rs45466197

chr1-201361327-C-A

Variant summary

Our verdict is Likely benign. Variant got -2 ACMG points: 2P and 4B. PM1 BS2

The NM_001276345.2(TNNT2):c.762G>T(p.Glu254Asp) variant causes a missense change involving the alteration of a non-conserved nucleotide. The variant allele was found at a frequency of 0.000191 in 1,614,050 control chromosomes in the GnomAD database, with no homozygous occurrence. Variant has been reported in ClinVar as Conflicting classifications of pathogenicity (no stars).

• Frequency:
  • Genomes: 𝑓 0.000066 (0 hom., cov: 33)
  • Exomes 𝑓: 0.00020 (0 hom.)
• Consequence: TNNT2 NM_001276345.2 missense
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications U:14 B:2
• Conservation: PhyloP100: 1.71

Genes affected

TNNT2 (HGNC:11949): (troponin T2, cardiac type) This gene encodes the cardiac isoform of troponin T. The encoded protein is the tropomyosin-binding subunit of the troponin complex, which is located on the thin filament of striated muscles and regulates muscle contraction in response to alterations in intracellular calcium ion concentration. Mutations in this gene have been associated with familial hypertrophic cardiomyopathy as well as with dilated cardiomyopathy. [provided by RefSeq, May 2022]

ACMG classification

Verdict is Likely_benign. Variant got -2 ACMG points.

• PM1: In a chain Troponin T, cardiac muscle (size 296) in uniprot entity TNNT2_HUMAN there are 174 pathogenic changes around while only 18 benign (91%) in NM_001276345.2
• BS2: High AC in GnomAd at 10 AD gene.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
TNNT2	NM_001276345.2	c.762G>T	p.Glu254Asp	missense_variant	15/17	ENST00000656932.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
TNNT2	ENST00000656932.1	c.762G>T	p.Glu254Asp	missense_variant	15/17		NM_001276345.2	A2

Frequencies

GnomAD3 genomes AF: 0.0000657 AC: 10 AN: 152182 Hom.: 0 Cov.: 33

Gnomad AFR AF: 0.00

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: 0.000147

Gnomad OTH AF: 0.00

GnomAD3 exomes AF: 0.000223 AC: 56 AN: 251480 Hom.: 0 AF XY: 0.000243 AC XY: 33 AN XY: 135914

Gnomad AFR exome AF: 0.0000615

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: 0.000483

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.000205 AC: 299 AN: 1461868 Hom.: 0 Cov.: 31 AF XY: 0.000191 AC XY: 139 AN XY: 727240

Gnomad4 AFR exome AF: 0.0000597

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: 0.0000374

Gnomad4 NFE exome AF: 0.000258

Gnomad4 OTH exome AF: 0.000132

GnomAD4 genome AF: 0.0000657 AC: 10 AN: 152182 Hom.: 0 Cov.: 33 AF XY: 0.0000942 AC XY: 7 AN XY: 74348

