13-48456349-G-C

Variant names:

• chr13-48456349-G-C
• rs483352690
• NM_000321.3:c.1960G>C

Variant summary

Our verdict is Pathogenic. The variant received 18 ACMG points: 18P and 0B. PM1 PM2 PM5 PP3_Strong PP5_Very_Strong

The NM_000321.3(RB1):​c.1960G>C​(p.Val654Leu) variant causes a missense, splice region change. The variant was absent in control chromosomes in GnomAD project. 2/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Pathogenic (★★). Another nucleotide change resulting in the same amino acid substitution has been previously reported as Likely pathogenic in ClinVar. Another variant affecting the same amino acid position, but resulting in a different missense (i.e. V654G) has been classified as Uncertain significance.

• Frequency: Genomes: not found (cov: 32)
• Consequence: RB1 NM_000321.3 missense, splice_region
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic criteria provided, multiple submitters, no conflicts P:3
• Conservation: PhyloP100: 5.92
• Publications: 33 publications found

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]

RB1 Gene-Disease associations (from GenCC):

• hereditary retinoblastoma

  Inheritance: AD Classification: DEFINITIVE, STRONG, SUPPORTIVE Submitted by: Labcorp Genetics (formerly Invitae), Orphanet, G2P, Ambry Genetics
• retinoblastoma

  Inheritance: AD Classification: DEFINITIVE Submitted by: ClinGen
• melanoma

  Inheritance: AD Classification: MODERATE Submitted by: Ambry Genetics

ACMG classification

Our verdict: Pathogenic. The variant received 18 ACMG points.

• PM1: In a hotspot region, there are 4 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 3 benign, 24 uncertain 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 chr13-48456349-G-A is described in ClinVar as Pathogenic. ClinVar VariationId is 100808.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars.
• PP3: Splicing predictors support a deleterious effect. Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF. Scorers claiming Uncertain: max_spliceai. No scorers claiming Benign.
• PP5: Variant 13-48456349-G-C is Pathogenic according to our data. Variant chr13-48456349-G-C is described in ClinVar as Pathogenic. ClinVar VariationId is 410929.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
RB1	NM_000321.3	c.1960G>C	p.Val654Leu	missense_variant, splice_region_variant	Exon 19 of 27	ENST00000267163.6	NP_000312.2
RB1	NM_001407165.1	c.1960G>C	p.Val654Leu	missense_variant, splice_region_variant	Exon 19 of 27		NP_001394094.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
RB1	ENST00000267163.6	c.1960G>C	p.Val654Leu	missense_variant, splice_region_variant	Exon 19 of 27	1	NM_000321.3	ENSP00000267163.4

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 32

ClinVar

Significance: Pathogenic

Submissions summary: Pathogenic:3

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Retinoblastoma Pathogenic:2

May 20, 2024

Genetic Diagnostic Laboratory, University of Pennsylvania School of Medicine

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Case and Pedigree Information: BILATERAL CASES:3, UNILATERAL CASES:2, TOTAL CASES:5, PEDIGREES:4 (one pedigree contains both unilateral and bilateral cases). ACMG Codes Applied:PVS1, PM2, PS4SUP -

Jun 25, 2024

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This sequence change affects codon 654 of the RB1 mRNA. It is a 'silent' change, meaning that it does not change the encoded amino acid sequence of the RB1 protein. RNA analysis indicates that this variant induces altered splicing and may result in an absent or disrupted protein product. This variant is not present in population databases (gnomAD no frequency). This variant has been observed in individual(s) with retinoblastoma (PMID: 15605413, 19280657, 36729443). In at least one individual the variant was observed to be de novo. ClinVar contains an entry for this variant (Variation ID: 410929). An algorithm developed to predict the effect of missense changes on protein structure and function (PolyPhen-2) suggests that this variant is likely to be disruptive. 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 skipping of exon 19 and introduces a premature termination codon (Invitae). The resulting mRNA is expected to undergo nonsense-mediated decay. This variant disrupts the c.1960G nucleotide in the RB1 gene. Other variant(s) that disrupt this nucleotide have been determined to be pathogenic (PMID: 14722923, 15884040, 22084214, 26925970, 29568217). This suggests that this nucleotide is clinically significant, and that variants that disrupt this position are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic. -

Hereditary cancer-predisposing syndrome Pathogenic:1

Nov 12, 2018

Ambry Genetics

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The c.1960G>C pathogenic mutation (also known as p.V654L), located in coding exon 19 of the RB1 gene, results from a G to C substitution at nucleotide position 1960. The amino acid change results in valine to leucine at codon 654, an amino acid with highly similar properties. However, this change occurs in the last base pair of coding exon 19, which makes it likely to have some effect on normal mRNA splicing. This mutation has been described in multiple individuals with retinoblastoma (Alonso J et al. Hum. Mutat. 2005 Jan;25(1):99; Li T et al. Int J Clin Exp Pathol 2016;9(2):2120-2126; Ambry internal data). Using two different splice site prediction tools, this alteration is predicted by ESEfinder to abolish the native splice donor site, but is predicted to weaken (but not abolish) the efficiency of the native splice donor site by BDGP. RNA studies using patient RNA have shown that both this alteration and another mutation at this nucleotide position (c.1960G>A) cause exon 19 skipping, resulting in a truncated protein (Houdayer C et al. Hum. Mutat. 2008 Jul;29(7):975-82). Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Uncertain	0.49
BayesDel_addAF	Pathogenic	0.31	D
BayesDel_noAF	Pathogenic	0.21
CADD	Uncertain	25
DANN	Uncertain	0.99
DEOGEN2	Uncertain	0.68	D;.
Eigen	Uncertain	0.19
Eigen_PC	Uncertain	0.38
FATHMM_MKL	Pathogenic	0.98	D
LIST_S2	Uncertain	0.89	D;D
M_CAP	Benign	0.052	D
MetaRNN	Uncertain	0.67	D;D
MetaSVM	Uncertain	0.18	D
MutationAssessor	Benign	0.55	N;.
PhyloP100		5.9
PrimateAI	Uncertain	0.76	T
PROVEAN	Benign	-1.3	N;.
REVEL	Uncertain	0.48
Sift	Benign	0.28	T;.
Sift4G	Benign	0.97	T;.
Polyphen		0.35	B;.
Vest4		0.44
MutPred		0.78		Gain of catalytic residue at Y659 (P = 0.0117);Gain of catalytic residue at Y659 (P = 0.0117);
MVP		0.97
MPC		1.3
ClinPred		0.86	D
GERP RS		6.0
Varity_R		0.58
gMVP		0.72
Mutation Taster		=6/94	disease causing (ClinVar)

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.99
SpliceAI score (max)		0.21		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.21		Position offset: 0

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

Other links and lift over

dbSNP: rs483352690; hg19: chr13-49030485; COSMIC: COSV57309017; COSMIC: COSV57309017;