rs397517735

Positions:

• chr2-178559309-A-T

Variant summary

Our verdict is Pathogenic. Variant got 16 ACMG points: 16P and 0B. PVS1_Strong PP3_Strong PP5_Very_Strong

The NM_001267550.2(TTN):​c.86821+2T>A variant causes a splice donor change involving the alteration of a conserved nucleotide. The variant allele was found at a frequency of 0.000012 in 1,581,920 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: 𝑓 0.000046 (0 hom., cov: 33)
  • Exomes 𝑓: 0.0000084 (0 hom.)
• Consequence: TTN NM_001267550.2 splice_donor
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:12
• Conservation: PhyloP100: 9.16

Genes affected

TTN (HGNC:12403): (titin) This gene encodes a large abundant protein of striated muscle. The product of this gene is divided into two regions, a N-terminal I-band and a C-terminal A-band. The I-band, which is the elastic part of the molecule, contains two regions of tandem immunoglobulin domains on either side of a PEVK region that is rich in proline, glutamate, valine and lysine. The A-band, which is thought to act as a protein-ruler, contains a mixture of immunoglobulin and fibronectin repeats, and possesses kinase activity. An N-terminal Z-disc region and a C-terminal M-line region bind to the Z-line and M-line of the sarcomere, respectively, so that a single titin molecule spans half the length of a sarcomere. Titin also contains binding sites for muscle associated proteins so it serves as an adhesion template for the assembly of contractile machinery in muscle cells. It has also been identified as a structural protein for chromosomes. Alternative splicing of this gene results in multiple transcript variants. Considerable variability exists in the I-band, the M-line and the Z-disc regions of titin. Variability in the I-band region contributes to the differences in elasticity of different titin isoforms and, therefore, to the differences in elasticity of different muscle types. Mutations in this gene are associated with familial hypertrophic cardiomyopathy 9, and autoantibodies to titin are produced in patients with the autoimmune disease scleroderma. [provided by RefSeq, Feb 2012]

TTN-AS1 (HGNC:44124): (TTN antisense RNA 1) This gene encodes a non-coding RNA transcribed from the opposite strand to the titin gene. [provided by RefSeq, Aug 2016]

ACMG classification

Verdict is Pathogenic. Variant got 16 ACMG points.

• PVS1: Splicing +-2 bp (donor or acceptor) variant, product NOT destroyed by NMD, known LOF gene, truncates exone, which is 0.15841483 fraction of the gene. No cryptic splice site detected. Exon removal is inframe change.
• PP3: Splicing scoreres supports a deletorius effect: Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF, max_spliceai. No scorers claiming Uncertain. No scorers claiming Benign.
• PP5: Variant 2-178559309-A-T is Pathogenic according to our data. Variant chr2-178559309-A-T is described in ClinVar as [Likely_pathogenic]. Clinvar id is 47458.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars. Variant chr2-178559309-A-T is described in Lovd as [Pathogenic]. Variant chr2-178559309-A-T is described in Lovd as [Likely_pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
TTN	NM_001267550.2	c.86821+2T>A		splice_donor_variant		ENST00000589042.5
TTN-AS1	NR_038272.1	n.2043+16948A>T		intron_variant, non_coding_transcript_variant

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
TTN	ENST00000589042.5	c.86821+2T>A		splice_donor_variant		5	NM_001267550.2	P1
TTN-AS1	ENST00000659121.1	n.416+35673A>T		intron_variant, non_coding_transcript_variant

Frequencies

GnomAD3 genomes AF: 0.0000460 AC: 7 AN: 152174 Hom.: 0 Cov.: 33

Gnomad AFR AF: 0.0000483

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: 0.0000735

Gnomad OTH AF: 0.00

GnomAD3 exomes AF: 0.00000437 AC: 1 AN: 228782 Hom.: 0 AF XY: 0.00 AC XY: 0 AN XY: 123794

Gnomad AFR exome AF: 0.00

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: 0.00000944

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.00000839 AC: 12 AN: 1429746 Hom.: 0 Cov.: 30 AF XY: 0.0000113 AC XY: 8 AN XY: 707908

