Long QT Syndrome Research Articles

The endocannabinoid ARA-S acts as an endogenous rescuing factor of Kv7.1/KCNE1 carrying Long QT syndrome mutations and paves the way for drug development strategies

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – EU funding. Main funding source(s): European Union’s Horizon 2020 research and innovation program Long QT Syndrome (LQTS) is a cardiac arrhythmia characterized by delayed ventricular repolarization. LQTS occurs when there are loss-of-function mutations in genes encoding for voltage-gated channels and their beta subunits that contribute to the cardiac action potential. Among them, the most mutated channel is Kv7.1/KCNE1, which encodes for the slowly activating component of the delayed rectifier K+ current (IKs). Families with inherited mutations in the Kv7.1/KCNE1 exhibit inconsistent correlations between genotypes and phenotypes making the clinical management of LQTS challenging. A possible strategy to help guide clinical approaches involves exploring endogenous compounds capable of modulating the Kv7.1/KCNE1 channel function, acting as potential disease modifiers. Using the two-electrode voltage clamp technique, we previously found that the endogenous endocannabinoid-like compound ARA-S, which has an arachidonic acid tail and a serine head group, facilitates activation of the wild-type Kv7.1/KCNE1 channel expressed in Xenopus oocytes. This is seen as a left shift in the voltage dependence of channel opening and an increase in conductance. Moreover, we showed that ARA-S restores a physiological QT interval in isolated guinea pig hearts suffering from drug-induced LQTS. Hence, ARA-S is an interesting endogenous Kv7.1/KCNE1 channel modulator that may influence disease severity. However, it is uncertain whether ARA-S also activates the Kv7.1/KCNE1 when it carries LQTS-associated mutations. Therefore, the current study involves studying the effect of ARA-S on mutated Kv7.1/KCNE1 channels using two-electrode voltage clamp electrophysiology. We find that ARA-S facilitates the activation of all tested LQTS mutants. However, the magnitude of the effect varies among them, suggesting that the compound´s effect is dependent on the genetic background. To understand why ARA-S has different effects, we use computational simulation approaches to study how mutations in the putative ARA-S binding sites change ARA-S efficacy. This understanding aids in developing new substances that can activate different mutations, by, for example, modifying the ARA-S tail to enhance its efficacy. We also try to understand the concentration of ARA-S required to restore a wild-type-like function of each mutant, considering the mutant's intrinsic behavior and the apparent affinity of ARA-S for the specific mutant. Altogether, this work finds ARA-S to be an efficient activator of LQTS-associated mutant Kv7.1/KCNE1 channels and is a first step towards understanding how the genetic background determines the effect of this endogenous modulator.

In vivo calcium and voltage mapping of the zebrafish heart

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Research Foundation Flanders (FWO) Purpose Inherited cardiac arrhythmias are characterized by a disturbance in the normal electrical functioning of the heart due to a dysfunction of cardiac ion channels, accessory proteins and/or structural components. This complex process is difficult to model in vitro, while in vivo mouse models differ substantially from humans with regards to cardiac electrophysiology. Thus, there is a need for alternative disease models in cardiac arrhythmia research. The zebrafish heart shows remarkable similarity to humans in its electrocardiogram and action potential (AP) morphology. Due to the optical translucency of zebrafish larvae, it is possible visualize the cardiac calcium and voltage dynamics across their entire hearts in vivo by use of genetically encoded calcium and voltage indicators (GECI and GEVI). We combined the fast kinetics of a next generation voltage reporter (Ace2N-mNeon) with a spectrally compatible calcium sensor (R-GECO) to create a sensitive high throughput method for in vivo characterization of the cardiac AP. Methods Ace2N-mNeon and R-GECO were expressed under control of a myocardial specific promoter via the Tol2kit system. The effect of sensor expression on normal physiology was assessed by patch clamp experiments. Fluorescence recordings were obtained with a Leica SP8 DLS microscope and a custom-built light sheet microscope. Our zebrafish sensor line was validated by experiments with quinidine drug exposure, as well as a novel model for long QT syndrome (LQTS) with a knock-in (KI) mutation in the cacna1c gene. Results Sensor expression did not significantly alter cardiac electrophysiology. The tg(myl7:Ace2N-mNeon/R-GECO) line demonstrated alterations in the heart rate, depolarization and repolarization induced by quinidine exposure. We observed a significantly prolonged ventricular repolarization in the LQTS KI line. Three-dimensional optical maps of AP characteristics across the entire zebrafish heart exhibited the physiological characteristics of the zebrafish cardiac electrophysiology, with activation initiated at the atrial venous pole and propagated towards the ventricular outflow tract, prolonged repolarization in the ventricles compared to the atria and slowed depolarization in the atrio-ventricular canal. Conclusion By expressing Ace2N-mNeon and R-GECO in the zebrafish heart, we were able to accurately detect drug- and mutation induced alterations in the cardiac AP, as well as AP characteristics across the entire heart, without compromising normal cardiac electrophysiology. Our transgenic line will provide a promising assay for future studies of cardiac arrhythmia.

