17-43104867-C-T

Position:

• chr17-43104867-C-T

Variant summary

Our verdict is Likely pathogenic. Variant got 6 ACMG points: 6P and 0B. PVS1_Moderate PP3_Strong

The NM_007294.4(BRCA1):​c.301+1G>A variant causes a splice donor change. The variant allele was found at a frequency of 0.00000479 in 1,460,460 control chromosomes in the GnomAD database, with no homozygous occurrence. 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 Uncertain significance (★★).

• Frequency:
  • Genomes: not found (cov: 32)
  • Exomes 𝑓: 0.0000048 (0 hom.)
• Consequence: BRCA1 NM_007294.4 splice_donor
• Scores: Splicing: ADA: 0.9999
• Clinical Significance: Uncertain significance criteria provided, multiple submitters, no conflicts P:1 U:9 O:1
• Conservation: PhyloP100: 4.63

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 Likely_pathogenic. Variant got 6 ACMG points.

• PVS1: Splicing +-2 bp (donor or acceptor) variant, product NOT destroyed by NMD, known LOF gene, truncates exone, which is 0.015736766 fraction of the gene. Cryptic splice site detected, with MaxEntScore 5.6, offset of -9, new splice context is: cagGTttgg. Cryptic site results in inframe change. If cryptic site found is not functional and variant results in exon loss, it results in frameshift change.
• PP3: Splicing scoreres supports a deletorius effect: Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF, max_spliceai. No scorers claiming Uncertain. No scorers claiming Benign.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
BRCA1	NM_007294.4	c.301+1G>A		splice_donor_variant		ENST00000357654.9

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
BRCA1	ENST00000357654.9	c.301+1G>A		splice_donor_variant		1	NM_007294.4	P4

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD3 exomes AF: 0.00000398 AC: 1 AN: 251180 Hom.: 0 AF XY: 0.00000737 AC XY: 1 AN XY: 135766

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: 0.00000881

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.00000479 AC: 7 AN: 1460460 Hom.: 0 Cov.: 30 AF XY: 0.00000413 AC XY: 3 AN XY: 726646

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: 0.00000630

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome Cov.: 32

ExAC AF: 0.00000824 AC: 1

ClinVar

Significance: Uncertain significance

Submissions summary: Pathogenic:1 Uncertain:9 Other:1

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

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

Pathogenic, flagged submission	clinical testing	Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA), c/o University of Cambridge	Oct 02, 2015	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Baylor Genetics	Jan 19, 2024	- -
Uncertain significance, criteria provided, single submitter	clinical testing	All of Us Research Program, National Institutes of Health	Jan 11, 2024	- -
not provided, no classification provided	in vitro	Brotman Baty Institute, University of Washington	-	- -

not provided Uncertain:3

Uncertain significance, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Oct 27, 2017	- -
Uncertain significance, criteria provided, single submitter	clinical testing	GeneDx	Apr 13, 2022	Canonical splice site variant with an unclear effect on protein function; a different variant at the same splice site, c.301+1G>C, demonstrated to result in an in-frame isoform that replaces 3 amino acids with 1 incorrect one in the critical RING domain (Borg 2010, Paul 2014, Leman 2018); Observed in individuals with breast, pancreatic, or prostate cancer (Rebbeck 2016, Na 2017, Alimirzaie 2018); Not observed at significant frequency in large population cohorts (gnomAD); Also known as c.420+1G>A and IVS6+1G>A; This variant is associated with the following publications: (PMID: 29940740, 25085752, 21769658, 27836010, 28526081, 29446198, 27989354, 24389207, 20104584, 30209399, 29750258) -
Uncertain significance, criteria provided, single submitter	clinical testing	Revvity Omics, Revvity	Apr 07, 2022	- -

