Anti-CCR8 antibodies and uses thereof

Abstract

The present disclosure relates to anti-CCR8 (chemokine (CC motif) receptor 8) antibodies, antigen-binding fragments, and uses thereof.

Description

[Technical Field]

[0001] Priority claims This application claims priority to PCT / CN2022 / 128366, filed October 28, 2022, and PCT / CN2023 / 114451, filed August 23, 2023. The entire contents of the above-mentioned applications are incorporated herein by reference.

[0002] Technical Field The present disclosure relates to anti-CCR8 (chemokine (CC motif) receptor 8) antibodies and uses thereof. [Background technology]

[0003] Regulatory T (Treg) cells suppress aberrant / excessive immune responses to self- and non-self-antigens to maintain immune homeostasis. In tumor immunity, Treg cells are involved in tumor development and progression by inhibiting anti-tumor immunity.

[0004] The recent clinical and commercial success of anti-cancer antibodies has generated significant interest in antibody-based therapies, including those targeting Tregs. There is a need to develop anti-cancer antibodies that target Tregs for use in various antibody-based therapies for cancer treatment. Summary of the Invention

[0005] The present disclosure relates to anti-CCR8 antibodies, antigen-binding fragments thereof, and uses thereof. In one aspect, the present disclosure provides fully human anti-CCR8 antibodies or antigen-binding fragments thereof (e.g., antibodies or antigen-binding fragments thereof produced by a genetically modified non-human animal having a humanized heavy chain immunoglobulin locus and a humanized light chain immunoglobulin locus).

[0006] In one aspect, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to CCR8 (chemokine (CC motif) receptor 8), a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VH CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR1, the VH CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR2, and the VH CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR3; a light chain variable region (VL) comprising CDR1, 2, and 3, wherein the VL CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR1, the VL CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR2, and the VL CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR3; The amino acid sequences of the selected VH CDR1, 2, and 3 are as follows: (1) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 1, 2, and 3, respectively; (2) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 4, 5, and 6, respectively; (3) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 7, 8, and 9, respectively; (4) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 10, 11, and 12, respectively; (5) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 13, 14, and 15, respectively; (6) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 16, 17, and 18, respectively; (7) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 72, 73, and 74, respectively; (8) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 75, 76, and 77, respectively; (9) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 78, 79, and 80, respectively; (10) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 81, 82, and 83, respectively; (11) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 84, 85, and 86, respectively; (12) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 87, 88, and 89, respectively; (13) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 90, 91, and 92, respectively; (14) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 93, 94, and 95, respectively; (15) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 96, 97, and 98, respectively; (16) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 99, 100, and 101, respectively; (17) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 102, 103, and 104, respectively; (18) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 105, 106, and 107, respectively; (19) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 144, 145, and 146, respectively; (20) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 147, 148, and 149, respectively; (21) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 150, 151, and 152, respectively; (22) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 153, 154, and 155, respectively; (23) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 156, 157, and 158, respectively. (24) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 159, 160, and 161, respectively; (25) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 162, 163, and 164, respectively; (26) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 165, 166, and 167, respectively; (27) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 168, 169, and 170, respectively; (28) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 171, 172, and 173, respectively. (29) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 174, 175, and 176, respectively; (30) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 177, 178, and 179, respectively; (31) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 219, 220, and 221. (32) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 222, 223, and 224, respectively. (33) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 225, 226, and 227, respectively; (34) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 228, 229, and 230, respectively; (35) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 231, 232, and 233, respectively; (36) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 234, 235, and 236, respectively. (37) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 237, 238, and 239, respectively. (38) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 240, 241, and 242, respectively. (39) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 243, 244, and 245, respectively. (40) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 246, 247, and 248, respectively; (41) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 249, 250, and 251, respectively. (42) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 252, 253, and 254, respectively. (43) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 255, 256, and 257, respectively; and (44) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 258, 259, and 260, respectively. One of the following: The amino acid sequences of the selected VL CDR1, 2, and 3 are as follows: (1) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 19, 20, and 21, respectively; (2) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 22, 23, and 24, respectively; (3) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 25, 26, and 27, respectively; (4) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 28, 29, and 30, respectively; (5) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 31, 32, and 33, respectively; (6) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 34, 35, and 36, respectively; (7) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 108, 109, and 110, respectively; (8) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 111, 112, and 113, respectively; (9) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 114, 115, and 116, respectively; (10) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 117, 118, and 119, respectively; (11) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 120, 121, and 122, respectively; (12) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 123, 124, and 125, respectively; (13) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 126, 127, and 128, respectively; (14) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 129, 130, and 131, respectively; (15) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 132, 133, and 134, respectively; (16) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 135, 136, and 137, respectively; (17) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 138, 139, and 140, respectively; (18) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 141, 142, and 143, respectively; (19) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 180, 181, and 182, respectively; (20) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 183, 184, and 185, respectively; (21) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 186, 187, and 188, respectively; (22) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 189, 190, and 191, respectively; (23) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 192, 193, and 194, respectively; (24) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 195, 196, and 197, respectively; (25) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 198, 199, and 200, respectively; (26) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 201, 202, and 203, respectively. (27) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 204, 205, and 206, respectively. (28) The amino acid sequences of the selected VL CDR1, 2, and 3 are represented by SEQ ID NOs: 207, 208, and 209, respectively. (29) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 210, 211, and 212, respectively; and (30) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 213, 214, and 215, respectively. The present invention relates to an antibody or antigen-binding fragment thereof, which is any one of:

[0007] In one aspect, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to CCR8 (chemokine (CC motif) receptor 8), a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VH CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR1, the VH CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR2, and the VH CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR3; a light chain variable region (VL) comprising CDR1, 2, and 3, wherein the VL CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR1, the VL CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR2, and the VL CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR3; The amino acid sequences of the selected VH CDR1, 2, and 3 are as follows: (1) the selected VH CDR1 is SYVMH (SEQ ID NO: 99), the selected VH CDR2 is VISYX1X2SDKYYADSVKG (SEQ ID NO: 100), and the selected VH CDR3 is GRGYRYGQSYYYGMDV (SEQ ID NO: 101); (2) the selected VH CDR1 is SYVVH (SEQ ID NO: 102), the selected VH CDR2 is VISYX1X2DNKFYADSVKG (SEQ ID NO: 103), and the selected VH CDR3 is GRX5YX6X7YYGLDV (SEQ ID NO: 104); (3) the selected VH CDR1 is TYVMH (SEQ ID NO: 105), the selected VH CDR2 is VISYX1X2NNKYYADSVKG (SEQ ID NO: 106), and the selected VH CDR3 is GRSYVNYYGLDV (SEQ ID NO: 107); (4) The selected VH CDR1 is GFSFSSY (SEQ ID NO: 171), the selected VH CDR2 is SYX1X2SD (SEQ ID NO: 172), and the selected VH CDR3 is GRGYRYGQSYYYGMDV (SEQ ID NO: 173); (5) the selected VH CDR1 is GFTFSSY (SEQ ID NO: 174), the selected VH CDR2 is SYX1X2DN (SEQ ID NO: 175), and the selected VH CDR3 is GRX5YX6X7YYGLDV (SEQ ID NO: 176); and (6) The selected VH CDR1 is GFTFSTY (SEQ ID NO: 177), the selected VH CDR2 is SYX1X2NN (SEQ ID NO: 178), and the selected VH CDR3 is GRSYVNYYGLDV (SEQ ID NO: 179); Among them, the amino acid sequences of the selected VL CDR1, 2, and 3 are any one of the following: (1) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 135), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 136), and the selected VL CDR3 is MQSIKLPLT (SEQ ID NO: 137); (2) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 138), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 139), and the selected VL CDR3 is MQSLKVPPT (SEQ ID NO: 140); (3) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 141), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 142), and the selected VL CDR3 is MQSVKIPLT (SEQ ID NO: 143); (4) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 207), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 208), and the selected VL CDR3 is MQSIKLPLT (SEQ ID NO: 209); (5) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 210), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 211), and the selected VL CDR3 is MQSLKVPPT (SEQ ID NO: 212); and (6) The selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 213), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 214), and the selected VL CDR3 is MQSVKIPLT (SEQ ID NO: 215). One of the following, provided that: X1 and X3 in each sequence are independently selected from the group consisting of D, S, and A; X2 and X4 in each sequence are independently selected from the group consisting of G and A; X5 of SEQ ID NO: 104 or 176 is independently selected from the group consisting of N and S; X6 of SEQ ID NO: 104 or 176 is independently selected from the group consisting of Y and V; X7 of SEQ ID NO: 104 or 176 is independently selected from the group consisting of Y and N.

[0008] In some embodiments, the selected VH CDR1, 2, and 3 amino acid sequences and the selected VL CDR1, 2, and 3 amino acid sequences are any one of the following: (1) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively; (2) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 4, 5, and 6, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (3) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 7, 8, and 9, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; (4) According to the definition of Chothia, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 28, 29, and 30, respectively; (5) According to the definition of Chothia, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 13, 14, and 15, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (6) According to the definition of Chothia, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 16, 17, and 18, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively; (7) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (8) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; (9) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 4, 5, and 6, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively; (10) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 4, 5, and 6, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; (11) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 7, 8, and 9, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively; (12) According to the definition of Kabat, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 7, 8, and 9, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (13) According to the definition of Chothia, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (14) According to the definition of Chothia, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively; (15) According to the definition of Chothia, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 13, 14, and 15, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 28, 29, and 30, respectively; (16) According to the definition of Chothia, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 13, 14, and 15, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively; (17) According to the definition of Chothia, the VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 16, 17, and 18, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 28, 29, and 30, respectively; and (18) According to the definition of Chothia, the VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 16, 17, and 18, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively.

[0009] In some embodiments, the selected VH CDR1, 2, and 3 amino acid sequences and the selected VL CDR1, 2, and 3 amino acid sequences are any one of the following: (1) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 114, 115, and 116, respectively; (2) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 72, 73, and 74, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 114, 115, and 116, respectively; (3) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 72, 73, and 74, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 108, 109, and 110, respectively; (4) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 78, 79, and 80, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 114, 115, and 116, respectively; (5) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 75, 76, and 77, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 111, 112, and 113, respectively; (6) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 186, 187, and 188, respectively; (7) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 144, 145, and 146, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 186, 187, and 188, respectively; (8) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 144, 145, and 146, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 180, 181, and 182, respectively; (9) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 150, 151, and 152, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 186, 187, and 188, respectively; (10) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 147, 148, and 149, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 183, 184, and 185, respectively; (11) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 72, 73, and 74, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; (12) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 144, 145, and 146, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively; (13) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 219, 220, and 221, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (14) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 222, 223, and 224, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (15) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 225, 226, and 227, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (16) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 228, 229, and 230, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (17) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 231, 232, and 233, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (18) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 234, 235, and 236, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (19) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 237, 238, and 239, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (20) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 240, 241, and 242, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (21) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 243, 244, and 245, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (22) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 246, 247, and 248, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (23) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 249, 250, and 251, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (24) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 252, 253, and 254, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (25) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 255, 256, and 257, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; and (26) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 258, 259, and 260, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively.

[0010] In some embodiments, the antibody or antigen-binding fragment specifically binds to human CCR8, mouse CCR8, or monkey CCR8.

[0011] In some embodiments, the antibody or antigen-binding fragment is a human or humanized antibody or antigen-binding fragment thereof (eg, a human IgG1 antibody).

[0012] In some embodiments, the antibody or antigen-binding fragment is a single-chain variable fragment (scFv), a one-armed antibody, or a multispecific antibody (eg, a bispecific antibody).

[0013] In one aspect, the disclosure relates to a nucleic acid comprising a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, comprising a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VH binds to CCR8 when paired with a light chain variable region (VL); The amino acid sequences of the VH CDR1, 2, and 3 are as follows: (1) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 1, 2, and 3, respectively; (2) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 4, 5, and 6, respectively; (3) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 7, 8, and 9, respectively; (4) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 10, 11, and 12, respectively; (5) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 13, 14, and 15, respectively; (6) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 16, 17, and 18, respectively; (7) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 72, 73, and 74, respectively; (8) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 75, 76, and 77, respectively; (9) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 78, 79, and 80, respectively; (10) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 81, 82, and 83, respectively; (11) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 84, 85, and 86, respectively; (12) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 87, 88, and 89, respectively; (13) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 90, 91, and 92, respectively; (14) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 93, 94, and 95, respectively; (15) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 96, 97, and 98, respectively; (16) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 99, 100, and 101, respectively; (17) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 102, 103, and 104, respectively; (18) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 105, 106, and 107, respectively; (19) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 144, 145, and 146, respectively; (20) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 147, 148, and 149, respectively; (21) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 150, 151, and 152, respectively; (22) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 153, 154, and 155, respectively; (23) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 156, 157, and 158, respectively. (24) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 159, 160, and 161, respectively; (25) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 162, 163, and 164, respectively; (26) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 165, 166, and 167, respectively; (27) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 168, 169, and 170, respectively; (28) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 171, 172, and 173, respectively. (29) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 174, 175, and 176, respectively; (30) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 177, 178, and 179, respectively; (31) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 219, 220, and 221, respectively; (32) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 222, 223, and 224, respectively. (33) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 225, 226, and 227, respectively; (34) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 228, 229, and 230, respectively; (35) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 231, 232, and 233, respectively; (36) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 234, 235, and 236, respectively. (37) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 237, 238, and 239, respectively. (38) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 240, 241, and 242, respectively. (39) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 243, 244, and 245, respectively. (40) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 246, 247, and 248, respectively; (41) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 249, 250, and 251, respectively. (42) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 252, 253, and 254, respectively. (43) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 255, 256, and 257, respectively; and (44) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 258, 259, and 260, respectively. One of the following: The VL amino acid sequence is selected from the group consisting of SEQ ID NOs: 40, 41, 42, 66, 67, and 68.

[0014] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide, wherein the polypeptide comprises an immunoglobulin heavy chain or a fragment thereof, comprising a VH comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3, respectively.

[0015] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide, wherein the polypeptide comprises an immunoglobulin heavy chain or a fragment thereof, comprising a VH comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6, respectively.

[0016] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide, wherein the polypeptide comprises an immunoglobulin heavy chain or a fragment thereof, comprising a VH comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 7, 8, and 9, respectively.

[0017] In one aspect, the disclosure relates to a nucleic acid comprising a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, comprising a light chain variable region (VL) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VL binds to CCR8 when paired with a heavy chain variable region (VH); The amino acid sequences of the selected VL CDR1, 2, and 3 are as follows: (1) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 19, 20, and 21, respectively; (2) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 22, 23, and 24, respectively; (3) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 25, 26, and 27, respectively; (4) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 28, 29, and 30, respectively; (5) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 31, 32, and 33, respectively; (6) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 34, 35, and 36, respectively; (7) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 108, 109, and 110, respectively; (8) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 111, 112, and 113, respectively; (9) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 114, 115, and 116, respectively; (10) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 117, 118, and 119, respectively; (11) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 120, 121, and 122, respectively; (12) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 123, 124, and 125, respectively; (13) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 126, 127, and 128, respectively; (14) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 129, 130, and 131, respectively; (15) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 132, 133, and 134, respectively; (16) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 135, 136, and 137, respectively; (17) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 138, 139, and 140, respectively; (18) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 141, 142, and 143, respectively; (19) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 180, 181, and 182, respectively; (20) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 183, 184, and 185, respectively; (21) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 186, 187, and 188, respectively; (22) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 189, 190, and 191, respectively; (23) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 192, 193, and 194, respectively; (24) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 195, 196, and 197, respectively; (25) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 198, 199, and 200, respectively; (26) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 201, 202, and 203, respectively; (27) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 204, 205, and 206, respectively; (28) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 207, 208, and 209, respectively; (29) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 210, 211, and 212, respectively; and (30) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 213, 214, and 215, respectively. One of the following: The VH amino acid sequence is selected from the group consisting of SEQ ID NOs: 37, 38, 39, 63, 64, 65, 69, 70, 71, 216, 261, 262, 263, 264, 265, 266, and 267.

[0018] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide, wherein the polypeptide comprises an immunoglobulin light chain or a fragment thereof, comprising a VL comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 19, 20, and 21, respectively.

[0019] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide, wherein the polypeptide comprises an immunoglobulin light chain or a fragment thereof, comprising a VL comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 22, 23, and 24, respectively.

[0020] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide, wherein the polypeptide comprises an immunoglobulin light chain or a fragment thereof, comprising a VL comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 25, 26, and 27, respectively.

[0021] In some embodiments, the VH, when paired with the VL, specifically binds to human CCR8, mouse CCR8, or monkey CCR8, or the VL, when paired with the VH, specifically binds to human CCR8, mouse CCR8, or monkey CCR8.

[0022] In some embodiments, the immunoglobulin heavy chain or fragment thereof is a human or humanized immunoglobulin heavy chain or fragment thereof, and the immunoglobulin light chain or fragment thereof is a human or humanized immunoglobulin light chain or fragment thereof.

[0023] In some embodiments, the nucleic acid encodes a single-chain variable fragment (scFv), a one-armed antibody, or a multispecific antibody (eg, a bispecific antibody).

[0024] In some embodiments, the nucleic acid is cDNA.

[0025] In one aspect, the disclosure relates to a vector comprising any one or more of the nucleic acids described herein.

[0026] In one aspect, the disclosure relates to a vector comprising two nucleic acids described herein, said vector encoding a VH region and a VL region that binds CCR8 generically.

[0027] In one aspect, the disclosure relates to a pair of vectors, each vector comprising any one of the nucleic acids described herein, wherein the pair of vectors collectively encodes a VH region and a VL region that collectively bind to CCR8.

[0028] In one aspect, the disclosure relates to a cell comprising a vector described herein or a pair of vectors described herein.

[0029] In some embodiments, the cells are CHO cells.

[0030] In one aspect, the disclosure relates to a cell comprising any one or more of the nucleic acids described herein.

[0031] In one aspect, the disclosure relates to a cell comprising any two of the nucleic acids described herein.

[0032] In some embodiments, the two nucleic acids comprehensively encode a VH region and a VL region that comprehensively bind to CCR8.

[0033] In one aspect, the disclosure relates to a method for producing an antibody or antigen-binding fragment thereof, the method comprising: (a) culturing a cell described herein under conditions sufficient for said cell to produce said antibody or said antigen-binding fragment; (b) harvesting the antibody or antigen-binding fragment produced by the cell.

[0034] In one aspect, the disclosure provides an antibody or antigen-binding fragment thereof that binds to CCR8, a heavy chain variable region (VH) comprising an amino acid sequence that is at least 80% identical to a selected VH sequence, and a light chain variable region (VL) comprising an amino acid sequence that is at least 80% identical to a selected VL sequence; the selected VH sequence is selected from the group consisting of SEQ ID NOs: 37, 38, 39, 63, 64, 65, 69, 70, 71, 216, 261, 262, 263, 264, 265, 266, and 267; The selected VL sequence is selected from the group consisting of SEQ ID NOs: 40, 41, 42, 66, 67, and 68. The present invention relates to the above antibody or antigen-binding fragment thereof.

[0035] In some embodiments, the VH sequence and the VL sequence are any one of the following: (1) the VH comprises the sequence of SEQ ID NO: 37, and the VL comprises the sequence of SEQ ID NO: 40; (2) the VH comprises the sequence of SEQ ID NO: 38, and the VL comprises the sequence of SEQ ID NO: 41; (3) The VH comprises the sequence of SEQ ID NO: 39, and the VL comprises the sequence of SEQ ID NO: 42; (4) The VH comprises the sequence of SEQ ID NO: 37, and the VL comprises the sequence of SEQ ID NO: 41; (5) The VH comprises the sequence of SEQ ID NO: 37, and the VL comprises the sequence of SEQ ID NO: 42; (6) The VH comprises the sequence of SEQ ID NO: 38, and the VL comprises the sequence of SEQ ID NO: 40; (7) The VH comprises the sequence of SEQ ID NO: 38, and the VL comprises the sequence of SEQ ID NO: 42; (8) The VH comprises the sequence of SEQ ID NO: 39, and the VL comprises the sequence of SEQ ID NO: 40; (9) The VH comprises the sequence of SEQ ID NO: 39, and the VL comprises the sequence of SEQ ID NO: 41; (10) The VH comprises the sequence of SEQ ID NO: 37, and the VL comprises the sequence of SEQ ID NO: 68; (11) The VH comprises the sequence of SEQ ID NO: 63, and the VL comprises the sequence of SEQ ID NO: 68; (12) The VH comprises the sequence of SEQ ID NO: 63, and the VL comprises the sequence of SEQ ID NO: 66; (13) The VH comprises the sequence of SEQ ID NO: 65, and the VL comprises the sequence of SEQ ID NO: 68; (14) The VH comprises the sequence of SEQ ID NO: 64, and the VL comprises the sequence of SEQ ID NO: 67; (15) The VH comprises the sequence of SEQ ID NO: 63, and the VL comprises the sequence of SEQ ID NO: 42; (16) The VH comprises the sequence of SEQ ID NO: 216, and the VL comprises the sequence of SEQ ID NO: 42; (17) The VH comprises the sequence of SEQ ID NO: 261, and the VL comprises the sequence of SEQ ID NO: 41; (18) The VH comprises the sequence of SEQ ID NO: 262, and the VL comprises the sequence of SEQ ID NO: 41; (19) The VH comprises the sequence of SEQ ID NO: 263, and the VL comprises the sequence of SEQ ID NO: 41; (20) The VH comprises the sequence of SEQ ID NO: 264, and the VL comprises the sequence of SEQ ID NO: 41; (21) The VH comprises the sequence of SEQ ID NO: 265, and the VL comprises the sequence of SEQ ID NO: 41; (22) The VH comprises the sequence of SEQ ID NO: 266, and the VL comprises the sequence of SEQ ID NO: 41; and (23) The VH comprises the sequence of SEQ ID NO: 267, and the VL comprises the sequence of SEQ ID NO: 41.

[0036] In some embodiments, the antibody or antigen-binding fragment specifically binds to human CCR8, mouse CCR8, or monkey CCR8.

[0037] In some embodiments, the antibody or antigen-binding fragment is a human or humanized antibody or antigen-binding fragment thereof.

[0038] In some embodiments, the antibody or antigen-binding fragment is a single-chain variable fragment (scFv), a one-armed antibody, or a multispecific antibody (eg, a bispecific antibody).

[0039] In one aspect, the present disclosure relates to an antibody or antigen-binding fragment thereof that binds to CCR8, comprising a heavy chain variable region (VH) comprising VH CDR1, 2, and 3, and a light chain variable region comprising VL CDR1, 2, and 3, wherein the VH CDR1, 2, and 3 and the VL CDR1, 2, and 3 are identical to the complementarity-determining regions in an antibody or antigen-binding fragment thereof described herein.

[0040] In some embodiments, the VH CDR1, 2, 3 are identical to the complementarity determining regions of SEQ ID NO: 37, 38, 39, 63, 64, 65, 69, 70, 71, 216, 261, 262, 263, 264, 265, 266, or 267, and the VL CDR1, 2, 3 are identical to the complementarity determining regions of SEQ ID NO: 40, 41, 42, 66, 67, or 68.

[0041] In one aspect, the present disclosure relates to antibodies, or antigen-binding fragments thereof, that cross-compete with the antibodies, or antigen-binding fragments thereof, described herein.

[0042] In one aspect, the present disclosure relates to an antibody drug conjugate comprising an antibody, or antigen-binding fragment thereof, described herein, covalently attached to a therapeutic agent.

[0043] In some embodiments, the therapeutic agent is a cytotoxic or cytostatic agent.

