Preparation method and application of compound containing zinc binding group and quinoline skeleton
阅读说明：本技术 含锌结合基以及喹啉骨架的化合物的制备方法和用途 (Preparation method and application of compound containing zinc binding group and quinoline skeleton ) 是由 宫平 秦铭泽 赵燕芳 刘亚婧 侯云雷 胡浩 于 2020-05-07 设计创作，主要内容包括：本发明涉及药物化学领域,特别涉及一种新的含锌结合基的喹啉类化合物、其几何异构体及其药学上可接受的盐、溶剂化物或前药,他们的制备方法以及含有所述化合物的药物组合物。本发明还涉及该类化合物用于制备治疗和/或预防由c-Met酪氨酸激酶和HDAC所介导的疾病的药物中的应用。所述的喹啉类化合物、其几何异构体及其药学上可接受的盐、溶剂化物或前药的结构如下所示,其中,R-(1)、m、Q、X如权利要求书和说明书所述。(The invention relates toAnd the pharmaceutical chemistry field, in particular to a new quinoline compound containing a zinc binding group, a geometrical isomer thereof, a pharmaceutically acceptable salt, a solvate or a prodrug thereof, a preparation method thereof and a pharmaceutical composition containing the compound. The invention also relates to the application of the compounds in preparing medicines for treating and/or preventing diseases mediated by c-Met tyrosine kinase and HDAC. The structures of the quinoline compound, the geometric isomer thereof, and the pharmaceutically acceptable salt, solvate or prodrug thereof are shown as follows, wherein R 1 M, Q, X are as described in the claims and the description.)
1. A compound of formula (I) and pharmaceutically acceptable salts, solvates, geometric isomers, enantiomers, diastereomers, racemates or prodrugs thereof,
wherein the content of the first and second substances,
R1hydrogen or (C)1-C6) An alkoxy group,
m＝1-6，
the number X is O, NH, and the number X is O, NH,
q is
R2Selected from the group consisting of hydrogen, hydroxy, halogen, nitro, amino, cyano, carbamoyl, aminosulfonyl, (C)1-C6) Alkyl, (C)1-C6) Alkoxy group, (C)2-C6) Alkynyl, (C)1-C6) Acyl, optionally hydroxy-, amino-, cyano-or halogen-substituted (C)1-C6) Alkyl or (C)1-C6) Alkoxy, mono-or di (C)1-C6Alkyl) substituted amino, (C)1-C6) Alkylamide group, (C)1-C6) Alkyl acyl group, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl and free, salified, esterified and amidated carboxyl groups;
wherein R is3Is halogen or hydrogen;
b is
R4Is selected from hydrogen, (C)1-C6) Alkyl or halo (C)1-C6) An alkyl group;
R5selected from hydrogen, (C)1-C6) Alkyl or halo (C)1-C6) An alkyl group;
cy is (C)1-C6) Alkyl, (C)3-C7) Cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 6-10 membered arylmethyl, 5-10 membered heteroarylmethyl, said aryl or heteroaryl optionally substituted with 0-3R which may be the same or different6Substitution;
R6is hydroxy, halogen, nitro, amino, cyano, (C)1-C6) Alkyl, (C)2-C6) Alkenyl, (C)2-C6) Alkynyl, (C)1-C6) Alkoxy, optionally hydroxy, amino or halo (C)1-C6) Alkyl or (C)1-C6) Alkoxy radical, 1-2 (C)1-C6) Alkyl-substituted amino group, (C)1-C6) Alkylamido, free, salified, esterified and amidated carboxyl, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl group, (C)1-C6) Alkoxy group, (C)1-C6) Alkyl, (C)1-C6) Alkyl acyl group, (C)1-C6) Carbamoyl radical, substituted by 1-2 (C)1-C6) Alkyl-substituted carbamoyl group, (C)1-C3) Alkylenedioxy, allyl.
2. A compound of formula (I) and pharmaceutically acceptable salts, solvates, geometric isomers, enantiomers, diastereomers, racemates or prodrugs thereof,
wherein the content of the first and second substances,
R1is (C)1-C4) An alkoxy group;
m＝1-6；
when X is NH; q is
R2Selected from the group consisting of hydrogen, hydroxy, halogen, nitro, amino, cyano, carbamoyl, aminosulfonyl, (C)1-C6) Alkyl, (C)1-C6) Alkoxy group, (C)2-C6) Alkynyl, (C)1-C6) Acyl, optionally hydroxy-, amino-, cyano-or halogen-substituted (C)1-C6) Alkyl or (C)1-C6) Alkoxy, mono-or di (C)1-C6Alkyl) substituted amino, (C)1-C6) Alkylamide group, (C)1-C6) Alkyl acyl group, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl and free, salified, esterified and amidated carboxyl groups.
When X is O, Q is
Wherein R is3Preferably hydrogen or fluorine
Wherein B is
R4Is hydrogen, (C)1-C4) An alkyl group;
R5is hydrogen, (C)1-C4) Alkyl or halo (C)1-C4) An alkyl group;
cy is phenyl optionally substituted by 0 to 3 identical or different R6And (4) substitution.
R6Is hydroxy, halogen, halo (C)1-C6) Alkyl, halo (C)1-C6) Alkoxy, nitro, amino, cyano, (C)1-C6) Alkyl, (C)2-C6) Alkenyl, (C)2-C6) Alkynyl, (C)1-C6) Alkoxy, optionally hydroxy, amino or halo (C)1-C6) Alkyl or (C)1-C6) Alkoxy radical, 1-2 (C)1-C6) Alkyl-substituted amino group, (C)1-C6) Alkylamido, free, salified, esterified and amidated carboxyl, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl group, (C)1-C6) Alkoxy group, (C)1-C6) Alkyl, (C)1-C6) Alkyl acyl group, (C)1-C6) Carbamoyl radical, substituted by 1-2 (C)1-C6) Alkyl-substituted carbamoyl group, (C)1-C3) Alkylenedioxy, allyl.
3. A compound of formula (I) and pharmaceutically acceptable salts, solvates, geometric isomers, enantiomers, diastereomers, racemates or prodrugs thereof,
R1is methoxy;
m＝1-6；
x is O, NH;
when X is NH; q is
R2Selected from the group consisting of hydrogen, hydroxy, halogen, nitro, amino, cyano, carbamoyl, aminosulfonyl, (C)1-C6) Alkyl, (C)1-C6) Alkoxy group, (C)2-C6) Alkynyl, (C)1-C6) Acyl, optionally hydroxy-, amino-, cyano-or halogen-substituted (C)1-C6) Alkyl or (C)1-C6) Alkoxy, mono-or di (C)1-C6Alkyl) substituted amino, (C)1-C6) Alkylamide group, (C)1-C6) Alkyl acyl group, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl and free, salified, esterified and amidated carboxyl groups.
When X is O, Q is
Wherein R is3Preferably hydrogen or fluorine;
wherein B is
R4Preferably hydrogen, methyl, ethyl;
R5selected from hydrogen, methyl or trifluoromethyl;
cy is phenyl optionally substituted by 0 to 3 identical or different R6Substitution;
R6is hydroxy, halogen, nitro, amino, cyano, (C)1-C6) Alkyl, (C)2-C6) Alkenyl, (C)2-C6) Alkynyl, (C)1-C6) Alkoxy, optionally hydroxy, amino or halo (C)1-C6) Alkyl or (C)1-C6) Alkoxy radical, 1-2 (C)1-C6) Alkyl-substituted amino group, (C)1-C6) Alkylamido, free, salified, esterified and amidated carboxyl, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl group, (C)1-C6) Alkoxy group, (C)1-C6) Alkyl, (C)1-C6) Alkyl acyl group, (C)1-C6) Carbamoyl radical, substituted by 1-2 (C)1-C6) Alkyl-substituted carbamoyl group, (C)1-C3) Alkylenedioxy, allyl.
