基于网络药理学及实验验证探究通关藤治疗卵巢癌的作用机制

Mechanism of Marsdenia tenacissima against ovarian cancer based on network pharmacology and experimental verification

通过网络药理学、分子对接技术和体外细胞实验,探究通关藤Marsdenia tenacissima治疗卵巢癌(ovarian cancer,OC)的主要活性成分和潜在作用机制。文献检索获取通关藤中活性成分,利用SwissTargetPrediction数据库获得药物的潜在靶点;借助Therapeutic Target Database(TTD)、Online Mendelian Inheritance in Man Database(OMIM)、GeneCards、PharmGKB等数据库检索OC相关靶点,并利用韦恩在线分析工具将药物靶点和疾病靶点取交集;运用Cytoscape软件构建“成分-靶点-疾病”网络,依据其节点度值筛选出核心成分;根据筛选出来的药物与疾病交集靶点,利用STRING平台和Cytoscape软件构建交集靶点的蛋白相互作用(PPI)网络,依据节点度值筛选出核心靶点;借助DAVID数据库对潜在治疗靶点进行GO功能富集和KEGG通路分析;并采用AutoDock等软件对部分活性成分与潜在靶点进行分子对接验证;最后选用SKOV3细胞在体外对通关藤提取物的抗卵巢癌活性进行验证,并依据GO功能和KEGG通路分析结果,选择PI3K/AKT信号通路进行体外实验验证。结果筛选出山柰酚(kaempferol)、11α-O-苯甲酰-12β-O-乙酰通关藤苷元(11α-O-benzoyl-12β-O-acetyltenacigenin B)、苦绳苷元Q(drevogenin Q)等39个有效活性成分,涉及AKT1、VEGFA、EGFR等25个核心靶点,靶点蛋白富集的通路主要为PI3K/AKT信号通路。分子对接结果显示前10位核心成分与前10位核心靶点均有较好的结合能力。体外实验结果表明,通关藤提取物对卵巢癌细胞增殖有显著的抑制作用,可通过线粒体途径诱导卵巢癌细胞凋亡,下调PI3K/AKT信号通路相关蛋白表达。该研究表明通关藤治疗卵巢癌的多成分、多靶点、多途径协同作用的特点,为深入研究其药效物质基础、作用机制以及临床应用提供理论依据。

The present study aimed to explore the main active components and underlying mechanisms of Marsdenia tenacissima in the treatment of ovarian cancer(OC)through network pharmacology,molecular docking,and in vitro cell experiments.The active components of M.tenacissima were obtained from the literature search,and their potential targets were obtained from SwissTargetPrediction.The OC-related targets were retrieved from Therapeutic Target Database(TTD),Online Mendelian Inheritance in Man(OMIM),GeneCards,and PharmGKB.The common targets of the drug and the disease were screened out by Venn diagram.Cytoscape was used to construct an"active component-target-disease"network,and the core components were screened out according to the node degree.The protein-protein interaction(PPI)network of the common targets was constructed by STRING and Cytoscape,and the core targets were screened out according to the node degree.GO and KEGG enrichment analyses of potential therapeutic targets were carried out with DAVID database.Molecular docking was used to determine the binding activity of some active components to key targets by AutoDock.Finally,the anti-OC activity of M.tenacissima extract was verified based on SKOV3 cells in vitro.The PI3K/AKT signaling pathway was selected for in vitro experimental verification according to the results of GO function and KEGG pathway analyses.Network pharmacology results showed that 39 active components,such as kaempferol,11α-O-benzoyl-12β-O-acetyltenacigenin B,and drevogenin Q,were screened out,involving 25 core targets such as AKT1,VEGFA,and EGFR,and the PI3K-AKT signaling pathway was the main pathway of target protein enrichment.The results of molecular docking also showed that the top ten core components showed good binding affinity to the top ten core targets.The results of in vitro experiments showed that M.tenacissima extract could significantly inhibit the proliferation of OC cells,induce apoptosis of OC cells through the mitochondrial pathway,and down-regulate the expression of proteins related to the PI3K/AKT signaling pathway.This study shows that M.tenacissima has the characteristics of multi-component,multi-target,and multi-pathway synergistic effect in the treatment of OC,which provides a theoretical basis for in-depth research on the material basis,mechanism,and clinical application.