[0044] In one aspect, the present disclosure relates to a method of treating a subject afflicted with cancer, the method comprising administering to the subject a therapeutically effective amount of an antibody or antigen-binding fragment thereof described herein, or a composition comprising an antibody drug conjugate described herein.

[0045] In some embodiments, the subject is a patient with a solid tumor.

[0046] In some embodiments, the cancer is breast cancer, malignant melanoma, non-small cell lung cancer, lung adenocarcinoma, colon cancer, or colorectal cancer (eg, colorectal adenocarcinoma).

[0047] In some embodiments, the cancer is resistant to PD-1 pathway inhibitor treatment.

[0048] In some embodiments, the method further comprises administering to the subject a PD-1 pathway inhibitor.

[0049] In some embodiments, the PD-1 pathway inhibitor is an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, or cemiplimab).

[0050] In some embodiments, the PD-1 pathway inhibitor is an anti-PD-L1 antibody (e.g., atezolizumab, avelumab, or durvalumab).

[0051] In one aspect, the present disclosure relates to a method of reducing tumor growth rate, the method comprising contacting tumor cells with an effective amount of an antibody or antigen-binding fragment thereof described herein, or a composition comprising an antibody drug conjugate described herein.

[0052] In one aspect, the present disclosure relates to a method of killing tumor cells, the method comprising contacting the tumor cells with an effective amount of an antibody or antigen-binding fragment thereof described herein, or a composition comprising an antibody drug conjugate described herein.

[0053] In one aspect, the present disclosure relates to a pharmaceutical composition comprising an antibody or antigen-binding fragment thereof described herein and a pharmaceutically acceptable carrier.

[0054] In one aspect, the present disclosure relates to a pharmaceutical composition comprising an antibody drug conjugate described herein and a pharmaceutically acceptable carrier.

[0055] As used herein, the term "cancer" refers to cells capable of autonomous proliferation. Examples of such cells include cells with an abnormal state or pathology characterized by rapidly proliferating cell proliferation. This term is intended to include cancerous growths, e.g., tumors, oncogenic processes, metastatic tissues, and malignantly transformed cells, tissues, or organs, regardless of the type of tissue change or stage of invasiveness. It also includes malignant tumors of various organ systems, such as the respiratory, cardiovascular, renal, reproductive, hematological, nervous, liver, gastrointestinal, and endocrine systems, as well as adenocarcinomas, including most colon cancers, renal cell carcinoma, prostate and / or testicular cancer, non-small cell lung cancer, and small intestine cancer. "Naturally occurring" cancers include any cancer that is not experimentally induced by implanting cancer cells into a subject, such as naturally occurring cancers, cancers caused by exposure of a patient to a carcinogen, cancers resulting from the insertion of a transgenic oncogene or the knockout of a tumor suppressor gene, and cancers caused by infectious diseases, e.g., viral infections. The term "carcinoma" is art-recognized and refers to a malignant tumor of epithelial or endocrine tissue. It also includes carcinosarcomas, which include malignant tumors composed of carcinomatous and sarcomatous tissue. "Adenocarcinoma" refers to a carcinoma derived from glandular tissue or in which the tumor cells form recognizable glandular structures. The term "sarcoma" is art-recognized and refers to a malignant tumor of mesenchymal origin. The term "hematopoietic neoplastic disease" includes diseases involving hyperplastic / neoplastic cells of hematopoietic origin. Hematopoietic neoplastic diseases can arise from the myeloid, lymphoid, or erythroid lineages, or their precursor cells. Blood cancers are cancers that begin in blood-forming tissues, such as the bone marrow, or in cells of the immune system. Examples of blood cancers include leukemia, lymphoma, and multiple myeloma.

[0056] As used herein, the term "antibody" refers to any antigen-binding molecule that contains at least one (e.g., one, two, three, four, five, or six) complementarity-determining region (CDR) (e.g., any of the three CDRs from an immunoglobulin light chain or any of the three CDRs from an immunoglobulin heavy chain) and is capable of specifically binding to an epitope. Non-limiting examples of antibodies include monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), single-chain antibodies, chimeric antibodies, human antibodies, and humanized antibodies. In some embodiments, an antibody can contain the Fc region of a human antibody. The term antibody also includes derivatives, e.g., bispecific antibodies, single-chain antibodies, heavy-chain antibodies, diabodies, linear antibodies, and multispecific antibodies formed from antibody fragments.

[0057] As used herein, the term "antigen-binding fragment" refers to a portion of a full-length antibody, which portion of the antibody is capable of specifically binding to an antigen. In some embodiments, the antigen-binding fragment contains at least one variable domain (e.g., a heavy chain variable domain or a light chain variable domain). Non-limiting examples of antibody fragments include, for example, Fab, Fab', F(ab')2, VHH, and Fv fragments.

[0058] As used herein, the term "human antibody" means an antibody encoded by endogenous nucleic acid derived from a human (e.g., a rearranged human immunoglobulin heavy or light chain locus). In some embodiments, a human antibody is obtained from a human or produced in human cell culture (e.g., in human hybridoma cells). In some embodiments, a human antibody is produced in a non-human cell (e.g., a mouse or hamster cell line). In some embodiments, a human antibody is produced in a bacterial cell or yeast cell. In some embodiments, a human antibody is produced in a transgenic non-human animal (e.g., a cow) containing unrearranged or rearranged human immunoglobulin loci (e.g., a heavy or light chain human immunoglobulin locus).

[0059] As used herein, the term "chimeric antibody" refers to an antibody that contains sequences present in at least two different species (e.g., an antibody derived from two different mammalian species, such as a human and a murine antibody). A non-limiting example of a chimeric antibody is an antibody that contains variable domain sequences (e.g., all or part of the light and / or heavy chain variable domain sequences) of a non-human (e.g., murine) antibody and the constant domain of a human antibody. Further examples of chimeric antibodies are described herein and are well known in the art.

[0060] As used herein, the term "humanized antibody" refers to a non-human antibody that contains minimal sequence derived from non-human (e.g., murine) immunoglobulin and contains sequence derived from human immunoglobulin. In a non-limiting example, a humanized antibody is a human antibody (recipient antibody) in which hypervariable (e.g., CDR) region residues of the recipient antibody are replaced by hypervariable (e.g., CDR) region residues from a non-human antibody (e.g., donor antibody) such as a mouse, rat, or rabbit antibody possessing the desired specificity, affinity, and capacity. In some embodiments, Fv framework residues of the human immunoglobulin are replaced by corresponding non-human (e.g., murine) immunoglobulin residues. In some embodiments, humanized antibodies can contain residues not found in the recipient antibody or the donor antibody. These modifications can be made to further refine antibody performance. In some embodiments, a humanized antibody contains substantially all of at least one, and typically two, variable domains, in which all or substantially all of the hypervariable loops (CDRs) correspond to those of a non-human (e.g., murine) immunoglobulin, and all or substantially all of the framework regions are those of a human immunoglobulin. The humanized antibody can also contain at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin. Humanized antibodies can be produced using molecular biology methods well known in the art. Non-limiting examples of methods for generating humanized antibodies are described herein.

[0061] As used herein, the term "single-chain antibody" means a single polypeptide containing at least two immunoglobulin variable domains (e.g., the variable domains of a mammalian immunoglobulin heavy or light chain) that are capable of specifically binding to an antigen. Non-limiting examples of single-chain antibodies are described herein.

[0062] As used herein, the term "multimeric antibody" refers to an antibody that contains four or more (e.g., six, eight, or ten) immunoglobulin variable domains. In some embodiments, the multimeric antibody is capable of cross-linking one target molecule (e.g., CCR8) to at least one second target molecule (e.g., PD-L1) on the surface of a mammalian cell (e.g., a Treg cell and a cancer cell).

[0063] As used herein, the terms "subject" and "patient" are used interchangeably throughout the specification and refer to an animal, human, or non-human to whom treatment according to the methods of the present invention is provided. Veterinary and non-veterinary applications are contemplated by this disclosure. A human patient can be an adult human or a juvenile human (e.g., a human under the age of 18). In addition to humans, treatment subjects include, but are not limited to, mice, rats, hamsters, guinea pigs, rabbits, ferrets, cats, dogs, and primates. For example, non-human primates (e.g., monkeys, chimpanzees, gorillas, etc.), rodents (e.g., rats, mice, gerbils, hamsters, ferrets, rabbits), lagomorphs, swine (e.g., pigs, minipigs), equines, canines, felines, bovines, and other domestic, livestock, and zoo animals.

[0064] As used herein, the phrase "specifically binds" when referring to an antibody means that the antibody interacts preferably with its target molecule (e.g., CCR8) over other molecules because the interaction depends on the presence of a specific structure (i.e., an antigenic determinant or epitope) on the target molecule; in other words, the reagent recognizes and binds to molecules containing a specific structure, rather than all molecules in general. An antibody that specifically binds to a target molecule can be referred to as a target-specific antibody. For example, an antibody that specifically binds to a CCR8 molecule can be referred to as a CCR8-specific antibody or an anti-CCR8 antibody.

[0065] As used herein, the terms "polypeptide," "peptide," and "protein" are used interchangeably and refer to polymers of amino acids of any length, of at least two amino acids.

[0066] As used herein, the terms "polynucleotide," "nucleic acid molecule," and "nucleic acid sequence" are used interchangeably herein and refer to polymers of nucleotides of any length, of at least two nucleotides, including, but not limited to, DNA, RNA, DNA / RNA hybrids, and modifications thereof.

[0067] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Methods and materials for use in the present invention are described herein; other suitable methods and materials well-known in the art can also be used. The materials, methods, and examples are illustrative only and are not intended to be limiting. All publications, patent applications, patents, sequences, database entries, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control.

[0068] Other features and advantages of the invention will become apparent from the following detailed description and drawings, and from the claims. [Brief explanation of the drawings]

[0069] [Figure 1] 1 shows the effect of anti-CCR8 antibodies on tumor growth in vivo in a colon cancer model. [Figure 2A] The CDR sequences corresponding to anti-human CCR8 mAbs 11B12, 11F5, and 11F8, and CDR-modified anti-human CCR8 mAbs, as defined by Kabat's definition, are shown. [Figure 2B] Same as above [Figure 2C] Same as above [Figure 3A] The CDR sequences corresponding to anti-human CCR8 mAbs 11B12, 11F5, and 11F8, and CDR-modified anti-human CCR8 mAbs, as defined by Chothia's definition, are shown. [Figure 3B] Same as above [Figure 3C] Same as above [Figure 4-1] Some of the relevant amino acid sequences discussed in this disclosure are shown. [Figure 4-2] Same as above [Figure 4-3] Same as above [Figure 4-4] Same as above [Figure 5-1] Some VH and VL combinations in this disclosure are shown. [Figure 5-2] Same as above [Figure 5-3] Same as above [Figure 6] Shown are the mean tumor volumes for each group of B-hPD-1 / hPD-L1 / hCCR8 mice subcutaneously injected with MC38 cells (murine colon cancer cells) and treated with PBS (G1), pembrolizumab analog (G2), anti-CCR8 antibody 11F5 (G3), or a combination of pembrolizumab analog and 11F5 (G4). [Figure 7]Shown are the mean tumor volumes for each group of B-hPD-1 / hPD-L1 / hCCR8 mice subcutaneously injected with MC38 cells and treated with PBS (G1), pembrolizumab analog (G2), anti-CCR8 antibody 11F5-SI (G3), a combination of pembrolizumab analog and 11F5-SI (G4), anti-CCR8 antibody 11B12-SI (G5), or a combination of pembrolizumab analog and 11B12-SI (G6). [Figure 8] 1 shows the blocking of hCCL1 binding to hCCR8 by anti-CCR8 antibody 11F5 and CDR3-modified antibodies of 11F5, as tested by a calcium (Ca) flux assay using human CCR8-expressing 293T cells. DETAILED DESCRIPTION OF THE INVENTION

[0070] Chemokines and chemokine receptors play an important role in the immune defense system by regulating leukocyte migration, activation, differentiation, and survival. The 50 human chemokines are classified into C, CC, CXC, and CX3C classes based on the number and spacing of conserved cysteine ​​residues in their N-terminal regions. Chemokine receptors belong to family A of G protein-coupled receptors (GPCRs), characterized by seven transmembrane (7TM) helical domains. There are 18 human chemokine receptors, activated by different chemokine subfamilies: C (XCR1), CC (CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, CCR10), CXC (CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6), or CX3C (CX3CR1), and four atypical decoy chemokine receptors (ACKR: ACKR1, ACKR2, ACKR3 / CXCR7, and ACKR4). Chemokine receptors are thought to interact with chemokine ligands via a two-step binding mechanism. In this mechanism, (i) the structured C-terminal region of the chemokine first binds to the N-terminal region and extracellular loop (ECL) of the receptor (chemokine recognition site 1, CRS1), and (ii) the unstructured N-terminus of the chemokine then targets the 7TM helix bundle (chemokine recognition site 2, CRS2) and stabilizes the receptor in an active conformation that facilitates intracellular signaling, for example, by G proteins and arrestins. Chemokine receptors play an important role in cell migration and are therefore important therapeutic targets for inflammatory diseases and cancer.

[0071] Chemokine (CC motif) receptor 8 (CCR8, or CDw198) is a seven-transmembrane G protein-coupled receptor (GPCR) and belongs to the CC subfamily of chemokine receptors. CCR8 is one of the most important chemokine receptors, primarily expressed on Tregs, with minor expression on T helper 2 cells and monocytes. CCR8 has four known ligands: CCL1, CCL8, CCL16, and CCL18. Human CCL1 is the primary ligand for human CCR8, binding simultaneously to extracellular loop 2 and the N-terminal domain of the receptor. Specifically, CCL1 enhances the immunosuppressive activity of Tregs by recruiting CCR8, FOXP3, and IL-10 through a positive feedback mechanism. CCR8 has also been reported to be correlated with poor prognosis in non-small cell lung cancer and colorectal cancer.

[0072] CCR8 is selectively upregulated in tumor Treg cells of multiple solid cancers, but is rarely observed in peripheral blood mononuclear cells (PBMCs) or normal organ Treg cells. Treg cells are immunosuppressive and generally suppress or downregulate the induction and proliferation of effector T cells. These cells play an important role in maintaining self-tolerance and immune homeostasis. Tumor Treg cells can suppress antitumor immunity in the tumor microenvironment. High CCR8 expression is often associated with reduced survival in cancer patients. Therefore, CCR8 is a potential target for Treg cells. Targeting CCR8 to inhibit and / or deplete Treg cells has several advantages. First, because CCR8 expression is restricted to Treg cells but not to CD8 T cells, the antitumor activity of T effector cells is not attenuated. Second, systemic Treg cell depletion can often lead to autoimmune responses. CCR8 is rarely expressed on peripheral blood mononuclear cells (PBMCs) or normal organ Treg cells, reducing the risk of autoimmune responses. Indeed, treatment with a rat anti-mouse CCR8 blocking antibody can produce significant antitumor activity and improve long-term survival in a mouse model of colon tumors. Therefore, CCR8 is a superior target among various known Treg targets, as its expression has been found to be selectively observed on Treg cells in tumors. Anti-CCR8 antibodies can exhibit monotherapy activity in PD-1-resistant settings and can also restore PD-1 activity as a combination partner.

[0073] For a detailed description of CCR8 and its functions, see, for example, Plitas G. et al., "Regulatory T cells exhibit distinct features in human breast cancer," Immunity 45.5 (2016): 1122-1134; Villarreal. et al., "Targeting CCR8 induces protective antitumor immunity and enhances vaccine-induced responses in colon cancer," Cancer research 78.18 (2018): 5340-5348; Arimont et al., "Structural analysis of chemokine receptor-ligand interactions," Journal of Medicinal Chemistry 60.12 (2017): 4735-4779; and Li et al., "Clinical and prognostic significance of CC chemokine receptor type 8 protein expression in gastrointestinal stromal tumors," World Journal of Gastroenterology. 26.31(2020):4656, which references are incorporated herein by reference in their entirety.

[0074] The present disclosure provides anti-CCR8 antibodies, antigen-binding fragments thereof, and methods for inhibiting tumor growth and treating cancer using these anti-CCR8 antibodies and antigen-binding fragments, and in particular, the present disclosure provides fully human anti-CCR8 antibodies (e.g., antibodies produced in mice having a humanized heavy chain immunoglobulin locus and a humanized light chain immunoglobulin locus).

[0075] Anti-CCR8 antibodies and antigen-binding fragments The present disclosure provides antibodies and antigen-binding fragments thereof that specifically bind to CCR8. The antibodies and antigen-binding fragments described herein are capable of binding to CCR8. In some embodiments, these antibodies can block the binding of CCL1 to CCR8, thereby inhibiting the CCL1-induced suppressive function of tumor Treg cells. In some embodiments, these antibodies can inhibit CCL1-CCR8-mediated downstream signaling. In some embodiments, these antibodies can initiate complement-dependent cytotoxicity (CDC) or antibody-dependent cellular cytotoxicity (ADCC). In some embodiments, these antibodies and antigen-binding fragments thereof can block the interaction between CCR8 and a CCR8 ligand (e.g., CCL1, CCL8, CCL16, or CCL18).

[0076] The disclosure provides, for example, murine anti-CCR8 antibodies 11B12, 11F5, and 11F8, chimeric antibodies thereof, and human or humanized antibodies thereof.

[0077] CDR sequences for 11B12 and antibodies derived from 11B12 (e.g., humanized antibodies) include the heavy chain variable domain CDRs set forth in SEQ ID NOS: 1, 2, and 3, and the light chain variable domain CDRs set forth in SEQ ID NOS: 19, 20, and 21, as defined by the Kabat definition. CDRs can also be defined by the Chothia definition. Under the Chothia definition, the heavy chain variable domain CDR sequences are set forth in SEQ ID NOS: 10, 11, and 12, and the light chain variable domain CDR sequences are set forth in SEQ ID NOS: 28, 29, and 30.

[0078] Similarly, CDR sequences for 11F5 and antibodies derived from 11F5 include, as defined by the Kabat definition, heavy chain variable domain CDRs SEQ ID NOS: 4, 5, 6, and light chain variable domain CDRs SEQ ID NOS: 22, 23, 24. Under the Chothia definition, heavy chain variable domain CDR sequences are set forth in SEQ ID NOS: 13, 14, 15, and light chain variable domain CDRs are set forth in SEQ ID NOS: 31, 32, 33.

[0079] Similarly, CDR sequences for 11F8 and antibodies derived from 11F8 include the heavy chain variable domain CDRs SEQ ID NOs: 7, 8, 9 and light chain variable domain CDRs SEQ ID NOs: 25, 26, 27, as defined by the Kabat definition. Under the Chothia definition, the heavy chain variable domain CDR sequences are set forth in SEQ ID NOs: 16, 17, 18, and the light chain variable domain CDRs are set forth in SEQ ID NOs: 34, 35, 36.

[0080] The amino acid sequence for the heavy chain variable region of the 11B12 antibody is set forth in SEQ ID NO: 37. The amino acid sequence for the light chain variable region of the 11B12 antibody is set forth in SEQ ID NO: 40. The amino acid sequence for the heavy chain variable region of the 11F5 antibody is set forth in SEQ ID NO: 38. The amino acid sequence for the light chain variable region of the 11F5 antibody is set forth in SEQ ID NO: 41. The amino acid sequence for the heavy chain variable region of the 11F8 antibody is set forth in SEQ ID NO: 39. The amino acid sequence for the light chain variable region of the 11F8 antibody is set forth in SEQ ID NO: 42.

[0081] Amino acid sequences for the heavy and light chain variable regions of the humanized antibody are also provided. Because there are several methods for humanizing mouse antibodies (e.g., sequences can be modified with different amino acid substitutions), multiple versions of the humanized antibody heavy and light chain sequences may exist. In some embodiments, the humanized heavy chain variable region is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 37, 38, or 39. In some embodiments, the humanized light chain variable region is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 40, 41, or 42. The heavy chain variable region sequence is paired with the corresponding light chain variable region sequence to bind to CCR8.

[0082] Percent humanization refers to the percent identity of a heavy or light chain variable region sequence compared to human antibody sequences in the International Immunogenetics Database (IMGT). A top hit means that the heavy or light chain variable region sequence is closer to a particular species than to other species. For example, a top hit with humans means that the sequence is closer to humans than to other species. A top hit with humans and cynomolgus monkeys means that the sequence has the same percent identity with the human sequence and the cynomolgus monkey sequence, and these percent identities are the highest compared to sequences from other species. In some embodiments, the percent humanization is greater than 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, or 95%. Detailed descriptions of how to determine the humanization rate and how to determine the top hits are well known in the art and are described, for example, in Jones, et al. "The INNs and outs of antibody nonproprietary names," MAbs. Vol. 8. No. 1. Taylor & Francis, 2016, which references are incorporated herein by reference in their entirety. A high humanization rate often has various advantages, such as being safer and more effective in humans, more likely to be tolerated by human subjects, and / or less likely to have side effects. In some embodiments, the antibody is a fully human antibody.

[0083] Furthermore, in some embodiments, the antibodies or antigen-binding fragments thereof described herein may also contain one, two, or three heavy chain variable region CDRs selected from the group consisting of SEQ ID NOs: 1 to 3, SEQ ID NOs: 4 to 6, SEQ ID NOs: 7 to 9, SEQ ID NOs: 10 to 12, SEQ ID NOs: 13 to 15, and SEQ ID NOs: 16 to 18, and / or one, two, or three light chain variable region CDRs selected from the group consisting of SEQ ID NOs: 19 to 21, SEQ ID NOs: 22 to 24, SEQ ID NOs: 25 to 27, SEQ ID NOs: 28 to 30, SEQ ID NOs: 31 to 33, and SEQ ID NOs: 34 to 36.

[0084] In some embodiments, the antibody may have a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the CDR1 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to the amino acid sequence of a selected VH CDR1; the CDR2 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to the amino acid sequence of a selected VH CDR2; and the CDR3 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to the amino acid sequence of a selected VH CDR3. In some embodiments, the antibody may have a light chain variable region (VL) comprising CDR1, 2, and 3, wherein the CDR1 region comprises or consists of an amino acid sequence at least 80%, 85%, 90%, or 95% identical to the amino acid sequence of a selected VL CDR1, the CDR2 region comprises or consists of an amino acid sequence at least 80%, 85%, 90%, or 95% identical to the amino acid sequence of a selected VL CDR2, and the CDR3 region comprises or consists of an amino acid sequence at least 80%, 85%, 90%, or 95% identical to the amino acid sequence of a selected VL CDR3. The amino acid sequences of the selected VH CDR1, 2, and 3 and the selected VL CDR1, 2, and 3 are shown in Figures 2A-2C (Kabat CDRs) and 3A-3C (Chothia CDRs).

[0085] In some embodiments, the present disclosure relates to CDR-modified VH or VL. In some embodiments, "DG" in the VH CDR2 of an antibody described herein is replaced with "SG," "AG," or "DA" to create a CDR-modified VH. For example, in one embodiment, "DG" in the VH CDR2 of 11B12 is replaced with "SG," and the resulting CDR-modified VH is designated 11B12-SG VH. The sequences of some of these CDR-modified VHs are shown in Figure 4.

[0086] In some embodiments, the "DG" in the VL CDR1 of an antibody described herein is replaced with "SG," "AG," or "DA" to create a CDR-modified VL. For example, in one embodiment, the "DG" in the VL CDR1 of 11B12 is replaced with SG, and the resulting CDR-modified VL is designated 11B12-SG VL. The sequences of some of these CDR-modified VLs are shown in Figure 4.

