Clinical and molecular implications of RGS2 promoter genetic variation in severe asthma.

BACKGROUND: Regulator of G protein signaling (RGS) 2 terminates bronchoconstrictive Gαq signaling; murine RGS2 knockout demonstrate airway hyperresponsiveness. While RGS2 promoter variants rs2746071 and rs2746072 associate with a clinical mild asthma phenotype, their impact on human airway smooth muscle (HASM) contractility and asthma severity outcomes is unknown. OBJECTIVE: We sought to determine whether reductions in RGS2 expression seen with these 2 RGS2 promoter variants augment HASM contractility and associate with an asthma severity phenotype. METHODS: We transfected HASM with a range of RGS2-specific small interfering RNA (siRNA) concentrations and determined RGS2 protein expression by Western blot analysis and intracellular calcium flux induced by histamine (a Gαq-coupled H1 receptor bronchoconstrictive agonist). We conducted regression-based genotype association analyses of RGS2 variants from 611 patients from the National Heart, Lung, and Blood Institute Severe Asthma Research Program 3. RESULTS: RGS2-specific siRNA caused dose-dependent increases in histamine-stimulated bronchoconstrictive intracellular calcium signaling (2-way ANOVA, P < .0001) with a concomitant decrease in RGS2 protein expression. RGS2-specific siRNA did not affect Gαq-independent ionomycin-induced intracellular calcium signaling (P = .42). The minor allele frequency of rs2746071 and rs2746072 was 0.46 and 0.28 among African American/non-Hispanic Black patients and was 0.28 and 0.27 among non-Hispanic White patients, among whom these single nucleotide polymorphisms were in stronger linkage disequilibrium (r2 = 0.97). Among non-Hispanic White patients, risk allele homozygotes for rs2746072 and rs2746071 each had nearly 2-fold greater asthma exacerbation rates relative to alternative genotypes with wild-type alleles (Padditive = 2.86 × 10-5/Precessive = 5.22 × 10-6 and Padditive = 3.46 × 10-6/Precessive = 6.74 × 10-7, respectively) at baseline, which was confirmed by prospective longitudinal exacerbation data. CONCLUSION: RGS2 promoter variation associates with a molecular and clinical phenotype characterized by enhanced bronchoconstrictive stimulation in vitro and higher asthma exacerbations rates in non-Hispanic White patients.

Multi-ancestry genome-wide association study of asthma exacerbations.

BACKGROUND: Asthma exacerbations are a serious public health concern due to high healthcare resource utilization, work/school productivity loss, impact on quality of life, and risk of mortality. The genetic basis of asthma exacerbations has been studied in several populations, but no prior study has performed a multi-ancestry meta-analysis of genome-wide association studies (meta-GWAS) for this trait. We aimed to identify common genetic loci associated with asthma exacerbations across diverse populations and to assess their functional role in regulating DNA methylation and gene expression. METHODS: A meta-GWAS of asthma exacerbations in 4989 Europeans, 2181 Hispanics/Latinos, 1250 Singaporean Chinese, and 972 African Americans analyzed 9.6 million genetic variants. Suggestively associated variants (p ≤ 5 × 10-5 ) were assessed for replication in 36,477 European and 1078 non-European asthma patients. Functional effects on DNA methylation were assessed in 595 Hispanic/Latino and African American asthma patients and in publicly available databases. The effect on gene expression was evaluated in silico. RESULTS: One hundred and twenty-six independent variants were suggestively associated with asthma exacerbations in the discovery phase. Two variants independently replicated: rs12091010 located at vascular cell adhesion molecule-1/exostosin like glycosyltransferase-2 (VCAM1/EXTL2) (discovery: odds ratio (ORT allele ) = 0.82, p = 9.05 × 10-6 and replication: ORT allele  = 0.89, p = 5.35 × 10-3 ) and rs943126 from pantothenate kinase 1 (PANK1) (discovery: ORC allele  = 0.85, p = 3.10 × 10-5 and replication: ORC allele  = 0.89, p = 1.30 × 10-2 ). Both variants regulate gene expression of genes where they locate and DNA methylation levels of nearby genes in whole blood. CONCLUSIONS: This multi-ancestry study revealed novel suggestive regulatory loci for asthma exacerbations located in genomic regions participating in inflammation and host defense.

Genome-wide association study of asthma, total IgE, and lung function in a cohort of Peruvian children.

