BRCA somatic and germline mutation detection in paraffin embedded ovarian cancers by next-generation sequencing

Oncotarget. 2016 Jan 12;7(2):1076-83. doi: 10.18632/oncotarget.6834.


BRCA mutated ovarian cancers respond better to platinum-based therapy and to the recently approved PARP-inhibitors. There is the need for efficient and timely methods to detect both somatic and germline mutations using formalin-fixed paraffin-embedded (FFPE) tissues and commercially available technology. We used a commercial kit exploring all exons and 50bp exon-intron junctions of BRCA1 and BRCA2 genes, and semiconductor next-generation sequencing (NGS) on DNA from 47 FFPE samples of high-grade serous ovarian cancers. Pathogenic mutations were found in 13/47 (28%) cancers: eight in BRCA1 and five in BRCA2. All BRCA1 and two BRCA2 mutations were germline; three BRCA2 mutations were somatic. All mutations were confirmed by Sanger sequencing. To evaluate the performance of the NGS panel, we assessed its capability to detect the 6,953 variants described for BRCA1 and BRCA2 in ClinVar and COSMIC databases using callability analysis. 6,059 (87.1%) variants were identified automatically by the software; 829 (12.0%) required visual verification. The remaining 65 (0.9%) variants were uncallable, and would require 15 Sanger reactions to be resolved. Thus, the sensitivity of the NGS-panel was 99.1%. In conclusion, NGS performed with a commercial kit is highly efficient for detection of germline and somatic mutations in BRCA genes using routine FFPE tissue.

Keywords: BRCA1-BRCA2; PARP inhibitor; next generation sequencing; olaparib; ovarian carcinoma.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aged
  • BRCA1 Protein / genetics*
  • BRCA2 Protein / genetics*
  • DNA, Neoplasm / chemistry
  • DNA, Neoplasm / genetics
  • Exons / genetics
  • Female
  • Genetic Testing / methods
  • Germ-Line Mutation
  • High-Throughput Nucleotide Sequencing / methods*
  • Humans
  • Introns / genetics
  • Middle Aged
  • Mutation*
  • Ovarian Neoplasms / diagnosis
  • Ovarian Neoplasms / genetics*
  • Paraffin Embedding
  • RNA Splice Sites / genetics
  • Reagent Kits, Diagnostic
  • Reproducibility of Results
  • Sensitivity and Specificity


  • BRCA1 Protein
  • BRCA1 protein, human
  • BRCA2 Protein
  • BRCA2 protein, human
  • DNA, Neoplasm
  • RNA Splice Sites
  • Reagent Kits, Diagnostic