A dual linker compound and applications thereof

Abstract

This invention relates to a dual-linker compound and its applications. The dual-linker compound of this invention has two linkers capable of conjugating to antibodies and a linker capable of conjugating to cytotoxic drugs. The dual-linker compound exhibits excellent conjugation to antibodies and demonstrates very good uniformity in its DAR value.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and more specifically, to a class of dual linkers used in ADC drugs. Background Technology

[0002] Traditional antibody-drug conjugates (ADCs) utilize the amino group of lysine in antibodies or the thiol group of cysteine ​​obtained by breaking interchain disulfide bonds for conjugation. The ADCs obtained by this method are highly heterogeneous mixtures with poor homogeneity and low stability, which affects efficacy and therapeutic window. Therefore, it is particularly important to obtain ADCs with controllable drug-antibody ratio (DAR) and high homogeneity.

[0003] Re-bridging technology is based on the reduction and re-bridging of disulfide bonds in antibodies. Cysteine-selective cross-linking reagents (such as TCEP or DTT) are used to reduce interchain disulfide bonds in IgG1 antibodies. Subsequently, a two-reaction reagent is used to simultaneously re-link the polypeptide chains, thereby loading small molecule payloads or further modifying the antibody.

[0004] This technology boasts advantages such as high uniformity, good reactivity, and stability. It maintains the stability of disulfide bonds by covalently relinking cysteine ​​residues, while simultaneously achieving controlled coupling of one payload per disulfide bond. Re-bridging coupling technology can be used for direct localization modification of natural antibodies, has broad applicability, controllable DAR values, and produces highly uniform ADCs. For example, Igenica Biotherapeutics' SNAP platform and Abzena's Thiobridge platform utilize disulfide bond reduction modification technology. Despite the significant advantages of re-bridging coupling technology, some challenges remain, such as the risk of intra-chain misbridging, which may affect the quality and efficacy of ADCs.

[0005] With technological advancements, rebridge coupling techniques are constantly being optimized and improved. For example, TetraDVP technology overcomes the formation of fragmented "half-antibody" species by increasing the number of cysteine ​​reactive groups in each linker, thereby improving coupling efficiency.

[0006] Therefore, it is of great significance to design a dual linker that can better match antibodies and form stable and highly homogeneous antibody-drug conjugates. Summary of the Invention

[0007] This invention provides bilinker compounds for the preparation of ADC drugs.

[0008] In a first aspect, the present invention provides a dual-linker compound or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates, or isotopically labeled analogs thereof, said dual-linker compound having the structure shown in formula (I):

[0009]

[0010] in,

[0011] A1 and A2 may be the same or different, and are selected independently from:

[0012] R1 is independently selected from: C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 1-6 Alkylamine alkyl, C 1-6 Alkoxyalkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene;

[0013] Cy is independently selected from: substituted or unsubstituted C 3-6 Cycloalkyl, substituted or unsubstituted 3-6 membered heterocyclic groups, substituted or unsubstituted aryl groups, substituted or unsubstituted heteroaryl groups;

[0014] L A1 L A2 Same or different, independently selected from: single bond, substituted or unsubstituted C 1-12 Subchain hydrocarbon groups;

[0015] L1, L2, and L3 are linking groups;

[0016] M represents a functional group or leaving group that can be linked to cytotoxic drugs.

[0017] In another preferred embodiment, R1 is independently selected from: C 1-6 Alkyl, C 1-6 Deuterated alkyl, C 3-6 Cycloalkyl, aryl, arylalkylene.

[0018] In another preferred embodiment, Cy is independently selected from: substituted or unsubstituted: phenyl, naphthyl, furanyl, thiophene, pyrrole, oxazolyl, thiazolyl, imidazolyl, benzofuranyl, benzothiophene, benzopyrrole, benzooxazolyl, benzothiazolyl, benzoimidazolyl, pyridyl, pyrazinyl, pyridazinyl, quinolinyl, isoquinolinyl, quinoxalinyl, quinazolinyl.

[0019] In another preferred embodiment, Cy is independently selected from one of the following structures:

[0020]

[0021] Where x is the same or different, and is independently selected from CR a Or N, and L A1 The connected x is selected from C; R a Whether the same or different, they are independently selected from hydrogen, deuterium, and carbon. 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 1-6 alkylamine group, C 1-6 Alkoxy, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene;

[0022] Ring A represents an unsubstituted or substituted aromatic ring, or a substituted or unsubstituted heteroaromatic ring;

[0023] Y is selected from O, S, NR Y , where R Y Independently selected from hydrogen, deuterium, and C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene;

[0024] "*1" indicates the connection site with SO2R1; Indicates with L A1 The connection site.

[0025] Preferably, ring A represents a substituted or unsubstituted ring: benzene ring, naphthalene ring, pyridine ring, pyrimidine ring.

[0026] Preferred, Choose one of the following structures:

[0027]

[0028] Where y is the same or different, it is independently selected from CR. b Or N, and L A1 The connected y is selected from C; R b Whether the same or different, they are independently selected from hydrogen, deuterium, and carbon. 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6Deuterated alkyl, C 1-6 alkylamine group, C 1-6 Alkoxy, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene.

[0029] Preferably, Cy is independently selected from one of the following structures:

[0030]

[0031] Preferably, Cy is independently selected from one of the following structures:

[0032]

[0033] Preferably, Y is independently selected from O, S, and NH, and more preferably from O or S.

[0034] In another preferred embodiment, L A1 L A2 Same or different, independently selected from: single bond, substituted or unsubstituted C 1-8 Subchain hydrocarbon group, the C 1-8 The subchain hydrocarbon group may optionally contain 1, 2 or 3 carbon-carbon double bonds or carbon-carbon triple bonds.

[0035] Preferred, L A1 L A2 Whether the bonds are the same or different, they are independently selected from a single bond and one of the following structures:

[0036]

[0037] a is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

[0038] b is selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[0039] Preferred, The double or triple bond is connected to Cy.

[0040] In another preferred embodiment, A1 and A2 may be the same or different, and are independently selected from one of the following structures:

[0041]

[0042] t is selected from 0, 1, 2, 3, 4, 5, or 6;

[0043] q is selected from 0, 1, 2, 3, 4, 5 or 6.

[0044] Preferably, R1 is independently selected from methyl, ethyl, propyl, butyl, and benzyl;

[0045] In another preferred embodiment, L1 and L2 may be the same or different, and are independently selected from: -L a C 1-12 Alkylene L b -, the C 1-12 The alkylene group may have an intercalation between any two C and C groups selected from -O-, -S-, or -NR. L -, -CO-, -COO-, -OCO-, -OCOO-, -CONR L -、-NR L CO-、-OCONR L -、-NR L COO-、-NR L CONR L -、-NR L SO-、-SONR L -、-NR L S(O)2-、-S(O)2NR L - one of the groups;

[0046] L a Whether the same or different, it is independently selected from single bonds, -CO-, -SO-, -S(O)2-, -OCO-, -NR L CO-; L a It is attached to the N atom in the middle of the compound;

[0047] L b Same or different, independently selected from single bond, -O-, -S-, -NR L -, -CO-, -COO-, -OCO-, -OCOO-, -CONR L -、-NR L CO-、-OCONR L -、-NR L COO-、-NR L CONR L -、-NR L SO-、-SONR L -、-NR L S(O)2-、-S(O)2NR L -;L b Connected to L A1 or L A2 superior;

[0048] R L Independently selected from hydrogen, deuterium, and C 1-6 alkyl.

