Benchmarking computational variant effect predictors by their ability to infer human traits.

BACKGROUND: Computational variant effect predictors offer a scalable and increasingly reliable means of interpreting human genetic variation, but concerns of circularity and bias have limited previous methods for evaluating and comparing predictors. Population-level cohorts of genotyped and phenotyped participants that have not been used in predictor training can facilitate an unbiased benchmarking of available methods. Using a curated set of human gene-trait associations with a reported rare-variant burden association, we evaluate the correlations of 24 computational variant effect predictors with associated human traits in the UK Biobank and All of Us cohorts. RESULTS: AlphaMissense outperformed all other predictors in inferring human traits based on rare missense variants in UK Biobank and All of Us participants. The overall rankings of computational variant effect predictors in these two cohorts showed a significant positive correlation. CONCLUSION: We describe a method to assess computational variant effect predictors that sidesteps the limitations of previous evaluations. This approach is generalizable to future predictors and could continue to inform predictor choice for personal and clinical genetics.

Multiplexed Assays of Variant Effect and Automated Patch-clamping Improve KCNH2-LQTS Variant Classification and Cardiac Event Risk Stratification.

Background: Long QT syndrome (LQTS) is a lethal arrhythmia syndrome, frequently caused by rare loss-of-function variants in the potassium channel encoded by KCNH2. Variant classification is difficult, often owing to lack of functional data. Moreover, variant-based risk stratification is also complicated by heterogenous clinical data and incomplete penetrance. Here, we sought to test whether variant-specific information, primarily from high-throughput functional assays, could improve both classification and cardiac event risk stratification in a large, harmonized cohort of KCNH2 missense variant heterozygotes. Methods: We quantified cell-surface trafficking of 18,796 variants in KCNH2 using a Multiplexed Assay of Variant Effect (MAVE). We recorded KCNH2 current density for 533 variants by automated patch clamping (APC). We calibrated the strength of evidence of MAVE data according to ClinGen guidelines. We deeply phenotyped 1,458 patients with KCNH2 missense variants, including QTc, cardiac event history, and mortality. We correlated variant functional data and Bayesian LQTS penetrance estimates with cohort phenotypes and assessed hazard ratios for cardiac events. Results: Variant MAVE trafficking scores and APC peak tail currents were highly correlated (Spearman Rank-order ρ = 0.69). The MAVE data were found to provide up to pathogenic very strong evidence for severe loss-of-function variants. In the cohort, both functional assays and Bayesian LQTS penetrance estimates were significantly predictive of cardiac events when independently modeled with patient sex and adjusted QT interval (QTc); however, MAVE data became non-significant when peak-tail current and penetrance estimates were also available. The area under the ROC for 20-year event outcomes based on patient-specific sex and QTc (AUC 0.80 [0.76-0.83]) was improved with prospectively available penetrance scores conditioned on MAVE (AUC 0.86 [0.83-0.89]) or attainable APC peak tail current data (AUC 0.84 [0.81-0.88]). Conclusion: High throughput KCNH2 variant MAVE data meaningfully contribute to variant classification at scale while LQTS penetrance estimates and APC peak tail current measurements meaningfully contribute to risk stratification of cardiac events in patients with heterozygous KCNH2 missense variants.

Multifocal Ectopic Purkinje Premature Contractions due to neutralization of an SCN5A negative charge: structural insights into the gating pore hypothesis.

