INHIBITORS OF MAST CELLS DEGRANULATION AND TRPM CATION CHANNELS FROM MARINE ORG

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The structural diversity of secondary metabolites of marine organisms makes their extracts effective chemical libraries for bioassay screening to identify biologically active small molecules. The overall goal of the proposed research is (a) to develop robust methods for the production of high-quality prefractionated marine natural product chemical libraries using high performance liquid chromatography with mass spectrometry and evaporate light scattering detection (HPLC-MS-ELSD) and (b) to implement these methods toward the discovery of potent specific inhibitors of mast cell degranulation and TRPM-class cation channels from extracts of marine organisms. Mast cells are recognized as potent effector cells in inflammatory diseases, including allergic asthma and arthritis. TRPM cation channels are implicated in a range of cellular functions in cells of the immune and nervous systems. Specific inhibitors of mast cell activation, or TRPM channels, are of potential importance as therapeutics or research tools. The first specific aim of the proposed research comprises the establishment of a cell culture laboratory at HPU, and development of assays for mast cell activation and TRPM channel function that can be used to screen marine organism-derived chemical libraries. Concurrently, methods will be developed for the prefractionation of organism extracts by HPLC-MS-ELSD into 96-well plate format, replication of plates for bioassay, archiving of collected fractions, and analysis of MS and bioassay data. Acquisition and documentation of taxonomically diverse samples from cultured and collected marine cyanobacteria, algae, and invertebates will be on-going. Prioritization and dereplication strategies will be employed utilizing correlations between MS profiles and bioactivity, database searches, and nuclear magnetic resonance (NMR) spectroscopy. Active components from highly promising extracts will be isolated by bioassay-and MS-directed fractionation and structually characterized primarily by NMR and MS.