J. Gustav Smith

Novel and conventional biomarkers for prediction of incident cardiovascular events in the community.

CONTEXT Prior studies have demonstrated conflicting results regarding how much information novel biomarkers add to cardiovascular risk assessment. OBJECTIVE To evaluate the utility of contemporary biomarkers for predicting cardiovascular risk when added to conventional risk factors. DESIGN, SETTING, AND PARTICIPANTS Cohort study of 5067 participants (mean age, 58 years; 60% women) without cardiovascular disease from Malmö, Sweden, who attended a baseline examination between 1991 and 1994. Participants underwent measurement of C-reactive protein (CRP), cystatin C, lipoprotein-associated phospholipase 2, midregional proadrenomedullin (MR-proADM), midregional proatrial natriuretic peptide, and N-terminal pro-B-type natriuretic peptide (N-BNP) and underwent follow-up until 2006 using the Swedish national hospital discharge and cause-of-death registers and the Stroke in Malmö register for first cardiovascular events (myocardial infarction, stroke, coronary death). MAIN OUTCOME MEASURES Incident cardiovascular and coronary events. RESULTS During median follow-up of 12.8 years, there were 418 cardiovascular and 230 coronary events. Models with conventional risk factors had C statistics of 0.758 (95% confidence interval [CI], 0.734 to 0.781) and 0.760 (0.730 to 0.789) for cardiovascular and coronary events, respectively. Biomarkers retained in backward-elimination models were CRP and N-BNP for cardiovascular events and MR-proADM and N-BNP for coronary events, which increased the C statistic by 0.007 (P = .04) and 0.009 (P = .08), respectively. The proportion of participants reclassified was modest (8% for cardiovascular risk, 5% for coronary risk). Net reclassification improvement was nonsignificant for cardiovascular events (0.0%; 95% CI, -4.3% to 4.3%) and coronary events (4.7%; 95% CI, -0.76% to 10.1%). Greater improvements were observed in analyses restricted to intermediate-risk individuals (cardiovascular events: 7.4%; 95% CI, 0.7% to 14.1%; P = .03; coronary events: 14.6%; 95% CI, 5.0% to 24.2%; P = .003). However, correct reclassification was almost entirely confined to down-classification of individuals without events rather than up-classification of those with events. CONCLUSIONS Selected biomarkers may be used to predict future cardiovascular events, but the gains over conventional risk factors are minimal. Risk classification improved in intermediate-risk individuals, mainly through the identification of those unlikely to develop events.

Meta-analysis identifies six new susceptibility loci for atrial fibrillation

Atrial fibrillation is a highly prevalent arrhythmia and a major risk factor for stroke, heart failure and death. We conducted a genome-wide association study (GWAS) in individuals of European ancestry, including 6,707 with and 52,426 without atrial fibrillation. Six new atrial fibrillation susceptibility loci were identified and replicated in an additional sample of individuals of European ancestry, including 5,381 subjects with and 10,030 subjects without atrial fibrillation (P < 5 × 10−8). Four of the loci identified in Europeans were further replicated in silico in a GWAS of Japanese individuals, including 843 individuals with and 3,350 individuals without atrial fibrillation. The identified loci implicate candidate genes that encode transcription factors related to cardiopulmonary development, cardiac-expressed ion channels and cell signaling molecules.

