人参皂苷CP-80选择性抑制IDH2/R140Q突变体的抗急性髓性白血病作用与机制研究

Acute myeloid leukemia (AML) is a kind of malignant tumor that seriously harms human health. In recent years, isocitrate dehydrogenase mutant IDH2/R140Q has been found to have close relationship with the initiation and maintenance of AML. It is demonstrated that the inhibition of IDH2/R140Q can induce cellular differentiation. However, no effective IDH2/R140Q inhibitors are available in the clinic. Many of the active ingredients in ginseng exhibit anti-tumor activity. In the previous study, the applicant discovered a ginsenoside CP-80 from the natural products database through high-throughput virtual screening, which showed high inhibitory activity (IC50) against IDH2/R140Q at nanomolar level, but no activity against the wild-type IDH. Based on these existing results, we plan to study the selectivity of CP-80 to other enzymes, and combine molecular simulation and experimental site-directed mutagenesis to explore its inhibitory mechanism on IDH2/R140Q at molecular level. Then, further investigations about CP-80 including its pharmacological activity on TF-1 cells and effect on the cell proliferation and differentiation of primary AML patient samples will be carried out at cellular level. Finally, the security of CP-80 in vivo will also be estimated. This project will not only provide a fundamental clue in using ginsenoside CP-80 as a therapeutic intervention against AML, but also facilitate the optimization of lead compounds and discovery of new IDH2/R140Q inhibitors.

急性髓性白血病（AML）是一类严重危害人类健康的恶性肿瘤。近年发现，异柠檬酸脱氢酶突变体IDH2/R140Q与AML的发生发展密切相关，抑制IDH2/R140Q活性能够诱导AML细胞分化，但是临床上尚无有效抑制剂。人参中许多活性成分具有抗肿瘤作用，申请人前期通过高通量虚拟筛选，从天然产物数据库中获得一个对IDH2/R140Q的抑制活性（IC50）在nM级水平而对野生型IDH无活性的抑制剂——人参皂苷CP-80。本项目在既有研究基础上，拟从分子水平考察CP-80对其他酶的选择性，并结合分子模拟和实验定点突变技术研究其对IDH2/R140Q的抑制机理；然后从细胞水平研究CP-80对TF-1细胞的药理活性、对临床AML病人样本细胞增殖和分化的影响；最后评价CP-80在动物体内的安全性。本项目将为人参皂苷CP-80在抗AML中的应用奠定基础，为先导化合物的改造及新型抑制剂的发现提供一定的科学依据。

急性髓性白血病（AML）是一类严重危害人类健康的恶性肿瘤。近年发现，异柠檬酸脱氢酶突变体IDH2/R140Q与AML的发生发展密切相关，抑制IDH2/R140Q活性能够诱导AML细胞分化，因此，IDH2突变体是治疗AML的有效靶点。本项目通过计算机辅助高通量虚拟筛选方法，从商业化合物数据库中获得一个对IDH2/R140Q的抑制活性（IC50）在nM级水平（140 nM）而对野生型IDH几乎无活性的抑制剂CP80，其选择性远高于已知抑制剂AGI-6780和Enasidenib。分子模拟结果表明，CP80结合在IDH2/R140Q双亚基的界面处，通过别构调节作用将此酶锁定在一个失活的开放构象，降低了辅酶NADPH与酶的结合自由能，从而发挥抑制活性。细胞实验证明CP80能够降低TF-1(IDH2/R140Q) 细胞的2-HG水平，抑制细胞增殖并能诱导TF-1(IDH2/R140Q) 细胞分化，上调细胞红化相关的血红蛋白和 Kruppel 样因子的表达。然而CP80 在小鼠体内并不稳定，需要进一步做结构改造。本项目的实施为CP80的后续结构改造提供数据支持，为新型抗AML抑制剂的研发奠定基础。