Antibodies and fusion proteins that bind to CCR8 and uses thereof

Abstract

To provide antibodies and fusion proteins that bind to CCR8, and uses thereof.SOLUTION: Provided herein are various embodiments relating to antibodies and fusion proteins and to uses thereof. Some of the embodiments include antibodies that bind to CCR8. Some of the embodiments include fusion proteins that bind to CCR8. Such antibodies and fusion proteins can be used in methods for treating, e.g., cancer. There remains a need for antibodies and fusion proteins that bind to CCR8 for treatment of cancer and other diseases and disorders.SELECTED DRAWING: None

Description

[Technical Field]

[0001] (Cross Reference Statement) This application claims priority under 35 U.S.C. §119(e) to U.S. Provisional Patent Application No. 62 / 976,869, filed February 14, 2020, and U.S. Provisional Patent Application No. 63 / 130,157, filed December 23, 2020. The entire contents of both provisional applications are incorporated herein by reference.

[0002] (Sequence Listing) This application contains a Sequence Listing that has been submitted electronically in ASCII format and is incorporated herein by reference in its entirety. The ASCII copy created on February 7, 2021 is named 119835-0178_SL.txt and is 157KB in size.

[0003] FIELD OF THE INVENTION Antibodies and fusion proteins that bind to CCR8 are provided, as are methods of treatment that include administering such antibodies and / or fusion proteins. [Background technology]

[0004] Chemokine (CC motif) receptor 8 (CCR8) belongs to the G protein-coupled receptor (GPCR) family. CCR8 is primarily expressed on tumor regulatory T (Treg) cells, a type of immune suppressor cell found in the tumor microenvironment.

[0005] There remains a need for antibodies and fusion proteins that bind to CCR8 for the treatment of cancer and other diseases and disorders. Summary of the Invention

[0006] The present disclosure describes an isolated antibody that binds to human CCR8, the antibody comprising: a) HCDR1 comprising the amino acid sequence of SEQ ID NO: 12, HCDR2 comprising the amino acid sequence of SEQ ID NO: 13, HCDR3 comprising the amino acid sequence of SEQ ID NO: 14, LCDR1 comprising the amino acid sequence of SEQ ID NO: 15, LCDR2 comprising the amino acid sequence of SEQ ID NO: 16, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 17; b) HCDR1 comprising the amino acid sequence of SEQ ID NO: 24, HCDR2 comprising the amino acid sequence of SEQ ID NO: 25, HCDR3 comprising the amino acid sequence of SEQ ID NO: 26, LCDR1 comprising the amino acid sequence of SEQ ID NO: 27, LCDR2 comprising the amino acid sequence of SEQ ID NO: 28, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 29; c) HCDR1 comprising the amino acid sequence of SEQ ID NO: 36, HCDR2 comprising the amino acid sequence of SEQ ID NO: 37, HCDR3 comprising the amino acid sequence of SEQ ID NO: 38, LCDR1 comprising the amino acid sequence of SEQ ID NO: 39, LCDR2 comprising the amino acid sequence of SEQ ID NO: 40, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 41; d) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 48, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 49, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 50, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 51, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 52, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 53, or e) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65, or f) Comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0007] The present application also describes an isolated antibody, the antibody comprising: a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65, or b) Comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0008] In some embodiments, the antibody a) a heavy chain variable region (VH) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 68 or 74, and a light chain variable region (VL) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 69 or 75; or b) a heavy chain variable region (VH) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 92 or 96, and a light chain variable region (VL) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 93 or 97.

[0009] In some embodiments, the antibody a) a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 68 or 74, and a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO: 69 or 75, or b) A heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 92 or 96, and a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO: 93 or 97.

[0010] In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is a humanized antibody. In some embodiments, the antibody is a full-length antibody. In some embodiments, the antibody is an IgG1 or IgG3 antibody.

[0011] In some embodiments, the antibody a) a heavy chain (HC) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 70 or 76, and a light chain (LC) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 71 or 77, or b) a heavy chain (HC) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 94 or 98, and a light chain (LC) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 95 or 99.

[0012] In some embodiments, the antibody comprises a heavy chain (HC) comprising the amino acid sequence of SEQ ID NO: 70 or 76, and a light chain (LC) comprising the amino acid sequence of SEQ ID NO: 71 or 77, or a heavy chain (HC) comprising the amino acid sequence of SEQ ID NO: 94 or 98, and a light chain (LC) comprising the amino acid sequence of SEQ ID NO: 95 or 99.

[0013] In some embodiments, the antibody comprises at least one modification that enhances cell killing. In some embodiments, the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). In some embodiments, the at least one modification is afucosylation. In some embodiments, the at least one modification is one or more heavy chain constant region mutations at one or more positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396. In some embodiments, the one or more heavy chain constant region mutations are one or more mutations selected from S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L. In some embodiments, the one or more heavy chain constant region mutations are selected from F243L / R292P / Y300L / V305I / P396L, S239D / I332E, S239D / I332E / A330L, S298A / E333A / K334A, L234Y / L235Q / G236W / S239M / H268D / D270E / S298A, and D270E / K326D / A330M / K334E. In some embodiments, at least one modification is galactosylation.

[0014] In some embodiments, the antibody has an affinity (K) (e.g., as determined by a Kinetic Exclusion Assay (i.e., KinExA)) of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75 pM, or less than 50 pM, or less than 25 pM. D ) In some embodiments, the antibody binds to human CCR8 with an on-cell affinity (K) of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75, or less than 50 pM, or less than 25 pM, as determined, for example, by an equilibrium binding exclusion assay (i.e., KinExA). D ) binds to human CCR8.

[0015] The present disclosure also describes isolated nucleic acids encoding the antibodies. In some embodiments, a vector comprises the isolated nucleic acid. In some embodiments, a host cell comprises the nucleic acid or vector. In some embodiments, the host cell expresses the antibody. In some embodiments, the host cell is engineered to produce an afucosylated antibody.

[0016] In some embodiments, the method for producing an antibody that binds to CCR8 comprises culturing a host cell under conditions suitable for expressing the antibody. In some embodiments, the method further comprises isolating the antibody.

[0017] In some embodiments, the fusion protein comprises (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region. In some embodiments, the Fc region is an IgG1 or IgG3 Fc region. In some embodiments, the Fc region comprises the amino acid sequence of SEQ ID NO: 4.

[0018] In some embodiments, the Fc region comprises at least one modification that enhances cell killing. In some embodiments, the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). In some embodiments, the at least one modification is afucosylation. In some embodiments, the at least one modification is one or more Fc region mutations at one or more positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396. In some embodiments, the one or more Fc region mutations are one or more mutations selected from S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L. In some embodiments, the one or more Fc region mutations are selected from F243L / R292P / Y300L / V305I / P396L, S239D / I332E, S239D / I332E / A330L, S298A / E333A / K334A, L234Y / L235Q / G236W / S239M / H268D / D270E / S298A, and D270E / K326D / A330M / K334E. In some embodiments, at least one modification is galactosylation. In some embodiments, the fusion protein comprises CCL1 or an active fragment thereof. In some embodiments, the CCL1 or an active fragment thereof comprises the amino acid sequence of SEQ ID NO:2 or amino acids 24-96 of SEQ ID NO:2. In some embodiments, the fusion protein comprises the amino acid sequence of SEQ ID NO:1 or amino acids 24-316 of SEQ ID NO:1. In some embodiments, the fusion protein comprises MC148 or an active fragment thereof. In some embodiments, the MC148 or an active fragment thereof comprises the amino acid sequence of SEQ ID NO: 6. In some embodiments, the fusion protein comprises the amino acid sequence of SEQ ID NO: 5.

[0019] In some embodiments, the isolated nucleic acid encodes a fusion protein. In some embodiments, a vector comprises the isolated nucleic acid. In some embodiments, a host cell comprises the nucleic acid or vector. In some embodiments, the host cell expresses the fusion protein. In some embodiments, the host cell is engineered to produce an afucosylated antibody.

[0020] In some embodiments, the method of producing a fusion protein comprises culturing a host cell under conditions suitable for expressing the fusion protein. In some embodiments, the method further comprises isolating the fusion protein.

[0021] In some embodiments, the method of treating cancer comprises administering to a subject with cancer an effective amount of an antibody provided herein (e.g., 7-B16, 1-K17, etc.). In some embodiments, the method of treating cancer comprises administering to a subject with cancer an effective amount of a fusion protein provided herein that binds to CCR8.

[0022] In some embodiments, the cancer comprises tumor-infiltrating Treg cells. In some embodiments, the cancer comprises tumor-infiltrating Treg cells. In some embodiments, the cancer comprises tumor-infiltrating Treg cells. cell sorting, FACS), gene expression analysis (Q-PCR or RT-PCR, etc.), In some embodiments, the CCR8-expressing cells include cells that express CCR8 (as determined by Western blot, ELISA, etc.). In some embodiments, the CCR8-expressing cells are Treg cells. In some embodiments, Treg cells are a subpopulation of T cells that are immunosuppressive and generally suppress or downregulate the induction and proliferation of effector T cells. In some embodiments, Treg cells express CD4, FOXP3, and CD25 (IL-2 receptor alpha chain). In some embodiments, CCR8 is expressed on the surface of Treg cells at less than 10,000 copies per cell, which can be determined, for example, by fluorescence-activated cell sorting (FACS) and flow cytometry. In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells express less than 10,000 copies of CCR8 per cell on their surface. In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells in a sample obtained from a tumor express less than 10,000 copies of CCR8 per cell. In some embodiments, the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma, hi some embodiments, the cancer is breast cancer (e.g., triple-negative breast cancer).

[0023] In some embodiments, the cancer comprises cells that express CCR8 (e.g., as determined by immunohistochemistry, fluorescence-activated cell sorting (FACS), gene expression analysis (such as Q-PCR or RT-PCR), Western blot, ELISA, etc.). In some embodiments, the CCR8-expressing cells are Treg cells. In some embodiments, Treg cells are a subpopulation of T cells that are immunosuppressive and generally suppress or downregulate the induction and proliferation of effector T cells. In some embodiments, Treg cells express CD4, FOXP3, and CD25 (IL-2 receptor alpha chain). In some embodiments, the cancer is a solid cancer. In some embodiments, the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. In some embodiments, the cancer is breast cancer (e.g., triple-negative breast cancer). In some embodiments, a method of treating a solid cancer comprises administering to a subject having a solid cancer an effective amount of an antibody that binds to human CCR8 (e.g., 7-B16, 1-K17, etc.). In some embodiments, the antibody inhibits binding of CCL1 to CCR8. In some embodiments, the cancer is a hematological cancer. In some embodiments, a method of treating a hematological cancer comprises administering to a subject having a hematological cancer an effective amount of an antibody that binds to human CCR8. In some embodiments, the antibody inhibits binding of CCL1 to CCR8. In some embodiments, the hematological cancer expresses CCR8. In some embodiments, the subject has not previously been treated with a checkpoint inhibitor (CPI). and optionally, the cancer is resistant to a CPI. In some embodiments, the cancer is a refractory cancer or is resistant to checkpoint inhibitor (CPI) therapy. In some embodiments, the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimberelimab; the anti-CTLA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is tiragolumab, vibostolimab, domvanalimab, Selected from AB308, BMS-986207, and durvalumab.

[0024] In some embodiments, the method for selecting a subject having a solid cancer for treatment with an antibody that binds to human CCR8 comprises detecting CCR8 expression in a sample from the subject. In some embodiments, the method further comprises administering an effective amount of an antibody that binds to human CCR8.

[0025] In some embodiments, the method for selecting a subject having a hematological cancer for treatment with an antibody that binds to human CCR8 comprises detecting CCR8 expression in a sample from the subject. In some embodiments, the method further comprises administering an effective amount of an antibody that binds to human CCR8.

[0026] In some embodiments, the method of treating a hematological cancer comprises administering to a subject with the hematological cancer an effective amount of a fusion protein provided herein that binds to CCR8. In some embodiments, the method of selecting a subject with a hematological cancer for treatment with a fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region comprises detecting CCR8 expression in a sample from the subject. In some embodiments, the method further comprises administering an effective amount of a fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region. In some embodiments, the hematological cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma.

[0027] In one aspect, the disclosure provides an isolated antibody that binds to human CCR8, wherein the antibody comprises an HCDR3 comprising SEQ ID NO: 86 or a variant of SEQ ID NO: 86 comprising one, two, or three mutations, wherein the antibody binds to human CCR8 and has ADCC activity. In some embodiments, the mutations are substitutions (e.g., conservative and / or non-conservative substitutions), deletions, or insertions. In some embodiments, the one, two, or three mutations are located at at least one of amino acid positions 1-4, 6, 7, or 12 of SEQ ID NO: 86. In some embodiments, the substitutions are conservative substitutions. In some embodiments, the conservative substitution is at amino acid position 1, 4, or 12 of SEQ ID NO: 86. In some embodiments, the substitution is a non-conservative substitution. In some embodiments, the non-conservative substitution is at amino acid position 7 of SEQ ID NO: 86. In some embodiments, the antibody comprises at least two substitutions in HCDR3. In some embodiments, the at least two substitutions are located at at least one of amino acid positions 1-4, 6, 7, or 12 of SEQ ID NO: 86. In some embodiments, at least two substitutions are conservative. In some embodiments, at least one of the conservative substitutions is at amino acid position 1, 4, or 12 of SEQ ID NO: 86. In some embodiments, at least two substitutions are non-conservative. In some embodiments, at least one of the non-conservative substitutions is at amino acid position 7 of SEQ ID NO: 86. In some embodiments, the mutations comprise a conservative substitution and a non-conservative substitution when more than one substitution mutation is present. In another aspect, the disclosure provides an isolated antibody that binds to human CCR8, wherein the antibody comprises an HCDR3 that is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 86, and the antibody binds to human CCR8 and has ADCC activity. In some embodiments, the HCDR3 comprises an amino acid sequence selected from SEQ ID NO: 86 and any one of SEQ ID NOs: 104-119. In some embodiments, the antibody comprises an HCDR1 comprising SEQ ID NO: 84 or SEQ ID NO: 123. In some embodiments, the antibody comprises an HCDR2 comprising SEQ ID NO:85 or SEQ ID NO:124.In some embodiments, the antibody comprises an LCDR1 comprising SEQ ID NO: 87 or SEQ ID NO: 120. In some embodiments, the antibody comprises an LCDR2 comprising SEQ ID NO: 88 or SEQ ID NO: 121. In some embodiments, the antibody comprises an LCDR3 comprising SEQ ID NO: 89 or SEQ ID NO: 122. In some embodiments, the ADCC activity is mechanism of action (MOA) based on the ADCC reporter. In some embodiments, the ADCC activity is more potent than the 7-B16 antibody. In some embodiments, the ADCC activity is at least as potent as the 7-B16 antibody. In some embodiments, the antibody has a K for human CCR8 that is equal to or less than the 7-B16 antibody (e.g., as determined by an equilibrium binding displacement assay (i.e., KinExA)). D In some embodiments, the antibody has an on-cell K for human CCR8 that is equal to or less than that of the 7-B16 antibody (e.g., as determined by a binding equilibrium exclusion assay (i.e., KinExA)). DIn some embodiments, the antibody comprises at least one modification that enhances cell killing. In some embodiments, the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). In some embodiments, the at least one modification is afucosylation. In some embodiments, the at least one modification is one or more heavy chain constant region mutations at one or more positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396. In some embodiments, the one or more heavy chain constant region mutations are one or more mutations selected from S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L. In some embodiments, the one or more heavy chain constant region mutations are selected from F243L / R292P / Y300L / V305I / P396L, S239D / I332E, S239D / I332E / A330L, S298A / E333A / K334A, L234Y / L235Q / G236W / S239M / H268D / D270E / S298A, and D270E / K326D / A330M / K334E. In some embodiments, at least one modification is galactosylation. In some embodiments, the antibody has an affinity (K) (e.g., as determined by equilibrium binding displacement assay (i.e., KinExA)) of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, then less than 250 pM, or less than 100 pM, or less than 75 pM, or less than 50 pM, or less than 25 pM. D In some embodiments, the antibody binds to human CCR8 with an on-cell affinity (K) of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75, or less than 50 pM, or less than 25 pM, as determined, for example, by an equilibrium binding exclusion assay (i.e., KinExA). D) binds to human CCR8. In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is a human antibody or a humanized antibody. In some embodiments, the antibody is a full-length antibody. In some embodiments, the antibody is an IgG1 or IgG3 antibody.

[0028] The present disclosure also provides an isolated antibody or fusion protein of any one of the aspects or embodiments described herein for treating cancer. In some embodiments, the cancer is a hematological cancer or a solid cancer / tumor. In some embodiments, the cancer expresses CCR8. In some embodiments, the hematological cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. In some embodiments, the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. In some embodiments, the cancer is breast cancer (e.g., triple-negative breast cancer). In some embodiments, the method of treatment may further include administering one or more additional therapeutic agents (e.g., other anti-cancer agents) to the subject. In some embodiments, the one or more additional therapeutic agents are an anti-Trop-2 antibody (e.g., sacituzumab govitecan, SKB-264, JS-108 (DAC-002), datopotamab deruxtecan, BAT-800). 3), anti-CD47 antibodies or CD47 blocking agents (e.g., magrolimab, DSP-107, AO-176, ALX-148, IBI-188, lemzoparlimab, TTI-621, TTI-622), anti-SIRPα antibodies (e.g., GS-0189), FLT3L-Fc fusion proteins (e.g., GS-3583), anti-PD-1 antibodies (pembrolizumab, nivolumab, dimvelelimab), small molecule PD-L1 inhibitors (e.g., GS-4224), anti-PD-L1 antibodies (e.g., atezolizumab), small molecule MCL1 inhibitors (e.g., GS-9716), small molecule HPK1 inhibitors (e.g., GS-6451), HPK1 degraders (PROTACs, e.g., ARV-766), small molecule DGKa inhibitors, small molecule CD73 inhibitors (e.g., AB680), anti-CD73 antibodies (e.g., oleclumab )), Dual A 2a / A 2b Adenosine receptor antagonists (e.g., etrumadenant (AB928)), anti-TIGIT antibodies (e.g., tiragolumab, vibostolimab, domvanalimab, AB308), anti-TREM1 antibodies (e.g., PY159), anti-TREM2 antibodies (e.g., PY314), TGFβ traps (e.g., bintrafusp alpha, AGEN-1423), anti-TGFβ1 antibodies (e.g., SRK-181), and CAR-T cell therapy (e.g., axicabtagene ciloleucel, brexucabtagene In some embodiments, the one or more additional therapeutic agents are selected from sacituzumab govitecan-hziy, magrolimab, GS-0189, GS-3583, dimverelimab, GS-4224, GS-9716, GS-6451, AB680, etormadenant (AB928), domvanalimab, AB308, PY159, PY314, SRK-181, axicabtadine ciloleucel, and brexcabutadine autoleucel. In some embodiments, the administration of the one or more additional therapeutic agents is concurrent with the administration of the antibody, while in some embodiments, the administration of the one or more additional therapeutic agents is before or after the administration of the antibody. In some embodiments, the cancer is a refractory cancer or is resistant to checkpoint inhibitor (CPI) therapy. In some embodiments, the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab; the anti-CTLA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab.

[0029] The present disclosure also provides a nucleic acid encoding an antibody according to any one of the aspects or embodiments described herein.

[0030] The present disclosure also provides a method of treating cancer, comprising administering an antibody according to any one of the aspects or embodiments described herein to a subject with cancer. In some embodiments, the cancer is a hematological cancer or a solid cancer / tumor. In some embodiments, the cancer expresses CCR8. In some embodiments, the hematological cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. In some embodiments, the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. In some embodiments, the cancer is breast cancer (e.g., triple-negative breast cancer). In some embodiments, the treatment method may further comprise administering to the subject one or more additional therapeutic agents (e.g., other anti-cancer agents). In some embodiments, the administration of the one or more additional therapeutic agents is concurrent with the administration of the antibody, while in some embodiments, the administration of the one or more additional therapeutic agents occurs before or after the administration of the antibody. In some embodiments, the one or more additional therapeutic agents are an anti-Trop-2 antibody (e.g., sacituzumab govitecan, SKB-264, JS-108 (DAC-002), datopotamab delquistecan, BAT-8003), an anti-CD47 antibody or CD47 blocker (e.g., magrolimab, DSP-107, AO-176, ALX-148, IBI-188, lemzoparlimab, TTI-621, TTI-622), an anti-SIRPα antibody (e.g., GS-0189), an FLT3L-Fc fusion protein (e.g., e.g., GS-3583), anti-PD-1 antibodies (pembrolizumab, nivolumab, dimvelelimab), small molecule PD-L1 inhibitors (e.g., GS-4224), anti-PD-L1 antibodies (e.g., atezolizumab), small molecule MCL1 inhibitors (e.g., GS-9716), small molecule HPK1 inhibitors (e.g., GS-6451), HPK1 degraders (PROTACs, e.g., ARV-766), small molecule DGKa inhibitors, small molecule CD73 inhibitors (e.g., AB680), anti-CD73 antibodies (e.g., oleculab), dual A 2a / A2b The antibody is selected from an adenosine receptor antagonist (e.g., etormadenant (AB928)), an anti-TIGIT antibody (e.g., tiragolumab, vibostolimab, domvanalimab, AB308), an anti-TREM1 antibody (e.g., PY159), an anti-TREM2 antibody (e.g., PY314), a TGFβ trap (e.g., vintrafusp alfa, AGEN-1423), an anti-TGFβ1 antibody (e.g., SRK-181), and a CAR-T cell therapy (e.g., axicabtadine ciloleucel, brexcabtadine autoleucel, tisagenlecleucel). In some embodiments, the one or more additional therapeutic agents are selected from sacituzumab govitecan-hziy, magrolimab, GS-0189, GS-3583, dimverelimab, GS-4224, GS-9716, GS-6451, AB680, etrumadenant (AB928), domvanalimab, AB308, PY159, PY314, SRK-181, axicabtadin ciloleucel, and brexcabtadin autoleucel.

[0031] The present disclosure also provides a use for treating cancer, comprising administering to a subject with cancer an antibody according to any one of the aspects or embodiments described herein. In some embodiments, the cancer is a hematological cancer or a solid cancer / tumor. In some embodiments, the cancer expresses CCR8. In some embodiments, the hematological cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. In some embodiments, the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. In some embodiments, the cancer is breast cancer (e.g., triple-negative breast cancer). In some embodiments, the treatment method may further include administering to the subject one or more additional therapeutic agents (e.g., other anti-cancer agents). In some embodiments, the administration of the one or more additional therapeutic agents is concurrent with the administration of the antibody, while in some embodiments, the administration of the one or more additional therapeutic agents occurs before or after the administration of the antibody. In some embodiments, the cancer is a refractory cancer or is resistant to checkpoint inhibitor (CPI) therapy. In some embodiments, the CPI therapy includes an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab; the anti-CLTA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab.

