Hypertrophic Cardiomyopathy Mutations Research Articles

Cardiac MRI traits linked to cardiomyopathy mutations in phenotype naive carriers

Abstract Hypertrophic cardiomyopathy (HCM) and dilated cardiomyopathy (DCM) are prevalent forms of cardiomyopathies, characterised by changes in cardiac structure and function, associated with an increased risk of atrial fibrillation (AF), heart failure (HF), and sudden cardiac death (SCD). Due to these potential life-threatening consequences close monitoring of genetic mutation carriers of HCM and DCM, is important. However, disease onset in these subjects is highly variable, with a substantial subset of people only developing disease late-in-life, or not at all, making risk stratification challenging. We set out to identify cardiac MRI (CMR) based imaging biomarkers of HCM and DCM carriership in phenotype naïve individuals, which were benchmarked against associations with AF and HF. CMR and whole genome sequencing (n: 42,723) data were sourced from a UK biobank sample free of heart failure at the time of CMR measurements. A previously validated deep learning(DL) network was used to derive 22 CMR traits(1). Adjusted cox regression models were used to determine the CMR associations with the onset of heart failure and atrial fibrillation, providing empirical validation of the DL CMR network. Univariable and multivariable generalised linear models were employed to identify associations with genetic carriership conditional on cardiac risk factors. 14 and 19 CMR measurements were associated with the time until HF (274 cases), AF (704 cases), respectively. These variables included left-ventricle (LV) ejection fraction (EF), LV end systolic volume (ESV), Right-ventricular (RV) EF and right atrial (RA) EF. Using WGS data we identified 154 people with an HCM mutation and 220 with a DCM mutation, where HCM carriership associated with five and DCM with four CMR traits. This included RA-EF (OR 1.26 95%CI 1.10; 1.45), and RV-EF (OR 1.41 95%CI 1.23;1,63) for HCM, and LV-EF (OR 0.72 95%CI 0.62; 0.84), and RV stroke volume (OR 0.75 95CI 0.62; 0.91) for DCM. By combining deep learning of CMR with large sample size WGS we were able to identify cardiac imaging biomarkers of HCM and DCM carriership in apparently healthy subjects. This could prove relevant for early identification and monitoring of phenotype naïve people.

Structural dynamics of the intrinsically disordered linker region of cardiac troponin T.

Troponin plays a central role in regulating heart contraction, and mutations in troponin can cause human cardiomyopathies. There are several functionally-significant regions of troponin that have not been structurally resolved, including the troponin T linker region that contains multiple cardiomyopathy mutations. In these unresolved regions, it is not possible to understand how changes in sequence affect function. We used computational and experimental techniques to demonstrate that this linker is dynamic and intrinsically disordered both in isolation and in the fully regulated thin filament. Moreover, we show how a cardiomyopathy mutation in this region affects function via allosteric disruption of intermolecular interactions. Our results highlight the need to consider how key mutations affect troponin disorder rather than the structure-function relationship.

Arg92Leu-cTnT Alters the cTnC-cTnI Interface Disrupting PKA-Mediated Relaxation.

