Live-Cell Assays for Lysosomal Enzyme Activity

描述（由申请人提供）：该小型企业创新研究II期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靶向荧光底物。如果成功，拟议的研究将提供所需的突破，以推动发现有前途的新疗法和调节药物，用于溶酶体贮积病和相关的医学应用。在该项目的第一阶段，Marker Gene Technologies，Inc.通过制备用于溶酶体酶的新的荧光糖苷酶、酯酶、磷酸酶、脂肪酶和硫酸酯酶底物，确立了该技术的可行性，并在来自正常或疾病来源的活细胞中或在诱导抑制溶酶体酶活性时证明了差异染色。在II期，将在体外测定这些和其他新底物对活细胞中溶酶体酶的特异性和局部抑制或诱导以及以细胞或组织特异性方式区分单个酶活性的能力。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，并用于监测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也将提供创新的方法来定量溶酶体酶的功能，并筛选二次药物或蛋白质给药的影响，使其成为各种重要的生物化学和医学应用的有用的医学研究工具。该公司已经与几个著名的研究实验室和机构以及该竞技场的主要制药公司合作，他们渴望在现有的临床应用中测试这些方法和系统。由此产生的分析和产品将销售给研究，制药，生物技术和诊断行业。 公共卫生相关性：该项目的成功通过发现用于调节这些疾病和其他相关疾病中的溶酶体酶活性的新蛋白质和药物，在溶酶体贮积症如桑德霍夫病、泰-萨克斯综合征、克拉比病和戈谢病的医学干预领域中开辟了巨大的商业可能性。这些检测还将使临床医生能够确定当前和新兴疗法的疗效，并为患者评估增加一套新的临床诊断检测。此外，它将导致这些领域的商业和许可产品。

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. PUBLIC HEALTH RELEVANCE: The success of this project opens up enormous commercial possibilities in the fields of medical intervention in lysosomal storage disorders such as Sandhoff's disease, Tay-Sachs syndrome, Krabbi's disease and Gaucher's disease through discovery of new proteins and drugs for use in modulating lysosomal enzyme activity in these and other related diseases. These assays will also enable clinicians to determine the efficacy of current and emerging therapies as well as add a new set of clinical diagnostic assays for patient assessment. In addition, it will lead to commercial and licensable products in these areas.