Discovery and Optimization of Novel Tricyclic Ubiquitin-Specific Protease 1 Inhibitors for the Treatment of BRCA-Mutated Breast Cancer

Ubiquitin-specific protease 1 (USP1) represents an emerging therapeutic target for BRCA-deficient malignancies. Using a scaffold-hopping strategy derived from <b>KSQ-4279</b>, we designed a novel series of USP1 inhibitors featuring a tricyclic core. Among them, two lead compounds <b>43</b> and <b>47</b> were identified with potent USP1 inhibitory and cellular antiproliferative activity. Further studies in MDA-MB-436 cells demonstrated that compounds <b>43</b> and <b>47</b> suppressed colony formation and induced S-phase arrest, with time- and concentration-dependently stabilizing ubiquitinated PCNA and amplifying <i>p</i>-H2AX. Importantly, both compounds showed synergistic antiproliferative effects in combination with the PARP inhibitor Olaparib. Supported by its favorable pharmacokinetic properties, compound <b>43</b> exhibited significant <i>in vivo</i> anticancer efficacy in an MDA-MB-436 xenograft model, achieving superior tumor growth inhibition both as monotherapy and in combination with Olaparib compared to <b>KSQ-4279</b>. Collectively, compound <b>43</b> emerges as a promising preclinical candidate with translational potential for BRCA-mutated breast cancer.