Enhanced Radiation Response in Human Carcinoma Cell Lines by the Cardiovascular Drug Hydralazine

Abstract

Purpose/Objective(s)Hydralazine has vascular dilating effects, and has been used for 30 years as an oral antihypertensive. One rare side effect of this drug is a lupus-like syndrome. It is likely that this side effect is caused by the demethylating properties of this drug. As recent studies have shown that demethylating agents can lead to an enhanced radio response we examined the effect of hydralazine on response to radiation (IR) in a variety of human cancer cell lines.Materials/MethodsThree human cancer cell lines (HCT116, DU145, and MiaPaca) were treated for 72 hour with hydralazine (10, 30, and 50μmol/L, respectively) pre-irradiation in all in vitro experiments. The effects of hydralazine on the in vitro radiosensitivity were evaluated using clonogenic assay. DNA damage and repair were evaluated using γH2AX at 1, 6, and 24 hours after 2 Gy IR. Neutral comet assay was performed at 0, 3, 6, 16, 24 hours after 10 Gy IR to further evaluate DNA damage and repair. Mechanism of cell death after DNA damage was determined by immunostaining for mitotic catastrophe and flow cytometry for apoptosis. Drug induced cell cycle changes were evaluated by staining of phospho-H3 and flow cytometry.ResultsExposure of HCT116, DU145, and MiaPaca to hydralazine for 72 hour prior to radiation resulted in an increase in radiosensitivity with dose enhancement factors at a surviving fraction of 0.1 of 1.36, 1.83, and 1.35, respectively. In HCT116, hydralazine had no effect on the number of cells in each phase of the cell cycle nor on the activation of the G2 cell cycle checkpoint after IR. However, hydralazine did modify the time course of damaged DNA repairing after radiation. The number of γH2AX foci per cell was significantly greater at 6 and 24 hours after hydralazine and RT as compared to either individual treatment. Mitotic catastrophe, measured at 72 hours, was also significantly increased in cells receiving the hydralazine + IR combination as compared to the single treatments.ConclusionsThese results indicate that hydralazine can enhance tumor cell radiosensitivity in vitro and suggests that this effect involves an inhibition of DNA double strand breaks repair leading to an increase in mitotic catastrophe. Purpose/Objective(s)Hydralazine has vascular dilating effects, and has been used for 30 years as an oral antihypertensive. One rare side effect of this drug is a lupus-like syndrome. It is likely that this side effect is caused by the demethylating properties of this drug. As recent studies have shown that demethylating agents can lead to an enhanced radio response we examined the effect of hydralazine on response to radiation (IR) in a variety of human cancer cell lines. Hydralazine has vascular dilating effects, and has been used for 30 years as an oral antihypertensive. One rare side effect of this drug is a lupus-like syndrome. It is likely that this side effect is caused by the demethylating properties of this drug. As recent studies have shown that demethylating agents can lead to an enhanced radio response we examined the effect of hydralazine on response to radiation (IR) in a variety of human cancer cell lines. Materials/MethodsThree human cancer cell lines (HCT116, DU145, and MiaPaca) were treated for 72 hour with hydralazine (10, 30, and 50μmol/L, respectively) pre-irradiation in all in vitro experiments. The effects of hydralazine on the in vitro radiosensitivity were evaluated using clonogenic assay. DNA damage and repair were evaluated using γH2AX at 1, 6, and 24 hours after 2 Gy IR. Neutral comet assay was performed at 0, 3, 6, 16, 24 hours after 10 Gy IR to further evaluate DNA damage and repair. Mechanism of cell death after DNA damage was determined by immunostaining for mitotic catastrophe and flow cytometry for apoptosis. Drug induced cell cycle changes were evaluated by staining of phospho-H3 and flow cytometry. Three human cancer cell lines (HCT116, DU145, and MiaPaca) were treated for 72 hour with hydralazine (10, 30, and 50μmol/L, respectively) pre-irradiation in all in vitro experiments. The effects of hydralazine on the in vitro radiosensitivity were evaluated using clonogenic assay. DNA damage and repair were evaluated using γH2AX at 1, 6, and 24 hours after 2 Gy IR. Neutral comet assay was performed at 0, 3, 6, 16, 24 hours after 10 Gy IR to further evaluate DNA damage and repair. Mechanism of cell death after DNA damage was determined by immunostaining for mitotic catastrophe and flow cytometry for apoptosis. Drug induced cell cycle changes were evaluated by staining of phospho-H3 and flow cytometry. ResultsExposure of HCT116, DU145, and MiaPaca to hydralazine for 72 hour prior to radiation resulted in an increase in radiosensitivity with dose enhancement factors at a surviving fraction of 0.1 of 1.36, 1.83, and 1.35, respectively. In HCT116, hydralazine had no effect on the number of cells in each phase of the cell cycle nor on the activation of the G2 cell cycle checkpoint after IR. However, hydralazine did modify the time course of damaged DNA repairing after radiation. The number of γH2AX foci per cell was significantly greater at 6 and 24 hours after hydralazine and RT as compared to either individual treatment. Mitotic catastrophe, measured at 72 hours, was also significantly increased in cells receiving the hydralazine + IR combination as compared to the single treatments. Exposure of HCT116, DU145, and MiaPaca to hydralazine for 72 hour prior to radiation resulted in an increase in radiosensitivity with dose enhancement factors at a surviving fraction of 0.1 of 1.36, 1.83, and 1.35, respectively. In HCT116, hydralazine had no effect on the number of cells in each phase of the cell cycle nor on the activation of the G2 cell cycle checkpoint after IR. However, hydralazine did modify the time course of damaged DNA repairing after radiation. The number of γH2AX foci per cell was significantly greater at 6 and 24 hours after hydralazine and RT as compared to either individual treatment. Mitotic catastrophe, measured at 72 hours, was also significantly increased in cells receiving the hydralazine + IR combination as compared to the single treatments. ConclusionsThese results indicate that hydralazine can enhance tumor cell radiosensitivity in vitro and suggests that this effect involves an inhibition of DNA double strand breaks repair leading to an increase in mitotic catastrophe. These results indicate that hydralazine can enhance tumor cell radiosensitivity in vitro and suggests that this effect involves an inhibition of DNA double strand breaks repair leading to an increase in mitotic catastrophe.