Betaine induces ferroptotic stress by enhancing KEAP1-mediated NRF2 degradation in breast cancer cells.

BACKGROUND

Breast cancer remains a leading cause of cancer-related mortality in women, largely due to therapeutic resistance. Ferroptosis, a regulated form of cell death driven by lipid peroxidation, represents a promising strategy to suppress tumor progression, yet its regulatory mechanisms in breast cancer are not fully defined. This study aimed to investigate the anti-cancer effects of betaine (BET) and its potential involvement in ferroptosis.

METHODS

Network pharmacology, bioinformatics analyses of The Cancer Genome Atlas and Gene Expression Omnibus datasets, and molecular docking were used to identify potential BET targets and construct a BET-target-breast cancer interaction network. Functional assays in MDA-MB-231 and MCF-7 cells evaluated proliferation, migration, oxidative stress, mitochondrial function, and ferroptosis.

RESULTS

BET treatment was associated with inhibition of cell proliferation, clonogenicity, and migration, while increasing reactive oxygen species and lipid peroxidation. Mechanistically, BET enhanced KEAP1-NRF2 interaction, thereby increasing NRF2 ubiquitination and proteasomal degradation. Consequently, NRF2 nuclear accumulation was reduced, leading to suppression of downstream antioxidant and ferroptosis-protective targets, including xCT and GPX4. The resulting depletion of glutathione and accumulation of lipid peroxidation products promoted ferroptosis-associated cellular damage. BET also disrupted mitochondrial membrane potential and oxidative phosphorylation, further exacerbating oxidative stress. Notably, the KEAP1 R483S mutation or NRF2 overexpression partially rescued BET-induced oxidative stress and ferroptosis-associated phenotypes, supporting a role for enhanced KEAP1-dependent NRF2 turnover in mediating BET activity.

CONCLUSIONS

BET may promote ferroptosis-associated cell death and suppress breast cancer cell malignant phenotypes, potentially through modulation of the KEAP1/NRF2/xCT/GPX4 pathway. These findings suggest a possible therapeutic relevance of BET in breast cancer, although further validation in additional in vivo models and clinical studies is required.