Degradation agent-antibody conjugates, preparation method therefor, and use thereof

Abstract

The present application provides a class of degradation agent-antibody conjugates, a preparation method therefor, and use thereof. Specifically, provided are degradation agent-antibody conjugates represented by formula (I), a preparation method therefor, and use thereof in the preparation of drugs for treating and / or preventing cancer. Also provided are a drug linker represented by formula (II), a preparation method therefor, and use thereof in the preparation of degradation agent-antibody conjugates.

Description

A class of degradation agent antibody conjugates, preparation methods and applications

[0001] Cross-referencing

[0002] This application is based on and claims priority to CN application No. 202510123164.1 filed on January 24, 2025, the contents of which are incorporated herein by reference in their entirety. Technical Field

[0003] This disclosure pertains to the pharmaceutical field, specifically relating to a class of degradation agent antibody conjugates, their preparation methods, and their applications. Background Technology

[0004] Degrader-Antibody Conjugates (DACs) are a novel type of targeted therapy drug, consisting of an antibody and a protein degrader linked by a linker unit. Utilizing the high specificity of antibodies, they recognize and bind to specific antigens on the cell surface, thereby precisely delivering the protein degrader into the target cell to degrade the target protein. This design not only improves drug targeting but also addresses the issues of low bioavailability and poor pharmacokinetic properties of protein degraders themselves through antibody internalization.

[0005] The ubiquitin-proteasome system (UPS) is one of the two naturally occurring protein degradation pathways in eukaryotic cells, mediating 80-90% of protein degradation in the body and is the most important protein degradation system in eukaryotes. Ubiquitin-mediated protein degradation mainly utilizes ubiquitin activator E1, ubiquitin conjugate E2, and ubiquitin-protein ligase E3 to load ubiquitin onto target proteins for labeling and then deliver it to the 26S proteasome for degradation.

[0006] Currently, protein degraders developed using the ubiquitin-proteasome system mainly include two types: PROTACs and molecular gels. These protein degraders induce or stabilize protein-protein interactions between E3 ubiquitinase and target proteins, forming ternary complexes that lead to ubiquitination and degradation of the target protein. The earliest approved molecular gel drug was thalidomide. One part of the thalidomide molecule interacts with the E3 ubiquitin ligase CRBN, while another part links to IKZF1 / 3 proteins, inducing CRBN and IKZF1 / 3 proteins to approach each other. Subsequently, IKZF1 / 3 proteins are ubiquitinated and ultimately degraded by the proteasome. Building upon thalidomide, researchers have obtained molecular gels with different degradation properties through reasonable structural optimization, and these gels can recruit different novel substrates to lead to their degradation, such as CK1α and GSPT1.

[0007] G1-to-S phase transition protein 1 (GSPT1) belongs to the eukaryotic peptide chain releasing factor (eRF3a) class and is a GTPase primarily located in the cytoplasm, widely expressed in various organs and tissues of the human body. GSPT1 is a translation termination factor that recognizes stop codons by forming a translation termination complex (eRF1 / eRF3 / GTP complex), forcing the protein to dissociate from the ribosome after translation. GSPT1 protein is highly expressed in various tumors, and downregulation of its expression can inhibit tumor cell proliferation, migration, and invasion. Summary of the Invention

[0008] This disclosure provides a class of degradation agent antibody-drug conjugates (DACs) using GSPT1 protein degraders as loads, their preparation methods, and their applications in the prevention and / or treatment of cancer.

[0009] In a first aspect, this application provides compounds of formula (I) or pharmaceutically acceptable salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, isotopically labeled compounds, metabolites, or prodrugs thereof:

[0010] in,

[0011] Ab 1 For targeting groups;

[0012] M represents the junction site that connects to the target group;

[0013] L is the connector that connects M and D;

[0014] m is selected from 1-20;

[0015] D is selected from the structures of formulas (DI-1) and (DI-2);

[0016] X 1 Selected from CH2 and -C(=O)-;

[0017] X 2 X 3 X 4 Each is independently selected from O, S, Se, N, C, and CR. 3 And when X 2 X 3 X 4 When one of the elements is independently selected from O, S, and Se, the other two are independently selected from N, C, and CR. 3 ;

[0018] It can be a single bond or a double bond;

[0019] X 5 Selected from O, S, C1-6 Alkylene and C 1-6 Halogenated alkylene;

[0020] Ring A is selected from phenyl and 5-6-membered heteroaryl groups;

[0021] Y is selected from C 6-10 Aryl and 5-10 heteroaryl groups, wherein the aryl and heteroaryl groups are optionally separated by one or more R groups. 8 replace;

[0022] Z is selected from C 6-10 Aryl and 5-10 heteroaryl groups, said aryl and heteroaryl groups being R 7 While replacing, it can be optionally replaced by one or more R 9 replace;

[0023] L 1 Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Haloalkylene -, -O-(CH2) e -、-N(R a )-(CH2) e -、-(CH2-CH2-O) f -、C 3-10 Cycloalkyl groups, 3-12 membered heterocyclic groups, and divalent structures composed of one or more of the above structural units;

[0024] R 1 Selected from C 6-10 Aryl and 5-10 heteroaryl, said aryl and heteroaryl being -L a -L b -L c -R 5 While replacing, it can be optionally replaced by one or more R 4 replace;

[0025] R 2 Selected from H and C 1-6 alkyl;

[0026] R 3 Selected from H, halogen, hydroxyl, cyano, C 1-6 Alkyl, C 1-6 Halogenated alkyl groups and -OC 1-6 alkyl;

[0027] R 4 Each is independently selected from H, halogen, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, C3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 6 replace;

[0028] R 5 Selected from

[0029] R 6 Each is independently selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Halogenated alkyl groups, -OC 1-6 Alkyl, C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more groups selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2- 6-acetylinyl, C 1-6 Haloalkyl, C 1-6 Alkoxy and C 1-6 Substituents of haloalkoxy groups;

[0030] L a Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Halogenated alkylene groups -, -(CH2) e -O-、-(CH2) e -N(R a )-、-(CH2-CH2-O) f -、-(CH2-CH2-O) f -(CH2) e -N(R a )-、-(CH2) e -O-(CH2) e -N(R a - and -(CH2) e -C(=O)-O-;

[0031] L b Selected from covalent bonds, C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 Aryl and 5-10 heteroaryl groups;

[0032] L c Selected from covalent bonds, -O-, -S-, -S(=O)-, -S(=O)2-, -N-(R a )-、-C(=O)N(R a )-、-C(=S)N(R a )-, -C(=O)-, -C(=S)-, -C(=O)-O-, -OC(=O)-O-, -OC(=O)-N(R a )-、-N(R a )-C(=O)-N(R a )-、-S(=O)2-N(R a )-, -S(=O)2-NH-C(=O)-O- and -S(=O)2-NH-C(=O)-N(R a )-;

[0033] R 8 Each is independently selected from H, halogen, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b -OR a -C 1-6 Alkylene-OR a -C 1-6 Alkylene-N(R) a )R b C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 10 replace;

[0034] R 10 Each is independently selected from H, halogen, cyano, and C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b and -OR a ;

[0035] R 7 Selected from

[0036] R 9 Each is independently selected from H, halogen, cyano, and C. 1-6 Alkyl, C2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b -OR a C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 11 replace;

[0037] R 11 Each is independently selected from H, halogen, cyano, and C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b and -OR a ;

[0038] R a Each is independently selected from H and C. 1-6 alkyl;

[0039] R b Each is independently selected from H and C. 1-6 alkyl;

[0040] z is selected from 1, 2, and 3;

[0041] f are each independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10;

[0042] e is independently selected from 0, 1, 2, 3, 4, 5, and 6;

[0043] In this context, position 1 of each substituent indicates the connection point with L.

[0044] On the other hand, this disclosure provides compounds of Formula II or their pharmaceutically acceptable salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, isotopically labeled compounds, metabolites, or prodrugs:

[0045] M 1 -LD

[0046] II

[0047] in,

[0048] L and D are as defined in any of the preceding items;

[0049] M 1 It is a precursor for the junction site that connects to the target group.

[0050] In another aspect, this disclosure provides a pharmaceutical composition comprising any of the compounds described herein or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, and one or more pharmaceutical excipients.

[0051] In another aspect, this disclosure provides the use of the compound of Formula II or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug thereof in the preparation of conjugates (e.g., the compound of Formula I or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug thereof).

[0052] On the other hand, this disclosure provides for the use of the compounds described herein or their pharmaceutically acceptable salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, isotopically labeled compounds, metabolites or prodrugs, or combinations thereof in the preparation of medicaments, particularly in medicaments for the treatment and / or prevention of cancer (e.g., cancers associated with GSPT1 protein dysregulation, cancers associated with HER2 expression, and / or cancers associated with B7H3 expression).

[0053] In another aspect, this disclosure provides a method for treating and / or preventing cancer (e.g., cancer associated with GSPT1 protein dysregulation, cancer associated with HER2 expression, and / or cancer associated with B7H3 expression), comprising administering to a subject in need a therapeutically and / or preventively effective amount of the compound described herein or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug, or a combination thereof. Detailed Implementation

[0054] definition

[0055] Unless otherwise defined below, all technical and scientific terms used herein are intended to have the same meaning as commonly understood by one of ordinary skill in the art. References to technical terms herein refer to techniques commonly understood in the art, including variations or equivalent substitutions of techniques that are obvious to one of ordinary skill in the art. While it is believed that the following terms will be well understood by one of ordinary skill in the art, the following definitions are set forth to better interpret this disclosure.

[0056] The terms “including,” “comprising,” “having,” “containing,” or “involving,” and their other variations herein, are inclusive or open-ended and do not exclude other unlisted elements or method steps.

[0057] The terms "conjugate" and "combination" are used interchangeably, referring to a substance obtained by linking one or more heterologous molecules(s) to a target moiety. These heterologous molecules can be, for example, bioactive molecules or drugs, including but not limited to cytotoxic agents and protein degraders. Conjugates targeting antibodies are also called antibody-drug conjugates (ADCs). The term "degrader-antibody conjugates (DACs)" specifically refers to conjugates obtained by linking a protein degrader to an antibody.

[0058] The one or more heterologous molecules can be linked to a target moiety (e.g., an antibody) via a linker. The linker can be linked to the target moiety via various chemical bonds. For example, in some embodiments, when the target moiety is an antibody, the linker is linked by forming a thioether bond with the thiol group of the antibody. In some specific ADC molecular structures, -S- only represents the thioether bond formed between the linker and the thiol group of the antibody, and does not mean that -S- is part of the linker.

[0059] The structure of the conjugates in this application can be represented by general formula I, where m refers to the number of bioactive molecular fragments linked to each target molecule. During the preparation of antibody-drug conjugates, each antibody molecule may link to different numbers of bioactive molecular fragments; therefore, generally speaking, antibody-drug conjugates are mixtures of antibody-drug conjugates with different drug-antibody conjugation ratios. In practice, DAR is usually used to represent the average number of drugs linked to the antibody.

[0060] As used herein, the term “DAR” or “drug-antibody ratio” refers to: (a) the number of linker / drug moieties linked to the antibody in a single antibody-drug conjugate (e.g., a degrader antibody-drug conjugate) molecule, which is an integer from 0 to 20, such as an integer from 1 to 10; or (b) the average number of linker / drug moieties linked to the antibody in a composition comprising more than one antibody-drug conjugate (e.g., a degrader antibody-drug conjugate) molecule, which is an integer or decimal from 0 to 20, such as an integer or decimal from 1 to 10. Methods for determining DAR are well known to those skilled in the art, including methods using reversed-phase chromatography or HPLC-MS. As used herein, the term “antibody” is used in the broadest sense to include various antibody structures, including but not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments, provided they exhibit the desired antigen-binding activity. For example, an immunoglobulin molecule may consist of two pairs of polypeptide chains, each pair having one light chain (LC) and one heavy chain (HC) Antibody light chains can be classified into κ (kappa) and λ (lambda) light chains. Heavy chains can be classified into μ, δ, γ, α, or ε, and antibody isotypes are defined as IgM, IgD, IgG, IgA, and IgE, respectively. Within both light and heavy chains, variable and constant regions are linked by a "J" region of approximately 12 or more amino acids, and the heavy chain also contains a "D" region of approximately 3 or more amino acids. Each heavy chain consists of a heavy chain variable region (VH) and a heavy chain constant region (CH). The heavy chain constant region consists of three domains (CH1, CH2, and CH3). Each light chain consists of a light chain variable region (VL) and a light chain constant region (CL). The light chain constant region consists of one domain, CL. Constant domains do not directly participate in antibody-antigen binding but exhibit various effector functions, such as mediating the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system. The VH and VL regions can be further subdivided into highly degenerated regions (called complementarity-determining regions (CDRs)) interspersed with more conserved regions called framework regions (FRs). Each VH and VL consists of three CDRs and four FRs arranged from the amino terminus to the carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions (VH and VL) of each heavy / light chain pair form the antigen-binding sites.The allocation of amino acids in different regions or domains can follow the definitions in Kabat, Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987 and 1991)), or Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883, or related studies by AbM and Martin (Martin ACR, Cheetham JC, Rees AR (1989) Modelling antibody hypervariable loops: A combined algorithm. Proc Natl Acad Sci USA 86:9268–9272). In this paper, unless the context clearly indicates otherwise, when referring to the term "antibody," it includes not only the complete antibody but also the antigen-binding fragment of the antibody.

[0061] As used in this article, the term "complementarity-determining region" or "CDR" refers to the amino acid residues in the antibody variable region responsible for antigen binding. The precise boundaries of these amino acid residues can be defined according to various numbering systems known in the art, such as the Kabat numbering system (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991), the Chothia numbering system (Chothia & Lesk (1987) J. Mol. Biol. 196: 901-917; Chothia et al. (1989) Nature 342: 878-883), the IMGT numbering system (Lefranc et al., Dev. Comparat. Immunol. 27: 55-77, 2003), or Martin's related research (Martin ACR, Cheetham JC, Rees AR (1989) Modelling antibody hypervariable loops: A combined algorithm. Proc Natl Acad Sci USA). 86:9268–9272), this definition method integrates parts of the definitions from Kabat and Chothia, and was first applied in the Oxford Molecular antibody modeling software (Martin AC R. Protein sequence and structure analysis of antibody variable domains[M] / / Antibody engineering. Springer, Berlin, Heidelberg, 2010:33-51.). For a given antibody, those skilled in the art will easily identify the CDRs defined by each numbering system. Furthermore, the correspondence between different numbering systems is well known to those skilled in the art (e.g., see Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003).

[0062] The CDR contained in the antibodies or antigen-binding fragments thereof disclosed herein can be determined according to various numbering systems known in the art. In some embodiments, the CDR contained in the antibodies or antigen-binding fragments thereof disclosed herein is preferably determined using the Chothia numbering system.

[0063] As used herein, the term “antigen-binding fragment” of an antibody refers to a molecule other than the full-length antibody, which includes a portion of the full-length antibody that binds to the antigen bound to the full-length antibody. For example, a polypeptide fragment of a full-length antibody that retains the ability to specifically bind to the same antigen bound to the full-length antibody, and / or competes with the full-length antibody for specific binding to the antigen, is also referred to as an “antigen-binding moiety.” See Fundamental Immunology, Ch. 7 (Paul, W., ed., 2nd ed., Raven Press, NY (1989), which is incorporated herein by reference in its entirety for all purposes. Antigen-binding fragments of antibodies can be generated by recombinant DNA technology or by enzymatic or chemical cleavage of intact antibodies. Non-limiting examples of antigen-binding fragments include Fab, Fab', Fab'-SH, F(ab')2, Fd, Fv, dAb and complementarity-determining region (CDR) fragments, single-chain antibodies (e.g., scFv), chimeric antibodies, diabody, linear antibody, nanobody (technology from Domantis), domain antibody (technology from Ablynx), and peptides containing at least a portion of an antibody sufficient to confer specific antigen-binding ability to the peptide. Engineered antibody variants are reviewed in Holliger et al., 2005; Nat In Biotechnol, 23:1126-1136, the term "Fd" refers to an antibody fragment composed of VH and CH1 domains; the term "dAb fragment" refers to an antibody fragment composed of VH domains (Ward et al., Nature 341:544-546 (1989)); the term "Fab fragment" refers to an antibody fragment composed of VL, VH, CL, and CH1 domains; the term "F(ab')2 fragment" refers to an antibody fragment containing two Fab fragments connected by disulfide bridges on the hinge region; the term "Fab' fragment" refers to the fragment obtained by reducing the disulfide bonds connecting the two heavy chain fragments in the F(ab')2 fragment, which consists of a complete light chain and heavy chain Fd fragment (composed of VH and CH1 domains).

[0064] The term "Fv" refers to an antibody fragment consisting of the VL and VH domains of a single arm of the antibody. Fv fragments are generally considered to be the smallest antibody fragment capable of forming a complete antigen-binding site. It is generally believed that six CDRs confer antigen-binding specificity to the antibody. However, even a variable region (such as the Fd fragment, which contains only three antigen-specific CDRs) can recognize and bind to the antigen, although its affinity may be lower than that of a complete binding site.

[0065] The term "Fc" refers to an antibody fragment formed by disulfide bonds connecting the second and third constant regions of the first heavy chain to the second and third constant regions of the second heavy chain. The Fc fragment of an antibody has various functions but does not participate in antigen binding.

[0066] The term "scFv" refers to a single polypeptide chain containing VL and VH domains linked by a linker (see, for example, Bird et al., Science 242:423-426 (1988); Huston et al., Proc. Natl. Acad. Sci. USA 85:5879-5883 (1988); and Pluckthun, The Pharmacology of Monoclonal Antibodies, Vol. 113, edited by Roseburg and Moore, Springer-Verlag, New York, pp. 269-315 (1994)). Such scFv molecules may have a general structure: NH2-VL-linker-VH-COOH or NH2-VH-linker-VL-COOH. Suitable prior art linkers consist of a repeating GGGGS (SEQ ID NO:64) amino acid sequence or a variant thereof. For example, a linker having the amino acid sequence (GGGGS)4 (SEQ ID NO: 65) can be used, but variants thereof can also be used (Holliger et al. (1993), Proc. Natl. Acad. Sci. USA 90:6444-6448). Other linkers that can be used in this disclosure are described by Alfthan et al. (1995), Protein Eng. 8:725-731, Choi et al. (2001), Eur. J. Immunol. 31:94-106, Hu et al. (1996), Cancer Res. 56:3055-3061, Kipriyanov et al. (1999), J. Mol. Biol. 293:41-56, and Roovers et al. (2001), Cancer Immunol. In some cases, a disulfide bond may also exist between the VH and VL of scFv. In some implementations, the VH and VL domains can be positioned relative to each other in any suitable arrangement. For example, scFv containing NH2-VH-VH-COOH and NH2-VL-VL-COOH.

[0067] As used herein, the term "full-length antibody" refers to an antibody composed of two "full-length heavy chains" or "heavy chains" and two "full-length light chains" or "light chains". A "full-length heavy chain" or "heavy chain" refers to a polypeptide chain that, in the N-terminal to C-terminal direction, comprises a heavy chain variable region (VH), a heavy chain constant region CH1 domain, a hinge region (HR), a heavy chain constant region CH2 domain, and a heavy chain constant region CH3 domain; and, optionally, when the full-length antibody is an IgE isotype, it also includes a heavy chain constant region CH4 domain. Preferably, the "full-length heavy chain" is a polypeptide chain composed of VH, CH1, HR, CH2, and CH3 in the N-terminal to C-terminal direction. A "full-length light chain" or "light chain" is a polypeptide chain composed of a light chain variable region (VL) and a light chain constant region (CL) in the N-terminal to C-terminal direction. The two pairs of full-length antibody chains are linked together by disulfide bonds between CL and CH1 and between the HRs of the two full-length heavy chains. The full-length antibody disclosed herein can be derived from a single species, such as humans; it can also be a chimeric antibody or a humanized antibody. The full-length antibody disclosed herein comprises two antigen-binding sites formed by VH and VL pairs, respectively, which specifically recognize / bind to the same antigen.

[0068] In this document, the techniques for obtaining antibodies may use conventional techniques known to those skilled in the art (e.g., recombinant DNA techniques or enzymatic or chemical fragmentation methods) to obtain antigen-binding fragments of the antibody (e.g., the antibody fragments described above) from a given antibody (e.g., the antibody provided in this disclosure), and the antigen-binding fragments of the antibody may be specifically screened in the same manner as those used for intact antibodies.

[0069] As used herein, the terms “monoclonal antibody,” “monoclonal antibody,” and “mAb” have the same meaning and are used interchangeably. They refer to an antibody or a fragment of an antibody derived from a group of highly homologous antibody molecules; that is, a group of identical antibody molecules except for the possibility of spontaneous natural mutations. Monoclonal antibodies have high specificity for a single epitope on an antigen. Polyclonal antibodies, as opposed to monoclonal antibodies, typically contain at least two or more different antibodies that typically recognize different epitopes on an antigen. Furthermore, the modifier “monoclonal” only indicates that the antibody is derived from a highly homologous group of antibodies and should not be construed as requiring preparation by any particular method.

[0070] The monoclonal antibodies disclosed herein can be prepared using a variety of techniques, such as hybridoma techniques (see, for example, Kohler et al. Nature, 256:495, 1975), recombinant DNA techniques (see, for example, U.S. Patent Application 4,816,567), or phage antibody library techniques (see, for example, Clackson et al. Nature 352:624-628, 1991, or Marks et al. J. Mol. Biol. 222:581-597, 1991).

[0071] For example, monoclonal antibodies can be prepared as follows: First, immunize mice or other suitable host animals with an immunogen (with adjuvants added if necessary). The immunogen or adjuvant is usually injected subcutaneously at multiple sites or intraperitoneally. The immunogen can be pre-conjugated to certain known proteins, such as serum albumin or soybean trypsin inhibitors, to enhance the immunogenicity of the antigen in the host. The adjuvant can be Freund's adjuvant or MPL-TDM, etc. After immunization, the animal will produce lymphocytes that secrete antibodies specifically binding to the immunogen. Alternatively, lymphocytes can be obtained through in vitro immunization. Collect the target lymphocytes and fuse them with myeloma cells using a suitable fusion agent, such as PEG, to obtain hybridoma cells (Goding, Monoclonal Antibodies: Principles and Practice, pp. 59-103, Academic Press, 1996). The hybridoma cells prepared above can be seeded into a suitable culture medium for growth, preferably containing one or more substances that can inhibit the growth of unfused maternal myeloma cells. For example, for maternal myeloma cells lacking hypoxanthine-guanine phosphotransferase (HGPRT or HPRT), adding substances such as hypoxanthine, aminopterin, and thymine (HAT medium) to the culture medium can inhibit the growth of HGPRT-deficient cells. Preferred myeloma cells should possess characteristics such as high fusion rate, stable antibody secretion capacity, and sensitivity to HAT culture medium. Among these, murine myeloma cells are preferred, such as MOP-21 or MC-11 mouse tumor-derived lines (THE Salk Institute Cell Distribution Center, San Diego, Calif.USA), and SP-2 / 0 or X63-Ag8-653 cell lines (American Type Culture Collection, Rockville, Md.USA). Other studies have reported the preparation of human monoclonal antibodies using human myeloma and human-mouse heterologous myeloma cell lines (Kozbor, J. Immunol., 133:3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp.51-63, Marcel Dekker, Inc., New York, 1987). The culture medium for growing hybridoma cells is used to detect the production of monoclonal antibodies against specific antigens. Methods for determining the binding specificity of monoclonal antibodies produced by hybridoma cells include, for example, immunoprecipitation or in vitro binding assays such as radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA).For example, the affinity of monoclonal antibodies can be determined using the Scatchard assay described by Munson et al. in Anal. Biochem. 107:220 (1980). Once the specificity, affinity, and reactivity of the antibodies produced by the hybridoma have been determined, the target cell line can be subcloned using the standard limiting dilution method described in Goding, Monoclonal Antibodies: Principles and Practice, pp. 59-103, Academic Press, 1996. Suitable culture media may include DMEM or RPMI-1640. Additionally, hybridoma cells can also grow in animals in the form of ascites tumors. Monoclonal antibodies secreted by subclonal cells can be separated from cell culture medium, ascites, or serum using conventional immunoglobulin purification methods such as protein A agarose gel chromatography, hydroxyapatite chromatography, gel electrophoresis, dialysis, or affinity chromatography.

[0072] Monoclonal antibodies can also be obtained through recombinant genetic engineering. PCR amplification using nucleic acid primers that specifically bind to the heavy and light chain genes of monoclonal antibodies allows the isolation of DNA molecules encoding these genes from hybridoma cells. The resulting DNA molecules are then inserted into an expression vector, transfected into host cells (such as E. coli cells, COS cells, CHO cells, or other non-immunoglobulin-producing myeloma cells), and cultured under suitable conditions to obtain recombinantly expressed target antibodies.

[0073] Antibodies can be purified using known techniques, such as affinity chromatography with protein A or protein G. Subsequently, or alternatively, the specific antigen (the target molecule recognized by the antibody) or its epitope can be immobilized on a column and purified by immunoaffinity chromatography to purify the immunospecific antibody. For purification of immunoglobulins, see, for example, D. Wilkinson (The Scientist, published by The Scientist, Inc., Philadelphia Pa., Vol. 14, No. 8 (Apr. 17, 2000), pp. 25-28).

[0074] As used in this article, the term "mouse antibody" refers to a mouse hybrid fusion cell derived from the fusion of B cells and myeloma cells from immunized mice. These cells are then screened to identify those capable of unlimited proliferation and antibody secretion, followed by screening, antibody preparation, and purification. Alternatively, antibodies may be produced by the differentiation and proliferation of B cells after antigen invasion of the mouse, forming plasma cells that can secrete antibodies. Antibodies are also produced by stimulation with specific antigens. Antibody production occurs because antigen invasion of the human body triggers interactions among various immune cells, causing B cells in lymphocytes to differentiate and proliferate, forming plasma cells that can secrete antibodies.

[0075] As used herein, the term "humanized antibody" refers to a genetically engineered non-human antibody whose amino acid sequence has been modified to increase sequence homology with human antibodies. Typically, all or part of the CDR region of a humanized antibody is derived from a non-human antibody (donor antibody), and all or part of the non-CDR region (e.g., the variable region FR and / or constant region) is derived from a human immunoglobulin (receptor antibody). Humanized antibodies generally retain the intended properties of the donor antibody, including but not limited to antigen specificity, affinity, reactivity, ability to enhance immune cell activity, and ability to enhance immune responses. Donor antibodies can be mouse, rat, rabbit, or non-human primate (e.g., cynomolgus monkey) antibodies with the intended properties (e.g., antigen specificity, affinity, reactivity, ability to enhance immune cell activity, and / or ability to enhance immune responses).

[0076] Humanized antibodies are particularly advantageous because they retain the intended properties of non-human donor antibodies (e.g., murine antibodies) while effectively reducing their immunogenicity in human subjects. However, due to the matching problem between the CDR of the donor antibody and the FR of the receptor antibody, the intended properties of humanized antibodies (e.g., antigen specificity, affinity, reactivity, ability to enhance immune cell activity, and / or ability to enhance the immune response) are generally lower than those of non-human donor antibodies (e.g., murine antibodies).

[0077] Therefore, although researchers in this field have conducted in-depth research on antibody humanization and made some progress (see, for example, Jones et al., Nature, 321:522 525 (1986); Reichmann et al., Nature, 332:323 329 (1988); Presta, Curr. Op. Struct. Biol., 2:593 596 (1992); and Clark, Immunol. Today 21:397 402 (2000)), existing technologies do not provide detailed guidance on how to fully humanize a donor antibody so that the resulting humanized antibody has the highest possible degree of humanization while retaining the expected properties of the donor antibody as much as possible. Technicians need to explore, investigate, and modify specific donor antibodies, and it takes a lot of creative work to obtain humanized antibodies that have a high degree of humanization (e.g., at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%) while retaining the expected properties of the specific donor antibody.

[0078] In this disclosure, in order to preserve as much of the properties of the donor antibody as possible (including, for example, antigen specificity, affinity, reactivity, ability to enhance immune cell activity and / or ability to enhance immune response), the framework region (FR) of the humanized antibody of this disclosure may contain both amino acid residues of the human receptor antibody and the corresponding non-human donor antibody.

[0079] The humanized antibodies disclosed herein can be prepared based on the sequence of the mouse monoclonal antibody prepared as described above. The DNA encoding the heavy and light chains can be obtained from the target mouse hybridoma and engineered using standard molecular biology techniques to contain non-mouse (e.g., human) immunoglobulin sequences.

[0080] To prepare humanized antibodies, mouse CDR regions can be inserted into human framework sequences using methods known in the art (see Winter's U.S. Patent No. 5,225,539; Queen et al.'s U.S. Patent Nos. 5,530,101, 5,585,089, 5,693,762, and 6,180,370; and Lo, Benny, KC, editor, in Antibody Engineering: Methods and Protocols, volume 248, Humana Press, New Jersey, 2004). Alternatively, transgenic animals that do not produce endogenous immunoglobulins after immunization and can generate a complete human antibody library can be used. For example, it has been reported that homozygous deletion of the antibody heavy chain linker (JH) gene in chimeric and germline mutant mice can completely suppress the production of endogenous antibodies, and then transfer of human germline immunoglobulin gene arrays into said germline mutant mice will cause the mice to produce human antibodies upon encountering antigen stimulation (see, for example, Jakobovits et al., 1993, Proc. Natl. Acad. Sci. USA 90: 2551; Jakobovits et al., 1993, Nature 362: 255-258; Bruggermann et al., 1993, Year in Immunology 7: 33; and Duchosal et al., 1992, Nature 355: 258). Non-limiting examples of the aforementioned transgenic animals include the HuMAb mouse (Medarex, Inc.) containing a miniloci of the human immunoglobulin gene encoding unrearranged human heavy chain (μ and γ) and κ light chain immunoglobulin sequences, coupled with targeted mutations that inactivate the endogenous μ and κ chain loci (see, for example, Lonberg et al. (1994) Nature 368(6474):856-859); or the “KM mouse™” carrying human heavy chain transgenes and human light chain transchromosomes (see patent application WO02 / 43478). Other methods for humanizing antibodies include phage display technology (Hoogenboom et al., 1991, J. Mol. Biol. 227:381; Marks et al., J. Mol. Biol. 1991, 222:581-597; Vaughan et al., 1996, Nature Biotech 14:309).

