Liuwei Dihuang pills suppress hepatocellular carcinoma via PI3K/AKT/TP53 pathway: Integrating network pharmacology, metabolomics, and experimental validation.

Hepatocellular carcinoma (HCC) remains a significant global health challenge with limited treatment options. Liuwei Dihuang Pills (LWPs), a classic Traditional Chinese Medicine formula, have demonstrated potential anti-tumor effects; however, their bioactive components and mechanisms of action against HCC remain unclear. Network pharmacology identified 20 core targets of LWPs, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses indicated pro-apoptotic activity via the PI3K/AKT and TP53 pathways. In vivo studies confirmed tumor growth inhibition, biochemical index regulation, and regulation of the PI3K/AKT/TP53 axis. Metabolomics revealed alterations in amino acid and carbohydrate metabolism. In vitro, LWPs-containing serum (LWPs-S) on the PI3K/AKT/TP53 pathway, suppressing Huh7 cell proliferation, migration, and invasion, inducing G1-phase arrest, elevating reactive oxygen species (ROS) levels, and engaging apoptosis-related pathways. Among various extracts, LWPs alcohol extract emerged as the principal anti-HCC fraction. A total of 77 components were identified in LWPs alcohol extract, including 17 blood‑absorbed and 6 tumor‑absorbed plant monomer compounds. Molecular docking demonstrated strong binding affinities of specific monomers from LWPs alcohol extract to PI3K, AKT and TP53. Collectively, these findings elucidate how LWPs promote HCC apoptosis through key pathways and provide experimental evidence supporting the pivotal role of the alcohol extract, offering a scientific basis for the rational development of LWPs.