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Genetic cardiomyopathies are disorders of the cardiac muscle, most often explained by pathogenic mutations in genes encoding sarcomere, cytoskeleton, or ion channel proteins. Clinical phenotypes such as heart failure and arrhythmia are classically treated with generic drugs, but aetiology-specific and targeted treatments are lacking. As a result, cardiomyopathies still present a major burden to society, and affect many young and older patients. The Translational Committee of the Heart Failure Association (HFA) and the Working Group of Myocardial Function of the European Society of Cardiology (ESC) organized a workshop to discuss recent advances in molecular and physiological studies of various forms of cardiomyopathies. The study of cardiomyopathies has intensified after several new study setups became available, such as induced pluripotent stem cells, three-dimensional printing of cells, use of scaffolds and engineered heart tissue, with convincing human validation studies. Furthermore, our knowledge on the consequences of mutated proteins has deepened, with relevance for cellular homeostasis, protein quality control and toxicity, often specific to particular cardiomyopathies, with precise effects explaining the aberrations. This has opened up new avenues to treat cardiomyopathies, using contemporary techniques from the molecular toolbox, such as gene editing and repair using CRISPR-Cas9 techniques, antisense therapies, novel designer drugs, and RNA therapies. In this article, we discuss the connection between biology and diverse clinical presentation, as well as promising new medications and therapeutic avenues, which may be instrumental to come to precision medicine of genetic cardiomyopathies. [ABSTRACT FROM AUTHOR]

The ubiquitin-proteasome system (UPS) and the autophagy-lysosomal pathway (ALP) are the main proteolytic systems involved in cellular homeostasis. Since cardiomyocytes, as terminally differentiated cells, lack the ability to share damaged proteins with their daughter cells, they are especially reliant on these protein degradation systems for their proper function. Alterations of the UPS and ALP have been reported in a wide range of cardiac diseases, including cardiomyopathies. In this study, we determined whether the UPS and ALP are altered in a mouse model of eccentric left ventricular (LV) hypertrophy expressing both cyclin T1 and Gαq under the control of the cardiac-specific α-myosin heavy chain promoter (double transgenic; DTG). Compared to wild-type (WT) littermates, DTG mice showed higher end-diastolic (ED) LV wall thicknesses and diameter with preserved ejection fraction (EF). The cardiomyopathic phenotype was further confirmed by an upregulation of the fetal gene program and genes associated with fibrosis as well as a downregulation of genes involved in Ca2+ handling. Likewise, higher NT-proBNP levels were detected in DTG mice. Investigation of the UPS showed elevated steady-state levels of (poly)ubiquitinated proteins without alterations of all proteasomal activities in DTG mice. Evaluation of ALP key marker revealed a mixed pattern with higher protein levels of microtubule-associated protein 1 light chain 3 beta (LC3)-I and lysosomal-associated membrane protein-2, lower protein levels of beclin-1 and FYVE and coiled-coil domain-containing protein 1 (FYCO1) and unchanged protein levels of p62/SQSTM1 in DTG mice when compared to WT. At transcriptional level, a > 1.2-fold expression was observed for Erbb2 , Hdac6 , Lamp2 , Nrg1 , and Sqstm1 , while a < 0.8-fold expression was revealed for Fyco1 in DTG mice. The results related to the ALP suggested overall a repression of the ALP during the initiation process, but an induction of the ALP at the level of autophagosome-lysosome fusion and the delivery of ubiquitinated cargo to the ALP for degradation. [ABSTRACT FROM AUTHOR]

The underlying genetic mechanisms and early pathological events of children with primary cardiomyopathy (CMP) are insufficiently characterized. In this study, we aimed to characterize the mutational spectrum of primary CMP in a large cohort of patients ≤18 years referred to a tertiary center. Eighty unrelated index patients with pediatric primary CMP underwent genetic testing with a panel‐based next‐generation sequencing approach of 89 genes. At least one pathogenic or probably pathogenic variant was identified in 30/80 (38%) index patients. In all CMP subgroups, patients carried most frequently variants of interest in sarcomere genes suggesting them as a major contributor in pediatric primary CMP. In MYH7, MYBPC3, and TNNI3, we identified 18 pathogenic/probably pathogenic variants (MYH7 n = 7, MYBPC3 n = 6, TNNI3 n = 5, including one homozygous (TNNI3 c.24+2T>A) truncating variant. Protein and transcript level analysis on heart biopsies from individuals with homozygous mutation of TNNI3 revealed that the TNNI3 protein is absent and associated with upregulation of the fetal isoform TNNI1. The present study further supports the clinical importance of sarcomeric mutation—not only in adult—but also in pediatric primary CMP. TNNI3 is the third most important disease gene in this cohort and complete loss of TNNI3 leads to severe pediatric CMP. [ABSTRACT FROM AUTHOR]

