the
presence of N-hydroxy succinimide (NHS). This active ester was then
reacted with a lysine or the N-terminus of a cystine-dense peptide
to create a dexamethasone-carboxylic acid-peptide conjugate.
Methods such as those described in "Functionalized derivatives of
hyaluronic acid oligosaccharides: drug carriers and novel
biomaterials" Bioconjugate Chemistry 1994, 5, 339-347, Pouyani and
Prestwich, and Bioconjugate Techniques by Greg Hermanson can be
used (Elsevier Inc., 3.sup.rd Edition, 2013).
[0270] Peptide-dexamethasone conjugates were prepared by coupling
dexamethasone to the peptides of this disclosure using standard
coupling-reagent chemistry. For example, dexamethasone conjugates
were made by reacting dexamethasone hemigluterate with 1.05 molar
equivalents of 1,1'-carbonyldiimidazole in anhydrous DMSO in an
inert atmosphere. After 30 minutes, excess dexamethasone in
anhydrous DMSO was added along with two molar equivalents of
anhydrous trimethylamine. The N-hydroxysuccinimide ester of the
peptide-dexamethasone conjugate was generated to form a
shelf-stable intermediate for later reaction with an
amine-containing carrier. The N-terminal dexamethasone-peptide
conjugate (SEQ ID NO: 108B) was verified by electrospray mass
spectrometry (ES-MS) within a 10 ppm error.
[0271] A peptide of any of the sequences of this disclosure
including SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO:
564, are conjugated to dexamethasone using the methods described
above.
[0272] Such peptide-drug conjugates can be made using either a
cleavable or stable linker as described herein (e.g., EXAMPLES 21
and 22).
Example 26
Beclomethasone Monopropionate Peptide Conjugates
[0273] This example describes conjugation of a peptide of SEQ ID
NO: 27 or SEQ ID NO: 108 of this disclosure to beclomethasone
monopropionate. Beclomethasone monopropionate is readily conjugated
to any peptide disclosed herein via a dicarboxylic acid linker. The
dicarboxylic acid linker is a linear dicarboxylic acid, such as
succinic acid, or a related cyclic anhydride, such as succinic
anhydride. Reactions with anhydrides can proceed under simple
conditions. For example, the reaction of beclomethasone
monopropionate with five molar equivalents of glutaric anhydride is
carried out in anhydrous pyridine at room temperature. Reactions
with dicarboxylic acids can occur using standard carbodiimide
coupling methods. For example, beclomethasone monopropionate is
reacted with one molar equivalent dimethylsuccinic acid, one molar
equivalent 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (or
another carbodiimide), and 0.2 molar equivalents of
40-dimethylamino pyridine.
[0274] The same methods as described in EXAMPLE 20 are used to
adjust the rate of hydrolysis of peptide-beclomethasone
monopropionate conjugates, preventing premature cleavage and
ensuring that the beclomethasone monopropionate of
peptide-beclomethasone monopropionate conjugates accumulate in
cartilage.
[0275] Peptide-beclomethasone monopropionate conjugates are
prepared by coupling beclomethasone monopropionate to the peptides
of this disclosure using standard coupling-reagent chemistry. The
peptide-beclomethasone monopropionate conjugate was made by first
converting beclomethasone monopropionate to a hemisuccinate by
reacting it with succinic anhydride. The hemisuccinate was then
converted to a succinate carboxylic acid containing an active
ester, using dicyclohexyl carbodiimide (DCC) or
1-ethyl-3-(3-dimethylamninopropyl)carbodiimide (EDC) in the
presence of N-hydroxy succinimide (NHS). This active ester was then
reacted with a lysine or the N-terminus of a peptide to create a
beclomethasone monopropionate-carboxylic acid-peptide conjugate.
Methods such as those described in "Functionalized derivatives of
hyaluronic acid oligosaccharides: drug carriers and novel
biomaterials" Bioconjugate Chemistry 1994, 5, 339-347, Pouyani and
Prestwich, and Bioconjugate Techniques by Greg Hermanson can be
used (Elsevier Inc., 3.sup.rd Edition, 2013).
[0276] Peptide-beclomethasone monopropionate conjugates were
prepared by coupling beclomethasone monopropionate to the peptides
of this disclosure using standard coupling-reagent chemistry. For
example, beclomethasone monopropionate conjugates were made by
reacting beclomethasone monopropionate hemigluterate with 1.05
molar equivalents of 1,1'-carbonyldiimidazole in anhydrous DMSO in
an inert atmosphere. After 30 minutes, excess beclomethasone
monopropionate in anhydrous DMSO was added along with two molar
equivalents of anhydrous trimethylamine. The N-hydroxysuccinimide
ester of the peptide-beclomethasone monopropionate conjugate was
generated to form a shelf-stable intermediate for later reaction
with an amine-containing carrier.
[0277] Beclomethasone monopropionate is also readily conjugated to
any peptide disclosed herein via a dicarboxylic acid linker. The
dicarboxylic acid linker is a linear dicarboxylic acid, such as
succinic acid, or a related cyclic anhydride, such as succinic
anhydride. Reactions with anhydrides can proceed under simple
conditions. For example, the reaction of beclomethasone
monopropionate with five molar equivalents of glutaric anhydride is
carried out in anhydrous pyridine at room temperature. Reactions
with dicarboxylic acids can occur using standard carbodiimide
coupling methods. For example, beclomethasone monopropionate is
reacted with one molar equivalent dimethylsuccinic acid, one molar
equivalent 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (or
another carbodiimide), and 0.2 molar equivalents of
40-dimethylamino pyridine. The peptide-beclomethasone
monopropionate conjugates are administered to a subject in need
thereof and home, target, are directed to, are retained by,
accumulate in, migrate to, and/or bind to cartilage and/or kidneys.
The subject is a human or animal and has inflammation in the
cartilage or kidney tissues. Upon administration of the
peptide-beclomethasone monopropionate conjugates, the cartilage
and/or kidney inflammation is alleviated.
[0278] The peptide can also be a peptide of SEQ ID: NO: 33. The
peptide can be any peptide with the sequence selected SEQ ID NO:
24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564.
[0279] Such peptide-drug conjugates are made using either a
cleavable or stable linker as described herein (e.g., EXAMPLES 21
and 22).
Example 27
Desciclesonide Peptide Conjugates
[0280] This example describes conjugation of a peptide of SEQ ID
NO: 199 or SEQ ID NO: 187 of this disclosure to desciclesonide.
Ciclesonide is a prodrug that is metabolized in vivo to the active
metabolite desciclesonide. By conjugating desciclesonide to a
peptide via an ester linker, upon hydrolysis the released drug
would be desciclesonide, just as after systemic administration of
ciclesonide the active metabolite desciclesonide is present and
active. Desciclesonide is readily conjugated to any peptide
disclosed herein via a dicarboxylic acid linker. The dicarboxylic
acid linker is a linear dicarboxylic acid, such as succinic acid,
or a related cyclic anhydride, such as succinic anhydride.
Reactions with anhydrides can proceed under simple conditions. For
example, the reaction of desciclesonide with five molar equivalents
of glutaric anhydride is carried out in anhydrous pyridine at room
temperature. Reactions with dicarboxylic acids can occur using
standard carbodiimide coupling methods. For example, desciclesonide
is reacted with one molar equivalent dimethylsuccinic acid, one
molar equivalent 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (or
another carbodiimide), and 0.2 molar equivalents of
40-dimethylamino pyridine.
[0281] The same methods as described in EXAMPLE 20 are used to
adjust the rate of hydrolysis of peptide-desciclesonide conjugates,
preventing premature cleavage and ensuring that the desciclesonide
of peptide-desciclesonide conjugates accumulate in cartilage.
[0282] Desciclesonide is also readily conjugated to any peptide
disclosed herein via a dicarboxylic acid linker. The dicarboxylic
acid linker is a linear dicarboxylic acid, such as succinic acid,
or a related cyclic anhydride, such as succinic anhydride.
Reactions with anhydrides can proceed under simple conditions. For
example, the reaction of desciclesonide with five molar equivalents
of glutaric anhydride is carried out in anhydrous pyridine at room
temperature. Reactions with dicarboxylic acids can occur using
standard carbodiimide coupling methods. For example, desciclesonide
is reacted with one molar equivalent dimethylsuccinic acid, one
molar equivalent 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (or
another carbodiimide), and 0.2 molar equivalents of
40-dimethylamino pyridine. The peptide-desciclesonide conjugates
are administered to a subject in need thereof and home, target, are
directed to, are retained by, accumulate in, migrate to, and/or
bind to cartilage and/or kidneys. The subject is a human or animal
and has inflammation in the cartilage or kidney tissues. Upon
administration of the peptide-desciclesonide conjugates, the
cartilage and/or kidney inflammation is alleviated.
[0283] The peptide-desciclesonide conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage
and/or kidneys. The subject is a human or animal and has
inflammation in the cartilage or kidney tissues. Upon
administration of the peptide-desciclesonide conjugates, the
cartilage and/or kidney inflammation is alleviated.
[0284] The peptide can also be a peptide of SEQ ID: NO: 196. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564.
[0285] Such peptide-drug conjugates are made using either a
cleavable or stable linker as described herein (e.g., EXAMPLES 21
and 22).
Example 28
Desciclesonide Peptide Conjugates
[0286] This example describes conjugation of a peptide of SEQ ID
NO: 199 or SEQ ID NO: 187 of this disclosure to desciclesonide.
Ciclesonide is a prodrug that is metabolized in vivo to the active
metabolite desciclesonide. By conjugating desciclesonide to a
peptide via an ester linker, upon hydrolysis the released drug
would be desciclesonide, just as after systemic administration of
ciclesonide the active metabolite desciclesonide is present and
active. Desciclesonide is readily conjugated to any peptide
disclosed herein via a stable linker.
[0287] The peptide-desciclesonide conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage
and/or kidneys. The subject is a human or animal and has
inflammation in the cartilage or kidney tissues. Upon
administration of the peptide-desciclesonide conjugates, the
cartilage and/or kidney inflammation is alleviated.
[0288] The peptide can also be a peptide of SEQ ID: NO: 196. The
peptide can be any peptide with the sequence selected SEQ ID NO:
24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564.
[0289] Such peptide-drug conjugates are made using either a
cleavable or stable linker as described herein (e.g., EXAMPLES 21
and 22).
Example 29
Peptide-Ustekinumab Conjugates
[0290] This example describes conjugation of a peptide of SEQ ID
NO: 106 this disclosure to ustekinumab. Ustekinumab is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). Alternatively the peptide-active agent of this Example can
be expressed as a fusion protein. From one to eight peptides are
linked to ustekinumab.
[0291] The peptide-ustekinumab conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has psoriatic arthritis. Upon
administration of the peptide-ustekinumab conjugates, the psoriatic
arthritis condition is alleviated.
[0292] The peptide can also be a peptide of SEQ ID NO: 36. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564.
Example 30
Peptide-Xeljanz Conjugates
[0293] This example describes conjugation of a peptide of SEQ ID
NO: 187 this disclosure to xeljanz. Xeljanz is readily conjugated
to any peptide disclosed herein via standard chemistries such as
those described in, but not limited to, Bioconjugate Techniques by
Greg Hermanson (Elsevier Inc., 3.sup.rd edition, 2013). From one to
eight peptides are linked to xeljanz.
[0294] The peptide-xeljanz conjugates are administered to a subject
in need thereof and home, target, are directed to, are retained by,
accumulate in, migrate to, and/or bind to cartilage. The subject is
a human or animal and has rheumatoid arthritis. Upon administration
and homing of peptide-xeljanz conjugates, the rheumatoid arthritis
condition is alleviated.
[0295] The peptide can also be a peptide of SEQ ID NO: 185. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 31
Peptide-IL-17 Inhibitor Conjugates
[0296] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to an IL-17 inhibitor. An IL-17 inhibitor
is readily conjugated to any peptide disclosed herein via standard
chemistries such as those described in, but not limited to,
Bioconjugate Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd
edition, 2013).
[0297] The peptide-IL-17 inhibitor conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has ankylosing spondylitis.
Upon administration and homing of peptide-IL-17 inhibitor
conjugates, the ankylosing spondylitis condition is alleviated.
[0298] The peptide can also be a peptide of SEQ ID NO: 111. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 32
Peptide-Iguratimod Conjugates
[0299] This example describes conjugation of a peptide of SEQ ID
NO: 199 this disclosure to iguratimod. Iguratimod is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013).
[0300] The peptide-iguratimod conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has rheumatoid arthritis. Upon
administration and homing of peptide-iguratimod conjugates, the
rheumatoid arthritis condition is alleviated.
[0301] The peptide can also be a peptide of SEQ ID NO: 26. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 33
Peptide Mycophenolic Acid Conjugates
[0302] This example describes conjugation of a peptide of SEQ ID
NO: 27 this disclosure to mycophenolic acid. Mycophenolic acid is
readily conjugated to any peptide disclosed herein via standard
chemistries such as those described in, but not limited to,
Bioconjugate Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd
edition, 2013).
[0303] The peptide-mycophenolic acid conjugates are administered to
a subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has organ transplantation,
infection, cancer, or other kidney disorders. Upon administration
and homing of peptide-mycophenolic acid conjugates, the organ
transplantation, infection, cancer, other kidney disorders
condition is alleviated.
[0304] The peptide can also be a peptide of SEQ ID NO: 107. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 34
Peptide-Tacrolimus Conjugates
[0305] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to tacrolimus. Tacrolimus is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013).
[0306] The peptide-tacrolimus conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has organ transplantation, any
other kidney disease. Upon administration and homing of
peptide-tacrolimus conjugates, the organ transplantation, any other
kidney disease condition is alleviated.
[0307] The peptide can also be a peptide of SEQ ID NO: 111. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 35
Peptide-Secukinumab Conjugates
[0308] This example describes conjugation of a peptide of SEQ ID
NO: 106 this disclosure to secukinumab. Secukinumab is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to secukinumab.
Alternatively the peptide-active agent of this Example can be
expressed as a fusion protein.
[0309] The peptide-secukinumab conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has ankylosing spondylitis.
Upon administration and homing of peptide-secukinumab acid
conjugates, the ankylosing spondylitis condition is alleviated.
[0310] The peptide can also be a peptide of SEQ ID NO: 24. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 36
Peptide-Sirukumab Conjugates
[0311] This example describes conjugation of a peptide of SEQ ID
NO: 199 this disclosure to sirukumab. Sirukumab is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to sirukumab.
Alternatively the peptide-active agent of this Example can be
expressed as a fusion protein.
[0312] The peptide-sirukumab conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has rheumatoid arthritis, immune
diseases of the kidneys. Upon administration and homing of
peptide-sirukumab conjugates, the rheumatoid arthritis, immune
diseases of the kidneys condition is alleviated.
[0313] The peptide can also be a peptide of SEQ ID NO: 36. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 37
Peptide-Anifrolumab Conjugates
[0314] This example describes conjugation of a peptide of SEQ ID
NO: 27 this disclosure to anifrolumab. Anifrolumab is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to anifrolumab.
Alternatively the peptide-active agent of this Example can be
expressed as a fusion protein.
[0315] The peptide-anifrolumab conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has lupus nephritis. Upon
administration and homing of peptide-anifrolumab conjugates, the
lupus nephritis condition is alleviated.
[0316] The peptide can also be a peptide of SEQ ID NO: 185. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 38
Peptide-Denosumab Conjugates
[0317] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to denosumab. Denosumab is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to denosumab.
Alternatively the peptide-active agent of this Example can be
expressed as a fusion protein.
[0318] The peptide-denosumab conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has osteoporosis. Upon
administration and homing of peptide-denosumab conjugates, the
osteoporosis condition is alleviated.
[0319] The peptide can also be a peptide of SEQ ID NO: 25. The
peptide can be any peptide with the sequence selected SEQ ID NO:
24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 39
Peptide-Rituximab Conjugates
[0320] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to rituximab. Rituximab is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to rituximab.
Alternatively the peptide-active agent of this Example can be
expressed as a fusion protein.
[0321] The peptide-rituximab conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage or
kidneys. The subject is a human or animal and has rheumatoid
arthritis, kidney transplant. Upon administration and homing of
peptide-rituximab conjugates, the rheumatoid arthritis, kidney
transplant condition is alleviated.
[0322] The peptide can also be a peptide of SEQ ID NO: 26. The
peptide can be any peptide with the sequence selected SEQ ID NO:
24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 40
Peptide-Omalizumab Conjugates
[0323] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to omalizumab. Omalizumab is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to omalizumab.
Alternatively the peptide-active agent of this Example can be
expressed as a fusion protein.
[0324] The peptide-omalizumab conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has kidney inflammation. Upon
administration and homing of peptide-omalizumab conjugates, the
kidney inflammation condition is alleviated.
[0325] The peptide can also be a peptide of SEQ ID NO: 107. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 41
Peptide-Abatacept Conjugates
[0326] This example describes conjugation of a peptide of SEQ ID
NO: 199 this disclosure to abatacept. Abatacept is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to abatacept.
Alternatively the peptide-active agent of this Example can be
expressed as a fusion protein.
[0327] The peptide-abatacept conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has rheumatoid arthritis, lupus
nephritis, organ transplant, focal segmental glomerulosclerosis.
Upon administration and homing of peptide-abatacept conjugates, the
rheumatoid arthritis, lupus nephritis, organ transplant, focal
segmental glomerulosclerosis condition is alleviated.
[0328] The peptide can also be a peptide of SEQ ID NO: 111. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 42
Peptide-Oxycodone Conjugates
[0329] This example describes conjugation of a peptide of SEQ ID
NO: 187 this disclosure to oxycodone. Oxycodone is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013).
[0330] The peptide-oxycodone conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has cartilage or
kidney-related pain. Upon administration and homing of
peptide-oxycodone conjugates, the cartilage-related pain condition
is alleviated.
[0331] The peptide can also be a peptide of SEQ ID NO: 24. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 43
Peptide Capsaicin Conjugates
[0332] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to capsaicin. Capsaicin is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013).
[0333] The peptide-capsaicin conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has cartilage-related pain.
Upon administration and homing of peptide-capsaicin conjugates, the
cartilage or kidney-related pain condition is alleviated.
[0334] The peptide can also be a peptide of SEQ ID NO: 36. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 44
Peptide-GSK2193874 Conjugates
[0335] This example describes conjugation of a peptide of SEQ ID
NO: 106 this disclosure to GSK2193874. GSK2193874 is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013).
[0336] The peptide-GSK2193874 conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to GSK2193874.
The subject is a human or animal and has cartilage-related pain.
Upon administration and homing of peptide-GSK2193874 conjugates,
the cartilage-related pain condition is alleviated.
[0337] The peptide can also be a peptide of SEQ ID NO: 185. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 45
Peptide BIIB023 Conjugates
[0338] This example describes conjugation of a peptide of SEQ ID
NO: 199 this disclosure to BIIB023. BIIB023 is readily conjugated
to any peptide disclosed herein via standard chemistries such as
those described in, but not limited to, Bioconjugate Techniques by
Greg Hermanson (Elsevier Inc., 3.sup.rd edition, 2013). From one to
eight peptides are linked to BIIB023. Alternatively the
peptide-active agent of this Example can be expressed as a fusion
protein.
[0339] The peptide-BIIB023 conjugates are administered to a subject
in need thereof and home, target, are directed to, are retained by,
accumulate in, migrate to, and/or bind to cartilage. The subject is
a human or animal and has lupus nephritis or rheumatoid arthritis.
Upon administration and homing of peptide-BIIB023 conjugates, the
lupus nephritis or rheumatoid arthritis condition is
alleviated.
[0340] The peptide can also be a peptide of SEQ ID NO: 25. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 46
Peptide-Anakinra Conjugates
[0341] This example describes conjugation or fusion of a peptide of
SEQ ID NO: 187 or SEQ ID NO: 550-564 of this disclosure to
anakinra. A linker is optionally used to conjugate the peptide to
anakinra. Anakinra is readily conjugated to any peptide disclosed
herein via standard chemistries such as those described in, but not
limited to, Bioconjugate Techniques by Greg Hermanson (Elsevier
Inc., 3.sup.rd edition, 2013). From one to eight peptides are
linked to anakinra. Alternatively the peptide-active agent of this
Example can be expressed as a fusion protein.
[0342] The peptide-anakinra conjugates or fusions are administered
to a subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has lupus nephritis or
rheumatoid arthritis. Upon administration and homing of
peptide-anakinra conjugates or fusions, the lupus nephritis or
rheumatoid arthritis condition is alleviated.
[0343] The peptide can also be a peptide of SEQ ID NO: 26. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 47
Peptide-IGF-1 Conjugates
[0344] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to IGF-1. IGF-1 is readily conjugated to
any peptide disclosed herein via standard chemistries such as those
described in, but not limited to, Bioconjugate Techniques by Greg
Hermanson (Elsevier Inc., 3.sup.rd edition, 2013). From one to
eight peptides are linked to IGF-1. Alternatively the
peptide-active agent (where the active agent is the biologic of
this Example) can be expressed as a fusion protein.
[0345] The peptide-IGF-1 conjugates are administered to a subject
in need thereof and home, target, are directed to, are retained by,
accumulate in, migrate to, and/or bind to cartilage. The subject is
a human or animal and has renal cancer or arthritis. Upon
administration and homing of peptide-IGF-1 conjugates, the renal
cancer or arthritis condition is alleviated.
[0346] The peptide can also be a peptide of SEQ ID NO: 107. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 48
Peptide-Romosozumab Conjugates
[0347] This example describes conjugation of a peptide of SEQ ID
NO: 106 this disclosure to Romosozumab. Romosozumab is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to romosozumab.
Alternatively the peptide-active agent of this Example can be
expressed as a fusion protein.
[0348] The peptide-romosozumab conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has osteoporosis. Upon
administration and homing of peptide-romosozumab conjugates, the
osteoporosis condition is alleviated.
[0349] The peptide can also be a peptide of SEQ ID NO: 111. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 49
Peptide-ZVAD-fmk Conjugates
[0350] This example describes conjugation of a peptide of SEQ ID
NO: 187 this disclosure to ZVAD-fmk. ZVAD-fmk is readily conjugated
to any peptide disclosed herein via standard chemistries such as
those described in, but not limited to, Bioconjugate Techniques by
Greg Hermanson (Elsevier Inc., 3.sup.rd edition, 2013). From one to
eight peptides are linked to ZVAD-fmk. The peptide-ZVAD-fmk
conjugates are administered to a subject in need thereof and home,
target, are directed to, are retained by, accumulate in, migrate
to, and/or bind to cartilage. The subject is a human or animal and
has cartilage grafting, arthritis, surgical intervention, surgery
for cartilage repair. Upon administration and homing of
peptide-ZVAD-fmk conjugates, the cartilage grafting, arthritis,
surgical intervention, surgery for cartilage repair condition is
alleviated.
[0351] The peptide can also be a peptide of SEQ ID NO: 24. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 50
Peptide-S-Methylisothiourea Conjugates
[0352] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to S-methylisothiourea. S-methylisothiourea
is readily conjugated to any peptide disclosed herein via standard
chemistries such as those described in, but not limited to,
Bioconjugate Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd
edition, 2013).
[0353] The peptide-S-methylisothiourea conjugates are administered
to a subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has arthritis surgery, kidney
iron overload, renal ischemia reperfusion injury, or acute kidney
injury. Upon administration and homing of
peptide-S-methylisothiourea conjugates, the arthritis surgery,
kidney iron overload, renal ischemia reperfusion injury, or acute
kidney injury condition is alleviated.
[0354] The peptide can also be a peptide of SEQ ID NO: 33. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 51
Peptide-P188 Conjugates
[0355] This example describes conjugation of a peptide of SEQ ID
NO: 106 this disclosure to P188. P188 is readily conjugated to any
peptide disclosed herein via standard chemistries such as those
described in, but not limited to, Bioconjugate Techniques by Greg
Hermanson (Elsevier Inc., 3.sup.rd edition, 2013).
[0356] The peptide-P188 conjugates are administered to a subject in
need thereof and home, target, are directed to, are retained by,
accumulate in, migrate to, and/or bind to cartilage. The subject is
a human or animal and has arthritis surgery. Upon administration
and homing of peptide-P188 conjugates, the arthritis surgery
condition is alleviated.
[0357] The peptide can also be a peptide of SEQ ID NO: 185. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 52
Peptide-Alendronate Conjugates
[0358] This example describes conjugation of a peptide of SEQ ID
NO: 187 this disclosure to alendronate. Alendronate is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013).
[0359] The peptide-alendronate conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has bone erosion. Upon
administration and homing of peptide-alendronate conjugates, the
bone erosion condition is alleviated.
[0360] The peptide can also be a peptide of SEQ ID NO: 22. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 53
Peptide-MIP-3a Conjugates
[0361] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to MIP-3a. MIP-3a is readily conjugated to
any peptide disclosed herein via standard chemistries such as those
described in, but not limited to, Bioconjugate Techniques by Greg
Hermanson (Elsevier Inc., 3.sup.rd edition, 2013). From one to
eight peptides are linked to MIP-3a. Alternatively the
peptide-active agent of this Example can be expressed as a fusion
protein
[0362] The peptide-MIP-3a conjugates are administered to a subject
in need thereof and home, target, are directed to, are retained by,
accumulate in, migrate to, and/or bind to cartilage. The subject is
a human or animal and has Joint injury, repair and regeneration of
cartilage and bone. Upon administration and homing of
peptide-MIP-3a conjugates, the Joint injury, repair and
regeneration of cartilage and bone condition is alleviated.
[0363] The peptide can also be a peptide of SEQ ID NO: 26. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 54
Peptide-BMP-2 Conjugates
[0364] This example describes conjugation of a peptide of SEQ ID
NO: 106 this disclosure to BMP-2. BMP-2 is readily conjugated to
any peptide disclosed herein via standard chemistries such as those
described in, but not limited to, Bioconjugate Techniques by Greg
Hermanson (Elsevier Inc., 3.sup.rd edition, 2013). From one to
eight peptides are linked to BMP-2. Alternatively the
peptide-active agent of this Example can be expressed as a fusion
protein.
[0365] The peptide-BMP-2 conjugates are administered to a subject
in need thereof and home, target, are directed to, are retained by,
accumulate in, migrate to, and/or bind to cartilage. The subject is
a human or animal and has Joint repair. Upon administration and
homing of peptide-BMP-2 conjugates, the Joint repair condition is
alleviated.
[0366] The peptide can also be a peptide of SEQ ID NO: 107. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 55
Peptide-Icariin Conjugates
[0367] This example describes conjugation of a peptide of SEQ ID
NO: 187 this disclosure to icariin. Icariin is readily conjugated
to any peptide disclosed herein via standard chemistries such as
those described in, but not limited to, Bioconjugate Techniques by
Greg Hermanson (Elsevier Inc., 3.sup.rd edition, 2013).
[0368] The peptide-icariin conjugates are administered to a subject
in need thereof and home, target, are directed to, are retained by,
accumulate in, migrate to, and/or bind to cartilage. The subject is
a human or animal and has Joint repair. Upon administration and
homing of peptide-icariin conjugates, the Joint repair condition is
alleviated.
[0369] The peptide can also be a peptide of SEQ ID NO: 108. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or a
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 56
Peptide-Captopril Conjugates
[0370] This example describes conjugation of a peptide of SEQ ID
NO: 27 this disclosure to captopril. Captopril is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013).
[0371] The peptide-captopril conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has diabetic nephropathy. Upon
administration and homing of peptide-captopril conjugates, the
diabetic nephropathy condition is alleviated.
[0372] The peptide can also be a peptide of SEQ ID NO: 24. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 57
Peptide-Tofacitinib Conjugates
[0373] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to tofacitinib. Tofacitinib is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013). From one to eight peptides are linked to tofacitinib.
[0374] The peptide-tofacitinib conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage.
The subject is a human or animal and has rheumatoid arthritis and
kidney transplant, ankyloses spondylitis. Upon administration and
homing of peptide-tofacitinib conjugates, the rheumatoid arthritis
and kidney transplant, ankyloses spondylitis condition is
alleviated.
[0375] The peptide can also be a peptide of SEQ ID NO: 36. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 58
Peptide-Dimethyl Fumarate Conjugates
[0376] This example describes conjugation of a peptide of SEQ ID
NO: 108 this disclosure to dimethyl fumarate. Dimethyl fumarate is
readily conjugated to any peptide disclosed herein via standard
chemistries such as those described in, but not limited to,
Bioconjugate Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd
edition, 2013). Alternatively, peptide-dimethyl fumarate conjugates
can be synthesized by Michael addition of a thiol (on the peptide
of linker) to dimethyl fumarate as described by Schmidt et al.
(Bioorg Med Chem. 2007 Jan. 1; 15(1):333-42. Epub 2006 Sep.
29).
[0377] The peptide-dimethyl fumarate conjugates are administered to
a subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to kidneys. The
subject is a human or animal and has Kidney fibrosis, psoriatic
arthritis, rheumatoid arthritis. Upon administration and homing of
peptide-dimethyl fumarate conjugates, the Kidney fibrosis,
psoriatic arthritis, rheumatoid arthritis condition is
alleviated.
[0378] The peptide can also be a peptide of SEQ ID NO: 187. The
peptide can be any peptide with the sequence selected from SEQ ID
NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 59
Intra-Articular Administration of Peptides and Peptide
Conjugates
[0379] This example illustrates intra-articular administration of
peptides or peptide conjugates of this disclosure. A peptide of
this disclosure is expressed recombinantly or chemically
synthesized. In some cases, the peptide is subsequently conjugated
to a detectable agent or an active agent. The peptide or peptide
conjugate is administered to a subject in need thereof via
intra-articular administration. The cartilage is penetrated by the
peptide or peptide conjugate due to the small size of the peptide
or peptide conjugate, and due to binding of cartilage components by
the peptide or peptide conjugate. The peptide or peptide conjugate
is bound to cartilage and the residence time in the cartilage is
longer due to this binding. Optionally, the injected material is
aggregated, is crystallized, or complexes are formed, further
extending the depot effect and contributing to longer residence
time.
[0380] The peptide can be a peptide of SEQ ID NO: 108. The peptide
can also be a peptide of SEQ ID NO: 24. The peptide can be any
peptide with the sequence selected SEQ ID NO: 24-SEQ ID NO: 274 or
SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates can be
made using either a cleavable or stable linker as described herein
(e.g., EXAMPLES 21 and 22).
Example 60
Treatment of Osteoarthritis
[0381] This example describes a method for treating osteoarthritis
using peptides of the present disclosure. This method is used as a
treatment for acute and/or chronic symptoms associated with
osteoarthritis. A peptide of the present disclosure is expressed
recombinantly or chemically synthesized and then is used directly
or conjugated to an anti-inflammatory compound, such as aspirin,
desciclesonide, or secukinumab. The resulting peptide or
peptide-drug conjugate is administered in a pharmaceutical
composition subcutaneously, intravenously, or orally, or is
injected directly into a joint of a patient and targeted to
cartilage. The formulation can be modified physically or chemically
to increase the time of exposure in the cartilage. One or more
anti-inflammatory peptide conjugates are administered to a human or
animal.
[0382] The peptide can be a peptide of SEQ ID NO: 106. The peptide
can also be a peptide of SEQ ID NO: 33. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 61
Treatment of Cartilage Degradation
[0383] This example describes a method for treating and/or
preventing cartilage degradation using a peptide of the present
disclosure. This method is used as a treatment for acute and/or
chronic symptoms associated with cartilage degradation. Progressive
degradation or thinning of the cartilage is difficult to treat in
part because molecules such as small molecule drugs and antibodies
typically do not reach the avascular cartilage. A peptide of the
present disclosure is used for its homing and/or native activity,
or is mutated to generate activity such as MMP protease inhibition.
It is expressed recombinantly or chemically synthesized and then is
used directly or conjugated to an extracellular matrix targeting
active agent, such as an inhibitor of MMP activity or an
anti-apoptosis agent (e.g., osteoprotegrin, romosozumab, P188,
ZVAD-fmk, quercetin, dasatinib, dimethyl fumarate, bortezomib,
carilzomib, or navitoclax). The resulting peptide or peptide-drug
conjugate is administered in a pharmaceutical composition
subcutaneously, intravenously, or orally, or is injected directly
into a joint of a patient and targeted to extracellular matrix. One
or more extracellular matrix targeting conjugates are administered
to a human or animal.
[0384] The peptide can be a peptide of SEQ ID NO: 187. The peptide
can also be a peptide of SEQ ID NO: 27. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 62
Treatment of a Cartilage Injury
[0385] This example describes a method for treating a cartilage
injury using a peptide of the present disclosure. A peptide of the
present disclosure is expressed recombinantly or chemically
synthesized and then is used directly or conjugated to a
therapeutic compound, such as those described herein, including,
but not limited to BMP-2, BMP-7, BMP-9, BMP-13, PDGF, PTH, PTHrP,
IL-8, MIP-3a. The resulting peptide or peptide-drug conjugate is
administered in a pharmaceutical composition to a patient and
targeted to cartilage. One or more therapeutic compound-peptide
conjugates are administered to a human or animal.
