Anti-CCR8 antibodies and uses thereof

Abstract

Provided herein are isolated monoclonal antibodies to CCR8 antibodies, antigen binding fragments thereof, mutants thereof, variants thereof, and uses thereof for the treatment of diseases such as cancer. The mutant antibodies have complementarity determining region 3 modified to eliminate post-translational modifications and improve the stability and potency of the antibodies. The variant antibodies have different levels of defucosylation. Also provided herein are methods of using the anti-CCR8 antibodies, including methods of treating cancer. Also provided are polynucleotides encoding the heavy or light chain or antigen-binding portion thereof described herein, as well as vectors, particularly expression vectors, comprising the polynucleotides described herein.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims the benefit of priority under 35 U.S.C. §119(e) to U.S. Provisional Application No. 63 / 399,483, filed on August 19, 2022. The disclosure of the prior application is considered part of the disclosure of the present application and is incorporated by reference in its entirety into the disclosure of the present application.

[0003] Incorporation of Sequence Listing

[0004] The material in the attached sequence listing is hereby incorporated into this application by reference.The attached sequence listing xml file named 148640-004502_SL.xml was created on August 18, 2023 and is 39,844 bytes. Summary of the Invention

[0005] The present invention relates to isolated anti-chemokine (C-C motif) receptor 8 (CCR8) monoclonal antibodies or antigen-binding fragments thereof, nucleic acids encoding the antibodies and expression vectors, recombinant cells containing the vectors, and compositions comprising the antibodies. Also provided are methods for preparing the antibodies and methods for using the antibodies to treat diseases, including cancer and / or related complications.

[0006] CCR8 is a chemokine receptor that mediates cell migration under CCL1 or CCL18 gradients (Islam et al., JEM [Journal of Experimental Medicine] 210 (10): 1889-1898 (2013)). Recently, CCR8 has been identified as a highly specific cell surface marker of tumor-infiltrating regulatory T cells (TITR) in human cancers. Compared with Treg in the circulation, its expression in Treg present in the tumor is significantly higher, and it is not expressed or expressed very low on other T cell populations (cytotoxic T cells or effector T cells, respectively). In addition, CCR8 is mainly expressed on highly immunosuppressive Tregs that express FoxP3 high, CD25 high, TIGIT+, LAG3+ and release high IL-10 and TGF-β. Exhaustion of CCR8+Treg will reduce immunosuppressive cytokines and regulate the tumor-promoting microenvironment to restore anti-tumor immunity. Interestingly, in patients with breast cancer or pancreatic cancer, a high number of CCR8+Treg is associated with a more advanced stage of the disease and may reduce overall survival. Therefore, CCR8 is an ideal cancer immunotherapy target for treating and potentially curing CCR8-positive cancers.

[0007] Aspects of the invention described herein relate to humanized antibodies directed against CCR8, mutants and variants thereof, and their use in treating diseases such as cancer.

[0008] In one embodiment, the present invention provides an isolated monoclonal antibody or antigen-binding fragment thereof, comprising an HCVR comprising an HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), an HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and an HCVR CDR3 sequence having the amino acid sequence of TRGSEQQYYYAMDY (SEQ ID NO: 19); and an LCVR comprising an LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), an LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and an LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15), wherein the antibody or antigen-binding fragment thereof specifically binds CCR8. In a specific embodiment thereof, the antibody comprises the HCVR of SEQ ID NO: 36 and the LCVR of SEQ ID NO: 32. In a specific embodiment thereof, the antibody is Ab001_M4 and comprises the heavy chain of SEQ ID NO:6 and the light chain of SEQ ID NO:2.

[0009] In another embodiment, the present invention provides an isolated monoclonal antibody or antigen-binding fragment thereof, comprising: a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 6; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 2.

[0010] In another embodiment, the present invention provides an isolated monoclonal antibody or antigen-binding fragment thereof comprising (a) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSEQNYYYAMDY (SEQ ID NO: 16); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (b) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a LCVR CDR3 sequence having the amino acid sequence of TRGSEQNYYYAMDY (SEQ ID NO: 16). NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSENNYYYAMDY (SEQ ID NO: 17); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (c) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSENNYYYAMDY (SEQ ID NO: 18); and a LCVR comprising a LCVR having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13). CDR1 sequence, a LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO: 15);or (d) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSDQNYYYAMDY (SEQ ID NO: 20); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (e) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a LCVR CDR3 sequence having the amino acid sequence of TRGSDNQYYYAMDY (SEQ ID NO: 21). NO: 21); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (f) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSDQQYYYAMDY (SEQ ID NO: 22);and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO: 15), wherein the antibody or antigen-binding fragment thereof specifically binds CCR8. In certain embodiments thereof, the antibody comprises (a) a HCVR of SEQ ID NO: 33 and a LCVR of SEQ ID NO: 32, (b) a HCVR of SEQ ID NO: 34 and a LCVR of SEQ ID NO: 32, (c) a HCVR of SEQ ID NO: 35 and a LCVR of SEQ ID NO: 32, (d) a HCVR of SEQ ID NO: 37 and a LCVR of SEQ ID NO: 32, (e) a HCVR of SEQ ID NO: 38 and a LCVR of SEQ ID NO: 32, or (f) a HCVR of SEQ ID NO: 39 and a LCVR of SEQ ID NO: 32. In a specific embodiment thereof, the antibody is (a) Ab001_M1 and comprises a heavy chain of SEQ ID NO: 3 and a light chain of SEQ ID NO: 2, (b) Ab001_M2 and comprises a heavy chain of SEQ ID NO: 4 and a light chain of SEQ ID NO: 2, (c) Ab001_M3 and comprises a heavy chain of SEQ ID NO: 5 and a light chain of SEQ ID NO: 2, (d) Ab001_M5 and comprises a heavy chain of SEQ ID NO: 7 and a light chain of SEQ ID NO: 2, (e) Ab001_M6 and comprises a heavy chain of SEQ ID NO: 8 and a light chain of SEQ ID NO: 2, or (f) Ab001_M7 and comprises a heavy chain of SEQ ID NO: 9 and a light chain of SEQ ID NO: 2.;

[0011] In another embodiment, the present invention provides an isolated monoclonal antibody or antigen-binding fragment thereof, comprising: (a) a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 3; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 2; (b) a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 4; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 2; (c) a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 5; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 2; (d) a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 7; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 8. NO:2; (e) a HC comprising a sequence at least 80% identical to SEQ ID NO:8 and having the antigen binding specificity thereof; and a LC comprising a sequence at least 80% identical to SEQ ID NO:2 and having the antigen binding specificity thereof; or (f) a HC comprising a sequence at least 80% identical to SEQ ID NO:9 and having the antigen binding specificity thereof; and a LC comprising a sequence at least 80% identical to SEQ ID NO:2 and having the antigen binding specificity thereof, wherein the antibody or antigen binding fragment thereof specifically binds to CCR8.

[0012] In one aspect, the HCVR sequence is SEQ ID NO:33 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:34 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:35 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:37 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:38 and / or the LCVR sequence is SEQ ID NO:32, or the HCVR sequence is SEQ ID NO:39 and / or the LCVR sequence is SEQ ID NO:32. In some aspects, the HCVR sequence is SEQ ID NO:33 and the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:34 and the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:35 and the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:37 and the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:38 and the LCVR sequence is SEQ ID NO:32, or the HCVR sequence is SEQ ID NO:39 and the LCVR sequence is SEQ ID NO:32.

[0013] In another aspect, the present invention provides an isolated monoclonal antibody comprising: (a) a heavy chain comprising SEQ ID NO: 3 and a light chain comprising SEQ ID NO: 2; (b) a heavy chain comprising SEQ ID NO: 4 and a light chain comprising SEQ ID NO: 2; (c) a heavy chain comprising SEQ ID NO: 5 and a light chain comprising SEQ ID NO: 2; (d) a heavy chain comprising SEQ ID NO: 7 and a light chain comprising SEQ ID NO: 2; (e) a heavy chain comprising SEQ ID NO: 8 and a light chain comprising SEQ ID NO: 2; (f) a heavy chain comprising SEQ ID NO: 9 and a light chain comprising SEQ ID NO: 2, wherein the antibody specifically binds to CCR8.

[0014] In another aspect, the antibody is a humanized antibody. In one aspect, its antigen-binding fragment is Fab, Fab', F(ab')2, F d , single-chain Fv or scFv, disulfide-linked F v, V-NAR domain, IgNar, intrabody, IgGACH2, minibody, F(ab')3, tetrabody, triabody, diabody, single domain antibody, DVD-Ig, Fcab, mAb2, (scFv)2 or scFv-Fc. In other aspects, the monoclonal antibody or its antigen-binding fragment has a K of less than about 25nM, 20nM, 15nM, 10nM, 5nM, 2nM, 1nM, 0.5nM or 0.01nM. d Binds to human CCR8. In one aspect, the antibody induces antibody-dependent cell-mediated cytotoxicity (ADCC). In another aspect, the antibody has an afucosylation level of at least 50%. In various aspects, the antibody has an afucosylation level of at least 85%.

[0015] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of any of the isolated monoclonal antibodies or antigen-binding fragments described herein, thereby treating the cancer in the subject.

[0016] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of at least one isolated monoclonal antibody or antigen-binding fragment thereof as provided herein. In one aspect, the method comprises administering an antibody comprising a HCVR comprising a CDR1 sequence comprising the amino acid sequence GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence comprising the amino acid sequence IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence comprising the amino acid sequence TRGSEQQYYYAMDY (SEQ ID NO: 19); and a LCVR comprising a LCVR CDR1 sequence comprising the amino acid sequence QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence comprising the amino acid sequence YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence comprising the amino acid sequence MQHLEYPFT (SEQ ID NO: 15), thereby treating cancer in the subject. In a specific embodiment thereof, the antibody comprises a HCVR of SEQ ID NO: 36 and a LCVR of SEQ ID NO: 32. In a specific embodiment thereof, the antibody is Ab001_M4 and comprises the heavy chain of SEQ ID NO:6 and the light chain of SEQ ID NO:2.

[0017] In one aspect, the cancer is a blood cancer. In some aspects, the blood cancer is selected from the group consisting of acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia, chronic myelogenous leukemia, hairy cell leukemia, AIDS-related lymphoma, cutaneous T-cell lymphoma, Hodgkin's lymphoma, Hodgkin's lymphoma, mycosis fungoides, non-Hodgkin's lymphoma, primary central nervous system lymphoma, Sezary syndrome, cutaneous T-cell lymphoma, Waldenstrom's macroglobulinemia, diffuse large B-cell lymphoma (DLBCL), and multiple myeloma.

[0018] On the other hand, cancer is a solid tumor. In some aspects, the solid tumor is selected from the group consisting of: breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colorectal cancer, renal cancer, urothelial cancer, ovarian cancer, endometrial cancer, uterine cancer, liver cancer, pancreatic cancer, bile duct cancer, gastric cancer, gastroesophageal cancer, esophageal cancer, cervical cancer, squamous cell carcinoma, prostate cancer or bladder cancer. On the one hand, the method further includes administering a chemotherapeutic agent, an anti-angiogenic agent, a growth inhibitor, an immuno-oncology agent, a checkpoint inhibitor and / or an anti-tumor composition to the patient.

[0019] In additional embodiments, the present invention provides polynucleotides encoding the heavy or light chain, or antigen-binding portion thereof, of any of the antibodies or antigen-binding fragments thereof described herein.

[0020] In one embodiment, the present invention provides a vector comprising a polynucleotide described herein, wherein the vector is an expression vector selected from the group consisting of a mammalian expression vector, a yeast expression vector, an insect expression vector, and a bacterial expression vector. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 is a graph illustrating the ADCC potency of antibodies with different levels of defucosylation.

[0022] Figure 2A-2C Deamination of the Ab001_Wt antibody is shown. Figure 2A The graph shows the results of a binding assay using CHO-hCCR8 cells. Figure 2B The ADCC reporter assay results are shown. Figure 2C Peptide mapping results are shown.

[0023] Figure 3A-Figure 3B The activity of Ab001_Wt antibody over time is shown. Figure 3A The results of binding assays over time are shown graphically. Figure 3B The ADCC reporter assay results over time are shown graphically.

[0024] Figures 4A-4CGraphed are the results of binding assays using CHO.hCCR8 cells for AbOOl_M1-M4 compared to AbOOl_Wt. Figure 4A The graph shows the binding assay results of Ab001_M1 and Ab001_M2 compared to Ab001_Wt. Figure 4B The graph shows the binding assay results of Ab001_M3 and Ab001_M4 compared to Ab001_Wt. Figure 4C It shows that Figure 4A and Figure 4B Table of EC50 calculated in .

[0025] Figures 5A-5C Graph depicts ADCC reporter assay results obtained using CHO.hCCR8 as target cells for AbOOl_M1-M4 compared to AbOOl_Wt. Figure 5A Shown are the ADCC reporter assay results for Ab001_M1 and Ab001_M2 compared to Ab001_Wt. Figure 5B Shown are the ADCC reporter assay results for Ab001_M3 and Ab001_M4 compared to Ab001_Wt. Figure 5C It shows that Figure 5A and Figure 5B Table of EC50 calculated in . DETAILED DESCRIPTION

[0026] Aspects of the invention described herein relate to humanized antibodies directed against CCR8 and their use in treating diseases such as cancer.

[0027] Before describing the compositions and methods of the present invention, it should be understood that the present invention is not limited to the specific compositions, methods and experimental conditions described, as such compositions, methods and conditions may vary. It should also be understood that the terminology used herein is for the purpose of describing specific embodiments only and is not intended to be limiting, as the scope of the present invention will be defined only in the appended claims.

[0028] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "the method" includes one or more methods and / or steps of the type described herein that will become apparent to those skilled in the art upon reading this disclosure.

[0029] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

[0030] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, it is understood that modifications and variations are encompassed within the spirit and scope of this disclosure. Preferred methods and materials are now described.

[0031] Unless otherwise indicated, any numerical value such as concentration or concentration range described herein is understood to be modified by the term "about" in all cases. Therefore, numerical values ​​typically include ± 10% of the listed value. For example, a concentration of 1 mg / mL includes 0.9 mg / mL to 1.1 mg / mL. Similarly, a concentration range of 1% to 10% (w / v) includes 0.9% (w / v) to 11% (w / v). Unless the context clearly indicates otherwise, as used herein, the use of numerical ranges clearly includes all possible subranges, all individual numerical values ​​within the range, including integers and fractions of values ​​within such ranges.

[0032] Unless otherwise indicated, the term "at least" preceding a series of elements should be understood to refer to every element in the series. Those skilled in the art will recognize or be able to ascertain many equivalents to the specific embodiments of the invention described herein using only routine experimentation. Such equivalents are intended to be encompassed by the present invention.

[0033] As used herein, the terms "comprises," "comprising," "includes," "including," "has," "having," "contains," or "containing," or any other variation thereof, will be understood to imply the inclusion of the stated integer or groups of integers, but not the exclusion of any other integer or groups of integers, and are intended to be non-exclusive or open-ended. For example, a composition, mixture, process, method, article, or apparatus that comprises a list of elements is not necessarily limited to only those elements but may include other elements not expressly listed or inherent to such composition, mixture, process, method, article, or apparatus. Furthermore, unless expressly stated to the contrary, "or" refers to an inclusive or and not an exclusive or. For example, any of the following satisfies condition A or B: A is true (or present) and B is false (or absent), A is false (or absent) and B is true (or present), and both A and B are true (or present).

[0034] As used herein, the connecting term "and / or" between multiple listed elements should be understood to cover both individual and combined options. For example, where two elements are connected by "and / or," the first option refers to the applicability of the first element without the second element. The second option refers to the applicability of the second element without the first element. The third option refers to the applicability of the first element and the second element together. Any of these options should be understood to fall within this meaning and therefore meet the requirements of the term "and / or" as used herein. The simultaneous applicability of more than one option should also be understood to fall within this meaning and therefore meet the requirements of the term "and / or."

[0035] As used herein, the term "consists of" or variations such as "consist of" or "consisting of" as used throughout the specification and claims indicates that any recited integer or group of integers is included, but additional integers or groups of integers cannot be added to the specified method, structure, or composition.

[0036] As used herein, the term "consists essentially of," or variations such as "consist essentially of," or "consisting essentially of," as used throughout the specification and claims, includes any recited integer or group of integers, and optionally includes any recited integer or group of integers that does not materially change the basic or novel characteristic of the specified method, structure, or composition. See MPEP §2111.03.

[0037] The present invention is based on the recognition of deficiencies within the anti-CCR8 antibodies disclosed in International Publication No. WO 2023 / 020621, particularly the antibody defined as H11-10 having a heavy chain of SEQ ID NO: 1 and a light chain of SEQ ID NO: 2. These deficiencies include asparaginyl residues in the antigen-binding complementarity-determining region (CDR) 3 of the HCVR, which are susceptible to deamidation and loss of activity over time, and fucosylation in the Fc region, which inhibits antibody-dependent cellular cytotoxicity (ADCC). Due to the critical nature of CDRs for antigen binding, any modification of these asparaginyl residues in the HCVR CDR3 may reduce the effectiveness of the antibody in CCR8 target engagement.

