Wenbo Tang

Genome-Wide Association Studies Identify CHRNA5/3 and HTR4 in the Development of Airflow Obstruction

RATIONALEnGenome-wide association studies (GWAS) have identified loci influencing lung function, but fewer genes influencing chronic obstructive pulmonary disease (COPD) are known.nnnOBJECTIVESnPerform meta-analyses of GWAS for airflow obstruction, a key pathophysiologic characteristic of COPD assessed by spirometry, in population-based cohorts examining all participants, ever smokers, never smokers, asthma-free participants, and more severe cases.nnnMETHODSnFifteen cohorts were studied for discovery (3,368 affected; 29,507 unaffected), and a population-based family study and a meta-analysis of case-control studies were used for replication and regional follow-up (3,837 cases; 4,479 control subjects). Airflow obstruction was defined as FEV(1) and its ratio to FVC (FEV(1)/FVC) both less than their respective lower limits of normal as determined by published reference equations.nnnMEASUREMENTS AND MAIN RESULTSnThe discovery meta-analyses identified one region on chromosome 15q25.1 meeting genome-wide significance in ever smokers that includes AGPHD1, IREB2, and CHRNA5/CHRNA3 genes. The region was also modestly associated among never smokers. Gene expression studies confirmed the presence of CHRNA5/3 in lung, airway smooth muscle, and bronchial epithelial cells. A single-nucleotide polymorphism in HTR4, a gene previously related to FEV(1)/FVC, achieved genome-wide statistical significance in combined meta-analysis. Top single-nucleotide polymorphisms in ADAM19, RARB, PPAP2B, and ADAMTS19 were nominally replicated in the COPD meta-analysis.nnnCONCLUSIONSnThese results suggest an important role for the CHRNA5/3 region as a genetic risk factor for airflow obstruction that may be independent of smoking and implicate the HTR4 gene in the etiology of airflow obstruction.

Large-Scale Genome-Wide Association Studies and Meta-Analyses of Longitudinal Change in Adult Lung Function

Background Genome-wide association studies (GWAS) have identified numerous loci influencing cross-sectional lung function, but less is known about genes influencing longitudinal change in lung function. Methods We performed GWAS of the rate of change in forced expiratory volume in the first second (FEV1) in 14 longitudinal, population-based cohort studies comprising 27,249 adults of European ancestry using linear mixed effects model and combined cohort-specific results using fixed effect meta-analysis to identify novel genetic loci associated with longitudinal change in lung function. Gene expression analyses were subsequently performed for identified genetic loci. As a secondary aim, we estimated the mean rate of decline in FEV1 by smoking pattern, irrespective of genotypes, across these 14 studies using meta-analysis. Results The overall meta-analysis produced suggestive evidence for association at the novel IL16/STARD5/TMC3 locus on chromosome 15 (P u200a=u200a 5.71 × 10-7). In addition, meta-analysis using the five cohorts with ≥3 FEV1 measurements per participant identified the novel ME3 locus on chromosome 11 (P u200a=u200a 2.18 × 10-8) at genome-wide significance. Neither locus was associated with FEV1 decline in two additional cohort studies. We confirmed gene expression of IL16, STARD5, and ME3 in multiple lung tissues. Publicly available microarray data confirmed differential expression of all three genes in lung samples from COPD patients compared with controls. Irrespective of genotypes, the combined estimate for FEV1 decline was 26.9, 29.2 and 35.7 mL/year in never, former, and persistent smokers, respectively. Conclusions In this large-scale GWAS, we identified two novel genetic loci in association with the rate of change in FEV1 that harbor candidate genes with biologically plausible functional links to lung function.

Molecular mechanisms underlying variations in lung function: a systems genetics analysis

