Hypertrophic Cardiomyopathy Genes Research Articles

A personalized mRNA signature for predicting hypertrophic cardiomyopathy applying machine learning methods

Hypertrophic cardiomyopathy (HCM) may lead to cardiac dysfunction and sudden death. This study was designed to develop a HCM signature applying bioinformatics and machine learning methods. Data of HCM and normal tissues were obtained from public databases to screen differentially expressed genes (DEGs) using the R software limma package. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were performed for enrichment analysis of HCM-associated DEGs. Hub genes for HCM were determined using weighted gene co-expression network analysis (WGCNA) together with two machine learning algorithms (SVM-RFE and LASSO). Finally, we introduced a zebrafish model to simulate changes in the hub genes in the HCM and to observe their effects on cardiac disease development. The mRNA expression data from a total of 106 HCM tissues and 39 normal samples were collected and we screened 157 DEGs. Enrichment analysis showed that immune pathways played an important role in the pathogenesis of HCM. Three hub genes (FCN3, MYH6 and RASD1) were identified using WGCNA, SVM-RFE, and LASSO analysis. In a zebrafish model, knockdown of MYH6 and RASD1 resulted in cardiac malformations with reduced ventricular capacity and heart rate, which validated the clinical significance of these genes in the diagnosis of HCM. Based on machine learning algorithms, our study created a signature with potential impact on cardiac function and cardiac quality index for HCM. The current findings had important implications for the early diagnosis and treatment of HCM.

Unveiling Immune Infiltration Characterizing Genes in Hypertrophic Cardiomyopathy Through Transcriptomics and Bioinformatics.

Hypertrophic cardiomyopathy (HCM) is a dominantly inherited disease associated with sudden immune cell associations that remain unclear. The aim of this study was to comprehensively screen candidate markers associated with HCM and immune cells and explore potential pathogenic pathways. First, download the GSE32453 dataset to identify differentially expressed genes (DEGs) and perform Gene Ontology and pathway enrichment analysis using DAVID and GSEA. Next, construct protein-protein interaction (PPI) networks using String and Cytoscape to identify hub genes. Afterward, use CIBERSORT to determine the proportion of immune cells attributed to key genes in HCM and conduct ROC analysis based on the external dataset GSE36961 to evaluate their diagnostic value. Finally, validate the expression of key genes in the hypertrophic cardiomyocyte model through qRT-PCR using data from the HPA database. Comprehensive analysis revealed that there were 254 upregulated genes and 181 downregulated genes in HCM. The enrichment study underscored pathways of inflammatory signaling, including MAPK and PI3K-Akt pathways. Pathways abundant in genes associated with HCM encompassed myocardial contraction and NADH dehydrogenase activity. Additionally, the analysis of immune infiltration revealed a notable increase in macrophages, NK cells, and monocytes in the HCM group, showing statistically significant variances in CD4 memory resting T cell infiltration when compared to the healthy control group. Within the validation dataset GSE36961, the Area Under the Curve (AUC) scores for eight crucial genes (FOS, CD86, CD68, BDNF, PIK3R1, PLEK, RAC2, CCL2) each exceeded 0.8. The HPA database revealed the positioning traits and paths of these eight crucial genes in smooth muscle cells, myocardial cells, and fibroblasts. The outcomes of the qRT-PCR were aligned with the sequencing findings. Bioinformatics analysis unveiled pivotal genes, pathways, and immune involvement, illuminating the molecular underpinnings of HCM. These findings suggest promising therapeutic targets for clinical applications.

Abstract 15631: A Precision Medicine Approach to Predicting Pathogenicity Among Population Variants in Hypertrophic Cardiomyopathy-Associated Genes in the UK Biobank

Introduction: Genomic and precision medicine have opened the door to predictive genetic testing to identify those at risk of cardiac disease before its development. This is important for hypertrophic cardiomyopathy (HCM), the most prevalent heritable heart muscle disease. A major hurdle to accurate predictive testing is the markedly higher prevalence of HCM-associated variants than of disease. New approaches are needed to discern which disease-causative, “actionable,” variants. Objective: To develop and validate a precision medicine approach for predicting pathogenicity of population-identified HCM-associated variants. Methods: We used phenotypic and genotypic data from the UK Biobank (UKBB). We filtered for missense variants in “definitive” (ClinGen) HCM genes. Our novel precision medicine tool, DiscoVari , provides signal-to-noise (S:N) analysis comparing disease to population variant frequency to identify pathogenic genetic ’hotspots’. We used reverse phenotyping to correlate evidence of HCM and sequelae (sudden death, heart failure, ventricular arrhythmias, atrial fibrillation, chest pain, pacemaker/ICD) to S:N hotspots. Variants were re-evaluated using American College of Medical Genetics and Genomics (ACMG) criteria; those in S:N hotspots met PM1 criteria. Results: Variants of uncertain significance (VUSs) in HCM genes had a prevalence of 21.8% in the UKBB, with PM1 determining variant call in 25.7% [24.9-26.6] of HCM-gene variants. Using SN enriched for HCM (0.35% with HCM without PM1 and 0.83% with PM1, p < 0.01). InterVar did not enrich for HCM (0.71% with HCM with PM1 and 0.51% without PM1). DiscoVari reduced the likely pathogenic/pathogenic (LPP) variant burden in individuals without HCM (11.8% compared to InterVar (25.9%, p < 0.0001). DiscoVari enriched for LPP variants in those with HCM (36.1%, p < 0.0001) compared to healthy individuals, while InterVar did not. Conclusion: VUSs are common in HCM genes in the population and PM1 is crucial for accurate variant classification. Incorporating DiscoVari S:N into ACMG guidelines improves prediction of variants associated with HCM, or subclinical cardiomyopathy.