[0038] In one embodiment, the invention provides an isolated monoclonal antibody or antigen-binding fragment thereof, comprising a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSEQQYYYAMDY (SEQ ID NO: 19); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVRCDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15), wherein the antibody or antigen-binding fragment thereof specifically binds CCR8. In specific embodiments thereof, the antibody comprises the HCVR of SEQ ID NO: 36 and / or the LCVR of SEQ ID NO: 32, particularly the HCVR of SEQ ID NO: 36 and the LCVR of SEQ ID NO: 32. In a specific embodiment thereof, the antibody is Ab001_M4 and comprises the heavy chain of SEQ ID NO: 6 and the light chain of SEQ ID NO: 2.

[0039] In another embodiment, the present invention provides an isolated monoclonal antibody or antigen-binding fragment thereof, comprising: a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 6; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 2.

[0040] In another embodiment, the present invention provides an isolated monoclonal antibody or antigen-binding fragment thereof comprising (a) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSEQNYYYAMDY (SEQ ID NO: 16); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (b) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a LCVR CDR3 sequence having the amino acid sequence of TRGSEQNYYYAMDY (SEQ ID NO: 16). NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSENNYYYAMDY (SEQ ID NO: 17); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (c) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSENNYYYAMDY (SEQ ID NO: 18); and a LCVR comprising a LCVR having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13). CDR1 sequence, a LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO: 15);or (d) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSDQNYYYAMDY (SEQ ID NO: 20); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (e) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a LCVR CDR3 sequence having the amino acid sequence of TRGSDNQYYYAMDY (SEQ ID NO: 21). NO: 21); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (f) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSDQQYYYAMDY (SEQ ID NO: 22);and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO: 15), wherein the antibody or antigen-binding fragment thereof specifically binds CCR8. In certain embodiments thereof, the antibody comprises (a) a HCVR of SEQ ID NO: 33 and a LCVR of SEQ ID NO: 32, (b) a HCVR of SEQ ID NO: 34 and a LCVR of SEQ ID NO: 32, (c) a HCVR of SEQ ID NO: 35 and a LCVR of SEQ ID NO: 32, (d) a HCVR of SEQ ID NO: 37 and a LCVR of SEQ ID NO: 32, (e) a HCVR of SEQ ID NO: 38 and a LCVR of SEQ ID NO: 32, or (f) a HCVR of SEQ ID NO: 39 and a LCVR of SEQ ID NO: 32. In a specific embodiment thereof, the antibody is (a) Ab001_M1 and comprises a heavy chain of SEQ ID NO: 3 and a light chain of SEQ ID NO: 2, (b) Ab001_M2 and comprises a heavy chain of SEQ ID NO: 4 and a light chain of SEQ ID NO: 2, (c) Ab001_M3 and comprises a heavy chain of SEQ ID NO: 5 and a light chain of SEQ ID NO: 2, (d) Ab001_M5 and comprises a heavy chain of SEQ ID NO: 7 and a light chain of SEQ ID NO: 2, (e) Ab001_M6 and comprises a heavy chain of SEQ ID NO: 8 and a light chain of SEQ ID NO: 2, or (f) Ab001_M7 and comprises a heavy chain of SEQ ID NO: 9 and a light chain of SEQ ID NO: 2.;

[0041] In another embodiment, the present invention provides an isolated monoclonal antibody or antigen-binding fragment thereof, comprising: (a) a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 3; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 2; (b) a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 4; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 2; (c) a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 5; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 2; (d) a HC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 7; and a LC comprising a sequence at least 80% identical to and having the antigen-binding specificity of SEQ ID NO: 8. NO:2; (e) a HC comprising a sequence at least 80% identical to SEQ ID NO:8 and having the antigen binding specificity thereof; and a LC comprising a sequence at least 80% identical to SEQ ID NO:2 and having the antigen binding specificity thereof; or (f) a HC comprising a sequence at least 80% identical to SEQ ID NO:9 and having the antigen binding specificity thereof; and a LC comprising a sequence at least 80% identical to SEQ ID NO:2 and having the antigen binding specificity thereof, wherein the antibody or antigen binding fragment thereof specifically binds to CCR8.

[0042] The terms "antibody", "ab", "immunoglobulin" or "Ig" refer in the broadest sense to glycoproteins with the same structural features (i.e., molecules containing an antigen binding site that immunospecifically binds to an antigen), and encompass various antibody structures, including natural or artificial, monovalent or multivalent antibodies, including but not limited to monoclonal antibodies (including chimeric monoclonal antibodies, humanized monoclonal antibodies and human monoclonal antibodies, in particular humanized monoclonal antibodies), polyclonal antibodies, single-chain antibodies, antibody fragments and multispecific antibodies (e.g., bispecific antibodies). Although antibodies exhibit binding specificity to specific antigens, immunoglobulins include both antibodies and other antibody-like molecules that lack antigen specificity. "Antibody" encompasses any polypeptide comprising an antigen binding site, regardless of source, species of origin, method of production and characteristics. The term "antibody" can also broadly refer to molecules comprising CDR1, CDR2 and CDR3 of a heavy chain and CDR1, CDR2 and CDR3 of a light chain, wherein the molecule is capable of binding to an antigen. In some embodiments, the antibody comprises an HCVR and an LCVR. In some embodiments, the antibody comprises at least one heavy chain (HC) comprising an HCVR and at least a portion of a heavy chain constant region and at least one light chain (LC) comprising an LCVR and at least a portion of a light chain constant region. In some embodiments, the antibody comprises two heavy chains and two light chains, wherein each heavy chain comprises an HCVR and at least a portion of a heavy chain constant region, and wherein each light chain comprises an LCVR and at least a portion of a light chain constant region.

[0043] In some aspects, the isolated monoclonal antibody or antigen-binding fragment thereof comprises a HCVR sequence of SEQ ID NO:33 and / or a LCVR sequence of SEQ ID NO:32, a HCVR sequence of SEQ ID NO:34 and / or a LCVR sequence of SEQ ID NO:32, a HCVR sequence of SEQ ID NO:35 and / or a LCVR sequence of SEQ ID NO:32, a HCVR sequence of SEQ ID NO:37 and / or a LCVR sequence of SEQ ID NO:32, a HCVR sequence of SEQ ID NO:8 and / or a LCVR sequence of SEQ ID NO:32, or the HCVR sequence is SEQ ID NO:39 and / or the LCVR sequence is SEQ ID NO:32.

[0044] In other aspects, the isolated monoclonal antibody or antigen-binding fragment thereof comprises a HCVR sequence of SEQ ID NO:33 and a LCVR sequence of SEQ ID NO:32, a HCVR sequence of SEQ ID NO:34 and a LCVR sequence of SEQ ID NO:32, a HCVR sequence of SEQ ID NO:35 and a LCVR sequence of SEQ ID NO:32, a HCVR sequence of SEQ ID NO:37 and a LCVR sequence of SEQ ID NO:32, a HCVR sequence of SEQ ID NO:38 and a LCVR sequence of SEQ ID NO:32, or a HCVR sequence of SEQ ID NO:39 and a LCVR sequence of SEQ ID NO:32.

[0045] "Native antibodies" and "complete immunoglobulins" are typically heterotetrameric glycoproteins of about 150,000 daltons, composed of two identical light (L) chains and two identical heavy (H) chains. The light chains from any vertebrate species can be assigned to one of two clearly distinct types, called kappa (κ) and lambda (λ), based on the amino acid sequence of their constant domains. Immunoglobulins can be assigned to different classes based on the amino acid sequence of the constant domains of their heavy chains. There are five major immunoglobulin classes: IgA, IgD, IgE, IgG, and IgM, and several of these can be further divided into subclasses (isotypes). For example, IgG antibodies include, but are not limited to, IgG1 (comprising a γ1 constant region), IgG2 (comprising a γ2 constant region), IgG3 (comprising a γ3 constant region), and IgG4 (comprising a γ4 constant region) antibodies; IgA antibodies include, but are not limited to, IgA1 (comprising an α1 constant region) and IgA2 (comprising an α2 constant region) antibodies; and IgM antibodies include, but are not limited to, IgM1 (comprising a μ1 constant region) and IgM2 (comprising a μ2 constant region) antibodies. The heavy chain constant domains corresponding to different classes of immunoglobulins are referred to as α, δ, ε, γ, and μ, respectively. The subunit structures and three-dimensional configurations of different classes of immunoglobulins are well known.

[0046] Intact antibodies may have one or more "effector functions," which are those biological activities attributable to the Fc region (a native sequence Fc region or an amino acid sequence variant Fc region or any other modified Fc region) of an antibody. Examples of antibody effector functions include C1q binding; complement-dependent cytotoxicity; Fc receptor binding; ADCC; phagocytosis; downregulation of cell surface receptors (e.g., B cell receptor (BCR)); and cross-presentation of antigens by antigen-presenting cells or dendritic cells.

[0047] Each light chain is linked to a heavy chain by one covalent disulfide bond, while the number of disulfide linkages varies between the heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain has a variable domain (V H ), followed by a number of constant domains. Each light chain has a variable domain at one end (V L ) and have a constant domain at its other end; the constant domain of the light chain is aligned with the first constant domain of the heavy chain, and the light chain variable domain is aligned with the variable domain of the heavy chain. It is believed that specific amino acid residues form an interface between the light chain and the heavy chain variable domain. Each variable region includes three segments called complementary determining regions (CDRs) or hypervariable regions, and the more highly conserved parts of the variable domains are called framework regions (FRs). The variable domains of the heavy and light chains each include four FR regions, mainly adopting a β-sheet configuration, connected by three CDRs, which form a loop that connects the β-sheet structure and forms a part of the β-sheet structure in some cases. The CDRs in each chain are tightly held together by FR and, together with the CDRs from the other chain, contribute to the formation of the antigen binding site of the antibody (see Kabat et al., NIH Publication No. 91-3242, Volume 1, Pages 647-669

[1991] ). The constant domains do not directly participate in the binding of the antibody to the antigen, but exhibit various effector functions, such as the involvement of antibodies in antibody-dependent cellular toxicity.

[0048] As used herein, the term "heavy chain variable region (HCVR)" refers to at least the region comprising the heavy chain CDR1 (CDR-H1), framework 2 (HFR2), CDR2 (CDR-H2), FR3 (HFR3), and CDR3 (CDR-H3). In some embodiments, the HCVR further comprises at least a portion (e.g., all) of FR1 (HFR1) at the N-terminus of CDR-H1 and / or at least a portion (e.g., all) of FR4 (HFR4) at the C-terminus of CDR-H3. In various embodiments, any HCVR sequence disclosed herein further comprises an HFR1 sequence of SEQ ID NO: 23, an HFR2 sequence of SEQ ID NO: 24, an HFR3 sequence of SEQ ID NO: 25, and / or an HFR4 sequence of SEQ ID NO: 26.

[0049] As used herein, the term "heavy chain constant region" refers to a region comprising at least three heavy chain constant domains, CH1, CH2, and CH3. Non-limiting exemplary heavy chain constant regions include γ, δ, and α. Non-limiting exemplary heavy chain constant regions also include ε and μ. Each heavy chain constant region corresponds to an antibody isotype. For example, an antibody comprising a γ constant region is an IgG antibody, an antibody comprising a δ constant region is an IgD antibody, an antibody comprising an α constant region is an IgA antibody, an antibody comprising an ε constant region is an IgE antibody, and an antibody comprising a μ constant region is an IgM antibody.

[0050] As used herein, the term "light chain variable region (LCVR)" refers to the region comprising the light chain CDR1 (CDR-L1), framework (FR) 2 (LFR2), CDR2 (CDR-L2), FR3 (LFR3), and CDR3 (CDR-L3). In some embodiments, the LCVR further comprises at least a portion (e.g., all) of FR1 (LFR1) and / or at least a portion (e.g., all) of FR4 (LFR4). In various embodiments, any LCVR sequence disclosed herein further comprises the LFR1 sequence of SEQ ID NO: 27, the LFR2 sequence of SEQ ID NO: 28, the LFR3 sequence of SEQ ID NO: 29, and / or the LFR4 sequence of SEQ ID NO: 30.

[0051] As used herein, the term "light chain constant region" refers to the region comprising the light chain constant domain, CL. Non-limiting exemplary light chain constant regions include λ and κ.

[0052] As used herein, the term "light chain" refers to a polypeptide comprising at least an LCVR, with or without a leader sequence. In some embodiments, a light chain 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 an LCVR and a light chain constant region, with or without a leader sequence.

[0053] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a substantially homogeneous antibody population, that is, except for the possible naturally occurring mutations that may exist in a small amount, the individual antibodies constituting the population are identical. Monoclonal antibodies have a high degree of specificity and are directed against a single antigenic site. In addition, in contrast to the traditional (polyclonal) antibody preparations that typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. In addition to its specificity, the advantage of monoclonal antibodies is that they are synthesized by hybridoma culture and are not contaminated by other immunoglobulins. The modifier "monoclonal" indicates that the characteristic of the antibody is obtained from a substantially homogeneous antibody population, and should not be interpreted as requiring antibodies to be produced by any ad hoc method. For example, the monoclonal antibody to be used according to the present invention can be prepared by the hybridoma method described first by Kohler et al., Nature [nature], 256:495 (1975), or can be prepared by recombinant DNA methods (see, for example, U.S. Patent number 4,816,567). For example, "monoclonal antibodies" can also be isolated from phage antibody libraries using the techniques described in Clackson et al., Nature, 352:624-628 (1991) and Marks et al., J. Mol. Biol., 222:581-597 (1991).

[0054] In a general aspect, the present invention relates to isolated monoclonal antibodies or antigen-binding fragments thereof that bind to chemokine (CC motif) receptor 8 (CCR8).

[0055] As used herein, an antibody that "specifically binds to CCR8" refers to an antibody that binds to CCR8 at a concentration of 1×10 -7 M or less, preferably 1×10 -8 M or less, more preferably 5×10 -9 M or smaller, 1×10 -9 M or smaller, 5×10 -10 M or smaller or 1×10 -10 In some embodiments, the antibody and / or antigen binding domain binds to CCR8, preferably human CCR8, with a KD of M or less. In some embodiments, the antibody and / or antigen binding domain binds to cynomolgus monkey CCR8. The term "KD" refers to the dissociation constant, which is obtained from the ratio of Kd to Ka (i.e., Kd / Ka) and is expressed as a molar concentration (M). Based on the present disclosure, the KD value of an antibody can be determined using methods in the art. For example, the KD value can be determined by using surface plasmon resonance (e.g., by using, for example, The KD of the antibody can be determined by using a bio-layer interferometry system (e.g., a biosensor system such as the Octet RED96 system) or by using bio-layer interferometry technology (e.g., the Octet RED96 system).

[0056] The smaller the KD value of an antibody, the higher the affinity of the antibody for binding to the target antigen.

[0057] As used herein, the term "IC 50 " refers to the half-maximal inhibitory concentration of the monoclonal or bispecific antibody or antigen-binding fragment thereof of the present invention. IC 50 is a measure of the potency of the monoclonal or bispecific antibody or antigen-binding fragment thereof of the invention to inhibit the binding of CCL1 to CCR8 or inhibit the function of CCR8 in a cell. In certain embodiments, the KD of the monoclonal antibody or antigen-binding fragment thereof or the bispecific antibody or antigen-binding fragment thereof is less than about 10 -7 M, less than about 10 -8 M, less than about 10 -9 M, less than about 10 -10 M, less than about 10 -11 M, less than about 10 -12 M or less than about 10 -13 M.

[0058] As used herein, the term "EC 50 ” refers to the half-maximal effective concentration of the monoclonal or bispecific antibody or antigen-binding fragment thereof of the present invention. EC 50 EC refers to the concentration of a monoclonal or bispecific antibody or antigen-binding fragment thereof that induces a biological response (e.g., cell death) at halfway between baseline and maximum over a specified exposure time. In certain embodiments, the EC of a monoclonal antibody or antigen-binding fragment thereof or a bispecific antibody or antigen-binding fragment thereof is 50 Less than about 1 μM, about 1000 nM to about 100 nM, about 100 nM to about 10 nM, about 10 nM to about 1 nM, about 1000 pM to about 500 pM, about 500 pM to about 200 pM, less than about 200 pM, about 200 pM to about 150 pM, about 200 pM to about 100 pM, about 100 pM to about 10 pM, or about 10 pM to about 1 pM.

[0059] In one aspect, the HCVR sequence is SEQ ID NO:33 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:34 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:35 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:36 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:37 and / or the LCVR sequence is SEQ ID NO:32, the HCVR sequence is SEQ ID NO:38 and / or the LCVR sequence is SEQ ID NO:32, or the HCVR sequence is SEQ ID NO:39 and / or the LCVR sequence is SEQ ID NO:32.

