Antibodies that bind interleukin 13 and methods of use

Abstract

Described herein are novel and improved antibodies that bind to interleukin 13 (IL-13) and methods of use thereof. In certain aspects, methods of inhibiting the biological activity of IL-13 are described herein. In certain aspects, pharmaceutical compositions comprising the anti-IL-13 antibodies are described herein. In certain aspects, the antibodies and methods described herein are used to treat inflammatory diseases or disorders associated with elevated levels of IL-13 and / or IgE.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims the benefit of and priority to U.S. Provisional Application No. 63 / 353,367 filed on June 17, 2022, U.S. Provisional Application No. 63 / 462,822 filed on April 28, 2023, and U.S. Provisional Application No. 63 / 469,167 filed on May 26, 2023, the entire disclosures of which are hereby incorporated by reference in their entirety for all purposes. Background Art

[0003] Interleukin (IL) -13 is a T helper cell subclass 2 (Th2) cytokine and belongs to the type I cytokine family, which exhibits pleiotropic effects in multiple cell pathways. IL-13 is related to the differentiation of initial T cells into Th2 cells. When co-stimulated with CD40 / CD40L, IL-13 promotes B cell proliferation and induces immunoglobulin isotype class conversion to IgG4 and IgE. It also raises FcεRI, and therefore contributes to the IgE stimulation of mast cells. In monocytes / macrophages, IL-13 raises the expression of CD23 and MHC class I and class II antigens, lowers CD14 expression, inhibits antibody-dependent cellular toxicity, and promotes eosinophil survival, activation and recruitment. IL-13 also shows important functions on non-hematopoietic cells (such as smooth muscle cells, epithelial cells, endothelial cells and fibroblasts). IL-13 enhances the proliferation of smooth muscle and the contraction induced by cholinergics. In epithelial cells, IL-13 is a potent inducer of chemokine production, alters mucociliary differentiation, reduces ciliary beat frequency of ciliated epithelial cells, and causes goblet cell metaplasia. In endothelial cells, IL-13 is a potent inducer of vascular cell adhesion molecule 1 (VCAM-1), which is critical for eosinophil recruitment. In epithelial keratinocytes, IL-13 reduces the expression of barrier integrity molecules such as filamentous protein and loricrin, while stimulating the secretion of CCL26 and CCL2, which are responsible for recruiting several myeloid lineage inflammatory cells. In human dermal fibroblasts, IL-13 induces type 1 collagen synthesis in human dermal fibroblasts.

[0004] Inhibition of IL-13 can be used to treat or prevent inflammatory diseases and disorders, such as those associated with elevated IgE levels, including but not limited to asthma, allergic rhinitis, urticaria, and allergic or atopic dermatitis. Therefore, there is a need to develop potent and specific IL-13 inhibitors (e.g., inhibitors that retain activity for a longer period of time when administered to a subject) to prevent and / or treat IL-13 and IgE-mediated diseases or disorders. Summary of the invention

[0005] In certain aspects, described herein is an isolated antibody that binds IL-13, comprising i) a variable heavy (VH) chain sequence comprising three heavy chain CDR sequences CDR-H1, CDR-H2, and CDR-H3; and ii) a variable light (VL) chain sequence comprising three light chain CDR sequences CDR-L1, CDR-L2, and CDR-L3; wherein: a) CDR-H1 comprises a sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 58-99 and 121; b) CDR-H2 comprises a sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 100-111; c) CDR-H3 comprises a sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 112-120 and 130-140; d) CDR-L1 comprises a sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 141-144 and 149-152; e) CDR-L2 comprises a sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: NO: 153-158 and the amino acid sequence LAS; and f) CDR-L3 comprises a sequence selected from the group consisting of the sequences shown in SEQ ID NO: 165-172.

[0006] In certain embodiments, the isolated antibody comprises: a) CDR-H1 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 58-66; b) CDR-H2 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 100-103; c) CDR-H3 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 112-120; d) CDR-L1 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 141-144; e) CDR-L2 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 153-158; and f) CDR-L3 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 165-172.

[0007] In certain embodiments, the isolated antibody comprises: a) CDR-H1 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 67-83; b) CDR-H2 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 104-107; c) CDR-H3 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 112-120; d) CDR-L1 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 141-144; e) CDR-L2 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 153-158; and f) CDR-L3 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 165-172.

[0008] In certain embodiments, the isolated antibody comprises: a) CDR-H1 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 84-99 and 121; b) CDR-H2 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 108-111; c) CDR-H3 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 130-140; d) CDR-L1 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 149-152; e) CDR-L2 comprising the amino acid sequence LAS; and f) CDR-L3 comprising a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 165-172.

[0009] In certain embodiments, the isolated antibody does not comprise: a) CDR-H1 set forth in SEQ ID NO: 58; CDR-H2 set forth in SEQ ID NO: 100; CDR-H3 set forth in SEQ ID NO: 112; CDR-L1 set forth in SEQ ID NO: 141; CDR-L2 set forth in SEQ ID NO: 153; and CDR-L3 set forth in SEQ ID NO: 165; or b) CDR-H1 set forth in SEQ ID NO: 67; CDR-H2 set forth in SEQ ID NO: 104; CDR-H3 set forth in SEQ ID NO: 112; CDR-L1 set forth in SEQ ID NO: 141; CDR-L2 set forth in SEQ ID NO: 153; and CDR-L3 set forth in SEQ ID NO: 165; or c) CDR-H1 set forth in SEQ ID NO: 84; CDR-H2 set forth in SEQ ID NO: 108; CDR-H3 set forth in SEQ ID NO: 130; CDR-L1 shown in NO:149; CDR-L2 shown in the amino acid sequence LAS; and CDR-L3 shown in SEQ ID NO:165.

[0010] In certain embodiments, the antibody does not comprise any combination of: a) CDR-H1 shown in any one of SEQ ID NOs: 58, 67 or 84; b) CDR-H2 shown in any one of SEQ ID NOs: 100, 104 or 108; c) CDR-H3 shown in any one of SEQ ID NOs: 112 or 130; d) CDR-L1 shown in any one of SEQ ID NOs: 141 or 149; e) CDR-L2 shown in any one of SEQ ID NOs: 153 or 154; and f) CDR-L3 shown in SEQ ID NO: 165.

[0011] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising a sequence as set forth in any one of SEQ ID NOs: 58, 67 or 68; a CDR-H2 comprising a sequence as set forth in any one of SEQ ID NOs: 100 or 104; a CDR-H3 comprising a sequence as set forth in SEQ ID NO: 112; a CDR-L1 comprising a sequence as set forth in any one of SEQ ID NOs: 141 or 149; a CDR-L2 comprising a sequence as set forth in any one of SEQ ID NOs: 153 or amino acid sequence LAS; and a CDR-L3 comprising a sequence as set forth in SEQ ID NO: 165.

[0012] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO: 58; a CDR-H2 comprising the sequence shown in SEQ ID NO: 100; a CDR-H3 comprising the sequence shown in SEQ ID NO: 112; a CDR-L1 comprising the sequence shown in SEQ ID NO: 141; a CDR-L2 comprising the sequence shown in SEQ ID NO: 153; and a CDR-L3 comprising the sequence shown in SEQ ID NO: 165.

[0013] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO: 67; a CDR-H2 comprising the sequence shown in SEQ ID NO: 104; a CDR-H3 comprising the sequence shown in SEQ ID NO: 112; a CDR-L1 comprising the sequence shown in SEQ ID NO: 141; a CDR-L2 comprising the sequence shown in SEQ ID NO: 153; and a CDR-L3 comprising the sequence shown in SEQ ID NO: 165.

[0014] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO: 68; a CDR-H2 comprising the sequence shown in SEQ ID NO: 104; a CDR-H3 comprising the sequence shown in SEQ ID NO: 112; a CDR-L1 comprising the sequence shown in SEQ ID NO: 141; a CDR-L2 comprising the sequence shown in SEQ ID NO: 153; and a CDR-L3 comprising the sequence shown in SEQ ID NO: 165.

[0015] In certain embodiments, the isolated antibody comprises: CDR-H1, which comprises the sequence shown in SEQ ID NO: 67; CDR-H2, which comprises the sequence shown in SEQ ID NO: 104; CDR-H3, which comprises the sequence shown in SEQ ID NO: 112; CDR-L1, which comprises the sequence shown in SEQ ID NO: 149; CDR-L2, which comprises the amino acid sequence LAS; and CDR-L3, which comprises the sequence shown in SEQ ID NO: 165.

[0016] In certain embodiments, the isolated antibody comprises: CDR-H1, which comprises the sequence shown in SEQ ID NO: 68; CDR-H2, which comprises the sequence shown in SEQ ID NO: 104; CDR-H3, which comprises the sequence shown in SEQ ID NO: 112; CDR-L1, which comprises the sequence shown in SEQ ID NO: 149; CDR-L2, which comprises the amino acid sequence LAS; and CDR-L3, which comprises the sequence shown in SEQ ID NO: 165.

[0017] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising a sequence as set forth in any one of SEQ ID NOs: 58, 67, 68, 84 or 85; a CDR-H2 comprising a sequence as set forth in any one of SEQ ID NOs: 100, 104 or 108; a CDR-H3 comprising a sequence as set forth in any one of SEQ ID NOs: 112 or 130; a CDR-L1 comprising a sequence as set forth in any one of SEQ ID NOs: 141 or 149; a CDR-L2 comprising a sequence as set forth in any one of SEQ ID NOs: 153 or amino acid sequence LAS; and a CDR-L3 comprising a sequence as set forth in SEQ ID NO: 165.

[0018] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO: 68; a CDR-H2 comprising the sequence shown in SEQ ID NO: 104; a CDR-H3 comprising the sequence shown in SEQ ID NO: 112; a CDR-L1 comprising the sequence shown in SEQ ID NO: 141; a CDR-L2 comprising the sequence shown in SEQ ID NO: 153; and a CDR-L3 comprising the sequence shown in SEQ ID NO: 165.

[0019] In certain embodiments, the isolated antibody comprises: CDR-H1, which comprises the sequence shown in SEQ ID NO: 84; CDR-H2, which comprises the sequence shown in SEQ ID NO: 108; CDR-H3, which comprises the sequence shown in SEQ ID NO: 130; CDR-L1, which comprises the sequence shown in SEQ ID NO: 149; CDR-L2, which comprises the amino acid sequence LAS; and CDR-L3, which comprises the sequence shown in SEQ ID NO: 165.

[0020] In certain embodiments, the isolated antibody comprises: CDR-H1, which comprises the sequence shown in SEQ ID NO: 85; CDR-H2, which comprises the sequence shown in SEQ ID NO: 108; CDR-H3, which comprises the sequence shown in SEQ ID NO: 130; CDR-L1, which comprises the sequence shown in SEQ ID NO: 149; CDR-L2, which comprises the amino acid sequence LAS; and CDR-L3, which comprises the sequence shown in SEQ ID NO: 165.

[0021] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising a sequence as shown in any one of SEQ ID NOs: 58, 67, 68, 84 or 85; a CDR-H2 comprising a sequence as shown in any one of SEQ ID NOs: 100, 104 or 108; a CDR-H3 comprising a sequence as shown in any one of SEQ ID NOs: 112 or 130; a CDR-L1 comprising a sequence as shown in any one of SEQ ID NOs: 141 or 149; a CDR-L2 comprising a sequence as shown in any one of SEQ ID NOs: 157 or amino acid sequence LAS; and a CDR-L3 comprising a sequence as shown in SEQ ID NO: 165.

[0022] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO: 58; a CDR-H2 comprising the sequence shown in SEQ ID NO: 100; a CDR-H3 comprising the sequence shown in SEQ ID NO: 112; a CDR-L1 comprising the sequence shown in SEQ ID NO: 141; a CDR-L2 comprising the sequence shown in SEQ ID NO: 157; and a CDR-L3 comprising the sequence shown in SEQ ID NO: 165.

[0023] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO: 68; a CDR-H2 comprising the sequence shown in SEQ ID NO: 104; a CDR-H3 comprising the sequence shown in SEQ ID NO: 112; a CDR-L1 comprising the sequence shown in SEQ ID NO: 141; a CDR-L2 comprising the sequence shown in SEQ ID NO: 157; and a CDR-L3 comprising the sequence shown in SEQ ID NO: 165.

[0024] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising a sequence as shown in any one of SEQ ID NOs: 58, 67, 68, 84 or 85; a CDR-H2 comprising a sequence as shown in any one of SEQ ID NOs: 100, 104 or 108; a CDR-H3 comprising a sequence as shown in any one of SEQ ID NOs: 112 or 130; a CDR-L1 comprising a sequence as shown in any one of SEQ ID NOs: 141 or 149; a CDR-L2 comprising a sequence as shown in any one of SEQ ID NOs: 157 or amino acid sequence LAS; and a CDR-L3 comprising a sequence as shown in SEQ ID NO: 165.

[0025] In certain embodiments, the isolated antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO: 68; a CDR-H2 comprising the sequence shown in SEQ ID NO: 104; a CDR-H3 comprising the sequence shown in SEQ ID NO: 112; a CDR-L1 comprising the sequence shown in SEQ ID NO: 141; a CDR-L2 comprising the sequence shown in SEQ ID NO: 157; and a CDR-L3 comprising the sequence shown in SEQ ID NO: 165.

[0026] In certain embodiments, the isolated antibody comprises: CDR-H1, which comprises the sequence shown in SEQ ID NO: 84; CDR-H2, which comprises the sequence shown in SEQ ID NO: 108; CDR-H3, which comprises the sequence shown in SEQ ID NO: 130; CDR-L1, which comprises the sequence shown in SEQ ID NO: 149; CDR-L2, which comprises the amino acid sequence LAS; and CDR-L3, which comprises the sequence shown in SEQ ID NO: 165.

[0027] In certain embodiments, the isolated antibody comprises: CDR-H1, which comprises the sequence shown in SEQ ID NO: 85; CDR-H2, which comprises the sequence shown in SEQ ID NO: 108; CDR-H3, which comprises the sequence shown in SEQ ID NO: 130; CDR-L1, which comprises the sequence shown in SEQ ID NO: 149; CDR-L2, which comprises the amino acid sequence LAS; and CDR-L3, which comprises the sequence shown in SEQ ID NO: 165.

[0028] In certain embodiments, the isolated antibody comprises a VH sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 1-32 and 470.

[0029] In certain embodiments, the isolated antibody comprises a VL sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 33-57 and 471.

[0030] In certain embodiments, the isolated antibody comprises a VH sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 1-32 and 470 and a VL sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 33-57 and 471.

[0031] In certain embodiments, the isolated antibody comprises a VH sequence selected from the group consisting of SEQ ID NOs: 1-32 and 470 and a VL sequence set forth in SEQ ID NO:49.

[0032] In certain embodiments, the isolated antibody comprises a VH sequence selected from the group consisting of SEQ ID NOs: 1-32 and 470 and a VL sequence as shown in SEQ ID NO:51.

[0033] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:1 and the VL sequence shown in SEQ ID NO:33.

[0034] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:2 and the VL sequence shown in SEQ ID NO:33.

[0035] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:35.

[0036] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:4 and the VL sequence shown in SEQ ID NO:35.

[0037] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:5 and the VL sequence shown in SEQ ID NO:35.

[0038] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:6 and the VL sequence shown in SEQ ID NO:35.

[0039] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:35.

[0040] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:36.

[0041] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:4 and the VL sequence shown in SEQ ID NO:36.

[0042] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:5 and the VL sequence shown in SEQ ID NO:36.

[0043] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:6 and the VL sequence shown in SEQ ID NO:36.

[0044] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:36.

[0045] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:39.

[0046] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:4 and the VL sequence shown in SEQ ID NO:39.

[0047] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:5 and the VL sequence shown in SEQ ID NO:39.

[0048] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:6 and the VL sequence shown in SEQ ID NO:39.

[0049] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:39.

[0050] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:40.

[0051] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:4 and the VL sequence shown in SEQ ID NO:40.

[0052] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:5 and the VL sequence shown in SEQ ID NO:40.

[0053] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:6 and the VL sequence shown in SEQ ID NO:40.

[0054] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:40.

[0055] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:8 and the VL sequence shown in SEQ ID NO:42.

[0056] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:9 and the VL sequence shown in SEQ ID NO:43.

[0057] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:39.

[0058] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:44.

[0059] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:45.

[0060] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:46.

[0061] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:7 and the VL sequence set forth in SEQ ID NO:47.

[0062] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:7 and the VL sequence set forth in SEQ ID NO:48.

[0063] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:7 and the VL sequence set forth in SEQ ID NO:49.

[0064] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:50.

[0065] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:51.

[0066] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:51.

[0067] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:52.

[0068] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:53.

[0069] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:54.

[0070] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:55.

[0071] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:56.

[0072] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:7 and the VL sequence set forth in SEQ ID NO:57.

[0073] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:10 and the VL sequence shown in SEQ ID NO:39.

[0074] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:11 and the VL sequence shown in SEQ ID NO:39.

[0075] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:12 and the VL sequence shown in SEQ ID NO:39.

[0076] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:13 and the VL sequence shown in SEQ ID NO:39.

[0077] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:14 and the VL sequence shown in SEQ ID NO:39.

[0078] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:15 and the VL sequence shown in SEQ ID NO:39.

[0079] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:16 and the VL sequence shown in SEQ ID NO:39.

[0080] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:17 and the VL sequence shown in SEQ ID NO:39.

[0081] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:18 and the VL sequence shown in SEQ ID NO:39.

[0082] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:19 and the VL sequence set forth in SEQ ID NO:39.

[0083] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:20 and the VL sequence shown in SEQ ID NO:39.

[0084] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:21 and the VL sequence shown in SEQ ID NO:39.

[0085] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:22 and the VL sequence shown in SEQ ID NO:39.

[0086] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:23 and the VL sequence shown in SEQ ID NO:39.

[0087] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:24 and the VL sequence shown in SEQ ID NO:39.

[0088] In certain embodiments, the antibody comprises the VH sequence shown in SEQ ID NO:25 and the VL sequence shown in SEQ ID NO:39.

[0089] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:26 and the VL sequence shown in SEQ ID NO:39.

[0090] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:27 and the VL sequence shown in SEQ ID NO:39.

[0091] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:28 and the VL sequence shown in SEQ ID NO:39.

[0092] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:28 and the VL sequence shown in SEQ ID NO:39.

[0093] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:29 and the VL sequence set forth in SEQ ID NO:39.

[0094] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:30 and the VL sequence shown in SEQ ID NO:39.

[0095] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:31 and the VL sequence shown in SEQ ID NO:39.

[0096] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:32 and the VL sequence shown in SEQ ID NO:39.

[0097] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:8 and the VL sequence shown in SEQ ID NO:39.

[0098] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:8 and the VL sequence shown in SEQ ID NO:51.

[0099] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:470 and the VL sequence shown in SEQ ID NO:471.

[0100] In certain embodiments, the separated antibodies are humanized antibodies, human antibodies or chimeric antibodies. In certain embodiments, the separated antibodies are humanized antibodies. In certain embodiments, the separated antibodies comprise a heavy chain human constant region of a class selected from IgG, IgA, IgD, IgE and IgM. In certain embodiments, the human Fc region comprises a class IgG and a human heavy chain constant region of a subclass selected from IgG1, IgG2, IgG3 and IgG4. In certain embodiments, the human Fc region comprises human IgG1 Fc. In certain embodiments, the human Fc region comprises human IgG4 Fc. In certain embodiments, the human Fc region comprises human IgG2 Fc.

[0101] In certain embodiments of the antibodies described herein, the heavy chain comprises a constant heavy chain sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 425-468 and 484-539.

[0102] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 1 and a VL sequence as shown in SEQ ID NO: 33; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0103] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 2 and a VL sequence as shown in SEQ ID NO: 33; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0104] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0105] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 4 and a VL sequence as shown in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0106] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 5 and a VL sequence as shown in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0107] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 6 and a VL sequence as shown in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0108] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0109] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0110] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 4 and a VL sequence as shown in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0111] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 5 and a VL sequence as shown in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0112] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 6 and a VL sequence as shown in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0113] In certain embodiments, the isolated antibody comprises a VH sequence shown in SEQ ID NO: 7 and a VL sequence shown in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NO: 439, 440, 446, 457 and 460.

[0114] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0115] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 4 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0116] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 5 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0117] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 6 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0118] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0119] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0120] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 4 and a VL sequence as shown in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0121] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 5 and a VL sequence as shown in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0122] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 6 and a VL sequence as shown in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0123] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0124] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 8 and a VL sequence as shown in SEQ ID NO: 42; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0125] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 9 and a VL sequence as shown in SEQ ID NO: 43; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0126] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0127] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 44; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0128] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 45; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0129] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 46; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0130] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 47; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0131] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 48; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0132] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 49; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0133] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 50; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0134] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0135] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0136] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 52; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0137] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 53; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0138] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 54; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0139] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 55; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0140] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 56; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0141] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 57; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0142] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 10 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0143] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 11 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0144] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 12 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0145] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 13 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0146] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 14 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0147] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 15 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0148] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 16 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0149] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 17 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0150] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 18 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0151] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 19 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0152] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 20 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0153] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 21 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0154] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 22 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0155] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 23 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0156] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 24 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0157] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 25 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0158] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 26 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0159] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 27 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0160] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 28 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0161] In certain embodiments, the isolated antibody comprises a VH sequence as set forth in SEQ ID NO: 28 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NOs: 436-468 and 484-539.

[0162] In certain embodiments, the isolated antibody comprises a VL sequence as set forth in SEQ ID NO: 29 and in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 436-468 and 484-539.

[0163] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 30 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0164] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 31 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0165] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 32 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0166] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO: 8 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0167] In certain embodiments, the isolated antibody comprises a VH sequence as set forth in SEQ ID NO: 8 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the isolated antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539.

[0168] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:439.

[0169] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:446.

[0170] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:457.

[0171] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:460.

[0172] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:439.

[0173] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:446.

[0174] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:457.

[0175] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:460.

[0176] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:439.

[0177] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:446.

[0178] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:457.

[0179] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:460.

[0180] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:439.

[0181] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:446.

[0182] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:457.

[0183] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:460.

[0184] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:439.

[0185] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:446.

[0186] In certain embodiments, the isolated antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:457.

[0187] In certain embodiments, the isolated antibody comprises a VH sequence shown in SEQ ID NO:470 and a VL sequence shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:460.

[0188] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:15 and the VL sequence set forth in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence set forth in SEQ ID NO:439.

[0189] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:15 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:457.

[0190] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:15 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:460.

[0191] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:15 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:446.

[0192] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:439.

[0193] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:457.

[0194] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:460.

[0195] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:3 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:446.

[0196] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:7 and the VL sequence set forth in SEQ ID NO:51; and wherein the human Fc region comprises the human IgG sequence set forth in SEQ ID NO:439.

[0197] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:51; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:457.

[0198] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:51; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:460.

[0199] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:7 and the VL sequence shown in SEQ ID NO:51; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:446.

[0200] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:8 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:439.

[0201] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:8 and the VL sequence shown in SEQ ID NO:39; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:457.

[0202] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:8 and the VL sequence shown in SEQ ID NO:51; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:460.

[0203] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:8 and the VL sequence shown in SEQ ID NO:51; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:446.

[0204] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:470 and the VL sequence set forth in SEQ ID NO:471; and wherein the human Fc region comprises the human IgG sequence set forth in SEQ ID NO:439.

[0205] In certain embodiments, the isolated antibody comprises the VH sequence set forth in SEQ ID NO:470 and the VL sequence set forth in SEQ ID NO:471; and wherein the human Fc region comprises the human IgG sequence set forth in SEQ ID NO:457.

[0206] In certain embodiments, the isolated antibody comprises the VH sequence shown in SEQ ID NO:470 and the VL sequence shown in SEQ ID NO:471; and wherein the human Fc region comprises the human IgG sequence shown in SEQ ID NO:460.

[0207] In certain embodiments, the isolated antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 446. In certain embodiments, the isolated antibody light chain comprises a constant light chain sequence as set forth in SEQ ID NO: 469.

[0208] In certain embodiments, the separated antibody Fc region comprises one or more amino acid substitutions, wherein compared to other equivalent antibodies comprising Fc without the one or more substitutions, the one or more substitutions cause changes (such as increases or decreases) in antibody half-life, ADCC activity, ADCP activity or CDC activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, changes are made to an increase in antibody half-life, an increase or decrease in ADCC activity, an increase in ADCP activity or an increase in CDC activity. In certain embodiments, compared to the antibody comprising wild-type Fc region, the one or more amino acid substitutions cause an increase in antibody half-life. In certain embodiments, the half-life of the separated antibody comprising Fc region with one or more amino acid substitutions in humans is about 80 to 110 days.

[0209] In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the changes are an increase or decrease in antibody half-life, an increase or decrease in ADCC activity, an increase or decrease in ADCP activity, or an increase or decrease in CDC activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the changes are an increase in antibody half-life. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the changes are a decrease in antibody half-life. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the changes are an increase in ADCC activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the changes are a decrease in ADCC activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the changes are an increase in ADCP activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the changes are a decrease in ADCP activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the change is an increase in CDC activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the change is a reduction in CDC activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the change is an increase in antibody half-life, an increase in ADCC activity, an increase in ADCP activity and an increase in CDC activity. In certain embodiments, compared to other equivalent antibodies comprising Fc without the one or more substitutions, the change is an increase in antibody half-life, a reduction in ADCC activity, an increase in ADCP activity and an increase in CDC activity.

[0210] In certain embodiments, the change is an increase in antibody half-life compared to an otherwise equivalent antibody comprising an Fc not having said one or more substitutions.

[0211] In certain embodiments, the change is an increase in ADCC activity compared to an otherwise equivalent antibody comprising an Fc not having said one or more substitutions.

[0212] In certain embodiments, the change is a decrease in ADCC activity compared to an otherwise equivalent antibody comprising an Fc not having said one or more substitutions.

[0213] In certain embodiments, the Fc region binds to the neonatal Fc receptor (FcRn). In certain embodiments, the Fc region binds to FcRn with a higher affinity at pH 6.0 than an antibody comprising a wild-type Fc region. In certain embodiments, the Fc region binds to FcRn with a higher affinity at pH 6.0 than an antibody comprising a wild-type Fc region.-7 M of K D Binds to FcRn.

[0214] In certain embodiments, the isolated antibody is a monoclonal antibody.

[0215] In certain embodiments, the antibody binds to the IL-13 sequence set forth in SEQ ID NOs: 472-475.

[0216] In certain embodiments, the isolated antibody has a density of less than or equal to about 1, 2, 3, 4, 5, 6, 7, 8, 9 x 10 -9 M of K D Binds to the IL-13 sequence set forth in SEQ ID NOs: 472-475. In certain embodiments, the isolated antibody has a specific affinity for IL-13 as measured by SPR at a concentration of less than or equal to about 1 x 10 -10 M of K D Binds to the IL-13 sequence set forth in SEQ ID NOs: 472-475. In certain embodiments, the antibody has a binding affinity of less than or equal to about 1 x 10 -9 M of K D Binds to human IL-13.