[0087] In some embodiments, the present disclosure relates to a VH with framework region modifications (FR-modified). In some embodiments, "DNS" in the framework of the VH of an antibody described herein is replaced with "DNA" to create a FR-modified VH. For example, in one example, "DNS" in the framework of the VH of 11B12 is replaced with "DNA," and the resulting FR-modified VH is designated 11B12-DNA VH. The sequences of some of these FR-modified VHs are shown in Figure 4 (e.g., SEQ ID NOS: 69-71). Thus, in some embodiments, the antibody has a S74A mutation (Kabat numbering). In some embodiments, the amino acid at position 74 (Kabat numbering) is A.

[0088] In some embodiments, a VH has both FR modifications (e.g., DNA) and CDR modifications (e.g., SG). In one example, "DNS" in the framework of 11B12 VH is replaced with "DNA," and "DG" in 11B12 VH CDR2 is replaced with "SG," and the resulting FR- and CDR-modified VH is designated 11B12-SG-DNA VH.

[0089] In some embodiments, an 11F5 CDR3-modified VH can be generated. In some embodiments, the amino acid "N" at position 3 in the VH CDR3 of 11F5 (e.g., SEQ ID NO: 6 or 15) is substituted with "S". In some embodiments, the amino acid "Y" at position 5 in the VH CDR3 of 11F5 (e.g., SEQ ID NO: 6 or 15) is substituted with "V". In some embodiments, the amino acid "Y" at position 6 in the VH CDR3 of 11F5 (e.g., SEQ ID NO: 6 or 15) is substituted with "N". For example, the amino acid "N" at position 3 in the VH CDR3 of 11F5 can be substituted with "S", and the resulting CDR-modified VH is designated 11F5-N3S VH; the amino acid "Y" at position 5 in the VH CDR3 of 11F5 can be substituted with "V", and the resulting CDR-modified VH is designated 11F5-Y5V VH; the amino acid "Y" at position 6 in the VH CDR3 of 11F5 can be substituted with "N", and the resulting CDR-modified VH is designated 11F5-Y6N VH; the amino acid "N" at position 3 and the amino acid "Y" at position 5 in the VH CDR3 of 11F5 can be substituted with "S" and "V", respectively, and the resulting CDR-modified VH is designated 11F5-N3S-Y5V VH; In CDR3, the amino acid "N" at position 3 and the amino acid "Y" at position 6 can be replaced with "S" and "N", respectively, and the resulting CDR-modified VH is designated 11F5-N3S-Y6N VH; in VH CDR3 of 11F5, the amino acid "Y" at position 5 and the amino acid "Y" at position 6 can be replaced with "V" and "N", respectively, and the resulting CDR-modified VH is designated 11F5-Y5V-Y6N VH; and in VH CDR3 of 11F5, the amino acid "N" at position 3, the amino acid "Y" at position 5, and the amino acid "Y" at position 6 can be replaced with "S", "V", and "N", respectively, and the resulting CDR-modified VH is designated 11F5-N3S-Y5V-Y6N VH. The sequences of these CDR-modified VHs are shown in Figure 4.

[0090] In some embodiments, the VH of any antibody described herein can be paired with the VL of any antibody described herein to form an antibody. For example, in one embodiment, the VH of 11B12 is paired with the VL of 11F8, and the resulting antibody is designated 11B12-11F8. In one embodiment, the VH of 11B12-SG is paired with the VL of 11F8-SG, and the resulting antibody is designated 11B12-SG-11F8-SG. In one embodiment, the VH of 11B12-SG is paired with the VL of 11B12-SG, and the resulting antibody is designated 11B12-SG-11B12-SG. The binding affinities of some of these antibodies are shown in Tables 15, 16, and 19. Some of the disclosed VH and VL combinations are listed in Figure 5. As shown in Figure 5, at least 163 VH and VL combinations have been disclosed. In addition, the VH CDRs and VL CDRs in the VH and VL can also be combined, as shown in FIG.

[0091] In one aspect, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to CCR8 (chemokine (CC motif) receptor 8), a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VH CDR1 region comprises an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of a selected VH CDR1; and the VH CDR2 region comprises an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of a selected VH CDR1; a heavy chain variable region comprising an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of the selected VH CDR3; a light chain variable region (VL) comprising CDR1, 2, and 3, wherein the VL CDR1 region comprises an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of a selected VL CDR1; and the VL CDR2 region comprises an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of a selected VL CDR2; and a light chain variable region as described above, wherein the CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR3, Among them, the amino acid sequences of the selected VH CDR1, 2, and 3 are any one of the following: (1) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 1, 2, and 3, respectively; (2) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 4, 5, and 6, respectively; (3) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 7, 8, and 9, respectively; (4) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 10, 11, and 12, respectively; (5) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 13, 14, and 15, respectively; (6) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 16, 17, and 18, respectively; (7) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 72, 73, and 74, respectively; (8) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 75, 76, and 77, respectively; (9) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 78, 79, and 80, respectively; (10) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 81, 82, and 83, respectively; (11) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 84, 85, and 86, respectively; (12) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 87, 88, and 89, respectively; (13) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 90, 91, and 92, respectively; (14) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 93, 94, and 95, respectively; (15) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 96, 97, and 98, respectively; (16) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 99, 100, and 101, respectively; (17) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 102, 103, and 104, respectively; (18) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 105, 106, and 107, respectively; (19) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 144, 145, and 146, respectively; (20) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 147, 148, and 149, respectively; (21) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 150, 151, and 152, respectively; (22) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 153, 154, and 155, respectively; (23) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 156, 157, and 158, respectively; (24) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 159, 160, and 161, respectively; (25) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 162, 163, and 164, respectively; (26) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 165, 166, and 167, respectively; (27) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 168, 169, and 170, respectively; (28) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 171, 172, and 173, respectively; (29) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 174, 175, and 176, respectively; (30) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 177, 178, and 179, respectively; (31) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 219, 220, and 221, respectively; (32) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 222, 223, and 224, respectively; (33) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 225, 226, and 227, respectively; (34) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 228, 229, and 230, respectively; (35) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 231, 232, and 233, respectively; (36) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 234, 235, and 236, respectively; (37) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 237, 238, and 239, respectively; (38) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 240, 241, and 242, respectively; (39) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 243, 244, and 245, respectively; (40) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 246, 247, and 248, respectively; (41) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 249, 250, and 251, respectively; (42) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 252, 253, and 254, respectively; (43) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 255, 256, and 257, respectively; and (44) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 258, 259, and 260, respectively; Among them, the amino acid sequences of the selected VL CDR1, 2, and 3 are any one of the following:

[0092] (1) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 19, 20, and 21, respectively; (2) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 22, 23, and 24, respectively; (3) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 25, 26, and 27, respectively; (4) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 28, 29, and 30, respectively; (5) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 31, 32, and 33, respectively; (6) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 34, 35, and 36, respectively; (7) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 108, 109, and 110, respectively; (8) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 111, 112, and 113, respectively; (9) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 114, 115, and 116, respectively; (10) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 117, 118, and 119, respectively; (11) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 120, 121, and 122, respectively; (12) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 123, 124, and 125, respectively; (13) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 126, 127, and 128, respectively; (14) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 129, 130, and 131, respectively; (15) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 132, 133, and 134, respectively; (16) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 135, 136, and 137, respectively; (17) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 138, 139, and 140, respectively; (18) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 141, 142, and 143, respectively; (19) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 180, 181, and 182, respectively; (20) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 183, 184, and 185, respectively; (21) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 186, 187, and 188, respectively; (22) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 189, 190, and 191, respectively; (23) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 192, 193, and 194, respectively; (24) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 195, 196, and 197, respectively; (25) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 198, 199, and 200, respectively; (26) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 201, 202, and 203, respectively; (27) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 204, 205, and 206, respectively; (28) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 207, 208, and 209, respectively; (29) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 210, 211, and 212, respectively; and (30) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 213, 214, and 215, respectively.

[0093] In one aspect, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to CCR8 (chemokine (CC motif) receptor 8), a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VH CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR1, the VH CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR2, and the VH CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR3; a light chain variable region (VL) comprising CDR1, 2, and 3, wherein the VL CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR1, the VL CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR2, and the VL CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR3, Among them, the amino acid sequences of the selected VH CDR1, 2, and 3 are any one of the following:

[0094] (1) the selected VH CDR1 is SYVMH (SEQ ID NO: 99), the selected VH CDR2 is VISYX1X2SDKYYADSVKG (SEQ ID NO: 100), and the selected VH CDR3 is GRGYRYGQSYYYGMDV (SEQ ID NO: 101); (2) the selected VH CDR1 is SYVVH (SEQ ID NO: 102), the selected VH CDR2 is VISYX1X2DNKFYADSVKG (SEQ ID NO: 103), and the selected VH CDR3 is GRX5YX6X7YYGLDV (SEQ ID NO: 104); (3) the selected VH CDR1 is TYVMH (SEQ ID NO: 105), the selected VH CDR2 is VISYX1X2NNKYYADSVKG (SEQ ID NO: 106), and the selected VH CDR3 is GRSYVNYYGLDV (SEQ ID NO: 107); (4) The selected VH CDR1 is GFSFSSY (SEQ ID NO: 171), the selected VH CDR2 is SYX1X2SD (SEQ ID NO: 172), and the selected VH CDR3 is GRGYRYGQSYYYGMDV (SEQ ID NO: 173); (5) the selected VH CDR1 is GFTFSSY (SEQ ID NO: 174), the selected VH CDR2 is SYX1X2DN (SEQ ID NO: 175), and the selected VH CDR3 is GRX5YX6X7YYGLDV (SEQ ID NO: 176); and (6) The selected VH CDR1 is GFTFSTY (SEQ ID NO: 177), the selected VH CDR2 is SYX1X2NN (SEQ ID NO: 178), and the selected VH CDR3 is GRSYVNYYGLDV (SEQ ID NO: 179); Among them, the amino acid sequences of the selected VL CDR1, 2, and 3 are any one of the following:

[0095] (1) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 135), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 136), and the selected VL CDR3 is MQSIKLPLT (SEQ ID NO: 137); (2) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 138), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 139), and the selected VL CDR3 is MQSLKVPPT (SEQ ID NO: 140); (3) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 141), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 142), and the selected VL CDR3 is MQSVKIPLT (SEQ ID NO: 143); (4) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 207), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 208), and the selected VL CDR3 is MQSIKLPLT (SEQ ID NO: 209); (5) the selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 210), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 211), and the selected VL CDR3 is MQSLKVPPT (SEQ ID NO: 212); and (6) The selected VL CDR1 is KSSQSLLYSX3X4KTYLY (SEQ ID NO: 213), the selected VL CDR2 is EVSNRFS (SEQ ID NO: 214), and the selected VL CDR3 is MQSVKIPLT (SEQ ID NO: 215); wherein X1 and X3 in each sequence are independently selected from the group consisting of D, S, and A; wherein X2 and X4 in each sequence are independently selected from the group consisting of G and A; wherein X5 of SEQ ID NO: 104 or 176 is independently selected from the group consisting of N and S; wherein X6 of SEQ ID NO: 104 or 176 is independently selected from the group consisting of Y and V; wherein X7 of SEQ ID NO: 104 or 176 is independently selected from the group consisting of Y and N.

[0096] In some embodiments, humanization may involve limited modifications in the CDRs. However, these modifications have only a limited effect on binding affinity. In some embodiments, the antibodies or antigen-binding fragments described herein may contain a heavy chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 1 with 0, 1, or 2 amino acid insertions, deletions, or substitutions, SEQ ID NO: 2 with 0, 1, or 2 amino acid insertions, deletions, or substitutions, or SEQ ID NO: 3 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0097] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a light chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 19 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 20 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 21 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0098] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a heavy chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 4 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 5 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 6 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0099] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a light chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 22 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 23 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 24 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0100] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a heavy chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 7 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 8 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 9 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0101] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a light chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 25 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 26 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 27 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0102] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a heavy chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 10 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 11 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 12 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0103] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a light chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 28 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 29 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 30 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0104] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a heavy chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 13 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 14 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 15 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0105] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a light chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 31 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 32 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 33 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0106] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a heavy chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 16 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; SEQ ID NO: 17 with 0, 1, or 2 amino acid insertions, deletions, or substitutions; or SEQ ID NO: 18 with 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0107] In some embodiments, the antibodies or antigen-binding fragments described herein may contain a light chain variable domain containing one, two, or three of the CDRs of SEQ ID NO: 34 with zero, one, or two amino acid insertions, deletions, or substitutions; SEQ ID NO: 35 with zero, one, or two amino acid insertions, deletions, or substitutions; or SEQ ID NO: 36 with zero, one, or two amino acid insertions, deletions, or substitutions.

[0108] Insertions, deletions, and substitutions can occur within the CDR sequences or at either or both ends of the CDR sequences. In some embodiments, CDRs are determined according to the Kabat definition. In some embodiments, CDRs are determined according to the Chothia definition.

[0109] In some embodiments, there is no more than one insertion, deletion, or substitution. In some embodiments, there is no more than two insertions, deletions, or substitutions. In some embodiments, there are no more than three insertions, deletions, or substitutions. In some embodiments, there are no more than four insertions, deletions, or substitutions. In some embodiments, the VH is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 37-39, 63-65, 69-71, 216, and 261-267. In some embodiments, the VL is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 40-42 and 66-68.

[0110] The present disclosure also provides an antibody or antigen-binding fragment thereof that binds to CCR8. The antibody or antigen-binding fragment thereof comprises a heavy chain variable region (VH) comprising, or consisting of, an amino acid sequence at least 80%, 85%, 90%, or 95% identical to a selected VH sequence, and a light chain variable region (VL) comprising, or consisting of, an amino acid sequence at least 80%, 85%, 90%, or 95% identical to a selected VL sequence. In some embodiments, the selected VH sequence is SEQ ID NO: 37 and the selected VL sequence is SEQ ID NO: 40. In some embodiments, the selected VH sequence is SEQ ID NO: 38 and the selected VL sequence is SEQ ID NO: 41. In some embodiments, the selected VH sequence is SEQ ID NO: 39 and the selected VL sequence is SEQ ID NO: 42. In some embodiments, the selected VH sequence is SEQ ID NO: 261 and the selected VL sequence is SEQ ID NO: 41. In some embodiments, the selected VH sequence is SEQ ID NO: 262 and the selected VL sequence is SEQ ID NO: 41. In some embodiments, the selected VH sequence is SEQ ID NO:263 and the selected VL sequence is SEQ ID NO:41. In some embodiments, the selected VH sequence is SEQ ID NO:264 and the selected VL sequence is SEQ ID NO:41. In some embodiments, the selected VH sequence is SEQ ID NO:265 and the selected VL sequence is SEQ ID NO:41. In some embodiments, the selected VH sequence is SEQ ID NO:266 and the selected VL sequence is SEQ ID NO:41. In some embodiments, the selected VH sequence is SEQ ID NO:267 and the selected VL sequence is SEQ ID NO:41.

[0111] To compare two amino acid sequences or determine the percent homology of two nucleic acid sequences, the sequences are aligned (e.g., gaps may be introduced into one or both of the first and second amino acid or nucleic acid sequences to ensure optimal alignment, and non-homologous sequences may be ignored). The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. If a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, the molecules are identical at that position. The percent homology between two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps that need to be introduced to optimally align the two sequences and the length of each gap. Illustratively, sequence comparison and determination of percent homology between two sequences can be performed using the Blossum62 scoring matrix with a gap penalty of 12, a gap extension penalty of 4, and a frameshift gap penalty of 5.

[0112] The present disclosure also provides a nucleic acid comprising a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or an immunoglobulin light chain. The immunoglobulin heavy chain or immunoglobulin light chain comprises the CDRs shown in Figures 2A-2C or 3A-3C, or has the sequence shown in Figure 4. When the polypeptide pairs with a corresponding polypeptide (e.g., a corresponding heavy chain variable region or a corresponding light chain variable region), the paired polypeptide binds to CCR8 (e.g., human CCR8).

[0113] Anti-CCR8 antibodies and antigen-binding fragments can also be antibody variants (including derivatives and conjugates) of antibodies or antibody fragments, as well as multispecific (e.g., bispecific) antibodies or antibody fragments. Additional antibodies provided herein include polyclonal, monoclonal, multispecific (multimeric, e.g., bispecific), human, chimeric (e.g., human-mouse chimeras), single-chain, and intracellularly produced antibodies (i.e., intrabodies), and antigen-binding fragments thereof. The antibody or antigen-binding fragment thereof can be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass. In some embodiments, the antibody or antigen-binding fragment thereof is an IgG antibody or antigen-binding fragment thereof.

[0114] Antibody fragments can be used in the provided methods as long as they retain the desired affinity and specificity of the full-length antibody. Thus, antibody fragments that bind to CCR8 retain the ability to bind to CCR8. Fv fragments are antibody fragments that contain the complete antigen recognition and binding site. This region consists of a tightly associated dimer of one heavy-chain variable domain and one light-chain variable domain, which can be essentially covalently linked, for example, in scFv. In this configuration, the three CDRs of each variable domain interact to define an antigen-binding site on the surface of the VH-VL dimer. Collectively, the six CDRs, or a subset thereof, confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of an Fv containing only three CDRs specific for a given antigen) can still recognize and bind to an antigen, although usually with lower affinity than the entire binding site. Single-chain Fv or (scFv) antibody fragments comprise the VH and VL domains (or regions) of an antibody, which are present in a single polypeptide chain. Generally, the scFv polypeptide further comprises a polypeptide linker between the VH and VL domains that enables the scFv to form the desired structure for antigen binding. In some embodiments, the polypeptide comprises a linker having the sequence GGGS (SEQ ID NO: 61) or GGGSGGGS (SEQ ID NO: 62).

[0115] The present disclosure also provides antibodies or antigen-binding fragments thereof that cross-compete with any of the antibodies or antigen-binding fragments described herein. Cross-competition assays are well known in the art and are described, for example, in Moore et al., "Antibody cross-competition analysis of the human immunodeficiency virus type 1 gp120 exterior envelope glycoprotein," Journal of Virology 70.3 (1996):1863-1872, which are incorporated herein by reference in their entireties. In one aspect, the present disclosure also provides antibodies or antigen-binding fragments thereof that bind to the same epitope or region as any of the antibodies or antigen-binding fragments described herein. Epitope binding assays are well known in the art and are described, for example, in Estep et al., "High throughput solution-based measurement of antibody-antigen affinity and epitope binning," MAbs. Vol. 5. No. 2. Taylor & Francis, 2013, which are incorporated herein by reference in their entireties.

[0116] Antibodies and antigen-binding fragments The present disclosure provides various antibodies and antigen-binding fragments thereof derived from the anti-CCR8 antibodies described herein. Generally, antibodies (also called immunoglobulins) are composed of two classes of polypeptide chains: light chains and heavy chains. Non-limiting examples of antibodies of the present disclosure can be intact four immunoglobulin chain antibodies, comprising two heavy chains and two light chains. The heavy chain of the antibody can be of any isotype, including IgM, IgG, IgE, IgA, or IgD, or a subisotype, including IgG1, IgG2, IgG2a, IgG2b, IgG3, IgG4, IgE1, IgE2, etc. The light chain can be a kappa light chain or a lambda light chain. The antibody can comprise two identical copies of the light chain and two identical copies of the heavy chain. Heavy chains, each containing one variable domain (or variable region, VH) and multiple constant domains (or constant regions), are linked to each other via disulfide bonds within their constant domains to form the antibody "stem." Light chains, each containing one variable domain (or variable region, VL) and one constant domain (or constant region), are linked to one heavy chain via disulfide bonds. The variable region of each light chain aligns with the variable region of the heavy chain to which it is attached. Both light and heavy chain variable regions contain three hypervariable regions sandwiched between more conserved framework regions (FR).

[0117] The hypervariable regions, known as complementarity-determining regions (CDRs), form the loops that comprise the antigen-binding surface of an antibody. The four framework regions largely adopt a β-sheet structure, and the CDRs form loops that connect, and in some cases form part of, the β-sheet structure. The CDRs of each chain are held in close proximity by the framework regions and, together with the CDRs of the other chain, contribute to the formation of the antigen-binding region.

[0118] Methods for identifying CDR regions of antibodies by analyzing their amino acid sequences are well known, and several definitions of CDRs are commonly used: the Kabat definition is based on sequence variability, and the Chothia definition is based on the location of structural loop regions. These methods and definitions are described, for example, in Martin, "Protein sequence and structure analysis of antibody variable domains," Antibody engineering, Springer Berlin Heidelberg, 2001, pp. 422-439; Abhinandan, et al., "Analysis and improvements to Kabat and structurally correct numbering of antibody variable domains," Molecular immunology 45.14 (2008): 3832-3839; Wu, TT and Kabat, EA (1970) J. Exp. Med. 132: 211-250; Martin et al., Methods Enzymol. 203: 121-53 (1991); Morea et al., Biophys Chem. 68 (1-3): 9-16 (Oct. 1997); Morea et al., J. Mol. Biol. 275 (2): 269-94 (Jan. 1998); Chothia et al. al., Nature 342(6252):877-83 (Dec. 1989), Ponomarenko and Bourne, BMC Structural Biology 7:64 (2007), which references are incorporated herein by reference in their entirety.

[0119] CDRs are important for recognizing the epitope of an antigen. As used herein, "epitope" refers to the smallest portion of a target molecule that can be specifically bound by the antigen-binding domain of an antibody. The minimum size of an epitope can be approximately 3, 4, 5, 6, or 7 amino acids, but these amino acids do not need to be in a contiguous linear sequence in the primary structure of the antigen, as the epitope may depend on the three-dimensional structure of the antigen based on the secondary and tertiary structure of the antigen.

[0120] In some embodiments, antibodies are intact immunoglobulin molecules (e.g., IgG1, IgG2a, IgG2b, IgG3, IgM, IgD, IgE, IgA). The IgG subclasses (IgG1, IgG2, IgG3, and IgG4) are highly conserved and differ in their constant regions, particularly the hinge and upper CH2 domains. The sequences and differences of IgG subclasses are well known in the art and are described, for example, in Vidarsson, et al., "IgG subclasses and allotypes: from structure to effector functions." Frontiers in immunology 5 (2014); Irani, et al., "Molecular properties of human IgG subclasses and their implications for designing therapeutic monoclonal antibodies against infectious diseases." Molecular immunology 67.2 (2015): 171-182; and Shakib, Farouk, ed., The human IgG subclasses: molecular analysis of structure, function and regulation. Elsevier, 2016, the entire contents of which are incorporated herein by reference.

[0121] An antibody can also be an immunoglobulin molecule from any species (e.g., human, rodent, murine, camelid). Antibodies disclosed herein include, but are not limited to, polyclonal, monoclonal, monospecific, multispecific antibodies, and chimeric antibodies comprising an immunoglobulin binding domain fused to another polypeptide. The term "antigen-binding domain" or "antigen-binding fragment" refers to any portion of an antibody that retains the specific binding activity of the intact antibody, i.e., any portion of an antibody that is capable of specifically binding to an epitope on the intact antibody's target molecule. This includes, for example, Fab, Fab', F(ab')2, and variants of these fragments. Thus, in some embodiments, an antibody or antigen-binding fragment thereof can be, for example, an scFv, Fv, Fd, dAb, diabody, bispecific scFv, diabody, linear antibody, single-chain antibody molecule, multispecific antibody formed from antibody fragments, and any polypeptide comprising a binding domain that is, or is homologous to, an antibody-binding domain. Non-limiting examples of antigen-binding domains include, for example, the heavy and / or light chain CDRs of an intact antibody, the heavy and / or light chain variable regions of an intact antibody, the full-length heavy or light chain of an intact antibody, or individual CDRs derived from either the heavy or light chain of an intact antibody.

