Prevalence of TTR variants detected by whole-exome sequencing in hypertrophic cardiomyopathy

Abstract

Background: A proportion of patients with hypertrophic cardiomyopathy (HCM) have a diagnosis of cardiac amyloidosis. Hereditary transthyretin amyloid cardiomyopathy (ATTRv-CM) is caused by mutations in the TTR gene. Our aim was to study the prevalence of potentially amyloidogenic TTR variants in a whole-exome sequencing (WES) study of a large HCM cohort.

Methods and results: 770 consecutive HCM probands underwent WES and clinical characterisation. Patients with rare or known pathogenic variants in TTR underwent 99mTechnetium labelled 3,3-diphosphono-1,2-propanodicarboxylic acid (DPD) scintigraphy and were retrospectively re-assessed for clinical features of amyloidosis. Two patients had rare TTR variants of unknown significance and four had the known pathogenic V122I (p.V142I) variant (one homozygous and also heterozygous for a likely pathogenic MYL3 variant and another double heterozygous for a likely pathogenic MYBPC3 variant). Four out of 6 patients with TTR variants underwent DPD scintigraphy; the only positive study was in the patient with the homozygous V122I (p.V142I) variant.

Conclusions: Pathogenic TTR variants are rare in carefully assessed HCM patients and may occur in double heterozygosity with pathogenic sarcomere variants. The lack of evidence for an amyloidosis phenotype in all but one TTR variant carrier illustrates the importance of complete clinical evaluation of HCM patients that harbour pathogenic TTR variants.     

Rentabilidad del estudio genético mediante técnicas de next-generation sequencing masiva de pacientes con miocardiopatía arritmogénica de alto riesgo

Introduction and objectives

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited cardiomyopathy characterized by progressive fibrofatty replacement of predominantly right ventricular myocardium. This cardiomyopathy is a frequent cause of sudden cardiac death in young people and athletes. The aim of our study was to determine the incidence of pathological or likely pathological desmosomal mutations in patients with high-risk definite ARVC.

LMNA‐Mediated Arrhythmogenic Right Ventricular Cardiomyopathy and Charcot‐Marie‐Tooth Type 2B1: A Patient‐Discovered Unifying Diagnosis

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an uncommon cardiomyopathy most classically associated with mutations in genes encoding desmosomal proteins. Recent literature has identified mutations in several non‐desmosomal proteins including lamins that may result in the ARVC phenotype. We describe a patient who discovered her own pathogenic LMNA mutation that offered a unifying diagnosis explaining her ARVC and Charcot‐Marie‐Tooth phenotypes as well as musculoskeletal abnormalities. Suspicion for LMNA‐mediated cardiomyopathy should arise in patients with extracardiac manifestations of laminopathies and testing for specific gene mutations may be helpful in establishing an unifying diagnosis.     

Compound and heterozygous mutations of DSG2 identified by Whole Exome Sequencing in arrhythmogenic right ventricular cardiomyopathy/dysplasia with ventricular tachycardia

Backgrounds

This study was designed to identify the pathogenic mutations in two Chinese families of arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D) using the Whole Exome Sequencing (WES).

Deleterious Rare Desmosomal Variants Contribute to Hypertrophic Cardiomyopathy and Are Associated With Distinctive Clinical Features

