Guiding ATR and PARP inhibitor combinations with chemogenomic screens

Combinations of ataxia telangiectasia- and Rad3-related kinase inhibitors (ATRis) and poly(ADP-ribose) polymerase inhibitors (PARPis) synergistically kill tumor cells through modulation of complementary DNA repair pathways, but their tolerability is limited by hematological toxicities. To address this, we performed a genome-wide CRISPR-Cas9 screen to identify genetic alterations that hypersensitize cells to a combination of the ATRi RP-3500 with PARPi, including deficiency in RNase H2, RAD51 paralog mutations, or the “alternative lengthening of telomeres” telomere maintenance mechanism. We show that RP-3500 and PARPi combinations kill cells carrying these genetic alterations at doses sub-therapeutic as single agents. We also demonstrate the mechanism of combination hypersensitivity in RNase H2-deficient cells, where we observe an irreversible replication catastrophe, allowing us to design a highly efficacious and tolerable in vivo dosing schedule. We present a comprehensive dataset to inform development of ATRi and PARPi combinations and an experimental framework applicable to other drug combination strategies. [Display omitted] •A CRISPR screen identifies cancer alterations that sensitize to ATRis/PARPis•Low doses of ATRis/PARPis are effective in tumor models carrying these alterations•Mechanistic insights allow designing a well-tolerated intermittent dosing schedule•Together, these data can guide the use of ATRi/PARPi combinations in the clinic Combinations of PARP inhibitor (PARPi) and ATR inhibitor (ATRi) anti-cancer drugs show promise for cancer therapy but are limited by bone marrow toxicities. Zimmermann et al. identify multiple genetic alterations in tumors that sensitize to ATRi/PARPi combinations, potentially allowing use of lowered, tolerated doses to kill tumor cells.