Abstract 14403: Echocardiographic Predictors of Phenotype Development in Hypertrophic Cardiomyopathy Mutation-Carriers

Introduction: Hypertrophic cardiomyopathy (HCM) is a hereditary disease characterized by unexplained left ventricular (LV) hypertrophy (LVH). Genetic family screening of probands identifies mutation carriers (Gen+) without phenotype (Phen-), who require clinical follow-up. Hypothesis: Identification of early phenotype signs may facilitate clinical management of Gen+/Phen-. We assessed if echocardiography LV global longitudinal strain (GLS) and strain rate (SR) may predict LVH development in Gen+/Phen- HCM. Methods: We identified 16 Gen+/Phen- (25% women; age 20±11 years). Echocardiogram was performed at baseline, when no phenotype was evident, and repeated yearly. The end of follow up was the HCM diagnosis date in subjects who developed HCM (FU-Phen+) and the last visit date in those who didn’t develop HCM (FU-Phen-). Mitral inflow peak E and A velocities and Tissue Doppler E’ and A’ peak velocities were measured. LV GLS and SR-peak during isovolumic relaxation (SRIVR) were calculated offline independently by two investigators blind to the clinical data. Results: GLS and SRIVR were not age-dependent (p=0.08 and 0.55, respectively), whereas E/E’ ratio was directly related to age (p=0.011). During follow-up (8±5 years, interquartile range 5-10 years) 5 subjects became FU-Phen+. At baseline, FU-Phen+ compared to FU-Phen- were older and had higher maximal wall thickness, higher EF, less negative GLS and lower SRIVR (Table). Abnormal baseline GLS, despite a normal EF, and reduced SRIVR indicate reduced systolic deformation and covert diastolic dysfunction which may be caused by disarray preceding LVH and fibrosis. Conclusions: This is the first HCM preclinical study to compare at baseline HCM Gen+/Phen- subjects who will develop to those who will not develop LVH. We identified sensitive, easily obtainable, age- and load-independent echocardiographic predictors of phenotype development in HCM gene carriers who may undergo early preventive treatment.

Patients With Hypertrophic Cardiomyopathy and Normal Genetic Investigations Have Few Affected Relatives

BackgroundCurrent guidelines recommend that relatives of index patients with hypertrophic cardiomyopathy (HCM) are offered clinical investigations to identify individuals at risk of adverse disease complications and sudden cardiac death. However, the value of family screening in relatives of index patients with a normal genetic investigation of recognized HCM genes is largely unknown. ObjectivesThe purpose of this study was to perform family screening among relatives of HCM index patients with a normal genetic investigation to establish the frequency of familial disease and the clinical characteristics of affected individuals. MethodsClinical and genetic investigations were performed in consecutive and unrelated HCM index patients. Relatives of index patients who did not carry pathogenic/likely pathogenic variants in recognized HCM genes were invited for clinical investigations. ResultsIn total, 60% (270 of 453) of HCM index patients had a normal genetic investigation. A total of 80% of their relatives (751 of 938, median age 44 years) participated in the study. Of these, 5% (34 of 751) were diagnosed with HCM at baseline, whereas 0.3% (2 of 717 [751–34]) developed the condition during 5 years of follow-up. Their median age at diagnosis was 57 years (IQR: 51-70 years). Two-thirds (22 of 36) were diagnosed following family screening, whereas one-third (14 of 36) had been diagnosed previously because of cardiac symptoms, a murmur, or an abnormal electrocardiogram. None of the affected relatives experienced adverse disease complications. The risk of SCD was low. ConclusionsSystematic family screening of index patients with HCM and normal genetic investigations was associated with a low frequency of affected relatives who appeared to have a favorable prognosis.

