Unlocking the potential of Traditional Chinese Medicine (TCM): Shipi Xiaoji formula (SPXJF) as a novel ferroptosis inducer in hepatocellular carcinoma

ETHNOPHARMACOLOGICAL RELEVANCE: Hepatocellular Carcinoma (HCC) is a major health concern with limited treatment options. Traditional Chinese Medicine (TCM) offers potential therapeutic approaches for HCC, and SPXJF, a TCM formula, has shown promise in clinical observations for prolonging the survival of liver cancer patients. AIM OF THE STUDY: To investigate the anti-tumor effects of SPXJF on HCC cells and explore its potential mechanism, focusing on ferroptosis induction. MATERIALS AND METHODS: LC/Q-TOF-MS was used for compound identification. Cell viability assays, EdU proliferation assay, colony formation assay, wound healing assay, Transwell assay, and Western-blotting were conducted to evaluate the effects of SPXJF on HCC cell proliferation, migration, and invasion. Bioinformatics analysis and RT-PCR were employed to identify potential ferroptosis-related genes and validate the results. Ferroptosis induction was investigated using ferroptosis inhibitors, ROS and lipid peroxidation detection, and TEM. In vivo experiments using a subcutaneous xenograft tumor model confirmed the anti-tumor effects of SPXJF and its ability to induce ferroptosis in HCC. RESULTS: and ROS levels, decreased GSH levels, altered mitochondrial morphology, and upregulation of ferroptosis-inducing proteins ACSL4 and LPCAT3, along with downregulation of ferroptosis-inhibiting proteins xCT and GPX4. Bioinformatics analysis and RT-PCR further identified GSTZ1, CDC25A, AURKA, NOX4, and CAPG as potential ferroptosis-related genes regulated by SPXJF. In vivo experiments confirmed the anti-tumor effects of SPXJF and its ability to induce ferroptosis in HCC. CONCLUSIONS: SPXJF exerts anti-tumor effects on HCC cells by inducing ferroptosis, and its mechanism of action involves the regulation of ferroptosis-related genes and proteins. This study provides a theoretical basis for the clinical treatment of HCC and the development of new anti-cancer drugs, offering a valuable contribution to the field of ethnopharmacology.