[0049] Preferably, the C1-12 Alkyl groups are selected from: -(CH2) n -, n is independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12.

[0050] Preferably, when L a When attached to the N atom in the middle of a compound, it is attached to the N atom with the -CO-, -SO-, or -S(O)2- ends.

[0051] In another preferred embodiment, L3 is selected from: -L c C 1-12 alkylene-, the C 1-12 The alkylene group may have an intercalation between any two C and C groups selected from -O-, -S-, or -NR. L -, -CO-, -COO-, -OCO-, -OCOO-, -CONR L -、-NR L CO-、-OCONR L -、-NR L COO-、-NR L CONR L -、-NR L SO-、-SONR L -、-NR L S(O)2-、-S(O)2NR L - one of the groups;

[0052] L c Independently selected from single bonds, -CO-, -SO-, -S(O)2-, -OCO-, -NR L CO-; L c It is attached to the N atom in the middle of the compound;

[0053] R L Independently selected from hydrogen, deuterium, and C 1-6 alkyl.

[0054] Preferably, the C 1-12 Alkyl groups are selected from: -(CH2) m -, m is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12.

[0055] Preferably, when L c When attached to the N atom in the middle of a compound, it is attached to the N atom with the -CO-, -SO-, or -S(O)2- ends.

[0056] In another preferred example, M is selected from -OR M -SR M -N(R) M )2、-COORM -CON(R) M )2、-SO(R M )-、-S(O)2(R M )2;

[0057] R M Independently selected from hydrogen, deuterium, and C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 1-6 Alkylamine alkyl, C 1-6 Alkoxyalkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene.

[0058] Preferred, R M Independently selected from hydrogen, deuterium, and C 1-6 alkyl.

[0059] In another preferred embodiment, the dual-connector compound has the structure shown in formula (I-1):

[0060]

[0061] Among them, A1, A2, L a L b n, m, and M are as described in this article.

[0062] In another preferred embodiment, the dual-connector compound has the structure shown in formula (I-2):

[0063]

[0064] Wherein, A1, A2, n, m, and M are as described in this document.

[0065] In another preferred embodiment, the dual-connector compound is selected from:

[0066]

[0067]

[0068] In another aspect, the present invention provides the use of the said dual linker compound or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates or isotopically labeled analogs in the manufacture of pharmaceutical linker compounds.

[0069] In another aspect, the present invention provides the use of the said dual-linker compound or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates or isotopically labeled analogs in the manufacture of antibody-drug conjugates.

[0070] In another aspect, the present invention provides a drug-dual linker compound or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, prodrugs, hydrates, solvates, or isotopically labeled analogs thereof, said drug-dual linker compound having the structure shown in formula (II):

[0071]

[0072] Where D is the cytotoxic drug fragment (i.e. the fragment after the cytotoxic drug reacts with M); M' is the residue of M after the reaction with the cytotoxic drug;

[0073] A1, A2, L1, L2, L3, and M are as described in this article.

[0074] In another aspect of the present invention, the use of the compound of formula (II) or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, prodrugs, hydrates, solvates or isotopically labeled analogs in the manufacture of antibody-drug conjugates is provided.

[0075] In another preferred embodiment, the antibody is selected from at least one of Trastuzumab and Sacituzumab.

[0076] Preferably, in the Trastuzumab, the light chain amino acid sequence is: DIQMTQSPSSLSASVGD RVTITCRASQDVNTAVAWYQQKPGKAPKLLIYSASFLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQHYTTPPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC; the heavy chain amino acid sequence is: EVQLVESGGGLVQPGGSLRLSCAASGFNIKDTYIHWVRQAPGKGLEWVARIYPTNGYTRYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCSRWGGDGFYAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG;

[0077] The light chain amino acid sequence of the Sacituzumab is: DIQLTQSPSSLSASVGDRVSITCKAS QDVSIAVAWYQQKPGKAPKLLIYSASYRYTGVPDRFSGSGSGTDFTLTISSLQPEDFAVYYCQQHYITPLTFGAGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC; the heavy chain amino acid sequence is: VQLQQSGSELKKPGASVKVSCKASGYTFTNYGMNWVKQAPGQGLKWMGWINTYTGEPTYTDDFKGRFAFSLDTSVSTAYLQISSLKADDTAVYFCARGGFGSSYWYFDVWGQGSLVTVSSASTKGP SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVH NAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK.

[0078] In another aspect, the present invention provides a pharmaceutical composition comprising the drug-dual linker compound or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, prodrugs, hydrates, solvates, or isotopically labeled analogs thereof.

[0079] In this invention, the pharmaceutical composition further comprises pharmaceutically acceptable excipients and carriers.

[0080] definition:

[0081] Unless otherwise specified, the term "alkyl" refers to a monovalent saturated aliphatic hydrocarbon group, a straight-chain or branched group comprising 1 to 12 (specifically 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12) carbon atoms, preferably comprising 1 to 6 carbon atoms (i.e., C1-6 alkyl) or 1 to 4 carbon atoms (i.e., C1-4 alkyl). Examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, etc.

[0082] Unless otherwise specified, the term "subchain hydrocarbon group" refers to a divalent saturated or unsaturated chain hydrocarbon group containing 1 to 12 (specifically 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12) carbon atoms, such as a saturated or partially saturated subchain hydrocarbon group containing 1 to 8 carbon atoms; when the subchain hydrocarbon group is unsaturated, it may contain 1, 2, 3, or 4 unsaturated groups selected from carbon-carbon double bonds and carbon-carbon triple bonds. Subchain hydrocarbon groups are, for example, alkylene, alkenylene, and ynylene groups.

[0083] Unless otherwise specified, the term "alkylene" refers to a divalent saturated aliphatic hydrocarbon group obtained by further removing one hydrogen atom from an alkyl group, comprising a straight-chain or branched group containing 1 to 12 (specifically 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12) carbon atoms. Examples include, but are not limited to, methylene, ethylene, n-propylene, n-butylene, n-pentylene, and n-hexylene.

[0084] Unless otherwise specified, the term "alkenyl" refers to an unsaturated aliphatic hydrocarbon group consisting of a straight or branched chain of carbon and hydrogen atoms, having at least one double bond. Alkenyl groups can contain 2-12 carbon atoms (i.e., C2-12 alkenyl), 2-6 carbon atoms (i.e., C2-12 alkenyl), 2-4 carbon atoms (i.e., C2-4 alkenyl), 2-3 carbon atoms (i.e., C2-3 alkenyl), or 2 carbon atoms (i.e., C2 alkenyl). For example, "C2-6 alkenyl" indicates that the group is alkenyl and the number of carbon atoms in the carbon chain is between 2 and 6 (specifically 2, 3, 4, 5, or 6). Non-limiting examples of alkenyl groups include, but are not limited to, vinyl, 1-propenyl, 2-propenyl, 1-butenyl, isobutenyl, and 1,3-butadienyl.

