I-138 (I 138) is a potent, specific small molecule USP1-UAF1 inhibitor with IC50 of 4.1 nM, binds to USP1-UAF1 at an allosteric pocket synergistically with ubiquitin.
I-138 displays exquisite selectivity (>2,000-fold) across a panel of 45 deubiquitinating enzymes (Ubiquigent DUBprofiler) at 10 uM, including the closely related USP12-UAF1 and USP46-UAF1 complexes.
I-138 treatment (500 nM, 4 hours) increased PCNA and FANCD2 monoubiquitination in HAP-1 USP1 WT but not knockout cells, induced the monoubiquitination of FANCD2 and PCNA in MDA-MB-436 cells, also ablated USP1 autocleavage in cells.
I-138 (0-10 uM) selectively caused loss of viability (IC50=50 nM) and replication stress in BRCA1/2 mutant cells (MDA-MB-436 cell), but not the WT breast cancer cell line HCC1954.
I-138 displayed improved potency and physiochemical properties suitable for in vivo studies compared with ML323.
I-138 demonstarted strongly synergistic effect with PARP inhibitor Olaparib in a BRCA1/2-dependent manner in vitro, and in mice bearing MDA-MB-436 tumors.