Maturing hiPSC-cardiomyocytes in cardiac microtissues for accurate preclinical in vitro modelling of long-QT syndrome type 1

Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): reNEW - Novo Nordisk Foundation Center for Stem Cell Medicine Background Long-QT syndrome type 1 (LQT1), a cardiac arrhythmia often leading to sudden cardiac death, is due to mutations in KCNQ1 gene. Human iPSC-derived cardiomyocytes (hiPSC-CMs) have been widely used to model LQT1, promoting the concept of their use in preclinical studies. However, KCNQ1 locus is subjected to developmentally regulated genomic imprinting in the heart: in fetal cardiomyocytes the paternal allele is repressed, while both alleles become expressed after birth. Since hiPSC-CMs are typically immature fetal-like cells, residual imprinting may affect evaluation of KCNQ1 mutations in this system. Purpose This study wants to address the imprinting status of KCNQ1 in hiPSC-CMs and propose maturing hiPSC-CMs as a solution for reliable LQT1 modelling. Methods We derived hiPSC-CMs from a LQT1 patient carrying a heterozygous KCNQ1 mutation on the paternal allele and compared their electrical properties with the isogenic corrected hiPSC-CMs by patch clamp. We analysed KCNQ1 expression by ddPCR and DNA methylation. We used three-dimensional cardiac microtissues (MTs) composed by hiPSC-CMs, hiPSC-derived cardiac fibroblasts and endothelial cells to induce cardiomyocte maturation. Results LQT1 and isogenic corrected hiPSC-CMs showed no difference in action potential duration (APD). We hypothesized that residual KCNQ1 imprinting in hiPSC-CMs due to their immaturity was masking the phenotype. When we analyzed different hiPSC-CM lines, we found in all an unbalanced KCNQ1 allelic expression, with paternal allele accounting for 5-15% of the total. We then investigated whether maturing hiPSC-CMs could remove KCNQ1 imprinting. We included hiPSC-CMs in MTs, as this system previously showed to promote functional maturation and upregulation of KCNQ1. Here, we observed a substantial increased expression of the paternal allele and a reduction in the expression of KCNQ1OT1, the long non-coding RNA inducing paternal allele repression. LQT1 hiPSC-CMs dissociated from MTs showed prolonged APD compared to the corrected line. We also demonstrated that this was due to the reduction of the ionic current mediated by KCNQ1 (IKs) in LQT1 hiPSC-CMs compared to the corrected controls, thus revealing the mutation causative effect. Conclusions Our findings show that residual KCNQ1 imprinting in immature hiPSC-CMs can lead to underestimate (or overestimate) functional effects of pathological variants based on the parental allele that carries the mutation. This is overcome by maturation of hiPSC-CMs in tri-cellular cardiac microtissue which promotes KCNQ1 bi-allelic expression and allow dissecting mutation mechanisms. This study brings attention to the epigenetic regulation of hiPSC models and demonstrates the utility of using matured hiPSC-CMs as in vitro preclinical model for cardiac arrhythmias.

Current real-world evidence of genetic testing diagnostic yield in a large cohort of long QT syndrome

Abstract Background Diagnostic yield in long QT syndrome (LQTS) ranges between 30-70%, depending on the studied population. LQTS can lead to an unexpected sudden cardiac death. For this reason and due to its wider current availability, genetic testing in patients with suspicion of LQTS has spread. Purpose The aim of this study is to investigate the genetic testing yield among a large cohort of heterogeneous index cases with LQTS phenotype. Methods Diagnostic yield was restrospectively evaluated in a cohort of index cases referred to our laboratory for genetic study with LQTS phenotype. Genetic study was performed with customized libraries through next generation sequencing (NGS), which included copy number variations (CNVs) analysis. Patients with LQTS phenotype studied both with "long QT panels" and with broader cardiovascular panels were included, but those with additional cardiac phenotype (cardiomyopathies, heart defects or catecholaminergic polymorphic ventricular tachycardia) were excluded. Positive genetic reports were considered when a likely pathogenic (LP) or pathogenic (P) variant could explain patient´s phenotype. Inconclusive reports were described when a variant considered risk factor (RF) or a variant of uncertain significance variant (VUS) or a hot-VUS was identified in a clinically relevant gene. Results 1183 index cases were genotyped for LQTS phenotype. Only 164 index cases (13,8%) were studied with broader panels which included between 77 and 368 genes. The rest (n=1019) were studied with LQTS panels (either with small panels which included KCNQ1, KCNH2, KCNE1, KCNE2, SCN5A, KCNJ2, CACNA1C and with up to 36 genes-QT panels). Around 27% (n=321) of the index cases had a positive result. They carried LP/P variants in the following genes: KCNQ1 (n=166, 52%), KCNH2 (n=112), SCN5A (n=25), RYR2 (n=5), CACNA1C (n=3), HCN4 (n=4), KCNJ2 (n=2), KCNE2 (n=2) and CALM2 (n=1). 53,3% of the reports were negative. However, there were some incidental findings: 2 index cases carried P variants in LDLR, 2 carried P variants in PKP2, 2 carried a P variant in MYBPC3 and 1 index case each carried a P variant in DES, TTN and DSG2. From the 231 inconclusive reports, 37 index cases carried the RF variant in KCNE1 (p.Asp85Asn), 2 carried the RF variant in SCN5A (p.Arg1193Gln) and 50 patients carried hot-VUS in KCNQ1, KCNH2, CACNA1C, SCN5a or RYR2 gene. Conclusions In our large cohort of index cases referred for genetic testing for LQTS, diagnostic yield was around 27% when considering only LP/P variants in LQTS or arrhythmic phenotype related genes. Even by including the inconclusive but potentially relevant genetic results (RF and hot VUS), the diagnostic yield in our cohort reaches 35%. This is lower than expected for LQTS cohort. It should be taken into consideration that our cohort consists of heterogeneous patients referred for LQTS genetic testing, which would probably reflects better the real world patients seen in every day practice.

