Bufalin Induces Ferroptosis in Triple-Negative Breast Cancer via Inhibition of the NRF2/HO-1/GPX4 Pathway

Triple-negative breast cancer (TNBC), a highly aggressive subtype of breast cancer characterized by poor prognosis and limited therapeutic options, remains a major clinical challenge. Bufalin, a principal bioactive component of Venenum Bufonis, exhibits potent anticancer activity against breast cancer. Nevertheless, the precise molecular mechanism by which bufalin triggers ferroptosis in TNBC cells remains poorly elucidated. In the present study, both in vitro and in vivo investigations were performed. Cellular ferroptosis was comprehensively evaluated by detecting intracellular Fe[Formula: see text], ROS, MDA, and reduced GSH levels, together with ultrastructural observation under transmission electron microscopy. Transcriptome sequencing combined with bioinformatic analysis was applied to screen the core regulatory mechanism of bufalin-induced ferroptosis in TNBC, and subsequent molecular biological experiments were conducted for further validation. The results demonstrated that bufalin significantly suppressed the proliferation and migration of TNBC cells. Bufalin treatment markedly increased Fe[Formula: see text], ROS, and MDA levels, depleted intracellular GSH, and induced mitochondrial shrinkage and cristae damage. Transcriptome analysis identified the NRF2/HO-1/GPX4 signaling axis as the central pathway mediating the pharmacological effect of bufalin. Mechanistically, bufalin downregulated the expression of NRF2, HO-1, SLC7A11, and GPX4, whereas it upregulated KEAP1 expression. In vivo studies verified that bufalin effectively inhibited tumor growth with no obvious systemic toxicity. Moreover, bufalin decreased the expression of GPX4 and Ki-67, increased the number of TUNEL-positive cells, and blocked the activation of the NRF2/HO-1/GPX4 pathway in tumor tissues. In conclusion, bufalin induces ferroptosis in TNBC by inhibiting the NRF2/HO-1/GPX4 signaling pathway, thereby exerting antitumor effects. These findings indicate that bufalin serves as a promising therapeutic agent for TNBC treatment.