Publication Date: 
Thursday, June 25, 2015
Digital PCR quantification of MGMT methylation refines prediction of clinical benefit from alkylating agents in glioblastoma and metastatic colorectal cancer.

Ann Oncol. 2015 Jun 25. pii: mdv272. [Epub ahead of print]
Barault L, Amatu A, Bleeker FE, Moutinho C, Falcomatà C, Fiano V, Cassingena A, Siravegna G, Milione M, Cassoni P, De Braud F, Rudà R, Soffietti R, Venesio T, Bardelli A, Wesseling P, de Witt Hamer P, Pietrantonio F, Siena S, Esteller M, Sartore-Bianchi A, Di Nicolantonio F.
O6-Methyl-Guanine-Methyl-Transferase (MGMT) silencing by promoter methylation may identify cancer patients responding to the alkylating agents dacarbazine or temozolomide.
We evaluated the prognostic and predictive value of MGMT methylation testing both in tumor and cell-free circulating DNA (cfDNA) from plasma samples using an ultra-sensitive two-step digital PCR technique (Methyl-BEAMing). Results were compared to two established techniques, Methylation specific PCR (MSP) and Bs-pyrosequencing.
Thresholds for MGMT methylated status for each technique were established in a training-set of 98 glioblastoma patients. The prognostic and the predictive value of MGMT methylated status was validated in a second cohort of 66 glioblastoma patients treated with temozolomide in which Methyl-BEAMing displayed a better specificity than the other techniques. Cut-off values of MGMT methylation specific for metastatic colorectal cancer (mCRC) tissue samples were established in a cohort of 60 patients treated with dacarbazine. In mCRC, both quantitative assays Methyl-BEAMing and Bs-pyrosequencing outperformed MSP, providing better prediction of treatment response and improvement in progression-free survival (PFS)(p<0.001). Ability of Methyl-BEAMing to identify responding patients was validated in a cohort of 23 mCRC patients treated with temozolomide and pre-selected for MGMT methylated status according to MSP. In mCRC patients treated with dacarbazine, exploratory analysis of cfDNA by Methyl-BEAMing showed that MGMT methylation was associated with better response and improved median PFS (p=0.008).
Methyl-BEAMing showed high reproducibility, specificity and sensitivity and was applicable to formalin fixed paraffin embedded tissues and cfDNA. This study supports the quantitative assessment of MGMT methylation for clinical purposes since it could refine prediction of response to alkylating agents.
© The Author 2015. Published by Oxford University Press on behalf of the European Society for Medical Oncology. All rights reserved. For permissions, please email:
DNA methylation; MGMT; Metastatic colorectal cancer; alkylating agent; cell free circulating DNA; digital PCR



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