Temporal variability of electromechanical-window negativity in patients with inherited long-QT syndrome or drug-induced QT prolongation: relation to torsades de pointes

Abstract Background Electromechanical window (EMW) negativity is a novel parameter that identifies long-QT syndrome (LQTS) patients at increased arrhythmia risk. Whether the EMW remains stable or alters, especially close to arrhythmic events, is unknown. Purpose To study temporal variability of the EMW in controls and in patients with LQTS or drug-induced QT prolongation (di-QTprol), in relation to the timing of torsades de pointes (TdP) or ventricular fibrillation (VF). Methods In a retrospective cohort study with 5 participating centers, we assessed the EMW (ms) at 2 or 3 timepoints (if applicable, one of them close to TdP/VF) from cw-Doppler echocardiography in the apical long-axis view and concomitant ECG. EMW was calculated by subtracting the QT time from the aortic-valve closure time measured from QRS onset (surrogate of mechanical systole). Data are shown as mean±SD or median (25th to 75th percentile). Results Forty patients (75% female, 43±18 years old) were included: 8 controls, 27 LQTS (10 symptomatic: 9 TdP, 1 VF), and 5 di-QTprol (all TdP). LQTS genotypes were LQT1 (n=11), LQT2 (n=6), LQT3 (n=7), LQT7 (n=2), and genotype-negative (n=1). Overall, 67% of LQTS patients and 20% of di-QTprol were treated with β-blockers. Baseline QTc was 409±32 ms in controls, 444±32 ms in asymptomatic LQTS, 491±41 ms in symptomatic LQTS, and 498±49 ms in di-QTprol. Baseline EMW was 6±14 ms in controls, -12±29 ms in asymptomatic LQTS, -52±16 ms in symptomatic LQTS, and -5±69 ms in di-QTprol. Over years of follow-up, EMW remained stable in controls (8±10 ms; Fig. 1A) and the asymptomatic LQTS group (-17±20 ms; Fig. 1B). In LQTS patients with TdP/VF, EMW (baseline measured at -534 (-962 to -143) days before arrhythmia) had become profoundly more negative, -127±42 ms, (p=0.01 vs. baseline) at 1 (0 to 5) days after the arrhythmia and returned to basal values, -52±18 ms, (p=0.005) after 722 (155 to 3100) days (Fig. 1C). There was a significant correlation between time-to-event and EMW negativity (r=0.72, p<0.0001), indicating stronger EMW negativity when measured closer to TdP/VF. For the di-QTprol patients (n=2 amiodarone; n=2 escitalopram; n=1 sotalol) a trend towards more exaggerated EMW negativity was observed: from baseline (at -19 (-785 to -13) days before the event) to -125±33 ms (p=0.08 vs. baseline) at 1 (1 to 5) days after TdP (Fig. 1D). Again, significant correlation was found between time-to-event and EMW negativity (r=0.6, p=0.03). Conclusion Episodes of electrical instability in patients with LQTS or di-QTprol are associated with profound but reversible exaggeration of EMW negativity. If also present just prior to TdP/VF (as in experimental models of torsadogenic di-QTprol and anecdotal LQTS cases), imminent ventricular arrhythmias may be anticipated as electromechanical reciprocity may contribute to arrhythmogenesis.Figure 1Graphical Abstract

Exercise-induced regional heterogeneity of myocardial left ventricular contraction in hereditary long QT syndrome

Abstract Background Hereditary long QT syndrome (LQTS) is characterized by prolonged ventricular repolarization and an increased risk for ventricular arrhythmias (VA) and sudden cardiac death. Dispersion of ventricular repolarization in LQTS may be more pronounced during exercise and not only impacts the heart’s electrical functions but also the pattern of mechanical contraction. Little is known about the intraventricular heterogeneity of wall motion abnormalities in LQTS patients. Methods In this cross-sectional study we quantify regional, transmural, and longitudinal alterations in left ventricular (LV) contraction at rest, during exercise, and recovery in genetically confirmed LQTS patients and healthy controls to evaluate their value in differentiating high risk patients as defined by a history of VA. ECG and echocardiographic assessments were conducted during rest, peak ergometer activity (105 to 125 bpm), and four minutes into recovery phases. Longitudinal strain (LStr) parameters were measured for each LV segment separately. We differentiated LStr variations as heterogeneity of LStr amplitude (hetLStr; standard deviation [SD] or mean range) and dispersion of time-to-peak LStr (disLStr; SD or mean range). Here, we present the analyses of disLStr. Results At present, 15 patients with genetically confirmed LQTS (LQT1: 8, LQTS 2: 7; mean QTc 475 ± 46 ms) have been included of which 9 had a history of VA. In addition, we collected data from 8 age- and gender-matched control patients (mean QTc 401 ± 18 ms). LV ejection fraction and LStr were not significantly different between the LQTS and control groups. However, LQTS patients exhibited a significantly lower mean LStr in both the endocardial and epicardial layers across all phases compared to control. We observed a significantly more pronounced disLStr across all segments in the LQTS group during the rest and recovery phases in the endocardial layer and during all phases in the epicardial layer. Regional differences (apical vs. basal) were found in all LQTS patients only endocardially at rest. Globally, disLStr showed transmural differences (epicardial vs. endocardial) in both asymptomatic and symptomatic LQTS patients during peak exercise and recovery. Upon segmental analysis, it was observed that symptomatic patients exhibited these transmural differences apically already at rest (p=.004), as well as in the basal region during peak exercise (p=.01), and in the apical region during recovery (p=.02). Conclusion LQTS patients show regional (base to apex) and transmural (endocardial to epicardial) heterogeneities in myocardial contraction at rest, during exercise, and recovery as compared to healthy patients. These regional variations in myocardial contraction patterns may be more pronounced in high risk LQTS patients. Ongoing analyses of hetLStr are in progress to corroborate our findings.

