Detection of MGMT promoter methylation in glioblastoma using pyrosequencing.

Abstract	Recent clinical trials on patients with glioblastoma revealed that O6-Methylguanine-DNA methyltransferase (MGMT) methylation status significantly predicts patient's response to alkylating agents. In this study, we sought to develop and validate a quantitative MGMT methylation assay using pyrosequencing on glioblastoma. We quantified promoter methylation of MGMT using pyrosequencing on paraffin-embedded fine needle aspiration biopsy tissues from 43 glioblastoma. Using a 10% cutoff, MGMT methylation was identified in 37% cases of glioblastoma and 0% of the non-neoplastic epileptic tissue. Methylation of any individual CpG island in MGMT promoter ranged between 33% and 95%, with a mean of 65%. By a serial dilution of genomic DNA of a homogenously methylated cancer cell line with an unmethylated cell line, the analytical sensitivity is at 5% for pyrosequencing to detect MGMT methylation. The minimal amount of genomic DNA required is 100 ng (approximately 3,000 cells) in small fine needle biopsy specimens. Compared with methylation-specific PCR, pyrosequencing is comparably sensitive, relatively specific, and also provides quantitative information for each CpG methylation.
Authors	Hao Xie, Raymond Tubbs, Bin Yang
Journal	International journal of clinical and experimental pathology (Int J Clin Exp Pathol) Vol. 8 Issue 2 Pg. 1790-6 ( 2015) ISSN: 1936-2625 [Electronic] United States
PMID	25973069 (Publication Type: Journal Article)
Chemical References	Tumor Suppressor Proteins DNA Modification Methylases MGMT protein, human DNA Repair Enzymes
Topics	Biopsy, Fine-Needle Brain Neoplasms (enzymology, genetics, pathology) Case-Control Studies Cell Line, Tumor CpG Islands DNA Methylation DNA Modification Methylases (genetics) DNA Repair Enzymes (genetics) Glioblastoma (enzymology, genetics, pathology) Humans Paraffin Embedding Promoter Regions, Genetic Sequence Analysis, DNA (methods) Tumor Suppressor Proteins (genetics)

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