Rare loss-of-function variants in matrisome genes are enriched in Ebstein's anomaly.

Ebstein's anomaly, a rare congenital heart disease, is distinguished by the failure of embryological delamination of the tricuspid valve leaflets from the underlying primitive right ventricle myocardium. Gaining insight into the genetic basis of Ebstein's anomaly allows a more precise definition of its pathogenesis. In this study, two distinct cohorts from the Chinese Han population were included: a case-control cohort consisting of 82 unrelated cases and 125 controls without cardiac phenotypes and a trio cohort comprising 36 parent-offspring trios. Whole-exome sequencing data from all 315 participants were utilized to identify qualifying variants, encompassing rare (minor allele frequency < 0.1% from East Asians in the gnomAD database) functional variants and high-confidence (HC) loss-of-function (LoF) variants. Various statistical models, including burden tests and variance-component models, were employed to identify rare variants, genes, and biological pathways associated with Ebstein's anomaly. Significant associations were noted between Ebstein's anomaly and rare HC LoF variants found in genes related to the matrisome, a collection of extracellular matrix (ECM) components. Specifically, 47 genes with HC LoF variants were exclusively or predominantly identified in cases, while nine genes showed such variants in the probands. Over half of unrelated cases (n = 42) and approximately one-third of probands (n = 12) were found to carry one or two LoF variants in these prioritized genes. These results highlight the role of the matrisome in the pathogenesis of Ebstein's anomaly, contributing to a better understanding of the genetic architecture underlying this condition. Our findings hold the potential to impact the genetic diagnosis and treatment approaches for Ebstein's anomaly.

Heterozygous nonsense variants in laminin subunit 3α resulting in Ebstein's anomaly.

Ebstein's anomaly is a rare congenital heart disease characterized by tricuspid valve downward displacement and is associated with additional cardiac phenotypes such as left ventricle non-compaction. The genetic basis of Ebstein's anomaly has yet to be fully elucidated, although several genes (e.g., NKX2-5, MYH7, TPM1, and FLNA) may contribute to Ebstein's anomaly. Here, in two Ebstein's anomaly families (a three-generation family and a trio), we identified independent heterozygous nonsense variants in laminin subunit 3 α (LAMA3), cosegregated with phenotypes in families with reduced penetrance. Furthermore, knocking out Lama3 in mice revealed that haploinsufficiency of Lama3 led to Ebstein's malformation of the tricuspid valve and an abnormal basement membrane structure. In conclusion, we identified a novel gene-disease association of LAMA3 implicated in Ebstein's anomaly, and the findings extended our understanding of the role of the extracellular matrix in Ebstein's anomaly etiology.

Altered contractility, Ca2+ transients, and cell morphology seen in a patient-specific iPSC-CM model of Ebstein's anomaly with left ventricular noncompaction.

Patients with two congenital heart diseases (CHDs), Ebstein's anomaly (EA) and left ventricular noncompaction (LVNC), suffer higher morbidity than either CHD alone. The genetic etiology and pathogenesis of combined EA/LVNC remain largely unknown. We investigated a familial EA/LVNC case associated with a variant (p.R237C) in the gene encoding Kelch-like protein 26 (KLHL26) by differentiating induced pluripotent stem cells (iPSCs) generated from affected and unaffected family members into cardiomyocytes (iPSC-CMs) and assessing iPSC-CM morphology, function, gene expression, and protein abundance. Compared with unaffected iPSC-CMs, CMs containing the KLHL26 (p.R237C) variant exhibited aberrant morphology including distended endo(sarco)plasmic reticulum (ER/SR) and dysmorphic mitochondria and aberrant function that included decreased contractions per minute, altered calcium transients, and increased proliferation. Pathway enrichment analyses based on RNASeq data indicated that the "structural constituent of muscle" pathway was suppressed, whereas the "ER lumen" pathway was activated. Taken together, these findings suggest that iPSC-CMs containing this KLHL26 (p.R237C) variant develop dysregulated ER/SR, calcium signaling, contractility, and proliferation.NEW & NOTEWORTHY We demonstrate here that iPSCs derived from patients with Ebstein's anomaly and left ventricular noncompaction, when differentiated into cardiomyocytes, display significant structural and functional changes that offer insight into disease pathogenesis, including altered ER/SR and mitochondrial morphology, contractility, and calcium signaling.

A case of capillary malformation-arteriovenous malformation and Ebstein's anomaly in a child with EphB4 mutation.

