Identification of Genetic Variants in Patients with Idiopathic Ventricular Arrhythmia by Taiwan Arrhythmia Gene Panel Implying Underlying Mechanisms

Identification of Genetic Variants in Patients with Idiopathic Ventricular Arrhythmia by Taiwan Arrhythmia Gene Panel Implying Underlying Mechanisms | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Identification of Genetic Variants in Patients with Idiopathic Ventricular Arrhythmia by Taiwan Arrhythmia Gene Panel Implying Underlying Mechanisms Jien Jiun Chen, Sheng-Nan Chang, Fu-Chun Chiu, Pang-Shuo Huang, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4458113/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background Sudden cardiac death (SCD) due to idiopathic ventricular tachycardia or ventricular fibrillation is a catastrophic disease. Its genetic basis is heterogeneous and has been rarely addressed in Asia. We aim to find variants in an Asian cohort of idiopathic ventricular arrhythmia (IVA). Methods Nationwide patients with IVA were consecutively recruited. We designed a SCD gene panel (134 genes) to detect variants by next-generation sequencing (NGS) including most of the channelopathy and cardiomyopathy genes. Results A total of 40 IVA patients were included. Thirteen variants with unknown significance (VUS) and 7 pathogenic/likely pathogenic (P/LP) variants were identified in 20 patients (50%). All variants were novel and not found in dbSNP, ExAC and our general population. The identified variants were in genes implicated for long QT or Brugada syndromes (SCN5A, KCNH2, CACNA1C and ANK2), cardiomyopathy (MYH6, DSP and TTN) and catecholaminergic polymorphic ventricular tachycardia (RYR2). Patients with P/LP were younger, and more were women than those with VUS. Conclusions A high yield rate of genetic test was found in the largest NGS cohort of Asian IVA patients. These patients should be vigorously followed up for possible channelopathy or cardiomyopathy with repeated provocative test and myocardial imaging. sudden arrhythmia death genetic test idiopathic ventricular arrhythmia next-generation sequencing variants Figures Figure 1 Introduction Cardiac arrest due to idiopathic ventricular tachycardia (IVT) or idiopathic ventricular fibrillation (IVF), naming idiopathic ventricular arrhythmia (IVA) in the absence of structural heart disease and other identifiable causes of ventricular arrhythmia is a catastrophic disease. Patients die suddenly and unexpectedly as the initial presentation of the disease, which is called sudden cardiac death (SCD).(1) Up to 11% of SCD survivors recurred within 1 year of the diagnosis. Importantly, there is no clinical predictor for IVA, and it is very difficult to identify or even treat these patients before SCD occurs. Therefore, identification of genetic predictor(s) is critical. Alders et al reported up to 20% of probands with this disorder had a family history, suggesting that at least a subset of IVA is hereditary.(2) Earlier candidate gene approach and functional studies have demonstrated sodium channel as one of the contributions to the risk of developing familial IVA.(3-5) Though there was no report of potassium channel involvement in IVA, KCNE5 variants had been reported as a modulator.(6) With the evolvement of technology, using a genome-wide haplotype-sharing analysis, Alders et al had identified a haplotype, on chromosome 7q36, harboring part of the DPP6 gene, increased DPP6 mRNA levels in the myocardium of carriers.(2) Followed up study revealed that DPP6 gain of function selectively enhancing Purkinje fiber Ito current and might play a role of arrhythmogenesis. Another whole exome sequencing study identified ACTN2 gene variant segregated with the disease in a family.(7) Finally, the latest report of calmodulin mutation had been reported to be the causative variant in families with young-onset IVA.(8) Therefore, multiple genes may underlie the mechanism of IVA. All these studies have been focusing on familial cases. However, clinically we have met many sporadic cases without family history. Sporadic IVA is also more unpredictable, and identification of genetic risk factors is of paramount importance. However, so far, the genetic study for these sporadic IVA without family history has been rarely reported before. We hypothesize that the genetic architecture of sporadic IVA is like familial IVA because of their similar presentation pattern or phenotypes. As mentioned above rare variants are usually the genetic causes of familial disease (e.g., familial IVA). It has also been demonstrated that rare variants are also usually the genetic causes of rare sporadic cases with severe phenotypes (e.g., sporadic IVA) because rare variants usually have a large effect on phenotype.(9) For example, based on this hypothesis, Ng et al. have identified disease-causing genes that were enriched for rare variants in a very small number of patients with extremely rare diseases (4 patients with Miller syndrome).(10) Therefore, the diseasing causing variants may be rare variants for those patients with sporadic IVA. Recently, parallelized high-throughput technologies, e.g., next-generation sequencing (NGS), have become available to efficiently sequence large segments of DNA, including the processes of whole genome or whole exome sequencing to identify disease-causing variants or rare variants.(11) However, in-depth whole genome or exome sequencing may reveal a significantly large number of possible variants, even with a very small number of cases. As such, sequencing candidate genes might be a more reasonable approach, which raises the important issue of how to select the candidate genes a priori . Therefore, we sought to perform a candidate gene approach in 40 unrelated patients with documented IVA. Accordingly, we used NGS to do a deep sequencing of the coding regions of our self-established arrhythmia or SCD gene panel. We also performed in silico functional studies for these identified rare variants.(12) The result of the present proposed study may help identify disease-causing variants of IVA in Taiwan and may shed light on the possible disease mechanism of IVA. Methods Study population This nationwide registry recruited patients with ventricular arrhythmia. The clinical diagnosis of IVA was based on history of ventricular arrhythmia (ventricular fibrillation or hemodynamic unstable ventricular tachycardia, excluding ventricular tachycardia ordinated from outflow tract or fascicles.) or aborted sudden death due to ventricular arrhythmia, negative myocardial image studies (echocardiography and/or cardiac magnetic resonance imaging(CRM)), negative coronary angiography and electrocardiographic (ECG) findings (resting, provocative or exercise) of known causes of SCD by certified electrophysiologists in our country according to the HRS/ACC/AHA(13) or EHRA/APHRS(14) guidelines. Oral flecainide loading test (oral flecainide 400 mg loading and standard and high-lead ECGs were performed at baseline, 30 mins, every hour till 12 hours and 24 hours after oral flecainide)(15) was arranged for those with ECG suspicious of Brugada syndrome (BrS) and intravenous epinephrine test (initial 0.025 ug/kg/min and increased to 0.05, 0.1 and 0.2 ug/kg/min, ECGs were performed at baseline, 5 mins after each dose increase, and 5 and 10 minutes after discontinuation of epinephrine, and the difference between the minimal and maximal QTc during infusion 30 ms was considered positive) for long QT syndrome (LQTS).(16) Toxicological screening was arranged as individual physician’s discretion because illegal drug use has been rarely the cause of SCD in our Taiwanese population. All patients or his delegates signed informed consent for genetic analyses. The study complied with declaration of Helsinki and was approved by Institution Review Board of National Taiwan University Hospital (201705122RINC). DNA extraction and next-generation exon sequencing Genomic DNA was extracted as described previously(17-19). For the generation of standard exome capture libraries, we used the Agilent SureSelect XT Reagent kit protocol for Illumina Hiseq paired-end sequencing library (catalog# G9611A). In all cases, the SureSelect XT Human All Exon Version 4 +UTR (71Mb) probe set was used. We sonicated 3 μg genomic DNA to 150-200bp, then constructed a library with Agilent SureSelect XT Reagent kit. The amplification adapter-ligated sample was purified using Agencourt AMPure XP beads (Beckman Coulter; Brea, CA, USA) and analyzed on a Bioanalyzer DNA1000 chip. Five hundred nanograms of the sample were prepared for hybridization with the capture baits, and the sample was hybridized for 24 h at 65°C, captured with the Dynabeads MyOne Streptavidin T1 (Life Technologies; USA), and purified using Agencourt AMPure XP beads. Finally, we pooled samples together for sequencing by Illumina Hiseq and achieved 100-fold sequence coverage of targeted exonic regions. The sequence reads were aligned to human genome hg19 (GRCh37) with DNAnexus software (Palo Alto, CA) using default parameters. Establishment of the Taiwanese Arrhythmia-Sudden Cardiac Death Gene Panel and Variant Classification We did a deep search of all the published literature and have selected candidate genes related to cardiac arrhythmia or SCD for our Arrhythmia-Sudden Cardiac Death Gene Panel . (20,21) We updated the LQTS genes as core LQTS genes ( KCNQ1, KCNH2, SCN5A ), genes with strong or definitive evidence for causality in LQTS ( CALM1, CALM2, CALM3, TRDN ) and one gene with moderate level evidence ( CACNA1C ).(22) We also include genes that are identified by our own genetic studies and may be unique to our Taiwanese populations. First, in our preliminary and unpublished genetic study for ventricular arrhythmia, we found BAZ2B was associated with ventricular arrhythmia in our Taiwanese population. Second, in Taiwan, we found KCNIP1 gene was highly associated with atrial fibrillation (AF).(23) It is well-known that some SCD patients also have AF and SCD may be associated with AF. Therefore, we included BAZ2B and KCNIP 1 to 4 in our SCD gene panel. The candidate genes are summarized in the supplemental data. The variant classification was done according to the American College of Medical Genetics (ACMG) guidelines.(24) We focused on variants that might destroy the protein function, such as changing the amino acid sequence (mis-sense mutation), truncating or premature stop mutations. Furthermore, when an identified variant could not be found in the Taiwanese population (1000 normal subjects), this indicated that this variant might be a novel mutation that was only specific or private to the affected proband. Finally, we also attempted to make sure the identified novel sequence variations or variants had also never been reported in the public references, such as dbSNP (http://www.ncbi.nlm.nih.gov/projects/SNP/) or ExAC.(25) DNA sequencing by Sanger method To verify the sequence variation identified by NGS, all variants were sequence-verified by conventional Sanger sequencing, as described previously.