Antibodies targeting TIM-3 and uses thereof

Abstract

Disclosed herein are anti-TIM-3 antibodies and antigen-binding fragments, polynucleotides encoding the antibodies and antigen-binding fragments, and pharmaceutical compositions comprising the antibodies and antigen-binding fragments. Also disclosed are uses of the anti-TIM-3 antibodies and antigen-binding fragments described herein in treating cancer.

Description

[Technical Field]

[0001] This application claims priority to PCT patent application No. PCT / CN2021 / 089261, filed April 23, 2021, and PCT patent application No. PCT / CN2021 / 120140, filed September 24, 2021, each of which is incorporated herein by reference in its entirety.

[0002] (1. Field) The present invention relates to molecular biology and immuno-oncology. Provided herein are anti-TIM-3 antibodies and their use in treating tumors or cancer. [Background technology]

[0003] (2.Background) Immune checkpoint inhibitors (e.g., those targeting PD-1, TIM-3, and CTLA-4) have become a promising class of molecules for therapeutic development. Despite the success of checkpoint inhibitors such as Yervoy®, Keytruda®, and Opdivo®, only a small fraction of patients experience a durable clinical response to these therapies. Some tumor types have shown little response to anti-CTLA-4 or anti-PD-1 / PD-L1 monotherapy in clinical trials. Although TIM-3 expression has been associated with cancer, there has been limited success in developing therapeutic options that target TIM-3. Thus, there is an unmet need for additional treatment options for cancer patients, particularly for TIM-3-targeting agents. The compositions and methods provided herein fulfill these needs and offer other related advantages. Summary of the Invention

[0004] (3. Overview) Provided herein is an antibody or antigen-binding fragment thereof that specifically binds to human TIM-3, comprising: (a) a light chain variable region (VL) comprising (1) a light chain CDR1 (VL CDR1) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 86-93 and 129-137; (2) a light chain CDR2 (VL CDR2) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 94-100 and 138-144; and (3) a light chain CDR3 (VL CDR3) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47-55, 145-153, and 198-206; or a variant thereof having up to about three amino acid substitutions, additions, and / or deletions in the VL CDR; and / or (b) (1) a heavy chain CDR1 (VH CDR1) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 101-108 and 154-161. (2) a heavy chain CDR2 (VH CDR2) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 109 to 118 and 162 to 170; and (3) a heavy chain CDR3 (VH CDR3) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 119 to 128 and 171 to 179; or a heavy chain variable region (VH) comprising these variants having up to about three amino acid substitutions, additions, and / or deletions in the VH CDR.

[0005] In some embodiments of the anti-TIM-3 antibodies or antigen-binding fragments provided herein, (a) VL CDR1, CDR2, and CDR3 are (1) the amino acid sequences of SEQ ID NOs: 86, 94, and 47, respectively; (2) the amino acid sequences of SEQ ID NOs: 87, 95, and 48, respectively; (3) the amino acid sequences of SEQ ID NOs: 88, 96, and 49, respectively; (4) the amino acid sequences of SEQ ID NOs: 89, 97, and 50, respectively; (5) the amino acid sequences of SEQ ID NOs: 90, 94, and 51, respectively; (6) the amino acid sequences of SEQ ID NOs: 91, 98, and 52, respectively; (7) the amino acid sequences of SEQ ID NOs: 91, 98, and 53, respectively. (8) the amino acid sequences of SEQ ID NOs: 92, 99, and 54, respectively; (9) the amino acid sequences of SEQ ID NOs: 93, 100, and 55, respectively; (10) the amino acid sequences of SEQ ID NOs: 129, 138, and 145, respectively; (11) the amino acid sequences of SEQ ID NOs: 130, 139, and 146, respectively; (12) the amino acid sequences of SEQ ID NOs: 131, 140, and 147, respectively; (13) the amino acid sequences of SEQ ID NOs: 132, 141, and 148, respectively; (14) the amino acid sequences of SEQ ID NOs: 133, 139, and 146, respectively. and 149; (15) the amino acid sequences of SEQ ID NOs: 134, 142, and 150, respectively; (16) the amino acid sequences of SEQ ID NOs: 135, 143, and 151, respectively; (17) the amino acid sequences of SEQ ID NOs: 136, 144, and 152, respectively; (18) the amino acid sequences of SEQ ID NOs: 137, 100, and 153, respectively; (19) the amino acid sequences of SEQ ID NOs: 86, 94, and 198, respectively; (20) the amino acid sequences of SEQ ID NOs: 86, 94, and 199, respectively; (21) the amino acid sequences of SEQ ID NOs: 137, 100, and 153, respectively. (22) the amino acid sequences of SEQ ID NOs: 86, 94, and 201, respectively; (23) the amino acid sequences of SEQ ID NOs: 86, 94, and 202, respectively; (24) the amino acid sequences of SEQ ID NOs: 86, 94, and 203, respectively; (25) the amino acid sequences of SEQ ID NOs: 86, 94, and 204, respectively; (26) the amino acid sequences of SEQ ID NOs: 86, 94, and 205, respectively; or (27) the amino acid sequences of SEQ ID NOs: 86, 94, and 206, respectively; or a variant thereof having up to about three amino acid substitutions, additions, and / or deletions in the VL CDR;and / or (b) VH CDR1, CDR2, and CDR3 are (1) the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (2) the amino acid sequences of SEQ ID NOs: 102, 110, and 120, respectively; (3) the amino acid sequences of SEQ ID NOs: 103, 111, and 121, respectively; (4) the amino acid sequences of SEQ ID NOs: 104, 112, and 122, respectively; (5) the amino acid sequences of SEQ ID NOs: 105, 113, and 123, respectively; (6) the amino acid sequences of SEQ ID NOs: 106, 114, and 124, respectively; (7) the amino acid sequences of SEQ ID NOs: 106, 115, and 125, respectively; (8) the amino acid sequences of SEQ ID NOs: 107, 116, and 126, respectively; (9) the amino acid sequences of SEQ ID NOs: 108, 117, and 127, respectively; (10) the amino acid sequences of SEQ ID NOs: 106, 118, and 129, respectively; (11) the amino acid sequences of SEQ ID NOs: 106, 162, and 171, respectively; (12) the amino acid sequences of SEQ ID NOs: 154, 163, and 172, respectively; (13) the amino acid sequences of SEQ ID NOs: 155, 164, and 173, respectively; (14) the amino acid sequences of SEQ ID NOs: 156, 165, and 174, respectively; (15) the amino acid sequences of SEQ ID NOs: 157, 166, and 175, respectively; (16) the amino acid sequences of SEQ ID NOs: 158, 167, and 176, respectively; (17) the amino acid sequences of SEQ ID NOs: 159, 168, and 177, respectively; (18) the amino acid sequences of SEQ ID NOs: 160, 169, and 178, respectively; or (19) the amino acid sequences of SEQ ID NOs: 161, 170, and 179, respectively; or the VH These variants have up to about three amino acid substitutions, additions, and / or deletions in the CDRs;

[0006] In some embodiments of the anti-TIM-3 antibodies or antigen-binding fragments provided herein, (1) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 47, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (2) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 87, 95, and 48, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 102, 110, and 120, respectively; (3) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 88, 96, and 49, respectively; and / or VH (4) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 89, 97, and 50, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 104, 112, and 122, respectively; (5) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 90, 94, and 51, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 105, 113, and 123, respectively; (6) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 91, 98, and 52, respectively; and / or VH (7) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 91, 98, and 53, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 106, 115, and 125, respectively; (8) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 92, 99, and 54, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 107, 116, and 126, respectively;(9) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 93, 100, and 55, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 108, 117, and 127, respectively; (10) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 91, 98, and 52, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 106, 118, and 128, respectively; (11) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 129, 138, and 145, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 106, 162, and 171, respectively; (12) VL (13) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 131, 140, and 147, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 155, 164, and 173, respectively; (14) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 132, 141, and 148, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 156, 165, and 174, respectively; (15) VL (16) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 134, 142, and 150, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 158, 167, and 176, respectively;(17) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 135, 143, and 151, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 159, 168, and 177, respectively; (18) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 136, 144, and 152, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 160, 169, and 178, respectively; (19) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 137, 100, and 153, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 161, 170, and 179, respectively; (20) VL (21) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 198, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (22) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 200, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (23) VL (23) CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 201, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (24) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 202, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively;(25) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 203, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (26) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 204, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (27) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 205, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; or (28) VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 206, respectively; and / or VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively;

[0007] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise VL CDR1, VL CDR2, VH CDR1, VH CDR2, and VH CDR3 having the amino acid sequences of SEQ ID NOs: 86, 94, 101, 109, and 119, respectively, and a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47 and 198-206; or variants thereof having up to about three amino acid substitutions, additions, and / or deletions in the VL CDRs and up to about three amino acid substitutions, additions, and / or deletions in the VH CDRs. In some embodiments, the F residue of VH CDR1 (amino acid 3 of SEQ ID NO: 101), the H and S residues of VH CDR2 (amino acids 4 and 5 of SEQ ID NO: 109), the Y, R, S, and W residues of VH CDR3 (amino acids 2, 3, 4, and 6 of SEQ ID NO: 119), and the S residue of VL CDR2 (amino acid 7 of SEQ ID NO: 94) are not mutated. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 having the amino acid sequences of SEQ ID NOs: 86, 94, 47, 101, 109, and 119, respectively.

[0008] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to human TIM-3, comprising: (a) a VL having at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-10 and 180-188; and / or (b) a VH having at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 11-20 and 189-197.

[0009] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise a VL and a VH, wherein the VL and VH are each selected from the group consisting of: (1) SEQ ID NOs: 1 and 11, respectively; (2) SEQ ID NOs: 2 and 12, respectively; (3) SEQ ID NOs: 3 and 13, respectively; (4) SEQ ID NOs: 4 and 14, respectively; (5) SEQ ID NOs: 5 and 15, respectively; (6) SEQ ID NOs: 6 and 16, respectively; (7) SEQ ID NOs: 7 and 17, respectively; (8) SEQ ID NOs: 8 and 18, respectively; (9) SEQ ID NOs: 9 and 19, respectively; (10) SEQ ID NOs: 10 and 20, respectively; (11) and (12) SEQ ID NOs: 181 and 190, respectively; (13) SEQ ID NOs: 182 and 191, respectively; (14) SEQ ID NOs: 183 and 192, respectively; (15) SEQ ID NOs: 184 and 193, respectively; (16) SEQ ID NOs: 185 and 194, respectively; (17) SEQ ID NOs: 186 and 195, respectively; (18) SEQ ID NOs: 187 and 196, respectively; or (19) SEQ ID NOs: 188 and 197, respectively. In some embodiments, the VL and VH each have at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity to the amino acid sequence of SEQ ID NOs: 1 and 11, respectively.

[0010] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to human TIM-3, comprising: (a) a VL having at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity to SEQ ID NOs: 21 and 207-215; and / or (b) a VH having at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 22-29.

[0011] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to human TIM-3, comprising: (a) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-10 and 180-188; and / or (b) a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 11-20 and 189-197.

[0012] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are selected from the group consisting of: (1) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 1, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 11; (2) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 2, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 12; (3) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 3, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 13; (4) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 4, and / or a VH derived from a VH having the amino acid sequence of SEQ ID NO: 14. (5) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 5, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 15; (6) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 6, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 16; (7) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 7, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 17; (8) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 8, and / or a VH derived from a VH having the amino acid sequence of SEQ ID NO: 18. (9) a VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 9, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 19;(10) VL comprising VL CDR1, CDR2, and CDR3 derived from VL having the amino acid sequence of SEQ ID NO: 10, and / or VH comprising VH CDR1, CDR2, and CDR3 derived from VH having the amino acid sequence of SEQ ID NO: 20; (11) VL comprising VL CDR1, CDR2, and CDR3 derived from VL having the amino acid sequence of SEQ ID NO: 180, and / or VH comprising VH CDR1, CDR2, and CDR3 derived from VH having the amino acid sequence of SEQ ID NO: 189; (12) VL comprising VL CDR1, CDR2, and CDR3 derived from VL having the amino acid sequence of SEQ ID NO: 181, and / or VH comprising VH CDR1, CDR2, and CDR3 derived from VH having the amino acid sequence of SEQ ID NO: 190; (13) VL comprising VL CDR1, CDR2, and CDR3 derived from VL having the amino acid sequence of SEQ ID NO: 182, and / or VH derived from VH having the amino acid sequence of SEQ ID NO: 191. (14) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 183, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 192; (15) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 184, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 193; (16) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 185, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 194; (17) a VL derived from a VL having the amino acid sequence of SEQ ID NO: 186. (18) a VL comprising CDR1, CDR2, and CDR3, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 195; (19) a VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 187, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 196;or (19) a VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 188, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 197;

[0013] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to human TIM-3, comprising a VL and a VH, wherein the VL comprises VL CDR1, CDR2, and CDR3 derived from a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 21, and 207-215, and the VH comprises VH CDR1, CDR2, and CDR3 derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 11 and 22-29.

[0014] In some embodiments, provided herein are antibodies or antigen-binding fragments thereof that compete with any of the anti-TIM-3 antibodies or antigen-binding fragments disclosed herein for binding to human TIM-3.

[0015] In some embodiments, provided herein are antibodies or antigen-binding fragments thereof that specifically bind to human TIM-3, wherein the antibodies or antigen-binding fragments specifically bind to an epitope comprising at least one of amino acids 71-82 of human TIM-3. In some embodiments, the antibodies or antigen-binding fragments disclosed herein specifically bind to at least one of the following amino acid residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the antibodies or antigen-binding fragments disclosed herein specifically bind to at least two, at least three, at least four, at least five, at least six, at least seven, or eight of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the antibodies or antigen-binding fragments disclosed herein specifically bind to at least D71, N76, or Y82 of human TIM-3. In some embodiments, the antibodies or antigen-binding fragments disclosed herein do not specifically bind to an epitope outside of amino acids 71-82 of human TIM-3.

[0016] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are administered in a concentration of 5×10 -8 In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein bind to human TIM-3 with a KD that is 10 M or less. -11 M~5×10 -9 It binds to human TIM-3 with a KD that spans the range of M.

[0017] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein block the interaction between TIM-3 and a TIM-3 ligand. In some embodiments, the TIM-3 ligand is phosphatidylserine, CEACAM1, HMGB1, or any combination thereof.

[0018] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein (1) inhibit TIM-3-mediated T cell suppression, (2) inhibit TIM-3-mediated myeloid cell suppression, (3) inhibit TIM-3-mediated suppression of inflammasome activation, or any combination thereof.

[0019] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are monoclonal antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are selected from the group consisting of IgG1 antibodies, IgG2 antibodies, IgG3 antibodies, and IgG4 antibodies. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are IgG1 antibodies. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are selected from the group consisting of Fab, Fab', F(ab')2, Fv, scFv, (scFv)2, single domain antibodies (sdAbs), and heavy chain antibodies (HCAbs).

[0020] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are chimeric antibodies or antigen-binding fragments, humanized antibodies or antigen-binding fragments, or human antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are humanized antibodies or antigen-binding fragments.

[0021] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are bispecific or multispecific antibodies, ie, bispecific antibodies that additionally specifically bind to PD-1, PD-L1, CEACAM1, or CEACAM5.

[0022] In some embodiments, provided herein are polynucleotides encoding the anti-TIM-3 antibodies or antigen-binding fragments provided herein. In some embodiments, provided herein are vectors comprising the polynucleotides provided herein. In some embodiments, provided herein are host cells comprising the polynucleotides provided herein or the vectors provided herein.

[0023] In some embodiments, provided herein is a pharmaceutical composition comprising a therapeutically effective amount of an anti-TIM-3 antibody or antigen-binding fragment provided herein and a pharmaceutically acceptable carrier.

[0024] In some embodiments, provided herein are methods for inducing or stimulating immune cell activation and / or proliferation, comprising contacting an immune cell with an effective amount of an anti-TIM-3 antibody or antigen-binding fragment provided herein. In some embodiments, provided herein are methods for reducing TIM-3-mediated suppression of an immune cell, comprising contacting an immune cell with an effective amount of an anti-TIM-3 antibody or antigen-binding fragment provided herein. In some embodiments, the immune cell is a T cell, an NK cell, an NKT cell, or a myeloid cell. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is an NK cell. In some embodiments, the immune cell is a myeloid cell, wherein the myeloid cell is a macrophage or a dendritic cell.

[0025] In some embodiments, provided herein are methods of stimulating anti-tumor immunity in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of an anti-TIM-3 antibody or antigen-binding fragment provided herein, or a pharmaceutical composition provided herein. In some embodiments, provided herein are methods of inhibiting tumor cell growth in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of an anti-TIM-3 antibody or antigen-binding fragment provided herein, or a pharmaceutical composition provided herein.

[0026] In some embodiments, provided herein is a method of treating cancer in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of an anti-TIM-3 antibody or antigen-binding fragment provided herein, or a pharmaceutical composition provided herein.

[0027] In some embodiments, the methods provided herein further comprise administering to the subject an additional therapy. In some embodiments, the additional therapy comprises an antibody that specifically binds to PD-L1, PD-1, CEACAM1, CTLA4, CEACAM5, latent TGF-β, a TGF-β receptor, CD70, B7H4, or B7H3. In some embodiments, the additional therapy comprises radiation or chemotherapy.

[0028] In some embodiments of the methods provided herein, the subject is a human.

[0029] In some embodiments, provided herein is the use of an anti-TIM-3 antibody or antigen-binding fragment provided herein in the treatment of cancer. In some embodiments, provided herein is the use of an anti-TIM-3 antibody or antigen-binding fragment for the preparation of a medicament for the treatment of cancer.

[0030] In some embodiments, the cancer is a blood cancer. In some embodiments, the blood cancer is acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS). In some embodiments, the cancer is a solid tumor. In some embodiments, the cancer is melanoma, lung cancer, head and neck cancer, colorectal cancer, pancreatic cancer, gastric cancer, kidney cancer, bladder cancer, prostate cancer, breast cancer, ovarian cancer, uterine / cervical cancer, testicular cancer, thyroid cancer, esophageal cancer, soft tissue sarcoma, liver cancer, gallbladder cancer, cervical cancer, duodenal cancer, bone cancer, neuroendocrine cancer, intestinal cancer, skin cancer, or germ cell cancer. In some embodiments, the cancer is selected from the group consisting of renal cell carcinoma (RCC), non-small cell lung cancer (NSCLC), squamous cell carcinoma of the head and neck (SCCHN), triple-negative breast cancer (TNBC), gastric / gastric adenocarcinoma (STAD), pancreatic adenocarcinoma (PAAD), colon adenocarcinoma (COAD), or rectal adenocarcinoma (READ). In some embodiments, the cancer has a high level of microsatellite instability. In some embodiments, the cancer is a metastatic cancer, a refractory cancer, or a recurrent cancer. [Brief explanation of the drawings]

[0031] (4. Brief description of the drawings) [Figure 1] Figure 1A provides flow cytometry data showing cross-reactivity of various anti-TIM-3 antibodies with cynomolgus monkey TIM-3, and Figure 1B provides flow cytometry data showing that none of the tested anti-TIM-3 antibodies cross-reacted with mouse TIM-3.

[0032] [Figure 2] FIG. 2 provides flow cytometry data showing that several humanized anti-TIM-3 antibodies did not cross-react with cynomolgus monkey TIM-3.

[0033] [Figure 3] FIG. 3 provides flow cytometry data showing that an anti-TIM-3 antibody blocked TIM-3 binding to phosphatidylserine on apoptotic cells.

[0034] [Figure 4] FIG. 4 provides flow cytometry data showing that certain anti-TIM-3 antibodies induced internalization of TIM-3.

[0035] [Figure 5] Figure 5A provides ELISA data showing that anti-TIM-3 antibodies dose-dependently increased IFN-γ secretion in cocultures of 293T / OS8 target and effector cells, and Figure 5B provides the results of the same assay for the affinity-matured 3E6 antibody.

[0036] [Figure 6] FIG. 6 provides flow cytometry data showing that an anti-TIM-3 antibody increased CD107a expression on NK cells in a dose-dependent manner.

[0037] [Figure 7] Figures 7A-7B provide data from the biomolecular interaction system Gator (Probe Life) measuring binding between the humanized anti-TIM-3 antibody 3E6 of various isotypes (IgG1, IgG1 LALA, IgG2, and IgG4) and various Fc receptors. Figure 7A (human Fc receptor): CD32a H167, CD32a R167, CD32b, Cd16a 176F, Cd16a 176V, and FCRN; Figure 7B (mouse Fc receptor): CD16, CD32B, FCRN, or FCGR4. IgG1 LALA: IgG1 with L234A and L235A mutations.

[0038] [Figure 8] Figure 8 provides results from a BxPC-3 / hCD34+ humanized mouse model showing that humanized anti-TIM-3 antibody 3E6 of various isotypes (IgG1 LALA, IgG2, and IgG4) effectively inhibited tumor growth.

[0039] [Figure 9]Figure 9 provides the levels of cytokines (LEGENDplex™ Human Essential Immune Response Panel) detected in endpoint serum samples in the BxPC-3 / hCD34+ humanized mouse model, showing that the humanized anti-TIM-3 antibody (h3E6 IgG1 LALA) enhanced serum levels of several cytokines in the mouse model.

[0040] [Figure 10] Figure 10 provides results from the MC38 / TIM3-humanized C57BL / 6 mouse model showing that humanized anti-TIM-3 antibodies of various isotypes (IgG1, IgG1 LALA, IgG2, and IgG4) effectively inhibited tumor growth.

[0041] [Figure 11] Figure 11A provides the crystal structure of humanized 3E6 Fab bound to human TIM-3 (left: hTIM-3; right: humanized 3E6 Fab (top: VH; bottom: VL)). Figure 11B shows the interactions between specific residues of the TIM-3 antigen and specific residues of the VH and VL CDRs.

[0042] [Figure 12] Figures 12A-12B provide the crystal structure of the complex formed by human TIM-3 simultaneously binding humanized 3E6 Fab and another anti-TIM-3 antibody (Figure 12A: 6TXZ; Figure 12B: 7KQL). DETAILED DESCRIPTION OF THE INVENTION

[0043] (5. Detailed Description) Before the present disclosure is further described, it is to be understood that the present disclosure is not limited to particular embodiments described herein, and that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.

[0044] The present disclosure provides novel antibodies, including antigen-binding fragments, that specifically bind to TIM-3 (e.g., human TIM-3). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments disclosed herein bind to a novel epitope on human TIM-3 and can inhibit tumor growth by reducing or inhibiting TIM-3-mediated suppression of both innate and adaptive immune responses. Pharmaceutical compositions comprising a therapeutically effective amount of such antibodies or antigen-binding fragments are also disclosed herein. Also disclosed herein are uses of such pharmaceutical compositions for treating cancer (e.g., cancers that express TIM-3) and methods of cancer treatment.

[0045] T-cell immunoglobulin and mucin domain-3 (TIM-3), also known as hepatitis A virus cellular receptor 2 (HAVCR2), is an immune checkpoint protein. First identified as a molecule selectively expressed on IFN-γ-producing CD4+ T helper 1 (Th1) and CD8+ T cytotoxic 1 (Te1) T cells (Monney et al., 2002, Nature, 415(6871):536-41), TIM-3 is a cytotoxic protein that inhibits the immune response to FOXP3. + CD4 + It is also expressed on the surface of many immune cell types, including certain subsets of T cells such as T regulatory cells (Tregs), natural killer (NK) cells, NKT cells, monocytes, and tumor-associated dendritic cells (TADCs) (Clayton et al. (2014) J. Immunol., 192(2):782-791; Anderson et al. (2007) Science 318(5853): 1141- 1143; Baitsch et al. (2012) Plos One 7(2):e30852; Ndhlovu et al. (2012) Blood 119(16):3734-3743).

[0046] T-cell immunoglobulin and mucin domain-containing-3 (TIM-3), also known as hepatitis A virus cellular receptor 2 (HAVCR2), TIM-3, TIMD3, TIMD-3, kidney injury molecule-3, KIM-3, and CD366, is a type I transmembrane protein that functions as a key regulator of the immune response. TIM-3 initially targets activated IFN-γ-producing T cells (e.g., type 1 helper CD4 + T cells and cytotoxic CD8 + TIM-3 was identified on T cells and shown to induce T cell death or exhaustion after binding to galectin-9 (Monney et al., 2002). More recent studies have shown that TIM-3 expression is also important in regulating the activity of many innate immune cells (e.g., macrophages, monocytes, dendritic cells, mast cells, and natural killer cells) (Han et al., 2013).