Genetic Associations with Valvular Calcification and Aortic Stenosis

BACKGROUND Limited information is available regarding genetic contributions to valvular calcification, which is an important precursor of clinical valve disease. METHODS We determined genomewide associations with the presence of aortic-valve calcification (among 6942 participants) and mitral annular calcification (among 3795 participants), as detected by computed tomographic (CT) scanning; the study population for this analysis included persons of white European ancestry from three cohorts participating in the Cohorts for Heart and Aging Research in Genomic Epidemiology consortium (discovery population). Findings were replicated in independent cohorts of persons with either CT-detected valvular calcification or clinical aortic stenosis. RESULTS One SNP in the lipoprotein(a) (LPA) locus (rs10455872) reached genomewide significance for the presence of aortic-valve calcification (odds ratio per allele, 2.05; P=9.0×10(-10)), a finding that was replicated in additional white European, African-American, and Hispanic-American cohorts (P<0.05 for all comparisons). Genetically determined Lp(a) levels, as predicted by LPA genotype, were also associated with aortic-valve calcification, supporting a causal role for Lp(a). In prospective analyses, LPA genotype was associated with incident aortic stenosis (hazard ratio per allele, 1.68; 95% confidence interval [CI], 1.32 to 2.15) and aortic-valve replacement (hazard ratio, 1.54; 95% CI, 1.05 to 2.27) in a large Swedish cohort; the association with incident aortic stenosis was also replicated in an independent Danish cohort. Two SNPs (rs17659543 and rs13415097) near the proinflammatory gene IL1F9 achieved genomewide significance for mitral annular calcification (P=1.5×10(-8) and P=1.8×10(-8), respectively), but the findings were not replicated consistently. CONCLUSIONS Genetic variation in the LPA locus, mediated by Lp(a) levels, is associated with aortic-valve calcification across multiple ethnic groups and with incident clinical aortic stenosis. (Funded by the National Heart, Lung, and Blood Institute and others.).

Genetic risk, coronary heart disease events, and the clinical benefit of statin therapy: an analysis of primary and secondary prevention trials.

BACKGROUND Genetic variants have been associated with the risk of coronary heart disease. In this study, we tested whether or not a composite of these variants could ascertain the risk of both incident and recurrent coronary heart disease events and identify those individuals who derive greater clinical benefit from statin therapy. METHODS A community-based cohort study (the Malmo Diet and Cancer Study) and four randomised controlled trials of both primary prevention (JUPITER and ASCOT) and secondary prevention (CARE and PROVE IT-TIMI 22) with statin therapy, comprising a total of 48,421 individuals and 3477 events, were included in these analyses. We studied the association of a genetic risk score based on 27 genetic variants with incident or recurrent coronary heart disease, adjusting for traditional clinical risk factors. We then investigated the relative and absolute risk reductions in coronary heart disease events with statin therapy stratified by genetic risk. We combined data from the different studies using a meta-analysis. FINDINGS When individuals were divided into low (quintile 1), intermediate (quintiles 2-4), and high (quintile 5) genetic risk categories, a significant gradient in risk for incident or recurrent coronary heart disease was shown. Compared with the low genetic risk category, the multivariable-adjusted hazard ratio for coronary heart disease for the intermediate genetic risk category was 1·34 (95% CI 1·22-1·47, p<0·0001) and that for the high genetic risk category was 1·72 (1·55-1·92, p<0·0001). In terms of the benefit of statin therapy in the four randomised trials, we noted a significant gradient (p=0·0277) of increasing relative risk reductions across the low (13%), intermediate (29%), and high (48%) genetic risk categories. Similarly, we noted greater absolute risk reductions in those individuals in higher genetic risk categories (p=0·0101), resulting in a roughly threefold decrease in the number needed to treat to prevent one coronary heart disease event in the primary prevention trials. Specifically, in the primary prevention trials, the number needed to treat to prevent one such event in 10 years was 66 in people at low genetic risk, 42 in those at intermediate genetic risk, and 25 in those at high genetic risk in JUPITER, and 57, 47, and 20, respectively, in ASCOT. INTERPRETATION A genetic risk score identified individuals at increased risk for both incident and recurrent coronary heart disease events. People with the highest burden of genetic risk derived the largest relative and absolute clinical benefit from statin therapy. FUNDING National Institutes of Health.

Assessment of conventional cardiovascular risk factors and multiple biomarkers for the prediction of incident heart failure and atrial fibrillation.