[0060] In another aspect, the present invention provides an isolated monoclonal antibody comprising: (a) a heavy chain comprising SEQ ID NO: 3 and a light chain comprising SEQ ID NO: 2; (b) a heavy chain comprising SEQ ID NO: 4 and a light chain comprising SEQ ID NO: 2; (c) a heavy chain comprising SEQ ID NO: 5 and a light chain comprising SEQ ID NO: 2; (d) a heavy chain comprising SEQ ID NO: 7 and a light chain comprising SEQ ID NO: 2; (e) a heavy chain comprising SEQ ID NO: 8 and a light chain comprising SEQ ID NO: 2; (f) a heavy chain comprising SEQ ID NO: 9 and a light chain comprising SEQ ID NO: 2, wherein the antibody specifically binds to CCR8.

[0061] On the one hand, antibody is a humanized antibody. " humanization " form of non-human (for example, mouse) antibody is chimeric immunoglobulin, immunoglobulin chain or its fragment (such as Fv, Fab, Fab ', F (ab ') 2 or other antigen binding subsequences of antibody) containing the minimum sequence being derived from non-human immunoglobulin.In most cases, humanized antibody is such human immunoglobulin (receptor antibody), wherein the residue from the CDR of acceptor is replaced by the residue of the CDR with required specificity, affinity and ability from non-human species (donor antibody) (such as mouse, rat or rabbit).In some cases, the Fv FR residues of human immunoglobulin are replaced by corresponding non-human residues.In addition, humanized antibody can comprise residues that are neither in receptor antibody nor in the CDR or FR sequence of input, find.Carry out these modifications to further improve and maximize 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 CDRs correspond to those of a non-human immunoglobulin and all or substantially all of the FRs are those of a human immunoglobulin sequence. The humanized antibody will also optimally comprise at least a portion of an immunoglobulin constant region (Fc), typically at least a portion of a human immunoglobulin constant region. For further details, see Jones et al., Nature, 321:522-525 (1986); Reichmann et al., Nature, 332:323-329 (1988); and Presta, Curr. Op. Struct. Biol., 2:593-596 (1992). Humanized antibodies include PRIMATIZED TM The antibody, wherein the antigen-binding region of the antibody is derived from an antibody produced by immunizing a macaque with an antigen of interest.

[0062] Methods for humanizing non-human antibodies are well known in the art. Preferably, a humanized antibody has one or more amino acid residues introduced therein from a non-human source. These non-human amino acid residues are generally referred to as "input" residues, which are typically taken from the "input" variable domain. Humanization can be performed essentially according to the method of Winter and colleagues (Jones et al., Nature [Nature], 321: 522-525 (1986); Riechmann et al., Nature [Nature], 332: 323-327 (1988); Verhoeyen et al., Science [Science], 239: 1534-1536

[1988] ) by replacing rodent CDRs or CDR sequences with corresponding sequences of human antibodies. Therefore, such "humanized" antibodies are chimeric antibodies (U.S. Patent No. 4,816,567), in which far fewer than complete human variable domains have been replaced by corresponding sequences from non-human species. In practice, humanized antibodies are typically human antibodies in which some CDR residues and possibly some FR residues are substituted by residues from analogous sites in rodent antibodies.

[0063] The selection of human variable domains (both light and heavy chains) to be used for preparing humanized antibodies is very important for reducing antigenicity. According to the so-called "best fit" method, the sequence of the variable domains of rodent antibodies is screened for the entire library of known human variable domain sequences. The human sequence closest to the rodent sequence is then accepted as the human FR of the humanized antibody (Sims et al., J. Immunol. [Journal of Immunology], 151: 2296 (1993); Chothia et al., J. Mol. Biol. [Journal of Molecular Biology], 196: 901

[1987] ). Another method uses the specific FR of the consensus sequence of all human antibodies derived from a specific subgroup of light or heavy chains. The same FRs can be used for several different humanized antibodies (Carter et al., Proc. Natl. Acad. Sci. USA, 89:4285 (1992); Presta et al., J. Immunol., 151:2623

[1993] ).

[0064] It is further important to humanize antibodies when retaining high affinity to antigen and other favorable biological properties. In order to achieve this goal, according to a preferred method, humanized antibodies are prepared by analyzing the process of parental sequence and various conceptual humanized products using the three-dimensional model of the parental and humanized sequences. The three-dimensional immunoglobulin model is normally available and is familiar to those skilled in the art. The computer program that illustrates and displays the possible three-dimensional conformational structure of selected candidate immunoglobulin sequences is available. Check that these displays allow analysis of the possible effects of residues in the function of candidate immunoglobulin sequences, i.e., analyze the residues that affect the ability of candidate immunoglobulin to bind its antigen. In this way, FR residues can be selected and combined from receptor and input sequence to achieve desired antibody characteristics, such as the affinity to the increase of one or more target antigens. Usually, CDR residues directly and most substantially participate in affecting antigen binding.

[0065] Alternatively, it is now possible to produce transgenic animals (e.g., mice) that can produce a complete library of human antibodies in the absence of endogenous immunoglobulins after immunization. For example, homozygous deletions of antibody heavy chain joining region (JH) genes in chimeric and germline mutant mice have been described, resulting in complete suppression of endogenous antibody production. Transferring the human germline immunoglobulin gene array in such germline mutant mice will result in the production of human antibodies after antigen stimulation. See, for example, Jakobovits et al., Proc. Natl. Acad. Sci. USA [Proceedings of the National Academy of Sciences of the United States], 90: 2551 (1993); Jakobovits et al., Nature [Nature], 362: 255-258 (1993); Bruggermann et al., Year in Immuno. [Year in Immunology], 7: 33 (1993). Human antibodies can also be derived from phage display libraries (Hoogenboom et al., J. Mol. Biol., 227:381 (1991); Marks et al., J. Mol. Biol., 222:581-597

[1991] ).

[0066] Humanized antibodies can be made by replacing sequences of Fv variable regions that are not directly involved in antigen binding with equivalent sequences from human Fv variable regions. A general overview of humanized chimeric antibodies is provided by Morrison et al. (Science [Science] 229:1202-1207 (1985)) and Oi et al. (BioTechniques [Biotechnology] 4:214 (1986)). These methods include separating, manipulating, and expressing nucleic acid sequences encoding all or part of the immunoglobulin Fv variable region from at least one of the heavy or light chains. The source of such nucleic acids is well known to those skilled in the art and can, for example, be obtained from hybridomas that produce antibodies. The recombinant DNA encoding the humanized or chimeric antibody or its fragment can then be cloned into an appropriate expression vector.

[0067] Alternatively, humanized antibodies can be generated by CDR substitution (U.S. Patent No. 5,225,539; Jones, Nature 321:552-525 (1986); Verhoeyan et al., Science 239:1534 (1988); and Beidler, J. Immunol. 141:4053-4060 (1988)).

[0068] In another aspect, the antigen-binding fragment thereof is Fab, Fab', F(ab')2, F d , single-chain Fv or scFv, disulfide-linked F v , V-NAR domain, IgNar, intracellular antibody, IgGACH2, minibody, F(ab')3, tetrabody, triabody, diabody, single domain antibody, DVD-Ig, Fcab, mAb2, (scFv)2 or scFv-Fc.

[0069] Experimentally, antibodies can be cleaved with the proteolytic enzyme papain, which results in the breakage of each heavy chain, producing three separate antibody fragments. An "antibody fragment" comprises a portion of an intact antibody, preferably the antigen-binding region or variable region of an intact antibody. Examples of antibody fragments include Fab, Fab' and F(ab')2, Fc fragments or Fc fusion products, single-chain Fv (scFv), disulfide-linked Fv (sdFv), and fragments comprising a VL or VH domain; diabodies, triabodies, etc. (Zapata et al. Protein Eng. [Protein Engineering] 8(10): 1057-1062

[1995] ). The term "antibody fragment" or "antigen-binding portion" (of an antibody) includes, but is not limited to, a fragment that is capable of binding to an antigen. Two units consisting of a light chain and a heavy chain fragment that is approximately equal in mass to the light chain are referred to as Fab fragments (i.e., "antigen-binding" fragments). A third unit consisting of two equal segments of a heavy chain is referred to as an Fc fragment. The Fc fragment is typically not involved in antigen-antibody binding, but is important in later processes involved in clearance of antigen from the body.

[0070] The Fab fragment contains the constant domain of the light chain and the first constant domain (CH1) of the heavy chain. "Fab" refers to an antibody fragment with a molecular weight of about 50,000 daltons and has the activity of binding to an antigen. It comprises about half of the N-terminal side of the heavy chain connected by a disulfide bridge and a whole light chain. Fab can be obtained in particular by treating immunoglobulins with a protease (papain). The difference between the Fab' fragment and the Fab fragment is that several residues are added to the carboxyl terminus of the heavy chain CH1 domain, including one or more cysteines from the antibody hinge region. Fab'-SH is the name of such Fab' in this article, in which the one or more cysteine ​​residues of the constant domains carry a free sulfhydryl group.

[0071] F(ab')2 antibody fragments were originally produced as pairs of Fab' fragments with hinge cysteines between them. Other chemical couplings of antibody fragments are also known. The term "F(ab')2" refers to a fragment of approximately 100,000 daltons that has antigen-binding activity. This fragment is slightly larger than two Fab fragments linked by a disulfide bridge in the hinge region. These fragments are obtained by treating immunoglobulins with a protease (pepsin). Fab fragments can be obtained from F(ab')2 fragments by cleaving the disulfide bridge in the hinge region.

[0072] The Fc region of an antibody is the tail region of the antibody that interacts with cell surface receptors and some proteins of the complement system. This characteristic allows antibodies to activate the immune system. In IgG, IgA and IgD antibody isotypes, the Fc region is composed of two identical protein fragments derived from the second and third constant domains of the two heavy chains of the antibody; IgM and IgFc regions contain three heavy chain constant domains (CH domains 2-4) in each polypeptide chain. The Fc region of IgG carries highly conserved N-glycosylation sites. Glycosylation of the Fc fragment is essential for Fc receptor-mediated activity. The N-glycans attached to this site are mainly complex core fucosylated diantennary structures. In addition, a small amount of these N-glycans also carry bisecting GlcNAc and α-2,6 linked sialic acid residues.

[0073] Fc fusion proteins (also known as Fc chimeric fusion proteins, Fc-Ig, Ig-based chimeric fusion proteins and Fc-tagged proteins) are composed of the Fc domain of IgG genetically linked to a peptide or protein of interest. Fc fusion proteins have become valuable reagents for in vivo and in vitro studies. The binding partners of Fc fusions can range from single peptides, ligands activated upon binding to cell surface receptors, signaling molecules, extracellular domains of receptors activated upon dimerization, or as bait proteins for identifying binding partners in protein microarrays. One of the most valuable features of the Fc domain in vivo is that it can significantly extend the plasma half-life of the protein of interest, which, for biotherapeutics, leads to improved therapeutic efficacy; this attribute makes Fc fusion proteins attractive biotherapeutics.

[0074] The Fc fusion protein can be part of a pharmaceutical composition comprising the Fc fusion protein and a pharmaceutically acceptable excipient or carrier. Pharmaceutically acceptable carriers, excipients, or stabilizers are well known in the art (Remington's Pharmaceutical Sciences, 16th edition, Osol, A. ed. (1980)). Acceptable carriers, excipients, or stabilizers are nontoxic to the recipient at the dosages and concentrations employed and may include buffers such as phosphates, citrates, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl alcohol, or benzyl alcohol; alkyl parabens such as methyl paraben or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates, including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or nonionic surfactants such as TWEEN®. TM 、PLURONICS TM or polyethylene glycol (PEG).

[0075] "Fv" is the smallest antibody fragment containing a complete antigen recognition and antigen binding site. The dimer of "scFv" corresponds to two scFv molecules linked together by a peptide bond. The Fv chain is often the result of fusion gene expression, which includes genes encoding VH and VL connected by a linker sequence encoding a peptide. Human scFv fragments can include CDR regions preferably maintained in an appropriate conformation by means of genetic recombination technology. This region is composed of a dimer of a heavy chain variable domain and a light chain variable domain in a tight non-covalent association. It is in this configuration that the three CDRs of each variable domain interact to limit the antigen binding site on the surface of the VH-VL dimer. Together, six CDRs confer antigen binding specificity to antibodies. However, even a single variable domain (or half of an Fv comprising only three CDRs specific for an antigen) has the ability to recognize and bind to an antigen, but the affinity is lower than the entire binding site.

[0076] "Single-chain Fv" or "scFv" antibody fragments comprise the VH and VL domains of an antibody, wherein these domains are present in a single polypeptide chain. Preferably, the Fv polypeptide further comprises a polypeptide linker between the VH and VL domains, which enables the scFv to form the desired structure for antigen binding. For a review of scFv, see Pluckthun, The Pharmacology of Monoclonal Antibodies, Vol. 113, Rosenburg and Moore, eds., Springer-Verlag, New York, pp. 269-315 (1994).

[0077] A "dsFv" fragment is a VH-VL heterodimer stabilized by disulfide bridges; it can be bivalent (dsFV2). Divalent Sc(Fv)2 or multivalent antibody fragments can be formed spontaneously by the association of monovalent scFvs or by linking scFv fragments via peptide binding sequences.

[0078] The Fc fragment supports the biological properties of antibodies, especially their ability to be recognized by immune effectors or to activate complement. It consists of the constant segments outside the hinge region of the heavy chain.

[0079] The term "diabodies" refers to small antibody fragments with two antigen-binding sites. These fragments contain a variable heavy chain domain (VH) connected to a variable light chain domain (VL) within the same VH-VL polypeptide chain. Using binding sequences that are too short to allow for matching between the two domains of the same chain inevitably results in matching with two complementary domains of another chain, thus creating two antigen-binding sites.

[0080] Various techniques for producing antibody fragments have been developed. Traditionally, these fragments are obtained via proteolytic digestion of intact antibodies (see, for example, Morimoto et al., Journal of Biochemical and Biophysical Methods [Journal of Biochemical and Biophysical Methods] 24: 107-117 (1992) and Brennan et al., Science [Science], 229: 81

[1985] ). However, these fragments can now be directly produced by recombinant host cells. For example, antibody fragments can be isolated from the antibody phage library discussed above. Alternatively, Fab'-SH fragments can be directly recovered from Escherichia coli and chemically coupled to form F(ab')2 fragments (Carter et al., Bio / Technology [Biology / Technology] 10: 163-167

[1992] ). According to another approach, F(ab')2 fragments can be directly isolated from recombinant host cell cultures. Other techniques for producing antibody fragments will be clear to skilled practitioners. In other embodiments, the antibody of choice is a single-chain Fv fragment (scFv). See WO 93 / 16185.

[0081] In other aspects, the monoclonal antibody or antigen-binding fragment thereof has a K of less than about 25 nM, 20 nM, 15 nM, 10 nM, 5 nM, 2 nM, 1 nM, 0.5 nM, or 0.01 nM. d Binds to human CCR8.

[0082] The term "antigen binding domain" refers to the portion of an antibody molecule that contains a region that specifically binds to or is complementary to a portion or all of an antigen. In the case of a larger antigen, an antibody may only bind to a specific portion of the antigen. An "epitope" or "antigenic determinant" is the portion of an antigen molecule that is responsible for interacting with the antigen binding domain of an antibody. The antigen binding domain can be provided by one or more antibody variable domains (e.g., a so-called Fd antibody fragment consisting of a VH domain). The antigen binding domain may comprise an antibody LCVR and an antibody HCVR.

[0083] " Antigen " according to the present invention covers any substance that will cause an immune response. In particular, " antigen " relates to any substance with antibody or T lymphocyte (T cell) specific reaction, preferably peptide or protein. According to the present invention, the term " antigen " includes any molecule comprising at least one epitope. Preferably, the antigen in the context of the present invention is a molecule that optionally induces an immune response after treatment. According to the present invention, any suitable antigen can be used, which is a candidate for immune response, wherein the immune response is preferably a cellular immune response. In the context of an embodiment of the present invention, antigen is preferably presented by cell, preferably by antigen presenting cells, and the antigen presenting cells include diseased cells, particularly cancer cells (in the case of MHC molecules), which result in an immune response against the antigen. Antigen is preferably a product corresponding to a naturally occurring antigen, or is derived from a naturally occurring antigen. Such naturally occurring antigens include tumor antigens.

[0084] The term "epitope" refers to an antigenic determinant in a molecule (such as an antigen), that is, a part or fragment of a molecule that is recognized by the immune system. An epitope of a protein (such as a tumor antigen) preferably comprises a continuous or discontinuous portion of the protein.

