Michael V. Holmes

Mendelian randomization in cardiometabolic disease: challenges in evaluating causality

Mendelian randomization (MR) is a burgeoning field that involves the use of genetic variants to assess causal relationships between exposures and outcomes. MR studies can be straightforward; for example, genetic variants within or near the encoding locus that is associated with protein concentrations can help to assess their causal role in disease. However, a more complex relationship between the genetic variants and an exposure can make findings from MR more difficult to interpret. In this Review, we describe some of these challenges in interpreting MR analyses, including those from studies using genetic variants to assess causality of multiple traits (such as branched-chain amino acids and risk of diabetes mellitus); studies describing pleiotropic variants (for example, C-reactive protein and its contribution to coronary heart disease); and those investigating variants that disrupt normal function of an exposure (for example, HDL cholesterol or IL-6 and coronary heart disease). Furthermore, MR studies on variants that encode enzymes responsible for the metabolism of an exposure (such as alcohol) are discussed, in addition to those assessing the effects of variants on time-dependent exposures (extracellular superoxide dismutase), cumulative exposures (LDL cholesterol), and overlapping exposures (triglycerides and non-HDL cholesterol). We elaborate on the molecular features of each relationship, and provide explanations for the likely causal associations. In doing so, we hope to contribute towards more reliable evaluations of MR findings.

Association Between Diabetes and Cause-Specific Mortality in Rural and Urban Areas of China.

Importance In China, diabetes prevalence has increased substantially in recent decades, but there are no reliable estimates of the excess mortality currently associated with diabetes. Objectives To assess the proportional excess mortality associated with diabetes and estimate the diabetes-related absolute excess mortality in rural and urban areas of China. Design, Setting, and Participants A 7-year nationwide prospective study of 512 869 adults aged 30 to 79 years from 10 (5 rural and 5 urban) regions in China, who were recruited between June 2004 and July 2008 and were followed up until January 2014. Exposures Diabetes (previously diagnosed or detected by screening) recorded at baseline. Main Outcomes and Measures All-cause and cause-specific mortality, collected through established death registries. Cox regression was used to estimate adjusted mortality rate ratio (RR) comparing individuals with diabetes vs those without diabetes at baseline. Results Among the 512 869 participants, the mean (SD) age was 51.5 (10.7) years, 59% (n = 302 618) were women, and 5.9% (n = 30 280) had diabetes (4.1% in rural areas, 8.1% in urban areas, 5.8% of men, 6.1% of women, 3.1% had been previously diagnosed, and 2.8% were detected by screening). During 3.64 million person-years of follow-up, there were 24 909 deaths, including 3384 among individuals with diabetes. Compared with adults without diabetes, individuals with diabetes had a significantly increased risk of all-cause mortality (1373 vs 646 deaths per 100 000; adjusted RR, 2.00 [95% CI, 1.93-2.08]), which was higher in rural areas than in urban areas (rural RR, 2.17 [95% CI, 2.07-2.29]; urban RR, 1.83 [95% CI, 1.73-1.94]). Presence of diabetes was associated with increased mortality from ischemic heart disease (3287 deaths; RR, 2.40 [95% CI, 2.19-2.63]), stroke (4444 deaths; RR, 1.98 [95% CI, 1.81-2.17]), chronic liver disease (481 deaths; RR, 2.32 [95% CI, 1.76-3.06]), infections (425 deaths; RR, 2.29 [95% CI, 1.76-2.99]), and cancer of the liver (1325 deaths; RR, 1.54 [95% CI, 1.28-1.86]), pancreas (357 deaths; RR, 1.84 [95% CI, 1.35-2.51]), female breast (217 deaths; RR, 1.84 [95% CI, 1.24-2.74]), and female reproductive system (210 deaths; RR, 1.81 [95% CI, 1.20-2.74]). For chronic kidney disease (365 deaths), the RR was higher in rural areas (18.69 [95% CI, 14.22-24.57]) than in urban areas (6.83 [95% CI, 4.73-9.88]). Among those with diabetes, 10% of all deaths (16% rural; 4% urban) were due to definite or probable diabetic ketoacidosis or coma (408 deaths). Conclusions and Relevance Among adults in China, diabetes was associated with increased mortality from a range of cardiovascular and noncardiovascular diseases. Although diabetes was more common in urban areas, it was associated with greater excess mortality in rural areas.

Cannabis use and risk of schizophrenia: a Mendelian randomization study.