[0386] The peptide can be a peptide of SEQ ID NO: 108. The peptide
can also be a peptide of SEQ ID NO: 185. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 63
Treatment of Rheumatoid Arthritis
[0387] This example describes a method for treating rheumatoid
arthritis. This method is used as a treatment for acute and/or
chronic symptoms associated with rheumatoid arthritis. A peptide of
the present disclosure is expressed recombinantly or chemically
synthesized and then is used directly, or is conjugated to an
anti-inflammatory compound, such as adalimumab, certolizumab,
golimumab, thalidomide, lenalidomide, pomalidomide, pentocifylline,
bupropion. When the peptide is used directly, the peptide can, for
example, bind or inhibit ion channels such as Kv 1. 3. The
resulting peptide or peptide-drug conjugate is administered in a
pharmaceutical composition to a patient and is targeted to
cartilage. One or more anti-inflammatory compound-peptide
conjugates are administered to a human or animal subcutaneously,
intravenously, or orally, or is injected directly into a joint
[0388] The peptide can be a peptide of SEQ ID NO: 106. The peptide
can also be a peptide of SEQ ID NO: 25. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 64
Treatment of Gout
[0389] This example describes a method for treating gout using
peptides of the present disclosure. This method is used as a
treatment for acute and/or chronic symptoms associated with gout. A
peptide of the present disclosure is expressed and administered in
a pharmaceutical composition to a patient as a therapeutic for
gout. A peptide of the disclosure is recombinantly or chemically
synthesized and then is used directly or conjugated to pegloticase
to treat a cartilage disorder. A peptide of the disclosure is
recombinantly or chemically synthesized and then is used directly
or conjugated to probenecid to treat a kidney disorder. The peptide
is administered in a pharmaceutical composition to a patient and
the peptide is targeted to the cartilage or kidney affected by
gout. One or more peptides are administered to a human or animal
subcutaneously, intravenously, or orally, or is injected directly
into a joint.
[0390] The peptide can be a peptide of SEQ ID NO: 187. The peptide
can also be a peptide of SEQ ID NO: 24. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 65
Treatment or Management of Pain
[0391] This example describes a method for treating or managing
pain associated with a cartilage injury or disorder. This method is
used as a treatment for acute and/or chronic symptoms associated
with a cartilage injury or disorder. A peptide of the disclosure is
expressed and administered in a pharmaceutical composition to a
patient as a therapeutic for pain as a result of injury or other
cartilage or joint condition as described herein. The peptide of
the present disclosure inhibits ion channels, such as Nav 1.7. The
peptide is expressed recombinantly or chemically synthesized,
wherein the peptide selected from SEQ ID NO: 24-SEQ ID NO: 274 or
SEQ ID NO: 314-SEQ ID NO: 564. Alternatively, the peptides of SEQ
ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564 are
mutated to maintain the cartilage homing function, but to add or
increase ion channel inhibition, such as to Nav 1. 7. Following
expression or synthesis, the peptide is used directly or conjugated
to a narcotic (e.g., oxycodone), a non-narcotic analgesic, a
natural counter-irritant (capsaicin), or a pain receptor channel
inhibitor (such as the TRPV4 inhibitor GSK2193874). Following
administration of the peptide, the peptide targets to the cartilage
affected by pain. One or more peptides are administered to a human
or animal subcutaneously, intravenously, or orally, or is injected
directly into a joint.
[0392] The peptide can be a peptide of SEQ ID NO: 108. The peptide
can also be a peptide of SEQ ID NO: 107. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 66
Treatment or Management of Pain with Peptides Only
[0393] This example describes a method for treating or managing
pain associated with a cartilage injury or disorder. This method is
used as a treatment for acute and/or chronic symptoms associated
with a cartilage injury or disorder. A peptide of the disclosure is
expressed and administered in a pharmaceutical composition to a
patient as a therapeutic for pain as a result of injury or other
cartilage or joint condition as described herein. The peptide of
the present disclosure inhibits ion channels, such as Nav 1.7. The
peptide is expressed recombinantly or chemically synthesized,
wherein the peptide selected from SEQ ID NO: 24-SEQ ID NO: 274 or
SEQ ID NO: 314-SEQ ID NO: 564. Alternatively, the peptides of SEQ
ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564 are
mutated to maintain the cartilage homing function, but to add or
increase ion channel inhibition, such as to Nav 1.7. Following
expression or synthesis, the peptide is used directly. Following
administration of the peptide, the peptide targets to the cartilage
affected by pain. One or more peptides are administered to a human
or animal subcutaneously, intravenously, or orally, or is injected
directly into a joint.
[0394] The peptide can be a peptide of SEQ ID NO: 108. The peptide
can also be a peptide of SEQ ID NO: 107. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564.
Example 67
Treatment of Chondrosarcoma
[0395] This example illustrates treatment of chondrosarcoma using
peptides of the present disclosure. A peptide of the present
disclosure is recombinantly expressed or chemically synthesized and
are used directly, after radiolabeling, or after conjugation to a
fluorophore or therapeutic compound, such as dasatinib. The peptide
or peptide conjugate is administered in a pharmaceutical
composition to a subject as a therapeutic for chondrosarcoma. One
or more peptides or peptide conjugates of the present disclosure
are administered to a subject. A subject can be a human or an
animal. The pharmaceutical composition is administered
subcutaneously, intravenously, orally, or injected directly into a
joint. The peptides or peptide conjugates target cartilage affected
by chondrosarcoma.
[0396] The peptide can be a peptide of SEQ ID NO: 106. The peptide
can also be a peptide of SEQ ID NO: 108. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 68
Treatment of Chordoma
[0397] This example illustrates treatment of chordoma using
peptides of the present disclosure. A peptide of the present
disclosure is recombinantly expressed or chemically synthesized and
are used directly, after radiolabeling, or after conjugation to a
fluorophore or therapeutic compound, such as dasatinib. The peptide
or peptide conjugate is administered in a pharmaceutical
composition to a subject as a therapeutic for chordoma. One or more
peptides or peptide conjugates of the present disclosure are
administered to a subject. A subject can be a human or an animal.
The pharmaceutical composition is administered subcutaneously,
intravenously, orally, or injected directly into a joint. The
peptides or peptide conjugates target cartilage affected by
chordoma.
[0398] The peptide can be a peptide of SEQ ID NO: 187. The peptide
can also be a peptide of SEQ ID NO: 24. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 69
Treatment for Rapid Pain Relief
[0399] This example illustrates rapid pain relief in patients
treated for rheumatoid arthritis or osteoarthritis with the
peptides or peptide conjugates of this disclosure. A peptide of
this disclosure is expressed recombinantly or chemically
synthesized, and then the N-terminus of the peptide is conjugated
to an active agent via an NHS ester to produce a peptide-active
agent conjugate. In some aspects the active agent such as a kidney
therapeutic from TABLE 4, TABLE 5, or TABLE 6. In some cases, the
peptide alone is administered to the subject.
[0400] The peptide or peptide-active agent conjugate is
administered to a subject in need thereof. The subject is a human
or non-human animal. The subject in need thereof has rheumatoid
arthritis or osteoarthritis. The peptide or peptide conjugate is
delivered via intravenous administration. Upon administration, the
peptide or peptide conjugate rapidly homes to cartilage. Rapid pain
relief within five minutes to an hour is experienced by the
subject, and pain relieve can last as long as over 3 hours.
[0401] The peptide can be a peptide of SEQ ID NO: 108. The peptide
can also be a peptide of SEQ ID NO: 33. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 70
Treatment for Lupus Nephritis
[0402] This example illustrates treatment of lupus nephritis using
peptides or peptide conjugates of this disclosure. A peptide of the
present disclosure is recombinantly expressed or chemically
synthesized and are used directly, after radiolabeling, or after
conjugation to a fluorophore or therapeutic compound, such as
abatacept or BIIB023.
[0403] The peptide or peptide conjugate is administered in a
pharmaceutical composition to a subject as a therapeutic for lupus
nephritis. The peptide is selected from any one of the peptides of
SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. One
or more peptides or peptide conjugates of the present disclosure
are administered to a subject. A subject can be a human or an
animal. The pharmaceutical composition is administered
subcutaneously, intravenously, orally, or injected directly. The
peptides or peptide conjugates target kidney affected by lupus
nephritis.
[0404] The peptide can be a peptide of SEQ ID NO: 27. The peptide
can also be a peptide of SEQ ID NO: 24. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 71
Treatment for Acute Kidney Injury (AHI)
[0405] This example illustrates treatment of acute kidney injury
(AM) using peptides or peptide conjugates of this disclosure. A
peptide of the present disclosure is recombinantly expressed or
chemically synthesized and are used directly, after radiolabeling,
or after conjugation to a fluorophore or therapeutic compound, such
as such as a kidney therapeutic from TABLE 4, TABLE 5, or TABLE
6.
[0406] The peptide or peptide conjugate is administered in a
pharmaceutical composition to a subject as a therapeutic for acute
kidney injury (AM). The peptide is selected from any one of the
peptides of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID
NO: 564. One or more peptides or peptide conjugates of the present
disclosure are administered to a subject. A subject can be a human
or an animal. The pharmaceutical composition is administered
subcutaneously, intravenously, orally, or injected directly into a
joint. The peptides or peptide conjugates target cartilage affected
by acute kidney injury (AM).
[0407] The peptide can be a peptide of SEQ ID NO: 108. The peptide
can also be a peptide of SEQ ID NO: 36. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 72
Treatment for Chronic Kidney Disease (CKD)
[0408] This example illustrates treatment of chronic kidney disease
(CKD) using peptides or peptide conjugates of this disclosure. A
peptide of the present disclosure is recombinantly expressed or
chemically synthesized and are used directly, after radiolabeling,
or after conjugation to a fluorophore or therapeutic compound, such
as a kidney therapeutic from TABLE 4, TABLE 5, or TABLE 6.
[0409] The peptide or peptide conjugate is administered in a
pharmaceutical composition to a subject as a therapeutic for
chronic kidney disease (CKD). The peptide is selected from any one
of the peptides of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO:
314-SEQ ID NO: 564. One or more peptides or peptide conjugates of
the present disclosure are administered to a subject. A subject can
be a human or an animal. The pharmaceutical composition is
administered subcutaneously, intravenously, orally, or injected
directly into a joint. The peptides or peptide conjugates target
cartilage affected by chronic kidney disease (CKD).
[0410] The peptide can be a peptide of SEQ ID NO: 199. The peptide
can be any peptide with the sequence selected from SEQ ID NO:
24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. Such
peptide-drug conjugates can be made using either a cleavable or
stable linker as described herein (e.g., EXAMPLES 21 and 22).
Example 73
Treatment for Hypertensive Kidney Damage
[0411] This example illustrates treatment of hypertensive kidney
damage using peptides or peptide conjugates of this disclosure. A
peptide of the present disclosure is recombinantly expressed or
chemically synthesized and are used directly, after radiolabeling,
or after conjugation to a fluorophore or therapeutic compound, such
as such as a kidney therapeutic from TABLE 4, TABLE 5, or TABLE
6.
[0412] The peptide or peptide conjugate is administered in a
pharmaceutical composition to a subject as a therapeutic for
hypertensive kidney damage. The peptide is selected from any one of
the peptides of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ
ID NO: 564. One or more peptides or peptide conjugates of the
present disclosure are administered to a subject. A subject can be
a human or an animal. The pharmaceutical composition is
administered subcutaneously, intravenously, orally, or injected
directly into a joint. The peptides or peptide conjugates target
cartilage affected by hypertensive kidney damage.
[0413] The peptide can be a peptide of SEQ ID NO: 27. The peptide
can also be a peptide of SEQ ID NO: 185. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 74
Treatment for Diabetic Nephropathy
[0414] This example illustrates treatment of diabetic nephropathy
using peptides or peptide conjugates of this disclosure. A peptide
of the present disclosure is recombinantly expressed or chemically
synthesized and are used directly, after radiolabeling, or after
conjugation to a fluorophore or therapeutic compound, such as such
as a kidney therapeutic from TABLE 4, TABLE 5, or TABLE 6.
[0415] The peptide or peptide conjugate is administered in a
pharmaceutical composition to a subject as a therapeutic for
diabetic nephropathy. The peptide is selected from any one of the
peptides of SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID
NO: 564. One or more peptides or peptide conjugates of the present
disclosure are administered to a subject. A subject can be a human
or an animal. The pharmaceutical composition is administered
subcutaneously, intravenously, orally, or injected directly into a
joint. The peptides or peptide conjugates target cartilage affected
by diabetic nephropathy.
[0416] The peptide can be a peptide of SEQ ID NO: 108. The peptide
can also be a peptide of SEQ ID NO: 22. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 75
Treatment for Renal Fibrosis
[0417] This example illustrates treatment of renal fibrosis using
peptides or peptide conjugates of this disclosure. A peptide of the
present disclosure is recombinantly expressed or chemically
synthesized and are used directly, after radiolabeling, or after
conjugation to a fluorophore or therapeutic compound, such as such
as a kidney therapeutic from TABLE 4, TABLE 5, or TABLE 6.
[0418] The peptide or peptide conjugate is administered in a
pharmaceutical composition to a subject as a therapeutic for renal
fibrosis. The peptide is selected from any one of the peptides of
SEQ ID NO: 24-SEQ ID NO: 274 or SEQ ID NO: 314-SEQ ID NO: 564. One
or more peptides or peptide conjugates of the present disclosure
are administered to a subject. A subject can be a human or an
animal. The pharmaceutical composition is administered
subcutaneously, intravenously, orally, or injected directly into a
joint. The peptides or peptide conjugates target cartilage affected
by renal fibrosis.
[0419] The peptide can be a peptide of SEQ ID NO: 199. The peptide
can also be a peptide of SEQ ID NO: 26. The peptide can be any
peptide with the sequence selected from SEQ ID NO: 24-SEQ ID NO:
274 or SEQ ID NO: 314-SEQ ID NO: 564. Such peptide-drug conjugates
can be made using either a cleavable or stable linker as described
herein (e.g., EXAMPLES 21 and 22).
Example 76
Peptide Variants Based on Multiple Sequence Alignment
[0420] This example illustrates using multiple sequence alignment
to design peptide variants with increased stability and decreased
immunogenicity. An alignment was generated using R language and an
"msa" software package, which codes for R language specific for
multiple alignments (Bodenhofer, U et al. Bioinformatics, 31 (24):
3997-3999 (2015)). FIG. 11 illustrates a multiple sequence
alignment of SEQ ID NO: 316, SEQ ID NO: 317, SEQ ID NO: 321, SEQ ID
NO: 333, SEQ ID NO: 337, SEQ ID NO: 338, SEQ ID NO: 340, SEQ ID NO:
398, SEQ ID NO: 474, SEQ ID NO: 483, SEQ ID NO: 486, and SEQ ID NO:
543-SEQ ID NO: 549. The alignment identified permissive or
preferred amino acids at a given location, and provided a guide for
discovery of novel peptide variants that could be generated and
that could retain essential properties such as structure, function,
peptide folding, biodistribution, or stability. SEQ ID NO: 21 and
SEQ ID NO: 295 are consensus sequences based on the above multiple
sequence alignment. SEQ ID NO: 21 is the same sequence as SEQ ID
NO: 295 but with an N-terminal "GS." Furthermore, based on the
ability to substitute K residues to R residues, the multiple
sequence alignment identified peptides of the family of sequences
of SEQ ID NO: 22 and SEQ ID NO: 296 as potential peptide variants
that could be generated and that could retain essential properties
such as structure, function, peptide folding, biodistribution, or
stability. Additionally, the multiple sequence alignment identified
SEQ ID NO: 312 as a conserved region within the sequences of the
alignment, which may, at least in part, be important for
maintaining the essential properties such as structure, function,
peptide folding, biodistribution, binding, accumulation, retention,
or stability.
Example 77
Peptide Immunogenicity
[0421] This example illustrates the testing of the immunogenicity
of a peptide. NetMHC II version 2.3 prediction software was used to
identify immunogenic peptides based on a neural network alignment
algorithm that predicts peptide binding to MHC Class II molecules.
The NetMHC II prediction software was utilized to determine the
putative peptide binding capability to DR, DQ, and DP MHC II
alleles and the strength of the interaction between peptide and MHC
II molecules. TABLE 7 shows the resulting immunogenicity score of
select peptides. The numbers of strong versus weak peptides were
tallied into each major MHC allele group (DR, DQ, and DP).
Additionally, the numbers of `unique strong` and `unique weak core`
peptides were also tallied. These data were used to predict which
peptides are less likely to induce an immunogenic response in
patients. For example, the stronger a peptide binds to an allele,
the more likely it is to be presented in a MHC/peptide combination
on an antigen presenting cell, thus triggering an immune response,
and a peptide that is predicted to bind to fewer alleles is more
likely to have weaker binding to given alleles and should be less
immunogenic.
TABLE-US-00034 TABLE 7 Immunogenicity Scores of Peptides Strong
Binding Unique Unique Alleles Strong Weak Binding Weak SEQ ID (DR +
DQ + Core Alleles Core NO: DP) Peptides (DR + DQ + DP) Peptides 108
1 + 0 + 0 1 + 0 + 0 7 + 1 + 0 7 + 2 + 0 260 0 + 0 + 0 0 + 0 + 0 4 +
1 + 3 6 + 1 + 1 261 0 + 0 + 0 0 + 0 + 0 4 + 1 + 3 6 + 2 + 1 262 1 +
0 + 0 2 + 0 + 0 5 + 1 + 3 7 + 1 + 1 263 1 + 0 + 0 2 + 0 + 0 6 + 1 +
3 5 + 1 + 1 264 0 + 0 + 0 0 + 0 + 0 4 + 1 + 3 6 + 2 + 1 265 0 + 0 +
0 0 + 0 + 0 5 + 1 + 3 7 + 2 + 1 271 0 + 0 + 1 0 + 0 + 1 7 + 4 + 1 8
+ 6 + 2 274 0 + 0 + 0 0 + 0 + 0 5 + 4 + 1 5 + 4 + 2 398 1 + 0 + 0 1
+ 0 + 0 7 + 1 + 0 7 + 1 + 0 550 0 + 0 + 0 0 + 0 + 0 4 + 0 + 3 6 + 0
+ 1 551 0 + 0 + 0 0 + 0 + 0 4 + 1 + 3 6 + 1 + 1 552 1 + 0 + 0 2 + 0
+ 0 5 + 0 + 3 7 + 0 + 1 553 1 + 0 + 0 2 + 0 + 0 6 + 0 + 3 5 + 0 + 1
554 0 + 0 + 0 0 + 0 + 0 4 + 1 + 3 6 + 1 + 1 555 0 + 0 + 0 0 + 0 + 0
5 + 1 + 3 7 + 1 + 1 561 0 + 0 + 1 0 + 0 + 1 7 + 4 + 1 8 + 6 + 2 564
0 + 0 + 0 0 + 0 + 0 5 + 4 + 1 5 + 5 + 2
Example 78
Peptide Variants
[0422] This example illustrates the design of variant peptide
sequences with increased stability, decreased regions of
immunogenicity, and the substitution of a tyrosine for
spectrophotometric reporting as compared to a parent peptide
sequence. Potential mutations to the parent peptide sequence, SEQ
ID NO: 108, that may result in a peptide with increased stability,
decreased immunogenicity, or increased absorbance at 270-280 nm
(such as the substitution to a tyrosine or tryptophan residue for
spectrophotometric reporting) were identified based on information
from multiple sequence alignment from EXAMPLE 76 and immunogenicity
testing from EXAMPLE 77.
[0423] In SEQ ID NO: 108, residue N7 is at risk for deamidation.
Based on the multiple sequence alignment of SEQ ID NO: 316, SEQ ID
NO: 317, SEQ ID NO: 321, SEQ ID NO: 333, SEQ ID NO: 337, SEQ ID NO:
338, SEQ ID NO: 340, SEQ ID NO: 398, SEQ ID NO: 483, SEQ ID NO:
486, and SEQ ID NO: 543-SEQ ID NO: 549, the candidate residue
mutations to best reduce this risk were N7S and N7G. N7S was
determined to be more likely to result in a peptide with desirable
properties such as folding and stability as shown by matches in the
alignment and conservationist presence in a peptide with high
stability (SEQ ID NO: 474).
[0424] Residue D18 is at risk for cleavage. Based on the multiple
sequence alignment, the candidate residue mutations to best reduce
cleavage at D18 are D18E and D18Q. D18E is the preferred choice
based on retaining charge.
[0425] Residue M25 is at risk for oxidation. Based on the multiple
sequence alignment, the candidate residue mutations to best reduce
oxidation were M25T and M25A. Based on the immunogenicity score of
peptides with each mutation, it was determined that M25T is the
better mutation, as it eliminates a significant source of
immunogenicity as compared to SEQ ID NO: 108 as well as the variant
with M25A, which did not eliminate the predicted immunogenicity of
the parent peptide of SEQ ID NO: 108.
[0426] Residue N32 is at risk for deamidation, at least in part due
to the neighboring residue S33. However, N32 is conserved across
Kv1. 3 binding cystine-dense peptides in the alignment of EXAMPLE
76, and implicated in receptor binding (Peigneur, S., Biochemistry,
55(32): 2927-35 (2016)). For certain applications, peptides are
designed to maintain this binding interaction, and for other
applications, peptides are designed to remove this binding
interaction. To maintain functionality, one candidate residue
mutation based on the multiple sequence alignment is S33R, which
would impact deamidation. However, it resulted in a predicted
increased immunogenicity score. Another candidate residue mutation
is S33G, but this may result in higher deamidation rates. If N32 is
mutated, the best candidate residue mutation based the multiple
sequence alignment in combination with the immunogenicity score was
N32Q despite it having a slight increase in immunogenicity. Other
options are N32A, N32S, or N32T. Alternatively, to remove
functionality, candidate mutations based on the multiple sequence
alignment are N32A and N32L, which are the preferred choices.
[0427] For the substitution to a tyrosine for spectrophotometric
reporting, the best candidate locations were T38Y (which had the
strongest precedence in the multiple sequence alignment and is
found in several of the stable peptides (e.g., SEQ ID NO: 474, SEQ
ID NO: 544, and SEQ ID NO: 545)), L17Y, and H36Y. However, T38Y may
slightly increase immunogenicity with respect to the DR allele.
Another option for spectrophometric absorbance is to substitute Trp
for the Leu at position 17.
[0428] Based on the above analysis, the following short list of
potential mutations for SEQ ID NO: 108 were compiled: N7S; D18E;
M25T; N32Q, N32A, N32S, N32T, N32L, S33G, and S33R (variants both
to retain function and to remove function of binding ion channel);
and L17Y, H36Y, and T38Y.
[0429] TABLE 8 provides some exemplary sequences using various
combinations of these mutations.
TABLE-US-00035 TABLE 8 Exemplary Sequence Variants of SEQ ID NO:
108 SEQ ID NO: Mutations 108 Parent 550 N5S, D16E, M23T, S31G 551
N5S, D16E, M23T, N30Q 552 N5S, D16E, M23T, S31R 553 D16E, M23T 554
D16E, M23T, N30Q 555 D16E, M23T, N30Q, T36Y 556 L15Y, D16E, M23T,
N30Q 557 D16E, M23T, N30Q, H34Y 558 N5S, D16E, M23T, N30Q, T36Y 559
N5S, L15Y, D16E, M23T, N30Q 560 N5S, D16E, M23T, N30Q, H34Y 561
D16E, M23T, N32A, T36Y 562 D16E, M23T, N32S, T36Y 563 D16E, M23T,
N32T, T36Y 564 D16E, M23T, T36Y
Example 79
Peptide-Budesonide Conjugate
[0430] This example describes conjugation of a peptide of any one
of SEQ ID NO: 260-SEQ ID NO: 274 or SEQ ID NO: 550-SEQ ID NO: 564
to budesonide. Budesonide is readily conjugated to any peptide
disclosed herein via standard chemistries such as those described
in, but not limited to, Bioconjugate Techniques by Greg Hermanson
(Elsevier Inc., 3rd edition, 2013) or by any of the methods
described in EXAMPLES 25-28.
[0431] The peptide-budesonide conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage
and/or kidneys. The subject is a human or animal and has
inflammation in the cartilage or kidney tissues. Upon
administration and homing of peptide-budesonide conjugates, the
inflammation in the cartilage and/or kidney tissues is
alleviated.
Example 80
Peptide-Dexamethasone Conjugate
[0432] This example describes conjugation of a peptide of any one
of SEQ ID NO: 260-SEQ ID NO: 274 or SEQ ID NO: 550-SEQ ID NO: 564
to dexamethasone. Dexamethasone is readily conjugated to any
peptide disclosed herein via standard chemistries such as those
described in, but not limited to, Bioconjugate Techniques by Greg
Hermanson (Elsevier Inc., 3.sup.rd edition, 2013) or by any of the
methods described in EXAMPLES 25-28.
[0433] The peptide-dexamethasone conjugates are administered to a
subject in need thereof and home, target, are directed to, are
retained by, accumulate in, migrate to, and/or bind to cartilage
and/or kidneys. The subject is a human or animal and has
inflammation in the cartilage or kidney tissues. Upon
administration and homing of peptide-dexamethasone conjugates, the
inflammation in the cartilage and/or kidney tissues is
alleviated.
Example 81
Peptide-Triamicinalone Acetonide Conjugate
[0434] This example describes conjugation of a peptide of any one
of SEQ ID NO: 260-SEQ ID NO: 274 or SEQ ID NO: 550-SEQ ID NO: 564
to triamicinalone acetonide. Triamicinalone acetonide is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition, 2013
or by any of the methods described in EXAMPLES 25-28.
[0435] The peptide-triamicinalone acetonide conjugates are
administered to a subject in need thereof and home, target, are
directed to, are retained by, accumulate in, migrate to, and/or
bind to cartilage and/or kidneys. The subject is a human or animal
and has inflammation in the cartilage or kidney tissues. Upon
administration and homing of peptide-triamicinalone acetonide
conjugates, the inflammation in the cartilage and/or kidney tissues
is alleviated.
Example 82
Peptide-Desciclesonide Acetonide Conjugate
[0436] This example describes conjugation of a peptide of any one
of SEQ ID NO: 260-SEQ ID NO: 274 or SEQ ID NO: 550-SEQ ID NO: 564
to desciclesonide acetonide. Desciclesonide acetonide is readily
conjugated to any peptide disclosed herein via standard chemistries
such as those described in, but not limited to, Bioconjugate
Techniques by Greg Hermanson (Elsevier Inc., 3.sup.rd edition,
2013) or by any of the methods described in EXAMPLES 25-28.
[0437] The peptide-desciclesonide acetonide conjugates are
administered to a subject in need thereof and home, target, are
directed to, are retained by, accumulate in, migrate to, and/or
bind to cartilage and/or kidneys. The subject is a human or animal
and has inflammation in the cartilage or kidney tissues. Upon
administration and homing of peptide-desciclesonide acetonide
conjugates, the inflammation in the cartilage and/or kidney tissues
is alleviated.
Example 83
Method of Peptide Synthesis
[0438] This example describes the synthesis of SEQ ID NO: 106, SEQ
ID NO: 108, and SEQ ID NO: 187.
[0439] A peptide of SEQ ID NO: 108 was made using Solid Phase
Peptide Synthesis (SPPS). After release of the peptide from the
solid phase, the peptide was purified prior to folding by oxidation
in solution. The folded peptide was further purified by
reversed-phase chromatography and lyophilized as a TFA salt. The
final SEQ ID NO: 108 peptide product had a purity of 96.1% and a
mass of 4,301.7 Da, which confirmed its identity as a peptide of
SEQ ID NO: 108.
[0440] A peptide of SEQ ID NO: 106 was made using Solid Phase
Peptide Synthesis (SPPS). After release of the peptide from the
solid phase, the peptide was folded by oxidation in solution. The
folded peptide was purified by reversed-phase chromatography and
lyophilized as a TFA salt. The final SEQ ID NO: 106 had a purity of
95.6% and a mass of 4,503.0 Da, which confirmed its identity as a
peptide of SEQ ID NO: 106.
[0441] A peptide of SEQ ID NO: 187 was made using Solid Phase
Peptide Synthesis (SPPS). After release of the peptide from the
solid phase, the peptide was folded by oxidation in solution. The
folded peptide was purified by reversed-phase chromatography and
lyophilized as a TFA salt. The final SEQ ID NO: 187 peptide product
had a purity of 95.5% and a mass of 4,154.0 Da, which confirmed its
identity as a peptide of SEQ ID NO: 187.
Example 84
Whole Body Autoradiography of Cartilage Homing Peptides
[0442] This example illustrates peptide homing to cartilage mice 5
minutes to 48 hours after administration of a radiolabeled peptide.
Signal from the radiolabeled peptides was found in all types of
cartilage at each time point examined. Each peptide was
radiolabeled by methylating lysines at the N-terminus as described
in EXAMPLE 2. As such, the peptide may contain methyl or dimethyl
lysines and a methylated or dimethylated amino terminus. A dose of
100 nmol radiolabeled peptide was administered via tail vein
injection in Female Harlan athymic nude mice, weighing 20-25 g. The
experiment was done in duplicate (n=2 animals per group). Each
radiolabeled peptide was allowed to freely circulate within the
animal for the described time period before the animals were
euthanized and sectioned.
[0443] Whole body autoradiography (WBA) sagittal sectioning was
performed as follows. At the end of the dosing period, mice were
frozen in a hexane/dry ice bath and then embedded in a frozen block
of carboxymethylcellulose. Whole animal sagittal slices were
prepared that resulted in thin frozen sections for imaging.
Sections were allowed to dessicate in a freezer prior to imaging.
For the autoradiography imaging, tape mounted thin sections were
freeze dried and radioactive samples were exposed to phosphoimager
plates. These plates were developed and the signal (densitometry)
from each organ was normalized to the signal found in the cardiac
blood of each animal. A signal in tissue darker than the signal
expected from blood in that tissue indicates accumulation in a
region, tissue, structure, or cell.
[0444] FIG. 14 illustrates autoradiography image of frozen sections
from a mouse, 3 hours after administration of 100 nmol of a
radiolabeled peptide of SEQ ID NO: 108. FIG. 14A illustrates the
.sup.17C signal in a frozen section of a mouse, 3 hours after
administration of 100 nmol of a radiolabeled peptide of SEQ ID NO:
108. The .sup.14C signal identifies the radiolabeled peptide
distribution in the cartilage of the mouse. FIG. 14B illustrates
the .sup.14C signal in a different frozen section of a mouse, 3
hours after administration of 100 nmol of a radiolabeled peptide of
SEQ ID NO: 108. The .sup.14C signal identifies the radiolabeled
peptide distribution in the cartilage of the mouse.
[0445] TABLE 9 shows the signal of radiolabeled peptides of SEQ ID
NO: 27 and SEQ ID NO: 108 in intervertebral discs (IVD) and knee
joints as a percentage of the blood. Because the peptides may
arrive at the joint within five minutes, a therapeutic effect from
the peptide or a conjugated active agent may begin quickly. A
therapeutic effect could be long lasting, due to continued presence
of detected agents at 48 hours and/or due to long lasting
pharmacodynamics effects.
TABLE-US-00036 TABLE 9 Signal of Radiolabeled Peptides of SEQ ID
NO: 27 and SEQ ID NO: 108 in IVD and Knee Joints as a Percentage of
Blood SEQ ID SEQ ID SEQ ID Hours NO: 27 IVD NO: 108 IVD NO: 108
Knee 0.08 164 404 0.5 369 510 1 961 1114 3 1779 3213 4059 8 3777
4990 24 833 5391 2137 48 3320 843
[0446] FIG. 15 illustrates autoradiography images of frozen
sections from a mouse, 3 hours after administration of 100 nmol of
a radiolabeled peptide of SEQ ID NO: 106. FIG. 15A illustrates the
.sup.14C signal in a frozen section of a mouse, 3 hours after
administration of 100 nmol of a radiolabeled peptide of SEQ ID NO:
106. The .sup.14C signal identifies the radiolabeled peptide
distribution in the cartilage of the mouse. FIG. 15B illustrates
the .sup.14C signal in a frozen section of a different mouse, 3
hours after administration of 100 nmol of a radiolabeled peptide of
SEQ ID NO: 106. The .sup.14C signal identifies the radiolabeled
peptide distribution in the cartilage of the mouse.
[0447] FIG. 16 illustrates autoradiography images of frozen
sections from a mouse, 3 hours after administration of 100 nmol of
a radiolabeled peptide of SEQ ID NO: 187. FIG. 16A illustrates the
.sup.14C signal in a frozen section of the mouse, 3 hours after
administration of 100 nmol of a radiolabeled peptide of SEQ ID NO:
187. The .sup.14C signal identifies the radiolabeled peptide
distribution in the cartilage of the mouse. FIG. 16B illustrates
the .sup.14C signal in a frozen section of a different mouse, 3
hours after administration of 100 nmol of a radiolabeled peptide of
SEQ ID NO: 187. The .sup.14C signal identifies the radiolabeled
peptide distribution in the cartilage of the mouse.
[0448] This data illustrates peptides of SEQ ID NO: 27, SEQ ID NO:
108, SEQ ID NO: 106 and SEQ ID NO: 187 homed to and accumulated in
the cartilage of the animals. The peptide of SEQ ID NO: 108 is a K
to R variant of a peptide of SEQ ID NO: 27. These data show that K
to R variants of cartilage homing peptides retained their cartilage
homing properties.
