Comparative Analysis of Peptide Nucleic Acid (PNA)-Mediated Real-Time PCR Clamping and DNA Direct Sequencing for EGFR Mutation Detection.

Tuberculosis and Respiratory Diseases

article

Kim, Hee Joung¹  Kim, Wan Seop²  Shin, Kyeong Cheol³  Lee, Gwan Ho³  Kim, Mi Jin⁴  Lee, Jeong Eun⁵  Song, Kyu Sang⁶  Kim, Sun Young⁵  Lee, Kye Young⁷ 

[1] Departments of Internal Medicine, Konkuk University School of Medicine;Departments of Pathology, Konkuk University School of Medicine;Departments of Internal Medicine, Yeungnam University College of Medicine;Departments of Pathology, Yeungnam University College of Medicine;Departments of Internal Medicine, Chungnam National University College of Medicine;Departments of Pathology, Chungnam National University College of Medicine;Departments of Chungnam National University College of Medicine

关键词: Peptide Nucleic Acids;    Molecular Sequencing Data;    Receptor;    Epidermal Growth Factor;    Epidermal Growth Factor;   

DOI  :  10.4046/trd.2011.70.1.21

学科分类：医学（综合）

来源: The Korean Academy of Tuberculosis and Respiratory Diseases

【 摘 要 】

BACKGROUND Although the gold standard method for research trials on epidermal growth factor receptor (EGFR) mutations has been direct sequencing, this approach has the limitations of low sensitivity and of being time-consuming. Peptide nucleic acid (PNA)-mediated polymerase chain reaction (PCR) clamping is known to be a more sensitive detection tool. The aim of this study was to compare the detection rate of EGFR mutation and EGFR-tyrosine kinase inhibitor (TKI) responsiveness according to EGFR mutation status using both methodologies. METHODS: Clinical specimens from 112 NSCLC patients were analyzed for EGFR mutations in exons 18, 19, 20, and 21. All clinical data and tumor specimens were obtained from 3 university hospitals in Korea. After genomic DNA was extracted from paraffin-embedded tissue specimens, both PNA-mediated PCR clamping and direct-sequencing were performed. The results and clinical response to EGFR-TKIs were compared. RESULTS: Sequencing revealed a total of 35 (22.9%) mutations: 8 missense mutations in exon 21 and 26 deletion mutations in exon 19. PNA-mediated PCR clamping showed the presence of genomic alterations in 45 (28.3%) samples, including the 32 identified by sequencing plus 13 additional samples (6 in exon 19 and 7 in exon 21). CONCLUSION: PNA-mediated PCR clamping is simple and rapid, as well as a more sensitive method for screening of genomic alterations in EGFR gene compared to direct sequencing. This data suggests that PNA-mediated PCR clamping should be implemented as a useful screening tool for detection of EGFR mutations in clinical setting.

【 授权许可】

CC BY-NC