[0122] Also provided are antibody fragments suitable for use in the methods described herein. Fab fragments contain the variable and constant domains of the light chain and the variable and first constant domain (CH1) of the heavy chain. F(ab')2 antibody fragments comprise a pair of Fab fragments covalently linked, generally near their carboxy termini, by hinge cysteines between them. Other chemical linkages of antibody fragments are also well known in the art.

[0123] Diabodies are small antibody fragments with two antigen-binding sites, comprising a VH connected to a VL (VH and VL) in the same polypeptide chain. By using a linker that is too short to allow pairing between the two domains on the same chain, the domains can pair with complementary domains on another chain and create two antigen-binding sites.

[0124] Linear antibodies comprise a pair of tandem Fd segments (VH-CH1-VH-CH1) that, together with complementary light chain polypeptides, form a pair of antigen-binding regions. Linear antibodies may be bispecific or monospecific.

[0125] The antibodies and antibody fragments of the disclosure can be modified in the Fc region to have desired effector functions or serum half-lives.

[0126] Antibody multimerization can be achieved by natural antibody aggregation or by chemical or recombinant conjugation techniques well known in the art. For example, a certain percentage of purified antibody preparations (e.g., purified IgG1 molecules) naturally form protein aggregates containing antibody homodimers and other higher order antibody multimers.

[0127] Alternatively, antibody homodimers can be formed by chemical coupling techniques well known in the art. For example, heterobifunctional cross-linkers, including but not limited to SMCC (succinimidyl 4-(maleimidomethyl)cyclohexane-1-carboxylate) and SATA (N-succinimidyl S-acetylthioacetate), can be used to form antibody multimers. An exemplary procedure for forming antibody homodimers is described in Ghetie et al. (Proc. Natl. Acad. Sci. USA 94:7509-7514, 1997). Antibody homodimers can be converted to Fab'2 homodimers by pepsin digestion. Another method for forming antibody homodimers is by using the self-affinity T15 peptide, as described in Zhao et al. (J. Immunol. 25:396-404, 2002).

[0128] In some embodiments, multispecific antibodies are bispecific antibodies. Bispecific antibodies can be produced by engineering the interface between a pair of antibody molecules to maximize the percentage of heterodimers recovered from recombinant cell culture. For example, the interface can comprise at least a portion of the CH3 domain of an antibody constant domain. In this method, one or more small amino acid side chains from the interface of a first antibody molecule are replaced with larger side chains (e.g., tyrosine or tryptophan). By replacing the large amino acid side chains with smaller ones (e.g., alanine or threonine), alternative "cavities" of the same or similar size as the large side chains are created in the interface of the second antibody molecule. This provides a mechanism for increasing the yield of heterodimers over other unwanted end-products, such as homodimers. This method is described, for example, in WO 96 / 27011, the entire disclosure of which is incorporated by reference.

[0129] Bispecific antibodies include cross-linked or "heteroconjugate" antibodies. For example, one of the antibodies in the heteroconjugate can be conjugated to avidin, the other to biotin. Heteroconjugate antibodies can be made using any convenient cross-linking method. Suitable cross-linking agents and techniques are well known in the art and are disclosed in U.S. Pat. No. 4,676,980, the disclosure of which is incorporated herein by reference in its entirety.

[0130] Methods for generating bispecific antibodies from antibody fragments are also well known in the art. For example, bispecific antibodies can be prepared using chemical linkage. Brennan et al. (Science 229:81, 1985) describe a procedure in which intact antibodies are proteolytically cleaved to generate F(ab')2 fragments. These fragments are reduced in the presence of the dithiol complexing agent sodium arsenite to stabilize vicinal dithiols and prevent intermolecular disulfide formation. The resulting Fab' fragments are then converted to thionitrobenzoate (TNB) derivatives. One of the Fab'TNB derivatives is then reconverted to the Fab' thiol by reduction with mercaptoethylamine and mixed with an equimolar amount of another Fab'TNB derivative to form the bispecific antibody.

[0131] Any of the antibodies or antigen-binding fragments described herein can be conjugated to a stabilizing molecule (e.g., a molecule that increases the half-life of the antibody or antigen-binding fragment thereof in a subject or in solution). Non-limiting examples of stabilizing molecules include polymers (e.g., polyethylene glycol) or proteins (e.g., serum albumins, such as human serum albumin). Conjugation of a stabilizing molecule can increase the half-life or prolong the biological activity of the antibody or antigen-binding fragment in vitro (e.g., in tissue culture or when stored as a pharmaceutical composition) or in vivo (e.g., in humans).

[0132] In some embodiments, the antibodies or antigen-binding fragments described herein can be conjugated to a therapeutic agent. The antibody-drug conjugate comprising the antibody or antigen-binding fragment thereof can be covalently or non-covalently bound to the therapeutic agent. In some embodiments, the therapeutic agent is a cytotoxic or cytostatic agent (e.g., cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, teniposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxyanthracene, maytansinoids (such as DM-1 and DM-4), dione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, puromycin, epirubicin, and cyclophosphamide and analogs).

[0133] In some embodiments, the antigen-binding fragment can form part of a chimeric antigen receptor (CAR). In some embodiments, the chimeric antigen receptor is a fusion of a single-chain variable fragment (scFv) described herein fused to the CD3ζ transmembrane and endodomain. In some embodiments, the chimeric antigen receptor also comprises intracellular signaling domains from various costimulatory protein receptors (e.g., CD28, 41BB, ICOS). In some embodiments, the chimeric antigen receptor comprises multiple signaling domains, e.g., CD3z-CD28-41BB or CD3z-CD28-OX40, for increased efficacy. Thus, in one aspect, the present disclosure further provides a cell (e.g., a T cell) expressing a chimeric antigen receptor described herein.

[0134] In some embodiments, an scFv has one heavy chain variable domain and one light chain variable domain, hi some embodiments, an scFv has two heavy chain variable domains and two light chain variable domains.

[0135] In some embodiments, the antibodies or antigen-binding fragments thereof described herein comprise a heavy chain constant region (e.g., CH1, CH2, and / or CH3) selected from mouse IgG1, IgG2a, and IgG2b. In some embodiments, the antibodies or antigen-binding fragments thereof described herein comprise a light chain constant region (e.g., CL) selected from mouse kappa or lambda CL.

[0136] In some embodiments, the antibodies or antigen-binding fragments thereof described herein comprise a heavy chain constant region (e.g., CH1, CH2, and / or CH3) selected from human IgG (e.g., human IgG1, IgG2, IgG3, or IgG4). In some embodiments, the antibodies or antigen-binding fragments thereof described herein comprise a light chain constant region (e.g., CL) selected from human kappa or lambda CL.

[0137] Antibody characteristics The antibodies or antigen-binding fragments thereof described herein can block the binding between CCR8 and a CCR8 ligand (e.g., CCL1). In some embodiments, the antibodies can inhibit the CCL1-induced suppressive function of tumor Treg cells by binding to CCR8. In some embodiments, the antibodies can upregulate an immune response. In some embodiments, the antibodies can deplete Treg cells (e.g., tumor Treg cells). In some embodiments, the antibodies have a reduced risk of inducing an autoimmune response and a reduced risk of inducing systemic Treg cell depletion.

[0138] In some embodiments, the antibodies or antigen-binding fragments thereof described herein increase anti-tumor immunity by at least or about 50%, 60%, 70%, 80%, 90%, 95%, 98%, or 100%.

[0139] In some embodiments, the antibodies or antigen-binding fragments thereof described herein have a reduced risk of inducing an autoimmune response or causing systemic Treg cell depletion. In some embodiments, the antibodies or antigen-binding fragments thereof described herein have at least or about a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% reduced risk of inducing autoimmunity compared to other therapeutic agents that target Tregs (e.g., anti-CCR4, CTLA4, OX40, or GITR antibodies).

[0140] In some embodiments, the antibodies or antigen-binding fragments thereof described herein can reduce the activity or number of Treg cells (e.g., Treg cells in the tumor and / or in the tumor microenvironment) by at least or about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%, e.g., compared to the activity or number of Treg cells when the antibody or antigen-binding fragment thereof is not administered.

[0141] In some embodiments, the antibodies or antigen-binding fragments thereof described herein can increase an immune response, activity or number of immune cells (e.g., T cells, CD8+ T cells, CD4+ T cells, macrophages, antigen-presenting cells) by at least or about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 2-fold, 3-fold, 5-fold, 10-fold, or 20-fold compared to when the antibody or antigen-binding fragment thereof is not administered.

[0142] In some embodiments, the antibodies or antigen-binding fragments thereof described herein can effectively increase the Teff:Treg ratio, e.g., by at least or about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 2-fold, 3-fold, 5-fold, 10-fold, or 20-fold, compared to when the antibody or antigen-binding fragment thereof is not administered.

[0143] In some embodiments, the antibody (or antigen-binding fragment thereof) binds to CCR8 (e.g., human CCR8, monkey CCR8 (e.g., cynomolgus monkey (Macaca fascicularis)), mouse CCR8, and / or chimeric CCR8) in a 0.1 s -1 Less than 0.01s -1 Less than 0.001s -1 Less than 0.0001s -1 Less than or equal to 0.00001s -1 In some embodiments, the koff is less than 0.01 s -1 Super, 0.001s -1 Super, 0.0001s -1 Super, 0.00001s -1 Over or 0.000001s -1 It's super.

[0144] In some embodiments, the kinetic association rate (k) is 1×10 2 / Ms super, 1×10 3 / Ms super, 1×10 4 / Ms super, 1×10 5 / Ms or more than 1 × 10 6 In some embodiments, the kinetic association rate (k) is greater than 1×10 5 / Ms less than 1 × 10 6 / Ms or less than 1 x 10 7 / Ms is less than.

[0145] Affinity can be estimated from the quotient of the kinetic rate constants (K = k / k). In some embodiments, K is greater than or equal to 1 x 10 -6 Less than M, 1 x 10 -7 Less than M, 1 x 10 -8 Less than M, 1 x 10 -9 Less than M or 1 x 10 -10 In some embodiments, the KD is less than 50 nM, 30 nM, 20 nM, 15 nM, 10 nM, 9 nM, 8 nM, 7 nM, 6 nM, 5 nM, 4 nM, 3 nM, 2 nM, or 1 nM. In some embodiments, the KD is less than 1 x 10 -7Super M, 1×10 -8 Super M, 1×10 -9 Super M, 1×10 -10 Super M, 1×10 -11 Over M or 1 x 10 -12 It's over M.

[0146] In some embodiments, the antibody or antigen-binding fragment thereof binds to human CCR8 at Y17, F21, D97, W99, F101, G102, C106, G179, C183, W194, K195, I196, and / or L278.

[0147] In some embodiments, the binding affinity of an antibody or antigen-binding fragment thereof described herein can be measured by FACS. An exemplary procedure is shown in Example 4. In some embodiments, the EC50 is less than 25 μg / mL, less than 15 μg / mL, less than 10 μg / mL, less than 5 μg / mL, less than 2.5 μg / mL, less than 1 μg / mL, less than 0.5 μg / mL, less than 0.4 μg / mL, less than 0.3 μg / mL, less than 0.2 μg / mL, less than 0.15 μg / mL, less than 0.1 μg / mL, or less than 0.05 μg / mL. In some embodiments, the EC50 is about 0.05 to 0.5 μg / mL, about 0.1 to 1 μg / mL, about 0.1 to 0.8 μg / mL, about 0.1 to 0.5 μg / mL, about 0.2 to 2 μg / mL, about 0.2 to 1 μg / mL, or about 0.3 to 5 μg / mL.

[0148] In some embodiments, blockage of hCCL1 binding to hCCR8 by an antibody or antigen-binding fragment thereof described herein can be measured using a Fluorescence Imaging Plate Reader (FLIPR) calcium 6 assay kit. In some embodiments, the EC50 is less than 30 nM, 20 nM, 10 nM, 5 nM, 2.5 nM, 1.5 nM, or 1 nM. In some embodiments, the EC50 is about 1-10 nM, 1.5-20 nM, or 2.5-30 nM. In some embodiments, maximal inhibition is greater than 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 95%. In some embodiments, maximal inhibition is less than 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 95%.

[0149] In some embodiments, the antibody can bind to any one or more of human CCR2, human CCR4, human CCR5, human CCR8, and human CX3CR1. In some embodiments, the antibody cannot bind to any one or more of human CCR2, human CCR4, human CCR5, human CCR8, and human CX3CR1.

[0150] In some embodiments, antibody-dependent cellular cytotoxicity (ADCC) can be determined by reporter gene bioassay (ADCC reporter bioassay). In some embodiments, the EC50 is less than 25 μg / mL, less than 15 μg / mL, less than 10 μg / mL, less than 5 μg / mL, less than 2.5 μg / mL, less than 1 μg / mL, less than 0.5 μg / mL, less than 0.4 μg / mL, less than 0.3 μg / mL, less than 0.2 μg / mL, less than 0.15 μg / mL, less than 0.1 μg / mL, less than 0.05 μg / mL, less than 0.025 μg / mL, less than 0.01 μg / mL, less than 0.005 μg / mL, or less than 0.0025 μg / mL. In some embodiments, the EC50 is about 0.0025 to 0.05 μg / mL, about 0.005 to 0.1 μg / mL, about 0.005 to 0.05 μg / mL, about 0.005 to 0.01 μg / mL, about 0.01 to 0.2 μg / mL, or about 0.025 to 0.3 μg / mL.

[0151] Common techniques for measuring the affinity of an antibody for an antigen include, for example, flow cytometry, FACS, ELISA, RIA, and surface plasmon resonance (SPR). In some embodiments, the antibody binds to human CCR8 (SEQ ID NO: 43).

[0152] In some embodiments, the thermal stability is determined. The antibodies or antigen-binding fragments described herein can have a Tm of 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, or 95°C or greater. In some embodiments, the Tm is less than 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, or 95°C.

[0153] In some embodiments, the antibody has a percent tumor growth inhibition (TGI%) of more than 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190%, or 200%. In some embodiments, the antibody has a percent tumor growth inhibition of less than 60%, 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190%, or 200%. TGI% can be determined, for example, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 days after the start of treatment, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months after the start of treatment. As used herein, tumor growth inhibition percentage (TGI%) is calculated using the following formula: TGI(%)=[1-(Ti-T0) / (Vi-V0)]×100 Ti is the mean tumor volume in the treatment group on day i. T0 is the mean tumor volume in the treatment group on day 0. Vi is the mean tumor volume in the control group on day i. V0 is the mean tumor volume in the control group on day 0.

[0154] In some embodiments, the antibodies or antigen-binding fragments thereof described herein block the binding of CCR8 to its ligand (e.g., CCL1). In some embodiments, the antibodies or antigen-binding fragments thereof described herein inhibit the CCL-1-induced suppressive function of Treg cells in tumors.

[0155] In some embodiments, the antibody or antigen-binding fragment can induce complement-dependent cytotoxicity (CDC) and / or antibody-dependent cellular cytotoxicity (ADCC) and kill cells expressing CCR8 (e.g., Treg cells).

[0156] In some embodiments, the antibody or antigen-binding fragment has a functional Fc region. In some embodiments, the effector function of the functional Fc region is antibody-dependent cellular cytotoxicity (ADCC). In some embodiments, the antibody or antigen-binding fragment is capable of inducing complement-mediated cytotoxicity (CMC).

[0157] In some embodiments, the Fc region is human IgG1, human IgG2, human IgG3, or human IgG4, hi some embodiments, the antibody is a human IgG1 antibody.

[0158] In some embodiments, the antibody or antigen-binding fragment does not have a functional Fc region. For example, the antibody or antigen-binding fragment is a Fab, Fab', F(ab')2, or Fv fragment. In some embodiments, the Fc region has a LALA mutation (L234A and L235A mutations in EU numbering) or a LALA-PG mutation (L234A, L235A, P329G mutations in EU numbering).

[0159] In some embodiments, the Fc region is human IgG1, human IgG2, human IgG3, or human IgG4. In some embodiments, the antibody is a human IgG1 antibody, optionally with an SI mutation, a LALA mutation, an N297A mutation, a YTE mutation, and / or a FLAA mutation. In some embodiments, the antibody is a human IgG4 antibody, optionally with an SI mutation, a LALA mutation, an N297A mutation, a YTE mutation, and / or a FLAA mutation.

[0160] In some embodiments, the antibody or antigen-binding fragment does not have a functional Fc region. For example, the antibody or antigen-binding fragment is a Fab, Fab', F(ab')2, or Fv fragment. In some embodiments, the Fc region has a LALA mutation (L234A and L235A mutations according to EU numbering) or a LALA-PG mutation (L234A, L235A, P329G mutations according to EU numbering). In some embodiments, the Fc region has a FLAA mutation (F234A and L235A mutations according to EU numbering). In some embodiments, the Fc region has an SI mutation (S239D and I332E mutations according to EU numbering). In some embodiments, the Fc region has an N297A mutation according to EU numbering. In some embodiments, the Fc region has a YTE mutation (M252Y, S254T, and T256E mutations according to EU numbering).

[0161] Method for producing anti-CCR8 antibody Isolated membrane protein of human CCR8 can be used as an immunogen to generate antibodies using standard techniques for polyclonal and monoclonal antibody preparation. Polyclonal antibodies can be raised in animals by multiple injections (e.g., subcutaneous or intraperitoneal) of the antigenic peptide or protein. In some embodiments, the antigenic peptide or protein is injected with at least one adjuvant. In some embodiments, the antigenic peptide or protein can be conjugated to an agent that is immunogenic in the species being immunized. Animals can be injected with the antigenic peptide or protein two or more times (e.g., two, three, or four times).

[0162] The full-length polypeptide or protein can be used, or an antigenic peptide fragment thereof can be used as an immunogen. The antigenic peptide of the protein contains at least 8 (e.g., at least 10, 15, 20, or 30) amino acid residues of the amino acid sequence of CCR8 and encompasses an epitope of the protein such that antibodies raised against the peptide form specific immune complexes with the protein. As mentioned above, the full-length sequence of human CCR8 is well known in the art (SEQ ID NO: 43).

[0163] The immunogen is typically used to prepare antibodies by immunizing a suitable subject (e.g., a human or transgenic animal expressing at least one human immunoglobulin locus). An appropriate immunogenic preparation can contain, for example, a recombinantly expressed or chemically synthesized polypeptide (e.g., a fragment of human CCR8). The preparation can further include an adjuvant, such as Freund's complete or incomplete adjuvant, or a similar immunostimulant.

[0164] Polyclonal antibodies can be prepared as described above by immunizing a suitable subject with a CCR8 polypeptide or its antigenic peptide (e.g., a portion of CCR8) as an immunogen. Antibody titers in immunized subjects can be monitored over time by standard techniques, such as enzyme-linked immunosorbent assay (ELISA), using immobilized CCR8 polypeptide or peptide. If desired, antibody molecules can be isolated from the mammal (e.g., from the blood) and further purified by well-known techniques, such as protein G or protein A chromatography, to obtain an IgG fraction. At an appropriate time after immunization, e.g., when the specific antibody titer is at its highest, antibody-producing cells are harvested from the immunized animal and used to prepare monoclonal antibodies by standard techniques, such as the hybridoma technique originally described by Kohler et al. (Nature 256:495-497, 1975), the human B cell hybridoma technique (Kozbor et al., Immunol. Today 4:72, 1983), the EBV-hybridoma technique (Cole et al., Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pp. 77-96, 1985), or the trioma technique. Hybridoma production techniques are well known (see generally, Current Protocols in Immunology, 1994, Coligan et al. (Eds.), John Wiley & Sons, Inc., New York, NY)). For example, hybridoma cells producing monoclonal antibodies are detected by screening the hybridoma culture supernatants for antibodies that bind the polypeptide or epitope of interest using a standard ELISA assay.

[0165] The antibodies disclosed herein can be derived from any species of animal, including mammals. Non-limiting examples of natural antibodies include antibodies derived from humans, primates, such as monkeys and apes, cows, pigs, horses, sheep, camelids (e.g., camels and llamas), chickens, goats, and rodents (e.g., rats, mice, hamsters, and rabbits), including transgenic rodents that have been genetically engineered to produce human antibodies.

[0166] Human and humanized antibodies include antibodies having variable and constant regions derived from (or having the same amino acid sequences derived from) human germline immunoglobulin sequences. Human antibodies can include, for example, amino acid residues within the CDRs that are not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo).

[0167] A humanized antibody typically has a human framework (FR) into which non-human CDRs have been grafted. Thus, a humanized antibody has one or more amino acid sequences introduced into it from a non-human source. These non-human amino acid residues are often referred to as "import" residues, which are typically taken from an "import" variable domain. Humanization can essentially be performed, for example, by substituting rodent CDRs or CDR sequences with the corresponding sequences of a human antibody. These methods are described, for example, in Jones et al. "Replacing the complementarity-determining regions in a human antibody with those from a mouse," Nature 321.6069 (1986):522; Riechmann et al. "Reshaping human antibodies for therapy," Nature 332.6162 (1988):323; and Dall'Acqua et al. "Antibody humanization by framework shuffling," Methods 36.1 (2005):43-60, the entire contents of which are incorporated by reference. Thus, a "humanized" antibody is a chimeric antibody in which substantially less than an intact human V domain has been substituted by the corresponding sequence from a non-human species. In practice, humanized antibodies are typically murine antibodies in which some CDR residues and some FR residues have been substituted by residues from analogous sites in human antibodies.

[0168] The selection of human VH and VL domains used in the production of humanized antibodies is crucial to reducing immunogenicity. According to the so-called "best-fit" method, the sequence of the V domain of a mouse antibody is screened against the entire library of known human domain sequences. The human sequence that is closest to the mouse sequence is then accepted as the human FR for the humanized antibody (Sims et al., "A humanized CD18 antibody can block function without cell destruction," The Journal of Immunology 151.4 (1993):2296-2308; Chothia, et al., "Canonical structures for the hypervariable regions of immunoglobulins," Journal of molecular biology 196.4 (1987):901-917).

[0169] Furthermore, it is important to humanize antibodies while retaining high specificity and affinity for the antigen and other favorable biological properties. To achieve this goal, humanized antibodies can be prepared by a process of analysis of the parental sequences and various conceptual humanized products using three-dimensional models of the parental and humanized sequences. Three-dimensional immunoglobulin models are commonly available and are familiar to those skilled in the art. Computer programs are available that can illustrate and display probable three-dimensional conformations of selected candidate immunoglobulin sequences. These permit analysis of the possible role of residues in the functioning of the candidate immunoglobulin sequence, i.e., analysis of residues that influence the ability of the candidate immunoglobulin to bind to its antigen. In this way, FR residues can be selected and combined from the recipient and import sequences to achieve desired antibody characteristics, such as increased affinity for the target antigen.

[0170] Typically, an amino acid sequence variant of a human, humanized, or chimeric anti-CCR8 antibody contains an amino acid sequence that has at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% percent identity with the sequence present in the light or heavy chain of the original antibody.

[0171] In some embodiments, mice with a humanized heavy chain immunoglobulin locus and a humanized kappa chain immunoglobulin locus (e.g., RenMab TM The heavy chain immunoglobulin locus is a region on a chromosome that contains the gene for the heavy chain of an antibody. Examples of the locus include the human IGHV (variable) gene, the human IGHD (diversity) gene, the human IGHJ (joining) gene, and the mouse heavy chain constant domain gene. The kappa chain immunoglobulin locus is a region on a chromosome that contains the gene encoding the light chain (kappa chain) of an antibody. Examples of the kappa chain immunoglobulin locus include the human IGKV (variable) gene, the human IGKJ (joining) gene, and the mouse light chain constant domain gene. RenMab TM A detailed description of the mice can be found in PCT / CN2020 / 075698 or US20200390073A1, which references are incorporated herein in their entirety.