BACKGROUND: Genetic ancestry plays a role in asthma health disparities. OBJECTIVE: Our aim was to evaluate the impact of ancestry on and identify genetic variants associated with asthma, total serum IgE level, and lung function. METHODS: A total of 436 Peruvian children (aged 9-19 years) with asthma and 291 without asthma were genotyped by using the Illumina Multi-Ethnic Global Array. Genome-wide proportions of indigenous ancestry populations from continental America (NAT) and European ancestry from the Iberian populations in Spain (IBS) were estimated by using ADMIXTURE. We assessed the relationship between ancestry and the phenotypes and performed a genome-wide association study. RESULTS: The mean ancestry proportions were 84.7% NAT (case patients, 84.2%; controls, 85.4%) and 15.3% IBS (15.8%; 14.6%). With adjustment for asthma, NAT was associated with higher total serum IgE levels (P < .001) and IBS was associated with lower total serum IgE levels (P < .001). NAT was associated with higher FEV1 percent predicted values (P < .001), whereas IBS was associated with lower FEV1 values in the controls but not in the case patients. The HLA-DR/DQ region on chromosome 6 (Chr6) was strongly associated with total serum IgE (rs3135348; P = 3.438 × 10-10) and was independent of an association with the haplotype HLA-DQA1∼HLA-DQB1:04.01∼04.02 (P = 1.55 × 10-05). For lung function, we identified a locus (rs4410198; P = 5.536 × 10-11) mapping to Chr19, near a cluster of zinc finger interacting genes that colocalizes to the long noncoding RNA CTD-2537I9.5. This novel locus was replicated in an independent sample of pediatric case patients with asthma with similar admixture from Brazil (P = .005). CONCLUSION: This study confirms the role of HLA in atopy, and identifies a novel locus mapping to a long noncoding RNA for lung function that may be specific to children with NAT.

The Effects of Rare SERPINA1 Variants on Lung Function and Emphysema in SPIROMICS.

Rationale: The role of PI (protease inhibitor) type Z heterozygotes and additional rare variant genotypes in the gene encoding alpha-1 antitrypsin, SERPINA1 (serpin peptidase inhibitor, clade A, member 1), in determining chronic obstructive pulmonary disease risk and severity is controversial.Objectives: To comprehensively evaluate the effects of rare SERPINA1 variants on lung function and emphysema phenotypes in subjects with significant tobacco smoke exposure using deep gene resequencing and alpha-1 antitrypsin concentrations.Methods: DNA samples from 1,693 non-Hispanic white individuals, 385 African Americans, and 90 Hispanics with ≥20 pack-years smoking were resequenced for the identification of rare variants (allele frequency < 0.05) in 16.9 kB of SERPINA1.Measurements and Main Results: White PI Z heterozygotes confirmed by sequencing (MZ; n = 74) had lower post-bronchodilator FEV1 (P = 0.007), FEV1/FVC (P = 0.003), and greater computed tomography-based emphysema (P = 0.02) compared with 1,411 white individuals without PI Z, S, or additional rare variants denoted as VR. PI Z-containing compound heterozygotes (ZS/ZVR; n = 7) had lower FEV1/FVC (P = 0.02) and forced expiratory flow, midexpiratory phase (P = 0.009). Nineteen white heterozygotes for five non-S/Z coding variants associated with lower alpha-1 antitrypsin had greater computed tomography-based emphysema compared with those without rare variants. In African Americans, a 5' untranslated region insertion (rs568223361) was associated with lower alpha-1 antitrypsin and functional small airway disease (P = 0.007).Conclusions: In this integrative deep sequencing study of SERPINA1 with alpha-1 antitrypsin concentrations in a heavy smoker and chronic obstructive pulmonary disease cohort, we confirmed the effects of PI Z heterozygote and compound heterozygote genotypes. We demonstrate the cumulative effects of multiple SERPINA1 variants on alpha-1 antitrypsin deficiency, lung function, and emphysema, thus significantly increasing the frequency of SERPINA1 variation associated with respiratory disease in at-risk smokers.

Comparison of Proteomic Assessment Methods in Multiple Cohort Studies.

Novel proteomics platforms, such as the aptamer-based SOMAscan platform, can quantify large numbers of proteins efficiently and cost-effectively and are rapidly growing in popularity. However, comparisons to conventional immunoassays remain underexplored, leaving investigators unsure when cross-assay comparisons are appropriate. The correlation of results from immunoassays with relative protein quantification is explored by SOMAscan. For 63 proteins assessed in two chronic obstructive pulmonary disease (COPD) cohorts, subpopulations and intermediate outcome measures in COPD Study (SPIROMICS), and COPDGene, using myriad rules based medicine multiplex immunoassays and SOMAscan, Spearman correlation coefficients range from -0.13 to 0.97, with a median correlation coefficient of ≈0.5 and consistent results across cohorts. A similar range is observed for immunoassays in the population-based Multi-Ethnic Study of Atherosclerosis and for other assays in COPDGene and SPIROMICS. Comparisons of relative quantification from the antibody-based Olink platform and SOMAscan in a small cohort of myocardial infarction patients also show a wide correlation range. Finally, cis pQTL data, mass spectrometry aptamer confirmation, and other publicly available data are integrated to assess relationships with observed correlations. Correlation between proteomics assays shows a wide range and should be carefully considered when comparing and meta-analyzing proteomics data across assays and studies.

Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) and receptors in type 1, type 2 and type 17 inflammation in cross-sectional asthma study.

Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) reportedly promotes, or conversely, resolves inflammation in asthma. In this study of TRAIL and cell receptors in sputum, bronchoalveolar lavage and biopsy from subjects in the Severe Asthma Research Program at Wake Forest, the high TRAIL group had significant increases in all leucocytes, and was associated with increased type 1, type 2 and type 17 cytokines, but not type 9 interleukin 9. Two variants at loci in the TRAIL gene were associated with higher sputum levels of TRAIL. Increased TRAIL decoy receptor R3/DcR1 was observed on sputum leucocytes compared with death receptor R1/DR4, suggesting reduced apoptosis and prolonged cellular inflammation.

Overlap of Genetic Risk between Interstitial Lung Abnormalities and Idiopathic Pulmonary Fibrosis.

Rationale: Interstitial lung abnormalities (ILAs) are associated with the highest genetic risk locus for idiopathic pulmonary fibrosis (IPF); however, the extent to which there are unique associations among individuals with ILAs or additional overlap with IPF is not known.Objectives: To perform a genome-wide association study (GWAS) of ILAs.Methods: ILAs and a subpleural-predominant subtype were assessed on chest computed tomography (CT) scans in the AGES (Age Gene/Environment Susceptibility), COPDGene (Genetic Epidemiology of Chronic Obstructive Pulmonary Disease [COPD]), Framingham Heart, ECLIPSE (Evaluation of COPD Longitudinally to Identify Predictive Surrogate End-points), MESA (Multi-Ethnic Study of Atherosclerosis), and SPIROMICS (Subpopulations and Intermediate Outcome Measures in COPD Study) studies. We performed a GWAS of ILAs in each cohort and combined the results using a meta-analysis. We assessed for overlapping associations in independent GWASs of IPF.Measurements and Main Results: Genome-wide genotyping data were available for 1,699 individuals with ILAs and 10,274 control subjects. The MUC5B (mucin 5B) promoter variant rs35705950 was significantly associated with both ILAs (P = 2.6 × 10-27) and subpleural ILAs (P = 1.6 × 10-29). We discovered novel genome-wide associations near IPO11 (rs6886640, P = 3.8 × 10-8) and FCF1P3 (rs73199442, P = 4.8 × 10-8) with ILAs, and near HTRE1 (rs7744971, P = 4.2 × 10-8) with subpleural-predominant ILAs. These novel associations were not associated with IPF. Among 12 previously reported IPF GWAS loci, five (DPP9, DSP, FAM13A, IVD, and MUC5B) were significantly associated (P < 0.05/12) with ILAs.Conclusions: In a GWAS of ILAs in six studies, we confirmed the association with a MUC5B promoter variant and found strong evidence for an effect of previously described IPF loci; however, novel ILA associations were not associated with IPF. These findings highlight common genetically driven biologic pathways between ILAs and IPF, and also suggest distinct ones.

Exacerbation-prone asthma in the context of race and ancestry in Asthma Clinical Research Network trials.

BACKGROUND: Minority groups of African descent experience disproportionately greater asthma morbidity compared with other racial groups, suggesting that genetic variation from a common ancestry could influence exacerbation risk. OBJECTIVE: We evaluated clinical trial measures in the context of self-reported race and genetic ancestry to identify risk factors for asthma exacerbations. METHODS: One thousand eight hundred forty multiethnic subjects from 12 Asthma Clinical Research Network and AsthmaNet trials were analyzed for incident asthma exacerbations with Poisson regression models that included clinical measures, self-reported race (black, non-Hispanic white, and other), and estimates of global genetic African ancestry in a subgroup (n = 760). RESULTS: Twenty-four percent of 1840 subjects self-identified as black. Black and white subjects had common risk factors for exacerbations, including a history of 2 or more exacerbations in the previous year and FEV1 percent predicted values, whereas chronic sinusitis, allergic rhinitis, and gastroesophageal reflux disease were only associated with increased exacerbation risk in black subjects. In the combined multiethnic cohort, neither race (P = .30) nor percentage of genetic African ancestry as a continuous variable associated with exacerbation risk (adjusted rate ratio [RR], 1.26 [95% CI, 0.94-1.70; P = .13]; RR per 1-SD change [32% ancestry], 0.97 [95% CI, 0.78-1.19; P = .74]). However, in 161 black subjects with genetic data, those with African ancestry greater than the median (≥82%) had a significantly greater risk of exacerbation (RR, 3.06 [95% CI, 1.09-8.6; P = .03]). CONCLUSION: Black subjects have unique risk factors for asthma exacerbations, of which global African genetic ancestry had the strongest effect.