[0085] The terms "epitope", "antigenic peptide", "antigenic epitope", "immunogenic peptide" and "MHC binding peptide" may be used interchangeably herein and preferably relate to an incomplete representation of an antigen that is preferably capable of eliciting an immune response against the antigen or a cell that expresses or contains and preferably presents the antigen.

[0086] The term "binding affinity" generally 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. A variety of methods for measuring binding affinity or binding activity are known in the art, any of which can be used for the purposes of the present invention. Specific illustrative embodiments are described below.

[0087] As used herein, "specific binding" refers to the binding of an antibody to a predetermined antigen. Typically, an antibody binds to a specific antigen in an amount corresponding to about 10 -8 M or smaller K D The antibody binds with such affinity (as determined by K D The antibody may bind to the predetermined antigen with an affinity that is at least 10 times smaller, and preferably at least 100 times smaller, than the affinity of the antibody for binding to a nonspecific antigen other than the predetermined antigen or a closely related antigen (e.g., BSA, casein). Alternatively, the antibody may bind to the predetermined antigen with an affinity that corresponds to about 10 6 M -1 , or about 10 7 M -1 , or about 10 8 M -1 , or 109 M -1 or higher K A The antibody binds with such affinity (as determined by K A The antibody binds to the predetermined antigen with an affinity that is at least 10 times, and preferably at least 100 times, greater than the affinity with which the antibody binds to a nonspecific antigen other than the predetermined antigen or a closely related antigen (e.g., BSA, casein).

[0088] As used herein, the term "k d ”(sec -1 ) is intended to refer to the dissociation rate constant for a particular antibody-antigen interaction. This value is also referred to as the off value. As used herein, the term "K D ”(M -1 ) is intended to refer to the equilibrium dissociation constant for a specific antibody-antigen interaction.

[0089] In some embodiments, the dissociation constant (K) of a CCR8-binding antibody of the invention for CCR8 (e.g., for human CCR8) is d )≤1 μM, ≤100 nM, ≤10 nM, ≤1 nM, ≤0.1 nM, ≤0.01 nM or ≤0.001 nM (e.g., 10 -8 M or smaller, for example, from 10 -8 M to 10 -13 M, for example, from 10 -9 M to 10 -13 In certain embodiments, the dissociation constant (K) of a CCR8-binding antibody for CCR8 (e.g., for human CCR8) is d )≤1 μM, ≤100 nM, ≤10 nM, ≤1 nM, ≤0.1 nM, ≤0.01 nM or ≤0.001 nM (e.g., 10 -8 M or smaller, for example, from 10 -8 M to 10 -13 M, for example, from 10 -9 M to 10 -13 M).

[0090] As used herein, the term "k a ”(M -1 sec -1 ) is intended to refer to the association rate constant for a particular antibody-antigen interaction. As used herein, the term "K A "(M) is intended to refer to the equilibrium association constant for a particular antibody-antigen interaction.

[0091] As described herein, anti-CCR8 antibodies may also be referred to as WT anti-CCR8 antibodies (having a HCVR comprising the sequence of SEQ ID NO: 31 and / or a LCVR comprising the sequence of SEQ ID NO: 32), as well as mutants thereof. As used herein, the term "mutant" antibody refers to an antibody having a mutation in CDR3 to eliminate post-translational modification (deamidation and isomerization) of the asparaginyl residues at N104 and / or N105 in the wild-type sequence. Mutations in the 3-amino acid sequence 103-105 (DNN) reduce or prevent deamidation and isomerization, thereby improving the stability and efficacy of the antibody. In particular, mutations in the DNN sequence to EQQ prevent deamidation and retain equivalent binding to CCR8 as the parent antibody. This modification extends the active half-life of the mutant antibody, greatly exceeding the active half-life of the parent antibody.

[0092] Antibodies having a HCVR sequence of any one of SEQ ID NOs: 33-39 and a LCVR sequence of SEQ ID NO: 32 are referred to as mutant anti-CCR8 antibodies. In particular, antibodies having a HCVR sequence of SEQ ID NO: 36 and a LCVR sequence of SEQ ID NO: 32 are preferred mutant anti-CCR8 antibodies.

[0093] On the one hand, antibodies induce antibody-dependent cell-mediated cytotoxicity (ADCC). ADCC is a cell-mediated immune defense mechanism in which effector cells of the immune system actively lyse target cells whose membrane surface antigens have been bound by specific antibodies. This is one of the mechanisms by which antibodies, as part of the humoral immune response, can act to limit and curb infection. ADCC is independent of the immune complement system, which also lyses targets but does not require any other cells. ADCC requires effector cells, which are generally known to be natural killer (NK) cells that typically interact with immunoglobulin G (IgG) antibodies. However, macrophages, neutrophils, and eosinophils can also mediate ADCC, such as eosinophils that kill certain parasites called helminths via IgE antibodies. Generally, ADCC is typically described as an immune response to antibody-coated cells, ultimately leading to the lysis of infected cells or non-host cells. In recent literature, its importance in treating cancer cells and a deeper understanding of its seemingly complex pathways have been topics of increasing interest to medical researchers.

[0094] As used herein, the term "variant" antibody refers to an antibody with variable levels of fucosylation. Defucosylated monoclonal antibodies are monoclonal antibodies produced in engineered cells or under selected conditions such that the oligosaccharides in the Fc region of the defucosylated monoclonal antibody have reduced or no fucose units compared to production in non-engineered cells or under normal conditions. When an antibody is defucosylated, ADCC is increased. Most approved monoclonal antibodies are of the IgG1 isotype, in which two N-linked biantennary complex oligosaccharides are bound to the Fc region. The Fc region exerts the effector function of ADCC through its interaction with leukocyte receptors of the FcγR family. ADCC is important in the efficacy of cancer antibodies, but for many approved cancer antibodies, ADCC is less than might be expected due to nonspecific IgG competing with the drug for binding to FcγIIIa on natural killer cells. Defucosylated monoclonal antibodies overcome this problem through improved FcγIIIa binding.

[0095] In their review, Pereira et al. (MABS [Monoclonal Antibodies] 2018, Vol. 10, No. 5, 693-711) discussed the correlation between antibody core fucosylation and ADCC, different strategies for generating defucosylated antibodies, and updates on defucosylated antibody drugs currently in clinical trials and those already approved. The disclosures in that article are hereby incorporated into this application in their entirety.

[0096] In another aspect, the antibody has a defucosylation level of at least 50%. For example, the antibody has a defucosylation level of at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more.

[0097] In various aspects, the antibody has a defucosylation level of at least 85%. For example, the antibody has a defucosylation level of at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more.

[0098] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of any of the isolated monoclonal antibodies or antigen-binding fragments described herein, thereby treating the cancer in the subject.

[0099] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, the method comprising administering to the subject an effective amount of at least one isolated monoclonal antibody or antigen-binding fragment thereof as provided herein, thereby treating the cancer in the subject. In one aspect, the method comprises administering an antibody having a HCVR comprising a CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVRCDR3 sequence having the amino acid sequence of TRGSEQQYYYAMDY (SEQ ID NO: 19); and a LCVR comprising a LCVRCDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15), thereby treating the cancer in the subject. In a specific embodiment thereof, the antibody comprises the HCVR of SEQ ID NO: 36 and the LCVR of SEQ ID NO: 32. In a specific embodiment thereof, the antibody is Ab001_M4 and comprises the heavy chain of SEQ ID NO: 6 and the light chain of SEQ ID NO: 2.

[0100] As used herein, the term "subject" refers to any individual or patient to whom the subject method is performed. Typically, the subject is a human, but as will be appreciated by those skilled in the art, the subject can be an animal. Thus, other animals (including vertebrates such as rodents (including mice, rats, hamsters, and guinea pigs), cats, dogs, rabbits, farm animals (including cattle, horses, goats, sheep, pigs, chickens, etc.), and primates (including monkeys, chimpanzees, orangutans, and gorillas) are included within the definition of a subject.

[0101] The term "treatment" is used interchangeably with the term "treatment method" herein and refers to 1) a therapeutic treatment or measure that cures, slows down, alleviates the symptoms of a diagnosed pathological condition or disorder and / or stops the progression of a diagnosed pathological condition or disorder, and 2) a preventive / preventative measure. Those individuals in need of treatment may include individuals who already have a specific medical disorder and those individuals who may eventually develop a disorder (i.e., those individuals in need of preventive measures). "Treatment" covers any administration or application of therapeutic agents for diseases (also referred to herein as "disorders" or "disorders") in mammals (including humans), and includes inhibiting disease or disease progression, inhibiting or slowing disease or its progression, preventing its development, partially or completely alleviating disease, partially or completely alleviating one or more symptoms of a disease, or restoring or repairing lost, missing or defective functions; or stimulating an inefficient process. The term "treatment" also includes reducing the severity of any phenotypic characteristic and / or reducing the incidence, degree or likelihood of the characteristic.

[0102] The terms "therapeutically effective amount," "effective dose," "therapeutically effective dose," "effective amount," and the like refer to the amount of the subject compound that will elicit a biological or medical response in a tissue, system, animal, or human that is being sought by a researcher, veterinarian, physician, or other clinician. Typically, the response is an improvement in a patient's symptoms or a desired biological outcome (e.g., treatment of cancer). The therapeutically effective amount of a CCR8 antagonist of the present invention can vary depending on factors such as the disease state, age, sex, and weight of the individual, as well as the ability of the antagonist to elicit a desired response in the individual. A therapeutically effective amount encompasses an amount in which the therapeutically beneficial effects outweigh any toxic or deleterious effects of the CCR8 antagonist.

[0103] The terms "administration of" and / or "administering" should be understood to mean providing a therapeutically effective amount of a pharmaceutical composition to a subject in need of treatment. The route of administration can be enteral, topical, or parenteral. Thus, routes of administration include, but are not limited to, intradermal, subcutaneous, intravenous, intraperitoneal, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, transdermal, transtracheal, subcutaneous, intraarticular, subcapsular, subarachnoid, intraspinal and intrasternal, oral, sublingual, buccal, rectal, vaginal, nasal, ocular administration, as well as infusion, inhalation, and aerosolization. As used herein, the phrases "parenteral administration" and "administered parenterally" mean modes of administration other than enteral and topical administration. In various embodiments, the anti-CCR8 antibody can be administered subcutaneously or intravenously.

[0104] The antibodies and antigen-binding fragments thereof described herein can be prepared as pharmaceutical compositions comprising the antibodies or their antigen-binding fragments and a pharmaceutically acceptable carrier. 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 a therapeutic agent, which together with the therapeutic agent constitute a "pharmaceutical composition" for administration to a subject. A pharmaceutically acceptable carrier is non-toxic to the recipient at the dosage and concentration employed and is compatible with the other ingredients of the dosage form. A pharmaceutically acceptable carrier is suitable for the formulation employed. For example, if the therapeutic agent is to be administered orally, the carrier can be a gel capsule. If the therapeutic agent is to be administered subcutaneously, the carrier is ideally non-allergenic to the skin and does not cause injection site reactions.

[0105] The pharmaceutical composition can be administered in a variety of unit dosage forms according to the method of administration. Suitable unit dosage forms include, but are not limited to, powders, tablets, pills, capsules, lozenges, suppositories, patches, nasal sprays, injections, implantable sustained-release formulations, lipid complexes, and the like.

[0106] The subject compositions may be formulated into preparations in solid, semisolid, liquid or gaseous form; including but not limited to tablets, capsules, powders, granules, ointments, solutions, suppositories, enemas, injections, inhalants and aerosols.

[0107] In various embodiments, compositions comprising a CCR8 antagonist are provided in formulations with a variety of pharmaceutically acceptable carriers (see, e.g., Gennaro, Drugfacts Plus, 20th ed. (2003); Ansel et al., Pharmaceutical Dosage Forms and Drug Delivery Systems, 7th ed., Lippencott Williams and Wilkins (2004); Kibbe et al., Handbook of Pharmaceutical Excipients, 3rd ed., 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 adjusters and buffers, tonicity adjusters, stabilizers, wetting agents, and the like, are also available. Non-limiting exemplary carriers include saline, buffered saline, dextrose, water, glycerol, ethanol, and combinations thereof.

[0108] In various embodiments, compositions comprising CCR8 antagonists can be formulated for injection (including subcutaneous administration) by dissolving, suspending or emulsifying them in an aqueous or non-aqueous solvent (such as vegetable oil or other oil, synthetic fatty acid glyceride, higher fatty acid ester or propylene glycol); and if necessary, together with traditional additives (such as solubilizers, isotonic agents, suspending agents, emulsifiers, stabilizers and preservatives).

[0109] In various embodiments, the compositions can be formulated for inhalation, for example, using pressurized acceptable propellants (eg, dichlorodifluoromethane, propane, nitrogen, etc.).

[0110] In various embodiments, compositions can also be configured to slow-release microcapsules, as with biodegradable or non-biodegradable polymers. Non-limiting exemplary biodegradable formulations include polylactic-glycolic acid (PLGA) polymers. Non-limiting exemplary non-biodegradable formulations include polyglycerol fatty acid esters. Some methods of preparing such formulations are described in, for example, EP 1125584 A1.

[0111] Also provided is a pharmaceutical dosage package comprising one or more containers, each container containing one or more doses of an anti-CCR8 antibody and its antigen-binding fragment. In some embodiments, a unit dose is provided, wherein the unit dose contains a predetermined amount of a composition comprising an anti-CCR8 antibody and its antigen-binding fragment, with or without one or more additional agents. In some embodiments, such a unit dose is provided in a disposable prefilled syringe for injection. In various embodiments, the composition contained in the unit dose may include saline, sucrose, etc.; a buffer such as phosphate, etc.; and / or may be formulated within a stable and effective pH range. Alternatively, in some embodiments, the composition may be provided as a lyophilized powder that can be reconstituted after adding an appropriate liquid (e.g., sterile water). In some embodiments, the composition comprises one or more substances that inhibit protein aggregation, including but not limited to sucrose and arginine. In some embodiments, the composition of the present invention comprises heparin and / or proteoglycans.

[0112] The pharmaceutical composition is administered in an amount effective to treat or prevent the particular indication. A therapeutically effective amount typically depends on the weight of the subject being treated, his or her physical or health condition, the extensiveness of the condition to be treated, or the age of the subject being treated.

[0113] In some embodiments, the anti-CCR8 antibodies and antigen-binding fragments thereof may be administered in an amount ranging from about 50 μg / kg body weight to about 50 mg / kg body weight per dose. In some embodiments, the anti-CCR8 antibodies and antigen-binding fragments thereof may be administered in an amount ranging from about 100 μg / kg body weight to about 50 mg / kg body weight per dose. In some embodiments, the anti-CCR8 antibodies and antigen-binding fragments thereof may be administered in an amount ranging from about 100 μg / kg body weight to about 20 mg / kg body weight per dose. In some embodiments, the anti-CCR8 antibodies and antigen-binding fragments thereof may be administered in an amount ranging from about 0.5 mg / kg body weight to about 20 mg / kg body weight per dose.

[0114] In some embodiments, the anti-CCR8 antibodies and antigen-binding fragments thereof may be administered in an amount ranging from about 10 mg to about 1,000 mg per dose. In some embodiments, the anti-CCR8 antibodies and antigen-binding fragments thereof may be administered in an amount ranging from about 20 mg to about 500 mg per dose. In some embodiments, the anti-CCR8 antibodies and antigen-binding fragments thereof may be administered in an amount ranging from about 20 mg to about 300 mg per dose. In some embodiments, the anti-CCR8 antibodies and antigen-binding fragments thereof may be administered in an amount ranging from about 20 mg to about 200 mg per dose.

[0115] Anti-CCR8 antibodies and antigen-binding fragments thereof can be administered to the subject as needed. In some embodiments, an effective dose of anti-CCR8 antibodies and antigen-binding fragments thereof are administered to the subject once or multiple times. In various embodiments, an effective dose of anti-CCR8 antibodies and antigen-binding fragments thereof are administered to the subject once a month, less than once a month (such as, for example, every two months, every three months, or every six months). In other embodiments, an effective dose of anti-CCR8 antibodies and antigen-binding fragments thereof are administered more than once a month (such as, for example, every two weeks, every week, twice a week, three times a week, every day, or multiple times a day). An effective dose of anti-CCR8 antibodies and antigen-binding fragments thereof are administered to the subject at least once. In some embodiments, an effective dose of CCR8 antagonists can be administered multiple times, including a period of at least one month, at least six months, or at least one year. In some embodiments, anti-CCR8 antibodies and antigen-binding fragments thereof are administered to the subject as needed to alleviate one or more symptoms of the disease.

[0116] The antibodies described herein can be used to treat cancer.

[0117] Cancer is a group of diseases involving abnormal cell growth that has the potential to invade or spread to other parts of the body. In 2015, approximately 90.5 million people were living with cancer, with approximately 14.1 million new cases occurring annually and causing approximately 8.8 million deaths (15.7% of all deaths). The most common types of cancer in men are lung, prostate, colorectal, and stomach cancers. In women, the most common types are breast, colorectal, lung, and cervical cancers.

