Oops, you're using an old version of your browser so some of the features on this page may not be displaying properly.

MINIMAL Requirements: Google Chrome 24+Mozilla Firefox 20+Internet Explorer 11Opera 15–18Apple Safari 7SeaMonkey 2.15-2.23

The development of a narrow target gene panel makes next generation sequencing effective for circulating free DNA analysis

Date

10 Oct 2016

Session

Poster display

Presenters

Umberto Malapelle

Citation

Annals of Oncology (2016) 27 (6): 15-42. 10.1093/annonc/mdw363

Authors

U. Malapelle1, C. Mayo2, D. Rocco3, M. Garzon2, P. Pisapia4, R. Sgariglia4, C. De Luca4, N.J. Ariza2, F. Pepe4, D.M. Espinosa5, A.M. Bueno5, M. González-Cao6, N. Karachaliou7, S. Viteri5, M.A. Molina Vila2, R. Rosell5, G. Troncone4

Author affiliations

  • 1 Public Health, Azienda Ospedaliera Universitaria Policlinico Federico II-AOU Federico II, 80131 - Napoli/IT
  • 2 Laboratory Of Cellular And Molecular Biology, Pangaea Biotech SL, IOR Quirón-Dexeus University Institute, 08028 - Barcelona/ES
  • 3 Oncology, Azienda Ospedaliera Dei Colli-Monaldi, Napoli/IT
  • 4 Public Health, Azienda Ospedaliera Universitaria Policlinico Federico II-AOU Federico II, Napoli/IT
  • 5 Medical Oncology Service, Instituto Oncológico Dr Rosell (IOR), Hospital Universitario Quirón-Dexeus, 08028 - Barcelona/ES
  • 6 Medical Oncology, Intituto Oncológico Rosell, Quirón-Dexeus University Hospital, 08028 - Barcelona/ES
  • 7 Medical Oncology Service, Instituo Oncológico Dr Rosell (IOR), Hospital Universitario Quirón-Dexeus, 08028 - Barcelona/ES
More

Resources

Abstract 4034

Background

Since tissue is not always available, biomarkers testing, can be performed on circulating free DNA (cfDNA). Compared to real time PCR, next-generation sequencing (NGS), covers also less common and novel variants. To increase sensitivity NGS can be narrowed to target a limited number of actionable genes. This strategy, known as ultra-deep sequencing, requires a careful validation. Here we validated a narrowed gene panel to produce a DNA library covering 568 actionable mutations in six gene (EGFR, KRAS, NRAS, BRAF, cKIT and PDGFRα) involved in non small cell lung cancer, gastrointestinal stromal tumor, metastatic colo rectal carcinoma and melanoma (SiRe).

Methods

This study had a retrospective and prospective design. After in – vitro studies on cell lines aimed to assess the assay analytical performance, cfDNA from a retrospective series of cases including, lung (n = 51) and colon (n = 3) neoplasms and melanoma (n = 9) previously well characterized on both tissue and matched cfDNA was employed to validate the SiRe panel; then, blood samples prospectically collected from NSCLC patients (n = 87) were tested to assess this panel performance in daily clinical practice.

Results

On cell lines, the SiRe had high intra – and inter – run reproducibility with 0.2% lower limit of mutation detection. In the retrospective series of cfDNA, a total of 54 mutations were detected by SiRe showing 100% specificity, confirming 39 EGFR, 9 KRAS, 1 NRAS, 5 BRAF mutations previously detected on matched tissue. Noteworthy, in 4 cases SiRe detected mutations that had been missed on cfDNA by real time PCR. On prospectically collected cfDNA, SiRe detected in 4/46 patients, without baseline tissue availability, activating EGFR mutation; at the time of tumor progression following the treatment with gefitinib, erlotinib and afatinib SiRe detected T790M in 30% (9/30). On the overall, the SiRe panel showed a sensibility of 93.4% and specificity of 100%.

Conclusions

The SiRe panel is an effective tool enabling a cost - effective implementation of NGS for cfDNA mutational profiling in molecular pathology practice

Clinical trial identification

Legal entity responsible for the study

University of Naples Federico II, Department of Public Health

Funding

University of Naples Federico II, Department of Public Health.

Disclosure

All authors have declared no conflicts of interest.

Resources from the same session

This site uses cookies. Some of these cookies are essential, while others help us improve your experience by providing insights into how the site is being used.

For more detailed information on the cookies we use, please check our Privacy Policy.

Customise settings