Hsu Chih-Yi, Ho Hsiang-Ling, Lin Shih-Chieh, Chang-Chien Yi-Chun, Chen Ming-Hsiung, Hsu Sanford Ping-Chuan, Yen Yu-Shu, Guo Wan-You, Ho Donald Ming-Tak
Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, No. 201, Sec 2, Shih-Pai Road, Taipei, 11217, Taiwan.
J Neurooncol. 2015 Mar;122(1):179-88. doi: 10.1007/s11060-014-1701-1. Epub 2015 Jan 10.
Methylation-specific polymerase chain reaction (MSP) for the promoter methylation status of O(6)-methylguanine-DNA-methyltranferase (MGMT) gene theoretically provides a positive or negative result. However, the faint MSP product is difficult to interpret. The aim of this study was to evaluate the significance of faint MSP product in glioblastoma (GBM). Critical concentrations of methylated control DNA, i.e., 100, 1, 0.5 and 0 % were run parallel with 116 newly diagnosed GBMs in order to standardize the interpretation and to distinguish positive (+), equivocal (±), and negative (-; unmethylated) results. Cases with the faint MSP product and its intensity between those of 1 and 0.5 % DNA controls were considered equivocal (±). MGMT methylation quantifications were also determined by quantitative real-time MSP (qMSP) and pyrosequencing (PSQ), and protein expression was detected by immunohistochemistry. There were significant correlations between MSP and all the aforementioned studies. The concordance rates between the MSP+ and qMSP+ cases, as well as the MSP- and qMSP- cases were 100 %, and the MSP± cases comprised 76.5 % of qMSP+ cases and 23.5 % of qMSP- cases. PSQ study showed that heterogeneous methylation was more frequently encountered in the MSP± cases. Multivariate analyses disclosed that although the overall survival of the MSP± cases was indistinct from that of the MSP+ cases, its progression free survival was significantly worse and was indistinct from that of the MSP- cases. In conclusion, GBMs with faint MGMT MSP products should be distinguished from MSP+ cases as their behaviors were different.
用于检测O(6)-甲基鸟嘌呤-DNA甲基转移酶（MGMT）基因启动子甲基化状态的甲基化特异性聚合酶链反应（MSP）理论上会给出阳性或阴性结果。然而，微弱的MSP产物难以解读。本研究的目的是评估胶质母细胞瘤（GBM）中微弱MSP产物的意义。将甲基化对照DNA的临界浓度，即100%、1%、0.5%和0%与116例新诊断的GBM平行检测，以规范解读并区分阳性（+）、可疑（±）和阴性（-；未甲基化）结果。微弱MSP产物及其强度介于1%和0.5%DNA对照之间的病例被视为可疑（±）。MGMT甲基化定量也通过定量实时MSP（qMSP）和焦磷酸测序（PSQ）测定，蛋白质表达通过免疫组织化学检测。MSP与上述所有研究之间存在显著相关性。MSP+与qMSP+病例以及MSP-与qMSP-病例之间的一致率为100%，MSP±病例占qMSP+病例的76.5%和qMSP-病例的23.5%。PSQ研究表明，MSP±病例中更常出现异质性甲基化。多变量分析显示，虽然MSP±病例的总生存期与MSP+病例无明显差异，但其无进展生存期明显更差，与MSP-病例无明显差异。总之，MGMT MSP产物微弱的GBM应与MSP+病例区分开来，因为它们的行为不同。
