Cardiomyopathy-related Genes Research Articles

Molecular Diagnosis of Primary Cardiomyopathy in 231 Unrelated Pediatric Cases by Panel-Based Next-Generation Sequencing: A Major Focus on Five Carriers of Biallelic TNNI3 Pathogenic Variants.

Pediatric cardiomyopathies are clinically heterogeneous heart muscle disorders associated with significant morbidity and mortality for which substantial evidence for a genetic contribution was previously reported. We present a detailed molecular investigation of a cohort of 231 patients presenting with primary cardiomyopathy below the age of 18 years. Cases with pediatric cardiomyopathies were analyzed using a next-generation sequencing (NGS) workflow based on a virtual panel including 57 cardiomyopathy-related genes. This molecular approach led to the identification of 69 cases (29.9% of the cohort) genotyped as a carrier of at least one pathogenic or likely pathogenic variant. Fourteen patients were carriers of two mutated alleles (homozygous or compound heterozygous) on the same cardiomyopathy-related gene, explaining the severe clinical disease with early-onset cardiomyopathy. Homozygous TNNI3 pathogenic variants were detected for five unrelated neonates (2.2% of the cohort), with four of them carrying the same truncating variant, i.e. p.Arg69Alafs*8. Our study confirmed the importance of genetic testing in pediatric cardiomyopathies. Discovery of novel pathogenic variations is crucial for clinical management of affected families, as a positive genetic result might be used by a prospective parent for prenatal genetic testing or in the process of pre-implantation genetic diagnosis.

Chemotherapy induced cardiotoxicity: on the way to personalized risk stratification based on genetic variants

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): 1.Flanders Research Foundation- FWO (doctoral research grant) 2.UZA Foundation-Wolvenbos Grant 2019 Background Anthracycline chemotherapy is a cornerstone in the treatment of several malignancies. However, it causes substantial toxicity, of which cardiotoxicity is the most frequent and most severe (1). Ideally, patients at risk for cardiotoxicity are identified prior to treatment, however, traditional patient- and treatment-related factors fail to fully predict the individual risk at the moment. Improved risk stratification for ANT-induced cardiotoxicity could possibly be reached by taking genetic risk factors into account. Purpose We assessed the prevalence of variants in genes encoding for cardiac proteins in the development of anthracycline-induced cardiotoxicity. Methods Patients presenting with early (< 1y) or late (> 1y) anthracycline-induced cardiotoxicity between 1995 and 2020 were included. Cardiotoxicity was defined as a decline in left ventricular ejection fraction (LVEF) to <50% and a ≥10% reduction from baseline by echocardiography. Genetic analysis was offered using a haloplex gene panel composed of 59 known cardiomyopathy-related genes. Variants were classified according to ACMG guidelines(2). Both variants of unknown significance (VUS, class 3) and likely pathogenic/pathogenic variants (class 4 and 5) were taken into account. Frequency of genetic variants was compared to a matched positive control cohort of patients with dilated cardiomyopathy (DCM). Results Forty-two patients that had developed cardiotoxicity (mean age at time of diagnosis 54yrs) fulfilled the inclusion criteria and agreed to genetic testing. The majority of patients had been treated for lymphoma (52.4%) or breast cancer (38.1%). Total equivalent dose of anthracyclines averaged 285.5 mg/m² doxorubicin. Patients showed an average reduction in LVEF of 27.1% (+- 10.1). In 18 patients (42.9 %), a variant could be identified; 15 patients carried a variant of unknown significance (VUS) and 3 carried a likely pathogenic variant. In total 29 variants different variants were identified in 18 distinct individuals. Genetic yield was independent of anthracycline-dose and the presence of other cardiovascular risk factors. The genetic yield in cardiotoxicity patients did not differ significantly from this in DCM-controls (18/42 VUS, 5/42 (likely) pathogenic variants, p = 0.19). After initiation of standard heart failure therapy, 6 (33.3%) of variant carriers showed incomplete recovery of cardiac function, compared to only 3 (12.5%) of patients who did not carry a variant (p=0.103). Conclusion Prevalence of genetic variants in anthracycline-induced cardiotoxicity is similar to that in isolated dilated cardiomyopathy. This finding supports a second-hit mechanism in which anthracycline chemotherapy, uncovers a genetically determined cardiomyopathy. As such, genetic variants in cardiomyopathy genes could improve individualized risk stratification prior to anthracycline chemotherapy and can assist in personalized prevention of this feared side effect of chemotherapy.

