Characterization of cytochrome P450 enzymes and P-glycoprotein involved in the metabolism and transport of a new anti-angiogenic agent KR-31831

Abstract

The in vitro metabolism and the transport of a novel anti-angiogenic agent KR-31831, (2R,3R,4S)-6-amino-4-[N-(4-chlorophenyl)-N-(1H-imidazol-2-ylmethyl)amino]-3-hydroxy-2-dimethyoxymethyl-3,4-dihydro-2-methyl-2H-1-benzopyran were investigated. Liquid chromatography-mass spectrometry and tandem mass spectrometry were used for qualitative and quantitative analysis. The bidirectional transport studies of KR-31831 using Caco-2 cell monolayers showed the efflux to be significantly higher than influx (29.1 × 10−6 compared to 11.5 × 10−6 cm/s). P-glycoprotein inhibitors significantly increased the influx of KR-31831 and decreased the efflux of KR-31831. These data indicate that KR-313831 is a substrate for an efflux pump, P-glycoprotein. The incubations of KR-31831 with human liver microsomes produced three metabolites, M1, M2, and M3. M1 and M2 were identified as N-(4-chlorophenyl)-N-(1H-imidazol-2-ylmethyl)amine and (2R,3R,4S)-6-amino-4-[N-(4-chlorophenyl)-N-(1H-imidazol-2-ylmethyl)amino]-3-hydroxy-2-hydroxymethyl-3,4-dihydro-2-methyl-2H-1-benzopyran by comparison with the authentic standards. M3 was tentatively characterized as hydroxy-KR-31831. CYP3A4 was identified as the major enzyme responsible for KR-31831 metabolism to a major metabolite M1 using the combination of correlation analysis, immuno-inhibition, chemical inhibition in human liver microsomes, and metabolism by cDNA expressed CYP enzymes. There is the possibility of drug–drug interactions when prescribing KR-31831 concomitantly with known inhibitors or inducers of CYP3A4 and P-glycoprotein. KR-31831 was found to inhibit potently the metabolism of CYP2D6 substrate, suggesting that coadministration of KR-31831 with CYP2D6 substrates may have significant effects on the pharmacokinetics of CYP2D6 substrates. Drug Dev. Res. 66:40–49. 2006. © 2006 Wiley-Liss, Inc.