ANTIBODIES AGAINST HUMAN IL-22 AND USES THEREFOR
CROSS REFERENCE TO RELATED APPLICATIONS
TECHNICAL FIELD
[0001]This invention relates to antibodies, e.g., human antibodies, and
antigen-binding fragments thereof that bind interleukin-22 (IL-22), in particular,
human IL-22, and their use in regulating IL-22-associated activities. The antibodies
disclosed herein are useful in diagnosing, preventing, and/or treating IL-22
associated disorders, e.g., autoimmune disorders, including arthritis.
BACKGROUND OF THE INVENTION
[0002]Antigens initiate immune responses and activate the two largest
populations of lymphocytes: T cells and B cells. After encountering antigen, T cells
proliferate and differentiate into effector cells, while B cells proliferate and
differentiate into antibody-secreting plasma cells. These effector cells secrete
and/or respond to cytokines, which are small proteins (< about 30 kDa) secreted by
lymphocytes and other cell types.
[0003]Interleukin-22 (IL-22) is a class II cytokine that shows sequence
homology to IL-10. its expression is up-regulated in T cells by IL-9 or ConA
(Dumoutier L. et al. (2000) Proc Natl Acad Sci USA 97(18):10144-9). Further
studies have shown that expression of IL-22 mRNA is induced in vivo in response
to LPS administration, and that IL-22 modulates parameters indicative of an acute
phase response (Dumoutier L. et al. (2000); Pittman D. et al. (2001) Genes and
Immunity 2:172). In addition, IL-22 enhances the expression of antimicrobial
peptides associated with host defense, including β-defensin, S100A7, S100A8, and
S100A. Wolk et al., Immunity, 21:241-54 (2004); Boniface et al., J. Immunol.
174:3695-3702 (2005); Liang et al., J. Exp. Med., 203(10):2271-79 (2006). Taken
together, these observations indicate that IL-22 plays a role in inflammation
(Kotenko S.V. (2002) Cytokine & Growth Factor Reviews 13(3):223-40).
[0004JIL-22 is believed to bind to a receptor complex consisting of IL-22R
and IL-10R2, two members of the type II cytokine receptor family (CRF2) (Xie M.H.
et al. (2000) J Biol Chem 275(40):31335-9; Kotenko S.V. et al. (2001) J Biol Chem
276(4):2725-32). Both chains of the IL-22 receptor are expressed constitutively in
a number of organs. Epithelial cell lines derived form these organs are responsive

to IL-22 in vitro (Kotenko S.V. (2002) Cytokine & Growth Factor Reviews
13(3):223-40). IL-22 induces activation of the JAK/STAT3 and ERK pathways, as
well as intermediates of other MAPK pathways (Dumoutier L. et al. (2000) supra;
Xie M.H. et al. (2000) supra; Dumoutier L. et al. (2000) J Immunol 164(4):1814-9;
Kotenko S.V. et al. (2001) J Bio/ Chem 276(4):2725-32; Lejeune, D. et al. (2002) J
Biol Chem 277(37):33676-82).
[0005]CRF2 members are receptors for IFNα/β, IFNγ, coagulation factor
VIla, IL-10 and the IL-10 related proteins IL-19, IL-20, IL-22, IL-24, IL-26, as well as
the recently identified IFN-like cytokines, IL-28 and IL-29 (Kotenko S.V. (2002)
Cytokine & Growth Factor Reviews 13(3):223-40; Kotenko, S.V. et al. (2000)
Oncogene 19(21):2557-65; Sheppard, P. etal. (2003) Nature Immunology 4(1):63-
8; Kotenko, S.V. et al. (2003) Nature Immunology 4(1):69-77). In addition to these
membrane receptors, the CRF2 family also includes a soluble protein, IL-22
binding protein (IL-22BP), which is specific for IL-22 and blocks its activity
(Dumoutier, L. et al. (2001) J Immunol 166(12):7090-5; Kotenko, S.V. et al. (2001)
J /mmunol166(12):709G-103; Xu, W. etal. (2001) ProcNatlAcadSdUSA
98(17):9511-6; Gruenberg, B.H. etal. (2001) Genes & Immunity2(6):329-34; Wei
C-C et al. (2003) Genes & Immunity 4:204-211). While the IL-22 receptor complex
is unique for IL-22, each chain (i.e., IL-22R and IL-10R2) is shared with other CRF2
members to define functional receptors for other cytokines, including IL-20, IL-24
(IL-22R/IL-20R2), IL-28, IL-29 (IFN-A.R1/IL-10R2) and IL-10 (IL-10R1/IL-10R2)
(Dumoutier, L. et al. (2001) J. Immunol. 167(7):3545-9; Wang, M. et al. (2002) J
Biol Chem 277(9):7341 -7; Parrish-Novak, J. et al. (2002) J Biol Chem
277(49):47517-23; Kotenko, S.V. et al. (1997) EMBOJ. 16(19):5894-903; Spencer,
S.D. et al. (1998) J Exp Med 187(4):571-8).
[0006jBoth chains of the CRF2-composed receptor are necessary for signal
transduction. One chain of the composed receptor has been historically defined as
a ligand binding chain (e.g., IFNγRI) based on its high affinity for the cytokine. The
other chain (e.g., IFNγR2) has been characterized as a helper or accessory chain,
and shows minimal affinity for the cytokine alone (Kotenko, S.V. et al. (2000)
Oncogene 19(21):2557-65). For IL-22, IL-22R is the high affinity receptor subunit
with IL-10R2 subsequently binding to the IL-22/IL-22R complex (Li, J. et al. (2004)

Int. Immunopharmacol. 4(5):673-708; Logsdon, N. J. et al. (2002) J. Interferon
Cytokine Res 22(11):1099-1112).
SUMMARY OF THE INVENTION
[0007]The present application provides, at least in part, IL-22 binding agents
such as antibodies and antigen-binding fragments thereof that bind to interleukin-
22 ("IL-22"), in particular, human IL-22, with high affinity and specificity. The
antibodies and antigen-binding fragments thereof of the present invention are also
referred to herein as "anti-IL-22 antibodies" and "fragments thereof," respectively.
In one embodiment, the antibody or fragment thereof reduces, inhibits, or
antagonizes IL-22 activity. Such antibodies can be used to regulate immune
responses or IL-22-associated disorders by antagonizing IL-22 activity. In other
embodiments, the anti-IL-22 antibody can be used diagnostically, or as a targeting
antibody to deliver a therapeutic or a cytotoxic agent to an IL-22-expressing cell.
Thus, the anti-IL-22 antibodies of the invention are useful in diagnosing, treating,
and/or preventing IL-22-associated disorders, e.g., autoimmune disorders, e.g.,
arthritis (including rheumatoid arthritis, juvenile rheumatoid arthritis, osteoarthritis,
psoriatic arthritis, lupus-associated arthritis or ankylosing spondylitis), scleroderma,
systemic lupus erythematosis, HIV, Sjogren's syndrome, vasculitis, multiple
sclerosis, autoimmune thyroiditis, dermatitis (including atopic dermatitis and
eczematous dermatitis), myasthenia gravis, inflammatory bowel disease (IBD),
Crohn's disease, colitis, diabetes mellitus (type I); inflammatory conditions of, e.g.,
the skin (e.g., psoriasis), cardiovascular system (e.g., atherosclerosis), nervous
system (e.g., Alzheimer's disease), liver (e.g., hepatitis), kidney (e.g., nephritis) and
pancreas (e.g., pancreatitis); cardiovascular disorders, e.g., cholesterol metabolic
disorders, oxygen free radical injury, ischemia; disorders associated with wound
healing; respiratory disorders, e.g., asthma and COPD (e.g., cystic fibrosis); acute
inflammatory conditions (e.g., endotoxemia, sepsis and septicaemia, toxic shock
syndrome and infectious disease); transplant rejection and allergy. In one
embodiment, the IL-22-associated disorder is, an arthritic disorder, e.g., a disorder
chosen from one or more of rheumatoid arthritis, juvenile rheumatoid arthritis,
osteoarthritis, psoriatic arthritis, or ankylosing spondylitis; a respiratory disorder
(e.g., asthma, chronic obstructive pulmonary disease (COPD); or an inflammatory
condition of, e.g., the skin (e.g., psoriasis), cardiovascular system (e.g.,

atherosclerosis), nervous system (e.g., Alzheimer's disease), liver (e.g., hepatitis),
kidney (e.g., nephritis), pancreas (e.g., pancreatitis), and gastrointestinal organs,
e.g., colitis, Crohn's disease and IBD.
[0008]Accordingly, in one aspect, the invention features an isolated antibody
that binds to IL-22, in particular, human IL-22. In certain embodiments, the anti-IL-
22 antibody may have at least one of the following characteristics: (1) it is a
monoclonal or single specificity antibody; (2) it is a human antibody; (3) it is an in
vitro generated antibody; (4) it is an in vivo generated antibody (e.g., transgenic
mouse system); (5) it binds to IL-22 with an association constant of at least 1012 M-
1; (6) it binds to IL-22 with an association constant of at least 1011 M-1 (7) it binds to
IL-22 with an association constant of at least 1010 M-1; (8) it binds to IL-22 with an
association constant of at least 109 M-1; (9) it binds to IL-22 with an association
constant of at least 106 M-1; (10) it binds to IL-22 with a dissociation constant of 500
nM or less; (11) it binds to IL-22 with a dissociation constant of 10 nM or less; (12)
it binds to IL-22 with a dissociation constant of 150 pM or less; (13) it binds to IL-22
with a dissociation constant of 60 pM or less; (14) it inhibits binding of IL-22 to IL-
22R or a receptor complex of IL-22R and IL-10R2 with an ICso of 10 nM or less;
(15) it blocks IL-22 mediated proliferation of IL-22 receptor engineered BaF3 cells
with an IC50 of 1 nM or less in one embodiment, with an IC50 of 150 pM or less in
another embodiment, with an IC50 of 100 pM or less in another embodiment, and
with an IC50 of 10 pM or less in another embodiment; and (16) it blocks IL-22
mediated GROa secretion from HT29 cells with an IC50 of 1 nM or less in one
embodiment, with an IC50 of 150 pM or less in another embodiment, and with an
IC50 of 10 pM or less in another embodiment. IL-22 mediated BaF3 cell
proliferation and IL-22 mediated GROa secretion from HT29 cells can be measured
as described in the examples.
[0009]Nonlimiting illustrative embodiments of the antibodies of the invention
are referred to herein as "GIL01," "GIL16," "GIL45," "GIL60," "GIL68," "GIL92,"
°097D09," "062A09," "062G05," "087B03," "367D04," "368D04," "166B06,"
"166G05," "375G06," "376B10," "354A08," "355B06," "355E04," and "356A11."
These antibodies can be germlined or non-germlined. In another embodiment, the
antibody is chosen from 356A11, 354A08, 087B03, and 368D04. The antibodies of
the invention may specifically bind to the same IL-22 epitope or a similar epitope

(e.g., an overlapping epitope) that GIL01, GIL16, GIL45, GIL60, GIL68, GIL92,
097D09, 062A09, 062G05, 087B03, 367D04, 368D04, 166B06, 166G05, 375G06,
376B10, 354A08, 355B06, 355E04, or 356A11 binds to. In other embodiments, the
antibodies specifically bind to a fragment of an IL-22, e.g., a fragment of at least
10, 20, 50, 75, 100, 150, or 200 amino acids contiguous to the amino acid
sequence set forth in SEQ ID NO:1, or a sequence that is at least 85%, 90%, 95%,
96%, 97%, 98%, 99% or more identical thereto. In other embodiments, the
antibody competitively inhibits the binding of at least one of GIL01, GIL16, GIL45,
GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03, 367D04, 368D04,
166B06, 166G05, 375G06, 376B10, 354A08, 355B06, 355E04, or 356A11 to its
target epitope.
[0010]In one embodiment, the antibody of the present invention includes a
VH domain, VL domain, or combination thereof, of the Fv fragment of GIL01, G1L16,
GIL45, GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03, 367D04,
368D04, 166B06, 166G05, 375G06, 376B10, 354A08, 355B06, 355E04, or
356A11. For example, the antibody includes a VH and/or a VL domain having
amino add sequence as set forth in Tables 1 and 7 (SEQ ID NO:5, 23, 41, 59, 77,
95,113, 131, 149, 167, 185, 203, 221, 239, 257, 275, 293, 311, 329, 347, 365,
383, 401,419, 437,455, 473, 491, 509, 527, 545, 563, 581, 599, or 617 for VH and
SEQ ID NO:6, 24, 42, 60, 78, 96, 114, 132,150, 168, 186, 204, 222, 240, 258, 276,
294, 312, 330, 348, 366, 384, 402, 420, 438, 456, 474, 492, 510, 528, 546, 564,
582, 600, or 618 for VL), or a sequence substantially identical thereto (e.g., a
sequence at least about 85%, 90%, 95%, 96%, 97%, 98%, 99% or more identical
thereto, or which differs by no more than 1, 2, 5, 10 or 15 amino acid residues from
SEQ ID NO:5, 6, 23, 24, 41, 42, 59, 60, 77, 78, 95, 96, 113, 114,131,132,149,
150, 167, 168, 185, 186, 203, 204, 221, 222, 239, 240, 257, 258, 275, 276, 293,
294, 311,312, 329, 330, 347, 348, 365, 366, 383, 384, 401, 402, 419, 420,437,
438, 455,456, 473, 474, 491, 492, 509, 510, 527, 528, 545, 546, 563, 564, 581,
582, 599,600, 617, or 618).
[0011] ln another embodiment, the antibody of the present invention includes
a VH domain, VL domain, or combination thereof, of the Fv fragment of an antibody
chosen from 356A11,354A08, 087B03, and 368D04. In this embodiment, the
antibody, or antigen-binding fragment thereof, comprises:

[0012] a VH domain comprising the amino acid sequence set out in SEQ ID
NO:167 or 491 and/or a VL domain comprising the amino acid sequence set out in
SEQ ID NO:168 or 492 (087B03);
[0013]a VH domain comprising the amino acid sequence set out in SEQ ID
NO:293 or 545 and/or a VL domain having the amino acid sequence set out in SEQ
ID NO:294 or 546 (354A08);
[0014]a VH domain comprising the amino acid sequence set out in SEQ ID
NO:203 or 617 and/or a VL domain comprising the amino acid sequence set out in
SEQ ID NO:204 or 618 (368D04); or
[0015]a VH domain comprising the amino acid sequence set out in SEQ ID
NO:347 or 599 and/or a VL domain comprising the amino acid sequence set out in
SEQ ID NO:348 or 600 {356A11).
[0016]In another embodiment, the antibody includes a VH and/or VL domain
encoded by a nucleic acid having a nucleotide sequence as set forth in Tables 1
and 7 (SEQ ID NO:14, 32, 50, 68, 86,104,122,140,158, 176, 194, 212, 230, 248,
266, 284, 302, 320, 338, 356, 374, 392, 410, 428, 446, 464, 482, 500, 518, 536,
554, 572, 590, 608, or 626 for VH and SEQ ID NO: 15, 33, 51, 69, 87,105,123,
141,159,177, 195, 213, 231, 249, 267, 285, 303, 321, 339, 357, 375, 393, 411,
429, 447, 465, 483, 501, 519, 537, 555, 573, 591, 609, or 627 for V), or a
sequence substantially identical thereto (e.g., a sequence at least about 85%, 90%,
95%, 96%, 97%, 98%, 99% or more identical thereto, or which differs by no more
than 1, 2, 3, 6, 15, 30 or 45 nudeotides from SEQ ID NO: 14, 15,32,33, 50, 51,
68, 69, 86, 87, 104, 105, 122, 123, 140, 141, 158, 159,176, 177, 194, 195, 212,
213, 230, 231, 248, 249, 266, 267, 284, 285, 302, 303, 320, 321, 338, 339, 356,
357, 374, 375, 392, 393, 410, 411, 428, 429, 446, 447, 464, 465, 482, 483, 500,
501, 518, 519, 536, 537, 554, 555, 572, 573, 590, 591, 608, 609, 626, Of 627).
[0017] In other embodiments, the antibody includes an Fv domain having an
amino acid sequence as set forth in Tables 1 and 7 (SEQ ID NO:7,25, 43, 61,79,
97, 115, 133, 151, 169, 187, 205, 223, 241, 259,277, 295, 313, 331, 349, 367,
385, 403, 421,439,457, 475, 493, 511, 529, 547, 565, 583, 601, or 619), or a
sequence substantially identical thereto (e.g., a sequence at least about 85%, 90%,
95%, 96%, 97%, 98%, 99% or more identical thereto, or which differs by no more
than 1, 2, 5, 10,15,20, 30 or 35 amino acid residues from SEQ ID NO:7, 25,43,

61, 79, 97,115, 133, 151,169, 187, 205, 223, 241, 259, 277, 295, 313, 331, 349,
367, 385, 403, 421, 439, 457, 475, 493, 511, 529, 547, 565, 583, 601, or 619). In
another embodiment, the antibody of the present invention includes an Fv domain
of an antibody chosen from 356A11 (SEQ ID NO:349 or 601), 354A08 (SEQ ID
NO:295 or 547), 087B03 (SEQ ID NO:169 or 493), and 368D04 (SEQ ID NO:205
or 619). in another embodiment, the antibody includes an Fv domain encoded by a
nucleic acid having a nucleotide sequence as set forth in Tables 1 and 7 (SEQ ID
NO:16, 34, 52, 70, 88, 106, 124, 142, 160, 178, 196, 214, 232, 250, 268, 286, 304,
322, 340, 358, 376, 394, 412, 430, 448, 466, 484, 502, 520,538, 556, 574, 592,
610, or 628), or a sequence substantially identical thereto (e.g., a sequence at
least about 85%, 90%, 95%, 96%, 97%, 98%, 99% or more identical thereto, or
which differs by no more than 1, 2, 3, 6,15, 30, 45, 60, 90 or 105 nucieotides from
SEQ ID NO: 16, 34, 52, 70, 88, 106,124,142, 160, 178,196, 214, 232, 250, 268,
286, 304, 322, 340, 358, 376, 394, 412, 430, 448, 466, 484, 502, 520, 538, 556,
574, 592,610, or 628). In yet other embodiments, the antibody comprises at least
one complementarity determining region (CDR) of these VH and VL domains. For
example, the antibody can include one, two, or three CDR's of the VH domain
having an amino acid sequence as set forth in or included within the sequences in
Tables 1 and 7 (SEQ ID NO:5, 7, 8, 9, 10, 23, 25, 26, 27, 28, 41, 43, 44, 45, 46, 59,
61, 62, 63, 64, 77, 79, 80, 81, 82, 95, 97, 98, 99, 100, 113,115, 116, 117, 118,
131, 133,134,135, 136, 149,151, 152, 153,154, 167, 169,170, 171, 172, 185,
187, 188, 189, 190, 203, 205, 206, 207, 208, 221, 223, 224, 225, 226, 239, 241,
242, 243, 244, 257, 259, 260, 261, 262, 275, 277, 278, 279. 280, 293, 295, 296,
297, 298, 311, 313, 314, 315, 316, 329, 331,332, 333, 334,347, 349, 350, 351,
352, 365, 367, 368, 369, 370, 383, 385, 386, 387, 388, 401,403, 404, 405, 406,
419, 421, 422, 423, 424, 437, 439, 440, 441, 442, 455, 457, 458, 459, 460, 473,
475, 476, 477, 478, 491, 493, 494, 495, 496, 509, 511, 512, 513, 514, 527, 529,
530, 531, 532, 545, 547, 548, 549, 550, 563, 565, 566, 567, 568, 581, 583, 584,
585, 586, 599, 601, 602, 603, 604, 617, 619, 620, 621, or 622), or a sequence
substantially homologous thereto (e.g., a sequence at least about 85%, 90%, 95%,
96%, 97%, 98%, 99% or more identical thereto). In another embodiment, the
antibody of the present invention includes one, two, or three CDR's of the VH
domain of an antibody chosen from 356A11,354A08, 087B03, and 368D04. In this

embodiment, the antibody, or antigen-binding fragment thereof, comprises a heavy
chain variable region comprising:
[0018]a) SEQ ID NO: 170 or494; b) SEQ ID NO: 171 or495; and/or c) SEQ
ID NO: 172 or 496 (087B03);
[0019Ja) SEQ ID NO:296 or 548; b) SEQ ID NO:297 or 549; and/or c) SEQ
ID NO:298 or 550 (354A08);
[0020]a) SEQ ID NO:206 or 620; b) SEQ ID NO:207 or 621; and/or c) SEQ
ID NO:208 or 622 (368D04); or
[0021 ]a) SEQ ID NO:350 or 602; b) SEQ ID NO:351 or 603; and/or c) SEQ
ID NO:352 or 604 (356A11).
[0022] In another embodiment, the antibody can include one, two, or three
CDR's of the VL domain having an amino acid sequence as set forth in or included
within the sequences in Tables 1 and 7 (SEQ ID NO:6, 7, 11,12,13, 24, 25, 29,
30, 31, 42, 43, 47, 48, 49, 60, 61, 65, 66, 67, 78, 79, 83, 84, 85, 96, 97, 101, 102,
103, 114, 115, 119, 120, 121, 132, 133, 137, 138, 139, 150,151, 155, 156, 157,
168, 169, 173, 174, 175, 186, 187, 191, 192, 193, 204, 205,209, 210, 211, 222,
223, 227, 228, 229, 240, 241, 245, 246, 247, 258, 259, 263, 264, 265. 276, 277,
281, 282, 283, 294, 295, 299, 300, 301, 312, 313, 317, 318, 319, 330, 331, 335,
336, 337, 348, 349, 353, 354, 355, 366, 367, 371, 372, 373, 384, 385. 389, 390,
391, 402, 403, 407, 408, 409, 420, 421, 425, 426, 427, 438, 439, 443, 444, 445,
456, 457, 461, 462, 463, 474, 475, 479, 480, 481, 492, 493, 497, 498, 499, 510,
511, 515, 516, 517, 528, 529, 533, 534, 535, 546, 547, 551, 552, 553, 564, 565,
569, 570, 571, 582, 583, 587, 588, 589. 600, 601, 605, 606, 607, 618, 619, 623,
624, or 625), or a sequence substantially identical thereto (e.g., a sequence at
least about 85%, 90%, 95%, 96%, 97%, 98%, 99% or more identical thereto). In
another embodiment, the antibody of the present invention includes one, two, or
three CDR's of the VL domain of an antibody chosen from 356A11,354A08,
087B03, and 368D04. In this embodiment, the antibody, or antigen-binding
fragment thereof, comprises a light chain variable region comprising:
[0023]a) SEQ ID NO: 173 or 497; b) SEQ ID NO: 174 or 498; and/or c) SEQ
ID NO: 175 or 499 (087B03);
[0024]a) SEQ ID NO-299 or 551; b) SEQ ID NO:300 or 552; and/or c) SEQ
ID NO 301 or 553 (354A08);

