13-48459708-C-T

Position:

• chr13-48459708-C-T

Variant summary

Our verdict is Pathogenic. Variant got 18 ACMG points: 18P and 0B. PM1 PM2 PM5 PP3_Strong PP5_Very_Strong

The NM_000321.3(RB1):​c.1981C>T​(p.Arg661Trp) variant causes a missense change. The variant allele was found at a frequency of 0.00000137 in 1,461,854 control chromosomes in the GnomAD database, with no homozygous occurrence. In-silico tool predicts a pathogenic outcome for this variant. Variant has been reported in ClinVar as Pathogenic (★★). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. R661Q) has been classified as Likely pathogenic.

• Frequency:
  • Genomes: not found (cov: 30)
  • Exomes 𝑓: 0.0000014 (0 hom.)
• Consequence: RB1 NM_000321.3 missense
• Clinical Significance: Pathogenic criteria provided, multiple submitters, no conflicts P:11 O:2
• Conservation: PhyloP100: 4.65

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 18 ACMG points.

• PM1: In a helix (size 24) in uniprot entity RB_HUMAN there are 11 pathogenic changes around while only 1 benign (92%) in NM_000321.3
• PM2: Very rare variant in population databases, with high coverage
• PM5: Other missense variant is known to change same aminoacid residue: Variant chr13-48459709-G-A is described in Lovd as [Likely_pathogenic].
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.956
• PP5: Variant 13-48459708-C-T is Pathogenic according to our data. Variant chr13-48459708-C-T is described in ClinVar as [Pathogenic]. Clinvar id is 13087.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.1981C>T	p.Arg661Trp	missense_variant	20/27	ENST00000267163.6	NP_000312.2
RB1	NM_001407165.1	c.1981C>T	p.Arg661Trp	missense_variant	20/27		NP_001394094.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
RB1	ENST00000267163.6	c.1981C>T	p.Arg661Trp	missense_variant	20/27	1	NM_000321.3	ENSP00000267163.4
RB1	ENST00000650461.1	c.1981C>T	p.Arg661Trp	missense_variant	20/27			ENSP00000497193.1
RB1	ENST00000643064.1	c.192+78265C>T		intron_variant				ENSP00000496005.1

Frequencies

GnomAD3 genomes Cov.: 30

GnomAD3 exomes AF: 0.0000159 AC: 4 AN: 251324 Hom.: 0 AF XY: 0.00000736 AC XY: 1 AN XY: 135856

Gnomad AFR exome AF: 0.0000616

Gnomad AMR exome AF: 0.00

Gnomad ASJ exome AF: 0.0000992

Gnomad EAS exome AF: 0.00

Gnomad SAS exome AF: 0.00

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.0000176

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.00000137 AC: 2 AN: 1461854 Hom.: 0 Cov.: 34 AF XY: 0.00000138 AC XY: 1 AN XY: 727232

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: 0.00000180

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome Cov.: 30

ExAC AF: 0.00000824 AC: 1

ClinVar

Significance: Pathogenic

Submissions summary: Pathogenic:11 Other:2

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Retinoblastoma Pathogenic:6 Other:1