[0217] In certain embodiments, the isolated antibody exhibits a melting temperature greater than 68° C. as measured by Differential Scanning Fluorometry (DSF). In certain embodiments, the antibody exhibits a melting temperature greater than 75° C. as measured by DSF. In certain embodiments, the antibody exhibits an aggregation temperature equal to or greater than 71.2° C. as measured by DSF.

[0218] In certain embodiments, the isolated antibody has a retention time of 15.2 minutes or less as measured by hydrophobic interaction chromatography.

[0219] In certain embodiments, the isolated antibody does not have the heavy chain variable region sequence shown in SEQ ID NO:470.

[0220] In certain embodiments, the separated antibodies are used to treat inflammatory conditions or diseases. In certain embodiments, the separated antibodies are used to treat atopic dermatitis. In certain embodiments, the treatment reduces the severity of the disease in the subject and wherein the severity of the disease is assessed by atopic dermatitis disease severity outcome measurement. In certain embodiments, the separated antibodies are used to treat asthma. In certain embodiments, the separated antibodies are used to treat idiopathic pulmonary fibrosis. In certain embodiments, the separated antibodies are used to treat alopecia areata. In certain embodiments, the separated antibodies are used to treat chronic sinusitis with nasal polyps. In certain embodiments, the separated antibodies are used to treat chronic sinusitis without nasal polyps (CRSsNP). In certain embodiments, the separated antibodies are used to treat eosinophilic esophagitis (EoE). In certain embodiments, the isolated antibody is used to treat an eosinophilic gastrointestinal disorder or disease (ENID) selected from the group consisting of eosinophilic gastritis (EoG), eosinophilic enteritis (EoN), eosinophilic colitis (EoC), and eosinophilic gastroenteritis (EGE). In certain embodiments, the isolated antibody is used to treat Churg-Strauss syndrome / eosinophilic granulomatosis with polyangiitis (EGPA). In certain embodiments, the isolated antibody is used to treat prurigo nodularis (PN). In certain embodiments, the isolated antibody is used to treat chronic spontaneous urticaria (CSU). In certain embodiments, the isolated antibody is used to treat chronic pruritus of unknown origin (CPUO). In certain embodiments, the isolated antibody is used to treat bullous pemphigoid (BP). In certain embodiments, the isolated antibody is used to treat cold-induced urticaria (ColdU). In certain embodiments, the isolated antibodies are used to treat allergic fungal sinusitis (AFRS). In certain embodiments, the isolated antibodies are used to treat allergic bronchopulmonary aspergillosis (ABPA). In certain embodiments, the isolated antibodies are used to treat chronic obstructive pulmonary disease (COPD). In certain embodiments, the isolated antibodies are used to treat inflammatory bowel disease, such as Crohn's disease or ulcerative colitis. In certain embodiments, the isolated antibodies are used to treat psoriasis. In certain embodiments, the isolated antibodies are used to treat lupus. In certain embodiments, the isolated antibodies are used to treat rheumatoid arthritis.

[0221] In some aspects, described herein are isolated polynucleotides or polynucleotide groups encoding antibodies, VH thereof, VL thereof, light chains thereof, heavy chains thereof, or antigen-binding portions thereof, and optionally wherein said polynucleotides or polynucleotide groups comprise cDNA. In some aspects, described herein are vectors or vector groups comprising said polynucleotides or polynucleotide groups. In some aspects, described herein are a host cell comprising said polynucleotides or polynucleotide groups, or said vectors or vector groups.

[0222] In certain aspects, described herein is a method of making an antibody, the method comprising expressing the antibody using a host cell described herein and isolating the expressed antibody.

[0223] In certain aspects, described herein is a pharmaceutical composition comprising an antibody described herein and a pharmaceutically acceptable excipient.

[0224] In certain aspects, described herein is a kit comprising an antibody described herein or a pharmaceutical composition described herein, and instructions for use.

[0225] In certain aspects, described herein is a method for treating an inflammatory condition or disease in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody described herein or a pharmaceutical composition described herein. In certain embodiments of the methods described herein, the inflammatory condition or disease is atopic dermatitis. In certain embodiments, the inflammatory condition or disease is asthma. In certain embodiments, the inflammatory condition or disease is idiopathic pulmonary fibrosis. In certain embodiments, the inflammatory condition or disease is alopecia areata. In certain embodiments, the inflammatory condition or disease is chronic sinusitis with nasal polyps. In certain embodiments, the inflammatory condition or disease is chronic sinusitis without nasal polyps (CRSsNP). In certain embodiments, the inflammatory condition or disease is eosinophilic esophagitis (EoE). In certain embodiments, the inflammatory disorder or disease is an eosinophilic gastrointestinal disorder or disease (ENID) selected from the group consisting of eosinophilic gastritis (EoG), eosinophilic enteritis (EoN), eosinophilic colitis (EoC), and eosinophilic gastroenteritis (EGE). In certain embodiments, the inflammatory disorder or disease is Tschag-Strauss syndrome / eosinophilic granulomatosis with polyangiitis (EGPA). In certain embodiments, the inflammatory disorder or disease is prurigo nodularis (PN). In certain embodiments, the inflammatory disorder or disease is chronic spontaneous urticaria (CSU). In certain embodiments, the inflammatory disorder or disease is chronic pruritus of unknown origin (CPUO). In certain embodiments, the inflammatory disorder or disease is bullous pemphigoid (BP). In certain embodiments, the inflammatory disorder or disease is cold-induced urticaria (ColdU). In certain embodiments, the inflammatory condition or disease is allergic fungal sinusitis (AFRS). In certain embodiments, the inflammatory condition or disease is allergic bronchopulmonary aspergillosis (ABPA). In certain embodiments, the inflammatory condition or disease is chronic obstructive pulmonary disease (COPD). In certain embodiments, the inflammatory condition or disease is inflammatory bowel disease, such as Crohn's disease or ulcerative colitis. In certain embodiments, the inflammatory condition or disease is psoriasis. In certain embodiments, the inflammatory condition or disease is lupus. In certain embodiments, the inflammatory condition or disease is rheumatoid arthritis.

[0226] In certain aspects, described herein is a method for treating a pathology associated with elevated IL-13 levels in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody described herein or a pharmaceutical composition described herein.

[0227] In certain aspects, described herein is a method of reducing the biological activity of IL-13 in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody described herein or a pharmaceutical composition described herein.

[0228] In certain aspects, described herein is a method of inhibiting a TH2 type allergic response in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody described herein or a pharmaceutical composition described herein.

[0229] In certain aspects, described herein is a method of reducing thymus and activation-regulated chemokine (TARC) / CCL17 levels in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody described herein or a pharmaceutical composition described herein.

[0230] In certain aspects, described herein is a method of preventing an inflammatory disorder or disease in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody described herein or a pharmaceutical composition described herein. BRIEF DESCRIPTION OF THE DRAWINGS

[0231] These and other features, aspects and advantages of the present invention will be better understood with respect to the following description and accompanying drawings, in which:

[0232] Figure 1 3D rendering of human IL-13. The “1” gray highlights epitopes of 1eb rikizumab that overlap with the epitopes of construct 133 disclosed herein (see, e.g., Tables 2-8). These epitopes also overlap with the IL-4Rα epitope on IL-13 (shown in “2” gray). Tralokunumab-1drm (Adbry TM ) is shown in gray at "3". IL-13R α The overlapping epitope of IL-13R1 / IL-13R2 is shown in gray in “4”, and the IL-13R α 2 (non-overlapping) epitopes are shown in “5| in grey.

[0233] Figure 2 Graph depicting the percent inhibition of IL-13 binding to HEK293 cells overexpressing IL13Rα1 / IL-4Rα that had been incubated with the indicated engineered anti-IL-13 antibodies as determined by FACs.

[0234] Figure 3Graph depicting the percent inhibition of IL-13-induced STAT6 phosphorylation in HT-29 cells incubated with the indicated engineered anti-IL-13 antibodies as determined by FACs.

[0235] Figure 4 Graph depicting the percent inhibition of IL-13-induced thymic regulatory and activation-regulated chemokine (TARC / CCL17) release in supernatants of A549 cell cultures incubated with the indicated engineered anti-IL-13 antibodies as determined by enzyme-linked immunosorbent assay (ELISA).

[0236] Figure 5 Graph depicting the percent inhibition of IL-13-induced TARC release in supernatants of A549 cell cultures incubated with the indicated engineered anti-IL-13 antibodies as determined by ELISA.

[0237] Figure 6 Graph depicting the percent inhibition of IL-13-induced proliferation of TF-1 cells incubated with the indicated engineered anti-IL-13 antibodies as quantified by CellTiter-Glo assay.

[0238] Figure 7 Graph depicting the percent inhibition of IL-13-induced STAT6 phosphorylation in human peripheral blood mononuclear cells (PBMCs) incubated with the indicated engineered anti-IL-13 antibodies as determined by FACs.

[0239] Figure 8 Graph depicting the percent inhibition of IL-13-induced CD23 expression in human peripheral blood mononuclear cells (PBMCs) incubated with the indicated engineered anti-IL-13 antibodies as determined by FACs.

[0240] Fig. 9 Graph depicting serum concentrations (ng / mL) of construct 133 and lerezumab in non-human primates (NHPs) over time (days post injection). The half-life of construct 15 was 27.6 days compared to 17 to 18 days for lerezumab.

[0241] Fig.10 To depict the normalized AUC from dosing to infinity in the antibody with YTE substitution 0-∞ (C norm*day ) or area under the curve (AUC).

[0242] Fig.11 is a graph depicting serum concentrations (ng / mL) of the indicated engineered anti-IL-13 antibodies administered intravenously (IV) in NHPs.

[0243] Fig.12 Graph depicting serum concentrations (ng / mL) of the indicated engineered anti-IL-13 antibodies administered subcutaneously (SQ) in NHPs.

[0244] Fig.13 Graph depicting the percent inhibition of IL-13-induced CCL2 secretion in HaCaT cells incubated with the indicated engineered anti-IL-13 antibodies as determined by Luminex.

[0245] Fig.14 Graph depicting the percent inhibition of CCL26 secretion in HaCaT cells incubated with the indicated engineered anti-IL-13 antibodies as determined by Luminex.

[0246] Fig.15 Graph depicting the percent inhibition of NTRK1 gene expression in HaCaT cells incubated with the indicated engineered anti-IL-13 antibodies as determined by QuantiGene assay. DETAILED DESCRIPTION

[0247] IL-13 signaling begins with the binding of IL-13 to IL-13Rα1, forming an inactive complex, which then binds to IL-4Rα to form a fully active receptor heterodimer. This active receptor heterodimer contributes to the pathogenesis of atopic dermatitis. The present disclosure relates, in part, to anti-IL-13 antibodies that prevent this heterodimer formation.

[0248] like Figure 1 (3D rendering of human IL-13), "1" grey highlights epitopes of leromemab that overlap with epitopes of certain antibodies disclosed herein. Importantly, these epitopes also overlap with the IL-4Rα epitope on IL-13. Without wishing to be bound by theory, it is thought that antibodies binding to this region may prevent the formation of IL-13Rα1-IL-4Rα heterodimers, thereby limiting inflammatory signaling that leads to atopic dermatitis. In contrast, the tralokumab-ldrm (Adbry TM ) does not overlap with the IL-4Rα epitope on IL-13 and may therefore have a more limited ability to prevent heterodimerization.

[0249] definition

[0250] Unless otherwise defined, all technical terms, symbols and other scientific terms used herein are intended to have meanings commonly understood by those skilled in the art. In some cases, for clarity and / or convenient reference, terms with commonly understood meanings are defined herein, and including such definitions herein should not necessarily be interpreted as indicating differences commonly understood in the art. Techniques and procedures described or referenced herein are generally fully understood by those skilled in the art and are generally adopted using conventional methods, such as Sambrook et al., Molecular Cloning: A Laboratory Manual 4th edition (2012) Cold Spring Harb or Laboratory Press, Cold Spring Harbor, NY, a widely used molecular cloning method described. Where appropriate, unless otherwise noted, procedures involving the use of commercial kits and reagents are generally performed according to the protocols and conditions defined by the manufacturer.

[0251] As used herein, the singular form "a," "an," and "the" include plural referents unless otherwise indicated.

[0252] It is to be understood that aspects and embodiments of the present invention described herein include "comprising," "consisting of," and "consisting essentially of" aspects and embodiments.

[0253] For all compositions described herein and all methods of using the compositions described herein, the composition may comprise the listed components or steps, or may "consist essentially of" the listed components or steps. When a composition is described as "consisting essentially of" the listed components, the composition contains the listed components and may contain other components that do not substantially affect the condition being treated, but does not contain any other components that substantially affect the condition being treated other than those components explicitly listed; or if the composition contains additional components other than those listed that substantially affect the condition being treated, the composition does not contain the additional components at sufficient concentration or amount to substantially affect the condition being treated. When a method is described as "consisting essentially of" the listed steps, the method contains the listed steps and may contain other steps that do not substantially affect the condition being treated, but the method does not contain any other steps that substantially affect the condition being treated other than those steps explicitly listed. As a non-limiting specific example, when a composition is described as "consisting essentially of" a component, the composition may additionally contain any amount of pharmaceutically acceptable carriers, vehicles or diluents and other such components that do not substantially affect the condition being treated.

[0254] As used herein, the term "vector" refers to a nucleic acid molecule capable of propagating another nucleic acid to which it is linked. The term includes vectors that are self-replicating nucleic acid structures as well as vectors that are incorporated into the genome of a host cell into which they have been introduced. Certain vectors are capable of directing the expression of nucleic acids to which they are operably linked. Such vectors are referred to herein as "expression vectors."

[0255] The terms "host cell", "host cell line" and "host cell culture" are used interchangeably and refer to cells into which exogenous nucleic acids have been introduced and the progeny of such cells. Host cells include "transformants" (or "transformed cells") and "transfectants" (or "transfected cells"), each of which includes the primary transformed or transfected cells and the progeny derived therefrom. The nucleic acid content of such progeny may not be exactly the same as that of the parent cell, and may contain mutations. "Recombinant host cell" or "host cell" refers to a cell that includes an exogenous polynucleotide, regardless of the method used for insertion (e.g., direct uptake, transduction, f-mating, or other methods known in the art for producing recombinant host cells).

[0256] As used herein, the term "eukaryote" refers to organisms belonging to the phylogenetic domain Eukaryota, such as animals (including but not limited to mammals, insects, reptiles, birds, etc.), ciliates, plants (including but not limited to monocots, dicots, algae, etc.), fungi, yeasts, flagellates, microsporidia, protists, etc.

[0257] As used herein, the term "prokaryote" refers to a prokaryotic organism. For example, the non-eukaryotic organism can belong to the Eubacteria (including but not limited to Escherichia coli, Thermus thermophilus, Bacillus stearothermophilus, Pseudomonasfluorescens, Pseudomonas aeruginosa, Pseudomonas putida, etc.) phylogenetic domain or the Archaea (including but not limited to Methanococcusjannaschii, Methanobacterium thermoautotrophicum, Halobacterium (such as Haloferax volcanii and Halobacterium species NRC-1), Archaeoglobus fulgidus, Pyrococcus furiosus, Pyrococcus horikoshii, Aeurop yrum pernix) etc.) phylogenetic domain.

[0258] As used herein, "effective amount" or "therapeutically effective amount" refers to an amount of a therapeutic compound (e.g., an anti-IL-13 antibody) administered to an individual as a single dose or as part of a series of doses (alone or in combination with another therapeutic modality) that is effective to produce or promote a desired therapeutic effect. Examples of desired therapeutic effects are enhancing immune response; slowing or delaying tumor development; stabilizing disease; improving one or more symptoms. An effective amount can be given in one or more doses.

[0259] The term "treating" (and variations thereof, such as "treat" or "treatment") refers to clinical intervention in an attempt to alter the natural course of a disease or disorder in a subject in need thereof. Treatment may be performed during the course of clinical pathology. Desirable effects of treatment include preventing disease recurrence, alleviating symptoms, alleviating any direct or indirect pathological consequences of the disease, preventing cancer metastasis, slowing the rate of disease progression, ameliorating or palliating the disease state, and alleviating or improving prognosis.

[0260] The term "sufficient amount" means an amount sufficient to produce a desired effect, such as an amount sufficient to modulate an immune response in a subject.

[0261] As used herein, the term "subject" or "individual" means a mammalian subject. Exemplary subjects include humans, monkeys, dogs, cats, mice, rats, cattle, horses, camels, goats, rabbits, and sheep. In certain embodiments, the subject is human. In some embodiments, the subject suffers from a disease or illness that can be treated with the antibodies provided herein. In some embodiments, the disease or illness is cancer. In some embodiments, the disease or illness is a viral infection.

[0262] The term "in vitro" refers to processes occurring in living cells grown separately from living organisms, such as in tissue culture.

[0263] The term "in vivo" refers to processes that occur within a living organism.

[0264] The term "package insert" is used to refer to instructions customarily included in commercial packages of therapeutic or diagnostic products (e.g., kits), that contain information about the indications, usage, dosage, administration, combination therapy, contraindications and / or warnings concerning the use of such therapeutic or diagnostic products.

[0265] The term "pharmaceutical composition" refers to a preparation that is in such form as to permit the biological activity of the active ingredients contained therein to be effective in treating a subject and contains no additional components that are unacceptably toxic to the subject in the amounts provided in the pharmaceutical composition.

[0266] The terms "co-administration", "co-administer" and "incombination with" include the simultaneous, concurrent or sequential administration of two or more therapeutic agents within unspecified time limits. In one embodiment, the agents are present in a cell or in a subject at the same time, or exert their biological or therapeutic effects at the same time. In one embodiment, the therapeutic agents are in the same composition or unit dosage form. In other embodiments, the therapeutic agents are in separate compositions or unit dosage forms. In certain embodiments, the first agent may be administered prior to the administration of the second therapeutic agent.

[0267] The terms "modulate" and "modulation" refer to the decrease or inhibition or the activation or increase of the recited variable.

[0268] The terms "increase" and "activate" refer to an increase of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 50-fold, 100-fold or more of the recited variable.

[0269] The terms "reduce" and "inhibit" refer to a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 50-fold, 100-fold or more reduction in the recited variable.

[0270] The term "about" indicates and encompasses the indicated value and the range above and below the value. In certain embodiments, the term "about" indicates a specified value ± 10%, ± 5% or ± 1%. In certain embodiments, where applicable, the term "about" indicates a specified value ± one standard deviation of the value.

[0271] The term "agonism" refers to the activation of receptor signaling to induce a biological response associated with receptor activation. An "agonist" is an entity that binds to and agonizes a receptor.

[0272] The term "antagonism" refers to the inhibition of receptor signaling to inhibit the biological response associated with receptor activation. An "antagonist" is an entity that binds to and antagonizes a receptor.

[0273] For any of the structural and functional characteristics described herein, methods of determining these characteristics are known in the art.

[0274] The term "optionally" when used in sequence is meant to include one to all of the listed combinations and encompass all sub-combinations.

[0275] The term "amino acid" refers to the twenty common naturally occurring amino acids. Naturally occurring amino acids include alanine (Ala; A), arginine (Arg; R), asparagine (Asn; N), aspartic acid (Asp; D), cysteine ​​(Cys; C); glutamic acid (Glu; E); glutamine (Gln; Q); glycine (Gly; G); histidine (His; H); isoleucine (Ile; I); leucine (Leu; L); lysine (Lys; K); methionine (Met; M); phenylalanine (Phe; F); proline (Pro; P); serine (Ser; S); threonine (Thr; T); tryptophan (Trp; W); tyrosine (Tyr; Y); and valine (Val; V).

[0276] The term "affinity" refers to the total non-covalent interaction strength between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen or epitope). Unless otherwise indicated, as used herein, "affinity" refers to intrinsic binding affinity, which reflects a 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen or epitope).

[0277] As used herein, the term "kd" (sec-1) refers to the dissociation rate constant for a specific antibody-antigen interaction. This value is also known as the koff value.

[0278] As used herein, the term "ka" (Ml x sec-1) refers to the association rate constant for a specific antibody-antigen interaction. This value is also known as the kon value.

[0279] As used herein, the term "KD" (M) refers to the dissociation equilibrium constant of a particular antibody-antigen interaction. KD = kd / ka. In some embodiments, the affinity of an antibody is described in terms of the KD of the interaction between such an antibody and its antigen. For clarity, as known in the art, a smaller KD value indicates a higher affinity interaction, while a larger KD value indicates a lower affinity interaction.

[0280] As used herein, the term "KA" (M- 1 ) refers to the binding equilibrium constant of a specific antibody-antigen interaction. KA = ka / kd.

[0281] The term "antibody" is used herein in its broadest sense and includes certain types of immunoglobulin molecules comprising one or more antigen binding domains that specifically bind to an antigen or epitope. Antibodies particularly include complete antibodies (e.g., complete immunoglobulins), antibody fragments, and multispecific antibodies.

[0282] An "anti-IL-13 antibody," "IL-13 antibody," or "IL-13-specific antibody" is an antibody that specifically binds to the antigen IL-13 as provided herein.

[0283] The term "epitope" means the portion of an antigen to which an antibody specifically binds.

[0284] As used herein, the term "hypervariable region" or "HVR" refers to the regions of an antibody variable domain that are hypervariable in sequence and / or form structurally defined loops ("hypervariable loops").

[0285] The term "antigen binding domain" means the portion of an antibody that is capable of specifically binding to an antigen or epitope.

[0286] The term "chimeric antibody" refers to an antibody in which a portion of the heavy chain and / or light chain is derived from a particular source or species, while the remainder of the heavy chain and / or light chain is derived from a different source or species.

[0287] The term "human antibody" refers to an antibody having an amino acid sequence corresponding to an antibody produced by a human or human cell or derived from an amino acid sequence of a non-human source utilizing a human antibody repertoire or human antibody encoding sequence (e.g., obtained from a human source or redesigned). Human antibodies particularly exclude humanized antibodies.

[0288] The term "humanized antibody" refers to a protein that has a sequence that differs from the sequence of an antibody derived from a non-human species by one or more amino acid substitutions, deletions and / or additions, such that when administered to a human subject, the humanized antibody is less likely to induce an immune response and / or induces a less severe immune response than the non-human species antibody.

[0289] The term "multispecific antibody" refers to an antibody comprising two or more different antigen binding domains that collectively specifically bind to two or more different epitopes.

[0290] A "monospecific antibody" is an antibody that contains one or more binding sites that specifically bind to a single epitope. An example of a monospecific antibody is a naturally occurring IgG molecule, which, although bivalent (i.e., having two antigen binding domains), recognizes the same epitope at each of the two antigen binding domains. The binding specificity may exist at any suitable valency.

[0291] The term "monoclonal antibody" refers to an antibody from a population of substantially homogeneous antibodies. A population of substantially homogeneous antibodies comprises antibodies that are substantially similar and bind to the same epitope, except for variants that may generally occur during the production of monoclonal antibodies. Such variants are generally present only in small amounts. Monoclonal antibodies are typically obtained by methods including selecting a single antibody from a variety of antibodies. For example, a selection method may be selected from a variety of clones, such as a group of hybridoma clones, phage clones, yeast clones, bacterial clones or other recombinant DNA clones to select a unique clone. The selected antibody may be further changed, for example, to improve affinity for the target ("affinity maturation"), to humanize the antibody, to improve its yield in cell culture, and / or to reduce its immunogenicity in a subject.

[0292] The term "single chain" refers to a molecule comprising amino acid monomers linearly connected by peptide bonds. In a specific embodiment of this type, in a single-chain Fab molecule, the C-terminus of the Fab light chain is connected to the N-terminus of the Fab heavy chain. As described in more detail herein, scFv has a light chain variable domain (VL) connected to the N-terminus of the heavy chain variable domain (VH) from its C-terminus by a polypeptide chain. Alternatively, scFv is composed of a polypeptide chain in which the C-terminus of VH is connected to the N-terminus of VL by a polypeptide chain.

[0293] "Fab fragments" (also called fragment antigen binding) contain the constant domain of the light chain (CL) and the first constant domain of the heavy chain (CH1) and the variable domains VL and VH on the light and heavy chains, respectively. The variable domains contain the complementarity determining loops (CDRs, also called hypervariable regions) involved in antigen binding. Fab' fragments differ from Fab fragments by the addition of a few residues at the carboxyl terminus of the heavy chain CH1 domain including one or more cysteines from the antibody hinge region.

[0294] "F(ab')2" fragments contain two Fab' fragments joined by a disulfide bond near the hinge region. F(ab')2 fragments can be produced, for example, by recombinant methods or by pepsin digestion of intact antibodies. F(ab') fragments can be dissociated, for example, by treatment with B-mercaptoethanol.

[0295] An "Fv" fragment comprises a non-covalently linked dimer of one heavy- and one light-chain variable domain.

[0296] "Single-chain Fv" or "sFv" or "scFv" includes the VH and VL domains of an antibody, wherein these domains are present in a single polypeptide chain. In one embodiment, the Fv polypeptide further comprises a polypeptide linker between the VH and VL domains, which enables the scFv to form the desired structure for antigen binding. For a review of scFv, see Pluckthun, The Pharmacology of Monoclonal Antibodies, Vol. 113, Rosenburg and Moore, ed., Springer-Verlag, New York, pp. 269-315 (1994). HER2 antibody scFv fragments are described in WO93 / 16185; U.S. Pat. No. 5,571,894; and U.S. Pat. No. 5,587,458.

[0297] "scFv-Fc" fragments comprise a scFv connected to an Fc domain. For example, the Fc domain can be connected to the C-terminus of the scFv. Depending on the orientation of the variable domains in the scFv (i.e., VH-VL or VL-VH), the Fc domain can be after the VH or VL. Any suitable Fc domain known in the art or described herein can be used. In some cases, the Fc domain comprises an IgG4 Fc domain.

[0298] The term "single domain antibody" or "sdAb" refers to a molecule in which one variable domain of an antibody specifically binds to an antigen in the absence of other variable domains. Single domain antibodies and fragments thereof are described in Arabi Ghahroudi et al., FEBS Letters, 1998, 414: 521-526 and Muyldermans et al., Trends in Biochem. Sci., 2001, 26: 230-245, each of which is incorporated by reference in its entirety. Single domain antibodies are also referred to as sdAbs or nanobodies. SdAbs are extremely stable and are easily expressed with the Fc chain of an antibody as a fusion partner (Harmsen MM, De Haard HJ (2007). "Properties, production, and applications of camelid single-domain antibody fragments". Appl. Microbiol Biotechnol. 77 (1): 13-22).

[0299] The terms "full-length antibody," "intact antibody," and "complete antibody" are used interchangeably herein to refer to an antibody having a structure substantially similar to a naturally occurring antibody structure and having heavy chains comprising an Fc region. For example, when used to refer to an IgG molecule, a "full-length antibody" is an antibody comprising two heavy chains and two light chains.