[0081] As used herein, the term "specific binding" refers to a non-random binding reaction between two molecules, such as the reaction between an antibody and its target antigen. The strength or affinity of a specific binding interaction can be expressed as the equilibrium dissociation constant (KD) or half-maximal effective concentration (EC50) of the interaction.

[0082] The specific binding properties between two molecules can be determined using methods known in the art. One method involves measuring the rate of formation and dissociation of the antigen binding site / antigen complex. Both the “binding rate constant” (ka or kon) and the “dissociation rate constant” (kdis or koff) can be calculated from the concentration and the actual rates of association and dissociation (see Malmqvist M, Nature, 1993, 361:186-187). The ratio of kdis / kon is equal to the dissociation constant KD (see Davies et al., Annual Rev Biochem, 1990; 59:439-473). The values ​​of KD, kon, and kdis can be measured using any effective method. In some embodiments, the dissociation constant can be measured using bioluminescent interferometry (e.g., the ForteBio Octet method). Alternatively, surface plasmon resonance techniques (e.g., Biacore) or Kinexa can be used to measure the dissociation constant.

[0083] The twenty common amino acids referred to herein are written in accordance with conventional usage. See, for example, Immunology-A Synthesis (2nd Edition, E.S. Golub and D.G. Ren, Eds., Sinauer Associates, Sunderland, Mass. (1991)), which is incorporated herein by reference. In this disclosure, the terms “polypeptide” and “protein” have the same meaning and are used interchangeably. And in this disclosure, amino acids are generally represented by single-letter and three-letter abbreviations known in the art. For example, alanine may be represented by A or Ala.

[0084] The "sequence identity percentage" or "identity percentage" between two polynucleotide or polypeptide sequences refers to the number of identical matching positions shared by sequences within a comparison window, taking into account additions or deletions (i.e., vacancies) that must be introduced for optimal alignment of the two sequences. A matching position is any location where the same nucleotide or amino acid is present in both the target and reference sequences. Vacancies are not nucleotides or amino acids and are not counted in the target sequence. Similarly, vacancies in the reference sequence are not counted because nucleotides or amino acids from the target sequence are included, but those from the reference sequence are excluded.

[0085] The percentage of sequence identity can be calculated as follows: determine the number of positions in both sequences where the same amino acid residue or nucleic acid base appears (the number of matching positions), divide the number of matching positions by the total number of positions in the comparison window, and multiply the result by 100 to obtain the percentage of sequence identity. Sequence comparison and determination of the percentage of sequence identity between two sequences can be accomplished using software that is readily available online and downloadable. Suitable software programs are available from various sources for protein and nucleotide sequence alignment. A suitable program for determining the percentage of sequence identity is bl2seq, which is part of the BLAST program suite available from the National Center for Biotechnology Information (NCBI) website (blast.ncbi.nlm.nih.gov). Bl2seq uses either the BLASTN or BLASTP algorithm for comparing two sequences. BLASTN is used to compare nucleic acid sequences, while BLASTP is used to compare amino acid sequences. Other suitable programs are, for example, Needle, Stretcher, Water, or Matcher, which are part of the EMBOSS suite of bioinformatics programs and are also available from the European Institute of Bioinformatics (EBI) at www.ebi.ac.uk / Tools / psa.

[0086] As used herein, the term "conservative substitution" refers to an amino acid substitution that does not adversely affect or alter the intended properties of a protein / peptide containing an amino acid sequence. For example, conservative substitutions can be introduced using standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions include substitutions of amino acid residues with amino acid residues having similar side chains, such as substitutions with residues that are physically or functionally similar to the corresponding amino acid residues (e.g., having similar size, shape, charge, chemical properties, including the ability to form covalent or hydrogen bonds). Families of amino acid residues with similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, and histidine), acidic side chains (e.g., aspartic acid and glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, and tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, and methionine), β-branched side chains (e.g., threonine, valine, and isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, and histidine). Therefore, it is preferable to replace the corresponding amino acid residue with another amino acid residue from the same side chain family. Methods for identifying conserved amino acid substitutions are well known in the art (see, for example, Brummell et al., Biochem. 32:1180-1187 (1993); Kobayashi et al., Protein Eng. 12(10):879-884 (1999); and Burks et al., Proc. Natl Acad. Set USA 94:412-417 (1997), which are incorporated herein by reference).

[0087] As used herein, the term “pharmaceuticalally acceptable carrier” means a carrier that is pharmacologically and / or physiologically compatible with the subject and the active ingredient, and is well known in the art (see, for example, Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th ed. Pennsylvania: Mack Publishing Company, 1995).

[0088] As used herein, the term “prevention” refers to methods implemented to prevent or delay the occurrence of a disease or condition or symptom (e.g., a tumor) in a subject.

[0089] As used herein, the term "treatment" refers to a method performed to achieve a beneficial or desired clinical outcome. For the purposes of this disclosure, a beneficial or desired clinical outcome includes, but is not limited to, relief of symptoms, reduction of the extent of disease, stabilization (i.e., cessation of disease progression) of the disease state, delay or slowing of disease progression, improvement or relief of the disease state, and relief of symptoms (whether partial or complete), whether detectable or undetectable. Furthermore, "treatment" can also mean prolongation of survival compared to expected survival (if no treatment was received).

[0090] As used herein, the term "subject" refers to a mammal, such as a primate mammal, like a human. In some embodiments, the subject (e.g., a human) has a tumor, or is at risk of having the aforementioned disease.

[0091] As used herein, the term "effective amount" means an amount sufficient to achieve, or at least partially achieve, the desired effect. For example, an effective amount for preventing disease (e.g., cancer) means an amount sufficient to prevent, stop, or delay the onset of disease (e.g., cancer); an effective amount for treating disease means an amount sufficient to cure or at least partially stop the disease and its complications in a patient already suffering from the disease. Determining such an effective amount is entirely within the capabilities of those skilled in the art. For example, an effective amount for therapeutic purposes will depend on the severity of the disease to be treated, the overall state of the patient's own immune system, the patient's general characteristics such as age, weight, and sex, the manner of administration of the drug, and other concurrent treatments, etc.

[0092] The terms "cancer" and "tumor" are used interchangeably to refer to a large class of diseases characterized by the uncontrolled growth of abnormal cells in the body. Uncontrolled cell division can lead to the formation of malignant tumors or cells that invade adjacent tissues and can metastasize to distant sites of the body via the lymphatic system or bloodstream. Cancer includes benign and malignant cancers, as well as dormant tumors or micrometastases. Cancer also includes hematologic malignancies, especially hematologic malignancies.

[0093] As used herein, the term "alkyl" is defined as a straight-chain or branched saturated aliphatic hydrocarbon group. For example, as used herein, the term "C" is... 1-6 "Alkyl" refers to a straight-chain or branched group having 1 to 6 carbon atoms (e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, or n-hexyl), optionally substituted with one or more (such as 1 to 3) suitable substituents such as halogens. The term "C" 1-4 "Alkyl" refers to a straight-chain or branched group having 1 to 4 carbon atoms (e.g., methyl, ethyl, n-propyl, isopropyl), which is optionally substituted by one or more (such as 1 to 3) suitable substituents such as halogens.

[0094] As used herein, the term "alkoxy" refers to "alkyl-O-", where "alkyl" is as defined above. For example, the term "C" 1-6 "Alkoxy" refers to "C 1-6 alkyl-O-", the "C" 1-6 "alkyl" is as defined above. "C" as used in this disclosure 1-4 "Alkoxy" refers to "C 1-4 alkyl-O-", the "C" 1-4 "Alkyl" is as defined above. An exemplary C 1-6 Alkoxy groups include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, tert-butoxy, n-pentoxy, or n-hexoxy.

[0095] The term "alkylene" refers to a divalent group formed by the loss of two hydrogen atoms from a straight-chain or branched alkane, including, for example, "C". 1-20 Alkylene, C 1- 10 Alkylene, C 1-6 Alkylene, C 1-4 Alkylene, C 1-3 "alkylene", etc., specific examples include but are not limited to: methylene, ethylene, 1,3-propylene, 1,4-butylene, 1,5-pentylene or 1,6-hexylene, etc.

[0096] The term "alkenyl" refers to a divalent group formed by the loss of two hydrogen atoms from a straight-chain or branched hydrocarbon group containing at least one carbon-carbon double bond, including, for example, "C". 2-6 "Ideinyl", "C" 2-4 Examples of these include, but are not limited to: vinylidene, 1-propenyne, 2-propenyne, 1-butenyne, 2-butenyne, 1,3-butadiene, 1-pentenyne, 2-pentenyne, 3-pentenyne, 1,3-pentadiene, 1,4-pentadiene, 1-hexenyne, 2-hexenyne, 3-hexenyne, 1,4-hexadiene, etc.

[0097] The term "acetylenic" refers to a divalent group formed by the loss of two hydrogen atoms from a straight-chain or branched hydrocarbon group containing at least one carbon-carbon triple bond. This includes, for example, "C..." 2-6 "Isynyne", "C" 2-4 "Isynyne", "C" 4-6 Examples of "ethynyl" include, but are not limited to: ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl, 1,3-butynyl, 1-pentynyl, 2-pentynyl, 3-pentynyl, 1,3-pentynyl, 1,4-pentynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 1,4-hexynyl, etc.

[0098] As used herein, the term "aryl" or "aromatic ring" refers to a fully carbon monocyclic or fused-ring polycyclic aromatic group having a conjugated π-electron system. For example, the term "C 6-10 "Aryl" or "C" 6-10 "Aromatic ring" refers to an aromatic group containing 6 to 10 carbon atoms, such as phenyl (cyclic) or naphthyl (cyclic). The aryl group is optionally substituented by one or more (such as 1 to 3) suitable substituents (e.g., halogen, -OH, -CN, -NO2, C). 1-6 Alkyl groups, etc., are substituted.

[0099] In this document, the term "heteroaryl" refers to an aromatic cyclic group in which at least one ring atom is a heteroatom, such as a nitrogen atom, an oxygen atom, or a sulfur atom. Optionally, the ring atom (e.g., a carbon atom, nitrogen atom, or sulfur atom) in the cyclic structure may be oxidized. Specific examples include, but are not limited to, 5-10-membered heteroaryl, 5-10-membered nitrogen-containing heteroaryl, 6-10-membered oxygen-containing heteroaryl, 6-8-membered nitrogen-containing heteroaryl, 5-8-membered oxygen-containing heteroaryl, etc., such as furanyl, thiophene, pyrrole, thiazolyl, isothiazolyl, thiadiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, imidazole, pyrazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl Azolyl, pyridyl, 2-pyridone, 4-pyridone, pyrimidinyl, 1,4-dioxazinyl, 2H-1,2-oxazinyl, 4H-1,2-oxazinyl, 6H-1,2-oxazinyl, 4H-1,3-oxazinyl, 6H-1,3-oxazinyl, 4H-1,4-oxazinyl, pyridazinyl, 1,2,3-triazinyl, 1,3,5-triazinyl, 1,2,4,5-tetraazinyl, aziridine-heptanetrienyl, 1,3-diazacycloheptanetrienyl, aziridine-octatetraenyl, etc.

[0100] The term "cycloalkyl" refers to a saturated cyclic hydrocarbon group, including but not limited to monocycloalkyl and bicycloalkyl groups (such as spirocycloalkyl, fused cycloalkyl, and bridged cycloalkyl). The term "C" 3-10 cycloalkyl (e.g., C10) 3-8 "Cycloalkyl" refers to cycloalkyl groups having 3 to 10 (e.g., 3-8) cyclic carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, etc.

[0101] The term "heterocyclic group" refers to a group formed by a saturated or partially saturated non-aromatic cyclic structure containing at least one ring member selected from N, O, and S. The term "3-12 membered heterocyclic group (e.g., 3-10 membered heterocyclic group, 3-8 membered heterocyclic group)" refers to a heterocyclic group having 3-12 (e.g., 3-10, 3-8) ring atoms. Specific examples include, but are not limited to, 5-6 membered heterocyclic groups, 5-6 membered nitrogen-containing heterocyclic groups, and 5-6 membered oxygen-containing heterocyclic groups, such as ethylene oxide, aziridinyl, azetidinyl, oxetanyl, thiohexacyclic, pyrrolyl, hexahydro-1H-pyrrololine, pyrrolidone, imidazoalkyl, pyrazolyl, dihydropyrroleyl, dihydroimidazoyl, dihydrofuranyl, dihydropyrazolyl, dihydropyridinyl, tetrahydrofuranyl, tetrahydrothiophenyl, tetrahydropyranyl, tetrahydropyridinyl, piperidinyl, morpholinyl, dithianyl, thiomorpholinyl, or piperazine, etc.

[0102] As used herein, the term "halogen" is defined as including fluorine, chlorine, bromine, or iodine.

[0103] As used in this article, the term "halogenation" refers to the substitution of one or more (such as 1 to 3) identical or different halogen atoms.

[0104] As used herein, the term "haloalkyl" refers to an alkyl group substituted with one or more (such as 1 to 3) identical or different halogen atoms. For example, the term "C" 1-6 "Halogenated alkyl" and "C" 1-4 "Halogenated alkyl" refers to alkyl halogens having 1 to 6 carbon atoms and 1 to 4 carbon atoms, such as -CF3, -C2F5, -CHF2, -CH2F, -CH2CF3, -CH2Cl or -CH2CH2CF3, etc.

[0105] Optionally, the hydrogen in the groups involved in this disclosure may be replaced by deuterium.

[0106] The term "substitution" refers to the selective replacement of one or more (e.g., 1, 2, 3, or 4) hydrogen atoms on a specified atom by a designated group, provided that the substitution does not exceed the normal valence of the specified atom in the present case and the substitution forms a stable compound. Combinations of substituents and / or variables are permitted only if such combinations form a stable compound.

[0107] Unless otherwise defined, the terms "substituent" or "suitable substituent" as used herein refer to modifications of a compound that can be made by those skilled in the art to suit the needs of the compound's substituents, including oxo (=O), halogen, cyano, amino, C... 1-6 Alkylamino, carboxyl, mercapto, hydroxyl, ester (e.g., -C) 1-6Alkyl-C(=O)-OC 1-6 Alkyl), C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Alkyl-OC 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-10 membered heterocyclic, 5-10 membered heteroaryl, C 6-10 Aryl, benzyl, hydroxy-substituted benzyl, indolylmethylene and C 1-6 Halogenated alkoxy groups, etc.

[0108] If a substituent is described as “optionally substituted with…”, then the substituent may be (1) unsubstituted or (2) substituted. If the carbon of the substituent is described as being optionally substituted with one or more of the substituents in the list, then one or more hydrogens on the carbon (to the extent that any hydrogens are present) may be substituted individually and / or together with independently selected substituents or not substituted. If the nitrogen of the substituent is described as being optionally substituted with one or more of the substituents in the list, then one or more hydrogens on the nitrogen (to the extent that any hydrogens are present) may each be substituted with independently selected substituents or not substituted.

[0109] If a substituent is described as being "independently selected" from a group of groups, then each substituent is selected independently of the others. Therefore, each substituent may be the same as or different from another (other) substituent.

[0110] As used herein, the term "one or more" means one or more under reasonable conditions, such as two, three, four, five, six, seven, eight, nine, or ten.

[0111] Unless otherwise specified, as used herein, the connection point of a substituent may be located at any suitable position of the substituent.

[0112] When the bond of a substituent is such that it passes through the ring and connects two atoms, then such a substituent can be bonded to any cyclic atom in the substituted ring.

[0113] This disclosure also includes all pharmaceutically acceptable isotopically labeled compounds identical to those of this disclosure, except that one or more atoms are replaced by atoms having the same atomic number but with an atomic mass or mass number different from the dominant atomic mass or mass number in nature. Examples of isotopes suitable for inclusion in the compounds of this disclosure include, but are not limited to, isotopes of hydrogen (e.g., 2 H, 3 H, deuterium (D), tritium (T); carbon isotopes (e.g., H, deuterium (D), tritium (T)); 11 C 13 C and14 C); isotopes of chlorine (e.g. 37 Cl); isotopes of fluorine (e.g., Cl); 18 F); isotopes of iodine (e.g., F); 123 I and 125 I); nitrogen isotopes (e.g.) 13 N and 15 N); isotopes of oxygen (e.g., N); 15 O、 17 O and 18 O); isotopes of phosphorus (e.g., O); phosphorus isotopes (e.g., O); 32 P); and isotopes of sulfur (e.g., ... 35 S). Certain isotope-labeled compounds of this disclosure (e.g., those doped with radioisotopes) can be used in drug and / or substrate tissue distribution studies (e.g., analysis). Radioisotope tritium (i.e. 3 H) and carbon-14 (i.e. 14 C) It is particularly suitable for this purpose due to its ease of incorporation and detection. Using positron-emitting isotopes (e.g.) 11 C 18 F, 15 O and 13 Substitution of N) can be used in positron emission tomography (PET) studies to examine substrate acceptor occupancy. Isotopically labeled compounds of this disclosure can be prepared by methods similar to those described in the accompanying routes and / or examples and preparations, by replacing previously used unlabeled reagents with appropriate isotopically labeled reagents. Pharmaceutically acceptable solvates of this disclosure include those in which the crystallization solvent can be isotopically substituted, for example, D2O, acetone-d6, or DMSO-d6.

[0114] The term "stereoisomer" refers to an isomer formed due to at least one asymmetric center. In compounds having one or more (e.g., 1, 2, 3, or 4) asymmetric centers, racemic mixtures, single enantiomers, diastereomer mixtures, and individual diastereomers can be produced. Specific individual molecules can also exist as geometric isomers (cis / trans). Similarly, the compounds of this disclosure can exist as mixtures of two or more structurally different forms in rapid equilibrium (commonly referred to as tautomers). Representative examples of tautomers include keto-enol tautomers, phenol-keto tautomers, nitroso-oxime tautomers, imine-enamine tautomers, etc. It is to be understood that the scope of this application covers all such isomers or mixtures thereof in any proportion (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%).

[0115] This disclosure covers all possible crystalline forms or polymorphs of the compounds disclosed herein, which may be a single polymorph or a mixture of more than one polymorph in any proportion.

[0116] It should also be understood that certain compounds of this disclosure may exist in their free form for therapeutic purposes, or, where appropriate, in their pharmaceutically acceptable derivative forms. In this disclosure, pharmaceutically acceptable derivatives include, but are not limited to, pharmaceutically acceptable salts, solvates, metabolites, or prodrugs that, upon administration to a patient in need, can directly or indirectly provide the compounds of this disclosure or their metabolites or residues. Therefore, when referring herein to “compounds of this disclosure,” it is also intended to encompass the various derivative forms of the compounds described above.

[0117] Pharmaceutically acceptable salts of the compounds disclosed herein include their acid addition salts and base addition salts. Suitable acid addition salts are formed by acids that form pharmaceutically acceptable salts. Suitable base addition salts are formed by bases that form pharmaceutically acceptable salts. A review of suitable salts can be found in Stahl and Wermuth's "Handbook of Pharmaceutical Salts: Properties, Selection, and Use" (Wiley-VCH, 2002). Methods for preparing pharmaceutically acceptable salts of the compounds disclosed herein are known to those skilled in the art.

[0118] The compounds disclosed herein may exist as solvates (preferably hydrates), wherein the compounds of this disclosure contain a polar solvent as a structural element of the compound's crystal lattice. The amount of the polar solvent, particularly water, may be stoichiometric or non-stoichiometric.

[0119] Those skilled in the art will understand that not all nitrogen-containing heterocycles can form N-oxides because nitrogen requires available lone pairs of electrons to be oxidized into oxides; those skilled in the art will identify nitrogen-containing heterocycles that can form N-oxides. Those skilled in the art will also recognize that tertiary amines can form N-oxides. Synthetic methods for preparing N-oxides of heterocycles and tertiary amines are well known to those skilled in the art, including the oxidation of heterocycles and tertiary amines with peroxy acids such as peracetic acid and m-chloroperoxybenzoic acid (MCPBA), hydrogen peroxide, alkyl peroxides such as tert-butyl peroxide, sodium perborate, and dioxiranes such as dimethyldioxirane. These methods for preparing N-oxides have been extensively described and reviewed in the literature, see, for example: T.L. Gilchrist, Comprehensive Organic Synthesis, vol. 7, pp. 748-750; A.R. Katritzky and A.J. Boulton, Eds., Academic Press; and G.W. H. Heeseman and E.S. G. Wierstiuk, Advances in Heterocyclic Chemistry, vol. 22, pp. 390-392, A.R. Katritzky and A.J. Boulton, Eds., Academic Press.

[0120] The scope of this disclosure also includes metabolites of the compounds of this disclosure, i.e., substances formed in the body upon administration of the compounds of this disclosure. Such products can be generated, for example, by oxidation, reduction, hydrolysis, amidation, deamidation, esterification, enzymatic hydrolysis, etc., of the administered compound. Therefore, this disclosure includes metabolites of the compounds of this disclosure, including compounds prepared by methods that expose the compounds of this disclosure to mammals for a time sufficient to produce their metabolites.

[0121] This disclosure further includes, within its scope, prodrugs of the compounds of this disclosure, which are certain derivatives of the compounds of this disclosure that may themselves have little or no pharmacological activity, and which, when administered to or onto the body, can be converted, for example, by hydrolysis and cleavage into the compounds of this disclosure having the desired activity. Typically, such prodrugs are functional group derivatives of the compounds that are readily converted in vivo into the compounds with the desired therapeutic activity. Further information regarding the use of prodrugs can be found in “Pro-drugs as Novel Delivery Systems,” Vol. 14, ACS Symposium Series (T. Higuchi and V. Stella) and “Bioreversible Carriers in Drug Design,” Pergamon Press, 1987 (EB Roche, editor, American Pharmaceutical Association). Prodrugs of this disclosure can be prepared, for example, by replacing suitable functional groups present in the compounds of this disclosure with certain portions known to those skilled in the art as “pro-moiety” (e.g., as described in “Design of Prodrugs,” H. Bundgaard (Elsevier, 1985)).

[0122] This disclosure also covers compounds of this disclosure containing protecting groups. In any process of preparing the compounds of this disclosure, protection of sensitive or reactive groups on any relevant molecule may be necessary and / or desired, thereby forming a form of chemical protection for the compounds of this disclosure. This can be achieved by conventional protecting groups, for example, those described in *Protective Groups in Organic Chemistry*, ed. J.F.W. McOmie, Plenum Press, 1973; and T.W. Greene & P. ​​G.W. M. Uts, *Protective Groups in Organic Synthesis*, John Wiley & Sons, 1991, which are incorporated herein by reference. Protecting groups can be removed at appropriate subsequent stages using methods known in the art.

[0123] In this document (specification and claims), the singular forms "a," "an," and "the" are used unless the context clearly indicates otherwise, including their plural forms, such as "a host cell" which includes multiple such host cells. Furthermore, there are no direct equivalent singular / plural grammatical rules in Chinese; the singular or plural form of a noun must be determined based on the context or actual situation. Therefore, in the English translation, adding "one or more than one" before a noun is likely correct.

[0124] Whether explicitly stated or not, all numerical values ​​in this application are modified by the term “about”. The term “about” means within ±20%, ±10%, ±5%, or ±2% of the stated numerical value.

[0125] Coupled

[0126] In one aspect, this disclosure provides a conjugate wherein the target portion Ab 1 The payload D is coupled to the load via a connecting unit. In some embodiments, the load D is the GSPT1 protein degrader portion.

[0127] In some embodiments, this disclosure provides compounds of formula (I) or pharmaceutically acceptable salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, isotopically labeled compounds, metabolites, or prodrugs thereof:

[0128] in,

[0129] Ab 1 For targeting groups;

[0130] M represents the junction site that connects to the target group;

[0131] L is the connector that connects M and D;

[0132] m is selected from 1-20;

[0133] D is selected from the structures of formulas (DI-1) and (DI-2);

[0134] X 1 Selected from CH2 and -C(=O)-;

[0135] X 2 X 3 X 4 Each is independently selected from O, S, Se, N, C, and CR. 3 And when X 2 X 3 X 4When one of the elements is independently selected from O, S, and Se, the other two are independently selected from N, C, and CR. 3 ;

[0136] It can be a single bond or a double bond;

[0137] X 5 Selected from O, S, C 1-6 Alkylene and C 1-6 Halogenated alkylene;

[0138] Ring A is selected from phenyl and 5-6-membered heteroaryl groups;

[0139] Y is selected from C 6-10 Aryl and 5-10 heteroaryl groups, wherein the aryl and heteroaryl groups are optionally separated by one or more R groups. 8 replace;

[0140] Z is selected from C 6-10 Aryl and 5-10 heteroaryl groups, said aryl and heteroaryl groups being R 7 While replacing, it can be optionally replaced by one or more R 9 replace;

[0141] L 1 Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Haloalkylene -, -O-(CH2) e -、-N(R a )-(CH2) e -、-(CH2-CH2-O) f -、C 3-10 Cycloalkyl groups, 3-12 membered heterocyclic groups, and divalent structures composed of one or more of the above structural units;

[0142] R 1 Selected from C 6-10 Aryl and 5-10 heteroaryl, said aryl and heteroaryl being -L a -L b -L c -R 5 While replacing, it can be optionally replaced by one or more R 4 replace;

[0143] R 2 Selected from H and C 1-6 alkyl;

[0144] R 3 Selected from H, halogen, hydroxyl, cyano, C 1-6 Alkyl, C 1-6 Halogenated alkyl groups and -OC1-6 alkyl;

[0145] R 4 Each is independently selected from H, halogen, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 6 replace;

[0146] R 5 Selected from

[0147] R 6 Each is independently selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Halogenated alkyl groups, -OC 1-6 Alkyl, C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more groups selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2- 6-acetylinyl, C 1-6 Haloalkyl, C 1-6 Alkoxy and C 1-6 Substituents of haloalkoxy groups;

[0148] L a Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Halogenated alkylene groups -, -(CH2) e -O-、-(CH2) e -N(R a )-、-(CH2-CH2-O) f -、-(CH2-CH2-O) f -(CH2) e -N(R a )-、-(CH2) e -O-(CH2) e -N(R a- and -(CH2) e -C(=O)-O-;

[0149] L b Selected from covalent bonds, C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 Aryl and 5-10 heteroaryl groups;

[0150] L c Selected from covalent bonds, -O-, -S-, -S(=O)-, -S(=O)2-, -N-(R a )-、-C(=O)N(R a )-、-C(=S)N(R a )-, -C(=O)-, -C(=S)-, -C(=O)-O-, -OC(=O)-O-, -OC(=O)-N(R a )-、-N(R a )-C(=O)-N(R a )-、-S(=O)2-N(R a )-, -S(=O)2-NH-C(=O)-O- and -S(=O)2-NH-C(=O)-N(R a )-;

[0151] R 8 Each is independently selected from H, halogen, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b -OR a -C 1-6 Alkylene-OR a -C 1-6 Alkylene-N(R) a )R b C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 10 replace;

[0152] R 10 Each is independently selected from H, halogen, cyano, and C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b and -ORa ;

[0153] R 7 Selected from

[0154] R 9 Each is independently selected from H, halogen, cyano, and C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b -OR a C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 11 replace;

[0155] R 11 Each is independently selected from H, halogen, cyano, and C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b and -OR a ;

[0156] R a Each is independently selected from H and C. 1-6 alkyl;

[0157] R b Each is independently selected from H and C. 1-6 alkyl;

[0158] z is selected from 1, 2, and 3;

[0159] f are each independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10;

[0160] e is independently selected from 0, 1, 2, 3, 4, 5, and 6;

[0161] In this context, position 1 of each substituent indicates the connection point with L.

[0162] In some embodiments, in the conjugate, the cellular active molecule portion D can be linked to the target portion via the "-ML-" structure shown in this application.

[0163] In some implementations, m is selected from 1 to 10, such as 1 to 8 or 4 to 8.

[0164] In some implementations, M is selected from the following structures:

[0165] Where, the 2nd position of M is related to Ab 1 Connected, position 3 is connected to L; each a is independently selected from 1, 2, 3, 4, 5 and 6, and b is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10.

[0166] In some implementations, M is selected from the following structures:

[0167] In some implementations, M is

[0168] In some embodiments, L is selected from non-breakable linkers and breakable linkers, wherein the breakable linker is cleaved by an enzyme present in the pathological environment, wherein the enzyme is selected from proteases, phosphatases, pyrophosphatases, β-glucuronidase, β-galactosidase, and sulfatases.

[0169] In some implementations, L is -L 2 -L 3 -L 4 -,in:

[0170] L 2 It is a covalent bond or selected from the following structures:

[0171] R 12 Selected from H and -L 2a -L 2b -(Q) n ,in;

[0172] L 2a It is a covalent bond or selected from the following structures:

[0173] L 2b It is a covalent bond or selected from the following structures:

[0174] Q is a covalent bond or selected from the following structures:

[0175] Where n is selected from 1, 2 and 3; each c is independently selected from 1, 2, 3, 4, 5 and 6; and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10.