[0047] Reported ligands for TIM-3 (TIM-3L) include phosphatidylserine ("PtdSer"; Nakayama et al. (2009) Blood 113(16):3821-30), galectin-9 (Gal-9) (Zhu et al. (2005) Nat Immunol 6(12):1245-52), high mobility group protein 1 (HMGB1) (Chiba et al. (2012) Nat Immunol 13(9):832-42), carcinoembryonic antigen cell adhesion molecule 1 (CEACAM1) (Huang et al. (2015) Nature 517(7534):386-90), semaphorin-4A, and ILT-4. PtdSer is an important plasma membrane component and is normally localized to the inner leaflet of the plasma membrane. However, when cells undergo apoptosis, PtdSer is redistributed to the outer membrane and exposed. This redistribution is also observed in many tumor cell lines. The binding of TIM-3 to PtdSer may be crucial for phagocytosis and cross-presentation (Nakayama 2009, supra).

[0048] TIM-3 regulates various aspects of the immune response. The interaction of TIM-3 with its ligand, galectin-9 (Gal-9), induces cell death. Blockade of this interaction in vivo exacerbates autoimmunity and abolishes tolerance in experimental models, indicating that the TIM-3 / Gal-9 interaction negatively regulates immune responses (Zhu et al. (2005) supra; Kanzaki et al. (2012) Endocrinology 153(2):612-620). Inhibition of TIM-3 also enhanced the severity of pathology in experimental autoimmune encephalomyelitis in vivo (Monney et al. (2002) Nature 415:536-541; Das et al. (2017) Immunol Rev 276(1):97-11). In studies using material from human patients with multiple sclerosis (Koguchi et al. (2006) J Exp Med 203(6): 1413-1418), Crohn's disease (CD) (Morimoto et al. (2011) Scand J Gastroenterol 46(6):701-709), and rheumatoid arthritis (RA) (Liu et al. (2010) Clin Immunol 137(2):288-295; Li et al. (2014) PLoS ONE 9(2):e85784), the observation that TIM-3 expression levels on T cells inversely correlated with the progression of autoimmune disease indicates an immunosuppressive role for TIM-3 on T cells.

[0049] TIM-3 is considered a promising candidate for cancer immunotherapy, in part because it is upexpressed on tumor-infiltrating lymphocytes, including Foxp3+CD4+ Tregs and exhausted CD8+ T cells, two important immune cell populations that constitute immunosuppressive components in the tumor environment of many human cancers (McMahan et al. (2010) J. Clin. Invest. 120(12):4546-4557; Jin et al. (2010) Proc Natl Acad Sci USA 107(33): 14733-8; Zhou et al. (2011) Blood 117(17):4501-4510; Yan et al. (2013) PLoS ONE 8(3):e58006). It has been reported that interaction of TIM-3 on CD8+ T cells with its ligand, galectin-9 on tumor cells, results in phosphorylation of the TIM-3 cytoplasmic tail at tyrosines 256 and 263, resulting in the release of HLA-B-associated transcript 3 (Bat3) and catalytically active lymphocyte-specific protein tyrosine kinase (Lck) from the TIM-3 cytoplasmic tail. Dissociation of Bat3 and Lck from TIM-3 leads to the accumulation of inactive phosphorylated Lck, which may be a major cause of the observed T cell dysfunction (Rangachari et al. (2012) Nat. Med. 18(9): 1394-400).

[0050] Furthermore, intratumoral TIM-3 + FoxP3 +Treg cells have been reported to express large amounts of Treg effector molecules (IL-10, perforin, and granzymes). TIM-3+ Tregs have been reported to promote the development of a dysfunctional phenotype of CD8+ tumor-infiltrating lymphocytes (TILs) in the tumor environment (Sakuishi et al. (2013) Oncoimmunology 2(4):e23849). TIM-3 has also been reported to exert effects on the myeloid compartment. T cell expression of TIM-3 has been shown to promote CD1lb+Gr-1+ myeloid-derived suppressor cells (MDSCs) in a galectin-9-dependent manner (Dardalhon et al. (2010) J Immunol 185(3):1383-92). Furthermore, TIM-3 is specifically upregulated in tumor-associated dendritic cells (TADCs) and can interfere with the sensing of DNA released by cells undergoing necrotic cell death. TIM-3 binds to high mobility group protein 1 (HMGB1), thereby preventing HMGB1 from binding to DNA released from dying cells and mediating its delivery to innate cells via the receptor for advanced glycation end products (RAGE) and / or Toll-like receptor (TLR) 2 and 4 pathways. Binding of TIM-3 to HMGB1 suppresses the activation of innate immune responses in tumor tissues (Chiba et al., 2012, supra). TIM-3 potentially also acts as a gatekeeper of inflammation and suppresses antitumor immunity by regulating inflammasome activation (Gayden et al., Nature Genetics 50.12(2018): 1650-1657; Dixon et al., Nature 595.7865(2021): 101-106). Taken together, these observations indicate that TIM-3 can further suppress antitumor T cell responses through T cell-extrinsic mechanisms involving myeloid cells and various TIM-3 / ligand interactions.

[0051] Like many inhibitory receptors (e.g., PD-1 and CTLA-4), expression of TIM-3 is associated with many types of chronic diseases, including cancer. +T cells have been detected in patients with advanced melanoma, non-small cell lung cancer, and follicular B-cell non-Hodgkin's lymphoma. + The presence of regulatory T cells has been described as a useful indicator of lung cancer progression (Anderson, 2014, Cancer Immunol. Res. 2, 393-98). Studies have shown a close relationship between TIM-3 and the inhibitory receptor PD-1. For example, many tumor-specific T cells express both PD-1 and TIM-3, and these T cells have been shown to be more dysfunctional than T cells that express only PD-1 or TIM-3 (Fourcade et al., 2010, J. Exp. Med. 207, 2175-2186).

[0052] The term "TIM-3" includes any variant or isoform of TIM-3 that is naturally expressed by a cell. Some antibodies described herein can cross-react with TIM-3 from certain species other than human (e.g., cynomolgus monkey TIM-3), but not with that of certain other species, such as mouse TIM-3. Some antibodies described herein (e.g., humanized 3E6) do not cross-react with TIM-3 from species other than human, including cynomolgus monkey TIM-3 and mouse TIM-3. TIM-3 or any variants and isoforms thereof can be isolated from cells or tissues that naturally express them, or can be recombinantly produced using techniques well known in the art and / or described herein.

[0053] Two isoforms of human TIM-3 have been identified. Isoform 1 (NCBI Reference Sequence: NP_116171.3; SEQ ID NO: 84) consists of 301 amino acids and represents the canonical sequence. The extracellular region of human TIM-3 comprises amino acid residues 22-202 of SEQ ID NO: 84. The transmembrane domain of human TIM-3 comprises amino acid residues 203-223 of SEQ ID NO: 84. The cytoplasmic domain of human TIM-3 comprises amino acid residues 224-301 of SEQ ID NO: 84. [ka]

[0054] Human TIM-3 isoform 2 (accession number AAH20843; SEQ ID NO: 85) is soluble and consists of 142 amino acids. It lacks amino acid residues 143-301 of isoform 1, which encode the transmembrane, cytoplasmic, and part of the extracellular domain of TIM-3. Amino acid residues 132-142 also differ from the standard sequence. [ka]

[0055] (5.1 Definition) Unless otherwise defined herein, scientific and technical terms used in this disclosure shall have the meanings commonly understood by those of ordinary skill in the art. Furthermore, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular. Overall, the nomenclature used in connection with, and techniques of, cell and tissue culture, molecular biology, immunology, microbiology, genetics, and protein and nucleic acid chemistry, and hybridization described herein are those well known and commonly used in the art.

[0056] The terms "a" or "an" entity refer to one or more of that entity; for example, "an antibody" is understood to refer to one or more antibodies.

[0057] The term "and / or" when used herein should be interpreted as a specific disclosure of each of the two specified features or components, regardless of the presence or absence of the other. Thus, the term "and / or" as used herein in phrases such as "A and / or B" is intended to include "A and B," "A or B," "A" (only), and "B" (only). Similarly, the term "and / or" as used in phrases such as "A, B, and / or C" is intended to encompass each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (only); B (only); and C (only).

[0058] As used herein, the term "antibody" and its grammatical equivalents refer to an immunoglobulin molecule that recognizes and specifically binds to a target, e.g., a protein, polypeptide, peptide, carbohydrate, polynucleotide, lipid, or any combination of the foregoing, via at least one antigen-binding site (wherein the antigen-binding site is located within the variable region of the immunoglobulin molecule). As used herein, the term encompasses intact polyclonal antibodies, intact monoclonal antibodies, single domain antibodies (sdAbs; camelid antibodies, alpaca antibodies), single-chain Fv (scFv) antibodies, heavy chain antibodies (HCAbs), light chain antibodies (LCAbs), multispecific antibodies, bispecific antibodies, monospecific antibodies, monovalent antibodies, and any other modified immunoglobulin molecule containing an antigen-binding site (e.g., dual variable domain immunoglobulin molecules), so long as the antibody exhibits the desired biological activity. Antibodies also include, but are not limited to, murine antibodies, camelid antibodies, chimeric antibodies, humanized antibodies, and human antibodies. Antibodies can be any of the five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, based on the identity of their heavy chain constant domains, called alpha, delta, epsilon, gamma, and mu, respectively, or their subclasses (isotypes) (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2). Unless expressly indicated otherwise, the term "antibody" as used herein includes "antigen-binding fragments" of intact antibodies. As used herein, the term "antigen-binding fragment" refers to portions or fragments of intact antibodies that are the antigen-determining variable regions of the intact antibody. Examples of antigen-binding fragments include, but are not limited to, Fab, Fab', F(ab'), Fv, linear antibodies, heavy-chain antibody molecules (e.g., scFv), heavy-chain antibodies (HCAbs), light-chain antibodies (LCAbs), disulfide-linked scFv (dsscFv), diabodies, triabodies, tetrabodies, minibodies, dual variable domain antibodies (DVDs), single variable domain antibodies (sdAbs; e.g., camelid antibodies, alpaca antibodies), and single variable domains of heavy-chain antibodies (VHHs), as well as bispecific or multispecific antibodies formed from antibody fragments.A "bispecific" antibody is an artificial hybrid antibody having two different antigen-binding sites that recognize and specifically bind to two different targets. Bispecific antibodies can be produced by a variety of methods, including fusion of hybridomas or linking of Fab' fragments. See, for example, Songsivilai and Lachmann, Clin. Exp. Immunol. 79:315-321 (1990); Kostelny et al., J. Immunol. 148, 1547-1553 (1992).

[0059] The term "humanized antibody," as used herein, refers to forms of non-human (e.g., murine) antibodies that are specific immunoglobulin chains, chimeric immunoglobulins, or fragments thereof that contain minimal non-human sequences. Typically, humanized antibodies are human immunoglobulins. In some cases, Fv framework region residues of a human immunoglobulin are replaced with corresponding residues in an antibody from a non-human species. In some cases, CDR residues are replaced with residues from a CDR of a non-human species (e.g., mouse, rat, hamster, camel) that has the desired specificity, affinity, and / or binding capacity. Humanized antibodies can be further modified by substitution of additional residues either in the Fv framework regions and / or within the replaced non-human residues to improve and optimize the antibody's specificity, affinity, and / or binding capacity. As used herein, the term "human antibody" refers to an antibody having an amino acid sequence corresponding to an antibody produced by a human or produced using any technique known in the art.

[0060] The term "heavy chain" when used in reference to an antibody refers to a polypeptide chain of approximately 50 to 70 kDa, the amino-terminal portion of which contains a variable region of approximately 120 to 130 or more amino acids, and the carboxy-terminal portion of which contains a constant region. The constant region can be one of five different types (e.g., isotypes), called alpha (α), delta (δ), epsilon (ε), gamma (γ), and mu (μ), based on the amino acid sequence of the heavy chain constant region. Different heavy chains vary in size: α, δ, and γ contain approximately 450 amino acids, while μ and ε contain approximately 550 amino acids. When combined with light chains, these different types of heavy chains give rise to the five well-known classes of antibodies: IgA, IgD, IgE, IgG, and IgM, including the four subclasses of IgG, namely, IgG1, IgG2, IgG3, and IgG4, respectively. The heavy chain can be a human heavy chain.

[0061] The term "light chain" when used in reference to an antibody refers to a polypeptide chain of approximately 25 kDa, the amino-terminal portion of which contains a variable region of about 100 to about 110 or more amino acids, and the carboxy-terminal portion of which contains a constant region. The approximate length of a light chain is 211 to 217 amino acids. Based on the amino acid sequence of the constant domain, there are two different types, called lambda (λ) and kappa (κ). The light chain can be a human light chain.

[0062] The term "variable domain" or "variable region" refers to the portion of an antibody light or heavy chain, usually located at the amino terminus of the light or heavy chain, approximately 120-130 amino acids in length for heavy chains and approximately 100-110 amino acids in length for light chains, and is used in relation to the binding and specificity of each particular antibody to its specific antigen. Variable domains vary significantly in sequence among different antibodies. Sequence variability is concentrated in the CDRs, while less variable portions of variable domains are called framework regions (FRs). The CDRs of the light and heavy chains are primarily responsible for the interaction of the antibody with the antigen. The numbering of amino acid positions used herein is according to the EU index, as found in Kabat et al. (1991), Sequences of Proteins of Immunological Interest (US Department of Health and Human Services, Washington, DC), 5th ed. The variable region can be a human variable region.

[0063] CDR refers to one of the three hypervariable regions (H1, H2, or H3) within the non-framework region of an immunoglobulin (Ig or antibody) VH β-sheet framework, or one of the three hypervariable regions (L1, L2, or L3) within the non-framework region of an antibody VL β-sheet framework. Thus, CDRs are variable region sequences interspersed within framework region sequences. CDR regions are well known to those skilled in the art and have been defined by various methods / systems. These systems and / or definitions have been developed and refined over decades and include Kabat, Chothia, IMGT, AbM, and contact. For example, Kabat defined the most hypervariable regions within antibody variable (V) domains (Kabat et al., J. Biol. Chem. 252:6609-6616 (1977); Kabat, Adv. Prot. Chem. 32: 1-75 (1978)). The Chothia definition is based on the location of structural loop regions, which are not part of the conserved β-sheet framework and therefore define CDR region sequences as residues that can adopt various conformations (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)). Both terminologies are well recognized in the art. Furthermore, the IMGT system is based on sequence variability and location within the structure of the variable region. The AbM definition is a compromise between Kabat and Chothia. The contact definition is based on analysis of available antibody crystal structures. Software programs (e.g., abYsis) for analysis of antibody sequences and determination of CDRs are available and known to those skilled in the art. The locations of CDRs within canonical antibody variable domains have been determined by numerous structural comparisons (Al-Lazikani et al., J. Mol. Biol. 273:927-948 (1997); Morea et al., Methods 20:267-279 (2000)).Because the number of residues within a hypervariable region varies among different antibodies, standard variable domain numbering systems conventionally number additional residues relative to the canonical positions with a, b, c, etc. next to the residue number (Al-Lazikani et al., supra (1997)). Such nomenclature is also well known to those skilled in the art.

[0064] For example, CDRs defined according to either the Kabat (hypervariable) notation or the Chothia (structural) notation are set forth in the table below. [Table 1] 1 Residue numbering follows the nomenclature of Kabat et al. (supra). 2 Residue numbering follows the nomenclature of Chothia et al. (supra).

[0065] One or more CDRs can be incorporated into a molecule, either covalently or noncovalently, to form an immunoadhesin. Immunoadhesins can incorporate the CDR(s) as part of a larger polypeptide chain, can be covalently linked to another polypeptide chain, or can incorporate the CDR(s) noncovalently. The CDR(s) enable the immunoadhesin to bind to a specific antigen of interest. CDR regions can be analyzed, for example, at the abysis website (http: / / abysis.org / ).

[0066] The terms "epitope" and "antigenic determinant" are used interchangeably herein and refer to a site on the surface of a target molecule to which an antibody or antigenic fragment binds, e.g., a localized region on the surface of an antigen. Target molecules can include proteins, peptides, nucleic acids, carbohydrates, or lipids. An epitope with immunogenic activity is a portion of a target molecule that elicits an immune response in an animal. An epitope of a target molecule with antigenic activity is a portion of a target molecule to which an antibody binds, as determined by any method known in the art, including, for example, by immunoassay. An antigenic epitope is not necessarily immunogenic. Epitopes often consist of chemically active surface groupings of molecules, such as amino acids or sugar side chains, and have specific three-dimensional structural characteristics and specific charge characteristics. The term "epitope" includes linear epitopes and conformational epitopes. The region of a target molecule (e.g., a polypeptide) that contributes to an epitope may be contiguous amino acids of the polypeptide, or the epitope may be comprised of two or more non-contiguous regions of the target molecule. An epitope may or may not be a three-dimensional surface feature of the target molecule. Epitopes formed from contiguous amino acids (also called linear epitopes) are typically retained upon protein denaturation, whereas epitopes formed by tertiary folding (also called conformational epitopes) are typically lost upon protein denaturation. An epitope typically contains at least three, and more commonly, five, six, seven, or eight to ten amino acids in a unique spatial arrangement.

[0067] As used herein, the term "specifically binds" means that a polypeptide or molecule interacts with an epitope, protein, or target molecule more frequently, more rapidly, for a longer duration, with greater affinity, or with any combination of the above, than with alternative substances, including related and unrelated proteins. Binding moieties (e.g., antibodies) that specifically bind to a target molecule (e.g., an antigen) can be identified, for example, by immunoassays, ELISA, biolayer interferometry ("BLI"), SPR (e.g., Biacore), or other techniques known to those skilled in the art. Typically, a specific response is at least two times the background signal or noise, and may be more than 10 times the background. For a discussion of antibody specificity, see, e.g., Paul (ed.), 1989, Fundamental Immunology, 2nd ed., Raven Press, New York, pp. 332-336. A binding moiety that specifically binds to a target molecule can bind to the target molecule with an affinity that is higher than its affinity for a different molecule. In some embodiments, a binding moiety that specifically binds to a target molecule can bind to the target molecule with an affinity that is at least 20 times greater, at least 30 times greater, at least 40 times greater, at least 50 times greater, at least 60 times greater, at least 70 times greater, at least 80 times greater, at least 90 times greater, or at least 100 times greater than its affinity for a different molecule. In some embodiments, a binding moiety that specifically binds to a particular target molecule binds to a different molecule with such low affinity that binding cannot be detected using assays described herein or otherwise known in the art. In some embodiments, "specifically binds" refers to, for example, a binding moiety that binds to a different molecule with a K D In some embodiments, "specifically binds" means that the polypeptide or molecule binds to a target molecule with a K of about 10 μM or less, or about 1 μM or less. DIn some embodiments, "specifically binds" means that the polypeptide or molecule binds to a target with a K of about 0.1 μM or less, about 0.01 μM or less, or about 1 nM or less. D Specific binding means binding to a target at a specific site. Due to sequence identity between homologous proteins in different species, specific binding can include polypeptides or molecules that recognize proteins or targets in multiple species. Similarly, due to homology within certain regions of the polypeptide sequences of different proteins, specific binding can include polypeptides or molecules that recognize multiple proteins or targets. It is understood that in some embodiments, a binding moiety (e.g., an antibody) that specifically binds to a first target may or may not specifically bind to a second target. Thus, "specific binding" does not necessarily require (although it can include) exclusive binding, i.e., binding to a single target. Thus, a binding moiety (e.g., an antibody) can, in some embodiments, specifically bind to multiple targets. For example, an antibody may, in some cases, contain two identical antigen-binding sites, each capable of specifically binding to the same epitope on two or more proteins. In certain alternative embodiments, an antibody may be bispecific and contain at least two antigen-binding sites with different specificities.

[0068] As used herein, the term "binding affinity" typically refers to the strength of the sum of non-covalent interactions between a binding moiety and a target molecule (e.g., an antigen). Binding between a binding moiety and a target molecule is a reversible process, and the affinity of binding is typically measured by the equilibrium dissociation constant (K D ) is reported as K D is the dissociation rate (k off or k d ) and association rate (k on or k a ) is the ratio of the K DThe lower the K, the higher the affinity. Various methods for measuring binding affinity are known in the art, any of which may be used for purposes of the present disclosure. Specific exemplary embodiments include the following: In some embodiments, "K" is used to measure binding affinity. D " or "K D The "K value" can be measured by assays known in the art, for example, by binding assays. D can be measured by radiolabeled antigen binding assay (RIA) (Chen et al. (1999) J. Mol Biol 293:865-881). D or K D K values ​​can also be measured by using biolayer interferometry (BLI), for example, using the Gator system (Probe Life) or the Octet-96 system (Sartorius AG). D or K D Values ​​can also be measured by using a surface plasmon resonance assay by Biacore, for example, using a BIAcore™-2000 or BIAcore™-3000 (BIAcore, Inc., Piscataway, NJ).

[0069] The term "variant," as used herein with respect to a protein or polypeptide having particular sequence characteristics ("reference polypeptide" or "reference polypeptide"), refers to a different protein or polypeptide that has one or more (e.g., about 1 to about 25, about 1 to about 20, about 1 to about 15, about 1 to about 10, or about 1 to about 5) amino acid substitutions, deletions, and / or additions when compared to the reference protein or polypeptide. The changes to the amino acid sequence can be amino acid substitutions. The changes to the amino acid sequence can be conservative amino acid substitutions. A functional fragment or functional variant of a protein or polypeptide retains essential structural and functional properties of the reference protein or polypeptide.

[0070] The terms "polypeptide," "peptide," and "protein," as used interchangeably herein, and their grammatical equivalents, refer to polymers of amino acids of any length, which can be linear or branched, which may include non-natural or modified amino acids, or which may be interrupted by non-amino acids. A polypeptide, peptide, or protein may also be modified, for example, by disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification.

[0071] The terms "polynucleotide," "nucleic acid," and grammatical equivalents, used interchangeably herein, refer to a polymer of nucleotides of any length, including DNA and RNA. The nucleotides can be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases, and / or their analogs, or any substrate that can be incorporated into a polymer by DNA or RNA polymerase.

[0072] As used herein, the terms "identical," percent "identity," and their grammatical equivalents in the context of two or more polynucleotides or polypeptides refer to two or more sequences or subsequences that are the same or have a specified percentage of the same nucleotide or amino acid residues when compared and aligned for maximum correspondence (introducing gaps, if necessary), not considering conservative amino acid substitutions as part of the sequence identity. Percent identity can be measured using sequence comparison software or algorithms or by visual inspection. Various algorithms and software that can be used to align amino acid or nucleotide sequences are well known in the art. These include, but are not limited to, BLAST, ALIGN, Megalign, BestFit, GCG Wisconsin Package, and variants thereof. In some embodiments, two polynucleotides or polypeptides provided herein are substantially identical, i.e., they share 70%, at least 75%, at least 80%, at least 85%, at least 90%, and in some embodiments, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% nucleotide or amino acid residue identity when compared and aligned for maximum correspondence, as determined using a sequence comparison algorithm or by visual inspection. In some embodiments, identity exists over a region of the amino acid sequence that is at least about 10 residues, at least about 20 residues, at least about 40-60 residues, at least about 60-80 residues, or any integer value therebetween. In some embodiments, identity exists over a region longer than 60-80 residues, e.g., about 80-100 residues, and in some embodiments, the sequences are substantially identical over the entire length of the sequences being compared, e.g., the coding regions of the target proteins or antibodies. In some embodiments, the identity exists over a region of the nucleotide sequence that is at least about 10 bases, at least about 20 bases, at least about 40-60 bases, at least about 60-80 bases in length, or any integer value therebetween.In some embodiments, the identity exists over a region longer than 60-80 bases, e.g., over at least about 80-1000 bases or more, and in some embodiments, the sequences are substantially identical over the entire length of the sequence being compared, e.g., the nucleotide sequence encoding a protein of interest.

[0073] As used herein, the term "vector" and its grammatical equivalents refer to a vehicle used to carry genetic material (e.g., a polynucleotide sequence) that can be introduced into a host cell and replicated and / or expressed therein. Vectors applicable for use include, for example, expression vectors, plasmids, phage vectors, viral vectors, episomes, artificial chromosomes, etc., which can contain operable selection sequences or markers for stable integration into a host cell chromosome. Furthermore, a vector can contain one or more selectable marker genes and appropriate expression control sequences. Selectable marker genes that can be included, for example, provide resistance to antibiotics or toxins, complement auxotrophic deficiencies, or supply critical nutrients not present in the culture medium. Expression control sequences can include constitutive and inducible promoters, transcription enhancers, transcription terminators, and the like, which are well known in the art. When two or more polynucleotides are to be coexpressed, both polynucleotides can be inserted, for example, into a single expression vector or into separate expression vectors. For single vector expression, the encoding polynucleotides can be operably linked to a common expression control sequence or can be linked to different expression control sequences, such as one inducible promoter and one constitutive promoter. Introduction of the polynucleotides into the host cell can be confirmed using methods well known in the art. It will be understood by those skilled in the art that the polynucleotides will be expressed in an amount sufficient to produce the desired product (e.g., an anti-TIM-3 antibody or antigen-binding fragment described herein), and further that expression levels can be optimized to obtain sufficient expression using methods well known in the art.

