Prognosis of glioblastoma with faint MGMT methylation-specific PCR product

Abstract

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.

Prognostic Relevance of Tumor Purity and Interaction with MGMT Methylation in Glioblastoma

“…Using pyrosequencing, Dunn and colleagues found a survival advantage of highly methylated tumors in a cohort of 109 patients homogeneously treated with temozolomide and standard radiotherapy (1). Similarly, another study reported an intermediate overall survival for patients with tumors producing a faint band in methylation-specific PCR compared with both unmethylated and distinctly methylated cases (32). Our analysis of TCGA data suggests that tumor purity is a predictor of survival, raising the question of which biological mechanism is responsible for this phenomenon.…”

MGMT Gene Promoter Methylation Status – Assessment of Two Pyrosequencing Kits and Three Methylation-specific PCR Methods for their Predictive Capacity in Glioblastomas

“…A major disadvantage with using MSP and qMSP is that both these techniques only detect fully methylated MGMT promoter at the annealing sites of primers and probe (see Figure 1). This may lead to a problem in sensitivity; however, both these methods have in previous studies been found to be of prognostic value (1,(21)(22)(23)(24)(25)(26)(27)(28)(29)(30)39).…”

“…MGMT promoter methylation, regardless of whether measured by MSP, qMSP, PSQ, MS-HRM, MS-MLPA, or MethyLight qMSP, has been demonstrated to predict clinical outcome (1,13,(21)(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32)(33). PSQ has been claimed to be best technique to identify patients with GBM likely to benefit from therapy with temozolomide (21)(22)(23)(24).…”

Detection of human cytomegalovirus in glioblastoma among Taiwanese subjects

“…Cox regression model was used to adjust the influence of age, gender, KPS, extent of resection, bevacizumab treatment, isocitrate dehydrogenase 1 (IDH1) mutation and O 6 -methylguanine-DNA-methyltranferase (MGMT) status. The data of the MGMT status were from our previous study [2] and samples with methylation-specific PCR products of any intensity were regarded as a positive result. P -values were derived from two-tailed tests and p < 0.05 was considered significant.…”

“…Patients’ survival has significantly improved since the use of radiotherapy with concomitant and adjuvant temozolomide (TMZ) chemotherapy and its median survival is 20.0 months at our institute [1, 2]. However, in comparing with other common malignancies in Taiwan, such as colon and breast cancers, its survival is still poor.…”