[0300] The term "antibody fragment" refers to an antibody comprising a portion of an intact antibody, such as the antigen binding region or variable region of an intact antibody. Antibody fragments include, for example, Fv fragments, Fab fragments, F(ab')2 fragments, Fab' fragments, scFv (sFv) fragments, and scFv-Fc fragments.

[0301] The term "Fc domain" or "Fc region" is used herein to define the C-terminal region of an immunoglobulin heavy chain that contains at least a portion of the constant region. The term includes native sequence Fc regions and variant Fc regions.

[0302] The term "substantially purified" refers to a construct described herein or variants thereof that are substantially or essentially free of components that normally accompany or interact with the protein as found in its naturally occurring environment (i.e., native cells; or in the case of recombinantly produced antibodies, host cells, which in certain embodiments are essentially free of cellular material), including preparations of protein having less than about 30%, less than about 25%, less than about 20%, less than about 15%, less than about 10%, less than about 5%, less than about 4%, less than about 3%, less than about 2%, or less than about 1% (by dry weight) of contaminating protein.

[0303] In the case of two or more nucleic acid or polypeptide sequences, the term percentage "identity" refers to two or more sequences or subsequences having a specified percentage of identical nucleotides or amino acid residues when compared and aligned for maximum correspondence, as measured using one of the sequence comparison algorithms described below (e.g., using publicly available computer software such as BLAST, BLASTP, BLASTN, BLAST-2, ALIGN, MEGALIGN (DNASTAR), CLUSTALW, CLUSTAL OMEGA or MUSCLE software or other algorithms available to those skilled in the art) or by visual inspection. Software for performing BLAST analysis is publicly available via the National Center for Biotechnology Information (ncbi.n1m.nih.gov). One skilled in the art can determine the parameters suitable for aligning sequences, including any algorithm required to achieve maximum alignment over the full length of the compared sequences. Depending on the application, the percentage "identity" may be present in a region of the compared sequences, such as within a functional domain, or within the full length of the two sequences to be compared.

[0304] Regarding sequence comparison, usually a sequence serves as a reference sequence compared to a test sequence. When using a sequence comparison algorithm, the test and reference sequences are input into a computer, subsequence coordinates are specified if necessary, and sequence algorithm program parameters are specified. Then, the sequence comparison algorithm calculates the sequence identity percentage of the test sequence relative to the reference sequence based on the specified program parameters.

[0305] Sequences for comparison can be optimally aligned, for example, by the local homology algorithm of Smith and Waterman, Adv. Appl. Math. 2:482 (1981); the homology alignment algorithm of Needleman and Wunsch, J Mol. Biol. 48:443 (1970); the similarity search method of Pearson and Lipman, Proc. Nat'l. Acad. Sci. USA 85:2444 (1988); computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, Wis.); or by visual inspection (see generally, Ausubel et al., see below).

[0306] The ranges recited herein are to be understood as shorthand for all values ​​within the range, including the recited endpoints. For example, a range of 1 to 50 is to be understood to include any value, combination of values, or subrange from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, and 50.

[0307] It must be noted that, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0308] Anti-IL-13 antibodies

[0309] Antibody structure

[0310] The present application provides antibodies and compositions comprising antibodies that bind IL-13.

[0311] The immunoglobulin genes identified include kappa, lambda, alpha, gamma, delta, epsilon and mu constant region genes and a variety of immunoglobulin variable region genes. Light chains are classified as kappa or lambda. The "class" of an antibody or immunoglobulin refers to the type of constant domain or constant region possessed by its heavy chain. There are five major classes of antibodies: IgA, IgD, IgE, IgG and IgM, and some of these antibodies can be further divided into subclasses (isotypes), such as IgG1, IgG2, IgG3, IgG4, IgA1 and IgA2. The heavy chain constant domains corresponding to different classes of immunoglobulins are referred to as α, δ, epsilon, γ and μ, respectively.

[0312] An exemplary immunoglobulin (antibody) structural unit is composed of two pairs of polypeptide chains, each pair having a "light" chain (about 25kD) and a "heavy" chain (about 50-70kD). The N-terminal domain of each chain defines a variable region of about 100 to 110 or more amino acids, which is primarily responsible for antigen recognition. The terms variable light chain (VL) and variable heavy chain (VH) refer to these light chain and heavy chain domains, respectively. The IgG1 heavy chain is composed of VH, CH1, CH2, and CH3 domains from the N-terminus to the C-terminus, respectively. The light chain is composed of VL and CL domains from the N-terminus to the C-terminus. The IgG1 heavy chain comprises a hinge between the CH1 and CH2 domains. In certain embodiments, the immunoglobulin construct comprises at least one immunoglobulin domain from IgG, IgM, IgA, IgD, or IgE connected to a therapeutic polypeptide. In some embodiments, the immunoglobulin domains found in the antibodies provided herein are from or derived from immunoglobulin-based constructs, such as bifunctional antibodies or nanobodies. In certain embodiments, the immunoglobulin constructs described herein comprise at least one immunoglobulin domain from a heavy chain antibody such as a camel antibody. In certain embodiments, the immunoglobulin constructs provided herein comprise at least one immunoglobulin domain from a mammalian antibody such as a bovine antibody, a human antibody, a camel antibody, a mouse antibody, or any chimeric antibody.

[0313] In some embodiments, the antibodies provided herein comprise a heavy chain. In one embodiment, the heavy chain is IgA. In one embodiment, the heavy chain is IgD. In one embodiment, the heavy chain is IgE. In one embodiment, the heavy chain is IgG. In one embodiment, the heavy chain is IgM. In one embodiment, the heavy chain is IgG1. In one embodiment, the heavy chain is IgG2. In one embodiment, the heavy chain is IgG3. In one embodiment, the heavy chain is IgG4. In one embodiment, the heavy chain is IgA1. In one embodiment, the heavy chain is IgA2.

[0314] In some embodiments, the antibody is an IgG1 antibody. In some embodiments, the antibody is an IgG3 antibody. In some embodiments, the antibody is an IgG2 antibody. In some embodiments, the antibody is an IgG4 antibody.

[0315] Typically, native four-chain antibodies contain six hypervariable regions (HVRs); three in VH (H1, H2 and H3), and three in VL (L1, L2 and L3). HVRs typically contain amino acid residues from hypervariable loops and / or from complementarity determining regions (CDRs), the latter having the highest sequence variability and / or being involved in antigen recognition. In addition to CDR1 in VH, CDRs typically contain amino acid residues that form hypervariable loops. HVRs are also referred to as CDRs, and these terms are used interchangeably herein with respect to portions of the variable regions that form antigen binding regions. This particular region has been described by Kabat et al., USDept. of Health and Human Services, Sequences of Proteins of Immunological Interest (1983) and Chothia et al., J Mol Biol 196: 901-917 (1987), wherein each definition includes overlaps or subsets of amino acid residues when compared to each other. However, the application of any definition of CDRs involving antibodies or variants thereof is intended to be within the scope of the terms as defined and used herein. The exact number of residues encompassing a particular CDR will vary depending on the CDR sequence and size. Given the variable region amino acid sequence of an antibody, one skilled in the art can routinely determine which residues comprise a particular CDR.

[0316] The amino acid sequence boundaries of a CDR can be determined by one of skill in the art using any of a variety of known numbering schemes, including those described by Kabat et al., supra ("Kabat" numbering scheme); Al-Lazikani et al., 1997, J Mol. Biol., 273:927-948 ("Chothia" numbering scheme); MacCallum et al., 1996, J Mol. Biol. 262:732-745 ("Contact" numbering scheme); Lefranc et al., Dev. Comp. Immunol., 2003, 27:55-77 ("IMGT" numbering scheme); and Honegge and Plückthun, J. Mol. Biol., 2001, 309:657-70 ("AHo" numbering scheme); each of which is incorporated by reference in its entirety.

[0317] Table 1 provides the positions of CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2 and CDR-H3 identified by the Kabat and Chothia schemes. For CDR-H1, the residue numbering is provided using both the Kabat and Chothia numbering schemes.

[0318] CDRs may be assigned, for example, using antibody numbering software, such as Abnum, available at www.bioinf.org.uk / abs / abnum / and described in Abhinandan and Martin, Immunology, 2008, 45:3832-3839, incorporated by reference in its entirety.

[0319] Table 1. Residues in the CDRs according to the Kabat and Chothia numbering schemes.

[0320]

[0321]

[0322] *When numbering using the Kabat numbering convention, the C-terminus of CDR-H1 varies between H32 and H34, depending on the length of the CDR.

[0323] When referring to residues in the antibody heavy chain constant region, the "EU numbering scheme" is generally used (eg, as reported in Kabat et al., supra). Unless otherwise stated, the EU numbering scheme is used to refer to residues in the antibody heavy chain constant region described herein.

[0324] An example of an antigen binding domain is an antigen binding domain formed by a VH-VL dimer of an antibody. Another example of an antigen binding domain is an antigen binding domain formed by diversification of certain loops from the tenth fibronectin type III domain of Adnectin. The antigen binding domain may include CDRs 1, 2, and 3 from the heavy chain, in the order described; and CDRs 1, 2, and 3 from the light chain, in the order described.

[0325] Epitopes are often composed of surface accessible amino acid residues and / or sugar side chains and may have specific three-dimensional structural features and charge-to-mass ratio characteristics. The difference between conformational and non-conformational epitopes is that in the presence of denaturing solvents, binding to the former but not the latter may be lost. The epitope may include amino acid residues that are directly involved in binding and other amino acid residues that are not directly involved in binding. Epitopes that bind to antibodies can be determined using known techniques for epitope determination, such as testing antibodies for binding to IL-13 variants with different point mutations or to chimeric IL-13 variants.

[0326] To screen for antibodies that bind to an epitope on an antigen of interest (e.g., IL-13) to which an antibody of interest binds, a conventional cross-blocking assay, such as that described in Antibodies, A Laboratory Manual, Cold Spring Harbor Laboratory, Ed Harlow and David Lane (1988) can be performed. Alternatively or additionally, epitope mapping can be performed by methods known in the art.

[0327] Chimeric antibodies are antibodies in which a portion of the heavy chain and / or light chain is derived from a particular source or species, while the remainder of the heavy chain and / or light chain is derived from a different source or species.

[0328] A human antibody is an antibody having an amino acid sequence corresponding to an antibody produced by a human or human cell or derived from an amino acid sequence of a non-human source utilizing a human antibody repertoire or human antibody encoding sequence (e.g., obtained from a human source or de novo). Human antibodies specifically exclude humanized antibodies.

[0329] Humanized antibodies have sequences that differ from the sequences of antibodies derived from non-human species by one or more amino acid substitutions, deletions and / or additions, so that when they are applied to human subjects, compared to non-human species antibodies, humanized antibodies are unlikely to induce immune responses and / or induce less severe immune responses. In one embodiment, certain amino acids in the framework and constant domain of the heavy and / or light chains of non-human species antibodies are mutated to produce humanized antibodies. In another embodiment, the constant domains from human antibodies are fused to the variable domains of non-human species. In another embodiment, one or more amino acid residues in one or more CDR sequences of non-human antibodies are changed to reduce the possible immunogenicity of the antibodies when non-human antibodies are applied to human subjects, wherein the amino acid residues changed are not critical for the immunospecific binding of antibodies to their antigens, or the changes in the amino acid sequences performed are conservative changes, so that the binding of humanized antibodies to antigens is not significantly worse than the binding of non-human antibodies to antigens. Examples of how to prepare humanized antibodies can be found in U.S. Pat. Nos. 6,054,297, 5,886,152, and 5,877,293. For additional details, see Jones et al., Nature, 1986, 321: 522-525; Riechmann et al., Nature, 1988, 332: 323-329; and Presta, Curr. Op. Struct. Biol., 1992, 2: 593-596, each of which is incorporated by reference in its entirety.

[0330] The two or more different epitopes can be epitopes on the same antigen (e.g., a single IL-13) or on different antigens (e.g., different IL-13 molecules, or IL-13 molecules and non-IL-13 molecules). In some embodiments, the multispecific antibody binds to two different epitopes (i.e., a "bispecific antibody"). In some embodiments, the multispecific antibody binds to three different epitopes (i.e., a "trispecific antibody").

[0331] Anti-IL-13 antibodies may include those described herein, such as clones shown in the accompanying drawings and / or tables. In some embodiments, the antibodies comprise alternative scaffolds. In some embodiments, the antibodies consist of alternative scaffolds. In some embodiments, the antibodies consist essentially of alternative scaffolds. In some embodiments, the antibodies comprise antibody fragments. In some embodiments, the antibodies consist of antibody fragments. In some embodiments, the antibodies consist essentially of antibody fragments.

[0332] In some embodiments, the antibody is a monoclonal antibody.

[0333] In some embodiments, the antibody is a polyclonal antibody.

[0334] In some embodiments, the antibodies are produced by hybridomas. In other embodiments, the antibodies are produced by recombinant cells engineered to express the desired variable and constant domains.

[0335] In some embodiments, the antibody may be a single chain antibody or other antibody derivative that retains antigen specificity and the lower hinge region or a variant thereof.

[0336] In some embodiments, the antibody may be a multifunctional antibody, a recombinant antibody, a human antibody, a humanized antibody, a fragment or a variant thereof. In a specific embodiment, the antibody fragment or derivative thereof is selected from a Fab fragment, a Fab'2 fragment, a CDR and a ScFv.

[0337] In some embodiments, the antibody is capable of forming an immune complex. For example, the immune complex can be a tumor cell coated with the antibody.

[0338] Regarding sequence comparison, usually a sequence serves as a reference sequence compared to a test sequence. When using a sequence comparison algorithm, the test and reference sequences are input into a computer, subsequence coordinates are specified if necessary, and sequence algorithm program parameters are specified. Then, the sequence comparison algorithm calculates the sequence identity percentage of the test sequence relative to the reference sequence based on the specified program parameters.

[0339] Sequences for comparison can be optimally aligned, for example, by the local homology algorithm of Smith and Waterman, Adv. Appl. Math. 2:482 (1981); the homology alignment algorithm of Needleman and Wunsch, J Mol. Biol. 48:443 (1970); the similarity search method of Pearson and Lipman, Proc. Nat'l. Acad. Sci. USA 85:2444 (1988); computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, Wis.); or by visual inspection (see generally, Ausubel et al., see below).

[0340] One example of an algorithm suitable for determining percent sequence identity and sequence similarity is the BLAST algorithm, which is described in Altschul et al., J Mol. Biol. 215:403-410 (1990). Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information (www.ncbi.nlm.nih.gov / ).

[0341] Sequence of IL-13 antibody

[0342] VH domain

[0343] In some embodiments, the antibodies provided herein comprise a VH sequence selected from SEQ ID NOs: 1-32 and 470.

[0344] In some embodiments, the antibodies provided herein comprise a VH sequence that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the illustrative VH sequences provided in SEQ ID NOs: 1-32 and 470. In some embodiments, the antibodies provided herein comprise a VH sequence that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the illustrative VH sequences provided in SEQ ID NOs: 1-32 and 470. HSequence, it has at most 1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19 or 20 amino acid substitutions.In some embodiments, the amino acid substitution is a conservative amino acid substitution.In some embodiments, the antibody described in this paragraph is referred to as "variant" in this article.In some embodiments, such variants are derived from the sequence provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis or any other method known in the art or described herein.In some embodiments, such variants are not derived from the sequence provided herein, and can be separated again, for example, according to the method for obtaining antibodies provided herein.

[0345] VL domain

[0346] In some embodiments, the antibodies provided herein comprise a VL sequence selected from SEQ ID NOs: 33-57 and 471.

[0347] In some embodiments, the antibodies provided herein comprise a VL sequence having at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to the illustrative VL sequences provided in SEQ ID NOs: 33-57 and 471. In some embodiments, the antibodies provided herein comprise a VL sequence provided in SEQ ID NOs: 33-57 and 471 having at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0348] VH-VL combination

[0349] In some embodiments, the antibodies provided herein comprise a VH sequence selected from SEQ ID NOs: 1-32 and 470; and a VL sequence selected from SEQ ID NOs: 33-57 and 471, such as the VH-VL combinations shown in Table 2 below.

[0350] In certain aspects, any one of SEQ ID NOs: 1-32 and 470 can be combined with any one of SEQ ID NOs: 33-57 and 471.

[0351] In certain embodiments, the antibody comprises a VH sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 1-32 and 470 and a VL sequence shown in SEQ ID NO:49.

[0352] In certain embodiments, the antibody comprises a VH sequence selected from the group consisting of SEQ ID NOs: 1-32 and 470 and a VL sequence as set forth in SEQ ID NO:51.

[0353] In some embodiments, the antibodies provided herein comprise a VH sequence that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the illustrative VH sequences provided in SEQ ID NOs: 1-32 and 470; and a VL sequence that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the illustrative VL sequences provided in SEQ ID NOs: 33-57 and 471. In some embodiments, the antibodies provided herein comprise VH sequences provided in SEQ ID NOs: 1-32 and 470 having at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acid substitutions; and VL sequences provided in SEQ ID NOs: 33-57 and 471 having at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0354] In some embodiments, the antibodies provided herein comprise a VH sequence and a VL sequence selected from the combination shown in Table 2 below. In certain embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO: 1 and a VL sequence as shown in SEQ ID NO: 33. In certain embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO: 2 and a VL sequence as shown in SEQ ID NO: 33. In certain embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 35. In certain embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO: 4 and a VL sequence as shown in SEQ ID NO: 35. In certain embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO: 5 and a VL sequence as shown in SEQ ID NO: 35. In certain embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO: 6 and a VL sequence as shown in SEQ ID NO: 35. In certain embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 35. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:36. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:4 and a VL sequence as set forth in SEQ ID NO:36. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:5 and a VL sequence as set forth in SEQ ID NO:36. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:6 and a VL sequence as set forth in SEQ ID NO:36. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:36. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:4 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:5 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 6 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 39.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:40. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:4 and a VL sequence as set forth in SEQ ID NO:40. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:5 and a VL sequence as set forth in SEQ ID NO:40. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:6 and a VL sequence as set forth in SEQ ID NO:40. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:40. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:8 and a VL sequence as set forth in SEQ ID NO:42. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:9 and a VL sequence as set forth in SEQ ID NO:43. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 44. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 45. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 46. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 47. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 48. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 49. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 50. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 51. In certain embodiments, the antibody comprises the VH sequence set forth in SEQ ID NO:7 and the VL sequence set forth in SEQ ID NO:52.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:53. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:54. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:55. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:56. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:57. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:10 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:11 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:12 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 13 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences as set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 14 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 15 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 16 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 17 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 18 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 19 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 20 and a VL sequence as set forth in SEQ ID NO: 39.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 21 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 22 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 23 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 24 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 25 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 26 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 27 and a VL sequence as set forth in SEQ ID NO: 39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:28 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:28 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:29 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:30 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:31 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:32 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:8 and a VL sequence as set forth in SEQ ID NO:39. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:8 and a VL sequence as set forth in SEQ ID NO:51. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 3 and a VL sequence as set forth in SEQ ID NO: 51. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471.

[0355] In certain embodiments, the isolated antibody comprises a heavy chain variable domain comprising a framework region sequence selected from the group consisting of: SEQ ID NO: 198-229, 255-256, 258-259, 261-285, 311-315, 317-342, 368-369, 371-399 and 540-580. In certain embodiments, the isolated antibody comprises a heavy chain variable domain comprising 1, 2, 3 or 4 framework region sequences selected from the group consisting of: SEQ ID NO: 198-229, 255-256, 258-259, 261-285, 311-315, 317-342, 368-369, 371-399 and 540-580.

[0356] In certain embodiments, the isolated antibody comprises a light chain variable domain comprising a framework region sequence selected from the group consisting of SEQ ID NOs: 230-231, 233-235, 239, 241-254, 286, 288, 290-291, 293, 296-310, 343-345, 347, 400-424, and 581-609. In certain embodiments, the isolated antibody comprises a light chain variable domain comprising 1, 2, 3 or 4 framework region sequences selected from the group consisting of SEQ ID NOs: 230-231, 233-235, 239, 241-254, 286, 288, 290-291, 293, 296-310, 343-345, 347, 400-424, and 581-609.

[0357] In certain embodiments, the isolated antibody comprises a heavy chain variable domain comprising 1, 2, 3 or 4 framework region sequences selected from the group consisting of: SEQ ID NOs: 198-229, 255-256, 258-259, 261-285, 311-315, 317-342, 368-369, 371-399 and 540-580, and comprises a light chain variable domain comprising 1, 2, 3 or 4 framework region sequences selected from the group consisting of: SEQ ID NOs: 230-231, 233-235, 239, 241-254, 286, 288, 290-291, 293, 296-310, 343-345, 347, 400-424 and 581-609.

[0358] Table 2. Anti-IL-13 antibody VH-VL sequences

[0359]

[0360]

[0361]

[0362]

[0363]

[0364]

[0365]

[0366]

[0367]

[0368]

[0369]

[0370]

[0371]

[0372]

[0373]

[0374] *Names correspond to those in the informal sequence listing

[0375] In some embodiments, such an IgG4-SP HC constant domain has the following sequence:

[0376]

[0377] In some embodiments, such hIgG1-LALA-YTE HC constant domain has the following sequence:

[0378]

[0379]

[0380] In some embodiments, the hIgG1-LAGA YTE HC constant domain has the following sequence:

[0381]

[0382] In some embodiments, such hIgG1-LALA-LS HC constant domain has the following sequence:

[0383]

[0384] In some embodiments, such an IgG4-YTE HC constant domain has the following sequence:

[0385]

[0386] In some embodiments, such an IgG4-LS HC constant domain has the following sequence:

[0387]

[0388] In some embodiments, such a human κ LC constant domain has the following sequence:

[0389]

[0390] CDR

[0391] In some embodiments, the antibodies provided herein comprise one to three CDRs of a VH domain selected from SEQ ID NOs: 1-32 and 470, such as any of the CDRs listed in Table 3, Table 4, or Table 5 below. In some embodiments, the antibodies provided herein comprise two to three CDRs of a VH domain selected from SEQ ID NOs: 1-32 and 470. In some embodiments, the antibodies provided herein comprise three CDRs of a VH domain selected from SEQ ID NOs: 1-32 and 470. In some embodiments, the CDRs are exemplary CDRs. In some embodiments, the CDRs are Kabat CDRs. In some embodiments, the CDRs are Chothia CDRs. In some embodiments, the CDRs are IMGT CDRs. In some embodiments, the CDRs are AbM CDRs. In some embodiments, the CDRs are Contact CDRs.

[0392] In some embodiments, the CDR is a CDR that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to CDR-H1, CDR-H2 or CDR-H3 of SEQ ID NOs: 58-140. In some embodiments, CDR-H1 is a CDR-H1 of a VH domain selected from SEQ ID NOs: 1-32 and 470, which has at most 1, 2, 3, 4 or 5 amino acid substitutions. In some embodiments, CDR-H2 is a CDR-H2 of a VH domain of SEQ ID NOs: 1-32 and 470, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, CDR-H3 is a CDR-H3 of a VH domain selected from SEQ ID NOs: 1-32 and 470, having at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be re-isolated, for example, according to the methods for obtaining antibodies provided herein.

[0393] In some embodiments, the antibodies provided herein comprise one to three CDRs of the VL domain of SEQ ID NOs: 33-57 and 471, such as any of the CDRs listed in Table 6, Table 7, or Table 8 below. In some embodiments, the antibodies provided herein comprise two to three CDRs of the VL domain of SEQ ID NOs: 33-57 and 471. In some embodiments, the antibodies provided herein comprise three CDRs of the VL domain of SEQ ID NOs: 33-57 and 471. In some embodiments, the CDRs are exemplary CDRs. In some embodiments, the CDRs are Kabat CDRs. In some embodiments, the CDRs are Chothia CDRs. In some embodiments, the CDRs are IMGT CDRs. In some embodiments, the CDRs are AbM CDRs. In some embodiments, the CDRs are Contact CDRs.

[0394] In some embodiments, the CDR is a CDR having at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to CDR-L1, CDR-L2 or CDR-L3 of SEQ ID NOs: 141-188. In some embodiments, CDR-L1 is CDR-L1 of the VL domain of SEQ ID NOs: 33-57 and 471, which has at most 1, 2, 3, 4 or 5 amino acid substitutions. In some embodiments, CDR-L2 is CDR-L2 of the VL domain of SEQ ID NOs: 33-57 and 471, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, CDR-L3 is SEQ ID NO:33-57 and 471 VL domain CDR-L3, it has at most 1,2,3,4,5,6,7 or 8 amino acid substitutions.In some embodiments, the amino acid substitution is a conservative amino acid substitution.In some embodiments, the antibody described in this paragraph is referred to as "variant" in this article.In some embodiments, such variants are derived from the sequence provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis or any other method known in the art or described herein.In some embodiments, such variants are not derived from the sequence provided herein, and can be re-isolated, for example, according to the method for obtaining antibodies provided herein.

[0395] In some embodiments, the antibodies provided herein comprise one to three CDRs of the VH domain selected from SEQ ID NOs: 1-32 and 470 and one to three CDRs of the VL domain of SEQ ID NOs: 33-57 and 471. In some embodiments, the antibodies provided herein comprise two to three CDRs of the VH domain selected from SEQ ID NOs: 1-32 and 470 and two to three CDRs of the VL domain of SEQ ID NOs: 33-57 and 471. In some embodiments, the antibodies provided herein comprise three CDRs of the VH domain selected from SEQ ID NOs: 1-32 and 470 and three CDRs of the VL domain of SEQ ID NOs: 33-57 and 471. In some embodiments, the CDRs are exemplary CDRs. In some embodiments, the CDRs are Kabat CDRs. In some embodiments, the CDRs are Chothia CDRs. In some embodiments, the CDRs are IMGT CDRs. In some embodiments, the CDRs are AbM CDRs. In some embodiments, the CDRs are Contact CDRs.

[0396] In some embodiments, the antibodies provided herein comprise a CDR-H3 selected from SEQ ID NOs: 112-120 and 130-40. In some embodiments, the CDR-H3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to the CDR-H3 of SEQ ID NOs: 112-120 or 130-40. In some embodiments, the CDR-H3 is a CDR-H3 selected from SEQ ID NOs: 112-120 and 130-40 having at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0397] In some embodiments, the antibodies provided herein comprise the CDR-H1 of SEQ ID NOs: 58-99 and 121. In some embodiments, the CDR-H1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to the CDR-H1 of SEQ ID NOs: 58-99 or 121. In some embodiments, the CDR-H1 is the CDR-H1 of SEQ ID NOs: 58-99 or 121 having at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0398] In some embodiments, the antibodies provided herein comprise a CDR-H2 of any one of SEQ ID NOs: 100-111. In some embodiments, the CDR-H2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to the CDR-H2 of any one of SEQ ID NOs: 100-111. In some embodiments, the CDR-H2 is a CDR-H2 of any one of SEQ ID NOs: 100-111 having at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0399] In some embodiments, the antibodies provided herein comprise a CDR-L3 selected from SEQ ID NOs: 165-172. In some embodiments, the CDR-L3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to the CDR-L3 of SEQ ID NOs: 165-172. In some embodiments, the CDR-L3 is the CDR-L3 of SEQ ID NOs: 165-172, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be re-isolated, for example, according to the methods provided herein for obtaining antibodies.

