Anti-CCR8 antibody or antigen binding fragment thereof, and use thereof

Abstract

The present application relates to the technical field of biology, and discloses an anti-CCR8 antibody or an antigen binding fragment thereof, and a use thereof. The antibody can specifically bind to at least one epitope of CCR8, can also bind to Treg cells, has ADCC activity, and provides new possibilities for the treatment and / or prevention of cancer.

Description

Anti-CCR8 antibodies or their antigen-binding fragments and their applications

[0001] Cross-references to related applications

[0002] This disclosure claims priority to Chinese Patent Application No. 202411604626.3, filed on November 11, 2024, entitled "Anti-CCR8 antibody or antigen-binding fragment thereof and its application", the entire contents of which are incorporated herein by reference. Technical Field

[0003] This invention relates to the field of biotechnology, and in particular to an anti-CCR8 antibody or its antigen-binding fragment and its applications. Background Technology

[0004] The following statements are provided only as background information in relation to the present invention and do not necessarily constitute prior art.

[0005] Treg cells infiltrating tumor tissue have been identified as a crucial mechanism for tumor immune escape. In various mouse tumor models, targeting and killing tumor-infiltrating Treg cells with antibodies can significantly inhibit tumor growth; therefore, eliminating Treg cells has become an important strategy in anti-tumor drug development. CCR8 is a G protein-coupled receptor with a seven-transmembrane structure, belonging to the CC subfamily of chemokine receptors. Recent studies have shown that CCR8 is specifically expressed on tumor-infiltrating Treg cells, but not on peripheral blood Treg cells or other PBMCs, thus it is considered a specific marker of tumor-infiltrating Treg cells. Developing knockout antibodies targeting CCR8 holds promise for targeting and eliminating Treg cells in tumor tissue, thereby enhancing the efficacy of anti-tumor therapy.

[0006] In view of this, the present invention is hereby proposed. Summary of the Invention

[0007] One of the objectives of this invention is to provide an antibody or its antigen-binding fragment that can specifically bind to CCR8 and its application.

[0008] To solve the above-mentioned technical problems, the present invention adopts the following technical solution:

[0009] In a first aspect, an anti-CCR8 antibody or an antigen-binding fragment thereof is provided, wherein the antibody or the antigen-binding fragment thereof contains a heavy chain variable region and / or a light chain variable region, the complementarity-determining regions HCDR1, HCDR2 and HCDR3 of the heavy chain variable region, and the complementarity-determining regions LCDR1, LCDR2 and LCDR3 of the light chain variable region.

[0010] The HCDR1 comprises the amino acid residue DY, or an amino acid sequence as shown in either SEQ ID NO.29 or 30;

[0011] The HCDR2 comprises an amino acid sequence as shown in either SEQ ID NO. 41 or 42; or, comprises the following amino acid sequence NPN-X1-X2-D (SEQ ID NO. 195), where X1 is N or T and X2 is D or G;

[0012] The HCDR3 comprises an amino acid sequence as shown in either SEQ ID NO. 74 or 76;

[0013] The LCDR1 comprises the following amino acid sequence X3-H-X4-NGNTY (SEQ ID NO.196), where X3 is V or F and X4 is S or N;

[0014] The LCDR2 includes the amino acid residue KV;

[0015] The LCDR3 comprises the following amino acid sequence SQ-X5-X6-HVP-X7 (SEQ ID NO.196), where X5 is S or G, X6 is T or A, and X7 is F or W.

[0016] In a second aspect, an anti-CCR8 antibody or its antigen-binding fragment is also provided, wherein the antibody or its antigen-binding fragment contains complementarity-determining regions HCDR1, HCDR2 and HCDR3 of the heavy chain variable region, and / or complementarity-determining regions LCDR1, LCDR2 and LCDR3 of the light chain variable region.

[0017] The HCDR1, HCDR2, and HCDR3 comprise amino acid sequences consistent with the heavy chain variable regions shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17, or 20.

[0018] The LCDR1, LCDR2, and LCDR3 comprise amino acid sequences consistent with the light chain variable regions shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22, or 23.

[0019] Optionally, the above-mentioned HCDR1, HCDR2 and HCDR3, and LCDR1, LCDR2 and LCDR3 are determined according to the Kabat definition method, Chothia definition method, AbM definition method, Contact definition method or IMGT definition method.

[0020] Thirdly, an anti-CCR8 antibody or its antigen-binding fragment is also provided, said anti-CCR8 antibody or its antigen-binding fragment having an amino acid sequence as shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20 or a heavy chain variable region having at least 67% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20;

[0021] And / or, the anti-CCR8 antibody or its antigen-binding fragment has an amino acid sequence as shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23 or a light chain variable region having at least 93% sequence identity with an amino acid sequence as shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23.

[0022] Fourthly, an anti-CCR8 antibody or its antigen-binding fragment is provided, which competitively binds to CCR8 with the anti-CCR8 antibody of the first or second aspect, or the epitope of the anti-CCR8 antigen bound by the fragment is the same as the epitope of the anti-CCR8 antigen bound by the anti-CCR8 antibody of the first or second aspect.

[0023] Fifthly, a dual-epitope anti-CCR8 antibody or its antigen-binding fragment is also provided, which contains two antigen-binding portions that bind to different antigenic epitopes of CCR8, each antigen-binding portion containing at least one antigen-binding domain; each antigen-binding domain consists of a light chain variable region and a heavy chain variable region paired with the light chain variable region, which together bind to the same antigenic epitope.

[0024] The light chain variable region includes complementarity-determining regions LCDR1, LCDR2, and LCDR3. LCDR1, LCDR2, and LCDR3 are the same between light chain variable regions belonging to different antigen-binding regions.

[0025] In a sixth aspect, a biomaterial is also provided, said biomaterial comprising any one of the following (i) to (vi):

[0026] (i) A multispecific antibody containing the anti-CCR8 antibody or its antigen-binding fragment as shown above;

[0027] (ii) a chimeric antigen receptor, wherein the chimeric antigen receptor comprises an extracellular region, a transmembrane region and an intracellular region;

[0028] The extracellular region includes the aforementioned anti-CCR8 antibody or its antigen-binding fragment;

[0029] (iii) A nucleic acid molecule encoding the above-mentioned anti-CCR8 antibody or its antigen-binding fragment, (i) the multispecific antibody, recombinant protein and immunoconjugate, (i) the chimeric antigen receptor, or the above-mentioned dual epitope anti-CCR8 antibody or its antigen-binding fragment.

[0030] (iv) A vector carrying the nucleic acid molecule described in (iii);

[0031] (v) Recombinant cells, characterized in that they comprise the nucleic acid molecules described in (iii), or the vector described in (iv), or express the anti-CCR8 antibody or its antigen-binding fragment described above, or express the multispecific antibody, recombinant protein and immunoconjugate described in (i), express the chimeric antigen receptor described in (ii), or the dual epitope anti-CCR8 antibody or its antigen-binding fragment described above.

[0032] (vi) Engineered immune effector cells that express the chimeric antigen receptor described in (ii) or contain a nucleic acid molecule encoding the chimeric antigen receptor described in (ii).

[0033] In a seventh aspect, a pharmaceutical composition is also provided, the pharmaceutical composition comprising the anti-CCR8 antibody or its antigen-binding fragment as described in the first, second, third or fourth aspects, or the biepisode anti-CCR8 antibody or its antigen-binding fragment as described in the fifth aspect, or the biological material as described in the sixth aspect.

[0034] The eighth aspect also provides the use of the anti-CCR8 antibody or its antigen-binding fragment as described in the first, second, third, or fourth aspects, or the dual-epitope anti-CCR8 antibody or its antigen-binding fragment as described in the fifth aspect, or the biological material as described in the sixth aspect, or the pharmaceutical composition as described in the seventh aspect, in any of the following:

[0035] (I) Detect CCR8 or cells expressing CCR8;

[0036] (II) Prepare products for detecting CCR8 or cells expressing CCR8;

[0037] (III) Used for the treatment, prevention or relief of diseases, symptoms or conditions related to CCR8;

[0038] (IV) Prepare pharmaceutical compositions for treating, preventing or alleviating diseases, symptoms or conditions associated with CCR8;

[0039] (V) Used for the treatment, prevention or relief of tumors;

[0040] (VI) Prepare pharmaceutical compositions for the treatment, prevention or relief of tumors.

[0041] A ninth aspect also provides a method for producing the anti-CCR8 antibody or its antigen-binding fragment as described in the first, second, third, or fourth aspects, or the dual-epitope anti-CCR8 antibody or its antigen-binding fragment as described in the fifth aspect, the method comprising the following steps:

[0042] (a) Culture the recombinant cells described in aspect eight under conditions expressing the antibody or its antigen-binding fragment;

[0043] (b) Separation and purification of the antibody or its antigen-binding fragment obtained in step (a).

[0044] In a tenth aspect, a kit for detecting CCR8 is also provided, the kit comprising the anti-CCR8 antibody or its antigen-binding fragment as described in the first, second, third or fourth aspects, or the dual epitope anti-CCR8 antibody or its antigen-binding fragment as described in the fifth aspect.

[0045] Eleventhly, a method for treating, preventing, or alleviating a disease, symptom, or condition is also provided, characterized in that the method comprises: administering to a subject a therapeutically effective amount of the above-mentioned anti-CCR8 antibody or its antigen-binding fragment, or the above-mentioned dual-epitope anti-CCR8 antibody or its antigen-binding fragment, or the above-mentioned biological material, or the above-mentioned pharmaceutical composition.

[0046] Preferably, the disease, symptom, or condition includes tumors, inflammation, autoimmune diseases, infections, and immune rejection due to organ transplantation. Attached Figure Description

[0047] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0048] Figure 1 shows the flow cytometry analysis of the binding activity of hybridoma purified antibodies F022-102, F022-104, F022-119 and F022-125 to Raji-hCCR8 cells in Example 2.

[0049] Figure 2 shows the flow cytometry analysis of the binding activity of hybridoma purified antibodies F022-104 and F022-106 to Raji-hCCR8 cells in Example 2.

[0050] Figure 3 shows the flow cytometry analysis of the binding activity of hybridoma purified antibodies F022-102, F022-104, F022-119 and F022-125 to 293T-cynoCCR8 cells in Example 2.

[0051] Figure 4 shows the flow cytometry analysis of the binding activity of hybridoma purified antibodies F022-104 and F022-106 to 293T-cynoCCR8 cells in Example 2.

[0052] Figure 5 shows the flow cytometry analysis of the binding activity of recombinant antibodies R2281, R2282 and R2283 to Raji-hCCR8 cells in Example 4.

[0053] Figure 6 shows the flow cytometry analysis of the binding activity of recombinant antibodies R2281, R2292 and R2293 with Raji-hCCR8 cells in Example 4.

[0054] Figure 7 shows the flow cytometry analysis of the killing activity of NK-92 cells against Treg cells mediated by recombinant antibodies R2281, R2282, R2283, R2292 and R2293 in Example 4.

[0055] Figure 8 shows the flow cytometry analysis of the binding activity of antibodies R2337, R2339 and R2281 to Raji-hCCR8 cells in Example 6.

[0056] Figure 9 shows the flow cytometry analysis of the binding activity of antibodies R2349, R2350 and R2282 to Raji-hCCR8 cells in Example 6.

[0057] Figure 10 shows the flow cytometry analysis of the binding activity of antibodies R2359, R2360 and R2283 to Raji-hCCR8 cells in Example 6.

[0058] Figure 11 shows the flow cytometry analysis of the binding activity of humanized antibodies R2337, R2339, R2349, R2350, R2359 and R2360 to Treg cells in Example 6;

[0059] Figure 12 shows the flow cytometry analysis of the binding activity of humanized antibodies R2337, R2339, R2349, R2350, R2359 and R2360 to 293T-cynoCCR8 cells in Example 6.

[0060] Figure 13 shows the flow cytometry analysis of the killing activity of humanized antibodies R2337, R2339, R2349, R2350, R2359 and R2360 against Treg cells in Example 6.

[0061] Figure 14 shows the epitope competition between F022-102, F022-119, and F022-125 detected by flow cytometry in Example 7;

[0062] Figure 15 is a schematic diagram of antibodies binding to antigens with different structures;

[0063] Figure 16 is a schematic diagram of the structures of antibody dual epitope co-light chain antibodies R2701, R2702 and R2703 provided in Example 8;

[0064] Figure 17 shows the flow cytometry analysis of the binding activity of antibodies R2701, R2702, R2703, R2481 and R1418 to Treg cells in Example 9;

[0065] Figure 18 shows the flow cytometry analysis of the killing activity of PBMCs against Treg cells mediated by antibodies R2701, R2702, R2703, R2481 and R1418 in Example 9. Detailed Implementation

[0066] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0067] The articles “a / an,” “an,” and “described” in this invention include plural references unless the context clearly indicates otherwise. For example, “an antibody” refers to one or more antibodies.

[0068] In this document, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of indicated technical features. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one of that feature. In the description of this invention, "a plurality of" means at least two, such as two, three, etc., unless otherwise explicitly specified.

[0069] In this document, the term "CCR8" refers to CC chemokine receptor 8 (CCR8), a G protein-coupled receptor belonging to the CC chemokine receptor family. CCR8 is primarily expressed in skin immune cells, Treg cells in the tumor microenvironment, and a small number of effector T cells, monocytes, and NK cells, with lower expression in peripheral blood and normal tissues. Unless otherwise stated, the term "CCR8" includes human CCR8, its subtypes, and species homologs expressed by cells naturally expressed or transfected with the CCR8 gene. The term also includes artificially modified CCR8, including but not limited to mutated, truncated, or fused peptides or proteins with other domains, while retaining the necessary antigenic epitopes for antibody binding. The anti-CCR8 antibody or its antigen-binding fragment provided by this invention can specifically bind to CCR8.

[0070] In this paper, a "domain" refers to a specific region within a molecule that folds into a relatively independent structural unit in three-dimensional space, possessing specific functions and stability. A domain typically consists of one or more continuous portions of a polypeptide chain. These portions may not be continuous in the amino acid sequence of a protein, but they are close to each other in the three-dimensional structure, folding into a relatively independent structural unit. Generally, a domain is responsible for a single functional property and, in many cases, can be added to, removed from, or transferred to other molecules without losing the function of the rest of the molecule and / or the domain itself.

[0071] In this document, an antigen-binding domain refers to a domain in a molecule used to bind a target antigen. The antigen-binding domain can be a variable domain (variable region) of an antibody or a variant thereof, such as a nanobody (VHH), a heavy chain variable region (VH) or a light chain variable region (VL) of an antibody, or a variant thereof. The number of antigen-binding domains of an antigen or its antigen-binding fragment can be 2, 3, 4, or more. The number of antigen-binding domains binding different antigenic epitopes can be the same or different. The amino acid sequences between multiple antigen-binding domains binding the same antigenic epitope can be the same or different, for example, having different CDRs, or having the same CDR but different FRs. When the amino acid sequence of an antigen-binding domain is described herein, it indicates that at least one antigen-binding domain in the multispecific binding molecule has that amino acid sequence, without requiring all antigen-binding domains targeting the same antigenic epitope to have exactly the same amino acid sequence. In some alternative embodiments, the antigen-binding domains binding the same antigenic epitope in the multispecific binding molecule have the same amino acid sequence.

[0072] In this document, the term "antibody" includes any immunoglobulin capable of binding to a specific antigen. The term "antibody" is used in the broadest sense to encompass a wide range of antibody structures, including but not limited to monoclonal / polyclonal antibodies, monospecific / multispecific antibodies, full-length antibodies, and antigen-binding fragments, provided they exhibit the desired antigen-binding activity. Typically, a natural, complete antibody consists of two heavy (H) chains and two light (L) chains. Based on the presence or absence of α, δ, ε, γ, and μ heavy chains, antibodies can be classified into five main categories or isotypes: IgA, IgD, IgE, IgG, and IgM. Several major antibody categories can also be subdivided into subclasses, such as IgG1 (γ1 heavy chain), IgG2 (γ2 heavy chain), IgG3 (γ3 heavy chain), IgG4 (γ4 heavy chain), IgA1 (α1 heavy chain), or IgA2 (α2 heavy chain), etc. Each heavy chain consists of a variable region (VH) and first, second, third, and fourth (optionally) constant regions (CH1, CH2, CH3, and CH4, respectively). Light chains in mammals can be divided into λ or κ, and each light chain consists of a variable region (VL) and a constant region (CL).

[0073] The "variable region" or "variable domain" of an antibody refers to the domain at the amino terminus of the antibody's heavy or light chain that recognizes and binds to antigens. The composition and arrangement of the amino acids in this region determine the antibody's specificity in recognizing antigens. The heavy chain variable region can be called "VH," and the light chain variable region can be called "VL." The variable region contains antigen-binding sites. Both the heavy and light chain variable regions consist of three complementarity-determining regions (CDRs) (also known as hypervariable regions) connected by four framework regions (FRs). The extent of the backbone region and CDRs has been precisely defined, for example, in Kabat (see Sequences of Proteins of Immunological Interest, E. Kabat et al.) and Chothia. Any CDR determination method well-known in the art, including combinations of methods, can identify CDRs of variable domains. CDRs in each chain are held together closely by FRs to form variable regions. Typically, the variable regions VL / VH of the heavy and light chains can be obtained by linking the following numbered CDRs with FRs in the following combination: FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4.