[0074] As used herein, the term "encode" and its grammatical equivalents refer to the inherent property of a particular sequence of nucleotides in a polynucleotide or nucleic acid, such as a gene, cDNA, or mRNA, to serve as a template for the synthesis of other polymers and macromolecules in biological processes having either a defined sequence of nucleotides (i.e., rRNA, tRNA, mRNA) or a defined sequence of amino acids, and the biological properties resulting therefrom. Thus, a gene encodes a protein if transcription and translation of the mRNA corresponding to that gene produces the protein. Unless otherwise specified, a "nucleotide sequence encoding an amino acid sequence" includes all nucleotide sequences that are degenerate versions of each other and encode the same amino acid sequence. Nucleotide sequences that encode proteins and RNAs can contain introns.

[0075] An "isolated" polypeptide, peptide, protein, antibody, polynucleotide, vector, cell, or composition is a polypeptide, peptide, protein, antibody, polynucleotide, vector, cell, or composition in a form not found in nature. Isolated polypeptides, peptides, proteins, antibodies, polynucleotides, vectors, cells, or compositions include those that have been purified to the extent that they are no longer in the form in which they are found in nature. In some embodiments, an isolated polypeptide, peptide, protein, antibody, polynucleotide, vector, cell, or composition is substantially pure.

[0076] The term "treating" and its grammatical equivalents, as used herein in connection with a disease or disorder, or a subject having a disease or disorder, refers to the act of suppressing, eliminating, alleviating, and / or ameliorating the symptoms, the severity of the symptoms, and / or the frequency of the symptoms associated with the disease or disorder being treated. For example, when used with respect to a cancer or tumor, the term "treating" and its grammatical equivalents refer to the act of reducing the severity of the cancer or tumor or delaying or slowing the progression of the cancer or tumor, including (a) inhibiting the growth of the cancer or tumor or halting the development of the cancer or tumor, (b) causing the regression of the cancer or tumor, or (c) delaying, ameliorating, or minimizing one or more symptoms associated with the presence of the cancer or tumor.

[0077] As used herein, the term "administering" and its grammatical equivalents refer to the act of delivering or causing to be delivered a therapeutic agent or pharmaceutical composition into the body of a subject, by methods described herein or otherwise known in the art. A therapeutic agent can be a compound, a polypeptide, an antibody, a cell, or a population of cells. Administering a therapeutic agent or pharmaceutical composition includes formulating the therapeutic agent or pharmaceutical composition to be delivered into the body of a subject. Exemplary dosage forms include oral dosage forms such as tablets, capsules, syrups, suspensions, etc.; injectable dosage forms such as intravenous (IV), intramuscular (IM), or intraperitoneal (IP); transdermal dosage forms such as creams, jellies, powders, patches, etc.; buccal dosage forms; inhalation powders, sprays, suspensions, and rectal suppositories.

[0078] As used herein, the terms "effective amount," "therapeutically effective amount," and their grammatical equivalents refer to the administration of an agent to a subject, either alone or as part of a pharmaceutical composition, in an amount that, when administered to a subject, is capable of having any detectable positive effect on any symptom, aspect, or characteristic of a disease, disorder, or condition, in a single dose or as part of a series of doses. A therapeutically effective amount can be ascertained by measuring the relevant physiological effect. The precise amount required will vary from subject to subject, depending on the subject's age, weight, and general condition, the severity of the condition being treated, the judgment of the clinician, and the like. An appropriate "effective amount" in any individual case can be determined by one of ordinary skill in the art using routine experimentation.

[0079] The term "pharmaceutically acceptable carrier" or "pharmaceutically acceptable excipient" refers to a material that is suitable for drug administration to an individual along with an active agent without causing any undesired biological effects or interacting in a deleterious manner with any of the other components of the pharmaceutical composition.

[0080] As used herein, the term "subject" refers to any animal (e.g., mammal) that is to be the recipient of a particular treatment, including, but not limited to, humans, non-human primates, dogs, cats, rodents, etc. The subject can be a human. The subject can have a particular disease or disorder.

[0081] Ranges: Throughout this disclosure, various aspects of the invention may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the invention. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values ​​within that range. For example, the description of a range such as 1 to 6 should be considered to have specifically disclosed subranges such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, etc., as well as individual numbers within that range, e.g., 1, 2, 2.7, 3, 4, 5, 5.3, and 6. This applies regardless of the broadness of the range.

[0082] Exemplary genes and polypeptides are described herein by reference to GenBank numbers, GI numbers, and / or SEQ ID NOs. It will be appreciated that one of skill in the art can readily identify homologous sequences by reference to sequence sources, including, but not limited to, GenBank (ncbi.nlm.nih.gov / genbank / ) and EMBL (embl.org / ).

[0083] 5.2 Anti-TIM-3 Antibodies and Antigen-Binding Fragments Provided herein are antibodies or antigen-binding fragments thereof that specifically bind to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are anti-TIM-3 antibodies. In some embodiments, the antibody is an IgA, IgD, IgE, IgG, or IgM antibody. In some embodiments, the antibody is an IgA antibody. In some embodiments, the antibody is an IgD antibody. In some embodiments, the antibody is an IgE antibody. In some embodiments, the antibody is an IgG antibody. In some embodiments, the antibody is an IgM antibody. In some embodiments, the antibody provided herein can be an IgG1 antibody, an IgG2 antibody, an IgG3 antibody, or an IgG4 antibody. In some embodiments, the antibody is an IgG1 antibody. In some embodiments, the antibody is an IgG2 antibody. In some embodiments, the antibody is an IgG3 antibody. In some embodiments, the antibody is an IgG4 antibody.

[0084] In some embodiments, provided herein are antigen-binding fragments of anti-TIM-3 antibodies. In some embodiments, the antigen-binding fragments provided herein can be a single domain antibody (sdAb), a heavy chain antibody (HCAb), a Fab, a Fab', a F(ab')2, an Fv, a single-chain variable fragment (scFv), or an (scFv)2. In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is a single domain antibody (sdAb). In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is a heavy chain antibody (HCAb). In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is a Fab. In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is a Fab'. In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is a F(ab')2. In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is an Fv. In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is an scFv. In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is a disulfide-linked scFv [(scFv)2]. In some embodiments, the antigen-binding fragment of an anti-TIM-3 antibody is a diabody (dAb).

[0085] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise recombinant antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise monoclonal antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise polyclonal antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise camelid (e.g., camel, dromedary, and llama) antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise chimeric antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise humanized antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise human antibodies or antigen-binding fragments. In some embodiments, provided herein are anti-TIM-3 human scFvs.

[0086] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are isolated. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein are substantially pure.

[0087] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise multispecific antibodies or antigen-binding fragments. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise bispecific antibodies or antigen-binding fragments. In some embodiments, provided herein are bispecific T cell engagers (BiTEs). BiTEs are bispecific antibodies that bind to a T cell antigen (e.g., CD3) and a tumor antigen. BiTEs have been shown to induce directed lysis of targeted tumor cells, thus offering significant potential therapies for cancer and other disorders. In some embodiments, provided herein are BiTEs that specifically bind to CD3 and TIM-3. In some embodiments, the BiTEs comprise the anti-TIM-3 antibodies or antigen-binding fragments provided herein. In some embodiments, the BiTEs comprise the anti-TIM-3 scFvs provided herein.

[0088] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise a monovalent antigen-binding site. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments comprise a monospecific binding site. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments comprise a bivalent binding site.

[0089] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment is a monoclonal antibody or antigen-binding fragment. Monoclonal antibodies can be prepared by any method known to those skilled in the art. One exemplary approach is screening a protein expression library, such as a phage or ribosome display library. Phage display is described, for example, in Ladner et al., U.S. Pat. No. 5,223,409; Smith (1985) Science 228:1315-1317; and WO 92 / 18619. In some embodiments, recombinant monoclonal antibodies are isolated from a phage display library expressing the variable regions or CDRs of the desired species. Screening of phage libraries can be accomplished by various techniques known in the art.

[0090] In some embodiments, monoclonal antibodies are prepared using hybridoma methods known to those skilled in the art. For example, using hybridoma methods, a mouse, rat, rabbit, hamster, or other suitable host animal is immunized as described above. In some embodiments, lymphocytes are immunized in vitro. In some embodiments, the immunizing antigen is a human protein or a fragment thereof. In some embodiments, the immunizing antigen is a human protein or a fragment thereof.

[0091] After immunization, lymphocytes are isolated and fused with a suitable myeloma cell line, e.g., using polyethylene glycol. Hybridoma cells are selected using specialized media known in the art; unfused lymphocytes and myeloma cells do not survive the selection process. Hybridomas producing monoclonal antibodies against the selected antigen can be identified by various methods, including, but not limited to, immunoprecipitation, immunoblotting, and in vitro binding assays (e.g., flow cytometry, FACS, ELISA, BLI, SPR (e.g., Biacore), and radioimmunoassays). Once hybridoma cells producing antibodies of the desired specificity, affinity, and / or activity are identified, clones can be subcloned by limiting dilution or other techniques. Hybridomas can be grown in vitro in culture using standard methods or in vivo as ascites tumors in animals. Monoclonal antibodies can be purified from the culture medium or ascites fluid according to standard methods in the art, including, but not limited to, affinity chromatography, ion exchange chromatography, gel electrophoresis, and dialysis.

[0092] In some embodiments, monoclonal antibodies are produced using recombinant DNA techniques known to those skilled in the art. For example, polynucleotides encoding the antibodies are isolated from mature B cells or hybridoma cells, e.g., by RT-PCR using oligonucleotide primers that specifically amplify genes encoding the heavy and light chains of the antibody, and their sequences are determined using standard techniques. The isolated polynucleotides encoding the heavy and light chains are then cloned into a suitable expression vector that produces the monoclonal antibody when transfected into host cells that do not otherwise produce immunoglobulin proteins, such as E. coli, monkey COS cells, Chinese hamster ovary (CHO) cells, or myeloma cells.

[0093] In some embodiments, monoclonal antibodies are modified using recombinant DNA technology to generate alternative antibodies. In some embodiments, the constant domains of the light and heavy chains of a mouse monoclonal antibody are replaced with the constant regions of a human antibody to generate chimeric antibodies. In some embodiments, the constant regions are truncated or removed to generate desired antibody fragments of the monoclonal antibody. In some embodiments, site-directed or high-density mutagenesis of the variable regions is used to optimize the specificity and / or affinity of the monoclonal antibody.

[0094] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment is a humanized antibody or antigen-binding fragment. Various methods for generating humanized antibodies are known in the art. Methods for achieving high-affinity binding with humanized antibodies are known in the art. A non-limiting example of such a method is hypermutation of the variable regions and selection of cells expressing such high-affinity antibodies (affinity maturation). In addition to using display libraries, a specific antigen (e.g., recombinant TIM-3 or an epitope thereof) can be used to immunize a non-human animal, e.g., a rodent. In certain embodiments, rodent antigen-binding fragments (e.g., murine antigen-binding fragments) can be generated and isolated using methods known in the art and / or disclosed herein. In some embodiments, mice can be immunized with the antigen (e.g., recombinant TIM-3 or an epitope thereof).

[0095] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment is a human antibody or antigen-binding fragment. Human antibodies can be prepared using various techniques known in the art. In some embodiments, human antibodies are produced from immortalized human B lymphocytes immunized in vitro. In some embodiments, human antibodies are produced from lymphocytes isolated from immunized individuals. In either case, cells producing antibodies against the target antigen can be generated and isolated. In some embodiments, human antibodies are selected from phage libraries, where the phage library expresses human antibodies. Alternatively, phage display technology can be used to generate human antibodies and antibody fragments in vitro from immunoglobulin variable region gene repertoires derived from unimmunized donors. Techniques for generating and using antibody phage libraries are well known in the art. Once an antibody is identified, affinity maturation strategies known in the art, including but not limited to chain shuffling and site-directed mutagenesis, can be used to generate higher affinity human antibodies. In some embodiments, human antibodies are produced in transgenic mice containing human immunoglobulin loci. Upon immunization, these mice are capable of producing a full repertoire of human antibodies in the absence of endogenous immunoglobulin production.

[0096] The specific CDR sequences defined herein are generally based on a combination of the Kabat and Chothia definitions, however, it is understood that reference to the heavy chain CDR(s) and / or light chain CDR(s) of a specific antibody encompasses all CDR definitions known to those skilled in the art.

[0097] The anti-TIM-3 antibodies or antigen-binding fragments provided herein include the following clones: 3E6, 4H2, 16H1, 18C6, 19D11, CH5#, CH8#, CH9#, CH10#, CH11#, 3F2, 36A2, 36B11, 38F8, 38A8, 40F11, 50H9, 84G10, and 39D1. Sequence characteristics are described below. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise one, two, three, four, five, and / or six CDRs of any one of the antibodies described herein. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise one, two, three, four, five, and / or six CDRs of 3E6, 4H2, 16H1, 18C6, 19D11, CH5#, CH8#, CH9#, CH10#, CH11#, 3F2, 36A2, 36B11, 38F8, 38A8, 40F11, 50H9, 84G10, or 39D1. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise a VL that comprises one, two, and / or three VL CDRs of Tables 1a-1c. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise a VH that comprises one, two, and / or three VH CDRs of Tables 2a-2b. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise one, two, and / or three VL CDRs of Table 1 and one, two, and / or three VH CDRs of Tables 2a-2b. Table 1a. Amino acid sequences of the light chain variable region CDRs (VL CDRs; Kabat notation) of anti-TIM-3 Abs [Table 2] Table 1b. Amino acid sequences of VL CDRs (alternative designations) of anti-TIM-3 Abs [Table 3] Table 1c. Amino acid sequence of 3E6 VL CDR (affinity matured) [Table 4] Table 2a. Amino acid sequences of the heavy chain variable region CDRs (VH CDRs; Kabat notation) of anti-TIM-3 Abs [Table 5] Table 2b. Amino acid sequences of VH CDRs (alternative designations) of anti-TIM-3 Abs [Table 6]

[0098] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof comprises a humanized antibody or antigen-binding fragment. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof comprises a VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and / or VH CDR3 from an antibody or antigen-binding fragment described herein. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof comprises a variant of the anti-TIM-3 antibody or antigen-binding fragment described herein. In some embodiments, the variant of the anti-TIM-3 antibody or antigen-binding fragment comprises 1 to 30 amino acid substitutions, additions, and / or deletions in the anti-TIM-3 antibody or antigen-binding fragment. In some embodiments, the variant of the anti-TIM-3 antibody or antigen-binding fragment comprises 1 to 25 amino acid substitutions, additions, and / or deletions in the anti-TIM-3 antibody or antigen-binding fragment. In some embodiments, the variant anti-TIM-3 antibody or antigen-binding fragment comprises 1 to 20 substitutions, additions, and / or deletions in the anti-TIM-3 antibody or antigen-binding fragment. In some embodiments, the variant anti-TIM-3 antibody or antigen-binding fragment comprises 1 to 15 substitutions, additions, and / or deletions in the anti-TIM-3 antibody or antigen-binding fragment. In some embodiments, the variant anti-TIM-3 antibody or antigen-binding fragment comprises 1 to 10 substitutions, additions, and / or deletions in the anti-TIM-3 antibody or antigen-binding fragment. In some embodiments, the variant anti-TIM-3 antibody or antigen-binding fragment comprises 1 to 5 conservative amino acid substitutions, additions, and / or deletions in the anti-TIM-3 antibody or antigen-binding fragment. In some embodiments, the variant anti-TIM-3 antibody or antigen-binding fragment comprises 1 to 3 amino acid substitutions, additions, and / or deletions in the anti-TIM-3 antibody or antigen-binding fragment. In some embodiments, the amino acid substitutions, additions, and / or deletions are conservative amino acid substitutions. In some embodiments, the conservative amino acid substitutions are in the CDRs of the antibody or antigen-binding fragment. In some embodiments, the conservative amino acid substitutions are not in the CDRs of the antibody or antigen-binding fragment. In some embodiments, the conservative amino acid substitutions are in the framework regions of the antibody or antigen-binding fragment.

[0099] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a light chain variable region (VL) comprising: (1) a light chain CDR1 (VL CDR1) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 86-93; (2) a light chain CDR2 (VL CDR2) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 94-100; or (3) a light chain CDR3 (VL CDR3) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47-55 and 198-206; or a variant thereof having up to about 3, about 5, about 8, about 10, about 12, or about 15 amino acid substitutions, additions, and / or deletions in the VL CDR. In some embodiments, the variant has about 5 amino acid substitutions, additions, and / or deletions in the VL CDR. In some embodiments, the antibody or antigen-binding fragment comprises all three VL CDRs.

[0100] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a heavy chain variable region (VH) comprising: (1) a heavy chain CDR1 (VH CDR1) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 101-108; (2) a heavy chain CDR2 (VH CDR2) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 109-118; or (3) a heavy chain CDR3 (VH CDR3) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 119-128; or a variant thereof having up to about 3, about 5, about 8, about 10, about 12, or about 15 amino acid substitutions, additions, and / or deletions in the VH CDR. In some embodiments, the variant has about 5 amino acid substitutions, additions, and / or deletions in the VH CDR. In some embodiments, the antibody or antigen-binding fragment comprises all three VH CDRs.

[0101] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising: (a) a VL comprising: (1) a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 86-93; (2) a VL CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 94-100; and (3) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47-55 and 198-206; or a variant thereof having up to about five amino acid substitutions, additions, and / or deletions in the VL CDR; and (b) a VH comprising: (1) a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 101-108; (2) a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 109-118; and (3) a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 119-128; or a variant thereof having up to about five amino acid substitutions, additions, and / or deletions in the VH CDR.

[0102] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3 having a VL, wherein the VL comprises a VL CDR1, CDR2, and CDR3 having specific sequences. The VL CDR1, CDR2, and CDR3 can have (1) the amino acid sequences of SEQ ID NOs: 86, 94, and 47, respectively. The VL CDR1, CDR2, and CDR3 can have (2) the amino acid sequences of SEQ ID NOs: 87, 95, and 48, respectively. The VL CDR1, CDR2, and CDR3 can have (3) the amino acid sequences of SEQ ID NOs: 88, 96, and 49, respectively. The VL CDR1, CDR2, and CDR3 can have (4) the amino acid sequences of SEQ ID NOs: 89, 97, and 50, respectively. The VL CDR1, CDR2, and CDR3 can have (5) the amino acid sequences of SEQ ID NOs: 90, 94, and 51, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (6) SEQ ID NOs: 91, 98, and 52, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (7) SEQ ID NOs: 91, 98, and 53, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (8) SEQ ID NOs: 92, 99, and 54, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (9) SEQ ID NOs: 93, 100, and 55, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (10) SEQ ID NOs: 129, 138, and 145, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (11) SEQ ID NOs: 130, 139, and 146, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (12) SEQ ID NOs: 131, 140, and 147, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (13) SEQ ID NOs: 132, 141, and 148, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (14) SEQ ID NOs: 133, 139, and 149, respectively.The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (15) SEQ ID NOs: 134, 142, and 150, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (16) SEQ ID NOs: 135, 143, and 151, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (17) SEQ ID NOs: 136, 144, and 152, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (18) SEQ ID NOs: 137, 100, and 153, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (19) SEQ ID NOs: 86, 94, and 198, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (20) SEQ ID NOs: 86, 94, and 199, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (21) SEQ ID NOs: 86, 94, and 200, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (22) SEQ ID NOs: 86, 94, and 201, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (23) SEQ ID NOs: 86, 94, and 202, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (24) SEQ ID NOs: 86, 94, and 203, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (25) SEQ ID NOs: 86, 94, and 204, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (26) SEQ ID NOs: 86, 94, and 205, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (27) SEQ ID NOs: 86, 94, and 206, respectively. The VL can be a variant of the above VL having up to about 3, about 5, about 8, about 10, about 12, or about 15 amino acid substitutions, additions, and / or deletions in the VL CDRs. In some embodiments, the variant has up to about 5 amino acid substitutions, additions, and / or deletions in the VL CDRs.

[0103] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, having a VH, wherein the VH comprises a VH CDR1, CDR2, and CDR3 having specific sequences. The VH CDR1, CDR2, and CDR3 can have (1) the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively. The VH CDR1, CDR2, and CDR3 can have (2) the amino acid sequences of SEQ ID NOs: 102, 110, and 120, respectively. The VH CDR1, CDR2, and CDR3 can have (3) the amino acid sequences of SEQ ID NOs: 103, 111, and 121, respectively. The VH CDR1, CDR2, and CDR3 can have (4) the amino acid sequences of SEQ ID NOs: 104, 112, and 122, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (5) SEQ ID NOs: 105, 113, and 123, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (6) SEQ ID NOs: 106, 114, and 124, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (7) SEQ ID NOs: 106, 115, and 125, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (8) SEQ ID NOs: 107, 116, and 126, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (9) SEQ ID NOs: 108, 117, and 127, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (10) SEQ ID NOs: 106, 118, and 128, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (11) SEQ ID NOs: 106, 162, and 171, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (12) SEQ ID NOs: 154, 163, and 172, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (13) SEQ ID NOs: 155, 164, and 173, respectively.The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (14) SEQ ID NOs: 156, 165, and 174, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (15) SEQ ID NOs: 157, 166, and 175, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (16) SEQ ID NOs: 158, 167, and 176, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (17) SEQ ID NOs: 159, 168, and 177, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (18) SEQ ID NOs: 160, 169, and 178, respectively. The VH CDR1, CDR2, and CDR3 can have the amino acid sequences of (19) SEQ ID NOs: 161, 170, and 179, respectively. The VH can be a variant of the above VH having up to about 3, about 5, about 8, about 10, about 12, or about 15 amino acid substitutions, additions, and / or deletions in the VH CDRs. In some embodiments, the variant has up to about 5 amino acid substitutions, additions, and / or deletions in the VH CDRs.

[0104] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL and a VH. In some embodiments, the VL and VH are connected by a linker. In some embodiments, the linker has the amino acid sequence of (GGGGS)n, where n=1, 2, 3, 4, or 5 (SEQ ID NO: 216). In some embodiments, the linker has the amino acid sequence of (EAAAK)n, where n=1, 2, 3, 4, or 5 (SEQ ID NO: 217). In some embodiments, the linker is [ka] It has the amino acid sequence:

[0105] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3 having a VL and a VH, wherein (a) the VL is selected from the group consisting of: (1) SEQ ID NOs: 86, 94, and 47, respectively; (2) SEQ ID NOs: 87, 95, and 48, respectively; (3) SEQ ID NOs: 88, 96, and 49, respectively; (4) SEQ ID NOs: 89, 97, and 50, respectively; and (5) SEQ ID NOs: 90, 94, and 51, respectively. (6) SEQ ID NOs: 91, 98, and 52, respectively; (7) SEQ ID NOs: 91, 98, and 53, respectively; (8) SEQ ID NOs: 92, 99, and 54, respectively; (9) SEQ ID NOs: 93, 100, and 55, respectively; (10) SEQ ID NOs: 129, 138, and 145, respectively; (11) SEQ ID NOs: 130, 139, and 146, respectively; (12) SEQ ID NOs: 131, 140, and 147, respectively; (13) SEQ ID NOs: 132, 133, and 134, respectively. 41, and 148; (14) SEQ ID NOs: 133, 139, and 149, respectively; (15) SEQ ID NOs: 134, 142, and 150, respectively; (16) SEQ ID NOs: 135, 143, and 151, respectively; (17) SEQ ID NOs: 136, 144, and 152, respectively; (18) SEQ ID NOs: 137, 100, and 153, respectively; (19) SEQ ID NOs: 86, 94, and 198, respectively; (20) SEQ ID NOs: 86, 94, and 19, respectively. 9; (21) SEQ ID NOs: 86, 94, and 200, respectively; (22) SEQ ID NOs: 86, 94, and 201, respectively; (23) SEQ ID NOs: 86, 94, and 202, respectively; (24) SEQ ID NOs: 86, 94, and 203, respectively; (25) SEQ ID NOs: 86, 94, and 204, respectively; (26) SEQ ID NOs: 86, 94, and 205, respectively; or (27) VLs having the amino acid sequences of SEQ ID NOs: 86, 94, and 206, respectively. or variants thereof having up to about three amino acid substitutions, additions, and / or deletions in the VL CDRs; and / or (b) VH CDR1, CDR2, and CDR3 are (1) SEQ ID NOs: 101, 109, and 119, respectively; (2) SEQ ID NOs: 102, 110, and 120, respectively; (3) SEQ ID NOs: 103, 111, and 121, respectively; (4) SEQ ID NOs: 104, 112, and 122, respectively; (5) SEQ ID NOs: 105, 113, and 123, respectively;(6) SEQ ID NOs: 106, 114, and 124, respectively; (7) SEQ ID NOs: 106, 115, and 125, respectively; (8) SEQ ID NOs: 107, 116, and 126, respectively; (9) SEQ ID NOs: 108, 117, and 127, respectively; (10) SEQ ID NOs: 106, 118, and 128, respectively; (11) SEQ ID NOs: 106, 162, and 171, respectively; (12) SEQ ID NOs: 154, 163, and 172, respectively; (13) SEQ ID NOs: 154, 163, and 172, respectively. (14) SEQ ID NOs: 156, 165, and 174, respectively; (15) SEQ ID NOs: 157, 166, and 175, respectively; (16) SEQ ID NOs: 158, 167, and 176, respectively; (17) SEQ ID NOs: 159, 168, and 177, respectively; (18) SEQ ID NOs: 160, 169, and 178, respectively; or (19) SEQ ID NOs: 161, 170, and 179, respectively; or variants thereof having up to about five amino acid substitutions, additions, and / or deletions in the VH CDRs.