[0400] In some embodiments, the antibodies provided herein include a CDR-L2 selected from SEQ ID NO: 153-158 and an amino acid sequence LAS. In some embodiments, the CDR-L2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to a CDR-L2 selected from SEQ ID NO: 153-158 and an amino acid sequence LAS. In some embodiments, the CDR-L2 is a CDR-L2 selected from SEQ ID NO: 153-158 and an amino acid sequence LAS, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0401] In some embodiments, the antibodies provided herein comprise a CDR-L1 selected from SEQ ID NOs: 141-144 and 149-152. In some embodiments, the CDR-L1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to a CDR-L1 selected from SEQ ID NOs: 141-144 and 149-152. In some embodiments, the CDR-L1 is a CDR-L1 selected from SEQ ID NOs: 141-144 and 149-152 having at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions. In some embodiments, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein, and can be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0402] In some embodiments, the antibodies provided herein comprise: a CDR-H3 selected from SEQ ID NOs: 112-120 and 130-140; a CDR-H2 selected from SEQ ID NOs: 100-111; a CDR-H1 selected from SEQ ID NOs: 58-99 and 121; a CDR-L3 selected from SEQ ID NOs: 165-172; a CDR-L2 selected from SEQ ID NOs: 153-158 and the amino acid sequence LAS; and a CDR-L1 selected from SEQ ID NOs: 141-144 and 149-152. In some embodiments, the CDR-H3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to a CDR-H3 selected from SEQ ID NOs: 112-120 and 130-140, the CDR-H2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to a CDR-H2 of SEQ ID NOs: 100-111, the CDR-H1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to a CDR-H1 selected from SEQ ID NOs: 58-99 and 121, and the CDR-L3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to a CDR-L3 selected from SEQ ID NOs: NO:165-172 has a CDR-L3 that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical, the CDR-L2 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with a CDR-L2 selected from SEQ ID NO:153-158 and amino acid sequence LAS, and the CDR-L1 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with a CDR-L1 selected from SEQ ID NO:141-144 and 149-152.In some embodiments, the CDR-H3 is a CDR-H3 selected from SEQ ID NOs: 112-120 and 130-140, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H2 is a CDR-H2 of SEQ ID NOs: 100-111, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H1 is a CDR-H1 selected from SEQ ID NOs: 58-99 and 121, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L3 is a CDR-L3 selected from SEQ ID NOs: 165-172, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L2 is a CDR-H2 selected from SEQ ID NOs: 100-111, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; NO: 153-158 and a CDR-L2 of the amino acid sequence LAS, which has up to 1, 2, 3 or 4 amino acid substitutions; and the CDR-L1 is a CDR-L1 selected from SEQ ID NO: 141-144 and 149-152, which has up to 1, 2, 3, 4, 5 or 6 amino acid substitutions.

[0403] In some embodiments, the antibodies provided herein comprise: CDR-H3 of SEQ ID NOs: 112, 121 and 130; CDR-H2 of SEQ ID NOs: 100, 104 and 108; CDR-H1 of SEQ ID NOs: 58, 68 and 85; CDR-L3 of SEQ ID NOs: 168, 173 and 181; CDR-L2 of SEQ ID NO: 153 and amino acid sequence LAS; and CDR-L1 of SEQ ID NOs: 141 and 149. In some embodiments, the CDR-H3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H3 of SEQ ID NO: 112 or 130, the CDR-H2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H2 of SEQ ID NO: 100, 104, or 108, the CDR-H1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H1 of SEQ ID NO: 58, 68, or 85, and the CDR-L3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-L3 of SEQ ID NO: The CDR-L3 of SEQ ID NO: 168 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical, the CDR-L2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to the CDR-L2 of SEQ ID NO: 153 or the amino acid sequence LAS, and the CDR-L1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to the CDR-L1 of SEQ ID NO: 141 or 149.In some embodiments, the CDR-H3 is the CDR-H3 of SEQ ID NO: 112 or 130, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H2 is the CDR-H2 of SEQ ID NO: 100, 104 or 108, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H1 is the CDR-H1 of SEQ ID NO: 58, 68 or 85, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L3 is the CDR-L3 of SEQ ID NO: 168, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L2 is the CDR-L2 of SEQ ID NO: 153 or the amino acid sequence LAS, which has at most 1, 2, 3 or 4 amino acid substitutions; and the CDR-L1 is the CDR-H1 of SEQ ID NO: 58, 68 or 85, which has at most 1, 2, 3, 4 or 5 amino acid substitutions. The CDR-L1 of NO: 141 or 149 having up to 1, 2, 3, 4, 5 or 6 amino acid substitutions.

[0404] In some embodiments, the antibodies provided herein comprise: a CDR-H3 of SEQ ID NO: 112, 121 or 130; a CDR-H2 of SEQ ID NO: 100, 104 or 108; a CDR-H1 of SEQ ID NO: 58, 68 or 85; a CDR-L3 of SEQ ID NO: 165; a CDR-L2 of SEQ ID NO: 153 or amino acid sequence LAS; and a CDR-L1 of SEQ ID NO: 141 or 149. In some embodiments, the CDR-H3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H3 of SEQ ID NO: 112 or 130, the CDR-H2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H2 of SEQ ID NO: 100, 104, or 108, the CDR-H1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H1 of SEQ ID NO: 58, 68, or 85, and the CDR-L3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-L3 of SEQ ID NO: NO:165 has a CDR-L3 that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical, the CDR-L2 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L2 of SEQ ID NO:153 or the amino acid sequence LAS, and the CDR-L1 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L1 of SEQ ID NO:141 or 149.In some embodiments, the CDR-H3 is the CDR-H3 of SEQ ID NO: 112 or 130, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H2 is the CDR-H2 of SEQ ID NO: 100, 104 or 108, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H1 is the CDR-H1 of SEQ ID NO: 58, 68 or 85, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L3 is the CDR-L3 of SEQ ID NO: 165, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L2 is the CDR-L2 of SEQ ID NO: 153 or the amino acid sequence LAS, which has at most 1, 2, 3 or 4 amino acid substitutions; and the CDR-L1 is the CDR-H1 of SEQ ID NO: 58, 68 or 85, which has at most 1, 2, 3, 4 or 5 amino acid substitutions. The CDR-L1 of NO: 141 or 149 having up to 1, 2, 3, 4, 5 or 6 amino acid substitutions.

[0405] In some embodiments, the antibodies provided herein comprise: CDR-H3 of SEQ ID NO: 112 or 130; CDR-H2 of SEQ ID NO: 100, 104 or 108; CDR-H1 of SEQ ID NO: 58, 68 or 85; CDR-L3 of SEQ ID NO: 165; CDR-L2 of SEQ ID NO: 158 or amino acid sequence LAS; and CDR-L1 of SEQ ID NO: 141 or 149. In some embodiments, the CDR-H3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H3 of SEQ ID NO: 112 or 130; the CDR-H2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H2 of SEQ ID NO: 100, 104, or 108; the CDR-H1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H1 of SEQ ID NO: 58, 68, or 85; the CDR-L3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-L3 of SEQ ID NO: NO:165 has a CDR-L3 that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical; the CDR-L2 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L2 of SEQ ID NO:158 or amino acid sequence LAS; and the CDR-L1 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L1 of SEQ ID NO:141 or 149.In some embodiments, the CDR-H3 is the CDR-H3 of SEQ ID NO: 112 or 130, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H2 is the CDR-H2 of SEQ ID NO: 100, 104 or 108, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H1 is the CDR-H1 of SEQ ID NO: 58, 68 or 85, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L3 is the CDR-L3 of SEQ ID NO: 165, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L2 is the CDR-L2 of SEQ ID NO: 158 or the amino acid sequence LAS, which has at most 1, 2, 3 or 4 amino acid substitutions; and the CDR-L1 is the CDR-L1 of SEQ ID NO: The CDR-L1 of NO: 141 or 149 having up to 1, 2, 3, 4, 5 or 6 amino acid substitutions.

[0406] In some embodiments, the antibodies provided herein comprise: a CDR-H3 of SEQ ID NO: 112, 121 or 130; a CDR-H2 of SEQ ID NO: 100, 104 or 108; a CDR-H1 of SEQ ID NO: 58, 67 or 84; a CDR-L3 of SEQ ID NO: 165; a CDR-L2 of SEQ ID NO: 153 or amino acid sequence LAS; and a CDR-L1 of SEQ ID NO: 141 or 149. In some embodiments, the CDR-H3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H3 of SEQ ID NO: 112 or 130, the CDR-H2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H2 of SEQ ID NO: 100, 104, or 108, the CDR-H1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H1 of SEQ ID NO: 58, 67, or 84, and the CDR-L3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-L3 of SEQ ID NO: NO:165 has a CDR-L3 that is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical, the CDR-L2 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L2 of SEQ ID NO:153 or the amino acid sequence LAS, and the CDR-L1 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L1 of SEQ ID NO:141 or 149.In some embodiments, the CDR-H3 is the CDR-H3 of SEQ ID NO: 112 or 130, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H2 is the CDR-H2 of SEQ ID NO: 100, 104 or 108, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H1 is the CDR-H1 of SEQ ID NO: 58, 67 or 84, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L3 is the CDR-L3 of SEQ ID NO: 165, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L2 is the CDR-L2 of SEQ ID NO: 153 or the amino acid sequence LAS, which has at most 1, 2, 3 or 4 amino acid substitutions; and the CDR-L1 is the CDR-H1 of SEQ ID NO: 58, 67 or 84, which has at most 1, 2, 3, 4 or 5 amino acid substitutions. The CDR-L1 of NO: 141 or 149 having up to 1, 2, 3, 4, 5 or 6 amino acid substitutions.

[0407] In some embodiments, the antibodies provided herein comprise: CDR-H3 of SEQ ID NO: 112 or 130; CDR-H2 of SEQ ID NO: 100, 104 or 108; CDR-H1 of SEQ ID NO: 58, 67 or 84; CDR-L3 of SEQ ID NO: 165; CDR-L2 of SEQ ID NO: 158 or amino acid sequence LAS; and CDR-L1 of SEQ ID NO: 141 or 149. In some embodiments, the CDR-H3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H3 of SEQ ID NO: 112 or 130; the CDR-H2 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H2 of SEQ ID NO: 100, 104, or 108; the CDR-H1 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-H1 of SEQ ID NO: 58, 67, or 84; the CDR-L3 is at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the CDR-L3 of SEQ ID NO: NO:165 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L3; the CDR-L2 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L2 of SEQ ID NO:158 or amino acid sequence LAS; and the CDR-L1 has at least about 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the CDR-L1 of SEQ ID NO:141 or 149.In some embodiments, the CDR-H3 is the CDR-H3 of SEQ ID NO: 112 or 130, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H2 is the CDR-H2 of SEQ ID NO: 100, 104 or 108, which has at most 1, 2, 3, 4, 5, 6, 7 or 8 amino acid substitutions; the CDR-H1 is the CDR-H1 of SEQ ID NO: 58, 67 or 84, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L3 is the CDR-L3 of SEQ ID NO: 165, which has at most 1, 2, 3, 4 or 5 amino acid substitutions; the CDR-L2 is the CDR-L2 of SEQ ID NO: 158 or the amino acid sequence LAS, which has at most 1, 2, 3 or 4 amino acid substitutions; and the CDR-L1 is the CDR-L1 of SEQ ID NO: The CDR-L1 of NO: 141 or 149 having up to 1, 2, 3, 4, 5 or 6 amino acid substitutions.

[0408] In some embodiments, the amino acid substitution is a conservative amino acid substitution. In some embodiments, the antibodies described in the present invention are referred to herein as "variants" or "clones". In some embodiments, such variants or clones are derived from sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants or clones are non-derived from sequences provided herein, and can be isolated again, for example, according to the method for obtaining antibodies provided herein.

[0409] In certain aspects, the antibodies disclosed herein do not include the antibodies disclosed in US Pat. No. 9,067,994.

[0410]

[0411]

[0412]

[0413]

[0414]

[0415]

[0416]

[0417]

[0418]

[0419]

[0420]

[0421]

[0422]

[0423]

[0424]

[0425]

[0426]

[0427]

[0428]

[0429]

[0430]

[0431]

[0432]

[0433]

[0434]

[0435]

[0436]

[0437]

[0438]

[0439]

[0440]

[0441]

[0442]

[0443]

[0444]

[0445]

[0446]

[0447]

[0448]

[0449]

[0450]

[0451] Fc region

[0452] The structures of the Fc regions of different immunoglobulins and the glycosylation sites contained therein are known in the art. See Schroeder and Cavacini, J. Allergy Clin. Immunol., 2010, 125: S41-52, which is incorporated by reference in its entirety. The Fc region may be a naturally occurring Fc region or a modified Fc region as described in the art or elsewhere in this disclosure.

[0453] Unless otherwise specified herein, the numbering of amino acid residues in the Fc region or constant region is according to the EU numbering system, also known as the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th edition. Public Health Service, National Institutes of Health, Bethesda, MD, 1991. As used herein, the "Fc polypeptide" of a dimeric Fc refers to one of the two polypeptides that form the dimeric Fc domain, i.e., a polypeptide comprising a C-terminal constant region of an immunoglobulin heavy chain that is capable of stabilizing self-association. For example, the Fc polypeptide of a dimeric IgG Fc comprises IgGCH2 and IgGCH3 constant domain sequences. Fc can have classes IgA, IgD, IgE, IgG, and IgM, and several of these can be further divided into subclasses (isotypes), such as IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2.

[0454] The terms "Fc receptor" and "FcR" are used to describe receptors that bind to the Fc region of an antibody. For example, an FcR may be a native sequence human FcR. Typically, an FcR is one that binds to an IgG antibody (gamma receptor) and includes receptors of FcγRI, FcγRII, and FcγRIII subclasses, including allelic variants and alternative splicing forms of these receptors. FcγRII receptors include FcγRIIA ("activating receptor") and FcγRIIB ("inhibiting receptor"), both of which have similar amino acid sequences that differ primarily in their cytoplasmic domains. Other isotypes of immunoglobulins may also be bound by certain FcRs (see, e.g., Janeway et al., ImmunoBiology: the immune system in health and disease, (Elsevier Science Ltd., NY) (4th edition, 1999)). The activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain. Inhibiting receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibition motif (ITIM) in its cytoplasmic domain (reviewed in Annu. Rev. Immunol. 15: 203-234 (1997)). FcRs are reviewed in Ravetch and Kinet, Annu. Rev. Immunol 9: 457-92 (1991); Capel et al., Immunomethods 4: 25-34 (1994); and de Haas et al., J. Lab. Clin. Med. 126: 330-41 (1995). The term "FcR" herein encompasses other FcRs including FcRs identified in the future. The term also includes the neonatal receptor FcRn, which is responsible for transporting maternal IgG to the fetus (Guyer et al., J Immunol. 117: 587 (1976); and Kim et al., J Immunol. 24: 249 (1994)).

[0455] Modifications in the CH2 domain may affect the binding of FcR to Fc. Various amino acid modifications in the Fc region are known in the art for selectively changing the affinity of Fc for different Fcγ receptors. In some embodiments, Fc comprises one or more modifications that promote selective binding of Fc-γ receptors.

[0456] Exemplary mutations that alter FcR binding to Fc are listed below:

[0457] S298A / E333A / K334A, S298A / E333A / K334A / K326A (Lu Y, Vernes JM, Chiang N et al., J Immunol Methods. 2011 Feb 28;365(1-2):132-41);

[0458] F243L / R292P / Y300L / V305I / P396L, F243L / R292P / Y300L / L235V / P396L (Stavenhagen JB, Gorlatov S, Tuaillon N, et al., Cancer Res. 2007 Sep 15;67(18):8882-90; Nordstrom JL, Gorlatov S, Zhang W, et al., Breast Cancer Res. 2011 Nov 30;13(6):R123);

[0459] F243L (Stewart R, Thom G, Levens M et al., Protein Eng Des Sel. 2011 Sep; 24(9): 671-8.), S298A / E333A / K334A (Shields RL, Namenuk AK, Hong K et al., J Biol Chem. 2001 Mar 2; 276(9): 6591-604);

[0460] S239D / I332E / A330L, S239D / I332E (Lazar GA, Dang W, Karki S et al., Proc Natl Acapd Sci US A. 2006 Mar 14; 103(11):4005-10);

[0461] S239D / S267E, S267E / L328F (Chu SY, Vostiar I, Karki S et al., Mol Immunol. 2008 Sep;45(15):3926-33); and

[0462] S239D / D265S / S298A / I332E, S239E / S298A / K326A / A327H, G237F / S298A / A330L / I332E, S239D / I332E / S298A, S239D / K326E / A330L / I332E / S298A, G236A / S239D / D270L / I332E, S239E / S267E / H268D, L234F / S267E / N325L, G237F / V266L / S267D and other mutations listed in WO2011 / 120134 and WO2011 / 120135, which are incorporated herein by reference. The rapeutic Antibody Engineering (William R. Strohl and Lila M. Strohl, Woodhead Publishing series in Biomedicine No. 11, ISBN 1907568379, October 2012) lists mutations on page 283.

[0463] In some embodiments, the antibodies described herein include modifications to improve their ability to mediate effector functions. Such modifications are known in the art and include defucosylation or engineering of the affinity of the Fc for activating receptors, primarily FCGR3a for ADCC and C1q for CDC. Table 9 below summarizes the different designs reported in the literature for effector function engineering.

[0464] Methods for generating antibodies with little or no fucose at the Fc glycosylation position (Asn 297 EU numbering) without changing the amino acid sequence are well known in the art. The technology (ProBioGen AG) is based on the introduction of genes for enzymes that bias fucose biosynthesis into cells used for antibody production. This prevents antibody-producing cells from adding the sugar "fucose" to N-linked antibody carbohydrate moieties (von Horsten et al. (2010) Glycobiology. 2010 Dec; 20(12): 1607-18). Another approach to obtaining antibodies with reduced fucosylation levels can be found in U.S. Pat. No. 8,409,572, which teaches selecting a cell line capable of producing lower fucosylation levels on antibodies for antibody production. The antibody may be completely defucosylated (meaning it contains no detectable fucose) or it may be partially defucosylated, meaning the fucose of the isolated antibody contains less than 95%, less than 85%, less than 75%, less than 65%, less than 55%, less than 45%, less than 35%, less than 25%, less than 15% or less than 5% of the amount of fucose normally detected for similar antibodies produced by mammalian expression systems.

[0465] Thus, in one embodiment, an antibody described herein may include a dimeric Fc comprising one or more amino acid modifications conferring improved effector function as indicated in Table 9. In another embodiment, the antibody may be defucosylated to improve effector function.

[0466] Table 9: CH2 domain and effector function engineering

[0467]

[0468] Fc modifications that reduce FcgR and / or complement binding and / or effector function are known in the art. Recent publications describe strategies that have been used to engineer antibodies with lower or silent effector activity (see Strohl, WR (2009), Curr Opin Biotech 20: 685-691 and Strohl, WR and Strohl LM, "Antibody Fc engineering for optimal antibody performance", Therapeutic Antibody Engineering, Cambridge: Woodhead Publishing (2012), pp. 225 to 249). These strategies include reducing effector function by changing glycosylation, using IgG2 / IgG4 scaffolds or introducing mutations in the hinge or CH2 domains of Fc. For example, U.S. Patent Publication No. 2011 / 0212087 (Strohl), International Patent Publication No. WO2006 / 105338 (Xencor), U.S. Patent Publication No. 2012 / 0225058 (Xencor), U.S. Patent Publication No. 2012 / 0251531 (Genentech), and Strop et al. ((2012) J Mol. Biol. 420:204-219) describe specific modifications for reducing binding of FcgR or complement to Fc.

[0469] Specific non-limiting examples of amino acid modifications known to reduce binding of FcgR or complement to Fc include those identified in Table 10 below:

[0470] Table 10: Modifications for reducing binding of FcgR or complement to Fc

[0471] company mutation GSK N297A Ortho Biotech L234A / L235A Protein Design labs IGG2 V234A / G237A Wellcome Labs IGG4 L235A / G237A / E318A GSK IGG4 S228P / L236E Alexion IGG2 / IGG4combo Merck IGG2 H268Q / V309L / A330S / A331S Bristol-Myers C220S / C226S / C229S / P238S Seattle Genetics C226S / C229S / E3233P / L235V / L235A Amgen Produced by E. coli, non-sugar Medimune L234F / L235E / P331S Trubion Hinge mutant, probably C226S / P230S

[0472] Examples of cell lines capable of producing defucosylated antibodies include CHO-DG44 with stable overexpression of the bacterial oxidoreductase GDP-6-deoxy-D-lyxose-4-hexose reductase (RMD) (see Henning von Horsten et al., Glycobiol 2010, 20: 1607-1618) or Lec13 CHO cells, which lack protein fucosylation (see Ripka et al., Arch. Biochem. Biophys., 1986, 249: 533-545; U.S. Patent Publication No. 2003 / 0157108; WO 2004 / 056312; each of which is incorporated by reference in its entirety) and gene knockout cell lines, such as α-1,6-fucosyltransferase gene or FUT8 gene knockout CHO cells (see Yamane-Ohnuki et al., Biotech. Bioeng., 2004, 87: 614-622; Kanda et al., Biotechnol. Bioeng., 2006, 94: 680-688; and WO 2003 / 085107; each of which is incorporated by reference in its entirety). Another approach to obtaining antibodies with reduced fucosylation levels can be found in U.S. Pat. No. 8,409,572, which teaches selecting cell lines capable of producing lower fucosylation levels on antibodies for antibody production.

[0473] The antibody may be fully defucosylated, meaning that it contains no detectable fucose, or it may be partially defucosylated, meaning that the isolated antibody contains less than 95%, less than 85%, less than 75%, less than 65%, less than 55%, less than 45%, less than 35%, less than 25%, less than 15%, or less than 5% of the amount of fucose normally detected for a similar antibody produced by a mammalian expression system.

[0474] In some embodiments, the antibodies provided herein comprise an IgG1 domain having reduced fucose content at position Asn297 compared to a naturally occurring IgG1 domain. Such Fc domains are known to have improved ADCC. See Shields et al., J Biol. Chem., 2002, 277: 26733-26740, which is incorporated by reference in its entirety. In some embodiments, such antibodies will not comprise any fucose at position Asn 297. The amount of fucose can be determined using any suitable method, for example as described in WO 2008 / 077546, which is incorporated by reference in its entirety.

[0475] In certain embodiments, the antibodies provided herein comprise an Fc region having one or more amino acid substitutions that improve ADCC, such as substitutions at one or more of positions 298, 333, and 334 of the Fc region. In some embodiments, the antibodies provided herein comprise an Fc region having one or more amino acid substitutions at positions 239, 332, and 330, as described in Lazar et al., Proc. Natl. Acad Sci. USA, 2006, 103: 4005-4010, which is incorporated by reference in its entirety.

[0476] Other illustrative glycosylation variants that can be incorporated into the antibodies provided herein are described in, e.g., U.S. Patent Publication Nos. 2003 / 0157108, 2004 / 0093621, 2003 / 0157108, 2003 / 0115614, 2002 / 0164328, 2004 / 0093621, 2004 / 0132140, 2004 / 0110704, 2004 / 0164328, 2004 / 0093621, 2004 / 0132140, 2004 / 0110704, 2004 / 0164328, 2004 / 0093621, 2004 / 0132141, 2004 / 0132143 2004 / 0110282, 2004 / 0109865; International Patent Publication Nos. 2000 / 61739, 2001 / 29246, 2003 / 085119, 2003 / 084570, 2005 / 035586, 2005 / 035778, 2005 / 053742, 2002 / 031140; Okazaki et al., J Mol. Biol., 2004, 336: 1239-1249; and Yamane-Ohnuki et al., Biotech. Bioeng., 2004, 87: 614-622; each of which is incorporated by reference in its entirety.

[0477] In some embodiments, the antibodies provided herein include at least one galactose residue in the oligosaccharide connected to the Fc region of the Fc region. Such antibody variants may have improved CDC function. Examples of such antibody variants are described in, for example, WO1997 / 30087, WO1998 / 58964, and WO1999 / 22764; each of the documents is incorporated by reference in its entirety.

[0478] In some embodiments, the antibodies provided herein comprise one or more alterations that improve or reduce C1q binding and / or CDC. See U.S. Pat. No. 6,194,551; WO 99 / 51642; and Idusogie et al., J. Immunol., 2000, 164: 4178-4184; each of which is incorporated by reference in its entirety.

[0479] In certain embodiments, the antibodies provided herein comprise a heavy chain comprising a constant heavy chain sequence selected from the group consisting of those set forth in SEQ ID NOs: 425-468 and 484-539.

[0480] In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 1 and a VL sequence as set forth in SEQ ID NO: 33; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 2 and a VL sequence as set forth in SEQ ID NO: 33; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 3 and a VL sequence as set forth in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 4 and a VL sequence as set forth in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 5 and a VL sequence as set forth in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 6 and a VL sequence as set forth in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 3 and a VL sequence as set forth in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 4 and a VL sequence as set forth in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 5 and a VL sequence as set forth in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 6 and a VL sequence as set forth in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 439, 440, 446, 457, and 460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 3 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 4 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 5 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 6 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 3 and a VL sequence as set forth in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 4 and a VL sequence as set forth in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 5 and a VL sequence as set forth in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 6 and a VL sequence as set forth in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 8 and a VL sequence as set forth in SEQ ID NO: 42; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 9 and a VL sequence as set forth in SEQ ID NO: 43; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 44; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 45; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 46; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 47; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 48; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 49; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 50; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 52; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 53; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 54; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 55; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 56; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 57; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 10 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 11 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 12 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 13 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 14 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 15 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NO: 436-468 and 484-539.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 16 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 17 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 18 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 19 and a VL sequence as set forth in SEQ ID NOO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 20 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 21 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 22 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 23 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 24 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 25 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 26 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 27 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 28 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 28 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 29 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 30 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 31 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 32 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequences set forth in SEQ ID NOs: 436-468 and 484-539.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 8 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 8 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 3 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences set forth in SEQ ID NOs: 436-468 and 484-539. In certain embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO: 470 and a VL sequence as shown in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

[0481] In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 15 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 15 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 15 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 15 and a VL sequence as set forth in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 446. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO:439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO:457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO:460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO:446. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 7 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 457.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO:460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO:446. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:8 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO:439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:8 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO:457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 8 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 8 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 446. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence as set forth in SEQ ID NO: 446.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:446. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:446. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 3 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO: 439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 3 and a VL sequence as set forth in SEQ ID NO: 51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO: 446.In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:3 and a VL sequence as set forth in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:446. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:7 and a VL sequence as set forth in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:460. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:470 and a VL sequence as set forth in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:439. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO:470 and a VL sequence as set forth in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO:446. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO: 457. In certain embodiments, the antibody comprises a VH sequence as set forth in SEQ ID NO: 470 and a VL sequence as set forth in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of the sequence set forth in SEQ ID NO: 460.