[0172] In some embodiments, mice with a humanized heavy chain immunoglobulin locus and a humanized kappa chain immunoglobulin locus (e.g., RenLite TM The heavy chain immunoglobulin locus is a region on a chromosome that contains the gene for the heavy chain of an antibody. Examples of the locus include the human IGHV (variable) gene, the human IGHD (diversity) gene, the human IGHJ (joining) gene, and the mouse heavy chain constant domain gene. The kappa chain immunoglobulin locus is a region on a chromosome that contains the gene encoding the common light chain. Examples of the kappa chain immunoglobulin locus include the human IGKV (variable) gene, the human IGKJ (joining) gene, and the mouse light chain constant domain gene. RenLite TMA detailed description of the mice can be found in PCT / CN2021 / 097652, which references are incorporated herein in their entirety.

[0173] Antibodies generated by mice have a fully human VH, a fully human VL, and a mouse constant region. In some embodiments, the human VH and human VL bind to a human IgG constant region (e.g., IgG1, IgG2, IgG3, and IgG4).

[0174] Identity or homology to the original sequence is typically the percentage of amino acid residues present in the candidate sequence that are identical to sequences present in a human, humanized, or chimeric anti-CCR8 antibody or fragment, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, without considering conservative substitutions as part of the sequence identity.

[0175] Further modifications can be made to the anti-CCR8 antibody or antigen-binding fragment. For example, cysteine ​​residues can be introduced into the Fc region, thereby allowing interchain disulfide bond formation within this region. The homodimeric antibody thus generated may have some increased in vitro and / or in vivo half-life. Homodimeric antibodies with increased in vitro and / or in vivo half-lives can also be prepared using heterobifunctional cross-linkers, for example, as described by Wolff et al. ("Monoclonal antibody homodimers: enhanced antitumor activity in nude mice," Cancer research 53.11 (1993):2560-2565). Alternatively, antibodies having dual Fc regions can be engineered.

[0176] In some embodiments, covalent modifications can be made to anti-CCR8 antibodies or antigen-binding fragments thereof. These covalent modifications can be made by chemical or enzymatic synthesis, or by enzymatic or chemical cleavage. Other types of covalent modifications of antibodies or antibody fragments are introduced into the molecule by reacting targeted amino acid residues of the antibody or fragment with organic derivatizing agents capable of reacting with selected side chains or the N- or C-terminal residues.

[0177] In some embodiments, antibody variants are provided that have carbohydrate structures lacking fucose attached (directly or indirectly) to the Fc region. For example, the amount of fucose in such antibody compositions may be 1% to 80%, 1% to 65%, 5% to 65%, or 20% to 40%. The amount of fucose is determined by calculating the average amount of fucose in the glycan at Asn297 relative to the sum of all glycans (e.g., complex, hybrid, and high-mannose structures) attached to Asn297, as measured, for example, by MALDI-TOF mass spectrometry as described in WO 2008 / 077546. Asn297 refers to the asparagine residue located at position 297 (position 314 in the EU numbering of Fc region residues or Kabat numbering) within the Fc region; however, Asn297 may also be located approximately ±3 amino acids upstream or downstream from position 297, i.e., between positions 294 and 300, due to minor sequence variations in antibodies. Such fucosylation variants may have improved ADCC function. In some embodiments, the Fc region of the antibody can be further modified to replace the asparagine at position 297 with alanine (N297A) to reduce glycan heterogeneity.

[0178] In some embodiments, to promote production efficiency by avoiding Fab-arm exchange, the Fc region of the antibody is further modified to replace the serine at position 228 (EU numbering) of IgG4 with a proline (S228P). A detailed description of the S228 mutation is provided, for example, in Silva et al. "The S228P mutation prevents in vivo and in vitro IgG4 Fab-arm exchange as demonstrated using a combination of novel quantitative immunoassays and physiological matrix preparation," Journal of Biological Chemistry 290.9 (2015):5462-5469, the entire contents of which are incorporated herein by reference.

[0179] Recombinant vector The present disclosure also provides recombinant vectors (e.g., expression vectors) comprising the isolated polynucleotides disclosed herein (e.g., polynucleotides encoding the polypeptides disclosed herein), host cells into which the recombinant vectors have been introduced (i.e., such that the host cell contains the polynucleotides and / or vectors comprising the polynucleotides), and the production of recombinant antibody polypeptides or fragments thereof by recombinant techniques.

[0180] As used herein, a "vector" is any construct capable of delivering one or more polynucleotides of interest to a host cell when the vector is introduced into the host cell. An "expression vector" is capable of delivering and expressing one or more polynucleotides of interest as encoded polypeptides in a host cell into which the expression vector is introduced. Thus, within an expression vector, a polynucleotide of interest is positioned for expression in the vector by being operably linked to control elements, such as a promoter, enhancer, and / or polyA tail, within the vector or in the genome of a host cell at, near, or on either side of the integration site of the polynucleotide of interest, such that the polynucleotide of interest is translated in a host cell into which the expression vector is introduced.

[0181] Vectors can be introduced into host cells by methods well known in the art, such as electroporation, chemical transfection (e.g., DEAE-dextran), transformation, transfection, and infection and / or transduction (e.g., by recombinant viruses). Thus, non-limiting examples of vectors include viral vectors (which can be used to generate recombinant viruses), naked DNA or RNA, plasmids, cosmids, phage vectors, and DNA or RNA expression vectors associated with cationic condensing agents.

[0182] In some embodiments, a polynucleotide disclosed herein (e.g., a polynucleotide encoding a polypeptide disclosed herein) is introduced using a viral expression system (e.g., vaccinia or other poxvirus, retrovirus, or adenovirus), which may involve the use of a non-pathogenic (defective), replication-competent virus, or may employ a replication-defective virus, in which case viral propagation generally occurs only in complementary viral packaging cells. Preferred systems are described, for example, in Fisher-Hoch et al., 1989, Proc. Natl. Acad. Sci. USA 86:317-321; Flexner et al., 1989, Ann. N.Y. Acad. Sci. 569:86-103; Flexner et al., 1990, Vaccine, 8:17-21; U.S. Pat. Nos. 4,603,112, 4,769,330, and 5,017,487; WO 89 / 01973; U.S. Pat. No. 4,777,127; GB 2,200,651; EP 0,345,242; WO 91 / 02805; Berkner-Biotechniques, 6:616-627, 1988; Rosenfeld et al. al., 1991, Science, 252:431-434; Kolls et al., 1994, Proc. Natl. Acad. Sci. USA, 91:215-219; Kass-Eisler et al., 1993, Proc. Natl. Acad. Sci. USA, 90:11498-11502; Guzman et al., 1993, Circulation, 88:2838-2848; and Guzman et al., 1993, Cir. Res., 73:1202-1207. Techniques for incorporating DNA into such expression systems are well known to those skilled in the art. DNA can also be "naked," as described, for example, in Ulmer et al., 1993, Science, 259:1745-1749, and Cohen, 1993, Science, 259:1691-1692. Uptake of naked DNA can be enhanced by coating the DNA onto biodegradable beads that are efficiently transported into cells.

[0183] For expression, a DNA insert containing an antibody-encoding or polypeptide-encoding polynucleotide disclosed herein can be operably linked to a suitable promoter (e.g., a heterologous promoter), such as the phage lambda PL promoter, the E. coli lac, trp, and tac promoters, the SV40 early and late promoters, and promoters of retroviral long terminal repeats, to name a few. Other suitable promoters are known to those of skill in the art. In some embodiments, the promoter is a cytomegalovirus (CMV) promoter. The expression construct can further contain sites for transcription initiation and termination, and, within the transcribed region, a ribosome binding site for translation. The coding portion of the mature transcripts expressed by the construct can include a translation initiation codon at the beginning and a termination codon (UAA, UGA, or UAG) appropriately positioned at the end of the polypeptide to be translated.

[0184] As indicated, expression vectors can contain at least one selectable marker. Such markers include dihydrofolate reductase or neomycin resistance genes for eukaryotic cell culture, and tetracycline or ampicillin resistance genes for E. coli and other bacterial culture. Representative examples of suitable hosts include, but are not limited to, bacterial cells such as E. coli, Streptomyces, and Salmonella typhimurium cells, fungal cells such as yeast cells, insect cells such as Drosophila melanogaster S2 and Spodoptera litura Sf9 cells, animal cells such as CHO, COS, Bowes malignant melanoma, and HK 293 cells, and plant cells. Appropriate culture media and conditions for the host cells described herein are well known in the art.

[0185] Non-limiting vectors for use in bacteria include pQE70, pQE60, and pQE-9 available from Qiagen, pBS vectors, Phagescript vectors, Bluescript vectors, pNH8A, pNH16a, pNH18A, pNH46A available from Stratagene, and ptrc99a, pKK223-3, pKK233-3, pDR540, and pRIT5 available from Pharmacia. Non-limiting eukaryotic vectors include pWLNEO, pSV2CAT, pOG44, pXT1, and pSG available from Stratagene, and pSVK3, pBPV, pMSG, and pSVL available from Pharmacia. Other suitable vectors will be readily apparent to those of skill in the art.

[0186] Non-limiting bacterial promoters suitable for use include the E. coli lacI and lacZ promoters, the T3 and T7 promoters, the gpt promoter, the lambda PR and PL promoters, and the trp promoter. Suitable eukaryotic promoters include the CMV immediate early promoter, the HSV thymidine kinase promoter, the early and late SV40 promoters, promoters of retroviral LTRs such as those of Rous sarcoma virus (RSV), and metallothionein promoters such as the mouse metallothionein-I promoter.

[0187] In the yeast Saccharomyces cerevisiae, several vectors containing constitutive or inducible promoters, such as alpha factor, alcohol oxidase, and PGH, can be used.

[0188] Introduction of the construct into the host cell can be accomplished by calcium phosphate transfection, DEAE-dextran-mediated transfection, cationic lipid-mediated transfection, electroporation, transduction, infection, or other methods. Such methods are described in many standard laboratory manuals, such as Davis et al., Basic Methods In Molecular Biology (1986), the entire contents of which are incorporated herein by reference.

[0189] Transcription of DNA encoding the antibodies of the present disclosure by higher eukaryotes can be increased by inserting an enhancer sequence into the vector. Enhancers are cis-acting elements of DNA, usually about 10 to 300 bp, that serve to increase transcriptional activity of a promoter in a given host cell type. Examples of enhancers include the SV40 enhancer, which is located on the late side of the replication origin at base pairs 100 to 270, the cytomegalovirus early promoter enhancer, the polyoma enhancer on the late side of the replication origin, and adenovirus enhancers.

[0190] For secretion of the translated protein into the lumen of the endoplasmic reticulum, the periplasmic space or the extracellular environment, appropriate secretion signals can be incorporated into the expressed polypeptide. The signals can be endogenous to the polypeptide or they can be heterologous signals.

[0191] Polypeptides (e.g., antibodies) can be expressed in modified forms, such as fusion proteins (e.g., GST fusions) or histidine-tagged, and can contain not only secretion signals but also additional heterologous functional regions. For example, a region of additional amino acids, particularly charged amino acids, can be added to the N-terminus of a polypeptide to improve stability and durability in host cells during purification or during subsequent handling and storage. Peptide moieties can also be added to polypeptides to facilitate purification. Such regions can be removed before final preparation of the polypeptide. Adding peptide moieties to polypeptides to cause secretion or excretion, improve stability, and facilitate purification is, inter alia, a well-known and routine technique in the art.

[0192] Treatment method The antibodies or antigen-binding fragments thereof of the present disclosure can be used for a variety of therapeutic purposes. In one aspect, the present disclosure provides methods of treating cancer in a subject, reducing the rate of growth of tumor volume in a subject over time, reducing the risk of developing metastases, or reducing the risk of developing further metastases in a subject. In some embodiments, treatment can halt, slow, prevent, or inhibit the progression of cancer. In some embodiments, treatment can result in a reduction in the number, severity, and / or duration of one or more symptoms of cancer in a subject.

[0193] In one aspect, the disclosure features a method including administering a therapeutically effective amount of an antibody or antigen-binding fragment thereof disclosed herein to a subject in need thereof (e.g., a subject suffering from, or identified or diagnosed as suffering from, cancer), e.g., breast cancer (e.g., triple-negative breast cancer), carcinoid cancer, cervical cancer, endometrial cancer, glioma, head and neck cancer, liver cancer, lung cancer, small cell lung cancer, lymphoma, malignant melanoma, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, colorectal cancer, gastric cancer, testicular cancer, thyroid cancer, bladder cancer, urethral cancer, stomach cancer, testicular cancer, gallbladder cancer, bile duct cancer, esophageal cancer, thymic cancer, acute myeloid leukemia, lymphoblastic lymphoma, transitional cell carcinoma, or a hematological malignancy. In some embodiments, the cancer is unresectable or metastatic melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), bladder cancer, or metastatic hormone-refractory prostate cancer. In some embodiments, the subject is a patient with a solid tumor. In some embodiments, the cancer is squamous cell carcinoma of the head and neck (SCCHN), renal cell carcinoma (RCC), triple-negative breast cancer (TNBC), or colorectal carcinoma. In some embodiments, the subject is a patient with Hodgkin's lymphoma. In some embodiments, the subject is a patient with triple-negative breast cancer (TNBC), gastric cancer, urothelial carcinoma, Merkel cell carcinoma, or head and neck cancer. In some embodiments, the cancer is melanoma, pancreatic cancer, mesothelioma, a hematological malignancy, particularly non-Hodgkin's lymphoma, lymphoma, chronic lymphocytic leukemia, or an advanced solid tumor.

[0194] In some embodiments, the subject has a PD-1 inhibitor-resistant cancer. In some embodiments, the subject is unresponsive to an anti-PD-1 antibody or anti-PD-1 antibody treatment.

[0195] In some embodiments, the compositions and methods disclosed herein can be used to treat patients at risk for cancer. Patients with cancer can be identified by a variety of methods known in the art.

[0196] In one aspect, the present disclosure provides methods for treating, preventing, or reducing the risk of developing a disease associated with an aberrant or unwanted immune response, such as an autoimmune disease.

[0197] As used herein, "effective amount" means an amount or dosage sufficient to bring about beneficial or desired results, including halting, slowing, preventing, or inhibiting the progression of a disease, e.g., cancer. The effective amount will vary depending on, for example, the age and weight of the subject to whom the antibody, antigen-binding fragment, polynucleotide encoding the antibody, vector comprising the polynucleotide, and / or composition thereof is administered, the severity of symptoms, and the route of administration, and thus, dosing can be determined on an individual basis.

[0198] An effective amount can be administered in one or more administrations. For example, an effective amount of an antibody or antigen-binding fragment is an amount sufficient to palliate, arrest, stabilize, reverse, inhibit, slow, and / or delay the progression of an autoimmune disease or cancer in a patient, or to palliate, arrest, stabilize, reverse, slow, and / or delay the proliferation of cells (e.g., biopsy cells, any of the cancer cells described herein, or cell lines (e.g., cancer cell lines)) in vitro. As understood in the art, an effective amount of an antibody or antigen-binding fragment may vary depending on other factors, such as, inter alia, the patient's medical history, as well as the type (and / or dosage) of antibody used.

[0199] Effective amounts and schedules for administering the antibodies, antibody-encoding polynucleotides, and / or compositions disclosed herein can be determined empirically, and making such determinations is within the skill of one in the art. One of skill in the art will understand that the dosage required to be administered will vary depending, for example, on the mammal receiving the antibodies, antibody-encoding polynucleotides, and / or compositions disclosed herein, the route of administration, the particular type of antibody, antibody-encoding polynucleotide, antigen-binding fragment, and / or composition disclosed herein used, and other agents administered to the mammal.

[0200] A typical daily dose of an effective amount of antibody is 0.01 mg / kg to 100 mg / kg (mg per kg of patient body weight). In some embodiments, the dose can be less than 100 mg / kg, 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.5 mg / kg, or 0.1 mg / kg. In some embodiments, the dose can be greater than 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.5 mg / kg, 0.1 mg / kg, 0.05 mg / kg, or 0.01 mg / kg. In some embodiments, the dose is about 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.9 mg / kg, 0.8 mg / kg, 0.7 mg / kg, 0.6 mg / kg, 0.5 mg / kg, 0.4 mg / kg, 0.3 mg / kg, 0.2 mg / kg, or 0.1 mg / kg.

[0201] In any of the methods described herein, at least one antibody, antigen-binding fragment thereof, or pharmaceutical composition (e.g., any of the antibodies, antigen-binding fragments, or pharmaceutical compositions described herein), and optionally at least one additional therapeutic agent, can be administered to a subject at least once a week (e.g., once a week, twice a week, three times a week, four times a week, once a day, twice a day, or three times a day). In some embodiments, at least two different antibodies and / or antigen-binding fragments are administered in the same composition (e.g., a liquid composition). In some embodiments, at least one antibody or antigen-binding fragment and at least one additional therapeutic agent are administered in the same composition (e.g., a liquid composition). In some embodiments, at least one antibody or antigen-binding fragment and at least one additional therapeutic agent are administered in two different compositions (e.g., a liquid composition containing at least one antibody or antigen-binding fragment and a solid oral composition containing at least one additional therapeutic agent). In some embodiments, at least one additional therapeutic agent is administered as a pill, tablet, or capsule. In some embodiments, the at least one additional therapeutic agent is administered in a sustained release oral formulation.

[0202] In some embodiments, one or more additional therapeutic agents can be administered to a subject before or after administration of at least one antibody, antigen-binding antibody fragment, or pharmaceutical composition (e.g., any of the antibodies, antigen-binding antibody fragments, or pharmaceutical compositions described herein). In some embodiments, the one or more additional therapeutic agents and at least one antibody, antigen-binding antibody fragment, or pharmaceutical composition (e.g., any of the antibodies, antigen-binding antibody fragments, or pharmaceutical compositions described herein) are administered to a subject such that there is overlap in the period of biological activity of the one or more additional therapeutic agents with the period of biological activity of the at least one antibody or antigen-binding fragment (e.g., any of the antibodies or antigen-binding fragments described herein) in the subject.

[0203] In some embodiments, a subject can be administered at least one antibody, antigen-binding antibody fragment, or pharmaceutical composition (e.g., any of the antibodies, antigen-binding antibody fragments, or pharmaceutical compositions described herein) for an extended period of time (e.g., for a period of at least 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, 1 year, 2 years, 3 years, 4 years, or 5 years). A skilled medical professional can determine the length of treatment period using any of the methods described herein to diagnose or track the effectiveness of the treatment (e.g., to observe at least one symptom of cancer). As described herein, a skilled medical professional can also vary (e.g., increase or decrease) the identity and number of antibodies or antigen-binding antibody fragments (and / or one or more additional therapeutic agents) administered to a subject, and can adjust (e.g., increase or decrease) the dosage or frequency of administration of at least one antibody or antigen-binding antibody fragment (and / or one or more additional therapeutic agents) to a subject based on an evaluation of the effectiveness of the treatment (e.g., using any of the methods described herein and known in the art).

[0204] In some embodiments, one or more additional therapeutic agents can be administered to the subject. The additional therapeutic agents can include one or more inhibitors selected from the group consisting of B-Raf inhibitors, EGFR inhibitors, MEK inhibitors, ERK inhibitors, K-Ras inhibitors, c-Met inhibitors, anaplastic lymphoma kinase (ALK) inhibitors, phosphatidylinositol 3-kinase (PI3K) inhibitors, Akt inhibitors, mTOR inhibitors, dual PI3K / mTOR inhibitors, Bruton's tyrosine kinase (BTK) inhibitors, and isocitrate dehydrogenase 1 (IDH1) and / or isocitrate dehydrogenase 2 (IDH2) inhibitors. In some embodiments, the additional therapeutic agent is an indoleamine 2,3-dioxygenase-1 (IDO1) inhibitor (e.g., epacadostat).

[0205] In some embodiments, the additional therapeutic agent can comprise one or more inhibitors selected from the group consisting of inhibitors of HER3, inhibitors of LSD1, inhibitors of MDM2, inhibitors of BCL2, inhibitors of CHK1, inhibitors of the activated hedgehog signaling pathway, and agents that selectively degrade the estrogen receptor.

[0206] In some embodiments, the additional therapeutic agent is trabectedin, nab-paclitaxel, trebananib, pazopanib, cediranib, palbociclib, everolimus, fluoropyrimidine, IFL, regorafenib, leolysin, Alimta, Dicaida, Sutent, temsirolimus, axitinib, everolimus, sorafenib, Votrient, pazopanib, IMA-901, AGS-00 The present invention may include one or more therapeutic agents selected from the group consisting of 3, cabozantinib, vinflunine, Hsp90 inhibitors, Ad-GM-CSF, temozolomide, IL-2, IFNa, vinblastine, thalomid, dacarbazine, cyclophosphamide, lenalidomide, azacitidine, lenalidomide, bortezomib, amrubicin, carfilzomib, pralatrexate, and enzastaurin.

[0207] In some embodiments, the additional therapeutic agent can comprise one or more therapeutic agents selected from the group consisting of an adjuvant, a TLR agonist, tumor necrosis factor (TNF) alpha, IL-1, HMGB1, an IL-10 antagonist, an IL-4 antagonist, an IL-13 antagonist, an IL-17 antagonist, an HVEM antagonist, an ICOS agonist, a CX3CL1 targeted therapy, a CXCL9 targeted therapy, a CXCL10 targeted therapy, a CCL5 targeted therapy, an LFA-1 agonist, an ICAM1 agonist, and a selectin agonist.

[0208] In some embodiments, carboplatin, nab-paclitaxel, paclitaxel, cisplatin, pemetrexed, gemcitabine, FOLFOX, or FOLFIRI is administered to the subject.

[0209] In some embodiments, the additional therapeutic agent is an anti-OX40 antibody, an anti-PD-1 antibody, an anti-PD-L1 antibody, an anti-PD-L2 antibody, an anti-LAG-3 antibody, an anti-TIGIT antibody, an anti-BTLA antibody, an anti-CTLA-4 antibody, an anti-B7-H3 antibody, an anti-CLDN18 antibody, an anti-SIGLEC15 antibody, an anti-41BB antibody, an anti-CD40 antibody, or an anti-GITR antibody.

[0210] In some embodiments, the additional therapeutic agent is a PD-1 pathway inhibitor, such as an anti-PD-1 antibody or an anti-PD-L1 antibody. Exemplary anti-PD-1 antibodies include, for example, pembrolizumab, nivolumab, cemiplimab, spartalizumab, camrelizumab, sintilimab, tislelizumab, toripalimab, and dostarlimab. Exemplary anti-PD-L1 antibodies include, for example, atezolizumab, avelumab, durvalumab, and KN035.

[0211] Pharmaceutical Compositions and Routes of Administration Also provided herein are pharmaceutical compositions containing at least one (e.g., 1, 2, 3, or 4) of the antibodies or antigen-binding fragments described herein. Two or more (e.g., 2, 3, or 4) of any of the antibodies or antigen-binding fragments described herein can be present in the pharmaceutical composition, in any combination. Pharmaceutical compositions can be formulated in any manner known in the art.