(17,18) Prediction of the functional significance of the identified variants For the variants within the proximal promoters (exon sequencing covers some lengths of the 5’UTR), we mapped the location of the mutation. We also investigated whether the mutation localized in the regulatory regions which were hypersensitive to DNase I, associated with transcriptional factor bindings, or epigenetic modification,(26,27) because it has been demonstrated that those regions are transcriptionally active and are important to determine the transcriptional activity of the downstream gene. For the variants within the 5UTR but not in the promoter (un-translated region in the first exon), we also focused on whether the mutation localized in the DNase I hypersensitive region. For the variants within the coding sequences, we used the PolyPhen-2 program to predict whether the variants resulted in a damage of the structure of the encoded protein. PolyPhen-2 is an automatic tool for prediction of possible impact of an amino acid substitution on the structure and function of a human protein. This prediction is based on a number of features comprising the sequence, phylogenetic and structural information characterizing the substitution. A variant is appraised based on pairs of false positive rate thresholds. Variants with their posterior probability scores associated with estimated false positive rates at or below the false positive rate value are predicted to be damaging. Variants with estimated false positive rates above the false positive rate value are classified as benign. If the lack of data does not allow to make a prediction, the outcome is reported as unknown. Statistical methods All data were expressed as mean ± standard deviation. Continued data from independent group were compared using unpaired Student t test and categorical data using chi square test. A p value or post-hoc p value < 0.05 was considered statistically significant. Results Clinical characteristics A total of 40 patients with IVA were prospectively enrolled. The mean age was 63.8±10.9 and 82.5% (33/40) were men. The clinical presentation was 17 patients with Out of hospital cardiac arrest (OHCA)/SCD, 12 with pre-syncope/syncope and 11 non-syncopal symptoms such as severe palpitation, weakness, dizziness, diaphoresis, or chest tightness, i.e. the ventricular arrhythmia compromised the patients’ hemodynamics. Three patients had family history of SCD. In the resting ECGs, the PQ interval was ≤200 ms and QTc ≤ 480 ms in all patients. The mean QTc was 419 ± 33 ms. Based on the results of echocardiography, the left ventricular ejection fraction was ≥50% in all patients either after the episode or during the follow-up. All the patients received coronary angiography and no significant coronary stenosis or coronary anomaly was found. Identification of novel variants A total of 20 novel variants or variants were identified in 20 patients. According to the American College of Medical Genetics and Genomics (ACMG) guidelines (24), 7 variants were classified as pathogenic or likely pathogenic (P/LP) variants and 13 as variants of uncertain significance (VUSs). Table 1 shows the clinical characteristics of the patients and all the variants. All of these novel non-synonymous exon variants are heterozygous with only one mutant allele in each locus. We did not find any rare variant or mutation in those 5 proposed Taiwan-specific SCD genes ( BAZ2B and KCNIP 1 to 4 ) among all of our study subjects. Table 1. Novel variants identified by next-generation sequencing in an Asian cohort of patients with idiopathic ventricular arrhythmia No. Age Gender CoM LVEF (%) Zygosity Gene Variant type Mutation class CV Medication ECG 1 74 F HTN 68.6 Het NEXN VUS missense Amio, CB RBBB 2 73 M HTN 73.1 Het FHL2 VUS missense Amio, ARB NP 3 67 M HPL 70.1 Het TTN VUS missense ARB, Amio NP 4 64 M - 63.1 Het TTN VUS missense ARB, Amio NP 5 59 M - 62.8 Het KCNH2 P/LP missense BB NP 6 66 M - 58.4 Het TTN VUS missense ARB, BB, Amio Diffuse TWI 7 64 M - 69.5 Het TTN VUS missense ARB, BB, Amio TWI in V1-3 8 71 M HTN 77.8 Het SLC8A1 VUS missense BB, ARB NP 9 60 M - 61.5 Het ANK2 VUS missense BB, Propa AF 10 67 M HTN, HPL 58.3 Het NUP155 VUS missense BB, CB 1 ° AV Block, diffuse TWF 11 52 F - 62.1 Het MYH6 P/LP missense BB, CB, ARB NP 12 60 F HPL 78.8 Het CACNA1H P/LP missense BB, ARB diffuse TWI 13 75 M HTN, DM 73.6 Het RBM20 VUS missense Amio, ARB, CB AF 14 60 M - 63.1 Het CACNA1H P/LP missense CB, BB, ARB NP 15 41 M - 70.0 Het CACNA1I VUS missense CB, ARB NP 16 46 F - 69.7 Het CACNA1C P/LP missense CB, BB NP 17 59 M HPL 77.5 Het DSP VUS missense Amio NP 18 47 F - 67.5 Het SCN5A P/LP missense BB NP 19 45 M - 72.4 Het RYR2 VUS missense BB NP 20 54 M - 75.9 Het RYR2 P/LP missense BB NP Abbreviations: AF, atrial fibrillation; Amio, amiodarone; ARB, angiotensin receptor blocker; AVB, atrioventricular conduction block; BB, beta blocker; Het, heterozygous; CB, calcium channel blocker; CoM, comorbidity; CV, cardiovascular; DM, diabetes mellitus; HPL, hyperlipidemia; HTN, hypertension; LVEF, left ventricular ejection fraction; NP, nothing particular; P/LP, pathogenic or likely pathogenic variant; Propa, propafenone; RBBB, right bundle branch block; TWF, T wave flattening; TWI, T wave inversion; VUS, variant with uncertain significance. We have verified all these important variants by conventional Sanger sequencing. We have also confirmed that these variants were not found in the Taiwanese general population (Taiwan Biobank), dbSNP and ExAC. which indicates that these variants are novel. We have inferred whether these variants might result in any functional change of the expressed genes or encoded proteins by in silico prediction of protein structural change due to an amino acid change by using Polyphen2 and SIFT, and revealed all these variants are damaging or probable damaging. Patients with KCNH2 , SCN5A or CACNA1C mutation also received epinephrine test for possible long QT syndrome (LQTS), and all were negative. Patients with SCN5A or CACNA1C mutation also received oral flecainide loading test for possible BrS, and all were also negative. Pathogenic and likely pathogenic variants Seven P/LP variants were identified in our IVA cohort (Table 1), and all these variants are novel. Two patients carried two novel missense variants in the heart failure related SCD gene CACNA1H , and one patient in LQTS-associated KCNH2 gene. One patient carried a novel missense mutation in the hypertrophic cardiomyopathy (HCM)-related MYH6 gene and one in the ARVC (arrhythmogenic right ventricular cardiomyopathy)-related DSP gene. Two patients carried a novel missense mutation in the Brugada-related SCN5A or CACNA1C gene and one in the catecholaminergic polymorphic ventricular tachycardia (CPVT)-related RYR2 gene. The demographics of patients carrying P/LP variants were significant different from those with VUS. The patients with P/LP were younger (54.0±6.0 vs 63.5±10.4 years, P=0.039), and more were women (4/7 vs 1/13, P=0.031). The illustration summarizing the study designs and conclusion could be found in Figure 1. Discussion In this nationwide cohort of patients with IVA, we identified several novel genes and variants, which have never been reported before and include ionic channel genes such as SCN5A, KCNH2 , CACNA1C , and CACNA1H and cardiac myofilaments or connexin genes such as MYBPC3, MYH6, NEXN , TTN and DSP genes. Genetic studies of idiopathic ventricular fibrillation (IVF) or unexplained SCD had been conducted in various populations. The positive rate are 1.6 % to 23% (28,29) in the Asian population, and 0 % to 10% (30-32) in the Caucasian populations. The VUS positive rate may be up to 20% in a recent population of genetics of Caucasian IVF patients by Pannone et al.(33) The genes identified were highly variable. The common genes in IVF are CCR7 and PKN2 , SCN5A , KCNQ1 and RYR2 (29,34) in the Asian populations, and CALM1~3 , RYR2 , TRDN , CACNA1C , SCN5A , KCNE5 in the Caucasian populations.(35) Searching for the cause of unexplained SCD is a great challenge.(32) In the current era, genetic test or molecular autopsy may provide some implications. A high yield rate of genetic test was found in our NGS cohort of Asian IVA patients. Around 18% of our IVA patients had P/LP variants, which is similar to that reported by Lahrouchi et al., in which molecular autopsy (genetic test) for electrical disorder and cardiomyopathy genes, using ACMG guidelines for variant classification, identified a modest but realistic yield in patients with sudden arrhythmic death syndrome (SADS)(13% of patients had P/LP variants).(36) Importantly, we also found the important roles of CPVT and LQTS genes in unexplained SCD as in the CASPER study.(20) Fang et al. conducted the first genome-wide association study of idiopathic ventricular tachycardia only in the Chinese Han population.(29) They identified 4 novel loci and 2 risk genes ( PKN2 and CCR7 ) for idiopathic ventricular tachycardia, implicating the role of cardiac calcium homeostasis in rhythm maintenance. However, whether these genes also play a role in IVF or unexplained SCD remains unknown. In addition to BrS, genetic variants in SCN5A and CACNA1C may also be implicated in early repolarization syndrome (ERS). ERS is a newly identified entity with J wave elevation in inferior leads.(37,38) Many different variants had been reported, probably due to heterogeneity of the disease. In our patients with these variants, no J wave elevation or early repolarization was noted in screening ECG. However, J wave elevation or early repolarization is a dynamic process and more close monitoring of ECG may reveal them. In our patients, some have LQTS gene variants such as KCNH2, SCN5A, and CACNA1C . If these patients do have LQTS, it indicates the limitation of the diagnostic value of traditional ECG, especially resting ECG. The QT interval may change with drug challenge or epinephrine and exercise tests, but the sensitivity and specificity are not well documented.(16,39) Whether a molecular or genetic diagnosis of LQTS is feasible without ECG confirmation remains unknown, especially in the Asian or our Taiwanese populations. According to the guidelines, an unknown SCD patients should be diagnosed with LQTS if a pathogenic variant is found in major LQTS genes, even the QT or QTc interval is not prolonged. In Taiwan or Asia, the incidence of LQTS may be lower compared to the Caucasian populations(40-43) but the rates of background or incidental pathogenic variants of LQTS genes in our Taiwanese population are relatively high (especially SCN5A and KCNH2 genes).(44) Therefore, if we use the sole criteria of having LQTS gene pathogenic variant to diagnose LQTS, there will be much more LQTS patients in Taiwan, which is really not the case. Nevertheless, all our patients with LQTS gene mutations had undergone ICD implantation. Furthermore, empirical use of beta blocker may not be harmful, and may also be advantageous in patients with unknown cause of SCD. Therefore, all these patients received beta blocker therapy. They will also be closely followed up for the possibility of LQTS. There are also overlaps of genes responsible for different hereditary arrhythmia, and it is also possible these patients with LQTS gene mutations might have BrS or ERS. The ECG manifestation of these diseases may change in different circumstances, such as fever unmasking Brudaga pattern.(45) Again, close follow-up of these patients is mandatory. We also found two patients having CACNA1H variants. CACNA1H mutation has not been reported in genetic SCD. CACNA1H encodes for the alpha subunit of T type calcium channel, which has been implicated to play a possible role in the mechanism of ventricular arrhythmia in the setting of heart failure.(46) Somatic mutation of CACNA1H has also been found to cause primary aldosteronism.(47) MYH6 gene encodes for the cardiac contractile filament protein, mutation of which causes HCM with myocyte disarray, ventricular hypertrophy, and increased susceptibility to ventricular arrhythmia.(48) TTN gene encodes for a very large protein called Titin,(49) which functions as a molecular spring responsible for passive elasticity of muscle.(49) TTN truncating variants are a common cause of dilated cardiomyopathy, occurring in approximately 25% of genetic dilated cardiomyopathy and in 18% of sporadic cases.(50,51) In our patients with IVA, although no structural heart disease had been identified, this did not exclude the possibility that these patients would eventually develop cardiomyopathy. It is also possible that arrhythmia precedes structural changes, and the patients may later manifest structural change in the heart. This finding was similar to that reported by Isbister et al. looking at genetic causes of cardiomyopathy in patients with autopsy negative sudden death, also suggesting that the earliest presentation of cardiomyopathy could be arrhythmic events.(52) This warrants periodic surveillance of myocardial imaging and function. Whether treatment for specific cardiomyopathy should be instituted in these patients remain unknown. Patients with HCM may have favorable response to beta blocker which is also helpful for a wide spectrum of cause of SCD.(53) Finally, it is also possible that the genetic variant was just a by-stander, based on the high rate of incidental variant in cardiac myosin genes in our Taiwanese population.(44) We could not rule out the possibility that the high positive rate of genetic test in our study is due to many genes were tested simultaneously. In those patients with frank phenotypes, such as HCM or LQTS, only meaningful target genes will be tested. In patients with IVA, we did not pre-specify genes or select genes in priori , thus many genes related to SCD (such as our SCD panel) were tested which might generate many unrelated variants, especially VUS. In addition to this Asian study, VUS was also found in a high proportion of Caucasian patients with unexplained SCD.