[0032] The present disclosure also provides a method of treating refractory or checkpoint inhibitor (CPI)-resistant cancer, comprising administering to a subject having cancer an isolated antibody according to any one of the foregoing aspects or embodiments or a fusion protein according to any one of the foregoing aspects or embodiments, wherein the subject has previously been treated with chemotherapy or CPI therapy without responding to the chemotherapy or CPI therapy. In some embodiments, the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimverelimab; the anti-CLTA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. In some embodiments, the cancer is a hematological cancer or a solid cancer. In some embodiments, the cancer expresses CCR8. In some embodiments, the cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. In some embodiments, the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. In some embodiments, the cancer is breast cancer (e.g., triple-negative breast cancer). In some embodiments, the method may further comprise administering one or more additional therapeutic agents to the subject.In some embodiments, the one or more additional therapeutic agents are an anti-Trop-2 antibody (e.g., sacituzumab govitecan, SKB-264, JS-108 (DAC-002), datopotamab delquistecan, BAT-8003), an anti-CD47 antibody or CD47 blocker (e.g., magrolimab, DSP-107, AO-176, ALX-148, IBI-188, lemzoparlimab, TTI-621, TTI-622), an anti-SIRPα antibody (e.g., GS-0189), an FLT3L-Fc fusion protein (e.g., e.g., GS-3583), anti-PD-1 antibodies (pembrolizumab, nivolumab, dimvelelimab), small molecule PD-L1 inhibitors (e.g., GS-4224), anti-PD-L1 antibodies (e.g., atezolizumab), small molecule MCL1 inhibitors (e.g., GS-9716), small molecule HPK1 inhibitors (e.g., GS-6451), HPK1 degraders (PROTACs, e.g., ARV-766), small molecule DGKa inhibitors, small molecule CD73 inhibitors (e.g., AB680), anti-CD73 antibodies (e.g., oleculab), and dual A. 2a / A 2b The antibody is selected from an adenosine receptor antagonist (e.g., etormadenant (AB928)), an anti-TIGIT antibody (e.g., tiragolumab, vibostolimab, domvanalimab, AB308), an anti-TREM1 antibody (e.g., PY159), an anti-TREM2 antibody (e.g., PY314), a TGFβ trap (e.g., vintrafusp alfa, AGEN-1423), an anti-TGFβ1 antibody (e.g., SRK-181), and a CAR-T cell therapy (e.g., axicabtadine ciloleucel, brexcabtadine autoleucel, tisagenlecleucel). In some embodiments, the one or more additional therapeutic agents are selected from sacituzumab govitecan-hziy, magrolimab, GS-0189, GS-3583, dimverelimab, GS-4224, GS-9716, GS-6451, AB680, etormadenant (AB928), domvanalimab, AB308, PY159, PY314, SRK-181, axicabtadine ciloleucel, and brexcabutadine autoleucel. In some embodiments, the administration of the one or more additional therapeutic agents is concurrent with the administration of the antibody. In some embodiments, the administration of the one or more additional therapeutic agents is before or after the administration of the antibody. The present invention provides, for example, the following items. (Item 1) 1. An isolated antibody that binds to human CCR8, said antibody comprising: i) HCDR1 comprising the amino acid sequence of SEQ ID NO: 12, HCDR2 comprising the amino acid sequence of SEQ ID NO: 13, HCDR3 comprising the amino acid sequence of SEQ ID NO: 14, LCDR1 comprising the amino acid sequence of SEQ ID NO: 15, LCDR2 comprising the amino acid sequence of SEQ ID NO: 16, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 17; ii) HCDR1 comprising the amino acid sequence of SEQ ID NO: 24, HCDR2 comprising the amino acid sequence of SEQ ID NO: 25, HCDR3 comprising the amino acid sequence of SEQ ID NO: 26, LCDR1 comprising the amino acid sequence of SEQ ID NO: 27, LCDR2 comprising the amino acid sequence of SEQ ID NO: 28, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 29; iii) HCDR1 comprising the amino acid sequence of SEQ ID NO: 36, HCDR2 comprising the amino acid sequence of SEQ ID NO: 37, HCDR3 comprising the amino acid sequence of SEQ ID NO: 38, LCDR1 comprising the amino acid sequence of SEQ ID NO: 39, LCDR2 comprising the amino acid sequence of SEQ ID NO: 40, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 41; iv) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 48, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 49, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 50, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 51, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 52, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 53, or v) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65, or vi) An isolated antibody comprising an HCDR1 having the amino acid sequence of SEQ ID NO: 84 or 100, an HCDR2 having the amino acid sequence of SEQ ID NO: 85, an HCDR3 having the amino acid sequence of SEQ ID NO: 86, an LCDR1 having the amino acid sequence of SEQ ID NO: 87, an LCDR2 having the amino acid sequence of SEQ ID NO: 88, and an LCDR3 having the amino acid sequence of SEQ ID NO: 89. (Item 2) The antibody i) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65, or ii) An isolated antibody according to item 1, comprising an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85, an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86, an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89. (Item 3) The antibody i) a heavy chain variable region (VH) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 68 or 74, and a light chain variable region (VL) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 69 or 75; or ii) the isolated antibody of item 2, comprising a heavy chain variable region (VH) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 92 or 96, and a light chain variable region (VL) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 93 or 97. (Item 4) The antibody i) a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 68 or 74, and a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO: 69 or 75; or ii) The isolated antibody of item 2 or 3, comprising a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 92 or 96, and a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO: 93 or 97. (Item 5) 5. The isolated antibody of any one of items 1 to 4, wherein the antibody is a monoclonal antibody. (Item 6) Item 7. The isolated antibody according to any one of Items 1 to 5, wherein the antibody is a humanized antibody. Item 8. The isolated antibody according to any one of Items 1 to 6, wherein the antibody is a full-length antibody. 8. The isolated antibody according to any one of items 1 to 7, wherein the antibody is an IgG1 or IgG3 antibody. (Item 9) The antibody i) a heavy chain (HC) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 70 or 76, and a light chain (LC) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 71 or 77; or ii) the isolated antibody of any one of items 1 to 8, comprising a heavy chain (HC) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 94 or 98, and a light chain (LC) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 95 or 99. (Item 10) The antibody i) a heavy chain (HC) comprising the amino acid sequence of SEQ ID NO: 70 or 76, and a light chain (LC) comprising the amino acid sequence of SEQ ID NO: 71 or 77, or ii) The isolated antibody according to any one of items 1 to 9, comprising a heavy chain (HC) comprising the amino acid sequence of SEQ ID NO: 94 or 98, and a light chain (LC) comprising the amino acid sequence of SEQ ID NO: 95 or 99. (Item 11) 11. The isolated antibody of any one of paragraphs 1 to 10, wherein the antibody comprises at least one modification that enhances cell killing. (Item 12) 12. The isolated antibody of item 11, wherein the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). (Item 13) 13. The isolated antibody of item 11 or 12, wherein the at least one modification is afucosylation. (Item 14) 13. The isolated antibody of paragraph 11 or 12, wherein the at least one modification is one or more heavy chain constant region mutations at one or more positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396. (Item 15) 15. The isolated antibody of item 14, wherein the one or more heavy chain constant region mutations are one or more mutations selected from S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L. (Item 16) 15. The isolated antibody of claim 14, wherein the one or more heavy chain constant region mutations are selected from F243L / R292P / Y300L / V305I / P396L, S239D / I332E, S239D / I332E / A330L, S298A / E333A / K334A, L234Y / L235Q / G236W / S239M / H268D / D270E / S298A, and D270E / K326D / A330M / K334E. (Item 17) 13. The isolated antibody of item 11 or 12, wherein the at least one modification is galactosylation. (Item 18) The antibody has an affinity (K) of less than 10 nM, or less than 5 nM, or less than 1 nM, then less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75 pM, or less than 50 pM, or less than 25 pM. D 18. The isolated antibody of any one of items 1 to 17, which binds to human CCR8 at the CCR8 domain. (Item 19) The antibody has an on-cell affinity (K) of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75, or less than 50 pM, or less than 25 pM. D 19. The isolated antibody of any one of items 1 to 18, which binds to human CCR8 at the CCR8 domain. (Item 20) The affinity (K D 20. The isolated antibody of item 18 or 19, wherein the binding affinity of the antibody to the antibody is determined by an equilibrium binding exclusion assay (i.e., KinExA). (Item 21) An isolated nucleic acid encoding the antibody of any one of items 1 to 20. (Item 22) A vector comprising the isolated nucleic acid of Item 21. (Item 23) 23. A host cell comprising the nucleic acid of Item 21 or the vector of Item 22. (Item 24) A host cell expressing the antibody according to any one of items 1 to 20. (Item 25) Item 26. The host cell according to Item 23 or 24, which is engineered to produce an afucosylated antibody. 26. A method for producing an antibody that binds to CCR8, the method comprising culturing the host cell of any one of items 23 to 25 under conditions suitable for expressing the antibody. (Item 27) 27. The method of claim 26, further comprising isolating the antibody. (Item 28) A fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region. (Item 29) 29. The fusion protein of item 28, wherein the Fc region is an IgG1 or IgG3 Fc region. (Item 30) 30. The fusion protein of item 28 or 29, wherein the Fc region comprises the amino acid sequence of SEQ ID NO: 4. (Item 31) 31. The fusion protein of any one of items 28 to 30, wherein the Fc region comprises at least one modification that enhances cell killing. (Item 32) 32. The fusion protein of item 31, wherein the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). (Item 33) 33. The fusion protein according to item 31 or 32, wherein the at least one modification is afucosylation. (Item 34) 33. The fusion protein of item 31 or 32, wherein the at least one modification is one or more Fc region mutations at one or more positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396. (Item 35) 35. The fusion protein of item 34, wherein the one or more Fc region mutations are one or more mutations selected from S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L. (Item 36) 35. The fusion protein of item 34, wherein the one or more Fc region mutations are selected from F243L / R292P / Y300L / V305I / P396L, S239D / I332E, S239D / I332E / A330L, S298A / E333A / K334A, L234Y / L235Q / G236W / S239M / H268D / D270E / S298A, and D270E / K326D / A330M / K334E. (Item 37) 33. The fusion protein according to item 31 or 32, wherein the at least one modification is galactosylation. (Item 38) 38. The fusion protein according to any one of items 28 to 37, wherein the fusion protein comprises CCL1 or an active fragment thereof. (Item 39) 39. The fusion protein according to item 38, wherein the CCL1 or an active fragment thereof comprises the amino acid sequence of SEQ ID NO: 2 or amino acids 24 to 96 of SEQ ID NO: 2. (Item 40) 40. The fusion protein according to item 38 or 39, wherein the fusion protein comprises the amino acid sequence of SEQ ID NO: 1 or amino acids 24 to 316 of SEQ ID NO: 1. (Item 41) 38. The fusion protein according to any one of items 28 to 37, wherein the fusion protein comprises MC148 or an active fragment thereof. (Item 42) 42. The fusion protein of item 41, wherein the MC148 or an active fragment thereof comprises the amino acid sequence of SEQ ID NO: 6. (Item 43) 43. The fusion protein according to item 41 or 42, wherein the fusion protein comprises the amino acid sequence of SEQ ID NO: 5. (Item 44) 44. An isolated nucleic acid encoding the fusion protein according to any one of Items 28 to 43. (Item 45) A vector comprising the isolated nucleic acid of item 44. (Item 46) 46. ​​A host cell comprising the nucleic acid according to item 44 or the vector according to item 45. (Item 47) A host cell that expresses the fusion protein according to any one of Items 28 to 43. (Item 48) Item 49. The host cell according to Item 46 or 47, which is engineered to produce an afucosylated antibody. 49. A method for producing a fusion protein, comprising culturing a host cell according to any one of items 46 to 48 under conditions suitable for expressing said fusion protein. (Item 50) 50. The method of claim 49, further comprising isolating the fusion protein. (Item 51) A method for treating cancer, comprising administering to a subject having cancer an effective amount of the antibody according to any one of items 1 to 20. (Item 52) A method for treating cancer, comprising administering to a subject having cancer an effective amount of the fusion protein according to any one of items 28 to 43. (Item 53) 53. The method of paragraph 51 or 52, wherein the subject has previously been treated with a checkpoint inhibitor (CPI), and optionally the cancer was resistant to the CPI. (Item 54) 54. The method of any one of items 51 to 53, wherein the cancer comprises tumor-infiltrating Treg cells. (Item 55) 54. The method of any one of items 51 to 53, wherein the cancer comprises cells that express CCR8. (Item 56) 56. The method of item 55, wherein the cells expressing CCR8 are Treg cells. (Item 57) 57. The method of item 56, wherein CCR8 is expressed on the surface of the Treg cells at less than 10,000 copies per cell (as determined by fluorescence-activated cell sorting (FACS) and / or flow cytometry). (Item 58) 58. The method of any one of items 51 to 57, wherein the cancer comprises tumor cells that express CCR8. (Item 59) 59. The method according to any one of items 51 to 58, wherein the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. (Item 60) 60. The method according to any one of items 51 to 59, wherein the cancer is a refractory cancer or is resistant to checkpoint inhibitor (CPI) therapy. (Item 61) 61. The method of claim 60, wherein the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. (Item 62) 62. The method of item 61, wherein the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab, the anti-CLTA4 antibody is ipilimumab or tremelimumab, or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. (Item 63) A method for treating blood cancer, comprising administering to a subject having the blood cancer an effective amount of an antibody that binds to human CCR8. (Item 64) 64. The method of claim 63, wherein the subject has previously been treated with a checkpoint inhibitor (CPI), and optionally the cancer was resistant to the CPI. (Item 65) 65. The method of item 63 or 64, wherein the antibody inhibits binding of CCL1 to CCR8. (Item 66) The method according to any one of Items 63 to 65, wherein the blood cancer expresses CCR8. (Item 67) 67. The method according to any one of items 63 to 66, wherein the blood cancer is a refractory cancer or is resistant to checkpoint inhibitor (CPI) therapy. (Item 68) 68. The method of item 67, wherein the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. (Item 69) 69. The method of item 68, wherein the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab, the anti-CLTA4 antibody is ipilimumab or tremelimumab, or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. (Item 70) 1. A method for selecting a subject having a blood cancer for treatment with an antibody that binds to human CCR8, the method comprising detecting CCR8 expression in a sample from the subject. (Item 71) 71. The method of item 70, further comprising administering an effective amount of an antibody that binds to human CCR8. (Item 72) 72. The method according to any one of items 63 to 71, wherein the antibody that binds to human CCR8 is the antibody according to any one of items 1 to 20. (Item 73) A method for treating blood cancer, comprising administering to a subject having blood cancer an effective amount of the fusion protein according to any one of items 28 to 43. (Item 74) 1. A method for selecting a subject having a blood cancer for treatment with a fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region, the method comprising detecting CCR8 expression in a sample from the subject. (Item 75) 75. The method of item 74, further comprising administering an effective amount of a fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region. (Item 76) Item 76. The method according to Item 75, wherein the fusion protein is the fusion protein according to any one of Items 28 to 43. (Item 77) 77. The method according to any one of Items 73 to 76, wherein the blood cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. (Item 78) An isolated antibody that binds to human CCR8, wherein the antibody comprises an HCDR3 comprising SEQ ID NO: 86 or a variant of SEQ ID NO: 86 containing one, two, or three mutations, and wherein the antibody binds to human CCR8 and has ADCC activity. (Item 79) 79. The isolated antibody of item 78, wherein the mutation is selected from a substitution, an insertion, or a deletion. (Item 80) 80. The isolated antibody of item 78 or 79, wherein the antibody comprises at least two substitutions in HCDR3. (Item 81) 81. The isolated antibody of any one of items 78 to 80, wherein the one, two, or three mutations are located at at least one of amino acid positions 1 to 4, 6, 7, or 12 of SEQ ID NO: 86. (Item 82) 82. The isolated antibody of any one of items 79 to 81, wherein the substitution is a conservative substitution. (Item 83) 83. The isolated antibody of item 82, wherein the conservative substitution is at amino acid position 1, 4, or 12 of SEQ ID NO: 86. (Item 84) 82. The isolated antibody of any one of items 79 to 81, wherein the substitution is a non-conservative substitution. (Item 85) 85. The isolated antibody of item 84, wherein the non-conservative substitution is at amino acid position 7 of SEQ ID NO: 86. (Item 86) 82. The isolated antibody of any one of items 79 to 81, wherein the mutations comprise conservative and non-conservative substitutions when more than one substitution mutation is present. (Item 87) An isolated antibody that binds to human CCR8, wherein the antibody comprises an HCDR3 that is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 86, and the antibody binds to human CCR8 and has ADCC activity. (Item 88) 88. The isolated antibody of item 78 or 87, wherein the HCDR3 comprises an amino acid sequence selected from any one of SEQ ID NOs: 104 to 119. (Item 89) 88. The isolated antibody according to any one of Items 78 to 87, wherein the HCDR3 comprises an amino acid sequence selected from SEQ ID NO: 86 and any one of SEQ ID NOs: 104 to 119. (Item 90) 90. The isolated antibody of any one of items 78 to 89, wherein the antibody comprises an HCDR1 comprising SEQ ID NO: 84 or SEQ ID NO: 123. (Item 91) 91. The isolated antibody of any one of items 78 to 90, wherein the antibody comprises an HCDR2 comprising SEQ ID NO: 85 or SEQ ID NO: 124. (Item 92) 92. The isolated antibody of any one of items 78 to 91, wherein the antibody comprises an LCDR1 comprising SEQ ID NO: 87 or SEQ ID NO: 120. (Item 93) 93. The isolated antibody of any one of items 78 to 92, wherein the antibody comprises an LCDR2 comprising SEQ ID NO: 88 or SEQ ID NO: 121. (Item 94) 94. The isolated antibody of any one of items 78 to 93, wherein the antibody comprises an LCDR3 comprising SEQ ID NO: 89 or SEQ ID NO: 122. (Item 95) 95. The isolated antibody of any one of Aspects 78 to 94, wherein the antibody comprises a heavy chain variable region (VH) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of any one of SEQ ID NOs: 80, 92, and 96, and a light chain variable region (VL) comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of any one of SEQ ID NOs: 81, 93, and 97. (Item 96) 96. The isolated antibody of any one of paragraphs 78 to 95, wherein the antibody comprises a heavy chain that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of any one of SEQ ID NOs: 82, 90, 94, and 98, and a light chain comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of any one of SEQ ID NOs: 83, 91, 95, and 99. (Item 97) 97. The isolated antibody of any one of paragraphs 78 to 96, wherein the ADCC activity comprises an EC50 value of less than 200, 175, 150, 125, 100, 75, 50, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 ng / ml as measured by an ADCC reporter mechanism of action (MOA)-based bioassay. (Item 98) 98. The isolated antibody according to any one of items 78 to 97, wherein the ADCC activity is stronger than that of 7-B16 antibody. (Item 99) The antibody has a K activity against human CCR8 that is equal to or less than 7-B16 antibody. D 99. The isolated antibody of any one of items 78 to 98, having the formula: (Item 100) The antibody is the 7-B16 antibody or less. D 90. The isolated antibody according to any one of items 78 to 99, comprising: (Item 101) The above K D 101. The isolated antibody of item 99 or 100, wherein the binding affinity of the antibody is determined by equilibrium binding exclusion assay (i.e., KinExA). (Item 102) 102. The isolated antibody of any one of paragraphs 78 to 101, wherein the antibody comprises at least one modification that enhances cell killing. (Item 103) 103. The isolated antibody of paragraph 102, wherein the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). (Item 104) 104. The isolated antibody of item 102 or 103, wherein the at least one modification is afucosylation. (Item 105) 104. The isolated antibody of paragraph 102 or 103, wherein the at least one modification is one or more heavy chain constant region mutations at one or more positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396. (Item 106) 106. The isolated antibody of claim 105, wherein the one or more heavy chain constant region mutations are one or more mutations selected from S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L. (Item 107) 106. The isolated antibody of claim 105, wherein the one or more heavy chain constant region mutations are selected from F243L / R292P / Y300L / V305I / P396L, S239D / I332E, S239D / I332E / A330L, S298A / E333A / K334A, L234Y / L235Q / G236W / S239M / H268D / D270E / S298A, and D270E / K326D / A330M / K334E. (Item 108) 98. The isolated antibody of item 96 or 97, wherein the at least one modification is galactosylation. (Item 109) The antibody has an affinity (K) of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75 pM, or less than 50 pM, or less than 25 pM. D 109. The isolated antibody of any one of items 78 to 108, which binds to human CCR8 at the CCR8 domain. (Item 110) The antibody has an on-cell affinity (K) of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75, or less than 50 pM, or less than 25 pM. D 101. The isolated antibody of any one of items 78 to 109, which binds to human CCR8 at the CCR8 domain. (Item 111) The affinity (K D 111. The isolated antibody of item 109 or 110, wherein the binding affinity of the antibody to the antibody is determined by equilibrium binding exclusion assay (i.e., KinExA). (Item 112) 112. The isolated antibody of any one of items 78 to 111, wherein the antibody is a monoclonal antibody. (Item 113) 113. The isolated antibody of any one of items 78 to 112, wherein the antibody is a human antibody or a humanized antibody. (Item 114) 114. The isolated antibody of any one of items 78 to 113, wherein the antibody is a full-length antibody. (Item 115) 115. The isolated antibody of any one of items 78 to 114, wherein the antibody is an IgG1 or IgG3 antibody. (Item 116) A nucleic acid encoding the antibody according to any one of Items 78 to 115. (Item 117) A method for treating cancer, comprising administering to a subject having cancer the antibody of any one of Items 78 to 115. (Item 118) Item 118. The method of item 117, wherein the cancer is a blood cancer or a solid cancer. (Item 119) 119. The method of item 117 or 118, wherein the cancer expresses CCR8. (Item 120) 119. The method of any one of Items 117 to 119, wherein the cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. (Item 121) 119. The method according to any one of items 117 to 119, wherein the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. (Item 122) 122. The method of any one of items 117 to 121, further comprising administering to the subject one or more additional therapeutic agents. (Item 123) The one or more additional therapeutic agents may be an anti-Trop-2 antibody (e.g., sacituzumab govitecan, SKB-264, JS-108 (DAC-002), datopotamab delquistecan, BAT-8003), an anti-CD47 antibody or CD47 blocker (e.g., magrolimab, DSP-107, AO-176, ALX-148, IBI-188, lemzoparlimab, TTI-621, TTI-622), an anti-SIRPα antibody (e.g., GS-0189), an FLT3L-Fc fusion protein (e.g., GS- 3583), anti-PD-1 antibodies (pembrolizumab, nivolumab, dimvelelimab), small molecule PD-L1 inhibitors (e.g., GS-4224), anti-PD-L1 antibodies (e.g., atezolizumab), small molecule MCL1 inhibitors (e.g., GS-9716), small molecule HPK1 inhibitors (e.g., GS-6451), HPK1 degraders (PROTACs, e.g., ARV-766), small molecule DGKa inhibitors, small molecule CD73 inhibitors (e.g., AB680), anti-CD73 antibodies (e.g., oleculab), dual A 2a / A 2b 123. The method of claim 122, wherein the therapeutic agent is selected from an adenosine receptor antagonist (e.g., etormadenant (AB928)), an anti-TIGIT antibody (e.g., tiragolumab, vibostolimab, domvanalimab, AB308), an anti-TREM1 antibody (e.g., PY159), an anti-TREM2 antibody (e.g., PY314), a TGFβ trap (e.g., vintrafusp alfa, AGEN-1423), an anti-TGFβ1 antibody (e.g., SRK-181), and a CAR-T cell therapy (e.g., axicabtadine ciloleucel, brexcabtadine autoleucel, tisagenlecleucel). (Item 124) 123. The method of claim 122, wherein the one or more additional therapeutic agents are selected from sacituzumab govitecan-hziy, magrolimab, GS-0189, GS-3583, dimverelimab, GS-4224, GS-9716, GS-6451, AB680, etormadenant (AB928), domvanalimab, AB308, PY159, PY314, SRK-181, axicabtadin ciloleucel, and brexcabtadin autoleucel. (Item 125) 125. The method of any one of items 122 to 124, wherein the administration of the one or more additional therapeutic agents is concurrent with the administration of the antibody. (Item 126) 125. The method of any one of items 122 to 124, wherein the administration of the one or more additional therapeutic agents precedes or follows the administration of the antibody. (Item 127) 127. The method of any one of items 117 to 126, wherein the cancer is a refractory cancer or is resistant to checkpoint inhibitor (CPI) therapy. (Item 128) 128. The method of claim 127, wherein the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. (Item 129) Item 129. The method of item 128, wherein the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab, the anti-CLTA4 antibody is ipilimumab or tremelimumab, or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. (Item 130) 11. Use of the isolated antibody of any one of items 1 to 20 or 78 to 115 or the fusion protein of any one of items 28 to 43 for treating cancer. (Item 131) 131. The use according to item 130, wherein the cancer is a blood cancer or a solid tumor. (Item 132) 132. The use according to item 130 or 131, wherein the cancer expresses CCR8. (Item 133) 133. The use according to any one of items 130 to 132, wherein the cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. (Item 134) 133. The use according to any one of items 130 to 132, wherein the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. (Item 135) 135. The use of any one of items 130 to 134, further comprising administering to the subject one or more additional therapeutic agents. (Item 136) 136. The use of item 135, wherein the administration of the one or more additional therapeutic agents is concurrent with the administration of the antibody. (Item 137) 136. The use of item 135, wherein the administration of the one or more additional therapeutic agents occurs before or after the administration of the antibody. (Item 138) 138. The use according to any one of items 130 to 137, wherein the cancer is a refractory cancer or is resistant to checkpoint inhibitor (CPI) therapy. (Item 139) 139. The use of item 138, wherein the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. (Item 140) 139. The use of item 139, wherein the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab, the anti-CLTA4 antibody is ipilimumab or tremelimumab, or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. (Item 141) 11. Use of the isolated antibody of any one of items 1 to 20 or 78 to 115 or the fusion protein of any one of items 28 to 43 for treating cancer. (Item 142) 142. The isolated antibody or fusion protein of claim 141, wherein the cancer is a hematological cancer or a solid tumor. (Item 143) 143. The isolated antibody or fusion protein of paragraph 141 or 142, wherein the cancer expresses CCR8. (Item 144) 144. The isolated antibody or fusion protein of any one of paragraphs 141 to 143, wherein the cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. (Item 145) 144. The isolated antibody or fusion protein of any one of items 141 to 143, wherein the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. (Item 146) 146. The isolated antibody or fusion protein of any one of paragraphs 141 to 145, further comprising administering to the subject one or more additional therapeutic agents. (Item 147) 147. The isolated antibody or fusion protein of claim 146, wherein the administration of the one or more additional therapeutic agents is concurrent with the administration of the antibody. (Item 148) 147. The isolated antibody or fusion protein of claim 146, wherein administration of the one or more additional therapeutic agents occurs before or after administration of the antibody. (Item 149) 149. The isolated antibody or fusion protein of any one of paragraphs 141 to 148, wherein the cancer is a refractory cancer or is resistant to checkpoint inhibitor (CPI) therapy. (Item 150) 10. A method of treating refractory or checkpoint inhibitor (CPI) resistant cancer, comprising administering to a subject having cancer the isolated antibody of any one of items 1 to 20 or 78 to 115 or the fusion protein of any one of items 28 to 43, wherein the subject has been previously treated with chemotherapy or CPI therapy without responding to said chemotherapy or CPI therapy. (Item 151) 151. The method of claim 150, wherein the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. (Item 152) Item 152. The method of item 151, wherein the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab, the anti-CLTA4 antibody is ipilimumab or tremelimumab, or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. (Item 153) 153. The method according to any one of items 150 to 152, wherein the cancer is a blood cancer or a solid cancer. (Item 154) The method of any one of Items 150 to 153, wherein the cancer expresses CCR8. 155. The method of any one of items 150 to 154, wherein the cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma. (Item 156) 155. The method according to any one of items 150 to 154, wherein the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. (Item 157) 157. The method of any one of items 150 to 156, further comprising administering to the subject one or more additional therapeutic agents. (Item 158) The one or more additional therapeutic agents may be an anti-Trop-2 antibody (e.g., sacituzumab govitecan, SKB-264, JS-108 (DAC-002), datopotamab delquistecan, BAT-8003), an anti-CD47 antibody or CD47 blocker (e.g., magrolimab, DSP-107, AO-176, ALX-148, IBI-188, lemzoparlimab, TTI-621, TTI-622), an anti-SIRPα antibody (e.g., GS-0189), an FLT3L-Fc fusion protein (e.g., GS- 3583), anti-PD-1 antibodies (pembrolizumab, nivolumab, dimvelelimab), small molecule PD-L1 inhibitors (e.g., GS-4224), anti-PD-L1 antibodies (e.g., atezolizumab), small molecule MCL1 inhibitors (e.g., GS-9716), small molecule HPK1 inhibitors (e.g., GS-6451), HPK1 degraders (PROTACs, e.g., ARV-766), small molecule DGKa inhibitors, small molecule CD73 inhibitors (e.g., AB680), anti-CD73 antibodies (e.g., oleculab), dual A 2a / A 2b 158. The method of claim 157, wherein the therapeutic agent is selected from an adenosine receptor antagonist (e.g., etormadenant (AB928)), an anti-TIGIT antibody (e.g., tiragolumab, vibostolimab, domvanalimab, AB308), an anti-TREM1 antibody (e.g., PY159), an anti-TREM2 antibody (e.g., PY314), a TGFβ trap (e.g., vintrafusp alfa, AGEN-1423), an anti-TGFβ1 antibody (e.g., SRK-181), and a CAR-T cell therapy (e.g., axicabtadine ciloleucel, brexcabtadine autoleucel, tisagenlecleucel). (Item 159) 158. The method of claim 157, wherein the one or more additional therapeutic agents are selected from sacituzumab govitecan-hziy, magrolimab, GS-0189, GS-3583, dimverelimab, GS-4224, GS-9716, GS-6451, AB680, etormadenant (AB928), domvanalimab, AB308, PY159, PY314, SRK-181, axicabtadin ciloleucel, and brexcabtadin autoleucel. (Item 160) 159. The method of any one of items 157 to 159, wherein the administration of the one or more additional therapeutic agents is concurrent with the administration of the antibody. (Item 161) 159. The method of any one of items 157 to 159, wherein the administration of the one or more additional therapeutic agents precedes or follows the administration of the antibody. [Brief explanation of the drawings]

[0033] [Figure 1] Scatter plots of CCR8 mRNA expression levels determined by microarray are shown. On the y-axis is CCR8 gene expression measured by fluorescence intensity (AU). The x-axis represents leukemia subtypes or healthy bone marrow. For each group on the x-axis, the line represents the median CCR8 expression level for that leukemia subtype. Leukemia subtypes indicated include acute lymphocytic leukemia (ALL), T-cell acute lymphocytic leukemia (TALL), B-cell acute lymphocytic leukemia (BALL), acute myeloid leukemia (AML), and chronic lymphocytic leukemia (chronic lymphocytic leukemia). lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), and myelodysplastic syndromes (MDS).

[0034] [Figure 2] Box plots showing CCR8 mRNA expression levels determined by RNA sequencing are shown. On the y-axis is CCR8 gene expression measured by log-transformed transcripts per million (TPM). On the x-axis is lymphoma subtype, leukemia subtype, or healthy blood or tissue.

[0035] [Figure 3A]Human tumor-infiltrating (TIL) Tregs preferentially express CCR8, but at low receptor numbers on a per-cell basis. An exemplary flow cytometry plot (gated on tumor T CD4+ cells) (A) and quantitative plot (B) are shown. [Figure 3B] Human tumor-infiltrating (TIL) Tregs preferentially express CCR8, but at low receptor numbers on a per-cell basis. An exemplary flow cytometry plot (gated on tumor T CD4+ cells) (A) and quantitative plot (B) are shown.

[0036] [Figure 4A] Binding of anti-CCR8 monoclonal antibodies to human CCR8-expressing CHO-S cells (A) and CCR4-expressing CHO-S cells (B) is shown. [Figure 4B] Binding of anti-CCR8 monoclonal antibodies to human CCR8-expressing CHO-S cells (A) and CCR4-expressing CHO-S cells (B) is shown.

[0037] [Figure 5] 1 shows the antagonist activity of anti-CCR8 chimeric antibodies using the DiscoverX assay (Eurofins).

[0038] [Figure 6] Figure 1 shows antibody-dependent cellular cytotoxicity (ADCC) of certain anti-CCR8 chimeric antibodies using an ADCC reporter bioassay (Promega).

[0039] [Figure 7A] Figure 1 shows that afucosylation of a human IgG1 anti-CCR8 antibody is required for killing of target cells expressing low levels of CCR8. Fold change in ADCC reporter activity of titrated human IgG1 (A) and afucosylated human IgG1 (B) versions of the anti-human CCR8 clone 1K17. CHO target cells expressed 300,000 (filled circles) or 10,000 (open circles) cell surface human CCR8 molecules per cell. [Figure 7B] Figure 1 shows that afucosylation of a human IgG1 anti-CCR8 antibody is required for killing of target cells expressing low levels of CCR8. Fold change in ADCC reporter activity of titrated human IgG1 (A) and afucosylated human IgG1 (B) versions of the anti-human CCR8 clone 1K17. CHO target cells expressed 300,000 (filled circles) or 10,000 (open circles) cell surface human CCR8 molecules per cell.

[0040] [Figure 8A] These results demonstrate that the Fc effector function of anti-mouse CCR8 mAb is required for efficient killing of tumor-infiltrating (TIL) Tregs in vivo. The Fc-competent (mouse IgG2a, open circles) anti-mouse CCR8 mIgG2a antibody was able to deplete TIL Tregs in vivo in the murine MC38 syngeneic model, whereas the Fc-competent (mouse IgG1, filled circles) anti-mouse CCR8 mIgG1 antibody was not. An exemplary flow cytometry plot (gated on tumor T CD4+ cells) (A) and quantitative plots (B and C) show the frequency of TIL Tregs 3 days after injection of 200 μg of the indicated antibody (n=4). [Figure 8B] These results demonstrate that the Fc effector function of anti-mouse CCR8 mAb is required for efficient killing of tumor-infiltrating (TIL) Tregs in vivo. The Fc-competent (mouse IgG2a, open circles) anti-mouse CCR8 mIgG2a antibody was able to deplete TIL Tregs in vivo in the murine MC38 syngeneic model, whereas the Fc-competent (mouse IgG1, filled circles) anti-mouse CCR8 mIgG1 antibody was not. An exemplary flow cytometry plot (gated on tumor T CD4+ cells) (A) and quantitative plots (B and C) show the frequency of TIL Tregs 3 days after injection of 200 μg of the indicated antibody (n=4). [Figure 8C]These results demonstrate that the Fc effector function of anti-mouse CCR8 mAb is required for efficient killing of tumor-infiltrating (TIL) Tregs in vivo. The Fc-competent (mouse IgG2a, open circles) anti-mouse CCR8 mIgG2a antibody was able to deplete TIL Tregs in vivo in the murine MC38 syngeneic model, whereas the Fc-competent (mouse IgG1, filled circles) anti-mouse CCR8 mIgG1 antibody was not. An exemplary flow cytometry plot (gated on tumor T CD4+ cells) (A) and quantitative plots (B and C) show the frequency of TIL Tregs 3 days after injection of 200 μg of the indicated antibody (n=4).

[0041] [Figure 9A] Figure 1 shows that Fc-competent (mouse IgG2a, open circles) anti-mouse CCR8 mIgG2a antibody reduced tumor growth (A) and increased survival (B) in the MC38 syngeneic mouse model, whereas Fc-competent (mouse IgG1, closed circles) anti-mouse CCR8 mIgG1 antibody did not. Tumor growth curves (A) and survival curves (B) of MC38 mice after therapeutic treatment with isotype control, Fc-competent (mouse IgG2a), or Fc-incompetent (mouse IgG1) anti-mouse CCR8 antibodies (n = 10-15) are shown. [Figure 9B] Figure 1 shows that Fc-competent (mouse IgG2a, open circles) anti-mouse CCR8 mIgG2a antibody reduced tumor growth (A) and increased survival (B) in the MC38 syngeneic mouse model, whereas Fc-competent (mouse IgG1, closed circles) anti-mouse CCR8 mIgG1 antibody did not. Tumor growth curves (A) and survival curves (B) of MC38 mice after therapeutic treatment with isotype control, Fc-competent (mouse IgG2a), or Fc-incompetent (mouse IgG1) anti-mouse CCR8 antibodies (n = 10-15) are shown.

[0042] [Figure 10]Figure 1 shows CCR8 mRNA expression in various hematological malignancies compared to normal healthy blood. * indicates cancer types with FDR-adjusted p-values ​​less than 0.05. N indicates the number of samples in each cancer type. Cancers indicated include acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), diffuse large B cell lymphoma (DLBCL), chronic myeloid leukemia (CML), mantle cell lymphoma (MCL), and angioimmunoblastic T cell lymphoma (MCL). lymphoma, ATCL), hepatosplenic T cell lymphoma (HTCL), peripheral T cell lymphoma not otherwise specified (PTCL NOS), Burkitt lymphoma (BL), adult T cell leukemia / lymphoma (ATLL), anaplastic large cell lymphoma lymphoma, ALCL), chronic myelomonocytic leukemia (chronic These include myelomonocytic leukemia (CMML), follicular lymphoma (FL), T cell lymphoblastic leukemia / lymphoma (TLLL), extranodal NK / T cell lymphoma (NKTCL), primary effusion lymphoma (PEL), acute lymphocytic leukemia / acute myeloid leukemia (ALL, AML), histiocytic lymphoma (HL), cutaneous T cell lymphoma (CTCL), and marginal zone lymphoma (MZL).

[0043] [Figure 11] 1 shows antibody-dependent cellular cytotoxicity (ADCC) of CDR H3 variants of anti-CCR8 chimeric antibody 7-B16 using an ADCC reporter bioassay (Promega). DETAILED DESCRIPTION OF THE INVENTION

[0044] Antibodies that bind to CCR8 are provided. Antibody heavy and light chains capable of forming antibodies that bind to CCR8 are also provided. Additionally, antibodies, heavy chains, and light chains that contain one or more specific complementarity determining regions (CDRs) are provided. Polynucleotides encoding antibodies against CCR8 are provided. Polynucleotides encoding the antibody heavy or light chains are also provided. Methods for producing and / or purifying antibodies against CCR8 are provided. Fusion proteins comprising CCL1 or MC148 and an Fc region are also provided. Polynucleotides encoding such fusion proteins are also provided. Methods for producing and / or purifying fusion proteins are provided. Therapeutic methods using antibodies and / or fusion proteins are provided. Such methods include, but are not limited to, methods for treating cancer. Methods for detecting CCR8 are provided. Such methods include methods for identifying individuals who may benefit from treatment with antibodies or fusion proteins provided herein, monitoring treatment of individuals with antibodies or fusion proteins provided herein, and improving the therapeutic efficacy of antibodies or fusion proteins provided herein in individuals.

[0045] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0046] All references cited herein, including patent applications, patent publications, and Genbank accession numbers, are incorporated by reference herein as if each individual reference was specifically and individually indicated to be incorporated by reference in its entirety.