High-resolution single-cell transcriptomic survey of cardiomyocytes from patients with hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is a common inherited cardiovascular disease, which can cause heart failure and lead to death. In this study, we performed high-resolution single-cell RNA-sequencing of 2115 individual cardiomyocytes obtained from HCM patients and normal controls. Signature up- and down-regulated genes in HCM were identified by integrative analysis across 37 patients and 41 controls from our data and published human single-cell and single-nucleus RNA-seq datasets, which were further classified into gene modules by single-cell co-expression analysis. Using our high-resolution dataset, we also investigated the heterogeneity among individual cardiomyocytes and revealed five distinct clusters within HCM cardiomyocytes. Interestingly, we showed that some extracellular matrix (ECM) genes were up-regulated in the HCM cardiomyocytes, suggesting that they play a role in cardiac remodelling. Taken together, our study comprehensively profiled the transcriptomic programs of HCM cardiomyocytes and provided insights into molecular mechanisms underlying the pathogenesis of HCM.

Major Cardiac Events in Patients and Relatives With Hereditary Hypertrophic Cardiomyopathy

BackgroundLittle evidence is available on the disease expression in relatives of index patients with hypertrophic cardiomyopathy (HCM). This information has important implications for family screening programs, genetic counseling, and management of affected families. ObjectivesThe purpose of this study was to investigate the disease expression and penetrance in relatives of index patients carrying pathogenic/likely pathogenic (P/LP) variants in recognized HCM genes. MethodsA total of 453 consecutive and unrelated HCM index patients underwent clinical and genetic investigations. A total of 903 relatives of genotype-positive index patients were invited for clinical investigations and genetic testing. Penetrance, disease expression, and incidence rates of major adverse cardiac events (MACEs) were investigated in individuals carrying P/LP variants. ResultsForty percent (183/453) of index patients carried a P/LP variant. Eighty-four percent (757/903) of all relatives of index patients with P/LP variants were available for the investigation, of whom 54% (407/757) carried a P/LP variant. The penetrance of HCM among relatives was 39% (160/407). Relatives with HCM and index patients were diagnosed at a similar age (43 ± 18 years vs 46 ± 15 years; P = 0.11). There were no differences in clinical characteristics or incidence rates of MACE during 8 years of follow-up. ConclusionsThe disease expression of HCM among index patients and affected relatives carrying P/LP variants in recognized disease genes was similar, with an equal risk of experiencing MACE. These findings provide evidence to support family screening and follow-up of genotype-positive HCM families to improve management and diminish the number of adverse disease complications among relatives.

Identification and analysis of key hypoxia- and immune-related genes in hypertrophic cardiomyopathy

BackgroundHypertrophic cardiomyopathy (HCM), an autosomal dominant genetic disease, is the main cause of sudden death in adolescents and athletes globally. Hypoxia and immune factors have been revealed to be related to the pathology of HCM. There is growing evidence of a role for hypoxia and inflammation as triggers and enhancers in the pathology in HCM. However, the role of hypoxia- and immune-related genes in HCM have not been reported.MethodsFirstly, we obtained four HCM-related datasets from the Gene Expression Omnibus (GEO) database for differential expression analysis. Immune cells significantly expressed in normal samples and HCM were then screened by a microenvironmental cell population counter (MCP-counter) algorithm. Next, hypoxia- and immune-related genes were screened by the LASSO + support vector machine recursive feature elimination (SVM-RFE) and weighted gene co-expression network analysis (WGCNA). Single-gene enrichment analysis and expression validation of key genes were then performed. Finally, we constructed a competing endogenous RNA (ceRNA) network of key genes.ResultsIn this study, 35 differentially expressed hypoxia genes were found. By using LASSO + SVM-RFE analysis, 10 more targets with differentially expressed hypoxia genes were identified. The MCP-count algorithm yielded five differentially expressed immune cells, and after assessing them for WGCNA characteristics, 612 immune genes were discovered. When hypoxia and immune genes were combined for cross-tabulation analysis, three hypoxia- and immune-related genes (ATP2A2, DDAH1, and OMA1) were identified.ConclusionBased on hypoxia characteristic genes, three key genes were identified. These were also significantly related to immune activation, which proves a theoretical basis and reference value for studying the relationship between HCM and hypoxia and immunity.Graphical

Abstract P3038: Novel MicroRNAs Targeting Myosin Binding Protein-C3 Gene In Hypertrophic Cardiomyopathy