The Genetic Architecture of Hypertrophic Cardiomyopathy in Hungary: Analysis of 242 Patients with a Panel of 98 Genes.

Hypertrophic cardiomyopathy (HCM) is a primary disease of the myocardium most commonly caused by mutations in sarcomeric genes. We aimed to perform a nationwide large-scale genetic analysis of a previously unreported, representative HCM cohort in Hungary. A total of 242 consecutive HCM index patients (127 men, 44 ± 11 years) were studied with next generation sequencing using a custom-designed gene-panel comprising 98 cardiomyopathy-related genes. A total of 90 patients (37%) carried pathogenic/likely pathogenic (P/LP) variants. The percentage of patients with P/LP variants in genes with definitive evidence for HCM association was 93%. Most of the patients with P/LP variants had mutations in MYBPC3 (55 pts, 61%) and in MYH7 (21 pts, 23%). Double P/LP variants were present in four patients (1.7%). P/LP variants in other genes could be detected in ≤3% of patients. Of the patients without P/LP variants, 46 patients (19%) carried a variant of unknown significance. Non-HCM P/LP variants were identified in six patients (2.5%), with two in RAF1 (p.Leu633Val, p.Ser257Leu) and one in DES (p.Arg406Trp), FHL1 (p.Glu96Ter), TTN (p.Lys23480fs), and in the mitochondrial genome (m.3243A>G). Frameshift, nonsense, and splice-variants made up 82% of all P/LP MYBPC3 variants. In all the other genes, missense mutations were the dominant form of variants. The MYBPC3 p.Gln1233Ter, the MYBPC3 p.Pro955ArgfsTer95, and the MYBPC3 p.Ser593ProfsTer11 variants were identified in 12, 7, and 13 patients, respectively. These three variants made up 36% of all patients with identified P/LP variants, raising the possibility of a possible founder effect for these mutations. Similar to other HCM populations, the MYBPC3 and the MYH7 genes seemed to be the most frequently affected genes in Hungarian HCM patients. The high prevalence of three MYBPC3 mutations raises the possibility of a founder effect in our HCM cohort.

Identification and Validation of Dilated Cardiomyopathy-Related Genes via Bioinformatics Analysis.

PurposeDilated cardiomyopathy (DCM) is a type of cardiomyopathy that can easily cause heart failure and has a high mortality rate. Therefore, there is an urgent need to study the underlying mechanism of action of dilated cardiomyopathy. In the present study, we aimed to explore potential miRNA–mRNA pairs and drugs related to DCM.MethodsThe Microarray data were collected from the Gene Expression Omnibus (GEO) database. Bioinformatics analysis differentially expressed miRNAs and mRNAs in each microarray were obtained. The target genes of miRNAs were obtained from the miRWalk 2.0 database, and the intersection of these two gene sets (miRNA target genes and differentially expressed mRNAs in the microarray) was obtained. Pathway and Gene Ontology (GO) enrichment analyses were performed in the KOBAS database. Cytoscape software was used to construct the miRNA–mRNA network, and the final hub genes were obtained. Furthermore, we predicted several candidate drugs related to hub genes using DSigDB database. To confirm the abnormal expression of hub genes, qRT-PCR was performed.ResultsIn total, eight differentially expressed miRNAs and 92 differentially expressed mRNAs were identified. In addition, 47 differentially expressed miRNA target genes were identified. According to the analysis results of the miRNA-mRNA network, we identified hsa-miR-551b-3p, hsa-miR-770-5p, hsa-miR-363-3p, PIK3R1, DDIT4, and CXCR4 as hub genes in DCM. Several candidate drugs, which are related to the hug genes, were identified.ConclusionIn conclusion, in our study, we identified several hub genes that may be involved in the pathogenesis of DCM. Several drugs related to these hub genes may be used as clinical therapeutic candidates.