[0449] SEQ ID NO: 567 (GSGVPINVRSRGSRDSLDPSRRAGMRFGRSINSRSHSTP) is
a linearized version of SEQ ID NO: 108, where the knotted scaffold
of the peptide was removed by mutating out the cysteine residues
that form the disulfide bonds of the peptide to serine residues,
but retaining the rest of the sequence. TABLE 10 shows
quantification of signal as a percentage of signal in blood from a
linearized radiolabeled SEQ ID NO: 567 peptide in intervertebral
discs (IVD).
TABLE-US-00037 TABLE 10 Signal of Radiolabled Peptides of SEQ ID
NO: 567 in IVD as a Percentage of Blood 3 hr Ligated Kidneys 3 hr
Intact Kidneys 24 hr Intact Kidneys IVD 117 177 104
[0450] The peptide of SEQ ID NO: 567, a linearized version of the
peptide of SEQ ID NO: 108, homed to cartilage to a much lesser
extent than the folded knotted peptide (SEQ ID NO: 108). The signal
of the folded knotted peptide of SEQ ID NO: 108 was .about.20-fold
greater at 3 hours and .about.50-fold greater at 24 hours (TABLE 9)
as compared to the linearized peptide of SEQ ID NO: 567 (TABLE 10).
These results indicate that in addition to changes in primary
sequence or peptide charge, homing to cartilage can also be related
to changes in conformation, or tertiary structure. Namely, in some
cases, folded cystine-dense peptides can be exemplary cartilage
homers in comparison to unfolded, linearized peptides of the same
primary sequence (except for the mutated cysteine residues).
Example 85
Fluorescence of Cartilage Homing Peptides
[0451] This example illustrates peptide homing to cartilage mice
after administration of a peptide fluorophore conjugate. A peptide
of SEQ ID NO: 108 was chemically conjugated to one molecule of
Cyanine 5.5, and then imaged using the methods of EXAMPLE 13.
[0452] FIG. 10 shows white light images and corresponding whole
body fluorescence images of a mouse administered 10 nmol of a
peptide of SEQ ID NO: 108 conjugated to a Cy5.5 fluorophore (SEQ ID
NO: 108A) at 24 hours post-administration. FIG. 10A illustrates an
image of a frozen section of a mouse, 24 hours after administration
of 10 nmol of a peptide of SEQ ID NO: 108 conjugated to a Cy5.5
fluorophore (SEQ ID NO: 108A). FIG. 10B illustrates the
fluorescence signal in the mouse, corresponding to the section
shown in FIG. 10A, 24 hours after administration of 10 nmol of a
peptide of SEQ ID NO: 108 conjugated to a Cy5.5 fluorophore (SEQ ID
NO: 108A). FIG. 10C illustrates an image of a different frozen
section of the mouse, 24 hours after administration of 10 nmol of a
peptide of SEQ ID NO: 108 conjugated to a Cy5.5 fluorophore (SEQ ID
NO: 108A). FIG. 10D illustrates the fluorescence signal in the
mouse, corresponding to the section shown in FIG. 10C, 24 hours
after administration of 10 nmol of a peptide of SEQ ID NO: 108
conjugated to a Cy5.5 fluorophore (SEQ ID NO: 108A). FIG. 10E
illustrates an image of a different frozen section of the mouse, 24
hours after administration of 10 nmol of a peptide of SEQ ID NO:
108 conjugated to a Cy5.5 fluorophore (SEQ ID NO: 108A). FIG. 10F
illustrates a fluorescence signal in the mouse, corresponding to
the section shown in FIG. 10E, 24 hours after administration of 10
nmol of a peptide of SEQ ID NO: 108 conjugated to a Cy5.5
fluorophore (SEQ ID NO: 108A).
[0453] FIG. 13 shows IVIS fluorescence imaging of an isolated hind
limb from a first mouse and an isolated hind limb from a second
mouse after administration of 10 nmol SEQ ID NO: 108 peptide
conjugated to a Cy5.5 fluorophore (SEQ ID NO: 108A). FIG. 13A shows
the right hind limb with skin removed from a first mouse and from a
second mouse 3 hours after peptide administration. FIG. 13B shows
the right hind limb with muscle removed from a first mouse and from
a second mouse 3 hours after peptide administration. FIG. 13C shows
the right hind limb with skin removed from a first mouse and from a
second mouse 24 hours after peptide administration. FIG. 13D shows
the right hind limb with muscle removed from a first mouse and from
a second mouse 24 hours after peptide administration. FIG. 13E
shows the right hind limb with skin removed from a first mouse and
from a second mouse 48 hours after peptide administration. FIG. 13F
shows the right hind limb with muscle removed from a first mouse
and from a second mouse 48 hours after peptide administration. FIG.
13G shows the right hind limb with skin removed from a first mouse
and from a second mouse 72 hours after peptide administration. FIG.
13H shows the right hind limb with muscle removed from a first
mouse and from a second mouse 72 hours after peptide
administration. Peptide fluorescence was observed in the knee
joints of isolated right hind limbs at all time points tested.
Example 86
Peptide Resistance Under Various Conditions
[0454] This example illustrates peptide stability under various
stress conditions such as high temperature, low pH, reducing
agents, and proteases. To determine resistance to high
temperatures, cystine-dense peptides (CDPs) were incubated at 0.5
mM in PBS at 75.degree. C. or 100.degree. C. for 1 h and pelleted,
and the supernatant was analyzed with reversed-phase chromatography
(RPC). To determine resistance to proteolytic digestion, CDPs were
mixed with 50 U of porcine pepsin, in simulated gastric fluid at pH
1.0, or 50 U of porcine trypsin in PBS, incubated for 30 minutes at
37.degree. C. and analyzed with RPC. Oxidized and reduced forms
(prepared through addition 10 mM DTT) were compared. Circular
Dichroism spectroscopy was used in order to measure the secondary
structure of peptides with a Jasco J-720W spectropolarimeter in a
cell with a 1.0-mm path length, and CDPs were diluted into 20 mM
phosphate buffer, pH 7.4, at a concentration of 15-25 .mu.M. These
conditions were expected to denature or degrade conventional
globular proteins and many peptides. In TABLE 11, "high" resistance
indicated a high amount of the peptide remained or was retained as
unmodified under the given experimental conditions and "low"
resistance indicated a low amount of the peptide remained or was
retained unmodified under the given experimental conditions.
Notably, the experimental conditions described in this example were
more extreme stress conditions than to many standard in vivo or
physiologic conditions, in vitro conditions, conditions during
manufacturing, and handling conditions. As such, even "low"
resistance can indicate meaningful resistance to these stress
conditions that may have applicability for a number of uses
described herein. The data from these studies are shown in TABLE
11. The peptides tested, SEQ ID NO: 315, SEQ ID NO: 317 and SEQ ID
NO: 482, showed high resistance to one or more of the conditions
tested.
TABLE-US-00038 TABLE 11 Resistance of SEQ ID NO: 317, SEQ ID NO:
315, and SEQ ID NO: 482 to Various Conditions SEQ Resistance to
Resistance Resistance Resistance Resistance ID NO: Reduction to
75.degree. C. to 100.degree. C. to Pepsin to Trypsin 27 High High
High High High 25 Low High Low High Low 192 Low High Low High
Low
[0455] While certain embodiments of the present disclosure have
been exemplified or shown and described herein, it will be apparent
to those skilled in the art that such embodiments are provided by
way of example only. It is not intended that the disclosure be
limited by the specific examples provided within the specification.
While the disclosure has been described with reference to the
aforementioned specification, the descriptions and illustrations of
the embodiments herein are not meant to be construed in a limiting
sense. Numerous variations, changes, and substitutions will now
occur to those skilled in the art without departing from the
disclosure. Furthermore, it shall be understood that all
embodiments of the disclosure are not limited to the specific
depictions, configurations or relative proportions set forth herein
which depend upon a variety of conditions and variables. It should
be understood that various alternatives to the embodiments of the
disclosure described herein may be employed in practicing the
disclosure. It is therefore contemplated that the disclosure shall
also cover any such alternatives, modifications, variations or
equivalents. It is intended that the following claims define the
scope of the disclosure and that methods and structures within the
scope of these claims and their equivalents be covered thereby.
Sequence CWU 1
1
570142PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(5)..(5)Any amino acid, amino acid
analogue or nullMOD_RES(7)..(8)Any amino acid, amino acid analogue
or nullMOD_RES(11)..(11)Any amino acid, amino acid analogue or
nullMOD_RES(17)..(17)Any amino acid, amino acid analogue or
nullMOD_RES(22)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(28)..(28)Any amino acid, amino acid analogue or
nullMOD_RES(32)..(32)Any amino acid, amino acid analogue or
nullMOD_RES(39)..(42)Any amino acid, amino acid analogue or null
1Gly Ser Gly Val Xaa Ile Xaa Xaa Lys Cys Xaa Gly Ser Lys Gln Cys1 5
10 15Xaa Asp Pro Cys Lys Xaa Xaa Xaa Gly Xaa Arg Xaa Gly Lys Cys
Xaa 20 25 30Asn Lys Lys Cys Lys Cys Xaa Xaa Xaa Xaa 35
40242PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(5)..(5)P or RMOD_RES(7)..(7)P or
NMOD_RES(8)..(8)V or IMOD_RES(11)..(11)S, T, R or
KMOD_RES(17)..(17)Y or LMOD_RES(22)..(22)Q, R or
KMOD_RES(23)..(23)A, K or RMOD_RES(24)..(24)T or
AMOD_RES(26)..(26)C or MMOD_RES(28)..(28)F or NMOD_RES(32)..(32)M
or IMOD_RES(39)..(39)Y or TMOD_RES(40)..(40)G or
PMISC_FEATURE(41)..(42)May or may not be present 2Gly Ser Gly Val
Xaa Ile Xaa Xaa Lys Cys Xaa Gly Ser Lys Gln Cys1 5 10 15Xaa Asp Pro
Cys Lys Xaa Xaa Xaa Gly Xaa Arg Xaa Gly Lys Cys Xaa 20 25 30Asn Lys
Lys Cys Lys Cys Xaa Xaa Cys Gly 35 40341PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(3)..(6)Any amino acid, amino acid analogue or
nullMOD_RES(8)..(8)Any amino acid, amino acid analogue or
nullMOD_RES(10)..(10)Any amino acid, amino acid analogue or
nullMOD_RES(17)..(17)Any amino acid, amino acid analogue or
nullMOD_RES(21)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(28)Any amino acid, amino acid analogue or
nullMOD_RES(31)..(32)Any amino acid, amino acid analogue or
nullMOD_RES(34)..(34)Any amino acid, amino acid analogue or null
3Gly Ser Xaa Xaa Xaa Xaa Ile Xaa Cys Xaa Gly Ser Lys Gln Cys Tyr1 5
10 15Xaa Pro Cys Lys Xaa Xaa Thr Gly Cys Xaa Xaa Xaa Lys Cys Xaa
Xaa 20 25 30Lys Xaa Cys Lys Cys Tyr Gly Cys Gly 35
40441PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMISC_FEATURE(3)..(4)May or may not be
presentMOD_RES(5)..(5)E, G or nullMOD_RES(6)..(6)V, S or
nullMOD_RES(8)..(8)R or SMOD_RES(10)..(10)S or TMOD_RES(17)..(17)G
or DMOD_RES(21)..(21)Q or RMOD_RES(22)..(22)Q or
KMOD_RES(26)..(26)T or PMOD_RES(27)..(27)N or QMOD_RES(28)..(28)S
or AMOD_RES(31)..(31)M or LMOD_RES(32)..(32)N or
QMOD_RES(34)..(34)V or S 4Gly Ser Gly Ser Xaa Xaa Ile Xaa Cys Xaa
Gly Ser Lys Gln Cys Tyr1 5 10 15Xaa Pro Cys Lys Xaa Xaa Thr Gly Cys
Xaa Xaa Xaa Lys Cys Xaa Xaa 20 25 30Lys Xaa Cys Lys Cys Tyr Gly Cys
Gly 35 40542PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptideMOD_RES(3)..(5)Any amino acid, amino
acid analogue or nullMOD_RES(7)..(7)Any amino acid, amino acid
analogue or nullMOD_RES(9)..(9)Any amino acid, amino acid analogue
or nullMOD_RES(11)..(11)Any amino acid, amino acid analogue or
nullMOD_RES(13)..(14)Any amino acid, amino acid analogue or
nullMOD_RES(16)..(17)Any amino acid, amino acid analogue or
nullMOD_RES(20)..(20)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(33)..(33)Any amino acid, amino acid analogue or
nullMOD_RES(35)..(35)Any amino acid, amino acid analogue or
nullMOD_RES(38)..(38)Any amino acid, amino acid analogue or
nullMOD_RES(42)..(42)Any amino acid, amino acid analogue or null
5Gly Ser Xaa Xaa Xaa Val Xaa Ile Xaa Val Xaa Cys Xaa Xaa Ser Xaa1 5
10 15Xaa Cys Leu Xaa Pro Cys Lys Xaa Ala Gly Met Arg Phe Gly Lys
Cys 20 25 30Xaa Asn Xaa Lys Cys Xaa Cys Thr Pro Xaa 35
40642PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMISC_FEATURE(3)..(3)May or may not be
presentMOD_RES(4)..(4)G, S or nullMOD_RES(5)..(5)G, S or
nullMOD_RES(7)..(7)P or RMOD_RES(9)..(9)N or PMOD_RES(11)..(11)K or
SMOD_RES(13)..(13)R or KMOD_RES(14)..(14)G or HMOD_RES(16)..(16)R
or GMOD_RES(17)..(17)D or QMOD_RES(20)..(20)D or
KMOD_RES(24)..(24)K or DMOD_RES(33)..(33)I or MMOD_RES(35)..(35)S
or GMOD_RES(38)..(38)H or DMISC_FEATURE(42)..(42)May or may not be
present 6Gly Ser Gly Xaa Xaa Val Xaa Ile Xaa Val Xaa Cys Xaa Xaa
Ser Xaa1 5 10 15Xaa Cys Leu Xaa Pro Cys Lys Xaa Ala Gly Met Arg Phe
Gly Lys Cys 20 25 30Xaa Asn Xaa Lys Cys Xaa Cys Thr Pro Lys 35
40737PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(3)..(3)Any amino
acidMOD_RES(5)..(5)Any amino acidMOD_RES(9)..(9)Any amino
acidMOD_RES(15)..(15)Any amino acidMOD_RES(20)..(20)Any amino
acidMOD_RES(22)..(22)Any amino acidMOD_RES(24)..(24)Any amino
acidMOD_RES(32)..(32)Any amino acidMOD_RES(37)..(37)Any amino
acidSee specification as filed for detailed description of
substitutions and preferred embodiments 7Gly Ser Xaa Val Xaa Val
Lys Cys Xaa Gly Ser Lys Gln Cys Xaa Pro1 5 10 15Cys Lys Arg Xaa Gly
Xaa Arg Xaa Gly Lys Cys Ile Asn Lys Lys Xaa 20 25 30Cys Lys Cys Tyr
Xaa 35834PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(3)..(3)Any amino
acidMOD_RES(7)..(7)Any amino acidMOD_RES(13)..(13)Any amino
acidMOD_RES(15)..(15)Any amino acidMOD_RES(18)..(18)Any amino
acidMOD_RES(20)..(20)Any amino acidMOD_RES(29)..(29)Any amino
acidMOD_RES(34)..(34)Any amino acidSee specification as filed for
detailed description of substitutions and preferred embodiments
8Gly Ser Xaa Gly Cys Val Xaa Lys Cys Arg Pro Gly Xaa Lys Xaa Cys1 5
10 15Cys Xaa Pro Xaa Lys Arg Cys Ser Arg Arg Phe Gly Xaa Lys Lys
Cys 20 25 30Lys Xaa943PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptideMOD_RES(3)..(3)Any amino
acid, amino acid analogue or nullMOD_RES(5)..(7)Any amino acid,
amino acid analogue or nullMOD_RES(11)..(11)Any amino acid, amino
acid analogue or nullMOD_RES(17)..(18)Any amino acid, amino acid
analogue or nullMOD_RES(23)..(24)Any amino acid, amino acid
analogue or nullMOD_RES(26)..(26)Any amino acid, amino acid
analogue or nullMOD_RES(28)..(28)Any amino acid, amino acid
analogue or nullMOD_RES(36)..(36)Any amino acid, amino acid
analogue or nullMOD_RES(41)..(43)Any amino acid, amino acid
analogue or null 9Gly Ser Xaa Val Xaa Xaa Xaa Val Lys Cys Xaa Gly
Ser Lys Gln Cys1 5 10 15Xaa Xaa Pro Cys Lys Arg Xaa Xaa Gly Xaa Arg
Xaa Gly Lys Cys Ile 20 25 30Asn Lys Lys Xaa Cys Lys Cys Tyr Xaa Xaa
Xaa 35 401047PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptideMOD_RES(3)..(5)Any amino
acidMOD_RES(9)..(12)Any amino acidMOD_RES(18)..(18)Any amino
acidMOD_RES(20)..(20)Any amino acidMOD_RES(23)..(23)Any amino
acidMOD_RES(25)..(25)Any amino acidMOD_RES(34)..(37)Any amino
acidMOD_RES(42)..(47)Any amino acid 10Gly Ser Xaa Xaa Xaa Gly Cys
Val Xaa Xaa Xaa Xaa Lys Cys Arg Pro1 5 10 15Gly Xaa Lys Xaa Cys Cys
Xaa Pro Xaa Lys Arg Cys Ser Arg Arg Phe 20 25 30Gly Xaa Xaa Xaa Xaa
Lys Lys Cys Lys Xaa Xaa Xaa Xaa Xaa Xaa 35 40 451142PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(5)..(5)Any amino acid, amino acid analogue or
nullMOD_RES(7)..(8)Any amino acid, amino acid analogue or
nullMOD_RES(11)..(11)Any amino acid, amino acid analogue or
nullMOD_RES(17)..(17)Any amino acid, amino acid analogue or
nullMOD_RES(22)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(28)..(28)Any amino acid, amino acid analogue or
nullMOD_RES(32)..(32)Any amino acid, amino acid analogue or
nullMOD_RES(39)..(42)Any amino acid, amino acid analogue or null
11Gly Ser Gly Val Xaa Ile Xaa Xaa Arg Cys Xaa Gly Ser Arg Gln Cys1
5 10 15Xaa Asp Pro Cys Arg Xaa Xaa Xaa Gly Xaa Arg Xaa Gly Arg Cys
Xaa 20 25 30Asn Arg Arg Cys Arg Cys Xaa Xaa Xaa Xaa 35
401242PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(5)..(5)P or RMOD_RES(7)..(7)P or
NMOD_RES(8)..(8)V or IMOD_RES(11)..(11)S, T, R or
KMOD_RES(17)..(17)Y or LMOD_RES(22)..(22)Q, R or
KMOD_RES(23)..(23)A, K or RMOD_RES(24)..(24)T or
AMOD_RES(26)..(26)C or MMOD_RES(28)..(28)F or NMOD_RES(32)..(32)M
or IMOD_RES(39)..(39)Y or TMOD_RES(40)..(40)G or
PMISC_FEATURE(41)..(42)May or may not be present 12Gly Ser Gly Val
Xaa Ile Xaa Xaa Arg Cys Xaa Gly Ser Arg Gln Cys1 5 10 15Xaa Asp Pro
Cys Arg Xaa Xaa Xaa Gly Xaa Arg Xaa Gly Arg Cys Xaa 20 25 30Asn Arg
Arg Cys Arg Cys Xaa Xaa Cys Gly 35 401341PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(3)..(6)Any amino acid, amino acid analogue or
nullMOD_RES(8)..(8)Any amino acid, amino acid analogue or
nullMOD_RES(10)..(10)Any amino acid, amino acid analogue or
nullMOD_RES(17)..(17)Any amino acid, amino acid analogue or
nullMOD_RES(21)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(28)Any amino acid, amino acid analogue or
nullMOD_RES(31)..(32)Any amino acid, amino acid analogue or
nullMOD_RES(34)..(34)Any amino acid, amino acid analogue or null
13Gly Ser Xaa Xaa Xaa Xaa Ile Xaa Cys Xaa Gly Ser Arg Gln Cys Tyr1
5 10 15Xaa Pro Cys Arg Xaa Xaa Thr Gly Cys Xaa Xaa Xaa Arg Cys Xaa
Xaa 20 25 30Arg Xaa Cys Arg Cys Tyr Gly Cys Gly 35
401441PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMISC_FEATURE(3)..(4)May or may not be
presentMOD_RES(5)..(5)E, G or nullMOD_RES(6)..(6)V, S or
nullMOD_RES(8)..(8)R or SMOD_RES(10)..(10)S or TMOD_RES(17)..(17)G
or DMOD_RES(21)..(21)Q or RMOD_RES(22)..(22)Q, R or
KMOD_RES(26)..(26)T or PMOD_RES(27)..(27)N or QMOD_RES(28)..(28)S
or AMOD_RES(31)..(31)M or LMOD_RES(32)..(32)N or
QMOD_RES(34)..(34)V or S 14Gly Ser Gly Ser Xaa Xaa Ile Xaa Cys Xaa
Gly Ser Arg Gln Cys Tyr1 5 10 15Xaa Pro Cys Arg Xaa Xaa Thr Gly Cys
Xaa Xaa Xaa Arg Cys Xaa Xaa 20 25 30Arg Xaa Cys Arg Cys Tyr Gly Cys
Gly 35 401542PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptideMOD_RES(3)..(5)Any amino acid, amino
acid analogue or nullMOD_RES(7)..(7)Any amino acid, amino acid
analogue or nullMOD_RES(9)..(9)Any amino acid, amino acid analogue
or nullMOD_RES(11)..(11)Any amino acid, amino acid analogue or
nullMOD_RES(13)..(14)Any amino acid, amino acid analogue or
nullMOD_RES(16)..(17)Any amino acid, amino acid analogue or
nullMOD_RES(20)..(20)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(33)..(33)Any amino acid, amino acid analogue or
nullMOD_RES(35)..(35)Any amino acid, amino acid analogue or
nullMOD_RES(38)..(38)Any amino acid, amino acid analogue or
nullMOD_RES(42)..(42)Any amino acid, amino acid analogue or null
15Gly Ser Xaa Xaa Xaa Val Xaa Ile Xaa Val Xaa Cys Xaa Xaa Ser Xaa1
5 10 15Xaa Cys Leu Xaa Pro Cys Arg Xaa Ala Gly Met Arg Phe Gly Arg
Cys 20 25 30Xaa Asn Xaa Arg Cys Xaa Cys Thr Pro Xaa 35
401642PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMISC_FEATURE(3)..(3)May or may not be
presentMOD_RES(4)..(4)G, S or nullMOD_RES(5)..(5)G, S or
nullMOD_RES(7)..(7)P or RMOD_RES(9)..(9)N or PMOD_RES(11)..(11)R, K
or SMOD_RES(13)..(13)R or KMOD_RES(14)..(14)G or
HMOD_RES(16)..(16)R or GMOD_RES(17)..(17)D or QMOD_RES(20)..(20)D,
R or KMOD_RES(24)..(24)K, R or DMOD_RES(33)..(33)I or
MMOD_RES(35)..(35)S or GMOD_RES(38)..(38)H or DMOD_RES(42)..(42)K,
R or null 16Gly Ser Gly Xaa Xaa Val Xaa Ile Xaa Val Xaa Cys Xaa Xaa
Ser Xaa1 5 10 15Xaa Cys Leu Xaa Pro Cys Arg Xaa Ala Gly Met Arg Phe
Gly Arg Cys 20 25 30Xaa Asn Xaa Arg Cys Xaa Cys Thr Pro Xaa 35
401737PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(3)..(3)Any amino
acidMOD_RES(5)..(5)Any amino acidMOD_RES(9)..(9)Any amino
acidMOD_RES(15)..(15)Any amino acidMOD_RES(20)..(20)Any amino
acidMOD_RES(22)..(22)Any amino acidMOD_RES(24)..(24)Any amino
acidMOD_RES(32)..(32)Any amino acidMOD_RES(37)..(37)Any amino
acidSee specification as filed for detailed description of
substitutions and preferred embodiments 17Gly Ser Xaa Val Xaa Val
Arg Cys Xaa Gly Ser Arg Gln Cys Xaa Pro1 5 10 15Cys Arg Arg Xaa Gly
Xaa Arg Xaa Gly Arg Cys Ile Asn Arg Arg Xaa 20 25 30Cys Arg Cys Tyr
Xaa 351834PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(3)..(3)Any amino
acidMOD_RES(7)..(7)Any amino acidMOD_RES(13)..(13)Any amino
acidMOD_RES(15)..(15)Any amino acidMOD_RES(18)..(18)Any amino
acidMOD_RES(20)..(20)Any amino acidMOD_RES(29)..(29)Any amino
acidMOD_RES(34)..(34)Any amino acidSee specification as filed for
detailed description of substitutions and preferred embodiments
18Gly Ser Xaa Gly Cys Val Xaa Arg Cys Arg Pro Gly Xaa Arg Xaa Cys1
5 10 15Cys Xaa Pro Xaa Arg Arg Cys Ser Arg Arg Phe Gly Xaa Arg Arg
Cys 20 25 30Arg Xaa1943PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptideMOD_RES(3)..(3)Any amino
acid analogue or nullMOD_RES(5)..(7)Any amino acid analogue or
nullMOD_RES(11)..(11)Any amino acid analogue or
nullMOD_RES(17)..(18)Any amino acid analogue or
nullMOD_RES(23)..(24)Any amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid analogue or
nullMOD_RES(28)..(28)Any amino acid analogue or
nullMOD_RES(36)..(36)Any amino acid analogue or
nullMOD_RES(41)..(43)Any amino acid analogue or null 19Gly Ser Xaa
Val Xaa Xaa Xaa Val Arg Cys Xaa Gly Ser Arg Gln Cys1 5 10 15Xaa Xaa
Pro Cys Arg Arg Xaa Xaa Gly Xaa Arg Xaa Gly Arg Cys Ile 20 25 30Asn
Arg Arg Xaa Cys Arg Cys Tyr Xaa Xaa Xaa 35 402047PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(3)..(5)Any amino acid analogue or
nullMOD_RES(9)..(12)Any amino acid analogue or
nullMOD_RES(18)..(18)Any amino acid analogue or
nullMOD_RES(20)..(20)Any amino acid analogue or
nullMOD_RES(23)..(23)Any amino acid analogue or
nullMOD_RES(25)..(25)Any amino acid analogue or
nullMOD_RES(34)..(37)Any amino acid analogue or
nullMOD_RES(42)..(47)Any amino acid analogue or null 20Gly Ser Xaa
Xaa Xaa Gly Cys Val Xaa Xaa Xaa Xaa Arg Cys Arg Pro1 5 10 15Gly Xaa
Arg Xaa Cys Cys Xaa Pro Xaa Arg Arg Cys Ser Arg Arg Phe 20 25 30Gly
Xaa Xaa Xaa Xaa Arg Arg Cys Arg Xaa Xaa Xaa Xaa Xaa Xaa 35 40
452142PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(3)..(3)Any amino acid, amino acid
analogue or nullMOD_RES(5)..(5)Any amino acid, amino acid analogue
or nullMOD_RES(7)..(7)Any amino acid, amino acid analogue or
nullMOD_RES(11)..(11)Any amino acid, amino acid analogue or
nullMOD_RES(14)..(14)Any amino acid, amino acid analogue or
nullMOD_RES(18)..(18)Any amino acid, amino acid analogue or
nullMOD_RES(22)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(28)..(28)Any amino acid, amino acid analogue or
nullMOD_RES(37)..(37)Any amino acid, amino acid analogue or
nullMOD_RES(39)..(39)Any amino acid, amino acid analogue or
nullMOD_RES(41)..(42)Any amino acid, amino acid analogue or null
21Gly Ser Xaa Val Xaa Ile Xaa Val Lys Cys Xaa Gly Ser Xaa Gln Cys1
5 10 15Leu Xaa Pro Cys Lys Xaa Ala Xaa Gly Xaa Arg Xaa Gly Lys Cys
Met 20 25 30Asn Gly Lys Cys Xaa Cys Xaa Pro Xaa Xaa 35
402242PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(3)..(3)Any amino acid, amino acid
analogue or nullMOD_RES(5)..(5)Any amino acid, amino acid analogue
or nullMOD_RES(7)..(7)Any amino acid, amino acid analogue or
nullMOD_RES(11)..(11)Any amino acid, amino acid analogue or
nullMOD_RES(14)..(14)Any amino acid, amino acid analogue or
nullMOD_RES(18)..(18)Any amino acid, amino acid analogue or
nullMOD_RES(22)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(28)..(28)Any amino acid, amino acid analogue or
nullMOD_RES(37)..(37)Any amino acid, amino acid analogue or
nullMOD_RES(39)..(39)Any amino acid, amino acid analogue or
nullMOD_RES(41)..(42)Any amino acid, amino acid analogue or null
22Gly Ser Xaa Val Xaa Ile Xaa Val Arg Cys Xaa Gly Ser Xaa Gln Cys1
5 10 15Leu Xaa Pro Cys Arg Xaa Ala Xaa Gly Xaa Arg Xaa Gly Arg Cys
Met 20 25 30Asn Gly Arg Cys Xaa Cys Xaa Pro Xaa Xaa 35
402339PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(7)..(7)N, S or GMOD_RES(17)..(17)L or
YMOD_RES(18)..(18)D or EMOD_RES(25)..(25)M or TMOD_RES(32)..(32)N,
Q, A, S, T or LMOD_RES(33)..(33)S, G or RMOD_RES(36)..(36)H or
YMOD_RES(38)..(38)T or Y 23Gly Ser Gly Val Pro Ile Xaa Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Xaa Xaa Pro Cys Arg Arg Ala Gly Xaa
Arg Phe Gly Arg Cys Ile Xaa 20 25 30Xaa Arg Cys Xaa Cys Xaa Pro
352434PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideSee specification as filed for detailed
description of substitutions and preferred embodiments 24Gly Ser
Gly Ile Val Cys Lys Val Cys Lys Ile Ile Cys Gly Met Gln1 5 10 15Gly
Lys Lys Val Asn Ile Cys Lys Ala Pro Ile Lys Cys Lys Cys Lys 20 25
30Lys Gly2537PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptideSee specification as filed for
detailed description of substitutions and preferred embodiments
25Gly Ser Ser Glu Lys Asp Cys Ile Lys His Leu Gln Arg Cys Arg Glu1
5 10 15Asn Lys Asp Cys Cys Ser Lys Lys Cys Ser Arg Arg Gly Thr Asn
Pro 20 25 30Glu Lys Arg Cys Arg 352639PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptideSee
specification as filed for detailed description of substitutions
and preferred embodiments 26Gly Ser Val Arg Ile Pro Val Ser Cys Lys
His Ser Gly Gln Cys Leu1 5 10 15Lys Pro Cys Lys Asp Ala Gly Met Arg
Phe Gly Lys Cys Met Asn Gly 20 25 30Lys Cys Asp Cys Thr Pro Lys
352739PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideSee specification as filed for detailed
description of substitutions and preferred embodiments 27Gly Ser
Gly Val Pro Ile Asn Val Lys Cys Arg Gly Ser Arg Asp Cys1 5 10 15Leu
Asp Pro Cys Lys Lys Ala Gly Met Arg Phe Gly Lys Cys Ile Asn 20 25
30Ser Lys Cys His Cys Thr Pro 352835PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptideSee
specification as filed for detailed description of substitutions
and preferred embodiments 28Gly Ser Ala Val Cys Val Tyr Arg Thr Cys
Asp Lys Asp Cys Lys Arg1 5 10 15Arg Gly Tyr Arg Ser Gly Lys Cys Ile
Asn Asn Ala Cys Lys Cys Tyr 20 25 30Pro Tyr Gly 352937PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptideSee
specification as filed for detailed description of substitutions
and preferred embodiments 29Gly Ser Ile Ser Cys Thr Gly Ser Lys Gln
Cys Tyr Asp Pro Cys Lys1 5 10 15Arg Lys Thr Gly Cys Pro Asn Ala Lys
Cys Met Asn Lys Ser Cys Lys 20 25 30Cys Tyr Gly Cys Gly
