CN115260195B - EGFR degrading agent - Google Patents

EGFR degrading agent Download PDF

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CN115260195B
CN115260195B CN202210451380.5A CN202210451380A CN115260195B CN 115260195 B CN115260195 B CN 115260195B CN 202210451380 A CN202210451380 A CN 202210451380A CN 115260195 B CN115260195 B CN 115260195B
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cancer
methyl
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CN115260195A (en
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潘建峰
孙大庆
闫琪
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Shanghai Qilu Pharmaceutical Research and Development Centre Ltd
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/10—Spiro-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D519/00—Heterocyclic compounds containing more than one system of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring system not provided for in groups C07D453/00 or C07D455/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic System
    • C07F9/02—Phosphorus compounds
    • C07F9/547—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom
    • C07F9/6561—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom containing systems of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring or ring system, with or without other non-condensed hetero rings

Abstract

The present invention provides a novel compound which can degrade EGFR protein, a pharmaceutical composition containing the compound, a useful intermediate for preparing the compound and a method for treating a cell proliferative disease such as cancer using the compound of the present invention.

Description

EGFR degrading agent
Technical Field
The present invention is in the field of pharmaceutical chemistry, and in particular relates to a novel class of compounds that degrade EGFR proteins, pharmaceutical compositions containing the compounds, useful intermediates for preparing the compounds, and methods of treating cell proliferative disorders, such as cancer, using the compounds of the present invention.
Background
EGFR, the EGFR receptor (epidermal growth factor receptor), is widely distributed on the cell surface of mammalian epithelial cells, fibroblasts, glial cells, and the like. The EGFR signaling pathway plays an important role in physiological processes such as cell growth, proliferation and differentiation. EGFR mutations are also one of the most common types of mutations in NSCLC patients, and can account for 40% to 50% of Asian populations in particular. EGFR has therefore been one of the hottest targets in the field of drug development.
Currently, EGFR inhibitors on the market are divided into the first, second and third generation. The first generation is reversible targeted drugs such as gefitinib, erlotinib, and icotinib. The second generation is irreversible targeted drugs such as afatinib and dacatinib. Although the first and second generation targeting drugs have remarkable curative effects, most patients can have drug resistance after using the drugs for 1-2 years. Of the EGFR inhibitor resistant patients, 50% were associated with the T790M mutation. The third generation EGFR targeting drug, namely the Ornitinib, can overcome tumor resistance caused by T790M mutation, and brings better survival benefit to more lung cancer patients. However, the third generation of targeting drugs inevitably generates drug resistance, and the reason for drug resistance is mainly C797S mutation. The C797S mutation is manifested as a mutation of a cysteine residue to serine, which disrupts EGFR protein binding to third generation targeting drugs, thereby failing to prevent EGFR protein phosphorylation and downstream signaling pathway activation. At present, no mature treatment means exists for the treatment of the Ornitinib drug resistance, the clinical requirement is urgent, and the invention is based on solving the problem.
Disclosure of Invention
The invention aims to provide a novel compound capable of degrading EGFR protein, a pharmaceutical composition containing the compound, a useful intermediate for preparing the compound and application of the compound in preparing medicines for treating cancers.
The invention provides a compound shown as a formula (I-1) or pharmaceutically acceptable salt thereof,
Figure BDA0003618772920000011
wherein R is 1 Selected from H, C 1-4 Alkyl, -C (O) OR 5 、-(CH 2 )s-OR 6 、-(CH 2 )t-C(O)NR 7 R 8 、-C(O)R 9 、-S(O) 2 R 10 Or 5-7 membered heterocycloalkyl, and said C 1-4 Alkyl or 5-7 membered heterocycloalkyl optionally substituted with one or more R 11 The groups are substituted.
X 1 Selected from-C (=o) or N;
X 2 selected from N or CH;
X 3 selected from N or C;
X 1 and X 3 Connected with each other
Figure BDA0003618772920000021
Represents a single bond or a double bond;
L 1 is C 1-4 Alkylene group, wherein said C 1-4 The alkylene group may be further optionally substituted with one or more U groups selected from O, S, NH or NR ua Wherein R is ua Is C 1-4 An alkyl group;
alternatively, L 1 Is a connecting key;
R 2 h, C of a shape of H, C 1-6 Alkyl or C 3-5 Cycloalkyl;
L 2 is C 1-4 Alkylene group, wherein said C 1-4 The alkylene group may be further optionally substituted with one or more Q groups selected from C.ident. C, O, S, NH, NR qa -NHC (O) -or-C (O) NH-, wherein R qa Is C 1-4 An alkyl group;
alternatively, L 2 Is a connecting key;
ring A is selected from-C 6-10 Aryl or 6-10 membered heteroaryl;
ring B is C 6-10 Aryl, 5-10 membered heteroaryl, 5-6 membered heterocycloalkyl or partially saturated 5-6 membered heterocycloalkyl;
R 3 selected from H, halogen, C 2-4 Alkynyl, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-5 Cycloalkyl, -S (O) 2 R 10 、-P(O)(R 13 )R 14 Or C 6-10 An aryl group;
alternatively, two adjacent R 3 Cyclisation to C 4-6 Cycloalkyl is fused to ring a;
R 4 selected from H, halogen or C 1-4 An alkyl group;
m and n are each independently selected from 0, 1, 2 or 3;
s and t are each independently selected from 0, 1 or 2;
R 5 and R is 6 Each independently selected from H or C 1-4 An alkyl group;
R 7 and R is 8 Each independently selected from H or C 1-4 An alkyl group;
alternatively, R 7 And R is 8 Together with the N atom to which they are attached form a 3-5 membered heterocycloalkyl, said 3-5 membered heterocycloalkyl optionally being further substituted with one or more R 12 Substituted with a group;
R 9 selected from C 1-4 Alkyl or C 3-5 Cycloalkyl;
R 10 is C 1-4 An alkyl group;
R 11 selected from OH, halogen, C 1-4 Alkyl, C 6-10 Aryl or 5-7 membered heteroaryl;
R 12 selected from OH, halogen or C 1-4 An alkyl group;
R 13 and R is 14 Each independently selected from OH or C 1-4 An alkyl group.
In some aspects of the invention, L as described above 1 Selected from-CH 2 -、-CH 2 CH 2 -or-CH 2 NH-; or L 1 Is a connecting key.
In some aspects of the invention, L as described above 2 Selected from-CH 2 -、-CH 2 CH 2 -、-NHCH 2 -、-CH 2 NH-, -NHC (O) -; or L 2 Is a connecting key.
In some aspects of the invention, R is as defined above 2 H.
In some embodiments of the invention, the ring B is C 6-10 Aryl groups.
In some embodiments of the invention, the above-described compounds, or pharmaceutically acceptable salts thereof, are selected from,
Figure BDA0003618772920000031
wherein R is 1 Selected from H, C 1-4 Alkyl, -C (O) OR 5 、-(CH 2 )s-OR 6 、-(CH 2 )t-C(O)NR 7 R 8 、-C(O)R 9 、-S(O) 2 R 10 Or 5-7 membered heterocycloalkyl, and said C 1-4 Alkyl or 5-7 membered heterocycloalkyl optionally substituted with one or more R 11 The groups are substituted.
X 1 Selected from-C (=o) or N;
X 2 selected from N or CH;
X 3 selected from N or C;
X 1 and X 3 Connected with each other
Figure BDA0003618772920000032
Represents a single bond or a double bond;
R 2 is H;
ring A is selected from C 6-10 Aryl or 6-10 membered heteroaryl;
ring B is C 6-10 An aryl group;
R 3 selected from H, halogen, C 2-4 Alkynyl, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-5 Cycloalkyl, -S (O) 2 R 10 、-P(O)(R 13 )R 14 Or C 6-10 An aryl group;
alternatively, two adjacent R 3 Cyclisation to C 4-6 Cycloalkyl is fused to ring a;
R 4 selected from H, halogen or C 1-4 An alkyl group;
m and n are each independently selected from 0, 1, 2 or 3;
s and t are each independently selected from 0, 1 or 2;
R 5 and R is 6 Each independently selected from H or C 1-4 An alkyl group;
R 7 and R is 8 Each independently selected from H or C 1-4 An alkyl group;
alternatively, R 7 And R is 8 Together with the N atom to which it is attachedForming a 3-5 membered heterocycloalkyl, said 3-5 membered heterocycloalkyl optionally being further substituted by one or more R 12 Substituted with a group;
R 9 selected from C 1-4 Alkyl or C 3-5 Cycloalkyl;
R 10 is C 1-4 An alkyl group;
R 11 selected from OH, halogen, C 1-4 Alkyl, C 6-10 Aryl or 5-7 membered heteroaryl;
R 12 Selected from OH, halogen or C 1-4 An alkyl group;
R 13 and R is 14 Each independently selected from OH or C 1-4 An alkyl group.
