An automated high throughput phenotypic screen for schistosomiasis drug discovery

描述（申请人提供）：血吸虫病是一种热带寄生虫病，感染人口超过2亿。治疗依赖于单一药物吡喹酮（PZQ）。在缺乏具有PZQ治疗谱的备用药物的情况下，PZQ耐药和最终药物失败的风险是一个主要问题。传统的表型筛选，使用成虫期曼氏梭菌，是低通量和不兼容的现代高通量筛选（HTS）系统。为了与NIAID的使命保持一致，本提案旨在将新开发的中等通量表型筛选（MTS）转变为全自动、定量的HTS，以加速这种传染病的药物发现。该提案涉及三个正在进行合作的pi，以及各自的生物，筛选技术和生物计算技能，他们专注于这一目标。作为该提案的第一个研究方向，我们将利用内部自动化和高含量筛选（HCS）系统来显著提高我们已发布的MTS方法的吞吐量。该提案将涉及；扩大寄生虫的机器人电镀，开发基于亮场和荧光的显微镜协议，并采用商业图像分析软件来识别（片段），定量描述和跟踪寄生虫的运动，以优先考虑化合物进行进一步的临床前开发。由于商业HCS分析工具不太可能优化记录这种多细胞寄生虫所显示的复杂和动态表型，我们也将追求第二和平行的研究轨道。具体来说，我们将开发一种自动图像分析筛选技术来定义、识别和量化这种寄生虫可能的表型反应（形态和行为）范围。最终，实验和计算轨道将共同产生一个标准化的HTS协议和一个全面的、定量的图像分析程序套件，用于对寄生虫表型进行分类。这种严谨性将有助于筛选大量潜在化合物，并将其优先纳入内部可用的二级和三级筛选分析。我们还打算公开算法框架，包括其方法和实现。

DESCRIPTION (provided by applicant): Schistosomiasis is a tropical parasitic disease infecting over 200 million people. Treatment relies on a single drug, praziquantel (PZQ). In the absence of back-up drugs with PZQ's therapeutic spectrum, the risk of resistance to PZQ and eventual drug failure is a major concern. Traditional phenotypic screens, using adult- stage S. mansoni, are low-throughput and incompatible with modern high-throughput screen (HTS) systems. In keeping with the NIAID's mission, the present proposal aims to turn a newly developed, moderate- throughput phenotypic screen (MTS), into a fully automated, quantitative HTS to accelerate drug discovery for this infectious disease. The proposal involves three PIs with ongoing collaborations and respective biological, screening-technology and bio-computational skills who are focused on just this goal. As a first research track for this proposal, we will utilize in-house automation and a high-content screening (HCS) system to significantly increase throughput over our published MTS approach. The proposal will involve; expanding robotic plating of the parasite, developing protocols for bright-field and fluorescence-based microscopy and adapting commercial image-analysis software to identify (segment), quantitatively describe and track the motion of parasites with a view to prioritizing compounds for further pre-clinical development. Because commercial HCS analysis tools are not likely optimized for recording the complex and dynamic phenotypes displayed by this multicellular parasite, we will also pursue a second and parallel track of research. Specifically, we will develop de novo an automated image-analysis screening technology to define, identify, and quantify the range of phenotypic responses (morphological and behavioral) possible in this parasite. Ultimately, both the experimental and computation tracks will together produce a standardized HTS protocol and a comprehensive, quantitative suite of image-analysis programs to categorize parasite phenotypes. Such rigor will facilitate the screening of large numbers of potential compounds and their prioritization into the secondary and tertiary screening assays available in-house. We also intend to make the algorithmic framework including its methods and implementations, publicly available. PUBLIC HEALTH RELEVANCE: The major goal of the project is to turn a moderate-throughput phenotypic screen (MTS) system for schistosomiasis, into a fully automated, quantitative high-throughput screen (HTS). By so doing, the rate of discovery of drugs to treat this global tropical disease will be increased.