Anti-her3 antibody drug conjugate, composition thereof, and use thereof

Abstract

Provided is an anti-HER3 antibody drug conjugate, specifically comprising an antibody moiety, an intermediate linker moiety, and a cytotoxic drug moiety which are linked together. The provided antibody drug conjugate achieves excellent anti-tumor activity or / and better safety. The provided antibody drug conjugate can be used for the treatment of cancer.

Description

Anti-HER3 antibody drug conjugates and their compositions and uses Technical Field

[0001] The present invention relates to an antibody-drug conjugate comprising an antibody portion, an intermediate linker portion and a cytotoxic drug portion connected to each other. The present invention also relates to the use of the antibody-drug conjugate in preparing a drug for treating cancer. Background Art

[0002] HER3 (human epidermal growth factor receptor 3), also known as ErbB-3 (Receptor tyrosine-protein kinase erbB-3), is a member of the human epidermal growth factor receptor (HER / EGFR / ERBB) family, which includes EGFR (ErbB-1), HER2 / c-neu (ErbB-2), HER3 (ErbB-3), and HER4 (ErbB-4). HER3 possesses an extracellular ligand-binding domain (ECD), a dimerization domain within the ECD, a transmembrane domain, a protein tyrosine kinase domain (TKD), and a C-terminal phosphorylation domain, but lacks intracellular tyrosine kinase activity.

[0003] The HER3 ligand, neuregulin (NRG; also known as heregulin, HRG), binds to the extracellular domain of HER3 and activates receptor-mediated signaling pathways by promoting dimerization with other HER family members and transphosphorylation of their intracellular domains. Dimerization of HER3 with other HER family members expands the signaling potential of HER3 and serves not only as a means of signal diversification but also as a means of signal amplification. For example, HER2 / HER3 heterodimers induce one of the most important mitogenic signals among HER family members. HER3 is ubiquitously expressed in various cancers, including breast, ovarian, colon, gastric, lung, skin, and pancreatic cancers.

[0004] Antibody-drug conjugates (ADCs) are a class of drugs that combine the high specificity of therapeutic antibodies with the high cytotoxic activity of cytotoxic drugs. The therapeutic antibody and cytotoxic drug are connected via a linker. Currently, at least ten ADCs are marketed globally. The antibody components of brentuximab vedotin, polatuzumab vedotin, and enfortumab vedotin target CD30, CD79b, and Nectin-4, respectively; trastuzumab emtansine and trastuzumab deruxtecan target HER2; gemtuzumab ozogamicin and inotuzumab ozogamicin target CD33 and CD22, respectively; and sacituzumab govitecan targets TROP2. The newly approved belantamab mafodotin and loncastuximab tesirine target BCMA and CD19, respectively. Cytotoxic agents: brentuximab vedotin, polatuzumab vedotin, enfortumab vedotin, and belantamab mafodotin all utilize microtubule-targeting auristatins; trastuzumab emtansine utilizes a microtubule-targeting maytansinoid toxin; gemtuzumab ozogamicin and inotuzumab ozogamicin utilize DNA-targeting calicheamicins; trastuzumab deruxtecan and sacituzumab govitecan utilize camptothecins; and loncastuximab tesirine utilizes a DNA-targeting PBD dimer. Regarding the intermediate linker: trastuzumab emtansine and belantamab mafodotin utilize a non-cleavable linker, while the remaining eight molecules utilize a cleavable linker.

[0005] Eribulin (Formula I) is a synthetic analog of the natural marine product halichondrin B. It inhibits the microtubule growth phase through a tubulin-based antimitotic mechanism, leading to G2 / M cell cycle arrest, disruption of the mitotic spindle, and ultimately, apoptosis after prolonged mitotic arrest. Eribulin is currently approved for the treatment of metastatic breast cancer and soft tissue sarcoma.

[0006]

[0007] ADC drugs combine the dual advantages of the high efficacy of cytotoxic small molecules and the high selectivity of antibodies for specific tumor cells. There is still a need to develop high-efficiency and low-toxic ADC drugs that can target more indications.

[0008] Summary of the Invention

[0009] Antibody-drug conjugates (ADCs)

[0010] The present invention provides an antibody-drug conjugate, wherein an antibody or an antigen-binding fragment thereof is coupled to the cytotoxic drug eribulin or a derivative thereof; preferably, the antibody or the antigen-binding fragment thereof specifically binds to HER3.

[0011] In one aspect, the present invention provides a compound having the general formula Ab-(LU) n The present invention relates to an antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, wherein Ab represents an antibody portion, L represents a linker portion, U represents a cytotoxic drug portion, and n is an integer or decimal selected from 1 to 10. In certain embodiments, the antibody portion Ab and the linker portion are connected through a specific functional group, and the antibody portion can specifically bind to an antigen.

[0012] In one aspect, the present invention provides a compound of the formula Ab-(LU) n The present invention provides an antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, wherein the cytotoxic drug portion U is coupled to the antibody portion Ab via a linker portion L. In some specific embodiments, the present invention provides an antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, wherein each cytotoxic drug portion U is coupled to the antibody portion Ab via a linker portion L. The linker portion L of the present invention can be linked to the antibody portion by any method known in the art, preferably the linker portion is linked to the antibody portion via a sulfhydryl group and / or an amino group. In some more preferred embodiments, the linker portion of the present invention is linked to the antibody portion via a sulfhydryl group.

[0013] In one aspect, the present invention provides a compound of the formula Ab-(LU) nThe invention provides an antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, wherein the cytotoxic drug portion U is coupled to the antibody portion Ab via a linker portion L, which may be a cleavable linker or a non-cleavable linker. In some embodiments, the linker portion of the present invention is a cleavable linker, for example, it may be a low pH-dependent degradation type (including hydrazone bonds, carbonate bonds, etc.), a proteolytic type (including peptidyl bonds), or a high glutathione concentration-dependent degradation type (including disulfide bonds). The cleavable linker can be broken in the target cell, thereby releasing the cytotoxic drug. In other embodiments, the linker portion of the present invention is a non-cleavable linker, for example, it may be a maleimidocaproyl group, etc.

[0014] In one aspect, the present invention provides a compound of the formula Ab-(LU) n The antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, wherein the antibody portion Ab is coupled to one or more cytotoxic drug portions U, and the cytotoxic drug can be selected from alkaloids, antimetabolites, antitumor antibiotics, alkylating agents and platinums, etc., and preferably the cytotoxic drug is a microtubule inhibitor cytotoxic drug (including maytansines, auristatins, eribulins, etc.) or a DNA-acting cytotoxic drug (including calicheamicins, duocarmycins, PBDs (pyrrolobenzodiazepines), topoisomerase I inhibitors, etc.).

[0015] In some specific embodiments, the present invention provides the general formula Ab-(LU) n The cytotoxic drug portion U of the antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof is a microtubule inhibitor.

[0016] In some specific embodiments, the present invention provides the general formula Ab-(LU) n The cytotoxic drug portion U of the antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof is eribulin or a derivative thereof.

[0017] In certain embodiments, the cytotoxic drug portion is connected to the linker portion via a functional group, and the cytotoxic drug molecules are released in tumor cells, thereby exerting an anti-tumor effect.

[0018] In one aspect, the present invention provides a compound having the general formula Ab-(LU) n An antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, wherein Ab represents an antibody portion, L represents a linker portion, U represents a cytotoxic drug portion, and n is an integer or decimal selected from 1 to 10, wherein the antibody-drug conjugate comprises the structure shown in the following Formula IIa:

[0019]

[0020] in,

[0021] R a selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0022] R b selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0023] or,

[0024] R a With R b and the atoms to which they are attached form an optionally substituted 5- to 8-membered heterocyclyl. a With R b Each is independently selected from a hydrogen atom, a methyl group, an ethyl group, a propyl group or an isopropyl group. a and R b A hydrogen atom.

[0025] In some embodiments, the antibody drug conjugate comprises the structure depicted in Formula IIa-1 below:

[0026]

[0027] In some embodiments, the present invention provides a compound having the general formula Ab-(LU) n An antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, wherein Ab represents an antibody portion, L represents a linker portion, U represents a cytotoxic drug portion, n is an integer or decimal selected from 1 to 10, wherein -U is a structure represented by Formula IIa,

[0028] in,

[0029] R a selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0030] R b selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0031] or,

[0032] R a With R b and the atoms to which they are attached form an optionally substituted 5- to 8-membered heterocyclyl. a With R b Each is independently selected from a hydrogen atom, a methyl group, an ethyl group, a propyl group or an isopropyl group. a and R b In some embodiments, the -U is a structure shown in Formula IIa-1.

[0033] In some embodiments, the present invention provides a general formula of Ab-(LU) n The antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof comprises the structure shown in the following formula IIIa:

[0034]

[0035] in,

[0036] R a selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0037] R b selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0038] or,

[0039] R a With R b and the atoms to which they are attached form an optionally substituted 5- to 8-membered heterocyclyl. a With R b Each is independently selected from a hydrogen atom, a methyl group, an ethyl group, a propyl group or an isopropyl group. a and R b A hydrogen atom.

[0040] In some embodiments, the antibody drug conjugate comprises the structure depicted by the following formula IIIa-1:

[0041]

[0042] In some embodiments, the present invention provides a general formula of Ab-(LU) n The antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof has a structure shown in the following formula IV:

[0043]

[0044] Wherein, Ab represents the antibody portion;

[0045] n is an integer or decimal selected from 1-10;

[0046] R a selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0047] R b selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0048] or,

[0049] R a With R b and the atoms to which they are attached form an optionally substituted 5- to 8-membered heterocyclyl. a With R b Each independently selected from a hydrogen atom or C1-5 Alkyl (preferably C 1-4 Alkyl groups, such as C 1-3 In some embodiments, the R a With R b Each is independently selected from a hydrogen atom, a methyl group, an ethyl group, a propyl group or an isopropyl group. a and R b A hydrogen atom.

[0050] In a specific embodiment, the present invention provides an antibody drug conjugate represented by the following formula IV-1 or a pharmaceutically acceptable salt or solvate thereof:

[0051]

[0052] in,

[0053] Ab is the antibody part,

[0054] n is an integer or decimal selected from 1 to 10.

[0055] In some embodiments, in the above-mentioned antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof, n is 2-4.8, 2.6-4.8, 3.5-4.8, 4-4.8, 2-4.5, 2.6-4.5, 3.5-4.5, 4-4.5, 3.5-4.2, 3.5-4, 4-4.2, 7-8, 7-7.9, 7-7.6, 7-7.5, 7.1-8, 7.1-7.9, 7.1-7.6, 7.5-8, 7.6-8, or 7.6-7.9. In some embodiments, n is about 2.6, about 4, about 4.2, about 4.8, about 7, about 7.1, about 7.5, about 7.6, about 7.9 or about 8.

[0056] The number of cytotoxic drugs conjugated to the antibody portion of the antibody-drug conjugate (ADC) of the present invention can vary, such that the antibody-drug conjugates or pharmaceutically acceptable salts or solvates thereof provided herein can be heterogeneous, i.e., the antibody-drug conjugates or pharmaceutically acceptable salts or solvates thereof provided herein include antibodies or antigen-binding fragments thereof conjugated to different numbers of cytotoxic drugs, for example, one molecule of antibody or antigen-binding fragment thereof is conjugated to 0 (i.e., no cytotoxic drug), 1, 2, 3, 4, 5, 6, 7, 8, or other more molecules of cytotoxic drug.