[0047] The techniques and procedures described or referenced herein are generally well understood and can be found, for example, in Sambrook et al., Molecular Cloning: A Laboratory Manual 3rd edition (2001), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; CURRENT PROTOCOLS IN MOLECULAR BIOLOGY (F.M.A.usubel, et al. eds., (2003)); the series METHODS IN ENZYMOLOGY (Academic Press, Inc.); PCR 2: A PRACTICAL APPROACH (M.J. MacPherson, B.D. Hames and G.R. Taylor eds. (1995)), Harlow and Lane, eds. (1988); ANTIBODIES, A LABORATORY MANUAL, and ANIMAL CELL CULTURE (R.I. Freshney, ed. (1987)); Oligonucleotide Synthesis (M.J. Gait, ed., 1984); Methods in Molecular Biology, Humana Press; Cell Biology: A Laboratory Notebook (JECellis, ed., 1998) Academic Press; Animal Cell Culture (RIFreshney), ed., 1987); Introduction to Cell and Tissue Culture (JP Mather and PE Roberts, 1998) Plenum Press; Cell and Tissue Culture Laboratory Procedures (A. DGNewell, eds., 1993-8) J. Wiley and Sons; Handbook of Experimental Immunology (DMWeir and CC Blackwell, eds.); Gene Transfer Vectors for Mammalian Cells (JMMiller and MPCalos, eds., 1987); PCR: The Polymerase Chain Reaction, (Mullis et al., eds., 1994); Current Protocols in Immunology (JEColigan et al., eds., 1991); Short Protocols in Molecular Biology (Wiley and Sons, 1999); Immunobiology (CA Janeway and P. Travers, 1997); Antibodies (P. Finch, 1997); Antibodies: A Practical Approach (D. Catty., ed., IRL Press, 1988-1989); Monoclonal Antibodies: A Practical Approach (P. Shepherd and C. Dean, eds., Oxford University Press, 2000); Using Antibodies: A Laboratory Manual (E. Harlow and D.Lane(Cold Spring Harbor Laboratory Press,1999);The Antibodies(M.Zanetti and JDCapra, eds., Harwood Academic. Publishers, 1995); and Cancer: Principles and Practice of Oncology (VT DeVita et al., eds., J.B. Lippincott Company, 1993); and updated versions thereof, are commonly employed using conventional methodologies by those skilled in the art. I. Definition

[0048] Unless otherwise defined, scientific and technical terms used in connection with this disclosure shall have the meanings commonly understood by those of ordinary skill in the art. Further, unless otherwise required by context or expressly indicated, singular terms shall include the plural and plural terms shall include the singular. In the event of discrepancies in definitions among various sources or references, the definitions provided herein shall control.

[0049] It is understood that embodiments of the invention described herein include embodiments "consisting of" and / or "consisting essentially of." As used herein, the singular forms "a," "an," and "the" include plural references unless otherwise indicated. The use of the term "or" herein is not meant to imply that alternatives are mutually exclusive.

[0050] In this application, the use of "or" means "and / or" unless expressly stated otherwise or understood by one of ordinary skill in the art. In the context of multiple dependent claims, the use of "or" refers back to one or more preceding independent or dependent claims.

[0051] As will be understood by one of ordinary skill in the art, reference herein to "about" a value or parameter includes (and describes) embodiments relating to that value or parameter per se. For example, a description referring to "about X" includes a description of "X."

[0052] The terms "nucleic acid molecule," "nucleic acid," and "polynucleotide" may be used interchangeably and refer to a polymer of nucleotides. Such polymers of nucleotides may contain natural and / or non-natural nucleotides and include, but are not limited to, DNA, RNA, and PNA. A "nucleic acid sequence" refers to the linear sequence of nucleotides that comprises a nucleic acid molecule or polynucleotide.

[0053] The terms "polypeptide" and "protein" are used interchangeably to refer to polymers of amino acid residues and are not limited to a minimum length. Such polymers of amino acid residues may contain natural or unnatural amino acid residues and include, but are not limited to, peptides, oligopeptides, dimers, trimers, and multimers of amino acid residues. Both full-length proteins and fragments thereof are encompassed by the definition. The term also includes post-expression modifications of the polypeptide, such as glycosylation, sialylation, acetylation, phosphorylation, and the like. Furthermore, for purposes of this disclosure, "polypeptide" refers to proteins containing modifications, such as deletions, additions, and substitutions (generally conservative in nature), relative to the native sequence, so long as the protein maintains the desired activity. These modifications may be deliberate, through site-directed mutagenesis, or may be accidental, such as through mutation of the host that produces the protein or errors by PCR amplification.

[0054] "CCR8" and "CC chemokine receptor type 8" and "chemokine receptor 8," as used herein, refer to any native CCR8 resulting from expression and processing of CCR8 in a cell. The terms include CCR8 from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys) and rodents (e.g., mice and rats), unless otherwise indicated. The terms also include naturally occurring variants of CCR8, such as splice variants or allelic variants. The amino acid sequence of an exemplary human CCR8 protein is set forth in SEQ ID NO: 101 (UniProt identifier P51685). The amino acid sequence of an exemplary mouse CCR8 protein is set forth in SEQ ID NO: 102 (UniProt identifier P56484). The amino acid sequence of an exemplary cynomolgus monkey CCR8 protein is set forth in SEQ ID NO: 103 (UniProt identifier G7NYJ2).

[0055] "CCL1" and "CC motif chemokine 1," as used herein, refer to any native CCR1 resulting from expression and processing of CCR1 in cells. The terms include CCR1 from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys) and rodents (e.g., mice and rats), unless otherwise indicated. The terms also include naturally occurring variants of CCR1, such as splice variants or allelic variants. The amino acid sequence of an exemplary human CCR1 protein is set forth in SEQ ID NO: 2 (UniProt identifier P22362.1). An exemplary mature CCR1 protein comprises amino acids 24-96 of SEQ ID NO: 2.

[0056] "MC148," as used herein, refers to any native MC148 resulting from expression and processing of MC148 in a cell. The term includes MC148 from any viral source, including molluscum contagiosum virus (subtype 1 or subtype 2), unless otherwise indicated. The term also includes naturally occurring variants of MC148, such as truncated or allelic variants. The amino acid sequence of an exemplary MC148 protein is set forth in SEQ ID NO: 6 (amino acids 25-104 of UniProt identifier Q98314.1).

[0057] As used herein, "7-B16 antibody" should be understood as any antibody that binds to CCR8 and comprises (i) a heavy chain comprising SEQ ID NO: 82 and a light chain comprising SEQ ID NO: 83, (ii) a variable heavy chain region comprising SEQ ID NO: 80 and a variable light chain region comprising SEQ ID NO: 81, or (iii) HCDR1, HCDR2, and HCDR3 comprising SEQ ID NOs: 84, 85, and 86, respectively, and LCDR1, LCDR2, and LCDR3 comprising SEQ ID NOs: 87, 88, and 89, respectively, and chimeric, human, or humanized versions of any of the foregoing (i), (ii), or (iii). In some embodiments, "7-B16 antibody" may be used to specifically refer to an antibody comprising a heavy chain of SEQ ID NO: 82 and a light chain of SEQ ID NO: 83.

[0058] The term "specifically binds" to an antigen or epitope is a term well understood in the art, and methods for determining such specific binding are also well known in the art. A molecule is said to exhibit "specific binding" or "preferential binding" if it reacts or associates with a particular cell or substance more frequently, more rapidly, with a longer duration, and / or with a higher affinity than it reacts or associates with alternative cells or substances. An antibody "specifically binds" or "preferentially binds" to a target if it binds with higher affinity, avidity, more readily, and / or with a longer duration than it binds to other substances. For example, an antibody that specifically or preferentially binds to a CCR8 epitope is an antibody that binds to this epitope with greater affinity, avidity, more readily, and / or with a longer duration than it binds to other CCR8 epitopes or non-CCR8 epitopes. It should also be understood by reading this definition that, for example, an antibody (or portion or epitope) that specifically or preferentially binds to a first target may or may not specifically or preferentially bind to a second target. Thus, "specific binding" or "preferential binding" does not necessarily require (but can include) exclusive binding. Generally, although not necessarily, reference to binding means preferential binding. "Specificity" refers to the ability of a binding protein to selectively bind to an antigen.

[0059] As used herein, "substantially pure" refers to a material that is at least 50% pure (i.e., free from contaminants), more preferably at least 90% pure, more preferably at least 95% pure, even more preferably at least 98% pure, and most preferably at least 99% pure.

[0060] As used herein, the term "epitope" refers to a site on a target molecule (e.g., an antigen, such as a protein, nucleic acid, carbohydrate, or lipid) to which an antigen-binding molecule (e.g., an antibody, antibody fragment, or scaffold protein containing an antibody-binding region) binds. Epitopes often comprise chemically active surface groupings of molecules, such as amino acids, polypeptides, or sugar side chains, and have specific three-dimensional structural and charge characteristics. Epitopes can be formed from both adjacent and / or juxtaposed non-adjacent residues (e.g., amino acids, nucleotides, sugars, and lipid moieties) of a target molecule. Epitopes formed from adjacent residues (e.g., amino acids, nucleotides, sugars, and lipid moieties) typically persist on exposure to denaturing solvents, whereas epitopes formed by tertiary folding typically are lost upon treatment with denaturing solvents. Epitopes can include, but are not limited to, at least 3, at least 5, or 8-10 residues (e.g., amino acids or nucleotides). In some examples, an epitope is less than 20 residues (e.g., amino acids or nucleotides) in length, less than 15 residues, or less than 12 residues. Two antibodies may bind to the same epitope within an antigen if they exhibit competitive binding to the antigen. In some embodiments, an epitope can be identified by a specific minimum distance to CDR residues on an antigen-binding molecule. In some embodiments, an epitope can be identified by the above distance and further limited to residues involved in binding (e.g., hydrogen bonding) between antibody and antigen residues. Epitopes can also be identified by various scans; for example, an alanine or arginine scan can reveal one or more residues with which an antigen-binding molecule can interact. Unless explicitly indicated, identifying a set of residues as an epitope does not exclude other residues from being part of the epitope for a particular antibody. Rather, the existence of such a set indicates a minimal series (or species set) of epitopes. Thus, in some embodiments, the set of residues identified as an epitope refers to a minimal epitope associated with the antigen, rather than an exclusive list of residues of the epitope on the antigen.

[0061] A "nonlinear epitope" or "conformational epitope" comprises noncontiguous polypeptides, amino acids, and / or sugars within an antigenic protein to which an epitope-specific antibody binds. In some embodiments, at least one of the residues is noncontiguous with other listed residues of the epitope; however, one or more of the residues may also be contiguous with other residues.

[0062] A "linear epitope" comprises contiguous polypeptides, amino acids, and / or sugars within an antigenic protein to which an antibody specific for the epitope binds. Note that in some embodiments, not all of the residues within a linear epitope need directly bind (or be involved in binding) to an antibody. In some embodiments, a linear epitope can result from immunization with a peptide that effectively consists of the sequence of the linear epitope, or from only a structural section of the protein, a sequence section of the protein that is relatively isolated from the rest of the protein (so that an antibody can at least primarily interact with it).

[0063] The term "antibody" as used herein is used in the broadest sense and encompasses a variety of antibody structures, including, but not limited to, monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific and trispecific antibodies (such as bispecific T cell engagers)), and antibody fragments so long as they exhibit the desired antigen-binding activity.

[0064] The term antibody includes, but is not limited to, fragments capable of binding antigen, such as Fv, single-chain Fv (scFv), Fab, Fab', di-scFv, sdAb (single domain antibody), and (Fab')2 (including chemically linked F(ab')2). Papain digestion of antibodies produces two identical antigen-binding fragments, called "Fab" fragments, each with a single antigen-binding site, and a residual "Fc" fragment, the name reflecting its ability to readily crystallize. Pepsin treatment yields an F(ab')2 fragment, which has two antigen-binding sites and is capable of cross-linking antigen. The term antibody includes, but is not limited to, chimeric antibodies, humanized antibodies, and antibodies of various species, such as murine, human, and cynomolgus monkey. Furthermore, for all antibody constructs provided herein, variants with sequences derived from other organisms are also contemplated. Thus, if a human version of an antibody is disclosed, one of skill in the art would understand how to convert the human sequence-based antibody to a mouse, rat, cat, dog, horse, etc. sequence. Antibody fragments also include any orientation of single-chain scFvs, tandem di-scFvs, diabodies, tandem tri-sdcFvs, minibodies, etc. Antibody fragments also include nanobodies (sdAbs, which are antibodies with a single monomer domain, such as a pair of lower limit domains of a heavy chain without a light chain). Antibody fragments may in some embodiments be referred to as being of a specific species (e.g., human scFv or mouse scFv). This indicates the sequence of at least a portion of the non-CDR regions, not the source of the construct.

[0065] The term "monoclonal antibody" refers to an antibody from a substantially homogeneous antibody population, i.e., the individual antibodies comprising the population are identical except for possible minor naturally occurring mutations. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. Thus, a sample of monoclonal antibody is capable of binding to the same epitope on the antigen. The modifier "monoclonal" indicates the character of the antibody as being obtained from a substantially homogeneous antibody population and should not be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies can be made by the hybridoma method first described by Kohler and Milstein, 1975, Nature 256:495, or by recombinant DNA methods such as those described in U.S. Pat. No. 4,816,567. The monoclonal antibodies may also be isolated from phage libraries generated using the techniques described in McCafferty et al., 1990, Nature 348:552-554, for example.

[0066] The term "CDR" refers to a complementarity-determining region defined by at least one mode of identification to one of ordinary skill in the art. In some embodiments, CDRs may be defined according to any of the following: the Chothia numbering scheme, the Kabat numbering scheme, a combination of Kabat and Chothia, the AbM definition, the contact definition, and / or a combination of the Kabat, Chothia, AbM, and / or contact definitions. Exemplary CDRs (CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2, and CDR-H3) occur at amino acid residues 24-34 of L1, 50-56 of L2, 89-97 of L3, 31-35B of H1, 50-65 of H2, and 95-102 of H3. (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD (1991)). The AbM definition may include, for example, CDRs at amino acid residues 24-34 of L1, 50-56 of L2, 89-97 of L3, H26-H35B of H1, 50-58 of H2, and 95-102 of H3 (CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2, and CDR-H3).The contact definition may include, for example, CDRs at amino acid residues 30-36 of L1, 46-55 of L2, 89-96 of L3, 30-35 of H1, 47-58 of H2, and 93-101 of H3 (CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2, and CDR-H3). The Chothia definition may include, for example, CDRs at amino acid residues 24-34 of L1, 50-56 of L2, 89-97 of L3, 26-32...34 of H1, 52-56 of H2, and 95-102 of H3 (CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2, and CDR-H3). CDRs may also be provided as shown in any one or more of the accompanying figures. HWith the exception of CDR1, CDRs generally comprise amino acid residues that form hypervariable loops. The various CDRs within an antibody may be designated by their appropriate number and chain type, including, but not limited to, a) CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2, and CDR-H3; b) CDRL1, CDRL2, CDRL3, CDRH1, CDRH2, and CDRH3; c) LCDR-1, LCDR-2, LCDR-3, HCDR-1, HCDR-2, and HCDR-3; or d) LCDR1, LCDR2, LCDR3, HCDR1, HCDR2, and HCDR3. The term "CDR" is used herein to encompass HVRs or "hypervariable regions," which comprise the hypervariable loops. Exemplary hypervariable loops occur at amino acid residues 26-32 (L1), 50-52 (L2), 91-96 (L3), 26-32 (H1), 53-55 (H2), and 96-101 (H3) (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)).

[0067] The term "heavy chain variable region," as used herein, refers to a region comprising at least three heavy chain CDRs. In some embodiments, a heavy chain variable region comprises three CDRs and at least FR2 and FR3. In some embodiments, a heavy chain variable region comprises at least heavy chain HCDR1, framework (FR) 2, HCDR2, FR3, and and HCDR3. In some embodiments, the heavy chain variable region also comprises at least a portion of FR1 and / or at least a portion of FR4.

[0068] The term "heavy chain constant region" as used herein refers to a region comprising at least three heavy chain constant domains: H 1. C H 2, and C H3. Of course, deletions and modifications that do not alter function within the domain are encompassed within the term "heavy chain constant region" unless otherwise specified. Non-limiting exemplary heavy chain constant regions include gamma, delta, and alpha. Non-limiting exemplary heavy chain constant regions also include epsilon and mu. Each heavy constant region corresponds to an antibody isotype. For example, an antibody containing a gamma constant region is an IgG antibody, an antibody containing a delta constant region is an IgD antibody, and an antibody containing an alpha constant region is an IgA antibody. Furthermore, an antibody containing a mu constant region is an IgM antibody, and an antibody containing an epsilon constant region is an IgE antibody. A particular isotype can be further subdivided into subclasses. For example, IgG antibodies include, but are not limited to, IgG1 (containing a gamma 1 constant region), IgG2 (containing a gamma 2 constant region), IgG3 (containing a gamma 3 constant region), and IgG4 (containing a gamma 4 constant region) antibodies; IgA antibodies include, but are not limited to, IgA1 (containing an alpha 1 constant region) and IgA2 (containing an alpha 2 constant region) antibodies; and IgM antibodies include, but are not limited to, IgM1 and IgM2.

[0069] As used herein, the term "heavy chain" refers to a polypeptide comprising at least a heavy chain variable region, with or without a leader sequence. In some embodiments, a heavy chain includes at least a portion of a heavy chain constant region. As used herein, the term "full-length heavy chain" refers to a polypeptide comprising a heavy chain variable region and a heavy chain constant region, with or without a leader sequence.

[0070] As used herein, the term "light chain variable region" refers to a region comprising at least three light chain CDRs. In some embodiments, a light chain variable region comprises three CDRs and at least FR2 and FR3. In some embodiments, a light chain variable region comprises at least a light chain LCDR1, framework (FR) 2, LCDR2, FR3, and LCDR3. For example, a light chain variable region may comprise a light chain CDR1, framework (FR) 2, CDR2, FR3, and CDR3. In some embodiments, a light chain variable region also comprises at least a portion of FR1 and / or at least a portion of FR4.

[0071] As used herein, the term "light chain constant region" refers to a light chain constant domain C L "light chain constant region" refers to a region comprising: a) a region comprising a light chain constant region (e.g., ...

[0072] As used herein, the term "light chain" refers to a polypeptide comprising at least a light chain variable region, with or without a leader sequence. In some embodiments, a light chain also comprises at least a portion of a light chain constant region. As used herein, the term "full-length light chain" refers to a polypeptide comprising a light chain variable region and a light chain constant region, with or without a leader sequence.

[0073] For purposes herein, an "acceptor human framework" refers to a light chain variable domain (V) derived from a human immunoglobulin framework or a human consensus framework, as defined below. L ) framework or heavy chain variable domain (V H) framework. The acceptor human framework, derived from a human immunoglobulin framework or a human consensus framework, can comprise the same amino acid sequence or can contain amino acid sequence changes. In some embodiments, the number of amino acid changes is 10 or less, 9 or less, 8 or less, 7 or less, 6 or less, 5 or less, 4 or less, 3 or less, or 2 or less. In some embodiments, the V L Acceptor framework is V L The sequence is identical to a human immunoglobulin framework sequence or a human consensus framework sequence.

[0074] "Affinity" refers to the strength of the sum of non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). The affinity of molecule X for partner Y is generally determined by the dissociation constant (K D Affinity can be expressed by common methods known in the art (e.g., ELISA, including those described herein). D , KinExA, biolayer interferometry (BLI), and / or surface plasmon resonance devices (such as BIAcore® devices).

[0075] "K D The term "" as used herein refers to the equilibrium dissociation constant of an antibody-antigen interaction.

[0076] In some embodiments, the "K" of the antibody D "," "K d"," "Kd," or "Kd value" is measured by using a surface plasmon resonance assay with a BIACORE®-2000 or BIACORE®-3000 (BIAcore, Inc., Piscataway, NJ) on an antigen-immobilized CM5 chip at approximately 10 response units (RU) at 25°C. Briefly, carboxymethylated dextran biosensor chips (CM5, BIACORE, Inc.) were prepared by adding N-ethyl-N'-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC) and carboxymethylated dextran biosensor chips (CM5, BIACORE, Inc.) according to the supplier's instructions. The antibody is activated with N-hydroxysuccinimide (NHS). The antigen is diluted to 5 μg / ml (approximately 0.2 μM) in 10 mM sodium acetate, pH 4.8, and then injected at a flow rate of 5 μL / min to achieve approximately 10 response units (RU) of bound protein. After antigen injection, 1 M ethanolamine is injected to block unreacted groups. For kinetic measurements, serial dilutions of a polypeptide, e.g., a full-length antibody, are injected in PBS containing 0.05% TWEEN®-20 surfactant (PBST) at 25°C at a flow rate of approximately 25 μL / min. The association rate (k on ) and dissociation rate (k off The equilibrium dissociation constant (K) is calculated using a simple one-to-one Langmuir binding model (BIACORE® evaluation software version 3.2) by simultaneously fitting the association and dissociation sensorgrams. d ) is k off / k on See, e.g., Chen et al., J. Mol. Biol. 293:865-881 (1999). If the on-rate is 10 by the surface plasmon resonance assay described above, 6 M -1 s -1If the on-rate exceeds 20 nM, the on-rate can be determined at 25°C of 20 nM anti-antigen antibody in PBS, pH 7.2, in the presence of increasing concentrations of antigen, by using a fluorescence quenching technique that measures the increase or decrease in fluorescence emission intensity (excitation = 295 nm, emission = 340 nm, 16 nm bandpass) as measured in a spectrometer such as a spectrophotometer equipped with stopped flow (Aviv Instruments) or an 8000 Series SLM-AMINCO™ spectrophotometer (ThermoSpectronic) equipped with a stirred cuvette.

[0077] In some embodiments, the two values ​​(e.g., K d The difference between the reference / comparison values ​​is substantially the same, e.g., less than about 50%, less than about 40%, less than about 30%, less than about 20%, and / or less than about 10%, as a function of the reference / comparison value.

[0078] In some embodiments, the two values ​​(e.g., K d The difference between the values) is substantially different as a function of the value of the reference / comparator molecule, e.g., greater than about 10%, greater than about 20%, greater than about 30%, greater than about 40%, and / or greater than about 50%.

[0079] "Surface plasmon resonance" refers to an optical phenomenon that allows for the analysis of real-time biospecific interactions by detecting alterations in protein concentration within a biosensor matrix, for example, using the BIAcore™ system (BIAcore International AB, a GE Healthcare company, Uppsala, Sweden and Piscataway, NJ). For further explanation, see Jonsson et al. (1993) Ann. Biol. Clin. 51:19-26.

[0080] "Bio-layer interferometry" refers to an optical analysis technique that analyzes the interference pattern of light reflected from a layer of immobilized proteins on a biosensor tip and an internal reference layer. Changes in the number of molecules bound to the biosensor tip cause a shift in the interference pattern that can be measured in real time. A non-limiting exemplary device for bio-layer interferometry is the ForteBio Octet® RED96 system (Pall Corporation). See, e.g., Abdiche et al., 2008, Anal. Biochem. 377:209-277.

[0081] "k on The term "as used herein" refers to the rate constant for the association of an antibody to an antigen. Specifically, using IgG (bivalent) with monovalent CCR8 antigen, the rate constant (k on and k off ) and the equilibrium dissociation constant are measured. on "," k on The terms "kA", "association rate constant", or "kA" are used interchangeably herein. The value indicates the rate of binding of a binding protein to its target antigen or the rate of complex formation between an antibody and an antigen, and is given by the formula: antibody ("Ab") + antigen ("Ag") → Ab-Ag. "k off The term k as used herein refers to the rate constant for dissociation of an antibody from the antibody / antigen complex. off is "K off " or "dissociation rate constant." This value indicates the rate of dissociation of an antibody from its target antigen or the separation of the Ab-Ag complex into free antibody and antigen over time, as shown by the following equation: Ab+Ag←Ab-Ag.

[0082] The term "biological activity" refers to any one or more biological properties of a molecule (whether naturally occurring as found in vivo or provided or enabled by recombinant means). Biological properties include, but are not limited to, binding to cytokines, inducing cell proliferation, inhibiting cell growth, inducing other cytokines, inducing apoptosis, and inducing enzymatic activity. In some embodiments, the biological activities of CCR8 include anti-apoptotic activity, cell chemotaxis, immunosuppressive function, and the ability to polarize cells toward various cell differentiation pathways.

[0083] An "affinity matured" antibody refers to an antibody with one or more alterations in one or more CDRs compared to a parent antibody that does not possess such alterations, which alterations result in an improvement in the affinity of the antibody for antigen.

[0084] "Chimeric antibody," as used herein, refers to an antibody in which a portion of the heavy and / or light chain is derived from a particular source or species, while at least a portion of the remaining heavy and / or light chain is derived from a different source or species. In some embodiments, a chimeric antibody refers to an antibody comprising at least one variable region derived from a first species (mouse, rat, cynomolgus monkey, etc.) and at least one constant region derived from a second species (human, cynomolgus monkey, etc.). In some embodiments, a chimeric antibody comprises at least one mouse variable region and at least one human constant region. In some embodiments, a chimeric antibody comprises at least one cynomolgus monkey variable region and at least one human constant region. In some embodiments, all of the variable regions of a chimeric antibody are derived from a first species, and all of the constant regions of a chimeric antibody are derived from a second species. As noted above, chimeric constructs can also be functional fragments.

[0085] As used herein, "humanized antibody" refers to an antibody in which at least one amino acid in a framework region of a non-human variable region is replaced with the corresponding amino acid from a human variable region. In some embodiments, a humanized antibody comprises at least one human constant region or fragment thereof. In some embodiments, a humanized antibody is an antibody fragment such as a Fab, scFv, or (Fab')2. The term humanized also refers to forms of non-human (e.g., murine) antibodies that are chimeric immunoglobulins, immunoglobulin chains, or fragments thereof (such as Fv, Fab, Fab', F(ab')2, or other antigen-binding subsequences of antibodies) that contain minimal sequence of a non-human immunoglobulin. Humanized antibodies can include a human immunoglobulin (recipient antibody) in which residues from the recipient's complementarity-determining region (CDR) are replaced by residues from a CDR of a non-human species (donor antibody), e.g., mouse, rat, or rabbit, possessing the desired specificity, affinity, and capacity. In some instances, Fv framework region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues. Furthermore, humanized antibodies can comprise residues found neither in the recipient antibody nor in the imported CDR or framework sequences, but which are included to further refine and optimize antibody performance. Generally, a humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the CDR regions correspond to those of a non-human immunoglobulin and all or substantially all of the FR regions are those of a human immunoglobulin consensus sequence. In some embodiments, the humanized antibody will also comprise at least a portion of an immunoglobulin constant region or domain (Fc), typically that of a human immunoglobulin. Other forms of humanized antibodies have one or more CDRs (CDRL1, CDRL2, CDRL3, CDRH1, CDRH2, and / or CDRH3) that are altered with respect to the original antibody, also referred to as one or more CDRs "derived from" one or more CDRs from the original antibody. It will be understood that a humanized sequence can be identified by its primary sequence and does not necessarily indicate the process by which the antibody was produced.

[0086] "CDR-grafted antibody," as used herein, refers to a humanized antibody in which one or more complementarity-determining regions (CDRs) of a first (non-human) species have been grafted onto framework regions (FRs) of a second (human) species.

[0087] As used herein, "human antibody" encompasses antibodies produced in humans, antibodies produced in non-human animals containing human immunoglobulin genes, such as XenoMouse® mice, and antibodies selected using in vitro methods such as phage display (Vaughan et al., 1996, Nature Biotechnology, 14:309-314; Sheets et al., 1998, Proc. Natl. Acad. Sci. (USA) 95:6157-6162; Hoogenboom and Winter, 1991, J. Mol. Biol., 227:381; Marks et al., 1991, J. Mol. Biol., 222:581), where the antibody repertoire is based on human immunoglobulin sequences. The term "human antibody" refers to the genus of sequences that are human sequences. Thus, the term does not designate the process by which the antibody is made, but rather the genus of related sequences.

[0088] A "functional Fc region" possesses an "effector function" of a native sequence Fc region. Exemplary "effector functions" include Fc receptor binding, C1q binding, CDC, ADCC, phagocytosis, and down-regulation of a cell surface receptor (e.g., B cell receptor, BCR). Such effector functions generally require an Fc region in combination with a binding domain (e.g., an antibody variable domain) and can be assessed using a variety of assays.

[0089] A "native sequence Fc region" comprises an amino acid sequence identical to that of an Fc region found in nature. Native sequence human Fc regions include native sequence human IgG1 Fc regions (non-A and A allotypes), native sequence human IgG2 Fc regions, native sequence human IgG3 Fc regions, and native sequence human IgG4 Fc regions, as well as naturally occurring variants thereof.

[0090] A "variant Fc region" comprises an amino acid sequence that differs from that of a native-sequence Fc region by at least one amino acid modification. In some embodiments, a "variant Fc region" comprises an amino acid sequence that differs from that of a native-sequence Fc region by at least one amino acid modification, and retains at least one effector function of the native-sequence Fc region. In some embodiments, the variant Fc region has at least one amino acid substitution compared to a native-sequence Fc region or the Fc region of a parent polypeptide, e.g., about 1 to about 10 amino acid substitutions, preferably about 1 to about 5 amino acid substitutions, in the native-sequence Fc region or the Fc region of a parent polypeptide. In some embodiments, the variant Fc region herein has at least 80% sequence identity to a native sequence Fc region and / or the Fc region of a parent polypeptide, at least about 90% sequence identity to a native sequence Fc region and / or the Fc region of a parent polypeptide, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% sequence identity to a native sequence Fc region and / or the Fc region of a parent polypeptide.

[0091] "Fc receptor" or "FcR" describes a receptor that binds to the Fc region of an antibody. In some embodiments, the FcγR is a native human FcR. In some embodiments, the FcR binds IgG antibodies (gamma receptors) and includes receptors of the FcγRI, FcγRII, and FcγRIII subclasses, including allelic variants and alternatively spliced ​​forms of those receptors. FcγRII receptors include FcγRIIA (an "activating receptor") and FcγRIIB (an "inhibiting receptor"), which have similar amino acid sequences that differ primarily in their cytoplasmic domains. Activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain, and inhibitory receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibition motif (ITIM) in its cytoplasmic domain. (See, e.g., Daeron, Annu. Rev. Immunol. 15:203-234 (1997).) FcRs are reviewed, for example, in Ravetch and Kinet, Annu. Rev. Immunol 9:457-92 (1991), Capel et al., Immunomethods 4:25-34 (1994), and de Haas et al., J. Lab. Clin. Med. 126:330-41 (1995). Other FcRs, including those identified in the future, are encompassed by the term "FcR" herein.

[0092] The term "Fc receptor" or "FcR" also includes the neonatal receptor FcRn, which is involved in the transfer of maternal IgG to the fetus (Guyer et al., J. Immunol. 117:587 (1976) and Kim et al., J. Immunol. 24:249 (1994)) and in regulating immunoglobulin homeostasis. Methods for measuring binding to FcRn are known (see, e.g., Ghetie and Ward, Immunol. Today 18(12):592-598 (1997); Ghetie et al., Nature Biotechnology, 15(7):637-640 (1997); Hinton et al., J. Biol. Chem. 279(8):6213-6216 (2004); WO 2004 / 92219 (Hinton et al.)).

[0093] "Effector functions" refer to biological activities attributable to the Fc region of an antibody, which vary depending on the antibody isotype. Examples of antibody effector functions include Clq binding and complement-dependent cytotoxicity (CDC), Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, and binding to cell surface receptors (e.g., These include downregulation of the B cell receptor (B cell receptor), as well as B cell activation.

[0094] A "human effector cell" is a leukocyte that expresses one or more FcRs and performs effector function. In some embodiments, the cells express at least FcγRIII and perform ADCC effector function. Examples of human leukocytes that mediate ADCC include peripheral blood mononuclear cells (PBMCs), natural killer (NK) cells, monocytes, cytotoxic T cells, and neutrophils. Effector cells can be isolated from a native source, for example, from blood.

