Research on the mechanism of eugenol in the treatment of liver cancer based on network pharmacology, molecular docking technology, and in vitro experiments

Eugenol, a phenolic natural product with diverse pharmacological activities, remains unexplored in liver cancer. Using network pharmacology, we investigated eugenol's therapeutic mechanisms in liver cancer. We obtained eugenol's molecular structure from PubChem and screened its targets using similarity ensemble approach in Swiss Target Predictiondatabases. Overlapping genes with liver cancer-related genes from GeneCards were identified. Protein-protein interaction networks, Gene Ontology annotations, and Kyoto Encyclopedia of Genes and Genomes pathway analyses were conducted. A target-pathway network revealed eugenol's interaction with 122 liver cancer-related genes. Molecular docking confirmed eugenol's high affinity for mitochondrial nicotinamide adenine dinucleotide, reduced form (NADH) dehydrogenase 1 (MT-ND1), AKT1, NDUFB7, and NADH dehydrogenase (complex I) subunit S3 (NDUFS3). Expression levels of these targets in normal liver and liver cancer tissues were examined using GEPIA2 and HPA databases. The CCK-8 assay and colony formation assay demonstrated that eugenol significantly inhibited the proliferation of hepatocellular carcinoma cells. Western blot analysis confirmed that eugenol upregulated MT-ND1 while downregulating the expression of targets such as AKT1, NDUFB7, and NDUFS3. Furthermore, it was found that eugenol could influence the expression of the AKT1 target through the AKT/p70 S6K pathway. This study provides new insights into the potential mechanisms of eugenol in liver cancer and offers novel perspectives for network-based liver cancer research.