Background: Mutations in cardiac myosin binding protein-C3 ( MYBPC3 ) constitute a significant cause of hypertrophic cardiomyopathy (HCM) with incomplete penetrance, and later age of onset. MYBPC3 variant with 25 base pair deletion in intron 32 ( MYBPC3 Δ25bp ) leads to exon 33 skipping and 3’-untranslated region truncation. Independent transcriptomics studies in HCM individuals carrying c.927-2A>G, c.2373insG, c.3624delC and c.3369_3370insC MYBPC3 variants reported altered levels of cardiac and circulating microRNAs (miRNAs). However, the miRNA-mediated post-transcriptional regulation of the MYBPC3 and MYBPC3 Δ25bp in the HCM phenotype remains largely unknown. Methods: Using a computational consensus approach, top-scoring cardiac-specific miRNAs predicted to target wild-type MYBPC3 (WT) and MYBPC3 Δ25bp were identified. To further characterize the potential regulators, the miRNAs with earlier reports on cardiac and skeletal diseases were selected and their targets were intersected with a compiled set of HCM and heart failure (HF) candidate genes. The overlapping genes were subjected to enrichment analysis for biological processes. Results: The computational analyses predicted 7 high scoring miRNAs for MYBPC3, among which miR-146b-3p, miR-2355-5p, miR-4508, miR-4800-5p and miR-214-5p were predicted to regulate both the WT and MYBPC3 Δ25bp variant, whereas the binding sites for miR-5588-5p, and miR-6500-3p were lost in MYBPC3 Δ25bp variant. In specific, miR-146-3p, miR-214-5p, and miR-2355-5p have been reported to be associated with skeletal muscle dysfunction, HCM and myocardial ischemia-reperfusion injury, respectively. The putative targets of these 3 miRNAs intersected with HCM and HF candidate genes, in addition the intersected targets displayed enrichment for cardiac disease-associated biological processes. Conclusions: Computational prediction suggests that predicted MYBPC3 miRNAs affect biological processes and pathways associated with HCM and HF. Importantly, MYBPC3 Δ25bp show a loss of miRNA regulation compared to the WT. Further studies and experimental validations are required to determine the influence of loss of miRNA regulation in the MYBPC3 Δ25bp and their impact on HCM interventions.

Ethnicity, consanguinity, and genetic architecture of hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is characterized by phenotypic heterogeneity that is partly explained by the diversity of genetic variants contributing to disease. Accurate interpretation of these variants constitutes a major challenge for diagnosis and implementing precision medicine, especially in understudied populations. The aim is to define the genetic architecture of HCM in North African cohorts with high consanguinity using ancestry-matched cases and controls. Prospective Egyptian patients (n = 514) and controls (n = 400) underwent clinical phenotyping and genetic testing. Rare variants in 13 validated HCM genes were classified according to standard clinical guidelines and compared with a prospective HCM cohort of majority European ancestry (n = 684). A higher prevalence of homozygous variants was observed in Egyptian patients (4.1% vs. 0.1%, P = 2 × 10-7), with variants in the minor HCM genes MYL2, MYL3, and CSRP3 more likely to present in homozygosity than the major genes, suggesting these variants are less penetrant in heterozygosity. Biallelic variants in the recessive HCM gene TRIM63 were detected in 2.1% of patients (five-fold greater than European patients), highlighting the importance of recessive inheritance in consanguineous populations. Finally, rare variants in Egyptian HCM patients were less likely to be classified as (likely) pathogenic compared with Europeans (40.8% vs. 61.6%, P = 1.6 × 10-5) due to the underrepresentation of Middle Eastern populations in current reference resources. This proportion increased to 53.3% after incorporating methods that leverage new ancestry-matched controls presented here. Studying consanguineous populations reveals novel insights with relevance to genetic testing and our understanding of the genetic architecture of HCM.

Pathogenic BCS1L Mutation Resulting in Hypertrophic Cardiomyopathy: A Unique Presentation of Nuclear Mitochondrial Disease.

A 21-year-old man with sensorineural hearing loss and glaucoma presented with severely limited exercise capacity since childhood. He was found to have biventricular concentric hypertrophy with greatest wall thickening at the posterior and lateral walls of the left ventricle apex (1.7 cm) and the free wall of the right ventricle (1.1 cm). There was no inducible left ventricular outflow tract obstruction. Metabolic testing revealed marked lactic aciduria (1,650.1 μmol/mmol creatinine) and plasma lactate (3.9 mmol/L). A sarcomeric hypertrophic cardiomyopathy gene panel was unremarkable, but mitochondrial gene analysis revealed a homozygous c.385G>A (p.Gly129Arg) pathogenic mutation in the BCS1L gene. This gene is responsible for an assembly subunit of cytochrome complex III in the respiratory transport chain and is the rarest respiratory chain defect. This gene has not frequently been implicated in cardiomyopathy. Mitochondrial hypertrophic cardiomyopathy is more rare than hypertrophic cardiomyopathy resulting from sarcomeric mutations and is more likely to be symmetric, less frequently results in left ventricular outflow tract obstruction, and is more likely to progress to dilated cardiomyopathy. Evidence-based screening protocols have not been established; treatment follows guideline-directed medical therapy for congestive heart failure, including evaluation for heart transplantation. This report expands the phenotype of the BCS1L mutation and suggests that affected patients may need screening for underlying cardiomyopathy.

