OBJECTIVE: To analyze the reliability and reproducibility of PCR-based testing of O6-methylguanine-DNA methyltransferase gene (MGMT) promoter methylation status in formalin-fixed and paraffin-embedded (FFPE) neurosurgical biopsy specimens. MATERIALS: We used 6 FFPE neurosurgical temporal lobe specimens of children and young adults with drug-resistant epilepsy. Histopathologically, all specimens showed CNS tissue with gliosis but no tumor tissue. METHODS: For MGMT promoter methylation analysis, we used methylation specific PCR (MGMT MSP). In all 6 tissue specimens, 4 repetitive runs of MGMT MSP were performed. RESULTS: We obtained conclusive results only in 13/24 (54.2%) MGMT MSP analyses. In 5/13 (38.5%) successful MSP runs, the results indicated presence of a methylated MGMT promoter. In only 1/6 specimens, MSP yielded consistent results in all 4 repetitive runs. CONCLUSIONS: In our hands, MGMT MSP shows poor reliability and reproducibility of test results on FFPE neurosurgical tissue specimens. A more reliable method for diagnostic MGMT promoter methylation testing needs to be identified and validated by systematic testing of intra- and interlaboratory reliability and reproducibility. Alternatively, methods of tissue fixation that do not impair DNA quality but at the same time warrant high quality histopathology could facilitate molecular diagnostics.
OBJECTIVE: To analyze the reliability and reproducibility of PCR-based testing of O6-methylguanine-DNA methyltransferase gene (MGMT) promoter methylation status in formalin-fixed and paraffin-embedded (FFPE) neurosurgical biopsy specimens. MATERIALS: We used 6 FFPE neurosurgical temporal lobe specimens of children and young adults with drug-resistant epilepsy. Histopathologically, all specimens showed CNS tissue with gliosis but no tumor tissue. METHODS: For MGMT promoter methylation analysis, we used methylation specific PCR (MGMT MSP). In all 6 tissue specimens, 4 repetitive runs of MGMT MSP were performed. RESULTS: We obtained conclusive results only in 13/24 (54.2%) MGMT MSP analyses. In 5/13 (38.5%) successful MSP runs, the results indicated presence of a methylated MGMT promoter. In only 1/6 specimens, MSP yielded consistent results in all 4 repetitive runs. CONCLUSIONS: In our hands, MGMT MSP shows poor reliability and reproducibility of test results on FFPE neurosurgical tissue specimens. A more reliable method for diagnostic MGMT promoter methylation testing needs to be identified and validated by systematic testing of intra- and interlaboratory reliability and reproducibility. Alternatively, methods of tissue fixation that do not impair DNA quality but at the same time warrant high quality histopathology could facilitate molecular diagnostics.
Authors: Gisela N Castro; Niubys Cayado-Gutiérrez; Vera L Moncalero; Patricia Lima; Rodolfo Lucero De Angelis; Victor Chávez; F Darío Cuello-Carrión; Daniel R Ciocca Journal: Cell Stress Chaperones Date: 2012-07-18 Impact factor: 3.667
Authors: Mark G Hamilton; Gloria Roldán; Anthony Magliocco; John B McIntyre; Ian Parney; Jacob C Easaw Journal: J Neurooncol Date: 2010-07-21 Impact factor: 4.130