Sanguinarine exerts anti-hepatocellular carcinoma activity by targeting FDX1 to induce FDX1/LIAS/DLAT/HSP70 axis-dependent cuproptosis

Hepatocellular carcinoma (HCC), the predominant type of primary liver cancer, represents an extremely aggressive malignancy. The induction of cuproptosis has developed into a favorable therapeutic direction for HCC, considering its strong association with HCC. Sanguinarine (San), a benzophenanthridine alkaloid derived from traditional herbs such as Chelidonium majus L., demonstrates broad-spectrum anticancer activities against various cancer cell types. However, the precise molecular mechanisms underlying its effects in the treatment of HCC remain largely undefined. This investigation seeks to examine the anti-HCC effects of San and to explore the mechanisms underlying these effects through the induction of cuproptosis. In vitro experiments demonstrate that San markedly inhibits the proliferation, movement, and epithelial-mesenchymal transition of HCC cells while enhancing their apoptosis. In vivo, San notably impedes tumor growth and upregulates the cuproptosis signature markers ferredoxin 1 (FDX1), oligomeric dihydrolipoamide S-acetyltransferase (DLAT), and heat shock protein 70 (HSP70) in HCC xenograft tumor models. Mechanistically, San induces proteotoxic stress and cuproptosis in HCC cells by increasing copper concentration, upregulating the expression of FDX1, lipoic acid synthetase (LIAS), HSP70, and lipoylated DLAT aggregation, and simultaneously reducing mitochondrial membrane potential and intracellular glutathione and pyruvate levels. Moreover, the combination of San with copper ionophores (Elesclomol-CuCl₂) exhibits synergistic effects in promoting cuproptosis. FDX1 silencing markedly diminishes San-induced suppression of cell proliferation and FDX1 and HSP70 levels in HCC cells. Additionally, molecular docking analysis predicts that San exhibits the highest potential for binding with FDX1. Surface plasmon resonance experiments and cellular thermal shift assay confirm that San strongly interacts with FDX1 and markedly enhances the thermostability of FDX1. In conclusion, our findings indicate that San substantially inhibits the progression of HCC by targeting FDX1/LIAS/DLAT/HSP70 axis-dependent cuproptosis.