Variant classified as Uncertain Significance - Favor Pathogenic. The p.Arg159Gln variant in TNNT2 has been identified in 1 Caucasian individual with peripartum DCM, 1 Caucasian individual with LVNC, and 1 Black individual with DCM. It segre gated with disease in 1 affected relative (Hershberger 2008, Morales 2010, LMM u npublished data). Studies have shown that the p.Arg159Gln variant may affect pro per myocyte function (Hershberger 2009); however, this in vitro assay may not ac curately represent biological function. This variant has not been identified by large population studies and was predicted to be pathogenic using a computationa l tool clinically validated by our laboratory. This tool's pathogenic prediction is estimated to be correct 94% of the time (Jordan 2011). In summary, while the re is some suspicion for a pathogenic role, the clinical significance of the p.A rg159Gln variant is uncertain.

The c.476G>A (p.R159Q) alteration is located in exon 11 (coding exon 10) of the TNNT2 gene. This alteration results from a G to A substitution at nucleotide position 476, causing the arginine (R) at amino acid position 159 to be replaced by a glutamine (Q). Based on insufficient or conflicting evidence, the clinical significance of this alteration remains unclear.

Reported in association with cardiomyopathy (PMID: 19412328, 21483645, 27532257, 31983221); Reported to decrease calcium sensitivity of the myofilaments, which could lead to altered contractility dynamics of the heart; however, it is not known whether these findings are biological or clinically relevant in vivo (PMID: 20031601); Not observed at significant frequency in large population cohorts (gnomAD); In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; De novo variant with confirmed parentage in a patient referred for genetic testing at GeneDx; however, the reported clinical features are only partially consistent with the features typically observed in individuals with pathogenic variants in this gene; This variant is associated with the following publications: (PMID: 20458009, 27532257, 21483645, 31983221, 27535533, 33500567, 36166435, 37466024, 35653365, 19412328, 20031601)

Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as Likely pathogenic. 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) 0115 - Variants in this gene are known to have variable expressivity. p.(Arg92Gln) has been described in families which has both dilated cardiomyopathy (DCM) and hypertrophic cardimyopathy (PMID: 26507537). (I) 0200 - Variant is predicted to result in a missense amino acid change from arginine to glutamine. (I) 0251 - This variant is heterozygous. (I) 0301 - Variant is absent from gnomAD (both v2 and v3). (SP) 0502 - Missense variant with conflicting in silico predictions and uninformative conservation. (I) 0600 - Variant is located in the annotated troponin domain (NCBI). (I) 0705 - No comparable missense variants have previous evidence for pathogenicity. (I) 0808 - Previous reports of pathogenicity for this variant are conflicting. This variant has been reported in at least three unrelated individuals with DCM (ClinVar, PMID: 20458009, 19412328, 20031601, 27532257, 31983221), and has been described as both likely pathogenic and as a VUS (ClinVar, LOVD). Additionally, it has been observed in another individual with DCM and classified using ACMG guidelines as pathogenic. However, this publication is yet to be peer reviewed (Lesurf R, et al. (2020)). (I) 0905 - No published segregation evidence has been identified for this variant. (I) 1002 - This variant has moderate functional evidence supporting abnormal protein function. Functional studies using porcine cardiac fibres demonstrated that this variant results in reduced calcium sensitivity (PMID: 20031601). (SP) 1208 - Inheritance information for this variant is not currently available in this individual. (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign

This sequence change replaces arginine, which is basic and polar, with glutamine, which is neutral and polar, at codon 159 of the TNNT2 protein (p.Arg159Gln). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individuals with dilated cardiomyopathy (PMID: 20031601, 27532257, 31983221, 33500567, 35653365, 36166435; internal data). This variant is also known as p.Arg169Gln. ClinVar contains an entry for this variant (Variation ID: 43647). Invitae Evidence Modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) indicates that this missense variant is not expected to disrupt TNNT2 protein function with a negative predictive value of 80%. Experimental studies have shown that this missense change affects TNNT2 function (PMID: 20031601). For these reasons, this variant has been classified as Pathogenic.