Quinuclidine N-Oxygenation Mediated by Flavin-Containing Monooxygenases 1 and 3 in Kidney and Liver Microsomes from Humans, Monkeys, Dogs, and Pigs.

Abstract

Flavin-containing monooxygenases (FMOs) are a family of enzymes that are involved in the oxygenation of heteroatom-containing molecules. In humans, FMO3 is the major hepatic form, whereas FMO1 is predominant in the kidneys. FMO1 and FMO3 have also been identified in monkeys, dogs, and pigs. The predicted contribution of human FMO3 to drug candidate N-oxygenation could be estimated using the classic base dissociation constants of the N-containing moiety. A basic quinuclidine moiety was found in natural quinine and medicinal products. Consequently, N-oxygenation of quinuclidine was evaluated using liver and kidney microsomes from humans, monkeys, dogs, and pigs as well as recombinant FMO1, FMO3, and FMO5 enzymes. Experiments using simple reversed-phase liquid chromatography with fluorescence monitoring revealed that recombinant FMO1 mediated quinuclidine N-oxygenation with a high capacity in humans. Moreover, recombinant FMO1, FMO3, and/or FMO5 in monkeys, dogs, and pigs exhibited relatively broad substrate specificity toward quinuclidine N-oxygenation. Kinetic analysis showed that human FMO1 efficiently, and pig FMO1 moderately, mediated quinuclidine N-oxygenation with high capacity, which is consistent with the reported findings for larger substrates readily accepted by pig FMO1 but excluded by human FMO1. In contrast, human FMO3-mediated quinuclidine N-oxygenation was slower than that of the typical FMO3 substrate trimethylamine. These results suggest that some species differences exist in terms of FMO-mediated quinuclidine N-oxygenation in humans and some animal models (monkeys, dogs, and minipigs); however, the potential for quinuclidine, which has a simple chemical structure, to be inhibited clinically by co-administered drugs should be relatively low, especially in human livers. SIGNIFICANCE STATEMENT: The high capacity of human flavin-containing monooxygenase (FMO) 1 to mediate quinuclidine N-oxygenation, a basic moiety in natural products and medicines, was demonstrated by simple reversed-phase liquid chromatography using fluorescence monitoring. The substrate specificity of FMO1 and FMO3 toward quinuclidine N-oxygenation in monkeys, dogs, and pigs was suggested to be relatively broad. Human FMO3-mediated quinuclidine N-oxygenation was slower than trimethylamine N-oxygenation. The likelihood of quinuclidine, with its simple chemical structure, being clinically inhibited by co-administered drugs is relatively low.