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Optimization of quantitative MGMT promoter methylation analysis using pyrosequencing and combined bisulfite restriction analysis

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dc.contributor.author El Maarri, Osman
dc.contributor.author Mikeska, Thomas
dc.contributor.author Bock, Christoph
dc.contributor.author Hubner, Anika
dc.contributor.author Ehrentraut, Denise
dc.contributor.author Schramm, Johannes
dc.contributor.author Felsberg, Jorg
dc.contributor.author Khl, Philip
dc.contributor.author Buttner, Reinhard
dc.contributor.author Pietsch, Torsten
dc.contributor.author Waha, Andreas
dc.date.accessioned 2017-09-13T09:03:11Z
dc.date.available 2017-09-13T09:03:11Z
dc.date.copyright 2007 en_US
dc.identifier.issn 1943-7811 en_US
dc.identifier.uri http://hdl.handle.net/10725/6179
dc.description.abstract Resistance to chemotherapy is a major complication during treatment of cancer patients. Hypermethylation of the MGMT gene alters DNA repair and is associated with longer survival of glioblastoma patients treated with alkylating agents. Therefore, MGMT promoter methylation plays an important role as a predictive biomarker for chemotherapy resistance. To adopt this established correlation into a molecular diagnosis procedure, we compared and optimized three experimental techniques [combined bisulfite restriction analysis, a primer extension- and denaturing high-performance liquid chromatography-based method named SIRPH (SNuPE ion pair-reverse phase high-performance liquid chromatography), and pyrosequencing] with regard to their accuracy of detecting MGMT promoter methylation. Initially, bisulfite sequencing was used to obtain a comprehensive methylation profile of the MGMT promoter region in 22 glioblastoma samples and in three normal brain controls. Next, we statistically identified CpG sites that best discriminate between methylated and unmethylated MGMT promoters. These results were then used to design optimal combined bisulfite restriction analysis, SIRPH, and pyrosequencing assays for accurate and cost-efficient assessment of MGMT promoter methylation. We compared all three techniques with regard to their reliability and reproducibility on well-characterized tumor samples. The optimized pyrosequencing assay performed best and provides a sensitive, robust, and easy-to-use method for quantitative assessment of MGMT methylation, for both snap-frozen and paraffin-embedded specimens. en_US
dc.language.iso en en_US
dc.title Optimization of quantitative MGMT promoter methylation analysis using pyrosequencing and combined bisulfite restriction analysis en_US
dc.type Article en_US
dc.description.version Published en_US
dc.author.school SAS en_US
dc.author.idnumber 201508713 en_US
dc.author.department Natural Sciences en_US
dc.description.embargo N/A en_US
dc.relation.journal The Journal of Molecular Diagnostics en_US
dc.journal.volume 9 en_US
dc.journal.issue 3 en_US
dc.article.pages 368-381 en_US
dc.identifier.doi https://doi.org/10.2353/jmoldx.2007.060167 en_US
dc.identifier.ctation Mikeska, T., Bock, C., El-Maarri, O., Hübner, A., Ehrentraut, D., Schramm, J., ... & Waha, A. (2007). Optimization of quantitative MGMT promoter methylation analysis using pyrosequencing and combined bisulfite restriction analysis. The Journal of Molecular Diagnostics, 9(3), 368-381. en_US
dc.author.email osman.elmaarri@lau.edu.lb en_US
dc.identifier.tou http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.php en_US
dc.identifier.url http://www.sciencedirect.com/science/article/pii/S1525157810604060 en_US
dc.author.affiliation Lebanese American University en_US


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