Twinning International Pediatric Cardiology Fellowship Programs: A Transformative Educational Experience for Trainees with Potential for Global Adoption.

Over the last decade, having endured the COVID-19 pandemic, education and training in pediatric cardiology have undergone a profound disruptive transformation. Trainees experience considerable stress achieving all the competencies required to become a competent pediatric cardiologist. Often the quality of the training experienced by trainees, the approach to patients, and potential institutional preference in management strategy is heavily influenced by the center in which they train. We developed an online live twin program of education between Texas Children's Hospital, Houston, Texas and Children's Health at Crumlin Dublin Ireland in 2019. We explored using grounded theory whether a regular scheduled shared teaching program improved fellow education and training between both centers. Trainees were surveyed to evaluate the benefits and disadvantages of such a twin program. The majority (93%) found the sessions helpful from an educational standpoint with many trainees reporting it to be a transformative experience. Three important learning themes emerged: practice variation between centers, managing uncertainty in clinical practice and cognitive overload. This pedagogical model could be replicated across multiple international pediatric cardiology units and facilitate "collaborative learning" among centers across the globe. Furthermore, this novel educational model could also be adopted by other medical specialties.

Efficacy of RyR2 inhibitor EL20 in induced pluripotent stem cell-derived cardiomyocytes from a patient with catecholaminergic polymorphic ventricular tachycardia.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited cardiac arrhythmia syndrome that often leads to sudden cardiac death. The most common form of CPVT is caused by autosomal-dominant variants in the cardiac ryanodine receptor type-2 (RYR2) gene. Mutations in RYR2 promote calcium (Ca2+ ) leak from the sarcoplasmic reticulum (SR), triggering lethal arrhythmias. Recently, it was demonstrated that tetracaine derivative EL20 specifically inhibits mutant RyR2, normalizes Ca2+ handling and suppresses arrhythmias in a CPVT mouse model. The objective of this study was to determine whether EL20 normalizes SR Ca2+ handling and arrhythmic events in induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) from a CPVT patient. Blood samples from a child carrying RyR2 variant RyR2 variant Arg-176-Glu (R176Q) and a mutation-negative relative were reprogrammed into iPSCs using a Sendai virus system. iPSC-CMs were derived using the StemdiffTM kit. Confocal Ca2+ imaging was used to quantify RyR2 activity in the absence and presence of EL20. iPSC-CMs harbouring the R176Q variant demonstrated spontaneous SR Ca2+ release events, whereas administration of EL20 diminished these abnormal events at low nanomolar concentrations (IC50  = 82 nM). Importantly, treatment with EL20 did not have any adverse effects on systolic Ca2+ handling in control iPSC-CMs. Our results show for the first time that tetracaine derivative EL20 normalized SR Ca2+ handling and suppresses arrhythmogenic activity in iPSC-CMs derived from a CPVT patient. Hence, this study confirms that this RyR2-inhibitor represents a promising therapeutic candidate for treatment of CPVT.

TBX5-encoded T-box transcription factor 5 variant T223M is associated with long QT syndrome and pediatric sudden cardiac death.

Long QT syndrome (LQTS) is a genetic disease resulting in a prolonged QT interval on a resting electrocardiogram, predisposing affected individuals to polymorphic ventricular tachycardia and sudden death. Although a number of genes have been implicated in this disease, nearly one in four individuals exhibiting the LQTS phenotype are genotype-negative. Whole-exome sequencing identified a missense T223M variant in TBX5 that cosegregates with prolonged QT interval in a family with otherwise genotype-negative LQTS and sudden death. The TBX5-T223M variant was absent among large ostensibly healthy populations (gnomAD) and predicted to be pathogenic by in silico modeling based on Panther, PolyPhen-2, Provean, SIFT, SNAP2, and PredictSNP prediction tools. The variant was located in a highly conserved region of TBX5 predicted to be part of the DNA-binding interface. A luciferase assay identified a 57.5% reduction in the ability of TBX5-T223M to drive expression at the atrial natriuretic factor promotor compared to wildtype TBX5 in vitro. We conclude that the variant is pathogenic in this family, and we put TBX5 forward as a disease susceptibility allele for genotype-negative LQTS. The identification of this familial variant may serve as a basis for the identification of previously unknown mechanisms of LQTS with broader implications for cardiac electrophysiology.

