Integrative bioinformatics and experimental validation reveal quercetin as a potential multi-target therapeutic agent...

Hepatocellular carcinoma (HCC) is the most common form of primary liver cancer worldwide, with increasing incidence and mortality rates. Although several targeted therapies are currently available, the therapeutic outcomes remain unsatisfactory due to the high heterogeneity and drug resistance of HCC. Therefore, novel molecular mechanisms and therapeutic strategies urgently need to be explored. In this study, we obtained the GSE39791 dataset from the GEO database and identified 1,186 differentially expressed genes (DEGs). Weighted gene co-expression network analysis (WGCNA) was conducted to obtain 776 key module genes, which were intersected with 11,671 HCC-related genes from the GeneCards database, resulting in 226 candidate genes. A protein-protein interaction (PPI) network was constructed using the STRING database, and the top 20 hub genes were identified using the MNC algorithm in Cytoscape. Among these, the five most significant hub genes-RFC4, TOP2A, AURKA, HSP90AA1, and MCM4-were selected for further analysis. KEGG enrichment analysis was performed to explore their functional pathways. Potential therapeutic agents were predicted using the CMap database, and molecular docking was conducted via AutoDock Vina. To validate the computational predictions, a quercetin intervention model was established. The optimal dose was determined through CCK-8 assays in HepG2 cells, and the expression of the five hub genes was examined in normal liver cells (LO2), HepG2 cells, and HepG2 cells treated with quercetin using RT-qPCR. The five hub genes-RFC4, TOP2A, AURKA, HSP90AA1, and MCM4-were significantly overexpressed in both HCC tissues and cell lines. Enrichment analysis revealed that these genes were mainly involved in cancer-related pathways, including the cell cycle, p53 signaling pathway, and FoxO signaling pathway. Drug prediction analysis showed that quercetin exhibited a negative regulatory pattern with respect to HCC and displayed binding energies below - 5 kcal/mol with all five hub proteins. CCK-8 assays confirmed the dose-dependent inhibitory effect of quercetin on HepG2 cell viability. RT-qPCR results demonstrated that quercetin significantly downregulated the expression of the five hub genes, consistent with the bioinformatics predictions. This study integrated multi-omics analysis and experimental validation to identify five core genes closely associated with HCC and suggested that quercetin may exert anti-HCC effects partly associated with the regulation of these genes. Our findings offer new insights into the molecular mechanisms of HCC and provide a promising strategy for the development of targeted therapeutics.

The online version contains supplementary material available at 10.1007/s10616-026-00993-x.