[0212] Pharmaceutical compositions are formulated to be compatible with their intended route of administration (e.g., intravenous, intraarterial, intramuscular, intradermal, subcutaneous, or intraperitoneal). The compositions may contain a sterile diluent (e.g., sterile water or saline), fixed oils, polyethylene glycol, glycerin, propylene glycol, or other synthetic solvents, antibacterial or antifungal agents (e.g., benzyl alcohol, methylparaben, chlorobutanol, phenol, ascorbic acid, thimerosal, etc.), antioxidants (e.g., ascorbic acid or sodium bisulfite), chelating agents (e.g., ethylenediaminetetraacetic acid), buffers (e.g., acetate, citrate, or phosphate), and isotonic agents (e.g., sugars (e.g., dextrose), polyalcohols (e.g., mannitol or sorbitol), or salts (e.g., sodium chloride)), or any combination thereof. Liposomal suspensions may also be used as pharmaceutically acceptable carriers. The composition preparation can be formulated and enclosed in ampoules, disposable syringes, or multiple-dose vials. Where necessary (e.g., in injectable formulations), proper fluidity can be maintained, for example, by the use of a coating such as lecithin or a surfactant. Absorption of the antibody or antigen-binding fragment thereof can be prolonged by including an agent that delays absorption (e.g., aluminum monostearate and gelatin). Alternatively, sustained release can be achieved by implants and microencapsulated delivery systems, which can include biodegradable, biocompatible polymers (e.g., ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid).

[0213] Compositions containing one or more of any of the antibodies or antigen-binding fragments described herein can be formulated for parenteral (e.g., intravenous, intraarterial, intramuscular, intradermal, subcutaneous, or intraperitoneal) administration in unit dosage form (i.e., physically discrete units containing a predetermined amount of active compound(s) for ease of administration and uniformity of dosage).

[0214] Pharmaceutical compositions for parenteral administration are preferably sterile, substantially isotonic, and manufactured under Good Manufacturing Practice (GMP) conditions. Pharmaceutical compositions may be provided in unit dosage form (i.e., a dose for a single administration). Pharmaceutical compositions may be formulated using one or more physiologically acceptable carriers, diluents, excipients, or adjuvants. The formulation depends on the chosen route of administration. For injection, the antibody may be formulated in an aqueous solution, preferably in a physiologically compatible buffer, to reduce discomfort at the injection site. The solution may contain formulating agents such as suspending agents, stabilizing agents, and / or dispersing agents. Alternatively, the antibody may be in lyophilized form for constitution with a suitable vehicle, e.g., sterile, pyrogen-free water, before use.

[0215] The toxicity and therapeutic efficacy of a composition can be determined by standard pharmaceutical procedures in cell cultures or experimental animals (e.g., monkeys). For example, the LD50 (the dose lethal to 50% of the population) and the ED50 (the dose therapeutically effective in 50% of the population) can be determined, and the therapeutic index is the ratio of LD50:ED50. Drugs that exhibit a high therapeutic index are preferred. If a drug exhibits undesirable side effects, care should be taken to minimize the potential for harm (i.e., reduce the undesirable side effects). Toxicity and therapeutic efficacy can be determined by other standard pharmaceutical procedures.

[0216] Data obtained from cell culture assays and animal studies can be used in formulating an appropriate dose of any given agent for use in a subject (e.g., a human). A therapeutically effective amount of one or more (e.g., one, two, three, or four) antibodies or antigen-binding fragments thereof (e.g., any of the antibodies or antibody fragments described herein) is an amount that treats the disease in a subject (e.g., a human subject identified as having cancer) or a subject identified as at risk for developing the disease (e.g., a subject who previously developed cancer but has now been cured), (e.g., an amount that reduces the severity, frequency, and / or duration of one or more symptoms of the disease in the subject (e.g., a human). The efficacy and administration of any of the antibodies or antigen-binding fragments described herein can be determined by a health care professional or veterinary professional using methods well known in the art, as well as by observing one or more symptoms of the disease in the subject (e.g., a human). Certain factors can affect the dosage and timing required to effectively treat a subject (e.g., the severity of the disease or disorder, previous treatments, the overall health and / or age of the subject, and the presence of other diseases).

[0217] Exemplary doses include amounts (milligrams or micrograms) of any of the antibodies or antigen-binding fragments described herein per kilogram of subject body weight (e.g., about 1 μg / kg to about 500 mg / kg, about 100 μg / kg to about 500 mg / kg, about 100 μg / kg to about 50 mg / kg, about 10 μg / kg to about 5 mg / kg, about 10 μg / kg to about 0.5 mg / kg, about 1 μg / kg to about 50 μg / kg, about 1 mg / kg to about 10 mg / kg, or about 1 mg / kg to about 5 mg / kg). While these doses cover a wide range, those of skill in the art will understand that therapeutic agents, including antibodies and antigen-binding fragments thereof, vary in their potency and that effective amounts can be determined by methods well known in the art. Typically, a relatively low dose is administered initially, and the attending medical or veterinary professional (for therapeutic uses) or researcher (for developmental stages) can gradually increase the dose until an appropriate response is obtained. In addition, it is understood that the specific dose level for any particular subject will depend on a variety of factors, including the activity of the specific compound used, the subject's age, weight, general health, sex, and diet, the time of administration, the route of administration, the rate of excretion, and the half-life of the antibody or antibody fragment in vivo.

[0218] The pharmaceutical compositions can be included in a container, pack, or dispenser together with instructions for administration.The present disclosure also provides methods for producing antibodies or antigen-binding fragments thereof for various uses described herein. [Example]

[0219] The invention is further described in the following examples, which do not limit the scope of the invention described in the claims.

[0220] Example 1. Generation of anti-CCR8 antibodies RenMice to generate antibodies against human CCR8 TM (i.e., recombinant mice containing DNA encoding human immunoglobulin heavy chain and kappa light chain variable regions, e.g., RenMab TM Mouse, RenLiteTM Mice were immunized with human CCR8 protein, and antibody responses were monitored by antigen-specific immunoassays.

[0221] A total of four immunizations were administered, with immunizations given at 2-week intervals. Five to six days after the final immunization, retroorbital bleeds were collected, and serum antibody titers were determined by FACS.

[0222] Spleen tissue was then harvested and pulverized. Spleen cells were first selected with CD3ε microbeads and anti-mouse IgM microbeads (to generate hybridoma cells) and then fused with SP2 / 0 cells. The cells were then plated in 96-well plates with hypoxanthine-aminopterin-thymidine (HAT) medium.

[0223] Primary screening of hybridoma supernatants in 96-well plates was performed using FACS according to standard procedures. Subcloning was performed using ClonePix® 2. Positive wells identified during primary screening were transferred to semi-solid medium, and IgG-positive clones were identified and tested. FITC anti-mouse IgG Fc antibody was used. Positive hybridoma cells were then injected intraperitoneally into mice. Monoclonal antibodies were produced by growing the hybridoma cells in the mouse peritoneal cavity. The hybridoma cells proliferated and produced ascites in the abdominal cavity of the mouse. This fluid contained a high concentration of antibodies and could be harvested for later use. The antibodies in the ascites were collected using GE AKTA TM Purification was performed using protein chromatography (GE Healthcare, Chicago, IL, USA).

[0224] In another experiment, when a desired immune response was achieved, antigen-specific immune cells were isolated from the immunized mice to obtain further anti-CCR8 antibodies or to obtain the light and heavy chain variable region sequences of anti-CCR8 antibodies. For example, single-cell technology (e.g., using the Beacon® Optofluidic System, Berkeley Lights Inc.) was used to screen and identify plasma cells secreting antigen-specific monoclonal antibodies, and antibody variable region sequences were obtained using reverse transcription and PCR sequencing. The obtained variable region sequences were used for antibody expression. Binding affinity to CCR8 was determined using FACS.

[0225] In another experiment, phage display was performed to screen and find antigen-specific monoclonal antibodies.

[0226] Exemplary antibodies obtained by this method include 11B12, 11F8, and 11F5.

[0227] In another experiment, the "DG" in the VH CDR2 of the antibodies described herein was replaced with "SG," "AG," or "DA" to create CDR-modified VHs. For example, in one embodiment, the "DG" in the VH CDR2 of 11B12 was replaced with "SG," and the resulting CDR-modified VH is designated 11B12-SG VH. The sequences of some of these CDR-modified VHs are shown in Figure 4.

[0228] In another experiment, "DG" in the VL CDR1 of the antibodies described herein was replaced with "SG," "AG," or "DA" to create CDR-modified VLs. For example, in one embodiment, "DG" in the VL CDR1 of 11B12 was replaced with SG, and the resulting CDR-modified VL is designated 11B12-SG VL. The sequences of some of these CDR-modified VLs are shown in Figure 4.

[0229] In another experiment, "DNS" in the VH framework regions of the antibodies described herein was replaced with "DNA" to create FR-modified VHs. For example, "DNS" in the framework of the VH of 11B12 was replaced with "DNA", and the resulting FR-modified VH is designated 11B12-DNA VH. The sequences of some of these FR-modified VHs are shown in Figure 4 (e.g., SEQ ID NOS: 69-71). Thus, the antibodies have a S74A mutation (Kabat numbering).

[0230] In another experiment, VH was modified by both FR (e.g., DNA) and CDR (e.g., SG) modifications. In one example, "DNS" in the framework of 11B12 VH was replaced with "DNA," and "DG" in 11B12 VH CDR2 was replaced with "SG," and the resulting FR- and CDR-modified VH is designated 11B12-SG-DNA VH. The sequence of the resulting FR- and CDR-modified VH is shown in Figure 4 (e.g., SEQ ID NO: 216).

[0231] In another experiment, to generate 11F5 CDR3-modified VH, amino acid "N" at position 3 in VH CDR3 of 11F5 was substituted with "S", amino acid "Y" at position 5 in VH CDR3 of 11F5 was substituted with "V", and / or amino acid "Y" at position 6 in VH CDR3 of 11F5 was substituted with "N". For example, amino acid "N" at position 3 in VH CDR3 of 11F5 was substituted with "S", and the resulting CDR-modified VH is designated 11F5-N3S VH. Amino acid "Y" at position 5 in VH CDR3 of 11F5 is substituted with "V", and the resulting CDR-modified VH is designated 11F5-Y5V VH. The amino acid "Y" at position 6 in the VH CDR3 of 11F5 is replaced with "N", and the resulting CDR-modified VH is designated 11F5-Y6N VH. The amino acid "N" at position 3 and the amino acid "Y" at position 5 in the VH CDR3 of 11F5 are replaced with "S" and "V", respectively, and the resulting CDR-modified VH is designated 11F5-N3S-Y5V VH. The amino acid "N" at position 3 and the amino acid "Y" at position 6 in the VH CDR3 of 11F5 are replaced with "S" and "N", respectively, and the resulting CDR-modified VH is designated 11F5-N3S-Y6N VH. In the VH CDR3 of 11F5, the amino acid "Y" at position 5 and the amino acid "Y" at position 6 were replaced with "V" and "N", respectively, and the resulting CDR-modified VH was designated 11F5-Y5V-Y6N VH. In the VH CDR3 of 11F5, the amino acid "N" at position 3, the amino acid "Y" at position 5, and the amino acid "Y" at position 6 were replaced with "S," "V," and "N," respectively, and the resulting CDR-modified VH was designated 11F5-N3S-Y5V-Y6N VH. The sequences of these CDR-modified VHs are shown in Figure 4.

[0232] In another experiment, the VH of an antibody described herein was paired with the VL of another antibody described herein to produce an antibody.For example, in one embodiment, the VH of 11B12 was paired with the VL of 11F8, and the resulting antibody was named 11B12-11F8.In one embodiment, the VH of 11B12-SG was paired with the VL of 11F8-SG, and the resulting antibody was named 11B12-SG-11F8-SG.In one embodiment, the VH of 11B12-SG was paired with the VL of 11B12-SG, and the resulting antibody was named 11B12-SG-11B12-SG.In one embodiment, the VH of 11B12-SG-DNA was paired with the VL of 11F8, and the resulting antibody was named 11B12-SG-DNA-11F8. Some of the disclosed VH and VL combinations are listed in Figure 5. The sequences of the heavy and light chain variable regions of these antibodies are shown in Figure 4. Figures 2A and 3A show the heavy and light chain CDR sequences of 11B12, 11F8, and 11F5, respectively, according to the Kabat and Chothia definitions.

[0233] Various IgG1, IgG2, and IgG4 antibodies were generated. When the VH and VL of 11B12 are connected to an IgG1 constant region, the antibody is designated 11B12. When the VH and VL of 11B12 are connected to an IgG4 constant region, the antibody is designated 11B12-IgG4. The constant region can also contain some mutations. For example, when SI mutations (EU numbering: S239D and I332E mutations) are introduced into the Fc region of 11B12, the resulting antibody is designated 11B12-SI.

[0234] Example 2. Cross-reactivity of anti-CCR8 antibodies CHO-S cells expressing human CCR8 (CHO-S-hCCR8), CHO-S cells expressing mouse CCR8 (CHO-S-mCCR8), HEK293T cells expressing human CCR8 (293T-hCCR8), and HEK293T cells expressing cynomolgus monkey CCR8 (293T-cynCCR8) were cultured in a 96-well plate at 5 × 10 4The cells were plated at a density of 1000 cells / well. Diluted sample anti-CCR8 antibodies (1 μg / mL) were added to the 96-well plate and incubated at 4°C for 30 minutes. The cells were then incubated with secondary antibody anti-hIgG-Fc-Alexa Flour® 647 (RL1-H) (Jackson ImmunoResearch Laboratories, Inc., Cat. No. 109-606-170) in the dark for 15 minutes at 4°C before flow cytometry analysis.

[0235] CHO-S-hCCR8 cells were obtained by transfecting CHO-S cells with a vector encoding human CCR8 (hCCR8, SEQ ID NO: 43). CHO-S-mCCR8 cells were obtained by transfecting CHO-S cells with a vector encoding mouse CCR8 (mCCR8, SEQ ID NO: 44). 293T-hCCR8 cells were obtained by transfecting HEK293T cells (ATCC, Cat. No. CRL-3216) with a vector encoding human CCR8. TM 293T-cynCCR8 cells were obtained by transfecting HEK293T cells with a vector encoding a construct containing the amino acid sequence of cynomolgus monkey (monkey) CCR8 (cynCCR8, SEQ ID NO: 45).

[0236] The test results are shown in the following tables (Table 1 and Table 2): All three anti-CCR8 antibodies can bind to hCCR8, cynCCR8, and mCCR8.

[0237] [Table 1]

[0238] [Table 2]

[0239] Example 3. Cross-reactivity of anti-CCR antibodies to CCR8 family proteins In each experiment, CHO cells were transfected to express human CCR8 (CHO-S-hCCR8), human CCR2 (hCCR2, SEQ ID NO: 46) (CHO-S-hCCR2), human CCR4 (hCCR4, SEQ ID NO: 47) (CHO-S-hCCR4), human CCR5 (hCCR5, SEQ ID NO: 48) (CHO-S-hCCR5), or human CX3CR1 (hCX3CR1, SEQ ID NO: 49) (CHO-S-hCX3CR1).

[0240] CHO-S-hCCR8, CHO-S-hCCR2, CHO-S-hCCR4, CHO-S-hCCR5, and CHO-S-hCX3CR1 were cultured in a 96-well plate at 5 × 10 4 The cells were plated at a density of 1000 cells / well. Diluted sample anti-CCR8 antibodies (1 μg / mL) were added to the 96-well plate and incubated at 4°C for 30 minutes. The cells were then incubated with the secondary antibody Anti-hIgG-Fc-Alexa Flour® 647 (RL1-H) (Jackson ImmunoResearch Laboratories, Inc., Catalog No. 109-606-170) in the dark for 15 minutes at 4°C before flow cytometry analysis.

[0241] The test results are shown in the table below: None of the three anti-CCR8 antibodies can bind to hCCR2, hCCR4, hCCR5, or hCX3CR1.

[0242] [Table 3]

[0243] Prosalizuzumab (MLN1202) is a humanized immunoglobulin G1 (IgG1) anti-CCR2 monoclonal antibody developed by Millennium Pharmaceuticals (Takeda Pharmaceuticals). The VH and VL sequences of the prosalizuzumab analog are shown in SEQ ID NO: 50 and SEQ ID NO: 51, respectively.

[0244] Mogamulizumab (KW-0761) is a humanized immunoglobulin G1 (IgG1) anti-CCR4 monoclonal antibody developed by Kyowa Kirin Co., Ltd. The VH and VL sequences of mogamulizumab analog-SI are shown in SEQ ID NO: 52 and SEQ ID NO: 53, respectively.

[0245] Leronlimab is a fully humanized IgG4 monoclonal antibody targeting CCR5 developed by CytoDyn, Inc. The VH and VL sequences of the leronlimab analog are shown in SEQ ID NO: 54 and SEQ ID NO: 55, respectively.

[0246] BII-00313 is a single heavy chain variable domain (VHH) antibody developed by Boehringer-Ingelheim that targets the human CX3C chemokine receptor 1 (CX3CR1) mutant (VH SEQ ID NO: 56). BII-00313-FC-LALA was constructed by linking the BII-00313 variable domain to an IgG1 Fc with LALA mutations (EU numbering: L234A and L235A mutations).

[0247] Example 4. Binding affinity verification of anti-CCR8 antibodies 11F5, 11F8, and 11B12 The binding activity of anti-CCR8 antibodies (11B12, 11F8, 11F5, and 7B16 analogs) to 293T-hCCR8 cells (HEK293T cells expressing human CCR8) or 293T-cynCCR8 (HEK293T cells expressing cynomolgus monkey CCR8) was verified by flow cytometry. The secondary antibody used in the experiment was Alexa Fluor® 647 anti-human IgG Fcγ (Jackson ImmunoResearch Laboratories, Inc., catalog number: 109-606-170). EC50 was determined using serially diluted sample anti-CCR8 antibodies. The results are shown in the table below.

[0248] 7B16 is a humanized immunoglobulin G1 (IgG1) anti-CCR8 monoclonal antibody developed by JOUNCE THERAPEUTICS, Inc. (see WO2021163064A2). The VH and VL sequences of the 7B16 analog are set forth in SEQ ID NO: 59 and SEQ ID NO: 60, respectively.

[0249] [Table 4]

[0250] All three anti-CCR8 antibodies showed binding affinity to both human and monkey CCR8. Among them, 11B12 bound to both monkey and human CCR8 with similar affinity. 11F8 and 11F5 bound to human CCR8 with higher affinity than monkey CCR8. However, the 7B16 analog showed binding affinity to human CCR8 but not to monkey CCR8 (data not shown).

[0251] In a separate experiment, the binding activity of anti-CCR8 antibodies (11B12, 11F8, 11F5, 10A11 analogs, and 7B16 analogs) to 293T-hCCR8 cells (HEK293T cells expressing human CCR8) or 293T-mCCR8 (HEK293T cells expressing mouse CCR8) was verified by flow cytometry. The secondary antibody used in the experiment was PE Goat α-Human IgG Fc (SouthernBiotech, Catalog No.: 2014-09). EC50 values ​​were determined using serially diluted sample anti-CCR8 antibodies. The results are shown in the table below.

[0252] [Table 5]

[0253] 10A11 is a humanized immunoglobulin G1 (IgG1) anti-CCR8 monoclonal antibody developed by Shionogi & Co., Ltd. (see WO2020138489A1). The VH and VL sequences of the 10A11 analog are shown in SEQ ID NO: 57 and SEQ ID NO: 58, respectively.

[0254] The anti-CCR8 antibodies 11B12, 11F8, and 11F5 showed good binding affinity to human and mouse CCR8. The 10A11 and 7B16 analogs showed binding affinity to human CCR8 but not to mouse CCR8 (data not shown).

[0255] Example 5. Blockade of calcium flux in hCCR8-expressing cells by anti-hCCR8 mAbs Blockade of hCCL1 binding to hCCR8 by human IgG1 anti-CCR8 antibodies and CDR- and FR-modified anti-CCR8 antibodies was tested by performing a calcium (Ca) flux assay using human CCR8-expressing 293T cells. A Fluorescence Imaging Plate Reader (FLIPR) Calcium 6 Assay Kit (Molecular Devices, catalog number: R8190) was used. Cells were seeded the day before the experiment and incubated overnight at 37°C and 5% CO2. Next, the cells were loaded with FLIPR Calcium 6 Dye (Molecular Devices) containing probenecid (Invitrogen) and PowerLoad (Invitrogen) and incubated at 37°C for 30 minutes. Different anti-CCR8 mAbs were added to the cells and further incubated at 37°C for 30 minutes. After incubation, the Ca flux fluorescence signal induced by 1 μg / mL recombinant human CCL1 (BioLegend, Cat. No. 582706) was measured for 1 minute using a SpectraMax® iD3 (Molecular Devices). The results are shown in the table below.

[0256] [Table 6]

[0257] 11B12, 11F8, 11B12-11F8, 11F8-SG-11F8-SG, 11F5-SG-11F5-SG, 11B12-SG-11F8, and 11B12-DNA-11B12 showed nearly complete inhibition, whereas the 10A11 analog showed only partial inhibition even at 10 μg / mL.

[0258] Example 6. Biophysical analysis and stability of anti-CCR8 antibodies The biophysical properties and stability of three anti-CCR8 antibodies, 11B12, 11F8, and 11F5, were evaluated.

[0259] Three anti-CCR8 antibodies, 11B12, 11F8, and 11F5, were buffer-exchanged to pH 6.0 (3 mg / mL histidine, 80 mg / mL sucrose, and 0.2 mg / mL Tween 80). The antibodies were placed in sealed Eppendorf tubes and stored at 40±2°C, 60%±5% RH (hereafter referred to as 40°C) and 4±3°C (hereafter referred to as 4°C) for 14 days, and their thermal stability was evaluated.

[0260] Three anti-CCR8 antibodies, 11B12, 11F8, and 11F5, were loaded onto a Protein A column and eluted with pH 3.5 buffer (0.1 mol / L HAc). Half of the antibodies were added to 2 M Tris buffer, and the pH was immediately adjusted to 7.5. The remaining half was kept at pH 3.5 for 6 hours, and then the pH was adjusted to 7.5. The diluted antibodies were placed in sealed Eppendorf tubes and stored at pH 3.5 ± 0.1 and 25 ± 2°C (hereafter referred to as pH 3.5) for 6 hours to test their stability at low pH.

[0261] Three anti-CCR8 antibodies, 11B12, 11F8, and 11F5, were buffer-exchanged to pH 6.0 (3 mg / mL histidine, 80 mg / mL sucrose, 0.2 mg / mL Tween® 80) and NH4HCO3 stock was added to a final concentration of 0.94% NH4HCO3. To test stability, samples were placed in sealed Eppendorf tubes and stored at 40±2°C and 60%±5% RH for 6 hours (hereafter referred to as NH4HCO36h) or 24 hours (hereafter referred to as NH4HCO324h).

[0262] Specifically, the following tests were performed: (1) size-exclusion high-performance liquid chromatography (SEC-HPLC) to detect antibody purity (expressed as the ratio of the main peak area to the sum of all peak areas (purity, %)); (2) hydrophobic interaction chromatography-high-performance liquid chromatography (HIC-HPLC) to detect the apparent hydrophobicity of the antibody (expressed as the retention time of the main peak (HIC, minutes)); (3) capillary isoelectric focusing (cIEF) to detect the pI (isoelectric point) and charge variants of the antibody (expressed as the ratio of the main component, acidic component, and alkaline component); (4) capillary electrophoresis-sodium dodecyl sulfate (CE-SDS) under non-reducing (CE-SDS (NR)) conditions to detect changes in antibody purity (expressed as the ratio of the main peak area to the sum of all peak areas (purity, %)); and (5) high-throughput multifunctional protein stability analyzer (UNcle, Unchained Labs) to detect the thermodynamic stability of the antibody (expressed as the melting temperature (Tm) and aggregation temperature (Tagg)).

