NZ623425B2 - Anti il-36r antibodies - Google Patents

Abstract

Disclosed are antibodies for anti Interleukin (IL) 36R. Including therapeutic and diagnostic methods.

Description

WO 74569 Anti IL-36R Antibodies Sequence Listing The instant application contains a Sequence Listing which has been ted in ASCII format via EFS-Web and is hereby orated by reference in its entirety. Said ASCII copy, created on November 12, 2012, is named 09-0583WO.txt and is 147,390 bytes in size.

Technical Field of the Invention This invention generally relates to anti-IL-36R antibodies for diagnostic and therapeutic use. The antibodies can be used in pharmaceutical compositions and kits sing such compounds. The antibodies are useful in s for the treatment of various diseases or disorders, for example immunological, inflammatory, autoimmune, fibrotic and respiratory diseases in humans.

Background of the Invention The IL-1 family of cytokines is composed of 11 different ligands, , IL-1or (also termed IL-1F1), IL-1B (IL-1F2), IL-1 receptor antagonist (IL-1Ra or IL-1F3), IL-18 (IL- 1F4), IL-1F5 to IL-1F10, and IL-1F11 (or IL-33). IL-1or and IL-1B are known to induce pro-inflammatory activities on g to type I IL-1 receptor (IL-1Rl) and recruitment of the common co-receptor IL-1 receptor accessory protein (IL-1RAcP), whereas IL-1Ra acts as a competitive inhibitor of IL-1 binding to IL-1Rl, thus ng nflammatory activity. Numerous studies reported that IL-18 is a pro-inflammatory cytokine that is an inducer of IFN-v, whereas IL-33 was described as an immunoregulatory cytokine ed in particular in the control of Th2 ses. New members of the IL-1 family, including IL-1F5, IL-1F6, IL-1F8, and IL-1F9, were identified through searches in DNA databases for homologs of IL-1. In humans and mice, all the genes encoding these cytokines map to less than 300 kb of chromosome 2q, where they are flanked by the IL1A, IL1B, and IL1RN genes. IL-1 F6, IL-1 F8, and IL-1 F9 share 21% to 37% amino acid sequence homology with lL-1 and lL-1Ra, whereas lL-1F5 displays 52% amino acid sequence homology with lL-1Ra, suggesting that lL-1F5 might ent an endogenous receptor antagonist. lL-1 F6, lL-1F8, and lL-1F9 bind to lL-1Rrp2, a receptor of the lL-1 R family, and use lL- 1RAcP as a co-receptor to stimulate intracellular signals r to those induced by IL- 1, whereas lL-1F5 was shown to inhibit lL-1F9—induced NF-KB activation in Jurkat T cells that over-express lL-1Rrp2. Like lL-1B, all these lL-1 homologs lack a leader peptide and cannot be released through the conventional secretory pathway, although studies suggest that release of lL-1Rrp2 agonists may be controlled by mechanisms ent from those regulating lL-1B secretion. To acknowledge the specific biologic effects of these cytokines and to recognize that they all bind to the same receptor, it has recently been proposed to amend the nomenclature of lL-1 homologs. Thus, lL-1Rrp2 is now termed lL-36R and its ligands are named lL-36d (IL-1 F6), lL-3BB 8), and IL- 36y (lL-1F9). In addition, lL-1F5, which has been shown to exert receptor antagonist activities, has been renamed lL-36Ra.

Messenger RNAs for lL-36d, lL-3BB, and lL-36y are highly expressed in several tissues, particularly in internal epithelial s, which are exposed to pathogens and in skin.

Interestingly, expression of lL-36Ra and lL-36d is significantly up-regulated in IL- 1B/TNF-d—stimulated human nocytes, and lL-36Ra and lL-36y mRNA are highly increased in lesional psoriasis skin. Moreover, lL-36y n production is enhanced in human keratinocytes after TNF-d and lFN-y stimulation. Elevated lL-36d mRNA and protein sion was reported also in chronic kidney disease.

Transgenic mice overexpressing lL-36d in nocytes exhibit inflammatory skin lesions sharing some features with psoriasis. This phenotype was more severe when transgenic mice were crossed with lL-36Ra—deficient mice, ting a regulatory function of lL-36Ra in vivo. The inflammatory skin condition in keratinocyte-specific lL- 36d transgenic is even more similar to human psoriasis if the mice are treated with 12- O-tetradecanoylphorbol tate, resembling the human disease histologically, molecularly, and in its se to therapeutics. Moreover, human psoriatic lesional skin transplanted onto immunodeficient mice is normalized when the mice are treated with anti–IL-36R dy, arguing that the IL-36 axis is required to maintain the lesional phenotype in human psoriatic skin. Taken together, these data indicate that IL-36R s, including IL- 36α, IL-36β, and , exert proinflammatory effects in vitro and in vivo and that IL-36Ra acts as a natural antagonist, thus mimicking the IL-1/IL-1Ra system.

There is therefore ce that IL-36R ligands are involved in a number of disease conditions, and there is a need for new therapeutic agents targeting this y, in particular for use in the treatment of inflammatory diseases.

Summary of the invention The present invention addresses the above need by providing biotherapeutics, in particular antibodies, which bind to IL-36R. In one aspect, the antibodies of the present invention block IL36 ligand-mediated signaling (α, β and/or γ). In one aspect the antibodies of the present invention are useful, for example for the treatment of epithelial-mediated inflammation/fibrosis in diseases such as sis, inflammatory bowel disease, scleroderma, COPD, and chronic kidney disease.

According to a first aspect, the present invention provides an anti-interleukin-36R (anti-IL-36R) antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 35, 102, 103, 104, 105, 106 or 140 (L-CDR2); the amino acid ce of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 2); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In one aspect, the present invention provides an anti-IL-36R antibody having one or more of the properties below.

In one aspect, an anti-IL-36R antibody of the t invention has high molecular/cellular binding potency. In one aspect, an anti-IL-36R antibody of the present invention binds to human IL-36R at a KD < 0.1 nM. In a further aspect, an anti-IL-36R antibody of the present invention, in particular a humanized anti-IL-36R antibody, binds to human IL-36R at a KD < 50 pM. In one aspect, an anti-IL-36R dy of the present invention binds to IL-36R expressing cells at an EC90 <5 nM.

In another , an anti-IL-36R antibody of the present invention has high cell-based functional ng potency. In one aspect, an anti-IL-36R dy of the present AH26(10651560_1):RTK 2012/064933 invention blocks all three lL-36R agonistic ligands (or, B, y) at an I090 55 nM, in disease- relevant cell lines and primary cells.

In one aspect, an anti-lL-36R antibody of the present invention has the molecular/cellular binding potency and the cell-based functional blocking potency set forth above.

In a further aspect, an anti-lL-36R antibody of the present invention has high selectivity for example greater than 1000-fold selectivity against human lL-1R1 or lL-36R negative cell lines. In a further aspect, an anti-lL-36R antibody of the present ion does not bind to human lL-1 R1 or lL-36R negative cell lines.

In embodiment one, the present invention provides an anti-lL-36R antibody or antigen- binding fragment thereof, which binds to human lL-36R at a KD equal to or< 0.1 nM. ln embodiment two, the present invention provides an anti-lL-36R antibody or antigen- binding fragment thereof according to embodiment one, wherein the said antibody or antigen-binding fragment is a monoclonal dy or n-binding fragment f.

In embodiment three, the present invention es an anti-lL-36R antibody or antigen- binding fragment thereof according to embodiment one or two, wherein the said antibody or antigen-binding fragment is a zed antibody or antigen-binding fragment thereof.

In embodiment four, the present invention provides an anti-lL-36R antibody or antigen- binding fragment thereof ing to embodiment three, which binds to human lL-36R at a KD equal to or< 50 pM. ln embodiment five, the t invention provides an anti-lL-36R antibody or antigen- binding fragment f according to any one of ment one to four, which does not bind to human lL-1R1.

In embodiment six, the present invention provides an anti-IL-36R antibody or antigen- binding fragment thereof according to embodiment one, wherein the antibody or antigen-binding fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 35, 102, 103, 104, 105106 or 140 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In embodiment seven, the present invention provides an anti-IL-36R antibody or antigen-binding fragment thereof ing to ment six, wherein the antibody or antigen-binding fragment fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 102 (L-CDR2); the amino acid ce of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid ce of SEQ ID NO: 72 (H-CDR3).

In embodiment eight, the present invention provides an anti-IL-36R dy or n- binding fragment thereof according to embodiment six, wherein the antibody or antigenbinding fragment nt thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 103 (L-CDR2); the amino acid ce of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid ce of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In embodiment nine, the t invention provides an anti-IL-36R antibody or antigen- binding fragment thereof according to embodiment six, wherein the antibody or antigenbinding fragment fragment thereof comprises: a) a light chain le region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 104 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In embodiment ten, the present invention provides an anti-IL-36R antibody or antigen- binding fragment thereof ing to embodiment six, n the dy or antigen- binding fragment fragment thereof comprises: a) a light chain variable region sing the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 105 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid ce of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In embodiment eleven, the present invention provides an anti-IL-36R antibody or antigen-binding fragment f ing to embodiment six, wherein the antibody or antigen-binding fragment fragment thereof comprises: a) a light chain variable region sing the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 106 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In embodiment twelve, the present invention provides an anti-IL-36R antibody or antigen-binding fragment thereof according to embodiment six, wherein the antibody or antigen-binding fragment fragment thereof comprises: a) a light chain le region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 140 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In embodiment thirteen, the present invention provides an anti-IL-36R dy or antigen-binding fragment thereof according to embodiment one, wherein the antibody or antigen-binding fragment fragment thereof comprises alight chain variable region comprising the amino acid sequence of any one of SEQ ID NO: 76, 77, 78, 79, 80, 81, 82 or 83; and a heavy chain variable region comprising the amino acid sequence of any one of SEQ ID NO: 87, 88, 89, 90, 91, 92, 93, 94 or 95.

In embodiment fourteen, the present invention es an L-36R antibody or antigen-binding fragment thereof according to embodiment thirteen, wherein the antibody or antigen-binding fragment fragment thereof comprises alight chain le region comprising the amino acid sequence of SEQ ID NO: 77; and a heavy chain variable region sing the amino acid sequence of SEQ ID NO: 87;or alight chain variable region comprising the amino acid sequence of SEQ ID NO: 77; and a heavy chain le region sing the amino acid sequence of SEQ ID NO: 88; or alight chain variable region sing the amino acid sequence of SEQ ID NO: 77; and a heavy chain variable region sing the amino acid sequence of SEQ ID NO: In embodiment fifteen, the present invention provides an in the antibody or antigen- binding fragment fragment thereof comprises alight chain variable region comprising the amino acid sequence of SEQ ID NO: 80; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 87;or a light chain variable region comprising the amino acid ce of SEQ ID NO: 80; and a heavy chain variable region comprising the amino acid ce of SEQ ID NO: 88; or alight chain variable region comprising the amino acid sequence of SEQ ID NO: 80; and a heavy chain le region comprising the amino acid sequence of SEQ ID NO: In embodiment sixteen, the present invention es an anti-IL-36R antibody or antigen-binding fragment thereof ing to embodiment one, wherein the antibody or antigen-binding fragment fragment thereof comprises: a light chain variable region comprising the amino acid sequence of SEQ ID NO: 27 (L- CDRf); the amino acid sequence of SEQ ID NO: 36 (L-CDR2); the amino acid sequence of SEQ ID NO: 45 (L-CDR3); and a heavy chain variable region sing the amino acid sequence of SEQ ID NO: 107 (H-CDRt); the amino acid sequence of SEQ ID NO: 63 (H-CDR2); the amino acid sequence of SEQ ID NO: 73 (H-CDR3); or alight chain variable region comprising the amino acid sequence of SEQ ID NO: 27 (L- CDRf); the amino acid sequence of SEQ ID NO: 36 (L-CDR2); the amino acid sequence of SEQ ID NO: 45 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 107 (H-CDRt); the amino acid sequence of SEQ ID NO: 64 (H-CDR2); the amino acid sequence of SEQ ID NO: 73 (H-CDR3); or or alight chain le region sing the amino acid sequence of SEQ ID NO: 27 (L- CDRf); the amino acid sequence of SEQ ID NO: 36 (L-CDR2); the amino acid sequence of SEQ ID NO: 45 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 54 (H-CDRt); the amino acid sequence of SEQ ID NO: 63 or 64 (H-CDR2); the amino acid sequence of SEQ ID NO: 73 (H- CDR3).

In embodiment seventeen, the present ion provides an anti-IL-36R antibody or antigen-binding nt thereof according to embodiment one, wherein the antibody or antigen-binding fragment fragment thereof comprises alight chain variable region comprising the amino acid sequence of any one of SEQ ID NO: 84, 85 or 86; and a heavy chain variable region comprising the amino acid sequence of any one of SEQ ID NO: 96, 97, 98, 99, 100 or 101.

In embodiment eighteen, the present invention provides an anti-IL-36R antibody or antigen-binding fragment thereof according to embodinet seventeen, wherein the antibody or antigen-binding fragment fragment thereof ses alight chain variable region sing the amino acid sequence of SEQ ID NO: 85; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 100; or alight chain variable region comprising the amino acid sequence of SEQ ID NO: 85; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:101.

In embodiment nineteen, the t invention provides an anti-IL-36R antibody or antigen-binding nt thereof according to embodiment seventeen, wherein the antibody or antigen-binding fragment fragment thereof ses alight chain le region sing the amino acid sequence of SEQ ID NO: 86; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 100; or alight chain variable region comprising the amino acid sequence of SEQ ID NO: 86; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:101.

In embodiment twenty, the present invention es an anti-IL-36R antibody, wherein the antibody comprises alight chain comprising the amino acid sequence of any one of SEQ ID NO:114,115,116,117,118,119,12O or 121; and a heavy chain comprising the amino acid sequence of any one of SEQ ID NO: 125, 126, 127, 128, 129, 130, 131, 132 or 133.

In embodiment twenty one, the present invention provides an L-36R antibody according to embodiment twenty, wherein the antibody comprises a light chain sing the amino acid sequence of SEQ ID NO: 115; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 125.

In embodiment twenty two, the present invention provides an anti-IL-36R antibody according to embodiment twenty, n the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 115; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 126.

In embodiment twenty three, the present invention es an anti-IL-36R antibody according to embodiment twenty, wherein the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 115; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 127.

In embodiment twenty four, the present ion provides an L-36R antibody according to embodiment twenty, n the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 125.

In embodiment twenty five, the present invention provides an anti-IL-36R antibody according to embodiment twenty, wherein the antibody ses a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 126.

In embodiment twenty six, the present invention provides an anti-IL-36R antibody according to embodiment twenty, wherein the dy comprises a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 127.

In embodiment twenty seven, the present invention es an anti-IL-36R antibody, wherein the antibody comprises a light chain comprising the amino acid sequence of any one of SEQ ID NO: 122, 123 or 124; and a heavy chain comprising the amino acid sequence of any one of SEQ ID NO: 134, 135, 136, 137, 138 or 139.

In embodiment twenty eight, the t invention provides an anti-IL-36R antibody according to embodiment twenty seven, wherein the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 123; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 138.

In embodiment twenty nine, the present invention provides an anti-IL-36R antibody according to embodiment twenty seven, wherein the antibody comprises a light chain sing the amino acid sequence of SEQ ID NO: 123; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 139.

In embodiment thirty, the present invention provides an anti-IL-36R dy according to twenty seven, wherein the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 124; and a heavy chain comprising the amino acid ce of SEQ ID NO: 138.

In embodiment thirty one, the present invention provides an anti-IL-36R dy or antigen-binding fragment thereof according to embodiment one, wherein the antibody or antigen-binding nt fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid ce of SEQ ID NO: 103 2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62 (H-CDR2); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In embodiment thirty two, the present invention es an anti-IL-36R antibody or antigen-binding fragment thereof according to embodiment one, n the antibody or antigen-binding fragment fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDRt); the amino acid sequence of SEQ ID NO: 104(L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid ce of SEQ ID NO: 53 (H-CDRt); the amino acid sequence of SEQ ID NO: 62 (H-CDR2); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

In embodiment thirty three, the present invention provides an anti-IL-36R antibody or antigen-binding fragment f ing to embodiment one, wherein the antibody or n-binding nt fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 27 (L-CDRt); the amino acid sequence of SEQ ID NO: 36(L-CDR2); the amino acid sequence of SEQ ID NO: 45 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 107 t); the amino acid sequence of SEQ ID NO: 63 (H-CDR2); the amino acid sequence of SEQ ID NO: 73 (H-CDR3).

In embodiment thirty four, the present invention provides an anti-IL-36R antibody or antigen-binding fragment thereof according to embodiment one, wherein the antibody or antigen-binding nt fragment f comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 27 (L-CDRt); the amino acid sequence of SEQ ID NO: 36(L-CDR2); the amino acid sequence of SEQ ID NO: 45 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 107 (H-CDRt); the amino acid sequence of SEQ ID NO: 64 (H-CDR2); the amino acid sequence of SEQ ID NO: 73 (H-CDR3).

In one embodiment, an antibody or antigen-binding fragment thereof according to any one of embodiments one to thirty-four is a onal antibody. In one embodiment, an antibody or antigen-binding fragment thereof ing to any one of ments one to -four is a humanized antibody. In one embodiment, an antibody or antigen- binding fragment thereof according to any one of embodiments one to thirty-four is a monoclonal humanized antibody.

In embodiment thirty five, the present ion provides an anti-IL-36R antibody or antigen-binding fragment f, wherein the antibody or n-binding fragment fragment thereof comprises: a light chain variable region comprising the amino acid sequence of SEQ ID NO: 21 (L- CDR1); the amino acid sequence of SEQ ID NO: 30 (L-CDR2); the amino acid ce of SEQ ID NO: 39 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 48 (H-CDR1); the amino acid sequence of SEQ ID NO: 57 (H-CDR2); the amino acid sequence of SEQ ID NO: 67 (H-CDR3); or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 22 (L- CDR1); the amino acid ce of SEQ ID NO: 31 (L-CDR2); the amino acid sequence of SEQ ID NO: 40 (L-CDR3); and a heavy chain le region comprising the amino acid sequence of SEQ ID NO: 49 (H-CDR1); the amino acid sequence of SEQ ID NO: 58 (H-CDR2); the amino acid sequence of SEQ ID NO: 68 (H-CDR3); or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 23 (L- CDR1); the amino acid sequence of SEQ ID NO: 32 (L-CDR2); the amino acid sequence of SEQ ID NO: 41 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 50 (H-CDR1); the amino acid sequence of SEQ ID NO: 59 (H-CDR2); the amino acid sequence of SEQ ID NO: 69 (H-CDR3); or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 24 (L- CDR1); the amino acid sequence of SEQ ID NO: 33 (L-CDR2); the amino acid sequence of SEQ ID NO: 42 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 51 (H-CDR1); the amino acid sequence of SEQ ID NO: 60 (H-CDR2); the amino acid sequence of SEQ ID NO: 70 (H-CDR3); or 2012/064933 a light chain le region comprising the amino acid sequence of SEQ ID NO: 25 (L- CDRt); the amino acid sequence of SEQ ID NO: 34 (L-CDR2); the amino acid sequence of SEQ ID NO: 43 3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 52 (H-CDRt); the amino acid sequence of SEQ ID NO: 61 (H-CDR2); the amino acid sequence of SEQ ID NO: 71 3); or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L- CDRt); the amino acid sequence of SEQ ID NO: 35 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDRt); the amino acid sequence of SEQ ID NO: 62 (H-CDR2); the amino acid sequence of SEQ ID NO: 72 3); or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 27 (L- CDRt); the amino acid sequence of SEQ ID NO: 36 (L-CDR2); the amino acid sequence of SEQ ID NO: 45 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 54 (H-CDRt); the amino acid sequence of SEQ ID NO: 63 (H-CDR2); the amino acid sequence of SEQ ID NO: 73 (H-CDR3); or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 27 (L- CDRt); the amino acid sequence of SEQ ID NO: 36 (L-CDR2); the amino acid sequence of SEQ ID NO: 45 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 54 (H-CDRt); the amino acid sequence of SEQ ID NO: 64 (H-CDR2); the amino acid sequence of SEQ ID NO: 73 (H-CDR3); or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 28 (L- CDRt); the amino acid sequence of SEQ ID NO: 37 (L-CDR2); the amino acid ce of SEQ ID NO: 46 (L-CDR3); and a heavy chain le region comprising the amino acid sequence of SEQ ID NO: 55 t); the amino acid sequence of SEQ ID NO: 65 (H-CDR2); the amino acid sequence of SEQ ID NO: 74 (H-CDR3); or a light chain variable region comprising the amino acid ce of SEQ ID NO: 29 (L- CDRt ); the amino acid sequence of SEQ ID NO: 38 (L-CDR2); the amino acid sequence of SEQ ID NO: 47 (L-CDR3); and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 56 (H-CDRt); the amino acid sequence of SEQ ID NO: 66 (H-CDR2); the amino acid sequence of SEQ ID NO: 75 (H-CDR3).

In ment thirty six, the present invention provides an L-36R antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment nt thereof comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 1; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 11; or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 2; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 12; a light chain variable region comprising the amino acid sequence of SEQ ID NO: 3; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 13; a light chain variable region comprising the amino acid sequence of SEQ ID NO: 4; and a heavy chain variable region comprising the amino acid ce of SEQ ID NO: 14; a light chain variable region comprising the amino acid sequence of SEQ ID NO: 5; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 15; a light chain variable region sing the amino acid sequence of SEQ ID NO: 6; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 16; a light chain variable region comprising the amino acid sequence of SEQ ID NO: 7; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 17; a light chain variable region comprising the amino acid sequence of SEQ ID NO: 8; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 18; a light chain variable region comprising the amino acid ce of SEQ ID NO: 9; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 19; a light chain variable region comprising the amino acid sequence of SEQ ID NO: 10; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: In a further ment thirty seven, the present invention provides a pharmaceutical composition comprising an antibody or antigen-binding fragment according to any one of the previous ments and a pharmaceutically acceptable carrier.

In a further embodiment thirty eight the present invention provides an antibody or antigen-binding fragment or pharmaceutical composition according to any one of the previous embodiments, for use in ne.

In a further embodiment thirty nine the t invention provides an antibody or antigen-binding fragment or pharmaceutical composition according to any one of the embodiments 1-37, wherein the use is the treatment of an inflammatory disease, of an autoimmune disease, of a respiratory disease, of a metabolic disorder, of an lial mediated inflammatory disorder, fibrosis or of cancer.

In a further ment forty the present invention provides an antibody or antigen- binding fragment or pharmaceutical composition ing to any one of the embodiments 1-37, wherein the use is for the treatment of psoriasis, inflammatory bowel disease, psoriatic tis, multiple sclerosis, rheumatoid arthritis, COPD, chronic asthma or ankylosing litis.

In a further embodiment forty one, the present invention provides an antibody or antigen-binding fragment or pharmaceutical composition according to any one of the embodiments 1-37, wherein the use is for the treatment of inflammatory bowel disease.

In a still further embodiment forty two, the present invention es an antibody or n-binding fragment or pharmaceutical composition according to ment 41, wherein the the disease is Crohns disease.

In another embodiment forty three, the present invention provides a method of treating a disease comprising administering the antibody or n-binding fragment or pharmaceutical ition according to any one of the embodiments 1-37, to a patient in need thereof, wherein the disease is selected from an inflammatory e, an autoimmune disease, a respiratory disease, a metabolic disorder, an epithelial mediated inflammatory disorder, fibrosis and cancer.

In another embodiment forty four, the present invention provides a method according to embodiment 43 wherein the disease is selected from psoriasis, inflammatory bowel disease, psoriatic tis, multiple sclerosis, rheumatoid arthritis, COPD, chronic asthma and ankylosing litis.

In a still further embodiment forty five, the present invention provides a method for treating Crohns disease.

Further embodiments of the invention encompass: - An isolated polynucleotide comprising a ce encoding an anti-lL-36R antibody or antigen-binding fragment according to the invention, preferably a DNA or RNA sequence; - an isolated polynucleotide according to the invention, encoding a sequence as defined by one or more of SEQ ID NOs. 1 to 140; - a vector comprising a cleotide according to the invention, preferably an expression vector, more preferred a vector comprising the polynucleotide according to the invention in functional association with an expression control ce; - a host cell comprising a polynucleotide according to the invention and/or a vector according to the invention; - a method for the production of an anti-lL-36R dy or antigen-binding fragment according to the invention, preferably a inant production method comprising the use of a polynucleotide according to the invention, and/or of a vector according to the invention and/or of a host cell according to the invention; - such a method preferably comprises the steps (a) cultivating the host cell under conditions allowing the expression of the anti-lL-36R antibody or antigen-binding fragment and (b) recovering the anti-lL-36R antibody or antigen-binding fragment; - a diagnostic kit or diagnostic method comprising an anti-lL-36R antibody or antigen-binding fragment according to the invention, or the use thereof; - a Diagnostic kit or diagnostic method according the invention, for the diagnosis of an matory disease, an autoimmune e, a respiratory disease, a metabolic disorder, an epithelial mediated inflammatory disorder, fibrosis, cancer, psoriasis, inflammatory bowel disease, psoriatic arthritis, multiple sclerosis, toid arthritis, COPD, chronic , ankylosing litis, or Crohns disease.

Brief Description of the Figures Figure 1: lL-36 antagonist ligands (lL-36RA/IL1 F5, lL-38/ILF10) inhibit the signaling cascade.

Figure 2: Gene chip es demonstrate lL-36R ligands are upregulated in psoriatic skin (IL-36 RA, lL-36 or and lL-36 y).

Figure 3: sion profile using human skin sections.

Formalin-fixed paraffin embedded with antibody titrations using antibody 33D10 Figure 4: Method to Evaluate Epidermal thickness of human skin sections Description of the invention This invention s to anti-lL-36R antibodies. In one aspect, the antibodies of the t ion are for diagnostic and therapeutic use, for example in humans.

The present invention provides antibodies that bind to lL-36R, in particular human lL- 36R. The present invention also relates to humanized antibodies that bind lL-36R. ln specific ments, the sequence of these humanized antibodies has been identified based on the sequences of certain lead mouse antibodies.

Without wishing to be bound by this theory it is believed that anti-lL-36R antibodies or antigen-binding fragments thereof bind to human lL-36R and thus interfer with the binding of lL-36 agonists, and in doing so block at least partially the signaling cascade from the lL-36R to inflammatory mediators. This is illustrated by Figure 1.

In one aspect, the antibodies of the present invention are for use in models of human disease. lL-36R is also known as lL-1RL2 and lL-1Rrp2. It has been reported that agonistic lL-36 ligands (or, B, or y) initiate the signaling cascade by engaging the lL-36 receptor which then forms a heterodimer with the lL-1 receptor accessory n (lL- 1RAcP). lL-36 antagonist ligands (lL-36RA/IL1 F5, ILF10) t the ing cascade (see figure 1).

In one , the present ion provides an anti-lL-36R antibody having one or more of the properties below.

In one , an anti-lL-36R antibody of the present invention has high molecular/cellular binding potency. In one aspect, an anti-lL-36R antibody of the present invention binds to human lL-36R at a KD< 0.1 nM. In a further , an anti-lL-36R antibody of the present invention, in particular a humanized anti-lL-36R antibody, binds to human lL-36R at a KD < 50 pM. In one aspect, an anti-lL-36R antibody of the present invention binds to lL-36R expressing cells at an E090 <5 nM.

In another aspect, an anti-lL-36R antibody of the present invention has high cell-based functional blocking potency. In one aspect, an anti-lL-36R dy of the present ion blocks all three lL-36R agonistic ligands (or, B, y) at an |Cgo 55 nM, in disease- relevant cell lines and primary cells.

In one aspect, an anti-lL-36R antibody of the t invention has the molecular/cellular binding potency and the cell-based functional blocking potency set forth above.

In one aspect, an anti-lL-36R antibody of the present invention is a humanized antibody.

