Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN 55905, USA.
Department of Biomedical Engineering and Institute for Computational Medicine, Johns Hopkins University, Baltimore, MD 21205, USA.
Am J Hum Genet. 2018 Feb 1;102(2):233-248. doi: 10.1016/j.ajhg.2017.12.013. Epub 2018 Jan 25.
Many variants of uncertain significance (VUS) have been identified in BRCA2 through clinical genetic testing. VUS pose a significant clinical challenge because the contribution of these variants to cancer risk has not been determined. We conducted a comprehensive assessment of VUS in the BRCA2 C-terminal DNA binding domain (DBD) by using a validated functional assay of BRCA2 homologous recombination (HR) DNA-repair activity and defined a classifier of variant pathogenicity. Among 139 variants evaluated, 54 had ?99% probability of pathogenicity, and 73 had ?95% probability of neutrality. Functional assay results were compared with predictions of variant pathogenicity from the Align-GVGD protein-sequence-based prediction algorithm, which has been used for variant classification. Relative to the HR assay, Align-GVGD significantly (p < 0.05) over-predicted pathogenic variants. We subsequently combined functional and Align-GVGD prediction results in a Bayesian hierarchical model (VarCall) to estimate the overall probability of pathogenicity for each VUS. In addition, to predict the effects of all other BRCA2 DBD variants and to prioritize variants for functional studies, we used the endoPhenotype-Optimized Sequence Ensemble (ePOSE) algorithm to train classifiers for BRCA2 variants by using data from the HR functional assay. Together, the results show that systematic functional assays in combination with in silico predictors of pathogenicity provide robust tools for clinical annotation of BRCA2 VUS.
通过临床基因检测，已在 BRCA2 中发现了许多意义不明的变体 (VUS)。VUS 带来了重大的临床挑战，因为这些变体对癌症风险的贡献尚未确定。我们通过使用经过验证的 BRCA2 同源重组 (HR) DNA 修复活性的功能测定法，对 BRCA2 C 末端 DNA 结合域 (DBD) 中的 VUS 进行了全面评估，并定义了一种变体致病性分类器。在评估的 139 种变体中，有 54 种具有?99%的致病性概率，73 种具有?95%的中性概率。功能测定法的结果与基于 Align-GVGD 蛋白序列的预测算法（用于变体分类）对变体致病性的预测进行了比较。与 HR 测定法相比，Align-GVGD 显著（p < 0.05）高估了致病性变体。随后，我们在贝叶斯层次模型（VarCall）中将功能和 Align-GVGD 预测结果相结合，以估计每个 VUS 的总体致病性概率。此外，为了预测所有其他 BRCA2 DBD 变体的影响并为功能研究优先考虑变体，我们使用了内表型优化序列集（ePOSE）算法，通过 HR 功能测定法的数据来训练 BRCA2 变体的分类器。总之，这些结果表明，系统的功能测定法与致病性的计算预测器相结合，为 BRCA2 VUS 的临床注释提供了强大的工具。