In some embodiments of the present invention,
Figure BDA0003618772920000041
wherein R is 1 Selected from H, C 1-4 Alkyl, -C (O) OR 5 、-(CH 2 )s-OR 6 、-(CH 2 )t-C(O)NR 7 R 8 、-C(O)R 9 、-S(O) 2 R 10 Or 5-7 membered heterocycloalkyl, and said C 1-4 Alkyl or 5-7 membered heterocycloalkyl optionally substituted with one or more R 11 Substituted with a group;
X 1 selected from-C (=o) or N;
X 2 selected from N or CH;
X 3 selected from N or C;
X 1 and X 3 Connected with each other
Figure BDA0003618772920000042
Represents a single bond or a double bond;
R 2 h, C of a shape of H, C 1-6 Alkyl or C 3-5 Cycloalkyl;
ring A is selected from C 6-10 Aryl or 6-10 membered heteroaryl;
ring B is C 6-10 An aryl group;
R 3 selected from halogen、C 2-4 Alkynyl, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-5 Cycloalkyl, -S (O) 2 R 10 、-P(O)(R 13 )R 14 Or C 6-10 An aryl group;
alternatively, two adjacent R 3 Cyclisation to C 4-6 Cycloalkyl is fused to ring a;
R 4 selected from halogen or C 1-4 An alkyl group;
m and n are each independently selected from 0, 1, 2 or 3;
s and t are each independently selected from 0, 1 or 2;
R 5 and R is 6 Each independently selected from H or C 1-4 An alkyl group;
R 7 and R is 8 Each independently selected from H or C 1-4 An alkyl group;
alternatively, R 7 And R is 8 Together with the N atom to which they are attached form a 3-5 membered heterocycloalkyl, said 3-5 membered heterocycloalkyl optionally being further substituted with one or more R 12 Substituted with a group;
R 9 selected from C 1-4 Alkyl or C 3-5 Cycloalkyl;
R 10 is C 1-4 An alkyl group;
R 11 selected from OH, halogen, C 1-4 Alkyl, C 6-10 Aryl or 5-7 membered heteroaryl;
R 12 Selected from OH, halogen or C 1-4 An alkyl group;
R 13 and R is 14 Each independently selected from OH or C 1-4 An alkyl group.
In some aspects of the invention, R is as defined above 5 Is tert-butyl.
In some aspects of the invention, R is as defined above 6 Methyl, s is 2.
In some aspects of the invention, R is as defined above 7 And R is 8 Each independently selected from H, methyl or ethyl;
alternatively, R 7 And R is 8 Together with the N atom to which it is attached form an azetidinyl group, said azetidinyl groupOptionally further by one or more R 12 Substituted with a group;
R 12 f is the same as F;
t is selected from 0 or 1.
In some aspects of the invention, R is as defined above 9 Selected from methyl or cyclopropyl.
In some aspects of the invention, R is as defined above 10 Selected from methyl or isopropyl.
In some aspects of the invention, R is as defined above 13 And R is 14 Each independently is methyl.
In some aspects of the invention, R is as defined above 1 Selected from H, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, -C (O) OR 5 、-(CH 2 )s-OR 6 、-(CH 2 )t-C(O)NR 7 R 8 、-C(O)R 9 、-S(O) 2 R 10 Or tetrahydropyranyl, and said methyl or isobutyl group is optionally substituted with one or more R 11 Substituted with a group;
R 11 selected from OH, phenyl or pyridyl.
In some aspects of the invention, R is as defined above 1 Selected from H, methyl, ethyl, isopropyl,
Figure BDA0003618772920000051
In some embodiments of the invention, the ring A is selected from
Figure BDA0003618772920000052
Figure BDA0003618772920000053
In some aspects of the invention, R is as defined above 3 Selected from F, cl, ethynyl, methyl, ethyl, methoxy, cyclopropyl, -S (O) 2 CH 3 、-P(O)(CH 3 )CH 3 Or phenyl;
alternatively, two adjacent R' s 3 Cyclisation toCyclopentyl is fused to ring a, which is as defined in any one of claims 1 to 9; m is selected from 0, 1 or 2.
In some aspects of the invention, the structural units described above
Figure BDA0003618772920000061
Selected from the group consisting of
Figure BDA0003618772920000062
Figure BDA0003618772920000063
In some embodiments of the invention, the ring B is phenyl.
In some aspects of the invention, R is as defined above 4 Selected from Br or methyl; n is selected from 0 or 1.
In some aspects of the invention, the structural units described above
Figure BDA0003618772920000064
Selected from->
Figure BDA0003618772920000065
In some embodiments of the invention, the above compound, or a pharmaceutically acceptable salt thereof, is selected from the group consisting of:
Figure BDA0003618772920000066
wherein R is 1 、R 3 、R 4 、X 1 、X 3 M, n and ring A are as defined above.
In some embodiments of the invention, the above compound, or a pharmaceutically acceptable salt thereof, is selected from the group consisting of:
Figure BDA0003618772920000071
wherein R is 1 、R 3 、R 4 、m、n and ring A are as defined above.
In some embodiments of the invention, the above compound, or a pharmaceutically acceptable salt thereof, is selected from the group consisting of:
Figure BDA0003618772920000072
wherein R is 1 、R 3 、R 4 M and n are as defined above.
The present invention also provides the following compounds, or pharmaceutically acceptable salts thereof, selected from:
Figure BDA0003618772920000073
Figure BDA0003618772920000081
Figure BDA0003618772920000091
the present invention also provides a pharmaceutical composition comprising a therapeutically effective amount of the above compound or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, diluent or excipient.
The invention also provides application of the compound or pharmaceutically acceptable salt thereof or the pharmaceutical composition in preparing a medicament for treating cancer.
In some embodiments of the invention, the cancer comprises lymphoma, non-hodgkin's lymphoma, ovarian cancer, cervical cancer, prostate cancer, colorectal cancer, breast cancer, pancreatic cancer, glioma, glioblastoma, melanoma, leukemia, gastric cancer, endometrial cancer, lung cancer, hepatocellular carcinoma, gastric cancer, gastrointestinal stromal tumor (GIST), acute Myelogenous Leukemia (AML), cholangiocarcinoma, renal cancer, thyroid cancer, anaplastic large cell lymphoma, mesothelioma, multiple myeloma, or melanoma.
In some aspects of the invention, the cancer is lung cancer.
The invention also provides an intermediate compound shown as a formula (Z-3), a formula (Z-5) or a formula (Z-6) or stereoisomer and pharmaceutically acceptable salt thereof,
Figure BDA0003618772920000101
wherein PG is selected from tert-butyloxycarbonyl, benzyloxycarbonyl or p-toluenesulfonyl;
R 15 selected from H or C 1-4 An alkyl group;
R 2 、R 3 、R 4 、X 1 、X 2 、X 3 m, n, ring A and ring B are as defined above.
In some embodiments of the invention, the above intermediate is selected from:
Figure BDA0003618772920000102
/>
Figure BDA0003618772920000111
wherein PG is tert-butyloxycarbonyl;
R 15 selected from H, methyl or ethyl;
R 3 、R 4 M and n are as defined above.
The invention also provides a preparation method of the compound of the formula (I), which is characterized in that,
Figure BDA0003618772920000112
deprotecting a compound represented by the formula (Z-5) under acidic conditions to give a compound represented by the formula (Z-6), and introducing R 1 Preparing a compound shown in a formula (I),
wherein PG is selected from tert-butyloxycarbonyl or benzyloxycarbonyl, and the acid is selected from hydrochloric acid, acetic acid, trifluoroacetic acid or hydrobromic acid;
R 1 、R 2 、R 3 、R 4 、X 1 、X 2 、X 3 m, n, ring A and ring B are as defined above.
The invention also provides a process for the preparation of the compounds of formula (Z-5), characterized in that,
Figure BDA0003618772920000121
the compound shown in the formula (Z-3) or salt thereof is subjected to acylation reaction with the compound shown in the formula ((Z-4) or salt thereof under the action of a condensing agent,
wherein the condensing agent is selected from 2- (7-aza-benzotriazol) -N, N, N ', N' -tetramethyl urea hexafluorophosphate, 4- (4, 6-dimethoxy triazine) -4-methyl morpholine hydrochloride, dicyclohexylcarbodiimide, diisopropylcarbodiimide or 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide;
PG、R 2 、R 3 、R 4 、X 1 、X 2 、X 3 m, n, ring A and ring B are as defined above.
The invention also provides a process for the preparation of a compound of formula (Z-3), characterized in that:
Figure BDA0003618772920000122
the compound shown in the formula (Z-1) or salt thereof and the compound shown in the formula ((Z-2) or salt thereof are subjected to coupling reaction under the catalysis of palladium/copper,
Wherein R is 15 Selected from H or ethyl;
R 16 is that
Figure BDA0003618772920000123
The palladium/copper catalytic system is tetra (triphenylphosphine) palladium/thiophene-2-carboxylic acid cuprous (I), dichloro di (triphenylphosphine) palladium/thiophene-2-carboxylic acid cuprous (I) or palladium acetate/bis (2-diphenylphosphinophenyl) ether/thiophene-2-carboxylic acid cuprous (I);
PG、R 4 、X 1 、X 2 、X 3 n and ring B are as defined above.
The invention also provides a preparation method of the compound or the pharmaceutically acceptable salt thereof, and a representative preparation route is shown in the following scheme:
Figure BDA0003618772920000131
wherein PG, R 15 、R 16 、R 1 、R 2 、R 3 、R 4 、X 1 、X 2 、X 3 M, n, ring A and ring B are as defined above.