Identification of cardiomyopathy-related core genes through human metabolic networks and expression data

BackgroundCardiomyopathy is a complex type of myocardial disease, and its incidence has increased significantly in recent years. Dilated cardiomyopathy (DCM) and ischemic cardiomyopathy (ICM) are two common and indistinguishable types of cardiomyopathy.ResultsHere, a systematic multi-omics integration approach was proposed to identify cardiomyopathy-related core genes that could distinguish normal, DCM and ICM samples using cardiomyopathy expression profile data based on a human metabolic network. First, according to the differentially expressed genes between different states (DCM/ICM and normal, or DCM and ICM) of samples, three sets of initial modules were obtained from the human metabolic network. Two permutation tests were used to evaluate the significance of the Pearson correlation coefficient difference score of the initial modules, and three candidate modules were screened out. Then, a cardiomyopathy risk module that was significantly related to DCM and ICM was determined according to the significance of the module score based on Markov random field. Finally, based on the shortest path between cardiomyopathy known genes, 13 core genes related to cardiomyopathy were identified. These core genes were enriched in pathways and functions significantly related to cardiomyopathy and could distinguish between samples of different states.ConclusionThe identified core genes might serve as potential biomarkers of cardiomyopathy. This research will contribute to identifying potential biomarkers of cardiomyopathy and to distinguishing different types of cardiomyopathy.

Impact of genetic tests on survivors of paediatric sudden cardiac arrest

ObjectiveTo retrospectively investigate the clinical spectrum, genetic profiles and outcomes of survivors of paediatric sudden cardiac arrest (SCA).Design and patientsAll 66 patients (aged 1–20 years), with unexpected SCA or syncope...

Long-term outcomes and left ventricular diastolic function of sarcomere mutation-positive and mutation-negative patients with hypertrophic cardiomyopathy: a prospective cohort study.

Hypertrophic cardiomyopathy (HCM) is an inheritable disease that leads to sudden cardiac death and heart failure (HF). Sarcomere mutations (SMs) have been associated with HF. However, the differences in ventricular function between SM-positive and SM-negative HCM patients are poorly characterized. Of the prospectively enrolled 374 unrelated HCM patients in Taiwan, 115 patients underwent both 91 cardiomyopathy-related gene screening and cardiovascular magnetic resonance (45.6 ± 10.6 years old, 76.5% were male). Forty pathogenic/likely pathogenic mutations were identified in 52 patients by next-generation sequencing. The SM-positive group were younger at first cardiovascular event (P = 0.04) and progression to diastolic HF (P = 0.02) with higher N-terminal pro-brain natriuretic peptide (NT-proBNP) [New York Heart Association (NYHA) Class III/IV symptoms with left ventricular ejection fraction > 55%] than the SM-negative group (P < 0.001). SM-positive patients had a greater extent of late gadolinium enhancement (P = 0.01), larger left atrial diameter (P = 0.03), higher normalized peak filling rate (PFR) and PFR ratio, and a greater reduction in global longitudinal strain than SM-negative patients (all P ≤ 0.01). During mean lifelong follow-up time (49.2 ± 15.6 years), SM-positive was a predictor of earlier HF (NYHA Class III/IV symptoms) after multivariate adjustment (hazard ratio 3.5; 95% confidence interval 1.3-9.7; P = 0.015). SM-positive HCM patients had a higher extent of myocardial fibrosis and more severe ventricular diastolic dysfunction than those without, which may contribute to earlier onset of advanced HF, suggesting the importance of close surveillance and early treatment throughout life.

Lymphocytic myocarditis: a genetically predisposed disease?

