Brugada Syndrome and Pulmonary Atresia with Intact Interventricular Septum: Fortuitous Finding or New Genetic Connection?

Brugada syndrome is a rare arrhythmogenic syndrome associated mainly with pathogenic variants in the SCN5A gene. Right ventricle outflow tract fibrosis has been reported in some cases of patients diagnosed with Brugada syndrome. Pulmonary atresia with an intact ventricular septum is characterized by the lack of a functional pulmonary valve, due to the underdevelopment of the right ventricle outflow tract. We report, for the first time, a 4-year-old boy with pulmonary atresia with an intact ventricular septum who harbored a pathogenic de novo variant in SCN5A, and the ajmaline test unmasked a type-1 Brugada pattern. We suggest that deleterious variants in the SCN5A gene could be implicated in pulmonary atresia with an intact ventricular septum embryogenesis, leading to overlapping phenotypes.

Patient-specific iPSC-derived cardiomyocytes reveal variable phenotypic severity of Brugada syndrome.

BACKGROUND: Brugada syndrome (BrS) is a cardiac channelopathy that can result in sudden cardiac death (SCD). SCN5A is the most frequent gene linked to BrS, but the genotype-phenotype correlations are not completely matched. Clinical phenotypes of a particular SCN5A variant may range from asymptomatic to SCD. Here, we used comparison of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) derived from a SCN5A mutation-positive (D356Y) BrS family with severely affected proband, asymptomatic mutation carriers (AMCs) and healthy controls to investigate this variation. METHODS: 26 iPSC lines were generated from skin fibroblasts using nonintegrated Sendai virus. The generated iPSCs were differentiated into cardiomyocytes using a monolayer-based differentiation protocol. FINDINGS: D356Y iPSC-CMs exhibited increased beat interval variability, slower depolarization, cardiac arrhythmias, defects of Na+ channel function and irregular Ca2+ signaling, when compared to controls. Importantly, the phenotype severity observed in AMC iPSC-CMs was milder than that of proband iPSC-CMs, an observation exacerbated by flecainide. Interestingly, the iPSC-CMs of the proband exhibited markedly decreased Ca2+ currents in comparison with control and AMC iPSC-CMs. CRISPR/Cas9-mediated genome editing to correct D356Y in proband iPSC-CMs effectively rescued the arrhythmic phenotype and restored Na+ and Ca2+ currents. Moreover, drug screening using established BrS iPSC-CM models demonstrated that quinidine and sotalol possessed antiarrhythmic effects in an individual-dependent manner. Clinically, venous and oral administration of calcium partially reduced the malignant arrhythmic events of the proband in mid-term follow-up. INTERPRETATION: Patient-specific and genome-edited iPSC-CMs can recapitulate the varying phenotypic severity of BrS. Our findings suggest that preservation of the Ca2+ currents might be a compensatory mechanism to resist arrhythmogenesis in BrS AMCs. FUNDING: National Key R&D Program of China (2017YFA0103700), National Natural Science Foundation of China (81922006, 81870175), Natural Science Foundation of Zhejiang Province (LD21H020001, LR15H020001), National Natural Science Foundation of China (81970269), Key Research and Development Program of Zhejiang Province (2019C03022) and Natural Science Foundation of Zhejiang Province (LY16H020002).

Transmembrane protein 168 mutation reduces cardiomyocyte cell surface expression of Nav1.5 through αB-crystallin intracellular dynamics.

