APOBEC3A (A3A) expression induces DNA replication stress without slowing down replication forks, leading to single-stranded DNA (ssDNA) gaps through PrimPol-mediated repriming. These ssDNA gaps are repaired by multiple pathways involving ATR, RAD51, and translesion synthesis (TLS). Inhibition of ATR or trapping of poly(ADP-ribose) polymerase (PARP) impairs the repair of A3A-induced gaps, preferentially killing A3A-expressing cells. Combining ATR and PARP inhibitors synergistically kills A3A-expressing cells, suggesting that therapies targeting gap-associated DNA repair pathways may be effective against A3A-expressing tumor cells. The study provides insights into the mechanisms of A3A-induced replication stress and identifies gap-targeted therapies as a potential strategy to eliminate A3A-expressing cancer cells.APOBEC3A (A3A) expression induces DNA replication stress without slowing down replication forks, leading to single-stranded DNA (ssDNA) gaps through PrimPol-mediated repriming. These ssDNA gaps are repaired by multiple pathways involving ATR, RAD51, and translesion synthesis (TLS). Inhibition of ATR or trapping of poly(ADP-ribose) polymerase (PARP) impairs the repair of A3A-induced gaps, preferentially killing A3A-expressing cells. Combining ATR and PARP inhibitors synergistically kills A3A-expressing cells, suggesting that therapies targeting gap-associated DNA repair pathways may be effective against A3A-expressing tumor cells. The study provides insights into the mechanisms of A3A-induced replication stress and identifies gap-targeted therapies as a potential strategy to eliminate A3A-expressing cancer cells.