Antibody-drug conjugates containing N-methylene amide linkers

Abstract

The present invention relates to a linking structure for linking an antibody to a drug toxic to malignant cells, and provides an antibody-drug conjugate represented by formula (I) obtained by linking an antibody to a cytotoxic drug via the linking structure. The definitions of each group in the formula are as described in the specification, and the antibody-drug conjugate has a significant antitumor effect. [Formula 1] JPEG2026501616000130.jpg23170

Description

[Technical Field]

[0001] The present invention relates to the field of biomedicine. Specifically, the present invention provides antibody-drug conjugates containing linkers of methylene amide and similar structures, their preparation methods, and uses. [Background technology]

[0002] Antibody-targeting conjugates are a highly effective class of therapeutic agents, particularly in the treatment of tumor diseases. Linking the drug to the antibody requires a linker, which not only functions as a linker but also controls the method and rate of intracellular drug release, thereby affecting drug efficacy.

[0003] Existing linker release technologies include para-aminobenzyl alcohol carboxylic acid ester amide (PAB-carbonyl fragment), hemiaminal, cleavable peptide chain, non-cleavable peptide chain, etc. It is of great significance to develop more antibody-drug conjugates containing intracellularly cleavable linkers. Summary of the Invention

[0004] The object of the present invention is to provide a linking structure for linking an antibody to a drug toxic to malignant cells, and to obtain an antibody-drug conjugate in which the antibody is linked to a cytotoxic drug via the linking structure, and the antibody-drug conjugate has a significant antitumor effect.

[0005] In one embodiment of the present invention, there is provided an antibody-drug conjugate according to formula (I): [ka] (In the formula, D is a cytotoxic drug and Ab is an antibody; L D is absent or selected from a C1-C3 alkylene group; X is selected from N, O or S; R 1 is selected from absent, hydrogen, deuterium, a C1-C6 alkyl group, or a halogen-substituted C1-C6 alkyl group; R 2 is selected from hydrogen, deuterium, a C1-C6 alkyl group, a C1-C6 alkoxy group, an acyl group, and a sulfonyl group; L 1 -L 11 -L 12 -L 13 - selected from, where L 11 , L 12 , and L 13 are each independently selected from absent, —C═O—, a C1-C6 alkylene group, or —O—C1-C6 alkylene; L 2 is selected from a C1-C6 alkylene group or a C1-C6 acyl group, and the C1-C6 alkylene group or the C1-C6 acyl group is selected from one or more R 3 optionally substituted with R 3 is selected from phenyl substituted or unsubstituted C1-C6 alkyl groups and C1-C6 alkoxy groups; L P is a peptide residue consisting of 2 to 7 amino acids, Z is -L z -L j - selected from, where L z is a -C(=O)-C1-C8 alkylene group or -C(=O)-(CH2CH2O) 2~6 -CH2CH2NH-, L j is a linker that can be linked to an antibody.

[0006] In the antibody-drug conjugate of formula (I), the drug-antibody ratio (DAR) is selected from the range of 2 to 8, and may further be 6 to 8 or 6.2 to 7.8, for example, 6.2, 6.3, 6.5, 6.6, 6.8, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.78, or 7.8.

[0007] The antibody-drug conjugate shown in formula (I) is an antibody-drug conjugate shown in formula (I'). [ka] wherein n' is selected from 2 to 8, and may further be 6 to 8 or 6.2 to 7.8, such as 6.2, 6.3, 6.5, 6.6, 6.8, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.78, or 7.8; The definitions of each group in the formula are the same as above.

[0008] In some specific embodiments, R 1 is selected from absent, hydrogen, deuterium, a C1-C3 alkyl group, or a halogen-substituted C1-C3 alkyl group.

[0009] In some specific embodiments, R 1 is selected from absent, hydrogen, deuterium, or a halogen-substituted C1-C3 alkyl group.

[0010] In some specific embodiments, R 1 is selected from absent, hydrogen, deuterium, or a C1-C3 alkyl group substituted with F, Cl, Br, or I.

[0011] In some specific embodiments, R 1 is selected from absent, hydrogen, deuterium, or an F-substituted C1-C3 alkyl group.

[0012] In some specific embodiments, R 1 is selected from absent, hydrogen, deuterium, a fluoromethyl group, a difluoromethyl group, a trifluoromethyl group, a fluoroethyl group, a difluoroethyl group, or a trifluoroethyl group. In some specific embodiments, R 1 is selected from absent, hydrogen, a difluoroethyl group, or a trifluoroethyl group.

[0013] In some specific embodiments, R 2 is selected from hydrogen, deuterium, a C1-C6 alkyl group, a C1-C6 alkoxy group, a -C(=O)C1-C6 alkyl group, or a -S(=O)2C1-C6 alkyl group.

[0014] In some specific embodiments, R 2 is selected from hydrogen, deuterium, a C1-C6 alkyl group, a C1-C6 alkoxy group, a -C(=O)C1-C6 alkyl group, or a -S(=O)2C1-C3 alkyl group.

[0015] In some specific embodiments, R 2 is selected from hydrogen, deuterium, a C1-C3 alkyl group, a C1-C3 alkoxy group, a -C(=O)C1-C3 alkyl group, or a -S(=O)2C1-C3 alkyl group.

[0016] In some specific embodiments, R 2 is selected from hydrogen, deuterium, methyl, ethyl, methoxy, ethoxy, formyl, acetyl, methanesulfonyl or ethanesulfonyl.

[0017] In some specific embodiments, R 2 is selected from hydrogen, methyl, methoxy, formyl, or methanesulfonyl.

[0018] In some specific embodiments, R 2 is selected from hydrogen, formyl, or methanesulfonyl.

[0019] In some specific embodiments, L 11 , L 12 , and L 13 are each independently selected from absent, —C(═O)—, a C1-C2 alkylene group, or a C1-C2 alkylene-O—, with the proviso that L 11 When is -C(=O)-, R 2 is not hydrogen.

[0020] In some specific embodiments, L 11 , L 12 , and L 13 are each independently selected from absent, —C═O—, —CH2—, or —OCH2—.

[0021] In some specific embodiments, L 1 is selected from -C(=O)CH2OCH2-, -C(=O)CH2O- or -CH2-.

[0022] In some specific embodiments, L 2 is selected from a C1-C3 alkylene group or a C1-C3 acyl group, and the C1-C3 alkylene group or the C1-C3 acyl group is selected from one or more R 3 is optionally replaced by

[0023] In some specific embodiments, L 2 is selected from a methylene group, an ethylene group, or —C(═O)CH—, and the methylene group, ethylene group, or —C(═O)CH— is selected from one or more R 3 is optionally replaced by

[0024] In some specific embodiments, R 3 is selected from a C1 to C6 alkyl group, a C1 to C6 alkoxy group, or a phenyl-substituted C1 to C6 alkyl group.

[0025] In some specific embodiments, R 3 is selected from C1 to C6 alkyl groups or phenyl-substituted C1 to C6 alkyl groups.

[0026] In some specific embodiments, R 3 is selected from C1 to C4 alkyl groups or phenyl-substituted C1 to C3 alkyl groups.

[0027] In some specific embodiments, R 3is selected from a methyl group, an ethyl group, a propyl group, an isopropyl group, a butyl group, an isobutyl group, a phenylmethyl group or a phenylethyl group.

[0028] In some specific embodiments, R 3 is selected from a methyl group, an isopropyl group, an isobutyl group, or a phenylmethyl group.

[0029] In some specific embodiments, L 2 is selected from -CH2-, -CH2CH2-, or a group represented by the following formula: [ka]

[0030] In some specific embodiments, L 2 is selected from -CH2-, -CH2CH2-, or a group represented by the following formula: [ka]

[0031] In some specific embodiments, L 2 is selected from -CH2-, -CH2CH2-, or a group represented by the following formula: [ka]

[0032] In some specific embodiments, the amino acid is selected from phenylalanine (Phe), glycine (Gly), valine (Val), alanine (Ala), leucine (Leu), lysine (Lys), or glutamic acid (Glu).

[0033] In some specific embodiments, the amino acid is selected from phenylalanine (Phe), glycine (Gly), valine (Val), alanine (Ala), or leucine (Leu).

[0034] In some specific embodiments, L p is selected from peptide residues consisting of 2 to 5 amino acids selected from phenylalanine (Phe), glycine (Gly), valine (Val), alanine (Ala), and leucine (Leu).

[0035] In some specific embodiments, L p is selected from peptide residues consisting of 2, 3 or 4 amino acids selected from phenylalanine (Phe), glycine (Gly), valine (Val), alanine (Ala) or leucine (Leu).

[0036] In some specific embodiments, L p is selected from -Val-Cit-, -Gly-Lys-, -Gly-Leu-, -Val-Ala-, -Gly-Phe-, -GLy-Gly-Lys-, -Gly-Gly-Phe-, -Gly-Val-Ala-, -Gly-Gly-Val-, -Gly-Leu-Val-, -Gly-Phe-Gly-, or -Gly-Gly-Leu-.

[0037] In some specific embodiments, L p is selected from -Gly-Leu-, -Gly-Phe-, -Gly-Gly-Phe-, -Gly-Val-Ala-, -Gly-Gly-Val-, -Gly-Leu-Val-, -Gly-Phe-Gly or -Gly-Gly-Leu-.

[0038] In some specific embodiments, L p is selected from the groups represented by the following formulas: [ka]

[0039] In some specific embodiments, L p is selected from the groups represented by the following formulas: [ka]

[0040] In some specific embodiments, L j is selected from groups represented by the following formulas: [ka] [ka] The position indicated by indicates that it is linked to an antibody, [ka] The position indicated by L z represents that the group is linked to the

[0041] In some specific embodiments, L z is a -C(=O)-C1-C8 alkenyl group or -C(=O)-(CH2CH2O) 2~6 -CH2CH2NH-.

[0042] In some specific embodiments, L z is -C(=O)-(CH2CH2O) 2~6 -CH2CH2NH-.

[0043] In some specific embodiments, L z is -C(=O)-(CH2CH2O) 2~4 -CH2CH2NH-.

[0044] In some specific embodiments, L z is selected from —C(═O)—(CH2CH2O)2—CH2CH2NH—.

[0045] In some specific embodiments, Z is [ka] is selected from.

[0046] In some specific embodiments, said cytotoxic drug is selected from camptothecin and its derivatives.

[0047] In some specific embodiments, the cytotoxic drug is selected from those shown in the following formula: [ka]

[0048] In some specific embodiments, the antibody is an anti-tumor-associated antigen antibody.

[0049] In some specific embodiments, the tumor-associated antigen antibody is selected from an anti-Her2 antibody, an anti-Trop2 antibody, an anti-B7H3 antibody, an anti-5T4 antibody, an anti-Nectin-4 antibody, an anti-CD20 antibody, and an anti-ROR1 antibody.

[0050] In some specific embodiments, the structure of the compound of formula (I) is as shown in formula (II) or formula (II'). [ka] (In the formula, R 2 , L 1 , L 2 , L p , Z, n', and Ab are defined as in the antibody-drug conjugate of formula (I) or formula (I'), respectively.)

[0051] In some specific embodiments, L 1 is selected from -C(=O)CH2OCH2-, or -C(=O)CH2O.

[0052] In some specific embodiments, L 2 is selected from -CH2- or -CH2CH2-.

[0053] In some specific embodiments, [ka] teeth, [ka] is selected from.

[0054] In some specific embodiments, the structure of the antibody-drug conjugate shown in formula (I) is shown in formula (III) or formula (III'). [ka] (In the formula, R 1 , R 2 , R 4 , L p , Z, n', and Ab are defined as in the antibody-drug conjugate of formula (I) or formula (I'), respectively.)

[0055] In some specific embodiments, R 4 is selected from hydrogen, phenyl substituted or unsubstituted C1 to C6 alkyl groups, and C1 to C6 alkoxy groups.

[0056] In some specific embodiments, R 4 is selected from hydrogen, a C1-C6 alkyl group, a C1-C6 alkoxy group, or a phenyl-substituted C1-C6 alkyl group.

[0057] In some specific embodiments, R 4 is selected from hydrogen, a C1-C6 alkyl group, or a phenyl-substituted C1-C6 alkyl group.

[0058] In some specific embodiments, R 4 is selected from hydrogen, a C1-C4 alkyl group, or a phenyl-substituted C1-C3 alkyl group.

[0059] In some specific embodiments, R 4is selected from hydrogen, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, phenylmethyl or phenylethyl.

[0060] In some specific embodiments, R 4 is selected from hydrogen, a methyl group, an isopropyl group, an isobutyl group, or a phenylmethyl group.

[0061] In some specific embodiments, R 4 is selected from hydrogen, a methyl group, or an isopropyl group.

[0062] In some specific embodiments, the structure of the antibody-drug conjugate shown in formula (I) is shown in formula (IV) or formula (IV'). [ka] (In the formula, R 2 , L 1 , L 2 , L p , Z, n', and Ab are defined as in the antibody-drug conjugate of formula (I) or formula (I'), respectively.

[0063] In some specific embodiments, L 1 and L 2 are each independently selected from C1 to C3 alkylene groups.

[0064] In some specific embodiments, L 1 is -CH2-, and L 2 is -CH2CH2-.

[0065] In some specific embodiments, [ka] is selected from the following structures: [ka]

[0066] In some specific embodiments, [ka] is selected from the following structures: [ka] (In the formula, R 1 is defined as in the antibody-drug conjugate of formula (I).

[0067] Antibody-drug conjugates obtained by linking the following compounds with antibodies: JPEG2026501616000023.jpg159170JPEG2026501616000024.jpg144170

[0068] Antibody-drug conjugates obtained by linking the following compounds with antibodies: JPEG2026501616000025.jpg155170JPEG2026501616000026.jpg142170

[0069] In some specific embodiments, the antibody is an HS627 antibody or an IP140B antibody, wherein the heavy chain amino acid sequence of HS627 is set forth in SEQ ID NO:1 and the light chain amino acid sequence is set forth in SEQ ID NO:2, and the heavy chain amino acid sequence of IP140B antibody is set forth in SEQ ID NO:3 and the light chain amino acid sequence is set forth in SEQ ID NO:4.

[0070] The following antibody-drug conjugates: JPEG2026501616000027.jpg161170JPEG2026501616000028.jpg144170where, 2 <n<8である。

[0071] In one embodiment, n is selected from 6 to 8 or 6.2 to 7.8, and may be, for example, 6.2, 6.3, 6.5, 6.6, 6.8, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.78 or 7.8.

[0072] In some specific embodiments, the Ab is selected from tumor-associated antigen antibodies, and further selected from anti-Her2 antibody, anti-Trop2 antibody, anti-B7H3 antibody, anti-5T4, anti-Nectin-4 antibody, anti-CD20 antibody, and anti-ROR1 antibody.