The method comprises the steps of (1) carrying out coupling reaction on a compound shown in a formula (Z-1) or salt thereof and a compound shown in a formula ((Z-2) or salt thereof under a palladium/copper catalytic system to obtain a compound shown in a formula (Z-3), wherein the palladium/copper catalytic system is tetra (triphenylphosphine) palladium/thiophene-2-cuprous (I) formate, dichloro di (triphenylphosphine) palladium/thiophene-2-cuprous (I) formate or palladium acetate/bis (2-diphenylphosphino) ether/thiophene-2-cuprous (I) formate;
the compound shown in the formula (Z-3) or salt thereof is subjected to acylation reaction with the compound shown in the formula ((Z-4) or salt thereof under the action of a condensing agent and alkali, wherein the condensing agent is selected from 4- (4, 6-dimethoxy triazine) -4-methylmorpholine hydrochloride (DMTMM), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC) and 1- (3-dimethylaminopropyl) -3-Ethylcarbodiimide (EDCI) or 2- (7-aza-benzotriazol) -N, N, N ', N' -tetramethyl urea Hexafluorophosphate (HATU);
Deprotecting a compound represented by the formula (Z-5) under acidic conditions to give a compound represented by the formula (Z-6), and introducing R 1 Preparing a compound shown in a formula (I), R 1 The groups may be introduced by nucleophilic substitution reactionsBy, for example, using R 1 X is halogen as a reactant, including but not limited to introduction of 2-methoxyethyl by using 1-iodo-2 methoxyethane, introduction of methanesulfonyl chloride into methanesulfonyl, introduction of methyl iodide into methyl, and the like. The methyl and ethyl groups can be introduced by using formaldehyde and aqueous solution of acetaldehyde, and then reducing the mixture by sodium borohydride acetate and the like.
Interpretation of the terms
The following terms and phrases used herein are intended to have the following meanings unless otherwise indicated. A particular term or phrase, unless otherwise specifically defined, should not be construed as being ambiguous or otherwise clear, but rather should be construed in a generic sense.
The term "pharmaceutically acceptable" refers to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
The term "pharmaceutically acceptable salts" refers to derivatives of the compounds of the present invention prepared with relatively non-toxic acids or bases. These salts may be prepared during synthesis, isolation, purification of the compound, or the purified compound may be used alone in free form to react with a suitable acid or base. When the compound contains relatively acidic functional groups, reaction with alkali metal, alkaline earth metal hydroxides or organic amines yields base addition salts, including cations based on alkali metals and alkaline earth metals, such as sodium, lithium, potassium, calcium, magnesium, and the like, as well as non-toxic ammonium, quaternary ammonium, and amine cations, and the like. When the compound contains a relatively basic functional group, it is reacted with an organic acid or an inorganic acid to give an acid addition salt.
The compounds provided herein also include pro-drug forms, meaning compounds that are rapidly converted in vivo to the parent compounds of the above formula, and converted to the compounds of the present invention by chemical or biochemical means in an in vivo or in vitro environment, for example by hydrolysis in blood.
The compounds of the invention can exist in unsolvated as well as solvated forms, including hydrated forms. In general, solvated forms are equivalent to unsolvated forms and are intended to be encompassed within the scope of the present invention.
The compounds of the present invention exist as geometric isomers as well as stereoisomers, such as cis-trans isomers, enantiomers, diastereomers, and racemic and other mixtures thereof, all of which are within the scope of the present invention.
The term "enantiomer" refers to stereoisomers that are mirror images of each other.
The term "diastereoisomer" refers to a stereoisomer of a molecule having two or more chiral centers and having a non-mirror image relationship between the molecules.
The term "cis-trans isomer" refers to a configuration in which a double bond or a single bond of a ring-forming carbon atom in a molecule cannot rotate freely.
Stereoisomers of the compounds of the invention may be prepared by chiral syntheses or chiral reagents or other conventional techniques. For example, one enantiomer of a compound of the invention may be prepared by asymmetric catalytic techniques or chiral auxiliary derivatization techniques. Or by chiral resolution techniques, a single configuration of the compound is obtained from the mixture. Or directly prepared by chiral starting materials. The separation of the optically pure compounds in the invention is usually accomplished by using preparative chromatography, and chiral chromatographic columns are used to achieve the purpose of separating chiral compounds.
The invention also includes isotopically-labeled compounds comprising isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, and chlorine, respectively, e.g. 2 H、 3 H、 13 C、 11 C、 14 C、 15 N、 18 O、 17 O、 31 P、 32 P、 35 S、 18 F and F 36 Cl. Compounds of the present invention containing the above isotopes and/or other isotopes of other atoms are within the scope of this invention.
Figure BDA0003618772920000141
Represents a single bond or a double bond, for example +.>
Figure BDA0003618772920000142
Middle X 1 And X 3 The two can be connected by single bond or double bond.
Where a bond of a substituent may cross-connect to two atoms on a ring, the substituent may be bonded to any atom on the ring.
For example
Figure BDA0003618772920000151
Represents R 3 Substitution can occur at any position of ring A, likewise, -/-, etc.>
Figure BDA0003618772920000152
Represents R 4 Substitution may occur at any position of ring B.
Figure BDA0003618772920000153
Represents R 3 Substitution can occur at any position on the quinoline ring, < >>
Figure BDA0003618772920000154
Represents R 3 Substitution can occur at any position on the pyridine ring, < >>
Figure BDA0003618772920000155
Represents R 4 Substitution may occur at any position on the benzene ring.
The term "pharmaceutically acceptable carrier" refers to a medium commonly accepted in the art for delivery of biologically active agents to animals, particularly mammals, and includes, for example, adjuvants, excipients or vehicles, such as diluents, preservatives, fillers, flow modifiers, disintegrants, wetting agents, emulsifying agents, suspending agents, sweetening, flavoring, perfuming, antibacterial, antifungal, lubricating and dispersing agents, depending on the mode of administration and nature of the dosage form. Pharmaceutically acceptable carriers are formulated within the purview of one of ordinary skill in the art according to a number of factors. Including but not limited to: the type and nature of the active agent formulated, the subject to which the composition containing the agent is to be administered, the intended route of administration of the composition, and the therapeutic indication of interest. Pharmaceutically acceptable carriers include both aqueous and nonaqueous media and a variety of solid and semi-solid dosage forms. Such carriers include many different ingredients and additives in addition to the active agent, and such additional ingredients included in the formulation for a variety of reasons (e.g., stabilizing the active agent, adhesive, etc.) are well known to those of ordinary skill in the art. The term "excipient" generally refers to the carrier, diluent, and/or medium required to make an effective pharmaceutical composition. The term "prophylactically or therapeutically effective amount" means that the compound of the invention, or a pharmaceutically acceptable salt thereof, is a sufficient amount of the compound to treat a disorder at a reasonable effect/risk ratio applicable to any medical treatment and/or prophylaxis. It will be appreciated that the total daily amount of the compounds of formula I or pharmaceutically acceptable salts and compositions of the present invention will be determined by the physician within the scope of sound medical judgment. For any particular patient, the particular therapeutically effective dose level will depend on a variety of factors including the disorder being treated and the severity of the disorder; the activity of the particular compound employed; the specific composition employed; age, weight, general health, sex and diet of the patient; the time of administration, route of administration and rate of excretion of the particular compound employed; duration of treatment; a medicament for use in combination with or simultaneously with the particular compound employed; and similar factors well known in the medical arts. For example, it is common in the art to start doses of the compound at levels below that required to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved. In general, the compounds of formula I or pharmaceutically acceptable salts thereof of the present invention may be administered to mammals, particularly humans, at a dosage of from 0.001 to 1000mg/kg body weight/day, for example from 0.01 to 100mg/kg body weight/day, for example from 0.01 to 10mg/kg body weight/day.
The term "optionally substituted" means that it may be substituted or notIn an unsubstituted, unless otherwise specified, substituent species and numbers may be arbitrary in that they are chemically realizable, e.g., the term "optionally substituted with one or more R 11 Substituted "means that one or more R's may be substituted 11 Substituted or not by R 11 And (3) substitution. When any variable (e.g. R 11 ) Where the composition or structure of a compound occurs more than once, its definition is independent in each case. For example, if a group is substituted with 0-2R 11 Substituted, the radicals may optionally be substituted by up to two R 11 Substituted, and R in each case 11 There are independent options.
Unless otherwise specified, "ring" refers to saturated, partially saturated or unsaturated monocyclic and polycyclic, and "polycyclic" includes bicyclic, spiro, and fused or bridged rings. Representative "rings" include substituted or unsubstituted cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, cycloalkynyl, heterocycloalkynyl, aryl, or heteroaryl. The term "hetero" refers to substituted or unsubstituted heteroatoms, typically selected from N, O, S, and oxidized forms of heteroatoms, typically including NO, SO, S (O) 2 The nitrogen atom may be substituted, i.e., NR (R is H or other substituent as defined herein); the number of atoms on the ring is generally defined as the number of ring elements, e.g., "5-7 membered heterocycloalkyl" means a mono-, bi-, spiro-, and heterocyclic or bridged-heterocyclic ring of 5-7 atoms arranged around, each ring optionally containing 1-3 heteroatoms, i.e., N, O, S, NO, SO, S (O) 2 Or NR.
