title: 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.
authors.affiliation: Department of Genetics, Portuguese Oncology Institute, Porto, Portugal.
authors.forename: Catarina
authors.initials: C
authors.lastname: Santos
authors.affiliation: Department of Genetics, Portuguese Oncology Institute, Porto, Portugal.
authors.forename: Ana
authors.initials: A
authors.lastname: Peixoto
authors.affiliation: Department of Genetics, Portuguese Oncology Institute, Porto, Portugal.
authors.forename: Patrícia
authors.initials: P
authors.lastname: Rocha
authors.affiliation: Department of Genetics, Portuguese Oncology Institute, Porto, Portugal.
authors.forename: Pedro
authors.initials: P
authors.lastname: Pinto
authors.affiliation: Department of Genetics, Portuguese Oncology Institute, Porto, Portugal.
authors.forename: Susana
authors.initials: S
authors.lastname: Bizarro
authors.affiliation: Department of Genetics, Portuguese Oncology Institute, Porto, Portugal.
authors.forename: Manuela
authors.initials: M
authors.lastname: Pinheiro
authors.affiliation: Department of Genetics, Portuguese Oncology Institute, Porto, Portugal.
authors.forename: Carla
authors.initials: C
authors.lastname: Pinto
authors.affiliation: Department of Pathology, Portuguese Oncology Institute, Porto, Portugal; Biomedical Sciences Institute (ICBAS), University of Porto, Porto, Portugal.
authors.forename: Rui
authors.initials: R
authors.lastname: Henrique
authors.affiliation: Department of Genetics, Portuguese Oncology Institute, Porto, Portugal; Biomedical Sciences Institute (ICBAS), University of Porto, Porto, Portugal. Electronic address: manuel.teixeira@ipoporto.min-saude.pt.
authors.forename: Manuel R
authors.initials: MR
authors.lastname: Teixeira
mesh_terms: D019313:BRCA1 Protein / Q000235:genetics*; D024682:BRCA2 Protein / Q000235:genetics*; D001940:Breast / Q000378:metabolism / Q000473:pathology; D001943:Breast Neoplasms / Q000175:diagnosis / Q000453:epidemiology / Q000235:genetics; D005260:Female; D005820:Genetic Testing / Q000379:methods; D014644:Genetic Variation; D006801:Humans; D010051:Ovarian Neoplasms / Q000175:diagnosis / Q000453:epidemiology / Q000235:genetics*; D010053:Ovary / Q000378:metabolism / Q000473:pathology; D011174:Portugal / Q000453:epidemiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012984:Software; D035501:Uncertainty
chemical_list: D019313:BRCA1 Protein; D024682:BRCA2 Protein