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Variants of uncertain clinical significance in hereditary breast and ovarian cancer genes: best practices in functional analysis for clinical annotation
  1. Alvaro N Monteiro1,
  2. Peter Bouwman2,
  3. Arne N Kousholt2,
  4. Diana M Eccles3,
  5. Gael A Millot4,
  6. Jean-Yves Masson5,
  7. Marjanka K Schmidt2,
  8. Shyam K Sharan6,
  9. Ralph Scully7,
  10. Lisa Wiesmüller8,
  11. Fergus Couch9,
  12. Maaike P G Vreeswijk10
  1. 1 Cancer Epidemiology, H. Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA
  2. 2 Division of Molecular Pathology, Oncode Institute, The Netherlands Cancer Institute, Amsterdam, The Netherlands
  3. 3 Cancer Sciences, University of Southampton Faculty of Medicine, Southampton, UK
  4. 4 Hub-DBC, Institut Pasteur, USR 3756 CNRS, Paris, France
  5. 5 CHU de Québec-Université Laval, Oncology Division, Laval University Cancer Research Center, Quebec City, Quebec, Canada
  6. 6 National Cancer Institute at Frederick, Frederick, Maryland, USA
  7. 7 Beth Israel Deaconess Medical Center, Boston, Massachusetts, USA
  8. 8 Ulm University, Ulm, Baden-Württemberg, Germany
  9. 9 Mayo Clinic, Rochester, Minnesota, USA
  10. 10 Human Genetics, Leiden University Medical Center, Leiden, The Netherlands
  1. Correspondence to Dr Alvaro N Monteiro, Cancer Epidemiology, H. Lee Moffitt Cancer Center and Research Institute, Tampa, FL 33612, USA; Alvaro.Monteiro{at}; Dr Maaike P G Vreeswijk, Human Genetics, Leiden University Medical Center, Leiden, 2300RC, The Netherlands; M.P.G.Vreeswijk{at}

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Since the identification and cloning of BRCA1 in 1994,1 and shortly thereafter of BRCA2,2 genetic tests of germline DNA to identify pathogenic variants in genes linked to hereditary breast and ovarian cancer (HBOC) have become mainstream.3 These tests are critical to identify women at increased risk relative to the general population. Women at moderate risk (2≤relative risk (RR) <4) may benefit from enhanced screening and chemoprevention while those at high risk (RR >4), including those with BRCA1 and BRCA2 pathogenic variants, may also benefit from preventive surgery. Germline mutation testing is also becoming increasingly relevant in the cancer treatment setting because carriers of pathogenic variants in BRCA1/2 may benefit from poly-ADP ribose polymerase (PARP) inhibitors.4 5 Importantly, genetic tests can identify individuals in HBOC families who do not carry the relevant predisposing allele and are not at elevated risk of cancer.6

A significant fraction of documented variants in BRCA1 and BRCA2 are considered variants of uncertain clinical significance (VUS), for which cancer association has not been assessed or could not be determined due to insufficient information (table 1). In ClinVar (, a clinically oriented database, currently ~37% of BRCA1 and ~45% of BRCA2 unique variants recorded are VUS. Thus, there is a critical need to classify variants according to their pathogenicity.

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Table 1

Fraction of VUS in BRCA1 and BRCA2

Over the past decade, functional assays have emerged that can be included as a source of evidence to classify variants according to their pathogenicity, with the potential to greatly accelerate classification.7 Here, we discuss several technical and conceptual aspects relevant for the use of functional assays in the classification of variants. We present best practice recommendations to improve annotation quality and accuracy, and to provide a basis for the comparison and integration of functional data …

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  • Contributors This manuscript emerged from discussions held at a Netherlands Cancer Institute (NKI) workshop on Functional Analysis of Sequence Variants in Hereditary Breast and Ovarian Cancer Genes (Amsterdam, The Netherlands). AM and MV planned the study and coordinated the discussions and are responsible for the overall content as guarantors. All authors contributed to the discussions, to the preparation and editing of the manuscript, reviewed and approved the final manuscript.

  • Funding The meeting was supported by funds from the Cancer Genomics Center Netherlands through the Netherlands Organization for Scientific Research (NWO), the Royal Netherlands Academy of Arts and Sciences (KNAW), the Danish Counsil for Independent Research (DFF), the Netherlands Cancer Institute and from IDT and BIOKÉ.

  • Competing interests LW is an inventor and owner of a patent on a test system for determining genotoxicities and cancer risk.

  • Patient consent for publication Not required.

  • Provenance and peer review Not commissioned; externally peer reviewed.