MMPA: A Novel Multiplexing Methylation Analysis Technology

描述（由申请人提供）：该快速SBIR项目旨在开发一种新型的多重甲基化分析（MMPA）技术，用于临床标本DNA的甲基化分析。具体来说，我们将开发MMPA检测（每种癌症一种），分别用于分析结直肠癌、肺癌和乳腺癌（三种最致命的癌症）中的DNA甲基化。每种检测方法可以同时测定10，000倍过量的未甲基化DNA中20个基因的甲基化状态。该检测方法还可以测定临床标本中甲基化程度和甲基化基因的相对丰度。此外，这些分析将具有成本效益，易于操作。第一阶段的目标是确定MMPA测定是否可以准确地揭示来自临床样品的DNA中的甲基化。在实验上，我们将使用我们现有的MMPA检测来分析来自粪便（最复杂的标本）的DNA中8个基因的甲基化状态。具体的里程碑是证明该检测试剂盒可分别达到90%或更高的检测灵敏度和特异性。第二阶段是开发MMPA检测方法，分别用于分析结直肠癌、肺癌和乳腺癌中的DNA甲基化。每个MMPA检测试剂盒“开一枪”以“杀死四只鸟”。首先，该测定法可以确定单个基因的甲基化状态;其次，该测定法描绘了许多基因中的DNA甲基化;第三，该测定法具有确定甲基化基因的甲基化程度的能力;以及第四，该测定法提供了对临床标本中甲基化基因丰度的了解。显然，这些独特的功能将使MMPA成为甲基化分析的首选方法，因此该项目的成功将对癌症筛查和基础癌症研究产生深远的影响。

DESCRIPTION (provided by applicant): This fast-track SBIR project is to develop a novel multiplexed methylation profiling assay (MMPA) technology for methylation analysis of DNA derived from clinical specimens. Specifically, we will develop the MMPA assays (one for each type of cancer) for analysis of DNA methylation in colorectal, lung, and breast cancer, the three deadliest cancers, respectively. Each assay can simultaneously determine the methylation status of 20 genes in a 10,000 fold excess of unmethylated DNA. The assays can also determine the degree of methylation and the relative abundance of methylated genes in clinic specimens. In addition, the assays will be cost-effective and easy to operate. The goal of Phase I is to determine if the MMPA assay can accurately reveal methylation in DNA derived from clinic samples. Experimentally, we will use our existing MMPA assay to profile the methylation status of eight genes in DNA derived from stools, the most complex specimen. The specific milestone is to demonstrate that this assay can achieve the detection sensitivity and specificity of 90% or better, respectively. Phase II is to develop the MMPA assays for analysis of DNA methylation in colorectal, lung, and breast cancers, respectively. Each MMPA assay "fires one shot" to "kill four birds". First, the assay can determine the methylation status of an individual gene; second, the assay profiles DNA methylation among a number of genes; third, the assay has the capability of determining the degree of methylation of methylated genes; and fourth, the assay provides insights into the abundance of methylated genes in clinical specimens. Clearly, these unique features will make MMPA the method of choice for methylation analysis, and thus the success of this project will have a profound impact on both cancer screening and basic cancer research.