13-32362521-A-G

Position:

• chr13-32362521-A-G

Variant summary

Our verdict is Pathogenic. Variant got 16 ACMG points: 16P and 0B. PVS1_Moderate PM2 PP3_Strong PP5_Very_Strong

The NM_000059.4(BRCA2):​c.7806-2A>G variant causes a splice acceptor change involving the alteration of a conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. In-silico tool predicts a pathogenic outcome for this variant. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Pathogenic (★★★).

• Frequency: Genomes: not found (cov: 32)
• Consequence: BRCA2 NM_000059.4 splice_acceptor
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic reviewed by expert panel P:22
• Conservation: PhyloP100: 9.18

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]

ACMG classification

Verdict is Pathogenic. Variant got 16 ACMG points.

• PVS1: Splicing variant, NOT destroyed by nmd, known LOF gene, truncates exone, which is 0.016574048 fraction of the gene. Cryptic splice site detected, with MaxEntScore 6, offset of 20, new splice context is: ggctctgtgtgacactccAGgtg. Cryptic site results in frameshift change. If cryptic site found is not functional and variant results in exon loss, it results in inframe change.
• PM2: Very rare variant in population databases, with high coverage
• PP3: Splicing scoreres supports a deletorius effect: Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF, max_spliceai. No scorers claiming Uncertain. No scorers claiming Benign.
• PP5: Variant 13-32362521-A-G is Pathogenic according to our data. Variant chr13-32362521-A-G is described in ClinVar as [Pathogenic]. Clinvar id is 52418.Status of the report is reviewed_by_expert_panel, 3 stars. Variant chr13-32362521-A-G is described in Lovd as [Pathogenic]. Variant chr13-32362521-A-G is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
BRCA2	NM_000059.4	c.7806-2A>G		splice_acceptor_variant		ENST00000380152.8

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
BRCA2	ENST00000380152.8	c.7806-2A>G		splice_acceptor_variant		5	NM_000059.4	A2

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Cov.: 33

GnomAD4 genome Cov.: 32

ClinVar

Significance: Pathogenic

Submissions summary: Pathogenic:22

Revision: reviewed by expert panel

Submissions by phenotype

Breast-ovarian cancer, familial, susceptibility to, 2 Pathogenic:9

Pathogenic, criteria provided, single submitter	research	Centre for Mendelian Genomics, University Medical Centre Ljubljana	Dec 09, 2022	PVS1, PS3, PS4_STR, PM2_SUP -
Pathogenic, no assertion criteria provided	clinical testing	Breast Cancer Information Core (BIC) (BRCA2)	May 29, 2002	- -
Pathogenic, reviewed by expert panel	curation	Evidence-based Network for the Interpretation of Germline Mutant Alleles (ENIGMA)	Jun 18, 2019	IARC class based on posterior probability from multifactorial likelihood analysis, thresholds for class as per Plon et al. 2008 (PMID: 18951446). Class 5 based on posterior probability = 0.998459 -
Likely pathogenic, no assertion criteria provided	clinical testing	Sharing Clinical Reports Project (SCRP)	Aug 10, 2007	- -
Pathogenic, criteria provided, single submitter	clinical testing	Human Genome Sequencing Center Clinical Lab, Baylor College of Medicine	Feb 28, 2019	This c.7806-2A>G variant in the BRCA2 gene disrupts the canonical splice donor site in intron 16 and is predicted to result in abnormal mRNA splicing. This variant is absent from large databases of genetic variation in the general population. This variant has been reported in multiple patients with breast, ovarian and pancreatic cancers (PMID 10449599, 12097290, 16764716, 18783588). Therefore, this c.7806-2A>G variant in the BRCA2 gene is classified as pathogenic. -
Pathogenic, no assertion criteria provided	clinical testing	BRCAlab, Lund University	Aug 26, 2022	- -
Pathogenic, criteria provided, single submitter	clinical testing	All of Us Research Program, National Institutes of Health	Dec 18, 2023	This variant causes an A to G nucleotide substitution at the -2 position of intron 16 of the BRCA2 gene. RNA studies have detected aberrant splicing products in carrier RNA (PMID: 10449599, 12461697, 22505045, 30832263) and in a minigene splicing assay (PMID: 28339459) that include out-of-frame and in-frame splicing products. To our knowledge, functional studies have not been reported for this variant. This variant has been reported in at least 10 individuals affected with breast and/or ovarian cancer and 1 individual affected with pancreatic cancer (PMID: 10449599, 12097290, 16764716, 18439106, 29470806, 33891299) and has a segregation likelihood ratio for pathogenicity of 7.1109 (PMID: 31131967). Haplotype analyses suggest that this variant is a founder mutation in Slovenia and Italy (PMID: 12461697, 18439106, 26852130). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Loss of BRCA2 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Pathogenic. -
Pathogenic, no assertion criteria provided	clinical testing	Department of Pathology and Laboratory Medicine, Sinai Health System	-	The c.7806-2A>G variant was identified in 14 of 872 proband chromosomes (frequency: 0.016) from individuals or families with hereditary breast or ovarian cancer (Santarosa 1999, Novakovic 2012). The variant was also identified in dbSNP (ID: rs81002836) “With likely pathogenic allele”, HGMD, the ClinVar database (classified as a Pathogenic/Likely pathogenic variant by the Sharing Clinical Reports Project, derived from Myriad reports), GeneInsight through the Canadian Open Genetics Repository (http://opengenetics.ca/) (1X, classified as “pathogenic” by a clinical laboratory), the BIC database (5X with “pending” clinical importance), and UMD (5X as a causal variant).The c.7806-2A>G variant is predicted to cause abnormal splicing because the nucleotide substitution occurs in the invariant region of the splice consensus sequence. In addition, 5 of 5 in silico or computational prediction software programs (SpliceSiteFinder, MaxEntScan, NNSPLICE, GeneSplicer, HumanSpliceFinder) predict a greater than 10% difference in splicing. Also, mini-gene splicing assays show the variant to have a severe impact on splicing (Houdayer 2012). In summary, based on the above information, this variant meets our laboratory’s criteria to be classified as pathogenic. -
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:5