Contribution of clinically relevant copy number variations in long QT syndrome: a large cohort study

Abstract Background Copy number variations (CNVs) have increasingly been recognized in the cardiovascular setting. Their role in hypertrophic cardiomyopathy or sudden cardiac death has been described, but their involvement in Long QT syndrome (LQTS) has been scarcely studied. Most of these descriptions are confined to case reports or small series. Purpose The aim of this study is to investigate the proportion of clinically relevant CNVs among a large cohort of index cases referred for genetic testing due to LQTS. Methods Retrospective study in which the presence of likely pathogenic (LP) or pathogenic (P) CNVs was evaluated in a cohort of index cases referred for genetic study with LQTS phenotype. Index cases were genotyped with customized libraries through next generation sequencing (NGS). CNVs analysis was conducted by a proprietary method by means of comparative analysis of normalized read depth data. Clinically relevant (LP or P) CNVs were then confirmed by other techniques. Results 1183 index cases were referred for genetic testing to our laboratory with long QT syndrome phenotype. Some of these index cases were referred for suspicion of Jervell-Lange-Nielsen syndrome (JNLS) or for LQTS associated with congenital deafness (n=10) or Anderson-Tawil syndrome (n=1). Around 27% (n=322) of the index cases had a positive result. CNVs analysis was run on 1071 (91%) of the index cases. Most of the CNVs were considered negative (n=994, 93%), 59 could not be analysed (6%) and in 11 patients (1%), a LP/P CNV was identified. All CNVs but three (72.72%), were confirmed by orthogonal molecular techniques (Sanger sequencing or multiplex ligation-dependent probe amplification (MLPA)). In all, but two, the genetic study was considered positive due to the CNV findings. One of these patients carried a pathogenic lost-of-function (LOF) CNV in the LDLR gene (incidental finding) but the report was negative because he was referred for suspicion of JLNS. In another case a CNV in the DSP gene was identified in mosaicism (30-40%). However, in this case, the positive report was attributed to the presence of an additional pathogenic variant in CACNA1C. Regarding the other index cases with LP/P CNVs: 4 carried a deletion in the KCNH2 gene, 3 carried a deletion in KCNQ1 gene (one of them in homozygosis and was studied for suspicion of JLNS) and 1 carried a duplication in the KCQN1 (LOF variant). In addition, one patient carried a deletion of exon 3 in the RYR2 gene. Even if the patient was referred for suspicion of LQTS, the report was considered positive. These clinically relevant CNVs represented around 3% of the positive genetic results. Conclusions In our large cohort of patients referred for genetic testing for LQTS, around 1% of clinically relevant CNVs were identified. As such large genomic rearrangements underlie a non-neglectable portion of cases, we consider that their analysis should be performed as part of the routine genetic testing.Deletion exons 6-14 in KCNH2 gene (CNV)MLPA with deletion exons 6-14 in KCNH2

Molecular mechanism for the interaction of MOG1 with the intracellular Loop II of cardiac sodium channel Nav1.5 and its role in Brugada syndrome

