Preferential potentiation of topoisomerase I poison cytotoxicity by PARP inhibition in S phase.

BACKGROUND: Topoisomerase I (Topo I) poisons (e.g., camptothecin (CPT)), used to treat cancer, cause DNA breaks that are most cytotoxic during S phase. PARP-1 promotes DNA repair and PARP inhibitors (PARPi) sensitise cells to Topo I poisons. We aimed to determine whether chemosensitisation is also S phase specific using rucaparib, a potent PARPi in advanced clinical evaluation. METHODS: The impact of rucaparib, on CPT-induced cytotoxicity was measured in human colon cancer (LoVo) and leukaemic (K562) cells in asynchronous and cell cycle phase-separated cultures. Topoisomerase I and PARP levels and activity and the effect of rucaparib on DNA single-strand breaks (SSBs), double-strand breaks (DSBs) and collapsed replication fork induction and repair were determined in cell cycle phase-separated cells. RESULTS: The cytotoxicity of CPT was greatest during S phase, partially attributable to high Topo I activity, and rucaparib preferentially sensitised S-phase cells. Rucaparib increased CPT-induced DNA SSBs in all phases of the cell cycle, and increased DSB and γH2AX foci in S and G2, with γH2AX foci being highest in S-phase cells. Repair of SSBs and DSBs was most rapid during S then G2 phases and was substantially hindered by rucaparib. CONCLUSIONS: Rucaparib preferentially sensitises S-phase cells by increasing the frequency of collapsed replication forks.