Long-term reliability of the phospholamban (PLN) p.(Arg14del) risk model in predicting major ventricular arrhythmia: a landmark study.

AIMS: Recently, a genetic variant-specific prediction model for phospholamban (PLN) p.(Arg14del)-positive individuals was developed to predict individual major ventricular arrhythmia (VA) risk to support decision-making for primary prevention implantable cardioverter defibrillator (ICD) implantation. This model predicts major VA risk from baseline data, but iterative evaluation of major VA risk may be warranted considering that the risk factors for major VA are progressive. Our aim is to evaluate the diagnostic performance of the PLN p.(Arg14del) risk model at 3-year follow-up. METHODS AND RESULTS: We performed a landmark analysis 3 years after presentation and selected only patients with no prior major VA. Data were collected of 268 PLN p.(Arg14del)-positive subjects, aged 43.5 ± 16.3 years, 38.9% male. After the 3 years landmark, subjects had a mean follow-up of 4.0 years (± 3.5 years) and 28 (10%) subjects experienced major VA with an annual event rate of 2.6% [95% confidence interval (CI) 1.6-3.6], defined as sustained VA, appropriate ICD intervention, or (aborted) sudden cardiac death. The PLN p.(Arg14del) risk score yielded good discrimination in the 3 years landmark cohort with a C-statistic of 0.83 (95% CI 0.79-0.87) and calibration slope of 0.97. CONCLUSION: The PLN p.(Arg14del) risk model has sustained good model performance up to 3 years follow-up in PLN p.(Arg14del)-positive subjects with no history of major VA. It may therefore be used to support decision-making for primary prevention ICD implantation not merely at presentation but also up to at least 3 years of follow-up.

Longitudinal analysis of serum neurofilament light chain levels as marker for neuronal damage in hereditary transthyretin amyloidosis.

OBJECTIVE: To evaluate serum neurofilament light chain (sNfL) as biomarker of disease onset, progression and treatment effect in hereditary transthyretin (ATTRv) amyloidosis patients and TTR variant (TTRv) carriers. METHODS: sNfL levels were assessed longitudinally in persistently asymptomatic TTRv carriers (N = 12), persistently asymptomatic ATTRv amyloidosis patients (defined as asymptomatic patients but with amyloid detectable in subcutaneous abdominal fat tissue) (N = 8), in TTRv carriers who developed polyneuropathy (N = 7) and in ATTRv amyloidosis patients with polyneuropathy on treatment (TTR-stabiliser (N = 20) or TTR-silencer (N = 18)). Polyneuropathy was confirmed by nerve conduction studies or quantitative sensory testing. sNfL was analysed using a single-molecule array assay. RESULTS: sNfL increased over 2 years in persistently asymptomatic ATTRv amyloidosis patients, but did not change in persistently asymptomatic TTRv carriers. In all TTRv carriers who developed polyneuropathy, sNfL increased from 8.4 to 49.8 pg/mL before the onset of symptoms and before polyneuropathy could be confirmed neurophysiologically. In symptomatic ATTRv amyloidosis patients on a TTR-stabiliser, sNfL remained stable over 2 years. In patients on a TTR-silencer, sNfL decreased after 1 year of treatment. CONCLUSION: sNfL is a biomarker of early neuronal damage in ATTRv amyloidosis already before the onset of polyneuropathy. Current data support the use of sNfL in screening asymptomatic TTRv carriers and in monitoring of disease progression and treatment effect.

High-Sensitivity Cardiac Troponin T to Exclude Cardiac Involvement in TTR Variant Carriers and ATTRv Amyloidosis Patients.