Unless otherwise specified, "cycloalkyl" refers to a saturated monocyclic or polycyclic hydrocarbon group, including spirocyclic groups, fused ring groups, or bridged ring groups, which are equivalent to fused ring groups when the bridge atom in the bridged ring group is zero. 3-8 membered cycloalkyl is preferably 3-8 membered monocycloalkyl, more preferably 3-5 membered monocycloalkyl, examples of which include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and the like;
unless otherwise specified, "heterocycloalkyl" means that a number is included in the ringMono-and polyheterocycloalkyl radicals of heteroatoms of the order, said heteroatoms being generally selected from N, O, S, NO, SO, S (O) 2 And NR. The polyheterocyclic alkyl group includes spiroheterocyclyl, and heterocyclyl or bridged heterocyclyl, which is equivalent to a bridged heterocyclyl when the bridging atom in the bridged heterocyclyl is zero. The 3-8 membered heterocycloalkyl group is preferably a 3-8 membered mono-heterocycloalkyl group, and examples of such mono-heterocycloalkyl groups include, but are not limited to, oxiranyl, azetidinyl, tetrahydropyranyl, piperidinyl, piperazinyl, morpholinyl, tetrahydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, 1, 3-dioxolane, 1, 4-dioxane, and the like.
Unless otherwise specified, the term "aryl" refers to an unsaturated, typically aromatic, hydrocarbon group that may be a single ring or multiple rings fused together. Examples of aryl groups include, but are not limited to, phenyl, naphthyl.
Unless otherwise specified, the term "heteroaryl" means a stable monocyclic or polycyclic aromatic hydrocarbon containing at least one heteroatom (N, O, S, NO, SO, S (O) 2 Or NR. ). Preferably a 5-12 membered heteroaryl, more preferably a 5, 6, 7 membered monocyclic or bicyclic or 7, 8, 9 or 10 membered bicyclic heteroaryl; preferably comprising carbon atoms and 1, 2, 3 or 4 ring heteroatoms independently selected from N, O and S. Examples of heteroaryl groups include, but are not limited to, pyrrolyl, pyrazolyl, imidazolyl, pyrazinyl, oxazolyl, benzoxazolyl, isoxazolyl, thiazolyl, furanyl, thienyl, pyrimidinyl, benzothiazolyl, purinyl, benzimidazolyl, indolyl, isoquinolyl,
Figure BDA0003618772920000171
Figure BDA0003618772920000172
The term "alkyl" is used to denote a straight or branched saturated hydrocarbon group unless otherwise specified. Preferably C 1-6 More preferably C1-4 alkyl, examples of alkyl include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, isopentyl, neopentyl, n-hexyl and the like.
Unless otherwise specified, "alkenyl" refers to an alkyl group having one or more carbon-carbon double bonds. Preferably C 2-8 Examples of alkenyl groups, alkenyl groups include, but are not limited to, ethenyl, propenyl, butenyl, pentenyl, hexenyl, and the like.
Unless otherwise specified, "alkynyl" refers to an alkyl group having one or more carbon-carbon triple bonds. Preferably C 2-8 Examples of alkynyl groups, alkynyl groups include, but are not limited to, ethynyl, propynyl, butynyl, pentynyl, and the like.
The term "halogen" means a fluorine, chlorine, bromine or iodine atom unless otherwise specified.
It is specifically stated that combinations of all substituents and/or variants thereof are permissible only if such combinations result in stable compounds.
In the examples of the present invention, the title compound is named after the compound structure is converted by Chemdraw. If the compound name is inconsistent with the compound structure, the compound name can be determined in an auxiliary way by combining the related information and the reaction route; cannot be confirmed by other methods, and the structural formula of the given compound is subject to. The preparation method of some compounds in the present invention refers to the preparation method of the aforementioned analogous compounds. It will be appreciated by those skilled in the art that the ratio of the reactants, the reaction solvent, the reaction temperature, etc. may be appropriately adjusted depending on the reactants when using or referring to the preparation method to which they are applied.
The compounds of the present invention may be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments set forth below, embodiments formed by combining with other chemical synthetic methods, and equivalent alternatives well known to those skilled in the art, preferred embodiments including but not limited to the examples of the present invention.
Abbreviations used in the examples of the present invention and their corresponding chemical names are as follows:
abbreviations (abbreviations) Chemical name
Boc Tert-butyloxycarbonyl group
CuTC Thiophene-2-carboxylic acid cuprous (I)
Lawson reagent 2, 4-bis (p-methoxyphenyl) -1, 3-dithio-diphosphazetidine-2, 4-sulfide
DMF N, N-dimethylformamide
HATU 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethylurea hexafluorophosphate
DIPEA N, N-diisopropylethylamine
Detailed Description
The present invention is described in detail below by way of examples, but is not meant to be limiting in any way. The present invention has been described in detail herein, and specific embodiments thereof are also disclosed, it will be apparent to those skilled in the art that various changes and modifications can be made to the specific embodiments of the invention without departing from the spirit and scope of the invention.
The structure of the compounds of the present invention is determined by Nuclear Magnetic Resonance (NMR) or/and liquid chromatography-mass spectrometry (LC-MS). NMR chemical shifts (δ) are given in parts per million (ppm). NMR measurements were performed using Bruker Neo 400M or Bruker Ascend 400 nuclear magnetic instruments with deuterated dimethyl sulfoxide (DMSO-d 6), deuterated methanol (CD 3 OD) and/or deuterated chloroform (CDCl 3) as the measurement solvent and Tetramethylsilane (TMS) as the internal standard.
LC-MS was performed using an Agilent 1260-6125B single quadrupole mass spectrometer or a Waters H-Class SQD2 mass spectrometer (electrospray ionization as the ion source). HPLC determinations used Waters e2695-2998 or Waters ARC and Agilent 1260 or Agilent Poroshell HPH high performance liquid chromatography.
The HPLC was performed using Waters 2555-2489 (10 μm, ODS 250 cm. Times.5 cm) or GILSON Trilution LC, and the column was a Welch XB-C18 column (5 um, 21.2. Times.150 mm).
The thin layer chromatography silica gel plate uses a smoke table Jiang You silica gel development company GF254 silica gel plate or a new material company GF254 silica gel plate on the market of the nissan, the specification adopted by TLC is 0.15-0.20 mm, the preparation is 20x 20cm, and column chromatography is generally used for forming 200-300 mesh silica gel as a carrier.
The starting materials in the examples of the present invention are known and commercially available or may be synthesized using or according to methods known in the art.
All reactions of the invention were carried out under continuous magnetic stirring under dry nitrogen or argon atmosphere, with the solvent being a dry solvent and the reaction temperature being in degrees celsius, without specific description.
Example 1
(E) -3- (3- (4-bromophenyl) -8-methyl-1, 4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide
Figure BDA0003618772920000181
The reaction flow is as follows:
Figure BDA0003618772920000191
the reaction steps are as follows:
step 1: 2-amino-2- (4-bromophenyl) acetic acid (5.0 g,21.8 mmol) was dissolved in methanol (100 mL), and thionyl chloride (3.9 g,33.0 mmol) was slowly added dropwise and stirred at room temperature overnight. The reaction solution was concentrated under reduced pressure to obtain crude 2-amino-2- (4-bromophenyl) acetic acid methyl ester hydrochloride, which was directly used in the next reaction.
MS(ESI)M/Z:244.1[M+H] + .
Step 2: methyl 2-amino-2- (4-bromophenyl) acetate hydrochloride (6.2 g, crude) was dissolved in concentrated aqueous ammonia (40 mL) and stirred at room temperature for 2 days. White precipitate was precipitated and filtered. The filter cake was dried under vacuum to give 2.6g of 2-amino-2- (4-bromophenyl) acetamide.
MS(ESI)M/Z:229.0[M+H] + .
Step 3: 2-amino-2- (4-bromophenyl) acetamide (2.4 g,10.5 mmol) and tert-butyl 4-piperidone-1-carboxylate (2.1 g,10.5 mmol) were dissolved in ethanol (80 mL) and the reaction was heated to reflux under nitrogen overnight. The reaction was cooled to room temperature and concentrated under reduced pressure, the residue was dissolved in dichloromethane (80 mL), NBS (1.9 g,10.7 mmol) was added and stirred at room temperature overnight. After dilution with methylene chloride (100 mL), the mixture was washed with saturated aqueous sodium bicarbonate (100 mL. Times.2). The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue obtained was purified by column chromatography on silica gel (eluent: petroleum ether/ethyl acetate=2/1) to give 2.1g of tert-butyl 2- (4-bromophenyl) -3-oxo-1, 4, 8-triazaspiro [4.5] dec-1-ene-8-carboxylate.
MS(ESI)M/Z:408.1[M+H] + .
Step 4: tert-butyl 2- (4-bromophenyl) -3-oxo-1, 4, 8-triazaspiro [4.5] dec-1-ene-8-carboxylate (1.0 g,2.5 mmol) was dissolved in anhydrous toluene (30 mL), and Lawson reagent (993 mg,2.5 mmol) was added in portions and the mixture was stirred at 100deg.C for 3 hours. The reaction solution was cooled to room temperature and concentrated under reduced pressure. The resulting residue was purified by column chromatography on silica gel (eluent: petroleum ether/ethyl acetate=4/1) to give 800mg of tert-butyl 2- (4-bromophenyl) -3-thio-1, 4, 8-triazaspiro [4.5] dec-1-ene-8-carboxylate.
MS(ESI)M/Z:422.2[M-H]﹣.