Background: We identified a novel SCN5A variant, E171Q, in a neonate with very frequent ectopy and reduced ejection fraction which normalized after arrhythmia suppression by flecainide. This clinical picture is consistent with multifocal ectopic Purkinje-related premature contractions (MEPPC). Most previous reports of MEPPC have implicated SCN5A variants such as R222Q that neutralize positive charges in the S4 voltage sensor helix of the channel protein NaV1.5 and generate a gating pore current. Methods and Results: E171 is a highly conserved negatively-charged residue located in the S2 transmembrane helix of NaV1.5 domain I. E171 is a key component of the Gating Charge Transfer Center, a region thought to be critical for normal movement of the S4 voltage sensor helix. We used heterologous expression, CRISPR-edited induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), and molecular dynamics simulations to demonstrate that E171Q generates a gating pore current, which was suppressed by a low concentration of flecainide (IC50 = 0.71±0.07 µM). R222Q shifts voltage dependence of activation and inactivation in a negative direction but we observed positive shifts with E171Q. E171Q iPSC-CMs demonstrated abnormal spontaneous activity and prolonged action potentials. Molecular dynamics simulations revealed that both R222Q and E171Q proteins generate a water-filled permeation pathway that underlies generation of the gating pore current. Conclusion: Previously identified MEPPC-associated variants that create gating pore currents are located in positively-charged residues in the S4 voltage sensor and generate negative shifts in the voltage dependence of activation and inactivation. We demonstrate that neutralizing a negatively charged S2 helix residue in the Gating Charge Transfer Center generates positive shifts but also create a gating pore pathway. These findings implicate the gating pore pathway as the primary functional and structural determinant of MEPPC and widen the spectrum of variants that are associated with gating pore-related disease in voltage-gated ion channels.

Detection of distant relatedness in biobanks for identification of undiagnosed carriers of a Mendelian disease variant: application to Long QT Syndrome.

Rare genetic diseases are typically studied in referral populations, resulting in underdiagnosis and biased assessment of penetrance and phenotype. To address this, we developed a generalizable method of genotype inference based on distant relatedness and deployed this to identify undiagnosed Type 5 Long QT Syndrome (LQT5) rare variant carriers in a non-referral population. We identified 9 LQT5 families referred to a single specialty clinic, each carrying p.Asp76Asn, the most common LQT5 variant. We uncovered recent common ancestry and a single shared haplotype among probands. Application to a non-referral population of 69,819 BioVU biobank subjects identified 22 additional subjects sharing this haplotype, subsequently confirmed to carry p.Asp76Asn. Referral and non-referral carriers had prolonged QTc compared to controls, and, among carriers, QTc polygenic score additively associated with QTc prolongation. Thus, our novel analysis of shared chromosomal segments identified undiagnosed cases of genetic disease and refined the understanding of LQT5 penetrance and phenotype.

Detection of distant familial relatedness in biobanks for identification of undiagnosed carriers of a Mendelian disease variant: application to Long QT syndrome.

Importance: The diagnosis and study of rare genetic disease is often limited to referral populations, leading to underdiagnosis and a biased assessment of penetrance and phenotype. Objective: To develop a generalizable method of genotype inference based on distant relatedness and to deploy this to identify undiagnosed Type 5 Long QT Syndrome (LQT5) rare variant carriers in a non-referral population. Participants: We identified 9 LQT5 probands and 3 first-degree relatives referred to a single Genetic Arrhythmia clinic, each carrying D76N (p.Asp76Asn), the most common variant implicated in LQT5. The non-referral population consisted of 69,879 ancestry-matched subjects in BioVU, a large biobank that links electronic health records to dense array data. Participants were enrolled from 2007-2022. Data analysis was performed in 2022. Exposures: We developed and applied a novel approach to genotype inference (Distant Relatedness for Identification and Variant Evaluation, or DRIVE) to identify shared, identical-by-descent (IBD) large chromosomal segments in array data. Main Outcomes and Measures: We sought to establish genetic relatedness among the probands and to use genomic segments underlying D76N to identify other potential carriers in BioVU. We then further studied the role of D76N in LQT5 pathogenesis. Results: Genetic reconstruction of pedigrees and distant relatedness detection among clinic probands using DRIVE revealed shared recent common ancestry and identified a single long shared haplotype. Interrogation of the non-referral population in BioVU identified a further 23 subjects sharing this haplotype, and sequencing confirmed D76N carrier status in 22, all previously undiagnosed with LQT5. The QTc was prolonged in D76N carriers compared to BioVU controls, with 40% penetrance of QTc ≥ 480 msec. Among D76N carriers, a QTc polygenic score was additively associated with QTc prolongation. Conclusions and Relevance: Detection of IBD shared chromosomal segments around D76N enabled identification of distantly related and previously undiagnosed rare-variant carriers, demonstrated the contribution of polygenic risk to monogenic disease penetrance, and further established LQT5 as a primary arrhythmia disorder. Analysis of shared chromosomal regions spanning disease-causing mutations can identify undiagnosed cases of genetic diseases.