Abstract Background SCN5A encodes the cardiac sodium channel a-subunit Nav1.5 with an essential role in generation and conduction of cardiac action potential. Mutations in SCN5A cause Brugada syndrome (BrS) and long QT syndrome (LQTS). The function of Nav1.5, including its cell membrane trafficking and clustering in caveolae, is regulated by MOG1. We recently showed that gene therapy with AAV9-MOG1 reversed disease phenotypes of BrS, dilated cardiomyopathy, cardiac conduction disease and sick sinus syndrome associated with SCN5A mutations. MOG1 is a small chaperon that interacts with multiple sites of Nav1.5, including intracellular Loop I between transmembrane domain I and II and Loop II between domain II and III. We previously characterized the interaction between MOG1 and the intracellular Loop I of Nav1.5, and defined amino acids D24, E36, D44, E53, and E101A of MOG1 as critical residues for its interaction with Nav1.5 Loop I. However, the mechanistic understanding of the interaction between MOG1 and intracellular Loop II of Nav1.5 is needed. Purpose The aim of this study is to perform systematic mutational and electrophysiological studies to identify the key structural elements involved in the interaction between MOG1 and Nav1.5 Loop II and determine the significance of this interaction to the pathogenesis of BrS. Methods Patch-clamping was used to record sodium currents. Glutathione S-transferase (GST) pull-down were used to characterize protein-protein interactions. Results Nav1.5 Loop II contains 765 amino acids, and the domain between amino acids 1157 and 1200 was shown to be the interaction domain for MOG1. Six serial deletions were created, including LoopII△940-982, LoopII△983-1025, LoopII△1026-1068, LoopII△1069-1112, LoopII△1113-1156, and LoopII△1157-1200. GST-pull-down showed that only LoopII△1157-1200 failed to interact with MOG1. MOG1 enhanced sodium current densities all Nav1.5 deletion mutants except for LoopII△1157-1200. Further deletions by every 10-11 amino acids defined the MOG1-interacting domain to 1190-1200. Point mutation analysis defined the MOG1-Nav1.5 Loop II interaction domain to an 8-amino-acid domain of R1193-Y1200 of Nav1.5 Loop II. Mutations R1193W, L1194M and Y1199S associated with BrS reduced the interaction between MOG1 and Nav1.5. MOG1 enhanced sodium current densities from wild type Nav1.5, but not from mutant Nav1.5 with mutations R1193W, L1194M and Y1199S. Conclusions This study identifies the critical elements of Nav1.5 Loop II for interaction with MOG1, and reveals the molecular mechanism by which Nav1.5 mutations R1193W, L1194M and Y1199S cause BrS. Mutations R1193W, L1194M and Y1199S weaken the interaction between Nav1.5 and MOG1, and reduce MOG1-enhanced cardiac sodium current densities.

In vivo KCNQ1-suppression-replacement gene therapy in transgenic LQT1 rabbits restores a physiological QT interval at baseline and under catecholamine infusion

Abstract Background Long-QT syndrome type 1 (LQT1) is a genetic channelopathy characterized by loss-of-function variants in the repolarizing potassium channel KCNQ1/IKs which lead to QT-prolongation and predisposes patients to adverse events such as cardiac arrhythmias. Clincally, during catecholamine infusion or exercise stress test, patients with LQT1 present a paradoxical QT prolongation. Purpose To investigate whether a gene-therapy approach targeting the defective K+-channel, through a KCNQ1 suppression-and-replacement (SupRep) strategy, can restore the healthy phenotype at baseline and under catecholamine infusion in LQT1 transgenic rabbits. Methods Our proprietary dual component suppression-and-replacement (SupRep) gene therapy was created by combining a custom-designed KCNQ1 shRNA and a shRNA-immune (shIMM) KCNQ1 cDNA into AAV9. In vivo KCNQ1-SupRep gene therapy was performed by targeted intra-aortic root injection during balloon occlusion. Two or three weeks post gene therapy, 12-lead ECGs were assessed in adult transgenic LQT1 (KCNQ1-Y315S, loss-of IKs) and wild-type (WT) rabbits to determine the effect of SupRep gene therapy on QT-duration at baseline and under catecholamine infusion (10nM isoproterenol, 5mL/h). Ventricular cardiomyocytes (VCMs) were isolated to investigate the effect of SupRep gene therapy on action potential duration (APD) and calcium transients (Ca2+T) at baseline and under isoproterenol infusion. Results At the cellular level, VCMs isolated from SupRep treated LQT1 rabbits showed a significant shortening of the APD90 as compared to UT-LQT1 (p<0.0001). At the whole animal level, at baseline, the QTi (%) of the UT-LQT1 transgenic rabbits (122±3) was significantly longer compared to the WT rabbits (105±2, p<0.0001). Treatment with AAV9-KCNQ1-SupRep shortened significantly the QTi of the transgenic LQT1 rabbits as compared to the UT-LQT1 rabbits (111±2, p=0.0115). To further investigate the effect of SupRep gene therapy, 12-lead ECGs were obtained in UT-LQT1, sham-treated LQT1, SupRep-treated LQT1, and WT rabbits under ISO infusion. While reaching a 20% increase in heart rate during ISO stimulation, the UT-LQT1 and the sham treated LQT1 rabbits showed a marked QT prolongation, with ΔQTi of 31.4% and 24.5%, respectively, as compared to baseline. In contrast, the SupRep-treated LQT1 rabbit behaved nearly identical to the WT rabbit during ISO infusion, showing a ΔQTi of 16.9% as compared to 16.5% in the WT rabbit. Similarly, in contrast to the UT-LQT1, VCMs isolated from SupRep-treated LQT1 rabbits demonstrated a normalization of the APD and Ca2+T under ISO stimulation, showing a significantly shortened APD90 (p<0.0001) and Ca2+T90 (p=0.0025) which was similar to what was observed in WT VCMs. Conclusion In vivo KCNQ1 suppression-and-replacement gene therapy normalizes the pathologically prolonged APD/QT interval and restores a physiological response to catecholamine infusion in treated LQT1 transgenic rabbits.