Impaired left ventricular relaxation, high filling pressures, and dysregulation of Ca2+ homeostasis are common findings contributing to diastolic dysfunction in hypertrophic cardiomyopathy (HCM). Studies have shown that impaired relaxation is an early observation in the sarcomere-gene-positive preclinical HCM cohort, which suggests the potential involvement of myofilament regulators in relaxation. A molecular-level understanding of mechanism(s) at the level of the myofilament is lacking. We hypothesized that mutation-specific, allosterically mediated, changes to the cTnC (cardiac troponin C)-cTnI (cardiac troponin I) interface can account for the development of early-onset diastolic dysfunction via decreased PKA accessibility to cTnI. HCM mutations R92L-cTnT (cardiac troponin T; Arg92Leu) and Δ160E-cTnT (Glu160 deletion) were studied in vivo, in vitro, and in silico via 2-dimensional echocardiography, Western blotting, ex vivo hemodynamics, stopped-flow kinetics, time-resolved fluorescence resonance energy transfer, and molecular dynamics simulations. The HCM-causative mutations R92L-cTnT and Δ160E-cTnT result in different time-of-onset diastolic dysfunction. R92L-cTnT demonstrated early-onset diastolic dysfunction accompanied by a localized decrease in phosphorylation of cTnI. Constitutive phosphorylation of cTnI (cTnI-D23D24) was sufficient to recover diastolic function to non-Tg levels only for R92L-cTnT. Mutation-specific changes in Ca2+ dissociation rates associated with R92L-cTnT reconstituted with cTnI-D23D24 led us to investigate potential involvement of structural changes in the cTnC-cTnI interface as an explanation for these observations. We probed the interface via time-resolved fluorescence resonance energy transfer revealing a repositioning of the N-terminus of cTnI, closer to cTnC, and concomitant decreases in distance distributions at sites flanking the PKA consensus sequence. Implementing time-resolved fluorescence resonance energy transfer distances as constraints into our atomistic model identified additional electrostatic interactions at the consensus sequence. These data show that the early diastolic dysfunction observed in a subset of HCM is attributable to allosterically mediated structural changes at the cTnC-cTnI interface that impair accessibility of PKA, thereby blunting β-adrenergic responsiveness and identifying a potential molecular target for therapeutic intervention.

Abstract We133: In Silico Model of Cardiomyopathy Caused by a MYBPC3-induced HCM Mutation

Introduction: Hypertrophic cardiomyopathy (HCM) is the most frequent hereditary cardiomyopathy and a leading cause of sudden cardiac death, particularly in adolescent. Despite recent improved prognosis and FDA approved drugs, current cardiac therapeutics cannot efficiently prevent remodeling across all HCM patient populations due to genetic variability. Therefore, we require a systems biology approach to understand how the complex cardiac signaling networks regulate cardiomyocyte shape and size due to an HCM mutation. This study leverages a computational systems approach to explore MYBPC3-induced HCM mutation regulation of cardiomyocyte hypertrophy. Method: For the signaling network model, we employed a logic-based differential equations model (NETFLUX) to construct a signaling network validated using in vitro data from published studies. This model captured signaling dynamics and cross talks between pathways governing cardiomyocyte size and shape. Additionally, a MYBPC3 HCM mutation is integrated in the model as an input providing a comparison between a control and simulated HCM responses. Sensitivity analyses are performed to investigate the most influential signaling pathways that mediate changes in mass. Results: We created a MYBPC3-induced HCM model by integrating an additional cardiac myosin binding protein-C node to a previously validated inherited cardiomyopathy signaling network. This updated model predicted an increase in mass when the HCM mutation is simulated. The simulated inhibition of each node in the model revealed that inhibition of calcium pathway, growth factor receptor, and titin stiffness could mitigate the HCM phenotype by reducing mass. The sensitivity analysis results also suggested PI3K, Ras, and ERK1/2 pathways as potential mechanisms mediating the decrease in mass and served as potential therapeutic targets. Discussion and Future Work: Our systems approach gave insights on the mechanisms governing the cardiomyocyte response to a MYBPC3-induced HCM mutation. This will be further validated via a MYBPC3-HCM model using human iPSC-CMs with MYBPC3-induced mutations. This will create a cell line that will provide genetically precise comparisons for identifying new molecular mechanisms of disease, thereby offering a strategy for advancing precision medicine.

Abstract Mo080: Clinically Variable Penetrant MYH7 G256E Mutation Shows Gene-Dose-Dependent HCM Disease Phenotype on a Transcriptomic, Proteomic, and Functional Level