[0106] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, having a VL and a VH, wherein the VL comprises a VL CDR1, CDR2, and CDR3, and the VH comprises a VH CDR1, CDR2, and CDR3, and wherein the VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have specific sequences. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (1) SEQ ID NOs: 86, 94, 47, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (2) SEQ ID NOs: 87, 95, 48, 102, 110, and 120, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (3) SEQ ID NOs: 88, 96, 49, 103, 111, and 121. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (4) SEQ ID NOs: 89, 97, 50, 104, 112, and 122, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (5) SEQ ID NOs: 90, 94, 51, 105, 113, and 123, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (6) SEQ ID NOs: 91, 98, 52, 106, 114, and 124, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (7) SEQ ID NOs: 91, 98, 53, 106, 115, and 125, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (8) SEQ ID NOs: 92, 99, 54, 107, 116, and 126, respectively.The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (9) SEQ ID NOs: 93, 100, 55, 108, 117, and 127, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (10) SEQ ID NOs: 91, 98, 52, 106, 118, and 128, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (11) SEQ ID NOs: 129, 138, 145, 106, 162, and 171, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (12) SEQ ID NOs: 130, 139, 146, 154, 163, and 172, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (13) SEQ ID NOs: 131, 140, 147, 155, 164, and 173, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (14) SEQ ID NOs: 132, 141, 148, 156, 165, and 174, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (15) SEQ ID NOs: 133, 139, 149, 157, 166, and 175, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (16) SEQ ID NOs: 134, 142, 150, 158, 167, and 176, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (17) SEQ ID NOs: 135, 143, 151, 159, 168, and 177, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (18) SEQ ID NOs: 136, 144, 152, 160, 169, and 178, respectively.The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (19) SEQ ID NOs: 137, 100, 153, 161, 170, and 179, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (20) SEQ ID NOs: 86, 94, 198, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (21) SEQ ID NOs: 86, 94, 199, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (22) SEQ ID NOs: 86, 94, 200, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (23) SEQ ID NOs: 86, 94, 201, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (24) SEQ ID NOs: 86, 94, 202, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (25) SEQ ID NOs: 86, 94, 203, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (26) SEQ ID NOs: 86, 94, 204, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (27) SEQ ID NOs: 86, 94, 205, 101, 109, and 119, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 can have (28) SEQ ID NOs: 86, 94, 206, 101, 109, and 119, respectively.The antibody or antigen-binding fragment can be a variant of the above-described antibody or antigen-binding fragment having up to about 5 amino acid substitutions, additions, and / or deletions in the CDRs.

[0107] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL comprising: (1) a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 31-38; (2) a VL CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 39-46; or (3) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47-55; or a variant thereof having up to about 3, about 5, about 8, about 10, about 12, or about 15 amino acid substitutions, additions, and / or deletions in the VL CDR. In some embodiments, the variant has about 5 amino acid substitutions, additions, and / or deletions in the VL CDR. In some embodiments, the antibody or antigen-binding fragment comprises all three VL CDRs.

[0108] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VH comprising: (1) a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 56-63; (2) a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 64-73; or (3) a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 74-83; or a variant thereof having up to about 3, about 5, about 8, about 10, about 12, or about 15 amino acid substitutions, additions, and / or deletions in the VH CDR. In some embodiments, the variant has about 5 amino acid substitutions, additions, and / or deletions in the VH CDR. In some embodiments, the antibody or antigen-binding fragment comprises all three VH CDRs.

[0109] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising: (a) a VL comprising: (1) a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 31 to 38; (2) a VL CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 39 to 46; and (3) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47 to 55; or a variant thereof having up to about five amino acid substitutions, additions, and / or deletions in the VL CDR; and (b) a VH comprising: (1) a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 56 to 63; (2) a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 64 to 73; and (3) a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 74 to 83; or a variant thereof having up to about five amino acid substitutions, additions, and / or deletions in the VH CDR.

[0110] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3 having a VL, wherein the VL comprises a VL CDR1, CDR2, and CDR3 having a specific sequence. The VL CDR1, CDR2, and CDR3 can have (1) the amino acid sequences of SEQ ID NOs: 31, 39, and 47, respectively. The VL CDR1, CDR2, and CDR3 can have (2) the amino acid sequences of SEQ ID NOs: 32, 40, and 48, respectively. The VL CDR1, CDR2, and CDR3 can have (3) the amino acid sequences of SEQ ID NOs: 33, 41, and 49, respectively. The VL CDR1, CDR2, and CDR3 can have (4) the amino acid sequences of SEQ ID NOs: 34, 42, and 50, respectively. The VL CDR1, CDR2, and CDR3 can have (5) the amino acid sequences of SEQ ID NOs: 35, 43, and 51, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (6) SEQ ID NOs: 36, 44, and 52, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (7) SEQ ID NOs: 36, 44, and 53, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (8) SEQ ID NOs: 37, 45, and 54, respectively. The VL CDR1, CDR2, and CDR3 can have the amino acid sequences of (9) SEQ ID NOs: 38, 46, and 55, respectively. The VL can be a variant of the VL described above having up to about 3, about 5, about 8, about 10, about 12, or about 15 amino acid substitutions, additions, and / or deletions in the VL CDRs. In some embodiments, the variant has up to about 5 amino acid substitutions, additions, and / or deletions in the VL CDRs.

[0111] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3 having a VH, wherein the VH comprises VH CDR1, CDR2, and CDR3 having the amino acid sequences of (1) SEQ ID NOs: 56, 64, and 74, respectively. The VH CDR1, CDR2, and CDR3 can have (2) SEQ ID NOs: 57, 65, and 75, respectively. The VH CDR1, CDR2, and CDR3 can have (3) SEQ ID NOs: 58, 66, and 76, respectively. The VH CDR1, CDR2, and CDR3 can have (4) SEQ ID NOs: 59, 67, and 77, respectively. The VH CDR1, CDR2, and CDR3 can have (5) SEQ ID NOs: 60, 68, and 78, respectively. The VH CDR1, CDR2, and CDR3 can have (6) SEQ ID NOs: 61, 69, and 79, respectively. The VH CDR1, CDR2, and CDR3 can have (7) SEQ ID NOs: 61, 70, and 80, respectively. The VH CDR1, CDR2, and CDR3 can have (8) SEQ ID NOs: 62, 71, and 81, respectively. The VH CDR1, CDR2, and CDR3 can have (9) SEQ ID NOs: 63, 72, and 82, respectively. The VH CDR1, CDR2, and CDR3 can have (10) SEQ ID NOs: 61, 73, and 83, respectively. In some embodiments, the VH can be a variant of the VH described above having up to about 3, about 5, about 8, about 10, about 12, or about 15 amino acid substitutions, additions, and / or deletions in the VH CDRs. In some embodiments, the variant has up to about 5 amino acid substitutions, additions, and / or deletions in the VH CDRs.

[0112] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL and a VH. In some embodiments, the VL and VH are connected by a linker. In some embodiments, the linker has the amino acid sequence of (GGGGS)n, where n=1, 2, 3, 4, or 5 (SEQ ID NO: 216). In some embodiments, the linker has the amino acid sequence of (EAAAK)n, where n=1, 2, 3, 4, or 5 (SEQ ID NO: 217). In some embodiments, the linker is [ka] It has the amino acid sequence:

[0113] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, having a VL and a VH, wherein (a) the VL has VL CDR1, CDR2, and CDR3 having the amino acid sequences of: (1) SEQ ID NOs: 31, 39, and 47, respectively; (2) SEQ ID NOs: 32, 40, and 48, respectively; (3) SEQ ID NOs: 33, 41, and 49, respectively; (4) SEQ ID NOs: 34, 42, and 50, respectively; (5) SEQ ID NOs: 35, 43, and 51, respectively; (6) SEQ ID NOs: 36, 44, and 52, respectively; (7) SEQ ID NOs: 36, 44, and 53, respectively; (8) SEQ ID NOs: 37, 45, and 54, respectively; or (9) SEQ ID NOs: 38, 46, and 55, respectively; or (9) SEQ ID NOs: 63, 72, and 82, respectively; or (10) VH CDR1, CDR2, and CDR3 having the amino acid sequences of SEQ ID NOs: 61, 73, and 83, respectively; or variants thereof having up to about five amino acid substitutions, additions, and / or deletions in the VH CDRs; and (b) VH comprises VH CDR1, CDR2, and CDR3 having the amino acid sequences of SEQ ID NOs: 61, 73, and 83, respectively; or variants thereof having up to about five amino acid substitutions, additions, and / or deletions in the VH CDRs.

[0114] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, having a VL and a VH, wherein the VL comprises VL CDR1, CDR2, and CDR3, and the VH comprises VH CDR1, CDR2, and CDR3, and wherein the VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (1) SEQ ID NOs: 31, 39, 47, 56, 64, and 74, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (2) SEQ ID NOs: 32, 40, 48, 57, 65, and 75, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (3) SEQ ID NOs: 33, 41, 49, 58, 66, and 76, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (4) SEQ ID NOs: 34, 42, 50, 59, 67, and 77, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (5) SEQ ID NOs: 35, 43, 51, 60, 68, and 78, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (6) SEQ ID NOs: 36, 44, 52, 61, 69, and 79, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (7) SEQ ID NOs: 36, 44, 53, 61, 70, and 80, respectively. The VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (8) SEQ ID NOs: 37, 45, 54, 62, 71, and 81, respectively. VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (9) SEQ ID NOs: 38, 46, 55, 63, 72, and 82, respectively.VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2, and VH CDR3 have the amino acid sequences of (10) SEQ ID NOs: 36, 44, 52, 61, 73, and 83, respectively. The antibody or antigen-binding fragment can be a variant of the above-described antibody or antigen-binding fragment having up to about 5 amino acid substitutions, additions, and / or deletions in the CDRs.

[0115] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise the VL and / or VH of any one of the antibodies described herein. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments provided herein comprise the VL and / or VH of 3E6, 4H2, 16H1, 18C6, 19D11, CH5#, CH8#, CH9#, CH10#, CH11#, 3F2, 36A2, 36B11, 38F8, 38A8, 40F11, 50H9, 84G10, or 39D1. Table 3. Amino acid sequences of the light chain variable region (VL) and heavy chain variable region (VH) of anti-TIM-3 antibodies [Table 7] TIFF2024516616000012.tif225170TIFF2024516616000013.tif125170

[0116] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are humanized versions of any one of the antibodies described herein. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are humanized versions of 3E6, 4H2, 16H1, 18C6, 19D11, CH5#, CH8#, CH9#, CH10#, CH11#, 3F2, 36A2, 36B11, 38F8, 38A8, 40F11, 50H9, 84G10, or 39D1. Exemplary sequences are provided below. Table 4a. Amino acid sequences of VL and VH of humanized anti-TIM-3 antibodies [Table 8] Table 4b. Amino acid sequences of affinity-matured 3E6 VL0s [Table 9]

[0117] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL having at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-10 and 180-188. In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VH having at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 11-20 and 189-197. In some embodiments, the antibody or antigen-binding fragment comprises both a VL and a VH.

[0118] An anti-TIM-3 antibody or antigen-binding fragment thereof can comprise a combination of any VL disclosed herein and any VH disclosed herein. In some embodiments, the VL and VH are connected by a linker. In some embodiments, the linker has the amino acid sequence (GGGGS)n, where n=1, 2, 3, 4, or 5 (SEQ ID NO: 216). In some embodiments, the linker has the amino acid sequence (EAAAK)n, where n=1, 2, 3, 4, or 5 (SEQ ID NO: 217). In some embodiments, the linker is [ka] It has the amino acid sequence:

[0119] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL and a VH, wherein the VH has an amino acid sequence selected from the group consisting of SEQ ID NOs: 11-20 and 189-197, and the VL has a specific sequence selected from the group consisting of SEQ ID NOs: 1-10 and 180-188. The VL can have the amino acid sequence of SEQ ID NO: 1. The VL can have the amino acid sequence of SEQ ID NO: 2. The VL can have the amino acid sequence of SEQ ID NO: 3. The VL can have the amino acid sequence of SEQ ID NO: 4. The VL can have the amino acid sequence of SEQ ID NO: 5. The VL can have the amino acid sequence of SEQ ID NO: 6. The VL can have the amino acid sequence of SEQ ID NO: 7. The VL can have the amino acid sequence of SEQ ID NO: 8. The VL can have the amino acid sequence of SEQ ID NO: 9. The VL can have the amino acid sequence of SEQ ID NO: 10. The VL can have the amino acid sequence of SEQ ID NO: 180. The VL can have the amino acid sequence of SEQ ID NO: 181. The VL can have the amino acid sequence of SEQ ID NO: 182. The VL can have the amino acid sequence of SEQ ID NO: 183. The VL can have the amino acid sequence of SEQ ID NO: 184. The VL can have the amino acid sequence of SEQ ID NO: 185. The VL can have the amino acid sequence of SEQ ID NO: 186. The VL can have the amino acid sequence of SEQ ID NO: 187. The VL can have the amino acid sequence of SEQ ID NO: 188.

[0120] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL and a VH, wherein the VL has an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-10 and 180-188, and the VH has a specific sequence selected from the group consisting of SEQ ID NOs: 11-20 and 189-197. The VH can have the amino acid sequence of SEQ ID NO: 11. The VH can have the amino acid sequence of SEQ ID NO: 12. The VH can have the amino acid sequence of SEQ ID NO: 13. The VH can have the amino acid sequence of SEQ ID NO: 14. The VH can have the amino acid sequence of SEQ ID NO: 15. The VH can have the amino acid sequence of SEQ ID NO: 16. The VH can have the amino acid sequence of SEQ ID NO: 17. The VH can have the amino acid sequence of SEQ ID NO: 18. The VH can have the amino acid sequence of SEQ ID NO: 19. The VH can have the amino acid sequence of SEQ ID NO: 20. The VH can have the amino acid sequence of SEQ ID NO: 189. The VH can have the amino acid sequence of SEQ ID NO: 190. The VH can have the amino acid sequence of SEQ ID NO: 191. The VH can have the amino acid sequence of SEQ ID NO: 192. The VH can have the amino acid sequence of SEQ ID NO: 193. The VH can have the amino acid sequence of SEQ ID NO: 194. The VH can have the amino acid sequence of SEQ ID NO: 195. The VH can have the amino acid sequence of SEQ ID NO: 196. The VH can have the amino acid sequence of SEQ ID NO: 197.

[0121] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL, wherein the VL has at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 1. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 2. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 3. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 4. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:5.The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 6. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 7. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 8. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:9. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:10.The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 180. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 181. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 182. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 183. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 184.The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 185. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 186. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 187. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 188.

[0122] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VH, wherein the VH has at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 11. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 12. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 13. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 14. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 15.The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 16. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 17. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 18. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 19. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:20.The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 189. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 190. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 191. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 192. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 193.The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 194. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 195. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 196. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 197.

[0123] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL and a VH, wherein the VL and VH have the amino acid sequences of SEQ ID NOs: 1 and 11, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 2 and 12, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 3 and 13, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 4 and 14, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 5 and 15, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 6 and 16, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 7 and 17, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 8 and 18, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 9 and 19, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 10 and 20, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 180 and 189, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 181 and 190, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 182 and 191, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 183 and 192, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 184 and 193, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 185 and 194, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 186 and 195, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 187 and 196, respectively. In some embodiments, the VL and VH have the amino acid sequences of SEQ ID NOs: 188 and 197, respectively.

[0124] In some embodiments, provided herein is a humanized antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL having at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 21 and 207-215. In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VH having at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29. In some embodiments, the humanized antibody or antigen-binding fragment comprises both a VL and a VH.

[0125] An anti-TIM-3 antibody or antigen-binding fragment thereof can comprise a combination of any VL disclosed herein and any VH disclosed herein. In some embodiments, the VL and VH are connected by a linker. In some embodiments, the linker has the amino acid sequence (GGGGS)n, where n=1, 2, 3, 4, or 5 (SEQ ID NO: 216). In some embodiments, the linker has the amino acid sequence (EAAAK)n, where n=1, 2, 3, 4, or 5 (SEQ ID NO: 217). In some embodiments, the linker is [ka] It has the amino acid sequence:

[0126] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL, wherein the VL has at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 21. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 207. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 208. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 209. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:210.The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 211. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 212. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 213. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 214. The VL can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:215.

[0127] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VH, wherein the VH has at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 22. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 23. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 24. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 25. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:26.The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 27. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 28. The VH can have at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:29.

[0128] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL and a VH, wherein the VL and VH have the amino acid sequences of SEQ ID NOs: 21 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 21 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 21 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 21 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 21 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 21 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 21 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 21 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 207 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 207 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 207 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 207 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 207 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 207 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 207 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 207 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 208 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 208 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 208 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 208 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 208 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 208 and 27, respectively.The VL and VH can have the amino acid sequences of SEQ ID NOs: 208 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 208 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 209 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 209 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 209 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 209 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 209 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 209 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 209 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 209 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 210 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 210 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 210 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 210 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 210 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 210 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 210 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 210 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 211 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 211 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 211 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 211 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 211 and 26, respectively.The VL and VH can have the amino acid sequences of SEQ ID NOs: 211 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 211 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 211 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 212 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 212 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 212 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 212 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 212 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 212 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 212 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 212 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 213 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 213 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 213 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 213 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 213 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 213 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 213 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 213 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 214 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 214 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 214 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 214 and 25, respectively.The VL and VH can have the amino acid sequences of SEQ ID NOs: 214 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 214 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 214 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 214 and 29, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 215 and 22, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 215 and 23, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 215 and 24, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 215 and 25, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 215 and 26, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 215 and 27, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 215 and 28, respectively. The VL and VH can have the amino acid sequences of SEQ ID NOs: 215 and 29, respectively.

[0129] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising: (a) a VL comprising VL CDRs 1, 2, and 3 derived from a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-10 and 180-188; and / or (b) a VH comprising VH CDRs 1, 2, and 3 derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 11-20 and 189-197. In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising: (a) a VL comprising VL CDRs 1, 2, and 3 derived from a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 21 and 207-215; and / or (b) a VH comprising VH CDRs 1, 2, and 3 derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22-29.

[0130] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL, wherein the VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 1. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 2. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 3. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 4. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 5. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 6. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 7. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 8. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 9. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 10. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 180. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 181. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 182. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 183. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 184. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 185. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 186. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 187. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 188. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 21. The VL comprises VL CDRs 1, 2, and 3 derived from a VL having the amino acid sequence of SEQ ID NO: 207.The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 208. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 209. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 210. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 211. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 212. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 213. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 214. The VL comprises VL CDRs 1, 2, and 3 from a VL having the amino acid sequence of SEQ ID NO: 215.

[0131] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VH, wherein the VH comprises VH CDRs 1, 2, and 3 from a VH having the amino acid sequence of SEQ ID NO: 11. The VH comprises VH CDRs 1, 2, and 3 from a VH having the amino acid sequence of SEQ ID NO: 12. The VH comprises VH CDRs 1, 2, and 3 from a VH having the amino acid sequence of SEQ ID NO: 13. The VH comprises VH CDRs 1, 2, and 3 from a VH having the amino acid sequence of SEQ ID NO: 14. The VH comprises VH CDRs 1, 2, and 3 from a VH having the amino acid sequence of SEQ ID NO: 15. The VH comprises VH CDRs 1, 2, and 3 from a VH having the amino acid sequence of SEQ ID NO: 16. The VH comprises VH CDRs 1, 2, and 3 from a VH having the amino acid sequence of SEQ ID NO: 17. The VH comprises VH CDRs 1, 2, and 3 from a VH having the amino acid sequence of SEQ ID NO: 18. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 19. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 20. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 189. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 190. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 191. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 192. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 193. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 194. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 195. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 196. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 197. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 22. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 23.The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 24. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 25. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 26. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 27. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 28. The VH comprises VH CDRs 1, 2, and 3 derived from a VH having the amino acid sequence of SEQ ID NO: 29.

[0132] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL and a VH, wherein the VL comprises a VL CDR1, CDR2, and CDR3 derived from a VL of a specific sequence, and the VH comprises a VH CDR1, CDR2, and CDR3 derived from a VH of a specific sequence. In some embodiments, the VL CDRs and VH CDRs are derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 1 and 11, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 2 and 12, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 3 and 13, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 4 and 14, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 5 and 15, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 6 and 16, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 7 and 17, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 8 and 18, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 9 and 19, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 10 and 20, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 180 and 189, respectively. The VL CDRs and VH CDRs can be derived from a VL and a VH having the amino acid sequences of SEQ ID NOs: 181 and 190, respectively. The VL CDRs and VH CDRs can be derived from a VL and VH having the amino acid sequences of SEQ ID NOs: 182 and 191, respectively. The VL CDRs and VH CDRs can be derived from a VL and VH having the amino acid sequences of SEQ ID NOs: 183 and 192, respectively.The VL CDRs and VH CDRs can be derived from a VL and VH having the amino acid sequences of SEQ ID NOs: 184 and 193, respectively. The VL CDRs and VH CDRs can be derived from a VL and VH having the amino acid sequences of SEQ ID NOs: 185 and 194, respectively. The VL CDRs and VH CDRs can be derived from a VL and VH having the amino acid sequences of SEQ ID NOs: 186 and 195, respectively. The VL CDRs and VH CDRs can be derived from a VL and VH having the amino acid sequences of SEQ ID NOs: 187 and 196, respectively. The VL CDRs and VH CDRs can be derived from a VL and VH having the amino acid sequences of SEQ ID NOs: 188 and 197, respectively.

[0133] In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to TIM-3, comprising a VL and a VH, wherein the VL comprises VL CDR1, CDR2, and CDR3 derived from a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 21 and 207-215, and the VH comprises VH CDR1, CDR2, and CDR3 derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22-29. The VL CDRs can be derived from a VL having the amino acid sequence of SEQ ID NO: 21, and the VH CDRs can each be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22-29 (e.g., SEQ ID NO: 22). The VL CDRs can be derived from a VL having the amino acid sequence of SEQ ID NO: 207, and the VH CDRs can each be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22-29 (e.g., SEQ ID NO: 22). The VL CDR can be derived from a VL having the amino acid sequence of SEQ ID NO: 208, and the VH CDR can be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29 (e.g., SEQ ID NO: 22). The VL CDR can be derived from a VL having the amino acid sequence of SEQ ID NO: 209, and the VH CDR can be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29 (e.g., SEQ ID NO: 22). The VL CDR can be derived from a VL having the amino acid sequence of SEQ ID NO: 210, and the VH CDR can be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29 (e.g., SEQ ID NO: 22). The VL CDR can be derived from a VL having the amino acid sequence of SEQ ID NO: 211, and the VH CDR can be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29 (e.g., SEQ ID NO: 22). The VL CDRs can be derived from a VL having the amino acid sequence of SEQ ID NO: 212, and the VH CDRs can each be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29 (e.g., SEQ ID NO: 22).The VL CDR can be derived from a VL having the amino acid sequence of SEQ ID NO: 213, and the VH CDR can be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29 (e.g., SEQ ID NO: 22), respectively. The VL CDR can be derived from a VL having the amino acid sequence of SEQ ID NO: 214, and the VH CDR can be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29 (e.g., SEQ ID NO: 22), respectively. The VL CDR can be derived from a VL having the amino acid sequence of SEQ ID NO: 215, and the VH CDR can be derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29 (e.g., SEQ ID NO: 22), respectively.