(20,33) Interpreting VUS is usually a great challenge and dilemma either in patients with unexplained SCD or in those with clinical diagnosis confirmed SCD case (such as a MYH7 VUS in a clinically confirmed LQTS patient). But VUS implicating a SCD syndrome in a patient with unexplained SCD or implicating the other cause of SCD in a patient with clinically confirmed SCD case may encourage the physician to follow-up the patient more closely and also to perform molecular functional studies of this variant to define its role in SCD. Nevertheless, the rates of VUS or P/LP variants were significantly higher in our IVA patients than those of incidental variants in our Taiwanese general population.(44) This result may imply that the identified variants do mean something and are not incidental. Finally, we stress the importance of complete work-up before diagnosing a patient with IVF or the genetic test results will be very misleading and confusing. In this study, we worked up these patients according to the reported SCD guidelines to avoid as many occult or concealed SCD syndromes included in our study population as possible. Most importantly, the genetic findings might stimulate re-evaluation of the original clinical data and possibly lead to a renewed clinical diagnosis.(54) There are limitations in the present study. First, there was no cascade family screening done in this study, either clinical screening or genetic screening. Phenotypes corresponding to the identified variant in other asymptomatic family members may confirm the role of the identified variant. Co-segregation of variant may also confirm the role of identified variants. All the patients received history taking and most of them reported no family history of SCD or unexplained syncope, probably because of the nature of the Asian people not willing to reveal detailed family history, particular those histories related to severe diseases. Second, our IVA patients were significantly older compared to other studies. It is possible that it is easier to make a definite diagnosis in Asian young patients with SCD, such as BrS, or our IVA population represents a milder form of ventricular arrhythmia manifesting late or at older age. Third, because the cost of genetic test was covered by the research grant and nearly all the patients agreed to receive a genetic test, we believed in our study there was no selection bias, such as those with a more severe phenotype more intending to take the genetic test. Finally, not all our patients received sodium channel blocker provocative test and we also only used oral flecainide loading test as the provocative test. Also, not all patients received CMR examination because early in this study, CMR was not a routine for unexplained SCD. Therefore, there might be concealed BrS or cardiomyopathy patients in our study subjects. That is the reason why we used IVA but not IVF in our study. In conclusion, in this largest IVA cohort with NGS genetic testing in Asia, we identified genes coding for channelopathy or cardiomyopathy that are not clinically manifested. Because of high yield rate of variants in our IVA patients, genetic test may be considered in Asian patients with IVA. Periodic surveillance for myocardial morphology and function is important in these patients. Repeated provocative test could also be considered. Abbreviations ACMG: American College of Medical Genetics AF: atrial fibrillation ARVC: arrhythmogenic right ventricular cardiomyopathy BrS: Brudaga syndrome CMR: cardiac magnetic resonance imaging CPVT: catecholaminergic polymorphic ventricular tachycardia ECG: electrocardiography ERS: early repolarization syndrome HCM: hypertrophic cardiomyopathy IVA: idiopathic ventricular arrhythmia IVT: idiopathic ventricular tachycardia IVF: idiopathic ventricular fibrillation LQTS: long QT syndrome NGS: next-generation sequencing OHCA: Out of hospital cardiac arrest P/LP: pathogenic/likely pathogenic SADS: sudden arrhythmic death syndrome SCD: sudden cardiac death VUS: variants of uncertain significance Declarations Author contribution statement: Jien-Jiun Chen, designed the study, collected patients’ data, and wrote the manuscript. Seng-Nan Chang, Fu-Chun Chiu, Pang-Shuo Huang, Cho-Kai Wu, Yi-Chih Wang, collected patients’ data and gave a critical review of the manuscript. Juey-Jen Hwang and Chia-Ti Tsai designed the study and genetic panel, in charge of the whole study. Funding This study was supported by a grant from the National Science and Technology Council (NSTC)(105-2314-B-002-153-MY2). Acknowledgments We would thank all the patients and intensivists who cared these patients This research has been conducted using data from UK Biobank (www.ukbiobank.ac.uk), a major biomedical database, approved under project # 54423. All authors have read the journal's policy on disclosure of potential conflicts of interest. All authors have disclosed any financial or personal relationship with organizations that could potentially be perceived as influencing the described research. Conflict of interests All authors declared no competing interests. Data availability The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request. Ethics statements Ethics approval This study involves human participants and was approved by the Research ethics committee of National Taiwan University Hospital (201705122RINC). Written informed consent was provided by all participants/legal guardians. References Viskin S, Belhassen B. Idiopathic ventricular fibrillation. Am Heart J 1990;120:661–71. Alders M, Koopmann TT, Christiaans I et al. Haplotype-sharing analysis implicates chromosome 7q36 harboring DPP6 in familial idiopathic ventricular fibrillation. Am J Hum Genet 2009;84:468–76. Chen Q, Kirsch GE, Zhang D et al. Genetic basis and molecular mechanism for idiopathic ventricular fibrillation. Nature 1998;392:293–6. Wan X, Chen S, Sadeghpour A, Wang Q, Kirsch GE. Accelerated inactivation in a mutant Na(+) channel associated with idiopathic ventricular fibrillation. Am J Physiol Heart Circ Physiol 2001;280:H354-60. Valdivia CR, Medeiros-Domingo A, Ye B et al. Loss-of-function mutation of the SCN3B-encoded sodium channel {beta}3 subunit associated with a case of idiopathic ventricular fibrillation. Cardiovasc Res 2010;86:392–400. Ohno S, Zankov DP, Ding WG et al. KCNE5 (KCNE1L) variants are novel modulators of Brugada syndrome and idiopathic ventricular fibrillation. Circ Arrhythm Electrophysiol 2011;4:352–61. Bagnall RD, Molloy LK, Kalman JM, Semsarian C. Exome sequencing identifies a mutation in the ACTN2 gene in a family with idiopathic ventricular fibrillation, left ventricular noncompaction, and sudden death. BMC Med Genet 2014;15:99. Nomikos M, Thanassoulas A, Beck K et al. Altered RyR2 regulation by the calmodulin F90L mutation associated with idiopathic ventricular fibrillation and early sudden cardiac death. FEBS Lett 2014;588:2898–902. Manolio TA, Collins FS, Cox NJ et al. Finding the missing heritability of complex diseases. Nature 2009;461:747–53. Ng SB, Buckingham KJ, Lee C et al. Exome sequencing identifies the cause of a mendelian disorder. Nat Genet 2010;42:30–5. Cirulli ET, Goldstein DB. Uncovering the roles of rare variants in common disease through whole-genome sequencing. Nat Rev Genet 2010;11:415–25. Chen YH, Pai CW, Huang SW et al. Inactivation of Myosin binding protein C homolog in zebrafish as a model for human cardiac hypertrophy and diastolic dysfunction. J Am Heart Assoc 2013;2:e000231. Al-Khatib SM, Stevenson WG, Ackerman MJ et al. 2017 AHA/ACC/HRS Guideline for Management of Patients With Ventricular Arrhythmias and the Prevention of Sudden Cardiac Death: Executive Summary: A Report of the American College of Cardiology/American Heart Association Task Force on Clinical Practice Guidelines and the Heart Rhythm Society. J Am Coll Cardiol 2018;72:1677–1749. Pedersen CT, Kay GN, Kalman J et al. EHRA/HRS/APHRS expert consensus on ventricular arrhythmias. Heart Rhythm 2014;11:e166-96. Dubner S, Azocar D, Gallino S et al. Single oral flecainide dose to unmask type 1 Brugada syndrome electrocardiographic pattern. Ann Noninvasive Electrocardiol 2013;18:256–61. Vyas H, Hejlik J, Ackerman MJ. Epinephrine QT stress testing in the evaluation of congenital long-QT syndrome: diagnostic accuracy of the paradoxical QT response. Circulation 2006;113:1385–92. Chang SN, Tsai CT, Wu CK et al. A functional variant in the promoter region regulates the C-reactive protein gene and is a potential candidate for increased risk of atrial fibrillation. J Intern Med 2012;272:305–15. Tsai CT, Lai LP, Lin JL et al. Renin-angiotensin system gene polymorphisms and atrial fibrillation. Circulation 2004;109:1640–6. Chiang FT, Hsu KL, Tseng CD et al. Molecular variant M235T of the angiotensinogen gene is associated with essential hypertension in Taiwanese. J Hypertens 1997;15:607–11. Grondin S, Davies B, Cadrin-Tourigny J et al. Importance of genetic testing in unexplained cardiac arrest. Eur Heart J 2022;43:3071–3081. Hellenthal N, Gaertner-Rommel A, Klauke B et al. Molecular autopsy of sudden unexplained deaths reveals genetic predispositions for cardiac diseases among young forensic cases. Europace 2017;19:1881–1890. Adler A, Novelli V, Amin AS et al. An International, Multicentered, Evidence-Based Reappraisal of Genes Reported to Cause Congenital Long QT Syndrome. Circulation 2020;141:418–428. Tsai CT, Hsieh CS, Chang SN et al. Genome-wide screening identifies a KCNIP1 copy number variant as a genetic predictor for atrial fibrillation. Nat Commun 2016;7:10190. Erikson GA, Deshpande N, Kesavan BG, Torkamani A. SG-ADVISER CNV: copy-number variant annotation and interpretation. Genet Med 2015;17:714–8. Karczewski KJ, Weisburd B, Thomas B et al. The ExAC browser: displaying reference data information from over 60 000 exomes. Nucleic Acids Res 2017;45:D840-D845. Thurman RE, Rynes E, Humbert R et al. The accessible chromatin landscape of the human genome. Nature 2012;489:75–82. Mercer TR, Edwards SL, Clark MB et al. DNase I-hypersensitive exons colocalize with promoters and distal regulatory elements. Nat Genet 2013;45:852–9. Son MK, Ki CS, Park SJ, Huh J, Kim JS, On YK. Genetic mutation in Korean patients of sudden cardiac arrest as a surrogating marker of idiopathic ventricular arrhythmia. J Korean Med Sci 2013;28:1021–6. Fang C, Wang P, Yu D et al. Genome-Wide Association Study for Idiopathic Ventricular Tachyarrhythmias Identifies Key Role of CCR7 and PKN2 in Calcium Homeostasis and Cardiac Rhythm Maintenance. Circ Genom Precis Med 2022;15:e003603. Herman AR, Cheung C, Gerull B et al. Outcome of Apparently Unexplained Cardiac Arrest: Results From Investigation and Follow-Up of the Prospective Cardiac Arrest Survivors With Preserved Ejection Fraction Registry. Circ Arrhythm Electrophysiol 2016;9:e003619. Bai R, Napolitano C, Bloise R, Monteforte N, Priori SG. Yield of genetic screening in inherited cardiac channelopathies: how to prioritize access to genetic testing. Circ Arrhythm Electrophysiol 2009;2:6–15. Visser M, van der Heijden JF, Doevendans PA, Loh P, Wilde AA, Hassink RJ. Idiopathic Ventricular Fibrillation: The Struggle for Definition, Diagnosis, and Follow-Up. Circ Arrhythm Electrophysiol 2016;9. Pannone L, Gauthey A, Conte G et al. Genetics in Probands With Idiopathic Ventricular Fibrillation: A Multicenter Study. JACC Clin Electrophysiol 2023;9:1296–1306. Zhong X, Guo W, Wei J et al. Identification of loss-of-function RyR2 mutations associated with idiopathic ventricular fibrillation and sudden death. Biosci Rep 2021;41. Wang M, Tu X. The Genetics and Epigenetics of Ventricular Arrhythmias in Patients Without Structural Heart Disease. Front Cardiovasc Med 2022;9:891399. Lahrouchi N, Raju H, Lodder EM et al. Utility of Post-Mortem Genetic Testing in Cases of Sudden Arrhythmic Death Syndrome. J Am Coll Cardiol 2017;69:2134–2145. Nademanee K, Haissaguerre M, Hocini M et al. Mapping and Ablation of Ventricular Fibrillation Associated with Early Repolarization Syndrome. Circulation 2019. Mahapatra SR, Chakraborty P. An update on early repolarization(ER) syndrome. Indian Pacing Electrophysiol J 2015;15:265–7. Churet M, Luttoo K, Hocini M, Haissaguerre M, Sacher F, Duchateau J. Diagnostic reproducibility of epinephrine drug challenge interpretation in suspected long QT syndrome. J Cardiovasc Electrophysiol 2019;30:896–901. Yoshinaga M, Kucho Y, Nishibatake M, Ogata H, Nomura Y. Probability of diagnosing long QT syndrome in children and adolescents according to the criteria of the HRS/EHRA/APHRS expert consensus statement. Eur Heart J 2016;37:2490–7. Uhm JS, Hwang IU, Oh YS et al. Prevalence of electrocardiographic findings suggestive of sudden cardiac death risk in 10,867 apparently healthy young Korean men. Pacing Clin