[0073] In another aspect of the present invention, there is provided a method for producing the above antibody-drug conjugate, the method being selected from the following synthetic routes:

[0074] Synthetic Route 1: [ka] The compound of formula 1-1 is reacted to give a compound of formula 1-2, the compound of formula 1-2 is reacted with D-OH to give a compound of formula 1-3, the Fmoc protecting group is removed from the compound of formula 1-3 to give a compound of formula 1-4, and the compound of formula 1-4 is reacted with HL p -OH to give a compound of formula 1-5, which is condensed with Z'-OH to give a compound of formula 1-6, which is coupled to an antibody to give a compound of formula 1-7.

[0075] Synthetic Route 2: [ka] The compound of formula 1-2 is reacted to obtain a compound of formula 2-2, and the compound of formula 2-2 is reacted with compound R 1 -NH-L D -D to give a compound of formula 2-3, the Fmoc protecting group is removed from the compound of formula 2-3 to give a compound of formula 2-4, and the compound of formula 2-4 is reacted with HL p -OH to give a compound of formula 2-5, which is condensed with Z'-OH to give a compound of formula 2-6, which is coupled to an antibody to give a compound of formula 2-7.

[0076] Synthetic Route 3: [ka] The compound of formula 3-1 is R 2 -Cl to give a compound of formula 3-2, reacting the compound of formula 3-2 to give a compound of formula 3-3, reacting the compound of formula 3-3 with D-OH to give a compound of formula 3-4, removing the Fmoc protecting group from the compound of formula 3-4 to give a compound of formula 3-5, and converting the compound of formula 3-5 into HL p -OH to give a compound of formula 3-6, which is condensed with Z'-OH to give a compound of formula 3-7, which is coupled to an antibody to give a compound of formula 3-8.

[0077] Synthetic Route 4: [ka] The compound of formula 4-1 is R 2 -Cl to give a compound of formula 4-2, which is reacted to give a compound of formula 4-3, which is further reacted to give a compound of formula 4-4, which is reacted with R 1 -NH-L D -D to give a compound of formula 4-5, and the compound of formula 4-5 is reacted to give a compound of formula 4-6, and the compound of formula 4-6 is reacted with HL p -OH to give a compound of formula 4-7, which is condensed with Z'-OH to give a compound of formula 4-8, which is coupled to an antibody to give a compound of formula 4-9.

[0078] Synthetic Route 5: [ka] The compound of formula 5-1 is reacted to give a compound of formula 5-2, and the compound of formula 5-2 is reacted with R 1 -NH-L D -D to give a compound of formula 5-3, the compound of formula 5-3 is reacted to give a compound of formula 5-4, and the compound of formula 5-4 is reacted with HL p -OH to give a compound of formula 5-5, which is condensed with Z'-OH to give a compound of formula 5-6, which is coupled to an antibody to give a compound of formula 5-7.

[0079] In each of the above synthetic routes, R 1 , R 2 , R 3 , L D , D, L p , Z, and Ab are each defined as above.

[0080] Z' is -L z -L j’ where L z is defined as before, and L j is selected from groups represented by the following formulas: [ka]

[0081] In another aspect of the present invention, there is provided a pharmaceutical composition comprising the above antibody-drug conjugate and a pharmaceutically acceptable carrier.

[0082] In another aspect of the present invention, there is provided a use of the above antibody-drug conjugate or pharmaceutical composition in the manufacture of an anti-tumor medicament.

[0083] In another aspect of the present invention, there is provided a method for suppressing or treating a tumor disease in a patient in need thereof, the method comprising administering to a patient in need thereof the above-described antibody-drug conjugate or pharmaceutical composition.

[0084] In some specific embodiments, the amount of the antibody-drug conjugate or pharmaceutical composition administered is a therapeutically effective amount.

[0085] In some specific embodiments, the tumor is selected from a solid tumor.

[0086] In some specific embodiments, the tumor is selected from lung cancer, non-small cell lung cancer, esophageal squamous cell carcinoma, ovarian cancer, esophageal adenocarcinoma, or breast cancer.

[0087] In some specific embodiments, the lung cancer is non-small cell lung cancer. [Brief explanation of the drawings]

[0088] [Figure 1] 1 is a graph showing the tumor growth curve of the NCI-H1975 lung cancer CDX model with in vivo inhibition by ADC3b in Test Example 1. [Figure 2] 1 shows photographs of tumors after dissection in an NCI-H1975 lung cancer CDX model with in vivo inhibition by ADC3b in Test Example 1. [Figure 3] 1 is a graph showing tumor growth curves in an NCI-H1975 lung cancer CDX model due to in vivo inhibition by ADC11b and ADC15b in Test Example 1. [Figure 4] 1 shows photographs of tumors after dissection in an NCI-H1975 lung cancer CDX model with in vivo inhibition by ADC11b and ADC15b in Test Example 1. [Figure 5] 1 is a graph showing the tumor growth curve of a Calu-6 non-small cell lung cancer CDX model with in vivo inhibition by ADC16b in Test Example 1. [Figure 6] 1 shows photographs of tumors after dissection in a Calu-6 non-small cell lung cancer CDX model with in vivo inhibition by ADC16b in Test Example 1. [Figure 7] 1 is a graph showing the tumor growth curve of the KYSE-150 esophageal squamous cell carcinoma CDX model with in vivo inhibition by ADC16b in Test Example 1. [Figure 8] 1 shows photographs of tumors after dissection in a KYSE-150 esophageal squamous cell carcinoma CDX model with in vivo inhibition by ADC16b in Test Example 1. [Figure 9] 1 is a graph showing tumor growth curves in a KYSE-150 esophageal squamous cell carcinoma CDX model with in vivo inhibition by ADC11b, ADC15b, and ADC2b in Test Example 1. [Figure 10]1 shows photographs of tumors after dissection in a KYSE-150 esophageal squamous cell carcinoma CDX model with in vivo inhibition by ADC11b, ADC15b, and ADC2b in Test Example 1. [Figure 11] 1 is a graph showing the tumor growth curve of the ES-2 ovarian cancer CDX model with in vivo inhibition by ADC16b in Test Example 1. [Figure 12] 1 shows photographs of tumors after dissection in an ES-2 ovarian cancer CDX model with in vivo inhibition by ADC16b in Test Example 1. [Figure 13] 1 is a graph showing tumor growth curves in an OE-21 esophageal adenocarcinoma CDX model with in vivo inhibition by ADC7b, ADC9b, and ADC8b in Test Example 1. [Figure 14] 1 shows photographs of tumors after dissection in an OE-21 esophageal adenocarcinoma CDX model with in vivo inhibition by ADC7b, ADC9b, and ADC8b in Test Example 1. [Figure 15] 1 is a graph showing tumor growth curves in an MDA-MB-231 triple-negative human breast cancer CDX model with in vivo inhibition by ADC11b, ADC15b, ADC2b, and ADC10b in Test Example 1. [Figure 16] 1 shows photographs of tumors after dissection in an MDA-MB-231 triple-negative human breast cancer CDX model with in vivo inhibition by ADC11b, ADC15b, ADC2b, and ADC10b in Test Example 1. DETAILED DESCRIPTION OF THE INVENTION

[0089] I. Definition In the present invention, unless otherwise specified, the scientific and technical terms used herein have the meanings that are commonly understood by those skilled in the art.In addition, the relevant terms and experimental procedures used herein are terms and routine steps that are widely used in the corresponding fields.In addition, in order to better understand the present invention, the definitions and explanations of relevant terms are provided below.

[0090] As used herein, unless otherwise specified, the terms "comprise," "include," "have," and "contain," including their grammatical equivalents, are generally to be understood as open-ended and non-limiting, e.g., not excluding other unrecited elements or steps.

[0091] The compounds of the present disclosure may be asymmetric, e.g., may have one or more stereoisomers. Unless otherwise specified, all stereoisomers, such as enantiomers and diastereomers, are included. The stereoisomers include geometric isomers (e.g., cis- and trans-configurations) and optical isomers (e.g., enantiomers), and are therapeutic agents consisting of individual compounds, racemates, racemic mixtures, and pharmaceutically acceptable salts thereof. Compounds of the present disclosure containing asymmetric carbon atoms can be isolated in optically pure or racemic form. Optically pure forms can be separated from racemic mixtures or synthesized using chiral starting materials or chiral reagents. Racemates, diastereomers, and enantiomers are all within the scope of the present invention.

[0092] Numerical ranges herein refer to individual integers within the specified range, for example, "C1-C6" 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.

[0093] When any variable (e.g., Rn) occurs more than one time in any composition or structure of a compound, its definition at each occurrence is independent. Thus, for example, if a group is substituted with 1 to 5 R, then that group is optionally substituted with up to 5 R, and each occurrence of R is an independent option. Further, combinations of substituents and / or variants thereof are permissible only if such combinations result in stable compounds.

[0094] The term "C1-C6 alkyl group" refers to a straight-chain or branched-chain alkyl group having 1 to 6 carbon atoms. Specific examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, and various branched-chain isomers thereof.

[0095] The term "alkoxy group" refers to -O-(alkyl group) and -O-(unsubstituted cycloalkyl group), where alkyl group is defined above. Non-limiting examples of alkoxy groups include methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy, cyclopentyloxy, and cyclohexyloxy groups. An alkoxy group may be optionally substituted or unsubstituted, and when substituted, the substituents are preferably one or more groups independently selected from alkyl groups, alkenyl groups, alkynyl groups, alkoxy groups, alkylthio groups, alkylamino groups, halogens, sulfhydryl groups, hydrogen groups, nitro groups, chloro groups, cycloalkyl groups, heterocycloalkyl groups, aryl groups, heteroaryl groups, cycloalkoxy groups, heterocycloalkoxy groups, cycloalkylthio groups, heterocycloalkylthio groups, carboxylic acid groups, and carboxylic acid ester groups.

[0096] The term "acyl group" refers to -C(=O)R, where R is a substituted or unsubstituted alkyl group, a substituted or unsubstituted cycloalkyl group, or a substituted or unsubstituted heteroalkyl group.

[0097] The term "sulfonyl" refers to -S(=O)R, where R is a substituted or unsubstituted alkyl group, a substituted or unsubstituted cycloalkyl group, or a substituted or unsubstituted heteroalkyl group.

[0098] [ka] indicates the point of attachment of a chemical bond. 11 -L 12 -L 13 -or-L z -L j -) represents a linking of the corresponding groups in left to right order unless otherwise specified. For example, -L 12 is C1-C2 alkylene-O-, this means that the left side of C1-C2 alkylene-O- is -L 11 -L on the right 13 - indicates that it will be linked to.

[0099] As will be understood by those skilled in the art, in the antibody-drug conjugates of the present application, for example, those represented by formula (I), (II), (III), or (IV), the "-" between the drug-linker fragment and the antibody is intended to represent the linkage between the antibody and the fragment, and is not intended to limit the linkage of one antibody to only one drug-linker fragment. As is well known in the art, one antibody can have multiple intermolecular disulfide bonds, and therefore one antibody can have one or more drugs linked to it.

[0100] Drug or pharmaceutical composition The drugs or pharmaceutical compositions of the present invention can be administered orally, topically, parenterally, or mucosally (e.g., sublingually, by inhalation, or rectally) in dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers. Oral administration is generally preferred. The active agent can be administered orally in the form of capsules, tablets, or the like.

[0101] The term "treatment" includes inhibiting, alleviating, preventing or eliminating one or more symptoms or side effects associated with the disease, condition or disorder being treated.

[0102] The use of the term "inhibition" refers to a control. Those skilled in the art can easily determine the appropriate control for each experiment. For example, the reduction in the response in subjects or cells treated with a compound is compared with the response in subjects or cells not treated with a compound.

[0103] The term "patient" refers to an animal, preferably a mammal, and more preferably a human.

[0104] The term "pharmaceutical composition" means a composition comprising the compound of the present invention or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable ingredient selected from the group consisting of, but not limited to, carriers, diluents, adjuvants, excipients, preservatives, fillers, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, fragrances, antibacterial agents, antifungal agents, lubricants, dispersing agents, temperature-sensitive materials, temperature regulators, adhesives, stabilizers, and suspension aids, depending on the nature of the administration method and dosage form.

[0105] The term "effective amount" or "therapeutically effective amount" refers to a nontoxic but sufficient amount of a drug or agent to achieve a desired effect. In embodiments of the present invention, when treating a patient according to the present invention, the amount of a given drug to be administered will depend on many factors, such as the specific administration protocol, the type and severity of the disease or condition, and the specific characteristics (e.g., body weight) of the subject or host requiring treatment. However, dosages can be routinely determined by methods known in the art depending on the specific circumstances, including, for example, the specific drug employed, the route of administration, the condition being treated, and the subject or host being treated. Generally, dosages used in adult treatment typically range from 0.02 to 5000 mg / day, e.g., about 1 to 1500 mg / day. This required dose can conveniently be expressed as a single dose or as divided doses administered simultaneously (or shortly thereafter) or at appropriate intervals, e.g., two, three, four, or more divided doses per day. Although the above dosage ranges are given, it will be understood by those skilled in the art that the specific effective amount may be appropriately adjusted depending on the condition of the patient and in conjunction with the diagnosis of a physician.

[0106] The term "antibody-drug conjugate (ADC)" refers to a monoclonal antibody to which a biologically active small molecule drug is linked via a chemical bond, and the monoclonal antibody functions as a carrier for delivering the small molecule drug to target cells. In this specification, the meaning of "antibody-conjugated drug" is the same as that of antibody-drug conjugate.

[0107] The term "drug-to-antibody ratio (DAR)" refers to the average number of active ingredient molecules (anti-tumor compound or drug) bound to a single monoclonal antibody.

[0108] Abbreviation: MS: methanesulfonyl group AC: acetyl group Fmoc: 9-fluorenylmethoxycarbonyl group The amino acid constituting the peptide residue LP is Val: valine, and its structural formula is [ka] and Ala: Alanine, its structural formula is [ka] and Gly: Glycine, whose structural formula is [ka] and Phe: Phenylalanine, its structural formula is [ka] and Leu: Leucine, whose structural formula is [ka] is.

[0109] In the present invention, the peptide residue L P can be obtained by an amino acid condensation method known in the art, and the present invention is not particularly limited thereto.

[0110] In the present invention, a linker L for linking antibodies j can be obtained by linking by methods known in the art. For example, L j The structure of [ka] (or [ka] ), then it is [ka] (or [ka] ) with a reactive group such as thiol on the antibody, and the present invention is not particularly limited thereto.

[0111] II. MODE FOR CARRYING OUT THE INVENTION The antibodies used to produce the ADC in this example are not limited to pertuzumab and IP140B antibodies. The heavy chain amino acid sequence of pertuzumab (HS627, PERTUZUMAB) is as follows (SEQ ID NO: 1): [ka]

[0112] The light chain amino acid sequence is as follows (SEQ ID NO:2): [ka]

[0113] The heavy chain amino acid sequence of the IP140B antibody is as follows (SEQ ID NO:3): [ka]

[0114] The light chain amino acid sequence is as follows (SEQ ID NO:4): [ka]

[0115] Unless otherwise specified, the materials and equipment used in specific embodiments of the present invention are known products and can be obtained commercially.