[0134] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as 3E6. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL (SEQ ID NO: 1) derived from 3E6. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH (SEQ ID NO: 11) derived from 3E6. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from 3E6. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 1) derived from 3E6. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 11) derived from 3E6. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3, respectively, from the VL and VH of 3E6. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 3E6. The 3E6 variants can have a VL that is a variant of the VL of 3E6 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 1. The 3E6 variants can have a VH that is a variant of the VH of 3E6 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 11. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 3E6 variants have up to about five conservative amino acid substitutions. In some embodiments, the variant of 3E6 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 3E6.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 3E6. In some embodiments, provided herein are affinity-matured variants of 3E6 having one, two, or three mutations in the VL CDR3 region as shown in Table 1c.

[0135] As provided below, epitope mapping revealed that 3E6 binds to amino acids 71-82 of human TIM-3, a fragment within the extracellular domain of human TIM-3. Human TIM-3 isoform 1 (SEQ ID NO: 84) and isoform 2 (SEQ ID NO: 85) share the same amino acids 1-131. Thus, 3E6 binds to the same epitope on both isoforms. As used herein, an antibody or antigen-binding fragment that specifically binds to one or more amino acid residues comprising amino acids 1-131, or amino acids 71-82, of human TIM-3 means that the antibody or antigen-binding fragment binds to the listed amino acid residues on both isoform 1 and isoform 2 of human TIM-3.

[0136] Specifically, 3E6 binds to human TIM-3 via CDR1 to CDR3 of VH and CDR2 of VL. As shown in Figure 11B and Table 9 below: the F residue of VH CDR1 (amino acid 3 of SEQ ID NO:101) binds to N76 of human TIM-3; the H residue of VH CDR2 (amino acid 4 of SEQ ID NO:109) binds to W78 of human TIM-3; the S residue of VH CDR2 (amino acid 5 of SEQ ID NO:109) binds to W78 of human TIM-3; the Y residue of VH CDR3 (amino acid 2 of SEQ ID NO:119) binds to D71 of human TIM-3; the R residue of VH CDR3 (amino acid 3 of SEQ ID NO:119) binds to V75, T79, and Y82 of human TIM-3; the S residue of VH CDR3 (amino acid 4 of SEQ ID NO:119) binds to D74 of human TIM-3; the W residue of VH CDR3 (amino acid 6 of SEQ ID NO:119) binds to D74 of human TIM-3; The S residue of CDR2 (amino acid 7 of SEQ ID NO: 94) binds to R73 of human TIM-3.

[0137] Thus, in some embodiments, provided herein are variants of 3E6 having up to about three amino acid substitutions, additions, and / or deletions in the VL CDR and up to about three amino acid substitutions, additions, and / or deletions in the VH CDR. In some embodiments, provided herein are affinity-matured variants of 3E6 having up to about three amino acid substitutions, additions, and / or deletions in the VL CDR and up to about three amino acid substitutions, additions, and / or deletions in the VH CDR. In some embodiments, provided herein are anti-TIM-3 antibodies or antigen-binding fragments comprising VL CDR1, VL CDR2, VH CDR1, VH CDR2, and VH CDR3 having the amino acid sequences of SEQ ID NOs: 86, 94, 101, 109, and 119, respectively, and a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47 and 198-206; or variants thereof having up to about three amino acid substitutions, additions, and / or deletions in the VL CDR and up to about three amino acid substitutions, additions, and / or deletions in the VH CDR.

[0138] In some embodiments of the antibodies or antigen-binding fragments provided herein, the F residue of VH CDR1 (amino acid 3 of SEQ ID NO: 101), the H and S residues of VH CDR2 (amino acids 4 and 5 of SEQ ID NO: 109), the Y, R, S, and W residues of VH CDR3 (amino acids 2, 3, 4, and 6 of SEQ ID NO: 119), and the S residue of VL CDR2 (amino acid 7 of SEQ ID NO: 94) are not mutated. In some embodiments, the 3E6 variant does not have a mutation in at least one of the following sites: the F residue of VH CDR1 (amino acid 3 of SEQ ID NO: 101), the H and S residues of VH CDR2 (amino acids 4 and 5 of SEQ ID NO: 109), the Y, R, S, and W residues of VH CDR3 (amino acids 2, 3, 4, and 6 of SEQ ID NO: 119), and the S residue of VL CDR2 (amino acid 7 of SEQ ID NO: 94). In some embodiments, the 3E6 variant does not have mutations in at least two, at least three, at least four, at least five, at least six, at least seven, or all eight of the following sites: the F residue of VH CDR1 (amino acid 3 of SEQ ID NO: 101), the H and S residues of VH CDR2 (amino acids 4 and 5 of SEQ ID NO: 109), the Y, R, S, and W residues of VH CDR3 (amino acids 2, 3, 4, and 6 of SEQ ID NO: 119), and the S residue of VL CDR2 (amino acid 7 of SEQ ID NO: 94). In some embodiments, the 3E6 variant does not have a mutation in the R residue of VH CDR3 (amino acid 3 of SEQ ID NO: 119).

[0139] In some embodiments, provided herein are humanized 3E6 and affinity-matured humanized 3E6. In some embodiments, the humanized anti-TIM-3 antibody or antigen-binding fragment thereof provided herein comprises a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 21 and 207-215. In some embodiments, the humanized anti-TIM-3 antibody or antigen-binding fragment thereof provided herein comprises a VH having an amino acid sequence selected from SEQ ID NOs: 22-29. In some embodiments, the humanized anti-TIM-3 antibody or antigen-binding fragment thereof provided herein comprises a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 21 and 207-215 and a VH having an amino acid sequence selected from SEQ ID NOs: 22-29. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a variant of the humanized 3E6 provided herein. The variant can have a VL that is a variant of the VL of humanized 3E6, having up to about five amino acid substitutions, additions, and / or deletions in an amino acid sequence selected from the group consisting of SEQ ID NOs: 21 and 207-215. The variant can have a VH that is a variant of the VH of humanized 3E6 having up to about 5 amino acid substitutions, additions, and / or deletions in an amino acid sequence selected from the group consisting of SEQ ID NOs: 22 to 29. In some embodiments, the humanized 3E6 variant has up to about 5 conservative amino acid substitutions.

[0140] In some embodiments, the humanized 3E6 variant has up to three conservative amino acid substitutions. In some embodiments, provided herein is an IgG1 antibody having a VH and VL derived from 3E6 or a 3E6 variant. In some embodiments, the IgG1 heavy chain constant region contains amino acid substitutions L234A and L235A according to EU numbering. In some embodiments, provided herein is an IgG2 antibody having a VH and VL derived from 3E6 or a 3E6 variant. In some embodiments, provided herein is an IgG4 antibody having a VH and VL derived from 3E6 or a 3E6 variant. By way of example, in some embodiments, provided herein is a 3E6 variant comprising VL0 (SEQ ID NO: 21) and VH6 (SEQ ID NO: 28) described herein. In some embodiments, provided herein is a 3E6 variant comprising VL0-v5 (SEQ ID NO: 211) and VH6 (SEQ ID NO: 28) described herein.

[0141] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as 4H2. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL derived from 4H2 (SEQ ID NO: 2). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH derived from 4H2 (SEQ ID NO: 12). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from 4H2. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from the VL derived from 4H2 (SEQ ID NO: 2). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from the VH derived from 4H2 (SEQ ID NO: 12). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of 4H2, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 4H2. The 4H2 variant can have a VL that is a variant of the VL of 4H2 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 2. The 4H2 variant can have a VH that is a variant of the VH of 4H2 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 12. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 4H2 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 4H2 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 4H2.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 4H2.

[0142] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as 16H1. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL derived from 16H1 (SEQ ID NO: 3). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH derived from 16H1 (SEQ ID NO: 13). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from 16H1. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from the VL derived from 16H1 (SEQ ID NO: 3). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from the VH derived from 16H1 (SEQ ID NO: 13). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3, respectively, from the VL and VH of 16H1. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 16H1. The 16H1 variant can have a VL that is a variant of the VL of 16H1 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 3. The 16H1 variant can have a VH that is a variant of the VH of 16H1 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 13. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 16H1 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 16H1 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 16H1.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 16H1.

[0143] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is the antibody designated 18C6. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL (SEQ ID NO: 4) derived from 18C6. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH (SEQ ID NO: 14) derived from 18C6. The anti-TIM-3 antibody or antigen-binding fragment thereof provided herein can have both a VL and a VH derived from 18C6. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 4) derived from 18C6. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 14) derived from 18C6. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3, respectively, from the VL and VH of 18C6. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 18C6. The 18C6 variant can have a VL that is a variant of the VL of 18C6 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO:4. The 18C6 variant can have a VH that is a variant of the VH of 18C6 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO:14. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 18C6 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 18C6 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 18C6.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 18C6.

[0144] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is the antibody designated 19D11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL (SEQ ID NO: 5) derived from 19D11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH (SEQ ID NO: 15) derived from 19D11. The anti-TIM-3 antibody or antigen-binding fragment thereof provided herein can have both a VL and a VH derived from 19D11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 5) derived from 19D11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 15) derived from 19D11. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3, respectively, from the VL and VH of 19D11. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 19D11. The 19D11 variant can have a VL that is a variant of the VL of 19D11 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 5. The 19D11 variant can have a VH that is a variant of the VH of 19D11 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 15. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 19D11 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 19D11 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 19D11.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 19D11.

[0145] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as CH 5#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL derived from CH 5# (SEQ ID NO: 6). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH derived from CH 5# (SEQ ID NO: 16). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from CH 5#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from a VL derived from CH 5# (SEQ ID NO: 6). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from a VH derived from CH 5# (SEQ ID NO: 16). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of CH5#, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of CH5#. The CH5# variants can have a VL that is a variant of the VL of CH5# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 6. The CH5# variants can have a VH that is a variant of the VH of CH5# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 16. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the CH5# variants have up to about five conservative amino acid substitutions. In some embodiments, the variant of CH 5# has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from CH 5#.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from CH 5#.

[0146] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as CH 8#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL derived from CH 8# (SEQ ID NO: 7). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH derived from CH 8# (SEQ ID NO: 17). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from CH 8#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from the VL derived from CH 8# (SEQ ID NO: 7). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from the VH derived from CH 8# (SEQ ID NO: 17). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of CH 8#, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of CH 8#. The CH 8# variants can have a VL that is a variant of the VL of CH 8# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 7. The CH 8# variants can have a VH that is a variant of the VH of CH 8# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 17. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the CH 8# variants have up to about five conservative amino acid substitutions. In some embodiments, the variant of CH 8# has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from CH 8#.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from CH 8#.

[0147] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as CH 9#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL derived from CH 9# (SEQ ID NO: 8). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH derived from CH 9# (SEQ ID NO: 18). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from CH 9#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from a VL derived from CH 9# (SEQ ID NO: 8). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from a VH derived from CH 9# (SEQ ID NO: 18). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of CH9#, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of CH9#. The CH9# variants can have a VL that is a variant of the VL of CH9# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO:8. The CH9# variants can have a VH that is a variant of the VH of CH9# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO:18. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the CH9# variants have up to about five conservative amino acid substitutions. In some embodiments, the variant of CH 9# has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from CH 9#.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from CH 9#.

[0148] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as CH 10#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL derived from CH 10# (SEQ ID NO: 9). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH derived from CH 10# (SEQ ID NO: 19). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from CH 10#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from a VL derived from CH 10# (SEQ ID NO: 9). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from a VH derived from CH 10# (SEQ ID NO: 19). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of CH 10#, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of CH 10#. The CH 10# variants can have a VL that is a variant of the VL of CH 10# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 9. The CH 10# variants can have a VH that is a variant of the VH of CH 10# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 19. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the CH 10# variants have up to about five conservative amino acid substitutions. In some embodiments, the variant of CH 10# has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from CH 10#.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from CH 10#.

[0149] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as CH 11#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL derived from CH 11# (SEQ ID NO: 10). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH derived from CH 11# (SEQ ID NO: 20). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from CH 11#. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from a VL derived from CH 11# (SEQ ID NO: 10). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from a VH derived from CH 11# (SEQ ID NO: 20). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of CH 11#, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of CH 11#. The CH 11# variant can have a VL that is a variant of the VL of CH 11# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 10. The CH 11# variant can have a VH that is a variant of the VH of CH 11# with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 20. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the CH 11# variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of CH 11# has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from CH 11#.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from CH 11#.

[0150] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as 3F2. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL derived from 3F2 (SEQ ID NO: 180). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH derived from 3F2 (SEQ ID NO: 189). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from 3F2. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from the VL derived from 3F2 (SEQ ID NO: 180). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from the VH derived from 3F2 (SEQ ID NO: 189). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of 3F2, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 3F2. The 3F2 variants can have a VL that is a variant of the VL of 3F2 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 180. The 3F2 variants can have a VH that is a variant of the VH of 3F2 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 189. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 3F2 variants have up to about five conservative amino acid substitutions. In some embodiments, the variant of 3F2 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 3F2.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 3F2.

[0151] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is the antibody designated 36A2. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL (SEQ ID NO: 181) derived from 36A2. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH (SEQ ID NO: 190) derived from 36A2. The anti-TIM-3 antibody or antigen-binding fragment thereof provided herein can have both a VL and a VH derived from 36A2. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 181) derived from 36A2. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 190) derived from 36A2. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3, respectively, from the VL and VH of 36A2. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 36A2. The 36A2 variant can have a VL that is a variant of the VL of 36A2 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 181. The 36A2 variant can have a VH that is a variant of the VH of 36A2 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 190. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 36A2 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 36A2 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 36A2.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 36A2.

[0152] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is the antibody designated 36B11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL (SEQ ID NO: 182) derived from 36B11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH (SEQ ID NO: 191) derived from 36B11. The anti-TIM-3 antibody or antigen-binding fragment thereof provided herein can have both a VL and a VH derived from 36B11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 182) derived from 36B11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 191) derived from 36B11. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of 36B11, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 36B11. The 36B11 variant can have a VL that is a variant of the VL of 36B11 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 182. The 36B11 variant can have a VH that is a variant of the VH of 36B11 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 191. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, variants of 36B11 have up to about 5 conservative amino acid substitutions. In some embodiments, variants of 36B11 have up to 3 conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are humanized antibodies or antigen-binding fragments derived from 36B11.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 36B11.

[0153] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is the antibody designated 38F8. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL (SEQ ID NO: 183) derived from 38F8. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH (SEQ ID NO: 192) derived from 38F8. The anti-TIM-3 antibody or antigen-binding fragment thereof provided herein can have both a VL and a VH derived from 38F8. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 183) derived from 38F8. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 192) derived from 38F8. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3, respectively, from the VL and VH of 38F8. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 38F8. The 38F8 variant can have a VL that is a variant of the VL of 38F8 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 183. The 38F8 variant can have a VH that is a variant of the VH of 38F8 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 192. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 38F8 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 38F8 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 38F8.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 38F8.

[0154] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is the antibody designated 38A8. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL (SEQ ID NO: 184) derived from 38A8. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH (SEQ ID NO: 193) derived from 38A8. The anti-TIM-3 antibody or antigen-binding fragment thereof provided herein can have both a VL and a VH derived from 38A8. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 184) derived from 38A8. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 193) derived from 38A8. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3, respectively, from the VL and VH of 38A8. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 38A8. The 38A8 variant can have a VL that is a variant of the VL of 38A8 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 184. The 38A8 variant can have a VH that is a variant of the VH of 38A8 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 193. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 38A8 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 38A8 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 38A8.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 38A8.

[0155] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is the antibody designated 40F11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL (SEQ ID NO: 185) derived from 40F11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH (SEQ ID NO: 194) derived from 40F11. The anti-TIM-3 antibody or antigen-binding fragment thereof provided herein can have both a VL and a VH derived from 40F11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 185) derived from 40F11. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 194) derived from 40F11. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of 40F11, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 40F11. The 40F11 variant can have a VL that is a variant of the VL of 40F11 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 185. The 40F11 variant can have a VH that is a variant of the VH of 40F11 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 194. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, variants of 40F11 have up to about 5 conservative amino acid substitutions. In some embodiments, variants of 40F11 have up to 3 conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are humanized antibodies or antigen-binding fragments derived from 40F11.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 40F11.

[0156] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are antibodies designated as 50H9. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL (SEQ ID NO: 186) derived from 50H9. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH (SEQ ID NO: 195) derived from 50H9. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from 50H9. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 186) derived from 50H9. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 195) derived from 50H9. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of 50H9, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 50H9. The 50H9 variant can have a VL that is a variant of the VL of 50H9 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 186. The 50H9 variant can have a VH that is a variant of the VH of 50H9 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 195. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 50H9 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 50H9 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 50H9.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 50H9.

[0157] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are the antibodies designated 84G10. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL (SEQ ID NO: 187) derived from 84G10. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH (SEQ ID NO: 196) derived from 84G10. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have both a VL and a VH derived from 84G10. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VL comprising VL CDRs 1, 2, and 3 derived from the VL (SEQ ID NO: 187) derived from 84G10. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein have a VH comprising VH CDRs 1, 2, and 3 derived from the VH (SEQ ID NO: 196) derived from 84G10. The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3 from the VL and VH of 84G10, respectively. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 84G10. The 84G10 variant can have a VL that is a variant of the VL of 84G10 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 187. The 84G10 variant can have a VH that is a variant of the VH of 84G10 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 196. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, variants of 84G10 have up to about 5 conservative amino acid substitutions. In some embodiments, variants of 84G10 have up to 3 conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are humanized antibodies or antigen-binding fragments derived from 84G10.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 84G10.

[0158] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is the antibody designated 39D1. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL derived from 39D1 (SEQ ID NO: 188). In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH derived from 39D1 (SEQ ID NO: 197). The anti-TIM-3 antibody or antigen-binding fragment thereof provided herein can have both a VL and a VH derived from 39D1. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VL comprising VL CDRs 1, 2, and 3 derived from the VL derived from 39D1 (SEQ ID NO: 188). In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein has a VH comprising VH CDRs 1, 2, and 3 derived from the VH derived from 39D1 (SEQ ID NO: 197). The anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein can have a VL comprising VL CDRs 1, 2, and 3 and a VH comprising VH CDRs 1, 2, and 3, respectively, derived from the VL and VH of 39D1. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are variants of 39D1. The 39D1 variant can have a VL that is a variant of the VL of 39D1 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 188. The 39D1 variant can have a VH that is a variant of the VH of 39D1 with up to about five amino acid substitutions, additions, and / or deletions in SEQ ID NO: 197. The amino acid substitutions, additions, and / or deletions can be in the VH CDR or the VL CDR. In some embodiments, the amino acid substitutions, additions, and / or deletions are not in the CDRs. In some embodiments, the 39D1 variant has up to about five conservative amino acid substitutions. In some embodiments, the variant of 39D1 has up to three conservative amino acid substitutions. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment thereof provided herein is a humanized antibody or antigen-binding fragment derived from 39D1.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments thereof provided herein are human antibodies or antigen-binding fragments derived from 39D1.

[0159] In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with the antibodies or antigen-binding fragments provided above for binding to TIM-3 (e.g., human TIM-3). An antibody that "competes with another antibody for binding to a target" refers to an antibody that inhibits (partially or completely) the binding of the other antibody to a target. Whether two antibodies compete with each other for binding to a target, i.e., whether one antibody inhibits the binding of the other antibody to a target and the extent to which one antibody inhibits the binding of the other antibody to a target, can be determined using known competition experiments, such as BLI analysis or BIACORE® surface plasmon resonance (SPR) analysis. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment competes with and inhibits the binding of another antibody or antigen-binding fragment to TIM-3 (e.g., human TIM-3) by at least 50%, 60%, 70%, 80%, 90%, or 100%. The level of inhibition or competition can vary depending on which antibody is the "blocking antibody" (i.e., the cold antibody that is first incubated with the target). Competition assays can be performed, for example, as described in Ed Harlow and David Lane, Cold Spring Haib Protoc; 2006; doi: 10.1101 / pdb.prot 4277, or Chapter 11 of "Using Antibodies," Ed Harlow and David Lane, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA 1999. Two antibodies "cross-compete" if they block each other by at least 50% in both directions, i.e., regardless of whether one or the other antibody first contacts the antigen in the competition experiment.

[0160] Competitive binding assays to determine whether two antibodies compete or cross-compete for binding include, for example, competition for binding to TIM-3-expressing T cells by, e.g., flow cytometry, as described in the Examples. Other methods include biolayer interferometry (BLI), SPR (e.g., BIACORE®), solid-phase direct or indirect radioimmunoassay (RIA), solid-phase direct or indirect enzyme immunoassay (EIA), sandwich competition assay (see Stahli et al., Methods in Enzymology 9:242 (1983)); solid-phase direct biotin-avidin EIA (see Kirkland et al., J. Immunol. 137:3614 (1986)); solid-phase direct label assay, solid-phase direct label sandwich assay (see Harlow and Lane, ANTIBODIES: A LABORATORY MANUAL, Cold Spring Harbor Press (1988)); solid-phase direct label RIA using I-125 label (see Morel et al., J. Immunol. 25(1):7 (1988)); solid-phase direct biotin-avidin EIA (Cheung et al., Virology 176:546 (1990)); and direct label RIA (Moldenhauer et al., Scand. J. Immunol. 32:77 (1990)).

[0161] In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 3E6 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 4H2 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 16H1 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 18C6 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 19D11 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with CH5# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with CH 8# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with CH 9# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with CH 10# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with CH 11# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with 3F2 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with 36A2 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with 36B11 for binding to TIM-3 (eg, human TIM-3).In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 38F8 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 38A8 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 40F11 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 50H9 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 84G10 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with 39D1 for binding to TIM-3 (e.g., human TIM-3).

[0162] In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 3E6 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 4H2 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 16H1 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 18C6 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 19D11 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized CH5# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized CH 8# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized CH 9# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized CH 10# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized CH 11# for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 3F2 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 36A2 for binding to TIM-3 (e.g., human TIM-3).In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 36B11 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 38F8 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 38A8 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 40F11 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 50H9 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein is an antibody or antigen-binding fragment that competes with humanized 84G10 for binding to TIM-3 (e.g., human TIM-3). In some embodiments, provided herein are antibodies or antigen-binding fragments that compete with humanized 39D1 for binding to TIM-3 (eg, human TIM-3).

[0163] In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as the antibodies or antigen-binding fragments provided above. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 3E6 or humanized 3E6. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 4H2 or humanized 4H2. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 16H1 or humanized 16H1. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 18C6 or humanized 18C6. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 19D11 or humanized 19D11. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as CH 5# or humanized CH 5#. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as CH 8# or humanized CH 8#. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as CH 9# or humanized CH 9#. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as CH 10# or humanized CH 10#.In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as CH11# or humanized CH11#. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 3F2 or humanized 3F2. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 36A2 or humanized 36A2. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 36B11 or humanized 36B11. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 38F8 or humanized 38F8. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 38A8 or humanized 38A8. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 40F11 or humanized 40F11. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 50H9 or humanized 50H9. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 84G10 or humanized 84G10. In some embodiments, provided herein are antibodies or antigen-binding fragments that bind to the same epitope on TIM-3 (e.g., human TIM-3) as 39D1 or humanized 39D1.

[0164] The present disclosure further contemplates additional variants and equivalents that are substantially homologous to the recombinant, monoclonal, chimeric, humanized, and human antibodies described herein, or antibody fragments thereof. In some embodiments, it is desirable to improve the binding affinity of the antibody. In some embodiments, it is desirable to modulate the biological properties of the antibody, including, but not limited to, specificity, thermostability, expression level, effector function, glycosylation, immunogenicity, and / or solubility. Those skilled in the art will understand that amino acid changes can alter post-translational processing of the antibody, for example, by changing the number or location of glycosylation sites or altering membrane anchoring characteristics.