Capillary malformation-arteriovenous malformation (CM-AVM) is a rare condition characterized by multiple cutaneous capillary malformations with potential associated arteriovenous malformations. RAS p21 protein activator 1 (RASA1) and ephrin type-B receptor 4 (EPHB4) genes are implicated. We present a child with CM-AVM, due to EPHB4 mutation, and Ebstein's anomaly. Although EPHB4 is a known effector of vascular remodeling, its contribution to cardiogenesis is still being explored. Further research is needed to determine causality of Ebstein's anomaly in the setting of CM-AVM due to EPHB4 mutation.

Integrated microRNA and mRNA Expression Profiling Identifies Novel Targets and Networks Associated with Ebstein's Anomaly.

Little is known about abundance level changes of circulating microRNAs (miRNAs) and messenger RNAs (mRNA) in patients with Ebstein's anomaly (EA). Here, we performed an integrated analysis to identify the differentially abundant miRNAs and mRNA targets and to identify the potential therapeutic targets that might be involved in the mechanisms underlying EA. A large panel of human miRNA and mRNA microarrays were conducted to determine the genome-wide expression profiles in the blood of 16 EA patients and 16 age and gender-matched healthy control volunteers (HVs). Differential abundance level of single miRNA and mRNA was validated by Real-Time quantitative PCR (RT-qPCR). Enrichment analyses of altered miRNA and mRNA abundance levels were identified using bioinformatics tools. Altered miRNA and mRNA abundance levels were observed between EA patients and HVs. Among the deregulated miRNAs and mRNAs, 76 miRNAs (49 lower abundance and 27 higher abundance, fold-change of ≥2) and 29 mRNAs (25 higher abundance and 4 lower abundance, fold-change of ≥1.5) were identified in EA patients compared to HVs. Bioinformatics analysis identified 37 pairs of putative miRNA-mRNA interactions. The majority of the correlations were detected between the lower abundance level of miRNA and higher abundance level of mRNA, except for let-7b-5p, which showed a higher abundance level and their target gene, SCRN3, showed a lower abundance level. Pathway enrichment analysis of the deregulated mRNAs identified 35 significant pathways that are mostly involved in signal transduction and cellular interaction pathways. Our findings provide new insights into a potential molecular biomarker(s) for the EA that may guide the development of novel targeting therapies.

Ebstein anomaly associated with cri du chat (cat's cry) syndrome and 20q duplication.

Ebstein anomaly is a congenital heart defect with a low prevalence and high mortality in the early stages of life. In medical literature, there is no reported association between Ebstein anomaly and cri du chat syndrome. Here, we report the case of a full-term newborn with a low weight for his age and who had a prenatal diagnosis of Ebstein anomaly and a postnatal diagnosis of cri du chat syndrome and 20q duplication detected on array CGH. The patient required medical treatment with inotropic support, high-frequency ventilation and nitric oxide, with an adequate response. Surgical intervention was not needed.

Novel KLHL26 variant associated with a familial case of Ebstein's anomaly and left ventricular noncompaction.

BACKGROUND: Ebstein's anomaly (EA) is a rare congenital heart disease of the tricuspid valve and right ventricle. Patients with EA often manifest with left ventricular noncompaction (LVNC), a cardiomyopathy. Despite implication of cardiac sarcomere genes in some cases, very little is understood regarding the genetic etiology of EA/LVNC. Our study describes a multigenerational family with at least 10 of 17 members affected by EA/LVNC. METHODS: We performed echocardiography on all family members and conducted exome sequencing of six individuals. After identifying candidate variants using two different bioinformatic strategies, we confirmed segregation with phenotype using Sanger sequencing. We investigated structural implications of candidate variants using protein prediction models. RESULTS: Exome sequencing analysis of four affected and two unaffected members identified a novel, rare, and damaging coding variant in the Kelch-like family member 26 (KLHL26) gene located on chromosome 19 at position 237 of the protein (GRCh37). This variant region was confirmed by Sanger sequencing in the remaining family members. KLHL26 (c.709C > T p.R237C) segregates only with EA/LVNC-affected individuals (FBAT p < .05). Investigating structural implications of the candidate variant using protein prediction models suggested that the KLHL26 variant disrupts electrostatic interactions when binding to part of the ubiquitin proteasome, specifically Cullin3 (CUL3), a component of E3 ubiquitin ligase. CONCLUSION: In this familial case of EA/LVNC, we have identified a candidate gene variant, KLHL26 (p.R237C), which may have an important role in ubiquitin-mediated protein degradation during cardiac development.

Left ventricular non-compaction with Ebstein anomaly attributed to a TPM1 mutation.

