RT Journal Article SR Electronic T1 Phosphocholine Conjugation: An Unexpected In Vivo Conjugation Pathway Associated with Hepatitis C NS5B Inhibitors Featuring A Bicyclo[1.1.1]Pentane JF Drug Metabolism and Disposition JO Drug Metab Dispos FD American Society for Pharmacology and Experimental Therapeutics SP 1332 OP 1340 DO 10.1124/dmd.115.069062 VO 44 IS 8 A1 Xiaoliang Zhuo A1 Joseph L. Cantone A1 Yingzi Wang A1 John E. Leet A1 Dieter M. Drexler A1 Kap-Sun Yeung A1 Xiaohua Stella Huang A1 Kyle J. Eastman A1 Kyle E. Parcella A1 Kathleen W. Mosure A1 Matthew G. Soars A1 John F. Kadow A1 Benjamin M. Johnson YR 2016 UL http://dmd.aspetjournals.org/content/44/8/1332.abstract AB During a medicinal chemistry campaign to identify inhibitors of the hepatitis C virus nonstructural protein 5B (RNA-dependent RNA polymerase), a bicyclo[1.1.1]pentane was introduced into the chemical scaffold to improve metabolic stability. The inhibitors bearing this feature, compound 1 [5-(3-(bicyclo[1.1.1]pentan-1-ylcarbamoyl)-4-fluorophenyl)-2-(4-fluorophenyl)-N-methyl-6-(3,3,3-trifluoropropyl)furo[2,3-b]pyridine-3-carboxamide] and compound 2 [5-(3-(bicyclo[1.1.1]pentan-1-ylcarbamoyl)phenyl)-2-(4-fluorophenyl)-N-methyl-6-(3,3,3-trifluoropropyl)furo[2,3-b]pyridine-3-carboxamide], exhibited low turnover in incubations with liver S9 or hepatocytes (rat, human), with hydroxylation of the bicyclic moiety being the only metabolic pathway observed. In subsequent disposition studies using bile duct–cannulated rats, the metabolite profiles of bile samples revealed, in addition to multiple products of bicyclopentane oxidation, unexpected metabolites characterized by molecular masses that were 181 Da greater than those of compound 1 or 2. Further liquid chromatography/multiple-stage mass spectrometry and nuclear magnetic resonance analysis of the isolated metabolite of compound 1 demonstrated the presence of a phosphocholine (POPC) moiety bound to the methine carbon of the bicyclic moiety through an ester bond. The POPC conjugate of the nonstructural protein 5B inhibitors was assumed to result from two sequential reactions: hydroxylation of the bicyclic methine to a tertiary alcohol and addition of POPC by cytidine-diphosphocholine:1,2-diacylglycerol cholinephosphotransferase, an enzyme responsible for the final step in the biosynthesis of phosphatidylcholine. However, this pathway could not be recapitulated using cytidine-diphosphocholine–supplemented liver S9 or hepatocytes because of inadequate formation of the hydroxylation product in vitro. The observation of this unexpected pathway prompted concerns about the possibility that compounds 1 and 2 might interfere with routine phospholipid synthesis. These results demonstrate the participation in xenobiotic metabolism of a process whose function is ordinarily limited to the synthesis of endogenous compounds.