rs587778850

Positions:

• chr13-48473391-G-A
• chr13-48473391-G-C
• chr13-48473391-G-T

Variant summary

Our verdict is Pathogenic. Variant got 12 ACMG points: 12P and 0B. PVS1_Moderate PM2 PP5_Very_Strong

The NM_000321.3(RB1):​c.2520+1G>A variant causes a splice donor, intron 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)
  • Exomes 𝑓: 0.0 (0 hom.)
  • Failed GnomAD Quality Control
• Consequence: RB1 NM_000321.3 splice_donor, intron
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic criteria provided, multiple submitters, no conflicts P:4 O:1
• Conservation: PhyloP100: 8.77

Genes affected

RB1 (HGNC:9884): (RB transcriptional corepressor 1) The protein encoded by this gene is a negative regulator of the cell cycle and was the first tumor suppressor gene found. The encoded protein also stabilizes constitutive heterochromatin to maintain the overall chromatin structure. The active, hypophosphorylated form of the protein binds transcription factor E2F1. Defects in this gene are a cause of childhood cancer retinoblastoma (RB), bladder cancer, and osteogenic sarcoma. [provided by RefSeq, Jul 2008]

ACMG classification

Verdict is Pathogenic. Variant got 12 ACMG points.

• PVS1: Splicing +-2 bp (donor or acceptor) variant, product NOT destroyed by NMD, known LOF gene, truncates exone, which is 0.010764263 fraction of the gene. Cryptic splice site detected, with MaxEntScore 8.5, offset of 39, new splice context is: attGTaagt. Cryptic site results in inframe change. If cryptic site found is not functional and variant results in exon loss, it results in frameshift change.
• PM2: Very rare variant in population databases, with high coverage
• PP5: Variant 13-48473391-G-A is Pathogenic according to our data. Variant chr13-48473391-G-A is described in ClinVar as [Pathogenic]. Clinvar id is 126805.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
RB1	NM_000321.3	c.2520+1G>A		splice_donor_variant, intron_variant		ENST00000267163.6	NP_000312.2
RB1	NM_001407165.1	c.2520+1G>A		splice_donor_variant, intron_variant			NP_001394094.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
RB1	ENST00000267163.6	c.2520+1G>A		splice_donor_variant, intron_variant		1	NM_000321.3	ENSP00000267163.4
RB1	ENST00000650461.1	c.2520+1G>A		splice_donor_variant, intron_variant				ENSP00000497193.1
RB1	ENST00000643064.1	c.192+91948G>A		intron_variant				ENSP00000496005.1

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Data not reliable, filtered out with message: AC0 AF: 0.00 AC: 0 AN: 1409452 Hom.: 0 Cov.: 28 AF XY: 0.00 AC XY: 0 AN XY: 703868

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: 0.00

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome Cov.: 32

ClinVar

Significance: Pathogenic

Submissions summary: Pathogenic:4 Other:1

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Retinoblastoma Pathogenic:3

Pathogenic, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Oct 13, 2019	Disruption of this splice site has been observed in several individuals affected with retinoblastoma (PMID: 12541220, 18000883, 28193182, 16595082). This variant is also described as c.2658+1G>A and g.170403G>A in the literature. ClinVar contains an entry for this variant (Variation ID: 126805). For these reasons, this variant has been classified as Pathogenic. Donor and acceptor splice site variants typically lead to a loss of protein function (PMID: 16199547), and loss-of-function variants in RB1 are known to be pathogenic (PMID: 17096365). Experimental studies have shown that this variant disrupts mRNA splicing (PMID: 18000883). This variant is not present in population databases (ExAC no frequency). This sequence change affects a donor splice site in intron 24 of the RB1 gene. It is expected to disrupt RNA splicing and likely results in an absent or disrupted protein product. -
Pathogenic, criteria provided, single submitter	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Apr 27, 2018	The c.2520+1G>A variant in RB1 has been reported in at least 2 individuals with retinoblastoma (Richter 2003, Houdayer 2008) and was absent from large populatio n studies. This variant occurs in the invariant region (+/- 1,2) of the splice c onsensus sequence and is predicted to cause altered splicing leading to an abnor mal or absent protein. However, this information is not predictive enough to det ermine pathogenicity. Functional analysis using patient cell lines demonstrated that the c.2520+1G>A variant results in skipping of exon 24 (Houdayer 2008, John son 2014). This results in a frameshift, which is predicted to lead to loss of function via nonsense mediated decay (NMD). Loss of function is an established mechanism of disease for the RB1 gene. In summary, this variant meets criteria to be classified as pathogenic for retinoblastoma in an autosomal dominant manne r. ACMG criteria applied: PVS1, PM2, PS4_Supporting. -
Pathogenic, criteria provided, single submitter	clinical testing	Genetic Diagnostic Laboratory, University of Pennsylvania School of Medicine	May 20, 2024	Case and Pedigree Information: BILATERAL CASES:3, UNILATERAL CASES:0, TOTAL CASES:3, PEDIGREES:3. ACMG Codes Applied:PVS1, PM2, PS4SUP -

Hereditary cancer-predisposing syndrome Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Ambry Genetics

Dec 03, 2024

The c.2520+1G>A intronic pathogenic mutation results from a G to A substitution one nucleotide(s) after coding exon 24 of the RB1 gene. This variant has been reported in individual(s) with features consistent with RB1-related hereditary retinoblastoma (Rodríguez-Martín C et al. J Hum Genet, 2020 Jan;65:165-174; Richter S et al. Am J Hum Genet, 2003 Feb;72:253-69; Tsai T et al. Arch Ophthalmol, 2004 Feb;122:239-48; Houdayer C et al. Hum Mutat, 2004 Feb;23:193-202; Ambry internal data). In multiple assays testing RB1 function, this variant showed functionally abnormal results (Gámez-Pozo A et al. Hum Mutat, 2007 Dec;28:1245; Johnson BE et al. Science, 2014 Jan;343:189-193; Houdayer C et al. Hum Mutat, 2008 Jul;29:975-82). Of note, this variant is also referred to as c.2658+1G>A and g.170403G>A in the 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 donor site. 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. -

Neoplasm Other:1

-, criteria provided, single submitter

clinical testing

Center for Genomic Medicine, Rigshospitalet, Copenhagen University Hospital

Jul 31, 2024

- -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.63	D
BayesDel_noAF	Pathogenic	0.41
CADD	Pathogenic	34
DANN	Uncertain	1.0
Eigen	Pathogenic	1.2
Eigen_PC	Pathogenic	1.1
FATHMM_MKL	Pathogenic	1.0	D
GERP RS		5.7
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		2.8

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.93
SpliceAI score (max)		0.76		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.76		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: rs587778850; hg19: chr13-49047527; COSMIC: COSV57294549; COSMIC: COSV57294549;