[0057] By controlling the ratio of the antibodies or antigen-binding fragments thereof coupled with different numbers of cytotoxic drugs, antibody-drug conjugates or pharmaceutically acceptable salts or solvates thereof having different drug-antibody ratios (DAR) can be produced. In this article, "DAR" and "n" are used interchangeably. It should be understood that the DAR or n is the average molar ratio of the cytotoxic drug to the antibody or antigen-binding fragment thereof in the ADC, that is, the average number of cytotoxic drugs coupled per molecule of the antibody or antigen-binding fragment thereof. For example, "DAR is about 4" or "n is about 4" refers to an antibody-drug conjugate or pharmaceutically acceptable salt or solvate thereof: comprising a heterogeneous mixture in which each molecule of the antibody or antigen-binding fragment thereof is coupled with different numbers of cytotoxic drugs (e.g., each antibody or antigen-binding fragment thereof is coupled with 0, 1, 2, 3, 4, 5, 6, 7 or 8 cytotoxic drugs), but the average molar ratio of the cytotoxic drug to the antibody or antigen-binding fragment thereof is about 4. Similarly, "DAR is about 8" or "n is about 8" means that the average molar ratio of cytotoxic drug to antibody or antigen-binding fragment thereof in the ADC is about 8.

[0058] In one aspect, the present invention provides a compound having the general formula Ab-(LU) n The present invention provides an antibody drug conjugate of formula IV-1 or a pharmaceutically acceptable salt or solvate thereof, wherein Ab (antibody portion) can specifically bind to a tumor antigen, and the tumor antigen can be selected from any target for tumor treatment known in the art, and non-limiting examples of the target include HER2, HER3, EGFR, CD20, CD30, CD33, CD47, CD79b, VEGF, VEGFR, MET, RET, PD-1 or PD-L1. In some embodiments, the present invention provides an antibody drug conjugate of formula IV-1 or a pharmaceutically acceptable salt or solvate thereof, wherein Ab (antibody portion) can specifically bind to a tumor antigen, and the tumor antigen can be selected from any target for tumor treatment known in the art, and non-limiting examples of the target include HER2, HER3, EGFR, CD20, CD30, CD33, CD47, CD79b, VEGF, VEGFR, MET, RET, PD-1 or PD-L1.

[0059] In some embodiments, in the antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, Ab is an anti-HER3 antibody or an antigen-binding fragment thereof.

[0060] In some embodiments, the Ab is an anti-HER3 antibody or an antigen-binding fragment thereof, comprising a heavy chain CDR (HCDR) 1 comprising the amino acid sequence of SEQ ID NO: 1, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 2, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 3, a light chain CDR (LCDR) 1 comprising the amino acid sequence of SEQ ID NO: 4, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 5, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0061] The CDR amino acid sequences of the anti-HER3 antibodies or antigen-binding fragments thereof are provided in Table S1 below.

[0062] In some embodiments, the present invention provides a general formula of Ab-(LU) n The antibody portion of the antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof is Patritumab, which has the sequence shown in Table S1 below.

[0063] Table S1 CDR and variable region amino acid sequences of patrastuzumab

[0064]

[0065] In some embodiments, the anti-HER3 antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO:7; and a light chain variable region comprising an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO:8. In some embodiments, the anti-HER3 antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 7, and the light chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 8. In some embodiments, the amino acid sequence of the heavy chain variable region of the anti-HER3 antibody or antigen-binding fragment thereof is set forth in SEQ ID NO: 7, and the amino acid sequence of the light chain variable region is set forth in SEQ ID NO: 8.

[0066] In some embodiments, the anti-HER3 antibody or its antigen-binding fragment may further comprise a constant region of an immunoglobulin, or a fragment, analog, variant or derivative of the constant region. In some embodiments, the constant region is from a human immunoglobulin heavy chain, such as a heavy chain of IgG1, IgG2, IgG3 and IgG4 or other classes of immunoglobulins, preferably a heavy chain of IgG1. In some embodiments, the constant region may comprise modifications as described in any text, such as insertion, deletion, substitution or chemical modification of amino acids. In some embodiments, the constant region comprises a mutation that changes effector function. In some embodiments, any amino acid residue in the constant region may be substituted with an amino acid residue of any allotype.

[0067] In some embodiments, the anti-HER3 antibody or antigen-binding fragment thereof comprises a heavy chain comprising an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 9, and a light chain comprising an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 10. In some embodiments, the anti-HER3 antibody or antigen-binding fragment thereof comprises a heavy chain and a light chain, the heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 9, and the light chain comprising the amino acid sequence set forth in SEQ ID NO: 10. In some embodiments, the amino acid sequence of the heavy chain of the anti-HER3 antibody or antigen-binding fragment thereof is set forth in SEQ ID NO: 9, and the amino acid sequence of the light chain is set forth in SEQ ID NO: 10.

[0068]

[0069]

[0070] In some embodiments, the anti-HER3 antibody is selected from the group consisting of: Patritumab, Seribantumab, Elgemtumab, Duligotuzumab, CDX-3379, Lumretuzumab, or GSK2849330. In some specific embodiments, the anti-HER3 antibody is Patritumab.

[0071] In some embodiments, the anti-HER3 antibody or antigen-binding fragment thereof is selected from a monoclonal antibody, a multispecific antibody, a Fab fragment, a Fab' fragment, a F(ab)'2 fragment, a Fd fragment, a Fv fragment, a dAb fragment, an isolated CDR region, a scFv, a nanobody, or a fusion protein.

[0072] In some embodiments, the antibody drug conjugate of the present invention or a pharmaceutically acceptable salt or solvate thereof has the general formula Ab-(LU) n , wherein the Ab can be modified, for example, including changes, additions or reductions in one or more amino acid sequences. In this context, the modified Ab still retains the activity of specifically binding to its corresponding antigen.

[0073] In some embodiments, the antibody drug conjugate provided by the present invention or a pharmaceutically acceptable salt or solvate thereof has the structure shown below:

[0074] In some such embodiments, n is 2-4.8, 2.6-4.8, 3.5-4.8, 4-4.8, 2-4.5, 2.6-4.5, 3.5-4.5, 4-4.5, 3.5-4.2, 3.5-4, 4-4.2, 7-8, 7-7.9, 7-7.6, 7-7.5, 7.1-8, 7.1-7.9, 7.1-7.6, 7.5-8, 7.6-8, or 7.6-7.9. In some specific embodiments, n is about 2.6, about 4, about 4.2, about 4.8, about 7, about 7.1, about 7.5, about 7.6, about 7.9, or about 8.

[0075] In one aspect, the present invention provides a compound having the general formula Ab-(LU) n The antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof exhibits one or more of the following properties:

[0076] (a) Binding to HER3;

[0077] (b) blocking the binding of HER3 to its ligand;

[0078] (c) Demonstrated endocytosis in cells expressing HER3;

[0079] (d) has cytotoxic activity against HER3-expressing tumor cells;

[0080] (e) There is a bystander effect.

[0081] In some embodiments, the antibody drug conjugate, or a pharmaceutically acceptable salt or solvate thereof, binds to human HER3.

[0082] In some embodiments, the antibody drug conjugates provided by the present invention or pharmaceutically acceptable salts or solvates thereof show strong endocytosis in cells with different HER3 expression levels, and can continuously accumulate the amount of internalized ADC as the endocytosis time increases.

[0083] Pharmaceutical composition

[0084] In one aspect, the present invention provides a pharmaceutical composition comprising an antibody drug conjugate of the present invention or a pharmaceutically acceptable salt or solvate thereof. In some embodiments, the present invention provides a pharmaceutical composition comprising an antibody drug conjugate of the present invention or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers include, for example, excipients, diluents, encapsulating materials, fillers, buffers, or other agents.

[0085] use

[0086] In one aspect, the present invention provides the use of an antibody-drug conjugate of the present invention, or a pharmaceutically acceptable salt or solvate thereof, in the preparation of a medicament for treating cancer. In one aspect, the present invention provides the use of a pharmaceutical composition comprising an antibody-drug conjugate of the present invention, or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier in the preparation of a medicament for treating cancer. In one aspect, the present invention provides the use of a pharmaceutical composition of the present invention in the preparation of a medicament for treating cancer.

[0087] In one aspect, the present invention provides an antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition thereof for treating cancer.

[0088] In one aspect, the present invention provides a method for treating cancer, comprising administering to a patient in need thereof a therapeutically effective amount of an antibody drug conjugate of the present invention or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition comprising an antibody drug conjugate of the present invention or a pharmaceutically acceptable salt or solvate thereof and a pharmaceutically acceptable carrier. In one aspect, the present invention provides a method for treating cancer, comprising administering to a patient in need thereof a therapeutically effective amount of a pharmaceutical composition of the present invention. In some embodiments, the method comprises contacting tumor cells with the antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof, or the pharmaceutical composition, thereby killing tumor cells or inhibiting tumor cell growth.

[0089] In some embodiments, administering a therapeutically effective amount of an antibody-drug conjugate of the present invention, or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition of the present invention to a patient can kill tumor cells or inhibit tumor cell growth.

[0090] In one aspect, the present invention provides the use of the antibody-drug conjugate of the present invention, or a pharmaceutically acceptable salt or solvate thereof, for treating cancer. In one aspect, the present invention provides the use of a pharmaceutical composition comprising the antibody-drug conjugate of the present invention, or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier for treating cancer. In one aspect, the present invention also provides the use of the pharmaceutical composition of the present invention for treating cancer.

[0091] In some embodiments, in the above methods or uses, the patient is not suitable for treatment with a drug targeting HER2. In some embodiments, in the above methods or uses, the patient is resistant to a drug targeting HER2.

[0092] In some embodiments, in the above methods or uses, the cancer is a HER3-positive cancer. In some embodiments, the antibody drug conjugate of the present invention or a pharmaceutically acceptable salt or solvate thereof can be used to treat cancers that express the HER3 protein. In some embodiments, administering a therapeutically effective amount of the antibody drug conjugate of the present invention or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition of the present invention to a patient can kill HER3-expressing tumor cells or inhibit the growth of HER3-expressing tumor cells. Examples of cancer include, but are not limited to, biliary tract cancer, carcinosarcoma, esophageal cancer, gastroesophageal junction cancer, breast cancer, gastric cancer, pancreatic cancer, head and neck cancer, colorectal cancer, renal cancer, cervical cancer, ovarian cancer, endometrial cancer, uterine cancer, melanoma, pharyngeal cancer, oral cancer, skin cancer, lung cancer, glioblastoma multiforme, glioblastoma, urothelial carcinoma, prostate cancer, bladder cancer, gastrointestinal stromal tumor, squamous cell carcinoma, peritoneal cancer, liver cancer, salivary gland cancer, vulvar cancer, thyroid cancer, testicular cancer, anal cancer, or penile cancer.

[0093] Linker-drug intermediate compounds

[0094] In some aspects, the present invention provides a linker-drug intermediate compound having a structure shown in the following formula III:

[0095]

[0096] in,

[0097] R a selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0098] R bselected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0099] or R a With R b Together with the atoms to which they are attached, they form an optionally substituted 5- to 8-membered heterocyclyl.

[0100] In some embodiments, the linker-drug intermediate compound of the structure shown in Formula III, wherein R a With R b Each is independently selected from a hydrogen atom, a methyl group, an ethyl group, a propyl group or an isopropyl group.