Electrophysiol 2011;34:717–23. Hayashi K, Fujino N, Uchiyama K et al. Long QT syndrome and associated gene mutation carriers in Japanese children: results from ECG screening examinations. Clin Sci (Lond) 2009;117:415–24. Murakoshi N, Aonuma K. Epidemiology of arrhythmias and sudden cardiac death in Asia. Circ J 2013;77:2419–31. Huang PS, Hsieh CS, Chang SN et al. Prevalence of sudden arrhythmic death syndrome-related genetic mutations in an Asian cohort of whole genome sequence. Europace 2020;22:1287–1297. Argenziano M, Antzelevitch C. Recent advances in the treatment of Brugada syndrome. Expert Rev Cardiovasc Ther 2018;16:387–404. Kinoshita H, Kuwahara K, Takano M et al. T-type Ca2 + channel blockade prevents sudden death in mice with heart failure. Circulation 2009;120:743–52. Nanba K, Blinder AR, Rege J et al. Somatic CACNA1H Mutation As a Cause of Aldosterone-Producing Adenoma. Hypertension 2020;75:645–649. Carrier L, Mearini G, Stathopoulou K, Cuello F. Cardiac myosin-binding protein C (MYBPC3) in cardiac pathophysiology. Gene 2015;573:188–97. Labeit S, Kolmerer B. Titins: giant proteins in charge of muscle ultrastructure and elasticity. Science 1995;270:293–6. Herman DS, Lam L, Taylor MR et al. Truncations of titin causing dilated cardiomyopathy. N Engl J Med 2012;366:619–28. Fang HJ, Liu BP. Prevalence of TTN mutations in patients with dilated cardiomyopathy: A meta-analysis. Herz 2019. Isbister JC, Nowak N, Yeates L et al. Concealed Cardiomyopathy in Autopsy-Inconclusive Cases of Sudden Cardiac Death and Implications for Families. J Am Coll Cardiol 2022;80:2057–2068. Drazner MH. Angiotensin Receptor-Neprilysin Inhibition (ARNI) Therapy and Reverse Remodeling in Heart Failure With Reduced Ejection Fraction. JAMA 2019:1–3. Landstrom AP, Dailey-Schwartz AL, Rosenfeld JA et al. Interpreting Incidentally Identified Variants in Genes Associated With Catecholaminergic Polymorphic Ventricular Tachycardia in a Large Cohort of Clinical Whole-Exome Genetic Test Referrals. Circ Arrhythm Electrophysiol 2017;10. Additional Declarations No competing interests reported. Supplementary Files SupplementaldataR1.docx Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4458113","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":310200213,"identity":"c3d0000f-0a72-42b4-98b4-32ffb0803ba2","order_by":0,"name":"Jien Jiun Chen","email":"","orcid":"","institution":"National Taiwan University Hospital Yun-Lin Branch","correspondingAuthor":false,"prefix":"","firstName":"Jien","middleName":"Jiun","lastName":"Chen","suffix":""},{"id":310200214,"identity":"77af5920-33ce-4166-aeb2-cf5b65e8f47b","order_by":1,"name":"Sheng-Nan Chang","email":"","orcid":"","institution":"National Taiwan University Hospital Yun-Lin Branch","correspondingAuthor":false,"prefix":"","firstName":"Sheng-Nan","middleName":"","lastName":"Chang","suffix":""},{"id":310200215,"identity":"b5e76a59-3695-4758-a185-8047ab23138b","order_by":2,"name":"Fu-Chun Chiu","email":"","orcid":"","institution":"National Taiwan University Hospital Yun-Lin Branch","correspondingAuthor":false,"prefix":"","firstName":"Fu-Chun","middleName":"","lastName":"Chiu","suffix":""},{"id":310200216,"identity":"ea9d968b-79c0-40cf-ada5-7719d52581bf","order_by":3,"name":"Pang-Shuo Huang","email":"","orcid":"","institution":"National Taiwan University Hospital Yun-Lin Branch","correspondingAuthor":false,"prefix":"","firstName":"Pang-Shuo","middleName":"","lastName":"Huang","suffix":""},{"id":310200217,"identity":"da10c5fa-5d36-49b2-82f3-d2bea1e70218","order_by":4,"name":"Cho-Kai Wu","email":"","orcid":"","institution":"National Taiwan University Hospital","correspondingAuthor":false,"prefix":"","firstName":"Cho-Kai","middleName":"","lastName":"Wu","suffix":""},{"id":310200218,"identity":"25543bdc-e5ef-4ed5-b953-f93e22afa02d","order_by":5,"name":"Yi-Chih Wang","email":"","orcid":"","institution":"National Taiwan University Hospital","correspondingAuthor":false,"prefix":"","firstName":"Yi-Chih","middleName":"","lastName":"Wang","suffix":""},{"id":310200219,"identity":"6e1c250b-9507-4239-a28d-419e4cfedb41","order_by":6,"name":"Juey-Jen Hwang","email":"","orcid":"","institution":"National Taiwan University College of Medicine and Hospital","correspondingAuthor":false,"prefix":"","firstName":"Juey-Jen","middleName":"","lastName":"Hwang","suffix":""},{"id":310200220,"identity":"dc905d9f-1d04-4189-a9d0-f7e403eac523","order_by":7,"name":"Chia-Ti Tsai","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA4ElEQVRIiWNgGAWjYNACAwk5fvbmA0CWhAxh1WzMIC0WxpI9xxJAWniI1MJQkbjhRo4BiEVYC//8/sMffhRIgLR8fnWjxoKHgf3w0Q34tEgcY2aT7DGQMJ555u0265xjQIfxpKXdwKfFgI2ZjYHHQEK273juNuMcNqAWCR4zQlqYP/4xkGBsOJDzzDjnH3FaGKSBtihOOJHD/Di3jQgtEseSzaRlgH4BBrIZc26fBA8bIb/wNx98/PHNnzpQVD7+nPMNxDh8DK8WZMAmASaJVQ4CzB9IUT0KRsEoGAUjBwAAQZ9DgJoFn90AAAAASUVORK5CYII=","orcid":"","institution":"National Taiwan University College of Medicine and Hospital","correspondingAuthor":true,"prefix":"","firstName":"Chia-Ti","middleName":"","lastName":"Tsai","suffix":""}],"badges":[],"createdAt":"2024-05-22 04:21:56","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4458113/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4458113/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":57953443,"identity":"8766c945-b07a-43e1-a726-2d9880fe20da","added_by":"auto","created_at":"2024-06-07 23:02:29","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":453475,"visible":true,"origin":"","legend":"\u003cp\u003eIn this large idiopathic ventricular arrhythmia (IVA) cohort with NGS genetic testing in Asia, we identified genes coding for channelopathy or cardiomyopathy that are not clinically manifested. Because of high yield rate of variants in our IVA patients, genetic test may be considered in Asian patients with IVA. Periodic surveillance for myocardial morphology and function is important in these patients. Repeated provocative test could also be considered.\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-4458113/v1/d067e31e80e7a9c19dc41b25.jpeg"},{"id":67523976,"identity":"99504cab-05f3-4381-9157-a0916c76cd3e","added_by":"auto","created_at":"2024-10-26 04:38:39","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":916625,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4458113/v1/6cf2f6b5-9878-4f0e-9814-f8e736a913f3.pdf"},{"id":57953442,"identity":"5784e214-2ff8-4940-8895-df18445ec19e","added_by":"auto","created_at":"2024-06-07 23:02:29","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":129164,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaldataR1.docx","url":"https://assets-eu.researchsquare.com/files/rs-4458113/v1/ee31a8e734e6b067fd66341c.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Identification of Genetic Variants in Patients with Idiopathic Ventricular Arrhythmia by Taiwan Arrhythmia Gene Panel Implying Underlying Mechanisms","fulltext":[{"header":"Introduction","content":"\u003cp\u003eCardiac arrest\u0026nbsp;due to idiopathic ventricular tachycardia (IVT) or idiopathic ventricular fibrillation (IVF), naming idiopathic ventricular arrhythmia (IVA)\u0026nbsp;in the absence of structural heart disease and other identifiable causes of ventricular\u0026nbsp;arrhythmia\u0026nbsp;is a catastrophic disease. Patients die suddenly and unexpectedly as the initial presentation of the disease, which is called\u0026nbsp;sudden cardiac death (SCD).(1)\u0026nbsp;Up to 11% of SCD survivors recurred within 1 year of the diagnosis. Importantly, there is no clinical predictor for IVA, and it is very difficult to identify or even treat these patients before SCD occurs. Therefore, identification of genetic predictor(s) is critical. Alders et al reported up to 20% of probands with this disorder had a family history, suggesting that at least a subset of IVA is hereditary.(2)\u003c/p\u003e\n\u003cp\u003eEarlier candidate gene approach and functional studies have demonstrated sodium channel as one of the contributions to the risk of developing familial\u0026nbsp;IVA.(3-5)\u0026nbsp;Though there was no report of potassium channel involvement in IVA, \u003cem\u003eKCNE5\u003c/em\u003e variants had been reported as a modulator.(6)\u0026nbsp;With the evolvement of technology, using a genome-wide haplotype-sharing analysis, Alders et al had identified a haplotype, on chromosome 7q36, harboring part of the DPP6 gene, increased \u003cem\u003eDPP6\u003c/em\u003e mRNA levels in the myocardium of carriers.(2)\u0026nbsp;Followed up study revealed that DPP6 gain of function selectively enhancing Purkinje fiber Ito current and might play a role of arrhythmogenesis. Another whole exome sequencing study identified \u003cem\u003eACTN2\u003c/em\u003e gene variant segregated with the disease in a family.(7)\u0026nbsp;Finally, the latest report of calmodulin mutation had been reported to be the causative variant in families with young-onset IVA.(8)\u0026nbsp;Therefore, multiple genes may underlie the mechanism of IVA.\u003c/p\u003e\n\u003cp\u003eAll these studies have been focusing on familial cases. However, clinically we have met many sporadic cases without family history. Sporadic IVA is also more unpredictable, and identification of genetic risk factors is of paramount importance. However, so far, the genetic study for these sporadic IVA without family history has been rarely reported before. We hypothesize that the genetic architecture of sporadic IVA is like familial IVA because of their similar presentation pattern or phenotypes. As mentioned above rare variants are usually the genetic causes of familial disease (e.g., familial IVA). It has also been demonstrated that rare variants are also usually the genetic causes of rare sporadic cases with severe phenotypes (e.g., sporadic IVA) because rare variants usually have a large effect on phenotype.(9)\u0026nbsp;For example, based on this hypothesis, Ng \u003cem\u003eet al.\u003c/em\u003e have identified disease-causing genes that were enriched for rare variants in a very small number of patients with extremely rare diseases (4 patients with Miller syndrome).(10)\u0026nbsp;Therefore, the diseasing causing variants may be rare variants for those patients with sporadic IVA.\u003c/p\u003e\n\u003cp\u003eRecently, parallelized high-throughput technologies, e.g., next-generation sequencing (NGS), have become available to efficiently sequence large segments of DNA, including the processes of whole genome or whole exome sequencing to identify disease-causing variants or rare variants.(11)\u0026nbsp;However, in-depth whole genome or exome sequencing may reveal a significantly large number of possible variants, even with a very small number of cases. As such, sequencing candidate genes might be a more reasonable approach, which raises the important issue of how to select the candidate genes \u003cem\u003ea priori\u003c/em\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTherefore, we\u0026nbsp;sought\u0026nbsp;to perform a candidate gene approach in\u0026nbsp;40\u0026nbsp;unrelated patients with documented IVA. Accordingly, we used NGS to do a deep sequencing of the coding regions of our self-established arrhythmia or SCD gene panel. We also performed\u003cem\u003e\u0026nbsp;in silico\u003c/em\u003e functional studies for these identified rare variants.(12) The result of the present proposed study may help identify disease-causing variants of IVA in Taiwan and may shed light on the possible disease mechanism of IVA.\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003e\u003cem\u003eStudy population\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThis nationwide registry recruited patients with ventricular arrhythmia. The clinical diagnosis of IVA was based on history of ventricular arrhythmia (ventricular fibrillation or hemodynamic unstable ventricular tachycardia, excluding ventricular tachycardia ordinated from outflow tract or fascicles.) or aborted sudden death due to ventricular arrhythmia, negative myocardial image studies (echocardiography and/or cardiac magnetic resonance imaging(CRM)), negative coronary angiography and electrocardiographic (ECG) findings (resting, provocative or exercise) of known causes of SCD by certified electrophysiologists in our country according to the HRS/ACC/AHA(13)\u0026nbsp;or EHRA/APHRS(14)\u0026nbsp;guidelines. Oral flecainide loading test (oral flecainide 400 mg loading and standard and high-lead ECGs were performed at baseline, 30 mins, every hour till 12 hours and 24 hours after oral flecainide)(15)\u0026nbsp;was arranged for those with ECG suspicious of Brugada syndrome (BrS) and intravenous epinephrine test (initial\u0026nbsp;0.025 ug/kg/min and increased to 0.05, 0.1 and 0.2 ug/kg/min, ECGs were performed at baseline, 5 mins after each dose increase, and 5 and 10 minutes after discontinuation of epinephrine, and the difference between the minimal and maximal QTc during infusion\u0026nbsp;30 ms was considered positive) for long QT syndrome (LQTS).