Step 5: tert-butyl 2- (4-bromophenyl) -3-thio-1, 4, 8-triazaspiro [4.5] dec-1-ene-8-carboxylate (400 mg,0.95 mmol) and ethyl (E) -3-boronic acid pinacol ester-acrylate (320 mg,1.4 mmol) were dissolved in anhydrous tetrahydrofuran (20 mL) and CuTC (360 mg,1.9 mmol) and tetrakis (triphenylphosphine) palladium (109 mg,0.09 mmol) were added. The reaction was heated to reflux under nitrogen overnight. The reaction mixture was cooled to room temperature, diluted with ethyl acetate (30 mL), and washed with 25% concentrated aqueous ammonia (20 ml×2 times). The organic phase was washed with saturated brine (20 mL. Times.2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by column chromatography on silica gel (eluent: petroleum ether/ethyl acetate=4/1) to give 270mg of (E) -2- (4-bromophenyl) -3- (3-ethoxy-3-oxypropyl-1-en-1-yl) -1,4, 8-triazaspiro [4.5] decan-1, 3-diene-8-carboxylic acid tert-butyl ester.
MS(ESI)M/Z:490.4[M+H] + .
Step 6: to a mixed solution of (E) -2- (4-bromophenyl) -3- (3-ethoxy-3-oxypropyl-1-en-1-yl) -1,4, 8-triazaspiro [4.5] dec-1, 3-diene-8-carboxylic acid tert-butyl ester (260 mg,0.53 mmol) in tetrahydrofuran (10 mL) and water (5 mL) was added sodium hydroxide (86 mg,2.1 mmol) at room temperature. The reaction mixture was stirred at room temperature for 2 hours, the pH of the reaction mixture was adjusted to 5 with 1M diluted hydrochloric acid, and extracted with ethyl acetate (20 mL. Times.3). The organic phases were combined, washed with saturated brine (20 mL. Times.2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 245mg of (E) -3- (3- (4-bromophenyl) -8- (t-butoxycarbonyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylic acid.
MS(ESI)M/Z:460.3[M-H] ﹣ .
Step 7: (E) -3- (3- (4-bromophenyl) -8- (t-butoxycarbonyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylic acid (245 mg,0.53 mmol) and 3-aminoquinoline (115 mg,0.80 mmol) were dissolved in DMF (10 mL) and HATU (262 mg,0.69 mmol) and DIPEA (205 mg,1.6 mmol) were added. The reaction was stirred at room temperature overnight, poured into water (90 mL) and extracted with ethyl acetate (50 mL. Times.3). The organic phases were combined, washed with saturated brine (80 ml×2 times), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by column chromatography on silica gel (eluent: petroleum ether/ethyl acetate=1/1) to give 190mg of (E) -2- (4-bromophenyl) -3- (3-oxo-3- (quinolin-3-ylamino) propyl-1-en-1-yl) -1,4, 8-triazaspiro [4.5] decan-1, 3-diene-8-carboxylic acid tert-butyl ester.
MS(ESI)M/Z:588.4[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ10.99(s,1H),8.97(d,J＝2.4Hz,1H),8.83(d,J＝2.0Hz,1H),7.99-7.95(m,2H),7.82-7.80(m,2H),7.73-7.71(m,2H),7.68-7.53(m,3H),7.25(d,J＝15.2Hz,1H),3.72(br s,4H),1.73(br s,4H),1.46(s,9H).
Step 8: to a solution of (E) -2- (4-bromophenyl) -3- (3-oxo-3- (quinolin-3-ylamino) propyl-1-en-1-yl) -1,4, 8-triazaspiro [4.5] dec-1, 3-diene-8-carboxylic acid tert-butyl ester (180 mg,0.31 mmol) in dichloromethane (10 mL) was added trifluoroacetic acid (1 mL) and the mixture was stirred at room temperature for 1 hour. Saturated aqueous sodium bicarbonate (20 mL) was added and the mixture extracted with dichloromethane (20 mL. Times.3). The organic phases were combined, washed with saturated brine (30 mL. Times.2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue obtained is purified by column chromatography on silica gel (eluent: dichloromethane/methanol=10/1) to give 140mg of (E) -3- (3- (4-bromophenyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide.
MS(ESI)M/Z:488.3[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ11.29(s,1H),9.05(d,J＝2.4Hz,1H),8.85(d,J＝2.4Hz,1H),7.99-7.93(m,2H),7.83-7.73(m,4H),7.69-7.52(m,3H),7.35(d,J＝15.2Hz,1H),3.36(br s,4H),2.03(br s,4H).
Step 9: (E) -3- (3- (4-bromophenyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide (140 mg,0.29 mmol), 36% aqueous formaldehyde (76 mg,0.92 mmol) and acetic acid (37 mg,0.62 mmol) were dissolved in tetrahydrofuran (15 mL), and sodium borohydride acetate (195 mg,0.92 mmol) was added and stirred at room temperature for 2 hours. Saturated aqueous sodium bicarbonate (30 mL) was added and extracted with ethyl acetate (50 mL. Times.3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue obtained is purified by column chromatography on silica gel (eluent: dichloromethane/methanol=10/1) to give 105mg of the final product (E) -3- (3- (4-bromophenyl) -8-methyl-1, 4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide.
MS(ESI)M/Z:502.3[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ11.01(s,1H),8.97(d,J＝2.4Hz,1H),8.83(d,J＝2.4Hz,1H),7.99-7.95(m,2H),7.82-7.60(m,6H),7.54(d,J＝15.2Hz,1H),7.26(d,J＝15.6Hz,1H),2.85(br s,4H),2.49(br s,3H),1.90(br s,4H).
The following target product was prepared by the synthetic method of reference example 1:
Figure BDA0003618772920000211
/>
Figure BDA0003618772920000221
example 6
(E) -3- (8-methyl-3- (p-tolyl) -1,4, 8-triazaspiro [4.5] decan-1, 3-dien-2-methyl) -N- (quinolin-3-yl) acrylamide
Figure BDA0003618772920000222
The reaction flow is as follows:
Figure BDA0003618772920000231
the reaction steps are as follows:
150mg of the final product (E) -3- (8-methyl-3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-methyl) -N- (quinolin-3-yl) acrylamide was prepared using 2-amino-2- (p-tolyl) acetic acid as a starting material according to the procedure of example 1.
MS(ESI)M/Z:438.5[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ11.05(s,1H),8.90(d,J＝2.4Hz,1H),8.84(d,J＝2.0Hz,1H),7.99-7.95(m,2H),7.70-7.55(m,5H),7.40(d,J＝8.0Hz,2H),7.30(d,J＝15.2Hz,1H),2.85(br s,4H),2.51(s,3H),2.50(br s,3H),1.85(br s,4H).
Example 7
(E) -3- (8-methyl-3-phenyl-1, 4, 8-triazaspiro [4.5] dec-1, 3-dien-2-methyl) -N- (quinolin-3-yl) acrylamide
Figure BDA0003618772920000232
The reaction flow is as follows:
Figure BDA0003618772920000241
the reaction steps are as follows:
using 2-amino-2-phenylacetic acid as a starting material, 20mg of (E) -3- (8-methyl-3-phenyl-1, 4, 8-triazaspiro [4.5] dec-1, 3-dien-2-methyl) -N- (quinolin-3-yl) acrylamide as an end product was prepared according to the procedure of example 1.
MS(ESI)M/Z:424.4[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ10.07(s,1H),8.99(s,1H),8.84(d,J＝2.0Hz,1H),7.96(t,J＝8.4Hz,2H),7.77-7.55(m,8H),7.31(d,J＝15.2Hz,1H),2.89(br s,4H),2.50(s,3H),1.89(br s,4H).
Example 8
(E) -3- (8-ethyl-3- (p-tolyl) -1,4, 8-triazaspiro [4.5] decan-1, 3-dien-2-methyl) -N- (quinolin-3-yl) acrylamide
Figure BDA0003618772920000242
The reaction flow is as follows:
Figure BDA0003618772920000251
the reaction steps are as follows:
step 1: tert-butyl (E) -2- (3-oxo-3- (quinolin-3-amino) propyl-1-enyl) -3-p-tolyl-1, 4, 8-triazaspiro [4.5] decan-1, 3-diene-8-carboxylate (303 mg,0.58 mmol) was dissolved in ethyl acetate (5 mL), and an ethyl acetate solution (6M, 0.6mL,3.6 mmol) containing hydrogen chloride gas was added thereto and stirred at room temperature for 1 hour. A white solid precipitated, filtered and dried under vacuum to give 230mg of (E) -N- (quinolin-3-yl) -3- (3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide hydrochloride.
MS(ESI)M/Z:424.2[M+H] + .
Step 2: (E) -N- (quinolin-3-yl) -3- (3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide hydrochloride (100 mg,0.22 mmol) and triethylamine (89 mg,0.88 mmol) were dissolved in tetrahydrofuran (3 mL), acetaldehyde (29 mg,0.66 mmol) and sodium borohydride acetate (208 mg,0.98 mmol) were added, and the mixture was stirred at room temperature overnight. Water (15 mL) was added for dilution, and extraction was performed with ethyl acetate (20 mL. Times.3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue obtained was purified by high performance liquid chromatography to give 6.8mg of the final product (E) -3- (8-ethyl-3- (p-tolyl) -1,4, 8-triazaspiro [4.5] decan-1, 3-dien-2-methyl) -N- (quinolin-3-yl) acrylamide.
MS(ESI)M/Z:452.1[M+H] + .
1 H NMR(400MHz,CDCl 3 ):δ8.91(br s,2H),8.04(d,J＝8.4Hz,1H),7.83-7.75(m,2H),7.64-7.54(m,4H),7.29-7.23(m,3H),2.99(br s,4H),2.72(q,J＝7.2Hz,2H),2.40(s,3H),2.26(br s,4H),1.23(t,J＝7.2Hz,3H).
