Amy R. Bentley

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

RATIONALE Genome-wide association studies (GWAS) have identified loci influencing lung function, but fewer genes influencing chronic obstructive pulmonary disease (COPD) are known. OBJECTIVES Perform 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. METHODS Fifteen 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. MEASUREMENTS AND MAIN RESULTS The 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. CONCLUSIONS These 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.

Genetic variation and gene expression in antioxidant related enzymes and risk of COPD: a systematic review

Background: Observational epidemiological studies of dietary antioxidant intake, serum antioxidant concentration and lung outcomes suggest that lower levels of antioxidant defences are associated with decreased lung function. Another approach to understanding the role of oxidant/antioxidant imbalance in the risk of chronic obstructive pulmonary disease (COPD) is to investigate the role of genetic variation in antioxidant enzymes, and indeed family based studies suggest a heritable component to lung disease. Many studies of the genes encoding antioxidant enzymes have considered COPD or COPD related outcomes, and a systematic review is needed to summarise the evidence to date, and to provide insights for further research. Methods: Genetic association studies of antioxidant enzymes and COPD/COPD related traits, and comparative gene expression studies with disease or smoking as the exposure were systematically identified and reviewed. Antioxidant enzymes considered included enzymes involved in glutathione metabolism, in the thioredoxin system, superoxide dismutases (SOD) and catalase. Results: A total of 29 genetic association and 15 comparative gene expression studies met the inclusion criteria. The strongest and most consistent effects were in the genes GCL, GSTM1, GSTP1 and SOD3. This review also highlights the lack of studies for genes of interest, particularly GSR, GGT and those related to TXN. There were limited opportunities to evaluate the contribution of a gene to disease risk through synthesis of results from different study designs, as the majority of studies considered either association of sequence variants with disease or effect of disease on gene expression. Conclusion: Network driven approaches that consider potential interaction between and among genes, smoke exposure and antioxidant intake are needed to fully characterise the role of oxidant/antioxidant balance in pathogenesis.

UGT1A1 is a major locus influencing bilirubin levels in African Americans

Total serum bilirubin is associated with several clinical outcomes, including cardiovascular disease, diabetes and drug metabolism. We conducted a genome-wide association study in 619 healthy unrelated African Americans in an attempt to replicate reported findings in Europeans and Asians and to identify novel loci influencing total serum bilirubin levels. We analyzed a dense panel of over two million genotyped and imputed SNPs in additive genetic models adjusting for age, sex, and the first two significant principal components from the sample covariance matrix of genotypes. Thirty-nine SNPs spanning a 78 kb region within the UGT1A1 displayed P-values <5 × 10−8. The lowest P-value was 1.7 × 10−22 for SNP rs887829. None of SNPs in the UGT1A1 remained statistically significant in conditional association analyses that adjusted for rs887829. In addition, SNP rs10929302 located in phenobarbital response enhancer module was significantly associated with bilirubin level with a P-value of 1.37 × 10−11; this enhancer module is believed to have a critical role in phenobarbital treatment of hyperbilirubinemia. Interestingly, the lead SNP, rs887829, is in strong linkage disequilibrium (LD) (r2≥0.74) with rs10929302. Taking advantage of the lower LD and shorter haplotypes in African-ancestry populations, we identified rs887829 as a more refined proxy for the causative variant influencing bilirubin levels. Also, we replicated the reported association between variants in SEMA3C and bilirubin levels. In summary, UGT1A1 is a major locus influencing bilirubin levels and the results of this study promise to contribute to understanding of the etiology and treatment of hyperbilirubinaemia in African-ancestry populations.

Dietary antioxidants and forced expiratory volume in 1 s decline: the Health, Aging and Body Composition study

Increased antioxidant defences are hypothesised to decrease age- and smoking-related decline in lung function. The relationship between dietary antioxidants, smoking and forced expiratory volume in 1 s (FEV1) was investigated in community-dwelling older adults in the Health, Aging and Body Composition study. 1,443 participants completed a food frequency questionnaire, self-reported smoking history and had measurements taken of FEV1 at both baseline and after 4 yrs of follow-up. The association of dietary intake of nutrients and foods with antioxidant properties and rate of FEV1 decline was investigated using hierarchical linear regression models. In continuing smokers (current smokers at both time-points), higher vitamin C intake and higher intake of fruit and vegetables were associated with an 18 and 24 mL·yr−1 slower rate of FEV1 decline compared with a lower intake (p<0.0001 and p=0.003, respectively). In quitters (a current smoker at study baseline who had quit during follow-up), higher intake was associated with an attenuated rate of decline for each nutrient studied (p≤0.003 for all models). In nonsmoking participants, there was little or no association of diet and rate of decline in FEV1. The intake of nutrients with antioxidant properties may modulate lung function decline in older adults exposed to cigarette smoke.

