Cancer is the second leading cause of death in the United States and involves multiple stages including, initiation, promotion, progression, and metastasis. This Program will dissect the discrete molecular events associated with early components of this multistage disease by using complete carcinogens as chemical tweezers [polycyclic aromatic hydrocarbons (PAH)]. The hypothesis to be tested is that reactive and redox active PAH o-quinones generated by constitutively expressed aldo-keto reductase (AKR?s)contribute to PAHmultistage carcinogenesis. We propose that the electrophilic PAH-o-quinones plus the reactive oxygen species (ROS) they generate lead to covalent modifications of DNA in PAH target tissues and this may have mutational consequences leading to initiation. We also propose that the PAH o-quinones and ROS have direct effects on protein kinase C and its downstream signals. These epigenetic effects may have consequences for cell proliferation leading to promotion. These events could explain how PAH act as complete carcinogens. The Program consists of three interactive Projects and two Cores. Each project will use common human cell lines which are either capable of PAH activation (MCF-7 cells) or are PAH targets (bronchoalveolar cells). Project # 1 (Dr. Penning) will compare the roles of human aldehyde reductase (AKR1A1), CYP1A1/CYP1B1, and CYP-peroxidase in the metabolic activation of PAH using a stable expression strategy. Project #2 (Dr. Blair) will develop quantitative LC/MS methods to measure covalent modifications to DNA by reactive PAH-metabolites (o-quinones, anti-diol epoxides and radical cations), ROS and decomposition products of lipid hydroperoxides. The relative abundance of these DNA-adducts will be measured in bronchoalveolar cells (Project #1) and in sites of PAH carcinogenesis i.e, SENCAR mouse skin (Project #2). This will identify the metabolic pathway most responsible for adduct formation. Project #3 (Drs. Kazanietz and Assoian) will determine whether PAH o-quinones and ROS activate or inhibit individual PKC isoforms in vitro and in vivo and measure the phenotypic outcome on PMA-induced cell proliferation or growth inhibition. Changes in growth properties will be correlated to effects on cell cycle signaling pathways. The Bioanalytical Core (Core B) will provide LC/MS support to monitor the purity of PAH metabolites used in all projects, and will provide authentic PAH-DNA adduct reference compounds for Projects 1 and 2. The administrative core (Core A) will provide oversight to the program, a share Program Database, and biostatistical support to tract trends within and between projects. By studying the discrete molecular events responsible for the causation of cancer this Program may lead to the early prevention and intervention of this disease.
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