[0085] Unless otherwise specified, the term "alkenyl" refers to a divalent unsaturated aliphatic hydrocarbon group obtained by further removing one hydrogen atom from an alkenyl group. An alkenyl group may contain 2-12 carbon atoms, 2-6 carbon atoms, 2-4 carbon atoms, 2-3 carbon atoms, or 2 carbon atoms. Non-limiting examples of alkenyl groups include, but are not limited to, -(CH2)sCH=CH(CH2)s-, -(CH2)sCH=CH(CH2)sCH=CH(CH2)s- (where s may be the same or different, and are independently selected from 0, 1, 2, 3, 4, 5, or 6), etc.

[0086] Unless otherwise specified, the term "alkynyl" refers to an unsaturated aliphatic hydrocarbon group consisting of a straight or branched chain of carbon and hydrogen atoms, having at least one triple bond. An alkynyl group can contain 2-12 carbon atoms (i.e., C2-12 alkynyl), 2-6 carbon atoms (i.e., C2-12 alkynyl), 2-4 carbon atoms (i.e., C2-4 alkynyl), 2-3 carbon atoms (i.e., C2-3 alkynyl), or 2 carbon atoms (i.e., C2 alkynyl). For example, "C2-6 alkynyl" indicates that the group is alkynyl and the number of carbon atoms in the carbon chain is between 2 and 6 (specifically 2, 3, 4, 5, or 6). Non-limiting examples of alkynyl groups include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, and 1-butynyl.

[0087] Unless otherwise specified, the term "alkynyl" refers to the divalent unsaturated aliphatic hydrocarbon group obtained by further removing one hydrogen atom from an alkynyl group. The alkynyl group may contain 2-12 carbon atoms, 2-6 carbon atoms, 2-4 carbon atoms, 2-3 carbon atoms, or 2 carbon atoms. Non-limiting examples of alkynyl groups include, but are not limited to, -(CH2)sC≡C(CH2)s-, -(CH2)sC≡C(CH2)sC≡C(CH2)s-, -(CH2)sC≡C(CH2)sC≡C(CH2)s- (s may be the same or different, and are independently selected from 0, 1, 2, 3, 4, 5, or 6), etc.

[0088] Unless otherwise specified, the term "cycloalkyl" refers to a monocyclic saturated aliphatic hydrocarbon group having a specific number of carbon atoms, preferably containing 3-6 carbon atoms (i.e., C3-6 cycloalkyl), examples of which include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, methylcyclopropyl, 2-ethyl-cyclopentyl, dimethylcyclobutyl, etc.

[0089] Unless otherwise specified, the term "alkoxy" refers to an -O-alkyl group, which is defined as above as containing 1 to 6 carbon atoms (specifically 1, 2, 3, 4, 5, or 6). Examples include, but are not limited to, methoxy, ethoxy, propoxy, isopropoxy, butoxy, 1-methylpropoxy, 2-methylpropoxy, tert-butoxy, pentoxy, 1-methylbutoxy, 2-methylbutoxy, 3-methylbutoxy, 1,1-dimethylpropoxy, 1,2-dimethylpropoxy, 2,2-dimethylpropoxy, and 1-ethylpropoxy.

[0090] Unless otherwise specified, the term "alkylamine" refers to -NR′R″, where R′ and R″ may be the same or different, and may be H or an alkyl group as defined above. The alkyl group is defined as above, i.e., containing 1 to 20 carbon atoms, preferably 1 to 6 carbon atoms (specifically 1, 2, 3, 4, 5, or 6). Representative examples include, but are not limited to, -NH(CH3), -N(CH3)(CH3), -N(CH2CH3)(CH3), -N(CH2CH3)[CH(CH3)2], etc.

[0091] Unless otherwise specified, the terms "halogen" or "halogenated" refer to F, Cl, Br, and I. The term "halogenated alkyl" refers to an alkyl group as defined above in which one, two, or more hydrogen atoms, or all hydrogen atoms, are replaced by a halogen. Representative examples of halogenated alkyl groups include CCl3, CF3, CHCl2, CH2Cl, CH2Br, CH2I, CH2CF3, and CF2CF3.

[0092] Unless otherwise specified, the term "heterocyclic group" or "heterocycle" refers to a saturated or partially unsaturated monocyclic or polycyclic non-aromatic substituent having a ring carbon atom and 1 to 4 ring heteroatoms, comprising 3-6 ring atoms (3-6 membered heterocyclic group), 4-6 ring atoms (4-6 membered heterocyclic group), or 5-6 ring atoms (5-6 membered heterocyclic group). The number of heteroatoms is preferably 1-4, more preferably 1-3 (i.e., 1, 2, or 3). Examples of monocyclic heterocyclic groups include pyrrolidinyl, imidazoalkyl, tetrahydrofuranyl, tetrahydrothiophenyl, dihydropyrrolidinyl, piperidinyl, piperazinyl, pyranyl, morpholinyl, thiocarolinyl, etc. Polycyclic heterocyclic groups include spirocyclic, fused-ring, and bridged-ring heterocyclic groups. A "heterocyclic group" can be a monocyclic ("monocyclic heterocyclic group") or a fused ("fused heterocyclic group" or "heterofused-cyclic group"), bridged ("heterobridged heterocyclic group" or "bridged-ring heterocyclic group") or spiro-fused ("heterospirocyclic group" or "spirocyclic heterocyclic group") ring system, such as a bicyclic system ("bicyclic heterocyclic group"), and can be saturated or partially unsaturated. A heterocyclic bicyclic system can include one or more heteroatoms in one or both rings. "Heterocyclic group" also includes a ring system in which the heterocyclic ring as defined above is fused with one or more carbocyclic groups, wherein the attachment point is on the carbocyclic or heterocyclic ring; or "heterocyclic group" also includes a ring system in which the heterocyclic ring as defined above is fused with one or more aryl or heteroaryl groups, or a ring system in which a cycloalkyl ring as defined above is fused with one or more heteroaryl groups, wherein the attachment point is on the heterocyclic or cycloalkyl ring, and in such cases, the number of members in the heterocyclic ring system is the number of atoms in the fused ring system. In some embodiments, each example of a heterocyclic group is independently optionally substituted, for example, unsubstituted (an "unsubstituted heterocyclic group") or substituted with one or more substituents (a "substituted heterocyclic group"). Exemplary 3-membered heterocyclic groups containing one heteroatom include, but are not limited to, azirropropyl, oxiranyl, and thiorenyl. Exemplary 4-membered heterocyclic groups containing one heteroatom include, but are not limited to, azahexacyclobutane, oxacyclobutane, and thiohexacyclobutane. Exemplary 5-membered heterocyclic groups containing one heteroatom include, but are not limited to, tetrahydrofuranyl, dihydrofuranyl, tetrahydrophenylthio, dihydrophenylthio, pyrrolidinyl, dihydropyrrolidinyl, and pyrrolidin-2,5-dione. Exemplary 5-membered heterocyclic groups containing two heteroatoms include, but are not limited to, dioxolanecyclopentane, oxathiocyclopentane, dithiocyclopentane, and oxazolidinyl-2-one. Exemplary 5-membered heterocyclic groups containing three heteroatoms include, but are not limited to, triazolinyl, oxadiazolinyl, and thiadiazolinyl. Exemplary 6-membered heterocyclic groups containing one heteroatom include, but are not limited to, piperidinyl, tetrahydropyranyl, dihydropyridinyl, and thianyl.Exemplary 6-membered heterocyclic groups containing two heteroatoms include, but are not limited to, piperazine, morpholinyl, dithiocyclohexyl, and dioxazinanyl. Exemplary 6-membered heterocyclic groups containing three heteroatoms include, but are not limited to, triazacyclohexyl, oxadiazine, thiadiazine, oxathiazine, and dioxazinanyl. Exemplary 5-membered heterocyclic groups fused to a C6 aryl ring (also referred to herein as a 5,6-bicyclic heterocycle) include, but are not limited to, dihydroindolyl, isodihydroindolyl, dihydrobenzofuranyl, dihydrobenzothiophenyl, benzoxazolinone, etc. Exemplary 6-membered heterocyclic groups fused to an aryl ring (also referred to herein as a 6,6-bicyclic heterocycle) include, but are not limited to, tetrahydroquinolinyl, tetrahydroisoquinolinyl, etc.