4. A compound of the structure or a geometric isomer, enantiomer, diastereomer, racemate, pharmaceutically acceptable salt, prodrug thereof:
5. a process for the preparation of compounds of general formula (I) according to claim 1 and pharmaceutically acceptable salts thereof:
6. a pharmaceutical composition comprising a compound of any one of claims 1-4 or a geometric isomer, enantiomer, diastereomer, racemate, pharmaceutically acceptable salt, prodrug thereof, and a pharmaceutically acceptable carrier or excipient.
7. Use of a compound according to any one of claims 1 to 4 or a geometric isomer, enantiomer, diastereomer, racemate, pharmaceutically acceptable salt, prodrug, or pharmaceutical composition according to claim 6 thereof in the manufacture of a c-Met tyrosine kinase inhibitor or HDAC inhibitor.
8. Use of a compound according to any one of claims 1 to 4 or a geometric isomer, enantiomer, diastereomer, racemate, pharmaceutically acceptable salt, prodrug, or pharmaceutical composition according to claim 6 thereof in the manufacture of a c-Met tyrosine kinase inhibitor and an HDAC inhibitor.
9. Use of a compound according to any one of claims 1 to 4 or a geometric isomer, enantiomer, diastereomer, racemate, pharmaceutically acceptable salt, prodrug, or pharmaceutical composition according to claim 6 thereof in the manufacture of an anti-neoplastic medicament.
10. The use according to claim 9, wherein the neoplasm is breast cancer, lung cancer, liver cancer, kidney cancer, colon cancer, rectal cancer, stomach cancer, prostate cancer, bladder cancer, uterine cancer, pancreatic cancer, bone marrow cancer, testicular cancer, ovarian cancer, lymphatic cancer, soft tissue cancer, head and neck cancer, thyroid cancer, esophageal cancer, leukemia, and neuroblastoma.
The technical field is as follows:
the invention relates to the field of pharmaceutical chemistry, in particular to a novel quinoline compound containing a zinc binding group, a geometric isomer thereof, a pharmaceutically acceptable salt, a solvate or a prodrug thereof, a preparation method thereof and a pharmaceutical composition containing the compound. The invention also relates to the application of the compounds in preparing medicines for treating and/or preventing diseases mediated by c-Met tyrosine kinase and HDAC.
Background art:
cancer, also known as malignant tumor, is a disease caused by the malfunction of the mechanism controlling cell growth and proliferation. In the last 90 s of the last century, with the rapid development of cell biology, molecular biology, genetics and other disciplines, the research on relevant targets and signal transduction pathways in the process of tumorigenesis and development has been deepened, and molecular targeted therapy (molecular targeted therapy) which uses cell receptors, key genes and regulatory molecules as targets has begun. In 1986, Park et al [ Park M, Dean M, Cooper, CS, et al, mechanism of oncogene activation [ J ]. Cell,1986,45(6):895-904 ] reported the cDNA sequence of c-Met kinase and determined that it belongs to the tyrosine kinase family. Subsequently, c-Met kinase was identified as the only high affinity Receptor in the Receptor tyrosine kinase family that binds to Hepatocyte Growth Factor (Hepatocyte Growth Factor), and hence is also referred to as Hepatocyte Growth Factor Receptor (HGFR), the expression product of which is a transmembrane Receptor protein with tyrosine kinase activity. Under normal physiological conditions, both c-Met kinase and HGF are expressed in a large number of tissues, but c-Met RNA is expressed at low levels, and only rises briefly after tissue damage, and then returns to normal levels, indicating the ability of normal cells to control their response to HGF by reducing expression of c-Met kinase. C-Met kinase can promote epithelial Cell dispersion, enhance Cell motility, promote Cell growth, proliferation, differentiation, contraction, movement, secretion and mitosis, and has important biological significance for promoting the development of placenta and embryo, regulating the development and structure formation of organs such as lung, nervous system, kidney and mammary gland [ Giordano S, Ponzetto C, Di Renzo MF, et al. tyrosinase receptor operable from the C-Met protein [ J ]. Nature,1989,339: 155: 156. Birchmeier C, Birchmeier W, Gherrdi E, et al. Met kinase, mobility and more [ J ]. Nat Rev Mol Biol, 2003,4(12): 925-Christen J, Burrows J, Burkitt R.c. nucleic acid J.for culture J., 2005,225(1):1-26]
Histone Deacetylases (HDACs) are closely related to tumorigenesis and regulate gene transcription and Chromatin recombination (Chromatin remodelling) [ Groselj B, Sharma NL, Hamdy FC, et al, Histone deacetylase inhibiting as Radiosensisers: efffects on DNA damagesingling and repair [ J ]. Br J Cancer,2013,108 (108) J-754. Bertrand P.Inside HDAC inhibitors [ J ]. Eur J Med Chem,2010,45(6):2095-2116. DelcuGP, Khan DH, video JR.targeting class I acetylases [ J ]. D J.D. C.E.E., 2010,45(6):2095-2116. DelcuGP, Khan DH, J.E.S.E.T. III acetylation of Histone I proteins [ J.: proteins ] proteins, hsp.S.S.S.S.S.J.: proteins, 23, 3, 23, 3, 23, 3, 2,3, 2,3, 2,3, 2,3, 2,3, 2,3, 2,3, 2, one, three, one, three, one, three. Inhibiting HDACs can induce tumor cell cycle arrest, differentiation and apoptosis. In 1999, Finnin [ Paris M, Porceloni M, Binaschi M, et al, Histone deacylase inhibitors from benzene to clinic [ J ]. J Med Chem,2008,51(6):1505 cake 1529] reported for the first time the co-crystal structure of HDLP (Histone deacylase like protein) and SAHA, which established the basis for the rational design of HDAC inhibitors. Studies have shown that the structural features of HDAC inhibitors are as follows: 1) contains a Zinc-binding group (ZBG) for chelating Zinc ions at the bottom of the HDAC pocket; 2) contains a surface recognition group, called CAP moiety (CAP); 3) a hydrophobic linker linking ZBG and CAP.
Clinical test data show that HDAC is effective alone, has wide antitumor activity, can play a synergistic role in mediating apoptosis when being used together with antitumor drugs with different pharmacological and biological mechanisms, has an action mechanism of simultaneously activating a plurality of apoptosis pathways when being used together, and can reduce the apoptosis threshold of tumor cells and inhibit the anti-apoptosis effect of other factors. The c-Met oncogene encoding HGF controls cell formation, invasion and protects cells from apoptosis. Therefore, the abnormal activation of c-Met is not only important for the generation of tumors, but also important for the invasion and metastasis of the tumors, and the c-Met inhibitor inhibits the generation and metastasis of the tumors by blocking the abnormal activation of the c-Met, thereby inducing the apoptosis of tumor cells.
Based on the above, when the HDAC and the c-Met inhibitor are used together, the HDAC and the c-Met inhibitor reduce the apoptosis threshold of tumor cells, and inhibit the anti-apoptosis effect of other factors to promote the c-Met inhibitor to block the activation of c-Met, inhibit the formation and metastasis of tumors and induce the apoptosis of tumor cells. In fact, HDAC and c-Met are both proteins with general biological significance for tumor cell formation regulation, and thus both are abnormally highly expressed in many cancers, for example, HDAC and c-Met inhibitors can treat many of the same tumors, such as prostate cancer, leukemia, breast cancer, colon cancer and liver cancer. Clearly, designing both drugs simultaneously to target both has many advantages over combining HDAC and c-Met inhibitors.