[0482] In certain embodiments, the isolated antibodies described herein comprise the constant light chain sequence set forth in SEQ ID NO:469.

[0483] In certain embodiments, the Fc region comprises one or more amino acid substitutions, wherein the one or more substitutions cause an increase in antibody half-life, an increase in ADCC activity, an increase in ADCP activity, or an increase in CDC activity compared to an Fc without the one or more substitutions. In certain embodiments, the one or more amino acid substitutions cause an increase in antibody half-life at pH 6.0 compared to an antibody comprising a wild-type Fc region. In certain embodiments, the half-life of an isolated antibody comprising an Fc region having one or more amino acid substitutions in humans is about 80 to 110 days.

[0484] In certain embodiments, the increased half-life of the antibody is about 10,000-fold, 1,000-fold, 500-fold, 100-fold, 50-fold, 20-fold, 10-fold, 9-fold, 8-fold, 7-fold, 6-fold, 5-fold, 4.5-fold, 4-fold, 3.5-fold, 3-fold, 2.5-fold, 2-fold, 1.95-fold, 1.9-fold, 1.85-fold, 1.8-fold, 1.75-fold, 1.7-fold, 1.65-fold, 1.6-fold, 1.55-fold, 1.50-fold, 1.45-fold, 1.4-fold, 1.35-fold, 1-3-fold, 1.25-fold, 1.2-fold, 1.15-fold, 1.1-fold, or 1.05-fold longer than that of an antibody comprising a wild-type Fc region. In certain embodiments, the increased half-life of the antibody is about 10,000-fold, 1,000-fold, 500-fold, 100-fold, 50-fold, 20-fold, 10-fold, 9-fold, 8-fold, 7-fold, 6-fold, 5-fold, 4.5-fold, 4-fold, 3.5-fold, 3-fold, 2.5-fold, 2-fold, 1.95-fold, 1.9-fold, 1.85-fold, 1.8-fold, 1.75-fold, 1.7-fold, 1.65-fold, 1.6-fold, 1.55-fold, 1.50-fold, 1.45-fold, 1.4-fold, 1.35-fold, 1.3-fold, 1.25-fold, 1.2-fold, 1.15-fold, 1.1-fold, or 1.05-fold longer than that of leruzumab.

[0485] In certain embodiments, the Fc region comprises one or more amino acid substitutions, wherein the one or more substitutions cause an increase in antibody half-life compared to an Fc without the one or more substitutions; one or more of ADCC activity, ADCP activity, or CDC activity is reduced. In certain embodiments, the one or more amino acid substitutions cause an increase in antibody half-life at pH 6.0 compared to an antibody comprising a wild-type Fc region. In certain embodiments, the half-life of an isolated antibody comprising an Fc region having one or more amino acid substitutions in humans is about 80 to 110 days.

[0486] In certain embodiments, the one or more amino acid substitutions are selected from the group consisting of S228P (SP); M252Y, S254T, T256E, T256D, T250Q, H285D, T307A, T307Q, T307R, T307W, L309D, Q411H, Q311V, A378V, E380A, M428L, N434A, N434S, N297A, D265A, L234A, L235A, and N434W. In certain embodiments, the one or more amino acid substitutions comprise a plurality of amino acid substitutions selected from the group consisting of: M428L / N434S (LS); M252Y / S254T / T256E (YTE); T250Q / M428L; T307A / E380A / N434A; T256D / T307Q (DQ); T256D / T307W (DW); M252Y / T256D (YD); T307Q / Q311V / A378V (QVV); T256D / H285D / T307R / Q311V / A378V (D DRVV); L309D / Q311H / N434S(DHS); S228P / L235E(SPLE); L234A / L235A(LA), M428L / N434A L234A / G237A(LALA), L234A / L235A / G237A, L234A / L235A / P329G, N297A, D265A / YTE, LALA / YT E. LAGA / YTE, LALAGA / YTE, LALAPG / YTE, N297A / LS; D265A / LS; LALA / LS; LALAGA / LS; LALAPG / LS; N297A / DHS; D265A / DHS; LALA / DHS; LAGA / DHS ;LALAGA / DHS;LALAPG / DHS;SP / YTE;SPLE / YTE;SP / LS;SPLE / LS, SP / DHS;SPLE / DHS;N297A / LA;D265A / LA, LALA / LA, LAGA / LA, LALAGA / LA, LALA PG / LA, N297A / N434A; D265A / N434A;LALA / N434A, LAGA / N434A, LALAGA / N434A, LALAPG / N434A, N297A / N434W, D265A / N434W, LALA / N434W, LAGA / N434W, LALAGA / N434W, LALA PG / N434W, N297A / DQ, D265A / DQ, LALA / DQ, LAGA / DQ, LAL AGA / DQ, LALAPG / DQ, N297A / DW, D265A / DW, LALA / DW, LAGA / DW, LALAGA / DW, LALAPG / DW N297A / YD, D265A / YD, LALA / YD, LAGA / YD, LALAGA / YD, LALAPG / YD, T307Q / Q311V / A378V(QVV), N297A / QVV, D265A / QVV, LAL A / QVV, LAGA / QVV, LALAGA / QVV, LALAPG / QVV, DDRVV, N297A / DDRVV, D265A / DDRVV, LALA / DDRVV, LAGA / DDRVV, LALAGA / DDRV V and LALAPG / DDRVV. ;

[0487] In certain embodiments, the one or more amino acid substitutions are selected from the group consisting of LALA / YTE, LAGA / YTE, LALA / LS, YTE, and LS.

[0488] In certain embodiments, the one or more amino acid substitutions comprise or consist of LALA / YTE. In certain embodiments, the one or more amino acid substitutions comprise or consist of LAGA / YTE. In certain embodiments, the one or more amino acid substitutions comprise or consist of LALA / LS. In certain embodiments, the one or more amino acid substitutions comprise or consist of YTE. In certain embodiments, the one or more amino acid substitutions comprise or consist of LS.

[0489] In certain embodiments, the Fc region binds an Fcγ receptor selected from the group consisting of: FcγRI, FcγRIIa, FcγRIIb, FcγRIIc, FcγRIIIa, and FcγRIIIb. In certain embodiments, the Fc region binds an Fcγ receptor with a higher affinity at pH 6.0 than an antibody comprising a wild-type Fc region.

[0490] Combination

[0491] The affinity of a molecule X for its partner Y can be expressed by the dissociation constant (K D) is represented. The kinetic components that contribute to the dissociation equilibrium constant are described in more detail below. The kinetic components that contribute to the dissociation equilibrium constant can be determined by common methods known in the art, including those described herein, such as surface plasmon resonance (SPR) techniques (e.g. ) or biolayer interferometry (e.g. ) to measure affinity.

[0492] With respect to the binding of an antibody to a target molecule, the terms "bind", "specifically bind", "binds specifically to", "has specificity for", "selectively binds", and "is selective for" a particular antigen (e.g., a polypeptide target) or an epitope on a particular antigen mean binding that is measurably different from a non-specific or non-selective interaction (e.g., with a non-target molecule). Specific binding can be measured, for example, by measuring binding to a target molecule (i.e., IL-13) and comparing it to binding to a non-target molecule. Specific binding can also be determined by competition with a control molecule that mimics the epitope recognized on the target molecule. In such cases, if the binding of the antibody to the target molecule is competitively inhibited by the control molecule, specific binding is indicated. In some embodiments, the affinity of the anti-IL-13 antibody for the non-target molecule is less than about 50% of the affinity for IL-13. In some embodiments, the affinity of the anti-IL-13 antibody for the non-target molecule is less than about 40% of the affinity for IL-13. In some embodiments, the affinity of the anti-IL-13 antibody for the non-target molecule is less than about 30% of the affinity for IL-13. In some embodiments, the anti-IL-13 antibody has an affinity for a non-target molecule that is less than about 20% of its affinity for IL-13. In some embodiments, the anti-IL-13 antibody has an affinity for a non-target molecule that is less than about 10% of its affinity for IL-13. In some embodiments, the anti-IL-13 antibody has an affinity for a non-target molecule that is less than about 10% of its affinity for IL-13. In some embodiments, the anti-IL-13 antibody has an affinity for a non-target molecule that is less than about 1% of its affinity for IL-13. In some embodiments, the anti-IL-13 antibody has an affinity for a non-target molecule that is less than about 0.1% of its affinity for IL-13.

[0493] When used in the context of two or more antibodies herein, the term "competes with" or "cross-competes with" indicates that the two or more antibodies compete for binding to an antigen (e.g., IL-13). In one exemplary assay, IL-13 is coated on a surface and contacted with a first anti-IL-13 antibody, after which a second anti-IL-13 antibody is added. In another exemplary assay, a first anti-IL-13 antibody is coated on a surface and contacted with IL-13, and then a second anti-IL-13 antibody is added. If the presence of the first anti-IL-13 antibody reduces the binding of the second anti-IL-13 antibody in either assay, the antibodies compete with each other. The term "competes with" also includes antibody combinations in which one antibody reduces the binding of another antibody, but in which no competition is observed when the antibodies are added in the opposite order. However, in some embodiments, the first antibody and the second antibody inhibit binding to each other, regardless of the order in which they are added. In some embodiments, one antibody reduces the binding of another antibody to its antigen by at least 25%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99%, as measured in a competitive binding assay. One skilled in the art can select the concentration of the antibody to be used in a competitive assay based on the affinity of the antibody for IL-13 and the antibody valency. The assays described in this definition are illustrative, and one skilled in the art can utilize any suitable assay to determine whether antibodies compete with each other. Suitable assays are described, for example, in Cox et al., "Immunoassay Methods," Assay Guidance Manual [Internet], updated December 24, 2014 (ncbi.nlm.nih.gov / books / NBK92434 / ; accessed September 29, 2015); Silman et al., Cytometry, 2001, 44:30-37; and Finco et al., J. Pharm. Biomed. Anal., 2011, 54:351-358; each of which is incorporated by reference in its entirety.

[0494] If an excess of the test antibody (e.g., at least 2x, 5x, 10x, 20x, or 100x) inhibits or blocks the binding of the reference antibody, for example, by at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%, as measured in a competitive binding assay, then the test antibody competes with the reference antibody. Antibodies identified by competition assays (competing antibodies) include antibodies that bind to the same epitope as the reference antibody and antibodies that bind to an adjacent epitope that is sufficiently close to the epitope bound by the reference antibody to cause steric hindrance. For example, a second competing antibody may be identified as competing with the first antibody described herein for binding to IL-13. In some cases, the second antibody may block or inhibit the binding of the first antibody, for example, by at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%, as measured in a competitive binding assay. In certain cases, the second antibody may replace greater than 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% of the first antibody.

[0495] In certain embodiments, the antibody binds human IL-13.

[0496] In certain embodiments, the antibody binds to the IL-13 sequence set forth in SEQ ID NOs: 472-475.

[0497] In certain embodiments, the antibody is cross-reactive with cynomolgus monkey IL-13.

[0498] In certain embodiments, the antibody has a specific activity as measured by SPR at less than or equal to about 1, 2, 3, 4, 5, 6, 7, 8, 9 x 10 -9 M of K D Binds to the IL-13 sequence set forth in SEQ ID NOs: 472-475. In certain embodiments, the antibody has a binding affinity of less than or equal to about 1 x 10 -10 M of K D Binds to the IL-13 sequence set forth in SEQ ID NOs: 472-475. In certain embodiments, the antibody has a binding affinity of less than or equal to about 1 x 10 -9 M of K D Binds to human IL-13.

[0499] In some embodiments, the antibodies provided herein are expressed as follows, as measured by ELISA or any other suitable method known in the art: DBinding to IL-13: less than or equal to about 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 1.95, 2, 2.5, 3, 3.5, 4, 4.5, 5, 6, 7, 8, 9, or 10 x 10 -8 M. In some embodiments, the antibodies provided herein are expressed as follows, as measured by ELISA or any other suitable method known in the art: D Binding to IL-13: less than or equal to about 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 1.95, 2, 2.5, 3, 3.5, 4, 4.5, 5, 6, 7, 8, 9, or 10 x 10 -9 M.

[0500] In some embodiments, the antibodies provided herein bind to IL-13 with a K of 1.5, 2.0, or 1.5, as measured by ELISA or any other suitable method known in the art. D Between: about 0.001-0.01, 0.01-0.1, 0.01-0.05, 0.05-0.1, 0.1-0.5, 0.5-1, 0.25-0.75, 0.25-0.5, 0.5-0.75, 0.75-1, 0.75-2, 1.1-1.2, 1.2-1.3, 1.3-1.4, 1.4-1.5, 1.5-1.6, 1.6-1.7, 1.7-1.8, 1.8-1.9, 1.9-2, 1-2, 1-5, 2-7, 3-8, 3-5, 4-6, 5-7, 6-8, 7-9, 7-10, or 5-10 x 10 -8 In some embodiments, the antibodies provided herein have a level of less than or equal to about 1 x 10 -8 M or less than or equal to 1x10 -9 M of K D Binds to IL-13.

[0501] In some embodiments, the antibodies provided herein are expressed as follows, as measured by ELISA or any other suitable method known in the art: DBinding to IL-13: less than or equal to about 10, 9, 8, 7, 6, 5, 4.5, 4, 3.5, 3, 2.5, 2, 1.98, 1.95, 1.9, 1.85, 1.8, 1.75, 1.7, 1.65, 1.6, 1.55, 1.50, 1.45, 1.4, 1.3, 1.2, 1.1, 1, 0.9, 0.85, 0.8, 0.75, 0.7, 0.65, 0.6, 0.55, 0.5, 0.45, 0.4, 0.35, 0.3, 0.25, 0.2, 0.15, 0.1, 0.05, 0.01, 0.005, 0.001, 0.0005, or 0.0001 x 10 -8 In some embodiments, the antibodies provided herein have a K between: D Binding to IL-13: 5-3, 4-2, 3-1, 1.9-1.8, 1.8-1.7, 1.7-1.6, 1.6-1.5, 1.9-1.5, 1.5-1, 1-0.8, 1-0.5, 0.9-0.6, 0.7-0.4, 0.6-0.2, 0.5-0.3, 0.3-0.2, 0.2-0.1, 0.1-0.01, 0.01-0.001, or 0.001-0.0001 x 10 -8 M.

[0502] Function

[0503] "Effector functions" refer to those biological activities mediated by the Fc region of an antibody, which may vary depending on the antibody isotype. Examples of antibody effector functions include receptor ligand blocking, agonism, or antagonism, C1q binding to activate complement dependent cytotoxicity (CDC), Fc receptor binding to activate antibody dependent cellular cytotoxicity (ADCC), and antibody dependent cellular phagocytosis (ADCP). In some embodiments, the effector function of the anti-IL-13 antibodies described herein is antagonism and blocks the binding of the IL-13 receptor to IL-13.

[0504] Pharmaceutical composition

[0505] The application provides a composition comprising an antibody, which includes a pharmaceutical composition comprising any one or more of the antibodies described herein and one or more pharmaceutically acceptable excipients. In some embodiments, the composition is sterile. The pharmaceutical composition generally comprises an effective amount of an antibody.

[0506] In addition to one or more of the antibodies disclosed herein, these compositions may contain pharmaceutically acceptable excipients, carriers, buffers, stabilizers or other materials well known to those skilled in the art. Such materials should be non-toxic and should not interfere with the efficacy of the active ingredient. The exact nature of the carrier or other material may depend on the route of administration, such as oral, intravenous, cutaneous or subcutaneous, nasal, intramuscular, intraperitoneal routes.

[0507] Pharmaceutical compositions for oral administration may be in tablet, capsule, powder or liquid form. Tablets may include solid carriers such as gelatin or adjuvants. Liquid pharmaceutical compositions typically include liquid carriers such as water, petroleum, animal or vegetable oils, mineral oils or synthetic oils. Physiological saline solutions, dextrose or other sugar solutions or glycols such as ethylene glycol, propylene glycol or polyethylene glycol may be included.

[0508] For intravenous, cutaneous or subcutaneous injection or injection at the site of affliction, the active ingredient will be in the form of a parenterally acceptable aqueous solution that is pyrogen-free and has suitable pH, isotonicity and stability. Those skilled in the art are well able to prepare suitable solutions using, for example, isotonic vehicles (e.g., sodium chloride injection, Ringer's Injection, lactated Ringer's Injection). Preservatives, stabilizers, buffers, antioxidants and / or other additives may be included as needed.

[0509] The anti-IL-13 antibody to be administered to an individual is preferably administered in a "therapeutically effective amount" or a "prophylactically effective amount" (as the case may be, but prevention may be considered treatment), which is sufficient to show benefit to the individual. The actual amount administered and the rate and time course of administration will depend on the nature and severity of the protein aggregation disease being treated. Treatment prescriptions, such as decisions on dosage, are within the responsibility of general practitioners and other physicians, and typically take into account the condition to be treated, the individual patient's condition, the delivery site, the method of administration, and other factors known to professionals. Examples of the techniques and protocols mentioned above can be found in Remington's Pharmaceutical Sciences, 16th edition, Osol, A. (ed.), 1980.

[0510] Depending on the condition to be treated, the compositions may be administered alone or in combination with other treatments, either simultaneously or sequentially.

[0511] method

[0512] Preparation method

[0513] The antibodies described herein can be produced using recombinant methods and compositions, for example as described in U.S. Pat. No. 4,816,567. In one embodiment, an isolated nucleic acid encoding an antibody described herein is provided. Such nucleic acids may encode an amino acid sequence comprising the VL of an antibody and / or an amino acid sequence comprising the VH of an antibody (e.g., a light chain and / or a heavy chain of an antibody) or an amino acid sequence comprising the VHH of a single domain antibody. In another embodiment, one or more vectors (e.g., expression vectors) comprising such nucleic acids are provided. In one embodiment, the nucleic acid is provided in a polycistronic vector. In another embodiment, a host cell comprising such nucleic acids is provided. In one such embodiment, the host cell comprises (e.g., has been transformed as follows): (1) a vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of an antibody and an amino acid sequence comprising the VH of an antigen-binding polypeptide construct; or (2) a first vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of an antigen-binding polypeptide construct, and a second vector comprising a nucleic acid encoding an amino acid sequence comprising the VH of an antigen-binding polypeptide construct. In one embodiment, the host cell is a eukaryotic cell, such as a Chinese hamster ovary (CHO) cell or a human embryonic kidney (HEK) cell or a lymphoid cell (e.g., a Y0, NS0, Sp20 cell). In one embodiment, a method for making an antibody is provided, wherein the method comprises culturing a host cell comprising a nucleic acid encoding an antibody as provided above under conditions suitable for expressing the antibody and optionally recovering the antibody from the host cell (or host cell culture medium).

[0514] For recombination production of antibodies, nucleic acids encoding antibodies such as those described above are separated and inserted into one or more vectors for further cloning and / or expression in host cells. Such nucleic acids can be easily separated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that can specifically bind to genes encoding antibody heavy and light chains).

[0515] When the antibody or variant thereof is produced by a host cell in a recombinant manner, in certain embodiments, the protein is present at about 30%, about 25%, about 20%, about 15%, about 10%, about 5%, about 4%, about 3%, about 2%, or about 1% or less of the dry weight of the cells. When the antibody or variant thereof is produced by a host cell in a recombinant manner, in certain embodiments, the protein is present in the culture medium at about 5 g / L, about 4 g / L, about 3 g / L, about 2 g / L, about 1 g / L, about 750 mg / L, about 500 mg / L, about 250 mg / L, about 100 mg / L, about 50 mg / L, about 10 mg / L, or about 1 mg / L or less of the dry weight of the cells. In certain embodiments, "substantially purified" antibodies produced by the methods described herein have a purity level of at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, specifically, at least about 75%, 80%, 85%, and more specifically, a purity level of at least about 90%, at least about 95%, at least about 99% or greater, as determined by an appropriate method (e.g., SDS / PAGE analysis, RP-HPLC, SEC, and capillary electrophoresis).

[0516] Suitable host cells for cloning or expressing antibody-encoding vectors include prokaryotic or eukaryotic cells described herein.

[0517] Recombinant host cells or host cells are cells comprising exogenous polynucleotides, regardless of the method used for insertion (e.g., direct uptake, transduction, f-pairing, or other methods known in the art for producing recombinant host cells). Exogenous polynucleotides can be maintained as non-integrated vectors, such as plasmids, or can be integrated into the host genome. Host cells can include CHO, CHO derivatives, NS0, Sp20, CV-1, VERO-76, HeLa, HepG2, Per.C6, or BHK.

[0518] For example, antibodies can be produced in bacteria, especially when glycosylation and Fc effector functions are not required. Regarding the expression of antibody fragments and polypeptides in bacteria, see, for example, U.S. Patent Nos. 5,648,237, 5,789,199 and 5,840,523 (see also Charlton, Methods in Molecular Biology, Vol. 248 (BKCLo, Humana Press, Totowa, NJ, 2003), pp. 245 to 254, which describes the expression of antibody fragments in Escherichia coli). After expression, antibodies can be separated from bacterial cell paste in soluble fractions and can be further purified.

[0519] In addition to prokaryotes, eukaryotic microorganisms such as filamentous fungi or yeast are suitable cloning or expression hosts for antibody encoding vectors, including fungi and yeast strains whose glycosylation pathways have been "humanized" so that the antibodies produced have a partially or fully human glycosylation pattern. See Gerngross, Nat. Biotech. 22: 1409-1414 (2004), and Li et al., Nat. Biotech. 24: 210-215 (2006).

[0520] Suitable host cells for expressing glycosylated antibodies also come from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant and insect cells. A variety of baculovirus strains have been identified that can be used in conjunction with insect cells, particularly for transfecting Spodoptera frugiperda cells.

[0521] Plant cell cultures can also be used as hosts. See, e.g., U.S. Pat. Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429 (describing PLANTIBODIES for producing antibodies in transgenic plants). TM technology).

[0522] Vertebrate cells can also be used as hosts. For example, mammalian cell lines adapted to growth in suspension may be suitable. Other examples of suitable mammalian host cell lines are monkey kidney CV1 cell line transformed by SV40 (COS-7); human embryonic kidney cell line (293 or 293 cells, as described, e.g., in Graham et al., J Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK); mouse sertoli cells (TM4 cells, as described, e.g., in Mather, Biol. Reprod. 23:243-251 (1980)); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical carcinoma cells (HELA); canine kidney cells (MDCK); buffalo rat liver cells (BRL 3A); human lung cells (W138); human hepatocytes (Hep G2); mouse mammary tumor (MMT 060562); TRI cells, such as, e.g., Annals NY Acad. Sci. 383: 44-68 (1982); MRC5 cells; and FS4 cells. Other suitable mammalian host cell lines include: Chinese hamster ovary (CHO) cells, including DHFR-CHO cells (Urlaub et al., Proc. Natl. Acad Sci. USA 77: 4216 (1980)); and myeloma cell lines, such as Y0, NS0 and Sp2 / 0. For a review of certain mammalian host cell lines suitable for antibody production, see, for example, Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (BKCLo ed., Humana Press, Totowa, NJ), pp. 255-268 (2003).

[0523] In one embodiment, the antibodies described herein are produced in stable mammalian cells by a method comprising: transfecting at least one stable mammalian cell with: nucleic acids encoding the antibodies at a predetermined ratio; and expressing the nucleic acids in at least one mammalian cell. In some embodiments, the predetermined ratio of nucleic acids is determined in a transient transfection experiment to determine the relative ratio of input nucleic acids that produces the highest percentage of antibody in the expression product.

[0524] In some embodiments are methods of producing an antibody in a stable mammalian cell as described herein, wherein the expression product of at least one stable mammalian cell comprises a greater percentage of the desired glycosylated antibody compared to a monomeric heavy or light chain polypeptide or other antibody.

[0525] In some embodiments, a method for producing a glycosylated antibody in a stable mammalian cell as described herein comprises identifying and purifying the desired glycosylated antibody. In some embodiments, the identification is by one or both of liquid chromatography and mass spectrometry.

[0526] If necessary, the antibody can be purified or isolated after expression. Proteins can be isolated or purified by a variety of methods known to those skilled in the art. Standard purification methods include chromatography techniques, including ion exchange, hydrophobic interaction, affinity, size determination or gel filtration and reverse phase, which are performed at atmospheric pressure or under high pressure using systems such as FPLC and HPLC. Purification methods also include electrophoresis, immunoassay, precipitation, dialysis and chromatofocusing techniques. Ultrafiltration and diafiltration techniques combined with protein concentration are also applicable. As is well known in the art, a variety of natural proteins bind to Fc and antibodies, and these proteins can be used in the present invention to purify antibodies. For example, bacterial proteins A and G bind to the Fc region. Similarly, bacterial protein L binds to the Fab region of some antibodies. Purification can often be achieved by specific fusion partners. For example, glutathione resins (if GST fusions are used), Ni +2 The antibody is purified by affinity chromatography (if a His tag is employed) or by immobilized anti-flag antibody (if a flag tag is used). For general guidance in suitable purification techniques, see, e.g., Protein Purification: Principles and Practice, 3rd Edition, Scopes, Springer-Verlag, NY, 1994, which is incorporated by reference in its entirety. The degree of purification required will vary depending on the use of the antibody. In some cases, no purification is required.

[0527] In certain embodiments, the antibodies are purified using anion exchange chromatography, including but not limited to chromatography on Q-Sepharose, DEAE Sepharose, poros HQ, poros DEAF, Toyopearl Q, Toyopearl QAE, ToyopearlDEAE, Resource / Source Q and DEAE, Fractogel Q, and DEAE columns.

[0528] In specific embodiments, the proteins described herein are purified using cation exchange chromatography, including but not limited to SP-Sepharose, CM Sepharose, poros HS, poros CM, Toyopearl SP, Toyopearl CM, Resource / Source Q and CM, Fractogel S and CM columns, and equivalents and analogs thereof.

[0529] In addition, the antibodies described herein can be chemically synthesized using techniques known in the art (see, for example, Creighton, 1983, Proteins: Structures and Molecular Principles, WH Freeman & Co., NY, and Hunkapiller et al., Nature, 310: 105-111 (1984)). For example, a polypeptide corresponding to a polypeptide fragment can be synthesized using a peptide synthesizer. In addition, if necessary, non-classical amino acids or chemical amino acid analogs can be introduced as substitutions or additions to the polypeptide sequence. In general, atypical amino acids include, but are not limited to, D-isomers of common amino acids, 2,4-diaminobutyric acid, α-aminoisobutyric acid, 4-aminobutyric acid, Abu, 2-aminobutyric acid, g-Abu, e-Ahx, 6-aminocaproic acid, Aib, 2-aminoisobutyric acid, 3-aminopropionic acid, ornithine, norleucine, norvaline, hydroxyproline, sarcosine, citrulline, homocitrulline, oxidized cysteine, tert-butylglycine, tert-butylalanine, phenylglycine, alanine cyclohexyl ester, alanine, fluoroamino acids, designer amino acids such as methyl amino acids, C-methyl amino acids, N-methyl amino acids, and general amino acid analogs. In addition, amino acids can be D (dextrorotatory) or L (levorotatory).

