Performance of in silico prediction tools for the classification of rare BRCA1/2 missense variants in clinical diagnostics

BMC Med Genomics. 2018 Mar 27;11(1):35. doi: 10.1186/s12920-018-0353-y.


Background: The use of next-generation sequencing approaches in clinical diagnostics has led to a tremendous increase in data and a vast number of variants of uncertain significance that require interpretation. Therefore, prediction of the effects of missense mutations using in silico tools has become a frequently used approach. Aim of this study was to assess the reliability of in silico prediction as a basis for clinical decision making in the context of hereditary breast and/or ovarian cancer.

Methods: We tested the performance of four prediction tools (Align-GVGD, SIFT, PolyPhen-2, MutationTaster2) using a set of 236 BRCA1/2 missense variants that had previously been classified by expert committees. However, a major pitfall in the creation of a reliable evaluation set for our purpose is the generally accepted classification of BRCA1/2 missense variants using the multifactorial likelihood model, which is partially based on Align-GVGD results. To overcome this drawback we identified 161 variants whose classification is independent of any previous in silico prediction. In addition to the performance as stand-alone tools we examined the sensitivity, specificity, accuracy and Matthews correlation coefficient (MCC) of combined approaches.

Results: PolyPhen-2 achieved the lowest sensitivity (0.67), specificity (0.67), accuracy (0.67) and MCC (0.39). Align-GVGD achieved the highest values of specificity (0.92), accuracy (0.92) and MCC (0.73), but was outperformed regarding its sensitivity (0.90) by SIFT (1.00) and MutationTaster2 (1.00). All tools suffered from poor specificities, resulting in an unacceptable proportion of false positive results in a clinical setting. This shortcoming could not be bypassed by combination of these tools. In the best case scenario, 138 families would be affected by the misclassification of neutral variants within the cohort of patients of the German Consortium for Hereditary Breast and Ovarian Cancer.

Conclusion: We show that due to low specificities state-of-the-art in silico prediction tools are not suitable to predict pathogenicity of variants of uncertain significance in BRCA1/2. Thus, clinical consequences should never be based solely on in silico forecasts. However, our data suggests that SIFT and MutationTaster2 could be suitable to predict benignity, as both tools did not result in false negative predictions in our analysis.

Keywords: BRCA; Classification; Missense variant; Prediction tools; Variant of uncertain significance.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • BRCA1 Protein / genetics*
  • BRCA2 Protein / genetics*
  • Breast Neoplasms / diagnosis
  • Breast Neoplasms / genetics
  • Computational Biology / methods*
  • Computer Simulation*
  • Female
  • Humans
  • Molecular Diagnostic Techniques*
  • Mutation, Missense*
  • Ovarian Neoplasms / diagnosis
  • Ovarian Neoplasms / genetics


  • BRCA1 Protein
  • BRCA1 protein, human
  • BRCA2 Protein
  • BRCA2 protein, human