In one aspect, an anti-lL-36R antibody of the present invention is a monoclonal antibody. In one , an anti-lL-36R antibody of the t invention is a full length antibody. In one , an anti-lL-36R antibody of the present invention is a humanized monoclonal antibody, for e a full length humanized monoclonal dy.

An antibody or antigen-binding nt thereof of the present invention recognizes specific "IL-36R antigen epitope" or " lL-36R epitope". As used herein these terms refer to a molecule (e.g., a peptide) or a nt of a molecule capable of immunoreactivity with an anti-lL-36R antibody.

The epitopes are most commonly proteins, short oligopeptides, oligopeptide mimics (i.e., organic compounds that mimic antibody g properties of the lL-36R antigen), or combinations thereof. The minimum size of a peptide or polypeptide epitope for an antibody is thought to be about four to five amino acids. Peptide or polypeptide epitopes n for example at least seven amino acids or for example at least nine amino acids or for example between about 15 to about 20 amino acids. Since an antibody can recognize an antigenic peptide or polypeptide in its tertiary form, the amino acids comprising an epitope need not be contiguous, and in some cases, may not even be on the same peptide chain. Epitopes may be determined by various techniques known in the art, such as X-ray crystallography, Hydrogen/Deuterium Exchange Mass Spectrometry (HXMS), irected mutagenesis, alanine ng mutagenesis, and peptide screening methods.

The generalized structure of antibodies or immunoglobulin is well known to those of skill in the art. These les are heterotetrameric glycoproteins, typically of about 150,000 daltons, composed of two identical light (L) chains and two identical heavy (H) chains and are typically referred to as full length antibodies. Each light chain is covalently linked to a heavy chain by one disulfide bond to form a heterodimer, and the heterotrameric molecule is formed through a covalent disulfide linkage between the two identical heavy chains of the heterodimers. Although the light and heavy chains are linked together by one disulfide bond, the number of disulfide es between the two heavy chains varies by immunoglobulin isotype. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain has at the amino- terminus a variable domain (VH), followed by three or four constant domains (CH1, CH2, CH3, and CH4), as well as a hinge region between CH1 and CH2. Each light chain has two domains, an amino-terminal variable domain (VL) and a carboxy-terminal constant domain (CL). The VL domain associates non-covalently with the VH , whereas the CL domain is commonly covalently linked to the CH1 domain via a disulfide bond.

Particular amino acid es are believed to form an interface between the light and heavy chain variable domains (Chothia et al., 1985, J. Mol. Biol. 186:651-663). Variable domains are also referred herein as variable regions.

Certain domains within the variable domains differ ively between different antibodies i.e., are "hypervariable." These hypervariable domains contain residues that are ly involved in the binding and specificity of each particular dy for its specific nic determinant. Hypervariability, both in the light chain and the heavy chain variable s, is concentrated in three segments known as mentarity determining s (CDRs) or hypervariable loops (HVLs). CDRs are defined by sequence comparison in Kabat et al., 1991, In: Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of , da, Md., whereas HVLs (also referred herein as CDBs) are structurally defined according to the three-dimensional structure of the variable domain, as described by Chothia and Lesk, 1987, J. Mol. Biol. 196: 901-917. These two methods result in slightly different fications of a CDB. As defined by Kabat, CDR-L1 is positioned at about es 24-34, CDB-L2, at about residues 50-56, and CDR-L3, at about residues 89-97 in the light chain variable domain; CDB-H1 is positioned at about residues 31-35, CDR-H2 at about residues 50-65, and CDR-H3 at about residues 95-102 in the heavy chain variable domain. The exact residue numbers that encompass a particular CDR will vary depending on the sequence and size of the GDP. Those skilled in the art can routinely determine which residues comprise a particular CDR given the variable region amino acid sequence of the antibody. The CDR1, CDR2, CDR3 of the heavy and light chains therefore define the unique and functional properties specific for a given antibody.

The three CDRs within each of the heavy and light chains are separated by framework regions (FR), which n sequences that tend to be less variable. From the amino us to the carboxy terminus of the heavy and light chain variable domains, the FRs and CDRs are arranged in the order: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4.

The largely B-sheet uration of the FRs brings the CDRs within each of the chains into close proximity to each other as well as to the CDRs from the other chain. The resulting conformation contributes to the antigen binding site (see Kabat et al., 1991, NIH Publ. No. 91-3242, Vol. I, pages 647-669), although not all CDR residues are necessarily directly ed in antigen binding.

FR residues and lg constant domains are not directly involved in antigen g, but contribute to antigen binding and/or mediate antibody effector function. Some FR residues are thought to have a significant effect on antigen binding in at least three ways: by noncovalently binding directly to an epitope, by interacting with one or more CDR residues, and by affecting the interface n the heavy and light chains. The constant domains are not directly involved in antigen binding but mediate various lg or ons, such as ipation of the antibody in antibody dependent cellular cytotoxicity (ADCC), complement dependent cytotoxicity (CDC) and antibody dependent cellular phagocytosis (ADCP).

The light chains of vertebrate immunoglobulins are assigned to one of two clearly distinct s, kappa (K) and lambda (7»), based on the amino acid sequence of the constant domain. By comparison, the heavy chains of mammalian immunoglobulins are assigned to one of five major classes, according to the ce of the constant domains: lgA, lgD, lgE, lgG, and lgM. lgG and lgA are further divided into subclasses pes), e.g., lgG1, lng, lgG3, lgG4, lgA1, and lgAz. The heavy chain constant domains that correspond to the different classes of immunoglobulins are called or, 6, s, y, and u, respectively. The subunit structures and three-dimensional configurations of the classes of native immunoglobulins are well known.

WO 74569 The terms, ody", "anti-lL-36R antibody", "humanized anti-lL-36R dy", "humanized anti-lL-36R epitope antibody", and "variant humanized anti-lL-36R epitope antibody" specifically encompass monoclonal antibodies (including full length monoclonal antibodies), polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments such as variable domains and other portions of antibodies that exhibit a desired biological activity, e.g., lL-36R binding.The term "monoclonal antibody" (mAb) refers to an antibody that is highly specific, being directed against a single antigenic inant, an “epitope”. Therefore, the er "monoclonal" is tive of antibodies directed to the identical epitope and is not to be construed as requiring production of the antibody by any ular . It should be understood that monoclonal antibodies can be made by any technique or methodology known in the art; including e.g., the hybridoma method ( Kohler et al., 1975, Nature 256:495), or recombinant DNA methods known in the art (see, e.g., U.S. Pat. No. 4,816,567), or s of isolation of monoclonal inantly produced using phage dy libraries, using techniques described in on et al., 1991, Nature 352: 624- 628, and Marks et al., 1991, J. Mol. Biol. 222: 581-597.

The term “monomer” refers to a homogenous form of an antibody. For example, for a full-length antibody, monomer means a monomeric antibody having two identical heavy chains and two identical light chains.

Chimeric antibodies consist of the heavy and light chain variable regions of an dy from one species (e.g., a non-human mammal such as a mouse) and the heavy and light chain constant regions of another species (e.g., human) antibody and can be obtained by linking the DNA sequences encoding the variable regions of the antibody from the first species (e.g., mouse) to the DNA sequences for the constant regions of the antibody from the second (e.g. human) species and transforming a host with an expression vector ning the linked sequences to allow it to produce a chimeric antibody. Alternatively, the chimeric antibody also could be one in which one or more regions or domains of the heavy and/or light chain is identical with, homologous to, or a variant of the corresponding ce in a monoclonal antibody from another immunoglobulin class or isotype, or from a consensus or germline sequence. Chimeric antibodies can include fragments of such antibodies, provided that the antibody 2012/064933 fragment exhibits the desired biological activity of its parent dy, for example binding to the same epitope (see, e.g., U.S. Pat. No. 4,816,567; and Morrison et al., 1984, Proc. Natl. Acad. Sci. USA 81: 6851-6855).

The terms, "antibody fragment anti-lL-36R antibody fragment", "anti-lL-36R epitope antibody fragment", "humanized anti-lL-36R antibody fragment", "humanized anti-IL- 36R epitope antibody fragment", "variant humanized anti-lL-36R epitope dy fragment" refer to a portion of a full length anti-lL-36R antibody, in which a variable region or a functional capability is retained, for e, specific lL-36R epitope binding. es of antibody fragments include, but are not limited to, a Fab, Fab', F(ab')2, Fd, Fv, scFv and scFv-Fc fragment, a diabody, a linear antibody, a single-chain antibody, a minibody, a diabody formed from antibody fragments, and multispecific antibodies formed from antibody fragments.

Full length antibodies can be treated with enzymes such as papain or pepsin to generate useful dy fragments. Papain digestion is used to produces two cal antigen-binding dy nts called "Fab" fragments, each with a single antigen- binding site, and a residual "Fc" fragment. The Fab fragment also contains the constant domain of the light chain and the CH1 domain of the heavy chain. Pepsin treatment yields a F(ab')2 fragment that has two antigen-binding sites and is still capable of cross- linking antigen.

Fab' fragments differ from Fab fragments by the presence of additional residues including one or more cysteines from the antibody hinge region at the C-terminus of the CH1 domain. F(ab')2 dy fragments are pairs of Fab' fragments linked by cysteine residues in the hinge region. Other chemical couplings of antibody fragments are also known.

"Fv" fragment contains a complete antigen-recognition and g site consisting of a dimer of one heavy and one light chain variable domain in tight, non-covalent association. In this configuration, the three CDRs of each le domain interact to define an antigen-biding site on the surface of the VH-VL dimer. Collectively, the six CDRs confer antigen-binding specificity to the antibody.

A "single-chain Fv" or "scFv" antibody fragment is a single chain Fv variant comprising the VH and VL domains of an antibody where the domains are present in a single polypeptide chain. The single chain Fv is capable of recognizing and g antigen.

The scFv polypeptide may optionally also contain a polypeptide linker positioned n the VH and VL domains in order to facilitate formation of a desired three- dimensional structure for antigen binding by the scFv (see, e.g., hun, 1994, In The Pharmacology of monoclonal Antibodies, Vol. 113, Rosenburg and Moore eds., Springer-Verlag, New York, pp. 269-315).

A "diabody" refers to small antibody fragments with two antigen-binding sites, which nts se a heavy chain variable domain (V.sub.H) connected to a light chain variable domain (V.sub.L) in the same polypeptide chain (V.sub.H-V.sub.L or V.sub.L- V.sub.H). Diabodies are described more fully in, e.g., Holliger et al. (1993) Proc. Natl.

Acad. Sci. USA 90: 6444-6448.

Other recognized antibody fragments include those that comprise a pair of tandem Fd segments (VH-CH1-VH-CH1) to form a pair of antigen binding regions. These “linear antibodies” can be bispecific or monospecific as described in, for example, Zapata et al. 1995, Protein Eng. 1057-1062.

A “humanized antibody” or a “humanized antibody fragment” is a specific type of chimeric antibody which includes an immunoglobulin amino acid sequence variant, or fragment f, which is capable of binding to a predetermined n and which, comprises one or more FRs having substantially the amino acid sequence of a human immunoglobulin and one or more CDRs having substantially the amino acid sequence of a non-human immunoglobulin. This man amino acid sequence often referred to as an "import" ce is typically taken from an "import" antibody domain, particularly a variable domain. In general, a humanized antibody includes at least the CDRs or HVLs of a non-human antibody, inserted n the PBS of a human heavy or light chain variable domain. The t invention describes specific humanized anti- lL-36R antibodies which contain CDRs derived from the mouse monoclonal antibodies or humanized CDRs ed between the PBS of human ne sequence heavy and light chain variable domains. It will be understood that certain mouse FR residues may be important to the function of the zed antibodies and therefore certain of the human germline sequence heavy and light chain variable domains residues are modified to be the same as those of the corresponding mouse sequence.

In another aspect, a humanized anti-lL-36R antibody comprises substantially all of at least one, and typically two, variable domains (such as contained, for example, in Fab, Fab', F(ab')2, Fabc, and Fv fragments) in which all, or ntially all, of the CDRs correspond to those of a non-human immunoglobulin, and specifically herein, all of the CDRs are mouse or humanized sequences as detailed herein below and all, or substantially all, of the FRs are those of a human immunoglobulin consensus or germline sequence. In r aspect, a humanized anti- lL-36R antibody also es at least a n of an immunoglobulin Fc region, typically that of a human immunoglobulin. Ordinarily, the antibody will contain both the light chain as well as at least the variable domain of a heavy chain. The antibody also may include one or more of the CH1, hinge, CH2, CH3, and/or CH4 regions of the heavy chain, as appropriate.

A humanized anti-lL-36r antibody can be selected from any class of immunoglobulins, including lgM, lgG, lgD, lgA and lgE, and any isotype, ing lgG1, lng, lgG3, lgG4, lgA1 and lgA2. For example, the constant domain can be a complement fixing constant domain where it is desired that the humanized antibody exhibit cytotoxic activity, and the isotype is typically lgG1. Where such cytotoxic activity is not desirable, the nt domain may be of another isotype, e.g., lng. An alternative humanized anti-lL-36R antibody can comprise sequences from more than one immunoglobulin class or e, and selecting particular constant domains to optimize desired or functions is within the ry skill in the art. In specific embodiments, the present invention provides dies that are lth antibodies and more particularly, are lth antibodies in which there is a knock-out of effector functions.

The FRs and CDRs, or HVLs, of a humanized anti-lL-36R antibody need not correspond precisely to the parental sequences. For e, one or more residues in the import CDR, or HVL, or the consensus or germline FR sequence may be altered (e.g., mutagenized) by substitution, insertion or deletion such that the resulting amino acid residue is no longer identical to the original e in the corresponding position in either parental sequence but the antibody nevertheless retains the function of binding to IL-36R. Such alteration typically will not be extensive and will be conservative alterations. Usually, at least 75% of the humanized antibody residues will correspond to those of the parental consensus or germline FR and import CDR sequences, more often at least 90%, and most frequently greater than 95%, or greater than 98% or r than 99%. lmmunoglobulin residues that affect the interface between heavy and light chain variable regions ("the VL-VH interface") are those that affect the proximity or orientation of the two chains with respect to one another. Certain residues that may be involved in interchain interactions include VL residues 34, 36, 38, 44, 46, 87, 89, 91, 96, and 98 and VH es 35, 37, 39, 45, 47, 91, 93, 95, 100, and 103 (utilizing the numbering system set forth in Kabat et al., Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md., 1987)). U.S. Pat. No. 6,407,213 also discusses that es such as VL residues 43 and 85, and VH residues 43 and 60 also may be involved in this interaction. While these residues are indicated for human IgG only, they are applicable across species. Important antibody es that are reasonably ed to be ed in interchain interactions are selected for substitution into the consensus sequence.

The terms "consensus sequence" and "consensus antibody" refer to an amino acid sequence which comprises the most frequently occurring amino acid residue at each location in all immunoglobulins of any ular class, isotype, or subunit structure, e.g., a human immunoglobulin variable domain. The consensus sequence may be based on immunoglobulins of a particular species or of many species. A "consensus" sequence, structure, or antibody is understood to encompass a consensus human ce as described in certain embodiments, and to refer to an amino acid sequence which ses the most frequently occurring amino acid residues at each location in all human immunoglobulins of any particular class, isotype, or subunit structure. Thus, the sus sequence contains an amino acid sequence having at each position an amino acid that is present in one or more known immunoglobulins, but which may not exactly duplicate the entire amino acid sequence of any single immunoglobulin. The le region consensus ce is not obtained from any naturally produced antibody or globulin. Kabat et al., 1991, Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., and variants thereof.

Human germline sequences are found naturally in the human population. A combination of those germline genes generates antibody diversity. ne antibody sequences for the light chain of the antibody come from conserved human germline kappa or lambda v-genes and j-genes. Similarly the heavy chain sequences come from germline v-, d- and j-genes (LeFranc, M-P, and LeFranc, G, “The Immunoglobulin Facts Book” Academic Press, 2001).

As used herein, "variant", "anti- IL-36R variant", "humanized anti- IL-36R variant", or "variant humanized anti- IL-36R" each refers to a humanized anti-IL-36R dy having at least a light chain variable murine CDR. Variants include those having one or more amino acid changes in one or both light chain or heavy chain variable domains, provided that the amino acid change does not substantially impair binding of the antibody to IL-36R.

An "isolated" antibody is one that has been identified and separated and/or recovered from a component of its natural environment. Contaminant components of the antibody's natural environment are those materials that may interfere with diagnostic or therapeutic uses of the dy, and can be enzymes, hormones, or other proteinaceous or nonproteinaceous solutes. In one , the dy will be purified to at least greater than 95% isolation by weight of antibody.

An isolated dy includes an antibody in situ within recombinant cells in which it is produced, since at least one ent of the antibody's natural environment will not be present. Ordinarily r, an isolated antibody will be prepared by at least one purification step in which the recombinant cellular material is removed.

The term "antibody performance" refers to s that contribute to antibody recognition of n or the iveness of an antibody in vivo. Changes in the amino acid sequence of an antibody can affect antibody properties such as folding, and can influence physical factors such as initial rate of antibody binding to antigen (ka), dissociation constant of the antibody from n (kd), affinity constant of the antibody for the antigen (Kd), conformation of the antibody, protein stability, and half life of the antibody.

The term "epitope tagged" when used herein, refers to an anti-lL-36R antibody fused to an "epitope tag". An "epitope tag" is a polypeptide having a sufficient number of amino acids to provide an epitope for antibody production, yet is designed such that it does not interfere with the d activity of the zed anti-lL-36R antibody. The e tag is y sufficiently unique such that an antibody raised t the epitope tag does not substantially cross-react with other epitopes. le tag polypeptides generally contain at least 6 amino acid residues and usually contain about 8 to 50 amino acid residues, or about 9 to 30 residues. Examples of epitope tags and the antibody that binds the epitope include the flu HA tag polypeptide and its antibody 12CA5 (Field et al., 1988 Mol. Cell. Biol. 8: 2159-2165; c-myc tag and 8F9, 3C7, 6E10, G4, B7 and 9E10 antibodies thereto (Evan et al., 1985, Mol. Cell. Biol. 5(12):3610-3616; and Herpes simplex virus glycoprotein D (gD) tag and its antibody (Paborsky et al. 1990, Protein Engineering 3(6): 3). In certain embodiments, the epitope tag is a "salvage receptor binding e". As used herein, the term "salvage receptor binding epitope" refers to an epitope of the Fc region of an lgG molecule (such as lgG1, lng, lgGs, or lgG4) that is responsible for sing the in vivo serum half-life of the lgG molecule.

In some embodiments, the antibodies of the present invention may be ated to a cytotoxic agent. This is any substance that inhibits or prevents the function of cells and/or causes destruction of cells. The term is intended to include radioactive isotopes (such as l‘3‘, I125, Y90 and Re , 186) , chemotherapeutic agents, and toxins such as enzymatically active toxins of bacterial, fungal, plant, or animal origin, and fragments thereof. Such cytotoxic agents can be coupled to the zed antibodies of the t invention using standard procedures, and used, for example, to treat a patient indicated for therapy with the antibody.

A "chemotherapeutic agent" is a chemical compound useful in the treatment of cancer.

There are numerous examples of chemotherapeutic agents that could be conjugated with the therapeutic antibodies of the present invention. Examples of such chemotherapeutic agents include ting agents such a thiotepa and cyclosphosphamide; alkyl sulfonates such as busulfan, improsulfan, and piposulfan; aziridines such as benzodopa, carboquone, meturedopa, and uredopa; ethylenimines and methylamelamines including altretamine, triethylenemelamine, trietylenephosphoramide, triethylenethiophosphoramide, and trimethylolomelamine; acetogenins ially bullatacin and bullatacinone); camptothecin (including the synthetic analogue can); bryostatin; callystatin; CC-1065 (including its adozelesin, esin, and bizelesin synthetic analogues); cryptophycines (particularly cryptophycin 1 and phycin 8); dolastatin, auristatins, (including analogues monomethyl- auristatin E and monomethyl-auristatin F); duocarmycin (including the synthetic analogues, 9 and CBl-TMI); eleutherobin; pancratistatin; sarcodictyin; spongistatin; nitrogen mustards such as chlorambucil, chlomaphazine, cholophosphamide, ustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, phenesterine, prednimustine; trofosfamide, uracil mustard; ureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, stine; antibiotics such as the enediyne antibiotics (e.g., calicheamicin, especially calichemicin gamma1l and calicheamicin phil1, see for example, Agnew, Chem. lntl. Ed. Engl., 33:183-186; dynemicin, including dynemicin A; bisphosphonates, such as clodronate; esperamicin; as well as neocarzinostatin chromophore and d chromoprotein enediyne antibiotic chromomophores), nomysins, actinomycin, authramycin, azaserine, bleomycins, cactinomycin, carabicin, caminomycin, carzinophilin, chromomycins, dactinomycin, daunorubicin, detorubicin, 6—diazooxo-L- norleucine, doxorubicin (AdriamycinT'V') ding morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2—pyrrolino-doxorubicin, and deoxydoxorubicin), epirubucin, esorubicin, idarubicin, marcellomycin, mitomycins such as mitomycin C, enolic acid, nogalamycin, olivomycins, peplomycin, omycin, puromycine, quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ex, zinostatin, zorubicin; anti-metabolites such a methotrexate and 5-fluorouracil (5-FU); folic acid analogues such as denopterin, methotrexate, pteropterin, rexate; purine analogs such as fludarabine, aptopurine, prine, thioguanine; pyrimidine analogs such as ancitabine, azacitidine, 6—azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine; androgens such as calusterone, dromostanolone nate, epitiostanol, mepitiostane, testolactone; anti-adranals such as aminoglutethimide, mitotane, trilostane; folic acid replenisher such as frolinic acid; aceglatone; aldophosphamide glycoside; aminolevulinic acid; eniluracil; ine; bestrabucil; bisantrene; edatraxate; defofamine; lcine; diaziquone; elfomithine; elliptinium e; an lone; etoglucid; gallium nitrate; hydroxyurea; lentinan; Ionidamine; maytansinoids such as maytansine and ansamitocins; mitoguazone, mitoxantrone; mopidamol; nitracrine; pentostatin; phenamet; pirarubicin; Iosoxantrone; yllinic acid; 2—ethylhydrazide; procarbazine; PSK®; razoxane; rhizoxin; sizofuran; spirogermanium; tenuazonic acid; quone; 2,2',2"-trichlorotriethylamine; trichothecenes ially T-2 toxin, verracurin A, roridin A and ine); urethan; vindesine; dacarbazine; mannomustine; mitabronitol; mitolactol; pipobroman; gacytosine; arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxoids, e.g., paclitaxel (TAXOL®, l-Myers Squibb Oncology, Princeton, NJ.) and doxetaxel (TAXOTERE®, Rhone-Poulenc Rorer, Antony, France); chlorambucil; gemcitabine (GemzarT'V'); 6-thioguanine; mercaptopurine; methotrexate; platinum analogs such as cisplatin and carboplatin; vinblastine; platinum; etoposide ); ifosfamide; mitoxantrone; vincristine; vinorelbine NavelbineT'V'); novantrone; teniposide; edatrexate; daunomycin; aminopterin; xeloda; ibandronate; OPT-11; topoisomerase inhibitor RFS 2000; difluoromethylornithine (DMFO); retinoids such as ic acid; capecitabine; and pharmaceutically acceptable salts, acids, or derivatives of any of the above. Also included in this tion are ormonal agents that act to regulate or inhibit hormone action on tumors such as anti-estrogens and ive estrogen receptor modulators (SERMs), including, for example, tamoxifen (including NolvadexT'V'), raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, and toremifene (FarestonTM); aromatase inhibitors that inhibit the enzyme aromatase, which regulates estrogen production in the adrenal glands, such as, for example, 4(5)-imidazoles, aminoglutethimide, megestrol acetate (MegaceT'V'), exemestane, formestane, fadrozole, vorozole (RivisorT'V'), letrozole (FemaraT'V'), and anastrozole (ArimidexT'V'); and anti-androgens such as flutamide, mide, bicalutamide, leuprolide, and goserelin; and pharmaceutically acceptable salts, acids, or tives of any of the above. Any one or more of these agents may be conjugated to the humanized antibodies of the present invention to provide a useful therapeutic agent for the treatment of various disorders.

The dies also may be ated to prodrugs. A "prodrug" is a precursor or derivative form of a pharmaceutically active substance that is less cytotoxic to tumor cells compared to the parent drug and is capable of being enzymatically activated or converted into the more active form. See, for example, Wilman, 1986, ugs in Cancer Chemotherapy", ln Biochemical Society Transactions, 14, pp. 375-382, 615th Meeting Belfast and Stella et al., 1985, "Prodrugs: A Chemical Approach to Targeted Drug Delivery, ln: "Directed Drug Delivery, Borchardt et al., (ed.), pp. 247-267, Humana Press. Useful prodrugs include, but are not d to, phosphate-containing gs, thiophosphate-containing prodrugs, sulfate-containing prodrugs peptide-containing prodrugs, o acid-modified prodrugs, ylated prodrugs, B-lactam-containing prodrugs, optionally tuted phenoxyacetamide-containing prodrugs, and optionally substituted phenylacetamide-containing prodrugs, 5-fluorocytosine and other 5- fluorouridine prodrugs that can be converted into the more active cytotoxic free drug.

Examples of cytotoxic drugs that can be derivatized into a prodrug form include, but are not limited to, those chemotherapeutic agents described above.

For diagnostic as well as eutic monitoring es, the antibodies of the invention also may be conjugated to a label, either a label alone or a label and an additional second agent (prodrug, chemotherapeutic agent and the like). A label, as distinguished from the other second agents refers to an agent that is a detectable compound or composition and it may be conjugated directly or indirectly to a zed antibody of the present invention. The label may itself be detectable (e.g., radioisotope labels or fluorescent labels) or, in the case of an enzymatic label, may catalyze chemical alteration of a substrate compound or composition that is detectable. Labeled humanized anti-lL-36R antibody can be prepared and used in various ations including in vitro and in vivo diagnostics.

The antibodies of the present ion may be formulated as part of a liposomal preparation in order to affect ry thereof in vivo. A "liposome" is a small vesicle composed of various types of lipids, phospholipids, and/or surfactant. Liposomes are useful for delivery to a mammal of a compound or formulation, such as a humanized anti-lL-36R antibody disclosed herein, optionally, coupled to or in combination with one or more pharmaceutically active agents and/or labels. The components of the liposome are commonly arranged in a bilayer formation, similar to the lipid arrangement of biological membranes.

Certain aspects of the present ion related to isolated c acids that encode one or more domains of the humanized antibodies of the present invention. An "isolated" nucleic acid molecule is a nucleic acid molecule that is identified and separated from at least one contaminant nucleic acid molecule with which it is ordinarily associated in the natural source of the antibody c acid. An isolated nucleic acid molecule is distinguished from the nucleic acid molecule as it exists in natural cells.

In various aspects of the present invention one or more domains of the humanized antibodies will be recombinantly expressed. Such recombinant expression may employ one or more control sequences, i.e., polynucleotide sequences necessary for expression of an operably linked coding sequence in a particular host organism. The control sequences suitable for use in yotic cells e, for example, promoter, operator, and ribosome binding site sequences. Eukaryotic control sequences include, but are not limited to, promoters, polyadenylation signals, and enhancers. These control sequences can be utilized for sion and production of humanized anti-lL-36R antibody in yotic and eukaryotic host cells.

A c acid sequence is "operably linked" when it is placed into a functional relationship with another nucleic acid sequence. For example, a nucleic acid presequence or secretory leader is operably linked to a nucleic acid encoding a ptide if it is expressed as a tein that participates in the secretion of the polypeptide; a promoter or enhancer is operably linked to a coding sequence if it affects the ription of the sequence; or a ribosome binding site is operably linked to a coding sequence if it is positioned so as to tate translation. Generally, "operably linked" means that the DNA sequences being linked are contiguous, and, in the case of a secretory leader, uous and in g frame. However, enhancers are optionally uous. Linking can be accomplished by ligation at convenient ction sites. If such sites do not exist, synthetic oligonucleotide adaptors or linkers can be used.

