13-32326613-G-C

Position:

• chr13-32326613-G-C

Variant summary

Our verdict is Pathogenic. Variant got 12 ACMG points: 12P and 0B. PM2 PM5 PP5_Very_Strong

The NM_000059.4(BRCA2):​c.631G>C​(p.Val211Leu) variant causes a missense, splice region change. The variant allele was found at a frequency of 0.000000702 in 1,424,234 control chromosomes in the GnomAD database, with no homozygous occurrence. In-silico tool predicts a benign outcome for this variant. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Likely pathogenic (★★). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. V211A) has been classified as Likely benign.

• Frequency:
  • Genomes: not found (cov: 33)
  • Exomes 𝑓: 7.0e-7 (0 hom.)
• Consequence: BRCA2 NM_000059.4 missense, splice_region
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:11
• Conservation: PhyloP100: 3.91

Genes affected

BRCA2 (HGNC:1101): (BRCA2 DNA repair associated) Inherited mutations in BRCA1 and this gene, BRCA2, confer increased lifetime risk of developing breast or ovarian cancer. Both BRCA1 and BRCA2 are involved in maintenance of genome stability, specifically the homologous recombination pathway for double-strand DNA repair. The largest exon in both genes is exon 11, which harbors the most important and frequent mutations in breast cancer patients. The BRCA2 gene was found on chromosome 13q12.3 in human. The BRCA2 protein contains several copies of a 70 aa motif called the BRC motif, and these motifs mediate binding to the RAD51 recombinase which functions in DNA repair. BRCA2 is considered a tumor suppressor gene, as tumors with BRCA2 mutations generally exhibit loss of heterozygosity (LOH) of the wild-type allele. [provided by RefSeq, May 2020]

BRCA2 (HGNC:):

ACMG classification

Verdict is Pathogenic. Variant got 12 ACMG points.

• PM2: Very rare variant in population databases, with high coverage
• PM5: Other missense variant is known to change same aminoacid residue: Variant chr13-32326613-G-A is described in Lovd as [Pathogenic].
• PP5: Variant 13-32326613-G-C is Pathogenic according to our data. Variant chr13-32326613-G-C is described in ClinVar as [Likely_pathogenic]. Clinvar id is 38035.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars. Variant chr13-32326613-G-C is described in Lovd as [Pathogenic]. Variant chr13-32326613-G-C is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
BRCA2	NM_000059.4	c.631G>C	p.Val211Leu	missense_variant, splice_region_variant	7/27	ENST00000380152.8	NP_000050.3

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
BRCA2	ENST00000380152.8	c.631G>C	p.Val211Leu	missense_variant, splice_region_variant	7/27	5	NM_000059.4	ENSP00000369497.3
BRCA2	ENST00000530893.7	c.262G>C	p.Val88Leu	missense_variant, splice_region_variant	7/27	1		ENSP00000499438.2
BRCA2	ENST00000614259.2	n.631G>C		splice_region_variant, non_coding_transcript_exon_variant	6/26	2		ENSP00000506251.1

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome AF: 7.02e-7 AC: 1 AN: 1424234 Hom.: 0 Cov.: 29 AF XY: 0.00 AC XY: 0 AN XY: 710436

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.00

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.00

Gnomad4 SAS exome AF: 0.00

Gnomad4 FIN exome AF: 0.00

Gnomad4 NFE exome AF: 9.27e-7

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome Cov.: 33

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:11

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Breast-ovarian cancer, familial, susceptibility to, 2 Pathogenic:3

Pathogenic, no assertion criteria provided	clinical testing	Breast Cancer Information Core (BIC) (BRCA2)	May 29, 2002	- -
Pathogenic, no assertion criteria provided	clinical testing	Sharing Clinical Reports Project (SCRP)	Oct 09, 2013	- -
Pathogenic, criteria provided, single submitter	clinical testing	Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA), c/o University of Cambridge	Oct 02, 2015	- -