Left atrial appendage dimension predicts elevated brain natriuretic peptide in nonvalvular atrial fibrillation.

INTRODUCTION: BNP elevation in patients with AF is observed in the absence of heart failure; however, prior mechanistic studies have not included direct left atrial pressure measurements. This study sought to understand how emptying function of the left atrial appendage (LAA) and LAA dimension contributes to brain-natriuretic peptide elevations (BNP) in atrial fibrillation (AF) accounting for left atrial pressure (LAP). METHODS: 132 patients referredfor left atrial appendage occlusion (LAAO) were prospectively enrolled in this study. BNP levels and LAP were measured just before LAAO. Statistical analysis considered BNP, rhythm at time of procedure, LAP, LAA morphology, LAA size (ostial diameter, depth, volume), LAA emptying velocity, CHADS2-VASc score, body mass index (BMI), left ventricular ejection fraction (LVEF), estimated glomerular filtration rate (eGFR), and obstructive sleep apnea (OSA) diagnosis as covariates. RESULTS: Bivariate statistical analysis demonstrated positive associations with age, LAA ostial diameter, depth, and volume, LAP, AF status at time of measurement, OSA, and CHADS2-VASc score. BNP was negatively associated with LVEF, eGFR, LAA emptying velocity and BMI. With multivariate logistic regression including LAP as covariate, significant relationships between BNP and AF/AFL(OR 1.99 [1.03, 3.85]), LAP (OR 1.13 [1.06, 1.20]), LAA diameter (OR 1.14 [1.03, 1.27]), LAA depth (OR 1.14 [1.07, 1.22]), and LAA emptying velocity (OR 0.97 [0.96,0.99]) were observed; however, no significant associations were seen with LAA morphology or CHADS2-VASc score. CONCLUSIONS: BNP elevations in AF are associated with LAA size and function, but not CHADS2-VASc score or appendage morphology after accounting for changes in LAP.

Effectiveness of Intense Accent Modification Training With Refugees From Burma.

PURPOSE: This pilot research project sought to determine if an intensive accent modification training program that included See the Sound-Visual Phonics and prosodic gestures improved articulation, prosody, and intelligibility measures in refugees from Burma. PARTICIPANTS: Four individuals (two men, two women) aged 20-67 participated in this study, and they were recruited from a state organization supporting refugees who have resettled in the United States. METHOD: All participants completed the Proficiency in Oral English Communication (POEC) and Assessment of Intelligibility of Dysarthric Speech (AIDS) to measure pre- and posttraining changes. The duration of this study was 6 weeks and consisted of 1 week of pretesting, 4 weeks of accent modification training, and 1 week of posttesting. Participants attended a total of twelve 50-min accent modification training sessions, including eight individual sessions (twice per week) and four group sessions (once per week), which provided a functional way to practice newly acquired skills in a scripted conversational-type format. Trained and untrained articulation and prosody probes were used to establish baselines and measure change. RESULTS: All four participants showed gains across articulation and prosody (in untrained and trained items). On pre- and posttest measures, three of the four participants also made gains on the broad measures of the AIDS and the POEC. CONCLUSION: Findings support that a brief and intensive multimodality accent modification program can be beneficial.

Arrhythmias as Presentation of Genetic Cardiomyopathy.