As used herein, the sions "cell", "cell line", and "cell culture" are used interchangeably and all such designations include the y thereof. Thus, "transformants" and "transformed cells" include the primary subject cell and cultures derived rom without regard for the number of transfers.

The term "mammal" for purposes of treatment refers to any animal classified as a mammal, including humans, domesticated and farm animals, and zoo, sports, or pet animals, such as dogs, horses, cats, cows, and the like. Preferably, the mammal is human.

A "disorder", as used herein, is any condition that would t from treatment with a zed anti-lL-36R antibody described herein. This includes chronic and acute disorders or diseases including those pathological conditions that predispose the mammal to the disorder in question. Non-limiting examples or disorders to be treated herein include inflammatory, angiogenic, autoimmune and immunologic disorders, respiratory disorders, cancer, hematological malignancies, benign and malignant tumors, leukemias and lymphoid ancies.

The terms "cancer" and "cancerous" refer to or be the physiological condition in mammals that is typically terized by lated cell growth. Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and leukemia.

An lL-36R-associated disorder includes diseases and disorders of the immune system, such as autoimmune disorders and inflammatory disorders. Such conditions include, but are not limited to, rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), scleroderma, Sjogren's syndrome, multiple sclerosis, sis, psoriatic arthritis, inflammatory bowel disease (e.g., ulcerative colitis and Crohn's disease), pulmonary inflammation, asthma, idiopathic ocytopenic purara (lTP) epithelial inflammatory disorders, fibrosis and ankylosing spondylitis.

The term "intravenous infusion" refers to introduction of an agent into the vein of an animal or human patient over a period of time greater than approximately 15 minutes, lly between approximately 30 to 90 s.

The term venous bolus" or "intravenous push" refers to drug stration into a vein of an animal or human such that the body receives the drug in approximately 15 minutes or less, generally 5 minutes or less.

The term "subcutaneous administration" refers to introduction of an agent under the skin of an animal or human patient, preferable within a pocket between the skin and ying tissue, by relatively slow, sustained delivery from a drug receptacle. Pinching or drawing the skin up and away from underlying tissue may create the pocket.

The term "subcutaneous infusion" refers to introduction of a drug under the skin of an animal or human patient, preferably within a pocket between the skin and underlying tissue, by relatively slow, ned delivery from a drug receptacle for a period of time including, but not limited to, 30 minutes or less, or 90 minutes or less. Optionally, the infusion may be made by subcutaneous implantation of a drug delivery pump implanted under the skin of the animal or human patient, wherein the pump delivers a predetermined amount of drug for a predetermined period of time, such as 30 minutes, 90 minutes, or a time period spanning the length of the treatment regimen.

The term "subcutaneous bolus" refers to drug administration beneath the skin of an animal or human patient, where bolus drug delivery is less than approximately 15 s; in another aspect, less than 5 minutes, and in still another aspect, less than 60 seconds. In yet even another aspect, administration is within a pocket between the skin and underlying tissue, where the pocket may be created by ng or drawing the skin up and away from underlying tissue.

The term "therapeutically ive amount" is used to refer to an amount of an active agent that relieves or ameliorates one or more of the symptoms of the disorder being treated. In r aspect, the therapeutically effective amount refers to a target serum concentration that has been shown to be effective in, for e, slowing disease progression. Efficacy can be measured in conventional ways, depending on the condition to be treated.

WO 74569 The terms "treatment" and "therapy" and the like, as used herein, are meant to include therapeutic as well as prophylactic, or suppressive measures for a disease or disorder leading to any clinically desirable or beneficial effect, including but not limited to alleviation or relief of one or more symptoms, sion, slowing or cessation of progression of the disease or disorder. Thus, for example, the term treatment includes the stration of an agent prior to or following the onset of a m of a disease or disorder thereby preventing or removing one or more signs of the disease or disorder. As another example, the term includes the administration of an agent after clinical manifestation of the disease to combat the symptoms of the disease. Further, administration of an agent after onset and after clinical symptoms have developed where administration affects al ters of the disease or disorder, such as the degree of tissue injury or the amount or extent of metastasis, r or not the treatment leads to amelioration of the disease, comprises "treatment" or "therapy" as used herein. Moreover, as long as the itions of the invention either alone or in combination with another therapeutic agent alleviate or ameliorate at least one symptom of a disorder being treated as compared to that m in the absence of use of the humanized L-36R antibody composition, the result should be considered an effective ent of the ying disorder regardless of whether all the symptoms of the disorder are alleviated or not.

The term "package insert" is used to refer to instructions customarily included in commercial packages of therapeutic products, that contain information about the indications, usage, administration, contraindications and/or warnings concerning the use of such therapeutic products.

Antibodies In one , described and disclosed herein are anti-lL-36R antibodies, in particular humanized anti-lL-36R antibodies, and compositions and articles of manufacture comprising one or more anti-lL-36R antibody, in particular one or more humanized anti- lL-36R antibody of the present invention. Also described are binding agents that include an antigen-binding fragment of an anti-lL-36 antibody, in particular a zed anti-IL- 36R antibody.

Variable regions and CDRs of representative antibodies of the present invention are disclosed below: Anti-lL-36R Mouse Antibody Seguences Variable regions and CDRs of representative mouse lead antibodies of the t invention (mouse leads) are shown below: Light Chain le Region (VK) Amino Acid Sequences >33D10B12VK Protein (antibody 33D 1 0) QIVLTQSPAIMSASLGERVTMTCTASSSVSSSYLHWYQKKPGSSPKLWVYSTSNLASGV PVRFSGSGSGTSYSLTISSMEAEDAATYYCHQHHRSPVTFGSGTKLEMK (SEQ ID NO: 1) >172C8B 12 VK protein ody 172C8) SPASQSASLGESVTFTCLASQTIGTWLAWYQQRPGKSPQLLIYAATSLADGVPS RFSGSGSGTQFSFNIRSLQAEDFASYYCQQVYTTPLTFGGGTKLEIK (SEQ ID NO: 2) >67E7E8 VK protein (antibody 67E7) DIQMTQSPASQSASLGESVTFTCLASQTIGTWLGWYQQKPGKSPQLLIYRSTTLADGVPS RFSGSGSGTKFSFKISSLQAADFASYYCQQLYSAPYTFGGGTKLEIR (SEQ ID NO: 3) >78C8D1 VK Protein (antibody 78C8) DVLLTQTPLSLPVSLGDQASISCRSSQNIVHSNGNTYLQWYLQKPGQSPKLLIYKVSNRF SGVPDRFSGSGSGTDFTLKISRVEAEDLGVYYCFQGSHVPFTFGAGTKLELK (SEQ ID NO: 4) >81A1D1 VK Protein (antibody 81A1) DIQMTQTTSSLSASLGDRVTISCRASQDIYKYLNWYQQKPDGTLKLLIYYTSGLHSGVPS RFSGSGSGTDFSLTISNLEPEDIATYFCQQDSKFPWTFGGDTKLEIK (SEQ ID NO: 5) >81B4E11 VK Protein (antibody 81B4) QIVLTQSPAIMSASLGERVTMTCTASSSVSSSYFHWYQQKPGSSPKLWIYRTSNLASGVP GSGTSYSLTISSMEAEDAATYYCHQFHRSPLTFGAGTKLELK (SEQ ID NO: 6) >73C5C10 VK protein (antibody 73C5) DIVMTQSQKFLSTSVGVRVSVTCKASQDVGTNVLWYQQKIGQSPKPLIYSASYRHSGVP DRFTGSGSGTDFTLIISNVQSEDLAEYFCQQYSRYPLTFGPGTKLELK (SEQ ID NO: 7) >73F6F8 VK protein (antibody 73F6) DIVMTQSQKFLSTSVGVRVSVTCKASQDVGTNVLWYQQKIGQSPKALIYSASYRHSGVP GSGTDFTLIITNVQSEDLAEYFCQQYSRYPLTFGPGTKLELK (SEQ ID NO: 8) >76E10E8 VK protein (antibody 76E10) DIVMTQSQKFMSATVGGRVNITCKASQNVGRAVAWYQQKPGQSPKLLTHSASNRYTG GSGSGTDFTLTITNMQSEDLADYFCQQYSSYPLTFGAGTKLDLK (SEQ ID NO: >89A12B8 VK protein (antibody 89A12) DIQMTQSPASQSASLGESVTFSCLASQTIGTWLGWYQQKPGKSPQLLIYRATSLADGVPS RFSGSGSGTNFSFKISSLQAEDLASYYCQQLYSGPYTFGGGTKLEIR (SEQ ID NO: 10) Heavy Chain Variable Region (VH) Amino Acid Sequences >33D10B12VH Protein (antibody 33D 1 0) QVQLQQSGTELLKPGASVKLSCKASGNTVTSYWMHWVKQRPGQGLEWIGEILPSTGRT NYNENFKGKAMLTVDKSSSTAYMQLSSLASEDSAVYYCTIVYFGNPWFAYWGQGTLV TVSA (SEQ ID NO: 11) >172C8B12 VH n (antibody 172C8) SGPELVKPGASVKLSCKASGYTFTDNYMNWVRQSHGKSLEWIGRVNPSNGD TKYNQNFKGKATLTVDKSLSTAYMQLNGLTSEDSAVYYCGRTKNFYSSYSYDDAMDY WGQGTSVTVSS (SEQ ID NO: 12) >67E7E8 VH protein (antibody 67E7) EVQLQQSGAEFVRPGASVKFSCTASGFNIKDDYIHWVRQRPEQGLEWVGRIDPANGNT QDKATITADTSSNTAYLQLSSLTSEDTAVYYCAKSFPNNYYSYDDAFAYWGQ GTLVTVSA (SEQ ID NO: 13) >78C8D1 VH Protein (antibody 78C8) QVQLKESGPVLVAPSQSLSITCTVSGFSLTKFGVHWIRQTPGKGLEWLGVIWAGGPTNY NSALMSRLTISKDISQSQVFLRIDSLQTDDTAMYYCAKQIYYSTLVDYWGQGTSVTVSS (SEQ ID NO: 14) >81A1D1 VH Protein (antibody 81A1) QVQLKESGPGLVAPSQSLFITCTVSGFSLSSYEINWVRQVPGKGLEWLGVIWTGITTNYN SALISRLSISKDNSKSLVFLKMNSLQTDDTAIYYCARGTGTGFYYAMDYWGQGTSVTVS S (SEQ ID NO: 15) >81B4E11 VH Protein (antibody 81B4) QVQLQQPGADFVRPGASMRLSCKASGYSFTSSWIHWVKQRPGQGLEWIGEINPGNVRT NYNENFRNKATLTVDKSSTTAYMQLRSLTSADSAVYYCTVVFYGEPYFPYWGQGTLVT VSA (SEQ ID NO: 16) >73C5C10 VH Protein (antibody 73C5) QVQLKESGPGLVAPSQSLSITCTVSGFSLTNYAVHWVRQFPGKGLEWLGVIWSDGSTDF NAPFKSRLSINKDNSKSQVFFKMNSLQIDDTAIYYCARKGGYSGSWFAYWGQGTLVTV SA (SEQ ID NO: 17) >73F6F8 VH protein ody 73F6) QVQLKESGPGLVAPSQSLSITCTVSGFSLTNYAVHWVRQFPGKGLEWLGVIWSDGSTDY NAPFKSRLSINKDNSKSQVFFKMNSLQTDDTAIYYCARKGGYSGSWFAYWGQGTLVTV SA (SEQ ID NO: 18) >76E10E8 VH n (antibody 76E10) SGPVLVAPSQSLSITCTVSGFSLTNYGVHWVRQPPGKGLEWLGVIWPVGSTNY NSALMSRLSIHKDNSKSQVFLRMNSLQTDDTAIYYCAKMDWDDFFDYWGQGTTLTVSS (SEQ ID NO: 19) >89A12B8 VH Protein (antibody 89A12) EVQLQQSGAELVRPGASVRLSCTASGENIKDDYIHWVRQRPKQGLEWLGRIDPANGNT KYDPRFQDKATITADTSSNTAYLHLSSLTSEDTAVYYCAKSFPDNYYSYDDAFAYWGQ GTLVTVSA (SEQ ID NO: 20) Light chain CDR-l (L-CDRl) Amino Acid Sequences >33D10G1 L—CDR1 TASSSVSSSYLH (SEQ ID NO: 21) >172C8B12 L-CDRl LASQTIGTWLA (SEQ ID NO: 22) >67E7E8 L-CDR1 LASQTIGTWLG (SEQ ID NO: 23) >78C8D1 L-CDRl RSSQNIVHSNGNTYLQ (SEQ ID NO: 24) >81A1D1L—CDR1 YKYLN (SEQ ID NO: 25) 11 L—CDRI TASSSVSSSYFH (SEQ ID NO: 26) >73C5C10 L—CDRl KASQDVGTNVL (SEQ ID NO: 27) >73F6F8 L—CDRl KASQDVGTNVL (SEQ ID NO: 27) >76E10E8 L—CDRI KASQNVGRAVA (SEQ ID NO: 28) B8 L—CDRI LASQTIGTWLG (SEQ ID NO: 29) Light chain CDR-2 (L-CDR2) Amino Acid Sequences >33D10B12 L—CDR2 STSNLAS (SEQ ID NO: 30) >172C8B12 L—CDR2 AATSLAD ( SEQ ID NO: 31) >67E7E8 L—CDR2 RSTTLAD (SEQ ID NO: 32) >78C8D1 L—CDR2 KVSNRFS (SEQ ID NO: 33) >81A1D1 L—CDR2 YTSGLHS (SEQ ID NO: 34) >81B4E11 L—CDR2 RTSNLAS (SEQ ID NO: 35) >73C5C10 L—CDR2 SASYRHS (SEQ ID NO: 36) >73F6F8 L—CDR2 SASYRHS (SEQ ID NO: 36) >76E10E8 L—CDR2 SASNRYT (SEQ ID NO: 37) >89A12B8 L—CDR2 RATSLAD (SEQ ID NO: 38) Light chain CDR-3 (L-CDR3) Amino Acid Sequences >33D10B12 L—CDR3 HQHHRSPVT (SEQ ID NO: 39) >172C8B12 L—CDR3 QQVYTTPLT (SEQ ID NO: 40) >67E7E8 L—CDR3 QQLYSAPYT (SEQ ID NO: 41) >78C8D1 L—CDR3 FQGSHVPFT (SEQ ID NO: 42) 1 L—CDR3 QQDSKFPWT (SEQ ID NO: 43) >81B4E11 L—CDR3 HQFHRSPLT (SEQ ID NO: 44) >73C5C10 L—CDR3 QQYSRYPLT (SEQ ID NO: 45) >73F6F8 L—CDR3 QQYSRYPLT (SEQ ID NO: 45) >76E10E8 L—CDR3 PLT (SEQ ID NO: 46) >89A12B8 L—CDR3 QQLYSGPYT (SEQ ID NO: 47) Heavy chain CDR-l (H-CDRl) Amino Acid Sequences >33D10B12 H—CDRI GNTVTSYWMH (SEQ ID NO: 48) >172C8B 12 H-CDRI GYTFTDNYMN (SEQ ID NO: 49) WO 74569 >67E7E8 H—CDR1 GFNIKDDYIH (SEQ ID NO: 50) >78C8D1 H—CDR1 GFSLTKFGVH (SEQ ID NO: 51) >81A1D1 H—CDR1 GFSLSSYEIN (SEQ ID NO: 52) >81B4E11 H—CDR1 GYSFTSSWIH (SEQ ID NO: 53) >73C5C10 H—CDRl GFSLTNYAVH (SEQ ID NO: 54) >73F6F8 H-CDR1 GFSLTNYAVH (SEQ ID NO: 54) >76E10E8 H—CDR1 GFSLTNYGVH (SEQ ID NO: 55) >89A12B8 H-CDR1 GFNIKDDYIH (SEQ ID NO: 56) Heavy chain CDR-2 (H-CDR2) Amino Acid Sequences >33D10B12 H-CDR2 EILPSTGRTNYNENFKG (SEQ ID NO: 57) >172C8B 12 H—CDR2 GDTKYNQNFKG (SEQ ID NO: 58) >67E7E8 H—CDR2 RIDPANGNTKYAPKFQD (SEQ ID NO: 59) >78C8D1 H—CDR2 VIWAGGPTNYNSALMS (SEQ ID NO: 60) 1 H-CDR2 VIWTGITTNYNSALIS (SEQ ID NO: 61) >81B4E11 H-CDR2 EINPGNVRTNYNENF (SEQ ID NO: 62) >73C5C10 H—CDR2 STDFNAPFKS (SEQ ID NO: 63) >73F6F8 H—CDR2 VIWSDGSTDYNAPFKS (SEQ ID NO: 64) >76E10E8 H-CDR2 VIWPVGSTNYNSALMS (SEQ ID NO: 65) >89A12B8 H-CDR2 RIDPANGNTKYDPRFQD (SEQ ID NO: 66) Heavy chain CDR-3 (H-CDR3) Amino Acid Sequences >33D10B12 H-CDR3 VYFGNPWFAY (SEQ ID NO: 67) >172C8B 12 H-CDR3 TKNFYSSYSYDDAMDY (SEQ ID NO: 68) 8 H-CDR3 SFPNNYYSYDDAFAY (SEQ ID NO: 69) >78C8D1 H-CDR3 LVDY (SEQ ID NO: 70) >81A1D1 H—CDR3 GTGTGFYYAMDY (SEQ ID NO: 71) >81B4E11 H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >73C5C10 H—CDR3 KGGYSGSWFAY (SEQ ID NO: 73) >73F6F8 H—CDR3 KGGYSGSWFAY (SEQ ID NO: 73) >76E10E8 H—CDR3 MDWDDFFDY (SEQ ID NO: 74) >89A12B8 H-CDR3 SFPDNYYSYDDAFAY (SEQ ID NO: 75) Anti-lL-36R Mouse CDR Sequences A summary of the CDR seguences of the lead mouse dies is shown below: Antibody H-CDR Sequences L—CDR Sequences 33D10 GNTVTSYWMH (H—CDRI) TASSSVSSSYLH (L—CDRI) SEQ ID No: 48 SEQ ID No: 21 EILPSTGRTNYNENFKG STSNLAS (L—CDR2) SEQ ID 2) SEQ ID No: 57 No: 30 VYFGNPWFAY (H—CDR3) HQHHRSPVT (L—CDR3) SEQ ID No: 67 SEQ ID No: 39 172C8 GYTFTDNYMN (H—CDRI) LASQTIGTWLA (L—CDRI) SEQ ID No: 49 SEQ ID No: 22 RVNPSNGDTKYNQNFKG AATSLAD (L—CDR2) SEQ (H-CDR2) SEQ ID No: 58 ID No: 31 TKNFYSSYSYDDAMDY QQVYTTPLT (L—CDR3) 3) SEQ ID No: 68 SEQ ID No: 40 67E7 GFNIKDDYIH (H—CDRI) LASQTIGTWLG (L—CDRI) SEQ ID No: 50 SEQ ID No: 23 RIDPANGNTKYAPKFQD RSTTLAD (L—CDR2) SEQ ID (H-CDR2) SEQ ID No: 59 No: 32 YSYDDAFAY (H- QQLYSAPYT 3) CDR3) SEQ ID NO: 69 SEQ ID No: 41 GFSLTKFGVH (H—CDRI) RSSQNIVHSNGNTYLQ (L— SEQ ID No: 51 CDRl) SEQ ID No: 24 VIWAGGPTNYNSALMS KVSNRFS (L—CDR2) SEQ ID (H-CDR2) SEQ ID No: 60 No: 33 QIYYSTLVDY (H—CDR3) FQGSHVPFT (L—CDR3) SEQ ID No: 42 SEQ ID No: 70 YEIN (H—CDRI) RASQDIYKYLN I) SEQ ID No: 52 SEQ ID No: 25 YTSGLHS (L—CDR2) SEQ ID VIWTGITTNYNSALIS (H— No: 34 CDR2) SEQ ID No: 61 QQDSKFPWT 3) SEQ ID No: 43 GTGTGFYYAMDY (H— CDR3) SEQ ID No: 71 GYSFTSSWIH (H—CDRI) TASSSVSSSYFH (L—CDRI) SEQ ID No: 53 SEQ ID No: 26 RTSNLAS (L—CDR2) SEQ ID EINPGNVRTNYNENF (H- No: 35 CDR2) SEQ ID No: 62 HQFHRSPLT (L—CDR3) SEQ ID No: 44 VFYGEPYFPY (H—CDR3) SEQ ID No: 72 GFSLTNYAVH (H—CDRI) KASQDVGTNVL (L—CDRI) SEQ ID No: 54 SEQ ID No: 27 VIWSDGSTDFNAPFKS (H- SASYRHS (L—CDR2) SEQ ID CDR2) SEQ ID NO: 63 No: 36 KGGYSGSWFAY (H—CDR3) QQYSRYPLT (L—CDR3) SEQ ID No: 73 SEQ ID No: 45 GFSLTNYAVH (H—CDRI) KASQDVGTNVL (L—CDRI) SEQ ID No: 54 SEQ ID N0:27 VIWSDGSTDYNAPFKS (H- SASYRHS (L—CDR2) SEQ ID CDR2) SEQ ID No: 64 No: 36 KGGYSGSWFAY (H—CDR3) PLT (L—CDR3) SEQ ID No: 73 SEQ ID No: 45 YGVH I) KASQNVGRAVA (L-CDRI) SEQ ID NO: 55 SEQ ID No: 28 VIWPVGSTNYNSALMS (H— SASNRYT (L—CDR2) SEQ CDR2) SEQ ID No: 65 ID NO: 37 MDWDDFFDY (H—CDR3) QQYSSYPLT (L—CDR3) SEQ “3 N03 46 SEQ ID No: 74 GFNIKDDYIH (H—CDRI) LASQTIGTWLG (L—CDRI) SEQ ID NO: 56 SEQ ID No: 29 RIDPANGNTKYDPRFQD RATSLAD (L—CDR2) SEQ (H-CDR2) SEQ ID No: 66 ID No: 38 QQLYSGPYT (L—CDR3) SFPDNYYSYDDAFAY (H— SEQ ID No: 47 CDR3) SEQ ID No: 75 Anti-lL-36R Humanized Antibody Seguences Human framework sequences were selected for the mouse leads based on the framework homology, CDR structure, ved canonical residues, conserved interface g residues and other parameters to produce humanized variable regions (see Example 5).

Representative zed variable regions derived from antibodies 81 B4 and 7305 are shown below.

Light Chain Variable Region (VK) Amino Acid Sequences K32_3 VK protein EIVLTQSPGTLSLSPGERATMSCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSTLASGIPD RFSGSGSGTDFTLTISRLEPEDAATYYCHQFHRSPLTFGQGTKLEIK (SEQ ID NO: 76) >81B4VK32_105 VK protein EIVLTQSPGTLSLSPGERATMSCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSILASGVPD RFSGSGSGTDFTLTISRLEPEDFATYYCHQFHRSPLTFGQGTKLEIK (SEQ ID NO: 77) >81B4VK32_116 VK n EIVLTQSPGTLSLSPGERATMSCTASSSVSSSYFHWYQQKPGQAPRLWIYRTSRLASGVP DRFSGSGSGTDFTLTISRLEPEDAATYYCHQFHRSPLTFGQGTKLEIK (SEQ ID NO: 78) >81B4VK32_127 VK n EIVLTQSPGTLSLSPGERATMTCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSRLASGVP DRFSGSGSGTDFTLTISRLEPEDFAVYYCHQFHRSPLTFGQGTKLEIK (SEQ ID NO: 79) >81B4VK32_13 8 VK protein QIVLTQSPGTLSLSPGERATMTCTASSSVSSSYFHWYQQKPGQAPRLWIYRTSRLASGVP DRFSGSGSGTDFTLTISRLEPEDAATYYCHQFHRSPLTFGAGTKLEIK (SEQ ID NO: 80) >81B4VK32_140 VK protein QIVLTQSPGTLSLSPGERVTMSCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSQLASGIPD RFSGSGSGTDFTLTISRLEPEDAATYYCHQFHRSPLTFGQGTKLEIK (SEQ ID NO: 81) >81B4VK32_141 VK protein SPGTLSLSPGERATMTCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSKLASGVP DRFSGSGSGTDFTLTISRLEPEDFATYYCHQFHRSPLTFGQGTKLEIK (SEQ ID NO: 82) >81B4VK32_147 VK protein EIVLTQSPGTLSLSPGERATMSCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSHLASGIPG RFSGSGSGTDFTLTISRLEPEDAAVYYCHQFHRSPLTFGQGTKLEIK (SEQ ID NO: 83) >73C5VK39_2 VK n EIVMTQSPATLSVSPGVRATLSCKASQDVGTNVLWYQQKPGQAPRPLIYSASYRHSGIP DRFSGSGSGTEFTLTISSLQSEDFAEYFCQQYSRYPLTFGQGTKLEIK (SEQ ID NO: 84) >73C5VK39_7 VK protein EIVMTQSPATLSVSPGVRATLSCKASQDVGTNVLWYQQKPGQAPRPLIYSASYRHSGIP DRFSGSGSGTEFTLTISSLQSEDFAVYYCQQYSRYPLTFGQGTKLEIK (SEQ ID NO: 85) >73C5VK39_15 VK protein EIVMTQSPATLSVSPGVRATLSCKASQDVGTNVLWYQQKPGQAPRPLIYSASYRHSGIP ARFSGSGSGTEFTLTISSLQSEDFAEYYCQQYSRYPLTFGQGTKLEIK (SEQ ID NO: 86) Heavy Chain le Region (VH) Amino Acid Sequences >81B4VH33_49 VH Protein QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQAPGQGLEWIGEINPGNVRT NYNENFRNKATMTVDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLVT VSS (SEQ ID NO: 87) >81B4VH33_85T VH Protein QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQRPGQGLEWIGEINPGNVRT RNRVTMTVDTSISTAYMELSRLRSDDTAVYYCTVVFYGEPYFPYWGQGTLVT VSS (SEQ ID NO: 88) >81B4VH33_90 VH Protein QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVKQAPGQGLEWMGEINPGNVR TNYNENFRNKVTMTVDTSISTAYMELSRLRSDDTAVYYCTVVFYGEPYFPYWGQGTLV TVSS (SEQ ID NO: 89) >81B4VH33_93 VH Protein QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQRPGQGLEWMGEINPGNVR TNYNENFRNRATLTRDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLV TVSS (SEQ ID NO: 90) >81B4VH50_22 VH Protein QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQRPGQGLEWMGEILPGVVR TNYNENFRNKVTMTVDTSISTAYMELSRLRSDDTAVYYCTVVFYGEPYFPYWGQGTLV TVSS (SEQ ID NO: 91) >81B4VH50_30 VH Protein SGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQRPGQGLEWIGEINPGAVRT NYNENFRNRVTMTVDTSISTAYMELSRLRSDDTAVYYCTVVFYGEPYFPYWGQGTLVT VSS (SEQ ID NO: 92) >81B4VH51_13 VH Protein QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQAPGQGLEWIGEINPGLVRT NYNENFRNKVTMTVDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLVT VSS (SEQ ID NO: 93) >81B4VH51_15 VH Protein QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQAPGQGLEWIGEINPGAVRT NYNENFRNKVTMTVDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLVT VSS (SEQ ID NO: 94) >81B4VH52_83 VH Protein QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQAPGQGLEWIGEINPGSVRT NYNENFRNKATMTVDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLVT VSS (SEQ ID NO: 95) >73C5VH46_4 VH Protein QVQLQESGPGLVKPSETLSITCTVSGFSLTDYAVHWIRQPPGKGLEWIGVIWSDGSTDYN APFKSRVTINKDTSKSQVSFKMSSVQAADTAVYYCARKGGYSGSWFAYWGQGTLVTV SS (SEQ ID NO: 96) >73C5VH46_19 VH Protein QVQLQESGPGLVKPSETLSITCTVSGFSLTDYAVHWIRQPPGKGLEWIGVIWSDGSTDYN APFKSRVTISKDTSKNQVSLKMNSLTTDDTAVYYCARKGGYSGSWFAYWGQGTLVTV SS (SEQ ID NO: 97) >73C5VH46_40 VH Protein SGPGLVKPSETLSITCTVSGFSLTDYAVHWIRQPPGKGLEWIGVIWSDGSTDYN APFKSRVTISKDNSKSQVSLKMNSVTVADTAVYYCARKGGYSGSWFAYWGQGTLVTV SS (SEQ ID NO: 98) >73C5VH47_65 VH Protein QVQLQESGPGLVKPSETLSITCTVSGFSLTDYAVHWVRQPPGKGLEWIGVIWSDGSTDY NAPFKSRVTISKDTSKNQVSFKLSSVTVDDTAVYYCARKGGYSGSWFAYWGQGTLVTV SS (SEQ ID NO: 99) >73C5VH47_77 VH Protein SGPGLVAPSETLSLTCTVSGFSLTDYAVHWIRQFPGKGLEWIGVIWSDGSTDF RVTISKDTSKNQVSFKLSSVTTDDTAVYYCARKGGYSGSWFAYWGQGTLVTV SS (SEQ ID NO: 100) >73C5VH58_91 VH Protein QVQLQESGPGLVKPSETLSITCTVSGFSLTDYAVHWIRQPPGKGLEWIGVIWSDGSTDYN APFKSRVTISKDNSKSQVSFKMSSVTADDTAVYYCARKGGYSGSWFAYWGQGTLVTVS s (SEQ ID NO: 101) The CDR sequences from the humanized variable regions derived from antibodies 81 B4 and 7305 shown above are depicted below.

