Abstract The goal of this project is to develop small molecule modulators of RuvBL1/L2 complex as potential therapies for triple negative breast cancer (TNBC) by targeting DNA double-strand breaks (DSB) repair pathways. TNBC is a subgroup of breast cancer and is molecularly characterized by the lack of expression of estrogen receptor (ER), progesterone receptor (PR) or HER2 (human epidermal growth factor receptor 2) amplification. DNA replication stress and reactive oxygen species are prevalent in TNBC cells due to activation of various oncogenes. Thus, TNBC cells constantly generate DSBs. In order for the TNBC cells to survive, these lethal DSBs must be repaired. Accordingly, over the course of development of cancer, TNBC cells have co-evolved efficient DSB repair mechanisms that protect them from the endogenous lethal DSBs. Therefore, targeting DNA repair pathways has been proposed as a potentially powerful strategy to develop novel therapeutics for TNBC. By exploiting this unique feature of endogenous DSBs prevalent only in TNBC cells, such therapeutics can offer selective toxicity to TNBC cells without harming normal cells. We recently identified a novel small molecule lead compound to target RuvBL1/L2 to inhibit DSB repair in TNBC cells. In this application, we propose to further develop this small molecule lead compound for the treatment of TNBC with 3 specific aims. In aim 1, we will study the mechanism of action of this novel lead compound. In aim 2, we will optimize this lead compound using different medicinal chemistry strategies. In aim 3, we will investigate the optimized compounds' anti-TNBC efficacy in different preclinical models alone and in combination with PARP inhibitors.