(1) Background: Individuals carrying a pathogenic transthyretin gene variant (TTRv) are at high risk for developing hereditary transthyretin (ATTRv) amyloidosis and are routinely screened for the development of cardiomyopathy (ATTRv-CM). This study aims to evaluate whether the cardiac biomarkers N-terminal pro B-type natriuretic peptide (NT-proBNP) and high-sensitivity cardiac troponin T (hs-cTnT) can be used to rule out ATTRv-CM. (2) Methods: In this retrospective case-control study, data from 46 ATTRv-CM patients and 101 TTRv carriers and ATTRv amyloidosis patients without cardiomyopathy were included. Binary logistic regression models were used to assess the ability of NT-proBNP and hs-cTnT to predict the diagnosis of ATTRv-CM. An optimal cutoff for the relevant biomarker(s) was determined based on a sensitivity of ≥99% and the highest possible percentage of additional tests avoided (%ATA) in the index dataset. (3) Results: Hs-cTnT demonstrated the highest predictive capabilities for ATTRv-CM. The addition of NT-proBNP did not improve the predictive model. A hs-cTnT cutoff of <6 ng/L resulted in a 97% sensitivity and a negative predictive value of 95% with a %ATA of 30% in the validation dataset. (4) Conclusion: In conclusion, hs-cTnT is a useful biomarker for excluding cardiac involvement in TTRv carriers and ATTRv amyloidosis patients and it has the potential to prevent unnecessary diagnostic procedures.

ECG-only explainable deep learning algorithm predicts the risk for malignant ventricular arrhythmia in phospholamban cardiomyopathy.

BACKGROUND: Phospholamban (PLN) p.(Arg14del) variant carriers are at risk for development of malignant ventricular arrhythmia (MVA). Accurate risk stratification allows timely implantation of intracardiac defibrillators and is currently performed with a multimodality prediction model. OBJECTIVE: This study aimed to investigate whether an explainable deep learning-based approach allows risk prediction with only electrocardiogram (ECG) data. METHODS: A total of 679 PLN p.(Arg14del) carriers without MVA at baseline were identified. A deep learning-based variational auto-encoder, trained on 1.1 million ECGs, was used to convert the 12-lead baseline ECG into its FactorECG, a compressed version of the ECG that summarizes it into 32 explainable factors. Prediction models were developed by Cox regression. RESULTS: The deep learning-based ECG-only approach was able to predict MVA with a C statistic of 0.79 (95% CI, 0.76-0.83), comparable to the current prediction model (C statistic, 0.83 [95% CI, 0.79-0.88]; P = .054) and outperforming a model based on conventional ECG parameters (low-voltage ECG and negative T waves; C statistic, 0.65 [95% CI, 0.58-0.73]; P < .001). Clinical simulations showed that a 2-step approach, with ECG-only screening followed by a full workup, resulted in 60% less additional diagnostics while outperforming the multimodal prediction model in all patients. A visualization tool was created to provide interactive visualizations (https://pln.ecgx.ai). CONCLUSION: Our deep learning-based algorithm based on ECG data only accurately predicts the occurrence of MVA in PLN p.(Arg14del) carriers, enabling more efficient stratification of patients who need additional diagnostic testing and follow-up.

Phenotypic variability of filamin C-related cardiomyopathy: Insights from a novel Dutch founder variant.

BACKGROUND: Dilated cardiomyopathy (DCM) can be caused by truncating variants in the filamin C gene (FLNC). A new pathogenic FLNC variant, c.6864_6867dup, p.(Val2290Argfs∗23), was recently identified in Dutch patients with DCM. OBJECTIVES: The report aimed to evaluate the phenotype of FLNC variant carriers and to determine whether this variant is a founder variant. METHODS: Clinical and genetic data were retrospectively collected from variant carriers. Cardiovascular magnetic resonance studies were reassessed. Haplotypes were reconstructed to determine a founder effect. The geographical distribution and age of the variant were determined. RESULTS: Thirty-three individuals (of whom 23 [70%] were female) from 9 families were identified. Sudden cardiac death was the first presentation in a carrier at the age of 28 years. The median age at diagnosis was 41 years (range 19-67 years). The phenotype was heterogeneous. DCM with left ventricular dilation and reduced ejection fraction (<45%) was present in 11 (33%) individuals, 3 (9%) of whom underwent heart transplantation. Cardiovascular magnetic resonance showed late gadolinium enhancement in 13 (65%) of the assessed individuals, primarily in a ringlike distribution. Nonsustained ventricular arrhythmias were detected in 6 (18%), and 5 (15%) individuals received an implantable cardioverter-defibrillator. A shared haplotype spanning 2.1 Mb was found in all haplotyped individuals. The variant originated between 275 and 650 years ago. CONCLUSION: The pathogenic FLNC variant c.6864_6867dup, p.(Val2290Argfs∗23) is a founder variant originating from the south of the Netherlands. Carriers are susceptible to developing heart failure and ventricular arrhythmias. The cardiac phenotype is characterized by ringlike late gadolinium enhancement, even in individuals without significantly reduced left ventricular function.