Left ventricular non-compaction (cardiomyopathy) (LVN(C)) is a rare hereditary cardiac condition, resulting from abnormal embryonic myocardial development. While it mostly occurs as an isolated condition, association with other cardiovascular manifestations such as Ebstein anomaly (EA) has been reported. This congenital heart defect is characterized by downward displacement of the tricuspid valve and leads to diminished ventricular size and function. In an autosomal dominant LVN(C) family consisting of five affected individuals, of which two also presented with EA and three with mitral valve insufficiency, we pursued the genetic disease cause using whole exome sequencing (WES). WES revealed a missense variant (p.Leu113Val) in TPM1 segregating with the LVN(C) phenotype. TPM1 encodes α-tropomyosin, which is involved in myocardial contraction, as well as in stabilization of non-muscle cytoskeletal actin filaments. So far, LVN(C)-EA has predominantly been linked to pathogenic variants in MYH7. However, one sporadic LVN(C)-EA case with a de novo TPM1 variant has recently been described. We here report the first LVN(C)-EA family segregating a pathogenic TPM1 variant, further establishing the association between EA predisposition and TPM1-related LVN(C). Consequently, we recommend genetic testing for both MYH7 and TPM1 in patients or families in which LVN(C)/non-compaction and EA coincide.

Copy number variants in Ebstein anomaly.

BACKGROUND: Ebstein anomaly (EA) is a rare congenital defect characterized by apical displacement of the septal tricuspid leaflets and atrialization of the right ventricle. The etiology of EA is unclear; however, recurrence in families and the association of EA with genetic syndromes and copy number variants (CNVs) suggest a genetic component. OBJECTIVE: We performed a population-based study to search for recurrent and novel CNVs in a previously unreported set of EA cases. METHODS: We genotyped 60 EA cases identified from all live births (2,891,076) from selected California counties (1991-2010) using the Illumina HumanOmni2.5-8 array. We identified 38 candidate CNVs in 28 (46%) cases and prioritized and validated 11 CNVs based on the genes included. RESULTS: Five CNVs (41%) overlapped or were close to genes involved in early myocardial development, including NODAL, PDLIM5, SIX1, ASF1A and FGF12. We also replicated a previous association of EA with CNVs at 1p34.1 and AKAP12. Finally, we identified four CNVs overlapping or in close proximity to the transcription factors HES3, TRIM71, CUX1 and EIF4EBP2. CONCLUSIONS: This study supports the relationship of genetic factors to EA and demonstrates that defects in cardiomyocytes and myocardium differentiation may play a role. Abnormal differentiation of cardiomyocytes and how genetic factors contribute should be examined for their association with EA.

Familial Ebstein Anomaly: Whole Exome Sequencing Identifies Novel Phenotype Associated With FLNA.

BACKGROUND: Familial Ebstein anomaly is a rare form of congenital heart disease. We report 7 individuals among 2 generations of 1 family with Ebstein anomaly. This family was first reported in 1991 by Balaji et al in which family members were also reported to have a mild skeletal phenotype. The most likely mechanism of inheritance was concluded to be autosomal dominant. We sought to identify the genetic pathogenesis in this family using a next generation sequencing approach. METHODS AND RESULTS: Whole exome sequencing was performed in 2 cousins in this family using the Agilent SureSelect Human all Exon 51 Mb version 5 capture kit. Data were processed through an analytic in-house pipeline. Whole exome sequencing identified a missense mutation in FLNA (Filamin A), an actin-binding protein located at Xq28, mutations in which are associated with the skeletal phenotypes Frontometaphyseal dysplasia, Otopalatodigital, and Melnick-Needles syndrome, with X-linked periventricular nodular heterotopia and FG syndrome (Omim, 305450). Review of the phenotypes of those with the mutation in this family shows increased severity of the cardiac phenotype and associated skeletal features in affected males, consistent with X-linked inheritance. CONCLUSIONS: Although congenital heart disease is reported in families with mutations in FLNA, this is the first report of individuals being affected by Ebstein anomaly because of a mutation in this gene and details the concurrent skeletal phenotype observed in this family.

Ebstein's Anomaly: Genetics, Clinical Manifestations, and Management.

Ebstein's anomaly is uncommon. Genetic bases of this congenital heart defect may be related to the mutations in myosin heavy chain 7 and NKX2.5, among others. Asymptomatic patients with Ebstein's anomaly can be conservatively treated and kept under close follow-up, whereas surgical operation is indicated for those patients with evidence of right heart dilation and progressively impaired ventricular systolic function. A biventricular repair consisting of the reconstruction of a competent monocuspid tricuspid valve, right ventriculorrhaphy, subtotal atrial septal defect closure, and aggressive reduction atrioplasty is suitable for most patients, and 1.5-ventricular repair (bidirectional Glenn shunt) is indicated for patients with poor right ventricular function; by contrast, heart transplantation is used in patients with severe left ventricular dysfunction.

