Drug activation and DNA repair as determinants of cellular responses to mitomycin C.

摘要

Mitomycin C is a drug currently in use for the treatment of some solid tumors. As with any drug, cellular response will be determined by a number of factors. My thesis contains studies on two of these factors: metabolic drug activation and DNA excision repair. The two topics will be dealt with separately.; The studies on DNA excision repair have been aimed at the cloning of a human DNA repair gene. This gene is involved in repairing mitomycin C damage. Lack of gene function causes extreme mitomycin C sensitivity. The approach to cloning involved complementation of the repair defect in a Chinese hamster ovary cell mutant with human DNA. Eight transfectants were isolated, which, while they exhibited repair proficiency, did not exhibit the expected marker of human DNA uptake. This finding necessitated alternate strategies for gene cloning. The isolation and characterization of the transfectants suggest several strategies for gene cloning.; During the isolation of repair proficient transfectants, cell lines were isolated that exhibited mitomycin C resistance independent of repair capability. This resistance was accompanied by a decrease in drug induced lesions. Altered drug transport and detoxification were ruled out as mechanisms of resistance. Enzymes responsible for MMC activation were studied. It was found that the resistant cells had decreased levels of DT-diaphorase activity, as well as decreased DT-diaphorase mRNA transcription. The isolation of these MMC resistant cell lines should illuminate the pathways of bioreductive drug activation.