Citations

Background: Long-QT syndrome (LQTS) is characterized by QT prolongation and increased risk for syncope, seizures, and sudden cardiac death. The majority of LQTS stems from pathogenic mutations in KCNQ1 , KCNH2 , or SCN5A . However, ≈10% of patients with LQTS remain genetically elusive. We utilized genome sequencing to identify a novel LQTS genetic substrate in a multigenerational genotype-negative LQTS pedigree. Methods: Genome sequencing was performed on 5 affected family members. Only rare nonsynonymous variants present in all affected family members were considered. The candidate variant was characterized functionally in patient-derived induced pluripotent stem cell and gene-edited, variant corrected, isogenic control induced pluripotent stem cell–derived cardiomyocytes. Results: A missense variant (p.G6S) was identified in ALG10B -encoded α-1,2-glucosyltransferase B protein. ALG10B (alpha-1,2-glucosyltransferase B protein) is a known interacting protein of KCNH2 -encoded K v 11.1 (HERG [human Ether-à-go-go–related gene]). Compared with isogenic control, ALG10B-p.G6S induced pluripotent stem cell–derived cardiomyocytes showed (1) decreased protein expression of ALG10B (p.G6S, 0.7±0.18, n=8 versus control, 1.25±0.16, n=9; P <0.05), (2) significant retention of HERG in the endoplasmic reticulum ( P <0.0005), and (3) a significantly prolonged action potential duration confirmed by both patch clamp (p.G6S, 531.1±38.3 ms, n=15 versus control, 324.1±21.8 ms, n=13; P <0.001) and multielectrode assay ( P <0.0001). Lumacaftor—a compound known to rescue HERG trafficking—shortened the pathologically prolonged action potential duration of ALG10B-p.G6S induced pluripotent stem cell–derived cardiomyocytes by 10.6% (n=31 electrodes; P <0.001). Conclusions: Here, we demonstrate that ALG10B-p.G6S downregulates ALG10B, resulting in defective HERG trafficking and action potential duration prolongation. Therefore, ALG10B is a novel LQTS-susceptibility gene underlying the LQTS phenotype observed in a multigenerational pedigree. ALG10B mutation analysis may be warranted, especially in genotype-negative patients with an LQT2-like phenotype.

Background: Hypertrophic cardiomyopathy (HCM) is characterized by asymmetric left ventricular hypertrophy. Currently, hypertrophy pathways responsible for HCM have not been fully elucidated. Their identification could serve as a nidus for the generation of novel therapeutics aimed at halting disease development or progression. Herein, we performed a comprehensive multi-omic characterization of hypertrophy pathways in HCM. Methods: Flash-frozen cardiac tissues were collected from genotyped HCM patients (n=97) undergoing surgical myectomy and tissue from 23 controls. RNA sequencing and mass spectrometry–enabled deep proteome and phosphoproteomic assessment were performed. Rigorous differential expression, gene set enrichment, and pathway analyses were performed to characterize HCM-mediated alterations with emphasis on hypertrophy pathways. Results: We identified transcriptional dysregulation with 1246 (8%) differentially expressed genes and elucidated downregulation of 10 hypertrophy pathways. Deep proteomic analysis identified 411 proteins (9%) that differed between HCM and controls with strong dysregulation of metabolic pathways. Seven hypertrophy pathways were upregulated with antagonistic upregulation of 5 of 10 hypertrophy pathways shown to be downregulated in the transcriptome. Most upregulated hypertrophy pathways encompassed the rat sarcoma-mitogen-activated protein kinase signaling cascade. Phosphoproteomic analysis demonstrated hyperphosphorylation of the rat sarcoma-mitogen-activated protein kinase system suggesting activation of this signaling cascade. There was a common transcriptomic and proteomic profile regardless of genotype. Conclusions: At time of surgical myectomy, the ventricular proteome, independent of genotype, reveals widespread upregulation and activation of hypertrophy pathways, mainly involving the rat sarcoma-mitogen-activated protein kinase signaling cascade. In addition, there is a counterregulatory transcriptional downregulation of the same pathways. Rat sarcoma-mitogen-activated protein kinase activation may serve a crucial role in hypertrophy observed in HCM.