[0025]a) SEQ ID NO:209 or 623; b) SEQ ID NO:210 or 624; and/or c) SEQ
ID NO:211 or 625 (368D04); or
[0026]a) SEQ ID NO.353 or 605; b) SEQ ID NO.354 or 606; and/or c) SEQ
ID NO:355 or 607 (356A11).
[0027] In a still further embodiment, the antibody comprises an H3 fragment
of the VH domain of GIL01, GIL16, GIL45, GIL60, GIL68, GIL92, 097D09, 062A09,
062G05, 087B03, 367D04, 368D04, 166B06, 166G05, 375G06, 376B10, 354A08,
355B06, 355E04, or 356A11, e.g., an H3 fragment having the amino acid
sequence as set forth in Tables 1 and 7 (SEQ ID NO:10, 28, 46, 64, 82,100, 118,
136, 154, 172, 190, 208, 226, 244, 262, 280, 298, 316, 334, 352, 370, 388, 406,
424, 442, 460,478, 496, 514, 532, 550, 568, 586, 604, or 622), or a sequence
substantially identical thereto (e.g., a sequence at least about 85%, 90%, 95%,
96%, 97%, 98%, 99% or more identical thereto).
[0028]The antibody of the invention can be full-length (e.g., include at least
one complete heavy chain and at least one complete light chain) or can include
only an antigen-binding fragment (e.g., a Fab, F(ab')2, Fv, a single chain Fv
fragment, a Fd fragment, or a dAb fragment). The antibody can include a constant
region, or a portion thereof, chosen from any of: the kappa, lambda, alpha,
gamma, delta, epsilon and mu constant region genes. For example, heavy chain
constant regions of the various isotypes can be used, including: IgG1, lgG2, lgG3,
lgG4, IgM, IgA1, lgA2, IgD, and IgE. The light chain constant region can be chosen
from kappa or lambda. The antibody may be an IgG, or it may also be IgG1k or
IgG1y.
[0029] The anti-IL-22 antibody described herein can be derivatized or linked
to another functional molecule (such as another peptide or protein (e.g., a Fab
fragment)). For example, an antibody of the invention can be functionally linked
(e.g., by chemical coupling, genetic fusion, non-covalent association or otherwise)
to at least one other molecular entity, such as another antibody (e.g., a bispecific or
a multispecific antibody), toxin, radioisotope, cytotoxic or cytostatic agent, among
others.
[0030]In another aspect, the invention features a pharmaceutical
composition containing at least one anti-IL-22 antibody and a pharmaceutically
acceptable carrier. The pharmaceutical composition can further include a

combination of at (east one anti-IL-22 antibody and at least one therapeutic agent
(e.g., cytokine and growth factor inhibitors, immunosuppressants, anti-inflammatory
agents, metabolic inhibitors, enzyme inhibitors, cytotoxic agents, cytostatic agents,
or combinations thereof, as described in more detail herein). Combinations of the
anti-IL-22 antibody and a therapeutic agent are also within the scope of the
invention. The compositions and combinations of the invention can be used to
regulate IL-22-associated inflammatory conditions, e.g., by modulating IL-22
signaling through its receptors located on epithelial cells of a variety of tissues,
including, but not limited to, those of the pancreas, skin, lung, gut, liver, kidney,
salivary gland, and vascular endothelia, in addition to potentially activated and
tissue localized immune cells.
[0031]ln another aspect, the invention features a method of treating a
subject with an IL-22-associated disorder. The method includes administering to
the subject an anti-IL-22 antibody in an amount sufficient to inhibit at least one IL-
22 activity of immune cells, thereby treating the IL-22-associated disorder.
[0032]The anti-IL-22 antibody can be administered to the subject, alone or
in combination, with other therapeutic agents as described herein. The subject
may be a mammal, e.g. human. For example, the method can be used to treat a
subject with an IL-22-associated disorder such as autoimmune disorders, e.g.,
arthritis (including rheumatoid arthritis, juvenile rheumatoid arthritis, osteoarthritis,
psoriatic arthritis, lupus-associated arthritis or ankylosing spondylitis), scleroderma,
systemic lupus erythematosis, HIV, Sjogren's syndrome, vasculitis, multiple
sclerosis, autoimmune thyroiditis, dermatitis (including atopic dermatitis and
eczematous dermatitis), myasthenia gravis, inflammatory bowel disease (IBD),
Crohn's disease, colitis, diabetes mellitus (type I); inflammatory conditions of, e.g.,
the skin (e.g., psoriasis), cardiovascular system (e.g., atherosclerosis), nervous
system (e.g., Alzheimer's disease), liver (e.g., hepatitis), kidney (e.g., nephritis) and
pancreas (e.g., pancreatitis); cardiovascular disorders, e.g., cholesterol metabolic
disorders, oxygen free radical injury, ischemia; disorders associated with wound
healing; respiratory disorders, e.g., asthma and COPD (e.g., cystic fibrosis); acute
inflammatory conditions (e.g., endotoxemia, sepsis and septicaemia, toxic shock
syndrome and infectious disease); transplant rejection and allergy. In one
embodiment, the IL-22-associated disorder is, an arthritic disorder, e.g., a disorder

chosen from one or more of rheumatoid arthritis, juvenile rheumatoid arthritis,
osteoarthritis, psoriatic arthritis, or ankylosing spondylitis; a respiratory disorder
(e.g., asthma, chronic obstructive pulmonary disease (COPD); or an inflammatory
condition of, e.g., the skin (e.g., psoriasis), cardiovascular system (e.g.,
atherosclerosis), nervous system (e.g., Alzheimer's disease), liver (e.g., hepatitis),
kidney (e.g., nephritis), pancreas (e.g., pancreatitis), and gastrointestinal organs,
e.g., colitis, Crohn's disease and IBD.
[0033]ln another aspect, the invention features a method of decreasing,
inhibiting or reducing an acute phase response in a subject. The method includes
administering to the subject an IL-22 binding agent, e.g., an IL-22 antagonist, (e.g.,
an anti-IL-22 antibody or fragment thereof as described herein), in an amount
sufficient to decrease, inhibit or reduce the acute phase response in the subject. In
one embodiment, the subject is a mammal, e.g., a human suffering from an IL-22-
associated disorder, including, e.g., respiratory disorders, inflammatory disorders
and autoimmune disorders. In one embodiment, the IL-22 binding agent is
administered locally, e.g., topically, subcutaneously, or other administrations that
are not in the general circulation.
[0034]ln another aspect, an IL-22 binding agent can be used to alter the
type of immune respone and/or increase the efficacy of a vaccine formulation used
to immunize a subject. For example, an anti-IL-22 antibody of the present
invention can be administered before, during and/or after an immunization to
increase vaccine efficacy. In one embodiment, the vaccine formulation contains
one or more IL-22 antagonists and an antigen, i.e., an immunogen, including, for
example, viral, bacterial, or tumor antigens. In another embodiment, the IL-22
antagonist and the immunogen are administered separately, e.g., within one hour,
three hours, one day or two days of each other.
[0035] In another aspect, the invention provides a method for detecting the
presence of IL-22 in a sample in vitro. Samples may include biological samples
such as serum, plasma, tissue and biopsy. The subject method can be used to
diagnose a disorder, such as an IL-22-associated disorder as described herein.
The method includes: (1) contacting the sample or a control sample with an anti-IL-
22 antibody, and (2) detecting formation of a complex between the anti-IL-22
antibody and the sample or the control sample, wherein a statistically significant

change in the formation of the complex in the sample relative to a control sample,
is indicative of the presence of the IL-22 in the sample.
[0036]In another aspect, the invention provides a method for detecting the
presence of IL-22 in vivo (e.g., in vivo imaging in a subject). The method can be
used to diagnose a disorder, e.g., an IL-22-associated disorder as described
herein. The method includes: (1) administering an anti-IL-22 antibody to a subject
or a control subject under conditions that allow binding of the antibody to IL-22, and
(2) detecting formation of a complex between the antibody and IL-22, wherein a
statistically significant change in the formation of the complex in the subject relative
to a control, e.g., a control subject, is indicative of the presence of IL-22.
[0037]The antibody may be directly or indirectly labeled with a detectable
substance to facilitate detection of the bound or unbound antibody. Suitable
detectable substances include various enzymes, prosthetic groups, fluorescent
materials, luminescent materials and radioactive materials.
[0038]In another aspect, the invention provides a method for delivering or
targeting an agent, e.g., a therapeutic or a cytotoxic agent, to an IL-22-expressing
cell in vivo. The method includes administering an anti-IL-22 antibody to a subject
under conditions that allow binding of the antibody to IL-22. The antibody may be
coupled to a second therapeutic moiety, such as a toxin.
[0039]The disclosure provides nucleic acid sequences from the VH and VL
domains of GIL01, GIL16, GIL45, GIL60, GIL68, GIL92, 097D09, 062A09, 062G05,
087B03, 367D04, 368D04,166B06, 166G05, 375G06, 376B10, 354A08, 355B06,
355E04, and 356A11. Also provided are nucleic acid sequences that comprise at
least one CDR from GIL01, GIL16, GIL45, GIL60, GIL68, GIL92, 097D09, 062A09,
062G05, 087B03, 367D04, 368D04, 166B06, 166G05, 375G06, 376B10, 354A08,
355B06, 355E04, and 356A11. The disclosure also provides vectors and host cells
comprising such nucleic acids.
[0040]The disclosure further provides methods of producing new VH and VL
domains and functional antibodies comprising all or a portion of such domains
derived from the VH or VLdomains of GIL01, GIL16, GIL45, GIL60, GIL68, GIL92,
097D09, 062A09, 062G05, 087B03, 367D04, 368D04,166B06, 166G05, 375G06,
376B10, 354A08, 355B06, 355E04, or 356A11.

[0041]Additional aspects of the disclosure will be set forth in part in the
description, and in part will be obvious from the description, or may be learned by
practicing the invention. The invention is set forth and particularly pointed out in
the claims, and the disclosure should not be construed as limiting the scope of the
claims. The following detailed description includes exemplary representations of
various embodiments of the invention, which are not restrictive of the invention as
claimed. The accompanying figures constitute a part of this specification and,
together with the description, serve only to illustrate embodiments and not limit the
invention.
BRIEF DESCRIPTION OF THE FIGURES
[0042] Figure 1. Potency of parent anti-IL-22 scFv clones in the IL-22
receptor complex assay: bio.IL.-22 binding IL-22 receptor complex DELFIA
competition assay.
[0043]Figure 2. Profiling of lead scFv clones in IL-22 receptor complex
assay: bio.IL-22 binding IL-22 receptor complex DELFIA competition assay. (A)
GIL 1 derived. (B) GIL 16 derived. (C) GIL 16, GIL 60, and GIL 68 derived. (D)
GIL 60 derived. (E) GIL 68 derived. (F) GIL 63 derived. (G) GIL 92 derived.
[0044]Figure 3. IgG potency in GROa cell based assays. Optimized GIL-
IgGs in hulL-22 GROa assay. (A) Germlined IgG. (B) Non-germlined IgG.
[0045]Figure 4. Cross species reactivity of IL-22 antibodies by ELISA.
Optimized GIL-lgGs specifically bind to IL-22. (A) Germlined IgG. (B) Non-
germlined IgG.
[0046]Figure 5. Amino acid and nucleotide sequences of human IL-22. The
nucleotide sequence of human IL-22 is SEQ ID NO:2 and includes a poly (A) tail.
The disclosed nucleotide sequence includes an open reading frame and the amino
acid sequence of full-length IL-22 protein corresponding to the foregoing nucleotide
sequence is reported in SEQ ID NO:1. The amino acid sequence of mature IL-22
corresponds to about amino acids 34-179 of SEQ ID NO.1.
[0047] Figure 6. Amino acid and nucleotide sequences of mouse IL-22.
[0048]Figure 7. Amino acid and nucleotide sequences of non-germlined
GIL01, GIL16, GIL45, GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03,
367D04, 368D04,166B06, 166G05, 375G06, 376B10, 354A08, 355B06, 355E04,

and 356A11, including VH and VL domains, and CDRs (H1, H2, H3, L1, L2, and
L3).
[0049] Figure 8. Amino acid and nucleotide sequences of germlined GIL01,
GIL16, GIL45, GIL60, GIL68, GIL92, 062A09, 087B03, 166B06, 166G05, 354A08,
355B06,355E04, 356A11, and 368D04, including VH and VL domains, and CDRs
(H1,H2,H3,L1,L2, and L3).
[0050] Figure 9. Amino acid and nucleotide sequences of scFv's for non-
germlined GIL01, GIL16, GIL45, GIL60, GIL68, GIL92, 097D09, 062A09, 062G05,
087B03, 367D04, 368D04, 166B06,166G05, 375G06, 376B10, 354A08, 355B06,
355E04, and 356A11, with CDRs underlined (H1, H2, H3, L1, L2, and L3).
[0051]Rgure 10. Amino acid and nucleotide sequences of scFv's for
germlined GIL01, GIL16, GIL45, GIL60, GIL68, GIL92, 062A09, 087B03,166B06,
166G05, 354A08, 355B06, 355E04, 356A11, and 368D04 with CDRs underlined
(H1, H2, H3, L1,L2, and L3).
DETAILED DESCRIPTION
I. Definitions
[0052]In order that the present invention may be more readily understood,
certain terms are first defined. Additional definitions are set forth throughout the
detailed description.
[0053]The term "antibody" refers to an immunoglobulin or fragment thereof,
and encompasses any polypeptide comprising an antigen-binding fragment or an
antigen-binding domain. The term includes but is not limited to pofyclonal,
monoclonal, monospecific, polyspecific, non-specific, humanized, human,
single-chain, chimeric, synthetic, recombinant, hybrid, mutated, grafted, and in vitro
generated antibodies. Unless preceded by the word "intact", the term "antibody"
includes antibody fragments such as Fab, F(ab')2, Fv, scFv, Fd, dAb, and other
antibody fragments that retain antigen-binding function. Typically, such fragments
would comprise an antigen-binding domain.
[0054]The terms "antigen-binding domain" and "antigen-binding
fragment" refer to a part of an antibody molecule that comprises amino acids
responsible for the specific binding between antibody and antigen. The part of the
antigen that is specifically recognized and bound by the antibody is referred to as
the "epitope." An antigen-binding domain may comprise an antibody light chain

variable region (VL) and an antibody heavy chain variable region (VH); however, it
does not have to comprise both. Fd fragments, for example, have two VH regions
and often retain some antigen-binding function of the intact antigen-binding
domain. Examples of antigen-binding fragments of an antibody include (1) a Fab
fragment, a monovalent fragment having the VL, VH, CL and CH1 domains; (2) a
F(ab')2 fragment, a bivalent fragment having two Fab fragments linked by a
disulfide bridge at the hinge region; (3) a Fd fragment having the two VH and CH1
domains; (4) a Fv fragment having the VL and VH domains of a single arm of an
antibody, (5) a dAb fragment (Ward et al., (1989) Nature 341:544-546), which has
a VH domain; (6) an isolated complementarity determining region (CDR), and (7) a
single chain Fv (scFv). Although the two domains of the Fv fragment, VL and VH,
are coded for by separate genes, they can be joined, using recombinant methods,
by a synthetic linker that enables them to be made as a single protein chain in
which the VL and VH regions pair to form monovalent molecules (known as single
chain Fv (scFv); see e.g., Bird et al. (1988) Science 242:423-426; and Huston et al.
(1988) Proc. Natl. Acad. Sci. USA 85:5879-5883). These antibody fragments are
obtained using conventional techniques known to those with skill in the art, and the
fragments are evaluated for function in the same manner as are intact antibodies.
[0055]The term "effective amount" refers to a dosage or amount that is
sufficient to regulate IL-22 activity to ameliorate clinical symptoms or achieve a
desired biological outcome, e.g., decreased T cell and/or B cell activity,
suppression of autoimmunity, suppression of transplant rejection, etc.
[0056]The term "human antibody" includes antibodies having variable and
constant regions corresponding substantially to human germline immunoglobulin
sequences known in the art, including, for example, those described by Kabat et al.
(See Kabat, et a!. (1991) Sequences of Proteins of Immunological interest, Fifth
Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-
3242). The human antibodies of the invention may include amino acid residues not
encoded by human germline immunoglobulin sequences (e.g., mutations
introduced by random or site-specific mutagenesis in vitro or by somatic mutation
in vivo), for example in the CDRs, and in particular, CDR3. The human antibody
can have at least one, two, three, four, five, or more positions replaced with an

amino acid residue that is not encoded by the human germline immunoglobulin
sequence.
[0057]The phrase "inhibit" or "antagonize" IL-22 activity and its cognates
refer to a reduction, inhibition, or otherwise diminution of at least one activity of
IL-22 due to binding an anti-IL-22 antibody, wherein the reduction is relative to the
activity of IL-22 in the absence of the same antibody. The activity can be
measured using any technique known in the art, including, for example, as
described in Examples 7 and 9. Inhibition or antagonism does not necessarily
indicate a total elimination of the IL-22 polypeptide biological activity. A reduction
in activity may be about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or
more.
[0058]The term "inter Jeukin-22" or "IL-22" refers to a class II cytokine
(which may be mammalian) capable of binding to IL-22R and/or a receptor
complex of IL-22R and IL-10R2, and has at least one of the following features: (1)
an amino acid sequence of a naturally occurring mammalian IL-22 polypeptide (full
length or mature form) or a fragment thereof, e.g., an amino acid sequence shown
as SEQ ID NO:1 (human) or SEQ ID NO:3 (murine) or a fragment thereof; (2) an
amino acid sequence substantially identical to, e.g., at least 85%, 90%, 95%, 96%,
97%, 98%, 99% identical to, an amino acid sequence shown as SEQ ID NO:1 or
amino acids 34-179 thereof (human) or SEQ ID NO:3 (murine) or a fragment
thereof; (3) an amino acid sequence which is encoded by a naturally occurring
mammalian IL-22 nucleotide sequence or a fragment thereof (e.g., SEQ ID NO:2 or
nucleotides 71 to 610 (human) or SEQ ID NO:4 (murine) or a fragment thereof); (4)
an amino acid sequence encoded by a nucleotide sequence which is substantially
identical to, e.g., at least 85%, 90%, 95%, 96%, 97%, 98%, 99% identical to, a
nucleotide sequence shown as SEQ ID NO:2 or nucleotides 71 to 610 thereof
(human) or SEQ ID NO:4 (murine) or a fragment thereof; (5) an amino acid
sequence encoded by a nucleotide sequence degenerate to a naturally occurring
IL-22 nucleotide sequence or a fragment thereof, e.g., SEQ ID NO:2 (human) or
SEQ ID NO:4 (murine) or a fragment thereof; or (6) a nucleotide sequence that
hybridizes to one of the foregoing nucleotide sequences under stringent conditions,
e.g., highly stringent conditions. The IL-22 may bind to IL-22R and/or a receptor
complex of IL-22R and IL-10R2 of mammalian origin, e.g., human or mouse.

[0059]The nudeotide sequence and the predicted amino acid sequence of
human IL-22 are shown in SEQ ID NO:2 and SEQ ID NO:1, respectively. The
amino acid sequence of mature human IL-22 corresponds to amino acids 34-179 of
SEQ ID NO:1. Analysis of recombinant human IL-22 reveals many structural
domains. (Nagem et al. (2002) Structure, 10:1051-62; U.S. Patent Application No.
US 2002/0187512 A1).
[0060]The term "IL-22 activity" refers to at least one cellular process
initiated or interrupted as a result of IL-22 binding to a receptor complex consisting
of IL-22R and IL-10R2 on the cell. IL-22 activities include at least one of, but are
not limited to: (1) binding IL-22R or a receptor complex of IL-22R and IL-10R2
(e.g., human IL-22R with or without human IL-10R2); (2) associating with signal
transduction molecules (e.g., JAK-1); (3) stimulating phosphorylation of STAT
proteins (e.g., STAT5, STAT3, or combination thereof); (4) activating STAT
proteins; and (5) modulating (e.g., increasing or decreasing) proliferation,
differentiation, effector cell function, cytolytic activity, cytokine secretion, survival, or
combinations thereof, of epithelial cells, fibroblasts, or immune cells. Epithelial
cells include, but are not limited to, cells of the skin, gut, liver, and kidney, as well
as endothelial ceils. Fibroblasts include, but are not limited to, synovia! fibroblasts.
Immune cells may include CD8+ and CD4+ T cells, NK cells, B cells,
macrophages, and megakaryocytes. IL-22 activity can be determined in vitro, for
example, using the IL-22 receptor inhibition assay as described in Examples 2 and
6, the GROa secretion assay in Example 9, or the BAF3 proliferation assay of
Example 7. IL-22 activity can also be determined in vivo, for example, by scoring
progression of an immune response or disorder as described in Example 13.
[0061] As used herein, "in vitro generated antibody" refers to an antibody
where all or part of the variable region (e.g., at least one CDR) is generated in a
non-immune cell selection (e.g., an in vitro phage display, protein chip or any other
method in which candidate sequences can be tested for their ability to bind to an
antigen). This term excludes sequences generated by genomic rearrangement in
an immune cell.
[0062]The term "isolated" refers to a molecule that is substantially free of its
natural environment. For instance, an isolated protein is substantially free of
cellular material or other proteins from the ceil or tissue source from which it was

derived. The term also refers to preparations where the isolated protein is
sufficiently pure for pharmaceutical compositions; or at least 70-80% (w/w) pure; or
at least 80-90% (w/w) pure; or at least 90-95% pure; or at least 95%, 96%, 97%,
98%, 99%, or 100% (w/w) pure.
[0063]The phrase "percent identical" or "percent identity" refers to the
similarity between at least two different sequences. This percent identity can be
determined by standard alignment algorithms, for example, the Basic Local
Alignment Tool (BLAST) described by Altshul et al. ((1990) J. Mol. Biol., 215:
403-410); the algorithm of Needleman et al. ((1970) J. Mol. Biol., 48:444-453); or
the algorithm of Meyers et al. ((1988) Comput. Appl. Biosci., 4: 11-17). A set of
parameters may be the Blosum 62 scoring matrix with a gap penalty of 12, a gap
extend penalty of 4, and a frameshift gap penalty of 5. The percent identity
between two amino acid or nucleotide sequences can also be determined using the
algorithm of E. Meyers and W. Miller ((1989) CABIOS, 4:11-17) which has been
incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue
table, a gap length penalty of 12 and a gap penalty of 4. The percent identity is
usually calculated by comparing sequences of similar length.
[0064]The term "repertoire" refers to at least one nucleotide sequence
derived wholly or partially from at least one sequence encoding at least one
immunoglobulin. The sequence(s) may be generated by rearrangement in vivo of
the V, D, and J segments of heavy chains, and the V and J segments of light
chains. Alternatively, the sequence(s) can be generated from a cell in response to
which rearrangement occurs, e.g., in vitro stimulation. Alternatively, part or all of
the sequence(s) may be obtained by DNA splicing, nucleotide synthesis,
mutagenesis, and other methods, see, e.g., U.S. Patent 5,565,332. A repertoire
may include only one sequence or may include a plurality of sequences, including
ones in a genetically diverse collection.
[0065] The terms "specific binding" or "specifically binds" refers to two
molecules forming a complex that is relatively stable under physiologic conditions.
Specific binding is characterized by a high affinity and a low to moderate capacity
as distinguished from nonspecific binding which usually has a low affinity with a
moderate to high capacity. Typically, binding is considered specific when the
association constant KA is higher than 106M-1. If necessary, nonspecific binding

can be reduced without substantially affecting specific binding by varying the
binding conditions. The appropriate binding conditions, such as concentration of
antibodies, ionic strength of the solution, temperature, time allowed for binding,
concentration of a blocking agent (e.g., serum albumin, milk casein), etc., may be
optimized by a skilled artisan using routine techniques. Illustrative conditions are
set forth in Example 3, but other conditions known to the person of ordinary skill in
the art fall within the scope of this invention.
[0066]As used herein, the term "stringent" describes conditions for
hybridization and washing. Stringent conditions are known to those skilled in the
art and can be found in Current Protocols in Molecular Biology, John Wiley & Sons,
N.Y. (1989), 6.3.1-6.3.6. Aqueous and nonaqueous methods are described in that
reference and either can be used. One example of stringent hybridization
conditions is hybridization in 6X sodium chloride/sodium citrate (SSC) at about
45°C, followed by at least one wash in 0.2X SSC, 0.1% SDS at 50°C. A second
example of stringent hybridization conditions is hybridization in 6X SSC at about
45°C, followed by at least one wash in 0.2X SSC, 0.1% SDS at 55°C. Another
example of stringent hybridization conditions is hybridization in 6X SSC at about
45°C, followed by at least one wash in 0.2X SSC, 0.1% SDS at 60°C. A further
example of stringent hybridization conditions is hybridization in 6X SSC at about
45°C, followed by at least one wash in 0.2X SSC, 0.1% SDS at 65°C. High
stringent conditions include hybridization in 0.5M sodium phosphate, 7% SDS at
65°C, followed by at least one wash at 0.2X SSC, 1 % SDS at 65°C.
[0067]The phrase "substantially as set out," "substantially identical" or
"substantially homologous" means that the relevant amino acid or nucleotide
sequence (e.g., CDR(s), VH, or VL domain) will be identical to or have insubstantial
differences (through conserved amino acid substitutions) in comparison to the
sequences which are set out. Insubstantial differences include minor amino acid
changes, such as 1 or 2 substitutions in a 5 amino acid sequence of a specified
region. In the case of antibodies, the second antibody has the same specificity and
has at least 50% of the affinity of the first antibody.
[0068]Sequences substantially identical or homologous (e.g., at least about
85% sequence identity) to the sequences disclosed herein are also part of this
application. In some embodiment, the sequence identity can be about 85%, 90%,

