Autoantibody- and microRNA-based Next-Gen Multiplex Test for Type 1 Diabetes

基于自身抗体和 microRNA 的下一代 1 型糖尿病多重检测

• 批准号: 8981928
• 负责人: Taiho Kim
• 金额: $ 15万
• 依托单位: NESHER TECHNOLOGIES, INC.
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-15 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8981928
• 关键词: Affinity; Antibodies; Antigens; Archives; Area; Autoantibodies; Autoimmune Diseases; Automation; Beta Cell; Binding; Biological Assay; Biological Markers; Biophysics; Blinded; Body Fluids; Clinical; Clinical Sensitivity; Color; Communities; Complex; Conduct Clinical Trials; DNA; Detection; Development; Diabetes Mellitus; Diagnosis; Diagnostic; Diagnostic tests; Disease Management; Dyes; Early Diagnosis; Early Intervention; Energy Transfer; Enzyme-Linked Immunosorbent Assay; Epidemic; Evaluation; Evaluation Studies; Exclusion; Fluorescence; Fluorescence Spectroscopy; Fluorescent Dyes; Future; Geometry; Goals; Head; Health Care Costs; Human; Hyperglycemia; Individual; Insulin; Insulin-Dependent Diabetes Mellitus; Intellectual Property; Interferon Type II; Kinetics; Label; Laboratories; Lasers; Legal patent; Licensing; Life Expectancy; Measures; Medical; Methodology; Methods; MicroRNAs; Microfluidics; Miniaturization; Monitor; Morbidity - disease rate; Noise; Nucleic Acids; Outcome; Pancreas; Pathogenesis; Patient Monitoring; Patients; Phase; Plasma; Procedures; Property; Proteins; Quality of life; Reagent; Research; Research Design; Risk; Sample Size; Sampling; Savings; Sensitivity and Specificity; Serological; Site; Small Business Innovation Research Grant; Societies; Solutions; Sorting - Cell Movement; Spectrum Analysis; Stratification; Structure of beta Cell of islet; System; Technical Expertise; Technology; Testing; Therapeutic; Time; Translating; Tumor Markers; Tumor Necrosis Factor-alpha; Validation; Work; adiponectin; base; circulating microRNA; clinical assay development; cost; cytokine; diagnostic panel; endocrine pancreas development; fluorophore; improved; innovation; insulin dependent diabetes mellitus onset; interest; islet; mortality; nanobiotechnology; nanolitre; next generation; outcome forecast; prognostic; prospective; prototype; public health relevance; research clinical testing; sample collection; single molecule; tool; user-friendly; virtual