[0176] L 3Fragments selected from amino acids or their derivatives and peptide fragments formed from two or more amino acids or their derivatives, wherein the amino acids are selected from Val, Cit, Glu, Lys, Arg, Phe, Leu, Gly, Ala, and Asn, and the amino acid derivatives are selected from... R' is independently selected from hydrogen and C. 1-6 Alkyl, -C 1-6 Alkylene-OC 1-6 Alkyl, -C 1-6 Alkylene-NHC 1-6 Alkyl, -C 1-6 Alkylene-N(C) 1-6 Alkyl)2;

[0177] L 4 It is a covalent bond or selected from the following structures:

[0178] In some implementations, L is -L 2 -L 3 -L 4 -,in:

[0179] L 2 It is a covalent bond or selected from the following structures:

[0180] R 12 Selected from H and -L 2a -L 2b -(Q) n ,in;

[0181] L 2a It is a covalent bond or selected from the following structures:

[0182] L 2b It is a covalent bond or selected from the following structures:

[0183] Q is a covalent bond or selected from the following structures:

[0184] Where n is selected from 1, 2 and 3; each c is independently selected from 1, 2, 3, 4, 5 and 6; and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10.

[0185] L 3 Fragments selected from amino acids or their derivatives and peptide fragments formed from two or more amino acids or their derivatives, wherein the amino acids are selected from Val, Cit, Glu, Lys, Arg, Phe, Leu, Gly, Ala, and Asn, and the amino acid derivatives are selected from... Where R' is selected from hydrogen, C 1-6 Alkyl, -C 1-6 Alkylene-OC 1-6 Alkyl, -C 1-6 Alkylene-NHC 1-6 Alkyl, -C 1-6 Alkylene-N(C) 1-6 Alkyl)2;

[0186] L 4 It is a covalent bond or selected from the following structures:

[0187] In some implementations, L 2 It is a covalent bond or selected from the following structures:

[0188] In some implementations, L 2 It is a covalent bond or selected from the following structures:

[0189] In some implementations, L 2a It is a covalent bond or selected from the following structures:

[0190] Each 'c' is independently selected from 1, 2, 3, 4, 5, and 6, and each 'd' is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.

[0191] In some implementations, L 2b Covalent bond or selected from: Each 'c' is independently selected from 1, 2, 3, 4, 5, and 6, and each 'd' is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.

[0192] In some implementations, L 2b Covalent bond or selected from: Each 'c' is independently selected from 1, 2, 3, 4, 5, and 6, and each 'd' is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.

[0193] In some implementations, L 2b Covalent bond or selected from: Position 6 is related to L 2a The 7th position is connected to Q, each c is independently selected from 1, 2, 3, 4, 5 and 6, and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10.

[0194] In some implementations, L 2bCovalent bond or selected from: Position 6 is related to L 2a The 7th position is connected to Q, each c is independently selected from 1, 2, 3, 4, 5 and 6, and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10.

[0195] In some implementations, Q is a covalent bond or selected from the following structures:

[0196] In some implementations, Q is selected from the following structures:

[0197] In some implementations, Q is a covalent bond or selected from the following structures:

[0198] In some implementations, -L 2a -L 2b -(Q) n Selected from the following structures:

[0199] In some implementations, L 2 It is a covalent bond or selected from the following structures: In this context, each 'c' is independently selected from 1, 2, 3, 4, 5, and 6, and each 'd' is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.

[0200] In some implementations, L 2 It is a covalent bond or selected from the following structures: In this context, each 'c' is independently selected from 1, 2, 3, 4, 5, and 6, and each 'd' is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.

[0201] In some implementations, L 3 Selected from the following structures:

[0202] In some implementations, L 3 Selected from the following structures:

[0203] In some implementations, L 3 Selected from the following structures:

[0204] In some implementations, L 4 For covalent bonds or

[0205] In some implementations, L is selected from the following structures:

[0206] In this case, position 4 of L is connected to M, and position 5 is connected to D; each c is independently selected from 1, 2, 3, 4, 5 and 6, and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10.

[0207] In some implementations, L is selected from the following structures:

[0208] In this case, position 4 of L is connected to M, and position 5 is connected to D; each c is independently selected from 1, 2, 3, 4, 5 and 6, and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10.

[0209] In some implementations, L is selected from the following structures:

[0210] In this case, position 4 of L is connected to M, and position 5 is connected to D.

[0211] In some implementations, L is selected from the following structures:

[0212] In this case, position 4 of L is connected to M, and position 5 is connected to D.

[0213] In some embodiments, D is the remainder obtained after the GSPT1 protein degrader molecule loses one or more atoms or groups of atoms (e.g., hydrogen or hydroxyl groups).

[0214] In some embodiments, D is a monovalent structure obtained by losing an H from the -OH, -NH2, or secondary amino group on the GSPT1 protein degrader.

[0215] In some embodiments, the GSPT1 protein degrader is described in CN202511226632.4 and PCT / CN2025 / 132988, the contents of which are incorporated herein by reference.

[0216] In some embodiments, D is selected from the structures of formulas (D-II-1), (D-II-2), (D-II-3), (D-II-4), (D-II-5), (D-II-6), (D-II-7), (D-II-8), (D-II-9), and (D-II-10):

[0217] Among them, X 2 X3 X 4 Each is independently selected from O, S, and Se; X 5 R 1 R 2 R 3 Y, Z, L 1 And z as defined in any of the preceding items.

[0218] In some implementations, D is selected from the structures of formulas (D-IV-1), (D-IV-2), and (D-IV-3):

[0219] Among them, X 5 R 1 R 2 R 3 Y, Z, L 1 One or more of them are each independently defined as in any of the preceding items;

[0220] Optionally, X 5 Selected from O, S, C 1-6 Alkylene and C 1-6 Halogenated alkylene;

[0221] Y is selected from C 6-10 Aryl and 5-10 heteroaryl groups, wherein the aryl and heteroaryl groups are optionally separated by one or more R groups. 8 replace;

[0222] Z is selected from C 6-10 Aryl and 5-10 heteroaryl groups, said aryl and heteroaryl groups being R 7 While replacing, it can be optionally replaced by one or more R 9 replace;

[0223] L 1 Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Haloalkylene -, -O-(CH2) e -、-N(R a )-(CH2) e -、-(CH2-CH2-O) f -、C 3-10 Cycloalkyl groups, 3-12 membered heterocyclic groups, and divalent structures composed of one or more of the above structural units;

[0224] R 1 Selected from C 6-10 Aryl and 5-10 heteroaryl, said aryl and heteroaryl being -L a -L b -Lc -R 5 While replacing, it can be optionally replaced by one or more R 4 replace;

[0225] R 2 Selected from H and C 1-4 alkyl;

[0226] R 3 Selected from H, halogen, hydroxyl, cyano, C 1-4 Alkyl, C 1-4 Halogenated alkyl groups and -OC 1-4 alkyl;

[0227] R 4 Each is independently selected from H, halogen, amino, C 1-6 Alkyl, 3-10 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 6 replace;

[0228] R 5 Selected from

[0229] R 6 Each is independently selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-6 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Halogenated alkyl groups, -OC 1-6 Alkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more groups selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-4 Alkyl, C 2-4 alkenyl, C 2- 4-Alynyl group, C 1-4 Haloalkyl, C 1-4 Alkoxy and C 1-4 Substituents of haloalkoxy groups;

[0230] L a Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Halogenated alkylene groups -, -(CH2) e -O-、-(CH2) e -N(R a )-、-(CH2-CH2-O) f-、-(CH2-CH2-O) f -(CH2) e -N(R a - and -(CH2) e -O-(CH2) e -N(R a )-;

[0231] L b Selected from covalent and 3-10 member heterocyclic groups;

[0232] L c Selected from covalent bonds, -O-, -S-, -S(=O)-, -S(=O)2-, -N-(R a )-、-C(=O)N(R a )-, -C(=O)-, -C(=O)-O-, -S(=O)2-N(R a )-and -S(=O)2-NH-C(=O)-O-;

[0233] R 8 Each is independently selected from H, halogen, cyano, nitro, C 1-4 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Haloalkyl, -N(R) a )R b -OR a -C 1-4 Alkylene-OR a -C 1-4 Alkylene-N(R) a )R b C 3-8 Cycloalkyl, 3-10 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 10 replace;

[0234] R 10 Each is independently selected from H, halogen, cyano, and C. 1-4 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Haloalkyl, -N(R) a )R b and -OR a ;

[0235] R 7 Selected from

[0236] R 9Each is independently selected from H, halogen, cyano, and C. 1-4 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Haloalkyl, -N(R) a )R b -OR a C 3-8 Cycloalkyl, 3-10 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 11 replace;

[0237] R 11 Each is independently selected from H, halogen, cyano, and C. 1-4 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Haloalkyl, -N(R) a )R b and -OR a ;

[0238] R a Each is independently selected from H and C. 1-4 alkyl;

[0239] R b Each is independently selected from H and C. 1-4 alkyl;

[0240] f are each independently selected from 1, 2, 3, 4, 5, and 6;

[0241] e is independently selected from 0, 1, 2, 3, 4 and 5;

[0242] In this context, position 1 of each substituent indicates the connection point with L.

[0243] In some embodiments of formulas (DI-1), (DI-2), (D-II-1), (D-II-2), (D-II-3), (D-II-4), (D-II-5), (D-II-6), (D-II-7), (D-II-8), (D-II-9), and / or (D-II-10), X 1 X 2 X 3 X 4 X 5 R 1 R 2 R 3 Y, Z, L 1 ,z,R 4 R5 R 6 R 7 R 8 R 9 R 10 R 11 R a R b One or more of them are as defined above for equations (D-IV-1) and (D-IV-2).

[0244] In some embodiments, D is selected from the structures of formulas (D-IV-1) and (D-IV-2) above. In some embodiments, D is selected from the following structures.

[0245] In some implementations, D is selected from the following structures In some implementation schemes, Ab 1 It is a targeting group that targets cell surface receptors or tumor surface antigens.

[0246] In some implementation schemes, Ab 1 The antibody is an antibody or its antigen-binding fragment; the antibody is selected from monoclonal antibodies, polyclonal antibodies, linear antibodies, bispecific antibodies, multispecific antibodies, chimeric antibodies, murine antibodies, humanized antibodies, fully human antibodies, and fusion proteins containing the antigen-binding portion of the antibody; the antigen-binding fragment is selected from Fab, Fab', F(ab')2, Fv, disulfide-linked Fv and scFv.

[0247] In Ab 1 When the substance is an antibody or its antigen-binding fragment, the compound represented by Formula I is a degradation agent antibody conjugate.

[0248] In some implementation schemes, Further for Where Ab-(W- is Ab) 1 , represents an antibody or its antigen-binding fragment, and -(W- represents the linking portion of the amino acid residues in the antibody or its antigen-binding fragment to M.

[0249] In some embodiments, the amino acid residue is a cysteine, lysine, serine, or threonine residue.

[0250] In some implementations, W is S, NH, or O.

[0251] In some embodiments, the antibody or its antigen-binding fragment is an anti-Her-2 antibody or its antigen-binding fragment.

[0252] In some embodiments, the anti-Her-2 antibody or its antigen-binding fragment comprises:

[0253] (1) The following heavy chain variable regions (VH) and / or light chain variable regions (VL):

[0254] (1a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:28 or a variant thereof, CDR-H2 with sequence SEQ ID NO:29 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or,

[0255] (1b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:43 or a variant thereof, CDR-H2 with sequence SEQ ID NO:44 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof;

[0256] or,

[0257] (2) The following heavy chain variable regions (VH) and / or light chain variable regions (VL):

[0258] (2a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence SEQ ID NO:41 or a variant thereof, CDR-H2 with the sequence SEQ ID NO:42 or a variant thereof, and CDR-H3 with the sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with the sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with the sequence SEQ ID NO:33 or a variant thereof; or,

[0259] (2b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:56 or a variant thereof, CDR-H2 with sequence SEQ ID NO:57 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof;

[0260] or,

[0261] (3) The following heavy chain variable regions (VH) and / or light chain variable regions (VL):

[0262] (3a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:34 or a variant thereof, CDR-H2 with sequence SEQ ID NO:35 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or,

[0263] (3b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:49 or a variant thereof, CDR-H2 with sequence SEQ ID NO:50 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof;

[0264] or,

[0265] (4) The following heavy chain variable regions (VH) and / or light chain variable regions (VL):

[0266] (4a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:36 or a variant thereof, CDR-H2 with sequence SEQ ID NO:37 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:38 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:39 or a variant thereof, CDR-L2 with sequence SEQ ID NO:40 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or,

[0267] (4b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:51 or a variant thereof, CDR-H2 with sequence SEQ ID NO:52 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:53 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:54 or a variant thereof, CDR-L2 with sequence SEQ ID NO:55 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof;

[0268] The variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it originates, or the variant has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to the sequence from which it originates; preferably, the substitutions are conservative substitutions.

[0269] In some embodiments, the anti-Her-2 antibody or its antigen-binding fragment comprises:

[0270] (1) The following heavy chain variable regions (VH) and / or light chain variable regions (VL), wherein the CDR is defined according to the Chothia numbering system:

[0271] (1a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:28 or a variant thereof, CDR-H2 with sequence SEQ ID NO:29 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or,

[0272] (1b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:43 or a variant thereof, CDR-H2 with sequence SEQ ID NO:44 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof;

[0273] or,

[0274] (2) The following heavy chain variable regions (VH) and / or light chain variable regions (VL), wherein the CDR is defined according to the AbM numbering system:

[0275] (2a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence SEQ ID NO:41 or a variant thereof, CDR-H2 with the sequence SEQ ID NO:42 or a variant thereof, and CDR-H3 with the sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with the sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with the sequence SEQ ID NO:33 or a variant thereof; or,

[0276] (2b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:56 or a variant thereof, CDR-H2 with sequence SEQ ID NO:57 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof;

[0277] or,

[0278] (3) The following heavy chain variable regions (VH) and / or light chain variable regions (VL), wherein the CDR is defined according to the Kabat numbering system:

[0279] (3a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:34 or a variant thereof, CDR-H2 with sequence SEQ ID NO:35 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or,

[0280] (3b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:49 or a variant thereof, CDR-H2 with sequence SEQ ID NO:50 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof;

[0281] or,

[0282] (4) The following heavy chain variable regions (VH) and / or light chain variable regions (VL), wherein the CDR is defined according to the IMGT numbering system:

[0283] (4a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:36 or a variant thereof, CDR-H2 with sequence SEQ ID NO:37 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:38 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:39 or a variant thereof, CDR-L2 with sequence SEQ ID NO:40 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or,

[0284] (4b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:51 or a variant thereof, CDR-H2 with sequence SEQ ID NO:52 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:53 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:54 or a variant thereof, CDR-L2 with sequence SEQ ID NO:55 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof;

[0285] The variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it originates, or the variant has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to the sequence from which it originates; preferably, the substitutions are conservative substitutions.

[0286] In some embodiments, the anti-Her-2 antibody or its antigen-binding fragment comprises:

[0287] (a) VH or a variant thereof shown in SEQ ID NO: 24, and / or VL or a variant thereof shown in SEQ ID NO: 25; or

[0288] (b) VH or a variant thereof shown in SEQ ID NO: 26, and / or VL or a variant thereof shown in SEQ ID NO: 27;

[0289] The variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it originates, or the variant has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to the sequence from which it originates; preferably, the substitutions are conservative substitutions.

[0290] In some embodiments, the anti-Her-2 antibody or its antigen-binding fragment comprises:

[0291] (1) A heavy chain comprising the VH region of the sequence shown in SEQ ID NO: 24 and the heavy chain constant region (CH) shown in SEQ ID NO: 58, and a light chain comprising the VL region of the sequence shown in SEQ ID NO: 25 and the light chain constant region (CL) shown in SEQ ID NO: 59; or

[0292] (2) A heavy chain including the VH of the sequence shown in SEQ ID NO: 26 and the heavy chain constant region (CH) shown in SEQ ID NO: 58, and a light chain including the VL of the sequence shown in SEQ ID NO: 27 and the light chain constant region (CL) shown in SEQ ID NO: 59.

[0293] In some embodiments, the anti-Her-2 antibody or its antigen-binding fragment comprises:

[0294] (1) The heavy chain comprising the sequence shown in SEQ ID NO: 60, and the light chain comprising the sequence shown in SEQ ID NO: 61; or

[0295] (2) The heavy chain comprising the sequence shown in SEQ ID NO: 62, and the light chain comprising the sequence shown in SEQ ID NO: 63.

[0296] In some embodiments, the N-terminal glutamine of the VH or variant thereof, as shown in SEQ ID NO:24 or 26, or the heavy chain or variant thereof, as shown in SEQ ID NO:60 or 62, undergoes cyclization to form pyroglutamic acid or pyroglutamic acid salt.

[0297] In some embodiments, the heavy chain constant region (CH) of the sequence shown in SEQ ID NO: 58 or a variant thereof, or the heavy chain of the sequence shown in SEQ ID NO: 60 or 62 or a variant thereof, lacks a C-terminal lysine residue.

[0298] In some embodiments, the anti-Her-2 antibody is a trastuzumab antibody, a pertuzumab antibody, or a dual epitope antibody composed of these, wherein the amino acid sequence of the trastuzumab antibody has an IMGT / mAb-DB ID of 97 and the amino acid sequence of the pertuzumab antibody has an IMGT / mAb-DB ID of 80.

[0299] In some embodiments, the anti-Her-2 antibody is trastuzumab.

[0300] In some embodiments, the anti-Her-2 antibody is pertuzumab.

[0301] In some embodiments, the antibody or its antigen-binding fragment is an anti-B7H3 antibody or its antigen-binding fragment.

[0302] In some embodiments, the anti-B7H3 antibody or its antigen-binding fragment comprises:

[0303] (1) The following heavy chain variable regions (VH) and / or light chain variable regions (VL):

[0304] (1a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:11 or a variant thereof, CDR-H2 with sequence SEQ ID NO:12 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof; or

[0305] (1b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence of SEQ ID NO:11 or a variant thereof, CDR-H2 with the sequence of SEQ ID NO:68 or a variant thereof, and CDR-H3 with the sequence of SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence of SEQ ID NO:6 or a variant thereof, CDR-L2 with the sequence of SEQ ID NO:7 or a variant thereof, and CDR-L3 with the sequence of SEQ ID NO:5 or a variant thereof;

[0306] or

[0307] (2) The following heavy chain variable regions (VH) and / or light chain variable regions (VL):

[0308] (2a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO:14 or a variant thereof, CDR-H2 of SEQ ID NO:15 or a variant thereof, and CDR-H3 of SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO:6 or a variant thereof, CDR-L2 of SEQ ID NO:7 or a variant thereof, and CDR-L3 of SEQ ID NO:5 or a variant thereof; or

[0309] (2b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:14 or a variant thereof, CDR-H2 with sequence SEQ ID NO:69 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof;

[0310] or

[0311] (3) The following heavy chain variable regions (VH) and / or light chain variable regions (VL):

[0312] (3a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO:8 or a variant thereof, CDR-H2 of SEQ ID NO:9 or a variant thereof, and CDR-H3 of SEQ ID NO:10 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO:3 or a variant thereof, CDR-L2 of SEQ ID NO:4 or a variant thereof, and CDR-L3 of SEQ ID NO:5 or a variant thereof; or

[0313] (3b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:8 or a variant thereof, CDR-H2 with sequence SEQ ID NO:67 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:10 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:3 or a variant thereof, CDR-L2 with sequence SEQ ID NO:4 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof;

[0314] or

[0315] (4) The following heavy chain variable regions (VH) and / or light chain variable regions (VL):

[0316] (4a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:16 or a variant thereof, CDR-H2 with sequence SEQ ID NO:17 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof; or

[0317] (4b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence of SEQ ID NO:16 or a variant thereof, CDR-H2 with the sequence of SEQ ID NO:70 or a variant thereof, and CDR-H3 with the sequence of SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence of SEQ ID NO:6 or a variant thereof, CDR-L2 with the sequence of SEQ ID NO:7 or a variant thereof, and CDR-L3 with the sequence of SEQ ID NO:5 or a variant thereof.

[0318] In some embodiments, the anti-B7H3 antibody or its antigen-binding fragment comprises:

[0319] (1) The following heavy chain variable regions (VH) and / or light chain variable regions (VL), wherein the CDR is defined according to the Chothia numbering system:

[0320] (1a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:11 or a variant thereof, CDR-H2 with sequence SEQ ID NO:12 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof; or

[0321] (1b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence of SEQ ID NO:11 or a variant thereof, CDR-H2 with the sequence of SEQ ID NO:68 or a variant thereof, and CDR-H3 with the sequence of SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence of SEQ ID NO:6 or a variant thereof, CDR-L2 with the sequence of SEQ ID NO:7 or a variant thereof, and CDR-L3 with the sequence of SEQ ID NO:5 or a variant thereof;

[0322] or

[0323] (2) The following heavy chain variable regions (VH) and / or light chain variable regions (VL), wherein the CDR is defined according to the Kabat numbering system:

[0324] (2a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO:14 or a variant thereof, CDR-H2 of SEQ ID NO:15 or a variant thereof, and CDR-H3 of SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO:6 or a variant thereof, CDR-L2 of SEQ ID NO:7 or a variant thereof, and CDR-L3 of SEQ ID NO:5 or a variant thereof; or

[0325] (2b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:14 or a variant thereof, CDR-H2 with sequence SEQ ID NO:69 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof;

[0326] or

[0327] (3) The following heavy chain variable regions (VH) and / or light chain variable regions (VL), wherein the CDR is defined according to the IMGT numbering system:

[0328] (3a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO:8 or a variant thereof, CDR-H2 of SEQ ID NO:9 or a variant thereof, and CDR-H3 of SEQ ID NO:10 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO:3 or a variant thereof, CDR-L2 of SEQ ID NO:4 or a variant thereof, and CDR-L3 of SEQ ID NO:5 or a variant thereof; or

[0329] (3b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:8 or a variant thereof, CDR-H2 with sequence SEQ ID NO:67 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:10 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:3 or a variant thereof, CDR-L2 with sequence SEQ ID NO:4 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof;

[0330] or

[0331] (4) The following heavy chain variable regions (VH) and / or light chain variable regions (VL), wherein the CDR is defined according to the AbM numbering system:

[0332] (4a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:16 or a variant thereof, CDR-H2 with sequence SEQ ID NO:17 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof; or

[0333] (4b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence of SEQ ID NO:16 or a variant thereof, CDR-H2 with the sequence of SEQ ID NO:70 or a variant thereof, and CDR-H3 with the sequence of SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence of SEQ ID NO:6 or a variant thereof, CDR-L2 with the sequence of SEQ ID NO:7 or a variant thereof, and CDR-L3 with the sequence of SEQ ID NO:5 or a variant thereof.

[0334] In some embodiments, the anti-B7H3 antibody or its antigen-binding fragment comprises:

[0335] (a) VH or a variant thereof shown in SEQ ID NO:23 or 1, and / or VL or a variant thereof shown in SEQ ID NO:2; or

[0336] (b) VH or a variant thereof shown in SEQ ID NO:66, and / or VL or a variant thereof shown in SEQ ID NO:2.

[0337] In some embodiments, the antibody or its antigen-binding fragment comprises:

[0338] (a) The heavy chain of the VH or variant thereof shown in SEQ ID NO:23 or 1 and the heavy chain constant region (CH) or variant thereof shown in SEQ ID NO:18, and / or, the light chain of the VL or variant thereof shown in SEQ ID NO:2 and the light chain constant region (CL) or variant thereof shown in SEQ ID NO:19; or

[0339] (b) The heavy chain of the VH or variant thereof shown in SEQ ID NO:1 and the heavy chain constant region (CH) or variant thereof shown in SEQ ID NO:71, and / or the light chain of the VL or variant thereof shown in SEQ ID NO:2 and the light chain constant region (CL) or variant thereof shown in SEQ ID NO:19;

[0340] (c) The VH or variant thereof shown in SEQ ID NO:66 and the heavy chain of the heavy chain constant region (CH) or variant thereof shown in SEQ ID NO:71 or 18, and / or the VL or variant thereof shown in SEQ ID NO:2 and the light chain constant region (CL) or variant thereof shown in SEQ ID NO:19.

[0341] In some embodiments, the amino acid sequence of the heavy chain of the anti-B7H3 antibody or its antigen-binding fragment is as shown in SEQ ID NO:22 or 20, 72, 73 or 74, and the amino acid sequence of the light chain of the anti-B7H3 antibody or its antigen-binding fragment is as shown in SEQ ID NO:21.

[0342] In some embodiments, the anti-B7H3 antibody or its antigen-binding fragment comprises: VH or a variant thereof shown in SEQ ID NO:66, and / or, VL or a variant thereof shown in SEQ ID NO:2; preferably, the anti-B7H3 antibody or its antigen-binding fragment comprises: the heavy chain of the VH or a variant thereof shown in SEQ ID NO:66 and the heavy chain constant region (CH) or a variant thereof shown in SEQ ID NO:18, and / or, the light chain of the VL or a variant thereof shown in SEQ ID NO:2 and the light chain constant region (CL) or a variant thereof shown in SEQ ID NO:19; preferably, the amino acid sequence of the heavy chain of the anti-B7H3 antibody or its antigen-binding fragment is as shown in SEQ ID NO:74, and the amino acid sequence of the light chain of the antibody or its antigen-binding fragment is as shown in SEQ ID NO:21.

[0343] In some embodiments, the N-terminal glutamine of the heavy chain or heavy chain variable region and / or the light chain or light chain variable region undergoes cyclization to form pyroglutamic acid or pyroglutamate; and / or, the heavy chain or heavy chain constant region (CH) or its variants do not contain a C-terminal lysine.

[0344] In some embodiments, the variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with respect to its source sequence, or has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to its source sequence; preferably, the substitutions are conservative substitutions.

[0345] In some embodiments, the anti-B7H3 antibody or its antigen-binding fragment is Ab-01, Ab-02, Ab-03, Ab-04, or Ab-05.

[0346] In some embodiments, the anti-B7H3 antibody or its antigen-binding fragment is Ab-05.

[0347] In some embodiments, the compound represented by Formula I has the following structure:

[0348] Each m is selected from 1 to 20, for example 1 to 10, and Ab-(S-) is an antibody or its antigen-binding fragment as defined in any of the preceding items.

[0349] In some implementations, the S in Ab-(S-) originates from the thiol group in the antibody or its antigen-binding fragment.

