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NGS-based BRCA1/2 mutation testing of high-grade serous ovarian cancer tissue: results and conclusions of the first international round robin trial

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Abstract

With the approval of olaparib as monotherapy treatment in platinum-sensitive, relapsed high-grade serous ovarian cancer by the European Medical Agency (EMA), comprehensive genotyping of BRCA1 and BRCA2 in tumor tissue has become a mandatory pre-therapeutic test. This requires significant advances in routine tumor test methodologies due to the large size of both genes and the lack of mutational hot spots. Classical focused screening approaches, like Sanger sequencing, do not allow for a sensitive, rapid, and economic analysis of tumor tissue. Next-generation sequencing (NGS) approaches employing targeted panels for BRCA1/2 to interrogate formalin-fixed and paraffin-embedded tumor samples from either surgical resection or biopsy specimens can overcome these limitations. Although focused NGS methods have been implemented by few centers in routine molecular diagnostics for the analysis of some druggable oncogenic mutations, the reliable diagnostic testing of the entire coding regions of BRCA1 and BRCA2 was a new challenge requiring extensive technological improvement and quality management. Here, we describe the implementation and results of the first round robin trial for BRCA1/2 mutation testing in tumor tissue that was conducted in central Europe on May 2015, shortly after the approval and prior to the official release of olaparib. The high success rate of 81 % (21/26 test centers) demonstrates that BRCA1/2 multicenter mutation testing is well feasible in FFPE tumor tissue, extending to other tumor entities beyond ovarian cancer. The high number of test centers passing the trial demonstrates the success of the concerted efforts by German, Swiss, and Austrian pathology centers to ensure quality-controlled NGS-based testing and proves the potential of this technology in routine molecular pathology. On the basis of our results, we provide recommendations for predictive testing of tumor tissue for BRCA1/2 to clinical decision making in ovarian cancer patients.

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Author notes

  1. Nicole Pfarr & Wilko Weichert

    Present address: Institute of Pathology, Technical University of Munich (TUM), Munich, Germany

Authors and Affiliations

  1. Institute of Pathology, University Hospital Heidelberg, Im Neuenheimer Feld, 224, Heidelberg, Germany

    Volker Endris, Albrecht Stenzinger, Nicole Pfarr, Roland Penzel, Wilko Weichert & Peter Schirmacher

  2. Institute of Pathology, Charité Universitaetsmedizin Berlin, Berlin, Germany

    Markus Möbs, Dido Lenze, Silvia Darb-Esfahani, Michael Hummel & Manfred Dietel

  3. Institute of Pathology, University of Cologne, Cologne, Germany

    Sabine-Merkelbach-Bruse & Reinhard Büttner

  4. Institute of Pathology, Ludwig-Maximilians University (LMU), Munich, Germany

    Andreas Jung & Thomas Kirchner

  5. Institute of Pathology, Hannover Medical School, Hannover, Germany

    Ulrich Lehmann & Hans Kreipe

  6. Institute of Pathology, Kantonsspital St. Gallen, St. Gallen, Switzerland

    Wolfram Jochum

  7. Institute of Pathology, Medical University Graz, Graz, Austria

    Gerald Höfler

Authors
  1. Volker Endris
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Corresponding author

Correspondence to Peter Schirmacher.

Ethics declarations

Usage of the tissue was approved by the local ethics committee in Heidelberg (No. 206/2005) and Berlin (2013 amendment of No. EA1 139/05). In compliance with the current version of the German Gene Law, for none of the cases, the specific germline status of BRCA1/2 was determined.

Conflict of interest

VE, SMB, AJ, HK, TK, RB, SDE, MD, WW, and PS are advisory board members of Astra Zeneca. VE, MD, SDE, WW, TK, and PS received honoraria from Astra Zeneca for serving as speakers.

Electronic supplementary material

Supplementary Table 1

Packaging schemes for the test samples that were sent to each participating test center of the phase II test (XLSX 12 kb)

Supplementary Table 2

Detailed overview of genotyping results of the phase I trial for each test center. Somatic mutations including IARC score and allele frequencies are provided for each tumor sample (XLSX 12 kb)

Supplementary Table 3

Detailed overview of genotyping results of the phase II trial for each test center. Somatic mutations including IARC score and allele frequencies are provided for each tumor sample (XLSX 98 kb)

Supplementary Table 4

Overview of different analyses and annotation softwares used in the round robin trial (where available) (XLSX 10 kb)

Supplementary Fig. 1

Allelic frequencies and coverage of BRCA1/2 mutations across all test centers of the phase II test. Data are visualized as boxplots, upper and lower quartiles interquartile range and mean. Whiskers indicate outliers. a Reported allelic frequencies per mutation. b Reported depth of coverage per mutation (GIF 95 kb)

High resolution image (TIFF 726 kb)

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Endris, V., Stenzinger, A., Pfarr, N. et al. NGS-based BRCA1/2 mutation testing of high-grade serous ovarian cancer tissue: results and conclusions of the first international round robin trial. Virchows Arch 468, 697–705 (2016). https://doi.org/10.1007/s00428-016-1919-8

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  • DOI: https://doi.org/10.1007/s00428-016-1919-8

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