[0116] Preparation Example 1: Preparation of Intermediate 1-5 JPEG2026501616000049.jpg122170

[0117] Synthesis of Intermediate 1 [ka] Intermediate 1 was synthesized according to the method described in the literature (WO2019195665A1). Fmoc-Gly-Gly-OH (5 g, 14.11 mol) and DMF (50 mL) were added to a 250 mL reaction flask and stirred to dissolve. Copper acetate (768 mg, 4.23 mmol), acetic acid (1.69 g, 28.22 mmol), and lead tetraacetate (6.872 g, 15.5 mmol) were added sequentially. The mixture was heated to 60°C and stirred for 20 minutes. The reaction mixture was poured into ice water, extracted with ethyl acetate, dried, and recrystallized to obtain an off-white solid product, Intermediate 1 (3.2 g, 66% yield). LCMS: (M+1) + 369.21 (theoretical value: 368.14).

[0118] Synthesis of Intermediate 2 [ka] Intermediate 2 was synthesized according to the method described in the literature (WO2019195665A1). Intermediate 1 (1 g, 2.7 mmol), DCM (10 mL), and TMSCl (8 mL) were added to a 100 mL three-necked flask, and the mixture was stirred at room temperature for 2 hours under argon gas protection. The mixture was then concentrated under reduced pressure to obtain Intermediate 2 (yield: approximately 100%) as a white solid, which was used directly in the next reaction without purification. LCMS: (M+1) + 344.80 (theoretical value: 344.09).

[0119] Synthesis of Intermediates 3a and 3b (1) Synthesis of intermediate 3a [ka] A 250 mL reaction flask was charged with Fmoc-NHCHCHNHCHCOOH (5 g, 14.7 mmol), DCM (70 mL), TEA (4.46 g, 44.07 mmol), and acetyl chloride (2.31 g, 29.38 mmol). The mixture was stirred at room temperature for 4 hours, and the solvent was removed under reduced pressure to give the solid product Fmoc-NHCHCHN(Ac)CHCOOH (5.62 g, 100% yield). LCMS: (M+1) + 383.16 (theoretical value: 382.15).

[0120] To a reaction flask was added Fmoc-NHCHCHN(Ac)CHCOOH (2 g, 5.23 mmol), THF (30 mL), Cu(OAc) (351.5 mg, 1.94 mmol), Pb(OAc) (3.48 g, 7.84 mmol), and acetic acid (691 mg, 11.51 mmol). The reaction was stirred at room temperature for 0.5 h, and the solvent was removed under reduced pressure. Recrystallization afforded the solid product intermediate 3a Fmoc-NHCHCHN(Ac)CHOAc (1.95 g, 94.1% yield). 1 H NMR(500MHz,DMSO-d6,ppm):δ7.89(d,J=7.5Hz,2H),7.68(t,J=6.3Hz,2H),7.42(t,J=7.5Hz,2H),7.34(t,J=7.5Hz,2H),5.31(d,J =18.7Hz,2H),4.34(dd,J=20.0,6.9Hz,2H),4.23(d,J=6.9Hz,1H),3.46-3.39(m,2H),3.12-3.20(m,2H),2.11(s,3H),2.04(s,3H); 13 CNMR(126MHz,DMSO-d6,ppm)δ171.67,170.74,156.65,144.37,141.22,128.08,127 .53,125.62,120.57,74.41,65.81,47.21,46.01,39.08,21.66,21.19;LCMS:(M+1) + 397.17 (calculated value: 396.17).

[0121] (2) Synthesis of intermediate 3b [ka] To a 250 mL reaction flask was added Fmoc-NHCHCHNHCHCOOH (2 g, 5.88 mmol), DCM (70 mL), TEA (1.78 g, 17.7 mmol), and MsCl (1.35 g, 11.8 mmol). The reaction was stirred at room temperature for 4 h, and the solvent was removed under reduced pressure to give the Fmoc-NHCHCHN(Ac)CHCOOH solid product (2.46 g, 100% yield). LCMS: (M+1) 419.12. + (Theoretical value: 418.12).

[0122] A reaction flask was charged with Fmoc-NHCHCHN(Ms)CHCOOH (2 g, 4.78 mmol), THF (30 mL), Cu(OAc) (321.5 mg, 1.77 mmol), Pb(OAc) (3.18 g, 7.17 mmol), and acetic acid (631 mg, 10.51 mmol), and the mixture was stirred at room temperature for 0.5 h. The solvent was then removed under reduced pressure to give solid 3b Fmoc-NHCHCHN(Ms)CHOAc (1.95 g, 94.3% yield). 1 H NMR(500MHz,DMSO-d6,ppm):δ7.89(d,J=7.5Hz,2H),7.69(d,J=7.5Hz,2H),7.42(t,J=7.6Hz,2H),7.33(t,J=7.1Hz,2H),5.30 (s,2H),4.32(d,J=6.7Hz,2H),4.23(d,J=6.8Hz,1H),3.32(t,J=6.1Hz,2H),3.23(t,J=6.1Hz,2H),3.02(s,3H),2.05(s,3H); 13 CNMR(126MHz,DMSO-d6,ppm)δ170.74,156.67,144.34,141.21,128.08,127 .54,125.61,120.57,72.53,65.90,47.17,46.31,39.97,36.24;LCMS:(M+1) + 433.14 (calculated value: 432.14).

[0123] Synthesis of Intermediates 4a and 4b (1) Synthesis of intermediate 4a [ka] N-(aminoethyl)phthalimide (0.6 g, 2.65 mmol), DCM (50 mL), DIEA (0.684 g, 5.29 mmol), and AcCl (0.25 g, 3.18 mmol) were added to a reaction flask and stirred at room temperature for 4 hours. Saturated brine (50 mL) was added, the DCM layer was separated, the aqueous layer was extracted twice with dichloromethane (50 mL x 2), the organic layers were combined, dried, and the solvent was removed under reduced pressure to give compound N-(aminoethyl)phthalimide (550 mg, 89.4% yield). LCMS: (M+1) + 233.02 (theoretical value: 232.08).

[0124] The above N-(acetylamidoethyl)phthalimide (0.55 g, 2.37 mmol), paraformaldehyde (95.4 mg, 3.18 mmol), DCM (12 mL), and TMSCl (3 mL) were added to a reaction flask, and the mixture was stirred at 50° C. for 12 hours to react. The solvent was removed under reduced pressure to give intermediate 4a (585 mg, 88% yield). 1 H NMR(600MHz,DMSO-d6,ppm):δ7.88-7.71(m,4H),4.76-4.51(m,2H),3.77-3.42(m,4H),1.91(s,3H);LCMS:(M+1) + 280.95 (theoretical value: 280.06).

[0125] (2) Synthesis of intermediate 4b [ka] N-(aminoethyl)phthalimide (0.6 g, 2.65 mmol), DCM (50 mL), DIEA (0.684 g, 5.29 mmol), and MsCl (0.364 g, 3.18 mmol) were added to a reaction flask and stirred at room temperature for 4 hours. Saturated brine (50 mL) was added, the DCM layer was separated, and the aqueous phase was extracted twice with methane dichloride (50 mL x 2). The combined organic phases were dried, and the solvent was removed under reduced pressure to give compound N-(methanesulfonylamidoethyl)phthalamide (700 mg, 98% yield). LCMS: (M+1)+ 269.05 (theoretical value: 268.05).

[0126] The above N-(methanesulfonylamidoethyl)phthalamide (0.71 g, 2.65 mmol), paraformaldehyde (95.4 mg, 3.18 mmol), DCM (12 mL), and TMSCl (3 mL) were added to a reaction flask, and the mixture was stirred at 50° C. for 12 hours. The solvent was removed under reduced pressure to give intermediate 4b (838.5 mg, 100% yield). 1 H NMR (600MHz, DMSO-d6, ppm): δ7.98-7.65(m,4H),4.85-4.54(m,2H),3.85-3.64(m,2H),3.50-3.30(m,2H),2.84(s,3H); 13 C NMR(150MHz,DMSO-d6,ppm)δ168.2,168.2,134.8,134.8,132.1,132.1,123.5,123.5,70.0,42.5,40.1,36.2;LCMS:(M+1) + 318.03 (theoretical value: 316.76).

[0127] Synthesis of Intermediates 5a-c (1) Synthesis of intermediate 5a [ka] Phthaloylglycine (5 g, 24.39 mmol) was added to a 500 mL single-neck flask and dissolved in DCM (175 mL). DMF (100 μL) was added. The reaction mixture was cooled to 0°C in an ice bath. Oxalyl chloride (6.15 g, 48.78 mmol) was added dropwise under argon gas protection. The reaction mixture was then stirred at 0°C for 1.5 hours, concentrated under reduced pressure, and the concentrate was dissolved in DCM (90 mL). The mixture was cooled to 0°C. 7 M ammonia methanol solution (10.45 mL, 73 mmol) was then added dropwise. The mixture was further stirred at room temperature for 2 hours. n-Hexane was added, the mixture was filtered, and the filter cake was dried to obtain phthaloylglycine amide (4.05 g, yield 81%). 1HNMR(600MHz,DMSO-d6):δ7.87-7.86(m,4H),7.84(s,1H),7.26(s,1H),4.17(s,2H); 13 CNMR(151MHz,DMSO-d6):δ168.54,168.05,162.96,134.98,132.15,45.79;LCMS:(M+1) + 204.87 (theoretical value: 204.05).

[0128] A reaction flask was charged with 5 mL of anhydrous DCM, compound 2-(phthaloylimino)acetamide (200 mg, 979.5 μmol), paraformaldehyde (470.15 mg, 3.92 mmol), and TMSCl (1.25 mL). The reaction mixture was stirred at 50°C for 2 h, and the solvent was removed under reduced pressure. The resulting mixture was purified by silica gel column chromatography to give intermediate 5a (R = H) (220 mg, 89% yield, 95% HPLC yield). 1 H NMR (500MHz, CDCl3, ppm): δ7.91-7.89(m,2H),7.77-7.75(m,2H),5.44(d,2H),4.73(s,2H); 13 CNMR(151MHz,DMSO-d6):δ170.20,167.84,134.86,132.25,123.52,48.57,15.24;LCMS:(M+1) + 253.15 (theoretical value: 252.15).

[0129] (2) Synthesis of intermediate 5b [ka] Phthaloyl valine (5 g, 20.22 mmol), dry DCM (175 mL), and DMF (100 μL) were added to a 500 mL single-neck flask. The reaction mixture was cooled to 0°C in an ice bath. Oxalyl chloride (5.13 g, 40.44 mmol) was added dropwise under argon gas protection. The mixture was stirred at 0°C for 1.5 hours. The solvent was removed under reduced pressure, and the concentrate was dissolved in DCM (90 mL). The mixture was cooled to 0°C. 7M ammonia in methanol (8.66 mL, 60.6 mmol) was added dropwise. The mixture was stirred at room temperature for 2 hours. n-Hexane was added, the mixture was filtered, and the filter cake was dried to give phthaloyl valine amide (2.98 g, 61% yield). LCMS: (M+1) + 247.02 (theoretical value: 246.10).

[0130] The product (1 g, 4.06 mmol) prepared by the above method was dissolved in DCM (10 mL), TMSCl (5 mL) and paraformaldehyde (550.4 mg, 18 mmol) were added, the tube was sealed, heated to 40 °C and reacted for 1.5 h, cooled to room temperature, filtered and concentrated to give 5b (1.19 g, 100% yield). 1 H NMR(600MHz,DMSO-d6,ppm):δ8.01-7.70(m,4H),4.75-4.42(m,2H),4.42-4.15(m,1H),2.82-2.60(m,1H),1.08-0.83(m,6H);LCMS:(M+1) + 294.97 (theoretical value: 294.08).

[0131] (3) Synthesis of intermediate 5c [ka] Phthaloylalanine (5 g, 22.8 mmol), DCM (175 mL), and DMF (100 μL) were added to a 500 mL single-neck flask. The reaction mixture was cooled to 0°C in an ice bath, and oxalyl chloride (5.79 g, 45.6 mmol) was added dropwise under argon gas protection. The mixture was stirred at 0°C for 1.5 hours. The solvent was removed under reduced pressure, and the concentrate was dissolved in DCM (90 mL). The mixture was cooled to 0°C, and 7 M ammonia methanol solution (9.78 mL, 68.5 mmol) was added dropwise. The mixture was further stirred at room temperature for 2 hours. n-Hexane was added, the mixture was filtered, and the filter cake was dried to obtain phthaloylalanine amide (4.5 g, 92% yield). 1 H NMR(600MHz,DMSO-d6,ppm):δ7.84(d,J=4.9Hz,3H),7.63(s,1H),7.45(s,1H),7.19(d,J=10.6Hz,1H),4.69(q,J=7.2Hz,1H),1.56(d,J=7.3Hz,3H); 13 CNMR(151MHz,DMSO-d6,ppm):δ171.23,167.96,134.81,132.25,123.45,48.58,15.42;LCMS:(M+1) + 219.05 (theoretical value: 218.07).

[0132] The phthaloylalaninamide (1 g, 4.5 mmol) prepared above was dissolved in DCM (10 mL), TMSCl (5 mL) and paraformaldehyde (550.4 mg, 18 mmol) were added, the tube was sealed, heated to 40 °C and reacted for 1.5 h, cooled to room temperature, filtered, and concentrated to give 5c (1.22 g, 100% yield). 1 H NMR(500MHz, CDCl3, ppm): δ7.85(dd,J=5.4,3.1Hz,2H),7.78-7.72(m,2H),7.19 (s,1H),5.23-5.13(m,2H),5.03-4.74(m,2H),1.69(d,J=7.3Hz,3H);LCMS:(M+1) + 266.98 (theoretical value: 266.05).

[0133] Synthesis of Intermediates 6a and 6b (1) Synthesis of intermediate 6a [ka] Trifluoroethylamine hydrochloride (100 mg, 0.74 mmol) and 7-methyl-10,11-methylenedioxycamptothecin (100 mg, 0.25 mmol) were dissolved in DMSO (3 mL), heated to 120°C with stirring, and reacted for 1 hour. After cooling, methyl tert-butyl ether was added, filtered, and purified by silica gel column chromatography to obtain the product 7-(N-2,2,2-trifluoroethyl)aminoethyl-10,11-methylenedioxycamptothecin (49 mg, yield 38%, HPLC 99%). 1 H NMR(500MHz,DMSO-d6)δ7.71(s,1H),7.54(s,1H),7.29(s,1H),6.55(s,1H),6.34(d,J=2.0Hz,2H),5.48(d,J=3.0Hz,2H) ),5.30(s,2H),3.42(s,2H),3.42(s,2H),3.29(s,2H),1.92(dd,J=14.4,7.3Hz,2H),0.94(t,J=7.3Hz,4H);LC-MS(M+H) + 518.34 (theoretical value 17.15).

