Abstract 5723: XL309 is a potent, selective, and orally bioavailable USP1 inhibitor active as monotherapy and in combination with PARP inhibitors or irinotecan

Abstract Background: Ubiquitin-specific protease 1 (USP1) catalyzes selective removal of ubiquitin from its substrate proteins. During translesion synthesis (TLS), USP1 cleaves ubiquitin from proliferating cell nuclear antigen (PCNA), facilitating exchange of TLS polymerases for polymerase delta. Inhibition of USP1 prevents the repair of DNA lesions by TLS, resulting in single-stranded DNA gaps which are synthetically lethal with BRCA1/2 mutations. Poly (ADP-ribose) polymerase (PARP) inhibitors have shown clinical benefit in BRCA-mutated cancers, but some patients do not respond, and many responders develop resistance to treatment. USP1 inhibition could potentially benefit patients by synergizing with PARP inhibition or bypassing resistance mechanisms. We report preclinical data for XL309, a potent and selective orally bioavailable small molecule inhibitor of USP1. Methods: XL309 selectivity was assessed using a DUBprofiler-Cell™ assay. Viability was measured using CellTiter-Glo® in MDA-MB-436 cells. Ubiquitinated PCNA (Ub-PCNA) and PCNA levels after XL309 treatment of cell lines were quantified using global ubiquitination levels and total proteomic analysis (PTMScan and TMT). The antitumor activity of XL309 alone or in combination with saruparib, olaparib, or irinotecan was evaluated in the BRCA1 mutant MDA-MB-436 cell-derived xenograft (CDX) model or in BRCA1/2 mutant patient-derived xenograft (PDX) models. Drug-like properties were assessed, and pharmacokinetic (PK) parameters were determined following administration of XL309 in rat, mouse, and dog. Results: XL309 is a potent inhibitor of USP1 in biochemical and viability assays. XL309 showed selectivity for USP1 versus other ubiquitin-specific proteases, increased Ub-PCNA levels while decreasing PCNA in the global ubiquitination profiling, and exhibited dose-dependent antitumor activity and induction of Ub-PCNA in a CDX model. XL309 exhibited single-agent activity in BRCA1/2 mutant PDX models. XL309 also elicited durable tumor regression in combination with saruparib and irinotecan in a CDX model, and with olaparib in a PDX model. XL309 showed favorable drug-like properties including high permeability in Caco-2 cells. Low clearance and high bioavailability were observed for XL309 in rat, mouse and dog. Preliminary predictions of human PK parameters estimate a low clearance and long half-life for XL309 in humans. Conclusions: XL309 is potentially a best-in-class USP1 inhibitor. PK assessments in conjunction with preclinical antitumor activity suggest that once daily oral administration of XL309 may achieve clinical activity alone or in combination with PARP inhibitors. XL309 is currently under investigation in a phase 1 first-in-human trial as monotherapy and in combination with olaparib in patients with advanced solid tumors (NCT05932862). Citation Format: Alex Charruyer-Reinwald, Aditya Kumar, Jimmy Lai, Scott Simonovich, Christopher Kochansky, Amy DeMicco, Tina Huynh, Avinash Kumar, Amit A. Dhudashiya, Kishore Narayanan, Kiran Aithal, Shailesh V. Jadhav, Anthony Flores, Richard Mansfield, Randy Wu, Jae H. Chang, Andrew Simmons, Paige Mahaney, Lisa D. Belmont. XL309 is a potent, selective, and orally bioavailable USP1 inhibitor active as monotherapy and in combination with PARP inhibitors or irinotecan [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5723.