Fully Automated Angiogenesis Drug Discovery Assay in Zebrafish

描述（由申请人提供）：斑马鱼，Danio rerio，已被广泛接受为人类疾病的模型。小分子筛选已被用于识别血管模式的修饰剂，但由于全自动处理的工具尚未可用，因此该过程的吞吐量相对较低。我们建议开发一种新的、高通量的血管生成促进剂筛选方法。该试验将基于使用“SideView”微孔板来收集血管中表达GFP的动物血管模式的清晰图像。在第一阶段，我们：1)开发了调节神经丛血管生成促进因子的新方法；2)开发软件，定位斑马鱼，识别感兴趣的区域，并将该区域移动到视野中心；3)开发了自动采集后图像处理软件，用于区分野生型斑马鱼胚胎和神经丛畸形的XXX突变胚胎。在第二阶段，方法、软件和仪器将完全集成，该分析将使用FDA批准的化合物的Prestwick库在模型筛选中进行验证。

DESCRIPTION (provided by applicant): The zebrafish, Danio rerio, has become widely accepted as a model for human disease. Small molecule screens have been performed to identify modifiers of vascular patterning but this process has been performed at relatively low throughput since the tools for fully automated processing are not yet available. We propose to develop a novel, high-throughput screening assay for angiogenesis promoters. The assay would be based on the use of the SideView" microplate to collect clear images of the vascular pattern in animals that express GFP in their vasculature. During Phase I we: 1) developed novel methods to modulate angiogenesis promoters in the plexus; 2) developed software that locates the zebrafish, identifies the region of interest, and moves that region to the center of the field of view; and 3) developed automated, post-acquisition image processing software that distinguishes wild-type zebrafish embryos from xxx mutant embryos with deformities in the plexus. During Phase II the methods, software and instrumentation will be fully integrated and the assay will be validated in a model screen using the Prestwick Library of FDA- approved compounds. PUBLIC HEALTH RELEVANCE: The Phase I and Phase II programs will develop a fully automated tool for discovery of potential therapeutic compounds that affect developing vasculature, which is relevant to embryonic diseases such as vascular anomalies. In addition, compounds that promote or inhibit growth of new blood vessels as a treatment for numerous diseases including cancer and wet macular degeneration will also be discovered. The assay will be performed in zebrafish larvae, which will result in a higher probability that identified compounds, will proceed through the drug development process.