Background: Mitral annular disjunction (MAD) has recently been recognized as an arrhythmogenic entity. Data on the electrophysiological substrate as well as the outcomes of catheter ablation of ventricular arrhythmias in patients with MAD is limited. Methods: Forty patients with MAD (mean age 47±15 years; 70% female) underwent catheter ablation for ventricular arrhythmias. Detailed clinical, electrocardiographic, cardiac imaging, and procedural data were collected. Clinical outcomes were compared between patients who had substrate modification in the MAD area and those who did not. Results: Twenty-three (57.5%) patients had ablation for premature ventricular contractions, 10 (25%) patients for sustained ventricular tachycardia, and 7 (17.5%) patients for premature ventricular contraction-triggered ventricular fibrillation. Mean end-systolic MAD length was 10.58±3.49 mm on transthoracic echocardiography. Seventeen (42.5%) patients had preprocedural cardiac magnetic resonance imaging, and 5 (29%) patients had late gadolinium enhancement. Among the 18 (45%) patients who had abnormal local electrograms (low voltage, long-duration, fractionated, isolated mid-diastolic potentials) during electroanatomical mapping, 10 (25%) patients had abnormal electrograms in the anterolateral mitral annulus and/or MAD area. Substrate modification was performed in 10 (25%) patients. Catheter ablation was acutely successful in 36 (90%) patients (elimination of premature ventricular contraction or noninducibility of ventricular tachycardia). After a median follow-up duration of 54.08 (interquartile range, 10.67–89.79) months, premature ventricular contraction burden decreased from a median of 9.75% (interquartile range, 3.25–14) before the ablation to a median of 4% (interquartile range, 1–7.75) after the ablation ( P =0.03 [95% CI, 0.055–6.5]). Eight (20.5%) patients had repeat ablation for ventricular arrhythmias. Substrate modification of the MAD was associated with a trend toward lower rates of repeat ablation (0% versus 26.7%; P =0.16). Conclusions: Patients with MAD have a complex arrhythmogenic substrate, and catheter ablation is effective in reducing recurrence of ventricular arrhythmias. Substrate mapping and ablation may be considered in these patients.

Background: The transient outward current (Ito) that mediates early (phase 1) repolarization is conducted by the KCND3 -encoded Kv4.3 pore-forming α-subunit. KCND3 gain-of-function mutations have been reported previously as a pathogenic substrate for J wave syndromes (JWS), including the Brugada syndrome and early repolarization syndrome, as well as autopsy-negative sudden unexplained death (SUD). Acacetin, a natural flavone, is a potent Ito current blocker. Acacetin may be a novel therapeutic for KCND3 -mediated J wave syndrome. Methods: KCND3-V392I was identified in an 18-year-old male with J wave syndrome/early repolarization syndrome, and a history of cardiac arrest including ventricular tachycardia/ventricular fibrillation and atrial fibrillation/atrial flutter. Pathogenic KCND3 mutation was engineered by site-directed mutagenesis and co-expressed with wild-type KChIP2 in TSA201 cells. Gene-edited/variant-corrected isogenic control and patient-specific pluripotent stem cell–derived cardiomyocytes (iPSC-CMs) from the p. Val392Ile-KCND3-positive patient were generated. I to currents and action potentials were recorded before and after treatment with Acacetin using the whole cell patch-clamp and multielectrode array technique. Western blot and immunocytochemistry were performed to investigate KCND3 expression. Results: KCND3-V392I demonstrated a marked gain-of-function phenotype, increasing peak I to current density by 92.2% ( P <0.05 versus KCND3-WT). KCND3 expression was significantly increased in KCND3-V392I-derived iPSC-CMs ( P <0.05 versus isogenic control). While KCND3-WT revealed an IC50 of 7.2±1.0 µmol/L for acacetin effect, 30 µmol/L acacetin dramatically inhibited KCND3-V392I peak Ito current density by 96.2% ( P <0.05 versus before Acacetin). Ito was also increased by 60.9% in Kv4.3-V392I iPSC-CM ( P <0.05 versus isogenic control iPSC-CM). Ten micromoles per liter acacetin, a concentration approaching its IC50 value, inhibited Ito by ≈50% in patient-derived iPSC-CMs and reduced the accentuated action potential notch displayed in KCND3-V392I-derived iPSC-CMs. Conclusions: This preclinical study provides pharmacological and functional evidence to suggest that Acacetin may be a novel therapeutic for patients with KCND3 gain-of-function-associated J wave syndrome by inhibiting I to and abolishing the accentuated action potential notch in patient-derived iPSC-CMs.