Transmembrane protein 168 (TMEM168) was found to be localized on the nuclear membrane. A heterozygous mutation (c.1616G>A, p. R539Q) in TMEM168 was identified in patients with Brugada syndrome. This mutation reduced expression of cardiomyocyte sodium channel Nav1.5 via Nedd4-2 E3 ubiquitin ligase-induced ubiquitination and degradation. However, the detailed molecular mechanism provoked by the TMEM168 mutant remains unclear. Here, we demonstrated that small heat shock protein αB-crystallin, which can bind to Nav1.5 and Nedd4-2 and interfere with the association of both proteins, was strongly recruited from the cell surface to the perinuclear region because of the much higher affinity of αB-crystallin with the TMEM168 mutant than with wild-type TMEM168. Following knockdown of αB-crystallin in HL-1 cardiomyocytes, the interaction of Nav1.5 with Nedd4-2 was increased, despite the reduced expression of Nav1.5. Moreover, reduction of Nav1.5 expression by αB-crystallin knockdown was rescued in the presence of a proteasome inhibitor MG-132, suggesting the importance of the αB-crystallin-modulated ubiquitin-proteasome system for the stability of Nav1.5 expression. Collectively, the balance of molecular interactions among Nav1.5, Nedd4-2 and αB-crystallin plays a role in the regulation of cardiomyocyte cell surface expression of Nav1.5, and the TMEM168 mutant disturbs this balance, resulting in a decrease in Nav1.5 expression.

iPSC-Cardiomyocyte Models of Brugada Syndrome-Achievements, Challenges and Future Perspectives.

Brugada syndrome (BrS) is an inherited cardiac arrhythmia that predisposes to ventricular fibrillation and sudden cardiac death. It originates from oligogenic alterations that affect cardiac ion channels or their accessory proteins. The main hurdle for the study of the functional effects of those variants is the need for a specific model that mimics the complex environment of human cardiomyocytes. Traditionally, animal models or transient heterologous expression systems are applied for electrophysiological investigations, each of these models having their limitations. The ability to create induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), providing a source of human patient-specific cells, offers new opportunities in the field of cardiac disease modelling. Contemporary iPSC-CMs constitute the best possible in vitro model to study complex cardiac arrhythmia syndromes such as BrS. To date, thirteen reports on iPSC-CM models for BrS have been published and with this review we provide an overview of the current findings, with a focus on the electrophysiological parameters. We also discuss the methods that are used for cell derivation and data acquisition. In the end, we critically evaluate the knowledge gained by the use of these iPSC-CM models and discuss challenges and future perspectives for iPSC-CMs in the study of BrS and other arrhythmias.

Fibrosis and Conduction Abnormalities as Basis for Overlap of Brugada Syndrome and Early Repolarization Syndrome.

Brugada syndrome and early repolarization syndrome are both classified as J-wave syndromes, with a similar mechanism of arrhythmogenesis and with the same basis for genesis of the characteristic electrocardiographic features. The Brugada syndrome is now considered a conduction disorder based on subtle structural abnormalities in the right ventricular outflow tract. Recent evidence suggests structural substrate in patients with the early repolarization syndrome as well. We propose a unifying mechanism based on these structural abnormalities explaining both arrhythmogenesis and the electrocardiographic changes. In addition, we speculate that, with increasing technical advances in imaging techniques and their spatial resolution, these syndromes will be reclassified as structural heart diseases or cardiomyopathies.

Incidence and etiology of sudden cardiac arrest in Koreans: A cohort from the national health insurance service database.

The incidence of sudden cardiac arrest (SCA) in Asians is lower than that seen in Western populations, but there are few available data on the incidence and associated cardiac etiology of SCA in Asians. From 2002 to 2013, patients with SCA were analyzed using a cohort from the South Korean National Health Insurance Service (NHIS) coded database. Sudden unexplained death syndrome (SUDS) was defined as cryptogenic arrest, excluding that of non-cardiac origin, coronary artery disease (CAD), cardiomyopathy (CM), and valvular heart disease. During the 12-year study period, 5,973 patients (0.53%) from the total cohort of 1,125,691 had a cardiac arrest code. The overall incidence of arrest was 48.7 per 100,000 person-years (95% CI 16.6-18.0). The incidence of primary SCA excluding those of non-cardiac origin was 16.1 per 100,000 person-years (95% CI 15.4-16.8). It was higher in males than in females (18.1 vs. 14.1 per 100,000 person-years). CAD was the most common cause of SCA (59.4%), and followed by CM (13.9%). SUDS accounted for 14.7% of SCA events. The risk of SCA had increased gradually from over 25 years old. Heart failure, atrial fibrillation and hypertension are major factors associated with SCA incidence. Our findings outline epidemiologic data for SCA and the proportion of associated cardiac etiology leads SCA in a large population.

