Evaluation of the Genetic Basis of Familial Aggregation of Pacemaker Implantation by a Large Next Generation Sequencing Panel

Abstract

BackgroundThe etiology of conduction disturbances necessitating permanent pacemaker (PPM) implantation is often unknown, although familial aggregation of PPM (faPPM) suggests a possible genetic basis. We developed a pan-cardiovascular next generation sequencing (NGS) panel to genetically characterize a selected cohort of faPPM.Materials and MethodsWe designed and validated a custom NGS panel targeting the coding and splicing regions of 246 genes with involvement in cardiac pathogenicity. We enrolled 112 PPM patients and selected nine (8%) with faPPM to be analyzed by NGS.ResultsOur NGS panel covers 95% of the intended target with an average of 229x read depth at a minimum of 15-fold depth, reaching a SNP true positive rate of 98%. The faPPM patients presented with isolated cardiac conduction disease (ICCD) or sick sinus syndrome (SSS) without overt structural heart disease or identifiable secondary etiology. Three patients (33.3%) had heterozygous deleterious variants previously reported in autosomal dominant cardiac diseases including CCD: LDB3 (p.D117N) and TRPM4 (p.G844D) variants in patient 4; TRPM4 (p.G844D) and ABCC9 (p.V734I) variants in patient 6; and SCN5A (p.T220I) and APOB (p.R3527Q) variants in patient 7.ConclusionFaPPM occurred in 8% of our PPM clinic population. The employment of massive parallel sequencing for a large selected panel of cardiovascular genes identified a high percentage (33.3%) of the faPPM patients with deleterious variants previously reported in autosomal dominant cardiac diseases, suggesting that genetic variants may play a role in faPPM.

Reevaluating the Mutation Classification in Genetic Studies of Bradycardia Using ACMG/AMP Variant Classification Framework

“…We studied 88 articles, including 99 variants and 34 genes, after searching the PubMed database and identified 13 high-priority genes causing familial bradycardia, as follows: ABCC9 [18], ACTN2 [19], CACNA1C [20,21], DES [22][23][24][25][26][27], HCN4 [28][29][30][31][32], KCNQ1 [33,34], KCNH2 [35], LMNA [36,37], MECP2 [38], LAMP2 [39], NPPA [40], SCN5A [41][42][43][44][45], and TRPM4 [5,[46][47][48] (Table 3).…”

Genetic analyses in a bonobo (Pan paniscus) with arrhythmogenic right ventricular cardiomyopathy

“…Given that we did not find any pathogenic/likely pathogenic variants using our Sanger approach, we used a custom NGS panel to look for variants in the coding and splicing regions of 246 genes associated with cardiovascular disorders in Lody’s whole blood and heart and Kitty’s whole blood DNA as described in the Methods section and in a previous study 22 . In total, using the human genome GRCh37/hg19 as reference, 20,756 variants were found in Lody’s whole blood DNA, 19,817 variants were found in Lody’s formalin-fixed paraffin-embedded (FFPE) heart DNA, and 20,040 variants were found in Kitty’s whole blood DNA.…”

“…A previously described custom next generation sequencing (NGS) panel containing probes for human DNA was used in the DNA from Lody’s whole blood and heart and from Kitty’s blood to sequence the coding and splicing regions of 246 genes associated with cardiovascular disorders including arrhythmias, cardiomyopathies, congenital heart defects, aortopathy, connective tissue disorders, Noonan spectrum disorders, pulmonary arterial hypertension, metabolic disorders that afflict the heart and lipid disorders 22 . Briefly, paired-end sequencing was performed using an Illumina MiSeq sequencer (Illumina, Inc., San Diego, CA), followed by read alignment using the BWA software, local realignment, base quality recalibration, and variant identification using the GATK software, and variant annotation using ANNOVAR 22 . A 300× average depth of coverage was obtained among variants for the three samples.…”

“…The HGMD variants were classified as being pathogenic/likely pathogenic, pathogenic/likely pathogenic in autosomal recessive disorders, modifier, VUS, or benign/likely benign based on our interpretation from the literature 22 . In order to filter the non-HGMD variants (variants that did not have a previous association with human diseases) and find those with higher likelihood of having disease association, we selected variants that were in coding regions, excluded synonymous and nonsynonymous that were found in more than 2% of the populations in the 1000 Genomes browser, the NHLBI Exome Sequencing Project, and the gnomAD browser beta 20 , excluded variants that had a damage score lower than 4 (based on software in silico prediction of damage to protein structure/function) 22 , excluded synonymous and non-frameshift TTN variants, excluded variants that mapped to regions of known segmental duplications, and excluded non-frameshift variants that were found in more than 2% of the populations in the 1000 Genomes browser, in the NHLBI Exome Sequencing Project, or in the gnomAD browser beta. Following, a subtractive analysis was performed to select only those resulting HGMD and non-HGMD variants of acceptable sequencing quality scores that were specific to Lody (not in Kitty, unless heterozygous in Kitty for a homozygous variant in Lody), as those in common between the two individuals were judged to be either unlikely to be clinically relevant or to be of low penetrance.…”

Genetic Discovery of ATP-Sensitive K + Channels in Cardiovascular Diseases

“…It was found that mutant Kir6.2/SUR2B channels, but not Kir6.2/SUR2A or Kir6.1/SUR2B channels, had reduced sensitivity to MgATP inhibition, suggestive of K ATP overactivity in the endothelial cell subunit combination. In addition, the V734I variant was reported as a gain-of-function mutation in four early repolarization syndrome (ERS) patients with bradycardia 79 and in a patient with a permanent pacemaker who presented with isolated cardiac conduction disease 80 , perhaps consistent with K ATP channels playing a unique role in pacemaker and conduction system cells.…”