Interspecies comparison and role of human cytochrome P450 and flavin-containing monooxygenase in hepatic metabolism of L-775,606, a potent 5-HT_1D receptor agonist

Abstract

1. Quantitative species differences and human liver enzymes involved in the metabolism of L-775,606, a potent and selective 5-HT_1D receptor agonist developed for the acute treatment of migraine headache, have been investigated in vitro. 2. In human, monkey, dog and rat liver microsomes, formation of the hydroxylated M1 and the N-dealkylated M2 was mediated by enzyme(s) of high-affinity (apparent K_m ~1-6 muM), and that of the two N-oxide isomers (M3) was catalysed by those of low affinity (apparent K_m ~50-110 muM). In dog, M3 constituted a major pathway (~40%), whereas in all other species it was a minor metabolite (<5%). 3. In human liver microsomes, a marked inhibition ( 80%) of M1 and M2 formation was observed by SKF525-A, troleandomycin, ketoconazole and anti-CYP3A antibodies, whereas the inhibition was modest (~20-40%) with quercetin. Of seven cDNA-expressed human P450 tested, only CYP3A4 and CYP2C8 were capable of oxidizing L-775,606, resulting primarily in M1 and M2. However, CYP3A4 possessed much higher affinity ( 20-fold) and much higher intrinsic activity (>100-fold) than CYP2C8. 4. In contrast, N-oxidation was not inhibited by any inhibitors of P450 tested, but rather was reduced significantly by heat treatment and methimazole, and was increased substantially with an incubation pH > 7.4. Human flavin-containing monooxygenase form 3 (FMO3) catalysed exclusively the N-oxidation to M3, with apparent K_m and optimum pH comparable with those observed in human liver microsomes. 5. These results demonstrated quantitative interspecies differences in the metabolism of L-775,606. In human, metabolism of L-775,606 to the principal metabolites, M1 and M2, was mediated primarily by CYP3A4 with minimal contribution from CYP2C8, whereas the minor N-oxidative pathway was catalysed mainly by FMO3.