Smaller right pulmonary artery is associated with longer survival time without scimitar vein repair.

INTRODUCTION: The optimal management of scimitar syndrome remains incompletely defined. We (1) evaluated the impact of aortopulmonary collateral (APC) occlusion, (2) compared outcomes according to surgical approach for patients who underwent surgery, and (3) identified anatomic factors associated with longer survival time without scimitar vein repair. METHODS: We conducted a single center, retrospective study of 61 patients diagnosed with scimitar syndrome between 1995 and 2019. Right pulmonary artery to total pulmonary artery cross-sectional area (RPA:PA CSA) quantitatively assessed right pulmonary artery size. Anatomical features were analyzed for association with longer survival time without scimitar vein repair. RESULTS: Median follow-up time was 6 years (Q1-Q3, 2-12), with 96% 5-year survival. Twenty-three patients underwent APC occlusion, which significantly decreased symptoms of overcirculation (100%-46%; p = .001) and systolic pulmonary artery pressure (median, 34-29 mmHg; p = .004). Twenty-three patients underwent scimitar vein repair; 5-year freedom from scimitar vein stenosis was 90% among patients who underwent a reimplantation compared with 42% in patients with baffle repair (p = .1). Three patients underwent surgery before the first year of age, with lower 5-year freedom from scimitar vein stenosis (0% vs. 84%; p < .001). On multivariate analysis, a lower RPA:PA CSA was associated with longer survival time without scimitar vein repair (p = .003). CONCLUSIONS: APC occlusion improves the clinical status of young and hemodynamically unstable patients. Repair at an early age is associated with an increased risk of scimitar vein stenosis. Scimitar vein repair might be avoided in patients with a smaller right pulmonary artery.

Meta-analysis of cardiomyopathy-associated variants in troponin genes identifies loci and intragenic hot spots that are associated with worse clinical outcomes.

INTRODUCTION: Troponin (TNN)-encoded cardiac troponins (Tn) are critical for sensing calcium and triggering myofilament contraction. TNN variants are associated with development of cardiomyopathy; however, recent advances in genetic analysis have identified rare population variants. It is unclear how certain variants are associated with disease while others are tolerated. OBJECTIVE: To compare probands with TNNT2, TNNI3, and TNNC1 variants and utilize high-resolution variant comparison mapping of pathologic and rare population variants to identify loci associated with disease pathogenesis. METHODS: Cardiomyopathy-associated TNN variants were identified in the literature and topology mapping conducted. Clinical features were compiled and compared. Rare population variants were obtained from the gnomAD database. Signal-to-noise (S:N) normalized pathologic variant frequency against population variant frequency. Abstract review of clinical phenotypes was applied to "significant" hot spots. RESULTS: Probands were compiled (N = 70 studies, 224 probands) as were rare variants (N = 125,748 exomes; 15,708 genomes, MAF <0.001). TNNC1-positive probands demonstrated the youngest age of presentation (20.0 years; P = .016 vs TNNT2; P = .004 vs TNNI3) and the highest death, transplant, or ventricular fibrillation events (P = .093 vs TNNT2; P = .024 vs TNNI3; Kaplan Meir: P = .025). S:N analysis yielded hot spots of diagnostic significance within the tropomyosin-binding domains, α-helix 1, and the N-Terminus in TNNT2 with increased sudden cardiac death and ventricular fibrillation (P = .004). The inhibitory region and C-terminal region in TNNI3 exhibited increased restrictive cardiomyopathy (P =.008). HCM and RCM models tended to have increased calcium sensitivity and DCM decreased sensitivity (P < .001). DCM and HCM studies typically showed no differences in Hill coefficient which was decreased in RCM models (P < .001). CM models typically demonstrated no changes to Fmax (P = .239). CONCLUSION: TNNC1-positive probands had younger ages of diagnosis and poorer clinical outcomes. Mapping of TNN variants identified locations in TNNT2 and TNNI3 associated with heightened pathogenicity, RCM diagnosis, and increased risk of sudden death.

Bridging the Cardiac Needs of a Large, Underserved Immigrant and Resettled Refugee Population.