The following target product was prepared by the synthetic method of reference example 8:
Figure BDA0003618772920000252
/>
Figure BDA0003618772920000261
example 12
(E) -3- (8- (2-methoxyethyl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide
Figure BDA0003618772920000262
The reaction flow is as follows:
Figure BDA0003618772920000263
the reaction steps are as follows:
(E) -N- (quinolin-3-yl) -3- (3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide hydrochloride (50 mg,0.11 mmol) was dissolved in N, N-dimethylformamide (6 mL), followed by addition of potassium carbonate (60 mg,0.44 mmol) and 1-iodo-2 methoxyethane (20 mg,0.27 mmol), and the reaction was stirred at room temperature overnight. LCMS detection showed the disappearance of starting material, quench by adding water (10 mL) to the reaction, extract the mixture with ethyl acetate (20 mL x 3 times), combine the organic phases, wash the organic phases first with saturated brine (10 mL), then dry over anhydrous sodium sulfate, filter, and concentrate under reduced pressure. The residue obtained was purified by high performance liquid chromatography to give 14.6mg of the final product (E) -3- (8- (2-methoxyethyl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide.
MS(ESI)M/Z:482.3[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ10.98(s,1H),8.97(s,1H),8.83(s,1H),7.96(t,J＝7.6Hz,2H),7.67-7.54(m,5H),7.39(d,J＝8.0Hz,2H),7.28(d,J＝15.6Hz,1H),3.53(t,J＝6.0Hz,2H),3.28(s,3H),2.83(br s,4H),2.67(br s,2H),2.49(s,3H),1.79(br s,4H).
The following target product was prepared by the synthetic method of reference example 12:
Figure BDA0003618772920000271
example 17
(E) -3- (8- (2- (3, 3-difluoroazetidin-1-yl) -2-oxoethyl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide
Figure BDA0003618772920000281
The reaction flow is as follows:
Figure BDA0003618772920000282
the reaction steps are as follows:
step 1: (E) -N- (quinolin-3-yl) -3- (3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide hydrochloride (50 mg,0.11 mmol) was dissolved in tetrahydrofuran (6 mL), N-diisopropylethylamine (56 mg,0.44 mmol) and tert-butyl bromoacetate (21 mg,0.11 mmol) were added in this order, and the mixture was stirred at room temperature overnight. LCMS detection showed the starting material disappeared, quenched by addition of water (10 mL) to the reaction, and the mixture was extracted with ethyl acetate (20 mL x 3 times). The organic phases were combined, washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give crude (E) -tert-butyl 2- (2- (3-oxo-3- (quinolin-3-ylamino) propyl-1-en-1-yl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-8-yl) acetate, which was used directly in the next reaction.
MS(ESI)M/Z:538.3[M+H] + .
Step 2: tert-butyl (58 mg, crude) of (E) -2- (2- (3-oxo-3- (quinolin-3-ylamino) propyl-1-en-1-yl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-8-yl) acetate was dissolved in tetrahydrofuran and water (3 mL/3 mL), lithium hydroxide monohydrate (27 mg,0.66 mmol) was added, and stirred at room temperature overnight. LCMS detection showed the disappearance of starting material, water (10 mL) was added to the reaction system and ph=6-7 was adjusted with 3N dilute hydrochloric acid. The mixture was extracted with ethyl acetate (20 mL. Times.3), the organic phases were combined, washed with saturated brine (30 mL), then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure to give crude (E) -2- (2- (3-oxo-3- (quinolin-3-ylamino) propyl-1-en-1-yl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-8-yl) acetic acid, which was used directly in the next reaction.
MS(ESI)M/Z:482.3[M+H] + .
Step 3: (E) -2- (2- (3-oxo-3- (quinolin-3-ylamino) propyl-1-en-1-yl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-8-yl) acetic acid (52 mg, crude) and N, N-diisopropylethylamine (70 mg,0.55 mmol) were dissolved in dichloromethane (6 mL), HATU (62 mg,0.16 mmol) and 3, 3-difluorotrimethyleneimine hydrochloride (14 mg,0.11 mmol) were added and stirred at room temperature overnight. LCMS detection showed the disappearance of starting material, water (10 mL) was added to the reaction, and the mixture was extracted with dichloromethane (20 ml×3 times). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by high performance liquid chromatography to give 22.8mg of the final product (E) -3- (8- (2- (3, 3-difluoroazetidin-1-yl) -2-oxoethyl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide.
MS(ESI)M/Z:557.4[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ11.01(s,1H),8.98(s,1H),8.84(s,1H),7.97(t,J＝7.2Hz,2H),7.69-7.54(m,5H),7.39(d,J＝8.0Hz,2H),7.27(d,J＝15.6Hz,1H),4.77(t,J＝13.2Hz,2H),4.33(t,J＝12.8Hz,2H),3.51(s,2H),2.83(br s,4H),2.41(s,3H),1.99-1.75(m,4H).
Example 18
(E) -N-ethyl-2- (3-oxo-3- (quinolin-3-ylamino) -propyl-1-en-1-yl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] decan-1, 3-diene-8-carboxamide
Figure BDA0003618772920000291
The reaction flow is as follows:
Figure BDA0003618772920000292
the reaction steps are as follows:
(E) -N- (quinolin-3-yl) -3- (3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide (50 mg,0.12 mmol) was dissolved in dichloromethane (6 mL), triethylamine (26 mg,0.26 mmol) and ethyl isocyanate (17 mg,0.24 mmol) were added in this order, and the mixture was stirred at room temperature overnight. LCMS detection showed the disappearance of starting material, water (10 mL) was added to the reaction, and the mixture was extracted with dichloromethane (20 ml×3 times). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by high performance liquid chromatography to give 7.2mg of the final product (E) -N-ethyl-2- (3-oxo-3- (quinolin-3-ylamino) -propyl-1-en-1-yl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-diene-8-carboxamide.
MS(ESI)M/Z:495.3[M+H] + .
1 H NMR(400MHz,CDCl 3 ):δ10.13(s,1H),9.04(s,2H),8.09(d,J＝8.4Hz,1H),7.85-7.77(m,2H),7.66-7.54(m,4H),7.38(d,J＝15.2Hz,1H),7.30(d,J＝8.0Hz,2H),4.73(br s,1H),3.89-3.84(m,4H),3.38(q,J＝7.2Hz,2H),2.42(s,3H),1.89-1.86(m,4H),1.20(t,J＝7.2Hz,3H).
Example 19
(E) -3- (8-acetyl-3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide
Figure BDA0003618772920000301
The reaction flow is as follows:
Figure BDA0003618772920000302
the reaction steps are as follows:
(E) -N- (quinolin-3-yl) -3- (3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide hydrochloride (100 mg,0.22 mmol) was dissolved in methylene chloride (3 mL), and triethylamine (97 mg,0.96 mmol) and acetyl chloride (17 mg,0.22 mmol) were added in this order and stirred at room temperature for 1 hour. LCMS detection showed the disappearance of starting material, water (10 mL) was added to the reaction, and the mixture was extracted with dichloromethane (10 ml×3 times). The organic phases were combined, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue obtained was purified by high performance liquid chromatography to give 23.9mg of the final product (E) -3- (8-acetyl-3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) -N- (quinolin-3-yl) acrylamide.
MS(ESI)M/Z:466.2[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ11.01(s,1H),8.98(s,1H),8.83(s,1H),7.97(t,J＝7.2Hz,2H),7.69-7.56(m,5H),7.40(d,J＝8.0Hz,2H),7.28(d,J＝15.6Hz,1H),3.86-3.81(m,4H),2.41(s,3H),2.12(s,3H),1.80(t,J＝5.2Hz,2H),1.69(t,J＝5.2Hz,2H).
The following target product was prepared by the synthetic method of reference example 19:
Figure BDA0003618772920000311
example 24
(E) -N- (6-ethynylpyridin-3-yl) -3- (8-methyl-3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide
Figure BDA0003618772920000321
The reaction flow is as follows:
Figure BDA0003618772920000322
the reaction steps are as follows:
step 1: (E) -3- (8- (tert-Butoxycarbonyl) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylic acid (100 mg,0.25 mmol) was dissolved in dichloromethane (10 mL), N-diisopropylethylamine (97 mg,0.75 mmol) and HATU (143 mg,0.38 mmol) were added sequentially, and after stirring for 0.5 h, 6-ethynylpyridin-3-amine (33 mg,0.28 mmol) was added and stirred at room temperature overnight. LCMS detection showed the starting material disappeared, quenched by addition of water (10 mL) to the reaction, and the mixture was extracted with dichloromethane (20 mL x 3 times). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by column chromatography on silica gel (eluent: petroleum ether/ethyl acetate=1/1) to give 100mg of (E) -2- (3- ((6-ethynylpyridin-3-yl) amino) -3-acrylamido) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] decan-1, 3-diene-8-carboxylic acid tert-butyl ester.
MS(ESI)M/Z:498.3[M+H] + .
Step 2: (E) -2- (3- ((6-Acetylpyridin-3-yl) amino) -3-acrylamido) -3- (p-tolyl) -1,4, 8-triazaspiro [4.5] decan-1, 3-diene-8-carboxylic acid tert-butyl ester (100 mg,0.20 mmol) was dissolved in ethyl acetate (3 mL) under ice-bath, and a hydrogen chloride/ethyl acetate solution (4M, 1 mL) was added and stirred at room temperature for 2 hours. LCMS detection showed the disappearance of starting material and the reaction solution was concentrated under reduced pressure to give crude (E) -N- (6-ethynylpyridin-3-yl-3- (3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide hydrochloride, which was used directly in the next reaction.