[0093] Unless otherwise specified, the term "aryl" or "aromatic ring group" refers to a monocyclic, bicyclic, or tricyclic aromatic carbocyclic system containing 6-16 carbon atoms, or 6-14 carbon atoms, or 6-12 carbon atoms, preferably 6-10 carbon atoms. The term "aryl" may be used interchangeably with the term "aromatic ring." Examples of aryl groups may include, but are not limited to, phenyl, naphthyl, anthraceneyl, phenanthryl, or pyrene.

[0094] Unless otherwise specified, the term "arylalkylene" means aryl-alkylene-*, which is attached to other groups or segments by an alkylene group.

[0095] Unless otherwise specified, the term "heteroaryl" or "heteroary cycloyl" refers to an aromatic monocyclic or polycyclic system containing a 5-14 member structure, or preferably a 5-10 member structure, or preferably a 5-8 member structure, more preferably a 5-6 member structure, wherein one, two, three or more ring atoms are heteroatoms and the remaining atoms are carbon atoms, the heteroatoms being independently selected from O, N or S, and the number of heteroatoms is preferably one, two or three. Examples of heteroaryl groups include, but are not limited to, furanyl, thiophene, oxazolyl, thiazolyl, isoxazolyl, oxadiazolyl, thiazolyl, pyrroleyl, pyrazolyl, imidazoleyl, triazolyl, tetrazolyl, pyridyl, pyrimidinyl, pyrazinyl, thiodiazolyl, triazinyl, phthalazinyl, quinolinyl, isoquinolinyl, pteridinyl, purine, indoleyl, isoindoleyl, indazoleyl, benzofuranyl, benzothiophene, benzopyridyl, benzopyrimidinyl, and benzo[] Pyrazinyl, benzimidazolyl, benziphthalazolyl, pyrrolo[2,3-b]pyridyl, imidazo[1,2-a]pyridyl, pyrazolo[1,5-a]pyridyl, pyrazolo[1,5-a]pyrimidinyl, imidazo[1,2-b]pyridazinyl, [1,2,4]triazolo[4,3-b]pyridazinyl, [1,2,4]triazolo[1,5-a]pyrimidinyl, [1,2,4]triazolo[1,5-a]pyridyl, etc.

[0096] Unless otherwise specified, the term "heteroarylalkylene" means heteroaryl-alkylene-*, which is attached to other groups or segments by an alkylene group.

[0097] Unless otherwise specified, the terms "pharmaceutically acceptable salt" or "medicinal salt" refer to a salt that, within the bounds of reasonable medical judgment, is suitable for contact with the tissues of mammals, particularly humans, without excessive toxicity, irritation, allergic reactions, etc., and is proportionate to a reasonable benefit / risk ratio. Medically acceptable salts of amines, carboxylic acids, and other types of compounds are well known in the art. The salts can be prepared in situ during the final separation and purification of the compounds of this invention, or solely by reacting a free base or free acid with a suitable reagent.

[0098] Unless otherwise specified, the term "isotope derivative" refers to compounds of the present invention that can exist in an isotopically traced or enriched form, containing one or more atoms whose atomic weights or mass numbers differ from the atomic weights or mass numbers of the most abundant atoms found in nature. Isotopes can be radioactive or non-radioactive. Commonly used isotopes for isotopic labeling are: hydrogen isotopes, 2 H and 3 H; Carbon isotopes: 13 C and 14 C; Chlorine isotopes: 35 Cl and 37 Cl; Fluorine isotopes: 18 F; Iodine isotopes: 123 I and 125 I; Nitrogen isotopes: 13 N and 15 N; oxygen isotopes: 15 O, 17 O and 18 O and sulfur isotopes 35 S. These isotope-labeled compounds can be used to study the distribution of pharmaceutical molecules in tissues. Especially 2 H and 13 C, because they are easy to label and convenient to detect, are more widely used. Some heavy isotopes, such as deuterium (… 2 Substitution with H can enhance metabolic stability and prolong the half-life, thereby achieving the goal of reducing dosage and providing therapeutic advantages. Isotope-labeled compounds are generally synthesized from labeled starting materials using known synthetic techniques, just like non-isotope-labeled compounds.

[0099] Unless otherwise specified, the terms "solvent" or "solvent compound" refer to the physical association of the compound of the present invention with one or more solvent molecules (organic or inorganic). This physical association includes hydrogen bonding. In some cases, such as when one or more solvent molecules are incorporated into the crystal lattice of a crystalline solid, the solvate can be separated. The solvent molecules in the solvate may be present in a regular and / or disordered arrangement. The solvate may contain stoichiometric or non-stoichiometric solvent molecules. "Solvent compound" encompasses both solution phases and separable solvates. Exemplary solvates include, but are not limited to, hydrates, ethanol compounds, methanol compounds, and isopropanol compounds. Solvation methods are well known in the art.

[0100] Unless otherwise specified, the term "stereoisomer" refers to compounds having the same chemical structure but with different spatial arrangements of atoms or groups. Stereoisomers include enantiomers, diastereomers, conformational isomers (rotational isomers), geometric isomers (cis / trans) isomers, and inhibited isomers. Any mixture of stereoisomers obtained can be separated into pure or substantially pure geometric isomers, enantiomers, and diastereomers based on differences in the physicochemical properties of the components, for example, by chromatography and / or fractional crystallization.

[0101] Unless otherwise specified, the term "tautomer" refers to structural isomers with different energies that can interconvert through a low energy barrier. If tautomerism is possible (e.g., in solution), chemical equilibrium can be achieved in the tautomer. For example, proton tautomers (also called proton transfer tautomers) involve interconversions via proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers involve interconversions via the rearrangement of some bonding electrons.

[0102] Unless otherwise indicated, the structural formulas described in this invention include all isomers (e.g., enantiomers, diastereomers, and geometric isomers (or conformational isomers)): for example, R and S configurations containing an asymmetric center, (Z) and (E) isomers of double bonds, and (Z) and (E) conformational isomers. Therefore, any single stereochemical isomer of the compounds of this invention, or its enantiomers, diastereomers, or mixtures of geometric isomers (or conformational isomers), is within the scope of this invention.

