Whole-exome sequencing in familial atrial fibrillation.

AIMS: Positional cloning and candidate gene approaches have shown that atrial fibrillation (AF) is a complex disease with familial aggregation. Here, we employed whole-exome sequencing (WES) in AF kindreds to identify variants associated with familial AF. METHODS AND RESULTS: WES was performed on 18 individuals in six modestly sized familial AF kindreds. After filtering very rare variants by multiple metrics, we identified 39 very rare and potentially pathogenic variants [minor allele frequency (MAF) ≤0.04%] in genes not previously associated with AF. Despite stringent filtering >1 very rare variants in the 5/6 of the kindreds were identified, whereas no plausible variants contributing to familial AF were found in 1/6 of the kindreds. Two candidate AF variants in the calcium channel subunit genes (CACNB2 and CACNA2D4) were identified in two separate families using expression data and predicted function. CONCLUSION: By coupling family data with exome sequencing, we identified multiple very rare potentially pathogenic variants in five of six families, suggestive of a complex disease mechanism, whereas none were identified in the remaining AF pedigree. This study highlights some important limitations and challenges associated with performing WES in AF including the importance of having large well-curated multi-generational pedigrees, the issue of potential AF misclassification, and limitations of WES technology when applied to a complex disease.

Candidate gene approach to identifying rare genetic variants associated with lone atrial fibrillation.

BACKGROUND: Rare variants in candidate atrial fibrillation (AF) genes have been associated with AF in small kindreds. The extent to which such polymorphisms contribute to AF is unknown. OBJECTIVE: The purpose of this study was to determine the spectrum and prevalence of rare amino acid coding (AAC) variants in candidate AF genes in a large cohort of unrelated lone AF probands. METHODS: We resequenced 45 candidate genes in 303 European American (EA) lone AF probands (186 lone AF probands screened for each gene on average [range 89-303], 63 screened for all) identified in the Vanderbilt AF Registry (2002-2012). Variants detected were screened against 4300 EAs from the Exome Sequencing Project (ESP) to identify very rare (minor allele frequency ≤0.04%) AAC variants and these were tested for AF co-segregation in affected family members where possible. RESULTS: Median age at AF onset was 46.0 years [interquartile range 33.0-54.0], and 35.6% had a family history of AF. Overall, 63 very rare AAC variants were identified in 60 of 303 lone AF probands, and 10 of 19 (52.6%) had evidence of co-segregation with AF. Among the 63 lone AF probands who had 45 genes screened, the very rare variant burden was 22%. Compared with the 4300 EA ESP, the proportion of lone AF probands with a very rare AAC variant in CASQ2 and NKX2-5 was increased 3-5-fold (P <.05). CONCLUSION: No very rare AAC variants were identified in ~80% of lone AF probands. Potential reasons for the lack of very rare AAC variants include a complex pattern of inheritance, variants in as yet unidentified AF genes or in noncoding regions, and environmental factors.

A common variant on chromosome 4q25 is associated with prolonged PR interval in subjects with and without atrial fibrillation.

Single nucleotide polymorphisms (SNPs) at chromosome 4q25 (near PITX2) are strongly associated with atrial fibrillation (AF). We assessed whether a 4q25-tagging SNP (rs2200733) is associated with PR interval duration in patients with lone and typical AF and controls. Patients with lone (n = 169) and typical (n = 269) AF enrolled in the Vanderbilt AF registry and controls (n = 1,403) derived from the Vanderbilt DNA Biobank were studied. Carriage of the rs2200733T allele (CT or TT genotype) was more common in patients with lone (39%) than typical (25%) AF or controls (21%, p <0.01 for both comparisons). In both AF cohorts, we observed an association between genotype and PR interval duration (median PR interval for CC, CT, and TT: 162, 178, and 176 ms, respectively, for lone, p = 0.038 and 166, 180, and 196 ms, respectively, for typical, p = 0.001). After adjustment for covariates, the association between T allele and PR prolongation persisted, with mean effect size of 10.9, 12.8, and 4.4 ms for patients with lone and typical AF and controls, respectively (p <0.05 for each comparison). We found that a common 4q25 AF susceptibility allele (rs2200733) is associated with PR interval prolongation in patients with lone and typical AF and controls with no AF. Given that prolonged PR interval is an established risk factor for AF, this observation, in the context of previously described functional effects of PITX2 deficiency, provides further knowledge about the pathophysiological link of 4q25 variants with AF.