A genome-wide association and admixture mapping study of bronchodilator drug response in African Americans with asthma.

Short-acting ß2-adrenergic receptor agonists (SABAs) are the most commonly prescribed asthma medications worldwide. Response to SABAs is measured as bronchodilator drug response (BDR), which varies among racial/ethnic groups in the United States. However, the genetic variation that contributes to BDR is largely undefined in African Americans with asthma. To identify genetic variants that may contribute to differences in BDR in African Americans with asthma, we performed a genome-wide association study (GWAS) of BDR in 949 African-American children with asthma, genotyped with the Axiom World Array 4 (Affymetrix, Santa Clara, CA) followed by imputation using 1000 Genomes phase III genotypes. We used linear regression models adjusting for age, sex, body mass index (BMI) and genetic ancestry to test for an association between BDR and genotype at single-nucleotide polymorphisms (SNPs). To increase power and distinguish between shared vs. population-specific associations with BDR in children with asthma, we performed a meta-analysis across 949 African Americans and 1830 Latinos (total = 2779). Finally, we performed genome-wide admixture mapping to identify regions whereby local African or European ancestry is associated with BDR in African Americans. We identified a population-specific association with an intergenic SNP on chromosome 9q21 that was significantly associated with BDR (rs73650726, p = 7.69 × 10-9). A trans-ethnic meta-analysis across African Americans and Latinos identified three additional SNPs within the intron of PRKG1 that were significantly associated with BDR (rs7903366, rs7070958 and rs7081864, p ≤ 5 × 10-8). Our results failed to replicate in three additional populations of 416 Latinos and 1615 African Americans. Our findings indicate that both population-specific and shared genetic variation contributes to differences in BDR in minority children with asthma, and that the genetic underpinnings of BDR may differ between racial/ethnic groups.

Genome-wide association study of lung function and clinical implication in heavy smokers.

BACKGROUND: The aim of this study is to identify genetic loci associated with post-bronchodilator FEV1/FVC and FEV1, and develop a multi-gene predictive model for lung function in COPD. METHODS: Genome-wide association study (GWAS) of post-bronchodilator FEV1/FVC and FEV1 was performed in 1645 non-Hispanic White European descent smokers. RESULTS: A functional rare variant in SERPINA1 (rs28929474: Glu342Lys) was significantly associated with post-bronchodilator FEV1/FVC (p = 1.2 × 10- 8) and FEV1 (p = 2.1 × 10- 9). In addition, this variant was associated with COPD (OR = 2.3; p = 7.8 × 10- 4) and severity (OR = 4.1; p = 0.0036). Heterozygous subjects (CT genotype) had significantly lower lung function and higher percentage of COPD and more severe COPD than subjects with the CC genotype. 8.6% of the variance of post-bronchodilator FEV1/FVC can be explained by SNPs in 10 genes with age, sex, and pack-years of cigarette smoking (P <  2.2 × 10- 16). CONCLUSIONS: This study is the first to show genome-wide significant association of rs28929474 in SERPINA1 with lung function. Of clinical importance, heterozygotes of rs28929474 (4.7% of subjects) have significantly reduced pulmonary function, demonstrating a major impact in smokers. The multi-gene model is significantly associated with CT-based emphysema and clinical outcome measures of severity. Combining genetic information with demographic and environmental factors will further increase the predictive power for assessing reduced lung function and COPD severity.

Expression of asthma susceptibility genes in bronchial epithelial cells and bronchial alveolar lavage in the Severe Asthma Research Program (SARP) cohort.