Genetic Variants in Isolated Ebstein Anomaly Implicated in Myocardial Development Pathways.

Ebstein anomaly (EA) is a rare heart defect in which the tricuspid valve is malformed and displaced. The tricuspid valve abnormalities can lead to backflow of blood from the right ventricle to the right atrium, preventing proper circulation of blood to the lungs. Although the etiology of EA is largely unresolved, increased prevalence of EA in those with a family history of congenital heart disease suggests EA has a genetic component. Copy number variants (CNVs) are a major source of genetic variation and have been implicated in a range of congenital heart defect phenotypes. We performed a systematic, genome-wide search for CNVs in 47 isolated EA cases using genotyping microarrays. In addition, we used a custom HaloPlex panel to sequence three known EA genes and 47 candidate EA genes. We identified 35 candidate CNVs in 24 (51%) EA cases. Rare sequence variants in genes associated with cardiomyopathy were identified in 11 (23%) EA cases. Two CNVs near the transcriptional repressor HEY1, a member of the NOTCH signaling pathway, were identified in three unrelated cases. All other candidate CNVs were each identified in a single case. At least 11 of 35 candidate CNVs include genes involved in myocardial development or function, including multiple genes in the BMP signaling pathway. We identified enrichment of gene sets involved in histone modification and cardiomyocyte differentiation, supporting the involvement of the developing myocardium in the etiology of EA. Gene set enrichment analysis also identified ribosomal RNA processing, a potentially novel pathway of altered cardiac development in EA. Our results suggest an altered myocardial program may contribute to abnormal tricuspid valve development in EA. Future studies should investigate abnormal differentiation of cardiomyocytes as a potential etiological factor in EA.

Ebstein anomaly, left ventricular non-compaction, and early onset heart failure associated with a de novo α-tropomyosin gene mutation.

Ebstein anomaly of the tricuspid valve (EA) can be associated with left ventricular non-compaction (LVNC), a rare congenital cardiomyopathy. We report a 2 year-old female with EA and severe tricuspid regurgitation, LVNC, pulmonary hypertension, and chronic biventricular systolic heart failure, who died during evaluation for cardiac transplantation. Gene panel testing revealed a heterozygous de novo missense mutation in TPM1, which encodes the cardiac sarcomeric thin filament protein α-tropomyosin. The c.475G>A variant results in a p.Asp159Asn substitution, altering a highly conserved residue predicted to be damaging to protein structure and function. TPM1 is the second gene linked to EA with LVNC in humans, implicating overlap in the molecular basis of structural and myopathic heart disease. © 2016 Wiley Periodicals, Inc.

[Altered Z-disks of myofibrils in the cardiomyocytes from patients with Ebstein's anomaly].

UNLABELLED: The aim of the study was to examine the peculiarities of the changes in Z-bands of myofibrils in the cardiomyocytes from patients with Ebstein's anomaly. MATERIAL AND METHODS: Electron microscopy assay of intraoperative biopsies of the right heart chambers in 41 patients aged from 9 months to 57 years was performed. RESULTS: Some patients exhibited Z-disk alterations of two types in individual cardiomyocytes, namely: local symmetrical bead-like expansions of Z-disks or longitudinal deposits of Z-material of different lengths along the myofibrils. Z-disk alterations were more common in atrial cardiomyocytes than in the ventricle. The presence of Z-disk alterations in the cardiomyocytes correlated with a number of clinical parameters. In particular, the occurrence of longitudinal deposits of Z-material in atrial cardiomyocytes directly correlated with the manifestation of the Wolff-Parkinson-White syndrome in the patients. CONCLUSIONS: Above characteristic ultrastructural changes in Z-bands of myofibrils in the cardiomyocytes from patients with Ebstein's anomaly have a certain similarity to Z-band diseases in skeletal muscle at sarcomeric protein gene mutations described in the literature, which suggests the mutations in the genes of proteins included in Z-bands of myofibrils in the cardiomyocytes from patients with Ebstein's anomaly.

Genetics of valvular heart disease.

Valvular heart disease is associated with significant morbidity and mortality and often the result of congenital malformations. However, the prevalence is increasing in adults not only because of the growing aging population, but also because of improvements in the medical and surgical care of children with congenital heart valve defects. The success of the Human Genome Project and major advances in genetic technologies, in combination with our increased understanding of heart valve development, has led to the discovery of numerous genetic contributors to heart valve disease. These have been uncovered using a variety of approaches including the examination of familial valve disease and genome-wide association studies to investigate sporadic cases. This review will discuss these findings and their implications in the treatment of valvular heart disease.