[0165] Mutations can be substitutions, deletions, or insertions of one or more nucleotides encoding the antibody or polypeptide, resulting in a change in the amino acid sequence compared to the native antibody or polypeptide sequence. In some embodiments, the amino acid substitutions result in the replacement of one amino acid with another amino acid having similar structural and / or chemical properties, e.g., conservative amino acid substitutions, such as a leucine for a serine. Insertions or deletions can range from about 1 to 5 amino acids. In some embodiments, substitutions, deletions, or insertions comprise fewer than 25 amino acid substitutions, fewer than 20 amino acid substitutions, fewer than 15 amino acid substitutions, fewer than 10 amino acid substitutions, fewer than 5 amino acid substitutions, fewer than 4 amino acid substitutions, fewer than 3 amino acid substitutions, or fewer than 2 amino acid substitutions compared to the parent molecule. In some embodiments, biologically useful and / or relevant amino acid sequence variations can be determined by systematically making insertions, deletions, or substitutions in the sequence and testing the resulting mutant proteins for activity compared to the parent protein.

[0166] It is known in the art that the constant region of an antibody mediates several effector functions, and these effector functions may vary depending on the antibody isotype. For example, the binding of the C1 component of complement to the Fc region of an IgG or IgM antibody (bound to an antigen) activates the complement system. Complement activation is important for the opsonization and lysis of cellular pathogens. Complement activation also stimulates the inflammatory response and can be involved in autoimmune hypersensitivity. Furthermore, the Fc region of an antibody can bind to cells expressing Fc receptors (FcRs). There are several Fc receptors specific for different classes of antibodies, including IgG (gamma receptors), IgE (epsilon receptors), IgA (alpha receptors), and IgM (mu receptors). Binding of antibodies to cell surface Fc receptors elicits several important and diverse biological responses, including the uptake and destruction of antibody-coated particles, clearance of immune complexes, lysis of antibody-coated target cells by killer cells (called antibody-dependent cellular cytotoxicity or ADCC), antibody-dependent cellular phagocytosis (ADCP), release of inflammatory mediators, placental transfer, and regulation of immunoglobulin production. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgA antibody. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgD antibody. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgE antibody. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgG antibody. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgM antibody. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgG1 antibody. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgG2 antibody.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgG3 antibody. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise at least one constant region of a human IgG4 antibody.

[0167] Also provided are engineered, modified antibodies that can be prepared using antibodies having one or more of the VH and / or VL sequences disclosed herein as starting materials for engineering the modified antibodies, which can have altered properties from the starting antibody. The antibodies can be engineered by modifying one or more residues in one or both variable regions (i.e., VH and / or VL), for example, in one or more CDR regions and / or one or more framework regions. Additionally or alternatively, the antibodies can be engineered by modifying residues in the constant region, for example, to alter the effector function of the antibody. One type of variable region engineering that can be performed is CDR grafting. Antibodies interact with target antigens primarily through amino acid residues located in the six CDRs. Because the CDR sequences are responsible for most antibody-antigen interactions, recombinant antibodies that mimic the properties of a particular naturally occurring antibody can be expressed by constructing expression vectors containing the CDR sequences from that particular naturally occurring antibody grafted onto framework sequences from a different antibody with different properties (see, e.g., Riechmann, L. et al. (1998) Nature 332:323-327; Jones, P. et al. (1986) Nature 321:522-525; Queen, C. et al. (1989) Proc. Natl. Acad Sci. USA 86:10029-10033; Winter, U.S. Pat. No. 5,225,539, and Queen et al., U.S. Pat. Nos. 5,530,101; 5,585,089; 5,693,762, and 6,180,370).

[0168] Accordingly, some embodiments described herein relate to an isolated monoclonal antibody, or antigen-binding portion thereof, comprising: a light chain variable region comprising CDR1, CDR2, and CDR3 sequences, and comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-10, 21, 180-188, and 207-215, respectively; and a heavy chain variable region comprising CDR1, CDR2, and CDR3 sequences, and comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 11-20, 22-29, and 189-197, respectively. Thus, such antibodies contain the VH and VL CDR sequences of monoclonal antibodies 3E6, 4H2, 16H1, 18C6, 19D11, CH5#, CH8#, CH9#, CH10#, CH11#, 3F2, 36A2, 36B11, 38F8, 38A8, 40F11, 50H9, 84G10, or 39D1, and can further contain different framework sequences from these antibodies.

[0169] Such framework sequences can be obtained from public DNA databases or published references that contain germline antibody gene sequences. For example, germline DNA sequences for human heavy and light chain variable region genes can be found in the "Vbase" human germline sequence database (available on the Internet at www.mrc-cpe.cam.ac.uk / vbase), as well as in Kabat, E. A. et al. (1991), Sequences of Proteins of Immunological Interest, 5th ed., US Department of Health and Human Services, NIH Publication No. 91-3242; Tomlinson, I. M. et al. (1992) Mol. Biol. 227:776-798; and Cox, J. P. L. et al. (1994), Eur. J. Immunol. 24:827-836, the contents of each of which are expressly incorporated herein by reference.

[0170] In some embodiments, framework sequences for use in the anti-TIM-3 antibodies or antigen-binding fragments described herein are structurally similar to the framework sequences used by the anti-TIM-3 antibodies described herein. The VH CDR1, 2, and 3 sequences and the VL CDR1, 2, and 3 sequences can be grafted into framework regions having sequences identical to those found in the germline immunoglobulin gene from which the framework sequences are derived, or the CDR sequences can be grafted into framework regions containing one or more mutations compared to the germline sequences. For example, in some cases, it has been found beneficial to mutate residues within the framework regions to maintain or enhance the antigen-binding ability of the antibody (see, e.g., U.S. Patent Nos. 5,530,101; 5,585,089; 5,693,762; and 6,180,370 to Queen et al.).

[0171] The modified anti-TIM-3 antibodies or antigen-binding fragments described herein include those in which modifications have been made to framework residues within the VH and / or VL, for example, to improve the properties of the antibody. Typically, such framework modifications are made to reduce the immunogenicity of the antibody. For example, one approach is to "backmutate" one or more framework residues to the corresponding germline sequence. More specifically, an antibody that has undergone somatic mutation may contain framework residues that differ from the germline sequence from which the antibody is derived. Such residues can be identified by comparing the antibody framework sequence to the germline sequence from which the antibody is derived. To return the framework region sequences to their germline configuration, somatic mutations can be "backmutated" to the germline sequence, for example, by site-directed mutagenesis or PCR-mediated mutagenesis. Such "backmutated" antibodies are also intended to be encompassed. Another type of framework modification involves mutating one or more residues within the framework region, or even one or more CDR regions, to remove T cell epitopes, thereby reducing the potential immunogenicity of the antibody. This approach, also referred to as "deimmunization," is described in further detail in US Patent Publication No. 20030153043 by Carr et al.

[0172] Another type of variable region modification is to mutate amino acid residues within the CDR1, CDR2, and / or CDR3 regions of VH and / or VL, thereby improving one or more binding characteristics (e.g., affinity) of the antibody of interest. Site-directed mutagenesis or PCR-mediated mutagenesis can be performed to introduce the mutations, and the effect on antibody binding or other functional properties of interest can be assessed using in vitro or in vivo assays described herein and provided in the Examples. In some embodiments, conservative modifications (discussed above) are introduced. Mutations can be amino acid substitutions, additions, or deletions. Furthermore, typically, no more than one, two, three, four, or five residues within the CDR regions are altered.

[0173] Methionine residues in the CDRs of antibodies can be oxidized, resulting in potential chemical degradation and a resulting reduction in antibody potency. Therefore, anti-TIM-3 antibodies or antigen-binding fragments are also provided in which one or more methionine residues in the heavy and / or light chain CDRs are substituted with amino acid residues that are not susceptible to oxidative degradation. In some embodiments, the methionine residues in the CDRs of antibodies 3E6, 4H2, 16H1, 18C6, 19D11, CH5#, CH8#, CH9#, CH10#, CH11#, 3F2, 36A2, 36B11, 38F8, 38A8, 40F11, 50H9, 84G10, or 39D1 are substituted with amino acid residues that are not susceptible to oxidative degradation. Similarly, deamidation sites can be removed from anti-TIM-3 antibodies or antigen-binding fragments, particularly in the CDRs.

[0174] The anti-TIM-3 variable regions described herein include Fc, e.g., IgG1, IgG2, IgG3, or IgG4 Fc (which may be of any allotype or isoallotype, e.g., for IgG1: G1m, G1m1(a), G1m2(x), G1m3(f), G1m17(z); for IgG2: G2m, G2m23(n); for IgG3: G3m, G3m21(g1), G3m28(g5), G3m11(b0), G3m5(b1), G3m13(b3), G3m14(b4), G3m10(b5), G3m15(s), G3m16(t), G3m6(c3), G3m24(c5), G3m26(u), G3m27(v); and for K: The antibody can be linked (e.g., covalently linked or fused) to a mAb (which can be Km, Km1, Km2, Km3) (see, e.g., Jefferies et al. (2009) mAbs 1:1).

[0175] In some embodiments, the anti-TIM-3 variable regions described herein are linked to an Fc with no or little effector function, such as an IgG4.

[0176] Generally, the variable regions described herein can be linked to an Fc region that typically includes one or more modifications to alter one or more functional properties of the antibody, such as serum half-life, complement fixation, Fc receptor binding, and / or antigen-dependent cellular cytotoxicity. Additionally, the antibodies described herein can be chemically modified (e.g., one or more chemical moieties can be attached to the antibody) or modified to alter its glycosylation to alter one or more functional properties of the antibody. Each of these embodiments is described in further detail below. The numbering of residues in the Fc region is that of the EU index of Kabat.

[0177] The Fc region encompasses domains derived from immunoglobulin constant regions, including fragments, analogs, variants, mutants, or derivatives of the constant region. Suitable immunoglobulins include IgG1, IgG2, IgG3, IgG4, and other classes, such as IgA, IgD, IgE, and IgM. The constant region of an immunoglobulin is defined as a naturally occurring or synthetically produced polypeptide homologous to the C-terminal region of an immunoglobulin and can include a CH1 domain, hinge, CH2 domain, CH3 domain, or CH4 domain, either separately or in combination. In some embodiments, at least one or more of the constant regions are modified or deleted in the anti-TIM-3 antibodies or antigen-binding fragments described herein. In some embodiments, the antibodies include modifications to one or more of the three heavy chain constant regions (CH1, CH2, or CH3) and / or to the light chain constant region (CL). In some embodiments, the heavy chain constant region of the modified antibody comprises at least one human constant region. In some embodiments, the heavy chain constant region of the modified antibody comprises multiple human constant regions. In some embodiments, modifications to the constant region comprise the addition, deletion, or substitution of one or more amino acids in one or more regions. In some embodiments, one or more regions are partially or completely deleted from the constant region of the modified antibody. In some embodiments, the entire CH2 domain is removed from the antibody (ΔCH2 construct). In some embodiments, the deleted constant region is replaced with a short amino acid spacer that provides some of the molecular flexibility normally conferred by the absent constant region. In some embodiments, the modified antibody comprises a CH3 domain fused directly to the hinge region of the antibody. In some embodiments, the modified antibody comprises a peptide spacer inserted between the hinge region and the modified CH2 and / or CH3 domain.

[0178] Ig molecules interact with multiple classes of cellular receptors. For example, IgG molecules interact with three classes of Fcγ receptors (FcγRs), namely, FcγRI, FcγRII, and FcγRIII, which are specific for the IgG class of antibody. It has been reported that sequences important for binding between IgG and FcγR receptors reside in the CH2 and CH3 domains. The serum half-life of an antibody is influenced by its ability to bind to an Fc receptor (FcR). In some embodiments, an anti-TIM-3 antibody or antigen-binding fragment comprises an Fc region. In some embodiments, the Fc region is fused via a hinge. The hinge can be an IgG1 hinge, an IgG2 hinge, or an IgG3 hinge. The amino acid sequences of the Fc regions of human IgG1, IgG2, IgG3, and IgG4 are known to those skilled in the art. In some examples, Fc regions with amino acid mutations have been identified in native antibodies. In some embodiments, the modified antibody (e.g., modified Fc region) provides altered effector function, which in turn affects the biological profile of the antibody. For example, in some embodiments, deletion or inactivation of the constant region (by point mutation or other means) reduces Fc receptor binding when the modified antibody circulates. In some embodiments, the constant region modification reduces the immunogenicity of the antibody. In some embodiments, the constant region modification increases the serum half-life of the antibody. In some embodiments, the constant region modification reduces the serum half-life of the antibody. In some embodiments, the constant region modification reduces or eliminates the ADCC, ADCP, and / or complement-dependent cytotoxicity (CDC) of the antibody. In some embodiments, specific amino acid substitutions in the human IgG1 Fc region with corresponding IgG2 or IgG4 residues reduce effector functions (e.g., ADCC, ADCP, and CDC) in the modified antibody. In some embodiments, the antibody does not have one or more effector functions (e.g., an "effectorless" antibody). In some embodiments, the antibody does not have ADCC activity and / or CDC activity. In some embodiments, the antibody does not bind to Fc receptors and / or complement factors. In some embodiments, the antibody does not have effector function.In some embodiments, the constant region modifications increase or enhance the ADCC, ADCP, and / or CDC of the antibody. In some embodiments, the constant region is modified to remove disulfide bonds or oligosaccharide moieties. In some embodiments, the constant region is modified to add / substitute one or more amino acids to provide one or more cytotoxin, oligosaccharide, or carbohydrate attachment sites. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment comprises a variant Fc region that is altered with substitutions at specific amino acid positions compared to the native Fc region.

[0179] In some embodiments, the Fc region is a variant Fc region, e.g., an Fc sequence, that has been modified (e.g., by amino acid substitution, deletion, and / or insertion) relative to a parent Fc sequence (e.g., an unmodified Fc polypeptide that is subsequently modified to create the variant) to provide desired structural characteristics and / or biological activity. Typically, variants of a constant region or portion thereof, e.g., a CH1, CL, hinge, CH2, or CH3 domain, can contain 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more mutations, and / or at most 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 mutation, or 1-10 or 1-5 mutations, or comprise an amino acid sequence that is at least about 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of the corresponding wild-type region or domain (CH1, CL, hinge, CH2, or CH3 domain, respectively), provided that the heavy chain constant region containing the particular variant retains the required biological activity.

[0180] For example, modifications can be made in the Fc region to generate Fc variants that, compared to the parent Fc, have (a) increased or decreased antibody-dependent cell-mediated cytotoxicity (ADCC), (b) increased or decreased antibody-dependent cell-mediated phagocytosis (ACDP), (c) increased or decreased complement-mediated cytotoxicity (CDC), (d) increased or decreased affinity for C1q, and / or (e) increased or decreased affinity for Fc receptors. Such Fc region variants typically contain at least one amino acid modification in the Fc region. Combinations of amino acid modifications may be particularly desirable. For example, a variant Fc region can contain substitutions therein, e.g., at two, three, four, five, etc., of the specific Fc region positions identified herein.

[0181] The variant Fc region can also contain sequence modifications in which amino acids involved in disulfide bond formation are removed or replaced with other amino acids. Such removal can avoid reaction with other cysteine-containing proteins present in the host cells used to produce the anti-TIM-3 antibodies or antigen-binding fragments described herein. Even when cysteine ​​residues are removed, the single-chain Fc domain can still form a non-covalently held dimeric Fc domain. In some embodiments, the Fc region can be modified to make it more compatible with a selected host cell. For example, the PA sequence near the N-terminus of a typical native Fc region, which can be recognized by E. coli digestive enzymes such as proline iminopeptidase, can be removed. In some embodiments, one or more glycosylation sites within the Fc domain can be removed. Typically glycosylated residues (e.g., asparagine) can confer a cytolytic response. Such residues can be deleted or substituted with non-glycosylated residues (e.g., alanine). In some embodiments, sites involved in complement interaction, such as the C1q binding site, can be removed from the Fc region. For example, the EKK sequence of human IgG1 can be deleted or replaced. In some embodiments, sites affecting binding to Fc receptors, preferably sites other than the salvage receptor binding site, can be removed. In some embodiments, the Fc region can be modified to remove the ADCC site. In some embodiments, the Fc region can be modified to remove the ADCP site. ADCC sites and ADCP sites are known in the art; for example, see Molec. Immunol. 29(5): 633-9 (1992) for ADCC sites in IgG1, and Herbrand, U. (2016). BioProcessing, 15(1), 1538-8786 for ADCP sites in IgG1. Specific examples of mutant Fc domains are disclosed, for example, in WO 97 / 34631 and WO 96 / 32478.

[0182] In some embodiments, the hinge region of an Fc is modified to change, e.g., increase or decrease, the number of cysteine ​​residues in the hinge region. This approach is further described in U.S. Patent No. 5,677,425 by Bodmer et al. The number of cysteine ​​residues in the hinge region of an Fc is altered, for example, to facilitate association of the light and heavy chains or to increase or decrease the stability of the antibody. In some embodiments, the Fc hinge region of an antibody is mutated to decrease the biological half-life of the antibody. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc-hinge fragment so that the antibody has impaired Staphylococcus aureus protein A (SpA) binding compared to native Fc hinge domain SpA binding. This approach is further described in U.S. Patent No. 6,165,745 by Ward et al.

[0183] In some embodiments, the Fc region is modified by replacing at least one amino acid residue with a different amino acid residue to alter the effector function of the antibody. For example, one or more amino acids selected from amino acid residues 234, 235, 236, 237, 297, 318, 320, 322, 330, and / or 331 can be substituted with a different amino acid residue so that the antibody has altered affinity for an effector ligand but retains the antigen-binding ability of the parent antibody. The effector ligand for which affinity is altered can be, for example, an Fc receptor or the C1 component of complement. This approach is described in further detail in U.S. Patent Nos. 5,624,821 and 5,648,260, both by Winter et al.

[0184] In another example, one or more amino acids selected from amino acid residues 329, 331, and 322 can be substituted with a different amino acid residue such that the antibody has altered C1q binding and / or reduced or abolished complement dependent cytotoxicity (CDC). This approach is described in further detail in U.S. Patent No. 6,194,551 by Idusogie et al.

[0185] In another example, one or more amino acid residues within amino acid positions 231 and 239 are altered to thereby alter the ability of the antibody to fix complement. This approach is further described in PCT Publication WO 94 / 29351 by Bodmer et al.

[0186] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein comprise an IgG1 heavy chain constant region comprising one or more amino acid substitutions selected from the group consisting of K214R, L234A, L235E, G237A, D356E, and L358M according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises one or more amino acid substitutions selected from the group consisting of K214R, L234A, L234F, L235A, L235E, G236R, G237A, D265A, N297A, N297Q, N297G, E318A, L328R, P329G, A330S, P331S, D356E, and L358M according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises one or more amino acid substitutions selected from the group consisting of K214R, C226S, C229S, and P238S according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises one or more amino acid substitutions selected from the group consisting of K214R, D356E, and L358M according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises one or more amino acid substitutions selected from the group consisting of S131C, K133R, G137E, G138S, Q196K, I199T, N203D, K214R, C226S, C229S, and P238S according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises an amino acid substitution selected from the group consisting of N297A, N297Q, and N297G according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises amino acid substitutions of L234A and L235A according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises amino acid substitutions of G236R and L328R according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises amino acid substitutions of L234F, L235E, and P331S according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises amino acid substitutions of L234A, L235A, and P329G according to EU numbering. In some embodiments, the IgG1 heavy chain constant region comprises amino acid substitutions of L234F, L235E, and D265A according to EU numbering.

[0187] In some embodiments, the Fc region comprises a nucleotide sequence selected from the group consisting of: 234, 235, 236, 237, 238, 239, 240, 241, 243, 244, 245, 247, 248, 249, 252, 254, 255, 256, 258, 262, 263, 264, 265, 267, 268, 269, 270, 272, 276, 278, 280, 283, 285, 286, 289, 290, 292, 293, 294, 295, 296, 297, 298, 299, 301, 303, 305, 307, 309, 312, 313, 315, 318, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 358, 359, 360, 361, 362, 363, 364, 365, Modifications can be made to decrease ADCC, ADCP, and / or decrease affinity for Fcγ receptors by modifying one or more amino acids at positions 2, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 337, 338, 340, 360, 373, 376, 378, 382, ​​388, 389, 398, 414, 416, 419, 430, 433, 434, 435, 436, 437, 438, or 439. Exemplary substitutions include 234A, 235A, 236A, 239D, 239E, 268D, 267E, 268E, 268F, 324T, 332D, 332E, and any combination thereof. Exemplary variants include 234A / 235A, 239D / 332E, 236A / 332E, 236A / 239D / 332E, 268F / 324T, 267E / 268F, 267E / 324T, and 267E / 268F7324T. Other modifications to enhance FcγR and complement interactions include, but are not limited to, the substitutions 298A, 333A, 334A, 326A, 247I, 339D, 339Q, 280H, 290S, 298D, 298V, 243L, 292P, 300L, 396L, 305I, and 396L. These and other modifications are reviewed in Strohl, 2009, Current Opinion in Biotechnology 20:685-691.

[0188] Fc modifications that increase binding to Fc receptors include those at amino acid positions 238, 239, 248, 249, 252, 254, 255, 256, 258, 265, 267, 268, 269, 270, 272, 279, 280, 283, 285, 298, 289, 290, 292, 293, 294, 295, 296, 298, 301, 303, 305, 307, 312, 315, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, ​​383, 384, 385, 386, 387, 388 and / or 439, wherein the numbering of residues in the Fc region is that of the EU index as found in abat (WO 00 / 42072).

[0189] Other Fc modifications that can be made to the Fc are those that reduce or eliminate binding to FcγRs and / or complement proteins, thereby reducing or eliminating Fc-mediated effector functions such as ADCC, ADCP, and CDC. Exemplary modifications include, but are not limited to, substitutions, insertions, and deletions at positions 234, 235, 236, 237, 267, 269, 325, 328, 330, and / or 331 (e.g., 330 and 331) (where numbering is according to the EU index). Exemplary substitutions include, but are not limited to, 234A, 235E, 236R, 237A, 267R, 269R, 325L, 328R, 330S, and 331S (e.g., 330S and 331S) (where numbering is according to the EU index). An Fc variant can include 236R / 328R. Other modifications to reduce FcγR and complement interactions include substitutions of 297A, 234A, 235A, 237A, 318A, 228P, 236E, 268Q, 309L, 330S, 331S, 220S, 226S, 229S, 238S, 233P, and 234V, as well as removal of glycosylation at position 297 by mutational or enzymatic means or by production in organisms such as bacteria that do not glycosylate proteins. These and other modifications are reviewed in Strohl, 2009, Current Opinion in Biotechnology 20:685-691.

[0190] Optionally, the Fc region can include non-naturally occurring amino acid residues at additional and / or alternative positions known to those of skill in the art (e.g., U.S. Patent Nos. 5,624,821; 6,277,375; 6,737,056; 6,194,551; 7,317,091; 8,101,720; PCT Patent Publications WO 00 / 42072; WO 01 / 58957; WO 02 / 06919; WO 04 / 016750; WO 04 / 029207; WO 04 / 035752; WO 04 / 074455; WO 04 / 099249; WO 04 / 063351; WO 05 / 070963; WO (See WO No. 05 / 040217, WO No. 05 / 092925 and WO No. 06 / 020114).

[0191] Fc variants that enhance affinity for the inhibitory receptor FcγRIIb can also be used. Such variants can provide Fc fusion proteins with immunomodulatory activity associated with FcγRIIb cells, including, for example, B cells and monocytes. In some embodiments, the Fc variants provide selectively enhanced affinity for FcγRIIb relative to one or more activating receptors. Modifications to alter binding to FcγRIIb include one or more modifications at positions selected from the group consisting of 234, 235, 236, 237, 239, 266, 267, 268, 325, 326, 327, 328, 330, 331, and 332 according to the EU index. Exemplary substitutions for enhancing FcγRIIb affinity include, but are not limited to, 234A, 234D, 234E, 234F, 234W, 235A, 235D, 235E, 235F, 235R, 235Y, 236D, 236N, 237A, 237D, 237N, 239D, 239E, 266M, 267D, 267E, 268D, 268E, 327D, 327E, 328F, 328W, 328Y, 330S, 331S, and 332E. Exemplary substitutions include 235Y, 236D, 239D, 266M, 267E, 268D, 268E, 328F, 328W, and 328Y. Other Fc variants for enhancing binding to FcγRIIb include 235Y / 267E, 236D / 267E, 239D / 268D, 239D / 267E, 267E / 268D, 267E / 268E, and 267E / 328F.

