17-43057137-T-C

Variant summary

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

The NM_007294.4(BRCA1):c.5194-2A>G variant causes a splice acceptor change. 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 Likely pathogenic (★★).

• Frequency: Genomes: not found (cov: 31)
• Consequence: BRCA1 NM_007294.4 splice_acceptor
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:16 O:1
• Conservation: PhyloP100: 4.74

Genes affected

BRCA1 (HGNC:1100): (BRCA1 DNA repair associated) This gene encodes a 190 kD nuclear phosphoprotein that plays a role in maintaining genomic stability, and it also acts as a tumor suppressor. The BRCA1 gene contains 22 exons spanning about 110 kb of DNA. The encoded protein combines with other tumor suppressors, DNA damage sensors, and signal transducers to form a large multi-subunit protein complex known as the BRCA1-associated genome surveillance complex (BASC). This gene product associates with RNA polymerase II, and through the C-terminal domain, also interacts with histone deacetylase complexes. This protein thus plays a role in transcription, DNA repair of double-stranded breaks, and recombination. Mutations in this gene are responsible for approximately 40% of inherited breast cancers and more than 80% of inherited breast and ovarian cancers. Alternative splicing plays a role in modulating the subcellular localization and physiological function of this gene. Many alternatively spliced transcript variants, some of which are disease-associated mutations, have been described for this gene, but the full-length natures of only some of these variants has been described. A related pseudogene, which is also located on chromosome 17, has been identified. [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 0.014842632 fraction of the gene. Cryptic splice site detected, with MaxEntScore 7.3, offset of -49, new splice context is: ctgctccacttccattgaAGgaa. 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 17-43057137-T-C is Pathogenic according to our data. Variant chr17-43057137-T-C is described in ClinVar as [Likely_pathogenic]. Clinvar id is 37647.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars. Variant chr17-43057137-T-C is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
BRCA1	NM_007294.4	c.5194-2A>G		splice_acceptor_variant		ENST00000357654.9

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
BRCA1	ENST00000357654.9	c.5194-2A>G		splice_acceptor_variant		1	NM_007294.4	P4

Frequencies

GnomAD3 genomes Cov.: 31

GnomAD3 exomes AF: 0.00000398 AC: 1 AN: 251484 Hom.: 0 AF XY: 0.00000736 AC XY: 1 AN XY: 135912

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.00

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.00

Gnomad SAS exome AF: 0.00

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.00000879

Gnomad OTH exome AF: 0.00

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 31

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:16 Other:1

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Breast-ovarian cancer, familial, susceptibility to, 1 Pathogenic:6 Other:1

Pathogenic, no assertion criteria provided	clinical testing	Sharing Clinical Reports Project (SCRP)	Apr 11, 2011	- -
not provided, no classification provided	in vitro	Brotman Baty Institute, University of Washington	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	University of Washington Department of Laboratory Medicine, University of Washington	Nov 20, 2015	- -
Pathogenic, criteria provided, single submitter	clinical testing	Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA), c/o University of Cambridge	Oct 02, 2015	- -
Pathogenic, criteria provided, single submitter	clinical testing	All of Us Research Program, National Institutes of Health	Oct 09, 2023	This variant disrupts a canonical splice site and is predicted to result in abnormal splicing. Aberrant splicing and/or loss of function is an established mechanism of disease. Functional studies suggest that this variant results in a deleterious effect on the protein (PMID: 30209399). This variant has been reported in multiple individuals with hereditary breast and ovarian cancer (PMID: 14684619, 26845104, 29446198, 31771539, 32566972, 35918668). This variant is present in 1/251484 total alleles in the Genome Aggregation Database (http://gnomad.broadinstitute.org/). -
Pathogenic, criteria provided, single submitter	clinical testing	Baylor Genetics	May 16, 2023	- -
Pathogenic, no assertion criteria provided	clinical testing	Breast Cancer Information Core (BIC) (BRCA1)	Jun 21, 1999	- -

Hereditary breast ovarian cancer syndrome Pathogenic:3

Pathogenic, no assertion criteria provided	research	Research Molecular Genetics Laboratory, Women's College Hospital, University of Toronto	Jan 31, 2014	- -
Pathogenic, criteria provided, single submitter	clinical testing	Invitae	Dec 24, 2023	This sequence change affects an acceptor splice site in intron 18 of the BRCA1 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 present in population databases (rs80358069, gnomAD 0.0009%). Disruption of this splice site has been observed in individual(s) with breast cancer (PMID: 10923033, 14684619, 21520333, 26845104). It has also been observed to segregate with disease in related individuals. This variant is also known as 5313-2A>G or IVS19-2A>G. ClinVar contains an entry for this variant (Variation ID: 37647). Studies have shown that disruption of this splice site results in activation of a cryptic splice site and introduces a premature termination codon (Invitae). The resulting mRNA is expected to undergo nonsense-mediated decay. For these reasons, this variant has been classified as Pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Oct 29, 2019	Variant summary: BRCA1 c.5194-2A>G is located in a canonical splice-site and is predicted to affect mRNA splicing resulting in a significantly altered protein due to either exon skipping, shortening, or inclusion of intronic material. 5/5 computational tools predict that the variant abolishes a 3' acceptor site. However, these predictions have yet to be confirmed by functional studies. The variant allele was found at a frequency of 4e-06 in 251484 control chromosomes (gnomAD). c.5194-2A>G has been reported in the literature in multiple individuals affected with Hereditary Breast and Ovarian Cancer (e.g. Esteban-Cardenosa_2004, Shirts_2015, Rebbeck_2018). These data indicate that the variant is very likely to be associated with disease. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. Five other clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar (evaluation after 2014) and cited the variant as pathogenic/likely pathogenic. Based on the evidence outlined above, the variant was classified as pathogenic. -

