Pathogenicity Evaluation of BRCA1 and BRCA2 Unclassified Variants Identified in Portuguese Breast/Ovarian Cancer Families

Abstract

Hereditary breast/ovarian cancer syndrome is caused by germline deleterious mutations in BRCA1 and BRCA2. A major problem of genetic testing and counseling is the finding of variants of uncertain significance (VUS). We sought to ascertain the pathogenicity of 25 BRCA1 and BRCA2 VUS identified in Portuguese families during genetic testing. We performed cosegregation analysis of VUS with cancer in families, evaluated their frequency in unaffected controls, and looked for loss of heterozygosity in tumors. In addition, three different bioinformatic algorithms were used (Interactive Biosoftware, ESEfinder, and PolyPhen). Finally, six VUS located in exon-intron boundaries were analyzed by RT-PCR. We found that seven variants segregated with the disease, six variants co-occurred with a pathogenic mutation in the same gene, and four variants co-occurred with a deleterious mutation in the other BRCA gene. By RT-PCR, we observed that four variants (BRCA1 c.4484G>T, BRCA2 c.682-2A>C, BRCA2 c.8488-1G>A, and BRCA2 c.8954-5A>G) disrupted splicing. After the combined analysis, we were able to classify 4 splicing variants as pathogenic mutations, 16 variants as neutral, and 3 variants as polymorphisms; only 2 variants remained classified as VUS. This work highlights the contribution of DNA, RNA, and in silico data to assess the pathogenicity of BRCA1/2 VUS, which, in turn, allows more accurate genetic counseling and clinical management of the families carrying them.

The role of targeted BRCA1/BRCA2 mutation analysis in hereditary breast/ovarian cancer families of Portuguese ancestry

“…Thus, 119 pathogenic mutations were detected in total, of which 41.2% occurred in BRCA1 and 58.8% in BRCA2 . We also identified 70 different VUS, of which two ( BRCA1 c.5122G>C; BRCA2 c.2T>G) have recently been studied in more detail .…”

“…Thus, 119 pathogenic mutations were detected in total, of which 41.2% occurred in BRCA1 and 58.8% in BRCA2. We also identified 70 different VUS, of which two (BRCA1 c.5122G>C; BRCA2 c.2T>G) have recently been studied in more detail (7). Two mutations in BRCA1 (c.2037delinsCC and c.3331_3334del) and one in BRCA2 (c.156_157insAlu) together represent about 50% of all deleterious mutations found in Portuguese HBOC families.…”

“…Seventy different VUS (26 and 44 different variants in BRCA1 and BRCA2 genes, respectively) were identified, of which seven were detected in probands with BRCA1/BRCA2 pathogenic mutations ( Table 2). Nineteen variants were previously classified as either neutral or polymorphisms (7) and were therefore not included here.…”

BRCA Share: A Collection of Clinical BRCA Gene Variants

“…Co-occurrence of a VUS with a known disease-causing mutation, either from the same or the other BRCA gene, can provide evidence to classify the variant, especially if it is reported in multiple patients [Goldgar et al, 2008;Cherbal et al, 2012;Santos et al, 2014]. Cooccurrence data are used in two ways: in a Posterior Probability calculation [Goldgar et al, 2004], as it is one of its elements, and as a standalone criterion for classification.…”

Novel Mutations in BMPR2, ACVRL1 and KCNA5 Genes and Hemodynamic Parameters in Patients with Pulmonary Arterial Hypertension

“…We detected 53 different nucleotide changes in BMPR2, ACVRL1 and KCNA5 genes in 40 out of 41 PAH patients. Twenty two of these mutations (found in 51% of patients) were considered pathogenic according to the in silico analysis that was performed with several programs to reach a high reliability [21].…”