OBJECTIVE: Genome-wide association studies (GWASs) have identified genes associated with asthma, however expression of these genes in asthma-relevant tissues has not been studied. This study tested expression and correlation between GWAS-identified asthma genes and asthma or asthma severity. METHODS: Correlation analyses of expression levels of GWAS-identified asthma genes and asthma-related biomarkers were performed in cells from human bronchial epithelial biopsy (BEC, n = 107) and bronchial alveolar lavage (BAL, n = 94). RESULTS: Expression levels of asthma genes between BEC and BAL and with asthma or asthma severity were weakly correlated. The expression levels of IL18R1 were consistently higher in asthma than controls or in severe asthma than mild/moderate asthma in BEC and BAL (p < 0.05). In RAD50-IL13 region, the expression levels of RAD50, not IL4, IL5, or IL13, were positively correlated between BEC and BAL (ρ = 0.53, P = 4.5 × 10(-6)). The expression levels of IL13 were positively correlated with IL5 in BEC (ρ = 0.35, P = 1.9 × 10(-4)) and IL4 in BAL (ρ = 0.42, P = 2.5 × 10(-5)), respectively. rs3798134 in RAD50, a GWAS-identified SNP, was correlated with IL13 expression and the expression levels of IL13 were correlated with asthma (P = 0.03). rs17772583 in RAD50 was significantly correlated with RAD50 expression in BAL and BEC (P = 7.4 × 10(-7) and 0.04) but was not associated with asthma. CONCLUSIONS: This is the first report studying the expression of GWAS-identified asthma genes in BEC and BAL. IL13, rather than RAD50, IL4, or IL5, is more likely to be the asthma susceptibility gene. Our study illustrates tissue-specific expression of asthma-related genes. Therefore, whenever possible, disease-relevant tissues should be used for transcription analysis.

Genome-wide association study identifies TH1 pathway genes associated with lung function in asthmatic patients.

BACKGROUND: Recent meta-analyses of genome-wide association studies in general populations of European descent have identified 28 loci for lung function. OBJECTIVE: We sought to identify novel lung function loci specifically for asthma and to confirm lung function loci identified in general populations. METHODS: Genome-wide association studies of lung function (percent predicted FEV1 [ppFEV1], percent predicted forced vital capacity, and FEV1/forced vital capacity ratio) were performed in 4 white populations of European descent (n = 1544), followed by meta-analyses. RESULTS: Seven of 28 previously identified lung function loci (HHIP, FAM13A, THSD4, GSTCD, NOTCH4-AGER, RARB, and ZNF323) identified in general populations were confirmed at single nucleotide polymorphism (SNP) levels (P < .05). Four of 32 loci (IL12A, IL12RB1, STAT4, and IRF2) associated with ppFEV1 (P < 10(-4)) belong to the TH1 or IL-12 cytokine family pathway. By using a linear additive model, these 4 TH1 pathway SNPs cumulatively explained 2.9% to 7.8% of the variance in ppFEV1 values in 4 populations (P = 3 × 10(-11)). Genetic scores of these 4 SNPs were associated with ppFEV1 values (P = 2 × 10(-7)) and the American Thoracic Society severe asthma classification (P = .005) in the Severe Asthma Research Program population. TH2 pathway genes (IL13, TSLP, IL33, and IL1RL1) conferring asthma susceptibility were not associated with lung function. CONCLUSION: Genes involved in airway structure/remodeling are associated with lung function in both general populations and asthmatic subjects. TH1 pathway genes involved in anti-virus/bacterial infection and inflammation modify lung function in asthmatic subjects. Genes associated with lung function that might affect asthma severity are distinct from those genes associated with asthma susceptibility.

Genome-wide association studies of asthma indicate opposite immunopathogenesis direction from autoimmune diseases.

BACKGROUND: Genome-wide association studies (GWASs) of asthma have consistently implicated the ORM1-like 3 and gasdermin B (ORMDL3-GSDMB), IL33, IL-1 receptor-like 1 and IL-18 receptor 1 (IL1RL1-IL18R1), RAD50-IL13, thymic stromal lymphopoietin and WD repeat domain 36 region (TSLP-WDR36), and HLA-DR/DQ regions. OBJECTIVE: A GWAS of asthma was performed in a non-Hispanic white population. METHODS: A GWAS was performed in 813 Severe Asthma Research Program/Collaborative Studies on the Genetics of Asthma/Chicago Asthma Genetics Study cases and 1564 control subjects. The GWAS results were compared with those of the published GWASs of autoimmune diseases. RESULTS: Multiple single nucleotide polymorphisms in the TNFAIP3 interacting protein 1 (TNIP1) gene, which interacts with TNFAIP3 and inhibits the TNF-α-induced nuclear factor κB inflammation pathway, were associated with asthma: rs1422673 (P = 3.44 × 10(-7)) and rs10036748 (P = 1.41 × 10(-6), r(2) = 0.67). rs1422673 was also associated with asthma in the published GABRIEL (P = .018) and EVE (P = 1.31 × 10(-5)) studies. The minor allele T of rs20541 in IL13 is the risk allele for asthma but the protective allele for psoriasis. The minor allele T of rs2395185 in HLA-DRA is the risk allele for asthma but the protective allele for ulcerative colitis. The minor allele A of rs2872507 in GSDMB is the protective allele for asthma but the risk allele for rheumatoid arthritis, Crohn disease, and ulcerative colitis. The T allele of rs10036748 in the TNIP1 gene is the minor protective allele for asthma but the minor or major risk allele for systemic lupus erythematosus and systemic sclerosis in non-Hispanic white or Chinese subjects, respectively. CONCLUSIONS: Our study suggests that single nucleotide polymorphisms associated with both asthma and autoimmune diseases might have opposite effects on immunopathogenesis.