[0101] In a specific embodiment, the present invention provides a linker-drug intermediate compound having a structure shown in the following formula III-1:

[0102]

[0103] The linker structure employed in the present invention connects the anti-tumor compound eribulin or its derivative to an antibody or its antigen-binding fragment, resulting in an antibody-drug conjugate that achieves excellent anti-tumor efficacy and / or safety. In some embodiments, the antibody-drug conjugate exhibits superior anti-tumor efficacy and / or safety compared to eribulin. In some embodiments, the antibody-drug conjugate exhibits superior anti-tumor efficacy and / or safety compared to Daiichi Sankyo Co., Ltd.'s anti-HER3 antibody-drug conjugate U3-1402 (Patritumab deruxtecan). In some embodiments, the provided anti-HER3 antibody-drug conjugate exhibits good tumor cell cytotoxicity, particularly against a variety of tumor cells and / or cells with varying HER3 expression levels (high, intermediate, and / or low). In some embodiments, the provided anti-HER3 antibody-drug conjugate exhibits good cytotoxicity against trastuzumab-resistant tumor cells. In some embodiments, the provided anti-HER3 antibody-drug conjugate has excellent safety. In some embodiments, the provided antibody-drug conjugate, such as the anti-HER3 antibody-drug conjugate, is less susceptible to aggregation. In some experiments, it was found that using the linker structure of the present invention to connect the anti-tumor compound eribulin or its derivatives to an antibody or its antigen-binding fragment can improve the anti-aggregation property of the antibody-drug conjugate. BRIEF DESCRIPTION OF THE DRAWINGS

[0104] Figures 1A-1G show the binding activities of pertrastuzumab-eribulin conjugates, pertrastuzumab-DDDXd-D8, and pertrastuzumab with different DAR values ​​to cells with different HER3 expression levels;

[0105] Figures 2A-2D show the endocytosis of pertuzumab-eribulin conjugates with different DAR values, pertuzumab-DDDXd-D8, and pertuzumab in cells with different HER3 expression levels;

[0106] Figures 3A-3I show the cell killing rates of pertrastuzumab-eribulin conjugates with different DAR values ​​and pertrastuzumab-DDDXd-D8 against cells with different HER3 expression levels;

[0107] FIG4 shows the effects of different DAR values ​​of pertrastuzumab-eribulin conjugates, pertrastuzumab-DDDXd-D8, and vehicle control on the changes in tumor volume in a nude mouse subcutaneous transplanted tumor model of JIMT-1 human breast cancer cells;

[0108] FIG5 shows the effects of different DAR values ​​of pertrastuzumab-eribulin conjugates, pertrastuzumab-DDDXd-D8, and vehicle control on tumor weight in nude mice subcutaneously transplanted with JIMT-1 human breast cancer cells;

[0109] Figure 6 shows the effects of different DAR values ​​of pertuzumab-eribulin conjugates, pertuzumab-DDDXd-D8 and vehicle control on the changes in mouse body weight in the JIMT-1 human breast cancer cell subcutaneous transplanted tumor model in nude mice.

[0110] Explanation and Definition

[0111] Unless otherwise indicated, the following terms used in this application have the following meanings. A particular term should not be construed as undefined or unclear unless specifically defined, but rather should be understood according to its ordinary meaning in the art. When a trade name appears in this document, it is intended to refer to the corresponding commercial product or its active ingredient.

[0112] The term "substituted" refers to the replacement of any one or more hydrogen atoms on a particular atom by a substituent, as long as the valence state of the particular atom is normal and the substituted compound is stable. When the substituent is an oxo (i.e., =O), it means that two hydrogen atoms are replaced. Oxo does not occur on aromatic groups.

[0113] The term "optionally" or "optionally" means that the event or situation described subsequently may or may not occur, and the description includes both the occurrence of the event or situation and the non-occurrence of the event or situation. The term "optionally substituted" means substituted or unsubstituted, for example, an ethyl group is "optionally" substituted with a halogen, meaning that the ethyl group may be unsubstituted (CH2CH3), monosubstituted (such as CH2CH2F), polysubstituted (such as CHFCH2F, CH2CHF2, etc.), or fully substituted (CF2CF3). It will be understood by those skilled in the art that for any group comprising one or more substituents, no substitution or substitution pattern that would be sterically impossible and / or incomposable to synthesize will be introduced.

[0114] In this article, C m-n , means that the part has an integer or decimal number of carbon atoms in a given range. For example, "C 1-6 ” means that the group can have 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms.

[0115] When any variable (e.g., R) occurs more than once in a compound's composition or structure, its definition on each occurrence is independent. Thus, for example, if a group is substituted with two R's, each R has an independent alternative.

[0116] When the number of a linking group is 0, such as -(CH2)0-, it means that the linking group is a covalent bond.

[0117] When one of the variables is selected from a covalent bond, it means that the two groups it connects are directly connected. For example, when L in ALZ represents a covalent bond, it means that the structure is actually AZ.

[0118] The term "halo" or "halogen" refers to fluorine, chlorine, bromine and iodine.

[0119] The term "hydroxy" refers to an -OH group.

[0120] The term "cyano" refers to a -CN group.

[0121] The term "mercapto" refers to a -SH group.

[0122] The term "amino" refers to a -NH2 group.

[0123] The term "nitro" refers to a -NO2 group.

[0124] The term "alkyl" refers to a group of the formula C n H 2n+1The alkyl group may be straight-chain or branched. For example, the term "C1-6 alkyl" refers to an alkyl group containing 1 to 6 carbon atoms (e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, neopentyl, hexyl, 2-methylpentyl, etc.). Similarly, the alkyl portion (i.e., alkyl) of alkoxy, alkylamino, dialkylamino, alkylsulfonyl, and alkylthio has the same definition as above.

[0125] The term "alkoxy" refers to an -O-alkyl group.

[0126] The term "alkylamino" refers to an -NH-alkyl group.

[0127] The term "dialkylamino" refers to -N(alkyl)2.

[0128] The term "alkylsulfonyl" refers to an -SO2-alkyl group.

[0129] The term "alkylthio" refers to an -S-alkyl group.

[0130] The term "alkenyl" refers to a linear or branched unsaturated aliphatic hydrocarbon group consisting of carbon atoms and hydrogen atoms and having at least one double bond. Non-limiting examples of alkenyl include, but are not limited to, ethenyl, 1-propenyl, 2-propenyl, 1-butenyl, isobutenyl, 1,3-butadienyl, and the like.

[0131] The term "alkynyl" refers to a straight or branched unsaturated aliphatic hydrocarbon group consisting of carbon atoms and hydrogen atoms and having at least one triple bond. Non-limiting examples of alkynyl groups include, but are not limited to, ethynyl (-C≡CH), 1-propynyl (-C≡C-CH3), 2-propynyl (-CH2-C≡CH), 1,3-butadiynyl (-C≡C≡CH), and the like.

[0132] The term "cycloalkyl" refers to a fully saturated carbocyclic ring that can exist as a monocyclic, bridged, or spirocyclic ring. Unless otherwise indicated, the carbocyclic ring is typically a 3- to 10-membered ring. Non-limiting examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, norbornyl (bicyclo[2.2.1]heptyl), bicyclo[2.2.2]octyl, adamantyl, and the like.

[0133] The term "cycloalkenyl" refers to a non-aromatic carbocyclic ring that is not fully saturated and can exist as a monocyclic, bridged, or spirocyclic ring. Unless otherwise indicated, the carbocyclic ring is typically a 5- to 8-membered ring. Non-limiting examples of cycloalkenyl include, but are not limited to, cyclopentenyl, cyclopentadienyl, cyclohexenyl, cyclohexadienyl, cycloheptenyl, cycloheptadienyl, etc.

[0134] The term "heterocyclic radical" refers to a non-aromatic ring that is fully saturated or partially undersaturated (but not fully unsaturated heteroaromatic) and can exist as a monocycle, a bridged ring or a spirocycle. Unless otherwise indicated, the heterocycle is typically a 3 to 7 ring containing 1 to 3 heteroatoms (preferably 1 or 2 heteroatoms) independently selected from sulphur, oxygen and / or nitrogen. The limiting examples of heterocyclic radicals include but are not limited to oxiranyl, tetrahydrofuranyl, dihydrofuranyl, pyrrolidinyl, N-methylpyrrolidinyl, dihydropyrrolyl, piperidinyl, piperazinyl, pyrazolidinyl, 4H-pyranyl, morpholinyl, thiomorpholinyl, tetrahydrothienyl etc.

[0135] The term "heterocycloalkyl" refers to a cyclic group that is fully saturated and can exist as a monocyclic, bridged or spirocyclic ring. Unless otherwise indicated, the heterocycle is typically a 3 to 7 membered ring containing 1 to 3 heteroatoms (preferably 1 or 2 heteroatoms) independently selected from sulfur, oxygen and / or nitrogen. Examples of 3-membered heterocycloalkyl groups include, but are not limited to, oxirane, thioethane, and aziridine groups; non-limiting examples of 4-membered heterocycloalkyl groups include, but are not limited to, azetidinyl, oxetanyl, and thietanyl groups; examples of 5-membered heterocycloalkyl groups include, but are not limited to, tetrahydrofuranyl, tetrahydrothiophenyl, pyrrolidinyl, isoxazolidinyl, oxazolidinyl, isothiazolidinyl, thiazolidinyl, imidazolidinyl, and tetrahydropyrazolyl groups; examples of 6-membered heterocycloalkyl groups include, but are not limited to, piperidinyl, tetrahydropyranyl, tetrahydrothiopyranyl, morpholinyl, piperazinyl, 1,4-thioxanyl, 1,4-dioxanyl, thiomorpholinyl, 1,3-dithianyl, and 1,4-dithianyl groups; examples of 7-membered heterocycloalkyl groups include, but are not limited to, azepanyl, oxetanyl, and thiepanyl groups. Preferably, the heterocycloalkyl group is a monocyclic group having 5 or 6 ring atoms.

[0136] The term "aryl" refers to an all-carbon monocyclic or fused polycyclic aromatic ring group having a conjugated π electron system. For example, an aryl group can have 6-20 carbon atoms, 6-14 carbon atoms, or 6-12 carbon atoms. Non-limiting examples of aryl groups include, but are not limited to, phenyl, naphthyl, anthracenyl, and 1,2,3,4-tetrahydronaphthalene.

[0137] The term "heteroaryl" refers to a monocyclic or fused polycyclic ring system containing at least one ring atom selected from N, O or S, with the remaining ring atoms being C and having at least one aromatic ring. Preferred heteroaryl groups have single 4 to 8-membered rings, especially 5 to 8-membered rings, or multiple fused rings containing 6 to 14, especially 6 to 10, ring atoms. Non-limiting examples of heteroaryl groups include, but are not limited to, pyrrolyl, furyl, thienyl, imidazolyl, oxazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, quinolyl, isoquinolyl, tetrazolyl, triazolyl, triazinyl, benzofuranyl, benzothienyl, indolyl, isoindolyl, etc.

[0138] "Derivative": A compound formed by replacing atoms or atomic groups in the parent compound molecule with other atoms or atomic groups is called a derivative of the parent compound.