353040PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideSee specification as filed for detailed
description of substitutions and preferred embodiments 30Gly Ser
Gln Val Gln Thr Asn Val Lys Cys Gln Gly Gly Ser Cys Ala1 5 10 15Ser
Val Cys Arg Arg Glu Ile Gly Val Ala Ala Gly Lys Cys Ile Asn 20 25
30Gly Lys Cys Val Cys Tyr Arg Asn 35 403139PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptideSee
specification as filed for detailed description of substitutions
and preferred embodiments 31Gly Ser Glu Val Ile Arg Cys Ser Gly Ser
Lys Gln Cys Tyr Gly Pro1 5 10 15Cys Lys Gln Gln Thr Gly Cys Thr Asn
Ser Lys Cys Met Asn Lys Val 20 25 30Cys Lys Cys Tyr Gly Cys Gly
353235PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideSee specification as filed for detailed
description of substitutions and preferred embodiments 32Gly Ser
Ala Cys Lys Gly Val Phe Asp Ala Cys Thr Pro Gly Lys Asn1 5 10 15Glu
Cys Cys Pro Asn Arg Val Cys Ser Asp Lys His Lys Trp Cys Lys 20 25
30Trp Lys Leu 353340PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptideSee specification as filed for
detailed description of substitutions and preferred embodiments
33Gly Ser Gln Ile Tyr Thr Ser Lys Glu Cys Asn Gly Ser Ser Glu Cys1
5 10 15Tyr Ser His Cys Glu Gly Ile Thr Gly Lys Arg Ser Gly Lys Cys
Ile 20 25 30Asn Lys Lys Cys Tyr Cys Tyr Arg 35 403437PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptideSee
specification as filed for detailed description of substitutions
and preferred embodiments 34Gly Ser Gly Cys Leu Glu Phe Trp Trp Lys
Cys Asn Pro Asn Asp Asp1 5 10 15Lys Cys Cys Arg Pro Lys Leu Lys Cys
Ser Lys Leu Phe Lys Leu Cys 20 25 30Asn Phe Ser Phe Gly
353539PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 35Gly Ser Asp Cys Val Arg Phe Trp Gly Lys Cys
Ser Gln Thr Ser Asp1 5 10 15Cys Cys Pro His Leu Ala Cys Lys Ser Lys
Trp Pro Arg Asn Ile Cys 20 25 30Val Trp Asp Gly Ser Val Gly
353635PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideSee specification as filed for detailed
description of substitutions and preferred embodiments 36Gly Ser
Gly Cys Phe Gly Tyr Lys Cys Asp Tyr Tyr Lys Gly Cys Cys1 5 10 15Ser
Gly Tyr Val Cys Ser Pro Thr Trp Lys Trp Cys Val Arg Pro Gly 20 25
30Pro Gly Arg 353762PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 37Gly Ser Met Asn Ala Lys Phe Ile
Leu Leu Leu Val Leu Thr Thr Met1 5 10 15Met Leu Leu Pro Asp Thr Lys
Gly Ala Glu Val Ile Arg Cys Ser Gly 20 25 30Ser Lys Gln Cys Tyr Gly
Pro Cys Lys Gln Gln Thr Gly Cys Thr Asn 35 40 45Ser Lys Cys Met Asn
Lys Val Cys Lys Cys Tyr Gly Cys Gly 50 55 603863PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
38Gly Ser Met Asn Ala Lys Leu Ile Tyr Leu Leu Leu Val Val Thr Thr1
5 10 15Met Thr Leu Met Phe Asp Thr Ala Gln Ala Val Asp Ile Met Cys
Ser 20 25 30Gly Pro Lys Gln Cys Tyr Gly Pro Cys Lys Lys Glu Thr Gly
Cys Pro 35 40 45Asn Ala Lys Cys Met Asn Arg Arg Cys Lys Cys Tyr Gly
Cys Val 50 55 603964PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 39Gly Ser Met Asn Ala Lys Leu Ile
Tyr Leu Leu Leu Val Val Thr Thr1 5 10 15Met Met Leu Thr Phe Asp Thr
Thr Gln Ala Gly Asp Ile Lys Cys Ser 20 25 30Gly Thr Arg Gln Cys Trp
Gly Pro Cys Lys Lys Gln Thr Thr Cys Thr 35 40 45Asn Ser Lys Cys Met
Asn Gly Lys Cys Lys Cys Tyr Gly Cys Val Gly 50 55
604063PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 40Gly Ser Met Asn Thr Lys Phe Ile Phe Leu Leu
Leu Val Val Thr Asn1 5 10 15Thr Met Met Leu Phe Asp Thr Lys Pro Val
Glu Gly Ile Ser Cys Thr 20 25 30Gly Ser Lys Gln Cys Tyr Asp Pro Cys
Lys Arg Lys Thr Gly Cys Pro 35 40 45Asn Ala Lys Cys Met Asn Lys Ser
Cys Lys Cys Tyr Gly Cys Gly 50 55 604140PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
41Gly Ser Gly Val Pro Ile Asn Val Lys Cys Ser Gly Ser Arg Asp Cys1
5 10 15Leu Glu Pro Cys Lys Lys Ala Gly Met Arg Phe Gly Lys Cys Ile
Asn 20 25 30Arg Lys Cys His Cys Thr Pro Lys 35 404240PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
42Gly Ser Gly Val Pro Ile Asn Val Lys Cys Thr Gly Ser Pro Gln Cys1
5 10 15Leu Lys Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Ile
Asn 20 25 30Gly Lys Cys His Cys Thr Pro Lys 35 404340PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
43Gly Ser Gly Val Ile Ile Asn Val Lys Cys Lys Ile Ser Arg Gln Cys1
5 10 15Leu Glu Pro Cys Lys Lys Ala Gly Met Arg Phe Gly Lys Cys Met
Asn 20 25 30Gly Lys Cys His Cys Thr Pro Lys 35 404440PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
44Gly Ser Gly Val Pro Ile Asn Val Lys Cys Arg Gly Ser Pro Gln Cys1
5 10 15Ile Gln Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Lys Cys Met
Asn 20 25 30Gly Lys Cys His Cys Thr Pro Gln 35 404540PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
45Gly Ser Gly Val Glu Ile Asn Val Lys Cys Thr Gly Ser His Gln Cys1
5 10 15Ile Lys Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Ile
Asn 20 25 30Arg Lys Cys His Cys Thr Pro Lys 35 404640PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
46Gly Ser Gly Val Glu Ile Asn Val Lys Cys Ser Gly Ser Pro Gln Cys1
5 10 15Leu Lys Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met
Asn 20 25 30Arg Lys Cys His Cys Thr Pro Lys 35 404740PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
47Gly Ser Gly Val Pro Thr Asp Val Lys Cys Arg Gly Ser Pro Gln Cys1
5 10 15Ile Gln Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met
Asn 20 25 30Gly Lys Cys His Cys Thr Pro Lys 35 404840PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
48Gly Ser Gly Val Pro Ile Asn Val Ser Cys Thr Gly Ser Pro Gln Cys1
5 10 15Ile Lys Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met
Asn 20 25 30Arg Lys Cys His Cys Thr Pro Lys 35 404940PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
49Gly Ser Gly Val Pro Ile Asn Val Pro Cys Thr Gly Ser Pro Gln Cys1
5 10 15Ile Lys Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met
Asn 20 25 30Arg Lys Cys His Cys Thr Pro Lys 35 405039PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
50Gly Ser Val Gly Ile Asn Val Lys Cys Lys His Ser Gly Gln Cys Leu1
5 10 15Lys Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Ile Asn
Gly 20 25 30Lys Cys Asp Cys Thr Pro Lys 355139PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
51Gly Ser Val Gly Ile Asn Val Lys Cys Lys His Ser Gly Gln Cys Leu1
5 10 15Lys Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn
Gly 20 25 30Lys Cys Asp Cys Thr Pro Lys 355239PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
52Gly Ser Val Gly Ile Pro Val Ser Cys Lys His Ser Gly Gln Cys Ile1
5 10 15Lys Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn
Arg 20 25 30Lys Cys Asp Cys Thr Pro Lys 355338PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
53Gly Ser Arg Lys Gly Cys Phe Lys Glu Gly His Ser Cys Pro Lys Thr1
5 10 15Ala Pro Cys Cys Arg Pro Leu Val Cys Lys Gly Pro Ser Pro Asn
Thr 20 25 30Lys Lys Cys Thr Arg Pro 355434PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
54Gly Ser Ser Phe Cys Ile Pro Phe Lys Pro Cys Lys Ser Asp Glu Asn1
5 10 15Cys Cys Lys Lys Phe Lys Cys Lys Thr Thr Gly Ile Val Lys Leu
Cys 20 25 30Arg Trp5538PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 55Gly Ser Leu Lys Gly Cys
Leu Pro Arg Asn Arg Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys
Cys Ser Gly Leu Arg Cys Lys Glu Leu Ser Ile 20 25 30Trp Ala Ser Lys
Cys Leu 355636PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 56Gly Ser Gly Asn Tyr Cys Leu Arg
Gly Arg Cys Leu Pro Gly Gly Arg1 5 10 15Lys Cys Cys Asn Gly Arg Pro
Cys Glu Cys Phe Ala Lys Ile Cys Ser 20 25 30Cys Lys Pro Lys
355731PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 57Gly Ser Thr Val Lys Cys Gly Gly Cys Asn Arg
Lys Cys Cys Pro Gly1 5 10 15Gly Cys Arg Ser Gly Lys Cys Ile Asn Gly
Lys Cys Gln Cys Tyr 20 25 305831PRTArtificial SequenceDescription
of Artificial Sequence Synthetic polypeptide 58Gly Ser Gly Cys Met
Lys Glu Tyr Cys Ala Gly Gln Cys Arg Gly Lys1 5 10 15Val Ser Gln Asp
Tyr Cys Leu Lys His Cys Lys Cys Ile Pro Arg 20 25
305936PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 59Gly Ser Ala Cys Leu Gly Phe Gly Glu Lys Cys
Asn Pro Ser Asn Asp1 5 10 15Lys Cys Cys Lys Ser Ser Ser Leu Val Cys
Ser Gln Lys His Lys Trp 20 25 30Cys Lys Tyr Gly 356039PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
60Gly Ser Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu1
5 10 15Ser Gly Pro Arg Cys Cys
Ser Gly Leu Arg Cys Lys Glu Leu Ser Ile 20 25 30Arg Asp Ser Arg Cys
Leu Gly 356138PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 61Gly Ser Arg Gly Gly Cys Leu Pro
Arg Asn Lys Phe Cys Asn Pro Ser1 5 10 15Ser Gly Pro Arg Cys Cys Ser
Gly Leu Thr Cys Lys Glu Leu Asn Ile 20 25 30Trp Ala Ser Lys Cys Leu
356235PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 62Gly Ser Gln Arg Ser Cys Ala Lys Pro Gly Asp
Met Cys Met Gly Ile1 5 10 15Lys Cys Cys Asp Gly Gln Cys Gly Cys Asn
Arg Gly Thr Gly Arg Cys 20 25 30Phe Cys Lys 356342PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
63Gly Ser Ala Arg Gly Cys Ala Asp Ala Tyr Lys Ser Cys Asn His Pro1
5 10 15Arg Thr Cys Cys Asp Gly Tyr Asn Gly Tyr Lys Arg Ala Cys Ile
Cys 20 25 30Ser Gly Ser Asn Cys Lys Cys Lys Lys Ser 35
406439PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 64Gly Ser Arg Gly Gly Cys Leu Pro His Asn Arg
Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys Cys Ser Gly Leu Arg
Cys Lys Glu Leu Ser Ile 20 25 30Trp Asp Ser Arg Cys Leu Gly
356539PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 65Gly Ser Arg Gly Gly Cys Leu Pro His Asn Arg
Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys Cys Ser Gly Leu Lys
Cys Lys Glu Leu Ser Ile 20 25 30Tyr Asp Ser Arg Cys Leu Gly
356639PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 66Gly Ser Arg Gly Gly Cys Leu Pro His Asn Arg
Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys Cys Ser Arg Leu Lys
Cys Lys Glu Leu Ser Ile 20 25 30Trp Asp Ser Arg Cys Leu Gly
356739PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 67Gly Ser Arg Gly Gly Cys Leu Pro His Asn Arg
Phe Cys Asn Ala Leu1 5 10 15Thr Gly Pro Arg Cys Cys Ser Arg Leu Arg
Cys Lys Glu Leu Ser Ile 20 25 30Trp Asp Ser Ile Cys Leu Gly
356836PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 68Gly Ser Ser Cys Ala Asp Ala Tyr Lys Ser Cys
Asp Ser Leu Lys Cys1 5 10 15Cys Asn Asn Arg Thr Cys Met Cys Ser Met
Ile Gly Thr Asn Cys Thr 20 25 30Cys Arg Lys Lys 356932PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
69Gly Ser Glu Arg Arg Cys Leu Pro Ala Gly Lys Thr Cys Val Arg Gly1
5 10 15Pro Met Arg Val Pro Cys Cys Gly Ser Cys Ser Gln Asn Lys Cys
Thr 20 25 307033PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 70Gly Ser Leu Cys Ser Arg Glu Gly
Glu Phe Cys Tyr Lys Leu Arg Lys1 5 10 15Cys Cys Ala Gly Phe Tyr Cys
Lys Ala Phe Val Leu His Cys Tyr Arg 20 25 30Asn7125PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 71Gly
Ser Ala Cys Gly Ser Cys Arg Lys Lys Cys Lys Gly Ser Gly Lys1 5 10
15Cys Ile Asn Gly Arg Cys Lys Cys Tyr 20 257225PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 72Gly
Ser Ala Cys Gly Ser Cys Arg Lys Lys Cys Lys Gly Pro Gly Lys1 5 10
15Cys Ile Asn Gly Arg Cys Lys Cys Tyr 20 257334PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
73Gly Ser Ala Cys Gln Gly Tyr Met Arg Lys Cys Gly Arg Asp Lys Pro1
5 10 15Pro Cys Cys Lys Lys Leu Glu Cys Ser Lys Thr Trp Arg Trp Cys
Val 20 25 30Trp Asn7432PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 74Gly Ser Gly Arg Tyr Cys
Gln Lys Trp Met Trp Thr Cys Asp Ser Lys1 5 10 15Arg Ala Cys Cys Glu
Gly Leu Arg Cys Lys Leu Trp Cys Arg Lys Ile 20 25
307536PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 75Gly Ser Asn Ala Lys Cys Arg Gly Ser Pro Glu
Cys Leu Pro Lys Cys1 5 10 15Lys Glu Ala Ile Gly Lys Ala Ala Gly Lys
Cys Met Asn Gly Lys Cys 20 25 30Lys Cys Tyr Pro 357636PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
76Gly Ser Asn Val Lys Cys Arg Gly Ser Lys Glu Cys Leu Pro Ala Cys1
5 10 15Lys Ala Ala Val Gly Lys Ala Ala Gly Lys Cys Met Asn Gly Lys
Cys 20 25 30Lys Cys Tyr Pro 357736PRTArtificial SequenceDescription
of Artificial Sequence Synthetic polypeptide 77Gly Ser Asn Val Lys
Cys Arg Gly Ser Pro Glu Cys Leu Pro Lys Cys1 5 10 15Lys Glu Ala Ile
Gly Lys Ser Ala Gly Lys Cys Met Asn Gly Lys Cys 20 25 30Lys Cys Tyr
Pro 357836PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 78Gly Ser Asn Ala Lys Cys Arg Gly Ser Pro Glu
Cys Leu Pro Lys Cys1 5 10 15Lys Gln Ala Ile Gly Lys Ala Ala Gly Lys
Cys Met Asn Gly Lys Cys 20 25 30Lys Cys Tyr Pro 357935PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
79Gly Ser Arg Gly Tyr Cys Ala Glu Lys Gly Ile Lys Cys His Asn Ile1
5 10 15His Cys Cys Ser Gly Leu Thr Cys Lys Cys Lys Gly Ser Ser Cys
Val 20 25 30Cys Arg Lys 358033PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 80Gly Ser Glu Arg Gly Cys
Lys Leu Thr Phe Trp Lys Cys Lys Asn Lys1 5 10 15Lys Glu Cys Cys Gly
Trp Asn Ala Cys Ala Leu Gly Ile Cys Met Pro 20 25
30Arg8141PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 81Gly Ser Lys Lys Lys Cys Ile Ala Lys Asp Tyr
Gly Arg Cys Lys Trp1 5 10 15Gly Gly Thr Pro Cys Cys Arg Gly Arg Gly
Cys Ile Cys Ser Ile Met 20 25 30Gly Thr Asn Cys Glu Cys Lys Pro Arg
35 408231PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 82Gly Ser Gly Cys Lys Leu Thr Phe Trp Lys Cys
Lys Asn Lys Lys Glu1 5 10 15Cys Cys Gly Trp Asn Ala Cys Ala Leu Gly
Ile Cys Met Pro Arg 20 25 308334PRTArtificial SequenceDescription
of Artificial Sequence Synthetic polypeptide 83Gly Ser Ala Cys Lys
Gly Leu Phe Val Thr Cys Thr Pro Gly Lys Asp1 5 10 15Glu Cys Cys Pro
Asn His Val Cys Ser Ser Lys His Lys Trp Cys Lys 20 25 30Tyr
Lys8438PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 84Gly Ser Ile Ala Cys Ala Pro Arg Gly Leu Leu
Cys Phe Arg Asp Lys1 5 10 15Glu Cys Cys Lys Gly Leu Thr Cys Lys Gly
Arg Phe Val Asn Thr Trp 20 25 30Pro Thr Phe Cys Leu Val
358538PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 85Gly Ser Ala Cys Ala Gly Leu Tyr Lys Lys Cys
Gly Lys Gly Val Asn1 5 10 15Thr Cys Cys Glu Asn Arg Pro Cys Lys Cys
Asp Leu Ala Met Gly Asn 20 25 30Cys Ile Cys Lys Lys Lys
358640PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 86Gly Ser Phe Thr Cys Ala Ile Ser Cys Asp Ile
Lys Val Asn Gly Lys1 5 10 15Pro Cys Lys Gly Ser Gly Glu Lys Lys Cys
Ser Gly Gly Trp Ser Cys 20 25 30Lys Phe Asn Val Cys Val Lys Val 35
408735PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 87Gly Ser Gly Phe Cys Ala Gln Lys Gly Ile Lys
Cys His Asp Ile His1 5 10 15Cys Cys Thr Asn Leu Lys Cys Val Arg Glu
Gly Ser Asn Arg Val Cys 20 25 30Arg Lys Ala 358844PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
88Gly Ser Cys Ala Lys Lys Arg Asn Trp Cys Gly Lys Asn Glu Asp Cys1
5 10 15Cys Cys Pro Met Lys Cys Ile Tyr Ala Trp Tyr Asn Gln Gln Gly
Ser 20 25 30Cys Gln Ser Thr Ile Thr Gly Leu Phe Lys Lys Cys 35
408930PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 89Gly Ser Tyr Cys Gln Lys Trp Met Trp Thr Cys
Asp Ser Ala Arg Lys1 5 10 15Cys Cys Glu Gly Leu Val Cys Arg Leu Trp
Cys Lys Lys Ile 20 25 309039PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 90Gly Ser Arg Gly Gly Cys
Leu Pro His Asn Lys Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys
Cys Ser Gly Leu Lys Cys Lys Glu Leu Thr Ile 20 25 30Trp Asn Thr Lys
Cys Leu Glu 359136PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 91Gly Ser Asn Val Lys Cys Thr Gly
Ser Lys Gln Cys Leu Pro Ala Cys1 5 10 15Lys Ala Ala Val Gly Lys Ala
Ala Gly Lys Cys Met Asn Gly Lys Cys 20 25 30Lys Cys Tyr Thr
359235PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 92Gly Ser Gln Arg Ser Cys Ala Lys Pro Gly Glu
Met Cys Met Arg Ile1 5 10 15Lys Cys Cys Asp Gly Gln Cys Gly Cys Asn
Arg Gly Thr Gly Arg Cys 20 25 30Phe Cys Lys 359338PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
93Gly Ser Gly Cys Ile Pro Lys His Lys Arg Cys Thr Trp Ser Gly Pro1
5 10 15Lys Cys Cys Asn Asn Ile Ser Cys His Cys Asn Ile Ser Gly Thr
Leu 20 25 30Cys Lys Cys Arg Pro Gly 359435PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
94Gly Ser Asn Tyr Cys Val Ala Lys Arg Cys Arg Pro Gly Gly Arg Gln1
5 10 15Cys Cys Ser Gly Lys Pro Cys Ala Cys Val Gly Lys Val Cys Lys
Cys 20 25 30Pro Arg Asp 359539PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 95Gly Ser Glu Arg Gly Cys
Ser Gly Ala Tyr Lys Arg Cys Ser Ser Ser1 5 10 15Gln Arg Cys Cys Glu
Gly Arg Pro Cys Val Cys Ser Ala Ile Asn Ser 20 25 30Asn Cys Lys Cys
Arg Lys Thr 359638PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 96Gly Ser Arg Tyr Cys Pro Arg Asn
Pro Glu Ala Cys Tyr Asn Tyr Cys1 5 10 15Leu Arg Thr Gly Arg Pro Gly
Gly Tyr Cys Gly Gly Arg Ser Arg Ile 20 25 30Thr Cys Phe Cys Phe Arg
359735PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 97Gly Ser Gln Arg Ser Cys Ala Lys Pro Gly Glu
Met Cys Met Gly Ile1 5 10 15Lys Cys Cys Asp Gly Gln Cys Gly Cys Asn
Arg Gly Thr Gly Arg Cys 20 25 30Phe Cys Lys 359838PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
98Gly Ser Arg Arg Gly Cys Phe Lys Glu Gly Lys Trp Cys Pro Lys Ser1
5 10 15Ala Pro Cys Cys Ala Pro Leu Lys Cys Lys Gly Pro Ser Ile Lys
Gln 20 25 30Gln Lys Cys Val Arg Glu 359933PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
99Gly Ser Thr Val Lys Cys Gly Gly Cys Asn Arg Lys Cys Cys Ala Gly1
5 10 15Gly Cys Arg Ser Gly Lys Cys Ile Asn Gly Lys Cys Gln Cys Tyr
Gly 20 25 30Arg10031PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 100Gly Ser Glu Arg Arg Cys Glu Pro
Ser Gly Lys Pro Cys Arg Pro Leu1 5 10 15Met Arg Ile Pro Cys Cys Gly
Ser Cys Val Arg Gly Lys Cys Ala 20 25 3010138PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
101Gly Ser Arg Gly Gly Cys Leu Pro Arg Asn Lys Phe Cys Asn Pro Ser1
5 10 15Ser Gly Pro Arg Cys Cys Ser Gly Leu Thr Cys Lys Glu Leu Asn
Ile 20 25 30Trp Ala Asn Lys Cys Leu 3510244PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
102Gly Ser Cys Ala Lys Lys Arg Asn Trp Cys Gly Lys Asn Glu Asp Cys1
5 10 15Cys Cys Pro Met Lys Cys Ile Tyr Ala Trp Tyr Asn Gln Gln Gly
Ser 20 25 30Cys Gln Thr Thr Ile Thr Gly Leu Phe Lys Lys Cys 35
4010339PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 103Gly Ser Val Arg Ile Pro Val Ser Cys Lys
His Ser Gly Gln Cys Leu1 5 10 15Lys Pro Cys Lys Asp Ala Gly Met Arg
Thr Gly Lys Cys Met Asn Gly 20 25 30Lys Cys Asp Cys Thr Pro Lys
3510422PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 104Gly Ser Val Lys Cys Thr Thr Ser Lys Asp Cys
Trp Pro Pro Cys Lys1 5 10 15Lys Val Thr Gly Arg Ala
2010534PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 105Gly Ser Gly Ile Val Cys Arg Val Cys Arg
Ile Ile Cys Gly Met Gln1 5 10 15Gly Arg Arg Val Asn Ile Cys Arg Ala
Pro Ile Arg Cys Arg Cys Arg 20 25 30Arg Gly10637PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
106Gly Ser Ser Glu Arg Asp Cys Ile Arg His Leu Gln Arg Cys Arg Glu1
5 10 15Asn Arg Asp Cys Cys Ser Arg Arg Cys Ser Arg Arg Gly Thr Asn
Pro 20 25 30Glu Arg Arg Cys Arg 3510739PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
107Gly Ser Val Arg Ile Pro Val Ser Cys Arg His Ser Gly Gln Cys Leu1
5 10 15Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn
Gly 20 25 30Arg Cys Asp Cys Thr Pro Arg 3510839PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
108Gly Ser Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys1
5 10 15Leu Asp Pro Cys Arg Arg Ala Gly Met Arg Phe Gly Arg Cys Ile
Asn 20 25 30Ser Arg Cys His Cys Thr Pro 3510935PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
109Gly Ser Ala Val Cys Val Tyr Arg Thr Cys Asp Arg Asp Cys Arg Arg1
5 10 15Arg Gly Tyr Arg Ser Gly Arg Cys Ile Asn Asn Ala Cys Arg Cys
Tyr 20 25 30Pro Tyr Gly 3511037PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 110Gly Ser Ile Ser Cys
Thr Gly Ser Arg Gln Cys Tyr Asp Pro Cys Arg1 5 10 15Arg Arg Thr Gly
Cys Pro Asn Ala Arg Cys Met Asn Arg Ser Cys Arg 20 25 30Cys Tyr Gly
Cys Gly 3511140PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 111Gly Ser Gln Val Gln Thr Asn Val
Arg Cys Gln Gly Gly Ser Cys Ala1 5 10 15Ser Val Cys Arg Arg Glu Ile
Gly Val Ala Ala Gly Arg Cys Ile Asn 20 25 30Gly Arg Cys Val Cys Tyr
Arg Asn 35 4011239PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 112Gly Ser Glu Val Ile
Arg Cys Ser Gly Ser Arg Gln Cys Tyr Gly Pro1 5 10 15Cys Arg Gln Gln
Thr Gly Cys Thr Asn Ser Arg Cys Met Asn Arg Val 20 25 30Cys Arg Cys
Tyr Gly Cys Gly 3511335PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 113Gly Ser Ala Cys Arg
Gly Val Phe Asp Ala Cys Thr Pro Gly Arg Asn1 5 10 15Glu Cys Cys Pro
Asn Arg Val Cys Ser Asp Arg His Arg Trp Cys Arg 20 25 30Trp Arg Leu
3511440PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 114Gly Ser Gln Ile Tyr Thr Ser Arg Glu Cys
Asn Gly Ser Ser Glu Cys1 5 10 15Tyr Ser His Cys Glu Gly Ile Thr Gly
Arg Arg Ser Gly Arg Cys Ile 20 25 30Asn Arg Arg Cys Tyr Cys Tyr Arg
35 4011537PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 115Gly Ser Gly Cys Leu Glu Phe Trp Trp Arg
Cys Asn Pro Asn Asp Asp1 5 10 15Arg Cys Cys Arg Pro Arg Leu Arg Cys
Ser Arg Leu Phe Arg Leu Cys 20 25 30Asn Phe Ser Phe Gly
3511639PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 116Gly Ser Asp Cys Val Arg Phe Trp Gly Arg
Cys Ser Gln Thr Ser Asp1 5 10 15Cys Cys Pro His Leu Ala Cys Arg Ser
Arg Trp Pro Arg Asn Ile Cys 20 25 30Val Trp Asp Gly Ser Val Gly
3511735PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 117Gly Ser Gly Cys Phe Gly Tyr Arg Cys Asp
Tyr Tyr Arg Gly Cys Cys1 5 10 15Ser Gly Tyr Val Cys Ser Pro Thr Trp
Arg Trp Cys Val Arg Pro Gly 20 25 30Pro Gly Arg
3511862PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 118Gly Ser Met Asn Ala Arg Phe Ile Leu Leu
Leu Val Leu Thr Thr Met1 5 10 15Met Leu Leu Pro Asp Thr Arg Gly Ala
Glu Val Ile Arg Cys Ser Gly 20 25 30Ser Arg Gln Cys Tyr Gly Pro Cys
Arg Gln Gln Thr Gly Cys Thr Asn 35 40 45Ser Arg Cys Met Asn Arg Val
Cys Arg Cys Tyr Gly Cys Gly 50 55 6011963PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
119Gly Ser Met Asn Ala Arg Leu Ile Tyr Leu Leu Leu Val Val Thr Thr1
5 10 15Met Thr Leu Met Phe Asp Thr Ala Gln Ala Val Asp Ile Met Cys
Ser 20 25 30Gly Pro Arg Gln Cys Tyr Gly Pro Cys Arg Arg Glu Thr Gly
Cys Pro 35 40 45Asn Ala Arg Cys Met Asn Arg Arg Cys Arg Cys Tyr Gly
Cys Val 50 55 6012064PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 120Gly Ser Met Asn Ala
Arg Leu Ile Tyr Leu Leu Leu Val Val Thr Thr1 5 10 15Met Met Leu Thr
Phe Asp Thr Thr Gln Ala Gly Asp Ile Arg Cys Ser 20 25 30Gly Thr Arg
Gln Cys Trp Gly Pro Cys Arg Arg Gln Thr Thr Cys Thr 35 40 45Asn Ser
Arg Cys Met Asn Gly Arg Cys Arg Cys Tyr Gly Cys Val Gly 50 55
6012163PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 121Gly Ser Met Asn Thr Arg Phe Ile Phe Leu
Leu Leu Val Val Thr Asn1 5 10 15Thr Met Met Leu Phe Asp Thr Arg Pro
Val Glu Gly Ile Ser Cys Thr 20 25 30Gly Ser Arg Gln Cys Tyr Asp Pro
Cys Arg Arg Arg Thr Gly Cys Pro 35 40 45Asn Ala Arg Cys Met Asn Arg
Ser Cys Arg Cys Tyr Gly Cys Gly 50 55 6012240PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
122Gly Ser Gly Val Pro Ile Asn Val Arg Cys Ser Gly Ser Arg Asp Cys1
5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Met Arg Phe Gly Arg Cys Ile
Asn 20 25 30Arg Arg Cys His Cys Thr Pro Arg 35 4012340PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
123Gly Ser Gly Val Pro Ile Asn Val Arg Cys Thr Gly Ser Pro Gln Cys1
5 10 15Leu Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Ile
Asn 20 25 30Gly Arg Cys His Cys Thr Pro Arg 35 4012440PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
124Gly Ser Gly Val Ile Ile Asn Val Arg Cys Arg Ile Ser Arg Gln Cys1
5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Met Arg Phe Gly Arg Cys Met
Asn 20 25 30Gly Arg Cys His Cys Thr Pro Arg 35 4012540PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
125Gly Ser Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Pro Gln Cys1
5 10 15Ile Gln Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met
Asn 20 25 30Gly Arg Cys His Cys Thr Pro Gln 35 4012640PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
126Gly Ser Gly Val Glu Ile Asn Val Arg Cys Thr Gly Ser His Gln Cys1
5 10 15Ile Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Ile
Asn 20 25 30Arg Arg Cys His Cys Thr Pro Arg 35 4012740PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
127Gly Ser Gly Val Glu Ile Asn Val Arg Cys Ser Gly Ser Pro Gln Cys1
5 10 15Leu Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met
Asn 20 25 30Arg Arg Cys His Cys Thr Pro Arg 35 4012840PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
128Gly Ser Gly Val Pro Thr Asp Val Arg Cys Arg Gly Ser Pro Gln Cys1
5 10 15Ile Gln Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met
Asn 20 25 30Gly Arg Cys His Cys Thr Pro Arg 35 4012940PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
129Gly Ser Gly Val Pro Ile Asn Val Ser Cys Thr Gly Ser Pro Gln Cys1
5 10 15Ile Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met
Asn 20 25 30Arg Arg Cys His Cys Thr Pro Arg 35 4013040PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
130Gly Ser Gly Val Pro Ile Asn Val Pro Cys Thr Gly Ser Pro Gln Cys1
5 10 15Ile Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met
Asn 20 25 30Arg Arg Cys His Cys Thr Pro Arg 35 4013139PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
131Gly Ser Val Gly Ile Asn Val Arg Cys Arg His Ser Gly Gln Cys Leu1
5 10 15Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Ile Asn
Gly 20 25 30Arg Cys Asp Cys Thr Pro Arg 3513239PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
132Gly Ser Val Gly Ile Asn Val Arg Cys Arg His Ser Gly Gln Cys Leu1
5 10 15Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn
Gly 20 25 30Arg Cys Asp Cys Thr Pro Arg 3513339PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
133Gly Ser Val Gly Ile Pro Val Ser Cys Arg His Ser Gly Gln Cys Ile1
5 10 15Arg Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn
Arg 20 25 30Arg Cys Asp Cys Thr Pro Arg 3513438PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
134Gly Ser Arg Arg Gly Cys Phe Arg Glu Gly His Ser Cys Pro Arg Thr1
5 10 15Ala Pro Cys Cys Arg Pro Leu Val Cys Arg Gly Pro Ser Pro Asn
Thr 20 25 30Arg Arg Cys Thr Arg Pro 3513534PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
135Gly Ser Ser Phe Cys Ile Pro Phe Arg Pro Cys Arg Ser Asp Glu Asn1
5 10 15Cys Cys Arg Arg Phe Arg Cys Arg Thr Thr Gly Ile Val Arg Leu
