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Arsenic trioxide (ATO), a compound clinically proven for acute promyelocytic leukemia (APL), has garnered attention for its emerging efficacy in solid tumors, including HCC. However, the molecular mechanisms driving ATO\u2019s antitumor activity in HCC remain incompletely understood. In this study, we aimed to elucidate the ferroptosis-dependent effects of ATO on HCC and and propose a potential therapeutic strategy.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec002\">\n<jats:title>Methods<\/jats:title>\n<jats:p>The response of HCC cells to ATO was evaluated using cell viability, wound healing, colony formation, Transwell migration assays, and cell cycle analysis. ATO-induced ferroptosis was assessed by measuring lipid peroxidation (via C11-BODIPY staining), intracellular iron levels, and malondialdehyde (MDA) production. Western blotting was performed to quantify protein levels of NRF2, HO-1, SLC7A11, and GPX4; immunofluorescence staining was employed to determine NRF2 subcellular localization.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec003\">\n<jats:title>Results<\/jats:title>\n<jats:p>ATO exhibited significant cytotoxicity and inhibited the progression of HCC cells. Treatment with ATO resulted in a notable increase in lipid ROS and MDA levels, which were subsequently reversed by the ferroptosis inhibitors Fer-1 and DFO. Mechanistically, ATO induced ferroptosis by inhibiting GPX4. Furthermore, NRF2 and its downstream targets, HO-1 and SLC7A11, were upregulated during ferroptosis. NRF2 knockdown enhanced lipid peroxidation and ATO-induced cell death.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec004\">\n<jats:title>Conclusions<\/jats:title>\n<jats:p>ATO significantly promoted ferroptosis in HCC cells, and NRF2 knockdown enhanced the cytotoxic effects of ATO.<\/jats:p>\n<\/jats:sec>","DOI":"10.1371\/journal.pone.0322746","type":"journal-article","created":{"date-parts":[[2025,5,22]],"date-time":"2025-05-22T17:29:56Z","timestamp":1747934996000},"page":"e0322746","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":4,"title":["Silencing NRF2 enhances arsenic trioxide-induced ferroptosis in hepatocellular carcinoma 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