Ebselen sensitizes multidrug-resistant cancer cells to doxorubicin by disrupting cellular redox homeostasis and inducing ferroptosis.

AIM: Multidrug resistance (MDR), driven by P-glycoprotein (P-gp/ABCB1)-mediated drug efflux and enhanced antioxidant defenses, remains a major obstacle to effective chemotherapy. This study investigated whether thioredoxin reductase (TrxR) inhibition by ebselen induces ferroptosis, suppresses NRF2/Keap1 signaling and restores doxorubicin sensitivity in MDR cancer cells and xenograft models. MATERIALS AND METHODS: MDR KB ChR 8-5 and MCF-7/ADR cells were treated with ebselen, doxorubicin, or their combination. TrxR inhibition was evaluated by molecular docking and DTNB assay. Ferroptosis was assessed by clonogenic survival, combination index analysis, oxidative DNA damage (8-OHdG), intracellular and mitochondrial Fe2+ accumulation, lipid peroxidation, aconitase activity, RT-qPCR, western blotting and immunofluorescence. Deferoxamine and ferrostatin-1 rescue experiments confirmed ferroptosis involvement. Antitumor efficacy and systemic safety were further evaluated in a KB ChR 8-5 xenograft mouse model. KEY FINDINGS: Ebselen directly inhibited TrxR and synergistically enhanced doxorubicin cytotoxicity (CI < 1), significantly reducing clonogenic survival and increasing oxidative DNA damage. Combination treatment promoted intracellular and mitochondrial Fe2+ accumulation, aconitase inactivation and lipid peroxidation, while upregulating Keap1, suppressing NRF2 nuclear translocation, downregulating GPX4, GCLC, HO1, NQO1, FTH1 and ABCB1/P-gp and increasing IRP1 and TFR1 expression, thereby enhancing intracellular doxorubicin retention. In vivo, combination therapy markedly suppressed tumor growth without significant systemic toxicity. SIGNIFICANCE: Ebselen sensitizes MDR cancer cells to doxorubicin by targeting the TrxR-NRF2/Keap1 axis, disrupting redox homeostasis, inducing iron-dependent ferroptosis and suppressing drug efflux. These findings identify TrxR as a promising therapeutic target for overcoming chemoresistance.