[0095] "Antibody-dependent cell-mediated cytotoxicity" or "ADCC" refers to a form of cytotoxicity in which secreted Ig binds to Fc receptors (FcRs) present on certain cytotoxic cells (e.g., NK cells, neutrophils, and macrophages) so that these cytotoxic effector cells can specifically bind to antigen-bearing target cells and subsequently kill them with cytotoxins. NK cells, the primary cells for mediating ADCC, express only FcγRIII, whereas monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol 9:457-92 (1991). To assess ADCC activity of a molecule of interest, an in vitro ADCC assay, such as that described in U.S. Patent No. 5,500,362 or U.S. Patent No. 5,821,337, or U.S. Patent No. 6,737,056 (Presta), can be performed. Useful effector cells for such assays include PBMCs and NK cells. Alternatively, or additionally, ADCC activity of a molecule of interest can be assessed in vivo in an animal model, such as that disclosed in Clynes et al. Proc. Natl. Acad. Sci. (USA) 95:652-656 (1998). Additional polypeptide variants with altered Fc region amino acid sequences (polypeptides with variant Fc regions) and increased or decreased ADCC activity are described, for example, in U.S. Patent Nos. 7,923,538 and 7,994,290.

[0096] "Complement-dependent cytotoxicity" or "CDC" refers to the lysis of target cells in the presence of complement. Activation of the classical complement pathway is initiated by the binding of the first component of the complement system (C1q) to antibodies (of the appropriate subclass) that are bound to their cognate antigen. To assess complement activation, for example, a CDC assay as described in Gazzano-Santoro et al., J. Immunol. Methods 202:163 (1996) can be performed. Polypeptide variants with altered Fc region amino acid sequences (polypeptides with variant Fc regions) and increased or decreased C1q binding ability are described, for example, in U.S. Pat. No. 6,194,551 (B1), U.S. Pat. No. 7,923,538, U.S. Pat. No. 7,994,290, and WO 1999 / 51642. See, e.g., Idusogie et al., J. Immunol. 164:4178-4184 (2000).

[0097] Polypeptide variants with "altered" FcR binding affinity or ADCC activity are those that have either enhanced or decreased FcR binding activity and / or ADCC activity compared to a parent polypeptide or a polypeptide comprising a native-sequence Fc region. Polypeptide variants that "exhibit increased binding" to an FcR bind to at least one FcR with better affinity than the parent polypeptide. Polypeptide variants that "exhibit decreased binding" to an FcR bind to at least one FcR with lower affinity than the parent polypeptide. Such variants that exhibit decreased binding to an FcR may exhibit little or no binding to an FcR compared to a native-sequence IgG Fc region, e.g., 0-20% binding to an FcR.

[0098] A polypeptide variant that "more effectively mediates antibody-dependent cell-mediated cytotoxicity (ADCC) in the presence of human effector cells" than a parent antibody is one that is more effective at mediating ADCC in vitro or in vivo when essentially the same amounts of polypeptide variant and parent antibody are used in the assay. Generally, such variants are identified using the in vitro ADCC assays disclosed herein, although other assays or methods for determining ADCC activity, such as in animal models, are contemplated.

[0099] The terms "substantially similar" or "substantially the same," as used herein, refer to a sufficiently high degree of similarity between two or more numerical values ​​such that one of skill in the art would regard the difference between the two or more values ​​as having little or no biological and / or statistical significance within the context of the biological property measured by the values. In some embodiments, two or more substantially similar values ​​differ by no more than about any one of 5%, 10%, 15%, 20%, 25%, or 50%.

[0100] The phrase "substantially different," as used herein, refers to a sufficiently high degree of difference between two numerical values ​​such that one of skill in the art would consider the difference between the two values ​​within the context of the biological property measured by the values ​​to be statistically significant. In some embodiments, two substantially different numerical values ​​differ by more than any one of about 10%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0101] The phrase "substantially reduced," as used herein, refers to a sufficiently high degree of reduction between a numerical value and a reference numerical value such that one of skill in the art would consider the difference between the two values ​​within the context of the biological property measured by the values ​​to be statistically significant. In some embodiments, a substantially reduced numerical value is a reduction of more than about any one of 10%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90%, or 100% compared to the reference value.

[0102] The term "leader sequence" refers to a sequence of amino acid residues located at the N-terminus of a polypeptide that facilitates secretion of the polypeptide from mammalian cells. Leader sequences may be cleaved upon export of the polypeptide from mammalian cells to form the mature protein. Leader sequences may be natural or synthetic, and they may be heterologous or homologous to the protein to which they are attached.

[0103] A "native sequence" polypeptide includes a polypeptide having the same amino acid sequence as a polypeptide found in nature. Thus, a native sequence polypeptide can have the amino acid sequence of a naturally occurring polypeptide from any mammal. Such native sequence polypeptides can be isolated from nature or produced by recombinant or synthetic means. The term "native sequence" polypeptide specifically encompasses naturally occurring truncated or secreted forms of a polypeptide (e.g., extracellular domain sequences), naturally occurring variant forms (e.g., alternatively spliced ​​forms), and naturally occurring allelic variants of a polypeptide.

[0104] A polypeptide "variant" means a biologically active polypeptide having at least about 80% amino acid sequence identity with a native sequence polypeptide after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity and not considering any conservative substitutions as part of the sequence identity. Such variants include, for example, polypeptides in which one or more amino acid residues are added or deleted at the N- or C-terminus of the polypeptide. In some embodiments, a variant will have at least about 80% amino acid sequence identity. In some embodiments, a variant will have at least about 90% amino acid sequence identity. In some embodiments, a variant will have at least about 95% amino acid sequence identity with a native sequence polypeptide.

[0105] As used herein, "percent (%) amino acid sequence identity" and "homology" with respect to a peptide, polypeptide, or antibody sequence are defined as the percentage of amino acid residues in a candidate sequence that are identical to the amino acid residues of a particular peptide or polypeptide sequence after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be achieved in a variety of ways that are within the skill of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or MEGALIGN™ (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared.

[0106] Amino acid substitutions can include, but are not limited to, substituting one amino acid for another in a polypeptide. Exemplary conservative substitutions are shown in Table 1. Amino acid substitutions can be introduced into an antibody of interest and the products can be screened for a desired activity, such as retained / improved antigen binding, reduced immunogenicity, or improved ADCC or CDC. [Table 1]

[0107] Amino acids can be grouped according to common side chain properties. (1) Hydrophobic: Norleucine, Met, Ala, Val, Leu, Ile, (2) Neutral hydrophilicity: Cys, Ser, Thr, Asn, Gln, (3) Acidification: Asp, Glu, (4) Bases: His, Lys, Arg, (5) Residues that affect chain orientation: Gly, Pro, (6) Aromatic: Trp, Tyr, Phe.

[0108] Non-conservative substitutions would involve exchanging a member of one of these classes for another class.

[0109] The term "vector" is used to describe a polynucleotide that can be manipulated to contain cloned polynucleotide(s) that can be propagated in a host cell. A vector can contain one or more of the following elements: an origin of replication, one or more regulatory sequences (such as, for example, a promoter and / or enhancer) that control the expression of a polypeptide of interest, and / or one or more selectable marker genes (e.g., antibiotic resistance genes and genes that can be used in colorimetric assays, e.g., β-galactosidase). The term "expression vector" refers to a vector used to express a polypeptide of interest in a host cell.

[0110] A "host cell" refers to a cell that can be or has been the recipient of a vector or isolated polynucleotide. Host cells can be prokaryotic or eukaryotic. Exemplary eukaryotic cells include mammalian cells, such as primate or non-primate cells, fungal cells such as yeast, plant cells, and insect cells. Non-limiting exemplary mammalian cells include, but are not limited to, NSO cells, PER.C6,® cells (Crucell), and 293 and CHO cells, as well as their derivatives, such as 293-6E and DG44 cells, respectively. A host cell includes the progeny of a single host cell; the progeny may not necessarily be completely identical (in morphology or genomic DNA complement) to the original parent cell due to natural, accidental, or deliberate mutation. Host cells include cells transfected in vivo with a polynucleotide provided herein.

[0111] The term "isolated," as used herein, refers to a molecule that is separated from at least some of the components typically found or produced in nature. For example, a polypeptide is referred to as "isolated" if it is separated from at least some of the components of the cell in which it is produced. If the polypeptide is secreted by a cell after expression, physically separating the supernatant containing the polypeptide from the cell in which it was produced is considered to "isolate" the polypeptide. Similarly, a polynucleotide is referred to as "isolated" if it is not part of a larger polynucleotide in which it is typically found in nature (e.g., in the case of a DNA polynucleotide, genomic DNA or mitochondrial DNA, etc.), or, for example, in the case of an RNA polynucleotide, if it is separated from at least some of the components of the cell in which it was produced. Thus, a DNA polynucleotide contained in a vector within a host cell can be referred to as "isolated."

[0112] The terms "individual" or "subject" are used interchangeably herein and refer to animals, e.g., mammals. In some embodiments, methods are provided for treating mammals, including, but not limited to, humans, rodents, apes, cats, dogs, horses, cows, pigs, sheep, goats, mammalian laboratory animals, mammalian farm animals, mammalian sport animals, and mammalian pets. In some examples, "individual" or "subject" refers to an individual or subject in need of treatment for a disease or disorder. In some embodiments, the subject receiving treatment may be a patient, which designates the fact that the subject has been identified as having a disorder relevant to the treatment or is at sufficient risk for the disorder.

[0113] "Disease" or "disorder," as used herein, refers to a condition for which treatment is necessary and / or desirable.

[0114] As used herein, "cancer" and "tumor" are interchangeable terms that refer to any abnormal cell or tissue growth or proliferation in an animal. As used herein, the terms "cancer" and "tumor" encompass solid and hematologic / lymphatic cancers, and also encompass malignant, premalignant, and benign growths such as dysplasia. Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and leukemia. More specific, non-limiting examples of such cancers include squamous cell carcinoma, small cell lung cancer, pituitary cancer, esophageal cancer, astrocytoma, soft tissue sarcoma, non-small cell lung cancer, adenocarcinoma of the lung, squamous cell carcinoma of the lung, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, and colorectal cancer. endometrial or uterine cancer, salivary gland cancer, kidney cancer, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, hepatic carcinoma, brain cancer, These include endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, gastric cancer, melanoma, mesothelioma, and various types of head and neck cancer. In some embodiments, hematological / lymphatic cancers are referred to as "blood cancers." Non-limiting exemplary blood cancers include mixed B-cell and T-cell leukemia, B-cell lymphoma, chronic myelogenous leukemia (CML), chronic myelomonocytic leukemia, diffuse large B-cell lymphoma (DLBC), lymphoma, mantle cell lymphoma (MCL), multiple myeloma, myelodysplastic syndrome (MDS), myeloproliferative disorders, peripheral T-cell lymphoma, T-cell leukemia, acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma, and the like. lymphoma, SLL), CLL / SLL, mature T-cell and NK-cell lymphoma, follicular lymphoma, acute lymphocytic leukemia (ALL), T-cell acute lymphocytic leukemia (TALL), T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, cutaneous T-cell lymphoma (CTCL), adult T-cell leukemia / lymphoma (ATLL), T-cell lymphoblastic leukemia / lymphoma (TLLL), angioimmunoblastic T-cell lymphoma (ATCL), hepatosplenic T-cell lymphoma (HTCL), peripheral T-cell lymphoma not otherwise specified (PTCL) These include NOS), Burkitt's lymphoma (BL), chronic myelomonocytic leukemia (CMML), extranodal NK / T-cell lymphoma (NKTCL), primary effusion lymphoma (PEL), acute lymphocytic leukemia / acute myeloid leukemia (ALL, AML), histiocytic lymphoma (HL), marginal zone lymphoma (MZL), B-cell acute lymphocytic leukemia, and anaplastic large cell lymphoma (ALCL).

[0115] As used herein, "treatment" is an approach to obtain beneficial or desired clinical results. "Treatment," as used herein, covers any administration or application of a therapeutic agent for a disease in a mammal, including a human. For purposes of this disclosure, beneficial or desired clinical results include, but are not limited to, any one or more of the following: alleviation of one or more symptoms; reduction in the extent of the disease; prevention or delay of disease spread (e.g., metastasis, e.g., to the lungs or lymph nodes); prevention or delay of disease recurrence; delay or slowing of disease progression; amelioration of the condition; inhibition of the disease or disease progression; inhibition or slowing of the disease or its progression; arrest of its development; and remission (whether partial or total). Also encompassed by "treatment" is the reduction of the pathological consequences of a proliferative disease. The methods provided herein contemplate any one or more of these aspects of treatment. As above, the term treatment does not require 100 percent elimination of all aspects of the disorder.

[0116] "Ameliorate" means that one or more symptoms are alleviated or improved compared to when the anti-CCR8 antibody is not administered. "Ameliorate" also includes shortening or reducing the duration of the symptoms.

[0117] In the context of cancer, the term "treating" includes any or all of inhibiting cancer cell growth, inhibiting cancer cell replication, reducing overall tumor burden, and ameliorating one or more symptoms associated with the disease.

[0118] As used herein, the term "regulatory T cells" (also known as "Treg" or "Treg cells" or "suppressor T cells") refers to a subpopulation of T cells that are immunosuppressive and generally suppress or downregulate the induction and proliferation of effector T cells. Tregs express CD4, FOXP3, and CD25 (IL-2 receptor α chain). Human Foxp3+CD4+ T cells have been divided into three subfractions based on the expression levels of Foxp3 and the cell surface molecules CD25 and CD45RA. The Foxp3hiCD45RA-CD25hi and Foxp3loCD45RA+CD25lo phenotypes correspond to suppressive Treg cells, whereas the Foxp3loCD45RA-CD25lo fraction marks activated T effector (Teff) cells without suppressive activity. Furthermore, in healthy subjects, Compared with Treg cells from cancer patients, Treg cells from cancer patients are typically characterized by distinct expression profiles of chemokine receptors, such as CCR4, CXCR4, and CCR5, which promote migration to tumors in response to corresponding chemokine ligands derived from the tumor microenvironment (see, e.g., Liu, et al., FEBS J. (2016) 283(14):2731-48 and Miyara, et al., Immunity (2009) 30, 899-911).

[0119] "Conventional T cells" or "Tconv" are a population of T cells that are generally CD4 positive (i.e., CD4+), but are distinguishable from Tregs in that Tconv are generally FoxP3 negative (i.e., FoxP3-).

[0120] The term "biological sample" refers to a quantity of material from a living or formerly living thing, including, but not limited to, blood (e.g., whole blood), plasma, serum, urine, amniotic fluid, synovial fluid, endothelial cells, leukocytes, monocytes, other cells, organs, tissues, bone marrow, lymph nodes, and spleen.

[0121] The term "control" refers to a composition that is known to contain either no analyte (a "negative control") or to contain the analyte (a "positive control"). A positive control may contain a known concentration of analyte. "Control," "positive control," and "calibrator" may be used interchangeably herein to refer to a composition that contains a known concentration of analyte. A "positive control" can be used to establish assay performance characteristics and is a useful indicator of the integrity of the reagents (e.g., analyte).

[0122] "Predetermined cutoff" and "predetermined level" generally refer to assay cutoff values ​​used to evaluate diagnostic / prognostic / therapeutic efficacy results by comparing assay results to the predetermined cutoff / level, which have already been linked or associated with various clinical parameters (e.g., disease severity, progression / non-progression / improvement, etc.). While the present disclosure may provide exemplary predetermined levels, it is well known that cutoff values ​​may vary depending on the nature of the immunoassay (e.g., the antibody used, etc.). It is further within the skill of one of ordinary skill in the art to adapt the present disclosure herein to other immunoassays to obtain immunoassay-specific cutoff values ​​for those immunoassays based on the present disclosure. While the exact value of the predetermined cutoff / level may vary between assays, the correlations (if any) described herein may generally be applicable.

[0123] The term "inhibition" or "inhibiting" refers to the reduction or cessation of any phenotypic characteristic, or the reduction or cessation of the incidence, degree, or likelihood of that characteristic. "Decrease" or "inhibiting" refers to the reduction, decrease, or cessation of an activity, function, and / or amount compared to a reference. In some embodiments, "decrease" or "inhibiting" refers to the ability to cause an overall reduction of 20% or more. In some embodiments, "decrease" or "inhibiting" refers to the ability to cause an overall reduction of 50% or more. In some embodiments, "decrease" or "inhibiting" refers to the ability to cause an overall reduction of 75%, 85%, 90%, 95%, or more. In some embodiments, the above amounts are inhibited or reduced over a period of time relative to a control dose (such as a placebo) over the same period of time. "Reference," as used herein, refers to any sample, standard, or level used for comparison purposes. A reference can be obtained from a healthy and / or non-diseased sample. In some examples, a reference can be obtained from an untreated sample. In some examples, a reference is obtained from a non-diseased over untreated sample of the individual of interest. In some instances, the reference is obtained from one or more healthy individuals who are not the subject or patient.

[0124] As used herein, "delaying the onset of a disease" refers to deferring, hindering, slowing, retarding, stabilizing, inhibiting, and / or preventing the onset of a disease (such as cancer). "Delay" means to delay or postpone the progression of a cancer. This delay can be of varying lengths of time, depending on the disease being treated and / or the individual's medical history. As will be apparent to those skilled in the art, a sufficient or significant delay can, in effect, encompass prevention, in that the individual does not develop the disease. For example, late-stage cancer, such as the onset of metastases, can be delayed.

[0125] "Prevention," as used herein, includes providing protection against the onset or recurrence of a disease in a subject who is susceptible to the disease but has not yet been diagnosed with the disease. Unless otherwise specified, the terms "reducing," "inhibiting," or "preventing" do not indicate or require complete prevention all the time.

[0126] As used herein, "inhibiting" a function or activity refers to decreasing the function or activity as compared to the same conditions except for the condition or parameter of interest, or alternatively, as compared to another condition. For example, an antibody that inhibits tumor growth will decrease the rate of tumor growth as compared to the rate of tumor growth in the absence of the antibody.

[0127] A "therapeutically effective amount" of a substance / molecule, agonist, or antagonist can vary depending on factors such as the individual's medical condition, age, sex, and weight, and the ability of the substance / molecule, agonist, or antagonist to elicit a desired response in the individual. A therapeutically effective amount is also one in which any toxic or detrimental effects of the substance / molecule, agonist, or antagonist are outweighed by the therapeutically beneficial effects. A therapeutically effective amount can be delivered in one or more administrations. A "therapeutically effective amount" refers to an amount effective to achieve the desired therapeutic and / or prophylactic result, at the dosages and for the duration necessary.

[0128] A "prophylactically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired prophylactic result. Typically, although not necessarily, since a prophylactic dose is used in subjects prior to or at an earlier stage of disease, the prophylactically effective amount will be less than the therapeutically effective amount.

[0129] The terms "pharmaceutical formulation" and "pharmaceutical composition" refer to a preparation that is in a form that allows the biological activity of the active ingredient to be effective and that does not contain additional components that are unacceptably toxic to the subject to which the formulation is administered. Such formulations may be sterile.

[0130] A "pharmaceutically acceptable carrier" refers to a non-toxic solid, semi-solid, or liquid filler, diluent, encapsulating material, formulation aid, or carrier conventional in the art for use with therapeutic agents that together comprise a "pharmaceutical composition" for administration to a subject. A pharmaceutically acceptable carrier is non-toxic to recipients at the dosages and concentrations employed and is compatible with other ingredients of the formulation. A pharmaceutically acceptable carrier is appropriate for the formulation with which it is employed.

[0131] A "sterile" formulation is sterile or essentially free of living microorganisms and their spores.

[0132] The term "IDO inhibitor" refers to an agent that inhibits the activity of indoleamine 2,3-dioxygenase (IDO), thereby reversing IDO-mediated immunosuppression. IDO inhibitors can inhibit IDO1 and / or IDO2 (INDOL1). IDO inhibitors can be reversible or irreversible IDO inhibitors. A "reversible IDO inhibitor" is a compound that reversibly inhibits IDO enzyme activity at either the catalytic or non-catalytic site, while an "irreversible IDO inhibitor" is a compound that irreversibly inhibits IDO enzyme activity by forming a covalent bond with the enzyme. Non-limiting exemplary IDO inhibitors include Indoximod (New Link Genetics), INCB024360 (Incyte Corp.), 1-methyl-D-tryptophan (New Link Genetics), and GDC-0919 (Genentech, Inc.).

[0133] "Chimeric antigen receptor T-cell therapy" or "CAR-T therapy" refers to a therapeutic agent comprising T cells genetically engineered to express a receptor that recognizes an antigen expressed by tumor cells. The antigen can be an antigen specifically expressed by tumors or an antigen expressed by both cancerous cells and healthy tissue. In some embodiments, CAR-T therapy is adoptive CAR-T therapy, in which a patient's T cells are removed, modified to express a chimeric antigen receptor, and then returned to the patient. See, for example, Dai et al., 2016, J Natl Cancer Inst, 108(7):djv439, doi:10.1093 / jnci / djv439; Gill et al., 2015, Blood Rev, pii:S0268-960X(15)00080-6, doi:10.1016 / j.blre.2015.10.003; Gill et al., 2015, Immunol See Rev,263(1):68~89.doi:10.1111 / imr.12243.

[0134] Administration "in combination with" one or more further therapeutic agents includes simultaneous (concurrent) and consecutive or sequential administration in any order.

[0135] The term "concurrently" is used herein to refer to the administration of two or more therapeutic agents, where at least a portion of the administration overlaps in time, or the administration of one therapeutic agent is within a short period of time relative to the administration of another therapeutic agent, e.g., the two or more therapeutic agents are administered within a certain time interval of no more than a certain number of minutes.

[0136] The term "sequentially" is used herein to refer to the administration of two or more therapeutic agents, where the administration of one or more agents continues after the administration of one or more other agents is discontinued, or where the administration of one or more agents begins before the administration of one or more other agents, e.g., where the two or more therapeutic agents are administered at intervals of more than a certain number of minutes.

[0137] As used herein, "in conjunction with" refers to the administration of one therapeutic modality in addition to another therapeutic modality. Thus, "in conjunction with" refers to the administration of one therapeutic modality before, during, or after the administration of another therapeutic modality to an individual.

[0138] The term "package insert" is used to refer to instructions typically included in commercial packaging of a therapeutic product that contain information about the directions, uses, dosages, administration, concomitant therapies, contraindications, and / or warnings regarding the use of such therapeutic product.

[0139] An "article of manufacture" is any product (e.g., package or container) or kit that includes at least one reagent, e.g., an agent for the treatment of a disease or disorder (e.g., cancer), or a probe for specifically detecting a biomarker described herein. In some embodiments, the product or kit is promoted, distributed, or sold as a unit for performing a method described herein.

[0140] The terms "label" and "detectable label" refer to a moiety attached to an antibody or its analyte that renders the reaction (e.g., binding) between members of a specific binding pair detectable. A labeled member of a specific binding pair is said to be "detectably labeled." Thus, the term "labeled binding protein" refers to a protein into which a label has been incorporated that provides identification of the binding protein. In some embodiments, the label is a detectable marker that can produce a signal that is detectable by visual or instrumental means, for example, incorporation of a radiolabeled amino acid, or binding to a polypeptide that can be detected by marked avidin (e.g., streptavidin containing a fluorescent marker or enzymatic activity that can be detected optically or colorimetrically). Examples of labels for polypeptides include, but are not limited to, the following: radioisotopes or radionuclides (e.g., 3 H, 14 C. 35 S, 90 Y, 99 Tc, 111 In, 125I, 131 I, 177 Lu, 166 Ho, or 153 Sm), chromogens, fluorescent labels (e.g., FITC, rhodamine, lanthanide fluorophores), enzyme labels (e.g., horseradish peroxidase, luciferase, alkaline phosphatase), chemiluminescent markers, biotinyl groups, predetermined polypeptide epitopes recognized by secondary reporters (e.g., leucine zipper pair sequences, binding sites for secondary antibodies, metal binding domains, epitope tags), and magnetic agents such as gadolinium chelates. Representative examples of labels commonly used in immunoassays include light-producing moieties, e.g., acridinium compounds, and fluorescence-producing moieties, e.g., fluorescein. In this regard, the moiety itself need not be detectably labeled, but may become detectable upon reaction with yet another moiety.

[0141] The term "conjugate" refers to an antibody chemically linked to a second chemical moiety, such as a therapeutic or cytotoxic agent. The term "agent" includes a chemical compound, a mixture of chemical compounds, a biological macromolecule, or an extract made from biological materials. In some embodiments, the therapeutic or cytotoxic agent includes, but is not limited to, pertussis toxin, taxol, cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxyanthracin dione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, and puromycin, as well as analogs or homologs thereof. When used in the context of an immunoassay, the conjugated antibody can be a detectably labeled antibody used as a detection antibody. II. Anti-CCR8 antibody

[0142] Novel antibodies directed against CCR8 are provided. Anti-CCR8 antibodies include, but are not limited to, humanized antibodies, chimeric antibodies, murine antibodies, human antibodies, and antibodies comprising the heavy and / or light chain CDRs discussed herein. In some embodiments, isolated antibodies that bind to CCR8 are provided. In some embodiments, monoclonal antibodies that bind to CCR8 are provided. In some embodiments, the anti-CCR8 antibodies are antagonistic anti-CCR8 antibodies. In some embodiments, the anti-CCR8 antibodies provided herein inhibit the binding of CCR8 to CCL1. In some embodiments, administration of the anti-CCR8 antibodies described herein reduces infiltrating Treg cells in cancer in a subject. In some embodiments, administration of the anti-CCR8 antibodies described herein treats a hematological cancer that expresses CCR8.

[0143] In some embodiments, the anti-CCR8 antibody comprises at least one, two, three, four, five, or six CDRs selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 12, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 13, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 14, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 15, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 16, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 17.

[0144] In some embodiments, the anti-CCR8 antibody comprises at least one, two, three, four, five, or six CDRs selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 24, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 25, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 26, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 27, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 28, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 29.

[0145] In some embodiments, the anti-CCR8 antibody comprises at least one, two, three, four, five, or six CDRs selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 36, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 37, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 38, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 39, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 40, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 41.

[0146] In some embodiments, the anti-CCR8 antibody comprises at least one, two, three, four, five, or six CDRs selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 48, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 49, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 50, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 51, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 52, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 53.

[0147] In some embodiments, the anti-CCR8 antibody comprises at least one, two, three, four, five, or six CDRs selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65.

[0148] In some embodiments, the anti-CCR8 antibody comprises at least one, two, three, four, five, or six CDRs selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0149] In some embodiments, the anti-CCR8 antibody comprises a heavy chain variable region and a light chain variable region. In some embodiments, the anti-CCR8 antibody comprises at least one heavy chain comprising a heavy chain variable region and at least a portion of a heavy chain constant region, and at least one light chain comprising a light chain variable region and at least a portion of a light chain constant region. In some embodiments, the anti-CCR8 antibody comprises two heavy chains, each comprising a heavy chain variable region and at least a portion of a heavy chain constant region, and two light chains, each comprising a light chain variable region and at least a portion of a light chain constant region. As used herein, a single-chain Fv (scFv), or any other antibody comprising, for example, a single polypeptide chain comprising all six CDRs (three heavy chain CDRs and three light chain CDRs), is considered to have a heavy chain and a light chain. In some embodiments, the heavy chain is the region of the anti-CCR8 antibody comprising three heavy chain CDRs. In some embodiments, the light chain is the region of the anti-CCR8 antibody comprising three light chain CDRs.

[0150] In some embodiments, the anti-CCR8 antibody comprises six CDRs, including (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 12, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 13, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 14, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 15, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 16, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 17.

[0151] In some embodiments, the anti-CCR8 antibody comprises six CDRs, including (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 24, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 25, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 26, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 27, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 28, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 29.

[0152] In some embodiments, the anti-CCR8 antibody comprises six CDRs, including (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 36, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 37, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 38, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 39, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 40, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 41.

[0153] In some embodiments, the anti-CCR8 antibody comprises six CDRs, including (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 48, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 49, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 50, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 51, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 52, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 53.

[0154] In some embodiments, the anti-CCR8 antibody comprises six CDRs, including (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65.

[0155] In some embodiments, the anti-CCR8 antibody comprises six CDRs, including (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0156] In some embodiments, the anti-CCR8 antibody comprises the six CDRs described above and binds to CCR8. In some embodiments, the anti-CCR8 antibody comprises the six CDRs described above, binds to CCR8, and inhibits binding of CCR8 to CCL1. In some embodiments, the anti-CCR8 antibody comprises the six CDRs described above, binds to CCR8, and enhances an immune response in a subject and / or increases T cell activation in a subject after administration of the antibody to the subject.

[0157] In some embodiments, anti-CCR8 antibodies are provided that compete with the anti-CCR8 antibodies described herein for binding to CCR8. In some embodiments, antibodies can be made and / or used that compete for binding with any of the antibodies provided herein.

[0158] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VH CDR sequences selected from: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 12; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 13; and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 14.

[0159] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VH CDR sequences selected from: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 24; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 25; and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 26.

[0160] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VH CDR sequences selected from: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 36; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 37; and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 38.

[0161] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VH CDR sequences selected from: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 48; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 49; and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 50.

[0162] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VH CDR sequences selected from: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78; and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79.

[0163] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VH CDR sequences selected from: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85; and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86.

[0164] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VL CDR sequences selected from: (a) LCDR1 comprising the amino acid sequence of SEQ ID NO: 15; (b) LCDR2 comprising the amino acid sequence of SEQ ID NO: 16; and (c) LCDR3 comprising the amino acid sequence of SEQ ID NO: 17.

[0165] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VL CDR sequences selected from: (a) LCDR1 comprising the amino acid sequence of SEQ ID NO: 27; (b) LCDR2 comprising the amino acid sequence of SEQ ID NO: 28; and (c) LCDR3 comprising the amino acid sequence of SEQ ID NO: 29.

[0166] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VL CDR sequences selected from: (a) LCDR1 comprising the amino acid sequence of SEQ ID NO: 39; (b) LCDR2 comprising the amino acid sequence of SEQ ID NO: 40; and (c) LCDR3 comprising the amino acid sequence of SEQ ID NO: 41.

[0167] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VL CDR sequences selected from: (a) LCDR1 comprising the amino acid sequence of SEQ ID NO: 51; (b) LCDR2 comprising the amino acid sequence of SEQ ID NO: 52; and (c) LCDR3 comprising the amino acid sequence of SEQ ID NO: 53.

[0168] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VL CDR sequences selected from: (a) LCDR1 comprising the amino acid sequence of SEQ ID NO: 63; (b) LCDR2 comprising the amino acid sequence of SEQ ID NO: 64; and (c) LCDR3 comprising the amino acid sequence of SEQ ID NO: 65.

[0169] In some embodiments, the anti-CCR8 antibody comprises at least one, at least two, or all three VL CDR sequences selected from: (a) LCDR1 comprising the amino acid sequence of SEQ ID NO: 87; (b) LCDR2 comprising the amino acid sequence of SEQ ID NO: 88; and (c) LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0170] In some embodiments, any of the six CDRs provided herein can be partially combined with any of the other CDRs provided herein, for a total of six CDRs in the construct. Thus, in some embodiments, two CDRs (e.g., HCDR1 and HCDR2) from a first antibody can be combined with four CDRs (HCDR3, LCDR1, LCDR2, and LCDR3) from a second antibody. In some embodiments, no more than two residues in one or more of the CDRs can be replaced to obtain a variant thereof. In some embodiments, no more than two residues in one, two, three, four, five, or six of the CDRs can be replaced.