[0134] (2) Synthesis of intermediate 6b [ka] Difluoroethylamine (1.0 g, 12 mmol), hydrochloric acid (1.4 mL, 16 mmol), 7-methyl-10,11-methylenedioxycamptothecin (1.0 g, 2.46 mmol), and DMSO (5 mL) were added to a reaction flask, heated to 120°C with stirring, and reacted for 1 hour. After cooling, isopropyl alcohol was added, and the mixture was suction filtered and purified by silica gel column chromatography to give 7-(N-2,2-difluoroethyl)aminoethyl-10,11-methylenedioxycamptothecan (393 mg, 32% yield, 95% HPLC yield). LC-MS (M+H) analysis revealed that the 7-(N-2,2-difluoroethyl)aminoethyl-10,11-methylenedioxycamptothecan (393 mg, 32% yield, 95% HPLC yield) was obtained. + 500.13 (theoretical value 499.16). 1H NMR(500MHz,DMSO-d6)δ7.68(s,1H),7.52(s,1H),7.28(s,1H),6.58(s,1H),6.33(s,2H),6.02(t,J=5 6.6Hz,1H),5.47(s,2H),5.26(s,2H),3.26(s,3H),2.94(s,4H),1.92(s,2H),0.93(s,3H);LC-MS(M+H) + 500.13 (theoretical value 499.16).

[0135] Example 1: 7-[N-(2,2-difluoroethyl)-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Gly-Phe-Gly-methylene)]aminoethyl-10,11-methylenedioxycamptothecan (1) [ka] 7-(N-2,2-difluoroethyl)aminoethyl-10,11-methylenedioxycamptothecin (payload 1,100 mg, 0.2 mmol), DCM (5 mL), triethylamine, and intermediate 2 (0.93 g, 2.7 mmol) were added to a reaction flask, and the reaction mixture was allowed to react at room temperature for 2 hours. Water (10 mL) was added, and the mixture was extracted with DCM (20 mL x 3). The organic phase was concentrated and purified by silica gel column chromatography to give compound 10a (121 mg, 75%). LCMS: (M+1) + 808.13 (theoretical value: 807.28).

[0136] To a reaction flask was added 1a (121 mg, 0.149 mg), DMF (1 mL), and piperidine (100 μL). The reaction mixture was stirred at room temperature for 1 hour, methyl tert-butyl ether (20 mL) was added, the mixture was centrifuged, the supernatant was removed, and the mixture was dried to give 1b (68 mg, 78% yield). LCMS: (M+1) + 586.21 (theoretical value: 585.20).

[0137] To a reaction flask were added 1b (68 mg, 0.11 mmol), DMF (2 mL), and DIPEA (15 mg, 0.11 mmol). The reaction mixture was stirred at room temperature, and then Fmoc-Gly-Gly-Phe-OH (56 mg, 0.11 mmol) and HATU (42 mg, 0.11 mmol) were added sequentially. The reaction mixture was left overnight at room temperature, and the solvent was removed under reduced pressure. The mixture was purified by silica gel column chromatography to give product 1c (107 mg, 86% yield). LCMS: (M+1) + 1069.09 (theoretical value: 1068.38).

[0138] To a reaction flask was added 1c (107 mg, 0.1 mmol), DMF (1 mL), and piperidine (100 μL). The reaction mixture was stirred at room temperature for 1 hour, methyl tert-butyl ether (20 mL) was added, the mixture was centrifuged, the supernatant was removed, and the mixture was dried to give 1d (56 mg, 66 percent yield). LCMS: (M+1) + 847.23 (theoretical value: 846.31).

[0139] To a reaction flask was added DCM (3 mL), 1d (56 mg, 0.066 mmol), bromoacetylaminoethoxyethoxypropionic acid (19.2 mg, 0.066 mmol), and DIC (8.4 mg, 0.066 mmol). The reaction was stirred at room temperature for 90 min, concentrated, and purified by silica gel column chromatography to give a yellow solid 1 (12 mg, 16% yield). LCMS: (M+1) + 1126.24 (theoretical value: 1125.33).

[0140] ADC-1a: An antibody conjugate consisting of Compound 1 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken, and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0141] Compound 1 (0.90 mg, 0.8 mmol) was dissolved in 0.09 ml of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM L-histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva), to obtain ADC-1a (3.4 mg / ml, 2 ml). Average value calculated by UV-HPLC: n=7.5

[0142] ADC-1b: an antibody conjugate consisting of Compound 1 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0143] Compound 1 (0.90 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-1b (3.1 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0144] Example 2: 7-[N-(2,2,2-trifluoroethyl)-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Gly-Phe-Gly-methylene)]aminoethyl-10,11-methylenedioxycamptothecan (2) [ka] 7-(N-2,2,2-trifluoroethyl)aminoethyl-10,11-methylenedioxycamptothecin (payload 2,100 mg, 0.19 mmol), DCM (5 mL), and intermediate 2 (665.3 mg, 1.9 mmol) were added to a reaction flask, and the reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was concentrated and purified by silica gel column chromatography to give compound 2a (47 mg, 30% yield). LCMS: (M+1) + 826.19 (theoretical value: 825.26).

[0145] 2a (47 mg, 0.056 mmol) was added to a reaction flask, DMF (1 mL) and piperidine (100 μL) were added, and the mixture was stirred at room temperature for 1 hour. Methyl tert-butyl ether (20 mL) was added, the mixture was centrifuged, the supernatant was removed, and the mixture was dried to give 2b (21 mg, 60% yield). LCMS: (M+1) + 604.03 (theoretical value: 603.19).

[0146] To a reaction flask, 2b (21 mg, 0.034 mmol), DMF (2 mL), and DIPEA (4.49 mg, 0.034 mmol) were added. The reaction mixture was stirred at room temperature until clear. Fmoc-aminoethoxyethoxypropionyl-Gly-Leu (19.8 mg, 0.034 mmol) and HATU (13.23 mg, 0.034 mmol) were added sequentially. The reaction mixture was stirred at room temperature for 1 hour, concentrated, and purified by silica gel column chromatography to give compound 2c (35 mg, 87.5% yield). LCMS: (M+1) + 1155.22 (theoretical value: 1154.46).

[0147] 2c (35 mg, 0.03 mmol) was added to a reaction flask, DMF (1 mL) and piperidine (100 μL) were added, and the mixture was stirred at room temperature for 1 hour. Methyl tert-butyl ether (20 mL) was added, the mixture was centrifuged, the supernatant was removed, and the mixture was dried to give 2d (22 mg, 78.9% yield). LCMS: (M+1) + 933.65 (theoretical value: 932.95).

[0148] To the reaction flask was added DCM (3 mL), 2d (22 mg, 0.023 mmol), bromoacetic acid (7 mg, 0.023 mmol), and DIC (2.97 mg, 0.023 mmol). The reaction was stirred at room temperature for 90 min, concentrated, and purified by silica gel column chromatography to give a yellow solid 2 (19 mg, 77% yield). LCMS: (M+1) + 1053.27 (theoretical value: 1052.31).

[0149] ADC-2a: An antibody conjugate consisting of Compound 2 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0150] Compound 2 (0.84 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM L-histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20 using a NAP-5 gel column (Cytiva) to obtain ADC-2a (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.1

[0151] ADC-2b: an antibody conjugate drug consisting of Compound 2 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0152] Compound 2 (0.84 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-2b (3.1 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.0

[0153] Example 3: 7-[N-(2,2,2-trifluoroethyl)-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Gly-Phe-Gly-methylene)]aminoethyl-10,11-methylenedioxycamptothecan (3) [ka] A reaction flask was charged with 2b (70 mg, 0.11 mmol), DMF (2 mL), HATU (41.8 mg, 0.11 mmol), DIPEA (14.19 mg, 0.11 mmol), and Fmoc-Gly-Gly-Phe-OH (55.11 mg, 0.11 mmol). The reaction mixture was stirred at room temperature for 6 h, and then DCM (200 mL) was added. The mixture was washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to give a yellow solid, 3a (108 mg, 85% yield).

[0154] A reaction flask was charged with 3a (108 mg, 0.099 mmol), DMF (1 mL), and piperidine (77 mg, 0.99 mmol), and the mixture was stirred at room temperature for 1 hour. Methyl tert-butyl ether (30 mL) was added, the mixture was centrifuged, the supernatant was removed, and the solvent was removed under reduced pressure. The resulting solid 3b (43 mg, 50% yield) was used directly in the next reaction. LCMS: (M+1) + 865.29 (theoretical value: 864.31).

[0155] DCM (2 mL), 3b (43 mg, 0.049 mmol), bromoacetamidoethoxyethoxypropionic acid (15 mg, 0.049 mmol), and DIC (6.27 mg, 0.049 mmol) were added to the reaction flask sequentially and stirred at room temperature for 90 min. The reaction mixture was then loaded onto a 25 g C18 precolumn (equilibrated first with acetonitrile and then with water, containing 0.1% TFA in the aqueous phase). The mixture was then eluted by medium-pressure reverse-phase C18 chromatography (gradient: 5% to 40% acetonitrile in water, 30 min), and lyophilized to give a yellow solid 3 (21 mg, 37.5% yield). LCMS: (M+1) + 1144.28 (theoretical value: 1143.32).

[0156] ADC-3a: An antibody conjugate consisting of Compound 3 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0157] Compound 3 (0.92 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva) to obtain ADC-3a (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=6.2

[0158] ADC-3b: an antibody conjugate consisting of Compound 3 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0159] Compound 3 (0.92 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-3b (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=6.5

[0160] Example 4: 7-[N-(2,2,2-trifluoroethyl)-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Val-Ala-Gly-methylene)]aminoethyl-10,11-methylenedioxycamptothecan (4) [ka] A reaction flask was charged with 2b (70 mg, 0.11 mmol), DMF (2 mL), HATU (41.8 mg, 0.11 mmol), DIPEA (14.19 mg, 0.11 mmol), and Fmoc-Gly-Val-Ala-OH (51.37 mg, 0.11 mmol). The mixture was stirred at room temperature for 6 hours, and then DCM (200 mL) was added. The mixture was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to give a yellow solid 13a (100.1 mg, 87% yield). LCMS: (M+1) + 1053.06 (theoretical value: 1052.39).

[0161] A reaction flask was charged with 4a (100.1 mg, 0.095 mmol), DMF (0.5 mL), and piperidine (74 mg, 0.95 mmol), and the mixture was stirred at room temperature for 1 hour. Methyl tert-butyl ether (30 mL) was added, the mixture was centrifuged, the supernatant was removed, and the solvent was removed under reduced pressure. The resulting solid 4b (49 mg, 62% yield) was used directly in the next reaction. LCMS: (M+1) + 831.27 (theoretical value: 830.32).

[0162] DCM (2 mL), 4b (49 mg, 0.059 mmol), bromoacetamidoethoxyethoxypropionic acid (17.6 mg, 0.059 mmol), and DIC (7.43 mg, 0.059 mmol) were added to the reaction flask sequentially and stirred at room temperature for 90 min. The reaction mixture was then loaded onto a 25 g C18 precolumn (equilibrated first with acetonitrile and then with water, containing 0.1% TFA in the aqueous phase). The mixture was then eluted by medium-pressure reverse-phase C18 chromatography (gradient: 5% to 40% acetonitrile in water, 30 min), and lyophilized to give a yellow solid 4 (15 mg, 22% yield). LCMS: (M+1). + 1110.28 (theoretical value: 1109.33).

[0163] ADC-4a: An antibody conjugate consisting of Compound 4 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0164] Compound 4 (0.89 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM L-histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20 using a NAP-5 gel column (Cytiva) to obtain ADC-4a (3.1 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=6.3

[0165] ADC-4b: an antibody conjugate consisting of Compound 4 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0166] Compound 4 (0.89 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotating turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-4b (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=6.6

[0167] Example 5: Bromoacetamidoethoxyethoxypropionyl-Gly-Leu-ethylenediamine(Ac)-methyleneoxyacetylexatecan (5) [ka] A reaction flask was charged with intermediate 3a (1.95 g, 4.92 mmol), DCM (25 mL), benzyl hydroxyacetate (8.17 g, 49.2 mmol), and PPTS (123.6 mg, 0.492 mmol). The reaction mixture was heated and refluxed with stirring overnight, after which 200 mL of EtOAc was added, washed with brine (3×200 mL), dried over anhydrous MgSO, filtered, concentrated in vacuo, and purified by silica gel column chromatography to give 5a (2.43 g, 98.3% yield) as a white powder. LCMS: (M+1). + 503.21 (theoretical value: 502.21).

[0168] 5a (200 mg, 0.398 mmol) and DBU (30.3 mg, 0.199 mmol) were added sequentially to the reaction flask and stirred at room temperature for 1 hour. 5b was then directly used in the next reaction. PPTS (50 mg, 0.298 mmol), DIPEA (60.9 mg, 0.471 mmol), PEG2-Gly-Leu (224 mg, 0.392 mmol), and HATU (298.4 mg, 0.785 mmol) were added to the reaction mixture of 5b and stirred at room temperature for 2 hours. The solvent was removed under reduced pressure to give 5c (236.8 mg, 72.5% yield). LCMS: (M+1) + 832.41 (theoretical value: 831.41).

[0169] Compound 5c (0.3 g, 0.36 mmol) obtained above was dissolved in MeOH (5 mL), palladium-charcoal catalyst (90 mg) was added, and the mixture was stirred at room temperature. Triethylsilane (0.42 g, 3.61 mmol) was then added, and the mixture was stirred at room temperature for 0.5 hours. The mixture was then filtered through diatomaceous earth to remove insoluble matter, and the solvent in the filtrate was removed under reduced pressure to give solid 5d (267.5 mg, 100% yield). LCMS: (M+1) + 742.36 (theoretical value: 741.36).

[0170] Exatecan (100 mg, 0.188 mmol) was dissolved in DMF (2 mL), and then DIPEA (48.6 mg, 0.376 mmol), 5d (167.5 mg, 0.226 mmol), and HATU (143 mg, 0.376 mmol) were added sequentially. The reaction was stirred at room temperature for 2 hours, and the solvent was removed under reduced pressure to give 5e (196.4 mg, 90.1% yield) as a solid. LCMS: (M+1) + 1159.51 (theoretical value: 1158.51).