L-CDRI Amino Acid Seguences >81B4VK32_3 L—CDR1 TASSSVSSSYFH (SEQ ID NO: 26) >81B4VK32_105 L-CDR1 TASSSVSSSYFH (SEQ ID NO: 26) K32_116 L-CDR1 TASSSVSSSYFH (SEQ ID NO: 26) >81B4VK32_127 L-CDR1 TASSSVSSSYFH (SEQ ID NO: 26) K32_138 L-CDR1 TASSSVSSSYFH (SEQ ID NO: 26) >81B4VK32_140 L-CDR1 TASSSVSSSYFH (SEQ ID NO: 26) >81B4VK32_141 L-CDR1 TASSSVSSSYFH (SEQ ID NO: 26) >81B4VK32_147 L-CDR1 TASSSVSSSYFH (SEQ ID NO: 26) >73C5VK39_2 L-CDR1 KASQDVGTNVL (SEQ ID NO: 27) >73C5VK39_7 L—CDRl KASQDVGTNVL (SEQ ID NO: 27) >73C5VK39_15 L—CDRl KASQDVGTNVL (SEQ ID NO: 27) L-CDR2 Amino Acid ces >81B4VK32_3 L-CDR2 (SEQ ID 102) RTSTLAS >81B4VK32_105 L-CDR2 (SEQ ID 103) RTSILAS >81B4VK32_116 L-CDR2 (SEQ ID 104) RTSRLAS >81B4VK32_127 L-CDR2 (SEQ ID 104) RTSRLAS WO 74569 >81B4VK32_138 L—CDR2 (SEQ ID 104) RTSRLAS >81B4VK32_140 L—CDR2 (SEQ ID 105) RTSQLAS >81B4VK32_141 L—CDR2 (SEQ ID 106) RTSKLAS >81B4VK32_147 L-CDR2 (SEQ ID 140) RTSHLAS >73C5VK39_2 L—CDR2 SASYRHS (SEQ ID NO: 36) >73C5VK39_7 L—CDR2 SASYRHS (SEQ ID NO: 36) >73C5VK39_15 L-CDR2 SASYRHS (SEQ ID NO: 36) L-CDR3 Amino Acid Seguences >81B4VK32_3 L-CDR3 HQFHRSPLT (SEQ ID NO: 44) >81B4VK32_105 L-CDR3 HQFHRSPLT (SEQ ID NO: 44) K32_116 L-CDR3 HQFHRSPLT (SEQ ID NO: 44) >81B4VK32_127 L—CDR3 HQFHRSPLT (SEQ ID NO: 44) >81B4VK32_138 L-CDR3 HQFHRSPLT (SEQ ID NO: 44) >81B4VK32_140 L-CDR3 HQFHRSPLT (SEQ ID NO: 44) >81B4VK32_141 L-CDR3 HQFHRSPLT (SEQ ID NO: 44) >81B4VK32_147 L—CDR3 HQFHRSPLT (SEQ ID NO: 44) >73C5VK39_2 L-CDR3 QQYSRYPLT (SEQ ID NO: 45) >73C5VK39_7 L-CDR3 QQYSRYPLT (SEQ ID NO: 45) >73C5VK39_15 L—CDR3 QQYSRYPLT (SEQ ID NO: 45) H-CDRI Amino Acid Seguences >81B4VH33_49 H—CDRI GYSFTSSWIH (SEQ ID NO: 53) >81B4VH33_85T H—CDRI GYSFTSSWIH (SEQ ID NO: 53) >81B4VH33_90 H—CDRI SWIH (SEQ ID NO: 53) >81B4VH33_93 H—CDRI GYSFTSSWIH (SEQ ID NO: 53) >81B4VH50_22 H—CDRI GYSFTSSWIH (SEQ ID NO: 53) >81B4VH50_30 H—CDRI SWIH (SEQ ID NO: 53) H51_13 H—CDRI GYSFTSSWIH (SEQ ID NO: 53) >81B4VH51_15 H—CDRI GYSFTSSWIH (SEQ ID NO: 53) >81B4VH52_83 H—CDRI GYSFTSSWIH (SEQ ID NO: 53) >73C5VH46_4 H-CDR1 GFSLTDYAVH (SEQ ID NO: 107) >73C5VH46_19 H-CDR1 GFSLTDYAVH (SEQ ID NO: 107) >73C5VH46_40 H-CDR1 GFSLTDYAVH (SEQ ID NO: 107) >73C5VH47_65 H-CDR1 GFSLTDYAVH (SEQ ID NO: 107) H47_77 H—CDR1 GFSLTDYAVH (SEQ ID NO: 107) >73C5VH58_91 H-CDR1 GFSLTDYAVH (SEQ ID NO: 107) H-CDR2 Amino Acid Seguences H33_49 H-CDR2 EINPGNVRTNYNENF (SEQ ID NO: 62) >81B4VH33_85T H-CDR2 EINPGNVRTNYNENF (SEQ ID NO: 62) >81B4VH33_90 H-CDR2 EINPGNVRTNYNENF (SEQ ID NO: 62) >81B4VH33_93 H-CDR2 EINPGNVRTNYNENF (SEQ ID NO: 62) >81B4VH50_22 H-CDR2 EILPGVVRTNYNENF (SEQ ID NO: 108) >81B4VH50_30 H-CDR2 EINPGAVRTNYNENF (SEQ ID NO: 109) >81B4VH51_13 H—CDR2 VRTNYNENF (SEQ ID NO: 110) >81B4VH51_15 H-CDR2 EINPGAVRTNYNENF (SEQ ID NO: 109) >81B4VH52_83 H—CDR2 EINPGSVRTNYNENF (SEQ ID NO: 111) >73C5VH46_4 H-CDR2 STDYNAPFKS (SEQ ID NO: 64) >73C5VH46_19 H-CDR2 VIWSDGSTDYNAPFKS (SEQ ID NO: 64) >73C5VH46_40 H-CDR2 VIWSDGSTDYNAPFKS (SEQ ID NO: 64) >73C5VH47_65 H-CDR2 VIWSDGSTDYNAPFKS (SEQ ID NO: 64) >73C5VH47_77 H—CDR2 VIWSDGSTDFNAPFKS (SEQ ID NO: 63) >73C5VH58_91 H-CDR2 VIWSDGSTDYNAPFKS (SEQ ID NO: 64) H-CDR3 Amino Acid Seguences H33_49 H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >81B4VH33_85T H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >81B4VH33_90 H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >81B4VH33_93 H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >81B4VH50_22 H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >81B4VH50_30 H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >81B4VH51_13 H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >81B4VH51_15 H-CDR3 VFYGEPYFPY (SEQ ID NO: 72) >81B4VH52_83 H—CDR3 VFYGEPYFPY (SEQ ID NO: 72) >73C5VH46_4 H—CDR3 SWFAY (SEQ ID NO: 73) >73C5VH46_19 H—CDR3 KGGYSGSWFAY (SEQ ID NO: 73) >73C5VH46_40 H—CDR3 KGGYSGSWFAY (SEQ ID NO: 73) >73C5VH47_65 H—CDR3 KGGYSGSWFAY (SEQ ID NO: 73) >73C5VH47_77 H—CDR3 KGGYSGSWFAY (SEQ ID NO: 73) >73C5VH58_91 H—CDR3 KGGYSGSWFAY (SEQ ID NO: 73) In one aspect, a variable region of the present ion is linked to a constant region.

For example, a variable region of the present invention is linked to a constant region shown below to form a heavy chain or a light chain of an antibody.

Heavy Chain Constant region linked downstream of a zed variable heavy region: ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSS GLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAAGG PSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQY NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSR EEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 112) Light Chain Constant region linked downstream of a humanized variable light region: SVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQ DSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO:113) Representative light chain and heavy chain sequences of the present invention are shown below (humanized variable regions derived from antibodies 81 B4 and 7305 linked to constant regions).

Light Chain Amino Acid Seguences >81B4VK32_3 Light Chain EIVLTQSPGTLSLSPGERATMSCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSTLASGIPD SGTDFTLTISRLEPEDAATYYCHQFHRSPLTFGQGTKLEIKRTVAAPSVFIFPPSD EQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTL SKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 114) >81B4VK32_105 Light Chain EIVLTQSPGTLSLSPGERATMSCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSILASGVPD RFSGSGSGTDFTLTISRLEPEDFATYYCHQFHRSPLTFGQGTKLEIKRTVAAPSVFIFPPSD EQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTL SKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 115) >81B4VK32_116 Light Chain EIVLTQSPGTLSLSPGERATMSCTASSSVSSSYFHWYQQKPGQAPRLWIYRTSRLASGVP DRFSGSGSGTDFTLTISRLEPEDAATYYCHQFHRSPLTFGQGTKLEIKRTVAAPSVFIFPPS DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 116) >81B4VK32_127 Light Chain EIVLTQSPGTLSLSPGERATMTCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSRLASGVP DRFSGSGSGTDFTLTISRLEPEDFAVYYCHQFHRSPLTFGQGTKLEIKRTVAAPSVFIFPPS DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 117) >81B4VK32_13 8 Light Chain QIVLTQSPGTLSLSPGERATMTCTASSSVSSSYFHWYQQKPGQAPRLWIYRTSRLASGVP DRFSGSGSGTDFTLTISRLEPEDAATYYCHQFHRSPLTFGAGTKLEIKRTVAAPSVFIFPPS DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 118) >81B4VK32_140 Light Chain SPGTLSLSPGERVTMSCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSQLASGIPD RFSGSGSGTDFTLTISRLEPEDAATYYCHQFHRSPLTFGQGTKLEIKRTVAAPSVFIFPPSD EQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTL SKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 119) >81B4VK32_141 Light Chain QIVLTQSPGTLSLSPGERATMTCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSKLASGVP DRFSGSGSGTDFTLTISRLEPEDFATYYCHQFHRSPLTFGQGTKLEIKRTVAAPSVFIFPPS DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 120) >81B4VK32_147 Light Chain EIVLTQSPGTLSLSPGERATMSCTASSSVSSSYFHWYQQKPGQAPRLLIYRTSHLASGIPG RFSGSGSGTDFTLTISRLEPEDAAVYYCHQFHRSPLTFGQGTKLEIKRTVAAPSVFIFPPSD TASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTL SKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 121) >73C5VK39_2 Light Chain SPATLSVSPGVRATLSCKASQDVGTNVLWYQQKPGQAPRPLIYSASYRHSGIP DRFSGSGSGTEFTLTISSLQSEDFAEYFCQQYSRYPLTFGQGTKLEIKRTVAAPSVFIFPPS DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 122) >73C5VK39_7 Light Chain EIVMTQSPATLSVSPGVRATLSCKASQDVGTNVLWYQQKPGQAPRPLIYSASYRHSGIP DRFSGSGSGTEFTLTISSLQSEDFAVYYCQQYSRYPLTFGQGTKLEIKRTVAAPSVFIFPPS DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 123) >73C5vK39_15 Light Chain EIVMTQSPATLSVSPGVRATLSCKASQDVGTNVLWYQQKPGQAPRPLIYSASYRHSGIP ARFSGSGSGTEFTLTISSLQSEDFAEYYCQQYSRYPLTFGQGTKLEIKRTVAAPSVFIFPPS DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 124) Heavy Chain Amino Acid Sequences >81B4vH33_49 Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQAPGQGLEWIGEINPGNVRT NYNENFRNKATMTVDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL SLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEA AGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPR EEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTL PPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 125) >81B4vH33_85T Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQRPGQGLEWIGEINPGNVRT NYNENFRNRVTMTVDTSISTAYMELSRLRSDDTAVYYCTVVFYGEPYFPYWGQGTLVT KGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEA AGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPR TYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTL PPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 126) >81B4vH33_90 Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVKQAPGQGLEWMGEINPGNVR TNYNENFRNKVTMTVDTSISTAYMELSRLRSDDTAVYYCTVVFYGEPYFPYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAV LQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPE AAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKP REEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY TLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSK LTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 127) >81B4vH33_93 Heavy Chain WO 74569 QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQRPGQGLEWMGEINPGNVR FRNRATLTRDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAV LQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPE AAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKP REEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY TLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSK LTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 128) >81B4vH50_22 Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQRPGQGLEWMGEILPGVVR TNYNENFRNKVTMTVDTSISTAYMELSRLRSDDTAVYYCTVVFYGEPYFPYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAV LQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPE AAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKP STYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY TLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSK LTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 129) >81B4vH50_30 Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQRPGQGLEWIGEINPGAVRT NYNENFRNRVTMTVDTSISTAYMELSRLRSDDTAVYYCTVVFYGEPYFPYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL SLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEA AGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPR EEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTL PPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 130) >81B4vH51_13 Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQAPGQGLEWIGEINPGLVRT NYNENFRNKVTMTVDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEA AGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPR EEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTL PPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 131) >81B4vH51_15 Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQAPGQGLEWIGEINPGAVRT NYNENFRNKVTMTVDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEA 2012/064933 AGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPR EEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTL MTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 132) >81B4vH52_83 Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYSFTSSWIHWVRQAPGQGLEWIGEINPGSVRT NYNENFRNKATMTVDTSISTAYMELSRLRSDDTAVYYCAVVFYGEPYFPYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEA FLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPR EEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTL PPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 133) >73C5VH46_4 Heavy Chain SGPGLVKPSETLSITCTVSGFSLTDYAVHWIRQPPGKGLEWIGVIWSDGSTDYN APFKSRVTINKDTSKSQVSFKMSSVQAADTAVYYCARKGGYSGSWFAYWGQGTLVTV SSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQ SSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAA GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREE QYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPP SREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 134) >73C5VH46_19 Heavy Chain QVQLQESGPGLVKPSETLSITCTVSGFSLTDYAVHWIRQPPGKGLEWIGVIWSDGSTDYN APFKSRVTISKDTSKNQVSLKMNSLTTDDTAVYYCARKGGYSGSWFAYWGQGTLVTV SSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQ SSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAA GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREE QYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPP SREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 135) >73C5VH46_40 Heavy Chain QVQLQESGPGLVKPSETLSITCTVSGFSLTDYAVHWIRQPPGKGLEWIGVIWSDGSTDYN APFKSRVTISKDNSKSQVSLKMNSVTVADTAVYYCARKGGYSGSWFAYWGQGTLVTV SSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQ SSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAA GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREE QYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPP SREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 136) >73C5VH47_65 Heavy Chain QVQLQESGPGLVKPSETLSITCTVSGFSLTDYAVHWVRQPPGKGLEWIGVIWSDGSTDY NAPFKSRVTISKDTSKNQVSFKLSSVTVDDTAVYYCARKGGYSGSWFAYWGQGTLVTV SSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQ LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAA GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREE QYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPP SREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 137) >73C5VH47_77 Heavy Chain QVQLQESGPGLVAPSETLSLTCTVSGFSLTDYAVHWIRQFPGKGLEWIGVIWSDGSTDF NAPFKSRVTISKDTSKNQVSFKLSSVTTDDTAVYYCARKGGYSGSWFAYWGQGTLVTV SSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQ SSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAA GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREE QYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPP SREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV QGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 138) >73C5VH58_91 Heavy Chain QVQLQESGPGLVKPSETLSITCTVSGFSLTDYAVHWIRQPPGKGLEWIGVIWSDGSTDYN APFKSRVTISKDNSKSQVSFKMSSVTADDTAVYYCARKGGYSGSWFAYWGQGTLVTVS SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQS SGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAAG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQ YNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPS REEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVD KSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 139) The CDRs listed above are defined using the Chothia ing system (Al-Lazikani et al., (1997) JMB 273, 927-948).

In one aspect, an antibody of the present invention comprises 3 light chain CDRs and 3 heavy chain CDRs, for example as set forth above.

In one aspect, an antibody of the present ion comprises a light chain and a heavy chain le region as set forth above. In one aspect, a light chain variable region of the invention is fused to a light chain constant region, for e a kappa or lambda nt region. In one aspect, a heavy chain variable region of the invention is fused to a heavy chain constant region, for example lgA, lgD, lgE, lgG or lgM, in particular, lgG1, lng, lgG3 or lgG4.

The t invention provides an L-36R antibody comprising a light chain comprising the amino acid sequence of SEQ ID NO: 115; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 125 (Antibody B1).

The present invention provides an anti-lL-36R antibody comprising a light chain comprising the amino acid sequence of SEQ ID NO: 115; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 126 (Antibody B2).

The present invention provides an anti-lL-36R antibody comprising a light chain comprising the amino acid sequence of SEQ ID NO: 115; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 127 (Antibody B3).

The present invention provides an anti-lL-36R antibody comprising a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 125 ody B4).

The present invention provides an anti-lL-36R antibody comprising a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 126 ody B5).

The present invention provides an anti-lL-36R antibody comprising a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid ce of SEQ ID NO: 127 Antibody B6).

WO 74569 The present invention provides an anti-lL-36R antibody sing a light chain comprising the amino acid sequence of SEQ ID NO: 123; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 138 (Antibody C3).

The present invention provides an anti-lL-36R antibody comprising a light chain comprising the amino acid sequence of SEQ ID NO: 123; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 139 (Antibody C2).

The present invention provides an anti-lL-36R antibody comprising a light chain comprising the amino acid sequence of SEQ ID NO: 124; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 138 (Antibody C1) Representative antibodies of the t invention are shown below.

Table A.

Anti Light Chain Sequences Heavy Chain Sequences body B1 SPGTLSLSPGERATMSCTASSS QVQLVQSGAEVKKPGASVKVSCKASGYSFTS VSSSYFHWYQQKPGQAPRLLIYRTSIL SWIHWVRQAPGQGLEWIGEINPGNVRTNYNE ASGVPDRFSGSGSGTDFTLTISRLEPE NFRNKATMTVDTSISTAYMELSRLRSDDTAV DFATYYCHQFHRSPLTFGQGTKLEIK YYCAVVFYGEPYFPYWGQGTLVTVSSASTK RTVAAPSVFIFPPSDEQLKSGTASVV GPSVFPLAPSSKSTSGGTAALGCLVKDYFPEP CLLNNFYPREAKVQWKVDNALQSGN SGALTSGVHTFPAVLQSSGLYSLSS SQESVTEQDSKDSTYSLSSTLTLSKA VVTVPSSSLGTQTYICNVNHKPSNTKVDKRV VYACEVTHQGLSSPVTKSF EPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKP NRGEC (SEQ ID NO: 115) KDTLMISRTPEVTCVVVDVSHEDPEVKFNWY VDGVEVHNAKTKPREEQYNSTYRVVSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKA KGQPREPQVYTLPPSREEMTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVLDSD GSFFLYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPGK (SEQ ID NO: 125) EIVLTQSPGTLSLSPGERATMSCTASSS QVQLVQSGAEVKKPGASVKVSCKASGYSFTS VSSSYFHWYQQKPGQAPRLLIYRTSIL SWIHWVRQRPGQGLEWIGEINPGNVRTNYNE RFSGSGSGTDFTLTISRLEPE NFRNRVTMTVDTSISTAYMELSRLRSDDTAV DFATYYCHQFHRSPLTFGQGTKLEIK YYCTVVFYGEPYFPYWGQGTLVTVSSASTKG RTVAAPSVFIFPPSDEQLKSGTASVV PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPV CLLNNFYPREAKVQWKVDNALQSGN TVSWNSGALTSGVHTFPAVLQSSGLYSLSSV SQESVTEQDSKDSTYSLSSTLTLSKA VTVPSSSLGTQTYICNVNHKPSNTKVDKRVEP DYEKHKVYACEVTHQGLSSPVTKSF HTCPPCPAPEAAGGPSVFLFPPKPKD NRGEC (SEQ ID NO: 115) TLMISRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPREEQYNSTYRVVSVLTVLH QDWLNGKEYKCKVSNKALPAPIEKTISKAKG VYTLPPSREEMTKNQVSLTCLVKGF YPSDIAVEWESNGQPENNYKTTPPVLDSDGSF FLYSKLTVDKSRWQQGNVFSCSVMHEALHN HYTQKSLSLSPGK (SEQ ID NO: 126) BS EIVLTQSPGTLSLSPGERATMSCTASSS QVQLVQSGAEVKKPGASVKVSCKASGYSFTS VSSSYFHWYQQKPGQAPRLLIYRTSIL SWIHWVKQAPGQGLEWMGEINPGNVRTNYN ASGVPDRFSGSGSGTDFTLTISRLEPE ENFRNKVTMTVDTSISTAYMELSRLRSDDTA DFATYYCHQFHRSPLTFGQGTKLEIK VYYCTVVFYGEPYFPYWGQGTLVTVSSASTK SVFIFPPSDEQLKSGTASVV GPSVFPLAPSSKSTSGGTAALGCLVKDYFPEP CLLNNFYPREAKVQWKVDNALQSGN VTVSWNSGALTSGVHTFPAVLQSSGLYSLSS SQESVTEQDSKDSTYSLSSTLTLSKA VVTVPSSSLGTQTYICNVNHKPSNTKVDKRV DYEKHKVYACEVTHQGLSSPVTKSF EPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKP NRGEC (SEQ ID NO: 115) KDTLMISRTPEVTCVVVDVSHEDPEVKFNWY VDGVEVHNAKTKPREEQYNSTYRVVSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKA KGQPREPQVYTLPPSREEMTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVLDSD SKLTVDKSRWQQGNVFSCSVMHEAL KSLSLSPGK (SEQ ID NO: 127) B4 QIVLTQSPGTLSLSPGERATMTCTASS QVQLVQSGAEVKKPGASVKVSCKASGYSFTS SVSSSYFHWYQQKPGQAPRLWIYRTS SWIHWVRQAPGQGLEWIGEINPGNVRTNYNE RLASGVPDRFSGSGSGTDFTLTISRLEP NFRNKATMTVDTSISTAYMELSRLRSDDTAV EDAATYYCHQFHRSPLTFGAGTKLEI YYCAVVFYGEPYFPYWGQGTLVTVSSASTK KRTVAAPSVF| FP PSDEQLKSGTASV GPSVFPLAPSSKSTSGGTAALGCLVKDYFPEP VCLLNNFYPREAKVQWKVDNALQSG VTVSWNSGALTSGVHTFPAVLQSSGLYSLSS NSQESVTEQDSKDSTYSLSSTLTLSK VVTVPSSSLGTQTYICNVNHKPSNTKVDKRV ADYEKHKVYACEVTHQGLSSPVTKS KTHTCPPCPAPEAAGGPSVFLFPPKP FNRGEC (SEQ ID NO: 118) KDTLMISRTPEVTCVVVDVSHEDPEVKFNWY VDGVEVHNAKTKPREEQYNSTYRVVSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKA KGQPREPQVYTLPPSREEMTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVLDSD GSFFLYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPGK (SEQ ID NO: 125) B5 QIVLTQSPGTLSLSPGERATMTCTASS QVQLVQSGAEVKKPGASVKVSCKASGYSFTS SVSSSYFHWYQQKPGQAPRLWIYRTS SWIHWVRQRPGQGLEWIGEINPGNVRTNYNE RLASGVPDRFSGSGSGTDFTLTISRLEP NFRNRVTMTVDTSISTAYMELSRLRSDDTAV EDAATYYCHQFHRSPLTFGAGTKLEI YYCTVVFYGEPYFPYWGQGTLVTVSSASTKG KRTVAAPSVF| FP PSDEQLKSGTASV PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPV VCLLNNFYPREAKVQWKVDNALQSG GALTSGVHTFPAVLQSSGLYSLSSV NSQESVTEQDSKDSTYSLSSTLTLSK VTVPSSSLGTQTYICNVNHKPSNTKVDKRVEP ADYEKHKVYACEVTHQGLSSPVTKS KSCDKTHTCPPCPAPEAAGGPSVFLFPPKPKD FNRGEC (SEQ ID NO: 118) TLMISRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPREEQYNSTYRVVSVLTVLH QDWLNGKEYKCKVSNKALPAPIEKTISKAKG QPREPQVYTLPPSREEMTKNQVSLTCLVKGF YPSDIAVEWESNGQPENNYKTTPPVLDSDGSF FLYSKLTVDKSRWQQGNVFSCSVMHEALHN HYTQKSLSLSPGK (SEQ ID NO: 126) SPGTLSLSPGERATMTCTASS QVQLVQSGAEVKKPGASVKVSCKASGYSFTS SVSSSYFHWYQQKPGQAPRLWIYRTS SWIHWVKQAPGQGLEWMGEINPGNVRTNYN RLASGVPDRFSGSGSGTDFTLTISRLEP ENFRNKVTMTVDTSISTAYMELSRLRSDDTA EDAATYYCHQFHRSPLTFGAGTKLEI VYYCTVVFYGEPYFPYWGQGTLVTVSSASTK KRTVAAPSVF| FP PSDEQLKSGTASV GPSVFPLAPSSKSTSGGTAALGCLVKDYFPEP VCLLNNFYPREAKVQWKVDNALQSG VTVSWNSGALTSGVHTFPAVLQSSGLYSLSS NSQESVTEQDSKDSTYSLSSTLTLSK VVTVPSSSLGTQTYICNVNHKPSNTKVDKRV KVYACEVTHQGLSSPVTKS EPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKP FNRGEC (SEQ ID NO: 118) KDTLMISRTPEVTCVVVDVSHEDPEVKFNWY VDGVEVHNAKTKPREEQYNSTYRVVSVLTV NGKEYKCKVSNKALPAPIEKTISKA KGQPREPQVYTLPPSREEMTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVLDSD GSFFLYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPGK (SEQ ID NO: 127) Table B Light Chain Seguences Heavy Chain Seguences EIVMTQSPATLSVSPGVRATLSCKASQ QVQLQESGPGLVAPSETLSLTCTVSGFSLTDY DVGTNVLWYQQKPGQAPRPLIYSASY AVHWIRQFPGKGLEWIGVIWSDGSTDFNAPF RHSGIPARFSGSGSGTEFTLTISSLQSE KSRVTISKDTSKNQVSFKLSSVTTDDTAVYYC CQQYSRYPLTFGQGTKLEIK ARKGGYSGSWFAYWGQGTLVTVSSASTKGP RTVAAPSVFIFPPSDEQLKSGTASVVC SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVT LLNNFYPREAKVQWKVDNALQSGNS VSWNSGALTSGVHTFPAVLQSSGLYSLSSVV QESVTEQDSKDSTYSLSSTLTLSKADY TVPSSSLGTQTYICNVNHKPSNTKVDKRVEPK ACEVTHQGLSSPVTKSFNRG SCDKTHTCPPCPAPEAAGGPSVFLFPPKPKDT EC (SEQ ID NO: 124) LMISRTPEVTCVVVDVSHEDPEVKFNWYVDG VEVHNAKTKPREEQYNSTYRVVSVLTVLHQ DWLNGKEYKCKVSNKALPAPIEKTISKAKGQ PREPQVYTLPPSREEMTKNQVSLTCLVKGFYP SDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHY TQKSLSLSPGK (SEQ ID NO: 138) EIVMTQSPATLSVSPGVRATLSCKASQ QVQLQESGPGLVKPSETLSITCTVSGFSLTDY DVGTNVLWYQQKPGQAPRPLIYSASY AVHWIRQPPGKGLEWIGVIWSDGSTDYNAPF RHSGIPDRFSGSGSGTEFTLTISSLQSE KSRVTISKDNSKSQVSFKMSSVTADDTAVYY CQQYSRYPLTFGQGTKLEIK CARKGGYSGSWFAYWGQGTLVTVSSASTKG RTVAAPSVFIFPPSDEQLKSGTASVVC PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPV LLNNFYPREAKVQWKVDNALQSGNS TVSWNSGALTSGVHTFPAVLQSSGLYSLSSV QESVTEQDSKDSTYSLSSTLTLSKADY VTVPSSSLGTQTYICNVNHKPSNTKVDKRVEP EKHKVYACEVTHQGLSSPVTKSFNRG KSCDKTHTCPPCPAPEAAGGPSVFLFPPKPKD EC (SEQ ID NO: 123) TPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPREEQYNSTYRVVSVLTVLH QDWLNGKEYKCKVSNKALPAPIEKTISKAKG QPREPQVYTLPPSREEMTKNQVSLTCLVKGF YPSDIAVEWESNGQPENNYKTTPPVLDSDGSF TVDKSRWQQGNVFSCSVMHEALHN HYTQKSLSLSPGK (SEQ ID NO: 139) EIVMTQSPATLSVSPGVRATLSCKASQ QVQLQESGPGLVAPSETLSLTCTVSGFSLTDY DVGTNVLWYQQKPGQAPRPLIYSASY AVHWIRQFPGKGLEWIGVIWSDGSTDFNAPF RHSGIPDRFSGSGSGTEFTLTISSLQSE KSRVTISKDTSKNQVSFKLSSVTTDDTAVYYC DFAVYYCQQYSRYPLTFGQGTKLEIK ARKGGYSGSWFAYWGQGTLVTVSSASTKGP RTVAAPSVFIFPPSDEQLKSGTASVVC SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVT LLNNFYPREAKVQWKVDNALQSGNS VSWNSGALTSGVHTFPAVLQSSGLYSLSSVV QESVTEQDSKDSTYSLSSTLTLSKADY TVPSSSLGTQTYICNVNHKPSNTKVDKRVEPK EKHKVYACEVTHQGLSSPVTKSFNRG SCDKTHTCPPCPAPEAAGGPSVFLFPPKPKDT EC (SEQ ID NO: 123) LMISRTPEVTCVVVDVSHEDPEVKFNWYVDG VEVHNAKTKPREEQYNSTYRVVSVLTVLHQ DWLNGKEYKCKVSNKALPAPIEKTISKAKGQ PREPQVYTLPPSREEMTKNQVSLTCLVKGFYP SDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHY TQKSLSLSPGK (SEQ ID NO: 138) The antibodies of the present invention are useful in methods for the treatment of various es or disorders, for e immunological, inflammatory, autoimmune diseases and respiratory diseases in . For example, the antibodies of the present invention are useful in methods for the treatment of psoriasis, rheumatoid arthritis, inflammatory bowel disease or psoriatic arthritis. For example, the dies of the present invention are useful in methods for the treatment of chronic obstructive pulmonary disorder (COPD) or asthma. For example, the antibodies of the present invention are useful in s for the treatment of scleroderma, palmoplantar pustulosis, generalized pustular psoriasis, diabetic nephropathy, lupus nephritis, scleroderma, ankylosing spondylitis, deficiency in the lL-36 receptor antagonist autoimmune disease (DITRA), deficiency in the lL-1 receptor nist autoimmune disease (DIRA) or cryopyrin associated periodic syndromes (CAPS).

