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Live-Cell Assays for Lysosomal Enzyme Activity

溶酶体酶活性的活细胞测定

基本信息

批准号：
8620726
负责人：
JOHN J NALEWAY
金额：
$ 47.56万
依托单位：
MARKER GENE TECHNOLOGIES
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-03-01 至 2016-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8620726
关键词：
Acid Phosphatase Ally Area Biochemical Biological Assay Biological Monitoring Biotechnology Cathepsins Cell Culture Techniques Cells Clinical Collaborations Cytolysis Detection Diagnostic Diagnostic tests Disease Disease Progression Drug usage Effectiveness Enzymes Evaluation Fluorescence Fluorescence Microscopy Fluorescence Spectroscopy Fluorogenic Substrate Fucosidase Galactosidase Gaucher Disease Genes Glycoside Hydrolases Glycosphingolipids Goals In Vitro Individual Industry Institution Intervention Laboratories Laboratory Markers Laboratory Research Lead Leucine Aminopeptidase Libraries Life Lipase Lysosomal Storage Diseases Lysosomes Mannosidase Marketing Measurement Measures Medical Medical Research Methods Monitor Morphologic artifacts Neuraminidase Normal tissue morphology Organelles Output Patients Pharmaceutical Preparations Pharmacologic Substance Phase Phosphoric Monoester Hydrolases Proteins Protocols documentation Radiolabeled Reporting Reproducibility Research Sandhoff Disease Small Business Innovation Research Grant Staining method Stains Sulfatases Syndrome System Techniques Technology Testing Therapeutic Therapeutic Intervention Tissues Treatment Efficacy Treatment Protocols United States National Institutes of Health Work chemical standard chemical synthesis clinical application commercialization drug discovery drug efficacy efficacy testing enzyme activity esterase fluorophore glucosidase high throughput screening in vitro Assay inhibitor/antagonist innovation meetings novel therapeutics radiotracer research clinical testing scale up screening small molecule success therapy development tool

项目摘要

DESCRIPTION (provided by applicant): This Small Business Innovation Research Phase II project aims to develop new targeted fluorogenic substrates capable of measuring lysosomal enzyme activity in 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 lysosomal storage diseases and allied medical applications. In Phase I of this project, Marker Gene Technologies, Inc. established the feasibility of the technology by preparing new fluorogenic glycosidase, esterase, phosphatase, lipase and sulfatase substrates for lysosomal enzymes and demonstrated differential staining in living cells that were from normal or were of disease origin or upon induction of inhibition of lysosomal enzyme activities. In Phase II, these and additional new substrates will be assayed in vitro for their ability to measure specific and localized inhibition or induction of lysosomal enzymes in living cells as well as differentiate individual enzyme activities in a cell- or tissue-specific manner. These new systems will be validated for use in high-throughput screening for drug discovery, for use in clinical diagnostics to evaluate the occurrence and progression of disease, and for use in monitoring the effectiveness of existing or emerging therapeutic interventions for these disorders. The new substrates and the resulting detection systems will also provide innovative methods to quantitate lysosomal enzyme function and to screen for the influence of secondary drug or protein administration, making them useful medical research tools for a variety of significant biochemical and medical applications. The company has engaged the collaboration of several noted research laboratories and institutions as well as major pharmaceutical companies in this arena, who are eager to test the methods and systems in their existing clinical applications. The resulting assays and products will be marketed to the research, pharmaceutical, biotechnology and diagnostic industries.

描述（由申请人提供）：该小型企业创新研究第二阶段项目旨在开发能够测量活细胞和组织中溶酶体酶活性的新型靶向荧光底物。如果成功，拟议的研究将为促进溶酶体贮积病和相关医疗应用的有前景的新疗法和调节药物的发现提供所需的突破。在该项目的第一阶段，Marker Gene Technologies, Inc. 通过制备溶酶体酶的新型荧光糖苷酶、酯酶、磷酸酶、脂肪酶和硫酸酯酶底物，确定了该技术的可行性，并在正常或疾病来源的活细胞中或在诱导溶酶体酶活性抑制后证明了差异染色。在第二阶段，将在体外测试这些和其他新底物测量活细胞中溶酶体酶的特异性和局部抑制或诱导的能力，以及以细胞或组织特异性方式区分个体酶活性的能力。这些新系统将被验证用于药物发现的高通量筛选、用于临床诊断以评估疾病的发生和进展，以及用于监测针对这些疾病的现有或新兴治疗干预措施的有效性。新的底物和由此产生的检测系统还将提供定量溶酶体酶功能和筛选二次药物或蛋白质施用的影响的创新方法，使其成为各种重要生化和医学应用的有用的医学研究工具。该公司与该领域的几个著名研究实验室和机构以及主要制药公司进行了合作，他们渴望在现有的临床应用中测试该方法和系统。由此产生的检测方法和产品将销往研究、制药、生物技术和诊断行业。