The goal of this research is to understand in greater detail the role of DNA damage and repair mechanisms in determining the cytotoxic response of mammalian cells to antitumor drugs. The rationale is based on previous reports showing that these processes can modulate the cytotoxic effects of such drugs by large factors. Because these mechanisms are enzymatic, their influence on drug effects can be evaluated either through the use of genetically defined mutant cell lines deficient in specific enzymes or of chemical inhibitors of these enzymatic processes. Furthermore, such inhibitors allow the extension of this basic approach to the in vivo situation in order to verify that the same mechanisms observed in vitro are operative in vivo. Various antitumor agents will be used in the investigations; however, DNA is believed to be the primary critical cellular target for the cytotoxicty of each of these agents. These would include: the bifunctional alkylating agents e.g., nitrogen mustard, melphalan, and 1,3-bis(2-chloroethyl)-1-nitrosourea; cis-diamminedichloroplatinum; and mitomycin-C. The proposed work is divided into 3 specific aims. The first proposes to isolate and characterize antitumor drug-sensitive mutant cell lines of Chinese hamster cells so that the enzymatic steps involved in drug sensitivity can be determined. The nature of the biochemical defect will be delineated using a variety of techniques in order to establish how it relates to modulation of cytotoxicity.
Aim 2 will be to evaluate the drug modification and repair pathways identified in aim 1 by the use of metabolic inhibitors in vitro. The inhibitors to be used include inhibitors of pathways involved in drug detoxification and inhibitors of DNA repair pathways. Finally, in aim 3, based on the results in aim 2, these inhibitors will be used in vivo to verify that the same biochemical processes are operative in normal and neoplastic mouse tissues. Moreover, modifications by the inhibitors that lead to an enhanced antitumor effect can be tested by correlating the changes in DNA damage or repair with effects on tumor growth delay. The experiments in aim 3 will take advantage of an adaptation of the technique of alkaline elution that allows sensitive characterization of the lesions induced in the cellular DNA of various tissues of mice treated with antitumor drugs in vivo. The results of this investigation, hopefully, will contribute to our further understanding of the basic biochemical processes in cells which affect cytotoxicity through modification of drug-induced DNA damage or its repair. Such knowledge could, in the future, be used to design more effective clinical regimens for the treatment of human cancer with antitumor drugs.