not provided Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Jan 27, 2022	Canonical splice site variant demonstrated to result in full length transcript as well as the in-frame deletion of exon 19, located in the critical BRCT1 and BRCT2 domains (Chevalier 2020, Billaud 2021, UniProt); Observed in individuals with BRCA1-related cancers in the published literature (Shirts 2016, Park 2018, Rebbeck 2018, Wang 2019); Published functional studies demonstrate a damaging effect: variant classified as non-functional based on a saturation genome editing (SGE) assay measuring cell survival (Findlay 2018); In silico analysis supports a deleterious effect on splicing; Not observed at significant frequency in large population cohorts (gnomAD); Also known as 5313-2A>G and IVS19-2A>G; This variant is associated with the following publications: (PMID: 28152038, 32124385, 14684619, 21523855, 26845104, 25085752, 25525159, 28111427, 31589614, 34749799, 30968603, 29673794, 29446198, 32341426, 33720054, 30209399) -
Pathogenic, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Jul 03, 2023	The BRCA1 c.5194-2A>G variant (also known as IVS19-2A>G and 5313-2A>G) disrupts a canonical splice-acceptor site, and is predicted to result in the in-frame skipping of exon 19 and removal of a portion of the gene important for its structure or function. In the published literature, this variant has been reported in individuals with breast and/or ovarian cancer (PMIDs: 35918668 (2022), 32341426 (2020), 30968603 (2019), 29673794 (2018), 26845104 (2016), 14684619 (2004)). A saturation genome editing assay measuring cell survival classified this variant as non-functional (PMID: 30209399 (2018)). The frequency of this variant in the general population, 0.000004 (1/251484 chromosomes (Genome Aggregation Database, http://gnomad.broadinstitute.org)), is consistent with pathogenicity. Based on the available information, this variant is classified as pathogenic. -

Hereditary cancer-predisposing syndrome Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Feb 28, 2023	This variant causes an A to G nucleotide substitution at the -2 position of intron 18 of the BRCA1 gene. Splice site prediction tools predict that this variant may have a significant impact on RNA splicing. Functional studies have reported conflicting results that this variant caused intermediate to loss of BRCA1 activity in haploid human cell proliferation assays (PMID: 30209399, 34749799). This variant has been reported in multiple individuals affected with breast cancer (PMID: 14684619, 22333603, 26845104, 30968603, 32341426; Color internal data). This variant has been identified in 1/251484 chromosomes in the general population by the Genome Aggregation Database (gnomAD). Loss of BRCA1 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Likely Pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Ambry Genetics	May 15, 2023	The c.5194-2A>G intronic pathogenic mutation results from an A to G substitution two nucleotides upstream from coding exon 18 in the BRCA1 gene. This mutation was detected in 2/82 Norwegian patients with breast and/or ovarian cancer who also had a family history that was concerning for hereditary breast and ovarian cancer (HBOC) syndrome (Møller P et al. Eur. J. Cancer. 2001 May;37:1027-32). One functional study found that this nucleotide substitution is non-functional in a high-throughput, genome editing, haploid cell survival assay (Findlay GM et al. Nature. 2018 Oct;562(7726):217-222). 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; however, direct evidence is insufficient at this time (Ambry internal data). In addition to the clinical data presented in the literature, alterations that disrupt the canonical splice site are expected to cause aberrant splicing, resulting in an abnormal protein or a transcript that is subject to nonsense-mediated mRNA decay. As such, this alteration is classified as a disease-causing mutation. -

Breast and/or ovarian cancer Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

CHEO Genetics Diagnostic Laboratory, Children's Hospital of Eastern Ontario

Sep 18, 2020

- -

Malignant tumor of breast Pathogenic:1

Pathogenic, no assertion criteria provided

clinical testing

Department of Pathology and Laboratory Medicine, Sinai Health System

-

The c.5194-2A>G variant has been previously reported as disease-causing in the BIC and UMD databases and in the literature (Esteban-Cardeñosa 2003), however, no normal population controls were included in this study. This variant is predicted to cause abnormal splicing because the nucleotide substitution occurs in the invariant -1 and -2 positions of the splice consensus sequence located in the 3' splice region of BRCA1. It is listed in the dbSNP database (ID#: rs80358069) but no frequency information was provided, and so the prevalence of this variant in the general population is not known. In summary, based on the above information this variant is classified as pathogenic. -

Familial cancer of breast Pathogenic:1

Likely pathogenic, no assertion criteria provided

literature only

Center for Precision Medicine, Meizhou People's Hospital

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Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.27	D
BayesDel_noAF	Pathogenic	0.15
Cadd	Pathogenic	35
Dann	Uncertain	0.99
Eigen	Pathogenic	0.99
Eigen_PC	Pathogenic	0.82
FATHMM_MKL	Uncertain	0.93	D
MutationTaster	Benign	1.0	D;D;D;D;D;D;D;D;D;D;D;D;D;D
GERP RS		5.1
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.1

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.94
SpliceAI score (max)		1.0		Details are displayed if max score is > 0.2
DS_AG_spliceai		0.69		Position offset: -15
DS_AL_spliceai		1.0		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: rs80358069; hg19: chr17-41209154;