[0074] The CDR boundaries of antibodies or their antigen-binding fragments described herein can be defined or identified according to the definitions of IMGT, Kabat, Chothia, AbM, and Contact. CDRs defined in other ways acceptable in the art are also within the scope of protection of this disclosure (Kaas, Q et al. IMGT unique numbering for immunoglobulin and T cell receptor constant domains and Ig superfamily C-like domains. Dev. Comp. Immunol. 29, 185-203, (2005); RM MacCallum et al., Antibody–antigen interactions: contact analysis and binding site topography J. Mol. Biol. (1996); Martin, ACR Protein sequence and structure analysis of antibody variable domains (Book chapter). In Antibody engineering lab manual Eds. Duebel, S. and Kontermann, R. (2001); Marie-Paule Lefranc et al. IMGT unique numbering for immunoglobulin and T cell receptor variable domains and Ig superfamily V-like). domains,Developmental and Comparative Immunology 27(2003)55–77).

[0075] In this document, the term "antigen-binding fragment" refers to a substance containing an antibody CDR that lacks some amino acids present in the full-length chain but still specifically binds to an antigen. Such fragments are biologically active because they bind to the target antigen and can compete with other antigen-binding molecules (including intact antibodies) for binding to a given epitope. Examples of antigen-binding fragments include, but are not limited to, Fab, Fab', F(ab')2, Fv fragments, disulfide-stabilized Fv fragments (dsFv), (dsFv)2, bispecific dsFv (dsFv-dsFv'), disulfide-stabilized bifunctional antibodies (ds diabody), single-chain antibody molecules (scFv), scFv dimers (bivalent bifunctional antibodies), and the smallest unit of antibody recognition. The aforementioned antigen-binding fragments can bind to the same antigen as the parent antibody.

[0076] In this paper, the term "Fab" in antibody refers to a portion of an antibody composed of a single light chain (including variable and constant regions) and a single heavy chain whose variable and first constant regions are linked by disulfide bonds. A "Fab' fragment" refers to a Fab fragment containing a portion of the hinge region. "F(ab')2" refers to a Fab' dimer. An "Fv fragment" is composed of the variable regions of a single light chain and / or a single heavy chain. A "single-chain Fv antibody" or "scFv" refers to an antibody fragment formed by the direct interconnection of light chain variable regions and heavy chain variable regions, or by linkage through peptide linker sequences. A "minimum recognition unit" refers to a structure containing only a single CDR within the variable region; although the minimum recognition unit has a small molecular weight and low affinity, it possesses the ability to bind to antigens.

[0077] In this paper, the term "full-length heavy chain" refers to a heavy chain in an antibody that meets the above definition and has a complete or substantially complete structure, i.e., a peptide chain containing the antibody heavy chain variable region (VH) and constant region (CH1, CH2, and CH3, and optionally CH4). "Substantially complete" can refer to a full-length heavy chain, or it can be a peptide chain of an antibody heavy chain that has been modified to retain the basic heavy chain function, such as a peptide chain with some amino acid residues deleted and / or mutated.

[0078] In this paper, the term "full-length light chain" refers to a light chain in an antibody that meets the above definition and has a complete or substantially complete structure, i.e., a peptide chain containing the variable region (VL) and constant region (CL) of the antibody heavy chain. "Substantially complete" can refer to a full-length light chain, or it can be a peptide chain in which the antibody light chain has been modified to retain the basic heavy chain function, such as a peptide chain with some amino acid residues deleted and / or mutated.

[0079] In this article, the term "amino acid" refers to naturally occurring amino acids and synthetic amino acids, as well as amino acid analogs and amino acid mimics that function in a similar manner to naturally occurring amino acids. Naturally occurring amino acids include amino acids encoded by the genetic code and their modified forms, such as hydroxyproline, γ-carboxyglutamic acid, and O-phosphoserine. Common natural amino acids include: alanine (Ala; A), arginine (Arg; R), asparagine (Asn; N), aspartic acid (Asp; D), cysteine ​​(Cys; C); glutamic acid (Glu; E), glutamine (Gln; Q), glycine (Gly; G); histidine (His; H), isoleucine (Ile; I), leucine (Leu; L), lysine (Lys; K), methionine (Met; M), phenylalanine (Phe; F), proline (Pro; P), serine (Ser; S), threonine (Thr; T), tryptophan (Trp; W), tyrosine (Tyr; Y), and valine (Val; V). Amino acid analogs are compounds that have the same basic chemical structure as naturally occurring amino acids (i.e., the α-carbon bound to hydrogen, carboxyl, amino, and R groups), such as homoserine, ortholeucine, methionine sulfoxide, and methionine methylsulfonium. Amino acid analogs typically have modified R groups (e.g., leucine) or modified peptide backbones, but retain the same basic chemical structure as naturally occurring amino acids. Amino acid mimics are chemical compounds that have a structure different from the general chemical structure of amino acids, but function in a manner similar to naturally occurring amino acids. In this document, the term "identity" percentage refers to the degree to which the amino acids of two polypeptides are identical at equivalent positions when two sequences are optimally aligned. The alignment of amino acid sequence identity percentages can be performed using various methods within the art, such as software well known in the art, such as BLAST, BLAST-2, ALIGN, MEGALIGN (DNASTAR), CLUSTALW, or CLUSTAL OMEGA. Those skilled in the art can determine appropriate parameters for the aligned sequences, including any algorithms required to achieve maximum alignment of the full length of the compared sequences.

[0080] In this document, the term "specific binding" or "specifically bound" refers to a non-random binding reaction between two molecules, such as the reaction between an antibody and an antigen. In some specific embodiments, for example, it is determined according to flow cytometry fluorescence sorting technology.

[0081] In this document, the term "anti-CCR8 antibody" refers to an antibody capable of specifically binding to CCR8. In some specific embodiments, the "anti-CCR8 antibody" specifically binds to CCR8 in humans and / or monkeys.

[0082] In this document, the term "affinity" or "affinity" refers to the strength of the non-covalent interaction between an immunoglobulin molecule (i.e., an antibody) or a fragment thereof and an antigen. The strength or affinity of an immune-binding interaction can be expressed as the equilibrium dissociation constant (KD) of the interaction, where a smaller KD value indicates higher affinity. KD can be determined using any conventional method known in the art, including but not limited to the Biacore assay, the Octet method, microthermophoresis, HPLC-MS, and flow cytometry fluorescence sorting techniques.

[0083] The binding of the antibody or its antigen-binding fragment provided by this invention to CCR8 can also be expressed as "half-maximum effective concentration (EC50)," which refers to the concentration of the drug or antibody that achieves 50% of the maximum biological effect after a specific exposure time. Generally, a smaller EC50 indicates better affinity, meaning that it can bind to the target protein at a lower concentration. The EC50 value can be determined using binding detection methods known in the art, such as direct or indirect binding detection methods (e.g., enzyme-linked immunosorbent assay (ELISA), flow cytometry, and other binding detection methods).

[0084] In this document, the term "epitaxy" refers to any antigenic determinant on an antigen that is bound to the complementary site of an antibody. An antigenic determinant is typically a specific chemical group with a defined composition and structure. Epitopes can be linear (i.e., continuous) or conformational (i.e., consisting of spaced-apart amino acid residues, discontinuous). Epitopes define the minimum binding site of an antibody and are therefore specific targets for antibodies or their antigen-binding fragments. Epitopes can be determined by any method well known in the art, such as conventional immunoassays, antibody competitive binding assays, or X-ray crystallography or related structural assays (e.g., nuclear magnetic resonance spectroscopy).

[0085] In this article, the term "multispecific antibody" refers to an antibody molecule that can bind to multiple (two or more) different antigenic epitopes of the same antigen or multiple (two or more) different antigens.

[0086] In this document, the term "nucleic acid molecule" refers to a polymeric form of nucleotides of any length, including ribonucleotides and / or deoxyribonucleotides. Examples of nucleic acid molecules include, but are not limited to, single-stranded, double-stranded, or multi-stranded DNA or RNA, genomic DNA, cDNA, DNA-RNA hybrids, or polymers containing purine and pyrimidine bases or other naturally occurring, chemically or biochemically modified, non-natural, or derived nucleotide bases. When a nucleic acid molecule encodes a protein or polypeptide, it may optionally encode either the sense or antisense strand. Nucleic acid molecules can be naturally occurring, synthetic, recombinant, or any combination thereof. The terms "nucleic acid molecule," "nucleic acid," and "polynucleotide" are used interchangeably.

[0087] In optional embodiments, the nucleic acid molecule is RNA or DNA, and can be single-stranded or double-stranded, preferably double-stranded DNA. When a nucleic acid molecule is placed in a functional relationship with another nucleic acid sequence, the nucleic acid molecule is "effectively linked." For example, if a promoter or enhancer affects the transcription of a coding sequence, then the promoter or enhancer is effectively linked to said coding sequence. DNA is preferably used when it is ligated into a vector.

[0088] In this document, the term "vector" refers to a delivery system that can operatively insert a genetic element (such as the aforementioned nucleic acid molecule) into and express that genetic element, for example, to produce a protein, RNA, or DNA encoded by the genetic element, or to replicate the genetic element. Vectors can be used to transform, transduce, or transfect host cells, enabling the expression of the genetic element they carry within the host cells. Examples of vectors include plasmids, phagemids, cosmids, artificial chromosomes such as yeast artificial chromosomes (YAC), bacterial artificial chromosomes (BAC), or P1-derived artificial chromosomes (PAC), bacteriophages such as λ phage or M13 phage, and animal viruses. Vectors may contain various elements that control expression, including promoter sequences, transcription initiation sequences, enhancer sequences, selection elements, and reporter genes. Additionally, vectors may contain a replication initiation site. Vectors may also include components that facilitate their entry into the cell, including but not limited to viral particles, liposomes, or protein coats. Vectors can be expression vectors or cloning vectors. In some embodiments, the vector (e.g., expression vector) provided by the present invention contains a nucleic acid sequence encoding an antibody or an antigen-binding fragment thereof as described in the present invention, at least one promoter operatively linked to the nucleic acid sequence (e.g., SV40, CMV, EF-1α), and at least one selection marker.

[0089] In this document, the terms “purified” or “isolated” associated with peptides or nucleic acids mean that the peptide or nucleic acid is not in its native medium or in its native form. Therefore, the term “isolated” includes peptides or nucleic acids removed from their original environment, such as if they are naturally occurring. Associated with nucleic acids, the terms “isolated” or “purified” indicate, for example, that the nucleic acid is not in its native genomic background (e.g., in a vector, as an expression cassette, linked to a promoter, or artificially introduced into a heterologous host cell).

[0090] In this document, the term "recombinant cell" refers to a cell into which exogenous polynucleotides and / or vectors can be introduced, or have already been introduced. The exogenous polynucleotides may or may not be integrated into the genome of the "recombinant cell." When the recombinant cell contains a vector, the vector can be introduced into mammalian cells to construct recombinant cells, which are then used to express the antibodies or antigen-binding fragments provided in this invention. The corresponding antibodies can be obtained by culturing the recombinant cells. Suitable mammalian cells include CHO cells or HEK cells, etc.

[0091] In this document, the term "pharmaceutical composition" refers to a form in which the biological activity of the active ingredient is permitted and which does not contain any additional ingredients that would have unacceptable toxicity to the subject to which the composition is administered. In some specific embodiments, the antibodies contained in or expressed in the pharmaceutical composition are capable of specifically targeting and binding to CCR8.

[0092] In this paper, the term “CCR8 binding” includes binding isolated CCR8 molecules and / or cells expressing CCR8.

[0093] In this article, "pharmaceutically acceptable carriers" can include any physiologically compatible solvent, dispersion medium, coating, antibacterial and antifungal agents, isotonic agents, and delayed absorption agents.

[0094] In this document, the terms "subject" or "patient" refer to a mammalian subject or patient. Exemplary subjects include, but are not limited to, humans, monkeys, dogs, cats, mice, rats, cattle, horses, camels, poultry, goats, and sheep. In some embodiments, the subject is a human. In some embodiments, the subject is a person suspected of having cancer.

[0095] In this document, the term "diagnosis" refers to the identification of a pathological state, disease, or condition, such as the identification of a CCR8-related disease, or the identification of a subject with a CCR8-related disease who may benefit from a specific treatment regimen. In some embodiments, a diagnosis includes identifying an abnormal level or activity of CCR8.

[0096] In this document, "CCR8-related" diseases, conditions, or states refer to changes in the expression or activity of CCR8 when a disease, condition, or state occurs, including increases or decreases. Changes in CCR8 expression or activity can be changes in the overall CCR8 expression or activity level in a patient, changes at the affected site, or changes in the expression or activity of CCR8 in other cells or tissues caused by the affected site. For example, changes in the expression or activity of CCR8 in cells or tissues at the affected site or in surrounding cells or tissues. Optionally, "CCR8-related" diseases, conditions, or states include tumors.

[0097] In this document, the term "effective dose" refers to a therapeutic dose sufficient to reduce or improve the severity and / or duration of a condition or one or more of its symptoms; prevent disease progression; cause disease remission; prevent recurrence, development, or progression of one or more disease-related symptoms; detect disease; or enhance or improve the preventive or therapeutic effect of another therapy (e.g., a prophylactic or therapeutic agent). The therapeutically effective dose of the antibody or its antigen-binding fragment described in this invention depends on a variety of factors known in the art, such as weight, age, medical history, current treatment, the subject's health status and potential for cross-infection, allergies, hypersensitivity, and side effects, as well as the route of administration and the extent of tumor development. Those skilled in the art (e.g., physicians or veterinarians) may proportionally reduce or increase the dose according to these or other conditions or requirements.

[0098] In a first aspect, an anti-CCR8 antibody or an antigen-binding fragment thereof is provided, wherein the antibody or the antigen-binding fragment thereof contains a heavy chain variable region and / or a heavy chain variable region, the complementarity-determining regions HCDR1, HCDR2 and HCDR3 of the heavy chain variable region, and the complementarity-determining regions LCDR1, LCDR2 and LCDR3 of the light chain variable region.

[0099] The HCDR1 comprises the amino acid residue DY, or an amino acid sequence as shown in either SEQ ID NO.29 or 30;

[0100] The HCDR2 comprises an amino acid sequence as shown in either SEQ ID NO. 41 or 42; or, comprises the following amino acid sequence NPN-X1-X2-D (SEQ ID NO. 195), where X1 is N or T and X2 is D or G;

[0101] The HCDR3 comprises an amino acid sequence as shown in either SEQ ID NO. 74 or 76;

[0102] The LCDR1 comprises the following amino acid sequence X3-H-X4-NGNTY (SEQ ID NO.196), where X3 is V or F and X4 is S or N;

[0103] The LCDR2 includes the amino acid residue KV;

[0104] The LCDR3 comprises the following amino acid sequence SQ-X5-X6-HVP-X7 (SEQ ID NO.196), where X5 is S or G, X6 is T or A, and X7 is F or W.

[0105] In an optional embodiment, the HCDR1 comprises the amino acid residue DY, or an amino acid sequence as shown in either SEQ ID NO. 29 or 30;

[0106] The HCDR2 comprises an amino acid sequence as shown in any of SEQ ID NO. 41, 42, 54, 55 and 56;

[0107] The HCDR3 comprises an amino acid sequence as shown in either SEQ ID NO. 74 or 76;

[0108] The LCDR1 comprises an amino acid sequence as shown in any of SEQ ID NO. 84, 87 and 92;

[0109] The LCDR2 includes the amino acid residue KV;

[0110] The LCDR3 includes an amino acid sequence as shown in any of SEQ ID NO. 100, 102, 104 and 106.

[0111] In an optional implementation, according to the IMGT definition:

[0112] The HCDR1 comprises an amino acid sequence as shown in any of SEQ ID NO. 28, 34 and 37;

[0113] The HCDR2 comprises an amino acid sequence as shown in any of SEQ ID NO. 43, 44, 51, 52 and 53;

[0114] The HCDR3 comprises an amino acid sequence as shown in any of SEQ ID NO. 71, 73, 79 and 81;

[0115] The LCDR1 comprises an amino acid sequence as shown in any of SEQ ID NO. 83, 86 and 88;

[0116] The LCDR2 includes the amino acid residue KV;

[0117] The LCDR3 includes an amino acid sequence as shown in any of SEQ ID NO. 101, 103, 105 and 107.

[0118] In an optional implementation, the complementary determination region of the heavy chain variable region includes any one of the following (a) to (e);

[0119] (a) HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.54, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.74;

[0120] (b) HCDR1 includes the amino acid sequence shown in SEQ ID NO.30, HCDR2 includes the amino acid sequence shown in SEQ ID NO.41, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.76;

[0121] (c)HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.56, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.74;

[0122] (d)HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.55, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.74;

[0123] (e)HCDR1 includes the amino acid sequence shown in SEQ ID NO.29, HCDR2 includes the amino acid sequence shown in SEQ ID NO.42, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.76.

