Isoform-specific function of NSD3 in DNA replication stress confers resistance to PARP inhibitors in prostate cancer

Poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) have been approved for the treatment of metastatic castration-resistant prostate cancer (mCRPC) in patients with deleterious BRCA1/2 alterations. Although this marks a significant milestone, intrinsic or acquired therapy resistance remains a major challenge that limits clinical efficacy. Here, we demonstrate that dysregulated ubiquitination and turnover by the cullin 3 (CUL3)^ZBTB2 E3 ligase complex induce the upregulation of the short isoform of nuclear-receptor-binding SET domain protein 3 (NSD3) (NSD3S), which confers PARPi resistance in prostate cancer cells and patient-derived mCRPC samples. Mechanistically, ATR drives the localization of NSD3S at stalled replication forks, where it antagonizes the PTIP-dependent recruitment of the MRE11 nuclease, thereby protecting nascent DNA from extensive degradation and ensuring fork stabilization. Importantly, pharmacological degradation of NSD3S using an NSD3-targeting proteolysis-targeting chimera (PROTAC) efficiently enhances PARPi sensitivity in both cell-line-derived xenograft and patient-derived xenograft (PDX) mouse models. These findings establish NSD3S as a key determinant of PARPi toxicity in mCRPC.