rs61751276

Positions:

• chr1-68449890-C-T

Variant summary

Our verdict is Pathogenic. Variant got 8 ACMG points: 8P and 0B. PP4_Moderate PP3 PP1 PM3 PM2_Supporting PS3_Supporting

This summary comes from the ClinGen Evidence Repository: The NM_000329.3(RPE65):c.11+5G>A variant is a putative splicing variant in intron 1 of the RPE65 gene. The computational splicing predictor SpliceAI lists a change score of 0.58 for splice donor gain, predicting a deleterious impact on splicing (PP3). VCEP-member provided data from a mini-gene assay in HEK-293 cells show that this variant reduces normal splicing and leads to >400x reduction of mature mRNA, relative to the wild-type control (PS3_Supporting, Guan et al., 2024). At least 3 patients with this variant have been reported with detailed phenotypes (PMID:21211845, PMID:11786058), one of whom exhibited nonrecordable ERG responses from rods (0.5 pts) and cones (1 pt), congenital night blindness (0.5 pts), absence of autofluorescence (2 pts), optic nerve pallor (0.5 pts), RPE granularity (0.5 pts), onset between birth and age 5 years (1 pt), OCT preserved at macula (1 pt), constricted Goldmann visual field (1 pt), decreased central visual acuity (1 pt), and nystagmus (1 pt), which together are highly specific for recessive RPE65 retinopathy (10 pts total, VCEP member-provided data, PP4_Moderate). This variant has been detected in at least 7 published cases with recessive retinopathy, including at least two individuals homozygous for the variant (1 pt) and three individuals compound heterozygous for this variant in trans with either the c.615_616del (p.Ile206Cysfs*27), p.Glu102Ter, or c.89dup (p.Thr31fs) variant, all which have been classified Pathogenic by this VCEP (4pts, PMID:11095629, PMID:17525851, PMID:35129589), (PM3_Very-Strong). Two publications contain segregation data (PMID:11786058 and PMID:35001204), with one set of genotype-positive siblings exhibiting the required phenotype of absent or severely decreased rod electroretinogram response (PP1). The GrpMax Filtering AF for this variant in gnomAD v2.1.1 is 0.0001017 (PM2_Supporting). In summary, this variant meets the criteria to be classified as pathogenic for RPE65-related recessive retinopathy based on the ACMG/AMP criteria applied, as specified by the ClinGen LCA/eoRD VCEP: PM3_Very-Strong, PP4_Moderate, PP1, PM2_Supporting, PP3, PS3_Supporting. (VCEP specifications version 1.0.0; date of approval 09/21/2023). LINK:https://erepo.genome.network/evrepo/ui/classification/CA226483/MONDO:0100368/120

• Frequency:
  • Genomes: 𝑓 0.00013 (0 hom., cov: 32)
  • Exomes 𝑓: 0.00016 (0 hom.)
• Consequence: RPE65 NM_000329.3 splice_region, intron
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic reviewed by expert panel P:24 O:1
• Conservation: PhyloP100: 5.18

Genes affected

RPE65 (HGNC:10294): (retinoid isomerohydrolase RPE65) The protein encoded by this gene is a component of the vitamin A visual cycle of the retina which supplies the 11-cis retinal chromophore of the photoreceptors opsin visual pigments. It is a member of the carotenoid cleavage oxygenase superfamily. All members of this superfamily are non-heme iron oxygenases with a seven-bladed propeller fold and oxidatively cleave carotenoid carbon:carbon double bonds. However, the protein encoded by this gene has acquired a divergent function that involves the concerted O-alkyl ester cleavage of its all-trans retinyl ester substrate and all-trans to 11-cis double bond isomerization of the retinyl moiety. As such, it performs the essential enzymatic isomerization step in the synthesis of 11-cis retinal. Mutations in this gene are associated with early-onset severe blinding disorders such as Leber congenital. [provided by RefSeq, Oct 2017]

LOC124904198 (HGNC:):

ACMG classification

Verdict is Pathogenic. Variant got 8 ACMG points.