Introduction: Loss-of-function variants in the RYR2-encoded cardiac calcium release channel underlie a small number of unexplained sudden cardiac arrest and death (SCA/SCD) cases. Objective: To determine if the yield of ultra-rare RYR2 variants in unexplained SCA patients with a default diagnosis of idiopathic ventricular fibrillation (IVF) depends on pre-defined IVF endophenotype and whether RYR2 variant-positive IVF cases have a discernible electrocardiographic phenotype. Methods: All SCA cases referred to Mayo Clinic’s Windland Smith Rice Genetic Heart Rhythm Clinic between 01/2000 and 04/2021 were reviewed. After exclusion of 116 cases where a probable SCA root cause was identified and/or RYR2 genetic testing was not pursued, a total of 64 unexplained SCA cases (36% female; 26 ± 11 years at the time of SCA) were included. Clinical data was extracted from the medical record and patients classified as having i) IVF (no diagnostic clues), N=38 (59%) ii) PVC-triggered ventricular fibrillation (PTVF; documented short coupled PVC-triggered ventricular arrhythmias), N=19 (30%), or iii) early repolarization syndrome (ERS; J-point elevation ≥0.1 mV in ≥2 contiguous leads, excluding leads V 1 -V 3 ), N=7 (11%). The yield of ultra-rare, non-synonymous RYR2 variants (MAF<0.0001 in the Genome Aggregation Database) and electrocardiographic profiles were then compared. Results: Overall, a qualifying, ultra-rare RYR2 non-synonymous variant was identified in 5 out of 64 (7.8%) unexplained SCA patients. Of note, the yield of ultra-rare RYR2 missense variants was significantly higher in individuals with an PTVF endophenotype than those without [4/19 (21%) versus 1/45 (2.2%), p=0.02]. Interestingly, the majority of ultra-rare RYR2 variant-positive unexplained SCA survivors [3/5 (60%)] displayed an exercise stress test pattern characterized by the onset of ventricular ectopy at ~120 bpm with suppression at heart rates ≥140 bpm. Conclusions: The burden of ultra-rare RYR2 variants is higher in unexplained SCA patients with documented PTVF. Collectively, this data suggests that i) a subset of PTVF may have an underlying genetic basis and ii) some IVF- and PTVF-associated RYR2 variants may result in an identifiable electrocardiographic profile.

Background: Multifocal ectopic Purkinje-related premature contractions (MEPPC), the most recent addition to the spectrum of SCN5A -mediated cardiac channelopathies, is characterized by frequent premature ventricular complexes (PVCs) originating from the Purkinje system, atrial arrhythmias, and a predilection for PVC-mediated dilated cardiomyopathy. MEPPC-causative variants tend to cluster within the voltage sensing domain (VSD) of Na v 1.5. Objective: To describe the clinical and cellular electrophysiological phenotype associated with a novel, putative MEPPC-causative SCN5A variant (p.Gln419Pro-SCN5A) that resides outside the VSD of Na v 1.5. Methods: p.Gln419Pro-SCN5A was identified in a 21-year-old male with a history of frequent, multifocal PVCs and atrial tachycardia. Subsequently, three p.Gln419Pro-SCN5A-positive relatives, each with early onset atrial arrhythmias and/or frequent, multifocal PVCs, were identified. All individuals had structurally normal hearts by echocardiography and/or cardiac magnetic resonance imaging. p.Gln419Pro-SCN5A was engineered by site directed mutagenesis and heterologously expressed in TSA201 cells. The standard whole-cell patch clamp technique was used to measure SCN5A wild-type (WT) and p.Gln419Pro-SCN5A sodium currents. Results: Analysis of the current-voltage relationship revealed that neither peak sodium current densities nor late inward sodium current were impacted by p.Gln419Pro-SCN5A. However, p.Gln419Pro-SCN5A significantly shifted V 1/2 in inactivation by -4.6 mV from -84.5±0.7 mV (WT, n=10) to -89.1±0.4 mV (Gln419Pro, n=12, p<0.05 vs. WT) and V 1/2 in activation by -11.6 mV from -30.9±1.3 mV (WT, n=10) to -42.5±1.0 mV (Gln419Pro, n=12, p<0.05 vs. WT) leading to a larger hyperpolarized window current. Flecainide dramatically reduced PVC burden in p.Gln419Pro-SCN5A-positive patients. Conclusions: p.Gln419Pro-SCN5A represents only the fifth MEPPC-causative variant described to date and the first associated with a classic MEPPC phenotype localizing outside Na v 1.5’s VSD. As class I antiarrhythmic drugs are highly efficacious in MEPPC, this clinical entity merits consideration in all young patients found to have a high burden of Purkinje-related PVCs.