[0124] In an optional implementation, the complementary determining region of the light chain variable region includes any one of the following (a') to (d'):

[0125] (a')LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.106;

[0126] (b')LCDR1 includes the amino acid sequence shown in SEQ ID NO.84, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.100;

[0127] (c')LCDR1 includes the amino acid sequence shown in SEQ ID NO.87, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.104;

[0128] (d')LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.102;

[0129] In an optional embodiment, the complementarity-determining region of the anti-CCR8 antibody or its antigen-binding fragment includes any one of the following (A) to (H);

[0130] (A)HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.56, HCDR3 includes the amino acid sequence shown in SEQ ID NO.74, LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.106.

[0131] (B)HCDR1 includes the amino acid sequence shown in SEQ ID NO.30, HCDR2 includes the amino acid sequence shown in SEQ ID NO.41, HCDR3 includes the amino acid sequence shown in SEQ ID NO.76, LCDR1 includes the amino acid sequence shown in SEQ ID NO.84, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.100.

[0132] (C)HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.56, HCDR3 includes the amino acid sequence shown in SEQ ID NO.74, LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.106.

[0133] (D)HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.55, HCDR3 includes the amino acid sequence shown in SEQ ID NO.74, LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.106.

[0134] (E)HCDR1 includes the amino acid sequence shown in SEQ ID NO.29, HCDR2 includes the amino acid sequence shown in SEQ ID NO.42, HCDR3 includes the amino acid sequence shown in SEQ ID NO.76, LCDR1 includes the amino acid sequence shown in SEQ ID NO.87, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.104.

[0135] (F)HCDR1 includes the amino acid sequence shown in SEQ ID NO.30, HCDR2 includes the amino acid sequence shown in SEQ ID NO.41, HCDR3 includes the amino acid sequence shown in SEQ ID NO.76, LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.102.

[0136] (G)HCDR1 includes the amino acid sequence shown in SEQ ID NO.30, HCDR2 includes the amino acid sequence shown in SEQ ID NO.41, HCDR3 includes the amino acid sequence shown in SEQ ID NO.76, LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.106.

[0137] (H)HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.54, HCDR3 includes the amino acid sequence shown in SEQ ID NO.74, LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.102.

[0138] In an optional implementation, according to the IMGT definition, the complementarity-determining region of the anti-CCR8 antibody or its antigen-binding fragment includes any one of the following (A) to (H);

[0139] (A')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.51, HCDR3 includes the amino acid sequence shown in SEQ ID NO.73, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.107.

[0140] (B')HCDR1 includes the amino acid sequence shown in SEQ ID NO. 34, HCDR2 includes the amino acid sequence shown in SEQ ID NO. 43, HCDR3 includes the amino acid sequence shown in SEQ ID NO. 81, LCDR1 includes the amino acid sequence shown in SEQ ID NO. 83, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO. 101; or,

[0141] (C')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.53, HCDR3 includes the amino acid sequence shown in SEQ ID NO.79, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.107.

[0142] (D')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.52, HCDR3 includes the amino acid sequence shown in SEQ ID NO.79, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.107.

[0143] (E')HCDR1 includes the amino acid sequence shown in SEQ ID NO.28, HCDR2 includes the amino acid sequence shown in SEQ ID NO.44, HCDR3 includes the amino acid sequence shown in SEQ ID NO.81, LCDR1 includes the amino acid sequence shown in SEQ ID NO.86, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.105.

[0144] (F')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.52, HCDR3 includes the amino acid sequence shown in SEQ ID NO.73, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.107.

[0145] (G')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.51, HCDR3 includes the amino acid sequence shown in SEQ ID NO.73, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.103.

[0146] (H')HCDR1 includes the amino acid sequence shown in SEQ ID NO.34, HCDR2 includes the amino acid sequence shown in SEQ ID NO.43, HCDR3 includes the amino acid sequence shown in SEQ ID NO.81, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.103.

[0147] (I')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.51, HCDR3 includes the amino acid sequence shown in SEQ ID NO.73, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.107.

[0148] (J')HCDR1 includes the amino acid sequence shown in SEQ ID NO.34, HCDR2 includes the amino acid sequence shown in SEQ ID NO.43, HCDR3 includes the amino acid sequence shown in SEQ ID NO.81, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.107.

[0149] In a second aspect, an anti-CCR8 antibody or its antigen-binding fragment is also provided, wherein the antibody or its antigen-binding fragment contains a heavy chain variable region and / or a light chain variable region, wherein the heavy chain variable region includes complementarity-determining regions HCDR1, HCDR2 and HCDR3, and the light chain variable region includes complementarity-determining regions LCDR1, LCDR2 and LCDR3.

[0150] The HCDR1, HCDR2, and HCDR3 comprise amino acid sequences consistent with the heavy chain variable regions shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17, or 20.

[0151] The LCDR1, LCDR2, and LCDR3 comprise amino acid sequences consistent with the light chain variable regions shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22, or 23.

[0152] The aforementioned HCDR1, HCDR2, and HCDR3, as well as LCDR1, LCDR2, and LCDR3, are determined according to the Kabat definition method, Chothia definition method, AbM definition method, Contact definition method, or IMGT definition method.

[0153] In an optional embodiment, the anti-CCR8 antibody or its antigen-binding fragments HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 can be independently selected from Tables 1 to 5.

[0154] Table 1 shows the exemplary antibody F022-102 (the amino acid sequence of the heavy chain variable region is shown in SEQ ID NO.1, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO.2).

[0155] Table 1

[0156] Table 2 shows the exemplary antibody F022-119 (the amino acid sequence of the heavy chain variable region is shown in SEQ ID NO.3, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO.4).

[0157] Table 2

[0158] Table 3 shows the exemplary antibody F022-104 (the amino acid sequence of the heavy chain variable region is shown in SEQ ID NO.5, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO.6).

[0159] Table 3

[0160] Table 4 shows the exemplary antibody F022-106 (the amino acid sequence of the heavy chain variable region is shown in SEQ ID NO.7, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO.8).

[0161] Table 4

[0162] Table 5 shows the exemplary antibody F022-125 (the amino acid sequence of the heavy chain variable region is shown in SEQ ID NO.9, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO.10).

[0163] Table 5

[0164] In an optional embodiment, the anti-CCR8 antibody or its antigen-binding fragment of the first or second aspect contains a heavy chain framework region and / or a light chain framework region, wherein the heavy chain framework region and / or light chain framework region are derived from at least one of a murine antibody, a human antibody, a primate antibody or a mutant thereof.

[0165] In an optional embodiment, the three CDRs of the anti-CCR8 antibody or its antigen-binding fragment are separated by flanking portions called frame regions (FR, where the light chain FR includes LFR1, LFR2, LFR3 and LFR4, and the heavy chain FR includes HFR1, HFR2, HFR3 and HFR4).

[0166] In an optional embodiment, the HFR1 comprises an amino acid sequence as shown in any one of SEQ ID NO. 108 to 113, or comprises a sequence having at least 72% (e.g., 72%, 75%, 78%, 80%, 82%, 85%, 88%, 90%, 93%, 95%, 97%, or 99%) sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 108 to 113.

[0167] In an optional embodiment, the HFR2 comprises an amino acid sequence as shown in any one of SEQ ID NO. 120 to 125, or comprises a sequence having at least 74% (e.g., 74%, 75%, 78%, 80%, 82%, 85%, 88%, 90%, 93%, 95%, 97%, or 99%) sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 120 to 125.

[0168] In an optional embodiment, the HFR3 comprises an amino acid sequence as shown in any one of SEQ ID NO. 127, 129, 131, 133, 135, 137, 139, 141, and 144, or comprises a sequence having at least 68% (e.g., 68%, 70%, 72%, 74%, 75%, 78%, 80%, 82%, 85%, 88%, 90%, 93%, 95%, 97%, or 99%) sequence identity with the amino acid sequence shown in any one of SEQ ID NO. 127, 129, 131, 133, 135, 137, 139, 141, and 144.

[0169] In an optional embodiment, the HFR4 comprises an amino acid sequence as shown in any one of SEQ ID NO. 145 and 146, or comprises a sequence having at least 90% (e.g., 90%, 93%, 95%, 97% or 99%) sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 145 and 146.

[0170] In an optional embodiment, the LFR1 comprises an amino acid sequence as shown in any one of SEQ ID NO. 148, 150, 152, 153, 155 and 157, or comprises a sequence having at least 86% (e.g. 86%, 88%, 90%, 93%, 95%, 97% or 99%) sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 148, 150, 152, 153, 155 and 157.

[0171] In an optional embodiment, the LFR2 comprises an amino acid sequence as shown in any one of SEQ ID NO. 161 and 162, or comprises a sequence having at least 88% (e.g., 88%, 90%, 93%, 95%, 97%, or 99%) sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 161 and 162.

[0172] In an optional embodiment, the LFR3 includes an amino acid sequence as shown in any one of SEQ ID NO. 163 to 166, or includes a sequence having at least 96% (e.g., 96%, 97%, 98%, or 99%) sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 163 to 166.

[0173] In an optional embodiment, the LFR4 comprises an amino acid sequence as shown in any one of SEQ ID NO. 175 to 180, or comprises a sequence having at least 67% (e.g., 67%, 68%, 70%, 72%, 74%, 75%, 78%, 80%, 82%, 85%, 88%, 90%, 93%, 95%, 97%, or 99%) sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 175 to 180.

[0174] In an optional implementation, the HFR1 comprises the HFR1 of the heavy chain variable region shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20.

[0175] In an optional implementation, the HFR2 comprises the HFR2 of the heavy chain variable region shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20.

[0176] In an optional implementation, the HFR3 comprises the HFR3 of the heavy chain variable region shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20.

[0177] In an optional implementation, the HFR4 comprises the HFR4 of the heavy chain variable region shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20.

[0178] In an optional implementation, the LFR1 comprises the LFR1 of the light chain variable region shown in any one of SEQ ID NO.2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23.

[0179] In an optional implementation, the LFR2 comprises the LFR2 of the light chain variable region shown in any one of SEQ ID NO.2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23.

[0180] In an optional implementation, the LFR3 comprises the LFR3 of the light chain variable region shown in any one of SEQ ID NO.2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23.

[0181] In an optional implementation, the LFR4 comprises the LFR4 of the light chain variable region shown in any one of SEQ ID NO.2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23.

[0182] In an optional embodiment, the HFR1 comprises the following amino acid sequence EVLLQQSGP-X8-LVKPGASV-X9-IPCKAS (SEQ ID NO.198), where X8 is E or V and X9 is Q or K; or comprises the amino acid sequence as described in SEQ ID NO.111.

[0183] In an optional embodiment, the HFR2 comprises an amino acid sequence as shown in any one of SEQ ID NO. 123, 125 and 121;

[0184] In an optional embodiment, the HFR3 comprises the following amino acid sequence: KATLTVDKSSX 10 -TAYMELRX 11 -LTSEDTAVYX 12 -C(SEQ ID NO.199), X 10 Is it T or S, X 11 Is it G or S, X 12 Is it F or Y; or,

[0185] Includes the following amino acid sequence RX 13 -TX14 -SRDTSNSX 15 -VFLKITSVDTX 16 -DSATYYC(SEQ ID NO.200), X 13 Is it I or V, X 14 Is it V or I, X 15 Is it Q or L, X 16 It is either E or A;

[0186] In an optional embodiment, the HFR4 comprises the amino acid sequence shown in SEQ ID NO.145;

[0187] In an optional embodiment, the LFR1 comprises the following amino acid sequence DVVX 17 -TQX 18 -PLSLPVSLGX 19 -X 20 -X 21 -SX 22 -SC(SEQ ID NO.201), X 17 Is it M or L, X 18 Is it T, F, or S, X 19 Is it D or N, X 20 Is it Q or H, X 21 Is it A or T, X 22 It is either I or F;

[0188] In an optional embodiment, the LFR2 comprises the amino acid sequence shown in SEQ ID NO.161;

[0189] In an optional embodiment, the LFR3 comprises an amino acid sequence as shown in either SEQ ID NO. 163 or 166;

[0190] In an optional embodiment, the LFR4 comprises the following amino acid sequence FGX 23 -GX 24 -X 25 -LEIX 26 (SEQ ID NO.202), X 23 Is it S or G, X 24 Is it T or S, X 25 Is it K or T, X 26 It is K or N;

[0191] In an optional embodiment, the antibody contains a heavy chain framework region and / or a light chain framework region, wherein the heavy chain framework region and / or light chain framework region are derived from a human antibody or a mutant thereof;

[0192] In an optional embodiment, the HFR1 comprises the amino acid sequence shown in SEQ ID NO.112;

[0193] In an optional embodiment, the HFR2 comprises an amino acid sequence as shown in either SEQ ID NO. 122 or 124;

[0194] In an optional embodiment, the HFR3 comprises the following amino acid sequence X 27 -VTMTTDX 28 -STSTAYMELRSLRSDDTAVYY-C(SEQ ID NO.203),X 27 Is it R or K, X 28 Is it K or T;

[0195] In an optional embodiment, the HFR4 comprises the amino acid sequence shown in SEQ ID NO.146;

[0196] In an optional embodiment, the LFR1 comprises the amino acid sequence shown in SEQ ID NO.155;

[0197] In an optional embodiment, the LFR2 comprises the amino acid sequence shown in SEQ ID NO.162;

[0198] In an optional embodiment, the LFR3 comprises an amino acid sequence as shown in either SEQ ID NO. 164 or 165;

[0199] In an optional embodiment, the LFR4 comprises an amino acid sequence as shown in SEQ ID NO.177.

[0200] In an optional implementation, HFR1, HFR2, HFR3, and HFR4 comprise the HFR1, HFR2, HFR3, and HFR4 of the heavy chain variable region shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17, and 20. These regions are defined as CDR regions according to the Kabat, Chothia, AbM, Contact, or IMGT definition schemes, and the FR regions of the corresponding definition schemes are obtained based on the structure of the heavy chain variable region.

[0201] Taking the IMGT definition as an example, the heavy chain framework region of the anti-CCR8 antibody or its antigen-binding fragment includes any one of the following (a) to (j):

[0202] (a) The HFR1 contains the amino acid sequence shown in SEQ ID NO. 109, the HFR2 contains the amino acid sequence shown in SEQ ID NO. 115, the HFR3 contains the amino acid sequence shown in SEQ ID NO. 126, and the HFR4 contains the amino acid sequence shown in SEQ ID NO. 145; or,

[0203] (b) HFR1 contains the amino acid sequence shown in SEQ ID NO. 111, HFR2 contains the amino acid sequence shown in SEQ ID NO. 119, HFR3 contains the amino acid sequence shown in SEQ ID NO. 128, and HFR4 contains the amino acid sequence shown in SEQ ID NO. 145; or,

[0204] (c) The HFR1 contains the amino acid sequence shown in SEQ ID NO. 110, the HFR2 contains the amino acid sequence shown in SEQ ID NO. 117, the HFR3 contains the amino acid sequence shown in SEQ ID NO. 130, and the HFR4 contains the amino acid sequence shown in SEQ ID NO. 145; or,

[0205] (d) The HFR1 contains the amino acid sequence shown in SEQ ID NO. 108, the HFR2 contains the amino acid sequence shown in SEQ ID NO. 115, the HFR3 contains the amino acid sequence shown in SEQ ID NO. 132, and the HFR4 contains the amino acid sequence shown in SEQ ID NO. 145; or,

[0206] (e) The HFR1 contains the amino acid sequence shown in SEQ ID NO. 111, the HFR2 contains the amino acid sequence shown in SEQ ID NO. 119, the HFR3 contains the amino acid sequence shown in SEQ ID NO. 134, and the HFR4 contains the amino acid sequence shown in SEQ ID NO. 145; or,

[0207] (f) The HFR1 contains the amino acid sequence shown in SEQ ID NO. 112, the HFR2 contains the amino acid sequence shown in SEQ ID NO. 114, the HFR3 contains the amino acid sequence shown in SEQ ID NO. 143, and the HFR4 contains the amino acid sequence shown in SEQ ID NO. 146; or,

[0208] (g) The HFR1 contains the amino acid sequence shown in SEQ ID NO. 112, the HFR2 contains the amino acid sequence shown in SEQ ID NO. 116, the HFR3 contains the amino acid sequence shown in SEQ ID NO. 136, and the HFR4 contains the amino acid sequence shown in SEQ ID NO. 146; or,

[0209] (h) The HFR1 contains the amino acid sequence shown in SEQ ID NO. 112, the HFR2 contains the amino acid sequence shown in SEQ ID NO. 116, the HFR3 contains the amino acid sequence shown in SEQ ID NO. 138, and the HFR4 contains the amino acid sequence shown in SEQ ID NO. 146; or,

[0210] (i) The HFR1 contains the amino acid sequence shown in SEQ ID NO.112, the HFR2 contains the amino acid sequence shown in SEQ ID NO.114, the HFR3 contains the amino acid sequence shown in SEQ ID NO.142, and the HFR4 contains the amino acid sequence shown in SEQ ID NO.146.