Hereditary cancer-predisposing syndrome Uncertain:2

Uncertain significance, criteria provided, single submitter	clinical testing	Ambry Genetics	Nov 02, 2023	The c.301+1G>A intronic variant results from a G to A substitution one nucleotide after coding exon 4 of the BRCA1 gene. This variant has been reported in individuals with breast, prostate, and pancreatic cancer (Rebbeck TR et al. Breast Cancer Res, 2016 11;18(1):112; Na R et al. Eur Urol, 2017 05;71:740-747; Alimirzaie S et al. Arch Iran Med, 2018 06;21:228-233), as well as individuals from control groups in multiple studies (Hu C et al. JAMA, 2018 Jun;319:2401-2409; Dorling et al. N Engl J Med. 2021 02;384:428-439; Nguyen-Dumont T et al. Cancers (Basel), 2021 Mar;13:; Southey MC et al. NPJ Breast Cancer, 2021 Dec;7:153). 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. Alterations at this donor site are known to produce an in frame transcript impacting four amino acids in the BRCA1 RING domain (Ambry internal data). One functional study found that nucleotide substitutions at this donor site are non-functional in a high-throughput, genome editing, haploid cell survival assay (Findlay GM et al. Nature, 2018 10;562:217-222). However, this alteration has also been identified in trans with a pathogenic mutation in BRCA1 in an individual without features of Fanconi Anemia (Ambry internal data, personal communication). Since supporting evidence is conflicting at this time, the clinical significance of this alteration remains unclear. -
Uncertain significance, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Dec 08, 2023	This variant changes a G to A nucleotide at the +1 position of the intron 5 splice donor site in the BRCA1 gene. Splice prediction tools indicate a significant disruption to the native splice donor site and the possible activation of a cryptic donor site, resulting in an in-frame deletion at the end of exon 5. The detection of in-frame deletion in the RNA of carriers of this variant has been reported (ClinVar accession: SCV000185671.7, SCV000635877.6). An RNA study for a different variant at this splice donor site, c.301+6T>C, has shown a disruption at the native donor site that is rescued by the use of a cryptic donor site resulting in the in-frame deletion of 9 nucleotides from the end of exon 5 (PMID: 21769658). A functional study has reported that this variant c.301+1G>A impacts BRCA1 function in a haploid cell proliferation assay (PMID: 30209399). This variant has been reported in one individual affected with pancreatic cancer (PMID: 29940740). However, this variant has been detected in a breast cancer case-control meta-analysis in 1/53461 unaffected individuals and absent in 60466 cases (PMID: 33471991; Leiden Open Variation Database DB-ID BRCA1_005049). This variant has been detected in an individual affected with breast cancer who has a BRCA2 truncation variant that is expected to be loss-of-function and explains the disease (PMID: 27836010), and it also has been observed in trans with a pathogenic BRCA1 variant in an individual without features of Fanconi anemia (ClinVar accession: SCV000185671.7). Another commercial laboratory also has reported this variant as benign based in part on health history evidence (PMID: 28408614). This variant has been identified in 1/251180 chromosomes in the general population by the Genome Aggregation Database (gnomAD). Although there is a suspicion that this variant may not be associated with disease, additional studies are necessary to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance. -

not specified Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine

May 22, 2017

The c.301+1G>A variant in BRCA1 has been reported in 1 Caucasian woman with brea st cancer who also carried another pathogenic variant in BRCA2 (Rebbeck 2016). T his variant has been identified in 1/111,504 European chromosomes by the genome Aggregation Database (gnomAD, http://gnomad.broadinstitute.org/; dbSNP rs5877821 73). This variant occurs in the invariant region (+/- 1,2) of the splice consens us sequence and variants in this region are predicted to cause altered splicing leading to an abnormal or absent protein. However, in vitro functional studies f or another variant at this exon-intron junction (301+6T>C) suggest the use of an alternative splice donor site resulting in an in-frame deletion of three amino acids (Thomassen 2012). Please note, these types of assays may not accurately re present biological function. In summary, due to conflicting and insufficient evi dence, the clinical significance of the c.301+1G>A variant is uncertain. -

Hereditary breast ovarian cancer syndrome Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Invitae

Oct 24, 2023

This sequence change affects a donor splice site in intron 5 of the BRCA1 gene. RNA analysis indicates that disruption of this splice site induces altered splicing and likely results in the loss of 4 and insertion of 1 amino acid residue(s), but is expected to preserve the integrity of the reading-frame. This variant is present in population databases (rs587782173, gnomAD 0.0009%). Disruption of this splice site has been observed in individual(s) with BRCA1-related conditions (PMID: 27836010, 29446198). ClinVar contains an entry for this variant (Variation ID: 142004). Algorithms developed to predict the effect of variants on protein structure and function are not available or were not evaluated for this variant. Experimental studies have shown that disruption of this splice site affects BRCA1 function (PMID: 30209399). Studies have shown that disruption of this splice site results in the activation of a cryptic splice site in exon 5 (Invitae). In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.36	D
BayesDel_noAF	Pathogenic	0.29
CADD	Pathogenic	29
DANN	Uncertain	0.99
Eigen	Pathogenic	1.0
Eigen_PC	Pathogenic	0.85
FATHMM_MKL	Pathogenic	0.99	D
MutationTaster	Benign	1.0	D;D;D;D;D;D;D;D;D;D;D;D;D;D
GERP RS		5.3
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)		0.67		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.67		Position offset: 1

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

Publications

LitVar

Other links and lift over

dbSNP: rs587782173; hg19: chr17-41256884;