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Open Access Technical advance

Detection and characterization of classical and “uncommon” exon 19 Epidermal Growth Factor Receptor mutations in lung cancer by pyrosequencing

Luisella Righi1*, Alessandra Cuccurullo3, Simona Vatrano1, Susanna Cappia1, Daniela Giachino2, Paolo De Giuli3, Mara Ardine4, Silvia Novello5, Marco Volante1, Giorgio V Scagliotti5 and Mauro Papotti1

Author Affiliations

1 Divisions of Pathology, University of Torino, Regione Gonzole 10, Torino, Orbassano, 10043, Italy

2 Department of Clinical and Biological Sciences, Medical Genetics, University of Torino, Regione Gonzole 10, Torino, Orbassano, 10043, Italy

3 Pathology Unit, ASLCN2, Cuneo, Alba, Italy

4 Oncology Unit, ASLTO5, Torino, Carmagnola, Italy

5 Department of Oncology, Medical Oncology, University of Torino, Regione Gonzole 10, Torino, Orbassano, 10043, Italy

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BMC Cancer 2013, 13:114  doi:10.1186/1471-2407-13-114

Published: 13 March 2013

Abstract

Background

The management of advanced stage non-small cell lung cancer is increasingly based on diagnostic and predictive analyses performed mostly on limited amounts of tumor tissue. The evaluation of Epidermal Growth Factor Receptor (EGFR) mutations have emerged as the strongest predictor of response to EGFR-tyrosine kinase inhibitors mainly in patients with adenocarcinoma. Several EGFR mutation detection techniques are available, having both sensitivity and specificity issues, being the Sanger sequencing technique the reference standard, with the limitation of a relatively high amount of mutated cells needed for the analysis.

Methods

A novel nucleotide dispensation order for pyrosequencing was established allowing the identification and characterization of EGFR mutation not definable with commercially and clinically approved kits, and validated in a consecutive series of 321 lung cancer patients (246 biopsies or cytology samples and 75 surgical specimens).

Results

61/321 (19%) mutated cases were detected, 17 (27.9%) in exon 21 and 44 (72.1%) in exon 19, these latter corresponding to 32/44 (72.7%) classical and 12/44 (27.3%) uncommon mutations. Furthermore, a novel, never reported, point mutation, was found, which determined a premature stop codon in the aminoacidic sequence that resulted in a truncated protein in the tyrosine kinase domain, thus impairing the inhibitory effect of specific therapy.

Conclusions

The novel dispensation order allows to detect and characterize both classical and uncommon EGFR mutations. Although several phase III studies in genotypically defined groups of patients are already available, further prospective studies assessing the role of uncommon EGFR mutations are warranted.

Keywords:
Lung cancer; EGFR mutation; Exon 19; Pyrosequencing; Adenocarcinoma