Genome-wide ancestry association testing identifies a common European variant on 6q14.1 as a risk factor for asthma in African American subjects.

BACKGROUND: Genetic variants that contribute to asthma susceptibility might be present at varying frequencies in different populations, which is an important consideration and advantage for performing genetic association studies in admixed populations. OBJECTIVE: We sought to identify asthma-associated loci in African American subjects. METHODS: We compared local African and European ancestry estimated from dense single nucleotide polymorphism genotype data in African American adults with asthma and nonasthmatic control subjects. Allelic tests of association were performed within the candidate regions identified, correcting for local European admixture. RESULTS: We identified a significant ancestry association peak on chromosome 6q. Allelic tests for association within this region identified a single nucleotide polymorphism (rs1361549) on 6q14.1 that was associated with asthma exclusively in African American subjects with local European admixture (odds ratio, 2.2). The risk allele is common in Europe (42% in the HapMap population of Utah residents with Northern and Western European ancestry from the Centre d'Etude du Polymorphisme Humain collection) but absent in West Africa (0% in the HapMap population of Yorubans in Ibadan, Nigeria), suggesting the allele is present in African American subjects because of recent European admixture. We replicated our findings in Puerto Rican subjects and similarly found that the signal of association is largely specific to subjects who are heterozygous for African and non-African ancestry at 6q14.1. However, we found no evidence for association in European American or Puerto Rican subjects in the absence of local African ancestry, suggesting that the association with asthma at rs1361549 is due to an environmental or genetic interaction. CONCLUSION: We identified a novel asthma-associated locus that is relevant to admixed populations with African ancestry and highlight the importance of considering local ancestry in genetic association studies of admixed populations.

Importance of hedgehog interacting protein and other lung function genes in asthma.

BACKGROUND: Two recent large meta-analyses of genome-wide association studies of lung function in general populations of European descent identified 11 candidate genes/regions. The importance of these genes in lung function in white and African American subjects with asthma is unknown. OBJECTIVES: To determine whether genes that regulate lung function in general populations are associated with lung function abnormalities in subjects with asthma from different racial groups. METHODS: Single nucleotide polymorphisms (SNPs) were tested in 5 asthma populations (N = 1441) for association with pulmonary function, and meta-analysis was performed across populations. The SNPs with the highest significance were then tested for association with bronchodilator reversibility and bronchial hyperresponsiveness to methacholine. A joint analysis of consistently replicated SNPs was performed to predict lung function in asthma. RESULTS: Hedgehog interacting protein (HHIP) on chromosome 4q31 was associated with lung function in all 5 populations (rs1512288: P(meta) = 9.62E-05 and 3.23E-05 for percent predicted FEV(1) [ppFEV(1)] and percent predicted forced vital capacity [ppFVC], respectively). The SNPs in HHIP were also associated with reversibility (P < .05) but not bronchial hyperresponsiveness to methacholine. Because of differences in linkage disequilibrium in the African American subjects, the most relevant SNPs in HHIP were identified. A subset of normal lung function genes, including HHIP, family with sequence similarity 13, member A (FAM13A), and patched homolog 1 (PTCH1), together predict lung function abnormalities, a measure of severity in white and African American subjects with asthma. CONCLUSION: A subset of the genes, including HHIP, that regulate lung function in general populations are associated with abnormal lung function in asthma in non-Hispanic white and African American subjects.

A genome-wide association study of bronchodilator response in participants of European and African ancestry from six independent cohorts.