[0171] In some embodiments, the anti-CCR8 antibody comprises: (I) a VH domain comprising at least one, at least two, or all three VH CDR sequences selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 12, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 13, and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 14; and (II) a VL domain comprising at least one, at least two, or all three VL CDR sequences selected from (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 15, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 16, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 17.

[0172] In some embodiments, the anti-CCR8 antibody comprises: (I) a VH domain comprising at least one, at least two, or all three VH CDR sequences selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 24, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 25, and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 26; and (II) a VL domain comprising at least one, at least two, or all three VL CDR sequences selected from (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 27, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 28, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 29.

[0173] In some embodiments, the anti-CCR8 antibody comprises: (I) a VH domain comprising at least one, at least two, or all three VH CDR sequences selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 36, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 37, and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 38; and (II) a VL domain comprising at least one, at least two, or all three VL CDR sequences selected from (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 39, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 40, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 41.

[0174] In some embodiments, the anti-CCR8 antibody comprises: (I) a VH domain comprising at least one, at least two, or all three VH CDR sequences selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 48, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 49, and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 50; and (II) a VL domain comprising at least one, at least two, or all three VL CDR sequences selected from (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 51, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 52, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 53.

[0175] In some embodiments, the anti-CCR8 antibody comprises: (I) a VH domain comprising at least one, at least two, or all three VH CDR sequences selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78; and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79; and (II) at least one, at least two, or all three VL CDR sequences selected from (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65. It contains a VL domain that includes the CDR sequences.

[0176] In some embodiments, the anti-CCR8 antibody comprises: (I) a VH domain comprising at least one, at least two, or all three VH CDR sequences selected from (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85, and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86; and (II) a VL domain comprising at least one, at least two, or all three VL CDR sequences selected from (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0177] In some embodiments, an anti-CCR8 antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 68 or 74. In some embodiments, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-CCR8 antibody comprising that sequence retains the ability to bind to CCR8. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 68 or 74. In some embodiments, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). Optionally, the anti-CCR8 antibody comprises the VH sequence of SEQ ID NO: 68 or 74, including post-translational modifications of that sequence.

[0178] In some embodiments, the VH comprises (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78, and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79.

[0179] In some embodiments, an anti-CCR8 antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 92 or 96. In some embodiments, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-CCR8 antibody comprising that sequence retains the ability to bind to CCR8. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 92 or 96. In some embodiments, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). Optionally, the anti-CCR8 antibody comprises the VH sequence of SEQ ID NO: 92 or 96, including post-translational modifications of that sequence.

[0180] In some embodiments, the VH comprises (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85, and (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86.

[0181] In some embodiments, an anti-CCR8 antibody comprises a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 69 or 75. In some embodiments, a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-CCR8 antibody comprising that sequence retains the ability to bind to CCR8. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 69 or 75. In some embodiments, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). Optionally, the anti-CCR8 antibody comprises the VL sequence of SEQ ID NO: 69 or 75, including post-translational modifications of that sequence.

[0182] In some embodiments, the VL comprises (a) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, (b) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and (c) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65.

[0183] In some embodiments, an anti-CCR8 antibody comprises a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 93 or 97. In some embodiments, a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-CCR8 antibody comprising that sequence retains the ability to bind to CCR8. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 93 or 97. In some embodiments, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). Optionally, the anti-CCR8 antibody comprises the VL sequence of SEQ ID NO: 93 or 97, including post-translational modifications of that sequence.

[0184] In some embodiments, the VL comprises (a) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, (b) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and (c) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0185] In some embodiments, the anti-CCR8 antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 68 or 74, and a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 69 or 75. In some embodiments, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, and a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-CCR8 antibody comprising the sequence retains the ability to bind to CCR8. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 68 or 74. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 69 or 75. In some embodiments, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). In some embodiments, the anti-CCR8 antibody comprises (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65.

[0186] In some embodiments, the anti-CCR8 antibody comprises a VH sequence of SEQ ID NO: 68 or 74, including post-translational modifications of one or both sequences, and a VL sequence of SEQ ID NO: 69 or 75, including post-translational modifications of one or both sequences.

[0187] In some embodiments, the anti-CCR8 antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 92 or 96, and a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 93 or 97. In some embodiments, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, and a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-CCR8 antibody comprising the sequence retains the ability to bind to CCR8. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 92 or 96. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) are substituted, inserted, and / or deleted in SEQ ID NO: 93 or 97. In some embodiments, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). In some embodiments, the anti-CCR8 antibody comprises (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0188] In some embodiments, the anti-CCR8 antibody comprises a VH sequence of SEQ ID NO: 92 or 96, including post-translational modifications of one or both sequences, and a VL sequence of SEQ ID NO: 93 or 97, including post-translational modifications of one or both sequences.

[0189] In some embodiments, the anti-CCR8 antibody comprises a VH as in any of the embodiments provided herein, and a VL as in any of the embodiments provided herein. In some embodiments, the antibody comprises the VH and VL sequences of SEQ ID NO: 68 or 74 and SEQ ID NO: 69 or 75, respectively, including post-translational modifications of those sequences. In some embodiments, the antibody comprises the VH and VL sequences of SEQ ID NO: 92 or 96 and SEQ ID NO: 93 or 97, respectively, including post-translational modifications of those sequences.

[0190] In some embodiments, the anti-CCR8 antibody comprises a heavy chain (HC) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of No. 70 or 76. Optionally, the anti-CCR8 antibody comprises the HC sequence of No. 70 or 76 including post-translational modifications.

[0191] In some embodiments, the anti-CCR8 antibody comprises an HC having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of No. 94 or 98. Optionally, the anti-CCR8 antibody comprises the HC sequence of No. 94 or 98 including a post-translational modification.

[0192] In some embodiments, the anti-CCR8 antibody comprises a light chain (LC) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of No. 71 or 77. Optionally, the anti-CCR8 antibody comprises the LC sequence of No. 71 or 77 including post-translational modifications.

[0193] In some embodiments, the anti-CCR8 antibody comprises an LC having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of No. 95 or 99. Optionally, the anti-CCR8 antibody comprises the LC sequence of No. 95 or 99 including a post-translational modification.

[0194] In some embodiments, the anti-CCR8 antibody comprises an HC as in any of the embodiments provided herein, and an LC as in any of the embodiments provided herein. In some embodiments, the antibody comprises the HC and LC sequences of SEQ ID NO: 70 or 76 and SEQ ID NO: 71 or 77, respectively, including post-translational modifications of those sequences. In some embodiments, the antibody comprises the HC and LC sequences of SEQ ID NO: 94 or 98 and SEQ ID NO: 95 or 99, respectively, including post-translational modifications of those sequences.

[0195] In some embodiments, anti-CCR8 antibodies are provided that compete with the anti-CCR8 antibodies described herein for binding to CCR8, hi some embodiments, the antibodies compete with the anti-CCR8 antibodies provided herein for binding to an epitope on CCR8.

[0196] In some embodiments, a competition assay can be used to identify monoclonal antibodies that compete with the anti-CCR8 antibodies described herein (e.g., 1-K16, 1-K17, 6-B09, 7-B16, 13-E16, and / or 19-O07) for binding to CCR8. A competition assay can be used to determine whether two antibodies bind to the same epitope by recognizing the same or sterically overlapping epitope, or by one antibody competitively inhibiting the binding of another antibody to an antigen. In some embodiments, such competing antibodies bind to the same epitope bound by an antibody described herein. Exemplary competition assays include, but are not limited to, routine assays such as those provided in Harlow and Lane (1988) Antibodies: A Laboratory Manual ch. 14 (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY). Detailed exemplary methods for mapping antibody-binding epitopes are provided in Morris (1996) "Epitope Mapping Protocols," in Methods in Molecular Biology, vol. 66 (Humana Press, Totowa, NJ). In some embodiments, two antibodies are said to bind to the same epitope if each blocks 50% or more of the binding of the other. In some embodiments, antibodies that compete with the anti-CCR8 antibodies described herein are chimeric, humanized, or human antibodies. In some embodiments, antibodies that compete with the chimeric, humanized, or human anti-CCR8 antibodies described herein are provided.

[0197] Additionally, the present disclosure also provides variants of the aforementioned disclosed antibodies, such as variants of the 7-B16 antibody. For example, in some embodiments, the present disclosure provides an isolated antibody that binds to human CCR8, wherein the antibody comprises an HCDR3 comprising SEQ ID NO: 86 or a variant of SEQ ID NO: 86 comprising one, two, or three mutations, and the antibody binds to human CCR8 and has ADCC activity. In some embodiments, the mutations are substitutions (e.g., conservative or non-conservative substitutions), deletions, or insertions. In some embodiments, the one, two, or three mutations are located at at least one of amino acids 1-4, 6, 7, or 12 of SEQ ID NO: 86. In some embodiments, the substitutions are conservative substitutions. In some embodiments, the conservative substitution is at amino acid position 1, 4, or 12 of SEQ ID NO: 86. In some embodiments, the substitution is a non-conservative substitution. In some embodiments, the non-conservative substitution is at amino acid position 7 of SEQ ID NO: 86. In some embodiments, the antibody comprises at least two substitutions in HCDR3. In some embodiments, the at least two substitutions are located at at least one of amino acids 1-4, 6, 7, or 12 of SEQ ID NO: 86. In some embodiments, the at least two substitutions are conservative substitutions. In some embodiments, at least one of the conservative substitutions is at amino acid position 1, 4, or 12 of SEQ ID NO: 86. In some embodiments, the at least two substitutions are non-conservative substitutions. In some embodiments, at least one of the non-conservative substitutions is at amino acid position 7 of SEQ ID NO: 86. In some embodiments, the mutations comprise a conservative substitution and a non-conservative substitution when more than one substitution mutation is present. In some embodiments, the disclosure provides an isolated antibody that binds to human CCR8, wherein the antibody comprises an HCDR3 that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 86, and wherein the antibody binds to human CCR8 and has ADCC activity. In some embodiments, the HCDR3 comprises an amino acid sequence selected from SEQ ID NO: 86 and any one of SEQ ID NOs: 104 to 119. In some embodiments, the antibody comprises an HCDR1 comprising SEQ ID NO: 84 or SEQ ID NO: 123.In some embodiments, the antibody comprises an HCDR2 comprising SEQ ID NO: 85 or SEQ ID NO: 124. In some embodiments, the antibody comprises an LCDR1 comprising SEQ ID NO: 87 or SEQ ID NO: 120. In some embodiments, the antibody comprises an LCDR2 comprising SEQ ID NO: 88 or SEQ ID NO: 121. In some embodiments, the antibody comprises an LCDR3 comprising SEQ ID NO: 89 or SEQ ID NO: 122. In some embodiments, the ADCC activity comprises an EC50 value of less than 200, 175, 150, 125, 100, 75, 50, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 ng / ml, as measured by an ADCC reporter mechanism of action (MOA)-based bioassay, such as the Promega reporter assay disclosed in Example 7 below. In some embodiments, the ADCC activity is more potent than the 7-B16 antibody. In some embodiments, the ADCC activity is at least as potent as the 7-B16 antibody. In some embodiments, the antibody has a K for human CCR8 that is equal to or less than that of the 7-B16 antibody (e.g., as determined by an equilibrium binding exclusion assay (i.e., KinExA)). D In some embodiments, the antibody has an on-cell K for human CCR8 that is equal to or less than that of the 7-B16 antibody (e.g., as determined by a binding equilibrium exclusion assay (i.e., KinExA)). DIn some embodiments, the antibody comprises at least one modification that enhances cell killing. In some embodiments, the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). In some embodiments, the at least one modification is afucosylation. In some embodiments, the at least one modification is one or more heavy chain constant region mutations at one or more positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396. In some embodiments, the one or more heavy chain constant region mutations are one or more mutations selected from S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L. In some embodiments, the one or more heavy chain constant region mutations are selected from F243L / R292P / Y300L / V305I / P396L, S239D / I332E, S239D / I332E / A330L, S298A / E333A / K334A, L234Y / L235Q / G236W / S239M / H268D / D270E / S298A, and D270E / K326D / A330M / K334E. In some embodiments, at least one modification is galactosylation. In some embodiments, the antibody has a binding affinity of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75 pM, or less than 50 pM, or less than 25 pM (e.g., as measured by a Kinetic Exclusion Assay (i.e., affinity (K) as determined by KinExA DIn some embodiments, the antibody binds to human CCR8 with an on-cell affinity (K) of less than 10 nM, or less than 5 nM, or less than 1 nM, or less than 500 pM, or less than 250 pM, or less than 100 pM, or less than 75, or less than 50 pM, or less than 25 pM, as determined, for example, by an equilibrium binding exclusion assay (i.e., KinExA). D ) binds to human CCR8. In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is a human antibody or a humanized antibody. In some embodiments, the antibody is a full-length antibody. In some embodiments, the antibody is an IgG1 or IgG3 antibody. Such variants can be used in methods of treating cancer, including both hematological cancers and solid tumors.

[0198] In some embodiments, antibodies are provided that bind to any one or more of the epitopes provided herein. In some embodiments, antibodies are provided that bind to an epitope that overlaps with that of the present antibodies. In some embodiments, antibodies are provided that compete with at least one of the antibodies provided herein. In some embodiments, antibodies are provided that compete with at least two of the antibodies provided herein. In some embodiments, antibodies are provided that compete with at least three of the antibodies provided herein. In some embodiments, the antibody binds to an overlapping epitope as an antibody described in the Examples herein. In some embodiments, the entire epitope is bound and / or blocked by the competing antibody. In some embodiments, only a portion of the epitope is bound and / or blocked by the competing antibody. In some embodiments, the paratope of the competing antibody binds to at least a portion of the epitope of the antibody provided herein. In some embodiments, the paratope of the competing antibody binds to the target, and a different section of the competing antibody's structure blocks at least a portion of the epitope of the antibody provided herein. Exemplary Chimeric Antibodies

[0199] In some embodiments, the antibodies provided herein are chimeric antibodies. Certain chimeric antibodies are described, for example, in U.S. Patent No. 4,816,567 and Morrison et al., (1984) Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984). In one example, a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non-human primate such as a monkey) and a human constant region. In a further example, a chimeric antibody is a "class-switched" antibody in which the class or subclass has been changed from that of the parent antibody. Chimeric antibodies include antigen-binding fragments thereof.

[0200] Non-limiting exemplary chimeric antibodies include those comprising the heavy and / or light chain variable regions of an antibody selected from antibodies 1-K16, 1-K17, 6-B09, 7-B16, 13-E16, and 19-O07, as disclosed herein. Additional non-limiting exemplary chimeric antibodies include those comprising the heavy chain CDR1, CDR2, and CDR3 and / or the light chain CDR1, CDR2, and CDR3 of an antibody selected from antibodies 1-K16, 1-K17, 6-B09, 7-B16, 13-E16, and 19-O07, as disclosed herein. In some embodiments, a chimeric anti-CCR8 antibody comprises the variable regions described above and binds to CCR8. In some embodiments, a chimeric anti-CCR8 antibody comprises the variable regions described above and binds to CCR8 and inhibits binding of CCR8 to CCL1. In some embodiments, the anti-CCR8 antibody comprises a variable region described above, binds to CCR8, enhances an immune response in a subject, and / or increases T cell activation in a subject after administration of the antibody to the subject. In some embodiments, administration of an anti-CCR8 antibody described herein stimulates immune cell activity, reduces immune cell downregulation, or increases T cell responses in a subject.

[0201] In some embodiments, the chimeric antibodies described herein comprise one or more human constant regions. In some embodiments, the human heavy chain constant region is of an isotype selected from IgA, IgG, IgD, and IgE. In some embodiments, the human light chain constant region is of an isotype selected from Kappa and Lambda. In some embodiments, the chimeric antibodies described herein comprise a human IgG constant region. In some embodiments, the chimeric antibodies described herein comprise a human IgG4 heavy chain constant region. In some embodiments, the chimeric antibodies described herein comprise a human IgG4 constant region and a human Kappa light chain.

[0202] As noted above, whether effector function is desirable can depend on the particular therapeutic method for which the antibody is intended. Thus, in some embodiments, if effector function is desirable, a chimeric anti-CCR8 antibody comprising a human IgG1 heavy chain constant region or a human IgG3 heavy chain constant region is selected. In some embodiments, if effector function is undesirable, a chimeric anti-CCR8 antibody comprising a human IgG4 or IgG2 heavy chain constant region is selected. In some embodiments, enhanced effector function is desirable. Exemplary Humanized Antibodies

[0203] In some embodiments, humanized antibodies that bind to CCR8 are provided. Immune responses to antibody therapeutics (such as human anti-mouse antibody (HAMA) responses) Humanized antibodies are useful as therapeutic molecules because they reduce or eliminate the human immune response compared to non-human antibodies, which can result in adverse reactions (e.g., inflammatory bowel disease, ulcers, and ulcers) and reduced therapeutic efficacy.

[0204] In some embodiments, a chimeric antibody is a humanized antibody. Typically, a non-human antibody is humanized to reduce immunogenicity to humans while retaining the specificity and affinity of the parent non-human antibody. Generally, a humanized antibody comprises one or more variable domains in which the CDRs (or portions thereof) are derived from a non-human antibody and the FRs (or portions thereof) are derived from human antibody sequences. A humanized antibody will optionally also comprise at least a portion of a human constant region. In some embodiments, some FR residues in a humanized antibody are substituted with corresponding residues from the non-human antibody (e.g., the antibody from which the CDR residues are derived), e.g., to restore or improve antibody specificity or affinity.

[0205] Humanized antibodies and methods for making them are reviewed, for example, in Almagro and Fransson, (2008) Front. Biosci. 13:1619-1633, and further described, for example, in Riechmann et al., (1988) Nature 332:323-329; Queen et al., (1989) Proc. Natl. Acad. Sci. USA 86:10029-10033; U.S. Patent Nos. 5,821,337, 7,527,791, 6,982,321, and 7,087,409; Kashmiri et al., (2005) Methods 36:25-34; Padlan, (1991) Mol. Immunol. 28:489-498 (describing "resurfacing"); Dall'Acqua et al., (2005) Methods 36:43-60 (describing "FR shuffling"); and Osbourn et al., (2005) Methods 36:61-68 and Klimka et al., (2000) Br. J. Cancer, 83:252-260 (describing a "guided selection" approach to FR shuffling).

[0206] Human framework regions that can be used for humanization include framework regions selected using "best fit" methods (see, e.g., Sims et al. (1993) J. Immunol. 151:2296), framework regions derived from consensus sequences of human antibodies of particular subgroups of light or heavy chain variable regions (see, e.g., Carter et al. (1992) Proc. Natl. Acad. Sci. USA, 89:4285 and Presta et al. (1993) J. Immunol, 151:2623), human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, (2008) Front. Biosci. 13:1619-1633), and framework regions derived from screening FR libraries (see, e.g., Baca et al., (1997) J. Biol. Chem. 272:10678-10684 and Rosok et al., (1996) J. Biol. Chem. 271:22611-22618).

[0207] In some embodiments, the humanized anti-CCR8 antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 68 or 74, and a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 69 or 75. In some embodiments, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, and a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-CCR8 antibody comprising the sequence retains the ability to bind to CCR8. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 68 or 74. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) are substituted, inserted, and / or deleted in SEQ ID NO: 69 or 75. In some embodiments, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). In some embodiments, the anti-CCR8 antibody comprises (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 60, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 61, 72, or 78, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 62, 73, or 79, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 65. In some embodiments, the antibody has ADCC activity.In some embodiments, the ADCC activity comprises an EC50 value of less than 200, 175, 150, 125, 100, 75, 50, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 ng / ml as measured by a mechanism of action (MOA)-based bioassay of an ADCC reporter. In some embodiments, the ADCC activity is more potent than the 7-B16 antibody. In some embodiments, the ADCC activity is at least as potent as the 7-B16 antibody. In some embodiments, the antibody has a K for human CCR8 that is equal to or less than the 7-B16 antibody (e.g., as determined by equilibrium binding displacement assay (i.e., KinExA)). D In some embodiments, the antibody has an on-cell K for human CCR8 that is equal to or less than that of the 7-B16 antibody (e.g., as determined by a binding equilibrium exclusion assay (i.e., KinExA)). D It has.

[0208] In some embodiments, the humanized anti-CCR8 antibody comprises a VH sequence of SEQ ID NO: 68 or 74, including post-translational modifications of one or both sequences, and a VL sequence of SEQ ID NO: 69 or 75, including post-translational modifications of one or both sequences.

[0209] In some embodiments, the humanized anti-CCR8 antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 92 or 96, and a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 93 or 97. In some embodiments, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, and a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-CCR8 antibody comprising the sequence retains the ability to bind to CCR8. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) have been substituted, inserted, and / or deleted in SEQ ID NO: 92 or 96. In some embodiments, a total of 1 to 10 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) are substituted, inserted, and / or deleted in SEQ ID NO: 93 or 97. In some embodiments, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). In some embodiments, the anti-CCR8 antibody comprises (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 84 or 100, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 85, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 86, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 89.

[0210] In some embodiments, the humanized anti-CCR8 antibody comprises a VH sequence of SEQ ID NO: 92 or 96, including post-translational modifications of one or both sequences, and a VL sequence of SEQ ID NO: 93 or 97, including post-translational modifications of one or both sequences.

[0211] Exemplary humanized anti-CCR8 antibodies include antibodies that compete for binding to CCR8 with the antibodies or fragments thereof described herein. Thus, in some embodiments, a humanized anti-CCR8 antibody is provided that competes for binding to CCR8 with an antibody selected from antibodies 1-K16, 1-K17, 6-B09, 7-B16, 13-E16, and 19-O07, or a fragment thereof. In some embodiments, the humanized anti-CCR8 antibody competes for binding to CCR8 with an antibody described herein and inhibits binding of CCR8 to CCL1. In some embodiments, the humanized anti-CCR8 antibody competes for binding to CCR8 with an antibody described herein. Exemplary Human Antibodies

[0212] In some embodiments, the anti-CCR8 antibodies provided herein are human antibodies. Human antibodies can be produced using various techniques known in the art. Human antibodies are generally described in van Dijk and van de Winkel, (2001) Curr. Opin. Pharmacol. 5:368-374 and Lonberg, (2008) Curr. Opin. Immunol. 20:450-459. In some embodiments, the human antibodies are not naturally occurring antibodies. In some embodiments, the human antibodies are monoclonal antibodies; thus, in some embodiments, each of the human antibodies in the set can bind to the same epitope on the antigen.

[0213] Human antibodies can be prepared by administering an immunogen to transgenic animals that have been modified to produce intact human antibodies or intact antibodies with human variable regions in response to antigen challenge. Such animals contain all or part of human immunoglobulin loci that replace endogenous immunoglobulin loci or that are present extrachromosomally or randomly integrated into the animal's chromosomes. In such transgenic mice, endogenous immunoglobulin loci are generally inactivated. For a review of methods for obtaining human antibodies from transgenic animals, see Lonberg, (2005) Nat. Biotech. 23:1117-1125. See also, e.g., U.S. Patent Nos. 6,075,181 and 6,150,584, which describe XENOMOUSE™ technology, U.S. Patent No. 5,770,429, which describes HUMAB® technology, U.S. Patent No. 7,041,870, which describes KM MOUSE® technology, and U.S. Patent Application Publication No. 2007 / 0061900, which describes VELOCIMOUSE® technology. The human variable regions from intact antibodies produced by such animals can be further modified, for example, by combining them with different human constant regions.

[0214] Human antibodies can also be produced by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for the production of human monoclonal antibodies have been described. (See, for example, Kozbor (1984) J. Immunol., 133:3001; Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp. 51-63 (Marcel Dekker, Inc., New York, 1987); and Boerner et al. (1991) J. Immunol., 147:86.) Human antibodies generated via human B cell hybridoma technology are also described in Li et al. (2006) Proc. Natl. Acad. Sci. USA, 103:3557-3562. Additional methods include those described, for example, in U.S. Patent No. 7,189,826 (which describes the production of monoclonal human IgM antibodies from hybridoma cell lines) and Ni, (2006) Xiandai Mianyixue, 26(4):265-268 (which describes human-human hybridomas). Human hybridoma technology (Trioma technology) is also described in Vollmers and Brandlein, (2005) Histology and Histopathology, 20(3):927-937 (2005) and Vollmers and Brandlein, (2005) Methods and Findings in Experimental and Clinical Pharmacology, 27(3):185-191.

[0215] Human antibodies can also be generated by isolating Fv clone variable domain sequences selected from human-derived phage display libraries. Such variable domain sequences can be combined with desired human constant domains. Techniques for selecting human antibodies from antibody libraries are described below.

[0216] Antibodies can be isolated by screening combinatorial libraries for antibodies with the desired activity(ies). For example, various methods are known in the art for generating phage display libraries and screening such libraries for antibodies with the desired binding characteristics. Such methods are reviewed, for example, in Hoogenboom et al. in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, NJ, 2001), and further described, for example, in McCafferty et al. (1990) Nature 348:552-554, Clackson et al. (1991) Nature 352:624-628, Marks et al. (1992) J. Mol. Biol. 222:581-597, Marks and Bradbury, in Methods in Molecular Biology 248:161-175 (Lo, ed., Human Press, Totowa, NJ, 2003), Sidhu et al. (2004) J. Mol. Biol. 338(2):299-310, Lee et al. al., (2004) J. Mol. Biol. 340(5):1073-1093, Fellouse, (2004) Proc. Natl. Acad. Sci. USA 101(34):12467-12472 and Lee et al., (2004) J. Immunol. Methods 284(1-2):119-132 and PCT Publication WO 99 / 10494.

[0217] In certain phage display methods, V H and V LGene repertoires can be separately cloned by polymerase chain reaction (PCR), randomly recombined in phage libraries, and then screened for antigen-binding phage as described by Winter et al. (1994) Ann. Rev. Immunol., 12:335-455. Phage typically display antibody fragments as either single-chain Fv (scFv) fragments or Fab fragments. Libraries from immunized sources provide high-affinity antibodies to immunogens without the need to construct hybridomas. Alternatively, naive repertoires can be cloned (e.g., from humans) to provide a single source of antibodies to a wide range of non-self and self antigens without any immunization, as described by Griffiths et al. (1993) EMBO J 12:725-734. Finally, naive libraries can be generated synthetically by cloning unrearranged V gene segments from stem cells and using PCR primers containing random sequences to encode the highly variable CDR3 regions and achieve rearrangement in vitro, as described by Hoogenboom and Winter (1992), J. Mol. Biol. 227:381-388. Patent publications describing human antibody phage libraries include, for example, U.S. Pat. No. 5,750,373, and U.S. Patent Application Publication Nos. 2005 / 0079574, 2005 / 0119455, 2005 / 0266000, 2007 / 0117126, 2007 / 0160598, 2007 / 0237764, 2007 / 0292936, and 2009 / 0002360.

[0218] In some embodiments, the human anti-CCR8 antibody binds to CCR8 and inhibits binding of CCR8 to CCL1.

[0219] Exemplary human anti-CCR8 antibodies also include antibodies that compete for binding to CCR8 with the human antibodies or fragments thereof described herein. Thus, in some embodiments, human anti-CCR8 antibodies are provided that compete for binding to CCR8 with an antibody selected from antibodies 1-K16, 1-K17, 6-B09, 7-B16, 13-E16, and 19-O07, or a fragment thereof. In some embodiments, the human anti-CCR8 antibody competes with an antibody described herein for binding to CCR8 and inhibits CCR8 binding to CCL1.

[0220] In some embodiments, chimeric human anti-CCR8 antibodies are provided, wherein the antibody comprises a variable region from a human antibody that binds to CCR8 and a constant region from a different human antibody. In some embodiments, chimeric human anti-CCR8 antibodies are provided, wherein the antibody comprises CDRs from a human antibody that binds to CCR8 and a framework region from a different human antibody. In some embodiments, the antibody is not a naturally occurring human antibody.

[0221] In some embodiments, a human anti-CCR8 antibody comprises one or more human constant regions. In some embodiments, the human heavy chain constant region is of an isotype selected from IgA, IgG, IgD, and IgE. In some embodiments, the human light chain constant region is of an isotype selected from kappa and lambda. In some embodiments, a human antibody described herein comprises a human IgG constant region. In some embodiments, a human antibody described herein comprises a human IgG4 heavy chain constant region. In some embodiments, a human antibody described herein comprises a human IgG4 constant region and a human kappa light chain.

[0222] In some embodiments, when effector function is desired, a human anti-CCR8 antibody comprising a human IgG1 heavy chain constant region or a human IgG3 heavy chain constant region is selected. In some embodiments, when effector function is undesired, a human anti-CCR8 antibody comprising a human IgG4 or IgG2 heavy chain constant region is selected.

[0223] As used herein, the term "human antibody" refers to the genus of possible sequences for antibody construction, not the source of the antibody. II. Fusion Proteins

[0224] In some embodiments, the fusion proteins described herein comprise CCL1 or an active fragment of CCL1. An "active fragment" of CCL1, as used herein, refers to a fragment of CCL1 that binds to CCR8. One of skill in the art can identify an active fragment of CCL1 that binds to CCR8. In some embodiments, an active fragment of CCL1 binds to CCR8 with an affinity that is at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% stronger than the affinity of native CCL1 for CCR8. For the avoidance of doubt, an affinity of 1 nM and an affinity that is at least 50% stronger are 2 nM or less. One of skill in the art can determine the affinity of an active fragment of CCL1 for CCR8 and compare it to the affinity of native CCL1 for CCR8. In some embodiments, the fusion proteins described herein comprise SEQ ID NO:2 or amino acids 24-96 of SEQ ID NO:2.

[0225] In some embodiments, a fusion protein comprising CCL1 or an active fragment thereof comprises an Fc region, such as an Fc region provided herein. In some embodiments, the Fc region comprises at least one modification that enhances cell killing. In some such embodiments, the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). Non-limiting exemplary modifications that enhance cell killing are provided herein and include, for example, afucosylation.

[0226] In some embodiments, the fusion proteins described herein comprise MC148 or an active fragment of MC148. As used herein, "active fragment" of MC148 refers to a fragment of MC148 that binds to CCR8. One of skill in the art can identify an active fragment of MC148 that binds to CCR8. In some embodiments, an active fragment of MC148 binds to CCR8 with an affinity that is at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% stronger than the affinity of native MC148 for CCR8. For the avoidance of doubt, an affinity of 1 nM and an affinity that is at least 50% stronger are 2 nM or less. One of skill in the art can determine the affinity of an active fragment of MC148 for CCR8 and compare it to the affinity of native MC148 for CCR8. In some embodiments, the fusion proteins described herein comprise SEQ ID NO:2 or amino acids 24-96 of SEQ ID NO:2.