Gnomad4 AFR AF: 0.00

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: 0.000147

Gnomad4 OTH AF: 0.00

Alfa AF: 0.000180 Hom.: 0

Bravo AF: 0.0000907

TwinsUK AF: 0.000809 AC: 3

ALSPAC AF: 0.000778 AC: 3

ESP6500AA AF: 0.000227 AC: 1

ESP6500EA AF: 0.00 AC: 0

ExAC AF: 0.000329 AC: 40

EpiCase AF: 0.000109

EpiControl AF: 0.000237

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Uncertain:14 Benign:2

Revision: criteria provided, conflicting classifications

Submissions by phenotype

not provided Uncertain:4

Uncertain significance, criteria provided, single submitter	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Apr 04, 2019	The p.Glu244Asp variant (NM_001001430.1 c.732G>T; also referred to as NM_000364.3 c.753G>T p.Glu251Asp) in TNNT2 has been reported in one individual of unspecified ancestry with HCM (Watkins 1995) and one individual of African American ancestry with DCM who carried another variant of unknown significance in the TPM1 gene (Hershberger 2008, Hershberger 2009, Hershberger 2010, Rampersaud 2011), and has been reported in ClinVar (Variation ID#43667). This variant has been identified in 3 individuals with cardiomyopathy tested by our laboratory, one of whom carries a second likely disease-causing variant and had an early onset of disease, and a second who had an additional TNNT2 variant of uncertain significance and an early onset of disease. This variant has been identified in 0.06% (40/66740) of European chromosomes by the Exome Aggregation Consortium (ExAC, http://exac.broadinstitute.org; dbSNP rs45466197). Although this variant has been seen in the general population, its frequency is not high enough to rule out a pathogenic role. Computational analyses (biochemical amino acid properties, conservation, AlignGVGD, PolyPhen2, and SIFT) do not provide strong support for or against an impact to the protein. In vitro studies have shown that the p.Glu244Asp variant may impact protein function (Watkins 1995, Yanaga 1995, Harada 2004, Matsumoto 2009), but it has not been demonstrated whether this can result in disease. The presence of a variant in HCM and DCM probands raises suspicion about its clinical significance as the two cardiomyopathies are caused by different defects at the cellular level. Given the early onset of disease in all individuals carrying this variant in addition to a second variant, it is possible that it is exacerbating disease severity in these cases. In summary, additional data is need to interpret the pathogenicity of this variant for causing primary disease as well as whether it may play a role in modifying the severity of disease due to other causes. -
Uncertain significance, criteria provided, single submitter	clinical testing	GeneDx	Jun 29, 2022	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: 7898523, 19412328, 28352236, 23299917, 19293840, 20215591, 19473338, 29121657, 14722098, 20031601, 25637381, 19275886, 10085122, 26774798, 27493864, 21483645, 30847666, 30565988, 33025817) -
Uncertain significance, no assertion criteria provided	clinical testing	Diagnostic Laboratory, Department of Genetics, University Medical Center Groningen	-	- -
Uncertain significance, no assertion criteria provided	clinical testing	Clinical Genetics, Academic Medical Center	-	- -

Cardiomyopathy Uncertain:3

Uncertain significance, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Apr 27, 2023	This missense variant replaces glutamic acid with aspartic acid at codon 244 of the TNNT2 protein. Computational prediction suggests that this variant may have a deleterious impact on protein structure and function (internally defined REVEL score threshold >= 0.7, PMID: 27666373). Experimental functional studies have shown that the variant does not cause changes in Ca2+ sensitivity of force development but increases maximal force development (PMID: 10085122, 10467159, 14722098, 19293840). Clinical relevance of this observation is not clear. This variant has been reported in individuals affected with hypertrophic cardiomyopathy (PMID: 7898523, 30565988), in individuals affected with dilated cardiomyopathy (PMID: 19412328, 20031601, 21483645), and in an individual affected with arrhythmogenic right ventricular cardiomyopathy (PMID: 30847666). This variant has been identified in 58/282838 chromosomes in the general population by the Genome Aggregation Database (gnomAD). Although this variant allele frequency is thought to be higher than expected for TNNT2-related disorders, additional studies are necessary to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance. -
Uncertain significance, criteria provided, single submitter	clinical testing	CHEO Genetics Diagnostic Laboratory, Children's Hospital of Eastern Ontario	May 26, 2023	- -
Uncertain significance, criteria provided, single submitter	clinical testing	All of Us Research Program, National Institutes of Health	Feb 05, 2024	This missense variant replaces glutamic acid with aspartic acid at codon 244 of the TNNT2 protein. Computational prediction suggests that this variant may have a deleterious impact on protein structure and function (internally defined REVEL score threshold >= 0.7, PMID: 27666373). Experimental functional studies have shown that the variant does not cause changes in Ca2+ sensitivity of force development but increases maximal force development (PMID: 10085122, 10467159, 14722098, 19293840). Clinical relevance of this observation is not clear. This variant has been reported in individuals affected with hypertrophic cardiomyopathy (PMID: 7898523, 30565988), in individuals affected with dilated cardiomyopathy (PMID: 19412328, 20031601, 21483645), and in an individual affected with arrhythmogenic right ventricular cardiomyopathy (PMID: 30847666). This variant has been identified in 58/282838 chromosomes in the general population by the Genome Aggregation Database (gnomAD). Although this variant allele frequency is thought to be higher than expected for TNNT2-related disorders, additional studies are necessary to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance. -

