Structural forms of phenprocoumon and warfarin that are metabolized at theactive site of CYP2C9

Possible reasons for the observed differences in metabolic behavior and dru g interaction liability between the structurally similar oral anticoagulant s warfarin and phenprocoumon were explored. Incubating (S)-phenprocoumon wi th human liver microsomes and cDNA-expressed CYP2C9 and determining its met abolism both in the absence and presence of the CYP2C9 inhibitor, sulfaphen azole, confirmed that phenprocoumon is a substrate for CYP2C9, Comparing th e metabolic behavior of (S)- and (R)-warfarin, (S)- and (R)-phenprocoumon, and fixed structural mimics of the various tautomeric forms [(S)- and (R)-4 -methoxyphenprocoumon, (S)- and (R)-2-methoxyphenprocoumon, (S)- and (R)-4- methoxywarfarin, (S)- and (R)-2methoxywarfarin, and 9(S)- and S(R)-cyclocou marol] available to these two drugs with expressed CYP2C9 provides compelli ng evidence indicating that the ring closed form of (S)-warfarin and the ri ng opened anionic form of (S)-phenprocoumon are the major and specific stru ctural forms of the two drugs that interact with the active site of CYP2C9, The conclusion that (S)-warfarin and (S)-phenprocoumon interact with CYP2C 9 in very different structural states provides a clear basis for the signif icant differences observed in their metabolic profiles. Moreover, in accord with a previously established CoMFA model these results are consistent wit h the hypothesis that the active site of CYP2C9 possesses at least two majo r substrate binding sites, a pi-stacking site for aromatic rings and an ion ic binding site for organic anions, An additional electrostatic binding sit e also appears to contribute to the orientation of coumarin analogs in the CYP2C9 active site by interacting with the Ca-carbonyl group of the coumari n nucleus. (C) 1999 Academic Press.