At present, relatively few reports are reported on the relevant documents aiming at the c-Met/HDAC inhibitor, and no c-Met/HDAC small molecule inhibitor is approved to be on the market. Among them, Dong Lu et al [ Design, Synthesis, and Biological Evaluation of the First c-Met/HDAC Inhibitors Based on polymeric Derivatives [ J].ACS Medicinal Chemistry Letters,2017,8(8):830-834.]Reported c-Met and HDAC1 kinase inhibitors, IC, of Compound 2m500.71nM and 38nM, respectively.
The inventor designs and synthesizes a plurality of series of quinoline compounds containing zinc binding groups on the basis of the reference literature, and the results of in vitro c-Met and HDAC kinase inhibitory activity and cancer cell proliferation activity tests show that the quinoline compounds have inhibitory activity to two enzymes and corresponding cancer cells.
Disclosure of Invention
A compound of formula (I) and pharmaceutically acceptable salts, solvates, geometric isomers, enantiomers, diastereomers, racemates or prodrugs thereof
Wherein the content of the first and second substances,
R1hydrogen or (C)1-C6) An alkoxy group;
m＝1-6；
x is O, NH;
q is
R2Selected from the group consisting of hydrogen, hydroxy, halogen, nitro, amino, cyano, carbamoyl, aminosulfonyl, (C)1-C6) Alkyl, (C)1-C6) Alkoxy group, (C)2-C6) Alkynyl, (C)1-C6) Acyl, optionally hydroxy-, amino-, cyano-or halogen-substituted (C)1-C6) Alkyl or (C)1-C6) Alkoxy, mono-or di (C)1-C6Alkyl) substituted amino, (C)1-C6) Alkylamide group, (C)1-C6) Alkyl acyl group, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl and free, salified, esterified and amidatedA carboxyl group;
R3is halogen or hydrogen;
wherein B is
R4Is selected from hydrogen, (C)1-C6) Alkyl or halo (C)1-C6) An alkyl group;
R5selected from hydrogen, (C)1-C6) Alkyl or halo (C)1-C6) An alkyl group;
cy is (C)1-C6) Alkyl, (C)3-C7) Cycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, 6-10 membered arylmethyl, 5-10 membered heteroarylmethyl, said aryl or heteroaryl optionally substituted with 0-3R which may be the same or different6Substitution;
R6is hydroxy, halogen, nitro, amino, cyano, (C)1-C6) Alkyl, (C)2-C6) Alkenyl, (C)2-C6) Alkynyl, (C)1-C6) Alkoxy, optionally hydroxy, amino or halo (C)1-C6) Alkyl or (C)1-C6) Alkoxy radical, 1-2 (C)1-C6) Alkyl-substituted amino group, (C)1-C6) Alkylamido, free, salified, esterified and amidated carboxyl, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl group, (C)1-C6) Alkoxy group, (C)1-C6) Alkyl, (C)1-C6) Alkyl acyl group, (C)1-C6) Carbamoyl radical, substituted by 1-2 (C)1-C6) Alkyl-substituted carbamoyl group, (C)1-C3) Alkylenedioxy, allyl.
The invention prefers the compound of the general formula (I) and the pharmaceutically acceptable salt, solvate, geometrical isomer, enantiomer, diastereoisomer, racemate or prodrug thereof,
wherein the content of the first and second substances,
R1is (C)1-C4) An alkoxy group;
m＝1-6；
x is O, NH;
when X is NH; q is
R2Selected from the group consisting of hydrogen, hydroxy, halogen, nitro, amino, cyano, carbamoyl, aminosulfonyl, (C)1-C6) Alkyl, (C)1-C6) Alkoxy group, (C)2-C6) Alkynyl, (C)1-C6) Acyl, optionally hydroxy-, amino-, cyano-or halogen-substituted (C)1-C6) Alkyl or (C)1-C6) Alkoxy, mono-or di (C)1-C6Alkyl) substituted amino, (C)1-C6) Alkylamide group, (C)1-C6) Alkyl acyl group, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl and free, salified, esterified and amidated carboxyl groups.
When X is O, Q is
Wherein R is3Preferably hydrogen or fluorine;
wherein B is
R4Preferably hydrogen, (C)1-C4) An alkyl group;
R5selected from hydrogen, (C)1-C4) Alkyl or halo (C)1-C4) An alkyl group;
cy is preferably phenyl, which is optionally substituted by 0 to 3 identical or different R6And (4) substitution.
R6Is hydroxy, halogen, nitro, amino, cyano, (C)1-C6) Alkyl, (C)2-C6) Alkenyl, (C)2-C6) Alkynyl, (C)1-C6) Alkoxy, optionally hydroxy, amino or halo (C)1-C6) Alkyl or (C)1-C6) Alkoxy radical, 1-2 (C)1-C6) Alkyl-substituted amino group, (C)1-C6) Alkylamido, free, salified, esterified and amidated carboxyl, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl group, (C)1-C6) Alkoxy group, (C)1-C6) Alkyl, (C)1-C6) Alkyl acyl group, (C)1-C6) Carbamoyl radical, substituted by 1-2 (C)1-C6) Alkyl-substituted carbamoyl group, (C)1-C3) Alkylenedioxy, allyl.
The invention prefers the compound of the general formula (I) and the pharmaceutically acceptable salt, solvate, geometrical isomer, enantiomer, diastereoisomer, racemate or prodrug thereof,
wherein the content of the first and second substances,
R1is methoxy;
m＝1-6；
x is O, NH;
when X is NH; q is
R2Selected from hydrogen, hydroxyl, halogen, nitro,Amino, cyano, carbamoyl, aminosulfonyl, (C)1-C6) Alkyl, (C)1-C6) Alkoxy group, (C)2-C6) Alkynyl, (C)1-C6) Acyl, optionally hydroxy-, amino-, cyano-or halogen-substituted (C)1-C6) Alkyl or (C)1-C6) Alkoxy, mono-or di (C)1-C6Alkyl) substituted amino, (C)1-C6) Alkylamide group, (C)1-C6) Alkyl acyl group, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl and free, salified, esterified and amidated carboxyl groups.
When X is O, Q is
Wherein R is3Preferably hydrogen or fluorine;
wherein B is
R4Preferably hydrogen, methyl, ethyl;
R5selected from hydrogen, methyl or trifluoromethyl;
cy is phenyl optionally substituted by 0 to 3 identical or different R6Substitution;
R6is hydroxy, halogen, nitro, amino, cyano, (C)1-C6) Alkyl, (C)2-C6) Alkenyl, (C)2-C6) Alkynyl, (C)1-C6) Alkoxy, optionally hydroxy, amino or halo (C)1-C6) Alkyl or (C)1-C6) Alkoxy radical, 1-2 (C)1-C6) Alkyl-substituted amino group, (C)1-C6) Alkylamido, free, salified, esterified and amidated carboxyl, (C)1-C6) Alkylsulfinyl (C)1-C6) Alkylsulfonyl group, (C)1-C6) Alkoxy radical,(C1-C6) Alkyl, (C)1-C6) Alkyl acyl group, (C)1-C6) Carbamoyl radical, substituted by 1-2 (C)1-C6) Alkyl-substituted carbamoyl group, (C)1-C3) Alkylenedioxy, allyl.
The invention preferably relates to the following compounds and pharmaceutically acceptable salts, solvates, geometrical isomers, enantiomers, diastereoisomers, racemates or prodrugs thereof,
the compounds of formula (I) of the present invention may form pharmaceutically acceptable salts thereof with acids according to some conventional methods in the art to which the present invention pertains. Preferred acids are hydrochloric, hydrobromic, hydrofluoric, sulfuric, phosphoric, nitric, formic, acetic, propionic, oxalic, malonic, succinic, fumaric, maleic, lactic, malic, tartaric, citric, picric, methanesulfonic, ethanesulfonic, toluenesulfonic, benzenesulfonic, naphthalenesulfonic, trifluoroacetic and aspartic acids; particularly preferred acids are hydrochloric acid and methanesulfonic acid.