OBJECTIVES the purpose of this study was to assess the predictive accuracy of conventional cardiovascular risk factors for incident heart failure and atrial fibrillation, and the added benefit of multiple biomarkers reflecting diverse pathophysiological pathways. BACKGROUND heart failure and atrial fibrillation are interrelated cardiac diseases associated with substantial morbidity and mortality and increasing incidence. Data on prediction and prevention of these diseases in healthy individuals are limited. METHODS in 5,187 individuals from the community-based MDCS (Malmö Diet and Cancer Study), we studied the performance of conventional risk factors and 6 biomarkers including midregional pro-atrial natriuretic peptide (MR-proANP), N-terminal pro-B-type natriuretic peptide (NT-proBNP), midregional pro-adrenomedullin, cystatin C, C-reactive protein (CRP), and copeptin. RESULTS during a mean follow-up of 14 years, 112 individuals were diagnosed with heart failure and 284 individuals with atrial fibrillation. NT-proBNP (hazard ratio [HR]: 1.63 per SD, 95% confidence interval [CI]: 1.29 to 2.06, p < 0.001), CRP (HR: 1.57 per SD, 95% CI: 1.28 to 1.94, p < 0.001), and MR-proANP (HR: 1.26 per SD, 95% CI: 1.02 to 1.56, p = 0.03) predicted incident heart failure independently of conventional risk factors and other biomarkers. MR-proANP (HR: 1.62, 95% CI: 1.42 to 1.84, p < 0.001) and CRP (HR: 1.18, 95% CI: 1.03 to 1.34, p = 0.01) independently predicted atrial fibrillation. Addition of biomarkers to conventional risk factors improved c-statistics from 0.815 to 0.842 for heart failure and from 0.732 to 0.753 for atrial fibrillation and the integrated discrimination improvement for both diseases (p < 0.001). Net reclassification improvement (NRI) with biomarkers was observed in 22% of individuals for heart failure (NRI, p < 0.001) and in 7% for atrial fibrillation (NRI, p = 0.06), mainly due to up-classification of individuals who developed disease (heart failure: 29%, atrial fibrillation: 19%). Addition of CRP to natriuretic peptides did not improve discrimination or reclassification. CONCLUSIONS conventional cardiovascular risk factors predict incident heart failure and atrial fibrillation with reasonable accuracy in middle-age individuals free from disease. Natriuretic peptides, but not other biomarkers, improve discrimination modestly for both diseases above and beyond conventional risk factors and substantially improve risk classification for heart failure.

Integrating Genetic, Transcriptional, and Functional Analyses to Identify 5 Novel Genes for Atrial Fibrillation

Background— Atrial fibrillation (AF) affects >30 million individuals worldwide and is associated with an increased risk of stroke, heart failure, and death. AF is highly heritable, yet the genetic basis for the arrhythmia remains incompletely understood. Methods and Results— To identify new AF-related genes, we used a multifaceted approach, combining large-scale genotyping in 2 ethnically distinct populations, cis-eQTL (expression quantitative trait loci) mapping, and functional validation. Four novel loci were identified in individuals of European descent near the genes NEURL (rs12415501; relative risk [RR]=1.18; 95% confidence interval [CI], 1.13–1.23; P=6.5×10−16), GJA1 (rs13216675; RR=1.10; 95% CI, 1.06–1.14; P=2.2×10−8), TBX5 (rs10507248; RR=1.12; 95% CI, 1.08–1.16; P=5.7×10−11), and CAND2 (rs4642101; RR=1.10; 95% CI, 1.06–1.14; P=9.8×10−9). In Japanese, novel loci were identified near NEURL (rs6584555; RR=1.32; 95% CI, 1.26–1.39; P=2.0×10−25) and CUX2 (rs6490029; RR=1.12; 95% CI, 1.08–1.16; P=3.9×10−9). The top single-nucleotide polymorphisms or their proxies were identified as cis-eQTLs for the genes CAND2 (P=2.6×10−19), GJA1 (P=2.66×10−6), and TBX5 (P=1.36×10−5). Knockdown of the zebrafish orthologs of NEURL and CAND2 resulted in prolongation of the atrial action potential duration (17% and 45%, respectively). Conclusions— We have identified 5 novel loci for AF. Our results expand the diversity of genetic pathways implicated in AF and provide novel molecular targets for future biological and pharmacological investigation.

