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A combination of immunohistochemistry and molecular approaches improves highly sensitive detection of BRAF mutations in papillary thyroid cancer

  • Endocrine Methods and Techniques
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Abstract

The optimal method for BRAF mutation detection remains to be determined despite advances in molecular detection techniques. The aim of this study was to compare, against classical Sanger sequencing, the diagnostic performance of two of the most recently developed, highly sensitive methods: BRAF V600E immunohistochemistry (IHC) and peptide nucleic-acid (PNA)-clamp qPCR. BRAF exon 15 mutations were searched in formalin-fixed paraffin-embedded tissues from 86 papillary thyroid carcinoma using the three methods. The limits of detection of Sanger sequencing in borderline or discordant cases were quantified by next generation sequencing. BRAF mutations were found in 74.4 % of cases by PNA, in 71 % of cases by IHC, and in 64 % of cases by Sanger sequencing. Complete concordance for the three methods was observed in 80 % of samples. Better concordance was observed with the combination of two methods, particularly PNA and IHC (59/64) (92 %), while the combination of PNA and Sanger was concordant in 55 cases (86 %). Sensitivity of the three methods was 99 % for PNA, 94.2 % for IHC, and 89.5 % for Sanger. Our data show that IHC could be used as a cost-effective, first-line method for BRAF V600E detection in daily practice, followed by PNA analysis in negative or uninterpretable cases, as the most efficient method. PNA-clamp quantitative PCR is highly sensitive and complementary to IHC as it also recognizes other mutations besides V600E and it is suitable for diagnostic purposes.

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Abbreviations

PNA-clamp qPCR:

Nucleic acid clamp quantitative polymerase chain reaction

IHC:

Immunohistochemistry

NGS:

Next generation sequencing

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    Authors and Affiliations

    1. Department of Internal Medicine and Medical Specialties (DiMI), University of Genoa, V.le Benedetto XV 6, 16132, Genoa, Italy

      Claudia Martinuzzi, Lorenza Pastorino, Virginia Andreotti, Anna Garuti, Giovanna Bianchi-Scarrà & Paola Ghiorzo

    2. Department of Surgical Sciences (DISC), University of Genoa, Genoa, Italy

      Claudia Martinuzzi, Michele Minuto, Roberto Fiocca, Federica Grillo & Luca Mastracci

    3. Surgery 1 Unit, IRCCS AOU San Martino-IST, Genoa, Italy

      Michele Minuto

    4. Pathology Unit, IRCCS AOU San Martino-IST, Genoa, Italy

      Roberto Fiocca, Federica Grillo & Luca Mastracci

    5. Genetics of Rare Tumors, IRCCS AOU San Martino-IST, University of Genoa, Genoa, Italy

      Giovanna Bianchi-Scarrà & Paola Ghiorzo

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    Martinuzzi, C., Pastorino, L., Andreotti, V. et al. A combination of immunohistochemistry and molecular approaches improves highly sensitive detection of BRAF mutations in papillary thyroid cancer. Endocrine 53, 672–680 (2016). https://doi.org/10.1007/s12020-015-0720-9

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