Cannabis use is observationally associated with an increased risk of schizophrenia, but whether the relationship is causal is not known. Using a genetic approach, we took 10 independent genetic variants previously identified to associate with cannabis use in 32 330 individuals to determine the nature of the association between cannabis use and risk of schizophrenia. Genetic variants were employed as instruments to recapitulate a randomized controlled trial involving two groups (cannabis users vs nonusers) to estimate the causal effect of cannabis use on risk of schizophrenia in 34 241 cases and 45 604 controls from predominantly European descent. Genetically-derived estimates were compared with a meta-analysis of observational studies reporting ever use of cannabis and risk of schizophrenia or related disorders. Based on the genetic approach, use of cannabis was associated with increased risk of schizophrenia (odds ratio (OR) of schizophrenia for users vs nonusers of cannabis: 1.37; 95% confidence interval (CI), 1.09–1.67; P-value=0.007). The corresponding estimate from observational analysis was 1.43 (95% CI, 1.19–1.67; P-value for heterogeneity =0.76). The genetic markers did not show evidence of pleiotropic effects and accounting for tobacco exposure did not alter the association (OR of schizophrenia for users vs nonusers of cannabis, adjusted for ever vs never smoker: 1.41; 95% CI, 1.09–1.83). This adds to the substantial evidence base that has previously identified cannabis use to associate with increased risk of schizophrenia, by suggesting that the relationship is causal. Such robust evidence may inform public health messages about cannabis use, especially regarding its potential mental health consequences.

Association of Body Mass Index With Cardiometabolic Disease in the UK Biobank: A Mendelian Randomization Study

Importance Higher body mass index (BMI) is a risk factor for cardiometabolic disease; however, the underlying causal associations remain unclear. Objectives To use UK Biobank data to report causal estimates of the association between BMI and cardiometabolic disease outcomes and traits, such as pulse rate, using mendelian randomization. Design, Setting, and Participants Cross-sectional baseline data from a population-based cohort study including 119 859 UK Biobank participants with complete phenotypic (medical and sociodemographic) and genetic data. Participants attended 1 of 22 assessment centers across the United Kingdom between 2006 and 2010. The present study was conducted from May 1 to July 11, 2016. Main Outcomes and Measures Prevalence of hypertension, coronary heart disease, and type 2 diabetes were determined at assessment, based on self-report. Blood pressure was measured clinically. Participants self-reported sociodemographic information pertaining to relevant confounders. A polygenic risk score comprising 93 single-nucleotide polymorphisms associated with BMI from previous genome-wide association studies was constructed, and the genetic risk score was applied to derive causal estimates using a mendelian randomization approach. Results Of the 119 859 individuals included in the study, 56 816 (47.4%) were men; mean (SD) age was 56.87 (7.93) years. Mendelian randomization analysis showed significant positive associations between genetically instrumented higher BMI and risk of hypertension (odds ratio [OR] per 1-SD higher BMI, 1.64; 95% CI, 1.48-1.83; P = 1.1 × 10−19), coronary heart disease (OR, 1.35; 95% CI, 1.09-1.69; P = .007) and type 2 diabetes (OR, 2.53; 95% CI, 2.04-3.13; P = 1.5 × 10−17), as well as systolic blood pressure (&bgr; = 1.65 mm Hg; 95% CI, 0.78-2.52 mm Hg; P = 2.0 × 10−04) and diastolic blood pressure (&bgr;  = 1.37 mm Hg; 95% CI, 0.88-1.85 mm Hg; P = 3.6 × 10−08). These associations were independent of age, sex, Townsend deprivation scores, alcohol intake, and smoking history. Conclusions and Relevance The results of this study add to the burgeoning evidence of an association between higher BMI and increased risk of cardiometabolic diseases. This finding has relevance for public health policies in many countries with increasing obesity levels.

Education and coronary heart disease: mendelian randomisation study

Objective To determine whether educational attainment is a causal risk factor in the development of coronary heart disease. Design Mendelian randomisation study, using genetic data as proxies for education to minimise confounding. Setting The main analysis used genetic data from two large consortia (CARDIoGRAMplusC4D and SSGAC), comprising 112 studies from predominantly high income countries. Findings from mendelian randomisation analyses were then compared against results from traditional observational studies (164 170 participants). Finally, genetic data from six additional consortia were analysed to investigate whether longer education can causally alter the common cardiovascular risk factors. Participants The main analysis was of 543 733 men and women (from CARDIoGRAMplusC4D and SSGAC), predominantly of European origin. Exposure A one standard deviation increase in the genetic predisposition towards higher education (3.6 years of additional schooling), measured by 162 genetic variants that have been previously associated with education. Main outcome measure Combined fatal and non-fatal coronary heart disease (63 746 events in CARDIoGRAMplusC4D). Results Genetic predisposition towards 3.6 years of additional education was associated with a one third lower risk of coronary heart disease (odds ratio 0.67, 95% confidence interval 0.59 to 0.77; P=3×10−8). This was comparable to findings from traditional observational studies (prevalence odds ratio 0.73, 0.68 to 0.78; incidence odds ratio 0.80, 0.76 to 0.83). Sensitivity analyses were consistent with a causal interpretation in which major bias from genetic pleiotropy was unlikely, although this remains an untestable possibility. Genetic predisposition towards longer education was additionally associated with less smoking, lower body mass index, and a favourable blood lipid profile. Conclusions This mendelian randomisation study found support for the hypothesis that low education is a causal risk factor in the development of coronary heart disease. Potential mechanisms could include smoking, body mass index, and blood lipids. In conjunction with the results from studies with other designs, these findings suggest that increasing education may result in substantial health benefits.