BACKGROUNDnLung function measures reflect the physiological state of the lung, and are essential to the diagnosis of chronic obstructive pulmonary disease (COPD). The SpiroMeta-CHARGE consortium undertook the largest genome-wide association study (GWAS) so far (n=48,201) for forced expiratory volume in 1 s (FEV1) and the ratio of FEV1 to forced vital capacity (FEV1/FVC) in the general population. The lung expression quantitative trait loci (eQTLs) study mapped the genetic architecture of gene expression in lung tissue from 1111 individuals. We used a systems genetics approach to identify single nucleotide polymorphisms (SNPs) associated with lung function that act as eQTLs and change the level of expression of their target genes in lung tissue; termed eSNPs.nnnMETHODSnThe SpiroMeta-CHARGE GWAS results were integrated with lung eQTLs to map eSNPs and the genes and pathways underlying the associations in lung tissue. For comparison, a similar analysis was done in peripheral blood. The lung mRNA expression levels of the eSNP-regulated genes were tested for associations with lung function measures in 727 individuals. Additional analyses identified the pleiotropic effects of eSNPs from the published GWAS catalogue, and mapped enrichment in regulatory regions from the ENCODE project. Finally, the Connectivity Map database was used to identify potential therapeutics in silico that could reverse the COPD lung tissue gene signature.nnnFINDINGSnSNPs associated with lung function measures were more likely to be eQTLs and vice versa. The integration mapped the specific genes underlying the GWAS signals in lung tissue. The eSNP-regulated genes were enriched for developmental and inflammatory pathways; by comparison, SNPs associated with lung function that were eQTLs in blood, but not in lung, were only involved in inflammatory pathways. Lung function eSNPs were enriched for regulatory elements and were over-represented among genes showing differential expression during fetal lung development. An mRNA gene expression signature for COPD was identified in lung tissue and compared with the Connectivity Map. This in-silico drug repurposing approach suggested several compounds that reverse the COPD gene expression signature, including a nicotine receptor antagonist. These findings represent novel therapeutic pathways for COPD.nnnINTERPRETATIONnThe system genetics approach identified lung tissue genes driving the variation in lung function and susceptibility to COPD. The identification of these genes and the pathways in which they are enriched is essential to understand the pathophysiology of airway obstruction and to identify novel therapeutic targets and biomarkers for COPD, including drugs that reverse the COPD gene signature in silico.nnnFUNDINGnThe research reported in this article was not specifically funded by any agency. See Acknowledgments for a full list of funders of the lung eQTL study and the Spiro-Meta CHARGE GWAS.

Genetic and Environmental Factors Are Associated with Serum 25-Hydroxyvitamin D Concentrations in Older African Americans

BACKGROUNDnLow circulating 25-hydroxyvitamin D [25(OH)D] is prevalent in African Americans, but predictors of vitamin D status are understudied compared to Caucasian populations.nnnOBJECTIVEnWe investigated whether certain environmental and genetic factors are predictors of circulating 25(OH)D in 989 elderly African Americans participating in the Health, Aging, and Body Composition (Health ABC) Study.nnnMETHODSnRegression analysis estimated the cross-sectional association of nongenetic (environmental) factors with 25(OH)D. Single nucleotide polymorphisms (SNPs) associated with 25(OH)D in Caucasian genome-wide association studies (GWASs) were analyzed for association with serum 25(OH)D, including analyses of all imputed SNPs in identified genomic regions. Genome-wide complex trait analysis (GCTA) evaluated the association of all (genome-wide) genotyped SNPs with serum 25(OH)D in the Health ABC Study with replication in the Multi-Ethnic Study of Atherosclerosis (MESA) cohort.nnnRESULTSnGender, study site, season of blood draw, body mass index, dietary supplement use, dairy and cereal consumption, Healthy Eating Index score, and walking >180 min/wk were associated with 25(OH)D (P < 0.05), jointly explaining 25% of the variation in circulating 25(OH)D. Multivitamin supplement use was the strongest predictor of circulating 25(OH)D, and supplement users had a 6.3-μg/L higher serum 25(OH)D concentration compared with nonusers. Previous GWAS-identified gene regions were not replicated in African Americans, but the nonsynonymous rs7041 SNP in group-specific component (vitamin D binding protein) was close to significance thresholds (P = 0.08), and there was evidence for an interaction between this SNP and use of multivitamin supplements in relation to serum 25(OH)D concentration (P = 0.04). Twenty-three percent (95% CI: 0%, 52%) of the variation in serum 25(OH)D was explained by total genetic variation in a pooled GCTA of 2087 Health ABC Study and MESA African-American participants, but population substructure effects could not be separated from other genetic influences.nnnCONCLUSIONSnModifiable dietary and lifestyle predictors of serum 25(OH)D were identified in African Americans. GCTA confirms that a proportion of 25(OH)D variability is attributable to genetic variation, but genomic regions associated with the 25(OH)D phenotype identified in prior GWASs of European Americans were not replicated in the Health ABC Study in African Americans.

