Abstract

Dapsone is used in the treatment of Pneumocystis carinii pneumonia, an opportunistic infection that afflicts acquired immunodeficiency syndrome (AIDS) patients. Inhibition of N-acetyltransferase (NAT)-dependent acetylation of dapsone could increase peak plasma concentrations of dapsone and shift the biotransformation pathway to the P450-mediated formation of a toxic metabolite of dapsone, the hydroxylamine. Therefore, we have determined using human liver cytosol and bacterially expressed NATs, the NAT isoform responsible for acetylating dapsone and the potential for antiopportunistic infection drugs to inhibit this metabolic pathway. Formation of monoacetyldiaminodiphenylsulfone (MADDS) was quantitated by HPLC/UV detection at 270 nm after incubation of dapsone with 100 microM acetyl coenzyme A regenerating system and human liver cytosol. The mean +/- SD apparent KM for the formation of MADDS in three different human livers predicted to be fast acetylators based on genotyping was 98 +/- 17.6 microM, and the Vmax was 190 +/- 20 pmol/min/mg cytosol protein. Eadie-Hofstee transformation of the substrate velocity data was linear, indicating acetylation by a kinetically single enzyme. Sulfamethazine (250 microM) inhibited dapsone acetylation by 100% and 80%, respectively, at dapsone concentrations of 3 and 100 microM, in both fast- and slow-acetylating liver cytosol preparations, whereas para-amino-benzoic acid (100 microM) did not inhibit MADDS formation at either of these dapsone concentrations. Lineweaver-Burk plots of dapsone acetylation in the presence of 0, 25, and 50 microM sulfamethazine showed an increase in the apparent KM, with increase in sulfamethazine concentration with no change in the Vmax, indicating competitive inhibition of dapsone acetylation by sulfamethazine. The apparent KM of dapsone acetylation by bacterially expressed NAT1 and NAT2 enzymes was 687 and 136 microM, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)