Novel Genes Involved in Hypertrophic Cardiomyopathy: Data of Transcriptome and Methylome Profiling

Hypertrophic cardiomyopathy (HCM) is the most common inherited heart disease; its pathogenesis is still being intensively studied to explain the reasons for the significant genetic and phenotypic heterogeneity of the disease. To search for new genes involved in HCM development, we analyzed gene expression profiles coupled with DNA methylation profiles in the hypertrophied myocardia of HCM patients. The transcriptome analysis identified significant differences in the levels of 193 genes, most of which were underexpressed in HCM. The methylome analysis revealed 1755 nominally significant differentially methylated positions (DMPs), mostly hypomethylated in HCM. Based on gene ontology enrichment analysis, the majority of biological processes, overrepresented by both differentially expressed genes (DEGs) and DMP-containing genes, are involved in the regulation of locomotion and muscle structure development. The intersection of 193 DEGs and 978 DMP-containing genes pinpointed eight common genes, the expressions of which correlated with the methylation levels of the neighboring DMPs. Half of these genes (AUTS2, BRSK2, PRRT1, and SLC17A7), regulated by the mechanism of DNA methylation, were underexpressed in HCM and were involved in neurogenesis and synapse functioning. Our data, suggesting the involvement of innervation-associated genes in HCM, provide additional insights into disease pathogenesis and expand the field of further research.

Identification of the Immune Status of Hypertrophic Cardiomyopathy by Integrated Analysis of Bulk- and Single-Cell RNA Sequencing Data.

Objectives Hypertrophic cardiomyopathy (HCM) is the most common hereditary cardiomyopathy and immune infiltration is considered an indispensable factor involved in its pathogenesis. In this study, we attempted to combine bulk sequencing and single-cell sequencing to map the immune infiltration-related genes in hypertrophic cardiomyopathy. Methods The GSE36961, GSE160997, and GSE122930 datasets were obtained from the Gene Expression Omnibus database. The compositional patterns of the 18 types of immune cell fraction and pathway enrichment score in control and HCM patients were estimated based on the GSE36961 cohort using xCell algorithm. The Weighted Gene Coexpression Network Analysis (WGCNA) was performed to identify genes associated with immune infiltration for hypertrophic cardiomyopathy. The area under the curve (AUC) value was obtained and used to evaluate the discriminatory ability of common immune-related DEGs. “NetworkAnalyst” platform was used to identify TF-gene and TF-miRNA interaction with identified common genes. Heat map was used to determine the association between common DEGs and various immune cells. Results Immune infiltration analysis by the xCell algorithm showed a higher level of CD8+ naive T cells, CD8+ T cells, as well as a lower level of activated dendritic cells (aDC), dendritic cells (DC), immature dendritic cells (iDC), conventional dendritic cells (cDC), macrophages, M1 macrophages, monocytes, and NKT cell in HCM compared with the control group in GSE36961 dataset. aDC, macrophages, and M1 macrophages were the top three discriminators between HCM and control groups with the area under the curve (AUC) of 0.907, 0.867, and 0.941. WGCNA analysis showed that 1258 immune-related genes were included in four different modules. Of these modules, the turquoise module showed a pivotal correlation with HCM. 13 common immune-related DEGs were found by intersecting common DEGs in GSE36961 and GSE160997 datasets with genes from the genes in turquoise module. 5 hub immune-related genes (S100A9, TYROBP, FCER1G, CD14, and S100A8) were identified by protein interaction network. Through analysis of single-cell sequencing data, S100a9, TYROBP, FCER1G, and S100a8 were mainly expressed by infiltrated M1 proinflammatory cells, especially Ccr2-M1 proinflammatory macrophage cells in the heart immune microenvironment while Cd14 was expressed by infiltrated M1 proinflammatory macrophage cells and M2 macrophages in transverse aortic constriction (TAC) mice at 1 week. Higher M2 macrophage and M1 proinflammatory macrophage infiltration as well as lower Ccr2-M1 proinflammatory macrophage and dendritic cells were shown in TAC 1week mice compared with sham mice. Conclusions There was a difference in immune infiltration between HCM patients and normal groups. aDC, macrophages, and M1 macrophages were the top three discriminator immune cell subsets between HCM and control groups. S100A9, TYROBP, FCER1G, CD14, and S100A8 were identified as potential biomarkers to discriminate HCM from the control group. S100a9, TYROBP, FCER1G, and S100a8 were mainly expressed by infiltrated M1 proinflammatory cells, especially Ccr2-M1 proinflammatory cells in the heart immune microenvironment while Cd14 was expressed by M2 macrophages in transverse aortic constriction (TAC) mice at 1 week.

