A Method for Quantitatively Retrieving Circulating miRNAs from Plasma

 描述(申请人提供)：这个SBIR第一阶段项目的目标是开发一种新的序列特异性捕获(SSC)技术来定量和可靠地从血液中提取循环miRNAs，该技术可以改变循环miRNAs检测在包括癌症和心血管疾病在内的疾病诊断中的作用。循环中的miRNAs已成为许多疾病的新生物标志物。然而，到目前为止，还没有可以用于临床的基于miRNA的检测方法。这是因为基于miRNA的检测是基于miRNA生物标志物的量化，而目前的检测方法不能准确地确定循环中miRNAs的水平。造成这种情况的一个主要原因是缺乏能够以稳健的方式定量检索循环中的miRNAs的方法。最常用的回收miRNAs的方法(基于硅胶的提取)是基于极性分子在硅胶上的吸附，并被开发用于提取大的DNA或RNA片段。由于miRNAs如此之小，它们与二氧化硅的相互作用比大片段与二氧化硅的相互作用弱得多。这意味着miRNA在二氧化硅上的吸附很容易被血液中的其他分子打断。显然，miRNAs在硅胶上的弱吸附是定量回收循环miRNAs所不能轻易解决的固有问题。最近，我们结合了我们发明的两项突破性技术，开发了一种新的SSC方法，并成功地将其用于从粪便和血液中定量提取DNA。不同于基于硅胶的萃取性能随着DNA或RNA尺寸的减小而恶化，SSC的回收率和稳健性实际上随着DNA或RNA尺寸的减小而增加。考虑到我们已经证明了我们的SSC方法可以定量和稳健地回收循环DNA，并且循环中的miRNAs比循环DNA短得多，我们期待我们的SSC技术也能定量和稳健地从血液中回收循环中的miRNAs。因此，我们提出了这个SBIR项目，以开发我们的SSC方法来定量提取血液中的miRNAs。显然，该项目的成功将对人类健康产生重大影响，因为它可以提供一种定量提取循环miRNAs的方法，从而改变基于循环miRNA的检测方法在疾病诊断中的应用。

 DESCRIPTION (provided by applicant): The objective of this SBIR Phase I project is to develop a novel sequence-specific capture (SSC) technology to quantitatively and robustly retrieve circulating miRNAs from blood, which can transform the utility of circulating miRNAs testing in diagnosis of diseases including cancer and cardiovascular disease. Circulating miRNAs have emerged as novel biomarkers for many diseases. To date, however, there are no miRNA-based assays that can be used clinically. This is because miRNA-based assay is based on quantification of miRNA biomarkers and current assays are unable to accurately determine levels of circulating miRNAs. One major reason for this is the lack of methods that can quantitatively retrieve circulating miRNAs in a robust manner. The most commonly used method (silica-based extraction) for retrieving miRNAs is based on adsorption of polar molecules on polar silica and was developed to extract large DNA or RNA fragments. Because miRNAs are so small, their interaction with silica is much weaker than that of large fragments with silica. Ths means that miRNA adsorption on silica can be easily interrupted by other molecules in blood. Clearly, weak adsorption of miRNAs on silica is an inherent problem that cannot be easily solved for quantitatively retrieving circulating miRNAs. Recently, we have developed a novel SSC method by incorporating two breakthrough technologies invented by us, and successfully used it to quantitatively retrieve DNA from stool and blood in a robust manner. Unlike silica-based extraction whose performance deteriorates with a decrease of the size of DNA or RNA, the recovery and robustness of SSC actually increase with a decrease of the size of DNA or RNA. Considering the fact that we have demonstrated that our SSC method could quantitatively and robustly retrieve circulating DNA and that circulating miRNAs are much shorter than circulating DNA, we expect that our SSC technology should work for quantitatively and robustly retrieving circulating miRNAs from blood as well. Therefore, we propose this SBIR project to develop our SSC method for quantitatively retrieving miRNAs from blood. Clearly, success of this project will have a great impact on human healthcare, for it can provide a method for quantitatively retrieving circulating miRNAs, hence transforming the utility of circulating miRNA- based assays in disease diagnosis.