[0192] The affinity and binding properties of an Fc region for its ligand can be determined by a variety of in vitro assay methods (biochemical or immunological-based assays) known in the art, including, but not limited to, equilibrium methods (e.g., enzyme-linked immunosorbent assay (ELISA), biolayer interferometry (BLI), or radioimmunoassay (RIA)), or kinetics (e.g., BIACORE analysis), and other methods such as indirect binding assays, competitive inhibition assays, fluorescence resonance energy transfer (FRET), gel electrophoresis, and chromatography (e.g., gel filtration). These and other methods can utilize labels on one or more components being tested and / or can utilize a variety of detection methods, including, but not limited to, chromogenic, fluorescent, luminescent, or isotopic labels. A detailed discussion of binding affinity and kinetics can be found in Fundamental Immunology, 4th ed., Paul, W. E. (ed.), Lippincott-Raven, Philadelphia (1999), which focuses on antibody-immunogen interactions.

[0193] In some embodiments, the antibody is modified to extend its biological half-life. Various approaches are possible. For example, this can be achieved by increasing the binding affinity of the Fc region to FcRn. For example, one or more of the following residues can be mutated as described in U.S. Patent No. 6,277,375: 252, 254, 256, 433, 435, and 436. Specific exemplary substitutions include one or more of the following: T252L, T254S, and / or T256F. Alternatively, to extend biological half-life, the antibody can be engineered within the CH or CL region to contain a salvage receptor binding epitope obtained from two loops of the CH2 domain of the IgG Fc region, as described in U.S. Patent Nos. 5,869,046 and 6,121,022 by Presta et al. Other exemplary variants that increase binding to FcRn and / or improve pharmacokinetic properties include substitutions at positions 259, 308, 428, and 434, including, for example, 259I, 308F, 428L, 428M, 434S, 434I, 434F, 434Y, and 434X1.Other variants that enhance binding to FcRn include 250E, 250Q, 428L, 428F, 250Q / 428L (Hinton et al., 2004, J. Biol. Chem. 279(8): 6213-6216, Hinton et al., 2006 Journal of Immunology 176:346-356), 256A, 272A, 286A, 305A, 307A, 307Q, 311A, 312A, 376A, 378Q, 380A, 382A, and 434A (Shields et al., Journal of Biological Chemistry, 2001). 276(9):6591-6604), 252F, 252T, 252Y, 252W, 254T, 256S, 256R, 256Q, 256E, 256D, 256T, 309P, 311S, 433R, 433S, 433I, 433P, 433Q, 434H, 434F, 434Y, 252Y / 254T / 256E, 433K / 434F / 436H, 308T / 309P / 311S (Dali Acqua et al., Journal of Immunology, 2002, 169:5171-5180; Dall Acqua et al., Journal of Biological Chemistry, 2006, 281:23514-23524). Other modifications to modulate FcRn binding are described in Yeung et al., 2010, J Immunol, 182:7663-7671.

[0194] In some embodiments, hybrid IgG isotypes with specific biological characteristics can be used. For example, IgG1 / IgG3 hybrid variants can be constructed by substituting IgG1 positions in the CH2 and / or CH3 regions with amino acids from IgG3 at positions that differ between the two isotypes. Thus, hybrid variant IgG antibodies can be constructed that contain one or more substitutions, for example, 274Q, 276K, 300F, 339T, 356E, 358M, 384S, 392N, 397M, 422I, 435R, and 436F. In some embodiments described herein, IgG1 / IgG2 hybrid variants can be constructed by substituting IgG2 positions in the CH2 and / or CH3 regions with amino acids from IgG1 at positions that differ between the two isotypes. Thus, hybrid variant IgG antibodies can be constructed that contain one or more substitutions, for example, one or more of the following amino acid substitutions: 233E, 234L, 235L, -236G (representing the insertion of glycine at position 236), and 327A.

[0195] Furthermore, the binding sites on human IgG1 for FcγRI, FcγRII, FcγRIII, and FcRn have been mapped, and mutants with improved binding have been described (see Shields, RL et al. (2001) J. Biol. Chem. 276:6591-6604). Specific mutations at positions 256, 290, 298, 333, 334, and 339 have been shown to improve binding to FcγRIII. Furthermore, the following combination mutants: T256A / S298A, S298A / E333A, S298A / K224A, and S298A / E333A / K334A have been shown to improve FcγRIII binding, and they have been shown to exhibit enhanced FcγRIIIa binding and ADCC activity (Shields et al., 2001). Other IgG1 mutants with strongly enhanced binding to FcγRIIIa have been identified, including mutants with S239D / I332E and S239D / I332E / A330L mutations, which showed the greatest increase in affinity for FcγRIIIa, reduced FcγRIIb binding, and strong cytotoxic activity in cynomolgus monkeys (Lazar et al., 2006). Introduction of triple mutations into antibodies such as alemtuzumab (CD52-specific), trastuzumab (HER2 / neu-specific), rituximab (CD20-specific), and cetuximab (EGFR-specific) led to greatly enhanced ADCC activity in vitro, and the S239D / I332E mutant showed enhanced ability to deplete B cells in monkeys (Lazar et al., 2006). Furthermore, IgG1 mutants containing the L235V, F243L, R292P, Y300L, and P396L mutations have been identified that showed enhanced binding to FcγRIIIa and concomitant enhanced ADCC activity in transgenic mice expressing human FcγRIIIa in models of B cell malignancies and breast cancer (Stavenhagen et al., 2007; Nordstrom et al., 2011). Other Fc mutants that can be used include: S298A / E333A / L334A, S239D / I332E, S239D / I332E / A330L, L235V / F243L / R292P / Y300L / P396L, and M428L / N434S.

[0196] In some embodiments, an Fc is selected that has reduced binding to FcγRs. An exemplary Fc with reduced FcγR binding, e.g., an IgG1 Fc, contains the following three amino acid substitutions: L234A, L235E, and G237A.

[0197] In some embodiments, an Fc is selected that has reduced complement fixation. An exemplary Fc with reduced complement fixation, e.g., an IgG1 Fc, has the following two amino acid substitutions: A330S and P331S.

[0198] In some embodiments, an Fc is selected that has essentially no effector function, i.e., it has reduced binding to FcγRs and reduced complement fixation. An exemplary effector-less Fc, e.g., an IgG1 Fc, contains the following five mutations: L234A, L235E, G237A, A330S, and P331S.

[0199] When an IgG4 constant domain is used, it can contain the substitution S228P, which mimics the hinge sequence in IgG1, thereby stabilizing the IgG4 molecule. In some embodiments, the IgG4 constant domain contains the substitutions S228P and L235E.

[0200] In some embodiments, the glycosylation of an antibody is modified. For example, an aglycosylated antibody can be generated (i.e., the antibody lacks glycosylation). Glycosylation can be altered, for example, to increase the affinity of the antibody for an antigen. Such carbohydrate modifications can be achieved, for example, by altering one or more sites of glycosylation within the antibody sequence. For example, one or more amino acid substitutions can be made that result in the elimination of one or more variable region framework glycosylation sites, thereby eliminating glycosylation at that site. Such aglycosylation can increase the affinity of the antibody for an antigen. Such approaches are described in further detail in U.S. Patent Nos. 5,714,350 and 6,350,861 by Co et al.

[0201] Glycosylation of the constant region at N297 can be prevented by mutating the N297 residue to another residue, e.g., N297A, and / or by mutating an adjacent amino acid, e.g., 298, thereby reducing glycosylation at N297.

[0202] Additionally or alternatively, antibodies can be generated with altered glycosylation, e.g., hypofucosylated antibodies with reduced amounts of fucosyl residues or antibodies with increased bisecting GlcNac structures. Such altered glycosylation patterns have been shown to increase the ADCC and / or ADCP capabilities of antibodies. Such carbohydrate modifications can be achieved, for example, by expressing the antibody in a host cell with altered glycosylation machinery. Cells with altered glycosylation machinery have been described in the art and can be used as host cells to express the recombinant anti-TIM-3 antibodies or antigen-binding fragments described herein, thereby producing antibodies with altered glycosylation. For example, EP 1,176,195 by Hanai et al. describes cell lines in which the FUT8 gene, encoding a fucosyltransferase, has been functionally disrupted, such that antibodies expressed in such cell lines exhibit hypofucosylation. PCT Publication WO 03 / 035835 by Presta describes a mutant CHO cell line, Led 3 cells, that has a reduced ability to attach fucose to Asn(297)-linked carbohydrates, which also results in hypofucosylation of antibodies expressed in the host cells (see also Shields, R.L. et al. (2002) J. Biol. Chem. 277:26733-26740). PCT Publication WO 99 / 54342 by Umana et al. describes cell lines engineered to express glycosyltransferases (e.g., β(1,4)-N-acetylglucosaminyltransferase III (GnTIII)) that modify glycoproteins such that antibodies expressed in the engineered cell lines exhibit an increase in bisecting GlcNac structures, which results in increased ADCC activity of the antibodies (see also Umana et al. (1999) Nat. Biotech. 17: 176-180).

[0203] Another modification of the anti-TIM-3 antibodies or antigen-binding fragments described herein is pegylation. Antibodies can be pegylated, for example, to extend the biological (e.g., serum) half-life of the antibody. To pegylate an antibody, the antibody or fragment thereof is typically reacted with polyethylene glycol (PEG), such as a reactive ester or aldehyde derivative of PEG, under conditions that result in one or more PEG groups being attached to the antibody or antibody fragment. In some embodiments, pegylation is carried out via an acylation reaction or an alkylation reaction with a reactive PEG molecule (or an analogous reactive water-soluble polymer). As used herein, the term "polyethylene glycol" is intended to encompass forms of PEG that have been used to derivatize other proteins, such as mono(CI-CIO)alkoxy- or aryloxy-polyethylene glycol or polyethylene glycol-maleimide. In some embodiments, the antibody to be pegylated is an aglycosylated antibody. Methods for pegylation of proteins are known in the art and can be applied to the anti-TIM-3 antibodies or antigen-binding fragments described herein. See, for example, EP 0 154 316 by Nishimura et al. and EP 0 401 384 by Ishikawa et al.

[0204] In some embodiments, variants can include the addition of amino acid residues at the amino and / or carboxyl termini of an antibody or polypeptide. The length of the additional amino acid residues can range from 1 residue to 100 or more residues. In some embodiments, variants include an N-terminal methionyl residue. In some embodiments, variants include an additional polypeptide / protein (e.g., an Fc region) to generate a fusion protein. In some embodiments, variants are modified to be detectable and can include a detectable label and / or protein (e.g., a fluorescent tag or an enzyme).

[0205] The variant antibodies or antigen-binding fragments described herein can be generated using methods known in the art, including but not limited to, site-directed mutagenesis, alanine scanning mutagenesis, and PCR mutagenesis.

[0206] In some embodiments, variants of the anti-TIM-3 antibodies or antigen-binding fragments disclosed herein can retain the ability to bind to TIM-3 to a similar, identical, or greater extent than the parent antibody or antigen-binding fragment. In some embodiments, the variants can be at least about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more identical in amino acid sequence to the parent antibody or antigen-binding fragment. In certain embodiments, variants of the anti-TIM-3 antibodies or antigen-binding fragments comprise the amino acid sequence of the parent anti-TIM-3 antibody or antigen-binding fragment with one or more conservative amino acid substitutions. Conservative amino acid substitutions are known in the art and include amino acid substitutions in which an amino acid with particular physical and / or chemical properties is replaced with another amino acid with the same or similar physical and / or chemical properties.

[0207] In some embodiments, a variant of an anti-TIM-3 antibody or antigen-binding fragment comprises the amino acid sequence of a parent antibody or antigen-binding fragment with one or more non-conservative amino acid substitutions. In some embodiments, a variant of an anti-TIM-3 antibody or antigen-binding fragment comprises the amino acid sequence of a parent binding antibody or antigen-binding fragment with one or more non-conservative amino acid substitutions, wherein the one or more non-conservative amino acid substitutions do not disrupt or inhibit one or more biological activities (e.g., TIM-3 binding) of the variant. In certain embodiments, the one or more conservative amino acid substitutions and / or the one or more non-conservative amino acid substitutions can enhance the biological activity of the variant, such that the biological activity of the functional variant is increased compared to the parent antibody or antigen-binding fragment.

[0208] In some embodiments, the variant can have one, two, three, four, or five amino acid substitutions in the CDRs of the antibody or antigen-binding fragment (e.g., VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and VL CDR3).

[0209] Described herein are antibodies, e.g., humanized antibodies, characterized by particular functional features or properties. For example, the antibodies specifically bind to human TIM-3, more specifically to a particular domain (e.g., a functional domain) within the extracellular domain of human TIM-3. In some embodiments, the antibodies are antagonistic antibodies, i.e., they inhibit or suppress the inhibitory activity of TIM-3 on cells, e.g., T cells or myeloid cells. In some embodiments, the anti-TIM-3 antibodies provided herein do not cross-react with TIM-3 from other species, e.g., cynomolgus monkey TIM-3 or mouse TIM-3. In some embodiments, the antibodies specifically bind to the extracellular region of human TIM-3. In some embodiments, the antibodies bind to human TIM-3 with high affinity.

[0210] The anti-TIM-3 antibodies or antigen-binding fragments described herein exhibit one or more of the following binding properties: (1) they block or inhibit binding of human TIM-3 to a TIM-3 ligand (e.g., PtdSer), e.g., as determined by the assays described herein; (2) they inhibit binding of immune cells, e.g., CD4 + , CD8 + (3) internalizing or down-regulating surface TIM-3 upon binding to TIM-3 on T cells, Th1 cells, NK cells, NKT cells, myeloid cells (e.g., dendritic cells, macrophages, monocytes, or tumor cells), or TILs; (4) inducing or stimulating an adaptive immune response; (5) inducing or stimulating an innate immune response; (6) activating immune cells, e.g., CD4 + , CD8 +(6) induce or stimulate T cells, Th1 cells, NK cells, NKT cells, myeloid cells (e.g., dendritic cells, macrophages, monocytes, or tumor cells), or TILs; (7) induce or stimulate immune cell proliferation, e.g., CD4 + , CD8 + (7) induce or stimulate cytokine (e.g., IFN-γ) production by T cells, e.g., Th1 cells, NK cells, NKT cells, myeloid cells (e.g., dendritic cells, macrophages, monocytes, or tumor cells), or TILs; (8) induce or stimulate production of cytokines activated by inflammasomes (e.g., IL-18, soluble CD25), inflammatory cytokines produced by monocyte / macrophage activation (e.g., human IL-1β, IL-18, IL-6, TNFα, or granulysin), and / or Treg effector molecules (e.g., IL-10, perforin); (9) induce or stimulate production of immune cells, e.g., CD4 T cells, mediated by TIM-3. + , CD8 + (10) reduce or inhibit the suppression of T cells, Th1 cells, NK cells, NKT cells, myeloid cells (e.g., dendritic cells, macrophages, monocytes, or tumor cells), or TILs; (11) reduce or inhibit the suppression of TIM-3-mediated inflammasome activation, as determined, for example, using the methods and assays described in Gayden 2018 and Dixon, 2021 (supra).

[0211] The anti-TIM-3 antibodies or antigen-binding fragments described herein exhibit one or more of the following binding properties: (1) specifically bind to soluble and / or membrane-bound human TIM-3; (2) do not cross-react with cynomolgus monkey TIM-3 or mouse TIM-3; and (4) compete with or cross-block the binding of antibodies that bind to TIM-3 described herein (e.g., 3E6, 4H2, 16H1, 18C6, 19D11, CH 5#, CH 8#, CH 9#, CH 10#, CH 11#, 3F2, 36A2, 36B11, 38F8, 38A8, 40F11, 50H9, 84G10, or 39D1) to human TIM-3, as determined, for example, by the assays described in the Examples.

[0212] Epitope mapping is a method for identifying the binding site, region, or epitope on a target protein to which an antibody binds. Various methods for mapping epitopes on a target protein are known in the art. These methods include, but are not limited to, mutagenesis, including shotgun mutagenesis, site-directed mutagenesis, and alanine scanning; domain or fragment scanning; peptide scanning (e.g., Pepscan technology); display methods (e.g., phage display, microbial display, and ribosome / mRNA display); methods involving proteolysis and mass spectrometry; and structure determination (e.g., X-ray crystallography and NMR). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein are characterized by assays including, but not limited to, N-terminal sequencing, amino acid analysis, HPLC, mass spectrometry, ion-exchange chromatography, and papain digestion. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to the epitope on human TIM-3 recognized by MBG453 (an anti-TIM-3 antibody developed by Novartis; see, e.g., Borate et al., Blood (2019): 570-570). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to the epitope on human TIM-3 recognized by TSR022 (an anti-TIM-3 antibody developed by Tesaro / GSK; see, e.g., Pollyea and Craig, Blood 129.12(2017): 1627-1635). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to the epitope on human TIM-3 recognized by both MBG453 and TSR022. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to the epitope on human TIM-3 recognized by MBG453. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to the epitope on human TIM-3 recognized by TSR022.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to an epitope on human TIM-3 recognized by MBG453 or TSR022. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to an epitope on human TIM-3 that is not recognized by MBG453. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to an epitope on human TIM-3 that is not recognized by TSR022. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to an epitope on human TIM-3 that is not recognized by MBG453 or TSR022. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to an epitope on human TIM-3 that is recognized by MBG453 and an epitope on human TIM-3 that is not recognized by MBG453. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment described herein binds to an epitope on human TIM-3 that is recognized by TSR022 and an epitope on human TIM-3 that is not recognized by TSR022.

[0213] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not compete with either Fab 6TXZ or Fab 7KQL for binding to human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to an epitope on human TIM-3 that is different from the epitope recognized by either Fab 6TXZ or Fab 7KQL. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to the epitope on human TIM-3 recognized by Fab 6TXZ. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to the epitope on human TIM-3 recognized by Fab 7KQL. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to an epitope on human TIM-3 that is not recognized by Fab 6TXZ. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to an epitope on human TIM-3 that is not recognized by Fab 7KQL.

[0214] As provided in the Experimental section below, the epitope of 3E6 was mapped to the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, provided herein is an antibody or antigen-binding fragment thereof that specifically binds to human TIM-3 at an epitope comprising at least one of amino acids 71-82 of human TIM-3. In some embodiments, the epitope of the antibody or antigen-binding fragment provided herein comprises at least two, at least three, at least four, at least five, at least six, at least seven, or all eight of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to at least one of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to D71 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to R73 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to D74 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to V75 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to N76 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to W78 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to T79 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to Y82 of human TIM-3.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to at least two of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to at least three of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to at least four of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to at least five of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to at least six of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to at least seven of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to all of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82.

[0215] In some embodiments, provided herein are anti-TIM-3 antibodies or antigen-binding fragments that compete with an anti-TIM-3 antibody described herein (e.g., 3E6) for binding to amino acids 71-82 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein compete for binding to at least one of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein compete for binding to at least two, at least three, at least four, at least five, at least six, at least seven, or all eight of the following residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82.

[0216] Without being bound by theory, in addition to promoting adaptive immunity, the anti-TIM-3 antibodies or antigen-binding fragments described herein can also promote innate immune activity by reducing or inhibiting TIM-3-mediated suppression of inflammasome activation. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein can also reduce or inhibit TIM-3-mediated suppression of myeloid cells, such as dendritic cells, macrophages, or monocytes. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein promote inflammasome activation or myeloid activation, at least in part, by specifically binding to D71, N76, or Y82 of human TIM-3, or any combination thereof. Mutations at some of these residues are known to result in sustained immune activation and increased cytokine production (Gayden, 2018).

[0217] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to D71, N76, or Y82 of human TIM-3. Also provided herein are anti-TIM-3 antibodies or antigen-binding fragments that compete with the anti-TIM-3 antibodies described herein for binding to D71, N76, or Y82 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to variants of human TIM-3 having an amino acid mutation at D71, N76, or Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to variants of human TIM-3 having an amino acid substitution at D71, N76, or Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to variants of human TIM-3 lacking D71, N76, or Y82.

[0218] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to Y82 of human TIM-3. Also provided herein are anti-TIM-3 antibodies or antigen-binding fragments that compete with the anti-TIM-3 antibodies described herein for binding to Y82 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 having an amino acid mutation at Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 having an amino acid substitution at Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 lacking Y82. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to D71 of human TIM-3. In some embodiments, provided herein are anti-TIM-3 antibodies or antigen-binding fragments that compete with the anti-TIM-3 antibodies described herein for binding to D71 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 having an amino acid mutation at D71. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 having an amino acid substitution at D71. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 in which D71 is deleted. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to N76 of human TIM-3. In some embodiments, provided herein are anti-TIM-3 antibodies or antigen-binding fragments that compete with the anti-TIM-3 antibodies described herein for binding to N76 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a variant of human TIM-3 having an amino acid mutation at N76.In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 having an amino acid substitution at N76. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 in which N76 is deleted. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein specifically bind to D71, N76, and Y82 of human TIM-3. In some embodiments, also provided herein are anti-TIM-3 antibodies or antigen-binding fragments that compete with the anti-TIM-3 antibodies described herein for binding to D71, N76, and Y82 of human TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not bind to a mutant of human TIM-3 having amino acid mutations at D71, N76, and Y82.

[0219] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind with high affinity, e.g., at least 10 -7 M or less, 10 -8 M or less, 5×10 -9 M or less, 10 -9 M or less, 5×10 -10 M or less, 10 -10 M or less, 5×10 -11 M or less, 10 -11 M or less, 5×10 -12 M or less, 10 -12 M or less, 10 -12 M~10 -7 M, 10 -11 M~10 -7 M, 10 -10 M~10 -7 M, 10 -9 M~10 -7 M, 10 -8 M~10 -7 M, 10 -10 M~10 -8 M, 10 -9 M~10 -8 M, 10 -11 M~10 -9 M or 10 -10 M~10 -9K of M D In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to human TIM-3 at 10 -11 M~5×10 -9 K of M D In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to human TIM-3 with high affinity, e.g., at least 10 as determined by BLI. -7 M or less, 10 -8 M or less, 5×10 -9 M or less, 10 -9 M or less, 5×10 -10 M or less, 10 -10 M or less, 5×10 -11 M or less, 10 -11 M or less, 5×10 -12 M or less, 10 -12 M or less, 10 -12 M~10 -7 M, 10 -11 M~10 -7 M, 10 -10 M~10 -7 M, 10 -9 M~10 -7 M, 10 -8 M~10 -7 , 10 -10 M~10 -8 M, 10 -9 M~10 -8 M, 10 -11 M~10 -9 M or 10 -10 M~10 -9 K of M D In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to soluble human TIM-3 at 10 -11 M~5×10 -9 K of M D In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to soluble human TIM-3 at 10, as determined, for example, by flow cytometry and Scatchard plots. -7 M or less, 10 -8 M or less, 5×10 -9 M or less, 10 -9M or less, 5×10 -10 M or less, 10 -10 M or less, 5×10 -11 M or less, 10 -11 M or less, 5×10 -12 M or less, 10 -12 M or less, 10 -12 M~10 -7 M, 10 -11 M~10 -7 M, 10 -10 M~10 -7 M, 10 -9 M~10 -7 M, 10 -8 M~10 -7 , 10 -10 M~10 -8 M, 10 -9 M~10 -8 M, 10 -11 M~10 -9 M or 10 -10 M~10 -9 K of M D and binds to bound (e.g., membrane-bound) human TIM-3 on, for example, activated human T cells. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment has an EC2 of 10 μg / mL or less, 5 μg / mL or less, 1 μg / mL or less, 0.9 μg / mL or less, 0.8 μg / mL or less, 0.7 μg / mL or less, 0.6 μg / mL or less, 0.5 μg / mL or less, 0.4 μg / mL or less, 0.3 μg / mL or less, 0.2 μg / mL or less, 0.1 μg / mL or less, 0.05 μg / mL or less, or 0.01 μg / mL or less, as determined, for example, by flow cytometry. 50 and binds to bound (e.g., cell membrane-bound) human TIM-3 on, for example, activated human T cells.

