Obesity is a well-established risk factor for a number of diseases, including type 2 diabetes and coronary heart disease. Existing drug and dietary treatments for obesity are only modestly effective. An approach that is very likely to be effective in treating obesity and with a good side effect profile is a drug that acts directly on lipid metabolism in the fat cell. The mission of AdipoGenix, Inc. is to discover and develop novel therapeutics acting at the level of the fat cell for treating obesity, diabetes, and related metabolic disorders. The enzyme carnitine palmitoyltransferase I (CPT I) is the critical control point for fatty acid metabolism in the cell and provides the main switch between free fatty acid (FFA) oxidation to CO2 and esterification to triglyceride (TG). A slight change in the balance between oxidation and synthesis could have a major impact on fat stores. Malonyl-CoA, an allosteric inhibitor of CPT I, regulates FFA oxidation and, consequently, fat storage. Compounds that interfere with the effect of malonyl-CoA on CPT I can promote CPT I activity and, thereby, FFA oxidation. We propose to characterize the differentiation- and fat depot-dependent expression of CPT I isoforms in human preadipocytes, and to develop high-throughput screens to identify compounds that relieve the inhibitory effect of malonyl-CoA on CPT I and thereby stimulate oxidation. Relative quantitative RTPCR and Western blotting will be used to analyze expression of CPT I isoforms in human adipocytes during differentiation and from different anatomical depots. A high-throughput assay to measure activity of CPT I in isolated mitochondrial preparations and in permeabilized human adipocytes in the presence and absence of the inhibitor malonyl-CoA will be developed. In parallel, a high-throughput assay to measure oxygen consumption in human adipocytes using an oxygen-sensing microplate-based system will be developed. The assay determined to be most suitable for HTS will be used to screen chemical libraries in Phase II. Achievement of these aims will establish a viable HTS assay and secondary assays for identifying compounds that activate CPT I and increase oxidation in human adipocytes. Development of such compounds may lead to therapeutic interventions that are effective for modulating lipid metabolism and treating obesity.