Introduction: Hypertrophic cardiomyopathy (HCM) is a monogenic disorder caused by heterozygous (HET) mutations in sarcomere genes such as MYH7 . Recently, human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) carrying HET HCM mutations have shed new insights into the molecular mechanisms of HCM. However, the cause of phenotype variability and the role of allelic imbalance in phenotype penetrance of HET missense mutations such as MYH7 G256E remain unclear. Hypothesis: We hypothesize that hiPSC CMs carrying clinically variable penetrant HCM missense mutations such as MYH7 G256E show a gene dose-dependent disease phenotype indicating their susceptibility for allelic imbalance as a cause for variable disease phenotype penetrance. Methods: We generated HET and homozygous (HOM) hiPSC-lines for variable penetrant HCM mutation MYH7 G256E. We performed multiplexed single-cell RNA sequencing and quantitative proteomics analyses on wildtype (WT), HET, and HOM iPSC-CMs at day 30 of cardiac differentiation. Additional Digital Droplet PCR studies were performed to investigate the MYH7 mutant allele expression. The functional contractile phenotype was assessed with Sony SI8000 Cell motion imaging system. Results: We detected a large clone-to-clone and batch-to-batch variability in HET iPSC-CMs on a transcriptomic and proteomic level that obscured the identification of a consistent mutant phenotype in HET MYH7 G256E vs WT CMs. These phenotypes correlated with a 1:1 bulk ratio of mutant vs WT mRNA and protein expression, while overall MYH7 gene and protein expression remained comparable. Transcriptomic and proteomic analyses of WT, HET, and HOM CMs show a gene dose-dependent regulation of known hypertrophy response markers such as NPPB (mRNA: HET vs WT: +1.76x p .adj=7.0*10 -7 , HOM vs WT: +5.42x p.adj=1.5*10 -48 ; protein: HET vs WT: +1.15x, 95% CI [0.86, 1.58]; HOM vs WT: +1.51x, 95% CI [1.12, 2.01]) which functionally correlated with enhanced contractility (contraction velocity in μm/s: WT: 5.84 +/-2.24, HET: 6.04 +/-2.89, HOM: 7.77 +/-3.352, p=0.3*10 -2 ANOVA) and the more frequent occurrence of arrhythmias (WT: 1/46, HET: 0/55, HOM: 5/41). Conclusions: Hypertrophic disease phenotype in MYH7 G256E mutation iPSC-CMs shows a gene dose-dependent penetrance on a transcriptomic, proteomic, and functional level. This indicates susceptibility to allelic imbalance as a potential cause for variable disease penetrance.

Abstract We051: Chronic mavacamten treatment prevents development of contractile dysfunction in cMyBP-C null engineered heart tissue constructs, but not in constructs carrying the cMyBP-C W792R missense mutation

Hypercontractility, caused by mutations that increase the number of myosin heads in the disordered relaxed (DRX) state, is a prevalent cause of hypertrophic cardiomyopathy (HCM). A subgroup of HCM mutations cause primary hypocontractility, by stabilizing super-relaxed crossbridges (SRX). Mavacamten (mava) directly targets hypercontractility by reducing the number of DRX heads and improves outcomes in ~70% patients with obstructive HCM. We here assessed the efficacy of chronic mava treatment at preventing phenotype development in human iPSC engineered heart tissue (EHT) in a cMyBP-C null model (-/-), that increases the number of DRX heads, and EHT carrying the W792R (RR) HCM mutation. While the primary contractile defect caused by the W792R mutation is unknown, we postulated that it differed from the cMyBP-C -/- model, as RR mice die prior to weaning while -/- mice do not. Twitch force (TF) measurements performed on day 7 on control (+/+), -/- and RR EHT show that -/- EHT were hypercontractile while RR EHT were hypocontractile (Fig. 1). Following measurement, EHT from each genotype were either treated with vehicle or 100nM mava and cultured for 21 days. By day 28, TF production was similar between +/+ and -/- vehicle treated EHT while RR EHT remained hypocontractile (Fig. 2). Chronic mava treatment significantly reduced TF production in +/+ EHT but not in -/- or RR EHT (Fig. 2). Consistent with diastolic dysfunction, a hallmark of HCM in humans, day 28 -/- and RR EHT displayed slow relaxation, though the effect was more pronounced in RR EHT (Fig. 3). While chronic mava treatment accelerated relaxation in -/- EHT to levels that were similar to that observed in vehicle treated +/+ EHT, it failed to do so in RR EHT (Fig. 3). cMyBP-C -/- EHTs initially present with a hypercontractile phenotype, consistent with an increase in DRX crossbridges, while W792R RR EHT present with a hypocontractile phenotype. By 4 weeks in EHT culture, both genotypes present with impaired relaxation, that is more severe in RR EHT. While chronic mava treatment prevents development of slow relaxation in -/- EHT, it failed to do so in RR EHT. These findings suggest that the primary disease mechanism of the W792R mutation is not an increase in DRX crossbridges and that mavacamten may not be an effective treatment for patients carrying this mutation, further highlighting the need for studying the pathomechansims of HCM mutations and the development of novel therapies.