Variation in APOL1 Contributes to Ancestry-Level Differences in HDLc-Kidney Function Association

Low levels of high-density cholesterol (HDLc) accompany chronic kidney disease, but the association between HDLc and the estimated glomerular filtration rate (eGFR) in the general population is unclear. We investigated the HDLc-eGFR association in nondiabetic Han Chinese (HC, n = 1100), West Africans (WA, n = 1497), and African Americans (AA, n = 1539). There were significant differences by ancestry: HDLc was positively associated with eGFR in HC (β = 0.13, P < 0.0001), but negatively associated among African ancestry populations (WA: −0.19, P < 0.0001; AA: −0.09, P = 0.02). These differences were also seen in nationally-representative NHANES data (among European Americans: 0.09, P = 0.005; among African Americans −0.14, P = 0.03). To further explore the findings in African ancestry populations, we investigated the role of an African ancestry-specific nephropathy risk variant, rs73885319, in the gene encoding HDL-associated APOL1. Among AA, an inverse HDLc-eGFR association was observed only with the risk genotype (−0.38 versus 0.001; P = 0.03). This interaction was not seen in WA. In summary, counter to expectation, an inverse HDLc-eGFR association was observed among those of African ancestry. Given the APOL1 × HDLc interaction among AA, genetic factors may contribute to this paradoxical association. Notably, these findings suggest that the unexplained mechanism by which APOL1 affects kidney-disease risk may involve HDLc.

Transferability and Fine Mapping of genome-wide associated loci for lipids in African Americans

BackgroundA recent, large genome-wide association study (GWAS) of European ancestry individuals has identified multiple genetic variants influencing serum lipids. Studies of the transferability of these associations to African Americans remain few, an important limitation given interethnic differences in serum lipids and the disproportionate burden of lipid-associated metabolic diseases among African Americans.MethodsWe attempted to evaluate the transferability of 95 lipid-associated loci recently identified in European ancestry individuals to 887 non-diabetic, unrelated African Americans from a population-based sample in the Washington, DC area. Additionally, we took advantage of the generally reduced linkage disequilibrium among African ancestry populations in comparison to European ancestry populations to fine-map replicated GWAS signals.ResultsWe successfully replicated reported associations for 10 loci (CILP2/SF4, STARD3, LPL, CYP7A1, DOCK7/ANGPTL3, APOE, SORT1, IRS1, CETP, and UBASH3B). Through trans-ethnic fine-mapping, we were able to reduce associated regions around 75% of the loci that replicated.ConclusionsBetween this study and previous work in African Americans, 40 of the 95 loci reported in a large GWAS of European ancestry individuals also influence lipid levels in African Americans. While there is now evidence that the lipid-influencing role of a number of genetic variants is observed in both European and African ancestry populations, the still considerable lack of concordance highlights the importance of continued ancestry-specific studies to elucidate the genetic underpinnings of these traits.

Genome-wide association study identifies African-ancestry specific variants for metabolic syndrome.

The metabolic syndrome (MetS) is a constellation of metabolic disorders that increase the risk of developing several diseases including type 2 diabetes and cardiovascular diseases. Although genome-wide association studies (GWAS) have successfully identified variants associated with individual traits comprising MetS, the genetic basis and pathophysiological mechanisms underlying the clustering of these traits remain unclear. We conducted GWAS of MetS in 1427 Africans from Ghana and Nigeria followed by replication testing and meta-analysis in another continental African sample from Kenya. Further replication testing was performed in an African American sample from the Atherosclerosis Risk in Communities (ARIC) study. We found two African-ancestry specific variants that were significantly associated with MetS: SNP rs73989312[A] near CA10 that conferred increased risk (P=3.86 × 10(-8), OR=6.80) and SNP rs77244975[C] in CTNNA3 that conferred protection against MetS (P=1.63 × 10(-8), OR=0.15). Given the exclusive expression of CA10 in the brain, our CA10 finding strengthens previously reported link between brain function and MetS. We also identified two variants that are not African specific: rs76822696[A] near RALYL associated with increased MetS risk (P=7.37 × 10(-9), OR=1.59) and rs7964157[T] near KSR2 associated with reduced MetS risk (P=4.52 × 10(-8), Pmeta=7.82 × 10(-9), OR=0.53). The KSR2 locus displayed pleiotropic associations with triglyceride and measures of blood pressure. Rare KSR2 mutations have been reported to be associated with early onset obesity and insulin resistance. Finally, we replicated the LPL and CETP loci previously found to be associated with MetS in Europeans. These findings provide novel insights into the genetics of MetS in Africans and demonstrate the utility of conducting trans-ethnic disease gene mapping studies for testing the cosmopolitan significance of GWAS signals of cardio-metabolic traits.

