Abstract P4-10-02: Targeted resequencing of BRCA1 and BRCA2 in inherited breast cancer
Authors: Wong-Brown MW , Li S , Wilkins M , Avery-Kiejda KA , Bowden NA , Scott RJ 
Published: August 2014

Abstract
Breast cancer is the leading cause of cancer-related deaths in women worldwide, affecting about 13,000 women every year in Australia. Inherited loss-of-function mutations in BRCA1 and BRCA2 predispose to high risk of breast and/or ovarian cancer. Since the discovery of breast cancer susceptibility genes BRCA1 and BRCA2 two decades ago, there have not been any other genes identified that play a significant role in predisposition to inherited breast cancer. A large proportion of individuals with inherited breast cancer are negative for BRCA mutations and despite numerous research efforts, further breast cancer susceptibility genes still remain elusive.
We hypothesize that genetic anomalies are present in the BRCA1 and BRCA2 genes in a subset of individuals with inherited breast cancer where no genetic anomalies where identified using traditional Sanger sequencing. This study aims were to identify genetic anomalies in BRCA1 and BRCA2 by completely re-sequencing 200 kilobases surrounding BRCA1 and BRCA2 using targeted massively parallel sequencing, or next-generation sequencing.
For this study, DNA was used from 10 individuals referred for genetic testing after meeting the criteria for inherited breast cancer, and had been screened for BRCA1 and BRCA2 mutations by the Hunter Area Pathology Service (Newcastle, NSW, Australia). All individuals used for this study did not harbour causative genetic changes in the coding regions of BRCA1 or BRCA2. Targeted next-generation paired-end sequencing of regions containing BRCA1 and BRCA2 was performed using Agilent SureSelect and an Illumina GAIIx (The Ramaciotti Centre for Gene Function Analysis). An average of 50x coverage was achieved across the targeted genomic region for all samples. The sequence data was aligned to the Human Reference Sequence 37.2. Single nucleotide polymorphisms present in dbSNP or the 1000 genomes project were removed from further analyses. Genetic differences in the form of single nucleotide variants (SNVs) and insertions/deletions (indels) were identified in most individuals tested in regions that had previously remained unexplored, such as the non-coding regions of BRCA1 and BRCA2, the 5′ UTR and promoter sites. Structural variations including large deletions, duplications, insertions, inversions and rearrangements were also investigated, but require further analysis and validation for confirmation.
This study has comprehensively investigated BRCA1 and BRCA2 and surrounding genomic regions in a mutation negative inherited breast cancer population. The issue of accuracy of mutation detection by traditional methods, such as Sanger sequencing alone, has also been addressed by this study. The outcome of this study is the increase in current knowledge of the genetic variations that results in the development and/or progression of inherited breast cancer, aid in the management of individuals with breast cancers by providing a more specific diagnosis of disease risk and provide information required for the development of personalized treatment.
Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P4-10-02.