UNLABELLED: Chromosomal instability (CIN) and epigenetic reprogramming are central drivers of breast cancer progression, yet the mechanisms connecting them remain elusive. Here, we uncover a direct role for EZH2 histone methyltransferase in promoting CIN in triple-negative breast cancer. Across breast cancers, EZH2 expression correlates with copy-number alterations, and its catalytic activity is associated with increased CIN in metastasis-initiating cells. Pharmacologic EZH2 inhibition suppresses CIN, revealing an unexpected vulnerability. Integrated chromatin and transcriptome profiling identified tankyrase (TNKS), a PARP, as a direct transcriptional target of EZH2. Mechanistically, EZH2-mediated TNKS suppression disrupts centrosomal P4.1-associated protein (CPAP), driving centrosome overduplication, multipolar mitosis, and exacerbated CIN. In vivo, CIN suppression is a critical mechanism underlying the antimetastatic effects of EZH2 inhibition. These findings delineate a previously unrecognized epigenetic mechanism governing CIN and establish EZH2 inhibitors as the first therapeutic agents capable of directly suppressing CIN, underscoring the need for trials with metastasis-focused endpoints.

SIGNIFICANCE: We elucidate epigenetic regulation of CIN through EZH2-TNKS-CPAP-axis and show that CIN suppression is important for the efficacy of EZH2 inhibition on metastasis. These mechanistic insights are informative for developing CIN-suppressing therapies.