[0103] Unless otherwise specified, the term "prodrug" refers to a drug that is converted into a parent drug in vivo. Prodrugs are generally useful because they can improve certain, undesirable physical or biological properties. Physical properties are generally related to solubility (excessive or insufficient lipid or water solubility) or stability, while problematic biological properties include metabolism that is too rapid or poor bioavailability, which may itself be related to physicochemical properties. For example, they can be bioavailable orally, whereas the parent drug cannot. Prodrugs also have improved solubility in pharmaceutical compositions compared to the parent drug. An example of a prodrug, but not limited thereto, can be any compound of the present invention administered as an ester ("prodrug") to facilitate transmembrane transport, where water solubility is detrimental to migration but beneficial once inside the cell, and which is subsequently metabolized and hydrolyzed into a carboxylic acid, the active entity. Another example of a prodrug can be a short peptide (polyamino acid) bound to an acid group, where the peptide is metabolized to exhibit the active moiety.

[0104] It should be understood that, within the scope of this invention, the above-described technical features of this invention and the technical features specifically described below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be described in detail here.

[0105] The general method for preparing the dicyclic compound described in this invention is as follows:

[0106] The dicyclic compound in this invention can be synthesized using the following synthetic route.

[0107]

[0108] Among them, G1 and G2 may be the same or different, and are independently selected from: H and reactive functional groups;

[0109] G a G b G c Whether the groups are the same or different, they are each independently selected from: H, reactive functional groups;

[0110] L1' is L1 excluding G1 and G... b The corresponding residue group;

[0111] L2' is L2 excluding G2 and G c The corresponding residue group;

[0112] L3' is L3 minus G a The corresponding residue group.

[0113] Preferably, the reactive functional group is selected from halogen, hydroxyl, amino, carboxyl, amide, acyl halide, ester, halogen, hydroxyl, and alkoxy.

[0114] In the method, steps 1 and 2 can be carried out in one reaction when A1 and A2 are the same substituents; in addition, protecting groups can be introduced / removed as needed, the reaction order of each step can be changed, or a compound of formula (I) can be transformed into a compound of formula (I).

[0115] In addition, the compound shown in formula (I-2) can be synthesized by the following synthetic route.

[0116]

[0117] Wherein, PG may be the same or different, and is independently selected from amino protecting groups;

[0118] X a X b X c Independently selected from halogens, OH, C 1-6 Alkoxy, -O benzyl.

[0119] Preferably, the PG is selected from -Boc.

[0120] In the method, steps 1 and 2 can be carried out in one reaction when A1 and A2 are the same substituents; in addition, protecting groups can be introduced / removed as needed, the reaction order of each step can be changed, or a compound represented by one formula (I-2) can be transformed into a compound represented by another formula (I-2).

[0121] Beneficial effects:

[0122] This invention provides a dual-linker compound and its application, wherein the dual-linker compound has two linkers capable of conjugating with antibodies and a linker capable of conjugating with cytotoxic drugs, and the dual-linker compound can conjugate well with antibodies and has very good uniformity in DAR value. Attached Figure Description

[0123] Figure 1 The LC-MS spectrum of the product conjugated with TBD1 and the antibody Trastuzumab;

[0124] Figure 2 for Figure 1 A magnified view of a portion of the image. Detailed Implementation

[0125] The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments, unless otherwise specified, are generally performed under conventional conditions or as recommended by the manufacturer. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as are familiar to those skilled in the art. Furthermore, any methods and materials similar to or equivalent to those described herein may be applied to the methods of the present invention. The preferred embodiments and materials shown herein are for illustrative purposes only.

[0126] The specific synthetic schemes for the C-1 and C-2 carboxylic acids used in this article are as follows:

[0127] C-1 synthesis route:

[0128]

[0129] C-1-1:

[0130]

[0131] Prepare a potassium hydroxide solution by slowly adding 78g KOH to 150ml of water. After dissolving, cool the solution and add it to a 1L single-necked reaction flask. Weigh 15g of 2-aminobenzothiazol-6-carboxylic acid and add it to the reaction flask. Stir until evenly dispersed and stir for 4 hours in the dark at 100℃-105℃ to obtain a milky yellow suspension. Cool the reaction mixture to rt and then cool it to 0-5℃ in an ice bath. Add concentrated HCl to the cooled solution, which will produce fumes. Stir slowly until acidic (pH=3-4), and a large amount of solid will precipitate. Keep the yellowish-brown suspension warm and stir for 15 minutes. Filter to obtain a filter cake, and wash the filter cake several times with deionized water. The crude 4-amino-3-mercaptobenzoic acid salt is a slightly moist light olive-greenish-gray solid. Transfer it to another single-necked flask (1000mL) and carry out the next cyclization reaction under N2 (be careful to avoid light). Calculate the yield as 100%.

[0132] C-1-2:

[0133]

[0134] The solid obtained in the previous step was first added to 50 mL of EtOH under dim lighting and stirred until homogeneous. Then, it was neutralized with a 2 M potassium hydroxide solution to pH 7-8 (pH paper test). Ethanol (100 mL) was added to the resulting light olive green solution, followed by potassium ethyl xanthate (18.57 g). The mixture was stirred at 100-105 °C for 3 h to obtain a fine suspension of pale olive green to yellow (deep orange solution with fine greenish-black precipitate). The reaction mixture was concentrated to obtain a viscous, deep orange residue, which was then cooled to room temperature and used directly in the next reaction step.

[0135] C-1-3:

[0136]

[0137] Add the materials from the previous step to a 1L single-necked flask, add 250ml of DMF, stir until dissolved, then add 32.01g of potassium carbonate, stir for 10 minutes, and then slowly add iodoethane dropwise to the reaction solution. Purge with nitrogen three times, stir overnight at room temperature, and monitor the TLC to ensure the C-1-2 reaction is complete. Quench the reaction solution slowly by adding 500ml of water, extract with EA (2*300ml), concentrate the organic phase, and elute with silica gel using a PE:EA ratio of 10:1 to obtain 14g of white solid. MS-ESI calculated value [M+H] + 267, the actual measured value is 267.

[0138] C-1-4:

[0139]

[0140] Weigh 13g of C-1-3 and add it to a 1L reaction flask. Add 72ml of MeOH and 48ml of THF and stir until dissolved. Dissolve 3.49g of LiOH in 24ml of water and slowly add it dropwise to the reaction solution. Stir at room temperature for 3 hours. Monitor the reaction of C-1-3 by TLC to ensure complete reaction. After the reaction is complete, adjust the pH to 5-6 with 1M dilute hydrochloric acid. Slowly add the reaction solution to 300ml of water, then extract with EA (3 x 200ml), wash with NaCl water, dry the organic phase with anhydrous sodium sulfate, and concentrate the organic phase to obtain 9g of white solid. MS-ESI calculated value [M+H] + 239, the actual measured value is 239.

