The importance of understanding drug interactions because of the frequency of their occurrence and the potential for serious therapeutic consequences is well documented. The ultimate aim of this proposal is to develop a structural framework from which drug interactions can be understood at the molecular level. Achievement of this goal requires that, correlation between in vivo and in vitro data be established. As a result, isolation and purification of the enzyme systems responsible for drug biotransformation is central to this approach. In Project I, the oral anticoagulants warfarin and phenprocoumon have been selected as model drugs to 1) probe the pharmacokinetic basis of drug interactions, 2) provide a foundation from which to build in vivo-in vitro correlations, 3) use inhibitor induced changes in their metabolic activities to assign specific activities to specific isozymes and 4) use their metabolic profiles to follow enzyme purification. Project II will focus on interactions between calcium blockers and beta-adrenoreceptor antagonists. Stereoselective effects of verapamil on in vivo metabolism of metoprolol in angina patients and in vitro metabolism in isolated human liver microsomes will be examined using stable isotope labeled pseudoracemates of metoprolol. The ability of verapamil to inhibit in vitro metabolism of debrisoquine-type polymorphic substrates in human liver microsomes will also be investigated. Project III will focus on drug interactions prevalent in anticonvulsant therapy. Concern has arisen over the role of drug interactions in the pathogenesis of a rare, but fatal liver injury associated with valproic acid (VPA) therapy. Recent evidence suggests that unsaturated metabolites of VPA may be causative agents. The linkage between olefin formation and cytochrome P-450 will be examined using purified isozymes. The effect of delta 4- VPA on 3-ketothiolase (a critical enzyme involved in fatty acid metabolism) will be examined. A second type of drug-drug interaction involving VPA will be examined. Unexpectedly, VPA and VPD were found to inhibit conversion of the epoxide metabolite of carbemazepine to the corresponding dihydrodiol. Purified epoxide hydrolase from human liver will be studied to define the scope and molecular basis for the inhibition. Project IV will consider drug- drug interactions which occur when one drug modulates basis for the inhibition. Project IV will consider drug-drug interactions which occur when one drug modulates the cytotoxicity of a second drug by altering concentrations of reactive metabolites of the second drug in a target tissue. Acetaminophen, a drug that is widely used in combination with other drugs, will be used in this model. To this end, the multiple effects of caffeine and isoniazid on acetaminophen hepatotoxicity, and the effects of acetaminophen on the metabolism and toxicity of ethanol and the AIDS drug, azidothymidine will be examined. The knowledge gained in the proposed studies can be expected to yield that impact many facets of drug therapy.
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