95%, 96%, 97%, 98%, 99% or higher. Alternatively, substantial identity or
homology exists when the nucleic acid segments will hybridize under selective
hybridization conditions (e.g., highly stringent hybridization conditions), to the
complement of the strand. The nucleic acids may be present in whole cells, in a
cell lysate, or in a partially purified or substantially pure form.
[0069]The term "therapeutic agent" is a substance that treats or assists in
treating a medical disorder. Therapeutic agents may include, but are not limited to,
substances that modulate immune cells or immune responses in a manner that
complements the IL-22 activity of anti-IL-22 antibodies. Non-limiting examples and
uses of therapeutic agents are described herein.
[0070]As used herein, a "therapeutically effective amount" of an anti-
IL-22 antibody refers to an amount of an antibody which is effective, upon single or
multiple dose administration to a subject (such as a human patient) at treating,
preventing, curing, delaying, reducing the severity of, and/or ameliorating at least
one symptom of a disorder or recurring disorder, or prolonging the survival of the
subject beyond that expected in the absence of such treatment.
[0071 ]The term treatment" refers to a therapeutic or preventative measure.
The treatment may be administered to a subject having a medical disorder or who
ultimately may acquire the disorder, in order to prevent, cure, delay, reduce the
severity of, and/or ameliorate one or more symptoms of a disorder or recurring
disorder, or in order to prolong the survival of a subject beyond that expected in the
absence of such treatment.
II. Anti-IL-22 Antibodies
[0072]The disclosure provides novel anti-IL-22 antibodies that comprise
novel antigen-binding fragments.
[0073]Numerous methods known to those skilled in the art are available for
obtaining antibodies or antigen-binding fragments thereof. For example, antibodies
can be produced using recombinant DNA methods (U.S. Patent 4,816,567).
Monoclonal antibodies may also be produced by generation of hybridomas (see
e.g., Kohler and Milstein (1975) Nature, 256:495-499) in accordance with known
methods. Hybridomas formed in this manner are then screened using standard
methods, such as enzyme-linked immunosorbent assay (ELISA) and surface
plasmon resonance (BIACORE™) analysis, to identify one or more hybridomas

that produce an antibody that specifically binds with a specified antigen. Any form
of the specified antigen may be used as the immunogen, e.g., recombinant
antigen, naturally occurring forms, any variants or fragments thereof, as well as
antigenic peptide thereof.
[0074]One exemplary method of making antibodies includes screening
protein expression libraries, e.g., phage or ribosome display libraries. Phage
display is described, for example, in Ladner et al., U.S. Patent No. 5,223,409;
Smith (1985) Science 228:1315-1317; Clackson et al. (1991) Nature, 352: 624-628;
Marks et al. (1991) J. Mo I. Bio!., 222: 581-597WO 92/18619; WO 91/17271; WO
92/20791; WO 92/15679; WO 93/01288; WO 92/01047; WO 92/09690; and WO
90/02809.
[0075]In addition to the use of display libraries, the specified antigen can be
used to immunize a non-human animal, e.g., a rodent, e.g., a mouse, hamster, or
rat. In one embodiment, the non-human animal includes at least a part of a human
immunoglobulin gene. For example, it is possible to engineer mouse strains
deficient in mouse antibody production with large fragments of the human Ig loci.
Using the hybridoma technology, antigen-specific monoclonal antibodies derived
from the genes with the desired specificity may be produced and selected. See,
e.g., XENOMOUSE™, Green etal. (1994) Nature Genetics 7:13-21, US 2003-
0070185, WO 96/34096, published Oct. 31, 1996, and PCT Application No.
PCT/US96/05928, filed Apr. 29,1996.
[0076]In another embodiment, a monoclonal antibody is obtained from the
non-human animal, and then modified, e.g., humanized, deimmunized, chimeric,
may be produced using recombinant DNA techniques known in the art. A variety of
approaches for making chimeric antibodies have been described. See e.g.,
Morrison et al., Proc. Natl. Acad. Sci. U.S.A. 81:6851, 1985; Takeda et al., Nature
314:452, 1985, Cabilly et al., U.S. Patent No. 4,816,567; Boss etal., U.S. Patent
No. 4,816,397; Tanaguchi et al., European Patent Publication EP171496;
European Patent Publication 0173494, United Kingdom Patent GB 2177096B.
Humanized antibodies may also be produced, for example, using transgenic mice
that express human heavy and light chain genes, but are incapable of expressing
the endogenous mouse immunoglobulin heavy and light chain genes. Winter
describes an exemplary CDR-grafting method that may be used to prepare the

humanized antibodies described herein (U.S. Patent No. 5,225,539). Alf of the
CDRs of a particular human antibody may be replaced with at least a portion of a
non-human CDR, or only some of the CDRs may be replaced with non-human
CDRs. It is only necessary to replace the number of CDRs required for binding of
the humanized antibody to a predetermined antigen.
[0077] Humanized antibodies or fragments thereof can be generated by
replacing sequences of the Fv variable domain that are not directly involved in
antigen binding with equivalent sequences from human Fv variable domains.
Exemplary methods for generating humanized antibodies or fragments thereof are
provided by Morrison (1985) Science 229:1202-1207; by Oi et al. (1986)
BioTechniques 4:214; and by US 5,585,089; US 5,693,761; US 5,693,762; US
5,859,205; and US 6,407,213. Those methods include isolating, manipulating, and
expressing the nucleic acid sequences that encode all or part of immunoglobulin Fv
variable domains from at least one of a heavy or light chain. Such nucleic acids
may be obtained from a hybridoma producing an antibody against a predetermined
target, as described above, as well as from other sources. The recombinant DNA
encoding the humanized antibody molecule can then be cloned into an appropriate
expression vector.
[0078] In certain embodiments, a humanized antibody is optimized by the
introduction of conservative substitutions, consensus sequence substitutions,
germiine substitutions and/or backmutations. Such altered immunogiobulin
molecules can be made by any of several techniques known in the art, (e.g., Teng
et al., Proc. Natl. Acad. Set. U.S.A., 80:7308-7312,1983; Kozbor et al.,
Immunology Today, 4: 7279, 1983; Olsson etai, Math. EnzymoL, 92: 3-16, 1982),
and may be made according to the teachings of PCT Publication WO92/06193 or
EP 0239400).
[0079]An antibody or fragment thereof may also be modified by specific
deletion of human T cell epitopes or "deimmunization" by the methods disclosed in
WO 98/52976 and WO 00/34317. Briefly, the heavy and light chain variable
domains of an antibody can be analyzed for peptides that bind to MHC Class II;
these peptides represent potential T-cell epitopes (as defined in WO 98/52976 and
WO 00/34317). For detection of potential T-cell epitopes, a computer modeling
approach termed "peptide threading" can be applied, and in addition a database of

human MHC class II binding peptides can be searched for motifs present in the VH
and VL sequences, as described in WO 98/52976 and WO 00/34317. These motifs
bind to any of the 18 major MHC class II DR allotypes, and thus constitute potential
T cell epitopes. Potential T-cell epitopes detected can be eliminated by substituting
small numbers of amino acid residues in the variable domains, or preferably, by
single amino acid substitutions. Typically, conservative substitutions are made.
Often, but not exclusively, an amino acid common to a position in human germline
antibody sequences may be used. Human germline sequences, e.g., are disclosed
in Tomlinson, et at. (1992) J. Mol. Biol. 227:776-798; Cook, G. P. et al. (1995)
Immunol. Today Vol. 16 (5): 237-242; Chothia, D. etal. (1992) J. Mol. Biol.
227:799-817; and Tomlinson et al. (1995) EMBO J. 14:4628-4638. The V BASE
directory provides a comprehensive directory of human immunoglobulin variable
region sequences (compiled by Tomlinson, LA. et al. MRC Centre for Protein
Engineering, Cambridge, UK). These sequences can be used as a source of
human sequence, e.g., for framework regions and CDRs. Consensus human
framework regions can also be used, e.g., as described in U.S. Patent No.
6,300,064.
[0080] In certain embodiments, an antibody can contain an altered
immunoglobulin constant or Fc region. For example, an antibody produced in
accordance with the teachings herein may bind more strongly or with more
specificity to effector molecules such as complement and/or Fc receptors, which
can control several immune functions of the antibody such as effector cell activity,
lysis, complement-mediated activity, antibody clearance, and antibody half-life.
Typical Fc receptors that bind to an Fc region of an antibody (e.g., an IgG antibody)
include, but are not limited to, receptors of the FcγRI, FcγRll, and FcγRIII and FcRn
subclasses, including alielic variants and alternatively spliced forms of these
receptors. Fc receptors are reviewed in Ravetch and Kinet, Annu. Rev. Immunol
9:457-92, 1991; Capel et al., Immunomethods 4:25-34,1994; and de Haas et al., J.
Lab. Clin. Med. 126:330-41,1995).
[0081]For additional antibody production techniques, see Antibodies: A
Laboratory Manual, eds. Harlow et al., Cold Spring Harbor Laboratory, 1988. The
present invention is not necessarily limited to any particular source, method of
production, or other special characteristics of an antibody.

[0082]Antibodies, also known as immunoglobulins, are typically tetrameric
glycosylated proteins composed of two light (L) chains of approximately 25 kDa
each and two heavy (H) chains of approximately 50 kDa each. Two types of light
chain, termed lambda and kappa, may be found in antibodies. Depending on the
amino acid sequence of the constant domain of heavy chains, immunoglobulins
can be assigned to five major classes: A, D, E, G, and M, and several of these may
be further divided into subclasses (isotypes), e.g., IgG1, lgG2, lgG3, lgG4, IgA1 and
lgA2. Each light chain includes an N-terminal variable (V) domain (VL) and a
constant (C) domain (CL). Each heavy chain includes an N-terminal V domain
(VH), three or four C domains (CHs), and a hinge region. The CH domain most
proximal to VH is designated as CH1. The VH and VL domains consist of four
regions of relatively conserved sequences called framework regions (FR1, FR2,
FR3, and FR4), which form a scaffold for three regions of hypervariable sequences
(complementarity determining regions, CDRs). The CDRs contain most of the
residues responsible for specific interactions of the antibody with the antigen.
CDRs are referred to as CDR1, CDR2, and CDR3. Accordingly, CDR constituents
on the heavy chain are referred to as H1, H2, and H3, while CDR constituents on
the light chain are referred to as L1, L2, and L3.
[0083]CDR3 is typically the greatest source of molecular diversity within the
antibody-binding site. H3, for example, can be as short as two amino acid residues
or greater than 26 amino acids. The subunit structures and three-dimensional
configurations of different classes of immunoglobulins are well known in the art.
For a review of the antibody structure, see Antibodies: A Laboratory Manual, Cold
Spring Harbor Laboratory, eds. Harlow et al., 1988. One of skill in the art will
recognize that each subunit structure, e.g., a CH, VH, CL, VL, CDR, FR structure,
comprises active fragments, e.g., the portion of the VH, VL, or CDR subunit the
binds to the antigen, i.e., the antigen-binding fragment, or, e.g., the portion of the
CH subunit that binds to and/or activates, e.g., an Fc receptor and/or complement
The CDRs typically refer to the Kabat CDRs, as described in Sequences of
Proteins of Immunological Interest, US Department of Health and Human Services
(1991), eds. Kabat et al. Another standard for characterizing the antigen binding
site is to refer to the hypervariable loops as described by Chothia. See, e.g.,
Chothia, D. et al. (1992; J. Mot. Blot. 227:799-817; and Tomlinson et al. (1995)

EMBO J. 14:4628-4638. Still another standard is the AbM definition used by
Oxford Molecular's AbM antibody modelling software. See, generally, e.g., Protein
Sequence and Structure Analysis of Antibody Variable Domains. In: Antibody
Engineering Lab Manual (Ed.: Duebel, S. and Kontermann, R., Springer-Verlag,
Heidelberg). Embodiments described with respect to Kabat CDRs can alternatively
be implemented using similar described relationships with respect to Chothia
hypervariable loops or to the AbM-defined loops.
[0084]The Fab fragment (Fragment antigen-binding) consists of VH-CH1 and
VL-CL domains covalently linked by a disulfide bond between the constant regions.
The Fv fragment is smaller and consists of VH and VL domains non-covalently
linked. To overcome the tendency of non-covalently linked domains to dissociate,
a single chain Fv fragment (scFv) can be constructed. The scFv contains a flexible
potypeptide that Jinks (1) the C-terminus of VH to the N-terminus of VL, or (2) the
C-terminus of VL to the N-terminus of VH. A 15-mer (Gly4Ser)3 peptide may be
used as a linker, but other linkers are known in the art.
[0085]The sequence of antibody genes after assembly and somatic
mutation is highly varied, and these varied genes are estimated to encode 1010
different antibody molecules (Immunoglobulin Genes, 2nd ed., eds. Jonio et at,
Academic Press, San Diego, CA, 1995).
[0086]A "bispecific" or "bifunctional antibody" Is an artificial hybrid antibody
having two different heavy/light chain pairs and two different binding sites.
Bispecific antibodies can be produced by a variety of methods including fusion of
hybridomas or linking of Fab' fragments. See, e.g., Songsivilai & Lachmann, Clin.
Exp. Immunol, 79:315-321 (1990); Kostelny et al., J. Immunol. 148,1547-1553
(1992). In one embodiment, the bispecific antibody comprises a first binding
domain polypeptide, such as a Fab' fragment, linked via an immunoglobulin
constant region to a second binding domain polypeptide.
[0087]Small Modular ImmunoPharmaceuticals (SMIP™) provide an
example of a variant molecule comprising a binding domain polypeptide. SMIPs
and their uses and applications are disclosed in, e.g., U.S. Published Patent
Application. Nos. 2003/0118592, 2003/0133939, 2004/0058445, 2005/0136049,
2005/0175614, 2005/0180970, 2005/0186216, 2005/0202012, 2005/0202023,
2005/0202028, 2005/0202534, and 2005/0238646, and related patent family

members thereof, all of which are hereby incorporated by reference herein in their
entireties.
[0088]A SMIP™ typically refers to a binding domain-immunoglobulin fusion
protein that includes a binding domain polypeptide that is fused or otherwise
connected to an immunoglobulin hinge or hinge-acting region polypeptide, which in
turn is fused or otherwise connected to a region comprising one or more native or
engineered constant regions from an immunoglobulin heavy chain, other than CH1,
for example, the CH2 and CH3 regions of IgG and IgA, or the CH3 and CH4
regions of IgE (see e.g., U.S. 2005/0136049 by Ledbetter, J. et al., which is
incorporated by reference, for a more complete description). The binding domain-
immunoglobulin fusion protein can further include a region that includes a native or
engineered immunoglobulin heavy chain CH2 constant region polypeptide (or CH3
in the case of a construct derived in whole or in part from IgE) that is fused or
otherwise connected to the hinge region polypeptide and a native or engineered
immunoglobulin heavy chain CH3 constant region polypeptide (or CH4 in the case
of a construct derived in whole or in part from IgE) that is fused or otherwise
connected to the CH2 constant region polypeptide (or CH3 in the case of a
construct derived in whole or in part from IgE). Typically, such binding domain-
immunoglobulin fusion proteins are capable of at least one immunological activity
selected from the group consisting of antibody dependent cell-mediated
cytotoxicity, complement fixation, and/or binding to a target, for example, a target
antigen, such as human IL-22.
[0089]Therapeutic proteins, i.e., a protein or peptide that has a biological
effect on a region in the body on which it acts or on a region of the body on which it
remotely acts via intermediates, are also useful for practicing the invention. A
therapeutic protein can include peptide mimetics. Mimetics are peptide-containing
molecules that mimic elements of protein secondary structure. See, for example,
Johnson et al., "Peptide Turn Mimetics11 in BIOTECHNOLOGY AND PHARMACY,
Pezzuto et al., Eds., Chapman and Hall, New York (1993), incorporated herein by
reference. The underlying rationale behind the use of peptide mimetics is that the
peptide backbone of proteins exists chiefly to orient amino acid side chains in such
a way as to facilitate molecular interactions, such as those of antibody and antigen.
A peptide mimetic is expected to permit molecular interactions similar to the natural

molecule. These principles may be used to engineer second generation molecules
having many of the natural properties of the targeting peptides disclosed herein,
but with altered and potentially improved characteristics.
[0090]Other embodiments of therapeutic proteins include fusion proteins.
These molecules generally have all or a substantial portion of a targeting peptide,
for example, 1L-22 or an anti IL-22 antibody, linked at the N- or C-terminus, to all or
a portion of a second polypeptide or protein. For example, fusions may employ
leader sequences from other species to permit the recombinant expression of a
protein in a heterologous host. Another useful fusion includes the addition of an
immunologically active domain, such as an antibody epitope, to facilitate
purification of the fusion protein. Inclusion of a cleavage site at or near the fusion
junction will facilitate removal of the extraneous polypeptide after purification.
Other useful fusions include linking of functional domains, such as active sites from
enzymes, glycosylation domains, cellular targeting signals or transmembrane
regions. Examples of proteins or peptides that may be incorporated into a fusion
protein include cytostatic proteins, cytocidal proteins, pro-apoptosis agents, anti-
angiogenic agents, hormones, cytokines, growth factors, peptide drugs, antibodies,
Fab fragments of antibodies, antigens, receptor proteins, enzymes, lectins, MHC
proteins, cell adhesion proteins and binding proteins. Methods of generating fusion
proteins are well known to those of skill in the art. Such proteins can be produced,
for example, by chemical attachment using bifunctional cross-linking reagents, by
de novo synthesis of the complete fusion protein, or by attachment of a DNA
sequence encoding the targeting peptide to a DNA sequence encoding the second
peptide or protein, followed by expression of the intact fusion protein.
[0091] ln one embodiment, the targeting peptide, for example, IL-22 or an
anti IL-22 antibody, is fused with an immunoglobulin heavy chain constant region,
such as an Fc fragment, which contains two constant region domains and a hinge
region but lacks the variable region (See, U.S. Pat. Nos. 6,018,026 and 5,750,375,
incorporated herein by reference). The Fc region may be a naturally occurring Fc
region, or may be altered to improve certain qualities, such as therapeutic qualities,
circulation time, reduced aggregation, etc. Peptides and proteins fused to an Fc
region typically exhibit a greater half-life in vivo than the unfused counterpart. Also,

a fusion to an Fc region permits dimerization/multimerization of the fusion
potypeptide.
[0092] VHH molecules (or nanobodies), as known to the skilled artisan, are
heavy chain variable domains derived from immunoglobulins naturally devoid of
light chains, such as those derived from Camelidae as described in WO9404678,
incorporated herein by reference. Such a VHH molecule can be derived from
antibodies raised in Camelidae species, for example in camel, llama, dromedary,
alpaca and guanaco and is sometomes called a camelid or carnalized variable
domain. See e.g., Muyldermans., J. Biotechnology (2001) 74(4):277-302,
incorporated herein by reference. Other species besides Camelidae may produce
heavy chain antibodies naturally devoid of light chain. VHH molecules are about
10 times smaller than IgG molecules. They are single polypeptides and very
stable, resisting extreme pH and temperature conditions. Moreover, they are
resistant to the action of proteases which is not the case for conventional
antibodies. Furthermore, in vitro expression of VHHs produces high yield, properly
folded functional VHHs. In addition, antibodies generated in Camelids will
recognize epitopes other than those recognized by antibodies generated in vitro
through the use of antibody libraries or via immunization of mammals other than
Camelids (see WO 9749805, which is incorporated herein by reference).
[0093]One aspect of the present invention comprises antibodies and antigen
binding fragments that bind IL-22. The disclosure provides novel CDRs derived
from human immunoglobulin gene libraries. The structure for carrying a CDR is
generally an antibody heavy or light chain or portion thereof, where the CDR is
located to a naturally occurring CDR region. The structures and locations of
variable domains may be determined as described in Kabat et al., Sequences of
Proteins of Immunological Interest, No. 91-3242, National Institutes of Health
Publications, Bethesda, MD (1991).
[0094]DNA and amino acid (AA) sequences of illustrative embodiments of
the anti-IL-22 antibodies of this invention, including their scFv fragments, VH and VL
domains, and CDRs, are set forth in Figures 7-10 and enumerated in Tables 1 and
7. Twenty specific embodiments of the non-germlined antibodies are identified as
GIL01, GIL16, GIL45, GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03,
367D04, 368D04, 166B06, 166G05, 375G06, 376B10, 354A08, 355B06, 355E04,

and 356A11. The CDR positions in the VH and VL domains of the non-germlined
antibodies are listed in Table 2. Fifteen specific embodiments of the germlined
antibodies are identified as GIL01, GIL16, GIL45, GIL60, GIL68, G1L92, 062A09,
087B03, 166B06, 166G05, 354A08, 355B06, 355E04, 356A11, and 368D04.