Echocardiographic Strain Abnormalities Precede Left Ventricular Hypertrophy Development in Hypertrophic Cardiomyopathy Mutation Carriers.

Hypertrophic cardiomyopathy (HCM) is a genetic disease characterized by unexplained left ventricular hypertrophy (LVH), diastolic dysfunction, and increased sudden-death risk. Early detection of the phenotypic expression of the disease in genetic carriers without LVH (Gen+/Phen-) is crucial for emerging therapies. This clinical study aims to identify echocardiographic predictors of phenotypic development in Gen+/Phen-. Sixteen Gen+/Phen- (one subject with troponin T, six with myosin heavy chain-7, and nine with myosin-binding protein C3 mutations), represented the study population. At first and last visit we performed comprehensive 2D speckle-tracking strain echocardiography. During a follow-up of 8 ± 5 years, five carriers developed LVH (LVH+). At baseline, these patients were older than those who did not develop LVH (LVH-) (30 ± 8 vs. 15 ± 8 years, p = 0.005). LVH+ had reduced peak global strain rate during the isovolumic relaxation period (SRIVR) (0.28 ± 0.05 vs. 0.40 ± 0.11 1/s, p = 0.048) and lower global longitudinal strain (GLS) (-19.8 ± 0.4 vs. -22.3 ± 1.1%; p < 0.0001) than LVH- at baseline. SRIVR and GLS were not correlated with age (overall, p > 0.08). This is the first HCM study investigating subjects before they manifest clinically significant or relevant disease burden or symptomatology, comparing at baseline HCM Gen+/Phen- subjects who will develop LVH with those who will not. Furthermore, we identified highly sensitive, easily obtainable, age- and load-independent echocardiographic predictors of phenotype development in HCM gene carriers who may undergo early preventive treatment.

Human cardiac β-myosin powerstroke energetics: Thin filament, Pi displacement, and mutation effects

The powerstroke of human cardiac β-myosin is an important stage of the cross-bridge cycle that generates force for muscle contraction. However, the starting structure of this process has never been resolved, and the relative timing of the powerstroke and inorganic phosphate (Pi) release is still controversial. In this study, we generated an atomistic model of myosin on the thin filament and utilized metadynamics simulations to predict the absent starting structure of the powerstroke. We demonstrated that the displacement of Pi from the active site during the powerstroke is likely necessary, reducing the energy barrier of the conformation change. The effects of the presence of the thin filament, the hypertrophic cardiomyopathy mutation R712L, and the binding of mavacamten on the powerstroke process were also investigated.

Clinical exome sequencing unravels the diverse spectrum of genetic heterogeneity and genotype-phenotype correlations in hypertrophic cardiomyopathy.

BackgroundCatalogues of pathogenic genetic mutations in hypertrophic cardiomyopathy (HCM) are disproportionately small when compared to that of the size of the population with South Asian ancestry and their collective increased risk of heart disease. MethodsWe conducted clinical exome sequencing of 200 HCM patients to identified cardiomyopathy-associated genetic mutations. The clinical and echocardiographic characteristics of genotype-positive and genotype-negative patients were compared, and the likelihood of detecting a positive genetic test result was evaluated. Allelic burden analysis was done to compare the minor allele frequencies (MAF) of the pathogenic or likely pathogenic (P/LP) variants and variants of uncertain significance (VUSs) identified in the cohort against various population genomics databases. ResultsThe genetic yield was 40% for P/LP variants, with MYBPC3 and MYH7 as the predominant sarcomere genes. Younger age-at-diagnosis, family history of HCM, asymmetric hypertrophic (ASH) pattern, the ratio of the interventricular septum to posterior wall thickness (IVS/PW ratio), left atrial (LA) dimensions, severe mitral regurgitation grade (MR grade), late gadolinium enhancement (LGE) detected fibrosis and absence of hypertension were associated with an increased likelihood of HCM-associated variants. Patients who experienced ventricular tachycardia and premature cardiovascular death were significantly likely to carry MYBPC3 or loss-of-function variants. LA and interventricular septal (IVS) dimensions were associated with MYH7 variants. The rare variant burden for P/LP variants and VUSs was significantly enriched in HCM cases compared to population controls. ConclusionOur study provides a comprehensive evaluation of HCM-associated genetic mutations from an Indian population. The identified genotype-phenotype associations could improve the yield of targeted genetic testing in HCM.

