Protein Identification for Stroke Progression via Mendelian Randomization in Million Veteran Program and UK Biobank.

BACKGROUND: Individuals who have experienced a stroke, or transient ischemic attack, face a heightened risk of future cardiovascular events. Identification of genetic and molecular risk factors for subsequent cardiovascular outcomes may identify effective therapeutic targets to improve prognosis after an incident stroke. METHODS: We performed genome-wide association studies for subsequent major adverse cardiovascular events (MACE; ncases=51 929; ncontrols=39 980) and subsequent arterial ischemic stroke (AIS; ncases=45 120; ncontrols=46 789) after the first incident stroke within the Million Veteran Program and UK Biobank. We then used genetic variants associated with proteins (protein quantitative trait loci) to determine the effect of 1463 plasma protein abundances on subsequent MACE using Mendelian randomization. RESULTS: Two variants were significantly associated with subsequent cardiovascular events: rs76472767 near gene RNF220 (odds ratio, 0.75 [95% CI, 0.64-0.85]; P=3.69×10-8) with subsequent AIS and rs13294166 near gene LINC01492 (odds ratio, 1.52 [95% CI, 1.37-1.67]; P=3.77×10-8) with subsequent MACE. Using Mendelian randomization, we identified 2 proteins with an effect on subsequent MACE after a stroke: CCL27 ([C-C motif chemokine 27], effect odds ratio, 0.77 [95% CI, 0.66-0.88]; adjusted P=0.05) and TNFRSF14 ([tumor necrosis factor receptor superfamily member 14], effect odds ratio, 1.42 [95% CI, 1.24-1.60]; adjusted P=0.006). These proteins are not associated with incident AIS and are implicated to have a role in inflammation. CONCLUSIONS: We found evidence that 2 proteins with little effect on incident stroke appear to influence subsequent MACE after incident AIS. These associations suggest that inflammation is a contributing factor to subsequent MACE outcomes after incident AIS and highlights potential novel targets.

Twin-based Mendelian Randomization Analyses Highlight Smoking's Effects on Blood DNA Methylation, with Putative Reverse Causation.

Epigenome-wide association studies (EWAS) aim to identify differentially methylated loci associated with complex traits and disorders. EWAS of cigarette smoking shows some of the most widespread DNA methylation (DNAm) associations in blood. However, traditional EWAS cannot differentiate between causation and confounding, leading to ambiguity in etiological interpretations. Here, we apply an integrated approach combining Mendelian Randomization and twin-based Direction-of-Causation analyses (MR-DoC) to examine causality underlying smoking-associated blood DNAm changes in the Netherlands Twin Register (N=2577). Evidence across models suggests that current smoking's causal effects on DNAm likely drive many of the previous EWAS findings, implicating functional pathways relevant to several adverse health outcomes of smoking, including hemopoiesis, cell- and neuro-development, and immune regulation. Additionally, we find evidence of potential reverse causal influences at some DNAm sites, with 17 of these sites enriched for gene regulatory functional elements in the brain. The top three sites with evidence of DNAm's effects on smoking annotate to genes involved in G protein-coupled receptor signaling (GNG7, RGS3) and innate immune response (SLC15A4), elucidating potential biological risk factors for smoking. This study highlights the utility of integrating genotypic and DNAm measures in twin cohorts to clarify the causal relationships between health behaviors and blood DNAm.

The association between genetically elevated polyunsaturated fatty acids and risk of cancer.