Long-term Monitoring to Detect Risk of Sudden Cardiac Death in Inherited Arrhythmia Patients

BackgroundRisk stratification in inherited arrhythmia syndromes is challenging. Implantable cardioverter defibrillators (ICDs) are effective in the prevention of sudden cardiac death but are associated with significant complications. We aimed to determine the value of long-term implantable loop recorder (ILR) monitoring to determine risk factors for arrhythmias in inherited arrhythmia patients. MethodsWe conducted a prospective multicentre study between 2015 and 2020 recruiting inherited arrhythmia probands and family members at intermediate arrhythmic risk, with no class 1 indication for ICD implantation. The primary endpoint was the detection by ILR of nonsustained ventricular tachycardia over ≥ 10 consecutive beats. Secondary endpoints included ICD insertion during follow-up, all-cause mortality, and ILR complication rates. ResultsA total of 45 individuals (30 female participants) were enrolled in the study. The most common diagnoses were long-QT syndrome (28%), Brugada syndrome (26%), and arrhythmogenic cardiomyopathy (11%). Following ILR insertion (mean follow-up 633 days; range, 387-969), cardiac symptoms occurred in 19 of 45 patients (42%), 5 of whom had nonsustained ventricular tachycardias (11%), which were symptomatic in 3 individuals. This situation led to ICD implantation based on ILR in 5 of 45 patients (11%). Fifty percent of symptomatic events occurred in ARVC patients. The median time from ILR insertion to ICD implantation was 152 days (interquartile range (25th, 75th percentiles) 55 of 209). No patient experienced sudden cardiac death. ConclusionsILRs enable the detection of high-risk arrhythmic features and facilitate selection of ICD candidates in inherited arrhythmia patients with borderline indications.

Handgrip strength test for the diagnosis of long QT syndrome

Long QT syndrome (LQTS) assessment is based on the QT/ÖRR equation (the so-called Bazett's formula) and calculating the corrected QT (QTc). However, this formula reliably estimates the QT interval in the heart rate range (HR) of 60-90 bpm, which makes it difficult to diagnose LQTS in patients with sinus bradycardia, typical of this disease.Aim. To improve the LQTS diagnosis in patients with sinus bradycardia using the handgrip strength test.Material and methods. A total of 188 patients aged 5-53 years (16 [13;17]) were examined: group I — 40 healthy children aged 9-17 years (14 [12;16]); group II — 98 athletes with sinus bradycardia <60 bpm aged 16 [16;17] years; group III — 50 patients with LQTS aged 5-53 years (12 [10;16]). The handgrip strength test consisted of regular rhythmic compression of a hand expander with a resistance of 20 kg until the heart rate increased >60 bpm. The patients had an electrocardiogram recorded twice before and after the handgrip test. Heart rate, QT and QTc intervals were analyzed.Results. In all groups, after using the expander, there was a significant increase in heart rate, QT interval shortening and QTc interval prolongation. QTc interval prolongation after the handgrip test >460 ms was a highly sensitive marker of LQTS (Se 96%, Sp 60%).Conclusion. Proposed handgrip test for sinus bradycardia makes it possible to increase heart rate, which ensures a more correct use of the Bazett's formula for assessing the corrected QT interval (QTc). QTc interval prolongation over 460 ms after the handgrip test is a highly sensitive additional marker for identifying patients with LQTS.

A PARTICULAR CASE OF CARDIAC ARREST

Abstract 88–year–old man with arterial hypertension, cognitive impairment, carrier of dual–chamber PM due to sinus node disease. Diagnosis of congenital long QT syndrome (LQT1) approximately 20 years ago during family screening (syncopal episodes in his son positive for LQT1), no history of syncope or major arrhythmias. In home therapy with mirtazapine and norvasc. He arrived in our cardiac intensive care unit for ACC on torsade de pointes trated with two cycles of CPR and 1 DC shock at 200 Joules with ROSC. On arrival ECG evidence of electrically induced ventricular rhythm, QTc 850 msec. On echocardiogram akinesia of the mid–basal segments of all the LV walls (takotsubo–like) with EF 25–30%. On coronary angiography non critical multivessel coronary disease. At laboratory tests increase of WBC, PCR and PCT, slight plateau movement of troponin, normal electrolytes. The patient underwent a chest CT scan with evidence of an inflammatory focus for which antibiotic therapy was started. Congenital long QT syndrome (LQTS) is an inherited condition characterized by prolongation of the QT interval (more than 450 ms in men and 460 ms in women), associated with repolarization abnormalities. There are three main forms transmitted via AD: LQT1, LQT2 and LQT3. The most common variant is LQT1 caused by a loss of function mutation in the KCNQ1 gene which causes a reduction in the outgoing potassium current during phase 3 of the action potential. In these patients, episodes of syncope caused by torsade de pointes, major arrhythmias and sudden cardiac death are frequent. Patients with LQTS should avoid taking drugs that can prolong the QT interval such as some antipsychotics, antidepressants (like mirtazapina that can cause QT prolongation, torsade de pointes, ventricular tachycardia and sudden cardiac death if administered at high doses or in patients with predisposing factors), antibiotics. Furthermore, there are various factors that can influence the QT prolongation: electrolyte alterations, physical stress, cerebrovascular events. The use of mirtazapine for a long period in association with takotsubo syndrome (probably triggered by underlying pneumonia) in a patient with LQTS, may have resulted in massive lengthening of the QT interval causing torsade de pointes and therefore ACC. During the hospitalization, mirtazapine was suspended and nadolol was introduced with a progressive reduction of the QT segment. No new arrhythmic events were recorded.