Homozygosity predominantly affects hypertrophic cardiomyopathy minor genes in an Egyptian clinical cohort

Abstract Background Consanguinity is prevalent in Egypt (35%) resulting in a high incidence of homozygosity. The influence of homozygosity on the genetics of Hypertrophic Cardiomyopathy (HCM) has not been adequately studied. Purpose The purpose of this study is to define the genetic architecture of HCM in Egypt using ethnically-matched case and control cohorts. Methods Prospective Egyptian patients (n=514) and controls (n=400) were recruited to Aswan Heart Centre for clinical phenotyping and genetic testing for 174 genes implicated in inherited cardiac conditions (Illumina). Rare variation (gnomAD filtering allele frequency ≤4x10–5) in 13 validated HCM genes were classified according to the American College of Medical Genetics (ACMG) guidelines and compared with a prospective HCM cohort of predominantly European ancestry (n=684). Results Significantly fewer rare variants detected in Egyptian patients could be classified as (likely) pathogenic compared to Europeans (40.8% vs. 61.6%, p-value=1.6x10–5). Incorporating analysis from these Egyptian case-control cohorts into the ACMG guidelines increased this yield to 53.8%. Homozygous variants were more frequently observed in Egyptian patients (4.1% vs 0.1%, p-value=2x10–7), with variants in the minor HCM genes MYL2, MYL3 and CSRP3 more likely to present in homozygosity than the major genes (MYH7, MYBPC3 and troponins), suggesting such variants are less penetrant in the heterozygous state. Conclusions The integration of Egyptian-specific genetic and phenotypic data significantly improves variant interpretation in HCM and consequently the precision of genetic testing. The observed prevalence of homozygosity and rare variation in minor HCM genes in Egyptian patients provides important insights into its disease-mechanisms and genetics. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Science and Technology Development FundAl Alfi Foundation

Representativeness of EXPLORER-HCM trial and prevalence of eligibility criteria for Mavacamten in a nationwide cohort

Abstract Introduction The EXPLORER-HCM trial evaluated the safety and efficacy of Mavacamten, a cardiac myosin inhibitor, in symptomatic obstructive hypertrophic cardiomyopathy (HCM). Mavacamten improved symptoms, exercise capacity and left ventricular outflow tract (LVOT) obstruction. However, the proportion of patients eligible for this therapy in real practice remains unclear. Purpose To determine the prevalence of eligibility criteria for Mavacamten and to describe the profile of patients complying with EXPLORER-HCM criteria in a nationwide cohort. Methods Cross-sectional study including all HCM patients enrolled in a European country-based nationwide registry. Presence of EXPLORER-HCM inclusion [≥18 years, obstructive HCM (LVOT gradient ≥50 mmHg), left ventricular ejection fraction (LVEF) ≥55%, New York Heart Association (NYHA) class II–III] and exclusion criteria [syncope or sustained ventricular tachycardia with exercise] were analyzed. Results Of 1042 patients included in this analysis (57.8% male, mean age at diagnosis 52 years, 39.6% obstructive HCM), 45 met the EXPLORER-HCM inclusion criteria and, after applying exclusion criteria, 30 patients were eligible for therapy with Mavacamten. Eligible patients were all Caucasian, mostly female (63.3%), with a mean age at diagnosis of 58±14 years. All patients presented with heart failure symptoms, and were mostly in NYHA class II (83.3%); 4 patients (13.3%) presented with atrial fibrillation. In the baseline echocardiogram, mean LVEF was 68±9%; mean interventricular septum thickness was 20±4 mm; most patients presented with systolic anterior motion of the mitral valve (96.7%) and mitral regurgitation (96.7%). In cardiac magnetic resonance imaging evaluation, maximal wall thickness was 22±4 mm and 76.9% had late gadolinium enhancement. A pathogenic or likely pathogenic HCM gene variant was identified in 5 patients (MYH7 and MYBPC3 gene mutations). In this group, 86.2% of patients were treated with beta-blockers and 34.5% with calcium channel antagonists. Only 1 patient was on Disopyramide. Septal reduction techniques were used in 43.3% of patients: alcohol septal ablation and septal myectomy were performed in 7 (23.3%) and 6 (20.0%) patients, respectively. Cardioverter-defibrillators were implanted in 2 patients for primary prevention of sudden cardiac death, and 6 patients received a pacemaker for bradycardia indications. Conclusion Specific therapies remain an unmet need in HCM. In this large nationwide cohort, only 2.9% of patients were eligible for therapy with Mavacamten according to EXPLORER-HCM criteria. Among patients complying with the study criteria, invasive septal reduction therapies were offered to a high proportion of patients. Further real-world data are warranted to estimate the proportion of HCM patients that will benefit from Mavacamten. Funding Acknowledgement Type of funding sources: None.