BackgroundDeleterious rare variants in genes encoding desmosome proteins have been identified as the essential basis of arrhythmogenic cardiomyopathy (ACM) and detected in dilated cardiomyopathy, but the relationship between deleterious rare desmosomal variants and hypertrophic cardiomyopathy (HCM) remains unknown.MethodsWhole exome sequencing was performed in 1000 patients with HCM and 761 non-HCM controls to search for deleterious rare variants in genes encoding desmosomal proteins including PKP2, JUP, DSC2, DSG2, and DSP. Clinical phenotypes were assessed in patients with HCM, and patients with deleterious rare desmosomal variants underwent evaluation of ACM revised Task Force Criteria.ResultsA total of 27 deleterious rare desmosomal variants were present in 24 (2.4%) patients with HCM and 5 (0.66%) controls. The variants were more prevalent in the patients with HCM than in the controls (P = 0.004). The majority of patients possessing deleterious rare desmosomal variants could not be diagnosed as ACM. Moreover, the patients with deleterious rare desmosomal variants possessed several distinctive clinical features compared with patients without such variants, including a higher incidence of nonsustained ventricular tachycardia (29.2% vs 4.5%, P < 0.001), left bundle branch block (33.3% vs 1.6%, P < 0.001), and right ventricular involvement for an HCM phenotype (29.2% vs 0.30%, P < 0.001).ConclusionsWe screened deleterious rare desmosomal variants in a large HCM case-control cohort and found deleterious rare desmosomal variants can be relevant to HCM. Moreover, our data indicated deleterious rare desmosomal variants were associated with distinctive clinical features of HCM. These findings require validation in other HCM cohorts.     

Arrhythmogenic right ventricular cardiomyopathy

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited cardiomyopathy accounting for life-threatening ventricular tachyarrhythmias and sudden death in young individuals and athletes. Over the past years, mutations in desmosomal genes have been identified as disease-causative. However, genetic heterogeneity and variable phenotypic expression alongside with diverse disease progression still render the evaluation of its prognostic implication difficult. ARVC was initially entered into the canon of cardiomyopathies of the World Health Organization in 1995, and international efforts have resulted in the 2010 modified diagnostic criteria for ARVC. Despite all additional insights into pathophysiology, clinical management, and modern risk stratification, under-/misdiagnosing of ARVC remains a problem and hampers reliable statements on the incidence, prevalence, and natural course of the disease. This review provides a comprehensive overview of the current literature on the pathogenesis, diagnosis, treatment, and prognosis of ARVC and sheds some light on potential new developments in these areas.     

Insights Into Hypertrophic Cardiomyopathy Evaluation Through Follow-up of a Founder Pathogenic Variant

Introduction and objectives

Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac disease. The current challenge relies on the accurate classification of the pathogenicity of the variants. Transthoracic echocardiography (TTE) is recommended at initial evaluation and cardiac magnetic resonance (CMR) imaging should also be considered. We aimed to reappraise the penetrance and clinical expression of the MYBPC3 p.G263* variant.

Absence of a primary role for TTN missense variants in arrhythmogenic cardiomyopathy: From a clinical and pathological perspective

Background

Arrhythmogenic cardiomyopathy (ACM) is an inheritable heart disease characterized by fibro‐fatty replacement of the myocardium. TTN missense variants were previously reported as a pathogenic factor for ACM.

Hypothesis

TTN missense variants are commonly identified in ACM, but have limited effect on the phenotype of ACM.

Methods

We sequenced 15 ACM‐related genes in 35 patients who had a heart transplantation and quantified myocardium, and fibrous and adipose tissue in blocks of the explanted heart. Clinical and pathological characteristics were compared between patients with TTN variants and others. Pedigree analysis was performed in 3 families with TTN variants.

Results

TTN variants were detected in 11 patients (all missense, 9 heterozygous and 2 oligogenic form). The TTN truncating variant was absent in the cohort. Patients with TTN variants had late onset age of the disease (31 ±13 years vs 17 ±3 years, P = 0.049) and age of heart transplantation (41 ±14 years vs 24 ±9 years, P = 0.027), larger left ventricle end‐diastolic diameter (62 ±10 mm vs 45 ±10 mm, P = 0.019), smaller right ventricular outflow tract (34 ±14 mm vs 50 ±15 mm, P = 0.046), more myocardium (40.8% ±29.4% vs 13.8% ±11.0%, P = 0.017), and less adipose tissue (43.0% ±30.9% vs 66.9% ±18.5%, P = 0.036) in right ventricle than those with desmosomal variants. There was few difference between patients with TTN variants and those without variants. Pedigrees showed none of the family members with TTN missense variants had a disease phenotype, indicating a very low penetrance.

Conclusions

TTN missense variants was commonly identified in ACM patients in this cohort, but hardly played a primary role in ACM as causative variants.     