Relationship between sodium channel function and clinical phenotype in SCN5A variants associated with Brugada syndrome.

The identification of a pathogenic SCN5A variant confers an increased risk of conduction defects and ventricular arrhythmias (VA) in Brugada syndrome (BrS). However, specific aspects of sodium channel function that influence clinical phenotype have not been defined. A systematic literature search identified SCN5A variants associated with BrS. Sodium current (INa ) functional parameters (peak current, decay, steady-state activation and inactivation, and recovery from inactivation) and clinical features (conduction abnormalities [CA], spontaneous VA or family history of sudden cardiac death [SCD], and spontaneous BrS electrocardiogram [ECG]) were extracted. A total of 561 SCN5A variants associated with BrS were identified, for which data on channel function and clinical phenotype were available in 142. In the primary analysis, no relationship was found between any aspect of channel function and CA, VA/SCD, or spontaneous BrS ECG pattern. Sensitivity analyses including only variants graded pathogenic or likely pathogenic suggested that reduction in peak current and positive shift in steady-state activation were weakly associated with CA and VA/SCD, although sensitivity and specificity remained low. The relationship between in vitro assessment of channel function and BrS clinical phenotype is weak. The assessment of channel function does not enhance risk stratification. Caution is needed when extrapolating functional testing to the likelihood of variant pathogenicity.

Mechanisms of Arrhythmias in the Brugada Syndrome.

Arrhythmias in Brugada syndrome patients originate in the right ventricular outflow tract (RVOT). Over the past few decades, the characterization of the unique anatomy and electrophysiology of the RVOT has revealed the arrhythmogenic nature of this region. However, the mechanisms that drive arrhythmias in Brugada syndrome patients remain debated as well as the exact site of their occurrence in the RVOT. Identifying the site of origin and mechanism of Brugada syndrome would greatly benefit the development of mechanism-driven treatment strategies.

A high number of 'natural' mitochondrial DNA polymorphisms in a symptomatic Brugada syndrome type 1 patient.

Brugada syndrome (BrS) is a rare genetic arrhythmic disorder with a complex model of transmission. At least 20 different genes have been identified as BrS-causal or susceptibility genes. Of these, SCN5A is the most frequently mutated. Coregulation of different mutations or genetic variants, including mitochondrial DNA (mtDNA), may contribute to the clinical phenotype of the disease. In thepresent study, we analysed the mitochondrial genome of a symptomatic BrS type 1 patient to investigate a possible mitochondrial involvement recently found in the arrhytmogenic diseases. No pathogenic mutation was identified; however, a high number of singlenucleotide polymorphisms were found (n=21) and some of them were already been reported in molecular autopsy case for sudden death.The results reported here further support our hypothesis on the potential role of mtDNA polymorphisms in mitochondrial dysfunction, which may represent a risk factor for arrhythmogenic disease.

GSTM3 variant is a novel genetic modifier in Brugada syndrome, a disease with risk of sudden cardiac death.

