Isolation and partial characterisation of a mammalian cell mutant hypersensitive to topoisomerase II inhibitors and X-rays.
We have isolated, following one-step mutagenesis, a Chinese hamster ovary cell mutant hypersensitive to the intercalating agent, adriamycin (4-fold compared to parental CHO-K1 cells). This agent exerts at least part of its cytotoxic action via inhibition of the nuclear enzyme, topoisomerase II. The mutant, designated ADR-3, showed hypersensitivity to all classes of topoisomerase II inhibitors, including actinomycin D, amsacrine (m-AMSA), etoposide (VP16) and mitoxantrone. ADR-3 cells also showed cross-sensitivity to ionizing radiation, but not to UV light. Cellular accumulation of radiolabeled actinomycin D was similar in parental and mutant cells. At equimolar doses, adriamycin induced more protein-associated DNA single- and double-strand breaks in ADR-3 cells than in CHO-K1 cells. Topoisomerase II activity was elevated to a small but significant degree in ADR-3 cells, and this was reflected in a 1.5-fold higher level of topoisomerase II protein in ADR-3 than in CHO-K1 cells, as judged by Western blotting. ADR-3 cells were hypersensitive to cumene hydroperoxide but cross-resistant to hydrogen peroxide, suggesting possible abnormality in the detoxification of peroxides by glutathione peroxidase or catalase. Glutathione peroxidase activity against hydrogen peroxide was similar in CHO-K1 and ADR-3 cell extracts, but activity against cumene hydroperoxide was evaluated to a small but significant extent in mutant cells. Catalase levels were not significantly different in ADR-3 and CHO-K1 cells. ADR-3 cells were recessive in hybrids with parental CHO-K1 cells with respect to sensitivity to topoisomerase II inhibitors and X-rays, and represent a different genetic complementation group from the previously reported adriamycin-sensitive mutant, ADR-1 [Davies et al., J. Biol. Chem., 263 (1988) 17724-17729].
Davies
S M
SM
Department of Clinical Oncology, Medical School, University of Newcastle upon Tyne, Great Britain.
Davies
S L
SL
Hall
A G
AG
Hickson
I D
ID
D000818:Animals; D015153:Blotting, Western; D002374:Catalase / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival; D006224:Cricetinae; D003412:Cricetulus; D004249:DNA Damage; D004317:Doxorubicin / Q000494:pharmacology; D004342:Drug Hypersensitivity / Q000235:genetics*; D005979:Glutathione Peroxidase / Q000378:metabolism; D006822:Hybrid Cells; D006861:Hydrogen Peroxide / Q000494:pharmacology; D009154:Mutation; D011836:Radiation Tolerance; D059005:Topoisomerase II Inhibitors
D059005:Topoisomerase II Inhibitors; D004317:Doxorubicin; D006861:Hydrogen Peroxide; D002374:Catalase; D005979:Glutathione Peroxidase