BACKGROUND: Alterations in autophagy have been reported in hypertrophic cardiomyopathy (HCM) caused by Danon disease, Vici syndrome, or LEOPARD syndrome, but not in HCM caused by mutations in genes encoding sarcomeric proteins, which account for most of HCM cases. MYBPC3, encoding cMyBP-C (cardiac myosin-binding protein C), is the most frequently mutated HCM gene. METHODS AND RESULTS: We evaluated autophagy in patients with HCM carrying MYBPC3 mutations and in a Mybpc3-targeted knockin HCM mouse model, as well as the effect of autophagy modulators on the development of cardiomyopathy in knockin mice. Microtubule-associated protein 1 light chain 3 (LC3)-II protein levels were higher in HCM septal myectomies than in nonfailing control hearts and in 60-week-old knockin than in wild-type mouse hearts. In contrast to wild-type, autophagic flux was blunted and associated with accumulation of residual bodies and glycogen in hearts of 60-week-old knockin mice. We found that Akt-mTORC1 (mammalian target of rapamycin complex 1) signaling was increased, and treatment with 2.24 mg/kg.d rapamycin or 40% caloric restriction for 9 weeks partially rescued cardiomyopathy or heart failure and restored autophagic flux in knockin mice. CONCLUSIONS: Altogether, we found that (1) autophagy is altered in patients with HCM carrying MYBPC3 mutations, (2) autophagy is impaired in Mybpc3-targeted knockin mice, and (3) activation of autophagy ameliorated the cardiac disease phenotype in this mouse model. We propose that activation of autophagy might be an attractive option alone or in combination with another therapy to rescue HCM caused by MYBPC3 mutations. [ABSTRACT FROM AUTHOR]

The clinical variability in patients with sarcomeric cardiomyopathies is striking: a mutation causes cardiomyopathy in one individual, while the identical mutation is harmless in a family member. Moreover, the clinical phenotype varies ranging from asymmetric hypertrophy to severe dilatation of the heart. Identification of a single phenotype-associated disease mechanism would facilitate the design of targeted treatments for patient groups with different clinical phenotypes. However, evidence from both the clinic and basic knowledge of functional and structural properties of the sarcomere argues against a 'one size fits all' therapy for treatment of one clinical phenotype. Meticulous clinical and basic studies are needed to unravel the initial and progressive changes initiated by sarcomere mutations to better understand why mutations in the same gene can lead to such opposing phenotypes. Ultimately, we need to design an 'integrative physiology' approach to fully realize patient/gene-tailored therapy. Expertise within different research fields (cardiology, genetics, cellular biology, physiology, and pharmacology) must be joined to link longitudinal clinical studies with mechanistic insights obtained from molecular and functional studies in novel cardiac muscle systems. New animal models, which reflect both initial and more advanced stages of sarcomeric cardiomyopathy, will also aid in achieving these goals. Here, we discuss current priorities in clinical and preclinical investigation aimed at increasing our understanding of pathophysiological mechanisms leading from mutation to disease. Such information will provide the basis to improve risk stratification and to develop therapies to prevent/rescue cardiac dysfunction and remodelling caused by sarcomere mutations. [ABSTRACT FROM AUTHOR]

