基于网络药理学、分子对接及体外实验探究木香烃内酯抗胃癌的作用及机制

Integrating Network Pharmacology, Molecular Docking and In Vitro Experiments to explore the pharmacological mechanisms of Costunolide against Gastric Cancer

关键词:  胃癌  木香烃内酯  网络药理学  PI3K/AKT 信号通路

Key words:  gastric cancer  costunolide  network pharmacology  PI3K/AKT signaling pathway

摘要:

目的 本研究旨在通过网络药理学分析结合实验验证，探讨藏木香（Inula racemosa Hook.f.）活性成分木香烃内酯（Costunolide，COS）抗胃癌（gastric cancer，GC）的药理作用及机制。方法 采用网络药理学方法，从五个数据库（即pharmMapper、SwissTargetPrediction、TargetNet、CTD和Super-PRED）中筛选COS的潜在靶点。利用GeneCards和OMIM数据库获取与GC相关的靶点。并利用Metascape数据库进行GO和KEGG富集分析。采用Autodock Vina 1.5.6对COS与靶向蛋白进行分子对接。通过CCK-8实验、集落形成实验及迁移实验，验证COS对胃癌细胞增殖和迁移能力的影响。同时采用流式细胞术和线粒体膜电位检测其对凋亡的影响，并利用qPCR和Western blot验证其作用机制。结果 网络药理学共筛选出100个潜在靶点，其中EGFR、CASP3、TNF、HSP90AA1、ESR1为5个核心靶点，PI3K/AKT信号通路为核心通路。体外实验结果显示，COS抑制了胃癌细胞AGS、HGC-27、NCI-N87的增殖，同时促进了HGC-27细胞的凋亡。qPCR和Western blot的结果还显示，COS可降低p-PI3K和p-AKT的mRNA及蛋白表达水平，并通过上调BAX、下调Bcl-2以及激活Caspase-3来诱导HGC-27细胞凋亡。结论 COS通过影响PI3K/AKT信号通路，抑制胃癌细胞增殖并促进其凋亡，从而发挥抗肿瘤作用。

Abstract:

OBJECTIVE To elucidate the pharmacological mechanisms underlying the anti-gastric cancer (GC) effects of costunolide (COS), an active sesquiterpene lactone isolated from Inula racemosa Hook.f., through integrative network pharmacology and experimental validation. METHODS Potential targets of COS were predicted from five databases (PharmMapper, SwissTargetPrediction, TargetNet, CTD, and Super-PRED). GC-related targets were retrieved from GeneCards and OMIM databases. GO and KEGG enrichment analyses were performed using Metascape. Molecular docking of COS with core targets was conducted using AutoDock Vina 1.5.6. The anti-proliferative and anti-migratory effects of COS on GC cells were evaluated by CCK-8 assay, colony formation assay, and wound healing assay. Apoptosis was assessed by Hoechst 33342 staining, Annexin V-FITC/PI flow cytometry, and JC-1 mitochondrial membrane potential assay. The underlying mechanisms were validated by qPCR and Western blot. RESULTS Network pharmacology identified 100 potential targets, with EGFR, CASP3, TNF, HSP90AA1, and ESR1 as the top five hub targets. The PI3K/AKT signaling pathway was identified as the most significantly enriched pathway. In vitro experimental results show that COS inhibited the proliferation of the gastric cancer cell lines AGS, HGC-27, and NCI-N87, while promoting apoptosis in HGC-27 cells. Mechanistically, COS downregulated p-PI3K and p-AKT at both mRNA and protein levels, upregulated BAX, downregulated Bcl-2, and activated cleaved caspase-3. CONCLUSION COS exerts anti-GC effects by inhibiting cell proliferation and inducing apoptosis through suppression of the PI3K/AKT signaling pathway.