title: Prognostic Role of Methylation Status of the MGMT Promoter Determined Quantitatively by Pyrosequencing in Glioblastoma Patients.
abstract: OBJECTIVE
  This study investigated whether pyrosequencing can be used to determine the methylation status of the MGMT promoter as a clinical biomarker using relatively old archival tissue samples of glioblastoma. We also examined other prognostic factors for survival of glioblastoma patients.
  
  
  METHODS
  The available study set included formalin-fixed paraffin-embedded (FFPE) tissue from 104 patients at two institutes from 1997 to 2012, all of which were diagnosed histopathologically as glioblastoma. Clinicopathologic data were collected by review of medical records. For pyrosequencing analysis, the PyroMark Q96 CpG MGMT kit (Qiagen, Hilden, Germany) was used to detect the level of methylation at exon 1 positions 17-39 of the MGMT gene, which contains 5 CpGs.
  
  
  RESULTS
  Methylation of the MGMT promoter was detected in 43 (41.3%) of 104 samples. The average percentage methylation was 14.0±16.8% overall and 39.0±14.7% for methylated cases. There was no significant pattern of linear increase or decrease according to the age of the FFPE block (p=0.687). In multivariate analysis, age, performance status, extent of surgery, method of adjuvant therapy, and methylation status estimated by pyrosequencing were independently associated with overall survival. Additionally, patients with a high level of methylation survived longer than those with low methylation (p=0.016).
  
  
  CONCLUSIONS
  In this study, the status and extent of methylation of the MGMT promoter analyzed by pyrosequencing were associated with overall survival in glioblastoma patients. Pyrosequencing is a quantitative method that overcomes the problems of MSP and a simple technique for accurate analysis of DNA sequences.
authors.affiliation: Department of Pathology, Dong-A University College of Medicine, Busan, Korea.
authors.forename: Dae Cheol
authors.initials: DC
authors.lastname: Kim
authors.affiliation: Department of Neurosurgery, Dong-A University College of Medicine, Busan, Korea.
authors.forename: Ki Uk
authors.initials: KU
authors.lastname: Kim
authors.affiliation: Division of Neurooncology, Department of Neurosurgery, Samsung Changwon Hospital, Sungkyunkwan University School of Medicine, Changwon, Korea.
authors.forename: Young Zoon
authors.initials: YZ
authors.lastname: Kim
keywords: Glioblastoma; MGMT promoter; Methylation; Prognosis; Pyrosequencing