Slow cardiac myosin regulatory light chain 2 (MYL2) was down-expressed in chronic heart failure patients

Background

Genetic studies have shown that many slow cardiac myosin regulatory light chain 2 (MYL2) gene mutations can cause hypertrophic cardiomyopathy, which is one of the most common causes of heart failure (HF). But until now there has been no pathological or histological evidence that MYL2 may be associated with HF development. Recent microarray studies indicated that myosin heavy chain expression changed in the pathological process of HF. Because MYL2 is a regulatory component of myosin heavy polypeptide, the role of MYL2 protein in HF needs to be studied.

Hypothesis

The level of expression of MYL2 may change in the heart tissue of patients with chronic HF.

Methods

We collected 28 human right auricle samples, 16 from chronic HF patients and 12 from healthy control subjects. Immunohistochemistry was carried out to observe the tissue‐expression pattern of the MYL2 protein and Western blot methods were performed to quantify the relative MYL2 expression level.

Results

In chronic HF patients, the MYL2 protein level decreased significantly compared with normal controls (t test P < 0.05). Among the 16 HF patients, MYL2 expression in the severe HF group (New York Heart Association [NYHA] class III or IV) was even lower than that of the moderate HF group (NYHA class II) (t test P < 0.05).

Conclusions

MYL2 was down‐expressed in HF tissues, and our findings suggested that MYL2 may play a role in the development and progression of chronic HF. Copyright © 2008 Wiley Periodicals, Inc. The first two authors contributed equally to this work. This study was supported by the National Natural Science Foundation of China (30800457, 30900803) and the Natural Science Program of Hubei Province (2009CDZ013). This study has no relationship with any kind of industry. The authors have no other funding, financial relationships, or conflicts of interest to disclose.     

Titin Mutation in Familial Restrictive Cardiomyopathy

Background

Familial restrictive cardiomyopathy (RCM) caused by a single gene mutation is the least common of the inherited cardiomyopathies. Only a few RCM-causing mutations have been described. Most mutations causing RCM are located in sarcomere protein genes which also cause hypertrophic cardiomyopathy (HCM).Other genes associated with RCM include the desmin and familial amyloidosis genes. In the present study we describe familial RCM with severe heart failure triggered by a de novo mutation in TTN, encoding the huge muscle filament protein titin.

Methods and results

Family members underwent physical examination, ECG and Doppler echocardiogram studies. The family comprised 6 affected individuals aged 12–35 years. Linkage to candidate loci was performed, followed by gene sequencing. Candidate loci/gene analysis excluded 18 candidate genes but showed segregation with a common haplotype surrounding the TTN locus. Sequence analysis identified a de novo mutation within exon 266 of the TTN gene, resulting in the replacement of tyrosine by cysteine. p.Y7621C affects a highly conserved region in the protein within a fibronectin-3 domain, belonging to the A/I junction region of titin. No other disease-causing mutation was identified in cardiomyopathy genes by whole exome sequencing.

Conclusions

Our study shows, for the first time, that mutations in TTN can cause restrictive cardiomyopathy. The giant filament titin is considered to be a determinant of a resting tension of the sarcomere and this report provides genetic evidence of its crucial role in diastolic function.     

The genetic basis of cardiomyopathy

Cardiomyopathies are the primary disorders of cardiac myocytes, and their cause is usually genetic. The three common forms are hypertrophic cardiomyopathy (HCM), dilated cardiomyopathy (DCM), and arrhythmogenic right ventricular cardiomyopathy (ARVC). Mutations in sarcomeric proteins typically cause HCM and, less commonly, DCM. Mutations in cytoskeletal proteins cause DCM, and those in desmosomal proteins cause ARVC. The pathways from mutations to the clinical phenotype could be categorized into three stages of initial functional defects leading to expression and activation of molecular events that mediate development of morphologic and structural phenotypes. Advances in understanding the molecular genetics and pathogenesis of cardiomyopathies could provide the opportunity for preclinical diagnosis and interventions to prevent, attenuate, or reverse the evolving phenotypes.     