Over the pastdecade, our understanding of cardiomyopathies has improved dramatically, due to improvements inscreening and detection of gene defects in the human genome as well as a variety of novel animal models (mouse, zebra fish, and drosophila) and in silico computational models. These novel experimental tools have created a platform that is highly complementary to the naturally occurring cardiomyopathies in cats and dogs that had been available for some time. A fully integrative approach, which incorporates all these modalities, is likely required for significant steps forward in understanding the molecular underpinnings and pathogenesis of cardiomyopathies. Finally, novel technologies, including CRISPR/Cas9, which have already been proved to work in zebra fish, are currently being employed to engineer sarcomeric cardiomyopathy in larger animals, including pigs and non-humanprimates. In the mouse, the increased speed with which these techniques can be employed to engineer precise 'knock-in' models that previously took years to make viamultiple rounds of homologous recombination-based gene targeting promises multiple and precise models of human cardiac disease for future study. Such novel genetically engineered animal models recapitulating human sarcomeric protein defects will help bridging the gap to translate therapeutic targets from small animal and in silico models to the human patient with sarcomeric cardiomyopathy. [ABSTRACT FROM AUTHOR]

A growing line of evidence indicates a dysfunctional ubiquitin-proteasome system (UPS) in cardiac diseases. Anti-hypertrophic effects and improved cardiac function have been reported after treatment with proteasome inhibitors in experimental models of cardiac hypertrophy. Here we tested whether proteasome inhibition could also reverse the disease phenotype in a genetically-modified mouse model of hypertrophic cardiomyopathy (HCM), which carries a mutation in Mybpc3, encoding the myofilament protein cardiac myosin-binding protein C. At 7 weeks of age, homozygous mutant mice (KI) have 39% higher left ventricular mass-to-body-weight ratio and 29% lower fractional area shortening (FAS) than wild-type (WT) mice. Both groups were treated with epoxomicin (0.5 mg/kg/day) or vehicle for 1 week via osmotic minipumps. Epoxomicin inhibited the chymotrypsinlike activity by ∼50% in both groups. All parameters of cardiac hypertrophy (including the fetal gene program) were not affected by epoxomicin treatment in both groups. In contrast, FAS was 12% and 35% higher in epoxomicin-treated than vehicle-treated WT and KI mice, respectively. To identify which genes or pathways could be involved in this positive effect, we performed a transcriptome analysis in KI and WT neonatal cardiac myocytes, treated or not with the proteasome inhibitor MG132 (1μM, 24 h). This revealed 103 genes (four-fold difference; 5% FDR) which are commonly regulated in both KI and WT cardiac myocytes. Thus, even in genetically-modified mice with manifest HCM, proteasome inhibition showed beneficial effects, at least with regard to cardiac function. Targeting the UPS in cardiac diseases remains therefore a therapeutic option. [ABSTRACT FROM AUTHOR]

Aims The transcription factor Islet-1 (ISL1) is a marker of cardiovascular progenitors and is essential for mammalian cardiogenesis. An ISL1 haplotype has recently been associated with congenital heart disease. In this study we evaluated whether ISL1 variants are associated with hypertrophic (HCM), dilated (DCM), arrhythmogenic right ventricular cardiomyopathy (ARVC), or with Emery–Dreifuss muscular dystrophy (EDMD). Methods and results The six exon and intron boundaries of ISL1 were screened for genetic variants in a cohort of 454 index cases. Eleven exonic variants were identified in HCM, DCM, ARVC, and/or EDMD. Out of the five novel variants, two are located in the 5'-untranslated region, two are silent (p.Arg171Arg and p.Asn189Asn), and one is a missense (p.Asn252Ser). The latter was identified in the homozygous state in one DCM patient, and in the heterozygous state in 11 relatives, who did not present with DCM but often with cardiovascular features. This variant was found in one HCM patient also carrying a MYH7 mutation and in 3/96 North-African Caucasian control individuals, but was absent in 138 European Caucasian control individuals. We investigated the effect of the ISL1 wild type and p.Asn252Ser mutant on myocyte enhancer factor 2C (Mef2c) promoter activity, an established ISL1 target. Mef2c promoter activity was ∼4-fold higher in the presence of wild-type and ∼6-fold higher in the presence of mutant ISL1 in both HEK and CHO cells. Conclusion This study describes a new gain-of-function p.Asn252Ser variant in the human ISL1 gene, which could potentially lead to greater activation of downstream targets involved in cardiac development, dilation, and hypertrophy. [ABSTRACT FROM AUTHOR]

The article focuses on a study which examined if hypertrophic cardiomyopathy (HCM) changes in sarcomeric properties depend on sarcomeric protein mutations. It states that HCM is characterized by asymmetrical left ventricular (LV) hypertrophy involving interventricular septum occurring in the absence of other cardiac disease like hypertension. The results showed that hypocontractile sarcomeres are a deficit in human HCM with normal systolic LV function thereby contributing to HCM progression.