[0139] The compounds and intermediates of the present application can also exist in different tautomeric forms, and all such forms are included in the scope of the present application. The term "tautomer" or "tautomeric form" refers to structural isomers of different energies that can interconvert via a low energy barrier. For example, proton tautomers (also referred to as prototropic tautomers) include interconversions via proton migration, such as keto-enol and imine-enamine isomerizations. The specific example of a proton tautomer is the imidazole moiety, in which a proton can migrate between two ring nitrogens. Valence tautomers include interconversions by reorganization of some bonding electrons.

[0140] The compounds of the present invention may be asymmetric, for example, having one or more stereoisomers. Unless otherwise indicated, all stereoisomers are included, such as enantiomers and diastereomers. The compounds of the present invention containing asymmetric carbon atoms can be isolated in optically pure forms or racemic forms. Optically pure forms can be resolved from racemic mixtures or synthesized by using chiral starting materials or chiral reagents.

[0141] Unless otherwise specified, any atom in the labeled synthetic compounds of the present invention may represent any stable isotope of that atom. Unless otherwise specified, when a position in a structure is defined as H, i.e., hydrogen (H-1), that position contains only naturally occurring isotopes. Similarly, unless otherwise specified, when a position in a structure is defined as D, i.e., deuterium (H-2), that position contains an isotope at least 3340 times greater than the naturally occurring isotope (0.015%) (i.e., at least 50.1% deuterium isotope). When one or more positions in the structure of a labeled synthetic compound are defined as D, i.e., deuterium (H-2), the content of the compound represented by that structure may be at least 52.5%, at least 60%, at least 67.5%, at least 75%, at least 82.5%, at least 90%, at least 95%, at least 97%, at least 98.5%, at least 99%, or at least 99.5%. The deuteration rate of a labeled synthetic compound herein refers to the ratio of the labeled synthetic isotope content to the naturally occurring isotope content. The deuteration rate per designated deuterium atom in a labeled synthetic compound herein may be at least 3500-fold (52.5%), at least 4000-fold (60%), at least 4500-fold (67.5%), at least 5000-fold (75%), at least 5500-fold (82.5%), at least 6000-fold (90%), at least 6333.3-fold (95%), at least 6466.7-fold (97%), at least 6566.7-fold (98.5%), at least 6600-fold (99%), or at least 6633.3-fold (99.5%). Isotopologues herein refer to compounds that differ in chemical structure only by their isotopic composition. The labeled compounds synthesized in the present invention have identical chemical structures, varying only in isotopic composition. Therefore, compounds synthesized in the present invention that contain deuterium at a specific position will also contain very low amounts of hydrogen isotopologues at that position. The amount of hydrogen isotopologues at a specific position in the labeled compounds synthesized in the present invention depends on many factors, including the deuterium isotopic purity of the deuterated reagent (D2O, D2, NaBD4, LiAlD4, etc.) and the effectiveness of the synthesis method for introducing the deuterium isotope. However, as mentioned above, the total amount of hydrogen isotopologues at a specific position will be less than 49.9%. The total amount of hydrogen isotopologues at a specific position in the labeled compounds synthesized in the present invention will be less than 47.5%, 40%, 32.5%, 25%, 17.5%, 10%, 5%, 3%, 1%, or 0.5%.

[0142] In the present invention, any atom not designated as deuterium is present at its natural isotopic abundance.

[0143] The term "bystander effect," as used herein, refers to an effect in which a cytotoxic drug coupled to an antibody or antigen-binding fragment thereof via a cleavable or non-cleavable linker has the ability to diffuse across the cell membrane after release from the antibody or antigen-binding fragment thereof, thereby causing killing of adjacent cells. The ability to diffuse across the cell membrane is related to the hydrophobicity of the cytotoxic drug or the combination of the cytotoxic drug and the linker. Such cytotoxic drugs may, for example, be eribulin or MMAE. A bystander effect may be desirable, particularly in tumors with heterogeneous target expression and in solid tumors where antibody penetration may be limited.

[0144] The term "treatment" means administering the compound or pharmaceutical composition described herein to prevent, improve or eliminate a disease or one or more symptoms associated with the disease, and includes but is not limited to:

[0145] (i) preventing a disease or disease state from occurring in a mammal, particularly where such mammal is susceptible to the disease state but has not yet been diagnosed as having the disease state;

[0146] (ii) inhibiting the disease or disease state, i.e., curbing its development;

[0147] (iii) alleviate the disease or condition, even if the disease or condition regresses;

[0148] (iv) reducing any direct or indirect pathological consequences of the disease or disease state.

[0149] The term "therapeutically effective amount" means an amount of a compound of the present invention that (i) treats or prevents a specific disease, condition, or disorder, (ii) alleviates, ameliorates, or eliminates one or more symptoms of a specific disease, condition, or disorder, or (iii) prevents or delays the onset of one or more symptoms of a specific disease, condition, or disorder described herein. The amount of a compound of the present invention or pharmaceutical composition that constitutes a "therapeutically effective amount" may vary depending on a number of factors, such as the compound or pharmaceutical composition and its ability to elicit a desired response in an individual, the disease state and its severity, the mode of administration, and the age, sex, and weight of the mammal to be treated. The effective amount can also be routinely determined by those skilled in the art based on their own knowledge and the present disclosure.

[0150] The term "pharmaceutically acceptable" refers to those compounds, materials, compositions and / or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0151] As the pharmaceutically acceptable salt, for example, metal salts, ammonium salts, salts with organic bases, salts with inorganic acids, salts with organic acids, salts with basic or acidic amino acids and the like can be mentioned.

[0152] The term "solvate" refers to an association of a compound with solvent molecules.

[0153] The term "antibody" is used in the broadest sense and specifically covers intact monoclonal antibodies, polyclonal antibodies, multispecific antibodies formed from at least two intact antibodies (e.g., bispecific antibodies), multifunctional antibodies, and antibody fragments, so long as they possess the desired biological activity.

[0154] The term "humanized antibody" refers to an antibody that comprises CDR regions derived from a non-human antibody, while the remainder of the antibody molecule is derived from one or more human antibodies.

[0155] The term "mutant" is used to refer to a peptide comprising an amino acid sequence derived from the amino acid sequence of the peptide as follows: one or two or more amino acids are substituted with an amino acid different from that of the original peptide, one or two or more wild-type amino acids are deleted, one or two or more amino acids not present in the wild type are inserted, and / or amino acids not present in the wild type are added to the amino terminus (N terminus) and / or carboxyl terminus (C terminus) of the wild type (collectively referred to as "mutations"). In the present invention, "insertion" may also be included in "addition".

[0156] The term "CDR" (complementarity determining region), also known as "hypervariable region", refers to each of the regions of an antibody variable domain that are highly variable in sequence and / or form structurally defined loops. A natural four-chain antibody typically contains six CDRs, three in the heavy chain variable region and three in the light chain variable region.

[0157] The term "variable region" refers to the domain of about 100 to 110 or more amino acids defined by the N-terminal domain of the light or heavy chain of an antibody that is primarily responsible for antigen recognition. The terms light chain variable region (VL) and heavy chain variable region (VH) refer to these light chain and heavy chain domains, respectively.

[0158] The term "Fab" refers to the constant domain (CL) of the light chain and the first constant domain (CH1) of the heavy chain, together with the variable domains VL (light chain variable region) and VH (heavy chain variable region) on the light chain and heavy chain, respectively. The variable domains contain the complementarity determining regions (CDRs) involved in antigen binding.

[0159] The term "scFv" includes the VH and VL domains of an antibody, wherein these domains are present in a single polypeptide chain. In some embodiments, the scFv further comprises a polypeptide linker between the VH and VL domains that enables the scFv to form the desired structure for antigen binding.

[0160] The term "antibody portion" refers to the antibody portion of the antibody drug conjugate. In certain specific embodiments, it is connected to the intermediate linker portion through a specific functional group, and the antibody portion can specifically bind to the antigen.

[0161] The term "linker moiety" refers to the portion of the antibody-drug conjugate that connects the antibody portion to the cytotoxic drug portion and can be cleavable or non-cleavable. The cleavable linker can be broken in the target cell, thereby releasing the cytotoxic drug.

[0162] The term "cytotoxic drug moiety" refers to the cytotoxic drug portion of an antibody-drug conjugate. In certain specific approaches, it is linked to an intermediate linker moiety via a functional group, which releases the cytotoxic drug molecules within tumor cells, thereby exerting an anti-tumor effect.

[0163] The term "trastuzumab" is a recombinant humanized monoclonal antibody that selectively targets the ECD4 of human epidermal growth factor receptor-2 (HER2) and can be used to treat HER2-positive cancers. An example of trastuzumab is sold under the trade name Marketed therapeutic monoclonal antibody products.

[0164] The term "Patritumab" or "Patritumomab" is a fully human anti-HER3 monoclonal antibody that can be used to treat cancers that express the HER3 protein.

[0165] The term "HER2" refers to the second member of the EGFR family and has tyrosine kinase activity. The HER2 expression level can be detected by immunohistochemistry. HER2-positive refers to IHC3+, and HER2-negative refers to IHC1+ / 0. For IHC2+, ISH testing should be performed for further clarification.

[0166] The term "HER3" (human epidermal growth factor receptor 3, also known as ErbB3) is a receptor protein tyrosine kinase and belongs to the epidermal growth factor receptor (EGFR) subfamily of receptor protein tyrosine kinases, which also includes HER1 (also known as EGFR), HER2 and HER4. HER3 is a transmembrane receptor and consists of an extracellular ligand binding domain (ECD), a dimerization domain within the ECD, a transmembrane domain, an intracellular protein tyrosine kinase domain (TKD), and a C-terminal phosphorylation domain. HER3 has been found to be overexpressed in several types of cancer (e.g., breast cancer, gastrointestinal cancer, and pancreatic cancer). The association between the expression of HER2 / HER3 and the progression from non-invasive to invasive stages has been shown.

[0167] The term "triple-negative breast cancer" refers to breast cancer that is negative for estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2.

[0168] The term "EC 50 ” refers to the effective concentration that elicits 50% of the maximal response of the antigen-binding construct. EC 50 Measurement can be performed by ELISA or FACS analysis or any other method known in the art.

[0169] The term "identity" is also known as consistency. The "percentage (%) identity" of an amino acid sequence refers to the percentage of amino acid residues in the sequence to be aligned that are identical to the amino acid residues in the specific amino acid sequence shown in this article, after comparing the sequence to be aligned and, if necessary, introducing gaps to achieve maximum sequence identity, and not considering any conservative substitutions as part of sequence identity. The alignment of amino acid sequences for identity can be performed in a variety of ways within the scope of the art, such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software. Those skilled in the art can determine the appropriate parameters for aligning sequences, including any algorithm needed to obtain maximum alignment over the full length of the comparison sequence.

[0170] As used herein, the terms "subject," "patient," or "subject" are used interchangeably. In some embodiments, the term "subject," "patient," or "subject" is a mammal. In some embodiments, the subject, patient, or subject is a mouse. In some embodiments, the subject, patient, or subject is a human.