BACKGROUND: The causal relevance of polyunsaturated fatty acids (PUFAs) for risk of site-specific cancers remains uncertain. METHODS: Using a Mendelian randomization (MR) framework, we assessed the causal relevance of PUFAs for risk of cancer in European and East Asian ancestry individuals. We defined the primary exposure as PUFA desaturase activity, proxied by rs174546 at the FADS locus. Secondary exposures were defined as omega 3 and omega 6 PUFAs that could be proxied by genetic polymorphisms outside the FADS region. Our study used summary genetic data on 10 PUFAs and 67 cancers, corresponding to 562,871 cases and 1,619,465 controls, collected by the Fatty Acids in Cancer Mendelian Randomization Collaboration. We estimated odds ratios (ORs) for cancer per standard deviation increase in genetically proxied PUFA exposures. FINDINGS: Genetically elevated PUFA desaturase activity was associated (P < 0.0007) with higher risk (OR [95% confidence interval]) of colorectal cancer (1.09 [1.07-1.11]), esophageal squamous cell carcinoma (1.16 [1.06-1.26]), lung cancer (1.06 [1.03-1.08]) and basal cell carcinoma (1.05 [1.02-1.07]). There was little evidence for associations with reproductive cancers (OR = 1.00 [95% CI: 0.99-1.01]; Pheterogeneity = 0.25), urinary system cancers (1.03 [0.99-1.06], Pheterogeneity = 0.51), nervous system cancers (0.99 [0.95-1.03], Pheterogeneity = 0.92) or blood cancers (1.01 [0.98-1.04], Pheterogeneity = 0.09). Findings for colorectal cancer and esophageal squamous cell carcinoma remained compatible with causality in sensitivity analyses for violations of assumptions. Secondary MR analyses highlighted higher omega 6 PUFAs (arachidonic acid, gamma-linolenic acid and dihomo-gamma-linolenic acid) as potential mediators. PUFA biosynthesis is known to interact with aspirin, which increases risk of bleeding and inflammatory bowel disease. In a phenome-wide MR study of non-neoplastic diseases, we found that genetic lowering of PUFA desaturase activity, mimicking a hypothetical intervention to reduce cancer risk, was associated (P < 0.0006) with increased risk of inflammatory bowel disease but not bleeding. INTERPRETATION: The PUFA biosynthesis pathway may be an intervention target for prevention of colorectal cancer and esophageal squamous cell carcinoma but with potential for increased risk of inflammatory bowel disease. FUNDING: Cancer Resesrch UK (C52724/A20138, C18281/A19169). UK Medical Research Council (MR/P014054/1). National Institute for Health Research (NIHR202411). UK Medical Research Council (MC_UU_00011/1, MC_UU_00011/3, MC_UU_00011/6, and MC_UU_00011/4). National Cancer Institute (R00 CA215360). National Institutes of Health (U01 CA164973, R01 CA60987, R01 CA72520, U01 CA74806, R01 CA55874, U01 CA164973 and U01 CA164973).

Design and quality control of large-scale two-sample Mendelian randomization studies.

Background: Mendelian randomization (MR) studies are susceptible to metadata errors (e.g. incorrect specification of the effect allele column) and other analytical issues that can introduce substantial bias into analyses. We developed a quality control (QC) pipeline for the Fatty Acids in Cancer Mendelian Randomization Collaboration (FAMRC) that can be used to identify and correct for such errors. Methods: We collated summary association statistics from fatty acid and cancer genome-wide association studies (GWAS) and subjected the collated data to a comprehensive QC pipeline. We identified metadata errors through comparison of study-specific statistics to external reference data sets (the National Human Genome Research Institute-European Bioinformatics Institute GWAS catalogue and 1000 genome super populations) and other analytical issues through comparison of reported to expected genetic effect sizes. Comparisons were based on three sets of genetic variants: (i) GWAS hits for fatty acids, (ii) GWAS hits for cancer and (iii) a 1000 genomes reference set. Results: We collated summary data from 6 fatty acid and 54 cancer GWAS. Metadata errors and analytical issues with the potential to introduce substantial bias were identified in seven studies (11.6%). After resolving metadata errors and analytical issues, we created a data set of 219 842 genetic associations with 90 cancer types, generated in analyses of 566 665 cancer cases and 1 622 374 controls. Conclusions: In this large MR collaboration, 11.6% of included studies were affected by a substantial metadata error or analytical issue. By increasing the integrity of collated summary data prior to their analysis, our protocol can be used to increase the reliability of downstream MR analyses. Our pipeline is available to other researchers via the CheckSumStats package (https://github.com/MRCIEU/CheckSumStats).

Applying Mendelian randomization to appraise causality in relationships between nutrition and cancer.