Long QT syndrome: importance of reassessing arrhythmic risk after treatment initiation.

Risk scores are proposed for genetic arrhythmias. Having proposed in 2010 one such score (M-FACT) for the long QT syndrome (LQTS), this study aims to test whether adherence to its suggestions would be appropriate. LQT1/2/3 and genotype-negative patients without aborted cardiac arrest (ACA) before diagnosis or cardiac events (CEs) below age 1 were included in the study, focusing on an M-FACT score ≥2 (intermediate/high risk), either at presentation (static) or during follow-up (dynamic), previously associated with 40% risk of implantable cardioverter defibrillator (ICD) shocks within 4 years. Overall, 946 patients (26 ± 19 years at diagnosis, 51% female) were included. Beta-blocker (βB) therapy in 94% of them reduced the rate of those with a QTc ≥500 ms from 18% to 12% (P < .001). During 7 ± 6 years of follow-up, none died; 4% had CEs, including 0.4% with ACA. A static M-FACT ≥2 was present in 110 patients, of whom 106 received βBs. In 49/106 patients with persistent dynamic M-FACT ≥2, further therapeutic optimization (left cardiac sympathetic denervation in 55%, mexiletine in 31%, and ICD at 27%) resulted in just 7 (14%) patients with CEs (no ACA), with no CEs in the remaining 57. Additionally, 32 patients developed a dynamic M-FACT ≥2 but, after therapeutic optimization, only 3 (9%) had CEs. According to an M-FACT score ≥2, a total of 142 patients should have received an ICD, but only 22/142 (15%) were implanted, with shocks reported in 3. Beta-blockers often shorten QTc, thus changing risk scores and ICD indications for primary prevention. Yearly risk reassessment with therapy optimization leads to fewer ICD implants (3%) without increasing life-threatening events.

Generation of induced pluripotent stem cell lines from patients with LQT1 caused by heterozygous mutations in the KCNQ1 gene

Long QT Syndrome (LQTS) is a genetic heart disorder that can induce cardiac arrhythmias. The most prevalent subtype, LQT1, stems from rare variants in the KCNQ1 gene. Utilizing induced pluripotent stem cells (iPSCs) enables detailed cellular studies and personalized medicine approaches for this life-threatening condition. We generated two LQT1 iPSC lines with single nucleotide nonsense mutations, c.1031 C > T and c.1121 T > A in KCNQ1. Both lines exhibited typical iPSC morphology, expressed high levels of pluripotent markers, maintained normal karyotype, and possessed the capability to differentiate into three germ layers. These cell lines serve as important tools for investigating the biological mechanisms underlying LQT1 due to mutations in the KCNQ1 gene.

Congenital Long QT Syndrome in Children and Adolescents: A General Overview.

Congenital long QT syndrome (LQTS) represents a disorder of myocardial repolarization characterized by a prolongation of QTc interval on ECG, which can degenerate into fast polymorphic ventricular arrhythmias. The typical symptoms of LQTS are syncope and palpitations, mainly triggered by adrenergic stimuli, but it can also manifest with cardiac arrest. At least 17 genotypes have been associated with LQTS, with a specific genotype-phenotype relationship described for the three most common subtypes (LQTS1, -2, and -3). β-Blockers are the first-line therapy for LQTS, even if the choice of the appropriate patients needing to be treated may be challenging. In specific cases, interventional measures, such as an implantable cardioverter-defibrillator (ICD) or left cardiac sympathetic denervation (LCSD), are useful. The aim of this review is to highlight the current state-of-the-art knowledge on LQTS, providing an updated picture of possible diagnostic algorithms and therapeutic management.

Utilizing median and maximum QTc values improves prediction of breakthrough cardiac events in pediatric long QT syndrome.

Although prior studies indicate that a QTc > 500 ms on a single baseline 12-lead electrocardiogram (ECG) is associated with significantly increased risk of arrhythmic events in long QT syndrome (LQTS), less is known about the risk of persistent QT prolongation. We sought to determine QTc persistence and its prognostic effect on breakthrough cardiac events (BCEs) among pediatric patients treated for LQTS. We performed a retrospective analysis of 433 patients with LQTS evaluated, risk-stratified, and undergoing active guideline-based LQTS treatment between 1999 and 2019. BCEs were defined as arrhythmogenic syncope/seizure, sudden cardiac arrest (SCA), appropriate VF-terminating ICD shock, and sudden cardiac death (SCD). During the median follow-up of 5.5 years (interquartile range [IQR] = 3-9), 32 (7%) patients experienced a total of 129 BCEs. A maximum QTc threshold of 520 ms and median QTc threshold of 490 ms were determined to be strong predictors for BCEs. A landmark analysis controlling for age, sex, genotype, and symptomatic status demonstrated models utilizing both the median QTc and maximum QTc demonstrated the highest discriminatory value (c-statistic = 0.93-0.95). Patients in the high-risk group (median QTc > 490 ms and maximum QTc > 520 ms) had a significantly lower BCE free survival (70%-81%) when compared to patients in both medium-risk (93%-97%) and low-risk (98%-99%) groups. The risk of BCE among patients treated for LQTS increases not only based upon their maximum QTc, but also their median QTc (persistence of QTc prolongation). Patients with a maximum QTc > 520 ms and median QTc > 490 ms over serial 12-lead ECGs are at the highest risk of BCE while on guideline-directed medical therapy.