The invention also includes prodrugs of the derivatives of the invention. Prodrugs of the derivatives of the invention are derivatives of formula (I) which may themselves have weak or even no activity, but which, upon administration, are converted under physiological conditions (e.g. by metabolism, solvolysis or otherwise) to the corresponding biologically active form.
The term "halogen" as used herein, unless otherwise indicated, refers to fluoro, chloro or bromo; "alkyl" refers to straight or branched chain alkyl; "cycloalkyl" refers to a substituted or unsubstituted cycloalkyl; "alkoxy" refers to straight or branched chain alkoxy groups; "alkynyl" refers to straight or branched chain alkynyl groups; "heteroaryl" refers to a monocyclic or polycyclic ring system containing one or more heteroatoms selected from N, O, S, the ring system being aromatic, such as imidazolyl, pyridyl, pyrazolyl, furanyl, thienyl, pyrrolyl, thiazolyl, benzothiazolyl, oxazolyl, isoxazolyl, naphthyl, quinolinyl, isoquinolinyl, benzimidazolyl, benzoxazolyl, and the like.
The invention also comprises a pharmaceutical composition which comprises the compound of the general formula I and pharmaceutically acceptable salts and/or solvates thereof as active ingredients and pharmaceutically acceptable carriers; the compounds of the present invention may be used in combination with other active ingredients as long as they do not produce other adverse effects, such as allergic reactions.
Carriers for the pharmaceutical compositions of the present invention are of the usual type available in the pharmaceutical art and include: binders, lubricants, disintegrants, solubilizing agents, diluents, stabilizers, suspending agents, non-coloring agents, flavoring agents, etc. for oral preparations; preservatives, solubilizers, stabilizers and the like for injectable preparations; bases for topical formulations, diluents, lubricants, preservatives, and the like. Pharmaceutical formulations may be administered orally or parenterally (e.g., intravenously, subcutaneously, intraperitoneally, or topically), and if certain drugs are unstable under gastric conditions, they may be formulated as enteric coated tablets.
Through in vitro inhibition activity tests of human breast cancer cells MCF7 and human colon cancer cells HCT 116, the compounds of the invention are found to have significant anti-tumor activity, so that the compounds of the invention can be used for preparing medicaments for treating and/or preventing various cancers, such as breast, lung, liver, kidney, colon, rectum, stomach, prostate, bladder, uterus, pancreas, bone marrow, testis, ovary, lymph, soft tissues, head and neck, thyroid, esophagus cancers, leukemia, neuroblastoma and the like. In particular for preparing medicaments for treating and/or preventing breast cancer and colon cancer.
A "therapeutically effective dose" of a compound of the invention is a dose of the compound that confers a therapeutic effect on the subject being treated, at a reasonable benefit/risk ratio for any drug treatment application. The therapeutic effect may be objective (e.g., measurable by some test or marker) or subjective (e.g., the subject gives an indication or experiences an effect). An effective dose of the compound described above may range from about 0.1mg/Kg to about 500mg/Kg, preferably from about 1mg/Kg to about 50 mg/Kg. Effective dosages will also vary depending upon the route of administration and the possibility of co-use with other agents, however, the daily usage of the compound and the compositions of the invention will be determined by the attending physician for adequate medical reasons. The specific therapeutically effective dose level for some particular patient will depend on a variety of factors, including the disorder being treated and the severity of the luxury; the activity of the specific compound employed; the specific composition employed; patient age, weight, general health, sex, and patient diet; time of administration, route of administration and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or concomitantly with specific compounds; and similar factors well known in medicine.
For single or divided dose administration to humans or other animals, the daily administration of the compounds of the invention may be in a dosage range, for example, from 0.01 to 50mg/Kg body weight or from 0.1 to 25mg/Kg body weight. Typically, the treatment regimen of the present invention comprises administration of a compound of the present invention in a single or multiple dose of from about 10mg to about 1000mg per day to a patient in need of such treatment.
The compounds of the general formula described herein may be administered by injection, intravenously, intraarterially, subcutaneously, intraperitoneally, intramuscularly, or subcutaneously; or orally, buccally, nasally, transmucosally, topically, ocularly or by inhalation, in a dose of from 0.1 to about 500mg/Kg body weight, alternatively in a dose of between 1mg to 1000mg/Kg of single dose, every 4 hours to 120 hours, or according to the needs of the particular drug. The desired or regular therapeutic effect can be achieved by administering an effective amount of the compound or compound composition. Generally, the pharmaceutical compositions of the present invention should be administered from about 1 to about 6 times per day or alternatively, as a continuous infusion. Such administration can be used as a chronic or acute treatment. The dosage of the active ingredient will depend on the host treated and the particular mode of administration, and the active ingredient may be combined with pharmaceutical excipients or carriers to form a single dosage form. Standard formulations will contain from about 5% to about 95% active compound (w/w). Alternatively, such formulations may contain from about 20% to 80% active compound. Lower or higher doses than those recited above may be required. The specific dose and treatment regimen for any particular patient will depend upon a variety of factors, including the activity of the specific compound employed; patient age, weight, general health, sex, and patient diet; time of administration, rate of excretion, drug combination, severity and course of the disease, physiological state or symptoms, patient disposition to the disease, physiological state or symptoms, and judgment of the treating physician.
Upon improvement of the physiological state of the patient, a maintenance dose of a compound, composition or combination of the invention may be administered, if necessary. Subsequently, the dosage or frequency of administration, or both, can be reduced, depending on the symptoms, to a level that maintains an improved physiological state when the symptoms have been alleviated to a desired level.
The examples and preparations provided below further illustrate and exemplify the compounds of the present invention and their methods of preparation. It should be understood that the scope of the following examples and preparations is not intended to limit the scope of the present invention in any way.
The following synthetic schemes describe the preparation of the derivatives of formula I of this invention, all starting materials are prepared by the methods described in these schemes, by methods well known to those of ordinary skill in the art of organic chemistry or are commercially available. All of the final compounds of the present invention are prepared by the methods described in these schemes or by methods analogous thereto, which are well known to those of ordinary skill in the art of organic chemistry. All of the variables used in these schemes are as defined below or in the claims.
The derivatives of general formula I according to the invention, in scheme a, are exemplified by the following compounds: when X is NH; q isR1Is methoxy, m is 6, wherein, the substituent R2Is as defined in the claims.
Route a:
in the route A, an intermediate 101 and an intermediate 102 are subjected to substitution reaction to prepare an intermediate 103, the intermediate 103 is subjected to nitration, buckle ring and chlorination to obtain an intermediate 107, R2The substituted aniline intermediate 107 undergoes a substitution reaction to obtain an intermediate 108, and the intermediate 108 is subjected to the action of hydroxylamine hydrochloride to obtain the compound shown in the general formula I.
Route B:
the derivatives of general formula I according to the invention, in scheme B, are exemplified by the following compounds: when X is O; q isR1Is methoxy, R3Is H, m-6, wherein substituent B is as defined in the claims.
In the route B, the intermediate 107 and the intermediate 201 undergo substitution reaction to prepare an intermediate 202, the intermediate 202 undergoes reduction reaction to obtain an intermediate 203, the intermediate 203 reacts with various types of intermediates B to prepare an intermediate 204, and the intermediate 204 is reacted with hydroxylamine hydrochloride to prepare the compound of the general formula I.
Route C: preparation of intermediate B-1
B is an intermediateR4Is H, Cy is unsubstituted phenyl
Aniline generates an intermediate 301 under the action of sodium nitrite and hydrochloric acid, then reacts with ethyl acetoacetate to generate an intermediate 302, the intermediate 302 generates an intermediate 303 through a buckle ring under the action of DMF-DMA, and then the intermediate B-1 is obtained through hydrolysis.