[0171] To the reaction flask, 5e (96.4 mg, 0.17 mmol) and piperidine (144.4 mg, 1.7 mmol) were added sequentially, and the mixture was stirred at room temperature for 0.5 hours. Methyl tert-butyl ether (30 mL) was added, the mixture was centrifuged, the supernatant was removed, and the solvent was removed under reduced pressure. The resulting solid 5f (135.6 mg, 85.4% yield) was used directly in the next reaction. LCMS: (M+1) + 937.44 (theoretical value: 936.44).

[0172] DCM (2 mL), 5f (100 mg, 0.049 mmol), bromoacetic acid (17.8 mg, 0.049 mmol), and DIC (16.2 mg, 0.049 mmol) were added to the reaction flask in this order and stirred at room temperature for 90 min. The reaction mixture was then loaded onto a 25 g C18 precolumn (first equilibrated with acetonitrile, then with water, containing 0.1% TFA in the aqueous phase). The mixture was then eluted by medium-pressure reverse-phase C18 chromatography (gradient: 5% to 40% acetonitrile in water, 30 min), and lyophilized to give a yellow solid 5 (30.1 mg, 26.6% yield). LCMS: (M+1) + 1058.26 (theoretical value: 1056.36).

[0173] ADC-5a: An antibody conjugate consisting of Compound 5 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken, and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0174] Compound 5 (0.85 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva) to obtain ADC-5a (3.1 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=6.6

[0175] ADC-5b: an antibody conjugate consisting of Compound 5 and the 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0176] Compound 5 (0.85 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-5b (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=6.5

[0177] Example 6: Bromoacetamidoethoxyethoxypropionyl-Gly-Leu-ethylenediamine(Ms)-methyleneoxyacetylexatecan (6) [ka] A reaction flask was charged with intermediate 3b (0.5 g, 1.16 mmol), DCM (5 mL), benzyl hydroxyacetate (8.17 g, 49.2 mmol), and PPTS (29.1 mg, 0.116 mmol). The reaction was heated and refluxed with stirring overnight, cooled to room temperature, added to EtOAc (200 mL), washed with water (3 x 200 mL), dried over anhydrous MgSO, filtered, concentrated in vacuo, and purified by silica gel column chromatography to give 6a (536.8 mg, 86.2% yield) as a white powder. LCMS: (M+1). + 539.18 (theoretical value: 538.18).

[0178] 6a (200 mg, 0.371 mmol) and DBU (28.3 mg, 0.186 mmol) were added to a reaction flask, and the mixture was stirred at room temperature for 1 hour to give 6b. PPTS (31.2 mg, 0.186 mmol), DIPEA (48 mg, 0.371 mmol), PEG2-Gly-Leu (179.8 mg, 0.316 mmol), and HATU (141.2 mg, 0.371 mmol) were added and the mixture was stirred at room temperature for 2 hours. The solvent was removed under reduced pressure to give 6c (215.6 mg, 66.9% yield). LCMS: (M+1) + 868.37 (theoretical value: 867.37).

[0179] Compound 6c (0.15 g, 0.173 mmol) obtained above was dissolved in MeOH (5 mL), palladium-charcoal catalyst (50 mg) was added, and the mixture was stirred at room temperature. Triethylsilane (0.2 g, 1.73 mmol) was then added, and the mixture was stirred at room temperature for 0.5 hours. The mixture was then filtered through diatomaceous earth to remove insoluble matter, and the solvent in the filtrate was removed under reduced pressure to give solid 6d (134.43 mg, 100% yield). LCMS: (M+1) + 778.33 (theoretical value: 777.33).

[0180] Exatecan (110 mg, 0.207 mmol) was dissolved in DMF (2 mL), DIPEA (80.2 mg, 0.62 mmol), 6d (161 mg, 0.207 mmol), and HATU (157.4 mg, 0.414 mmol) were added sequentially. The mixture was stirred at room temperature for 2 hours, and the solvent was removed under reduced pressure to give 6e (213.5 mg, 86.3% yield). LCMS: (M+1) + 1195.33 (theoretical value: 1194.47).

[0181] To the reaction flask, 6e (140 mg, 0.117 mmol) and piperidine (99.7 mg, 1.17 mmol) were added sequentially, and the mixture was stirred at room temperature for 0.5 hours. Methyl tert-butyl ether (30 mL) was added, the mixture was centrifuged, the supernatant was removed, and the solvent was removed under reduced pressure. The solid 6f (80 mg, 70.2% yield) was used directly in the next reaction. LCMS: (M+1) + 973.41 (theoretical value: 972.41).

[0182] DCM (2 mL), 6f (80 mg, 0.082 mmol), bromoacetic acid (13.7 mg, 0.099 mmol), and DIC (12.5 mg, 0.099 mmol) were added to the reaction flask in this order and stirred at room temperature for 90 min. 0.1 mL of TFA was added and the mixture was allowed to react for another 30 min. Finally, the reaction mixture was loaded onto a 25 g C18 precolumn (first equilibrated with acetonitrile, then with water, containing 0.1% TFA in the aqueous phase). The mixture was then eluted by medium-pressure reverse-phase C18 chromatography (gradient: 5% to 40% acetonitrile in water, 30 min), and lyophilized to give a yellow solid 6 (30.1 mg, 26.6% yield). LCMS: (M+1). + 1094.33 (theoretical value: 1092.33).

[0183] ADC-6a: An antibody conjugate consisting of Compound 6 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0184] Compound 6 (0.87 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva) to obtain ADC-6a (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=6.3

[0185] ADC-6b: an antibody conjugate consisting of compound 6 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0186] Compound 6 (0.87 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-6b (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=6.6

[0187] Example 7: 7-[N-(2,2,2-trifluoroethyl)-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Phe-Gly-methylene)]aminoethyl-10,11-methylenedioxycamptothecan (7) [ka] Anhydrous DCM (2 mL), 7-(N-2,2,2-trifluoroethyl)aminoethyl-10,11-methylenedioxycamptothecin (100 mg, 162.5 μmol), intermediate 5a (61.58 mg, 243.75 μmol), and DIPEA (42.01 mg, 325 μmol) were added to a reaction flask and stirred at 40° C. for 12 hours. The solvent was removed under reduced pressure, and the residue was purified by silica gel column chromatography to give compound 7a (100 mg, 83.9% yield). LCMS: (M+1) +734.41 (theoretical value: 733.41).

[0188] Anhydrous methanol (2 mL) was added to a reaction flask, followed by the addition of intermediate 7a (100 mg, 136.3 μmol) and hydrazine hydrate (40.94 mg, 817.82 μmol). The reaction mixture was stirred at 60°C for 12 hours. Saturated brine (10 mL) was added to the reaction mixture, which was then extracted four times with DCM / i-PrOH = 4:1. The mixture was dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give compound 2b (70 mg, 85% yield, 94% HPLC yield). LCMS: (M+1) + 604.16 (theoretical value: 603.16).

[0189] To a single-neck flask containing anhydrous DMF (1.5 mL), FmocPEG2-Gly-PheOH (70.01 mg, 115.98 μmol), HATU (66.15 mg, 173.97 μmol), and TEA (22.48 mg, 173.97 μmol) were added and stirred at room temperature for 5 minutes. Compound 2b (70 mg, 115.98 μmol) was added and stirred at room temperature for 2 hours. Saturated brine (10 mL) was added to the reaction mixture, which was extracted four times with DCM / i-PrOH = 4:1, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure to give compound 7c (90 mg, 65.3% yield, 94% HPLC yield). LCMS: (M+1) + 1189.42 (theoretical value: 1188.42).

[0190] Compound 7c (90 mg, 75.68 μmol), anhydrous DCM (1.5 mL), and piperidine (64.44 mg, 756.8 μmol) were added to a reaction flask and stirred at room temperature for 30 minutes. Methyl tert-butyl ether was added to the reaction mixture, and the solid was centrifuged twice. The resulting white precipitate was dried to give compound 7d (45 mg, 61.5% yield, 94% HPLC yield). LCMS: (M+1) + 967.23 (theoretical value: 966.23).

[0191] Bromoacetic acid (9.7 mg, 69.81 μmol) and DIC (8.81 mg, 69.81 μmol) were added to a flask containing anhydrous DCM (1.5 mL) and stirred at room temperature for 5 minutes. Compound 7d (45 mg, 46.54 μmol) was added and stirred at room temperature for an additional hour. The reaction was continued and purified by silica gel column chromatography to give 7 (27 mg, 53.3% yield, 97% HPLC). LCMS: (M+1) + 1087.35 (theoretical value: 1086.35).

[0192] ADC-7a: An antibody conjugate consisting of Compound 7 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0193] Compound 7 (0.87 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva) to obtain ADC-7a (3.1 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.5

[0194] ADC-7b: an antibody conjugate consisting of compound 7 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0195] Compound 7 (0.87 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-7b (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0196] Example 8: 7-[N-(2,2,2-trifluoroethyl)-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Gly-Val-methylene)]aminoethyl-10,11-methylenedioxycamptothecan (8) [ka] Anhydrous DCM (2 mL), 7-(N-2,2,2-trifluoroethyl)aminoethyl-10,11-methylenedioxycamptothecin (90 mg, 146.2 μmol), intermediate 5b (64.64 mg, 219.3 μmol), and DIPEA (37.79 mg, 292.4 μmol) were added to a reaction flask, and the reaction mixture was stirred at 40° C. for 12 hours. The reaction mixture was concentrated, and the solvent was removed under reduced pressure. The mixture was purified by silica gel column chromatography to give compound 8a (100 mg, 88.2% yield, 95% HPLC yield). LCMS: (M+1) + 776.25 (theoretical value: 775.25).

[0197] Anhydrous methanol (2 mL) was added to a reaction flask, and compound 8a (100 mg, 128.91 μmol) and hydrazine hydrate (38.72 mg, 773.46 μmol) were added. The reaction mixture was stirred at 60°C for 12 hours. Saturated brine (10 mL) was added to the reaction mixture, which was extracted four times with DCM / i-PrOH = 4:1, dried over anhydrous sodium sulfate, filtered, and the solvent was concentrated to give compound 8b (75 mg, 90% yield, HPLC 94%). LCMS: (M+1) + 646.3 (theoretical value: 645.3).

[0198] Anhydrous DMF (1.5 mL), FmocPEG2-GLy-Gly-OH (59.65 mg, 116.16 μmol), HATU (66.26 mg, 174.24 μmol), and TEA (17.63 mg, 174.24 μmol) were added to a reaction flask, and the reaction mixture was stirred at room temperature for 5 minutes. Compound 8b (75 mg, 116.16 μmol) was added, and the mixture was allowed to react at room temperature for 2 hours. Saturated brine (10 mL) was added to the reaction mixture, and the mixture was extracted four times with DCM / i-PrOH = 4:1, dried over anhydrous sodium sulfate, filtered, and the solvent was concentrated to give compound 8c (82 mg, 61.8% yield, 96% HPLC yield). LCMS: (M+1) + 1141.41 (theoretical value: 1140.41).

[0199] Compound 8c (82 mg, 71.85 μmol), anhydrous DCM (1.5 mL), and piperidine (61.18 mg, 718.5 μmol) were added to a reaction flask, and the reaction mixture was stirred at room temperature for 30 minutes. Methyl tert-butyl ether was added to the reaction mixture, and the solid was centrifuged twice. The resulting white precipitate was dried to give compound 8d (52 mg, 86.1% yield, HPLC 94%). LCMS: (M+1) + 919.2 (theoretical value: 918.2).

[0200] Anhydrous DCM (1.5 mL), bromoacetic acid (11.79 mg, 84.88 μmol), and DIC (10.71 mg, 84.88 μmol) were added to a reaction flask, and the reaction mixture was stirred at room temperature for 5 minutes. Compound 8d (52 mg, 56.58 μmol) was added, and the reaction mixture was allowed to react at room temperature for an additional hour. The solvent was concentrated under reduced pressure, and the mixture was purified by silica gel column chromatography to give 8 (42 mg, 71.4% yield, 98% HPLC). LCMS: (M+1) + 1039.23

[0201] ADC-8a: An antibody conjugate consisting of Compound 8 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0202] Compound 8 (0.83 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva) to obtain ADC-8a (3.1 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0203] ADC-8b: an antibody conjugate consisting of compound 8 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0204] Compound 8 (0.83 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-8b (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0205] Example 9: 7-[N-(2,2,2-trifluoroethyl)-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Val-Ala-methylene)]aminoethyl-10,11-methylenedioxycamptothecan (9) [ka] Anhydrous DCM (2 mL), 7-(N-2,2,2-trifluoroethyl)aminoethyl-10,11-methylenedioxycamptothecin (90 mg, 146.2 μmol), intermediate 5c (61.34 mg, 219.3 μmol), and DIPEA (37.79 mg, 292.4 μmol) were added to a reaction flask, and the reaction mixture was stirred at 40° C. for 12 hours. The solvent was removed under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give compound 9a (95 mg, 89.2% yield, 95% HPLC yield). LCMS: (M+1) + 748.22 (theoretical value: 747.22).

[0206] Compound 9a (95 mg, 128.91 μmol) and hydrazine hydrate (38.72 mg, 773.46 μmol) were added to a single-neck flask containing anhydrous methanol (2 mL). The reaction mixture was stirred at 60° C. for 12 hours. Saturated brine (10 mL) was added to the reaction mixture, which was then extracted four times with DCM / i-PrOH (4:1). The mixture was dried over anhydrous sodium sulfate, filtered, and concentrated to give compound 9b (75 mg, 88.5% yield, 93% HPLC yield). LCMS: (M+1) + 618.56 (theoretical value: 617.56).

[0207] To a single-neck flask containing anhydrous DMF (1.5 mL), FmocPEG2-GLy-Gly-OH (59.65 mg, 116.16 μmol), HATU (66.26 mg, 174.24 μmol), and TEA (17.63 mg, 174.24 μmol) were added. The reaction mixture was stirred at room temperature for 5 minutes, and compound 9b (75 mg, 116.16 μmol) was added. The mixture was allowed to react at room temperature for 2 hours. Saturated brine (10 mL) was added to the reaction mixture, which was then extracted four times with DCM / i-PrOH (4:1), dried over anhydrous sodium sulfate, and concentrated to give compound 9c (82 mg, 78.6% yield, 94% HPLC yield). LCMS: (M+1) + 1155.43 (theoretical value: 1154.43).

[0208] Compound 9c (82 mg, 71.85 μmol), anhydrous DCM (1.5 mL), and piperidine (61.18 mg, 718.5 μmol) were added to a reaction flask and stirred at room temperature for 30 minutes. Methyl tert-butyl ether was added to produce a white precipitate. The solid was centrifuged twice, and the white precipitate was dried to give compound 9d (52 mg, 85.6% yield, HPLC 95%). LCMS: (M+1) + 933.35 (theoretical value: 932.35).

