人参皂苷Rh2靶向结合ERβ募集NCOA1激活ERβ-TNFα通路的抗肿瘤机制研究

人参为传统名贵中药材，人参皂苷是其主要活性成分，其中人参皂苷Rh2（G-Rh2）抗肿瘤活性最强。课题组前期发现G-Rh2能上调ERβ，激活TNFα诱导肿瘤细胞凋亡，并获得国家发明专利，但其抗肿瘤药物靶点及上游信号机制不明。预实验提示G-Rh2与ERβ的配体结合结构域互作，促进ERβ募集转录共激活因子NCOA1，敲低ERβ或NCOA1可阻断G-Rh2激活TNFα诱导的肿瘤细胞凋亡。据此提出：G-Rh2靶向结合ERβ募集NCOA1激活ERβ-TNFα通路发挥广谱抗肿瘤作用。本项目将应用SPR与蛋白质共结晶技术解析G-Rh2与ERβ结合位点；使用CRISPR-Cas9和过表达技术阐释G-Rh2靶向ERβ募集NCOA1的机制；采用ChIP及荧光素酶报告活性分析及小鼠移植瘤模型阐明ERβ-NCOA1介导G-Rh2上调ERβ与TNFα的抗肿瘤机制，为G-Rh2抗肿瘤临床应用提供重要理论与科学依据。

Ginseng is one of the most valuable traditional Chinese medicines, Ginsenosides are the main active ingredients. Among them, Ginsenoside Rh2 (G-Rh2) has the strongest anti-tumor activity. Previously, we found that G-Rh2 could up-regulate ERβ, activate TNFα to induce cancer cell apoptosis. However, the anti-cancer targets and upstream signaling mechanism of G-Rh2 were not clear. Evidence showed that G-Rh2 and ERβ ligand binding domains interacted, promoting ERβ recruitment and transcripted co-activator NCOA1. Knocking down ERβ or NCOA1 could block G-Rh2 activating TNFα-induced various cancer cells apoptosis. Therefore, we put forward that G-Rh2 targets ERβ via activation of ERβ-TNFα pathway by recruiting NCOA1. In this project, SPR and small molecule-protein co-crystallization technology will be used to analyze the binding sites of G-Rh2 and ERβ, CRISPR-Cas9 and overexpression technology will be used to explain the mechanism of G-Rh2 targeting ERβ to recruit NCOA1, and then ChIP , luciferase reporter gene activity assay and Xenograft model will be used to elucidate the anti-tumor mechanism of ERβ-NCOA1 mediated G-Rh2 up-regulating ERβ and TNFα expression. Our study will provide important proof of concept for the clinical anti-cancer application of G-Rh2.