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: 0.0000100

Gnomad4 OTH exome AF: 0.0000170

GnomAD4 genome AF: 0.0000460 AC: 7 AN: 152174 Hom.: 0 Cov.: 33 AF XY: 0.0000538 AC XY: 4 AN XY: 74332

Gnomad4 AFR AF: 0.0000483

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: 0.0000735

Gnomad4 OTH AF: 0.00

Alfa AF: 0.0000314 Hom.: 0

Bravo AF: 0.0000302

ExAC AF: 0.00000828 AC: 1

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:12

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

not provided Pathogenic:4

Pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Aug 15, 2023	Not observed at significant frequency in large population cohorts (gnomAD); Canonical splice site variant predicted to result in an in-frame loss of the adjacent exon in a gene for which loss of function is a known mechanism of disease; Located in the A-band region of TTN in which the majority of loss of function variants have been associated with autosomal dominant titinopathies (Herman et al., 2012); This variant is associated with the following publications: (PMID: 22335739, 23418287, 30681174, 31402444, 33874732, 36264615, 34731015, 33226272) -
Pathogenic, criteria provided, single submitter	clinical testing	Revvity Omics, Revvity	Feb 08, 2022	- -
Pathogenic, criteria provided, single submitter	clinical testing	Mayo Clinic Laboratories, Mayo Clinic	Feb 25, 2020	PVS1, PM1, PP1 -
Pathogenic, criteria provided, single submitter	clinical testing	Eurofins Ntd Llc (ga)	May 30, 2017	- -

Primary dilated cardiomyopathy Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Jan 18, 2019	The c.79117+2T>A variant in TTN has been identified in 4 individuals with DCM and segregated with the disease in 5 affected relatives from 3 families (Norton 2013, Herman 2012, LMM data). It has also been identified 1/21992 African chromosomes by gnomAD (http://gnomad.broadinstitute.org); however, the quality of this region was low. This variant occurs within the canonical splice site (+/- 1,2) and is predicted to cause altered splicing leading to an abnormal or absent protein. Splice site variants and other truncating variants in TTN are strongly associated with DCM if they impact the exons encoding for the A-band (Herman 2012, Pugh 2014) and/or are located in an exon that is highly expressed in the heart (Roberts 2015). The c.79117+2T>A variant is located in A-band. Loss of function of the TTN gene is an established disease mechanism in autosomal dominant DCM. ACMG/AMP criteria applied: PVS1_Moderate, PS4_Moderate, PP1_Moderate. -
Pathogenic, criteria provided, single submitter	clinical testing	Center for Advanced Laboratory Medicine, UC San Diego Health, University of California San Diego	Apr 06, 2018	- -

TTN-related disorder Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

PreventionGenetics, part of Exact Sciences

Dec 11, 2023

The TTN c.86821+2T>A variant is predicted to disrupt the GT donor site and interfere with normal splicing. This variant was reported in the heterozygous and compound heterozygous states in individuals with TTN-related disorders (described as c.81898+2T>A, Herman et al. 2012. PubMed ID: 22335739; described as IVS 275+2T>A, Norton et al. 2013. PubMed ID: 23418287; Martinez-Thompson et al. 2019. PubMed ID: 30681174; Table S2, Goli et al. 2021. PubMed ID: 33874732). This variant is reported in 0.0042% of alleles in individuals of African descent in gnomAD. This variant is located in the canonical splice site within A-band region of the protein in which truncating TTN variants have been found more frequently in dilated cardiomyopathy patients than in controls (Herman et al. 2012. PubMed ID: 22335739). RNAseq studies from heart tissue indicate this exon is commonly included in TTN mRNA transcripts (PSI of 100%); however, this analysis in muscle tissue was not performed (Roberts et al. 2015. PMID: 25589632; https://cardiodb.org/titin/titin_transcripts.php). TTN truncating variants are reported in 1-2% of presumably healthy individuals and occur more frequently in exons with low PSI values, indicating this variant is more likely to be disease causing (Herman et al. 2012. PMID: 22335739; Roberts et al. 2015. PMID: 25589632). In summary, this variant is interpreted as pathogenic for TTN-related disorders. -

Autosomal recessive limb-girdle muscular dystrophy type 2J;C1858763:Dilated cardiomyopathy 1G Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Invitae