Cys 20 25 30Arg Trp13638PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 136Gly Ser Leu Arg Gly
Cys Leu Pro Arg Asn Arg Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg
Cys Cys Ser Gly Leu Arg Cys Arg Glu Leu Ser Ile 20 25 30Trp Ala Ser
Arg Cys Leu 3513736PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 137Gly Ser Gly Asn Tyr Cys Leu Arg
Gly Arg Cys Leu Pro Gly Gly Arg1 5 10 15Arg Cys Cys Asn Gly Arg Pro
Cys Glu Cys Phe Ala Arg Ile Cys Ser 20 25 30Cys Arg Pro Arg
3513831PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 138Gly Ser Thr Val Arg Cys Gly Gly Cys Asn
Arg Arg Cys Cys Pro Gly1 5 10 15Gly Cys Arg Ser Gly Arg Cys Ile Asn
Gly Arg Cys Gln Cys Tyr 20 25 3013931PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
139Gly Ser Gly Cys Met Arg Glu Tyr Cys Ala Gly Gln Cys Arg Gly Arg1
5 10 15Val Ser Gln Asp Tyr Cys Leu Arg His Cys Arg Cys Ile Pro Arg
20 25 3014036PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 140Gly Ser Ala Cys Leu Gly Phe Gly
Glu Arg Cys Asn Pro Ser Asn Asp1 5 10 15Arg Cys Cys Arg Ser Ser Ser
Leu Val Cys Ser Gln Arg His Arg Trp 20 25 30Cys Arg Tyr Gly
3514139PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 141Gly Ser Arg Gly Gly Cys Leu Pro His Asn
Arg Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys Cys Ser Gly Leu
Arg Cys Arg Glu Leu Ser Ile 20 25 30Arg Asp Ser Arg Cys Leu Gly
3514238PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 142Gly Ser Arg Gly Gly Cys Leu Pro Arg Asn
Arg Phe Cys Asn Pro Ser1 5 10 15Ser Gly Pro Arg Cys Cys Ser Gly Leu
Thr Cys Arg Glu Leu Asn Ile 20 25 30Trp Ala Ser Arg Cys Leu
3514335PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 143Gly Ser Gln Arg Ser Cys Ala Arg Pro Gly
Asp Met Cys Met Gly Ile1 5 10 15Arg Cys Cys Asp Gly Gln Cys Gly Cys
Asn Arg Gly Thr Gly Arg Cys 20 25 30Phe Cys Arg
3514442PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 144Gly Ser Ala Arg Gly Cys Ala Asp Ala Tyr
Arg Ser Cys Asn His Pro1 5 10 15Arg Thr Cys Cys Asp Gly Tyr Asn Gly
Tyr Arg Arg Ala Cys Ile Cys 20 25 30Ser Gly Ser Asn Cys Arg Cys Arg
Arg Ser 35 4014539PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 145Gly Ser Arg Gly Gly Cys Leu Pro
His Asn Arg Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys Cys Ser
Gly Leu Arg Cys Arg Glu Leu Ser Ile 20 25 30Trp Asp Ser Arg Cys Leu
Gly 3514639PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 146Gly Ser Arg Gly Gly Cys Leu Pro His Asn
Arg Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys Cys Ser Gly Leu
Arg Cys Arg Glu Leu Ser Ile 20 25 30Tyr Asp Ser Arg Cys Leu Gly
3514739PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 147Gly Ser Arg Gly Gly Cys Leu Pro His Asn
Arg Phe Cys Asn Ala Leu1 5 10 15Ser Gly Pro Arg Cys Cys Ser Arg Leu
Arg Cys Arg Glu Leu Ser Ile 20 25 30Trp Asp Ser Arg Cys Leu Gly
3514839PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 148Gly Ser Arg Gly Gly Cys Leu Pro His Asn
Arg Phe Cys Asn Ala Leu1 5 10 15Thr Gly Pro Arg Cys Cys Ser Arg Leu
Arg Cys Arg Glu Leu Ser Ile 20 25 30Trp Asp Ser Ile Cys Leu Gly
3514936PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 149Gly Ser Ser Cys Ala Asp Ala Tyr Lys Ser
Cys Asp Ser Leu Arg Cys1 5 10 15Cys Asn Asn Arg Thr Cys Met Cys Ser
Met Ile Gly Thr Asn Cys Thr 20 25 30Cys Arg Arg Arg
3515032PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 150Gly Ser Glu Arg Arg Cys Leu Pro Ala Gly
Arg Thr Cys Val Arg Gly1 5 10 15Pro Met Arg Val Pro Cys Cys Gly Ser
Cys Ser Gln Asn Arg Cys Thr 20 25 3015133PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
151Gly Ser Leu Cys Ser Arg Glu Gly Glu Phe Cys Tyr Arg Leu Arg Arg1
5 10 15Cys Cys Ala Gly Phe Tyr Cys Arg Ala Phe Val Leu His Cys Tyr
Arg 20 25 30Asn15225PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 152Gly Ser Ala Cys Gly Ser Cys Arg Arg
Arg Cys Arg Gly Ser Gly Arg1 5 10 15Cys Ile Asn Gly Arg Cys Arg Cys
Tyr 20 2515325PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 153Gly Ser Ala Cys Gly Ser Cys Arg Arg
Arg Cys Arg Gly Pro Gly Arg1 5 10 15Cys Ile Asn Gly Arg Cys Arg Cys
Tyr 20 2515434PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 154Gly Ser Ala Cys Gln Gly Tyr Met
Arg Arg Cys Gly Arg Asp Arg Pro1 5 10 15Pro Cys Cys Arg Arg Leu Glu
Cys Ser Arg Thr Trp Arg Trp Cys Val 20 25 30Trp
Asn15532PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 155Gly Ser Gly Arg Tyr Cys Gln Arg Trp Met
Trp Thr Cys Asp Ser Arg1 5 10 15Arg Ala Cys Cys Glu Gly Leu Arg Cys
Arg Leu Trp Cys Arg Arg Ile 20 25 3015636PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
156Gly Ser Asn Ala Arg Cys Arg Gly Ser Pro Glu Cys Leu Pro Arg Cys1
5 10 15Arg Glu Ala Ile Gly Arg Ala Ala Gly Arg Cys Met Asn Gly Arg
Cys 20 25 30Arg Cys Tyr Pro 3515736PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
157Gly Ser Asn Val Arg Cys Arg Gly Ser Arg Glu Cys Leu Pro Ala Cys1
5 10 15Arg Ala Ala Val Gly Arg Ala Ala Gly Arg Cys Met Asn Gly Arg
Cys 20 25 30Arg Cys Tyr Pro 3515836PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
158Gly Ser Asn Val Arg Cys Arg Gly Ser Pro Glu Cys Leu Pro Arg Cys1
5 10 15Arg Glu Ala Ile Gly Arg Ser Ala Gly Arg Cys Met Asn Gly Arg
Cys 20 25 30Arg Cys Tyr Pro 3515936PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
159Gly Ser Asn Ala Arg Cys Arg Gly Ser Pro Glu Cys Leu Pro Arg Cys1
5 10 15Arg Gln Ala Ile Gly Arg Ala Ala Gly Arg Cys Met Asn Gly Arg
Cys 20 25 30Arg Cys Tyr
Pro 3516035PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 160Gly Ser Arg Gly Tyr Cys Ala Glu Arg Gly
Ile Arg Cys His Asn Ile1 5 10 15His Cys Cys Ser Gly Leu Thr Cys Arg
Cys Arg Gly Ser Ser Cys Val 20 25 30Cys Arg Arg
3516133PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 161Gly Ser Glu Arg Gly Cys Arg Leu Thr Phe
Trp Arg Cys Arg Asn Arg1 5 10 15Arg Glu Cys Cys Gly Trp Asn Ala Cys
Ala Leu Gly Ile Cys Met Pro 20 25 30Arg16241PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
162Gly Ser Arg Arg Arg Cys Ile Ala Arg Asp Tyr Gly Arg Cys Arg Trp1
5 10 15Gly Gly Thr Pro Cys Cys Arg Gly Arg Gly Cys Ile Cys Ser Ile
Met 20 25 30Gly Thr Asn Cys Glu Cys Arg Pro Arg 35
4016331PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 163Gly Ser Gly Cys Arg Leu Thr Phe Trp Arg
Cys Arg Asn Arg Arg Glu1 5 10 15Cys Cys Gly Trp Asn Ala Cys Ala Leu
Gly Ile Cys Met Pro Arg 20 25 3016434PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
164Gly Ser Ala Cys Arg Gly Leu Phe Val Thr Cys Thr Pro Gly Arg Asp1
5 10 15Glu Cys Cys Pro Asn His Val Cys Ser Ser Arg His Arg Trp Cys
Arg 20 25 30Tyr Arg16538PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 165Gly Ser Ile Ala Cys
Ala Pro Arg Gly Leu Leu Cys Phe Arg Asp Arg1 5 10 15Glu Cys Cys Arg
Gly Leu Thr Cys Arg Gly Arg Phe Val Asn Thr Trp 20 25 30Pro Thr Phe
Cys Leu Val 3516638PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 166Gly Ser Ala Cys Ala Gly Leu Tyr
Arg Arg Cys Gly Arg Gly Val Asn1 5 10 15Thr Cys Cys Glu Asn Arg Pro
Cys Arg Cys Asp Leu Ala Met Gly Asn 20 25 30Cys Ile Cys Arg Arg Arg
3516740PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 167Gly Ser Phe Thr Cys Ala Ile Ser Cys Asp
Ile Arg Val Asn Gly Arg1 5 10 15Pro Cys Arg Gly Ser Gly Glu Arg Arg
Cys Ser Gly Gly Trp Ser Cys 20 25 30Arg Phe Asn Val Cys Val Arg Val
35 4016835PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 168Gly Ser Gly Phe Cys Ala Gln Arg Gly Ile
Arg Cys His Asp Ile His1 5 10 15Cys Cys Thr Asn Leu Arg Cys Val Arg
Glu Gly Ser Asn Arg Val Cys 20 25 30Arg Arg Ala
3516944PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 169Gly Ser Cys Ala Arg Arg Arg Asn Trp Cys
Gly Arg Asn Glu Asp Cys1 5 10 15Cys Cys Pro Met Arg Cys Ile Tyr Ala
Trp Tyr Asn Gln Gln Gly Ser 20 25 30Cys Gln Ser Thr Ile Thr Gly Leu
Phe Arg Arg Cys 35 4017030PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 170Gly Ser Tyr Cys Gln
Arg Trp Met Trp Thr Cys Asp Ser Ala Arg Arg1 5 10 15Cys Cys Glu Gly
Leu Val Cys Arg Leu Trp Cys Arg Arg Ile 20 25 3017139PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
171Gly Ser Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu1
5 10 15Ser Gly Pro Arg Cys Cys Ser Gly Leu Arg Cys Arg Glu Leu Thr
Ile 20 25 30Trp Asn Thr Arg Cys Leu Glu 3517236PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
172Gly Ser Asn Val Arg Cys Thr Gly Ser Arg Gln Cys Leu Pro Ala Cys1
5 10 15Arg Ala Ala Val Gly Arg Ala Ala Gly Arg Cys Met Asn Gly Arg
Cys 20 25 30Arg Cys Tyr Thr 3517335PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
173Gly Ser Gln Arg Ser Cys Ala Arg Pro Gly Glu Met Cys Met Arg Ile1
5 10 15Arg Cys Cys Asp Gly Gln Cys Gly Cys Asn Arg Gly Thr Gly Arg
Cys 20 25 30Phe Cys Arg 3517438PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 174Gly Ser Gly Cys Ile
Pro Arg His Arg Arg Cys Thr Trp Ser Gly Pro1 5 10 15Arg Cys Cys Asn
Asn Ile Ser Cys His Cys Asn Ile Ser Gly Thr Leu 20 25 30Cys Arg Cys
Arg Pro Gly 3517535PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 175Gly Ser Asn Tyr Cys Val Ala Arg
Arg Cys Arg Pro Gly Gly Arg Gln1 5 10 15Cys Cys Ser Gly Arg Pro Cys
Ala Cys Val Gly Arg Val Cys Arg Cys 20 25 30Pro Arg Asp
3517639PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 176Gly Ser Glu Arg Gly Cys Ser Gly Ala Tyr
Arg Arg Cys Ser Ser Ser1 5 10 15Gln Arg Cys Cys Glu Gly Arg Pro Cys
Val Cys Ser Ala Ile Asn Ser 20 25 30Asn Cys Arg Cys Arg Arg Thr
3517735PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 177Gly Ser Gln Arg Ser Cys Ala Arg Pro Gly
Glu Met Cys Met Gly Ile1 5 10 15Arg Cys Cys Asp Gly Gln Cys Gly Cys
Asn Arg Gly Thr Gly Arg Cys 20 25 30Phe Cys Arg
3517838PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 178Gly Ser Arg Arg Gly Cys Phe Arg Glu Gly
Arg Trp Cys Pro Arg Ser1 5 10 15Ala Pro Cys Cys Ala Pro Leu Arg Cys
Arg Gly Pro Ser Ile Arg Gln 20 25 30Gln Arg Cys Val Arg Glu
3517933PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 179Gly Ser Thr Val Arg Cys Gly Gly Cys Asn
Arg Arg Cys Cys Ala Gly1 5 10 15Gly Cys Arg Ser Gly Arg Cys Ile Asn
Gly Arg Cys Gln Cys Tyr Gly 20 25 30Arg18031PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
180Gly Ser Glu Arg Arg Cys Glu Pro Ser Gly Arg Pro Cys Arg Pro Leu1
5 10 15Met Arg Ile Pro Cys Cys Gly Ser Cys Val Arg Gly Arg Cys Ala
20 25 3018138PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 181Gly Ser Arg Gly Gly Cys Leu Pro
Arg Asn Arg Phe Cys Asn Pro Ser1 5 10 15Ser Gly Pro Arg Cys Cys Ser
Gly Leu Thr Cys Arg Glu Leu Asn Ile 20 25 30Trp Ala Asn Arg Cys Leu
3518244PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 182Gly Ser Cys Ala Arg Arg Arg Asn Trp Cys
Gly Arg Asn Glu Asp Cys1 5 10 15Cys Cys Pro Met Arg Cys Ile Tyr Ala
Trp Tyr Asn Gln Gln Gly Ser 20 25 30Cys Gln Thr Thr Ile Thr Gly Leu
Phe Arg Arg Cys 35 4018339PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 183Gly Ser Val Arg Ile
Pro Val Ser Cys Arg His Ser Gly Gln Cys Leu1 5 10 15Arg Pro Cys Arg
Asp Ala Gly Met Arg Thr Gly Arg Cys Met Asn Gly 20 25 30Arg Cys Asp
Cys Thr Pro Arg 3518440PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 184Gly Ser Gln Lys Ile
Leu Ser Asn Arg Cys Asn Asn Ser Ser Glu Cys1 5 10 15Ile Pro His Cys
Ile Arg Ile Phe Gly Thr Arg Ala Ala Lys Cys Ile 20 25 30Asn Arg Lys
Cys Tyr Cys Tyr Pro 35 4018534PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 185Gly Ser Ala Val Cys
Asn Leu Lys Arg Cys Gln Leu Ser Cys Arg Ser1 5 10 15Leu Gly Leu Leu
Gly Lys Cys Ile Gly Asp Lys Cys Glu Cys Val Lys 20 25 30His
Gly18639PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 186Gly Ser Ile Ser Ile Gly Ile Arg Cys Ser
Pro Ser Ile Asp Leu Cys1 5 10 15Glu Gly Gln Cys Arg Ile Arg Arg Tyr
Phe Thr Gly Tyr Cys Ser Gly 20 25 30Asp Thr Cys His Cys Ser Gly
3518735PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 187Gly Ser Gly Asp Cys Leu Pro His Leu Arg
Arg Cys Arg Glu Asn Asn1 5 10 15Asp Cys Cys Ser Arg Arg Cys Arg Arg
Arg Gly Ala Asn Pro Glu Arg 20 25 30Arg Cys Arg
3518836PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 188Gly Ser Ser Cys Glu Pro Gly Arg Thr Phe
Arg Asp Arg Cys Asn Thr1 5 10 15Cys Lys Cys Gly Ala Asp Gly Arg Ser
Ala Ala Cys Thr Leu Arg Ala 20 25 30Cys Pro Asn Gln
3518935PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 189Gly Ser Gly Asp Cys Leu Pro His Leu Lys
Arg Cys Lys Ala Asp Asn1 5 10 15Asp Cys Cys Gly Lys Lys Cys Lys Arg
Arg Gly Thr Asn Ala Glu Lys 20 25 30Arg Cys Arg
3519035PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 190Gly Ser Gly Asp Cys Leu Pro His Leu Lys
Arg Cys Lys Glu Asn Asn1 5 10 15Asp Cys Cys Ser Lys Lys Cys Lys Arg
Arg Gly Thr Asn Pro Glu Lys 20 25 30Arg Cys Arg
3519135PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 191Gly Ser Lys Asp Cys Leu Lys Lys Leu Lys
Leu Cys Lys Glu Asn Lys1 5 10 15Asp Cys Cys Ser Lys Ser Cys Lys Arg
Arg Gly Thr Asn Ile Glu Lys 20 25 30Arg Cys Arg
3519235PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 192Gly Ser Gly Asp Cys Leu Pro His Leu Lys
Arg Cys Lys Glu Asn Asn1 5 10 15Asp Cys Cys Ser Lys Lys Cys Lys Arg
Arg Gly Ala Asn Pro Glu Lys 20 25 30Arg Cys Arg
3519339PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 193Gly Ser Val Phe Ile Asn Val Lys Cys Arg
Gly Ser Pro Glu Cys Leu1 5 10 15Pro Lys Cys Lys Glu Ala Ile Gly Lys
Ser Ala Gly Lys Cys Met Asn 20 25 30Gly Lys Cys Lys Cys Tyr Pro
3519439PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 194Gly Ser Val Phe Ile Asn Ala Lys Cys Arg
Gly Ser Pro Glu Cys Leu1 5 10 15Pro Lys Cys Lys Glu Ala Ile Gly Lys
Ala Ala Gly Lys Cys Met Asn 20 25 30Gly Lys Cys Lys Cys Tyr Pro
3519538PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 195Gly Ser Val Ile Ile Asn Val Lys Cys Lys
Ile Ser Arg Gln Cys Leu1 5 10 15Glu Pro Cys Lys Lys Ala Gly Met Arg
Phe Gly Lys Cys Met Asn Gly 20 25 30Lys Cys His Cys Thr Pro
3519638PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 196Gly Ser Val Pro Thr Asp Val Lys Cys Arg
Gly Ser Pro Gln Cys Ile1 5 10 15Gln Pro Cys Lys Asp Ala Gly Met Arg
Phe Gly Lys Cys Met Asn Gly 20 25 30Lys Cys His Cys Thr Pro
3519738PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 197Gly Ser Val Arg Ile Pro Val Ser Cys Lys
His Ser Gly Gln Cys Leu1 5 10 15Lys Pro Cys Lys Asp Ala Gly Met Arg
Phe Gly Lys Cys Met Asn Gly 20 25 30Lys Cys Asp Cys Thr Pro
3519838PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 198Gly Ser Val Arg Ile Pro Val Ser Cys Arg
His Ser Gly Gln Cys Leu1 5 10 15Arg Pro Cys Arg Asp Ala Gly Met Arg
Phe Gly Arg Cys Met Asn Gly 20 25 30Arg Cys Asp Cys Thr Pro
3519935PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 199Gly Ser Thr Asn Val Ser Cys Thr Thr Ser
Lys Glu Cys Trp Ser Val1 5 10 15Cys Gln Arg Leu His Asn Thr Ser Arg
Gly Lys Cys Met Asn Lys Lys 20 25 30Cys Arg Cys
3520034PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 200Gly Ser Asn Val Lys Cys Thr Gly Ser Lys
Gln Cys Leu Pro Ala Cys1 5 10 15Lys Ala Ala Val Gly Lys Ala Ala Gly
Lys Cys Met Asn Gly Lys Cys 20 25 30Lys Cys20139PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
201Gly Ser Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys1
5 10 15Leu Asp Pro Cys Arg Gly Ala Gly Glu Arg His Gly Arg Cys Gly
Asn 20 25 30Ser Arg Cys His Cys Thr Pro 3520239PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
202Gly Ser Val Arg Ile Pro Val Ser Cys Arg His Ser Gly Gln Cys Leu1
5 10 15Arg Pro Cys Arg Asp Ala Gly Glu Arg His Gly Arg Cys Gly Gly
Gly 20 25 30Arg Cys Asp Cys Thr Pro Arg 3520340PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
203Gly Ser Gln Val Gln Thr Asn Val Arg Cys Gln Gly Gly Ser Cys Gly1
5 10 15Ser Val Cys Arg Arg Glu Gly Gly Gly Ala Gly Gly Gly Cys Gly
Asn 20 25 30Gly Arg Cys Gly Cys Tyr Arg Asn 35 4020434PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
204Gly Ser Ile Lys Cys Ser Glu Ser Tyr Gln Cys Phe Pro Val Cys Lys1
5 10 15Ser Arg Phe Gly Lys Thr Asn Gly Arg Cys Val Asn Gly Phe Cys
Asp 20 25 30Cys Phe20535PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 205Gly Ser Val Lys Cys
Ser Ser Pro Gln Gln Cys Leu Lys Pro Cys Lys1 5 10 15Ala Ala Phe Gly
Ile Ser Ala Gly Gly Lys Cys Ile Asn Gly Lys Cys 20 25 30Lys Cys Tyr
3520634PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 206Gly Ser Val Ser Cys Ser Ala Ser Ser Gln
Cys Trp Pro Val Cys Lys1 5 10 15Lys Leu Phe Gly Thr Tyr Arg Gly Lys
Cys Met Asn Ser Lys Cys Arg 20 25 30Cys Tyr20734PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
207Gly Ser Glu Ser Cys Thr Ala Ser Asn Gln Cys Trp Ser Ile Cys Lys1
5 10 15Arg Leu His Asn Thr Asn Arg Gly Lys Cys Met Asn Lys Lys Cys
Arg 20 25 30Cys Tyr20834PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 208Gly Ser Val Ser Cys
Thr Thr Ser Lys Glu Cys Trp Ser Val Cys Glu1 5 10 15Lys Leu Tyr Asn
Thr Ser Arg Gly Lys Cys Met Asn Lys Lys Cys Arg 20 25 30Cys
Tyr20934PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 209Gly Ser Met Arg Cys Lys Ser Ser Lys Glu
Cys Leu Val Lys Cys Lys1 5 10 15Gln Ala Thr Gly Arg Pro Asn Gly Lys
Cys Met Asn Arg Lys Cys Lys 20 25 30Cys Tyr21034PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
210Gly Ser Ile Lys Cys Thr Leu Ser Lys Asp Cys Tyr Ser Pro Cys
Lys1
5 10 15Lys Glu Thr Gly Cys Pro Arg Ala Lys Cys Ile Asn Arg Asn Cys
Lys 20 25 30Cys Tyr21134PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 211Gly Ser Ile Arg Cys
Ser Gly Ser Arg Asp Cys Tyr Ser Pro Cys Met1 5 10 15Lys Gln Thr Gly
Cys Pro Asn Ala Lys Cys Ile Asn Lys Ser Cys Lys 20 25 30Cys
Tyr21234PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 212Gly Ser Ile Arg Cys Ser Gly Thr Arg Glu
Cys Tyr Ala Pro Cys Gln1 5 10 15Lys Leu Thr Gly Cys Leu Asn Ala Lys
Cys Met Asn Lys Ala Cys Lys 20 25 30Cys Tyr21334PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
213Gly Ser Ile Ser Cys Thr Asn Pro Lys Gln Cys Tyr Pro His Cys Lys1
5 10 15Lys Glu Thr Gly Tyr Pro Asn Ala Lys Cys Met Asn Arg Lys Cys
Lys 20 25 30Cys Phe21434PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 214Gly Ser Ala Ser Cys
Arg Thr Pro Lys Asp Cys Ala Asp Pro Cys Arg1 5 10 15Lys Glu Thr Gly
Cys Pro Tyr Gly Lys Cys Met Asn Arg Lys Cys Lys 20 25 30Cys
Asn21533PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 215Gly Ser Thr Ser Cys Ile Ser Pro Lys Gln
Cys Thr Glu Pro Cys Arg1 5 10 15Ala Lys Gly Cys Lys His Gly Lys Cys
Met Asn Arg Lys Cys His Cys 20 25 30Met21633PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
216Gly Ser Lys Glu Cys Thr Gly Pro Gln His Cys Thr Asn Phe Cys Arg1
5 10 15Lys Asn Lys Cys Thr His Gly Lys Cys Met Asn Arg Lys Cys Lys
Cys 20 25 30Phe21734PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 217Gly Ser Ile Lys Cys Arg Thr Pro
Lys Asp Cys Ala Asp Pro Cys Arg1 5 10 15Lys Gln Thr Gly Cys Pro His
Ala Lys Cys Met Asn Lys Thr Cys Arg 20 25 30Cys
His21834PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 218Gly Ser Val Lys Cys Thr Thr Ser Lys Glu
Cys Trp Pro Pro Cys Lys1 5 10 15Ala Ala Thr Gly Lys Ala Ala Gly Lys
Cys Met Asn Lys Lys Cys Lys 20 25 30Cys Gln21934PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
219Gly Ser Leu Glu Cys Gly Ala Ser Arg Glu Cys Tyr Asp Pro Cys Phe1
5 10 15Lys Ala Phe Gly Arg Ala His Gly Lys Cys Met Asn Asn Lys Cys
Arg 20 25 30Cys Tyr22034PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 220Gly Ser Glu Lys Cys
Phe Ala Thr Ser Gln Cys Trp Thr Pro Cys Lys1 5 10 15Lys Ala Ile Gly
Ser Leu Gln Ser Lys Cys Met Asn Gly Lys Cys Lys 20 25 30Cys
Tyr22134PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 221Gly Ser Val Arg Cys Tyr Ala Ser Arg Glu
Cys Trp Glu Pro Cys Arg1 5 10 15Arg Val Thr Gly Ser Ala Gln Ala Lys
Cys Gln Asn Asn Gln Cys Arg 20 25 30Cys Tyr22234PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
222Gly Ser Val Lys Cys Ser Ala Ser Arg Glu Cys Trp Val Ala Cys Lys1
5 10 15Lys Val Thr Gly Ser Gly Gln Gly Lys Cys Gln Asn Asn Gln Cys
Arg 20 25 30Cys Tyr22334PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 223Gly Ser Val Lys Cys
Ile Ser Ser Gln Glu Cys Trp Ile Ala Cys Lys1 5 10 15Lys Val Thr Gly
Arg Phe Glu Gly Lys Cys Gln Asn Arg Gln Cys Arg 20 25 30Cys
Tyr22434PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 224Gly Ser Val Arg Cys Tyr Asp Ser Arg Gln
Cys Trp Ile Ala Cys Lys1 5 10 15Lys Val Thr Gly Ser Thr Gln Gly Lys
Cys Gln Asn Lys Gln Cys Arg 20 25 30Cys Tyr22534PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
225Gly Ser Val Asp Cys Thr Val Ser Lys Glu Cys Trp Ala Pro Cys Lys1
5 10 15Ala Ala Phe Gly Val Asp Arg Gly Lys Cys Met Gly Lys Lys Cys
Lys 20 25 30Cys Tyr22634PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 226Gly Ser Ala Lys Cys
Arg Gly Ser Pro Glu Cys Leu Pro Lys Cys Lys1 5 10 15Glu Ala Ile Gly
Lys Ala Ala Gly Lys Cys Met Asn Gly Lys Cys Lys 20 25 30Cys
Tyr22733PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 227Gly Ser Lys Lys Cys Gln Gly Gly Ser Cys
Ala Ser Val Cys Arg Arg1 5 10 15Val Ile Gly Val Ala Ala Gly Lys Cys
Ile Asn Gly Arg Cys Val Cys 20 25 30Tyr22834PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
228Gly Ser Lys Lys Cys Ser Asn Thr Ser Gln Cys Tyr Lys Thr Cys Glu1
5 10 15Lys Val Val Gly Val Ala Ala Gly Lys Cys Met Asn Gly Lys Cys
Ile 20 25 30Cys Tyr22934PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 229Gly Ser Val Lys Cys
Ser Gly Ser Ser Lys Cys Val Lys Ile Cys Ile1 5 10 15Asp Arg Tyr Asn
Thr Arg Gly Ala Lys Cys Ile Asn Gly Arg Cys Thr 20 25 30Cys
Tyr23034PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 230Gly Ser Asn Arg Cys Asn Asn Ser Ser Glu
Cys Ile Pro His Cys Ile1 5 10 15Arg Ile Phe Gly Thr Arg Ala Ala Lys
Cys Ile Asn Arg Lys Cys Tyr 20 25 30Cys Tyr23134PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
231Gly Ser Lys Glu Cys Asn Gly Ser Ser Glu Cys Tyr Ser His Cys Glu1
5 10 15Gly Ile Thr Gly Lys Arg Ser Gly Lys Cys Ile Asn Lys Lys Cys
Tyr 20 25 30Cys Tyr23231PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 232Gly Ser Ala Phe Cys
Asn Leu Arg Arg Cys Glu Leu Ser Cys Arg Ser1 5 10 15Leu Gly Leu Leu
Gly Lys Cys Ile Gly Glu Glu Cys Lys Cys Val 20 25
3023331PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 233Gly Ser Ala Val Cys Asn Leu Lys Arg Cys
Gln Leu Ser Cys Arg Ser1 5 10 15Leu Gly Leu Leu Gly Lys Cys Ile Gly
Asp Lys Cys Glu Cys Val 20 25 3023432PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
234Gly Ser Ala Ala Cys Tyr Ser Ser Asp Cys Arg Val Lys Cys Val Ala1
5 10 15Met Gly Phe Ser Ser Gly Lys Cys Ile Asn Ser Lys Cys Lys Cys
Tyr 20 25 3023529PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 235Gly Ser Ala Ile Cys Ala Thr Asp Ala
Asp Cys Ser Arg Lys Cys Pro1 5 10 15Gly Asn Pro Pro Cys Arg Asn Gly
Phe Cys Ala Cys Thr 20 2523629PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 236Gly Ser Thr Glu Cys Gln
Ile Lys Asn Asp Cys Gln Arg Tyr Cys Gln1 5 10 15Ser Val Lys Glu Cys
Lys Tyr Gly Lys Cys Tyr Cys Asn 20 2523729PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 237Gly
Ser Thr Gln Cys Gln Ser Val Arg Asp Cys Gln Gln Tyr Cys Leu1 5 10
15Thr Pro Asp Arg Cys Ser Tyr Gly Thr Cys Tyr Cys Lys 20
2523834PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 238Gly Ser Val Ser Cys Arg Tyr Gly Ser Asp
Cys Ala Glu Pro Cys Lys1 5 10 15Arg Leu Lys Cys Leu Leu Pro Ser Lys
Cys Ile Asn Gly Lys Cys Thr 20 25 30Cys Tyr23934PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
239Gly Ser Ile Lys Cys Arg Tyr Pro Ala Asp Cys His Ile Met Cys Arg1
5 10 15Lys Val Thr Gly Arg Ala Glu Gly Lys Cys Met Asn Gly Lys Cys
Thr 20 25 30Cys Tyr24034PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 240Gly Ser Ile Lys Cys
Ser Ser Ser Ser Ser Cys Tyr Glu Pro Cys Arg1 5 10 15Gly Val Thr Gly
Arg Ala His Gly Lys Cys Met Asn Gly Arg Cys Thr 20 25 30Cys
Tyr24134PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 241Gly Ser Val Lys Cys Thr Gly Ser Lys Gln
Cys Leu Pro Ala Cys Lys1 5 10 15Ala Ala Val Gly Lys Ala Ala Gly Lys
Cys Met Asn Gly Lys Cys Lys 20 25 30Cys Tyr24233PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
242Gly Ser Val Ser Cys Lys His Ser Gly Gln Cys Ile Lys Pro Cys Lys1
5 10 15Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn Arg Lys Cys Asp
Cys 20 25 30Thr24333PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 243Gly Ser Val Lys Cys Arg Gly Ser
Pro Gln Cys Ile Gln Pro Cys Arg1 5 10 15Asp Ala Gly Met Arg Phe Gly
Lys Cys Met Asn Gly Lys Cys His Cys 20 25 30Thr24435PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
244Gly Ser Val Lys Cys Thr Ser Pro Lys Gln Cys Leu Pro Pro Cys Lys1
5 10 15Ala Gln Phe Gly Ile Arg Ala Gly Ala Lys Cys Met Asn Gly Lys
Cys 20 25 30Lys Cys Tyr 3524535PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 245Gly Ser Val Lys Cys
Thr Ser Pro Lys Gln Cys Ser Lys Pro Cys Lys1 5 10 15Glu Leu Tyr Gly
Ser Ser Ala Gly Ala Lys Cys Met Asn Gly Lys Cys 20 25 30Lys Cys Tyr
3524635PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 246Gly Ser Val Lys Cys Thr Ser Pro Lys Gln
Cys Leu Pro Pro Cys Lys1 5 10 15Glu Ile Tyr Gly Arg His Ala Gly Ala
Lys Cys Met Asn Gly Lys Cys 20 25 30His Cys Ser
3524734PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 247Gly Ser Val Lys Cys Thr Gly Ser Lys Gln
Cys Trp Pro Val Cys Lys1 5 10 15Gln Met Phe Gly Lys Pro Asn Gly Lys
Cys Met Asn Gly Lys Cys Arg 20 25 30Cys Tyr24833PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
248Gly Ser Val Lys Cys Arg Gly Ser Arg Asp Cys Leu Asp Pro Cys Lys1
5 10 15Lys Ala Gly Met Arg Phe Gly Lys Cys Ile Asn Ser Lys Cys His
Cys 20 25 30Thr24929PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 249Gly Ser Val Arg Cys Val Thr Asp Asp
Asp Cys Phe Arg Lys Cys Pro1 5 10 15Gly Asn Pro Ser Cys Lys Arg Gly
Phe Cys Ala Cys Lys 20 2525034PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 250Gly Ser Val Pro Cys
Asn Asn Ser Arg Pro Cys Val Pro Val Cys Ile1 5 10 15Arg Glu Val Asn
Asn Lys Asn Gly Lys Cys Ser Asn Gly Lys Cys Leu 20 25 30Cys
Tyr25138PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 251Gly Ser Val Pro Ile Asn Val Lys Cys Arg
Gly Ser Arg Asp Cys Leu1 5 10 15Asp Pro Cys Lys Lys Ala Gly Met Arg
Phe Gly Lys Cys Ile Asn Ser 20 25 30Lys Cys His Cys Thr Pro
3525239PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 252Gly Ser Val Gln Thr Asn Val Lys Cys Gln
Gly Gly Ser Cys Ala Ser1 5 10 15Val Cys Arg Arg Glu Ile Gly Val Ala
Ala Gly Lys Cys Ile Asn Gly 20 25 30Lys Cys Val Cys Tyr Arg Asn
3525340PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 253Gly Ser Ala Glu Ile Ile Arg Cys Ser Gly
