以c-Met和ALK为靶点的新型小分子抑制剂的设计合成及生物活性初步研究

发布时间：2018-05-12 18:34

本文选题：ALK + c-Met　；参考：《南京中医药大学》2017年硕士论文

【摘要】：c-Met和ALK蛋白同属于受体酪氨酸激酶,研究表明:HGF/c-Met或ALK信号通路的异常激活是导致多种恶性肿瘤(如:胃癌、肺癌、肝癌、卵巢癌、乳腺癌、神经母细胞瘤、间变性细胞淋巴瘤等)的发生、发展以及继发性转移的重要原因。因此,以c-Met和ALK为靶点的抗癌药物是近几年的研究热点。克里唑替尼(crizotinib)是一种新型ATP竞争性小分子ALK和c-Met双靶点抑制剂,并被FDA批准治疗ALK+局部晚期或转移性非小细胞肺癌,其结构仍具有研究价值。因此以克里唑替尼的结构为基础,设计并合成两个新型系列的化合物并评价其对特定人源肿瘤细胞株的抑制活性。这两个系列的化合物分别以2-氨基吡嗪和3,6-二氯哒嗪为起始原料,分别经过卤代反应、与拆分得到的(R)-1-(2,6-二氯-3-氟苯基)乙醇发生Williamson成醚反应,在钯催化剂和碘化亚铜催化下发生Sonogashira反应以及发生选择性高的Huisgen-Click反应,最后在三氟乙酸作用下脱去Boc保护基得到目标产物。随后,对目标化合物进行体外抗肿瘤活性研究,评价其对人源神经母细胞瘤(SH-SY5Y)和胃癌细胞株(SNU-5)的抑制活性。此外,还运用Discovery Studio 4.0软件考察所合成的化合物与靶标蛋白ALK和c-Met的结合模式。抗肿瘤增殖结果表明:(1)吡嗪类化合物对两个肿瘤细胞株的抑制活性比哒嗪系列好;(2)吡嗪类化合物2-10a对SH-SY5Y的抑制活性最好(IC50=0.467 μmol·L-1),与阳性对照克里唑替尼活性相似(IC50= 0.341μmol·L-1);而化合物2-10e对SNU-5的抑制活性最好(148 nmol·L-1);(3)哒嗪系列化合物对SH-SY5Y的抗肿瘤活性不理想,但是对SNU-5细胞株显示了较强的抑制活性。其中3-9a对两种细胞株的抑制活性最高。同时,对两个系列化合物的构效关系进行了分析:(1)吡嗪类化合物1,2,3-三氮唑下缘取代基为环烷烃时活性比直链烷好,且环烷烃取代基活性顺序为五元环六元环四元环,对SNU-5细胞株抑制活性考察发现1,2,3-三氮唑下缘取代基构型为R时活性比S好;(2)哒嗪类化合物1,2,3-三氮唑下缘取代基为环烷烃时活性同样比直链烷好,对SH-SY5Y细胞株抑制活性分析发现1,2,3-三氮唑上取代基为五元环时活性最好,六元环次之,四元环最差。对SNU-5细胞株的抑制活性分析发现环烷烃取代基活性顺序为六元环五元环四元环,且取代基为R构型时比S构型活性好。最后,运用Discovery Studio 4.0软件模拟了所合成化合物与靶标蛋白ALK和c-Me活性位点的对接,并利用打分软件对对接结果进行打分,结果表明所测试的抗肿瘤活性结果与分子对接的结果基本一致。
[Abstract]:Both c-Met and ALK protein belong to receptor tyrosine kinase. The abnormal activation of the c-Met or ALK signaling pathway at 1: HGF / Met is responsible for many kinds of malignant tumors (such as gastric cancer, lung cancer, liver cancer, ovarian cancer, breast cancer, neuroblastoma, cancer of breast, neuroblastoma). Anaplastic cell lymphoma, et al.) the occurrence, development and secondary metastasis of important causes. Therefore, anticancer drugs targeting c-Met and ALK have been the focus of research in recent years. Crizotinib is a new type of ATP competitive small molecule ALK and c-Met double target inhibitor and has been approved by FDA for the treatment of ALK locally advanced or metastatic non-small cell lung cancer. Therefore, based on the structure of Crizoltini, two new series of compounds were designed and synthesized and their inhibitory activities against specific human tumor cell lines were evaluated. The two series of compounds, starting from 2-aminopyrazine and 3-tri-6-dichloropyridazine, were halogenated, respectively, and reacted with Williamson to form ether. The Sonogashira reaction and the high selectivity Huisgen-Click reaction took place under the catalysis of palladium and cuprous iodide. Finally, the target product was obtained by removing the Boc protection group under the action of trifluoroacetic acid. Then the antitumor activity of the target compound was studied in vitro to evaluate its inhibitory activity on human neuroblastoma SH-SY5Y) and gastric cancer cell line SNU-5). In addition, the binding patterns of the synthesized compounds to target proteins ALK and c-Met were investigated by Discovery Studio 4.0 software. The results of anti-tumor proliferation showed that the inhibitory activity of pyrazines on two tumor cell lines was higher than that on pyridazines from 2-10 years. The inhibitory activity of pyrazines on SH-SY5Y was the best (IC50467 渭 mol L -1), similar to that of crizoltini (0.341 渭 mol L -1). The inhibitory activity of compound 2-10e to SNU-5 was the best, and the antitumor activity of compound 2-10e to SH-SY5Y was not ideal. But it showed strong inhibitory activity on SNU-5 cell line. The inhibitory activity of 3-9 a on the two cell lines was the highest. At the same time, the structure-activity relationship of the two series of compounds was analyzed.) the activity of pyrazines was better than that of straight chain alkanes when the lower edge of pyrazines was cycloalkanes, and the order of activity of cycloalkyl substituents was quaternary ring. The inhibitory activity of SNU-5 cell line was also better than that of linear alkane when the lower edge substituent configuration of 1o 2H 3- triazole was R than that of S H 2) pyridazines. The inhibitory activity of SH-SY5Y cell line was found to be the best when the substituent group on the 1 titrazole was a quaternary ring, followed by a six-member ring and a quaternary ring. The inhibitory activity analysis of SNU-5 cell line showed that the sequence of cycloalkane substituent activity was six-member ring quaternary ring, and the substitution group was better than S configuration when the substituent was R configuration. Finally, Discovery Studio 4.0 software was used to simulate the docking of the synthesized compounds with the target protein ALK and c-Me active sites, and the results of the docking were scored by scoring software. The results showed that the results of anti-tumor activity were consistent with those of molecular docking.
【学位授予单位】：南京中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R914;R96

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