Tuberculosis and Respiratory Diseases

The Korean Academy of Tuberculosis and Respiratory Diseases (대한결핵및호흡기학회)
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Comparative Analysis of Peptide Nucleic Acid (PNA)-Mediated Real-Time PCR Clamping and DNA Direct Sequencing for EGFR Mutation Detection

EGFR 돌연변이 검출에 있어 PNA-Mediated Real-Time PCR Clamping과 직접 염기서열 분석법의 비교 분석

  • Kim, Hee-Joung (Department of Internal Medicine, Konkuk University School of Medicine) ;
  • Kim, Wan-Seop (Department of Pathology, Konkuk University School of Medicine) ;
  • Shin, Kyeong-Cheol (Department of Internal Medicine, Yeungnam University College of Medicine) ;
  • Lee, Gwan-Ho (Department of Internal Medicine, Yeungnam University College of Medicine) ;
  • Kim, Mi-Jin (Department of Pathology, Yeungnam University College of Medicine) ;
  • Lee, Jeong-Eun (Department of Internal Medicine, Chungnam National University College of Medicine) ;
  • Song, Kyu-Sang (Department of Pathology, Chungnam National University College of Medicine) ;
  • Kim, Sun-Young (Department of Internal Medicine, Chungnam National University College of Medicine) ;
  • Lee, Kye-Young (Department of Internal Medicine, Konkuk University School of Medicine)
  • 김희정 (건국대학교 의학전문대학원 내과학교실) ;
  • 김완섭 (건국대학교 의학전문대학원 병리학교실) ;
  • 신경철 (영남대학교 의과대학 내과학교실) ;
  • 이관호 (영남대학교 의과대학 내과학교실) ;
  • 김미진 (영남대학교 의과대학 병리학교실) ;
  • 이정은 (충남대학교 의과대학 내과학교실) ;
  • 송규상 (충남대학교 의과대학 병리학교실) ;
  • 김선영 (충남대학교 의과대학 내과학교실) ;
  • 이계영 (건국대학교 의학전문대학원 내과학교실)
  • Received : 2010.12.11
  • Accepted : 2011.01.14
  • Published : 2011.01.30
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Abstract

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.

Keywords

References

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