[0350] In some embodiments, this disclosure provides compositions of degrading agent antibody conjugates comprising the degrading agent antibody conjugate as defined in any of the preceding embodiments, wherein the DAR value of the composition is 1.0-20.0, preferably 1.0-10.0, and the following values ​​are provided: about 1 to 2, about 1 to 3, about 1 to 4, about 1 to 5, about 1 to 6, about 1 to 7, about 1 to 8, about 1 to 9, about 1 to 10, about 2 to 3, about 2 to 4, about 2 to 5, about 2 to 6, about 2 to 7, about 2 to 8, about 2 to 9, about 2 to 10, about 3 to 4, about 3 to 5, about 3 to 6, about 3 to 7, about 3 to 6, about 3 to 7, about 3 to 8, about 4 to 9, about 4 to 10, about 5 to 6, about 5 to 7, about 5 to 10. 8, about 5 to 9, about 5 to 10, about 6 to 7, about 6 to 8, about 6 to 9, about 6 to 10, about 7 to 8, about 7 to 9, about 7 to 10, about 8 to 9, about 8 to 10 or about 9 to 10, preferably 4.0 to 8.0, for example, 4.0 to 4.5, 4.0 to 5.0, 4.0 to 5.5, 4.0 to 6.0, 4.0 to 6.5, 4.0 ~7.0, 4.0~7.5, 4.0~8.0, 5.0~5.5, 5.0~6.0, 5.0~6.5, 5.0~7.0, 5.0~7.5, 5.0~8.0, 5.5~6.0, 5.5~6.5, 5.5~7.0, 5.5~7.5, 5.5~8.0, 6.0~6.5, 6.0~7.0, 6.0–7.5, 6.0–8.0, 6.5–7.0, 6.5–7.5, 6.5–8.0, 7.0–7.5, 7.0–8.0, 7.5–8.0, for example, approximately 4.0, approximately 4.01, approximately 4.02, approximately 4.03, approximately 4.04, approximately 4.05, approximately 4.06, approximately 4.07, approximately 4.08, approximately 4.09 Approximately 4.1, 4.11, 4.12, 4.13, 4.14, 4.15, 4.16, 4.17, 4.18, 4.19, 4.2, 4.21, 4.22, 4.23, 4.24, 4.25, 4.26, 4.27, 4.28, 4.29, 4.3, approximately 4.31, approximately 4.32, approximately 4.33, approximately 4.34, approximately 4.35, approximately 4.36, approximately 4.37, approximately 4.38, approximately 4.39, approximately 4.4, approximately 4.41, approximately 4.42, approximately 4.43, approximately 4.44, approximately 4.45, approximately 4.46, approximately 4.47, approximately 4.48, approximately 4.49, approximately 4.5, approximately 4.51, approximately 4 .52, approximately 4.53, approximately 4.54, approximately 4.55, approximately 4.56, approximately 4.57, approximately 4.58, approximately 4.59, approximately 4.6, approximately 4.61, approximately 4.62, approximately 4.63, approximately 4.64, approximately 4.65, approximately 4.66, approximately 4.67, approximately 4.68, approximately 4.69, approximately 4.7, approximately 4.71, approximately 4.72, approximately 4.73, approximately 4.74, approximately 4.75, approximately 4.76, approximately 4.77, approximately 4.78, approximately 4.79, approximately 4.8, approximately 4.81, approximately 4.82, approximately 4.83, approximately 4.84, approximately 4.85, approximately 4.86, approximately 4.87, approximately 4.88, approximately 4.89, approximately 4.9, approximately 4.91, approximately 4.92, approximately 4.93, approximately 4. 94, approximately 4.95, approximately 4.96, approximately 4.97, approximately 4.98, approximately 4.99, approximately 5.0, approximately 5.01, approximately 5.02, approximately 5.03, approximately 5.04, approximately 5.05, approximately 5.06, approximately 5.07, approximately 5.08, approximately 5.09, approximately 5.1, approximately 5.11, approximately 5.12, approximately 5.13, approximately 5.14, approximately 5.1 5. Approximately 5.16, 5.17, 5.18, 5.19, 5.2, 5.21, 5.22, 5.23, 5.24, 5.25, 5.26, 5.27, 5.28, 5.29, 5.3, 5.31, 5.32, 5.33, 5.34, 5.35, 5.3 6. Approximately 5.37, 5.38, 5.39, 5.4, 5.41, 5.42, 5.43, 5.44, 5.45, 5.46, 5.47, 5.48, 5.49, 5.5, 5.51, 5.52, 5.53, 5.54, 5.55, 5.56, 5.57 Approximately 5.58, 5.59, 5.6, 5.61, 5.62, 5.63, 5.64, 5.65, 5.66, 5.67, 5.68, 5.69, 5.7, 5.71, 5.72, 5.73, 5.74, 5.75, 5.76, 5.77, 5.78 Approximately 5.79, approximately 5.8, approximately 5.81, approximately 5.82, approximately 5.83, approximately 5.84, approximately 5.85, approximately 5.86, approximately 5.87, approximately 5.88, approximately 5.89, approximately 5.9, approximately 5.91, approximately 5.92, approximately 5.93, approximately 5.94, approximately 5.95, approximately 5.96, approximately 5.97, approximately 5.98, approximately 5.99. Approximately 6.0, 6.01, 6.02, 6.03, 6.04, 6.05, 6.06, 6.07, 6.08, 6.09, 6.1, 6.11, 6.12, 6.13, 6.14, 6.15, 6.16, 6.17, 6.18, 6.19, 6.2, 6 .21, about 6.22, about 6.23, about 6.24, about 6.25, about 6.26, about 6.27, about 6.28, about 6.29, about 6.3, about 6.31, about 6.32, about 6.33, about 6.34, about 6.35, about 6.36, about 6.37, about 6.38, about 6.39, about 6.4, about 6.41, about 6.42, approximately 6.43, approximately 6.44, approximately 6.45, approximately 6.46, approximately 6.47, approximately 6.48, approximately 6.49, approximately 6.5, approximately 6.51, approximately 6.52, approximately 6.53, approximately 6.54, approximately 6.55, approximately 6.56, approximately 6.57, approximately 6.58, approximately 6.59, approximately 6.6, approximately 6.61, approximately 6.62, approximately 6.63, approximately 6.64, approximately 6.65, approximately 6.66, approximately 6.67, approximately 6.68, approximately 6.69, approximately 6.7, approximately 6.71, approximately 6.72, approximately 6.73, approximately 6.74, approximately 6.75, approximately 6.76, approximately 6.77, approximately 6.78, approximately 6.79, approximately 6.8, approximately 6.81, Approximately 6.82, 6.83, 6.84, 6.85, 6.86, 6.87, 6.88, 6.89, 6.9, 6.91, 6.92, 6.93, 6.94, 6.95, 6.96, 6.97, 6.98, 6.99, approximately 7.0, 7.01, 7.02, 7.03, 7.04, 7.05, 7.06, 7.07, 7.08, 7.09, 7.1, 7.11, 7.12, 7.13, 7.14, 7.15, 7.16, 7.17, 7.18, 7.19, 7.2, 7.2 1. Approximately 7.22, 7.23, 7.24, 7.25, 7.26, 7.27, 7.28, 7.29, 7.3, 7.31, 7.32, 7.33, 7.34, 7.35, 7.36, 7.37, 7.38, 7.39, 7.4, 7.41, 7.42, 7.43, 7.44, 7.45, 7.46, 7.47, 7.48, 7.49, 7.5, 7.51, 7.52, 7.53, 7.54, 7.55, 7.56, 7.57, 7.58, 7.59, 7.6, 7. .61, approximately 7.62, approximately 7.63, approximately 7.64, approximately 7.65, approximately 7.66, approximately 7.67, approximately 7.68, approximately 7.69, approximately 7.7, approximately 7.71, approximately 7.72, approximately 7.73, approximately 7.74, approximately 7.75, approximately 7.76, approximately 7.77, approximately 7.78, approximately 7.79, approximately 7.8, approximately 7.81, approximately 7.82, approximately 7.83, approximately 7.84, approximately 7.85, approximately 7.86, approximately 7.87, approximately 7.88, approximately 7.89, approximately 7.9, approximately 7.91, approximately 7.92, approximately 7.93, approximately 7.94, approximately 7.95, approximately 7.96, approximately 7.97, approximately 7.98, approximately 7.99, approximately 8.0.

[0351] In some embodiments, the DAR value of the degradation agent antibody conjugate as defined in any of the preceding embodiments is 1.0-20.0, preferably 1.0-10.0, about 1 to 2, about 1 to 3, about 1 to 4, about 1 to 5, about 1 to 6, about 1 to 7, about 1 to 8, about 1 to 9, about 1 to 10, about 2 to 3, about 2 to 4, about 2 to 5, about 2 to 6, about 2 to 7, about 2 to 8, about 2 to 9, about 2 to 10, about 3 to 4, about 3 to 5, about 3 to 6, about 3 to 7, about 3 to 8, about 3 to 9, about 3 to 10, about 4 to 5, about 4 to 6, about 4 to 7, about 4 to 8, about 4 to 9, about 4 to 10, about 5 to 6, about 5 to 7, about 5 to 8, about 5 to 9, about 5 to 10, about 6 to 7, about 6 to 8, about 6 to 9, about 6 to 10, about 7 to 8, about 7 to 9, about 7 to 10, about 8 to 9, about 8 to 10 or about 9 to 10, preferably 4.0 to 8.0, for example, 4.0 to 4.5, 4.0 to 5.0, 4.0 to 5.5, 4.0 to 6.0, 4.0 to 6.5, 4.0 to 7.0, 4.0 to 7.5, 4.0 to 8.0, 5.0 to 5.5, 5 0~6.0, 5.0~6.5, 5.0~7.0, 5.0~7.5, 5.0~8.0, 5.5~6.0, 5.5~6.5, 5.5~7.0, 5.5~7.5, 5.5~8.0, 6.0~6.5, 6.0~7.0, 6.0~7.5, 6.0~8.0, 6.5~7.0, 6.5~7.0. 5, 6.5–8.0, 7.0–7.5, 7.0–8.0, 7.5–8.0, for example, approximately 4.0, approximately 4.01, approximately 4.02, approximately 4.03, approximately 4.04, approximately 4.05, approximately 4.06, approximately 4.07, approximately 4.08, approximately 4.09, approximately 4.1, approximately 4.11, approximately 4.12, approximately 4.13, approximately 4.14, Approximately 4.15, 4.16, 4.17, 4.18, 4.19, 4.2, 4.21, 4.22, 4.23, 4.24, 4.25, 4.26, 4.27, 4.28, 4.29, 4.3, 4.31, 4.32, 4.33, 4.34, 4.35, 4. 4.36, approximately 4.37, approximately 4.38, approximately 4.39, approximately 4.4, approximately 4.41, approximately 4.42, approximately 4.43, approximately 4.44, approximately 4.45, approximately 4.46, approximately 4.47, approximately 4.48, approximately 4.49, approximately 4.5, approximately 4.51, approximately 4.52, approximately 4.53, approximately 4.54, approximately 4.55, approximately 4.56, approximately 4 .57, approximately 4.58, approximately 4.59, approximately 4.6, approximately 4.61, approximately 4.62, approximately 4.63, approximately 4.64, approximately 4.65, approximately 4.66, approximately 4.67, approximately 4.68, approximately 4.69, approximately 4.7, approximately 4.71, approximately 4.72, approximately 4.73, approximately 4.74, approximately 4.75, approximately 4.76, approximately 4.77, approximately 4.78, approximately 4.79, approximately 4.8, approximately 4.81, approximately 4.82, approximately 4.83, approximately 4.84, approximately 4.85, approximately 4.86, approximately 4.87, approximately 4.88, approximately 4.89, approximately 4.9, approximately 4.91, approximately 4.92, approximately 4.93, approximately 4.94, approximately 4.95, approximately 4.96, approximately 4.97, approximately 4.98, approximately 4. 99, approximately 5.0, approximately 5.01, approximately 5.02, approximately 5.03, approximately 5.04, approximately 5.05, approximately 5.06, approximately 5.07, approximately 5.08, approximately 5.09, approximately 5.1, approximately 5.11, approximately 5.12, approximately 5.13, approximately 5.14, approximately 5.15, approximately 5.16, approximately 5.17, approximately 5.18, approximately 5.19, approximately 5.2 Approximately 5.21, 5.22, 5.23, 5.24, 5.25, 5.26, 5.27, 5.28, 5.29, 5.3, 5.31, 5.32, 5.33, 5.34, 5.35, 5.36, 5.37, 5.38, 5.39, 5.4, 5.41 Approximately 5.42, 5.43, 5.44, 5.45, 5.46, 5.47, 5.48, 5.49, 5.5, 5.51, 5.52, 5.53, 5.54, 5.55, 5.56, 5.57, 5.58, 5.59, 5.6, 5.61, 5.62. Approximately 5.63, 5.64, 5.65, 5.66, 5.67, 5.68, 5.69, 5.7, 5.71, 5.72, 5.73, 5.74, 5.75, 5.76, 5.77, 5.78, 5.79, 5.8, 5.81, 5.82, 5.83. Approximately 5.84, approximately 5.85, approximately 5.86, approximately 5.87, approximately 5.88, approximately 5.89, approximately 5.9, approximately 5.91, approximately 5.92, approximately 5.93, approximately 5.94, approximately 5.95, approximately 5.96, approximately 5.97, approximately 5.98, approximately 5.99, approximately 6.0, approximately 6.01, approximately 6.02, approximately 6.03, approximately 6.04, approximately 6.05, approximately 6.06, approximately 6.07, approximately 6.08, approximately 6.09, approximately 6.1, approximately 6.11, approximately 6.12, approximately 6.13, approximately 6.14, approximately 6.15, approximately 6.16, approximately 6.17, approximately 6.18, approximately 6.19, approximately 6.2, approximately 6.21, approximately 6.22, approximately 6.23, approximately 6.24, approximately 6.25, approximately 6 .26, about 6.27, about 6.28, about 6.29, about 6.3, about 6.31, about 6.32, about 6.33, about 6.34, about 6.35, about 6.36, about 6.37, about 6.38, about 6.39, about 6.4, about 6.41, about 6.42, about 6.43, about 6.44, about 6.45, about 6.46, about 6.47, approximately 6.48, approximately 6.49, approximately 6.5, approximately 6.51, approximately 6.52, approximately 6.53, approximately 6.54, approximately 6.55, approximately 6.56, approximately 6.57, approximately 6.58, approximately 6.59, approximately 6.6, approximately 6.61, approximately 6.62, approximately 6.63, approximately 6.64, approximately 6.65, approximately 6.66, approximately 6.67, approximately 6.68, approximately 6.69, approximately 6.7, approximately 6.71, approximately 6.72, approximately 6.73, approximately 6.74, approximately 6.75, approximately 6.76, approximately 6.77, approximately 6.78, approximately 6.79, approximately 6.8, approximately 6.81, approximately 6.82, approximately 6.83, approximately 6.84, approximately 6.85 Approximately 6.86, approximately 6.87, approximately 6.88, approximately 6.89, approximately 6.9, approximately 6.91, approximately 6.92, approximately 6.93, approximately 6.94, approximately 6.95, approximately 6.96, approximately 6.97, approximately 6.98, approximately 6.99, approximately 7.0, approximately 7.01, approximately 7.02, approximately 7.03, approximately 7.04, approximately 7.05, approximately 7.06, approximately 7.07, approximately 7.08, approximately 7.09, approximately 7.1, approximately 7.11, approximately 7.12, approximately 7.13, approximately 7.14, approximately 7.15, approximately 7.16, approximately 7.17, approximately 7.18, approximately 7.19, approximately 7.2, approximately 7.21, approximately 7.22, approximately 7.23, approximately 7.24, approximately 7.25, approximately 7.26, approximately 7.27, approximately 7.28, approximately 7.29, approximately 7.3, approximately 7.31, approximately 7.32, approximately 7.33, approximately 7.34, approximately 7.35, approximately 7.36, approximately 7.37, approximately 7.38, approximately 7.39, approximately 7.4, approximately 7.41, approximately 7.42, approximately 7.43, approximately 7.44, approximately 7.45, approximately 7.46, approximately 7.47, approximately 7.48, approximately 7.49, approximately 7.5, approximately 7.51, approximately 7.52, approximately 7.53, approximately 7.54, approximately 7.55, approximately 7.56, approximately 7.57, approximately 7.58, approximately 7.59, approximately 7.6, approximately 7.61, approximately 7. 62, approximately 7.63, approximately 7.64, approximately 7.65, approximately 7.66, approximately 7.67, approximately 7.68, approximately 7.69, approximately 7.7, approximately 7.71, approximately 7.72, approximately 7.73, approximately 7.74, approximately 7.75, approximately 7.76, approximately 7.77, approximately 7.78, approximately 7.79, approximately 7.8, approximately 7.81, approximately 7.82, approximately 7.83, approximately 7.84, approximately 7.85, approximately 7.86, approximately 7.87, approximately 7.88, approximately 7.89, approximately 7.9, approximately 7.91, approximately 7.92, approximately 7.93, approximately 7.94, approximately 7.95, approximately 7.96, approximately 7.97, approximately 7.98, approximately 7.99, approximately 8.0.

[0352] In some embodiments, the compound shown in this disclosure I is optionally substituted with one or more suitable substituents.

[0353] Drug linker

[0354] On the other hand, this disclosure provides compounds of Formula II or their pharmaceutically acceptable salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, isotopically labeled compounds, metabolites, or prodrugs:

[0355] M 1 -LD

[0356] II

[0357] in,

[0358] L and D are as defined in any of the preceding items;

[0359] M 1 It is a precursor for the junction site that connects to the target group.

[0360] In some implementations, the targeting group is as defined in any of the preceding descriptions.

[0361] In some implementation schemes, M 1 Selected from the following structures

[0362] In this context, each 'a' is independently selected from 1, 2, 3, 4, 5, and 6, and 'b' is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; LG represents a leaving group.

[0363] In some implementations, LG is selected from halogens (e.g., F, Cl, Br, I) and halogenated C. 1-6 Alkyl, C 1-6 Alkyl sulfonyl, halogenated C 1-6 Alkyl sulfonyl, halosulfonyl, C 1-6 Alkyl sulfonate group, halogenated C 1-6 Alkyl sulfonate group, C 1-6 Alkyl sulfinate group, C 1-6 Alkyl sulfoxide, halophenoxy, hydroxy, mercapto, amino, nitro, azide, cyano, alkenyl, alkynyl and alkynyl-containing structural fragments.

[0364] In some implementations, LG is C 1-6 Alkyl sulfonyl group, such as methanesulfonyl group.

[0365] In some implementation schemes, M 1 Selected from

[0366] In some implementation schemes, M 1 Selected from

[0367] In some embodiments, the compound represented by Formula II is selected from the following structures:

[0368] In some embodiments, the compound represented by Formula II is optionally substituted with one or more suitable substituents.

[0369] In some embodiments, this disclosure provides the use of a compound of Formula II or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof in the preparation of a conjugate (e.g., a compound of Formula I or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof).

[0370] In Formulas I and II of this disclosure, the functional groups in all embodiments can be suitably selected and combined in any way to obtain different general formula ranges or specific embodiments. These ranges and embodiments are all within the scope of this disclosure. This disclosure covers compounds obtained by arbitrarily combining the various embodiments.

[0371] Antibody or its antigen-binding fragment

[0372] In some embodiments, this disclosure provides an anti-B7H3 antibody or an antigen-binding fragment thereof, comprising: VH as shown in SEQ ID NO:66 or a variant thereof, and / or VL as shown in SEQ ID NO:2 or a variant thereof; preferably, the anti-B7H3 antibody or an antigen-binding fragment thereof comprises: the heavy chain of VH as shown in SEQ ID NO:66 or a variant thereof and the heavy chain constant region (CH) as shown in SEQ ID NO:18 or a variant thereof, and / or the light chain of VL as shown in SEQ ID NO:2 or a variant thereof and the light chain constant region (CL) as shown in SEQ ID NO:19 or a variant thereof; preferably, the amino acid sequence of the heavy chain of the anti-B7H3 antibody or an antigen-binding fragment thereof is as shown in SEQ ID NO:74, and the amino acid sequence of the light chain of the antibody or an antigen-binding fragment thereof is as shown in SEQ ID NO:21.

[0373] In some embodiments, the variant has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with respect to its source sequence, or has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to its source sequence; preferably, the substitutions are conservative substitutions.

[0374] Connection unit

[0375] In some embodiments, this disclosure provides a connecting unit of formula -ML-, wherein M and L are as described in any of the preceding embodiments. In some embodiments, the connecting unit -ML- is used to connect a target group and a load to obtain a coupling. In some embodiments, in the connecting unit -ML-, M is a connector site for connection with the target group, and L is a connection portion for connection with the load. In some embodiments, in the connecting unit -ML-, M is used for connection with the target group described in any of the present invention, and L is used for connection with D described in any of the present application.

[0376] In some embodiments, this disclosure provides a formula M 1 The connection unit shown in -L-, where M 1 And L as described in any of the above. In some embodiments, the connection unit M 1 -L- is used to connect the target group and the load to obtain a coupling. In some embodiments, the connecting unit M 1 In -L-, the M 1 To form a structure capable of reacting with and connecting to a target group, L is a connecting portion connected to the load. In some embodiments, the connecting unit M... 1 In -L-, the M 1 The L is used to react with and connect to the target group described in any one of the present invention, wherein the L is used to connect to the D described in any one of the present applications.

[0377] In some embodiments, this disclosure provides a connecting unit of formula -L-, wherein L is as described in any of the preceding embodiments. In some embodiments, the connecting unit -L- is used to connect a target group and a load to obtain a coupling. In some embodiments, one end of the connecting unit -L- is connected to the target group via a connector that connects to the target group, and the other end is connected to the load. In some embodiments, one end of the connecting unit -L- is connected to the target group described in any of the preceding embodiments via a connector that connects to the target group as described in any of the preceding embodiments, and the other end is connected to D as described in any of the preceding embodiments.

[0378] In some embodiments, this disclosure provides a formula-L 2 - The connection unit shown, where L 2 As described in any of the above. In some embodiments, the connection unit - L 2 - Used to connect the target group and the load to obtain a coupling compound.

[0379] In some implementations, a connection unit is provided that includes -L 2a -L 2b -(Q) n The structure shown, where L 2a L 2b Q and n are defined as in any of the preceding terms.

[0380] In some embodiments, this disclosure provides a connecting unit of formula -LD, wherein L and D are as described in any of the preceding embodiments. In some embodiments, the connecting unit is used to connect a target group via a connector site to obtain a coupling.

[0381] In some embodiments, this disclosure provides a linker unit of formula -MLD, wherein M, L, and D are as described in any of the preceding embodiments. In some embodiments, the linker unit is used to link a target group to obtain a conjugate.

[0382] In some implementations, the connection unit shown in -ML- has the following structure:

[0383] In this configuration, position 2 is connected to the target group, and position 5 is connected to the load.

[0384] In some embodiments, a compound or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof is provided, comprising -L-, -L- 2 -、-L 2a -L 2b -(Q) n -ML-,M 1 Connection units with structures shown as -L-, -LD, or -MLD.

[0385] In some embodiments, a compound or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof is provided, comprising -L-, -L- 2 -、-ML-、M 1 -L- The connection unit of the structure shown in Ab and Ab 1 And / or D, the connecting unit and Ab 1 And / or D connection; where Ab 1 Or D as defined in any of the preceding items.

[0386] In some embodiments, a compound or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug is provided, comprising a linker unit of the structure shown in -LD or -MLD and Ab. 1 The connection unit and Ab 1 Connect; where Ab 1 As defined in any of the preceding items.

[0387] In some embodiments, the aforementioned connecting unit is provided for use in the preparation of couplings (e.g., couplings as defined in any of the preceding items).

[0388] intermediate

[0389] In some embodiments, this disclosure provides intermediate compounds with the following structures or their salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, or isotopically labeled compounds:

[0390] in

[0391] PG 1 Each group is independently an H or amino protecting group, such as alkoxycarbonyl amino protecting groups, for example benzyloxycarbonyl (Cbz), tert-butoxycarbonyl (Boc), methoxycarbonyl (Fmoc), allyloxycarbonyl (Alloc), trimethylsilylethoxycarbonyl (Teoc), methoxycarbonyl (or ethoxycarbonyl); acyl amino protecting groups, such as phthaloyl (Pht), p-toluenesulfonyl (Tos), trifluoroacetyl (Tfa), o-(p-)nitrobenzenesulfonyl (Ns), tert-pentanoyl, benzoyl, tert-butoxycarbonyl, 9-fluorenylmethoxycarbonyl, allyloxycarbonyl, trichloroethoxycarbonyl, trimethylsilylethoxycarbonyl, benzyloxycarbonyl, p-methylbenzenesulfonyl, p-nitrobenzenesulfonyl, trifluoroacetyl, methoxycarbonyl, or ethoxycarbonyl; alkyl amino protecting groups, such as triphenylmethyl (Trt), C 1-6 Alkyl-substituted triphenylmethyl, p-methoxytriphenylmethyl (MMT), dimethoxytriphenylmethyl (DMT), 2,4-dimethoxybenzyl (Dmb), 4-methoxybenzyl (PMB), benzyl (Bn);

[0392] PG 2 Each is independently protected by an H or a carboxyl group, the carboxyl protecting group being, for example, a C group. 1-6Alkyl, allyl, benzyl, 2,4-dimethoxybenzyl, p-methoxybenzyl, methoxyethoxymethyl, pentafluorophenyl, 4-p-methylbenzyloxybenzyl;

[0393] LG is as defined in the previous item.

[0394] In some implementations, this disclosure provides a PG 1 -L 3 -L 4 -D、PG 1 -L 2b -(Q) n PG 1 -L 2a -L 2b -(Q) n PG 1 -L 2 -PG 3 or PG 1 -LD represents the intermediate compound or its salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, or isotopically labeled compound; wherein PG 3 -O-PG 2 ;PG 1 PG 2 L 2 L 3 L 4 L 2a L 2b L, Q, n, and D are as defined in any of the preceding terms.

[0395] In some embodiments, this disclosure provides intermediate compounds with the structures shown below, or their salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, or isotopically labeled compounds:

[0396] In some embodiments, this disclosure provides for the use of the intermediate compound as described above, or its salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, isotope-labeled compounds, in the preparation of the compound of this disclosure or its pharmaceutically acceptable salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, isotope-labeled compounds, metabolites, or prodrugs.

[0397] Pharmaceutical Composition

[0398] In another aspect, this disclosure provides a pharmaceutical composition comprising any of the compounds described in any of the preceding statements or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitride, isotope label, metabolite or prodrug thereof, and one or more pharmaceutically acceptable carriers.

[0399] The above-described pharmaceutical compositions can act systemically and / or locally, which can be achieved through suitable dosage forms. These dosage forms include, but are not limited to, tablets, capsules, lozenges, hard candies, powders, sprays, creams, ointments, suppositories, gels, pastes, lotions, aqueous suspensions, injectable solutions, elixirs, and syrups.

[0400] The pharmaceutical composition described above may contain 0.01 mg to 1000 mg of at least one of the compounds disclosed herein or a pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitrogen oxide, isotope label, metabolite or prodrug thereof.

[0401] In some embodiments, the above-described pharmaceutical composition may optionally further include one or more other therapeutic agents.

[0402] This disclosure also provides a method for preparing the above-described pharmaceutical composition or its corresponding formulation, comprising combining at least one of the compounds of this disclosure or its pharmaceutically acceptable salt, ester, stereoisomer, polymorph, solvate, nitride, isotope label, metabolite or prodrug with one or more pharmaceutically acceptable carriers.

[0403] Treatment methods and applications

[0404] In another aspect, this disclosure provides for the use of the compound of Formula II as described in any of the preceding statements, or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, in the preparation of the compound of Formula I, or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof.

[0405] In another aspect, this disclosure provides the use of any of the compounds described above (e.g., compounds represented by Formula I or Formula II) or pharmaceutically acceptable salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, isotopically labeled compounds, metabolites, or prodrugs or compositions thereof in the preparation of medicaments, particularly in the preparation of medicaments for the treatment and / or prevention of cancer.

[0406] In another aspect, this disclosure provides a compound described in any of the preceding statements (e.g., a compound represented by Formula I or Formula II) or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug or composition thereof for the treatment and / or prevention of diseases, such as cancer.

[0407] In another aspect, this disclosure provides a method of treating and / or preventing cancer, comprising administering to a subject in need a therapeutically and / or preventively effective amount of any of the preceding compounds (e.g., compounds represented by Formula I or Formula II) or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug or composition thereof.

[0408] In some embodiments, the cancer is cancer associated with GSPT1 protein dysregulation, cancer associated with HER2 expression, and / or cancer associated with B7H3 expression.

[0409] In some implementations, the cancer is selected from gastric cancer, breast cancer, or lung cancer (e.g., small cell lung cancer or non-small cell lung cancer). Beneficial effects

[0410] The compounds disclosed herein have one or more of the following beneficial effects, but are not limited to:

[0411] (1) It has a good degradation effect on GSPT1 protein;

[0412] (2) It has a strong inhibitory activity against tumor cell proliferation;

[0413] (3) It has a good tumor-suppressing effect;

[0414] (4) Excellent physicochemical properties (e.g., solubility, physical and / or chemical stability);

[0415] (5) Excellent pharmacokinetic properties (e.g., good bioavailability, appropriate half-life and duration of action);

[0416] (6) Excellent safety profile (lower toxicity and / or fewer side effects, wider therapeutic window), etc.

[0417] Example

[0418] To make the objectives and technical solutions of this disclosure clearer, the embodiments of this disclosure are described in detail below. However, those skilled in the art will understand that the following embodiments are for illustrative purposes only and should not be considered as limiting the scope of this disclosure. Unless otherwise specified, specific conditions in the embodiments are performed under conventional conditions or conditions recommended by the manufacturer. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0419] The structure of the compound was determined by nuclear magnetic resonance (NMR). 1 It was determined by 1H NMR or mass spectrometry (MS). 1 The 1H NMR was performed using a JEOL Eclipse400 NMR spectrometer. The solvents used were deuterated methanol (CD3OD), deuterated chloroform (CDCl3), or hexadeuterated dimethyl sulfoxide (DMSO-d6). The internal standard was tetramethylsilane (TMS). Chemical shifts (δ) were given in parts per million (ppm).

[0420] The instrument used for MS measurements was an Agilent (ESI) mass spectrometer, manufacturer: Agilent, model: Agilent 6120B.

[0421] Preparation method of high performance liquid chromatograph:

[0422] Instrument model: Agilent 1260; Column: Waters SunFire Prep C18 OBD (19mm×150mm×5.0μm); Column temperature: 25℃; Flow rate: 20.0mL / min; Detection wavelength: 214nm; Elution gradient: (0min: 10%A, 90%B; 16.0min: 90%A, 10%B); Mobile phase A: Acetonitrile; Mobile phase B: 0.05% formic acid aqueous solution.

[0423] The thin-layer chromatography silica gel plates (TLC) used were Merck aluminum plates (20×20cm), and the TLC separation and purification used Yantai-made GF 254 (1mm) plates.

[0424] The reaction was monitored using thin-layer chromatography (TLC) or LC-MS. The developing solvent systems used included dichloromethane and methanol, n-hexane and ethyl acetate, and petroleum ether and ethyl acetate. The volume ratio of the solvent was adjusted according to the polarity of the compound or by adding triethylamine, etc.

[0425] Preparation method of reverse column chromatography:

[0426] Preparation method A:

[0427] Instrument model: Biotage rapid medium-pressure preparative chromatography; Column: Agela C18 reverse-flow column (Spherical; 20-35 μm; 100A); Column temperature: 25℃; Flow rate: 28.0 mL / min; Detection wavelength: 220 nm; Mobile phase A: acetonitrile; Mobile phase B: water;

[0428] Preparation method B:

[0429] Instrument model: Biotage rapid medium-pressure preparative chromatography; Column: Agela C18 reverse-flow column (Spherical; 20-35μm; 100A); Column temperature: 25℃; Flow rate: 28.0mL / min; Detection wavelength: 220nm; Mobile phase A: acetonitrile; Mobile phase B: 0.05% formic acid aqueous solution;

[0430] Preparation method C:

[0431] Instrument model: Biotage rapid medium-pressure preparative chromatography; Column: Agela C18 reverse-flow column (Spherical; 20-35 μm; 100A); Column temperature: 25℃; Flow rate: 28.0 mL / min; Detection wavelength: 220 nm; Mobile phase A: acetonitrile; Mobile phase B: 0.05% NH4HCO3 aqueous solution;

[0432] Preparation method D:

[0433] Instrument model: Biotage rapid medium-pressure preparative chromatography; Column: Agela C18 reverse-flow column (Spherical; 20-35 μm; 100A); Column temperature: 25℃; Flow rate: 28.0 mL / min; Detection wavelength: 220 nm; Mobile phase A: acetonitrile; Mobile phase B: 0.05% trifluoroacetic acid aqueous solution;

[0434] The microwave reaction was performed using a Biotage Initiator+ (400W, RT~300℃) microwave reactor.