MS(ESI)M/Z:398.2[M+H] + .
Step 3: (E) -N- (6-Acetyl pyridin-3-yl-3- (3- (p-tolyl) -1,4, 8-triazaspiro [4.5] dec-1, 3-dien-2-yl) acrylamide hydrochloride (87 mg,0.20 mmol) was dissolved in dichloromethane (6 mL), triethylamine (120 mg,1.2 mmol) and sodium triacetoxyborohydride (192 mg,0.90 mmol) were added, followed by 37% aqueous formaldehyde solution (50 mg,0.60 mmol) and stirred overnight at room temperature LCMS showed the disappearance of starting material, water (10 mL) was added to the reaction system to quench the mixture, the organic phases were combined, washed with saturated brine (30 mL) then dried over anhydrous sodium sulfate, filtered, and finally concentrated under reduced pressure, the resulting residue was purified by high-performance preparative liquid chromatography to give 8.9mg of the final product (E) -N- (6-ethynyl pyridin-3-yl) -3- (8-methyl-3- (p-tolyl) -1,4, 8-triazaspiro [ 4.5-dec-2-yl ] acrylamide.
MS(ESI)M/Z:412.0[M+H] + .
1 H NMR(400MHz,CDCl 3 ):δ8.68(s,1H),8.48(s,1H),8.36(s,1H),8.17(br s,1H),7.75(d,J＝15.2Hz,1H),7.61(d,J＝8.4Hz,2H),7.30(d,J＝7.6Hz,2H),7.20-7.10(m,1H),3.22(s,1H),2.97(br s,4H),2.56(s,3H),2.43(s,3H),2.20-1.60(m,4H).
The following target product was prepared by the synthetic method of reference example 24:
Figure BDA0003618772920000331
Figure BDA0003618772920000341
example 30
(E) -3- (8-methyl-4-oxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-2-yl) -N- (quinolin-3-yl) acrylamide
Figure BDA0003618772920000342
The reaction flow is as follows:
Figure BDA0003618772920000343
the reaction steps are as follows:
step 1: methyl 4-aminopiperidine-1-carboxylate (1.0 g,3.9 mmol) and 4-methylbenzenesulfonate (560 mg,3.9 mmol) are dissolved in 1, 4-dioxane (80 mL), and the reaction system is heated to 90℃under nitrogen and stirred overnight. The reaction solution was cooled to room temperature and concentrated under reduced pressure, and the residue was purified by beating with a mixed solvent of petroleum ether/ethyl acetate (20 mL) at a volume ratio of 5/1 to give 1.3g of t-butyl 4-oxo-2-thioxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] decane-8-carboxylate.
MS(ESI)M/Z:376.3[M+H] + .
Step 2: tert-butyl 4-oxo-2-thioxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] decane-8-carboxylate (1.3 g,3.5 mmol) and (E) -3-boronic acid pinacol ester-ethyl acrylate (1.3 g,5.8 mmol) were dissolved in anhydrous tetrahydrofuran (50 mL), and CuTC (1.3 g,6.9 mmol) and tetrakis (triphenylphosphine) palladium (400 mg,0.35 mmol) were added. The reaction was heated to reflux under nitrogen overnight. The reaction mixture was cooled to room temperature, diluted with ethyl acetate (50 mL), and washed with 25% aqueous ammonia (50 ml×2 times). The organic phase was washed with saturated brine (50 mL. Times.2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by column chromatography on silica gel (eluent: petroleum ether/ethyl acetate=4/1) to give 600mg of (E) -2- (3-ethoxy-3-oxypropyl-1-en-1-yl) -4-oxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-8-carboxylic acid tert-butyl ester.
MS(ESI)M/Z:442.4[M+H] + .
Step 3: to a mixed solution of (E) -2- (3-ethoxy-3-oxypropyl-1-en-1-yl) -4-oxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-8-carboxylic acid tert-butyl ester (500 mg,1.1 mmol) in tetrahydrofuran (16 mL) and water (8 mL) was added sodium hydroxide (90 mg,2.3 mmol) at room temperature. The reaction was stirred at room temperature for 2 hours, the pH of the reaction mixture was adjusted to 5 with 1M diluted hydrochloric acid, and extracted with ethyl acetate (20 mL. Times.3). The organic phases were combined, washed with saturated brine (20 mL. Times.2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 160mg of (E) -3- (8- (tert-butoxycarbonyl) -4-oxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-2-yl) acrylic acid.
MS(ESI)M/Z:436.0[M+Na] + .
Step 4: (E) -3- (8- (tert-Butoxycarbonyl) -4-oxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-2-yl) acrylic acid (150 mg,0.36 mmol) and 3-aminoquinoline (58 mg,0.40 mmol) were dissolved in DMF (8 mL) and HATU (178 mg,0.47 mmol) and DIPEA (140 mg,1.1 mmol) were added. The reaction was stirred at room temperature overnight, poured into water (50 mL) and extracted with ethyl acetate (30 mL. Times.3). The organic phases were combined, washed with saturated brine (40 mL. Times.2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by column chromatography on silica gel (eluent: petroleum ether/ethyl acetate=1/2) to give 145mg of (E) -4-oxo-2- (3-oxo-3- (quinolin-3-ylamino) propyl-1-en-1-yl) -3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-8-carboxylic acid tert-butyl ester.
MS(ESI)M/Z:540.3[M+H] + .
Step 5: to a solution of (E) -4-oxo-2- (3-oxo-3- (quinolin-3-ylamino) propyl-1-en-1-yl) -3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-8-carboxylic acid tert-butyl ester (145 mg,0.27 mmol) in dichloromethane (8 mL) was added trifluoroacetic acid (1.3 mL) and the mixture was stirred at room temperature for 1 hour. Saturated aqueous sodium bicarbonate (20 mL) was added and the mixture extracted with dichloromethane (20 mL. Times.3). The organic phases were combined, washed with saturated brine (30 ml×2 times), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give crude (E) -3- (4-oxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-2-yl) -N- (quinolin-3-yl) acrylamide, which was used directly in the next reaction.
MS(ESI)M/Z:440.2[M+H] + .
Step 6: (E) -3- (4-oxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-2-yl) -N- (quinolin-3-yl) acrylamide (120 mg, crude), 36% aqueous formaldehyde (65 mg,0.81 mmol) and acetic acid (33 mg,0.55 mmol) were dissolved in tetrahydrofuran (10 mL), and sodium borohydride acetate (171 mg,0.81 mmol) was added and stirred at room temperature for 2 hours. Saturated aqueous sodium bicarbonate (20 mL) was added and extracted with ethyl acetate (20 mL. Times.3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue obtained was purified by high performance liquid chromatography to give 4.5mg of the final product (E) -3- (8-methyl-4-oxo-3- (p-tolyl) -1,3, 8-triazaspiro [4.5] dec-1-en-2-yl) -N- (quinolin-3-yl) acrylamide.
MS(ESI)M/Z:454.2[M+H] + .
1 H NMR(400MHz,DMSO-d6):δ11.15(s,1H),8.96(s,1H),8.77(s,1H),7.97-7.93(m,2H),7.68-7.57(m,2H),7.41-7.37(m,3H),7.25(d,J＝8.0Hz,2H),6.86(d,J＝15.2Hz,1H),2.84(br s,2H),2.50(br s,2H),2.40(s,3H),2.37(s,3H),2.01-1.91(m,2H),1.67-1.64(m,2H).
Biological test evaluation
Test example 1: evaluation of proliferation inhibition effect of the Compounds of the present invention on Ba/F3 cell lines stably expressing triple mutant EGFR
The experiment adopts a fluorescence method to measure the intracellular ATP content to detect the fineness of the compound for stably expressing the triple mutant epidermal growth factor receptor (EGFR triple mutants)Proliferation inhibition effect of cell strain, and half inhibition concentration IC of compound for inhibiting proliferation of triple mutant EGFR (EGFR triple mutants) cell strain 50 。
1. Experimental materials
RPMI-1640 medium, fetal Bovine Serum (FBS), 100 XPen/Strep, glutaMAX-I supply was purchased from GIBCO corporation. Cell Titer-Glo luminescence Cell viability assay reagents were purchased from Promega corporation.
2. Experimental method
1) Stably transfected Ba/F3 (DEL 19/T790M/C797S and L858R/T790M/C797S) cells were counted using a cytometer and plated in 96-well plates at a density of 3000 cells per well, 100. Mu.l per well. Placing in incubator (37 ℃,5% CO) 2 ) Incubate overnight.
2) Day 0: 500nL of a gradient diluted test compound (initial concentration of 30. Mu.M, 10 concentrations, 1:3 dilution) was added to the cells of the plates using D300e (TECAN) and the final DMSO concentration was 0.5%, and the plates were incubated in a cell incubator for 72 hours (37 ℃,5% CO) 2 ). Blank control was added to 500nL of DMSO per well.
3) Day 3: mu.L of Cell Titer-Glo reagent was added to each well, and the mixture was shaken at 500rpm for 2 minutes, centrifuged at 1000rpm for 1 minute, and incubated at room temperature for 10 minutes under dark conditions to stabilize the luminescence signal.
