BRCA2 NM_000059.3:c.9227G>T, Gly3076Val

This sequence change replaces glycine, which is neutral and non-polar, with valine, which is neutral and non-polar, at codon 3076 of the BRCA2 protein (p.Gly3076Val). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individual(s) with hereditary breast and/or ovarian cancer (PMID: 29161300, 30254663, 32814805). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 126203). Advanced modeling performed at Invitae incorporating data from internal and/or published experimental studies (PMID: 33609447) indicates that this missense variant is expected to disrupt BRCA2 function. Experimental studies have shown that this missense change affects BRCA2 function (PMID: 23108138, 29394989). This variant disrupts the p.Gly3076 amino acid residue in BRCA2. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 12569143, 22711857, 23108138, 29394989, 30078507). This suggests that this residue is clinically significant, and that variants that disrupt this residue are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic.

This missense variant replaces glycine with valine at codon 3076 in the DNA binding domain of the BRCA2 protein. Computational prediction tool suggests that this variant may have deleterious impact on protein structure and function (internally defined REVEL score threshold >=0.7, PMID: 27666373). Functional studies have shown that this variant results in the loss of homology directed repair activity of the BRCA2 protein (PMID: 23108138, 33609447). This variant has been reported in individuals affected with breast and ovarian cancer (PMID: 32814805, 33007869). It has been shown that this variant segregates with disease in at least one HBOC family (PMID: 32814805). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Based on the available evidence, this variant is classified as Likely Pathogenic.

The p.G3076V pathogenic mutation (also known as c.9227G>T), located in coding exon 23 of the BRCA2 gene, results from a G to T substitution at nucleotide position 9227. The glycine at codon 3076 is replaced by valine, an amino acid with dissimilar properties. This alteration was identified in a cohort of 418 Southern Brazilian HBOC probands (Alemar B et al. PLoS ONE. 2017 Nov;12:e0187630). Homology-directed DNA repair (HDR) assays have demonstrated this variant to be non-functional (Guidugli L et al. Am. J. Hum. Genet. 2018 02;102:233-248; Hart SN et al. Genet. Med. 2019 01;21:71-80). Internal structural analysis indicates that this amino acid substitution will be strongly destabilizing to the local structure and may affect binding to APRIN and PALB2 (Yang H et al. Science. 2002 Sep;297:1837-48; Ambry internal data). This amino acid position is completely conserved on sequence alignment. This alteration is predicted to be deleterious by in silico analysis. Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation.

Data used in classification: The frequency of this variant is 0/138,632 individuals (gnomAD) (PM2_mod). This variant is predicted deleterious on AlignGVGD (class: C65), SIFT (Deleterious), Polyphen2 HumVar (probably damaging) and CADD (25.3) (PP3_sup). The variant is in the DNA-binding domain of BRCA2 (PM1_sup). In the BRCA2 Homology-Directed Repair Activity assay for the DNA Binding Domain (Guidugli et al Cancer Res 2013;73:265-275,Couch Lab), the variant has a probability of pathogenicity of 1.0 (PS3_strong). Data not used in classification: There are additional reports of this variant in ClinVar (4), BIC (2), and BRCA2 LOVD (1).

The variant c.9227G>T in BRCA2 is a missense variant predicted to cause substitution of Glycine by Valine at amino acid 3076 (p.Gly3076Val). This variant is absent from gnomAD v2.1 (exomes only, non-cancer subset, read depth >=25) and gnomAD v3.1 (non-cancer subset, read depth >=25) (PM2_Supporting met). This BRCA2 missense variant is within a key functional domain and the computational predictor BayesDel (noAF) gives a score of 0.398, above the recommended threshold of 0.30 for prediction of impact on BRCA2 function via protein change. SpliceAI predictor score of 0.01 suggests that the variant has no impact on splicing (score threshold <0.10) (PP3 met). Reported by one calibrated study to exhibit protein function similar to pathogenic control variants (PMID: 33609447) (PS3 met). Multifactorial likelihood ratio analysis using clinically calibrated data produced a combined LR for this variant of 3104.98 (based on Cosegregation LR=770.85; Pathology LR=1.89; Co-occurrence LR=1.08; Family History LR=1.98), above the threshold for Very strong evidence towards pathogenicity (LR >350) (PP4_Very strong met; PMID: 31853058, Internal lab contributors). In summary, this variant meets the criteria to be classified as a Pathogenic variant for BRCA2-related cancer predisposition based on the ACMG/AMP criteria applied as specified by the ENIGMA BRCA1/2 VCEP (PM2_Supporting, PP3, PS3, PP4_Very strong).