REV1-Polζ maintains the viability of homologous recombination-deficient cancer cells through mutagenic repair of PRIMPOL-dependent ssDNA gaps

BRCA1/2 mutant tumor cells display an elevated mutation burden, the etiology of which remains unclear. Here, we report that these cells accumulate ssDNA gaps and spontaneous mutations during unperturbed DNA replication due to repriming by the DNA primase-polymerase PRIMPOL. Gap accumulation requires the DNA glycosylase SMUG1 and is exacerbated by depletion of the translesion synthesis (TLS) factor RAD18 or inhibition of the error-prone TLS polymerase complex REV1-Polζ by the small molecule JH-RE-06. JH-RE-06 treatment of BRCA1/2-deficient cells results in reduced mutation rates and PRIMPOL- and SMUG1-dependent loss of viability. Through cellular and animal studies, we demonstrate that JH-RE-06 is preferentially toxic toward HR-deficient cancer cells. Furthermore, JH-RE-06 remains effective toward PARP inhibitor (PARPi)-resistant BRCA1 mutant cells and displays additive toxicity with crosslinking agents or PARPi. Collectively, these studies identify a protective and mutagenic role for REV1-Polζ in BRCA1/2 mutant cells and provide the rationale for using REV1-Polζ inhibitors to treat BRCA1/2 mutant tumors. [Display omitted] •ssDNA gaps arise in BRCA1 mutant cancer cells due to PRIMPOL-mediated repriming•BRCA1/2 deficiency leads to mutagenic ssDNA gap repair by REV1-Polζ-dependent TLS•Targeted REV1-Polζ inhibition shows enhanced toxicity in HR-deficient cancer cells•ssDNA gaps formed by SMUG1 and PRIMPOL mediate the toxicity of REV1-Polζ inhibition Taglialatela et al. report that homologous recombination (HR)-deficient cancer cells, such as BRCA1/2 mutant cells, display increased reliance on error-prone translesion synthesis (TLS) for the repair of ssDNA gaps arising spontaneously during DNA replication. TLS inhibition shows exquisite toxicity in BRCA1/2-deficient cancer cells, providing the basis for alternative therapies against BRCA1/2 mutant tumors.