Pathogenic, no assertion criteria provided	clinical testing	Genome Diagnostics Laboratory, University Medical Center Utrecht	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Feb 02, 2022	Canonical splice site variant the produces multiple transcripts, leading to an abnormal message that is subject to nonsense-mediated mRNA decay or to an abnormal protein product (Santarosa 1999, Krajc 2002, Fraile-Bethencourt 2017, Gelli 2019); Observed in individuals with a personal or family history consistent with pathogenic variants in this gene, and is a founder variant in the Slovenian and North-Eastern Italian populations (Santarosa 1999, Krajc 2002, Marroni 2004, Besic 2008, Stegel 2011, Cini 2016); Not observed at significant frequency in large population cohorts (gnomAD); Truncating variants in this gene are considered pathogenic by a well-established clinical consortium and/or database; Also known as 8034-2A>G; This variant is associated with the following publications: (PMID: 19818148, 20104584, 26295337, 29907814, 12461697, 15340362, 22505045, 22923021, 25525159, 10449599, 10923033, 28339459, 24156927, 26852130, 22984553, 17301269, 16764716, 21232165, 18439106, 18783588, 23397983, 12097290, 24312913, 23199084, 25186627, 29470806, 29446198, 34399810, 33891299, 31131967, 12228710, 30832263, 30613976, 31360904) -
Pathogenic, criteria provided, single submitter	clinical testing	CeGaT Center for Human Genetics Tuebingen	Dec 01, 2020	- -
Pathogenic, no assertion criteria provided	clinical testing	Joint Genome Diagnostic Labs from Nijmegen and Maastricht, Radboudumc and MUMC+	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Jan 29, 2020	This variant is located in a canonical splice-acceptor site and interferes with normal BRCA2 mRNA splicing. In the published literature, this variant has been reported in individuals with hereditary breast and/or ovarian cancer (PMID: 29446198 (2018), 10449599 (1999)). Additionally, it has been described as a founder variant in Slovenian breast cancer populations (PMID: 24312913 (2013), PMID: 21232165 (2011), 12461697 (2002)). Functional studies have confirmed that this variant leads to aberrant splicing (PMID: 28339459 (2017), 10449599 (1999)). This variant has not been reported in large, multi-ethnic general populations (Genome Aggregation Database, http://gnomad.broadinstitute.org). Based on the available information, this variant is classified as pathogenic. -