[0530] In certain embodiments, the aggregation temperature of the antibodies described herein is greater than about 69° C., greater than about 70° C., greater than about 71° C., greater than about 72° C., greater than about 73° C., greater than about 74° C., greater than about 75° C., or greater than about 76° C., e.g., between about 69° C. and about 77° C., between about 70° C. and about 76° C., between about 71° C. and about 75° C. In certain embodiments, the aggregation temperature is measured using DSF.

[0531] In certain embodiments, the antibodies described herein have reduced hydrophobicity compared to Lerezumab as measured by hydrophobic interaction chromatography (HIC). In certain embodiments, the antibodies exhibit a HIC retention time of less than about 15.2 min. In certain embodiments, the antibodies exhibit a HIC retention time of between about 13 min and about 15 min.

[0532] How to use

[0533] In one aspect, the application provides methods of contacting IL-13 with an anti-IL-13 antibody (eg, a human antibody or a humanized antibody) that causes inhibition of IL-13 binding to an IL-13 receptor expressed on a cell.

[0534] In one aspect, the present application provides a method of using the isolated anti-IL-13 antibodies described herein to treat a subject's condition or disease. In certain aspects, a method of using an anti-IL-13 antibody to treat a subject in need thereof is described herein, the method comprising administering to the mammalian subject a therapeutically effective amount of an anti-IL-13 antibody or a pharmaceutical composition comprising an anti-IL-13 antibody described herein. In certain embodiments, the present application provides a method of treating a subject's condition or disease associated with elevated IL-13 and / or IgE levels.

[0535] In certain aspects, described herein are methods for treating a pathology associated with IL-13 activity, comprising administering to the mammalian subject a therapeutically effective amount of an isolated anti-IL-13 antibody or a pharmaceutical composition comprising an isolated anti-IL-13 antibody described herein.

[0536] In certain aspects, described herein is a method for treating an inflammatory condition or disease in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody described herein or a pharmaceutical composition described herein. In certain embodiments of the methods described herein, the inflammatory condition or disease is atopic dermatitis. In certain embodiments, the inflammatory condition or disease is asthma. In certain embodiments, the inflammatory condition or disease is idiopathic pulmonary fibrosis. In certain embodiments of the methods described herein, the inflammatory condition or disease is alopecia areata. In certain embodiments, the inflammatory condition or disease is chronic sinusitis with nasal polyps. In certain embodiments, the inflammatory condition or disease is chronic sinusitis without nasal polyps (CRSsNP). In certain embodiments, the inflammatory condition or disease is eosinophilic esophagitis (EoE). In certain embodiments, the inflammatory disorder or disease is an eosinophilic gastrointestinal disorder or disease (ENID) selected from the group consisting of eosinophilic gastritis (EoG), eosinophilic enteritis (EoN), eosinophilic colitis (EoC), and eosinophilic gastroenteritis (EGE). In certain embodiments, the inflammatory disorder or disease is Tschag-Strauss syndrome / eosinophilic granulomatosis with polyangiitis (EGPA). In certain embodiments, the inflammatory disorder or disease is prurigo nodularis (PN). In certain embodiments, the inflammatory disorder or disease is chronic spontaneous urticaria (CSU). In certain embodiments, the inflammatory disorder or disease is chronic pruritus of unknown origin (CPUO). In certain embodiments, the inflammatory disorder or disease is bullous pemphigoid (BP). In certain embodiments, the inflammatory disorder or disease is cold-induced urticaria (ColdU). In certain embodiments, the inflammatory condition or disease is allergic fungal sinusitis (AFRS). In certain embodiments, the inflammatory condition or disease is allergic bronchopulmonary aspergillosis (ABPA). In certain embodiments, the inflammatory condition or disease is chronic obstructive pulmonary disease (COPD). In certain embodiments, the inflammatory condition or disease is inflammatory bowel disease, such as Crohn's disease or ulcerative colitis. In certain embodiments, the inflammatory condition or disease is psoriasis. In certain embodiments, the inflammatory condition or disease is lupus. In certain embodiments, the inflammatory condition or disease is rheumatoid arthritis.

[0537] In certain aspects, described herein are methods for treating a pathology associated with elevated IL-13 levels in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody or pharmaceutical composition described herein.

[0538] In certain aspects, described herein are methods of reducing the biological activity of IL-13 in a mammalian subject in need thereof, the methods comprising administering to the mammalian subject a therapeutically effective amount of an antibody or pharmaceutical composition described herein.

[0539] In certain aspects, described herein are methods for inhibiting a TH2-type allergic response in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody or pharmaceutical composition described herein.

[0540] In certain aspects, described herein are methods for inhibiting IL-13-induced STAT6 phosphorylation in a cell, the method comprising contacting the cell with an antibody described herein.

[0541] In certain aspects, described herein are methods for inhibiting IL-13-induced CD23 expression in a cell, the method comprising contacting the cell with an antibody described herein.

[0542] In certain aspects, described herein are methods for inhibiting IL-13-induced secretion of CCL2 and CCL26 from a cell, the method comprising contacting the cell with an antibody described herein.

[0543] In certain aspects, described herein are methods for inhibiting IL-13-induced NTRK1 expression in a cell, the method comprising contacting the cell with an antibody described herein.

[0544] In certain aspects, described herein are methods for reducing thymus and activation-regulated chemokine (TARC) / CCL17 levels in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of an antibody or pharmaceutical composition described herein.

[0545] In certain aspects, described herein are methods of preventing an inflammatory disorder or disease in a mammalian subject in need thereof, the methods comprising administering to the mammalian subject a therapeutically effective amount of an antibody or pharmaceutical composition described herein.

[0546] Application method

[0547] In some embodiments, the methods provided herein are useful for treating a disease or disorder in an individual. In one embodiment, the individual is a human, and the antibody is an anti-IL-13 antibody described herein.

[0548] In some embodiments, the antibody is administered intravenously, intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorbitally, by implantation, by inhalation, intrathecally, intracerebroventricularly, or intranasally. An effective amount of the anti-IL-13 antibody may be administered for the treatment of a disease or disorder. The appropriate dose of the anti-IL-13 antibody may be determined based on the type of disease or disorder to be treated, the type of anti-IL-13 antibody, the severity and course of the disease or disorder, the individual's clinical condition, the individual's clinical history and response to treatment, and the judgment of the attending physician.

[0549] In some embodiments, the antibodies provided herein are administered with at least one additional therapeutic agent. Any suitable additional therapeutic agent or immunotherapeutic agent may be administered with the antibodies provided herein. Additional therapeutic agents include agents for treating or preventing diseases or conditions associated with elevated IL-13 and / or IgE levels (e.g., but not limited to, inflammatory diseases or conditions).

[0550] The additional therapeutic agent may be administered by any suitable means. In some embodiments, the antibody provided herein and the additional therapeutic agent are included in the same pharmaceutical composition. In some embodiments, the antibody provided herein and the additional therapeutic agent are included in different pharmaceutical compositions.

[0551] In embodiments where the antibody and additional therapeutic agent provided herein are included in different pharmaceutical compositions, antibody administration can be performed before, simultaneously with, and / or after the additional therapeutic agent is administered. In some embodiments, the administration of the antibody and additional therapeutic agent provided herein is performed within approximately one month of each other. In some embodiments, the administration of the antibody and additional therapeutic agent provided herein is performed within approximately one week of each other. In some embodiments, the administration of the antibody and additional therapeutic agent provided herein is performed within approximately one day of each other. In some embodiments, the administration of the antibody and additional therapeutic agent provided herein is performed within approximately twelve hours of each other. In some embodiments, the administration of the antibody and additional therapeutic agent provided herein is performed within approximately one hour of each other.

[0552] Medicine boxes and products

[0553] The application provides a kit comprising any one or more of the antibody compositions described herein and instructions for use. In some embodiments, the kit also contains a component selected from any one of the following: a secondary antibody, an immunohistochemical analysis reagent, a pharmaceutically acceptable excipient, and an instruction manual, and any combination thereof. In a specific embodiment, the kit comprises a pharmaceutical composition comprising any one or more of the antibody compositions described herein and one or more pharmaceutically acceptable excipients.

[0554] The present application also provides an article of manufacture comprising any of the antibody compositions or kits described herein. Examples of articles of manufacture include vials (including sealed vials).

[0555] Example

[0556] Below is an example of a specific embodiment of the present invention. The examples are provided for illustrative purposes only and are not intended to limit the scope of the present invention in any way. Every effort has been made to ensure accuracy with respect to the numbers used (e.g., amounts, temperatures, etc.), but of course some experimental errors and deviations should be allowed for.

[0557] Unless otherwise indicated, the practice of the present invention will employ conventional methods of protein chemistry, biochemistry, recombinant DNA technology, and pharmacology within the skill of the art. Such techniques are fully explained in the literature. See, for example, TECreighton, Proteins: Structures and Molecular Properties (WH Freeman and Company, 1993); ALL Lehninger, Biochemistry (Worth Publishers, Inc., current edition); Sambrook et al., Molecular Cloning: A Laboratory Manual (2nd edition, 1989); Methods In Enzymology (S. Colowick and N. Kaplan, ed., Academic Press, Inc.); Remington's Pharmaceutical Sciences, 18th edition (Easton, Pennsylvania: Mack Publishing Company, 1990); Carey and Sundberg Advanced Organic Chemistry 3rd edition. (Plenum Press) Volumes A and B (1992).

[0558] method

[0559] Humanization of the mouse hybridoma sequence of the anti-IL-13 antibody 228B / C-1

[0560] Complementary determining region (CDR) transplantation technology is used to make parent mouse anti-human IL-13228B / C-1 (parent monoclonal antibody of Lerui group monoclonal antibody) humanization.Parent mouse heavy chain and light chain sequence are modeled on human antibody framework, as described below.Select one group of human heavy chain and light chain for humanization.The target is to design the improved physiological properties of producing parent antibody while retaining the pair of these heavy chains and light chains combined.For the improved developability profile during the scale expansion in biotechnology, design these humanized molecules.

[0561] Humanization of the light chain

[0562] The parent mAb light chain sequence of the mouse hybridoma sequence of the anti-IL13 antibody (Lefranc monoclonal antibody) was compared with a set of human variable region light chain (VK) germline amino acid sequences (Lefranc, M.-PIMGT, the international ImMunoGeneTics database Nucleic Acids Res., 29, D207-209 (2001). DIO: 10.1093 / nar / 29.1.207. PMID: 11125093.). A total of 4 human VK germlines were selected. Among them, one belongs to the Vk4 family (IGKV4-1), two belong to the VK1 family (IGKV1-39 and IGKV3-15), and one belongs to the VK2 family (IGKV2-28). A substitution was also designed on the light chain framework 3, with R replaced by G. This back mutation from human to mouse can change binding. Based on the sequence similarity with the mouse sequence, human germline KJ4 was selected for the J region. The humanized VL domain was cloned into a vector encoding the kappa light chain constant domain.

[0563] The following nomenclature is used for light chains: "LC0" corresponds to the mouse hybridoma sequence. "LC1" corresponds to IGKV4-1 KJ4. "LC2" corresponds to IGKV1-39KJ4. "LC3" corresponds to IGKV3-15KJ4. "LC4" corresponds to IGKV2-28KJ4. "LC5" corresponds to IGKV4-1 R to G KJ. "LC6" corresponds to IGK V1-39R to G KJ4. "LC7" corresponds to IGKV3-15R to G KJ4. "LC8" corresponds to IGKV2-28R to G KJ4.

[0564] Humanization of the heavy chain

[0565] The parent mAb heavy chain sequence of the mouse hybridoma sequence of the anti-IL13 antibody (Lerezumab) was compared with a group of human variable region heavy chain (VH) germline amino acid sequences. A total of 5 human VH germlines were selected. Among them, one belongs to the VH4 family (IGHV4-59), two belong to the VH1 family (IGHV1-46, IGHV1-69), and two belong to the VH3 family (IGHV3-15, IGHV3-23). ​​The N-terminal Q in the heavy chain was replaced with E to prevent potential pyroglutamate conversion. Based on the sequence similarity with the mouse sequence, human germline HJ6 was selected for the J region. The humanized VH domain was cloned into a vector encoding the human IgG1 HC constant domain.

[0566] The following nomenclature is used for the heavy chains: "HCO" corresponds to the mouse hybridoma heavy chain. "HCO M" corresponds to HCO NIS to TIS in FR3 (to prevent potential glycosylation). "HCl" corresponds to the humanized sequence IGHV4-59HJ6. "HC2" corresponds to the humanized sequence IGHV1-46HJ6. "HC3" corresponds to the humanized sequence IGHV1-69HJ6. "HC4" corresponds to the humanized sequence IGHV3-15HJ6. "HC5" corresponds to the humanized sequence IGHV3-23HJ6.

[0567] Gene synthesis and plasmid construction

[0568] The coding sequences of the HC and LC of the antibody were generated by DNA synthesis and PCR, and then subcloned into a pTT5-based plasmid for protein expression in a mammalian cell system. The gene sequence in the expression vector was confirmed by DNA sequencing.

[0569] Expression of antibody constructs

[0570] Transient expression of antibodies was performed by co-transfection of paired HC and LC constructs into CHO cells using the PEI method. Briefly, approximately 5.5 × 10 6 / mL CHO cells were used as hosts. Transfection was initiated by: 1 mg / L DNA and 7 mg / L PEI in OptiMEM TM The mixture in the culture medium (Invitrogen) was added to the cells and then gently mixed. Then, the cells were cultured for 9 days at 120 rpm, 37 ° C and 8% CO2 in an incubator shaker. After 24 h and thereafter depending on the cell density and viability, peptone and glucose were fed every 2-3 days. When the cell viability decreased to <80%, the cell culture was terminated on the 9th day. The conditioned medium was harvested for protein purification.

[0571] Purification of antibody constructs

[0572] Protein purification was performed by affinity chromatography and ion exchange chromatography using an AKTA purification instrument (GE Lifesciences). Conditioned medium expressing the target antibody was harvested by centrifugation at 4000 rpm for 50 min and filtered with a 0.22 μm filter. The harvested supernatant was loaded into Mabselect TM SuRe TM (GE Healthcare) column. After washing the column with buffer A (PBS, pH 7.4), the protein was eluted with buffer B (1 M glycine, pH 2.7) and immediately neutralized with 1 / 10 volume of buffer D (1 M sodium citrate, pH 6.0). Next, the affinity purified antibody buffer was exchanged into 20 mM sodium acetate pH 5.5.

[0573] Table 11: Size Exclusion Chromatography of Anti-IL-13 Constructs

[0574]

[0575]

[0576]

[0577] *See construct sequences in Tables 2-8.

[0578] SEC-HPLC analysis of antibody constructs

[0579] Analytical SEC-HPLC was performed using a Shimadzu LC-10 HPLC instrument (Shimadzu Corp.). 20 μl of 1 mg / mL sample was loaded into 200Increase5 / 150GL column (GE Lifesciences). The mobile phase was 2*PBS, the flow rate was 0.3 mL / min, and the duration was 15 min.

[0580] Measuring antibody-IL13 binding kinetics using surface plasmon resonance

[0581] The Biacore 8K SPR system (GE HealthCare) equipped with S-series sensor chip Protein G (Cytiva, catalog number 29179315) was used to determine the binding kinetic rates and affinity constants at 25°C in HBS-EP+ (10 mM HEPES pH 7.4, 150 mM NaCl, 3 mM EDTA, 0.05% surfactant P20). After a stabilization period in the electrophoresis buffer, the anti-IL13 mAb construct (diluted to 1 μg / mL) was captured onto flow cell 2 (active) for 60 seconds at a flow rate of 10 μL / min. Recombinant human IL-13 protein, His tag (Acro catalog number IL3-H52H4) was prepared at concentrations of 0, 0.39, 0.78, 1.56, 3.13, 6.25, 12.5 and 0 nM and injected into flow cell 1 (reference) and flow cell 2 (active) for 180 seconds at a flow rate of 30 μL / min. Recombinant cynomolgus macaque IL-13 protein, His tag (SINO BIOLOGICAL, catalog number 11057-C07H) was prepared at concentrations of 0, 0.39, 0.78, 1.56, 3.13, 6.25, 12.5, 25 and 0 nM and injected into flow cell 1 (reference) and flow cell 2 (active) for 180 seconds at a flow rate of 30 μL / min. The capture surface was regenerated by injecting glycine pH 1.5 at a flow rate of 30 μL / min for 30 seconds, and the samples were injected onto the freshly captured mAb in a multi-cycle manner. Data were processed and analyzed using Biacore InsightEvaluation software version 2.0.15.12933 (GE Healthcare) as follows. The reaction of flow cell 1 (reference) was subtracted from the reaction of flow cell 2 (active). Next, the reaction of two buffer blank injections was subtracted from the reference-subtracted data (2-1) to obtain the data of the double reference, which was fitted to a 1:1 binding model to determine the apparent binding (ka) and dissociation rate constants (kd). Its ratio provides the apparent equilibrium dissociation constant or affinity constant (KD=kd / ka).

[0582] Determine antibody affinity for Fc receptors and C1q

[0583] The binding affinity (K) of the antibodies to Fc receptors and C1q was determined using Biacore 8K via surface plasmon resonance (SPR). D). Briefly, an SPR chip functionalized with an anti-κ light chain antibody was used to capture purified antibodies normalized to 5 mg / mL with a flow rate of 10 μL / min for 90 or 120 seconds. A paired channel with buffer only was used as a reference. Subsequently, varying concentrations of recombinant human CD32a(167H), CD32a(167R), CD32b, CD16a(176V), CD16a(176F), FcRn, CD64 and C1q were injected over the surface with the captured purified antibodies as well as the reference channel. Regeneration of the chip between different antigens at different concentrations was performed with 10 mM glycine HCl pH 1.5 and the antibodies were captured again. K was then derived from it using a 1:1 Langmuir binding model or a steady-state analysis model (whichever was applicable). D The values ​​of association and dissociation rate constants were determined by BiacoreInsight evaluation software.

[0584] Assessment of blocking by cell line-based assays

[0585] Multiple assays were used to assess the blocking of the complete signaling complex of IL-13 / IL13Rα1 / IL-4Rα and the prevention of downstream signaling. Briefly, HEK293 previously transduced to stably express both hIL-13Rα and hIL-4Rα were cultured and harvested. Cells were seeded at 200,000 cells per well (100 μL). The cells were washed and the supernatant was discarded. 100 μL of a previously prepared and incubated 1 hour biotinylated hIL-13 and purified antibody mixture (1:1, by volume) was added to resuspend the cells, resulting in a final concentration of 0.05 μg / mL hIL-13 and 0-100 nM purified antibody. The cells were stained in this mixture at 4°C for 1 hour. The cells were then washed and stained with 100 μL of avidin conjugated to Alexa Fluor 488 diluted 1:1000 to detect the binding of biotinylated hIL-13 on the cell surface. The cells were incubated at 4°C in the dark for 1 hour. Next, the cells were washed and the median fluorescence intensity (MFI) of the cells in each well was recorded by FACS using a BD FACSCanto II. Subsequent data were analyzed using GraphPad Prism. 50 The value was determined as the antibody concentration required to inhibit by 50% the maximum MFI detected on the biotinylated hIL-13 surface incubated with 0.05 μg / mL hIL-13 alone.

[0586] Cell line based assays include: inhibition of STAT6 phosphorylation in HT-29 cells, inhibition of TARC release in A549 cells, and inhibition of TF-1 cell proliferation. Primary human lymphocyte based assays include: inhibition of STAT6 phosphorylation and inhibition of CD23 expression.

[0587] Inhibition of STAT6 phosphorylation in HT-29 cells

[0588] The functional activity of the antibody blocking the biological activity induced by IL-13 was evaluated using the inhibition of STAT6 phosphorylation in HT-29 cells. In short, HT-29 cells were starved overnight in RMPI 1640+0.1% FBS. Cells were collected and inoculated with 50,000 cells / well (100 μL). At the same time, 100 μL hIL-13 and purified antibody mixture (1: 1, by volume) were added to the same well to produce a final concentration of 10 ng / mL hIL-13 and 0-50 nM purified antibody. Cells were incubated at 37°C for 1 hour, and then fixed, permeated, and stained with PE-conjugated anti-pSTAT6 antibodies. By FACS, BD FACSCanto II was used to record the MFI of cells in each well, and GraphPad Prism was used to analyze subsequent data. The IC50 value was determined as the antibody concentration required to inhibit 50% of the maximum MFI of pSTAT6 detected in the case of 10 ng / mL hIL-13 alone. The results are summarized below.

[0589] Inhibition of TARC release in A549 cells

[0590] The functional activity of the biological activity of antibody blocking IL-13 induction was evaluated using the inhibition of A549 cell secretion of TARC. In short, A549 cells were inoculated in DMEM+10% FBS with 20,000 cells (100 μL) and cultured overnight at 37°C. The next day, the cell culture medium was discarded, and the cells were gently washed with fresh culture medium. A mixture of 150 μL hIL-13, purified antibodies and hTNFa (1: 1: 1, by volume) was added to each well to produce a final concentration of 20 ng / mL hIL-13, 0-100 nM purified antibodies and 200 ng / mL. Cells were incubated in this mixture at 37°C for 20-24 hours. After incubation, culture supernatants were collected, and the presence of TARC was analyzed using a commercially available TARC ELISA kit (R&D Systems), analyzed according to the manufacturer's instructions. The TARC concentration measured in each well was analyzed using GraphPad Prism. IC50 values ​​were determined as the antibody concentration required to inhibit 50% of the maximum TARC concentration detected in the presence of 20 ng / mL hIL-13 alone and 200 ng / mL hTNFα incubation. The results are summarized below.

[0591] Inhibition of TF-1 cell proliferation

[0592] The functional activity of antibodies blocking IL-13-induced biological activity was evaluated using the proliferation of TF-1 cells or their inhibition. In brief, TF-1 cells were harvested and starved for 4 hours in RPMI1640 + 10% FBS without additional cytokines. During this time, a mixture of hIL-13 and purified antibodies was prepared (1: 1, by volume), and 50 μL was added to each well. After starvation, TF-1 cells were harvested again and inoculated at 15,000 cells / well (50 μL), resulting in a final concentration of 4 ng / mL hIL-13 and 0-5 nM purified antibodies. The cells were then incubated at 37°C for 72 hours, and CellTiter-Glo (Promega) was used to quantify cell proliferation according to the manufacturer's instructions. Luminescence was recorded by a SpectraMax M5 multi-mode plate reader, and the data was analyzed using GraphPad Prism. The IC50 value was determined as the antibody concentration required to cause 50% of the maximum luminescence detected when TF-1 cells were incubated and cultured with 4 ng / mL hIL-13 alone. The results are summarized below.

[0593] Inhibition of STAT6 phosphorylation and CD23 expression in primary human lymphocytes

[0594] To confirm the antagonistic activity of the antibodies herein in primary cells, human peripheral blood mononuclear cells (PBMCs) obtained from healthy donors were used to evaluate the ability of the antibodies herein to inhibit IL-13-induced STAT6 phosphorylation and upregulation of CD23 expression.

[0595] Frozen human PBMCs from previously identified IL-13 reactive donors were thawed and revived. In order to determine the in vitro efficacy of the antibodies herein in inhibiting STAT6 phosphorylation, total PBMCs were stimulated with 10 ng / mL IL-13 and purified antibodies (1: 1, by volume). These cells were incubated at 37°C for 15 minutes. Subsequently, STAT6 phosphorylation was analyzed by flow cytometry using commercially available anti-phosphoSTAT6 antibodies, and staining within a specific immune population gated by lineage-specific markers CD14 and CD19 was assessed. In a group of independent PBMCs, cells were incubated at 37°C for 24 hours. Subsequently, CD23 expression was analyzed by flow cytometry using anti-CD23 antibodies, and its staining intensity within a specific immune population gated by lineage-specific markers CD14 and CD19 was assessed.

[0596] Median fluorescence intensity (MFI) data of pSTAT6 or CD23 staining were analyzed using GraphPad Prism. 50 Values ​​were determined as the antibody concentration required to elicit 50% of the maximal MFI for each marker detected when primary human PBMCs were incubated and cultured with 10 ng / mL of hIL-13 alone.

[0597] Protein Thermal Stability Testing by Differential Scanning Fluorometry (DSF)

[0598] Orange (Thermo Fisher #S6651) is supplied at 5000X concentration in 100% DMSO and diluted to 40X in the appropriate formulation buffer. The antibody was mixed with the dye and nine microliters of this mixture was loaded into a UNi (Unchained Labs, catalog number 201-1010) and stained with a "T using SYPRO m The application was run on UNCLE (UnchainedLabs). The sample was subjected to a thermal ramp from 25 to 95°C with a ramp rate of 0.5°C / min and excited at 473 nm. The full spectrum was collected at 250-720 nm, and the area under the curve between 510-680 nm was measured using UNCLE software to calculate the inflection point (T m ).

[0599] HIC-HPLC analysis of antibody constructs

[0600] Using the Thermo UltiMate TM Analytical HIC-HPLC was performed on a Thermo Scientific 3000 instrument. 20 μl of 1 mg / mL sample was loaded into TM MAbPac TM HIC-Butyl HPLC column (5 μm, 4.6 mm×100 mm; catalog number 088558). Mobile phase A was 1.5 M ammonium sulfate + 50 mM PB buffer + 5% (v / v) isopropanol, pH 6.95; and mobile phase B was 50 mM PB buffer + 20% (v / v) isopropanol, pH 6.95. The gradient was 0% to 100% mobile phase B, 20 min, and the flow rate was 0.5 mL / min.

[0601] Inhibition of CCL26 and CCL2 secretion and NTRK1 expression

[0602] The functional activity of antibodies blocking IL-13-induced biological activity was evaluated using the CCL26 and CCL2 secretion of HaCaT cells and the inhibition of NTRK1 expression. HaCaT cells were seeded in 100 μL of DMEM + 10% FBS at 20,000 cells and cultured overnight at 37°C. The next day, a mixture of 150 μL of hIL-13 and purified antibodies was added to each well to produce a final concentration of 50 ng / mL IL-13 and 0-206.5 nM purified antibodies. The cells were then further incubated at 37°C for 48 hours. After incubation, the culture supernatant was collected and the levels of secretory CCL26 and CCL2 were measured using a commercially available Luminex-based immunoassay kit (R&D Systems) and analyzed according to the manufacturer's instructions. The measured concentrations of CCL26 and CCL2 in each well were analyzed using GraphPad Prism. IC 50 The value was determined as the antibody concentration required to inhibit 50% of the maximum concentration detected in the case of 50 ng / mL IL-13 incubated alone.

