Paired Box 5 Methylation Detection by Droplet Digital PCR for Ultra-Sensitive Deep Surgical Margins Analysis of Head and Neck Squamous Cell Carcinoma
Paired Box 5 Methylation Detection by Droplet Digital PCR for Ultra-Sensitive Deep Surgical Margins Analysis of Head and Neck Squamous Cell Carcinoma
复制标题
DOI：
10.1158/1940-6207.capr-15-0180
复制
发表时间：
2015-11-01
影响因子：
3.3
通讯作者：
Koch, Wayne M.
中科院分区：
文献类型：
--
作者：
Hayashi, Masamichi;Guerrero-Preston, Rafael;Koch, Wayne M.
分子深部手术切缘分析已被证明可以预测头颈部鳞状细胞癌（HNSCC）的局部复发。为了提高分析的准确性和通用性，我们使用了高度肿瘤特异性甲基化标记和高度灵敏的检测技术来测试手术切缘的DNA。通过印记程序（n = 75）和初次组织采集（n = 70），从82例符合条件的HNSCC手术中前瞻性采集组织学癌症阴性深部手术切缘样本。通过常规定量甲基化特异性PCR（QMSP）和通过靶向配对框5（PAX 5）基因启动子甲基化的液滴数字PCR（ddQMSP）的QMSP分析来自每个样品的亚硫酸氢盐处理的DNA。评估PAX 5甲基化的存在与局部无复发生存期（LRFS）之间的关联。PAX 5甲基化在印迹样品中的68.0%（51/75）的肿瘤和原发组织样品中的71.4%（50/70）中被发现。在未接受术后放疗的病例中（印记样本n = 31，组织样本n = 29），传统QMSP和ddQMSP均显示，通过单变量分析，PAX 5甲基化阳性边缘与LRFS不良显着相关。特别是，ddQMSP增加检测PAX 5标记从29%到71%的非辐射的印记的情况下。此外，通过多变量分析，PAX 5甲基化印迹边缘是LRFS较差的良好预测因子[HR，3.89; 95%置信区间（CI），1.19-17.52; P = 0.023]。PAX 5甲基化似乎是一个很好的肿瘤特异性标记物，用于HNSCC的分子深部手术切缘分析。此外，ddQMSP测定显示甲基化标志物检测的灵敏度增加。(C)2015年AACR。
Molecular deep surgical margin analysis has been shown to predict locoregional recurrences of head and neck squamous cell carcinoma (HNSCC). To improve the accuracy and versatility of the analysis, we used a highly tumor-specific methylation marker and highly sensitive detection technology to test DNA from surgical margins. Histologically cancer-negative deep surgical margin samples were prospectively collected from 82 eligible HNSCC surgeries by an imprinting procedure (n = 75) and primary tissue collection (n = 70). Bisulfite-treated DNA from each sample was analyzed by both conventional quantitative methylation-specific PCR (QMSP) and QMSP by droplet digital PCR (ddQMSP) targeting Paired box 5 (PAX5) gene promoter methylation. The association between the presence of PAX5 methylation and locoregional recurrence-free survival (LRFS) was evaluated. PAX5 methylation was found in 68.0% (51 of 75) of tumors in the imprint samples and 71.4% (50 of 70) in the primary tissue samples. Among cases that did not have postoperative radiation (n = 31 in imprint samples, n = 29 in tissue samples), both conventional QMSP and ddQMSP revealed that PAX5 methylation-positive margins was significantly associated with poor LRFS by univariate analysis. In particular, ddQMSP increased detection of the PAX5 marker from 29% to 71% in the nonradiated imprint cases. Also, PAX5 methylated imprint margins were an excellent predictor of poor LRFS [HR, 3.89; 95% confidence interval (CI), 1.19-17.52; P = 0.023] by multivariate analysis. PAX5 methylation appears to be an excellent tumor-specific marker for molecular deep surgical margin analysis of HNSCC. Moreover, the ddQMSP assay displays increased sensitivity for methylation marker detection. (C) 2015 AACR.