Antisense oligonucleotides as a potential therapeutic strategy in hypertrophic cardiomyopathy caused by MYBPC3 variants

Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): Sociedad Española de Cardiología Background/Introduction Non-sense or frameshift mutations in the MYBPC3 gene encoding cardiac myosin-binding protein C (cMyBP-C) are a major cause for inherited forms of Hypertrophic Cardiomyopathy (HCM), the most frequent inherited cardiac disease, affecting 0.2% of the general population and represents the leading cause for sudden cardiac death in young athletes. During the last decade, RNA-based therapeutic approaches have been studied for neuromuscular disorders and recently became to be considered to envision causal therapies for cardiomyopathies, utilizing antisense oligonucleotide mediated exon skipping, which attempts to reframe mutated mybpc3 transcripts, resulting in shortened but functional protein. MYBPC3 gene has a total of 34 coding exons and, exons 2, 22, and 24-27 are ideal targets for skipping. Purpose We aim to design and evaluate antisense oligonucleotides (AONs) for efficient exon-skipping in targeted MYBPC3 exons. Methods We designed two specific AONs to mask the MYBPC3 exon 26 and confirmed successfully mediated exon skipping without disrupting the MYBPC3 reading frame. We chose a previously identified MYBPC3 c.2671insG/p.R891Afs*160 variant in exon 26, which causes a frameshift mutation leading to truncated MYBPC3 in HCM. Primary Human Cardiac Myocytes (PHCM) (Promocell) were transfected with the splice-site targeting AONs. Results In this study we designed two chemically modified AONs (AON1 and AON2) through the addition of a 2′-O-methyl RNA phosphorothioate backbone, targeting exonic splicing enhancers (ESEs) or splice sites (SSs) can induce skipping of MYBPC3 exon 26. We evaluated AON1 (i26-3SS) and AON2 (ESE)-treated Primary Human Cardiac Myocytes (PHCM) show 26-27 exons skipping in comparison to control samples. The identity of the exon 26-27 skipped bands detected by RT-PCR was confirmed by Sanger sequencing and preservation of both nucleotide sequences of MYBPC3 exons 25 and 28 were observed. These results indicate that AONs specifically for mutated MYBPC3 target exons, expanding the framework of future advancements in the therapeutic potential of antisense-mediated exon skipping MYBPC3-based HCM. In addition, skipping exons 26 and 27 we eliminate a total of 22 possible exonic mutations, which represent 7.2% of the total known MYBPC3 exonic mutations. Conclusion(s) These results demonstrate that disruption of the MYBPC3 reading frame due to a truncating HCM mutation can be restored by exon skipping in cardiomyocytes in vitro indicating RNA-based strategies as a potential treatment option for HCM.Exon skipping scheme

Modeling cardiomyocyte signaling and metabolism predicts genotype-to-phenotype mechanisms in hypertrophic cardiomyopathy

Familial hypertrophic cardiomyopathy (HCM) is a significant precursor of heart failure and sudden cardiac death, primarily caused by mutations in sarcomeric and structural proteins. Despite the extensive research on the HCM genotype, the complex and context-specific nature of many signaling and metabolic pathways linking the HCM genotype to phenotype has hindered therapeutic advancements for patients. Here, we have developed a computational model of HCM encompassing cardiomyocyte signaling and metabolic networks and their associated interactions. Utilizing a stochastic logic-based ODE approach, we linked cardiomyocyte signaling to the metabolic network through a gene regulatory network and post-translational modifications. We validated the model against published data on activities of signaling species in the HCM context and transcriptomes of two HCM mouse models (i.e., R403Q-αMyHC and R92W-TnT). Our model predicts that HCM mutation induces changes in metabolic functions such as ATP synthase deficiency and a transition from fatty acids to carbohydrate metabolism. The model indicated major shifts in glutamine-related metabolism and increased apoptosis after HCM-induced ATP synthase deficiency. We predicted that the transcription factors STAT, SRF, GATA4, TP53, and FoxO are the key regulators of cardiomyocyte hypertrophy and apoptosis in HCM in alignment with experiments. Moreover, we identified shared (e.g., activation of PGC1α by AMPK, and FHL1 by titin) and context-specific mechanisms (e.g., regulation of Ca2+ sensitivity by titin in HCM patients) that may control genotype-to-phenotype transition in HCM across different species or mutations. We also predicted potential combination drug targets for HCM (e.g., mavacamten plus ROS inhibitors) preventing or reversing HCM phenotype (i.e., hypertrophic growth, apoptosis, and metabolic remodeling) in cardiomyocytes. This study provides new insights into mechanisms linking genotype to phenotype in familial hypertrophic cardiomyopathy and offers a framework for assessing new treatments and exploring variations in HCM experimental models.