[0141] C-1:

[0142]

[0143] Weigh 9g of C-1-4 and 10.39g of ammonium heptamolybdate into a 1L reaction flask. After adding 300ml of EtOH and stirring in an ice bath for 10min, slowly add 150ml of 30% hydrogen peroxide dropwise to the reaction solution while maintaining the temperature at 0-5℃. After the addition is complete, return to room temperature and stir for 12h. TLC monitoring shows that the C-1-4 reaction is complete. After the reaction is complete, slowly pour the reaction solution into 500ml of water to quench it, then wash with EA (3*300ml), NaCl, and water. Dry the organic phase with anhydrous sodium sulfate. After concentrating half the solvent, add 100ml of n-hexane to the organic phase to precipitate the product. Filter and dry to obtain 8g of white solid. The product has poor thermal stability (easily damaged when dried at 80℃). The drying temperature is 50℃. MS-ESI calculated value [M+H] + 271, the actual measured value is 271.

[0144] C-2 synthesis route:

[0145]

[0146] C-2-1:

[0147]

[0148] Add 5g of the raw material to 50ml of EtOH and stir until homogeneous. Then add 100mL of ethanol to the resulting reaction system, followed by 7.19g of potassium ethyl xanthate. Stir at 100-105℃ for 3 hours to obtain a pale olive-green to yellow fine suspension (deep orange solution with fine greenish-black precipitate). Concentrate the reaction mixture to obtain a viscous, deep orange residue. Cool to room temperature and use directly in the next reaction step. MS-ESI calculated value [M+H] + 209, the actual measured value is 209.

[0149] C-2-2:

[0150]

[0151] Add the materials from the previous step to a 500ml single-necked flask, add 200ml of DMF, stir until dissolved, then add 12.41g of potassium carbonate, stir for 10 minutes, and then slowly add 4.79ml of iodoethane dropwise to the reaction solution. Purge with nitrogen three times, stir overnight at room temperature, and monitor the C-2-1 reaction by TLC to ensure completeness. Quench the reaction solution slowly in 300ml of water, extract with Ea (2 x 150ml), concentrate the organic phase, and elute with silica gel using a PE:EA ratio of 10:1 to obtain 5g of white solid. MS-ESI calculated value [M+H] + 237, the actual measured value is 237.

[0152] C-2-3:

[0153]

[0154] Weigh 5g of C-2-2 and add it to a 500ml reaction flask. Add 24ml of MeOH and 16ml of THF and stir until dissolved. Dissolve 1.51g of LiOH in 8ml of water and slowly add it dropwise to the reaction solution. Stir at room temperature for 3 hours. Take a sample and monitor the reaction by TLC to ensure complete reaction of C-2-2. After the reaction is complete, adjust the pH to 5-6 with 1M dilute hydrochloric acid. Slowly add the reaction solution to 200ml of water, then extract with Ea (3 x 150ml), wash with NaCl water, dry the organic phase with anhydrous sodium sulfate, and concentrate the organic phase to obtain 4g of white solid. MS-ESI calculated value [M+H] + 223, the actual measured value is 223.

[0155] C-2:

[0156]

[0157] Weigh 4g of C-2-3 and 4.95g of ammonium heptamolybdate into a 250mL reaction flask. After adding 30mL of EtOH and stirring in an ice bath for 10min, slowly add 15mL of 30% hydrogen peroxide dropwise to the reaction solution while maintaining the temperature at 0-5℃. After the addition is complete, return to room temperature and stir for 12h. TLC monitoring shows that the C-2-3 reaction is complete. After the reaction is complete, slowly pour the reaction solution into 100mL of water to quench it, then wash with EA (3*50mL), NaCl, and water. Dry the organic phase with anhydrous sodium sulfate. After concentrating half the solvent, add 100mL of n-hexane to the organic phase to precipitate the product. Filter and dry to obtain 3g of white solid. The product has poor thermal stability (easily damaged when dried at 80℃). The drying temperature is 50℃. MS-ESI calculated value [M+H] + 255, the actual measured value is 255.

[0158] Example 1: Synthesis of compound TBD1:

[0159] Synthesis route:

[0160]

[0161] Experimental Step 1: Synthesis of TBD1-1

[0162]

[0163] Diethylenetriamine (5 g, 48.46 mmol) was added to DMF (120 mL) and stirred at room temperature. Tert-butylphenyl carbonate (20.71 g, 106.62 mmol) was then added, and the mixture was stirred at room temperature for 12 h under a dry atmosphere. The solvent DMF was evaporated under reduced pressure. The residue was washed with 1 N NaOH aq and brine, then dissolved in chloroform to remove insoluble matter, and the organic phase was filtered off. The organic phase was evaporated to dryness to obtain crude TBD1-1, which was directly used in the next reaction. MS-ESI calculated value [M+H] + 304, the actual measured value is 304.

[0164] Experimental Step 2: Synthesis of TBD1-2

[0165]

[0166] The crude compound TBD1-1 obtained in the previous step was dissolved in MeCN (120 mL), followed by the addition of ethyl bromoacetate (10.52 g, 62.99 mmol) and sodium carbonate (11.30 g, 106.59 mmol). The reaction mixture was stirred at 0 °C for 30 min, and then at room temperature under a dry atmosphere for 24 h. After evaporating the solvent under reduced pressure, the residue was dissolved in dichloromethane and washed with 1 N NaOH aq and brine. The organic layer was dehydrated with anhydrous Na2SO4, filtered, and evaporated. The residue was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 0-100%) to give TBD1-2 (13 g, 64.26%). MS-ESI calculated value [M+H] + 340, the actual measured value is 340.

[0167] Experimental Step 3: Synthesis of TBD1-3

[0168]

[0169] Compound TBD1-2 (13 g, 33.38 mmol) was added to HCl in dioxane (4 M, 100 mL) and stirred at room temperature for 3 h. The reaction mixture was cooled to 0 °C, and 50 mL of 1 N NaOH solution was added. The reaction system was diluted with 150 mL of ethyl acetate. The organic phase was washed with 100 mL of 1 N NaOH solution and 3 times of water (100 mL × 3). The mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the residue, yielding 5 g of TBD1-3 product (yield: 79.15%). MS-ESI calculated value [M+H] + 190, the actual measured value is 190.

[0170] Experimental Step 4: Synthesis of TBD1-4

[0171]

[0172] Carboxylic acid c-1 (300 mg, 1.11 mmol) was dissolved in 10 mL of DMF, cooled to 0-5 °C, and HATU (546.99 mg, 1.44 mmol) was added. Stirring continued for 25 min, followed by the dropwise addition of DIPEA (0.58 mL, 3.32 mmol). The reaction was maintained at this temperature for 0.5 h, and then TBD1-3 (104.64 mg, 0.55 mmol) was added. Stirring continued for 1 h, and the reaction was allowed to proceed at room temperature. A sample was taken and spotted onto a TLC plate after 1 h of reaction. The reactant carboxylic acid was then completely reacted. The reaction solution was directly purified using reverse-phase medium-pressure (C18) chromatography (Water (0.1%): ACN = 10%–75%) for 60 min to obtain product TBD1-4400 mg, yield: 51.99%. MS-ESI calculated value [M+H] + 696, the actual measured value is 696.