Exercise does not influence development of phenotype in PLN p.(Arg14del) cardiomyopathy.

BACKGROUND: Endurance and frequent exercise are associated with earlier onset of arrhythmogenic right ventricular cardiomyopathy (ARVC) and ventricular arrhythmias (VA) in desmosomal gene variant carriers. Individuals with the pathogenic c.40_42del; p.(Arg14del) variant in the PLN gene are frequently diagnosed with ARVC or dilated cardiomyopathy (DCM). The aim of this study was to evaluate the effect of exercise in PLN p.(Arg14del) carriers. METHODS: In total, 207 adult PLN p.(Arg14del) carriers (39.1% male; mean age 53 ± 15 years) were interviewed on their regular physical activity since the age of 10 years. The association of exercise with diagnosis of ARVC, DCM, sustained VA and hospitalisation for heart failure (HF) was studied. RESULTS: Individuals participated in regular physical activities with a median of 1661 metabolic equivalent of task (MET) hours per year (31.9 MET-hours per week) until clinical presentation. The 50% most and least active individuals had a similar frequency of sustained VA (18.3% vs 18.4%; p = 0.974) and hospitalisation for HF (9.6% vs 8.7%; p = 0.827). There was no relationship between exercise and survival free from (incident) sustained VA (p = 0.65), hospitalisation for HF (p = 0.81), diagnosis of ARVC (p = 0.67) or DCM (p = 0.39) during follow-up. In multivariate analyses, exercise was not associated with sustained VA or HF hospitalisation during follow-up in this relatively not-active cohort. CONCLUSION: There was no association between the amount of exercise and the susceptibility to develop ARVC, DCM, VA or HF in PLN p.(Arg14del) carriers. This suggested unaffected PLN p.(Arg14del) carriers can safely perform mild-moderate exercise, in contrast to desmosomal variant carriers and ARVC patients.

Phenotypic and Genetic Factors Associated with Absence of Cardiomyopathy Symptoms in PLN:c.40_42delAGA Carriers.

The c.40_42delAGA variant in the phospholamban gene (PLN) has been associated with dilated and arrhythmogenic cardiomyopathy, with up to 70% of carriers experiencing a major cardiac event by age 70. However, there are carriers who remain asymptomatic at older ages. To understand the mechanisms behind this incomplete penetrance, we evaluated potential phenotypic and genetic modifiers in 74 PLN:c.40_42delAGA carriers identified in 36,339 participants of the Lifelines population cohort. Asymptomatic carriers (N = 48) showed shorter QRS duration (- 5.73 ms, q value = 0.001) compared to asymptomatic non-carriers, an effect we could replicate in two different independent cohorts. Furthermore, symptomatic carriers showed a higher correlation (rPearson = 0.17) between polygenic predisposition to higher QRS (PGSQRS) and QRS (p value = 1.98 × 10-8), suggesting that the effect of the genetic variation on cardiac rhythm might be increased in symptomatic carriers. Our results allow for improved clinical interpretation for asymptomatic carriers, while our approach could guide future studies on genetic diseases with incomplete penetrance.

SEPT-GD: A decision tree to prioritise potential RNA splice variants in cardiomyopathy genes for functional splicing assays in diagnostics.

