Abstract
BackgroundThe 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.MethodsA 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).Results61/321 (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.ConclusionsThe 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.
Highlights
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 aim of the present study was to improve the performance of pyrosequencing assay for Epidermal Growth Factor Receptor (EGFR) mutation detection by setting up a novel dispensation order (NDO) capable to detect and to characterize the type of mutations of exon 19 associated to the responsiveness to treat NSCLC with EGFRTyrosine Kinase Inhibitors (TKI) therapy [14], and to validate its efficacy in the clinical setting in a consecutive prospectively collected series of lung cancer specimens
Validation of the novel dispensation order on cell lines and plasmids On cell lines, the EGFR TKI response kit and the NDO procedure were both able to detect the classical deletion in the HCC827 DNA cell line at the higher (50%) and lower (25%) dilutions in terms of peak profile alteration, whereas EGFR wt DNA cell line resulted in the normal profile
Summary
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. 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. For the management of lung cancer, a crucial issue is the availability of adequate and sufficient tumor tissue potentially impair the accuracy of tumor subtyping and molecular characterization [4]. This issue can be improved by the enrichment of tumor cells using tissue microdissection prior to mutational analysis [5]. Beyond the classical therapy-responsive mutations, some “uncommon” mutations were described whose clinical significance is still poorly understood [11]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