[0220] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein are administered intracellularly, e.g., at 10 -7 M or less, 10 -8 M or less, 5×10 -9 M or less, 10 -9 M or less, 5×10 -10 M or less, 10 -10 M or less, 5×10 -11 M or less, 10-11 M or less, 5×10 -12 M or less, 10 -12 M or less, 10 -12 M~10 -7 M, 10 -11 M~10 -7 M, 10 -10 M~10 -7 M, 10 -9 M~10 -7 M, 10 -8 M~10 -7 , 10 -10 M~10 -8 M, 10 -9 M~10 -8 M, 10 -11 M~10 -9 M or 10 -10 M~10 -9 K of M D In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment binds to cynomolgus monkey TIM-3 at 10, as determined, for example, by BLI (e.g., as described in the Examples). -7 M or less, 10 -8 M or less, 5×10 -9 M or less, 10 -9 M or less, 5×10 -10 M or less, 10 -10 M or less, 5×10 -11 M or less, 10 -11 M or less, 5×10 -12 M or less, 10 -12 M or less, 10 -12 M~10 -7 M, 10 -11 M~10 -7 M, 10 -10 M~10 -7 M, 10 -9 M~10 -7 M, 10 -8 M~10 -7 , 10 -10 M~10 -8 M, 10 -9 M~10 -8 M, 10 -11 M~10 -9 M or 10 -10 M~10 -9 K of M DThe anti-TIM-3 antibody or antigen-binding fragment binds to soluble cynomolgus monkey TIM-3 at an EC of, for example, 100 nM or less, 10 nM or less, 100 nM to 0.01 nM, 100 nM to 0.1 nM, 100 nM to 1 nM, or 10 nM to 1 nM, as measured, for example, by flow cytometry (e.g., as described in the Examples). 50 In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment can bind to membrane-bound cynomolgus monkey TIM-3 at, for example, 10, as determined, for example, by flow cytometry and Scatchard plots. -7 M or less, 10 -8 M or less, 5×10 -9 M or less, 10 -9 M or less, 5×10 -10 M or less, 10 -10 M or less, 5×10 -11 M or less, 10 -11 M or less, 5×10 -12 M or less, 10 -12 M or less, 10 -12 M~10 -7 M, 10 -11 M~10 -7 M, 10 -10 M~10 -7 M, 10 -9 M~10 -7 M, 10 -8 M~10 -7 , 10 -10 M~10 -8 M, 10 -9 M~10 -8 M, 10 -11 M~10 -9 M or 10 -10 M~10 -9 K of M D and binds to bound (e.g., membrane-bound) cynomolgus TIM-3 on activated human T cells.

[0221] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not detectably bind to cynomolgus monkey TIM-3.

[0222] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein stimulate or enhance an adaptive immune response. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein stimulate or enhance an immune response, e.g., by activating T cells, e.g., within a tumor. For example, the anti-TIM-3 antibodies or antigen-binding fragments can activate or costimulate cells, as evidenced by, e.g., enhanced cytokine (e.g., IFN-γ) secretion and / or enhanced proliferation, which may result from inhibition of TIM-3-mediated T cell inhibitory activity. In some embodiments, T cell activation or costimulation by the TIM-3 antibodies or antigen-binding fragments occurs in the presence of CD3 stimulation. In some embodiments, the anti-TIM-3 antibodies increase IFN-γ secretion by 50%, 100% (i.e., 2-fold), 3-fold, 4-fold, 5-fold, or more, optionally up to 10-fold, 30-fold, or 100-fold, e.g., as measured in primary human T cells and / or T cells expressing human TIM-3, e.g., TILs. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein block or reduce the inhibitory effect of TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein increased IFN-γ production in TIM-3-expressing T cells (e.g., Th1 cells or TILs). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein enhance the proliferation of TIM-3-expressing T cells (e.g., Th1 cells or TILs). In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein stimulate T cell proliferation in a mixed lymphocyte reaction (MLR) assay.

[0223] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein stimulate or enhance innate immune responses. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein induce or stimulate the activation of myeloid cells, such as dendritic cells, macrophages, or monocytes. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein reduce or inhibit TIM-3-mediated suppression of myeloid cells, such as dendritic cells, macrophages, or monocytes. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein reduce or inhibit TIM-3-mediated modulation or suppression of inflammasome activation. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein induce or stimulate the production of inflammasome-activated cytokines, such as IL-18 and soluble CD25. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein induce or stimulate the production of proinflammatory cytokines produced by monocyte / macrophage activation, such as human IL-1β, IL-18, IL-6, TNFα, or granulysin. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein induce or stimulate the production of Treg effector molecules, such as IL-10 and perforin.

[0224] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein, upon binding to TIM-3 on a cell, internalize or downregulate cell surface TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein bind to immune cells, e.g., CD4 + , CD8 +Upon binding to TIM-3 on T cells, Th1 cells, NK cells, NKT cells, myeloid cells (e.g., dendritic cells, macrophages, or monocytes), or TILs, they internalize or downregulate cell surface TIM-3. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein internalize or downregulate cell surface TIM-3 upon binding to TIM-3 on T cells. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein internalize or downregulate cell surface TIM-3 upon binding to TIM-3 on NK cells. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein internalize or downregulate cell surface TIM-3 upon binding to TIM-3 on tumor cells. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein do not internalize or downregulate cell surface TIM-3 upon binding to TIM-3 on tumor cells.

[0225] In some embodiments, the antibodies or antigen-binding fragments provided herein bind to the site on TIM-3 to which a TIM-3 ligand binds. In some embodiments, the antibodies or antigen-binding fragments provided herein block the interaction between TIM-3 and a TIM-3 ligand, wherein the TIM-3 ligand is PtdSer, HMGB1, CEACAM-1, or any combination thereof. In some embodiments, the TIM-3 ligand is PtdSer, and the antibodies or antigen-binding fragments provided herein bind to the site on TIM-3 to which PtdSer binds. In some embodiments, the antibodies or antigen-binding fragments provided herein block the interaction between PtdSer and TIM-3. In some embodiments, the TIM-3 ligand is CEACAM-1, and the antibodies or antigen-binding fragments provided herein bind to the site on TIM-3 to which CEACAM-1 binds. In some embodiments, the antibodies or antigen-binding fragments provided herein block the interaction between CEACAM-1 and TIM-3. In some embodiments, the TIM-3 ligand is HMGB1, and the antibodies or antigen-binding fragments provided herein bind to the site on TIM-3 to which HMGB1 binds. In some embodiments, the antibodies or antigen-binding fragments provided herein block the interaction between HMGB1 and TIM-3.In some embodiments, the binding of TIM-3 to its ligand, e.g., PtdSer, is measured using an art-recognized method, e.g., a FACS-based binding assay, e.g., using CHO cells transfected with human TIM-3 or activated T cells expressing TIM-3, to a concentration of about 1 μg / mL or less, e.g., about 0.9 pg / mL or less, about 0.85 pg / mL or less, about 0.8 pg / mL or less, about 0.75 pg / mL or less, about 0. The antibodies or antigen-binding fragments provided herein inhibit with an EC50 of 7 pg / mL or less, about 0.65 pg / mL or less, about 0.6 pg / mL or less, about 0.55 pg / mL or less, about 0.5 pg / mL or less, about 0.45 pg / mL or less, about 0.4 pg / mL or less, about 0.35 pg / mL or less, about 0.3 pg / mL or less, about 0.25 pg / mL or less, about 0.2 pg / mL or less, about 0.15 pg / mL or less, about 0.1 pg / mL or less, or about 0.05 pg / mL or less.

[0226] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein inhibit the binding of phosphatidylserine to TIM-3, e.g., as measured by a PS-hTIM-3 "in tandem" blocking assay. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments inhibit the binding of human TIM-3 to PtdSer on cells, e.g., CHO cells or activated T cells, expressing human TIM-3, with an EC of, e.g., 10 pg / mL or less, 1 pg / mL or less, 0.01 pg / mL to 10 pg / mL, 0.1 pg / mL to 10 pg / mL, or 0.1 pg / mL to 1 pg / mL. 50 and inhibits.

[0227] Thus, an anti-TIM-3 antibody or antigen-binding fragment exhibiting one or more of these functional properties (e.g., biochemical, immunochemical, cellular, physiological, or other biological activity, etc.) known in the art and determined according to the methodologies described herein will be understood to exhibit a statistically significant difference in the particular activity compared to that seen in the absence of the antibody (e.g., or in the presence of a control antibody of irrelevant specificity). In some embodiments, the increase in a measured parameter (e.g., T cell proliferation, cytokine production) induced by an anti-TIM-3 antibody or antigen-binding fragment in a given assay results in a statistically significant increase in the measured parameter of at least 10%, e.g., at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 100% (i.e., 2-fold), 3-fold, 5-fold, or 10-fold; in some embodiments, the antibodies described herein can increase the measured parameter by more than 92%, 94%, 95%, 97%, 98%, 99%, 100% (i.e., 2-fold), 3-fold, 5-fold, or 10-fold, compared to the same assay performed in the absence of the antibody. Conversely, the reduction in a measured parameter (e.g., tumor volume, TIM-3 ligand binding to human TIM-3) induced by an anti-TIM-3 antibody in a given assay results in a statistically significant reduction of the measured parameter by at least 10%, e.g., at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%, and in some embodiments, the antibodies described herein can reduce the measured parameter by greater than, e.g., 92%, 94%, 95%, 97%, 98%, or 99%, compared to the same assay performed in the absence of the antibody.

[0228] In some embodiments, the anti-TIM-3 antibody does not have agonist activity, e.g., as determined in cross-linking of the anti-TIM-3 antibody in a CHO-OKT3-CD32:T cell coculture experiment, and such an antibody does not enhance activity over anti-TIM-3 alone. In some embodiments, the anti-TIM-3 antibody blocks the interaction of TIM-3 with its ligand without promoting agonist activity.

[0229] In some embodiments, the anti-TIM-3 antibody enhances IL-12 production from monocytes or dendritic cells treated with lipopolysaccharide (LPS).

[0230] In some embodiments, the anti-TIM-3 antibody is combined with the treatment, thus inhibiting CD8 + Tumor-infiltrating CD8 co-expressing PD-1 and TIM-3 evade T cell exhaustion + Restore T cells.

[0231] Anti-TIM-3 antibodies or antigen-binding fragments can be analyzed for their physical, chemical, and / or biological properties by various methods known in the art. In some embodiments, anti-TIM-3 antibodies are tested for their ability to bind to TIM-3 (e.g., human TIM-3). Binding assays include, but are not limited to, BLI, SPR (e.g., Biacore), ELISA, FACS, Western blot, and RIA. Additionally, antibodies can be evaluated for solubility, stability, thermal stability, viscosity, expression level, expression quality, and / or purification efficiency. Assays to evaluate the effect of antibodies on the functional properties of TIM-3 (e.g., ligand binding, T cell proliferation, cytokine production) are described in further detail below and in the Examples.

[0232] In some embodiments, the anti-TIM-3 antibody is not a native or naturally occurring antibody, e.g., the anti-TIM-3 antibody has post-translational modifications that differ from those of naturally occurring antibodies, e.g., by having more, fewer, or different types of post-translational modifications.

[0233] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein are chemically modified, either naturally or by intervention. In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments are chemically modified by glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization with known protecting / blocking groups, proteolytic cleavage, and / or linkage to a cellular ligand or other protein. Any of a number of chemical modifications can be performed using known techniques. The anti-TIM-3 antibodies or antigen-binding fragments can include one or more analogs of an amino acid (e.g., including unnatural amino acids), as well as other modifications known in the art.

[0234] In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment is conjugated to a cytotoxic agent or moiety. In some embodiments, the anti-TIM-3 antibody or antigen-binding fragment is conjugated to a cytotoxic agent to form an ADC (antibody-drug conjugate). In some embodiments, the cytotoxic moiety is a chemotherapeutic agent, including but not limited to methotrexate, adriamycin / doxorubicin, melphalan, mitomycin C, chlorambucil, duocarmycin, daunorubicin, pyrrolobenzodiazepines (PBDs), or other intercalating agents. In some embodiments, the cytotoxic moiety is a microtubule inhibitor, including but not limited to auristatins, maytansinoids (e.g., DM1 and DM4), and tubulysins. In some embodiments, the cytotoxic moiety is an enzymatically active toxin or fragment thereof of bacterial, fungal, plant, or animal origin, including, but not limited to, diphtheria A chain, the nonbinding active portion of diphtheria toxin, exotoxin A chain, ricin A chain, abrin A chain, modeccin A chain, α-sarcin, Aleurites fordii protein, diansin protein, Phytolacca americana protein (PAPI, PAPII, and PAP-S), Momordica charantia inhibitor, curcin, crotin, Sapaonaria officinalis inhibitor, gelonin, mitogenin, restrictocin, phenomycin, enomycin, and a trichothecene. In some embodiments, the antibody is conjugated to one or more small molecule toxins, such as calicheamicin, maytansinoids, trichothenes, and CC1065.

[0235] In some embodiments, the anti-TIM-3 antibodies or antigen-binding fragments described herein are conjugated to a detectable substance or molecule, allowing the agent to be used for diagnosis and / or detection. Detectable substances include enzymes, such as horseradish peroxidase, alkaline phosphatase, β-galactosidase, and acetylcholinesterase; prosthetic groups, such as biotin and flavins; fluorescent substances, such as umbelliferone, fluorescein, fluorescein isothiocyanate (FITC), rhodamine, tetramethylrhodamine isothiocyanate (TRITC), dichlorotriazinylamine fluorescein, dansyl chloride, cyanine (Cy3), and phycoerythrin; bioluminescent substances, such as luciferase; radioactive substances, such as 212 Bi, 14 C. 57 Co, 51 Cr, 67 Cu, 18 F, 68 Ga, 67 Ga, 153 Gd, 159 Gd, 68 Ge, 3 H, 166 Ho, 131 I, 125 I, 123 I, 121 I, 115 In, 113 In, 112 In, 111 In, 140 La, 177 Lu, 54 Mn, 99 Mo, 32 P, 103 Pd, 149 Pm, 142 Pr, 186 Re, 188 Re, 105 Rh, 97 Ru, 35 S, 47 Sc, 75 Se, 153 Sm, 113 Sn, 117 Sn, 85 Sr, 99m Tc,201 Ti, 133 Xe, 90 Y, 69 Yb, 175 Yb, 65 Zn; positron-emitting metals; and magnetic metal ions. Positron-emitting metals; and magnetic metal ions can be included, but are not limited to these.

[0236] In some embodiments, the anti-T...

Claims

1. An antibody or antigen-binding fragment thereof that specifically binds to human TIM-3, wherein the antibody or antigen-binding fragment thereof specifically binds to an epitope comprising at least one of amino acids 71 ​​to 82 of human TIM-3.

2. The antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment thereof (1) specifically binds to at least one, at least two, at least three, at least four, at least five, at least six, at least seven, or eight of the following amino acid residues of human TIM-3: D71, R73, D74, V75, N76, W78, T79, and Y82; (2) specifically binds to at least D71, N76, or Y82 of human TIM-3; or (3) specifically binds to at least Y82 of human TIM-3, and optionally, the antibody or antigen-binding fragment thereof does not specifically bind to an epitope outside amino acids 71 ​​to 82 of human TIM-3.

3. An antibody or antigen-binding fragment thereof that specifically binds to human TIM-3, (a)(1) a light chain CDR1 (VL CDR1) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 86-93 and 129-137; (2) a light chain CDR2 (VL CDR2) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 94-100 and 138-144; and (3) a light chain variable region (VL) comprising a light chain CDR3 (VL CDR3) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47-55, 145-153, and 198-206; or a variant thereof having up to about 3 amino acid substitutions, additions, and / or deletions in said VL CDR; and / or (b)(1) a heavy chain CDR1 (VH CDR1) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 101-108 and 154-161; (2) a heavy chain CDR2 (VH CDR2) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 109-118 and 162-170; and (3) a heavy chain variable region (VH) comprising a heavy chain CDR3 (VH CDR3) having an amino acid sequence selected from the group consisting of SEQ ID NOs: 119-128 and 171-179; or and variants thereof having up to about 3 amino acid substitutions, additions, and / or deletions in said VH CDRs. The antibody or antigen-binding fragment thereof.

4. (1) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 47, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (2) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 87, 95, and 48, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 102, 110, and 120, respectively; (3) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 88, 96, and 49, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 103, 111, and 121, respectively; (4) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 89, 97, and 50, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 104, 112, and 122, respectively; (5) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 90, 94, and 51, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 105, 113, and 123, respectively; (6) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 91, 98, and 52, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 106, 114, and 124, respectively; (7) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 91, 98, and 53, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 106, 115, and 125, respectively; (8) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 92, 99, and 54, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 107, 116, and 126, respectively; (9) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 93, 100, and 55, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 108, 117, and 127, respectively; (10) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 91, 98, and 52, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 106, 118, and 128, respectively; (11) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 129, 138, and 145, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 106, 162, and 171, respectively; (12) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 130, 139, and 146, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 154, 163, and 172, respectively; (13) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 131, 140, and 147, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 155, 164, and 173, respectively; (14) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 132, 141, and 148, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 156, 165, and 174, respectively; (15) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 133, 139, and 149, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 157, 166, and 175, respectively; (16) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 134, 142, and 150, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 158, 167, and 176, respectively; (17) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 135, 143, and 151, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 159, 168, and 177, respectively; (18) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 136, 144, and 152, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 160, 169, and 178, respectively; (19) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 137, 100, and 153, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 161, 170, and 179, respectively; (20) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 198, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (21) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 199, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (22) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 200, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (23) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 201, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (24) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 202, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (25) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 203, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (26) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 204, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; (27) the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 205, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively; or (28) The antibody or antigen-binding fragment thereof described in claim 3, wherein the VL CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, and 206, respectively; and / or the VH CDR1, CDR2, and CDR3 have the amino acid sequences of SEQ ID NOs: 101, 109, and 119, respectively.

5. 4. The antibody or antigen-binding fragment thereof of claim 3, comprising VL CDR1, VL CDR2, VH CDR1, VH CDR2, and VH CDR3 having the amino acid sequences of SEQ ID NOs: 86, 94, 101, 109, and 119, respectively, and a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47 and 198-206; or a variant thereof having up to about three amino acid substitutions, additions, and / or deletions in the VL CDR and up to about three amino acid substitutions, additions, and / or deletions in the VH CDR.

6. The antibody or antigen-binding fragment thereof of claim 5, wherein the F residue of VH CDR1 (amino acid 3 of SEQ ID NO: 101), the H and S residues of VH CDR2 (amino acids 4 and 5 of SEQ ID NO: 109), the Y, R, S and W residues of VH CDR3 (amino acids 2, 3, 4 and 6 of SEQ ID NO: 119) and the S residue of VL CDR2 (amino acid 7 of SEQ ID NO: 94) are not mutated; or the VL CDR1, VL CDR2, VL CDR3, VH CDR1, VH CDR2 and VH CDR3 have the amino acid sequences of SEQ ID NOs: 86, 94, 47, 101, 109 and 119, respectively.

7. (a) a VL having at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 207-215, 1-10, and 180-188; and / or (b) a VH having at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 28, 22-27, 29, 11-20, and 189-197, or an antibody or antigen-binding fragment thereof according to claim 3.

8. (1) SEQ ID NO:1 and 11, respectively; (2) SEQ ID NO:2 and 12, respectively; (3) SEQ ID NO:3 and 13, respectively; (4) SEQ ID NO:4 and 14, respectively; (5) SEQ ID NO:5 and 15, respectively; (6) SEQ ID NO:6 and 16, respectively; (7) SEQ ID NO:7 and 17, respectively; (8) SEQ ID NO:8 and 18, respectively; (9) SEQ ID NO:9 and 19, respectively; (10) SEQ ID NO:10 and 20, respectively; (11) SEQ ID NO:180 and 189, respectively; (12) SEQ ID NO:181 and 190, respectively; (13) SEQ ID NO:182 and 191, respectively; (14) SEQ ID NO:183 and 192, respectively; (15) SEQ ID NO:184 and 193, respectively; (16) SEQ ID NO:185 and 194, respectively; (17) SEQ ID NO:186 and 196, respectively; 8. The antibody or antigen-binding fragment thereof of claim 7, wherein: (a) the VL has at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 21 and 207-215; and / or (b) the VH has at least 85%, at least 90%, at least 95%, at least 98%, or 100% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs: 22-29.

9. An antibody or antigen-binding fragment thereof that specifically binds to human TIM-3, (a) a VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-10 and 180-188; and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 11-20 and 189-197; or (b) a VL and a VH, wherein the VL comprises a VL CDR1, CDR2, and CDR3 from a VL having an amino acid sequence selected from the group consisting of SEQ ID NOs: 21 and 207-215, and the VH comprises a VH CDR1, CDR2, and CDR3 from a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 22-29; The antibody or antigen-binding fragment thereof.

10. (1) a VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 1, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 11; (2) a VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 2, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 12; (3) a VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 3, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 13; (4) a VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 4, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 14; (5) a VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO:5, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO:15; (6) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 6, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 16; (7) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 7, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 17; (8) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 8, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 18; (9) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 9, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 19; (10) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 10, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 20; (11) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 180, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 189; (12) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 181, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 190; (13) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 182, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 191; (14) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 183, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 192; (15) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 184, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 193; (16) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 185, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 194; (17) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 186, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 195; (18) A VL comprising a VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 187, and / or a VH comprising a VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 196; or (19) A VL comprising VL CDR1, CDR2, and CDR3 derived from a VL having the amino acid sequence of SEQ ID NO: 188, and / or a VH comprising VH CDR1, CDR2, and CDR3 derived from a VH having the amino acid sequence of SEQ ID NO: 197; The antibody or antigen-binding fragment thereof of claim 9,

11. An antibody or antigen-binding fragment thereof that competes with the antibody or antigen-binding fragment thereof of any one of claims 1 to 10 for binding to human TIM-3.

12. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 10, which (1) blocks the interaction between TIM-3 and a TIM-3 ligand, and optionally the TIM-3 ligand is phosphatidylserine, CEACAM1, HMGB1, or any combination thereof; (2) inhibits TIM-3-mediated T cell suppression; (3) inhibits TIM-3-mediated myeloid cell suppression; or (4) inhibits TIM-3-mediated suppression of inflammasome activation, or any combination of these (1) to (4).

13. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 10, which is (1) a monoclonal antibody or an antigen-binding fragment thereof; and / or (2) an IgG1 antibody, an IgG2 antibody, an IgG3 antibody, an IgG4 antibody, Fab, Fab', or F(ab'). 2 , Fv, scFv, (scFv) 2 (2) an antibody or antigen-binding fragment thereof selected from the group consisting of a chimeric antibody or antigen-binding fragment thereof, a humanized antibody or antigen-binding fragment thereof, or a human antibody or antigen-binding fragment thereof, and optionally wherein the antibody or antigen-binding fragment thereof is a humanized antibody or antigen-binding fragment thereof.

14. A polynucleotide encoding the antibody or antigen-binding fragment thereof according to any one of claims 1 to 10.

15. A vector comprising the polynucleotide of claim 14.

16. A host cell comprising the polynucleotide of claim 14.

17. A pharmaceutical composition comprising a therapeutically effective amount of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 10 and a pharma- ceutical acceptable carrier.

18. 11. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 10 for use in (1) inducing or stimulating activation and / or proliferation of immune cells, or (2) reducing TIM-3-mediated suppression of immune cells, optionally wherein the immune cells are T cells, NK cells, NKT cells, or myeloid cells, and optionally wherein the myeloid cells are macrophages or dendritic cells.

19. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 10 for use in: (1) stimulating anti-tumor immunity in a subject in need thereof; or (2) inhibiting tumor cell proliferation in a subject in need thereof.

20. 11. The antibody or antigen-binding fragment thereof of any one of claims 1 to 10 for use in treating cancer in a subject in need thereof.

21. 21. The antibody or antigen-binding fragment thereof for use according to claim 20, wherein the antibody or antigen-binding fragment thereof is used in combination with an additional therapy, optionally wherein the additional therapy comprises an antibody that specifically binds to PD-L1, PD-1, CEACAM1, CTLA4, CEACAM5, latent TGF-β, a TGF-β receptor, CD70, B7H4, or B7H3, or comprises radiation or chemotherapy.

22. 13. Use of an antibody or antigen-binding fragment thereof according to any one of claims 1 to 10 for the preparation of a medicament for the treatment of cancer.

23. 21. The antibody or antigen-binding fragment thereof for use according to claim 20, wherein (1) the cancer is a hematological cancer or a solid tumor, optionally the hematological cancer is acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS), and optionally the solid tumor is melanoma, lung cancer, head and neck cancer, colorectal cancer, pancreatic cancer, gastric cancer, renal cancer, bladder cancer, prostate cancer, breast cancer, ovarian cancer, uterine / cervical cancer, testicular cancer, thyroid cancer, esophageal cancer, soft tissue sarcoma, liver cancer, gallbladder cancer, cervical cancer, duodenal cancer, bone cancer, neuroendocrine cancer, intestinal cancer, skin cancer, or germ cell cancer. and / or (1) the cancer is selected from the group consisting of renal cell carcinoma (RCC), non-small cell lung cancer (NSCLC), squamous cell carcinoma of the head and neck (SCCHN), triple-negative breast cancer (TNBC), gastric / gastric adenocarcinoma (STAD), pancreatic adenocarcinoma (PAAD), colon adenocarcinoma (COAD), and rectal adenocarcinoma (READ); and / or (2) the cancer has high microsatellite instability; and / or (3) the cancer is metastatic, refractory, or recurrent cancer.

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