USE OF IMMUNOLOGICAL TECHNIQUES TO STUDY THE INTERACTION OF CARCINOGENS WITH DNA

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Antibodies specific for carcinogen-DNA adducts have probed the nature, extent, and consequences of in vitro and in vivo DNA modification. DNAs substituted with 2-acetylaminofluorene (AAF) and cis-diamiminedichloroplatinum II (cis-DDP) were analyzed by quantitative immunoassays able to detect about 100 adducts per cell and by immunohistochemical procedures developed to localize adducts in situ. In hepatic DNA of rats fed a carcinogenic dose of AAF for 4 weeks, adduct accumulation reached a plateau at 2-3 weeks. Since adduct removal was biphasic during 4 subsequent weeks on control diet, a computer-derived pharmacokinetic model proposed that adducts are formed into two genomic compartments, one from which they are removed rapidly and another from which they are removed slowly. Studies initiated to identify these two compartments involve adduct analysis in different liver cell types and different chromatin regions including the nuclear matrix. Immunohistochemical localization of AF-DNA adducts in livers of rats fed AAF was shown by fluorescence and microfluorometry to be concentrated primarily in periportal regions. In addition, there were no adducts detectable by immunofluorescence in preneoplastic foci induced by several different protocols. Nucleated peripheral blood cell DNA was obtained from cancer patients at multiple times during courses of cis-DDP therapy, and a total of 254 samples were analyzed. Of these, 23 untreated control samples were negative, and 43% of the 231 samples from patients receiving cis-DDP were positive. Adduct accumulation, in positive patients, occurred as a function of total cumulative dose, suggesting relatively slow adduct removal. Disease response data on 55 patients indicated that individuals with high adduct levels have a high rate of complete response to therapy. Parallel experiments in animal models have demonstrated that the same adduct forms in kidney, gonads, and tumors of rats and mice in direct relation to dose.