Ubiquitin-mediated degradation of ANP32A by Realgar licenses ferritinophagic ferroptosis in TNBC stem cells through an ATG5-dependent epigenetic program

Therapy-resistant cancer stem cells (CSCs) drive the poor prognosis of triple-negative breast cancer (TNBC). While inducing ferroptosis is a promising strategy, the upstream epigenetic checkpoints preventing it in CSCs are unknown. The mechanism of the traditional medicine Realgar against TNBC CSCs remains elusive. We integrated phenotypic screening in vitro and in vivo, quantitative proteomics, and mechanistic dissection using ubiquitination assays, immunoprecipitation, and pathway-specific genetic/pharmacological perturbations. Realgar selectively eradicated TNBC CSCs and suppressed tumor progression. Proteomics identified the epigenetic repressor ANP32A as its key downregulated target. Realgar promoted ubiquitin-mediated degradation of ANP32A via ubiquitination at K20. ANP32A degradation dismantled the INHAT repressor complex and enhanced CBP/p300 recruitment, relieving epigenetic repression at the ATG5 promoter through increased H3K27 acetylation. The resulting ATG5 upregulation triggered a ferritinophagic program, leading to iron overload, lipid peroxidation, and CSC ferroptosis. Genetic blockade of ANP32A degradation or ATG5 function abolished this cascade, establishing an ANP32A-ATG5-ferritinophagy axis. Our study has identified ANP32A as novel epigenetic target in TNBC that is degraded by Realgar and is essential for CSC survival. This discovery reveals a druggable axis (ANP32A–H3K27Ac–ATG5–ferritinophagy) and positions ANP32A as a candidate prognostic marker and a potential therapeutic target, thereby bridging traditional medicine and precision oncology in the suppression of TNBC stemness.