(16)\u0026nbsp;Toxicological screening was arranged as individual physician\u0026rsquo;s discretion because illegal drug use has been rarely the cause of SCD in our Taiwanese population.\u0026nbsp;All patients or his delegates signed informed consent for genetic analyses. The study complied with declaration of Helsinki and was approved by Institution Review Board of National Taiwan University Hospital (201705122RINC).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eDNA extraction and next-generation exon sequencing\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eGenomic DNA was extracted as described previously(17-19). For the generation of standard exome capture libraries, we used the Agilent SureSelect XT Reagent kit protocol for Illumina Hiseq paired-end sequencing library (catalog# G9611A). In all cases, the SureSelect XT Human All Exon Version 4 +UTR (71Mb) probe set was used. We sonicated 3 \u0026mu;g genomic DNA to 150-200bp, then constructed a library with Agilent SureSelect XT Reagent kit. The amplification adapter-ligated sample was purified using Agencourt AMPure XP beads (Beckman Coulter; Brea, CA, USA) and analyzed on a Bioanalyzer DNA1000 chip. Five hundred nanograms of the sample were prepared for hybridization with the capture baits, and the sample was hybridized for 24 h at 65\u0026deg;C, captured with the Dynabeads MyOne Streptavidin T1 (Life Technologies; USA), and purified using Agencourt AMPure XP beads.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFinally, we pooled samples together for sequencing by Illumina Hiseq and achieved 100-fold sequence coverage of targeted exonic regions. The sequence reads were aligned to human genome hg19 (GRCh37) with DNAnexus software (Palo Alto, CA) using default parameters.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e\u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eEstablishment of the Taiwanese Arrhythmia-Sudden Cardiac Death Gene Panel and Variant Classification\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eWe did a deep search of all the published literature and have selected candidate genes related to cardiac arrhythmia or SCD for our \u003cem\u003eArrhythmia-Sudden Cardiac Death Gene Panel\u003c/em\u003e.\u003cem\u003e(20,21)\u003c/em\u003e We updated the LQTS genes as core LQTS genes (\u003cem\u003eKCNQ1, KCNH2, SCN5A\u003c/em\u003e), genes with strong or definitive evidence for causality in LQTS (\u003cem\u003eCALM1, CALM2, CALM3, TRDN\u003c/em\u003e) and one gene with moderate level evidence (\u003cem\u003eCACNA1C\u003c/em\u003e).(22)\u003c/p\u003e\n\u003cp\u003eWe also include genes that are identified by our own genetic studies and may be unique to our Taiwanese populations. First,\u0026nbsp;in our preliminary and unpublished genetic study for ventricular arrhythmia, we found \u003cem\u003eBAZ2B\u003c/em\u003e was associated with ventricular arrhythmia in our Taiwanese population. Second, in Taiwan, we found \u003cem\u003eKCNIP1\u0026nbsp;\u003c/em\u003egene was highly associated with atrial fibrillation (AF).(23)\u0026nbsp;It is well-known that some SCD patients also have AF and SCD may be associated with AF. Therefore, we included \u003cem\u003eBAZ2B\u003c/em\u003e and \u003cem\u003eKCNIP 1 to 4\u0026nbsp;\u003c/em\u003ein our SCD gene panel.\u003c/p\u003e\n\u003cp\u003eThe candidate genes are summarized in the supplemental data. The variant classification was done according to the American College of Medical Genetics (ACMG) guidelines.(24)\u0026nbsp;We focused on variants that might destroy the protein function, such as changing the amino acid sequence (mis-sense mutation), truncating or premature stop mutations. Furthermore, when an identified variant could not be found in the Taiwanese population (1000 normal subjects), this indicated that this variant might be a novel mutation that was only specific or private to the affected proband. Finally, we also attempted to make sure the identified novel sequence variations or variants had also never been reported in the public references, such as dbSNP (http://www.ncbi.nlm.nih.gov/projects/SNP/) or ExAC.(25)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eDNA sequencing by Sanger method\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eTo verify the sequence variation identified by NGS, all variants were sequence-verified by conventional Sanger sequencing, as described previously.(17,18)\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003ePrediction of the functional significance of the identified variants\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eFor the variants within the proximal promoters (exon sequencing covers some lengths of the 5\u0026rsquo;UTR), we mapped the location of the mutation. We also investigated whether the mutation localized in the regulatory regions which were hypersensitive to DNase I, associated with transcriptional factor bindings, or epigenetic modification,(26,27) because it has been demonstrated that those regions are transcriptionally active and are important to determine the transcriptional activity of the downstream gene. For the variants within the 5UTR but not in the promoter (un-translated region in the first exon), we also focused on whether the mutation localized in the DNase I hypersensitive region.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFor the variants within the coding sequences, we used the PolyPhen-2 program to predict whether the variants resulted in a damage of the structure of the encoded protein. PolyPhen-2 is an automatic tool for prediction of possible impact of an amino acid substitution on the structure and function of a human protein. This prediction is based on a number of features comprising the sequence, phylogenetic and structural information characterizing the substitution. A variant is appraised based on pairs of false positive rate thresholds. Variants with their posterior probability scores associated with estimated false positive rates at or below the false positive rate value are predicted to be damaging. Variants with estimated false positive rates above the false positive rate value are classified as benign. If the lack of data does not allow to make a prediction, the outcome is reported as unknown. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eStatistical methods\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eAll data were expressed as mean \u0026plusmn; standard deviation. Continued data from independent group were compared using unpaired Student \u003cem\u003et\u003c/em\u003e test and categorical data using chi square test. A p value or post-hoc p value \u0026lt; 0.05 was considered statistically significant.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cem\u003eClinical characteristics\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eA total of 40 patients with IVA were prospectively enrolled. The mean age was\u0026nbsp;63.8\u0026plusmn;10.9\u0026nbsp;and 82.5% (33/40) were men. The clinical presentation was 17 patients with Out of hospital cardiac arrest (OHCA)/SCD, 12 with pre-syncope/syncope and 11 non-syncopal symptoms such as severe palpitation, weakness, dizziness, diaphoresis, or chest tightness, i.e. the ventricular arrhythmia compromised the patients\u0026rsquo; hemodynamics. Three patients had family history of SCD.\u003c/p\u003e\n\u003cp\u003eIn the resting ECGs, the PQ interval was \u0026le;200 ms and QTc \u0026le; 480 ms in all patients. The mean QTc was 419 \u0026plusmn; 33 ms. Based on the results of echocardiography, the left ventricular ejection fraction was \u0026ge;50% in all patients either after the episode or during the follow-up. All the patients received coronary angiography and no significant coronary stenosis or coronary anomaly was found. \u003cem\u003e\u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eIdentification of novel variants\u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eA total of 20 novel variants or variants were identified in 20 patients. According to the\u0026nbsp;American College of Medical Genetics and Genomics\u0026nbsp;(ACMG) guidelines\u0026nbsp;(24), 7 variants were classified as pathogenic or likely pathogenic (P/LP) variants and 13 as variants of uncertain significance (VUSs).\u0026nbsp;Table 1 shows the clinical characteristics of the patients and all the variants. All of these novel non-synonymous exon variants are heterozygous with only one mutant allele in each locus.\u0026nbsp;We did not find any rare variant or mutation in those 5 proposed Taiwan-specific SCD genes (\u003cem\u003eBAZ2B\u003c/em\u003e and \u003cem\u003eKCNIP 1 to 4\u003c/em\u003e) among all of our study subjects.\u003c/p\u003e\n\u003cp\u003eTable 1. Novel variants identified by next-generation sequencing in an Asian cohort of patients with idiopathic ventricular arrhythmia\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"709\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003eNo.\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003eAge\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eGender\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eCoM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eLVEF (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003eZygosity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eGene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVariant type\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003eMutation class\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eCV Medication\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eECG\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e74\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eHTN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e68.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eNEXN\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eAmio, CB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eRBBB\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eHTN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e73.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eFHL2\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eAmio, ARB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eHPL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e70.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eTTN\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eARB, Amio\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e63.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eTTN\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eARB, Amio\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e59\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e62.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eKCNH2\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eP/LP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e66\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e58.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eTTN\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eARB, BB, Amio\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eDiffuse TWI\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e69.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eTTN\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eARB, BB, Amio\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eTWI in V1-3\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e71\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eHTN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e77.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eSLC8A1\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB, ARB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e61.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eANK2\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB, Propa\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eAF\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eHTN, HPL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e58.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eNUP155\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB, CB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003e\u003cem\u003e1\u003c/em\u003e\u0026deg; \u003cem\u003eAV Block,\u003c/em\u003e\u003cem\u003e\u0026nbsp;\u003c/em\u003ediffuse TWF\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e62.