Route D: preparation of intermediate B-2
B isR5Is H; cy is unsubstituted phenyl
1, 1-cyclopropyl dicarboxylic acid reacts with acetone to produce intermediate 402, and intermediate 402 reacts with aniline to produce intermediate B-2.
Route E: preparation of intermediate B-3
B isR5Is CH3And Cy is unsubstituted phenyl.
Reacting aniline with sodium nitrite hydrochloric acid to generate an intermediate 502, reacting with a raw material 503 to generate an intermediate 504, performing loop-opening reaction on the intermediate 504 under the action of acetic acid/sodium acetate to generate an intermediate 505, and generating 506 under the action of dimethyl sulfate, and finally hydrolyzing to obtain an intermediate B-3.
The specific implementation mode is as follows:
the following examples are intended to illustrate, but not limit, the scope of the invention. The nuclear magnetic resonance hydrogen spectrum of the compound is determined by Bruker ARX-400, and the mass spectrum is determined by a Waters QAB-1837 series quadrupole liquid chromatograph; all reagents used were analytically or chemically pure.
Example 17- ((4- ((3-bromophenyl) amino) -6-methoxyquinolyl-7-yl) oxo) -N-hydroxyheptanamide
1.17- (4-acetyl-2-methoxyphenyl) heptanoic acid ethyl ester (103)
20.0g (121mmol) of 3-methoxy-4-hydroxyacetophenone (101) was dissolved in 200mL of N, N-dimethylacetamide at room temperature, 17.5g (127mmol) of anhydrous potassium carbonate and 30.0g (127mmol) of ethyl 7-bromoheptanoate (102) were added to the reaction mixture, and after the addition, the temperature was raised to 85 ℃ to react for 3 hours. After the reaction, the reaction solution was cooled to room temperature, poured into 500mL of ice water, stirred at room temperature for 20min, filtered, washed with 200mL of water, and dried to obtain 32.9g of a yellow solid with a yield of 84.5%.
1.27- (4-acetyl-2-methoxy-5-nitrophenyl) heptanoic acid ethyl ester (104)
30.0g (93mmol) of intermediate 103 was dissolved in 300mL of dry dichloromethane, the reaction was cooled to-10 deg.C, then 12mL (279mmol) of fuming nitric acid was slowly added dropwise, and the temperature of the reaction was controlled at-5 deg.C for 6 h. After the reaction, the reaction solution was poured into 500mL of ice water, the organic layer was separated, washed with water for 2 times, dried over anhydrous sodium sulfate, and evaporated to dryness to obtain 28.5g of a pale yellow solid with a yield of 83.8%.
1.3 Ethyl (E) -7- (4- (3- (dimethylamino) acryloyl) -2-methoxy-5-nitrophenyl) heptanoate (105)
Intermediate 104(25g, 68mmol) was added to 250mL of toluene, heated to 110 ℃ to completely dissolve intermediate 104, and N, N-dimethylformamide dimethyl acetal (48.5g, 408mmol) was added and heated under reflux for 10 h. After the reaction is finished, the reaction solution is cooled to room temperature and then is put into a cold trap to be stirred, solid is separated out, the filtration is carried out, and a yellow solid 25.0g is obtained after a filter cake is dried, wherein the yield is 87.1%.
1.4 ((6-methoxy-4-oxo-1, 4-dihydroquinolin-7-yl) oxo) heptanoic acid ethyl ester (106)
Intermediate 105(20g, 47mmol) was added to glacial acetic acid (8v/w, 160mL), warmed to 40 ℃ and after intermediate IV was completely dissolved, iron powder (13.2g, 237mmol) was added slowly in portions, warmed to 100 ℃ and reacted for 2h with mechanical stirring. And after the reaction is finished, carrying out suction filtration on the reaction solution while the reaction solution is hot, collecting filtrate, cooling the filtrate to separate out a large amount of solid, and carrying out suction filtration to obtain a khaki solid. Dissolving the filter cake in glacial acetic acid, stirring at 100 ℃ for about 30min, performing suction filtration while the solution is hot, collecting filtrate, cooling the filtrate to precipitate solid, performing suction filtration, washing the filter cake to be neutral, and drying to obtain 10.2g of solid with the yield of 61.8%.
1.57- ((4-chloro-6-methoxyquinolin-7-yl) oxo) heptanoic acid ethyl ester (107)
Intermediate 105(10g, 29mmol), phosphorus oxychloride (5v/w, 50mL) was added to acetonitrile (5v/w, 50mL) and the reaction was refluxed at 85 ℃ for 6 h. After the reaction is finished, the reaction product is decompressed and evaporated to dryness to obtain a gray viscous solid, the gray viscous solid is added into a large amount of ice-water mixed liquid to separate out a solid, the solid is filtered by suction, and a filter cake is washed by water to obtain 9.5g of an off-white solid with the yield of 90.5%.
MS(ESI)m/z:366.35[M+H]+
1.67- ((4- ((3-bromophenyl) amino) -6-methoxyquinolin-7-yl) oxo) heptanoic acid ethyl ester
Intermediate 107(1.0g,2.7mmol) and 3-bromoaniline (0.55g, 3.2mmol) were added to 7mL of isopropanol solution at room temperature and reacted at 90 ℃ for 2h at room temperature. After the reaction, the reaction solution was cooled to room temperature, a solid precipitated, filtered, and the filter cake was washed with water (10mL) and dried to give 1.1g of a white solid with a yield of 81.5%.
1.77- ((4- ((3-bromophenyl) amino) -6-methoxyquinolin-7-yl) oxo-N-hydroxyheptanamide
0.25g (0.5mmol) of ethyl 7- ((4- ((3-bromophenyl) amino) -6-methoxyquinolin-7-yl) oxo) heptanoate was added to 5mL of a hydroxylamine methanol solution (4.2g of hydroxylamine hydrochloride, 2.4g of NaOH, 25mL of methanol) at room temperature, and sodium hydroxide (0.04g, 1.0mmol) was added to the reaction solution under ice bath, and the reaction was allowed to stand at room temperature for 5 hours. After the reaction is finished, 5mL of water and 6mol/L of HCl solution are added into the residue to adjust the pH value to be neutral, and the crude product is obtained after suction filtration and drying. The crude product was purified by column chromatography (dichloromethane: methanol ═ 20:1) to give 0.15g of an off-white solid in 62.5% yield.
The compounds of examples 1-11 (see Table 1) were prepared by the procedure of example 1 starting from intermediate 107 and various substituted anilines.
Example 2: n- (4- ((7- ((7- (hydroxylamine) -7-oxoheptyl) oxo) -6-methoxyquinolin-4-yl) oxo) phenyl) -4-oxo-1-phenyl-1, 4-dihydropyridazine-3-carboxamide
1.7- ((6-methoxy-4- (4-nitrophenyl) quinolin-7-yl) oxo) heptanoic acid ethyl ester (202)
Intermediate 107(10g, 27mmol), 4-nitrophenol (5.6g, 40.5mmol) were added to dry chlorobenzene (5v/w, 50mL) and heated to 140 deg.C for 20 h. After the reaction was completed, the solvent was evaporated to dryness to obtain a gray solid, which was dissolved in dichloromethane and washed with saturated potassium carbonate solution, the organic layer was collected, dried, and the solvent was evaporated to dryness to obtain 6.5g of a solid with a yield of 51.6%.
