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

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。海洋生物次生代谢物的结构多样性使其提取物成为生物测定筛选的有效化学文库，可用于鉴定具有生物活性的小分子。该研究的总体目标是：(a)开发高效液相色谱-质谱和蒸发光散射检测（HPLC-MS-ELSD）生产高质量预分离海洋天然产物化学文库的可靠方法；(b)实施这些方法，从海洋生物提取物中发现肥大细胞脱颗粒和trpm类阳离子通道的有效特异性抑制剂。肥大细胞被认为是炎症性疾病的有效效应细胞，包括过敏性哮喘和关节炎。TRPM阳离子通道涉及免疫和神经系统细胞的一系列细胞功能。肥大细胞活化的特异性抑制剂，或TRPM通道，作为治疗或研究工具具有潜在的重要性。拟议研究的第一个具体目标包括在HPU建立一个细胞培养实验室，并开发可用于筛选海洋生物衍生化学文库的肥大细胞激活和TRPM通道功能的检测。同时，将开发生物提取物的HPLC-MS-ELSD预分离成96孔板格式，生物测定板的复制，收集的部分存档，以及MS和生物测定数据的分析方法。从培养和收集的海洋蓝藻，藻类和无脊椎动物的分类多样化样品的获取和文件将继续进行。优先级和反复制策略将利用质谱与生物活性、数据库搜索和核磁共振（NMR）光谱之间的相关性。从极具潜力的提取物中分离出活性成分，并通过生物测定和质谱定向分离，主要通过核磁共振和质谱进行结构表征。

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.