[0435] Column chromatography typically uses 200-300 mesh silica gel as the support. Eluent systems include dichloromethane and methanol systems, and petroleum ether and ethyl acetate systems. The volume ratio of the solvents is adjusted according to the polarity of the compounds, and a small amount of triethylamine can also be added for adjustment.

[0436] Unless otherwise specified in the examples, the reaction temperature is room temperature (20℃~35℃).

[0437] The reagents used in this disclosure were purchased from Acros Organics, Aldrich Chemical Company, TEB Chemicals, and other companies.

[0438] In the conventional synthesis methods and examples, as well as intermediate synthesis examples, the meanings of the abbreviations are as follows.

[0439] Information about the sequences involved in this disclosure is described in the table below:

[0440] I. Compound Examples

[0441] Example 1: Preparation of intermediate 1

[0442] Step 1: Preparation of tert-butyl 2-methylthiophene-3-carboxylate

[0443] 2-Methylthiophene-3-carboxylic acid (11.5 g, 80.89 mmol) was dissolved in dry tetrahydrofuran (60 mL), cooled to 0 °C, and O-tert-butyl-N,N'-diisopropylisourea (40.92 g, 202.22 mmol) was slowly added dropwise. The reaction mixture was reacted at 50 °C for 16 hours. The reaction mixture was cooled to room temperature, filtered, and the filter cake was washed with tetrahydrofuran. The filtrates were combined and concentrated to obtain a crude product, which was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 4 / 1 (v / v)) to give the title compound of this step (12.35 g, yield: 77.0%).

[0444] MS m / z (ESI): 199.1 [M+H] + .

[0445] 1 H NMR (400MHz, DMSO-d6) δ7.29-7.23(m,2H),2.64(s,3H),1.52(s,9H).

[0446] Step 2: Preparation of tert-butyl 5-bromo-2-methylthiophene-3-carboxylate

[0447] 12.35 g (62.29 mmol) of tert-butyl 2-methylthiophene-3-carboxylate was dissolved in a mixed solvent of anhydrous N,N-dimethylformamide (40 mL) and acetic acid (8 mL). The mixture was cooled to 0 °C, and N-bromosuccinimide (12.19 g, 68.51 mmol) was added in portions. The reaction mixture was allowed to react at room temperature for 16 hours. The reaction solution was diluted with water, extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous magnesium sulfate, filtered, and the filtrate was concentrated to give the title compound of this step (16.15 g, yield: 93.6%).

[0448] MS m / z(ESI): 279.0 [M+H] + .

[0449] 1 H NMR (400MHz, DMSO-d6) δ7.37(s,1H),2.63(s,3H),1.52(s,9H).

[0450] Step 3: Preparation of tert-butyl 5-bromo-2-(bromomethyl)thiophene-3-carboxylate

[0451] 5-Bromo-2-methylthiophene-3-carboxylic acid tert-butyl ester (16.15 g, 61.33 mmol) and benzoyl peroxide (1.49 g, 6.13 mmol) were dissolved in carbon tetrachloride (50 mL), and N-bromosuccinimide (10.92 g, 61.33 mmol) was added. The reaction mixture was reacted at 80 °C for 16 hours. The reaction solution was concentrated and diluted with an appropriate amount of water. The mixture was extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous magnesium sulfate, filtered, and the filtrate was concentrated to give the title compound of this step (19 g, yield: 87.0%).

[0452] MS m / z (ESI): 356.9 [M+H] + .

[0453] 1 H NMR (400MHz, DMSO-d6) δ7.40(s,1H),5.15(s,2H),1.54(s,9H).

[0454] Step 4: Preparation of tert-butyl 5-bromo-2-(((2,6-dioxopiperidin-3-yl)amino)methyl)thiophene-3-carboxylate

[0455] 10.9 g (30.61 mmol) of tert-butyl 5-bromo-2-(bromomethyl)thiophene-3-carboxylate was dissolved in anhydrous N,N-dimethylformamide (30 mL), and N,N-diisopropylethylamine (11.87 g (91.83 mmol) and 3-aminopiperidine-2,6-dione hydrochloride (4.03 g (24.49 mmol)) were added. The mixture was reacted at 80 °C for 3 hours. A suitable amount of water was added to the reaction solution, and the mixture was extracted three times with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous magnesium sulfate, filtered, and the crude product obtained by concentration was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 13 / 7 (v / v)) to give the title compound of this step (7.6 g, yield: 61.6%).

[0456] MS m / z(ESI): 405.0 [M+H] + .

[0457] Step 5: Preparation of 5-bromo-2-(((2,6-dioxopiperidin-3-yl)amino)methyl)thiophene-3-carboxylic acid

[0458] 1.27 g (3.15 mmol) of 5-bromo-2-(((2,6-dioxopiridine-3-yl)amino)methyl)thiophene-3-carboxylic acid tert-butyl ester was dissolved in dichloromethane (8 mL) and trifluoroacetic acid (4 mL) and reacted at room temperature for 6 hours. The reaction solution was concentrated to give the title compound of this step (1 g, yield: 91.2%).

[0459] MS m / z(ESI): 349.0 [M+H] + .

[0460] Step 6: Preparation of 3-(2-bromo-4-oxo-4,6-dihydro-5H-thieno[2,3-c]pyrrolo-5-yl)piperidine-2,6-dione

[0461] 5-Bromo-2-(((2,6-dioxopiperidin-3-yl)amino)methyl)thiophene-3-carboxylic acid (1 g, 2.87 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (1.64 g, 4.31 mmol) were added to anhydrous N,N-dimethylformamide (10 mL), the mixture was cooled to 0 °C, and N,N-diisopropylethylamine (1.11 g, 8.62 mmol) was added. The mixture was reacted at room temperature for 2 hours. The reaction solution was poured into ice water, filtered, and the filter cake was dried to give the title compound (871 mg, yield: 92.1%).

[0462] MS m / z(ESI): 330.9 [M+H] + .

[0463] Step 7: Preparation of 3-(2-(hydroxymethyl)-4-oxo-4,6-dihydro-5H-thieno[2,3-c]pyrrolo-5-yl)piperidine-2,6-dione

[0464] 3-(2-bromo-4-oxo-4,6-dihydro-5H-thieno[2,3-c]pyrrolo-5-yl)piperidine-2,6-dione (871 mg, 2.64 mmol), chloro(2-dicyclohexylphosphino-2',4',6'-triisopropyl-1,1'-biphenyl)[2-(2'-amino-1,1'-biphenyl)]palladium(II) (207.51 mg, 264.07 μmol), and (tributyltin)methanol (1.27 g, 3.96 mmol) were dissolved in anhydrous 1,4-dioxane (15 mL), purged three times with nitrogen, and reacted at 80 °C for 18 hours. The reaction mixture was poured into ice water, filtered, and the filter cake was dried to give the title compound (680 mg, yield: 91.9%).

[0465] MS m / z(ESI): 281.1 [M+H] + .

[0466] Example 2: Preparation of intermediate 2: Preparation of compound Int2

[0467] Step 1: Preparation of tert-butyl(2-chloro-4-nitrophenethoxy)diphenylsilane

[0468] 2-(2-chloro-4-nitrophenyl)ethane-1-ol (500 mg, 2.48 mmol), triethylamine (500 mg, 4.96 mmol), tert-butyldiphenylchlorosilane (1.02 g, 3.72 mmol), and dimethylaminopyridine (30.3 mg, 248.0 μmol) were added to dichloromethane (50 mL), and the reaction was carried out at room temperature for 16 hours. The crude product obtained by concentration of the reaction solution was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 1 / 1 (v / v)) to give the title compound of this step (800 mg, yield: 73.4%).

[0469] MS m / z (ESI): 440.1 [M+H] + .

[0470] Step 2: Preparation of 4-(2-((tert-butyldiphenylsilyl)oxy)ethyl)-3-chloroaniline

[0471] 100 mg of tert-butyl(2-chloro-4-nitrophenylethoxy)diphenylsilane (227.27 μmol) was dissolved in 7 mL of ethyl acetate. 10% palladium on carbon (82 mg) and zinc bromide (10.23 mg, 45.45 μmol) were added. The mixture was purged with hydrogen and reacted at room temperature for 2 hours under a hydrogen atmosphere. The reaction solution was filtered through diatomaceous earth. The crude product obtained by concentration of the filtrate was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 3 / 2 (v / v)) to give the title compound (72 mg, yield: 77.3%).

[0472] MS m / z (ESI): 410.2 [M+H] + .

[0473] Step 3: Preparation of phenyl(4-(2-((tert-butyldiphenylsilyl)oxy)ethyl)-3-chlorophenyl)carbamate

[0474] 4-(2-((tert-butyldiphenylsilyl)oxy)ethyl)-3-chloroaniline (34 mg, 82.92 μmol) was dissolved in acetonitrile (3 mL), and pyridine (32.80 mg, 414.62 μmol) was added. The mixture was cooled to 0 °C in an ice bath and reacted for 10 minutes. Then, phenyl chloroformate (19.48 mg, 124.38 μmol) was added, and the mixture was reacted at room temperature for 1 hour. The reaction solution was concentrated to give the title compound of this step (34 mg, yield: 77.3%).

[0475] MS m / z (ESI): 530.2 [M+H] + .

[0476] Step 4: Preparation of (5-(2,6-dioxopiperidin-3-yl)-4-oxo-5,6-dihydro-4H-thieno[2,3-c]pyrrolo-2-yl)methyl(4-(2-((tert-butyldiphenylsilyl)oxy)ethyl)-3-chlorophenyl)carbamate

[0477] Phenyl(4-(2-((tert-butyldiphenylsilyl)oxy)ethyl)-3-chlorophenyl)carbamate (22 mg, 41.50 μmol) and 3-(2-(hydroxymethyl)-4-oxo-4,6-dihydro-5H-thieno[2,3-c]pyrrolo-5-yl)piperidin-2,6-dione (17.45 mg, 62.25 μmol) were added to N,N-dimethylformamide (2 mL), purged with nitrogen, and cooled to 0 °C in an ice bath. 1,8-diazacyclo[5,4,0]undecene-7 (31.59 mg, 207.50 μmol) was added, and the reaction was maintained at this temperature for half an hour, followed by reaction at room temperature for 1 hour. The reaction mixture was extracted with water and ethyl acetate. The organic layers were combined and concentrated, and the crude product was purified by silica gel column chromatography (eluent: ethyl acetate) to give the title compound of this step (22.5 mg, yield: 75.7%).

[0478] MS m / z (ESI): 716.2 [M+H] + .

[0479] Step 5: Preparation of (5-(2,6-dioxopiperidin-3-yl)-4-oxo-5,6-dihydro-4H-thieno[2,3-c]pyrrolo-2-yl)methyl(3-chloro-4-(2-hydroxyethyl)phenyl)carbamate

[0480] (5-(2,6-dioxopiperidin-3-yl)-4-oxo-5,6-dihydro-4H-thieno[2,3-c]pyrrolo-2-yl)methyl(4-(2-((tert-butyldiphenylsilyl)oxy)ethyl)-3-chlorophenyl)carbamate (17 mg, 23.73 μmol) was dissolved in tetrahydrofuran (2 mL), and pyridine hydrogen fluoride (11.76 mg, 118.65 μmol) was added. The reaction was carried out at room temperature for 2 hours. The crude product obtained by concentration was purified by preparative high performance liquid chromatography to give the title compound (4.8 mg, yield: 41.9%).

[0481] MS m / z (ESI): 478.1 [M+H] + .

[0482] 1 H NMR (400MHz, DMSO-d6) δ10.95(s,1H),9.93(s,1H),7.58(s,1H),7.33(s,3H),5.35(s,2H),5.00(dd,J=12.0,4.0Hz,1H),4.69(t,J=8.0,4.0H z,1H),4.36-4.55(m,2H),3.58-3.53(m,2H),2.98-2.82(m,1H),2.88- 2.73(m,2H),2.71-2.64(m,1H),2.38-2.25(m,1H),2.05-1.92(m,1H).

[0483] Example 3: Preparation of intermediate Int3

[0484] Step 1: Preparation of 2-(3-chloro-4-fluorophenyl)-2-oxoacetic acid

[0485] 1-(3-chloro-4-fluorophenyl)ethane-1-one (2 g, 11.59 mmol) and selenium dioxide (2.83 g, 25.49 mmol) were added to dry pyridine (15 mL), purged three times with nitrogen, and reacted at 110 °C for 18 hours. The reaction mixture was cooled to room temperature, and 1 N hydrochloric acid was added to adjust the reaction system to acidity. The mixture was extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous magnesium sulfate, filtered, and the filtrate was concentrated to give the title compound of this step (2.1 g, yield: 85.0%).

[0486] MS m / z(ESI): 203.1 [M+H] + .

[0487] Step 2: Preparation of methyl 2-(3-chloro-4-fluorophenyl)-2-oxoacetate

[0488] 2-(3-chloro-4-fluorophenyl)-2-oxoacetic acid (1.0 g, 4.69 mmol) was dissolved in anhydrous methanol (5 mL), and thionyl chloride (2.23 g, 18.76 mmol) was slowly added. The reaction mixture was reacted at room temperature for 2 hours. The crude product obtained by concentration of the reaction solution was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 20 / 1 (v / v)) to give the title compound of this step (850 mg, yield: 79.5%).

[0489] MS m / z(ESI): 217.0 [M+H] + .

[0490] Step 3: Preparation of methyl 2-(3-chloro-4-(3-(4-nitrophenyl)azacyclobutane-1-yl)phenyl)-2-oxoacetate

[0491] 3-(4-nitrophenyl)azacyclobutane (200 mg, 1.12 mmol), methyl 2-(3-chloro-4-fluorophenyl)-2-oxoacetate (267.42 mg, 1.23 mmol), and potassium carbonate (465.37 mg, 3.37 mmol) were added to anhydrous N,N-dimethylformamide (5 mL), and reacted at 90 °C for 3 hours. The reaction solution was cooled to room temperature, diluted with water, extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous magnesium sulfate, filtered, and the crude product obtained by concentration was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 4 / 1 (v / v)) to give the title compound of this step (270 mg, yield: 61.0%).

[0492] MS m / z(ESI): 375.0 [M+H] + .

[0493] Step 4: Preparation of 2-(3-chloro-4-(3-(4-nitrophenyl)azacyclobutane-1-yl)phenyl)-2-oxoacetic acid

[0494] Methyl 2-(3-chloro-4-(3-(4-nitrophenyl)azacyclobutan-1-yl)phenyl)-2-oxoacetate (270 mg, 684.41 μmol) and lithium hydroxide monohydrate (143.60 mg, 3.42 mmol) were dissolved in a mixture of tetrahydrofuran (4 mL) and water (1 mL), and reacted at room temperature for 4 hours. The reaction solution was concentrated, and the pH was adjusted to 4–5 with 1 N hydrochloric acid. The solution was then filtered to give the title compound of this step (255 mg, yield: 98.1%).

[0495] MS m / z(ESI): 361.0 [M+H] + .

[0496] Step 5: Preparation of 2-(3-chloro-4-(3-(4-nitrophenyl)azacyclobutane-1-yl)phenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoline-5-yl)methyl)-2-oxoacetamide

[0497] 3-(5-(aminomethyl)-1-oxoisoindoline-2-yl)piperidin-2,6-dione (208 mg, 762 μmol), 2-(3-chloro-4-(3-(4-nitrophenyl)azacyclobutane-1-yl)phenyl)-2-oxoacetic acid (250 mg, 693 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (316 mg, 832 μmol), and N,N-diisopropylethylamine (269 mg, 2.08 mmol) were added to anhydrous N,N-dimethylformamide (3 mL), and the reaction was carried out under nitrogen protection at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 45%-90%) to give the title compound of this step (280 mg, yield: 62.3%).

[0498] MS m / z (ESI): 616.2 [M+H] + .

[0499] Step 6: Preparation of 2-(4-(3-(4-aminophenyl)azacyclobutane-1-yl)-3-chlorophenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoline-5-yl)methyl)-2-oxoacetamide

[0500] 2-(3-chloro-4-(3-(4-nitrophenyl)azacyclobutane-1-yl)phenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoline-5-yl)methyl)-2-oxoacetamide (50.0 mg, 81.1 μmol) and tetrahydroxydiboron (21.8 mg, 244 μmol) were added to anhydrous N,N-dimethylformamide (1.5 mL), followed by 4,4'-bipyridine (0.63 mg, 4.06 μmol). The reaction mixture was reacted at room temperature for 5 minutes. The crude product obtained by concentration was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-90%) to give the title compound of this step (16.0 mg, yield: 32.0%).

[0501] MS m / z (ESI): 586.3 [M+H] + .

[0502] 1H NMR (400MHz, DMSO-d6) δ10.99(s,1H),9.47(t,J=6.0Hz,1H),7.88(d,J=2.0Hz,1H),7.84(dd,J=8.0Hz,2.0Hz,1H),7.72( d,J=8.0Hz,1H),7.54(s,1H),7.47(d,J=8.0Hz,1H),7.09-7.01(m,2H),6.65(d,J=8.0Hz,1H),6.58-6.51(m,2H),5.11(dd ,J=14.0,6.0Hz,1H),5.00(s,2H),4.61(t,J=8.0Hz,2H),4.54(d,J=8.0Hz,2H),4.46(d,J=20.0Hz,1H),4.33(d,J=20.0H z,1H),4.12-4.10(m,2H),3.82-3.75(m,1H),2.97-2.85(m,1H),2.64-2.56(m,1H),2.41-2.34(m,1H),2.04-1.96(m,1H).

[0503] Example 4: Preparation of intermediate Int4

[0504] Step 1: Preparation of compound Int4-1

[0505] (5-(2,6-dioxopiperidin-3-yl)-4-oxo-5,6-dihydro-4H-thieno[2,3-c]pyrrolo-2-yl)methyl(3-chloro-4-(2-hydroxyethyl)phenyl)carbamate (150 mg, 313.86 μmol) and (S)-(2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propionamido)methyl acetate (600.1 mg, 1.57 mmol) were dissolved in... In tetrahydrofuran (6 mL), 4A molecular sieve (50 mg) was added and stirred for 5 minutes. Then, scandium trifluoromethanesulfonate (772 mg, 1.57 mmol) was added and reacted at room temperature for 3 hours. The reaction solution was filtered, and the crude product obtained by concentration of the filtrate was dissolved in methanol (2 mL) and purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 60%-65%) to obtain the title compound of this step (142.5 mg, yield: 56.7%).

[0506] MS m / z (ESI): 800.1 [M+H] + .

[0507] Step 2: Preparation of compound Int4-2

[0508] Compound Int4-1 (120 mg, 149.95 μmol) and diethylamine (54.8 mg, 749.74 μmol) were dissolved in N,N-dimethylformamide (3 mL) and reacted at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 35%-40%) to give the title compound of this step (51 mg, yield: 58.8%).

[0509] MS m / z (ESI): 578.1 [M+H] + .

[0510] Step 3: Preparation of compound Int4-3

[0511] Compound Int4-2 (51 mg, 88.4 μmol), (((9H-fluorene-9-yl)methoxy)carbonyl)-L-alanyl-L-alanine (23.1 mg, 88.4 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (50.4 mg, 132.6 μmol), and N,N-diisopropylethylamine (17.1 mg, 132.6 mmol) were added to anhydrous N,N-dimethylformamide (4 mL) under ice bath cooling, and the reaction was carried out at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 40%-50%) to give the title compound of this step (35 mg, yield: 42.0%).

[0512] MS m / z (ESI): 942.1 [M+H] + .

[0513] Step 4: Preparation of compound Int4

[0514] Compound Int4-3 (35 mg, 37.19 μmol) and diethylamine (13.6 mg, 185.95 μmol) were dissolved in N,N-dimethylformamide (3 mL) and reacted at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method A, elution gradient: mobile phase A% = 35%-40%) to give the title compound of this step (26 mg, yield: 97.2%).

[0515] MS m / z(ESI): 720.2 [M+H] + .

[0516] Example 5: Preparation of intermediate Int5

[0517] Step 1: Preparation of compound Int5-2

[0518] Compound Int5-1 (1 g, 2.12 mmol), 2-amino-2-(hydroxymethyl)propane-1,3-diol (1.8 g, 14.85 mmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (3.63 g, 9.54 mmol), and pyridine (1.68 g, 21.21 mmol) were added to anhydrous DMSO (12 mL) and tetrahydrofuran (12 mL) under ice bath cooling, and reacted at 40 °C for 12 hours. The crude product obtained by concentration of the reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-15%) to give the title compound of this step (800 mg, yield: 48.3%).

[0519] MS m / z (ESI): 781.3 [M+H] + .

[0520] Step 2: Preparation of compound Int5-3

[0521] Compound Int5-2 (800 mg, 1.03 mmol) and 10% palladium on carbon (297.95 mg, 0.49 mmol) were added to anhydrous methanol (10 mL), and the mixture was purged with hydrogen and reacted at room temperature for 2 hours. The reaction solution was filtered through a diatomaceous earth sieve, and the filtrate was concentrated to give the title compound of this step (648 mg, yield: 97.1%).

[0522] MS m / z (ESI): 647.3 [M+H] + .

[0523] Step 3: Preparation of compound Int5-4

[0524] Compound Int5-3 (0.5 g, 0.77 mmol), (S)-2-((((9H-fluorene-9-yl)methoxy)carbonyl)amino)-5-(allyloxy)-5-oxovaleric acid (316.6 mg, 0.77 mmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (323.4 mg, 0.85 mmol), and N,N-diisopropylethylamine (199.85 mg, 21.55 mmol) were added to anhydrous N,N-dimethylformamide (6 mL) under ice bath cooling, and the reaction was carried out at room temperature for 3 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 35%-40%) to give the title compound of this step (600 mg, yield: 74.8%).

[0525] MS m / z (ESI): 1038.5 [M+H] + .

[0526] Step 4: Preparation of compound Int5

[0527] Compound Int5-4 (600 mg, 0.58 mmol), phenylsilane (375.5 mg, 3.47 mmol), and tetrakis(triphenylphosphine)palladium (134 mg, 0.116 mmol) were added to anhydrous N,N-dimethylformamide (5 mL) under nitrogen protection and reacted at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation Method A, elution gradient: mobile phase A% = 25%-30%) to give the title compound of this step (500 mg, yield: 86.4%).

[0528] MS m / z (ESI): 998.4 [M+H] + .

[0529] Example 6: Preparation of intermediate 6

[0530] Step 1: Preparation of compound Int6-1

[0531] Compounds Int4 (40 mg, 55.54 μmol), Int5 (66.5 mg, 66.65 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (25.3 mg, 66.65 μmol), and N,N-diisopropylethylamine (21.5 mg, 166.62 μmol) were added to anhydrous tetrahydrofuran (3 mL) and anhydrous DMSO (1 mL) under ice bath cooling, and the reaction was carried out at room temperature for 2 hours. The crude product obtained by concentration of the reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 40%-50%) to give the title compound of this step (70 mg, yield: 74.1%).

[0532] MS m / z (ESI): 850.1 [M / 2+H] + .

[0533] Step 2: Preparation of compound Int6

[0534] Compound Int6-1 (70 mg, 341.17 μmol) and diethylamine (27.1 mg, 370.54 μmol) were added to N,N-dimethylformamide (3 mL) and reacted at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 15%-20%) to give the title compound of this step (45 mg, yield: 89.4%).

[0535] MS m / z(ESI): 1177.1 [M+H] + .

[0536] Example 7: Preparation of intermediate Int7

[0537] Step 1: Preparation of compound Int7-1

[0538] (2-(4-amino-2-chlorophenethoxy)ethyl)(methyl)carbamate tert-butyl ester (100 mg, 0.243 mmol) and N,N'-disuccinimidyl carbonate (155.0 mg, 0.608 mmol) were dissolved in dry dichloromethane (6 mL) under ice bath cooling. 50 mg of 4A molecular sieve was added, nitrogen was purged three times, and the reaction was carried out at room temperature for 2 hours. The reaction solution was filtered, and the crude product obtained by concentrating the filtrate was dissolved in dry N-methylpyrrolidone (2 mL) and used directly in the next step of the reaction.

[0539] MS m / z (ESI): 414.1 [M+H-56] + .

[0540] Step 2: Preparation of compound Int7-2

[0541] 3-(2-(hydroxymethyl)-4-oxo-4,6-dihydro-5H-thieno[2,3-c]pyrrolo-5-yl)piperidine-2,6-dione (116.32 mg, 414.97 μmol) and N,N-diisopropylethylamine (71.5 mg, 553.29 μmol) were dissolved in N-methylpyrrolidone (2 mL) under ice bath cooling. The N-methylpyrrolidone solution from the previous step was added, and the reaction was carried out at room temperature for 36 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 85%-90%) to give the title compound of this step (69 mg, yield: 39.3%).

[0542] MS m / z(ESI): 535.1 [M+H-100] + .

[0543] Step 3: Preparation of compound Int7-3

[0544] Compound Int7-2 (69 mg, 108.64 μmol) was added to a mixed solvent of trifluoroacetic acid (1.5 mL) and dichloromethane (3 mL), and reacted at room temperature for 2 hours. The reaction solution was concentrated to give the title compound of this step (58 mg, yield: 99.8%).

[0545] MS m / z (ESI): 535.1 [M+H] + .

[0546] Step 4: Preparation of compound Int7-4

[0547] Compounds Int7-3 (58 mg, 108.4 μmol), Int7-5 (70.32 mg, 130.1 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (49.5 mg, 130.1 μmol), and N,N-diisopropylethylamine (28.1 mg, 216.8 μmol) were added to anhydrous N,N-dimethylformamide (4 mL) under ice bath cooling, and the reaction was carried out at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 35%-40%) to give the title compound of this step (105 mg, yield: 91.6%).

[0548] MS m / z (ESI): 1057.3 [M+H] + .

[0549] Step 4: Preparation of compound Int7

[0550] Compound Int7-4 (50 mg, 47.28 μmol) was dissolved in anhydrous N,N-dimethylformamide (3 mL), followed by the addition of diethylamine (31.1 mg, 425.5 μmol), and the reaction was carried out at room temperature for 2 hours. The reaction solution was then purified directly by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 25%-30%) to obtain the title compound of this step (38 mg, yield: 96.2%).

[0551] MS m / z (ESI): 835.2 [M+H] + .

[0552] Example 8: Preparation of compound Int8

[0553] Following the synthetic route of compound Int6, compound Int4 was replaced with compound Int7 to obtain the title compound (13 mg).

[0554] MS m / z (ESI): 1592.6 [M+H] + .

[0555] Example 9: Preparation of intermediate: Int9

[0556] Step 1: Preparation of compound Int9-1

[0557] 2-(4-(3-(4-aminophenyl)azacyclobutane-1-yl)-3-chlorophenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoline-5-yl)methyl)-2-oxoacetamide (140 mg, 239 μmol), (((9H-fluorene-9-yl)methoxy)carbonyl)-L-valine-L-alanine (118 mg, 287 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (109 mg, 287 μmol), and N,N-diisopropylethylamine (92.6 mg, 717 μmol) were added to anhydrous N,N-dimethylformamide (4 mL), and the reaction was carried out under nitrogen protection at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-70%) to obtain the title compound of this step (38.0 mg, yield: 15.5%).

[0558] MS m / z (ESI): 978.3 [M+H] + .

[0559] Step 2: Preparation of compound Int9

[0560] Compound Int9-1 (38.0 mg, 38.8 μmol) and diethylamine (21.3 mg, 291.3 μmol) were added to anhydrous N,N-dimethylformamide (2 mL), and the reaction was carried out at room temperature for 2 hours. The reaction solution was concentrated to give the title compound of this step (30.0 mg, yield: 97.0%).

[0561] MS m / z (ESI): 757.2 [M+H] + .

[0562] Intermediate Preparation Example 10: Preparation of Compound Int10

[0563] Following the synthetic route of compound Int6, compound Int4 was replaced with compound Int9 to obtain the title compound (35 mg).

[0564] MS m / z (ESI): 1513.6 [M+H] + .

[0565] Example 11: Preparation of intermediate: Int11

[0566] Step 1: Preparation of compound Int11-2

[0567] Compound Int11-1 (2.43 g, 4.64 mmol), N,N-diisopropylethylamine (1.26 g, 9.28 mmol), and (S)-2-((((9H-fluorene-9-yl)methoxy)carbonyl)amino)-5-(allyloxy)-5-oxovaleric acid (2 g, 4.64 mmol) were dissolved in DMSO (20 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (2.64 g, 6.96 mmol) were added. The reaction mixture was reacted at room temperature for 3 hours. The reaction solution was directly purified by high-performance liquid chromatography to give the title compound of this step (3.5 g, yield: 80.6%).

[0568] MS m / z (ESI): 889.5 [M+H] + .

[0569] Step 2: Preparation of compound Int11-3

[0570] Compound Int11-2 (3.5 g, 3.74 mmol) was dissolved in dichloromethane (20 mL), and trifluoroacetic acid (10 mL) was added. The reaction mixture was reacted at room temperature for 2 hours. The reaction solution was filtered, and the crude product obtained by concentration was purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-50%) to give the title compound of this step (2.7 g, yield 72.4%).

[0571] MS m / z (ESI): 833.4 [M+H] + .

[0572] Step 3: Preparation of compound Int11-4

[0573] Compound Int11-3 (1.1 g, 1.25 mmol) and N,N-diisopropylethylamine (597.38 mg, 4.39 mmol) were dissolved in DMSO (10 mL). Compound Int5-3 (1 g, 1.47 mmol) and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (714.4 mg, 1.88 mmol) were added, and the reaction was carried out at room temperature for 3 hours. The reaction solution was filtered, and the crude product obtained by concentration was purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-50%) to give the title compound of this step (1.2 g, yield: 62.2%).