2.7- ((4- (4-Aminophenoxy) -6-methoxyquinolin-7-yl) oxo) heptanoic acid ethyl ester (203)
Intermediate 202(5.0g,10.7mmol) was dissolved in 50mL of methanol solution at room temperature,10% Pd/C (0.5g) was added and H was bubbled through2And reacting at room temperature for 2 h. After the reaction, the reaction solution was cooled to room temperature, filtered, and evaporated to dryness to obtain a dark gray solid (4.1 g) with a yield of 87.2%. MS (ESI) M/z 439.38[ M + H ]]+
3. Intermediate 4-oxo-1-phenyl-1, 4-dihydropyridazine-3-carboxylic acid (B-1)
3.13-oxo-2- (2-phenylhydrazono) butyric acid ethyl ester (302)
Aniline (5.58g, 60mmol) was added to 50mL of 20% hydrochloric acid solution, cooled to 0 ℃ in an ice-water bath, and 50mL of an aqueous solution of sodium nitrite (5g, 72mmol) was added dropwise to the reaction mixture at a rate controlled to bring the reaction temperature between 0 and 5 ℃. After dripping, the mixture reacts for 30min at the temperature of 0-5 ℃ for standby. Ethyl acetoacetate (8.19g, 63mmol) and anhydrous sodium acetate (14.76g, 180mmol) were added to a mixed solvent of 50mL of water and 200mL of ethanol, cooled to 0 ℃ in an ice-water bath, and then the diazonium salt solution prepared above was slowly dropped while maintaining the reaction temperature between 0 and 5 ℃. After dropping, l h is reacted at 0-5 ℃. After the reaction, the reaction solution was filtered, and the filter cake was washed with water and dried to obtain 12g of pale yellow solid with a yield of 85.7%.
3.24-oxo-1-phenyl-1, 4-dihydropyridazine-3-acetic acid ethyl ester (303)
Intermediate 302(17.8g, 76mmol) and DMF-DMA (18.1g, 152mmol) were added to 200mL of toluene and the reaction was warmed to reflux for 4 h. After the reaction solution was cooled to room temperature, the solvent was evaporated to dryness to obtain a pale yellow viscous substance, and diethyl ether was added to the viscous substance to solidify it to obtain 15g of an off-white solid with a yield of 77.5%.
3.34-oxo-1-phenyl-1, 4-dihydropyridazine-3-carboxylic acid (B-1)
The intermediate 303(12g, 49mmol) was added to 160mL of water, 50mL of 10% aqueous sodium hydroxide solution was added dropwise to the reaction solution, and the reaction was carried out at 50 ℃ for 3 hours. After the reaction, the reaction solution was concentrated, the residue was poured into water, the pH was adjusted to 2 with 15% hydrochloric acid solution, and a solid was precipitated, filtered, and dried to obtain 9.5g of a white solid with a yield of 90.5%.
4.7- ((6-methoxy-4- (4- (4-oxo-1-phenyl-1, 4-dihydropyridazine-3-carboxamide) phenoxy) quinolin-7-yl) oxy) heptanoic acid ethyl ester
Intermediate 203(0.3g, 0.6 mm) was added at room temperatureol), intermediate B-1(0.2g, 0.9mmol), HATU (0.34g, 0.9mmol) and triethylamine (0.09g, 0.9mmol) were added to 20mL of DCM and the reaction was warmed to 40 ℃ for 4 h. After completion of the reaction, the reaction mixture was washed with 100mL of 20% aqueous potassium carbonate solution for 3 times, the organic layer was washed with saturated brine twice, and the separated organic layer was dried over anhydrous sodium sulfate. Methylene chloride was distilled off under reduced pressure, and 0.25g of a yellow oil was obtained in a yield of 65.7%. MS (ESI) M/z 636.58[ M + H ]]+
N- (4- ((7- ((7- (hydroxylamine) -7-oxoheptyl) oxo) -6-methoxyquinolin-4-yl) oxo) phenyl) -4-oxo-1-phenyl-1, 4-dihydropyridazine-3-carboxamide
0.25g (0.39mmol) of ethyl 7- ((6-methoxy-4- (4- (4-oxo-1-phenyl-1, 4-dihydropyridazine-3-carboxamide) phenoxy) quinolin-7-yl) oxy) heptanoate was added to 5mL of a hydroxylamine methanol solution (4.2g of hydroxylamine hydrochloride, 2.4g of NaOH, 25mL of methanol) at room temperature, and sodium hydroxide (0.03g, 0.78mmol) was added to the reaction solution under ice bath, and the reaction was carried out at room temperature for 5 hours after completion of dropwise addition. After the reaction is finished, 5mL of water and 6mol/L of HCl solution are added into the residue to adjust the pH value to be neutral, and the crude product is obtained after suction filtration and drying. The crude product was purified by column chromatography (dichloromethane: methanol ═ 20:1) to give 0.15g of an off-white solid in 62.5% yield.
The compounds of examples 12-39 (see Table 1) were prepared by reacting intermediate 203 with various intermediates B-1 according to the procedure of example 2.
Example 3: (E) -2-benzylidene-N- (4- ((7- ((7- (hydroxylamine) -7-heptyloxy) oxo) -6-methoxyquinolin-4-yl) oxo) phenyl) hydrazine-1-carboxamide
1: 7- ((4- (4- (hydrazinamide) phenoxy) -6-methoxyquinolin-7-yl) oxy) heptanoic acid ethyl ester
Intermediate 203(3.06g,7mmol) and pyridine (1.65g,21mmol) were dissolved in 60mL of dichloromethane, and phenyl chloroformate (2.2g,0.014mol) was added dropwise under ice-bath, followed by reaction at room temperature for 2 h. TLC monitoring reaction is completed, reaction liquid is evaporated to dryness, 25mL of toluene solution and 80% (7.5mL,150mmol) of hydrazine hydrate are directly added, and the temperature is raised to 80 ℃ for reaction for 6 h. TLC monitoring reaction, separating organic layer, washing organic layer 2 times, washing once with saturated sodium chloride, drying with anhydrous sodium sulfate, desolventizing to obtain gray oily matter 2.5g, yield 71.4%. MS (ESI) m-z:497.42[M+H]+
2: (E) -ethyl 7- ((4- (4- (2-benzylidene hydrazide-1-amide) phenoxy) -6-methoxyquinolin-7-yl) oxo) heptanoate
The intermediate ethyl 7- ((4- (4- (hydrazinamide) phenoxy) -6-methoxyquinolin-7-yl) oxy) heptanoate (0.19g, 0.39mmol), benzaldehyde (0.055g, 0.39mmol) and 10mL of ethanol were added sequentially to a 25mL reaction flask. Stirring uniformly, refluxing at 85 ℃ for 1h, monitoring by TLC to finish the reaction, cooling to room temperature, and separating out a solid. Suction filtration and drying gave 0.16 g of an off-white solid in 69.5% yield.
3: (E) -2-benzylidene-N- (4- ((7- ((7- (hydroxylamine) -7-heptyloxy) oxo) -6-methoxyquinolin-4-yl) oxo) phenyl) hydrazine-1-carboxamide
0.15g (0.25mmol) of ethyl (E) -7- ((4- (4- (2-benzylidenehydrazide-1-amide) phenoxy) -6-methoxyquinolin-7-yl) oxo) heptanoate was added to 5mL of a hydroxylamine methanol solution (4.2g of hydroxylamine hydrochloride, 2.4g of NaOH, 25mL of methanol) at room temperature, and sodium hydroxide (0.02g, 0.5mmol) was added to the reaction solution under ice bath, and the reaction was stopped at room temperature for 5 hours. After the reaction is finished, 5mL of water and 6mol/L of HCl solution are added into the residue to adjust the pH value to be neutral, and the crude product is obtained after suction filtration and drying. The crude product was purified by column chromatography (dichloromethane: methanol ═ 20:1) to give 0.10g of an off-white solid in 71.4% yield.
The compounds of examples 40-43 (see Table 1) were prepared according to the procedure above by reacting intermediate 203 with various substituted benzaldehydes, starting from intermediate 3.
