Mouse embryonic stem cell–based functional assay to evaluate mutations in BRCA2

Abstract

Kuznetsov and his colleagues address a pressing problem in risk assessment for predisposition to breast cancer—whether a particular allele is cancer predisposing or not. Using a two-tiered approach, they have developed a functional assay for the classification of BRCA2 sequence variants of unknown importance. The assay may serve as a model to generate functional assays for other human disease genes. Individuals with mutations in breast cancer susceptibility genes BRCA1 and BRCA2 have up to an 80% risk of developing breast cancer by the age of 70. Sequencing-based genetic tests are now available to identify mutation carriers in an effort to reduce mortality through prevention and early diagnosis. However, lack of a suitable functional assay hinders the risk assessment of more than 1,900 BRCA1 and BRCA2 variants in the Breast Cancer Information Core database that do not clearly disrupt the gene product. We have established a simple, versatile and reliable assay to test for the functional significance of mutations in BRCA2 using mouse embryonic stem cells (ES cells) and bacterial artificial chromosomes and have used it to classify 17 sequence variants. The assay is based on the ability of human BRCA2 to complement the loss of endogenous Brca2 in mouse ES cells. This technique may also serve as a paradigm for functional analysis of mutations found in other genes linked to human diseases.

Cite this article

Kuznetsov, S., Liu, P. & Sharan, S. Mouse embryonic stem cell–based functional assay to evaluate mutations in BRCA2. Nat Med 14, 875–881 (2008). https://doi.org/10.1038/nm.1719

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