[0171] As used herein, "about" means within the acceptable error range for a particular value as determined by one of ordinary skill in the art, which depends in part on how the value is measured or determined, i.e., the limitations of the measurement system. For example, "about" may mean within 1 or more than 1 standard deviation according to practice in the art. Alternatively, "about" may mean a range of up to ±5%, such as fluctuations within ±2%, ±1%, or ±0.5% of the specific numerical range given. When a specific value is given in this application or in the scope of the invention patent application, unless otherwise stated, the meaning of "about" should be considered to be within the acceptable error range of the specific value. In this document, unless otherwise stated, the values ​​of step parameters or conditions are modified by "about" by default. DETAILED DESCRIPTION

[0172] The present invention also provides the following specific embodiments, but the protection scope of the present invention is not limited thereto:

[0173] Embodiment 1. A compound having the general formula Ab-(LU) n An antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, wherein Ab represents an antibody portion, L represents a linker portion, U represents a cytotoxic drug portion, and n is an integer or decimal selected from 1 to 10, wherein the antibody-drug conjugate comprises the structure shown in the following Formula IIa:

[0174]

[0175] in,

[0176] R a selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0177] R b selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0178] or,

[0179] R a With R b Together with the atoms to which they are attached, they form an optionally substituted 5- to 8-membered heterocyclyl.

[0180] Embodiment 2. The antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to Embodiment 1, wherein the antibody-drug conjugate comprises the structure represented by the following Formula IIIa:

[0181]

[0182] Embodiment 3. The antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to Embodiment 1 or 2, wherein the antibody-drug conjugate has a structure represented by the following Formula IV:

[0183]

[0184] in,

[0185] Ab represents the antibody part,

[0186] n is an integer or decimal selected from 1-10.

[0187] Embodiment 4. The antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to any one of Embodiments 1-3, wherein R a With R b Each is independently selected from a hydrogen atom, a methyl group, an ethyl group, a propyl group or an isopropyl group.

[0188] Embodiment 5. The antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to Embodiment 1 or 2, wherein the antibody-drug conjugate comprises the structure represented by the following Formula IIIa-1:

[0189]

[0190] Embodiment 6. The antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to Embodiment 3, wherein the antibody-drug conjugate has a structure represented by the following formula IV-1:

[0191]

[0192] Embodiment 7. The antibody drug conjugate or pharmaceutically acceptable salt or solvate thereof according to any one of embodiments 1-6, wherein n is 2-4.8, 2.6-4.8, 3.5-4.8, 4-4.8, 2-4.5, 2.6-4.5, 3.5-4.5, 4-4.5, 3.5-4.2, 3.5-4, 4-4.2, 7-8, 7-7.9, 7-7.6, 7-7.5, 7.1-8, 7.1-7.9, 7.1-7.6, 7.5-8, 7.6-8, or 7.6-7.9.

[0193] Embodiment 8. The antibody drug conjugate or pharmaceutically acceptable salt or solvate thereof according to Embodiment 7, wherein n is about 2.6, about 4, about 4.2, about 4.8, about 7, about 7.1, about 7.5, about 7.6, about 7.9 or about 8.

[0194] Embodiment 9. The antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to any one of Embodiments 1-8, wherein the Ab is an anti-HER3 antibody or an antigen-binding fragment thereof.

[0195] Embodiment 10. The antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to Embodiment 9, wherein the anti-HER3 antibody or antigen-binding fragment thereof comprises a HCDR1 comprising the amino acid sequence of SEQ ID NO: 1, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 2, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 3, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 4, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 5, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0196] Embodiment 11. The antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to Embodiment 10, wherein the anti-HER3 antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO: 7, and the light chain variable region comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO: 8; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 7, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 8.

[0197] Embodiment 12. The antibody-drug conjugate or pharmaceutically acceptable salt or solvate thereof according to embodiment 10 or 11, wherein the anti-HER3 antibody or antigen-binding fragment thereof comprises a heavy chain and a light chain, the heavy chain comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO: 9, and the light chain comprises an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO: 10.

[0198] Embodiment 13. The antibody-drug conjugate according to embodiment 9, or a pharmaceutically acceptable salt or solvate thereof, wherein the anti-HER3 antibody is patrastuzumab.

[0199] Embodiment 14. The antibody drug conjugate or pharmaceutically acceptable salt or solvate thereof according to any one of embodiments 9 to 13, wherein the antibody drug conjugate or pharmaceutically acceptable salt or solvate thereof exhibits one or more of the following properties:

[0200] (a) Binding to HER3;

[0201] (b) blocking the binding of HER3 to its ligand;

[0202] (c) Demonstrated endocytosis in cells expressing HER3;

[0203] (d) has cytotoxic activity against HER3-expressing tumor cells;

[0204] (e) There is a bystander effect.

[0205] Embodiment 15. A pharmaceutical composition comprising the antibody drug conjugate according to any one of Embodiments 1-14 or a pharmaceutically acceptable salt or solvate thereof; optionally, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.

[0206] Embodiment 16. Use of the antibody drug conjugate or a pharmaceutically acceptable salt or solvate thereof according to any one of Embodiments 1 to 14, or the pharmaceutical composition according to Embodiment 15, in the preparation of a medicament for treating cancer; preferably, the cancer is HER3-positive cancer; preferably, the cancer is biliary tract cancer, carcinosarcoma, esophageal cancer, gastroesophageal junction cancer, breast cancer, gastric cancer, pancreatic cancer, head and neck cancer, colorectal cancer, kidney cancer, cervical cancer, ovarian cancer, endometrial cancer, uterine cancer, melanoma, pharyngeal cancer, oral cancer, skin cancer, lung cancer, glioblastoma multiforme, glioblastoma, urothelial carcinoma, prostate cancer, bladder cancer, gastrointestinal stromal tumor, squamous cell carcinoma, peritoneal cancer, liver cancer, salivary gland cancer, vulvar cancer, thyroid cancer, testicular cancer, anal cancer, or penile cancer.

[0207] Embodiment 17. A method for treating cancer, comprising administering to a patient in need thereof a therapeutically effective amount of the antibody-drug conjugate of any one of Embodiments 1-14 or a pharmaceutically acceptable salt or solvate thereof, or the pharmaceutical composition of Embodiment 15; preferably, the cancer is HER3-positive cancer.

[0208] Embodiment 18. The method according to embodiment 17, comprising contacting tumor cells with the antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, or the pharmaceutical composition, thereby killing tumor cells or inhibiting tumor cell growth.

[0209] Embodiment 19. The method of embodiment 17 or 18, wherein the cancer is biliary tract cancer, carcinosarcoma, esophageal cancer, gastroesophageal junction cancer, breast cancer, gastric cancer, pancreatic cancer, head and neck cancer, colorectal cancer, kidney cancer, cervical cancer, ovarian cancer, endometrial cancer, uterine cancer, melanoma, pharyngeal cancer, oral cancer, skin cancer, lung cancer, glioblastoma multiforme, glioblastoma, urothelial carcinoma, prostate cancer, bladder cancer, gastrointestinal stromal tumor, squamous cell carcinoma, peritoneal cancer, liver cancer, salivary gland cancer, vulvar cancer, thyroid cancer, testicular cancer, anal cancer, or penile cancer.

[0210] Embodiment 20. A linker-drug intermediate compound having a structure shown in Formula III:

[0211]

[0212] in,

[0213] R a selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0214] R b selected from hydrogen atoms, deuterium atoms, optionally substituted C 1-6 Alkyl, optionally substituted C 3-7 Cycloalkyl, optionally substituted C 3-7 Heterocyclyl, optionally substituted C 6-10 Aryl, optionally substituted C 5-12 heteroaryl;

[0215] or R a With R b Together with the atoms to which they are attached, they form an optionally substituted 5- to 8-membered heterocyclyl.

[0216] Embodiment 21. The linker-drug intermediate compound according to embodiment 20, wherein R a With R b Each is independently selected from a hydrogen atom, a methyl group, an ethyl group, a propyl group or an isopropyl group.

[0217] For the sake of clarity, the present invention is further illustrated by examples, but the examples are not intended to limit the scope of the present application. The reagents used in the present application are generally commercially available and can be used without further purification.

[0218] The patritumab used in the examples of this application was prepared according to conventional methods for antibodies. An expression vector (including, for example, the pcDNA3.1 vector disclosed in CN107001463A and the pCHO1.0 vector disclosed in CN109422811A) was first constructed, and then expressed in Expi-CHO host cells after transfection. Purification was performed using Protein A affinity chromatography. The amino acid sequences of the heavy and light chains of patritumab are shown in SEQ ID NOs: 9 and 10, respectively. The synthesis of deuterated MC-GGFG-DXd (MC-GGFG-DDDXd) is described in Example 14 of patent publication WO2022033578A1.

[0219] The cells and their sources involved in the examples of this application are shown in the following table:

[0220] Cell sources: NCI-N87, Chinese Academy of Sciences Cell Bank BT474, Chinese Academy of Sciences SKBR3, Sino-US Crown Biotech SK-OV3, Chinese Academy of Sciences Cell Bank JIMT-1, ATCC Capan1, Beina Biotech MCF-7, Chinese Academy of Sciences Cell Bank KYSE410, Yaji Biotech MDA-MB-468, Beina Biotech HCC1569, Nanjing Kebai SW620, Nanjing Kebai A549, Basic Medical Cell Center, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences WiDr, Nanjing Kebai

[0221] .

[0222] Example 1 Preparation of Compound III-1

[0223]

[0224] 100 mg of compound A (0.12 mmol) and 75 mg of compound B (0.145 mmol) were weighed and added to a 20 mL reaction tube (Compound A CAS No. 2413428-36-9, Compound B CAS No. 441045-17-6). 2 mL of N,N-dimethylformamide was added, and the temperature was cooled to 0°C. 70 mg of 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (0.18 mmol) and 49 mg of N,N-diisopropylethylamine (0.36 mmol) were added. The reaction was continued at 0°C for 1 hour. Purification by preparative liquid phase chromatography gave approximately 70 mg of compound III-1 (MC-GGFG-Eribulin). ESI-MS analysis revealed m / z of compound III-1 = 1241.72 [M+H]. + The hydrogen spectrum of compound III-1 is as follows:

[0225] 1H NMR (500MHz, DMSO) δ8.22(t,J=5.3Hz,1H),8.12(d,J=8.0Hz,1H),8.06(t,J=5.4Hz,1H),7.9 9(t,J=5.3Hz,1H),7.65(d,J=5.2Hz,1H),7.30-7.21(m,4H),7.18(d,J=6.4Hz,1H),6.99(s, 2H),5.02(d,J=26.0Hz,2H),4.79(d,J=38.9Hz,2H),4.65-4.60(m,2H),4.56(d,J=3.7Hz,1H ),4.50-4.42(m,1H),4.26(d,J=10.1Hz,1H),4.21-4.14(m,1H),4.10(s,3H),4.05-3.98(m, 1H),3.86-3.64(m,8H),3.60-3.45(m,4H),3.37-3.30(m,2H),3.26(s,4H),3.17-3.09(m,1H ),3.08-2.99(m,2H),2.84(d,J=10.4Hz,1H),2.86-2.66(m,3H),2.56-2.50m,1H),2.37-2.1 8(m,5H),2.15-2.05(m,3H),2.05-1.96(m,2H),1.95-1.84(m,4H),1.75-1.57(m,6H),1.55- 1.38(m,6H),1.35-1.25(m,3H),1.23-1.12(m,3H),1.03(d,J=6.0Hz,3H),1.00-0.92(m,1H).

[0226] Example 2 Preparation of Antibody Drug Conjugates

[0227] Reagents:

[0228] Solution A: pH 7.4 PBS buffer

[0229] Solution B: 10 mM TCEP (tris(2-carboxyethyl)phosphine hydrochloride) aqueous solution

[0230] Solution C: DMSO

[0231] Solution D: Histidine buffer (containing 0.89 mg / mL L-histidine and 4.04 mg / mL L-histidine monohydrate)

[0232] Solution E: 700 mg / mL sucrose solution (prepared using solution D)

[0233] Solution F: 20 mg / mL Tween 80 (prepared with Solution D)

[0234] Experimental process:

[0235] 1. Antibody replacement

[0236] a. Fully wet the 30KD ultrafiltration centrifuge tube with solution A.