Example 4: 1-phenyl-N- (4- ((7- ((7- (hydroxylamine) -7-heptyloxy) oxo) -6-methoxyquinolin-4-yl) oxo) phenyl-2-oxopyrrolidine-3-carboxamide
1.2-oxo-1-phenylpyrrolidine-3-carboxylic acid (B-2)
1.1: 6, 6-dimethyl-5, 7-dioxaspiro [2.5] octane-4, 8-dione
401(10.4g, 80mmol), acetic anhydride (9.6mL, 100mmol) and concentrated sulfuric acid (0.32mL, 6mmol) are mixed uniformly at room temperature, acetone (8mL, 100mmol) is added dropwise to the solution under ice bath, reaction is carried out for 12h at room temperature, after the reaction is finished, the reaction solution is poured into 100mL of ice-water mixture, stirring is carried out for 30min, and suction filtration is carried out, so as to obtain 8.6g of white solid powder, wherein the yield is 63.2%.
1.2: 2-oxo-1-phenylpyrrolidine-3-carboxylic acid (B-2)
Intermediate 6, 6-dimethyl-5, 7-dioxaspiro [2.5] at room temperature]Octane-4, 8-dione (2.0g, 11.7mmol) and aniline (2.2g, 23.5mmol) were added to a 50mL acetonitrile solution, reacted at 60 ℃ for 10h, after completion of the reaction, the reaction solution was evaporated to dryness, and 50mL petroleum ether: pulping the mixed solution of ethyl acetate (3:1), separating out a solid, and performing suction filtration to obtain 1.8g of white powder, wherein the yield is as follows: 75.0 percent. MS (ESI) M/z 206.29[ M + H ]]+。
2.7- ((6-methoxy-4- (4- (2-oxo-1-phenylpyrrolidine-3-carboxamide) phenoxy) quinolin-7-yl) oxy) heptanoic acid ethyl ester
Intermediate 203(0.3g, 0.6mmol), intermediate B-2(0.18g, 0.9mmol), HATU (0.34g, 0.9mmol) and triethylamine (0.09g, 0.9mmol) were added to 20mL of LPCM at room temperature and the reaction was allowed to warm to 40 ℃ for 4 h. After completion of the reaction, the reaction mixture was washed with 100mL of a 20% aqueous potassium carbonate solution for 3 times, the organic layer was washed with saturated brine twice, and the separated organic layer was dried over anhydrous sodium sulfate. Methylene chloride was distilled off under reduced pressure, and 0.27 g of a yellow oil was obtained in a yield of 72.9%.
3.1-phenyl-N- (4- ((7- ((7- (hydroxylamine) -7-heptyloxy) oxo) -6-methoxyquinolin-4-yl) oxo) phenyl-2-oxopyrrolidine-3-carboxamide
0.25g (0.4mmol) of ethyl 7- ((6-methoxy-4- (4- (2-oxo-1-phenylpyrrolidine-3-carboxamide) phenoxy) quinolin-7-yl) oxy) heptanoate was added to 5mL of a hydroxylamine methanol solution (4.2g of hydroxylamine hydrochloride, 2.4g of NaOH, 25mL of methanol) at room temperature, and sodium hydroxide (0.03g, 0.8mmol) was added to the reaction solution under ice bath to react at room temperature for 5 hours at room temperature after completion of dropwise addition. After the reaction is finished, 5mL of water and 6mol/L of HCl solution are added into the residue to adjust the pH value to be neutral, and the crude product is obtained after suction filtration and drying. The crude product was purified by column chromatography (dichloromethane: methanol ═ 20:1) to give 0.18g of an off-white solid in 75.0% yield.
The compounds of examples 43-61 (see Table 1) were prepared according to the procedure above by reaction with various substituted intermediates B-2 starting from intermediate 203 according to the procedure of example 4.
Example 5: n- (3-fluoro-4- ((7- ((7- (hydroxylamine) -7-heptyloxy) oxo) -6-methoxyquinolin-4-yl) oxo) phenyl) -4-methyl-3, 5-dioxo-2-phenyl-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carboxamide
4-methyl-3, 5-dioxo-2-phenyl-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carboxylic acid (B-3)
1.1: (2-cyano-2- (2-phenylhydrazono) acetyl) carbamic acid ethyl ester (504)
Aniline (5.58g, 60mmol) was added to 50mL of 20% hydrochloric acid solution, cooled to 0 ℃ in an ice-water bath, and 50mL of an aqueous solution of sodium nitrite (5g, 72mmol) was added dropwise to the reaction mixture at a rate controlled to bring the reaction temperature between 0 and 5 ℃. After dripping, the mixture reacts for 30min at the temperature of 0-5 ℃ for standby. Intermediate (2-cyanoacetyl) urethane 503(9.36 g, 63mmol) and anhydrous sodium acetate (14.7g, 180mmol) were added to a mixed solvent of 50mL of water and 200mL of ethanol, cooled to 0 ℃ in an ice-water bath, and then the diazonium salt solution prepared above was slowly dropped while maintaining the reaction temperature between 0 and 5 ℃. After dropping, l h is reacted at 0-5 ℃. After the reaction, the reaction solution was filtered, and the filter cake was washed with water and dried to obtain 11.7g of a yellow solid with a yield of 75%.
1.2: 3, 5-dioxo-2-phenyl-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carbonitrile (504)
Intermediate 503(10g, 36mmol), NaOAc (3.2g, 40mmol) were added to 100mL of glacial acetic acid and reacted at 120 ℃ for 5 h. Evaporating most of the solvent to dryness under reduced pressure, cooling to room temperature, adding 40mL of water, stirring for 40min, filtering, and washing the filter cake with water to obtain a light yellow solid. The crude product was purified by column chromatography (eluent PE: EA ═ 1: 2) to give pure 5.1g, 66.2% yield. MS (ESI) M/z 214.7[ M + H ]]+
1.3: 4-methyl-3, 5-dioxo-2-phenyl-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carbonitrile (505)
Intermediate 504(5.0g, 23mmol) was dissolved in DMF, cooled to-10 deg.C and K was added to the reaction in portions2CO3(6.3g, 46mmol), the reaction was stirred for 20 min. Mixing Me with water2SO4(5.8g, 46mmol) was diluted with 10mL of DMF and slowly added dropwise to the reaction mixture, the reaction temperature was kept below-5 ℃ by controlling the dropping rate. After dropping, the reaction was carried out at-5 ℃ for 1.5 hours. Adding water and 5% NaOH solution into the reaction solution in sequence, and adjustingAdjusting the pH to be neutral, stirring for 2h, and performing suction filtration. The filter cake was washed with water to give 3.8g of a pale yellow solid with a yield of 73.1%.
1.4: 4-methyl-3, 5-dioxo-2-phenyl-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carboxylic acid (B-3)
Intermediate 505(3.5g,15mmol) was dissolved in 35mL of glacial acetic acid at room temperature, concentrated hydrochloric acid (5v/m,17.5mL) was added, and the temperature was gradually raised to 100 ℃ for 5 h. Cooling to room temperature, extracting with ethyl acetate, washing with saturated salt water, drying with anhydrous sodium sulfate, filtering, and evaporating to obtain light yellow solid 2.4g with yield 73.8%.
2- ((4- (2-fluoro-4- (4-methyl-3, 5-dioxo-2-phenyl-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-amide) phenoxy) -6-methoxyquinolin-7-yl) oxo) heptanoic acid ethyl ester
0.3g, 0.6mmol of intermediate ethyl 7- ((4- (4-amino-2-fluorophenoxy) -6-methoxyquinolin-7-yl) oxo) heptanoate, 0.3g, 0.9mmol of intermediate B-3(0.22g, 0.9mmol), HATU (0.34g, 0.9mmol) and triethylamine (0.09g, 0.9mmol) were added to 20mL of dichloromethane at room temperature, and the reaction was warmed to 40 ℃ for reaction for 6 h. After completion of the reaction, the reaction mixture was washed with 100mL of a 20% aqueous potassium carbonate solution for 3 times, the organic layer was washed with saturated brine twice, and the separated organic layer was dried over anhydrous sodium sulfate. Methylene chloride was distilled off under reduced pressure, and 0.29g of a yellow oil was obtained in a yield of 72.5%.