Population-based variation in cardiomyopathy genes

Background- Hypertrophic cardiomyopathy and dilated cardiomyopathy arise from mutations in genes encoding sarcomere proteins including MYH7, MYBPC3, and TTN. Genetic diagnosis of cardiomyopathy relies on complete sequencing of the gene coding regions, and most pathogenic variation is rare. The 1000 Genomes Project is an ongoing consortium designed to deliver whole genome sequence information from an ethnically diverse population and, therefore, is a rich source to determine both common and rare genetic variants. Methods and Results- We queried the 1000 Genomes Project database of 1092 individuals for exonic variants within 3 sarcomere genes MHY7, MYBPC3, and TTN. We focused our analysis on protein-altering variation, including nonsynonymous single nucleotide polymorphisms, insertion/deletion polymorphisms, or splice site altering variants. We identified known and predicted pathogenic variation in MYBPC3 and MYH7 at a higher frequency than what would be expected based on the known prevalence of cardiomyopathy. We also found substantial variation, including protein-disrupting sequences, in TTN. Conclusions- Cardiomyopathy is a genetically heterogeneous disorder caused by mutations in multiple genes. The frequency of predicted pathogenic protein-altering variation in cardiomyopathy genes suggests that many of these variants may be insufficient to cause disease on their own but may modify phenotype in a genetically susceptible host. This is suggested by the high prevalence of TTN insertion/deletions in the 1000 Genomes Project cohort. Given the possibility of additional genetic variants that modify the phenotype of a primary driver mutation, broad-based genetic testing should be employed.     

Value of genotyping and scar-phenotyping for VT ablation procedures in patients with nonischemic left ventricular cardiomyopathies

Introduction

Variants of cardiomyopathy genes in patients with nonischemic cardiomyopathy (NICM) generate various phenotypes of cardiac scar and delayed enhancement cardiac magnetic resonance (DE-CMR) imaging which may impact ventricular tachycardia (VT) management.

Methods

The objective was to compare the findings of cardiomyopathy genetic testing on DE-CMR imaging and long-term outcomes among patients with NICM undergoing VT ablation procedures. Image phenotyping and genotyping were performed in a consecutive series of patients referred for VT ablation and correlated to survival free of VT. Scar depth index (SDI) (% of scar at 0-3 mm, 3-5 mm and >5 mm projected on the closest endocardial surface) was determined.

Results

Forty-three patients were included (11 women, 55 ± 14 years, ejection fraction (EF) 45 ± 16%) and were followed for 3.4 ± 2.9 years. Pathogenic variants (PV) were identified in 16 patients (37%) in the following genes: LMNA (n = 5), TTN (n = 5), DSP (n = 2), AMLS1 (n = 1), MYBPC3 (n = 1), PLN (n = 1), and SCN5A (n = 1). A ring-like septal scar (RLSS) pattern was more often seen in patients with pathogenic variants (66% vs 15%, p = .001). RLSS was associated with deeper seated scars (SDI >5 mm 30.6 ± 22.6% vs 12.4 ± 16.2%, p = .005), and increased VT recurrence (HR 5.7 95% CI[1.8-18.4], p = .003). After adjustment for age, sex, EF, and total scar burden, the presence of a PV remained independently associated with worse outcomes (HR 4.7 95% CI[1.22-18.0], p = .02).

Conclusions

Preprocedural genotyping and scar phenotyping is beneficial to identify patients with a favorable procedural outcome. Some PVs are associated with an intramural, deeper seated scar phenotype and have an increase of VT recurrence after ablation.     

SCN5A mutation in Chinese patients with arrhythmogenic right ventricular dysplasia

Background

Arrhythmogenic right ventricular dysplasia (ARVD) is a genetically determined disorder, characterized by two components: cardiomyopathy and arrhythmia. To date, the ion channel-related pathogenesis underlying this phenomenon has been poorly understood. The aim of this study was to systematically evaluate the sodium channel variants in Chinese patients with ARVD.

