The long range objective is to evaluate the use of human lymphocytes as potential markers of human exposure and/or susceptibility. The effect of activation/ deactivation pathways on the formation of DNA adducts and the resulting consequences on genetic endpoints of DNA damage (assessed by various methods including gene mutation) are evaluated in human lymphocytes following in vitro exposure to chemicals. We have shown that human lymphocytes are polymorphic in one isozyme of glutathione S-transferase (GST) referred to as the mu form (GST-mu). Approximately 50% of the population has high GST-mu activity and 50% has low activity. A significant correlation is seen for GST-mu activity between human liver and lymphocytes suggesting that human lymphocytes are an appropriate surrogate tissue for liver GST-mu. GSTs play key roles in the metabolic detoxification of PAHs whereas this enzyme activates ethylene dibromide (EDB) to a DNA reactive species. A significant correlation between DNA adducts of benzo(a)pyrene (BP) and BP metabolism was seen in human lymphocytes for persons having high GST-mu activity. In cell-free experiments, the importance of GST-mu on adduct formation by BP and aflatoxin B1 (AFB1) has been evaluated. Our results indicate that much higher concentrations of AFB1-derived adducts are formed in systems containing low GST mu activity compared to systems containing high GST-mu. These data suggest an important role for GST-mu in the detoxication of AFB1. A 10-fold variation in DNA damage, measured by nucleoid sedimentation, is seen in human lymphocyte incubated in vitro with EDB. Moreover, there is a strong correlation between DNA damage and the level of EDB-DNA binding. We have verified by HPLC that the primary adduct produced by lymphocytes is the S-[2-N7-guanyl)-ethyl] glutathione and it is responsible for the DNA damage observed. Species comparisons of lymphocytes responses to EDB shows that the order of sensitivity is humans > rats > mice and this species comparison is correlated with GST activity. Current studies are evaluating the relationship between polymorphisms in drug-metabolizing enzymes, DNA adducts and mutation spectra of the hprt gene.