Dietary factors are assumed to play an important role in cancer risk, apparent in consensus recommendations for cancer prevention that promote nutritional changes. However, the evidence in this field has been generated predominantly through observational studies, which may result in biased effect estimates because of confounding, exposure misclassification, and reverse causality. With major geographical differences and rapid changes in cancer incidence over time, it is crucial to establish which of the observational associations reflect causality and to identify novel risk factors as these may be modified to prevent the onset of cancer and reduce its progression. Mendelian randomization (MR) uses the special properties of germline genetic variation to strengthen causal inference regarding potentially modifiable exposures and disease risk. MR can be implemented through instrumental variable (IV) analysis and, when robustly performed, is generally less prone to confounding, reverse causation and measurement error than conventional observational methods and has different sources of bias (discussed in detail below). It is increasingly used to facilitate causal inference in epidemiology and provides an opportunity to explore the effects of nutritional exposures on cancer incidence and progression in a cost-effective and timely manner. Here, we introduce the concept of MR and discuss its current application in understanding the impact of nutritional factors (e.g., any measure of diet and nutritional intake, circulating biomarkers, patterns, preference or behaviour) on cancer aetiology and, thus, opportunities for MR to contribute to the development of nutritional recommendations and policies for cancer prevention. We provide applied examples of MR studies examining the role of nutritional factors in cancer to illustrate how this method can be used to help prioritise or deprioritise the evaluation of specific nutritional factors as intervention targets in randomised controlled trials. We describe possible biases when using MR, and methodological developments aimed at investigating and potentially overcoming these biases when present. Lastly, we consider the use of MR in identifying causally relevant nutritional risk factors for various cancers in different regions across the world, given notable geographical differences in some cancers. We also discuss how MR results could be translated into further research and policy. We conclude that findings from MR studies, which corroborate those from other well-conducted studies with different and orthogonal biases, are poised to substantially improve our understanding of nutritional influences on cancer. For such corroboration, there is a requirement for an interdisciplinary and collaborative approach to investigate risk factors for cancer incidence and progression.

Polygenic risks for joint developmental trajectories of internalizing and externalizing problems: findings from the ALSPAC cohort.

BACKGROUND: Joint developmental trajectories of internalizing and externalizing problems show considerable heterogeneity; however, this can be parsed into a small number of meaningful subgroups. Doing so offered insights into risk factors that lead to different patterns of internalizing/externalizing trajectories. However, despite both domains of problems showing strong heritability, no study has yet considered genetic risks as predictors of joint internalizing/externalizing problem trajectories. METHODS: Using parallel process latent class growth analysis, we estimated joint developmental trajectories of internalizing and externalizing difficulties assessed across ages 4 to 16 using the Strengths and Difficulties Questionnaire. Multinomial logistic regression was used to evaluate a range of demographic, perinatal, maternal mental health, and child and maternal polygenic predictors of group membership. Participants included 11,049 children taking part in the Avon Longitudinal Study of Parents and Children. Polygenic data were available for 7,127 children and 6,836 mothers. RESULTS: A 5-class model was judged optimal: Unaffected, Moderate Externalizing Symptoms, High Externalizing Symptoms, Moderate Internalizing and Externalizing Symptoms and High Internalizing and Externalizing Symptoms. Male sex, lower maternal age, maternal mental health problems, maternal smoking during pregnancy, higher child polygenic risk scores for ADHD and lower polygenic scores for IQ distinguished affected classes from the unaffected class. CONCLUSIONS: While affected classes could be relatively well separated from the unaffected class, phenotypic and polygenic predictors were limited in their ability to distinguish between different affected classes. Results thus add to existing evidence that internalizing and externalizing problems have mostly shared risk factors.

Investigating DNA methylation as a potential mediator between pigmentation genes, pigmentary traits and skin cancer.

Pigmentation characteristics are well-known risk factors for skin cancer. Polymorphisms in pigmentation genes have been associated with these traits and with the risk of malignancy. However, the functional relationship between genetic variation and disease is still unclear. This study aims to assess whether pigmentation SNPs are associated with pigmentary traits and skin cancer via DNA methylation (DNAm). Using a meta-GWAS of whole-blood DNAm from 36 European cohorts (N = 27,750; the Genetics of DNA Methylation Consortium, GoDMC), we found that 19 out of 27 SNPs in 10 pigmentation genes were associated with 391 DNAm sites across 30 genomic regions. We examined the effect of 25 selected DNAm sites on pigmentation traits, sun exposure phenotypes and skin cancer and on gene expression in whole blood. We uncovered an association of DNAm site cg07402062 with red hair in the Avon Longitudinal Study of Parents and Children (ALSPAC). We also found that the expression of ASIP and CDK10 was associated with hair colour, melanoma and basal cell carcinoma. Our results indicate that DNAm and expression of pigmentation genes may play a role as potential mediators of the relationship between genetic variants, pigmentation phenotypes and skin cancer and thus deserve further scrutiny.

Evaluating shared genetic influences on nonsyndromic cleft lip/palate and oropharyngeal neoplasms.

