Garmany, Ramin

Background: Arrhythmogenic cardiomyopathy (ACM) is a genetic heart disease characterized by proclivity for ventricular arrhythmias and systolic dysfunction. Rarely, sudden cardiac arrest (SCA) is the presenting event in ACM. These individuals may have concealed disease with no appreciable structural manifestations on imaging. Aims: To determine the prevalence, clinical features, and outcomes of patients with genotype-positive ACM who presented with a sentinel SCA. Methods: In this retrospective study, 445 genotype-positive ACM patients evaluated from 1/2015-4/2024 were analyzed to identify those whose initial presentation was SCA. Pertinent demographic, clinical, genetic, and outcome data were extracted from the electronic medical record. Structural manifestations were defined by having one of the following on baseline TTE: EF < 50%, LVEDV >56 mL (male) or >51 mL (female), or regional RV akinesia, dyskinesia, or aneurysm. Categorical variables are presented as number (%), and continuous variables as median (IQR). Statistical analysis was done using Fisher’s exact test (categorical), t-test (continuous), and Logrank test (survival). Patients were censored at time of transplant, death, or last followup. Results: Overall, 14/445 (3%) ACM patients presented with sentinel SCA [71% male; median age at SCA 39 (IQR 30-46)]. The most common genetic substrates were DSP -encoded desmoplakin (n=4; 29%), LMNA -encoded lamin A/C (n=3; 21%), and PKP2 -encoded plakophilin-2 (n=3; 21%). During a median followup of 8.4 years, 12 (86%) experienced appropriate ICD shocks, 8 (57%) required catheter ablation for ventricular arrhythmias, 6 (42%) underwent transplant, and 2 (14%) died of cardiac causes. Notably, 5 (35%) patients did not have evidence of structural disease during baseline evaluation. Compared to those with structural disease, this group was younger [median age 28 (IQR 21-29) vs 45 (IQR 41-48), p = 0.002] and less likely to undergo transplant [0 (0%) vs 6 (42%), p = 0.03]. Conclusions: Sentinel SCA is a rare presentation in ACM and may precede overt structural cardiac changes in a subset of younger patients. This population remains at increased risk for recurrent ventricular arrhythmias and often require advanced therapies.

Background: Although exercise stress testing (EST) has shown to unmask latent electrical substrates in PKP2 -mediated arrhythmogenic cardiomyopathy (ACM), the prognostic role of EST-induced ventricular arrhythmia (VA) patterns in PKP2-ACM patients is unclear. Objective: To examine the exercise-induced VA patterns observed during EST in PKP2-ACM patients and explore their correlation with major VA outcomes. Methods: We retrospectively analyzed 621 patients with either genotype-positive ACM or genotype-positive dilated cardiomyopathy evaluated between 01/2015 and 04/2024 to identify those with a disease-causative variant in PKP2 (N = 108). After exclusion of 38 PKP2-ACM patients without EST data, demographic and electrocardiographic data was extracted from the electronic medical record. The arrhythmic burden [premature ventricular contractions (PVCs), PVCs in bigeminy, PVC couplets, non-sustained VT (NSVT), sustained VT, ventricular fibrillation (VF), and PVCs/minute] during each EST phase (baseline, peak, active and passive recovery) were reviewed and correlated with major VA outcomes [sudden cardiac arrest (SCA), sustained VT, and appropriate implantable cardioverter-defibrillator (ICD) shocks]. Results: A total of 186 exercise tests from 70/108 (65%) PKP2-ACM patients (mean age 36 ± 20 years, 40 % male) were analyzed. The presence and burden of PVCs occurring singly, bigeminy, couplets. and NSVT were more prevalent during the recovery phase, specifically the passive recovery phase ( Table ). PKP2-ACM patients with structural disease (64%) were more likely to have an increased VA burden during recovery than those without structural disease [28/45 (62%) vs 4/25 (16%); p<0.001]. Interestingly, the presence of an increased arrhythmic burden (worsening PVCs/min or complex PVCs) during the recovery was associated with major VA over a median follow-up of 47 months (HR 8.576 [1.015-72.449]; p= 0.019). Conclusion: Increased arrhythmic burden during the recovery phase (late > early) is associated with an increased risk of major VAs in PKP2-ACM. Further studies are needed to understand if this observation extends to other ACM genotypes and whether EST-induced VA patterns can be used to assess therapeutic efficacy in PKP2-ACM.