Circulating cardio-enriched microRNAs are associated with long-term prognosis following myocardial infarction

BackgroundIncreased levels of cardio-enriched microRNAs (miRNAs) have been described in patients with myocardial infarction (MI). We wanted to evaluate the diagnostic and prognostic potential of cardio-enriched miRNAs in patients presenting with a suspected acute coronary syndrome (ACS).MethodsCardio-enriched miRNAs (miR-1, miR-208b and miR-499-5p) were measured using real time PCR in plasma samples from 424 patients with suspected ACS treated in a coronary care unit. miRNAs were assessed for discrimination of a clinical diagnosis of myocardial infarction and for association with 30-day mortality and diagnosis of heart failure. Correlation with left ventricular systolic dysfunction as measured by the ejection fraction (LVEF) was also assessed. To confirm myocardial origin miRNA was measured during coronary artery bypass surgery.ResultsmiRNAs were higher in MI patients and correlated with LVEF (p < 0.001). Discrimination of MI was accurate for miR-208b (AUC = 0.82) and miR-499-5p (AUC = 0.79) but considerable lower than for Troponin T (AUC = 0.95). Increased miRNA levels were strongly associated with increased risk of mortality or heart failure within 30 days for miR-208b (OR 1.79, 95% CI = 1.38-2.23, p = 1 × 10-5) and miR-499-5p (OR 1.70, 95% CI = 1.31-2.20, p = 5 × 10-5) but the association was lost when adjusting for Troponin T. During surgery miR-208b and miR-499-5p was released in the coronary sinus after cardioplegia-reperfusion to markedly higher levels than in a peripheral vein.ConclusionsOur findings confirm increased levels of cardio-enriched miRNAs in the blood of MI patients and establish association of increased miRNA levels with reduced systolic function after MI and risk of death or heart failure.

Genome-wide association studies of the PR interval in African Americans

The PR interval on the electrocardiogram reflects atrial and atrioventricular nodal conduction time. The PR interval is heritable, provides important information about arrhythmia risk, and has been suggested to differ among human races. Genome-wide association (GWA) studies have identified common genetic determinants of the PR interval in individuals of European and Asian ancestry, but there is a general paucity of GWA studies in individuals of African ancestry. We performed GWA studies in African American individuals from four cohorts (n = 6,247) to identify genetic variants associated with PR interval duration. Genotyping was performed using the Affymetrix 6.0 microarray. Imputation was performed for 2.8 million single nucleotide polymorphisms (SNPs) using combined YRI and CEU HapMap phase II panels. We observed a strong signal (rs3922844) within the gene encoding the cardiac sodium channel (SCN5A) with genome-wide significant association (p<2.5×10−8) in two of the four cohorts and in the meta-analysis. The signal explained 2% of PR interval variability in African Americans (beta  = 5.1 msec per minor allele, 95% CI  = 4.1–6.1, p = 3×10−23). This SNP was also associated with PR interval (beta = 2.4 msec per minor allele, 95% CI = 1.8–3.0, p = 3×10−16) in individuals of European ancestry (n = 14,042), but with a smaller effect size (p for heterogeneity <0.001) and variability explained (0.5%). Further meta-analysis of the four cohorts identified genome-wide significant associations with SNPs in SCN10A (rs6798015), MEIS1 (rs10865355), and TBX5 (rs7312625) that were highly correlated with SNPs identified in European and Asian GWA studies. African ancestry was associated with increased PR duration (13.3 msec, p = 0.009) in one but not the other three cohorts. Our findings demonstrate the relevance of common variants to African Americans at four loci previously associated with PR interval in European and Asian samples and identify an association signal at one of these loci that is more strongly associated with PR interval in African Americans than in Europeans.

Association of Low-Density Lipoprotein Cholesterol–Related Genetic Variants With Aortic Valve Calcium and Incident Aortic Stenosis