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:5, UNILATERAL CASES:7, TOTAL CASES:12, PEDIGREES:12. ACMG Codes Applied:PM2, PS4M -
Pathogenic, no assertion criteria provided	literature only	OMIM	Oct 01, 1999	- -
Pathogenic, criteria provided, single submitter	research	Department of Pediatrics, Memorial Sloan Kettering Cancer Center	Dec 15, 2020	- -
Pathogenic, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Sep 30, 2023	This sequence change replaces arginine, which is basic and polar, with tryptophan, which is neutral and slightly polar, at codon 661 of the RB1 protein (p.Arg661Trp). This variant is present in population databases (rs137853294, gnomAD 0.01%). This variant has been reported in many individuals affected with retinoblastoma (PMID: 12541220, 16269091, 23532519, 28575107, 24225018). It has been reported to segregate with retinoblastoma in multiple families (PMID: 1352883, 26925970, 17096365). While all tested affected individuals in the families had this variant, penetrance was reduced in comparison to truncating variants in RB1 seen in other families, with relatively mild phenotypic expression observed in some cases. Penetrance appears to be lower when this variant is inherited from the mother than from the father (PMID: 26925970). ClinVar contains an entry for this variant (Variation ID: 13087). Advanced modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) performed at Invitae indicates that this missense variant is expected to disrupt RB1 protein function. Experimental studies have shown that this missense change has partial activity. It retains some ability to suppress retinoblastoma development, but is unstable with temperature-sensitive pocket protein-binding activity and defective in several aspects of cell cycle control (PMID: 18677112, 18682685, 10486322, 16449662, 15643604, 9632788). For these reasons, this variant has been classified as Pathogenic. -
not provided, no classification provided	phenotyping only	GenomeConnect, ClinGen	-	Variant classified as Pathogenic and reported on 08-06-2021 by Lab or GTR ID 500031. GenomeConnect assertions are reported exactly as they appear on the patient-provided report from the testing laboratory. GenomeConnect staff make no attempt to reinterpret the clinical significance of the variant. -
Pathogenic, criteria provided, single submitter	clinical testing	Genetics and Molecular Pathology, SA Pathology	Jul 24, 2019	This variant has been shown to be associated with the methylation status at CpG85 in RB1 intron 2 that is differentially methylated depending on parent -of -origin. Paternally transmitted mutation has low residual activity mimics a null mutation leading to the development of retinoblastoma. -
Pathogenic, criteria provided, single submitter	clinical testing	St. Jude Molecular Pathology, St. Jude Children's Research Hospital	Oct 25, 2023	The RB1 c.1981C>T (p.Arg661Trp) missense change has a maximum non-founder subpopulation frequency of 0.0060% in gnomAD v2.1.1 (https://gnomad.broadinstitute.org). This variant has been identified in individuals with retinoblastoma (PMID: 12541220, 16269091, 23532519, 24225018, 28575107) and has been found to segregate with disease in affected family members (PMID: 1352883, 26925970, 17096365). Interestingly, penetrance appears to be reduced when compared to truncating variants in RB1. It is thought to be associated with differential penetrance based on the sex of the transmitted parent, where paternally inherited alleles appear to show increased penetrance (PMID: 26925970). The in silico tool REVEL predicts a deleterious effect on protein function, and functional studies have shown that this variant is partially functional (PMID: 9632788, 10486322, 15643604, 16449662, 18677112, 18682685). In summary, this variant meets criteria to be classified as pathogenic. -

not provided Pathogenic:3

Pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Aug 30, 2024	Studies suggest this variant is associated with incomplete penetrance/reduced expression and a parent-of-origin effect: up to a 67.5% risk of retinoblastoma when paternally inherited and an increased risk for second primary cancers, most commonly sarcoma, leukemia, and melanoma; Published functional studies demonstrate reduced efficiency of Rb protein function without abolishing it (PMID: 9342358, 10486322, 9632788); Not observed at significant frequency in large population cohorts (gnomAD); In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; This variant is associated with the following publications: (PMID: 17960112, 9671401, 15643604, 18677112, 29568217, 34190019, 9342358, 1352883, 28575107, 12541220, 16269091, 9632788, 23532519, 16449662, 29662154, 34294096, 34277001, 34645364, 33057194, 35982159, 35960463, 36274096, 33466343, 28724667, 34308366, 31980526, 10486322, 24225018, Day alan_2006_Review, 31772335, 32191290, 26925970, 9311732, 17096365) -
Pathogenic, criteria provided, single submitter	clinical testing	Eurofins Ntd Llc (ga)	Nov 18, 2013	- -
Pathogenic, criteria provided, single submitter	clinical testing	CeGaT Center for Human Genetics Tuebingen	Nov 01, 2024	RB1: PP1:Strong, PM1, PM2, PS3:Moderate, PS4:Moderate -

Hereditary cancer-predisposing syndrome Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Ambry Genetics