Human versus machine: does artificial intelligence add value to identification of hypertrophic cardiomyopathy in pediatric patients?

Abstract Introduction An abnormal increase in left ventricular thickness is a hallmark of hypertrophic cardiomyopathy (HCM). Although standard measures of left ventricular voltage abnormalities on electrocardiograms (ECGs) have high false positive rates and poor correlation to left ventricular thickness, ECGs continue to be part of most screening programs. We developed a machine learning model for HCM classification from ECG in a pediatric cohort and compared its efficacy to that of clinician specialists. Purpose To compare clinician-based predictions of HCM to a machine learning model trained to classify pediatric patients with HCM utilizing 12-lead ECG. Methods ECGs from patients <19 years with HCM, including those with HCM gene mutations, were compared to those from age- and sex-matched controls with normal heart structure and function on echocardiogram. Patients with a known history of primary causes of left ventricular hypertrophy such as aortic stenosis or glycogen storage disease were excluded. A cascaded convolutional neural network was developed combining a residual neural network with a 2-layer dense neural network with 10-fold cross validation. The performance of the machine learning based HCM classification was compared to that of two independent clinicians. Results Analytic sample included data from 82 patients with clinical HCM compared to 91 healthy control subjects. The machine model AUC was 0.89 (0.84–0.94). Clinician inter-rater reliability was 0.8. The clinicians had a higher specificity (97% vs 82%) but lower sensitivity (50% vs 78%) than the machine learning model. Compared to clinician-classification, the positive predictive value was lower in the machine model (82% vs 93%). Conclusion In this preliminary study, machine learning classification of HCM utilizing 12-lead ECG had greater sensitivity than that of clinician interpretation. Machine learning may play a role in screening and diagnosis, including in subjects with normal-appearing ECG. Evaluation of its utility in a larger cohort is needed. Funding Acknowledgement Type of funding sources: Public hospital(s). Main funding source(s): Le Bonheur Children's Hospital

IRX2 activated by jumonji domain-containing protein 2A is crucial for cardiac hypertrophy and dysfunction in response to the hypertrophic stimuli

BackgroundIroquois homeobox 2 (IRX2) is a member of the Iroquois family whose upregulation has been potentially correlated to cardiac hypertrophy. This work studied the function of IRX2 and its related molecules in hypertrophic cardiomyopathy (HCM). MethodsA GEO dataset GSE32453 was analyzed to identify aberrantly expressed genes in HCM. Altered expression of IRX2 was induced in mice by lentivirus injection, followed by angiotensin II (Ang II) treatment to induce HCM. The function of IRX2 knockdown in ventricular dysfunction, heart volume and pathological changes in mice, and in surface area, oxidative stress and apoptosis of isolated cardiomyocytes were examined. Binding relationship between jumonji domain-containing protein 2A (JMJD2A) and IRX2 was predicted by online tools and validated. The interaction between JMJD2A and IRX2 in HCM development was examined by joint interventions. ResultsIRX2 was highly expressed in heart tissues with HCM. IRX2 knockdown prevented mice from Ang II-induced ventricular dysfunction, cardiac hypertrophy, inflammation and fibrosis in mouse heart, and it decreased the levels of cardiac hypertrophy-related markers, oxidative stress response, and apoptosis of Ang II-treated cardiomyocytes. JMJD2A catalyzed demethylation of H3K9me3 near the IRX2 promoter to activate its transcription. JMJD2A knockdown similarly exerted protective functions against cardiac hypertrophy in vivo and in vitro, but the protection was blocked upon further IRX2 upregulation. IRX2 was found to increase the Wnt/β-catenin signaling activation. ConclusionThis work reports that JMJD2A activates IRX2 transcription and the Wnt/β-catenin signaling to induce cardiac hypertrophy and dysfunction in HCM.

Genomic findings of hypertrophic and dilated cardiomyopathy characterized in a Thai clinical genetics service.