Deciphering the Causal Role of sPLA2s and Lp-PLA2 in Coronary Heart Disease

Over the last 10 to 15 years, animal and human observational studies have identified elevated levels of both proinflammatory secretory phospholipase A2-IIA and lipoprotein-associated phospholipase A2 as potential risk factors for coronary heart disease. However, Mendelian randomization, a genetic tool to test causality of a biomarker, and phase III randomized controlled trials of inhibitors of theses enzymes (varespladib and darapladib) converged to indicate that elevated levels are unlikely to be themselves causal of coronary heart disease and that inhibition had little or no clinical utility. The concordance of findings from Mendelian randomization and clinical trials suggests that for these 2 drugs, and for other novel biomarkers in future, validation of potential therapeutic targets by genetic studies (such as Mendelian randomization) before embarking on costly phase III randomized controlled trials could increase efficiency and offset the high risk of drug development, thereby facilitating discovery of new therapeutics and mitigating against the exuberant costs of drug development.

Evaluation of type 2 diabetes genetic risk variants in Chinese adults: findings from 93,000 individuals from the China Kadoorie Biobank.

Aims/hypothesisGenome-wide association studies (GWAS) have discovered many risk variants for type 2 diabetes. However, estimates of the contributions of risk variants to type 2 diabetes predisposition are often based on highly selected case–control samples, and reliable estimates of population-level effect sizes are missing, especially in non-European populations.MethodsThe individual and cumulative effects of 59 established type 2 diabetes risk loci were measured in a population-based China Kadoorie Biobank (CKB) study of 93,000 Chinese adults, including >7,100 diabetes cases.ResultsAssociation signals were directionally consistent between CKB and the original discovery GWAS: of 56 variants passing quality control, 48 showed the same direction of effect (binomial test, p = 2.3 × 10−8). We observed a consistent overall trend towards lower risk variant effect sizes in CKB than in case–control samples of GWAS meta-analyses (mean 19–22% decrease in log odds, p ≤ 0.0048), likely to reflect correction of both ‘winner’s curse’ and spectrum bias effects. The association with risk of diabetes of a genetic risk score, based on lead variants at 25 loci considered to act through beta cell function, demonstrated significant interactions with several measures of adiposity (BMI, waist circumference [WC], WHR and percentage body fat [PBF]; all pinteraction < 1 × 10−4), with a greater effect being observed in leaner adults.Conclusions/interpretationOur study provides further evidence of shared genetic architecture for type 2 diabetes between Europeans and East Asians. It also indicates that even very large GWAS meta-analyses may be vulnerable to substantial inflation of effect size estimates, compared with those observed in large-scale population-based cohort studies.Access to research materialsDetails of how to access China Kadoorie Biobank data and details of the data release schedule are available from www.ckbiobank.org/site/Data+Access.

Diabetes, plasma glucose and incidence of pancreatic cancer: A prospective study of 0.5 million Chinese adults and a meta-analysis of 22 cohort studies.

Diabetes is associated with an increased risk of pancreatic cancer (PC) in Western populations. Uncertainty remains, however, about the relevance of plasma glucose for PC among people without diabetes and about the associations of diabetes and high blood glucose with PC in China where the increase in diabetes prevalence has been very recent. The prospective China Kadoorie Biobank (CKB) study recruited 512,000 adults aged 30‐79 years from 10 diverse areas of China during 2004‐2008, recording 595 PC cases during 8 years of follow‐up. Cox regression yielded adjusted hazard ratios (HRs) for PC associated with diabetes (previously diagnosed or screen‐detected) and, among those without previously diagnosed diabetes, with levels of random plasma glucose (RPG). These were further meta‐analysed with 22 published prospective studies. Overall 5.8% of CKB participants had diabetes at baseline. Diabetes was associated with almost twofold increased risk of PC (adjusted HR = 1.87, 95% CI 1.48‐2.37), with excess risk higher in those with longer duration since diagnosis (p for trend = 0.01). Among those without previously diagnosed diabetes, each 1 mmol/L higher usual RPG was associated with a HR of 1.12 (1.04‐1.21). In meta‐analysis of CKB and 22 other studies, previously diagnosed diabetes was associated with a 52% excess risk (1.52, 1.43‐1.63). Among those without diabetes, each 1 mmol/L higher blood glucose was associated with a 15% (1.15, 1.09‐1.21) excess risk. In Chinese and non‐Chinese populations, diabetes and higher blood glucose levels among those without diabetes are associated with an increased risk of PC.