QT variability during initial exposure to sotalol: experience based on a large electronic medical record.

AIMS: A prolonged QT interval is associated with increased risk of Torsades de pointes (TdP) and may be fatal. We sought to investigate the extent to which clinical covariates affect the change in QT interval among 'real-world' patients treated with sotalol and followed in an electronic medical record (EMR) system. METHODS AND RESULTS: We used clinical alerts in our EMR system to identify all patients in whom a new prescription for sotalol was written (2001-11). Rate-corrected QT (QTc) was calculated by Bazett's formula. Correlates of sotalol-induced change in the QTc interval and sotalol discontinuation were examined using linear and logistic regression, respectively. Overall, 541 sotalol-exposed patients were identified (n = 200 women, 37%). The mean first sotalol dose was 86 ± 39 mg, age 64 ± 13 years, and BMI 30 ± 7 kg/m(2). Atrial fibrillation/flutter was the predominant indication (92.2%). After initial exposure, the change in the QTc interval from baseline was highly variable: ΔQTc after 2 h = 3 ± 42 ms (P = 0.17) and 11 ± 37 ms after ≥48 h (P < 0.001). Multivariable linear regression analysis identified female gender and age, reduced left ventricular ejection fraction, high sotalol dose, hypertrophic cardiomyopathy, and loop diuretic co-administration as correlates of increased ΔQTc at ≥48 h (P < 0.05 for all). Within 3 days of initiation, 12% discontinued sotalol of which 31% were because of exaggerated QTc prolongation. One percent developed TdP. CONCLUSION: In this EMR-based cohort, the increase in QTc with sotalol initiation was highly variable, and multiple clinical factors contributed. These data represent an important step in ongoing work to identify real-world patients likely to tolerate long-term therapy and reinforces the utility of EMR-based cohorts as research tools.

Common atrial fibrillation risk alleles at 4q25 predict recurrence after catheter-based atrial fibrillation ablation.

BACKGROUND: Common single nucleotide polymorphisms at chromosome 4q25 (rs2200733, rs10033464) are associated with both lone and typical atrial fibrillation (AF). Risk alleles at 4q25 have recently been shown to predict recurrence of AF after ablation in a population of predominately lone AF, but lone AF represents only 5%-30% of AF cases. OBJECTIVE: To test the hypothesis that 4q25 AF risk alleles can predict response to AF ablation in the majority of AF cases. METHODS: Patients enrolled in the Vanderbilt AF Registry underwent 378 catheter-based AF ablations (median age 60 years; 71% men; 89% typical AF) between 2004 and 2011. The primary end point was time to recurrence of any nonsinus atrial tachyarrhythmia (atrial tachycardia, atrial flutter, or AF). RESULTS: Two-hundred atrial tachycardia, atrial flutter, or AF recurrences (53%) were observed. In multivariable analysis, the rs2200733 risk allele predicted a 24% shorter recurrence-free time (survival time ratio 0.76; 95% confidence interval [CI] 0.6-0.95; P = .016) compared with wild type. The heterozygous haplotype demonstrated a 21% shorter recurrence-free time (survival time ratio 0.79; 95% CI 0.62-0.99) and the homozygous risk allele carriers a 39% shorter recurrence-free time (survival time ratio 0.61; 95% CI 0.37-1.0; P = .037). CONCLUSIONS: Risk alleles at the 4q25 loci predict impaired clinical response to AF ablation in a population of patients with predominately typical AF. Our findings suggest that the rs2200733 polymorphism may hold promise as an objectively measured patient characteristic that can be used as a clinical tool for selecting patients for AF ablation.

