Novel Factors Promoting Retinal Ganglion Cell Growth

描述（由申请人提供）：NIH路线图强调生物活性“小分子”的发现如何有助于探索“分子水平上的细胞功能”和开发新药。为了反映这类分子的重要性，分子文库路线图的倡议之一是支持小分子文库的构建和筛选。小分子和多肽通过调节各种细胞信号通路支持视网膜神经节细胞（RGC）的存活和神经突的生长。鉴定额外的这类分子是这项拨款的重点。Forskolin和BDNF是最初在培养中发现支持RGCs的分子的例子，后来在视神经损伤的动物模型中证明了它们的有效性。在细胞培养中鉴定支持rgc的新型小分子提供了两个潜在的好处。这样的分子将使在分子水平上研究RGC功能的新思路成为可能。其次，在细胞培养中支持RGCs的因素是候选因子的逻辑来源，可能在保护或恢复患有致盲性视神经疾病（如青光眼、视神经中风和视神经炎）的患者的视力方面具有转化和治疗益处。与大分子（如蛋白质）相比，小分子具有潜在的优势，因为它们更容易被开发成药物，也更容易被输送到眼睛。将灵敏的生物测定法与高通量筛选相结合是鉴定功能感兴趣的小分子的一种合理而全面的方法。我们已经成功启动了这样的筛选，用自动荧光图像分析系统检测免疫纯化大鼠RGCs的培养物，该系统测量细胞存活和神经突生长。本提案的目的是扩大筛选范围，以鉴定、验证和优化促进rgc存活和再生的新型药物化合物。更具体地说，这笔拨款将支持继续调查RGC存活和神经突生长因子的“小分子”文库，通过证明剂量反应关系来确认这些命中的生物活性，并进一步确定显示生物活性的基本元素。筛选中确定的试剂将成为未来假设驱动研究的主题，以表征其体外作用机制，并进一步探索其体内治疗潜力。

DESCRIPTION (provided by applicant): The NIH Roadmap emphasizes how the discovery of bioactive "small molecules" can help both in the exploration of "cellular functions at the molecular level" and the development of new drugs. Reflecting the importance of such molecules, one of the Molecular Libraries Roadmap's Initiatives is to support the construction and screening of small molecule libraries. Small molecules and peptides support retinal ganglion cell (RGC) survival and neurite outgrowth by modulating various cellular signaling pathways. Identification of additional such molecules is the focus of this grant. Forskolin and BDNF are examples of molecules that that were originally found to support RGCs in culture and later demonstrated to have efficacy in animal models of optic nerve injury. Identification of novel small molecules that support RGCs in cell culture offers two potential benefits. Such molecules will make possible new lines of inquiry into RGC function at the molecular level. Secondly, factors supporting RGCs in cell culture are a logical source of candidates that may have translational, therapeutic benefit in preserving or restoring vision in patients with blinding optic nerve diseases such as glaucoma, optic nerve stroke, and optic neuritis. Small molecules have potential advantages compared to larger molecules, such as proteins, in that they may be more readily developed into drugs and may be more easily delivered to the eye. A rational and comprehensive way to identify functionally interesting small molecules is to combine a sensitive bioassay with a high throughput screen. We have successfully initiated such a screen, assaying cultures of immunopurified rat RGCs with an automated fluorescence-based image analysis system that measures cell survival and neurite outgrowth. The objective of this proposal is to expand this screen so as to identify, validate, and optimize novel pharmaceutical compounds that promote survival and regeneration of RGCs. More specifically, this grant will support the continued survey of a "small molecule" library for RGC survival and neurite outgrowth factors, confirm the bioactivity of those hits by demonstrating a dose response relationship, and further determine the essential element that exhibits the bioactivity. Reagents identified in the screen will be the subject of future hypothesis driven research to characterize their mechanism of action in vitro and further explore their therapeutic potential in vivo.