Validation of real-time methylation-specific PCR to determine O6-methylguanine-DNA methyltransferase gene promoter methylation in glioma.

Détails

ID Serval
serval:BIB_CC92294BD2B5
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Titre
Validation of real-time methylation-specific PCR to determine O6-methylguanine-DNA methyltransferase gene promoter methylation in glioma.
Périodique
Journal of Molecular Diagnostics
Auteur(s)
Vlassenbroeck I., Califice S., Diserens A.C., Migliavacca E., Straub J., Di Stefano I., Moreau F., Hamou M.F., Renard I., Delorenzi M., Flamion B., DiGuiseppi J., Bierau K., Hegi M.E.
ISSN
1525-1578 (Print)
ISSN-L
1525-1578
Statut éditorial
Publié
Date de publication
2008
Peer-reviewed
Oui
Volume
10
Numéro
4
Pages
332-337
Langue
anglais
Notes
Publication types: Journal Article ; Research Support, Non-U.S. Gov't
Publication Status: ppublish
Résumé
Epigenetic silencing of the DNA repair protein O(6)-methylguanine-DNA methyltransferase (MGMT) by promoter methylation predicts successful alkylating agent therapy, such as with temozolomide, in glioblastoma patients. Stratified therapy assignment of patients in prospective clinical trials according to tumor MGMT status requires a standardized diagnostic test, suitable for high-throughput analysis of small amounts of formalin-fixed, paraffin-embedded tumor tissue. A direct, real-time methylation-specific PCR (MSP) assay was developed to determine methylation status of the MGMT gene promoter. Assay specificity was obtained by selective amplification of methylated DNA sequences of sodium bisulfite-modified DNA. The copy number of the methylated MGMT promoter, normalized to the beta-actin gene, provides a quantitative test result. We analyzed 134 clinical glioma samples, comparing the new test with the previously validated nested gel-based MSP assay, which yields a binary readout. A cut-off value for the MGMT methylation status was suggested by fitting a bimodal normal mixture model to the real-time results, supporting the hypothesis that there are two distinct populations within the test samples. Comparison of the tests showed high concordance of the results (82/91 [90%]; Cohen's kappa = 0.80; 95% confidence interval, 0.82-0.95). The direct, real-time MSP assay was highly reproducible (Pearson correlation 0.996) and showed valid test results for 93% (125/134) of samples compared with 75% (94/125) for the nested, gel-based MSP assay. This high-throughput test provides an important pharmacogenomic tool for individualized management of alkylating agent chemotherapy.
Mots-clé
DNA Methylation, Glioma/diagnosis, Glioma/genetics, Humans, O(6)-Methylguanine-DNA Methyltransferase/genetics, Polymerase Chain Reaction/methods, Promoter Regions, Genetic/genetics, Reproducibility of Results, Sensitivity and Specificity
Pubmed
Web of science
Création de la notice
26/02/2008 9:12
Dernière modification de la notice
03/03/2018 21:28
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