The long-term goal of this project is to develop tools for high throughput testing of drug compounds from combinatorial libraries that target G-Protein-Coupled Receptors (GPCR's) associated with appetite control and eating behavior. The development of drugs aimed at reducing obesity and controlling eating disorders represents a major target of the pharmaceutical industry. Novel targets for such drug development arise from the recent explosion of information on molecules and neural circuits involved in appetite regulation. This proposal focuses on developing testing platforms for two GPCR families associated with feeding behavior--the Melanocortin Receptors (MCRs) and the Orexin Receptors (OXR). ? ? High throughput testing of GPCR's requires an efficient and highly reliable system that allows multiplexing of receptors in order to test numerous compounds against numerous targets. This project will take advantage of a novel transfection technology that is licensed by Originus, Inc. from the University of Michigan termed Surface Transfection and Expression Protocol (STEP), which has significant advantages over transient and even stable expression methods ? ? In Phase I, we will use existing ligands to validate the platform. Future screening of new drugs will be carried out in a Phase I submission.
The Specific Aims for Phase I are: ? ? (1) The use and optimization of STEP transfection for 384 well microplate transfection to test the expression and ligand activation of individually expressed feeding-related receptors including MCRs and one of the OXR's. ? ? (2) Multiplexing via the co-expression of multiple feeding-related receptors belonging to the same receptor family (MCR's) in order to enhance efficiency of testing of potential targets. Phase II will build on the information from Phase I to test other receptors with different intracellular coupling, to multiplex both within and across receptor families and to facilitate searching for drugs that are agonists and antagonists and inverse agonists. ? ? Originus has worked with several partners from pharmaceutical industry in the past and has a specific partner interested in the development of multiplexed GPCR-expressing microplates. Moreover, this project will be the prototype for the future creation of multiplexed GPCR-bearing microplates beyond the feeding-related ones. These would be used for testing a wide range of compounds targeted at modifying cell signaling in a variety of disorders. ? ? ?