Patients and methods

Patients meeting the diagnostic guidelines of ARVD revised in 2010 were enrolled. All exons and exon-intron boundaries of the SCN5A gene and desmosomal genes known to be associated with ARVD, including DSC2, DSG2, DSP, JUP, and PKP2, were sequenced by direct DNA sequencing. A total of 12 unrelated index patients were included in the study.

Results

Eight of the patients developed ventricular tachycardia (VT) and ventricular fibrillation (VF), one of them showed epsilon wave, one of them showed type-1 Brugada wave, seven of them exhibited syncope or dizziness, and none of the patients had a family history of SCD. A new missense heterozygote mutation, I137M, in SCN5A was found in proband 5 with recurrent palpitations and a high incidence of VT. I137M is in exon 4 of SCN5A, at the S1 segment in domain I of Nav1.5, which predicted a substitution of isoleucine for methionine at codon site 137 (p. Ile137Met, I137M). I137M was not detected in 400 healthy control chromosomes from individuals of the same ethnic background, which indicated that this mutation was a conservative site in the SCN5A gene, and the encoded protein Nav1.5 might have a functional defect resulting in arrhythmia.

Conclusion

This was the first study to systematically investigate sodium channel variants in Chinese patients with ARVD; a new SCN5A mutation, I137M, was found. This finding may provide new evidence of the genetic pathogenesis of ARVD in Chinese patients, implying that the SCN5A gene should be screened in patients with ARVD and VT/VF.     

Molecular characterization of Portuguese patients with dilated cardiomyopathy

Introduction

Dilated cardiomyopathy (DCM) is a disease of the heart muscle characterized by ventricular dilatation and impaired systolic function. Familial forms account for 30-50% of cases. Autosomal dominant inheritance is the predominant pattern of transmission. Causal genetic variants have been identified in several genes and molecular diagnosis has implications for genetic counseling and risk stratification.

Human Molecular Genetic and Functional Studies Identify TRIM63, Encoding Muscle RING Finger Protein 1, as a Novel Gene for Human Hypertrophic Cardiomyopathy

Rationale: A delicate balance between protein synthesis and degradation maintains cardiac size and function. TRIM63 encoding Muscle RING Finger 1 (MuRF1) maintains muscle protein homeostasis by tagging the sarcomere proteins with ubiquitin for subsequent degradation by the ubiquitin-proteasome system (UPS). Objective: To determine the pathogenic role of TRIM63 in human hypertrophic cardiomyopathy (HCM). Methods and Results: Sequencing of TRIM63 gene in 302 HCM probands (250 white individuals) and 339 control subjects (262 white individuals) led to identification of 2 missense (p.A48V and p.I130M) and a deletion (p.Q247*) variants exclusively in the HCM probands. These 3 variants were absent in 751 additional control subjects screened by TaqMan assays. Likewise, rare variants were enriched in the white HCM population (11/250, 4.4% versus 3/262, 1.1%, respectively, P=0.024). Expression of the mutant TRIM63 was associated with mislocalization of TRIM63 to sarcomere Z disks, impaired auto-ubiquitination, reduced ubiquitination and UPS-mediated degradation of myosin heavy chain 6, cardiac myosin binding protein C, calcineurin (PPP3CB), and p-MTOR in adult cardiac myocytes. Induced expression of the mutant TRIM63 in the mouse heart was associated with cardiac hypertrophy, activation of the MTOR-S6K and calcineurin pathways, and expression of the hypertrophic markers, which were normalized on turning off expression of the mutant protein. Conclusions: TRIM63 mutations, identified in patients with HCM, impart loss-of-function effects on E3 ligase activity and are probably causal mutations in HCM. The findings implicate impaired protein degradation in the pathogenesis of HCM.     

Re‐evaluating pathogenicity of variants associated with the long QT syndrome

1 Introduction

Genetic testing for congenital long QT syndrome (LQTS) has become common. Recent studies have shown that some variants labelled as pathogenic might be misclassified due to sparse case reports and relatively common allele frequencies (AF) in the general population. This study aims to evaluate the presence of LQTS‐associated variants in the Genome Aggregation Database (gnomAD) population, and assess the functional impact of these variants.

