High-throughput screening identified a small molecule compound that induces ferroptosis and alleviates invasion and migration in hepatocellular carcinoma cells by targeting the STAT3/ GPX4 axis

Hepatocellular carcinoma (HCC) is a lethal malignancy. Despite considerable efforts in recent years, effective therapeutic drugs for HCC are still lacking. In this study, polyphyllin VI was identified as a potential therapeutic drug for HCC by screening natural herbal compounds. Cell proliferation was assessed using cell counting Kit-8, colony formation, lactate dehydrogenase release, and PI staining assays. The occurrence of ferroptosis was determined by assessing the accumulation of lipid reactive oxygen species, malondialdehyde, and intracellular ferrous iron. Wound healing and transwell assays were performed to detect migration and invasion abilities. Our study found that polyphyllin VI inhibited the proliferation, invasion, and metastasis of HCCLM3 and Huh7 cells by inducing ferroptosis. In addition, through network pharmacology and molecular docking analyses, we found that polyphyllin VI may target signal transducer and activator of transcription 3 (STAT3), and immunofluorescence revealed that polyphyllin VI significantly inhibited the nuclear translocation of P-STAT3 in HCCLM3 and Huh7 cells. HCCLM3 and Huh7 cells were treated with a STAT3 inhibitor (Stattic), which inhibited glutathione peroxidase 4 (GPX4) expression, promoted oxidative stress and ferroptosis, and inhibited invasion and metastasis. Overall, our findings suggest that polyphyllin VI may inhibit STAT3 phosphorylation, which in turn inhibits GPX4 expression, increases iron-dependent accumulation and lipid peroxidation, and induces ferroptosis in HCCLM3 and Huh7 cells, eventually inhibiting their invasion and metastasis. These data suggest that polyphyllin VI may be a candidate for the prevention and treatment of HCC.