Hypertrophic cardiomyopathy (HCM) and dilated cardiomyopathy (DCM) are the most common referrals in the Inherited Cardiovascular Condition (ICC) Genetics Service. Several issues must be discussed with patients and their families during the genetic consultation session, including the options for genetic testing and cardiovascular surveillance in family members. We developed an ICC registry and performed next-generation-based DNA sequencing for all patients affected by non-syndromic HCM and idiopathic DCM in our joint specialist genetics service. The target gene sequencing panel relied on the Human Phenotype Ontology with 237 genes for HCM (HP:0001639) and 142 genes for DCM (HP:0001644). All subjects were asked to contact their asymptomatic first-degree relatives for genetic counseling regarding their risks and to initiate cardiovascular surveillance and cascade genetic testing. The study was performed from January 1, 2014, to December 31, 2020, and a total of 62 subjects (31-HCM and 31-DCM) were enrolled. The molecular detection frequency was 48.39% (32.26% pathogenic/likely pathogenic, 16.13% variant of uncertain significance or VUS for HCM, and 25.81% (16.13% pathogenic/likely pathogenic, 9.68% VUS) for DCM. The most prevalent gene associated with HCM was MYBPC3. The others identified in this study included ACTN2, MYL2, MYH7, TNNI3, TPM1, and VCL. Among the DCM subjects, variants were detected in two cases with the TTN nonsense variants, while the others were missense and identified in MYH7, DRSP3, MYBPC3, and SCN5A. Following the echocardiogram surveillance and cascade genetic testing in the asymptomatic first-degree relatives, the detection rate of new cases was 8.82% and 6.25% in relatives of HCM and DCM subjects, respectively. Additionally, a new pre-symptomatic relative belonging to an HCM family was identified, although the genomic finding in the affected case was absent. Thus, ICC service is promising for the national healthcare system, aiming to prevent morbidity and mortality in asymptomatic family members.

The «Amish» NM_000256.3:c.3330+2T>G splice variant in MYBPC3 associated with hypertrophic cardiomyopathy is an ancient Swiss mutation

MYBPC3 is the most frequently mutated gene in hypertrophic cardiomyopathy (HCM). Several loss-of-function founder variants have been reported in MYBPC3 from various geographic regions, altogether suggestive of a modest or absent effect of these variants on reproductive fitness. One of them, a MYBPC3 splice variant, NM_000256.3:c.3330+2T > G, was first described in homozygous state in newborns presenting with a severe, recessive form of HCM among the Amish population and was later associated with adult-onset dominant HCM in heterozygous carriers. We here report this splice variant in heterozygous state in eight unrelated Swiss families with HCM, making it the most prevalent cardiomyopathy variant in western Switzerland. This variant was identified in patients using targeted (n = 5) or full-genome sequencing (n = 3). Given the prevalence of this variant in the Old Order Amish, Mennonites and Swiss populations, and given that both Amish and Mennonites founders originated from the Bern Canton in Switzerland, the MYBPC3, NM_000256.3:c.3330+2T > G variant appears to be of Swiss origin. Neighboring regions that hosted the first Amish settlements (Alsace, South Germany) should be on the lookout for that variant. The existence of MYBPC3 founder variants in different populations suggests that individuals with early-onset clinical disease may be the tip of the iceberg of a much larger number of asymptomatic carriers. Alternatively, reproductive fitness could even be slightly increased in some variant carriers to compensate for the reduction of fitness in the more severely affected ones, but this remains to be investigated.

Myosins and MyomiR Network in Patients with Obstructive Hypertrophic Cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is the most common genetic cardiomyopathy. The molecular mechanisms determining HCM phenotypes are incompletely understood. Myocardial biopsies were obtained from a group of patients with obstructive HCM (n = 23) selected for surgical myectomy and from 9 unused donor hearts (controls). A subset of tissue-abundant myectomy samples from HCM (n = 10) and controls (n = 6) was submitted to laser-capture microdissection to isolate cardiomyocytes. We investigated the relationship among clinical phenotype, cardiac myosin proteins (MyHC6, MyHC7, and MyHC7b) measured by optimized label-free mass spectrometry, the relative genes (MYH7, MYH7B and MYLC2), and the MyomiR network (myosin-encoded microRNA (miRs) and long-noncoding RNAs (Mhrt)) measured using RNA sequencing and RT-qPCR. MyHC6 was lower in HCM vs. controls, whilst MyHC7, MyHC7b, and MyLC2 were comparable. MYH7, MYH7B, and MYLC2 were higher in HCM whilst MYH6, miR-208a, miR-208b, miR-499 were comparable in HCM and controls. These results are compatible with defective transcription by active genes in HCM. Mhrt and two miR-499-target genes, SOX6 and PTBP3, were upregulated in HCM. The presence of HCM-associated mutations correlated with PTBP3 in myectomies and with SOX6 in cardiomyocytes. Additionally, iPSC-derived cardiomyocytes, transiently transfected with either miR-208a or miR-499, demonstrated a time-dependent relationship between MyomiRs and myosin genes. The transfection end-stage pattern was at least in part similar to findings in HCM myectomies. These data support uncoupling between myosin protein/genes and a modulatory role for the myosin/MyomiR network in the HCM myocardium, possibly contributing to phenotypic diversity and providing putative therapeutic targets.