Polybromo-1(PBRM1) loss of function mutations are present in a fraction of biliary tract cancers (BTCs).PBRM1, a subunit of the PBAF chromatin-remodeling complex, is involved in DNA damage repair. Herein, we aimed to decipher the molecular landscape ofPBRM1mutated (mut) BTCs and to define potential translational aspects. Totally, 1848 BTC samples were analyzed using next-generation DNA-sequencing and immunohistochemistry (Caris Life Sciences, Phoenix, AZ). siRNA-mediated knockdown ofPBRM1was performed in the BTC cell line EGI1 to assess the therapeutic vulnerabilities of ATR and PARP inhibitors in vitro.PBRM1mutations were identified in 8.1% (n= 150) of BTCs and were more prevalent in intrahepatic BTCs (9.9%) compared to gallbladder cancers (6.0%) or extrahepatic BTCs (4.5%). Higher rates of co-mutations in chromatin-remodeling genes (e.g.,ARID1A31% vs. 16%) and DNA damage repair genes (e.g.,ATRX4.4% vs. 0.3%) were detected inPBRM1-mutated (mut) vs.PBRM1-wildtype (wt) BTCs. No difference in real-world overall survival was observed betweenPBRM1-mut andPBRM1-wt patients (HR 1.043, 95% CI 0.821–1.325,p= 0.731). In vitro, experiments suggested that PARP ± ATR inhibitors induce synthetic lethality in thePBRM1knockdown BTC model. Our findings served as the scientific rationale for PARP inhibition in a heavily pretreatedPBRM1-mut BTC patient, which induced disease control. This study represents the largest and most extensive molecular profiling study ofPBRM1-mut BTCs, which in vitro sensitizes to DNA damage repair inhibiting compounds. Our findings might serve as a rationale for future testing of PARP/ATR inhibitors inPBRM1-mut BTCs.