Identification of protease activated receptor-2 modulators

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DESCRIPTION (provided by applicant): Activation of G protein-coupled protease-activated receptor-2 (PAR2) in peripheral organs by the serine proteases trypsin and mast cell-derived tryptase exacerbates inflammatory processes and inflammatory disorders such as arthritis, asthma, and allergies. Neuropathological conditions such as stroke, epilepsy, and neurodegenerative diseases- involve complex neuroinflammatory processes that can exacerbate brain damage. Recent studies suggest that levels of PAR2 and its activating proteases also increase in the brain during injury and damage, suggesting that PAR2 activation may contribute to the process of inflammation in brain injury. Neuroinflammatory responses are driven largely by reactive microglia and astrocytes, and both cell types exhibit dramatic morphological changes and proliferation in response to PAR2-selective activating peptides. These observations together give rise to the hypothesis that PAR2 activation plays a pro- inflammatory role in the brain through its effects on microglial and astrocytic functions. Despite the intriguing and important roles postulated for PAR2 in both peripheral and CMS, no small molecule antagonists or modulators have been reported for this receptor in the literature. Furthermore, no small molecule antagonists or modulators are available from commercial sources. To facilitate the evaluation of the role of PAR2 in CNS injury and explore therapeutic opportunities that follow PAR2 modulation, we propose to develop a robust assay for PAR2 allosteric regulators. We propose to specifically search for small molecules that regulate the receptor in a non-competitive fashion since such molecules often bind to sites with rich and selective pharmacology, and modulators can offer advantages including increased safety and reduced side effects in the clinic. The aims of this grant are: 1. To define an assay that will allow the identification of small molecule inhibitors and potentiators. 2. To convert assay to 384 well format and design secondary assays that will allow the identification of competitive inhibitors as well as positive and negative allosteric modulators.