Abstract Background Clinical presentation of myocarditis is extremely heterogeneous from asymptomatic to overt severe heart failure (HF). A complex interaction between pre-existing genetic background and inflammation might be responsible for this heterogeneity. Purpose The aim of the present study was to investigate whether positive genetic background for pathogenic cardiomyopathy-related variants might underlie a higher susceptibility to left ventricular dysfunction in patients with active lymphocytic myocarditis. Methods We prospectively performed genetic tests in 36 patients (46±15 years; 61% males; no relatives included) with biopsy-proven active lymphocytic myocarditis according to Dallas criteria and immunohistochemistry. Only pathogenic (P) or likely pathogenic (LP) variants were considered. Results After genetic test, 31% of patients (n=11) were carriers of P/LP truncating variants in structural Cardiomyopathy related genes: Titin (TTN, n=8, 73%), Desmoplakin (DSP, n=1), Filamin C (FLNC, n=1) and RNA binding protein 20 (RBM20, n=1). Among the 27 patients presenting with HF and LV dysfunction, the positive genetic yield was similar to the total cohort (n=9, 34%; 90% with TTN). Two out of six arrhythmic patients (30%) were carriers in arrhythmogenic genes (i.e. DSP and FLNC), whereas no patients with infarct-like presentation were carriers. During follow-up, 44% of patients (n=16) presented normal Left Ventricular Ejection Fraction (LVEF). Carriers had a lower rate of LVEF normalization compared to non-carriers (18% vs 56%, respectively; p=0.035). Conclusion Positive genetic testing for cardiomyopathy-related-genes might be found in a non-negligible percentage of patients with biopsy-proven myocarditis, especially if presenting with heart failure and LV dysfunction. Compared to non-carriers, carriers of P/LP variants show lower likelihood of LVEF normalization during follow-up. Funding Acknowledgement Type of funding source: None

Prevalence and Prognostic Impact of Pathogenic Variants in Patients With Dilated Cardiomyopathy Referred for Ventricular Tachycardia Ablation.

This study aimed to assess the frequency of (likely) pathogenic variants (LP/Pv) among dilated cardiomyopathy (DCM) ventricular tachycardia (VT) patients referred for CA and their impact on procedural outcome and long-term prognosis. The prevalence of genetic variants associated with monomorphic VT among DCM is unknown. Ninety-eight consecutive patients (age 56 ± 15 years; 84% men, left ventricular ejection fraction [LVEF] 39±12%) referred for DCM-VT ablation were included. Patients underwent electroanatomical mapping and testing of≥55 cardiomyopathy-related genes. Mapping data were analyzed for low-voltage areas and abnormal potentials. LP/Pv-positive (LP/Pv+) patients were compared with LP/Pv-negative (LP/Pv-) patients and followed for VT recurrence and mortality. In 37 (38%) patients, LP/Pv were identified, most frequently LMNA (n=11 of 37, [30%]), TTN (n=6 of 37, [16%]), PLN (n=6 of 37, [16%]), SCN5A (n=3 of 37, [8%]), RBM20 (n=2 of 37, [5%]) and DSP (n=2 of 37, [5%]). LP/Pv+ carriers had lower LVEF (35 ± 13% vs. LP/Pv-: 42 ± 11%; p=0.005) and were less often men (n=27 [73%] vs. n=55 [90%]; p=0.03). After a median follow-up of 2.4 years (interquartile range: 0.9 to 4.4 years), 63 (64%) patients had VT recurrence (LP/Pv+: 30 of 37 [81%] vs. LP/Pv-: 33 of 61 [54%]; p=0.007). Twenty-eight patients (29%) died (LP/Pv+: 19 of 37 [51%] vs. LP/Pv-: 9 of 61 [15%]; p<0.001). The cumulative 2-year VT-free survival was 41% in the total cohort (LP/Pv+: 16% vs. LP/Pv-: 54%; p=0.001). The presence of LP/Pv (hazard ratio: 1.9; 95% confidence interval: 1.1 to 3.4; p=0.02) and unipolar low-voltage area size/cm2 increase (hazard ratio: 2.5; 95% confidence interval: 1.6 to 4.0; p<0.001) were associated with a decreased 2-year VT-free survival. In patients with DCM-VT, a genetic cause is frequently identified. LP/Pv+ patients have a lower LVEF and more extensive VT substrates, which, in combination with disease progression, may contribute to the poor prognosis. Genetic testing in patients with DCM-VT should therefore be recommended.

Genetic Dissection of Hypertrophic Cardiomyopathy with Myocardial RNA-Seq.

