Background: Approximately 10% of urothelial cancer (UC) patients have defects in the NER pathway and at least another 10% have defects in the HRR pathway. Tumors with HRR or NER defects are more sensitive to platinum-based chemotherapy due to inability repair platinum-induced DNA damage. Two ADCs, sacituzumab govitecan (SG) and enfortumab vedotin (EV) are approved for treatment of metastatic UC. The cytotoxic payload attached to SG and EV are SN-38 and MMAE respectively. SN-38 directly damages DNA through inhibition of Topoisomerase 1 activity during DNA replication whereas MMAE is a potent microtubule disruptor but does not have any direct DNA damaging effects. Given the expanding clinical roles for EV and SG in UC, we wished to test the impact of HRR or NER deficiency on sensitivity to SN-38 and MMAE. We hypothesized that HRR or NER deficiency would result in a larger increase in sensitivity to SN-38 than MMAE. We also hypothesized that SN-38, but not MMAE, would result in synergistic cell killing when delivered with a PARP inhibitor. Methods: We assembled a series of isogenic NER- and HRR-proficient/deficient cell pairs and compared sensitivity to SN-38 and MMAE, as well as other clinically relevant agents including cisplatin, gemcitabine, and the PARP inhibitors olaparib and talazoparib. Results: HRR-deficient bladder cancer cell lines were significantly more sensitive to SN38 than their respective isogenic HRR-proficient counterpart whereas there was no significant difference in MMAE sensitivity between isogenic pairs of HRR-deficient and HRR-proficient cell lines. The largest difference in sensitivity between HRR-proficient vs HRR-deficient cell lines was observed with olaparib and talazoparib respectively. In contrast, NER-deficient cell lines were not more sensitive to SN38 or MMAE than their respective isogenic NER-proficient counterparts. Conclusions: HRR deficiency significantly increases sensitivity of bladder cancer cells to SN-38 but not MMAE. These findings suggest that UC patients with HRR deficiency may be more likely to benefit from treatment with SG than EV, and more generally with an ADC that has a Topoisomerase 1 inhibitor rather than a microtubule disruptor as the cytotoxic payload.