• PS3: For more information check the summary or visit ClinGen Evidence Repository.
• PM2: For more information check the summary or visit ClinGen Evidence Repository.
• PM3: For more information check the summary or visit ClinGen Evidence Repository.
• PP1: For more information check the summary or visit ClinGen Evidence Repository.
• PP3: For more information check the summary or visit ClinGen Evidence Repository.
• PP4: For more information check the summary or visit ClinGen Evidence Repository.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
RPE65	NM_000329.3	c.11+5G>A		splice_region_variant, intron_variant		ENST00000262340.6	NP_000320.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
RPE65	ENST00000262340.6	c.11+5G>A		splice_region_variant, intron_variant		1	NM_000329.3	ENSP00000262340.5

Frequencies

GnomAD3 genomes AF: 0.000131 AC: 20 AN: 152226 Hom.: 0 Cov.: 32

Gnomad AFR AF: 0.0000965

Gnomad AMI AF: 0.00

Gnomad AMR AF: 0.00

Gnomad ASJ AF: 0.00

Gnomad EAS AF: 0.00

Gnomad SAS AF: 0.00

Gnomad FIN AF: 0.00

Gnomad MID AF: 0.00

Gnomad NFE AF: 0.000235

Gnomad OTH AF: 0.00

GnomAD3 exomes AF: 0.0000756 AC: 19 AN: 251324 Hom.: 0 AF XY: 0.0000736 AC XY: 10 AN XY: 135870

Gnomad AFR exome AF: 0.0000617

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: 0.000158

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.000158 AC: 231 AN: 1461760 Hom.: 0 Cov.: 30 AF XY: 0.000153 AC XY: 111 AN XY: 727194

Gnomad4 AFR exome AF: 0.0000896

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: 0.000201

Gnomad4 OTH exome AF: 0.0000662

GnomAD4 genome AF: 0.000131 AC: 20 AN: 152226 Hom.: 0 Cov.: 32 AF XY: 0.000148 AC XY: 11 AN XY: 74372

Gnomad4 AFR AF: 0.0000965

Gnomad4 AMR AF: 0.00

Gnomad4 ASJ AF: 0.00

Gnomad4 EAS AF: 0.00

Gnomad4 SAS AF: 0.00

Gnomad4 FIN AF: 0.00

Gnomad4 NFE AF: 0.000235

Gnomad4 OTH AF: 0.00

Alfa AF: 0.000265 Hom.: 0

Bravo AF: 0.000147

EpiCase AF: 0.0000545

EpiControl AF: 0.000119

ClinVar

Significance: Pathogenic

Submissions summary: Pathogenic:24 Other:1

Revision: reviewed by expert panel

Submissions by phenotype

not provided Pathogenic:7 Other:1

Pathogenic, criteria provided, single submitter	clinical testing	Eurofins Ntd Llc (ga)	Oct 29, 2015	- -
not provided, no classification provided	literature only	Retina International	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	Institute of Medical Genetics and Applied Genomics, University Hospital Tübingen	Oct 23, 2020	- -
Pathogenic, criteria provided, single submitter	clinical testing	CeGaT Center for Human Genetics Tuebingen	Jun 01, 2024	RPE65: PM3:Very Strong, PM2, PS3:Moderate -
Pathogenic, criteria provided, single submitter	clinical testing	Center for Genomic Medicine, Rigshospitalet, Copenhagen University Hospital	Jul 31, 2024	- -
Pathogenic, criteria provided, single submitter	clinical testing	ARUP Laboratories, Molecular Genetics and Genomics, ARUP Laboratories	Oct 21, 2022	- -
Likely pathogenic, criteria provided, single submitter	clinical testing	Revvity Omics, Revvity	Jun 12, 2021	- -
Pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Jan 12, 2023	Published functional studies suggest that the variant causes a defect in RPE65 expression (Vazquez-Dominguez et al., 2022); This variant is associated with the following publications: (PMID: 11462243, 20683928, 23891399, 17964524, 32531858, 11786058, 19854499, 9326941, 20079931, 10766140, 25257057, 17525851, 21211845, 20811047, 18441370, 30268864, 31174678, 31736247, 32581362, 32865313, 12960219, 35121194, 34492281, 35129589, 34906171, 36429068, 29332120) -