Cardiomyopathies represent an important cause of cardiovascular morbidity and mortality due to heart failure, arrhythmias, and sudden death. Most forms of hypertrophic cardiomyopathy (HCM) are familial with an autosomal-dominant mode of inheritance. Over the last 20 years, the genetic basis of the disease has been largely unravelled. HCM is considered as a sarcomeropathy involving mutations in sarcomeric proteins, most often β-myosin heavy chain and cardiac myosin-binding protein C. ‘Missense’ mutations, more common in the former, are associated with dysfunctional proteins stably integrated into the sarcomere. ‘Nonsense’ and frameshift mutations, more common in the latter, are associated with low mRNA and protein levels derived from the diseased allele, leading to haploinsufficiency of the remaining healthy allele. The two quality control systems responsible for the removal of the affected mRNAs and proteins are the nonsense-mediated mRNA decay (NMD) and the ubiquitin-proteasome system (UPS), respectively. This review discusses clinical and genetic aspects of HCM and the role of NMD and UPS in the regulation of mutant proteins, evidence for impairment of UPS as a pathogenic factor, as well as potential therapies for HCM. [ABSTRACT FROM PUBLISHER]

The article presents a study that aims to investigate the control mechanism of the expression of myosin-binding protein C3 (MyBPC3) mutations. A cardiac myosin-binding protein-C (cMyBP-C) knock-in mouse model was developed to analyze the molecular mechanisms in hypertrophic cardiomyopathy (HCM). Results show that ubiquitin-proteasome and nonsense-mediated messenger RNA regulates cMyBP-C mutation in cardiomyopathic mouse.

Objective: Cardiac myosin-binding protein C (cMyBP-C) gene mutations are involved in familial hypertrophic cardiomyopathy (FHC). Many of these mutations produce truncated proteins, which are unstable in the cardiac tissue of patients, suggesting that haploinsufficiency could account for the development of the phenotype. However, existing mouse models of cMyBP-C gene mutations have represented hypomorphic alleles without evidence of asymmetric septal hypertrophy, a key FHC phenotypic feature. In the present study, we generated a new model of cMyBP-C null mice and characterized the phenotype in both homozygotes and heterozygotes at different ages. Methods: The mouse model was based upon the targeted deletion of exons 1 and 2, which contain the transcription initiation site, and the phenotype was determined by molecular, functional and morphological analyses. Results: Herein, we demonstrate that inactivation of one or two mouse cMyBP-C alleles leads to different cardiac disorders at different post-natal time windows. The homozygous cMyBP-C null mice do not express the cMyBP-C gene, develop eccentric left ventricular hypertrophy with decreased fractional shortening at 3–4 months of age and a markedly impaired relaxation after 9 months. This is associated with myocardial disarray and an increase of interstitial fibrosis. The heterozygous cMyBP-C null mice present a slight but significant decrease of cMyBP-C amount and develop asymmetric septal hypertrophy associated with fibrosis at 10–11 months of age. Conclusion: These data provide evidence that heterozygous cMyBP-C null mice represent the first model with a key feature of human FHC that is asymmetric septal hypertrophy. [Copyright &y& Elsevier]

Objective: Cardiac myosin-binding protein C (cMyBP-C) is a component of sarcomere that contains at least three putative myosin-binding sites. Mutations in its gene are implicated in familial hypertrophic cardiomyopathy (FHC) and most of them are predicted to produce C-terminal truncated cMyBP-Cs. The aim of the present study was to analyze whether cMyBP-C truncated mutants resulting from FHC mutations interact in vitro with human β-MyHC. Methods: Recombinant proteins were produced using the baculovirus/insect cell system, and wild type and three truncated cMyBP-Cs were purified using metal affinity chromatography. The interaction between recombinant proteins was analyzed in real time using biosensor technology on immobilized anti-β-MyHC antibodies. Results: Biomolecular interaction with β-MyHC was detected for both wild type cMyBP-C and a truncated mutant lacking half of the C-terminal C10 domain. In contrast, no interaction with β-MyHC was found for two truncated cMyBP-Cs lacking at least the C5–C9 region. Conclusions: Biosensor technology allows in vitro analysis of the interaction between human β-MyHC and cMyBP-C mutants resulting from FHC mutations. The data show that the interaction depends on the size of the truncation. This suggests that, in the context of FHC, impairment of suitable interaction between β-MyHC and some of the truncated cMyBP-Cs may promote degradation of the truncated proteins and therefore contribute to the development of the disease. [Copyright &y& Elsevier]