[0211] (j) HFR1 contains the amino acid sequence shown in SEQ ID NO.113, HFR2 contains the amino acid sequence shown in SEQ ID NO.118, HFR3 contains the amino acid sequence shown in SEQ ID NO.140, and HFR4 contains the amino acid sequence shown in SEQ ID NO.146.

[0212] In an optional implementation, LFR1, LFR2, LFR3, and LFR4 comprise the LFR1, LFR2, LFR3, and LFR4 of the light chain variable regions shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22, and 23. These regions are CDR regions determined according to the Kabat, Chothia, AbM, Contact, or IMGT definition schemes, and FR regions of the corresponding definition scheme are obtained based on the structure of the light chain variable region.

[0213] Taking the IMGT definition as an example, the light chain framework region of the anti-CCR8 antibody or its antigen-binding fragment includes any one of the following (a') to (l'):

[0214] (a') The LFR1 contains the amino acid sequence shown in SEQ ID NO.158, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.169, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.179;

[0215] (b') The LFR1 contains the amino acid sequence shown in SEQ ID NO.147, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.167, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.175;

[0216] (c') The LFR1 contains the amino acid sequence shown in SEQ ID NO.158, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.167, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.179;

[0217] (d') The LFR1 contains the amino acid sequence shown in SEQ ID NO.149, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.170, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.180;

[0218] (e') The LFR1 contains the amino acid sequence shown in SEQ ID NO.151, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.167, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.176;

[0219] (f') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.174, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177;

[0220] (g') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.171, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177;

[0221] (h') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.168, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177;

[0222] (i') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.172, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177;

[0223] (j') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.173, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177;

[0224] (k') The LFR1 contains the amino acid sequence shown in SEQ ID NO.154, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.168, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.178;

[0225] (l') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.174, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.178.

[0226] Thirdly, an anti-CCR8 antibody or its antigen-binding fragment is provided, said anti-CCR8 antibody or its antigen-binding fragment having an amino acid sequence as shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20 or a heavy chain variable region having at least 67% (e.g. 67%, 68%, 70%, 72%, 75%, 77%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity with the amino acid sequence shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20.

[0227] In an optional embodiment, the anti-CCR8 antibody or its antigen-binding fragment has an amino acid sequence as shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23, or a light chain variable region having at least 93% (e.g. 93%, 94%, 95%, 96%, 97%, 98% or 99%) sequence identity with the amino acid sequence shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23.

[0228] In an optional embodiment, the anti-CCR8 antibody or its antigen-binding fragment includes at least one of the following heavy chain variable regions and light chain variable regions:

[0229] (A) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.1 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.2;

[0230] (B) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.3 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.4;

[0231] (C) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.5 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.6;

[0232] (D) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.7 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.8;

[0233] (E) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.9 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.10;

[0234] (F) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.12;

[0235] (G) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.13;

[0236] (H) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.14 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.15;

[0237] (I) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.16 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.15;

[0238] (J) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.17 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.18;

[0239] (K) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.17 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.19;

[0240] (L) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.20 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.21;

[0241] (M) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.20 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.22;

[0242] (N) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.20 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.23;

[0243] (O) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.21;

[0244] (P) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.22;

[0245] (Q) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.23.

[0246] In an optional embodiment, the antigen-binding fragment includes one or more of F(ab')2, Fab', Fab, Fv, scFv, dsFv, and the smallest antibody recognition unit.

[0247] In an optional embodiment, the anti-CCR8 antibody or its antigen-binding fragment comprises part or all of a constant region sequence, wherein the constant region is derived from at least one of a murine antibody, a human antibody, a primate antibody, or a mutant thereof.

[0248] In an optional embodiment, the heavy chain constant region contains a sequence of part or all of the constant region of any one of IgG1, IgG2a, IgG2b, IgG3, IgG4, IgA, IgM, IgE or IgD.

[0249] In an optional embodiment, the constant region of the antibody light chain is a κ or λ chain.

[0250] In an optional implementation, the constant region source species include one or more combinations of mice, rats, guinea pigs, hamsters, rabbits, ferrets, cats, dogs, goats, sheep, cows, pigs, horses, monkeys, and humans.

[0251] In an optional embodiment, the heavy chain constant region contains a sequence of murine antibody IgG1 or a sequence of murine antibody IgG2a;

[0252] In an optional embodiment, the heavy chain constant region contains the sequence of human antibody IgG1 and has S239D, A330L and I332E mutations.

[0253] In an optional embodiment, the anti-CCR8 antibody or its antigen-binding fragment has one or more of the following properties (i) to (iv):

[0254] (i) The EC50 at a concentration not exceeding 10 nM (e.g., not exceeding 0.01 nM, 0.02 nM, 0.04 nM, 0.05 nM, 0.08 nM, 0.1 nM, 0.2 nM, 0.5 nM, 0.8 nM, 1 nM, 2 nM, 3 nM, 4 nM, 5 nM, 6 nM, 7 nM, 8 nM, 9 nM, or 10 nM) binds to human CCR8, and the binding is determined by flow cytometry fluorescence sorting, as described in Example 2.

[0255] In an optional embodiment, EC50 at a concentration not exceeding 10 nM (e.g., not exceeding 0.01 nM, 0.02 nM, 0.04 nM, 0.05 nM, 0.08 nM, 0.1 nM, 0.2 nM, 0.5 nM, 0.8 nM, 1 nM, 2 nM, 3 nM, 4 nM, 5 nM, 6 nM, 7 nM, 8 nM, 9 nM, or 10 nM) is bound to cells expressing human CCR8, and the binding is determined by flow cytometry fluorescence sorting.

[0256] (ii) Binding of EC50 to monkey CCR8 with no more than 3 nM (e.g., no more than 0.1 nM, 0.2 nM, 0.5 nM, 0.8 nM, 1 nM, 2 nM or 3 nM), the binding being determined by flow cytometry fluorescence sorting, as described in Example 2.

[0257] In an optional embodiment, EC50 is bound to monkey CCR8-expressing cells at a concentration not exceeding 3 nM (e.g., not exceeding 0.1 nM, 0.2 nM, 0.5 nM, 0.8 nM, 1 nM, 2 nM, or 3 nM), the binding being determined by flow cytometry fluorescence sorting.

[0258] (iii) EC50 is used to bind to Treg cells at concentrations not exceeding 5 nM (e.g., not exceeding 0.1 nM, 0.2 nM, 0.5 nM, 0.8 nM, 1 nM, 2 nM, 3 nM, 4 nM, or 5 nM), the binding being determined by flow cytometry fluorescence sorting. This determination method is described in Example 6.

[0259] (iv) Possesses ADCC activity (antibody-dependent cell-mediated cytotoxicity), the ADCC activity being detected by flow cytometry fluorescence sorting.

[0260] (v) Possesses ADCC activity, mediating NK cell killing of Treg cells with EC50 at no more than 10 nM (e.g., no more than 1 nM, 2 nM, 3 nM, 4 nM, 5 nM, 6 nM, 7 nM, 8 nM, 9 nM, or 10 nM), the binding being determined by flow cytometry fluorescence sorting. This determination method can be found in Example 4.

[0261] (vi) Possesses ADCC activity, mediating the killing of Treg cells by PBMC cells with EC50 at concentrations not exceeding 0.5 nM (e.g., not exceeding 0.01 nM, 0.02 nM, 0.04 nM, 0.05 nM, 0.08 nM, 0.1 nM, 0.2 nM, 0.3 nM, 0.4 nM, or 0.5 nM), the binding being determined by flow cytometry fluorescence sorting. This determination method can be found in Examples 6 or 9.

[0262] The anti-CCR8 antibodies or their antigen-binding fragments provided in the first, second, and third aspects can specifically bind to CCR8, and in some embodiments, can bind to the CCR8 protein expressed by mammalian cells (e.g., HEK cells, tumor cells, and Treg cells). The anti-CCR8 antibody or its antigen-binding fragment provided in the first aspect has ADCC activity, which can mediate the killing of Treg cells by immune cells such as PBMCs and NK cells. Because the ECD of CCR8 differs significantly between humans and monkeys, the discovery of human-monkey cross-binding antibodies for CCR8 is very difficult. However, human-monkey cross-binding antibodies have been successfully developed using the anti-CCR8 antibody or its antigen-binding fragment provided in the first aspect. For example, antibodies F022-102, F022-104, and F022-106 have shown binding activity to CCR8 in both humans and monkeys. Furthermore, one antibody (F022-102) was screened and found to be non-competitive with other antibodies, demonstrating the potential for dual epitope binding.

[0263] Fourthly, an anti-CCR8 antibody or its antigen-binding fragment is provided, which competitively binds to CCR8 with the anti-CCR8 antibodies of the first, second, and third aspects, or the epitope of the anti-CCR8 antigen bound by the anti-CCR8 antibody is the same as the epitope of the anti-CCR8 antigen bound by the anti-CCR8 antibodies of the first, second, and third aspects.

[0264] Fifthly, a dual-epitope anti-CCR8 antibody or its antigen-binding fragment is also provided, which contains two antigen-binding portions that bind to different CCR8 antigenic epitopes, each antigen-binding portion containing at least one antigen-binding domain; each antigen-binding domain consists of a light chain variable region and a heavy chain variable region paired with the light chain variable region, which together bind to the same antigenic epitope.

[0265] The light chain variable region includes complementarity-determining regions LCDR1, LCDR2, and LCDR3. LCDR1, LCDR2, and LCDR3 are the same between light chain variable regions belonging to different antigen-binding regions.

[0266] One or more antigen-binding domains belonging to the same antigen-binding moiety bind to the same antigenic epitope. Since the complementarity-determining regions (CDGs) of the light chain variable regions in a bi-epitope anti-CCR8 antibody or its antigen-binding fragment are identical—that is, the bi-epitope anti-CCR8 antibody or its antigen-binding fragment has a common light chain CDG—it is understandable that at least one of the HCDR1, HCDR2, and HCDR3 domains belonging to different antigen-binding moieties must differ to allow different antigen-binding moieties to bind different antigenic epitopes. However, differences in HCDR1, HCDR2, and HCDR3 domains between different heavy chain variable regions belonging to the same antigen-binding moiety are also permissible, as long as they can bind to the same antigenic epitope.

[0267] In an optional embodiment, the dual epitope anti-CCR8 antibody or its antigen-binding fragment contains at least two Fab fragments, such as, but not limited to, two, three, or four Fab fragments.

[0268] In an optional embodiment, at least two Fab fragments in the dual epitope anti-CCR8 antibody or its antigen-binding fragment are connected in series, that is, the C-terminus of one Fab fragment is connected to the N-terminus of an adjacent Fab fragment. The connection can be made by direct connection or by a linker.

[0269] In an optional implementation, at least two Fab fragments in tandem bind to different CCR8 antigenic epitopes.

[0270] In an optional embodiment, the dual-epitope anti-CCR8 antibody or its antigen-binding fragment comprises, in order from the N-terminus to the C-terminus, a first heavy chain peptide chain having the following structure: first VH-CH1-CH2-CH3; and a second heavy chain peptide chain having the following structure: second VH-CH1-CH2-CH3; the first VH and the second VH bind to different antigenic epitopes.

[0271] In an optional embodiment, the dual-epitope anti-CCR8 antibody or its antigen-binding fragment comprises, in order from the N-terminus to the C-terminus, a first heavy chain peptide chain having the following structure: first VH-CH1-second VH-CH1-CH2-CH3. The first VH and the second VH bind to different antigenic epitopes. Further optionally, the dual-epitope anti-CCR8 antibody or its antigen-binding fragment contains at least two first heavy chain peptide chains.

[0272] In an optional embodiment, the dual-epitope anti-CCR8 antibody or its antigen-binding fragment, in order from the N-terminus to the C-terminus, comprises a first heavy chain peptide chain having the structure: first VH-CH1-CH2-CH3; and a second heavy chain peptide chain having the structure: first VH-CH1-second VH-CH1-CH2-CH3. The first VH and the second VH bind to different antigenic epitopes.

[0273] It is understood that in any of the above embodiments, the first / second VH-CH1 portions of the first heavy chain peptide chain and the second heavy chain peptide chain both have paired VL-CL to form a Fab fragment.

[0274] In an optional embodiment, the heavy chain variable region includes complementary determinant regions HCDR1, HCDR2, and HCDR3, wherein the complementary determinant regions HCDR1, HCDR2, and HCDR3 of each heavy chain variable region are independently selected from HCDR1, HCDR2, and HCDR3 described in either the first or second aspect. The complementary determinant regions LCDR1, LCDR2, and LCDR3 of the light chain variable region are selected from LCDR1, LCDR2, and LCDR3 described in either the first or second aspect.

[0275] In an optional embodiment, the complementarity-determining regions HCDR1, HCDR2, and HCDR3 in one antigen-binding region of the dual epitope anti-CCR8 antibody or its antigen-binding fragment include amino acid sequences consistent with HCDR1, HCDR2, and HCDR3 of the heavy chain variable region shown in SEQ ID NO. 20.

[0276] The complementarity-determining regions HCDR1, HCDR2, and HCDR3 in another antigen-binding region include amino acid sequences consistent with those of the heavy chain variable regions HCDR1, HCDR2, and HCDR3 shown in SEQ ID NO. 11;

[0277] Furthermore, the complementary determinant regions LCDR1, LCDR2, and LCDR3 of the light chain variable region include amino acid sequences consistent with those of the light chain variable regions LCDR1, LCDR2, and LCDR3 shown in any one of SEQ ID NO. 21, 22, or 23.

[0278] The aforementioned HCDR1, HCDR2, and HCDR3, as well as LCDR1, LCDR2, and LCDR3, are determined according to the Kabat definition method, Chothia definition method, AbM definition method, Contact definition method, or IMGT definition method.

[0279] Table 6 shows a set of HCDR1, HCDR2, and HCDR3 for exemplary antibodies R2702, R2703, and R2701, with the amino acid sequences of the heavy chain variable region as shown in SEQ ID NO. 20 (Table 6) and SEQ ID NO. 11 (Table 7).

[0280] Table 6

[0281] Table 7

[0282] Table 8 shows the LCDR1, LCDR2, and LCDR3 of the exemplary antibody R2702, with the amino acid sequence of the light chain variable region as shown in SEQ ID NO.21 (Table 8).

[0283] Table 8

[0284] Table 9 shows the LCDR1, LCDR2, and LCDR3 of the exemplary antibody R2703, with the amino acid sequence of the light chain variable region as shown in SEQ ID NO. 22 (Table 9).

[0285] Table 9

[0286] Table 10 shows the LCDR1, LCDR2, and LCDR3 of the exemplary antibody R2701, with the amino acid sequence of the light chain variable region as shown in SEQ ID NO.23 (Table 10).

[0287] Table 10

[0288] In an optional embodiment, the complementarity-determining regions HCDR1, HCDR2, and HCDR3 of the heavy chain variable region in one antigen-binding portion of the dual epitope anti-CCR8 antibody or its antigen-binding fragment are selected from: HCDR1 comprising the amino acid sequence shown in SEQ ID NO.30, HCDR2 comprising the amino acid sequence shown in SEQ ID NO.41, and HCDR3 comprising the amino acid sequence shown in SEQ ID NO.76.

[0289] The complementarity-determining regions HCDR1, HCDR2, and HCDR3 of the heavy chain variable region in another antigen-binding moiety are selected from: HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.54, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.74;

[0290] The complementarity-determining regions LCDR1, LCDR2, and LCDR3 of the light chain variable region are selected from: LCDR1 including the amino acid sequence shown in SEQ ID NO.92, LCDR2 including the amino acid residue KV, and LCDR3 including the amino acid sequence shown in SEQ ID NO.106; or, LCDR1 including the amino acid sequence shown in SEQ ID NO.92, LCDR2 including the amino acid residue KV, and LCDR3 including the amino acid sequence shown in SEQ ID NO.102.

[0291] In an optional embodiment, according to the IMGT definition, the complementary determinant regions HCDR1, HCDR2 and HCDR3 of a set of heavy chain variable regions of the dual epitope anti-CCR8 antibody or its antigen-binding fragment are selected from: HCDR1 including the amino acid sequence shown in SEQ ID NO.34, HCDR2 including the amino acid sequence shown in SEQ ID NO.43, and HCDR3 including the amino acid sequence shown in SEQ ID NO.71;

[0292] The complementarity-determining regions of another set of heavy chain variable regions, HCDR1, HCDR2 and HCDR3, are selected from: HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.51, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.73;

[0293] The complementarity-determining regions LCDR1, LCDR2, and LCDR3 of the light chain variable region are selected from: LCDR1 comprising the amino acid sequence shown in SEQ ID NO.88, LCDR2 comprising the amino acid residue KV, and LCDR3 comprising the amino acid sequence shown in SEQ ID NO.103; or, LCDR1 comprising the amino acid sequence shown in SEQ ID NO.88, LCDR2 comprising the amino acid residue KV, and LCDR3 comprising the amino acid sequence shown in SEQ ID NO.107.

