Slow Dissociation from the PARP1-HPF1 Complex Drives Inhibitor Potency

PARP1, upon binding to damaged DNA, is activated to performpolyADP-ribosylation (PARylation) on itself and other proteins, whichleads to relaxation of chromatin and recruitment of DNA repair factors.HPF1 was recently discovered as a protein cofactor of PARP1 that directspreferential PARylation of histones over other targets by contributingto and altering the PARP1 active site. Inhibitors of PARP1 (PARPi)are used in the treatment of BRCA-/- cancers, but thebasis for their potency in cells, especially in the context of HPF1,is not fully understood. Here, we demonstrate the simple one-stepassociation for eight different PARPi to PARP1 with measured ratesof association (k (on)) of 0.8-6 & mu;M-1 s(-1). We find only minor differencesin these on rates when comparing PARP1 with the PARP1-HPF1complex. By characterizing the rates of dissociation (k (off)) and the binding constants (K (D)) for two more recently discovered PARPi, we find, for example,that saruparib has a half-life for dissociation of 22.5 h and fluzoparibhas higher affinity for PARP1 in the presence of HPF1, just like thestructurally related compound olaparib. By using the measured K (D) and k (on) to calculate k (off), we find that the potency of PARPi in cellscorrelates best with the k (off) from thePARP1-HPF1 complex. Our data suggest that dissociation of adrug compound from the PARP1-HPF1 complex should be the parameterof choice for guiding the development of next-generation PARPi.

Automated summary

Upon binding to DNA damage, PARP1 is activated to perform PARylation on itself and other proteins, leading to chromatin relaxation and DNA repair factor recruitment. HPF1 is a cofactor of PARP1 that directs preferential PARylation of histones. Understanding the dissociation of PARPi from the PARP1-HPF1 complex is important for the development of next-generation PARP inhibitors.