[0095]Anti-IL-22 antibodies of this invention may optionally comprise
antibody constant regions or parts thereof. For example, a VL domain may be
attached at its C-terminal end to a light chain constant domain like CΚ or Cλ.
Similarly, a VH domain or portion thereof may be attached to all or part of a heavy
chain like IgA, IgD, IgE, IgG, and IgM, and any isotype subclass. Constant regions
are known in the art (see, for example, Kabat et al., Sequences of Proteins of
Immunological Interest, No. 91-3242, National Institutes of Health Publications,
Bethesda, MD (1991)). Therefore, antibodies within the scope of this invention
include VH and VL domains, or a portion thereof, combined with constant regions
known in the art.
[0096]Certain embodiments comprise a VH domain, a VL domain, or a
combination thereof, of the Fv fragment from GIL01, GIL16, GIL45, GIL60, GIL68,
GIL92, 097D09, 062A09, 062G05, 087B03. 367D04, 368D04, 166B06, 166G05,
375G06,376B10, 354A08, 355B06, 355E04, or 356A11. Another embodiment
comprises a VH domain, a VL domain, or a combination thereof, of the Fv fragment
from an antibody chosen from 356A11, 354A08, 087B03, and 368D04. Further
embodiments comprise one, two, three, four, five or six complementarity
determining regions (CDRs) from the VH and VL domains. Antibodies whose CDR
sequences are included within SEQ ID NO:5-13, 23-31, 41-49, 59-67, 77-85, 95-
103, 113-121, 131-139, 149-157, 167-175, 185-193, 203-211, 221-229, 239-247,
257-265, 275-283, 293-301, 311-319, 329-337, 347-355, 365-373, 383-391, 401-
409, 419-427, 437-445, 455-463, 473-481, 491-499, 509-517, 527-535, 545-553,
563-571, 581-589, 599-607, or 617-625 are encompassed within the scope of this
invention. For example, in one embodiment, an antibody comprises a H3 fragment
of the VH domain of germlined or non-germlined GIL01, GIL16, GIL45, GIL60,
G1L68, GIL92, 097D09, 062A09, 062G05, 087B03, 367D04, 368D04, 166B06,
166G05, 375G06, 376B10, 354A08, 355B06, 355E04, or 356A11 or from an
antibody chosen from 356A11, 354A08, 087B03, and 368D04.
[0097] In certain embodiments, the VH and/or VL domains may be germlined,
i.e., the framework regions (FR) of these domains are mutated using conventional
molecular biology techniques to match those produced by the germline cells. In
other embodiments, the FR sequences remain diverged from the consensus

germline sequences. In one embodiment of this invention, germlined antibodies
are shown in Table 7.
[0098]In one embodiment, the invention provides amino acid and nucleic
acid sequences for the germlined GIL01, GIL16, GIL45, GIL60, GIL68, GIL92,
097D09, 062A09, 062G05, 087B03, 367D04, 368D04, 166B06, 166G05, 375G06,
376B10, 354A08, 355B06, 355E04, or 356A11. Amino acid and nucleotide
sequences for the VH domain of the germlined GIL01, GIL16, GIL45, GIL60, GIL68,
GIL92, 062A09, 087B03, 166B06, 166G05, 354A08, 355B06, 355E04, 356A11,
and 368D04 are depicted in Table 7 and Figure 8. Amino acid and nucleotide
sequences for the VL domain of the germlined GIL01, GIL 16, GIL45, GIL60, GIL68,
GIL92, 062A09, 087B03, 166B06, 166G05, 354A08, 355B06, 355E04, 356A11,
and 368D04 are also depicted in Table 7 and Figure 8.
[0099] In one embodiment, mutagenesis is used to make an antibody more
similar to one or more germline sequences. This may be desirable when mutations
are introduced into the framework region of an antibody through somatic
mutagenesis or through error prone PCR. Germline sequences for the VH and VL
domains can be identified by performing amino acid and nucleic acid sequence
alignments against the VBASE database (MRC Center for Protein Engineering,
UK). VBASE is a comprehensive directory of all human germline variable region
sequences compiled from over a thousand published sequences, including those in
the current releases of the Genbank and EMBL data libraries. In some
embodiments, the FR regions of the scFvs are mutated in conformity with the
closest matches in the VBASE database and the CDR portions are kept intact.
[00100] In certain embodiments, antibodies of this invention specifically
react with an epitope that is the same as the epitope recognized by GIL01, GIL16,
GIL45, GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03, 367D04,
368D04, 166B06, 166G05, 375G06, 376B10, 354A08, 355B06, 355E04, or
356A11, such that they competitively inhibit the binding of GIL01, GIL16, GIL45,
GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03, 367D04, 368D04,
166B06, 166G05, 375G06, 376B10, 354A08, 355B06, 355E04, or 356A11 to
human IL-22. Such antibodies can be determined in competitive binding assays.
In one embodiment, the antibody, or antigen binding fragment thereof, binds to an
IL-22 epitope that is recognized by 368D04, such that the antibody competitively

inhibits the binding of 368D04 to human IL-22. In another embodiment, the
antibody, or antigen binding fragment thereof, binds to an IL-22 epitope that is
recognized by 356A11, such that the antibody competitively inhibits the binding of
356A11 to human IL-22. In another embodiment, the antibody, or antigen binding
fragment thereof, binds to an IL-22 epitope that is recognized by 354A08, such that
the antibody competitively Inhibits the binding of 354A08 to human IL-22. In
another embodiment, the antibody, or antigen binding fragment thereof, binds to an
IL-22 epitope that is recognized by 087B03, such that the antibody competitively
inhibits the binding of 087B03 to human IL-22. In one embodiment, the association
constant (KA) of these antibodies for human IL-22 is at least 106 M-1. In another
embodiment, the association constant of these antibodies for human IL-22 is at
least 109 M-1. In other embodiments, the association constant of these antibodies
for human IL-22 is at least 1010 M-1. at least 1011 M-1 or at least 1012 M-1. The
binding affinity may be determined using techniques known in the art, such as
ELISA, biosensor technology, such as biospecific interaction analysis, or other
techniques including those described in this application.
[00101] it is contemplated that antibodies of this invention may bind
other proteins, such as, for example, recombinant proteins comprising all or a
portion of IL-22.
[00102] One of ordinary skill in the art will recognize that the disclosed
antibodies may be used to detect, measure, and/or inhibit proteins that differ
somewhat from IL-22. For example, these proteins may be homologs of IL-22.
Anti-IL-22 antibodies are expected to bind proteins that comprise a sequence
which is at least about 60%, 70%, 80%, 90%, 95%, or more identical to any
sequence of at least 100, 80, 60,40, or 20 contiguous amino acids in the sequence
set forth SEQ ID NO:1.
[00103] In addition to sequence homology analyses, epitope mapping
(see, e.g., Epitope Mapping Protocols, ed. Morris, Humana Press, 1996), and
secondary and tertiary structure analyses can be carried out to identify specific 3D
structures assumed by the presently disclosed antibodies and their complexes with
antigens. Such methods include, but are not limited to, X-ray crystallography
(Engstom (1974) Biochem. Exp. Biol., 11:7-13) and computer modeling of virtual
representations of the present antibodies (Fletterick et al. (1986) Computer

Graphics and Molecular Modeling, in Current Communications in Molecular
Biology, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).
[00104] The disclosure provides a method for obtaining anti-IL-22
antibodies that comprises creating antibodies with altered Table 1 VH and/or VL
sequence(s). Such antibodies may be derived by a skilled artisan using techniques
known in the art. For example, amino acid substitutions, deletions, or additions can
be introduced in FR and/or CDR regions. FR changes are usually designed to
improve the stability and immunogenicity of the antibody, while CDR changes are
typically designed to increase antibody affinity for its antigen. The changes that
increase affinity may be tested by altering CDR sequence and measuring antibody
affinity for its target (see Antibody Engineering, 2nd ed., Oxford University Press,
ed. Borrebaeck, 1995).
[00105] Antibodies whose CDR sequences differ insubstantially from
those included in or included within the sequences in SEQ ID NO: 5-13, 23-31, 41-
49, 59-67, 77-85, 95-103, 113-121,131-139, 149-157, 167-175, 185-193, 203-211,
221-229, 239-247, 257-265, 275-283, 293-301, 311-319, 329-337, 347-355, 365-
373, 383-391, 401-409, 419-427, 437-445, 455-463, 473-481, 491-499, 509-517,
527-535,545-553, 563-571, 581-589,599-607, or 617-625, are encompassed
within the scope of this invention. Typically, this involves substitution of an amino
acid with an amino acid having similar charge, hydrophobic, or stereochemical
characteristics. More drastic substitutions in FR regions, in contrast to CDR
regions, may also be made as long as they do not adversely affect (e.g., reduce
affinity by more than 50% as compared to unsubstituted antibody) the binding
properties of the antibody. Substitutions may also be made to germline the
antibody or stabilize the antigen binding site.
[00106] Conservative modifications will produce molecules having
functional and chemical characteristics similar to those of the molecule from which
such modifications are made. In contrast, substantial modifications in the
functional and/or chemical characteristics of the molecules may be accomplished
by selecting substitutions in the amino acid sequence that differ significantly in their
effect on maintaining (1) the structure of the molecular backbone in the area of the
substitution, for example, as a sheet or helical conformation, (2) the charge or
hydrophobicity of the molecule at the target site, or (3) the size of the molecule.

[00107] For example, a "conservative amino acid substitution" may
involve a substitution of a native amino acid residue with a nonnative residue such
that there is little or no effect ori the polarity or charge of the amino acid residue at
that position. (See, for example, MacLennan et al., 1998, Acta Physiol. Scand.
Suppl. 643:55-67; Sasaki et al., 1998, Adv. Biophys. 35:1-24).
[00108] Desired amino acid substitutions (whether conservative or non-
conservative) can be determined by those skilled in the art at the time such
substitutions are desired. For example, amino acid substitutions can be used to
identify important residues of the molecule sequence, or to increase or decrease
the affinity of the molecules described herein. Exemplary amino acid substitutions
include, but are not limited to, those set forth in Table 3.


[00109] In certain embodiments, conservative amino acid substitutions
also encompass non-naturally occurring amino acid residues which are typically
incorporated by chemical peptide synthesis rather than by synthesis in biological
systems.
[00110] In one embodiment, the method for making a variant VH domain
comprises adding, deleting, or substituting at least one amino acid in the disclosed

VH domains, or combining the disclosed VH domains with at least one V|_ domain,
and testing the variant VH domain for IL-22 binding or modulation of IL-22 activity.
[00111] An analogous method for making a variant Vi_ domain comprises
adding, deleting, or substituting at least one amino acid in the disclosed VL
domains, or combining the disclosed VL domains with at least one VH domain, and
testing the variant VL domain for JL-22 binding or modulation of IL-22 activity.
[00112] A further aspect of the disclosure provides a method for
preparing antibodies or antigen-binding fragments that specifically bind IL-22. The
method comprises:
(a) providing a starting repertoire of nucleic acids encoding a VH domain
which lacks at least one CDR or contains at least one CDR to be replaced;
(b) inserting into or replacing the CDR region of the starting repertoire with
at least one donor nucleic acid encoding an amino acid sequence as substantially
set out herein for a VH CDR, yielding a product repertoire;
(c) expressing the nucleic acids of the product repertoire;
(d) selecting a specific antigen-binding fragment that binds to IL-22; and
(e) recovering the specific antigen-binding fragment or nucleic acid
encoding it.
[00113] In an analogous method at least one VL CDR of the invention is
combined with a repertoire of nucleic acids encoding a VL domain which lacks at
least one CDR or contains at least one CDR to be replaced. The at least one VH or
VL CDR may be a CDR1, a CDR2, a CDR3, or a combination thereof, including
combinations of VH and VL CDRs, such as those set forth in Tables 1 or 7, including
those set out in SEQ ID NO:8, 9, 10,11, 12, 13, 26, 27, 28, 29, 30, 31, 44, 45, 46,
47, 48, 49, 62, 63, 64, 65, 66, 67, 80, 81, 82, 83, 84, 85, 98, 99, 100, 101, 102,
103 116, 117, 118, 119, 120, 121, 134, 135, 136, 137, 138, 139, 152, 153, 154,
155, 156, 157, 170,171, 172, 173, 174, 175, 188, 189, 190, 191, 192, 193,206,
207, 208, 209, 210, 211, 224, 225, 226, 227, 228, 229, 242, 243, 244, 245, 246,
247, 260, 261, 262, 263, 264, 265, 278, 279, 280, 281, 282, 283, 296, 297, 298,
299, 300, 301, 314, 315, 316, 317, 318, 319, 332, 333, 334, 335, 336, 337, 350,
351, 352, 353, 354, 355, 368, 369, 370, 371, 372, 373, 386, 387, 388, 389, 390,
391, 404, 405, 406, 407, 408, 409, 422, 423, 424, 425, 426, 427, 440, 441, 442,
443, 444, 445, 458, 459, 460, 461, 462, 463, 476, 477, 478, 479, 480, 481, 494,

495, 496, 497, 498, 499, 512, 513, 514, 515, 516, 517, 530, 531, 532, 533, 534,
535, 548, 549, 550, 551, 552, 553, 566, 567, 568, 569, 570, 571, 584, 585, 586,
587,588, 589, 602, 603, 604, 605,606, 607, 620, 621,622, 623, 624, or 625.
[00114] In one embodiment, the variable domain includes a CDR3 to be
replaced or lacks a CDR3 encoding region and the at least one donor nucleic acid
encodes an amino acid substantially as set out In SEQ ID NO:10,13, 28,31, 46,
49, 64,67, 82, 85,100, 103,118,121,136,139,154, 157,172, 175, 190, 193,
208, 211, 226, 229, 244, 247, 262, 265, 280, 283, 298, 301, 316, 319, 334, 337,
352, 355, 370, 373, 388, 391, 406,409, 424, 427, 442, 445, 460, 463, 478, 481,
496, 499, 514, 517, 532, 535, 550, 553, 568, 571, 586, 589, 604, 607, 622, or 625.
[00115] In another embodiment, the variable domain includes a CDR1 to
be replaced or lacks a CDR1 encoding region and the at least one donor nucleic
acid encodes an amino acid sequence substantially as set out in SEQ ID NO:8, 11,
26, 29, 44, 47, 62, 65, 80, 83, 98, 101, 116, 119, 134, 137, 152, 155, 170, 173,
188, 191, 206, 209, 224, 227, 242, 245, 260, 263, 278, 281, 296, 299, 314, 317,
32, 335, 350, 353, 368, 371, 386, 389, 404, 407, 422, 425, 440, 443, 458, 461,
476, 479, 494, 497, 512, 515, 530, 533, 548, 551, 566, 569, 584, 587, 602, 605,
620, or 623.
[00116] In another embodiment, the variable domain includes a CDR2 to
be replaced or lacks a CDR2 encoding region and the at least one donor nucleic
acid encodes an amino acid sequence substantially as set out in SEQ ID NO:9, 12,
27, 30, 45, 48, 63, 66, 81, 84, 99,102, 117, 120, 135, 138, 153, 156, 171, 174,
189, 192, 207, 210, 225, 228, 243, 246, 261, 264, 279, 282, 297, 300, 315, 318,
333, 336, 351, 354, 369, 372, 387, 390, 405, 408, 423, 426, 441, 444, 459, 462,
477, 480, 495, 498, 513, 516, 531, 534, 549, 552, 567, 570, 585, 588, 603, 606,
621, or 624.
[00117] In another embodiment, the variable domain includes a CDR3 to
be replaced or lacks a CDR3 encoding region and further comprises a CDR1 to be
replaced or lacks a CDR1 encoding region, where the at least one donor nucleic
acid encodes an amino acid sequence substantially as set out in Tables 1 or 7.
[00118] In another embodiment, the variable domain includes a CDR3 to
be replaced or lacks a CDR3 encoding region and further comprises a CDR2 to be

replaced or lacks a CDR2 encoding region, where the at least one donor nucleic
acid encodes an amino acid sequence substantially as set out in Tables 1 or 7.
[00119] In another embodiment, the variable domain includes a CDR3 to
be replaced or lacks a CDR3 encoding region and further comprises a CDR1 and a
CDR2 to be replaced or lacks a CDR1 and a CDR2 encoding region, where the at
least one donor nucleic acid encodes an amino acid sequence substantially as set
out in Tables 1 or 7.
[00120] Using recombinant DNA methodology, a disclosed CDR
sequence may be introduced into a repertoire of VH or VL domains lacking the
respective CDR (Marks et al. (BioTechnology (1992) 10: 779-783). For example, a
primer adjacent to the 5' end of the variable domain and a primer to the third FR
can be used to generate a repertoire of variable domain sequences lacking C0R3.
This repertoire can be combined with a CDR3 of a disclosed antibody. Using
analogous techniques, portions of a disclosed COR sequence may be shuffled with
portions of CDR sequences from other antibodies to provide a repertoire of
antigen-binding fragments that bind IL-22. Either repertoire can be expressed in a
host system such as phage display (described in WO 92/01047 and its
corresponding U.S. Patent No. 5,969,108) so suitable antigen-binding fragments
that bind to IL-22 can be selected.
[00121] A further alternative uses random mutagenesis of the disclosed
VH or VL sequences to generate variant VH or VL domains still capable of binding
IL-22. A technique using error-prone PCR is described by Gram et al. (Proc. Nat.
Acad. Sci. U.S.A. (1992) 89: 3576-3580).
[00122] Another method uses direct mutagenesis of the disclosed VH or
VL sequences. Such techniques are disclosed by Barbas et al. (Proc. Nat. Acad.
Sci. U.S.A. (1994) 91: 3809-3813) and Schier et al. (J. Mol. Biol. (1996) 263:
551-567).
[00123] A portion of a variable domain will comprise at least one CDR
region substantially as set out herein and, optionally, intervening framework
regions from the VH or VL domains as set out herein. The portion may include the
C-terminal half of FR1 and/or the N-terminal half of FR4. Additional residues at the
N-terminal or C-terminal end of the variable domain may not be the same residues
found in naturally occurring antibodies. For example, construction of antibodies by

recombinant DNA techniques often introduces N- or C-terminal residues from its
uses of linkers. Some linkers may be used to join variable domains to other variable
domains (e.g., diabodies), constant domains, or proteinaceous labels.
[00124] Although the embodiments illustrated in the Examples comprise
a "matching" pair of VH and VL domains, a skilled artisan will recognize that
alternative embodiments may comprise antigen-binding fragments containing only
a single CDR from either VL or VH domain. Either one of the single chain specific
antigen-binding domains can be used to screen for complementary domains
capable of forming a two-domain specific antigen-binding fragment capable of, for
example, binding to IL-22. The screening may be accomplished by phage display
screening methods using the so-called hierarchical dual combinatorial approach
disclosed in WO 92/01047. In this approach, an individual colony containing either
a H or L chain clone is used to infect a complete library of clones encoding the
other chain (L or H), and the resulting two-chain specific antigen-binding domain is
selected in accordance with phage display techniques as described.
[00125] In some alternative embodiments, the anti-IL-22 antibodies can
be linked to a protein (e.g., albumin) by chemical cross-linking or recombinant
methods. The disclosed antibodies may also be linked to a variety of
nonproteinaceous polymers (e.g., polyethylene glycol, polypropylene glycol, or
polyoxyalkylenes) in manners set forth in U.S. Patent Nos. 4,640,835; 4,496,689;
4,301,144; 4,670,417; 4,791,192; or 4,179,337. The antibodies can be chemically
modified by covalent conjugation to a polymer, for example, to increase their half-
life in blood circulation. Exemplary polymers and attachment methods are shown
in U.S. Pat. Nos. 4,766,106; 4,179,337; 4,495,285; and 4,609,546.
[00126] The disclosed antibodies can be modified to alter their
glycosylation; that is, at least one carbohydrate moiety can be deleted or added to
the antibody. Deletion or addition of glycosylation sites can be accomplished by
changing amino acid sequence to delete or create glycosylation consensus sites,
which are well known in the art. Another means of adding carbohydrate moieties is
the chemical or enzymatic coupling of glycosides to amino acid residues of the
antibody (see WO 87/05330 and Aplin et al. (1981) CRC Crit. Rev. Biochem., 22:
259-306). Removal of carbohydrate moieties can also be accomplished chemically
or enzymatically (see Hakimuddin et al. (1987) Arch. Biochem. Biophys., 259: 52;

Edge et al. (1981) Anal. Biochem., 118: 131; Thotakura et al. (1987) Meth.
Enzymol., 138: 350).
[00127] Methods for altering an antibody constant region are known in
the art. Antibodies with altered function (e.g., altered affinity for an effector ligand
such as FcR on a cell or the C1 component of complement) can be produced by
replacing at least one amino acid residue in the constant portion of the antibody
with a different residue (see e.g., EP 388,151 A1, US 5,624,821 and US
5,648,260). Similar types of alterations could be described which if applied to a
murine or other species antibody would reduce or eliminate similar functions.
[00128] For example, it is possible to alter the affinity of an Fc region of
an antibody (e.g., an IgG, such as a human IgG) for FcR (e.g., Fc gamma R1) or
C1q. The affinity may be altered by replacing at least one specified residue with at
least one residue having an appropriate functionality on its side chain, or by
introducing a charged functional group, such as glutamate or aspartate, or perhaps
an aromatic non-polar residue such as phenylalanine, tyrosine, tryptophan or
alanine (see e.g., US 5,624,821).
[00129] For example, replacing residue 297 (asparagine) with alanine in
the IgG constant region significantly inhibits recruitment of effector cells, while only
slightly reducing (about three fold weaker) affinity for Clq (see e.g., US 5,624,821).
The numbering of the residues in the heavy chain is that of the EU index (see
Kabat et al., 1991 supra). This alteration destroys the glycosylation site and it is
believed that the presence of carbohydrate is required for Fc receptor binding. Any
other substitution at this site that destroys the glycosylation site is believed to
cause a similar decrease in lytic activity. Other amino acid substitutions, e.g.,
changing any one of residues 318 (Glu), 320 (Lys) and 322 (Lys), to Ala, are also
known to abolish Clq binding to the Fc region of IgG antibodies (see e.g., US
5,624,821).
[00130] Modified antibodies can be produced which have a reduced
interaction with an Fc receptor. For example, it has been shown that in human
lgG3, which binds to the human Fc gamma R1 receptor, changing Leu 235 to Glu
destroys its interaction with the receptor. Mutations on adjacent or close sites in
the hinge link region of an antibody (e.g., replacing residues 234, 236 or 237 with
Ala) can also be used to affect antibody affinity for the Fc gamma R1 receptor. The

numbering of the residues in the heavy chain is based in the EU index (see Kabat
et al., 1991 supra).
[00131] Additional methods for altering the lytic activity of an antibody,
for example, by altering at least one amino acid in the N-terminal region of the CH2
domain, are described in WO 94/29351 by Morgan et al. and US 5,624,821.
[00132] The antibodies of this invention may be tagged with a detectable
or functional label. These labels include radiolabels (e.g., 131lor 99Tc), enzymatic
labels (e.g., horseradish peroxidase or alkaline phosphatase), and other chemical
moieties (e.g., biotin).
[00133] The invention may also feature an isolated antibody that binds to
IL-22, in particular, human IL-22. In certain embodiments, the anti-IL-22 antibody
may have at least one of the following characteristics: (1) it is a monoclonal or
single specificity antibody; (2) it is a human antibody; (3) it is an in vitro generated
antibody; (4) it is an in vivo generated antibody (e.g., transgenic mouse system);
(5) it binds to IL-22 with an association constant of at least 1012 M-1; (6) it binds to
IL-22 with an association constant of at least 1011 M-1; (7) it binds to IL-22 with an
association constant of at least 1010 M-1; (8) it binds to IL-22 with an association
constant of at least 109 M-1; (9) it binds to IL-22 with an association constant of at
least 106 M-1; (10) it binds to IL-22 with a dissociation constant of 500 nM or less;
(11) it binds to IL-22 with a dissociation constant of 10 nM or less; (12) it binds to
IL-22 with a dissociation constant of 150 pM or less; (13) it binds to IL-22 with a
dissociation constant of 60 pM or less; (14) it inhibits binding of IL-22 to IL-22R or a
receptor complex of IL-22R and IL-10R2 with an IC50 of 10 nM or less; (15) it
blocks IL-22 mediated proliferation of IL-22 receptor engineered BaF3 cells with an
IC50 of 1 nM or less in one embodiment, with an IC50 of 150 pM or less in another
embodiment, with an IC50 of 100 pM or less in another embodiment, and with an
IC50 of 10 pM or less in another embodiment; and (16) it blocks IL-22 mediated
GROa secretion from HT29 with an IC50 of 1 nM or less in one embodiment, with
an IC50 of 150 pM or less in another embodiment, and with an IC50 of 10 pM or less
in another embodiment.
[00134] One of skill in the art will appreciate that the modifications
described above are not all-exhaustive, and that many other modifications are
obvious to a skilled artisan in light of the teachings of the present disclosure.

