microRNA- and Autoantibody-based Multiplex Assay for Systemic Lupus Erythematosus

基于 microRNA 和自身抗体的系统性红斑狼疮多重检测

• 批准号: 8979824
• 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: 2015-08-01 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8979824
• 关键词: Acute Disease; Affect; Affinity; Antibodies; Antinuclear Antibodies; Antiphospholipid Antibodies; Archives; Area; Autoantibodies; Autoimmune Diseases; Automation; Binding; Biological Assay; Biological Markers; Biophysics; Blinded; Body Fluids; Clinical; Clinical Sensitivity; Color; Communities; Complex; Conduct Clinical Trials; DNA; Detection; Development; Diagnosis; Diagnostic; Diagnostic tests; Disease remission; Dyes; Early Diagnosis; Energy Transfer; Enzyme-Linked Immunosorbent Assay; Evaluation; Evaluation Studies; Exclusion; Flare; Fluorescence; Fluorescence Spectroscopy; Fluorescent Dyes; Future; Geometry; Goals; Head; Health Care Costs; Human; Immune; Individual; Intellectual Property; Kidney Diseases; Kinetics; Label; Laboratories; Lasers; Legal patent; Licensing; Longevity; Lupus; Measures; Medical; Methodology; Methods; MicroRNAs; Microfluidics; Miniaturization; Monitor; Morbidity - disease rate; Noise; Nucleic Acids; Organ; Outcome; Pathogenesis; Patient Monitoring; Patients; Phase; Plasma; Procedures; Property; Proteins; 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; System; Systemic Lupus Erythematosus; Technical Expertise; Technology; Testing; Therapeutic immunosuppression; Time; Translating; Tumor Markers; Validation; Work; base; circulating microRNA; clinical assay development; cost; diagnostic panel; ds-DNA; fluorophore; improved; innovation; interest; mortality; nanobiotechnology; nanolitre; next generation; novel marker; novel therapeutics; outcome forecast; prognostic; prospective; prototype; public health relevance; research clinical testing; sample collection; single molecule; systemic autoimmune disease; tool; user-friendly; virtual

 DESCRIPTION (provided by applicant): Systemic lupus erythematosus (SLE) is a systemic autoimmune disease with worldwide distribution. Despite medical treatment, morbidity and mortality from renal disease are still common in lupus patients. However, early diagnosis and prompt treatment can significantly improve long-term prognosis. Anti-double stranded (ds) DNA autoantibodies are a serologic hallmark of patients with SLE. In addition, circulating microRNAs (miRNAs) have been shown recently to be systematically altered, unveiling 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 both protein and nucleic acid targets 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 multi-foci 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 SLE testing. Our Specific Aims are: 1. Initial reagent development for a multiplex miRNA & autoantibody-based next-generation test for SLE 2. Separate as well as multiplexed biomarker detection and quantification using spiked samples 3. ALEX-based analysis of 42 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）：系统性红斑狼疮（SLE）是一种全身性自身免疫性疾病，分布于世界各地。尽管药物治疗，肾脏疾病的发病率和死亡率在狼疮患者中仍然很常见。但早期诊断和及时治疗可明显改善远期预后。抗双链DNA自身抗体是SLE患者的血清学标志。此外，循环microRNA（miRNA）最近已被证明是系统性改变，揭示了miRNA的诊断功能。越来越多的证据表明，与单一标志物检测相比，包含具有高临床敏感性和特异性的生物标志物组合的多标志物策略可能会提高未来诊断和预后的准确性。为了支持鉴定信息量最大的生物标志物组的努力，需要可靠的下一代平台技术，其允许在小样品中多重检测蛋白质和核酸靶标，并且适合于自动化和整合到临床实验室工作流程中。 Nesher Technologies，Inc. (NTI)拥有UCLA单分子生物物理学实验室（由Shimon韦斯教授领导）开发的超灵敏和特异性生物检测技术的独家知识产权，该技术具有高单孔多路复用潜力，最低样品要求和简化的处理程序（无分离/洗涤和扩增步骤）。它是基于交替激光激发（ALEX）单分子荧光光谱，其中目标识别分子标记不同颜色的荧光染料（和猝灭剂）。NTI最近实现了从2色到4色ALEX的扩展，大大扩展了其多路复用能力，并证明了直接蛋白质和miRNA定量的诊断实用性。此外，最近的工作由教授。Steve Quake和Shimon韦斯展示了i）基于微流体的样品处理与ALEX光谱学的组合，称为“单分子光流体”，和ii）使用多焦点激发/检测几何结构增强的通量。NTI的长期目标是开发快速，高度多重，超灵敏和特异性，以及全自动，核酸和蛋白质为基础的诊断测试，需要最小的样本量。在这里，我们提出了下一代检测的检测开发和临床验证，其具有显著改善的诊断，预后和治疗指导特性，实施一组自身抗体和miRNA生物标志物，并克服当前SLE检测的局限性。我们的具体目标是：1。基于多重miRNA和自身抗体的下一代SLE检测试剂的初步开发2.使用加标样品进行分离以及多重生物标志物检测和定量3.基于ALEX的42份存档临床样本分析和ELISA和qPCR方法的交叉验证SBIR第二阶段将提出分析扩展，以包括更多标记物、小型化和开发用户友好的“样本输入-回答”诊断系统，从而显著节省成本和患者样本。