BACKGROUND: Brugada syndrome (BrS) is a rare inherited disease causing sudden cardiac death (SCD). Copy number variants (CNVs) can contribute to disease susceptibility, but their role in Brugada syndrome (BrS) is unknown. We aimed to identify a CNV associated with BrS and elucidated its clinical implications. METHODS: We enrolled 335 unrelated BrS patients from 2000 to 2018 in the Taiwanese population. Microarray and exome sequencing were used for discovery phase whereas Sanger sequencing was used for the validation phase. HEK cells and zebrafish were used to characterize the function of the CNV variant. FINDINGS: A copy number deletion of GSTM3 (chr1:109737011-109737301, hg38) containing the eighth exon and the transcription stop codon was observed in 23.9% of BrS patients versus 0.8% of 15,829 controls in Taiwan Biobank (P < 0.001), and 0% in gnomAD. Co-segregation analysis showed that the co-segregation rate was 20%. Patch clamp experiments showed that in an oxidative stress environment, GSTM3 down-regulation leads to a significant decrease of cardiac sodium channel current amplitude. Ventricular arrhythmia incidence was significantly greater in gstm3 knockout zebrafish at baseline and after flecainide, but was reduced after quinidine, consistent with clinical observations. BrS patients carrying the GSTM3 deletion had higher rates of sudden cardiac arrest and syncope compared to those without (OR: 3.18 (1.77-5.74), P<0.001; OR: 1.76 (1.02-3.05), P = 0.04, respectively). INTERPRETATION: This GSTM3 deletion is frequently observed in BrS patients and is associated with reduced INa, pointing to this as a novel potential genetic modifier/risk predictor for the development of the electrocardiographic and arrhythmic manifestations of BrS. FUNDING: This work was supported by the Ministry of Science and Technology (107-2314-B-002-261-MY3 to J.M.J. Juang), and by grants HL47678, HL138103 and HL152201 from the National Institutes of Health to CA.

Validation and Disease Risk Assessment of Previously Reported Genome-Wide Genetic Variants Associated With Brugada Syndrome: SADS-TW BrS Registry.

BACKGROUND: Brugada syndrome (BrS) is an oligogenic arrhythmic disease with increased risk of sudden cardiac arrest. Several BrS or ECG traits-related single-nucleotide polymorphisms (SNPs) were identified through previous genome-wide association studies in white patients. We aimed to validate these SNPs in BrS patients in the Taiwanese population, assessing the cumulative effect of risk alleles and the BrS-polygenic risk score in predicting cardiac events. METHODS: We genotyped 190 unrelated BrS patients using the TWB Array, and Taiwan Biobank was used as controls. SNPs not included in the array were imputed by IMPUTE2. Cox proportional hazards model was used to evaluate the associations between each particular SNP, the collective BrS-polygenic risk score, and clinical outcomes. RESULTS: Of the 88 previously reported SNPs, 22 were validated in Taiwanese BrS patients (P<0.05). Of the 22 SNPs, 2 (rs10428132 and rs9388451) were linked with susceptibility to BrS, 10 were SNPs previously reaching genome-wide significance, and 10 were SNPs associated with ECG traits. For the 3 most commonly reported SNPs, disease risk increased consistently with the number of risk alleles (odds ratio, 3.54; Ptrend=1.38×10-9 for 5 risk alleles versus 1). Similar patterns were observed in both SCN5A mutation+ (odds ratio, 3.66; Ptrend=0.049) and SCN5A mutation- (odds ratio, 3.75; Ptrend=8.54×10-9) subgroups. Furthermore, BrS patients without SCN5A mutations had more risk alleles than BrS patients with SCN5A mutations regardless of the range of polygenic risk scores. Three SNPs (rs4687718, rs7784776, and rs2968863) showed significant associations with the composite outcome (sudden cardiac arrest plus syncope, hazard ratio, 2.13, 1.48, and 0.41; P=0.02, 0.006, and 0.008, respectively). CONCLUSIONS: Our findings suggested that some SNPs associated with BrS or ECG traits exist across multiple populations. The cumulative risk of the BrS-related SNPs is similar to that in white BrS patients, but it appears to correlate with the absence of SCN5A mutations.

Lamotrigine induced Brugada-pattern in a patient with genetic epilepsy associated with a novel variant in SCN9A.

