13-32376795-A-T

Position:

• chr13-32376795-A-T

Variant summary

Our verdict is Pathogenic. Variant got 10 ACMG points: 10P and 0B. PM2 PP5_Very_Strong

The NM_000059.4(BRCA2):​c.8754+4A>T variant causes a splice region, intron change involving the alteration of a non-conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. In-silico tool predicts a benign outcome for this variant. 2/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Likely pathogenic (★★).

• Frequency: Genomes: not found (cov: 32)
• Consequence: BRCA2 NM_000059.4 splice_region, intron
• Scores: Splicing: ADA: 0.9880
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:5 U:1
• Conservation: PhyloP100: 3.47

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 10 ACMG points.

• PM2: Very rare variant in population databases, with high coverage
• PP5: Variant 13-32376795-A-T is Pathogenic according to our data. Variant chr13-32376795-A-T is described in ClinVar as [Likely_pathogenic]. Clinvar id is 220353.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
BRCA2	NM_000059.4	c.8754+4A>T		splice_region_variant, intron_variant		ENST00000380152.8	NP_000050.3

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
BRCA2	ENST00000380152.8	c.8754+4A>T		splice_region_variant, intron_variant		5	NM_000059.4	ENSP00000369497.3
BRCA2	ENST00000530893.7	c.8385+4A>T		splice_region_variant, intron_variant		1		ENSP00000499438.2
BRCA2	ENST00000614259.2	n.*812+4A>T		splice_region_variant, intron_variant		2		ENSP00000506251.1

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Cov.: 32

GnomAD4 genome Cov.: 32

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:5 Uncertain:1

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Hereditary cancer-predisposing syndrome Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Nov 01, 2023	This variant causes an A to T nucleotide substitution at the +4 position of intron 21 of the BRCA2 gene. Splice site prediction tools predict that this variant may have a significant impact on RNA splicing. RNA studies have shown this variant produces a mutant transcript that partially retains 46 nucleotides of intron 21, creating a frameshift and premature translation stop signal (PMID: 31642931, 32133419, 36446827). This variant has not been reported in individuals affected with BRCA2-related disorders in the literature. However, a different variant at the same nucleotide position, BRCA2 c.8754+4A>G, has been reported as Pathogenic (ClinVar variation ID 52669). The BRCA2 c.8754+4A>T 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 Likely Pathogenic. -
Likely pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	Dec 13, 2023	The c.8754+4A>T intronic variant results from an A to T substitution 4 nucleotides after coding exon 20 in the BRCA2 gene. This nucleotide position is highly conserved in available vertebrate species. In silico splice site analysis predicts that this alteration will weaken the native splice donor site and will result in the creation or strengthening of a novel splice donor site. RNA studies have demonstrated this alteration results in abnormal splicing (Ambry internal data). Another alteration impacting the same donor site (c.8754+4A>G) has been shown to have a similar impact on splicing and is predicted to be pathogenic in multifactorial analyses (Bonatti F et al. Cancer Genet Cytogenet 2006 Oct;170(2):93-101; Acedo A et al. Hum Mutat 2015 Feb;36(2):210-21; Lindor NM et al. Hum Mutat 2012 Jan;33(1):8-21.) . Based on the majority of available evidence to date, this variant is likely to be 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	Sep 02, 2022	Variant summary: BRCA2 c.8754+4A>T alters a conserved nucleotide located close to a canonical splice site and therefore could affect mRNA splicing, leading to a significantly altered protein sequence. Several computational tools predict a significant impact on normal splicing: One predict the variant abolishes the canonical 5' splicing donor site. Three predict the variant weakens the canonical 5' splicing donor site. At least two publications report experimental evidence that this variant affects mRNA splicing by activation of a downstream intronic splice donor leading to a retention of 46 nucleotides (example, Karam_2019, Landrith_2020). The variant was absent in 250906 control chromosomes. To our knowledge, no occurrence of c.8754+4A>T in individuals affected with Hereditary Breast And Ovarian Cancer Syndrome has been reported. Three clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 (VUS, n=1; Likely pathogenic, n=2). Based on the evidence outlined above, the variant was classified as likely pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Apr 22, 2024	This sequence change falls in intron 21 of the BRCA2 gene. It does not directly 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 not been reported in the literature in individuals affected with BRCA2-related conditions. ClinVar contains an entry for this variant (Variation ID: 220353). Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Studies have shown that this variant results in activation of a cryptic splice site and introduces a premature termination codon (PMID: 32133419; Invitae). The resulting mRNA is expected to undergo nonsense-mediated decay. This variant disrupts the c.8754+4A nucleotide in the BRCA2 gene. Other variant(s) that disrupt this nucleotide have been determined to be pathogenic (PMID: 17011978, 17924331, 20927582, 21990134, 22505045, 27433846). 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. -

Familial cancer of breast Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Baylor Genetics

Mar 03, 2023

- -

Breast-ovarian cancer, familial, susceptibility to, 2 Uncertain:1

Uncertain significance, flagged submission

clinical testing

Counsyl

Sep 07, 2016

- -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_noAF	Benign	-0.34
CADD	Benign	17
DANN	Benign	0.92

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	0.99
dbscSNV1_RF	Benign	0.58
SpliceAI score (max)		0.77		Details are displayed if max score is > 0.2
DS_DG_spliceai		0.75		Position offset: 42
DS_DL_spliceai		0.77		Position offset: -4

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

Publications

LitVar

Other links and lift over

dbSNP: rs81002893; hg19: chr13-32950932;