Human liver microsomal N-hydroxylation of dapsone by cytochrome P-4503A4.
One of the major routes of elimination of dapsone (4,4'-diaminodiphenylsulfone) is by N-oxidation, to produce a hydroxylamine metabolite. The specific form of cytochrome P-450 (P-450) involved in this oxidation reaction was examined in human liver microsomal preparations previously characterized with respect to their content of several known P-450 enzymes. Among five preparations, the rank order of activity for dapsone hydroxylamine formation was most well correlated with the immunochemically determined level of P-4503A4 (r = 0.94, p less than 0.03). Moreover, inhibition of microsomal oxidation was observed with antibodies specific to P-4503A, with a maximum reduction of greater than 90%, but was not produced by antibodies specific to P-4501A2, P-4502CMP, or P-4502E1. Prior incubation of microsomes with gestodene (100 microM) or troleandomycin (20 microM), known selective mechanism-based inhibitors of P-4503A enzymes (in the presence of NADPH), led to 75% and 40% reductions in catalytic activity, respectively. In contrast, preincubation with increasing concentrations of alpha-naphthoflavone, a known activator of P-4503A4, increased dapsone N-hydroxylation in a concentration-dependent manner, with 5-fold activation being observed at 50 microM alpha-naphthoflavone. Finally, P-4503A4 isolated from human liver microsomes and cDNA-expressed P-4503A4 (in yeast) were both able to catalyze dapsone N-hydroxylation, with the latter preparation exhibiting a 3-fold activation in the presence of 100 microM alpha-naphthoflavone. Collectively, these findings demonstrate that N-oxidation of dapsone in human liver is predominantly mediated by P-4503A4, and they suggest that quantitative measurement of this metabolic pathway in vivo might serve as an index of the activity of this enzyme.
Fleming
C M
CM
Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232.
Branch
R A
RA
Wilkinson
G R
GR
Guengerich
F P
FP
D000906:Antibodies; D001571:Benzoflavones / Q000494:pharmacology; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006801:Humans; D006900:Hydroxylation; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D008862:Microsomes, Liver / Q000201:enzymology*; D009652:Norpregnenes / Q000494:pharmacology; D014217:Troleandomycin / Q000494:pharmacology
D000906:Antibodies; D001571:Benzoflavones; D007527:Isoenzymes; D009652:Norpregnenes; C033273:Gestodene; C011512:alpha-naphthoflavone; D003577:Cytochrome P-450 Enzyme System; D014217:Troleandomycin