Hypertrophic cardiomyopathy (HCM) is an inherited disorder of the myocardium, and pathogenic mutations in the sarcomere genes myosin heavy chain 7 (MYH7) and myosin-binding protein C (MYBPC3) explain 60%–70% of observed clinical cases. The heterogeneity of phenotypes observed in HCM patients, however, suggests that novel causative genes or genetic modifiers likely exist. Here, we systemically evaluated RNA-seq data from 28 HCM patients and 9 healthy controls with pathogenic variant identification, differential expression analysis, and gene co-expression and protein–protein interaction network analyses. We identified 43 potential pathogenic variants in 19 genes in 24 HCM patients. Genes with more than one variant included the following: MYBPC3, TTN, MYH7, PSEN2, and LDB3. A total of 2538 protein-coding genes, six microRNAs (miRNAs), and 1617 long noncoding RNAs (lncRNAs) were identified differentially expressed between the groups, including several well-characterized cardiomyopathy-related genes (ANKRD1, FHL2, TGFB3, miR-30d, and miR-154). Gene enrichment analysis revealed that those genes are significantly involved in heart development and physiology. Furthermore, we highlighted four subnetworks: mtDNA-subnetwork, DSP-subnetwork, MYH7-subnetwork, and MYBPC3-subnetwork, which could play significant roles in the progression of HCM. Our findings further illustrate that HCM is a complex disease, which results from mutations in multiple protein-coding genes, modulation by non-coding RNAs and perturbations in gene networks.

Expanding the clinical and genetic spectrum of ALPK3 variants: Phenotypes identified in pediatric cardiomyopathy patients and adults with heterozygous variants

Biallelic damaging variants in ALPK3, encoding alpha-protein kinase 3, cause pediatric-onset cardiomyopathy with manifestations that are incompletely defined. We analyzed clinical manifestations of damaging biallelic ALPK3 variants in 19 pediatric patients, including nine previously published cases. Among these, 11 loss-of-function (LoF) variants, seven compound LoF and deleterious missense variants, and one homozygous deleterious missense variant were identified. Among 18 live-born patients, 8 exhibited neonatal dilated cardiomyopathy (44.4%; 95% CI: 21.5%-69.2%) that subsequently transitioned into ventricular hypertrophy. The majority of patients had extracardiac phenotypes, including contractures, scoliosis, cleft palate, and facial dysmorphisms. We observed no association between variant type or location, disease severity, and/or extracardiac manifestations. Myocardial histopathology showed focal cardiomyocyte hypertrophy, subendocardial fibroelastosis in patients under 4 years of age, and myofibrillar disarray in adults. Rare heterozygous ALPK3 variants were also assessed in adult-onset cardiomyopathy patients. Among 1548 Dutch patients referred for initial genetic analyses, we identified 39 individuals with rare heterozygous ALPK3 variants (2.5%; 95% CI: 1.8%-3.4%), including 26 missense and 10 LoF variants. Among 149 U.S. patients without pathogenic variants in 83 cardiomyopathy-related genes, we identified six missense and nine LoF ALPK3 variants (10.1%; 95% CI: 5.7%-16.1%). LoF ALPK3 variants were increased in comparison to matched controls (Dutch cohort, P = 1.6×10-5; U.S. cohort, P = 2.2×10-13). Biallelic damaging ALPK3 variants cause pediatric cardiomyopathy manifested by DCM transitioning to hypertrophy, often with poor contractile function. Additional extracardiac features occur in most patients, including musculoskeletal abnormalities and cleft palate. Heterozygous LoF ALPK3 variants are enriched in adults with cardiomyopathy and may contribute to their cardiomyopathy. Adults with ALPK3 LoF variants therefore warrant evaluations for cardiomyopathy.

Cardiomyopathy associated with the Ala143Thr variant of the α-galactosidase A gene

ObjectiveTo investigate whether the Ala143Thr variant of the α-galactosidase A gene (A143T/GLA), with conflicting interpretations of pathogenicity, is associated with Fabry cardiomyopathy.MethodsThe index patient, a woman in her 60s with...

Vezf1 regulates cardiac structure and contractile function.

