目的 基于网络药理学结合分子对接技术探讨雷公藤多苷片（TPT）生物碱类成分抗肝癌作用机制以及细胞实验验证TPT生物碱类成分诱导肝癌细胞毒性的作用。方法 利用Swiss Target Prediction和Targetnet数据库预测TPT中9个生物碱类成分［雷公藤晋碱、雷公藤次碱、雷公藤春碱、雷公藤定碱、雷公藤宁碱A、18-O-(3-糠酰)雷公藤春碱、卫矛碱、peritassine A、雷公藤新碱］的作用靶点。借助Genecards、OMIM和TTD数据库检索肝癌的相关靶点，将两者输入Venny在线工具交集得到生物碱类成分治疗肝癌的作用靶点，通过Cytoscape软件构建“雷公藤多苷片-活性成分-靶点-肝癌”相互作用网络。运用String数据库对交集靶点进行蛋白质-蛋白质相互作用（PPI）网络分析，并使用Metascape数据库进行京都基因与基因组百科全书（KEGG）通路和基因本体（GO）富集分析。借助AutoDockTools和PyMol软件对核心靶点及9种生物碱类成分进行分子对接验证。通过体外人肝癌HepG2细胞CCK-8实验验证TPT种9种生物碱的细胞毒作用。结果 经筛选得到TPT生物碱成分潜在作用靶点119个，肝癌疾病靶点1 168个，交集靶点23个。经PPI分析筛选出10个关键靶点，包括EGFR、CASP3、HSP90AA1等，KEGG富集分析主要涉及通路有Pathways in cancer、Prostate cancer、PI3K-Akt signalingpathway等。GO富集发现TPT生物碱类成分促肝癌细胞凋亡与改变细胞形态和调节蛋白激酶活性有关。分子对接显示多种成分与多个关键靶点有较好的结合能力。细胞验证实验发现5种生物碱［雷公藤晋碱、雷公藤次碱、雷公藤宁碱A、18-O-(3-糠酰)雷公藤春碱、peritassine A］对HepG2细胞均具有细胞毒性。结论 TPT中9种生物碱类成分可能通过作用于EGFR、CASP3、HSP90AA1等关键靶点和调节APAF1/CASP9/CASP3信号通路，从而降低肝癌细胞增殖、激活体内内在凋亡途径，最终促进肝癌细胞凋亡，具有潜在的抗肝癌活性。

Objective To investigate the mechanism of anti-liver cancer effects of alkaloidal components of Tripterygium Polyglycoside Tablet (TPT) based on network pharmacology combined with molecular docking technique and to validate the cytotoxicity-inducing effect of TPT alkaloidal components in liver cancer by cellular experiment. Methods The Swiss Target Prediction and Targetnet databases were used to predict the targets of nine alkaloidal components (wilforgine, wilforine, wilfortrine, wilfordine, wilfornine A, triptonine B, euonymine, peritassine A, euonine) in TPT. The targets of liver cancer were retrieved from the Genecards, OMIM and TTD databases, and the targets of alkaloidal components for the treatment of liver cancer was were obtained by inputting the two into the Venny online tool for intersection. The interaction network of "TPT-active ingredient-target-liver cancer" was constructed by Cytoscape software. Protein-protein interaction (PPI) network analysis of the intersecting targets was performed using the String database, Protein-protein interaction (PPI) network analysis of the intersecting targets was performed using the String database, and the enrichment analysis of Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) pathways was performed using the Metascape database. Molecular docking validation of the core targets and nine alkaloidal components was performed by using AutoDockTools and PyMol software. Verification of the cytotoxicity of nine alkaloids of TPT by CCK-8 assay in human liver cancer HepG2 cells in vitro. Results The screening yielded 119 potential targets of TPT alkaloidal components, 1 168 liver cancer disease targets, and 23 intersecting targets. The PPI analysis identified ten key targets, including EGFR, CASP3, HSP90AA1, etc. The KEGG enrichment analysis mainly involved Pathways in cancer, Prostate cancer, PI3K-Akt signaling pathway, etc. The GO enrichment revealed that the TPT alkaloidal components promote apoptosis in liver cancer cells was associated with alteration of cell morphology and regulation of protein kinase activity. The molecular docking showed that multiple components have good binding ability to multiple key targets. Cellular validation experiments showed that five alkaloids (wilforgine, wilforine, wilfornine A, triptonine B, peritassine A) have cytotoxicity to HepG2 cells. Conclusion The nine alkaloidal components of TPT may act on key targets such as EGFR, CASP3, HSP90AA1 and modulate the APAF1/CASP9/CASP3 signaling pathway, thereby reducing cancer cell proliferation, activating the intrinsic apoptotic pathway in vivo and ultimately promoting apoptosis in liver cancer cells, having potential anti-liver cancer activity.

国家自然科学基金资助项目（82174209）