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eMYH6\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eP/LP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB, CB, ARB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eHPL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e78.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eCACNA1H\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eP/LP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB, ARB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003ediffuse TWI\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e75\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eHTN, DM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e73.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eRBM20\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eAmio, ARB, CB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eAF\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e63.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eCACNA1H\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eP/LP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eCB, BB, ARB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e41\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e70.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eCACNA1I\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eCB, ARB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e46\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e69.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eCACNA1C\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eP/LP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eCB, BB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e59\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003eHPL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e77.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eDSP\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eAmio\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e67.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eSCN5A\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eP/LP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e72.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eRYR2\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eVUS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9577464788732395%\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"3.8028169014084505%\"\u003e\n \u003cp\u003e54\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003e75.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.619718309859155%\"\u003e\n \u003cp\u003eHet\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.028169014084508%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eRYR2\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.492957746478874%\"\u003e\n \u003cp\u003eP/LP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.732394366197184%\"\u003e\n \u003cp\u003emissense\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.507042253521126%\"\u003e\n \u003cp\u003eBB\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.591549295774648%\"\u003e\n \u003cp\u003eNP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"2.9619181946403383%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"3.8081805359661494%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"6.6290550070521865%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"8.039492242595205%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"6.6290550070521865%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"6.6290550070521865%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"1.2693935119887165%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"6.6290550070521865%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"15.514809590973202%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"8.744710860366714%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"14.527503526093088%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd width=\"18.61777150916784%\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAbbreviations: AF, atrial fibrillation; Amio, amiodarone; ARB, angiotensin receptor blocker; AVB, atrioventricular conduction block; BB, beta blocker; Het, heterozygous; CB, calcium channel blocker; CoM, comorbidity; CV, cardiovascular; DM, diabetes mellitus; HPL, hyperlipidemia; HTN, hypertension; LVEF, left ventricular ejection fraction; NP, nothing particular; P/LP, pathogenic or likely pathogenic variant; Propa, propafenone; RBBB, right bundle branch block; TWF, T wave flattening; TWI, T wave inversion; VUS, variant with uncertain significance.\u003c/p\u003e\n\u003cp\u003eWe have verified all these important variants by conventional Sanger sequencing. We have also confirmed that these variants were not found in the Taiwanese general population (Taiwan Biobank), dbSNP and ExAC. which indicates that these variants are novel. We have inferred whether these variants might result in any functional change of the expressed genes or encoded proteins by\u003cem\u003e\u0026nbsp;in silico\u003c/em\u003e prediction of protein structural change due to an amino acid change by using Polyphen2 and SIFT, and revealed all these variants are damaging or probable damaging.\u003c/p\u003e\n\u003cp\u003ePatients with \u003cem\u003eKCNH2\u003c/em\u003e, \u003cem\u003eSCN5A\u003c/em\u003e or \u003cem\u003eCACNA1C\u003c/em\u003e mutation also received epinephrine test for possible long QT syndrome (LQTS), and all were negative. Patients with \u003cem\u003eSCN5A\u003c/em\u003e or\u003cem\u003e\u0026nbsp;CACNA1C\u003c/em\u003e mutation also received oral flecainide loading test for possible BrS, and all were also negative.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003ePathogenic and likely pathogenic variants\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eSeven P/LP variants were identified in our IVA cohort (Table 1), and all these variants are novel. Two patients carried two novel missense variants in the heart failure related SCD gene \u003cem\u003eCACNA1H\u003c/em\u003e, and one patient in LQTS-associated \u003cem\u003eKCNH2\u003c/em\u003e gene. One patient carried a novel missense mutation in the hypertrophic cardiomyopathy (HCM)-related \u003cem\u003eMYH6\u003c/em\u003e gene and one in the ARVC (arrhythmogenic right ventricular cardiomyopathy)-related \u003cem\u003eDSP\u003c/em\u003e gene. Two patients carried a novel missense mutation in the Brugada-related\u003cem\u003e\u0026nbsp;SCN5A\u003c/em\u003e or \u003cem\u003eCACNA1C\u003c/em\u003e gene and one in the catecholaminergic polymorphic ventricular tachycardia (CPVT)-related \u003cem\u003eRYR2\u003c/em\u003e gene.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe demographics of patients carrying P/LP variants were significant different from those with VUS. The patients with P/LP were younger (54.0\u0026plusmn;6.0 vs 63.5\u0026plusmn;10.4 years, P=0.039), and more were women (4/7 vs 1/13, P=0.031). The illustration summarizing the study designs and conclusion could be found in Figure 1.\u0026nbsp;\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eIn this nationwide cohort of patients with IVA, we identified several novel genes and variants, which have never been reported before and include ionic channel genes such as \u003cem\u003eSCN5A, KCNH2\u003c/em\u003e, \u003cem\u003eCACNA1C\u003c/em\u003e, and \u003cem\u003eCACNA1H\u003c/em\u003e and cardiac myofilaments or connexin genes such as \u003cem\u003eMYBPC3, MYH6, NEXN\u003c/em\u003e, \u003cem\u003eTTN\u003c/em\u003e and \u003cem\u003eDSP\u003c/em\u003e genes.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eGenetic studies of idiopathic ventricular fibrillation (IVF) or unexplained SCD had been conducted in various populations. The positive rate are 1.6 % to 23%\u0026nbsp;(28,29)\u0026nbsp;in the Asian population, and 0 % to 10%\u0026nbsp;(30-32)\u0026nbsp;in the Caucasian populations. The VUS positive rate may be up to 20% in a recent population of genetics of Caucasian IVF patients by Pannone et al.(33)\u0026nbsp;The genes identified were highly variable. The common genes in IVF are \u003cem\u003eCCR7\u003c/em\u003e and \u003cem\u003ePKN2\u003c/em\u003e, \u003cem\u003eSCN5A\u003c/em\u003e, \u003cem\u003eKCNQ1\u0026nbsp;\u003c/em\u003eand \u003cem\u003eRYR2\u003c/em\u003e (29,34)\u0026nbsp;in the Asian populations, and \u003cem\u003eCALM1~3\u003c/em\u003e, \u003cem\u003eRYR2\u003c/em\u003e, \u003cem\u003eTRDN\u003c/em\u003e, \u003cem\u003eCACNA1C\u003c/em\u003e, \u003cem\u003eSCN5A\u003c/em\u003e, \u003cem\u003eKCNE5\u003c/em\u003e in the Caucasian populations.(35)\u003c/p\u003e\n\u003cp\u003eSearching for the cause of unexplained SCD is a great challenge.(32)\u0026nbsp;In the current era, genetic test or molecular autopsy may provide some implications. A high yield rate of genetic test was found in our NGS cohort of Asian IVA patients. Around 18% of our IVA patients had\u0026nbsp;P/LP variants, which is similar to that reported by\u0026nbsp;Lahrouchi et al., in which\u0026nbsp;molecular autopsy (genetic test) for electrical disorder and cardiomyopathy genes, using ACMG guidelines for variant classification, identified a modest but realistic yield in patients with sudden arrhythmic death syndrome (SADS)(13% of patients had\u0026nbsp;P/LP\u0026nbsp;variants).(36)\u0026nbsp;Importantly, we also found the important roles of\u0026nbsp;CPVT and LQTS genes in unexplained SCD as in the CASPER study.(20)\u0026nbsp;Fang et al. conducted the first genome-wide association study of idiopathic ventricular tachycardia only in the Chinese Han population.(29)\u0026nbsp;They identified 4 novel loci and 2 risk genes (\u003cem\u003ePKN2\u003c/em\u003e and \u003cem\u003eCCR7\u003c/em\u003e) for\u0026nbsp;idiopathic ventricular tachycardia, implicating the role of\u0026nbsp;cardiac calcium homeostasis in rhythm maintenance. However, whether these genes also play a role in IVF or unexplained SCD remains unknown.\u003c/p\u003e\n\u003cp\u003eIn addition to BrS, genetic variants in \u003cem\u003eSCN5A\u003c/em\u003e and\u003cem\u003e\u0026nbsp;CACNA1C\u0026nbsp;\u003c/em\u003emay also be implicated in early repolarization syndrome (ERS). ERS is a newly identified entity with J wave elevation in inferior leads.(37,38)\u0026nbsp;Many different variants had been reported, probably due to heterogeneity of the disease. In our patients with these variants, no J wave elevation or early repolarization was noted in screening ECG. However, J wave elevation or early repolarization is a dynamic process and more close monitoring of ECG may reveal them. In our patients, some have LQTS gene variants such as \u003cem\u003eKCNH2, SCN5A,\u0026nbsp;\u003c/em\u003eand \u003cem\u003eCACNA1C\u003c/em\u003e. If these patients do have LQTS, it indicates the limitation of the diagnostic value of traditional ECG, especially resting ECG. The QT interval may change with drug challenge or epinephrine and exercise tests, but the sensitivity and specificity are not well documented.(16,39)\u0026nbsp;Whether a molecular or genetic diagnosis of LQTS is feasible without ECG confirmation remains unknown, especially in the Asian or our Taiwanese populations. According to\u0026nbsp;the\u0026nbsp;guidelines, an unknown SCD patients should be diagnosed with LQTS if a pathogenic variant is found in major LQTS genes, even the QT or QTc interval is not prolonged.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIn Taiwan\u0026nbsp;or Asia, the incidence of LQTS\u0026nbsp;may be\u0026nbsp;lower compared to the Caucasian populations(40-43)\u0026nbsp;but the rates of background or incidental pathogenic variants of LQTS genes in our Taiwanese population are relatively high (especially \u003cem\u003eSCN5A\u003c/em\u003e and \u003cem\u003eKCNH2\u003c/em\u003e genes).(44)\u0026nbsp;Therefore, if we use the sole criteria of having\u0026nbsp;LQTS\u0026nbsp;gene pathogenic variant\u0026nbsp;to diagnose LQTS, there will be much more LQTS patients in Taiwan, which is really not the case.\u0026nbsp;Nevertheless, all our patients with LQTS gene mutations had undergone ICD implantation. Furthermore, empirical use of beta blocker may not be harmful, and may also be advantageous in patients with unknown cause of SCD. Therefore, all these patients received beta blocker therapy. They will also be closely followed up for the possibility of LQTS.\u003c/p\u003e\n\u003cp\u003eThere are also overlaps of genes responsible for different hereditary arrhythmia, and it is also possible these patients with LQTS gene mutations might have BrS or ERS. The ECG manifestation of these diseases may change in different circumstances, such as fever unmasking Brudaga pattern.