Formation of thioether conjugates of the bladder carcinogen ANFT catalyzed by prostaglandin H synthase

Certain 2-aminothiazole-substituted 5-nitrofuran carcinogens specifically and potently induce tumors of the urinary tract. Cooxidation by the hydroperoxidase activity of prostaglandin H synthase (PHS) has been implicated in the initiation of this process in vivo. The molecular mechanism of this process was examined utilizing 2-amino-4-(5-nitro-2-furyl)thiazole (ANFT) as the organic cosubstrate. The ability of ANFT to release electrons was determined by cyclic voltammetry, which established that ANFT is oxidized to yield an extremely reactive intermediate by an apparent one-electron mechanism. In conventional incubations containing solubilized microsomal PHS at pH 7.8, glutathione was found to trap approximately 50% of the ANFT present as a soluble metabolite. Comparative u.v. spectrophotometry, cyclic voltammetry, proton and carbon magnetic resonance spectroscopy, and chemical synthesis established that conjugate formation occurs by S-substitution of the thiazole ring to yield 2-amino-4-(5-nitro-2-furyl)-5-(glutathion-S-yl)thiazole. The 5-nitrofuran ring is not altered by this pathway. This suggests that PHS may oxidize the aminothiazole ring of ANFT by direct electron withdrawal to generate a reactive electrophilic intermediate which may react with critical cellular nucleophiles. This conjugate, or a further metabolite thereof, may be a biologic marker of PHS activity and may be useful in assessing the effects of potential chemoprotective agents on hydroperoxidase activity in vivo.