Relation of morbid obesity and female gender to risk of procedural complications in patients undergoing atrial fibrillation ablation.

Obese patients with atrial fibrillation (AF) are frequently treated with AF ablation. We sought to examine whether a body mass index (BMI) threshold exists beyond which the odds of experiencing a complication from AF ablation increases. All patients enrolled in the Vanderbilt AF Registry who underwent catheter-based AF ablation from May 1999 to February 2012 were included. Major complications were recorded. Morbid obesity was defined as a BMI >40 kg/m(2) and examined in multivariable analysis. A total of 35 complications (6.8%) occurred in 512 ablations. Morbidly obese patients experienced a greater rate of complications (6 of 42, 14.3%) than the nonmorbidly obese (29 of 470, 6.2%; p = 0.046). Using a discrete BMI cutoff, the odds of complications increased 3.1-fold in those with morbid obesity (odds ratio [OR] 3.1, 95% confidence interval [CI] 1.1 to 8.4, p = 0.03) and 2.1-fold for female gender (OR 2.1, 95% CI 1.04 to 4.38, p = 0.04). With BMI as a continuous variable, the odds of complications increased by 5% per 1 unit increase in BMI (OR 1.05, 95% CI 1.0 to 1.11, p = 0.05), and the increase for female gender was 2.2-fold (OR 2.2, 95% CI 1.1 to 4.6, p = 0.03). In conclusion, morbid obesity represents a BMI threshold above which the odds of complications with AF ablation increase significantly. The increase in complications appears to be driven primarily by events in women, suggesting that morbidly obese women are a special population when considering AF ablation.

Chromosome 4q25 variants are genetic modifiers of rare ion channel mutations associated with familial atrial fibrillation.

OBJECTIVES: The aim of this study was to test the hypothesis that 2 common polymorphisms in the chromosome 4q25 region that have been associated with atrial fibrillation (AF) contribute to the variable penetrance of familial AF. BACKGROUND: Although mutations in ion channels, gap junction proteins, and signaling molecules have been described for Mendelian forms of AF, penetrance is highly variable. Recent studies have consistently identified 2 common single-nucleotide polymorphisms in the chromosome 4q25 region as independent AF susceptibility alleles. METHODS: Eleven families in which AF was present in ≥2 members who also shared a candidate gene mutation were studied. These mutations were identified in all subjects with familial lone AF (n = 33) as well as apparently unaffected family members (age >50 years with no AF; n = 17). RESULTS: Mutations were identified in SCN5A (n = 6), NPPA (n = 2), KCNQ1 (n = 1), KCNA5 (n = 1), and NKX2.5 (n = 1). In genetic association analyses, unstratified and stratified according to age of onset of AF and unaffected age >50 years, there was a highly statistically significant association between the presence of both common (rs2200733 and rs10033464) and rare variants and AF (unstratified p = 1 × 10(-8), stratified [age of onset <50 years and unaffected age >50 years] p = 7.6 × 10(-5)) (unstratified p < 0.0001, stratified [age of onset <50 years and unaffected age >50 years] p < 0.0001). Genetic association analyses showed that the presence of common 4q25 risk alleles predicted whether carriers of rare mutations developed AF (p = 2.2 × 10(-4)). CONCLUSIONS: Common AF-associated 4q25 polymorphisms modify the clinical expression of latent cardiac ion channel and signaling molecule gene mutations associated with familial AF. These findings support the idea that the genetic architecture of AF is complex and includes both rare and common genetic variants.

Symptomatic response to antiarrhythmic drug therapy is modulated by a common single nucleotide polymorphism in atrial fibrillation.

