PARP1，在结合损伤的DNA时被激活，进行对自身和其他蛋白质的多聚ADP-核糖基化（PARylation），导致染色质松弛和DNA修复因子的招募。HPF1最近被发现作为PARP1的蛋白质辅因子，通过促进并改变PARP1的活性位点，将PARylation优先作用于组蛋白而非其他靶蛋白。PARP1抑制剂（PARPi）被用于治疗BRCA-/-癌症，但在细胞中的有效性，尤其是在HPF1的背景下，其基础机制尚未完全理解。在这里，我们展示了8种不同PARPi与PARP1之间的简单单步结合，其结合速率（kon）在0.8-6 μM-1 s-1之间。当比较PARP1和PARP1-HPF1复合物时，我们发现这些结合速率只有微小的差异。通过表征两种最近发现的PARPi的解离速率（koff）和结合常数（KD），我们发现，例如，saruparib的解离半衰期为22.5小时，而fluzoparib在HPF1存在的情况下对PARP1具有更高的亲和力，就像结构相关的olaparib一样。通过使用测定的KD和kon计算koff，我们发现PARPi在细胞中的效力最好与PARP1-HPF1复合物的koff相关。我们的数据表明，药物化合物从PARP1-HPF1复合物中解离应是指导下一代PARPi开发的优选参数。

PARP1, upon binding to damaged DNA, is activated to perform poly ADP-ribosylation (PARylation) on itself and other proteins, which leads to relaxation of chromatin and recruitment of DNA repair factors. HPF1 was recently discovered as a protein cofactor of PARP1 that directs preferential PARylation of histones over other targets by contributing to and altering the PARP1 active site. Inhibitors of PARP1 (PARPi) are used in the treatment of BRCA-/- cancers, but the basis for their potency in cells, especially in the context of HPF1, is not fully understood. Here, we demonstrate the simple one-step association for eight different PARPi to PARP1 with measured rates of association (kon) of 0.8-6 μM-1 s-1. We find only minor differences in these on rates when comparing PARP1 with the PARP1-HPF1 complex. By characterizing the rates of dissociation (koff) and the binding constants (KD) for two more recently discovered PARPi, we find, for example, that saruparib has a half-life for dissociation of 22.5 h and fluzoparib has higher affinity for PARP1 in the presence of HPF1, just like the structurally related compound olaparib. By using the measured KD and kon to calculate koff, we find that the potency of PARPi in cells correlates best with the koff from the PARP1-HPF1 complex. Our data suggest that dissociation of a drug compound from the PARP1-HPF1 complex should be the parameter of choice for guiding the development of next-generation PARPi.