IMPORTANCE Plasma low-density lipoprotein cholesterol (LDL-C) has been associated with aortic stenosis in observational studies; however, randomized trials with cholesterol-lowering therapies in individuals with established valve disease have failed to demonstrate reduced disease progression. OBJECTIVE To evaluate whether genetic data are consistent with an association between LDL-C, high-density lipoprotein cholesterol (HDL-C), or triglycerides (TG) and aortic valve disease. DESIGN, SETTING, AND PARTICIPANTS Using a Mendelian randomization study design, we evaluated whether weighted genetic risk scores (GRSs), a measure of the genetic predisposition to elevations in plasma lipids, constructed using single-nucleotide polymorphisms identified in genome-wide association studies for plasma lipids, were associated with aortic valve disease. We included community-based cohorts participating in the CHARGE consortium (n = 6942), including the Framingham Heart Study (cohort inception to last follow-up: 1971-2013; n = 1295), Multi-Ethnic Study of Atherosclerosis (2000-2012; n = 2527), Age Gene/Environment Study-Reykjavik (2000-2012; n = 3120), and the Malmö Diet and Cancer Study (MDCS, 1991-2010; n = 28,461). MAIN OUTCOMES AND MEASURES Aortic valve calcium quantified by computed tomography in CHARGE and incident aortic stenosis in the MDCS. RESULTS The prevalence of aortic valve calcium across the 3 CHARGE cohorts was 32% (n = 2245). In the MDCS, over a median follow-up time of 16.1 years, aortic stenosis developed in 17 per 1000 participants (n = 473) and aortic valve replacement for aortic stenosis occurred in 7 per 1000 (n = 205). Plasma LDL-C, but not HDL-C or TG, was significantly associated with incident aortic stenosis (hazard ratio [HR] per mmol/L, 1.28; 95% CI, 1.04-1.57; P = .02; aortic stenosis incidence: 1.3% and 2.4% in lowest and highest LDL-C quartiles, respectively). The LDL-C GRS, but not HDL-C or TG GRS, was significantly associated with presence of aortic valve calcium in CHARGE (odds ratio [OR] per GRS increment, 1.38; 95% CI, 1.09-1.74; P = .007) and with incident aortic stenosis in MDCS (HR per GRS increment, 2.78; 95% CI, 1.22-6.37; P = .02; aortic stenosis incidence: 1.9% and 2.6% in lowest and highest GRS quartiles, respectively). In sensitivity analyses excluding variants weakly associated with HDL-C or TG, the LDL-C GRS remained associated with aortic valve calcium (P = .03) and aortic stenosis (P = .009). In instrumental variable analysis, LDL-C was associated with an increase in the risk of incident aortic stenosis (HR per mmol/L, 1.51; 95% CI, 1.07-2.14; P = .02). CONCLUSIONS AND RELEVANCE Genetic predisposition to elevated LDL-C was associated with presence of aortic valve calcium and incidence of aortic stenosis, providing evidence supportive of a causal association between LDL-C and aortic valve disease. Whether earlier intervention to reduce LDL-C could prevent aortic valve disease merits further investigation.

Genome-Wide Association Studies in an Isolated Founder Population from the Pacific Island of Kosrae

It has been argued that the limited genetic diversity and reduced allelic heterogeneity observed in isolated founder populations facilitates discovery of loci contributing to both Mendelian and complex disease. A strong founder effect, severe isolation, and substantial inbreeding have dramatically reduced genetic diversity in natives from the island of Kosrae, Federated States of Micronesia, who exhibit a high prevalence of obesity and other metabolic disorders. We hypothesized that genetic drift and possibly natural selection on Kosrae might have increased the frequency of previously rare genetic variants with relatively large effects, making these alleles readily detectable in genome-wide association analysis. However, mapping in large, inbred cohorts introduces analytic challenges, as extensive relatedness between subjects violates the assumptions of independence upon which traditional association test statistics are based. We performed genome-wide association analysis for 15 quantitative traits in 2,906 members of the Kosrae population, using novel approaches to manage the extreme relatedness in the sample. As positive controls, we observe association to known loci for plasma cholesterol, triglycerides, and C-reactive protein and to a compelling candidate loci for thyroid stimulating hormone and fasting plasma glucose. We show that our study is well powered to detect common alleles explaining ≥5% phenotypic variance. However, no such large effects were observed with genome-wide significance, arguing that even in such a severely inbred population, common alleles typically have modest effects. Finally, we show that a majority of common variants discovered in Caucasians have indistinguishable effect sizes on Kosrae, despite the major differences in population genetics and environment.