[0603] The remaining cells in the assay plate were lysed and mRNA was extracted using a commercially available Quantigene kit (ThermoFisher) for analysis of NTRK1 gene expression. The level of NTRK1 mRNA was determined according to the manufacturer's protocol and analyzed in GraphPad Prism. NTRK1 gene expression was quantified as the ratio of NTRK1 mRNA level relative to the housekeeping gene PPIB, and IC 50 Values ​​were calculated as the antibody concentration required to inhibit 50% of the maximal gene expression detected using 50 ng / mL hIL-13 alone.

[0604] Example 1: Engineered anti-IL-13 antibodies exhibit improved affinity for IL-13 and its blocking potency

[0605] result

[0606] Determination of antibody affinity for IL-13

[0607] Using the methods described above, the affinity and binding kinetics of construct 133 (see construct sequences in Tables 2-8) for IL-13 were evaluated using surface plasmon resonance (SPR) compared to dupilumab, lerezumab, and lerezumab variants with one or more amino acid substitutions in the heavy chain constant region (Construct 2 (lerezumab-HC; lerezumab-LC; hIgG1-LAGAYTE; human K LC); see construct sequences in Tables 2-8), and tralokinumab.

[0608] As measured by SPR, construct 133 had an affinity of 77 pM compared to 131 pM and 116 pM for leromezumab and troloxetine, respectively.

[0609] Variants of lerezumab with one or more amino acid substitutions in the heavy chain constant region (constructs 128-131), variants of constructs 15 or 98 with one or more amino acid substitutions in the heavy chain constant region (constructs 133-136 or 137-140, respectively), and variants with one or more amino acid substitutions in the heavy chain constant region (constructs 132 and 141-144) were also evaluated for affinity and binding kinetics to human and cynomolgus monkey IL-13 using SPR.

[0610] It was observed that all antibodies bound to human IL-13 with low picomolar affinities comparable to the variants of leromemab. In addition, all tested antibodies had cross-reactivity with cynomolgus monkey IL-13 with sub-nanomolar affinities (Table 12).

[0611] Table 12. Affinity of antibodies for IL-13

[0612]

[0613] *See construct sequences in Tables 2-8.

[0614] NT - Not tested

[0615] Antibody affinity for Fc receptors and C1q

[0616] Compared to lerezumab, all antibody hlgG1-LALAYTE variants showed near or complete abrogation of binding to all Fcγ receptors, significantly reduced binding to C1q, and significantly increased binding to FcRn at pH 5.8 (Table 13).

[0617] Table 13. Affinity of antibodies for Fc receptors and C1q

[0618]

[0619] *See construct sequences in Tables 2-8.

[0620] nd - Not Detectable

[0621] Blocking

[0622] A. Inhibition of IL-13 binding to cells overexpressing hIL-13Rα / hIL-4Rα

[0623] Binding of IL-13 to cells overexpressing hIL-13Rα / hIL-4Rα was used to evaluate the functional blocking of the antibodies against this binding interaction. The results of functional blocking of the antibodies described herein in blocking the binding of IL-13 to cells overexpressing hIL-13Rα / hIL-4Rα are provided in Tables 14 and Figure 2 In cell line-based assays, compared with the IC 50 1.11 nM, construct 133 exhibited an IC of 0.89 nM 50 , and inhibited IL-13 binding on cell lines overexpressing IL-13Rα1 / IL-4Rα.

[0624] Table 14. Blockade of IL-13 binding to cells overexpressing hIL-13Rα / hIL-4Rα

[0625]

[0626]

[0627] *See construct sequences in Tables 2-8.

[0628] B. Inhibition of IL-13-induced STAT6 phosphorylation in HT-29 cells

[0629] The inhibition of STAT6 phosphorylation in HT-29 cells was used to evaluate the functional activity of the antibodies in blocking IL-13-induced biological activity. The IC of construct 133 for inhibiting STAT6 phosphorylation in HT-29 cells was observed to be 0.16 nM for pilumab, 0.23 nM for lerumab, and 0.41 nM for troloxetine. 50It is 0.28nM.

[0630] Variants of nerezumab with one or more amino acid substitutions in the heavy chain constant region (constructs 128-131), variants of constructs 15 or 98 with one or more amino acid substitutions in the heavy chain constant region (constructs 133-136 or 137-140, respectively), and variants with one or more amino acid substitutions in the heavy chain constant region (constructs 132 and 141-144) were also tested in the same assay (Tables 15 and 15). Figure 3 ).

[0631] Table 15. Inhibition of IL-13-induced STAT6 phosphorylation in HT-29 cells

[0632]

[0633]

[0634] *See construct sequences in Tables 2-8.

[0635] C. Inhibition of IL-13TARC secretion by engineered anti-IL13 antibody variants

[0636] Human A549 cells express the IL4Rα / IL13Rα1 receptor, which responds to binding to human IL-13 by inducing pSTAT6 phosphorylation, triggering the expression of downstream genes involved in TH2-type allergic responses. Thymus and activation-regulated chemokine (TARC), also known as CCL17, is one such gene product that is secreted by a variety of cell types and plays a role in attracting effector immune cells involved in inflammation, such as eosinophils. A549 cells were contacted with an engineered anti-IL-13 antibody, and a TARC assay was performed ( Figure 4 ). Anti-IL-13 antibodies inhibited TARC secretion as measured by ELISA. TARC secretion IC 50 The profile (Table 16) was similar to that of lerezumab, thus confirming the retention of the potency of the anti-IL13 antibody for IL-13 chelator activity in the cell-based assay.

[0637] Table 16. TARC secretion

[0638]

[0639] *See construct sequences in Tables 2-8.

[0640] Variants of nerolizumab with one or more amino acid substitutions in the heavy chain constant region (constructs 128-131), variants of constructs 15 or 98 with one or more amino acid substitutions in the heavy chain constant region (constructs 133-136 or 137-140, respectively), and variants with one or more amino acid substitutions in the heavy chain constant region (constructs 132 and 141-144) were tested in the same assay (Tables 17 and 18). Figure 5 The IC of construct 133 for inhibiting TARC release in A549 cells was observed to be 1.11 nM for piloumab, 0.74 nM for lerumab, and 4.14 nM for troloxetine, respectively. 50 It is 0.86nM.

[0641] Table 17. Inhibition of IL-13-induced TARC release in A549 cells

[0642]

[0643]

[0644] *See construct sequences in Tables 2-8.

[0645] D. Inhibition of IL-13-induced TF-1 cell proliferation

[0646] The IC of construct 133 for inhibiting IL-13-induced TF-1 cell proliferation was observed to be 0.19 nM for piloumab, 0.20 nM for lerumab, and 0.59 nM for troloxetine, respectively. 50 is 0.16nM.

[0647] Variants of lerezumab with one or more amino acid substitutions in the heavy chain constant region (constructs 128-131), variants of constructs 15 or 98 with one or more amino acid substitutions in the heavy chain constant region (constructs 133-136 or 137-140, respectively), and variants with one or more amino acid substitutions in the heavy chain constant region (constructs 132 and 141-144) were also tested in the same assay (Tables 18 and 19). Figure 6 ).

[0648] Table 18. Inhibition of IL-13-induced TF-1 cell proliferation

[0649]

[0650]

[0651] *See construct sequences in Tables 2-8.

[0652] E Inhibition of IL-13-induced STAT6 phosphorylation and CD23 expression in primary human lymphocytes

[0653] In primary human lymphocytes, construct 133 potently blocked IL-13 activity in a dose-dependent manner, as evidenced by inhibition of STAT6 phosphorylation with an IC of 0.44 nM. 50 , compared to 0.38nM of Lerezumab; and the IC 50 0.85 nM, compared to 0.81 nM for leromemab. The results demonstrated that construct 133 had a strong antagonistic activity against IL-13-mediated signaling in primary human cells.

[0654] A variant of construct 98 with one or more amino acid substitutions in the heavy chain constant region (construct 137), and a variant with one or more amino acid substitutions in the heavy chain constant region (construct 141) were also tested in the same assay (Tables 19, Figure 7 and Figure 8 ).

[0655] Table 19. Inhibition of IL-13-induced STAT6 phosphorylation and CD23 in human PBMCs

[0656]

[0657]

[0658] Example 2: Improved expression of engineered anti-IL-13 antibodies relative to leromemab

[0659] Transient expression of antibodies was performed by co-transfection of paired HC and LC constructs into CHO cells using the PEI method as described above. Next, the relative expression of engineered antibody constructs compared to leromemab from CHO cell lysates was determined in a small-scale expression screening experiment (Table 20).

[0660] Table 20. Expression of engineered anti-IL-13 antibodies

[0661]

[0662]

[0663] *See construct sequences in Tables 2-8.

[0664] Example 3: Thermal stability of engineered anti-IL-13 antibodies

[0665] Differential scanning fluorimetry (DSF) was used to measure the melting temperature (Tm2) of the lerezumab and IL-13 antibodies described in Table 21. Although most variants exhibited a non-inferior Tm2 profile relative to lerezumab, there were a few variants that exhibited significantly higher melting temperatures, such as constructs 5 and 15 (Table 21). In addition, almost all variants exhibited a higher aggregation temperature Tagg than lerezumab. Tagg is a measure of the tendency of mAbs to form higher molecular weight aggregates, and higher values ​​are more desirable. Therefore, most engineered variants have more desirable aggregation temperatures.

[0666] Table 21. Thermal stability of engineered anti-IL-13 antibodies

[0667]

[0668]

[0669] *See construct sequences in Tables 2-8.

[0670] Example 4: Engineered anti-IL-13 antibodies exhibit reduced hydrophobicity

[0671] Hydrophobic interaction chromatography (HIC) was performed to measure the propensity of the engineered anti-IL-13 antibodies to interact with hydrophobic surfaces (Table 22). Shorter retention times indicate less hydrophobicity. All tested novel IL-13 antibodies showed shorter retention times (RT) compared to Lerezumab. Therefore, all tested engineered anti-13 antibodies exhibited reduced hydrophobicity compared to Lerezumab.

[0672] Table 22. Hydrophobicity of engineered anti-IL-13 antibodies

[0673]

[0674]

[0675] *See construct sequences in Tables 2-8.

[0676] Example 6: Engineered anti-IL13 antibody variants and Lerezumab have the same epitope on IL-13

[0677] Epitope binning describes a technique for characterizing whether two antibodies specific for the same target (in this case, IL-13) can each bind to the target at the same time. mAb pairs are grouped together if they block each other's ability to bind to the target antigen. mAb pairs grouped together typically bind to the same or overlapping epitopes on the antigen.

[0678] To characterize the binding of construct 133 (which comprises SEQ ID NOs: 3, 39, 439, and 469) relative to Lerezumab, Lerezumab was immobilized on the surface of a sensor chip capable of measuring mAb-antigen interactions. IL-13 was first injected into the flow channel, where the binding of IL-13 to Lerezumab produced a reaction. Subsequently, construct 133 was injected into the flow channel, and the interaction reaction was recorded. In these studies, no reaction was observed after injection of construct 133 (see results in Table 23). This indicates that construct 133 and Lerezumab are grouped together, and provides evidence supporting that the two mAbs may bind to similar or identical epitopes on IL-13.

[0679] Table 23. Antibody Binding

[0680] Competing Antibody or Construct ID* Immobilized lerezumab Lerezumab + Traloxinumab - Cendakimab - Construct 133 +

[0681] + indicates antibodies grouped with leromemab.

[0682] -Indicates antibodies not grouped with leromemab.

[0683] *See construct sequence 133 in Table 2-8.

[0684] In similar studies, it was found that tralokumab-ldrm (Adbry TM ) had a grouped reaction, indicating that it and lerezumab have different epitopes on IL-13.

[0685] To further characterize the epitopes of construct 133 and leruizumab, hydrogen-deuterium exchange mass spectrometry (HDX-MS) and cross-linking mass spectrometry (XL-MS) were performed as known in the art. In short, HDX-MS was performed using a mixed solution of IL-13 and purified antibodies at a concentration of 20 μM or a final concentration of 20 μM and 40 μM, respectively. Incubation times of 15 s, 60 s, 180 s, 600 s, 1800 s, and 7200 s were performed before the quenching exchange reaction, and the protein mixture was subjected to proteolysis, followed by LC-MS using Waters Q-ToF Xevo G2-XS. Determination of deuterium incorporation of both IL-13 and the mixture, and the peptide region in which deuterium incorporation was inhibited was considered to be possibly involved in binding to purified antibodies. In short, XL-MS was performed using a mixed solution of IL-13 and purified antibodies, wherein the final concentrations were 14.7 μM and 0.7 μM, respectively. 20 μL of this mixture was mixed with 2 μL of DSS (2 mg / mL stock in DCM) and the final solution was incubated at room temperature for 180 minutes. After this incubation, the samples were subjected to proteolysis using trypsin, chymotrypsin, ASP-N, elastase and thermolysin and analyzed using LC-MS using Q-Exactive MS. Peptides were referenced against peptides identified from the previous peptide mapping experiments of IL-13 alone. IL-13 peptide regions that showed cross-linking with corresponding peptides from the antibody variable region were considered to be potentially involved in binding to the purified antibody. Both methods provided complementary data indicating specific amino acid regions where construct 133 and Lerezumab may bind. This provides further evidence to support that the two mAbs may bind to highly overlapping epitopes on IL-13 ( Figure 1 ).

[0686] These studies provide evidence that construct 133 binds to the same region on IL-13 as lerezumab and is therefore more likely to have the same biological effect than if construct 133 recognized a different region.

[0687] Example 7: Engineered anti-IL-13 antibody variants demonstrate significantly extended half-life in pharmacokinetic analysis of NHP and anti-IL-13 antibodies

[0688] Anti-IL-13 antibody variants have extended half-lives in NHPs

[0689] To demonstrate the potential of engineered anti-IL13 antibody variants to improve dosing relative to current and anticipated standard of care for mAbs in Alzheimer's disease (AD) (among other diseases), construct 133, comprising SEQ ID NOs: 3, 39, 439, and 469, was studied in female non-human primates (NHPs) following a single bolus dose of 3 mg / kg given IV or SQ. Blood samples were collected serially beginning with sample pre-dose and then at 0.167, 1, 4, 8, 24, 48, 96, 168, 336, 504, 674, 840, 1334, 1680, and 2160 hours post-dose. The following PK parameters were calculated: Maximum observed serum concentration (C max ), the time to reach the maximum observed serum concentration (T max ), the area under the serum concentration versus time curve extrapolated from time 0 to infinity (AUC 0-inf ), clearance (CL), volume of distribution at steady state (V ss ), half-life (T 1 / 2 ) and absolute subcutaneous bioavailability (F). Data were analyzed to display mean serum concentrations with standard deviation over time and regression fits were performed.

[0690] In a head-to-head study of Construct 133 versus Lerezumab in NHPs, both IV and SQ formulations of Construct 133 showed significantly longer half-lives than Lerezumab. In these studies, the mean half-life of Construct 133 was 27.6 days compared to 18 days for Lerezumab. Fig. 9 In addition, construct 133 exhibited an average clearance rate (mL day ) of 1.45 in NHP. -1 kg -1 The observed steady-state distribution volume was 55.65 (mL kg -1 Construct 133 was well absorbed, with a subcutaneous bioavailability of 81.22%. Lerezumab exhibited an average clearance rate of 2.93 (mL·day) in NHPs. -1 kg -1 The observed steady-state distribution volume was 52.10 (mL kg -1). Lerezumab was well absorbed, with a subcutaneous bioavailability of 75.70%. Without being bound by theory, because construct 133 is engineered to have a YTE amino acid substitution in the Fc region, the half-life of IgG can be extended by increasing binding to the neonatal Fc receptor (FcRn) under acidic pH conditions. FcRn binds IgG to salvage recycling via lysosomes, causing IgG to return to circulation. The extended half-life of construct 133 may enable less frequent dosing than currently available treatments, which may reduce injection load and increase compliance in patients with atopic dermatitis and other IL-13 driven diseases.

[0691] In a non-head-to-head comparison with third-party NHP data, construct 133 exhibited the highest normalized AUC among antibodies with YTE substitutions 0-∞ (C norm*day ) or the area under the curve (AUC) from dosing to infinity, such as Fig.10 Thus, the PK profile of construct 133 appears to provide a maximum sustained concentration or level of drug in the bloodstream relative to the other antibodies with the YTE substitution.

[0692] The half-life extension of mAbs with YTE amino acid substitutions depends on the type of target (e.g., receptor vs. soluble). Therefore, the conversion of NHP half-life data to human half-life data for mAbs with soluble targets was investigated, and the human half-life was found to be approximately three to four times that of the NHP half-life (mean: 3.5x, median 3.1x; data not shown).

[0693] Based on this NHP half-life data it is expected that the antibodies disclosed herein (eg, construct 133) will have a human half-life of approximately 80 to 110 days based on a comparable mAb with a YTE amino acid substitution.

[0694] In addition, based on PK modeling, at a 33-day human half-life (which, to the applicant's knowledge, would be lower than the lowest half-life reported to date for mAbs with YTE amino acid substitutions and soluble targets), it is believed that the antibodies disclosed herein can be effectively administered every two months with a dosing schedule. At a 50-day half-life, it is believed that the antibodies disclosed herein can be effectively administered every three months with a dosing schedule.

[0695] In order to understand the maintenance dosing schedule that the antibodies disclosed herein may be able to achieve, the known PK parameters of Lerezumab were used. These PK parameters provide an understanding of how Lerezumab is distributed and cleared throughout the body. Based on these known parameters, a two-compartment PK model with first-order absorption, which is standard for mAbs, was constructed to predict the concentration or drug level of both Lerezumab and the antibodies disclosed herein over time. Key parameters included a clearance rate of 0.156 L / day, a central volume (Vc) of 4.10 L, a volume of 0.239 L / day, and a volume of 1.5 L / day. -1 The absorption rate (ka) and bioavailability of 85.6% were observed.

[0696] It is believed that the effects of C in inflammatory diseases such as AD are 谷 Therefore, based on the model described above, the target C of the antibodies disclosed herein is 谷 Set to be equal to once-monthly dosing of maintenance lerezumab C 谷 , which is 31.3 mg / L. Given the overlapping epitopes of leromemab and certain antibodies disclosed herein and the similarity of potency across multiple in vitro assays, the required exposure for potential clinical activity of the antibodies disclosed herein can be predicted. By modeling K 消除 , elimination rate constant or the fraction of drug eliminated in a given time, and a half-life to maintain a concentration of the disclosed antibodies above 31.3 mg / L, a half-life of at least 33 days should be required to maintain dosing of the disclosed antibodies every two months, and a half-life of at least 50 days should be required to maintain dosing of the disclosed antibodies every three months, assuming a dose of 300 mg.

[0697] Therefore, in the case of a 33-day human half-life, it is believed that the antibodies disclosed herein can be effectively administered every two months to maintain a dosing schedule. In the case of a 50-day half-life, it is believed that the antibodies disclosed herein can be effectively administered every three months to maintain a dosing schedule.

[0698] Pharmacokinetic Analysis of Anti-IL-13 Antibodies

[0699] In other experiments, multiple in vivo pharmacokinetic (PK) studies were performed in which additional variants of lerezumab, construct 15 or construct 98 having one or more amino acid substitutions in the heavy chain constant region (constructs 133-137 or 137 and 140, respectively) and / or additional variants having one or more amino acid substitutions in the heavy chain constant region (constructs 141 and 144) were also tested.

[0700] Studies were conducted using cynomolgus macaques (Macaca fascicularis) with either matched or subcutaneous (SQ) / intravenous (IV) groups of all females, ranging in weight from 1.5kg to 2.0kg; or all males, ranging in weight from 2.9kg to 3.3kg. Test agents were administered to animals at a dose of 3 mg / kg of each antibody by IV bolus and / or SQ injection on Day 0, with serum samples obtained regularly throughout the study.

[0701] PK parameters were determined from cynomolgus macaque serum samples up to day 56 (1334 hours) and in a small group up to day 90 (2160 hours), with the mean PK profile shown in Fig.11 (IV administration) and Fig.12 (SQ administration). PK analysis demonstrated that constructs 133, 134, 135, 136, 137, 140, 141 and 144 each had improved half-life and reduced serum clearance compared to those of lerezumab, as reported in Table 24. Where bioavailability (F) could be determined, constructs 133, 134 and 141 were shown to have equivalent bioavailability to lerezumab (Table 25).

[0702] Table 24. Half-life and serum clearance

[0703]

[0704]

[0705] *These values ​​were adjusted for bioavailability in the panels Lerezumab, Construct 133, Construct 134, and Construct 141.

[0706] # See construct sequences in Tables 2-8.

[0707] Table 25. Construct bioavailability

[0708] Construct ID* Bioavailability (%) Lerezumab 75.70％ Construct 133 81.22％ Construct 134 72.83％ Construct 141 75.57％

[0709] *See construct sequences in Tables 2-8.

[0710] Example 8. Inhibition of IL-13-induced CCL2 and CCL26 secretion and NTRK1 expression in HaCaT cells

[0711] The secretion of CCL26 (eotaxin-3) and CCL2 (MCP-1) by HaCaT cells (respectively Fig.13 and Fig.14 ) and NTRK1 expression ( Fig.15) was used to evaluate the functional activity of the antibody in blocking the biological activity induced by IL-13. HaCaT cells were seeded in 100 μL DMEM + 10% FBS at 20,000 cells and cultured overnight at 37°C. The next day, a mixture of 150 μL hIL-13 and purified antibodies was added to each well to produce a final concentration of 50 ng / mL IL-13 and 0-206.5 nM purified antibodies. The cells were then further incubated at 37°C for 48 hours. After incubation, the culture supernatant was collected and the levels of secretory CCL26 and CCL2 were measured using a commercially available Luminex-based immunoassay kit (R&D Systems) and analyzed according to the manufacturer's instructions. The measured concentrations of CCL26 and CCL2 in each well were analyzed using GraphPad Prism. IC 50 The value was determined as the antibody concentration required to inhibit 50% of the maximum concentration detected in the case of 50 ng / mL IL-13 incubated alone.

[0712] The remaining cells in the assay plate were lysed and mRNA was extracted using a commercially available Quantigene kit (ThermoFisher) for analysis of NTRK1 gene expression. The level of NTRK1 mRNA was determined according to the manufacturer's protocol and analyzed in GraphPad Prism. NTRK1 gene expression was quantified as the ratio of NTRK1 mRNA level relative to the housekeeping gene PPIB, and IC 50 Values ​​were calculated as the antibody concentration required to inhibit 50% of the maximal gene expression detected using 50 ng / mL hIL-13 alone.

[0713] Table 26. CCL26 and CCL2 secretion and NTRK1 expression

[0714]

[0715] *See construct sequences in Tables 2-8.

[0716] Example 9. Monomer Purity from Stable Pools after Affinity Capture

[0717] Monomer purity from one-step affinity capture is the determining factor of the final yield and unit cost of antibodies under cGMP production. In short, in order to characterize this, the CHO stable pool of each antibody is produced separately in the workflow that causes the master cell bank selection of cGMP manufacturing. Affinity capture step is carried out using Mabselect SuRe column. After one-step purification, novel IgG1 variants (such as constructs 132,133,136,137,140,141 and 144) maintain a clear solution with <15% aggregation. When using the elution buffer of 50mM sodium citrate, 150mM sodium chloride pH 3.0, novel IgG4 variants (constructs 134,135,138,139,142 and 143) have a milky appearance with some precipitation and aggregation sensitivity (>68% aggregation). The novel IgG4 variants had lower aggregation levels when eluted with 50 mM acetic acid pH 2.8 or with 100 mM sodium acetate, 800 mM arginine pH 3.5. At the higher pH of 3.5, arginine is required to retain proper reconstitution.

[0718] The results are summarized in Table 27. When compared to the leromemab variants (constructs 128-131), the novel IgG1 and IgG4 variants exhibited greater monomer purity directly beyond affinity capture with optimized elution conditions.

[0719] Table 27. Anti-IL-13 antibody monomer purity

[0720] Construct ID* One-step monomer purity (%) Construct 128 87.7 Construct 129 90.5 Construct 130 81.6 Construct 131 82.9 Construct 132 94.7 Construct 133 90.9 Construct 134 91.7 Construct 135 91.4 Construct 136 90.3 Construct 137 92.7 Construct 138 94.0 Construct 139 92.0 Construct 140 89.7 Construct 141 94.1 Construct 142 90.1 Construct 143 91.5 Construct 144 92.4

[0721] *See construct sequences in Tables 2-8.

[0722] Example 10. Accelerated stability

[0723] As shown by this example, the novel variants in the IgG1 construct have a lower tendency to aggregate and are more resistant to changes in basic species under various stress conditions, as shown in Table 28. The proportion of basic species was determined via capillary isoelectric focusing (ciEF), as is known in the art. Variants related to leruizumab (constructs 128-131) and IgG4 variants (constructs 134, 135, 142, and 143) as described in Examples 1-7 are more susceptible to changes in aggregation and basic species than novel IgG1 variants (e.g., constructs 132, 133, 136, 137, 140, 141, and 144).

[0724] Table 28. Changes in stability of anti-IL-13 antibodies

[0725]

[0726] *See construct sequences in Tables 2-8.

[0727] Evaluation of clones for monomer purity and changes in basic species were evaluated on days 0 and 14 for 40°C stability; and on days 0 and 2 for pH 3.5 stability.

[0728] NT - Not tested.

[0729] While the invention has been particularly shown and described with reference to a preferred embodiment and various alternative embodiments, it will be understood by those skilled in the relevant art that various changes in form and details may be made therein without departing from the spirit and scope of the invention.

[0730] All references, issued patents, and patent applications cited within the text of this specification are hereby incorporated by reference in their entirety for all purposes.

[0731] Informal Sequence Listing

[0732]

[0733]

[0734]

[0735]

[0736]

[0737]

[0738]

[0739]

[0740]

[0741]

[0742]

[0743]

[0744]

[0745]

[0746]

[0747]

[0748]

[0749]

[0750]

[0751]

[0752]

[0753]

[0754]

[0755]

[0756]

[0757]

[0758]

[0759]

[0760]

[0761]

[0762]

[0763]

[0764]

[0765]

[0766]

[0767]

[0768]

[0769]

[0770]

[0771]

[0772]

[0773]

[0774]

[0775]

[0776]

[0777]

[0778]

[0779]

[0780]

[0781]

[0782]

Claims

1. An isolated antibody that binds to interleukin 13 (IL-13), the antibody comprising a) a variable heavy (VH) chain sequence comprising three heavy chain CDR sequences CDR-H1, CDR-H2 and CDR-H3; and b) a variable light (VL) chain sequence comprising three light chain CDR sequences CDR-L1, CDR-L2 and CDR-L3; wherein: a. CDR-H1 comprises a sequence selected from the sequence shown in SEQ ID NO: 58-99 and 121; b. CDR-H2 comprises a sequence selected from the sequence shown in SEQ ID NO: 100-111; c. CDR-H3 comprises a sequence selected from the sequence shown in SEQ ID NO: 112-120 and 130-140, d. CDR-L1 comprises a sequence selected from the group consisting of sequences shown in SEQ ID NOs: 141-144 and 149-152, e. CDR-L2 comprises a sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 153-158 and the amino acid sequence LAS; and f. CDR-L3 comprises a sequence selected from the sequence shown in SEQ ID NO: 165-172.