Single-Center Case Series Study: Barostim Improves Quality of Life Outcomes of Heart Failure Patients

The list of medications known to induce QT prolongation is long. We report here a case of long QT syndrome (LQTS) caused by use of Doxylamine, which has previously only been reported once. The patient arrived to the emergency department after having ingested two bottles of Doxylamine (roughly 96 tablets) in a suicidal attempt and was noted to have seizure-like activity prior to arrival. Upon admission she was found to have severe rhabdomyolysis and significant QTc prolongation (&gt;650 ms). Her clinical status improved over the course of hospitalization with supportive treatment that included intravenous fluids as well as electrolyte monitoring and replacement. Given the relative ubiquity of Doxylamine, the purpose of this article is to raise awareness of a unique adverse effect of a widely available medication.

Antisense oligonucleotide therapeutic approach for Timothy syndrome

Timothy syndrome (TS) is a severe, multisystem disorder characterized by autism, epilepsy, long-QT syndrome and other neuropsychiatric conditions1. TS type 1 (TS1) is caused by a gain-of-function variant in the alternatively spliced and developmentally enriched CACNA1C exon 8A, as opposed to its counterpart exon 8. We previously uncovered several phenotypes in neurons derived from patients with TS1, including delayed channel inactivation, prolonged depolarization-induced calcium rise, impaired interneuron migration, activity-dependent dendrite retraction and an unanticipated persistent expression of exon 8A2–6. We reasoned that switching CACNA1C exon utilization from 8A to 8 would represent a potential therapeutic strategy. Here we developed antisense oligonucleotides (ASOs) to effectively decrease the inclusion of exon 8A in human cells both in vitro and, following transplantation, in vivo. We discovered that the ASO-mediated switch from exon 8A to 8 robustly rescued defects in patient-derived cortical organoids and migration in forebrain assembloids. Leveraging a transplantation platform previously developed7, we found that a single intrathecal ASO administration rescued calcium changes and in vivo dendrite retraction of patient neurons, suggesting that suppression of CACNA1C exon 8A expression is a potential treatment for TS1. Broadly, these experiments illustrate how a multilevel, in vivo and in vitro stem cell model-based approach can identify strategies to reverse disease-relevant neural pathophysiology.

Digital precision medicine in rhythmology : Risk prediction of recurrences, sudden cardiac death, and outcome

Digital precision medicine is gaining increasing importance in rhythmology, especially in the treatment of cardiac arrhythmias. This trend is driven by the advancing digitization in healthcare and the availability of large amounts of data from various sources such as electrocardiograms (ECGs), implants like pacemakers and implantable cardioverter-defibrillators (ICDs), as well as wearables like smartwatches and fitness trackers. Through the analysis of this data, physicians can develop more precise and individualized diagnoses and treatment strategies for patients with cardiac arrhythmias. For example, subtle changes in ECGs can be identified, indicating potentially dangerous arrhythmias. Genetic analyses and resulting large datasets also play an increasingly significant role, especially in hereditary ion channel disorders such as long QT syndrome (LQTS) and Brugada syndrome (BrS), as well as in lone atrial fibrillation (AF). Precision medicine enables the development of individualized treatment approaches tailored to the specific needs and risk factors of each patient. This can help improve screening strategies, reduce adverse events, and ultimately enhance the quality of life for patients. Technological advancements such as big data, artificial intelligence, machine learning, and predictive analytics play acrucial role in predicting the risk of arrhythmias and sudden cardiac death. These concepts enable more precise and personalized predictions and support physicians in the treatment and monitoring of their patients.

A novel mutation in hERG gene associated with azithromycin-induced acquired long QT syndrome.

Mutations in human ether-à-go-go-related gene (hERG) potassium channels are closely associated with long QT syndrome (LQTS). Previous studies have demonstrated that macrolide antibiotics increase the risk of cardiovascular diseases. To date, the mechanisms underlying acquired LQTS remain elusive. A novel hERG mutation I1025N was identified in an azithromycin-treated patient with acquired long QT syndrome via Sanger sequencing. The mutant I1025N plasmid was transfected into HEK-293 cells, which were subsequently incubated with azithromycin. The effect of azithromycin and mutant I1025N on the hERG channel was evaluated via western blot, immunofluorescence, and electrophysiology techniques. The protein expression of the mature hERG protein was down-regulated, whereas that of the immature hERG protein was up-regulated in mutant I1025N HEK-293 cells. Azithromycin administration resulted in a negative effect on the maturation of the hERG protein. Additionally, the I1025N mutation exerted an inhibitory effect on hERG channel current. Moreover, azithromycin inhibited hERG channel current in a concentration-dependent manner. The I1025N mutation and azithromycin synergistically decreased hERG channel expression and hERG current. However, the I1025N mutation and azithromycin did not alter channel gating dynamics. These findings suggest that hERG gene mutations might be involved in the genetic susceptibility mechanism underlying acquired LQTS induced by azithromycin.