DNA polymerase beta mediates protection of mammalian cells against ganciclovir-induced cytotoxicity and DNA breakage.
The efficacy of suicide herpes simplex virus-1 thymidine kinase (HSVtk)/ganciclovir (GCV) gene therapy is often limited by intrinsic resistance of tumor cells. Here we show that repair of GCV incorporated in DNA is a factor involved in GCV resistance. A protective role of DNA repair in GCV-induced cell killing is supported by the following findings: (a) GCV-exposed Chinese hamster ovary-HSVtk cells exhibited both reduced repair of GCV and cloning efficiency in the presence of a specific polymerase beta (beta-pol) inhibitor, prunasin; (b) DNA beta-pol-deficient mouse fibroblasts were more sensitive to the cytotoxic, apoptosis-inducing, and genotoxic (DNA breakage and chromosomal aberration-inducing) effects of GCV as compared with wild-type and beta-pol-complemented cell lines; (c) methoxyamine, an inhibitor of beta-pol-dependent short-patch base excision repair, sensitized wild-type and complemented beta-pol cells to GCV, whereas it had no effect on the sensitivity of beta-pol-null cells to GCV. Because methoxyamine-mediated sensitization of beta-pol wild-type and beta-pol-complemented cells to GCV did not reach the level of null cells, we suggest that both beta-pol-dependent short- and long-patch base excision repair are involved in protection of cells to GCV. Some implications for HSVtk/GCV gene therapy are being discussed.
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Division of Applied Toxicology, Institute of Toxicology, University of Mainz, Germany.
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Kaina
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D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D019951:DNA Polymerase beta / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology; D015774:Ganciclovir / Q000378:metabolism / Q000633:toxicity*; D018259:Herpesvirus 1, Human / Q000201:enzymology / Q000235:genetics; D006898:Hydroxylamines / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D009570:Nitriles / Q000494:pharmacology; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D004791:Enzyme Inhibitors; D006898:Hydroxylamines; D009570:Nitriles; C019063:prunasin; D004247:DNA; C005214:methoxyamine; D013937:Thymidine Kinase; D019951:DNA Polymerase beta; D015774:Ganciclovir