11-5225727-C-T

Variant names:

• chr11-5225727-C-T
• rs33952266
• NM_000518.5:c.316-1G>A

Variant summary

Our verdict is Pathogenic. Variant got 18 ACMG points: 18P and 0B. PVS1 PM2 PP5_Very_Strong

The NM_000518.5(HBB):​c.316-1G>A variant causes a splice acceptor, intron change. The variant allele was found at a frequency of 0.00000137 in 1,461,790 control chromosomes in the GnomAD database, with no homozygous occurrence. 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 Likely pathogenic (★★).

• Frequency:
  • Genomes: not found (cov: 33)
  • Exomes 𝑓: 0.0000014 (0 hom.)
• Consequence: HBB NM_000518.5 splice_acceptor, intron
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:7
• Conservation: PhyloP100: 7.10

Genes affected

HBB (HGNC:4827): (hemoglobin subunit beta) The alpha (HBA) and beta (HBB) loci determine the structure of the 2 types of polypeptide chains in adult hemoglobin, Hb A. The normal adult hemoglobin tetramer consists of two alpha chains and two beta chains. Mutant beta globin causes sickle cell anemia. Absence of beta chain causes beta-zero-thalassemia. Reduced amounts of detectable beta globin causes beta-plus-thalassemia. The order of the genes in the beta-globin cluster is 5'-epsilon -- gamma-G -- gamma-A -- delta -- beta--3'. [provided by RefSeq, Jul 2008]

ACMG classification

Verdict is Pathogenic. Variant got 18 ACMG points.

• PVS1: Splicing +-2 bp (donor or acceptor) variant, LoF is a know mechanism of disease,
• PM2: Very rare variant in population databases, with high coverage
• PP5: Variant 11-5225727-C-T is Pathogenic according to our data. Variant chr11-5225727-C-T is described in ClinVar as [Likely_pathogenic]. Clinvar id is 552712.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars. Variant chr11-5225727-C-T is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
HBB	NM_000518.5	c.316-1G>A		splice_acceptor_variant, intron_variant	Intron 2 of 2	ENST00000335295.4	NP_000509.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
HBB	ENST00000335295.4	c.316-1G>A		splice_acceptor_variant, intron_variant	Intron 2 of 2	1	NM_000518.5	ENSP00000333994.3
HBB	ENST00000647020.1	c.316-1G>A		splice_acceptor_variant, intron_variant	Intron 2 of 2			ENSP00000494175.1
HBB	ENST00000475226.1	n.248-1G>A		splice_acceptor_variant, intron_variant	Intron 1 of 1	2
HBB	ENST00000633227.1	n.*132-1G>A		splice_acceptor_variant, intron_variant	Intron 2 of 2	3		ENSP00000488004.1

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome AF: 0.00000137 AC: 2 AN: 1461790 Hom.: 0 Cov.: 31 AF XY: 0.00000275 AC XY: 2 AN XY: 727214

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.: 33

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:7

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

beta Thalassemia Pathogenic:4

Nov 25, 2019

The ITHANET community portal, The Cyprus Institute of Neurology and Genetics

Significance: Pathogenic

Review Status: no assertion criteria provided

Collection Method: curation

- -

Jul 01, 2018

College of Science, Al Muthanna University, Al Muthanna University

Significance: Pathogenic

Review Status: no assertion criteria provided

Collection Method: research

- -

Apr 05, 2024

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

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

Variant summary: HBB c.316-1G>A (also described in the literature as IVS-II-850G>A) is located in a canonical splice-site and is predicted to affect mRNA splicing resulting in a significantly altered protein due to either exon skipping, shortening, or inclusion of intronic material. Several computational tools predict a significant impact on normal splicing: Four predict the variant abolishes a 3 acceptor site. However, these predictions have yet to be confirmed by functional studies. The variant was absent in 251150 control chromosomes (gnomAD). c.316-1G>A has been reported in the literature in homozygous and heterozygous individuals affected with beta-thalassemia (e.g. Al Mosawi_2020). Furthermore, beta-thal trait individuals heterozygous for the variant, were reported with a history of mild anemia and microcytosis (e.g. Curuk_1995, Knott_2006). These data indicate that the variant is likely to be associated with disease. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. Other variants affecting the same splice-site nucleotide (c.316-1G>C, c.316-1G>T) have been reported internally and also, in ClinVar and HGMD as pathogenic/likely pathogenic and disease-associated. The following publications have been ascertained in the context of this evaluation (PMID: 31714438, 7558878, 16466947, 9101288). ClinVar contains an entry for this variant (Variation ID: 552712). Based on the evidence outlined above, the variant was classified as likely pathogenic. -

Aug 22, 2017

Counsyl

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

- -

Beta zero thalassemia Pathogenic:2

May 01, 1995

OMIM

Significance: Pathogenic

Review Status: no assertion criteria provided

Collection Method: literature only

- -

Jan 01, 2018

College of Science, Al Muthanna University, Al Muthanna University

Significance: Pathogenic

Review Status: no assertion criteria provided

Collection Method: research

- -

not provided Pathogenic:1

Feb 25, 2023

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

For these reasons, this variant has been classified as Pathogenic. 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. Studies have shown that disruption of this splice site alters HBB gene expression (PMID: 6583702). ClinVar contains an entry for this variant (Variation ID: 552712). Disruption of this splice site has been observed in individuals with autosomal recessive beta-thalassemia (PMID: 2123063, 6583702). It has also been observed to segregate with disease in related individuals. This variant is not present in population databases (gnomAD no frequency). This sequence change affects an acceptor splice site in intron 2 of the HBB gene. While this variant is not anticipated to result in nonsense mediated decay, it likely alters RNA splicing and results in a disrupted protein product. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.60	D
BayesDel_noAF	Pathogenic	0.34
CADD	Pathogenic	29
DANN	Uncertain	0.99
Eigen	Pathogenic	1.1
Eigen_PC	Pathogenic	0.93
FATHMM_MKL	Pathogenic	0.98	D
GERP RS		4.7

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.93
SpliceAI score (max)		0.98		Details are displayed if max score is > 0.2
DS_AL_spliceai		0.98		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: rs33952266; hg19: chr11-5246957;