Jan 31, 2024

This sequence change affects a donor splice site in intron 326 of the TTN gene. It is expected to disrupt RNA splicing and likely results in a truncated or disrupted TTN protein. This variant is present in population databases (rs397517735, gnomAD 0.01%). Disruption of this splice site has been observed in individual(s) with dilated cardiomyopathy (PMID: 22335739, 23418287, 25589632). It has also been observed to segregate with disease in related individuals. This variant is also known as IVS 275+2T>A. ClinVar contains an entry for this variant (Variation ID: 47458). Algorithms developed to predict the effect of sequence changes on RNA splicing suggest that this variant may disrupt the consensus splice site. This variant is located in the A band of TTN (PMID: 25589632). Truncating variants in this region are significantly overrepresented in patients affected with dilated cardiomyopathy (PMID: 25589632). Truncating variants in this region have also been reported in individuals affected with autosomal recessive centronuclear myopathy (PMID: 23975875). For these reasons, this variant has been classified as Pathogenic. -

Cardiomyopathy Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

CHEO Genetics Diagnostic Laboratory, Children's Hospital of Eastern Ontario

Jun 12, 2017

- -

Dilated cardiomyopathy 1G Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Illumina Laboratory Services, Illumina

Aug 14, 2023

The TTN c.86821+2T>A variant, also referred to as c.79117+2T>A, c.81898+2T>A, or IVS 275+2T>A, results in a substitution at the consensus splice donor site which is predicted to result in splicing defects. This variant affects exon 326 of the meta transcript of titin within the A-band, which is highly expressed in cardiac tissue (PMID: 25589632). In a meta-analysis of TTN truncating variants in patients and controls, variants in this region were associated with a significantly increased risk of developing dilated cardiomyopathy (DCM; odds ratio 49.8) (PMID: 27869827). This variant has been identified in individuals diagnosed with DCM, and has been shown to segregate with disease in multiple families (PMID: 22335739; 23418287; 24503780). This variant is not observed at a significant frequency in version 2.1.1 or version 3.1.2 of the Genome Aggregation Database. This variant has been classified as pathogenic by at least three submitters in ClinVar. Based on the available evidence, the c.86821+2T>A variant is classified as pathogenic for dilated cardiomyopathy. -

Autosomal recessive limb-girdle muscular dystrophy type 2J;C1838244:Tibial muscular dystrophy;C1858763:Dilated cardiomyopathy 1G;C1861065:Hypertrophic cardiomyopathy 9;C1863599:Myopathy, myofibrillar, 9, with early respiratory failure;C2673677:Early-onset myopathy with fatal cardiomyopathy Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Fulgent Genetics, Fulgent Genetics

Oct 02, 2021

- -

Cardiovascular phenotype Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Ambry Genetics

May 28, 2024

The c.59626+2T>A intronic variant results from a T to A substitution two nucleotides after coding exon 153 in the TTN gene. This alteration disrupts the canonical splice site and is expected to cause aberrant splicing, resulting in an abnormal protein or a transcript that is subject to nonsense-mediated mRNA decay. This exon is located in the A-band region of the N2-B isoform of the titin protein and is constitutively expressed in TTN transcripts (percent spliced in or PSI 100%). Based on data from gnomAD, the A allele has an overall frequency of 0.001% (2/260184) total alleles studied. The highest observed frequency was 0.004% (1/23844) of African alleles. This variant has been reported in several dilated cardiomyopathy patients and has been shown to segregate with disease in families (Herman, 2012; Norton, 2013; Pugh, 2014). 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. Based on the available evidence, this alteration is classified as pathogenic. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.23	D
BayesDel_noAF	Uncertain	0.090
CADD	Pathogenic	30
DANN	Uncertain	0.99
Eigen	Pathogenic	1.2
Eigen_PC	Pathogenic	1.0
FATHMM_MKL	Pathogenic	1.0	D
MutationTaster	Benign	1.0	D;D;D;D;D;D
GERP RS		5.6

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.93
SpliceAI score (max)		0.99		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.99		Position offset: 2

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

Publications

LitVar

Other links and lift over

dbSNP: rs397517735; hg19: chr2-179424036;