EZH2 Suppresses Ferroptosis in Hepatocellular Carcinoma and Reduces Sorafenib Sensitivity Through Epigenetic Regulation of TFR2

Abstract Background: Sorafenib is a first-line drug, and increasing the sensitivity to sorafenib can effectively prolong the time of sorafenib resistance, which has great benefits for the treatment of patients with hepatocellular carcinoma. One of the important mechanisms of sorafenib in treating HCC is to induce cell ferroptosis. Hence, it is of great importance to investigate the mechanisms that increase sorafenib-induced ferroptosis in hepatocellular carcinoma. Methods: Bioinformatic analysis approaches were used to assess EZH2 expression, prognosis and clinical characteristics. IHC staining assays were performed to assess the expression of EZH2 in tumors and normal tissues. CCK-8, EdU staining and colony formation assays were performed to assess cell proliferation. Western blot and q-PCR analysis of ferroptosis-associated genes. As indicators of ferroptosis, intracellular levels of glutathione, malondialdehyde, and iron were measured. ROS detection by the DCFH-DA probe and mitochondrial membrane potential were tested by JC-1. ChIP analysis was used to study histone modification. The antitumor effects of the combination of tazemetostat and sorafenib were investigated in both in vitro and in vivo studies. Results: EZH2 expression is upregulated in HCC and predicts an unfavorable prognosis. Overexpression of EZH2 can promote HCC cell proliferation and reduce ferroptosis. Further analysis showed that EZH2 can increase the modification of H3K37me3 and regulate the expression of TFR2. Reducing RNA polymerase II binding in the promoter region of TFR2 leads to decreased expression of TFR2. The decrease in intracellular iron levels inhibits the occurrence of the Fenton reaction and cell ferroptosis. Knockdown of EZH2 can increase sorafenib sensitivity in hepatocellular carcinoma cells, and tazemetostat, an inhibitor of EZH2, with sorafenib had significant synergistic antitumor effects both in vitro and in vivo Conclusions: EZH2 epigenetically regulates TFR2 expression through H3K27me3 and inhibits the occurrence of ferroptosis. The EZH2 inhibitor tazemetostat synergized with sorafenib and had superior synergistic effects in anticancer therapy in vitro and in vivo.