[0294] In an optional embodiment, the dual epitope anti-CCR8 antibody or its antigen-binding fragment contains a heavy chain framework region and / or a light chain framework region, wherein the heavy chain framework region and / or light chain framework region are derived from at least one of a murine antibody, a human antibody, a primate antibody, or a mutant thereof.

[0295] In an optional embodiment, the framework region of any heavy chain variable region of the dual-epitope anti-CCR8 antibody or its antigen-binding fragment includes HFR1, HFR2, HFR3, and HFR4, and the framework region of any light chain variable region includes LFR1, LFR2, LFR3, and LFR4, wherein,

[0296] HFR1, HFR2, HFR3, and HFR4 of each heavy chain variable region are independently selected from the HFR1, HFR2, HFR3, and HFR4 described in the anti-CCR8 antibody of the first aspect or its antigen-binding fragment; and / or, LFR1, LFR2, LFR3, and LFR4 of the light chain variable regions are selected from the LFR1, LFR2, LFR3, and LFR4 described in the anti-CCR8 antibody of the first or second aspect or its antigen-binding fragment.

[0297] In an optional implementation, the frame regions of the heavy chain variable region include HFR1, HFR2, HFR3, and HFR4, and the frame regions of the light chain variable region include LFR1, LFR2, LFR3, and LFR4, wherein...

[0298] The HFR1, HFR2, HFR3, and HFR4 of the framework region of the heavy chain variable region are selected from: HFR1 containing the amino acid sequence shown in SEQ ID NO. 113, HFR2 containing the amino acid sequence shown in SEQ ID NO. 120, HFR3 containing the amino acid sequence shown in SEQ ID NO. 141, and HFR4 containing the amino acid sequence shown in SEQ ID NO. 146; and / or the HFR1, HFR2, HFR3, and HFR4 of the framework region of the heavy chain variable region are selected from: HFR1 containing the amino acid sequence shown in SEQ ID NO. 112, HFR2 containing the amino acid sequence shown in SEQ ID NO. 122, HFR3 containing the amino acid sequence shown in SEQ ID NO. 144, and HFR4 containing the amino acid sequence shown in SEQ ID NO. 146;

[0299] And / or, the LFR1, LFR2, LFR3 and LFR4 of the framework region of the light chain variable region are selected from: the LFR1 containing the amino acid sequence shown in SEQ ID NO.153, the LFR2 containing the amino acid sequence shown in SEQ ID NO.162, the LFR3 containing the amino acid sequence shown in SEQ ID NO.165, and the LFR4 containing the amino acid sequence shown in SEQ ID NO.178.

[0300] In an optional implementation, according to the IMGT definition, HFR1, HFR2, HFR3, and HFR4 of the framework region of the heavy chain variable region are selected from: HFR1 containing the amino acid sequence shown in SEQ ID NO. 113, HFR2 containing the amino acid sequence shown in SEQ ID NO. 120, HFR3 containing the amino acid sequence shown in SEQ ID NO. 141, and HFR4 containing the amino acid sequence shown in SEQ ID NO. 146; and / or,

[0301] The HFR1, HFR2, HFR3 and HFR4 of the framework region of the heavy chain variable region are selected from: HFR1 containing the amino acid sequence shown in SEQ ID NO.112, HFR2 containing the amino acid sequence shown in SEQ ID NO.114, HFR3 containing the amino acid sequence shown in SEQ ID NO.143, and HFR4 containing the amino acid sequence shown in SEQ ID NO.146;

[0302] And / or, as defined by IMGT:

[0303] The LFR1, LFR2, LFR3, and LFR4 of the framework region of the light chain variable region are selected from: LFR1 containing the amino acid sequence shown in SEQ ID NO.154, LFR2 containing the amino acid sequence shown in SEQ ID NO.160, LFR3 containing the amino acid sequence shown in SEQ ID NO.168, and LFR4 containing the amino acid sequence shown in SEQ ID NO.178; or, LFR1 containing the amino acid sequence shown in SEQ ID NO.156, LFR2 containing the amino acid sequence shown in SEQ ID NO.160, LFR3 containing the amino acid sequence shown in SEQ ID NO.174, and LFR4 containing the amino acid sequence shown in SEQ ID NO.178.

[0304] In an optional embodiment, the amino acid sequence of the heavy chain variable region in one antigen-binding region is shown in SEQ ID NO. 20, and the amino acid sequence of the heavy chain variable region in the other antigen-binding region is shown in SEQ ID NO. 11; and / or, the amino acid sequences of the light chain variable regions are shown in SEQ ID NO. 21, 22 or 23.

[0305] In an optional embodiment, the dual epitope anti-CCR8 antibody or its antigen-binding fragment comprises part or all of a constant region sequence, wherein the constant region is derived from at least one of a murine antibody, a human antibody, a primate antibody, or a mutant thereof.

[0306] In an optional embodiment, the heavy chain constant region contains a sequence of part or all of the constant regions of any one of IgG1, IgG2a, IgG2b, IgG3, IgG4, IgA, IgM, IgE or IgD; the light chain constant region of the antibody is a κ or λ chain.

[0307] In an optional embodiment, the dual epitope anti-CCR8 antibody or its antigen-binding fragment contains two heavy chain constant regions, each containing a sequence of the constant region of IgG1.

[0308] In an optional implementation, the constant region of IgG1 is the constant region of human IgG1.

[0309] In an optional embodiment, the constant region of the human IgG1 contains S239D and I332E mutations.

[0310] In an optional embodiment, the amino acid sequences of the two heavy chain constant regions are shown in SEQ ID NO.181 and SEQ ID NO.182, respectively.

[0311] In an optional implementation, a light chain constant region is included.

[0312] In an optional embodiment, the amino acid sequence of the light chain constant region is shown in SEQ ID NO.183.

[0313] In an optional embodiment, the dual-epitope anti-CCR8 antibody or its antigen-binding fragment contains two full-length heavy chains and two full-length light chains, exhibiting a symmetrical structure.

[0314] In optional embodiments, the amino acid sequences of the heavy chain are shown in SEQ ID NO. 184 and 185, respectively, and the amino acid sequences of the two light chains are both shown in SEQ ID NO. 186; or, the amino acid sequences of the heavy chain are shown in SEQ ID NO. 184 and 185, respectively, and the amino acid sequences of the two light chains are both shown in SEQ ID NO. 187; or, the amino acid sequences of the heavy chain are shown in SEQ ID NO. 184 and 185, respectively, and the amino acid sequences of the two light chains are both shown in SEQ ID NO. 188.

[0315] In an optional embodiment, the Fc regions of the two heavy chains of the dual epitope anti-CCR8 antibody or its antigen-binding fragment contain Knob and Hole structures, respectively, and bind in a "Knob-into-Hole" manner.

[0316] In an optional embodiment, the complementarity-determining region includes a heavy chain variable region containing a Knob structure, where the amino acid sequence of the heavy chain variable region is consistent with HCDR1, HCDR2 and HCDR3 of the heavy chain variable region shown in SEQ ID NO.20 is located.

[0317] In an optional embodiment, the complementarity-determining region includes a light chain containing a Hole structure, where the heavy chain variable region contains an amino acid sequence consistent with HCDR1, HCDR2, and HCDR3 of the heavy chain variable region shown in SEQ ID NO. 11.

[0318] Conventional antibodies are bivalent, with one CCR8 antibody binding to two CCR8 molecules. However, for the biepisode antibody provided in the fifth aspect, since its two antigen-binding sites bind to two epitopes of the same antigen, one antibody binds only one CCR8 molecule. If a Treg cell has four CCR8 molecules, these four CCR8 molecules can be bound by two conventional CCR8 antibodies, but they can be bound by four biepisode CCR8 antibodies provided in the fifth aspect. Compared to standard monoclonal antibodies, the biepisode antibody provided in the fifth aspect can exert a stronger ADCC effect because more antibodies can be introduced onto the same cells. In the preferred embodiment, a biepisode co-light chain antibody with a common light chain was designed, which can be expressed simply and conveniently like a monoclonal antibody. Positive experimental results confirm that the biepisode antibody provided in the fifth aspect has better binding and killing activity.

[0319] In a sixth aspect, a biomaterial is also provided, said biomaterial comprising any one of the following (i) to (vi):

[0320] (i) A multispecific antibody containing the anti-CCR8 antibody or its antigen-binding fragment as shown above;

[0321] (ii) a chimeric antigen receptor, wherein the chimeric antigen receptor comprises an extracellular region, a transmembrane region and an intracellular region;

[0322] The extracellular region includes the aforementioned anti-CCR8 antibody or its antigen-binding fragment;

[0323] (iii) A nucleic acid molecule encoding the above-mentioned anti-CCR8 antibody or its antigen-binding fragment, (i) the multispecific antibody, recombinant protein and immunoconjugate, (i) the chimeric antigen receptor, or the above-mentioned dual epitope anti-CCR8 antibody or its antigen-binding fragment.

[0324] (iv) A vector carrying the nucleic acid molecule described in (iii);

[0325] (v) Recombinant cells, characterized in that they comprise the nucleic acid molecules described in (iii), or the vector described in (iv), or express the anti-CCR8 antibody or its antigen-binding fragment described above, or express the multispecific antibody, recombinant protein and immunoconjugate described in (i), express the chimeric antigen receptor described in (ii), or the dual epitope anti-CCR8 antibody or its antigen-binding fragment described above.

[0326] (vi) Engineered immune effector cells that express the chimeric antigen receptor described in (ii) or contain a nucleic acid molecule encoding the chimeric antigen receptor described in (ii).

[0327] In a seventh aspect, a pharmaceutical composition is also provided, the pharmaceutical composition comprising the anti-CCR8 antibody or its antigen-binding fragment as described in the first, second, third or fourth aspects, or the biepisode anti-CCR8 antibody or its antigen-binding fragment as described in the fifth aspect, or the biological material as described in the sixth aspect.

[0328] In an optional embodiment, the pharmaceutical composition further includes a pharmaceutically acceptable carrier and / or excipient. The acceptable carrier and pharmaceutically acceptable excipient may be any carrier and / or excipient known in the art and conventionally available. Examples of carriers include, but are not limited to, any physiologically compatible solvent, dispersion medium, coating, antibacterial and antifungal agents, isotonic agents, and delayed absorption agents; examples of excipients include, but are not limited to, fillers, disintegrants, preservatives, solubilizers, and emulsifiers.

[0329] In optional embodiments, the pharmaceutical composition may further include one or more active pharmaceutical ingredients having other therapeutic effects, including but not limited to one or more of the following: chemotherapeutic agents, anticancer drugs, radiotherapy agents, immunotherapy agents, antiangiogenic agents, targeted therapy agents, cell therapy agents, gene therapy agents, hormone therapy agents, antiviral agents, antibiotics, analgesics, antioxidants, metal chelators, and cytokines.

[0330] In an optional embodiment, the pharmaceutical composition is used to treat, prevent, or alleviate diseases, symptoms, or conditions associated with CCR8.

[0331] In an optional embodiment, the pharmaceutical composition is used to treat, prevent, or alleviate tumors.

[0332] The eighth aspect also provides the use of the anti-CCR8 antibody or its antigen-binding fragment as described in the first, second, third, or fourth aspects, or the dual-epitope anti-CCR8 antibody or its antigen-binding fragment as described in the fifth aspect, or the biological material as described in the sixth aspect, or the pharmaceutical composition as described in the seventh aspect, in any of the following:

[0333] (I) Detect CCR8 or cells expressing CCR8;

[0334] (II) Prepare products for detecting CCR8 or cells expressing CCR8;

[0335] (III) Used for the treatment, prevention or relief of diseases, symptoms or conditions related to CCR8;

[0336] (IV) Prepare pharmaceutical compositions for treating, preventing or alleviating diseases, symptoms or conditions associated with CCR8;

[0337] (V) Used for the treatment, prevention or relief of tumors;

[0338] (VI) Prepare pharmaceutical compositions for the treatment, prevention or relief of tumors.

[0339] In optional embodiments, the application of aspect (I) above can utilize the ability of anti-CCR8 antibodies or their antigen-binding fragments to specifically target and bind to CCR8, enabling the detection of CCR8 or cells expressing CCR8 based on immunoassay techniques. For example, when the antibody or its antigen-binding fragment chain is an immunoconjugate, such as one linked to a fluorescent group, a fluorescence detection device can be used to locate or detect CCR8 in real time. This can be used in techniques such as immunoblotting, immunoprecipitation, or flow cytometry, which involve utilizing the specific binding properties of CCR8 antigen and antibody to detect CCR8 or cells expressing CCR8. Correspondingly, in aspect (II) above, those skilled in the art can select the reagent composition in the kit according to the actual detection method, including but not limited to antagonists, anti-CCR8 antibodies or drug reference materials; protein purification columns; immunoglobulin affinity purification buffers; cell assay diluents; instructions or literature, etc. The kit can also be used to detect diseases, symptoms, or conditions related to CCR8.

[0340] In an optional implementation, the application of aspect (I) above is for non-diagnostic and non-therapeutic purposes.

[0341] In an optional embodiment, a method for treating, preventing, or alleviating a disease, symptom, or condition associated with CCR8 includes administering to a subject a therapeutically effective amount of the aforementioned anti-CCR8 antibody or its antigen-binding fragment and / or the aforementioned pharmaceutical composition.

[0342] In an optional implementation, the “CCR8-related” diseases, conditions or circumstances in any of the above aspects include: solid tumors or hematologic malignancies.

[0343] In an alternative implementation, a “CCR8-related” disease, symptom, or condition is characterized by high CCR8 expression.

[0344] In optional embodiments, the tumors involved in any of the above-mentioned diseases, conditions, or circumstances include, but are not limited to, one or more of the following: lymphoma, myeloma, brain tumor, squamous cell carcinoma of the head and neck, non-small cell lung cancer, nasopharyngeal carcinoma, esophageal cancer, gastric cancer, pancreatic cancer, gallbladder cancer, liver cancer, colorectal cancer, breast cancer, ovarian cancer, cervical cancer, endometrial cancer, uterine sarcoma, prostate cancer, bladder cancer, renal cell carcinoma, melanoma, pancreatic cancer, lung cancer, thyroid cancer, skin cancer, esophageal cancer, and cervical cancer.

[0345] In optional embodiments, the antibody or antigen-binding fragment or pharmaceutical composition as described in this invention may be administered at a therapeutically effective dose between about 0.001 mg / kg and about 1000 mg / kg. In some embodiments, the dosage may vary with the course of treatment. For example, in some embodiments, the initial dosage may be higher than subsequent dosages. In some embodiments, the dosage is adjusted during the course of treatment based on the subject's response.

[0346] In optional embodiments, the antibodies or antigen-binding fragments or pharmaceutical compositions disclosed in this invention can be administered alone or in combination with a therapeutically effective amount of a second therapeutic agent. For example, the antibodies or antigen-binding fragments or pharmaceutical compositions disclosed in this invention can be administered in combination with a second therapeutic agent (e.g., chemotherapeutic agents, anticancer drugs, radiotherapy agents, immunotherapy agents, antiangiogenic agents, targeted therapy agents, cell therapy agents, gene therapy agents, hormone therapy agents, antiviral agents, antibiotics, analgesics, antioxidants, metal chelators, or cytokines).

[0347] In optional embodiments, when the antibody or its antigen-binding fragment or pharmaceutical composition disclosed in this invention is used in combination with one or more other therapeutic agents, it may be administered simultaneously with the one or more other therapeutic agents. In some such embodiments, the antibody or its antigen-binding fragment and the other therapeutic agent may be administered simultaneously as part of the same composition. However, the antibody or its antigen-binding fragment or pharmaceutical composition "used in combination" with other therapeutic agents does not need to be administered simultaneously or in the same composition as the therapeutic agent. The meaning of "used in combination" in this invention also includes that an antibody or its antigen-binding fragment or pharmaceutical composition administered before or after another therapeutic agent is also considered to be "used in combination" with that therapeutic agent, i.e., the antibody or its antigen-binding fragment or pharmaceutical composition is administered with the second substance via a different route of administration.

[0348] A ninth aspect also provides a method for producing the anti-CCR8 antibody or its antigen-binding fragment as described in the first, second, third, or fourth aspects, or the dual-epitope anti-CCR8 antibody or its antigen-binding fragment as described in the fifth aspect, the method comprising the following steps:

[0349] (a) Culture the recombinant cells described in aspect eight under conditions expressing the antibody or its antigen-binding fragment;

[0350] (b) Separation and purification of the antibody or its antigen-binding fragment obtained in step (a).

[0351] In a tenth aspect, a kit for detecting CCR8 is also provided, the kit comprising the anti-CCR8 antibody or its antigen-binding fragment as described in the first, second, third or fourth aspects, or the dual epitope anti-CCR8 antibody or its antigen-binding fragment as described in the fifth aspect.

[0352] In an optional embodiment, the products in any of the above aspects, such as, but not limited to, anti-CCR8 antibodies or antigen-binding fragments thereof in reagents, kits, blocking agents, and pharmaceutical compositions, may be conjugated with at least one diagnostic agent and / or therapeutic agent to form an immunoconjugate.