Long-Term Survival and Freedom From Reintervention After Off-Pump Coronary Artery Bypass GraftingClinical Perspective

Background: The long-term outcomes of off-pump coronary artery bypass grafting (CABG) are the subject of speculation. Our institution has >15 years of experience performing CABG both off-pump (OPCAB) and on cardiopulmonary bypass (CPB). Our null hypothesis was that there would be no difference in a long-term composite of death and revascularisation between the 2 methods. Methods: We performed a retrospective cohort study of all isolated CABG at our institution from 2001 to 2015. We used an intention-to-treat analysis, performing risk adjustment with adjustment for and matching to propensity score. In total, 13 226 patients had CABG: 5882 had OPCAB and 7344 had CPB, with a median follow-up of 6.2 years. Results: Of the 5882 OPCAB, 76 (1.3%) converted to CPB. One-, 5-, and 10-year survivals in each group were similar (OPCAB vs CPB: 96.7%, 87.9%, 72.1% vs 96.2%, 87.4%, 72.8%). No difference was found in long-term survival (adjusted hazards ratio [HR] 1.03; 95% confidence interval [CI]: 0.94–1.11 for OPCAB vs CPB; P=0.56) or freedom from death and reintervention (HR 0.98; 95% CI: 0.92–1.06 for OPCAB vs CPB; P=0.23). Patients receiving OPCAB had higher EuroSCOREs (median [quartiles]: 2.81 [1.53–5.57] vs 2.73 [1.51–5.22]; P=0.01), fewer grafts (mean±SD: 3.0±0.9 vs 3.3±0.9; P<0.001), but more total arterial grafting (45.9% vs 8.4%; P<0.001). OPCAB also had more trainee first operators (15.3% vs 12.5%), lower cardiac enzyme rise, shorter length of stay, and fewer complications (such as myocardial infarction). Conclusions: OPCAB is associated with similar long-term outcomes to CABG performed on CPB in our institution. Our low conversion rate to CPB, while training junior surgeons, demonstrates that OPCAB can be taught safely. The number of grafts performed between the 2 approaches is clinically comparable, if statistically different, and appears to provide equal benefits to survival and freedom from reintervention as on-pump CABG.

Genetic Support for a Causal Role of Insulin Resistance on Circulating Branched-Chain Amino Acids and Inflammation

OBJECTIVE Insulin resistance has deleterious effects on cardiometabolic disease. We used Mendelian randomization analyses to clarify the causal relationships of insulin resistance (IR) on circulating blood-based metabolites to shed light on potential mediators of the IR to cardiometabolic disease relationship. RESEARCH DESIGN AND METHODS We used 53 single nucleotide polymorphisms associated with IR from a recent genome-wide association study (GWAS) to explore their effects on circulating lipids and metabolites. We used published summary-level data from two GWASs of European individuals; data on the exposure (IR) were obtained from meta-GWASs of 188,577 individuals, and data on the outcomes (58 metabolic measures assessed by nuclear magnetic resonance) were taken from a GWAS of 24,925 individuals. RESULTS One-SD genetically elevated IR (equivalent to 55% higher geometric mean of fasting insulin, 0.89 mmol/L higher triglycerides, and 0.46 mmol/L lower HDL cholesterol) was associated with higher concentrations of all branched-chain amino acids (BCAAs)—isoleucine (0.56 SD; 95% CI 0.43, 0.70), leucine (0.42 SD; 95% CI 0.28, 0.55), and valine (0.26 SD; 95% CI 0.12, 0.39)—as well as with higher glycoprotein acetyls (an inflammation marker) (0.47 SD; 95% CI 0.32, 0.62) (P < 0.0003 for each). Results were broadly consistent when using multiple sensitivity analyses to account for potential genetic pleiotropy. CONCLUSIONS We provide robust evidence that IR causally affects each individual BCAA and inflammation. Taken together with existing studies, this implies that BCAA metabolism lies on a causal pathway from adiposity and IR to type 2 diabetes.