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BioMed Research International
Volume 2013 (2013), Article ID 385087, 9 pages
http://dx.doi.org/10.1155/2013/385087
Research Article

Detection of EGFR Mutations by TaqMan Mutation Detection Assays Powered by Competitive Allele-Specific TaqMan PCR Technology

1Laboratory of Pharmacogenomics, Centro di Ricerche Oncologiche di Mercogliano (CROM), Istituto Nazionale per lo Studio e la Cura dei Tumori “Fondazione Giovanni Pascale”(IRCCS), 83013 Mercogliano, Italy
2Cell Biology and Biotherapy Unit, Istituto Nazionale per lo Studio e la Cura dei Tumori “Fondazione Giovanni Pascale”(IRCCS), 80131 Naples, Italy
3Surgical Pathology Unit, Istituto Nazionale per lo Studio e la Cura dei Tumori “Fondazione Giovanni Pascale”(IRCCS), 80131 Naples, Italy
4Department of Molecular and Clinical Medicine, Laboratory of Molecular and Cellular Biology, Università “La Sapienza”, 00189 Rome, Italy
5Department of Surgery “A. Valdoni”, Laboratory of Molecular and Cellular Biology, Università “La Sapienza”, 00161 Rome, Italy
6Medical Oncology Unit, SS Annunziata Hospital, 74123 Taranto, Italy
7Medical Oncology Unit, Department of Thoracic Surgical and Medical Oncology, Istituto Nazionale per lo Studio e la Cura dei Tumori “Fondazione Giovanni Pascale”(IRCCS), 80131 Naples, Italy

Received 30 August 2013; Accepted 4 October 2013

Academic Editor: Franco M. Buonaguro

Copyright © 2013 Cristin Roma et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Epidermal growth factor receptor (EGFR) mutations in non-small-cell lung cancer (NSCLC) are predictive of response to treatment with tyrosine kinase inhibitors. Competitive Allele-Specific TaqMan PCR (castPCR) is a highly sensitive and specific technology. EGFR mutations were assessed by TaqMan Mutation Detection Assays (TMDA) based on castPCR technology in 64 tumor samples: a training set of 30 NSCLC and 6 colorectal carcinoma (CRC) samples and a validation set of 28 NSCLC cases. The sensitivity and specificity of this method were compared with routine diagnostic techniques including direct sequencing and the EGFR Therascreen RGQ kit. Analysis of the training set allowed the identification of the threshold value for data analysis (0.2); the maximum cycle threshold ( ); and the cut-off ΔCt value (7) for the EGFR TMDA. By using these parameters, castPCR technology identified both training and validation set EGFR mutations with similar frequency as compared with the Therascreen kit. Sequencing detected rare mutations that are not identified by either castPCR or Therascreen, but in samples with low tumor cell content it failed to detect common mutations that were revealed by real-time PCR based methods. In conclusion, our data suggest that castPCR is highly sensitive and specific to detect EGFR mutations in NSCLC clinical samples.