[0209] Bromoacetic acid (11.79 mg, 84.88 μmol) and DIC (10.71 mg, 84.88 μmol) were added to a single-neck flask containing anhydrous DCM (1.5 mL) and stirred at room temperature for 5 minutes. Compound 9d (52 mg, 56.58 μmol) was added and the reaction was continued with stirring at room temperature for another hour. The reaction was then purified by silica gel column chromatography to give 9 (50 mg, 56.58 μmol).

[0210] ADC-9a: An antibody conjugate consisting of Compound 9 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0211] Compound 9 (0.84 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva) to obtain ADC-9a (3.1 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.7

[0212] ADC-9b: an antibody conjugate consisting of compound 9 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0213] Compound 9 (0.84 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-9b (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0214] Example 10: 10-(Bromoacetamidoethoxyethoxypropionyl-Gly-Leu-ethylenediamine(Ms)-methylene)oxy-7-ethylcamptothecin (10) [ka] 7-Ethyl-10-hydroxycamptothecin (0.1 g, 0.255 mmol), DCM (5 mL), DIEA (65.9 mg, 0.51 mmol), and intermediate 4 (84.8 mg, 0.268 mmol) were added to a reaction flask, and the reaction mixture was stirred at 40° C. for 1 hour. The solvent was removed under reduced pressure, and the mixture was purified by silica gel column chromatography to give compound 10a (145.1 mg, 84.6% yield). LCMS: (M+1) + 673.71 (theoretical value: 672.71)

[0215] Compound 10a (0.1 g, 0.149 mmol), MeOH (5 mL), and 80% hydrazine hydrate (55.8 mg, 0.892 mmol) were added to a reaction flask, and the reaction mixture was stirred at 60 °C for 2 hours. 20 mL of saturated brine was added, and the mixture was extracted three times with dichloromethane (10 mL × 3). The combined organic phases were dried, and the solvent was removed under reduced pressure to give compound 10b (73.2 mg, 90.8% yield). LCMS: (M+1) + 543.61 (theoretical value: 542.61).

[0216] Compound 10b (100 mg, 0.184 mmol) obtained above was dissolved in DMF (2 mL), and then DIPEA (47.6 mg, 0.369 mmol), Fmoc-PEG2-GLy-Leu-OH (105 mg, 0.184 mmol), and HATU (70.1 mg, 0.184 mmol) were added sequentially. The reaction mixture was stirred at room temperature for 2 hours, the solvent was removed under reduced pressure, and the mixture was purified by silica gel column chromatography to give compound 10c (184.5 mg, 91.5% yield). LCMS: (M+1) + 1195.25 (theoretical value: 1194.25).

[0217] Compound 10c (100 mg, 0.091 mmol) and piperidine (77.8 mg, 0.914 mmol) were added to the reaction flask, and the reaction mixture was stirred at room temperature for 0.5 hours. Methyl tert-butyl ether (30 mL) was added, and the mixture was centrifuged. The supernatant was removed. This was repeated three times to give compound 10d (80 mg, 100% yield). LCMS: (M+1) + 873.01 (theoretical value: 872.01).

[0218] DCM (2 mL), compound 10d (70 mg, 0.08 mmol), bromoacetic acid (22.3 mg, 0.16 mmol), and DIC (20.3 mg, 0.16 mmol) were added to a reaction flask in this order, and the mixture was stirred at room temperature for 90 minutes. The solvent was removed under reduced pressure, and the residue was purified by silica gel column chromatography to give a pale yellow solid 10 (65.3 mg, 81.9% yield). LCMS: (M+1) + 994.33 (theoretical value: 992.94).

[0219] ADC-10a: An antibody conjugate consisting of Compound 10 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0220] Compound 10 (0.79 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20 using a NAP-5 gel column (Cytiva) to obtain ADC-10a (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.1

[0221] ADC-10b: an antibody conjugate consisting of compound 10 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0222] Compound 10 (0.79 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-10b (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.2

[0223] Example 11: 2-[N-methyl-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Leu-Gly-methylene)]aminooxyacetylexatecan (11) [ka] Boc-N-methylhydroxylamine hydrochloride (15 g, 101.9 mmol) was dissolved in isopropyl alcohol (100 mL), bromoacetic acid benzyl ester (27.709 g, 121.4 mmol) and DIPEA (15.7 g, 121.7 mmol) were added, and the mixture was heated to 85 °C with stirring under argon gas protection for 3 hours. The reaction mixture was concentrated, dissolved in ethyl acetate (100 mL), washed with saturated brine (30 mL x 3), dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to give Intermediate 11a (24 g, 80% yield, 98% HPLC yield). LCMS: (M+1) + 296.08 (theoretical value: 295.14).

[0224] Dichloromethane (20 mL) and 11a (2 g, 6.8 mmol) were added to a 100 mL single-neck flask and stirred until uniform. 4N hydrochloric acid in ethyl acetate (10 mL) was added dropwise, and the mixture was stirred overnight at room temperature. The mixture was then concentrated to give 11b (1.32 g, 100% yield, 96% HPLC yield). LCMS: (M+1) + 195.99 (theoretical value: 195.06).

[0225] 5a (1 g, 3.9 mmol) was dissolved in DCM (5 mL), 11b (0.77 g, 3.9 mmol) and DIPEA (290.79 mg, 2.24 mmol) were added, the tube was sealed, and the mixture was heated to 50 °C and reacted overnight. The mixture was then cooled to room temperature, concentrated, and purified by silica gel column chromatography to give 11c (1.2 g, 75% yield, 91% HPLC yield). LCMS: (M+1) + 412.06 (theoretical value: 411.14).

[0226] 11c (1.2 g, 2.9 mmol) was added to a 100 mL single-neck flask, and methanol (5 mL) and 10% palladium on carbon (0.1 g) were added. The mixture was purged with hydrogen gas and reacted at room temperature and atmospheric pressure for 2 hours. The mixture was filtered and concentrated to give 11d (0.91 g, 97% yield, HPLC 93%). LCMS: (M+1) + 322.01 (theoretical value: 321.10).

[0227] A reaction flask was charged with exatecan sulfonate (0.26 g, 0.5 mmol), DMF (2 mL), HATU (0.19 g, 0.5 mmol), DIPEA (0.129 g, 1 mmol), and 11d (0.176 g, 0.55 mmol). The mixture was stirred at room temperature for 15 hours, and then DCM (200 mL) was added. The mixture was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to give a yellow solid 11e (0.46 g, 66% yield, 92% HPLC yield). LCMS: (M+1) + 739.11 (theoretical value: 738.24).

[0228] 11e (0.1 g, 0.12 mmol) was dissolved in MeOH (5 mL), hydrazine hydrate (66 mg, 1.32 mmol) was added, the tube was sealed, and the mixture was heated to 60 °C and reacted for 2 hours. After cooling to room temperature, the reaction mixture was poured into saturated brine (100 mL), extracted with ethyl acetate (200 mL), and concentrated to give 11f (0.06 g, 72% yield, HPLC 94%). LCMS: (M+1) + 609.14 (theoretical value: 608.24).

[0229] 11f (60 mg, 0.098 mmol) was dissolved in DMF (2 mL), DIPEA (25 mg, 0.197 mmol) was added, and Fmoc-PEG2-Gly-Leu-OH (55.76 mg, 0.098 mmol) and HATU (37.5 mg, 0.098 mmol) were added sequentially with stirring. The mixture was stirred at room temperature for 1 hour, concentrated, and purified by silica gel column chromatography to give compound 11g (85 mg, 75% yield, 91% HPLC yield). LCMS: (M+1) + 1160.27 (theoretical value: 1159.50).

[0230] 11g (85 mg, 0.069 mmol) was placed in a 10 mL single-neck flask, dissolved in 1 mL of DMF, and then piperidine (100 μL) was added. The mixture was allowed to react at room temperature for 1 hour, and then methyl tert-butyl ether (20 mL) was added. The mixture was centrifuged, the supernatant was removed, and the mixture was dried to give 11h (31 mg, 46% yield, 93% HPLC yield). LCMS: (M+1) + 938.02 (theoretical value: 937.43).

[0231] A reaction flask was charged with DCM (3 mL), 11h (31 mg, 0.032 mmol), bromoacetic acid (8.9 mg, 0.064 mmol), and DIC (8.01 mg, 0.064 mmol). The mixture was stirred at room temperature for 90 minutes, concentrated, and purified by silica gel column chromatography to give a yellow solid 11 (25 mg, 63% yield, 95% HPLC). LCMS: (M+1) + 1058.24 (theoretical value: 1057.36).

[0232] ADC-11a: An antibody conjugate consisting of Compound 11 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0233] Compound 11 (0.85 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM L-histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva), to obtain ADC-11a (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.5

[0234] ADC-11b: an antibody conjugate consisting of compound 11 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0235] Compound 11 (0.85 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution system, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-11b (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0236] Example 12: 2-[N-methyl-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Val-Ala-methylene)]aminooxyacetylexatecan (12) [ka] 5c (0.2 g, 0.75 mmol) was dissolved in DCM (5 mL), 11b (146 mg, 0.75 mmol) and DIPEA (290.79 mg, 2.24 mmol) were added, the tube was sealed, and the mixture was heated to 50 °C and reacted overnight. The mixture was then cooled to room temperature and purified by silica gel column chromatography to give product 12a (0.27 g, 84% yield, 95% HPLC yield). LCMS: (M+1) + 426.21 (theoretical value: 425.36).

[0237] 12a (0.27 g, 3.9 mmol) was added to a 100 mL single-neck flask, and methanol (5 mL) and 10% palladium on carbon (0.1 g) were added. The mixture was purged with hydrogen gas and reacted at room temperature and atmospheric pressure for 2 hours. The mixture was filtered and concentrated to give 12b (0.20 g, 95% yield, HPLC 93%). LCMS: (M+1) + 336.08 (theoretical value: 335.11).

[0238] A reaction flask was charged with exatecan sulfonate (0.26 g, 0.5 mmol), DMF (2 mL), HATU (0.19 g, 0.5 mmol), DIPEA (0.129 g, 1 mmol), and 12b (200 mg, 0.59 mmol). The mixture was stirred at room temperature for 15 hours, and then DCM (200 mL) was added. The mixture was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to give a yellow solid 12c (0.3 g, 81% yield, 92% HPLC yield). 1 HNMR(500MHz,DMSO-d6)δ8.72-8.66(m,1H),8.49(d,J=8.8Hz,1H),7.79-7.71(m,1H),7.70(s,3H),7.28 (s,1H),6.51(s,1H),5.39(d,J=17.2Hz,3H),5.23-5.19(s,1H),5.00(s,1H),4.68(q,J=7.3Hz,1H),4.18 (s,3H),4.12-4.01(m,1H),3.18(d,J=17.1Hz,1H),3.07(d,J=17.6Hz,1H),2.37(s,3H),2.00-1.92(m,2 H),1.87-1.80(m,2H),1.29(m,J=7.2Hz,3H),1.25-1.05(m,1H),0.84(q,J=13.5,7.5Hz,3H);LCMS:(M+1) + 753.12 (theoretical value: 752.26).

[0239] 12c (0.1 g, 0.13 mmol) was dissolved in MeOH (5 mL), hydrazine hydrate (39.9 mg, 0.79 mmol) was added, the tube was sealed, and the mixture was heated to 60°C and reacted for 2 hours. After cooling to room temperature, the reaction mixture was poured into saturated brine (100 mL), extracted with ethyl acetate (200 mL), and concentrated to give 12d (0.07 g, 77%, HPLC 92%). 1HNMR(500MHz,DMSO-d6)δ8.91(t,J=6.0Hz,1H),8.58(d,J=8.9Hz,1H),8.17(s,3H),7.66(d,J=10.8Hz,1H),7.27(s,1H),6.52(s, 1H),5.58(td,J=8.3,7.9,4.8Hz,1H),5.39(s,2H),5.19(d,J=19.0Hz,1H),5.01(d,J=18.9Hz,1H),4.35(dd,J=13.2,6.3Hz,1H),4 .25(dd,J=12.1,4.7Hz,1H),4.20(d,J=2.3Hz,2H),3.97-3.88(m,1H),3.18(dt,J=17.1,5.9Hz,1H),3.13-3.03(m,1H),2.53(s,3H) ),2.31(d,J=1.9Hz,3H),2.26-2.09(m,2H),1.93-1.77(m,J=7.2Hz,2H),1.37(d,J=6.9Hz,3H),0.86(t,J=7.3Hz,3H);LCMS:(M+1) + 623.15 (theoretical value: 622.26).

[0240] 12d (70 mg, 0.112 mmol) was dissolved in DMF (2 mL), DIPEA (29 mg, 0.224 mmol) was added, and Fmoc-PEG2-Gly-Val-OH (62.16 mg, 0.112 mmol) and HATU (42.56 mg, 0.112 mmol) were added sequentially with stirring. The mixture was stirred at room temperature for 1 hour, concentrated, and purified by silica gel column chromatography to give compound 12e (81 mg, 62% yield, 91% HPLC yield). 1HNMR(500MHz,DMSO-d6)δ8.50(d,J=8.9Hz,1H),8.24(t,J=6.3Hz,1H),8.11(t,J= 5.7Hz,1H),7.99(d,J=7.2Hz,1H),7.86(d,J=7.5Hz,2H),7.74(dd,J=9.8,7.4Hz, 2H),7.67(d,J=7.5Hz,2H),7.39(t,J=7.4Hz,2H),7.35-7.26(m,4H),6.52(s,1H) ,5.75(s,2H),5.59(dt,J=9.2,6.1Hz,1H),5.41(d,J=2.0Hz,2H),5.25(d,J=18.9H z,1H),5.09(d,J=18.8Hz,1H),4.33-3.99(m,8H),3.70(m,J=16.6,5.7Hz,2H),3. 57(t,J=6.5Hz,2H),3.46(s,4H),3.39(d,J=6.0Hz,2H),3.26-3.06(q,J=6.1Hz,5 H),2.48(d,J=2.4Hz,3H),2.41-2.29(m,5H),2.17(q,J=6.5Hz,2H),1.93-1.77(m ,3H),1.14(d,J=7.1Hz,3H),0.86(t,J=7.3Hz,3H),0.82-0.66(m,6H);LCMS:(M+1) + 1160.24 (theoretical value: 1159.50).

