CYTOCHROME-P450 2C9 IS RESPONSIBLE FOR HYDROXYLATION OF THE NAPHTHOQUINONE ANTIMALARIAL DRUG 58C80 IN HUMAN LIVER

(4-t-Butylcyclohexyl)-3-hydroxy-1,4-naphthoquinone (58C80) is an exper imental naphthoquinone antimalarial drug which undergoes extensive alk y hydroxylation in man. By means of purification, N-terminal amino aci d sequencing and inhibition by antibodies and sulfaphenazole, we have identified the form of cytochrome P450 primarily responsible for 58C80 hydroxylation in human liver, P450hB20-27, to be a member of the P450 2C9 subfamily. P450hB20-27 is a low-spin haemoprotein with molecular mass 54 kDa. 58C80 hydroxylation in human liver microsomes was depende nt on either NADPH or NADH, with the activity supported by NADH being 35% of that supported by NADPH. With purified P450hB20-27 cytochrome b 5 stimulated the NADH-dependent activity 8-fold but inhibited the NADP H-dependent activity by 30%. 58C80 is a novel substrate structure for human P450 2C and these results significantly broaden the range of dru gs which have been directly shown (i.e. using a purified enzyme as opp osed to expressed cDNA) to be metabolized by human P450 2C forms that are incontrovertibly expressed in human liver in vivo.