1] Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON, Canada [2] Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada.
Ontario Cancer Institute, Princess Margaret Cancer Center-University Health Network, Toronto, ON, Canada.
Sci Rep. 2014 Mar 21;4:4432. doi: 10.1038/srep04432.
Aberrant DNA methylation is a hallmark of cancer and is an important potential biomarker. Particularly, combined analysis of a panel of hypermethylated genes shows the most promising clinical performance. Herein, we developed, optimized and standardized a multiplex MethyLight assay to simultaneously detect hypermethylation of APC, HOXD3 and TGFB2 in DNA extracted from prostate cancer (PCa) cell lines, archival tissue specimens, and urine samples. We established that the assay is capable of discriminating between fully methylated and unmethylated alleles with 100% specificity and demonstrated the assay as highly accurate and reproducible as the singleplex approach. For proof of principle, we analyzed the methylation status of these genes in tissue and urine samples of PCa patients as well as PCa-free controls. These data show that the multiplex MethyLight assay offers a significant advantage when working with limited quantities of DNA and has potential applications in research and clinical settings.
异常的 DNA 甲基化是癌症的一个标志，也是一个重要的潜在生物标志物。特别是，一组超甲基化基因的联合分析显示出最有前途的临床性能。在此，我们开发、优化并标准化了一种多重 MethyLight 测定法，以同时检测从前列腺癌 (PCa) 细胞系、存档组织标本和尿液样本中提取的 DNA 中 APC、HOXD3 和 TGFB2 的过度甲基化。我们证实该测定法能够以 100%的特异性区分完全甲基化和非甲基化等位基因，并证明该测定法与单重测定法一样高度准确和可重复。为了验证原理，我们分析了 PCa 患者和无 PCa 对照组组织和尿液样本中这些基因的甲基化状态。这些数据表明，当使用有限量的 DNA 时，多重 MethyLight 测定法具有显著优势，并具有在研究和临床环境中的潜在应用。