BACKGROUND: Splice prediction algorithms currently used in routine DNA diagnostics have limited sensitivity and specificity, therefore many potential splice variants are classified as variants of uncertain significance (VUSs). However, functional assessment of VUSs to test splicing is labour-intensive and time-consuming. We developed a decision tree to prioritise potential splice variants for functional studies and functionally verified the outcome of the decision tree. MATERIALS AND METHODS: We built the decision tree, SEPT-GD, by setting thresholds for the splice prediction programs implemented in Alamut. A set of 343 variants with known effects on splicing was used as control for sensitivity and specificity. We tested SEPT-GD using variants from a Dutch cardiomyopathy cohort of 2002 patients that were previously classified as VUS and predicted to have a splice effect according to diagnostic rules. We then selected 12 VUSs ranked by SEPT-GD to functionally verify the predicted effect on splicing using a minigene assay: 10 variants predicted to have a strong effect and 2 with a weak effect. RT-PCR was performed for nine variants. Variant classification was re-evaluated based on the functional test outcome. RESULTS: Compared to similar individually tested algorithms, SEPT-GD shows higher sensitivity (91 %) and comparable specificity (88 %) for both consensus (dinucleotides at the start and end of the intron, GT at the 5' end and AG at the 3' end) and non-consensus splice-site variants (excluding middle of exon variants). Using clinical diagnostic criteria, 1295 unique variants in our cardiomyopathy cohort had originally been classified as VUSs, with 57 predicted by Alamut to have an effect on splicing. Using SEPT-GD, we prioritised 31 variants in 40 patients. In the minigene assay, all 12 variants showed results concordant with SEPT-GD predictions. RT-PCR confirmed the minigene results for two variants, TMEM43 c.1000 + 5G > T and TTN c.25922-6 T > G. Based on all outcomes, the SGCD c.4-1G > A and CSRP3 c.282-5_285del variants were reclassified as likely pathogenic. CONCLUSION: SEPT-GD outperforms the tools commonly used for RNA splicing prediction and improves prioritisation of variants in cardiomyopathy genes for functional splicing analysis in a diagnostic setting.

Validation of New Gene Variant Classification Methods: a Field-Test in Diagnostic Cardiogenetics.

Background: In the molecular genetic diagnostics of Mendelian disorders, solutions are needed for the major challenge of dealing with the large number of variants of uncertain significance (VUSs) identified using next-generation sequencing (NGS). Recently, promising approaches using constraint metrics to calculate case excess scores (CE), etiological fractions (EF), and gnomAD-derived constraint scores have been reported that estimate the likelihood of rare variants in specific genes or regions that are pathogenic. Our objective is to study the usability of these constraint data into variant interpretation in a diagnostic setting, using our cardiomyopathy cohort. Methods and Results: Patients (N = 2002) referred for clinical genetic diagnostics underwent NGS testing of 55-61 genes associated with cardiomyopathies. Previously classified likely pathogenic (LP) and pathogenic (P) variants were used to validate the use of data from CE, EF, and gnomAD constraint analyses for (re)classification of associated variant types in specific cardiomyopathy subtype-related genes. The classifications corroborated in 94% (354/378) of cases. Next, we reclassified 23 unique VUSs to LP, increasing the diagnostic yield by 1.2%. In addition, 106 unique VUSs (5.3% of patients) were prioritized for co-segregation or functional analyses. Conclusions: Our analysis confirms that the use of constraint metrics data can improve variant interpretation, and we, therefore, recommend using constraint scores on other cohorts and disorders and its inclusion in variant interpretation protocols.

Sex-specific aspects of phospholamban cardiomyopathy: The importance and prognostic value of low-voltage electrocardiograms.