OBJECTIVE: To describe a monthly outreach pediatric cardiology clinic established to better understand the cardiac needs of immigrant/resettled refugee children. STUDY DESIGN: Data obtained between 2014 and 2017 from a monthly pediatric cardiology clinic at a Federally Qualified Health Center were analyzed using descriptive statistics. RESULTS: A total of 366 patients (222 male, 61%) were evaluated. Indications for referral included murmur (242, 66%), nonexertional symptoms (31, 9%), exertional symptoms (16, 4%), history of cardiac surgery/transcatheter interventions (15, 4%), previous diagnosis of heart conditions without intervention (13, 4%), arrhythmia/bradycardia (13, 4%), and others (36, 10%). Echocardiograms were performed on 136 patients (67 were abnormal, 49%). The most common final diagnoses include innocent murmur in 201 (55%), simple congenital heart disease in 61 (16%), complex congenital heart disease in 3 (1%), and acquired heart disease in 3 (1%). A total of 15 patients (4%) were ultimately determined to require surgical or cardiac catherization as an intervention. Patients have been followed for a median of 0.7 years (range 0-3.3 years). CONCLUSIONS: Rates of abnormal echocardiograms suggest a greater likelihood of congenital or acquired heart disease at time of initial consultation compared with nonimmigrant/refugee populations. The most common indication for referral to the outreach pediatric cardiology clinic was a murmur. Collaborative efforts between physicians and support services are essential in assisting this vulnerable population access pediatric subspecialty care.

A genome-wide association study of congenital cardiovascular left-sided lesions shows association with a locus on chromosome 20.

Congenital heart defects involving left-sided lesions (LSLs) are relatively common birth defects with substantial morbidity and mortality. Previous studies have suggested a high heritability with a complex genetic architecture, such that only a few LSL loci have been identified. We performed a genome-wide case-control association study to address the role of common variants using a discovery cohort of 778 cases and 2756 controls. We identified a genome-wide significant association mapping to a 200 kb region on chromosome 20q11 [P= 1.72 × 10-8 for rs3746446; imputed Single Nucleotide Polymorphism (SNP) rs6088703 P= 3.01 × 10-9, odds ratio (OR)= 1.6 for both]. This result was supported by transmission disequilibrium analyses using a subset of 541 case families (lowest P in region= 4.51 × 10-5, OR= 1.5). Replication in a cohort of 367 LSL cases and 5159 controls showed nominal association (P= 0.03 for rs3746446) resulting in P= 9.49 × 10-9 for rs3746446 upon meta-analysis of the combined cohorts. In addition, a group of seven SNPs on chromosome 1q21.3 met threshold for suggestive association (lowest P= 9.35 × 10-7 for rs12045807). Both regions include genes involved in cardiac development-MYH7B/miR499A on chromosome 20 and CTSK, CTSS and ARNT on chromosome 1. Genome-wide heritability analysis using case-control genotyped SNPs suggested that the mean heritability of LSLs attributable to common variants is moderately high ([Formula: see text] range= 0.26-0.34) and consistent with previous assertions. These results provide evidence for the role of common variation in LSLs, proffer new genes as potential biological candidates, and give further insight to the complex genetic architecture of congenital heart disease.

Copy Number Variants of Undetermined Significance Are Not Associated with Worse Clinical Outcomes in Hypoplastic Left Heart Syndrome.

OBJECTIVE: To determine the prevalence, spectrum, and prognostic significance of copy number variants of undetermined significance (cnVUS) seen on chromosomal microarray (CMA) in neonates with hypoplastic left heart syndrome (HLHS). STUDY DESIGN: Neonates with HLHS who presented to Texas Children's Hospital between June 2008 and December 2016 were identified. CMA results were abstracted and compared against copy number variations (CNVs) in ostensibly healthy individuals gathered from the literature. Findings were classified as normal, consistent with a known genetic disorder, or cnVUS. Survival was then compared using Kaplan-Meier analysis. Secondary outcomes included tracheostomy, feeding tube at discharge, cardiac arrest, and extracorporeal membrane oxygenation (ECMO). RESULTS: Our study cohort comprised 105 neonates with HLHS, including 70 (66.7%) with normal CMA results, 9 (8.6%) with findings consistent with a known genetic disorder, and 26 (24.7%) with a cnVUS. Six of the 26 (23.0%) neonates with a cnVUS had a variant that localized to a specific region of the genome seen in the healthy control population. One-year survival was 84.0% in patients with a cnVUS, 68.3% in those with normal CMA results, and 33.3% in those with a known genetic disorder (P = .003). There were no significant differences in secondary outcomes among the groups, although notably ECMO was used in 15.7% of patients with normal CMA and was not used in those with cnVUS and abnormal results (P = .038). CONCLUSIONS: Among children with HLHS, cnVUSs detected on CMA are common. The cnVUSs do not localize to specific regions of the genome, and are not associated with worse outcomes compared with normal CMA results.