It has been hypothesised that nonsyndromic cleft lip/palate (nsCL/P) and cancer may share aetiological risk factors. Population studies have found inconsistent evidence for increased incidence of cancer in nsCL/P cases, but several genes (e.g., CDH1, AXIN2) have been implicated in the aetiologies of both phenotypes. We aimed to evaluate shared genetic aetiology between nsCL/P and oral cavity/oropharyngeal cancers (OC/OPC), which affect similar anatomical regions. Using a primary sample of 5,048 OC/OPC cases and 5,450 controls of European ancestry and a replication sample of 750 cases and 336,319 controls from UK Biobank, we estimate genetic overlap using nsCL/P polygenic risk scores (PRS) with Mendelian randomization analyses performed to evaluate potential causal mechanisms. In the primary sample, we found strong evidence for an association between a nsCL/P PRS and increased odds of OC/OPC (per standard deviation increase in score, odds ratio [OR]: 1.09; 95% confidence interval [CI]: 1.04, 1.13; p = .000053). Although confidence intervals overlapped with the primary estimate, we did not find confirmatory evidence of an association between the PRS and OC/OPC in UK Biobank (OR 1.02; 95% CI: 0.95, 1.10; p = .55). Mendelian randomization analyses provided evidence that major nsCL/P risk variants are unlikely to influence OC/OPC. Our findings suggest possible shared genetic influences on nsCL/P and OC/OPC.

Bayesian reassessment of the epigenetic architecture of complex traits.

Linking epigenetic marks to clinical outcomes improves insight into molecular processes, disease prediction, and therapeutic target identification. Here, a statistical approach is presented to infer the epigenetic architecture of complex disease, determine the variation captured by epigenetic effects, and estimate phenotype-epigenetic probe associations jointly. Implicitly adjusting for probe correlations, data structure (cell-count or relatedness), and single-nucleotide polymorphism (SNP) marker effects, improves association estimates and in 9,448 individuals, 75.7% (95% CI 71.70-79.3) of body mass index (BMI) variation and 45.6% (95% CI 37.3-51.9) of cigarette consumption variation was captured by whole blood methylation array data. Pathway-linked probes of blood cholesterol, lipid transport and sterol metabolism for BMI, and xenobiotic stimuli response for smoking, showed >1.5 times larger associations with >95% posterior inclusion probability. Prediction accuracy improved by 28.7% for BMI and 10.2% for smoking over a LASSO model, with age-, and tissue-specificity, implying associations are a phenotypic consequence rather than causal.

The association of DNA methylation with body mass index: distinguishing between predictors and biomarkers.

BACKGROUND: DNA methylation is associated with body mass index (BMI), but it is not clear if methylation scores are biomarkers for extant BMI or predictive of future BMI. Here, we explore the causal nature and predictive utility of DNA methylation measured in peripheral blood with BMI and cardiometabolic traits. METHODS: Analyses were conducted across the life course using the ARIES cohort of mothers (n = 792) and children (n = 906), for whom DNA methylation and genetic profiles and BMI at multiple time points (3 in children at birth, in childhood and in adolescence; 2 in mothers during pregnancy and in middle age) were available. Genetic and DNA methylation scores for BMI were derived using published associations between BMI and DNA methylation and genotype. Causal relationships between methylation and BMI were assessed using Mendelian randomisation and cross-lagged models. RESULTS: The DNA methylation scores in adult women explained 10% of extant BMI variance. However, less extant variance was explained by scores generated in the same women during pregnancy (2% BMI variance) and in older children (15-17 years; 3% BMI variance). Similarly, little extant variance was explained in younger children (at birth and at 7 years; 1% and 2%, respectively). These associations remained following adjustment for smoking exposure and education levels. The DNA methylation score was found to be a poor predictor of future BMI using linear and cross-lagged models, suggesting that DNA methylation variation does not cause later variation in BMI. However, there was some evidence to suggest that BMI is predictive of later DNA methylation. Mendelian randomisation analyses also support this direction of effect, although evidence is weak. Finally, we find that DNA methylation scores for BMI are associated with extant cardiometabolic traits independently of BMI and genetic score. CONCLUSION: The age-specific nature of DNA methylation associations with BMI, lack of causal relationship and limited predictive ability of future BMI indicate that DNA methylation is likely influenced by BMI and might more accurately be considered a biomarker of BMI and related outcomes rather than a predictor. Future epigenome-wide association studies may benefit from further examining associations between early DNA methylation and later health outcomes.