Dilated cardiomyopathy 1D Uncertain:2

Uncertain significance, no assertion criteria provided	clinical testing	Knight Diagnostic Laboratories, Oregon Health and Sciences University	Dec 19, 2015	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Victorian Clinical Genetics Services, Murdoch Childrens Research Institute	Feb 02, 2022	Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as VUS-3B. Following criteria are met: 0105 - The mechanism of disease for this gene is not clearly established. Functional studies have suggested loss-of-function, gain-of-function and dominant-negative mechanisms based on calcium sensitivity, contractibility and mouse models (PMID: 18612386, 32098556, 33025817). (I) 0107 - This gene is associated with autosomal dominant disease. (I) 0200 - Variant is predicted to result in a missense amino acid change from glutamic acid to aspartic acid. (I) 0251 - This variant is heterozygous. (I) 0302 - Variant is present in gnomAD (v2) <0.001 for a dominant condition (58 heterozygotes, 0 homozygotes). (SP) 0502 - Missense variant with conflicting in silico predictions and uninformative conservation. (I) 0600 - Variant is located in the annotated troponin domain (DECIPHER). (I) 0705 - No comparable missense variants have previous evidence for pathogenicity. (I) 0809 - Previous evidence of pathogenicity for this variant is inconclusive. This variant has been reported once as likely benign, and many times as a VUS, in several individuals with hypertrophic cardiomyopathy, dilated cardiomyopathy and arrhythmogenic right ventricular cardiomyopathy (ClinVar, LOVD, PMID: 30847666). In some of these individuals, additional variants in cardiac genes were found. (I) 1002 - This variant has moderate functional evidence supporting abnormal protein function. Transfected bovine and porcine fibres have been shown to result in a slight increase in maximum ATPase activity, and significant increase in maximum force. The biological significance of these findings is unclear (PMID: 19275886, PMID: 14722098). (SP) 1208 - Inheritance information for this variant is not currently available in this individual. (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign -

not specified Uncertain:1 Benign:1

Likely benign, criteria provided, single submitter	clinical testing	Blueprint Genetics	Apr 23, 2015	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Sep 11, 2018	Variant summary: TNNT2 c.732G>T (p.Glu244Asp) results in a conservative amino acid change in the encoded protein sequence. Four of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 0.00021 in 277176 control chromosomes, predominantly within the Non-Finnish European subpopulation at a frequency of 0.00047 in the gnomAD database. The observed variant frequency within Non-Finnish European control individuals in the gnomAD database is approximately 2.7 fold of the estimated maximal expected allele frequency for a pathogenic variant in TNNT2 causing Cardiomyopathy phenotype (0.00018), suggesting that the variant is a benign polymorphism found primarily in populations of Non-Finnish European origin. c.732G>T has been reported in the literature in one individual affected with Hypertrophic Cardiomyopathy (Watkins 1995) and in another individual with Dilated Cardiomyopathy, who also had another variant of unknown significance in the TPM1 gene (Hershberger 2008, Hershberger 2009, Rampersaud 2011). In none of these cases were the variant shown to be present in family members. Therefore these reports do not provide unequivocal conclusions about association of the variant with Cardiomyopathy. Several publications reported experimental evidence evaluating an impact on protein function. Multiple studies found that the variant protein increased the maximum level of ATPase activity with either increasing (Nakaura 1999) or not affecting Ca2+ sensitivity (Yanaga 1999, Harada 2004, Matsumoto 2009). A further study demonstrated that the variant caused abnormal troponin function, i.e. impaired troponin binding to the thin filament (Tobacman 1999) and disturbed the protein conformation (Matsuo 2015). However, it is unclear how these changes can result in disease. Four other clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. Multiple laboratories reported the variant with conflicting assessments (VUS (2x), Likely benign (1x), Likely pathogenic (1x)). Based on the evidence outlined above, the variant was classified as uncertain significance, until additional evidence becomes available. -

TNNT2-Related Cardiomyopathy Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Illumina Laboratory Services, Illumina

