Single molecule microarrays for the detection of mutant DNA in body fluids

描述（由申请人提供）：恶性肿瘤的形成与增强细胞增殖的基因突变的积累有关。检测体液中的这些突变可用于癌症管理和早期癌症检测。然而，目前的癌症诊断方法并不适合检测体液中的突变DNA，这阻碍了有效、经济的癌症筛查策略的发展。在此，我们提出了基于Twist-Biosensor技术的微阵列技术的发展，以显着改善和简化体液中生物标志物的检测，用于癌症筛查。Twist-Biosensor是一种新型的微阵列技术，它以单分子分辨率检测杂交。此外，Twist-Biosensor将破坏扭转应力应用于DNA杂交体，这是一种独特设计的新策略，可为微阵列提供高点突变选择性。因此，该技术是在存在大量野生型DNA的情况下检测低浓度突变DNA的理想技术。本应用程序的总体目标是展示twist -生物传感器微阵列用于癌症筛查的能力。我们将开发检测KRAS基因突变的方法，并用胰腺癌患者的血浆和尿液样本进行检测。本应用程序的每个目标都集中在建立微阵列的两个互补特性：检测多种癌症突变（目标1）和超灵敏检测突变DNA（目标2）。此外，这两个目标都将证明twist -生物传感器微阵列比其他诊断技术更快，所需步骤更少。

DESCRIPTION (provided by applicant): The formation of malignant tumors is associated with accumulation of genetic mutations that enhance cell proliferation. Detecting these mutations in body fluids can be used for cancer management and early cancer detection. However, current cancer diagnostic methods are not well suited for the detection of mutant DNA in body fluids which have prevented the development of effective, cost-efficient cancer screening strategies. Here, we propose the development of microarrays based on Twist-Biosensor technology to dramatically improve and simplify detection of biomarkers in body fluids for cancer screening. Twist-Biosensor is a novel microarray technique in which hybridization is detected with single molecule resolution. In addition, Twist-Biosensor applies disrupting torsional stress to DNA hybrids, a novel strategy uniquely designed to give microarrays high point mutation selectivity. Therefore, the technique is ideal for detection of mutated DNA at low concentration in the presence of a large proportion of wild-type DNA. The overall aim of this application is to demonstrate the capabilities of Twist-Biosensor microarrays for cancer screening. We will develop assays to detect mutations of the KRAS gene and test them with plasma and urine samples from pancreatic cancer patients. Each of the proposed aims of this application focuses on establishing two complementary properties of the microarrays: Detection of multiple cancer mutations (Aim 1) and ultra-sensitive detection of mutated DNA (Aim 2). In addition, both aims will demonstrate that Twist-Biosensor microarrays are faster and require fewer steps than alternative diagnostic techniques.