Evidence for causal effects of lifetime smoking on risk for depression and schizophrenia: a Mendelian randomisation study.

BACKGROUND: Smoking prevalence is higher amongst individuals with schizophrenia and depression compared with the general population. Mendelian randomisation (MR) can examine whether this association is causal using genetic variants identified in genome-wide association studies (GWAS). METHODS: We conducted two-sample MR to explore the bi-directional effects of smoking on schizophrenia and depression. For smoking behaviour, we used (1) smoking initiation GWAS from the GSCAN consortium and (2) we conducted our own GWAS of lifetime smoking behaviour (which captures smoking duration, heaviness and cessation) in a sample of 462690 individuals from the UK Biobank. We validated this instrument using positive control outcomes (e.g. lung cancer). For schizophrenia and depression we used GWAS from the PGC consortium. RESULTS: There was strong evidence to suggest smoking is a risk factor for both schizophrenia (odds ratio (OR) 2.27, 95% confidence interval (CI) 1.67-3.08, p < 0.001) and depression (OR 1.99, 95% CI 1.71-2.32, p < 0.001). Results were consistent across both lifetime smoking and smoking initiation. We found some evidence that genetic liability to depression increases smoking (ß = 0.091, 95% CI 0.027-0.155, p = 0.005) but evidence was mixed for schizophrenia (ß = 0.022, 95% CI 0.005-0.038, p = 0.009) with very weak evidence for an effect on smoking initiation. CONCLUSIONS: These findings suggest that the association between smoking, schizophrenia and depression is due, at least in part, to a causal effect of smoking, providing further evidence for the detrimental consequences of smoking on mental health.

The Parkinson's Disease Mendelian Randomization Research Portal.

BACKGROUND: Mendelian randomization is a method for exploring observational associations to find evidence of causality. OBJECTIVE: To apply Mendelian randomization between risk factors/phenotypic traits (exposures) and PD in a large, unbiased manner, and to create a public resource for research. METHODS: We used two-sample Mendelian randomization in which the summary statistics relating to single-nucleotide polymorphisms from 5,839 genome-wide association studies of exposures were used to assess causal relationships with PD. We selected the highest-quality exposure genome-wide association studies for this report (n = 401). For the disease outcome, summary statistics from the largest published PD genome-wide association studies were used. For each exposure, the causal effect on PD was assessed using the inverse variance weighted method, followed by a range of sensitivity analyses. We used a false discovery rate of 5% from the inverse variance weighted analysis to prioritize exposures of interest. RESULTS: We observed evidence for causal associations between 12 exposures and risk of PD. Of these, nine were effects related to increasing adiposity and decreasing risk of PD. The remaining top three exposures that affected PD risk were tea drinking, time spent watching television, and forced vital capacity, but these may have been biased and were less convincing. Other exposures at nominal statistical significance included inverse effects of smoking and alcohol. CONCLUSIONS: We present a new platform which offers Mendelian randomization analyses for a total of 5,839 genome-wide association studies versus the largest PD genome-wide association studies available (https://pdgenetics.shinyapps.io/MRportal/). Alongside, we report further evidence to support a causal role for adiposity on lowering the risk of PD. © 2019 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.

Appraising the causal relevance of DNA methylation for risk of lung cancer.

BACKGROUND: DNA methylation changes in peripheral blood have recently been identified in relation to lung cancer risk. Some of these changes have been suggested to mediate part of the effect of smoking on lung cancer. However, limitations with conventional mediation analyses mean that the causal nature of these methylation changes has yet to be fully elucidated. METHODS: We first performed a meta-analysis of four epigenome-wide association studies (EWAS) of lung cancer (918 cases, 918 controls). Next, we conducted a two-sample Mendelian randomization analysis, using genetic instruments for methylation at CpG sites identified in the EWAS meta-analysis, and 29 863 cases and 55 586 controls from the TRICL-ILCCO lung cancer consortium, to appraise the possible causal role of methylation at these sites on lung cancer. RESULTS: Sixteen CpG sites were identified from the EWAS meta-analysis [false discovery rate (FDR) < 0.05], for 14 of which we could identify genetic instruments. Mendelian randomization provided little evidence that DNA methylation in peripheral blood at the 14 CpG sites plays a causal role in lung cancer development (FDR > 0.05), including for cg05575921-AHRR where methylation is strongly associated with both smoke exposure and lung cancer risk. CONCLUSIONS: The results contrast with previous observational and mediation analysis, which have made strong claims regarding the causal role of DNA methylation. Thus, previous suggestions of a mediating role of methylation at sites identified in peripheral blood, such as cg05575921-AHRR, could be unfounded. However, this study does not preclude the possibility that differential DNA methylation at other sites is causally involved in lung cancer development, especially within lung tissue.