[0227] In some embodiments, a fusion protein comprising MC148 or an active fragment thereof comprises an Fc region, such as an Fc region provided herein. In some embodiments, the Fc region comprises at least one modification that enhances cell killing. In some such embodiments, the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). Non-limiting exemplary modifications that enhance cell killing are provided herein and include, for example, afucosylation. III. Exemplary Antibody Constant Regions and Fc Regions

[0228] In some embodiments, the antibodies described herein comprise one or more human constant regions. In some embodiments, the human heavy chain constant region is of an isotype selected from IgA, IgG, IgD, and IgE. In some embodiments, the antibodies described herein comprise a human IgG constant region. In some embodiments, when effector function is desired, an anti-CCR8 antibody comprising a human IgG1 heavy chain constant region or a human IgG3 heavy chain constant region is selected. In some embodiments, when effector function is undesired, an anti-CCR8 antibody comprising a human IgG4 or IgG2 heavy chain constant region is selected. In some embodiments, the human light chain constant region is of an isotype selected from kappa and lambda. In some embodiments, the antibodies described herein comprise a human IgG1 heavy chain constant region. In some embodiments, the antibodies described herein comprise a human IgG1 constant region and a human kappa light chain.

[0229] In some embodiments, the fusion proteins described herein comprise one or more human Fc regions. In some embodiments, the Fc region is of an isotype selected from IgA, IgG, IgD, and IgE. In some embodiments, the fusion proteins described herein comprise a human Fc region. In some embodiments, if effector function is desired, a fusion protein comprising a human IgG1 Fc region or a human IgG3 Fc region is selected. In some embodiments, if effector function is not desired, a fusion protein comprising a human IgG4 or IgG2 Fc region is selected.

[0230] Throughout this specification and claims, unless expressly stated or known to one of skill in the art, the numbering of residues in an immunoglobulin heavy chain is that of the EU index as in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991), expressly incorporated herein by reference. "EU index as in Kabat" refers to the residue numbering of the human IgG1 EU antibody.

[0231] As noted above, whether effector function is desirable can depend on the particular therapeutic method for which the antibody is intended. Thus, in some embodiments, if effector function is desirable, an anti-CCR8 antibody comprising a human IgG1 heavy chain constant region or a human IgG3 heavy chain constant region is selected. In some embodiments, if effector function is desirable, a fusion protein comprising a human IgG1 Fc region or a human IgG3 Fc region is selected.

[0232] In some embodiments, the antibody or fusion protein comprises a variant Fc region having at least one amino acid substitution compared to the Fc region of a wild-type IgG Fc region. In some embodiments, the variant Fc region has two or more amino acid substitutions compared to the wild-type Fc region. In some embodiments, the variant Fc region has three or more amino acid substitutions compared to the wild-type Fc region. In some embodiments, the variant Fc region has at least one, two, or three or more Fc region amino acid substitutions described herein. In some embodiments, the variant Fc region herein will have at least about 80% homology to a native sequence Fc region and / or the Fc region of the parent polypeptide. In some embodiments, the variant Fc region herein will have at least about 90% homology to a native sequence Fc region and / or the Fc region of the parent polypeptide. In some embodiments, the variant Fc region herein will have at least about 95% homology to a native sequence Fc region and / or the Fc region of the parent polypeptide. In some embodiments, the heavy chain constant region or Fc region lacks a C-terminal lysine (K) residue. In some such embodiments, the heavy chain constant region or Fc region may be referred to as "desK". In some embodiments, the heavy chain constant region or Fc region lacking a C-terminal lysine is an IgG, such as IgG1, IgG2, IgG3, or IgG4.

[0233] In some embodiments, the antibodies or fusion proteins provided herein are modified to increase or decrease the extent to which the antibody is glycosylated. Addition or deletion of glycosylation sites to an antibody can be conveniently accomplished by altering the amino acid sequence so that one or more glycosylation sites are created or removed.

[0234] The carbohydrate attached to the Fc region can be modified. Natural antibodies produced by mammalian cells typically contain branched, biantennary oligosaccharides that are generally attached by an N-linkage to Asn297 in the CH2 domain of the Fc region. See, e.g., Wright et al., TIBTECH 15:26-32 (1997). The oligosaccharides can contain various carbohydrates, such as mannose, N-acetylglucosamine (GlcNAc), galactose, and sialic acid, as well as fucose attached to the GlcNAc in the "stem" of the biantennary oligosaccharide structure. In some embodiments, modifications of the oligosaccharides in an antibody or fusion protein can be performed to generate antibody variants with specific improved properties.

[0235] In some embodiments, antibody or fusion protein variants are provided that have carbohydrate structures lacking fucose attached (directly or indirectly) to the Fc region (i.e., afucosylation). For example, the amount of fucose in such variants can 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 compared to the sum of all glycans (e.g., complex, hybrid, and high-mannose structures) attached to Asn297, as measured by MALDI-TOF mass spectrometry, e.g., as described in WO 2008 / 077546. Asn297 refers to the asparagine residue located at about position 297 (EU numbering of Fc region residues) of the Fc region; however, Asn297 can also be located approximately ±3 amino acids upstream or downstream of 297, i.e., between positions 294 and 300, due to slight sequence variation in antibodies. Such fucosylation variants may have improved ADCC function. See, for example, U.S. Patent Application Publication No. 2003 / 0157108 (Presta, L.) and U.S. Patent Application Publication No. 2004 / 0093621 (Kyowa Hakko Kogyo Co., Ltd.).Examples of publications related to "defucosylated" or "fucose-deficient" antibody variants include U.S. Patent Application Publication No. 2003 / 0157108, WO 2000 / 61739, WO 2001 / 29246, U.S. Patent Application Publication No. 2003 / 0115614, U.S. Patent Application Publication No. 2002 / 0164328, U.S. Patent Application Publication No. 2004 / 0093621, U.S. Patent Application Publication No. 2004 / 0132140 No. 2004 / 0110704, U.S. Patent Application Publication No. 2004 / 0110282, U.S. Patent Application Publication No. 2004 / 0109865, WO 2003 / 085119, WO 2003 / 084570, WO 2005 / 035586, WO 2005 / 035778, WO 2005 / 053742, WO 2002 / 031140, Okazaki et al. J. Mol. Biol. 336:1239-1249 (2004), Yamane-Ohnuki et al. Biotech. Bioeng. 87:614 (2004). Examples of cell lines capable of producing defucosylated antibodies include Lec13 CHO cells, which are deficient in protein fucosylation (Ripka et al. Arch. Biochem. Biophys. 249:533-545 (1986); U.S. Patent Application Publication No. 2003 / 0157108 (A1); Presta, L. and WO 2004 / 056312 (A1); Adams et al., especially Example 11), and knockout cell lines, such as alpha-1,6-fucosyltransferase gene, FUT8, knockout CHO cells (e.g., Yamane-Ohnuki et al. Biotech. Bioeng. 87:614 (2004); Kanda, Y. et al. al., Biotechnol. Bioeng., 94(4):680-688 (2006); and WO 2003 / 085107).

[0236] Further provided are antibody and fusion protein variants having bisected oligosaccharides, for example, biantennary oligosaccharides attached to the Fc region of the antibody, bisected by GlcNAc. Such antibody or fusion protein variants may have reduced fucosylation and / or improved ADCC function. Examples of such variants are described, for example, in WO 2003 / 011878 (Jean-Mairet et al.), U.S. Pat. No. 6,602,684 (Umana et al.), and U.S. Patent Application Publication No. 2005 / 0123546 (Umana et al.). Variants having at least one galactose residue in the oligosaccharide attached to the Fc region are also provided. Such variants may have improved CDC function. Such variants are described, for example, in WO 1997 / 30087 (Patel et al.), WO 1998 / 58964 (Raju, S.), and WO 1999 / 22764 (Raju, S.).

[0237] Antibodies or Fc region variants are provided that have Fc mutations that increase ADCC activity. In some embodiments, the antibodies or Fc region variants contain one or more mutations that enhance FcγRIIIa binding and / or decrease FcγRIIIb binding. Non-limiting exemplary such mutations may be made at one or more amino acid positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396. Non-limiting exemplary mutations include L234Y, L235Q, G236W, S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L. In some embodiments, the antibody or Fc region variant comprises the mutations F243L / R292P / Y300L / V305I / P396L. See, e.g., Stavenhagen et al., 2007, Cancer Res. 67:8882-8890. In some embodiments, the antibody or Fc region variant comprises the mutations S239D / I332E or S239D / I332E / A330L. See, e.g., Lazar et al., 2006, PNAS USA, 103:4005-4010. In some embodiments, the antibody or Fc region variant comprises the mutations S298A / E333A / K334A. See, e.g., Shields et al., 2001, J. Biol. Chem., 276:6591-6604. In some embodiments, the antibody or Fc region variant comprises the mutations L234Y / L235Q / G236W / S239M / H268D / D270E / S298A or the mutations D270E / K326D / A330M / K334E, or one heavy chain constant region or Fc comprises the mutations L234Y / L235Q / G236W / S239M / H268D / D270E / S298A and the other heavy chain constant region or Fc comprises the mutations D270E / K326D / A330M / K334E. See, e.g., Mimoto et al., 2013, MAbs, 5:229-236.

[0238] Antibody and Fc region variants with amino-terminal leader extensions are also provided. For example, one or more amino acid residues of an amino-terminal leader sequence are present at the amino terminus of any one or more heavy or light chains of the antibody. An exemplary amino-terminal leader extension comprises or consists of three amino acid residues, VHS, present on one or both light chains of the antibody variant.

[0239] The in vivo or serum half-life of a human FcRn high-affinity binding polypeptide can be assayed, for example, in transgenic mice, humans, or non-human primates to which the polypeptide having a variant Fc region is administered (see, e.g., Petkova et al., International Immunology 18(12):1759-1769 (2006)).

[0240] In some embodiments, the antibody or Fc region variant mediates ADCC in the presence of human effector cells more effectively than the parent antibody. In some embodiments, when the amounts of polypeptide variant and parent antibody or Fc region used in the assay are essentially the same, the antibody or Fc region variant is substantially more effective in mediating ADCC in vitro. In some embodiments, when the amounts of polypeptide variant and parent antibody or Fc region used in the assay are essentially the same, the antibody or Fc region variant is substantially more effective in mediating ADCC in vivo. Generally, such variants are identified using the in vitro ADCC assays disclosed herein, although other assays or methods for determining ADCC activity, such as in animal models, are contemplated. IV. Exemplary Conjugates

[0241] In some embodiments, the antibodies or fusion proteins provided herein are conjugated to another molecule. In some embodiments, the additional molecule can be a detectable marker, such as a label. In some embodiments, the additional molecule can be a therapeutic molecule, such as a cytotoxic agent. In some embodiments, a label and / or a cytotoxic agent can be conjugated to the antibodies or fusion proteins provided herein. As used herein, a label is a moiety that facilitates detection of the antibody or fusion protein and / or that facilitates detection of the molecule to which the antibody or fusion protein binds. Non-limiting exemplary labels include, but are not limited to, radioisotopes, fluorescent groups, enzymatic groups, chemiluminescent groups, biotin, epitope tags, metal-binding tags, etc. Those skilled in the art can select an appropriate label according to a particular application.

[0242] As used herein, a cytotoxic agent is a moiety that reduces the proliferative capacity of one or more cells. A cell's proliferative capacity is reduced when the cell is no longer able to proliferate, for example, because the cell undergoes apoptosis or otherwise dies, the cell fails to progress through the cell cycle and / or divides, the cell differentiates, etc. Non-limiting exemplary cytotoxic agents include, but are not limited to, radioisotopes, toxins, and chemotherapeutic agents. One skilled in the art can select an appropriate cytotoxic agent according to the intended use. In some embodiments, the cytotoxic agent is at least one of an antimetabolite, an alkylating agent, an antibiotic, a growth factor, a cytokine, an antiangiogenic agent, an antimitotic agent, an anthracycline, a toxin, or an apoptotic agent.

[0243] In some embodiments, the label and / or cytotoxic agent is conjugated to the antibody or fusion protein provided herein using in vitro chemical methods. Non-limiting exemplary chemical methods of conjugation are known in the art and include commercially available services, methods, and / or reagents from, for example, Thermo Scientific Life Science Research Produces (formerly Pierce; Rockford, Ill.), Prozyme (Hayward, Calif.), SACRI Antibody Services (Calgary, Canada), AbD Serotec (Raleigh, NC), and others. In some embodiments, when the label and / or cytotoxic agent is a polypeptide, the label and / or cytotoxic agent can be expressed from the same expression vector as at least one antibody chain or fusion protein to produce a polypeptide comprising the label and / or cytotoxic agent fused to the antibody or fusion protein. Those skilled in the art can select a suitable method for conjugating the label and / or cytotoxic agent to the antibody or fusion protein according to the intended use.

[0244] In some embodiments, the conjugation may be covalent. In some embodiments, the conjugation may be non-covalent. In some embodiments, the conjugation may be via a specific binding interaction, for example, via binding of a secondary antibody.

[0245] An exemplary embodiment of a conjugate comprising a drug (or drug derivative) and an antibody of the fusion protein disclosed herein can have the general formula 1: Mab-[linker]-drug (Formula 1) The linker is a cleavable or non-cleavable linker, the Mab is an antibody or fusion protein disclosed herein, and the drug is any drug or cytotoxic agent.

[0246] In some embodiments, the conjugate may comprise general formula 2: MAb-[L2]-[L1]-[AA] m -[A']-drug (Formula 2) wherein MAb is an antibody or fusion protein disclosed herein, L2 is a component of a crosslinker comprising an antibody-binding moiety and one or more acetylene (or azide) groups, L1 comprises a defined PEG having an azide (or acetylene) at one end that is complementary to the acetylene (or azide) moiety in L2 and a reactive group such as a carboxylic acid or hydroxyl group at the other end, AA is an L-amino acid, m is an integer having a value of 0, 1, 2, 3, or 4, and A' is an additional spacer selected from the group consisting of ethanolamine, 4-hydroxybenzyl alcohol, 4-aminobenzyl alcohol, or substituted or unsubstituted ethylenediamine. The L amino acid of "AA" is selected from alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine. When the A' group contains a hydroxyl, it is linked to the hydroxyl or amino group of the drug in the form of a carbonate or carbamate, respectively.

[0247] In some embodiments of Formula 2, A' is a substituted ethanolamine derived from an L-amino acid, where the carboxylic acid group of the amino acid is replaced by a hydroxymethyl moiety. A' can be derived from any one of the following L-amino acids: alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine.

[0248] In an example of a conjugate of such an embodiment of Formula 2, m is 0, A' is L-valinol, and the drug is exemplified by SN-38. The resulting structure is shown in Formula 3. [ka]

[0249] In another example of a conjugate of this embodiment of Formula 2, m is 1 and is represented by a derivatized L-lysine, A' is L-valinol, and the drug is exemplified by SN-38. The structure is shown in Formula 4. [ka]

[0250] In this embodiment, an amide bond is first formed between the carboxylic acid and amino groups of an amino acid, such as lysine, using an orthogonal protecting group for the lysine amino group. The protecting group on the N-terminus of lysine is removed, leaving the protecting group on the lysine side chain intact, and the carboxyl group of a defined PEG bearing an azide (or acetylene) at its N-terminus is attached to the other end. The hydroxyl group of valinol is then attached to a 20-chloroformate derivative of 10-hydroxy-protected SN-38, and this intermediate is attached to the L2 building block bearing the targeting vector binding moiety and the complementary acetylene (or azide) group involved in click cycloaddition chemistry. Finally, removal of the protecting groups on both the lysine side chain and SN-38 yields the product of this example, shown in Formula 3.

[0251] Without wishing to be bound by theory, the small MWSN-38 product generated after intracellular proteolysis, i.e., valinol-SN-38 carbonate, has an additional route for the liberation of intact SN-38 via intramolecular cyclization involving the amino group of valinol and the carbonyl of the carbonate.

[0252] In another embodiment, A' in general formula 2 is A-OH, whereby A-OH is 4-aminobenzyl alcohol or C-C at the benzylic position. 10 A cleavable moiety such as a substituted 4-aminobenzyl alcohol substituted with an alkyl group, the latter being attached via its amino group to an L-amino acid or a polypeptide containing up to four L-amino acid moieties, and the N-terminus being attached to a crosslinker terminating in a targeting moiety-binding group.

[0253] An example of such an embodiment is shown below. The A-OH embodiment of A' in general formula (2) is derived from a substituted 4-aminobenzyl alcohol, where "AA" consists of a single L-amino acid where m = 1 in general formula (2), and the drug is exemplified by SN-38. The structure is represented below (Formula 5, designated MAb-CLX-SN-38). The single amino acid in AA is selected from any one of the following L-amino acids: alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine. The substituent R on the 4-aminobenzyl alcohol moiety (the A-OH embodiment of A') is hydrogen or an alkyl group selected from C1-C10 alkyl groups. [ka]

[0254] An embodiment of MAb-CLX-SN-38 of Formula 5, where the single amino acid AA is L-lysine, R=H, and the drug is exemplified by SN-38 (Formula 6, designated MAb-CL2A-SN-38). [ka]

[0255] Other embodiments are possible within the context of 10-hydroxy-containing camptothecins, such as SN-38. In the example of SN-38 as the drug, the more reactive 10-hydroxy group of the drug is derivatized, leaving the 20-hydroxyl group unaffected. Within general formula 2, A' is a substituted ethylenediamine. An example of this embodiment is represented by formula '7' below, in which the phenolic hydroxyl group of SN-38 is derivatized as a carbamate with a substituted ethylenediamine, and the other amine of the diamine is derivatized as a carbamate with 4-aminobenzyl alcohol, with the latter amino group attached to a Phe-Lys dipeptide. In this structure (formula 7), R and R' are independently hydrogen or methyl. When R = R' = methyl, it is referred to as MAb-CL17-SN-38 or MAb-CL2E-SN-38. [ka]

[0256] In some embodiments, AA comprises a polypeptide moiety, such as a di-, tri-, or tetrapeptide, that is cleavable by an intracellular peptidase, examples being Ala-Leu, Leu-Ala-Leu, and Ala-Leu-Ala-Leu (SEQ ID NO: 129, Trouet et al., 1982).

[0257] In some embodiments, the L1 component of the conjugate contains a defined polyethylene glycol (PEG) spacer having 1 to 30 repeating monomer units. In further embodiments, the PEG is a defined PEG having 1 to 12 repeating monomer units. Introduction of PEG can include using commercially available heterobifunctional PEG derivatives. Heterobifunctional PEGs can contain azide or acetylene groups. An example of a heterobifunctional defined PEG containing 8 repeating monomer units, where "NHS" is succinimidyl, is shown below in Formula 8. [ka]

[0258] In some embodiments, L2 has a plurality of acetylene (or azide) groups, ranging from 2 to 40, preferably 2 to 20, more preferably 2 to 5, and a single targeting vector binding moiety.

[0259] A representative SN-38 conjugate of an antibody containing multiple drug molecules and a single targeting vector binding moiety is shown below. The "L2" component of this structure is attached to two acetylene groups, resulting in the attachment of two azide-tagged SN-38 molecules. The linkage to the MAb is depicted as a succinimide. [ka]

[0260] In some embodiments, when a bifunctional drug contains a thiol-reactive moiety as the antibody-binding group, thiols on the antibody are generated on the lysine groups of the antibody using a thiolation reagent. Methods for introducing thiol groups onto antibodies by modifying the lysine groups of MAbs are well known in the art (Wong in Chemistry of protein conjugation and cross-linking, CRC Press, Inc., Boca Raton, Fla. (1991), pp. 20-22). Alternatively, mild reduction of the interchain disulfide bonds on antibodies using a reducing agent such as dithiothreitol (DTT) (Willner et al., Bioconjugate Chem. 4:521-527 (1993)) can generate 7 to 10 thiols on the antibody, which has the advantage of allowing the incorporation of multiple drug moieties into the interchain region of MAbs, away from the antigen-binding region.

[0261] In some embodiments, the chemotherapeutic moiety is doxorubicin (DOX). , epirubicin, morpholinodoxorubicin (morpholino-DOX), cyanomorpholino-doxorubicin (cyanomorpholino-DOX), 2-pyrrolino-doxorubicin (2-PDOX), CPT, 10-hydroxycamptothecin, S The chemotherapeutic moiety is selected from the group consisting of N-38, topotecan, lutotecan, 9-aminocamptothecin, 9-nitrocamptothecin, taxanes, geldanmycin, ansamycins, and epothilones. In one embodiment, the chemotherapeutic moiety is SN-38. Preferably, in the conjugates of some embodiments, the targeting moiety is linked to at least one chemotherapeutic moiety, preferably 1 to about 12 chemotherapeutic moieties, and most preferably about 6 to about 12 chemotherapeutic moieties.

[0262] Furthermore, in some embodiments, the linker component "L2" contains a thiol group that reacts with a thiol-reactive residue introduced into one or more lysine side chain amino groups of the targeting moiety. In such cases, the antibody can be pre-derivatized with a thiol-reactive group such as maleimide, vinyl sulfone, bromoacetamide, or iodoacetamide by procedures well described in the art. Exemplary Leader Sequences

[0263] For the expression and secretion of large quantities of some secreted proteins, a leader sequence from a heterologous protein may be desirable. In some embodiments, the use of a heterologous leader sequence may be advantageous in that when the leader sequence is removed in the ER during the secretion process, the resulting mature polypeptide may remain unaltered. The addition of a heterologous leader sequence may be useful for the expression and secretion of some proteins.

[0264] Certain exemplary leader sequence sequences are described, for example, in the online leader sequence database maintained by the Department of Biochemistry, National University of Singapore. See Choo et al., BMC Bioinformatics, 6:249 (2005), and PCT Publication No. WO 2006 / 081430. V. Polypeptide Expression and Production Nucleic acid molecules encoding antibodies or fusion proteins

[0265] Provided herein are nucleic acid molecules comprising polynucleotides encoding one or more chains of an anti-CCR8 antibody. In some embodiments, the nucleic acid molecule comprises a polynucleotide encoding the heavy chain or the light chain of the anti-CCR8 antibody. In some embodiments, the nucleic acid molecule comprises both a polynucleotide encoding the heavy chain and a polynucleotide encoding the light chain of the anti-CCR8 antibody. In some embodiments, a first nucleic acid molecule comprises a first polynucleotide encoding the heavy chain, and a second nucleic acid molecule comprises a second polynucleotide encoding the light chain.

[0266] In some embodiments, the heavy and light chains are expressed as two separate polypeptides from one nucleic acid molecule or from two separate nucleic acid molecules, in some embodiments, a single polynucleotide encodes a single polypeptide comprising both the heavy and light chains linked together, such as when the antibody is an scFv.

[0267] In some embodiments, a polynucleotide encoding the heavy or light chain of an anti-CCR8 antibody comprises a nucleotide sequence encoding at least one of the CDRs provided herein. In some embodiments, a polynucleotide encoding the heavy or light chain of an anti-CCR8 antibody comprises a nucleotide sequence encoding at least three of the CDRs provided herein. In some embodiments, a polynucleotide encoding the heavy or light chain of an anti-CCR8 antibody comprises a nucleotide sequence encoding at least six of the CDRs provided herein. In some embodiments, a polynucleotide encoding the heavy or light chain of an anti-CCR8 antibody comprises a nucleotide sequence encoding a leader sequence, which, upon translation, is located at the N-terminus of the heavy or light chain. As described above, the leader sequence may be a native heavy or light chain leader sequence or another heterologous leader sequence.

[0268] Provided herein are nucleic acid molecules comprising a polynucleotide encoding a fusion protein. In some embodiments, the polynucleotide encoding the fusion protein comprises a nucleotide sequence encoding a leader sequence, which, upon translation, is located at the N-terminus of the fusion protein. The leader sequence may be a native leader sequence or another heterologous leader sequence.

[0269] In some embodiments, the nucleic acid encodes any of the amino acid sequences of the antibodies and fusion proteins in the sequence listing herein. In some embodiments, the nucleic acid is at least 80% identical, e.g., at least 80, 85, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% identical, to a nucleic acid encoding any of the amino acid sequences for the antibodies and fusion proteins in the sequence listing herein. In some embodiments, the nucleic acid hybridizes to any one or more of the nucleic acid sequences provided herein. In some embodiments, the hybridization is under moderate conditions. In some embodiments, the hybridization is under high stringency conditions, such as at least about 6X SSC and 1% SDS at 65°C, with an initial wash in about 20% (v / v) formamide in 0.1X SSC at about 42°C for 10 minutes, followed by a subsequent wash in 0.2X SSC and 0.1% SDS at 65°C.

[0270] The nucleic acid molecule can be constructed using recombinant DNA techniques conventional in the art. In some embodiments, the nucleic acid molecule is an expression vector suitable for expression in a selected host cell.

[0271] Vectors comprising polynucleotides encoding anti-CCR8 heavy chains and / or anti-CCR8 light chains are provided. Vectors comprising polynucleotides encoding anti-CCR8 heavy chains and / or anti-CCR8 light chains are also provided. Such vectors include, but are not limited to, DNA vectors, phage vectors, viral vectors, retroviral vectors, etc. In some embodiments, the vector comprises a first polynucleotide sequence encoding a heavy chain and a second polynucleotide sequence encoding a light chain. In some embodiments, the heavy chain and light chain are expressed from the vector as two separate polypeptides. In some embodiments, the heavy chain and light chain are expressed as part of a single polypeptide, for example, when the antibody is an scFv.

[0272] In some embodiments, a first vector comprises a polynucleotide encoding a heavy chain, and a second vector comprises a polynucleotide encoding a light chain. In some embodiments, the first vector and the second vector are transfected into host cells in similar amounts (e.g., similar molar amounts or similar masses). In some embodiments, a molar or mass ratio of the first vector and the second vector of 5:1 to 1:5 is transfected into host cells. In some embodiments, a mass ratio of 1:1 to 1:5 is used for the vector encoding the heavy chain and the vector encoding the light chain. In some embodiments, a mass ratio of 1:2 is used for the vector encoding the heavy chain and the vector encoding the light chain.

[0273] Vectors comprising a polynucleotide encoding the fusion protein are provided, including, but not limited to, DNA vectors, phage vectors, viral vectors, retroviral vectors, etc.

[0274] In some embodiments, a vector is selected that is optimized for expression of a polypeptide in CHO or CHO-derived cells or NSO cells. Exemplary such vectors are described, for example, in Running Deer et al., Biotechnol. Prog. 20:880-889 (2004). host cell

[0275] In some embodiments, the antibodies or fusion proteins provided herein can be expressed in prokaryotic cells, such as bacterial cells, or eukaryotic cells, such as fungal cells (such as yeast), plant cells, insect cells, and mammalian cells. Such expression can be carried out, for example, according to procedures known in the art. Exemplary eukaryotic cells that can be used to express polypeptides include, but are not limited to, COS cells, including COS7 cells; 293 cells, including 293-6E cells; CHO-S cells, including DG44, Lec13, and FUT8 CHO cells; PER.C6® cells (Crucell); and NSO cells. In some embodiments, the antibodies or fusion proteins provided herein can be expressed in yeast. See, for example, U.S. Patent Application Publication No. 2006 / 0270045 A1. In some embodiments, a particular eukaryotic host cell is selected based on its ability to make desired post-translational modifications to the antibodies or fusion proteins provided herein. For example, in some embodiments, CHO cells produce polypeptides that have higher levels of sialylation than the same polypeptides produced in 293 cells.

[0276] Introduction of one or more nucleic acids into a desired host cell can be accomplished by any method, including, but not limited to, calcium phosphate transfection, DEAE-dextran mediated transfection, cationic lipid mediated transfection, electroporation, transduction, infection, etc. Non-limiting exemplary methods are described, for example, in Sambrook et al., Molecular Cloning, A Laboratory Manual,3 rd Cold Spring Harbor Laboratory Press, 2001. The nucleic acid may be transiently or stably transfected into the desired host cell according to any suitable method.

[0277] Host cells containing any of the polynucleotides or vectors described herein are also provided. In some embodiments, host cells containing an antibody or fusion protein are provided. Any host cell capable of overexpressing heterologous DNA can be used for the purpose of isolating genes encoding antibodies, polypeptides, or proteins of interest. Non-limiting examples of mammalian host cells include, but are not limited to, COS, HeLa, and CHO cells. See also PCT Publication WO 87 / 04462. Suitable non-mammalian host cells include prokaryotes (such as E. coli or B. subtillis) and yeast (such as S. cerevisae, S. pombe, or K. lactis). Polypeptide purification

[0278] The antibodies and fusion proteins provided herein can be purified by any suitable method. Such methods include, but are not limited to, the use of affinity matrix or hydrophobic interaction chromatography. Suitable affinity ligands include ROR1 ECD and ligands that bind to antibody constant regions. For example, Protein A, Protein G, Protein A / G, or antibody affinity columns can be used to bind the constant region or Fc region and purify antibodies or fusion proteins. Hydrophobic interaction chromatography, such as butyl or phenyl columns, can also be suitable for purifying some polypeptides. Ion exchange chromatography (e.g., anion exchange chromatography and / or cation exchange chromatography) can also be suitable for purifying some polypeptides. Mixed-mode chromatography (e.g., reversed-phase / anion exchange, reversed-phase / cation exchange, hydrophilic interaction / anion exchange, hydrophilic interaction / cation exchange, etc.) can also be suitable for purifying some polypeptides. Many methods for purifying polypeptides are known in the art. Cell-free production of polypeptides

[0279] In some embodiments, the antibody or fusion protein is produced in a cell-free system. Non-limiting exemplary cell-free systems include those described, for example, in Sitaraman et al., Methods Mol.Biol.498:229-44(2009);Spirin,Trends Biotechnol. 22:538-45 (2004); Endo et al., Biotechnol. Adv. 21:695-713 (2003). composition

[0280] In some embodiments, an antibody or fusion protein prepared by the above method is provided. In some embodiments, the antibody or fusion protein is prepared in a host cell. In some embodiments, the antibody or fusion protein is prepared in a cell-free system. In some embodiments, the antibody or fusion protein is purified. In some embodiments, a cell culture medium comprising the antibody or fusion protein is provided. In some embodiments, a host cell culture medium comprising the antibody or fusion protein is provided.

[0281] In some embodiments, compositions are provided comprising an antibody or fusion protein prepared by the methods described above. In some embodiments, the composition comprises an antibody or fusion protein prepared in a host cell. In some embodiments, the composition comprises an antibody or fusion protein prepared in a cell-free system. In some embodiments, the composition comprises a purified antibody or fusion protein.

[0282] In some embodiments, compositions are provided that comprise an antibody or fusion protein at a concentration of greater than any one of about 10 mg / mL, 20 mg / mL, 30 mg / mL, 40 mg / mL, 50 mg / mL, 60 mg / mL, 70 mg / mL, 80 mg / mL, 90 mg / mL, 100 mg / mL, 125 mg / mL, 150 mg / mL, 175 mg / mL, 200 mg / mL, 225 mg / mL, or 250 mg / mL. VI. Therapeutic Compositions and Methods Methods of treating diseases using antibodies or fusion proteins

[0283] Antibodies (e.g., 7-B16, 1-K17, etc.) and fusion proteins, as well as compositions comprising the antibodies and fusion proteins, are provided for use in human or animal therapeutic methods. Pharmaceutical and therapeutic uses of the disclosed antibodies and fusion proteins are also provided herein. Methods and pharmaceutical / therapeutic applications for treating diseases that involve administering the antibodies and fusion proteins provided herein are also provided. Non-limiting exemplary diseases that can be treated with the antibodies and fusion proteins provided herein include, but are not limited to, cancer.