In some aspects, the humanized antibody ys blocking activity, whereby it decreases the binding of lL-36 ligand to lL-36 receptor by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, or by at least 95%. The ability of an antibody to block binding of lL-36 ligand to the lL-36 receptor can be measured using competitive binding assays known in the art. Alternatively, the blocking activity of an antibody can be ed by ing the biological s of lL-36, such as the production of lL-8, lL-6, and GM-CSF to determine if ing mediated by the lL-36 receptor is inhibited.

In a further aspect, the present invention provides a humanized anti-lL-36R antibody having favorable biophysical properties. In one aspect, a humanized anti-lL-36R dy of the present invention is present in at least 90% monomer form, or in at least 92% monomer form, or in at least 95% monomer form in a buffer. In a further aspect, a humanized anti-lL-36R antibody of the present invention remains in at least 90% r form, or in at least 92% r form, or in at least 95% r form in a buffer for one month or for four months.

In one aspect, a humanized antibody of the t invention is Antibody B1, Antibody B2, Antibody B3, Antibody B4, Antibody B5, Antibody B6, dy C1, Antibody C2, or Antibody C3. Accordingly, in one embodiment, a humanized dy of the present invention comprises the light chain ce of SEQ ID NO:115 and the heavy chain sequence of SEQ ID NO:125 (Antibody B1). In another embodiment, a humanized dy of the present invention comprises the light chain sequence of SEQ ID NO:115 and the heavy chain sequence of SEQ ID NO:126 (Antibody B2). In another embodiment, a humanized antibody of the present invention comprises the light chain sequence of SEQ ID NO:115 and the heavy chain sequence of SEQ ID NO:127 (Antibody B3). In another embodiment, a zed antibody of the present invention comprises the light chain sequence of SEQ ID NO:118 and the heavy chain sequence of SEQ ID NO:125 ody B4). In another embodiment, a humanized antibody of the present invention comprises the light chain sequence of SEQ ID NO:118 and the heavy chain sequence of SEQ ID NO:126 (Antibody B5). In another ment, a humanized antibody of the present invention comprises the light chain sequence of SEQ ID NO:118 and the heavy chain sequence of SEQ ID NO:127 (Antibody B6). In another embodiment, a humanized antibody of the present invention comprises the light chain sequence of SEQ ID NO:124 and the heavy chain sequence of SEQ ID NO:138 (Antibody C1). In another embodiment, a humanized antibody of the present invention comprises the light chain sequence of SEQ ID NO:123 and the heavy chain sequence of SEQ ID NO:139 (Antibody C2). In another embodiment, a zed antibody of the present invention comprises the light chain sequence of SEQ ID NO:123 and the heavy chain sequence of SEQ ID NO:138 (Antibody C3).

In a further embodiment, a humanized antibody of the present invention consists of the light chain sequence of SEQ ID NO:115 and the heavy chain sequence of SEQ ID NO:125 (Antibody B1). In r embodiment, a humanized antibody of the present invention consists of the light chain sequence of SEQ ID NO:115 and the heavy chain sequence of SEQ ID NO:126 (Antibody B2). In r ment, a humanized antibody of the present invention consists of the light chain sequence of SEQ ID NO:115 and the heavy chain sequence of SEQ ID NO:127 (Antibody B3). In r embodiment, a humanized antibody of the present invention consists of the light chain sequence of SEQ ID NO:118 and the heavy chain sequence of SEQ ID NO:125 (Antibody B4). In r embodiment, a humanized antibody of the present ion ts of the light chain sequence of SEQ ID NO:118 and the heavy chain sequence of SEQ ID NO:126 (Antibody B5). In another embodiment, a humanized antibody of the present invention consists of the light chain sequence of SEQ ID NO:118 and the heavy chain sequence of SEQ ID NO:127 (Antibody B6). In another embodiment, a humanized antibody of the t invention consists of the light chain sequence of SEQ ID NO:124 and the heavy chain sequence of SEQ ID NO:138 ody C1). In another embodiment, a humanized antibody of the present invention consists of the light chain sequence of SEQ ID NO:123 and the heavy chain sequence of SEQ ID NO:139 (Antibody C2). In another embodiment, a humanized antibody of the present invention consists of the light chain sequence of SEQ ID NO:123 and the heavy chain sequence of SEQ ID NO:138 (Antibody 03).

In some embodiments, the humanized anti-IL-36B antibodies, including antigen-binding fragments thereof, such as heavy and light chain variable s, comprise an amino acid sequence of the residues derived from Antibody B1, Antibody B2, Antibody B3, Antibody B4, Antibody B5, Antibody B6, Antibody C1, Antibody C2, or Antibody 03.

In a further embodiment, the present invention provides an anti-IL-36B antibody or antigen-binding fragment thereof that competitively binds to human IL-36R with an antibody of the present invention, for example Antibody B1, Antibody B2, Antibody B3, Antibody B4, Antibody B5, Antibody B6, Antibody C1, dy C2 or Antibody 03 bed herein. The ability of an dy or antigen-binding fragment to itively bind to IL-36B can be ed using competitive binding assays known in the art.

The humanized anti-IL-36B antibodies optionally include specific amino acid substitutions in the consensus or germline framework regions. The specific substitution of amino acid residues in these ork positions can improve various aspects of antibody performance ing binding affinity and/or stability, over that demonstrated in humanized antibodies formed by "direct swap" of CDRs or HVLs into the human germline framework regions.

In some embodiments, the t invention describes other monoclonal antibodies with alight chain variable region having the amino acid sequence set forth in any one of SEQ ID NO:1-10. In some embodiments, the present invention describes other monoclonal antibodies with a heavy chain variable region having the amino acid ce set forth in any one of SEQ ID NO:11-20. Placing such CDRs into FRs of the human consensus heavy and light chain variable domains will yield useful humanized antibodies of the t invention.

In particular, the present invention provides monoclonal antibodies with the combinations of light chain variable and heavy chain variable regions of SEQ ID NO:1/11, 2/12, 3/13, 4/14, 5/15, 6/16, 7/17, 8/18, 9/19, 10/20. Such variable regions can be combined with human constant regions.

In some embodiments, the present invention describes other humanized antibodies with light chain variable region sequences having the amino acid ce set forth in any one of SEQ ID NQ:76-86. In some embodiments, the present invention describes other humanized antibodies with heavy chain le region sequences having the amino acid ce set forth in any one of SEQ ID NQ:87-101. In particular, the present invention provides monoclonal dies with the ations of light chain variable and heavy chain variable regions of SEQ ID NO: 77/89, 80/88, 80/89, 77/87, 77/88, 80/87, 86/100, 85/101, 85/100. Such variable regions can be combined with human constant regions.

In a r embodiment, the present invention relates to an anti-IL-36R antibody or antigen-binding fragment thereof comprising a humanized light chain variable domain comprising the CDRs of SEQ ID NQ:77 and framework regions having an amino acid sequence at least 90% identical, at least 93% cal or at least 95% identical to the amino acid sequence of the framework regions of the variable domain light chain amino acid sequence of SEQ ID NQ:77 and a humanized heavy chain le domain comprising the CDRs of SEQ ID NQ:89 and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain heavy chain amino acid ce of SEQ ID NQ:89. In one embodiment, the anti-IL-36R antibody is a humanized monoclonal antibody.

In a further embodiment, the present invention relates to an anti-IL-36R antibody or antigen-binding fragment f comprising a humanized light chain variable domain comprising the CDRs of SEQ ID NQ:8O and framework regions having an amino acid sequence at least 90% identical, at least 93% cal or at least 95% identical to the amino acid sequence of the framework regions of the variable domain light chain amino acid sequence of SEQ ID NQ:8O and a humanized heavy chain variable domain comprising the CDRs of SEQ ID NQ:88 and framework s having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain heavy chain amino acid sequence of SEQ ID NQ:88. In one embodiment, the L-36R antibody is a humanized monoclonal antibody.

In a further embodiment, the present invention relates to an anti-IL-36R antibody or antigen-binding fragment thereof comprising a humanized light chain variable domain comprising the CDRs of SEQ ID NQ:8O and ork regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework s of the variable domain light chain amino acid sequence of SEQ ID NQ:8O and a humanized heavy chain le domain comprising the CDRs of SEQ ID NQ:89 and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain heavy chain amino acid ce of SEQ ID NQ:89. In one embodiment, the anti-IL-36R dy is a humanized monoclonal antibody.

In a further embodiment, the present invention relates to an anti-IL-36R dy or antigen-binding fragment thereof comprising a humanized light chain variable domain comprising the CDRs of SEQ ID NQ:77 and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework s of the variable domain light chain amino acid sequence of SEQ ID NO:77 and a humanized heavy chain variable domain comprising the CDRs of SEQ ID NO:87 and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain heavy chain amino acid sequence of SEQ ID NO:87. In one embodiment, the anti-IL-36R antibody is a humanized monoclonal antibody.

In a further embodiment, the t invention relates to an anti-IL-36R antibody or antigen-binding fragment thereof comprising a humanized light chain variable domain sing the CDRs of SEQ ID NO:77 and framework s having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain light chain amino acid sequence of SEQ ID NO:77 and a humanized heavy chain variable domain comprising the CDRs of SEQ ID NO:88 and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain heavy chain amino acid sequence of SEQ ID NO:88. In one embodiment, the anti-IL-36R antibody is a humanized monoclonal antibody.

In a further embodiment, the present invention relates to an L-36R antibody or n-binding fragment thereof comprising a humanized light chain variable domain comprising the CDRs of SEQ ID NO:8O and framework regions having an amino acid sequence at least 90% cal, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the le domain light chain amino acid sequence of SEQ ID NO:8O and a humanized heavy chain variable domain comprising the CDRs of SEQ ID NO:87 and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% cal to the amino acid sequence of the framework regions of the variable domain heavy chain amino acid sequence of SEQ ID NO:87. In one embodiment, the anti-IL-36R dy is a humanized monoclonal antibody.

In a further embodiment, the t invention s to an anti-lL-36R antibody or antigen-binding fragment thereof sing a humanized light chain variable domain comprising the CDRs of SEQ ID NO:86 and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain light chain amino acid sequence of SEQ ID NO:86 and a humanized heavy chain variable domain comprising the CDRs of SEQ ID NQ:1OO and ork regions having an amino acid ce at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain heavy chain amino acid sequence of SEQ ID NQ:100. In one embodiment, the anti-lL-36R dy is a humanized monoclonal antibody.

In a r embodiment, the present invention relates to an anti-lL-36R antibody or antigen-binding fragment thereof comprising a zed light chain variable domain comprising the CDRs of SEQ ID NO:85 and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain light chain amino acid sequence of SEQ ID NO:85 and a humanized heavy chain variable domain comprising the CDRs of SEQ ID NQ:101 and framework regions having an amino acid ce at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain heavy chain amino acid sequence of SEQ ID NQ:101. In one embodiment, the anti-lL-36R antibody is a humanized monoclonal antibody.

In a further embodiment, the present invention relates to an L-36R antibody or antigen-binding fragment thereof comprising a humanized light chain variable domain comprising the CDRs of SEQ ID NO:85 and framework regions having an amino acid ce at least 90% identical, at least 93% identical or at least 95% cal to the amino acid ce of the framework regions of the variable domain light chain amino acid sequence of SEQ ID NO:85 and a humanized heavy chain variable domain comprising the CDRs of SEQ ID NQ:1OO and framework regions having an amino acid sequence at least 90% identical, at least 93% identical or at least 95% identical to the amino acid sequence of the framework regions of the variable domain heavy chain 2012/064933 amino acid sequence of SEQ ID NQ:100. In one embodiment, the anti-IL-36R dy is a humanized monoclonal dy.

In some specific embodiments, the humanized anti-IL-36R antibodies disclosed herein comprise at least a heavy or a light chain variable domain comprising the CDRs or HVLs of the murine onal antibodies or zed antibodies as disclosed herein and the PBS of the human ne heavy and light chain variable domains.

In one further aspect, the present invention provides an L-36R antibody or antigen- binding fragment thereof comprising a light chain CDR1 (L-CDR1) sequence of any one of SEQ ID NQ:21-29; a light chain CDR2 (L-CDR2) sequence of any one of SEQ ID NQ:30-38; a light chain CDR3 (L-CDR3) sequence of any one of SEQ ID NQ:39-47; a heavy chain CDR1 (H-CDR1) sequence of any one of SEQ ID NQ:48-56; a heavy chain CDR2 (H-CDR2) sequence of any one of SEQ ID NQ:57-66; and a heavy chain CDR3 (H-CDR3) ce of any one of SEQ ID NQ:67-75. In one aspect, the anti-IL-36R antibody or antigen-binding fragment thereof comprises a light chain variable region comprising a L-CDR1 listed above, a L-CDR2 listed above and a L-CDR3 listed above, and a heavy chain variable region comprising a H-CDR1 listed above, a H-CDR2 listed above and a H-CDR3 listed above.

In a further aspect, the present invention provides an anti-IL-36R antibody or antigen- binding nt thereof comprising: a) a L-CDR1, a L-CDR2, a L-CDR3, a , a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:21, 30, 39, 48, 57 and 67, respectively; or b) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:22, 31, 40, 49, 58 and 68, respectively; or c) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:23, 32, 41, 50, 59 and 69, respectively; or d) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:24, 33, 42, 51, 60 and 70, respectively; or e) a L-CDR1, a L-CDR2, a L-CDR3, a , a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:25, 34, 43, 52, 61 and 71, respectively; or f) a L-CDR1, a L-CDR2, a , a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:26, 35, 44, 53, 62 and 72, respectively; or g) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:27, 36, 45, 54, 63 and 73, respectively; or h) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:27, 36, 45, 54, 64 and 74, respectively; or i) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:27, 36, 45, 54, 64 and 73, respectively; or j) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:28, 37, 46, 55, 65 and 74, respectively; or k) a L-CDR1, a L-CDR2, a L-CDR3, a , a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:29, 38, 47, 56, 66 and 75, tively.

In a further aspect, the present invention es an anti-IL-36R dy or antigen- binding fragment thereof comprising: a) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:26, 103, 44, 53, 62 and 72, respectively; or b) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:26, 104, 44, 53, 62 and 72, respectively; or c) a L-CDR1, a L-CDR2, a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:27, 36, 45, 107, 63 and 73, tively; or d) a L-CDR1, a , a L-CDR3, a H-CDR1, a H-CDR2 and a H-CDR3 sequence of SEQ ID NQ:27, 36, 45, 107, 64 or 73, respectively.

In one aspect, the anti-IL-36R antibody or antigen-binding fragment f comprises a light chain variable region comprising a L-CDR1, L-CDR2 and L-CDR3 combination listed above, and a heavy chain variable region comprising a H-CDR1, H-CDR2 and H- CDR3 combination listed above.

In specific embodiments, it is plated that chimeric antibodies with switched CDR regions (i.e., for example switching one or two CDRs of one of the mouse antibodies or humanized antibody derived therefrom with the analogous CDR from another mouse antibody or humanized dy derived therefrom) between these exemplary immunoglobulins may yield useful antibodies.

In certain embodiments, the humanized anti-lL-36R antibody is an antibody fragment.

Various antibody nts have been generally discussed above and there are techniques that have been developed for the production of antibody fragments. nts can be d via proteolytic digestion of intact antibodies (see, e.g., Morimoto et al., 1992, Journal of Biochemical and sical Methods 24:107-117; and Brennan et al., 1985, Science 229:81). Alternatively, the fragments can be produced directly in recombinant host cells. For example, Fab'-SH fragments can be directly recovered from E. coli and chemically d to form F(ab')2 fragments (see, e.g., Carter et al., 1992, Bio/Technology 10:163-167). By another approach, F(ab')2 fragments can be ed directly from recombinant host cell e. Other techniques for the production of antibody fragments will be apparent to the skilled practitioner.

Accordingly, in one aspect, the present ion provides dy fragments comprising the CDRs described , in particular one of the combinations of L- CDR1, L-CDR2, L-CDR3, H-CDR1, H-CDR2 and H-CDR3 described herein. In a further aspect, the present invention provides antibody fragments comprising the variable regions described herein, for example one of the combinations of light chain variable regions and heavy chain variable regions described herein.

Certain embodiments include an F(ab')2 fragment of a humanized anti-lL-36R antibody comprise a light chain sequence of any of SEQ ID NO: 115 or 118 in combination with a heavy chain sequence of SEQ ID NO: 125, 126 or 127. Such embodiments can include an intact antibody comprising such an F(ab')2.

Certain embodiments include an F(ab')2 fragment of a humanized anti-lL-36R antibody comprise a light chain ce of any of SEQ ID NO: 123 or 124 in combination with a heavy chain sequence of SEQ ID NO: 138 or 139. Such embodiments can include an intact antibody sing such an F(ab')2.

In some embodiments, the antibody or antibody fragment includes a constant region that mediates effector function. The constant region can provide antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP) and/or complement-dependent cytotoxicity (CDC) responses against an lL-36R sing target cell. The or domain(s) can be, for example, an Fc region of an lg molecule.

The effector domain of an antibody can be from any suitable vertebrate animal species and isotypes. The isotypes from different animal species differ in the abilities to mediate effector functions. For e, the ability of human globulin to mediate CDC and ADCC/ADCP is generally in the order of lglegG1zlgG3>lgG2>lgG4 and lgG1zlgG3>lgG2/lgM/lgG4, respectively. Murine immunoglobulins mediate CDC and ADCC/ADCP generally in the order of murine lglegG3>>lgG2b>lgG2a>>lgG1 and lgG2a>lgG1>>lgG3, respectively. In r example, murine lnga mediates ADCC while both murine lnga and lgM mediate CDC.

Antibody Modifications The humanized L-36R antibodies and agents can include modifications of the humanized anti-lL-36R dy or antigen-binding fragment thereof. For example, it may be desirable to modify the dy with respect to effector function, so as to enhance the effectiveness of the antibody in treating cancer. One such modification is the introduction of cysteine residue(s) into the Fc region, thereby allowing interchain disulfide bond formation in this region. The homodimeric antibody thus generated can have improved internalization capability and/or increased complement-mediated cell g and/or dy-dependent cellular cytotoxicity (ADCC). See, for example, Caron et al., 1992, J. Exp Med. 176:1191-1195; and Shopes, 1992, J. Immunol. 148:2918- 2922. meric antibodies having enhanced umor activity can also be prepared using heterobifunctional cross-linkers as described in Wolff et al., 1993, Cancer Research 53: 2560-2565. Alternatively, an antibody can be engineered to contain dual Fc regions, enhancing complement lysis and ADCC capabilities of the antibody. See Stevenson et al., 1989, Anti-Cancer Drug Design 3: 219-230.

Antibodies with improved ability to support ADCC have been generated by modifying the glycosylation pattern of their Fc region. This is possible since antibody glycosylation at the asparagine residue, N297, in the CH2 domain is involved in the interaction between lgG and Fcy receptors prerequisite to ADCC. Host cell lines have been engineered to express antibodies with d glycosylation, such as increased bisecting N-acetylglucosamine or reduced fucose. Fucose reduction provides r enhancement to ADCC activity than does increasing the presence of bisecting N- acetylglucosamine. Moreover, enhancement of ADCC by low fucose dies is independent of the Fclella V/F rphism.

Modifying the amino acid sequence of the Fc region of antibodies is an alternative to glycosylation engineering to enhance ADCC. The binding site on human lgG1 for Fcy receptors has been ined by extensive mutational analysis. This led to the generation of humanized lgG1 antibodies with PC mutations that increase the binding affinity for Fclella and enhance ADCC in vitro. Additionally, Fc variants have been obtained with many different permutations of binding properties, e.g., improved binding to specific chR receptors with unchanged or diminished binding to other FcyR receptors.

Another aspect includes conjugates sing the humanized antibody or fragments thereof conjugated to a cytotoxic agent such as a chemotherapeutic agent, a toxin (e.g., an enzymatically active toxin of bacterial, fungal, plant, or animal origin, or fragments thereof), or a radioactive e (Le, a radioconjugate).

Chemotherapeutic agents useful in the generation of such immunoconjugates have been described above. tically active toxins and fragments thereof that can be used to form useful immunoconjugates include diphtheria A chain, nonbinding active fragments of diphtheria toxin, exotoxin A chain (from Pseudomonas aeruginosa), ricin A chain, abrin A chain, in A chain, alpha-sarcin, Aleurites fordii proteins, dianthin proteins, Phytolaca americana proteins (PAPI, PAP”, and PAP-S), Momordica tia inhibitor, curcin, crotin, Sapaonaria officinalis inhibitor, gelonin, mitogellin, restrictocin, phenomycin, enomycin, the tricothecenes, and the like. A variety of radionuclides are available for the tion of radioconjugated humanized anti-lL-36R antibodies. Examples include 212Bi, 131I, 131In, 90Y, and 186Re.

Conjugates of the zed anti-lL-36R antibody and cytotoxic or chemotherapeutic agent can be made by known s, using a variety of bifunctional protein coupling agents such as N-succinimidyl(2—pyridyldithiol) propionate (SPDP), iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HCL), active esters (such as disuccinimidyl suberate), aldehydes (such as glutareldehyde), ido compounds (such as bis (p-azidobenzoyl) hexanediamine), bis-diazonium derivatives (such as bis-(p-diazoniumbenzoyl)-ethylenediamine), yanates (such as toluene 2,6—diisocyanate), and tive fluorine compounds (such as 1,5-difluoro-2,4- dinitrobenzene). For example, a ricin immunotoxin can be prepared as described in Vitetta et al., 1987, Science 238:1098. Carbonlabeled 1-isothiocyanatobenzyl methyldiethylene triaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent for conjugation of radionucleotide to the antibody. Conjugates also can be formed with a cleavable linker.

The humanized L-36R antibodies disclosed herein can also be ated as immunoliposomes. Liposomes ning the antibody are prepared by s known in the art, such as described in Epstein et al., 1985, Proc. Natl. Acad. Sci. USA 82:3688; Hwang et al., 1980, Proc. Natl. Acad. Sci. USA 77:4030; and US. Pat. Nos. 4,485,045 and 4,544,545. Liposomes having ed circulation time are disclosed, for example, in US. Pat. No. 556.

Particularly useful liposomes can be generated by the reverse phase evaporation method with a lipid composition comprising atidylcholine, cholesterol and PEG- derivatized phosphatidylethanolamine (PEG-PE). Liposomes are extruded through filters of defined pore size to yield liposomes with the desired diameter. Fab' fragments of an antibody disclosed herein can be conjugated to the liposomes as described in Martin et al., 1982, J. Biol. Chem. 257:286—288 via a disulfide interchange reaction. A chemotherapeutic agent (such as doxorubicin) is optionally contained within the liposome. See, e.g., Gabizon et al., 1989, J. National Cancer Inst. 81(19):1484.

