Wortmannin is a potent inhibitor of DNA double strand break but not single strand break repair in Chinese hamster ovary cells.
Wortmannin, an inhibitor of p110 PI 3-kinase, also inhibits DNA-dependent protein kinase, which is known to mediate DNA double strand break repair. It was recently demonstrated that wortmannin sensitized cells to ionizing radiation (IR) (Price and Youmell, Cancer Res., 56, 246-250, 1996). Wortmannin was used to determine if the potentiation of IR-induced cytotoxicity in Chinese hamster ovary cells could be accounted for by an inhibition of DNA double strand break (DSB) repair. Wortmannin, at concentrations which were non-toxic per se (5 and 20 microM), increased IR cytotoxicity with dose enhancement factors at 10% survival of 2.7+/-0.28 (5 microM) and 5.3+/-0.86 (20 microM). The effects of wortmannin on DSB levels were assessed by neutral elution. The effects of wortmannin on the kinetics of DSB repair were evaluated over a 3 h time course. Wortmannin (50 microM) completely inhibited DSB repair over this period, without having any effect on DSB levels itself. The concentration-dependent effects of wortmannin on DSB levels showed that inhibition of DSB repair was significant at 1 microM, and near-maximal at 20 microM. In marked contrast, it exerted no effect on the kinetics of single strand break (SSB) repair as assessed by alkaline elution, even at concentrations as high as 50 microM. There was an excellent correlation between the concentration-dependence and exposure time of wortmannin required to enhance IR cytotoxicity and inhibit DSB repair. These data implicate inhibition of DNA-dependent protein kinase, and the consequent inhibition of DSB repair, as the mechanism whereby wortmannin potentiates the cytotoxicity of IR.
Boulton
S
S
Cancer Research Unit, Medical School, The University of Newcastle upon Tyne, UK.
Kyle
S
S
Yalçintepe
L
L
Durkacz
B W
BW
D000730:Androstadienes / Q000494:pharmacology*; D000818:Animals; D016466:CHO Cells / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D006224:Cricetinae; D004247:DNA / Q000378:metabolism / Q000528:radiation effects; D004249:DNA Damage / Q000187:drug effects*; D004260:DNA Repair / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D000077191:Wortmannin
D000730:Androstadienes; D004791:Enzyme Inhibitors; D004247:DNA; D000077191:Wortmannin