Multicenter Evaluation of a Novel Automated Rapid Detection System of BRAF Status in Formalin-Fixed, Paraffin-Embedded Tissues

Abstract

The mutated BRAF oncogene represents a therapeutic target in malignant melanoma. Because BRAF mutations are also involved in the pathogenesis of other human malignancies, the use of specific BRAF inhibitors might also be extended to other diseases in the future. A prerequisite for the clinical application of BRAF inhibitors is the reliable detection of activating BRAF mutations in routine histopathological samples. In a multicenter approach, we evaluated a novel and fully automated PCR-based system (Idylla) capable of detecting BRAF V600 mutations in formalin-fixed, paraffin-embedded tissue within 90 minutes with high sensitivity. We analyzed a total of 436 samples with the Idylla system. Valid results were obtained in 421 cases (96.56%). Its performance was compared with conventional methods (pyrosequencing or Sanger sequencing). Concordant results were obtained in 406 cases (96.90%). Reanalysis of eight discordant samples by next-generation sequencing and/or pyrosequencing with newly extracted DNA and the BRAF RGQ Kit confirmed the Idylla result in seven cases, resulting in an overall agreement of 98.57%. In conclusion, the Idylla system is a highly reliable and sensitive platform for detection of BRAF V600 mutations in formalin-fixed, paraffin-embedded material, providing an efficient alternative to conventional diagnostic methods, particularly for routine diagnostics laboratories with limited experience in molecular pathology.

Rapid On‐site Molecular Evaluation in thyroid cytopathology: A same‐day cytological and molecular diagnosis

“…Many institutions all over the world have adopted this approach to rapidly assess the most common actionable oncogenes in a wide range of human neoplasms. [10][11][12][13][14][15][16][17][18][19][20] Acute deteriorating patients can be immediately treated and predictive biomarker testing can be carried out even where equipment and expertise are not available. 26 A recently developed combined NRAS-BRAF Mutation Test has only been applied to CRC and melanoma, to inform on targeted treatment response.…”

“…In any single case, the tissue block representative of the neoplasm was selected for the Idylla assay. The tissue area with the highest percentage of neoplastic cells (Table ) was marked on a hematoxylin and eosin (H&E) stained slide and microdissected on a corresponding FFPE section (10 μm thickness); then, the microdissected area was inserted into the Idylla cartridge and processed as previously described . The BRAF and NRAS genotyping results were compared with those obtained by processing the simulated FNA (Table ).…”

“…The (Table 1) was marked on a hematoxylin and eosin (H&E) stained slide and microdissected on a corresponding FFPE section (10 μm thickness); then, the microdissected area was inserted into the Idylla cartridge and processed as previously described. 10,[15][16][17][18][19][20] The BRAF and NRAS genotyping results were compared with those obtained by processing the simulated FNA ( Table 1).…”

Multi-center evaluation of the novel fully-automated PCR-based Idylla™ BRAF Mutation Test on formalin-fixed paraffin-embedded tissue of malignant melanoma

“…It is a fully-automated real-time-PCR-based test designed for the qualitative detection of the V600E/E2/D and V600K/R/M mutations, with high sensitivity (detection of 1% mutant in a background of wild type) and quick turnaround time (approximately 90 minutes) from FFPE sample to final result. 14,15 The single-use cartridge-based Idylla TM BRAF Mutation Test is performed directly on FFPE tissue sections, requiring no beforehand sample preparation and minimal hands-on time (less than 2 minutes per FFPE tissue section). The interpretation of results is fully automatic.…”

Can detection of Braf p.V600E mutation be improved? Comparison of allele specific multiplex sequencing to present tests

“…[4,5] In attempting to meet these objectives, methods with significantly increased sensitivities and specificities have been developed, including allele-specific PCR. [6][7][8][9][10][11][12][13] Although allele-specific PCR and ASMS both use allelespecific primers, they are two quite different methods. In allele-specific PCR, allele-specific primers are used in the amplification step.…”