[0284] In some embodiments, a method of treating cancer (or therapeutic / pharmaceutical use of the disclosed antibodies or fusion proteins) comprises administering to a subject with cancer an effective amount of an antibody (e.g., 7-B16, 1-K17, etc.) or fusion protein provided herein. In some embodiments, the cancer is a solid cancer. In some embodiments, the cancer is a hematological (i.e., liquid) cancer. In some embodiments, the cancer comprises tumor-infiltrating Treg cells. In some embodiments, the subject has previously received checkpoint inhibitor therapy (CPI) or has a CPI-resistant or refractory cancer, and many subjects with CPI-resistant or refractory cancer exhibit elevated levels of intratumoral Tregs. In some embodiments, the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab; the anti-CLTA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. In some embodiments, the cancer comprises cells that express CCR8 (e.g., CCR8-expressing Tregs). CCR8 expression can be determined, for example, by immunohistochemistry, fluorescence-activated cell sorting (FACS), gene expression analysis (such as Q-PCR or RT-PCR), Western blot, ELISA, or other known methods of assessing expression at the genetic or protein level. In some embodiments, the CCR8-expressing cells are Treg cells (e.g., intratumoral Tregs), and in some embodiments, the level of CCR8 expression in such intratumoral Tregs is elevated relative to peripheral Tregs. In some embodiments, CCR8 is expressed on the surface of Treg cells at less than 10,000 copies per cell, which can be determined, for example, by fluorescence-activated cell sorting (FACS) and flow cytometry. In some embodiments, the cancer comprises tumor cells that express CCR8.In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells express less than 10,000 copies of CCR8 per cell on their surface.

[0285] In some embodiments, the cancer is selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. In some embodiments, the cancer may be metastatic. In some embodiments, when the cancer is breast cancer, the breast cancer is triple-negative breast cancer (i.e., triple-negative breast cancer, TNBC). In some embodiments, when the cancer is breast cancer, the breast cancer is metastatic TNBC.

[0286] In some embodiments, a method for treating a solid cancer having tumor-infiltrating Tregs that express CCR8 comprises administering to a subject with cancer an effective amount of an antibody (e.g., 7-B16, 1-K17, etc.) or fusion protein provided herein. In some embodiments, the tumor-infiltrating Tregs express CCR8 at elevated levels relative to the CCR8 expression level of peripheral Tregs. CCR8 expression can be determined, for example, by immunohistochemistry, fluorescence-activated cell sorting (FACS), gene expression analysis (such as Q-PCR or RT-PCR), Western blot, ELISA, or other known methods for assessing expression at the genetic or protein level. In some embodiments, the solid cancer can be selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma.

[0287] In some embodiments, a method of treating hematological cancer (or therapeutic / pharmaceutical use) comprises administering to a subject with a hematological cancer an effective amount of an antibody that binds to human CCR8 (e.g., 7-B16, 1-K17, etc.). In some embodiments, the antibody inhibits binding of CCL1 to CCR8. In some embodiments, the hematological cancer expresses CCR8. In some embodiments, the subject has previously received checkpoint inhibitor therapy (CPI) or has a CPI-resistant or refractory cancer, and many subjects with CPI-resistant or refractory cancer will exhibit elevated levels of CCR8-expressing Tregs. In some embodiments, the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab; the anti-CLTA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. In some embodiments, the hematological cancer comprises cells that express CCR8 (e.g., CCR8-expressing Tregs). In some embodiments, the CCR8-expressing cells are Treg cells. In some embodiments, the hematological cancer may be metastatic.

[0288] In some embodiments, the method or use for selecting a subject having a hematological cancer for treatment with an antibody that binds to human CCR8 comprises detecting CCR8 expression in a sample from the subject, and in some embodiments, the method or use further comprises administering an effective amount of an antibody that binds to human CCR8.

[0289] In some embodiments, a method for treating a hematological cancer (or therapeutic / pharmaceutical use) comprises administering an effective amount of a fusion protein to a subject with the hematological cancer. In some embodiments, a method for selecting a subject with a hematological cancer for treatment with a fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region comprises detecting CCR8 expression in a sample from the subject. In some embodiments, the method further comprises administering an effective amount of a fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region. In some embodiments, the hematological cancer expresses CCR8. In some embodiments, the subject has previously received checkpoint inhibitor therapy (CPI) or has a CPI-resistant or refractory cancer, and many subjects with CPI-resistant or refractory cancer will exhibit elevated levels of CCR8-expressing Tregs. In some embodiments, the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab; the anti-CLTA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. In some embodiments, the hematological cancer comprises cells that express CCR8 (e.g., CCR8-expressing Tregs). In some embodiments, the CCR8-expressing cells are Treg cells. In some embodiments, Tregs express fewer than 10,000 copies of CCR8 per cell, which can be determined, for example, by fluorescence-activated cell sorting (FACS) and flow cytometry. In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells express less than 10,000 copies of CCR8 per cell on their surface.In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells in a sample obtained from the tumor express less than 10,000 copies of CCR8 per cell. In some embodiments, the hematological cancer may be metastatic.

[0290] In some embodiments, the hematological cancer is mixed B-cell and T-cell leukemia, B-cell lymphoma, chronic myelogenous leukemia (CML), chronic myelomonocytic leukemia, diffuse large B-cell lymphoma (DLBC), lymphoma, mantle cell lymphoma (MCL), multiple myeloma, myelodysplastic syndrome (MDS), myeloproliferative disorders, peripheral T-cell lymphoma, T-cell leukemia, acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), CLL / SLL, mature T-cell and NK cell leukemia. Follicular lymphoma, follicular lymphoma, acute lymphocytic leukemia (ALL), T-cell acute lymphocytic leukemia (TALL), T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, cutaneous T-cell lymphoma (CTCL), adult T-cell leukemia / lymphoma (ATLL), T-cell lymphoblastic leukemia / lymphoma (TLLL), angioimmunoblastic T-cell lymphoma (ATCL), hepatosplenic T-cell lymphoma (HTCL), peripheral T-cell lymphoma not otherwise specified (PTCL) NOS), Burkitt's lymphoma (BL), chronic myelomonocytic leukemia (CMML), extranodal NK / T-cell lymphoma (NKTCL), primary effusion lymphoma (PEL), acute lymphocytic leukemia / acute myeloid leukemia (ALL, AML), histiocytic lymphoma (HL), marginal zone lymphoma (MZL), B-cell acute lymphocytic leukemia, or anaplastic large cell lymphoma (ALCL).

[0291] In some embodiments, the hematological cancer is T-cell adult acute lymphocytic leukemia, T-cell childhood acute lymphocytic leukemia, lymphoblastic lymphoma, acute lymphocytic leukemia, cutaneous T-cell lymphoma, T-cell acute lymphocytic leukemia, adult T-cell leukemia / lymphoma, T-cell lymphoblastic leukemia / lymphoma, or anaplastic large cell lymphoma.

[0292] In some embodiments, a method of treating a solid tumor (or therapeutic / pharmaceutical use) comprises administering to a subject with a solid tumor an effective amount of an antibody that binds to human CCR8 (e.g., 7-B16, 1-K17, etc.). In some embodiments, the antibody inhibits binding of CCL1 to CCR8. In some embodiments, the solid tumor (or tumor-infiltrating Tregs) express CCR8. In some embodiments, the solid tumor comprises intratumoral Tregs. In some embodiments, the intratumoral Tregs have elevated levels of CCR8 expression relative to peripheral Tregs. In some embodiments, CCR8 is expressed on the surface of intratumoral Treg cells at less than 10,000 copies per cell (e.g., when the entire Treg population has low Treg expression), which can be determined, for example, by fluorescence-activated cell sorting (FACS) and flow cytometry. In some embodiments, the intratumoral Tregs comprise Tregs that express less than 10,000 copies of CCR8 per cell (referring to a subpopulation of Tregs with low CCR8 expression). In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells express less than 10,000 copies of CCR8 per cell on their surface. In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells in a sample obtained from a tumor express less than 10,000 copies of CCR8 per cell. In some embodiments, the subject has previously received checkpoint inhibitor therapy (CPI) or has a CPI-resistant or refractory cancer, and many subjects with CPI-resistant or refractory cancer will exhibit elevated levels of intratumoral Tregs. In some embodiments, the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody.In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab; the anti-CLTA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. In some embodiments, the intratumoral Tregs are depleted. Treg depletion can be the result of ADCC and / or CDC activity of the disclosed antibodies.

[0293] In some embodiments, the method or use for selecting a subject with a solid tumor for treatment with an antibody that binds to human CCR8 comprises detecting CCR8 expression in a sample from the subject. In some embodiments, CCR8 expression is detected in a population of tumor-infiltrating Tregs in the sample. In some embodiments, tumor-infiltrating Tregs express fewer than 10,000 copies of CCR8 per cell, which can be determined, for example, by fluorescence-activated cell sorting (FACS) and flow cytometry. In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells express fewer than 10,000 copies of CCR8 per cell on their surface. In some embodiments, the method or use further comprises administering an effective amount of an antibody that binds to human CCR8.

[0294] In some embodiments, a method for treating a solid tumor (or therapeutic / pharmaceutical use) comprises administering an effective amount of a fusion protein to a subject with the solid tumor. In some embodiments, a method for selecting a subject with a solid tumor for treatment with a fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region comprises detecting CCR8 expression in a sample from the subject. In some embodiments, the method further comprises administering an effective amount of a fusion protein comprising (a) CCL1 or an active fragment thereof or MC148 or an active fragment thereof, and (b) an Fc region. In some embodiments, the solid tumor expresses CCR8. In some embodiments, the subject has previously received checkpoint inhibitor therapy (CPI) or has CPI-resistant or refractory cancer, and many subjects with CPI-resistant or refractory cancer will exhibit elevated levels of CCR8-expressing Tregs. In some embodiments, the CPI therapy comprises an anti-PDL1 antibody, an anti-CTLA4 antibody, or an anti-TIGIT antibody. In some embodiments, the anti-PDL1 antibody is selected from pembrolizumab, nivolumab, atezolizumab, avelumab, durvalumab, cemiplimab-rwlc, and zimvelerimab; the anti-CLTA4 antibody is ipilimumab or tremelimumab; or the anti-TIGIT antibody is selected from tiragolumab, vibostolimab, domvanalimab, AB308, BMS-986207, and durvalumab. In some embodiments, the solid tumor comprises cells that express CCR8 (e.g., CCR8-expressing Tregs). In some embodiments, the CCR8-expressing cells are Treg cells. In some embodiments, the Tregs express fewer than 10,000 copies of CCR8 per cell (as determined, for example, by FACS and flow cytometry) and may be tumor-infiltrating Tregs. In some embodiments, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of tumor-infiltrating Treg cells express less than 10,000 copies of CCR8 per cell on their surface.In some embodiments, the hematological cancer may be metastatic.

[0295] In some embodiments, the solid cancer may be selected from breast cancer, colorectal cancer, head and neck cancer, lung cancer, ovarian cancer, gastric cancer, gastric adenocarcinoma, and thymoma. In some embodiments, the solid cancer may be metastatic. In some embodiments, when the solid cancer is breast cancer, the breast cancer is triple-negative breast cancer (i.e., TNBC). In some embodiments, when the solid cancer is breast cancer, the breast cancer is metastatic TNBC.

[0296] In some embodiments of any of the aforementioned methods or uses, the administered antibody (e.g., 7-B16, 1-K17, etc.) or fusion protein may deplete Tregs in the subject as a result of the ADCC and / or CDC properties of the disclosed antibodies and fusion proteins.

[0297] The antibody or fusion protein can be administered to the subject as needed. The frequency of administration can be determined by a skilled artisan, such as the attending physician, based on considerations of the condition being treated, the age of the subject being treated, the severity of the condition being treated, and the general state of the subject's health. In some embodiments, an effective dose of the antibody or fusion protein is administered to the subject one or more times. In some embodiments, an effective dose of the antibody or fusion protein is administered to the subject once a month, or less frequently, such as every two or three months. In some embodiments, an effective dose of the antibody or fusion protein is administered to the subject less frequently, such as once every three weeks, once every two weeks, or once a week. An effective dose of the antibody or fusion protein is administered to the subject at least once. In some embodiments, an effective dose of the antibody or fusion protein can be administered multiple times, including over a period of at least one month, at least six months, or at least one year.

[0298] In some embodiments, the pharmaceutical compositions are administered in an amount effective for treating (including preventing) cancer. A therapeutically effective amount typically depends on the weight of the subject being treated, their physical or health condition, the extent of the condition being treated, and the age of the subject being treated. Generally, antibodies and fusion proteins may be administered in an amount ranging from about 10 μg / kg to about 100 mg / kg of body weight per dose. In some embodiments, antibodies and fusion proteins may be administered in an amount ranging from about 50 μg / kg to about 5 mg / kg of body weight per dose. In some embodiments, antibodies and fusion proteins may be administered in an amount ranging from about 100 μg / kg to about 10 mg / kg of body weight per dose. In some embodiments, antibodies and fusion proteins may be administered in an amount ranging from about 100 μg / kg to about 20 mg / kg of body weight per dose. In some embodiments, antibodies and fusion proteins may be administered in an amount ranging from about 0.5 mg / kg to about 20 mg / kg of body weight per dose. Pharmaceutical Composition

[0299] In some embodiments, compositions comprising the antibodies and fusion proteins provided herein are provided in formulations with a wide variety of pharmaceutically acceptable carriers (see, e.g., Gennaro, Remington: The Science and Practice of Pharmacy with Facts and Comparisons:Drugfacts Plus,20th ed.(2003);Ansel et al.,Pharmaceutical Dosage Forms and Drug Delivery Systems,7 th ed., Lippencott Williams and Wilkins (2004); Kibbe et al., Handbook of Pharmaceutical Excipients, 3 rd(See, e.g., Pharmaceutical Press (2000)). A variety of pharmaceutically acceptable carriers, including vehicles, adjuvants, and diluents, are available. In addition, a variety of pharmaceutically acceptable auxiliary substances, such as pH adjusting and buffering agents, tonicity adjusting agents, stabilizers, wetting agents, and the like, are also available. Non-limiting exemplary carriers include saline, buffered saline, dextrose, water, glycerol, ethanol, and combinations thereof.

[0300] In some embodiments, a pharmaceutical composition comprising the antibody or fusion protein is provided. In some embodiments, the pharmaceutical composition comprises a humanized antibody. In some embodiments, the pharmaceutical composition comprises an antibody or fusion protein prepared in a host cell or cell-free system described herein. In some embodiments, the pharmaceutical composition comprises a pharmaceutically acceptable carrier.

[0301] In some embodiments, the pharmaceutical composition is administered in an amount effective to treat (including prevent) cancer. A therapeutically effective amount typically depends on the weight of the subject being treated, their physical or health condition, the extent of the condition being treated, or the age of the subject being treated. Administration route

[0302] In some embodiments, the antibodies and / or fusion proteins provided herein can be administered in vivo by various routes, including, but not limited to, intravenous, intraarterial, parenteral, intratumoral, intraperitoneal, or subcutaneous. The appropriate formulation and route of administration can be selected according to the intended use. Combination therapy

[0303] The antibodies and / or fusion proteins provided herein (e.g., 7-B16, 1-K17, etc.) can be administered alone or in combination with other therapeutic modalities. They can be provided before, substantially simultaneously with, and / or after the administration of other therapeutic modalities, e.g., surgery, chemotherapy, radiation therapy, or a biologic such as another therapeutic antibody. In some embodiments, the antibodies and / or fusion proteins provided herein (e.g., 7-B16, 1-K17, etc.) are administered in conjunction with another anti-cancer agent. In some embodiments, the antibodies and / or fusion proteins provided herein are administered in conjunction with radiation therapy, such as ablative or non-ablative radiation therapy.

[0304] In some embodiments, the antibody and / or fusion protein (e.g., 7-B16, 1-K17, etc.) is administered simultaneously with the second therapeutic agent. For example, the two or more therapeutic agents are administered at a time interval of about 60 minutes or less, such as about any of 30, 15, 10, 5, or 1 minute or less. In some embodiments, the antibody and / or fusion protein is administered sequentially with the second therapeutic agent. For example, the administration of the two or more therapeutic agents is at a time interval of more than about 15 minutes, such as about 20, 30, 40, 50, or 60 minutes, 1 day, 2 days, 3 days, 1 week, 2 weeks, or 1 month or more, apart.

[0305] In some embodiments, the antibodies and / or fusion proteins (e.g., 7-B16, 1-K17, etc.) are administered in conjunction with a second therapeutic method for treatment. Thus, administration of the antibodies provided herein can be combined with another therapeutic system.

[0306] In some embodiments, the antibodies and / or fusion proteins provided herein (e.g., 7-B16, 1-K17, etc.) are administered in conjunction with an anti-PD-1 or anti-PD-L1 therapy. In some embodiments, the antibodies and / or fusion proteins provided herein are administered in conjunction with an anti-PD-1 antibody or anti-PD-L1 antibody (e.g., atezolizumab).

[0307] In some embodiments, the antibodies and / or fusion proteins provided herein (e.g., 7-B16, 1-K17, etc.) are administered in conjunction with anti-ICOS therapy. In some embodiments, the antibodies and / or fusion proteins provided herein are administered in conjunction with an antibody that binds to Inducible T-Cell Costimulator (ICOS). In some embodiments, the antibodies and / or fusion proteins provided herein are administered with an isolated antibody that binds ICOS, where the anti-ICOS antibody is an agonist of CD4+ T cells (e.g., CD4+ T effector (Teff) cells). In some embodiments, the antibody that binds ICOS is an agonist of CD4+ T cells (e.g., CD4+ Teff cells) and depletes T regulatory (Treg) cells.

[0308] In some embodiments, antibodies and / or fusion proteins provided herein (e.g., 7-B16, 1-K17, etc.) are administered with an agonist anti-OX40 antibody (Medi6469, MedImmune; MOXR0916 / RG7888, Roche, etc.). In some embodiments, antibodies and / or fusion proteins provided herein are administered with an anti-CTLA4 antibody (ipilimumab, YERVOY®, BMS, etc.).

[0309] In some embodiments, the additional therapeutic agent is a chemotherapeutic agent. Exemplary chemotherapeutic agents that may be combined with the antibodies and / or fusion proteins provided herein include, but are not limited to, capectiabine, cyclophosphamide, dacarbazine, temozolomide, cyclophosphamide, docetaxel, doxorubicin, daunorubicin, cisplatin, carboplatin, epirubicin, eribulin, 5-FU, gemcitabine, irinotecan, ixabepilone, methotrexate, mitoxantrone, oxaliplatin, paclitaxel, nab-paclitaxel, ABRAXANE® (protein-bound paclitaxel), pemetrexed, vinorelbine, and vincristine. In some embodiments, the antibodies and / or fusion proteins provided herein are administered with at least one kinase inhibitor. Non-limiting exemplary kinase inhibitors include erlotinib, afatinib, gefitinib, crizotinib, dabrafenib, trametinib, vemurafenib, and cobimetanib.

[0310] In some embodiments, the additional therapeutic agent is an IDO inhibitor. Non-limiting exemplary IDO inhibitors are described, for example, in U.S. Patent Application Publication Nos. 2016 / 0060237 and 2015 / 0352206. Non-limiting exemplary IDO inhibitors include indoximod (New Link Genetic s), INCB024360 (Incyte Corp), 1-methyl-D-tryptophan (New Link Genetics), and GDC-0919 (Genentech).

[0311] In some embodiments, the antibodies and / or fusion proteins provided herein are administered in combination with an immune-modifying drug (IMiD). Non-limiting exemplary IMiDs include thalidomide, lenalidomide, and pomalidomide.

[0312] In some embodiments, the additional therapeutic agent is a cancer vaccine. Cancer vaccines are being investigated as a potential approach to antigen transfer and dendritic cell activation. In particular, vaccination combined with immunological checkpoints or agonists of costimulatory pathways has shown evidence of overcoming tolerance and increasing anti-tumor responses. A range of cancer vaccines using different approaches to promote immune responses to tumors are being tested (see, e.g., Emens LA, Expert Opin Emerg Drugs 13(2):295-308 (2008)). Approaches are designed to enhance the response of B cells, T cells, or professional antigen-presenting cells to tumors. Exemplary types of cancer vaccines include, but are not limited to, peptide-based vaccines that use targeting different tumor antigens, which can be delivered as peptides / proteins or genetically engineered DNA vectors, viruses, bacteria, etc., and cell biological approaches to less well-defined targets, including, but not limited to, vaccines developed from patient-derived dendritic cells, autologous tumor cells, or tumor cell lysates, allogeneic tumor cells, etc.

[0313] Thus, in certain embodiments, the antibodies and / or fusion proteins provided herein (e.g., 7-B16, 1-K17, etc.) can be used in combination with a cancer vaccine. Exemplary cancer vaccines include, but are not limited to, dendritic cell vaccines, oncolytic viruses, tumor cell vaccines, and the like. In some embodiments, such vaccines enhance anti-tumor responses. Examples of cancer vaccines that can be used in combination with the antibodies and / or fusion proteins provided herein include, but are not limited to, MAGE3 vaccines (e.g., for melanoma and bladder cancer), MUC1 vaccines (e.g., for breast cancer), EGFRv3 (such as Rindopepimut, e.g., for brain cancers, including glioblastoma multiforme), or ALVAC-CEA (e.g., for CEA+ cancers).

[0314] Non-limiting exemplary cancer vaccines also include Sipuleucel-T, which is derived from autologous peripheral blood mononuclear cells (PBMCs), which contain antigen-presenting cells (see, e.g., Kantoff PW (See, e.g., Carreno BM et al., N Engl J Med 363:411-22 (2010)). In generating sipuleucel-T, patient PBMCs are activated ex vivo with PA2024, a recombinant fusion protein of prostatic acid phosphatase (a prostate antigen) and granulocyte-macrophage colony-stimulating factor (an immune cell activator). Another approach to candidate cancer vaccines is to generate an immune response against specific peptides mutated in tumor tissue, such as melanoma (see, e.g., Carreno BM et al., Science 348:6236 (2015)). Such mutated peptides may, in some embodiments, be referred to as neoantigens. A non-limiting example of the use of neoantigens in tumor vaccines is the major histocompatibility complex protein HLA-A. * Neoantigens in tumors predicted to bind to 02:01 are identified for individual patients with cancer, such as melanoma. Dendritic cells from the patient are matured ex vivo and then incubated with the neoantigen. The activated dendritic cells are then administered to the patient. In some embodiments, robust T cell immunity against the neoantigen is detectable after administration of the cancer vaccine.

[0315] In some such embodiments, cancer vaccines are developed using neoantigens. In some embodiments, the cancer vaccine is a DNA vaccine. In some embodiments, the cancer vaccine is an engineered virus containing a cancer antigen, such as PROSTVAC (rilimogene galvacirepvec / rilimogene glafolivec). In some embodiments, the cancer vaccine comprises engineered tumor cells, such as GVAX, a granulocyte-macrophage colony-stimulating factor (GM-CSF) gene-transfected tumor cell vaccine (see, e.g., Nemunaitis, 2005, Expert Rev Vaccines, 4:259-74).

[0316] In some embodiments, the antibodies and / or fusion proteins described herein are administered before, simultaneously with, and / or after a cancer vaccine. In some embodiments, a cancer vaccine developed using a neoantigen is used in combination with the antibodies and / or fusion proteins described herein. In some such embodiments, the combination is used to treat cancers with high mutation burden, such as melanoma, lung, bladder, or colorectal cancer.

[0317] In some embodiments, the antibodies and / or fusion proteins provided herein are administered in combination with chimeric antigen receptor T cell (CAR-T) therapy.

[0318] In some embodiments, the antibodies and / or fusion proteins provided herein (e.g., 7-B16, 1-K17, etc.) are administered together with one or more (e.g., one, two, three, or four) additional therapeutic agents. In some embodiments, the additional therapeutic agent is, for example, an inhibitory immune checkpoint blocker or inhibitor, a stimulatory immune checkpoint stimulator, agonist, or activator, a chemotherapeutic agent, an anti-cancer agent, a radiotherapeutic agent, an anti-neoplastic agent, an anti-proliferative agent, an anti-angiogenic agent, an anti-inflammatory agent, an immunotherapeutic agent, a therapeutic antigen binding molecule (e.g., monospecific and multispecific antibodies in any format, such as DART®, Duobody®, BiTE®, BiKE, TriKE, XmAb®, TandAb®, scFv, Fab, Fab derivatives, and fragments thereof), a bispecific antibody, a non-immunoglobulin antibody mimic (e.g., adnectin, affibody molecule, affilin, affimer, afftin, alphabody, anticalin, peptide aptamer, armadillo repeat protein (ARM), atrimer, avimer, designed ankyrin repeat protein, antibody-drug conjugates (including antibodies against proteins, DARPins®), phenomers, knottins, Kunitz domain peptides, monobodies, and nanoCLAMPs; conjugate, ADC), antibody-peptide conjugate), oncolytic virus, genetic DNA modifying or editing agent, chimeric antigen receptor (CAR) The present invention also includes cells comprising an engineered T cell receptor (TCR-T), including a T cell immunotherapeutic, an NK cell immunotherapeutic, or a macrophage immunotherapeutic, or any combination thereof.