OBJECTIVES: This study tested the hypothesis that response to antiarrhythmic drugs (AADs) is modulated by 3 common loci associated with atrial fibrillation (AF). BACKGROUND: Recent genome-wide association studies have identified 3 loci, on chromosomes 4q25 (near PITX2), 16q22 (in ZFHX3), and 1q21 (in KCNN3), that associate with either typical or lone AF. These findings indicate that variable mechanisms contribute to AF susceptibility, and suggest that response to therapy may be genotype dependent. METHODS: We studied 478 and 198 Caucasian patients in the discovery cohort and validation cohort, respectively, who were prospectively enrolled in the Vanderbilt AF registry. Response was defined prospectively as successful rhythm control if the patient remained on the same AAD therapy for a minimum of 6 months with ≥75% reduction in symptomatic AF burden. We also evaluated AF recurrence by 12-lead electrocardiogram (ECG) at 3, 6, and 12 months. Symptomatic patients were also given a 24- to 48-h Holter monitor or 30-day event recorder when AF recurrence was not captured by 12-lead ECG. RESULTS: In the discovery cohort, 399 (83%) patients were successfully rhythm controlled. Multiple clinical variables (including age, hypertension, lone AF) failed to significantly predict response to AADs; however, single nucleotide polymorphism (SNP) rs10033464 at 4q25 was an independent predictor of successful rhythm control in patients with typical AF carrying the ancestral allele (wild type) versus carriers of variant allele (odds ratio [OR]: 4.7, 95% confidence interval [CI]: 1.83 to 12, p = 0.0013. In the validation cohort, 143 (72%) patients met the criteria for successful rhythm control, and rs10033464 was again an independent predictor of successful rhythm control, OR: 1.5, 95% CI: 1.02 to 3.06, p = 0.04. This SNP (rs10033464) was an independent predictor of AF recurrence in the discovery (39% AF recurrence) and validation (38% AF recurrence) cohorts; OR: 3.27, 95% CI: 1.7 to 6, p < 0.001 and OR: 4.3, 95% CI: 1.98 to 9.4, p < 0.001, respectively. CONCLUSIONS: These results suggest that a common SNP on chromosome 4q25 associated with AF modulates response to AAD therapy and points to a potential role for stratification of therapeutic approaches by genotype.

A sequence variant in ZFHX3 on 16q22 associates with atrial fibrillation and ischemic stroke.

We expanded our genome-wide association study on atrial fibrillation (AF) in Iceland, which previously identified risk variants on 4q25, and tested the most significant associations in samples from Iceland, Norway and the United States. A variant in the ZFHX3 gene on chromosome 16q22, rs7193343-T, associated significantly with AF (odds ratio OR = 1.21, P = 1.4 x 10(-10)). This variant also associated with ischemic stroke (OR = 1.11, P = 0.00054) and cardioembolic stroke (OR = 1.22, P = 0.00021) in a combined analysis of five stroke samples.

Cardiac sodium channel (SCN5A) variants associated with atrial fibrillation.

BACKGROUND: Genetic studies have identified ion channel gene variants in families segregating atrial fibrillation (AF), the most common arrhythmia in clinical practice. Here, we tested the hypothesis that vulnerability to AF is associated with variation in SCN5A, the gene encoding the cardiac sodium channel. METHODS AND RESULTS: We resequenced the entire SCN5A coding region in 375 subjects with either lone AF (n=118) or AF associated with heart disease (n=257). Controls (n=360) from the same population were then genotyped for the presence of mutations or rare variants identified in the AF cases. In 10 probands (2.7%), 8 novel variants not found in the control population (0%; P=0.001) were identified. All variants affect highly conserved residues in the SCN5A protein. In 6 families with >1 affected member, the novel variant cosegregated with AF. We also identified 11 rare missense variants in 12 probands (3.2%) that have previously been associated with inherited arrhythmia syndromes (eg, congenital long-QT syndrome and Brugada syndrome). CONCLUSIONS: Mutations or rare variants in SCN5A may predispose patients with or without underlying heart disease to AF. The findings of the present study expand the clinical spectrum of disorders of the cardiac sodium channel to include AF and represent important progress toward molecular phenotyping and directed rather than empirical therapy for this common arrhythmia.