Abstract
BackgroundAlterations in the highly penetrant cancer susceptibility gene BRCA1 are responsible for the majority of hereditary breast and/or ovarian cancers. However, the number of detected germline mutations has been lower than expected based upon genetic linkage data. Undetected deleterious mutations in the BRCA1 gene in some high-risk families could be due to the presence of intragenic rearrangements as deletions, duplications or insertions spanning whole exons. Standard PCR-based screening methods are mainly focused on detecting point mutations and small insertions/deletions, but large rearrangements might escape detection.The purpose of this study was to determine the type and frequency of large genomic rearrangements in the BRCA1 gene in hereditary breast and ovarian cancer cases in the Czech Republic.MethodsMultiplex ligation-dependent probe amplification (MLPA) was used to examine BRCA1 rearrangements in 172 unrelated patients with hereditary breast and/or ovarian cancer syndrome without finding deleterious mutation after complete screening of whole coding regions of BRCA1/2 genes. Positive MLPA results were confirmed and located by long-range PCR. The breakpoints of detected rearrangements were characterized by sequencing.ResultsSix different large deletions in the BRCA1 gene were identified in 10 out of 172 unrelated high-risk patients: exons 1A/1B and 2 deletion; partial deletion of exon 11 and exon 12; exons 18 and 19 deletion; exon 20 deletion; exons 21 and 22 deletion; and deletion of exons 5 to 14. The breakpoint junctions were localized and further characterized. Destabilization and global unfolding of the mutated BRCT domains explain the molecular and genetic defects associated with the exon 20 in-frame deletion and the exon 21 and 22 in-frame deletion, respectively.ConclusionUsing MLPA, mutations were detected in 6% of high-risk patients previously designated as BRCA1/2 mutation-negative. The breakpoints of five out of six large deletions detected in Czech patients are novel. Screening for large genomic rearrangements in the BRCA1 gene in the Czech high-risk patients is highly supported by this study.
Highlights
Alterations in the highly penetrant cancer susceptibility gene BRCA1 are responsible for the majority of hereditary breast and/or ovarian cancers
Screening for large genomic rearrangements in the BRCA1 gene in the Czech high-risk patients is highly supported by this study
The inclusion criteria were as follow: (i) unrelated index patients affected by invasive breast and/or ovarian cancer, (ii) at least three diagnoses of breast and/ or ovarian cancer in the family diagnosed at any age, and (iii) no deleterious mutation found during complete screening of the whole coding regions of BRCA1 and BRCA2 genes as described by Foretova et al [28]
Summary
Alterations in the highly penetrant cancer susceptibility gene BRCA1 are responsible for the majority of hereditary breast and/or ovarian cancers. Undetected deleterious mutations in the BRCA1 gene in some high-risk families could be due to the presence of intragenic rearrangements as deletions, duplications or insertions spanning whole exons. The purpose of this study was to determine the type and frequency of large genomic rearrangements in the BRCA1 gene in hereditary breast and ovarian cancer cases in the Czech Republic. Germline mutations in BRCA1 (OMIM#113705, Online Mendelian Inheritance in Man) and BRCA2 (OMIM#600185) genes are responsible for an important fraction of hereditary breast and ovarian cancers [2]. Pathogenic mutations in the coding region or in splice sites of the BRCA1 gene were found in approximately twothirds of BRCA1-linked families [2] This finding suggests that methods generally used for mutation scanning fail to detect certain types of BRCA1 germline defects, such as large intragenic rearrangements. Multiplex ligation-dependent probe amplification (MLPA) has been widely used as a highly sensitive method for detecting the relative copy number of all BRCA1 exons in a high-throughput format [16]
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