Thiamine-responsive megaloblastic anemia syndrome with Ebstein anomaly: a case report.

UNLABELLED: Thiamine-responsive megaloblastic anemia (TRMA) or Roger syndrome is a rare autosomal recessive disorder characterized by the occurrence of multiple clinical manifestations including megaloblastic anemia, diabetes mellitus, and sensorineural deafness. A few patients have been also described with congenital cardiac malformations. The patients usually respond to treatment with pharmacological doses of thiamine. Mutations in the SLC19A2 gene, located at chromosome 1q24.2, are responsible for this syndrome. Here, we present two new Iranian TRMA patients who were homozygous for c.697C > T mutation in the SLC19A2 gene. On follow-up, one of the patients showed Ebstein anomaly. CONCLUSION: The present study confirms the variability of the clinical manifestations caused by the same mutation within patients with TRMA syndrome. Therefore, follow-up of the affected children should be considered.

Familial ebstein anomaly, left ventricular hypertrabeculation, and ventricular septal defect associated with a MYH7 mutation.

Ebstein anomaly is a rare congenital heart defect that most often occurs sporadically within a kindred. Familial cases, although reported, are uncommon. At this time, the genetic etiology of Ebstein anomaly is not fully elucidated. Here, we describe clinical and molecular investigations of a rare case of familial Ebstein anomaly in association with a likely pathogenic mutation of the MYH7 gene. The severity of presentation varies, and Ebstein anomaly can be observed in association with such other heart defects as ventricular septal defect and left ventricular (LV) hypertrabeculation, as seen in our family of study. In our family of study, the 31-year-old father and four of his children have been diagnosed with Ebstein anomaly. Genetic testing revealed that the father was heterozygous for the Glu1220del variant detected in exon 27 of the MYH7 gene. The MYH7 gene encodes the ß-myosin heavy chain and is expressed in cardiac muscle. DNA sequencing of three of his affected children confirmed that they carried the same variant while the fourth affected child was not available for testing. This is the first report of familial Ebstein anomaly associated with the Glu1220del mutation of the MYH7 gene. The mutation segregates with disease in a family with autosomal dominant transmission of congenital heart defects including Ebstein anomaly and other associated cardiovascular defects including LV hypertrabeculation and ventricular septal defect.

Ebstein anomaly associated with left ventricular noncompaction: an autosomal dominant condition that can be caused by mutations in MYH7.

Left ventricular noncompaction (LVNC) is a relatively common genetic cardiomyopathy, characterized by prominent trabeculations with deep intertrabecular recesses in mainly the left ventricle. Although LVNC often occurs in an isolated entity, it may also be present in various types of congenital heart disease (CHD). The most prevalent CHD in LVNC is Ebstein anomaly, which is a rare form of CHD characterized by apical displacement and partial fusion of the septal and posterior leaflet of the tricuspid valve with the ventricular septum. Several reports of sporadic as well as familial cases of Ebstein anomaly associated with LVNC have been reported. Recent studies identified mutations in the MYH7 gene, encoding the sarcomeric ß-myosin heavy chain protein, in patients harboring this specific phenotype. Here, we will review the association between Ebstein anomaly, LVNC and mutations in MYH7, which seems to represent a subtype of Ebstein anomaly with autosomal dominant inheritance and variable penetrance.

Cardiac anomalies in individuals with the 18q deletion syndrome; report of a child with Ebstein anomaly and review of the literature.

Individuals with the 18q deletion syndrome are presented with various clinical characteristics, including cardiac anomalies in 24-36% of the reported cases. Nonetheless, genotype-phenotype correlations for cardiac anomalies in the 18q deletion syndrome have rarely been reported. We report on two girls with a terminal 18q deletion, one in whom an Ebstein anomaly and Wolff-Parkinson-White syndrome were detected and the other with multiple valve stenosis and a ventricular septal defect. The genotype and cardiac abnormalities of these girls and 17 other individuals with a de novo 18qter deletion reported in the literature are reviewed. All 19 individuals shared a small overlapping deletion region between 18q22.3q23. The most common cardiac defects detected were pulmonary valve anomalies and atrial septal defects. Ebstein anomaly, a rare cardiac malformation, was diagnosed in two individuals. Additional molecularly based genotype-phenotype studies are needed in order to pinpoint candidate genes within this region that contribute to normal cardiac development. A careful cardiac evaluation consisting of physical examination, ECG and ultrasound examination should be performed in all individuals diagnosed with the 18q deletion syndrome.