BACKGROUND: A pathogenic variant in the gene encoding phospholamban (PLN), a protein that regulates calcium homeostasis of cardiomyocytes, causes PLN cardiomyopathy. It is characterized by a high arrhythmic burden and can progress to severe cardiomyopathy. Risk assessment guides implantable cardioverter-defibrillator therapy and benefits from personalization. Whether sex-specific differences in PLN cardiomyopathy exist is unknown. OBJECTIVE: The purpose of this study was to improve the accuracy of PLN cardiomyopathy diagnosis and risk assessment by investigating sex-specific aspects. METHODS: We analyzed a multicenter cohort of 933 patients (412 male, 521 female) with the PLN p.(Arg14del) pathogenic variant following up on a recently developed PLN risk model. Sex-specific differences in the incidence of risk model components were investigated: low-voltage electrocardiogram (ECG), premature ventricular contractions, negative T waves, and left ventricular ejection fraction. RESULTS: Sustained ventricular arrhythmias (VAs) occurred in 77 males (18.7%) and 61 females (11.7%) (P = .004). Of the 933 cohort members, 287 (31%) had ≥1 low-voltage ECG during follow-up (180 females [63%], 107 males [37%]; P = .006). Female sex, age, age at clinical presentation, and proband status predicted low-voltage ECG during follow-up (area under the curve: 0.78). Sustained VA-free survival was lowest in males with low-voltage ECG (P <.001). CONCLUSION: Low-voltage ECGs predict sustained VA and are a component of the PLN risk model. Low-voltage ECGs are more common in females, yet prognostic value is greater in males. Future studies should determine the impact of this difference on the risk prediction of PLN cardiomyopathy and possibly other cardiomyopathies.

Prediction of ventricular arrhythmia in phospholamban p.Arg14del mutation carriers-reaching the frontiers of individual risk prediction.

AIMS: This study aims to improve risk stratification for primary prevention implantable cardioverter defibrillator (ICD) implantation by developing a new mutation-specific prediction model for malignant ventricular arrhythmia (VA) in phospholamban (PLN) p.Arg14del mutation carriers. The proposed model is compared to an existing PLN risk model. METHODS AND RESULTS: Data were collected from PLN p.Arg14del mutation carriers with no history of malignant VA at baseline, identified between 2009 and 2020. Malignant VA was defined as sustained VA, appropriate ICD intervention, or (aborted) sudden cardiac death. A prediction model was developed using Cox regression. The study cohort consisted of 679 PLN p.Arg14del mutation carriers, with a minority of index patients (17%) and male sex (43%), and a median age of 42 years [interquartile range (IQR) 27-55]. During a median follow-up of 4.3 years (IQR 1.7-7.4), 72 (10.6%) carriers experienced malignant VA. Significant predictors were left ventricular ejection fraction, premature ventricular contraction count/24 h, amount of negative T waves, and presence of low-voltage electrocardiogram. The multivariable model had an excellent discriminative ability {C-statistic 0.83 [95% confidence interval (CI) 0.78-0.88]}. Applying the existing PLN risk model to the complete cohort yielded a C-statistic of 0.68 (95% CI 0.61-0.75). CONCLUSION: This new mutation-specific prediction model for individual VA risk in PLN p.Arg14del mutation carriers is superior to the existing PLN risk model, suggesting that risk prediction using mutation-specific phenotypic features can improve accuracy compared to a more generic approach.

Diagnostic yield of targeted next generation sequencing in 2002 Dutch cardiomyopathy patients.