Background: Disease-causative variants in LMNA -encoded lamin A/C cause a genetic cardiomyopathy characterized by atrioventricular (AV) block, atrial fibrillation (AF) and ventricular arrhythmias (VA). Whether domain/variant-specific late gadolinium enhancement (LGE) patterns exist and carry prognostic significance is unclear. Objective: To determine if specific LGE patterns correlate with LMNA variant type/missense variant localization and clinical outcomes in a large cohort of cardiac laminopathy patients. Methods: Retrospectively, 621 genotype-positive arrhythmogenic/dilated cardiomyopathy patients were analyzed to identify those with a disease-causative variant in LMNA -encoded lamin A/C. After exclusion of patients without cardiac magnetic resonance data, pertinent demographic/clinical data was extracted from the electronic medical record. LGE distribution (subepicardial, midmyocardial, subendocardial, transmural) and location (septal, free wall, apical) patterns were analyzed by variant type (missense vs truncating) and missense variant localization (central rod domain vs tail domain) and correlated with advanced heart failure (AHF), MVA, AV block, AF and thromboembolic (TE) events. Results: 70 out of 116 (60%) LMNA variant-positive (mean age 39±12, 43% male and 94% Caucasian) were included. Missense variants localizing to the C-terminal tail region showed a higher prevalence of transmural and apical LGE (Table). Amongst the 5 (7%) patients with this distinct apical pseudo-infarct LGE pattern, p.Arg471His-LMNA was identified in all cases. Among the 48/70 (69%) of patients with clinically manifest disease, p.Arg471His-LMNA-positive patients (n=6) were noted to have an increased prevalence of TE events [3 (50%) vs 5 (12%), 0=0.050] despite a reduced prevalence of AF [2 (33%) vs 32 (76%), p= 0.031]. No difference in AHF or MVA were observed, but AV block was less prevalent [1 (20%) vs 29 (74%), p= 0.029]. Conclusion: p.Arg471His-LMNA causes a distinct apical pseudo-infarct LGE pattern associated with an increased risk of TE events and a decreased prevalence of AV block and AF. Future studies are needed to better refine the role of domain/variant-specific risk-stratification/anticoagulation in cardiac laminopathy.

Introduction: Cardiac differentiation protocols enable derivation of cardiomyocytes (CM). from human induced pluripotent stem cell (iPS). However, limited tools are available to ensure enhanced iPS-CM maturation without the need for prolonged and complex culture. Research Question: This study assessed whether the mesodermal and cardiopoiesis inductor, Brachyury (T), complements standard cardiac differentiation and augments structural and functional maturation of iPS-CM. Aims: The impact of T mRNA introduction prior to initiation of cardiac differentiation was characterized on: i) cardiac transcription factor expression; ii) structural and functional hallmarks of maturity; and iii) cardiotoxicity. Methods: RT-qPCR was employed to monitor the expression of cardiac-specific transcription factors. Transmission electron microscopy enabled the assessment of cellular ultrastructure. Seahorse measurements were used to determine basal respiration. Fluo4 staining was used to measure calcium transients with external field stimulation. Cardiotoxicity was measured as the impact of doxorubicin on calcium handling and cell survival. Results: Transfection with T prompted early and sustained upregulation of the cardiac transcription factors Nkx2.5 (10-fold; FDR<0.001) and MEF2C (7.2-fold; FDR<0.001). Mitochondrial area was increased with T transfection (P<0.001) and T-transfected iPS-CM exhibited greater basal respiration (FDR<0.05), greater calcium handling resilience in response to field stimulation, and a more mature transient morphology. Following doxorubicin exposure T transfected iPS-CM exhibited a greater compromise in calcium handling (FDR<0.001), and exaggerated cell death. Conclusion: Induction of human iPS with T prior to initiation of cardiac differentiation enhanced signatures of structural and functional maturity, and provided more robust readouts of drug toxicity.

BACKGROUND: Genetic testing for cardiac channelopathies is the standard of care. However, many rare genetic variants remain classified as variants of uncertain significance (VUS) due to lack of epidemiological and functional data. Whether deep protein language models may aid in VUS resolution remains unknown. Here, we set out to compare how 2 deep protein language models perform at VUS resolution in the 3 most common long-QT syndrome–causative genes compared with the gold-standard patch clamp. METHODS: A total of 72 rare nonsynonymous VUS (9 KCNQ1, 19 KCNH2 , and 50 SCN5A ) were engineered by site-directed mutagenesis and expressed in either HEK293 cells or TSA201 cells. Whole-cell patch-clamp technique was used to functionally characterize these variants. The protein language models, evolutionary scale modeling, version 1b and AlphaMissense, were used to predict the variant effect of missense variants and compared with patch clamp. RESULTS: Considering variants in all 3 genes, the evolutionary scale modeling, version 1b model had a receiver operating characteristic curve–area under the curve of 0.75 ( P =0.0003). It had a sensitivity of 88% and a specificity of 50%. AlphaMissense performed well compared with patch-clamp with an receiver operating characteristic curve-area under the curve of 0.85 ( P <0.0001), sensitivity of 80%, and specificity of 76%. CONCLUSIONS: Deep protein language models aid in VUS resolution with high sensitivity but lower specificity. Thus, these tools cannot fully replace functional characterization but can aid in reducing the number of variants that may require functional analysis.