Abstract: Cardiac myosin-binding protein C (cMyBP-C) is a component of the thick filaments of the sarcomere. Understanding the structural and functional role of cMyBP-C in the heart is clinically relevant since cMyBP-C gene mutations are a widely recognized cause of hypertrophic cardiomyopathy (HCM), which affects 0.2% of the general population. Nonsense and frameshift mutations are common in cMyBP-C and their expressions are regulated by three quality control systems, the nonsense-mediated mRNA decay, ubiquitin–proteasome system, and autophagy, which contribute to minimize the production of potential poison mutant proteins. This review discusses the structural and regulatory functions of cMyBP-C, the molecular mechanisms involved in cMyBP-C-related HCM, as well as potential causative therapies for HCM. [Copyright &y& Elsevier]

Abstract: Since proteins play crucial roles in all biological processes, the finely tuned equilibrium between their synthesis and degradation regulates cellular homeostasis. Controlling the quality of proteome informational content is essential for cell survival and function. After initial synthesis, membrane and secretory proteins are modified, folded, and assembled in the endoplasmic reticulum, whereas other proteins are synthesized and processed in the cytosol. Cells have different protein quality control systems, the molecular chaperones, which help protein folding and stabilization, and the ubiquitin–proteasome system (UPS) and lysosomes, which degrade proteins. It has generally been assumed that UPS and lysosomes are regulated independently and serve distinct functions. The UPS degrades both cytosolic, nuclear proteins, and myofibrillar proteins, whereas the lysosomes degrade most membrane and extracellular proteins by endocytosis as well as cytosolic proteins and organelles via autophagy. Over the last two decades, the UPS has been increasingly recognized as a major system in several biological processes including cell proliferation, adaptation to stress and cell death. More recently, activation or impairment of the UPS has been reported in cardiac disease and recent evidence indicate that autophagy is a key mechanism to maintain cardiac structure and function. This review mainly focuses on the UPS and its various components in healthy and diseased heart, but also summarizes recent data suggesting parallel activation of the UPS and autophagy in cardiac disease. [Copyright &y& Elsevier]

Autophagy (greek auto: self; phagein: eating) is a highly conserved process within eukaryotes that degrades long-lived proteins and organelles within lysosomes. Its accurate and constant operation in basal conditions ensures cellular homeostasis by degrading damaged cellular components and thereby acting not only as a quality control but as well as an energy supplier. An increasing body of evidence indicates a major role of autophagy in the regulation of cardiac homeostasis and function. In this review, we describe the different forms of mammalian autophagy, their regulations and monitoring with a specific emphasis on the heart. Furthermore, we address the role of autophagy in several forms of cardiomyopathy and the options for therapy. • Autophagy is a highly conserved process that degrades proteins and organelles within lysosomes. • Autophagy plays a major in the regulation of cardiac homeostasis and function. • Different forms, regulation and monitoring of cardiac autophagy are described. • Role of autophagy in several forms of cardiomyopathy and the options for therapy is discussed. [ABSTRACT FROM AUTHOR]

Introduction: Cardiomyopathy (CMP) is a heterogeneous disease group affecting heart function and may lead to heart failure. Genetic variants in more than 50 genes have been linked to CMP phenotypes. The understanding of the natural course and the underlying molecular mechanisms of pediatric CMP and heart failure is incomplete. Hypothesis: This study aims to establish the abundance of genetic variants in known CMP disease genes in a large cohort of pediatric CMP patients. Methods: To identify genetic defects in pediatric CMP we investigated a cohort of patients