not provided Pathogenic:3

Likely pathogenic, criteria provided, single submitter	clinical testing	Revvity Omics, Revvity	Aug 19, 2022	- -
Pathogenic, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Nov 20, 2017	- -
Pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Mar 18, 2015	This pathogenic variant is denoted BRCA2 c.631G>C at the cDNA level. Using alternate nomenclature, This variant would be defined as BRCA2 859G>C. Although the nucleotide substitution results in the change of a Valine to a Leucine at codon 211, and is called Val211Leu in the literature, we are using only the nucleotide nomenclature to refer to the mutation since the defect is determined to be one of splicing rather than a resulting missense pathogenic variant. Multiple splicing models predict that this variant may destroy the natural splice donor site for intron 7 and lead to abnormal splicing. Consistent with the splicing models, an in vitro minigene assay found that a significant proportion of transcripts produced from BRCA2 c.631G>C were missing exon 7 (Di Giacomo 2013). BRCA2 c.631G>C was not observed in approximately 6,500 individuals of European and African American ancestry in the NHLBI Exome Sequencing Project, indicating it is not a common benign variant in these populations. The nucleotide which is altered, a guanine (G) at base 631, is conserved across species. Based on the current evidence, we consider this variant to be pathogenic. -

Hereditary cancer-predisposing syndrome Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	Jun 13, 2023	The c.631G>C variant (also known as p.V211L), located in coding exon 6 of the BRCA2 gene, results from a G to C substitution at nucleotide position 631. The valine at codon 211 is replaced by leucine, an amino acid with highly similar properties. This alteration has been reported in multiple individuals undergoing genetic testing for hereditary breast and/or ovarian cancer (Lin PH et al. Oncotarget. 2016 Feb;7:8310-20; Crawford B et al. Breast Cancer Res Treat. 2017 Jun;163(2):383-390; Rebbeck TR et al. Hum Mutat. 2018 May;39(5):593-620; Bhaskaran SP et al. Int J Cancer. 2019 Aug;145(4):962-973; Laitman Y et al. Hum Mutat. 2019 Nov;40(11):e1-e23 ). This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). This change occurs in the last base pair of coding exon 6, which makes it likely to have some effect on normal mRNA splicing. In silico splice site analysis predicts that this alteration will weaken the native splice donor site. RNA studies have demonstrated that this alteration results in abnormal splicing in the set of samples tested (Ambry internal data; Di Giacomo D et al. Hum. Mutat. 2013 Nov;34:1547-57). Another alteration impacting the same donor site (c.631+2T>G) has been shown to have a similar impact on splicing in and has been identified in the homozygous state and in the compound heterozygous state with other pathogenic BRCA2 mutations in individuals with Fanconi anemia (Wagner JE et al. Blood. 2004 Apr;103:3226-9). This nucleotide position is highly conserved in available vertebrate species. Based on the majority of available evidence to date, this variant is likely to be pathogenic. However, because a variant with the same splicing profile as this variant has been identified in one or more patients with Fanconi Anemia it may be hypomorphic and thus, carriers of this variant and their families may present with reduced risks, and not with the typical clinical characteristics of a high-risk pathogenic BRCA2 alteration. As risk estimates are unknown at this time, clinical correlation is advised. -
Pathogenic, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Sep 08, 2021	This variant replaces c.G with c.C at c.631 nucleotide position of the BRCA2 gene. This variant alters the last nucleotide position of exon 7 and is predicted to impair RNA splicing. A functional RNA study using a mini-gene assay has shown that this variant causes out-of-frame skipping of exon 7 and use of an alternative splice site 70 nucleotides upstream (PMID: 23983145). Both abnormal transcript products would result in frameshift and premature truncation. This variant has been reported in individuals affected with breast cancer (PMID: 26824983, 31090900, 33471991) and has been identified in 9 families among CIMBA participants (PMID: 29446198). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). A different variant occurring at the same nucleotide position, c.631G>A, is a well documented pathogenic variant due to its deleterious impact on RNA splicing (ClinVar variation ID: 52058). Loss of BRCA2 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Pathogenic. -