BACKGROUND: Next-generation sequencing (NGS) is increasingly used for clinical evaluation of cardiomyopathy patients as it allows for simultaneous screening of multiple cardiomyopathy-associated genes. Adding copy number variant (CNV) analysis of NGS data is not routine yet and may contribute to the diagnostic yield. OBJECTIVES: Determine the diagnostic yield of our targeted NGS gene panel in routine clinical diagnostics of Dutch cardiomyopathy patients and explore the impact of exon CNVs on diagnostic yield. METHODS: Patients (N = 2002) referred for clinical genetic analysis underwent diagnostic testing of 55-61 genes associated with cardiomyopathies. Samples were analyzed and evaluated for single nucleotide variants (SNVs), indels and CNVs. CNVs identified in the NGS data and suspected of being pathogenic based on type, size and location were confirmed by additional molecular tests. RESULTS: A (likely) pathogenic (L)P variant was detected in 22.7% of patients, including 3 with CNVs and 25 where a variant was identified in a gene currently not associated with the patient's cardiomyopathy subtype. Only 15 out of 2002 patients (0.8%) were found to carry two (L)P variants. CONCLUSION: The yield of routine clinical diagnostics of cardiomyopathies was relatively low when compared to literature. This is likely due to the fact that our study reports the outcome of patients in daily routine diagnostics, therefore also including patients not fully fulfilling (subtype specific) cardiomyopathy criteria. This may also explain why (L)P variants were identified in genes not associated with the reported subtype. The added value of CNV analysis was shown to be limited but not negligible.

The effect of tropomyosin variants on cardiomyocyte function and structure that underlie different clinical cardiomyopathy phenotypes.

Background - Variants within the alpha-tropomyosin gene (TPM1) cause dominantly inherited cardiomyopathies, including dilated (DCM), hypertrophic (HCM) and restrictive (RCM) cardiomyopathy. Here we investigated whether TPM1 variants observed in DCM and HCM patients affect cardiomyocyte physiology differently. Methods - We identified a large family with DCM carrying a recently identified TPM1 gene variant (T201M) and a child with RCM with compound heterozygote TPM1 variants (E62Q and M281T) whose family members carrying single variants show diastolic dysfunction and HCM. The effects of TPM1 variants (T201M, E62Q or M281T) and of a plasmid containing both the E62Q and M281T variants on single-cell Ca2+ transients (CaT) in HL-1 cardiomyocytes were studied. To define toxic threshold levels, we performed dose-dependent transfection of TPM1 variants. In addition, cardiomyocyte structure was studied in human cardiac biopsies with TPM1 variants. Results - Overexpression of TPM1 variants led to time-dependent progressive deterioration of CaT, with the smallest effect seen for E62Q and larger and similar effects seen for the T201M and M281T variants. Overexpression of E62Q/M281T did not exacerbate the effects seen with overexpression of a single TPM1 variant. T201M (DCM) replaced endogenous tropomyosin dose-dependently, while M281T (HCM) did not. Human cardiac biopsies with TPM1 variants revealed loss of sarcomeric structures. Conclusion - All TPM1 variants result in reduced cardiomyocyte CaT amplitudes and loss of sarcomeric structures. These effects may underlie pathophysiology of different cardiomyopathy phenotypes.

Author Correction: The phospholamban p.(Arg14del) pathogenic variant leads to cardiomyopathy with heart failure and is unresponsive to standard heart failure therapy.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Publisher Correction: Relevance of Titin Missense and Non-Frameshifting Insertions/Deletions Variants in Dilated Cardiomyopathy.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The phospholamban p.(Arg14del) pathogenic variant leads to cardiomyopathy with heart failure and is unreponsive to standard heart failure therapy.

Phospholamban (PLN) plays a role in cardiomyocyte calcium handling as primary inhibitor of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA). The p.(Arg14del) pathogenic variant in the PLN gene results in a high risk of developing dilated or arrhythmogenic cardiomyopathy with heart failure. There is no established treatment other than standard heart failure therapy or heart transplantation. In this study, we generated a novel mouse model with the PLN-R14del pathogenic variant, performed detailed phenotyping, and tested the efficacy of established heart failure therapies eplerenone or metoprolol. Heterozygous PLN-R14del mice demonstrated increased susceptibility to ex vivo induced arrhythmias, and cardiomyopathy at 18 months of age, which was not accelerated by isoproterenol infusion. Homozygous PLN-R14del mice exhibited an accelerated phenotype including cardiac dilatation, contractile dysfunction, decreased ECG potentials, high susceptibility to ex vivo induced arrhythmias, myocardial fibrosis, PLN protein aggregation, and early mortality. Neither eplerenone nor metoprolol administration improved cardiac function or survival. In conclusion, our novel PLN-R14del mouse model exhibits most features of human disease. Administration of standard heart failure therapy did not rescue the phenotype, underscoring the need for better understanding of the pathophysiology of PLN-R14del-associated cardiomyopathy. This model provides a great opportunity to study the pathophysiology, and to screen for potential therapeutic treatments.