Thr Arg Glu Cys Tyr Ala1 5 10 15Pro Cys Gln Lys Leu Thr Gly Cys Leu
Asn Ala Lys Cys Met Asn Lys 20 25 30Ala Cys Lys Cys Tyr Gly Cys Val
35 4025438PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 254Gly Ser Arg Pro Thr Asp Ile Lys Cys Ser
Ala Ser Tyr Gln Cys Phe1 5 10 15Pro Val Cys Lys Ser Arg Phe Gly Lys
Thr Asn Gly Arg Cys Val Asn 20 25 30Gly Leu Cys Asp Cys Phe
3525539PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 255Gly Ser Gln Phe Thr Asp Val Lys Cys Thr
Gly Ser Lys Gln Cys Trp1 5 10 15Pro Val Cys Lys Gln Met Phe Gly Lys
Pro Asn Gly Lys Cys Met Asn 20 25 30Gly Lys Cys Arg Cys Tyr Ser
3525639PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 256Gly Ser Val Gly Ile Asn Val Lys Cys Lys
His Ser Arg Gln Cys Leu1 5 10 15Lys Pro Cys Lys Asp Ala Gly Met Arg
Phe Gly Lys Cys Thr Asn Gly 20 25 30Lys Cys His Cys Thr Pro Lys
3525737PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 257Gly Ser Val Val Ile Gly Gln Arg Cys Tyr
Arg Ser Pro Asp Cys Tyr1 5 10 15Ser Ala Cys Lys Lys Leu Val Gly Lys
Ala Thr Gly Lys Cys Thr Asn 20 25 30Gly Arg Cys Asp Cys
3525837PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 258Gly Ser Asn Phe Lys Val Glu Gly Ala Cys
Ser Lys Pro Cys Arg Lys1 5 10 15Tyr Cys Ile Asp Lys Gly Ala Arg Asn
Gly Lys Cys Ile Asn Gly Arg 20 25 30Cys His Cys Tyr Tyr
3525940PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 259Gly Ser Gln Ile Asp Thr Asn Val Lys Cys
Ser Gly Ser Ser Lys Cys1 5 10 15Val Lys Ile Cys Ile Asp Arg Tyr Asn
Thr Arg Gly Ala Lys Cys Ile 20 25 30Asn Gly Arg Cys Thr Cys Tyr Pro
35 4026039PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 260Gly Ser Gly Val Pro Ile Ser Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Asn 20 25 30Gly Arg Cys His Cys Thr Pro
3526139PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 261Gly Ser Gly Val Pro Ile Ser Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Gln 20 25 30Ser Arg Cys His Cys Thr Pro
3526239PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 262Gly Ser Gly Val Pro Ile Ser Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Asn 20 25 30Arg Arg Cys His Cys Thr Pro
3526339PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 263Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser
Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly
Arg Cys Ile Asn 20 25 30Ser Arg Cys His Cys Thr Pro
3526439PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 264Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Gln 20 25 30Ser Arg Cys His Cys Thr Pro
3526539PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 265Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Gln 20 25 30Ser Arg Cys His Cys Tyr Pro
3526639PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 266Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Tyr Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Gln 20 25 30Ser Arg Cys His Cys Thr Pro
3526739PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 267Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Gln 20 25 30Ser Arg Cys Tyr Cys Thr Pro
3526839PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 268Gly Ser Gly Val Pro Ile Ser Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Gln 20 25 30Ser Arg Cys His Cys Tyr Pro
3526939PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 269Gly Ser Gly Val Pro Ile Ser Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Tyr Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Gln 20 25 30Ser Arg Cys His Cys Thr Pro
3527039PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 270Gly Ser Gly Val Pro Ile Ser Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Gln 20 25 30Ser Arg Cys Tyr Cys Thr Pro
3527139PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 271Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Ala 20 25 30Ser Arg Cys His Cys Tyr Pro
3527239PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 272Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Ser 20 25 30Ser Arg Cys His Cys Tyr Pro
3527339PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 273Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Thr 20 25 30Ser Arg Cys His Cys Tyr Pro
3527439PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 274Gly Ser Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys1 5 10 15Leu Glu Pro Cys Arg Arg Ala Gly Thr
Arg Phe Gly Arg Cys Ile Asn 20 25 30Ser Arg Cys His Cys Tyr Pro
3527540PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(3)..(3)Any amino acid, amino acid
analogue or nullMOD_RES(5)..(6)Any amino acid, amino acid analogue
or nullMOD_RES(9)..(9)Any amino acid, amino acid analogue or
nullMOD_RES(15)..(15)Any amino acid, amino acid analogue or
nullMOD_RES(20)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(30)..(30)Any amino acid, amino acid analogue or
nullMOD_RES(37)..(40)Any amino acid, amino acid analogue or null
275Gly Val Xaa Ile Xaa Xaa Lys Cys Xaa Gly Ser Lys Gln Cys Xaa Asp1
5 10 15Pro Cys Lys Xaa Xaa Xaa Gly Xaa Arg Xaa Gly Lys Cys Xaa Asn
Lys 20 25 30Lys Cys Lys Cys Xaa Xaa Xaa Xaa 35 4027640PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(3)..(3)P or RMOD_RES(5)..(5)P or
NMOD_RES(6)..(6)V or IMOD_RES(9)..(9)S, T, R or KMOD_RES(15)..(15)Y
or LMOD_RES(20)..(20)Q, R or KMOD_RES(21)..(21)A, K or
RMOD_RES(22)..(22)T or AMOD_RES(24)..(24)C or MMOD_RES(26)..(26)F
or NMOD_RES(30)..(30)M or IMOD_RES(37)..(37)Y or
TMOD_RES(38)..(38)G or PMISC_FEATURE(39)..(40)May or may not be
present 276Gly Val Xaa Ile Xaa Xaa Lys Cys Xaa Gly Ser Lys Gln Cys
Xaa Asp1 5 10 15Pro Cys Lys Xaa Xaa Xaa Gly Xaa Arg Xaa Gly Lys Cys
Xaa Asn Lys 20 25 30Lys Cys Lys Cys Xaa Xaa Cys Gly 35
4027739PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(4)Any amino acid, amino acid
analogue or nullMOD_RES(6)..(6)Any amino acid, amino acid analogue
or nullMOD_RES(8)..(8)Any amino acid, amino acid analogue or
nullMOD_RES(15)..(15)Any amino acid, amino acid analogue or
nullMOD_RES(19)..(20)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(29)..(30)Any amino acid, amino acid analogue or
nullMOD_RES(32)..(32)Any amino acid, amino acid analogue or null
277Xaa Xaa Xaa Xaa Ile Xaa Cys Xaa Gly Ser Lys Gln Cys Tyr Xaa Pro1
5 10 15Cys Lys Xaa Xaa Thr Gly Cys Xaa Xaa Xaa Lys Cys Xaa Xaa Lys
Xaa 20 25 30Cys Lys Cys Tyr Gly Cys Gly 3527839PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMISC_FEATURE(1)..(2)May or may not be
presentMOD_RES(3)..(3)E, G or nullMOD_RES(4)..(4)V, S or
nullMOD_RES(6)..(6)R or SMOD_RES(8)..(8)S or TMOD_RES(15)..(15)G or
DMOD_RES(19)..(19)Q or RMOD_RES(20)..(20)Q or KMOD_RES(24)..(24)T
or PMOD_RES(25)..(25)N or QMOD_RES(26)..(26)S or
AMOD_RES(29)..(29)M or LMOD_RES(30)..(30)N or QMOD_RES(32)..(32)V
or S 278Gly Ser Xaa Xaa Ile Xaa Cys Xaa Gly Ser Lys Gln Cys Tyr Xaa
Pro1 5 10 15Cys Lys Xaa Xaa Thr Gly Cys Xaa Xaa Xaa Lys Cys Xaa Xaa
Lys Xaa 20 25 30Cys Lys Cys Tyr Gly Cys Gly 3527940PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(1)..(3)Any amino acid, amino acid analogue or
nullMOD_RES(5)..(5)Any amino acid, amino acid analogue or
nullMOD_RES(7)..(7)Any amino acid, amino acid analogue or
nullMOD_RES(9)..(9)Any amino acid, amino acid analogue or
nullMOD_RES(11)..(12)Any amino acid, amino acid analogue or
nullMOD_RES(14)..(15)Any amino acid, amino acid analogue or
nullMOD_RES(18)..(18)Any amino acid, amino acid analogue or
nullMOD_RES(22)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(31)..(31)Any amino acid, amino acid analogue or
nullMOD_RES(33)..(33)Any amino acid, amino acid analogue or
nullMOD_RES(36)..(36)Any amino acid, amino acid analogue or
nullMOD_RES(40)..(40)Any amino acid, amino acid analogue or null
279Xaa Xaa Xaa Val Xaa Ile Xaa Val Xaa Cys Xaa Xaa Ser Xaa Xaa Cys1
5 10 15Leu Xaa Pro Cys Lys Xaa Ala Gly Met Arg Phe Gly Lys Cys Xaa
Asn 20 25 30Xaa Lys Cys Xaa Cys Thr Pro Xaa 35 4028040PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMISC_FEATURE(1)..(1)May or may not be
presentMOD_RES(2)..(2)G, S or nullMOD_RES(3)..(3)G, S or
nullMOD_RES(5)..(5)P or RMOD_RES(7)..(7)N or PMOD_RES(9)..(9)K or
SMOD_RES(11)..(11)R or KMOD_RES(12)..(12)G or HMOD_RES(14)..(14)R
or GMOD_RES(15)..(15)D or QMOD_RES(18)..(18)D or
KMOD_RES(22)..(22)K or DMOD_RES(31)..(31)I or MMOD_RES(33)..(33)S
or GMOD_RES(36)..(36)H or DMISC_FEATURE(40)..(40)May or may not be
present 280Gly Xaa Xaa Val Xaa Ile Xaa Val Xaa Cys Xaa Xaa Ser Xaa
Xaa Cys1 5 10 15Leu Xaa Pro Cys Lys Xaa Ala Gly Met Arg Phe Gly Lys
Cys Xaa Asn 20 25 30Xaa Lys Cys Xaa Cys Thr Pro Lys 35
4028135PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(1)Any amino
acidMOD_RES(3)..(3)Any amino acidMOD_RES(7)..(7)Any amino
acidMOD_RES(13)..(13)Any amino acidMOD_RES(18)..(18)Any amino
acidMOD_RES(20)..(20)Any amino acidMOD_RES(22)..(22)Any amino
acidMOD_RES(30)..(30)Any amino acidMOD_RES(35)..(35)Any amino
acidSee specification as filed for detailed description of
substitutions and preferred embodiments 281Xaa Val Xaa Val Lys Cys
Xaa Gly Ser Lys Gln Cys Xaa Pro Cys Lys1 5 10 15Arg Xaa Gly Xaa Arg
Xaa Gly Lys Cys Ile Asn Lys Lys Xaa Cys Lys 20 25 30Cys Tyr Xaa
3528232PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(1)Any amino
acidMOD_RES(5)..(5)Any amino acidMOD_RES(11)..(11)Any amino
acidMOD_RES(13)..(13)Any amino acidMOD_RES(16)..(16)Any amino
acidMOD_RES(18)..(18)Any amino acidMOD_RES(27)..(27)Any amino
acidMOD_RES(32)..(32)Any amino acidSee specification as filed for
detailed description of substitutions and preferred embodiments
282Xaa Gly Cys Val Xaa Lys Cys Arg Pro Gly Xaa Lys Xaa Cys Cys Xaa1
5 10 15Pro Xaa Lys Arg Cys Ser Arg Arg Phe Gly Xaa Lys Lys Cys Lys
Xaa 20 25 3028341PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptideMOD_RES(1)..(1)Any amino acid, amino
acid analogue or nullMOD_RES(3)..(5)Any amino acid, amino acid
analogue or nullMOD_RES(9)..(9)Any amino acid, amino acid analogue
or nullMOD_RES(15)..(16)Any amino acid, amino acid analogue or
nullMOD_RES(21)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(34)..(34)Any amino acid, amino acid analogue or
nullMOD_RES(39)..(41)Any amino acid, amino acid analogue or null
283Xaa Val Xaa Xaa Xaa Val Lys Cys Xaa Gly Ser Lys Gln Cys Xaa Xaa1
5 10 15Pro Cys Lys Arg Xaa Xaa Gly Xaa Arg Xaa Gly Lys Cys Ile Asn
Lys 20 25 30Lys Xaa Cys Lys Cys Tyr Xaa Xaa Xaa 35
4028445PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(3)Any amino
acidMOD_RES(7)..(10)Any amino acidMOD_RES(16)..(16)Any amino
acidMOD_RES(18)..(18)Any amino acidMOD_RES(21)..(21)Any amino
acidMOD_RES(23)..(23)Any amino acidMOD_RES(32)..(35)Any amino
acidMOD_RES(40)..(45)Any amino acid 284Xaa Xaa Xaa Gly Cys Val Xaa
Xaa Xaa Xaa Lys Cys Arg Pro Gly Xaa1 5 10 15Lys Xaa Cys Cys Xaa Pro
Xaa Lys Arg Cys Ser Arg Arg Phe Gly Xaa 20 25 30Xaa Xaa Xaa Lys Lys
Cys Lys Xaa Xaa Xaa Xaa Xaa Xaa 35 40 4528540PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(3)..(3)Any amino acid, amino acid analogue or
nullMOD_RES(5)..(6)Any amino acid, amino acid analogue or
nullMOD_RES(9)..(9)Any amino acid, amino acid analogue or
nullMOD_RES(15)..(15)Any amino acid, amino acid analogue or
nullMOD_RES(20)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(30)..(30)Any amino acid, amino acid analogue or
nullMOD_RES(37)..(40)Any amino acid, amino acid analogue or null
285Gly Val Xaa Ile Xaa Xaa Arg Cys Xaa Gly Ser Arg Gln Cys Xaa Asp1
5 10 15Pro Cys Arg Xaa Xaa Xaa Gly Xaa Arg Xaa Gly Arg Cys Xaa Asn
Arg 20 25 30Arg Cys Arg Cys Xaa Xaa Xaa Xaa 35 4028640PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(3)..(3)P or RMOD_RES(5)..(5)P or
NMOD_RES(6)..(6)V or IMOD_RES(9)..(9)S, T, R or KMOD_RES(15)..(15)Y
or LMOD_RES(20)..(20)Q, R or KMOD_RES(21)..(21)A, K or
RMOD_RES(22)..(22)T or AMOD_RES(24)..(24)C or MMOD_RES(26)..(26)F
or NMOD_RES(30)..(30)M or IMOD_RES(37)..(37)Y or
TMOD_RES(38)..(38)G or PMISC_FEATURE(39)..(40)May or may not be
present 286Gly Val Xaa Ile Xaa Xaa Arg Cys Xaa Gly Ser Arg Gln Cys
Xaa Asp1 5 10 15Pro Cys Arg Xaa Xaa Xaa Gly Xaa Arg Xaa Gly Arg Cys
Xaa Asn Arg 20 25 30Arg Cys Arg Cys Xaa Xaa Cys Gly 35
4028739PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(4)Any amino acid, amino acid
analogue or nullMOD_RES(6)..(6)Any amino acid, amino acid analogue
or nullMOD_RES(8)..(8)Any amino acid, amino acid analogue or
nullMOD_RES(15)..(15)Any amino acid, amino acid analogue or
nullMOD_RES(19)..(20)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(29)..(30)Any amino acid, amino acid analogue or
nullMOD_RES(32)..(32)Any amino acid, amino acid analogue or null
287Xaa Xaa Xaa Xaa Ile Xaa Cys Xaa Gly Ser Arg Gln Cys Tyr Xaa Pro1
5 10 15Cys Arg Xaa Xaa Thr Gly Cys Xaa Xaa Xaa Arg Cys Xaa Xaa Arg
Xaa 20 25 30Cys Arg Cys Tyr Gly Cys Gly 3528839PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMISC_FEATURE(1)..(2)May or may not be
presentMOD_RES(3)..(3)E, G or nullMOD_RES(4)..(4)V, S or
nullMOD_RES(6)..(6)R or SMOD_RES(8)..(8)S or TMOD_RES(15)..(15)G or
DMOD_RES(19)..(19)Q or RMOD_RES(20)..(20)Q, R or
KMOD_RES(24)..(24)T or PMOD_RES(25)..(25)N or QMOD_RES(26)..(26)S
or AMOD_RES(29)..(29)M or LMOD_RES(30)..(30)N or
QMOD_RES(32)..(32)V or S 288Gly Ser Xaa Xaa Ile Xaa Cys Xaa Gly Ser
Arg Gln Cys Tyr Xaa Pro1 5 10 15Cys Arg Xaa Xaa Thr Gly Cys Xaa Xaa
Xaa Arg Cys Xaa Xaa Arg Xaa 20 25 30Cys Arg Cys Tyr Gly Cys Gly
3528940PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(3)Any amino acid, amino acid
analogue or nullMOD_RES(5)..(5)Any amino acid, amino acid analogue
or nullMOD_RES(7)..(7)Any amino acid, amino acid analogue or
nullMOD_RES(9)..(9)Any amino acid, amino acid analogue or
nullMOD_RES(11)..(12)Any amino acid, amino acid analogue or
nullMOD_RES(14)..(15)Any amino acid, amino acid analogue or
nullMOD_RES(18)..(18)Any amino acid, amino acid analogue or
nullMOD_RES(22)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(31)..(31)Any amino acid, amino acid analogue or
nullMOD_RES(33)..(33)Any amino acid, amino acid analogue or
nullMOD_RES(36)..(36)Any amino acid, amino acid analogue or
nullMOD_RES(40)..(40)Any amino acid, amino acid analogue or null
289Xaa Xaa Xaa Val Xaa Ile Xaa Val Xaa Cys Xaa Xaa Ser Xaa Xaa Cys1
5 10 15Leu Xaa Pro Cys Arg Xaa Ala Gly Met Arg Phe Gly Arg Cys Xaa
Asn 20 25 30Xaa Arg Cys Xaa Cys Thr Pro Xaa 35 4029040PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMISC_FEATURE(1)..(1)May or may not be
presentMOD_RES(2)..(2)G, S or nullMOD_RES(3)..(3)G, S or
nullMOD_RES(5)..(5)P or RMOD_RES(7)..(7)N or PMOD_RES(9)..(9)R, K
or SMOD_RES(11)..(11)R or KMOD_RES(12)..(12)G or
HMOD_RES(14)..(14)R or GMOD_RES(15)..(15)D or QMOD_RES(18)..(18)D,
R or KMOD_RES(22)..(22)K, R or DMOD_RES(31)..(31)I or
MMOD_RES(33)..(33)S or GMOD_RES(36)..(36)H or DMOD_RES(40)..(40)K,
R or null 290Gly Xaa Xaa Val Xaa Ile Xaa Val Xaa Cys
Xaa Xaa Ser Xaa Xaa Cys1 5 10 15Leu Xaa Pro Cys Arg Xaa Ala Gly Met
Arg Phe Gly Arg Cys Xaa Asn 20 25 30Xaa Arg Cys Xaa Cys Thr Pro Xaa
35 4029135PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(1)Any amino
acidMOD_RES(3)..(3)Any amino acidMOD_RES(7)..(7)Any amino
acidMOD_RES(13)..(13)Any amino acidMOD_RES(18)..(18)Any amino
acidMOD_RES(20)..(20)Any amino acidMOD_RES(22)..(22)Any amino
acidMOD_RES(30)..(30)Any amino acidMOD_RES(35)..(35)Any amino
acidSee specification as filed for detailed description of
substitutions and preferred embodiments 291Xaa Val Xaa Val Arg Cys
Xaa Gly Ser Arg Gln Cys Xaa Pro Cys Arg1 5 10 15Arg Xaa Gly Xaa Arg
Xaa Gly Arg Cys Ile Asn Arg Arg Xaa Cys Arg 20 25 30Cys Tyr Xaa
3529232PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(1)Any amino
acidMOD_RES(5)..(5)Any amino acidMOD_RES(11)..(11)Any amino
acidMOD_RES(13)..(13)Any amino acidMOD_RES(16)..(16)Any amino
acidMOD_RES(18)..(18)Any amino acidMOD_RES(27)..(27)Any amino
acidMOD_RES(32)..(32)Any amino acidSee specification as filed for
detailed description of substitutions and preferred embodiments
292Xaa Gly Cys Val Xaa Arg Cys Arg Pro Gly Xaa Arg Xaa Cys Cys Xaa1
5 10 15Pro Xaa Arg Arg Cys Ser Arg Arg Phe Gly Xaa Arg Arg Cys Arg
Xaa 20 25 3029341PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptideMOD_RES(1)..(1)Any amino acid
analogue or nullMOD_RES(3)..(5)Any amino acid analogue or
nullMOD_RES(9)..(9)Any amino acid analogue or
nullMOD_RES(15)..(16)Any amino acid analogue or
nullMOD_RES(21)..(22)Any amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid analogue or
nullMOD_RES(34)..(34)Any amino acid analogue or
nullMOD_RES(39)..(41)Any amino acid analogue or null 293Xaa Val Xaa
Xaa Xaa Val Arg Cys Xaa Gly Ser Arg Gln Cys Xaa Xaa1 5 10 15Pro Cys
Arg Arg Xaa Xaa Gly Xaa Arg Xaa Gly Arg Cys Ile Asn Arg 20 25 30Arg
Xaa Cys Arg Cys Tyr Xaa Xaa Xaa 35 4029445PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(1)..(3)Any amino acid analogue or
nullMOD_RES(7)..(10)Any amino acid analogue or
nullMOD_RES(16)..(16)Any amino acid analogue or
nullMOD_RES(18)..(18)Any amino acid analogue or
nullMOD_RES(21)..(21)Any amino acid analogue or
nullMOD_RES(23)..(23)Any amino acid analogue or
nullMOD_RES(32)..(35)Any amino acid analogue or
nullMOD_RES(40)..(45)Any amino acid analogue or null 294Xaa Xaa Xaa
Gly Cys Val Xaa Xaa Xaa Xaa Arg Cys Arg Pro Gly Xaa1 5 10 15Arg Xaa
Cys Cys Xaa Pro Xaa Arg Arg Cys Ser Arg Arg Phe Gly Xaa 20 25 30Xaa
Xaa Xaa Arg Arg Cys Arg Xaa Xaa Xaa Xaa Xaa Xaa 35 40
4529540PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptideMOD_RES(1)..(1)Any amino acid, amino acid
analogue or nullMOD_RES(3)..(3)Any amino acid, amino acid analogue
or nullMOD_RES(5)..(5)Any amino acid, amino acid analogue or
nullMOD_RES(9)..(9)Any amino acid, amino acid analogue or
nullMOD_RES(12)..(12)Any amino acid, amino acid analogue or
nullMOD_RES(16)..(16)Any amino acid, amino acid analogue or
nullMOD_RES(20)..(20)Any amino acid, amino acid analogue or
nullMOD_RES(22)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(35)..(35)Any amino acid, amino acid analogue or
nullMOD_RES(37)..(37)Any amino acid, amino acid analogue or
nullMOD_RES(39)..(40)Any amino acid, amino acid analogue or null
295Xaa Val Xaa Ile Xaa Val Lys Cys Xaa Gly Ser Xaa Gln Cys Leu Xaa1
5 10 15Pro Cys Lys Xaa Ala Xaa Gly Xaa Arg Xaa Gly Lys Cys Met Asn
Gly 20 25 30Lys Cys Xaa Cys Xaa Pro Xaa Xaa 35 4029640PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(1)..(1)Any amino acid, amino acid analogue or
nullMOD_RES(3)..(3)Any amino acid, amino acid analogue or
nullMOD_RES(5)..(5)Any amino acid, amino acid analogue or
nullMOD_RES(9)..(9)Any amino acid, amino acid analogue or
nullMOD_RES(12)..(12)Any amino acid, amino acid analogue or
nullMOD_RES(16)..(16)Any amino acid, amino acid analogue or
nullMOD_RES(20)..(20)Any amino acid, amino acid analogue or
nullMOD_RES(22)..(22)Any amino acid, amino acid analogue or
nullMOD_RES(24)..(24)Any amino acid, amino acid analogue or
nullMOD_RES(26)..(26)Any amino acid, amino acid analogue or
nullMOD_RES(35)..(35)Any amino acid, amino acid analogue or
nullMOD_RES(37)..(37)Any amino acid, amino acid analogue or
nullMOD_RES(39)..(40)Any amino acid, amino acid analogue or null
296Xaa Val Xaa Ile Xaa Val Arg Cys Xaa Gly Ser Xaa Gln Cys Leu Xaa1
5 10 15Pro Cys Arg Xaa Ala Xaa Gly Xaa Arg Xaa Gly Arg Cys Met Asn
Gly 20 25 30Arg Cys Xaa Cys Xaa Pro Xaa Xaa 35 4029737PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
polypeptideMOD_RES(5)..(5)N, S or GMOD_RES(15)..(15)L or
YMOD_RES(16)..(16)D or EMOD_RES(23)..(23)M or TMOD_RES(30)..(30)N,
Q, A, S, T or LMOD_RES(31)..(31)S, G or RMOD_RES(34)..(34)H or
YMOD_RES(36)..(36)T or Y 297Gly Val Pro Ile Xaa Val Arg Cys Arg Gly
Ser Arg Asp Cys Xaa Xaa1 5 10 15Pro Cys Arg Arg Ala Gly Xaa Arg Phe
Gly Arg Cys Ile Xaa Xaa Arg 20 25 30Cys Xaa Cys Xaa Pro
3529810PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 298Gly Lys Cys Ile Asn Lys Lys Cys Lys Cys1 5
102994PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 299Lys Cys Ile Asn13004PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 300Lys
Lys Cys Lys13014PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 301Pro Cys Lys Arg13026PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 302Lys
Arg Cys Ser Arg Arg1 53033PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 303Lys Gln
Cys130410PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 304Gly Arg Cys Ile Asn Arg Arg Cys Arg Cys1 5
103054PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 305Arg Cys Ile Asn13064PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 306Arg
Arg Cys Arg13074PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 307Pro Cys Arg Arg13086PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 308Arg
Arg Cys Ser Arg Arg1 53093PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 309Arg Gln
Cys13104PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 310Pro Cys Lys Lys13116PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 311Lys
Lys Cys Ser Lys Lys1 53128PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 312Gly Lys Cys Met Asn Gly
Lys Cys1 53138PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 313Gly Arg Cys Met Asn Gly Arg Cys1
531432PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 314Gly Ile Val Cys Lys Val Cys Lys Ile Ile
Cys Gly Met Gln Gly Lys1 5 10 15Lys Val Asn Ile Cys Lys Ala Pro Ile
Lys Cys Lys Cys Lys Lys Gly 20 25 3031535PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
315Ser Glu Lys Asp Cys Ile Lys His Leu Gln Arg Cys Arg Glu Asn Lys1
5 10 15Asp Cys Cys Ser Lys Lys Cys Ser Arg Arg Gly Thr Asn Pro Glu
Lys 20 25 30Arg Cys Arg 3531637PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 316Val Arg Ile Pro Val
Ser Cys Lys His Ser Gly Gln Cys Leu Lys Pro1 5 10 15Cys Lys Asp Ala
Gly Met Arg Phe Gly Lys Cys Met Asn Gly Lys Cys 20 25 30Asp Cys Thr
Pro Lys 3531737PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 317Gly Val Pro Ile Asn Val Lys Cys
Arg Gly Ser Arg Asp Cys Leu Asp1 5 10 15Pro Cys Lys Lys Ala Gly Met
Arg Phe Gly Lys Cys Ile Asn Ser Lys 20 25 30Cys His Cys Thr Pro
3531833PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 318Ala Val Cys Val Tyr Arg Thr Cys Asp Lys
Asp Cys Lys Arg Arg Gly1 5 10 15Tyr Arg Ser Gly Lys Cys Ile Asn Asn
Ala Cys Lys Cys Tyr Pro Tyr 20 25 30Gly31935PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
319Ile Ser Cys Thr Gly Ser Lys Gln Cys Tyr Asp Pro Cys Lys Arg Lys1
5 10 15Thr Gly Cys Pro Asn Ala Lys Cys Met Asn Lys Ser Cys Lys Cys
Tyr 20 25 30Gly Cys Gly 3532038PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 320Gln Val Gln Thr Asn
Val Lys Cys Gln Gly Gly Ser Cys Ala Ser Val1 5 10 15Cys Arg Arg Glu
Ile Gly Val Ala Ala Gly Lys Cys Ile Asn Gly Lys 20 25 30Cys Val Cys
Tyr Arg Asn 3532137PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 321Glu Val Ile Arg Cys Ser Gly Ser
Lys Gln Cys Tyr Gly Pro Cys Lys1 5 10 15Gln Gln Thr Gly Cys Thr Asn
Ser Lys Cys Met Asn Lys Val Cys Lys 20 25 30Cys Tyr Gly Cys Gly
3532233PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 322Ala Cys Lys Gly Val Phe Asp Ala Cys Thr
Pro Gly Lys Asn Glu Cys1 5 10 15Cys Pro Asn Arg Val Cys Ser Asp Lys
His Lys Trp Cys Lys Trp Lys 20 25 30Leu32338PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
323Gln Ile Tyr Thr Ser Lys Glu Cys Asn Gly Ser Ser Glu Cys Tyr Ser1
5 10 15His Cys Glu Gly Ile Thr Gly Lys Arg Ser Gly Lys Cys Ile Asn
Lys 20 25 30Lys Cys Tyr Cys Tyr Arg 3532435PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
324Gly Cys Leu Glu Phe Trp Trp Lys Cys Asn Pro Asn Asp Asp Lys Cys1
5 10 15Cys Arg Pro Lys Leu Lys Cys Ser Lys Leu Phe Lys Leu Cys Asn
Phe 20 25 30Ser Phe Gly 3532537PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 325Asp Cys Val Arg Phe
Trp Gly Lys Cys Ser Gln Thr Ser Asp Cys Cys1 5 10 15Pro His Leu Ala
Cys Lys Ser Lys Trp Pro Arg Asn Ile Cys Val Trp 20 25 30Asp Gly Ser
Val Gly 3532633PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 326Gly Cys Phe Gly Tyr Lys Cys Asp
Tyr Tyr Lys Gly Cys Cys Ser Gly1 5 10 15Tyr Val Cys Ser Pro Thr Trp
Lys Trp Cys Val Arg Pro Gly Pro Gly 20 25 30Arg32760PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
327Met Asn Ala Lys Phe Ile Leu Leu Leu Val Leu Thr Thr Met Met Leu1
5 10 15Leu Pro Asp Thr Lys Gly Ala Glu Val Ile Arg Cys Ser Gly Ser
Lys 20 25 30Gln Cys Tyr Gly Pro Cys Lys Gln Gln Thr Gly Cys Thr Asn
Ser Lys 35 40 45Cys Met Asn Lys Val Cys Lys Cys Tyr Gly Cys Gly 50
55 6032861PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 328Met Asn Ala Lys Leu Ile Tyr Leu Leu Leu
Val Val Thr Thr Met Thr1 5 10 15Leu Met Phe Asp Thr Ala Gln Ala Val
Asp Ile Met Cys Ser Gly Pro 20 25 30Lys Gln Cys Tyr Gly Pro Cys Lys
Lys Glu Thr Gly Cys Pro Asn Ala 35 40 45Lys Cys Met Asn Arg Arg Cys
Lys Cys Tyr Gly Cys Val 50 55 6032962PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
329Met Asn Ala Lys Leu Ile Tyr Leu Leu Leu Val Val Thr Thr Met Met1
5 10 15Leu Thr Phe Asp Thr Thr Gln Ala Gly Asp Ile Lys Cys Ser Gly
Thr 20 25 30Arg Gln Cys Trp Gly Pro Cys Lys Lys Gln Thr Thr Cys Thr
Asn Ser 35 40 45Lys Cys Met Asn Gly Lys Cys Lys Cys Tyr Gly Cys Val
Gly 50 55 6033061PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 330Met Asn Thr Lys Phe Ile Phe Leu
Leu Leu Val Val Thr Asn Thr Met1 5 10 15Met Leu Phe Asp Thr Lys Pro
Val Glu Gly Ile Ser Cys Thr Gly Ser 20 25 30Lys Gln Cys Tyr Asp Pro
Cys Lys Arg Lys Thr Gly Cys Pro Asn Ala 35 40 45Lys Cys Met Asn Lys
Ser Cys Lys Cys Tyr Gly Cys Gly 50 55 6033138PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
331Gly Val Pro Ile Asn Val Lys Cys Ser Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Lys Lys Ala Gly Met Arg Phe Gly Lys Cys Ile Asn Arg
Lys 20 25 30Cys His Cys Thr Pro Lys 3533238PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
332Gly Val Pro Ile Asn Val Lys Cys Thr Gly Ser Pro Gln Cys Leu Lys1
5 10 15Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Ile Asn Gly
Lys 20 25 30Cys His Cys Thr Pro Lys 3533338PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
333Gly Val Ile Ile Asn Val Lys Cys Lys Ile Ser Arg Gln Cys Leu Glu1
5 10 15Pro Cys Lys Lys Ala Gly Met Arg Phe Gly Lys Cys Met Asn Gly
Lys 20 25 30Cys His Cys Thr Pro Lys 3533438PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