[0118] The term "cancer" refers to a group of diseases characterized by abnormal, uncontrolled cell proliferation that begins in one site (the primary site) and has the potential to invade and spread to other sites (secondary sites, metastases), which distinguishes cancer (malignant tumors) from benign tumors. Virtually all organs can be affected, resulting in more than 100 types of cancer that can affect humans. Cancer can result from many causes, including genetic predisposition, viral infection, exposure to ionizing radiation, exposure to environmental pollutants, tobacco and / or alcohol use, obesity, poor diet, lack of physical activity, or any combination thereof.

[0119] As used herein, "neoplasm" or "tumor" (including grammatical variations thereof) means a new and abnormal growth of tissue, which may be benign or cancerous. In related aspects, a neoplasm is indicative of a neoplastic disease or disorder, including, but not limited to, various cancers. For example, such cancers may include prostate cancer, pancreatic cancer, bile duct cancer, colon cancer, rectal cancer, liver cancer, kidney cancer, lung cancer, testicular cancer, breast cancer, ovarian cancer, pancreatic cancer, brain cancer and head and neck cancer, melanoma, sarcoma, multiple myeloma, leukemia, lymphoma, and the like.

[0120] In one aspect, the cancer is a hematological cancer or a solid tumor.

[0121] Cancers that begin in cells of blood-forming tissues (such as the bone marrow) or the immune system are called blood cancers or leukemias. Blood cancers affect the production and function of blood cells and are classified into three main types: leukemias, lymphomas, and multiple myeloma.

[0122] As used herein, "leukemia" refers to a blood cancer caused by the rapid production of abnormal white blood cells. Examples of leukemias include acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia, chronic myeloid leukemia, and hairy cell leukemia.

[0123] As used herein, "lymphoma" refers to a type of blood cancer that affects the lymphatic system. Examples of lymphomas include AIDS-related lymphoma, cutaneous T-cell lymphoma, Hodgkin lymphoma, mycosis fungoides, non-Hodgkin lymphoma, primary central nervous system lymphoma, Sézary syndrome, cutaneous T-cell lymphoma, and Waldenstrom's macroglobulinemia.

[0124] As used herein, "myeloma" is a cancer of the plasma cells. Examples of myelomas include chronic myeloproliferative neoplasm, Langerhans cell histiocytosis, multiple myeloma, plasma cell neoplasm, myelodysplastic syndrome, and myelodysplastic / myeloproliferative neoplasm.

[0125] In some embodiments, the anti-CCR8 antibodies of the present invention may be used alone, or alternatively in combination with any other suitable compound known to be capable of treating a disease or indication.

[0126] In some aspects, administration can be combined with one or more additional therapeutic agents. The phrases "combination therapy," "in combination with," and the like refer to the simultaneous use of more than one drug or treatment to increase a response. The compositions of the present invention can, for example, be used in combination with other drugs or treatments for the treatment of cancer. In particular, administration of the compositions of the present invention to a subject can be combined with any anti-cancer therapy. Such therapies can be administered before, simultaneously with, or after administration of the compositions of the present invention.

[0127] In certain embodiments, an anti-CCR8 antibody is administered to a subject (e.g., a subject with cancer) simultaneously or continuously with another treatment. For example, an anti-CCR8 antibody can be administered together with one or more of the following: radiotherapy, surgery, or chemotherapy, such as targeted chemotherapy or immunotherapy. The administration of the two agents can be started, for example, 30 minutes, 60 minutes, 90 minutes, 120 minutes, 3 hours, 6 hours, 12 hours, 24 hours, 36 hours, 48 ​​hours, 3 days, 5 days, 7 days, or a week or weeks apart, or the administration of the second agent can be started, for example, 30 minutes, 60 minutes, 90 minutes, 120 minutes, 3 hours, 6 hours, 12 hours, 24 hours, 36 hours, 48 ​​hours, 3 days, 5 days, 7 days, or a week or weeks after the first agent has been administered.

[0128] In some aspects, the hematological cancer is selected from the group consisting of acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia, chronic myelogenous leukemia, hairy cell leukemia, AIDS-related lymphoma, cutaneous T-cell lymphoma, Hodgkin lymphoma, mycosis fungoides, non-Hodgkin lymphoma, primary central nervous system lymphoma, Sezary syndrome, cutaneous T-cell lymphoma, Waldenstrom's macroglobulinemia, diffuse large B-cell lymphoma (DLBCL), and multiple myeloma.

[0129] Examples of solid cancers include, but are not limited to, carcinoma, sarcoma, squamous cell carcinoma, small cell lung cancer, pituitary cancer, esophageal cancer, astrocytoma, soft tissue sarcoma, non-small cell lung cancer, lung adenocarcinoma, lung squamous cell carcinoma, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, renal cancer, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, hepatic carcinoma, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, stomach cancer, melanoma, and various types of head and neck cancer.

[0130] In other aspects, the solid tumor is selected from the group consisting of breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colon cancer, kidney cancer, ovarian cancer, liver cancer, and prostate cancer.

[0131] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment thereof having a HCVR comprising a CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSEQQYYYAMDY (SEQ ID NO: 19); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15), thereby treating cancer in the subject.

[0132] In some aspects, the methods described herein further comprise administering to the patient a chemotherapeutic agent, an anti-angiogenic agent, a growth inhibitory agent, an immuno-oncology agent, and / or an anti-tumor composition.

[0133] A "chemotherapeutic agent" is a chemical compound that can be used to treat cancer. Examples of chemotherapeutic agents include, but are not limited to, alkylating agents such as thiotepa and cyclosphosphamide; alkyl sulfonates such as busulfan, improsulfan, and piposulfan; aziridines such as benzodopa, carboquinone, meturedopa, and uredopa; ethyleneimines and methylamelamines, including hexamethylmelamine, triethylenemelamine, trietylenephosphoramide, triethiylenethiophosphoramide, and trimethylolomelamine; polyacetylamines (particularly bullatacin and bullatacinone); camptothecins (including synthetic analogs of topotecan); bryostatin; callystatin; CC-1065 (including its synthetic analogs of adolesin, carzelesin, and biszelesin); candidiasis (particularly candidiasis 1 and candidiasis 8); caudate; duocarmycin (including synthetic analogs KW-2189 and CB1-TM1); eleutherobin; pancratistatin; sarcodictyin; spongistatin; nitrogen mustards, such as chlorambucil, naphthyl mustard, cholophosphamide, estramustine, ifosfamide, mechlorethamine, chlorambucil oxide hydrochloride, melphalan, neomuscarin, phenylephrine, prednimustine, trofosfamide, uracil mustard mustard); nitrosureas, such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, and ranimnustine; antibiotics, such as enediyne antibiotics (e.g., calicheamicins, particularly calicheamicin γll and calicheamicin ωll (see, e.g., Agnew, Chem lntl. Ed. Engl [German Angewandte Chemie International English Edition], 33:183-186 (1994); dynemicins, including dynemicin A; bisphosphonates, such as clodronate; esperamicin;and neocarzinostatin chromophores and related chromoprotein enediyne antibiotic chromophores), aclacinomycin, actinomycin, authramycin, azaserine, bleomycin, actinomycin C, carabicin, carminomycin, carmosin, chromomycin, dactinomycin, daunomycin, detoxorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin (including morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin, and deoxydoxorubicin), epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins (such as mitomycin C), mycophenolic acid, nogamycin, olivomycin, peplomycin, porphyromycin, and potycin. firomycin), puromycin, quelamycin, rhodorubicin, streptozotocin, streptozotocin, tuberculin, ubenimex, zorubicin, antimetabolites, such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs, such as leucovorin, methotrexate, pteropterin, trimetrexate; purine analogs, such as fludarabine, 6-mercaptopropionate, Purines, thioamidine, thioguanine; pyrimidine analogs, such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, and floxuridine; androgens, such as caprotestosterone, drostanolone propionate, cyclothiocarbamate, melastane, and testolactone; antiadreners, such as aminoglutethimide, mitotane, and trolostane; folic acid supplements, such as folinic acid (folinic acid) acid; acetolactone; aldophosphamide glycoside; aminolevulinic acid; eniluracil; amsacrine; bestrabucil; bisantrene; edatraxate; defofamine; demeclocycline; diacrazine; elfomithine; elliptonium acetate; epothilone; etoglucose; gallium nitrate; hydroxyurea; lentinan; lonidainine; maytansine alkaloids, such as maytansine and ansamitocin; mitoxantrone; mitoxantrone; mopidanmol; nitraerine; pentostatin; methamidine; pirarubicin; losoxantrone; podophyllic acid; 2-ethylhydrazide; procarbazine; polysaccharide complex (JHS Natural Products Company, Inc.) Natural Products, Eugene, Oregon); razoxane; lisoxantrone; sizoran; spirogermanamine; tenuisporic acid; triazoquinone; 2,2′,2″-trichlorotriethylamine; trichothecenes (especially T-2 toxin, verracurin A, myclosan A, and anguidine); urethane; vindesine; dacarbazine; mannomustine; dibromomannitol; dibromodulcitol; pipobroman;gacytosine; arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxanes, such as paclitaxel (Bristol-Myers Squibb Oncology, Princeton, NJ), albumin-engineered nanoparticle formulations of paclitaxel without cremophor (American Pharmaceutical Partners, Schaumburg, IL), and doxetaxel (Rhone-Poulenc Rorer, Antoni, France); chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogs such as cisplatin, oxaliplatin, and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; vinorelbine; novantrone; teniposide; edatrexate; daunorubicin; aminopterin; xeloda; ibandronate; irinotecan (CPT-11) (including irinotecan with 5-FU and formyl tetracycline) leucovorin (LV); oxaliplatin, including oxaliplatin treatment regimens (FOLFOX); inhibitors of PKC-α, Raf, H-Ras, EGFR (e.g., erlotinib), and VEGF-A that reduce cell proliferation, and pharmaceutically acceptable salts, acids, or derivatives of any of the above.

[0134] Other non-limiting exemplary chemotherapeutic agents include antihormonal agents that act to modulate or inhibit the effects of hormones on cancer, such as antiestrogens and selective estrogen receptor modulators (SERMs), including, for example, tamoxifen, raloxifene, droloxifene, 4-hydroxytamoxifen, troloxifene, keoxifene, LY117018, onapristone, and toremifene; aromatase inhibitors that inhibit the aromatase enzyme, which regulates estrogen production in the adrenal glands, such as, for example, 4(5)-imidazole, aminoglutethimide, megestrol acetate, exemestane, formestanie, fadrozole, voriconazole, and valproate; Chlorazole, letrozole and anastrozole; and antiandrogens such as flutamide, nilutamide, bicalutamide, leuprolide and goserelin; and troxacitabine (1,3-dioxolane nucleoside cytosine analog); antisense oligonucleotides, particularly those that inhibit the expression of genes in signal transduction pathways involved in abnormal cell proliferation (such as, for example, PKC-α, Ralf and H-Ras); ribozymes such as VEGF expression inhibitors and HER2 expression inhibitors; vaccines such as gene therapy vaccines; rIL-2; topoisomerase 1 inhibitors; rmRH; and pharmaceutically acceptable salts, acids or derivatives of any of the above.

[0135] "Anti-angiogenic agent" or "angiogenesis inhibitor" refers to a small molecular weight substance, polynucleotide (including, for example, inhibitory RNA (RNAi or siRNA), polypeptide, isolated protein, recombinant protein, antibody, or conjugate or fusion protein thereof, that directly or indirectly inhibits angiogenesis, vasculogenesis, or undesirable vascular permeability. It is understood that anti-angiogenic agents include those agents that bind to angiogenic factors or their receptors and block the angiogenic activity of the angiogenic factors or their receptors. For example, anti-angiogenic agents are antibodies or other antagonists to angiogenic agents, such as antibodies to VEGF-A (e.g., bevacizumab) or VEGF-A receptors (e.g., KDR receptor or Flt-1 receptor), anti-PDGFR inhibitors such as (imatinib mesylate), small molecules that block VEGF receptor signaling (e.g., PTK787 / ZK2284, SU6668, SUI1248 (sunitinib malate), AMG706, or the agonists described, for example, in International Patent Application WO 2004 / 113304). Antiangiogenic agents also include natural angiogenesis inhibitors, such as angiostatin, endostatin, and the like. See, for example, Klagsbrun and D'Amore (Annu. Rev. Physiol. 1991, Vol. 53: 217-39); Streit and Detmar (Oncogene 2003, Vol. 22: 3172-3179) (e.g., Table 3 lists antiangiogenic therapies for malignant melanoma); Ferrara and Alitalo (Nature 2003, Vol. 22: 3172-3179). Medicine [Nature Medicine] 1999, Vol. 5(12): 1359-1364); Tonini et al. (Oncogene [Cancer Gene] 2003, Vol. 22: 6549-6556); and Sato (Int. J. Clin. Oncol. [International Journal of Clinical Oncology] 2003, Vol. 8: 200-206; for example, Table 1 lists antiangiogenic agents used in clinical trials).

[0136] As used herein, "growth inhibitor" refers to a compound or composition that inhibits the growth of cells (such as cells expressing VEGF) in vitro or in vivo. Thus, a growth inhibitor can be an agent that significantly reduces the percentage of cells in the S phase (such as cells expressing VEGF). Examples of growth inhibitors include, but are not limited to, agents that block cell cycle progression (outside the S phase), such as agents that induce G1 arrest and M phase arrest. Classical M phase blockers include vinca (vincristine and vinblastine), taxanes, and topoisomerase II inhibitors, such as doxorubicin, epirubicin, daunorubicin, etoposide, and bleomycin. Agents that cause G1 arrest also cause S phase arrest, such as DNA alkylating agents, such as tamoxifen, prednisone, dacarbazine, nitrogen mustard, cisplatin, methotrexate, 5-fluorouracil, and ara-C. Further information can be found in Mendelsohn and Israel, eds., The Molecular Basis of Cancer, Chapter 1, by Murakami et al., entitled "Cell cycle regulation, oncogenes, and antineoplastic drugs" (WB Saunders, Philadelphia, 1995), e.g., page 13. Taxanes (paclitaxel and docetaxel) are anticancer drugs both derived from the yew tree. Docetaxel (Rhone Blanc Roller), derived from the European yew tree, is a semisynthetic analog of paclitaxel (Bristol-Myers Squibb). Paclitaxel and docetaxel promote the assembly of microtubules from tubulin dimers and stabilize microtubules by preventing depolymerization, thereby inhibiting mitosis in cells.

[0137] The term "anti-tumor composition" refers to a composition comprising at least one active therapeutic agent that can be used to treat cancer. Examples of therapeutic agents include, but are not limited to, chemotherapeutic agents, growth inhibitors, cytotoxic agents, agents used in radiation therapy, anti-angiogenic agents, cancer immunotherapeutic agents (also known as immuno-oncology agents), apoptotic agents, anti-tubulin agents, and other agents for treating cancer, such as anti-HER-2 antibodies, anti-CD20 antibodies, epidermal growth factor receptor (EGFR) antagonists (e.g., tyrosine kinase inhibitors), HER1 / EGFR inhibitors (e.g., erlotinib), platelet-derived growth factor inhibitors (e.g., imatinib mesylate), COX-2 inhibitors (e.g., celecoxib), interferons, CTLA4 inhibitors (e.g., anti-CTLA antibody ipilimumab), PD-L1 inhibitors, and cytotoxic agents. -1 inhibitors (e.g., anti-PD1 antibodies, BMS-936558), PDL1 inhibitors (e.g., anti-PDL1 antibodies, MPDL3280A), PDL2 inhibitors (e.g., anti-PDL2 antibodies), VISTA inhibitors (e.g., anti-VISTA antibodies), cytokines, antagonists (e.g., neutralizing antibodies) that bind to one or more of the following targets (ErbB2, ErbB3, ErbB4, PDGFR-β, BlyS, APRIL, BCMA, PD-1, PDL1, PDL2, CTLA4, VISTA or VEGF receptors), TRAIL / Apo2 and other biologically active and organic chemical agents, etc. Combinations thereof are also included in the present invention.

[0138] Checkpoint inhibitor therapy is a current form of cancer treatment that uses immune checkpoints, which affect immune system function. Immune checkpoints can be either stimulatory or inhibitory. Tumors can use these checkpoints to protect themselves from immune system attack. Checkpoint therapy blocks inhibitory checkpoints and restores immune system function. Checkpoint proteins include programmed cell death 1 protein (PDCD1, PD-1; also known as CD279) and its ligand PD-1 ligand 1 (PD-L1, CD274), cytotoxic T lymphocyte-associated protein 4 (CTLA-4), A2AR (adenosine A2A receptor), B7-H3 (or CD276), B7-H4 (or VTCN1), BTLA (B and T lymphocyte attenuation factor or CD272), IDO (indoleamine 2,3-dioxygenase), KIR (killer cell immunoglobulin-like receptor), LAG3 (lymphocyte activation gene-3), TIM-3 (T cell immunoglobulin domain and mucin domain molecule 3), and VISTA (V-domain Ig inhibitor of T cell activation).