III. Nucleic Acids, Cloning and Expression Systems
[00135] The disclosure provides isolated nucleic acids encoding the
disclosed antibodies. The nucleic acids may comprise DNA or RNA, and they may
be synthetic (completely or partially) or recombinant (completely or partially).
Reference to a nucleotide sequence as set out herein encompasses a DNA
molecule with the specified sequence, and encompasses a RNA molecule with the
specified sequence in which U is substituted for T.
[00136] Also provided are nucleic acids that comprise a coding
sequence for one, two, or three CDR's, a VH domain, a VL domain, or combinations
thereof, as disclosed herein, or a sequence substantially identical thereto (e.g., a
sequence at least 85%, 90%, 95%, 96%, 97%, 98%, 99% or higher identical
thereto, or which is capable of hybridizing under stringent conditions to the
sequences disclosed).
[00137] In one embodiment, the isolated nucleic acids have nucleotide
sequences encoding heavy chain and light chain variable regions of an anti-IL-22
antibody having at least one CDR chosen from the amino acid sequences of SEQ
ID NO: 8-13, 26-31, 44-49,62-67, 80-85, 98-103,116-121, 134-139,152-157,170-
175, 188-193, 206-211, 224-229, 242-247, 260-265, 278-283, 296-301, 314-319,
332-337, 350-355, 368-373, 386-391, 404-409, 422-427, 440-445, 458-463, 476-
481, 494-499, 512-517, 530-535, 548-553, 566-571, 584-589, 602-607, or 620-
625; or sequence encoding a CDR which differs by one or two amino acids from
the sequences described herein.
[00138] The nucleic acid can encode only the light chain or the heavy
chain variable region, or can also encode an antibody light or heavy chain constant
region, operatively linked to the corresponding variable region. In one
embodiment, the light chain variable region is linked to a constant region chosen
from a kappa or a lambda constant region. The light chain constant region may
also be a human kappa or lambda type. In another embodiment, the heavy chain
variable region is linked to a heavy chain constant region of an antibody isotype
chosen from IgG (e.g., IgG,, lgG2, lgG3, lgG4), IgM, lgA1, lgA2, IgD, and IgE. The
heavy chain constant region may be an IgG (e.g., an IgG1) isotype.
[00139] The nucleic acid compositions of the present invention, while
often in the native sequence (of cDNA or genomic DNA or mixtures thereof) except

for modified restriction sites and the like, may be mutated in accordance with
standard techniques to provide gene sequences. For coding sequences, these
mutations, may affect amino acid sequence as desired. In particular, nucleotide
sequences substantially identical to or derived from native V, D, J, constant,
switches and other such sequences described herein are contemplated (where
"derived" indicates that a sequence is identical or modified from another
sequence).
[00140] In one embodiment, the nucleic acid differs (e.g., differs by
substitution, insertion, or deletion) from that of the sequences provided (e.g., as
follows: by at least one but less than 10, 20, 30, or 40 nucleotides; at least one but
less than 1%, 5%, 10% or 20% of the nucleotides in the subject nucleic acid). If
necessary for this analysis the sequences should be aligned for maximum homology.
"Looped" out sequences from deletions or insertions, or mismatches, are considered
differences. The difference may be at a nucleotide(s) encoding a non-essential
residue(s), or the difference may be a conservative substitution(s).
[00141] The disclosure also provides nucleic acid constructs in the form
of plasmids, vectors, transcription or expression cassettes, which comprise at least
one nucleic acid as described herein.
[00142] The disclosure further provides a host cell that comprises at
least one nucleic acid construct described herein.
[00143] Also provided are the methods of making the encoded protein(s)
from the nucleic acid(s) comprising sequence described herein. The method
comprises culturing host cells under appropriate conditions so they express the
protein from the nucleic acid. Following expression and production, the VH or VL
domain, or specific binding member may be isolated and/or purified using any
suitable technique, then used as appropriate. The method can also include the
steps of fusing a nucleic acid encoding a scFv with nucleic acids encoding a Fc
portion of an antibody and expressing the fused nucleic acid in a cell. The method
can also include a step of germlining.
[00144] Antigen-binding fragments, VH and/or VL domains, and encoding
nucleic acid molecules and vectors may be isolated and/or purified from their
natural environment, in substantially pure or homogenous form, or, in the case of

nucleic acid, free or substantially free of nucleic acid or genes of origin other than
the sequence encoding a polypeptide with the require function.
[00145] Systems for cloning and expressing polypeptides in a variety of
host cells are known in the art. Cells suitable for producing antibodies are
described in, for example, Fernandez et al. (1999) Gene Expression Systems,
Academic Press, eds. In brief, suitable host cells include mammalian cells, insect
cells, plant cells, yeast cells, or prokaryotic cells, e.g., E. coli. Mammalian cells
available in the art for heterologous polypeptide expression include lymphocytic cell
lines (e.g., NSO), HEK293 cells, Chinese hamster ovary (CHO) cells, COS cells,
HeLa cells, baby hamster kidney cells, oocyte cells, and cells from a transgenic
animal, e.g., mammary epithelial cell. In one embodiment, the GIL01, GIL16,
GIL45, GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03, 367D04,
368D04,166B06, 166G05, 375G06, 376B10, 354A08, 355B06, 355E04, and
356A11 antibodies are expressed in HEK293 or CHO cells. In another
embodiment, a selection of antibodies chosen from 365A11, 354A08, 087B03, and
368D04 are expressed in HEK293 or CHO cells. In other embodiments, the
nucleic acids encoding the antibodies of the invention are placed under the control
of a tissue-specific promoter (e.g., a mammary specific promoter) and the
antibodies are produced in transgenic animals. For example, the antibodies are
secreted into the milk of the transgenic animal, such as a transgenic cow, pig,
horse, sheep, goat or rodent.
[00146] Suitable vectors may be chosen or constructed to contain
appropriate regulatory sequences, including promoter sequences, terminator
sequences, polyadenylation sequences, enhancer sequences, marker genes, and
other sequences. The vectors may also contain a plasmid or viral backbone. For
details, see Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd ed.,
Cold Spring Harbor Laboratory Press (1989). Many established techniques used
with vectors, including the manipulation, preparation, mutagenesis, sequencing,
and transfection of DNA, are described in Current Protocols in Molecular Biology,
Second Edition, Ausubel et al. eds., John Wiley & Sons (1992).
[00147] A further aspect of the disclosure provides a method of
introducing the nucleic acid into a host cell. For eukaryotic cells, suitable
transfection techniques may include calcium phosphate, DEAE-Dextran,

electroporation, liposome-mediated transfection, and transduction using retro virus
or other viruses, e.g., vaccinia or baculovirus. For bacterial cells, suitable
techniques may include calcium chloride transformation, electroporation, and
transfection using bacteriophage. DNA introduction may be followed by a selection
method (e.g., drug resistance) to select cells that contain the nucleic acid.
IV. Uses of Anti-IL-22 Antibodies
[00148] Anti-IL-22 antibodies that act as antagonists to IL-22 can be
used to regulate at least one IL-22-mediated immune response, such as acting on
epithelial cells in solid tissue and indirectly modulating downstream immune
responses, such as blocking expansion of T cell subsets, including, for example,
TH17 T cells.. In one embodiment, antibodies of the invention are used in a
method for regulating an immune response, the method comprising contacting IL-
22 with an antibody of the invention thereby regulating the immune response. In
one embodiment, the immune response comprises cell proliferation, cytolytic
activity, cytokine secretion, or chemokine secretion.
[00149] Accordingly, the antibodies of the invention can be used to
directly or indirectly inhibit the activity (e.g., proliferation, differentiation, and/or
survival) of an immune or hematopoietic cell (e.g., a cell of myeloid, lymphoid, or
erythroid lineage, or precursor cells thereof), and, thus, can be used to treat a
variety of immune disorders and hyperproliferative disorders. Non-limiting
examples of immune disorders that can be treated include, but are not limited to,
autoimmune disorders, e.g., arthritis (including rheumatoid arthritis, juvenile
rheumatoid arthritis, osteoarthritis, psoriatic arthritis, lupus-associated arthritis or
ankylosing spondylitis), scleroderma, systemic lupus erythematosis, HIV, Sjogren's
syndrome, vasculitis, multiple sclerosis, autoimmune thyroiditis, dermatitis
(including atopic dermatitis and eczematous dermatitis), myasthenia gravis,
inflammatory bowel disease (IBD), Crohn's disease, colitis, diabetes mellitus (type
I); inflammatory conditions of, e.g., the skin (e.g., psoriasis), cardiovascular system
(e.g., atherosclerosis), nervous system (e.g., Alzheimer's disease), liver (e.g.,
hepatitis), kidney (e.g., nephritis) and pancreas (e.g., pancreatitis); cardiovascular
disorders, e.g., cholesterol metabolic disorders, oxygen free radical injury,
ischemia; disorders associated with wound healing; respiratory disorders, e.g.,
asthma and COPD (e.g., cystic fibrosis); acute inflammatory conditions (e.g.,

endotoxemia, sepsis and septicaemia, toxic shock syndrome and infectious
disease); transplant rejection and allergy. In one embodiment, the IL-22-
associated disorder is, an arthritic disorder, e.g., a disorder chosen from one or
more of rheumatoid arthritis, juvenile rheumatoid arthritis, osteoarthritis, psoriatic
arthritis, orankylosing spondylitis; a respiratory disorder (e.g., asthma, chronic
obstructive pulmonary disease (COPD); or an inflammatory condition of, e.g., the
skin (e.g., psoriasis), cardiovascular system (e.g., atherosclerosis), nervous system
(e.g., Alzheimer's disease), liver (e.g., hepatitis), kidney (e.g., nephritis), pancreas
(e.g., pancreatitis), and gastrointestinal organs, e.g., colitis, Crohn's disease and
IBD; acute inflammatory conditions, e.g., endotoxemia, sepsis and septicaemia,
toxic shock syndrome and infectious disease; multiple organ failure; respiratory
disease (ARD); amyloidosis; nephropathies such as glomerulosclerosis,
membranous neuropathy, renal arteriosclerosis, glomerulonephritis,
fibroproliferative diseases of the kidney, as well as other kidney disfunctions and
renal tumors. Because of IL-22's effects on epithelia, anti-lL-22 antibodies can be
used to treat epithelial cancers, e.g., carcinoma, melanoma and others. For a
description of a rationale for IL-22 inhibition in these and other disease states see
WO 03/083062 (pages 58-75).
[00150] Multiple sclerosis is a central nervous system disease that is
characterized by inflammation and loss of myelin sheaths—the fatty material that
insulates nerves and is needed for proper nerve function. Inflammation that results
from an immune response that is dependent on IL-22 can be treated with the
antibodies and compositions of this invention. In the experimental autoimmune
encephalitis (EAE) mouse model for multiple sclerosis (Tuohy et al. (J. Immunol.
(1988) 141: 1126-1130), Sobel etal. (J. Immunol. (1984) 132: 2393-2401), and
Tratigott (Cell Immunol. (1989) 119: 114-129), treatment of mice with GIL01,
GIL16, GIL45, GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03, 367D04,
368D04, 166B06, 166G05, 375G06, 376B10, 354A08, 355BO6, 355E04, or
356A11 injections prior (and continuously) to EAE induction may profoundly delay
the onset of the disease. This can serve as a model for confirming use of the
antibody of the invention. The antibodies of this invention may similarly be used to
treat multiple sclerosis in humans.

[00151] Arthritis is a disease characterized by inflammation in the joints.
Rheumatoid Arthritis (RA) is the most frequent form of arthritis, involving
inflammation of connective tissue and the synovial membrane, a membrane that
lines the joint. The inflamed synovial membrane often infiltrates the joint and
damages joint cartilage and bone. IL-22 and IL-22R protein and/or transcript is
associated with both human diseases. In RA synovial biopsies, IL-22 protein is
detected in vimentin+ synovial fibroblasts and some CD68+ macrophages while IL-
22R is detected in synovial fibroblasts. Treatment of synovial fibroblasts with IL-22
induces the production of monocyte chemoattractant protein-1, MCP-1, as well as
general metabolic activity (Ikeuchi, H., et al. (2005) Arthritis Rheum. 52:1037-46).
Inhibitors of IL-22 ameliorate symptoms of rheumatoid arthritis (WO 2005/000897
A2; U.S. Patent No. 6,939,545). Increased secretion of inflammatory cytokines and
chemokines, and more importantly, increased disease resulting from immune
responses that are dependent on IL-22 may be treated with the antibodies of this
invention. Similarly, the antibodies and compositions of this invention may be used
to treat RA or other arthritic diseases in humans.
[00152] Transplant rejection is the immunological phenomenon where
tissues from a donor are specifically "attacked" by immune cells of the host. The
principle "attacking" cells are T cells, whose T cell receptors recognize the donor's
MHC molecules as "foreign." This recognition activates the T cells, which
proliferate and secrete a variety of cytokines and cytolytic proteins that ultimately
destroy the transplant. MLR and transplantation models have been described by
Current Protocols in Immunology, Second Edition, Coligan et al. eds., John Wiley &
Sons, 1994; Kasaian et al. (Immunity (2002) 16: 559-569); Fulmer et al. (Am. J.
Anat. (1963) 113: 273-285), and Lenschow et al. (Science (1992) 257: 789-792).
The antibodies and compositions of this invention may be used to reduce the MLR
and treat transplant rejection and related diseases (e.g., graft versus host disease)
in humans that are dependent on IL-22.
[00153] The antibodies of this invention can also be used to treat
hyperproliferative disorders associated with aberrant activity of IL-22-responsive
cells and IL-22R/IL-10R2-responsive cells by adminstering the antibodies in an
amount sufficient to inhibit or reduce hyperproliferation of IL-22 and/or IL-22R
and/or IL-10R2-responsive cells in a subject and allowing the antibodies to treat or

prevent the disorder. IL-22 and 1L-22R expression is constitutive on epithelial cells
in a number of tissues including, but not limited to, pancreas, lung, skin, gut, liver,
kidney (Kotenko, S.V. et al. (2001) J. Biol Chem. 276:2725-32; Xie, M.H. et al.
(2000) J. Biol. Chem. 275:31335-9; Wolk, K. et al. (2004) Immunity 21:241 -54). In
addition, IL-22 receptor complex is also expressed on the surface of fibrobiasts
from the diseased joint and normal gut (Ikeuchi, H. et al. (2005) Arthritis Rheum.
52:1037-46; Andoh, A. et ah (2005) Gastroenterology 129:969-84). Neopiastic
derivatives of these cell types may be hyper responsive to IL-22, modulating these
cells ability to survive in the organism. Hence antibodies to IL-22 may be used to
inhibit the progression of such neoplasms, e.g. squamous cell carcinomas, basal
cell carcinomas, transitional cell papillomas and carcinomas, adenomas,
adenocarcinoma, linitis plastica, insulinoma, glucagonoma, gastrinoma, vipoma,
cholangiocarcinoma, hepatocellular carcinoma, adenoid cyctic carcinoma,
carcinoid tumor of appendix, prolactinoma, oncocytoma, hurthle cell adenoma,
renal cell carcinoma, Grawitz tumor, multiple endocrine adenomas, endometroid
adenoma, adnexal and skin appendage neoplasms, mucoepidermoid neoplams,
cystic, mucinous and serous neoplasms, cystadenoma, pseudomyxoma peritonei,
ductal, lobular and medullary neoplasms, acinar cell neoplasns, complex epithelial
neoplasms, Warthin's tumor, thymoma, specialized gonadal neoplasms, sex cord-
stromai tumor, thecoma, granulosa cell tumor, arrhenoblastoma, sertoli-leydig cell
tumor, paraganglioma, pheochromocytoma, glomus tumor, malanocytic nevus,
malignant melanoma, melanoma, nodular melanoma, dysplastic nevus, lentigo
maligna, superficial spreading melanoma, or acral lentiginous melanoma. While
the IL-22 receptor is not detected on ex vivo naive or activated immune cells,
dysregulation of the receptor might make such derivative neopiastic cells
responsive to IL-22 and thus inhibition by an antibody to IL-22.
[00154] In another aspect, the invention features a method of
decreasing, inhibiting or reducing an acute phase response in a subject. The
method includes administering to the subject an anti-IL-22 antibody or fragment
thereof as described herein, in an amount sufficient to decrease, inhibit or reduce
the acute phase response in the subject. In one embodiment, the subject is a
mammal, e.g., a human suffering from an IL-22-associated disorder as described
herein, including, e.g., respiratory disorders, inflammatory disorders and

autoimmune disorders. In one embodiment, the IL-22 binding agent is
administered locally, e.g., topically, subcutaneously, or other administrations that
are not in the general circulation.
[00155] IL-22 is believed to exert its inflammatory effects locally, e.g. by
acting (e.g., directly acting) as a modular or a regulator of tissue inflammation as
opposed to direct systemic effects. Accordingly, inhibition of IL-22 activity using,
e.g. an anti-IL-22 antibody of the present invention may provide a more effective
(e.g., less toxic) tissue-specific, anti-inflammatory agent than systemic anti-
inflammatory modalities. Furthermore, inhibition of local IL-22 using, e.g., an anti-
IL-22 antibody or fragment thereof described herein, may provide a useful
candidate for combination with systemic anti-inflammatory modalities.
V. Combination Therapy
[00156] In one embodiment, a pharmaceutical composition comprising at
least one anti-IL-22 antibody and at least one therapeutic agent is administered in
combination therapy. The therapy is useful for treating pathological conditions or
disorders, such as immune and inflammatory disorders. The term "in combination"
in this context means that the antibody composition and the therapeutic agent are
given substantially contemporaneously, either simultaneously or sequentially. In
one embodiment, if given sequentially, at the onset of administration of the second
compound, the first of the two compounds is still detectable at effective
concentrations at the site of treatment. In another embodiment, if given
sequentially, at the onset of administration of the second compound, the first of the
two compounds is not detectable at effective concentrations at the site of
treatment.
[00157] For example, the combination therapy can include at least one
anti-IL-22 antibody co-formulated with, and/or co-administered with, at least one
additional therapeutic agent. The additional agents may include at least one
cytokine inhibitor, growth factor inhibitor, immunosuppressant, anti-inflammatory
agent, metabolic inhibitor, enzyme inhibitor, cytotoxic agent, and cytostatic agent,
as described in more detail below. In one embodiment, the additional agent is a
standard treatment for arthritis, including, but not limited to, non-steroidal anti-
inflammatory agents (NSAIDs); corticosteroids, including prednisolone, prednisone,
cortisone, and triamcinolone; and disease modifying anti-rheumatic drugs

(DMARDs), such as methotrexate, hydroxychloroquine (Plaquenil) and
sulfasalazine, leflunomide (Arava), tumor necrosis factor inhibitors, including
etanercept (Enbrel), infliximab (Remicade) (with or without methotrexate), and
adalimumab (Humira), anti-CD20 antibody (e.g., Rituxan), soluble interleukin-1
receptor, such as anakinra (Kineret), gold, minocycline (Minocin), penicillamine,
and cytotoxic agents, including azathioprine, cyclophosphamide, and cyclosporine.
Such combination therapies may advantageously utilize lower dosages of the
administered therapeutic agents, thus avoiding possible toxicities or complications
associated with the various monotherapies. Moreover, the additional therapeutic
agents disclosed herein act on pathways in addition to or that differ from the IL-
22/IL-22R/1L-10R2 pathway, and thus are expected to enhance and/or synergize
with the effects of the anti-IL-22 antibodies.
[00158] Therapeutic agents used in combination with anti-IL-22
antibodies may be those agents that interfere at different stages in the autoimmune
and subsequent inflammatory response. In one embodiment, at least one anti-IL-
22 antibody described herein may be co-formulated with, and/or co-administered
with, at least one cytokine and/or growth factor antagonist. The antagonists may
include soluble receptors, peptide inhibitors, small molecules, ligand fusions,
antibodies and binding fragments thereof (that bind cytokines or growth factors or
their receptors or other cell surface molecules), and "anti-inflammatory cytokines"
and agonists thereof.
[00159] Non-limiting examples of the agents that can be used in
combination with the anti-IL-22 antibodies described herein, include, but are not
limited to, antagonists of at least one interteukin (e.g., IL-1, IL-2, IL-6, lL-7, IL-8, IL-
12 (or one of its subunits p35 or p40), IL-13, IL-15, IL-16, IL-17A-F (including
heterodimers thereof, for example, IL-17A/IL-17F heterodimer), IL-18, IL-19, IL-20,
IL-21, and IL-23 (or one of its subunits p19 or p40)); cytokine (e.g., TNFα, LT,
EMAP-II, and GM-CSF); and growth factor (e.g., FGF and PDGF). The agents
may also include, but not limited to, antagonists of at least one receptor for an
interieukin, cytokine, and growth factor. Anti-IL-22 antibodies can also be
combined with inhibitors (e.g., antibodies or binding fragments thereof) to cell
surface molecules such as CD2, CD3, CD4, CD8, CD20 (e.g. Rituxan), CD25,
CD28, CD30, CD40, CD45, CD69, CD80 (B7.1), CD86 (B7.2), CD90, or their

ligands (e.g., CD154 (gp39, CD40L)), or LFA-1/ICAM-1 and VLA-4/VCAM-1
(Yusuf-Makagiansar et al. (2002) Med Res Rev 22(2):146-67)). In certain
embodiments, antagonists that can be used in combination with anti-IL-22
antibodies described herein may include antagonists of IL-1, IL-12 (or one of its
subunits p35 or p40), TNFα, IL-15, IL-17A-F (including heterodimers thereof, for
example, IL-17A/IL-17F heterodimer), IL-18, IL-19, IL-20, IL-21, and IL-23 (or one
of its subunits p19 or p40), and their receptors.
[00160] Examples of those agents include IL-12 antagonists (such as
antibodies that bind IL-12 (see e.g., WO 00/56772) or one of its subunits p35 or
p40); IL-12 receptor inhibitors (such as antibodies to the IL-12 receptor); and
soluble IL-12 receptor and fragments thereof. Examples of IL-15 antagonists
include antibodies against IL-15 or its receptor, soluble fragments of the IL-15
receptor, and IL-15-binding proteins. Examples of IL-18 antagonists include
antibodies to IL-18, soluble fragments of the IL-18 receptor, and IL-18 binding
proteins (IL-18BP, Mallet et al. (2001) Circ. Res. 28). Examples of IL-1 antagonists
include lnterleukin-1-Converting Enzyme (ICE) inhibitors (such as Vx740), IL-1
antagonists (e.g., IL-1 RA (ANIKINRA, AMGEN)), sIL-IRII (Immunex), and anti-IL-1
receptor antibodies.
[00161] In one embodiment, the combination therapy includes at least
one anti-IL-22 antibody co-formulated with, and/or co-administered with an
antagonist, such as an antibody or antigen binding fragment thereof or a soluble
receptor, of at least one of IL-17A, IL-17F, IL-17A/IL-17F heterodimer, or IL-23 (or
one of its subunits p19 or p40).
[00162] Examples of TNF antagonists include antibodies to TNF (e.g.,
human TNFa), such as D2E7 (human anti-TNFα antibody, U.S. 6,258,562,
Humira™, BASF); CDP-571 /CDP-870/BAY-10-3356 (humanized anti-TNFα
antibodies, Celltech/Pharmacia); cA2 (chimeric anti-TNFa antibody, Remicade™,
Centocor); and anti-TNF antibody fragments (e.g., CPD870). Other examples
include soluble TNF receptor (e.g., human p55 or p75) fragments and derivatives,
such as p55 kdTNFR-IgG (55 kD TNF receptor-IgG fusion protein, Lenercept™)
and 75 kdTNFR-IgG (75 kD TNF receptor-IgG fusion protein, Enbrel™, Immunex,
see, e.g., Arthritis & Rheumatism (1994) Vol. 37, S295; J. Invest. Med. (1996) Vol.
44, 235A). Further examples include enzyme antagonists (e.g., TNFa converting

