Regulation of Drug Transport Proteins and Drug Metabolizing Enzymes by Nuclear Receptors

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Background - The intracellular concentration of chemical molecules determines their activity and toxicity. This applies not only to drugs but also to endogenous molecules such as vitamins and hormones. Membrane transport proteins act as a gateway to determine the concentration available to act on intracellular targets. Intracellular enzymes mediate the metabolism of numerous molecules to detoxify them and/or render them more suitable for elimination. Proteins called nuclear receptors tightly regulate the amount of these transporters and metabolic enzymes. The overall goal of this research program is to further our understanding of the processes that determine the activity of drug transporters and metabolic enzymes. This will help us understand the way the body handles drugs, vitamins and other hormones. Objectives - The objectives of the proposed research program are to: 1) Investigate the role of a hormone called erythropoietin in mediating the downregulation of metabolic enzymes, 2) To determine the role of intestinal bacteria and the metabolites they produce on transporters and metabolic enzymes, and 3) To determine the impact of nuclear receptor activity on intracellular metabolic pathways. Scientific Approach - Previous studies in our lab determined a hormone made in the kidney called erythropoietin (EPO), significantly decreases expression and activity of an essential metabolic enzyme (CYP3A). To elucidate the mechanism for this effect, we will isolate cells of the liver and treat them with EPO as well as various inhibitors of EPO signalling to determine the mechanism accounting for this effect. To evaluate the impact of intestinally produced bacterial metabolites on nuclear receptor mediated expression of transporters and metabolic enzymes, we will use germ free mice and antibiotics to alter the intestinal bacterial content. We will then use metabolomic analysis, which allows us to identify and measure the differences in metabolite formation. We will use this technology to determine the changes these treatments induce in target organs such as the intestine, liver and kidney. Although the contribution of nuclear receptors to regulation of enzymes and transporters is well appreciated in terms of response to drugs, the endogenous biological role of these processes is unknown. We will use established nuclear receptor knockout mouse models of two of important nuclear receptors (e.g. Pxr and Car) to perform genomic and metabolomic analysis in order to identify novel pathways regulated by these essential proteins. Expected Significance -The findings from this program of research will elucidate the mechanisms associated with modulation in gene expression of membrane transport proteins and metabolic enzymes at the molecular level. Ultimately these investigations will help us understand how the body handles and processes exogenous and endogenous compounds.