A High Throughput Screen for Inhibitors of Aggregation of the Alzheimer's Peptide

阿尔茨海默病肽聚集抑制剂的高通量筛选

• 批准号: 7256597
• 负责人: MICHAEL H HECHT
• 金额: $ 20.15万
• 依托单位: PRINCETON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-04-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7256597
• 关键词: Alzheimer' s Disease; Amyloid; Binding; Biochemical; Biochemical Genetics; Biological Assay; Brain; Cell-Free System; Cells; Chemicals; Chimeric Proteins; Coupled; Data; Development; Electron Microscopy; Environment; Etiology; Event; Fluorescence; Future; Genetic Transcription; Goals; Green Fluorescent Proteins; Human; In Vitro; Kinetics; Lead; Libraries; Measures; Methods; Molecular; Nerve Degeneration; Pathway interactions; Peptides; Pharmacologic Substance; Plasma; Play; Protein Precursors; Proteins; Proteolysis; Range; Rate; Role; Screening procedure; Senile Plaques; Series; Signal Transduction; Solubility; Structure; Structure-Activity Relationship; System; Testing; Therapeutic; Thinking; Thioflavin T; Tissues; Toxic effect; Translations; base; cytotoxicity; extracellular; high throughput screening; in vivo; inhibitor/antagonist; novel; novel strategies; peptide A; prevent; protein folding; rapid technique; small molecule libraries

DESCRIPTION (provided by applicant): A range of biochemical and genetic studies indicate that aggregation of the Alzheimer's peptide, A¿, plays a crucial role in the molecular etiology of Alzheimer's disease (AD). In particular, early steps of misfolding and oligomerization are thought to produce toxic species, which initiate a cascade of events ultimately leading to neurodegeneration. Inhibition of A¿ misfolding and oligomerization may provide a therapeutic strategy for the treatment of AD; however, finding compounds that specifically accomplish this goal without interfering with the folding of other proteins has remained a major challenge. To overcome this challenge, we propose to develop a rapid, reliable, and high throughput screen for specific inhibitors of A¿ misfolding and aggregation. This goal will be achieved thorough the following specific aims: (1) To construct and optimize a novel screen for inhibitors of A¿ aggregation. The proposed screen will use a fusion of A¿42 to Green Fluorescent Protein (GFP). When fused to GFP, AB42 causes the entire fusion protein to misfold and aggregate, thereby preventing the fluorescence of the GFP portion of the protein. Compounds that inhibit A¿ aggregation will enable GFP to fold into its native structure, and will be identified by the resulting fluorescent signal. (2) To implement the screen and test its ability to distinguish inhibitors of aggregation from inactive compounds. This will be demonstrated using fluorescence to screen a pilot library of compounds. (3) To validate the screen using biochemical and biophysical methods. These studies will verify that compounds isolated by the A¿42-GFP screen enhance solubility and inhibit aggregation of pure A¿42. (4) To probe structure-activity relationships (SAR). Compounds isolated by the screen will be compared to related compounds in the libraries. These studies will elucidate the chemical features responsible for inhibition and will facilitate future lead optimization. (5) To adapt the screen for use in either cell-based or cell-free expression systems. Completion of these aims will produce a rapid method of screening for novel and specific inhibitors of aggregation, thereby identifying lead compounds for the development of new pharmaceuticals that prevent or delay the onset of Alzheimer's disease.

描述（由申请人提供）：一系列生物化学和遗传学研究表明，阿尔茨海默氏肽A？的聚集在阿尔茨海默氏病（AD）的分子病因学中起着至关重要的作用。特别是，错误折叠和寡聚化的早期步骤被认为会产生有毒物质，这会引发一系列事件，最终导致神经变性。抑制A？错误折叠和寡聚化可以提供治疗AD的治疗策略;然而，找到特异性实现该目标而不干扰其他蛋白质折叠的化合物仍然是一个主要挑战。为了克服这一挑战，我们建议开发一种快速，可靠，高通量的筛选特异性抑制剂A？错误折叠和聚集。这一目标将通过以下具体目标实现： (1)构建并优化A ²聚集抑制剂的新型筛选方法。拟议的筛选将使用A42与绿色荧光蛋白（GFP）的融合。当与GFP融合时，AB 42导致整个融合蛋白错误折叠和聚集，从而阻止蛋白质的GFP部分的荧光。抑制A？聚集的化合物将使GFP折叠成其天然结构，并将通过产生的荧光信号进行鉴定。 (2)实施筛选并测试其区分聚集抑制剂和非活性化合物的能力。这将使用荧光来筛选化合物的试验文库来证明。 (3)采用生物化学和生物物理学方法验证筛选结果。这些研究将验证通过A 42-GFP筛选分离的化合物增强溶解度并抑制纯A 42的聚集。 (4)探讨构效关系（SAR）。将通过筛选分离的化合物与文库中的相关化合物进行比较。这些研究将阐明负责抑制的化学特征，并将促进未来的铅优化。 (5)使筛选适用于基于细胞或无细胞的表达系统。这些目标的完成将产生一种快速筛选新的和特异性的聚集抑制剂的方法，从而确定用于开发预防或延迟阿尔茨海默病发作的新药的先导化合物。