Introduction: Hypertrophic cardiomyopathy (HCM) is a major cause of sudden cardiac death in the young and especially in young athletes. The treadmill exercise test (TEST) has an important role in the functional assessment of patients with HCM. Herein, TEST-derived peak oxygen uptake (VO2 max) is a proxy for cardiopulmonary fitness. However, the correlation between symptoms and VO2 - especially in active young patients with HCM - is unclear. Objective: To explore the relationship between TEST-derived VO2 and symptomology in athletes and non-athletes with HCM. Methods: Overall, 91 patients (40% female) with HCM and a TEST with VO 2 data were included in this case-control study. Data were abstracted from the electronic health record using the first TEST performed at Mayo Clinic. Patients were defined as an athlete if they participated in an organized, competitive sport. Results: Overall, 60 HCM patients (33% female) were categorized as an athlete. HCM athletes were younger (17 vs. 27 years; p<0.0001) and less likely to be symptomatic at the time of their TEST (OR=0.24; p=0.01). Additionally, athletes had a higher VO2 max (86% vs. 68% of predicted; p<0.0001) and reached higher exercise times (93% vs 76% of predicted; p<0.0001) compared to the non-athletes with HCM. When comparing symptomatic (n = 18) to asymptomatic patients (n = 73), symptomatic patients had i) a reduced exercise time (68% vs. 92% of predicted; p=0.002) and ii) lower VO2 max (62% vs 84% of predicted; p<0.001). Finally, among the 27 patients (30%) with a VO 2 max < 70% predicted (n=27), they i) more likely had a history of cardiac arrest (4 vs 0; p=0.006) and ii) were more likely to be symptomatic (OR=7.7; p< 0.0001) than those with VO2 ≥ 70% of predicted (n=64). Importantly, however, 15/27 (56%) patients (8 athletes/7 non-athletes) with a VO2 max < 70% were asymptomatic. Conclusions: Reduced exercise capacity is common in athletes with HCM and does not correlate with symptom status. However, if the TEST-derived VO2 max is < 70%, treatment intensification may be necessary as it may portend an adverse outcome both arrhythmically and mechanically.

Introduction: Disease-causative variants in LMNA -encoded lamin A/C cause a penetrant dilated cardiomyopathy associated with conduction disease (CD) and ventricular arrhythmias (VA). Prior studies show conflicting results regarding the effect of sex, variant type, and CD on severity and sudden death risk in cardiac laminopathy. Methods: In this retrospective study, we utilized a single-center genetic arrhythmogenic cardiomyopathy registry (N=432) to identify patients with ultra-rare LMNA variants and ascertain the effect of sex, mutation type, and clinical presentation on disease severity. Advanced heart failure (AHF) events were defined as ventricular assist device placement or cardiac transplantation whereas VA events were defined as sudden cardiac arrest, sustained ventricular arrhythmia, or appropriate implantable cardioverter-defibrillator shock. Results: The LMNA variant-positive cohort (N=83) had an age of diagnosis of 36 ± 10 years with 80% showing a cardiac phenotype. No difference in the age at diagnosis (37 vs. 34 years; p=0.3) or prevalence of AHF events (14% vs. 18%; p=0.8) and VA events (34% vs. 36%; p=0.9) was observed between females (N=50) and males (N=33). Furthermore, there was no difference in age of diagnosis (37 vs. 33 years; p=0.1) or prevalence of AHF (14% vs. 12%; p=0.9) and VA events (33% vs. 34%; p=0.9) between individuals with missense (N=42) or frameshift (N=32) variants. Interestingly, in comparison to individuals that presented with isolated CD (N=11) those that first presented with left ventricular dysfunction or arrhythmias (atrial or ventricular) (N=41) were more likely to have VA events (9% vs. 53%; p=0.01) and AHF events (0% vs 29%; p=0.05). In addition, those with structural/arrhythmic presentation had earlier time to VA events (15 years vs. 22 years; p=0.04) and AHF events (23 vs undefined; p=0.09) than those with CD. Conclusions: Contrary to prior studies, in this large single-center cardiac laminopathy study, sex and mutation type were not associated with increased risk of AHF or VA events. However, patients presenting with isolated CD had a lower risk of AHF and VA events. Contemporary multi-center studies are needed to better ascertain risk factors associated with AHF and VA events in cardiac laminopathy.