[0319] Exemplary Targets

[0320] In some embodiments, the one or more additional therapeutic agents include, for example, inhibitors, agonists, antagonists, ligands, modulators, stimulators, blockers, activators, or suppressors of a target (e.g., a polypeptide or polynucleotide), such as: 2'-5'-oligoadenylate synthetase (OAS1; NCBI Gene ID: 4938); 5'-3' exoribonuclease 1 (XRN1; NCBI Gene ID: 54464); 5'-nucleotidase ecto (NT5E, CD73; NCBI Gene ID: 4907); ABL proto-oncogene 1, non-receptor tyrosine kinase (ABL1, BCR-ABL, c-ABL, v-ABL; NCBI Gene ID: 25); Absentee in melanoma 2 (AIM2; NCBI Gene ID: 9447 acetyl-CoA acyltransferase 2 (ACAA2; NCBI Gene ID: 10499); acid phosphatase 3 (ACP3; NCBI Gene ID: 55); adenosine deaminase (ADA, ADA1; NCBI Gene ID: 100); adenosine receptors (e.g., ADORA1 (A1), ADORA2A (A2a, A2AR), ADORA2B (A2b, A2BR), ADORA3 (A3); NCBI Gene IDs: 134, 135, 136, 137); AKT serine / stearate leonine kinase 1 (AKT1, AKT, PKB; NCBI Gene ID: 207); alanyl aminopeptidase, membrane (ANPEP, CD13; NCBI Gene ID: 290); ALK receptor tyrosine kinase (ALK, CD242; NCBI Gene ID: 238); alpha-fetoprotein (AFP; NCBI Gene ID: 174); amine oxidase copper-containing (e.g., AOC1 (DAO1), AOC2, AOC3 (VAP1); NCBI Gene IDs: 26, 314, 8639) ; androgen receptor (AR; NCBI Gene ID: 367); angiopoietin (ANGPT1, ANGPT2; NCBI Gene ID: 284, 285); angiotensin II receptor type 1 (AGTR1; NCBI Gene ID: 185); angiotensinogen (AGT; NCBI Gene ID: 183); apolipoprotein A1 (APOA1; NCBI Gene ID: 335); apoptosis-inducing factor mitochondrial-associated 1 (AIFM1, AIF; NCBI Gene ID: 9131);Arachidonate 5-lipoxygenase (ALOX5; NCBI Gene ID: 240); asparaginase (ASPG; NCBI Gene ID: 374569); asteraid homolog 1 (ASTE1; NCBI Gene ID: 28990); ATM serine / threonine kinase (ATM; NCBI Gene ID: 472); ATP-binding cassette subfamily B member 1 (ABCB1, CD243, GP170; NCBI Gene ID: 5243); ATP-dependent Clp-protease (CLPP; NCBI Gene ID: 8192); ATR serine / threonine kinase (ATR; NCBI Gene ID: 545); AXL receptor tyrosine kinase (AXL; NCBI Gene ID: 558); B and T lymphocyte-associated (BTLA, CD272; NCBI Gene ID: 151888); baculovirus IAP repeat-containing proteins (BIRC2 (cIAP1), BIRC3 (cIAP2), XIAP (BIRC4, IAP3), BIRC5 (survivin); NCBI Gene IDs: 329, 330, 331, 332); basigin (Ok blood group) (BSG, CD147; NCBI Gene ID: 68 2); B-cell lymphoma 2 (BCL2; NCBI Gene ID: 596); BCL2-binding protein 3 (BBC3, PUMA; NCBI Gene ID: 27113); Bcl2-like (e.g., Bcl2L1 (Bcl-x), Bcl2L2 (BIM); Bcl-x; NCBI Gene ID: 598, 10018); beta 3-adrenergic receptor (ADRB3; NCBI Gene ID: 155); bone gamma-carboxyglutamic acid protein (BGLAP; NCBI Gene ID: 632); bone morphogenetic protein-10 ligand (BMP10; NCBI Gene ID: 27302); bradykinin receptors (e.g., BDKRB1, BDKRB2; NCBI Gene IDs: 623, 624); B-RAF (BRAF; NCBI Gene ID: 273); breakpoint cluster region (BCR; NCBI Gene ID: 613); bromodomain and ectodomain (BET) bromodomain-containing proteins (e.g., BRD2, BRD3, BRD4, BRDT; NCBI Gene IDs: 6046, 8019, 23476, 676); Bruton's tyrosine kinase (BTK; NCBI Gene ID: 695);Cadherins (e.g., CDH3 (p-cadherin), CDH6 (k-cadherin); NCBI Gene IDs: 1001, 1004); cancer / testis antigens (e.g., CTAG1A, CTAG1B, CTAG2; NCBI Gene IDs: 1485, 30848, 246100); cannabinoid receptors (e.g., CNR1 (CB1), CNR2 (CB2); NCBI Gene IDs: 1268, 1269); carbohydrate sulfotransferase 15 (CHST15; NCBI Gene ID: 51363); carbonic anhydrases (CA1, CA2, CA3, CA4, CA5A, CA5 B, CA6, CA7, CA8, CA9, CA10, CA11, CA12, CA13, CA14; NCBI Gene IDs: 759, 760, 761, 762, 763, 765, 766, 767, 768, 770, 771, 11238, 23632, 56934, 377677); carcinoembryonic antigen-related cell adhesion molecules (e.g., CEACAM3 (CD66d), CEACAM5 (CD66e), CEACAM6 (CD66c); NCBI Gene IDs: 1048, 1084, 4680); casein kinases (e.g., CSNK1A1 (CK1), CSNK2A1 (CK 2); NCBI Gene ID: 1452, 1457); caspases (e.g., CASP3, CASP7, CASP8; NCBI Gene ID: 836, 840, 841, 864); catenin β1 (CTNNB1; NCBI Gene ID: 1499); cathepsin G (CTSG; NCBI Gene ID: 1511); Cbl proto-oncogene B (CBLB, Cbl-b; NCBI Gene ID: 868); CC motif chemokine ligand 21 (CCL21; NCBI Gene ID: 6366); CC motif chemokine receptor 2 (CCR2; NCBI Gene ID: 7292 30); CC motif chemokine receptors (e.g., CCR3 (CD193), CCR4 (CD194), CCR5 (CD195), CCR8 (CDw198); NCBI Gene ID: 1232, 1233, 1234, 1237); CCAAT enhancer-binding protein alpha (CEBPA, CEBP; NCBI Gene ID: 1050); cell adhesion molecule 1 (CADM1; NCBI Gene ID: 23705); cell division cycle 7 (CDC7; NCBI Gene ID: 8317); cell communication network factor 2 (CCN2; NCBI Gene ID: 1490);Cereblon (CRBN; NCBI Gene ID: 51185); checkpoint kinases (e.g., CHEK1 (CHK1), CHEK2 (CHK2); NCBI Gene ID: 1111, 11200); cholecystokinin B receptor (CCKBR; NCBI Gene ID: 887); chorionic somatomammotropic hormone 1 (CSH1; NCBI Gene ID: 1442); claudins (e.g., CLDN6, CLDN18; NCBI Gene ID: 9074, 51208); cluster of differentiation markers - (e.g., CD1A, CD1C, CD1D, CD1E, CD2, CD3 alpha (TRA), CD3 beta (TRB), CD3 gamma (TRG), CD3 delta (TRD), CD4, CD8A, CD8B, CD19, CD20 (MS4A1), CD22, CD24, CD25 (IL2RA, TCGFR), CD28, CD33 (SIGLEC3), CD37, CD38, CD39 (ENTPD1), CD40 (TNFRSF5), CD44 (MIC4, PGP1), CD47 (IAP), CD48 ( BLAST1), CD52, CD55(DAF), CD58(LFA3), CD74, CD79a, CD79b, CD80(B7-1), CD84, CD86(B7-2), CD96(TACTILE), CD99(MIC2), CD115(CS F1R), CD116 (GMCSFR, CSF2RA), CD122 (IL2RB), CD123 (IL3RA), CD128 (IL8R1), CD132 (IL2RG), CD135 (FLT3), CD137 (TNFRSF9, 4-1BB), CD142 (TF, TFA), CD152 (CTLA4), CD160, CD182 (IL8R2), CD193 (CCR3), CD194 (CCR4), CD195 (CCR5), CD207, CD221 (IGF1R), CD222 (IGF2) R), CD223(LAG3), CD226(DNAM1), CD244, CD247, CD248, CD276(B7-H3), CD331(FGFR1), CD332(FGFR2), CD333(FGFR3), CD334(FGFR4);NCBI gene IDs: 909, 911, 912, 913, 914, 919, 920, 923, 925, 926, 930, 931, 933, 940, 941, 942, 945, 951, 952, 953, 958, 960, 961, 962, 965, 972, 973, 974, 1043, 1232, 1233, 1234, 1237, 1436, 1438, 1493, 1604, 2152, 2260, 2261, 2263, 2322, 3480, 3482, 3559, 3560, 3561, 3563, 3577, 3579, 3604, 3902 , 4267, 6955, 6957, 6964, 6965, 8832, 10666, 11126, 50489, 51744, 80381, 100133941); clusterin (CLU; NCBI Gene ID: 1191); coagulation factors (e.g., F7, FXA; NCBI Gene IDs: 2155, 2159); collagen type IV alpha chain (e.g., COL4A1, COL4A2, COL4A3, COL4A4, COL4A5; NCBI Gene IDs: 1282, 1284, 1285, 1286, 1287); collectin subfamily members colony-stimulating factors (e.g., CSF1 (MCSF), CSF2 (GMCSF), CSF3 (GCSF); NCBI Gene IDs: 1435, 1437, 1440); complement factors (e.g., C3, C5; NCBI Gene IDs: 718, 727); COP9 signalosome subunit 5 (COPS5; NCBI Gene ID: 10987); C-type lectin domain family members (e.g., CLEC4C (CD303), CLEC9A (CD370), CLEC12A (CD3 71); CD371; NCBI Gene ID: 160364, 170482, 283420; C-X-C motif chemokine ligand 12 (CXCL12; NCBI Gene ID: 6387); C-X-C motif chemokine receptors (CXCR1 (IL8R1, CD128), CXCR2 (IL8R2, CD182), CXCR3 (CD182, CD183, IP-10R), CXCR4 (CD184); NCBI Gene ID: 2833, 3577, 3579, 7852); cyclin D1 (CCND1, BCL1; NCBI Gene ID: 595);Cyclin-dependent kinases (e.g., CDK1, CDK2, CDK3, CDK4, CDK5, CDK6, CDK7, CDK8, CDK9, CDK10, CDK12; NCBI Gene IDs: 983, 1017, 1018, 1019, 1020, 1021, 1022, 1024, 1025, 8558, 51755); cyclin G1 (CCNG1; NCBI Gene ID: 900); cytochrome P450 family members (e.g., CYP2D6, CYP3A4, CYP11A1, CYP11B2, CYP17A1, CYP19A1, CYP51A1; NCBI Gene IDs: 1565, 1576, 1583, 1585, 1586, 1588, 1595); cytochrome p450 oxidoreductases; protein (POR; NCBI Gene ID: 5447); cytokine-induced SH2-containing protein (CISH; NCBI Gene ID: 1154); cytotoxic T-lymphocyte-associated protein 4 (CTLA4, CD152; NCBI Gene ID: 1493); DEAD-box helicases (e.g., DDX5, DDX6, DDX58; NCBI Gene IDs: 1655, 1656, 23586); delta-like canonical Notch ligands (e.g., DLL3, DLL4; NCBI Gene IDs: 10683, 54567); diabetic IAP-binding mitochondrial protein (Di ablo, SMAC; NCBI Gene ID: 56616); diacylglycerol kinase (e.g., DGKA, DGKZ; NCBI Gene ID: 1606, 8525); Dickkopf WNT signaling pathway inhibitors (e.g., DKK1, DKK3; NCBI Gene ID: 22943, 27122); dihydrofolate reductase (DHFR; NCBI Gene ID: 1719); dihydropyrimidine dehydrogenase (DPYD; NCBI Gene ID: 1806); dipeptidyl peptidase 4 (DPP4; NCBI Gene ID: 1803); discoid domain receptor tyrosine kinases (e.g., DDR1 (CD167), DDR2; CD167; NCBI Gene ID: 780, 4921); DNA-dependent protein kinase (PRKDC; NCBI Gene ID: 5591); DNA topoisomerases (e.g., TOP1, TOP2A, TOP2B, TOP3A, TOP3B; NCBI Gene ID: 7150, 7153, 7155, 7156, 8940); dopachrome tautomer (DCT; NCBI Gene ID: 1638); dopamine receptor D2 (DRD2; NCBI Gene ID: 1318); DO T1-like histone lysine methyltransferase (DOT1L; NCBI Gene ID: 84444); ectonucleotide pyrophosphatase / phosphodiesterase 3 (ENPP3, CD203c; NCBI Gene ID: 5169); EMAP-like 4 (EML4; NCBI Gene ID: 27436); endoglin (ENG; NCBI Gene ID: 2022); endoplasmic reticulum aminopeptidases (e.g., ERAP1, ERAP2; NCBI Gene IDs: 51752, 64167); enhancer of zeste2 polycomb repressive complex 2 subunit (EZH2;NCBI Gene ID: 2146); ephrin receptors (e.g., EPHA1, EPHA2, EPHA3, EPHA4, EPHA5, EPHA7, EPHB4; NCBI Gene IDs: 1969, 2041, 2042, 2043, 2044, 2045, 2050); ephrins (e.g., EFNA1, EFNA4, EFNB2; NCBI Gene IDs: 1942, 1945, 1948); epidermal growth factor receptors (e.g., ERBB1 (HER1, EGFR), ERBB1 variant III (EGFRvIII), ERBB2 (HER2, NEU, CD340), E RBB3 (HER3), ERBB4 (HER4) NCBI Gene ID: 1956, 2064, 2065, 2066); epithelial cell adhesion molecule (EPCAM; NCBI Gene ID: 4072); epidermal mitogen-activating factor (EPGN; NCBI Gene ID: 255324); eukaryotic translation elongation factors (e.g., EEF1A2, EEF2; NCBI Gene ID: 1917, 1938); eukaryotic translation initiation factors (e.g., EIF4A1, EIF5A; NCBI Gene ID: 1973, 1984); exopolitin-1 (XPO1; NCBI Gene ID: 7514); falcon Neissoid X receptor (NR1H4, FXR; NCBI Gene ID: 9971); Fa ligand (FASLG, FASL, CD95L, CD178, TNFSF6; NCBI Gene ID: 356); fatty acid amide hydrolase (FAAH; NCBI Gene ID: 2166); fatty acid synthase (FASN; FAS; NCBI Gene ID: 2194); Fc fragment of Ig receptor (e.g., FCER1A, FCGRT, FCGR3A (CD16); NCBI Gene IDs: 2205, 2214, 2217); Fc receptor-like 5 (FCRL5, CD307; NCBI Gene ID: 2209); Gene ID: 83416); fibroblast activation protein alpha (FAP; NCBI Gene ID: 2191); fibroblast growth factor receptors (e.g., FGFR1 (CD331), FGFR2 (CD332), FGFR3 (CD333), FGFR4 (CD334); NCBI Gene IDs: 2260, 2261, 2263, 2264); fibroblast growth factors (e.g., FGF1 (FGF alpha), FGF2 (FGF beta), FGF4, FGF5; NCBI Gene IDs: 2246, 2247, 2249, 2250); fibronectin 1 (FN1, MSF;NCBI Gene ID: 2335); fms-related receptor tyrosine kinases (e.g., FLT1 (VEGFR1), FLT3 (STK1, CD135), FLT4 (VEGFR2); NCBI Gene ID: 2321, 2322, 2324); fms-related receptor tyrosine kinase 3 ligand (FLT3LG; NCBI Gene ID: 2323); focal adhesion kinase 2 (PTK2, FAK1; NCBI Gene ID: 5747); folate hydrolase 1 (FOLH1, PSMA; NCBI Gene ID: 2346); folate receptor 1 (FOLR1; NCBI Gene ID: 23 48); forkhead box protein M1 (FOXM1; NCBI Gene ID: 2305); furin (furin, PACE; NCBI Gene ID: 5045); FYN tyrosine kinase (FYN, SYN; NCBI Gene ID: 2534); galectins (e.g., LGALS3, LGALS8 (PCTA1), LGALS9; NCBI Gene IDs: 3958, 3964, 3965); glucocorticoid receptor (NR3C1, GR; NCBI Gene ID: 2908); glucuronidase beta (GUSB; NCBI Gene ID: 2990); glutamate Glycogen synthase kinase 3 beta (GSK3B; NCBI gene ID: 2932); Glypican 3 (GPC3; NCBI gene ID: 2719); Gonadotropin-releasing hormone 1 (GNRH1; NCBI gene ID: 2796); Gonadotropin-releasing hormone receptor (GNRHR; NCBI gene ID: 2798); G PNMB glycoprotein NMB (GPNMB, osteoactivin; NCBI Gene ID: 10457); growth differentiation factor 2 (GDF2, BMP9; NCBI Gene ID: 2658); growth factor receptor-bound protein 2 (GRB2, ASH; NCBI Gene ID: 2885); guanylate cyclase 2C (GUCY2C, STAR, MECIL, MUCIL; NCBI Gene ID: 2984); H19 imprinted maternally expressed transcript (H19; NCBI Gene ID: 283120); HCK pro-oncogene, Src family tyrosine kinase (HCK;NCBI Gene ID: 3055); heat shock proteins (e.g., HSPA5 (HSP70, BIP, GRP78), HSPB1 (HSP27), HSP90B1 (GP96); NCBI Gene IDs: 3309, 3315, 7184); heme oxygenases (e.g., HMOX1 (HO1), HMOX2 (HO1); NCBI Gene IDs: 3162, 3163); heparanase (HPSE; NCBI Gene ID: 10855); hepatitis A virus cellular receptor 2 (HAVCR2, TIM3, CD366; NCBI Gene ID: 84868); hepatic growth factor (HGF; NCBI Gene ID: 3082); HERV-H LTR-associated 2 (HHLA2, B7-H7; NCBI Gene ID: 11148); histamine receptor H2 (HRH2; NCBI Gene ID: 3274); histone deacetylases (e.g., HDAC1, HDAC7, HDAC9; NCBI Gene IDs: 3065, 9734, 51564); HRas proto-oncogene, GTPase (HRAS; NCBI Gene ID: 3265); hypoxia-inducible factors (e.g., HIF1A, HIF2A (EPAS1); NCBI Gene IDs: 2034, 3091); I-Kappa-B kinase (IKK beta; NCBI Gene IDs: 3551, 3553); IKAROS family zinc finger (IKZF1 (LYF1), IKZF3; NCBI Gene IDs: 10320, 22806); immunoglobulins immunoglobulin superfamily member 11 (IGSF11; NCBI Gene ID: 152404); indoleamine 2,3-dioxygenase (e.g., IDO1, IDO2; NCBI Gene ID: 3620, 169355); inducible T cell costimulatory molecule (ICOS, CD278; NCBI Gene ID: 29851); inducible T cell costimulatory molecule ligand (ICOSLG, B7-H2; NCBI Gene ID: 23308); insulin-like growth factor receptor (e.g., IGF1R, IGF2R; NCBI Gene ID: 3480, 3482); insulin-like growth factor (e.g., IGF1, IGF2; NCBI Gene ID: 3479, 3481); insulin receptor (INSR, CD220; NCBI Gene ID: 3643);Integrin subunits (e.g., ITGA5 (CD49e), ITGAV (CD51), ITGB1 (CD29), ITGB2 (CD18, LFA1, MAC1), ITGB7; NCBI Gene ID: 3678, 3685, 3688, 3695, 3698); intercellular adhesion molecule 1 (ICAM1, CD54; NCBI Gene ID: 3383); interleukin-1 receptor-associated kinase 4 (IRAK4; NCBI Gene ID: 51135); interleukin receptors (e.g., IL2RA (TCGFR, CD25), IL2RB (CD 122), IL2RG (CD132), IL3RA, IL6R, IL13RA2 (CD213A2), IL22RA1; NCBI Gene IDs: 3598, 3559, 3560, 3561, 3563, 3570, 58985); interleukins (e.g., IL1A, IL1B, IL2, IL3, IL6 (HGF), IL7, IL8 (CXCL8), IL10 (TGIF), IL12A, IL12B, IL15, IL17A (CTLA8), IL23A, IL24, IL-29 (IFNL1); NCBI Gene IDs: 3552, 3553 , 3558, 3562, 3565, 3569, 3574, 3586, 3592, 3593, 3600, 3605, 11009, 51561, 282618); isocitrate dehydrogenase (NADP()1) (e.g., IDH1, IDH2; NCBI Gene ID: 3417, 3418); Janus kinase (e.g., JAK1, JAK2, JAK3; NCBI Gene ID: 3716, 3717, 3718); kallikrein-related peptidase 3 (KLK3; NCBI Gene ID: 354); killer cell immunoglobulin-like receptor, Ig domain and long cytoplasmic tails (e.g., KIR2DL1 (CD158A), KIR2DL2 (CD158B1), KIR2DL3 (CD158B), KIR2DL4 (CD158D), KIR2DL5A (CD158F), KIR2DL5B, KIR3DL1 (CD158E1), KIR3DL2 (CD158K), KIR3DP1 (CD158c), KIR2DS2 (CD158J); NCBI Gene IDs: 3802, 3803, 3804, 3805, 3811, 3812, 57292, 553128, 548594, 100132285);Killer cell lectin-like receptors (e.g., KLRC1 (CD159A), KLRC2 (CD159c), KLRC3, KLRRC4, KLRD1 (CD94), KLRG1, KLRK1 (NKG2D, CD314); NCBI Gene ID: 3821, 3822, 3823, 3824, 8302, 10219, 22914); kinase insert domain receptor (KDR, CD309, VEGFR2; NCBI Gene ID: 3791); kinesin family member 11 (KIF11; NCBI Gene ID: 3832); KiSS-1 metastasis suppressor (KiSS1; NCBI Gene ID: 3833); ID: 3814); KIT proto-oncogene, receptor tyrosine kinase (KIT, c-KIT, CD117; NCBI Gene ID: 3815); KRAS proto-oncogene, GTPase (KRAS; NCBI Gene ID: 3845); lactotransferrin (LTF; NCBI Gene ID: 4057); LCK pro-oncogene, Src family tyrosine kinase (LCK; NCBI Gene ID: 3932); LDL receptor-related protein 1 (LRP1, CD91, IGFBP3R; NCBI Gene ID: 4035); Leucine-rich repeat-containing 15 (LRRC15; NCBI Gene ID: 131578); leukocyte immunoglobulin-like receptors (e.g., LILRB1 (ILT2, CD85J), LILRB2 (ILT4, CD85D); NCBI Gene ID: 10288, 10859); leukotriene A4 hydrolase (LTA4H; NCBI Gene ID: 4048); linker for activation of T cells (LAT; NCBI Gene ID: 27040); luteinizing hormone / chorionic gonadotropin receptor (LHCGR; NCBI Gene ID: 1 D:3973); LY6 / PLAUR domain-containing 3 (LYPD3; NCBI Gene ID: 27076); lymphocyte activation 3 (LAG3; CD223; NCBI Gene ID: 3902); lymphocyte antigens (e.g., LY9 (CD229), LY75 (CD205); NCBI Gene ID: 4063, 17076); LYN proto-oncogene, Src family tyrosine kinase (LYN; NCBI Gene ID: 4067); lymphocyte cytosolic protein 2 (LCP2; NCBI Gene ID: 3937); Lysine demethylase 1A (KDM1A; NCBI Gene ID: 23028); Lysophosphatidic acid receptor 1 (LPAR1, EDG2, LPA1, GPR26; NCBI Gene ID: 1902); Lysyl oxidase (LOX; NCBI Gene ID: 4015); Lysyl oxidase-like 2 (LOXL2, NCBI Gene ID: 4017); Macrophage migration inhibitory factor (MIF, GIF; NCBI Gene ID: 4282); Macrophage stimulating 1 receptor (MST1R, CD136; NCBI Gene ID: 4486);MAGE family members (e.g., MAGEA1, MAGEA2, MAGEA2B, MAGEA3, MAGEA4, MAGEA5, MAGEA6, MAGEA10, MAGEA11, MAGEC1, MAGEC2, MAGED1, MAGED2; NCBI Gene IDs: 4100, 4101, 4102, 4103, 4104, 4105, 4109, 4110, 9500, 9947, 10916, 51438, 266740); major histocompatibility complex (e.g., HLA-A, HLA-E, HLA-F, HLA-G; NCBI Gene IDs: 3105, 3133) , 3134, 3135); major vault protein (MVP, VAULT1; NCBI Gene ID: 9961); MALT1 paracaspase (MALT1; NCBI Gene ID: 10892); MAPK-activated protein kinase 2 (MAPKAPK2; NCBI Gene ID: 9261); MAPK-interacting serine / threonine kinases (e.g., MKNK1, MKNK2; NCBI Gene ID: 2872, 8569); matrix metallopeptidases (e.g., MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11) , MMP12, MMP13, MMP14, MMP15, MMP16, MMP17, MMP19, MMP20, MMP21, MMP24, MMP25, MMP26, MMP27, MMP28; NCBI Gene IDs: 4312, 4313, 4314, 4316, 4317, 4318, 4319, 4320, 4321, 4322, 4323, 4324, 4325, 4326, 4327, 9313, 10893, 56547, 64066, 64386, 79148, 118856); MCL1 apoptosis regulator, BCL2 family member (MCL1; NCBI I Gene ID: 4170); MDM2 oncogene (MDM2; NCBI Gene ID: 4193); MDM4 regulator of p53 (MDM4; BMFS6; NCBI Gene ID: 4194); mechanistic target of rapamycin kinase (MTOR, FRAP1; NCBI Gene ID: 2475); melan-A (MLANA; NCBI Gene ID: 2315); melanocortin receptors (MC1R, MC2R; NCBI Gene IDs: 4157, 4148); MER proto-oncogene, tyrosine kinase (MERTK; NCBI Gene ID: 10461); mesothelin (MSLN;NCBI Gene ID: 10232); MET proto-oncogene, receptor tyrosine kinase (MET, c-Met, HGFR; NCBI Gene ID: 4233); methionyl aminopeptidase 2 (METAP2, MAP2; NCBI Gene ID: 10988); MHC class I polypeptide-related sequence (e.g., MICA, MICB; NCBI Gene IDs: 4277, 100507436); mitogen-activated protein kinase (e.g., MAPK1 (ERK2), MAPK3 (ERK1), MAPK8 (JNK1), MAPK9 (JNK) K2), MAPK10 (JNK3), MAPK11 (p38 beta), MAPK12; NCBI Gene ID: 5594, 5595, 5599, 5600, 5601, 5602, 819251); mitogen-activated protein kinase kinase kinases (e.g., MAP3K5 (ASK1), MAP3K8 (TPL2, AURA2); NCBI Gene ID: 4217, 1326); mitogen-activated protein kinase kinase kinase kinase 1 (HPK1; NCBI Gene ID: 11184); mitogen-activated protein kinase MAP2K1 (MEK1), MAP2K2 (MEK2), MAP2K7 (MEK7); NCBI Gene IDs: 5604, 5605, 5609); MPL proto-oncogene, thrombopoietin receptor (MPL; NCBI Gene ID: 4352); mucins (e.g., MUC1 (including its splice variants (e.g., MUC1 / A, C, D, X, Y, Z, and REP)), MUC5AC, MUC16 (CA125); NCBI Gene IDs: 4582, 4586, 94025); MYC proto-oncogene, bH LH transcription factor (MYC; NCBI Gene ID: 4609); myostatin (MSTN, GDF8; NCBI Gene ID: 2660); myristoylated alanine-rich protein kinase C substrate (MARCKS; NCBI Gene ID: 4082); natriuretic peptide receptor 3 (NPR3; NCBI Gene ID: 4883); natural killer cytotoxicity receptor 3 ligand 1 (NCR3LG1, B7-H6; NCBI Gene ID: 374383); necdin, a MAGE family member (NDN; NCBI Gene ID: 4692);Nectin cell adhesion molecules (e.g., nectin2 (CD112, PVRL2), nectin4 (PVRL4); NCBI Gene IDs: 5819, 81607); neural cell adhesion molecule 1 (NCAM1, CD56; NCBI Gene ID: 4684); neuropilins (e.g., NRP1 (CD304, VEGF165R), NRP2 (VEGF165R2); NCBI Gene IDs: 8828, 8829); neurotrophic receptor tyrosine kinases (e.g., NTRK1 (TRKA), NTRK2 (TRKB), NTRK3 (TRKC); NCBI Gene IDs: 4914, 4915, 4916); NFKB-activating protein (NKAP; NCBI Gene ID: 79576); NIMA-related kinase 9 (NEK9; NCBI Gene ID: 91754); NLR family pyrin domain-containing 3 (NLRP3, NALP3; NCBI Gene ID: 114548); Notch receptors (e.g., notch1, notch2, notch3, notch4; NCBI Gene IDs: 4851, 4853, 4854, 4855); NRAS proto-oncogene, GTPase (NRAS; NCBI Gene ID: 4893); nuclear factor n...

Claims

1. 1. An isolated antibody that binds to human CCR8, said antibody comprising: An isolated antibody comprising a heavy chain complementarity determining region 1 (HCDR1) comprising the amino acid sequence of SEQ ID NO: 60, a heavy chain complementarity determining region 2 (HCDR2) comprising the amino acid sequence of SEQ ID NO: 61, 72 or 78, a heavy chain complementarity determining region 3 (HCDR3) comprising the amino acid sequence of SEQ ID NO: 62, a light chain complementarity determining region 1 (LCDR1) comprising the amino acid sequence of SEQ ID NO: 63, 73 or 79, a light chain complementarity determining region 2 (LCDR2) comprising the amino acid sequence of SEQ ID NO: 64, and a light chain complementarity determining region 3 (LCDR3) comprising the amino acid sequence of SEQ ID NO:

65.

2. The antibody (i) a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 56 and a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO: 57; (ii) a heavy chain variable region (VH) comprising an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 68, and a light chain variable region (VL) comprising an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 69; or (iii) a heavy chain variable region (VH) comprising an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 74, and a light chain variable region (VL) comprising an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 75; The isolated antibody of claim 1.

3. The antibody (i) a VH comprising the amino acid sequence of SEQ ID NO: 56 and a VL comprising the amino acid sequence of SEQ ID NO: 57; (ii) a VH comprising the amino acid sequence of SEQ ID NO: 68 and a VL comprising the amino acid sequence of SEQ ID NO: 69; or (iii) VH comprising the amino acid sequence of SEQ ID NO: 74 and VL comprising the amino acid sequence of SEQ ID NO: 75; The isolated antibody of claim 1.

4. The isolated antibody of any one of claims 1 to 3, wherein the antibody is a monoclonal antibody.

5. The isolated antibody of any one of claims 1 to 3, wherein the antibody is a humanized antibody.

6. The isolated antibody of any one of claims 1 to 3, wherein the antibody is a full-length antibody.

7. The isolated antibody of any one of claims 1 to 6, wherein the antibody is an IgG1 or IgG3 antibody.

8. The antibody (i) a heavy chain (HC) comprising the amino acid sequence of SEQ ID NO: 58 and a light chain (LC) comprising the amino acid sequence of SEQ ID NO: 59; (ii) a heavy chain (HC) comprising an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 70, and a light chain (LC) comprising an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 71; or (iii) a heavy chain (HC) comprising an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 76, and a light chain (LC) comprising an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 77; The isolated antibody of claim 1.

9. 2. The isolated antibody of claim 1, wherein the antibody comprises an HC comprising the amino acid sequence of SEQ ID NO: 58 and an LC comprising the amino acid sequence of SEQ ID NO:

59.

10. 2. The isolated antibody of claim 1, wherein the antibody comprises an HC comprising the amino acid sequence of SEQ ID NO: 70 and an LC comprising the amino acid sequence of SEQ ID NO:

71.

11. 2. The isolated antibody of claim 1, wherein the antibody comprises an HC comprising the amino acid sequence of SEQ ID NO: 76 and an LC comprising the amino acid sequence of SEQ ID NO:

77.

12. The isolated antibody of any one of claims 1 to 11, wherein the antibody comprises at least one modification that enhances cell killing.

13. 13. The isolated antibody of claim 12, wherein the enhanced cell killing is enhanced antibody-dependent cellular cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC).

14. 13. The isolated antibody of claim 12, wherein the at least one modification is afucosylation.

15. 15. The isolated antibody of any one of claims 1 to 14, wherein the antibody comprises a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 56 and a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO:

57.

16. 15. The isolated antibody of any one of claims 1 to 14, wherein the antibody comprises a heavy chain variable region (VH) comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO: 68, and a light chain variable region (VL) comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:

69.

17. 15. The isolated antibody of any one of claims 1 to 14, wherein the antibody comprises a heavy chain variable region (VH) comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO: 74, and a light chain variable region (VL) comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:

75.

18. 15. The isolated antibody of any one of claims 1 to 14, wherein the antibody comprises a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 68 and a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO:

69.

19. 15. The isolated antibody of any one of claims 1 to 14, wherein the antibody comprises a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 74 and a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO:

75.

20. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO: 70 and a light chain comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:

71.

21. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO: 76 and a light chain comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:

77.

22. 2. The isolated antibody of claim 1, wherein the HCDR1 comprises the amino acid sequence of SEQ ID NO: 60, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 61, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 62, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 63, the LCDR2 comprises the amino acid sequence of SEQ ID NO: 64, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 65, and the antibody is afucosylated.

23. 2. The isolated antibody of claim 1, wherein the HCDR1 comprises the amino acid sequence of SEQ ID NO: 60, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 72, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 62, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 73, the LCDR2 comprises the amino acid sequence of SEQ ID NO: 64, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 65, and the antibody is afucosylated.

24. 2. The isolated antibody of claim 1, wherein the HCDR1 comprises the amino acid sequence of SEQ ID NO: 60, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 78, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 62, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 79, the LCDR2 comprises the amino acid sequence of SEQ ID NO: 64, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 65, and the antibody is afucosylated.

25. 2. The isolated antibody of claim 1, wherein the antibody comprises a VH comprising the amino acid sequence of SEQ ID NO: 56 and a VL comprising the amino acid sequence of SEQ ID NO: 57, and the antibody is afucosylated.

26. 2. The isolated antibody of claim 1, wherein the antibody comprises a VH comprising the amino acid sequence of SEQ ID NO: 68 and a VL comprising the amino acid sequence of SEQ ID NO: 69, and the antibody is afucosylated.

27. 2. The isolated antibody of claim 1, wherein the antibody comprises a VH comprising the amino acid sequence of SEQ ID NO: 74 and a VL comprising the amino acid sequence of SEQ ID NO: 75, and the antibody is afucosylated.

28. 2. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 58 and a light chain comprising the amino acid sequence of SEQ ID NO: 59, and the antibody is afucosylated.

29. 2. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 70 and a light chain comprising the amino acid sequence of SEQ ID NO: 71, and the antibody is afucosylated.

30. 2. The isolated antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 76 and a light chain comprising the amino acid sequence of SEQ ID NO: 77, and the antibody is afucosylated.

31. 13. The isolated antibody of claim 12, wherein the at least one modification is one or more heavy chain constant region mutations at one or more positions selected from L234, L235, G236, S239, F243, H268, D270, R292, S298, Y300, V305, K326, A330, I332, E333, K334, and P396.

32. 32. The isolated antibody of claim 31 , wherein the one or more heavy chain constant region mutations are one or more mutations selected from S239D, S239M, F243L, H268D, D270E, R292P, S298A, Y300L, V305I, K326D, A330L, A330M, I332E, E333A, K334A, K334E, and P396L.

33. 32. The isolated antibody of claim 31 , wherein the one or more heavy chain constant region mutations are selected from F243L / R292P / Y300L / V305I / P396L, S239D / I332E, S239D / I332E / A330L, S298A / E333A / K334A, L234Y / L235Q / G236W / S239M / H268D / D270E / S298A, and D270E / K326D / A330M / K334E.

34. 13. The isolated antibody of claim 12, wherein the at least one modification is galactosylation.

35. The antibody has an affinity (K Ｄ 35. The isolated antibody of any one of claims 1 to 34, which binds to human CCR8 at the CCR8 domain.

36. The antibody has an on-cell affinity (K Ｄ 35. The isolated antibody of any one of claims 1 to 34, which binds to human CCR8 at the CCR8 domain.

37. The affinity (K Ｄ 36. The isolated antibody of claim 35, wherein the affinity of the antibody to the target protein is determined by an equilibrium binding displacement assay.

38. The isolated antibody of claim 36, wherein the on-cell affinity (K D ) is determined by an equilibrium binding exclusion assay.

39. An isolated nucleic acid encoding any one of the amino acid sequences of the isolated antibody of any one of claims 1 to 38.

40. 40. A vector comprising the isolated nucleic acid of claim 39.

41. A host cell comprising the vector of claim 40.

42. A host cell expressing the antibody of any one of claims 1 to 38.

43. 43. A method for producing an antibody that binds to CCR8, comprising culturing a host cell according to claim 41 or 42 under conditions suitable for expressing said antibody.

44. 40. Use of the isolated antibody of any one of claims 1 to 38 for the preparation of a medicament in the treatment of cancer.

45. A composition for treating cancer, comprising the antibody of any one of claims 1 to 38, wherein the composition is administered to a subject with cancer.

46. 39. A method for determining CCR8 expression in a sample from a subject having cancer as an indicator for selecting a subject to be treated with the antibody of any one of claims 1 to 38, the method comprising detecting CCR8 expression in a sample from the subject.

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