Eteplirsen for the treatment of Duchenne muscular dystrophy.

OBJECTIVE: In prior open-label studies, eteplirsen, a phosphorodiamidate morpholino oligomer, enabled dystrophin production in Duchenne muscular dystrophy (DMD) with genetic mutations amenable to skipping exon 51. The present study used a double-blind placebo-controlled protocol to test eteplirsen's ability to induce dystrophin production and improve distance walked on the 6-minute walk test (6MWT). METHODS: DMD boys aged 7 to 13 years, with confirmed deletions correctable by skipping exon 51 and ability to walk 200 to 400 m on 6 MWT, were randomized to weekly intravenous infusions of 30 or 50 mg/kg/wk eteplirsen or placebo for 24 weeks (n = 4/group). Placebo patients switched to 30 or 50 mg/kg eteplirsen (n=2/group) at week 25; treatment was open label thereafter. All patients had muscle biopsies at baseline and week 48. Efficacy included dystrophin-positive fibers and distance walked on the 6MWT. RESULTS: At week 24, the 30 mg/kg eteplirsen patients were biopsied, and percentage of dystrophin-positive fibers was increased to 23% of normal; no increases were detected in placebo-treated patients (p≤0.002). Even greater increases occurred at week 48 (52% and 43% in the 30 and 50 mg/kg cohorts, respectively), suggesting that dystrophin increases with longer treatment. Restoration of functional dystrophin was confirmed by detection of sarcoglycans and neuronal nitric oxide synthase at the sarcolemma. Ambulation-evaluable eteplirsen-treated patients experienced a 67.3 m benefit compared to placebo/delayed patients (p≤0.001). INTERPRETATION: Eteplirsen restored dystrophin in the 30 and 50 mg/kg/wk cohorts, and in subsequently treated, placebo-controlled subjects. Duration, more than dose, accounted for dystrophin production, also resulting in ambulation stability. No severe adverse events were encountered.

Signal-to-Noise Analysis Can Inform the Likelihood That Incidentally Identified Variants in Sarcomeric Genes Are Associated with Pediatric Cardiomyopathy.

Background: Hypertrophic cardiomyopathy (HCM) is the most common heritable cardiomyopathy and can predispose individuals to sudden death. Most pediatric HCM patients host a known pathogenic variant in a sarcomeric gene. With the increase in exome sequencing (ES) in clinical settings, incidental variants in HCM-associated genes are being identified more frequently. Diagnostic interpretation of incidental variants is crucial to enhance clinical patient management. We sought to use amino acid-level signal-to-noise (S:N) analysis to establish pathogenic hotspots in sarcomeric HCM-associated genes as well as to refine the 2015 American College of Medical Genetics (ACMG) criteria to predict incidental variant pathogenicity. Methods and Results: Incidental variants in HCM genes (MYBPC3, MYH7, MYL2, MYL3, ACTC1, TPM1, TNNT2, TNNI3, and TNNC1) were obtained from a clinical ES referral database (Baylor Genetics) and compared to rare population variants (gnomAD) and variants from HCM literature cohort studies. A subset of the ES cohort was clinically evaluated at Texas Children's Hospital. We compared the frequency of ES and HCM variants at specific amino acid locations in coding regions to rare variants (MAF < 0.0001) in gnomAD. S:N ratios were calculated at the gene- and amino acid-level to identify pathogenic hotspots. ES cohort variants were re-classified using ACMG criteria with S:N analysis as a correlate for PM1 criteria, which reduced the burden of variants of uncertain significance. In the clinical validation cohort, the majority of probands with cardiomyopathy or family history hosted likely pathogenic or pathogenic variants. Conclusions: Incidental variants in HCM-associated genes were common among clinical ES referrals, although the majority were not disease-associated. Leveraging amino acid-level S:N as a clinical tool may improve the diagnostic discriminatory ability of ACMG criteria by identifying pathogenic hotspots.