This research program focuses on the mechanisms by which cells respond to DNA damaging agents such as ionizing radiation, chemical carcinogens and anticancer drugs. The program is divided into three areas. The first is an analysis of the biochemical mechanisms by which chemical and physical agents induce the mutations that presumably initiate cells along the pathway toward malignancy. We examine the spectrum of mutations induced by a DNA damaging agents (for in the damaged DNA (adducts) may give rise to specific mutations. Using a combination of chemical synthesis and recombinant DNA tools, viral genomes are constructed containing the adducts suspected to have caused the mutations. Following replication of the site specifically modified genomes in bacterial or mammalian cells we determine the type, amount and genetic requirements for mutagenesis by each lesion studied. This work priorities the mutagenic potential of individual DNA adducts. The specific DNA adducts we proposed to study include those produced by oxidants and ionizing radiation, simple alkylating agents, aflatoxin, B1, cis- diamminedichloroplatinum (II) (cisplatin), 4-aminobiphenyl, 2-amino-3,8- dimethylimidazo94,5-f0quinoxaline (MeIQx), and vinyl chloride. The second area of proposed research is an examination of the mechanism of toxicity by the anticancer drug cisplatin. We propose to continue our investigation of a class of proteins we have termed """"""""DRPs"""""""" (for Damage Recognition Proteins). We hypothesize that DRPs are involved in the anticancer mechanism of cisplatin by either or both of the following models. The first model proposed that DRPs bind to therapeutically effective adducts of cisplatin and shield those adducts from DNA repair. The second model is based upon recent discovery that some of the DRPs have essential natural functions (one is the transcription factor, hUBF). We shall test the hypothesis that cisplating adducts divert DRPs from their natural functions, hence disrupting cellular homeostasis. Our third proposed area of investigation is the design of a novel anticancer agent that works by the """"""""shielding"""""""" mechanism proposed above for cisplatin. In this case, however, a DNA binding domain will be linked to a protein will bind to the adduct and shield it from repair, preserving the adduct so that its maximal lethal impact can be realized. In normal (nontumor) cells no such protection will be afforded owing to the absence of the tumor specific protein. In normal cells, therefore, the toxic effect of the adduct will be reduced by the repair system of the host.
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