BackgroundVascular endothelial zinc finger 1 (Vezf1) is a transcription factor previously shown to regulate vasculogenesis and angiogenesis. We aimed to investigate the role of Vezf1 in the postnatal heart.MethodsThe role of Vezf1 in regulating cardiac growth and contractile function was studied in zebrafish and in primary cardiomyocytes.FindingsWe find that expression of Vezf1 is decreased in diseased human myocardium and mouse hearts. Our experimental data shows that knockdown of zebrafish Vezf1 reduces cardiac growth and results in impaired ventricular contractile response to β-adrenergic stimuli. However, Vezf1 knockdown is not associated with dysregulation of cardiomyocyte Ca2+ transient kinetics. Gene ontology enrichment analysis indicates that Vezf1 regulates cardiac muscle contraction and dilated cardiomyopathy related genes and we identify cardiomyocyte Myh7/β-MHC as key target for Vezf1. We further identify a key role for an MCAT binding site in the Myh7 promoter regulating the response to Vezf1 knockdown and show that TEAD-1 is a binding partner of Vezf1.InterpretationWe demonstrate a role for Vezf1 in regulation of compensatory cardiac growth and cardiomyocyte contractile function, which may be relevant in human cardiac disease.

Genetic Dissection of Hypertrophic Cardiomyopathy with Myocardial RNA-Seq

Background: Hypertrophic cardiomyopathy (HCM) is an inherited disorder of the myocardium, and pathogenic mutations in the sarcomere genes MYH7 and MYBPC3 explain 60-70% of observed clinical cases. The heterogeneity of phenotypes observed in HCM patients, however, suggests that novel causative genes or genetic modifiers likely exist and that further elucidation of these genes is needed. Methods: In order to identify genes and potential networks that affect the HCM phenotype, we systemically evaluated RNA-seq data from 28 HCM patients and 9 healthy controls with pathogenic variant identification, differential expression analysis, and gene co-expression and protein-protein interaction network analyses. Findings: We identified 43 potential pathogenic variants in 19 genes in 24 HCM patients. Genes with more than one variant included the following: MYBPC3, TTN, MYH7, PSEN2, and LDB3. A total of 2538 protein-coding genes, 6 microRNAs (miRNAs), and 1617 long noncoding RNAs (lncRNAs) were identified differentially expressed between the groups, including several well-characterized cardiomyopathy-related genes (ANKRD1, FHL2, MYH6, TGFB3, miR-30d, and miR-154). Gene enrichment analysis revealed that those genes are significantly involved in heart development and physiology. Furthermore, by integrating gene co-expression and protein-protein interaction network analyses, we highlighted four subnetworks: Mitochondrial DNA (mtDNA)-subnetwork, Desmoplakin (DSP) -subnetwork, myosin heavy chain 7 (MYH7) -subnetwork, and cardiac myosin-binding protein C (MYBPC3)-subnetwork, which could play significant roles in the progression of HCM. Interpretation: Our findings further illustrate that HCM is a complex disease, which results from mutations in multiple protein-coding genes, modulation by non-coding RNAs and perturbations in gene networks. Funding Statement: The authors received no specific funding for this work. Declaration of Interests: All other authors declare no competing interests. Ethics Approval Statement: Not required.

P4797Genetic testing in 101 consecutive lone atrial fibrillation patients is able to identify a subgroup with increased severity

Abstract Introduction Atrial fibrillation (AF) is the most common arrhythmia and the cause of 15% of all strokes and up to 6% of medical admissions. It is estimated that currently about 2.0 million Brazilians, and 30 million individuals worldwide, are affected by the disease. It is a complex and multifactorial disease, and the mechanisms are still not well understood. Between 10–20% of AF patients do not have any known predisposing factors, a condition once called “lone AF”. The role of genetic testing still remains controversial in this clinical scenario. Purpose The aim of this study was to identify the occurrence of pathogenic genetic variants in patients with atrial fibrillation without known risk factors. Methods In a tertiary hospital, 101 young patients with apparent “lone AF” were screened with genetic testing by NGS using a custom genetic panel with 159 channelopathy and cardiomyopathy related genes. Variants found were classified according to the American College of Genetic and Genomic (ACMG) criteria. Subjects were evaluated with clinical and familial history, electrocardiogram, 24 hours Holter monitoring, echocardiogram, cardiac MRI and treatment response. Multivariate analyses were performed by logistic linear regression model. Results During an inclusion period of 4 years, 101 consecutive patients, with mean age of 38.6 years old, were classified as “lone AF” (78% male); 76% presented paroxysmal AF and 24% persistent/permanent AF. Family history of early sudden death (bellow 60 years old) was reported in 37% of cases (78% below 50 years old); 10% had family members with pacemakers; and 44% reported having family members with early AF onset. Genetic testing demonstrated that 14/101 (13.8%) of patients presented genetic variants classified as pathogenic or likely pathogenic (P/LP) according to ACMG criteria. The genes most frequently affected were LMNA (3/101), ANK2 (3/101) and truncating variants in TTN (3/101). Two variables were significantly associated with harboring a pathogenic mutation: family history of sudden death (OR: 5.58; 1,19–26,12 CI; p=0.029) and pacemaker reported in the family history (OR: 6.83; 1.11–42.04 CI; p=0.038). Conclusion Our data showed that approximately 15% of “lone AF” patients are carriers of known pathogenic mutations in genes associated with inherithed cardiomyopathy. In addition, we show that being a carrier is potentially associated with a more severe phenotype. These findings suggest that genetic testing in “lone AF” patients may be able to identify a subgroup with a more severe phenotype are for whom a different management strategy might be indicated. Acknowledgement/Funding National Council for Scientific and Technological Development (CNPq)