N- (3-fluoro-4- ((7- ((7- (hydroxylamine) -7-heptyloxy) oxo) -6-methoxyquinolin-4-yl) oxo) phenyl) -4-methyl-3, 5-dioxo-2-phenyl-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-carboxamide
0.25g (0.36mmol) of ethyl 7- ((4- (2-fluoro-4- (4-methyl-3, 5-dioxo-2-phenyl-2, 3,4, 5-tetrahydro-1, 2, 4-triazine-6-amide) phenoxy) -6-methoxyquinolin-7-yl) oxo) heptanoate was added to 5mL of a hydroxylamine methanol solution (4.2g of hydroxylamine hydrochloride, 2.4g of NaOH, 25mL of methanol) at room temperature, and sodium hydroxide (0.03g, 0.72mmol) was added to the reaction solution under ice bath, and the reaction was carried out at room temperature for 5 hours after completion of dropwise addition. After the reaction is finished, 5mL of water and 6mol/L of HCl solution are added into the residue to adjust the pH value to be neutral, and the crude product is obtained after suction filtration and drying. The crude product was purified by column chromatography (dichloromethane: methanol ═ 20:1) to give 0.13g of an off-white solid in 54.2% yield.
The compound of example 62 (see table 1) was prepared according to the procedure above, starting from intermediate 203 and reacting with various substituted intermediates B-3 according to the procedure of example 5.
Example 6: n-hydroxylamine-7- ((6-methoxy-4- (4- (3- (4- (trifluoromethoxy) phenyl) ureido) phenoxy) quinolin-7-yl) oxy) heptanamide
1: 7- ((6-methoxy-4- (4- (3- (4- (trifluoromethoxy) phenyl) ureido) phenoxy) quinolin-7-yl) oxy) heptanoic acid ethyl ester
Dissolving the intermediate 203(0.3g, 0.6mmol) in dry ethyl acetate at room temperature, then dropwise adding p-trifluoromethoxy isocyanate (0.15g, 0.72mmol), reacting at room temperature for 2h, evaporating the reaction solution after the reaction is finished, adding 10mL of isopropyl ether, pulping, and filtering to obtain 0.3g of off-white solid powder with the yield of 78.9%.
2: n-hydroxylamine-7- ((6-methoxy-4- (4- (3- (4- (trifluoromethoxy) phenyl) ureido) phenoxy) quinolin-7-yl) oxy) heptanamide
Ethyl 7- ((6-methoxy-4- (4- (3- (4- (trifluoromethoxy) phenyl) ureido) phenoxy) quinolin-7-yl) oxy) heptanoate (0.23g, 0.36mmol) was added to 5mL of a hydroxylamine methanol solution (4.2g of hydroxylamine hydrochloride, 2.4g of NaOH, 25mL of methanol) at room temperature, and sodium hydroxide (0.03g, 0.72mmol) was added to the reaction solution under ice bath to react at room temperature for 5 hours at room temperature after completion of dropwise addition. After the reaction is finished, 5mL of water and 6mol/L of HCl solution are added into the residue to adjust the pH value to be neutral, and the crude product is obtained after suction filtration and drying. The crude product was purified by column chromatography (dichloromethane: methanol ═ 20:1) to give 0.11g of an off-white solid in 47.8% yield.
The compounds of examples 63-65 (see Table 1) were prepared by the method of example 6 starting from intermediate 203 and reacting with various substituted isocyanates according to the above reaction procedure.
TABLE 1
The compounds containing a zinc binding group and a quinoline skeleton according to the above formula (I) of the present invention (concentration 0.1. mu.M) were tested for in vitro c-Met and HDAC1 kinase activity. Controls SAHA and cabozantinib were obtained as outsourced reagents.
1. Preparation of 1X buffer (tromethamine buffer)
2. Compounds were diluted according to the respective concentrations:
compounds were transferred to assay plates with 100% DMSO, the final content of DMSO is 1%.
3. Preparation of enzyme solution
The preparation method of HDAC buffer comprises the following steps: firstly, preparing 1M Tris-HCl stock solution, mixing 7.5mL of 1M Tris-HCl stock solution with 0.03654g of EDTA and 7.31g of NaCl, adding distilled water to 450mL, and then adding 50mL of glycerol to obtain 500mLHDAC buffer.
4. Preparing a substrate solution, namely adding trypsin and an ac peptide substrate into 1x detection buffer solution to prepare the substrate solution.
5. mu.L of enzyme solution was transferred to an enzyme conjugate plate.
6. Incubate for 15 minutes at room temperature.
7. To the above solution, 10. mu.L of a substrate solution was added to initiate a reaction.
8. Followed by incubation at room temperature for 60 minutes.
9. Fluorescence intensity was measured at 355nm and 460nm at Synergy MX.
10. And fitting Excel data by using a formula to obtain an inhibition value.
Equation (1): Inh% (Max-Signal)/(Max-Min) × 100
The results of in vitro kinase assays for c-Met and HDAC1 for some of the compounds are shown in table 2.
TABLE 2
In vitro anti-tumor cell activity of the product of the invention
The compound containing the zinc binding group and the quinoline skeleton of the formula I is screened for inhibiting the activity of human breast cancer cells MCF7 and human colon cancer cells HCT 116 in vitro. The control products SAHA and cabozantinib are obtained by using external purchase reagents
1. After cells were thawed and passaged for 2-3 stabilities, they were digested from the bottom of the flask with trypsin solution (0.25%). After pouring the cell digest into the centrifuge tube, the culture medium is added to stop the digestion. Centrifuging the tube at 800r/min for 10min, discarding supernatant, adding 5mL culture solution, blowing and beating the mixed cells, sucking 10 μ LAdding the cell suspension into a cell counting plate for counting, and adjusting the cell concentration to 104Per well. 100. mu.L of the cell suspension was added to the 96-well plate except that the A1 well was a blank well and no cells were added. The 96-well plate was placed in an incubator for 24 h.
2. The test sample was dissolved in 50. mu.L of dimethyl sulfoxide, and then an appropriate amount of culture medium was added to dissolve the sample to 2mg/mL of the liquid, and then the sample was diluted to 20, 4, 0.8, 0.16, 0.032. mu.g/mL in a 24-well plate.
3 wells were added for each concentration, two columns of cells surrounding each, which were greatly affected by the environment, and only used as blank wells. The 96-well plate was placed in an incubator for 72 h.
3. The drug-containing culture medium in the 96-well plate was discarded, the cells were washed twice with Phosphate Buffered Saline (PBS), 100. mu.L of MTT (0.5mg/mL) was added to each well, and the mixture was placed in an incubator for 4 hours, and then the MTT solution was discarded, and 100. mu.L of dimethyl sulfoxide was added thereto. And oscillating on a magnetic oscillator to fully dissolve the viable cells and the MTT reaction product formazan, and putting the formazan into an enzyme labeling instrument to measure the result. Determination of drug IC by Bliss method50The value is obtained.
The results of the partial compounds on inhibiting the activity of human breast cancer cells MCF7 and human colon cancer cells HCT 116 and human lung adenocarcinoma cells A549 are shown in Table 3.
TABLE 3
Preliminary in vitro kinase and cell activity test results show that the compound of the general formula I to be protected has good c-Met and HDAC inhibitory activity, and part of the compound is equivalent to or superior to positive control drugs of cabozantinib and SAHA.
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