NRF2-Driven CARM1 Promotes Malignant Progression and FerroptosisResistance in Breast Cancer

BACKGROUND: CARM1 has attracted significant attention due to its role in carcinogenesis across a variety of cancers. Ferroptosis has emerged as a prospective target in anticancer therapy. OBJECTIVE: This study aimed to investigate the role of CARM1 in TNBC progression and ferroptosis regulation, as well as to clarify the underlying molecular mechanism involving NRF2-mediated transcriptional activation of CARM1. METHODS: This study combined bioinformatics analysis, in vitro experiments, and in vivo validation to investigate the role of the NRF2-CARM1 axis in TNBC. Public datasets (UCSC Xena, GEPIA2) were analyzed for CARM1/NRF2 expression, correlation, and prognosis. CARM1 and NRF2 were manipulated in BC cells using siRNA/shRNA and overexpression plasmids. Proliferation, migration, and invasion were assessed by CCK-8, colony formation, wound healing, and Transwell assays. Cell apoptosis was assessed via flow cytometry and immunoblotting analysis. Ferroptosis was quantified by lipid peroxidation (TBARS), lipid ROS (BODIPY 581/591 C11), and total iron levels, together with immunoblotting of ferroptosis-related molecules. NRF2 binding to the CARM1 promoter was examined by ChIP and luciferase reporter assays using wild-type (WT) and NRF2-binding-site mutant (MUT) promoter constructs. A xenograft model was used to validate in vivo relevance. RESULTS: CARM1 and NRF2 were elevated in BC tissues and cells. CARM1 silencing inhibited proliferation, migration, and invasion in BC cells while facilitating cell apoptosis and ferroptosis, as evidenced by increased lipid peroxidation, lipid ROS accumulation, and iron levels. NRF2 positively regulated CARM1 expression and enhanced WT CARM1 promoter activity, whereas mutation of the NRF2-binding site abolished this effect. ChIP further confirmed enrichment of the CARM1 promoter region in NRF2 immunoprecipitates, supporting direct transcriptional activation of CARM1 by NRF2. Functionally, NRF2 overexpression partially rescued the anti-tumor and pro-ferroptotic effects caused by CARM1 depletion. In vivo, CARM1 knockdown inhibited tumor growth and enhanced lipid peroxidation, which was partially reversed by NRF2 overexpression. DISCUSSION: These findings identify a novel NRF2-CARM1 regulatory axis involved in TNBC progression and ferroptosis resistance. NRF2 directly activates CARM1 transcription, thereby promoting malignant behaviors and suppressing ferroptotic cell death in TNBC cells. These results improve the understanding of TNBC biology and suggest that the NRF2- CARM1 pathway may serve as a potential therapeutic target for ferroptosis-based treatment strategies. CONCLUSION: NRF2 directly binds to and transcriptionally activates CARM1, thereby enhancing ferroptosis resistance and promoting TNBC progression. These findings reveal a novel molecular mechanism underlying TNBC malignancy and suggest that targeting the NRF2-CARM1 axis, particularly in combination with ferroptosis inducers, may provide a potential strategy for TNBC treatment.