Introduction: Recently, we demonstrated transcriptional down-regulation of hypertrophy pathways in myectomy tissue derived from patients with obstructive hypertrophic cardiomyopathy (HCM) despite translational activation of associated hypertrophy pathways. Previous studies of other forms of cardiac hypertrophy demonstrate hypertrophy pathways are activated via histone acetylation and histone deacetylase (HDAC) inhibitors prevented development of hypertrophy. Whether histone modifications regulate hypertrophy pathways in HCM remains unexplored. Methods: Chromatin immunoprecipitation sequencing (ChIP-seq) was performed on myectomy tissues from 40 patients with HCM and 23 controls to characterize differential histone marks for H3K4me3 (active promoter), H3K9ac (active promoter), H3K9me3 (heterochromatin), H3K27ac (active enhancer), H3K27me3 (repression) across the genome. The differential histone marks were integrated with RNA-sequencing (RNA-seq) data derived from the same patients. Results: Overall, we identified 2,912 (7%) differential H3K4me3 peaks impacting 5 pathways, 23,339 (21%) differential H3K9ac peaks impacting 87 pathways, 33 (0.05%) differential H3K9me3 peaks, and 58,837 (42%) differential H3K27ac peaks impacting 50 pathways. There was increased H3K27ac (q-value = 0.002) and H3K9ac (q-value = 0.009) at RAS-MAPK loci. Univariate analysis of concordance between H3K9ac with RNA-seq data showed activation of cardiac hypertrophy signaling (z-score=4.5; -log [BH p-value] = 3.1), while H3K27me showed down-regulation of cardiac hypertrophy signaling (z-score=-2.4; -log [BH p-value] = 2.4). Multivariate integration of the ChIP-seq data with RNA-seq data identified 3,189 transcripts concordant with all five histone marks impacting 254 pathways, 18 (7%) of which are involved in cardiac hypertrophy. Conclusions: Myectomy tissue from patients with obstructive HCM shows enrichment of histone modifications at cardiac hypertrophy loci with activation via H3K27 and H3K9 acetylation and repression via H3K27 methylation. Further studies are needed to understand the balance and temporal relationship between these competing regulators in HCM pathobiology and the potential for epigenetic therapies.

Introduction: Treatment options for patients with recurrent ventricular arrhythmias (VAs) refractory to pharmacotherapy and/or ablation are minimal. Although left cardiac sympathetic denervation (LCSD) is well established in long QT syndrome (LQTS) and catecholaminergic polymorphic ventricular tachycardia (CPVT), LCSD’s role in idiopathic ventricular fibrillation (IVF) is less clear. Here, we report our single-center experience regarding the early outcomes of LCSD in patients with IVF. Hypothesis: To determine the efficacy of LCSD on reducing VAs in IVF patients refractory to conventional therapy. Methods: A retrospective analysis of the electronic health record of patients with IVF between January 2000 to April 2023 was performed. Cardiac event rates [defined as arrhythmic syncope/seizure, appropriate implantable cardioverter defibrillator (ICD) therapy, sudden cardiac arrest (SCA), or sudden cardiac death (SCD)] before and after LCSD for each patient were calculated and a Wilcoxon signed rank test was performed. Results: In total, 94 patients (41% female) were classified as IVF. Overall, 15/94 patients (16%) had LCSD due to refractory VAs (60% male; mean age at LCSD 36 ± 20 years) despite beta-blocker treatment (100%) or combined drug therapy (67%). In total, 14 patients (15%) had an electrophysiology study performed before LCSD of which 10 (67%) had VAs ablation. All patients had a structurally normal heart and genetic testing performed in 14/15 patients (93%) confirmed absence of genetic cardiomyopathies and channelopathies. SCA was the sentinel event in 10 of 15 patients (67%); 14 patients (93%) had an ICD in place. The average follow-up time after LCSD was 3 ± 4 years. The VA event rate before LCSD was 4 ± 3 events per year and was reduced to 1 ± 3 per year (p<0.04) after LCSD. Patients had a median of 3.2 less events per year after LCSD (p=0.02) with 7 patients (47%) having no post-LCSD recurrences thus far while 3 (20%) patients were deemed LCSD non-responders with no reduction of event rate. Conclusions: LCSD represents a substrate-independent, anti-fibrillatory treatment option for patients with life-threatening, genetic arrhythmias. While not curative, the early outcomes of LCSD for patients with medically refractory IVF are promising.

Introduction: Hypertrophic cardiomyopathy (HCM) is characterized by left ventricular hypertrophy. Approximately 40-50% of HCM is caused by pathogenic variants in genes encoding proteins of the cardiac sarcomere. Patients with sarcomeric HCM have earlier expression and more severe disease progression. Herein, we evaluated the proteomic differences between patients with sarcomeric HCM and genotype negative HCM. Methods: Mass-spectrometry was performed on cardiac tissue from genotyped patients with HCM obtained after surgical myectomy, followed by differential proteomics (log2fc ≥