P3688Familial recurrent autoimmune myocarditis associated with a truncating nonsense mutation of the desmoplakin gene

Abstract Background Arrhythmogenic cardiomyopathy (AC) is an important cause of ventricular arrhythmias in children and young adults. AC is associated with mutation of desmosomal proteins, however, cardiac disease penetrance is incomplete and the clinical course varies widely without recognizable exogenous or epi/genetic co-factors. Importantly, DSP mutation carriers may also display entirely non-cardiac e.g. dermatological phenotypes. Methods and results In two brothers with recurrent fulminant myocarditis, mutation screening of 218 cardiomyopathy-related genes identified a truncating mutation Arg1458* of desmoplakin (DSP). DSP immunhistology unexpectedly revealed complete loss (“knockout”) of DSP protein in endomyocardial biopsies (EMBs), but none of the histological anomalies of AC. Criteria for histological diagnosis of myocarditis were not either fulfilled, and cardiac MRI revealed no features associated with AC. Screening for infections was negative, there was no substance abuse, medication or vaccination. Possible disease triggers were competitive sport events. Myosin and troponin I autoantibodies were detected at titers up to 1:320. We used allele-specific RT-PCR to distinguish if the patients' allele classified as “normal” was actually defective due to promotor mutation or epigenetic silencing. RT-PCRs were done on EMBs and peripheral blood mononuclear cells (PBMCs). In a cohort of dilated cardiomyopathy (DCM) patients we were able to detect DSP transcripts in both, PBMC and left-ventricular heart tissue. RNA sequencing of human PBMC subpopulations suggested that DSP transcription may be restricted to certain immune cell subtypes. RT-PCRs revealed that both Arg1458* carriers have a functional second DSP allele, indicating that their “DSP knockout” occurs at the protein level and may be due to protein instability and degradation within desmosomes. We screened additional existing cohorts for such variants and identified stopgain variant Gln307Ter in a 37-yrs-old woman with ARVC. This patient's sister died from heart failure at the age of 39. In a 59-yrs-old female LVNC patient, stopgain variant Y1391X was identified. Here, family history was unclear, her brother probably died from coronary artery disease. In a 71-yrs-old female DCM patient with no family history, stopgain variant Tyr1512Ter was identified. Conclusions The described patients with DSP truncations strongly suggest the existence of additional genetic or exogenous modifiers driving pathogenesis either way. DSP defects may cause recurrent myocarditis, and mutation screening is advisable to enable early detection of high-risk patients with similar phenotypes. Our finding of complete myocardial DSP protein loss emphasizes that DNA sequencing may miss critical molecular disturbances. It is indispensable to also analyze transcriptome and protein level in the tissue actually affected in a patient in order to recognize his/her individual pathogenesis.

Clinical implications of sarcomere and nonsarcomere gene variants in patients with left ventricular noncompaction cardiomyopathy.

