Uracil processing by SMUG1 in the absence of UNG triggers homologous recombination and selectively kills BRCA1/2-deficient tumors

Resistance to poly (ADP-ribose) polymerase (PARP) inhibitors (PARPis) is the major obstacle to their effectiveness in the treatment of homologous recombination (HR)-deficient (HRD) tumors. Hence, developing alternative treatments for HRD tumors is critical. Here, we show that targeting the uracil excision pathway kills HRD tumors, including those with PARPi resistance. We found that the interplay between the two major uracil DNA glycosylases UNG and SMUG1 is regulated by nuclear nicotinamide adenine dinucleotide (NAD⁺), which maintains UNG at replication forks (RFs) and restrains SMUG1 chromatin binding. In the absence of UNG, SMUG1 retention on chromatin leads to persistent abasic sites, which incision by APE1 results in PARP1 hyperactivation, stalled RFs, and RAD51 foci. In HRD cells (i.e., BRCA1/2-deficient), this leads to under-replicated DNA that, when propagated throughout mitosis, results in chromosome fragmentation and cell death. Our findings open up unique possibilities for targeted therapies for HRD tumors based on UNG inhibition and uracil accumulation in the genome.