Integrative Pathway Genomics of Lung Function and Airflow Obstruction

Chronic respiratory disorders are important contributors to the global burden of disease. Genome-wide association studies (GWASs) of lung function measures have identified several trait-associated loci, but explain only a modest portion of the phenotypic variability. We postulated that integrating pathway-based methods with GWASs of pulmonary function and airflow obstruction would identify a broader repertoire of genes and processes influencing these traits. We performed two independent GWASs of lung function and applied gene set enrichment analysis to one of the studies and validated the results using the second GWAS. We identified 131 significantly enriched gene sets associated with lung function and clustered them into larger biological modules involved in diverse processes including development, immunity, cell signaling, proliferation and arachidonic acid. We found that enrichment of gene sets was not driven by GWAS-significant variants or loci, but instead by those with less stringent association P-values. Next, we applied pathway enrichment analysis to a meta-analyzed GWAS of airflow obstruction. We identified several biologic modules that functionally overlapped with those associated with pulmonary function. However, differences were also noted, including enrichment of extracellular matrix (ECM) processes specifically in the airflow obstruction study. Network analysis of the ECM module implicated a candidate gene, matrix metalloproteinase 10 (MMP10), as a putative disease target. We used a knockout mouse model to functionally validate MMP10s role in influencing lungs susceptibility to cigarette smoke-induced emphysema. By integrating pathway analysis with population-based genomics, we unraveled biologic processes underlying pulmonary function traits and identified a candidate gene for obstructive lung disease.

Genetic variation in antioxidant enzymes, cigarette smoking, and longitudinal change in lung function

Antioxidant enzymes play an important role in the defense against oxidative stress in the lung and in the pathogenesis of chronic obstructive pulmonary disease (COPD). Sequence variation in genes encoding antioxidant enzymes may alter susceptibility to COPD by affecting longitudinal change in lung function in adults. We genotyped 384 sequence variants in 56 candidate genes in 1281 African American and 1794 European American elderly adults in the Health, Aging, and Body Composition study. Single-marker associations and gene-by-smoking interactions with rate of change in FEV₁ and FEV₁/FVC were evaluated using linear mixed-effects models, stratified by race/ethnicity. In European Americans, rs17883901 in GCLC was statistically significantly associated with rate of change in FEV₁/FVC; the recessive genotype (TT) was associated with a 0.9% per year steeper decline (P = 4.50 × 10(-5)). Statistically significant gene-by-smoking interactions were observed for variants in two genes in European Americans: the minor allele of rs2297765 in mGST3 attenuated the accelerated decline in FEV₁/FVC in smokers by 0.45% per year (P = 1.13 × 10(-4)); for participants with greater baseline smoking pack-years, the minor allele of rs2073192 in IDH3B was associated with an accelerated decline in FEV₁/FVC (P = 2.10 × 10(-4)). For both genes, nominally significant interactions (P < 0.01) were observed at the gene level in African Americans (P = 0.007 and 4.60 × 10(-4), respectively). Nominally significant evidence of association was observed for variants in SOD3 and GLRX2 in multiple analyses. This study identifies two novel genes associated with longitudinal lung function phenotypes in both African and European Americans and confirms a prior finding for GCLC. These findings suggest novel mechanisms and molecular targets for future research and advance the understanding of genetic determinants of lung function and COPD risk.

Association of 25-Hydroxyvitamin D status and genetic variation in the vitamin D metabolic pathway with FEV1 in the Framingham Heart Study

BackgroundVitamin D is associated with lung function in cross-sectional studies, and vitamin D inadequacy is hypothesized to play a role in the pathogenesis of chronic obstructive pulmonary disease. Further data are needed to clarify the relation between vitamin D status, genetic variation in vitamin D metabolic genes, and cross-sectional and longitudinal changes in lung function in healthy adults.MethodsWe estimated the association between serum 25-hydroxyvitamin D [25(OH)D] and cross-sectional forced expiratory volume in the first second (FEV1) in Framingham Heart Study (FHS) Offspring and Third Generation participants and the association between serum 25(OH)D and longitudinal change in FEV1 in Third Generation participants using linear mixed-effects models. Using a gene-based approach, we investigated the association between 241 SNPs in 6 select vitamin D metabolic genes in relation to longitudinal change in FEV1 in Offspring participants and pursued replication of these findings in a meta-analyzed set of 4 independent cohorts.ResultsWe found a positive cross-sectional association between 25(OH)D and FEV1 in FHS Offspring and Third Generation participants (Pu2009=u20090.004). There was little or no association between 25(OH)D and longitudinal change in FEV1 in Third Generation participants (Pu2009=u20090.97). In Offspring participants, the CYP2R1 gene, hypothesized to influence usual serum 25(OH)D status, was associated with longitudinal change in FEV1 (gene-based Pu2009<u20090.05). The most significantly associated SNP from CYP2R1 had a consistent direction of association with FEV1 in the meta-analyzed set of replication cohorts, but the association did not reach statistical significance thresholds (Pu2009=u20090.09).ConclusionsSerum 25(OH)D status was associated with cross-sectional FEV1, but not longitudinal change in FEV1. The inconsistent associations may be driven by differences in the groups studied. CYP2R1 demonstrated a gene-based association with longitudinal change in FEV1 and is a promising candidate gene for further studies.

