USP7 inhibitor HBX19818 increased PARP1, protein PARylation, and nuclear apoptosis-inducing factor. (A) HBX19818 treatment (6 hours) elevates PARP1 and PARylated protein levels in the absence of caspase-3 cleavage (cl-CASP3) in both quiescent (without CD40L/IL-21) and proliferating (with CD40L/IL-21) primary CLL cells. CD40L/IL-21–stimulated proliferation was confirmed by the presence of cyclin A expression. SMC1 was the loading control. (B) Analysis of nuclear extracts from proliferating primary CLL cells demonstrated that HBX19818-induced translocation of apoptosis-inducing factor (tAIF) accumulation in the nucleus. (C) HBX19818 treatment for 72 hours induced CLL cells to swell and disperse their contents (arrows), morphological changes that are consistent with necrotic cell death. (D) In ATM/p53-defective primary CLL induced to proliferate (n = 6) the PARP inhibitor talazoparib did not potentiate the cytotoxic activity (left) of HBX19818 and acted in an antagonistic manner, as indicated by combination indices (right).