[0139] Programmed cell death protein 1 (also known as PD-1 and CD279 (cluster of differentiation 279)) is a cell surface receptor that plays an important role in downregulating the immune system and promoting self-tolerance by suppressing T cell inflammatory activity. PD-1 is an immune checkpoint and prevents autoimmunity through a dual mechanism of promoting apoptosis (programmed cell death) of antigen-specific T cells in the lymph nodes while reducing apoptosis of regulatory T cells (anti-inflammatory, suppressive T cells).

[0140] PD-1 has two ligands, PD-L1 and PD-L2, which are members of the B7 family. PD-L1 protein is upregulated on macrophages and dendritic cells (DCs) in response to LPS and GM-CSF treatment, and on T cells and B cells during TCR and B cell receptor signaling, while in resting mice, PD-L1 mRNA can be detected in the heart, lungs, thymus, spleen, and kidneys. After treatment with IFN-γ, PD-L1 is expressed on almost all murine tumor cell lines (including PA1 myeloma, P815 mastocytoma, and B16 melanoma). PD-L2 expression is more restricted and is mainly expressed by DCs and some tumor lines.

[0141] CTLA4 or CTLA-4 (cytotoxic T lymphocyte-associated protein 4) (also known as CD152 (cluster of differentiation 152)) is a protein receptor that acts as an immune checkpoint to downregulate the immune response. CTLA4 is constitutively expressed in regulatory T cells, but is only upregulated in conventional T cells after activation - a phenomenon that is particularly noteworthy in cancer. CTLA4 is a member of the immunoglobulin superfamily that is expressed by activated T cells and transmits inhibitory signals to T cells. CTLA4 is homologous to the T cell co-stimulatory protein CD28, and both molecules bind to CD80 and CD86 (also known as B7-1 and B7-2, respectively) on antigen-presenting cells. CTLA-4 binds to CD80 and CD86 with a greater affinity and avidity than it does to CD28, thus enabling it to outcompete CD28 compared to its ligand. CTLA4 transmits inhibitory signals to T cells, while CD28 transmits stimulatory signals. CTLA4 is also found in regulatory T cells and contributes to their inhibitory function. T cell activation via the T cell receptor and CD28 leads to increased expression of CTLA-4.

[0142] There are several checkpoint inhibitors currently used to treat cancer. PD-1 inhibitors include pembrolizumab (Keytruda) and nivolumab (Opdivo). PD-L1 inhibitors include atezolizumab (Tecentriq), avelumab (Bavencio), and durvalumab (Imfinzi). CTLA-4 inhibitors include iplimumab (Yervoy). There are several other checkpoint inhibitors in development, including an anti-B7-H3 antibody (MGA271), an anti-KIR antibody (lirilumab), and an anti-LAG3 antibody (BMS-986016).

[0143] In additional embodiments, the present invention provides polynucleotides encoding the heavy or light chain, or antigen-binding portion thereof, of any of the antibodies or antigen-binding fragments thereof described herein.

[0144] The present invention also provides nucleic acid molecules comprising polynucleotides encoding one or more chains of an antibody (e.g., an anti-CCR8 antibody) described herein. In some embodiments, the nucleic acid molecules comprise polynucleotides encoding the heavy chain or light chain of an antibody described herein. In some embodiments, the nucleic acid molecules comprise both polynucleotides encoding the heavy chain of an antibody described herein and polynucleotides encoding the light chain of an antibody described herein. In some embodiments, the first nucleic acid molecule comprises a first polynucleotide encoding a heavy chain, and the second nucleic acid molecule comprises a second polynucleotide encoding a light chain.

[0145] In some such embodiments, the heavy and light chains are expressed from one nucleic acid molecule, or are expressed as two separate polypeptides from two separate nucleic acid molecules. In some embodiments, such as when the antibody is an scFv, a single polynucleotide encodes a single polypeptide comprising both the heavy and light chains linked together.

[0146] In some embodiments, the polynucleotide encoding the heavy chain or light chain of the antibody described herein comprises a nucleotide sequence encoding a leader sequence that is located at the N-terminus of the heavy chain or light chain when translated. As discussed above, the leader sequence can be a natural heavy chain or light chain leader sequence, or can be another heterologous leader sequence.

[0147] As used herein, the term "nucleic acid" or "oligonucleotide" refers to a polynucleotide, such as deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). Nucleic acids include, but are not limited to, genomic DNA, cDNA, mRNA, iRNA, miRNA, tRNA, ncRNA, rRNA, and recombinantly produced and chemically synthesized molecules, such as aptamers, plasmids, antisense DNA strands, shRNA, ribozymes, conjugated nucleic acids, and oligonucleotides. According to the present invention, nucleic acids can exist as single-stranded or double-stranded linear or covalently circularly closed molecules. Nucleic acids can be isolated. The term "isolated nucleic acid" means that nucleic acids (i) are amplified in vitro, for example, via polymerase chain reaction (PCR), (ii) are produced by cloning and recombination, (iii) are purified, for example, by cutting and gel electrophoresis separation, (iv) are synthesized, for example, by chemical synthesis, or (v) are extracted from a sample. Nucleic acids can be used to introduce into cells (i.e., transfected cells), particularly in the form of RNA, which can be prepared from a DNA template by in vitro transcription. In addition, RNA can be modified by stabilizing sequences, capping, and polyadenylation before application.

[0148] In one embodiment, the present invention provides a vector comprising a polynucleotide described herein, wherein the vector is an expression vector selected from the group consisting of a mammalian expression vector, a yeast expression vector, an insect expression vector, and a bacterial expression vector.

[0149] Vectors are provided that contain polynucleotides encoding the heavy and / or light chains of the antibodies described herein. Such vectors include, but are not limited to, DNA vectors, phage vectors, viral vectors, retroviral vectors, and the like. In some embodiments, the vector contains a first polynucleotide sequence encoding the heavy chain and a second polynucleotide sequence encoding the light chain. In some embodiments, the heavy chain and the light chain are expressed as two separate polypeptides from the vector. In some embodiments, such as, for example, when the antibody is an scFv, the heavy chain and the light chain are expressed as part of a single polypeptide.

[0150] In some embodiments, the first vector comprises a polynucleotide encoding a heavy chain, and the second vector comprises a polynucleotide encoding a light chain. In some embodiments, the first vector and the second vector are transfected into a host cell in similar amounts (such as similar molar amounts or similar mass amounts). In some embodiments, a first vector and a second vector with a molar ratio or a mass ratio between 5:1 and 1:5 are transfected into a host cell. In some embodiments, for a vector encoding a heavy chain and a vector encoding a light chain, a mass ratio between 1:1 and 1:5 is used. In some embodiments, for a vector encoding a heavy chain and a vector encoding a light chain, a mass ratio of 1:2 is used.

[0151] In some embodiments, a vector is selected that is optimized for expressing a polypeptide in CHO or CHO-derived cells or in NSO cells. Exemplary such vectors are described, for example, in Running Deer et al., Biotechnol. Prog. 20:880-889 (2004). In some embodiments, a vector is selected for in vivo expression of a CCR8 antagonist in animals (including humans). In some such embodiments, expression of one or more polypeptides is under the control of one or more promoters that function in a tissue-specific manner. For example, liver-specific promoters are described, for example, in PCT Publication No. WO2006 / 076288.

[0152] The terms "vector," "expression vector," or "plasmid DNA" are used herein to refer to a recombinant nucleic acid construct that is manipulated by human intervention. A recombinant nucleic acid construct can contain two or more nucleotide sequences linked in such a way that the product is not found in cells in nature. In particular, two or more nucleotide sequences can be operably linked, such as genes encoding a protein of interest, one or more protein tags, functional domains, and the like.

[0153] Suitable vectors for use in the preparation of proteins and / or protein conjugates include those selected from the following vectors: baculovirus, phage, plasmid, phagemid, cosmid, F cosmid, bacterial artificial chromosome, viral DNA, P1-based artificial chromosome, yeast plasmid and yeast artificial chromosome. For example, viral DNA vectors can be selected from vaccinia, adenovirus, fowl pox virus, pseudorabies virus and SV40 derivatives. One type of vector is a genomic integration vector or an "integration vector" that can be integrated into the chromosomal DNA of a host cell. Another type of vector is an additional vector, such as a nucleic acid capable of extrachromosomal replication. Vectors that can direct the expression of genes to which they are operably linked are referred to herein as "expression vectors." Viral vectors include adenovirus, adeno-associated virus (AAV), retrovirus, lentivirus, vaccinia virus, measles virus, herpes virus, and bovine papilloma virus vectors (for a review of viral and non-viral vectors, see Kay et al., Proc. Natl. Acad. Sci. USA 94:12744-12746 (1997)). Viral vectors are modified so that the natural tropism and pathogenicity of the virus have been altered or removed. The viral genome can also be modified to increase its infectivity and accommodate the packaging of the nucleic acid encoding the polypeptide of interest.

[0154] The nucleic acid constructs of the present invention can be introduced into the cells to be changed, thereby allowing the chimeric protein to be expressed in the cells. A variety of methods are known in the art and are suitable for introducing nucleic acids into cells, including viral and non-viral mediated techniques. Examples of typical non-viral mediated techniques include, but are not limited to, electroporation, calcium phosphate mediated transfer, nuclear transfection, sonoporation, heat shock, magnetofection, liposome mediated transfer, microinjection, microparticle mediated transfer (nanoparticles), cationic polymer mediated transfer (DEAE-dextran, polyethyleneimine, polyethylene glycol (PEG), etc.), or cell fusion. Other transfection methods include proprietary transfection reagents such as Lipofectamine TM 、Dojindo Hilymax TM 、Fugene TM 、jetPEI TM 、Effectene TM and DreamFect TM .

[0155] Nucleic acid construct of the present invention can be introduced into the host cell to be changed, thereby allowing chimeric protein to be expressed in the cell. A variety of host cells are known in the art and are suitable for chimeric protein expression. Examples of typical cells for transfection include, but are not limited to, bacterial cells, eukaryotic cells, yeast cells, insect cells or plant cells. For example, Escherichia coli, Bacillus, Streptomyces, Pichia pastoris, Salmonella typhimurium, Drosophila S2, Spodoptera SJ9, CHO, COS (e.g., COS-7), 3T3-F442A, HeLa, HUVEC, HUAEC, NIH 3T3, Jurkat, 293, 293H or 293F.

[0156] In various embodiments, the heavy and / or light chains of the antibodies described herein can be expressed in prokaryotic cells (such as bacterial cells); or in eukaryotic cells (such as fungal cells (such as yeast), plant cells, insect cells and mammalian cells). Such expression can be performed, 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 COS 7 cells; 293 cells, including 293-6E cells; CHO cells, including CHO-S and DG44 cells; PER. cells (Crucell); and NSO cells. In some embodiments, the heavy and / or light chains of the antibodies described herein can be expressed in yeast. See, for example, U.S. Publication No. US2006 / 0270045A1. In some embodiments, a particular eukaryotic host cell is selected based on its ability to undergo the desired post-translational modification of the heavy and / or light chains of the anti-CCR8 antibody. For example, in some embodiments, the sialylation level of a polypeptide produced in CHO cells is higher than the sialylation level of the same polypeptide produced in 293 cells.

[0157] In various embodiments herein, the anti-CCR8 monoclonal antibodies of the present invention are expressed in cells in which fucosylation is limited or absent, thereby producing antibodies with increased defucosylation. In some aspects, the antibody is selected from (a) an antibody having a heavy chain comprising SEQ ID NO: 3 and a light chain comprising SEQ ID NO: 2; (b) an antibody having a heavy chain comprising SEQ ID NO: 4 and a light chain comprising SEQ ID NO: 2; (c) an antibody having a heavy chain comprising SEQ ID NO: 5 and a light chain comprising SEQ ID NO: 2; (d) an antibody having a heavy chain comprising SEQ ID NO: 6 and a light chain comprising SEQ ID NO: 2; (e) an antibody having a heavy chain comprising SEQ ID NO: 7 and a light chain comprising SEQ ID NO: 2; (f) an antibody having a heavy chain comprising SEQ ID NO: 8 and a light chain comprising SEQ ID NO: 2; and (h) an antibody having a heavy chain comprising SEQ ID NO: 9 and a light chain comprising SEQ ID NO: 2, wherein the antibody specifically binds to CCR8. In preferred aspects, the antibody is an antibody having a heavy chain comprising SEQ ID NO: 6 and a light chain comprising SEQ ID NO: 2. In these embodiments, the antibody has a defucosylation level of at least 50%. For example, the antibody has a defucosylation level of at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more.

[0158] One or more nucleic acids can be introduced into the desired host cells 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, 3rd ed. Cold Spring Harbor Laboratory Press (2001). Nucleic acids can be transiently or stably transfected into the desired host cells according to any suitable method.

[0159] In some embodiments, one or more polypeptides can be produced according to any suitable method in an animal that has been engineered or transfected with one or more nucleic acid molecules encoding the polypeptides.

[0160] Polynucleotides can be delivered to cells (e.g., a plurality of different cells or cell types comprising target cells or cell types and / or non-target cell types) in a vector (e.g., an expression vector). Examples of vectors include, but are not limited to, (a) non-viral vectors, such as nucleic acid vectors, including linear oligonucleotides and circular plasmids; artificial chromosomes, such as human artificial chromosomes (HACs), yeast artificial chromosomes (YACs), and bacterial artificial chromosomes (BACs or PACs); episomal vectors; transposons (e.g., PiggyBacs); and (b) viral vectors, such as retroviral vectors, lentiviral vectors, adenoviral vectors, and AAV vectors. Viral vectors have several advantages for delivering nucleic acids, including high infectivity and / or tropism for certain target cells or tissues. In some cases, viral vectors can be used to deliver the polynucleotides described herein.

[0161] Additional embodiments

[0162] Embodiment 1. An isolated monoclonal antibody or antigen-binding fragment thereof, comprising: a HCVR comprising a HCVRCDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSEQQYYYAMDY (SEQ ID NO: 19); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15), wherein the antibody or antigen-binding fragment thereof specifically binds CCR8.

[0163] Embodiment 2. The isolated monoclonal antibody or antigen-binding fragment thereof as described in embodiment 1, further comprising an HFR1 sequence of SEQ ID NO: 23, an HFR2 sequence of SEQ ID NO: 24, an HFR3 sequence of SEQ ID NO: 25 and / or an HFR4 sequence of SEQ ID NO: 26.

[0164] Embodiment 3. The isolated monoclonal antibody or antigen-binding fragment thereof according to embodiment 1 or 2, further comprising an LFR1 sequence of SEQ ID NO: 27, an LFR2 sequence of SEQ ID NO: 28, an LFR3 sequence of SEQ ID NO: 29 and / or an LFR4 sequence of SEQ ID NO: 30.

[0165] Embodiment 4. The isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-3, wherein the HCVR sequence is SEQ ID NO: 36 and / or the LCVR sequence is SEQ ID NO: 32.

[0166] Embodiment 5. The isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-4, wherein the HCVR sequence is SEQ ID NO: 36 and the LCVR sequence is SEQ ID NO: 32.

[0167] Embodiment 6. The isolated monoclonal antibody of any one of embodiments 1-5, wherein the heavy chain sequence is SEQ ID NO: 6 and the light chain sequence is SEQ ID NO: 2.

[0168] Embodiment 7. An isolated monoclonal antibody or antigen-binding fragment thereof, comprising (a) a HCVR comprising a HCVRR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSEQNYYYAMDY (SEQ ID NO: 16); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (b) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a LCVR CDR3 sequence having the amino acid sequence of TRGSEQNYYYAMDY (SEQ ID NO: 16). NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSENNYYYAMDY (SEQ ID NO: 17); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (c) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSENNYYYAMDY (SEQ ID NO: 18); and a LCVR comprising a LCVR having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13). CDR1 sequence, a LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO: 15);or (d) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSDQNYYYAMDY (SEQ ID NO: 20); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (e) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a LCVR CDR3 sequence having the amino acid sequence of TRGSDNQYYYAMDY (SEQ ID NO: 21). NO: 21); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); or (f) a HCVR comprising a HCVR CDR1 sequence having the amino acid sequence of GFRFNTNA (SEQ ID NO: 10), a HCVR CDR2 sequence having the amino acid sequence of IRSKSNSYATYY (SEQ ID NO: 11), and a HCVR CDR3 sequence having the amino acid sequence of TRGSDQQYYYAMDY (SEQ ID NO: 22); and a LCVR comprising a LCVR CDR1 sequence having the amino acid sequence of QSLLHSNGNTY (SEQ ID NO: 13), a LCVR CDR2 sequence having the amino acid sequence of YRMSNR (SEQ ID NO: 14), and a LCVR CDR3 sequence having the amino acid sequence of MQHLEYPFT (SEQ ID NO: 15); CDR2 sequence and a LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO: 15), wherein the antibody or antigen-binding fragment thereof specifically binds to CCR8.;

[0169] Embodiment 8. The isolated monoclonal antibody or antigen-binding fragment thereof of embodiment 7, further comprising an HFR1 sequence of SEQ ID NO: 23, an HFR2 sequence of SEQ ID NO: 24, an HFR3 sequence of SEQ ID NO: 25, and / or an HFR4 sequence of SEQ ID NO: 26.

