RT Journal Article SR Electronic T1 Investigation into MAO B-mediated formation of CC112273, a major circulating metabolite of ozanimod, in humans and preclinical species: Stereospecific oxidative deamination of (S)-enantiomer of indaneamine (RP101075) by MAO B JF Drug Metabolism and Disposition JO Drug Metab Dispos FD American Society for Pharmacology and Experimental Therapeutics SP DMD-AR-2021-000447 DO 10.1124/dmd.121.000447 A1 April Bai A1 Veerabahu .Shanmugasundaram A1 Julie V. Selkirk A1 Sekhar Surapaneni A1 Deepak Dalvie YR 2021 UL http://dmd.aspetjournals.org/content/early/2021/05/19/dmd.121.000447.abstract AB Ozanimod, recently approved for treating relapsing MS, produced a disproportionate, active, MAO B-catalyzed metabolite (CC112273) that showed remarkable interspecies differences and led to challenges in safety testing. This study explored the kinetics of CC112273 formation from its precursor RP101075. Incubations with human liver mitochondrial fractions revealed KMapp, Vmax and Clint for CC112273 formation to be 4.8 mM, 50.3 pmol/min/mg protein and 12 ml/min/mg, respectively, while KM with human recombinant MAO B was 1.1 mM. Studies with liver mitochondrial fractions from preclinical species led to KMapp, Vmax and Clint estimates of 3.0, 35 and 33 mM, 80.6, 114, 37.3 pmol/min/mg and 27.2, 3.25 and 1.14 ml/min/mg in monkey, rat and mouse, respectively, and revealed marked differences between rodents and primates, primarily attributable to differences in the KM. Comparison of Clint estimates revealed monkey to be ~two-fold more efficient and the mouse and rat to be 11 and 4-fold less efficient than humans in CC112273 formation. The influence of stereochemistry on MAO B-mediated oxidation was also investigated using the R-isomer of RP101075 (RP101074). This showed marked selectivity towards catalysis of the S-isomer (RP101075) only. Docking into MAO B crystal structure suggested that even though both the isomers occupied its active site, only the orientation of RP-101075 presented the C-H on the a-carbon that was ideal for the C-H bond cleavage, which is a requisite for oxidative deamination. These studies explain the basis for the observed interspecies differences in the metabolism of ozanimod as well as the substrate stereospecificity for formation of CC112273. Significance Statement This study evaluates the enzymology and the species differences of the major circulating metabolite of ozanimod, CC112273. Additionally, the study also explores the influence of stereochemistry on MAO B catalyzed reactions. The study is of significance to the DMD readers given that this oxidation is catalyzed by a non-CYP enzyme and that marked species difference and notable stereospecificity was observed in MAO B catalyzed biotransformation when the indaneamine enantiomers were used as a substrates.