Summary

Like ticlopidine, the ADP receptor antagonist clopidogrel is inactivein vitro and must be administered i.v. or orally to exhibit antiaggrega-toryand antithrombotic activities. We have previously shown that he-paticmetabolism is necessary for activity. This study demonstrates thatan active metabolite can be generated from human liver microsomesincubated with clopidogrel. Using several analytical methodologies(LC/MS, NMR, chiral supercritical fluid chromatography), we haveidentified its structure. In vitro, this highly unstable compound, differentfrom that formed from ticlopidine, exhibited all the biological activitiesof clopidogrel observed ex vivo: Irreversible inhibition of the bindingof ³³ P-2MeS-ADP to washed human platelets (IC ₅₀ = 0.53 μM), selectiveinhibition of ADP-induced platelet aggregation (IC ₅₀ = 1.8 μM)and ADP-induced adenylyl cyclase down-regulation. The irreversiblemodification of the ADP-receptor site which is responsible for thebiological activity could be explained by the formation of a disulfidebridge between the reactive thiol group of the active metabolite and acysteine residue of the platelet ADP receptor.