MYBPC3-c.772G>A mutation results in haploinsufficiency and altered myosin cycling kinetics in a patient induced stem cell derived cardiomyocyte model of hypertrophic cardiomyopathy

Approximately 40% of hypertrophic cardiomyopathy (HCM) mutations are linked to the sarcomere protein cardiac myosin binding protein-C (cMyBP-C). These mutations are either classified as missense mutations or truncation mutations. One mutation whose nature has been inconsistently reported in the literature is the MYBPC3-c.772G > A mutation. Using patient-derived human induced pluripotent stem cells differentiated to cardiomyocytes (hiPSC-CMs), we have performed a mechanistic study of the structure-function relationship for this MYBPC3-c.772G > A mutation versus a mutation corrected, isogenic cell line. Our results confirm that this mutation leads to exon skipping and mRNA truncation that ultimately suggests ∼20% less cMyBP-C protein (i.e., haploinsufficiency). This, in turn, results in increased myosin recruitment and accelerated myofibril cycling kinetics. Our mechanistic studies suggest that faster ADP release from myosin is a primary cause of accelerated myofibril cross-bridge cycling due to this mutation. Additionally, the reduction in force generating heads expected from faster ADP release during isometric contractions is outweighed by a cMyBP-C phosphorylation mediated increase in myosin recruitment that leads to a net increase of myofibril force, primarily at submaximal calcium activations. These results match well with our previous report on contractile properties from myectomy samples of the patients from whom the hiPSC-CMs were generated, demonstrating that these cell lines are a good model to study this pathological mutation and extends our understanding of the mechanisms of altered contractile properties of this HCM MYBPC3-c.772G > A mutation.

Detecting Polymorphism of Myosin-binding Protein C3 Gene in Persian Breed Cat With and Without Hypertrophic Cardiomyopathy

Background: In cats, hypertrophic cardiomyopathy (HCM) stands out as a prevailing heart disease. The mutations in the gene that encodes cardiac myosin-binding protein C (MYBPC3) have been detected in the Ragdoll and Maine Coon breeds. Objectives: HCM is believed to be hereditary in other breeds, too. Methods: Blood samples were collected for DNA extraction from 2 unaffected and 7 affected Persian breed cats with HCM. Besides accomplishing conventional polymerase chain reaction, DNA sequencing was performed. The sequence changes were utilized to detect single nucleotide polymorphisms in the MYBPC3 gene and predict amino acid substitutions based on the Acc. No. XM_019812396.1 and comparisons with the literature on identified breed variants and control samples. Results: Although many single nucleotide polymorphisms were found in the affected and unaffected Persian cats, no causative mutation for HCM was observed. Conclusion: In this breed, HCM does not seem to be caused solely by mutations in this cardiac gene. Potential cardiac genes should be investigated to uncover other genetic reasons for this cardiac disease in the Persian cat breed.

