title: Comparison of PNA clamping and direct sequencing for detecting KRAS mutations in matched tumour tissue, cell block, pleural effusion and serum from patients with malignant pleural effusion.
abstract: OBJECTIVE
  Peptide nucleic acid (PNA)-mediated real-time polymerase chain reaction clamping was recently developed to improve mutation detection sensitivity. Pleural effusion could be a good sample candidate for mutation analysis. To establish if PNA clamping could be used to detect KRAS mutation in particular in pleural effusion, we analysed its diagnostic performance.
  
  
  METHODS
  We studied 57 patients with malignant effusion. KRAS mutation was evaluated in samples of matched tumour tissue, cell block, pleural effusion and serum using PNA clamping and direct sequencing.
  
  
  RESULTS
  The detection rate of KRAS mutation using pleural effusion was 14% for PNA clamping and 10.5% for direct sequencing. The κ coefficient between the two methods was 0.76 (P value < 0.0001), 1.00 (P value < 0.0001) and 0.87 (P value < 0.0001) in pleural effusion, tissue and cell block, respectively. The diagnostic performance of KRAS mutation detection from pleural effusion compared with the results obtained for all samples combined showed that the sensitivity, specificity, positive predictive value and negative predictive value were as follows: 89, 100, 100 and 98%, respectively for PNA clamping; 67, 100, 100 and 94%, respectively for directing sequencing.
  
  
  CONCLUSIONS
  The current study suggests that PNA clamping had a good concordance with direct sequencing for the detection of KRAS mutation in patients with malignant effusion. Furthermore, the good diagnostic performance obtained from pleural effusion samples provides evidence that pleural effusion can be a useful source for detecting KRAS mutation in a clinical setting, in which the collection of tumour tissues is challenging.
authors.affiliation: Department of Internal Medicine, Seoul St Mary's Hospital, The Cancer Research Institute, Seoul, Korea.
authors.forename: Ji Young
authors.initials: JY
authors.lastname: Kang
authors.forename: Chan Kwon
authors.initials: CK
authors.lastname: Park
authors.forename: Chang Dong
authors.initials: CD
authors.lastname: Yeo
authors.forename: Hea Yeon
authors.initials: HY
authors.lastname: Lee
authors.forename: Chin Kook
authors.initials: CK
authors.lastname: Rhee
authors.forename: Seung Joon
authors.initials: SJ
authors.lastname: Kim
authors.forename: Seok Chan
authors.initials: SC
authors.lastname: Kim
authors.forename: Young Kyoon
authors.initials: YK
authors.lastname: Kim
authors.forename: Mi Sun
authors.initials: MS
authors.lastname: Park
authors.forename: Hyeon Woo
authors.initials: HW
authors.lastname: Yim
mesh_terms: D000368:Aged; D002289:Carcinoma, Non-Small-Cell Lung / Q000150:complications / Q000473:pathology; D004252:DNA Mutational Analysis / Q000379:methods; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D009154:Mutation; D020135:Peptide Nucleic Acids / Q000378:metabolism; D016066:Pleural Effusion, Malignant / Q000209:etiology / Q000235:genetics / Q000473:pathology; D011237:Predictive Value of Tests; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras); D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D018631:ras Proteins / Q000235:genetics*
chemical_list: C117307:KRAS protein, human; D020135:Peptide Nucleic Acids; D011518:Proto-Oncogene Proteins; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins
keywords: Kirsten rat sarcoma viral oncogene homologue (KRAS) mutation; direct sequencing; malignant pleural effusion; peptide nucleic acid-mediated real-time PCR clamping