334Gly Val Pro Ile Asn Val Lys Cys Arg Gly Ser Pro Gln Cys Ile Gln1
5 10 15Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn Gly
Lys 20 25 30Cys His Cys Thr Pro Gln 3533538PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
335Gly Val Glu Ile Asn Val Lys Cys Thr Gly Ser His Gln Cys Ile Lys1
5 10 15Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Ile Asn Arg
Lys 20 25 30Cys His Cys Thr Pro Lys 3533638PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
336Gly Val Glu Ile Asn Val Lys Cys Ser Gly Ser Pro Gln Cys Leu Lys1
5 10 15Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn Arg
Lys 20 25 30Cys His Cys Thr Pro Lys 3533738PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
337Gly Val Pro Thr Asp Val Lys Cys Arg Gly Ser Pro Gln Cys Ile Gln1
5 10 15Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn Gly
Lys 20 25 30Cys His Cys Thr Pro Lys 3533838PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
338Gly Val Pro Ile Asn Val Ser Cys Thr Gly Ser Pro Gln Cys Ile Lys1
5
10 15Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn Arg
Lys 20 25 30Cys His Cys Thr Pro Lys 3533938PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
339Gly Val Pro Ile Asn Val Pro Cys Thr Gly Ser Pro Gln Cys Ile Lys1
5 10 15Pro Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn Arg
Lys 20 25 30Cys His Cys Thr Pro Lys 3534037PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
340Val Gly Ile Asn Val Lys Cys Lys His Ser Gly Gln Cys Leu Lys Pro1
5 10 15Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Ile Asn Gly Lys
Cys 20 25 30Asp Cys Thr Pro Lys 3534137PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
341Val Gly Ile Asn Val Lys Cys Lys His Ser Gly Gln Cys Leu Lys Pro1
5 10 15Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn Gly Lys
Cys 20 25 30Asp Cys Thr Pro Lys 3534237PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
342Val Gly Ile Pro Val Ser Cys Lys His Ser Gly Gln Cys Ile Lys Pro1
5 10 15Cys Lys Asp Ala Gly Met Arg Phe Gly Lys Cys Met Asn Arg Lys
Cys 20 25 30Asp Cys Thr Pro Lys 3534336PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
343Arg Lys Gly Cys Phe Lys Glu Gly His Ser Cys Pro Lys Thr Ala Pro1
5 10 15Cys Cys Arg Pro Leu Val Cys Lys Gly Pro Ser Pro Asn Thr Lys
Lys 20 25 30Cys Thr Arg Pro 3534432PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
344Ser Phe Cys Ile Pro Phe Lys Pro Cys Lys Ser Asp Glu Asn Cys Cys1
5 10 15Lys Lys Phe Lys Cys Lys Thr Thr Gly Ile Val Lys Leu Cys Arg
Trp 20 25 3034536PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 345Leu Lys Gly Cys Leu Pro Arg Asn
Arg Phe Cys Asn Ala Leu Ser Gly1 5 10 15Pro Arg Cys Cys Ser Gly Leu
Arg Cys Lys Glu Leu Ser Ile Trp Ala 20 25 30Ser Lys Cys Leu
3534634PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 346Gly Asn Tyr Cys Leu Arg Gly Arg Cys Leu
Pro Gly Gly Arg Lys Cys1 5 10 15Cys Asn Gly Arg Pro Cys Glu Cys Phe
Ala Lys Ile Cys Ser Cys Lys 20 25 30Pro Lys34729PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 347Thr
Val Lys Cys Gly Gly Cys Asn Arg Lys Cys Cys Pro Gly Gly Cys1 5 10
15Arg Ser Gly Lys Cys Ile Asn Gly Lys Cys Gln Cys Tyr 20
2534829PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 348Gly Cys Met Lys Glu Tyr Cys Ala Gly Gln Cys
Arg Gly Lys Val Ser1 5 10 15Gln Asp Tyr Cys Leu Lys His Cys Lys Cys
Ile Pro Arg 20 2534934PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 349Ala Cys Leu Gly Phe
Gly Glu Lys Cys Asn Pro Ser Asn Asp Lys Cys1 5 10 15Cys Lys Ser Ser
Ser Leu Val Cys Ser Gln Lys His Lys Trp Cys Lys 20 25 30Tyr
Gly35037PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 350Arg Gly Gly Cys Leu Pro His Asn Arg Phe
Cys Asn Ala Leu Ser Gly1 5 10 15Pro Arg Cys Cys Ser Gly Leu Arg Cys
Lys Glu Leu Ser Ile Arg Asp 20 25 30Ser Arg Cys Leu Gly
3535136PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 351Arg Gly Gly Cys Leu Pro Arg Asn Lys Phe
Cys Asn Pro Ser Ser Gly1 5 10 15Pro Arg Cys Cys Ser Gly Leu Thr Cys
Lys Glu Leu Asn Ile Trp Ala 20 25 30Ser Lys Cys Leu
3535233PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 352Gln Arg Ser Cys Ala Lys Pro Gly Asp Met
Cys Met Gly Ile Lys Cys1 5 10 15Cys Asp Gly Gln Cys Gly Cys Asn Arg
Gly Thr Gly Arg Cys Phe Cys 20 25 30Lys35340PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
353Ala Arg Gly Cys Ala Asp Ala Tyr Lys Ser Cys Asn His Pro Arg Thr1
5 10 15Cys Cys Asp Gly Tyr Asn Gly Tyr Lys Arg Ala Cys Ile Cys Ser
Gly 20 25 30Ser Asn Cys Lys Cys Lys Lys Ser 35 4035437PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
354Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Ser Gly1
5 10 15Pro Arg Cys Cys Ser Gly Leu Arg Cys Lys Glu Leu Ser Ile Trp
Asp 20 25 30Ser Arg Cys Leu Gly 3535537PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
355Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Ser Gly1
5 10 15Pro Arg Cys Cys Ser Gly Leu Lys Cys Lys Glu Leu Ser Ile Tyr
Asp 20 25 30Ser Arg Cys Leu Gly 3535637PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
356Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Ser Gly1
5 10 15Pro Arg Cys Cys Ser Arg Leu Lys Cys Lys Glu Leu Ser Ile Trp
Asp 20 25 30Ser Arg Cys Leu Gly 3535737PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
357Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Thr Gly1
5 10 15Pro Arg Cys Cys Ser Arg Leu Arg Cys Lys Glu Leu Ser Ile Trp
Asp 20 25 30Ser Ile Cys Leu Gly 3535834PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
358Ser Cys Ala Asp Ala Tyr Lys Ser Cys Asp Ser Leu Lys Cys Cys Asn1
5 10 15Asn Arg Thr Cys Met Cys Ser Met Ile Gly Thr Asn Cys Thr Cys
Arg 20 25 30Lys Lys35930PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 359Glu Arg Arg Cys Leu
Pro Ala Gly Lys Thr Cys Val Arg Gly Pro Met1 5 10 15Arg Val Pro Cys
Cys Gly Ser Cys Ser Gln Asn Lys Cys Thr 20 25 3036031PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
360Leu Cys Ser Arg Glu Gly Glu Phe Cys Tyr Lys Leu Arg Lys Cys Cys1
5 10 15Ala Gly Phe Tyr Cys Lys Ala Phe Val Leu His Cys Tyr Arg Asn
20 25 3036123PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 361Ala Cys Gly Ser Cys Arg Lys Lys Cys
Lys Gly Ser Gly Lys Cys Ile1 5 10 15Asn Gly Arg Cys Lys Cys Tyr
2036223PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 362Ala Cys Gly Ser Cys Arg Lys Lys Cys Lys Gly
Pro Gly Lys Cys Ile1 5 10 15Asn Gly Arg Cys Lys Cys Tyr
2036332PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 363Ala Cys Gln Gly Tyr Met Arg Lys Cys Gly
Arg Asp Lys Pro Pro Cys1 5 10 15Cys Lys Lys Leu Glu Cys Ser Lys Thr
Trp Arg Trp Cys Val Trp Asn 20 25 3036430PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
364Gly Arg Tyr Cys Gln Lys Trp Met Trp Thr Cys Asp Ser Lys Arg Ala1
5 10 15Cys Cys Glu Gly Leu Arg Cys Lys Leu Trp Cys Arg Lys Ile 20
25 3036534PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 365Asn Ala Lys Cys Arg Gly Ser Pro Glu Cys
Leu Pro Lys Cys Lys Glu1 5 10 15Ala Ile Gly Lys Ala Ala Gly Lys Cys
Met Asn Gly Lys Cys Lys Cys 20 25 30Tyr Pro36634PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
366Asn Val Lys Cys Arg Gly Ser Lys Glu Cys Leu Pro Ala Cys Lys Ala1
5 10 15Ala Val Gly Lys Ala Ala Gly Lys Cys Met Asn Gly Lys Cys Lys
Cys 20 25 30Tyr Pro36734PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 367Asn Val Lys Cys Arg
Gly Ser Pro Glu Cys Leu Pro Lys Cys Lys Glu1 5 10 15Ala Ile Gly Lys
Ser Ala Gly Lys Cys Met Asn Gly Lys Cys Lys Cys 20 25 30Tyr
Pro36834PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 368Asn Ala Lys Cys Arg Gly Ser Pro Glu Cys
Leu Pro Lys Cys Lys Gln1 5 10 15Ala Ile Gly Lys Ala Ala Gly Lys Cys
Met Asn Gly Lys Cys Lys Cys 20 25 30Tyr Pro36933PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
369Arg Gly Tyr Cys Ala Glu Lys Gly Ile Lys Cys His Asn Ile His Cys1
5 10 15Cys Ser Gly Leu Thr Cys Lys Cys Lys Gly Ser Ser Cys Val Cys
Arg 20 25 30Lys37031PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 370Glu Arg Gly Cys Lys Leu Thr Phe
Trp Lys Cys Lys Asn Lys Lys Glu1 5 10 15Cys Cys Gly Trp Asn Ala Cys
Ala Leu Gly Ile Cys Met Pro Arg 20 25 3037139PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
371Lys Lys Lys Cys Ile Ala Lys Asp Tyr Gly Arg Cys Lys Trp Gly Gly1
5 10 15Thr Pro Cys Cys Arg Gly Arg Gly Cys Ile Cys Ser Ile Met Gly
Thr 20 25 30Asn Cys Glu Cys Lys Pro Arg 3537229PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 372Gly
Cys Lys Leu Thr Phe Trp Lys Cys Lys Asn Lys Lys Glu Cys Cys1 5 10
15Gly Trp Asn Ala Cys Ala Leu Gly Ile Cys Met Pro Arg 20
2537332PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 373Ala Cys Lys Gly Leu Phe Val Thr Cys Thr
Pro Gly Lys Asp Glu Cys1 5 10 15Cys Pro Asn His Val Cys Ser Ser Lys
His Lys Trp Cys Lys Tyr Lys 20 25 3037436PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
374Ile Ala Cys Ala Pro Arg Gly Leu Leu Cys Phe Arg Asp Lys Glu Cys1
5 10 15Cys Lys Gly Leu Thr Cys Lys Gly Arg Phe Val Asn Thr Trp Pro
Thr 20 25 30Phe Cys Leu Val 3537536PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
375Ala Cys Ala Gly Leu Tyr Lys Lys Cys Gly Lys Gly Val Asn Thr Cys1
5 10 15Cys Glu Asn Arg Pro Cys Lys Cys Asp Leu Ala Met Gly Asn Cys
Ile 20 25 30Cys Lys Lys Lys 3537638PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
376Phe Thr Cys Ala Ile Ser Cys Asp Ile Lys Val Asn Gly Lys Pro Cys1
5 10 15Lys Gly Ser Gly Glu Lys Lys Cys Ser Gly Gly Trp Ser Cys Lys
Phe 20 25 30Asn Val Cys Val Lys Val 3537733PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
377Gly Phe Cys Ala Gln Lys Gly Ile Lys Cys His Asp Ile His Cys Cys1
5 10 15Thr Asn Leu Lys Cys Val Arg Glu Gly Ser Asn Arg Val Cys Arg
Lys 20 25 30Ala37842PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 378Cys Ala Lys Lys Arg Asn Trp Cys
Gly Lys Asn Glu Asp Cys Cys Cys1 5 10 15Pro Met Lys Cys Ile Tyr Ala
Trp Tyr Asn Gln Gln Gly Ser Cys Gln 20 25 30Ser Thr Ile Thr Gly Leu
Phe Lys Lys Cys 35 4037928PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 379Tyr Cys Gln Lys Trp Met
Trp Thr Cys Asp Ser Ala Arg Lys Cys Cys1 5 10 15Glu Gly Leu Val Cys
Arg Leu Trp Cys Lys Lys Ile 20 2538037PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
380Arg Gly Gly Cys Leu Pro His Asn Lys Phe Cys Asn Ala Leu Ser Gly1
5 10 15Pro Arg Cys Cys Ser Gly Leu Lys Cys Lys Glu Leu Thr Ile Trp
Asn 20 25 30Thr Lys Cys Leu Glu 3538134PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
381Asn Val Lys Cys Thr Gly Ser Lys Gln Cys Leu Pro Ala Cys Lys Ala1
5 10 15Ala Val Gly Lys Ala Ala Gly Lys Cys Met Asn Gly Lys Cys Lys
Cys 20 25 30Tyr Thr38233PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 382Gln Arg Ser Cys Ala
Lys Pro Gly Glu Met Cys Met Arg Ile Lys Cys1 5 10 15Cys Asp Gly Gln
Cys Gly Cys Asn Arg Gly Thr Gly Arg Cys Phe Cys 20 25
30Lys38336PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 383Gly Cys Ile Pro Lys His Lys Arg Cys Thr
Trp Ser Gly Pro Lys Cys1 5 10 15Cys Asn Asn Ile Ser Cys His Cys Asn
Ile Ser Gly Thr Leu Cys Lys 20 25 30Cys Arg Pro Gly
3538433PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 384Asn Tyr Cys Val Ala Lys Arg Cys Arg Pro
Gly Gly Arg Gln Cys Cys1 5 10 15Ser Gly Lys Pro Cys Ala Cys Val Gly
Lys Val Cys Lys Cys Pro Arg 20 25 30Asp38537PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
385Glu Arg Gly Cys Ser Gly Ala Tyr Lys Arg Cys Ser Ser Ser Gln Arg1
5 10 15Cys Cys Glu Gly Arg Pro Cys Val Cys Ser Ala Ile Asn Ser Asn
Cys 20 25 30Lys Cys Arg Lys Thr 3538636PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
386Arg Tyr Cys Pro Arg Asn Pro Glu Ala Cys Tyr Asn Tyr Cys Leu Arg1
5 10 15Thr Gly Arg Pro Gly Gly Tyr Cys Gly Gly Arg Ser Arg Ile Thr
Cys 20 25 30Phe Cys Phe Arg 3538733PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
387Gln Arg Ser Cys Ala Lys Pro Gly Glu Met Cys Met Gly Ile Lys Cys1
5 10 15Cys Asp Gly Gln Cys Gly Cys Asn Arg Gly Thr Gly Arg Cys Phe
Cys 20 25 30Lys38836PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 388Arg Arg Gly Cys Phe Lys Glu Gly
Lys Trp Cys Pro Lys Ser Ala Pro1 5 10 15Cys Cys Ala Pro Leu Lys Cys
Lys Gly Pro Ser Ile Lys Gln Gln Lys 20 25 30Cys Val Arg Glu
3538931PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 389Thr Val Lys Cys Gly Gly Cys Asn Arg Lys
Cys Cys Ala Gly Gly Cys1 5 10 15Arg Ser Gly Lys Cys Ile Asn Gly Lys
Cys Gln Cys Tyr Gly Arg 20 25 3039029PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 390Glu
Arg Arg Cys Glu Pro Ser Gly Lys Pro Cys Arg Pro Leu Met Arg1 5 10
15Ile Pro Cys Cys Gly Ser Cys Val Arg Gly Lys Cys Ala 20
2539136PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 391Arg Gly Gly Cys Leu Pro Arg Asn Lys Phe
Cys Asn Pro Ser Ser Gly1 5
10 15Pro Arg Cys Cys Ser Gly Leu Thr Cys Lys Glu Leu Asn Ile Trp
Ala 20 25 30Asn Lys Cys Leu 3539242PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
392Cys Ala Lys Lys Arg Asn Trp Cys Gly Lys Asn Glu Asp Cys Cys Cys1
5 10 15Pro Met Lys Cys Ile Tyr Ala Trp Tyr Asn Gln Gln Gly Ser Cys
Gln 20 25 30Thr Thr Ile Thr Gly Leu Phe Lys Lys Cys 35
4039337PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 393Val Arg Ile Pro Val Ser Cys Lys His Ser
Gly Gln Cys Leu Lys Pro1 5 10 15Cys Lys Asp Ala Gly Met Arg Thr Gly
Lys Cys Met Asn Gly Lys Cys 20 25 30Asp Cys Thr Pro Lys
3539420PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 394Val Lys Cys Thr Thr Ser Lys Asp Cys Trp Pro
Pro Cys Lys Lys Val1 5 10 15Thr Gly Arg Ala 2039532PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
395Gly Ile Val Cys Arg Val Cys Arg Ile Ile Cys Gly Met Gln Gly Arg1
5 10 15Arg Val Asn Ile Cys Arg Ala Pro Ile Arg Cys Arg Cys Arg Arg
Gly 20 25 3039635PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 396Ser Glu Arg Asp Cys Ile Arg His
Leu Gln Arg Cys Arg Glu Asn Arg1 5 10 15Asp Cys Cys Ser Arg Arg Cys
Ser Arg Arg Gly Thr Asn Pro Glu Arg 20 25 30Arg Cys Arg
3539737PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 397Val Arg Ile Pro Val Ser Cys Arg His Ser
Gly Gln Cys Leu Arg Pro1 5 10 15Cys Arg Asp Ala Gly Met Arg Phe Gly
Arg Cys Met Asn Gly Arg Cys 20 25 30Asp Cys Thr Pro Arg
3539837PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 398Gly Val Pro Ile Asn Val Arg Cys Arg Gly
Ser Arg Asp Cys Leu Asp1 5 10 15Pro Cys Arg Arg Ala Gly Met Arg Phe
Gly Arg Cys Ile Asn Ser Arg 20 25 30Cys His Cys Thr Pro
3539933PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 399Ala Val Cys Val Tyr Arg Thr Cys Asp Arg
Asp Cys Arg Arg Arg Gly1 5 10 15Tyr Arg Ser Gly Arg Cys Ile Asn Asn
Ala Cys Arg Cys Tyr Pro Tyr 20 25 30Gly40035PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
400Ile Ser Cys Thr Gly Ser Arg Gln Cys Tyr Asp Pro Cys Arg Arg Arg1
5 10 15Thr Gly Cys Pro Asn Ala Arg Cys Met Asn Arg Ser Cys Arg Cys
Tyr 20 25 30Gly Cys Gly 3540138PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 401Gln Val Gln Thr Asn
Val Arg Cys Gln Gly Gly Ser Cys Ala Ser Val1 5 10 15Cys Arg Arg Glu
Ile Gly Val Ala Ala Gly Arg Cys Ile Asn Gly Arg 20 25 30Cys Val Cys
Tyr Arg Asn 3540237PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 402Glu Val Ile Arg Cys Ser Gly Ser
Arg Gln Cys Tyr Gly Pro Cys Arg1 5 10 15Gln Gln Thr Gly Cys Thr Asn
Ser Arg Cys Met Asn Arg Val Cys Arg 20 25 30Cys Tyr Gly Cys Gly
3540333PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 403Ala Cys Arg Gly Val Phe Asp Ala Cys Thr
Pro Gly Arg Asn Glu Cys1 5 10 15Cys Pro Asn Arg Val Cys Ser Asp Arg
His Arg Trp Cys Arg Trp Arg 20 25 30Leu40438PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
404Gln Ile Tyr Thr Ser Arg Glu Cys Asn Gly Ser Ser Glu Cys Tyr Ser1
5 10 15His Cys Glu Gly Ile Thr Gly Arg Arg Ser Gly Arg Cys Ile Asn
Arg 20 25 30Arg Cys Tyr Cys Tyr Arg 3540535PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
405Gly Cys Leu Glu Phe Trp Trp Arg Cys Asn Pro Asn Asp Asp Arg Cys1
5 10 15Cys Arg Pro Arg Leu Arg Cys Ser Arg Leu Phe Arg Leu Cys Asn
Phe 20 25 30Ser Phe Gly 3540637PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 406Asp Cys Val Arg Phe
Trp Gly Arg Cys Ser Gln Thr Ser Asp Cys Cys1 5 10 15Pro His Leu Ala
Cys Arg Ser Arg Trp Pro Arg Asn Ile Cys Val Trp 20 25 30Asp Gly Ser
Val Gly 3540733PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 407Gly Cys Phe Gly Tyr Arg Cys Asp
Tyr Tyr Arg Gly Cys Cys Ser Gly1 5 10 15Tyr Val Cys Ser Pro Thr Trp
Arg Trp Cys Val Arg Pro Gly Pro Gly 20 25 30Arg40860PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
408Met Asn Ala Arg Phe Ile Leu Leu Leu Val Leu Thr Thr Met Met Leu1
5 10 15Leu Pro Asp Thr Arg Gly Ala Glu Val Ile Arg Cys Ser Gly Ser
Arg 20 25 30Gln Cys Tyr Gly Pro Cys Arg Gln Gln Thr Gly Cys Thr Asn
Ser Arg 35 40 45Cys Met Asn Arg Val Cys Arg Cys Tyr Gly Cys Gly 50
55 6040961PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 409Met Asn Ala Arg Leu Ile Tyr Leu Leu Leu
Val Val Thr Thr Met Thr1 5 10 15Leu Met Phe Asp Thr Ala Gln Ala Val
Asp Ile Met Cys Ser Gly Pro 20 25 30Arg Gln Cys Tyr Gly Pro Cys Arg
Arg Glu Thr Gly Cys Pro Asn Ala 35 40 45Arg Cys Met Asn Arg Arg Cys
Arg Cys Tyr Gly Cys Val 50 55 6041062PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
410Met Asn Ala Arg Leu Ile Tyr Leu Leu Leu Val Val Thr Thr Met Met1
5 10 15Leu Thr Phe Asp Thr Thr Gln Ala Gly Asp Ile Arg Cys Ser Gly
Thr 20 25 30Arg Gln Cys Trp Gly Pro Cys Arg Arg Gln Thr Thr Cys Thr
Asn Ser 35 40 45Arg Cys Met Asn Gly Arg Cys Arg Cys Tyr Gly Cys Val
Gly 50 55 6041161PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 411Met Asn Thr Arg Phe Ile Phe Leu
Leu Leu Val Val Thr Asn Thr Met1 5 10 15Met Leu Phe Asp Thr Arg Pro
Val Glu Gly Ile Ser Cys Thr Gly Ser 20 25 30Arg Gln Cys Tyr Asp Pro
Cys Arg Arg Arg Thr Gly Cys Pro Asn Ala 35 40 45Arg Cys Met Asn Arg
Ser Cys Arg Cys Tyr Gly Cys Gly 50 55 6041238PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
412Gly Val Pro Ile Asn Val Arg Cys Ser Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Met Arg Phe Gly Arg Cys Ile Asn Arg
Arg 20 25 30Cys His Cys Thr Pro Arg 3541338PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
413Gly Val Pro Ile Asn Val Arg Cys Thr Gly Ser Pro Gln Cys Leu Arg1
5 10 15Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Ile Asn Gly
Arg 20 25 30Cys His Cys Thr Pro Arg 3541438PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
414Gly Val Ile Ile Asn Val Arg Cys Arg Ile Ser Arg Gln Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Met Arg Phe Gly Arg Cys Met Asn Gly
Arg 20 25 30Cys His Cys Thr Pro Arg 3541538PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
415Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Pro Gln Cys Ile Gln1
5 10 15Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn Gly
Arg 20 25 30Cys His Cys Thr Pro Gln 3541638PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
416Gly Val Glu Ile Asn Val Arg Cys Thr Gly Ser His Gln Cys Ile Arg1
5 10 15Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Ile Asn Arg
Arg 20 25 30Cys His Cys Thr Pro Arg 3541738PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
417Gly Val Glu Ile Asn Val Arg Cys Ser Gly Ser Pro Gln Cys Leu Arg1
5 10 15Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn Arg
Arg 20 25 30Cys His Cys Thr Pro Arg 3541838PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
418Gly Val Pro Thr Asp Val Arg Cys Arg Gly Ser Pro Gln Cys Ile Gln1
5 10 15Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn Gly
Arg 20 25 30Cys His Cys Thr Pro Arg 3541938PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
419Gly Val Pro Ile Asn Val Ser Cys Thr Gly Ser Pro Gln Cys Ile Arg1
5 10 15Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn Arg
Arg 20 25 30Cys His Cys Thr Pro Arg 3542038PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
420Gly Val Pro Ile Asn Val Pro Cys Thr Gly Ser Pro Gln Cys Ile Arg1
5 10 15Pro Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn Arg
Arg 20 25 30Cys His Cys Thr Pro Arg 3542137PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
421Val Gly Ile Asn Val Arg Cys Arg His Ser Gly Gln Cys Leu Arg Pro1
5 10 15Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Ile Asn Gly Arg
Cys 20 25 30Asp Cys Thr Pro Arg 3542237PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
422Val Gly Ile Asn Val Arg Cys Arg His Ser Gly Gln Cys Leu Arg Pro1
5 10 15Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn Gly Arg
Cys 20 25 30Asp Cys Thr Pro Arg 3542337PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
423Val Gly Ile Pro Val Ser Cys Arg His Ser Gly Gln Cys Ile Arg Pro1
5 10 15Cys Arg Asp Ala Gly Met Arg Phe Gly Arg Cys Met Asn Arg Arg
Cys 20 25 30Asp Cys Thr Pro Arg 3542436PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
424Arg Arg Gly Cys Phe Arg Glu Gly His Ser Cys Pro Arg Thr Ala Pro1
5 10 15Cys Cys Arg Pro Leu Val Cys Arg Gly Pro Ser Pro Asn Thr Arg
Arg 20 25 30Cys Thr Arg Pro 3542532PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
425Ser Phe Cys Ile Pro Phe Arg Pro Cys Arg Ser Asp Glu Asn Cys Cys1
5 10 15Arg Arg Phe Arg Cys Arg Thr Thr Gly Ile Val Arg Leu Cys Arg
Trp 20 25 3042636PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 426Leu Arg Gly Cys Leu Pro Arg Asn
Arg Phe Cys Asn Ala Leu Ser Gly1 5 10 15Pro Arg Cys Cys Ser Gly Leu
Arg Cys Arg Glu Leu Ser Ile Trp Ala 20 25 30Ser Arg Cys Leu
3542734PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 427Gly Asn Tyr Cys Leu Arg Gly Arg Cys Leu
Pro Gly Gly Arg Arg Cys1 5 10 15Cys Asn Gly Arg Pro Cys Glu Cys Phe
Ala Arg Ile Cys Ser Cys Arg 20 25 30Pro Arg42829PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 428Thr
Val Arg Cys Gly Gly Cys Asn Arg Arg Cys Cys Pro Gly Gly Cys1 5 10
15Arg Ser Gly Arg Cys Ile Asn Gly Arg Cys Gln Cys Tyr 20
2542929PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 429Gly Cys Met Arg Glu Tyr Cys Ala Gly Gln Cys
Arg Gly Arg Val Ser1 5 10 15Gln Asp Tyr Cys Leu Arg His Cys Arg Cys
Ile Pro Arg 20 2543034PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 430Ala Cys Leu Gly Phe
Gly Glu Arg Cys Asn Pro Ser Asn Asp Arg Cys1 5 10 15Cys Arg Ser Ser
Ser Leu Val Cys Ser Gln Arg His Arg Trp Cys Arg 20 25 30Tyr
Gly43137PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 431Arg Gly Gly Cys Leu Pro His Asn Arg Phe
Cys Asn Ala Leu Ser Gly1 5 10 15Pro Arg Cys Cys Ser Gly Leu Arg Cys
Arg Glu Leu Ser Ile Arg Asp 20 25 30Ser Arg Cys Leu Gly
3543236PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 432Arg Gly Gly Cys Leu Pro Arg Asn Arg Phe
Cys Asn Pro Ser Ser Gly1 5 10 15Pro Arg Cys Cys Ser Gly Leu Thr Cys
Arg Glu Leu Asn Ile Trp Ala 20 25 30Ser Arg Cys Leu
3543333PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 433Gln Arg Ser Cys Ala Arg Pro Gly Asp Met
Cys Met Gly Ile Arg Cys1 5 10 15Cys Asp Gly Gln Cys Gly Cys Asn Arg
Gly Thr Gly Arg Cys Phe Cys 20 25 30Arg43440PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
434Ala Arg Gly Cys Ala Asp Ala Tyr Arg Ser Cys Asn His Pro Arg Thr1
5 10 15Cys Cys Asp Gly Tyr Asn Gly Tyr Arg Arg Ala Cys Ile Cys Ser
Gly 20 25 30Ser Asn Cys Arg Cys Arg Arg Ser 35 4043537PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
435Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Ser Gly1
5 10 15Pro Arg Cys Cys Ser Gly Leu Arg Cys Arg Glu Leu Ser Ile Trp
Asp 20 25 30Ser Arg Cys Leu Gly 3543637PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
436Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Ser Gly1
5 10 15Pro Arg Cys Cys Ser Gly Leu Arg Cys Arg Glu Leu Ser Ile Tyr
Asp 20 25 30Ser Arg Cys Leu Gly 3543737PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
437Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Ser Gly1
5 10 15Pro Arg Cys Cys Ser Arg Leu Arg Cys Arg Glu Leu Ser Ile Trp
Asp 20 25 30Ser Arg Cys Leu Gly 3543837PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
438Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Thr Gly1
5 10 15Pro Arg Cys Cys Ser Arg Leu Arg Cys Arg Glu Leu Ser Ile Trp
Asp 20 25 30Ser Ile Cys Leu Gly 3543934PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
439Ser Cys Ala Asp Ala Tyr Lys Ser Cys Asp Ser Leu Arg Cys Cys Asn1
5 10 15Asn Arg Thr Cys Met Cys Ser Met Ile Gly Thr Asn Cys Thr Cys
Arg 20 25 30Arg Arg44030PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 440Glu Arg Arg Cys Leu
Pro Ala Gly Arg Thr Cys Val Arg Gly Pro Met1 5 10 15Arg Val Pro Cys
Cys Gly Ser Cys Ser Gln Asn Arg Cys Thr 20 25
3044131PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 441Leu Cys Ser Arg Glu Gly Glu Phe Cys Tyr
Arg Leu Arg Arg Cys Cys1 5 10 15Ala Gly Phe Tyr Cys Arg Ala Phe Val
Leu His Cys Tyr Arg Asn 20 25 3044223PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 442Ala
Cys Gly Ser Cys Arg Arg Arg Cys Arg Gly Ser Gly Arg Cys Ile1 5 10
15Asn Gly Arg Cys Arg Cys Tyr 2044323PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 443Ala
Cys Gly Ser Cys Arg Arg Arg Cys Arg Gly Pro Gly Arg Cys Ile1 5 10
15Asn Gly Arg Cys Arg Cys Tyr 2044432PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
444Ala Cys Gln Gly Tyr Met Arg Arg Cys Gly Arg Asp Arg Pro Pro Cys1
5 10 15Cys Arg Arg Leu Glu Cys Ser Arg Thr Trp Arg Trp Cys Val Trp
Asn 20 25 3044530PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 445Gly Arg Tyr Cys Gln Arg Trp Met
Trp Thr Cys Asp Ser Arg Arg Ala1 5 10 15Cys Cys Glu Gly Leu Arg Cys
Arg Leu Trp Cys Arg Arg Ile 20 25 3044634PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
446Asn Ala Arg Cys Arg Gly Ser Pro Glu Cys Leu Pro Arg Cys Arg Glu1
5 10 15Ala Ile Gly Arg Ala Ala Gly Arg Cys Met Asn Gly Arg Cys Arg
Cys 20 25 30Tyr Pro44734PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 447Asn Val Arg Cys Arg
Gly Ser Arg Glu Cys Leu Pro Ala Cys Arg Ala1 5 10 15Ala Val Gly Arg
Ala Ala Gly Arg Cys Met Asn Gly Arg Cys Arg Cys 20 25 30Tyr
Pro44834PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 448Asn Val Arg Cys Arg Gly Ser Pro Glu Cys
Leu Pro Arg Cys Arg Glu1 5 10 15Ala Ile Gly Arg Ser Ala Gly Arg Cys
Met Asn Gly Arg Cys Arg Cys 20 25 30Tyr Pro44934PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
449Asn Ala Arg Cys Arg Gly Ser Pro Glu Cys Leu Pro Arg Cys Arg Gln1
5 10 15Ala Ile Gly Arg Ala Ala Gly Arg Cys Met Asn Gly Arg Cys Arg
Cys 20 25 30Tyr Pro45033PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 450Arg Gly Tyr Cys Ala
Glu Arg Gly Ile Arg Cys His Asn Ile His Cys1 5 10 15Cys Ser Gly Leu
Thr Cys Arg Cys Arg Gly Ser Ser Cys Val Cys Arg 20 25