Retinitis pigmentosa 20 Pathogenic:4

Likely pathogenic, criteria provided, single submitter	clinical testing	Centre for Mendelian Genomics, University Medical Centre Ljubljana	Jun 12, 2019	This variant was classified as: Likely pathogenic. The following ACMG criteria were applied in classifying this variant: PS1,PM2,PM3,PP3. -
Pathogenic, criteria provided, single submitter	research	Ocular Genomics Institute, Massachusetts Eye and Ear	Apr 08, 2021	The RPE65 c.11+5G>A variant was identified in an individual with retinitis pigmentosa with a presumed recessive inheritance pattern. Through a review of available evidence we were able to apply the following criteria: PVS1, PM2, PM3, PP1-S, PP3, PM1. Based on this evidence we have classified this variant as Pathogenic. -
Pathogenic, no assertion criteria provided	literature only	OMIM	Sep 01, 2003	- -
Likely pathogenic, no assertion criteria provided	clinical testing	Joint Genome Diagnostic Labs from Nijmegen and Maastricht, Radboudumc and MUMC+	Sep 01, 2016	- -

Leber congenital amaurosis 2 Pathogenic:3

Pathogenic, no assertion criteria provided	research	Laboratory of Genetics in Ophthalmology, Institut Imagine	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	Baylor Genetics	Mar 27, 2024	- -
Pathogenic, no assertion criteria provided	literature only	OMIM	Sep 01, 2003	- -

Retinal dystrophy Pathogenic:3

Likely pathogenic, criteria provided, single submitter	clinical testing	Institute of Human Genetics, Univ. Regensburg, Univ. Regensburg	Jan 01, 2021	- -
Likely pathogenic, no assertion criteria provided	research	NIHR Bioresource Rare Diseases, University of Cambridge	Jan 01, 2015	- -
Pathogenic, criteria provided, single submitter	clinical testing	Blueprint Genetics	Oct 19, 2018	- -

Leber congenital amaurosis Pathogenic:2

Pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Aug 23, 2022	Variant summary: RPE65 c.11+5G>A 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: Two predict the variant abolishes the canonical 5' splicing donor site. Two predict the variant weakens the canonical 5' splicing donor site. However, these predictions have yet to be confirmed by functional studies. The variant allele was found at a frequency of 7.6e-05 in 251324 control chromosomes. This frequency is not significantly higher than estimated for a pathogenic variant in RPE65 causing Leber Congenital Amaurosis (7.6e-05 vs 0.0014), allowing no conclusion about variant significance. c.11+5G>A has been reported in the literature in multiple individuals affected with Leber Congenital Amaurosis/Visual impairment/Retinopathies (example, Galvin_2005, Astuti_2016, Haer-Wigman_2017, Testa_2022). These data indicate that the variant is very likely to be associated with disease. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. Twelve 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 pathogenic/likely pathogenic. Based on the evidence outlined above, the variant was classified as pathogenic. -
Pathogenic, no assertion criteria provided	clinical testing	Natera, Inc.	Sep 16, 2020	- -

RPE65-related disorder Pathogenic:1

Pathogenic, no assertion criteria provided

clinical testing

PreventionGenetics, part of Exact Sciences

Aug 14, 2024

The RPE65 c.11+5G>A variant is predicted to interfere with splicing. This variant has been reported as causative for severe congenital autosomal recessive retinal disorders (see for examples: Astuti et al. 2016. PubMed ID: 26626312; Coppieters et al. 2010. PubMed ID: 20683928; Ripamonti et al. 2014. PubMed ID: 25257057). This variant is one of the three most frequent causative variants in RPE65 (Astuti et al. 2016. PubMed ID: 26626312). This variant is reported in 0.016% of alleles in individuals of European (Non-Finnish) descent in gnomAD. This variant has been classified as pathogenic by a ClinGen Variant Curation Expert Panel (https://www.ncbi.nlm.nih.gov/clinvar/variation/98825/). Given all the evidence, we interpret this variant as pathogenic. -

RPE65-related recessive retinopathy Pathogenic:1

Pathogenic, reviewed by expert panel

curation

ClinGen Leber Congenital Amaurosis/early Onset Retinal Dystrophy Variant Curation Expert Panel, ClinGen

