Metabolic activation of arylamines, nitroaromatics, and related mutagens

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The focus of my research program is Molecular Toxicology. Our long-term goal is to understand the biochemical mechanisms that lead from exposure of a cell to a chemical to the ultimate toxic consequences of that exposure - e.g., induction of mutations, the raw material for evolutionary change. Most environmental mutagens are metabolized to DNA-reactive electrophiles. Consequently, I approach toxicology through biochemistry - especially, characterizing the enzymes that catalyze mutagen biotransformation. Fundamental questions about the metabolism and toxicology of aromatic amines, azo dyes, and nitroaromatic compounds (many of which are carcinogens) remain unanswered, but modern analytical and molecular-biological technologies are bringing the answers within reach.***Large knowledge gaps exist concerning the toxicology of azo dyes, their synthetic precursors, and the aromatic amines that are their putative reduction products. We recently examined cyanonitroanilines (CNAs), presumed to be formed by the reduction of azo Disperse Dyes. CNAs proved to be remarkably potent compounds in the Ames test, a widely used bacterial assay for mutagenic chemicals. This discovery - the starting point for the research proposed here - reveals an entirely unexpected effect: three distinct functional groups (cyano, amino, nitro) on a benzene ring interact to confer potent biological activity. Combining any two of these groups (e.g., 4-nitroaniline) yields at most weakly mutagenic compounds; but combining all three yields exceptionally potent mutagens. This is a fundamental challenge to our understanding of structure-activity relationships. To make sense of this effect, we need to elucidate the pathways of CNA bioactivation, and my lab is well positioned to do so. We have studied many of the enzymes that catalyze the biotransformation of N-aryl compounds, including nitroreductases (NRs, which reduce nitroaromatics to arylhydroxylamines and aromatic amines); azoreductases (ARs, which reduce azo compounds to the corresponding aromatic amines); and arylamine N-acetyltransferases (NATs, which transfer acetyl groups from acetyl CoA to aromatic amines and hydroxylamines). We will explore structure-activity relationships among CNAs and related compounds. We will study the enzymes that metabolize CNAs, including purified recombinant NRs, ARs, and NATs. Using LC-MS analysis, we will attempt to identify reactive metabolites and CNA-derived DNA adducts. Elucidating adduct structures will provide insight into mechanisms of activation.***These investigations will provide excellent training opportunities for graduate students, who will be well placed to obtain employment in areas such as regulatory toxicology, analytical and environmental chemistry, and the pharmaceutical industry.