Introduction: Bronchodilator response (BDR) is a measurement of acute bronchodilation in response to short-acting ß2-agonists, with a heritability between 10 and 40%. Identifying genetic variants associated with BDR may lead to a better understanding of its complex pathophysiology. Methods: We performed a genome-wide association study (GWAS) of BDR in six adult cohorts with participants of European ancestry (EA) and African ancestry (AA) including community cohorts and cohorts ascertained on the basis of obstructive pulmonary disease. Validation analysis was carried out in two paediatric asthma cohorts. Results: A total of 10 623 EA and 3597 AA participants were included in the analyses. No single nucleotide polymorphism (SNP) was associated with BDR at the conventional genome-wide significance threshold (p<5×10-8). Performing fine mapping and using a threshold of p<5×10-6 to identify suggestive variants of interest, we identified three SNPs with possible biological relevance: rs35870000 (within FREM1), which may be involved in IgE- and IL5-induced changes in airway smooth muscle cell responsiveness; rs10426116 (within ZNF284), a zinc finger protein, which has been implicated in asthma and BDR previously; and rs4782614 (near ATP2C2), involved in calcium transmembrane transport. Validation in paediatric cohorts yielded no significant SNPs, possibly due to age-genotype interaction effects. Conclusion: Ancestry-stratified and ancestry-combined GWAS meta-analyses of over 14 000 participants did not identify genetic variants associated with BDR at the genome-wide significance threshold, although a less stringent threshold identified three variants showing suggestive evidence of association. A common definition and protocol for measuring BDR in research may improve future efforts to identify variants associated with BDR.

Age by Single Nucleotide Polymorphism Interactions on Bronchodilator Response in Asthmatics.

An unaddressed and important issue is the role age plays in modulating response to short acting ß2-agonists in individuals with asthma. The objective of this study was to identify whether age modifies genetic associations of single nucleotide polymorphisms (SNPs) with bronchodilator response (BDR) to ß2-agonists. Using three cohorts with a total of 892 subjects, we ran a genome wide interaction study (GWIS) for each cohort to examine SNP by age interactions with BDR. A fixed effect meta-analysis was used to combine the results. In order to determine if previously identified BDR SNPs had an age interaction, we also examined 16 polymorphisms in candidate genes from two published genome wide association studies (GWAS) of BDR. There were no significant SNP by age interactions on BDR using the genome wide significance level of 5 × 10-8. Using a suggestive significance level of 5 × 10-6, three interactions, including one for a SNP within PRAG1 (rs4840337), were significant and replicated at the significance level of 0.05. Considering candidate genes from two previous GWAS of BDR, three SNPs (rs10476900 (near ADRB2) [p-value = 0.009], rs10827492 (CREM) [p-value = 0.02], and rs72646209 (NCOA3) [p-value = 0.02]) had a marginally significant interaction with age on BDR (p < 0.05). Our results suggest age may be an important modifier of genetic associations for BDR in asthma.

Pharmacogenetic studies of long-acting beta agonist and inhaled corticosteroid responsiveness in randomised controlled trials of individuals of African descent with asthma.

BACKGROUND: Pharmacogenetic studies in asthma cohorts, primarily made up of White people of European descent, have identified loci associated with response to inhaled beta agonists and corticosteroids (ICSs). Differences exist in how individuals from different ancestral backgrounds respond to long-acting beta agonist (LABA) and ICSs. Therefore, we sought to understand the pharmacogenetic mechanisms regulating therapeutic responsiveness in individuals of African descent. METHODS: We did ancestry-based pharmacogenetic studies of children (aged 5-11 years) and adolescents and adults (aged 12-69 years) from the Best African Response to Drug (BARD) trials, in which participants with asthma uncontrolled with low-dose ICS (fluticasone propionate 50 µg in children, 100 µg in adolescents and adults) received different step-up combination therapies. The hierarchal composite outcome of pairwise superior responsiveness in BARD was based on asthma exacerbations, a 31-day difference in annualised asthma-control days, or a 5% difference in percentage predicted FEV1. We did whole-genome admixture mapping of 15 159 ancestral segments within 312 independent regions, stratified by the two age groups. The two co-primary outcome comparisons were the step up from low-dose ICS to the quintuple dose of ICS (5 × ICS: 250 µg twice daily in children and 500 µg twice daily in adolescents and adults) versus double dose (2-2·5 × ICS: 100 µg twice daily in children, 250 µg twice daily in adolescents and adults), and 5 × ICS versus 100 µg fluticasone plus a LABA (salmeterol 50 µg twice daily). We used a genome-wide significance threshold of p<1·6 × 10-4, and tested for replication using independent cohorts of individuals of African descent with asthma. FINDINGS: We included 249 unrelated children and 267 unrelated adolescents and adults in the BARD pharmacogenetic analysis. In children, we identified a significant admixture mapping peak for superior responsiveness to 5 × ICS versus 100 µg fluticasone plus salmeterol on chromosome 12 (odds ratio [ORlocal African] 3·95, 95% CI 2·02-7·72, p=6·1 × 10-5) fine mapped to a locus adjacent to RNFT2 and NOS1 (rs73399224, ORallele dose 0·17, 95% CI 0·07-0·42, p=8·4 × 10-5). In adolescents and adults, we identified a peak for superior responsiveness to 5 × ICS versus 2·5 × ICS on chromosome 22 (ORlocal African 3·35, 1·98-5·67, p=6·8 × 10-6) containing a locus adjacent to TPST2 (rs5752429, ORallele dose 0·21, 0·09-0·52, p=5·7 × 10-4). We replicated rs5752429 and nominally replicated rs73399224 in independent African American cohorts. INTERPRETATION: BARD is the first genome-wide pharmacogenetic study of LABA and ICS response in clinical trials of individuals of African descent to detect and replicate genome-wide significant loci. Admixture mapping of the composite BARD trial outcome enabled the identification of novel pharmacogenetic variation accounting for differential therapeutic responses in people of African descent with asthma. FUNDING: National Institutes of Health, National Heart, Lung, and Blood Institute.