[0170] Embodiment 9. The isolated monoclonal antibody or antigen-binding fragment thereof of embodiment 7 or 8, further comprising an LFR1 sequence of SEQ ID NO: 27, an LFR2 sequence of SEQ ID NO: 28, an LFR3 sequence of SEQ ID NO: 28, and / or an LFR4 sequence of SEQ ID NO: 29.

[0171] Embodiment 10. An isolated monoclonal antibody or antigen-binding fragment thereof, comprising (a) a HCVR sequence of SEQ ID NO: 33 and a LCVR sequence of SEQ ID NO: 32; or (b) a HCVR sequence of SEQ ID NO: 34 and a LCVR sequence of SEQ ID NO: 32; or (c) a HCVR sequence of SEQ ID NO: 35 and a LCVR sequence of SEQ ID NO: 32; or (d) a HCVR sequence of SEQ ID NO: 37 and a LCVR sequence of SEQ ID NO: 32; or (e) a HCVR sequence of SEQ ID NO: 38 and a LCVR sequence of SEQ ID NO: 32; or (f) a HCVR sequence of SEQ ID NO: 39 and a LCVR sequence of SEQ ID NO: 32, wherein the antibody or antigen-binding fragment thereof specifically binds to CCR8.

[0172] Embodiment 11. An isolated monoclonal antibody comprising (a) a heavy chain sequence of SEQ ID NO: 3 and a light chain sequence of SEQ ID NO: 2; or (b) a heavy chain sequence of SEQ ID NO: 4 and a light chain sequence of SEQ ID NO: 2; or (c) a heavy chain sequence of SEQ ID NO: 5 and a light chain sequence of SEQ ID NO: 2; or (d) a heavy chain sequence of SEQ ID NO: 7 and a light chain sequence of SEQ ID NO: 2; or (e) a heavy chain sequence of SEQ ID NO: 8 and a light chain sequence of SEQ ID NO: 2; or (f) a heavy chain sequence of SEQ ID NO: 9 and a light chain sequence of SEQ ID NO: 2, wherein the antibody or antigen-binding fragment thereof specifically binds to CCR8.

[0173] Embodiment 12. An isolated monoclonal antibody or antigen-binding fragment thereof, which is at least 80% identical to the antibody of any one of embodiments 5, 6, 10 or 11, wherein the antibody or antigen-binding fragment thereof specifically binds CCR8.

[0174] Embodiment 13. The isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-12, wherein the antibody is a humanized antibody.

[0175] Embodiment 14. The isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-13, wherein the antigen-binding fragment thereof is Fab, Fab', F(ab')2, Fd, single-chain Fv or scFv, disulfide-linked Fv, V-NAR domain, IgNar, intrabody, IgGACH2, minibody, F(ab')3, tetrabody, triabody, diabody, domain antibody (dAb), DVD-Ig, Fcab, mAb2, (scFv)2, tandem scFv, TandAb, nanobody or scFv-Fc.

[0176] Embodiment 15. The isolated monoclonal antibody or antigen-binding fragment thereof of any one of Embodiments 1-14, wherein the monoclonal antibody or antigen-binding fragment thereof binds to human CCR8 with a Kd of less than about 25 nM, 20 nM, 15 nM, 10 nM, 5 nM, 2 nM, 1 nM, 0.5 nM, or 0.01 nM.

[0177] Embodiment 16. The isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-15, which induces ADCC.

[0178] Embodiment 17. The isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-16, wherein the antibody has a defucosylation level of at least 50%.

[0179] Embodiment 18. The isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-17, wherein the antibody has a defucosylation level of at least 85%.

[0180] Embodiment 19. A method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof according to any one of embodiments 1-18, thereby treating the cancer in the subject.

[0181] Embodiment 20. A method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof according to any one of embodiments 1-18 and an anti-cancer agent, thereby treating the cancer in the subject.

[0182] Embodiment 21. The method of embodiment 19 or 20, wherein the cancer is a hematological cancer or a solid tumor.

[0183] Embodiment 22. The method of embodiment 21, wherein the blood cancer is selected from the group consisting of acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia, chronic myeloid leukemia and hairy cell leukemia, AIDS-related lymphoma, cutaneous T-cell lymphoma, Hodgkin lymphoma, Hodgkin lymphoma, mycosis fungoides, non-Hodgkin lymphoma, primary central nervous system lymphoma, Sezary syndrome, cutaneous T-cell lymphoma, Waldenstrom's macroglobulinemia, diffuse large B-cell lymphoma (DLBCL) and multiple myeloma.

[0184] Embodiment 23. The method of embodiment 21, wherein the solid tumor is breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colon cancer, kidney cancer, ovarian cancer, liver cancer, or prostate cancer.

[0185] Embodiment 24. The method of embodiment 19, further comprising administering an anti-cancer agent to the patient.

[0186] Embodiment 25. The method of embodiment 20 or 24, wherein the anti-cancer agent is a chemotherapeutic agent, an anti-angiogenic agent, a growth inhibitory agent, an immuno-oncology agent, a checkpoint inhibitor and / or an anti-tumor composition.

[0187] Embodiment 26. A method of treating breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colon cancer, kidney cancer, ovarian cancer, liver cancer, and / or prostate cancer in a subject in need thereof, the method comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-18, thereby treating the cancer in the subject.

[0188] Embodiment 27. A method of treating breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colon cancer, kidney cancer, ovarian cancer, liver cancer, and / or prostate cancer in a subject in need thereof, the method comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof according to any one of embodiments 1-18 and an anti-cancer agent, thereby treating the cancer in the subject.

[0189] Embodiment 28. The method of embodiment 27, wherein the anti-cancer agent is a chemotherapeutic agent, an anti-angiogenic agent, a growth inhibitory agent, an immuno-oncology agent, a checkpoint inhibitor and / or an anti-tumor composition.

[0190] Embodiment 29. A method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof according to any one of embodiments 1-6, thereby treating the cancer in the subject.

[0191] Embodiment 30. A method of treating breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colon cancer, kidney cancer, ovarian cancer, liver cancer, and / or prostate cancer in a subject in need thereof, the method comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof of any one of embodiments 1-6, thereby treating the cancer in the subject.

[0192] Embodiment 31. A method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof according to any one of embodiments 1-6 and an anti-cancer agent, thereby treating the cancer in the subject.

[0193] Embodiment 32. A method of treating breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colon cancer, kidney cancer, ovarian cancer, liver cancer and / or prostate cancer in a subject in need thereof, the method comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof according to any one of embodiments 1-6 and an anti-cancer agent, thereby treating the cancer in the subject.

[0194] Embodiment 33. The method of embodiment 29 or 31, wherein the cancer is a hematological cancer or a solid tumor.

[0195] Embodiment 34. The method of embodiment 33, wherein the blood cancer is selected from the group consisting of acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia, chronic myeloid leukemia and hairy cell leukemia, AIDS-related lymphoma, cutaneous T-cell lymphoma, Hodgkin lymphoma, Hodgkin lymphoma, mycosis fungoides, non-Hodgkin lymphoma, primary central nervous system lymphoma, Sezary syndrome, cutaneous T-cell lymphoma, Waldenstrom's macroglobulinemia, diffuse large B-cell lymphoma (DLBCL) and multiple myeloma.

[0196] Embodiment 35. The method of embodiment 33, wherein the solid tumor is breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colon cancer, kidney cancer, ovarian cancer, liver cancer, or prostate cancer.

[0197] Embodiment 36. The method of embodiment 29 or 30, further comprising administering an anti-cancer agent to the patient.

[0198] Embodiment 37. The method of embodiment 36, wherein the anti-cancer agent is a chemotherapeutic agent, an anti-angiogenic agent, a growth inhibitory agent, an immuno-oncology agent, a checkpoint inhibitor and / or an anti-tumor composition.

[0199] Embodiment 38. A polynucleotide encoding the heavy chain or light chain or antigen binding portion thereof as described in any one of embodiments 1-18.

[0200] Embodiment 39. A vector comprising the polynucleotide of embodiment 38, wherein the vector is an expression vector selected from the group consisting of a mammalian expression vector, a yeast expression vector, an insect expression vector, and a bacterial expression vector.

[0201] Embodiment 40. A method of producing the monoclonal antibody of embodiment 6 or 11, wherein the antibody is at least 50%, 60%, 70%, 80%, 85%, 90%, 95% or 99% afucosylated, the method comprising expressing a polynucleotide expressing the antibody in a cell engineered to reduce or prevent fucosylation of the antibody.

[0202] Presented below are examples discussing isolated anti-CCR8 antibodies as described herein. The following examples are provided to further illustrate embodiments of the present invention and are not intended to limit the scope of the invention. While they are representative of those that can be used, other procedures, methods, or techniques known to those skilled in the art may alternatively be used.

[0203] Example

[0204] Example 1

[0205] Defucosylation of humanized anti-CCR8 antibody

[0206] To evaluate the impact of defucosylation levels on the properties of Ab001_WT, ATUM's proprietary hypofucosylation technology (miFuc) was applied during cell line generation. TM ) to increase the defucosylation level to >90%. TMThe second comparison cell line for the hypofucosylation technique produced approximately 11% defucosylation. The ADCC potency of the antibody was evaluated using a HuT78 cell reporter assay using Jurkat-CD16A-NFAT. An approximately 100-fold increase in ADCC activity was observed in the hypofucosylated cell line relative to the non-hypofucosylated cell line (see Tables 1 and 2). Figure 1 ).

[0207] Table 1: EC50 results of ADCC reporter assay using Hut78 target cells

[0208]

[0209] Example 2

[0210] CCR8 binding assay

[0211] The antibodies were tested for binding to CCR8 using the CHO-K1 human chemokine CCR8 receptor cell line (CHO.hCCR8; Perkin Elmer Corp., Waltham, MA) as follows. Approximately 5×10 4 Each CHO.hCCR8 cell was resuspended in 50 μL of serially diluted antibodies (antibody concentrations of 10, 3.3, 1.1, 0.37, 0.12, 0.04, and 0.01 nM) in a well of a V-bottom 96-well plate (Miltenyi Biotec, Inc., San Jose, California). After incubation on ice for 30 min, 150 μL of cold MACS buffer was added to each well, the plate was centrifuged at 500 x g for 5 min at 4 ° C, and the supernatant was discarded. The cells were washed twice in 200 μL of cold MACS buffer and then resuspended in 50 μL of anti-human IgG-Alexa Fluor 488 (The Jackson Laboratory, Bar Harbor, Maine) diluted 1:100 in MACS buffer. After incubation on ice for 30 min, 150 μL cold MACS buffer was added to each well, the plate was centrifuged at 500 x g for 5 min at 4 ° C, and the supernatant was discarded. The cells were washed twice in 200 μL cold MACS buffer and then resuspended in 80 μL cold MACS buffer with propidium iodide (PI, 1:1000 dilution, 1 μg / ml; ThermoFisher Scientific, Waltham, Massachusetts). Antibody binding (luminescence) was measured using the CellStream flow cytometry system (Cytek Biosciences, Fremont, California).

[0212] Example 3

[0213] Antibody-dependent cellular cytotoxicity (ADCC) assay

[0214] CHO.hCCR8 cells were cultured at 1x10 4 The cells were plated at a density of 100 cells / well in 100 μL Ham's F-12 medium (10% FBS, 0.4 mg / mL G418) in a white, clear-bottomed 96-well plate. After 24 hours, the culture medium was removed and 40 μL of assay medium (one-step luciferase assay system; BPS Bioscience, San Diego, California) was added. In the assay medium, the antibody was diluted 1:10, then serially diluted 1:4, and 10 μL of each was dispensed into the wells and incubated for 1 hour. 5x10 4 ADCC bioassay effector cell V variant (high affinity)-Jurkat recombinant cell line (BPS Biosciences) cells were added to each well in 50 μL aliquots of each cell / well. The plate was incubated at 37°C, 5% CO2 for 5 hours, followed by the addition of 100 μL luciferase assay working solution (luciferase reagent substrate diluted 1:100 in luciferase reagent buffer; BPS Biosciences), and the plate was gently shaken for 20 minutes at room temperature. Luminescence was then measured using a luminometer.

[0215] Example 4

[0216] Deamidation of AB001_WT antibody

[0217] Ab001_Wt was incubated at 37°C in PBS, pH 7.4, and samples were collected on days 0, 1, 4, 7, 14, 21, and 28. The samples were assayed for human CCR8 binding to EC at each time point. 50 、ADCC EC 50 and the percentage of deamidation at N104 and / or N105. Figure 2A and Figure 2B As shown in the figure, hCCR8 binding and ADCC activity decreased over time. Figure 2C As shown, the observed decrease is consistent with the deamidation of the Ab001_Wt antibody over the same time period. This demonstrates that Ab001_Wt has a defect that reduces the effectiveness of the antibody over time and suggests that frequent dosing of the antibody is required to maintain the activity level of Ab001_Wt.

[0218] In a second experiment, plasma samples from monkey pharmacokinetic (PK) studies demonstrated CCR8 binding ( Figure 3A ) and ADCC activity ( Figure 3B) Both of them decreased sharply over time. This proves that Ab001_Wt has the same defect in animals.

[0219] Example 5

[0220] Construction and testing of mutant antibodies

[0221] The HCVR CDR3 of Ab001_Wt was modified to generate a series of antibodies with reduced deamidation sensitivity. These antibodies modified amino acids 103 to 105 of the HCVR CDR3 sequence from DNN in the parent antibody Ab001_Wt to ​​EQN in Ab001_M1, ENN in Ab001_M2, ENQ in Ab001_M3, EQQ in Ab001_M4, DQN in Ab001_M5, DNQ in Ab001_M6, and DQQ in Ab001_M7. The mutant antibodies were tested for CCR8 binding affinity and ADCC activity relative to the parent antibody Ab001_Wt.

[0222] like Figure 4A and Figure 4B As shown in the figure, the CCR8 binding of Ab001-M1, M2, M3 and M4 was measured and compared with the binding activity of Ab001-Wt. EC50 is summarized in Figure 4C Although modifications in the CDR3 region have been shown to affect EC binding 50 104 and N105 in Ab001_M4 did not produce significant changes. Thus, Ab001_M4 eliminates the deamidation defect over time and retains CCR8 binding activity.

[0223] like Figure 5A and Figure 5B As shown in the figure, the ADCC activity of AbOOl-M1, M2, M3 and M4 was measured and compared with the ADCC activity of AbOOl-Wt. EC50 is summarized in Figure 5C As seen with CCR8 binding, although modifications in the CDR3 region showed an effect on ADCC ECs 50 104 and N105 in Ab001_M4 did not produce significant changes. Thus, Ab001_M4 eliminates the deamidation defect over time and retains ADCC activity.

[0224] Although the present invention has been described with reference to the above embodiments, it will be understood that modifications and variations are encompassed within the spirit and scope of the invention. Accordingly, the present invention is limited only by the following claims.

