抗肿瘤干细胞药物研究——选择性mTOR抑制剂的设计、合成与筛选

Cancer stem cells are kind of cells that have the characteristics of stem cells in cancer. They are intensively tumorigenic and transferrable, and resistant to conventional chemotherapy ordinarily. Selective mTOR inhibitors can inhibit the proliferation of cancer stem cells effectively, and possess the characteristics of strong targeting and low toxicity, which make it have great potential for development. The methods to improve the selectivity of these compounds summarize has been summarized and a serise of highly selective inhibitor of mTOR has been designed through analyzing the crystal structure of the protein mTOR in latest reports and researching the reports on the structural features of mTOR inhibitors over years. Then, the compounds has been optimized and synthesised by means of computer-aided drug design such as virtual docking, ADMET property prediction tools. Five different tumor cell systems are used in this project, and by the technology such as Western Blot, a set of comprehensive bio-activity evaluation system in binding enzyme levels, cellular level and molecular level has been done to quickly and efficiently evaluate the compounds on the selectively, the ability of inhibiting tumor stem cells, the molecular mechanism of action and the safety. A whole set of design, synthesis and screening system on mTOR inhibitors has been built, which not only provides a drug candidate for the inhibition of tumor stem cells and treatment of cancer, but also provides a reference for the study of other kinase inhibitors.

肿瘤干细胞是肿瘤中具有干细胞特性的一类细胞，它们的致瘤和转移能力极强，并且往往对传统化疗药物具有耐药性。选择性mTOR抑制剂可有效的抑制肿瘤干细胞的增殖，并且具有靶向性强和毒性小的特点，具有很大的发展潜力。通过分析最新报道的mTOR蛋白晶体结构，并总结历年来报道的mTOR抑制剂结构特点，我们归纳出提高该类化合物选择性的方法，设计出一组高选择性的mTOR抑制剂；然后借助分子对接、ADMET性质预测等计算机辅助药物设计手段优选出化合物，并进行合成。 本项目基于5种不同的肿瘤细胞系，运用Western Blot等技术，建立了一套结合酶水平、细胞水平、分子水平的生物活性综合评价体系，可对化合物的选择性、抑制肿瘤干细胞能力、分子作用机制、安全性进行快速高效评测。 本项目构建的mTOR抑制剂的设计、合成和筛选体系，不仅能为抑制肿瘤干细胞及治疗癌症提供候选药物，还可为其它激酶抑制剂的研究提供参考。

肿瘤干细胞在多种肿瘤的发生、发展中都扮演重要角色，抑制肿瘤干细胞是治疗肿瘤的关键所在。近年来，人们发现mTOR在肿瘤干细胞的生长，增殖中扮演重要角色，开发高效低毒的mTOR抑制剂已成为抑制肿瘤干细胞的新策略。. 本研究以开发新型mTOR抑制剂抑制肿瘤干细胞为目标。首先，基于对mTOR蛋白三维晶体结构的深入分析，确定了决定mTOR抑制剂活性和选择性的关键氨基酸残基VAL2240、ASP2195、GLN2194和LYS2187。然后，运用基于片段的虚拟设计技术，设计了包含四大结构类型的小分子化合物库。经过虚拟筛选，挑选出83个化合物进行合成。最后，对这些已获得的新结构化合物进行了分子作用机制，酶抑制活性，肿瘤干细胞抑制活性和动物体内肿瘤抑制活性方面的生物活性评价。经过多轮筛选，最后得到两个优选化合物CS-71（mTOR IC50=2.45）和CS-74（mTOR IC50=7.73），这两个化合物对mTOR具有高度选择性，而相对于PI3K达到100倍以上。细胞实验和动物实验均证明以上两个化合物具有较好的抗肿瘤干细胞活性。该项目的开展为肿瘤的治疗提供了较好的候选试剂，同时，建立了一套包含分子水平、酶水平、细胞水平和动物水平生物活性测试的抗肿瘤干细胞活性测试平台，该平台的搭建可为后续大规模分子活性筛选提供高效支持，具有较高的理论和应用价值。