Wertheim Gerald B W, Smith Catherine, Figueroa Maria E, Kalos Michael, Bagg Adam, Carroll Martin, Master Stephen R
Department of Pathology, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania; Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
Department of Pathology, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania.
J Mol Diagn. 2014 Mar;16(2):207-15. doi: 10.1016/j.jmoldx.2013.10.010. Epub 2013 Dec 25.
Aberrant regulation of DNA methylation is characteristic of cancer cells and clearly influences phenotypes of various malignancies. Despite clear correlations between DNA methylation and patient outcome, tests that directly measure multiple-locus DNA methylation are typically expensive and technically challenging. Previous studies have demonstrated that the prognosis of patients with acute myeloid leukemia can be predicted by the DNA methylation pattern of 18 loci. We have developed a novel strategy, termed microsphere HpaII tiny fragment enrichment by ligation-mediated PCR (MELP), to simultaneously analyze the DNA methylation pattern at these loci using methylation-specific DNA digestion, fluorescently labeled microspheres, and branched DNA hybridization. The method uses techniques that are inexpensive and easily performed in a molecular laboratory. MELP accurately reflects the methylation levels at each locus analyzed and segregates patients with acute myeloid leukemia into prognostic subgroups. Our results demonstrate the usefulness of MELP as a platform for simultaneous evaluation of DNA methylation of multiple loci.
DNA甲基化的异常调控是癌细胞的特征，并且明显影响各种恶性肿瘤的表型。尽管DNA甲基化与患者预后之间存在明确的相关性，但直接测量多位点DNA甲基化的检测通常成本高昂且技术上具有挑战性。先前的研究表明，急性髓系白血病患者的预后可以通过18个位点的DNA甲基化模式来预测。我们开发了一种新策略，称为通过连接介导的PCR进行微球HpaII小片段富集（MELP），利用甲基化特异性DNA消化、荧光标记微球和分支DNA杂交同时分析这些位点的DNA甲基化模式。该方法使用的技术成本低廉且易于在分子实验室中操作。MELP准确反映了所分析的每个位点的甲基化水平，并将急性髓系白血病患者分为不同的预后亚组。我们的结果证明了MELP作为同时评估多个位点DNA甲基化的平台的实用性。