BACKGROUND: A 30-year-old man presented with intellectual disability associated with epilepsy. The epilepsy was initially treated with sodium valproate and since he was 28 years-old with lamotrigine. With the addition of lamotrigine, a pattern of Brugada syndrome appeared on the electrocardiogram. The family history was positive for epilepsy from the mothers side, who had never been treated with lamotrigine. OBJECTIVE: Determine the genetic cause of the intellectual disability, epilepsy and Brugada syndrome of the patient and try to establish a possible correlation between the genetic background and the Brugada syndrome pattern under lamotrigine treatment. METHODS: A standard karyotype, array comparative genomic hybridization and two different NGS panels have done to the index case to identify the genetic causes of the intellectual disability, epilepsy and Brugada syndrome pattern. RESULTS: Genetic analyses in the family identified a de novo duplication of 1.3 Mb in 8p21.3 as well as two novel heterozygous rare variants in SCN9A and AKAP9 genes, both inherited from the mother. CONCLUSION: We hypothesize that in this family the SCN9A variant was responsible for the epileptic syndrome. In addition, given that SCN9A is lightly expressed in the heart tissue, we postulate that this SCN9A variant, alone or in combination with AKAP9 variant, might be responsible for the Brugada pattern when challenged by lamotrigine.

Clinical and Molecular Data Define a Diagnosis of Arrhythmogenic Cardiomyopathy in a Carrier of a Brugada-Syndrome-Associated PKP2 Mutation.

Plakophilin-2 (PKP2) is the most frequently mutated desmosomal gene in arrhythmogenic cardiomyopathy (ACM), a disease characterized by structural and electrical alterations predominantly affecting the right ventricular myocardium. Notably, ACM cases without overt structural alterations are frequently reported, mainly in the early phases of the disease. Recently, the PKP2 p.S183N mutation was found in a patient affected by Brugada syndrome (BS), an inherited arrhythmic channelopathy most commonly caused by sodium channel gene mutations. We here describe a case of a patient carrier of the same BS-related PKP2 p.S183N mutation but with a clear diagnosis of ACM. Specifically, we report how clinical and molecular investigations can be integrated for diagnostic purposes, distinguishing between ACM and BS, which are increasingly recognized as syndromes with clinical and genetic overlaps. This observation is fundamentally relevant in redefining the role of genetics in the approach to the arrhythmic patient, progressing beyond the concept of "one mutation, one disease", and raising concerns about the most appropriate approach to patients affected by structural/electrical cardiomyopathy. The merging of genetics, electroanatomical mapping, and tissue and cell characterization summarized in our patient seems to be the most complete diagnostic algorithm, favoring a reliable diagnosis.

Identification of transmembrane protein 168 mutation in familial Brugada syndrome.

Brugada syndrome (BrS) is an inherited channelopathy responsible for almost 20% of sudden cardiac deaths in patients with nonstructural cardiac diseases. Approximately 70% of BrS patients, the causative gene mutation(s) remains unknown. In this study, we used whole exome sequencing to investigate candidate mutations in a family clinically diagnosed with BrS. A heterozygous 1616G>A substitution (R539Q mutation) was identified in the transmembrane protein 168 (TMEM168) gene of symptomatic individuals. Similar to endogenous TMEM168, both TMEM168 wild-type (WT) and mutant proteins that were ectopically induced in HL-1 cells showed nuclear membrane localization. A significant decrease in Na+ current and Nav 1.5 protein expression was observed in HL-1 cardiomyocytes expressing mutant TMEM168. Ventricular tachyarrhythmias and conduction disorders were induced in the heterozygous Tmem168 1616G>A knock-in mice by pharmacological stimulation, but not in WT mice. Na+ current was reduced in ventricular cardiomyocytes isolated from the Tmem168 knock-in heart, and Nav 1.5 expression was also impaired. This impairment was dependent on increased Nedd4-2 binding to Nav 1.5 and subsequent ubiquitination. Collectively, our results show an association between the TMEM168 1616G>A mutation and arrhythmogenesis in a family with BrS.

Two Forms of Monomorphic Ventricular Tachycardia in a Patient with Brugada Syndrome.