(45)\u0026nbsp;Again, close follow-up of these patients is mandatory.\u0026nbsp;We also found two patients having\u003cem\u003e\u0026nbsp;CACNA1H\u0026nbsp;\u003c/em\u003evariants.\u003cem\u003e\u0026nbsp;CACNA1H\u003c/em\u003e mutation has not been reported in genetic SCD. \u003cem\u003eCACNA1H\u003c/em\u003e encodes for the alpha subunit of T type calcium channel, which has been implicated to play a possible role in the mechanism of ventricular arrhythmia in the setting of heart failure.(46)\u0026nbsp;Somatic mutation of\u0026nbsp;\u003cem\u003eCACNA1H\u003c/em\u003e has also been found to cause primary aldosteronism.(47)\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eMYH6\u0026nbsp;\u003c/em\u003egene encodes for the cardiac contractile filament protein, mutation of which causes HCM with myocyte disarray, ventricular hypertrophy, and increased susceptibility to ventricular arrhythmia.(48)\u0026nbsp;TTN gene encodes for a very large protein called Titin,(49)\u0026nbsp;which functions as a molecular spring responsible for passive elasticity of muscle.(49)\u0026nbsp;TTN truncating variants are a common cause of dilated cardiomyopathy, occurring in approximately 25% of genetic dilated cardiomyopathy and in 18% of sporadic cases.(50,51)\u0026nbsp;In our patients with IVA, although no structural heart disease had been identified, this did not exclude the possibility that these patients would eventually develop cardiomyopathy.\u0026nbsp;It is also possible that arrhythmia precedes structural changes, and the patients may later manifest structural change in the heart. This finding was\u0026nbsp;similar to that reported by\u0026nbsp;Isbister et al. looking at genetic causes of cardiomyopathy in patients with autopsy negative sudden death, also suggesting that the earliest presentation of cardiomyopathy could be arrhythmic events.(52)\u0026nbsp;This warrants periodic surveillance of myocardial imaging and function.\u0026nbsp;Whether treatment for specific cardiomyopathy should be instituted in these patients remain unknown. Patients with HCM may have favorable response to beta blocker which is also helpful for a wide spectrum of cause of SCD.(53)\u0026nbsp;Finally, it is also possible that the genetic variant was just a by-stander, based on the high rate of incidental variant in cardiac myosin genes in our Taiwanese population.(44)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eWe could not rule out the possibility that the high positive rate of genetic test in our study is due to many genes were tested simultaneously. In those patients with frank phenotypes, such as HCM or LQTS, only meaningful target genes will be tested. In patients with IVA, we did not pre-specify genes or select genes \u003cem\u003ein priori\u003c/em\u003e, thus many genes related to SCD (such as our SCD panel) were tested which might generate many unrelated variants, especially VUS.\u0026nbsp;In addition to this Asian study, VUS was also found in a high proportion of Caucasian patients with unexplained SCD.(20,33)\u0026nbsp;Interpreting VUS is usually a great challenge and dilemma either in patients with unexplained SCD or in those with clinical diagnosis confirmed SCD case (such as a MYH7 VUS in a clinically confirmed LQTS patient). But VUS implicating a SCD syndrome in a patient with unexplained SCD or implicating the other cause of SCD in a patient with clinically confirmed SCD case may encourage the physician to follow-up the patient more closely and also to perform molecular functional studies of this variant to define its role in SCD.\u003c/p\u003e\n\u003cp\u003eNevertheless, the rates of VUS or P/LP variants were significantly higher in our IVA patients than those of incidental variants in our Taiwanese general population.(44)\u0026nbsp;This result may imply that the identified variants do mean something and are not incidental.\u0026nbsp;Finally, we stress the importance of complete work-up before diagnosing a patient with IVF or the genetic test results will be very misleading and confusing. In this study, we worked up these patients according to the reported SCD guidelines to avoid as many occult or concealed SCD syndromes included in our study population as possible.\u0026nbsp;Most importantly, the genetic findings might stimulate re-evaluation of the original clinical data and possibly lead to a renewed clinical diagnosis.(54)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThere are limitations in the present study. First, there was no cascade family screening done in this study, either clinical screening or genetic screening. Phenotypes corresponding to the identified variant in other asymptomatic family members may confirm the role of the identified variant. Co-segregation of variant may also confirm the role of identified variants. All the patients received history taking and most of them reported no family history of SCD or unexplained syncope, probably because of the nature of the Asian people not willing to reveal detailed family history, particular those histories related to severe diseases. Second, our IVA patients were significantly older compared to other studies. It is possible that it is easier to make a definite diagnosis in Asian young patients with SCD, such as BrS, or our IVA population represents a milder form of ventricular arrhythmia manifesting late or at older age. Third, because the cost of genetic test was covered by the research grant and nearly all the patients agreed to receive a genetic test, we believed in our study there was no selection bias, such as those with a more severe phenotype more intending to take the genetic test. Finally,\u0026nbsp;not all our patients received sodium channel blocker provocative test and we also only used oral flecainide loading test as the provocative test. Also, not all patients received CMR examination because early in this study, CMR was not a routine for unexplained SCD. Therefore, there might be concealed BrS or cardiomyopathy patients in our study subjects. That is the reason why we used IVA but not IVF in our study.\u003c/p\u003e\n\u003cp\u003eIn conclusion, in this largest IVA cohort with NGS genetic testing in Asia, we identified genes coding for channelopathy or cardiomyopathy that are not clinically manifested. Because of high yield rate of variants in our IVA patients, genetic test may be considered in Asian patients with IVA. Periodic surveillance for myocardial morphology and function is important in these patients. Repeated provocative test could also be considered.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eACMG: American College of Medical Genetics\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAF: atrial fibrillation\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eARVC: arrhythmogenic right ventricular cardiomyopathy\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eBrS: Brudaga syndrome\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eCMR: cardiac magnetic resonance imaging\u003c/p\u003e\n\u003cp\u003eCPVT: catecholaminergic polymorphic ventricular tachycardia\u003c/p\u003e\n\u003cp\u003eECG: electrocardiography\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eERS: early repolarization syndrome\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eHCM: hypertrophic cardiomyopathy\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIVA: idiopathic ventricular arrhythmia\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIVT: idiopathic ventricular tachycardia\u003c/p\u003e\n\u003cp\u003eIVF: idiopathic ventricular fibrillation\u003c/p\u003e\n\u003cp\u003eLQTS: long QT syndrome\u003c/p\u003e\n\u003cp\u003eNGS: next-generation sequencing\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eOHCA: Out of hospital cardiac arrest\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eP/LP: pathogenic/likely pathogenic\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eSADS: sudden arrhythmic death syndrome\u003c/p\u003e\n\u003cp\u003eSCD: sudden cardiac death\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eVUS: variants of uncertain significance\u0026nbsp;\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003eAuthor contribution statement:\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eJien-Jiun Chen, designed the study, collected patients\u0026rsquo; data, and wrote the manuscript. Seng-Nan Chang, Fu-Chun Chiu, Pang-Shuo Huang, Cho-Kai Wu, Yi-Chih Wang, collected patients\u0026rsquo; data and gave a critical review of the manuscript. Juey-Jen Hwang and Chia-Ti Tsai designed the study and genetic panel, in charge of the whole study.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFunding\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThis study was supported by a grant from the National Science and Technology Council (NSTC)(105-2314-B-002-153-MY2).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAcknowledgments\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eWe would thank all the patients and intensivists who cared these patients\u003c/p\u003e\n\u003cp\u003eThis research has been conducted using data from UK Biobank (www.ukbiobank.ac.uk), a major biomedical database, approved under project # 54423.\u003c/p\u003e\n\u003cp\u003eAll authors have read the journal\u0026apos;s policy on disclosure of potential conflicts of interest. All authors have disclosed any financial or personal relationship with organizations that could potentially be perceived as influencing the described research.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eConflict of interests\u003c/p\u003e\n\u003cp\u003eAll authors declared no competing interests. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003eData availability\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eEthics statements\u003c/p\u003e\n\u003cp\u003eEthics approval\u003c/p\u003e\n\u003cp\u003eThis study involves human participants and was approved by the Research ethics committee of National Taiwan University Hospital (201705122RINC). Written informed consent was provided by all participants/legal guardians.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eViskin S, Belhassen B. Idiopathic ventricular fibrillation. Am Heart J 1990;120:661\u0026ndash;71.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAlders M, Koopmann TT, Christiaans I et al. Haplotype-sharing analysis implicates chromosome 7q36 harboring DPP6 in familial idiopathic ventricular fibrillation. Am J Hum Genet 2009;84:468\u0026ndash;76.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen Q, Kirsch GE, Zhang D et al. Genetic basis and molecular mechanism for idiopathic ventricular fibrillation. Nature 1998;392:293\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWan X, Chen S, Sadeghpour A, Wang Q, Kirsch GE. Accelerated inactivation in a mutant Na(+) channel associated with idiopathic ventricular fibrillation. Am J Physiol Heart Circ Physiol 2001;280:H354-60.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eValdivia CR, Medeiros-Domingo A, Ye B et al. Loss-of-function mutation of the SCN3B-encoded sodium channel {beta}3 subunit associated with a case of idiopathic ventricular fibrillation. Cardiovasc Res 2010;86:392\u0026ndash;400.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOhno S, Zankov DP, Ding WG et al. KCNE5 (KCNE1L) variants are novel modulators of Brugada syndrome and idiopathic ventricular fibrillation. Circ Arrhythm Electrophysiol 2011;4:352\u0026ndash;61.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBagnall RD, Molloy LK, Kalman JM, Semsarian C. Exome sequencing identifies a mutation in the ACTN2 gene in a family with idiopathic ventricular fibrillation, left ventricular noncompaction, and sudden death. BMC Med Genet 2014;15:99.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNomikos M, Thanassoulas A, Beck K et al. Altered RyR2 regulation by the calmodulin F90L mutation associated with idiopathic ventricular fibrillation and early sudden cardiac death. FEBS Lett 2014;588:2898\u0026ndash;902.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eManolio TA, Collins FS, Cox NJ et al. Finding the missing heritability of complex diseases. Nature 2009;461:747\u0026ndash;53.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNg SB, Buckingham KJ, Lee C et al. Exome sequencing identifies the cause of a mendelian disorder. Nat Genet 2010;42:30\u0026ndash;5.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCirulli ET, Goldstein DB. Uncovering the roles of rare variants in common disease through whole-genome sequencing. Nat Rev Genet 2010;11:415\u0026ndash;25.