Identification and Characterization of a Selective Human Carbonyl Reductase 1 Substrate.

Abstract

During drug discovery efforts targeting inhibition of cytochrome P450 11B2 (CYP11B2)-mediated production of aldosterone as a therapeutic approach for the treatment of chronic kidney disease and hypertension, (S)-6-(5-fluoro-4-(1-hydroxyethyl)pyridin-3-yl)-3,4-dihydro-1,8-naphthyridine-1(2H)-carboxamide (1) was identified as a potent and selective inhibitor of CYP11B2. Preclinical studies characterized 1 as low clearance in both in vitro test systems and in vivo in preclinical species. Despite low metabolic conversion, an active ketone metabolite (2) was identified from in vitro metabolite-identification studies. Due to the inhibitory activity of 2 against CYP11B2 as well as the potential for it to undergo reductive metabolism back to 1, the formation and elimination of 2 were characterized and are the focus of this manuscript. A series of in vitro investigations determined that 1 was slowly oxidized to 2 by cytochrome P450s 2D6, 3A4, and 3A5, followed by stereoselective reduction back to 1 and not its enantiomer (3). Importantly, reduction of 2 was mediated by an NADPH-dependent cytosolic enzyme. Studies with human cytosolic fractions from multiple tissues, selective inhibitors, and recombinantly expressed enzymes indicated that carbonyl reductase 1 (CBR1) is responsible for this transformation in humans. Carbonyl reduction is emerging as an important pathway for endogenous and xenobiotic metabolism. With a lack of selective substrates and inhibitors to enable characterization of the involvement of CBR1, 2 could be a useful probe to assess CBR1 activity in vitro in both subcellular fractions and in cell-based systems.