We herein report a 47-year-old man with relapsing polychondritis who developed monomorphic ventricular tachycardia (VT). His electrocardiogram in sinus rhythm showed a coved-type pattern, and there was no evidence of structural cardiac disease; therefore, he was diagnosed with Brugada syndrome. An electrophysiological study revealed a prolonged His-ventricular interval at the baseline. Two forms of VT were induced, which were shown to be bundle branch reentrant VT. A diagnosis of Brugada syndrome should not be ruled out in patients with monomorphic VTs, especially those with conduction abnormalities.

Brugada syndrome with SCN5A mutations exhibits more pronounced electrophysiological defects and more severe prognosis: A meta-analysis.

Whether the presence of SCN5A mutation is a predictor of BrS risk remains controversial, and patient selection bias may have weakened previous findings. Therefore, we performed this study to clarify the clinical characteristics and outcomes of BrS probands with SCN5A mutations. We systematically retrieved eligible studies published through October 2018. A total of 17 studies enrolling 1780 BrS patients were included. Overall, our results found that compared with BrS patients without SCN5A mutations, patients with SCN5A mutations exhibited a younger age at the onset of symptoms and higher rate of the spontaneous type-1 electrocardiogram pattern, more pronounced conduction or repolarization abnormalities, and increased atrial vulnerability. In addition, the presence of SCN5A mutations was associated with an elevated risk of major arrhythmic events in both Asian (odds ratio [OR] = 1.82, 95% confidence interval [CI] 1.07-3.11; P = .03) and Caucasian (OR = 2.24, 95% CI 1.02-4.90; P = .04) populations. In conclusions, patients with SCN5A mutations exhibit more pronounced electrophysiological defects and more severe prognosis. Clinicians should be cautious when utilizing genetic testing for risk stratification or treatment guidance before determining whether the causal relationship regarding SCN5A mutation status is an independent predictor of risk.

Novel SCN5A p.W697X Nonsense Mutation Segregation in a Family with Brugada Syndrome.

Brugada syndrome (BrS) is marked by an elevated ST-segment elevation and increased risk of sudden cardiac death. Variants in the SCN5A gene are considered to be molecular confirmation of the syndrome in about one third of cases, while the genetics remain a mystery in about half of the cases, with the remaining cases being attributed to variants in any of a number of genes. Before research models can be developed, it is imperative to understand the genetics in patients. Even data from humans is complicated, since variants in the most common gene in BrS, SCN5A, are associated with a number of pathologies, or could even be considered benign, depending on the variant. Here, we provide crucial human data on a novel NM_198056.2:c.2091G>A (p.Trp697X) point-nonsense heterozygous variant in the SCN5A gene, as well as its segregation with BrS. The results herein suggest a pathogenic effect of this variant. These results could be used as a stepping stone for functional studies to better understand the molecular effects of this variant in BrS.

Pharmacotherapy in inherited and acquired ventricular arrhythmia in structurally normal adult hearts.

Introduction: Ventricular arrhythmias are often seen in association with structural heart disease. However, approximately a tenth of affected patients have apparently normal hearts, where such arrhythmias typically occur in young patients, are sometimes inherited and can occasionally lead to sudden cardiac death (SCD). Over the past two decades, increased understanding of the underlying pathophysiology resulted in improved targeted pharmacological therapy.Areas covered: This article reviews current knowledge regarding drug therapy for inherited arrhythmia syndromes (Brugada, early repolarization, long QT and short QT syndromes, and catecholaminergic polymorphic ventricular tachycardia), and acquired arrhythmias (idiopathic ventricular fibrillation, short-coupled torsade de pointes, outflow tract ventricular tachycardia, idiopathic left, papillary muscle and annular ventricular tachycardias).Expert opinion: In inherited arrhythmia syndromes, appropriate clinical and genetic diagnoses followed by proper selection and dosing of antiarrhythmic drugs are of utmost importance to prevent SCD, most often without the need of implantable cardioverter-defibrillators. In acquired arrhythmias, appropriate pharmacotherapy in selected patients can also provide symptomatic relief and avoid the need for invasive therapy. Further research is needed to develop novel antiarrhythmic drugs or targeted therapy to increase efficacy and limit side effects.