enzyme inhibitors (TACE) such as alpha-sulfonyl hydroxamic acid derivative (WO
01/55112) or N-hydroxyformamide inhibitor (GW 3333, -005, or -022)) and TNF-
bp/s-TNFR (soluble TNF binding protein, see e.g., Arthritis & Rheumatism (1996)
Vol. 39, No. 9 (supplement), S284; and Am. J. Physiol. Heart Circ. Physiol. (1995)
Vol. 268, pp. 37-42). TNF antagonists may be soluble TNF receptor (e.g., human
p55 or p75) fragments and derivatives, such as 75 kdTNFR-IgG; and TNFα
converting enzyme (TACE) inhibitors.
[00163] In other embodiments, the anti-IL-22 antibodies described
herein can be administered in combination with at least one of the following: IL-13
antagonists, such as soluble IL-13 receptors and/or anti-IL-13 antibodies; and IL-2
antagonists, such as IL-2 fusion proteins (e.g., DAB 486-IL-2 and/or DAB 389-IL-2,
Seragen, see e.g., Arthritis & Rheumatism (1993) Vol. 36,1223) and anti-IL-2R
antibodies (e.g., anti-Tac (humanized antibody, Protein Design Labs, see Cancer
Res. 1990 Mar 1,-50(5).1495-502)). Another combination includes anti-IL-22
antibodies in combination with non-depleting anti-CD4 inhibitors such as IDEC-
CE9.1/SB 210396 (anti-CD4 antibody, IDEC/SmithKline). Yet other combinations
include anti-IL-22 antibodies with antagonists (such as antibodies, soluble
receptors, or antagonistic ligands) of costimulatory molecules, such as CD80
(B7.1) and CD86 (B7.2); ICOSL, ICOS, CD28, and CTLA4 (e.g., CTLA4-lg);
P-selectin glycoprotein iigand (PSGL); and anti-inflammatory cytokines and
agonists thereof (e.g., antibodies). The anti-inflammatory cytokines may include
IL-4 (DNAX/Schering); IL-10 (SCH 52000, recombinant IL-10, DNAX/Schering); IL-
13; and TGF.
[00164] In other embodiments, at least one anti-IL-22 antibody can be
co-formulated with, and/or co-administered with, at least one anti-inflammatory
drug, immunosuppressant, metabolic inhibitor, and enzymatic inhibitor. Non-
limiting examples of the drugs or inhibitors that can be used in combination with the
IL-22 antagonists described herein, include, but are not limited to, at least one of:
non-steroidal anti-inflammatory drug (NSAID) (such as ibuprofen, Tenidap (see
e.g., Arthritis & Rheumatism (1996) Vol. 39, No. 9 (supplement), S280)), Naproxen
(see e.g., Neuro Report (1996) Vol. 7, pp. 1209-1213), Meloxicam, Piroxicam,
Diclofenac, and (ndomethacin); Sulfasalazine (see e.g., Arthritis & Rheumatism
(1996) Vol. 39, No. 9 (supplement), S281); corticosteroid (such as prednisolone);

cytokine suppressive anti-inflammatory drug (CSAID); and an inhibitor of nudeotide
biosynthesis (such as an inhibitor of purine biosynthesis (e.g., folate antagonist
such as methotrexate) and an inhibitor of pyrimidine biosynthesis (e.g., a
dihydroorotate dehydrogenase (DHODH) inhibitor such as leflunomide (see e.g.,
Arthritis & Rheumatism (1996) Vol. 39, No. 9 (supplement), S131; Inflammation
Research (1996) Vol. 45, pp. 103-107)). Therapeutic agents for use in combination
with IL-22/IL-22R or IL-22/IL-10R2 antagonists may include NSAIDs, CSAIDs,
DHODH inhibitors (such as leflunomide), and folate antagonists (such as
methotrexate).
[00165] Examples of additional inhibitors include at least one of:
corticosteroid (oral, inhaled and local injection); immunosuppressant (such as
cyclosporin and tacrolimus (FK-506)); a mTOR inhibitor (such as sirolimus
(rapamycin) or a rapamycin derivative (e.g., ester rapamycin derivative such as
CC1-779 (Bit L. (2002) Current Opinion Investig. Drugs 3(8): 1249-53; Huang, S. et
al. (2002) Current Opinion Investig. Drugs 3(2):295-304))); an agent which
interferes with the signaling of proinflammatory cytokines such as TNFα and 1L-1
(e.g., IRAK, NIK, IKK, p38 or a MAP kinase inhibitor); a COX2 inhibitor (e.g.,
celecoxib and variants thereof (MK-966), see e.g., Arthritis & Rheumatism (1996)
Vol. 39, No. 9 (supplement), S81); a phosphodiesterase inhibitor (such as
R973401, see e.g., Arthritis & Rheumatism (1996) Vol. 39, No. 9 (supplement),
S282)); a phospholipase inhibitor (e.g., an inhibitor of cytosolic phosphoiipase 2
(cPLA2) such as trifluoromethyl ketone analogs (U.S. 6,350,892)); an inhibitor of
vascular endothelial cell growth factor (VEGF); an inhibitor of the VEGF receptor;
and an inhibitor of angiogenesis. Therapeutic agents for use in combination with
anti-IL-22 antibodies may include immunosuppresants (such as cyclosporine and
tacrolimus (FK-506)); and mTOR inhibitors (such as sirolimus (rapamycin) or
rapamycin derivatives (e.g., ester rapamycin derivatives such as CCI-779)); COX2
inhibitors (such as celecoxib and variants thereof); and phospholipase inhibitors
(such as inhibitors of cytosolic phospholipase 2 (cPLA2) (e.g., trifluoromethyl
ketone analogs)).
[00166] Examples of therapeutic agents that can be co-administered
and/or co-formulated with at least one anti-IL-22 antibody, include, but are not
limited to, at least one of: TNF antagonists (such as anti-TNF antibodies); soluble

fragments of TNF receptors (e.g., human p55 and p75) and derivatives thereof
(such as p55 kdTNFR-IgG (55 kD TNF receptor-IgG fusion protein, Lenercept™)
and 75 kdTNFR-IgG (75 kD TNF receptor-IgG fusion protein, Enbrel™)); TNF
enzyme antagonists (such as TACE inhibitors); antagonists of IL-12 (or one of its
subunits p35 or p40), IL-15, IL-17A-F (including heterodimers thereof, for example,
IL-17A/IL-17F heterodimer), IL-18, IL-19, IL-20, IL-21, IL-22, and IL-23 (or one of its
subunits p19 or p40); T cell and B cell depleting agents (such as anti-CD4 or anti-
CD22 antibodies); small molecule inhibitors (such as methotrexate and
leflunomide); sirolimus (rapamycin) and analogs thereof (such as CCI-779); Cox-2
and cPLA2 inhibitors; p38, TPL-2, Mk-2 and NFKB inhibitors; RAGE and soluble
RAGE; P-selectin and PSGL-1 inhibitors (such as antibodies to and small molecule
inhibitors); and estrogen receptor beta (ERB) agonists, and ERB-NFkb antagonists.
Therapeutic agents that can be co-administered and/or co-formulated with at least
one anti-IL-22 antibody may include at least one of: a soluble fragment of a TNF
receptor (e.g., human p55 or p75) such as 75 kdTNFR-IgG (75 kD TNF receptor-
IgG fusion protein, Enbrel™); methotrexate; leflunomide; and sirolimus (rapamycin)
and analogs thereof (such as CCI-779).
[00167] The anti-IL-22 antibodies disclosed herein can be used in
combination with other therapeutic agents to treat specific immune disorders as
discussed in further detail below.
[00168] Non-limiting examples of agents for treating arthritic disorders
(e.g., rheumatoid arthritis, inflammatory arthritis, rheumatoid arthritis, juvenile
rheumatoid arthritis, osteoarthritis and psoriatic arthritis), with which an anti-IL-22
antibody can be combined include at least one of the following: TNF antagonists
(such as anti-TNF antibodies); soluble fragments of TNF receptors (e.g., human
p55 and p75) and derivatives thereof (such as p55 kdTNFR-IgG (55 kD TNF
receptor-IgG fusion protein, Lenercept™) and 75 kdTNFR-IgG (75 kD TNF
receptor-IgG fusion protein, Enbrel™)); TNF enzyme antagonists (such as TACE
inhibitors); antagonists of IL-12 (or one of its subunits p35 or p40), IL-15, IL-17A-F
(including heterodimers thereof, for example, IL-17A/1L-17F heterodimer), IL-18, II-
19, IL-20, IL-21, IL-22, IL-23 (or one of its subunits p19 or p40), and IL-24; T cell
and B cell depleting agents (such as anti-CD4, anti-CD20, or anti-CD22
antibodies); small molecule inhibitors (such as methotrexate and leflunomide);

sirolimus (rapamycin) and analogs thereof (e.g., CCI-779); Cox-2 and cPLA2
inhibitors; NSAIDs; p38, TPL-2, Mk-2, and NFKB inhibitors; RAGE or soluble
RAGE; P-selectin or PSGL-1 inhibitors (such as small molecule inhibitors and
antibodies to); estrogen receptor beta (ERB) agonists, and ERB-NFKB antagonists.
Therapeutic agents that can be co-administered and/or co-formulated with at least
one IL-22/IL-22R/IL-10R2 antagonist may include at least one of: a soluble
fragment of a TNF receptor (e.g., human p55 or p75) such as 75 kdTNFR-IgG (75
kD TNF receptor-IgG fusion protein, Enbrel™); methotrexate; leflunomide; and
sirolimus (rapamycin) or an analog thereof (e.g., CCI-779).
[00169] Non-limiting examples of agents for treating multiple sclerosis
with which anti-IL-22 antibody can be combined include interferon-β for example,
IFNβ-1a and IFNβ-1b), copaxone, corticosteroids, IL-1 inhibitors, TNF inhibitors,
antibodies to CD40 ligand, antibodies to CD80, and IL-12 antagonists, including
antibodies that bind IL-12 (or one of its subunits p35 or p40).
[00170] Non-limiting examples of agents for treating inflammatory bowel
disease or Crohn's disease with which an anti-IL-22 antibody can be combined
include budenoside; epidermal growth factor; corticosteroids; cyclosporine;
sulfasalazine; aminosalicylates; 6-mercaptopurine; azathioprine; metronidazole;
lipoxygenase inhibitors; mesalamine; olsalazine; balsalazide; antioxidants;
thromboxane inhibitors; IL-1 receptor antagonists; anti-IL-1 monoclonal antibodies;
anti-IL-6 monoclonal antibodies; growth factors; elastase inhibitors; pyridinyl-
imidazole compounds; TNF antagonists as described herein; IL-4, IL-10, IL-13,
and/or TGFP or agonists thereof (e.g., agonist antibodies); IL-11; glucuronide- or
dextran-conjugated prodrugs of prednisolone, dexamethasone or budesonide;
ICAM-1 antisense phosphorothioate otigodeoxynucleotides (ISIS 2302; Isis
Pharmaceuticals, Inc.); soluble complement receptor 1 (TP10; T Cell Sciences,
Inc.); slow-release mesalazine; methotrexate; antagonists of Platelet Activating
Factor (PAF); ciprofloxacin; and lignocaine.
[00171] In other embodiments, an anti-IL-22 antibody can be used in
combination with at least one antibody directed at other targets involved in
regulating immune responses, e.g., transplant rejection or graft versus host
disease. Non-limiting examples of agents for treating immune responses with
which an IL-22/IL-22R/IL10R2 antagonist of the invention can be combined include

the following: antibodies against cell surface molecules, including but not limited to
CD25 (IL-2 receptor a), CD11a (LFA-1), CD54 (ICAM-1), CD4, CD45,
CD28/CTLA4, CD80 (B7-1), CD86 (B7-2), or combinations thereof. In another
embodiment, an anti-IL-22 antibody is used in combination with at least one
general immunosuppressive agent, such as cyclosporin A or FK506.
[00172] Another aspect of the present invention accordingly relates to
kits for carrying out the combined administration of the anti-IL-22 antibodies with
other therapeutic agents. In one embodiment, the kit comprises at least one anti-
IL-22 antibody formulated in a pharmaceutical carrier, and at least one therapeutic
agent, formulated as appropriate in one or more separate pharmaceutical
preparations.
VI. Diagnostic Uses
[00173] The antibodies may also be used to detect the presence of IL-22
in biological samples. By correlating the presence or level of these proteins with a
medical condition, one of skill in the art can diagnose the associated medical
condition. For example, IL-22 induces changes associated with those caused by
inflammatory cytokines (such as IL-1 and TNFα), and inhibitors of IL-22 ameliorate
symptoms of rheumatoid arthritis (WO 2005/000897 A2). Illustrative medical
conditions that may be diagnosed by the antibodies of this invention include
multiple sclerosis, rheumatoid arthritis, psoriasis, inflammatory bowel disease,
pancreatitis, and transplant rejection.
[00174] Antibody-based detection methods are well known in the art,
and include ELISA, radioimmunoassays, immunoblots, Western blots, flow
cytometry, immunofluorescence, immunoprecipitation, and other related
techniques. The antibodies may be provided in a diagnostic kit that incorporates at
least one of these procedures to detect IL-22. The kit may contain other
components, packaging, instructions, or other material to aid the detection of the
protein and use of the kit.
[00175] Antibodies may be modified with detectable markers, including
ligand groups (e.g., biotin), fluorophores and chromophores, radioisotopes,
electron-dense reagents, or enzymes. Enzymes are detected by their activity. For
example, horseradish peroxidase is detected by its ability to convert
tetramethylbenzidine (TMB) to a blue pigment, quantifiable with a

spectrophotometer. Other suitable binding partners include biotin and avidin, tgG
and protein A, and other receptor-ligand pairs known in the art.
[00176] Antibodies can also be functionally linked (e.g., by chemical
coupling, genetic fusion, non-covalent association or otherwise) to at least one
other molecular entity, such as another antibody (e.g., a bispecific or a multispecific
antibody), toxins, radioisotopes, cytotoxic or cytostatic agents, among others.
Other permutations and possibilities are apparent to those of ordinary skill in the
art, and they are considered equivalents within the scope of this invention
VII. Pharmaceutical Compositions and Methods of Administration
[00177] Certain embodiments of the invention include compositions
comprising the disclosed antibodies. The compositions may be suitable for
pharmaceutical use and administration to patients. The compositions comprise an
antibody of the present invention and a pharmaceutical excipient As used herein,
"pharmaceutical excipient includes solvents, dispersion media, coatings,
antibacterial and antifungal agents, isotonic and absorption delaying agents, etc.,
that are compatible with pharmaceutical administration. Use of these agents for
pharmaceutically active substances is well known in the art. The compositions
may also contain other active compounds providing supplemental, additional, or
enhanced therapeutic functions. The pharmaceutical compositions may also be
included in a container, pack, or dispenser together with instructions for
administration.
[00178] A pharmaceutical composition of the invention is formulated to
be compatible with its intended route of administration. Methods to accomplish the
administration are known to those of ordinary skill in the art. Pharmaceutical
compositions may be topically or orally administered, or capable of transmission
across mucous membranes. Examples of administration of a pharmaceutical
composition include oral ingestion or inhalation. Administration may also be
intravenous, intraperitoneal, intramuscular, intracavity, subcutaneous, cutaneous,
or transdermal.
[00179] Solutions or suspensions used for intradermal or subcutaneous
application typically include at least one of the following components: a sterile
diluent such as water, saline solution, fixed oils, polyethylene glycol, glycerine,
propylene glycol, or other synthetic solvent; antibacterial agents such as benzyl

alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite;
chelating agents such as ethylenediaminetetraacetic acid (EDTA); buffers such as
acetate, citrate, or phosphate; and tonicity agents such as sodium chloride or
dextrose. The pH can be adjusted with acids or bases. Such preparations may be
enclosed in ampoules, disposable syringes, or multiple dose vials.
[00180] Solutions or suspensions used for intravenous administration
include a carrier such as physiological saline, bacteriostatic water, Cremophor
EL™ (BASF, Parsippany, NJ), ethanol, or polyol. In all cases, the composition
must be sterile and fluid for easy syringabiiity. Proper fluidity can often be obtained
using lecithin or surfactants. The composition must also be stable under the
conditions of manufacture and storage. Prevention of microorganisms can be
achieved with antibacterial and antifungal agents, e.g., parabens, chlorobutanol,
phenol, ascorbic acid, thimerosal, etc. In many cases, isotonic agents (sugar),
polyalcohols (mannitol and sorbitol), or sodium chloride may be included in the
composition. Prolonged absorption of the composition can be accomplished by
adding an agent which delays absorption, e.g., aluminum monostearate and
gelatin.
[00181] Oral compositions include an inert diluent or edible carrier. The
composition can be enclosed in gelatin or compressed into tablets. For the
purpose of oral administration, the antibodies can be incorporated with excipients
and placed in tablets, troches, or capsules. Pharmaceutically compatible binding
agents or adjuvant materials can be included in the composition. The tablets,
troches, and capsules, may contain (1) a binder such as microcrystalline cellulose,
gum tragacanth or gelatin; (2) an excipient such as starch or lactose, (3) a
disintegrating agent such as alginic acid, Primogel, or corn starch; (4) a lubricant
such as magnesium stearate; (5) a glidant such as colloidal silicon dioxide; or (6) a
sweetening agent or a flavoring agent.
[00182] The composition may also be administered by a transmucosal or
transdermal route. For example, antibodies that comprise a Fc portion may be
capable of crossing mucous membranes in the intestine, mouth, or lungs (via Fc
receptors). Transmucosal administration can be accomplished through the use of
lozenges, nasal sprays, inhalers, or suppositories. Transdermal administration can
also be accomplished through the use of a composition containing ointments,

salves, gels, or creams known in the art. For transmucosal or transdermal
administration, penetrants appropriate to the barrier to be permeated are used. For
administration by inhalation, the antibodies are delivered in an aerosol spray from a
pressured container or dispenser, which contains a propellant (e.g., liquid or gas)
or a nebulizer.
[00183] In certain embodiments, the antibodies of this invention are
prepared with carriers to protect the antibodies against rapid elimination from the
body. Biodegradable polymers (e.g., ethylene vinyl acetate, polyanhydrides,
polyglycolic acid, collagen, polyorthoesters, polylactic acid) are often used.
Methods for the preparation of such formulations are known by those skilled in the
art. Liposomal suspensions can be used as pharmaceutically acceptable carriers
too. The liposomes can be prepared according to established methods known in
the art (U.S. Patent No. 4,522,811).
[00184] The antibodies or antibody compositions of the invention are
administered in therapeutically effective amounts as described. Therapeutically
effective amounts may vary with the subject's age, condition, sex, and seventy of
medical condition. Appropriate dosage may be determined by a physician based
on clinical indications. The antibodies or compositions may be given as a bolus
dose to maximize the circulating levels of antibodies for the greatest length of time.
Continuous infusion may also be used after the bolus dose.
[00185] As used herein, the term "subject" is intended to include human
and non-human animals. Subjects may include a human patient having a disorder
characterized by cells that express IL-22, e.g., a cancer cell or an immune cell.
The term "non-human animals" of the invention includes all vertebrates, such as
non-human primates, sheep, dogs, cows, chickens, amphibians, reptiles, etc.
[00186] Examples of dosage ranges that can be administered to a
subject can be chosen from: 1 µg/kg to 20 mg/kg, 1 µglkg to 10 mg/kg, 1 µg/kg to
1 mg/kg, 10 µg/kg to 1 mg/kg, 10 µg/kg to 100 µg/kg, 100 µg/kg to 1 mg/kg, 250
µg/kg to 2 mg/kg, 250 µg/kg to 1 mg/kg, 500 µg/kg to 2 mg/kg, 500 µg/kg to 1
mg/kg, 1 mg/kg to 2 mg/kg. 1 mg/kg to 5 mg/kg, 5 mg/kg to 10 mg/kg, 10 mg/kg to
20 mg/kg, 15 mg/kg to 20 mg/kg , 10 mg/kg to 25 mg/kg, 15 mg/kg to 25 mg/kg, 20
mg/kg to 25 mg/kg, and 20 mg/kg to 30 mg/kg (or higher). These dosages may be
administered daily, weekly, biweekly, monthly, or less frequently, for example,

biannually, depending on dosage, method of administration, disorder or
symptom(s) to be treated, and individual subject characteristics. Dosages can also
be administered via continuous infusion (such as through a pump). The
administered dose may also depend on the route of administration. For example,
subcutaneous administration may require a higher dosage than intravenous
administration.
[00187] In certain circumstances it may be advantageous to formulate
compositions in dosage unit form for ease of administration and uniformity of
dosage. Dosage unit form as used herein refers to physically discrete units suited
for the patient. Each dosage unit contains a predetermined quantity of antibody
calculated to produce a therapeutic effect in association with the carrier. The
dosage unit depends on the characteristics of the antibodies and the particular
therapeutic effect to be achieved.
[00188] Toxicity and therapeutic efficacy of the composition can be
determined by standard pharmaceutical procedures in cell cultures or experimental
animals, e.g., determining the LD50 (the dose lethal to 50% of the population) and
the ED50 (the dose therapeutically effective in 50% of the population). The dose
ratio between toxic and therapeutic effects is the therapeutic index and it can be
expressed as the ratio LD50/ED50. Antibodies that exhibit large therapeutic indices
may be less toxic and/or more therapeutically effective.
[00189] The data obtained from the cell culture assays and animal
studies can be used to formulate a dosage range in humans. The dosage of these
compounds may lie within the range of circulating antibody concentrations in the
blood, that includes an ED50 with little or no toxicity. The dosage may vary within
this range depending upon the dosage composition form employed and the route of
administration. For any antibody used in the present invention, the therapeutically
effective dose can be estimated initially using cell culture assays. A dose may be
formulated in animal models to achieve a circulating plasma concentration range
that includes the IC50 (i.e., the concentration of antibody which achieves a
half-maximal inhibition of symptoms). The effects of any particular dosage can be
monitored by a suitable bioassay. Examples of suitable bioassays include DNA
replication assays, transcription-based assays, IL-22/IL-22R binding assays, IL-