[0237] b. Replace the antibody into solution A

[0238] c. Add appropriate amount of solution A to adjust the antibody concentration

[0239] 2. Antibody reduction

[0240] a. Calculate the molar amount of antibody, recorded as N1

[0241] b. Add an appropriate amount of solution B to the antibody solution so that the molar amount of TCEP in the reaction system is N2

[0242] c. Wrap with aluminum foil, place on a rotary incubator and shake at low speed (20 rpm), incubate at 37°C in the dark for 1 hour

[0243] 3. Coupling

[0244] a. Take an appropriate amount of linker-payload and dissolve it in DMSO to a final concentration of 10 mg / mL

[0245] b. Add DMSO to the antibody solution to a concentration of 5%, then add an appropriate amount of linker-payload solution to a molar mass of N3.

[0246] c. Wrap with aluminum foil, place on a rotary incubator and shake at low speed (20 rpm), incubate at 22°C in the dark for 1.5 hours

[0247] 4. Coupling termination

[0248] a. Wet the ultrafiltration centrifuge tube with solution D

[0249] b. Replace the antibody into solution D, add appropriate amount of solution E and F, and freeze at -80℃

[0250] Determination of the DAR value (average number of drug linkages per antibody molecule) of antibody-drug conjugates

[0251] DAR values ​​were determined using LC-MS. A 50 μg ADC sample was added with 1 μL of glycosidase PNGase F (Ruian Biotechnology, China) and incubated at 37°C for 20 hours. The mass spectrometer used in the experiment was a high-resolution Xevo G2-XS (Waters, USA). The sample concentration was adjusted to 5 μM, and direct injection was used to acquire mass spectrometric data in positive ion mode. The native mass spectrometric data were analyzed and processed using UNIFI 1.8.2.169 (Waters, USA).

[0252] Protein concentration determination of antibody drug conjugates

[0253] The protein concentration was determined by Lowry method. The sample was measured at OD 650 The absorbance value of the sample was fitted into the standard curve to calculate the protein concentration.

[0254] The antibody drug conjugates of Formula IV-1 (including IV-1 (Patritumab)) and other antibody drug conjugates of Formula IV series were prepared by the above method:

[0255]

[0256]

[0257] Sample 1: DAR = 4.8, protein concentration = 1.71 mg / mL and sample 2: DAR = 7.1, protein concentration = 1.65 mg / mL were prepared respectively.

[0258] Example 3 Cellular Activity of Small Molecule Cytotoxic Compounds and Antibody Drug Conjugates

[0259] The small molecule cytotoxic compound was diluted to 9 concentrations ranging from 35000 ng / mL to 0.0896 ng / mL using culture medium. Tumor cells in the logarithmic growth phase were taken and the density was adjusted to 1×10 5 Cells / mL were plated, 100 μL was added to each well, and a blank well without cells was set up as a control. The above-mentioned gradient dilution samples were added at 50 μL per well. Culture was carried out in a 37°C, 5% CO2 carbon dioxide incubator for 5 days. The culture medium was discarded, and CCK-8 (Japan Dojin Chemical, product number: CK04) working solution was added at 100 μL per well. After incubation for 4-5 hours to develop color, the plate was placed in a microplate reader (manufacturer: Thermo, model: VarioskanFlash), and 630 nm was used as the reference wavelength. The absorbance value of the well plate at a wavelength of 450 nm was read and recorded. The tumor cell proliferation inhibition rate was calculated.

[0260] The antibody drug conjugate was diluted to 9 concentrations ranging from 5000 ng / mL to 0.0128 ng / mL using culture medium. Tumor cells in the logarithmic growth phase were taken and the density was adjusted to 2×10 4 Cells were plated at 400 μL per well, with 100 μL added to each well. A blank well without cells was set up as a control. 50 μL of the above-described serially diluted sample was added to each well. The cells were incubated at 37°C in a 5% CO2 incubator. The culture medium was discarded and 100 μL of CTG detection solution (Promega, Catalog No. G7572) was added to each well. After incubation for 10 minutes to develop color, the cells were placed on a multi-function plate reader (Thermo, Model: VarioskanFlash) and the chemiluminescence value was read. The tumor cell proliferation inhibition rate was calculated.

[0261] IV-1 (Patritumab) cell activity:

[0262]

[0263] Example 4 Preparation of Antibody Drug Conjugates

[0264] Reagents:

[0265] Solution G: histidine / histidine hydrochloride buffer (L-histidine 1.43 mg / mL, L-histidine hydrochloride monohydrate 2.27 mg / mL);

[0266] Solution H: 10 mM TCEP (tris(2-carboxyethyl)phosphine hydrochloride) aqueous solution;

[0267] Solution I: DMSO (dimethyl sulfoxide);

[0268] Solution J: 500 mg / mL sucrose solution (prepared using solution G);

[0269] Solution K: 30 mg / mL Tween 80 (prepared with Solution G);

[0270] Solution L: 10% DMSO in solution G;

[0271] Solution M: 0.3M Na2HPO4;

[0272] Antibody: Patratuzumab;

[0273] Linker-payload (linker-drug intermediate compound): MC-GGFG-eribulin (used in the preparation of pertuzumab-eribulin conjugate) of the compound of formula III-1 in Example 1; MC-GGFG-DDDXd (used in the preparation of pertuzumab-DDDXd conjugate).

[0274] (1) Pertratuzumab-eribulin conjugates with a DAR of 2.6 or 7.6 were prepared according to the following experimental procedure and named Pertratuzumab-eribulin-D2 and Pertratuzumab-eribulin-D8, respectively. Pertratuzumab-DDDXd conjugate with a DAR of 7.9 was prepared according to the following experimental procedure and named Pertratuzumab-DDDXd-D8:

[0275] Experimental process:

[0276] 1. Antibody replacement:

[0277] Fully wet a 30KD ultrafiltration centrifuge tube with Solution G, replace the antibody into Solution G, and add an appropriate amount of Solution G to adjust the antibody concentration to 10 mg / mL; then add an appropriate amount of Solution M to adjust the pH of the antibody solution to approximately 7.0.

[0278] 2. Antibody reduction:

[0279] Calculate the molar weight of the antibody, denoted as N1; add an appropriate amount of solution H to the antibody solution so that the molar weight of TCEP in the reaction system is N2; and shake the resulting mixture in the dark at 37°C for 1 hour to reduce the disulfide bonds of the antibody, thereby obtaining reaction solution 1.

[0280] 3. Conjugation of antibodies and linker-drug intermediate compounds:

[0281] An appropriate amount of linker-payload was dissolved in 50% acetone to a final concentration of 10 mg / mL. Solution I was added to reaction solution 1 (solution I:reaction solution 1 (v / v) = 1:10), mixed, and an appropriate amount of the linker-payload solution dissolved in acetone was added to obtain a molar weight of N3. The reaction mixture was shaken in the dark at 22°C for 1 h to obtain reaction solution 2.

[0282] 4. Termination of coupling reaction:

[0283] Wet the ultrafiltration centrifuge tube with solution L; successively ultrafilter the reaction solution 2 with 20 times the volume of solution L and 20 times the volume of solution G, add appropriate amounts of solutions J and K, and freeze at -80°C.

[0284] The experimental conditions and groups are shown in Table 1-1 below.

[0285] Table 1-1 Experimental conditions and groups

[0286] ADC antibody linker-payload N1:N2 N1:N3 Partrastuzumab-eribulin-D2 Partrastuzumab MC-GGFG-eribulin 1:2 1:3 Partrastuzumab-eribulin-D8 Partrastuzumab MC-GGFG-eribulin 1:8.5 1:10.5 Partrastuzumab-DDDXd-D8 Partrastuzumab MC-GGFG-DDDXd 1:8.5 1:10.5

[0287] (2) A perrituzumab-eribulin conjugate with a DAR of 4.0-4.2 was prepared according to the following experimental procedure and named perrituzumab-eribulin-D4:

[0288] Experimental process:

[0289] 1. Antibody replacement:

[0290] Fully wet a 30KD ultrafiltration centrifuge tube with Solution G, replace the antibody into Solution G, and add an appropriate amount of Solution G to adjust the antibody concentration to 10 mg / mL; then add an appropriate amount of Solution M to adjust the pH of the antibody solution to approximately 7.0.

[0291] 2. Antibody reduction:

[0292] Calculate the molar weight of the antibody, denoted as N1; add an appropriate amount of solution H to the antibody solution so that the molar weight of TCEP in the reaction system is N2; and react the resulting mixture at 5-10°C in the dark for 6 hours, thereby reducing the disulfide bonds of the antibody and obtaining reaction solution 3.

[0293] 3. Conjugation of antibodies and linker-drug intermediate compounds:

[0294] An appropriate amount of linker-payload was dissolved in 50% acetone aqueous solution to a final concentration of 10 mg / mL. An appropriate amount of the linker-payload solution dissolved in acetone aqueous solution was added to reaction solution 3 so that the molar weight of the linker-payload was N3. The reaction mixture was reacted in the dark at 5-10°C for 40 min to obtain reaction solution 4.

[0295] 4. Termination of coupling reaction:

[0296] Wet the ultrafiltration centrifuge tube with solution L; successively ultrafilter the reaction solution 4 with 20 times the volume of solution L and 20 times the volume of solution G, add appropriate amounts of solutions J and K, and freeze at -80°C.

[0297] The experimental conditions and groups are shown in Table 1-2 below.

[0298] Table 1-2 Experimental conditions and groups

[0299] ADC antibody linker-payload N1:N2 N1:N3 Patrastuzumab-Eribulin-D4 Patrastuzumab MC-GGFG-Eribulin 1:2.58 1:5.1

[0300] Example 5 Determination of DAR Value of Antibody Drug Conjugates

[0301] The components of the patrastuzumab-eribulin conjugate prepared in Example 4 above were separated using a butyl-bonded non-porous polystyrene / divinylbenzene (PS / DVB) filler. A neutral high-salt mobile phase was used to increase the hydrophobic properties of the protein molecules, thereby combining with the hydrophobic bonds in the chromatographic column. The substances were then eluted by gradually decreasing the salt concentration and gradually increasing the proportion of isopropanol. The less hydrophobic components were eluted first, and the more hydrophobic components were eluted later.

[0302] The chromatographic column specifications were Sepax HIC-Butyl, 4.6×100 mm, 5 μm, and the column temperature was 25°C. Mobile phase A consisted of 10 mM phosphate buffer-1.5 M ammonium sulfate, pH 7.0 (weigh 1.42 g of anhydrous disodium hydrogen phosphate and 198.21 g of ammonium sulfate, add approximately 800 mL of ultrapure water, stir until fully dissolved, adjust the pH to 7.0 ± 0.1 with phosphoric acid, make up to 1 L, mix thoroughly, and filter through a 0.22 μm filter). Mobile phase B consisted of 10 mM phosphate buffer, pH 7.0 (weigh 1.42 g of anhydrous disodium hydrogen phosphate, add approximately 800 mL of ultrapure water, stir until fully dissolved, adjust the pH to 7.0 ± 0.1 with phosphoric acid, make up to 1 L, mix thoroughly, and filter through a 0.22 μm filter). Mobile phase C consisted of 100% isopropanol. The flow rate was 0.5 mL / min, and the gradient elution was performed over 30 min. The mobile phase parameters were: 75% mobile phase A plus 25% mobile phase B to 75% mobile phase B plus 25% mobile phase C from 0-15 min, 75% mobile phase B plus 25% mobile phase C from 15-20 min, and 75% mobile phase A plus 25% mobile phase B from 20-30 min. The pertrastuzumab-eribulin conjugate was diluted 1-fold with the initial mobile phase at min 0 as the test solution. The injection volume was adjusted according to the pertrastuzumab-eribulin conjugate concentration and 50 μg of protein was injected. The absorbance was measured at 280 nm.