[0353] The diagnostic agents include one or more of the following: radioactive contrast agents, paramagnetic ions, metals, fluorescent labels, chemiluminescent labels, ultrasound contrast agents, and photosensitizers;

[0354] The therapeutic agents include one or more of the following: cytotoxic agents, drugs, radionuclides, boron atoms, immunomodulators, anti-apoptotic agents, photosensitizing therapeutic agents, immunoconjugates, and oligonucleotides.

[0355] Radioactive nuclides include, but are not limited to, 110 In、 111 In、 177 Lu、 18 F, 52 Fe、 62 Cu、 64 Cu、 67 Cu、 67 Ga、 68 Ga、 86 Y、 90 Y、 89 Zr、 94 mTc, 94 Tc, 99 mTc, 120 I, 123 I, 124 I, 125 I, 131 I, 154-158 Gd, 32 P, 11 C 13 N、 15 O、 186 Re、 188 Re、 51 Mn, 52 mMn, 55 Co、 72 As、 75 Br、 76 Br、 82 mRb and 83 One or more of Sr.

[0356] Paramagnetic ions include, but are not limited to, one or more of chromium (III), manganese (II), iron (III), cobalt (II), nickel (II), copper (II), neodymium (III), samarium (III), ytterbium (III), gadolinium (III), vanadium (II), terbium (III), dysprosium (III), holmium (III) and erbium (III).

[0357] Fluorescent markers include, but are not limited to, Alexa 350, Alexa 405, Alexa 430, Alexa 488, Alexa 555, Alexa 647, AMCA, aminoacridine, BODIPY 630 / 650, BODIPY 650 / 665, BODIPY-FL, BODIPY-R6G, BODIPY-TMR, BODIPY-TRX, 5-carboxy-4′,5′-dichloro-2′,7′-dimethoxyfluorescein, 5-carboxy-2′,4′,5′,7′-tetrachlorofluorescein, 5-carboxyfluorescein, 5-carboxyrhodamine, 6-carboxyrhodamine, 6-carboxytetramethylrhodamine, and Cascade. Blue, Cy2, Cy3, Cy5, Cy7, 6-FAM, Dansyl chloride, Fluorescein, HEX, 6-JOE, NBD (7-nitrobenzo-2-oxa-1,3-diazole), Oregon Green 488, Oregon Green 500, Oregon Green 514, Pacific Blue, Phthalic acid, Terephthalic acid, Isophthalic acid, Cresol Violet, Cresol Blue Violet, Brilliant Cresol Blue, p-Aminobenzoic acid, Erythrosine, Phthalocyanine, Azocyanine, Anthocyanin, Xanthine, Succinyl fluorescein, Rare earth metal cavitation compounds, Tribispyridyldiamine europium, europium cavitation compounds or chelates, Diamine, Dianthocyanin, La Jolla Blue dye, Allococyanin B. Phycocyanin C. Phycocyanin R. Thiamine, Phycoerythrin, Phycoerythrin R. REG. Rhodamine Green, Rhodamine Isothiocyanate, Rhodamine Red, ROX, TAMRA, TET, TRIT (Tetramethylrhodamine isothiol), Tetramethylrhodamine, and Texas Red.

[0358] Oligonucleotides include, but are not limited to, one or more of shRNA, miRNA, and siRNA.

[0359] Drugs include, but are not limited to, methotrexate, fluorouracil, mercaptopurine, hydroxyurea, cytarabine, nitrogen mustard, cyclophosphamide, thiotepa, cisplatin, mitomycin, bleomycin, camptothecin, podophyllotoxin, actinomycin D, doxorubicin, daunorubicin, vincristine, paclitaxel, cephalotaxine alkaloids, and L-asparaginase.

[0360] Immunomodulators include, but are not limited to, one or more of the following: cytokines, chemokines, stem cell growth factors, lymphotoxins, hematopoietic factors, colony-stimulating factors (CSF), interferons, erythropoietin, thrombopoietin, tumor necrosis factor (TNF), interleukins (IL), granulocyte-colony-stimulating factor (G-CSF), granulocyte-macrophage-colony-stimulating factor (GM-CSF), and stem cell growth factors.

[0361] Radioactive nuclides include, but are not limited to, 111 In、 111 At、 177 Lu、 211 Bi、 212 Bi、 213 Bi、 211 At、 62 Cu、 67 Cu、 90 Y、 125 I, 131 I, 133 I, 32 P, 33 P, 47 Sc、 111 Ag、 67 Ga、 153 Sm、 161 Tb, 152 Dy、 166 Dy、 161 Ho、 166 Ho、 186 Re、 188 Re、 189 Re、 211 Pb, 212 Pb, 223 Ra、 225 Ac、 77 As、 89 Sr、 99 Mo、 105 Rh、 149 Pm, 169 Er、 194 Ir、 58 Co、 80 mBr、 99 mTc, 103 mRh、 109 Pt, 119 Sb, 189 mOs, 192 Ir、 219 Rn、 215 Po、 221 Fr、 255 Fm、 11 C 13 N、 15 O、 75 Br、 198 Au、 199 Au、 224 Ac、 77 Br、 113 mIn, 95 Ru、 97 Ru、103 Ru、 105 Ru、 107 Hg, 203 Hg, 121 mTe, 122 mTe, 125 mTe, 165 Tm、 167 Tm、 168 Tm、 197 Pt, 109 Pd, 142 Pr、 143 Pr、 161 Tb, 57 Co、 58 Co、 51 Cr 59 Fe、 75 Se、 201 Tl、 76 Br and 169 One or more of Yb.

[0362] Eleventhly, a method for treating, preventing, or alleviating a disease, symptom, or condition is also provided, characterized in that the method comprises: administering to a subject a therapeutically effective amount of the above-mentioned anti-CCR8 antibody or its antigen-binding fragment, or the above-mentioned dual-epitope anti-CCR8 antibody or its antigen-binding fragment, or the above-mentioned biological material, or the above-mentioned pharmaceutical composition.

[0363] Preferably, the disease, symptom, or condition includes tumors, inflammation, autoimmune diseases, infections, and immune rejection due to organ transplantation.

[0364] The present invention will be further illustrated below with specific embodiments. However, it should be understood that these embodiments are merely for the purpose of more detailed illustration and should not be construed as limiting the present invention in any way.

[0365] Example 1. Preparation of anti-CCR8 hybridoma antibody

[0366] Balb / c mice were used, and multiple alternating immunizations were performed using HSA-conjugated chemically synthesized human or monkey CCR8 amino acids from positions 1 to 35 (Eurogentec). Immunizations were administered every one to two weeks, for a total of 6–10 immunizations. After the fourth immunization, orbital blood was collected from the mice, and serum titers of anti-human CCR8 antibodies were detected by ELISA and flow cytometry. Mouse serum was serially diluted and co-incubated with Raji cell lines (human Burkitt's lymphoma cells) and Raji cell lines overexpressing human CCR8 (Raji-hCCR8). The cells were stained with PE-labeled goat anti-mouse Fc antibody (Biolegend), and serum titers were detected by flow cytometry. Simultaneously, a fusion protein (CCR8N-term-hFc) consisting of amino acids 1-35 from the N-terminus of CCR8 and human Fc was coated onto 96-well plates. Serially diluted serum was added and the plates were incubated. HRP-labeled goat anti-mouse Fc antibody (Sigma) was then added, and serum titers were detected by ELISA. Mice with the highest titers were identified based on flow cytometry and ELISA results. These mice were euthanized, and their spleen cells were collected and fused with the SP20 cell line.

[0367] Monoclonal cells from the supernatant that exhibited strong binding activity to Raji-hCCR8 but did not bind to the Raji cell line were selected for large-scale culture. The supernatant was collected after one week, and the antibodies in the supernatant were purified.

[0368] Example 2. Screening for binding activity of anti-CCR8 antibodies

[0369] Raji-hCCR8 and HEK-293T cells overexpressing monkey CCR8 (293T-cynoCCR8, HEK-293T is a human kidney epithelial cell line) were counted and their density adjusted to 2E6 / ml. 100 μL of cell suspension was added to each well of a 96-well plate, and the cells were centrifuged at 400×g for 5 minutes, discarding the supernatant. Hybridoma-purified antibody at a concentration of 200 nM was prepared and serially diluted 3-fold. Cells were resuspended in 100 μL of the diluted hybridoma-purified antibody and incubated at 4°C for 30 minutes. After washing twice with PBS, cells were resuspended in 100 μL of PE-labeled goat anti-mouse Fc antibody diluted 1:500 and incubated at 4°C for 30 minutes. After washing once with PBS, cells were resuspended in 100 μL of PBS per well, and the PE fluorescence intensity was analyzed by flow cytometry.

[0370] The binding detection results of hybridoma purified antibodies with Raji-hCCR8 are shown in Figures 1 and 2. A total of 5 hybridoma purified antibodies (F022-102, F022-104, F022-106, F022-119, F022-125) showed strong binding activity with Raji-hCCR8.

[0371] The binding detection results of hybridoma purified antibodies to 293T-cynoCCR8 are shown in Figures 3 and 4. A total of three hybridoma purified antibodies (F022-102, F022-104, and F022-106) showed strong binding activity with 293T-cynoCCR8.

[0372] Example 3. Preparation of recombinant anti-CCR8 antibody

[0373] The sequences of hybridoma purified antibodies F022-102, F022-104, F022-106, F022-119, and F022-125 were retrieved. Sequencing results showed that the light chain constant region of all hybridoma purified antibodies was mouse κ type, the heavy chain constant region of F022-102, F022-106, F022-119, and F022-125 was mIgG1 type, and the heavy chain constant region of F022-104 was mIgG2a type. The variable region sequences are shown in Table 1. The heavy chain constant region of the hybridoma-purified antibody was replaced with human hIgG1 Fc (DLE) (human IgG1 Fc was mutated to S239D / A330L / I332E), while other sequences remained unchanged. The antibody was cloned into the pCDNA3.4A expression plasmid, which was then transfected into the EXPI293 cell line. After 7 days, the supernatant was collected and purified by protein A to obtain recombinant antibodies. The recombinant antibodies were named R2281 (F022-104), R2282 (F022-106), R2283 (F022-102), R2292 (F022-125), and R2293 (F022-119). For example, the recombinant antibody "R2281" represents the antibody formed by replacing the heavy chain constant region of the hybridoma purified antibody "F022-104" with the heavy chain constant region of human IgG1 (DLE) and the light chain constant region with the light chain constant region of human κ, and so on.

[0374] Example 4. In vitro activity analysis of anti-CCR8 antibody

[0375] The binding activity of the recombinant antibodies to Raji-hCCR8 was detected by flow cytometry, using the same method as in Example 2. R2141 was used as a negative control antibody (the amino acid sequence of the heavy chain variable region of R2141 is shown in SEQ ID NO. 193, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO. 194), and the sequences are shown in Table 11. The experimental results are shown in Figures 5 and 6. All five recombinant antibodies showed good binding activity to Raji-hCCR8.

[0376] Antibody-mediated ADCC function was detected by expressing hCD16 molecules (NK-92-hCD16) in NK-92 cells (natural killer cells from human patients with malignant non-Hodgkin lymphoma). Treg cells were isolated and cultured as target cells that naturally express CCR8.

[0377] The method for isolating and culturing Treg cells is as follows: Peripheral blood mononuclear cells (PBMCs) were isolated using Lymphoprep (Axis-Shield) according to the manufacturer's instructions. Tregs were then isolated from the PBMCs using the EasySep Human CD4+CD127lowCD25+Regulatory T Cell Isolation Kit (Stem Cell) according to the manufacturer's instructions. The isolated Tregs were mixed with Dynabeads Human T-Activator CD3 / CD28 (Gibco) at a 1:1 ratio and cultured in complete culture medium (90% X-VIVO 15 + 10% FBS + 500 IU / ml IL-2, X-VIVO 15: Lonza, FBS: Gibco, IL-2: Tetracycline) for 5 to 8 days to activate and expand the Treg cells.

[0378] Treg cells isolated and cultured were stained with CFSE. Treg cells were added to 96-well plates at a density of 2E4 cells / well. NK-92-hCD16 cells were added to 96-well plates at a density of 1E5 cells / well, followed by the addition of the CCR8 recombinant antibody containing hIgG1 Fc(DLE), and incubation at 37°C for 6 hours. The 96-well plates were then removed and PI staining was performed. CSFE and PI fluorescence were detected by flow cytometry, and the proportion of PI- and CFSE-positive cells in the CFSE-positive cell population was calculated, representing the cell killing rate. The experimental results are shown in Figure 7; all recombinant antibodies exhibited excellent ADCC activity.

[0379] Example 5. Humanization design of anti-CCR8 antibody

[0380] Given that R2281, R2282, and R2283 exhibit better binding activity and stronger ADCC capabilities, these three antibodies were designed for humanization. Humanization was performed using a "CDR transplantation" method, where the mouse VH CDR was transplanted into the frame region of the human antibody VH, and the mouse VL CDR was transplanted into the frame region of the human antibody VL. The heavy chain humanization of R2281, R2282, and R2283 used the IGHV1-18*01 germline gene sequence as a template, while the light chain humanization used the IGKV2-40*01 germline gene sequence. After generating the corresponding humanized sequences, key amino acids were examined one by one, and significant differentially expressed residues were reversed to generate the final humanized sequences. The humanized antibodies for R2281 are R2337 and R2339; the humanized antibodies for R2282 are R2349 and R2350; and the humanized antibodies for R2283 are R2359 and R2360. Sequence information is shown in Table 11.

[0381] Example 6. Activity analysis of anti-CCR8 humanized antibody

[0382] The designed humanized antibodies were expressed and purified. The binding activity of the humanized antibodies with Raji-hCCR8, Treg cells expressing natural CCR8, and 293T-cynoCCR8 was detected. The binding activity detection method was the same as in Example 2. The binding results of the humanized antibodies with Raji-hCCR8 are shown in Figures 8, 9, and 10; the binding results with Treg cells are shown in Figure 11; and the binding results with 293T-cynoCCR8 are shown in Figure 12. Each humanized antibody showed good binding activity with Raji-hCCR8, Treg cells, and 293T-cynoCCR8, and the binding activity was similar to that of the humanized parent antibody.

[0383] PBMCs were used as effector cells to detect antibody-mediated ADCC function. Treg cells were stained with CFSE. Treg cells were added to 96-well plates at a density of 2E4 / well. PBMC cells were added to 96-well plates at a density of 1E6 / well, followed by the addition of anti-CCR8 humanized antibody, and incubation at 37°C for 6 hours. PI staining was then performed on the 96-well plates, and CSFE and PI fluorescence were detected by flow cytometry. The proportion of PI- and CFSE-positive cells in the CFSE-positive cell population was calculated, representing the cell killing rate. The experimental results are shown in Figure 13; all recombinant antibodies exhibited excellent ADCC activity.

[0384] Example 7: Binding Epitope Competition Analysis of Anti-CCR8 Antibody

[0385] Sequence analysis of F0222-102, F022-104, F022-106, F022-119, and F022-125 revealed that the light chains of the five antibodies were quite similar, the heavy chains of F022-102, F022-104, and F022-106 were quite similar, and the heavy chains of F022-119 and F022-125 were quite similar.

[0386] Epitope competition between F022-102 and F022-119, F022-125 was investigated. The experimental method was as follows: Raji-hCCR8 cells were counted and their density adjusted to 2E6 / ml. 100 μL / well of cell suspension was added to a 96-well plate, and the cells were centrifuged at 400×g for 5 minutes, discarding the supernatant. Antibody working solution (containing F022-119, F022-125, the competitive antibody F022-63, and the isotype control antibody R0860; the amino acid sequence of the heavy chain variable region of R0860 is shown in SEQ ID NO.189, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO.190) was prepared at a concentration of 40 μg / mL. The cells were resuspended in 100 μL of the antibody working solution and incubated at 4°C for 30 minutes. Recombinant antibody R2283 (F022-102) or isotype control R0861 (heavy chain variable region amino acid sequence of R0861 is shown in SEQ ID NO.191, light chain variable region amino acid sequence is shown in SEQ ID NO.192) was added to a final concentration of 0.4 μg / mL, and the cells were incubated at 4°C for 30 min. After washing twice with PBS, 100 μL of PE-labeled goat anti-human Fc antibody diluted 1:500 was added to resuspend the cells, and the cells were incubated at 4°C for 30 min. After washing once with PBS, 100 μL / well of PBS was added to resuspend the cells, and the PE fluorescence intensity was analyzed by flow cytometry.

[0387] The experimental results are shown in Figure 14. When a high concentration of F022-119 antibody is present in the environment, R2283 (F022-102) can still bind to Raji-hCCR8, and the binding level is not significantly different from that of the control group, proving that R2283 (F022-102) and F022-119 have different binding epitopes.