[0263] For SEC-HPLC experiments, antibody samples were diluted to 1 mg / mL with purified water and run on an Agilent 1290 chromatographic system connected to an XBridge Protein BEH SEC column (200 Å, Waters Corporation). The following parameters were used: mobile phase: 0.1 M phosphate buffer (PB) + 10% ACN, pH 7.4; flow rate: 1.8 mL / min; column temperature: 25°C; detection wavelengths: 280 nm, 220 nm; injection volume: 10 μL; sample tray temperature: approximately 4°C; and run time: 7 min.

[0264] For HIC-HPLC experiments, an Agilent 1260 chromatographic system (connected to a ProPac HIC-10 column (4.6 × 250 mm, Thermo Scientific)) was used, and the sample was diluted 10-fold using mobile phase A. The following parameters were used: mobile phase A: 0.9 M ammonium sulfate, 0.1 M phosphate buffer (PB), 10% acetonitrile pH 6.5; mobile phase B: 0.1 M phosphate buffer (PB), 10% acetonitrile pH 6.5; flow rate: 0.8 mL / min; gradient: 0 min 100% A, 2 min 100% A, 32 min 100% B, 34 min 100% B, 35 min 100% A, 45 min 100% A; column temperature: 30 °C; detection wavelengths: 280 nm, 220 nm; injection volume: 10 μg; sample tray temperature: approximately 10 °C; and run time: 50 min.

[0265] For cIEF experiments, Maurice TM A cIEF method development kit (ProteinSimple, catalog number: PS-MDK01-C) was used. Specifically, 8 μL of 30 μg of protein sample was mixed with the following reagents in the kit: 1 μL of Maurice TM cIEF pI Marker-7.05, 1 µL Maurice TM cIEF pI Marker-10.10, 35 μL of 1% methylcellulose solution, 2 μL of Maurice TMcIEF 500 mM arginine, 1.33 μL Ampholytes (Pharmalyte pH range 3-10), and 6.66 μL Ampholytes (Pharmalyte pH range 8-10.5) water (add to a final volume of 100 μL). TM Maurice TM Imaging capillary isoelectric focusing spectra were generated using a cIEF cartridge (PS-MC02-C). Samples were focused for a total of 10 min.

[0266] In the CE-SDS experiment, Maurice TM (ProteinSimple, Maurice) and Maurice™ CE-SDS Size Application Kit (ProteinSimple, Cat. No. PS-MAK02-S) were used.

[0267] For CE-SDS(NR), 30 μL of sample buffer, 30 μL of 30 μg of antibody sample, 1.5 μL of 25× internal standard, and 3 μL of 250 nM iodoacetamide (SIGMA, Cat. No. 16125) were added to a microcentrifuge tube, followed by centrifugation at 3,000 rpm for 1 minute and heating in a 70°C water bath for 10 minutes. The sample was then cooled to room temperature and centrifuged at 10,000 rpm for 3 minutes. The supernatant sample preparation was then transferred to a 96-well plate and analyzed by Maurice. TM The parameters used were: injection voltage 4.6 kV, injection time 20 sec, separation voltage 5.75 kV, and separation time 40 min.

[0268] For UNCle experiments, 8 μL of protein sample was loaded into a UniTube, and the system was operated at thermal temperatures ranging from 25 °C to 95 °C at a heating rate of 1 °C / min. Particle size and polydispersity were detected by dynamic light scattering (DLS) before heating. Protein stability was characterized by differential scanning full-spectrum fluorescence (DSF), static light scattering (SLS), and dynamic light scattering (DLS).

[0269] Detailed results for 11B12, 11F8, and 11F5 are shown in the table below. 11B12, 11F8, and 11F5 showed good stability and biophysical properties.

[0270] [Table 7]

[0271] [Table 8]

[0272] Example 7. Determination of antibody-dependent cellular cytotoxicity (ADCC) In the reporter gene bioassay (ADCC reporter bioassay), recombinant Jurkat cells (Jurkat-Luc-hCD16A-158V) were used as effector cells, and HEK293T cells bearing the hCCR8 surface antigen were used as target cells. To determine ADCC activity, cells were incubated with each antibody at a 4:1 ratio for 6 hours. The results are shown in the table below.

[0273] [Table 9]

[0274] All anti-CCR8 antibodies had ADCC activity almost equivalent to that of the positive control drug (7B16 analog), and the SI mutants showed stronger ADCC effects compared to the WT.

[0275] In another experiment, the ADCC activity of human IgG1 anti-CCR8 antibody and the CDR- and FR-modified anti-CCR8 antibodies was determined. The results are shown in the table below.

[0276] The results showed that the anti-CCR8 antibodies 11B12-11F8, 11F5-SG-11F5-SG, 11F8-SG-11F8-SG, 11B12-SG-11F8, and 11B12-DNA-11B12 had ADCC activity, among which 11B12-11F8, 11F5-SG-11F5-SG, 11F8-SG-11F8-SG, and 11B12-SG-11F8 showed better ADCC activity than 11B12, 11F5, and 11F8.

[0277] [Table 10]

[0278] Example 8. Antitumor activity of anti-CCR8 antibodies Anti-CCR8 antibodies were tested for their effect on tumor growth in vivo in a colon cancer model. 5 MC38 cells (mouse-derived colon cancer cells) were subcutaneously injected into each B-hCCR8 mouse (Biocytogen Pharmaceuticals (Beijing) Co., Ltd., Beijing, China, Catalog No.: 110096). Tumors in the mice were approximately 100 mm 3 When tumors reached a volume of 1000 mg / kg, the mice were randomly divided into groups based on tumor volume, and then injected intraperitoneally (ip) with phosphate-buffered saline (PBS) or anti-human CCR8 antibody.

[0279] The injection volume was calculated based on the mouse body weight and the 10 mg / kg dose. The tumor's major and minor axis lengths were measured, and the tumor volume was calculated as 0.5 × (major axis) × (minor axis). 2 It was calculated as:

[0280] Tumor growth inhibition percentage (TGI%) is calculated using the following formula: (TGI%)=[1-(Ti-T0) / (Vi-V0)]×100. Ti is the mean tumor volume in the treatment group on day i. T0 is the mean tumor volume in the treatment group on day 0. Vi is the mean tumor volume in the control group on day i. V0 is the mean tumor volume in the control group on day 0.

[0281] Values ​​are expressed as mean ± SEM. T-test was performed for statistical analysis. TGI% above 60% indicates a clear inhibition of tumor growth. P<0.05 was the threshold for indicating a significant difference.

[0282] [Table 11] The weight of all mice in each group increased. On the day of grouping (day 0), the mean weight of each group ranged from 19.8 g to 20.3 g. At the end of the experiment (day 21), the mean weight of each group ranged from 21.6 g to 24.6 g, and the mean weight of each group ranged from 111.4% to 121.8%. The results showed that the tested antibodies were well tolerated and had no apparent toxicity to mice.

[0283] Tumor sizes in the anti-CCR8 antibody-treated groups are shown in Figure 1. The table below lists the results for this experiment, including tumor volumes on the day of grouping (day 0), 7 days after grouping (day 7), 14 days after grouping (day 14), and at the end of the experiment (day 21), mouse survival rates, tumor growth inhibition scores (TGI), and statistical differences (P values) in tumor volume and body weight between the treatment and control groups.

[0284] 11F5 showed better tumor growth inhibitory effects than the positive control analogs 10A11 and 7B16.

[0285] [Table 12]

[0286] In another similar experiment, anti-CCR8 antibodies 11F5, 11F5-SI, and / or 11F8-SI were tested for their effect on in vivo tumor growth in a colon cancer model at a dose of 3 mg / kg.

[0287] Tables 13 and 14 summarize the tumor volumes, tumor growth inhibition values ​​(TGI) on the day of grouping (day 0), 14 days after grouping (day 14), and 21 days after grouping (day 21), as well as statistical differences (P values) in tumor volumes and body weights between the treatment and control groups in two different experiments.

[0288] [Table 13]

[0289] [Table 14]

[0290] 11F5, 11F5-SI, and 11F8-SI exhibited good tumor growth inhibitory effects in colon cancer, and 11F5-SI exhibited a better tumor growth inhibitory effect than 11F5.

[0291] Furthermore, in a separate experiment, 11F8-SI and 11B12-SI were also tested for their effects on tumor growth in a colon cancer model. 5 MC38 cells were subcutaneously injected into B-hPD-1 / hPD-L1 / hCCR8 mice (Biocytogen Pharmaceuticals (Beijing) Co., Ltd., Beijing, China, Catalog No.: 131764). Tumors in the mice were approximately 100–150 mm 3 When tumors reached a volume of 1000 mg / kg, the mice were randomly divided into groups (6 mice per group) based on tumor volume. The mice were then intraperitoneally injected with PBS (G1, control), 3 mg / kg 11F8-SI (G2), and 3 mg / kg 11B12-SI (G3). The antibodies were administered on days 1 and 4 of each week for 3 weeks (a total of 6 injections).

[0292] The results showed that 11F8-SI and 11B12-SI exhibited good tumor growth inhibitory effects with TGI% of 44.34% and 37.60%, respectively.

[0293] Example 9. Binding affinity verification of anti-CCR8 antibodies The binding activity of anti-CCR8 antibodies to 293T-hCCR8 cells (obtained by transfecting HEK293T cells to express human CCR8) and 293T-mCCR8 cells (HEK293T cells expressing mouse CCR8) was verified by flow cytometry. PE Goat α-Human IgG Fc (SouthernBiotech, Catalog No. 2014-09) was used as the secondary antibody. EC50 values ​​were determined using serially diluted sample anti-CCR8 antibodies. The results are shown in the table below.

[0294] [Table 15]

[0295] In another experiment, the binding activity of the CDR- and FR-modified anti-CCR8 antibodies to 293T-hCCR8 cells and 293T-mCCR8 was verified by flow cytometry. The results are summarized in the table below.

[0296] [Table 16]

[0297] Example 10. Epitope analysis of anti-hCCR8 antibodies To analyze the amino acid residues involved in the binding activity of anti-hCCR8 antibodies to human CCR8, we generated human CCR8 point mutants of amino acids in the extracellular domain (N-terminus, ECL1, ECL2, and ECL3), and then evaluated the binding activity of 11F5, 11F8, 11F8-SG-11F8-SG, 11B12-11F8, and 11B12-SG-11F8 mutants by flow cytometry. Analysis revealed that multiple amino acids in each extracellular domain (N-terminus, ECL1, ECL2, and ECL3) contribute to the binding activity of the clones. In particular, significant decreases in the binding activity of clones were observed in the mutants Y17A (Y at position 17 of human CCR8 (SEQ ID NO: 43) is replaced with A; the same applies below), F21A, D97A, W99A, F101A, G102A, C106A, G179A, C183A, W194A, K195A, I196A, and L278A. The F at position 21 is conserved between human and mouse CCR8. Binding to F21 may be important for cross-reactivity to mouse CCR8. The F21A mutant did not alter the binding activity of the 10A11 and 7B16 analogs. This may explain why these clones do not cross-react with mouse CCR8.

[0298] Example 11. Combination therapy with anti-PD-1 antibody To evaluate the efficacy of the combination therapy, the anti-CCR8 antibody was administered to mice along with another therapeutic agent, a pembrolizumab analogue.

[0299] Pembrolizumab analog is a humanized immunoglobulin G1 (IgG1) anti-PD-1 monoclonal antibody. The VH and VL sequences of pembrolizumab analog are shown in SEQ ID NO: 217 and SEQ ID NO: 218, respectively.

[0300] Approximately 5×10 5MC38 cells (mouse-derived colon cancer cells) were subcutaneously injected into B-hPD-1 / hPD-L1 / hCCR8 mice (Biocytogen Pharmaceuticals (Beijing) Co., Ltd., Beijing, China, Catalog No.: 131764). Tumors in the mice were approximately 100–150 mm 3 When tumors reached a volume of 1000 mg / mL, the mice were randomly assigned to groups (6 mice per group) based on tumor volume. The mice were then intraperitoneally injected with PBS (G1, control), 0.3 mg / kg of a pembrolizumab analog (G2), 3 mg / kg of 11F5 (G3), or a combination of 0.3 mg / kg of a pembrolizumab analog and 3 mg / kg of 11F5 (G4). Antibodies were administered on days 1 and 4 of each week for 3 weeks (a total of 6 injections).

[0301] The weight of the mice was monitored throughout the treatment period. All mice in each group gained weight. On the day of grouping (day 0), the mean weight of each group ranged from 18.2 g to 19.2 g. At the end of the experiment (day 28), the mean weight of each group ranged from 21.7 g to 23.5 g, and the mean weight of each group ranged from 114.4% to 123.3%. The results showed that the tested antibodies were well tolerated and had no apparent toxicity to the mice.

[0302] The table below lists the results for this experiment, including tumor volumes on the day of grouping (day 0), 14 days after grouping (day 14), and at the end of the experiment (day 28), mouse survival rates, tumor growth inhibition scores (TGI), and statistical differences (P values) in tumor volumes and body weights between the treatment and control groups.

[0303] [Table 17]

[0304] The tumor size of each treatment group is also shown in Figure 6. Compared to the control group (G1), the treatment groups (G2-G4) exhibited different antitumor effects. Furthermore, the combination of 11F5 and a pembrolizumab analog (G4) inhibited tumor growth with a higher TGI% than either 11F5 (G3) or a pembrolizumab analog (G2).

[0305] In another similar experiment, the combination of 11F5-SI and pembrolizumab analogs or 11B12-SI and pembrolizumab analogs was tested for their effects on in vivo tumor growth in a colon cancer model to evaluate the efficacy of the combination therapy, details of which are shown in the table below.

[0306] [Table 18]

[0307] The tumor size for each treatment group is shown in Figure 7. The combination of 11F5-SI and a pembrolizumab analog (G4) inhibited tumor growth with a significantly higher TGI% (77.49%) than either 11F5-SI (G3) or a pembrolizumab analog (G2). Similarly, the combination of 11B12-SI and a pembrolizumab analog (G6) inhibited tumor growth with a significantly higher TGI% (74.52%) than either 11B12-SI (G5) or a pembrolizumab analog (G2).

[0308] Example 12. Verification of binding affinity of 11F5 and its CDR3-modified antibody The binding activity of 11F5 and its CDR3-modified antibodies to 293T-hCCR8 and 293T-mCCR8 cells was verified by flow cytometry. The secondary antibody used in the experiment was PE Goat α-Human IgG Fc (SouthernBiotech, Catalog No.: 2014-09). EC50 was determined using serially diluted sample antibodies. The results are shown in the table below.

[0309] [Table 19]

[0310] We demonstrated that the anti-CCR8 antibody 11F5 and its CDR3-modified antibody can bind to human and mouse CCR8 with high affinity.

[0311] Example 13. Determination of antibody-dependent cellular cytotoxicity (ADCC) of 11F5 and its CDR3-modified antibodies The ADCC activity of 11F5 and its CDR3-modified antibodies was determined, and the results are shown in the table below.

[0312] Specifically, recombinant Jurkat cells (Jurkat-Luc-hCD16A-158V) were used as effector cells, and HEK293T cells bearing the hCCR8 surface antigen were used as target cells. To determine ADCC activity, the cells were incubated with each antibody at a 4:1 ratio for 6 hours. The results are shown in the table below.

[0313] [Table 20]

[0314] 11F5 and all of the CDR3-modified antibodies exhibited potent ADCC activity.

[0315] In another similar experiment, 11F5 and its CDR3-modified antibody with SI mutations in the Fc region were also assayed. The results are shown in the table below, and 11F5 and its CDR3-modified antibody showed stronger ADCC activity when SI mutations were introduced into the Fc region.

[0316] [Table 21]

[0317] Example 14. Biophysical analysis and stability of 11F5 and / or its CDR3 modified antibodies The biophysical properties of 11F5 and its CDR3-modified antibodies were evaluated. Experiments similar to those described in Example 6 were performed. In addition, colloidal stability was detected using stand-up monolayer chromatography (SMAC) (shown as the retention time of the main peak (SMAC, minutes)) and interaction with human serum polyclonal antibodies was detected using interaction chromatography (CIC) (shown as the retention time of the main peak (CIC, minutes)).

[0318] For SMAC experiments, an Agilent 1260 chromatographic system (Zenix TM A column (4.6 mm x 30 cm, Sepax, catalog number: 213300-4630) was used, and the sample was diluted to 1 mg / mL using PBS. The following parameters were used: mobile phase: 150 mM sodium phosphate (pH 7.0), flow rate: 0.35 mL / min, column temperature: 30°C, detection wavelengths: 280 nm, 220 nm, run time: 25 min, and injection volume: 2 μg.

[0319] For CIC experiments, an Agilent 1260 chromatographic system (connected to a 1 mL HiTrap® (GE Healthcare, catalog number: 17-0716-01)) was used with a human serum polyclonal antibody (Sigma, catalog number: I4506). Samples were diluted to 1 mg / mL using PBS. The following parameters were used: mobile phase: PBS (pH 7.2-7.4), flow rate: 0.1 mL / min, detection wavelengths: 280 nm, 220 nm, run time: 50 min, and injection volume: 10 μg.

[0320] [Table 22]

[0321] These 11F5 CDR3 modified antibodies showed good biophysical properties.

[0322] In a separate experiment, the stability of 11F5-N3S-Y5V-11F5-SI and 11F5-N3S-Y5V-Y6N-11F5-SI was evaluated. The results are shown in the table below, and 11F5-N3S-Y5V-11F5-SI and 11F5-N3S-Y5V-Y6N-11F5-SI showed good stability.

[0323] [Table 23]

[0324] Example 15. Blockade of calcium flux in hCCR8-expressing cells by 11F5 and its CDR3-modified antibodies Blockade of hCCL1 binding to hCCR8 by 11F5 and its CDR3-modified antibodies was tested by calcium (Ca) flux assay using human CCR8-expressing 293T cells. A Fluorescence Imaging Plate Reader (FLIPR) Calcium 6 Assay Kit (Molecular Devices, Catalog No. R8190) was used. Cells were seeded the day before the experiment and incubated overnight at 37°C and 5% CO2. Next, the cells were loaded with FLIPR Calcium 6 Dye (Molecular Devices) containing probenecid (Invitrogen) and PowerLoad (Invitrogen) and incubated at 37°C for 30 minutes. Anti-CCR8 antibodies were added to the cells and further incubated at 37°C for 30 minutes. After incubation, Ca flux fluorescence signals induced by 1 μg / mL recombinant human CCL1 (BioLegend, Catalog No. 582706) were measured for 1 minute using a SpectraMax® iD3 (Molecular Devices). The results are shown in Figure 8 and in the table below.

[0325] [Table 24]

[0326] In calcium flux blocking experiments, the CDR3-modified 11F5 antibodies 11F5-N3S-11F5, 11F5-Y6N-11F5, 11F5-N3S-Y5V-11F5, and 11F5-N3S-Y5V-Y6N-11F5 showed superior blocking activity compared to 11F5. In particular, 11F5-N3S-Y5V-11F5 showed almost complete inhibition.

[0327] Example 16. Antitumor activity of 11F5 and its CDR3-modified antibodies 11F5 and its CDR3-modified antibodies were tested for their effects on in vivo tumor growth in a colon cancer model. 5 MC38 cells were injected subcutaneously into each B-hCCR8 mouse. Tumors in the mice were approximately 150 mm 3 When the tumor volume reached 100 μg / ml, the mice were randomly divided into groups based on tumor volume, and then injected intraperitoneally (ip) with phosphate-buffered saline (PBS), 11F5, or its CDR3-modified antibody.

[0328] The injection volume was calculated based on the mouse weight and a dose of 3 mg / kg (details shown in the table below).

[0329] [Table 25]

[0330] The table below summarizes the results of this experiment, including tumor volumes on the day of grouping (day 0), 14 days after grouping (day 14), and 21 days after the experiment (day 21), mouse survival rates, tumor growth inhibition scores (TGI), and statistical differences (P values) in tumor volumes and body weights between the treatment and control groups.

[0331] [Table 26]

[0332] 11F5-N3S-Y5V-11F5-SI showed a better tumor growth inhibitory effect than 11F5-SI.