The antibodies described and disclosed herein can also be used in ADEPT (Antibody- Directed Enzyme Prodrug Therapy) procedures by conjugating the antibody to a prodrug-activating enzyme that converts a prodrug (e.g., a peptidyl chemotherapeutic agent), to an active anti-cancer drug. See, for e, WO 81/01145, WO 88/07378, and US. Pat. No. 4,975,278. The enzyme component of the immunoconjugate useful for ADEPT is an enzyme e of acting on a prodrug in such a way so as to covert it 2012/064933 into its more active, cytotoxic form. Specific enzymes that are useful in ADEPT e, but are not limited to, alkaline phosphatase for converting phosphate-containing prodrugs into free drugs; lfatase for converting sulfate-containing prodrugs into free drugs; cytosine deaminase for converting non-toxic 5-fluorocytosine into the anti- cancer drug, 5-fluorouracil; ses, such as serratia protease, thermolysin, subtilisin, carboxypeptidases, and cathepsins (such as cathepsins B and L), for converting peptide-containing prodrugs into free drugs; D-alanylcarboxypeptidases, for converting prodrugs containing D-amino acid substituents; carbohydrate-cleaving enzymes such as B-galactosidase and neuraminidase for converting glycosylated prodrugs into free drugs; B-lactamase for converting drugs tized with B-lactams into free drugs; and penicillin es, such as penicillin V amidase or penicillin G amidase, for converting drugs derivatized at their amine nitrogens with phenoxyacetyl or phenylacetyl groups, respectively, into free drugs. Alternatively, antibodies having enzymatic activity ("abzymes") can be used to convert the prodrugs into free active drugs (see, for example, Massey, 1987, Nature 328: 457-458). Antibody-abzyme conjugates can be prepared by known methods for delivery of the abzyme to a tumor cell tion, for example, by covalently binding the enzyme to the humanized anti-lL-36R antibody/heterobifunctional crosslinking ts discussed above. Alternatively, fusion ns comprising at least the antigen binding region of an dy sed herein linked to at least a functionally active portion of an enzyme as described above can be ucted using recombinant DNA techniques (see, e.g., Neuberger et al., 1984, Nature 312:604-608).

In certain embodiments, it may be desirable to use a humanized anti-lL-36R antibody nt, rather than an intact antibody, to increase tissue penetration, for example. It may be desirable to modify the antibody fragment in order to increase its serum half life.

This can be achieved, for example, by incorporation of a salvage receptor binding epitope into the antibody fragment. In one method, the appropriate region of the antibody fragment can be altered (e.g., mutated), or the epitope can be incorporated into a peptide tag that is then fused to the antibody fragment at either end or in the middle, for e, by DNA or peptide synthesis. See, e.g., WO 96/32478.

In other ments, covalent modifications of the humanized anti-lL-36R antibody are also included. Covalent modifications include modification of nyl residues, histidyl residues, lysinyl and amino-terminal residues, arginyl residues, tyrosyl residues, carboxyl side groups (aspartyl or glutamyl), glutaminyl and asparaginyl es, or seryl, or threonyl residues. Another type of covalent modification involves chemically or enzymatically coupling glycosides to the antibody. Such modifications may be made by chemical synthesis or by enzymatic or chemical cleavage of the antibody, if applicable.

Other types of covalent modifications of the antibody can be introduced into the le by reacting targeted amino acid residues of the antibody with an organic derivatizing agent that is capable of reacting with selected side chains or the amino- or carboxy-terminal residues. l of any carbohydrate moieties present on the antibody can be accomplished chemically or enzymatically. al deglycosylation is described by Hakimuddin et al., 1987, Arch. Biochem. Biophys. 259:52 and by Edge et al., 1981, Anal. Biochem., 118:131. Enzymatic cleavage of carbohydrate moieties on antibodies can be achieved by the use of a variety of endo- and exo-glycosidases as described by Thotakura et al., 1987, Meth. Enzymol 138:350.

Another type of useful covalent modification comprises g the dy to one of a variety of nonproteinaceous polymers, e.g., hylene glycol, polypropylene glycol, or polyoxyalkylenes, in the manner set forth in one or more of US. Pat. No. 835, US Pat. No. 4,496,689, US. Pat. No. 4,301,144, US. Pat. No. 4,670,417, US. Pat.

No. 4,791,192 and US. Pat. No. 337.

Humanization and Amino Acid Sequence Variants Amino acid sequence variants of the anti-lL-36R antibody can be prepared by introducing appropriate nucleotide changes into the anti-lL-36R dy DNA, or by e synthesis. Such variants include, for example, deletions from, and/or insertions into and/or substitutions of, es within the amino acid sequences of the anti-lL-36R antibodies of the examples herein. Any combination of deletions, insertions, and substitutions is made to arrive at the final construct, provided that the final construct possesses the desired characteristics. The amino acid changes also may alter post- translational processes of the humanized or variant anti-lL-36R dy, such as changing the number or position of ylation sites.

A useful method for fication of certain residues or regions of the anti-lL-36R antibody that are preferred locations for mutagenesis is called "alanine scanning mutagenesis," as described by Cunningham and Wells (Science, 244:1081-1085 (1989)). Here, a residue or group of target es are identified (e.g., charged residues such as arg, asp, his, lys, and glu) and replaced by a neutral or negatively d amino acid (typically alanine) to affect the interaction of the amino acids with lL-36R antigen. Those amino acid locations demonstrating functional sensitivity to the substitutions then are d by ucing further or other variants at, or for, the sites of substitution. Thus, while the site for introducing an amino acid sequence variation is predetermined, the nature of the mutation per se need not be ermined. For e, to analyze the performance of a mutation at a given site, alanine scanning or random nesis is conducted at the target codon or region and the expressed anti- lL-36R antibody variants are screened for the desired activity.

Amino acid ce insertions include amino- and/or carboxyl-terminal fusions ranging in length from one e to polypeptides containing a hundred or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Examples of terminal insertions include an anti-lL-36R antibody fused to an epitope tag. Other insertional variants of the anti-lL-36R antibody molecule include a fusion to the N- or C- terminus of the anti-lL-36R antibody of an enzyme or a polypeptide which ses the serum half-life of the antibody.

Another type of variant is an amino acid substitution variant. These variants have at least one amino acid residue in the anti-lL-36R antibody molecule removed and a different residue inserted in its place. The sites of greatest interest for substitutional nesis include the hypervariable regions, but FR alterations are also contemplated. Conservative substitutions are shown in Table 5 under the heading of "preferred substitutions". If such substitutions result in a change in biological activity, then more substantial changes, denominated "exemplary substitutions", or as further described below in reference to amino acid classes, may be introduced and the products screened.

TABLE C: Original Residue Exemplary Substitutions Preferred Substitutions Ala (A) val; leu; ile val Arg (R) lys; gln; asn lys Asn (N) gln; his; asp, lys; arg gin Asp (D) glu; asn glu Cys (C) ser; ala ser Gin (Q) asn; glu asn Glu (E) asp; gln asp Gly (G) ala ala His (H) arg; asn; gln; lys; arg lie (I) leu; val; met; ala; phe; norleucine leu Leu (L) ile; norleucine; val; met; ala; phe ile Lys (K) arg; gln; asn arg Met (M) leu; phe; ile leu Phe (F) tyr; leu; val; ile; ala; tyr Pro (P) ala ala Ser (8) thr thr Thr (T) ser ser Trp (W) tyr; phe tyr Tyr (Y) p; thr; ser phe Val (V) leu; ile; met; phe ala; norleucine; leu In protein chemistry, it is generally accepted that the ical properties of the antibody can be accomplished by selecting substitutions that differ significantly in their effect on maintaining (a) the structure of the polypeptide backbone in the area of the substitution, for example, as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain.

Naturally occurring residues are d into groups based on common side-chain properties: (1) hydrophobic: norleucine, met, ala, val, leu, lie; (2) neutral hydrophilic: cys, ser, thr; (3) acidic: asp, glu; (4) basic: asn, gin, his, lys, arg; (5) residues that nce chain orientation: gly, pro; and (6) aromatic: trp, tyr, phe.

Non-conservative substitutions will entail exchanging a member of one of these s for another class.

Any cysteine residue not involved in maintaining the proper conformation of the humanized or variant anti-lL-36R antibody also may be substituted, generally with serine, to e the ive stability of the molecule, prevent aberrant crosslinking, or provide for established points of conjugation to a cytotoxic or cytostatic compound.

Conversely, cysteine bond(s) may be added to the antibody to improve its stability (particularly where the antibody is an antibody fragment such as an Fv fragment).

A type of substitutional variant involves substituting one or more hypervariable region residues of a parent antibody (e.g., a humanized or human antibody). Generally, the resulting variant(s) selected for further development will have improved biological properties ve to the parent antibody from which they are generated. A ient way for ting such substitutional variants is ty maturation using phage display. Briefly, several hypervariable region sites (e.g., 6-7 sites) are mutated to generate all possible amino substitutions at each site. The antibody variants thus generated are displayed in a lent fashion from filamentous phage particles as fusions to the gene lll product of M13 packaged within each particle. The phage- displayed variants are then ed for their biological activity (e.g., binding affinity). In order to identify candidate hypervariable region sites for modification, alanine ng mutagenesis can be performed to fy hypervariable region residues contributing significantly to antigen g. Alternatively, or in addition, it may be beneficial to analyze a crystal structure of the n-antibody complex to identify contact points between the dy and human lL-36R. Such contact residues and neighboring residues are candidates for substitution according to the techniques elaborated herein.

Once such variants are generated, the panel of variants is subjected to screening as described herein and antibodies with superior properties in one or more nt assays may be selected for r development.

Another type of amino acid t of the antibody alters the original glycosylation n of the antibody. By "altering" is meant deleting one or more carbohydrate moieties found in the antibody, and/or adding one or more glycosylation sites that are not present in the antibody.

In some embodiments, it may be desirable to modify the antibodies of the ion to add glycosylations sites. Glycosylation of antibodies is typically either N-linked or O- linked. N-linked refers to the attachment of the carbohydrate moiety to the side chain of an asparagine residue. The tripeptide sequences asparagine-X-serine and asparagine- X-threonine, where X is any amino acid except proline, are the recognition sequences for enzymatic attachment of the carbohydrate moiety to the asparagine side chain.

Thus, the presence of either of these tripeptide ces in a polypeptide creates a potential glycosylation site. O-linked ylation refers to the attachment of one of the sugars N-aceylgalactosamine, galactose, or xylose to a hydroxyamino acid, most commonly serine or threonine, although 5-hydroxyproline or 5-hydroxylysine may also be used. Thus, in order to glycosylate a given protein, e.g., an antibody, the amino acid sequence of the protein is engineered to contain one or more of the above-described tripeptide sequences (for N-linked glycosylation sites). The alteration may also be made by the addition of, or substitution by, one or more serine or threonine residues to the sequence of the original antibody (for O-linked glycosylation sites).

Nucleic acid les ng amino acid sequence variants of the anti-IL-36R antibody are prepared by a variety of methods known in the art. These methods include, but are not limited to, isolation from a natural source (in the case of naturally occurring amino acid sequence variants) or preparation by oligonucleotide-mediated (or site- ed) mutagenesis, PCR mutagenesis, and cassette mutagenesis of an earlier prepared variant or a non-variant n of the anti-IL-36R antibody.

Polynucleotides, Vectors, Host Cells, and Recombinant Methods Other embodiments encompass isolated polynucleotides that comprise a sequence encoding a humanized anti-IL-36R antibody, vectors, and host cells comprising the polynucleotides, and recombinant techniques for production of the humanized antibody.

The isolated polynucleotides can encode any desired form of the anti-IL-36R antibody including, for example, full length monoclonal antibodies, Fab, Fab', F(ab')2, and Fv fragments, diabodies, linear antibodies, single-chain antibody molecules, and multispecific antibodies formed from dy fragments.

Some embodiments include isolated polynucleotides comprising sequences that encode the light chain variable region of an antibody or antibody fragment having the amino acid sequence of any of SEQ ID NO: SEQ ID 0. Some embodiments include isolated polynucleotides comprising sequences that encode the heavy chain variable region of an antibody or dy fragment having the amino acid sequence of SEQ ID Some embodiments include ed polynucleotides comprising ces that encode the light chain variable region of an antibody or antibody fragment having the amino acid sequence of any of SEQ ID NQ:76-86. Some embodiments include isolated polynucleotides comprising ces that encode the heavy chain variable region of an antibody or antibody fragment having the amino acid ce of SEQ ID NO: 87- 101.

Some embodiments include isolated polynucleotides comprising sequences that encode the light chain of an dy having the amino acid sequence of any of SEQ ID NO:114-124. Some embodiments include isolated cleotides comprising sequences that encode the heavy chain of an antibody having the amino acid sequence of SEQ ID NO:125-139.

In one aspect, the isolated polynucleotide sequence(s) encodes an antibody or antibody fragment having a light chain and a heavy chain variable region sing the amino acid sequences of SEQ ID NO:115 and SEQ ID NO:127, respectively; SEQ ID NO:118 and SEQ ID NO:126, respectively; SEQ ID NO:118 and SEQ ID NO:127, respectively; SEQ ID NO:115 and SEQ ID NO:125, respectively; SEQ ID NO:115 and SEQ ID NO:126, respectively; SEQ ID NO:118 and SEQ ID NO:125, respectively; SEQ ID NO:124 and SEQ ID , tively; SEQ ID NO:123 and SEQ ID NO:139, tively; SEQ ID NO:123 and SEQ ID NO:138, respectively.

The polynucleotide(s) that comprise a sequence encoding a humanized anti-lL-36R antibody or a fragment or chain thereof can be fused to one or more regulatory or control sequence, as known in the art, and can be contained in suitable expression vectors or host cell as known in the art. Each of the polynucleotide molecules encoding the heavy or light chain variable domains can be ndently fused to a polynucleotide sequence ng a constant domain, such as a human constant domain, enabling the production of intact antibodies. Alternatively, polynucleotides, or ns thereof, can be fused together, providing a template for production of a single chain antibody.

For recombinant tion, a polynucleotide encoding the antibody is inserted into a replicable vector for cloning (amplification of the DNA) or for sion. Many suitable vectors for expressing the recombinant antibody are available. The vector components generally include, but are not limited to, one or more of the following: a signal sequence, an origin of replication, one or more marker genes, an enhancer element, a promoter, and a transcription termination sequence.

The humanized anti-lL-36R antibodies can also be produced as fusion ptides, in which the antibody is fused with a heterologous polypeptide, such as a signal sequence or other polypeptide having a specific cleavage site at the amino terminus of the mature protein or polypeptide. The heterologous signal sequence selected is typically one that is recognized and processed (i.e., cleaved by a signal ase) by the host cell. For prokaryotic host cells that do not recognize and process the humanized anti-lL-36R antibody signal sequence, the signal sequence can be substituted by a prokaryotic signal sequence. The signal sequence can be, for example, alkaline phosphatase, penicillinase, otein, heat-stable enterotoxin ll leaders, and the like. For yeast ion, the native signal sequence can be substituted, for example, with a leader sequence obtained from yeast invertase alpha-factor (including romyces and Kluyveromyces or leaders), acid phosphatase, C. albicans glucoamylase, or the signal described in 3646. In mammalian cells, mammalian signal sequences as well as viral secretory leaders, for example, the herpes simplex gD signal, can be used.

The DNA for such precursor region is ligated in reading frame to DNA encoding the humanized anti-lL-36R antibody.

Expression and cloning vectors contain a nucleic acid sequence that enables the vector to replicate in one or more selected host cells. Generally, in cloning vectors this sequence is one that enables the vector to replicate independently of the host somal DNA, and includes origins of replication or autonomously replicating sequences. Such sequences are well known for a variety of bacteria, yeast, and viruses. The origin of ation from the plasmid pBR322 is suitable for most Gram- negative bacteria, the 2—1). plasmid origin is le for yeast, and various viral origins (SV40, polyoma, adenovirus, VSV, and BPV) are useful for cloning vectors in mammalian cells. Generally, the origin of replication component is not needed for mammalian expression vectors (the SV4O origin may typically be used only because it contains the early promoter).

Expression and cloning vectors may contain a gene that encodes a selectable marker to facilitate fication of expression. Typical selectable marker genes encode proteins that confer resistance to antibiotics or other toxins, e.g., ampicillin, in, methotrexate, or tetracycline, or alternatively, are ment auxotrophic deficiencies, or in other atives supply specific nutrients that are not present in complex media, e.g., the gene ng D-alanine racemase for Bacilli.

One example of a selection scheme es a drug to arrest growth of a host cell. Those cells that are successfully transformed with a heterologous gene produce a n conferring drug ance and thus survive the selection regimen. Examples of such dominant selection use the drugs neomycin, mycophenolic acid, and hygromycin.

Common selectable markers for mammalian cells are those that enable the fication of cells competent to take up a nucleic acid encoding a humanized anti-IL- 36R antibody, such as DHFR (dihydrofolate reductase), thymidine kinase, metallothionein-l and -II (such as primate metallothionein genes), adenosine deaminase, ornithine decarboxylase, and the like. Cells transformed with the DHFR selection gene are first identified by culturing all of the transformants in a culture medium that contains methotrexate (Mtx), a competitive antagonist of DHFR. An appropriate host cell when wild-type DHFR is employed is the Chinese hamster ovary (CHO) cell line deficient in DHFR activity (e.g., DG44).

Alternatively, host cells (particularly wild-type hosts that contain endogenous DHFR) ormed or co-transformed with DNA sequences encoding anti-lL-36R antibody, wild-type DHFR protein, and another selectable marker such as aminoglycoside 3'- phosphotransferase (APH), can be selected by cell growth in medium containing a ion agent for the selectable marker such as an aminoglycosidic otic, e.g., kanamycin, neomycin, or G418. See, e.g., U.S. Pat. No. 4,965,199.

Where the inant production is performed in a yeast cell as a host cell, the TRP1 gene present in the yeast plasmid YRp7 (Stinchcomb et al., 1979, Nature 282: 39) can be used as a selectable marker. The TRP1 gene provides a ion marker for a mutant strain of yeast lacking the ability to grow in tryptophan, for example, ATCC No. 44076 or PEP4-1 (Jones, 1977, Genetics 85:12). The presence of the trp1 lesion in the yeast host cell genome then provides an effective environment for detecting transformation by growth in the absence of phan. Similarly, Leu2p-deficient yeast strains such as ATCC 20,622 and 38,626 are complemented by known plasmids bearing the LEU2 gene.

In addition, vectors derived from the 1.6 pm ar plasmid pKD1 can be used for transformation of Kluyveromyces . Alternatively, an expression system for large- scale production of recombinant calf chymosin was ed for K. lactis (Van den Berg, 1990, Bio/Technology 8:135). Stable multi-copy expression vectors for secretion of mature recombinant human serum albumin by industrial strains of Kluyveromyces have also been disclosed (Fleer et al., 1991, Bio/Technology 9:968—975).

Expression and cloning vectors usually contain a promoter that is recognized by the host organism and is operably linked to the nucleic acid molecule ng an anti-IL- 36R antibody or polypeptide chain thereof. Promoters suitable for use with prokaryotic hosts include phoA promoter, B-lactamase and lactose promoter systems, alkaline phosphatase, tryptophan (trp) promoter system, and hybrid promoters such as the tac promoter. Other known bacterial promoters are also suitable. Promoters for use in bacterial systems also will contain a Shine-Dalgarno (S.D.) sequence operably linked to the DNA encoding the humanized L-36R antibody.

Many eukaryotic promoter sequences are known. Virtually all eukaryotic genes have an AT-rich region d imately 25 to 30 bases upstream from the site where ription is initiated. Another sequence found 70 to 80 bases upstream from the start of transcription of many genes is a CNCAAT region where N may be any nucleotide. At the 3' end of most eukaryotic genes is an AATAAA sequence that may be the signal for addition of the poly A tail to the 3' end of the coding sequence. All of these sequences are suitably inserted into eukaryotic expression vectors.

Examples of suitable promoting sequences for use with yeast hosts include the promoters for 3-phosphoglycerate kinase or other glycolytic s, such as enolase, glyceraldehydephosphate dehydrogenase, hexokinase, te decarboxylase, ofructokinase, glucosephosphate isomerase, 3-phosphoglycerate mutase, pyruvate kinase, triosephosphate isomerase, phosphoglucose isomerase, and inase. lnducible ers have the additional advantage of transcription controlled by growth conditions. These include yeast promoter s for alcohol dehydrogenase 2, isocytochrome C, acid phosphatase, derivative enzymes ated with nitrogen lism, metallothionein, glyceraldehydephosphate dehydrogenase, and enzymes responsible for maltose and galactose utilization. Suitable vectors and promoters for use in yeast expression are further described in EP . Yeast enhancers also are advantageously used with yeast promoters.

Humanized anti-lL-36R antibody transcription from vectors in mammalian host cells is controlled, for example, by promoters obtained from the genomes of viruses such as polyoma virus, fowlpox virus, adenovirus (such as Adenovirus 2), bovine papilloma virus, avian sarcoma virus, cytomegalovirus, a retrovirus, hepatitis-B virus and Simian Virus 40 (SV40), from heterologous mammalian promoters, e.g., the actin promoter or an immunoglobulin promoter, or from heat-shock promoters, provided such ers are compatible with the host cell systems.

The early and late promoters of the SV40 virus are conveniently obtained as an SV4O restriction fragment that also contains the SV40 viral origin of replication. The ate early promoter of the human cytomegalovirus is conveniently obtained as a Hindlll E restriction fragment. A system for expressing DNA in mammalian hosts using the bovine papilloma virus as a vector is disclosed in U.S. Pat. No. 4,419,446. A modification of this system is described in U.S. Pat. No. 4,601,978. See also Reyes et al., 1982, Nature 297:598—601, disclosing expression of human p-interferon cDNA in mouse cells under the control of a thymidine kinase promoter from herpes simplex virus. Alternatively, the Rous a virus long terminal repeat can be used as the promoter.

Another useful element that can be used in a recombinant expression vector is an enhancer sequence, which is used to se the transcription of a DNA encoding a humanized anti-lL-36R antibody by higher eukaryotes. Many enhancer sequences are now known from ian genes (e.g., globin, elastase, albumin, d-fetoprotein, and insulin). Typically, however, an enhancer from a otic cell virus is used. es include the SV40 enhancer on the late side of the replication origin (bp 0), the cytomegalovirus early er enhancer, the polyoma enhancer on the late side of the replication origin, and adenovirus enhancers. See also Yaniv, 1982, Nature -18 for a description of ing elements for activation of eukaryotic ers. The enhancer may be spliced into the vector at a position 5' or 3' to the humanized anti-IL- 36R antibody-encoding sequence, but is preferably located at a site 5' from the promoter. 2012/064933 Expression vectors used in eukaryotic host cells (yeast, fungi, insect, plant, animal, human, or nucleated cells from other multicellular organisms) can also contain sequences necessary for the termination of transcription and for stabilizing the mRNA.

Such sequences are commonly available from the 5' and, occasionally 3', untranslated s of eukaryotic or viral DNAs or cDNAs. These regions contain nucleotide segments ribed as polyadenylated nts in the untranslated portion of the mRNA encoding anti-lL-36R antibody. One useful transcription termination component is the bovine growth hormone polyadenylation region. See WO94/11026 and the expression vector disclosed therein. In some embodiments, humanized anti-lL-36R antibodies can be expressed using the CHEF system. (See, e.g., US. Pat. No. ,888,809; the disclosure of which is incorporated by reference herein.) Suitable host cells for cloning or expressing the DNA in the vectors herein are the prokaryote, yeast, or higher ote cells described above. le prokaryotes for this purpose include eubacteria, such as Gram-negative or Gram-positive sms, for example, Enterobacteriaceae such as Escherichia, e.g., E. coli, Enterobacter, EnNinia, Klebsiella, Proteus, Salmonella, e.g., ella typhimurium, Serratia, e.g., Serratia marcescans, and Shigella, as well as Bacilli such as B. subtilis and B. licheniformis (e.g., B. licheniformis 41 P disclosed in DD 266,710 published Apr. 12, 1989), Pseudomonas such as P. aeruginosa, and omyces. One preferred E. coli cloning host is E. coli 294 (ATCC 31,446), although other strains such as E. coli B, E. coli X1776 (ATCC 31,537), and E. coli W3110 (ATCC 27,325) are suitable. These examples are illustrative rather than limiting.

In on to prokaryotes, eukaryotic microbes such as filamentous fungi or yeast are le g or expression hosts for humanized anti-lL-36Rantibody-encoding vectors. Saccharomyces cerevisiae, or common baker's yeast, is the most commonly used among lower eukaryotic host microorganisms. However, a number of other genera, species, and strains are commonly available and useful herein, such as Schizosaccharomyces pombe; Kluyveromyces hosts such as, e.g., K. lactis, K. fragilis (ATCC 12,424), K. bulgaricus (ATCC 16,045), K. wickeramii (ATCC 24,178), K. waltii (ATCC 56,500), K. drosophilarum (ATCC 36,906), K. thermotolerans, and K. marxianus; yarrowia (EP 402,226); Pichia pastors (EP 183,070); a; Trichoderma reesia (EP 244,234); Neurospora crassa; Schwanniomyces such as Schwanniomyces occidentalis; and filamentous fungi such as, e.g., Neurospora, Penicillium, Tolypocladium, and Aspergillus hosts such as A. nidulans and A. niger.

Suitable host cells for the expression of glycosylated humanized anti-lL-36R antibody are derived from multicellular organisms. Examples of invertebrate cells include plant and insect cells, including, e.g., numerous baculoviral strains and variants and corresponding permissive insect host cells from hosts such as Spodoptera frugiperda (caterpillar), Aedes aegypti (mosquito), Aedes albopictus (mosquito), Drosophila melanogaster (fruitfly), and Bombyx mori (silk worm). A variety of viral s for transfection are ly available, e.g., the L-1 variant of Autographa californica NPV and the Bm-5 strain of Bombyx mori NPV, and such viruses may be used, particularly for transfection of Spodoptera frugiperda cells.

Plant cell es of cotton, corn, potato, soybean, petunia, tomato, and tobacco can also be utilized as hosts.

In another aspect, expression of humanized anti-lL-36R is carried out in vertebrate cells. The propagation of vertebrate cells in culture (tissue e) has become routine procedure and techniques are widely ble. es of useful mammalian host cell lines are monkey kidney CV1 line transformed by SV40 (COS-7, ATCC CRL 1651), human embryonic kidney line (293 or 293 cells subcloned for growth in suspension e, (Graham et al., 1977, J. Gen Virol. 36: 59), baby r kidney cells (BHK, ATCC CCL 10), Chinese hamster ovary cells/-DHFR1 (CHO, Urlaub et al., 1980, Proc.

Natl. Acad. Sci. USA 77: 4216; e.g., DG44), mouse sertoli cells (TM4, Mather, 1980, Biol. Reprod. 23:243-251), monkey kidney cells (CV1 ATCC CCL 70), African green monkey kidney cells (VERO-76, ATCC CRL-1587), human al oma cells (HELA, ATCC CCL 2), canine kidney cells (MDCK, ATCC CCL 34), buffalo rat liver cells (BRL 3A, ATCC CRL 1442), human lung cells (W138, ATCC CCL 75), human liver cells (Hep G2, HB 8065), mouse mammary tumor (MMT 060562, ATCC CCL51), TR1 cells (Mather et al., 1982, Annals NY. Acad. Sci. 383: 44-68), MRC 5 cells, FS4 cells, and human hepatoma line (Hep G2).

Host cells are transformed with the above-described expression or cloning vectors for humanized anti-lL-36R antibody production and ed in conventional nutrient media modified as appropriate for inducing promoters, selecting transformants, or amplifying the genes encoding the desired sequences.

The host cells used to produce a humanized L-36R antibody described herein may be cultured in a variety of media. Commercially available media such as Ham's F10 (Sigma-Aldrich Co., St. Louis, Mo.), Minimal Essential Medium , (Sigma-Aldrich Co.), RPMl-1640 (Sigma-Aldrich Co.), and Dulbecco's Modified Eagle's Medium ((DMEM), Sigma-Aldrich Co.) are suitable for culturing the host cells. In addition, any of the media described in one or more of Ham et al., 1979, Meth. Enz. 58: 44, Barnes et al., 1980, Anal. Biochem. 102: 255, U.S. Pat. No. 4,767,704, U.S. Pat. No. 4,657,866, U.S. Pat. No. 4,927,762, U.S. Pat. No. 4,560,655, U.S. Pat. No. 5,122,469, WO 90/103430, and WO 87/00195 may be used as culture media for the host cells. Any of these media may be supplemented as ary with hormones and/or other growth factors (such as insulin, errin, or epidermal growth factor), salts (such as sodium chloride, calcium, magnesium, and phosphate), buffers (such as HEPES), nucleotides (such as adenosine and thymidine), antibiotics (such as gentamicin), trace elements (defined as inorganic compounds usually present at final concentrations in the micromolar range), and glucose or an equivalent energy source. Other supplements may also be included at appropriate concentrations that would be known to those skilled in the art. The e conditions, such as temperature, pH, and the like, are those usly used with the host cell selected for expression, and will be apparent to the ordinarily skilled n.