Polymorphisms in Serine Hydroxymethyltransferase 1 and Methylenetetrahydrofolate Reductase Interact to Increase Cardiovascular Disease Risk in Humans

The enzymes serine hydroxymethyltransferase 1 (gene name SHMT1) and methylenetetrahydrofolate reductase (gene name MTHFR) regulate key reactions in folate-mediated one-carbon metabolism. Common genetic variants with the potential to influence disease risk exist in both genes. A prior report from the Normative Aging Study indicated no association of the SHMT1 rs1979277 SNP with cardiovascular disease (CVD), but a strong gene-gene interaction was detected with MTHFR rs1801133. We investigated the effect of the SHMT1 rs1979277 SNP and the SHMT1 rs1979277-MTHFR rs1801133 interaction in 2 epidemiologic cohort studies. In the Nurses Health Study (NHS), the MTHFR rs1801133 variant genotypes were associated with an increased CVD risk and there was an interaction between SHMT1 and MTHFR such that the association of the MTHFR rs1801133 CT genotype (vs. CC; the TT genotype could not be evaluated) was stronger in the presence of the SHMT1 rs1979277 TT genotype (OR = 4.34, 95% CI = 1.2, 16.2; P = 0.049). In the Health Professionals Follow-Up Study, the MTHFR rs1801133 genotype was not associated with CVD risk, nor was there an interaction with SHMT1 rs1979277. The association of genetic variation in the SHMT1 gene, alone and in interaction with MTHFR, in relation to CVD risk is relatively understudied at the population level and results in the NHS confirmed a past report of gene-gene interaction, which is consistent with mechanisms suggested by basic science studies.

Evidence for large-scale gene-by-smoking interaction effects on pulmonary function

Abstract Background: Smoking is the strongest environmental risk factor for reduced pulmonary function. The genetic component of various pulmonary traits has also been demonstrated, and at least 26 loci have been reproducibly associated with either FEV1 (forced expiratory volume in 1 second) or FEV1/FVC (FEV1/forced vital capacity). Although the main effects of smoking and genetic loci are well established, the question of potential gene-by-smoking interaction effect remains unanswered. The aim of the present study was to assess, using a genetic risk score approach, whether the effect of these 26 loci on pulmonary function is influenced by smoking. Methods: We evaluated the interaction between smoking exposure, considered as either ever vs never or pack-years, and a 26-single nucleotide polymorphisms (SNPs) genetic risk score in relation to FEV1 or FEV1/FVC in 50u2009047 participants of European ancestry from the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) and SpiroMeta consortia. Results: We identified an interaction (βintu2009=u2009–0.036, 95% confidence interval, –0.040 to –0.032, Pu2009=u20090.00057) between an unweighted 26 SNP genetic risk score and smoking status (ever/never) on the FEV1/FVC ratio. In interpreting this interaction, we showed that the genetic risk of falling below the FEV1/FVC threshold used to diagnose chronic obstructive pulmonary disease is higher among ever smokers than among never smokers. A replication analysis in two independent datasets, although not statistically significant, showed a similar trend in the interaction effect. Conclusions: This study highlights the benefit of using genetic risk scores for identifying interactions missed when studying individual SNPs and shows, for the first time, that persons with the highest genetic risk for low FEV1/FVC may be more susceptible to the deleterious effects of smoking.

Multiethnic Meta-analysis Identifies New Loci for Pulmonary Function

Nearly 100 loci have been identified for pulmonary function, almost exclusively in studies of European ancestry populations. We extend previous research by meta-analyzing genome-wide association studies of 1000 Genomes imputed variants in relation to pulmonary function in a multiethnic population of 90,715 individuals of European (N=60,552), African (N=8,429), Asian (N=9,959), and Hispanic/Latino (N=11,775) ethnicities. We identified over 50 novel loci at genome-wide significance in ancestry-specific and/or multiethnic meta-analyses. Recent fine mapping methods incorporating functional annotation, gene expression, and/or differences in linkage disequilibrium between ethnicities identified potential causal variants and genes at known and newly identified loci. Sixteen of the novel genes encode proteins with predicted or established drug targets, including KCNK2 and CDK12.