[0333] Other embodiments While the present invention has been described in conjunction with its detailed description, it should be understood that the above description is intended to be illustrative and not limiting of the scope of the invention, which is defined by the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

Claims

1. An antibody or antigen-binding fragment thereof that binds to CCR8 (chemokine (C-C motif) receptor 8), a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VH CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR1, the VH CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR2, and the VH CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR3; a light chain variable region (VL) comprising CDR1, 2, and 3, wherein the VL CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR1, the VL CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR2, and the VL CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR3; wherein the selected VH CDR1, 2, and 3 amino acid sequences are as follows: (1) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 1, 2, and 3, respectively; (2) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 4, 5, and 6, respectively; (3) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 7, 8, and 9, respectively; (4) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 10, 11, and 12, respectively; (5) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 13, 14, and 15, respectively; (6) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 16, 17, and 18, respectively; (7) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 72, 73, and 74, respectively; (8) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 75, 76, and 77, respectively; (9) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 78, 79, and 80, respectively; (10) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 81, 82, and 83, respectively; (11) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 84, 85, and 86, respectively; (12) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 87, 88, and 89, respectively; (13) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 90, 91, and 92, respectively; (14) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 93, 94, and 95, respectively; (15) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 96, 97, and 98, respectively; (16) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 144, 145, and 146, respectively; (17) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 147, 148, and 149, respectively; (18) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 150, 151, and 152, respectively; (19) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 153, 154, and 155, respectively; (20) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 156, 157, and 158, respectively; (21) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 159, 160, and 161, respectively; (22) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 162, 163, and 164, respectively; (23) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 165, 166, and 167, respectively; (24) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 168, 169, and 170, respectively; (25) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 219, 220, and 221, respectively; (26) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 222, 223, and 224, respectively; (27) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 225, 226, and 227, respectively; (28) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 228, 229, and 230, respectively; (29) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 231, 232, and 233, respectively; (30) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 234, 235, and 236, respectively; (31) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 237, 238, and 239, respectively; (32) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 240, 241, and 242, respectively; (33) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 243, 244, and 245, respectively; (34) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 246, 247, and 248, respectively. (35) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 249, 250, and 251, respectively; (36) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 252, 253, and 254, respectively; (37) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 255, 256, and 257, respectively; and (38) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 258, 259, and 260, respectively. and The amino acid sequences of the selected VL CDR1, 2, and 3 are as follows: (1) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 19, 20, and 21, respectively; (2) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 22, 23, and 24, respectively; (3) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 25, 26, and 27, respectively; (4) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 28, 29, and 30, respectively; (5) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 31, 32, and 33, respectively; (6) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 34, 35, and 36, respectively; (7) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 108, 109, and 110, respectively; (8) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 111, 112, and 113, respectively; (9) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 114, 115, and 116, respectively; (10) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 117, 118, and 119, respectively; (11) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 120, 121, and 122, respectively; (12) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 123, 124, and 125, respectively; (13) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 126, 127, and 128, respectively; (14) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 129, 130, and 131, respectively; (15) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 132, 133, and 134, respectively; (16) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 180, 181, and 182, respectively; (17) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 183, 184, and 185, respectively; (18) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 186, 187, and 188, respectively; (19) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 189, 190, and 191, respectively; (20) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 192, 193, and 194, respectively; (21) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 195, 196, and 197, respectively; (22) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 198, 199, and 200, respectively; (23) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 201, 202, and 203, respectively; and (24) The amino acid sequences of the selected VL CDR1, 2, and 3 are shown in SEQ ID NOs: 204, 205, and 206, respectively. an antibody or antigen-binding fragment thereof,

2. An antibody or antigen-binding fragment thereof that binds to CCR8 (chemokine (C-C motif) receptor 8), a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VH CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR1, the VH CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR2, and the VH CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VH CDR3; a light chain variable region (VL) comprising CDR1, 2, and 3, wherein the VL CDR1 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR1, the VL CDR2 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR2, and the VL CDR3 region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of a selected VL CDR3; wherein the selected VH CDR1, 2, and 3 amino acid sequences are as follows: (1) The selected VH CDR1 is SYVMH (SEQ ID NO: 99) and the selected VH CDR2 is VISYX １ X ２ SDKYYADSVKG (SEQ ID NO: 100), and the selected VH CDR3 is GRGYRYGQSYYYGMDV (SEQ ID NO: 101); (2) the selected VH CDR1 is SYVVH (SEQ ID NO: 102) and the selected VH CDR2 is VISYX １ X ２ DNKFYADSVKG (SEQ ID NO: 103), and the selected VH CDR3 is GRX ５ YX ６ X ７ YYGLDV (SEQ ID NO: 104), (3) the selected VH CDR1 is TYVMH (SEQ ID NO: 105) and the selected VH CDR2 is VISYX １ X ２ NNKYYADSVKG (SEQ ID NO: 106), and the selected VH CDR3 is GRSYVNYYGLDV (SEQ ID NO: 107); (4) The selected VH CDR1 is GFSFSSY (SEQ ID NO: 171) and the selected VH CDR2 is SYX １ X ２ SD (SEQ ID NO: 172), and the selected VH CDR3 is GRGYRYGQSYYYGMDV (SEQ ID NO: 173); (5) The selected VH CDR1 is GFTFSSY (SEQ ID NO: 174) and the selected VH CDR2 is SYX １ X ２ DN (SEQ ID NO: 175), and the selected VH CDR3 is GRX ５ YX ６ X ７ YYGLDV (SEQ ID NO: 176), and (6) The selected VH CDR1 is GFTFSTY (SEQ ID NO: 177) and the selected VH CDR2 is SYX １ X ２ NN (SEQ ID NO: 178), and the selected VH CDR3 is GRSYVNYYGLDV (SEQ ID NO: 179); and The amino acid sequences of the selected VL CDR1, 2, and 3 are as follows: (1) The selected VL CDR1 is KSSQSLLYSX ３ X ４ KTYLY (SEQ ID NO: 135), wherein the selected VL CDR2 is EVSNRFS (SEQ ID NO: 136), and the selected VL CDR3 is MQSIKLPLT (SEQ ID NO: 137); (2) the selected VL CDR1 is KSSQSLLYSX ３ X ４ KTYLY (SEQ ID NO: 138), wherein the selected VL CDR2 is EVSNRFS (SEQ ID NO: 139), and the selected VL CDR3 is MQSLKVPPT (SEQ ID NO: 140); (3) the selected VL CDR1 is KSSQSLLYSX ３ X ４ KTYLY (SEQ ID NO: 141), wherein the selected VL CDR2 is EVSNRFS (SEQ ID NO: 142), and the selected VL CDR3 is MQSVKIPLT (SEQ ID NO: 143); (4) the selected VL CDR1 is KSSQSLLYSX ３ X ４ KTYLY (SEQ ID NO: 207), wherein the selected VL CDR2 is EVSNRFS (SEQ ID NO: 208), and the selected VL CDR3 is MQSIKLPLT (SEQ ID NO: 209); (5) The selected VL CDR1 is KSSQSLLYSX ３ X ４ KTYLY (SEQ ID NO:210), wherein the selected VL CDR2 is EVSNRFS (SEQ ID NO:211), and the selected VL CDR3 is MQSLKVPPT (SEQ ID NO:212); and (6) The selected VL CDR1 is KSSQSLLYSX ３ X ４ KTYLY (SEQ ID NO:213), wherein the selected VL CDR2 is EVSNRFS (SEQ ID NO:214), and the selected VL CDR3 is MQSVKIPLT (SEQ ID NO:215); One of the following, provided that: X in each sequence １ and X ３ is independently selected from the group consisting of D, S, and A; X in each sequence ２ and X ４ is independently selected from the group consisting of G and A; X of SEQ ID NO: 104 or 176 ５ are independently selected from the group consisting of N and S; X of SEQ ID NO: 104 or 176 ６ is independently selected from the group consisting of Y and V; X of SEQ ID NO: 104 or 176 ７ is independently selected from the group consisting of Y and N.

3. 2. The antibody or antigen-binding fragment thereof of claim 1, wherein the selected VH CDR1, 2, and 3 amino acid sequences and the selected VL CDR1, 2, and 3 amino acid sequences are any one of the following: (1) According to the Kabat definition, the VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively; (2) According to the Kabat definition, the VH comprises CDR1, 2, and 3 having the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences set forth in SEQ ID NOs: 22, 23, and 24, respectively; (3) According to the Kabat definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 7, 8, and 9, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; (4) According to the Chothia definition, the VH comprises CDR1, 2, 3 having the amino acid sequences set forth in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences set forth in SEQ ID NOs: 28, 29, and 30, respectively; (5) According to the Chothia definition, the VH comprises CDR1, 2, 3 having the amino acid sequences set forth in SEQ ID NOs: 13, 14, and 15, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences set forth in SEQ ID NOs: 31, 32, and 33, respectively; (6) According to the Chothia definition, the VH comprises CDR1, 2, 3 having the amino acid sequences set forth in SEQ ID NOs: 16, 17, and 18, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences set forth in SEQ ID NOs: 34, 35, and 36, respectively; (7) According to the Kabat definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (8) According to the Kabat definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; (9) According to the Kabat definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 4, 5, and 6, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively; (10) According to the Kabat definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 4, 5, and 6, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; (11) According to the Kabat definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 7, 8, and 9, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively; (12) According to the Kabat definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 7, 8, and 9, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (13) According to the Chothia definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (14) According to the Chothia definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively; (15) According to the Chothia definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 13, 14, and 15, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 28, 29, and 30, respectively; (16) According to the Chothia definition, the VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 13, 14, and 15, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively; (17) According to the Chothia definition, the VH comprises CDR1, 2, 3 having the amino acid sequences set forth in SEQ ID NOs: 16, 17, and 18, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences set forth in SEQ ID NOs: 28, 29, and 30, respectively; and (18) According to the Chothia definition, the VH comprises CDR1, 2, and 3 having the amino acid sequences set forth in SEQ ID NOs: 16, 17, and 18, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences set forth in SEQ ID NOs: 31, 32, and 33, respectively.

4. 2. The antibody or antigen-binding fragment thereof of claim 1, wherein the selected VH CDR1, 2, and 3 amino acid sequences and the selected VL CDR1, 2, and 3 amino acid sequences are any one of the following: (1) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 114, 115, and 116, respectively; (2) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 72, 73, and 74, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 114, 115, and 116, respectively; (3) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 72, 73, and 74, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 108, 109, and 110, respectively; (4) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 78, 79, and 80, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 114, 115, and 116, respectively; (5) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 75, 76, and 77, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 111, 112, and 113, respectively; (6) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 186, 187, and 188, respectively; (7) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 144, 145, and 146, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 186, 187, and 188, respectively; (8) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 144, 145, and 146, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 180, 181, and 182, respectively; (9) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 150, 151, and 152, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 186, 187, and 188, respectively; (10) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 147, 148, and 149, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 183, 184, and 185, respectively; (11) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 72, 73, and 74, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; (12) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 144, 145, and 146, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively; (13) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 219, 220, and 221, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (14) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 222, 223, and 224, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (15) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 225, 226, and 227, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (16) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 228, 229, and 230, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (17) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 231, 232, and 233, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (18) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 234, 235, and 236, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (19) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 237, 238, and 239, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; (20) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 240, 241, and 242, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (21) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 243, 244, and 245, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (22) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 246, 247, and 248, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (23) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 249, 250, and 251, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (24) The VH comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 252, 253, and 254, respectively, and the VL comprises CDR1, 2, 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; (25) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 255, 256, and 257, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; and (26) The VH comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 258, 259, and 260, respectively, and the VL comprises CDR1, 2, and 3 having the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively.

5. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 4, which specifically binds to human CCR8, mouse CCR8, or monkey CCR8.

6. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 5, wherein the antibody or antigen-binding fragment is a human or humanized antibody or antigen-binding fragment thereof (e.g., a human IgG1 antibody).

7. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, wherein the antibody or antigen-binding fragment is a single-chain variable fragment (scFv), a one-armed antibody, or a multispecific antibody (e.g., a bispecific antibody).

8. A nucleic acid comprising a polynucleotide encoding a polypeptide, said polypeptide comprising an immunoglobulin heavy chain or a fragment thereof comprising a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, said VH binding to CCR8 when paired with a light chain variable region (VL); The amino acid sequences of the VH CDR1, 2, and 3 are as follows: (1) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 1, 2, and 3, respectively; (2) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 4, 5, and 6, respectively; (3) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 7, 8, and 9, respectively; (4) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 10, 11, and 12, respectively; (5) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 13, 14, and 15, respectively; (6) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 16, 17, and 18, respectively; (7) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 72, 73, and 74, respectively; (8) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 75, 76, and 77, respectively; (9) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 78, 79, and 80, respectively; (10) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 81, 82, and 83, respectively; (11) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 84, 85, and 86, respectively; (12) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 87, 88, and 89, respectively; (13) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 90, 91, and 92, respectively; (14) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 93, 94, and 95, respectively; (15) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 96, 97, and 98, respectively; (16) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 99, 100, and 101, respectively; (17) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 102, 103, and 104, respectively; (18) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 105, 106, and 107, respectively; (19) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 144, 145, and 146, respectively; (20) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 147, 148, and 149, respectively; (21) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 150, 151, and 152, respectively; (22) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 153, 154, and 155, respectively; (23) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 156, 157, and 158, respectively; (24) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 159, 160, and 161, respectively; (25) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 162, 163, and 164, respectively; (26) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 165, 166, and 167, respectively; (27) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 168, 169, and 170, respectively; (28) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 171, 172, and 173, respectively; (29) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 174, 175, and 176, respectively; (30) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 177, 178, and 179, respectively; (31) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 219, 220, and 221, respectively; (32) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 222, 223, and 224, respectively; (33) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 225, 226, and 227, respectively. (34) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 228, 229, and 230, respectively; (35) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 231, 232, and 233, respectively; (36) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 234, 235, and 236, respectively; (37) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 237, 238, and 239, respectively; (38) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 240, 241, and 242, respectively; (39) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 243, 244, and 245, respectively; (40) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 246, 247, and 248, respectively; (41) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 249, 250, and 251, respectively; (42) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 252, 253, and 254, respectively; (43) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 255, 256, and 257, respectively; and (44) The amino acid sequences of the selected VH CDR1, 2, and 3 are set forth in SEQ ID NOs: 258, 259, and 260, respectively; and A nucleic acid wherein the VL amino acid sequence is selected from the group consisting of SEQ ID NOs: 40, 41, 42, 66, 67, and 68.

9. 9. The nucleic acid of claim 8, wherein the nucleic acid comprises a polynucleotide encoding a polypeptide, the polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, comprising a VH comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3, respectively.

10. 9. The nucleic acid of claim 8, wherein the nucleic acid comprises a polynucleotide encoding a polypeptide, the polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, comprising a VH comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6, respectively.

11. 9. The nucleic acid of claim 8, wherein the nucleic acid comprises a polynucleotide encoding a polypeptide, the polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, comprising a VH comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 7, 8, and 9, respectively.

12. A nucleic acid comprising a polynucleotide encoding a polypeptide, said polypeptide comprising an immunoglobulin heavy chain or a fragment thereof comprising a light chain variable region (VL) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein said VL binds to CCR8 when paired with a heavy chain variable region (VH); The amino acid sequences of the selected VL CDRs 1, 2, and 3 are as follows: (1) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 19, 20, and 21, respectively; (2) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 22, 23, and 24, respectively; (3) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 25, 26, and 27, respectively; (4) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 28, 29, and 30, respectively; (5) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 31, 32, and 33, respectively; (6) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 34, 35, and 36, respectively; (7) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 108, 109, and 110, respectively; (8) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 111, 112, and 113, respectively; (9) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 114, 115, and 116, respectively; (10) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 117, 118, and 119, respectively; (11) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 120, 121, and 122, respectively; (12) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 123, 124, and 125, respectively; (13) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 126, 127, and 128, respectively; (14) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 129, 130, and 131, respectively; (15) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 132, 133, and 134, respectively; (16) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 135, 136, and 137, respectively; (17) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 138, 139, and 140, respectively; (18) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 141, 142, and 143, respectively; (19) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 180, 181, and 182, respectively; (20) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 183, 184, and 185, respectively; (21) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 186, 187, and 188, respectively; (22) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 189, 190, and 191, respectively; (23) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 192, 193, and 194, respectively; (24) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 195, 196, and 197, respectively; (25) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 198, 199, and 200, respectively; (26) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 201, 202, and 203, respectively; (27) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 204, 205, and 206, respectively; (28) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 207, 208, and 209, respectively; (29) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 210, 211, and 212, respectively; and (30) The amino acid sequences of the selected VL CDR1, 2, and 3 are set forth in SEQ ID NOs: 213, 214, and 215, respectively; A nucleic acid wherein the VH amino acid sequence is selected from the group consisting of SEQ ID NOs: 37, 38, 39, 63, 64, 65, 69, 70, 71, 216, 261, 262, 263, 264, 265, 266, and 267.

13. 13. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding a polypeptide, the polypeptide comprising an immunoglobulin light chain or a fragment thereof, comprising a VL comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 19, 20, and 21, respectively.

14. 13. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding a polypeptide, the polypeptide comprising an immunoglobulin light chain or a fragment thereof, comprising a VL comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 22, 23, and 24, respectively.

15. 13. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding a polypeptide, the polypeptide comprising an immunoglobulin light chain or a fragment thereof, comprising a VL comprising CDR1, 2, and 3 comprising the amino acid sequences set forth in SEQ ID NOs: 25, 26, and 27, respectively.

16. The nucleic acid according to any one of claims 8 to 15, wherein the VH specifically binds to human CCR8, mouse CCR8, or monkey CCR8 when paired with the VL, or the VL specifically binds to human CCR8, mouse CCR8, or monkey CCR8 when paired with the VH.

17. 17. The nucleic acid of any one of claims 8 to 16, wherein the immunoglobulin heavy chain or fragment thereof is a human or humanized immunoglobulin heavy chain or fragment thereof, and the immunoglobulin light chain or fragment thereof is a human or humanized immunoglobulin light chain or fragment thereof.

18. 18. The nucleic acid of any one of claims 8 to 17, wherein the nucleic acid encodes a single-chain variable fragment (scFv), a one-armed antibody, or a multispecific antibody (e.g., a bispecific antibody).

19. The nucleic acid according to any one of claims 8 to 18, wherein the nucleic acid is a cDNA.

20. A vector comprising one or more of the nucleic acids according to any one of claims 8 to 19.

21. A vector comprising two of the nucleic acids according to any one of claims 8 to 19, wherein the vector encodes the VH region and the VL region that comprehensively bind to CCR8.

22. A pair of vectors, each of which comprises one of the nucleic acids according to any one of claims 8 to 19 and which comprehensively encode the VH region and the VL region that comprehensively bind to CCR8.

23. A cell comprising a vector according to claim 20 or 21, or a pair of vectors according to claim 22.

24. 24. The cell of claim 23, wherein the cell is a CHO cell.

25. A cell comprising any one or more of the nucleic acids according to any one of claims 8 to 19.

26. A cell comprising any two of the nucleic acids according to any one of claims 8 to 19.

27. The cell of claim 26 , wherein the two nucleic acids comprehensively encode the VH region and the VL region that comprehensively bind to CCR8.

28. 1. A method for producing an antibody or antigen-binding fragment thereof, comprising: (a) culturing the cell of any one of claims 23 to 27 under conditions sufficient for the cell to produce the antibody or antigen-binding fragment; (b) harvesting the antibody or antigen-binding fragment produced by the cell; and A method comprising:

29. An antibody or antigen-binding fragment thereof that binds to CCR8, a heavy chain variable region (VH) comprising an amino acid sequence that is at least 80% identical to a selected VH sequence, and a light chain variable region (VL) comprising an amino acid sequence that is at least 80% identical to a selected VL sequence; the selected VH sequence is selected from the group consisting of SEQ ID NOs: 37, 38, 39, 63, 64, 65, 69, 70, 71, 216, 261, 262, 263, 264, 265, 266, and 267; The selected VL sequence is selected from the group consisting of SEQ ID NOs: 40, 41, 42, 66, 67, and 68. An antibody or antigen-binding fragment thereof.

30. 30. The antibody or antigen-binding fragment thereof of claim 29, wherein the VH sequence and the VL sequence are any one of the following: (1) The VH comprises the sequence of SEQ ID NO: 37, and the VL comprises the sequence of SEQ ID NO: 40; (2) The VH comprises the sequence of SEQ ID NO: 38, and the VL comprises the sequence of SEQ ID NO: 41; (3) The VH comprises the sequence of SEQ ID NO: 39, and the VL comprises the sequence of SEQ ID NO: 42; (4) The VH comprises the sequence of SEQ ID NO: 37, and the VL comprises the sequence of SEQ ID NO: 41; (5) The VH comprises the sequence of SEQ ID NO: 37, and the VL comprises the sequence of SEQ ID NO: 42; (6) The VH comprises the sequence of SEQ ID NO: 38, and the VL comprises the sequence of SEQ ID NO: 40; (7) The VH comprises the sequence of SEQ ID NO: 38, and the VL comprises the sequence of SEQ ID NO: 42; (8) The VH comprises the sequence of SEQ ID NO: 39, and the VL comprises the sequence of SEQ ID NO: 40; (9) The VH comprises the sequence of SEQ ID NO: 39, and the VL comprises the sequence of SEQ ID NO: 41; (10) The VH comprises the sequence of SEQ ID NO: 37, and the VL comprises the sequence of SEQ ID NO: 68; (11) The VH comprises the sequence of SEQ ID NO: 63, and the VL comprises the sequence of SEQ ID NO: 68; (12) The VH comprises the sequence of SEQ ID NO: 63, and the VL comprises the sequence of SEQ ID NO: 66; (13) The VH comprises the sequence of SEQ ID NO: 65, and the VL comprises the sequence of SEQ ID NO: 68; (14) The VH comprises the sequence of SEQ ID NO: 64, and the VL comprises the sequence of SEQ ID NO: 67; (15) The VH comprises the sequence of SEQ ID NO: 63, and the VL comprises the sequence of SEQ ID NO: 42; (16) The VH comprises the sequence of SEQ ID NO: 216, and the VL comprises the sequence of SEQ ID NO: 42; (17) The VH comprises the sequence of SEQ ID NO: 261, and the VL comprises the sequence of SEQ ID NO: 41; (18) The VH comprises the sequence of SEQ ID NO: 262, and the VL comprises the sequence of SEQ ID NO: 41; (19) The VH comprises the sequence of SEQ ID NO: 263, and the VL comprises the sequence of SEQ ID NO: 41; (20) The VH comprises the sequence of SEQ ID NO: 264, and the VL comprises the sequence of SEQ ID NO: 41; (21) The VH comprises the sequence of SEQ ID NO: 265, and the VL comprises the sequence of SEQ ID NO: 41; (22) The VH comprises the sequence of SEQ ID NO: 266, and the VL comprises the sequence of SEQ ID NO: 41; and (23) The VH comprises the sequence of SEQ ID NO: 267, and the VL comprises the sequence of SEQ ID NO:

41.

31. The antibody or antigen-binding fragment thereof of claim 29 or 30, wherein the antibody or antigen-binding fragment specifically binds to human CCR8, mouse CCR8, or monkey CCR8.

32. The antibody or antigen-binding fragment thereof according to any one of claims 29 to 31, wherein the antibody or antigen-binding fragment is a human or humanized antibody or antigen-binding fragment thereof.

33. 33. The antibody or antigen-binding fragment thereof of any one of claims 29 to 32, wherein the antibody or antigen-binding fragment is a single-chain variable fragment (scFv), a one-armed antibody, or a multispecific antibody (e.g., a bispecific antibody).

34. 34. An antibody or antigen-binding fragment thereof that binds to CCR8, comprising a heavy chain variable region (VH) comprising VH CDR1, 2, and 3, and a light chain variable region comprising VL CDR1, 2, and 3, wherein the VH CDR1, 2, and 3 and the VL CDR1, 2, and 3 are identical to the complementarity-determining regions in the antibody or antigen-binding fragment thereof according to any one of claims 1 to 7 and 29 to 33.

35. 35. The antibody or antigen-binding fragment thereof of claim 34, wherein the VH CDR1, 2, and 3 are identical to the complementarity-determining regions of SEQ ID NO: 37, 38, 39, 63, 64, 65, 69, 70, 71, 216, 261, 262, 263, 264, 265, 266, or 267, and the VL CDR1, 2, and 3 are identical to the complementarity-determining regions of SEQ ID NO: 40, 41, 42, 66, 67, or 68.

36. An antibody or an antigen-binding fragment thereof that cross-competes with the antibody or antigen-binding fragment thereof according to any one of claims 1 to 7 and 29 to 35.

37. 37. An antibody drug conjugate comprising the antibody or antigen-binding fragment thereof of any one of claims 1 to 7 and 29 to 36 covalently attached to a therapeutic agent.

38. 38. The antibody drug conjugate of claim 37, wherein the therapeutic agent is a cytotoxic or cytostatic agent.

39. A method for treating a subject suffering from cancer, the method comprising administering to the subject a therapeutically effective amount of a composition comprising the antibody or antigen-binding fragment thereof of any one of claims 1 to 7 and 29 to 36, or the antibody-drug conjugate of claim 37 or 38.

40. 40. The method of claim 39, wherein the subject is a subject suffering from a solid cancer.

41. 41. The method of claim 39 or 40, wherein the cancer is breast cancer, malignant melanoma, non-small cell lung cancer, lung adenocarcinoma, colon cancer, or colorectal cancer (e.g., colorectal adenocarcinoma).

42. 42. The method of any one of claims 39 to 41, wherein the cancer is resistant to PD-1 pathway inhibitor therapy.

43. 43. The method of any one of claims 39 to 42, wherein the method further comprises administering to the subject a PD-1 pathway inhibitor.

44. 44. The method of claim 43, wherein the PD-1 pathway inhibitor is an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, or cemiplimab).

45. 44. The method of claim 43, wherein the PD-1 pathway inhibitor is an anti-PD-L1 antibody (e.g., atezolizumab, avelumab, or durvalumab).

46. 1. A method for reducing tumor growth rate, comprising: contacting tumor cells with an effective amount of a composition comprising the antibody or antigen-binding fragment thereof of any one of claims 1 to 7 and 29 to 36, or the antibody-drug conjugate of claim 37 or 38; method.

47. 1. A method for killing tumor cells, comprising: contacting tumor cells with an effective amount of a composition comprising the antibody or antigen-binding fragment thereof of any one of claims 1 to 7 and 29 to 36, or the antibody-drug conjugate of claim 37 or 38; method.

48. A pharmaceutical composition comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 7 and 29 to 36, and a pharmaceutically acceptable carrier.

49. 39. A pharmaceutical composition comprising the antibody-drug conjugate of claim 37 or 38 and a pharmaceutically acceptable carrier.

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