A Novel Method for Efficiently and Robustly Retrieving Circulating miRNAs

一种高效、稳健地检索循环 miRNA 的新方法

• 批准号: 10013246
• 负责人: Qipan Deng
• 金额: $ 75万
• 依托单位: GLC BIOTECHNOLOGY, INC.
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-09 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10013246
• 关键词: Adsorption; Biological Assay; Biological Markers; Biological Models; Blood; Cardiovascular Diseases; Clinical; DNA; Detection; Development; Diagnosis; Disease; FDA approved; Foundations; Health; Human; Individual; Interruption; Malignant Neoplasms; Methods; MicroRNAs; Monitor; Nucleic Acids; Phase; Plasma; Protocols documentation; RNA; Sampling; Serum; Small Business Innovation Research Grant; Specimen; Surface; Technology; Testing; base; cancer biomarkers; circulating microRNA; clinical application; colorectal cancer screening; commercialization; cost; cost effective; disease diagnosis; improved; microRNA biomarkers; new technology; novel; success

Abstract This direct SBIR Phase II project aims to develop a novel technology and associated assays for efficiently and robustly extracting circulating miRNAs from blood (i.e. plasma or serum), which could transform the utility of miRNA testing in the diagnosis and monitoring of major diseases such as cancer and cardiovascular disease. Our competitive advantage lies in the ability to retrieve circulating miRNAs more efficiently and robustly than current extraction methods, which has been a critical barrier in implementing miRNA testing clinically. This advantage is achieved by implementing sequence specific capture in conjunction with a propriety method of making novel capture beads. Circulating miRNAs are potential disease biomarkers. However, to date, there are no FDA approved miRNA tests available. A bottleneck problem is unreliable circulating miRNA extraction. Current extraction methods are based on adsorption of polar molecules on polar surfaces, which were originally developed to extract large DNA and RNA molecules. However, because miRNAs are so small (~22nt), their interaction with polar surfaces is much weaker, therefore their adsorption on polar surfaces can be easily interrupted by other molecules present in sample. Since the weak adsorption of miRNAs on polar surfaces is an inherent problem that cannot be fully solved even if all other conditions were optimized, new methods for retrieving circulating miRNAs based on a different mechanism are clearly needed. Sequence-specific capture (SSC) is another method to extract nucleic acids, but historically it performs poorly when used to extract circulating nucleic acids from clinical samples due to the lack of effective capture beads. In addition, the cost of making capture beads by current methods is very high, making SSC too expensive for clinical use. Recently, we made a major breakthrough by developing a proprietary method of making capture beads that can transform SSC from a concept to a practical method for extracting circulating miRNAs in an efficient, robust, and cost-effective manner. Therefore, we employed our capture beads to develop our own SSC assays, and conducted a systematic study to examine the feasibility of using our SSC assay in extracting circulating miRNAs. Our SSC assay was indeed found to efficiently and robustly extract circulating miRNAs. In the study, we also found that extraction efficiency of current kits varied by as much as 60-fold by plasma sample, further confirming that current methods are not robust. Considering the problem of current methods, unparalleled features of our SSC technology, and success of our early study, we propose this Phase II SBIR project to further develop and validate our SSC technology for extracting circulating miRNAs from plasma/serum, providing a foundation for its commercialization.

抽象的 这个直接的 SBIR 第二阶段项目旨在开发一种新技术和相关检测方法，以实现高效和 从血液（即血浆或血清）中强有力地提取循环 miRNA，这可能会改变 miRNA检测在癌症、心血管疾病等重大疾病的诊断和监测中的应用。 我们的竞争优势在于能够更有效、更稳健地检索循环 miRNA 目前的提取方法一直是临床实施 miRNA 检测的关键障碍。这 通过结合适当的方法实施序列特异性捕获来实现优势 制作新颖的捕获珠。 循环 miRNA 是潜在的疾病生物标志物。然而，迄今为止，还没有 FDA 批准的 miRNA 可用的测试。一个瓶颈问题是不可靠的循环 miRNA 提取。目前的提取方法 基于极性表面上极性分子的吸附，最初开发用于提取大分子 DNA 和 RNA 分子。然而，由于 miRNA 非常小（~22nt），它们与极性的相互作用 表面的吸附力要弱得多，因此它们在极性表面上的吸附很容易被其他物质干扰 样品中存在的分子。由于 miRNA 在极性表面上的弱吸附是一个固有的问题 即使优化所有其他条件也无法完全解决这个问题，回收循环的新方法 显然需要基于不同机制的 miRNA。序列特异性捕获 (SSC) 是另一种方法 提取核酸的方法，但历史上用于提取循环核酸时效果不佳 由于缺乏有效的捕获珠子而导致临床样品中的酸。此外，捕获的成本 目前方法的珠子非常高，使得 SSC 对于临床使用来说过于昂贵。 最近，我们通过开发一种制造捕获珠的专有方法取得了重大突破， 可以将 SSC 从一个概念转变为一种实用的方法，用于以高效、稳健、 和成本效益的方式。因此，我们使用捕获珠来开发我们自己的 SSC 检测方法，并且 进行了一项系统研究，以检验使用我们的 SSC 测定法提取循环中的可行性 miRNA。我们的 SSC 测定确实可以有效、稳健地提取循环 miRNA。在研究中， 我们还发现，当前试剂盒的提取效率因血浆样本的不同而变化多达 60 倍，进一步 确认当前方法并不稳健。 考虑到当前方法的问题、我们的SSC技术无与伦比的特点以及我们的成功 在早期研究中，我们提出了第二阶段 SBIR 项目，以进一步开发和验证我们的 SSC 技术 从血浆/血清中提取循环miRNA，为其商业化奠定基础。