When using recombinant techniques, the antibody can be produced intracellularly, in the periplasmic space, or directly secreted into the medium. If the dy is ed intracellularly, the cells may be ted to release protein as a first step. Particulate debris, either host cells or lysed fragments, can be removed, for e, by centrifugation or ultrafiltration. Carter et al., 1992, Bio/Technology 10:163-167 describes a procedure for isolating antibodies that are secreted to the asmic space of E. coli.

Briefly, cell paste is thawed in the presence of sodium acetate (pH 3.5), EDTA, and phenylmethylsulfonylfluoride (PMSF) over about 30 minutes. Cell debris can be WO 74569 removed by centrifugation. Where the antibody is secreted into the medium, supernatants from such expression systems are generally first concentrated using a commercially available protein concentration filter, for example, an Amicon or Millipore Pellicon iltration unit. A protease inhibitor such as PMSF may be ed in any of the foregoing steps to inhibit proteolysis and antibiotics may be included to prevent the growth of adventitious contaminants. A variety of methods can be used to e the antibody from the host cell.

The antibody composition ed from the cells can be purified using, for example, hydroxylapatite chromatography, gel electrophoresis, dialysis, and affinity chromatography, with affinity chromatography being a typical purification technique. The suitability of protein A as an affinity ligand depends on the s and isotype of any immunoglobulin Fc domain that is t in the antibody. Protein A can be used to purify antibodies that are based on human gamma1, gamma2, or gamma4 heavy chains (see, e.g., Lindmark et al., 1983 J. lmmunol. Meth. 62:1-13). Protein G is recommended for all mouse es and for human gamma3 (see, e.g., Guss et al., 1986 EMBO J. 5:1567-1575). A matrix to which an affinity ligand is attached is most often agarose, but other matrices are available. Mechanically stable matrices such as controlled pore glass or poly(styrenedivinyl)benzene allow for faster flow rates and shorter processing times than can be achieved with agarose. Where the antibody ses a CH3 domain, the Bakerbond ABXTM resin (J. T. Baker, Phillipsburg, NJ.) is useful for purification. Other techniques for protein purification such as fractionation on an ion-exchange column, ethanol precipitation, reverse phase HPLC, tography on silica, tography on heparin SEPHAROSETM chromatography on an anion or cation exchange resin (such as a polyaspartic acid column), chromatofocusing, SDS— PAGE, and ammonium sulfate precipitation are also available depending on the antibody to be recovered.

Following any preliminary cation step(s), the mixture comprising the dy of interest and contaminants may be subjected to low pH hydrophobic interaction chromatography using an elution buffer at a pH between about 2.5-4.5, typically performed at low salt concentrations (e.g., from about 0-0.25M salt).

Also included are nucleic acids that hybridize under low, moderate, and high stringency conditions, as d herein, to all or a portion (e.g., the portion encoding the variable ) of the nucleotide sequence represented by isolated polynucleotide ce(s) that encode an antibody or antibody fragment of the present invention.The hybridizing portion of the hybridizing nucleic acid is typically at least 15 (e.g., 20, 25, 30 or 50) nucleotides in length. The hybridizing portion of the izing nucleic acid is at least 80%, e.g., at least 90%, at least 95%, or at least 98%, identical to the sequence of a portion or all of a nucleic acid ng an anti-lL-36R polypeptide (e.g., a heavy chain or light chain variable region), or its complement. Hybridizing nucleic acids of the type described herein can be used, for e, as a cloning probe, a primer, e.g., a PCR , or a diagnostic probe.

Non-Therapeutic Uses The antibodies described herein are useful as affinity purification agents. In this process, the antibodies are immobilized on a solid phase such a Protein A resin, using methods well known in the art. The immobilized antibody is contacted with a sample containing the lL-36R protein (or fragment thereof) to be ed, and thereafter the support is washed with a suitable solvent that will remove substantially all the material in the sample except the lL-36R n, which is bound to the immobilized antibody. y, the t is washed with another suitable solvent that will release the lL-36R protein from the antibody.

Anti-lL-36R antibodies, for example humanized anti-lL-36R antibodies, are also useful in diagnostic assays to detect and/or quantify lL-36R protein, for example, detecting lL- 36R expression in specific cells, tissues, or serum. The anti-lL-36R dies can be used diagnostically to, for example, monitor the development or progression of a disease as part of a clinical testing procedure to, e.g., determine the cy of a given treatment and/or prevention regimen. Detection can be tated by coupling the anti- lL-36R antibody. Examples of detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials, bioluminescent materials, radioactive materials, positron emitting metals using various positron emission tomographies, and nonradioactive paramagnetic metal ions. See, for example, U.S. Patent No. 4,741,900 for metal ions which can be conjugated to antibodies for use as diagnostics according to the present ion.

The anti-lL-36R antibodies can be used in methods for diagnosing an lL-36R- associated disorder (e.g., a disorder characterized by abnormal expression of lL-36R) or to determine if a subject has an increased risk of ping an lL-36R-associated disorder. Such methods include ting a biological sample from a subject with an lL-36R antibody and detecting binding of the antibody to lL-36R. By "biological sample" is intended any biological sample obtained from an individual, cell line, tissue culture, or other source of cells potentially sing lL-36R. Methods for obtaining tissue biopsies and body fluids from mammals are well known in the art.

In some embodiments, the method can further comprise comparing the level of lL-36R in a patient sample to a control sample (e.g., a t that does not have an - associated er) to determine if the patient has an lL-36R-associated er or is at risk of developing an lL-36R-associated disorder.

It will be advantageous in some ments, for example, for diagnostic purposes to label the antibody with a detectable moiety. Numerous detectable labels are available, including radioisotopes, scent labels, enzyme substrate labels and the like. The label may be indirectly conjugated with the dy using various known techniques.

For example, the antibody can be conjugated with biotin and any of the three broad categories of labels mentioned above can be conjugated with avidin, or vice versa.

Biotin binds selectively to avidin and thus, the label can be conjugated with the antibody in this indirect manner. Alternatively, to achieve indirect conjugation of the label with the antibody, the antibody can be conjugated with a small hapten (such as digoxin) and one of the different types of labels mentioned above is conjugated with an anti-hapten antibody (e.g., anti-digoxin antibody). Thus, indirect conjugation of the label with the antibody can be achieved.

Exemplary sotopes labels include 358, 14C, 125l, 3H, and 131l. The dy can be labeled with the radioisotope, using the ques described in, for example, Current Protocols in Immunology, Volumes 1 and 2, 1991, Coligen et al., Ed. Wiley-lnterscience, New York, N.Y., Pubs. Radioactivity can be measured, for example, by scintillation couang.

Exemplary fluorescent labels include labels d from rare earth chelates (europium es) or fluorescein and its derivatives, rhodamine and its derivatives, dansyl, Lissamine, phycoerythrin, and Texas Red are available. The fluorescent labels can be conjugated to the antibody via known techniques, such as those disclosed in Current Protocols in Immunology, for example. Fluorescence can be quantified using a fluorimeter.

There are various well-characterized enzyme-substrate labels known in the art (see, e.g., U.S. Pat. No. 4,275,149 for a review). The enzyme generally catalyzes a al alteration of the chromogenic substrate that can be measured using various techniques.

For example, alteration may be a color change in a ate that can be ed spectrophotometrically. Alternatively, the enzyme may alter the fluorescence or chemiluminescence of the substrate. Techniques for fying a change in fluorescence are described above. The chemiluminescent substrate becomes electronically excited by a chemical reaction and may then emit light that can be measured, using a chemiluminometer, for example, or donates energy to a fluorescent acceptor.

Examples of enzymatic labels include luciferases such as firefly luciferase and bacterial luciferase (U.S. Pat. No. 4,737,456), luciferin, 2,3-dihydrophthalazinediones, malate dehydrogenase, urease, peroxidase such as horseradish dase (HRPO), alkaline phosphatase, B-galactosidase, glucoamylase, lysozyme, saccharide es (such as glucose e, galactose oxidase, and glucosephosphate dehydrogenase), heterocydic oxidases (such as uricase and xanthine oxidase), lactoperoxidase, eroxidase, and the like. Techniques for conjugating enzymes to dies are described, for example, in O'Sullivan et al., 1981, Methods for the Preparation of Enzyme-Antibody Conjugates for use in Enzyme Immunoassay, in Methods in Enzym.

(J. Langone & H. Van s, eds.), Academic press, N.Y., 73: 147-166.

Examples of enzyme-substrate combinations include, for example: Horseradish dase (HRPO) with hydrogen peroxidase as a substrate, wherein the hydrogen peroxidase oxidizes a dye precursor such as orthophenylene diamine (OPD) or 3,3',5,5'-tetramethyl benzidine hydrochloride (TMB); alkaline phosphatase (AP) with para-Nitrophenyl phosphate as chromogenic substrate; and B-D-galactosidase (B-D- Gal) with a chromogenic ate such as p-nitrophenyl-B-D-galactosidase or fluorogenic substrate 4-methylumbelliferyl-B-D-galactosidase.

Numerous other enzyme-substrate combinations are available to those skilled in the art.

For a general review of these, see U.S. Pat. No. 4,275,149 and U.S. Pat. No. 4,318,980.

In another embodiment, the humanized anti-lL-36R antibody is used unlabeled and ed with a labeled antibody that binds the humanized anti-lL-36R dy.

The antibodies described herein may be employed in any known assay method, such as itive binding assays, direct and indirect sandwich assays, and immunoprecipitation assays. See, e.g., Zola, Monoclonal Antibodies: A Manual of Techniques, pp. 147-158 (CRC Press, Inc. 1987).

The anti-lL-36R antibody or antigen g fragment thereof can be used to inhibit the binding of ligand to the lL-36 receptor. Such s comprise stering an anti- lL-36R antibody or antigen binding fragment thereof to a cell (e.g., a mammalian cell) or cellular environment, whereby signaling mediated by the lL-36 receptor is inhibited.

These s can be performed in vitro or in vivo. By lar environment” is intended the tissue, medium, or extracellular matrix surrounding a cell. The L-36R antibody or antigen binding fragment thereof is administered to the cellular nment of a cell in such a manner that the antibody or fragment is capable of binding to lL-36R molecules outside of and surrounding the cell, therefore, preventing the binding of lL-36 ligand to its or.

Diagnostic Kits An anti-lL-36R antibody can be used in a diagnostic kit, Le, a packaged combination of reagents in predetermined amounts with instructions for ming the diagnostic assay. Where the antibody is labeled with an enzyme, the kit may include substrates WO 74569 and cofactors required by the enzyme such as a ate precursor that provides the detectable chromophore or fluorophore. In addition, other additives may be included such as stabilizers, buffers (for example a block buffer or lysis buffer), and the like. The relative amounts of the various reagents may be varied widely to provide for concentrations in solution of the reagents that substantially optimize the sensitivity of the assay. The reagents may be provided as dry powders, usually lyophilized, including excipients that on dissolution will provide a reagent solution having the appropriate concentration.

Therapeutic Uses In another embodiment, a humanized anti-lL-36R antibody disclosed herein is useful in the treatment of various ers associated with the expression of lL-36R as described herein. Methods for treating an lL-36R associated disorder comprise administering a therapeutically effective amount of a humanized anti-lL-36R antibody to a t in need f.

The humanized anti-lL-36R antibody or agent is administered by any suitable means, including parenteral, subcutaneous, intraperitoneal, intrapulmonary, and asal, and, if desired for local immunosuppressive treatment, intralesional stration ding perfusing or otherwise contacting the graft with the antibody before transplantation). The humanized anti-lL-36R antibody or agent can be administered, for e, as an infusion or as a bolus. Parenteral infusions e uscular, intravenous, intraarterial, eritoneal, or subcutaneous administration. In addition, the humanized anti-lL-36R antibody is suitably administered by pulse infusion, particularly with declining doses of the antibody. In one , the dosing is given by injections, most preferably intravenous or subcutaneous injections, depending in part on whether the administration is brief or chronic.

For the prevention or treatment of disease, the appropriate dosage of antibody will depend on a variety of factors such as the type of disease to be treated, as defined above, the severity and course of the disease, whether the antibody is administered for preventive or therapeutic purposes, previous therapy, the patients clinical history and response to the antibody, and the discretion of the attending physician. The dy is suitably administered to the patient at one time or over a series of treatments.

Depending on the type and severity of the disease, about 1 ug/kg to 20 mg/kg (e.g., 0.1- mg/kg) of antibody is an initial candidate dosage for stration to the patient, whether, for example, by one or more separate administrations, or by continuous infusion. A typical daily dosage might range from about 1 ug/kg to 100 mg/kg or more, depending on the factors mentioned above. For repeated administrations over several days or longer, depending on the condition, the treatment is ned until a desired ssion of disease symptoms occurs. However, other dosage regimens may be . The progress of this therapy is easily monitored by conventional techniques and assays. An ary dosing regimen is that disclosed in WO 94/04188.

The term “suppression” is used herein in the same context as “amelioration” and “alleviation” to mean a lessening of one or more characteristics of the disease.

The antibody composition will be formulated, dosed, and administered in a fashion consistent with good medical practice. Factors for consideration in this context include the particular disorder being treated, the ular mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the site of delivery of the agent, the method of administration, the scheduling of administration, and other factors known to medical practitioners. The "therapeutically effective amount" of the antibody to be administered will be governed by such considerations, and is the minimum amount necessary to prevent, ameliorate, or treat the disorder associated with lL-36R expression.

The antibody need not be, but is ally, formulated with one or more agents currently used to prevent or treat the er in on. The effective amount of such other agents depends on the amount of humanized anti-lL-36R23p19 antibody present in the formulation, the type of disorder or treatment, and other factors discussed above.

These are generally used in the same dosages and with stration routes as used hereinbefore or about from 1 to 99% of the heretofore employed dosages. ceutical Compositions and Administration Thereof A composition comprising an IL-36R binding agent (e.g., an anti-IL-36R antibody) can be administered to a subject having or at risk of having an immunological disorder, respiratory disorder or a cancer. The invention further provides for the use of a IL-36R binding agent (e.g., an anti-IL-36R antibody) in the cture of a medicament for tion or treatment of a cancer , atory disorder or immunological disorder.

The term ct" as used herein means any mammalian patient to which an IL- 36Rbinding agent can be administered, including, e.g., humans and non-human mammals, such as primates, rodents, and dogs. Subjects specifically intended for treatment using the methods described herein include humans. The antibodies or agents can be administered either alone or in combination with other compositions in the prevention or treatment of the immunological disorder, respiratory disorder or cancer. Such compositions which can be administered in ation with the antibodies or agents include methotrexate (MTX) and immunomodulators, e.g. antibodies or small molecules.

Examples of antibodies for use in such ceutical compositions are those that comprise a antibody or antibody fragment having the light chain variable region amino acid sequence of any of SEQ ID NO: 1-10. Examples of antibodies for use in such pharmaceutical compositions are also those that comprise a zed antibody or antibody fragment having the heavy chain variable region amino acid sequence of any of SEQ ID NO:11-20. r es of antibodies for use in such pharmaceutical compositions are also those that comprise a humanized antibody or antibody fragment having the light chain variable region amino acid sequence of any of SEQ ID NQ:76-86. Preferred antibodies for use in such pharmaceutical compositions are also those that comprise a humanized antibody or antibody fragment having the heavy chain variable region amino acid sequence of any of SEQ ID NO:87-101.

Further es of antibodies for use in such pharmaceutical compositions are also those that comprise a humanized antibody or antibody fragment having the light chain le region and heavy chain variable region of any of SEQ ID NO: 77 and 89, SEQ ID NO: 80 and 88, SEQ ID NO: 80 and 89, SEQ ID NO: 77 and 87, SEQ ID NO: 77 and 88, SEQ ID NO: 80 and 87, SEQ ID NO: 86 and 100, SEQ ID NO: 85 and 101, or SEQ ID NO: 85 and 10.

Further examples of antibodies for use in such pharmaceutical compositions are also those that comprise a humanized antibody having the light chain region amino acid sequence of any of SEQ ID NO:115, 118, 123 or 124. Preferred dies for use in such pharmaceutical compositions are also those that comprise humanized dy having the heavy chain variable region amino acid sequence of any of SEQ ID NO:125, 126,127,138 or 139.

Further examples of antibodies for use in such pharmaceutical compositions are also those that comprise Antibody B1, Antibody B2, Antibody B3, Antibody B4, Antibody B5, Antibody B6, Antibody C1, dy C2 or dy C3.

Various delivery systems are known and can be used to administer the lL-36R g agent. Methods of introduction include but are not limited to intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The IL- 36R binding agent can be stered, for example by infusion, bolus or injection, and can be administered together with other biologically active agents such as herapeutic . Administration can be systemic or local. In preferred embodiments, the administration is by subcutaneous injection. Formulations for such injections may be prepared in for example prefilled syringes that may be administered once every other week.

In specific embodiments, the lL-36R binding agent composition is administered by injection, by means of a catheter, by means of a suppository, or by means of an implant, the implant being of a porous, non-porous, or gelatinous material, including a membrane, such as a sialastic ne, or a fiber. Typically, when administering the composition, materials to which the anti-lL-36R antibody or agent does not absorb are used.

In other embodiments, the anti-lL-36R antibody or agent is red in a controlled release system. In one embodiment, a pump may be used (see, e.g., Langer, 1990, Science 249:1527-1533; Sefton, 1989, CRC Crit. Ref. Biomed. Eng. 14:201; Buchwald et al., 1980, Surgery 88:507; Saudek et al., 1989, N. Engl. J. Med. 321 :574). In another embodiment, polymeric als can be used. (See, e.g., Medical Applications of Controlled Release (Langer and Wise eds., CRC Press, Boca Raton, Fla., 1974); lled Drug Bioavailability, Drug Product Design and Performance (Smolen and Ball eds., Wiley, New York, 1984); Ranger and Peppas, 1983, Macromol. Sci. Rev.

Macromol. Chem. 23:61. See also Levy et al., 1985, Science 228:190; During et al., 1989, Ann. Neurol. 25:351; Howard et al., 1989, J. urg. 71:105.) Other controlled release systems are discussed, for example, in Langer, supra.

An lL-36R binding agent (e.g., an L-36R antibody) can be administered as ceutical compositions comprising a therapeutically effective amount of the binding agent and one or more pharmaceutically ible ingredients.

In typical embodiments, the pharmaceutical composition is formulated in accordance with routine procedures as a pharmaceutical composition adapted for intravenous or subcutaneous administration to human beings. Typically, compositions for administration by injection are solutions in sterile ic s . Where necessary, the pharmaceutical can also include a solubilizing agent and a local anesthetic such as lignocaine to ease pain at the site of the injection. Generally, the ingredients are supplied either separately or mixed together in unit dosage form, for example, as a dry lyophilized powder or water free concentrate in a hermetically sealed container such as an ampoule or sachette indicating the quantity of active agent. Where the ceutical is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water or saline. Where the pharmaceutical is administered by injection, an ampoule of sterile water for injection or saline can be provided so that the ingredients can be mixed prior to administration.

Further, the pharmaceutical composition can be ed as a pharmaceutical kit comprising (a) a container containing a lL-36R binding agent (e.g., an anti-lL-36R antibody) in lyophilized form and (b) a second container containing a ceutically acceptable diluent (e.g., sterile water) for injection. The pharmaceutically acceptable diluent can be used for reconstitution or dilution of the lyophilized L-36R antibody or agent. Optionally associated with such container(s) can be a notice in the form ibed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects al by the agency of manufacture, use or sale for human administration.

The amount of the lL-36R binding agent (e.g., anti-lL-36R antibody) that is ive in the treatment or prevention of an immunological disorder or cancer can be ined by standard clinical techniques. In addition, in vitro assays may optionally be employed to help identify optimal dosage ranges. The precise dose to be employed in the formulation will also depend on the route of administration, and the stage of immunological disorder or cancer, and should be decided according to the judgment of the practitioner and each patient's circumstances. Effective doses may be extrapolated from dose-response curves derived from in vitro or animal model test systems.

Generally, the dosage of an anti-lL-36R dy or lL-36R g agent administered to a patient with an immunological disorder or lL-36R-expressing cancer is typically about 0.1 mg/kg to about 100 mg/kg of the subject's body weight. The dosage administered to a subject is about 0.1 mg/kg to about 50 mg/kg, about 1 mg/kg to about mg/kg, about 1 mg/kg to about 20 mg/kg, about 1 mg/kg to about 15 mg/kg, or about 1 mg/kg to about 10 mg/kg of the t's body weight.

Exemplary doses include, but are not limited to, from 1 ng/kg to 100 mg/kg. In some embodiments, a dose is about 0.5 mg/kg, about 1 mg/kg, about 2 mg/kg, about 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about 6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9 mg/kg, about 10 mg/kg, about 11 mg/kg, about 12 mg/kg, about 13 mg/kg, about 14 mg/kg, about 15 mg/kg or about 16 mg/kg. The dose can be administered, for example, daily, once per week (weekly), twice per week, thrice per week, four times per week, five times per week, six times per week, biweekly or monthly, every two months, or every three months. In specific embodiments, the dose is about 0.5 mg/kg/week, about 1 mg/kg/week, about 2 mg/kg/week, about 3 mg/kg/week, about 4 mg/kg/week, about 5 mg/kg/week, about 6 week, about 7 mg/kg/week, about 8 mg/kg/week, about 9 mg/kg/week, about 10 mg/kg/week, about 11 mg/kg/week, about 12 mg/kg/week, about 13 mg/kg/week, about 14 mg/kg/week, about 15 week or about 16 mg/kg/week.

In some embodiments, the dose ranges from about 1 mg/kg/week to about 15 mg/kg/week.

In some embodiments, the pharmaceutical compositions comprising the lL-36R binding agent can further se a therapeutic agent, either conjugated or unconjugated to the binding agent. The anti-lL-36R antibody or lL-36R g agent can be co- administered in combination with one or more eutic agents for the treatment or prevention of immunological disorders or cancers.

Such combination therapy administration can have an additive or synergistic effect on disease ters (e.g., severity of a symptom, the number of symptoms, or frequency of relapse).

With respect to therapeutic regimens for combinatorial administration, in a specific embodiment, an anti-lL-36R antibody or lL-36R g agent is administered concurrently with a therapeutic agent. In another specific embodiment, the therapeutic agent is administered prior or subsequent to administration of the L-36R antibody or lL-36R binding agent, by at least an hour and up to several months, for example at least an hour, five hours, 12 hours, a day, a week, a month, or three months, prior or subsequent to administration of the anti-lL-36R antibody or lL-36R binding agent.

Articles of Manufacture In another aspect, an article of manufacture containing materials useful for the treatment of the disorders described above is ed. The article of manufacture ses a container and a label. Suitable containers include, for example, bottles, vials, syringes, and test tubes. The containers may be formed from a variety of als such as glass or plastic. The container holds a composition that is effective for ng the condition and may have a sterile access port. For example, the container may be an enous on bag or a vial having a stopper pierceable by a hypodermic injection needle. The active agent in the composition is the humanized anti- lL-36R antibody. The label on or associated with the container indicates that the composition is used for treating the condition of choice. The article of manufacture may WO 74569 further comprise a second container comprising a pharmaceutically-acceptable buffer, such as phosphate-buffered saline, Ringer's solution, and dextrose solution. It may further e other materials desirable from a commercial and user standpoint, including other s, diluents, filters, needles, syringes, and package inserts with instructions for use.

The invention is further described in the following examples, which are not intended to limit the scope of the ion.

The antibodies of the present invention are further described in the Examples below.

Examples e 1: Identification of Anti-human |L-36R Antibodies Multiple mouse strains were immunized with recombinantly produced human lL-36R (ECD — extracellular domain: amino acids 20-332 of Genbank Accession #NP_OO3845) protein and those which generated a strong titer response taken into traditional hybridoma generation. Fusion products eliciting a strong g to human lL-36R (ECD) yet no binding to human lL-1R1 (the most d lL-1R family member) were ned and re-screened. Multiple hybridomas were identified to yield monoclonal antibodies which bound and lized signaling from lL-36R (see examples 2, 3, and 4). Variable domains were cloned from the hybridomas using standard PCR primer sets. The variable domains and specific CDRs of representative monoclonal antibodies are described above. All of the mouse antibodies were converted to chimeric antibodies consisting of the mouse variable domains on human constant domains (hu lthKO / kappa). The hu lngKO (Knock gut) has two replacement ons (Leu234Ala and Leu235Ala) that eliminate ADCC and CDC activity by reducing effector functions such as FcyR and complement binding. The variable domains of the mouse and chimeric antibodies are identical. Chimeric antibodies are ted to confirm the WO 74569 function of the antibody and to ensure the correct variable domain sequence has been obtained.

Example 2: Molecular g Affinities of Identified Mouse Anti-human |L-36R dies A) Kinetics and binding affinities of anti-lL-36R antibodies binding to recombinant human lL-36R were measured using the Proteon (Bio-Rad, Hercules, CA) using material generated from hybridoma following single column purification. The binding ties of all the mouse leads to human lL-36R run at a single lL-36R surface coat concentration was estimated to be . The binding affinities of the mouse antibodies to human lL-36R run at 7 different surface densities (globally fit) is shown in Table 1. Binding of the chimeric anti-lL36R lgGs is equivalent to the respective mouse leads.

Table 1: Binding affinity of mouse anti-human lL-36R antibodies.

Lead Binding KD (pM) Mouse 7305 57 Mouse 73F6 25 Mouse 7808 63 Mouse 81A1 16 Mouse 81 B4 24 Mouse 33D10 9 B) Molecular selectivity over mouse lL-36R and human lL-1R1 The mouse and chimeric L-36R antibodies were also injected over either a mouse lL-36R or a human lL-1R1 surface at a tration of 100 nM. The binding signal to mouse lL-36R and to human lL-1R1 for these antibodies measured using the Fortebio Octet (Fortebio, Menlo Park, CA) is zero, which indicates these antibodies selectively bind to human lL-36R. The binding of the anti-lL-36R antibodies to human lL-36R was 2012/064933 also analyzed in the ce of 50% human serum and no significant effect of serum on binding on-rate was observed demonstrating high specificity.

Example 3: y of mouse and chimeric anti-human |L-36R antibodies in functional human and cynomolgus assays Protocols: Human NCI/ADR-RES cells QNFKB/Cytokine release assa vs Reagents: R&D Systems: truncated rh IL36B R&D Systems: truncated rh lL36y R&D Systems: truncated rh lL36d: MA6000 Phospho- NFKB (Ser536) Whole Cell Lysate Kit Meso Scale Diagnostics, LLC MSD ELISA custom human 96-well 4-spot assay Meso Scale Diagnostics, LLC NCI/ADR-RES cells were plated at 45000 cells/well, in RPMI media with 0.25% serum in a 96well plate. One platewas utilized for the analysis of pNFKB and another for cytokine release. The plates were then incubated overnight at 37°C, 5% C02 Ligands (lL360c, B, or y) and antibodies were diluted at 4x desired concentration in serum starved (SS) media. Antagonists odies) were added to cells prior to . For pNFKB: NCI cells +/- ligand and nist were incubated for 1 hour, 37°C, 5% C02 Media was then aspirated and cells were lysed in 100 ul/well Complete lysis Buffer on ice 30min. Lysate was then centrifuged at 2500RPM, 20min, 4°C, and transferred to an MSD ELISA plate and assayed for pNFKB as per the manufacturer’s protocol. For Cytokine release: 18-24 hours after stimulation, supernatants were transferred to an MSD ELISA plate and assayed for cytokine as per manufacturer’s protocol.