Dec 22, 2023

The NM_000329.3(RPE65):c.11+5G>A variant is a putative splicing variant in intron 1 of the RPE65 gene. The computational splicing predictor SpliceAI lists a change score of 0.58 for splice donor gain, predicting a deleterious impact on splicing (PP3). VCEP-member provided data from a mini-gene assay in HEK-293 cells show that this variant reduces normal splicing and leads to >400x reduction of mature mRNA, relative to the wild-type control (PS3_Supporting, Guan et al., 2024). At least 3 patients with this variant have been reported with detailed phenotypes (PMID: 21211845, PMID: 11786058), one of whom exhibited nonrecordable ERG responses from rods (0.5 pts) and cones (1 pt), congenital night blindness (0.5 pts), absence of autofluorescence (2 pts), optic nerve pallor (0.5 pts), RPE granularity (0.5 pts), onset between birth and age 5 years (1 pt), OCT preserved at macula (1 pt), constricted Goldmann visual field (1 pt), decreased central visual acuity (1 pt), and nystagmus (1 pt), which together are highly specific for recessive RPE65 retinopathy (10 pts total, VCEP member-provided data, PP4_Moderate). This variant has been detected in at least 7 published cases with recessive retinopathy, including at least two individuals homozygous for the variant (1 pt) and three individuals compound heterozygous for this variant in trans with either the c.615_616del (p.Ile206Cysfs*27), p.Glu102Ter, or c.89dup (p.Thr31fs) variant, all which have been classified Pathogenic by this VCEP (4pts, PMID: 11095629, PMID: 17525851, PMID: 35129589), (PM3_Very-Strong). Two publications contain segregation data (PMID: 11786058 and PMID: 35001204), with one set of genotype-positive siblings exhibiting the required phenotype of absent or severely decreased rod electroretinogram response (PP1). The GrpMax Filtering AF for this variant in gnomAD v2.1.1 is 0.0001017 (PM2_Supporting). In summary, this variant meets the criteria to be classified as pathogenic for RPE65-related recessive retinopathy based on the ACMG/AMP criteria applied, as specified by the ClinGen LCA/eoRD VCEP: PM3_Very-Strong, PP4_Moderate, PP1, PM2_Supporting, PP3, PS3_Supporting. (VCEP specifications version 1.0.0; date of approval 09/21/2023). -

Inborn genetic diseases Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Ambry Genetics

Nov 20, 2024

The c.11+5G>A intronic alteration consists of a G to A substitution 5 nucleotides after coding exon 1 of the RPE65 gene. Based on data from gnomAD, the A allele has an overall frequency of 0.008% (22/282726) total alleles studied. The highest observed frequency was 0.016% (21/129112) of European (non-Finnish) alleles. This variant has been identified in the homozygous state and in conjunction with other RPE65 variant(s) in individual(s) with features consistent with RPE65-related retinopathy (Gu, 1997; Yzer, 2003; Astuti, 2016; Deng, 2022). This nucleotide position is highly conserved in available vertebrate species. RNA studies have demonstrated that this alteration results in abnormal splicing (Vazquez-Dominguez, 2022). In silico splice site analysis predicts that this alteration will result in the creation or strengthening of a novel splice donor site. Based on the available evidence, this alteration is classified as pathogenic. -

Leber congenital amaurosis 2;C3151086:Retinitis pigmentosa 20 Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Labcorp Genetics (formerly Invitae), Labcorp

Jan 04, 2024

This sequence change falls in intron 1 of the RPE65 gene. It does not directly change the encoded amino acid sequence of the RPE65 protein. It affects a nucleotide within the consensus splice site. This variant is present in population databases (rs61751276, gnomAD 0.02%). This variant has been observed in individual(s) with retinitis pigmentosa or Leber congenital amaurosis (PMID: 9326941, 11462243, 11786058, 19854499, 20683928, 21211845, 25257057). In at least one individual the data is consistent with being in trans (on the opposite chromosome) from a pathogenic variant. This variant is also known as 65+5G>A and IVS1+5G>A. ClinVar contains an entry for this variant (Variation ID: 98825). Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Algorithms developed to predict the effect of sequence changes on RNA splicing suggest that this variant may disrupt the consensus splice site. For these reasons, this variant has been classified as Pathogenic. -

Leber congenital amaurosis 2;C3151086:Retinitis pigmentosa 20;C5231465:Retinitis pigmentosa 87 with choroidal involvement Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Fulgent Genetics, Fulgent Genetics

Mar 10, 2022

- -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_noAF	Benign	-0.39
CADD	Benign	22
DANN	Benign	0.91

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.99
SpliceAI score (max)		0.050		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: rs61751276; hg19: chr1-68915573;