Live-Cell Assays of ER and Golgi Enzyme Activities

ER 和高尔基体酶活性的活细胞测定

• 批准号: 8591789
• 负责人: JOHN J NALEWAY
• 金额: $ 14.98万
• 依托单位: MARKER GENE TECHNOLOGIES
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8591789
• 关键词: Ally; Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Area; Ataxia; Biological; Biological Assay; Biological Monitoring; Biotechnology; Cell Culture System; Cell Culture Techniques; Cells; Centrifugation; Chemicals; Coculture Techniques; Collaborations; Creutzfeldt-Jakob Syndrome; Cytolysis; Data; Detection; Development; Diagnosis; Disease; Endoplasmic Reticulum; Enzyme Induction; Enzyme-Linked Immunosorbent Assay; Enzymes; Evaluation; Exhibits; Fluorescence Microscopy; Fluorescence Spectroscopy; Fluorogenic Substrate; Genes; Glioblastoma; Glycoside Hydrolases; Goals; Golgi Apparatus; High Pressure Liquid Chromatography; Huntington Disease; In Vitro; Individual; Intervention; Laboratories; Lead; Life; Mannosidase; Measurement; Measures; Medical; Medicine; Methodology; Methods; Modeling; Multiple System Atrophy; Myeloid Leukemia; NMR Spectroscopy; Neurodegenerative Disorders; Non-Insulin-Dependent Diabetes Mellitus; Normal Cell; Oculocerebrorenal Syndrome; Organelles; Parkinson Disease; Peptide Hydrolases; Peptides; Permeability; Pharmaceutical Preparations; Pharmacology; Phase; Phospholipids; Phosphoric Monoester Hydrolases; Proteins; Protocols documentation; Radiolabeled; Reproducibility; Research; Running; Sampling; Side; Small Business Innovation Research Grant; Staining method; Stains; System; Techniques; Technology; Testing; Therapeutic; Thin Layer Chromatography; Time; Tissues; analytical tool; base; chemical standard; drug discovery; drug efficacy; efficacy testing; enzyme activity; enzyme substrate; fluorophore; in vitro Assay; in vitro activity; in vivo; inhibitor/antagonist; innovation; meetings; novel; public health relevance; radiotracer; research study; screening; secretase; success; tissue culture; uptake

DESCRIPTION: This Small Business Innovation Research Phase I project aims to develop new targeted fluorogenic substrates capable of measuring enzyme activities in the Golgi apparatus and Endoplasmic Reticulum (ER) of living cells and tissues. If successful, the proposed research will provide breakthroughs needed to advance the discovery of promising new therapies and modulating drugs for neurodegenerative disorders including Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS), Parkinson's disease (PD), Type 2 diabetes, Lowe syndrome, Huntington's disease and allied medical conditions. In Phase I of this project, Marker Gene Technologies, Inc. will establish the feasibility of the technology by preparing new fluorogenic glycosidase, phosphatase and peptidase substrates for enzymes with localized activity in the Golgi and ER and demonstrating differential staining in living cells that are from normal or are of disease origin. In Phase II, these and additional new substrates will be assayed in vitro and in vivo for their ability to measure specific and localized inhibition or induction of these enzymes in living cells as well as differentiate individual enzyme activities in a cell- or tissue-specific manner. The new substrates and the resulting detection systems will provide innovative methods to quantitate Golgi and ER organelle enzyme function and to screen for the influence of secondary drug or protein administration, making them useful analytical tools for drug discovery and diagnosis in a variety of significant medical applications.

产品说明：该小型企业创新研究I期项目旨在开发新的靶向荧光底物，能够测量活细胞和组织的高尔基体和内质网（ER）中的酶活性。如果成功，拟议的研究将提供突破，以推动发现有前途的新疗法和调节神经退行性疾病的药物，包括阿尔茨海默病（AD），肌萎缩侧索硬化症（ALS），帕金森病（PD），2型糖尿病，Lowe综合征，亨廷顿病和相关的医疗条件。在该项目的第一阶段，Marker Gene Technologies，Inc.将通过制备新的荧光糖苷酶、磷酸酶和肽酶底物来确定该技术的可行性，所述底物用于在高尔基体和ER中具有局部活性的酶，并证明来自正常或疾病来源的活细胞中的差异染色。在第II阶段，将在体外和体内测定这些和其他新底物测量特异性和局部抑制或诱导的能力， 活细胞中的这些酶以及以细胞或组织特异性方式区分单个酶活性。新的底物和由此产生的检测系统将提供创新的方法来定量高尔基体和ER细胞器酶的功能，并筛选二次药物或蛋白质给药的影响，使其成为药物发现和诊断的有用分析工具，在各种重要的医学应用。