Automation of Assay Endpoints for Brain Slice Models of Neurodegenerative Disease

描述(由申请人提供)：对于神经和神经精神疾病的新药和药物靶点的发现，从基于细胞的分析的有效性过渡到在全动物模型中受益一直是困难和不确定的。理想情况下，药物发现研究将尽可能多地在疾病的全动物模型中进行，但体内动物实验非常昂贵和耗时。相反，尽管基于细胞系和原代培养的分析是快速和廉价的，但当神经元被培养和/或永生化时，它们会受到表型变化的影响，更重要的是，由于正常的三维环境和脑组织结构的局部细胞间相互作用不可避免地丢失。为了帮助弥合基于细胞的疗效研究和整个动物疗效研究之间的差距，我们开发了一系列基于完整脑组织的中枢神经系统疾病模型，包括中风、亨廷顿病(HD)和阿尔茨海默病(AD)。在我们已发表的研究中，我们已经展示了这种分析在推进一系列基因靶标识别和药物开发计划方面的效用。为了在更大规模的发现工作中支持这些脑片外植体模型的使用，我们在过去十年中开发了许多技术和工艺创新，包括高通量脑片和用于将疾病相关基因导入脑片的生物基因枪设备。 和化验报告构建。本提案的首要目标是解决该方法完全可扩展的最终速度限制障碍，即基于脑切片的分析终点的自动化。到目前为止，我们开发的所有大脑切片疾病模型都是使用繁琐的手动终点分析进行分析的；然而，即使是一个中等规模的科学团队，分析吞吐量也足以支持每年筛选数百到数千个化合物或基因靶标。“交钥匙”、无偏见的自动化显微镜和高含量分析(HCA)平台的实施将使这些分析的吞吐量增加10倍或更多，并使大规模系统生物学、生物信息学和药物发现计划能够得到全面支持。这种在细胞模型和全动物模型之间进行高通量生物学筛选的桥梁阶段应该会显著增加药物和药物靶点发现和开发计划的成功可能性，在充分的时间和财政承诺进行全面的体内疗效研究之前，提供完整神经组织分析中的有效性和潜在的不良靶外影响的预览。

DESCRIPTION (provided by applicant): For new drug and drug target discovery in neurological and neuropsychiatric disorders, transitioning from efficacy in cell-based assays to benefit in whole-animal models has always been difficult and uncertain. Ideally, drug discovery studies would be conducted, as much as possible, in whole-animal models of disease, but in vivo animal experiments are tremendously costly and time- consuming. Conversely, while cell line and primary culture-based assays are rapid and inexpensive, they are compromised by phenotypic changes when neurons are cultured and/or immortalized, and, importantly, by the normal 3-dimensional milieu and local inter-cellular interactions of brain tissue architecture being unavoidably lost. To help bridge this gap between cell-based and whole-animal efficacy studies, we have developed a series of intact brain tissue-based models for CNS disorders including stroke, Huntington's disease (HD), and Alzheimer's disease (AD). In our published studies, we have shown the utility of such assays in advancing a range of gene target identification and drug development programs. To support the use of these brain slice explant models in the context of larger-scale discovery efforts, we have developed numerous technological and process innovations over the last decade, including high-throughput brain slicing and biolistic gene gun devices for transfection of brain slices with disease-relevant genes and assay reporter constructs. The overarching goal of the present proposal is to solve the final rate-limiting barrier to full scalability of this approach, namely, the automation of brain slice-based assay endpoints. To date, all of the brain slice disease models we have developed have been analyzed using laborious manual endpoint assays; nevertheless, assay throughput has been sufficient to support the screening of hundreds to thousands of compounds or gene targets per year even with a modest-sized scientific team. Implementation of "turnkey" unbiased, automated microscopy and high-content analysis (HCA) platforms would increase the throughput of these assays by 10-fold or more, and enable full support of large-scale systems biology, bioinformatics, and drug discovery programs. Such a bridging stage of "high-throughput biology" screening between cell-based and whole-animal models should significantly increase the likelihood of success of drug and drug target discovery and development programs, by providing a preview of both efficacy as well as potential adverse off-target effects in intact neurl tissue assays before substantial time and financial commitments are made to full in vivo efficacy studies.