[0173] Experimental Step 5: Synthesis of TBD1

[0174]

[0175] The TBD1-4 product (300 mg, 0.43 mmol) was dissolved in 10 mL of ethanol. After cooling to 0 °C, 0.33 mL of 2N sodium hydroxide aqueous solution (0.65 mmol) was added dropwise to the reaction solution. The mixture was heated to room temperature and stirred for 2 h. The reaction solution was then cooled to 0 °C and acidified to pH 2 with 2N dilute hydrochloric acid aqueous solution. The reaction system was diluted with ethyl acetate (150 mL). The organic phase was washed successively with 2N dilute hydrochloric acid (100 mL × 1) and water (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the residue, yielding TBD1 product (200 mg) in 70.2% yield. MS-ESI calculated value [M+H] + 668, the actual measured value is 668.

[0176] Examples 2-4

[0177] Referring to the synthetic route in Example 1, using the common intermediate TBD1-3 and selecting the corresponding carboxylic acid compounds, a series of specific compounds of the present invention were prepared (see Table 1).

[0178] Table 1

[0179]

[0180] Example 5: Synthesis of TBD5

[0181] Synthesis route:

[0182]

[0183] Experimental Step 1: Synthesis of TBD5-1

[0184]

[0185] Carboxylic acid C-2 (150 mg, 0.59 mmol) was dissolved in 10 mL of DMF, cooled to 0-5 °C, and HATU (290.5 mg, 0.76 mmol) was added. Stirring continued for 25 min, followed by the dropwise addition of DIPEA (0.3 mL, 1.77 mmol). The reaction was maintained at this temperature for 0.5 h, and then TBD1-3 (111.22 mg, 0.59 mmol) was added. Stirring continued for 1 h, and the reaction was allowed to proceed at room temperature. A sample was taken and spotted onto a TLC plate after 1 h of reaction. The reactant carboxylic acid was then completely reacted. The reaction solution was directly purified using reverse-phase medium-pressure (C18) chromatography (Water (0.1%): ACN = 10%–75%) for 60 min to obtain product TBD5-1160 mg, yield: 63.84%. MS-ESI calculated value [M+H] + 426, the actual measured value is 426.

[0186] Experimental Step 2: Synthesis of TBD5-2

[0187]

[0188] Carboxylic acid c-1 (101.78 mg, 0.38 mmol) was dissolved in 10 mL of DMF, cooled to 0-5 °C, and HATU (189 mg, 0.46 mmol) was added. Stirring continued for 25 min, followed by the dropwise addition of DIPEA (0.24 mL, 1.41 mmol). The reaction was maintained at this temperature for 0.5 h, and then TBD5-1 (160 mg, 0.38 mmol) was added. Stirring continued for 1 h, and the reaction was allowed to proceed at room temperature. A sample was taken and spotted onto a TLC plate after 1 h of reaction. The reactant carboxylic acid was then completely reacted. The reaction solution was directly purified using reverse-phase medium-pressure (C18) chromatography (Water (0.1%): ACN = 10%–75%) for 60 min to obtain product TBD5-2 158 mg, yield: 61.95%. MS-ESI calculated value [M+H] + 680, the actual measured value is 680.

[0189] Experimental Step 3: Synthesis of TBD5

[0190]

[0191] The TBD5-2 product (158 mg, 0.23 mmol) was dissolved in 10 mL of ethanol. After cooling to 0 °C, 0.34 mL of 2N sodium hydroxide aqueous solution (0.68 mmol) was added dropwise to the reaction solution. The mixture was heated to room temperature and stirred for 2 h. The reaction solution was then cooled to 0 °C and acidified to pH 2 with 2N dilute hydrochloric acid aqueous solution. The reaction system was diluted with ethyl acetate (150 mL). The organic phase was washed successively with 2N dilute hydrochloric acid (100 mL × 1) and water (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the residue, yielding the TBD5 product (130 mg), with a yield of 85.82%. MS-ESI calculated value [M+H] + 652, the actual measured value is 652.

[0192] Examples 6-8

[0193] Referring to the synthetic route in Example 5, using the common intermediate TBD1-3 and selecting the corresponding carboxylic acid compounds, a series of specific compounds of the present invention were prepared (see Table 2).

[0194] Table 2

[0195]

[0196]

[0197] Example of antibody conjugation effect preparation:

[0198] Preparation Example 1

[0199] The following antibodies can be prepared using conventional antibody methods. For example, after constructing a vector, they can be transfected into eukaryotic cells such as HEK293 cells (Life Technologies Cat No. 11625019) and purified for expression.

[0200] Preparation Example 1.1 The following is the sequence of Trastuzumab:

[0201] Light chain (SEQ ID NO:1)

[0202]

[0203] Heavy chain (SEQ ID NO:2)

[0204]

[0205] Preparation Example 1.2 The following is the sequence of the hRS7 antibody (Sacituzumab).

[0206] Light chain (SEQ ID NO:3)

[0207]

[0208] Heavy chain (SEQ ID NO:4)

[0209]

[0210] Example 9

[0211] At 37°C, a prepared aqueous solution of tris(2-carboxyethyl)phosphine (5mM, 0.355mL, 1.77μmol) was added to a PB buffer solution of the antibody Trastuzumab (0.04M PB buffer solution with pH=7.0; 35mg, 15mg / mL, 0.236μmol). The solution was placed in a water bath and shaken at 37°C for 3 hours. The reaction was then stopped. The reaction solution was cooled to 25°C in a water bath, diluted to 5.0mg / mL, and 2.0mL of the solution was taken out and used for further reaction.

[0212] Compound TBD1 (1.64 mg, 2.36 μmol) was dissolved in 0.10 mL of DMSO and added to the above 2.0 mL solution. The mixture was placed in a water bath and shaken at 25 °C for 3 hours, after which the reaction was stopped. The reaction solution was purified by desalting using a Sephadex G25 gel column (elution phase: 0.04 M PB buffer aqueous solution at pH 7.0 containing 0.002 M EDTA) to obtain a PB solution of the exemplary product (7.00 mg / mL, 15 mg), which was stored frozen at 4 °C.

[0213] Example 10

[0214] At 25°C, a prepared aqueous solution of tris(2-carboxyethyl)phosphine (5 mM, 0.156 mL, 0.780 μmol) was added to a PB buffer solution of antibody Sacituzumab (0.04 M PB buffer solution of pH = 7.0; 35 mg, 11 mg / mL, 0.236 μmol). The solution was placed in a water bath shaker and shaken at 25°C for 3 hours. The reaction was then stopped. The reaction solution was diluted to 5.0 mg / mL.

[0215] Compound TBD1 (1.64 mg, 2.36 μmol) was dissolved in 0.10 mL DMSO and added to the above solution. The mixture was placed in a water bath and shaken at 25 °C for 3 hours, after which the reaction was stopped. The reaction solution was purified by desalting using a Sephadex G25 gel column (elution phase: 0.02 M histidine buffer aqueous solution at pH = 5.5) to obtain a histidine solution of the exemplary product ADC-II-3 (4.1 mg / mL, 22 mg), which was stored at 4 °C.

[0216] Referring to the synthesis schemes of Examples 9 and 10, the conjugation results of TBD3, TBD5, TBD7 with antibodies are shown in Table 3 below.