22/IL-10R2 binding assays, IL-22/IL-22R/IL-10R2, and other immunological
assays.
EXAMPLES
Example 1: Selection of Anti-IL-22 scFv's
Selection of Parents GIL01 and GIL68
[00190] GIL01 and GIL68 were isolated from scFv libraries by soluble
selection on IL-22. Soluble selections were carried out using biotinylated IL-22 with
an N-terminal His/FLAG tagged protein (bio.lL-22 H/F). Bio.lL-22 H/F was initially
used at a concentration of 100 nM. An scFv phagemid library, which is an
expanded version of the 1.38x1010 library described (Vaughan et al., 1996), was .
used to select antibodies specific for IL-22. Purified scFv phage (1012 transducing
units (tu)) were blocked for 30 minutes in 100 µl 3% MPBS (3% milk powder in
PBS), then bio.lL-22 H/F was added and incubated at room temperature for 1 hour.
Phage/antigen was added to 50 µl of Dynal M280 Streptavidin magnetic beads,
which had been blocked for 1 hour at 37oC in 1 ml of 3% MPBS, then incubated for
a further 15 minutes at room temperature. Beads were captured using a magnetic
rack and washed 4x in 1 ml of 3% MPBS/ 0.1% (v/v) Tween 20 followed by three
washes in PBS. After the last wash, beads were resuspended in 100 µl PBS and
used to infect 5 ml exponentially growing E. coli TG-1 cells. Cells and phage on
beads were incubated for 1 hour at 3720 (30 minutes stationary, 30 minutes
shaking at 250 rpm), then spread on 2TYAG plates. Plates were incubated at 30oC
overnight and colonies visualized the next day. Colonies were scraped off the
plates into 10 ml 2TY broth and 15% glycerol added for storage at-70oC.
[00191] Glycerol stock cultures from the first round panning selection
were superinfected with helper phage and rescued to give scFv antibody-
expressing phage particles for the second round of selection. A second and third
round of soluble selection was carried out as described above, dropping the
concentration of bio.lL-22 H/F to 50 nM.
Isolation of parents GIL16, GIL45, GIL60 and GIL92
[00192] GIL16, GIL45, GIL60 and GIL92 were isolated from scFv
libraries by a combination of panning on an IL-22 fusion protein and soluble
selection on bio.lL-22 H/F. Wells of a microtiter plate were coated with 10 µg/ml
(Dulbecco's PBS, pH 7.4) human IL-22 fusion protein and incubated overnight at

4oC. Wells were washed in PBS and blocked for 2 hours at 37aC in 3% MPBS.
Purified phage (1012 tu) in 100 µl of 3% MPBS were added to blocked wells and
incubated at room temperature for 1 hour. Wells were washed 10 times with PBST
(PBS containing 0.1% v/v Tween20), then 10 times with PBS. Bound phage
particles were eluted with 100 µl trypsin solution (0.5 µg/ml trypsin in 50 mM Tris
pH 8,1 mM CaCI2) for 30 minutes at 37oC. The eluted phage were used to infect
10 ml exponentially growing E. coli TG1. Infected cells were grown in 2TY broth for
1 hour at 37oC, as above, then streaked onto 2TYAG plates and incubated
overnight at 30oC. Output colonies were scraped off the plates and phage rescued
as described above. A second round of soluble selection was carried out as
described above, using 100 nM bio.lL-22 H/F.
Example 2: ScFv Blocks Binding of IL-22 to IL-22R
[00193] Inhibition assays were performed on the parent antibodies
GIL01, GIL16, GIL45, GIL60, GIL68, and GIL92 to identify antibodies that block or
alter binding of IL-22 to 1L-22R and/or IL-22 receptor complex. Crude scFv
containing periplasmic extracts were screened for the ability to inhibit the binding of
bio.lL-22 H/F to a human IL-22 receptor protein (hlL-22R). Output colonies from
selections were picked into 96 well plates containing 100 µl 2TYAG. ScFv
production was induced by addition of 1 mM IPTG to exponentially growing
cultures and overnight incubation at 30°C. Periplasmic extracts were prepared
(Griffiths et al., 1993) in 50 mM MOPS pH 7.4/ 0.5 mM EDTA / 0.5M Sorbitol.
[00194] Microtiter plates were coated with 1.25 µg/ml of an IL-22
receptor protein antibody (in PBS) for 1.5 hours at room temperature. Plates were
then washed three times in PBS, and blocked for 1 hour at room temperature with
PBS containing 2% milk powder (2% MPBS). After a further 3 washes, 50 µl of
25% cell conditioned medium containing an IL-22 receptor protein was added to
each well, and incubated overnight at 4°C. The following day, 25 µl of sample and
25µl of bio.IL-22 H/F (54 ng/ml in PBS/0.05% BSA/0.05% Tween) were added to
the washed plates, and incubated for 1.5 hours at room temperature. After 3
washes in PBST, binding of bio.lL-22 H/F was detected with Europium-Streptavidin
and TRF detected with the DELFIA® reagent kit and Victor 2™ Plate Reader
(Perkin Elmer).

[00195] Clones that showed inhibition of IL-22 binding were retested as
purified scFv. Both the JL-22/IL-22R binding assay (described above) and the IL-
22 / IL-22 receptor complex assay (described below) were used. ScFv
concentrations were titrated in order to establish the clone potencies as measured
by assay IC50 values. These were determined using GraphPad Prism software and
four-parameter logistic equation curve fitting. Sample results from the IL-22
receptor complex assay are shown in Figure 1.
Example 3: Verification of IL-22 binding by phage ELISA
[00196] To establish the specificity of the scFv's for IL-22, a phage
ELISA was performed using IL-22 fusion protein, IL-22 H/F and an unrelated
protein. Individual E. coli colonies containing phagemid were inoculated into 96
well plates containing 100 µl 2TYAG medium per well. M13K07 helper phage were
added to a multiplicity of infection (moi) of 10 to the exponentially growing culture
and the plates incubated an additional 1 hour at 37oC. Plates were centrifuged in a
benchtop centrifuge at 2000 rpm for 10 minutes. The supernatant was removed
and cell pellets were resuspended in 100 µl 2TYAK and incubated at 30oC
overnight with shaking. The next day, plates were centrifuged at 2000 rpm for 10
minutes and 100 µl phage-containing supernatant from each well was transferred
to a fresh 96 well plate. Phage samples were blocked in a final concentration of
3% MPBS for 1 hour at room temperature.
[00197] Microtiter plates were coated with 1 µg/ml IL-22 fusion protein,
IL-22 H/F or an unrelated protein and incubated overnight at 4oC. After coating,
the solutions were removed from the wells, and the plates blocked for 1 hour at
room temperature in 3% MPBS. Plates were rinsed with PBS then 50 µl of pre-
blocked phage was added to each well. The plates were incubated at room
temperature for 1 hour, then washed with 3 changes of PBST followed by 3
changes of PBS. To each well, 50 µl of a 1:5000 dilution of anti-M13-HRP
conjugate (Pharmacia) was added and the plates incubated at room temperature
for 1 hour. Plates were washed 3 times with PBST then 3 times with PBS. Fifty µl
of TMB substrate was added to each well and incubated until color development.
The reaction was stopped by the addition of 25 µl of 0.5 M H2SO4, and the
absorbance at 450 nm measured. These experiments confirmed the specific
binding of scFv clones to IL-22.

Example 4: Conversion of scFv to IgG
[00198] Heavy and light chain V regions from scFv clones were
amplified with clone-specific primers. PCR products were digested with
appropriate restriction enzymes and subcloned into vectors containing human lgG4
heavy chain constant domain (for VH domains) or vectors containing human
lambda or kappa light chain constant domains as appropriate (VL domains). The
closest human germlines of the VH and VL segments were determined and this
information was used to indicate whether kappa or lambda light chain constant
domains were used (Table 4). Correct insertion of V region domains into plasmids
was verified by sequencing of plasmid DNA from individual E. coli colonies.
Plasmids were prepared from E. coli cultures by standard techniques and heavy
and light chain constructs co-transfected into HEK 293 EBNA cells using standard
techniques. Secreted IgG was purified using protein A sepharose (Pharmacia) and
buffer exchanged into PBS.

[00199] Potency of purified IgG was verified in the biochemical IL-22
receptor complex inhibition assay as described below. IgG concentrations were

titrated in order to obtain potency values. Sample potency data is shown in Table
5.

Example 5: IL-22 Antibody Optimization
[00200] Large ribosome display libraries were created and screened for
scFv's that specifically recognized recombinant human IL-22, essentially as
described in Hanes et al. (2000). Initially the five parent clones (GIL01, GIL16,
GIL60, GIL68 and GIL92) were converted to ribosome display format, and this
template was subsequently used for library creation. On the DNA level, a T7
promoter was added at the 5'-end for efficient transcription to mRNA. On the
mRNA level, the construct contained a prokaryotic ribosome-binding site (Shine-
Dalgarno sequence) and 5' & 3' stem loops for mRNA stability. At the 3' end of the
single chain, the stop codon was removed and a portion of gIll was added to act as
a spacer, allowing folding of the scFv away from the ribosomal tunnel (Hanes et al.
2000).
[00201] Ribosome display libraries derived from the lead clones were
created by mutagenesis of the single chain complementarity determining regions
(CDRs) using PCR with non-proofreading Taq DNA polymerase. Affinity based
selections were performed where the library was incubated with bio.lL-22H/F,
followed by streptavidin coated paramagnetic beads (Dynal M280). Tertiary
complexes (mRNA-ribosome-scFv) were recovered by magnetic separation, while
unbound complexes were washed away. The mRNAs encoding the bound scFvs

were then rescued by RT-PCR as described (Hanes et al., 2000) and the selection
process repeated with decreasing concentrations (100 nM -10 pM over 5 rounds)
of bio.lL-22H/F.
[00202] Error prone PCR was introduced to further increase library size.
The error rate that was employed created, on average, 7.2 mutations per 1,000 bp
after a standard PCR reaction based on the method of Cadweil and Joyce (1992).
Initial error prone PCR reactions took place between the first and second rounds of
selection.
[00203] VH/VL recombination libraries for each parent clone were
prepared from the VH and VLCDR ribosome display outputs after either the second
or fourth round of selections. The VH portion of the VHCDR selection output for a
particular lineage was specifically PCR amplified, using clone specific primers. The
VL portion of the VLCDR selection output for the same lineage was amplified
separately. These two PCR products were recombined via an overlapping PCR
reaction. This created a complete library of scFv products containing all
components required for further rounds of ribosome display selection.
[00204] For some clones, phage display libraries were also utilized.
Phage libraries were created by mutagenesis of single chain CDRs using PCR
reactions with appropriate primers, and selected as described above. These
outputs were also combined with ribosome display selection outputs to create
VH/VL recombination libraries. The VH selection outputs from the fourth round of
ribosome display, together with the outputs from the third round of phage display,
were recombined with the VL outputs from the same lineage. This was achieved
using clone specific primers and over-lapping PCR to produce VH/VL recombination
libraries. Selections with soluble bio.IL.-22 H/F continued in ribosome display
format, as described above. The scFv regions of selection outputs were
directionally cloned into pCANTABS for production of scFv for biochemical high
throughput screening.
Example 6: Identification of optimized clones
[00205] Two assays were used for high throughput screening of
selection outputs. Outputs derived from clones GIL01, GIL16 and GIL68 were
screened in a homogeneous time resolved fluorescence assay (HTRF®, Cis

Biointernational), while G1L60 and GIL92 outputs were screened in a DELFIA®
(Perkin Elmer) assay.
HTRF® epitope competition assay
[00206] Crude scFv containing culture supernatants from GIL01, GIL16
and GIL68 output clones were prepared as described above and screened for
inhibition of bio.lL-22H/F binding GIL68 in an HTRF assay.
[00207] Cryptate labeled GIL68 IgG (labeling kit from Cis
Biointernational) was diluted 400 fold in assay buffer (PBS/0.4M KF/0.05%
BSA/0.05% Tween), followed by the addition of 7.5 nM Streptavidin XL665 (Xlent,
Cis Biointernational). This solution was mixed with crude scFv sample (diluted
125x), and bio.!L-22H/F in a Packard black 384 well Optiplate (Perkin Elmer).
Plates were incubated for 1 hour at room temperature then read using a Victor 2™
Plate Reader (Perkin Elmer). The 665 nM/620 nM emission ratio was used to
calculate the percentage of specific binding in each well.
DELFIA® Time Resolved Fluorescence assay
[00208J GIL60 and GIL92 output clones were screened for inhibition of
bio.ll_-22H/F binding to an IL-22 receptor complex.
[00209] Microtiter plates were coated with an IL-22 receptor complex
antibody (1 µg/ml in PBS), and incubated for 1.5 hours at room temperature. Plates
were washed three times in PBST, and blocked for 1 hour at room temperature
with 2% MPBS. After a further 3 washes, diluted cell conditioned medium
containing an IL-22 receptor complex was added and incubated overnight at 4°C.
Crude scFv supernatants were prepared as described above. The following day,
25 µl of diluted scFv sample and 25 µI of bio.IL-22 H/F (6 ng/ml) were added to the
washed plates, and incubated for 1.5 hours at room temperature. Plates were
washed 3 times in PBST, then binding of bio.lL-22H/F to the IL-22 receptor
complex was detected with Europium-Streptavidin and the DELFIA® reagent kit
(PerkinElmer). Time Resolved Fluorescence was measured using a Victor 2™
Plate Reader (Perkin Elmer).
[00210] Purified scFv from positive clones identified from the screening
were tested in the DELFIA® IL-22 receptor complex competition assay as
described above. A titration of scFv concentrations was used in order to establish
the clone potency as measured by IC50 values in the assay. Sample results are

shown in Figure 2. Fourteen optimized clones were designated 097D09, 062A09,
062G05, 087B03, 367D04, 368D04,166B06,166G05, 375G06, 376B10, 354A08,
355B06, 355E04, and 356A11.
Example 7: Ranking of Optimized Clones in the BAF3-IL-22 Proliferation
Assay
[00211] Proliferation assays were performed to assess the antibody's
ability to block the IL-22 mediated BaF3 cell proliferation. BaF3 cells expressing
hlL22R/hlL10R2 were generated by co-transfection of BaF3 cells with hlL22R-GFP
and hlL10R2-YFP. BaF3 cells expressing both ML22R and hlL10R2 (BaF3-IL-22
receptor cells) were sorted and collected by FACS.
[00212] BaF3-IL-22 receptor cells were routinely maintained in
RPM11640 with 10% FBS and 1 ng/mL murine IL-3. Immediately before assay
setup, cells were washed 4 times in assay medium (RPMI1640 with 10% FBS,
100U/ml Penicillin and 100ng/ml Streptomycin), resuspended in assay medium and
incubated at 37°C, 5% CO2 for 6-8 hours. To prepare assay plates, 100 µl of cells
(1x105/ml in assay medium) were added to the central 60 wells of a 96 well flat-
bottomed tissue culture plate (Costar). Test scFv or IgG samples were prepared
by diluting the stock sample In assay medium followed by filtration through a 0.22
µM filter. Serial 5-fold dilutions of samples were prepared in a separate dilution
plate. Cell containing wells were treated with 50 µl of sample followed by 50 µl of
human IL-22, (40 ng/ml in assay medium), and were then incubated for 40 hours at
37°C in 5% CO2. Control wells included media alone and cells either alone or in the
presence of 10 ng/mL human IL-22.
[00213] Cell proliferation was detected by the addition of 20 µl of Alamar
Blue (Serotec) to wells, followed by incubation for 5 hours ± 30 mins at 37°C in 5%
CO2. Plates were mixed by gentie tapping to ensure even signal throughout the
wells before measurement of fluorescence (excitation=560 nM, emission=590 nM).
EC50 and IC50 values were estimated using four-parameter logistic curve fitting
(Graphpad Prism 2 Software) and were used to rank antibodies. Sample potency
data for optimized scFvs and IgGs are shown in Table 6.

Table 6. IC50 values of scFv and IgG clones in BaF3-1L-22 proliferation assay

Clone Parent IC50 of scFv
(pM) IC50 of IgG
(PM)
097D09 GIL01 298±246 197±42
062A09 GIL16 267 83±37
062G05 GIL16 182 112±30
037B03 GIL60 212 105±17
367D04 GIL60 160±49 126±6
368D04 GIL60 186±66 127±10
166B06 GIL68 460 71 ±23
166G05 GIL68 204 97±23
375G06 GIL68 118±98 100±1
376B10 GIL68 104±47 119±6
354A08 GIL92 219±83 79±15*
355B06 GIL92 183±3 92±14*
355E04 GIL92 192±47 100±14*
356A11 GIL92 124*21 53±5*
*GIL92-derived clones were tested as germlined IgGs.
Example 8: Germlining
[00214] Sequence data for the six parent clones was used to identify the
nearest germline sequence for the heavy and light chain of each clone. Appropriate
mutations were made using standard site directed mutagenesis techniques with the
appropriate mutagenic primers. Mutation of sequences was confirmed by
sequence analysis. The sequences for the germlined clones and their scFv and VH
and VL domains are shown in Table 7. Purified scFv from the germlined parent
clones were tested in the biotinylated IL-22 binding IL-22 receptor complex
competition assay as described earlier, in order to establish the clone potency as
measured by IC50 values in the assay. Results are summarized in Table 8.

[00215] Nine of the optimized antibodies were germlined as described
above. Eight germlined IgGs were tested in the BaF3-IL-22 proliferation assay as
described above. Antibody IC50 values from a representative experiment are
showninTable 9.
[00216] Antibody sequences were then sent to GENEART North
America (28 Kirk Bradden Rd. East, Toronto, ON, Canada M8Y2E6), where they
were synthesized for optimized expression in CHO cells using GENEART's
proprietary optimization algorithm.


*sample contained precipitate ND= not determined
Example 9: Antibody inhibits IL-22 induced GROa secretion from HT29 cells
[00217] GROa assays were performed to assess the antibody's ability to
block the IL-22 induced GROa secretion from HT29 cells. HT29 cells were seeded
in 96 well flat bottom tissue culture plate (Coming Inc. Cat. #3595) in DMEM
medium (DMEM + 10% FBS + 100 unit/ml penicillin and streptomycin + 2 mM
Glutamine) at 5 x 104/well. 10 ng/ml IL-22 was mixed with serially diluted antibody
in DMEM medium and incubated for 30 min at 37°C. 24 hours after seeding,
medium was removed from HT29 cells and pre-mixed IL-22 and antibody were
added to the cells in 96 well plate.
[00218] After 48 hours of incubation at 37°C with 5% CO2, medium was
collected and secreted GROa was tested using Human GROa Immunoassay kit
(R&D Systems, Cat. DGR00), according to the manufacturer's directions. Results
are presented in Figure 3.
Example 10: Antibody binds to and inhibits different species IL-22
[00219] Cross species reactivity of germlined and non-germlined
optimized antibodies were determined as follows: ELISA plates (Costar, Cat.
#3590) were coated overnight with 1 µg/ml of rat, mouse, or human IL-22 or human
1L-26 in PBS buffer. Plates were washed with PBST buffer (0.05% Tween20 in

P8S) 3 times, then blocked with 1% BSA (Sigma A8918) / PBST for 1 hr at RT.
Antibodies were added at 1 µg/ml, incubated 1 hr at 25oC. The plates were
washed, then HRP-conjugated goat anti-human IgG antibody (Southern Biotech
Association, Cat. #2040-05) was added. The plates were incubated for 1 hour at
25oC, then washed with PBST, and developed with TMB (KPL, Cat. #50-76-04).
Reaction was stopped with 0.18 M H2SO4, Plates were read at OD 450 nm.
Results are presented in Figure 4.
[00220] These antibodies were also evaluated in both the GROa cell
assay and BaF3-IL-22 proliferation assay. As shown in Tables 10(a) and 10(b), the
antibodies blocked the activity of human, monkey, rat, and mouse lL-22 signalling
via a human IL-22 receptor. 356A11 and 368D04 also demonstrated cross-species
reactivity against murine, rat, and monkey IL-22 using real-time biospecific
interaction analysis (BIA), as discussed further in Example 11.
[00221] Table 10(a). IL-22 antibodies are highly potent for blocking
other species of IL-22 as shown in the GROa cell based assay system. Values
shown represent IC50 values in pM.

Protein ID human
IL-22 murine
IL-22 rat IL-22 monkey
IL-22
356A11 123.64 143.76 210.91 89.57
368D04 154.07 156.25 281.12 184.10
control 1 353.18 468.34 1161.57 343.19
control 2 1955.80 3399.79 10697.17 1459.27

[00222] Table 10(b). IL-22 antibodies are highly potent for blocking
other species of IL-22 as shown in the BaF3 cell based assay system. Values
shown represent IC50 values in pM.

Protein ID human
IL-22 murine
IL-22 rat IL-
22 monkey
IL-22
356A11 3.57 2.53 10.69 2.58
368D04 3.63 1.47 12.07 3.87
control 1 6.40 5-6 27.37 7.18
control 2 204.98 1033.26 2500.00 134.27
Example 11: Comparison of Binding Kinetics Between Rat Anti-IL-22
Monoclonal Antibodies and Human Anti-IL-22 Monoclonal
Antibodies
[00223] The binding kinetics of human, monoclonal anti-IL-22 antibodies
(356A11 and 368D04) and rat, monoclonal anti-IL-22 antibodies (P3/3 (Ab-02) and
P3/2 (Ab-04) from WO 2005/000897 and WO 02/068476) to human IL-22 were
evaluated by real-time biospecific interaction analysis (BIA) using surface plasmon
resonance technology.
[00224] To prepare the biosensor surface for the rat monoclonal
antibodies, Protein A/G (Pierce #21186, Rockford, 1L) was immobilized onto a
research-grade carboxymethyl dextran chip (CM5) using amine coupling. The
surface was activated with EDC/NHS. The protein A/G was injected at a
concentration of 50 µg/ml in sodium acetate buffer (pH 4.0). The immobilization
was done using the wizard tools with aim of 3000 (RUs) for the protein A/G.
Remaining activated groups were blocked with 1.0 M ethanolamine (pH 8.0). The
first flow cell was used as a reference surface to correct for bulk refractive index,
matrix effects, and non-specific binding. The second, third, and fourth flow cells
were coated with the capturing molecule. The rat monoclonal antibodies Ab-02
and Ab-04, which bind to protein A/G, were captured onto the protein A/G surface
by injecting 30 µl of a 1 µg/ml solution. The net difference between the baseline
and the point approximately 90 seconds after completing Ab-02 or Ab-04 injection
was used to represent the amount of ligand bound.

[00225] To prepare the biosensor surface for the human monoclonal
antibodies, either human monoclonal antibody (356A11 or 368D04) or control
antibody PD-1 (#17) were immobilized onto a research-grade carboxymethyl
dextran chip (CM5) using standard amine coupling. The surface was activated with
EDC/NHS. The capturing antibodies were injected at a concentration of 1 µg/ml in
sodium acetate buffer (pH 5.5). Remaining activated groups were blocked with 1.0
M ethanolamine (pH 8.0). The first flow cell was used as a reference surface to
correct for bulk refractive index, matrix effects, and non-specific binding. The
second, third, and fourth flow cells were coated with the capturing molecule.
[00226] For Ab-02 and Ab-04, solutions of human IL-22 at 300,100, 50,
25, 12.5, 6.4, 3.2, 1.6 and 0 nM concentrations were injected in triplicates at a flow
rate of 30 ul per minute for 3 minutes and the amount of bound material as a
function of time was recorded as sensorgrams. The dissociation phase was
monitored in HBS/EP buffer for 10 minutes at the same flow rate followed by a 5 µl
injection of 0.1 % TFA and a 5 µl injection of glycine pH 1.5 to regenerate a fully
active capturing surface.
[00227] For 356A11 and 368D04, solutions of human IL-22 at 400, 200,
100, 50, 25, 12.5, 6.25 and 0 nM were injected in triplicates at a flow rate of 100 µl
per minute (high flow to avoid non specific binding) for 3 minutes, and the amount
of bound material as a function of time was recorded as sensorgrams. The
dissociation phase was monitored in HBS/EP buffer for 60 minutes at the same
flow rate followed by two 5 µl injections of glycine pH 1.5 to regenerate a fully
active capturing surface.
[00228] All kinetic experiments were done at 22.5oC in HBS/EP buffer.
Blank and buffer effects were subtracted for each sensorgram using double
referencing. In control experiments the first injection contained buffer.
[00229] The kinetic data were analyzed using BIAevaluation software
3.0.2 applied to a 1:1 model. The apparent dissociation (Kd) and association (Ka)
rate constants were calculated from the appropriate regions of the sensorgrams
using a global analysis. The affinity constants of the interaction between antibody
and analyte were calculated from the kinetic rate constants by the following
formulae: KD = Kd / Ka, where KD is the dissociation constant and KA = Ka/Kd, where
KA is the association constant. The binding data for Ab-02 and AB-04 are

summarized in Tables 11A and 11B. The binding data for 356A11 and 368D04 are
summarized in Table 12.