[0303] Data processing and quantitative analysis of the results were performed using the area normalization method. The peak area percentages of the ADCs containing 0, 1, 2, 3, 4, 5, 6, 7, and 8 cytotoxic drugs were calculated, and the DAR values ​​were calculated. The calculation formula is: DAR value = (peak area percentage of ADC containing 0 cytotoxic drug × 0 + peak area percentage of ADC containing 1 cytotoxic drug × 1 + peak area percentage of ADC containing 2 cytotoxic drugs × 2 + peak area percentage of ADC containing 3 cytotoxic drugs × 3 + peak area percentage of ADC containing 4 cytotoxic drugs × 4 + peak area percentage of ADC containing 5 cytotoxic drugs × 5 + peak area percentage of ADC containing 6 cytotoxic drugs × 6 + peak area percentage of ADC containing 7 cytotoxic drugs × 7 + peak area percentage of ADC containing 8 cytotoxic drugs × 8) / 100%.

[0304] The perrituzumab-eribulin conjugate was prepared and determined by the methods of Examples 4 and 5, and had the following structure:

[0305]

[0306] Among them, the measured DAR value of patrastuzumab-eribulin-D2 was 2.6; the measured DAR value of patrastuzumab-eribulin-D4 was 4-4.2; and the measured DAR value of patrastuzumab-eribulin-D8 was 7.6.

[0307] Pertratuzumab-DDDXd-D8 was prepared and determined by the methods of Examples 4 and 5, and has the following structure:

[0308]

[0309] Its measured DAR value is 7.9.

[0310] Example 6 Aggregation Verification of Antibody Drug Conjugates

[0311] The components of the pertuzumab-drug conjugate prepared in Example 4 were separated using a gel chromatography column. A neutral pH buffer solution supplemented with 10% isopropanol was used as the mobile phase for elution, and the components were eluted in descending order of molecular weight. The column was an ACQUITY UPLC Protein BEH SEC Column. A 1.7 μm, 4.6 × 300 mm gel chromatography column was used at a column temperature of 25°C. The mobile phase consisted of 50 mM phosphate buffer-200 mM sodium chloride-10% isopropanol, pH 7.0 (12.53 g of disodium hydrogen phosphate dodecahydrate, 2.33 g of sodium dihydrogen phosphate dihydrate, and 11.69 g of sodium chloride were weighed, added to approximately 800 mL of ultrapure water, and stirred until fully dissolved. The mixture was then filled to 1000 mL with ultrapure water and set aside. 100 mL of isopropanol was added to the above solution to 1000 mL, mixed, and filtered through a 0.22 μm filter). A precise amount of 20 μg of the patrastuzumab-drug conjugate was injected into the liquid chromatograph and detected at a wavelength of 280 nm. The flow rate was 0.3 mL / min, and isocratic elution was performed for 15 min.

[0312] Data processing and quantitative analysis of the results were performed using the area normalization method. The peak area percentages of aggregates, immunoglobulin monomers, and low-molecular-weight impurities were calculated. Aggregates are present before the main peak, immunoglobulin monomers are present during the main peak, and low-molecular-weight impurities are present after the main peak. The percentages of monomers, aggregates, and low-molecular-weight impurities in the pertrastuzumab-drug conjugate are shown in Table 2.

[0313] Table 2 Contents of monomers, aggregates and low molecular weight impurities of patrastuzumab-drug conjugates

[0314]

[0315] Example 7 Cell Binding Activity of Antibody Drug Conjugates

[0316] Based on the FACS method, using pertrastuzumab-DDDXd-D8 and pertrastuzumab as controls, the binding activity of the pertrastuzumab-eribulin conjugate prepared in Example 4 above to cells with different HER3 expression levels was analyzed, including MCF-7, HCC1569, and BT474 cells with high HER3 expression levels, MDA-MB-468 and JIMT-1 cells with medium HER3 expression levels, SW620 cells with low HER3 expression levels, and HER3-negative A549 cells.

[0317] 1×10 5Cells were added to each well of a 96-well cell culture plate. The pastratumomab-eribulin conjugate was diluted four-fold in FACS buffer (Miltenyi Biotec, Catalog No. 130-091-221) at a starting concentration of 135.14 nM and then incubated at 4°C for 60 min. The cells were centrifuged at 1000 rpm for 5 min, the supernatant discarded, and the plates washed three times with pre-chilled PBS (pH 7.4). 100 μL / well of a 1:200 (v / v) goat anti-human IgG Fcγ-PE secondary antibody (Jackson Immunoresearch, Catalog No. 109-116-170) was added and incubated at 4°C for 30 min. The plates were washed three times with pre-chilled PBS (pH 7.4) and resuspended in 100 μL of PBS (pH 7.4). Fluorescence signals were then analyzed using a flow cytometer (Sartorius, iQUE). The binding activity of the patrastuzumab-eribulin conjugate and the control to the above cells was measured by the mean fluorescence intensity (MFI) of the staining. The data were analyzed using GraphPad Prism5, and the results are shown in Figures 1A-1G. The EC 50 The results are shown in Table 3. The results showed that the binding activity of the pertuzumab-eribulin conjugates at various DAR values ​​to cells with high, medium, and low HER3 expression levels was comparable to that of pertuzumab, but they did not bind to HER3-negative A549 cells.

[0318] Table 3 Binding activity of patrastuzumab-eribulin conjugate to cells with different HER3 expression levels

[0319]

[0320] *: Not tested.

[0321] Example 8 Endocytosis Experiment of Antibody Drug Conjugate

[0322] Based on the FACS method, using Pertratuzumab-DDDXd-D8 and Pertratuzumab as controls, the endocytosis of the Pertratuzumab-Eribulin conjugate prepared in Example 4 above in cells with different HER3 expression levels was analyzed, including MCF-7 and BT474 cells with high HER3 expression levels, NCI-N87 cells with medium HER3 expression levels, and SW620 cells with low HER3 expression levels.

[0323] Adjust the cell density to 1×10 6 / mL, 50μL / well was added to a 96-well cell culture plate. Sample preparation: The patrastuzumab-eribulin conjugate was pre-diluted to a concentration of 20μg / mL, marked as S1, and then diluted in a 3-fold concentration gradient to obtain 9 samples S1-S9; the gradient diluted sample solution was added to the cell culture plate, 50μL was added to each well, and incubated at 4°C for 30min. After the incubation, the 96-well cell culture plate was removed, placed at 4°C, centrifuged at 400g for 4min, and the supernatant was discarded. Dilute pHrodo at 1:200 (v / v) TM Green Maleimide (Green Maleimide; Invitrogen, Catalog No.: P35370) labeled AffiniPure Goat Anti-Human IgG, Fcγfragment specific (Goat anti-human IgG, Fc fragment specific antibody; Jackson Immuno; Catalog No.: 109-005-190) was added to each well and incubated at 4°C. After 30 minutes of washing, 50 μL of cell culture medium was added to each well, mixed, and incubated at 37°C for 2 hours. The cells were placed on a flow cytometer (Sartorius, iQUE) and the fluorescence readings of the BL1 channel were measured. Data were analyzed using GraphPad Prism5, and the results are shown in Figures 2A-2D. The calculated EC 50 The results are shown in Table 4. The results showed that the pascataract-eribulin conjugates with different DAR values ​​exhibited significant endocytosis in cells with high and medium HER3 expression levels, but had weak endocytosis in SW620 cells with low HER3 expression levels.

[0324] Table 4 Endocytosis of the patrastuzumab-eribulin conjugate in cells with different HER3 expression levels

[0325]

[0326] Example 9 Cell Killing Activity of Antibody Drug Conjugates

[0327] To detect the inhibitory effect of the pertuzumab-eribulin conjugate prepared in Example 4 on tumor cell proliferation, pertuzumab-DDDXd-D8 was used as a control, and the killing activity was detected using cells with different HER3 expression levels, including BT474, MCF-7, and HCC1569 cells with high HER3 expression levels, SKBR3, NCI-N87, JIMT-1, and MDA-MB-468 cells with medium HER3 expression levels, and SW620 and WiDr cells with low HER3 expression levels.

[0328] Cells in the logarithmic growth phase were added to a 96-well plate at a density of 1 × 10 4 / mL or 2×104 Cells were cultured overnight at 37°C and 5% CO2. Sample preparation: The patrastuzumab-eribulin conjugate was prepared as the test sample (starting at 5 μg / mL, with a 5-fold serial dilution series over nine steps) in basal medium containing 10% FBS. Cells were removed from the overnight adherent culture and 50 μL / well of the diluted test sample was added to the experimental group, while 50 μL / well of basal medium containing 10% FBS was added to the control group. After continued culture for 96 h, 120 h, or 144 h, the cells were assayed using the CellTiter-Glo Luminescent Cell Assay Kit (Promega, Catalog No. G7572). The 96-well plate was removed and 75 μL of CTG detection solution (Promega, Catalog No. G7572) was added to each well. The cells were mixed by vortexing and incubated in the dark at room temperature for 10 min. After that, 180 μL of each well was transferred to an opaque white plate. Air bubbles were removed, and the chemiluminescence value was read. The killing rate was calculated.

[0329] Killing rate (%) = (1-luminescence value of experimental group / luminescence value of control group) × 100%.

[0330] The data were analyzed and processed using Graphpad Prism5. The results are shown in Figures 3A-3I. The calculated EC 50 The results are shown in Tables 5-1 and 5-2 below. The results show that for cells with high, medium, and low HER3 expression levels, the larger the DAR value, the stronger the killing activity of the pertuzumab-eribulin conjugate, and the killing activity of the pertuzumab-eribulin conjugates at various DAR values ​​is superior to that of pertuzumab-DDDXd-D8.

[0331] Table 5-1 Cytotoxicity of Patrastuzumab-Eribulin Conjugate against Cells with High HER3 Expression

[0332]

[0333] Table 5-2 Cytotoxicity of patrastuzumab-eribulin conjugate against cells expressing HER3 at medium level

[0334]

[0335] Example 10 Pharmacodynamic Evaluation of Antibody Drug Conjugates in a JIMT-1 Human Breast Cancer Cell Subcutaneous Xenograft Tumor Model in Nude Mice

[0336] The in vivo efficacy of the trastuzumab-eribulin conjugate prepared in Example 4 was evaluated using a nude mouse subcutaneous transplantation tumor model of trastuzumab-resistant cell line JIMT-1 human breast cancer cells.

[0337] JIMT-1 cells were subcutaneously inoculated in the right axilla of SPF female nude mice (source: Changzhou Cavens Laboratory Animal Co., Ltd.) at a rate of 2 × 106 When the average tumor volume reaches 100-300mm 3 The animals were divided into 5 groups, with 6 animals in each group. The specific grouping and dosing schedule are shown in Table 6.