[0225] sequence:

[0226] Underline :Signal sequence

[0227] mutation

[0228] >Ab001_WT HC (wild type) (SEQ ID NO: 1)

[0229]

[0230] >Ab001_WT LC (wild type and all variants below) (SEQ ID NO: 2)

[0231] METDTLLLWVLLLWVPGSTG DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGNTYLYWFLQKPGQSPQLLIYR

[0232] MSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHLEYPFTFGQGTKLEIKRTVAAPSVFIFPPSDEQL

[0233] KSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQG

[0234] LSSPVTKSFNRGEC

[0235] >Ab001_M1 HC(EQN)(SEQ ID NO:3)

[0236]

[0237] >Ab001_M2 HC(ENN)(SEQ ID NO:4)

[0238]

[0239] >Ab001_M3 HC(ENQ)(SEQ ID NO:5)

[0240]

[0241] >Ab001_M4 HC(EQQ)(SEQ ID NO:6)

[0242]

[0243] >Ab001_M5 HC(DQN)(SEQ ID NO:7)

[0244]

[0245] >Ab001_M6 HC(DNQ)(SEQ ID NO:8)

[0246]

[0247] >Ab001_M7 HC(DQQ)(SEQ ID NO:9)

[0248]

[0249] LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0250] >Ab001_WT and M1-M7 HCDR1(SEQ ID NO:10)

[0251] GFRFNTNA

[0252] >Ab001_WT and M1-M7 HCDR2(SEQ ID NO:11)

[0253] IRSKSNSYATYY

[0254] >Ab001_WT HCDR3(SEQ ID NO:12)

[0255] TRGSDNNYYYAMDY

[0256] >Ab001_WT and M1-M7 LCDR1(SEQ ID NO:13)

[0257] QSLLHSNGNTY

[0258] >Ab001_WT and M1-M7 LCDR2(SEQ ID NO:14)

[0259] YRMSNR

[0260] >Ab001_WT and M1-M7 LCDR3(SEQ ID NO:15)

[0261] MQHLEYPFT

[0262] >Ab001_M1 HCDR3(SEQ ID NO:16)

[0263] TRGSEQNYYYAMDY

[0264] >Ab001 M2 HCDR3(SEQ ID NO:17)

[0265] TRGSENNYYYAMDY

[0266] >Ab001 M3 HCDR3(SEQ ID NO:18)

[0267] TRGSENQYYYAMDY

[0268] >Ab001 M4 HCDR3(SEQ ID NO:19)

[0269] TRGSEQQYYYAMDY

[0270] >Ab001 M5 HCDR3(SEQ ID NO:20)

[0271] TRGSDQNYYYAMDY

[0272] >Ab001 M6 HCDR3(SEQ ID NO:21)

[0273] TRGSDNQYYYAMDY

[0274] >Ab001 M7 HCDR3(SEQ ID NO:22)

[0275] TRGSDQQYYYAMDY

[0276] >Ab001 WT and M1-M7 HFR1(SEQ ID NO:23)

[0277] EVQLVESGGGLVQPGGSLKLSCAAS

[0278] >Ab001 WT and M1-M7 HFR2(SEQ ID NO:24)

[0279] MNWVRQASGKDLEWVAR

[0280] >Ab001 WT and M1-M7 HFR3(SEQ ID NO:25)

[0281] AASVKGRFTISRDDSKNTLYLQMNSLKTEDTAVYYC

[0282] >Ab001 WT and M1-M7 HFR4 (SEQ ID NO: 26)

[0283] WGQGTTVTVSS

[0284] >Ab001 WT and M1-M7 LFR1 (SEQ ID NO: 27)

[0285] DIVMTQSPLSLPVTPGEPASISCRSS

[0286] >Ab001 WT and M1-M7 LFR2 (SEQ ID NO: 28)

[0287] LYWFLQKPGQSPQLLI

[0288] >Ab001 WT and M1-M7 LFR3 (SEQ ID NO: 29)

[0289] ASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYC

[0290] >Ab001 WT and M1-M7 LFR4 (SEQ ID NO: 30)

[0291] FGQGTKLEIK

[0292] >Ab001_WT HCVR (wild type) (SEQ ID NO: 31)

[0293]

[0294] >Ab001_WT LCVR (wild type and all variants below) (SEQ ID NO: 32)

[0295] METDTLLLWVLLLWVPGSTG DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGNTYLYWFLQKPGQSPQLLIYRMSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHLEYPFTFGQGTKLEIK

[0296] >Ab001_M1 HC(EQN)(SEQ ID NO:33)

[0297]

[0298] >Ab001_M2 HC(ENN)(SEQ ID NO:34)

[0299]

[0300] >Ab001_M3 HC(ENQ)(SEQ ID NO:35)

[0301]

[0302] >Ab001_M4 HC(EQQ)(SEQ ID NO:36)

[0303]

[0304] >Ab001_M5 HC(DQN)(SEQ ID NO:37)

[0305]

[0306] >Ab001_M6 HC(DNQ)(SEQ ID NO:38)

[0307]

[0308] >Ab001_M7 HC(DQQ)(SEQ ID NO:39)

[0309]

Claims

1. An isolated monoclonal antibody or antigen-binding fragment thereof, said isolated monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain variable region (HCVR) comprising a HCVR complementarity-determining region (CDR1) sequence having the amino acid sequence GFRFNTNA (SEQ ID NO:10), a HCVR CDR2 sequence having the amino acid sequence IRSKSNSYATYY (SEQ ID NO:11), and a HCVR CDR3 sequence having the amino acid sequence TRGSEQQYYYAMDY (SEQ ID NO:19); and a light chain variable region (LCVR) comprising a LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO:13), a LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO:14), and a LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO:15), wherein said antibody or antigen-binding fragment thereof specifically binds to CCR8.

2. An isolated monoclonal antibody or antigen-binding fragment thereof, said isolated monoclonal antibody or antigen-binding fragment thereof comprising: a HC comprising a sequence having at least 80% identity to SEQ ID NO:6 and having its antigen-binding specificity; and a LC comprising a sequence having at least 80% identity to SEQ ID NO:2 and having its antigen-binding specificity.

3. The isolated monoclonal antibody or antigen-binding fragment thereof according to claim 2, wherein: the heavy chain variable region (HCVR) sequence is SEQ ID NO:36 and / or the light chain variable region (LCVR) sequence is SEQ ID NO:

32.

4. The isolated monoclonal antibody or antigen-binding fragment thereof according to claim 2 or 3, wherein: the HCVR sequence is SEQ ID NO:36 and the LCVR sequence is SEQ ID NO:

32.

5. The isolated monoclonal antibody or antigen-binding fragment thereof according to claim 2, 3 or 4, wherein: the HC sequence is SEQ ID NO:6 and the LC sequence is SEQ ID NO:

2.

6. An isolated monoclonal antibody or antigen-binding fragment thereof, said isolated monoclonal antibody or antigen-binding fragment thereof comprising: (a) A heavy chain variable region (HCVR) comprising an HCVR CDR1 sequence having the amino acid sequence GFRFNTNA (SEQ ID NO:10), an HCVR CDR2 sequence having the amino acid sequence IRSKSNSYATYY (SEQ ID NO:11), and an HCVR CDR3 sequence having the amino acid sequence TRGSEQNYYYAMDY (SEQ ID NO:16); and a light chain variable region (LCVR) comprising an LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO:13), an LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO:14), and an LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO:15); or (b) A heavy chain variable region (HCVR) comprising an HCVR CDR1 sequence having the amino acid sequence GFRFNTNA (SEQ ID NO:10), an HCVR CDR2 sequence having the amino acid sequence IRSKSNSYATYY (SEQ ID NO:11), and an HCVR CDR3 sequence having the amino acid sequence TRGSENNYYYAMDY (SEQ ID NO:17); and a light chain variable region (LCVR) comprising an LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO:13), an LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO:14), and an LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO:15); or (c) A heavy chain variable region (HCVR) comprising an HCVR CDR1 sequence having the amino acid sequence GFRFNTNA (SEQ ID NO:10), an HCVR CDR2 sequence having the amino acid sequence IRSKSNSYATYY (SEQ ID NO:11), and an HCVR CDR3 sequence having the amino acid sequence TRGSENQYYYAMDY (SEQ ID NO:18); and a light chain variable region (LCVR) comprising an LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO:13), an LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO:14), and an LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO:15); or (d) A heavy chain variable region (HCVR) comprising an HCVR CDR1 sequence having the amino acid sequence GFRFNTNA (SEQ ID NO:10), an HCVR CDR2 sequence having the amino acid sequence IRSKSNSYATYY (SEQ ID NO:11), and an HCVR CDR3 sequence having the amino acid sequence TRGSDQNYYYAMDY (SEQ ID NO:20); and a light chain variable region (LCVR) comprising an LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO:13), an LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO:14), and an LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO:15); or (e) A heavy chain variable region (HCVR) comprising an HCVR CDR1 sequence having the amino acid sequence GFRFNTNA (SEQ ID NO:10), an HCVR CDR2 sequence having the amino acid sequence IRSKSNSYATYY (SEQ ID NO:11), and an HCVR CDR3 sequence having the amino acid sequence TRGSDNQYYYAMDY (SEQ ID NO:21); and a light chain variable region (LCVR) comprising an LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO:13), an LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO:14), and an LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO:15); or (f) A heavy chain variable region (HCVR) comprising an HCVR CDR1 sequence having the amino acid sequence GFRFNTNA (SEQ ID NO:10), an HCVR CDR2 sequence having the amino acid sequence IRSKSNSYATYY (SEQ ID NO:11), and an HCVR CDR3 sequence having the amino acid sequence TRGSDQQYYYAMDY (SEQ ID NO:22); and a light chain variable region (LCVR) comprising an LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO:13), an LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO:14), and an LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO:15), wherein the antibody or antigen-binding fragment thereof specifically binds to CCR8.

7. An isolated monoclonal antibody or antigen-binding fragment thereof, said isolated monoclonal antibody or antigen-binding fragment thereof comprising: (a) A heavy chain (HC) comprising a sequence having at least 80% identity to SEQ ID NO:3 and having its antigen-binding specificity; and a light chain (LC) comprising a sequence having at least 80% identity to SEQ ID NO:2 and having its antigen-binding specificity; (b) A HC comprising a sequence having at least 80% identity to SEQ ID NO:4 and having its antigen-binding specificity; and a LC comprising a sequence having at least 80% identity to SEQ ID NO:2 and having its antigen-binding specificity; (c) A HC comprising a sequence having at least 80% identity to SEQ ID NO:5 and having its antigen-binding specificity; and a LC comprising a sequence having at least 80% identity to SEQ ID NO:2 and having its antigen-binding specificity; (d) A HC comprising a sequence having at least 80% identity to SEQ ID NO:7 and having its antigen-binding specificity; and a LC comprising a sequence having at least 80% identity to SEQ ID NO:2 and having its antigen-binding specificity; (e) A HC comprising a sequence having at least 80% identity to SEQ ID NO:8 and having its antigen-binding specificity; and a LC comprising a sequence having at least 80% identity to SEQ ID NO:2 and having its antigen-binding specificity; or (f) A HC comprising a sequence having at least 80% identity to SEQ ID NO:9 and having its antigen-binding specificity; and a LC comprising a sequence having at least 80% identity to SEQ ID NO:2 and having its antigen-binding specificity, wherein said antibody or antigen-binding fragment thereof specifically binds to CCR8.

8. The isolated monoclonal antibody or antigen-binding fragment thereof according to claim 6, wherein: (a) said HCVR sequence is SEQ ID NO:33 and / or said LCVR sequence is SEQ ID NO:32, (b) said HCVR sequence is SEQ ID NO:34 and / or said LCVR sequence is SEQ ID NO:32, (c) said HCVR sequence is SEQ ID NO:35 and / or said LCVR sequence is SEQ ID NO:32, (d) said HCVR sequence is SEQ ID NO:37 and / or said LCVR sequence is SEQ ID NO:32, (e) said HCVR sequence is SEQ ID NO:38 and / or said LCVR sequence is SEQ ID NO:32, (f) said HCVR sequence is SEQ ID NO:39 and / or said LCVR sequence is SEQ ID NO:

32.

9. The isolated monoclonal antibody or antigen-binding fragment thereof according to claim 6 or 8, wherein: (a) the HCVR sequence is SEQ ID NO:33 and the LCVR sequence is SEQ ID NO:32, (b) the HCVR sequence is SEQ ID NO:34 and the LCVR sequence is SEQ ID NO:32, (c) the HCVR sequence is SEQ ID NO:35 and the LCVR sequence is SEQ ID NO:32, (d) the HCVR sequence is SEQ ID NO:37 and the LCVR sequence is SEQ ID NO:32, (e) the HCVR sequence is SEQ ID NO:38 and the LCVR sequence is SEQ ID NO:32, (f) the HCVR sequence is SEQ ID NO:39 and the LCVR sequence is SEQ ID NO:

32.

10. The isolated monoclonal antibody or antigen-binding fragment thereof according to claim 6, 8 or 9, wherein: the HC sequence is SEQ ID NO:3 and the LC sequence is SEQ ID NO:2, the HC sequence is SEQ ID NO:4 and the LC sequence is SEQ ID NO:2, the HC sequence is SEQ ID NO:5 and the LC sequence is SEQ ID NO:2, the HC sequence is SEQ ID NO:7 and the LC sequence is SEQ ID NO:2, the HC sequence is SEQ ID NO:8 and the LC sequence is SEQ ID NO:2, or the HC sequence is SEQ ID NO:9 and the LC sequence is SEQ ID NO:

2.

11. The isolated monoclonal antibody or antigen-binding fragment thereof according to any one of claims 1-10, wherein the antibody is a humanized antibody.

12. The isolated monoclonal antibody or antigen-binding fragment thereof according to any one of claims 1 to 11, wherein the antigen-binding fragment thereof is Fab, Fab', F(ab') 2 、F d , single-chain Fv or scFv, disulfide-linked F v 、V-NAR domain、IgNar、Intracellular antibody、IgGACH 2 , mini antibodies, F(ab') 3 , tetrabodies, tribodies, diabodies, single domain antibodies, DVD-Ig, Fcab, mAb 2 , (scFv) 2 or scFv-Fc.

13. The isolated monoclonal antibody or antigen-binding fragment thereof according to any one of claims 1-12, wherein the monoclonal antibody or antigen-binding fragment thereof has a K of less than about 25 nM, 20 nM, 15 nM, 10 nM, 5 nM, 2 nM, 1 nM, 0.5 nM or 0.01 nM d binds to human CCR8.

14. The isolated monoclonal antibody or antigen-binding fragment thereof according to any one of claims 1-13, wherein the isolated monoclonal antibody or antigen-binding fragment thereof induces antibody-dependent cell-mediated cytotoxicity (ADCC).

15. The isolated monoclonal antibody or antigen-binding fragment thereof according to any one of claims 1-14, wherein the antibody has a fucosylation level of at least 50%.

16. The isolated monoclonal antibody or antigen-binding fragment thereof according to any one of claims 1-15, wherein the antibody has a fucosylation level of at least 85%.

17. A method of treating cancer in a subject in need thereof, the method comprising administering to the subject an effective amount of the isolated monoclonal antibody or antigen-binding fragment thereof according to any one of claims 1-16, thereby treating the cancer in the subject.

18. The method according to claim 17, wherein the cancer is a hematological cancer or a solid tumor.

19. The method according to claim 18, wherein the blood cancer is selected from the group consisting of: acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia, chronic myelogenous leukemia, hairy cell leukemia, AIDS-related lymphoma, cutaneous T-cell lymphoma, Hodgkin lymphoma, Hodgkin lymphoma, mycosis fungoides, non-Hodgkin lymphoma, primary central nervous system lymphoma, Sézary syndrome, cutaneous T-cell lymphoma, Waldenström macroglobulinemia, diffuse large B-cell lymphoma (DLBCL), and multiple myeloma.

20. The method according to claim 18, wherein the solid tumor is breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colon cancer, kidney cancer, ovarian cancer, liver cancer, or prostate cancer.

21. The method according to any one of claims 17-20, the method further comprising administering to the patient a chemotherapeutic agent, an anti-angiogenic agent, a growth inhibitor, an immuno-oncology agent, a checkpoint inhibitor, and / or an anti-tumor composition.

22. A method of treating cancer in a subject in need thereof, the method comprising administering to the subject an effective amount of an isolated monoclonal antibody or an antigen-binding fragment thereof, the isolated monoclonal antibody or an antigen-binding fragment thereof having an HCVR, the HCVR comprising a CDR1 sequence having the amino acid sequence GFRFNTNA (SEQ ID NO:10), an HCVR CDR2 sequence having the amino acid sequence IRSKSNSYATYY (SEQ ID NO:11), and an HCVR CDR3 sequence having the amino acid sequence TRGSEQQYYYAMDY (SEQ ID NO:19); and an LCVR, the LCVR comprising an LCVR CDR1 sequence having the amino acid sequence QSLLHSNGNTY (SEQ ID NO:13), an LCVR CDR2 sequence having the amino acid sequence YRMSNR (SEQ ID NO:14), and an LCVR CDR3 sequence having the amino acid sequence MQHLEYPFT (SEQ ID NO:15), thereby treating the cancer of the subject.

23. The method according to claim 22, wherein the cancer is a solid tumor.

24. The method according to claim 23, wherein the solid tumor is selected from the group consisting of: breast cancer, head and neck cancer, lung cancer, melanoma, uveal melanoma, colorectal cancer, kidney cancer, urothelial cancer, ovarian cancer, endometrial cancer, uterine cancer, liver cancer, pancreatic cancer, cholangiocarcinoma, gastric cancer, gastroesophageal cancer, esophageal cancer, cervical cancer, squamous cell carcinoma, prostate cancer, or bladder cancer.

25. The method according to any one of claims 22-24, the method further comprising administering to the patient a chemotherapeutic agent, an anti-angiogenic agent, a growth inhibitor, an immuno-oncology agent, a checkpoint inhibitor, and / or an anti-tumor composition.

26. A polynucleotide that encodes a heavy chain or a light chain or an antigen-binding portion thereof as recited in any one of claims 1-16.

27. A vector that comprises the polynucleotide as recited in claim 26, wherein the vector is an expression vector selected from the group consisting of: mammalian expression vectors, yeast expression vectors, insect expression vectors, and bacterial expression vectors.

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