2. The isolated antibody of claim 1, wherein the antibody comprises: a. CDR-H1, which comprises a sequence selected from the sequence shown in SEQ ID NO: 58-66; b. CDR-H2, which comprises a sequence selected from the sequence shown in SEQ ID NO: 100-103; c. CDR-H3, which comprises a sequence selected from the sequence shown in SEQ ID NO: 112-120, d. CDR-L1, which comprises a sequence selected from the sequence shown in SEQ ID NO: 141-144, e. CDR-L2, which comprises a sequence selected from the sequence shown in SEQ ID NO: 153-158; and f. CDR-L3, which comprises a sequence selected from the sequence shown in SEQ ID NO: 165-172.

3. The isolated antibody of claim 1, wherein the antibody comprises: a. CDR-H1, which comprises a sequence selected from the sequence shown in SEQ ID NO: 67-83; b. CDR-H2, which comprises a sequence selected from the sequence shown in SEQ ID NO: 104-107; c. CDR-H3, which comprises a sequence selected from the sequence shown in SEQ ID NO: 112-120, d. CDR-L1, which comprises a sequence selected from the sequence shown in SEQ ID NO: 141-144, e. CDR-L2, which comprises a sequence selected from the sequence shown in SEQ ID NO: 153-158; and f. CDR-L3, which comprises a sequence selected from the sequence shown in SEQ ID NO: 165-172.

4. The isolated antibody of claim 1, wherein the antibody comprises: a. CDR-H1, which comprises a sequence selected from the sequence shown in SEQ ID NO: 84-99 and 121; b. CDR-H2, which comprises a sequence selected from the sequence shown in SEQ ID NO: 108-111; c. CDR-H3, which comprises a sequence selected from the sequence shown in SEQ ID NO: 130-140, d. CDR-L1, which comprises a sequence selected from the sequence shown in SEQ ID NO: 149-152, e. CDR-L2, which comprises the amino acid sequence LAS; and f. CDR-L3, which comprises a sequence selected from the sequence shown in SEQ ID NO: 165-172.

5. The isolated antibody of any one of the above claims, wherein the antibody does not comprise: a. CDR-H1 as shown in SEQ ID NO: 58; CDR-H2 as shown in SEQ ID NO: 100; CDR-H3 as shown in SEQ ID NO: 112; CDR-L1 as shown in SEQ ID NO: 141; CDR-L2 as shown in SEQ ID NO: 153; and CDR-L3 as shown in SEQ ID NO: 165; or b. CDR-H1 shown in SEQ ID NO:67; CDR-H2 shown in SEQ ID NO:104; CDR-H3 shown in SEQ ID NO:112; CDR-L1 shown in SEQ ID NO:141; CDR-L2 shown in SEQ ID NO:153; and CDR-L3 shown in SEQ ID NO:165; or c. CDR-H1 shown in SEQ ID NO:84; CDR-H2 shown in SEQ ID NO:108; CDR-H3 shown in SEQ ID NO:130; CDR-L1 shown in SEQ ID NO:149; CDR-L2 shown in the amino acid sequence LAS; and CDR-L3 shown in SEQ ID NO:

165.

6. The isolated antibody of claim 5, wherein the antibody does not comprise any combination of: a. CDR-H1 shown in any one of SEQ ID NO: 58, 67 or 84; b. CDR-H2 shown in any one of SEQ ID NO: 100, 104 or 108; c. CDR-H3 shown in any one of SEQ ID NO: 112 or 130; d. CDR-L1 shown in any one of SEQ ID NO: 141 or 149; e. CDR-L2 shown in any one of SEQ ID NO: 153 or 154; and f. CDR-L3 shown in SEQ ID NO:

165.

7. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising a sequence as shown in any one of SEQ ID NO: 58 or 68; a CDR-H2 comprising a sequence as shown in any one of SEQ ID NO: 100 or 104; a CDR-H3 comprising a sequence as shown in SEQ ID NO: 112; a CDR-L1 comprising a sequence as shown in any one of SEQ ID NO: 141 or 149; a CDR-L2 comprising a sequence as shown in any one of SEQ ID NO: 153 or amino acid sequence LAS; and a CDR-L3 comprising a sequence as shown in SEQ ID NO:

165.

8. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:58; a CDR-H2 comprising the sequence shown in SEQ ID NO:100; a CDR-H3 comprising the sequence shown in SEQ ID NO:112; a CDR-L1 comprising the sequence shown in SEQ ID NO:141; a CDR-L2 comprising the sequence shown in SEQ ID NO:153; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

9. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:68; a CDR-H2 comprising the sequence shown in SEQ ID NO:104; a CDR-H3 comprising the sequence shown in SEQ ID NO:112; a CDR-L1 comprising the sequence shown in SEQ ID NO:141; a CDR-L2 comprising the sequence shown in SEQ ID NO:153; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

10. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:68; a CDR-H2 comprising the sequence shown in SEQ ID NO:104; a CDR-H3 comprising the sequence shown in SEQ ID NO:112; a CDR-L1 comprising the sequence shown in SEQ ID NO:149; a CDR-L2 comprising the amino acid sequence LAS; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

11. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising a sequence as shown in any one of SEQ ID NO: 58, 68 or 85; a CDR-H2 comprising a sequence as shown in any one of SEQ ID NO: 100, 104 or 108; a CDR-H3 comprising a sequence as shown in any one of SEQ ID NO: 112 or 130; a CDR-L1 comprising a sequence as shown in any one of SEQ ID NO: 141 or 149; a CDR-L2 comprising a sequence as shown in any one of SEQ ID NO: 153 or amino acid sequence LAS; and a CDR-L3 comprising a sequence as shown in SEQ ID NO:

165.

12. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:68; a CDR-H2 comprising the sequence shown in SEQ ID NO:104; a CDR-H3 comprising the sequence shown in SEQ ID NO:112; a CDR-L1 comprising the sequence shown in SEQ ID NO:141; a CDR-L2 comprising the sequence shown in SEQ ID NO:153; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

13. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:85; a CDR-H2 comprising the sequence shown in SEQ ID NO:108; a CDR-H3 comprising the sequence shown in SEQ ID NO:130; a CDR-L1 comprising the sequence shown in SEQ ID NO:149; a CDR-L2 comprising the amino acid sequence LAS; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

14. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising a sequence as shown in any one of SEQ ID NO: 58, 68 or 85; a CDR-H2 comprising a sequence as shown in any one of SEQ ID NO: 100, 104 or 108; a CDR-H3 comprising a sequence as shown in any one of SEQ ID NO: 112 or 130; a CDR-L1 comprising a sequence as shown in any one of SEQ ID NO: 141 or 149; a CDR-L2 comprising a sequence as shown in any one of SEQ ID NO: 157 or amino acid sequence LAS; and a CDR-L3 comprising a sequence as shown in SEQ ID NO:

165.

15. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:58; a CDR-H2 comprising the sequence shown in SEQ ID NO:100; a CDR-H3 comprising the sequence shown in SEQ ID NO:112; a CDR-L1 comprising the sequence shown in SEQ ID NO:141; a CDR-L2 comprising the sequence shown in SEQ ID NO:157; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

16. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:68; a CDR-H2 comprising the sequence shown in SEQ ID NO:104; a CDR-H3 comprising the sequence shown in SEQ ID NO:112; a CDR-L1 comprising the sequence shown in SEQ ID NO:141; a CDR-L2 comprising the sequence shown in SEQ ID NO:157; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

17. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising a sequence as shown in any one of SEQ ID NO: 58, 68 or 85; a CDR-H2 comprising a sequence as shown in any one of SEQ ID NO: 100, 104 or 108; a CDR-H3 comprising a sequence as shown in any one of SEQ ID NO: 112 or 130; a CDR-L1 comprising a sequence as shown in any one of SEQ ID NO: 141 or 149; a CDR-L2 comprising a sequence as shown in any one of SEQ ID NO: 157 or amino acid sequence LAS; and a CDR-L3 comprising a sequence as shown in SEQ ID NO:

165.

18. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:68; a CDR-H2 comprising the sequence shown in SEQ ID NO:104; a CDR-H3 comprising the sequence shown in SEQ ID NO:112; a CDR-L1 comprising the sequence shown in SEQ ID NO:141; a CDR-L2 comprising the sequence shown in SEQ ID NO:157; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

19. An isolated antibody as described in any one of the above claims, wherein the antibody comprises: a CDR-H1 comprising the sequence shown in SEQ ID NO:85; a CDR-H2 comprising the sequence shown in SEQ ID NO:108; a CDR-H3 comprising the sequence shown in SEQ ID NO:130; a CDR-L1 comprising the sequence shown in SEQ ID NO:149; a CDR-L2 comprising the amino acid sequence LAS; and a CDR-L3 comprising the sequence shown in SEQ ID NO:

165.

20. The isolated antibody of any one of the above claims, wherein the antibody comprises a VH sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 1-32 and 470.

21. The isolated antibody of any one of the above claims, wherein the antibody comprises a VL sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 33-57 and 471.

22. The isolated antibody of any one of the above claims, wherein the antibody comprises a VH sequence selected from the group consisting of sequences shown in SEQ ID NOs: 1-32 and 470 and a VL sequence selected from the group consisting of sequences shown in SEQ ID NOs: 33-57 and 471.

23. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence selected from the group consisting of sequences shown in SEQ ID NOs: 1-32 and 470 and a VL sequence shown in SEQ ID NO:

49.

24. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence selected from the group consisting of sequences shown in SEQ ID NOs: 1-32 and 470 and a VL sequence shown in SEQ ID NO:

51.

25. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:

51.

26. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:

471.

27. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 1 and a VL sequence as shown in SEQ ID NO:

33.

28. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:2 and a VL sequence as shown in SEQ ID NO:

33.

29. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO:

35.

30. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:4 and a VL sequence as shown in SEQ ID NO:

35.

31. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:5 and a VL sequence as shown in SEQ ID NO:

35.

32. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:6 and a VL sequence as shown in SEQ ID NO:

35.

33. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

35.

34. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO:

36.

35. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:4 and a VL sequence as shown in SEQ ID NO:

36.

36. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:5 and a VL sequence as shown in SEQ ID NO:

36.

37. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:6 and a VL sequence as shown in SEQ ID NO:

36.

38. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

36.

39. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO:

39.

40. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:4 and a VL sequence as shown in SEQ ID NO:

39.

41. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:5 and a VL sequence as shown in SEQ ID NO:

39.

42. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:6 and a VL sequence as shown in SEQ ID NO:

39.

43. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

39.

44. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:

40.

45. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:4 and a VL sequence as shown in SEQ ID NO:

40.

46. ​​The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:5 and a VL sequence as shown in SEQ ID NO:

40.

47. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:6 and a VL sequence as shown in SEQ ID NO:

40.

48. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

40.

49. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:

42.

50. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:9 and a VL sequence as shown in SEQ ID NO:

43.

51. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

39.

52. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

44.

53. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

45.

54. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

46.

55. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

47.

56. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

48.

57. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

49.

58. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

50.

59. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

51.

60. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

52.

61. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

53.

62. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

54.

63. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

55.

64. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

56.

65. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:

57.

66. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 10 and a VL sequence as shown in SEQ ID NO:

39.

67. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 11 and a VL sequence as shown in SEQ ID NO:

39.

68. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 12 and a VL sequence as shown in SEQ ID NO:

39.

69. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 13 and a VL sequence as shown in SEQ ID NO:

39.

70. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 14 and a VL sequence as shown in SEQ ID NO:

39.

71. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 15 and a VL sequence as shown in SEQ ID NO:

39.

72. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 16 and a VL sequence as shown in SEQ ID NO:

39.

73. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 17 and a VL sequence as shown in SEQ ID NO:

39.

74. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 18 and a VL sequence as shown in SEQ ID NO:

39.

75. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 19 and a VL sequence as shown in SEQ ID NO:

39.

76. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:20 and a VL sequence as shown in SEQ ID NO:

39.

77. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:21 and a VL sequence as shown in SEQ ID NO:

39.

78. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:22 and a VL sequence as shown in SEQ ID NO:

39.

79. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:23 and a VL sequence as shown in SEQ ID NO:

39.

80. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:24 and a VL sequence as shown in SEQ ID NO:

39.

81. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:25 and a VL sequence as shown in SEQ ID NO:

39.

82. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:26 and a VL sequence as shown in SEQ ID NO:

39.

83. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:27 and a VL sequence as shown in SEQ ID NO:

39.

84. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:28 and a VL sequence as shown in SEQ ID NO:

39.

85. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:28 and a VL sequence as shown in SEQ ID NO:

39.

86. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:29 and a VL sequence as shown in SEQ ID NO:

39.

87. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:30 and a VL sequence as shown in SEQ ID NO:

39.

88. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:31 and a VL sequence as shown in SEQ ID NO:

39.

89. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:32 and a VL sequence as shown in SEQ ID NO:

39.

90. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:

39.

91. The isolated antibody of claim 21, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:

51.

92. An isolated antibody as described in any one of the above claims, wherein the antibody is a humanized antibody, a human antibody, or a chimeric antibody.

93. An isolated antibody as described in any one of the above claims, wherein the antibody is a humanized antibody.

94. An isolated antibody as described in any of the above claims, wherein the antibody comprises a heavy chain human constant region of a class selected from IgG, IgA, IgD, IgE and IgM.

95. The isolated antibody of any of the above claims, wherein the human Fc region comprises a human heavy chain constant region of class IgG and a subclass selected from IgGl, IgG2, IgG3, and IgG4.

96. The isolated antibody of claim 95, wherein the human Fc region comprises a human IgG1 Fc.

97. The isolated antibody of claim 95, wherein the human Fc region comprises a human IgG4 Fc.

98. The isolated antibody of claim 95, wherein the human Fc region comprises a human IgG2 Fc.

99. An isolated antibody as described in any one of the above claims, wherein the heavy chain comprises a constant heavy chain sequence selected from the sequence shown in SEQ ID NOs: 425-468 and 484-539.

100. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

101. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 470 and a VL sequence as shown in SEQ ID NO: 471; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs: 436-468 and 484-539.

102. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 1 and a VL sequence as shown in SEQ ID NO: 33; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

103. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:2 and a VL sequence as shown in SEQ ID NO:33; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

104. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

105. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:4 and a VL sequence as shown in SEQ ID NO:35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

106. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:5 and a VL sequence as shown in SEQ ID NO:35; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

107. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:6 and a VL sequence as shown in SEQ ID NO:35; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

108. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:35; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

109. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:36; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NOs:436-468 and 484-539.

110. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:4 and a VL sequence as shown in SEQ ID NO:36; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

111. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:5 and a VL sequence as shown in SEQ ID NO:36; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

112. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:6 and a VL sequence as shown in SEQ ID NO:36; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

113. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:36; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:439, 440, 446, 457 and 460.

114. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

115. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:4 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

116. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:5 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

117. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:6 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

118. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

119. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 40; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs: 436-468 and 484-539.

120. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:4 and a VL sequence as shown in SEQ ID NO:40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

121. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:5 and a VL sequence as shown in SEQ ID NO:40; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NOs:436-468 and 484-539.

122. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:6 and a VL sequence as shown in SEQ ID NO:40; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

123. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:40; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

124. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 8 and a VL sequence as shown in SEQ ID NO: 42; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

125. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:9 and a VL sequence as shown in SEQ ID NO:43; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

126. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

127. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:44; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

128. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:45; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

129. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:46; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

130. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:47; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

131. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:48; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

132. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:49; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

133. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:50; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

134. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

135. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:52; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

136. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:53; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

137. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:54; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

138. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:55; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

139. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:56; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

140. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:57; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

141. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 10 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs: 436-468 and 484-539.

142. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 11 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

143. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 12 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

144. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 13 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

145. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 14 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

146. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 15 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs: 436-468 and 484-539.

147. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 16 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs: 436-468 and 484-539.

148. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 17 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

149. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 18 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs: 436-468 and 484-539.

150. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 19 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

151. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:20 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

152. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:21 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

153. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:22 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

154. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:23 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

155. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:24 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

156. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:25 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

157. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:26 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

158. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 27 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs: 436-468 and 484-539.

159. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:28 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

160. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:28 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

161. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:29 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

162. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:30 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the group consisting of sequences shown in SEQ ID NOs:436-468 and 484-539.

163. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:31 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

164. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:32 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

165. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

166. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequences shown in SEQ ID NOs:436-468 and 484-539.

167. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

439.

168. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

446.

169. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

457.

170. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

460.

171. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

439.

172. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

446.

173. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

457.

174. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

460.

175. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

439.

176. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

446.

177. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

457.

178. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

460.

179. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

439.

180. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

446.

181. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

457.

182. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

460.

183. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

439.

184. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

446.

185. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

457.

186. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence selected from the sequence shown in SEQ ID NO:

460.

187. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 15 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

439.

188. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 15 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

457.

189. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 15 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

460.

190. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO: 15 and a VL sequence as shown in SEQ ID NO: 39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

446.

191. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

439.

192. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

457.

193. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

460.

194. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:3 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

446.

195. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

439.

196. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

457.

197. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

460.

198. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:7 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

446.

199. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

439.

200. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:39; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

457.

201. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

460.

202. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:8 and a VL sequence as shown in SEQ ID NO:51; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

446.

203. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

439.

204. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

457.

205. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

460.

206. The isolated antibody of any one of claims 1-6, wherein the antibody comprises a VH sequence as shown in SEQ ID NO:470 and a VL sequence as shown in SEQ ID NO:471; and wherein the human Fc region comprises a human IgG sequence as shown in SEQ ID NO:

446.

207. An isolated antibody as described in any of the above claims, wherein the light chain comprises the constant light chain sequence shown by SEQ ID NO:

469.

208. An isolated antibody as described in any one of the above claims, wherein the Fc region comprises one or more amino acid substitutions, wherein the one or more substitutions cause a change in antibody half-life, ADCC activity, ADCP activity or CDC activity compared to an otherwise equivalent antibody comprising an Fc not having the one or more substitutions.

209. The isolated antibody of claim 208, wherein the change is an increase or decrease in antibody half-life, an increase or decrease in ADCC activity, an increase or decrease in ADCP activity, or an increase or decrease in CDC activity compared to an otherwise equivalent antibody comprising an Fc not having the one or more substitutions, Optionally wherein the change is: Increased antibody half-life; reduction in antibody half-life; Increased ADCC activity; Reduction of ADCC activity; Increased ADCP activity; Decreased ADCP activity; Increased CDC activity; Decreased CDC activity; Increased antibody half-life, increased ADCC activity, increased ADCP activity, and increased CDC activity; and / or Increased antibody half-life, decreased ADCC activity, increased ADCP activity and increased CDC activity.

210. The isolated antibody of claim 208, wherein the one or more amino acid substitutions result in an increase in antibody half-life compared to an antibody comprising a wild-type Fc region.

211. The isolated antibody of any of the above claims, wherein the Fc region binds to the neonatal Fc receptor (FcRn).

212. The isolated antibody of claim 211, wherein the Fc region binds FcRn with a higher affinity at pH 6.0 than an antibody comprising a wild-type Fc region.

213. The isolated antibody of claim 211 or 212, wherein the Fc region is at a pH of <1 x 10 -7 M of K D Binds to FcRn.

214. The isolated antibody of any of the above claims, wherein the antibody is a monoclonal antibody.

215. The antibody of any of the above claims, wherein the antibody binds to an IL-13 sequence selected from the group consisting of the sequences shown in SEQ ID NOs: 472-475.

216. An isolated antibody as described in any of the above claims, wherein the antibody has a density of less than or equal to about 1, 2, 3, 4, 5, 6, 7, 8, 9 x 10 -9 M of K D Binds to an IL-13 sequence selected from the group consisting of sequences shown in SEQ ID NOs: 472-475.

217. The isolated antibody of claim 216, wherein the antibody has a concentration of less than or equal to about 1 x 10 -10 M of K D Binds to an IL-13 sequence selected from the group consisting of sequences shown in SEQ ID NOs: 472-475.

218. The isolated antibody of claim 216, wherein the antibody has a concentration of less than or equal to about 1 x 10 -9 M of K D Binds to human IL-13.

219. The isolated antibody of any of the above claims, wherein the antibody exhibits a melting temperature greater than 68°C as measured by differential scanning fluorimetry (DSF).

220. The isolated antibody of claim 219, wherein the antibody exhibits a melting temperature greater than 75°C as measured by DSF.

221. The isolated antibody of any of the above claims, wherein the antibody exhibits an aggregation temperature equal to or greater than 71.2°C as measured by DSF.

222. The isolated antibody of any of the above claims, wherein the antibody has a retention time of 15.2 minutes or less as measured by hydrophobic interaction chromatography.

223. An isolated antibody as described in any of the above claims, wherein the antibody does not have the heavy chain variable region sequence shown in SEQ ID NO:

470.

224. The isolated antibody of any of the above claims for use in treating an inflammatory disorder or disease.

225. The isolated antibody of claim 224, for use in treating atopic dermatitis.

226. The isolated antibody of claim 225, wherein the treatment reduces disease severity in the subject, and wherein disease severity is assessed by an atopic dermatitis disease severity outcome measure.

227. The isolated antibody of claim 224, for use in treating asthma.

228. The isolated antibody of claim 224, for use in treating idiopathic pulmonary fibrosis.

229. The isolated antibody of claim 224, for use in treating alopecia areata.

230. The isolated antibody of claim 224, for use in treating chronic sinusitis with nasal polyps.

231. The isolated antibody of claim 224, for use in treating chronic rhinosinusitis without nasal polyps (CRSsNP).

232. The isolated antibody of claim 224, for use in treating eosinophilic esophagitis (EoE).

233. The isolated antibody of claim 224, for use in treating an eosinophilic gastrointestinal disorder or disease (ENID) selected from the group consisting of eosinophilic gastritis (EoG), eosinophilic enteritis (EoN), eosinophilic colitis (EoC), and eosinophilic gastroenteritis (EGE).

234. The isolated antibody of claim 224, for use in treating Chargar-Strauss syndrome / eosinophilic granulomatosis with polyangiitis (EGPA).

235. The isolated antibody of claim 224, for use in treating prurigo nodularis (PN).

236. The isolated antibody of claim 224, for use in treating chronic spontaneous urticaria (CSU).

237. The isolated antibody of claim 224, for use in treating chronic pruritus of unknown origin (CPUO).

238. The isolated antibody of claim 224, for use in treating bullous pemphigoid (BP).

239. The isolated antibody of claim 224, for use in treating cold-induced urticaria (ColdU).

240. The isolated antibody of claim 224, for use in treating allergic fungal sinusitis (AFRS).

241. The isolated antibody of claim 224, for use in treating allergic bronchopulmonary aspergillosis (ABPA).

242. The isolated antibody of claim 224, for use in treating chronic obstructive pulmonary disease (COPD).

243. An isolated polynucleotide or group of polynucleotides encoding the antibody, VH, VL, light chain, heavy chain or antigen binding portion thereof of any of the above claims, and optionally wherein the polynucleotide or group of polynucleotides comprises cDNA.

244. A vector or a vector set, comprising the polynucleotide or polynucleotide set of claim 243.

245. A host cell comprising the polynucleotide or polynucleotide set of claim 219 or the vector or vector set of claim 244.

246. A method for producing an antibody, the method comprising expressing the antibody using the host cell of claim 245 and isolating the expressed antibody.

247. A pharmaceutical composition comprising the antibody of any one of claims 1-223 and a pharmaceutically acceptable excipient.

248. A drug kit comprising the antibody of any one of claims 1-242 or the pharmaceutical composition of claim 247 and instructions for use.

249. A method for treating an inflammatory condition or disease in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of the antibody of any one of claims 1-223 or the pharmaceutical composition of claim 247.

250. The method of claim 249, wherein the inflammatory disorder or disease is atopic dermatitis.

251. The method of claim 249, wherein the inflammatory disorder or disease is asthma.

252. The method of claim 249, wherein the inflammatory disorder or disease is idiopathic pulmonary fibrosis.

253. The method of claim 249, wherein the inflammatory condition or disease is alopecia areata.

254. The method of claim 249, wherein the inflammatory condition or disease is chronic sinusitis with nasal polyps.

255. The method of claim 249, wherein the inflammatory disorder or disease is chronic rhinosinusitis without nasal polyps (CRSsNP).

256. The method of claim 249, wherein the inflammatory disorder or disease is eosinophilic esophagitis (EoE).

257. The method of claim 249, wherein the inflammatory disorder or disease is an eosinophilic gastrointestinal disorder or disease (ENID) selected from the group consisting of eosinophilic gastritis (EoG), eosinophilic enteritis (EoN), eosinophilic colitis (EoC), and eosinophilic gastroenteritis (EGE).

258. The method of claim 249, wherein the inflammatory disorder or disease is Chargar-Strauss syndrome / eosinophilic granulomatosis with polyangiitis (EGPA).

259. The method of claim 249, wherein the inflammatory disorder or disease is prurigo nodularis (PN).

260. The method of claim 249, wherein the inflammatory disorder or disease is chronic spontaneous urticaria (CSU).

261. The method of claim 249, wherein the inflammatory disorder or disease is chronic pruritus of unknown origin (CPUO).

262. The method of claim 249, wherein the inflammatory disorder or disease is bullous pemphigoid (BP).

263. The method of claim 249, wherein the inflammatory disorder or disease is cold-induced urticaria (ColdU).

264. The method of claim 249, wherein the inflammatory disorder or disease is allergic fungal sinusitis (AFRS).

265. The method of claim 249, wherein the inflammatory disorder or disease is allergic bronchopulmonary aspergillosis (ABPA).

266. The method of claim 249, wherein the inflammatory disorder or disease is chronic obstructive pulmonary disease (COPD).

267. A method for treating a pathology associated with elevated IL-13 levels in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of the antibody of any one of claims 1-242 or the pharmaceutical composition of claim 247.

268. A method of reducing IL-13 bioactivity in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of the antibody of any one of claims 1-242 or the pharmaceutical composition of claim 247.

269. A method of inhibiting a TH2 type allergic response in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of the antibody of any one of claims 1-242 or the pharmaceutical composition of claim 247.

270. A method of reducing thymus and activation-regulated chemokine (TARC) / CCL17 levels in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of the antibody of any one of claims 1-242 or the pharmaceutical composition of claim 247.

271. A method of preventing an inflammatory condition or disease in a mammalian subject in need thereof, the method comprising administering to the mammalian subject a therapeutically effective amount of the antibody of any one of claims 1-242 or the pharmaceutical composition of claim 247.

Images