Apr 20, 2018

The TNNT2 c.732G>T (p.Glu244Asp) missense variant has been reported in at least four studies in which it is found in a heterozygous state in three probands (Watkins et al. 1995; Hershberger et al. 2009; Rampersaud et al. 2011; Viswanathan et al. 2017). Two of the probands were diagnosed with hypertrophic cardiomyopathy and the third proband with dilated cardiomyopathy (DCM). The proband affected with DCM also carried a second missense variant in the TMP1 gene (Hershberger et al. 2010). The p.Glu244Asp variant was absent from at least 800 control chromosomes (Hershberger et al. 2009; Hershberger et al. 2010; Rampersaud et al. 2011) and is reported at a frequency of 0.000599 in the European (non-Finnish) population of the Exome Aggregation Consortium. Expression of p.Glu244Asp in bovine and rabbit cardiac myofibrils showed an enhanced maximum level of ATPase activity with no significant difference in calcium ion sensitivity (Yanaga et al. 1999; Matsumoto et al. 2009; Willott et al. 2009). Based on the evidence, the p.Glu244Asp variant is classified as a variant of unknown significance but suspicious for pathogenicity for TNNT2-related disorders. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population. -

Primary familial hypertrophic cardiomyopathy Uncertain:1

Uncertain significance, criteria provided, single submitter

research

CSER _CC_NCGL, University of Washington

Jun 01, 2014

Low GERP score may suggest that this variant may belong in a lower pathogenicity class -

Hypertrophic cardiomyopathy 2 Uncertain:1

Uncertain significance, no assertion criteria provided

clinical testing

Knight Diagnostic Laboratories, Oregon Health and Sciences University

Dec 19, 2015

- -

Dilated cardiomyopathy 1D;C1861864:Hypertrophic cardiomyopathy 2;C2676271:Cardiomyopathy, familial restrictive, 3 Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Invitae

Jan 08, 2024

This sequence change replaces glutamic acid, which is acidic and polar, with aspartic acid, which is acidic and polar, at codon 244 of the TNNT2 protein (p.Glu244Asp). This variant is present in population databases (rs45466197, gnomAD 0.05%), and has an allele count higher than expected for a pathogenic variant. This missense change has been observed in individual(s) with hypertrophic cardiomyopathy and dilated cardiomyopathy (PMID: 7898523, 19412328, 29121657, 30565988). This variant is also known as E251D. ClinVar contains an entry for this variant (Variation ID: 43667). 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 TNNT2 protein function with a positive predictive value of 95%. Experimental studies are conflicting or provide insufficient evidence to determine the effect of this variant on TNNT2 function (PMID: 10085122, 10467159, 10497196, 14722098, 19275886, 30565988). In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance. -

Cardiovascular phenotype Benign:1

Likely benign, criteria provided, single submitter

clinical testing

Ambry Genetics

May 07, 2021

This alteration is classified as likely benign based on a combination of the following: seen in unaffected individuals, population frequency, intact protein function, lack of segregation with disease, co-occurrence, RNA analysis, in silico models, amino acid conservation, lack of disease association in case-control studies, and/or the mechanism of disease or impacted region is inconsistent with a known cause of pathogenicity. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.98
CardioboostCm	Benign	0.095
BayesDel_addAF	Uncertain	0.15	D
BayesDel_noAF	Pathogenic	0.31
Cadd	Uncertain	24
Dann	Uncertain	1.0
Eigen	Benign	0.18
Eigen_PC	Benign	0.072
FATHMM_MKL	Uncertain	0.90	D
M_CAP	Pathogenic	0.60	D
MetaRNN	Uncertain	0.71	D;D;D;D;D;D;D;D;D;D;D
MetaSVM	Uncertain	0.63	D
MutationTaster	Benign	1.0	D;D;D;D;D;D;D;D;D;D
PrimateAI	Uncertain	0.57	T
PROVEAN	Uncertain	-2.5	N;N;.;.;.;.;.;.;D;N;N
REVEL	Pathogenic	0.81
Sift	Uncertain	0.0020	D;D;.;.;.;.;.;.;D;D;D
Sift4G	Uncertain	0.0080	D;D;D;D;T;D;D;D;D;D;.
Polyphen		0.95	.;.;.;.;P;.;.;.;.;.;.
Vest4		0.80
MutPred		0.67		.;.;.;.;Loss of methylation at K257 (P = 0.1193);.;.;.;.;.;.;
MVP		0.94
MPC		0.71
ClinPred		0.39	T
GERP RS		1.8
Varity_R		0.50
gMVP		0.93

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.0		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: rs45466197; hg19: chr1-201330455;