In silico, in vitro and case-control analyses as an effective combination for analyzing BRCA1 and BRCA2 unclassified variants in a population-based sample

Abstract

Ascertaining the clinical consequences of BRCA1 and BRCA2 variants of uncertain significance (VUS) is currently indispensable for providing effective genetic counseling and preventive actions for families with hereditary breast and ovarian cancer (HBOC). To this end, we conducted a combination of in silico prediction and cDNA splicing analyses of 13 BRCA1 and 10 BRCA2 VUS identified in our cohort as well as a case-control analysis in a population-based sample of 10 recurrent VUS. We observed consistent results between the in silico predictions and sequencing analyses for all analyzed VUS. An abnormal cDNA pattern was observed for variants c.212+1G>A and c.5278-1G>A in BRCA1 and c.516+2T>A and c.8168A>G in BRCA2 according to in silico splicing prediction. A case-control study of VUS confirmed the polymorphisms of the c.67+62A>G, c.7008-62A>G and c.8851G>A BRCA2 variants previously published. c.4068G>A in the BRCA2 gene can also be considered a polymorphism due to its occurrence at a frequency greater than 1% in our population. Our study shows that employing population-based analysis and a combination of several in silico methods yields highly accurate information, resulting in a reliable tool for selecting variants for cDNA sequencing analysis in routine cancer genetic counseling units.

Keywords: BRCA1; BRCA2; cDNA; cancer genetic counseling; splicing; variants of unknown significance.