Pyrosequencing Analysis of EGFR and KRAS Mutations in EUS and EBUS-Derived Cytologic Samples of Adenocarcinomas of the Lung
Patients with stage IV non–small-cell lung cancer harboring an activating epidermal growth factor receptor (EGFR) mutation are eligible for treatment with EGFR tyrosine kinase inhibitors. With pyrosequencing, low-frequency mutations may be detected more easily even in small diagnostic samples like e...
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Published in | Journal of thoracic oncology Vol. 8; no. 8; pp. 1012 - 1018 |
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Main Authors | , , , , |
Format | Journal Article |
Language | English |
Published |
United States
Elsevier Inc
01.08.2013
Copyright by the European Lung Cancer Conference and the International Association for the Study of Lung Cancer |
Subjects | |
Online Access | Get full text |
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Summary: | Patients with stage IV non–small-cell lung cancer harboring an activating epidermal growth factor receptor (EGFR) mutation are eligible for treatment with EGFR tyrosine kinase inhibitors. With pyrosequencing, low-frequency mutations may be detected more easily even in small diagnostic samples like endoscopic ultrasound-guided fine needle aspirations (EUS-FNA) and endobronchial ultrasound-guided transbronchial needle aspirations (EBUS-TBNA). The diagnostic performance of pyrosequencing in analyzing cytological specimens is compared with the routinely used high-resolution melting (HRM) and Sanger sequencing.
Patients diagnosed with adenocarcinoma of the lung were selected from a fine needle aspiration and transbronchial needle aspiration specimen database. If formalin-fixed paraffin-embedded tumor blocks were available, mutation analysis was performed for EGFR and V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog genes using both pyrosequencing and HRM. When HRM showed abnormalities, Sanger sequencing was used.
A total of 126 samples were available for mutation analysis. The analysis success rate for pyrosequencing and HRM were 97% and 93%, respectively. HRM failures were observed in fragmented DNA showing chains of 100 to 200 bp. A significant correlation between length of DNA fragments (100–300 bp versus 300–400 bp) and mean sample age (797 versus 317 days) was found (p < 0.0001), suggesting an influence of sample age on DNA quality.
Pyrosequencing on cytological blocks, especially older tumor blocks, is feasible with a high diagnostic success rate. Failures in HRM were observed in DNA samples with short fragments related to longer storage times. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
ISSN: | 1556-0864 1556-1380 |
DOI: | 10.1097/JTO.0b013e31829ce93e |