4) The luminescence signal was detected by an Envision enzyme-labeled instrument (PerkinElmer).
5) Data analysis using GraphPad Prism 6 software, calculation of IC for compounds 50 。
The proliferation inhibition effect result of the compound of the invention on the Ba/F3 cell strain stably expressing the triple mutant EGFR is shown in Table 1, and the activity data is divided into A, B, C, D four sections and IC 50 Compounds of less than or equal to 0.5. Mu.M are identified by A, 0.5. Mu.M < IC 50 Compounds of less than or equal to 1. Mu.M are identified by B, 1. Mu.M < IC 50 Compounds less than or equal to 2. Mu.M are marked with C, IC 50 > 2. Mu.M is identified by D.
TABLE 1 inhibition of Ba/F3 cell lines stably expressing triple mutant EGFR receptors
Figure BDA0003618772920000371
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Figure BDA0003618772920000381
Conclusion: as can be seen from Table 1, the compounds of the present invention have a good inhibitory effect on Ba/F3 cell lines stably expressing the triple mutant EGFR.

Claims (26)

1. A compound represented by the formula (I) or a pharmaceutically acceptable salt thereof,
Figure FDA0004230336180000011
wherein R is 1 Selected from C 1-4 Alkyl, -C (O) OR 5 、-(CH 2 )s-OR 6 、-(CH 2 )t-C(O)NR 7 R 8 、-C(O)R 9 、-S(O) 2 R 10 Or 5-7 membered heterocycloalkyl, and said C 1-4 Alkyl is optionally substituted with one or more R 11 Substituted with a group;
when X is 1 When N is N, X 3 Is C, X 1 And X 3 The space is double bond;
X 1 when the compound is-C (=O), X is 3 Is N, X 1 And X 3 A single bond is arranged between the two;
X 2 is N;
R 2 is H;
ring A is selected from C 6-10 Aryl or 6-10 membered heteroaryl;
ring B is phenyl;
R 3 selected from halogen, C 2-4 Alkynyl, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-5 Cycloalkyl, -S (O) 2 R 10 、-P(O)(R 13 )R 14 Or C 6-10 An aryl group;
alternatively, two adjacent R 3 Cyclisation and ring A fusion to form
Figure FDA0004230336180000012
R 4 Selected from halogen or C 1-4 An alkyl group;
m and n are each independently selected from 0, 1, 2 or 3;
s and t are each independently selected from 0, 1 or 2;
R 5 and R is 6 Each independently selected from C 1-4 An alkyl group;
R 7 and R is 8 Each independently selected from H or C 1-4 An alkyl group;
alternatively, R 7 And R is 8 Together with the N atom to which they are attached form a 3-5 membered heterocycloalkyl, said 3-5 membered heterocycloalkyl optionally being further substituted with one or more R 12 Substituted with a group;
R 9 selected from C 1-4 Alkyl or C 3-5 Cycloalkyl;
R 10 is C 1-4 An alkyl group;
R 11 selected from OH, halogen, C 6-10 Aryl or 5-7 membered heteroaryl;
R 12 selected from OH or halogen;
R 13 and R is 14 Each independently selected from OH or C 1-4 An alkyl group.
2. The compound of claim 1, wherein R 5 Is tert-butyl.
3. The compound of claim 1, wherein R 6 Methyl, s is 2.
4. The compound of claim 1, wherein R 7 And R is 8 Each independently selected from H, methyl or ethyl;
alternatively, R 7 And R is 8 Together with the N atom to which it is attached form an azetidinyl group, which is optionally further substituted with one or moreR 12 Substituted with a group;
R 12 f is the same as F;
t is selected from 0 or 1.
5. The compound of claim 1, wherein R 9 Selected from methyl or cyclopropyl.
6. The compound of claim 1, wherein R 10 Selected from methyl or isopropyl.
7. The compound of claim 1, wherein R 13 And R is 14 Each independently is methyl.
8. The compound of claim 1, wherein R 1 Selected from methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, -C (O) OR 5 、-(CH 2 )s-OR 6 、-(CH 2 )t-C(O)NR 7 R 8 、-C(O)R 9 、-S(O) 2 R 10 Or tetrahydropyranyl, and said methyl or isobutyl group is optionally substituted with one or more R 11 Substituted with a group;
R 11 selected from OH, phenyl or pyridyl.
9. The compound of claim 8, wherein R 1 Selected from methyl, ethyl, isopropyl,
Figure FDA0004230336180000021
10. The compound of claim 1, wherein ring a is selected from the group consisting of
Figure FDA0004230336180000022
Figure FDA0004230336180000023
11. The compound of claim 1, wherein R 3 Selected from F, cl, ethynyl, methyl, ethyl, methoxy, cyclopropyl, -S (O) 2 CH 3 、-P(O)(CH 3 )CH 3 Or phenyl;
alternatively, two adjacent R 3 Cyclisation and ring A fusion to form
Figure FDA0004230336180000024
m is selected from 0, 1 or 2.
12. A compound according to claim 10 or 11, wherein the structural unit
Figure FDA0004230336180000031
Selected from the group consisting of
Figure FDA0004230336180000032
13. The compound of claim 1, wherein R 4 Selected from Br or methyl;
n is selected from 0 or 1.
14. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein the compound has a structure according to formula (II):
Figure FDA0004230336180000033
wherein R is 1 、R 3 、R 4 、X 1 、X 3 M, n and ring A are as defined in claim 1.
15. The compound of claim 14, or a pharmaceutically acceptable salt thereof, selected from:
Figure FDA0004230336180000041
wherein R is 1 、R 3 、R 4 M, n and ring A are as defined in claim 1.
16. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, selected from:
Figure FDA0004230336180000042
wherein R is 1 、R 3 、R 4 M and n are as defined in claim 1.
17. A compound, or a pharmaceutically acceptable salt thereof, selected from:
Figure FDA0004230336180000043
Figure FDA0004230336180000051
Figure FDA0004230336180000061
18. a pharmaceutical composition comprising a compound of any one of claims 1-17, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, diluent, or excipient.
19. Use of a compound according to any one of claims 1 to 17, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 18, in the manufacture of a medicament for the treatment of cancer.
20. The use of claim 19, wherein the cancer comprises lymphoma, ovarian cancer, cervical cancer, prostate cancer, colorectal cancer, breast cancer, pancreatic cancer, glioma, leukemia, gastric cancer, endometrial cancer, lung cancer, hepatocellular cancer, gastrointestinal stromal tumor, cholangiocarcinoma, renal cancer, thyroid cancer, mesothelioma, multiple myeloma, or melanoma.
21. The use of claim 20, wherein the cancer is lung cancer.
22. An intermediate compound represented by the formula (Z-6) or a pharmaceutically acceptable salt thereof,
Figure FDA0004230336180000071
wherein R is 2 、R 3 、R 4 、X 1 、X 2 、X 3 M, n, ring A and ring B are as defined in claim 1.
23. An intermediate compound of formula (Z-6) according to claim 22, or a pharmaceutically acceptable salt thereof, selected from:
Figure FDA0004230336180000072
wherein R is 3 、R 4 M and n are as defined in claim 1.
24. A process for the preparation of a compound of formula (I) or a pharmaceutically acceptable salt thereof, characterized in that,
Figure FDA0004230336180000073
deprotecting a compound represented by the formula (Z-5) under acidic conditions to give a compound represented by the formula (Z-6), and introducing R 1 Preparing a compound shown in a formula (I),
wherein PG is selected from tert-butyloxycarbonyl or benzyloxycarbonyl, and the acid is selected from hydrochloric acid, acetic acid, trifluoroacetic acid or hydrobromic acid;
R 1 、R 2 、R 3 、R 4 、X 1 、X 2 、X 3 M, n, ring A and ring B are as defined in claim 1.
25. The process for the preparation of a compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 24, wherein the process for the preparation of a compound of formula (Z-5) is as follows:
Figure FDA0004230336180000081
the compound shown in the formula (Z-3) or salt thereof is subjected to acylation reaction with the compound shown in the formula ((Z-4) or salt thereof under the action of condensing agent and alkali,
wherein the condensing agent is selected from 2- (7-aza-benzotriazol) -N, N, N ', N' -tetramethyl urea hexafluorophosphate, 4- (4, 6-dimethoxy triazine) -4-methyl morpholine hydrochloride, dicyclohexylcarbodiimide, diisopropylcarbodiimide or 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide;
PG、R 2 、R 3 、R 4 、X 1 、X 2 、X 3 m, n, ring A and ring B are as defined in claim 24.
26. The process for the preparation of a compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 25, wherein the process for the preparation of a compound of formula (Z-3) is as follows:
Figure FDA0004230336180000082
the compound shown in the formula (Z-1) or salt thereof and the compound shown in the formula ((Z-2) or salt thereof are subjected to coupling reaction under the catalysis of palladium/copper,
wherein R is 15 Selected from H or ethyl;
R 16 is that
Figure FDA0004230336180000091
The palladium/copper catalytic system is tetra (triphenylphosphine) palladium/thiophene-2-carboxylic acid cuprous (I), dichloro di (triphenylphosphine) palladium/thiophene-2-carboxylic acid cuprous (I) or palladium acetate/bis (2-diphenylphosphinophenyl) ether/thiophene-2-carboxylic acid cuprous (I);
PG、R 4 、X 1 、X 2 、X 3 N and ring B are as defined in claim 24.
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