30Arg45131PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 451Glu Arg Gly Cys Arg Leu Thr Phe Trp Arg
Cys Arg Asn Arg Arg Glu1 5 10 15Cys Cys Gly Trp Asn Ala Cys Ala Leu
Gly Ile Cys Met Pro Arg 20 25 3045239PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
452Arg Arg Arg Cys Ile Ala Arg Asp Tyr Gly Arg Cys Arg Trp Gly Gly1
5 10 15Thr Pro Cys Cys Arg Gly Arg Gly Cys Ile Cys Ser Ile Met Gly
Thr 20 25 30Asn Cys Glu Cys Arg Pro Arg 3545329PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 453Gly
Cys Arg Leu Thr Phe Trp Arg Cys Arg Asn Arg Arg Glu Cys Cys1 5 10
15Gly Trp Asn Ala Cys Ala Leu Gly Ile Cys Met Pro Arg 20
2545432PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 454Ala Cys Arg Gly Leu Phe Val Thr Cys Thr
Pro Gly Arg Asp Glu Cys1 5 10 15Cys Pro Asn His Val Cys Ser Ser Arg
His Arg Trp Cys Arg Tyr Arg 20 25 3045536PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
455Ile Ala Cys Ala Pro Arg Gly Leu Leu Cys Phe Arg Asp Arg Glu Cys1
5 10 15Cys Arg Gly Leu Thr Cys Arg Gly Arg Phe Val Asn Thr Trp Pro
Thr 20 25 30Phe Cys Leu Val 3545636PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
456Ala Cys Ala Gly Leu Tyr Arg Arg Cys Gly Arg Gly Val Asn Thr Cys1
5 10 15Cys Glu Asn Arg Pro Cys Arg Cys Asp Leu Ala Met Gly Asn Cys
Ile 20 25 30Cys Arg Arg Arg 3545738PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
457Phe Thr Cys Ala Ile Ser Cys Asp Ile Arg Val Asn Gly Arg Pro Cys1
5 10 15Arg Gly Ser Gly Glu Arg Arg Cys Ser Gly Gly Trp Ser Cys Arg
Phe 20 25 30Asn Val Cys Val Arg Val 3545833PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
458Gly Phe Cys Ala Gln Arg Gly Ile Arg Cys His Asp Ile His Cys Cys1
5 10 15Thr Asn Leu Arg Cys Val Arg Glu Gly Ser Asn Arg Val Cys Arg
Arg 20 25 30Ala45942PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 459Cys Ala Arg Arg Arg Asn Trp Cys
Gly Arg Asn Glu Asp Cys Cys Cys1 5 10 15Pro Met Arg Cys Ile Tyr Ala
Trp Tyr Asn Gln Gln Gly Ser Cys Gln 20 25 30Ser Thr Ile Thr Gly Leu
Phe Arg Arg Cys 35 4046028PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 460Tyr Cys Gln Arg Trp Met
Trp Thr Cys Asp Ser Ala Arg Arg Cys Cys1 5 10 15Glu Gly Leu Val Cys
Arg Leu Trp Cys Arg Arg Ile 20 2546137PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
461Arg Gly Gly Cys Leu Pro His Asn Arg Phe Cys Asn Ala Leu Ser Gly1
5 10 15Pro Arg Cys Cys Ser Gly Leu Arg Cys Arg Glu Leu Thr Ile Trp
Asn 20 25 30Thr Arg Cys Leu Glu 3546234PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
462Asn Val Arg Cys Thr Gly Ser Arg Gln Cys Leu Pro Ala Cys Arg Ala1
5 10 15Ala Val Gly Arg Ala Ala Gly Arg Cys Met Asn Gly Arg Cys Arg
Cys 20 25 30Tyr Thr46333PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 463Gln Arg Ser Cys Ala
Arg Pro Gly Glu Met Cys Met Arg Ile Arg Cys1 5 10 15Cys Asp Gly Gln
Cys Gly Cys Asn Arg Gly Thr Gly Arg Cys Phe Cys 20 25
30Arg46436PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 464Gly Cys Ile Pro Arg His Arg Arg Cys Thr
Trp Ser Gly Pro Arg Cys1 5 10 15Cys Asn Asn Ile Ser Cys His Cys Asn
Ile Ser Gly Thr Leu Cys Arg 20 25 30Cys Arg Pro Gly
3546533PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 465Asn Tyr Cys Val Ala Arg Arg Cys Arg Pro
Gly Gly Arg Gln Cys Cys1 5 10 15Ser Gly Arg Pro Cys Ala Cys Val Gly
Arg Val Cys Arg Cys Pro Arg 20 25 30Asp46637PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
466Glu Arg Gly Cys Ser Gly Ala Tyr Arg Arg Cys Ser Ser Ser Gln Arg1
5 10 15Cys Cys Glu Gly Arg Pro Cys Val Cys Ser Ala Ile Asn Ser Asn
Cys 20 25 30Arg Cys Arg Arg Thr 3546733PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
467Gln Arg Ser Cys Ala Arg Pro Gly Glu Met Cys Met Gly Ile Arg Cys1
5 10 15Cys Asp Gly Gln Cys Gly Cys Asn Arg Gly Thr Gly Arg Cys Phe
Cys 20 25 30Arg46836PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 468Arg Arg Gly Cys Phe Arg Glu Gly
Arg Trp Cys Pro Arg Ser Ala Pro1 5 10 15Cys Cys Ala Pro Leu Arg Cys
Arg Gly Pro Ser Ile Arg Gln Gln Arg 20 25 30Cys Val Arg Glu
3546931PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 469Thr Val Arg Cys Gly Gly Cys Asn Arg Arg
Cys Cys Ala Gly Gly Cys1 5 10 15Arg Ser Gly Arg Cys Ile Asn Gly Arg
Cys Gln Cys Tyr Gly Arg 20 25 3047029PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 470Glu
Arg Arg Cys Glu Pro Ser Gly Arg Pro Cys Arg Pro Leu Met Arg1 5 10
15Ile Pro Cys Cys Gly Ser Cys Val Arg Gly Arg Cys Ala 20
2547136PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 471Arg Gly Gly Cys Leu Pro Arg Asn Arg Phe
Cys Asn Pro Ser Ser Gly1 5 10 15Pro Arg Cys Cys Ser Gly Leu Thr Cys
Arg Glu Leu Asn Ile Trp Ala 20 25 30Asn Arg Cys Leu
3547242PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 472Cys Ala Arg Arg Arg Asn Trp Cys Gly Arg
Asn Glu Asp Cys Cys Cys1 5 10 15Pro Met Arg Cys Ile Tyr Ala Trp Tyr
Asn Gln Gln Gly Ser Cys Gln 20 25 30Thr Thr Ile Thr Gly Leu Phe Arg
Arg Cys 35 4047337PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 473Val Arg Ile Pro Val Ser Cys Arg
His Ser Gly Gln Cys Leu Arg Pro1 5 10 15Cys Arg Asp Ala Gly Met Arg
Thr Gly Arg Cys Met Asn Gly Arg Cys 20 25 30Asp Cys Thr Pro Arg
3547438PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 474Gln Lys Ile Leu Ser Asn Arg Cys Asn Asn
Ser Ser Glu Cys Ile Pro1 5 10 15His Cys Ile Arg Ile Phe Gly Thr Arg
Ala Ala Lys Cys Ile Asn Arg 20 25 30Lys Cys Tyr Cys Tyr Pro
3547532PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 475Ala Val Cys Asn Leu Lys Arg Cys Gln Leu
Ser Cys Arg Ser Leu Gly1 5 10 15Leu Leu Gly Lys Cys Ile Gly Asp Lys
Cys Glu Cys Val Lys His Gly 20 25 3047637PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
476Ile Ser Ile Gly Ile Arg Cys Ser Pro Ser Ile Asp Leu Cys Glu Gly1
5 10 15Gln Cys Arg Ile Arg Arg Tyr Phe Thr Gly Tyr Cys Ser Gly Asp
Thr 20 25 30Cys His Cys Ser Gly 3547733PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
477Gly Asp Cys Leu Pro His Leu Arg Arg Cys Arg Glu Asn Asn Asp Cys1
5 10 15Cys Ser Arg Arg Cys Arg Arg Arg Gly Ala Asn Pro Glu Arg Arg
Cys 20 25 30Arg47834PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 478Ser Cys Glu Pro Gly Arg Thr Phe
Arg Asp Arg Cys Asn Thr Cys Lys1 5 10 15Cys Gly Ala Asp Gly Arg Ser
Ala Ala Cys Thr Leu Arg Ala Cys Pro 20 25 30Asn
Gln47933PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 479Gly Asp Cys Leu Pro His Leu Lys Arg Cys
Lys Ala Asp Asn Asp Cys1 5 10 15Cys Gly Lys Lys Cys Lys Arg Arg Gly
Thr Asn Ala Glu Lys Arg Cys 20 25 30Arg48033PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
480Gly Asp Cys Leu Pro His Leu Lys Arg Cys Lys Glu Asn Asn Asp Cys1
5 10 15Cys Ser Lys Lys Cys Lys Arg Arg Gly Thr Asn Pro Glu Lys Arg
Cys 20 25 30Arg48133PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 481Lys Asp Cys Leu Lys Lys Leu Lys
Leu Cys Lys Glu Asn Lys Asp Cys1 5 10 15Cys Ser Lys Ser Cys Lys Arg
Arg Gly Thr Asn Ile Glu Lys Arg Cys 20 25 30Arg48233PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
482Gly Asp Cys Leu Pro His Leu Lys Arg Cys Lys Glu Asn Asn Asp Cys1
5 10 15Cys Ser Lys Lys Cys Lys Arg Arg Gly Ala Asn Pro Glu Lys Arg
Cys 20 25 30Arg48337PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 483Val Phe Ile Asn Val Lys Cys Arg
Gly Ser Pro Glu Cys Leu Pro Lys1 5 10 15Cys Lys Glu Ala Ile Gly Lys
Ser Ala Gly Lys Cys Met Asn Gly Lys 20 25 30Cys Lys Cys Tyr Pro
3548437PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 484Val Phe Ile Asn Ala Lys Cys Arg Gly Ser
Pro Glu Cys Leu Pro Lys1 5 10 15Cys Lys Glu Ala Ile Gly Lys Ala Ala
Gly Lys Cys Met Asn Gly Lys 20 25 30Cys Lys Cys Tyr Pro
3548536PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 485Val Ile Ile Asn Val Lys Cys Lys Ile Ser
Arg Gln Cys Leu Glu Pro1 5 10 15Cys Lys Lys Ala Gly Met Arg Phe Gly
Lys Cys Met Asn Gly Lys Cys 20 25 30His Cys Thr Pro
3548636PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 486Val Pro Thr Asp Val Lys Cys Arg Gly Ser
Pro Gln Cys Ile Gln Pro1 5 10 15Cys Lys Asp Ala Gly Met Arg Phe Gly
Lys Cys Met Asn Gly Lys Cys 20 25 30His Cys Thr Pro
3548736PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 487Val Arg Ile Pro Val Ser Cys Lys His Ser
Gly Gln Cys Leu Lys Pro1 5 10 15Cys Lys Asp Ala Gly Met Arg Phe Gly
Lys Cys Met Asn Gly Lys Cys 20 25 30Asp Cys Thr Pro
3548836PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 488Val Arg Ile Pro Val Ser Cys Arg His Ser
Gly Gln Cys Leu Arg Pro1 5 10 15Cys Arg Asp Ala Gly Met Arg Phe Gly
Arg Cys Met Asn Gly Arg Cys 20 25 30Asp Cys Thr Pro
3548933PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 489Thr Asn Val Ser Cys Thr Thr Ser Lys Glu
Cys Trp Ser Val Cys Gln1 5 10 15Arg Leu His Asn Thr Ser Arg Gly Lys
Cys Met Asn Lys Lys Cys Arg 20 25 30Cys49032PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
490Asn Val Lys Cys Thr Gly Ser Lys Gln Cys Leu Pro Ala Cys Lys Ala1
5 10 15Ala Val Gly Lys Ala Ala Gly Lys Cys Met Asn Gly Lys Cys Lys
Cys 20 25 3049137PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 491Gly Val Pro Ile Asn Val Arg Cys
Arg Gly Ser Arg Asp Cys Leu Asp1 5 10 15Pro Cys Arg Gly Ala Gly Glu
Arg His Gly Arg Cys Gly Asn Ser Arg 20 25 30Cys His Cys Thr Pro
3549237PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 492Val Arg Ile Pro Val Ser Cys Arg His Ser
Gly Gln Cys Leu Arg Pro1 5 10 15Cys Arg Asp Ala Gly Glu Arg His Gly
Arg Cys Gly Gly Gly Arg Cys 20 25 30Asp Cys Thr Pro Arg
3549338PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 493Gln Val Gln Thr Asn Val Arg Cys Gln Gly
Gly Ser Cys Gly Ser Val1 5 10 15Cys Arg Arg Glu Gly Gly Gly Ala Gly
Gly Gly Cys Gly Asn Gly Arg 20
25 30Cys Gly Cys Tyr Arg Asn 3549432PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
494Ile Lys Cys Ser Glu Ser Tyr Gln Cys Phe Pro Val Cys Lys Ser Arg1
5 10 15Phe Gly Lys Thr Asn Gly Arg Cys Val Asn Gly Phe Cys Asp Cys
Phe 20 25 3049533PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 495Val Lys Cys Ser Ser Pro Gln Gln
Cys Leu Lys Pro Cys Lys Ala Ala1 5 10 15Phe Gly Ile Ser Ala Gly Gly
Lys Cys Ile Asn Gly Lys Cys Lys Cys 20 25 30Tyr49632PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
496Val Ser Cys Ser Ala Ser Ser Gln Cys Trp Pro Val Cys Lys Lys Leu1
5 10 15Phe Gly Thr Tyr Arg Gly Lys Cys Met Asn Ser Lys Cys Arg Cys
Tyr 20 25 3049732PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 497Glu Ser Cys Thr Ala Ser Asn Gln
Cys Trp Ser Ile Cys Lys Arg Leu1 5 10 15His Asn Thr Asn Arg Gly Lys
Cys Met Asn Lys Lys Cys Arg Cys Tyr 20 25 3049832PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
498Val Ser Cys Thr Thr Ser Lys Glu Cys Trp Ser Val Cys Glu Lys Leu1
5 10 15Tyr Asn Thr Ser Arg Gly Lys Cys Met Asn Lys Lys Cys Arg Cys
Tyr 20 25 3049932PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 499Met Arg Cys Lys Ser Ser Lys Glu
Cys Leu Val Lys Cys Lys Gln Ala1 5 10 15Thr Gly Arg Pro Asn Gly Lys
Cys Met Asn Arg Lys Cys Lys Cys Tyr 20 25 3050032PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
500Ile Lys Cys Thr Leu Ser Lys Asp Cys Tyr Ser Pro Cys Lys Lys Glu1
5 10 15Thr Gly Cys Pro Arg Ala Lys Cys Ile Asn Arg Asn Cys Lys Cys
Tyr 20 25 3050132PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 501Ile Arg Cys Ser Gly Ser Arg Asp
Cys Tyr Ser Pro Cys Met Lys Gln1 5 10 15Thr Gly Cys Pro Asn Ala Lys
Cys Ile Asn Lys Ser Cys Lys Cys Tyr 20 25 3050232PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
502Ile Arg Cys Ser Gly Thr Arg Glu Cys Tyr Ala Pro Cys Gln Lys Leu1
5 10 15Thr Gly Cys Leu Asn Ala Lys Cys Met Asn Lys Ala Cys Lys Cys
Tyr 20 25 3050332PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 503Ile Ser Cys Thr Asn Pro Lys Gln
Cys Tyr Pro His Cys Lys Lys Glu1 5 10 15Thr Gly Tyr Pro Asn Ala Lys
Cys Met Asn Arg Lys Cys Lys Cys Phe 20 25 3050432PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
504Ala Ser Cys Arg Thr Pro Lys Asp Cys Ala Asp Pro Cys Arg Lys Glu1
5 10 15Thr Gly Cys Pro Tyr Gly Lys Cys Met Asn Arg Lys Cys Lys Cys
Asn 20 25 3050531PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 505Thr Ser Cys Ile Ser Pro Lys Gln
Cys Thr Glu Pro Cys Arg Ala Lys1 5 10 15Gly Cys Lys His Gly Lys Cys
Met Asn Arg Lys Cys His Cys Met 20 25 3050631PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
506Lys Glu Cys Thr Gly Pro Gln His Cys Thr Asn Phe Cys Arg Lys Asn1
5 10 15Lys Cys Thr His Gly Lys Cys Met Asn Arg Lys Cys Lys Cys Phe
20 25 3050732PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 507Ile Lys Cys Arg Thr Pro Lys Asp
Cys Ala Asp Pro Cys Arg Lys Gln1 5 10 15Thr Gly Cys Pro His Ala Lys
Cys Met Asn Lys Thr Cys Arg Cys His 20 25 3050832PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
508Val Lys Cys Thr Thr Ser Lys Glu Cys Trp Pro Pro Cys Lys Ala Ala1
5 10 15Thr Gly Lys Ala Ala Gly Lys Cys Met Asn Lys Lys Cys Lys Cys
Gln 20 25 3050932PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 509Leu Glu Cys Gly Ala Ser Arg Glu
Cys Tyr Asp Pro Cys Phe Lys Ala1 5 10 15Phe Gly Arg Ala His Gly Lys
Cys Met Asn Asn Lys Cys Arg Cys Tyr 20 25 3051032PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
510Glu Lys Cys Phe Ala Thr Ser Gln Cys Trp Thr Pro Cys Lys Lys Ala1
5 10 15Ile Gly Ser Leu Gln Ser Lys Cys Met Asn Gly Lys Cys Lys Cys
Tyr 20 25 3051132PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 511Val Arg Cys Tyr Ala Ser Arg Glu
Cys Trp Glu Pro Cys Arg Arg Val1 5 10 15Thr Gly Ser Ala Gln Ala Lys
Cys Gln Asn Asn Gln Cys Arg Cys Tyr 20 25 3051232PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
512Val Lys Cys Ser Ala Ser Arg Glu Cys Trp Val Ala Cys Lys Lys Val1
5 10 15Thr Gly Ser Gly Gln Gly Lys Cys Gln Asn Asn Gln Cys Arg Cys
Tyr 20 25 3051332PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 513Val Lys Cys Ile Ser Ser Gln Glu
Cys Trp Ile Ala Cys Lys Lys Val1 5 10 15Thr Gly Arg Phe Glu Gly Lys
Cys Gln Asn Arg Gln Cys Arg Cys Tyr 20 25 3051432PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
514Val Arg Cys Tyr Asp Ser Arg Gln Cys Trp Ile Ala Cys Lys Lys Val1
5 10 15Thr Gly Ser Thr Gln Gly Lys Cys Gln Asn Lys Gln Cys Arg Cys
Tyr 20 25 3051532PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 515Val Asp Cys Thr Val Ser Lys Glu
Cys Trp Ala Pro Cys Lys Ala Ala1 5 10 15Phe Gly Val Asp Arg Gly Lys
Cys Met Gly Lys Lys Cys Lys Cys Tyr 20 25 3051632PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
516Ala Lys Cys Arg Gly Ser Pro Glu Cys Leu Pro Lys Cys Lys Glu Ala1
5 10 15Ile Gly Lys Ala Ala Gly Lys Cys Met Asn Gly Lys Cys Lys Cys
Tyr 20 25 3051731PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 517Lys Lys Cys Gln Gly Gly Ser Cys
Ala Ser Val Cys Arg Arg Val Ile1 5 10 15Gly Val Ala Ala Gly Lys Cys
Ile Asn Gly Arg Cys Val Cys Tyr 20 25 3051832PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
518Lys Lys Cys Ser Asn Thr Ser Gln Cys Tyr Lys Thr Cys Glu Lys Val1
5 10 15Val Gly Val Ala Ala Gly Lys Cys Met Asn Gly Lys Cys Ile Cys
Tyr 20 25 3051932PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 519Val Lys Cys Ser Gly Ser Ser Lys
Cys Val Lys Ile Cys Ile Asp Arg1 5 10 15Tyr Asn Thr Arg Gly Ala Lys
Cys Ile Asn Gly Arg Cys Thr Cys Tyr 20 25 3052032PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
520Asn Arg Cys Asn Asn Ser Ser Glu Cys Ile Pro His Cys Ile Arg Ile1
5 10 15Phe Gly Thr Arg Ala Ala Lys Cys Ile Asn Arg Lys Cys Tyr Cys
Tyr 20 25 3052132PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 521Lys Glu Cys Asn Gly Ser Ser Glu
Cys Tyr Ser His Cys Glu Gly Ile1 5 10 15Thr Gly Lys Arg Ser Gly Lys
Cys Ile Asn Lys Lys Cys Tyr Cys Tyr 20 25 3052229PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 522Ala
Phe Cys Asn Leu Arg Arg Cys Glu Leu Ser Cys Arg Ser Leu Gly1 5 10
15Leu Leu Gly Lys Cys Ile Gly Glu Glu Cys Lys Cys Val 20
2552329PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 523Ala Val Cys Asn Leu Lys Arg Cys Gln Leu Ser
Cys Arg Ser Leu Gly1 5 10 15Leu Leu Gly Lys Cys Ile Gly Asp Lys Cys
Glu Cys Val 20 2552430PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 524Ala Ala Cys Tyr Ser
Ser Asp Cys Arg Val Lys Cys Val Ala Met Gly1 5 10 15Phe Ser Ser Gly
Lys Cys Ile Asn Ser Lys Cys Lys Cys Tyr 20 25 3052527PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 525Ala
Ile Cys Ala Thr Asp Ala Asp Cys Ser Arg Lys Cys Pro Gly Asn1 5 10
15Pro Pro Cys Arg Asn Gly Phe Cys Ala Cys Thr 20
2552627PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 526Thr Glu Cys Gln Ile Lys Asn Asp Cys Gln Arg
Tyr Cys Gln Ser Val1 5 10 15Lys Glu Cys Lys Tyr Gly Lys Cys Tyr Cys
Asn 20 2552727PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 527Thr Gln Cys Gln Ser Val Arg Asp Cys
Gln Gln Tyr Cys Leu Thr Pro1 5 10 15Asp Arg Cys Ser Tyr Gly Thr Cys
Tyr Cys Lys 20 2552832PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 528Val Ser Cys Arg Tyr
Gly Ser Asp Cys Ala Glu Pro Cys Lys Arg Leu1 5 10 15Lys Cys Leu Leu
Pro Ser Lys Cys Ile Asn Gly Lys Cys Thr Cys Tyr 20 25
3052932PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 529Ile Lys Cys Arg Tyr Pro Ala Asp Cys His
Ile Met Cys Arg Lys Val1 5 10 15Thr Gly Arg Ala Glu Gly Lys Cys Met
Asn Gly Lys Cys Thr Cys Tyr 20 25 3053032PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
530Ile Lys Cys Ser Ser Ser Ser Ser Cys Tyr Glu Pro Cys Arg Gly Val1
5 10 15Thr Gly Arg Ala His Gly Lys Cys Met Asn Gly Arg Cys Thr Cys
Tyr 20 25 3053132PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 531Val Lys Cys Thr Gly Ser Lys Gln
Cys Leu Pro Ala Cys Lys Ala Ala1 5 10 15Val Gly Lys Ala Ala Gly Lys
Cys Met Asn Gly Lys Cys Lys Cys Tyr 20 25 3053231PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
532Val Ser Cys Lys His Ser Gly Gln Cys Ile Lys Pro Cys Lys Asp Ala1
5 10 15Gly Met Arg Phe Gly Lys Cys Met Asn Arg Lys Cys Asp Cys Thr
20 25 3053331PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 533Val Lys Cys Arg Gly Ser Pro Gln
Cys Ile Gln Pro Cys Arg Asp Ala1 5 10 15Gly Met Arg Phe Gly Lys Cys
Met Asn Gly Lys Cys His Cys Thr 20 25 3053433PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
534Val Lys Cys Thr Ser Pro Lys Gln Cys Leu Pro Pro Cys Lys Ala Gln1
5 10 15Phe Gly Ile Arg Ala Gly Ala Lys Cys Met Asn Gly Lys Cys Lys
Cys 20 25 30Tyr53533PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 535Val Lys Cys Thr Ser Pro Lys Gln
Cys Ser Lys Pro Cys Lys Glu Leu1 5 10 15Tyr Gly Ser Ser Ala Gly Ala
Lys Cys Met Asn Gly Lys Cys Lys Cys 20 25 30Tyr53633PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
536Val Lys Cys Thr Ser Pro Lys Gln Cys Leu Pro Pro Cys Lys Glu Ile1
5 10 15Tyr Gly Arg His Ala Gly Ala Lys Cys Met Asn Gly Lys Cys His
Cys 20 25 30Ser53732PRTArtificial SequenceDescription of Artificial
Sequence Synthetic polypeptide 537Val Lys Cys Thr Gly Ser Lys Gln
Cys Trp Pro Val Cys Lys Gln Met1 5 10 15Phe Gly Lys Pro Asn Gly Lys
Cys Met Asn Gly Lys Cys Arg Cys Tyr 20 25 3053831PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
538Val Lys Cys Arg Gly Ser Arg Asp Cys Leu Asp Pro Cys Lys Lys Ala1
5 10 15Gly Met Arg Phe Gly Lys Cys Ile Asn Ser Lys Cys His Cys Thr
20 25 3053927PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 539Val Arg Cys Val Thr Asp Asp Asp Cys
Phe Arg Lys Cys Pro Gly Asn1 5 10 15Pro Ser Cys Lys Arg Gly Phe Cys
Ala Cys Lys 20 2554032PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 540Val Pro Cys Asn Asn
Ser Arg Pro Cys Val Pro Val Cys Ile Arg Glu1 5 10 15Val Asn Asn Lys
Asn Gly Lys Cys Ser Asn Gly Lys Cys Leu Cys Tyr 20 25
3054136PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 541Val Pro Ile Asn Val Lys Cys Arg Gly Ser
Arg Asp Cys Leu Asp Pro1 5 10 15Cys Lys Lys Ala Gly Met Arg Phe Gly
Lys Cys Ile Asn Ser Lys Cys 20 25 30His Cys Thr Pro
3554237PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 542Val Gln Thr Asn Val Lys Cys Gln Gly Gly
Ser Cys Ala Ser Val Cys1 5 10 15Arg Arg Glu Ile Gly Val Ala Ala Gly
Lys Cys Ile Asn Gly Lys Cys 20 25 30Val Cys Tyr Arg Asn
3554338PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 543Ala Glu Ile Ile Arg Cys Ser Gly Thr Arg
Glu Cys Tyr Ala Pro Cys1 5 10 15Gln Lys Leu Thr Gly Cys Leu Asn Ala
Lys Cys Met Asn Lys Ala Cys 20 25 30Lys Cys Tyr Gly Cys Val
3554436PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 544Arg Pro Thr Asp Ile Lys Cys Ser Ala Ser
Tyr Gln Cys Phe Pro Val1 5 10 15Cys Lys Ser Arg Phe Gly Lys Thr Asn
Gly Arg Cys Val Asn Gly Leu 20 25 30Cys Asp Cys Phe
3554537PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 545Gln Phe Thr Asp Val Lys Cys Thr Gly Ser
Lys Gln Cys Trp Pro Val1 5 10 15Cys Lys Gln Met Phe Gly Lys Pro Asn
Gly Lys Cys Met Asn Gly Lys 20 25 30Cys Arg Cys Tyr Ser
3554637PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 546Val Gly Ile Asn Val Lys Cys Lys His Ser
Arg Gln Cys Leu Lys Pro1 5 10 15Cys Lys Asp Ala Gly Met Arg Phe Gly
Lys Cys Thr Asn Gly Lys Cys 20 25 30His Cys Thr Pro Lys
3554735PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 547Val Val Ile Gly Gln Arg Cys Tyr Arg Ser
Pro Asp Cys Tyr Ser Ala1 5 10 15Cys Lys Lys Leu Val Gly Lys Ala Thr
Gly Lys Cys Thr Asn Gly Arg 20 25 30Cys Asp Cys
3554835PRTArtificial SequenceDescription of Artificial Sequence
Synthetic
polypeptide 548Asn Phe Lys Val Glu Gly Ala Cys Ser Lys Pro Cys Arg
Lys Tyr Cys1 5 10 15Ile Asp Lys Gly Ala Arg Asn Gly Lys Cys Ile Asn
Gly Arg Cys His 20 25 30Cys Tyr Tyr 3554938PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
549Gln Ile Asp Thr Asn Val Lys Cys Ser Gly Ser Ser Lys Cys Val Lys1
5 10 15Ile Cys Ile Asp Arg Tyr Asn Thr Arg Gly Ala Lys Cys Ile Asn
Gly 20 25 30Arg Cys Thr Cys Tyr Pro 3555037PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
550Gly Val Pro Ile Ser Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Asn Gly
Arg 20 25 30Cys His Cys Thr Pro 3555137PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
551Gly Val Pro Ile Ser Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Gln Ser
Arg 20 25 30Cys His Cys Thr Pro 3555237PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
552Gly Val Pro Ile Ser Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Asn Arg
Arg 20 25 30Cys His Cys Thr Pro 3555337PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
553Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Asn Ser
Arg 20 25 30Cys His Cys Thr Pro 3555437PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
554Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Gln Ser
Arg 20 25 30Cys His Cys Thr Pro 3555537PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
555Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Gln Ser
Arg 20 25 30Cys His Cys Tyr Pro 3555637PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
556Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Tyr Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Gln Ser
Arg 20 25 30Cys His Cys Thr Pro 3555737PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
557Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Gln Ser
Arg 20 25 30Cys Tyr Cys Thr Pro 3555837PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
558Gly Val Pro Ile Ser Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Gln Ser
Arg 20 25 30Cys His Cys Tyr Pro 3555937PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
559Gly Val Pro Ile Ser Val Arg Cys Arg Gly Ser Arg Asp Cys Tyr Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Gln Ser
Arg 20 25 30Cys His Cys Thr Pro 3556037PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
560Gly Val Pro Ile Ser Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Gln Ser
Arg 20 25 30Cys Tyr Cys Thr Pro 3556137PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
561Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Ala Ser
Arg 20 25 30Cys His Cys Tyr Pro 3556237PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
562Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Ser Ser
Arg 20 25 30Cys His Cys Tyr Pro 3556337PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
563Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Thr Ser
Arg 20 25 30Cys His Cys Tyr Pro 3556437PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
564Gly Val Pro Ile Asn Val Arg Cys Arg Gly Ser Arg Asp Cys Leu Glu1
5 10 15Pro Cys Arg Arg Ala Gly Thr Arg Phe Gly Arg Cys Ile Asn Ser
Arg 20 25 30Cys His Cys Tyr Pro 35565292PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
565Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro1
5 10 15Gly Ser Thr Gly Gly Ser Gly Val Pro Ile Asn Val Arg Cys Arg
Gly 20 25 30Ser Arg Asp Cys Leu Asp Pro Cys Arg Arg Ala Gly Met Arg
Phe Gly 35 40 45Arg Cys Ile Asn Ser Arg Cys His Cys Thr Pro Gly Gly
Ser Gly Gly 50 55 60Ser Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala
Pro Glu Leu Leu65 70 75 80Gly Gly Pro Ser Val Phe Leu Phe Pro Pro
Lys Pro Lys Asp Thr Leu 85 90 95Met Ile Ser Arg Thr Pro Glu Val Thr
Cys Val Val Val Asp Val Ser 100 105 110His Glu Asp Pro Glu Val Lys
Phe Asn Trp Tyr Val Asp Gly Val Glu 115 120 125Val His Asn Ala Lys
Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr 130 135 140Tyr Arg Val
Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn145 150 155
160Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro
165 170 175Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu
Pro Gln 180 185 190Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr
Lys Asn Gln Val 195 200 205Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr
Pro Ser Asp Ile Ala Val 210 215 220Glu Trp Glu Ser Asn Gly Gln Pro
Glu Asn Asn Tyr Lys Thr Thr Pro225 230 235 240Pro Val Leu Asp Ser
Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 245 250 255Val Asp Lys
Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val 260 265 270Met
His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu 275 280
285Ser Pro Gly Lys 29056620PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 566Met Glu Thr Asp Thr Leu
Leu Leu Trp Val Leu Leu Leu Trp Val Pro1 5 10 15Gly Ser Thr Gly
2056739PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 567Gly Ser Gly Val Pro Ile Asn Val Arg Ser
Arg Gly Ser Arg Asp Ser1 5 10 15Leu Asp Pro Ser Arg Arg Ala Gly Met
Arg Phe Gly Arg Ser Ile Asn 20 25 30Ser Arg Ser His Ser Thr Pro
355684PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 568Gly Ala Gly Ala15694PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 569Asp
Glu Val Asp15704PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 570Leu Glu His Asp1
* * * * *