Novel mutations in the BRCA1 and BRCA2 genes in Iranian women with early-onset breast cancer

Abstract

Background: Breast cancer is the most common female malignancy and a major cause of death in middle-aged women. So far, germline mutations in the BRCA1 and BRCA2 genes in patients with early-onset breast and/or ovarian cancer have not been identified within the Iranian population.

Methods: With the collaboration of two main centres for cancer in Iran, we obtained clinical information, family history and peripheral blood from 83 women under the age of 45 with early-onset breast cancer for scanning of germline mutations in the BRCA1 and BRCA2 genes. We analysed BRCA1 exons 11 and BRCA2 exons 10 and 11 by the protein truncation test, and BRCA1 exons 2, 3, 5, 13 and 20 and BRCA2 exons 9, 17, 18 and 23 with the single-strand conformation polymorphism assay on genomic DNA amplified by polymerase chain reaction.

Results: Ten sequence variants were identified: five frameshifts (putative mutations - four novel); three missense changes of unknown significance and two polymorphisms, one seen commonly in both Iranian and British populations.

Conclusions: Identification of these novel mutations suggests that any given population should develop a mutation database for its programme of breast cancer screening. The pattern of mutations seen in the BRCA genes seems not to differ from other populations studied. Early-onset breast cancer (less than 45 years) and a limited family history is sufficient to justify mutation screening with a detection rate of over 25% in this group, whereas sporadic early-onset breast cancer (detection rate less than 5%) is unlikely to be cost-effective.

Figure 2

SDS-PAGE analysis of the whole of exon 11 of BRCA2, revealing the capacity of the PTT technique to detect mutations within 4959 bp coding sequences in a single reaction with the use of a coupled transcription-translation system, TNT^® T7 Quick for PCR DNA kit from Promega. Lanes 1 and 2 show the normal pattern of the full-length (181 kDa) protein from exon 11 of BRCA2; lanes 3–5 show three different sizes of truncated protein, which were identified on 6% SDS-PAGE. Arrows show the sizes and positions of the normal and truncated proteins. Lane 4 shows a mutation occurring close to the 3' end of exon 11 and producing a large protein for which the band migrated close to the top of the gel.

Figure 1

Single-strand conformation polymorphism (SSCP) assay for germline mutation in BRCA1 exon 2. Lanes 1 and 4 show normal patterns, and lane 2 and 3 depict the abnormal patterns of single-strand DNA mobility seen on a polyacrylamide gel (b). Frameshift mutation in sample (lane 2) confirmed by direct sequencing (a) that shows a 2 bp (AG) deletion in BRCA1 exon 2 at nucleotides 185–186 that leads to the formation of TGA at codon 39. (c) A frameshift mutation (lane 3) identified by direct sequencing, which revealed a 1 bp (T) insertion in BRCA1 exon 2 between nucleotides 181 and 182, leading to the formation of TGA at codon 40. These frameshift mutations are likely to disrupt the function of the BRCA1 proteins.