Cytochrome P450 3A enzymes are emerging as one of the most important enzyme systems involved in the metabolism of drugs, pesticides and carcinogens, particularly in humans. Our overall goal is to understand how the function and regulation of the P450 3A enzymes differ among mammalian species and individuals. This goal is an extension of our studies on the mechanism by which catatoxic steroids protect rats against chemical toxicity. Previously, we determined that treatment of rats with catatoxic steroids induces a P450 3A enzyme (Mr 50 kDa) that detoxifies digitoxin (by oxidative cleavage to dt2 and dt1). We hypothesize that this 50-kDa P450 3A enzyme is unique to rats, and that this enzyme also catalyzes the 4'-hydroxylation of (S)-mephenytoin. Our first specific aim is to test this hypothesis, which was formulated to explain why P450 3A enzymes play a major role in the metabolism of digitoxin and (S)-mephenytoin in rats, whereas the metabolism of these drugs in other species is either negligible or is not catalyzed by P450 3A enzymes. A second specific aim is to test the hypothesis that treatment with catatoxic steroids will afford no protection against the toxic effects of digitoxin in those species (hamster and mouse) that lack the 50-kDa P450 3A - enzyme and cannot convert digitoxin to dt2 and dt1. In testing this hypothesis, we aim to establish whether species differences in the function and regulation of P450 3A enzymes are responsible for species differences in the protective effects of catatoxic steroids. A third specific aim is to test the hypothesis that P450 3A levels in human and monkey liver are higher in females than males, and are lower in aged subjects than in young adults. To test this hypothesis, we propose to examine the rate of metabolism of several P450 3A substrates by liver microsomes from adult male and female cynomolgus monkeys and human subjects of various ages. This hypothesis will be tested in an attempt to establish whether certain age- and gender-related differences in drug metabolism (which have been observed clinically) are due to developmental changes and sexually dimorphic expression of P450 3A enzymes. A fourth specific aim is to test the hypothesis that P450 3A enzymes are widely distributed in extrahepatic tissues of primates but not rats. To test this hypothesis, the levels of P450 3A enzymes will be determined in kidney, lung, intestine and testis from rat, monkey and human. The hypothesis that the P450 3A enzymes in primate and rat liver are regulated differently will be tested by examining the effects of drugs and hormones on the expression of P450 3A enzymes in cultured hepatocytes from rat and monkey or human. These studies will explore the basis for species differences in the pharmacological and toxicological effects of drugs, which will contribute significantly to our understanding of the underlying cause of certain idiosyncratic responses to drugs, and the reason for gender- and age- dependent changes in the disposition of several clinically important drugs.
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