DHPLC-mutation scanning of the breast cancer predisposing gene BRCA1 in breast cancer patients from the south of Iran

Background: Germline mutations in Breast Cancer gene (BRCA1) confer anestimated cumulative lifetime risk of breast cancer of 56–84%. Furthermore, thecancer-founder effects of BRCA mutations have been suggested to be race and ethnicpopulation-specific. Previous investigations of our group indicated no association ofknown founder mutations in BRCA genes (185delAG, 5382insC, 6174delT) withfamilial breast cancer in Iranian populations. Objective: as a part of a larger projectsearching for BRCA founder mutations in Iranians, we aimed in the present study toinvestigate germ-line mutations in exon 16 of BRCA1 gene in females with familialbreast cancer in south of Iran. Subjects and Methods: DNA was extracted fromperipheral blood samples of 70 affected women with familial breast cancer, and 70age-sex matched healthy subjects. Denaturing high performance liquidchromatography (DHPLC) method, using Transgenomic WAVE 4500 system, wasapplied for genetic pre-screening of BRCA1 gene in order to reduce the cost and timefor the analysis of this gene. Chromatograms analysis was performed with NavigatorSoftware 2.2.0. Mutation positive amplicons had to be subsequently sequenced bydirect DNA sequencing using BigDye Terminator chemistry and ABI 310 sequencer.Results: Two point mutations were detected in exon 16 with different frequencies inboth patients and controls. The frequencies of A/G missence mutation (c.[4837A> G])causing serine 1613 with Glycin substitution were 60% vs. 50% in patients andcontrols, respectively. Moreover, a novel G/A mutation (c.[4956G> A]) thatsubstitutes Met 1652 with Ile was observed in 8 patients and only 1 control.Conclusion: The preliminary data indicated identification of new mutations inBRCA1 gene exon 16 in Iranians, suggesting appropriate targets for furtherinvestigations as the founder mutations in this population. In this regard, utilizing afast and reliable mutation pre-screening method like DHPLC can help to acceleratemutations scanning in BRCA genes of carrier females.