Functional assays for analysis of variants of uncertain significance in BRCA2

Abstract

Missense variants in the BRCA2 gene are routinely detected during clinical screening for pathogenic mutations in patients with a family history of breast and ovarian cancer. These subtle changes frequently remain of unknown clinical significance because of the lack of genetic information that may help establish a direct correlation with cancer predisposition. Therefore, alternative ways of predicting the pathogenicity of these variants are urgently needed. Since BRCA2 is a protein involved in important cellular mechanisms such as DNA repair, replication, and cell cycle control, functional assays have been developed that exploit these cellular activities to explore the impact of the variants on protein function. In this review, we summarize assays developed and currently utilized for studying missense variants in BRCA2. We specifically depict details of each assay, including variants of uncertain significance analyzed, and describe a validation set of (genetically) proven pathogenic and neutral missense variants to serve as a golden standard for the validation of each assay. Guidelines are proposed to enable implementation of laboratory-based methods to assess the impact of the variant on cancer risk.

Keywords: BRCA2; VUS; breast cancer; functional analysis; genetic testing; ovarian cancer; variants of uncertain significance.

Figure 1

Landscape of classified BRCA2 variants evaluated by functional assays. Schematic representation of BRCA2 protein and functional domains showing the approximate location of Class 1, 4 and 5 missense variants that have been analyzed in one or more functional assays and classified by the posterior probability model (Plon et al., 2008; Lindor et al., 2012). Class 1 or neutral variants are shown in green; Class 4/5 or pathogenic and likely pathogenic mutations are shown in red. H represents the helical domain; OB represents the oligonucleotide/oligosaccharide-binding fold; NLS depicts the nuclear localization signals.