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: 9405346
• 负责人: JOHN J NALEWAY
• 金额: $ 63.14万
• 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 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9405346
• 关键词: Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Area; Basic Science; Biochemical; Biological Assay; Biological Markers; Biological Monitoring; Biotechnology; Cell Culture System; Cells; Cellular biology; Chemicals; Clinical; Collaborations; Communities; Cytolysis; Defect; Detection; Development; Diagnostic; Diagnostic tests; Disease; Disease Progression; Dose; Drug usage; Endoplasmic Reticulum; Enzyme Induction; Enzymes; Evaluation; Flow Cytometry; Fluorescence; Fluorescence Microscopy; Fluorescence Spectroscopy; Fluorogenic Substrate; Funding; Galactosidase; Gaucher Disease; Genes; Globoid cell leukodystrophy; Glycoside Hydrolases; Glycosphingolipids; Goals; Golgi Apparatus; Huntington Disease; Image Analysis; Individual; Industrialization; Industry; Institution; Intervention; Laboratories; Laboratory Research; Lead; Legal patent; Lethal Dose 50; Libraries; Licensing; Literature; Lysosomal Storage Diseases; Mannosidase; Measurement; Measures; Medical; Metabolic; Methods; Monitor; Morphologic artifacts; Neurodegenerative Disorders; Neurologic; Non-Insulin-Dependent Diabetes Mellitus; Oculocerebrorenal Syndrome; Oregon; Organelles; Parkinson Disease; Pathway interactions; Patients; Peer Review; Peptide Hydrolases; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacology; Phase; Process; Proteins; Protocols documentation; Publications; Reporting; Reproducibility; Research; Sampling; Small Business Innovation Research Grant; Specificity; Staining method; Stains; System; Techniques; Technology; Testing; Therapeutic; Therapeutic Effect; Tissues; Treatment Efficacy; Treatment Protocols; United States National Institutes of Health; Universities; Validation; Work; base; beta-n-acetylhexosaminidase; cell immortalization; chemical standard; clinical diagnostics; commercialization; diagnostic assay; drug candidate; drug discovery; efficacy testing; endoplasmic reticulum stress; enzyme activity; enzyme substrate; fluorophore; genetic analysis; glucosidase; high throughput screening; human disease; immortalized cell; inhibitor/antagonist; innovation; meetings; multiplex detection; nervous system disorder; novel drug class; novel therapeutics; programs; protein folding; protein transport; research clinical testing; response; scale up; screening; small molecule; small molecule therapeutics; success; symposium; tool; trafficking

PROJECT SUMMARY/ABSTRACT This Small Business Innovation Research Phase II 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. The ultimate goal and the overall impact of this project is to provide an understanding of the dynamic processes that occur in intracellular enzyme and protein trafficking defects in human disease and to obtain information that will provide a pathway to more efficacious treatment options. If successful, the proposed research will provide breakthroughs needed to advance the discovery of promising new therapies and modulating drugs for neurodegenerative disorders including Gaucher disease and other lysosomal storage diseases, Parkinson's disease (PD), Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS), Type 2 diabetes, Lowe syndrome, Huntington's disease and allied medical conditions. In Phase I of this project, Marker Gene Technologies, Inc. established the feasibility of the technology by preparing new fluorogenic glycosidase 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 evaluated for their ability to measure specific and localized inhibition or induction of enzyme activities inside individual organelles in living cells as well as to elucidate the trafficking machinery that has a functional relationship to disease progression. The new substrates and the resulting detection systems will provide innovative methods to quantitate the influence of secondary drug or protein administration on organelle-specific lysosomal, Golgi or ER enzyme activities. Furthermore, their use in screening libraries of potential drug candidates for their ability to modulate enzyme function and localization will identify new small molecule therapeutic leads based on these parameters. This makes the combined systems useful as basic research tools for a variety of significant cell biology, biochemical and medical applications. The company has engaged the collaboration of several noted academic research laboratories and research institutions as well as major pharmaceutical companies in this arena, who are eager to test the methods and systems in their existing clinical, diagnostic or drug discovery applications. The resulting assays and products will be marketed to the research, pharmaceutical, biotechnology and diagnostic industries.

项目总结/文摘