Protocol: QNFKB (S5362 MSD ELISA for BaF/3 Cynomolgus lL-36R Cells BaF/3 cynomolgus lL-36R cells were plated at 90,000 cells/well in 88 media in a 96well plate 100 ul media was added to control wells. Antagonists (antibodies) were diluted at 4x d concentration and 50 ul was added to each well. Ligands (lL36d, B, or 7) were diluted at 4x desired tration in 88 media and Soul were added to each well (for final volume of 200 pl). Plates were incubated for 15 min, 37°C, 5% COZ Plates were fuged briefly, media was aspirated and cells were lysed in 100 ul/well Complete Lysis Buffer (see MSD pNFKB protocol) and incubated on ice 30min. Lysates were then centrifuged at 2500RPM, 20min, 4°C and transferred to an MSD ELISA plate.

Lysates were then evaluated for pNFKB activity using the MSD kit as described above. s: ngo results for mouse anti-human lL-36R antibodies in human functional assays (pNFKB and ne e) and a cynomolgus functional assay (pNFkB) with human lL-36 ligands are shown in Table 2. IC90 results for chimeric anti-human lL- 36R dies in human functional assays (pNFKB and cytokine release) and a cynomolgus functional assay (pNFKB) with human lL-36 ligands are shown in Table 3.

Table 2: Potency of mouse antibodies in functional cell assays (isotype controls demonstrated no inhibition of activity at highest concentration tested for samples) IIIIIIIIIII7376E RES 33D10 73C5 8 8 8D1 1D1 E11 B8 B12 8 “III-”Emutrun-lL-36a ----ImmllnflflIC90(nM) 1.3 3.4 3 4.2 1.4 ----III----fltrun-lL-36g ”"flfllflflflfltrun-lL-36a ---EIMII-flflfltrun-lL-36b ----I“mflflfltrun-lL-36g ”Illa-mun"trun-lL-36a 1 1 1 trun-IL-36b ND ND ND |c90 (nM) o .5 3.8 5.1 ND 5.8 5.3 0.3 III-mun“trun-IL-36g ”In-Inn“trun-IL-36a III-IIHMHHHtrun-IL-36b III-“MIHHHtrun-IL-36g IIIIII 36a -----E----pNFKB |C90 (nM) >4000 0.2 0.5 00 1 2.4 >4000 >4000 >4000 -trun-IL-geb -----m----|C90 (nM) >333 1.6 0.7 3 2.1 3.5 >333 >333 >333 -trun-IL-geg --H-------|C90 (nM) >333 1.5 3 1.5 1 5.1 >333 >333 >333 Table 3: Potency of chimeric antibodies in functional cell assays (isotype controls demonstrated no inhibition of activity at highest concentration tested for samples) NCI/ADR-RES— C33D10 C73C5 C3134 IIItrun-IL-36a |C90 13 III“trun-IL-36b |C90 -—--ntrun-IL-36g |C90 IIII“trun-IL-36a |C90 IIII“trun-IL-36b |C90 IIII“trun-IL-36g |C90 IIII“trun-IL-36a |C90 IIII“trun-IL-36b |C90 III“trun-IL-36g |C90 run-IL-36a |C90 IIItrun-IL-36b |C90 0,1 1 12 trun- |L-36g |C90 (nM) FOL (nM) -_n-“(nM) 7.9 -—n-n(nM) 1.3 _—___ -_“-pNFKB (nM) 1.2 -_“-n(nM) 4.1 M) 4.3 Example 4: Binding of mouse anti-human lL-36R antibodies to human lL-36R expressing cells Protocol for bind/n of antibodies b flow 0 fomez‘r HEK293 cells transfected with full-length human |L-36R or NCI/ADR-RES cells were passaged for 24 hours prior to staining. Cells were removed from flasks by g with ml of 5 mM EDTA in PBS, and then incubated at 37°C for 10 min with an additional ml of 5 mM EDTA and 2.5 ml of Accumax to declump/disperse cells. Antibodies were then diluted to ied concentrations in PBS + 2% BSA, and cells incubated for min at room temperature. Excess antibody was then washed by adding 200 pl of PBS and then centrifuged. Secondary reagent was then added at 50 pl per well and cells are incubated for 15 min at room temperature and then washed as above. Cells were resuspended in 200 pl PBS and analyzed by flow cytometry. The binding EC5o’s for the mouse anti-human |L-36R dies binding to human |L-36R HEK transfectants are shown in Table 4.

Table 4 Clone EC5O(M) binding to HEK-|L-36R cells 33D10 4.748e-O1O 67E7 5.321 e-O1 O 73F6 7.456e- 010 76E10 4.257e-010 7808 52896-010 81A1 2.7956-010 81B4 3.0166-010 89A12 6.0896-010 Example 5: Production of Humanized |L-36R Antibodies In order to reduce potential immunogenicity following administration in man the mouse anti-human lL-36R monoclonal antibodies 81 B4 and 7305 were ‘humanized’ through a design and screening process. Human framework sequences were ed for the mouse leads based on the framework homology, CDR structure, conserved canonical residues, conserved interface packing residues and other parameters. The specific substitution of amino acid residues in these framework ons can improve various aspects of antibody performance including binding ty and/or stability, over that demonstrated in humanized antibodies formed by "direct swap" of CDRs or HVLs into the human germline framework regions. Fabs that showed better or equal binding and improved expression as compared to the chimeric parent Fab were selected for further characterization. Representative humanized variable regions for antibody 81 B4 and 7305 are shown in are shown the specification n. In this manner, Antibody B1 to dy B6 were zed antibodies derived from mouse dy 81 B4 (cloned into a human lng KO (KO=knock-out)/kappa backbone. Antibodies B1 to B6 are shown in Table A. Antibody Ct to Antibody 03 were humanized dies derived from mouse antibody 7305 (cloned into a human lng -KO (KO=knock-out)/kappa backbone. Antibodies Ct to 03 are shown in Table C.

Example 6: Binding of humanized |L-36R antibodies Kinetics and binding affinities of humanized anti-lL-36R antibodies binding to recombinant human lL-36R were measured using the Proteon (Bio-Rad, Hercules, CA).

Human lL-36R was immobilized at 5 different surface densities and s analyzed using global fit (see Table 5 showing results of three experiments). Binding of the zed antibodies to NCl/ADR-RES cells via flow cytometry was ed using protocol described in Example 4 (See Table 5 for E090 values).

Table 5 — Molecular and Cellular Binding affinities of humanized uman |L- 36R antibodies.

Antibody (LC + HC) KB 1 Standard Deviation (pM) 1M) B1 - 32_138 + 33_49 27 12.5 32 - 32_138 + 33_85 32 15.4 B3 - 32_138 + 33_90 20:2.2 - 24:7 BG-32105+33 49 4114.9 Example 7: Potency of humanized anti-human |L-36R antibodies in functional human assays onal blockade of signaling with the humanized lL-36R variants from human NCl/ADR-RES cells were tested as described in Example 3. ngo results for the humanized anti-human lL-36R antibodies in human onal assays (pNFKB and cytokine release) with human lL-36 ligands are shown in Table 6.

Table 6: Potency [ngo (nM)] of humanized antibodies in human functional R-RES cell assays ts equal averages of at least 2 experiments. lsotype controls demonstrated no inhibition of activity at highest tration tested for samples) -------m I-I-I-I-IflflpNFKtrun-IL- III-“nuntrun-ILI-I-I-I-I-I36g I-I-I-I-I-n tr”“'”' 3.2 2.9 2.7 2.9 2.5 2.3 NA NA NA I-I-I-I-I-I Example 8: Potency of anti-lL-36R antibodies in functional human primary keratinocyte assays Protocols: Human rimar e idermal noc te NFkB/C tokine release assa s Cells were plated at 30,000 cells/well in culture media in 96 well plates and incubated overnight at 37°C, 5% COZ Assays were then performed as described in Example 3.

Results: ngo results for mouse, chimeric, and humanized anti-lL-36R antibodies in human primary keratinocyte assays (pNFkB and lL-8 release) stimulated with human IL- 36 ligands are shown in Table 7, Table 8, and Table 9, respectively.

Table 7 (isotype controls demonstrated no inhibition of activity at t concentration tested for samples) 33D10 73C5 73F6 78C8 81A1 8184 trun-IL- 36a |C90 1.3 (nM) trun-IL- 36b |C90 1.1 trun-IL- 36g |C90 . . 1.3 10.6 ND 0.5 (nM) -trun-IL- 36a |C90 . . 17.8 19.3 ND 0.7 (nM) trun-IL- 36b |C90 1.2 (nM) trun-IL- 36g |C90 0.8 2.4 2.1 3.6 ND 0.5 Table 8: Potency of chimeric anti-lL-36R antibodies in primary human keratinocyte assays (isotype controls demonstrated no inhibition of ty at highest concentration tested for s) C73C5 C81A1 C81B4 NHK - $232-$338 ND -Egg-m ND -raga-52b 87 Table 9: Potency of humanized anti-lL-36R antibodies in y human keratinocyte assays (isotype controls demonstrated no tion of activity at highest concentration tested for samples) 11111111 - - -11111111 - - 111111111 1.1111111 -.11111111 Example 9 Potency of anti-lL-36R dies in onal human primary intestinal epithelial cell assays Protocols: Human rimar intestinal e ithelial cell NFkB/C tokine release assa s Cells were plated at 30,000 cells/well in culture media in 96we|| plate and incubated overnight at 37°C, 5% COz_Assays were then performed as described in Example 3.

Results: ICgo results for mouse anti-IL-36R antibodies in human primary intestinal epithelial cell assays (pNFkB and IL-8 release) stimulated with human IL-36 ligands are shown in Table 10.

Table 10: Potency of mouse anti-lL-36R dies in primary human intestinal epithelial cell assays (isotype ls demonstrated no tion of ty at highest concentration tested for samples) 73C5 8184 trun-IL-36a |C9O 21 1.3 (nM) trun-IL-36b |C9O 7.4 ( ) ’(crunilL-36g |C9O 32 9.5 trun-IL-36a |C9O 123 5.8 (nM) ’(crunilL-36b |C9O 154 9.8 ’(crunilL-36g |C9O ND 16 Example 10: Potency of anti-lL-36R antibodies in functional human primary intestinal myofibroblast assays Protocols: Human primary intestinal myofibroblast QNFKB/Cytokine release assays Cells were plated at 30,000 cells/well in culture media in 96wel| plate and incubated at 37°C, 5% COz_Assays were then performed as described in Example 3.

Results: ICgo results for anti-lL-36R antibodies in human primary intestinal myofibroblast assays (pNFkB and lL-8 release) stimulated with human lL-36 ligands are shown in Table 11 and Table 12,.

Table 11 : Potency of mouse and chimeric anti-lL-36R dies in primary human inal roblast assays (isotype controls demonstrated no inhibition of activity at highest concentration tested for samples) trun-IVI|-L36a |C9O pNFKB trun-IVI|-L36b |C9O n(M) t run- |L-36g |C9O n(M) trun- |L-36a |C9O lL-8 n(M) trun- |L-36b |C9O trun-IL-36g |C9O (nM) Table 12: Potency of humanized L-36R antibodies in primary human intestinal myofibroblast assays (isotype controls demonstrated no inhibition of activity at highest concentration tested for samples) '33 BS BG pNFkB Emilia 9'8 5-4 3-2 T233153? 3-7 1.7 2.8 T233??? 2-6 1.7 3.7 Tabitha? 4'4 3-8 2-1 lL-8 T233153? 2-8 2.6 2.0 T531536)“; 5-8 6-7 5.2 Example 11 : Potency of anti-lL-36R antibodies in functional human primary dermal fibroblast assays ols: Human primary dermal last QNFKB/Cztokine release assays Cells were plated at 30,000 cells/well in culture media in 96well plate and incubated overnight at 37°C, 5% COz_Assays were then performed as described in Example 3.

Results: ngo results for anti-lL-36R antibodies in human primary dermal fibroblast assays (pNFkB and lL-8 release) stimulated with human lL-36 ligands are shown in Table 13 and Table 14,.

Table 13: y of mouse and chimeric anti-lL-36R antibodies in primary human dermal fibroblast assays (isotype controls demonstrated no inhibition of activity at highest concentration tested for samples) 81B4 C81B4 trun-lL-36a |C9O 1.9 1.8 pNFKB (nM) t -lL-36 IC9O L-36a |C9O t -lL-36blC9O t(2:2)-lL-36g IC9O 0.4 0.3 Table 14: Potency of humanized anti-lL-36R antibodies in y human dermal fibroblast assays (isotype ls demonstrated no inhibition of ty at highest concentration tested for samples) trun-lL-36a |C9O PNFKB ---n trun-IL-36b |C9O _trun-IL-36g IC90"‘1- WO 74569 (nM) —— ----- ---- Example 12: Potency of mouse anti-lL-36R antibodies in functional human primary proximal tubular cell assays Protocol: human rimar roximal tubular cells NFkB/C tokine release assa s.

Cells were plated at 5,000 cells/well in culture media in 96well plate and incubated overnight at 37°C, 5% COz_Assays were med as described in Example 3.

Results: ICgo results for mouse, chimeric, and humanized anti-lL-36R dies in human primary proximal tubular cell assays (IL-8 release) stimulated with human lL-36 ligands are shown in Table 15.

Table 15: Potency of mouse and human L-36R antibodies in primary human proximal tubular cell assays (isotype controls demonstrated no inhibition of ty at highest concentration tested for samples) BS 86 -trun-lL-36a 5 ND lL-8 |C9O (nM) trun-lL-36b 5 ND |C9O (nM) trun-lL-36g 3 ND |C9O (nM) 2012/064933 Example 13: Inhibition of lL-8 production from lL-367 stimulated reconstructed human mis Protocol reconstructed epidermis Anti-lL-36R antibodies (1.5 ug/ml) were pre-incubated with reconstructed human epidermis and stimulated with human recombinant lL-36y (20 ng/ml). Recombinant human lL-1B (20 ng/ml; R & D Systems) was used as a positive control. After 24 hours in culture, cell supernatants were collected and assayed for lL-8 s for lL-8 are described in Example 3). s were tested in triplicate and the average pg/ml 1 standard error is shown in the table below (Table 16).

Table 16 Average lL-8 (pg/ml) dy Cytokine Stimulation +/- Standard Error Noantibody 57.31153 Example 14: Inhibition of lL-36 ligand induced S100A7 and S100A12 gene expression in reconstructed human epidermis Stimulation of reconstructed human epidermis with agonsitic lL-36 s induces S100A7 and S100A12 gene expression. S100A7 and S100A12 are genes located within the epidermal differentiation complex.

Protocol: Reconstructed human epidermis were incubated with anti-lL-36R dies (1.5 ug/ml) and stimulated with human recombinant lL-36y (20 ng/ml). Recombinant human lL-1B (20 ng/mL; R & D Systems) was used as a positive control. After 24 hours in culture at 5% 002 and 37°C, RNA was isolated from the reconstructed human epidermis and assayed for gene expression by real-time reverse trancriptasepolymerase chain on. Relative expression was calculated using the 2AACt method.

Samples were tested in triplicate and the e expression 1 standard error is shown in the table below (Table 17).

Table 17 Mean S100A7 Mean S100A12 Antibody Cytokine Expression +/- sion +/- Stimulation Standard Error Standard Error No antibody None 1.00 :r 0.79 1.00 :r 0.47 33D10 None 3.92 :r 0.36 1.93 :r 0.02 No antibody 20 ng/mL IL-1B 76.03 1 24.66 47.84 i 9.24 33D1O 20 ng/mL IL-1B 95.83 i 11.83 76.41 i 6.92 No antibody 20 ng/mL lL-36y 19.57 i 3.26 20.53 i 5.21 33D1O 20 ng/mL |L-36v 3.47 i 1.37 2.01 i 0.35 Example 15: Efficacy of anti-lL-36R antibody in Xenotransplant Model of Psoriasis ol: Blood and sional skin biopsies were obtained from 24 psoriasis patients that were clinically diagnosed by a dermatologist. Skin biopsies were transplanted onto immune-deficient NIH-Ill mice and allowed to t for a period of four to five weeks.

Peripheral blood mononuclear cells (PBMC) were isolated from blood collected from each donor at the time of biopsy for intradermal injection into the engrafted skin. Prior to injection, PBMC were stimulated with with 1 ug/ml Staphylococcal Enterotoxin B (Toxin Technologies, Florida, USA) and 80 U/ml human inant lL-2 (Peprotech lnc., Oosterhout, The Netherlands). Autologous PBMC were intradermally injected with 7.5 x 10"5 cells in PBS to synchronize the induction of skin inflammation and the psoriasis phenotype. Three weeks after the injection of cells, the biopsies were retrieved from the mice and analyzed by histology.

Histological staining was performed on cryo-preserved skin tissue of all groups.

Diagonal cross sections (8 pm), covering all ayers, were prepared as described in Figure 4. For assessment of epidermal ess, two rial sections were randomly chosen from the center of the biopsy and stained with haematoxylin- eosin.Subsequently, sections were evaluated at a 100-fold magnification. Over the entire length of the biopsy, ridge lengths were measured in both sections using an Olympus DP71 camera and CellAD imaging software (V2.7, Munster, Germany).

Ridge length is defined as: the distance between the upper edge of the stratum granulosum to the bottom of the ridge. es were scored at random and in a blinded fashion.

Results for the average epidermal ess and maximum epidermal thickness for each treatment group are shown in Table 18. s for the net change in mal thickness in each treatment group are shown in Table 19.

Table 18 33D10 Table 19 _Average Epidermal Thickness Maximum Epidermal Thickness 2012/064933 —mmnmmmnmUntreated 36. 2 31. 9 52. 6 40.0 Vehicle 43.5 30.9 18 63.4 52.0 Example 16: The sub-chronic pulmonary inflammation after 3 weeks of cigarette smoke exposure in wild type and interleukin-1 receptor-like 2 homozygous knockout mice Protocol: Wild type or interleukin-1 receptor-like 2 mice were exposed to cigarette smoke for 3 weeks to induce ary distress. Weeks 1 and 2 consisted of 5 consecutive exposure days, while mice were exposed for 4 utive days during week 3. Mice were exposed to 5 cigarettes each day with 24 minute intervals of cigarette exposure (16 minutes) and fresh air (8 minutes). Eighteen hours following the final re, mice were lavaged with 2 x 0.8 ml of Hank’s Salt Solution (0.6mM EDTA). The supernatant and cell pellet were collected from the bronchial alveolar lavage following centrifugation for 10 s. Total macrophage and neutrophil cell counts in the bronchial alveolar lavage for each exposure group are shown in Table 20.

Table 20 Total Cells Cell Counts x 10A5 Mouse mean SD N SEM WT 2.21 1.47 9 0.49 |L1RL2 KO 2.45 0.87 6 0.36 WT + OS 9.07 2.83 10 0.90 IL-1RL2 KO + OS 5.32 1.03 10 0.32 Macrophages Cell Counts x 10A5 Mouse mean SD N SEM WT 2.08 1.62 9 0.54 WO 74569 |L1RL2 KO 2.40 0.86 0.35 WT + C8 3.36 1.46 10 0.46 IL-1RL2 KO + C8 3.22 0.86 10 0.27 Neutrophils Cell Countsx1 0‘5 Mouse mean SD SEM WT 0.002 0.004 0.001 |L1RL2 KO 0.013 0.016 0.007 WT + C8 5.698 2.751 10 0.870 IL-1RL2 KO + C8 2.083 0.749 10 0.237 WE

Claims (47)

CLAIM :

1. An anti-interleukin-36R (anti-IL-36R) antibody or antigen-binding fragment f, wherein the antibody or antigen-binding nt thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 35, 102, 103, 104, 105, 106 or 140 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain le region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid ce of SEQ ID NO: 72 (H-CDR3).

2. An anti-IL-36R dy or antigen-binding fragment thereof ing to claim 1, wherein the antibody or antigen-binding fragment thereof comprises: a) a light chain le region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 102 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

3. An anti-IL-36R antibody or antigen-binding fragment thereof according to claim 1, wherein the antibody or antigen-binding fragment thereof comprises: a) a light chain le region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 103 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid ce of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

4. An anti-IL-36R antibody or antigen-binding fragment f according to claim 1, wherein the antibody or antigen-binding fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 104 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and AH26(10651560_1):RTK b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

5. An anti-IL-36R antibody or antigen-binding fragment thereof according to claim 1, wherein the antibody or antigen-binding fragment thereof ses: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 105 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

6. An anti-IL-36R antibody or antigen-binding fragment thereof according to claim 1, wherein the antibody or antigen-binding fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid ce of SEQ ID NO: 106 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 3); and b) a heavy chain variable region comprising the amino acid ce of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

7. An anti-IL-36R antibody or antigen-binding fragment thereof according to claim 1, wherein the antibody or antigen-binding fragment thereof comprises: a) a light chain variable region sing the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 140 2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62, 108, 109, 110 or 111 (HCDR2 ); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

8. An L-36R antibody or antigen-binding nt thereof according to claim 1, wherein the antibody or antigen-binding fragment thereof comprises a light chain variable region comprising the amino acid sequence of any one of SEQ ID NO: 76, 77, 78, 79, 80, 81, 82 or 83; AH26(10651560_1):RTK and a heavy chain variable region comprising the amino acid ce of any one of SEQ ID NO: 87, 88, 89, 90, 91, 92, 93, 94 or 95.

9. An anti-IL-36R antibody or antigen-binding fragment thereof according to claim 8, n the antibody or antigen-binding fragment thereof comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 77; and a heavy chain variable region sing the amino acid ce of SEQ ID NO: 87;or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 77; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 88; or a light chain le region comprising the amino acid sequence of SEQ ID NO: 77; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 89.

10. An anti-IL-36R antibody or antigen-binding fragment thereof ing to claim 8, wherein the antibody or antigen-binding fragment thereof comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 80; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 87;or a light chain variable region comprising the amino acid ce of SEQ ID NO: 80; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 88; or a light chain variable region comprising the amino acid sequence of SEQ ID NO: 80; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 89.

11. An anti-IL-36R antibody according to claim 1, wherein the dy comprises a light chain comprising the amino acid sequence of any one of SEQ ID NO: 114, 115, 116, 117, 118, 119, 120 or 121; and a heavy chain comprising the amino acid sequence of any one of SEQ ID NO: 125, 126, 127, 128, 129, 130, 131, 132 or 133.

12. An anti-IL-36R antibody, wherein the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 115; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 125.

13. An anti-IL-36R antibody, wherein the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 115; and a heavy chain sing the amino acid sequence of SEQ ID NO: 126. AH26(10651560_1):RTK

14. An anti-IL-36R antibody, wherein the antibody comprises a light chain comprising the amino acid ce of SEQ ID NO: 115; and a heavy chain sing the amino acid sequence of SEQ ID NO: 127.

15. An anti-IL-36R antibody, wherein the antibody ses a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 125.

16. An L-36R antibody, wherein the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 126.

17. An anti-IL-36R antibody, wherein the antibody thereof comprises a light chain comprising the amino acid sequence of SEQ ID NO: 118; and a heavy chain comprising the amino acid sequence of SEQ ID NO: 127.

18. An anti-IL-36R antibody or antigen-binding fragment thereof, n the antibody or antigen-binding fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid sequence of SEQ ID NO: 103 (L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid sequence of SEQ ID NO: 62 (H-CDR2); the amino acid sequence of SEQ ID NO: 72 (H-CDR3).

19. An anti-IL-36R antibody or antigen-binding fragment f, wherein the antibody or antigen-binding fragment thereof comprises: a) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 26 (L-CDR1); the amino acid ce of SEQ ID NO: 104(L-CDR2); the amino acid sequence of SEQ ID NO: 44 (L-CDR3); and b) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 53 (H-CDR1); the amino acid ce of SEQ ID NO: 62 (H-CDR2); the amino acid sequence of SEQ ID NO: 72 (H-CDR3). AH26(10651560_1):RTK

20. An anti-IL-36R antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 80; and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 87.

21. An anti-IL-36R dy or antigen-binding fragment f, wherein the antibody or n-binding nt f comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 80; and a heavy chain variable region sing the amino acid sequence of SEQ ID NO: 88.

22. An anti-IL-36R antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 80; and a heavy chain variable region comprising the amino acid ce of SEQ ID NO: 89.

23. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 11 and a pharmaceutically acceptable carrier.

24. A pharmaceutical ition sing an antibody or antigen-binding fragment according to claim 12 and a pharmaceutically acceptable carrier.

25. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 13 and a pharmaceutically acceptable carrier.

26. A ceutical composition comprising an antibody or antigen-binding fragment according to claim 14 and a pharmaceutically acceptable carrier.

27. A ceutical composition comprising an antibody or antigen-binding fragment according to claim 15 and a pharmaceutically acceptable carrier.

28. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 16 and a pharmaceutically acceptable carrier.

29. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 17 and a pharmaceutically acceptable carrier. AH26(10651560_1):RTK

30. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 8 and a pharmaceutically acceptable carrier.

31. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 20 and a pharmaceutically acceptable carrier.

32. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 21 and a pharmaceutically acceptable r.

33. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 22 and a pharmaceutically acceptable carrier.

34. A ceutical composition comprising an antibody or antigen-binding fragment according to claim 18 and a pharmaceutically able carrier.

35. A pharmaceutical composition comprising an antibody or antigen-binding fragment according to claim 19 and a pharmaceutically acceptable carrier.

36. A pharmaceutical composition comprising an antibody or antigen-binding fragment ing to claim 1 and a pharmaceutically able r.

37. An antibody or n-binding fragment or pharmaceutical composition according to any one of claims 1 to 36, for use in a method for the treatment of ulcerative colitis.

38. Use of an antibody or antigen-binding nt according to any one of claims 1 to 22 or a pharmaceutical composition according to any one of claims 23 to 36, for the manufacture of a medicament for the treatment of a disease, wherein the disease is ed from an inflammatory disease, an autoimmune disease, a respiratory disease, a metabolic disorder, an epithelial mediated inflammatory disorder, fibrosis and cancer.

39. The use according to claim 38, wherein the disease is selected from psoriasis, inflammatory bowel e, psoriatic arthritis, le sclerosis, rheumatoid arthritis, COPD, c asthma and ankylosing spondylitis.

40. The use according to claim 38 and/or 39, wherein the disease is Crohn’s disease. AH26(10651560_1):RTK

41. The use according to claim 38 and/or 39, wherein the disease is ulcerative colitis.

42. An isolated polynucleotide comprising a sequence encoding an anti-IL-36R antibody or n-binding fragment according to any one of claims 1 to 22, preferably a DNA or RNA sequence.

43. An isolated polynucleotide according to claim 42, encoding a sequence as defined by one or more of SEQ ID NOs. 1 to 140.

44. A vector comprising a polynucleotide ing to claim 42 and/or 43, preferably an expression vector, more preferred a vector comprising the polynucleotide according to the invention in functional association with an expression control sequence.

45. A host cell comprising a polynucleotide according to claim 42 and/or 43, and/or a vector according to claim 44, wherein the host cell is not within a human.

46. A method for the production of an anti-IL-36R antibody or antigen-binding fragment ing to any one of claims 1 to 22, preferably a recombinant production method comprising the use of a polynucleotide according to claim 42 and/or 43, and/or of a vector according to claim 44 and/or of a host cell according to claim 45.

47. The method according to claim 46, comprising the steps (a) ating the host cell under conditions ng the expression of the anti-IL-36R antibody or antigen-binding fragment and (b) recovering the anti-IL-36R antibody or antigen-binding nt. Boehringer Ingelheim ational GmbH By the Attorneys for the Applicant SPRUSON & FERGUSON Per: AH26(10651560_1):RTK