Association of Schizophrenia Risk With Disordered Niacin Metabolism in an Indian Genome-wide Association Study.

Importance: Genome-wide association studies (GWASs) in European populations have identified more than 100 schizophrenia-associated loci. A schizophrenia GWAS in a unique Indian population offers novel findings. Objective: To discover and functionally evaluate genetic loci for schizophrenia in a GWAS of a unique Indian population. Design, Setting, and Participants: This GWAS included a sample of affected individuals, family members, and unrelated cases and controls. Three thousand ninety-two individuals were recruited and diagnostically ascertained via medical records, hospitals, clinics, and clinical networks in Chennai and surrounding regions. Affected participants fulfilled DSM-IV diagnostic criteria for schizophrenia. Unrelated control participants had no personal or family history of psychotic disorder. Recruitment, genotyping, and analysis occurred in consecutive phases beginning January 1, 2001. Recruitment was completed on February 28, 2018, and genotyping and analysis are ongoing. Main Outcomes and Measures: Associations of single-nucleotide polymorphisms and gene expression with schizophrenia. Results: The study population included 1321 participants with schizophrenia, 885 family controls, and 886 unrelated controls. Among participants with schizophrenia, mean (SD) age was 39.1 (11.4) years, and 52.7% were male. This sample demonstrated uniform ethnicity, a degree of inbreeding, and negligible rates of substance abuse. A novel genome-wide significant association was observed between schizophrenia and a chromosome 8q24.3 locus (rs10866912, allele A; odds ratio [OR], 1.27 [95% CI, 1.17-1.38]; P = 4.35 × 10-8) that attracted support in the schizophrenia Psychiatric Genomics Consortium 2 data (rs10866912, allele A; OR, 1.04 [95% CI, 1.02-1.06]; P = 7.56 × 10-4). This locus has undergone natural selection, with the risk allele A declining in frequency from India (approximately 72%) to Europe (approximately 43%). rs10866912 directly modifies the abundance of the nicotinate phosphoribosyltransferase gene (NAPRT1) transcript in brain cortex (normalized effect size, 0.79; 95% CI, 0.6-1.0; P = 5.8 × 10-13). NAPRT1 encodes a key enzyme for niacin metabolism. In Indian lymphoblastoid cell lines, (risk) allele A of rs10866912 was associated with NAPRT1 downregulation (AA: 0.74, n = 21; CC: 1.56, n = 17; P = .004). Preliminary zebrafish data further suggest that partial loss of function of NAPRT1 leads to abnormal brain development. Conclusions and Relevance: Bioinformatic analyses and cellular and zebrafish gene expression studies implicate NAPRT1 as a novel susceptibility gene. Given this gene's role in niacin metabolism and the evidence for niacin deficiency provoking schizophrenialike symptoms in neuropsychiatric diseases such as pellagra and Hartnup disease, these results suggest that the rs10866912 genotype and niacin status may have implications for schizophrenia susceptibility and treatment.

An integrative approach to detect epigenetic mechanisms that putatively mediate the influence of lifestyle exposures on disease susceptibility.

BACKGROUND: There is mounting evidence that our environment and lifestyle has an impact on epigenetic regulatory mechanisms, such as DNA methylation. It has been suggested that these molecular processes may mediate the effect of risk factors on disease susceptibility, although evidence in this regard has been challenging to uncover. Using genetic variants as surrogate variables, we have used two-sample Mendelian randomization (2SMR) to investigate the potential implications of putative changes to DNA methylation levels on disease susceptibility. METHODS: To illustrate our approach, we identified 412 CpG sites where DNA methylation was associated with prenatal smoking. We then applied 2SMR to investigate potential downstream effects of these putative changes on 643 complex traits using findings from large-scale genome-wide association studies. To strengthen evidence of mediatory mechanisms, we used multiple-trait colocalization to assess whether DNA methylation, nearby gene expression and complex trait variation were all influenced by the same causal genetic variant. RESULTS: We identified 22 associations that survived multiple testing (P < 1.89 × 10-7). In-depth follow-up analyses of particular note suggested that the associations between DNA methylation at the ASPSCR1 and REST/POL2RB gene regions, both linked with reduced lung function, may be mediated by changes in gene expression. We validated associations between DNA methylation and traits using independent samples from different stages across the life course. CONCLUSION: Our approach should prove valuable in prioritizing CpG sites that may mediate the effect of causal risk factors on disease. In-depth evaluations of findings are necessary to robustly disentangle causality from alternative explanations such as horizontal pleiotropy.

