Inhibition of ATR kinase with the selective inhibitor VE-821 results in radiosensitization of cells of promyelocytic leukaemia (HL-60)

Abstract

We compared the effects of inhibitors of kinases ATM (KU55933) and ATR (VE-821) (incubated for 30min before irradiation) on the radiosensitization of human promyelocyte leukaemia cells (HL-60), lacking functional protein p53. VE-821 reduces phosphorylation of check-point kinase 1 at serine 345, and KU55933 reduces phosphorylation of check-point kinase 2 on threonine 68 as assayed 4h after irradiation by the dose of 6Gy. Within 24h after gamma-irradiation with a dose of 3Gy, the cells accumulated in the G2 phase (67%) and the number of cells in S phase decreased. KU55933 (10μM) did not affect the accumulation of cells in G2 phase and did not affect the decrease in the number of cells in S phase after irradiation. VE-821 (2 and 10μM) reduced the number of irradiated cells in the G2 phase to the level of non-irradiated cells and increased the number of irradiated cells in S phase, compared to irradiated cells not treated with inhibitors. In the 144h interval after irradiation with 3Gy, there was a considerable induction of apoptosis in the VE-821 group (10μM). The repair of the radiation damage, as observed 72h after irradiation, was more rapid in the group exposed solely to irradiation and in the group treated with KU55933 (80 and 77% of cells, respectively, were free of DSBs), whereas in the group incubated with 10μM VE-821, there were only 61% of cells free of DSBs. The inhibition of kinase ATR with its specific inhibitor VE-821 resulted in a more pronounced radiosensitizing effect in HL-60 cells as compared to the inhibition of kinase ATM with the inhibitor KU55933. In contrast to KU55933, the VE-821 treatment prevented HL-60 cells from undergoing G2 cell cycle arrest. Taken together, we conclude that the ATR kinase inhibition offers a new possibility of radiosensitization of tumour cells lacking functional protein p53.