Use of cDNA-Expressed Human Cytochrome P450 Enzymes to Study Potential Drug-Drug Interactions

Publisher Summary

cDNA-expressed enzymes can be used to study the cytochrome P450 form-selective inhibition by drugs or drug candidates. This analysis is accomplished through the study of the inhibition of the metabolism of a model substrate by the drug or drug candidate. Through these analyses, apparent K_i values can be obtained and compared to K_i values for known, clinically significant inhibitors of the same enzyme. A coenzyme, cytochrome P450 NADPH oxidoreductase (OR), is essential for P450 catalytic function, and cytochrome bc can stimulate catalytic activities of some enzymes. The levels of these two coenzymes affect the rate of substrate metabolism per unit enzyme. Certain P450 forms are known to be polymorphic in humans and some are regulated in response to exposure to environmental agents. Two principal approaches have been developed to study form specificity in cytochrome P450-mediated metabolism that use either human tissue fractions or cloned/expressed enzymes. These two approaches are complementary and both are often used to establish form specificity. Cytochrome P450 cDNAs have been heterologously expressed in a variety of systems, including bacteria, yeast, mammalian cells, and insect cells. There appears to be no one “ideal” heterologous expression system. When multiple enzymes are involved in metabolism, the rates of metabolism can be expressed as nanomoles of metabolite per nanomole of P450. However, human liver does not contain equimolar concentrations of the different P450 forms, and a rigorous quantitation of the levels of all the different P450 forms has not been performed, particularly for the CYP2C subfamily.