[0574] MS m / z (ESI): 1461.7 [M+H] + .

[0575] Step 4: Preparation of compound Int11

[0576] Compound Int11-4 (1.2 g, 779.97 μmol) was dissolved in anhydrous N,N-dimethylformamide (10 mL), and 1,3-dimethylbarbituric acid (365.03 mg, 2.34 mmol) and tetrakis(triphenylphosphine)palladium (180.26 mg, 155.99 μmol) were added. The mixture was purged three times with nitrogen and reacted at 20 °C for 4 hours. The reaction solution was filtered, and the crude product obtained by concentration was purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-50%) to give the title compound of this step (1.1 g, yield 94.3%).

[0577] MS m / z (ESI): 1421.7 [M+H] + .

[0578] Example 12: Preparation of intermediate 2-(4-(4-(4-aminophenoxy)piperidin-1-yl)-3-chlorophenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-4-fluoro-1-oxoisoindoline-5-yl)methyl)-2-oxoacetamide (Int12)

[0579] Step 1: Preparation of methyl 2-(3-chloro-4-(4-(4-nitrophenoxy)piperidin-1-yl)phenyl)-2-oxoacetate

[0580] 4-(4-nitrophenoxy)piperidine (250 mg, 0.97 mmol), methyl 2-(3-chloro-4-fluorophenyl)-2-oxoacetate (209 mg, 0.97 mmol), and potassium carbonate (401 mg, 2.90 mmol) were added to anhydrous N,N-dimethylformamide (5 mL), and reacted at 100 °C for 1 hour. The reaction solution was cooled to room temperature, diluted with water, extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous magnesium sulfate, filtered, and the crude product obtained by concentration was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 4 / 1 (v / v)) to give the title compound of this step (210 mg, yield: 49.3%).

[0581] MS m / z (ESI): 419.2 [M+H] + .

[0582] Step 2: Preparation of 2-(3-chloro-4-(4-(4-nitrophenoxy)piperidin-1-yl)phenyl)-2-oxoacetic acid

[0583] 2-(3-chloro-4-(4-(4-nitrophenoxy)piperidin-1-yl)phenyl)-2-oxoacetate methyl ester (195 mg, 442 μmol) and lithium hydroxide monohydrate (92.8 mg, 2.21 mmol) were dissolved in a mixture of tetrahydrofuran (4 mL) and water (2 mL), and reacted at room temperature for 4 hours. The reaction solution was concentrated, and the pH was adjusted to 4–5 with 1 N hydrochloric acid. After filtration, the title compound of this step was obtained (147 mg, yield: 78.0%).

[0584] MS m / z(ESI): 405.0 [M+H] + .

[0585] Step 3: Preparation of 2-(3-chloro-4-(4-(4-nitrophenoxy)piperidin-1-yl)phenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-4-fluoro-1-oxoisoindoline-5-yl)methyl)-2-oxoacetamide

[0586] 3-(5-(aminomethyl)-4-fluoro-1-oxoisoindoline-2-yl)piperidine-2,6-dione hydrochloride (35.98 mg, 123.52 μmol), 2-(3-chloro-4-(4-(4-nitrophenoxy)piperidine-1-yl)phenyl)-2-oxoacetic acid (50 mg, 123.52 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (56.36 mg, 148.22 μmol), and N,N-diisopropylethylamine (47.89 mg, 370.55 μmol) were added to anhydrous N,N-dimethylformamide (1.5 mL), the mixture was purged with nitrogen, and the reaction was carried out at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-60%) to obtain the title compound of this step (45 mg, yield: 48.4%).

[0587] MS m / z (ESI): 678.2 [M+H] + .

[0588] Step 4: Preparation of 2-(4-(4-(4-aminophenoxy)piperidin-1-yl)-3-chlorophenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-4-fluoro-1-oxoisoindoline-5-yl)methyl)-2-oxoacetamide

[0589] 2-(3-chloro-4-(4-(4-nitrophenoxy)piperidin-1-yl)phenyl)-N-((2-(2,6-dioxopiperidin-3-yl)-4-fluoro-1-oxoisoindoline-5-yl)methyl)-2-oxoacetamide (40 mg, 58.99 μmol) and tin dichloride (335.57 mg, 1.77 mmol) were added to ethanol (6 mL) and reacted at 80 °C for 5 hours. The crude product obtained after concentration was purified by high performance liquid chromatography to give the title compound (6.6 mg, yield: 16.4%).

[0590] MS m / z (ESI): 648.2 [M+H] + .

[0591] 1 H NMR (400MHz, DMSO-d6) δ11.02(s,1H),9.55(t,J=8.0Hz,1H),7.98(d,J=2.0Hz,1H),7.93(dd,J=8.0,4.0Hz,1H), 7.61(d,J=8.0Hz,1H),7.59-7.53(m,1H),7.29(d,J=8.0Hz,1H),6.75-6.69(m,2H),6.53-6.48(m,2H),5.13(dd, J=12.0,4.0Hz,1H),4.74-4.55(m,5H),4.41(d,J=16.0Hz,1H),4.34-4.28(m,1H),3.42-3.37(m,2H),3.05(t,J= 8.0Hz,2H),2.96-2.87(m,1H),2.60(d,J=16.0Hz,1H),2.46-2.39(m,1H),2.02-1.98(m,3H),1.78-1.72(m,2H).

[0592] Example 13: Preparation of intermediate: Int13

[0593] Following the synthetic route of compound Int9, compound Int3 was replaced with compound Int12 to obtain the title compound (55 mg).

[0594] MS m / z (ESI): 819.2 [M+H] + .

[0595] Intermediate Preparation Example 14: Preparation of Compound Int14

[0596] Step 1: Preparation of compound Int14-1

[0597] Compound Int13 (54.0 mg, 66.0 μmol), compound Int11 (103 mg, 72.6 μmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (32.6 mg, 85.8 μmol), and N,N-diisopropylethylamine (34.1 mg, 264 μmol) were added to anhydrous N,N-dimethylformamide (2 mL), and the mixture was purged with nitrogen and reacted at room temperature for 2 hours. The reaction solution was then purified directly by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-50%) to give the title compound of this step (100 mg, yield: 64.8%).

[0598] MS m / z(ESI): 1111.2 [M / 2+H] + .

[0599] Step 2: Preparation of compound Int14

[0600] Compound Int14-1 (95.0 mg, 42.8 μmol) and diethylamine (31.3 mg, 428 μmol) were added to anhydrous N,N-dimethylformamide (3 mL) and reacted at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-30%) to give the title compound (75 mg, yield: 83.3%).

[0601] MS m / z(ESI): 1000.2 [M / 2+H] + .

[0602] Intermediate Preparation Example 15: Preparation of 3-(1-bromo-4-oxo-4H-thieno[3,4-c]pyrrole-5(6H)-yl)piperidine-2,6-dione (Int15)

[0603] Following the synthetic route of compound Int1, 2-methylthiophene-3-carboxylic acid was replaced with 4-methylthiophene-3-carboxylic acid to obtain the title compound (104.3 mg).

[0604] MS m / z(ESI): 281.1 [M+H] + .

[0605] Example 16: Preparation of compound Int16

[0606] Step 1: Preparation of 2-((tert-butyldiphenylsilyl)oxy)-N-(2-(2-chloro-4-nitrophenylethoxy)ethyl)-N-methylacetamide

[0607] 2-((tert-butyldiphenylsilyl)oxy)acetic acid (1.82 g, 5.80 mmol), 2-(2-chloro-4-nitrophenylethoxy)-N-methylethane-1-amine (1 g, 3.87 mmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (1.76 g, 4.64 mmol) and N,N-diisopropylethylamine (1.50 g, 11.60 mmol) were added to anhydrous N,N-dimethylformamide (5 mL) and reacted at room temperature for 2 hours. The crude product obtained by concentration of the reaction solution was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 3 / 1 (v / v)) to give the title compound of this step (2 g, yield: 74.6%).

[0608] MS m / z (ESI): 555.2 [M+H] + .

[0609] Step 2: Preparation of N-(2-(4-amino-2-chlorophenethoxy)ethyl)-2-((tert-butyldiphenylsilyl)oxy)-N-methylacetamide

[0610] 2-((tert-butyldiphenylsilyl)oxy)-N-(2-(2-chloro-4-nitrophenylethoxy)ethyl)-N-methylacetamide (1.08 g, 1.95 mmol) and tetrahydroxydiboron (525.18 mg, 5.86 mmol) were dissolved in N,N-dimethylformamide (3 mL), and 4,4'-bipyridine (15.25 mg, 97.63 μmol) was added. The mixture was reacted at room temperature for 5 minutes. The reaction solution was diluted with water, extracted with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the crude product obtained by concentration of the filtrate was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 3 / 2 (v / v)) to give the title compound of this step (940 mg, yield: 82.5%).

[0611] MS m / z (ESI): 525.2 [M+H] + .

[0612] Step 3: Preparation of 2,5-dioxopyrrolidone-1-yl(3-chloro-4-(2,2,7-trimethyl-6-oxo-3,3-diphenyl-4,10-dioxa-7-aza-3-silyldodecane-12-yl)phenyl)carbamate

[0613] N-(2-(4-amino-2-chlorophenethoxy)ethyl)-2-((tert-butyldiphenylsilyl)oxy)-N-methylacetamide (350 mg, 666.47 μmol) was dissolved in dichloromethane (10 mL), cooled to 0 °C, and N,N'-disuccinimidyl carbonate (273.17 mg, 1.07 mmol) was added. The reaction mixture was reacted at room temperature for 2 hours. The reaction solution was filtered, and the filtrate was concentrated to obtain the crude title compound of this step, which was used directly in the next step.

[0614] Step 4: Preparation of (5-(2,6-dioxopiperidin-3-yl)-4-oxo-5,6-dihydro-4H-thieno[3,4-c]pyrrolo-1-yl)methyl(3-chloro-4-(2,2,7-trimethyl-6-oxo-3,3-diphenyl-4,10-dioxa-7-aza-3-sildodecane-12-yl)phenyl)carbamate

[0615] 3-(1-(hydroxymethyl)-4-oxo-4H-thieno[3,4-c]pyrrolo-5(6H)-yl)piperidin-2,6-dione (180.91 mg, 645.42 μmol) and N,N-diisopropylethylamine (166.83 mg, 1.29 mmol) were added to N-methylpyrrolidone (5 mL), the mixture was purged with nitrogen, and 2,5-dioxopyrrolidin-1-yl(3-chloro-4-(2,2,7-trimethyl-6-oxo)-yl)piperidin-2,6-dione was added dropwise. A solution (1 mL) of N-methylpyrrolidone in 3,3-diphenyl-4,10-dioxa-7-aza-3-sildodecane-12-yl)phenyl)carbamate (430 mg, 645.42 μmol) was microwaved at 100 °C for 8 minutes. The reaction solution was then purified directly by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-70%) to give the title compound of this step (330 mg, yield: 61.5%).

[0616] MS m / z (ESI): 831.3 [M+H] + .

[0617] Step 5: Preparation of compound Int16

[0618] (240 mg, 288.65 μmol) of (5-(2,6-dioxopiperidin-3-yl)-4-oxo-5,6-dihydro-4H-thieno[3,4-c]pyrrolo-1-yl)methyl(3-chloro-4-(2,2,7-trimethyl-6-oxo-3,3-diphenyl-4,10-dioxa-7-aza-3-sildodecane-12-yl)phenyl)carbamate was dissolved in methanol (3 mL) and concentrated hydrochloric acid (90.00 μL) and reacted at room temperature for 2 hours. The crude product obtained by concentration of the reaction solution was purified by reverse-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-35%) to give the title compound (130 mg, yield: 75.9%).

[0619] MS m / z (ESI): 593.1 [M+H] + .

[0620] 1 H NMR(400MHz,DMSO-d6)δ(s,1H),9.95(s,1H),8.06(s,1H),7.56(s,1H),7.34-7.20(m,2H),5 .35-5.27(m,2H),5.03(dd,J=12.0,4.0Hz,1H),4.43-4.32(m,2H),4.26(d,J=16.0Hz,1H),4. 07(d,J=4.0Hz,1H),4.02(d,J=4.0Hz,1H),3.59-3.53(m,2H),3.53-3.47(m,2H),3.43(d,J= 4.0Hz,1H),2.88-2.81(m,6H),2.59(d,J=16.0Hz,1H),2.41-2.28(m,2H),2.04-1.95(m,1H).

[0621] Intermediate Preparation Example 17: Preparation of Compound Int17

[0622] Step 1: Preparation of compound Int17-1

[0623] Compound Int16 (120 mg, 202.34 μmol) and (S)-(2-((((9H-fluorene-9-yl)methoxy)carbonyl)amino)propionamido)methyl acetate (386.89 mg, 1.01 mmol) were dissolved in dichloromethane (10 mL), and 4A molecular sieves were added. After stirring for 5 minutes, trifluoroacetic acid (461.43 mg, 4.05 mmol) was added, and the reaction was carried out at room temperature for 3 hours. The reaction solution was filtered, and the crude product obtained by concentration was purified by reverse-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 50%-55%) to give the title compound of this step (50 mg, yield: 27.0%).

[0624] MS m / z (ESI): 915.3 [M+H] + .

[0625] Step 2: Preparation of compound Int17-2

[0626] Compound Int17-1 (50 mg, 54.62 μmol) and diethylamine (23.97 mg, 327.72 μmol) were added to anhydrous N,N-dimethylformamide (3 mL), and the reaction was carried out at room temperature for 1 hour. The reaction solution was concentrated to give the title compound of this step (35 mg, yield: 92.4%).

[0627] MS m / z (ESI): 693.1 [M+H] + .

[0628] Step 3: Preparation of compound Int17-3

[0629] Compound Int17-2 (30 mg, 43.28 μmol), N,N-diisopropylethylamine (11.19 mg, 86.56 μmol), and (((9H-fluorene-9-yl)methoxy)carbonyl)-L-alanyl-L-alanine (33.10 mg, 86.56 μmol) were added to anhydrous N,N-dimethylformamide (3 mL), followed by 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (24.68 mg, 64.92 μmol). The reaction mixture was reacted at room temperature for 1 hour. The reaction solution was then purified directly by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 45%-50%) to give the title compound of this step (40 mg, yield: 87.4%).

[0630] MS m / z (ESI): 1057.2 [M+H] + .

[0631] Step 4: Preparation of compound Int17

[0632] Compound Int17-3 (40 mg, 37.82 μmol) and diethylamine (16.60 mg, 226.94 μmol) were dissolved in anhydrous N,N-dimethylformamide (3 mL) and reacted at room temperature for 1 hour. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 20%-25%) to give the title compound (21 mg, yield: 66.5%).

[0633] MS m / z (ESI): 835.3 [M+H] + .

[0634] Intermediate Preparation Example 18: Preparation of Compound Int18

[0635] Step 1: Preparation of compound Int18-2

[0636] Compound Int18-1 (2 g, 8.04 mmol), N,N-disuccinimidyl carbonate (6.17 g, 24.11 mmol), and triethylamine (2.44 g, 24.11 mmol) were dissolved in dimethyl sulfoxide (30 mL) under ice bath conditions and reacted at room temperature for 2 hours. Then, 2-amino-2-(hydroxymethyl)propane-1,3-diol (2.95 g, 24.11 mmol) was added, and the reaction was carried out at room temperature for 3 hours. The reaction solution was purified by high performance liquid chromatography to give the title compound of this step (350 mg, yield: 8.1%).

[0637] MS m / z (ESI): 541.3 [M+H] + .

[0638] Step 2: Preparation of compound Int18

[0639] Compound Int18-2 (350 mg, 647.42 μmol) was dissolved in dichloromethane (1 mL), and a 1,4-dioxane solution of hydrogen chloride (4 M, 1 mL) was added. The reaction was carried out at room temperature for 1 hour. The reaction solution was concentrated to give the title compound (300 mg, yield: 97.2%).

[0640] MS m / z (ESI): 441.3 [M+H] + .

[0641] Intermediate Preparation Example 19: Preparation of Compound Int19

[0642] Step 1: Preparation of compound Int19-2

[0643] Compound Int19-1 (100 mg, 276.65 μmol) and (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(allyloxy)-5-oxovaleric acid (113.27 mg, 276.65 μmol) were dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (157.79 mg, 414.98 μmol) and N,N-diisopropylethylamine (107.26 mg, 829.95 μmol) were added. The reaction mixture was reacted at room temperature for 4 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-75%) to give the title compound of this step (181 mg, yield: 86.9%).

[0644] MS m / z (ESI): 753.4 [M+H] + .

[0645] Step 2: Preparation of compound Int19-3

[0646] Compound Int19-2 (181 mg, 240.41 μmol) was dissolved in anhydrous dichloromethane (6 mL), and trifluoroacetic acid (2 mL) was added. The reaction was carried out at room temperature for 3 hours. The crude product obtained by concentration of the reaction solution was purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-55%) to give the title compound of this step (136 mg, yield: 88.3%).

[0647] MS m / z (ESI): 641.3 [M+H] + .

[0648] Step 3: Preparation of compound Int19-4

[0649] Compounds Int19-3 (50 mg, 78.04 μmol) and Int18 (128.74 mg, 234.13 μmol) were dissolved in N,N-dimethylformamide (2 mL), and N,N,N′,N′-tetramethyl-O-(7-azabenzotriazol-1-yl)hexafluorophosphate urea (119.9 mg, 312.17 μmol) and N,N-diisopropylethylamine (71.32 mg, 546.3 μmol) were added. The reaction mixture was reacted at room temperature for 14 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-25%) to give the title compound of this step (60 mg, yield: 51.8%).

[0650] MS m / z (ESI): 1485.8 [M+H] + .

[0651] Step 4: Preparation of compound Int19-5

[0652] Compound Int19-4 (60 mg, 40.39 μmol), 1,3-dimethylbarbituric acid (12.61 mg, 80.77 μmol), and tetrakis(triphenylphosphine)palladium (4.66 mg, 4.04 μmol) were added to N,N-dimethylformamide (2 mL), and the mixture was purged with nitrogen and reacted at room temperature for 1 hour. The reaction solution was purified by preparative high-performance liquid chromatography to give the title compound of this step (22.0 mg, yield: 37.7%).

[0653] MS m / z (ESI): 1445.7 [M+H] + .

[0654] Step 5: Preparation of compound Int19-6

[0655] Compounds Int19-5 (22 mg, 15.22 μmol) and Int9 (11.51 mg, 15.22 μmol) were dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (8.77 mg, 22.83 μmol) and N,N-diisopropylethylamine (7.95 mg, 60.88 μmol) were added. The reaction mixture was reacted at room temperature for 14 hours. The reaction solution was purified by preparative high-performance liquid chromatography to obtain the title compound of this step (18 mg, yield: 54.2%).

[0656] MS m / z (ESI): 1902.6 [M / 2+H] + .

[0657] Step 6: Preparation of compound Int19

[0658] Compound Int19-6 (13 mg, 4.46 μmol) and diethylamine (1.0 mg, 13.67 μmol) were dissolved in N,N-dimethylformamide (2 mL) and reacted at room temperature for 4 hours. The reaction solution was purified by preparative high performance liquid chromatography to obtain the title compound (5 mg, yield: 55.8%).

[0659] MS m / z(ESI): 1960.9 [M+H] + .

[0660] Intermediate Preparation Example 20: Preparation of Compound Int20

[0661] Step 1: Preparation of compound Int20-2

[0662] Compound Int20-1 (200 mg, 857.26 μmol) and (S)-2-((((9H-fluorene-9-yl)methoxy)carbonyl)amino)-5-(allyloxy)-5-oxovaleric acid (350.99 mg, 857.26 μmol) were dissolved in N,N-dimethylformamide (3 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (380.0 mg, 999.40 μmol) and N,N-diisopropylethylamine (332.0 mg, 2.57 mmol) were added. The reaction mixture was reacted at room temperature for 4 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-50%) to give the title compound of this step (350 mg, yield: 67.8%).

[0663] MS m / z (ESI): 625.3 [M+H] + .

[0664] Step 2: Preparation of compound Int20-3

[0665] Compound Int20-2 (350 mg, 560.25 μmol) was dissolved in anhydrous dichloromethane (6 mL), and trifluoroacetic acid (2 mL) was added. The reaction was carried out at room temperature for 3 hours. The crude product obtained by concentration of the reaction solution was purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-45%) to give the title compound of this step (305 mg, yield: 95.7%).

[0666] MS m / z (ESI): 569.2 [M+H] + .

[0667] Step 3: Preparation of compound Int20-4

[0668] Compounds Int19-1 (100 mg, 276.65 μmol) and Int20-3 (157.3 mg, 276.11 μmol) were dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (152.95 mg, 402.26 μmol) and N,N-diisopropylethylamine (107.26 mg, 829.95 μmol) were added. The reaction mixture was reacted at room temperature for 4 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-65%) to give the title compound of this step (183 mg, yield: 72.5%).

[0669] MS m / z (ESI): 912.5 [M+H] + .

[0670] Step 4: Preparation of compound Int20-5

[0671] Compound Int20-4 (183 mg, 200.64 μmol) was dissolved in anhydrous dichloromethane (6 mL), and trifluoroacetic acid (2 mL) was added. The reaction was carried out at room temperature for 3 hours. The crude product obtained by concentration of the reaction solution was purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-35%) to give the title compound of this step (160 mg, yield: 99.7%).

[0672] MS m / z (ESI): 800.4 [M+H] + .

[0673] Step 5: Preparation of compound Int20-6

[0674] Compounds Int20-5 (80 mg, 100.02 μmol) and Int18 (128.74 mg, 234.13 μmol) were dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (96.03 mg, 252.56 μmol) and N,N-diisopropylethylamine (71.32 mg, 546.3 μmol) were added. The reaction mixture was reacted at room temperature for 14 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-30%) to give the title compound of this step (82 mg, yield: 49.8%).

[0675] MS m / z (ESI): 1644.9 [M+H] + .

[0676] Step 6: Preparation of compound Int20-7

[0677] Compound Int20-6 (82 mg, 49.85 μmol), 1,3-dimethylbarbituric acid (15.57 mg, 99.71 μmol), and tetrakis(triphenylphosphine)palladium (5.76 mg, 4.99 μmol) were added to N,N-dimethylformamide (2 mL), purged with nitrogen, and reacted at room temperature for 1 hour. The reaction solution was purified by preparative high-performance liquid chromatography to give the title compound of this step (65.0 mg, yield: 81.3%).

[0678] MS m / z (ESI): 1604.8 [M+H] + .

[0679] Step 7: Preparation of compound Int20-8

[0680] Compounds Int20-7 (65 mg, 40.5 μmol) and Int9 (35.25 mg, 46.61 μmol) were dissolved in N,N-dimethylformamide (2 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (23.34 mg, 61.38 μmol) and N,N-diisopropylethylamine (21.15 mg, 163.65 μmol) were added. The reaction mixture was reacted at room temperature for 14 hours. The reaction solution was purified by preparative high-performance liquid chromatography to obtain the title compound of this step (24 mg, yield: 25.3%).

[0681] MS m / z (ESI): 1171.1 [M / 2+H] + .

[0682] Step 8: Preparation of compound Int20

[0683] Compound Int20-8 (24 mg, 10.24 μmol) and diethylamine (1.69 mg, 23.05 μmol) were dissolved in N,N-dimethylformamide (1.5 mL) and reacted at room temperature for 4 hours. The reaction solution was purified by preparative high performance liquid chromatography to obtain the title compound (14 mg, yield: 84.1%).

[0684] MS m / z (ESI): 1060.7 [M / 2+H] + .

[0685] Intermediate Preparation Example 21: Preparation of Compound Int21

[0686] Following the synthetic route of compound Int20, compound Int20-1 was replaced with compound Int21-1 to obtain the title compound (10 mg).

[0687] MS m / z (ESI): 1104.7 [M+H] + .

[0688] Intermediate Preparation Example 22: Preparation of Compound Int22

[0689] Step 1: Preparation of compound Int22-2

[0690] (S)-2-((((9H-fluorene-9-yl)methoxy)carbonyl)amino)-5-(allyloxy)-5-oxovaleric acid (1.0 g, 2.44 mmol), N,N-diisopropylethylamine (0.63 g, 4.88 mmol), and compound Int22-1 (0.78 g, 2.44 mmol) were dissolved in dimethyl sulfoxide (10 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (1.39 g, 3.66 mmol) was added. The reaction mixture was reacted at room temperature for 3 hours. The reaction mixture was extracted dropwise into water, washed with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and the crude product obtained by concentration was purified by reversed-phase column chromatography (Preparation method D, elution gradient: mobile phase A% = 5%-30%) to give the title compound of this step (1.3 g, yield: 74.7%).

[0691] MS m / z (ESI): 713.4 [M+H] + .

[0692] Step 2: Preparation of compound Int22-3

[0693] Compound Int22-2 (1.3 g, 1.81 mmol) was dissolved in dichloromethane (15 mL), and trifluoroacetic acid (8 mL) was added. The reaction mixture was reacted at room temperature for 2 hours. The reaction solution was filtered, and the crude product obtained by concentration was purified by reversed-phase column chromatography (preparation method D, elution gradient: mobile phase A% = 5%-50%) to give the title compound of this step (1.1 g, yield: 91.7%).

[0694] MS m / z (ESI): 657.3 [M+H] + .

[0695] Step 3: Preparation of compound Int22-4

[0696] Compounds Int19-1 (300 mg, 788.44 μmol) and Int22-3 (545.04 mg, 788.44 μmol) were dissolved in N,N-dimethylformamide (10 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (458.86 mg, 1.18 mmol) and N,N-diisopropylethylamine (321.78 mg, 2.37 mmol) were added. The reaction mixture was reacted at room temperature for 4 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-50%) to give the title compound of this step (620 mg, yield: 74.7%).

[0697] MS m / z (ESI): 1001.4 [M+H] + .

[0698] Step 4: Preparation of compound Int22-5

[0699] Compound Int22-4 (620 mg, 619.89 μmol) was dissolved in anhydrous dichloromethane (6 mL), and trifluoroacetic acid (2 mL) was added. The reaction was carried out at room temperature for 3 hours. The crude product obtained by concentration of the reaction solution was purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-35%) to give the title compound of this step (530 mg, yield: 91.5%).

[0700] MS m / z (ESI): 888.3 [M+H] + .

[0701] Step 5: Preparation of compound Int22-6

[0702] Compounds Int22-5 (300 mg, 334.47 μmol) and Int5-3 (546.21 mg, 836.18 μmol) were dissolved in N,N-dimethylformamide (5 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (282.44 mg, 735.84 μmol) and N,N-diisopropylethylamine (174.66 mg, 1.34 mmol) were added. The reaction mixture was reacted at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-35%) to give the title compound of this step (620 mg, yield: 74.7%).

[0703] MS m / z (ESI): 1073.8 [M / 2+H] + .

[0704] Step 6: Preparation of compound Int22

[0705] Compound Int22-6 (180 mg, 83.9 μmol), 1,3-dimethylbarbituric acid (26.2 mg, 167.81 μmol), and tetrakis(triphenylphosphine)palladium (9.69 mg, 8.39 μmol) were added to N,N-dimethylformamide (2 mL), and the mixture was purged with nitrogen and reacted at room temperature for 1 hour. The reaction solution was purified by preparative high-performance liquid chromatography to give the title compound (110 mg, yield: 59.2%).

[0706] MS m / z (ESI): 1053.1 [M+H] + .

[0707] Example 23: Preparation of intermediate 23: Preparation of compound Int23

[0708] Step 1: Preparation of compound Int23-1

[0709] Compounds Int22 (10 mg, 4.75 μmol) and Int9 (4.13 mg, 5.46 μmol) were dissolved in N,N-dimethylformamide (1 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (2.74 mg, 7.2 μmol) and N,N-diisopropylethylamine (2.48 mg, 19.19 μmol) were added. The reaction mixture was reacted at room temperature for 14 hours. The reaction solution was purified by preparative high-performance liquid chromatography to obtain the title compound of this step (6 mg, yield: 44.4%).

[0710] MS m / z (ESI): 1422.2 [M / 2+H] + .

[0711] Step 2: Preparation of compound Int23

[0712] Compound Int23-1 (6 mg, 2.11 μmol) and diethylamine (463.0 μg, 6.33 μmol) were dissolved in N,N-dimethylformamide (0.5 mL) and reacted at room temperature for 4 hours. The reaction solution was purified by preparative high performance liquid chromatography to obtain the title compound (3 mg, yield: 53.3%).

[0713] MS m / z (ESI): 874.3 [M / 3+H] + .

[0714] Intermediate Preparation Example 24: Preparation of Compound Int24

[0715] Step 1: Preparation of compound Int24-1

[0716] Compound Int3 (118.27 mg, 201.81 μmol) and (((9H-fluorene-9-yl)methoxy)carbonyl)glycine (60.0 mg, 201.81 μmol) were dissolved in N,N-dimethylformamide (1 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (115.1 mg, 302.72 μmol) and N,N-diisopropylethylamine (78.25 mg, 605.44 μmol) were added. The mixture was reacted at room temperature for 2 hours, and the reaction solution was used directly in the next step.

[0717] MS m / z (ESI): 865.3 [M+H] + .

[0718] Step 2: Preparation of compound Int24

[0719] Compound Int24-1 (140 mg, 161.79 μmol) and diethylamine (35.3 mg, 482.59 μmol) were dissolved in N,N-dimethylformamide (1 mL) and reacted at room temperature for 4 hours. The reaction solution was then purified directly by reversed-phase column chromatography (Preparation method C, elution gradient: mobile phase A% = 5%-35%) to give the title compound of this step (34 mg, yield: 31.2%).

