rs273899694

Positions:

• chr17-43104865-T-TA

Variant summary

Our verdict is Uncertain significance. Variant got 3 ACMG points: 3P and 0B. PM2 PP3

The NM_007294.4(BRCA1):​c.301+2_301+3insT variant causes a splice region, intron change. The variant allele was found at a frequency of 0.00000274 in 1,460,016 control chromosomes in the GnomAD database, with no homozygous occurrence. 1/1 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: 0.0000027 (0 hom.)
• Consequence: BRCA1 NM_007294.4 splice_region, intron
• Clinical Significance: Uncertain significance criteria provided, multiple submitters, no conflicts P:1 U:6 B:1
• Conservation: PhyloP100: 5.49

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 Uncertain_significance. Variant got 3 ACMG points.

• PM2: Very rare variant in population databases, with high coverage
• PP3: Splicing scoreres supports a deletorius effect: Scorers claiming Pathogenic: 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+2_301+3insT		splice_region_variant, intron_variant		ENST00000357654.9

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
BRCA1	ENST00000357654.9	c.301+2_301+3insT		splice_region_variant, intron_variant		1	NM_007294.4	P4

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome AF: 0.00000274 AC: 4 AN: 1460016 Hom.: 0 Cov.: 31 AF XY: 0.00 AC XY: 0 AN XY: 726428

Gnomad4 AFR exome AF: 0.0000897

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: 9.01e-7

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome Cov.: 32

Bravo AF: 0.0000113

ClinVar

Significance: Uncertain significance

Submissions summary: Pathogenic:1 Uncertain:6 Benign:1

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

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

Uncertain significance, criteria provided, single submitter	clinical testing	All of Us Research Program, National Institutes of Health	Dec 13, 2023	This variant causes a 1-nucleotide insertion in the intron 5 splice donor site of the BRCA1 gene. Splice site prediction tools predict that this variant may have a significant impact on RNA splicing. Although this prediction has not been confirmed in published RNA studies, this variant is expected to result in an absent or disrupted protein product. However, a canonical splice site mutation c.301+1G>C that disrupts this splice donor site has been shown to cause the use of an in-frame cryptic donor site, resulting in a 3 amino acid deletion (PMID: 29750258). A different canonical splice site mutation c.301+1G>A also was reported as not disease-causing based on carrier family history (PMID: 28408614) and it has been found with a BRCA2 truncation variant expected to result in loss-of-function in two individuals with personal and/or family history of breast and/or ovarian cancer (PMID: 27836010, 28526081). These observations for the c.301+1G variants suggest that the deleterious splicing impact of this variant, c.301+2dup, could be ameliorated by alternative splicing. To our knowledge, functional and RNA studies have not been reported for this variant nor has this variant been reported in individuals affected with BRCA1-related disorders in the literature. This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). The available evidence is insufficient to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance. -
Uncertain significance, criteria provided, single submitter	clinical testing	Baylor Genetics	Mar 22, 2024	- -
Pathogenic, no assertion criteria provided	clinical testing	Breast Cancer Information Core (BIC) (BRCA1)	Feb 16, 2001	- -
Likely benign, no assertion criteria provided	clinical testing	Sharing Clinical Reports Project (SCRP)	Aug 28, 2012	- -

not specified Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Women's Health and Genetics/Laboratory Corporation of America, LabCorp

Jul 04, 2022

Variant summary: BRCA1 c.301+2dupT 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: Four predict the variant abolishes the canonical 5' splicing donor site. However, these predictions have yet to be confirmed by functional studies. The variant was absent in 251180 control chromosomes. The available data on variant occurrences in the general population are insufficient to allow any conclusion about variant significance. c.301+2dupT has been reported in the literature as c.301+2_301+3insT in at-least one individual of African American ancestry citing the BIC database (example, Friebel_2019). These report(s) do not provide unequivocal conclusions about association of the variant with Hereditary Breast And Ovarian Cancer Syndrome. At-least one co-occurrences with another pathogenic variant(s) have been observed at our laboratory (APC c.1958+3A>G), providing supporting evidence for a benign role. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. Three clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. All laboratories classified the variant as uncertain significance. Based on the evidence outlined above, the variant was classified as uncertain significance. -

not provided Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Quest Diagnostics Nichols Institute San Juan Capistrano

May 17, 2018

- -

Hereditary cancer-predisposing syndrome Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Ambry Genetics

Apr 13, 2023

The c.301+2dupT intronic variant results from a duplication of the T nucleotide located two nucleotides after coding exon 4 of the BRCA1 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. 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 an alteration with a similar splicing impact 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. -

Hereditary breast ovarian cancer syndrome Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Invitae

Oct 13, 2023

This sequence change falls in intron 5 of the BRCA1 gene. It does not directly change the encoded amino acid sequence of the BRCA1 protein. RNA analysis indicates that this variant induces altered splicing and likely results in the loss of 4 amino acid residue(s), but is expected to preserve the integrity of the reading-frame. This variant is not present in population databases (gnomAD no frequency). This variant has not been reported in the literature in individuals affected with BRCA1-related conditions. ClinVar contains an entry for this variant (Variation ID: 125664). 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 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

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.67		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.67		Position offset: 3

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

Publications

LitVar

Other links and lift over

dbSNP: rs273899694; hg19: chr17-41256882;