2 Methods and results

Variants associated with LQTS from the Human Gene Mutation Database were extracted and matched to the gnomAD to evaluate population‐based AF. We used MetaSVM to predict the function of LQTS variants. Allele distribution by protein topology in KCNQ1, KCNH2, and SCN5A was compared between gnomAD (n = 123,136) and a cohort of LQTS patients aggregated from eight published studies (n = 2,683). Among the 1,415 LQTS‐associated single nucleotide variants in 30 genes, 347 (25%) are present in gnomAD; 24% of the 347 variants were predicted as functionally tolerated compared with 4% of variants not present in gnomAD (P < 0.001). Of the 347 pathogenic variants in gnomAD, seven (2%) had an AF of ≥ 0.001 and 65 (19%) variants had an AF of ≥ 0.0001. In KCNQ1, KCNH2, and SCN5A, allele distribution by protein functional region was significantly different with gnomAD alleles appearing less frequently in highly pathogenic domains than case alleles.

3 Conclusion

A significant number of LQTS variants have insufficient evidence for pathogenicity and relatively common AF in the general population. Caution should be used when ascribing pathogenicity to these variants.     

Electrocardiographic (ECG) clues to differentiate idiopathic right ventricular outflow tract tachycardia (RVOTT) from arrhythmogenic right ventricular cardiomyopathy (ARVC)

Introduction

Arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D) is a genetic cardiomyopathy that most commonly affects young adults. The most commonly observed reason of death in patients suffering from ARVC/D is sudden cardiac death (SCD). On the other hand, idiopathic right ventricular outflow tract tachycardia (RVOT VT) usually has a benign course. Both of the entities may have ventricular tachycardia (VT) with left bundle branch block (LBBB) pattern and inferior axis. We tried to propose new discriminating electrocardiographic indices for differentiation of foretold entities.

Material and method

This was a retrospective study. We reviewed records of patients admitted between 2003 and 2012 with the diagnosis of either ARVC/D or RVOT VT that presented with VT (LBBB morphology).

Result

A total of fifty nine patients (30 RVOT VT and 29 ARVC/D) were enrolled. In ARVC/D group, men were dominant while the reverse was true of RVOT VT. Palpitation was more common in the RVOT VT group (90% vs. 66.7%), but aborted SCD and sustained VT were more common in ARVC/D group. The new ECG criteria proposed by us mean QRS duration in V1–V3, QRS difference in right and left precordial leads, S wave upstroke duration, JT interval dispersion, QRS and JT interval of right to left precordial leads were all significantly longer in ARVC/D when compared to RVOT VT patients (p < 0.001).

Conclusion

The proposed ECG criteria can be used for non-invasive diagnosis of ARVC/D and incorporation in the future updates of ARVC/D task force criteria.     

No variants in the cardiac actin gene in Finnish patients with dilated or hypertrophic cardiomyopathy

Background Dilated and hypertrophic cardiomyopathies are primary myocardial diseases that cause considerable morbidity and mortality. Although these cardiomyopathies are clinically heterogeneous, genetic factors play an important role in their etiology and pathogenesis. The defects in the cardiac actin (ACTC) gene can cause both cardiomyopathies. The aim of our study was to screen for variants in the ACTC gene in patients with dilated or hypertrophic cardiomyopathy from Eastern Finland. Materials and Methods Altogether, 32 patients with dilated and 40 patients with hypertrophic cardiomyopathy were included in the study. Commonly approved diagnostic criteria were applied, and secondary cardiomyopathies were carefully excluded. All 6 exons of the ACTC gene were amplified with polymerase chain reaction and screened for variants with single-strand conformation polymorphism analysis. Results and Conclusion We did not find any new or previously reported variants. Our results indicate that defects in the ACTC gene do not explain dilated cardiomyopathy or hypertrophic cardiomyopathy in subjects from Eastern Finland and confirm earlier results that the ACTC gene does not play an important role in the genetics of dilated or hypertrophic cardiomyopathies. (Am Heart J 2002;143:11-4.)