[0241] 12e (81 mg, 0.069 mmol) was added to a reaction flask, followed by DMF (1 mL) and piperidine (100 μL). The mixture was allowed to react at room temperature for 1 hour, methyl tert-butyl ether (20 mL) was added, the mixture was centrifuged, the supernatant was removed, and the mixture was dried to give 12f (32 mg, 49% yield, 93% HPLC yield). 1HNMR(500MHz,DMSO-d6)δ8.53(d,J=8.9Hz,1H),8.28(t,J=6.3Hz,1H),8.13(t,J=5.7Hz,1H),8.01(d,J=7.2Hz,1H),7.77(t,J=10.2Hz,5H),7.30( s,1H),6.53(s,1H),5.60(dt,J=8.9,6.0Hz,1H),5.41(s,2H),5.27(d,J= 18.9Hz,1H),5.13(d,J=19.0Hz,1H),4.28(dd,J=13.2,6.7Hz,1H),4.24-4 .12(m,3H),4.12-4.03(m,2H),3.79-3.64(m,2H),3.63-3.48(m,8H),3.2 7-3.08(m,2H),2.97(h,J=5.7Hz,2H),2.47(s,3H),2.37(dd,J=7.8,4.3H z,5H),2.18(q,J=6.4Hz,2H),1.85(qq,J=14.0,7.2Hz,3H),1.15(d,J=7.1Hz,3H),0.86(t,J=7.3Hz,3H),0.71(dd,J=13.3,6.8Hz,6H);LCMS:(M+1) + 938.02 (theoretical value: 937.43)

[0242] DCM (3 mL), 12f (32 mg, 0.034 mmol), bromoacetic acid (9.5 mg, 0.068 mmol), and DIC (8.6 mg, 0.068 mmol) were added to the reaction flask in this order, and the mixture was stirred at room temperature for 90 min. The resulting mixture was concentrated and purified by silica gel column chromatography to give a yellow solid 12 (22 mg, 61% yield, 95% HPLC yield). 1HNMR(500MHz,DMSO-d6)δ8.49(d,J=8.9Hz,1H),8.33(t,J=5.7Hz,1H),8.24(t,J=6.2Hz,1H),8.11(t,J=5.8Hz,1H),7.98(d,J=7.1Hz,1H),7.75(d,J=8 .5Hz,1H),7.70(d,J=10.8Hz,1H),7.28(s,1H),5.64-5.54(m,1H),5.40(s, 2H),5.23(d,J=18.9Hz,1H),5.04(d,J=18.9Hz,1H),4.33-3.97(m,6H),3.8 5(s,2H),3.70(qd,J=16.5,5.7Hz,3H),3.57(t,J=6.5Hz,4H),3.52-3.37(m ,4H),3.26-3.17(m,4H),3.14-3.06(m,1H),2.47(s,3H),2.41-2.28(m,5H) ,2.24-2.11(m,J=6.9,6.0Hz,2H),1.84(dh,J=20.8,7.0Hz,3H),1.14(d,J= 7.1Hz,3H),0.85(t,J=7.3Hz,3H),0.70(dd,J=11.4,6.8Hz,6H);LCMS:(M+1) + 1058.24 (theoretical value: 1057.36).

[0243] ADC-12a: An antibody conjugate consisting of Compound 12 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0244] Compound 12 (0.85 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM L-histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20 using a NAP-5 gel column (Cytiva) to obtain ADC-12a (3.3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0245] ADC-12b: an antibody conjugate consisting of compound 12 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0246] Compound 12 (0.85 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotating turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-12b (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.7

[0247] Example 13: 2-[N-methyl-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Gly-Val-methylene)]aminooxyacetylexatecan (13) [ka] 5b (0.2 g, 0.68 mmol) was dissolved in DCM (5 mL), 11a (133 mg, 0.68 mmol) and DIPEA (175 mg, 1.36 mmol) were added, the tube was sealed, and the mixture was heated to 50 °C and reacted overnight. The mixture was then cooled to room temperature, concentrated, and purified by silica gel column chromatography to give product 13a (0.27 g, 90% yield, HPLC 93%). LCMS: (M+1) + 454.11 (theoretical value: 453.19).

[0248] A 100 mL reaction flask was charged with 13a (0.27 g, 0.59 mmol), methanol (5 mL) and 10% palladium on carbon (0.1 g), and the mixture was purged with hydrogen gas. The mixture was reacted at room temperature and atmospheric pressure for 2 hours, filtered, and concentrated to give product 13b (0.20 g, 95% yield, HPLC 91%). LCMS: (M+1) + 364.03 (theoretical value: 363.14).

[0249] Exatecan sulfonate (0.26 g, 0.5 mmol) was dissolved in DMF (2 mL), HATU (0.19 g, 0.5 mmol), DIPEA (0.129 g, 1 mmol), and 13b (0.2 g, 0.55 mmol) were added, and the mixture was stirred at room temperature for 15 hours. DCM (200 mL) was added, and the mixture was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to give a yellow solid 13c (0.32 g, 84% yield, 94% HPLC yield). LCMS: (M+1) + 781.12 (theoretical value: 780.29).

[0250] 13c (0.1 g, 0.12 mmol) was dissolved in MeOH (5 mL), hydrazine hydrate (66 mg, 1.32 mmol) was added, the tube was sealed, and the mixture was heated to 60 °C and reacted for 2 hours. After cooling to room temperature, the reaction mixture was poured into saturated brine (100 mL), extracted with ethyl acetate (200 mL), and concentrated to give 13d (0.06 g, 72% yield, HPLC 90%). LCMS: (M+1) + 651.06 (theoretical value: 650.29).

[0251] 13d (60 mg, 0.092 mmol) was dissolved in DMF (2 mL), DIPEA (24 mg, 0.184 mmol) was added, and Fmoc-PEG2-Gly-Gly-OH (47.3 mg, 0.092 mmol) and HATU (35.07 mg, 0.092 mmol) were added sequentially with stirring. The mixture was stirred at room temperature for 1 hour, concentrated, and purified by silica gel column chromatography to give compound 13e (86 mg, 81% yield, 91% HPLC yield). LCMS: (M+1) + 1146.28 (theoretical value: 1145.49).

[0252] A reaction flask was charged with 13e (86 mg, 0.075 g), DMF (1 mL), and piperidine (100 μL). The mixture was stirred at room temperature for 1 hour, and methyl tert-butyl ether (20 mL) was added. The mixture was centrifuged, the supernatant was removed, and the mixture was dried to give 13f (36 mg, 52% yield, 89% HPLC purity). LCMS: (M+1) + 924.00 (theoretical value: 923.42).

[0253] A reaction flask was charged with DCM (3 mL), 13f (36 mg, 0.039 mmol), bromoacetic acid (10.6 mg, 0.078 mmol), and DIC (9.8 mg, 0.078 mmol). The mixture was stirred at room temperature for 90 minutes, concentrated, and purified by silica gel column chromatography to give a yellow solid 13 (17 mg, 42.5% yield, 95% HPLC). LCMS: (M+1) + 1044.25 (theoretical value: 1043.34).

[0254] ADC-13a: An antibody conjugate consisting of Compound 13 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0255] Compound 13 (0.83 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva) to obtain ADC-13a (3.1 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0256] ADC-13b: an antibody conjugate consisting of compound 13 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0257] Compound 13 (0.83 mg, 0.8 mmol) was dissolved in 0.08 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotating turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-13b (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.4

[0258] Example 14: 2-[N-methyl-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Leu-Val-methylene)]aminooxyacetylexatecan (14) [ka] 13a (0.10 g, 0.15 mmol) was dissolved in DMF (2 mL), HATU (1.17 g, 0.31 mmol), DIPEA (0.4 g, 0.31 mmol), and Fmoc-PEG2-Gly-Leu-OH (176 mg, 0.31 mmol) were added, and the reaction mixture was stirred at room temperature for 15 h. DCM (200 mL) was added, and the mixture was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to give yellow solid 14a (150 mg, 83% yield, 96% HPLC yield). LCMS: (M+1) + 1202.35 (theoretical value: 1201.55).

[0259] 14a (150 mg, 0.12 mmol) was dissolved in DMF (0.5 mL), piperidine (90 mg, 1.2 mmol) was added, and the mixture was stirred at room temperature for 1 hour. Methyl tert-butyl ether (30 mL) was added, the mixture was centrifuged, the supernatant was removed, the solvent was removed under reduced pressure, and the solid product 14b (63 mg, 49% yield, HPLC 93%) was dried and used directly in the next reaction. LCMS: (M+1) + 980.11 (theoretical value: 979.48).

[0260] To the reaction flask was added DCM (3 mL), 14b (63 mg, 0.023 mmol), bromoacetic acid (17 mg, 0.023 mmol), and DIC (16.2 mg, 0.023 mmol). The reaction was stirred at room temperature for 90 min, concentrated, and purified by silica gel column chromatography to give the yellow solid product 14 (34 mg, 49% yield). LCMS: (M+1) + 1100.04 (theoretical value: 1099.40).

[0261] ADC-14a: An antibody conjugate consisting of Compound 14 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0262] Compound 14 (0.88 mg, 0.8 mmol) was dissolved in 0.09 ml of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva), to obtain ADC-14a (3.2 mg / ml, 2 mL). Average value calculated by UV-HPLC: n=7.7

[0263] ADC-14b: an antibody conjugate consisting of compound 14 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0264] Compound 14 (0.88 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer solution was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7, using a NAP-5 gel column (Cytiva), to obtain ADC-14b (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.4

[0265] Example 15: 2-[N-methyl-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Phe-Gly-methylene)]aminooxyacetylexatecan (15) [ka] 11f (60 mg, 0.098 mmol) was dissolved in DMF (2 mL), DIPEA (25 mg, 0.197 mmol) was added, and while stirring, Fmoc-PEG2-Gly-Phe-OH (59.02 mg, 0.098 mmol) and HATU (37.5 mg, 0.098 mmol) were added sequentially. The reaction mixture was stirred at room temperature for 1 hour, concentrated, and purified by silica gel column chromatography to give compound 15a (92 mg, 79% yield, 93% HPLC yield). LCMS: (M+1) + 1194.36 (theoretical value: 1193.49).

[0266] 15a (92 mg, 0.077 mg) was added to a reaction flask, and DMF (1 mL) and piperidine (100 μL) were added. The reaction mixture was stirred at room temperature for 1 hour, and methyl tert-butyl ether (20 mL) was added. The mixture was centrifuged, the supernatant was removed, and the mixture was dried to give 15b (45 mg, 60% yield, 92% HPLC yield). LCMS: (M+1) + 972.38 (theoretical value: 971.42).

[0267] To a reaction flask were added DCM (3 mL), 15b (45 mg, 0.046 mmol), bromoacetic acid (12.60 mg, 0.092 mmol), and DIC (11.67 mg, 0.092 mmol). The reaction mixture was stirred at room temperature for 90 min, concentrated, and purified by silica gel column chromatography to give a yellow solid 15 (15 mg, 30% yield, 94% HPLC yield). LCMS: (M+1) + 1092.28 (theoretical value: 1091.34).

[0268] ADC-15a: An antibody conjugate consisting of Compound 15 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0269] Compound 15 (0.87 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.2 g / L polysorbate 20, using a NAP-5 gel column (Cytiva) to obtain ADC-15a (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.7

[0270] ADC-15b: an antibody conjugate consisting of compound 15 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (20.0 mg / mL, 10 mg, 0.066 mmol) was taken and the pH was adjusted to 7.2 using 1 M NaHPO solution. Then, 0.1 M ethylenediaminetetraacetic acid disodium solution (25 μL) was added, followed by the prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0271] Compound 15 (0.87 mg, 0.8 mmol) was dissolved in 0.09 mL of DMA, added to the above solution, mixed well, and then reacted on a rotary turntable at room temperature for 2 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine solution, 250 mM sorbitol, 0.02% Twenty-nine 80, pH 5.7 using a NAP-5 gel column (Cytiva) to obtain ADC-15b (3.2 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.6

[0272] Example 16: 2-[N-methyl-N-(bromoacetamidoethoxyethoxypropionyl-Gly-Val-Ala-Gly-methylene)]aminooxyacetylexatecan (16) [ka] 11f (50 mg, 0.082 mmol) was dissolved in DMF (2 mL), DIPEA (10.578 mg, 0.082 mmol) was added, and the mixture was stirred until clear. Fmoc-PEG2-Gly-Val-Ala-OH (51.41 mg, 0.082 mmol) and HATU (32 mg, 0.082 mmol) were added sequentially, and the mixture was stirred at room temperature for 1 hour. The reaction was then purified by silica gel column chromatography to give compound 16a (79 mg, 78% yield, 96% HPLC yield). LCMS: (M+1) + 1216.52 (theoretical value: 1217.32).

[0273] 16a (79 mg, 0.064 mg) was placed in a 10 mL single-neck flask and dissolved in DMF (1 mL). 100 μL of piperidine was added and the mixture was allowed to react at room temperature for 1 hour. 20 mL of methyl tert-butyl ether was added, the mixture was centrifuged, the supernatant was removed, and the resulting solid was purified again with a small amount of methanol and dried to give 16b (26 mg, 45% yield, 95% HPLC yield). LCMS: (M+1) + 994.46 (theoretical value: 995.08).

[0274] A 10 mL single-neck flask was charged with DCM (3 mL), 16b (26 mg, 0.026 mmol), bromoacetic acid (7.2 mg, 0.052 mmol), and DIC (6.6 mg, 0.052 mmol) in that order. The mixture was stirred at room temperature for 90 minutes, concentrated, and purified by silica gel column chromatography to give a yellow solid 16 (12 mg, 41% yield, HPLC 98.5%). LCMS: (M+1) + 1114.38 (theoretical value: 1115.01).

[0275] ADC-16a: An antibody conjugate consisting of Compound 16 and the HER2 monoclonal antibody HS627 [ka] HS627 antibody (10.0 mg / mL, 10 mg, 0.066 mmol) was taken, and the pH was adjusted to 7.2 using 1 M NaHPO solution. 5 mM ethylenediaminetetraacetic acid disodium solution (20 mg / mL, 84 μL) was added, followed by 6 eq of prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was allowed to react on a rotating turntable at room temperature (25°C) for 90 minutes.

[0276] Compound 16 (0.89 mg, 0.8 mmol) was dissolved in DMA (0.09 mL), added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 16 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.02% Twenty-nine 80, using a NAP-5 gel column (Cytiva), to obtain ADC-16a (3 mg / mL, 2 mL). Average value calculated by UV-HPLC: n = 7.80

[0277] ADC-16b: an antibody conjugate consisting of compound 16 and 5T4 monoclonal antibody IP140B [ka] IP140B antibody (10.0 mg / mL, 10 mg, 0.066 mmol) was taken, and the pH was adjusted to 7.2 using 1 M NaHPO solution. 5 mM ethylenediaminetetraacetic acid disodium solution (20 mg / mL, 84 μL) was added, followed by 6 eq of prepared TCEP·HCl (tris(2-carboxyethyl)phosphine hydrochloride) solution (10 mM, 0.04 mL). The mixture was then incubated on a rotating turntable at 25°C for 90 minutes.