[0720] MS m / z (ESI): 643.2 [M+H] + .

[0721] Intermediate Preparation Example 25: Preparation of Compound Int25

[0722] Step 1: Preparation of compound Int25-2

[0723] Compound Int25-1 (700 mg, 1.23 mmol) was dissolved in N,N-dimethylformamide (6 mL), cooled to 0 °C, and N,N-diisopropylethylamine (617.95 mg, 4.78 mmol) and benzyl bromide (613.34 mg, 3.59 mmol) were added. The mixture was reacted at room temperature for 14 hours. Water was added to the reaction solution, and the mixture was filtered. The filter cake was slurried with petroleum ether, filtered, and dried to give the title compound of this step (620 mg, yield: 76.4%).

[0724] MS m / z (ESI): 558.4 [M+H-100] + .

[0725] Step 2: Preparation of compound Int25-3

[0726] Compound Int25-2 (620 mg, 942.54 μmol) was dissolved in anhydrous dichloromethane (10 mL), and trifluoroacetic acid (4 mL) was added. The mixture was reacted at room temperature for 3 hours. The reaction solution was concentrated to obtain the title compound of this step (600 mg, yield: 94.8%).

[0727] MS m / z (ESI): 558.3 ​​[M+H] + .

[0728] Step 3: Preparation of compound Int25-4

[0729] Compound Int25-3 (595 mg, 1.07 mmol) was dissolved in tetrahydrofuran (7 mL), purged with nitrogen, and N,N-diisopropylethylamine (551.55 mg, 4.27 mmol) was added. The mixture was cooled to 0 °C, and a tetrahydrofuran solution of triphosgene (189.97 mg, 640.17 μmol) was added dropwise. The reaction was carried out at 0 °C for 3 hours. The crude product obtained by concentrating the reaction solution was used directly in the next step of the reaction.

[0730] MS m / z (ESI): 584.3 [M+H] + .

[0731] Step 4: Preparation of compound Int25-5

[0732] Compound Int25-4 (255 mg, 705.45 μmol) was dissolved in N,N-dimethylformamide (4 mL), and a solution of N,N-diisopropylethylamine (287.91 mg, 2.23 mmol) and compound Int19-1 (617.63 mg, 1.06 mmol) in N,N-dimethylformamide (4 mL) was added. The reaction was carried out at room temperature for 3 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-55%) to give the title compound of this step (460 mg, yield: 69.0%).

[0733] MS m / z (ESI): 945.5 [M+H] + .

[0734] Step 5: Preparation of compound Int25-6

[0735] Compound Int25-5 (460 mg, 486.70 μmol) was dissolved in anhydrous dichloromethane (8 mL), and trifluoroacetic acid (4 mL) was added. The reaction was carried out at room temperature for 3 hours. The crude product obtained by concentration of the reaction solution was purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 5%-35%) to give the title compound of this step (360 mg, yield: 88.8%).

[0736] MS m / z (ESI): 833.4 [M+H] + .

[0737] Step 6: Preparation of compound Int25-7

[0738] Compounds Int25-6 (280 mg, 336.16 μmol) and Int18 (370.20 mg, 840.4 μmol) were dissolved in N,N-dimethylformamide (5 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (387.33 mg, 1.01 mmol) and N,N-diisopropylethylamine (526.61 mg, 4.07 mmol) were added. The reaction mixture was reacted at room temperature for 4 hours. The reaction solution was purified by preparative high-performance liquid chromatography to give the title compound of this step (120 mg, yield: 21.3%).

[0739] MS m / z (ESI): 1677.9 [M+H] + .

[0740] Step 7: Preparation of compound Int25-8

[0741] Compound Int25-7 (120 mg, 71.52 μmol) was dissolved in methanol (6 mL), and 10% palladium on carbon (36.00 mg) was added. The mixture was then purged with hydrogen and reacted at room temperature for 1 hour. The reaction solution was filtered through diatomaceous earth, and the crude product obtained by concentration was purified by preparative high performance liquid chromatography to give the title compound of this step (47 mg, yield: 41.4%).

[0742] MS m / z (ESI): 1587.8 [M+H] + .

[0743] Step 8: Preparation of compound Int25-9

[0744] Compounds Int25-8 (40 mg, 25.19 μmol) and Int24 (16.2 mg, 25.19 μmol) were dissolved in N,N-dimethylformamide (1 mL), and 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (14.37 mg, 37.79 μmol) and N,N-diisopropylethylamine (9.77 mg, 75.58 μmol) were added. The reaction mixture was reacted at room temperature for 2 hours. The reaction solution was purified by preparative high-performance liquid chromatography to obtain the title compound of this step (10 mg, yield: 17.9%).

[0745] MS m / z(ESI): 1107.0 [M / 2+H] + .

[0746] Step 9: Preparation of compound Int25

[0747] Compound Int25-9 (10 mg, 4.52 μmol) and diethylamine (1.0 mg, 13.67 μmol) were dissolved in N,N-dimethylformamide (1 mL) and reacted at room temperature for 4 hours. The reaction solution was purified by preparative high performance liquid chromatography to obtain the title compound (7 mg, yield: 76.1%).

[0748] MS m / z (ESI): 995.9 [M / 2+H] + .

[0749] Drug linker preparation example 1-1: Preparation of compound C-1

[0750] Compound Int4 (26 mg, 36.16 μmol) and 2,5-dioxopyrrolidone-1-yl 6-(2-(methanesulfonyl)pyrimidin-5-yl)hex-5-acetylacetate (15.8 mg, 43.39 μmol) were dissolved in N,N-dimethylformamide (3 mL), and N,N-diisopropylethylamine (7 mg, 54.24 μmol) was added. The reaction mixture was reacted at 0 °C for 2 h. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 25%-30%) to give the title compound of this step (25 mg, yield: 71.3%).

[0751] MS m / z (ESI): 970.2 [M+H] + .

[0752] Drug linker preparation example 1-2: Preparation of compound C-2

[0753] Compound Int6 (45 mg, 30.45 μmol) and 2,5-dioxopyrrolidone-1-yl 6-(2-(methanesulfonyl)pyrimidin-5-yl)hex-5-acetylacetate (14.5 mg, 39.58 μmol) were dissolved in N,N-dimethylformamide (3 mL), and N,N-diisopropylethylamine (11.8 mg, 91.34 μmol) was added. The reaction mixture was reacted at 0 °C for 2 h. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 25%-30%) to give the title compound of this step (28.5 mg, yield: 54.2%).

[0754] MS m / z (ESI): 1727.5 [M+H] + .

[0755] Drug linker preparation examples 1-3: Preparation of compound C-3

[0756] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int7 to obtain the title compound (20 mg, yield: 34.2%).

[0757] MS m / z (ESI): 1085.3 [M+H] + .

[0758] Drug linker preparation examples 1-4: Preparation of compound C-4

[0759] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int8 to obtain the title compound (5 mg, yield: 33.2%).

[0760] MS m / z (ESI): 1843.1 [M+H] + .

[0761] Drug linker preparation examples 1-5: Preparation of compound C-5

[0762] Compound Int9 (30.0 mg, 39.7 μmol), 2,5-dioxopyrrolidone-1-yl 6-(2-(methanesulfonyl)pyrimidin-5-yl)hex-5-acetylacetate (17.4 mg, 47.6 μmol), and N,N-diisopropylethylamine (10.3 mg, 79.4 μmol) were added to anhydrous N,N-dimethylformamide (1.5 mL), and the reaction was carried out at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 10%-90%) to give the title compound of this step (21.4 mg, yield: 50.9%).

[0763] MS m / z (ESI): 1006.3 [M+H] + .

[0764] Drug linker preparation examples 1-6: Preparation of compound C-6

[0765] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int10 to obtain the title compound (20 mg, yield: 57.2%).

[0766] MS m / z (ESI): 882.1 [M / 2+H] + .

[0767] Drug linker preparation examples 1-7: Preparation of compound C-7

[0768] Step 1: Preparation of compound C-7-1

[0769] Compound Int11 (100 mg, 63.31 μmol) and diethylamine (9.35 mg, 126.62 μmol) were added to N,N-dimethylformamide (0.5 mL) and reacted at room temperature for 2 hours. The reaction solution was concentrated to obtain the title compound of this step (80 mg, crude product).

[0770] MS m / z (ESI): 1199.6 [M+H] + .

[0771] Step 2: Preparation of compound C-7-2

[0772] Compound C-7-1 (80 mg, 60.04 μmol) and 2,5-dioxopyrrolidone-1-yl 6-(2-(methanesulfonyl)pyrimidin-5-yl)hex-5-acetylacetate (24.37 mg, 60.04 μmol) were added to N,N-dimethylformamide (0.5 mL) and reacted at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 20%-25%) to give the title compound of this step (55 mg, yield: 56.8%).

[0773] MS m / z (ESI): 1449.6 [M+H] + .

[0774] Step 3: Preparation of compound C-7

[0775] Compound C-7-2 (31.09 mg, 19.83 μmol), N-hydroxysuccinimide (1.86 mg, 14.55 μmol), N,N'-diisopropylcarbodiimide (2.78 mg, 19.83 μmol), and compound Int9 (10 mg, 13.22 μmol) were added to N,N-dimethylformamide (0.5 mL) and reacted at room temperature for 2 hours. The reaction solution was directly purified by reversed-phase column chromatography (Preparation method B, elution gradient: mobile phase A% = 30%-40%) to give the title compound of this step (5 mg, yield: 16.1%).

[0776] MS m / z (ESI): 1094.9 [M / 2+H] + .

[0777] Drug linker preparation examples 1-8: Preparation of compound C-8

[0778] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int14 to obtain the title compound (51 mg, yield: 57.4%).

[0779] MS m / z(ESI): 1125.0 [M / 2+H] + .

[0780] Drug linker preparation examples 1-9: Preparation of compound C-9

[0781] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int17 to obtain the title compound (51 mg, yield: 57.4%).

[0782] MS m / z (ESI): 1085.3 [M+H] + .

[0783] Drug linker preparation examples 1-10: Preparation of compound C-10

[0784] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int19 to obtain the title compound (4.88 mg, yield: 84.2%).

[0785] MS m / z (ESI): 1106.2 [M / 2+H] + .

[0786] Drug linker preparation example 1-11: Preparation of compound C-11

[0787] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int20 to obtain the title compound (10.48 mg, yield: 68.4%).

[0788] MS m / z (ESI): 1185.7 [M / 2+H] + .

[0789] Drug linker preparation examples 1-12: Preparation of compound C-12

[0790] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int21 to obtain the title compound (8.21 mg, yield: 62.7%).

[0791] MS m / z (ESI): 1230.1 [M / 2+H] + .

[0792] Drug linker preparation example 1-13: Preparation of compound C-13

[0793] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int23 to obtain the title compound (1.45 mg, yield: 44.9%).

[0794] MS m / z (ESI): 1436.7 [M / 2+H] + .

[0795] Drug linker preparation examples 1-14: Preparation of compound C-14

[0796] Following the synthetic route of compound C-1, compound Int4 was replaced with compound Int25 to obtain the title compound (2.9 mg, yield: 37.7%).

[0797] MS m / z (ESI): 1120.7 [M / 2+H] + .

[0798] II. Preparation of Degrading Agent Antibody Conjugate (DAC)

[0799] Adjust the stock solution of monoclonal antibodies (Trastuzumab, Pertrzumab, or Ab-05) to pH 7.570 using 1M disodium hydrogen phosphate solution. Add 0.1M EDTA and TCEP (5.5 eq, 10 mM) to the resulting stock solution, mix well, and react at room temperature for 2 hours. Then, add DMSO solution (10 eq, 10 mM) of drug linkers C-1 to C-14 to the reduced monoclonal antibody buffer solution, mix well, and react at room temperature for 2 hours. Purify using a NAP-5 gel column and wash with histidine hydrochloride buffer solution at pH 6.0-6.1. Collect the filtrate to obtain histidine hydrochloride buffer for the degradation agent antibody conjugate (DAC), and store at -20°C.

[0800] III. Drug / Antibody Ratio of Degrading Agent Antibody Conjugates (DACs): Determination of DAR Value

[0801] The molecular weight of DAC was determined by LC-MS, and the drug / antibody ratio (DAR) was calculated.

[0802] Chromatographic determination conditions:

[0803] Liquid chromatography column: Thermo MAbPac RP 3.0*100mm;

[0804] Mobile phase A: 0.1% FA / H2O; Mobile phase B: 0.1% FA / ACN;

[0805] Flow rate: 0.25 ml / min; Sample chamber temperature: 8℃; Column temperature: 60℃; Injection volume: 1 μl;

[0806] Mass spectrometry determination conditions:

[0807] Mass spectrometer model: AB Sciex Triple TOF 5600+;

[0808] GS1 35; GS2 35; CUR 30; TEM 350; ISVF 5500; DP 250; CE 10; Accumulation time 0.5s;

[0809] m / z 600-4000; Time bins to sum 40.

[0810] The average DAR value of the degradation agent antibody conjugate (DAC) was calculated using CE-SDS, as shown in Table 1:

[0811] Table 1. Average DAR Value Test Results of Degrading Agent Antibody Conjugate (DAC)

[0812] Biological tests

[0813] Experimental Example 1: Inhibition of Proliferative Activity of Degradative Antibody-Drug Conjugate (DAC) on BT474 Breast Cancer Cells

[0814] 1. Experimental System:

[0815] Cell Name / Source: BT474 / ATCC

[0816] Reagent Name / Manufacturer: CellCounting-Lite 2.0 Luminescent Cell Viability Assay, Nanjing Novizan Biotechnology Co., Ltd.

[0817] 2. Experimental parameters:

[0818] Cell count: 3000 cells / well; Culture medium: DMEM + 10% FBS; Incubation conditions: 37℃, 5% CO2; Incubation time: 120 hours; Detection temperature: room temperature; Detection instrument: TECAN SPARK microplate reader.

[0819] 3. Experimental steps:

[0820] BT474 cells were cultured in vitro as a monolayer in DMEM medium with 10% FBS at 37°C in an incubator containing 5% CO2. Cells were seeded into 96-well plates at 3000 cells / well and cultured overnight. After incubation, pre-diluted antibody-delegate inhibitor (DAC) was added. DMSO was added to the negative control group, and culture medium was added to the blank control group. After incubation for 120 hours, the assay reagent was added to each well. 2.0, read the chemiluminescence unit value (RLU) in the chemiluminescence detection mode of the microplate reader.

[0821] 4. Data Processing:

[0822] Calculate the percentage inhibition rate of different concentrations of the test substance using the following formula:

[0823] Percentage inhibition rate = (1 - (chemiluminescence signal value of test substance - chemiluminescence signal value of blank control) / (chemiluminescence signal value of negative control - chemiluminescence signal value of blank control)) × 100%. The half-maximal inhibitory concentration (IC50) of the test substance is calculated by fitting a curve using a four-parameter model. 50 ).

[0824] 5. Experimental Results:

[0825] The inhibitory activity of the degradation agent antibody-drug conjugate (DAC) on the proliferation of BT474 cells was determined according to the above method, and the results are shown in Table 2.

[0826] Table 2. Inhibition of BT474 cell proliferation activity by the degradation agent antibody-drug conjugate (DAC).

[0827] Conclusion: The DAC of this application has strong cell proliferation inhibitory activity against BT474 cells.

[0828] Experiment Example 2: Inhibition of Proliferative Activity of Degrading Agent Antibody Conjugate (DAC) on Small Cell Lung Cancer Cell NCI-H146

[0829] 1. Experimental System:

[0830] Cell Name / Source: NCI-H146 / ATCC

[0831] Reagent Name / Manufacturer: CellCounting-Lite 2.0 Luminescent Cell Viability Assay, Nanjing Novizan Biotechnology Co., Ltd.

[0832] 2. Experimental parameters:

[0833] Cell count: 8000 cells / well; Culture medium: RPMI 1640 + 10% FBS; Incubation conditions: 37℃, 5% CO2; Incubation time: 96 hours; Detection temperature: room temperature; Instrument: TECAN SPARK microplate reader.

[0834] 3. Experimental steps:

[0835] NCI-H146 cells were cultured in suspension in vitro under the following conditions: RPMI 1640 + 10% FBS, 37°C, and an incubator containing 5% CO2. Cells were seeded into 96-well plates at 8000 cells / well, and then pre-diluted test reagents were added. DMSO was added to the negative control group, and culture medium was added to the blank control group. After incubation for 96 hours, the test reagent was added to each well. 2.0, read the chemiluminescence unit value (RLU) in the chemiluminescence detection mode of the microplate reader.

[0836] 4. Data Processing:

[0837] Calculate the percentage inhibition rate of different concentrations of the test substance using the following formula:

[0838] Percentage inhibition rate = (1 - (chemiluminescence signal value of test substance - chemiluminescence signal value of blank control) / (chemiluminescence signal value of negative control - chemiluminescence signal value of blank control)) × 100%. The half-maximal inhibitory concentration (IC50) of the test substance is calculated by fitting a curve using a four-parameter model. 50 ).

[0839] 5. Experimental Results:

[0840] The inhibitory activity of the degradation agent antibody-drug conjugate (DAC) on the proliferation of NCI-H146 cells was determined according to the above method, and the results are shown in Table 3.

[0841] Table 3. Results of inhibition of NCI-H146 cell proliferation activity by the degradation agent antibody-drug conjugate (DAC).

[0842] Conclusion: The DAC of this application has strong cell proliferation inhibitory activity against NCI-H146 cells.

[0843] Experimental Example 3: Degradation of GSPT1 protein in BT474 breast cancer cells by antibody-drug conjugate (DAC).

[0844] 1. Experimental System:

[0845] Cell Name / Source: BT474 / ATCC

[0846] 2. Experimental parameters

[0847] Cell count: 6*10 5 Cells / well, culture medium: DMEM + 10% FBS, test substance incubation conditions: 37℃, 5% CO2, incubation time: 24 hours, detection temperature: room temperature, testing instrument: ultra-high sensitivity chemiluminescence imaging system - manufacturer Bio-rad.

[0848] 3. Experimental steps:

[0849] BT474 cells were cultured in vitro as a monolayer in 10% FBS + DMEM medium at 37°C in an incubator containing 5% CO2. Cells were seeded into 6-well plates one day in advance, at a density of 6 x 10⁶ cells / well. 5 Cells / well; on the second day, add pre-diluted test sample, and add culture medium to the blank control group, incubate for 24 hours. Collect cells, add lysis buffer (Cell Signaling Technology 9803), lyse on ice for 25-30 minutes, centrifuge for 10 minutes, and collect the supernatant protein sample. Use Pierce... TM Protein quantification was performed using the BCA Protein Assay Kit (Thermo Scientific 23225). Protein concentration was normalized, and 1X loading buffer was added. The mixture was then denatured at 95°C for 10 minutes.

[0850] Take an appropriate amount of protein sample and perform SDS-PAGE gel electrophoresis. Electrophoresis for 30-50 minutes. After the protein sample is pressed into a line, continue electrophoresis until the loading buffer reaches the bottom of the separating gel, then stop electrophoresis.

[0851] After electrophoresis, proteins were transferred to a PVDF membrane using a wet transfer method, with transfer time of 45–90 minutes. The transferred PVDF membrane was then placed in blocking buffer and blocked for 1–2 hours. Bands were then cut from the PVDF membrane according to the molecular weight of the target proteins (GSPT-1 and internal control β-actin), and incubated with the corresponding primary antibodies overnight at 4°C with shaking. The membrane was washed three times with TBS containing 0.1% Tween 20. The membrane was then incubated with secondary antibody solution (horseradish peroxidase-labeled goat anti-rabbit or goat anti-mouse IgG, diluted 1:1000–5000 in 0.1% Tween 20 TBS) at room temperature for 1 hour. After washing three times as before, the membrane was developed using Western chemiluminescent HRP Substrate (Millipore / WBKL S0500) and photographed using an ultra-high sensitivity chemiluminescence imaging system. Grayscale values ​​of the GSPT1 and β-actin bands were obtained separately.

[0852] 4. Data Processing:

[0853] Protein degradation rate (D) = (1-G) 目的蛋白 / G0)×100%; where G 目的蛋白 The gray density (target protein band) / gray density (β-actin band) of the target protein degradation were observed in different concentrations of the test substance treatment group. G0 = gray density (target protein band) / gray density (corresponding β-actin band) of the blank control group.

[0854] 5. The experimental results are shown in Table 4.

[0855] Table 4. Degradative activity of the antibody-drug conjugate (DAC) in BT474 cells against GSPT1 protein degradation.

[0856] Among them, "++++" indicates a degradation effect between 100% and 80%; "++++" indicates a degradation effect between 80% and 60%; "+++" indicates a degradation effect between 60% and 30%; "++" indicates a degradation effect between 30% and 10%; and "+" indicates a degradation effect <10%.

[0857] The DACs of this application, such as DAC-1-A, DAC-2-A, DAC-3-A and DAC-6-A, have good degradation effects on GSPT1 protein.

[0858] Experiment Example 4: In vivo efficacy evaluation of the test substance in a BT474 cell CDX mouse model

[0859] 1. Experimental System:

[0860] Cell Name / Source: BT474 / ATCC

[0861] 2. Experimental parameters:

[0862] Number of cells inoculated: 1*10 7 Individuals / animal, inoculation medium: serum-free DMEM medium: matrix gel concentration = 6:4

[0863] 3. Experimental steps:

[0864] 100 μL of human breast cancer BT474 cells per mouse were subcutaneously injected into the right axilla of NOD SCID mice. One day prior to inoculation, estrogen tablets were administered to the backs of the mice. Eight days after inoculation, when the average tumor volume reached 150 mm², the mice were targeted for further treatment. 3Around this time, according to the measurement results, tumor-bearing mice with regular tumor shapes and uniform volumes were selected. The number of mice included in the group was determined according to the actual situation, and they were randomly divided into 6 groups. Screening criteria: Remove mice with irregular tumor shapes (such as long strip, branched shapes, etc.), overly large or small tumor volumes, and too light body weights. The remaining animals after grouping were euthanized at the end of the experiment. Administration started on the day of grouping. The test substance was administered by intravenous injection, once only. The tumor volume and body weight were measured 2 times a week, and the data were recorded.

[0865] 4. Data processing:

[0866] The formula for calculating the tumor volume (V): V = 1 / 2 × a × b 2 , where a and b represent the length and width respectively. The anti-tumor drug efficacy was evaluated by the tumor growth inhibition rate TGI (%), and the calculation formula: TGI (%) (瘤体积) = [1 - (T Vt - T V0 ) / (C Vt - C V0 )] × 100%, T V0 is the average tumor volume of the test substance group at the time of grouping and administration, T Vt is the average tumor volume of the test substance group t days after administration; C V0 is the average tumor volume of the vehicle group at the time of grouping and administration; C Vt is the average tumor volume of the vehicle group t days after administration. When the tumor regresses, TGI (%) (瘤体积) = 100% - (T [[ID=二十九]] Vt - T V0 ) / T V0 × 100%. <L

[0867] If the tumor shrinks compared to the initial volume, that is, when Vt < V0, it is defined as partial tumor regression (PR); if the tumor completely disappears, it is defined as complete tumor regression (CR).

[0868] 5. Experimental results:

[0869] After administration, each test substance could significantly inhibit tumor growth. Compared with the vehicle control group, at the dose of 5 mg / kg, the TGI of DAC-3-A 36 days after administration was 174.43%; the TGI of DAC-1-A 36 days after administration was 176.10%; the TGI of DAC-2-A 36 days after administration was 195.06%; the TGI of DAC-6-A 36 days after administration was 194.01% (Table 5). The results showed that in the subcutaneous xenograft tumor model of BT474 cells, DAC-1-A, DAC-3-A, DAC-2-A, and DAC-6-A had significant anti-tumor effects.

[0870] Table 5. Efficacy of the degradation agent antibody-drug conjugate (DAC) in BT474 cell subcutaneous xenografts.

[0871] PR: indicates partial regression; CR: indicates complete regression.

[0872] Although specific embodiments of this disclosure have been described in detail, those skilled in the art will understand that various modifications and substitutions can be made to those details based on all the teachings disclosed, and such changes are all within the scope of this disclosure. The full scope of this disclosure is given by the appended claims and any equivalents thereof.

Claims

1. A compound of formula (I) or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof: in, Ab 1 is a targeting group; M represents the junction site that connects to the target group; L is the connector that connects M and D; m is selected from 1-20; D is selected from the structures of formulas (DI-1) and (DI-2); X 1 is selected from CH2and -C(=O)-; X 2 X 3 X 4 Each is independently selected from O, S, Se, N, C, and CR. 3 And when X 2 X 3 X 4 When one of the elements is independently selected from O, S, and Se, the other two are independently selected from N, C, and CR. 3 ; It can be a single bond or a double bond; X 5 selected from O, S, C 1-6 alkylene and C 1-6 haloalkylene; Ring A is selected from phenyl and 5-6-membered heteroaryl groups; Y is selected from C 6-10 Aryl and 5-10 heteroaryl groups, wherein the aryl and heteroaryl groups are optionally separated by one or more R groups. 8 replace; Z is selected from C 6-10 aryl and 5-10 membered heteroaryl, said aryl and heteroaryl being substituted with R 7 substituted at the same time, optionally with one or more R 9 substituted; L 1 Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Haloalkylene -, -O-(CH2) e -、-N(R a )-(CH2) e -、-(CH2-CH2-O) f -、C 3-10 Cycloalkyl groups, 3-12 membered heterocyclic groups, and divalent structures composed of one or more of the above structural units; R 1 Selected from C 6-10 Aryl and 5-10 heteroaryl, said aryl and heteroaryl being -L a -L b -L c -R 5 While replacing, it can be optionally replaced by one or more R 4 replace; R 2 selected from H and C 1-6 alkyl; R 3 selected from H, halogen, hydroxyl, cyano, C 1-6 alkyl, C 1-6 haloalkyl and -O-C 1-6 alkyl; R 4 Each is independently selected from H, halogen, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 6 replace; R 5 selected from R 6 Each is independently selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Halogenated alkyl groups, -OC 1-6 Alkyl, C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more groups selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2- 6-acetylinyl, C 1-6 Haloalkyl, C 1-6 Alkoxy and C 1-6 Substituents of haloalkoxy groups; L a Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Halogenated alkylene groups -, -(CH2) e -O-、-(CH2) e -N(R a )-、-(CH2-CH2-O) f -、-(CH2-CH2-O) f -(CH2) e -N(R a )-、-(CH2) e -O-(CH2) e -N(R a - and -(CH2) e -C(=O)-O-; L b Selected from covalent bonds, C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 Aryl and 5-10 heteroaryl groups; L c selected from a covalent bond, -O-, -S-, -S(=O)-, -S(=O)2-, -N-(R a ), -, -C(=O)N(R a ), -, -C(=S)N(R a ), -, -C(=O)-, -C(=S)-, -C(=O)-O-, -O-C(=O)-O-, -O-C(=O)-N(R a ), -, -N(R a ), -C(=O)-N(R a ), -, -S(=O)2-N(R a ), -, -S(=O)2-NH-C(=O)-O- and -S(=O)2-NH-C(=O)-N(R a ); R 8 Each is independently selected from H, halogen, cyano, nitro, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b -OR a -C 1-6 Alkylene-OR a -C 1-6 Alkylene-N(R) a )R b C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 10 replace; R 10 Each is independently selected from H, halogen, cyano, C 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b and -OR a ; R 7 selected from R 9 Each is independently selected from H, halogen, cyano, and C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b -OR a C 3-10 Cycloalkyl, 3-12 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 11 replace; R 11 Each is independently selected from H, halogen, cyano, and C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 1-6 Haloalkyl, -N(R) a )R b and -OR a ; R a Each is independently selected from H and C. 1-6 alkyl; R b Each is independently selected from H and C. 1-6 alkyl; z is selected from 1, 2, and 3; f are each independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; e is independently selected from 0, 1, 2, 3, 4, 5, and 6; In this context, position 1 of each substituent indicates the connection point with L.