A Phenome-Wide Mendelian Randomization Study of Pancreatic Cancer Using Summary Genetic Data.

BACKGROUND: The 5-year mortality rate for pancreatic cancer is among the highest of all cancers. Greater understanding of underlying causes could inform population-wide intervention strategies for prevention. Summary genetic data from genome-wide association studies (GWAS) have become available for thousands of phenotypes. These data can be exploited in Mendelian randomization (MR) phenome-wide association studies (PheWAS) to efficiently screen the phenome for potential determinants of disease risk. METHODS: We conducted an MR-PheWAS of pancreatic cancer using 486 phenotypes, proxied by 9,124 genetic variants, and summary genetic data from a GWAS of pancreatic cancer (7,110 cancer cases, 7,264 controls). ORs and 95% confidence intervals per 1 SD increase in each phenotype were generated. RESULTS: We found evidence that previously reported risk factors of body mass index (BMI; 1.46; 1.20-1.78) and hip circumference (1.42; 1.21-1.67) were associated with pancreatic cancer. We also found evidence of novel associations with metabolites that have not previously been implicated in pancreatic cancer: ADpSGEGDFXAEGGGVR*, a fibrinogen-cleavage peptide (1.60; 1.31-1.95), and O-sulfo-l-tyrosine (0.58; 0.46-0.74). An inverse association was also observed with lung adenocarcinoma (0.63; 0.54-0.74). CONCLUSIONS: Markers of adiposity (BMI and hip circumference) are potential intervention targets for pancreatic cancer prevention. Further clarification of the causal relevance of the fibrinogen-cleavage peptides and O-sulfo-l-tyrosine in pancreatic cancer etiology is required, as is the basis of our observed association with lung adenocarcinoma. IMPACT: For pancreatic cancer, MR-PheWAS can augment existing risk factor knowledge and generate novel hypotheses to investigate.

Understanding the consequences of education inequality on cardiovascular disease: mendelian randomisation study.

OBJECTIVES: To investigate the role of body mass index (BMI), systolic blood pressure, and smoking behaviour in explaining the effect of education on the risk of cardiovascular disease outcomes. DESIGN: Mendelian randomisation study. SETTING: UK Biobank and international genome-wide association study data. PARTICIPANTS: Predominantly participants of European ancestry. EXPOSURE: Educational attainment, BMI, systolic blood pressure, and smoking behaviour in observational analysis, and randomly allocated genetic variants to instrument these traits in mendelian randomisation. MAIN OUTCOMES MEASURE: The risk of coronary heart disease, stroke, myocardial infarction, and cardiovascular disease (all subtypes; all measured in odds ratio), and the degree to which this is mediated through BMI, systolic blood pressure, and smoking behaviour respectively. RESULTS: Each additional standard deviation of education (3.6 years) was associated with a 13% lower risk of coronary heart disease (odds ratio 0.86, 95% confidence interval 0.84 to 0.89) in observational analysis and a 37% lower risk (0.63, 0.60 to 0.67) in mendelian randomisation analysis. As a proportion of the total risk reduction, BMI was estimated to mediate 15% (95% confidence interval 13% to 17%) and 18% (14% to 23%) in the observational and mendelian randomisation estimates, respectively. Corresponding estimates were 11% (9% to 13%) and 21% (15% to 27%) for systolic blood pressure and 19% (15% to 22%) and 34% (17% to 50%) for smoking behaviour. All three risk factors combined were estimated to mediate 42% (36% to 48%) and 36% (5% to 68%) of the effect of education on coronary heart disease in observational and mendelian randomisation analyses, respectively. Similar results were obtained when investigating the risk of stroke, myocardial infarction, and cardiovascular disease. CONCLUSIONS: BMI, systolic blood pressure, and smoking behaviour mediate a substantial proportion of the protective effect of education on the risk of cardiovascular outcomes and intervening on these would lead to reductions in cases of cardiovascular disease attributable to lower levels of education. However, more than half of the protective effect of education remains unexplained and requires further investigation.