Jul 21, 2022

The p.R661W pathogenic mutation with reduced penetrance (also known as c.1981C>T), located in coding exon 20 of the RB1 gene, results from a C to T substitution at nucleotide position 1981. The arginine at codon 661 is replaced by tryptophan, an amino acid with dissimilar properties. This alteration has been reported in several RB families showing reduced penetrance where there are some individuals who are unaffected carriers; where some have unilateral RB; where some have bilateral RB; and some have regressed RB tumors (Onadim Z et al. Proc. Natl. Acad. Sci. U.S.A.. 1992 Jul;89:6177-81; Dommering CJ et al. J. Med. Genet. 2014 Jun;51:366-74; Eloy P et al. PLoS Genet. 2016 Feb;12:e1005888). In another study, the p.R661W pathogenic mutation accounted for approximately 3% (7/235) of unrelated retinoblastoma probands with germline RB1 mutations and approximately 13% (4/30) of germline mutations identified in probands with unilateral retinoblastoma (Richter S et al. Am. J. Hum. Genet. 2003 Feb;72:253-69). In addition, the p.R661W alteration has been shown to result in reduced, but not abolished, retinoblastoma protein activity (Whitaker LL et al. Mol. Cell. Biol. 1998 Jul;18:4032-42) and confers a decreased, albeit significant tumor risk (DiCiommo D et al. Semin. Cancer Biol. 2000 Aug;10:255-69). In a recent study including ten families with the p.R661W alteration, authors reported that if this alteration was maternally inherited the probability of being affected with RB was just under 10% whereas if this alteration was paternally inherited the probability of being affected with RB was almost 68%. They speculate that this parent-of-origin effect is due to maternal imprinting of an internal promoter which produces an alternative RB1 transcript and may contribute to the reduced penetrance of this mutation (Eloy P et al. PLoS Genet. 2016 Feb;12:e1005888; Kanber D et al. PLoS Genet. 2009 Dec;5:e1000790). Based on the supporting evidence, p.R661W is interpreted as a disease-causing mutation with reduced penetrance. -

Malignant tumor of urinary bladder;C0035335:Retinoblastoma;C0149925:Small cell lung carcinoma;C0585442:Bone osteosarcoma Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Fulgent Genetics, Fulgent Genetics

Oct 31, 2018

- -

Vulvar adenocarcinoma of mammary gland type Other:1

-, no assertion criteria provided

research

Genome Sciences Centre, British Columbia Cancer Agency

Nov 13, 2024

- -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.99
BayesDel_addAF	Pathogenic	0.40	D
BayesDel_noAF	Pathogenic	0.51
CADD	Pathogenic	33
DANN	Pathogenic	1.0
DEOGEN2	Pathogenic	0.98	D;.
Eigen	Pathogenic	0.88
Eigen_PC	Pathogenic	0.80
FATHMM_MKL	Uncertain	0.96	D
LIST_S2	Pathogenic	0.97	D;D
M_CAP	Uncertain	0.22	D
MetaRNN	Pathogenic	0.96	D;D
MetaSVM	Pathogenic	1.1	D
MutationAssessor	Pathogenic	3.4	M;.
PrimateAI	Pathogenic	0.80	T
PROVEAN	Pathogenic	-7.2	D;.
REVEL	Pathogenic	0.94
Sift	Pathogenic	0.0	D;.
Sift4G	Pathogenic	0.0	D;.
Polyphen		1.0	D;.
Vest4		0.99
MutPred		0.85		Gain of catalytic residue at Y659 (P = 0.0164);Gain of catalytic residue at Y659 (P = 0.0164);
MVP		0.97
MPC		1.3
ClinPred		0.97	D
GERP RS		5.7
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.7
Varity_R		0.96
gMVP		0.95

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.010		Details are displayed if max score is > 0.2

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

Publications

LitVar

Other links and lift over

dbSNP: rs137853294; hg19: chr13-49033844; COSMIC: COSV57295487; COSMIC: COSV57295487;