 DESCRIPTION (provided by applicant): Diabetes is a major cause of morbidity and mortality in the U.S. Given that the diabetes epidemic continues to grow worldwide, there is a clear need for improvements in the management of the disease and its complications. Early diagnosis and intervention can significantly improve long-term prognosis. Autoantibodies against islet antigens are a serologic hallmark of patients with Type 1 Diabetes (T1D). In addition, circulating microRNAs (miRNAs) have been shown recently to be systematically altered, indicative of miRNA signatures with diagnostic utility. Growing evidence suggests that a multi-marker strategy, containing a combination of biomarkers with high clinical sensitivity and specificity, may enhance diagnostic and prognostic accuracy in the future compared to single marker tests. To support efforts of identifying the most informative biomarker panels, reliable next- generation platform technologies are needed that permit multiplexed detection of markers in small samples and are suitable for automation and integration into the clinical laboratory work flow. Nesher Technologies, Inc. (NTI) has exclusively licensed the intellectual property for an ultrasensitive and - specific biodetection technology, developed at the UCLA Single Molecule Biophysics Lab (headed by Prof. Shimon Weiss), with high single-well multiplexing potential, minimal sample requirements, and simplified handling procedures (no separation/washing and amplification steps). It is based on alternating laser excitation (ALEX) single molecule fluorescence spectroscopy, whereby target recognition molecules are tagged with different color fluorescent dyes (and quenchers). NTI recently achieved extension from 2-color to 4-color ALEX, substantially expanding its multiplexing power, and demonstrated diagnostic utility for direct protein as well as miRNA quantification. Furthermore, recent work by Profs. Steve Quake and Shimon Weiss shows i) combination of microfluidics-based sample handling with ALEX spectroscopy, termed "single molecule optofluidics", and ii) enhanced throughput using a multifoci excitation/detection geometry. NTI's long-term goal is to develop rapid, highly multiplexed, ultrasensitive and -specific, as well as fully automated, nucleic acid- and protein-based diagnostic tests that require minimal sample sizes. Here, we propose assay development and clinical validation of a next-generation test with significantly improved diagnostic, prognostic, and treatment- guiding properties, implementing a panel of autoantibody and miRNA biomarkers, and overcoming limitations of current T1D testing. Our Specific Aims are: 1. Initial reagent development for a multiplex autoantibody- and miRNA-based next-generation test for T1D 2. Separate as well as multiplexed biomarker detection and quantification using spiked samples 3. ALEX-based analysis of 200 archived clinical samples and cross-validation to ELISA and qPCR methods SBIR Phase II will propose assay expansion to include more markers, miniaturization, and development of a user-friendly, "sample in - answer out" diagnostic system offering significant cost and patient sample savings.

 描述（由申请人提供）：糖尿病是美国发病和死亡的主要原因。鉴于糖尿病流行在世界范围内持续增长，显然需要改进对该疾病及其并发症的管理。可以显着改善长期预后。针对胰岛抗原的自身抗体是 1 型糖尿病 (T1D) 患者的血清学标志。此外，循环 microRNA (miRNA) 也已被证实。最近显示被有意改变，表明 miRNA 特征具有诊断效用，越来越多的证据表明，与单一标记相比，包含具有高临床敏感性和特异性的生物标记组合的多标记策略可能会提高未来的诊断和预后准确性。为了支持识别信息最丰富的生物标记物组合的工作，需要可靠的下一代平台技术，该技术允许对小样本中的标记物进行多重检测，并且适合自动化和集成到临床实验室工作流程中。 ) 拥有独家授权超灵敏和特异性生物检测技术的知识产权，由加州大学洛杉矶分校单分子生物物理实验室（由 Shimon Weiss 教授领导）开发，具有高单孔复用潜力、最少的样品要求和简化的处理程序（无需分离/清洗）它基于交替激光激发 (ALEX) 单分子荧光光谱，其中目标识别分子用不同颜色的荧光染料（和猝灭剂）进行标记。最近实现了从 2 色到 4 色 ALEX 的扩展，大大扩展了其多重能力，并展示了直接蛋白质和 miRNA 定量的诊断实用性。此外，Steve Quake 和 Shimon Weiss 教授最近的工作表明 i) 微流体的组合。 - 基于 ALEX 光谱的样品处理，称为“单分子光流控”，以及 ii) 使用多焦点激发/检测几何结构提高通量。的目标是开发快速、高度多重、超灵敏和特异性以及全自动、基于核酸和蛋白质的诊断测试，这些测试需要最小的样本量。在这里，我们建议使用下一代测试进行检测开发和临床验证。显着改善诊断、预后和治疗指导特性，实施一系列自身抗体和 miRNA 生物标志物，并克服当前 T1D 检测的局限性： 1. 多重自身抗体和 miRNA 的初步试剂开发。基于 miRNA 的下一代 T1D 测试 2. 使用加标样品进行单独和多重生物标志物检测和定量 3. 对 200 个存档临床样品进行基于 ALEX 的分析，并对 ELISA 和 qPCR 方法进行交叉验证 SBIR II 期将提出检测扩展包括更多标记、小型化以及开发用户友好的“样本输入-答案输出”诊断系统，从而显着节省成本和患者样本。