Shared polygenic risk and causal inferences in amyotrophic lateral sclerosis.

OBJECTIVE: To identify shared polygenic risk and causal associations in amyotrophic lateral sclerosis (ALS). METHODS: Linkage disequilibrium score regression and Mendelian randomization were applied in a large-scale, data-driven manner to explore genetic correlations and causal relationships between >700 phenotypic traits and ALS. Exposures consisted of publicly available genome-wide association studies (GWASes) summary statistics from MR Base and LD-hub. The outcome data came from the recently published ALS GWAS involving 20,806 cases and 59,804 controls. Multivariate analyses, genetic risk profiling, and Bayesian colocalization analyses were also performed. RESULTS: We have shown, by linkage disequilibrium score regression, that ALS shares polygenic risk genetic factors with a number of traits and conditions, including positive correlations with smoking status and moderate levels of physical activity, and negative correlations with higher cognitive performance, higher educational attainment, and light levels of physical activity. Using Mendelian randomization, we found evidence that hyperlipidemia is a causal risk factor for ALS and localized putative functional signals within loci of interest. INTERPRETATION: Here, we have developed a public resource (https://lng-nia.shinyapps.io/mrshiny) which we hope will become a valuable tool for the ALS community, and that will be expanded and updated as new data become available. Shared polygenic risk exists between ALS and educational attainment, physical activity, smoking, and tenseness/restlessness. We also found evidence that elevated low-desnity lipoprotein cholesterol is a causal risk factor for ALS. Future randomized controlled trials should be considered as a proof of causality. Ann Neurol 2019;85:470-481.

Association Between Telomere Length and Risk of Cancer and Non-Neoplastic Diseases: A Mendelian Randomization Study.

IMPORTANCE: The causal direction and magnitude of the association between telomere length and incidence of cancer and non-neoplastic diseases is uncertain owing to the susceptibility of observational studies to confounding and reverse causation. OBJECTIVE: To conduct a Mendelian randomization study, using germline genetic variants as instrumental variables, to appraise the causal relevance of telomere length for risk of cancer and non-neoplastic diseases. DATA SOURCES: Genomewide association studies (GWAS) published up to January 15, 2015. STUDY SELECTION: GWAS of noncommunicable diseases that assayed germline genetic variation and did not select cohort or control participants on the basis of preexisting diseases. Of 163 GWAS of noncommunicable diseases identified, summary data from 103 were available. DATA EXTRACTION AND SYNTHESIS: Summary association statistics for single nucleotide polymorphisms (SNPs) that are strongly associated with telomere length in the general population. MAIN OUTCOMES AND MEASURES: Odds ratios (ORs) and 95% confidence intervals (CIs) for disease per standard deviation (SD) higher telomere length due to germline genetic variation. RESULTS: Summary data were available for 35 cancers and 48 non-neoplastic diseases, corresponding to 420 081 cases (median cases, 2526 per disease) and 1 093 105 controls (median, 6789 per disease). Increased telomere length due to germline genetic variation was generally associated with increased risk for site-specific cancers. The strongest associations (ORs [95% CIs] per 1-SD change in genetically increased telomere length) were observed for glioma, 5.27 (3.15-8.81); serous low-malignant-potential ovarian cancer, 4.35 (2.39-7.94); lung adenocarcinoma, 3.19 (2.40-4.22); neuroblastoma, 2.98 (1.92-4.62); bladder cancer, 2.19 (1.32-3.66); melanoma, 1.87 (1.55-2.26); testicular cancer, 1.76 (1.02-3.04); kidney cancer, 1.55 (1.08-2.23); and endometrial cancer, 1.31 (1.07-1.61). Associations were stronger for rarer cancers and at tissue sites with lower rates of stem cell division. There was generally little evidence of association between genetically increased telomere length and risk of psychiatric, autoimmune, inflammatory, diabetic, and other non-neoplastic diseases, except for coronary heart disease (OR, 0.78 [95% CI, 0.67-0.90]), abdominal aortic aneurysm (OR, 0.63 [95% CI, 0.49-0.81]), celiac disease (OR, 0.42 [95% CI, 0.28-0.61]) and interstitial lung disease (OR, 0.09 [95% CI, 0.05-0.15]). CONCLUSIONS AND RELEVANCE: It is likely that longer telomeres increase risk for several cancers but reduce risk for some non-neoplastic diseases, including cardiovascular diseases.