Interactions of two major metabolites of prasugrel, a thienopyridine antiplatelet agent, with the cytochromes P450

The biotransformation of prasugrel to R- 138727 ( 2-[ 1- 2- cyclopropyl-1-( 2- fluorophenyl)- 2- oxoethyl]- 4- mercapto- 3- piperidinylidene] acetic acid) involves rapid deesterification to R- 95913 ( 2-[ 2- oxo- 6,7dihydrothieno[ 3,2- c] pyridin- 5( 4H)- yl]- 1- cyclopropyl- 2-( 2- fluorophenyl) ethanone) followed by cytochrome P450 ( P450)- mediated formation of R- 138727, the metabolite responsible for platelet aggregation. For identification of the P450s responsible for the formation of the active metabolite, the current studies were conducted with R- 95913 as the substrate. Incubations required supplementation with reduced glutathione. Hyperbolic kinetics ( K-m 21 - 30 mu M), consistent with a single enzyme predominating, were observed after incubations with human liver microsomes. Correlation analyses revealed a strong relationship between R- 138727 formation and CYP3A- mediated midazolam 1 '- hydroxylation ( r(2) = 0.98; p < 0.001) in a bank of characterized human liver microsomal samples. The human lymphoblast-expressed enzymes capable of forming R- 138727, in rank order of rates, were CYP3A4 > CYP2B6 > CYP2C19 approximate to CYP2C9 > CYP2D6. A monoclonal antibody to CYP2B6 and the CYP3A inhibitor ketoconazole substantially inhibited R- 138727 formation, whereas inhibitors of CYP2C9 ( sulfaphenazole) and CYP2C19 ( omeprazole) did not. Scaling of in vitro intrinsic clearance values from expressed enzymes to the whole liver using a relative abundance approach indicated that either CYP3A4 alone or CYP3A4 and CYP2B6 are the major contributors to R- 138727 formation. R- 95913 and R- 138727 were also examined for their ability to inhibit metabolism mediated by five P450s. R- 138727 did not inhibit the P450s tested. In vitro, R- 95913 inhibited CYP2C9, CYP2C19, CYP2D6, and CYP3A, with K-i values ranging from 7.2 mu M to 82 mu M, but did not inhibit CYP1A2. These K-i values exceed circulating concentrations in humans by 3.8- to 43- fold. Therefore, neither R- 95913 nor R- 138727 is expected to substantially inhibit the P450- mediated metabolism of coadministered drugs.