Hereditary cancer-predisposing syndrome Pathogenic:3

Pathogenic, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Apr 18, 2023	This variant causes an A to G nucleotide substitution at the -2 position of intron 16 of the BRCA2 gene. RNA studies have detected aberrant splicing products in carrier RNA (PMID: 10449599, 12461697, 22505045, 30832263) and in a minigene splicing assay (PMID: 28339459) that include out-of-frame and in-frame splicing products. To our knowledge, functional studies have not been reported for this variant. This variant has been reported in at least 10 individuals affected with breast and/or ovarian cancer and 1 individual affected with pancreatic cancer (PMID: 10449599, 12097290, 16764716, 18439106, 29470806, 33891299) and has a segregation likelihood ratio for pathogenicity of 7.1109 (PMID: 31131967). Haplotype analyses suggest that this variant is a founder mutation in Slovenia and Italy (PMID: 12461697, 18439106, 26852130). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Loss of BRCA2 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	Jan 02, 2024	The c.7806-2A>G intronic pathogenic mutation results from an A to G substitution two nucleotides upstream from coding exon 16 in the BRCA2 gene. This alteration has been reported in splicing studies from both minigene and RT-PCR from patient cells to result in multiple aberrant transcripts including a majority of transcripts derived from the use of a cryptic acceptor site 20 nucleotides downstream in exon 17 (coding exon 16) leading to a predicted frameshift (Krajc M et al. Eur. J. Hum. Genet. 2002 Dec;10:879-82; Houdayer C et al. Hum. Mutat. 2012 Aug;33:1228-38; Fraile-Bethencourt E et al. PLoS Genet. 2017 Mar;13(3):e1006691; Gelli E et al. Cancers (Basel) 2019 Mar;11; Ambry internal data). This alteration has been described as a Slovenian founder mutation and has been identified in hereditary breast and ovarian cancer patients, male breast cancer patients, and familial pancreatic cancer patients (Santarosa M et al. Int. J. Cancer. 1999 Sep;83:5-9; Krajc M et al. Eur. J. Hum. Genet. 2002 Dec;10:879-82; Miolo G et al. BMC Cancer. 2006 Jun;6:156; Murphy KM et al. Cancer Res. 2002 Jul;62:3789-93; Miolo G et al. BMC Cancer. 2006 Jun;6:156; Besic N et al. Genet. Test. 2008 Jun;12:203-9). In addition, a high ratio of breast cancer versus ovarian cancer (relative risk: 16.33) has been noted in families harboring this alteration (Krajc M et al. BMC Med. Genet. 2008;9:83). This alteration was classified as pathogenic in a multifactorial model of variant interpretation that incorporates co-segregation, family history, co-occurrence, tumor pathology, and case-control data (Parsons MT et al. Hum Mutat. 2019 Sep;40(9):1557-1578). Of note, this alteration is also designated as IVS16-2A>G in published literature. This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). This nucleotide position is highly conserved in available vertebrate species. In silico splice site analysis predicts that this alteration will weaken the native splice acceptor site and will result in the creation or strengthening of a novel splice acceptor site. Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation. -
Pathogenic, criteria provided, single submitter	curation	Sema4, Sema4	Feb 10, 2022	- -

Hereditary breast ovarian cancer syndrome Pathogenic:2

Pathogenic, criteria provided, single submitter	clinical testing	Invitae	Jan 28, 2024	This sequence change affects an acceptor splice site in intron 16 of the BRCA2 gene. RNA analysis indicates that disruption of this splice site induces altered splicing and may result in an absent or disrupted protein product. This variant is not present in population databases (gnomAD no frequency). Disruption of this splice site has been observed in individual(s) with breast/ovarian cancer (PMID: 10449599, 12461697, 18439106, 18783588, 21232165, 23199084). It is commonly reported in individuals of Slovenian ancestry (PMID: 10449599, 12461697, 18439106, 18783588, 21232165, 23199084). ClinVar contains an entry for this variant (Variation ID: 52418). Studies have shown that disruption of this splice site results in activation of a cryptic splice site and introduces a premature termination codon (PMID: 10449599, 12461697; Invitae). The resulting mRNA is expected to undergo nonsense-mediated decay. For these reasons, this variant has been classified as Pathogenic. -
Pathogenic, no assertion criteria provided	research	Research Molecular Genetics Laboratory, Women's College Hospital, University of Toronto	Jan 31, 2014	- -

Breast-ovarian cancer, familial, susceptibility to, 1 Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Department of Molecular Diagnostics, Institute of Oncology Ljubljana

Apr 02, 2020

- -

Familial cancer of breast Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Baylor Genetics

Nov 14, 2022

- -

Ovarian neoplasm Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Centre for Mendelian Genomics, University Medical Centre Ljubljana

Oct 06, 2015

- -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.39	D
BayesDel_noAF	Pathogenic	0.32
CADD	Pathogenic	34
DANN	Uncertain	1.0
Eigen	Pathogenic	1.2
Eigen_PC	Pathogenic	1.0
FATHMM_MKL	Pathogenic	1.0	D
MutationTaster	Benign	1.0	D;D
GERP RS		5.7

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.93
SpliceAI score (max)		0.97		Details are displayed if max score is > 0.2
DS_AG_spliceai		0.50		Position offset: 22
DS_AL_spliceai		0.97		Position offset: 2

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

Publications

LitVar

Other links and lift over

dbSNP: rs81002836; hg19: chr13-32936658; COSMIC: COSV66452693;