[00230] These results show that the human monoclonal anti-IL-22
antibodies of this invention have a significantly higher affinity for human IL-22 than
the rat monoclonal anti-IL-22 antibodies Ab-02 and Ab-04, described in WO
2005/000897 and WO 02/068476 as having the ability to neutralize human IL-22.
Specifically, the dissociation constant of 356A11 (KD = 5.40 x 10-11 M or 0.054 nM)
for human IL-22 is approximately 1000-fold and more than 40-fold greater than the

dissociation constants of Ab-02 (KD = 5.22 x 10-8 M or 52 nM) and Ab-04 (KD = 2.38
x 10-9 M or 2.38 nM), respectively. Similarly, 368D04 (KD = 1.32 x 10-10 M or 0.132
nM) has an approximately 400-fold and 18-fold stronger affinity for human IL-22
than Ab-02 and Ab-04, respectively. The binding profiles of 356A11 and 36804 for
monkey, murine, and rat IL-22 were similar to that of human IL-22 (data not
shown).
[00231] The binding specificities of 356A11 and 368D04 were also
evaluated using BIA. Neither antibody showed cross reactivity with human IL-10,
human IL-19, human IL-20, human IL-24, human IL-28A, human IL-29, human IFN-
a2c, or human IFN-ω (data not shown).
Example 12: Model for Treatment of Arthritis
[00232] Arthritis is a disease characterized by inflammation in the joints.
Rheumatoid Arthritis (RA) is the most frequent form of arthritis, involving
inflammation of connective tissue and the synovial membrane, a membrane that
lines the joint. The inflamed synovial membrane often infiltrates the joint and
damages joint cartilage and bone. Both IL-22 and IL-22R protein and/or transcript
are associated with human disease. In RA synovial biopsies, IL-22 protein is
detected in vimentin+ synovial fibroblasts and some CD68+ macrophages while IL-
22R is detected in synovial fibroblasts. Treatment of synovial fibroblasts with IL-22
induces the production of monocyte chemoattractant protein-1, MCP-1, as well as
general metabolic activity (Ikeuchi, H. et al. (2005) Arthritis Rheum. 52:1037-46).
[00233] IL-22 is used to study its effect on cells from the synovial
membrane, the membrane that lines the joints. Human fibroblast-like synoviocytes
(HFLS) (Cell Applications (San Diego, CA)) are isolated from synovial tissues of
rheumatoid arthritis patients undergoing joint surgery. HFLS are cultured with
human IL-22 for 48 hours, and the supernatants are removed and tested for
chemokines and cytokines by ELISA. IL-22 will increase HFLS secretion of
chemokines MCP-1, Eotaxin, and IP-10, and cytokines TNFα, IL-6, and IL-8.
These chemokines and cytokines are known in the art to promote inflammation
through a number of activities, and increased concentrations in the joints caused
by IL-22 exacerbates inflammation and RA.
[00234] IL-22 is used to regulate the clinical progression of CIA
(Collagen Induced Arthritis). CIA is the standard mouse and rat model for studying

rheumatoid arthritis, see e.g., Holmdahl et al., (2002) Ageing Res. Rev., 1:135. On
day 0, mice are injected with 100 µg of Collagen Type II in complete Freund's
adjuvant, and on day 21, the mice are boosted with 100 µg of Collagen Type II in
incomplete Freund's adjuvant. On day 21, the mice are also injected daily with 1
µg of IL-22, and each day, the mice are examined for disease. The clinical signs
are scored as follows: 0 = no swelling, 1 = 1 to 2 swollen digits or swollen ankle, 2
= more than 2 swollen digits or mild paw swelling, 3 = extensive paw swelling, and
4 = ankylosis of paw. Mice injected with PBS after the collagen injections
progressively develop disease. Mice that are injected with IL-22 after the collagen
injections progressively develop more severe disease. Because treatment with IL-
22 specifically exacerbates CIA, treatment with anti-IL-22 antibodies, for example
with germlined 087B03, 368D04, 354A08 or 356A11, is expected to suppress or
delay CIA. Thus, since this model predicts treatment efficacy for RA, treatment
with anti-IL-22 antibodies, including germlined or non-germlined 087B03, 368D04,
354A08 or 356A11, is expected to suppress or delay RA in humans.
Example 13: Treatment of Patients
[00235] Patients with an autoimmune disorder, respiratory disorder,
inflammatory condition of the skin, cardiovascular system, nervous system,
kidneys, liver and pancreas or transplant patients are among the types of patients
that may be treated with the antibodies of the invention. Exemplary treatment
regimens and expected outcomes of antibodies according to this invention,
including 087B03, 368D04, 354A08, and 356A11, are provided below. Dosages
and frequencies of administration other than those in Table 13 may also be used.
The skilled artisan can adjust treatment regimens as necessary based on route of
administration or other known variables, such as the age, weight, condition, sex,
severity of medical condition, etc. of the patient to be treated.


[00236] The specification is most thoroughly understood in light of the
teachings of the references cited within the specification. The embodiments within
the specification provide an illustration of embodiments of the invention and should
not be construed to limit the scope of the invention. The skilled artisan readily
recognizes that many other embodiments are encompassed by the invention. All
publications and patents cited in this disclosure are incorporated by reference in

their entirety. To the extent the material incorporated by reference contradicts or is
inconsistent with this specification, the specification will supercede any such
material. The citation of any references herein is not an admission that such
references are prior art to the present invention.
[00237] Unless otherwise indicated, all numbers expressing quantities of
ingredients, reaction conditions, and so forth used in the specification, including
claims, are to be understood as being modified in all instances by the term "about."
Accordingly, unless otherwise indicated to the contrary, the numerical parameters
are approximations and may vary depending upon the desired properties sought to
be obtained by the present invention. At the very least, and not as an attempt to
limit the application of the doctrine of equivalents to the scope of the claims, each
numerical parameter should be construed in light of the number of significant digits
and ordinary rounding approaches.
[00238] Unless otherwise indicated, the term "at least" preceding a
series of elements is to be understood to refer to every element in the series.
Those skilled in the art will recognize, or be able to ascertain using no more than
routine experimentation, many equivalents to the specific embodiments of the
invention described herein. Such equivalents are intended to be encompassed by
the following claims.

CLAIMS
We claim:
1. An isolated antibody, or antigen-binding fragment thereof, that
specifically binds to IL-22, wherein the antibody, or antigen-binding fragment thereof,
comprises an amino acid sequence which is at least 95% identical to the amino acid
sequence shown in SEQ ID NO:5, 6, 7, 23, 24, 25,41, 42, 43, 59, 60, 61, 77, 78, 79,
95, 96, 97, 113,114, 115, 131, 132, 133, 149, 150, 151, 167, 168, 169, 185, 186,
187, 203, 204, 205, 221, 222, 223, 239, 240, 241, 257, 258, 259, 275, 276, 277, 293,
294, 295, 311, 312, 313, 329, 330, 331, 347, 348, 349, 365, 366, 367, 383, 384, 385,
401, 402, 403, 419, 420, 421, 437, 438, 439, 455, 456, 457, 473, 474, 475, 491, 492,
493, 509, 510, 511, 527, 528, 529, 545, 546, 547, 563, 564, 565, 581, 582, 583, 599,
600, 601, 617, 618, or 619.
2. An isolated antibody, or ant/gen-binding fragment thereof, that
specifically binds to IL-22, wherein the antibody, or antigen-binding fragment thereof,
comprises an amino acid sequence encoded by a nucleotide sequence which is at
least 95% identical to the nucleotide sequence shown in SEQ ID NO: 14, 15,16, 32,
33, 34, 50, 51, 52, 68, 69, 70, 86, 87, 88, 104, 105, 106, 122, 123, 124,140, 141,
142, 158, 159, 160, 176, 177, 178, 194, 195, 196, 212, 213, 214, 230, 231, 232, 248,
249, 250, 266, 267, 268, 284, 285, 286, 302, 303, 304, 320, 321, 322, 338, 339, 340,
356, 357, 358, 374, 375, 376, 392, 393, 394, 410, 411, 412, 428, 429, 430, 446, 447,
448, 464, 465, 466, 482, 483, 484, 500, 501, 502, 518, 519, 520, 536, 537, 538, 554,
555, 556, 572, 573, 574, 590, 591, 592, 608, 609, 610, 626, 627, or 628.
3. The antibody of claim 1, comprising the amino acid sequence as set
out in SEQ ID NO:5, 6, 7, 23, 24, 25, 41, 42, 43, 59, 60, 61, 77, 78, 79, 95, 96, 97,
113, 114, 115, 131, 132, 133, 149, 150, 151, 167, 168, 169, 185, 186, 187, 203, 204,
205, 221, 222, 223, 239, 240, 241, 257, 258, 259, 275, 276, 277, 293, 294, 295, 311,
312, 313, 329, 330, 331, 347, 348, 349, 365, 366, 367, 383, 384, 385, 401, 402, 403,
419, 420, 421, 437, 438, 439, 455, 456, 457, 473, 474, 475, 491, 492, 493, 509, 510,
511, 527, 528, 529, 545, 546, 547, 563, 564, 565, 581, 582, 583, 599, 600, 601, 617,
618, or 619.

4. An isolated antibody, or antigen-binding fragment thereof, that
specifically binds to IL-22, wherein the antibody, or antigen-binding fragment thereof,
comprises a VLdomain having a light chain variable region and a VH domain having
a heavy chain variable region, wherein the heavy chain variable region comprises
one or more of SEQ ID NO:8, 9, 10, 26, 27, 28, 44, 45, 46, 62, 63, 64, 80, 81, 82, 98,
99, 100, 116, 117, 118, 134, 135,136, 152, 153, 154, 170, 171, 172, 188, 189, 190,
206, 207, 208, 224, 225, 226, 242, 243, 244, 260, 261, 262, 278, 279, 280, 296, 297,
298, 314, 315, 316, 332, 333, 334, 350, 351, 352, 368, 369, 370, 386, 387, 388, 404,
405, 406, 422, 423, 424, 440, 441, 442, 458, 459, 460, 476, 477, 478, 494, 495, 496,
512, 513, 514, 530, 531, 532, 548, 549, 550, 566, 567, 568, 584, 585, 586, 602, 603,
604, 620, 621, or 622.
5. The antibody of claim 4, wherein the light chain variable region
comprises one or more of SEQ ID NO:11,12, 13, 29, 30, 31, 47, 48, 49, 65, 66, 67,
83, 84, 85, 101, 102, 103, 119, 120, 121, 137, 138, 139, 155, 156, 157, 173, 174,
175, 191, 192, 193, 209, 210, 211, 227, 228, 229, 245, 246, 247, 263, 264, 265, 281,
282, 283, 299, 300, 301, 317, 318, 319, 335, 336, 337, 353, 354, 355, 371, 372, 373,
389, 390, 391, 407, 408, 409, 425, 426, 427, 443, 444, 445, 461, 462, 463, 479, 480,
481, 497, 498, 499, 515, 516, 517, 533, 534, 535, 551, 552, 553, 569, 570, 571, 587,
588, 589, 605, 606, 607, 623, 624, and 625.
6. The antibody of claim 5, wherein the VH domain comprises the amino
acid sequence of any one of SEQ ID NO:5,23, 41, 59, 77, 95, 113, 131, 149, 167,
185, 203, 221, 239, 257, 275, 293, 311, 329, 347, 365, 383, 401, 419, 437, 455, 473,
491, 509, 527, 545, 563, 581, 599, or 617 and the VL domain comprises the amino
acid sequence of any one of SEQ ID NO:6, 24, 42, 60, 78, 96, 114, 132,150, 168,
186, 204, 222, 240, 258, 276, 294, 312, 330, 348, 366, 384, 402, 420, 438, 456, 474,
492, 510, 528, 546, 564, 582, 600, or 618.
7. The antibody of claim 6, wherein:
a) the VH domain comprises the amino acid sequence set out in SEQ ID
NO:167 or 491; and

b) the VL domain comprises the amino acid sequence set out in SEQ ID
NO:168 or 492.
8. The antibody of claim 6, wherein:
a) the VH domain comprises the amino acid sequence set out in SEQ ID
NO:293 or 545; and
b) the VL domain comprises the amino acid sequence set out in SEQ ID
NO:294 or 546.
9. The antibody of claim 6, wherein:
a) the VH domain comprises the amino acid sequence set out in SEQ ID
NO:203 or 617; and
b) the VL domain comprises the amino acid sequence set out in SEQ ID
NO:204 or 618.
10. The antibody of claim 6, wherein:
a) the VH domain comprises the amino acid sequence set out in SEQ ID
NO:347 or 599; and
b) the VL domain comprises the amino acid sequence set out in SEQ ID
NO:348 or 600.
11. The antibody of claim 4, wherein the heavy chain variable region
comprises:
a) SEQ ID NO:170 or 494,
b) SEQ ID NO:171 or 495; and
c) SEQ ID NO:172 or 496.
12. The antibody of claim 4, wherein the heavy chain variable region
comprises:
a) SEQ ID NO:296 or 548,
b) SEQ ID NO:297 or 549; and
c) SEQ ID NO:298 or 550.

13. The antibody of claim 4, wherein the heavy chain variable region
comprises:
a) SEQ ID NO:206 or 620,
b) SEQ ID NO:207 or 621; and
c) SEQ ID NO:208 or 622.
14. The antibody of claim 4, wherein the heavy chain variable region
comprises:
a) SEQ ID NO:350 or 602,
b) SEQ ID NO:351 or 603; and
c) SEQ ID NO:352 or 604.
15. The antibody of claim 11, wherein the light chain variable region
comprises:
a) SEQ ID NO: 173 or 497,
b) SEQ ID NO: 174 or 498; and
c) SEQ ID NO:175 or 499.
16. The antibody of claim 12, wherein the light chain variable region
comprises:
a) SEQ ID NO:299 or 551,
b) SEQ ID NO:300 or 552; and
c) SEQ ID NO:301 or 553.
17. The antibody of claim 13, wherein the light chain variable region
comprises:
a) SEQ ID NO:209 or 623,
b) SEQ ID NO:210 or 624; and
c) SEQ ID NO:211 or 625.
18. The antibody of claim 14, wherein the light chain variable region
comprises:

a) SEQ ID NO:353 or 605,
b) SEQ ID NO:354 or 606; and
c) SEQ ID NO:355 or 607.

19. An isolated antibody, or antigen-binding fragment thereof, that
specifically binds to an IL-22 epitope that is recognized by GIL01, GIL16, GIL45,
GIL60, GIL68, GIL92, 097D09, 062A09, 062G05, 087B03, 367D04, 368D04,
166B06, 166G05, 375G06, 376B10, 354A08, 355B06, 355E04, or356A11, such that
the antibody competitively inhibits the binding of GIL01, GIL16, GIL45, GIL60, GIL68,
GIL92, 097D09, 062A09, 062G05, 087B03, 367D04, 368D04, 166B06, 166G05,
375G06, 376B10, 354A08, 355B06, 355E04, or 356A11 to human IL-22.
20. The antibody of claim 19, comprising a VH domain, a VL domain, or a Fv
fragment as set out in SEQ ID NO:5, 6, 7, 23, 24, 25, 41,42, 43, 59,60, 61, 77, 78,
79;l 95,96, 97, 113,114, 115, 131, 132, 133, 149, 150, 151, 167, 168, 169, 185, 186,
187, 203, 204, 205, 221, 222, 223, 239, 240, 241, 257, 258, 259, 275, 276, 277, 293,
294, 295, 311, 312, 313, 329, 330, 331, 347, 348, 349, 365, 366, 367, 383, 384, 385,
401, 402, 403, 419, 420, 421, 437, 438, 439, 455, 456,457, 473, 474, 475, 491,492,
493, 509, 510, 511, 527, 528, 529, 545, 546, 547,563, 564, 565, 581, 582, 583, 599,
600, 601, 617, 618, or 619.
21. The antibody of claim 20, wherein the VH domain comprises SEQ ID
NO:167 or 491.
22. The antibody of claim 21, wherein the VL domain comprises SEQ ID
NO: 168 or 492.
23. The antibody of claim 20, wherein the VH domain comprises SEQ ID
NO:293 or 545.
24. The antibody of claim 23, wherein the VL domain comprises SEQ ID
NO:294 or 546.

25. The antibody of claim 20, wherein the VH domain comprises SEQ ID
NO:203 or 617.
26. The antibody of claim 25, wherein the VL domain comprises SEQ ID
NO:204 or 618.
27. The antibody of claim 20, wherein the VH domain comprises SEQ ID
NO:347 or 599.
28. The antibody of claim 27, wherein the VL domain comprises SEQ ID
NO:348 or 600.
29. The antibody of claim 19 that specifically binds to an IL-22 epitope that
is recognized by 087B03, such that the antibody competitively inhibits the binding of
087B03 to human IL-22.
30. The antibody of claim 19 that specifically binds to an IL-22 epitope that
is recognized by 354A08, such that the antibody competitively inhibits the binding of
354A08 to human IL-22.
31. The antibody of claim 19 that specifically binds to an IL-22 epitope that
is recognized by 368D04, such that the antibody competitively inhibits the binding of
368D04 to human IL-22.
32. The antibody of claim 19 that specifically binds to an IL-22 epitope that
is recognized by 356A11, such that the antibody competitively inhibits the binding of
356A11 to human IL-22.
33. The antibody of claim 22, wherein the antibody is 087B03.
34. The antibody of claim 24, wherein the antibody is 354A08.
35. The antibody of claim 26, wherein the antibody is 368D04.

36. The antibody of claim 28, wherein the antibody is 356A11.
37. The antibody of claim 1 or 2, wherein the association constant of the
antibody for human IL-22 is at least 1010 M-1.
38. The antibody of claim 1 or 2, wherein the antibody blocks IL-22
mediated proliferation of BaF3 cells with an IC50 of 150 pM or less and wherein the
BaF3 cells comprise a human IL-22 receptor.
39. The antibody of claim 1 or 2, wherein the antibody blocks IL-22
mediated GROa secretion from HT29 cells with an IC50 of 150 pM or less.
40. An isolated antibody, or antigen-binding fragment thereof, that
specifically binds to IL-22, wherein the association constant of the antibody for
human IL-22 is at least 1010 M-1.
41. An isolated antibody, or antigen-binding fragment thereof, wherein the
antibody blocks IL-22 mediated proliferation of BaF3 cells with an IC50 of 150 pM or
less and wherein the BaF3 cells comprise a human IL-22 receptor.
42. The antibody of claim 1 or 2, wherein the antibody specifically binds to
an amino acid sequence that is at least 95% identical to any sequence of at least
100 contiguous amino acids in the sequence set forth SEQ ID NO:1.
43. The antibody of claim 1 or 2, wherein the antibody inhibits the binding
of IL-22 to IL-22R or a receptor complex comprising IL-22R and IL-10R2.
44. The antibody of claim 1 or 2, wherein the antibody is human.
45. The antibody of claim 1 or 2, wherein the antibody is lgG1 or lgG4.
46. A pharmaceutical composition comprising the antibody of claim 1 or 2.

47. An isolated nucleic acid encoding the antibody of claim 1 or 2.
48. An expression vector comprising the nucleic acid of claim 47.
49. A host cell transformed with the vector of claim 48.
50. The host cell of claim 49, wherein the host cell is a bacteria,
mammalian cell, yeast cell, plant cell, or an insect cell.
51. A method of producing an antibody that binds to IL-22, comprising
culturing the host cell of claim 50 under conditions that allow expression of the
antibody, and isolating the antibody from the cell culture.
52. The nucleic acid of claim 47, wherein the nucleic acid encodes a
protein comprising an amino acid sequence set out in one of SEQ ID NO.-5-13, 23-
31, 41-49, 59-67, 77-85, 95-103, 113-121, 131-139, 149-157, 167-175,185-193,
203-211, 221-229, 239-247, 257-265, 275-283, 293-301, 311-319, 329-337, 347-
355, 365-373, 383-391, 401-409, 419-427, 437-445, 455-463, 473-481, 491-499,
509-517, 527-535, 545-553, 563-571, 581-589, 599-607, or 617-625.
53. The nucleic acid of claim 52, wherein the nucleic acid comprises a
nucleotide sequence of one of SEQ ID NO:14-22, 32-40, 50-58, 68-76, 86-94, 104-
112, 122-130, 140-148, 158-166, 176-184, 194-202, 212-220, 230-238, 248-256,
266-274, 284-292, 302-310, 320-328, 338-346, 356-364, 374-382, 392-400, 410-
418, 428-436, 446-454, 464-472, 482-490, 500-508, 518-526, 536-544, 554-562,
572-580, 590-598, 608-616, or 626-634.
54. The nucleic acid of claim 52, wherein the nucleic acid encodes a
protein comprising an amino acid sequence set out in SEQ ID NO: 167, 168, 169,
170, 171, 172, 173, 174, 175, 491, 492, 493, 494, 495, 496, 497, 498, or 499.

55. The nucleic acid of claim 52, wherein the nucleic acid encodes a
protein comprising an amino acid sequence set out in SEQ ID NO:293, 294, 295,
296, 297, 298, 299, 300, 301, 545, 546, 547, 548, 549, 550, 551, 552, or 553.
56. The nucleic acid of claim 52, wherein the nucleic acid encodes a
protein comprising an amino acid sequence set out in SEQ ID NO:203, 204, 205,
206, 207, 208, 209, 210, 211, 617, 618, 619, 620, 621, 622, 623, 624, or 625.
57. The nucleic acid of claim 52, wherein the nucleic acid encodes a
protein comprising an amino acid sequence set out in SEQ ID NO:347, 348, 349,
350, 351, 352, 353, 354, 355, 599, 600, 601, 602, 603, 604, 605, 606, or 607.
58. The nucleic acid of claim 53, wherein the the nucleic acid comprises a
nucleotide sequence of SEQ ID NO:176,177,178, 179,180,181,182, 183, 184,
500, 501, 502, 503, 504, 505, 506, 507, or 508.
59. The nucleic acid of claim 53, wherein the the nucleic acid comprises a
nucleotide sequence of SEQ ID NO:302, 303, 304, 305, 306, 307, 308, 309, 310,
554, 555, 556, 557, 558, 559, 560, 561, or 562.
60. The nucleic acid of claim 53, wherein the the nucleic acid comprises a
nucleotide sequence of SEQ ID NO:212, 213, 214, 215, 216, 217, 218, 219, 220,
626, 627, 628, 629, 630, 631, 632, 633, or 634.
61. The nucleic acid of claim 53, wherein the the nucleic acid comprises a
nucleotide sequence of SEQ ID NO:356, 357, 358, 359, 360, 361, 362, 363, 364,
608, 609, 610, 611, 612, 613, 614, 615, or 616.
62. A diagnostic kit comprising the antibody of claim 1 or 2.

The present application provides human antibodies and antigen binding fragments thereof that specifically bind to the human interleukin-22 (IL-22). The antibodies can act as antagonists of IL-22 activity, thereby modulating immune responses in general, and those mediated by IL-22 in particular. The disclosed compositions and methods may be used for example, in diagnosing, treating or preventing inflammatory disorders, autoimmune diseases, allergies, septic shock, infectious disorders, transplant rejection, cancer, and other immune system disorders.