PREDICTION OF DRUG INTERACTIONS IN VIVO AND IN VITRO

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The overall objective of this proposal is to evaluate a new framework for the prediction of inhibition based drug interactions. This framework is based on the ability to characterize the effects of inhibitors on specific cytochrome P450 isoforms using human liver microsomal preparations. Two hypotheses will be tested: Hypothesis 1: The mechanism of any drug-inhibitor in vivo interaction can be accounted for from (i) a determination of the specific cytochrome P450 isoforms responsible for that drug's major route of metabolism and (ii) a measurement of the inhibitor in vitro Ki for each isoform. Hypothesis 2: For any drug-inhibitor in vivo interaction, the Michaelis-Menton model can be used to calculate an isoform specific in vivo inhibition constant (Kiiv). This constant provides a quantitative estimate of the magnitude of interaction to be expected between that inhibitor and any substrate. These hypotheses will be evaluated with two classes of drugs for which inhibition based interactions constitute a critical clinical reality. The following Specific Aims will be pursued: 1(a) To determine, using human liver microsomes, if the clinical interactions between warfarin and 13 commonly prescribed drugs are a consequence of the inhibition of CYP2C9. 1(b) To determine, using human liver microsomes, if the clinical interactions between 4 major anticonvulsants and 33 other drugs are a consequence of inhibition of the specific isoforms responsible for the major fractional clearances of the anticonvulsants. If they are, to determine the in vitro Ki's for the inhibition of these isoforms and to compare these values to the corresponding Kiiv's estimated from literature data. 2(a) To demonstrate, for isoform CYP2C9, that (i) the degree of interaction between (S)-warfarin and fluconazole can be predicted quantitatively by the ratio, I/Kiiv, where I is the plasma inhibitor concentration and (ii) the same Kiiv for 2C9 is obtained for fluconazole when phenytoin is used as the substrate. 2(b) To demonstrate, for isoform 3A4, that (i) the degree of interaction between (R)-warfarin and fluconazole can be predicted quantitatively by the ratio, I/Kiiv, and (ii) the same Kiiv for 3A4 is obtained for fluconazole when carbamazepine is used as the substrate. 2(c) To demonstrate that the approach of specific aims 2(a) and 2(b) is inhibitor independent, a second inhibitor will be used with the same substrates for each isozyme. The experimental approaches include the use of human liver bank, reliable analytical techniques and a number of human studies in healthy subjects. If the proposed hypotheses are validated it will become possible to predict inhibition based interactions for any new drug.