[0388] Example 8: Construction of anti-CCR8 dual-epitope colight chain antibody

[0389] Because the CCR8 expression level in tumor-infiltrating Tregs varies among different patients, some Tregs with low CCR8 expression have fewer CCR8 cells on their surface, resulting in fewer bound antibodies and insufficient Fc counts, which cannot mediate the killing of Tregs by effector cells such as NK cells. A conventional antibody has two identical antigen-binding sites, meaning one antibody can bind to two antigens. However, if the antibody is modified so that its two antigen-binding sites bind to different epitopes of the antigen (i.e., a biepitaxy antibody), one antibody can bind to only one antigen, thereby introducing more antibody with the same amount of antigen and improving the antibody's ADCC effect (Figure 15).

[0390] However, preparing biepisode antibodies from two different epitopes often requires complex design or preparation processes due to light chain mismatch issues. In the aforementioned examples, it was found that the light chains of F022-102 and F022-119 are extremely similar, with a total of 4 amino acids different in the CDR region. Therefore, we designed mutations in the light chain through sequence alignment analysis and cloned it into the pCDNA3.4A expression plasmid, aiming to find a common light chain that can simultaneously match the heavy chains of F022-102 and F022-119 (Figure 16).

[0391] The heavy chain of F022-119 was humanized by replacing Fc with the hIgG1 (S239D / I332E) subtype and modifying it with "Knob". The result was cloned into the pCDNA3.4A expression plasmid.

[0392] The Fc of the humanized antibody R2359 (F022-102) was replaced with the hIgG1 (S239D / I332E) isotype, and "Hole" modification was performed before cloning into the pCDNA3.4A expression plasmid. The two heavy chain plasmids and a common light chain plasmid were transfected into the EXPI293 cell line at a ratio of 1:1:2 for antibody expression and purification. The resulting antibodies were named R2701, R2702, and R2703. Sequences are shown in Table 11.

[0393] Example 9: Activity Analysis of Anti-CCR8 Dual Epitope Co-light Chain Antibody

[0394] First, the binding activity of R2701, R2702, and R2703 with Treg cells naturally expressing CCR8 was detected. As a control, this example also expressed CCR8 antibody 4A19 from WO2021194942A1 (PCT application No. WO2021US23430, invention name: ANTI-CCR8 ANTIBODIES FOR TREATING CANCER), named R1418. Simultaneously, the Fc of the parent antibody R2359 was replaced with the hIgG1 (S239D / I332E) subtype, named R2481. The binding activity detection method was the same as in Example 2. The binding results of the dual-episode co-light chain antibodies with Treg cells are shown in Figure 17. The binding plateau values ​​of each dual-episode antibody with Treg cells were significantly higher than those of the parent antibody and the control antibody.

[0395] Furthermore, using PBMCs as effector cells and Treg cells as target cells, the ADCC activity of the dual-episode co-light chain antibody was detected. The experimental method was as described in Example 6, and the experimental results are shown in Figure 18. The ADCC activity of the dual-episode co-light chain antibody was significantly better than that of the parent antibody and the control antibody.

[0396] The sequences involved in the specific implementation are shown in Table 11:

[0397] Table 11

[0398] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention. Industrial applicability

[0399] The anti-CCR8 antibody disclosed herein can specifically bind to at least one epitope of CCR8. This antibody can also bind to Treg cells and exhibits ADCC activity, providing new possibilities for cancer treatment and / or prevention. Therefore, the anti-CCR8 antibody disclosed herein possesses excellent practical performance and broad market application prospects.

Claims

Anti-CCR8 antibody or its antigen-binding fragment, characterized in that, The antibody or its antigen-binding fragment contains a heavy chain variable region and / or a light chain variable region, wherein the heavy chain variable region includes complementarity-determining regions HCDR1, HCDR2 and HCDR3, and the light chain variable region includes complementarity-determining regions LCDR1, LCDR2 and LCDR3. The HCDR1 comprises the amino acid residue DY, or an amino acid sequence as shown in either SEQ ID NO.29 or 30; The HCDR2 comprises an amino acid sequence as shown in either SEQ ID NO. 41 or 42; or, comprises the following amino acid sequence NPN-X1-X2-D (SEQ ID NO. 195), where X1 is N or T and X2 is D or G; The HCDR3 comprises an amino acid sequence as shown in either SEQ ID NO. 74 or 76; The LCDR1 comprises the following amino acid sequence X3-H-X4-NGNTY (SEQ ID NO.196), where X3 is V or F and X4 is S or N; The LCDR2 includes the amino acid residue KV; The LCDR3 comprises the following amino acid sequence SQ-X5-X6-HVP-X7 (SEQ ID NO.196), where X5 is S or G, X6 is T or A, and X7 is F or W. The anti-CCR8 antibody or its antigen-binding fragment according to claim 1 is characterized in that, According to the IMGT definition, the complementarity-determining region of the anti-CCR8 antibody or its antigen-binding fragment includes any one of the following (A) to (H); (A')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.51, HCDR3 includes the amino acid sequence shown in SEQ ID NO.73, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

107. (B')HCDR1 includes the amino acid sequence shown in SEQ ID NO.34, HCDR2 includes the amino acid sequence shown in SEQ ID NO.43, HCDR3 includes the amino acid sequence shown in SEQ ID NO.81, LCDR1 includes the amino acid sequence shown in SEQ ID NO.83, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

101. (C')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.53, HCDR3 includes the amino acid sequence shown in SEQ ID NO.79, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

107. (D')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.52, HCDR3 includes the amino acid sequence shown in SEQ ID NO.79, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

107. (E')HCDR1 includes the amino acid sequence shown in SEQ ID NO.28, HCDR2 includes the amino acid sequence shown in SEQ ID NO.44, HCDR3 includes the amino acid sequence shown in SEQ ID NO.81, LCDR1 includes the amino acid sequence shown in SEQ ID NO.86, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

105. (F')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.52, HCDR3 includes the amino acid sequence shown in SEQ ID NO.73, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

107. (G')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.51, HCDR3 includes the amino acid sequence shown in SEQ ID NO.73, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

103. (H')HCDR1 includes the amino acid sequence shown in SEQ ID NO.34, HCDR2 includes the amino acid sequence shown in SEQ ID NO.43, HCDR3 includes the amino acid sequence shown in SEQ ID NO.81, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

103. (I')HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.51, HCDR3 includes the amino acid sequence shown in SEQ ID NO.73, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

107. (J')HCDR1 includes the amino acid sequence shown in SEQ ID NO.34, HCDR2 includes the amino acid sequence shown in SEQ ID NO.43, HCDR3 includes the amino acid sequence shown in SEQ ID NO.81, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

107. The anti-CCR8 antibody or its antigen-binding fragment according to claim 1 or 2 is characterized in that, The antibody contains a heavy chain framework region and / or a light chain framework region, wherein the heavy chain framework region and / or the light chain framework region are derived from at least one of a mouse antibody, a human antibody, a primate antibody or a mutant thereof; Optionally, the variable region of the heavy chain includes frame regions HFR1, HFR2, HFR3, and HFR4, and the variable region of the light chain includes frame regions LFR1, LFR2, LFR3, and LFR4, wherein... The HFR1 comprises an amino acid sequence as shown in any one of SEQ ID NO. 108 to 113, or comprises a sequence having at least 72% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 108 to 113; And / or, the HFR2 comprises an amino acid sequence as shown in any one of SEQ ID NO. 120 to 125, or comprises a sequence having at least 74% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 120 to 125; And / or, the HFR3 comprises an amino acid sequence as shown in any one of SEQ ID NO. 127, 129, 131, 133, 135, 137, 139, 141 and 144, or comprises a sequence having at least 68% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 127, 129, 131, 133, 135, 137, 139, 141 and 144; And / or, the HFR4 comprises an amino acid sequence as shown in any one of SEQ ID NO. 145 and 146, or comprises a sequence having at least 90% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 145 and 146; And / or, the LFR1 comprises an amino acid sequence as shown in any one of SEQ ID NO. 148, 150, 152, 153, 155 and 157, or comprises a sequence having at least 86% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 148, 150, 152, 153, 155 and 157; And / or, the LFR2 comprises an amino acid sequence as shown in any one of SEQ ID NO. 161 and 162, or comprises a sequence having at least 88% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 161 and 162; And / or, the LFR3 comprises an amino acid sequence as shown in any one of SEQ ID NO. 163 to 166, or comprises a sequence having at least 96% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 163 to 166; And / or, the LFR4 comprises an amino acid sequence as shown in any one of SEQ ID NO. 175 to 180, or comprises a sequence having at least 67% sequence identity with an amino acid sequence shown in any one of SEQ ID NO. 175 to 180; Optionally, the HFR1 comprises the following amino acid sequence EVLLQQSGP-X8-LVKPGASV-X9-IPCKAS (SEQ ID NO.198), where X8 is E or V and X9 is Q or K; or comprises the amino acid sequence as described in SEQ ID NO.111; And / or, the HFR2 comprises an amino acid sequence as shown in any one of SEQ ID NO. 123, 125 and 121; And / or, the HFR3 comprises the following amino acid sequence: KATLTVDKSSX 10 -TAYMELRX 11 -LTSEDTAVYX 12 -C(SEQ ID NO.199), X 10 Is it T or S, X 11 Is it G or S, X 12 Is it F or Y; or, Includes the following amino acid sequence RX 13 -TX 14 -SRDTSNSX 15 -VFLKITSVDTX 16 -DSATYYC(SEQ ID NO.200), X 13 Is it I or V, X 14 Is it V or I, X 15 Is it Q or L, X 16 It is either E or A; And / or, the HFR4 comprises the amino acid sequence shown in SEQ ID NO.145; And / or, the LFR1 comprises the following amino acid sequence DVVX 17 -TQX 18 -PLSLPVSLGX 19 -X 20 -X 21 -SX 22 -SC(SEQ ID NO.201), X 17 Is it M or L, X 18 Is it T, F, or S, X 19 Is it D or N, X 20 Is it Q or H, X 21 Is it A or T, X 22 It is either I or F; And / or, the LFR2 comprises the amino acid sequence shown in SEQ ID NO.161; And / or, the LFR3 comprises an amino acid sequence as shown in either SEQ ID NO. 163 or 166; And / or, the LFR4 comprises the following amino acid sequence FGX 23 -GX 24 -X 25 -LEIX 26 (SEQ ID NO.202), X 23 Is it S or G, X 24 Is it T or S, X 25 Is it K or T, X 26 It is K or N; Optionally, the antibody contains a heavy chain framework region and / or a light chain framework region, wherein the heavy chain framework region and / or light chain framework region are derived from a human antibody or a mutant thereof; And / or, the HFR1 comprises the amino acid sequence shown in SEQ ID NO.112; And / or, the HFR2 comprises an amino acid sequence as shown in either SEQ ID NO. 122 or 124; And / or, the HFR3 comprises the following amino acid sequence X 27 -VTMTTDX 28 -STSTAYMELRSLRSDDTAVYY-C(SEQ ID NO.203),X 27 Is it R or K, X 28 Is it K or T; And / or, the HFR4 comprises the amino acid sequence shown in SEQ ID NO.146; And / or, the LFR1 comprises the amino acid sequence shown in SEQ ID NO.155; And / or, the LFR2 comprises the amino acid sequence shown in SEQ ID NO.162; And / or, the LFR3 comprises an amino acid sequence as shown in either SEQ ID NO. 164 or 165; And / or, the LFR4 comprises the amino acid sequence shown in SEQ ID NO.177; Optionally, according to the IMGT definition, the heavy chain framework region of the anti-CCR8 antibody or its antigen-binding fragment includes any one of the following (a) to (j): (a) The HFR1 contains the amino acid sequence shown in SEQ ID NO.109, the HFR2 contains the amino acid sequence shown in SEQ ID NO.115, the HFR3 contains the amino acid sequence shown in SEQ ID NO.126, and the HFR4 contains the amino acid sequence shown in SEQ ID NO.145; (b) HFR1 contains the amino acid sequence shown in SEQ ID NO.111, HFR2 contains the amino acid sequence shown in SEQ ID NO.119, HFR3 contains the amino acid sequence shown in SEQ ID NO.128, and HFR4 contains the amino acid sequence shown in SEQ ID NO.145; (c) The HFR1 contains the amino acid sequence shown in SEQ ID NO.110, the HFR2 contains the amino acid sequence shown in SEQ ID NO.117, the HFR3 contains the amino acid sequence shown in SEQ ID NO.130, and the HFR4 contains the amino acid sequence shown in SEQ ID NO.145; (d) The HFR1 contains the amino acid sequence shown in SEQ ID NO.108, the HFR2 contains the amino acid sequence shown in SEQ ID NO.115, the HFR3 contains the amino acid sequence shown in SEQ ID NO.132, and the HFR4 contains the amino acid sequence shown in SEQ ID NO.145; (e) The HFR1 contains the amino acid sequence shown in SEQ ID NO.111, the HFR2 contains the amino acid sequence shown in SEQ ID NO.119, the HFR3 contains the amino acid sequence shown in SEQ ID NO.134, and the HFR4 contains the amino acid sequence shown in SEQ ID NO.145; (f) The HFR1 contains the amino acid sequence shown in SEQ ID NO. 112, the HFR2 contains the amino acid sequence shown in SEQ ID NO. 114, the HFR3 contains the amino acid sequence shown in SEQ ID NO. 143, and the HFR4 contains the amino acid sequence shown in SEQ ID NO. 146; or, (g) The HFR1 contains the amino acid sequence shown in SEQ ID NO.112, the HFR2 contains the amino acid sequence shown in SEQ ID NO.116, the HFR3 contains the amino acid sequence shown in SEQ ID NO.136, and the HFR4 contains the amino acid sequence shown in SEQ ID NO.146; (h) The HFR1 contains the amino acid sequence shown in SEQ ID NO.112, the HFR2 contains the amino acid sequence shown in SEQ ID NO.116, the HFR3 contains the amino acid sequence shown in SEQ ID NO.138, and the HFR4 contains the amino acid sequence shown in SEQ ID NO.146; (i) HFR1 contains the amino acid sequence shown in SEQ ID NO.112, HFR2 contains the amino acid sequence shown in SEQ ID NO.114, HFR3 contains the amino acid sequence shown in SEQ ID NO.142, and HFR4 contains the amino acid sequence shown in SEQ ID NO.146; (j) HFR1 contains the amino acid sequence shown in SEQ ID NO.113, HFR2 contains the amino acid sequence shown in SEQ ID NO.118, HFR3 contains the amino acid sequence shown in SEQ ID NO.140, and HFR4 contains the amino acid sequence shown in SEQ ID NO.146; And / or, according to the IMGT definition, the light chain framework region of the anti-CCR8 antibody or its antigen-binding fragment includes any one of the following (a') to (l'): (a') The LFR1 contains the amino acid sequence shown in SEQ ID NO.158, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.169, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.179; (b') The LFR1 contains the amino acid sequence shown in SEQ ID NO.147, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.167, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.175; (c') The LFR1 contains the amino acid sequence shown in SEQ ID NO.158, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.167, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.179; (d') The LFR1 contains the amino acid sequence shown in SEQ ID NO.149, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.170, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.180; (e') The LFR1 contains the amino acid sequence shown in SEQ ID NO.151, the LFR2 contains the amino acid sequence shown in SEQ ID NO.159, the LFR3 contains the amino acid sequence shown in SEQ ID NO.167, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.176; (f') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.174, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177; (g') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.171, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177; (h') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.168, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177; (i') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.172, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177; (j') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.173, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.177; (k') The LFR1 contains the amino acid sequence shown in SEQ ID NO.154, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.168, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.178; (l') The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.174, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.

178. Anti-CCR8 antibody or its antigen-binding fragment, characterized in that, The antibody or its antigen-binding fragment contains a heavy chain variable region and / or a light chain variable region, wherein the heavy chain variable region includes complementarity-determining regions HCDR1, HCDR2 and HCDR3, and the light chain variable region includes complementarity-determining regions LCDR1, LCDR2 and LCDR3. The HCDR1, HCDR2, and HCDR3 comprise amino acid sequences consistent with the heavy chain variable regions shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17, or 20. The LCDR1, LCDR2, and LCDR3 comprise amino acid sequences consistent with the light chain variable regions shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22, or 23. Optionally, the variable regions HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, or LCDR3 are defined by any one or a combination of multiple definition systems, such as Kabat, Chothia, IMGT, ABM, or Contact. Anti-CCR8 antibody or its antigen-binding fragment, characterized in that, The anti-CCR8 antibody or its antigen-binding fragment has an amino acid sequence as shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20 or a heavy chain variable region having at least 67% sequence identity with an amino acid sequence as shown in any one of SEQ ID NO. 1, 3, 5, 7, 9, 11, 14, 16, 17 and 20; And / or, the anti-CCR8 antibody or its antigen-binding fragment has an amino acid sequence as shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23 or a light chain variable region having at least 93% sequence identity with an amino acid sequence as shown in any one of SEQ ID NO. 2, 4, 6, 8, 10, 12, 13, 15, 18, 19, 21, 22 and 23. The anti-CCR8 antibody or its antigen-binding fragment according to any one of claims 1 to 5 is characterized in that, The anti-CCR8 antibody or its antigen-binding fragment includes at least one of the following heavy chain variable regions and light chain variable regions: (A) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.1 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.2; (B) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.3 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.4; (C) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.5 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.6; (D) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.7 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.8; (E) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.9 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.10; (F) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.12; (G) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.13; (H) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.14 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.15; (I) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.16 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.15; (J) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.17 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.18; (K) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.17 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.19; (L) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.20 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.21; (M) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.20 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.22; (N) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.20 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.23; (O) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.21; (P) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.22; (Q) The heavy chain variable region of the amino acid sequence shown in SEQ ID NO.11 and the light chain variable region of the amino acid sequence shown in SEQ ID NO.