There is increasing evidence regarding the prevalence of genetic cardiomyopathies, for which arrhythmias may be the first presentation. Ventricular and atrial arrhythmias presenting in the absence of known myocardial disease are often labelled as idiopathic, or lone. While ventricular arrhythmias are well-recognized as presentation for arrhythmogenic cardiomyopathy in the right ventricle, the scope of arrhythmogenic cardiomyopathy has broadened to include those with dominant left ventricular involvement, usually with a phenotype of dilated cardiomyopathy. In addition, careful evaluation for genetic cardiomyopathy is also warranted for patients presenting with frequent premature ventricular contractions, conduction system disease, and early onset atrial fibrillation, in which most detected genes are in the cardiomyopathy panels. Sudden death can occur early in the course of these genetic cardiomyopathies, for which risk is not adequately tracked by left ventricular ejection fraction. Only a few of the cardiomyopathy genotypes implicated in early sudden death are recognized in current indications for implantable cardioverter defibrillators which otherwise rely upon a left ventricular ejection fraction ≤0.35 in dilated cardiomyopathy. The genetic diagnoses impact other aspects of clinical management such as exercise prescription and pharmacological therapy of arrhythmias, and new therapies are coming into clinical investigation for specific genetic cardiomyopathies. The expansion of available genetic information and implications raises new challenges for genetic counseling, particularly with the family member who has no evidence of a cardiomyopathy phenotype and may face a potentially negative impact of a genetic diagnosis. Discussions of risk for both probands and relatives need to be tailored to their numeric literacy during shared decision-making. For patients presenting with arrhythmias or cardiomyopathy, extension of genetic testing and its implications will enable cascade screening, intervention to change the trajectory for specific genotype-phenotype profiles, and enable further development and evaluation of emerging targeted therapies.

A challenging VT ablation with a large cardiac tumor.

INTRODUCTION: Ventricular tachycardia (VT) normally occurs from an abnormal structural substrate. CASE: We report a case in which VT was caused by a large tumor in the interventricular septum. Surgical intervention was not an option due to the location of the tumor and its proximity to the coronary arteries. CONCLUSION: The patient underwent ablation and upgrade to CRT before ultimately receiving a heart transplant.

Phenotypic Clustering of Left Ventricular Diastolic Function Parameters: Patterns and Prognostic Relevance.

OBJECTIVES: This study sought to explore the natural clustering of echocardiographic variables used for assessing left ventricular (LV) diastolic dysfunction (DD) in order to isolate high-risk phenotypic patterns and assess their prognostic significance. BACKGROUND: Assessment of LV DD is important in the management and prognosis of cardiovascular diseases. Data-driven approaches such as cluster analysis may be useful in segregating similar cases without the constraint of an a priori algorithm for risk stratification. METHODS: The study included a convenience sample of 866 consecutive patients referred for myocardial function assessment (age 65 ± 17 years; 55.3% women; ejection fraction 60 ± 9%) for whom echocardiographic parameters of DD assessment were obtained per conventional guideline recommendations. Unsupervised, hierarchical cluster analysis of these parameters was conducted using the Ward linkage method. Major adverse cardiovascular events, hospitalization, and mortality were compared between conventional and cluster-based classifications. RESULTS: Clustering algorithms for screening the presence of DD in 559 of 866 patients identified 2 distinct groups and revealed modest agreement with conventional classification (kappa = 0.41, p < 0.001). Further cluster analysis in 387 patients with DD helped to classify the severity of DD into 2 groups, with good agreement with conventional classification (kappa = 0.619, p < 0.001). Survival analyses of patients assessed by both clustering algorithms for screening and grading DD showed improved prediction of event-free survival by clusters over conventional classification for all-cause mortality and cardiac mortality, even after accounting for a multivariable, balanced propensity score. CONCLUSIONS: An unsupervised assessment of echocardiographic variables for assessing LV DD revealed unique patterns of grouping. These natural patterns of clustering may better identify patient groups who have similar risk, and their incorporation into clinical practice may help eliminate indeterminate results and improve clinical outcome prediction.