BackgroundRobust data regarding genotype–phenotype correlations in left ventricular noncompaction cardiomyopathy (LVNC) are lacking.MethodsAbout 72 cardiomyopathy‐related genes were comprehensively screened in a cohort of LVNC patients using targeted sequencing. Baseline and follow‐up data were collected. The primary endpoint was a composite of death and heart transplantation.ResultsA total of 83 unrelated adult patients were included in analyses. Following stringent classification according to the American College of Medical Genetics and Genomics (ACMG) guidelines, 36 pathogenic variants of 14 genes were detected in 32 patients. Among them, 12 patients carried at least one nonsarcomere variant (NSV). At baseline, NSV carriers had a higher frequency of atrial fibrillation, but lower left ventricular ejection fraction, than did noncarriers. During a median follow‐up of 4.2 years, NSV carriers experienced a higher rate of the primary endpoint compared with noncarriers. There was no significant difference in the rate between carriers of sarcomere variant (SV) and noncarriers, as well as between carriers of SV and NSV. The presence of NSV was associated with an increased risk of the primary endpoint independent of age, sex, and cardiac function (hazard ratio: 3.61, 95% confidence interval: 1.42–9.19, p = .002).ConclusionNSV may act as a genetic modifier and worsen the clinical phenotype in patients with LVNC.

Genetics of cancer therapy-induced cardiomyopathy.

Patients with chemotherapy-induced cardiomyopathy have a high prevalence of rare variants of cardiomyopathy-related genes, in particular titin-truncating variants.

Digenic Inheritance of LAMA4 and MYH7 Mutations in Patient with Infantile Dilated Cardiomyopathy.

Background and objectives: Dilated cardiomyopathy (DCM) is a rare cardiac disease characterised by left ventricular enlargement, reduced left ventricular contractility, and impaired systolic function. Childhood DCM is clinically and genetically heterogenous and associated with mutations in over 100 genes. The aim of this study was to identify novel variations associated with infantile DCM. Materials and Methods: Targeted next generation sequencing (NGS) of 181 cardiomyopathy-related genes was performed in three unrelated consanguineous families from Saudi Arabia. Variants were confirmed and their frequency established in 50 known DCM cases and 80 clinically annotated healthy controls. Results: The three index cases presented between 7 and 10 months of age with severe DCM. In Family A, there was digenic inheritance of two heterozygous variants: a novel variant in LAMA4 (c.3925G > A, p.Asp1309Asn) and a known DCM mutation in MYH7 (c.2770G > A; p.Glu924Lys). The LAMA4 p.Asp1309Asn variant was predicted to be likely pathogenic according to international guidelines. The other two families had no identifiable potentially deleterious variants. Conclusions: Inheritance of two genetic variants may have a synergistic or dose effect to cause severe DCM. We report of a novel p.Asp1309Asn variation associated with DCM. Targeted NGS is useful in the molecular diagnosis of DCM and to guide whole-family management and counselling.

Whole-exome sequencing reveals doubly novel heterozygous Myosin Binding Protein C and Titin mutations in a Chinese patient with severe dilated cardiomyopathy.

Dilated Cardiomyopathy is a serious heart disorder that may induce sudden cardiac death and heart failure. Significant progress has been made in understanding the molecular basis of dilated cardiomyopathy. In previous studies, mutations in more than fifty genes have been identified in dilated cardiomyopathy patients. The purpose of this study was to detect the genetic lesion in a family from the central south of China affected by severe dilated cardiomyopathy. Whole-exome sequencing combined with cardiomyopathy-related genes list were used to analyse the mutations of the proband. Co-segregation analysis was performed by Sanger sequencing.Results and conclusionsTwo novel heterozygous mutations - Myosin Binding Protein C: p.L1014RfsX6 and Titin: p.R9793X - were identified in the proband. The deletion mutation c.3041delT/p.L1014RfsX6 caused a premature stop codon at position 1020 in exon 28 of the Myosin Binding Protein C. The nonsense mutation, c.29377 C>T/ p. R9793X, of Titin was located in the highly evolutionarily conserved domain, resulting in truncation of the Titin protein as well. Co-segregation analysis further revealed that the Myosin Binding Protein C mutation came from his mother and the Titin mutation came from his father. Both mutations are reported in dilated cardiomyopathy patients for the first time. Our study not only provides a unique example of the genes and molecular mechanisms involved in dilated cardiomyopathy but also expands the spectrum of Myosin Binding Protein C and Titin mutations and contributes to the genetic diagnosis and counselling of dilated cardiomyopathy patients.