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R. Langfort



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    P1.06 - Poster Session 1 - Prognostic and Predictive Biomarkers (ID 161)

    • Event: WCLC 2013
    • Type: Poster Session
    • Track: Biology
    • Presentations: 1
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      P1.06-044 - Diagnostic validation of PNA-LNA PCR clamp assay for detection of EGFR exon 19 and 21 mutations in non-small cell lung cancer specimens. (ID 2729)

      09:30 - 16:30  |  Author(s): R. Langfort

      • Abstract

      Background
      PNA-LNA PCR clamp is highly sensitive, real-time PCR-based laboratory technique developed to enable reliable detection of EGFR gene mutations in wide spectrum of tissue/biopsy samples from NSCLC patients. The aim of the study was to assess diagnostic reliability of PNA-LNA PCR clamp assay in EGFR mutations detection in different NSCLC samples.

      Methods
      Evaluation was performed: (i) in reference NSCLC tissue FFPE samples (n=10), (ii) in comparison to direct sequencing in resected NSCLC tissue (n=199) and biopsy material specimens (n=179) characterized by different tumor cells content (TCC) and fixation [Table 1].

      [Table 1.] NSCLC samples
      Resected tissue Biopsy material
      fresh -frozen FFPE FFPE Cytology smear
      84 115 115 64

      Results
      (i) PNA-LNA PCR clamp correctly detected all exon 19 deletions and L858R mutations in the reference FFPE materials, including those with meager TCC (5% and 10%). (ii) In total of 378 samples analyzed with PNA-LNA PCR clamp method EGFR mutations were detected in 36 (9.5%). No significant differences in detection efficiency were observed in reference to material (resected tissue vs biopsy, p=0,3972; OR=0,7405; CI=0,3694-1,4844) and fixation procedure (FFPE vs fresh-frozen tissue, p=0,5459; OR=0,7304; CI=0,2635-2,0248; biopsy material FFPE vs cytology smear, p=0,4366, OR=0,6908; CI=0,272-2,7544). (iii) PNA-LNA PCR clamp method and direct sequencing presented high conformity (overall percent agreement, OPA=99%; Cohen’s Kappa score of 0.94 (95% CI=0.9, 0.99) in n=100 samples with >50% TCC. (iv) PNA-LNA PCR clamp presented higher sensitivity in samples with TCC <50% (p=0.004). Reevaluation with direct sequencing proved positive only in 24 out of 36 (67%) mutation positive samples [Table 2].

      [Table 2. ]Comparison of PNA-LNA PCR clamp vs direct sequencing EGFR mutation detection sensitivity in 36 EGFR mutation positive materials with different %TCC.
      PNA-LNA PCR clamp direct sequencing
      ≥50% 25/25 (100%) 22/25 (88%)
      20<50% 6/6 (100%) 2/6 (33%)
      ≤20% 5/5 (100%) 0/5 (0%)
      total 36/36 (100%) 24/36 (67%)

      Conclusion
      PNA-LNA PCR clamp method is characterized by high sensitivity of EGFR exon 19 and 21 mutations detection in tissue and biopsy material, particularly in samples with TCC lower than 50%. Fixation procedures did not affect PNA-LNA PCR clamp method mutation detection effectiveness.

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    P3.06 - Poster Session 3 - Prognostic and Predictive Biomarkers (ID 178)

    • Event: WCLC 2013
    • Type: Poster Session
    • Track: Biology
    • Presentations: 1
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      P3.06-038 - New scoring system assessing the probability of malignancy as the cause of large pericardial effusion is able to improve the diagnostic algorithm of neoplastic pericarditis. (ID 2750)

      09:30 - 16:30  |  Author(s): R. Langfort

      • Abstract

      Background
      Neoplastic pericarditis is found in 60-80% of patients (pts) with large pericardial effusion (pe) (>2 cm on echocardiography). The most frequent causes of neoplastic pe are metastatic lung or breast cancer. Malignant pe is combined with high relapse rate after pericardiocentesis and poor prognosis. Early recognition of malignant pe and intensive local (+/-) systemic treatment is able to improve life expectancy. Nevertheless in 30-40% of pts the malignant cause of pe is not recognized during the first episode of disease. The aim of the study was to introduce the new scoring system assessing the probability of malignant pe in the pts requiring pericardial fluid drainage due to large pe/tamponade.

      Methods
      146 pts, median age 57 years (21-88), 80 with benign cause of pe, 66 with neoplastic cause of pe ( positive pe cytology or neoplastic infiltration in pericardial biopsy specimen) treated in the National Institute of Tuberculosis and Lung Diseases in 1982 - 2008 entered the study. Metastatic lung cancer was diagnosed in 67% of neoplastic pe.

      Results
      Based on previous results, the most important features distinguishing between neoplastic and benign pe were: cardiac tamponade on echocardiographic examination, HR>90 beats/min on ECG, enlarged mediastinal lymph nodes (>1cm) on chest CT scan, CEA> 5ng/ml in pe, Cyfra 21-1>95 ng/ml in pe, bloody pe. The sensitivity, specificity, PPV and NPV are listed in table 1.

      Parameter Diag. value HR>90/min Cardiac tamponade CEA>5 ng/ml (pe) Cyfra 21-1>95 ng/ml (pe) Bloody pe Lymph nodes>1 cm (CT)
      sensitivity 0.82 0.67 0.63 0.64 0.86 0.93
      specificity 0.55 0.65 0.94 0.95 0.57 0.70
      PPV 0.60 0.61 0.91 0.93 0.66 0.66
      NPV 0.79 0.70 0.71 0.71 0.80 0.94
      The original scoring system (-3 up to + 3 points) was developed based on PPV and NPV of the above mentioned parameters. The diagnostic value of the proposed scoring system was high, ( ROC: AUC 0.926 95%CI 0.85-0.96) and exceeding the value of single parameters (at a cut off of 0 points – sensitivity was 0.84 and specificity - 0.91).

      Conclusion
      Assessment of the probability of malignant pe according to the proposed original scoring system was able to improve the diagnostic algorithm based on single parameters, thus indicating pts in whom more invasive diagnostic methods should be applied to recognize malignant pe.