A number of known chemopreventive agents are hypothesized to work in part via induction of glutathione-S-transferase (GST) expression. In order to understand the functions of GST in detoxification of cytotoxic and mutagenic electrophiles, the investigators have utilized transgenic cell lines to show that GST expression can provide efficacious protection against DNA alkylation and, in some cases, cytotoxicity caused by electrophilic carcinogens that are GST substrates, including 4-NQO, B[a]P, BPDE, AFB1, CDNB, and specific drugs. However, the results indicated that the factors governing protection by GST are complex and vary with different agents and endpoints. The studies outlined in this proposal will provide new information on the efficacy and specificity of human GSTP1 or GSTM1 protection against DNA adduct formation or cytotoxicity caused by exposure to PAHs activated in situ by co-expressed rat rCYP1A1 or human hCYP1A1. Heterologous expression of the GST isozymes in V79 cells previously stably transfected with rCYP1A1 or hCYP1A1 will be used as the experimental model system. Importantly, this information will be directly compared with the effects of GST isoenzyme expression on metabolite accumulation and with cellular end-points, such as cytotoxicity, in cells. They hypothesize that the efficacy of the GST system is dependent on multiple factors and not only the enzymatic efficiency with a particular substrate. Several of these factors will be examined in the next funding period, including the relationship between protection by transfected GST isozymes against the above end-points and 1) the level of GST protein expressed, 2) rates and site of activation vs. detoxification, and resultant metabolite profiles and/or levels, 3) cellular factors: glutathione (GSH) supply, and/or efflux of GSH-conjugates, and 4) genetic polymorphisms that affect the active site architecture of hGSTP1-1. These studies will provide a detailed understanding of key parameters affecting the efficacy of GST protection in the transfected cells, and should help to identify the mechanisms of differential protection observed against the various cellular injury end-points examined.