2. The compound of claim 1 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein, M is selected from the following structures: Where, the 2nd position of M is related to Ab 1 Connected, position 3 is connected to L; each a is independently selected from 1, 2, 3, 4, 5 and 6, and b is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10; Preferably, M is selected from the following structures:

3. The compound of any one of claims 1-2 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein, L is selected from non-breakable linkers and breakable linkers, wherein the breakable linker is cleaved by an enzyme present in the pathological environment, wherein the enzyme is selected from proteases, phosphatases, pyrophosphatases, β-glucuronidase, β-galactosidase and sulfatase. Preferably, L is -L 2 -L 3 -L 4 -,in: L 2 is a covalent bond or is selected from the group consisting of: Preferably, L 2 is a covalent bond or selected from the following structures: R 12 Selected from H and -L 2a -L 2b -(Q) n ,in; L 2a is a covalent bond or is selected from the group consisting of: L 2b It is a covalent bond or selected from the following structures: Preferably, L 2b It is a covalent bond or selected from the following structures: Q is a covalent bond or is selected from the group consisting of the following structures: Where n is selected from 1, 2 and 3; each c is independently selected from 1, 2, 3, 4, 5 and 6; and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10. L 3 Fragments selected from amino acids or their derivatives and peptide fragments formed from two or more amino acids or their derivatives, wherein the amino acids are selected from Val, Cit, Glu, Lys, Arg, Phe, Leu, Gly, Ala, and Asn, and the amino acid derivatives are selected from... R' is independently selected from hydrogen and C. 1-6 Alkyl, -C 1-6 Alkylene-OC 1-6 Alkyl, -C 1-6 Alkylene-NHC 1-6 Alkyl, -C 1-6 Alkylene-N(C) 1-6 Alkyl)2; L 4 is a covalent bond or is selected from the group consisting of:

4. The compound of any one of claims 1-3 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug, wherein L is -L 2 -L 3 -L 4 -,in, L 2 is a covalent bond or is selected from the group consisting of: Preferably, L 2 is a covalent bond or selected from the following structures: R 12 Selected from H and -L 2a -L 2b -(Q) n ,in; L 2a is a covalent bond or is selected from the group consisting of: L 2b is a covalent bond or is selected from: Preferably, L 2b is a covalent bond or selected from: Q is a covalent bond or selected from the following structures: Where n is selected from 1, 2 and 3; each c is independently selected from 1, 2, 3, 4, 5 and 6; and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10. Preferably, L 2 is a covalent bond or selected from the group consisting of: Preferably, L 2 is a covalent bond or selected from the group consisting of: L 3 selected from the group consisting of: Preferably, L 3 selected from the following structures: Preferably, L 3 selected from the following structures: Preferably, L 4 is a covalent bond or 5. The compound of any one of claims 1-4 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein, L is selected from the following structures: In this case, position 4 of L is connected to M, and position 5 is connected to D; each c is independently selected from 1, 2, 3, 4, 5 and 6, and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10. Preferably, L is selected from the following structures: In this case, position 4 of L is connected to M, and position 5 is connected to D; L is selected from the following structures: In this case, position 4 of L is connected to M, and position 5 is connected to D; each c is independently selected from 1, 2, 3, 4, 5 and 6, and each d is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10. Preferably, L is selected from the following structures: In this case, position 4 of L is connected to M, and position 5 is connected to D.

6. The compound of any one of claims 1-5 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein, D is selected from the structures of formulas (D-II-1), (D-II-2), (D-II-3), (D-II-4), (D-II-5), (D-II-6), (D-II-7), (D-II-8), (D-II-9), and (D-II-10): Among them, X 2 X 3 X 4 Each is independently selected from O, S, and Se; X 5 R 1 R 2 R 3 Y, Z, L 1 And z as defined in claim 1.

7. The compound of any one of claims 1-6 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein, D is selected from the structures of formula (D-IV-1), formula (D-IV-2), and formula (D-IV-3): Among them, X 5 R 1 R 2 R 3 Y, Z, L 1 As defined in claim 1; Preferably, X 5 Selected from O, S, C 1-6 Alkylene and C 1-6 Halogenated alkylene; Y is selected from C 6-10 Aryl and 5-10 heteroaryl groups, wherein the aryl and heteroaryl groups are optionally separated by one or more R groups. 8 replace; Z is selected from C 6-10 Aryl and 5-10 heteroaryl groups, said aryl and heteroaryl groups being R 7 While replacing, it can be optionally replaced by one or more R 9 replace; L 1 Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Haloalkylene -, -O-(CH2) e -、-N(R a )-(CH2) e -、-(CH2-CH2-O) f -、C 3-10 Cycloalkyl groups, 3-12 membered heterocyclic groups, and divalent structures composed of one or more of the above structural units; R 1 Selected from C 6-10 Aryl and 5-10 heteroaryl, said aryl and heteroaryl being -L a -L b -L c -R 5 While replacing, it can be optionally replaced by one or more R 4 replace; R 2 Selected from H and C 1-4 alkyl; R 3 Selected from H, halogen, hydroxyl, cyano, C 1-4 Alkyl, C 1-4 Halogenated alkyl groups and -OC 1-4 alkyl; R 4 Each is independently selected from H, halogen, amino, C 1-6 Alkyl, 3-10 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 6 replace; R 5 Selected from R 6 Each is independently selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-6 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Halogenated alkyl groups, -OC 1-6 Alkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more groups selected from H, halogen, hydroxyl, amino, cyano, nitro, C 1-4 Alkyl, C 2-4 alkenyl, C 2- 4-Alynyl group, C 1-4 Haloalkyl, C 1-4 Alkoxy and C 1-4 Substituents of haloalkoxy groups; L a Selected from C 1-6 Alkylene, C 2-6 imidene group, C 2-6 Ethyne group, -C 1-6 Halogenated alkylene groups -, -(CH2) e -O-、-(CH2) e -N(R a )-、-(CH2-CH2-O) f -、-(CH2-CH2-O) f -(CH2) e -N(R a - and -(CH2) e -O-(CH2) e -N(R a )-; L b is selected from a covalent bond and a 3-10 membered heterocyclyl; L c Select your own key, -O-, -S-, -S(=O)-, -S(=O)2-, -N-(R a )-,-C(=O)N(R a )-, -C(=O)-, -C(=O)-O-, -S(=O)2-N(R a )-sum-S(=O)2-NH-C(=O)-O-; R 8 Each is independently selected from H, halogen, cyano, nitro, C 1-4 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Haloalkyl, -N(R) a )R b -OR a -C 1-4 Alkylene-OR a -C 1-4 Alkylene-N(R) a )R b C 3-8 Cycloalkyl, 3-10 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 10 replace; R 10 Each is independently selected from H, halogen, cyano, and C. 1-4 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Haloalkyl, -N(R) a )R b and -OR a ; R 7 selected from R 9 Each is independently selected from H, halogen, cyano, and C. 1-4 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Haloalkyl, -N(R) a )R b -OR a C 3-8 Cycloalkyl, 3-10 membered heterocyclic groups, C 6-10 aryl and 5-10-membered heteroaryl groups, wherein the alkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl groups are optionally surrounded by one or more R groups. 11 replace; R 11 Each is independently selected from H, halogen, cyano, C 1-4 Alkyl, C 2-4 alkenyl, C 2-4 alkynyl group, C 1-4 Haloalkyl, -N(R) a )R b and -OR a ; R a each independently selected from H and C 1-4 alkyl; R b each independently selected from H and C 1-4 alkyl; f are each independently selected from 1, 2, 3, 4, 5, and 6; e is independently selected from 0, 1, 2, 3, 4 and 5; Wherein, position 1 of each substituent indicates the connection point with L; Preferably, D is selected from the structures of the above formulas (D-IV-1) and (D-IV-2).

8. The compound of any one of claims 1-7 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein, D is selected from the following structures Preferably, D is selected from the following structures 9. The compound of any one of claims 1-8 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein Ab 1 is a targeting group that targets a cell surface receptor or a tumor surface antigen.

10. The compound of any one of claims 1-9 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein Ab 1 The antibody is an antibody or its antigen-binding fragment; the antibody is selected from monoclonal antibodies, polyclonal antibodies, linear antibodies, bispecific antibodies, multispecific antibodies, chimeric antibodies, murine antibodies, humanized antibodies, fully human antibodies, and fusion proteins containing the antigen-binding portion of the antibody; the antigen-binding fragment is selected from Fab, Fab', F(ab')2, Fv, disulfide-linked Fv and scFv.

11. The compound of any one of claims 1-10 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug thereof, wherein the antibody or antigen-binding fragment thereof is an anti-Her-2 antibody or an antigen-binding fragment thereof. Preferably, the antibody or its antigen-binding fragment comprises: (1) The following heavy chain variable regions (VH) and / or light chain variable regions (VL): (1a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:28 or a variant thereof, CDR-H2 with sequence SEQ ID NO:29 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or, (1b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:43 or a variant thereof, CDR-H2 with sequence SEQ ID NO:44 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof; or, (2) The following heavy chain variable regions (VH) and / or light chain variable regions (VL): (2a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence SEQ ID NO:41 or a variant thereof, CDR-H2 with the sequence SEQ ID NO:42 or a variant thereof, and CDR-H3 with the sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with the sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with the sequence SEQ ID NO:33 or a variant thereof; or, (2b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:56 or a variant thereof, CDR-H2 with sequence SEQ ID NO:57 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof; or, (3) The following heavy chain variable regions (VH) and / or light chain variable regions (VL): (3a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:34 or a variant thereof, CDR-H2 with sequence SEQ ID NO:35 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:30 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:31 or a variant thereof, CDR-L2 with sequence SEQ ID NO:32 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or, (3b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:49 or a variant thereof, CDR-H2 with sequence SEQ ID NO:50 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:45 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:46 or a variant thereof, CDR-L2 with sequence SEQ ID NO:47 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof; or, (4) The following heavy chain variable regions (VH) and / or light chain variable regions (VL): (4a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:36 or a variant thereof, CDR-H2 with sequence SEQ ID NO:37 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:38 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:39 or a variant thereof, CDR-L2 with sequence SEQ ID NO:40 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:33 or a variant thereof; or, (4b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:51 or a variant thereof, CDR-H2 with sequence SEQ ID NO:52 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:53 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:54 or a variant thereof, CDR-L2 with sequence SEQ ID NO:55 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:48 or a variant thereof; wherein The variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with its source sequence, or the variant has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to its source sequence; preferably, the substitutions are conservative substitutions; Preferably, the antibody or its antigen-binding fragment comprises: (a) VH or a variant thereof shown in SEQ ID NO: 24, and / or VL or a variant thereof shown in SEQ ID NO: 25; or (b) VH or a variant thereof shown in SEQ ID NO: 26, and / or VL or a variant thereof shown in SEQ ID NO: 27; The variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with its source sequence, or the variant has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to its source sequence; preferably, the substitutions are conservative substitutions; More preferably, the antibody or its antigen-binding fragment comprises: (1) A heavy chain comprising the VH region of the sequence shown in SEQ ID NO: 24 and the heavy chain constant region (CH) shown in SEQ ID NO: 58, and a light chain comprising the VL region of the sequence shown in SEQ ID NO: 25 and the light chain constant region (CL) shown in SEQ ID NO: 59; or (2) A heavy chain including the VH of the sequence shown in SEQ ID NO: 26 and the heavy chain constant region (CH) shown in SEQ ID NO: 58, and a light chain including the VL of the sequence shown in SEQ ID NO: 27 and the light chain constant region (CL) shown in SEQ ID NO: 59; More preferably, the antibody or its antigen-binding fragment comprises: (1) The heavy chain comprising the sequence shown in SEQ ID NO: 60, and the light chain comprising the sequence shown in SEQ ID NO: 61; or (2) The heavy chain comprising the sequence shown in SEQ ID NO: 62, and the light chain comprising the sequence shown in SEQ ID NO: 63; Optionally, the N-terminal glutamine of the VH or variant thereof of the sequence shown in SEQ ID NO:24 or 26, or the heavy chain or variant thereof of the sequence shown in SEQ ID NO:60 or 62, undergoes cyclization to form pyroglutamic acid or pyroglutamic acid salt; and / or, the constant region (CH) of the heavy chain of the sequence shown in SEQ ID NO:58, or the heavy chain or variant thereof of the sequence shown in SEQ ID NO:60 or 62, lacks a C-terminal lysine residue; More preferably, the antibody is a trastuzumab antibody, a pertuzumab antibody, or a dual epitope antibody composed of these.

12. The compound of any one of claims 1-10 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug thereof, wherein the antibody or antigen-binding fragment thereof is an anti-B7H3 antibody or an antigen-binding fragment thereof. Preferably, the antibody or its antigen-binding fragment comprises: (1) The following heavy chain variable regions (VH) and / or light chain variable regions (VL): (1a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:11 or a variant thereof, CDR-H2 with sequence SEQ ID NO:12 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof; or (1b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence of SEQ ID NO:11 or a variant thereof, CDR-H2 with the sequence of SEQ ID NO:68 or a variant thereof, and CDR-H3 with the sequence of SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence of SEQ ID NO:6 or a variant thereof, CDR-L2 with the sequence of SEQ ID NO:7 or a variant thereof, and CDR-L3 with the sequence of SEQ ID NO:5 or a variant thereof; or (2) The following heavy chain variable regions (VH) and / or light chain variable regions (VL): (2a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO:14 or a variant thereof, CDR-H2 of SEQ ID NO:15 or a variant thereof, and CDR-H3 of SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO:6 or a variant thereof, CDR-L2 of SEQ ID NO:7 or a variant thereof, and CDR-L3 of SEQ ID NO:5 or a variant thereof; or (2b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:14 or a variant thereof, CDR-H2 with sequence SEQ ID NO:69 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof; or (3) The following heavy chain variable regions (VH) and / or light chain variable regions (VL): (3a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO:8 or a variant thereof, CDR-H2 of SEQ ID NO:9 or a variant thereof, and CDR-H3 of SEQ ID NO:10 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO:3 or a variant thereof, CDR-L2 of SEQ ID NO:4 or a variant thereof, and CDR-L3 of SEQ ID NO:5 or a variant thereof; or (3b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:8 or a variant thereof, CDR-H2 with sequence SEQ ID NO:67 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:10 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:3 or a variant thereof, CDR-L2 with sequence SEQ ID NO:4 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof; or (4) The following heavy chain variable regions (VH) and / or light chain variable regions (VL): (4a) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with sequence SEQ ID NO:16 or a variant thereof, CDR-H2 with sequence SEQ ID NO:17 or a variant thereof, and CDR-H3 with sequence SEQ ID NO:13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with sequence SEQ ID NO:6 or a variant thereof, CDR-L2 with sequence SEQ ID NO:7 or a variant thereof, and CDR-L3 with sequence SEQ ID NO:5 or a variant thereof; or (4b) A heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with the sequence of SEQ ID NO:16 or a variant thereof, CDR-H2 with the sequence of SEQ ID NO:70 or a variant thereof, and CDR-H3 with the sequence of SEQ ID NO:13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with the sequence of SEQ ID NO:6 or a variant thereof, CDR-L2 with the sequence of SEQ ID NO:7 or a variant thereof, and CDR-L3 with the sequence of SEQ ID NO:5 or a variant thereof; Preferably, the antibody or its antigen-binding fragment comprises: (a) VH or a variant thereof shown in SEQ ID NO:23 or 1, and / or VL or a variant thereof shown in SEQ ID NO:2; or (b) VH or a variant thereof shown in SEQ ID NO:66, and / or VL or a variant thereof shown in SEQ ID NO:2; Preferably, the antibody or its antigen-binding fragment comprises: (a) The heavy chain of the VH or variant thereof shown in SEQ ID NO:23 or 1 and the heavy chain constant region (CH) or variant thereof shown in SEQ ID NO:18, and / or, the light chain of the VL or variant thereof shown in SEQ ID NO:2 and the light chain constant region (CL) or variant thereof shown in SEQ ID NO:19; or (b) The heavy chain of the VH or variant thereof shown in SEQ ID NO:1 and the heavy chain constant region (CH) or variant thereof shown in SEQ ID NO:71, and / or the light chain of the VL or variant thereof shown in SEQ ID NO:2 and the light chain constant region (CL) or variant thereof shown in SEQ ID NO:19; (c) The VH or variant thereof shown in SEQ ID NO:66 and the heavy chain of the heavy chain constant region (CH) or variant thereof shown in SEQ ID NO:71 or 18, and / or the VL or variant thereof shown in SEQ ID NO:2 and the light chain constant region (CL) or variant thereof shown in SEQ ID NO:19; Preferably, the amino acid sequence of the heavy chain of the antibody or its antigen-binding fragment is as shown in SEQ ID NO:22, 20, 72, 73 or 74, and the amino acid sequence of the light chain of the antibody or its antigen-binding fragment is as shown in SEQ ID NO:

21. Optionally, the N-terminal glutamine of the heavy chain or heavy chain variable region and / or the light chain or light chain variable region undergoes cyclization to form pyroglutamic acid or pyroglutamate; and / or, the heavy chain or heavy chain constant region (CH) or its variants do not contain a C-terminal lysine. Preferably, the variant has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it originates, or has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to the sequence from which it originates; preferably, the substitutions are conservative substitutions.

13. The compound of any one of claims 1-12, or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, selected from the following structures: Each m is selected from 1 to 20, for example 1 to 10, and Ab-(S-) is an antibody or an antigen-binding fragment thereof as defined in any one of claims 10-12.

14. A compound of formula (II) or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof: M 1 -L-D II in, L and D are as defined in any one of claims 1-13. M 1 precursors for the attachment site of the targeting group; Preferably, the targeting group is as defined in any one of claims 1-13; Preferably, M 1 selected from the group consisting of Wherein, each 'a' is independently selected from 1, 2, 3, 4, 5, and 6, and 'b' is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; LG represents a leaving group, preferably selected from halogens (e.g., F, Cl, Br, I) and halogenated C. 1-6 Alkyl, C 1-6 Alkyl sulfonyl, halogenated C 1-6 Alkyl sulfonyl, halosulfonyl, C 1-6 Alkyl sulfonate group, halogenated C 1-6 Alkyl sulfonate group, C 1-6 Alkyl sulfinate group, C 1-6 Structural fragments containing alkyl sulfoxide, halophenoxy, hydroxy, mercapto, amino, nitro, azide, cyano, alkenyl, alkynyl, and alkynyl groups, more preferably C10. 1-6 Alkyl sulfonyl group, for example, methanesulfonyl group; More preferably, M 1 selected from (For example )。 15. The compound of claim 14, or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein the compound is selected from the following structures:

16. A composition comprising the compound of any one of claims 1-13 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug thereof, wherein the composition has a DAR value of 1.0-20.0, preferably 1.0-10.0, and the following values ​​are also present: about 1 to 2, about 1 to 3, about 1 to 4, about 1 to 5, about 1 to 6, about 1 to 7, about 1 to 8, about 1 to 9, about 1 to 10, about 2 to 3, about 2 to 4, about 2 to 5, about 2 to 6, about 2 to 7, about 2 to 8, about 2 to 9, about 2 to 10, about 3 to 4, about 3 to 5, about 3 to 6, about 3 to 7, about 3 to 8, about 3 to 9, about 2 to 10, about 3 to 4, about 3 to 5, about 3 to 6, about 3 to 7, about 3 to 8, about 3 to 9, about 3 to 10, about 4 to 5, about 4 to 10. 6, about 4 to 7, about 4 to 8, about 4 to 9, about 4 to 10, about 5 to 6, about 5 to 7, about 5 to 8, about 5 to 9, about 5 to 10, about 6 to 7, about 6 to 8, about 6 to 9, about 6 to 10, about 7 to 8, about 7 to 9, about 7 to 10, about 8 to 9, about 8 to 10 or about 9 to 10, more preferably 4.0 to 8.0, for example, 4.0 to 4.5 4.0~5.0, 4.0~5.5, 4.0~6.0, 4.0~6.5, 4.0~7.0, 4.0~7.5, 4.0~8.0, 5.0~5.5, 5.0~6.0, 5.0~6.5, 5.0~7.0, 5.0~7.5, 5.0~8.0, 5.5~6.0, 5.5~6.5, 5.5~ 7.0, 5.5–7.5, 5.5–8.0, 6.0–6.5, 6.0–7.0, 6.0–7.5, 6.0–8.0, 6.5–7.0, 6.5–7.5, 6.5–8.0, 7.0–7.5, 7.0–8.0, 7.5–8.0, for example, approximately 4.0, approximately 4.01, approximately 4.02, approximately 4.03, Approximately 4.04, 4.05, 4.06, 4.07, 4.08, 4.09, 4.1, 4.11, 4.12, 4.13, 4.14, 4.15, 4.16, 4.17, 4.18, 4.19, 4.2, 4.21, 4.22, 4.23, 4.24, approximately 4.25, approximately 4.26, approximately 4.27, approximately 4.28, approximately 4.29, approximately 4.3, approximately 4.31, approximately 4.32, approximately 4.33, approximately 4.34, approximately 4.35, approximately 4.36, approximately 4.37, approximately 4.38, approximately 4.39, approximately 4.4, approximately 4.41, approximately 4.42, approximately 4.43, approximately 4.44, approximately 4.45, approximately 4 .46, about 4.47, about 4.48, about 4.49, about 4.5, about 4.51, about 4.52, about 4.53, about 4.54, about 4.55, about 4.56, about 4.57, about 4.58, about 4.59, about 4.6, about 4.61, about 4.62, about 4.63, about 4.64, about 4.65, about 4.66, about 4.67, approximately 4.68, approximately 4.69, approximately 4.7, approximately 4.71, approximately 4.72, approximately 4.73, approximately 4.74, approximately 4.75, approximately 4.76, approximately 4.77, approximately 4.78, approximately 4.79, approximately 4.8, approximately 4.81, approximately 4.82, approximately 4.83, approximately 4.84, approximately 4.85, approximately 4.86, approximately 4.87, approximately 4. 88, approximately 4.89, approximately 4.9, approximately 4.91, approximately 4.92, approximately 4.93, approximately 4.94, approximately 4.95, approximately 4.96, approximately 4.97, approximately 4.98, approximately 4.99, approximately 5.0, approximately 5.01, approximately 5.02, approximately 5.03, approximately 5.04, approximately 5.05, approximately 5.06, approximately 5.07, approximately 5.08, approximately 5.0 9. Approximately 5.1, 5.11, 5.12, 5.13, 5.14, 5.15, 5.16, 5.17, 5.18, 5.19, 5.2, 5.21, 5.22, 5.23, 5.24, 5.25, 5.26, 5.27, 5.28, 5.29, 5.3 Approximately 5.31, 5.32, 5.33, 5.34, 5.35, 5.36, 5.37, 5.38, 5.39, 5.4, 5.41, 5.42, 5.43, 5.44, 5.45, 5.46, 5.47, 5.48, 5.49, 5.5, 5.51 Approximately 5.52, 5.53, 5.54, 5.55, 5.56, 5.57, 5.58, 5.59, 5.6, 5.61, 5.62, 5.63, 5.64, 5.65, 5.66, 5.67, 5.68, 5.69, 5.7, 5.71, 5.

72. Approximately 5.73, 5.74, 5.75, 5.76, 5.77, 5.78, 5.79, 5.8, 5.81, 5.82, 5.83, 5.84, 5.85, 5.86, 5.87, 5.88, 5.89, 5.9, 5.91, 5.92, 5.93, approximately 5.94, approximately 5.95, approximately 5.96, approximately 5.97, approximately 5.98, approximately 5.99, approximately 6.0, approximately 6.01, approximately 6.02, approximately 6.03, approximately 6.04, approximately 6.05, approximately 6.06, approximately 6.07, approximately 6.08, approximately 6.09, approximately 6.1, approximately 6.11, approximately 6.12, approximately 6.13, approximately 6.14, approximately 6 .15, about 6.16, about 6.17, about 6.18, about 6.19, about 6.2, about 6.21, about 6.22, about 6.23, about 6.24, about 6.25, about 6.26, about 6.27, about 6.28, about 6.29, about 6.3, about 6.31, about 6.32, about 6.33, about 6.34, about 6.35, about 6.36, approximately 6.37, approximately 6.38, approximately 6.39, approximately 6.4, approximately 6.41, approximately 6.42, approximately 6.43, approximately 6.44, approximately 6.45, approximately 6.46, approximately 6.47, approximately 6.48, approximately 6.49, approximately 6.5, approximately 6.51, approximately 6.52, approximately 6.53, approximately 6.54, approximately 6.55, approximately 6.56, approximately 6.57, approximately 6.58, approximately 6.59, approximately 6.6, approximately 6.61, approximately 6.62, approximately 6.63, approximately 6.64, approximately 6.65, approximately 6.66, approximately 6.67, approximately 6.68, approximately 6.69, approximately 6.7, approximately 6.71, approximately 6.72, approximately 6.73, approximately 6.74, approximately 6.75, approximately 6.76, approximately 6. 77, approximately 6.78, approximately 6.79, approximately 6.8, approximately 6.81, approximately 6.82, approximately 6.83, approximately 6.84, approximately 6.85, approximately 6.86, approximately 6.87, approximately 6.88, approximately 6.89, approximately 6.9, approximately 6.91, approximately 6.92, approximately 6.93, approximately 6.94, approximately 6.95, approximately 6.96, approximately 6.97, approximately 6.98, approximately 6.99, approximately 7.0, approximately 7.01, approximately 7.02, approximately 7.03, approximately 7.04, approximately 7.05, approximately 7.06, approximately 7.07, approximately 7.08, approximately 7.09, approximately 7.1, approximately 7.11, approximately 7.12, approximately 7.13, approximately 7.14, approximately 7.15, approximately 7.16, approximately 7.17, approximately 7.1 8, approximately 7.19, approximately 7.2, approximately 7.21, approximately 7.22, approximately 7.23, approximately 7.24, approximately 7.25, approximately 7.26, approximately 7.27, approximately 7.28, approximately 7.29, approximately 7.3, approximately 7.31, approximately 7.32, approximately 7.33, approximately 7.34, approximately 7.35, approximately 7.36, approximately 7.37, approximately 7.38, approximately 7.39, approximately 7.4, approximately 7.41, approximately 7.42, approximately 7.43, approximately 7.44, approximately 7.45, approximately 7.46, approximately 7.47, approximately 7.48, approximately 7.49, approximately 7.5, approximately 7.51, approximately 7.52, approximately 7.53, approximately 7.54, approximately 7.55, approximately 7.56, approximately 7.57, approximately 7.58, approximately 7.5 9, approximately 7.6, approximately 7.61, approximately 7.62, approximately 7.63, approximately 7.64, approximately 7.65, approximately 7.66, approximately 7.67, approximately 7.68, approximately 7.69, approximately 7.7, approximately 7.71, approximately 7.72, approximately 7.73, approximately 7.74, approximately 7.75, approximately 7.76, approximately 7.77, approximately 7.78, approximately 7.79, approximately 7.8, approximately 7.81, approximately 7.82, approximately 7.83, approximately 7.84, approximately 7.85, approximately 7.86, approximately 7.87, approximately 7.88, approximately 7.89, approximately 7.9, approximately 7.91, approximately 7.92, approximately 7.93, approximately 7.94, approximately 7.95, approximately 7.96, approximately 7.97, approximately 7.98, approximately 7.99, approximately 8.

0. .

17. A pharmaceutical composition comprising a compound according to any one of claims 1-15 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug, or the composition according to claim 16, and one or more pharmaceutical excipients.

18. Use of the compound of any one of claims 14-15 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug thereof in the preparation of a conjugate (e.g., the compound of any one of claims 1-13 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug thereof).

19. Use of the compound of any one of claims 1-15 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite or prodrug, or composition of claim 16 or 17 in the preparation of a medicament, particularly in a medicament for the treatment and / or prevention of cancer (e.g., cancers associated with GSPT1 protein dysregulation, cancers associated with HER2 expression and / or cancers associated with B7H3 expression); Preferably, the cancer is a GSPT1 protein overexpressing cancer, a HER2 positive cancer, or a B7H3 positive cancer; Preferably, the cancer is selected from gastric cancer, breast cancer, or lung cancer (e.g., small cell lung cancer and non-small cell lung cancer).

20. A method of treating and / or preventing cancer (e.g., cancer associated with GSPT1 protein dysregulation, cancer associated with HER2 expression, and / or cancer associated with B7H3 expression), comprising administering to a subject in need a therapeutically and / or preventively effective amount of the compound of any one of claims 1-15 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, N-oxide, isotopically labeled compound, metabolite, or prodrug, or the composition of claim 16 or 17; Preferably, the cancer is a GSPT1 protein overexpressing cancer, a HER2 positive cancer, or a B7H3 positive cancer; Preferably, the cancer is selected from gastric cancer, breast cancer, or lung cancer (e.g., small cell lung cancer or non-small cell lung cancer).

21. PG 1 -L 3 -L 4 -D、PG 1 -L 2b -(Q) n PG 1 -L 2a -L 2b -(Q) n PG 1 -L 2 -PG 3 or PG 1 -LD represents the compound or its salts, esters, stereoisomers, tautomers, polymorphs, solvates, N-oxides, and isotopically labeled compounds; wherein L 2 L 3 L 4 L 2a L 2b L, Q, n, and D are as defined in any one of claims 1-13; PG 3 is -O-PG 2 ; PG 1 Each group is independently an H or amino protecting group, such as alkoxycarbonyl amino protecting groups, for example benzyloxycarbonyl (Cbz), tert-butoxycarbonyl (Boc), methoxycarbonyl (Fmoc), allyloxycarbonyl (Alloc), trimethylsilylethoxycarbonyl (Teoc), methoxycarbonyl (or ethoxycarbonyl); acyl amino protecting groups, such as phthaloyl (Pht), p-toluenesulfonyl (Tos), trifluoroacetyl (Tfa), o-(p-)nitrobenzenesulfonyl (Ns), pentanoyl, benzoyl, tert-butoxycarbonyl, 9-fluorenylmethoxycarbonyl, allyloxycarbonyl, trichloroethoxycarbonyl, trimethylsilylethoxycarbonyl, benzyloxycarbonyl, p-methylbenzenesulfonyl, p-nitrobenzenesulfonyl, trifluoroacetyl, methoxycarbonyl, or ethoxycarbonyl; alkyl amino protecting groups, such as triphenylmethyl (Trt), C 1-6 Alkyl-substituted triphenylmethyl, p-methoxytriphenylmethyl (MMT), dimethoxytriphenylmethyl (DMT), 2,4-dimethoxybenzyl (Dmb), 4-methoxybenzyl (PMB), benzyl (Bn); PG 2 Each is independently protected by an H or a carboxyl group, the carboxyl protecting group being, for example, a C group. 1-6 Alkyl, allyl, benzyl, 2,4-dimethoxybenzyl, p-methoxybenzyl, methoxyethoxymethyl, pentafluorophenyl, 4-p-methylbenzyloxybenzyl; Preferably, the compound is selected from: Preferably, the compound is selected from:

22. Equation -L-, -L 2 -、-L 2a -L 2b -(Q) n -ML-,M 1 The connection unit with the structure shown in -L-, -LD or -MLD, wherein L, L 2 -L 2a L 2b Q, n, M, M 1 D is as defined in any one of claims 1-15.

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