Homozygous damaging SOD2 variant causes lethal neonatal dilated cardiomyopathy.

BACKGROUND: Idiopathic dilated cardiomyopathy (DCM) is recognised to be a heritable disorder, yet clinical genetic testing does not produce a diagnosis in >50% of paediatric patients. Identifying a genetic cause is crucial because this knowledge can affect management options, cardiac surveillance in relatives and reproductive decision-making. In this study, we sought to identify the underlying genetic defect in a patient born to consanguineous parents with rapidly progressive DCM that led to death in early infancy. METHODS AND RESULTS: Exome sequencing revealed a potentially pathogenic, homozygous missense variant, c.542G>T, p.(Gly181Val), in SOD2. This gene encodes superoxide dismutase 2 (SOD2) or manganese-superoxide dismutase, a mitochondrial matrix protein that scavenges oxygen radicals produced by oxidation-reduction and electron transport reactions occurring in mitochondria via conversion of superoxide anion (O2-·) into H2O2. Measurement of hydroethidine oxidation showed a significant increase in O2-· levels in the patient's skin fibroblasts, as compared with controls, and this was paralleled by reduced catalytic activity of SOD2 in patient fibroblasts and muscle. Lentiviral complementation experiments demonstrated that mitochondrial SOD2 activity could be completely restored on transduction with wild type SOD2. CONCLUSION: Our results provide evidence that defective SOD2 may lead to toxic increases in the levels of damaging oxygen radicals in the neonatal heart, which can result in rapidly developing heart failure and death. We propose SOD2 as a novel nuclear-encoded mitochondrial protein involved in severe human neonatal cardiomyopathy, thus expanding the wide range of genetic factors involved in paediatric cardiomyopathies.

Arrhythmogenic Right Ventricular Cardiomyopathy-Associated Desmosomal Variants Are Rarely De Novo.

BACKGROUND: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is associated with pathogenic/likely pathogenic (P/LP) variants in genes encoding the cardiac desmosomal proteins. Origin of these variants, including de novo mutation rate and extent of founder versus recurrent variants has implications for variant adjudication and clinical care, yet this has never been systematically investigated. METHODS: We identified arrhythmogenic right ventricular cardiomyopathy probands who met 2010 Task Force Criteria and had undergone genotyping that included sequencing of the desmosomal genes (PKP2, DSP, DSG2, DSC2, and JUP) from 3 arrhythmogenic right ventricular cardiomyopathy registries in America and Europe. We classified the desmosomal variants, defined the contribution of unique versus nonunique (ie, not family-specific) P/LP variants, and identified the frequency and characteristics of de novo variants. Next, we haplotyped nonunique variants to determine how often they likely represent a single mutation event in a common ancestor (implied by shared haplotypes) versus multiple mutation events at the same genetic location. RESULTS: Of 501 arrhythmogenic right ventricular cardiomyopathy probands, 322 (64.3%) carried 327 desmosomal P/LP variants. Most variants (n=247, 75.6%, in 245 patients) were identified in more than one proband and, therefore, considered nonunique. For 212/327 variants (64.8%) genetic cascade screening was performed extensively enough to identify the parental origin of the P/LP variant. Only 3 variants were de novo, 2 of which were whole gene deletions. For 24 nonunique P/LP PKP2 variants, haplotyping was conducted in 183 available families. For all 24 variants, multiple seemingly unrelated families sharing identical haplotypes were identified, suggesting that these variants originate from common founders. CONCLUSIONS: Most desmosomal P/LP variants are inherited, nonunique, and originate from ancient founders. Two of 3 de novo variants were large deletions. These observations inform genetic testing, cascade screening, and variant adjudication.