Gene-Based Sequencing Identifies Lipid-Influencing Variants with Ethnicity-Specific Effects in African Americans

Although a considerable proportion of serum lipids loci identified in European ancestry individuals (EA) replicate in African Americans (AA), interethnic differences in the distribution of serum lipids suggest that some genetic determinants differ by ethnicity. We conducted a comprehensive evaluation of five lipid candidate genes to identify variants with ethnicity-specific effects. We sequenced ABCA1, LCAT, LPL, PON1, and SERPINE1 in 48 AA individuals with extreme serum lipid concentrations (high HDLC/low TG or low HDLC/high TG). Identified variants were genotyped in the full population-based sample of AA (n = 1694) and tested for an association with serum lipids. rs328 (LPL) and correlated variants were associated with higher HDLC and lower TG. Interestingly, a stronger effect was observed on a “European” vs. “African” genetic background at this locus. To investigate this effect, we evaluated the region among West Africans (WA). For TG, the effect size among WA was the same in AA with only African local ancestry (2–3% lower TG), while the larger association among AA with local European ancestry matched previous reports in EA (10%). For HDLC, there was no association with rs328 in AA with only African local ancestry or in WA, while the association among AA with European local ancestry was much greater than what has been observed for EA (15 vs. ∼5 mg/dl), suggesting an interaction with an environmental or genetic factor that differs by ethnicity. Beyond this ancestry effect, the importance of African ancestry-focused, sequence-based work was also highlighted by serum lipid associations of variants that were in higher frequency (or present only) among those of African ancestry. By beginning our study with the sequence variation present in AA individuals, investigating local ancestry effects, and seeking replication in WA, we were able to comprehensively evaluate the role of a set of candidate genes in serum lipids in AA.

Evaluation of Genome Wide Association Study Associated Type 2 Diabetes Susceptibility Loci in Sub Saharan Africans.

Genome wide association studies (GWAS) for type 2 diabetes (T2D) undertaken in European and Asian ancestry populations have yielded dozens of robustly associated loci. However, the genomics of T2D remains largely understudied in sub-Saharan Africa (SSA), where rates of T2D are increasing dramatically and where the environmental background is quite different than in these previous studies. Here, we evaluate 106 reported T2D GWAS loci in continental Africans. We tested each of these SNPs, and SNPs in linkage disequilibrium (LD) with these index SNPs, for an association with T2D in order to assess transferability and to fine map the loci leveraging the generally reduced LD of African genomes. The study included 1775 unrelated Africans (1035 T2D cases, 740 controls; mean age 54 years; 59% female) enrolled in Nigeria, Ghana, and Kenya as part of the Africa America Diabetes Mellitus (AADM) study. All samples were genotyped on the Affymetrix Axiom PanAFR SNP array. Forty-one of the tested loci showed transferability to this African sample (p < 0.05, same direction of effect), 11 at the exact reported SNP and 30 others at SNPs in LD with the reported SNP (after adjustment for the number of tested SNPs). TCF7L2 SNP rs7903146 was the most significant locus in this study (p = 1.61 × 10−8). Most of the loci that showed transferability were successfully fine-mapped, i.e., localized to smaller haplotypes than in the original reports. The findings indicate that the genetic architecture of T2D in SSA is characterized by several risk loci shared with non-African ancestral populations and that data from African populations may facilitate fine mapping of risk loci. The study provides an important resource for meta-analysis of African ancestry populations and transferability of novel loci.

Genetic variation in antioxidant enzymes and lung function.

Not all cigarette smokers develop chronic obstructive pulmonary disease, and discovering susceptibility factors is an important research priority. The oxidative burden of smoking may overwhelm antioxidant defenses, and vulnerabilities may exist as a result of sequence variants in genes encoding antioxidant enzymes. This study explored the association between genetic variation in a network of antioxidant enzymes and lung phenotypes. Linear models evaluated single-locus marker associations in 2387 European American and African American participants in the Health, Aging, and Body Composition Study. After corrections were made for multiple comparisons, 15 statistically significant associations were identified, all of which were for SNP by smoking interactions. The most statistically significant findings were for genes encoding members of the isocitrate dehydrogenase gene family (IDH3A, IDH3B, IDH2). For rs6107100 (IDH3B) the variant genotype was associated with a difference of 6% in the FEV(1)/FVC ratio in African American current smokers, but the SNP had little or no association with FEV(1)/FVC in former and never smokers (nominal p(interaction)=5×10(-6)). A variant of the peroxiredoxin gene (rs9787810, PRDX5) was associated with lower percentage predicted FEV(1) and a lower ratio in European American current smokers, with little or no association in other smoking groups (nominal p(interaction)=0.0001 and 0.0003, respectively). The studied genes have not been reported in previous candidate gene association studies, and thus the findings suggest novel mechanisms and targets for future research and provide evidence for a contribution of sequence variation in genes encoding antioxidant enzymes to susceptibility in smokers.