Preventing Atrial Fibrillation in Hypertrophic Cardiomyopathy using Angiotensin-Converting Enzyme (ACE) Inhibitors and Angiotensin Receptor Blockers (ARBs)

Hypertrophic cardiomyopathy (HCM), a genetic cardiovascular disease, is the leading cause of cardiac death in young people, often due to atrial fibrillation (AF). AF is generally treated using antiarrhythmics and anticoagulants, which have adverse side effects after long-term use, and are therefore unsuitable for young HCM patients. AF is characterized by a rapid and irregular atrial heartbeat, marked by a short action potential duration and atrial effective refractory period in atrial cardiomyocytes. Prior studies have indicated that the renin-angiotensin system is involved in lowering both, so it has been hypothesized that angiotensin-converting enzyme (ACE) inhibitors and angiotensin receptor blockers (ARBs), which inhibit the renin-angiotensin system, could prevent AF. Therefore, in this study, we propose a research protocol to examine the viability of ACE inhibitors and ARBs as prophylactic measures against the development of AF in HCM patients. To test this, we suggest extracting atrial cardiomyocytes from HCM patients by performing enzyme dissociation on myocardial tissue. The isolated cardiomyocytes will then be treated in vitro with an ACE inhibitor, an ARB, a combination of both, or a control saline solution, and the patch-clamp technique will be used to determine the frequency and duration of their action potentials. We expect action potential duration and atrial effective refractory period to be longer in treated cells, while neither medication will provide a greater advantage, and, as prior research suggests, the combination will not yield significant benefits. The study will continue by testing the effects of ACE inhibitors and ARBs on the function of atrial myocardial organoids created from differentiated stem cells with an HCM mutation. The results of this study could present a new preventative measure against AF for HCM patients which would be safe for long-term use.

Generation of induced pluripotent stem cell lines from two unrelated individuals with familial hypertrophic cardiomyopathy carrying MYBPC3 nonsense mutations

Familial hypertrophic cardiomyopathy (HCM) stands as a predominant heart condition, characterised by left ventricle hypertrophy in the absence of any associated loading conditions, with affected individuals having an increased risk of developing heart failure and sudden cardiac death (SCD). Two induced pluripotent stem cell (iPSC) lines were derived from peripheral blood mononuclear cells obtained from two unrelated individuals with previously reported nonsense mutations in the MYBPC3 gene. The first individual is a 48-year-old male (F26) with the MYBPC3 c.1731G > A HCM mutation, whereas the second individual is a 43-year-old female (F82) carrying the MYBPC3 c.2670G > A HCM mutation. The generated iPSCs exhibit appropriate expression of pluripotency markers, trilineage differentiation capacity and a normal karyotype. This resource contributes to gaining deeper insights into the pathophysiological mechanisms that underlie HCM.

Differentiation of Myocardial Properties in Physiological Athletic Cardiac Remodeling and Mild Hypertrophic Cardiomyopathy.

Clinical differentiation between athletes' hearts and those with hypertrophic cardiomyopathy (HCM) can be challenging. We aimed to explore the role of speckle tracking echocardiography (STE) and cardiac magnetic resonance imaging (CMR) in the differentiation between athletes' hearts and those with mild HCM. We compared 30 competitive endurance elite athletes (7% female, age 41 ± 9 years) and 20 mild phenotypic mutation-positive HCM carriers (15% female, age 51 ± 12 years) with left ventricular wall thickness 13 ± 1 mm. Mechanical dispersion (MD) was assessed by means of STE. Native T1-time and extracellular volume (ECV) were assessed by means of CMR. MD was higher in HCM mutation carriers than in athletes (54 ± 16 ms vs. 40 ± 11 ms, p = 0.001). Athletes had a lower native T1-time (1204 (IQR 1191, 1234) ms vs. 1265 (IQR 1255, 1312) ms, p < 0.001) and lower ECV (22.7 ± 3.2% vs. 25.6 ± 4.1%, p = 0.01). MD > 44 ms optimally discriminated between athletes and HCM mutation carriers (AUC 0.78, 95% CI 0.65-0.91). Among the CMR parameters, the native T1-time had the best discriminatory ability, identifying all HCM mutation carriers (100% sensitivity) with a specificity of 75% (AUC 0.83, 95% CI 0.71-0.96) using a native T1-time > 1230 ms as the cutoff. STE and CMR tissue characterization may be tools that can differentiate athletes' hearts from those with mild HCM.

Myosin with hypertrophic cardiomyopathy mutation M493I demonstrates altered crossbridge kinetics and force sensitivity despite normal working stroke

Myosin with hypertrophic cardiomyopathy mutation M493I demonstrates altered crossbridge kinetics and force sensitivity despite normal working stroke

Hypertrophic cardiomyopathy dysfunction mimicked in human engineered heart tissue and improved by sodium-glucose cotransporter 2 inhibitors.

Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiomyopathy, often caused by pathogenic sarcomere mutations. Early characteristics of HCM are diastolic dysfunction and hypercontractility. Treatment to prevent mutation-induced cardiac dysfunction is lacking. Sodium-glucose cotransporter 2 inhibitors (SGLT2i) are a group of antidiabetic drugs that recently showed beneficial cardiovascular outcomes in patients with acquired forms of heart failure. We here studied if SGLT2i represent a potential therapy to correct cardiomyocyte dysfunction induced by an HCM sarcomere mutation. Contractility was measured of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) harbouring an HCM mutation cultured in 2D and in 3D engineered heart tissue (EHT). Mutations in the gene encoding β-myosin heavy chain (MYH7-R403Q) or cardiac troponin T (TNNT2-R92Q) were investigated. In 2D, intracellular [Ca2+], action potential and ion currents were determined. HCM mutations in hiPSC-CMs impaired relaxation or increased force, mimicking early features observed in human HCM. SGLT2i enhance the relaxation of hiPSC-CMs, to a larger extent in HCM compared to control hiPSC-CMs. Moreover, SGLT2i-effects on relaxation in R403Q EHT increased with culture duration, i.e. hiPSC-CMs maturation. Canagliflozin's effects on relaxation were more pronounced than empagliflozin and dapagliflozin. SGLT2i acutely altered Ca2+ handling in HCM hiPSC-CMs. Analyses of SGLT2i-mediated mechanisms that may underlie enhanced relaxation in mutant hiPSC-CMs excluded SGLT2, Na+/H+ exchanger, peak and late Nav1.5 currents, and L-type Ca2+ current, but indicate an important role for the Na+/Ca2+ exchanger. Indeed, electrophysiological measurements in mutant hiPSC-CM indicate that SGLT2i altered Na+/Ca2+ exchange current. SGLT2i (canagliflozin > dapagliflozin > empagliflozin) acutely enhance relaxation in human EHT, especially in HCM and upon prolonged culture. SGLT2i may represent a potential therapy to correct early cardiac dysfunction in HCM.

Generation of induced pluripotent stem cell lines from two unrelated individuals with familial hypertrophic cardiomyopathy carrying the MYBPC3 missense c.1484G>A mutation

Familial hypertrophic cardiomyopathy (HCM) is the most common inherited heart condition. HCM patients show left ventricle hypertrophy without any associated loading conditions, being at risk for heart failure and sudden cardiac death. Two induced pluripotent stem cell (iPSC) lines were generated from peripheral blood mononuclear cells obtained from two unrelated individuals, a 54-year-old male (F81) and a 44-year-old female (F93), both carrying the MYBPC3 c.1484G>A HCM mutation. iPSCs show expression of pluripotency markers, trilineage differentiation capacity and a normal karyotype. This resource enables further assessment of the pathophysiological development of HCM.

Compound Heterozygosity for Late-Onset Cardiomyopathy-Causative ALPK3 Coding Variant and Novel Intronic Variant Cause Infantile Hypertrophic Cardiomyopathy.

Hypertrophic and dilated cardiomyopathy (HCM, DCM) are leading causes of cardiovascular morbidity and mortality in children. The pseudokinase alpha-protein kinase 3 (ALPK3) plays an essential role in sarcomere organization and cardiomyocyte differentiation. ALPK3 coding mutations are causative of recessively inherited pediatric-onset DCM and HCM with variable expression of facial dysmorphism and skeletal abnormalities and implicated in dominantly inherited adult-onset cardiomyopathy. We now report two variants in ALPK3-a coding variant and a novel intronic variant affecting splicing. We demonstrate that compound heterozygosity for both variants is highly suggestive to be causative of infantile-onset HCM with webbed neck, and heterozygosity for the coding variant presents with adult-onset HCM. Our data validate partial penetrance of heterozygous loss-of-function ALPK3 mutations in late-onset hypertrophic cardiomyopathy and expand the genotypic spectrum of autosomal recessive ALPK3-related cardiac disease with Noonan-like features.