Mechanism and in Vitro Experiment of Wogonin in Treatment of Hepatocellular Carcinoma Based on Network Pharmacology

Abstract

Background Hepatocellular carcinoma (HCC) is the leading cause of cancer-related deaths. The current prevention and treatment situation remains critical. It is of scientific significance to explore new therapeutic agents for HCC. Objective To analyze the mechanism of wogonin on HCC by network pharmacology and to verify it in vitro. Methods The drug targets of wogonin were searched in TCMSP database, and the disease targets of HCC were collected from TTD, GenCard, OMIM, DisGent databases. The collected drug targets and disease targets were intersected as potential targets for drug intervention in diseases. R software was used for enrichment analysis of intersection targets, STRING database and Cytoscape software were used to construct protein interaction network and screen core targets. The core targets were further analyzed in GIEPA database. Finally, the preliminary analysis results were verified by in vitro experiments, including cell activity determination using CCK-8 kit, cell proliferation determination using plate clone formation experiment, cell migration determination using scoring test, protein expression level determination using Western-blotting (WB) assay. Results The AMDE characteristics of wogonin were found to be in accordance with the rules for small molecule drug formation and the toxicity analysis showed no toxicity. A total of 135 wogonin targets and 8 238 HCC targets were collected, and 113 targets were intersected. Through the analysis of the core genes of TOP10 screened by the constructed protein interaction network, it was found that the mRNA levels of CDK1 and SRC in liver cancer tissues were higher than those in normal liver tissues (P<0.05), and the high expression levels in liver cancer patients were related to poor prognosis (P<0.05). KEGG enrichment analysis showed that the intersection genes were enriched in the PI3K/AKT signaling pathway, and the molecular docking results showed that wogonin had strong binding configuration activity with CDK1 and SRC. The results of CCK-8 kit showed that the activity of HepG2 cells in the 75.0, 150.0, and 300.0 μmol/L wogonin groups was lower than that in the control group (P<0.05). The results of plate clone formation experiment showed that the number of colony formation of HepG2 cells in the 37.5, 75.0, 150.0 μmol/L wogonin groups was lower than that in the control group (P<0.05). The results of scoring test showed that the migration rate of HepG2 cells in the 37.5, 75.0 and 150.0 μmol/L wogonin groups was lower than that in the control group (P<0.05). The results of the WB assay showed that the expression levels of PI3K, P-AKT/AKT, CDK1 and SRC proteins in the 75.0 and 150.0 μmol/L wogonin groups were lower than those in the control group (P<0.05) . Conclusion Wogonin inhibits the proliferation and migration of HCC cells and induces apoptosis by down-regulating the expression of CDK1 and SRC proteins and attenuating the PI3K/AKT pathway signaling, to achieve the purpose of interfering with the occurrence and progression of HCC.

Keywords

• carcinoma, hepatocellular|wogonin|network pharmacology|in vitro experiments