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen YH, Pai CW, Huang SW et al. Inactivation of Myosin binding protein C homolog in zebrafish as a model for human cardiac hypertrophy and diastolic dysfunction. J Am Heart Assoc 2013;2:e000231.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAl-Khatib SM, Stevenson WG, Ackerman MJ et al. 2017 AHA/ACC/HRS Guideline for Management of Patients With Ventricular Arrhythmias and the Prevention of Sudden Cardiac Death: Executive Summary: A Report of the American College of Cardiology/American Heart Association Task Force on Clinical Practice Guidelines and the Heart Rhythm Society. J Am Coll Cardiol 2018;72:1677\u0026ndash;1749.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePedersen CT, Kay GN, Kalman J et al. EHRA/HRS/APHRS expert consensus on ventricular arrhythmias. Heart Rhythm 2014;11:e166-96.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDubner S, Azocar D, Gallino S et al. Single oral flecainide dose to unmask type 1 Brugada syndrome electrocardiographic pattern. Ann Noninvasive Electrocardiol 2013;18:256\u0026ndash;61.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eVyas H, Hejlik J, Ackerman MJ. Epinephrine QT stress testing in the evaluation of congenital long-QT syndrome: diagnostic accuracy of the paradoxical QT response. Circulation 2006;113:1385\u0026ndash;92.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChang SN, Tsai CT, Wu CK et al. A functional variant in the promoter region regulates the C-reactive protein gene and is a potential candidate for increased risk of atrial fibrillation. J Intern Med 2012;272:305\u0026ndash;15.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTsai CT, Lai LP, Lin JL et al. Renin-angiotensin system gene polymorphisms and atrial fibrillation. Circulation 2004;109:1640\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChiang FT, Hsu KL, Tseng CD et al. Molecular variant M235T of the angiotensinogen gene is associated with essential hypertension in Taiwanese. J Hypertens 1997;15:607\u0026ndash;11.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGrondin S, Davies B, Cadrin-Tourigny J et al. Importance of genetic testing in unexplained cardiac arrest. Eur Heart J 2022;43:3071\u0026ndash;3081.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHellenthal N, Gaertner-Rommel A, Klauke B et al. Molecular autopsy of sudden unexplained deaths reveals genetic predispositions for cardiac diseases among young forensic cases. Europace 2017;19:1881\u0026ndash;1890.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAdler A, Novelli V, Amin AS et al. An International, Multicentered, Evidence-Based Reappraisal of Genes Reported to Cause Congenital Long QT Syndrome. Circulation 2020;141:418\u0026ndash;428.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTsai CT, Hsieh CS, Chang SN et al. Genome-wide screening identifies a KCNIP1 copy number variant as a genetic predictor for atrial fibrillation. Nat Commun 2016;7:10190.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eErikson GA, Deshpande N, Kesavan BG, Torkamani A. SG-ADVISER CNV: copy-number variant annotation and interpretation. Genet Med 2015;17:714\u0026ndash;8.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKarczewski KJ, Weisburd B, Thomas B et al. The ExAC browser: displaying reference data information from over 60 000 exomes. Nucleic Acids Res 2017;45:D840-D845.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eThurman RE, Rynes E, Humbert R et al. The accessible chromatin landscape of the human genome. Nature 2012;489:75\u0026ndash;82.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMercer TR, Edwards SL, Clark MB et al. DNase I-hypersensitive exons colocalize with promoters and distal regulatory elements. Nat Genet 2013;45:852\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSon MK, Ki CS, Park SJ, Huh J, Kim JS, On YK. Genetic mutation in Korean patients of sudden cardiac arrest as a surrogating marker of idiopathic ventricular arrhythmia. J Korean Med Sci 2013;28:1021\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFang C, Wang P, Yu D et al. Genome-Wide Association Study for Idiopathic Ventricular Tachyarrhythmias Identifies Key Role of CCR7 and PKN2 in Calcium Homeostasis and Cardiac Rhythm Maintenance. Circ Genom Precis Med 2022;15:e003603.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHerman AR, Cheung C, Gerull B et al. Outcome of Apparently Unexplained Cardiac Arrest: Results From Investigation and Follow-Up of the Prospective Cardiac Arrest Survivors With Preserved Ejection Fraction Registry. Circ Arrhythm Electrophysiol 2016;9:e003619.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBai R, Napolitano C, Bloise R, Monteforte N, Priori SG. Yield of genetic screening in inherited cardiac channelopathies: how to prioritize access to genetic testing. Circ Arrhythm Electrophysiol 2009;2:6\u0026ndash;15.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eVisser M, van der Heijden JF, Doevendans PA, Loh P, Wilde AA, Hassink RJ. Idiopathic Ventricular Fibrillation: The Struggle for Definition, Diagnosis, and Follow-Up. Circ Arrhythm Electrophysiol 2016;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePannone L, Gauthey A, Conte G et al. Genetics in Probands With Idiopathic Ventricular Fibrillation: A Multicenter Study. JACC Clin Electrophysiol 2023;9:1296\u0026ndash;1306.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhong X, Guo W, Wei J et al. Identification of loss-of-function RyR2 mutations associated with idiopathic ventricular fibrillation and sudden death. Biosci Rep 2021;41.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang M, Tu X. The Genetics and Epigenetics of Ventricular Arrhythmias in Patients Without Structural Heart Disease. Front Cardiovasc Med 2022;9:891399.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLahrouchi N, Raju H, Lodder EM et al. Utility of Post-Mortem Genetic Testing in Cases of Sudden Arrhythmic Death Syndrome. J Am Coll Cardiol 2017;69:2134\u0026ndash;2145.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNademanee K, Haissaguerre M, Hocini M et al. Mapping and Ablation of Ventricular Fibrillation Associated with Early Repolarization Syndrome. Circulation 2019.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMahapatra SR, Chakraborty P. An update on early repolarization(ER) syndrome. Indian Pacing Electrophysiol J 2015;15:265\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChuret M, Luttoo K, Hocini M, Haissaguerre M, Sacher F, Duchateau J. Diagnostic reproducibility of epinephrine drug challenge interpretation in suspected long QT syndrome. J Cardiovasc Electrophysiol 2019;30:896\u0026ndash;901.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYoshinaga M, Kucho Y, Nishibatake M, Ogata H, Nomura Y. Probability of diagnosing long QT syndrome in children and adolescents according to the criteria of the HRS/EHRA/APHRS expert consensus statement. Eur Heart J 2016;37:2490\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eUhm JS, Hwang IU, Oh YS et al. Prevalence of electrocardiographic findings suggestive of sudden cardiac death risk in 10,867 apparently healthy young Korean men. Pacing Clin Electrophysiol 2011;34:717\u0026ndash;23.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHayashi K, Fujino N, Uchiyama K et al. Long QT syndrome and associated gene mutation carriers in Japanese children: results from ECG screening examinations. Clin Sci (Lond) 2009;117:415\u0026ndash;24.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMurakoshi N, Aonuma K. Epidemiology of arrhythmias and sudden cardiac death in Asia. Circ J 2013;77:2419\u0026ndash;31.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHuang PS, Hsieh CS, Chang SN et al. Prevalence of sudden arrhythmic death syndrome-related genetic mutations in an Asian cohort of whole genome sequence. Europace 2020;22:1287\u0026ndash;1297.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eArgenziano M, Antzelevitch C. Recent advances in the treatment of Brugada syndrome. Expert Rev Cardiovasc Ther 2018;16:387\u0026ndash;404.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKinoshita H, Kuwahara K, Takano M et al. T-type Ca2\u0026thinsp;+\u0026thinsp;channel blockade prevents sudden death in mice with heart failure. Circulation 2009;120:743\u0026ndash;52.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNanba K, Blinder AR, Rege J et al. Somatic CACNA1H Mutation As a Cause of Aldosterone-Producing Adenoma. Hypertension 2020;75:645\u0026ndash;649.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCarrier L, Mearini G, Stathopoulou K, Cuello F. Cardiac myosin-binding protein C (MYBPC3) in cardiac pathophysiology. Gene 2015;573:188\u0026ndash;97.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLabeit S, Kolmerer B. Titins: giant proteins in charge of muscle ultrastructure and elasticity. Science 1995;270:293\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHerman DS, Lam L, Taylor MR et al. Truncations of titin causing dilated cardiomyopathy. N Engl J Med 2012;366:619\u0026ndash;28.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFang HJ, Liu BP. Prevalence of TTN mutations in patients with dilated cardiomyopathy: A meta-analysis. Herz 2019.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eIsbister JC, Nowak N, Yeates L et al. Concealed Cardiomyopathy in Autopsy-Inconclusive Cases of Sudden Cardiac Death and Implications for Families. J Am Coll Cardiol 2022;80:2057\u0026ndash;2068.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDrazner MH. Angiotensin Receptor-Neprilysin Inhibition (ARNI) Therapy and Reverse Remodeling in Heart Failure With Reduced Ejection Fraction. JAMA 2019:1\u0026ndash;3.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLandstrom AP, Dailey-Schwartz AL, Rosenfeld JA et al. Interpreting Incidentally Identified Variants in Genes Associated With Catecholaminergic Polymorphic Ventricular Tachycardia in a Large Cohort of Clinical Whole-Exome Genetic Test Referrals. Circ Arrhythm Electrophysiol 2017;10.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"sudden arrhythmia death, genetic test, idiopathic ventricular arrhythmia, next-generation sequencing, variants","lastPublishedDoi":"10.21203/rs.3.rs-4458113/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4458113/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSudden cardiac death (SCD) due to idiopathic ventricular tachycardia or ventricular fibrillation is a catastrophic disease. Its genetic basis is heterogeneous and has been rarely addressed in Asia. We aim to find variants in an Asian cohort of idiopathic ventricular arrhythmia (IVA).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNationwide patients with IVA were consecutively recruited. We designed a SCD gene panel (134 genes) to detect variants by next-generation sequencing (NGS) including most of the channelopathy and cardiomyopathy genes.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA total of 40 IVA patients were included. Thirteen variants with unknown significance (VUS) and 7 pathogenic/likely pathogenic (P/LP) variants were identified in 20 patients (50%). All variants were novel and not found in dbSNP, ExAC and our general population. The identified variants were in genes implicated for long QT or Brugada syndromes (SCN5A, KCNH2, CACNA1C and ANK2), cardiomyopathy (MYH6, DSP and TTN) and catecholaminergic polymorphic ventricular tachycardia (RYR2). Patients with P/LP were younger, and more were women than those with VUS.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA high yield rate of genetic test was found in the largest NGS cohort of Asian IVA patients. These patients should be vigorously followed up for possible channelopathy or cardiomyopathy with repeated provocative test and myocardial imaging.\u003c/p\u003e","manuscriptTitle":"Identification of Genetic Variants in Patients with Idiopathic Ventricular Arrhythmia by Taiwan Arrhythmia Gene Panel Implying Underlying Mechanisms","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-06-07 23:02:25","doi":"10.21203/rs.3.rs-4458113/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"a43d8b9a-8e2d-4815-b2db-6f299d9ac275","owner":[],"postedDate":"June 7th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2024-10-26T04:38:23+00:00","versionOfRecord":[],"versionCreatedAt":"2024-06-07 23:02:25","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4458113","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4458113","identity":"rs-4458113","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}