[0278] Compound 16 (0.89 mg, 0.8 mmol) was dissolved in DMA (0.09 mL), added to the above solution, and mixed well. The mixture was then reacted on a rotary turntable at room temperature for 16 hours. After the reaction was completed, the buffer was replaced with 20 mM histidine acetate buffer, pH 6.0, 120 mM sucrose, and 0.02% Twenty-nine 80, using a NAP-5 gel column (Cytiva), to obtain ADC-16b (2.9 mg / mL, 2 mL). Average value calculated by UV-HPLC: n=7.78

[0279] Test Example 1: In vivo tumor growth inhibitory activity of ADC Test method for in vitro inhibitory activity of ADC: Human non-small cell lung cancer cells Calu-6, human lung cancer cells NCI-H1975, esophageal adenocarcinoma cells OE-21, human esophageal squamous cell carcinoma cells KYSE-150, human ovarian cancer cells ES-2, and human breast cancer cells MDA-MB-231 were cultured in vitro as monolayers. When the cell density reached 80%-90%, they were digested with trypsin-EDTA, centrifuged, and the supernatant discarded. The cells were resuspended in PBS and adjusted to the appropriate cell concentration. Calu-6, NCI-H1975, KYSE-150, OE-21, and MDA-MB-231 cells (2-10 × 10) were cultured in vitro as monolayers. 6 BALB / c nude mice were subcutaneously inoculated with ES-2 cells (0.1 mL / cells), and NOD-SCID mice were subcutaneously inoculated with ES-2 cells. The animals and tumor growth were monitored regularly until the tumor volume reached 100–200 mm. 3 Once the tumors had grown to approximately 100% tumor size, they were randomly divided into groups based on tumor volume and body weight: a solvent control group (or vehicle group, injected with saline) and an ADC-treated group (dissolved in saline), with six animals in each group. The drugs were administered intravenously. The administration frequency was Q4D, with a total of two doses (the first dose was on Day 1, and the second on Day 5). See Table 1 for the mouse grouping and administration settings. The tumor's major axis (a) and minor axis (b) were measured twice a week using calipers, and the mouse's body weight was measured. Tumor volume (V) was calculated using the following formula: V = 1 / 2 × a × b 2 (mm 3), where a and b represent the length and width of the tumor, respectively. A growth curve was then created, and the tumors were finally excised and weighed. Statistical analysis was performed using GraphPad Prism software using the tumor volume and weight data of the tumor-bearing mice at the end of the experiment to obtain the results of the antitumor effect. In the figure, "" in the control group indicates that there was no result for that group or that the tumor status was not measured. In the experimental group, "no tumor" in the corresponding group, "tumor shrinkage to 0," and "tumor disappearance to 0" indicate that no tumor was found in the corresponding dissected animal. JPEG2026501616000109.jpg189170JPEG2026501616000110.jpg149170

[0280] The foregoing description of specific exemplary embodiments of the present invention has been presented for purposes of explanation and illustration. It is not intended that these descriptions be construed as limiting the invention to the precise forms disclosed, and it is apparent that many modifications and variations are possible in light of the above teachings. The purpose of selecting and describing the exemplary examples is to explain certain principles of the present invention and their practical application, so that those skilled in the art can realize and utilize various different exemplary embodiments of the present invention, as well as various alternatives and modifications. It is intended that the scope of the present invention be limited by the claims and their equivalents.

Claims

1. An antibody-drug conjugate as shown in formula (I): 【Chemistry 1】 (In the formula, D is a cytotoxic drug and Ab is an antibody; L Ｄ does not exist or C １ ~C ３ alkylene groups, X is selected from N, O or S; R １ does not exist, hydrogen, deuterium, C １ ~C ６ Alkyl or halogen-substituted C １ ~C ６ alkyl groups, R ２ is hydrogen, deuterium, C １ ~C ６ Alkyl group, C １ ~C ６ selected from an alkoxy group, an acyl group, and a sulfonyl group; L １ Is, -L １１ -L １２ -L １３ -, where L １１ , L １２ , and L １３ are each independently absent, —C═O—, C １ ~C ６ Alkylene group or —O—C １ ~C ６ alkylene; L ２ is C １ ~C ６ Alkylene group or C １ ~C ６ acyl group, １ ~C ６ Alkylene group or C １ ~C ６ The acyl group is formed by one or more R ３ is optionally replaced by R ３ is a phenyl-substituted or unsubstituted C １ ~C ６ Alkyl group, C １ ~C ６ alkoxy groups, L Ｐ is a peptide residue consisting of 2 to 7 amino acids, Z is -L ｚ -L ｊ -, where L ｚ is -C(=O)-C １ ~C ８ Alkylene group or —C(═O)—(CH ２ CH ２ O) ２～６ -CH ２ CH ２ NH—, and L ｊ is a linker that can be linked to an antibody.

2. R １ does not exist, hydrogen, deuterium, C １ ~C ３ Alkyl or halogen-substituted C １ ~C ３ alkyl groups, Preferably, R １ is absent, hydrogen, deuterium, or halogen-substituted C １ ~C ３ alkyl groups, Preferably, R １ is absent, hydrogen, deuterium, or C substituted with F, Cl, Br, or I １ ~C ３ alkyl groups, Preferably, R １ is absent, hydrogen, deuterium or F-substituted C １ ~C ３ alkyl groups, Preferably, R １ is selected from absent, hydrogen, deuterium, a fluoromethyl group, a difluoromethyl group, a trifluoromethyl group, a fluoroethyl group, a difluoroethyl group, or a trifluoroethyl group; Preferably, R １ is selected from absent, hydrogen, a difluoroethyl group, or a trifluoroethyl group; Preferably, R ２ is hydrogen, deuterium, C １ ~C ６ Alkyl group, C １ ~C ６ Alkoxy group, —C(═O)C １ ~C ６ Alkyl group, or -S(=O) ２ C １ ~C ６ alkyl groups, Preferably, R ２ is hydrogen, deuterium, C １ ~C ６ Alkyl group, C １ ~C ６ Alkoxy group, —C(═O)C １ ~C ６ Alkyl group, or -S(=O) ２ C １ ~C ３ alkyl groups, Preferably, R ２ is hydrogen, deuterium, C １ ~C ３ Alkyl group, C １ ~C ３ Alkoxy group, —C(═O)C １ ~C ３ Alkyl group, or -S(=O) ２ C １ ~C ３ alkyl groups, Preferably, R ２ is selected from hydrogen, deuterium, methyl, ethyl, methoxy, ethoxy, formyl, acetyl, methanesulfonyl, or ethanesulfonyl; Preferably, R ２ is selected from hydrogen, methyl, methoxy, formyl, or methanesulfonyl; Preferably, R ２ is selected from hydrogen, a formyl group or a methanesulfonyl group; The antibody-drug conjugate of claim 1.

3. L １１ , L １２ , and L １３ are each independently absent, —C(═O)—, C １ ~C ２ Alkylene group or C １ ~C ２ alkylene-O-, with the proviso that L １１ When is —C(═O)—, R ２ is not hydrogen, Preferably, L １１ , L １２ , and L １３ are each independently absent, —C═O—, —CH ２ - or -OCH ２ - is selected from, Preferably, L １ is -C(=O)CH ２ OCH ２ -, -C(=O)CH ２ O- or -CH ２ - is selected from, Preferably, L ２ is C １ ~C ３ Alkylene group or C １ ~C ３ acyl group, １ ~C ３ Alkylene group or C １ ~C ３ The acyl group is formed by one or more R ３ is optionally replaced by Preferably, L ２ is a methylene group, an ethylene group, or —C(═O)CH ２ -, wherein the methylene group, the ethylene group, or the -C(=O)CH ２ - represents one or more R ３ is optionally replaced by Preferably, R ３ is C １ ~C ６ Alkyl group, C １ ~C ６ Alkoxy group, or phenyl-substituted C １ ~C ６ alkyl groups, Preferably, R ３ is C １ ~C ６ Alkyl or phenyl substituted C １ ~C ６ alkyl groups, Preferably, R ３ is C １ ~C ４ Alkyl or phenyl substituted C １ ~C ３ alkyl groups, Preferably, R ３ is selected from a methyl group, an ethyl group, a propyl group, an isopropyl group, a butyl group, an isobutyl group, a phenylmethyl group, or a phenylethyl group; Preferably, R ３ is selected from a methyl group, an isopropyl group, an isobutyl group, or a phenylmethyl group; Preferably, L ２ is -CH ２ -, -CH ２ CH ２ -, a group selected from the group represented by the formula: 【Chemistry 2】 Preferably, L ２ is -CH ２ -, -CH ２ CH ２ -, a group selected from the group represented by the formula: 【Transformation 3】 Preferably, L ２ is -CH ２ -, -CH ２ CH ２ -, selected from the groups represented by the following formulas: 【Chemistry 4】 The antibody-drug conjugate according to claim 1 or 2.

4. Preferably, said amino acids are selected from phenylalanine (Phe), glycine (Gly), valine (Val), alanine (Ala), leucine (Leu), lysine (Lys) or glutamic acid (Glu); Preferably, said amino acids are selected from phenylalanine (Phe), glycine (Gly), valine (Val), alanine (Ala) or leucine (Leu); Preferably, L ｐ is selected from peptide residues consisting of 2 to 5 amino acids selected from phenylalanine (Phe), glycine (Gly), valine (Val), alanine (Ala) or leucine (Leu); Preferably, L ｐ is selected from peptide residues consisting of 2, 3 or 4 amino acids selected from phenylalanine (Phe), glycine (Gly), valine (Val), alanine (Ala) or leucine (Leu); Preferably, L ｐ is selected from -Val-Cit-, -Gly-Lys-, -Gly-Leu-, -Val-Ala-, -Gly-Phe-, -GLy-Gly-Lys-, -Gly-Gly-Phe-, -Gly-Val-Ala-, -Gly-Gly-Val-, -Gly-Leu-Val-, -Gly-Phe-Gly- or -Gly-Gly-Leu-; Preferably, L ｐ is selected from -Gly-Leu-, -Gly-Phe-, -Gly-Gly-Phe-, -Gly-Val-Ala-, -Gly-Gly-Val-, -Gly-Leu-Val-, -Gly-Phe-Gly or -Gly-Gly-Leu-; Preferably, L ｐ is selected from the group represented by the formula: 【Transformation 5】 Preferably, L ｊ is selected from the group represented by the formula: 【Transformation 6】 【Transformation 7】 The position indicated by indicates that it is linked to an antibody, 【Transformation 8】 The position indicated by L Ｚ represents being linked to a group, Preferably, L ｚ is -C(=O)-C １ ~C ８ Alkenyl group or —C(═O)—(CH ２ CH ２ O) ２～６ -CH ２ CH ２ NH-; Preferably, L ｚ is -C(=O)-(CH ２ CH ２ O) ２～６ -CH ２ CH ２ NH-; Preferably, L ｚ is -C(=O)-(CH ２ CH ２ O) ２～４ -CH ２ CH ２ NH-; Preferably, L ｚ is -C(=O)-(CH ２ CH ２ O) ２ -CH ２ CH ２ NH-; Preferably, Z is 【Chemistry 9】 Selected from: The antibody-drug conjugate according to any one of claims 1 to 3.

5. the cytotoxic drug is selected from camptothecin and its derivatives; Preferably, the cytotoxic drug is selected from the following formula: 【Chemistry 10】 Preferably, the antibody is an anti-tumor associated antigen antibody, Preferably, the tumor-associated antigen antibody is selected from an anti-Her2 antibody, an anti-Trop2 antibody, an anti-B7H3 antibody, an anti-5T4 antibody, an anti-Nectin-4 antibody, an anti-CD20 antibody, and an anti-ROR1 antibody. The antibody-drug conjugate according to any one of claims 1 to 4.

6. The antibody-drug conjugate according to any one of claims 1 to 5, wherein the structure of the compound represented by formula (I) is represented by formula (II), formula (III), or formula (IV). 【Chemistry 11】 (In formula (II), R ２ , L １ , L ２ , L ｐ , Z, and Ab are each defined as in the antibody-drug conjugate of formula (I), Preferably, L １ is -C(=O)CH ２ OCH ２ - or -C(=O)CH ２ O-, Preferably, L ２ Ha-CH ２ - or -CH ２ CH ２ - is selected from, Preferably, 【Chemistry 12】 teeth 【Chemistry 13】 is selected from 【Chemistry 14】 (In formula (III), R １ , R ２ , R ４ , L ｐ , Z, and Ab are each defined as in the antibody-drug conjugate of formula (I), Preferably, R ４ is hydrogen, phenyl substituted or unsubstituted C １ ~C ６ Alkyl group, C １ ~C ６ alkoxy groups, Preferably, R ４ is hydrogen, C １ ~C ６ Alkyl group, C １ ~C ６ Alkoxy group, or phenyl-substituted C １ ~C ６ alkyl groups, Preferably, R ４ is hydrogen, C １ ~C ６ Alkyl or phenyl substituted C １ ~C ６ alkyl groups, Preferably, R ４ is hydrogen, C １ ~C ４ Alkyl or phenyl substituted C １ ~C ３ alkyl groups, Preferably, R ４ is selected from hydrogen, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, phenylmethyl or phenylethyl; Preferably, R ４ is selected from hydrogen, a methyl group, an isopropyl group, an isobutyl group, or a phenylmethyl group; Preferably, R ４ is selected from hydrogen, a methyl group, or an isopropyl group. 【Chemistry 15】 (In formula (IV), R ２ , L １ , L ２ , L ｐ , Z, and Ab are each defined as in the antibody-drug conjugate of formula (I), Preferably, L １ and L ２ are each independently C １ ~C ３ alkylene groups, Preferably, L １ is -CH ２ - and L ２ is -CH ２ CH ２ - is.) 【Request Item 7】 【Chemistry 16】 The antibody-drug conjugate of any one of claims 1 to 6, wherein is selected from the following structures: 【Chemistry 17】 (where R １ is defined as in the antibody-drug conjugate of formula (I).

8. The compound is obtained by linking the following compound to an antibody: Preferably, the antibody is an HS627 antibody or an IP140B antibody, wherein the heavy chain amino acid sequence of HS627 is shown in SEQ ID NO: 1 and the light chain amino acid sequence thereof is shown in SEQ ID NO: 2, and the heavy chain amino acid sequence of the IP140B antibody is shown in SEQ ID NO: 3 and the light chain amino acid sequence thereof is shown in SEQ ID NO:

4. The antibody-drug conjugate according to any one of claims 1 to 7.

9. A pharmaceutical composition comprising the antibody-drug conjugate according to any one of claims 1 to 8 and a pharmaceutically acceptable carrier.

10. Use of the antibody-drug conjugate of any one of claims 1 to 8 or the pharmaceutical composition of claim 9 in the preparation of an antitumor drug, Preferably, the tumor is selected from solid tumors, more preferably selected from lung cancer, non-small cell lung cancer, esophageal squamous cell carcinoma, ovarian cancer, esophageal adenocarcinoma or breast cancer.

11. A method for treating a tumor disease, comprising the step of administering the antibody-drug conjugate according to any one of claims 1 to 8, or the pharmaceutical composition according to claim 9, to a patient in need thereof.

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