Hereditary breast ovarian cancer syndrome Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Feb 28, 2021	Variant summary: BRCA2 c.631G>C (p.Val211Leu) results in a conservative amino acid change in the encoded protein sequence. Four of five in-silico tools predict a benign effect of the variant on protein function. This variant alters the last nucleotide of exon 7 located adjacent to the canonical intronic splice donor site. Several computational tools predict a significant impact on normal splicing: Two predict the variant abolishes a 5' splicing donor site. One predict the variant weakens a 5' donor site. At least one publication reports experimental evidence that this variant affects mRNA splicing resulting in exon 7 skipping in an ex vivo pCAS2-BRCA2-Exon 7 minigene assay system (Di Giacomo_2013). This study also reported an activation of an upstream internal cryptic 5' splice site resulting in the excision of the last 70 nucleotides of the exon. To our knowledge, a confirmation of these findings in patient derived RNA has not been reported. The variant was absent in 250814 control chromosomes. c.631G>C has been reported in the literature in multiple individuals affected with Hereditary Breast And Ovarian Cancer Syndrome (example, Rebbeck_2018). These data indicate that the variant is very likely to be associated with disease. Three clinical diagnostic laboratories and one consortium (CIMBA, cited by Rebbeck_2018 above) have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. All submitters classified the variant as pathogenic (n=3)/likely pathogenic (n=1). Based on the evidence outlined above, until additional in-vivo evidence confirming the observed ex-vivo impact on exon 7 skipping is identified, the variant was classified as likely pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Oct 13, 2023	This sequence change affects codon 211 of the BRCA2 mRNA. It is a 'silent' change, meaning that it does not change the encoded amino acid sequence of the BRCA2 protein. RNA analysis indicates that this variant induces altered splicing and may result in an absent or disrupted protein product. This variant is not present in population databases (gnomAD no frequency). This variant has been observed in individual(s) with breast cancer and/or ovarian cancer (PMID: 26824983, 28281021, 31090900). ClinVar contains an entry for this variant (Variation ID: 38035). An algorithm developed to predict the effect of missense changes on protein structure and function (PolyPhen-2) suggests that this variant is likely to be disruptive. Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Studies have shown that this variant alters mRNA splicing and is expected to lead to the loss of protein expression (PMID: 23983145; Invitae). This variant disrupts the c.631G nucleotide in the BRCA2 gene. Other variant(s) that disrupt this nucleotide have been determined to be pathogenic (PMID: 19179552, 22962691, 23451180). This suggests that this nucleotide is clinically significant, and that variants that disrupt this position are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic. -

BRCA2-related cancer predisposition Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

All of Us Research Program, National Institutes of Health

May 07, 2024

This variant replaces c.G with c.C at c.631 nucleotide position of the BRCA2 gene. This variant alters the last nucleotide position of exon 7 and is predicted to impair RNA splicing. A functional RNA study using a mini-gene assay has shown that this variant causes out-of-frame skipping of exon 7 and use of an alternative splice site 70 nucleotides upstream (PMID: 23983145). Both abnormal transcript products would result in frameshift and premature truncation. This variant has been reported in individuals affected with breast cancer (PMID: 26824983, 31090900, 33471991) and has been identified in 9 families among CIMBA participants (PMID: 29446198). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). A different variant occurring at the same nucleotide position, c.631G>A, is a well documented pathogenic variant due to its deleterious impact on RNA splicing (ClinVar variation ID: 52058). Loss of BRCA2 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Pathogenic. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Uncertain	0.43
BayesDel_addAF	Benign	-0.019	T
BayesDel_noAF	Benign	-0.26
CADD	Pathogenic	26
DANN	Uncertain	1.0
Eigen	Uncertain	0.22
Eigen_PC	Uncertain	0.35
FATHMM_MKL	Uncertain	0.97	D
M_CAP	Uncertain	0.15	D
MetaRNN	Benign	0.33	T;T
MetaSVM	Benign	-0.86	T
PrimateAI	Uncertain	0.52	T
PROVEAN	Benign	-0.64	N;N
REVEL	Benign	0.16
Sift	Uncertain	0.029	D;D
Sift4G	Uncertain	0.017	D;D
Vest4		0.36
MutPred		0.24		Loss of sheet (P = 0.1158);Loss of sheet (P = 0.1158);
MVP		0.81
MPC		0.070
ClinPred		0.90	D
GERP RS		5.7
gMVP		0.35

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	1.0
SpliceAI score (max)		0.82		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.82		Position offset: 0

Find out detailed SpliceAI scores and Pangolin per-transcript scores at spliceailookup.broadinstitute.org

Publications

LitVar

Other links and lift over

dbSNP: rs80358871; hg19: chr13-32900750;