[0338] Table 6 Grouping and dosing regimen

[0339] Group Drug dose (mg / kg) Administration route Administration frequency 1 Vehicle control N / A Tail vein injection Q1W2 Pertratuzumab-DDDXd-D83 Tail vein injection Q1W3 Pertratuzumab-eribulin-D23 Tail vein injection Q1W4 Pertratuzumab-eribulin-D43 Tail vein injection Q1W5 Pertratuzumab-eribulin-D83 Tail vein injection Q1W

[0340] Q1W: Administer once a week.

[0341] The day of grouping was designated as day 0, and administration was performed via the tail vein on day 1. Tumor volume was measured 2-3 times per week, and mice were weighed and recorded. General performance of the mice was observed and recorded daily. Tumors were removed, weighed, and photographed at the end of the experiment.

[0342] Detection indicators include:

[0343] Tumor volume TV (mm 3 )=1 / 2×(a×b 2 ), where a is the major diameter and b is the minor diameter.

[0344] Relative tumor volume RTV = TV t / TV0; TV0 is the tumor volume on day d0, TV t is the tumor volume at each measurement.

[0345] Relative tumor growth rate T / C (%) = T RTV / C RTV ×100%; where T RTV RTV for the treatment group, C RTV RTV was used as the control group.

[0346] Tumor growth inhibition rate: 1-T / C.

[0347] Tumor inhibition rate TGI (%) = (1-TW / TW0) × 100%; wherein, TW is the tumor weight of the treatment group, and TW0 is the tumor weight of the control group.

[0348] Body weight change rate, WCR (%) = (Wt t -Wt0) / Wt0×100%; where Wt0 is the weight of the animal on day d0, Wt t is the animal weight at each measurement.

[0349] The effects of each drug on tumor volume, tumor weight, and mouse body weight are shown in Figures 4-6, and the test index results are shown in Table 7 below. By the end of the experiment on day 21, no animals had died, and the weight gain of mice in each treatment group was comparable to that of the model group. The drugs showed no significant toxic effects, demonstrating a good safety profile. Pertratuzumab-eribulin conjugates with various DAR values ​​exhibited significant in vivo tumor growth inhibition activity, and were superior to Pertratuzumab-DDDXd-D8. The greater the DAR value of the Pertratuzumab-eribulin conjugate, the stronger its in vivo tumor growth inhibition activity.

[0350] Table 7 Tumor inhibitory effect of patrastuzumab-eribulin conjugate in JIMT-1 human breast cancer cell xenograft model in nude mice

[0351] Tumor growth inhibition rate of group 1-T / C Tumor inhibition rate TGI 224.4% 26.6% 336.6% 41.1% 465.9% 68.9% 582.9% 84.3%

[0352] According to the contents disclosed in the present invention, although the method of the present invention has been described according to a preferred embodiment, it is apparent to those skilled in the art that the method described herein and the steps or the order of steps of the method may be changed without departing from the concept, spirit and scope of the present invention.

[0353] The disclosures of all documents cited herein are hereby incorporated by reference, to the extent that they provide exemplary, procedural and other details supplementary to those set forth herein.

Claims

1. A substance with the general formula Ab-(LU) n An antibody-drug conjugate or its pharmaceutically acceptable salt or solvate, wherein Ab represents the antibody moiety, L represents the linker moiety, U represents the cytotoxic drug moiety, and n is an integer or decimal selected from 1 to 10, characterized in that... The antibody-drug conjugate comprises the structure shown in Formula IIa: in, R a Selected from hydrogen atoms, deuterium atoms, and optionally substituted C atoms 1-6 Alkyl groups, optionally substituted C 3-7 cycloalkyl, optionally substituted C 3-7 Heterocyclic group, optionally substituted C 6-10 aryl, optional substituted C 5-12 Mixed aromatics; R b Selected from hydrogen atoms, deuterium atoms, and optionally substituted C atoms 1-6 Alkyl groups, optionally substituted C 3-7 cycloalkyl, optionally substituted C 3-7 Heterocyclic group, optionally substituted C 6-10 aryl, optional substituted C 5-12 Mixed aromatics; or, R a With R b Together with the atoms connected to it, they form optional 5- to 8-membered heterocyclic groups that can be substituted.

2. The antibody-drug conjugate according to claim 1, or a pharmaceutically acceptable salt or solvate thereof, wherein, The antibody-drug conjugate comprises the structure shown in Formula IIIa:

3. The antibody-drug conjugate according to claim 1 or 2, or a pharmaceutically acceptable salt or solvate thereof, wherein, The antibody-drug conjugate has the structure shown in Formula IV: in, Ab represents the antibody portion. n is an integer or decimal selected from 1 to 10.

4. The antibody-drug conjugate or its pharmaceutically acceptable salt or solvate according to any one of claims 1-3, wherein, R a With R b Each is independently selected from hydrogen atom, methyl, ethyl, propyl or isopropyl.

5. The antibody-drug conjugate according to claim 1 or 2, or a pharmaceutically acceptable salt or solvate thereof, wherein, The antibody-drug conjugate comprises the structure shown in Formula IIIa-1:

6. The antibody-drug conjugate according to claim 3, or a pharmaceutically acceptable salt or solvate thereof, wherein, The antibody-drug conjugate has the structure shown in Formula IV-1:

7. The antibody-drug conjugate or its pharmaceutically acceptable salt or solvate according to any one of claims 1-6, wherein, The value of n is 2-4.8, 2.6-4.8, 3.5-4.8, 4-4.8, 2-4.5, 2.6-4.5, 3.5-4.5, 4-4.5, 3.5-4.2, 3.5-4, 4-4.2, 7-8, 7-7.9, 7-7.6, 7-7.5, 7.1-8, 7.1-7.9, 7.1-7.6, 7.5-8, 7.6-8, or 7.6-7.

9.

8. The antibody-drug conjugate according to claim 7, or a pharmaceutically acceptable salt or solvate thereof, wherein, The n is approximately 2.6, approximately 4, approximately 4.2, approximately 4.8, approximately 7, approximately 7.1, approximately 7.5, approximately 7.6, approximately 7.9, or approximately 8.

9. The antibody-drug conjugate or its pharmaceutically acceptable salt or solvate according to any one of claims 1-8, wherein, The Ab is an anti-HER3 antibody or its antigen-binding fragment.

10. The antibody-drug conjugate according to claim 9, or a pharmaceutically acceptable salt or solvate thereof, wherein, The anti-HER3 antibody or its antigen-binding fragment comprises HCDR1 containing the amino acid sequence shown in SEQ ID NO:1, HCDR2 containing the amino acid sequence shown in SEQ ID NO:2, HCDR3 containing the amino acid sequence shown in SEQ ID NO:3, LCDR1 containing the amino acid sequence shown in SEQ ID NO:4, LCDR2 containing the amino acid sequence shown in SEQ ID NO:5, and LCDR3 containing the amino acid sequence shown in SEQ ID NO:

6.

11. The antibody-drug conjugate according to claim 10, or a pharmaceutically acceptable salt or solvate thereof, wherein, The anti-HER3 antibody or its antigen-binding fragment comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:7, and the light chain variable region comprises an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:8; or the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO:7, and the light chain variable region comprises the amino acid sequence shown in SEQ ID NO:

8.

12. The antibody-drug conjugate according to claim 10 or 11, or a pharmaceutically acceptable salt or solvate thereof, wherein, The anti-HER3 antibody or its antigen-binding fragment comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:9, and the light chain comprises an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:

10.

13. The antibody-drug conjugate according to claim 9, or a pharmaceutically acceptable salt or solvate thereof, wherein, The anti-HER3 antibody is pertrastuzumab.

14. The antibody-drug conjugate or its pharmaceutically acceptable salt or solvate according to any one of claims 9-13, wherein, The antibody-drug conjugate or its pharmaceutically acceptable salt or solvate exhibits one or a combination of the following properties: (a) Binding to HER3; (b) Blocking the binding of HER3 to its ligand; (c) Demonstrates endocytosis in cells expressing HER3; (d) It has killing activity against tumor cells expressing HER3; (e) It has the bystander effect.

15. A pharmaceutical composition comprising the antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof as described in any one of claims 1-14; optionally, the pharmaceutical composition further comprising a pharmaceutically acceptable carrier.

16. Use of the antibody-drug conjugate of any one of claims 1-14 or a pharmaceutically acceptable salt or solvate thereof, or the pharmaceutical composition of claim 15, in the preparation of a medicament for treating cancer; preferably, the cancer is HER3-positive cancer; preferably, the cancer is biliary tract cancer, carcinosarcoma, esophageal cancer, gastroesophageal junction cancer, breast cancer, gastric cancer, pancreatic cancer, head and neck cancer, colorectal cancer, kidney cancer, cervical cancer, ovarian cancer, endometrial cancer, uterine cancer, melanoma, pharyngeal cancer, oral cancer, skin cancer, lung cancer, glioblastoma multiforme, urothelial carcinoma, prostate cancer, bladder cancer, gastrointestinal stromal tumor, squamous cell carcinoma, peritoneal cancer, liver cancer, salivary gland cancer, vulvar cancer, thyroid cancer, testicular cancer, anal cancer, or penile cancer.

17. A method of treating cancer, comprising administering to a patient in need a therapeutically effective amount of the antibody-drug conjugate of any one of claims 1-14 or a pharmaceutically acceptable salt or solvation thereof, or the pharmaceutical composition of claim 15; preferably, the cancer is HER3-positive cancer; preferably, the cancer is biliary tract cancer, carcinosarcoma, esophageal cancer, gastroesophageal junction cancer, breast cancer, gastric cancer, pancreatic cancer, head and neck cancer, colorectal cancer, kidney cancer, cervical cancer, ovarian cancer, endometrial cancer, uterine cancer, melanoma, pharyngeal cancer, oral cancer, skin cancer, lung cancer, glioblastoma multiforme, urothelial carcinoma, prostate cancer, bladder cancer, gastrointestinal stromal tumor, squamous cell carcinoma, peritoneal cancer, liver cancer, salivary gland cancer, vulvar cancer, thyroid cancer, testicular cancer, anal cancer, or penile cancer.

18. The method of claim 17, wherein the method comprises contacting tumor cells with the antibody-drug conjugate or a pharmaceutically acceptable salt or solvate thereof, or the pharmaceutical composition, thereby killing tumor cells or inhibiting tumor cell growth.

19. A linker-pharmaceutical intermediate compound having the structure shown in Formula III: in, R a Selected from hydrogen atoms, deuterium atoms, and optionally substituted C atoms 1-6 Alkyl groups, optionally substituted C 3-7 cycloalkyl, optionally substituted C 3-7 Heterocyclic group, optionally substituted C 6-10 aryl, optional substituted C 5-12 Mixed aromatics; R b Selected from hydrogen atoms, deuterium atoms, and optionally substituted C atoms 1-6 Alkyl groups, optionally substituted C 3-7 cycloalkyl, optionally substituted C 3-7 Heterocyclic group, optionally substituted C 6-10 aryl, optional substituted C 5-12 Mixed aromatics; Or R a With R b Together with the atoms connected to it, they form optional 5- to 8-membered heterocyclic groups that can be substituted.

20. The linker-pharmaceutical intermediate compound according to claim 19, characterized in that, R a With R b Each is independently selected from hydrogen atom, methyl, ethyl, propyl or isopropyl.