Rapid detection of DNA-damaging agents using repair-deficient CHO cells.
A screening method is introduced to detect and classify DNA-damaging agents using DNA repair-deficient strains of Chinese hamster ovary cells. Differential cytotoxicity (relative growth) of the mutant cells compared to the wild-type cells was interpreted as a measure of lethal, potentially repairable damage to DNA. The assay consists of exposing the wild-type cells and three mutant strains to the test compound in a 24-well tray and using staining intensity to estimate growth after 72 h. The battery of mutants consists of two UV-sensitive strains (UV4 and UV5) that are deficient in different aspects of nucleotide excision repair, and strain EM9, which is defective in DNA-strand-break rejoining. The assay was highly reproducible, and the magnitude of the differential cytotoxicity response compared favorably with the amount of differential killing measured by colony-formation survival curves for several chemicals. 15 direct-acting and 7 metabolism-dependent agents that were expected to produce bulky, covalent DNA adducts were tested in the assay, and all produced a differential cytotoxicity response in at least two of the mutants. UV4 and UV5 showed a response to all of the test compounds whereas EM9 showed a response to 7 of the test compounds. Thus, the pattern of mutant responses presumably reflects the types of DNA damage produced by a compound. Although this aspect is still under development, these results indicate the potential of a larger battery of mutants to classify a wide spectrum of chemicals according to the lesions they produce. 13 non-DNA damaging agents were also tested and none produced a differential cytotoxicity response, suggesting that this endpoint is specific for DNA damage. We conclude that this assay may be a cost-effective alternative or adjunct to the existing short-term tests.
D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D055598:Chemical Phenomena; D002621:Chemistry; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA; D004260:DNA Repair; D005260:Female; D009152:Mutagenicity Tests / Q000379:methods*; D010053:Ovary
D004247:DNA