[0217] Table 3

[0218] Example Molecular code Antibody DAR value Example 9 TBD1 Antibody Trastuzumab 4.15 Example 10 TBD1 Antibody Sacituzumab 4.02 Example 11 TBD3 Antibody Trastuzumab 3.95 Example 12 TBD3 Antibody Sacituzumab 3.91 Example 13 TBD5 Antibody Trastuzumab 4.02 Example 14 TBD5 Antibody Sacituzumab 4.05 Example 15 TBD7 Antibody Trastuzumab 3.92 Example 16 TBD7 Antibody Sacituzumab 3.90

[0219] The experimental results show that the designed dual-linker compounds can be well conjugated with the antibody. The DAR values ​​of TBD1, TBD3, TBD5, and TBD7 conjugated with the antibody are all between 3.9 and 4.15, demonstrating very good homogeneity.

[0220] The embodiments of the present invention have been described above. However, the present invention is not limited to the above embodiments. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A dual-linker compound or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates, or isotopically labeled analogs thereof, said dual-linker compound having the structure shown in formula (I): in, A1 and A2 may be the same or different, and are selected independently from: R1 is independently selected from: C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 1-6 Alkylamine alkyl, C 1-6 Alkoxyalkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene; Cy is independently selected from: substituted or unsubstituted C 3-6 Cycloalkyl, substituted or unsubstituted 3-6 membered heterocyclic groups, substituted or unsubstituted aryl groups, substituted or unsubstituted heteroaryl groups; L A1 L A2 Same or different, independently selected from: single bond, substituted or unsubstituted C 1-12 Subchain hydrocarbon groups; L1, L2, and L3 are linking groups; M represents a functional group or leaving group that can be linked to cytotoxic drugs.

2. The dual-linker compound according to claim 1, or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates, or isotopically labeled analogs thereof, characterized in that, Cy is independently selected from one of the following structures: Where x is the same or different, and is independently selected from CR a Or N, and L A1 The connected x is selected from C; R a Whether the same or different, they are independently selected from hydrogen, deuterium, and carbon. 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 1-6 alkylamine group, C 1-6 Alkoxy, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene; Ring A represents an unsubstituted or substituted aromatic ring, or a substituted or unsubstituted heteroaromatic ring; Y is selected from O, S, NR Y , where R Y Independently selected from hydrogen, deuterium, and C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene; "*1" indicates the connection site with SO2R1; Indicates with L A1 Connection sites; L A1 L A2 Same or different, independently selected from: single bond, substituted or unsubstituted C 1-8 Subchain hydrocarbon group, the C 1-8 The subchain hydrocarbon group may optionally contain 1, 2 or 3 carbon-carbon double bonds or carbon-carbon triple bonds; Preferred, Choose one of the following structures: Where y is the same or different, it is independently selected from CR. b Or N, and L A1 The connected y is selected from C; R b Whether the same or different, they are independently selected from hydrogen, deuterium, and carbon. 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 1-6 alkylamine group, C 1-6 Alkoxy, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene; L A1 L A2 Whether the bonds are the same or different, they are independently selected from a single bond and one of the following structures: a is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12; b is selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

3. The dual-linker compound according to claim 1 or 2, or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates, or isotopically labeled analogs thereof, characterized in that, Cy is independently selected from one of the following structures: Preferably, Cy is independently selected from one of the following structures:

4. The dual-linker compound according to claim 1, or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates, or isotopically labeled analogs thereof, characterized in that, L1 and L2 may be the same or different, and can be selected independently from: -L a C 1-12 Alkylene L b -, the C 1-12 The alkylene group may have an intercalation between any two C and C groups selected from -O-, -S-, or -NR. L -, -CO-, -COO-, -OCO-, -OCOO-, -CONR L -、-NR L CO-、-OCONR L -、-NR L COO-、-NR L CONR L -、-NR L SO-、-SONR L -、-NR L S(O)2-、-S(O)2NR L - one of the groups; L a Whether the same or different, it is independently selected from single bonds, -CO-, -SO-, -S(O)2-, -OCO-, -NR L CO-;L a It is attached to the N atom in the middle of the compound; L b Same or different, independently selected from single bond, -O-, -S-, -NR L -, -CO-, -COO-, -OCO-, -OCOO-, -CONR L -、-NR L CO-、-OCONR L -、-NR L COO-、-NR L CONR L -、-NR L SO-、-SONR L -、-NR L S(O)2-、-S(O)2NR L -;L b Connected to L A1 or L A2 superior; R L Independently selected from hydrogen, deuterium, and C 1-6 alkyl; L3 is selected from: -L c C 1-12 alkylene-, the C 1-12 The alkylene group may have an intercalation between any two C and C groups selected from -O-, -S-, or -NR. L -, -CO-, -COO-, -OCO-, -OCOO-, -CONR L -、-NR L CO-、-OCONR L -、-NR L COO-、-NR L CONR L -、-NR L SO-、-SONR L -、-NR L S(O)2-、-S(O)2NR L - one of the groups; L c Independently selected from single bonds, -CO-, -SO-, -S(O)2-, -OCO-, -NR L CO-;L c It is attached to the N atom in the middle of the compound; R L Independently selected from hydrogen, deuterium, and C 1-6 alkyl.

5. The dual-linker compound according to claim 1, or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates, or isotopically labeled analogs thereof, characterized in that, M is selected from -OR M , -SR M , -N(R M )2, -COOR M , -CON(R M )2, -SO(R M )-, -S(O)2(R M )2; R M Independently selected from hydrogen, deuterium, and C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Deuterated alkyl, C 1-6 Alkylamine alkyl, C 1-6 Alkoxyalkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 3-6 Cycloalkyl, 3-6 membered heterocyclic, aryl, arylalkylene, heteroaryl, heteroarylalkylene.

6. The dual-linker compound according to claim 5, or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates, or isotopically labeled analogs thereof, characterized in that, The dual-connector compound has the structure shown in formula (I-1): Among them, A1, A2, L a L b n, m, and M are as described in claim 5; Alternatively, the dual-connector compound may have the structure shown in formula (I-2): Wherein, A1, A2, n, m, and M are as described in claim 5.

7. The dual-linker compound according to claim 1, or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates, or isotopically labeled analogs thereof, characterized in that, The dual-connector compounds are selected from:

8. Use of the dual linker compound according to any one of claims 1-7 or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, hydrates, solvates or isotopically labeled analogs thereof, for the manufacture of drug-linker compounds or for the manufacture of antibody-drug conjugates.

9. A drug-dual linker compound or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, prodrugs, hydrates, solvates, or isotopically labeled analogs thereof, said drug-dual linker compound having the structure shown in formula (II): in, D is the cytotoxic drug fragment; M' is the residue of M after the reaction with the cytotoxic drug; A1, A2, L1, L2, L3, and M are as described in any one of claims 1-7.

10. A pharmaceutical composition comprising the drug-dual linker compound of claim 9 or its stereoisomers, geometric isomers, tautomers, pharmaceutical salts, prodrugs, hydrates, solvates or isotopically labeled analogs thereof.

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