作用于SHP2变构位点2的抑制剂设计、合成及活性研究

【作者】 陈玉梅；

【导师】 周志旭；

【作者基本信息】 贵州大学 ， 药物化学， 2022， 硕士

【摘要】 SHP2是目前唯一被证实的原癌蛋白,参与调控多个癌症相关过程。目前对SHP2小分子抑制剂进行了大量的研发,主要分为催化位点抑制剂和变构抑制剂。由于PTP催化结构域的高度保守序列和带正电荷的性质,开发成药性好的SHP2催化位点抑制剂仍然非常困难。变构抑制剂是一种通过非催化位点选择性靶向SHP2的新方法。目前已报道的SHP2变构位点有4个,针对变构位点1的抑制剂研究较多,而作用于变构位点2的抑制剂目前只报道了成药性较差的SHP244及其衍生物,变构位点3和变构位点4暂无抑制剂报道。本课题选择SHP244化合物作为先导化合物,对其结构进行设计和改造,以期获得高选择性和高活性的化合物。本课题以SHP244结构为基础,针对溶解度低和活性差的缺陷,对其母核三唑并喹唑啉酮环和侧链进行改造,设计了A、B、C三个系列新型的目标化合物。通过分子对接表明所设计的化合物与SHP2蛋白均有相互作用。通过逆合成分析法设计出一条可操作性和合理性的合成路线,该合成路线最终以2-氨基烟酸作为原料,通过酰胺化、环合、取代、还原胺化等多种有机反应类型制备了30个目标化合物,其结构经¹H NMR、¹³C NMR和HRMS确证。以化合物SHP244作为阳性对照药,采用MTT法对30个化合物测试其对人肺腺癌细胞A549和人前列腺细胞PC-3的体外细胞抗增殖活性。初筛结果表明所有化合物对这两种细胞均有一定抑制活性。大多数化合物对A549细胞的抑制率高于阳性化合物SHP244,其中化合物B2和C2的抑制率相对较高。化合物B2>C5>A7>B1对PC-3细胞的抑制率高于阳性化合物SHP244。选取目标化合物B2测试其在人和大鼠肝微粒体中的代谢稳定性,结果表明化合物B2的代谢不是很稳定,猜测由于化合物B2结构中引入了多种胺结构导致在代谢过程中易氧化,从而导致其代谢不稳定。构效关系初步表明:（1）在C部分引入1-（4-溴苯基）吡咯烷片段片段或在D部分引入乙醇胺基团有利于提高化合物对A549细胞的抑制活性;（2）在D部分引入环己胺基团有利于提高化合物对A549细胞和PC-3细胞的抑制活性;（3）目标化合物中含有邻甲氧基苯环片段有利于提高化合物对PC-3细胞的抑制活性。更多还原

【Abstract】 SHP2 was currently the only confirmed proto-oncoprotein involved in the regulation of multiple cancer-related processes.At present,a large number of small molecule inhibitors of SHP2 have been developed,which were mainly divided into catalytic site inhibitors and allosteric inhibitors.Due to the highly conserved sequence and positively charged nature of the catalytic domain of PTP,the development of druggable SHP2 catalytic site inhibitors is still very difficult.Allosteric inhibitors are a novel approach to selectively target SHP2 through a non-catalytic site.At present,4allosteric sites of SHP2 have been reported,and there are many studies on the inhibitors targeting allosteric site 1.However,SHP244 and its derivatives with poor drugability have been reported for inhibitors acting on allosteric site 2,and no inhibitors has been reported for allosteric site 3 and allosteric site 4.In this project,the SHP244 compound was selected as the lead compound,and its structure was designed and modified in order to obtain the compounds with high selectivity and high activity.Based on the structure of SHP244,aiming at the defects of low solubility and poor activity,three series of new target compounds were designed by modifying the triazoloquinazolinone ring and side chain of its parent nucleus.Molecular docking showed that the designed compounds interacted with SHP2 protein.An operable and rational synthetic route was designed by retrosynthetic analysis.The synthetic route was finally prepared by using 2-aminonicotinic acid as a raw material through various organic reaction types such as amidation,cyclization,substitution,and reductive amination.30 target compounds were obtained,and their structures were confirmed by ¹H NMR,¹³C NMR and HRMS.Using compound SHP244 as a positive control drug,30 compounds were evaluated their antiproliferative activity against A549 cells（human lung adenocarcinoma cells）and PC-3 cells（human prostate cells）by MTT assay in vitro.Bioassay results showed that most compounds demonstrated good inhibitory effect against A549 cells and PC-3 cells.Especially,compounds B2 and C2 demonstrated the best inhibitory effect against A549 cells,which were even better than the positive compound SHP244.Compounds B2,C5,A7 and B1 had a better inhibitory effect against PC-3 cells than the positive compound SHP244.The target compound B2 was selected to test its metabolic stability in human and rat liver microsomes.The results showed that the metabolism of compound B2 was not very stable.It is speculated that the introduction of a variety of amine structures into the structure of compound B2leads to easy oxidation during the metabolic process,resulting in its metabolic instability.The preliminary structure-activity relationship showed that some interesting things.Firstly,the inhibition rates of the compounds that containing1-（4-bromophenyl）pyrrolidine fragment in the C part or ethanolamine group in the D part were generally higher than other compounds against A549 cells.What’s more,the inhibition rates of the compounds that containing cyclohexylamine group in the D part were generally higher than other compounds.Finally,Compounds containing o-methoxybenzene ring fragment had generally higher inhibition rates against PC-3cells than other compounds.更多还原