Genetic and non-genetic factors affecting the expression of COVID-19-relevant genes in the large airway epithelium.

BACKGROUND: The large airway epithelial barrier provides one of the first lines of defense against respiratory viruses, including SARS-CoV-2 that causes COVID-19. Substantial inter-individual variability in individual disease courses is hypothesized to be partially mediated by the differential regulation of the genes that interact with the SARS-CoV-2 virus or are involved in the subsequent host response. Here, we comprehensively investigated non-genetic and genetic factors influencing COVID-19-relevant bronchial epithelial gene expression. METHODS: We analyzed RNA-sequencing data from bronchial epithelial brushings obtained from uninfected individuals. We related ACE2 gene expression to host and environmental factors in the SPIROMICS cohort of smokers with and without chronic obstructive pulmonary disease (COPD) and replicated these associations in two asthma cohorts, SARP and MAST. To identify airway biology beyond ACE2 binding that may contribute to increased susceptibility, we used gene set enrichment analyses to determine if gene expression changes indicative of a suppressed airway immune response observed early in SARS-CoV-2 infection are also observed in association with host factors. To identify host genetic variants affecting COVID-19 susceptibility in SPIROMICS, we performed expression quantitative trait (eQTL) mapping and investigated the phenotypic associations of the eQTL variants. RESULTS: We found that ACE2 expression was higher in relation to active smoking, obesity, and hypertension that are known risk factors of COVID-19 severity, while an association with interferon-related inflammation was driven by the truncated, non-binding ACE2 isoform. We discovered that expression patterns of a suppressed airway immune response to early SARS-CoV-2 infection, compared to other viruses, are similar to patterns associated with obesity, hypertension, and cardiovascular disease, which may thus contribute to a COVID-19-susceptible airway environment. eQTL mapping identified regulatory variants for genes implicated in COVID-19, some of which had pheWAS evidence for their potential role in respiratory infections. CONCLUSIONS: These data provide evidence that clinically relevant variation in the expression of COVID-19-related genes is associated with host factors, environmental exposures, and likely host genetic variation.

Identification of PCDH1 as a novel susceptibility gene for bronchial hyperresponsiveness.

RATIONALE: Asthma is a chronic inflammatory airway disease that affects more than 300 million individuals worldwide. Asthma is caused by interaction of genetic and environmental factors. Bronchial hyperresponsiveness (BHR) is a hallmark of asthma and results from increased sensitivity of the airways to physical or chemical stimulants. BHR and asthma are linked to chromosome 5q31-q33. OBJECTIVES: To identify a gene for BHR on chromosome 5q31-q33. METHODS: In 200 Dutch families with asthma, linkage analysis and fine mapping were performed, and the Protocadherin 1 gene (PCDH1) was identified. PCDH1 was resequenced in 96 subjects from ethnically diverse populations to identify novel sequence variants. Subsequent replication studies were undertaken in seven populations from The Netherlands, the United Kingdom, and the United States, including two general population samples, two family samples, and three case-control samples. PCDH1 mRNA and protein expression was investigated using polymerase chain reaction, Western blotting, and immunohistochemistry. MEASUREMENTS AND MAIN RESULTS: In seven out of eight populations (n = 6,168) from The Netherlands, United Kingdom, and United States, PCHD1 gene variants were significantly associated with BHR (P values, 0.005-0.05) This association was present in both families with asthma and general populations. PCDH1 mRNA and protein were expressed in airway epithelial cells and in macrophages. CONCLUSIONS: PCDH1 is a novel gene for BHR in adults and children. The identification of PCDH1 as a BHR susceptibility gene may suggest that a structural defect in the integrity of the airway epithelium, the first line of defense against inhaled substances, contributes to the development of BHR.