23. The anti-CCR8 antibody or its antigen-binding fragment according to any one of claims 1 to 6 is characterized in that, The anti-CCR8 antibody or its antigen-binding fragment includes part or all of a constant region sequence, wherein the constant region is derived from at least one of a murine antibody, a human antibody, a primate antibody, or a mutant thereof; Optionally, the heavy chain constant region contains a sequence of part or all of the constant regions of any one of IgG1, IgG2a, IgG2b, IgG3, IgG4, IgA, IgM, IgE or IgD; the light chain constant region of the antibody is a κ or λ chain. The anti-CCR8 antibody or its antigen-binding fragment according to any one of claims 1 to 7 is characterized in that, The antigen-binding fragments include one or more of F(ab')2, Fab', Fab, Fv, scFv, dsFv, and the smallest antibody recognition unit. The anti-CCR8 antibody or its antigen-binding fragment according to any one of claims 1 to 8 is characterized in that, The anti-CCR8 antibody or its antigen-binding fragment has one or more of the following properties (i) to (vi): (i) Binding of human CCR8 with EC50 at a concentration not exceeding 10 nM, the binding being determined by flow cytometry fluorescence sorting. (ii) Binding of monkey CCR8 with EC50 of no more than 3 nM, the binding being determined by flow cytometry fluorescence sorting. (iii) Treg cells were bound with EC50 at a concentration not exceeding 5 nM, and the binding was determined by flow cytometry fluorescence sorting. (iv) It has ADCC activity; (v) It has ADCC activity and mediates NK cell killing of Treg cells with EC50 of no more than 10 nM, the binding of which is determined by flow cytometry fluorescence sorting. (vi) It has ADCC activity and mediates the killing of Treg cells by PBMC cells with EC50 of no more than 0.5 nM, the binding of which is determined by flow cytometry fluorescence sorting. An anti-CCR8 antibody or its antigen-binding fragment, characterized in that, It competitively binds to CCR8 with the anti-CCR8 antibody according to any one of claims 1 to 9, or its epitope binding to the CCR8 antigen is the same as the epitope binding to the CCR8 antigen of the anti-CCR8 antibody according to any one of claims 1 to 9. A dual-epitope anti-CCR8 antibody or its antigen-binding fragment, characterized in that, The antigen-binding moiety contains two antigen-binding regions that bind to different antigenic epitopes of CCR8, and each antigen-binding moiety contains at least one antigen-binding domain; each antigen-binding domain consists of a light chain variable region and a heavy chain variable region paired with the light chain variable region, which together bind the same antigenic epitope. The light chain variable region includes complementarity-determining regions LCDR1, LCDR2, and LCDR3. The LCDR1, LCDR2, and LCDR3 are the same between light chain variable regions belonging to different antigen-binding regions. Optionally, the dual-epitope anti-CCR8 antibody or its antigen-binding fragment contains at least two Fab fragments; Optionally, the dual-epitope anti-CCR8 antibody or its antigen-binding fragment comprises at least two Fab fragments tandemly. Optionally, at least two Fab fragments in tandem bind to different CCR8 antigenic epitopes. According to claim 11, the dual epitope anti-CCR8 antibody or its antigen-binding fragment is characterized in that, The heavy chain variable region includes complementarity-determining regions HCDR1, HCDR2 and HCDR3, and the complementarity-determining regions HCDR1, HCDR2 and HCDR3 of each heavy chain variable region are independently selected from HCDR1, HCDR2 and HCDR3 as described in any one of claims 1 to 18; The complementary determining regions LCDR1, LCDR2 and LCDR3 of the light chain variable region are selected from LCDR1, LCDR2 and LCDR3 as described in any one of claims 1 to 10; Optionally, the complementarity-determining regions HCDR1, HCDR2, and HCDR3 of the heavy chain variable region in an antigen-binding moiety include amino acid sequences consistent with HCDR1, HCDR2, and HCDR3 of the heavy chain variable region shown in SEQ ID NO. 20; The complementarity-determining regions HCDR1, HCDR2, and HCDR3 of the heavy chain variable region in another antigen-binding moiety include amino acid sequences consistent with those of HCDR1, HCDR2, and HCDR3 of the heavy chain variable region shown in SEQ ID NO. 11; The complementarity-determining regions LCDR1, LCDR2, and LCDR3 of the light chain variable region include amino acid sequences consistent with those of the light chain variable regions LCDR1, LCDR2, and LCDR3 shown in any one of SEQ ID NO. 21, 22, or 23; Optionally, according to the IMGT definition, the complementarity-determining regions HCDR1, HCDR2, and HCDR3 of the heavy chain variable region in an antigen-binding moiety are selected from: HCDR1 comprising the amino acid sequence shown in SEQ ID NO.34, HCDR2 comprising the amino acid sequence shown in SEQ ID NO.43, and HCDR3 comprising the amino acid sequence shown in SEQ ID NO.71; The complementarity-determining regions HCDR1, HCDR2, and HCDR3 of the heavy chain variable region in another antigen-binding moiety are selected from: HCDR1 includes the amino acid sequence shown in SEQ ID NO.37, HCDR2 includes the amino acid sequence shown in SEQ ID NO.51, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.73; The complementarity-determining regions LCDR1, LCDR2, and LCDR3 of the light chain variable region are selected from: LCDR1 including the amino acid sequence shown in SEQ ID NO.88, LCDR2 including the amino acid residue KV, and LCDR3 including the amino acid sequence shown in SEQ ID NO.103; Alternatively, LCDR1 includes the amino acid sequence shown in SEQ ID NO.88, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.107; Optionally, the complementarity-determining regions HCDR1, HCDR2, and HCDR3 of the heavy chain variable region in an antigen-binding moiety are selected from: HCDR1 comprising the amino acid sequence shown in SEQ ID NO.30, HCDR2 comprising the amino acid sequence shown in SEQ ID NO.41, and HCDR3 comprising the amino acid sequence shown in SEQ ID NO.76; The complementarity-determining regions HCDR1, HCDR2, and HCDR3 of the heavy chain variable region in another antigen-binding moiety are selected from: HCDR1 includes the amino acid residue DY, HCDR2 includes the amino acid sequence shown in SEQ ID NO.54, and HCDR3 includes the amino acid sequence shown in SEQ ID NO.74; The complementary determinant regions LCDR1, LCDR2, and LCDR3 of the light chain variable region are selected from: LCDR1 includes the amino acid sequence shown in SEQ ID NO. 92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO. 106; or, LCDR1 includes the amino acid sequence shown in SEQ ID NO.92, LCDR2 includes the amino acid residue KV, and LCDR3 includes the amino acid sequence shown in SEQ ID NO.

102. The dual-epitope anti-CCR8 antibody or its antigen-binding fragment according to claim 11 or 12 is characterized in that, The dual epitope anti-CCR8 antibody or its antigen-binding fragment contains a heavy chain framework region and / or a light chain framework region, wherein the heavy chain framework region and / or the light chain framework region are derived from at least one of a mouse antibody, a human antibody, a primate antibody or a mutant thereof; Optionally, the framework regions of the heavy chain variable region include HFR1, HFR2, HFR3, and HFR4, and the framework regions of the light chain variable region include LFR1, LFR2, LFR3, and LFR4, wherein... HFR1, HFR2, HFR3 and HFR4 of each heavy chain variable region are independently selected from HFR1, HFR2, HFR3 and HFR4 as described in any one of claims 1 to 18; and / or, LFR1, LFR2, LFR3 and LFR4 of each light chain variable region are independently selected from LFR1, LFR2, LFR3 and LFR4 as described in any one of claims 1 to 18. Optionally, the framework regions of the heavy chain variable region include HFR1, HFR2, HFR3, and HFR4, and the framework regions of the light chain variable region include LFR1, LFR2, LFR3, and LFR4, wherein HFR1, HFR2, HFR3, and HFR4 of the framework regions of the heavy chain variable region are selected from: HFR1 containing the amino acid sequence shown in SEQ ID NO.113, HFR2 containing the amino acid sequence shown in SEQ ID NO.120, HFR3 containing the amino acid sequence shown in SEQ ID NO.141, and HFR4 containing the amino acid sequence shown in SEQ ID NO.146; And / or, the HFR1, HFR2, HFR3 and HFR4 of the framework region of the heavy chain variable region are selected from: HFR1 containing the amino acid sequence shown in SEQ ID NO.112, HFR2 containing the amino acid sequence shown in SEQ ID NO.122, HFR3 containing the amino acid sequence shown in SEQ ID NO.144, and HFR4 containing the amino acid sequence shown in SEQ ID NO.146; And / or, the LFR1, LFR2, LFR3 and LFR4 of the framework region of the light chain variable region are selected from: the LFR1 containing the amino acid sequence shown in SEQ ID NO.153, the LFR2 containing the amino acid sequence shown in SEQ ID NO.162, the LFR3 containing the amino acid sequence shown in SEQ ID NO.165, and the LFR4 containing the amino acid sequence shown in SEQ ID NO.178; Alternatively, according to the IMGT definition: The HFR1, HFR2, HFR3 and HFR4 of the framework region of the heavy chain variable region are selected from: HFR1 containing the amino acid sequence shown in SEQ ID NO.113, HFR2 containing the amino acid sequence shown in SEQ ID NO.120, HFR3 containing the amino acid sequence shown in SEQ ID NO.141, and HFR4 containing the amino acid sequence shown in SEQ ID NO.146; And / or, the HFR1, HFR2, HFR3 and HFR4 of the framework region of the heavy chain variable region are selected from: HFR1 containing the amino acid sequence shown in SEQ ID NO.112, HFR2 containing the amino acid sequence shown in SEQ ID NO.114, HFR3 containing the amino acid sequence shown in SEQ ID NO.143, and HFR4 containing the amino acid sequence shown in SEQ ID NO.146; And / or, as defined by IMGT: The LFR1, LFR2, LFR3, and LFR4 of the framework region of the light chain variable region are selected from: LFR1 containing the amino acid sequence shown in SEQ ID NO. 154, LFR2 containing the amino acid sequence shown in SEQ ID NO. 160, LFR3 containing the amino acid sequence shown in SEQ ID NO. 168, and LFR4 containing the amino acid sequence shown in SEQ ID NO. 178; or, The LFR1 contains the amino acid sequence shown in SEQ ID NO.156, the LFR2 contains the amino acid sequence shown in SEQ ID NO.160, the LFR3 contains the amino acid sequence shown in SEQ ID NO.174, and the LFR4 contains the amino acid sequence shown in SEQ ID NO.

178. The dual-epitope anti-CCR8 antibody or its antigen-binding fragment according to any one of claims 11 to 13 is characterized in that, The amino acid sequence of the heavy chain variable region in one antigen-binding region is shown in SEQ ID NO.20, and the amino acid sequence of the heavy chain variable region in the other antigen-binding region is shown in SEQ ID NO.11; And / or, the amino acid sequences of the variable regions of the light chain are as shown in SEQ ID NO.21, 22 or 23. The dual-epitope anti-CCR8 antibody or its antigen-binding fragment according to any one of claims 11 to 14 is characterized in that, The dual epitope anti-CCR8 antibody or its antigen-binding fragment includes part or all of the sequence of a constant region, wherein the constant region is derived from at least one of a murine antibody, a human antibody, a primate antibody or a mutant thereof; Optionally, the heavy chain constant region contains a sequence of part or all of the constant regions of any one of IgG1, IgG2a, IgG2b, IgG3, IgG4, IgA, IgM, IgE or IgD; the light chain constant region of the antibody is a κ or λ chain; Optionally, the heavy chain constant region contains a sequence of murine antibody IgG1 or a sequence of murine antibody IgG2a; Optionally, the heavy chain constant region contains the sequence of human antibody IgG1 and has S239D, A330L and I332E mutations; Optionally, it contains two heavy chain constant regions, each containing the sequence of the heavy chain constant region of IgG1; Optionally, the heavy chain constant region of IgG1 is a constant region of human IgG1; Optionally, the heavy chain constant region of the human IgG1 contains S239D and I332E mutations; Optionally, the amino acid sequences of the two heavy chain constant regions are shown in SEQ ID NO.181 and SEQ ID NO.182, respectively; Optionally, it contains a light chain constant region; Optionally, the amino acid sequence of the light chain constant region is shown in SEQ ID NO.183; Optionally, it contains two full-length heavy chains and two full-length light chains, forming a symmetrical structure; Optionally, the Fc regions of the two heavy chains of the dual epitope anti-CCR8 antibody or its antigen-binding fragment contain Knob and Hole structures, respectively, and bind in a "Knob-into-Hole" manner; Optionally, the complementarity-determining region includes the heavy chain variable region containing a Knob structure, where the amino acid sequence of the heavy chain variable region is consistent with HCDR1, HCDR2 and HCDR3 of the heavy chain variable region shown in SEQ ID NO.20 is located. Optionally, the complementarity-determining region includes a light chain containing a Hole structure where the heavy chain variable region, which contains an amino acid sequence identical to HCDR1, HCDR2, and HCDR3 of the heavy chain variable region shown in SEQ ID NO.11, is located. Optionally, the amino acid sequences of the heavy chains are shown in SEQ ID NO.184 and 185, respectively, and the amino acid sequences of the two light chains are shown in SEQ ID NO.

186. Alternatively, the amino acid sequences of the heavy chains are shown in SEQ ID NO.184 and 185, respectively, and the amino acid sequences of the two light chains are shown in SEQ ID NO.

187. Alternatively, the amino acid sequences of the heavy chains are shown in SEQ ID NO.184 and 185, respectively, and the amino acid sequences of the two light chains are shown in SEQ ID NO.

188. A biomaterial, characterized in that, The biomaterial includes any one of the following (i) to (vi): (i) A multispecific antibody comprising the anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 1 to 10; (ii) a chimeric antigen receptor, wherein the chimeric antigen receptor comprises an extracellular region, a transmembrane region and an intracellular region; Wherein, the extracellular region includes the anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 1 to 10; (iii) A nucleic acid molecule encoding the anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 1 to 10, (i) the multispecific antibody, recombinant protein and immunoconjugate, (i) the chimeric antigen receptor, or the dual epitope anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 11 to 15. (iv) A vector carrying the nucleic acid molecule described in (iii); (v) Recombinant cells, characterized in that they comprise the nucleic acid molecule described in (iii), or the vector described in (iv), or express the anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 1 to 10, or express the multispecific antibody, recombinant protein and immunoconjugate described in (i), express the chimeric antigen receptor described in (ii), or the dual epitope anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 11 to 15; (vi) Engineered immune effector cells that express the chimeric antigen receptor described in (ii) or contain a nucleic acid molecule encoding the chimeric antigen receptor described in (ii). A pharmaceutical composition, characterized in that, The pharmaceutical composition contains the anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 1 to 10, or the dual epitope anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 11 to 15, or the biological material as described in claim 16; Optionally, the pharmaceutical composition may further include a pharmaceutically acceptable carrier and / or excipient. The use of the anti-CCR8 antibody or antigen-binding fragment thereof according to any one of claims 1 to 10, or the dual-epitope anti-CCR8 antibody or antigen-binding fragment thereof according to any one of claims 11 to 15, or the biological material according to claim 16, or the pharmaceutical composition according to claim 17, in any one of the following: (I) Detect CCR8 or cells expressing CCR8; (II) Prepare products for detecting CCR8 or cells expressing CCR8; (III) Used for the treatment, prevention or relief of diseases, symptoms or conditions related to CCR8; (IV) Prepare pharmaceutical compositions for treating, preventing or alleviating diseases, symptoms or conditions associated with CCR8; (V) Used for the treatment, prevention or relief of tumors; (VI) Prepare pharmaceutical compositions for the treatment, prevention or relief of tumors. A method for producing an anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 1-10, or a dual-epitope anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 11-15, characterized in that... The method includes the following steps: (a) The recombinant cells of claim 16(v) are cultured under conditions expressing an antibody or an antigen-binding fragment thereof; (b) Separation and purification of the antibody or its antigen-binding fragment obtained in step (a). A kit for detecting CCR8, characterized in that, It comprises the anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 1 to 10, or the dual epitope anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 11 to 15, or the biological material as described in claim 16. A method for treating, preventing, or alleviating a disease, symptom, or condition, characterized in that, The method comprises administering to a subject a therapeutically effective amount of the anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 1 to 10, or the dual epitope anti-CCR8 antibody or its antigen-binding fragment as described in any one of claims 11 to 15, or the biological material as described in claim 16, or the pharmaceutical composition as described in claim 17. Preferably, the disease, symptom, or condition includes tumors, inflammation, autoimmune diseases, infections, and immune rejection due to organ transplantation.

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