Modulating G6PD/PGD to overcome FSP1/DHODH-mediated ferroptosis defence: A novel oridonin derivative suppresses liver cancer

Background: Hepatocellular carcinoma (HCC), a globally prevalent malignancy with high mortality rates, presents an unmet need for innovative effective therapies. Purpose: This study aimed to explore the antitumour potential of compound XD, a novel oridonin derivative, on HCC and its underlying mechanism. Experimental Approach: The antitumour effects of compound XD were investigated in several HCC cells lines and mice models. The mechanism of XD was investigated using FACS, qPCR, WB, ELISA, IHC, siRNA and plasmid transfection. Key Results: Compound XD demonstrated potent inhibitory effects, surpassing sorafenib with a maximum of 10-fold lower IC₅₀ values against HCC cell lines. Its anticancer activities were ferroptosis dependent, which could be attenuated by ferroptosis inhibitors including deferoxamine, ferrostatin-1 and N-acetyl-cysteine. Unlike sorafenib, XD decreased two pivotal regulator FSP1 and DHODH to induce ferroptosis, while their overexpression partially mitigated XD-induced cytotoxicity and lipid peroxidation. In addition, XD treatment decreased cellular NADPH levels and inhibited the expression of G6PD and PGD in NADPH generation. Overexpression of G6PD or PGD reversed FSP1 and DHODH down-regulation, rescuing the ferroptosis induced by XD. Bioinformation analysis indicated the significant up-regulation of G6PD and PGD in clinical HCC patients and was positively correlated with cancer stages. Molecular docking and CETSA assay confirmed the binding capacity of XD with G6PD and PGD protein. Finally, XD dose-dependently inhibited liver tumour growth and induced ferroptosis-related markers in mice. Conclusion and Implications: This study suggests XD as a potential ferroptosis inducer and the potential role of G6PD/PGD/FSP1/DHODH axis in governing ferroptosis sensitivity in HCC.