title: Comparative evaluation of the new FDA approved THxID™-BRAF test with High Resolution Melting and Sanger sequencing.
abstract: BACKGROUND
  Since patients diagnosed with BRAF V600E and V600K mutated advanced melanoma show response to treatment with MAP kinase inhibitors, several sensitive methods have been developed to determine the V600 allele status of melanoma patients. Vemurafenib (Zelboraf) and dabrafenib (Tafinlar) are specific BRAF V600 inhibitors recently approved by the US FDA as single agent treatments for unresectable or metastatic melanoma in patients with the BRAF V600 mutation.
  
  
  METHODS
  We assessed the new CE THxID™-BRAF diagnostic test, which is also FDA-approved as a companion diagnostic test in the US under a specific reference and compared the results of this assay with both High Resolution Melting (HRM) and Sanger sequencing in 113 melanoma FFPE samples.
  
  
  RESULTS
  Invalid results were observed in 0/113 specimen with HRM, 5/113 (4.4%) with Sanger sequencing, and 1/113 (0.9%) with the THxID™-BRAF test. Positive percentage agreement (PPA) was 93.5% (95% CI 82.5 - 97.8) for V600E and V600K mutations combined for the THxID™-BRAF test and HRM, and negative percentage agreement (NPA) was 100.0% (95% CI 94.5 - 100.0). For the THxID™-BRAF test and Sanger, PPA was 100.0% (95% CI 92.1 - 100.0) and NPA 100.0% (95% CI 94.2 - 100.0). One V600E sample identified by THxID™-BRAF test was detected as wild-type by HRM and uninterpretable by Sanger. All V600K (n = 3) were detected using the 3 different approaches. Finally, percent agreement values were not significantly different when using punches (n = 77) vs. slides (n = 36) or depending on samples characteristics such as pigmentation, necrosis, and tumor content.
  
  
  CONCLUSIONS
  This study demonstrated the high agreement between the FDA approved THxID™-BRAF assay, HRM, and Sanger sequencing. It has also highlighted the potential of THxID™-BRAF to be applied to a broader range of sample types than claimed in the current "instructions for use", an extension that would require the ad hoc validation and approval.
authors.forename: Julie
authors.initials: J
authors.lastname: Marchant
authors.forename: Alain
authors.initials: A
authors.lastname: Mange
authors.forename: Marion
authors.initials: M
authors.lastname: Larrieux
authors.forename: Valérie
authors.initials: V
authors.lastname: Costes
authors.affiliation: Department of Biopathology, Center Hospital University, Avenue du Doyen Giraud, 34298 Montpellier Cedex 5, France. j-solassol@chu-montpellier.fr.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Solassol
mesh_terms: D005260:Female; D006801:Humans; D008297:Male; D008545:Melanoma / Q000175:diagnosis* / Q000235:genetics; D008875:Middle Aged; D009154:Mutation; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA / Q000379:methods*; D014481:United States; D014486:United States Food and Drug Administration
chemical_list: D011933:Reagent Kits, Diagnostic; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf