Peter Le Souef

Identification of IL6R and chromosome 11q13.5 as risk loci for asthma

BACKGROUND We aimed to identify novel genetic variants affecting asthma risk, since these might provide novel insights into molecular mechanisms underlying the disease. METHODS We did a genome-wide association study (GWAS) in 2669 physician-diagnosed asthmatics and 4528 controls from Australia. Seven loci were prioritised for replication after combining our results with those from the GABRIEL consortium (n=26,475), and these were tested in an additional 25,358 independent samples from four in-silico cohorts. Quantitative multi-marker scores of genetic load were constructed on the basis of results from the GABRIEL study and tested for association with asthma in our Australian GWAS dataset. FINDINGS Two loci were confirmed to associate with asthma risk in the replication cohorts and reached genome-wide significance in the combined analysis of all available studies (n=57,800): rs4129267 (OR 1·09, combined p=2·4×10(-8)) in the interleukin-6 receptor (IL6R) gene and rs7130588 (OR 1·09, p=1·8×10(-8)) on chromosome 11q13.5 near the leucine-rich repeat containing 32 gene (LRRC32, also known as GARP). The 11q13.5 locus was significantly associated with atopic status among asthmatics (OR 1·33, p=7×10(-4)), suggesting that it is a risk factor for allergic but not non-allergic asthma. Multi-marker association results are consistent with a highly polygenic contribution to asthma risk, including loci with weak effects that might be shared with other immune-related diseases, such as NDFIP1, HLA-B, LPP, and BACH2. INTERPRETATION The IL6R association further supports the hypothesis that cytokine signalling dysregulation affects asthma risk, and raises the possibility that an IL6R antagonist (tocilizumab) may be effective to treat the disease, perhaps in a genotype-dependent manner. Results for the 11q13.5 locus suggest that it directly increases the risk of allergic sensitisation which, in turn, increases the risk of subsequent development of asthma. Larger or more functionally focused studies are needed to characterise the many loci with modest effects that remain to be identified for asthma. FUNDING National Health and Medical Research Council of Australia. A full list of funding sources is provided in the webappendix.

Genome-Wide Association Studies of Asthma in Population-Based Cohorts Confirm Known and Suggested Loci and Identify an Additional Association near HLA

Rationale Asthma has substantial morbidity and mortality and a strong genetic component, but identification of genetic risk factors is limited by availability of suitable studies. Objectives To test if population-based cohorts with self-reported physician-diagnosed asthma and genome-wide association (GWA) data could be used to validate known associations with asthma and identify novel associations. Methods The APCAT (Analysis in Population-based Cohorts of Asthma Traits) consortium consists of 1,716 individuals with asthma and 16,888 healthy controls from six European-descent population-based cohorts. We examined associations in APCAT of thirteen variants previously reported as genome-wide significant (P<5x10−8) and three variants reported as suggestive (P<5×10−7). We also searched for novel associations in APCAT (Stage 1) and followed-up the most promising variants in 4,035 asthmatics and 11,251 healthy controls (Stage 2). Finally, we conducted the first genome-wide screen for interactions with smoking or hay fever. Main Results We observed association in the same direction for all thirteen previously reported variants and nominally replicated ten of them. One variant that was previously suggestive, rs11071559 in RORA, now reaches genome-wide significance when combined with our data (P = 2.4×10−9). We also identified two genome-wide significant associations: rs13408661 near IL1RL1/IL18R1 (P Stage1+Stage2 = 1.1x10−9), which is correlated with a variant recently shown to be associated with asthma (rs3771180), and rs9268516 in the HLA region (P Stage1+Stage2 = 1.1x10−8), which appears to be independent of previously reported associations in this locus. Finally, we found no strong evidence for gene-environment interactions with smoking or hay fever status. Conclusions Population-based cohorts with simple asthma phenotypes represent a valuable and largely untapped resource for genetic studies of asthma.

Evolutionary adaptation of inflammatory immune responses in human beings

Modern mans ancestors lived in an environment where infectious, tropical diseases would have been endemic. We postulate that this relatively hostile environment would have caused genetic selection for increased proinflammatory immune responses. On migrating to temperate regions, pronounced proinflammatory responses would have been less important and selected against due to increased mortality from overly vigorous responses to harmless environmental agents. This hypothesis is supported by the observation that proinflammatory alleles in several genes involved in inflammation are more prevalent in populations with long-term tropical ancestry than those with long-term residence in temperate regions. In addition, when the former populations relocate from a tropical to a temperate region, they have a higher incidence of allergic inflammatory diseases than the latter populations. These observations suggest that there may be general patterns of recent evolutionary adaptation of the human immune system to particular regions and that these adaptations can produce differences in disease susceptibility.

Genome-wide association analysis identifies 11 risk variants associated with the asthma with hay fever phenotype.

BACKGROUND To date, no genome-wide association study (GWAS) has considered the combined phenotype of asthma with hay fever. Previous analyses of family data from the Tasmanian Longitudinal Health Study provide evidence that this phenotype has a stronger genetic cause than asthma without hay fever. OBJECTIVE We sought to perform a GWAS of asthma with hay fever to identify variants associated with having both diseases. METHODS We performed a meta-analysis of GWASs comparing persons with both physician-diagnosed asthma and hay fever (n = 6,685) with persons with neither disease (n = 14,091). RESULTS At genome-wide significance, we identified 11 independent variants associated with the risk of having asthma with hay fever, including 2 associations reaching this level of significance with allergic disease for the first time: ZBTB10 (rs7009110; odds ratio [OR], 1.14; P = 4 × 10(-9)) and CLEC16A (rs62026376; OR, 1.17; P = 1 × 10(-8)). The rs62026376:C allele associated with increased asthma with hay fever risk has been found to be associated also with decreased expression of the nearby DEXI gene in monocytes. The 11 variants were associated with the risk of asthma and hay fever separately, but the estimated associations with the individual phenotypes were weaker than with the combined asthma with hay fever phenotype. A variant near LRRC32 was a stronger risk factor for hay fever than for asthma, whereas the reverse was observed for variants in/near GSDMA and TSLP. Single nucleotide polymorphisms with suggestive evidence for association with asthma with hay fever risk included rs41295115 near IL2RA (OR, 1.28; P = 5 × 10(-7)) and rs76043829 in TNS1 (OR, 1.23; P = 2 × 10(-6)). CONCLUSION By focusing on the combined phenotype of asthma with hay fever, variants associated with the risk of allergic disease can be identified with greater efficiency.

Robust Estimation of Experimentwise P Values Applied to a Genome Scan of Multiple Asthma Traits Identifies a New Region of Significant Linkage on Chromosome 20q13

Over 30 genomic regions show linkage to asthma traits. Six asthma genes have been cloned, but the putative loci in many linked regions have not been identified. To search for asthma susceptibility loci, we performed genomewide univariate linkage analyses of seven asthma traits, using 202 Australian families ascertained through a twin proband. House-dust mite sensitivity (Dpter) exceeded the empirical threshold for significant linkage at 102 cM on chromosome 20q13, near marker D20S173 (empirical pointwise P = .00001 and genomewide P = .005, both uncorrected for multiple-trait testing). Atopy, bronchial hyperresponsiveness (BHR), and forced expiratory volume in 1 s (FEV1) were also linked to this region. In addition, 16 regions were linked to at least one trait at the suggestive level, including 12q24, which has consistently shown linkage to asthma traits in other studies. Some regions were expected to be false-positives arising from multiple-trait testing. To address this, we developed a new approach to estimate genomewide significance that accounts for multiple-trait testing and for correlation between traits and that does not require a Bonferroni correction. With this approach, Dpter remained significantly linked to 20q13 (empirical genomewide P = .042), and airway obstruction remained linked to 12q24 at the suggestive level. Finally, we extended this method to show that the linkage of Dpter, atopy, BHR, FEV1, asthma, and airway obstruction to chromosome 20q13 is unlikely to be due to chance and may result from a quantitative trait locus in this region that affects several of these traits.

Urinary Cotinine Levels in Early Pregnancy

Summary: We tested the hypotheses that in early pregnancy smokers have lower urinary cotinine levels than nonpregnant smokers, and that pregnant nonsmokers exposed to passive smoke have higher cotinine levels than nonsmokers not exposed to passive smoke. This was a prospective, quantitative, comparative study of the urinary cotinine levels and smoking characteristics of pregnant and nonpregnant females. A urine specimen was collected from each subject and the cotinine/creatinine level determined using radio‐immunoassay. A questionnaire regarding smoking status, health issues, and demographic variables was administered to each subject. There was no difference in cotinine level between pregnant smokers and control smokers. Pregnant nonsmokers with passive smoke exposure had higher cotinine levels than the same group not passively exposed. Confidence in ability to stop smoking was associated with lower numbers of cigarettes consumed, but was not reflected by lower cotinine levels. These data suggest that if the mother smokes, the fetus is exposed from conception to levels of nicotine which are as high as those in adult female smokers who are not pregnant. The level of fetal exposure to nicotine during early gestation is independent of intention to alter smoking behaviour. Women should be targeted for antismoking public health messages before conception.

Gene-based analysis of regulatory variants identifies 4 putative novel asthma risk genes related to nucleotide synthesis and signaling

Background: Hundreds of genetic variants are thought to contribute to variation in asthma risk by modulating gene expression. Methods that increase the power of genome‐wide association studies (GWASs) to identify risk‐associated variants are needed. Objective: We sought to develop a method that aggregates the evidence for association with disease risk across expression quantitative trait loci (eQTLs) of a gene and use this approach to identify asthma risk genes. Methods: We developed a gene‐based test and software package called EUGENE that (1) is applicable to GWAS summary statistics; (2) considers both cis‐ and trans‐eQTLs; (3) incorporates eQTLs identified in different tissues; and (4) uses simulations to account for multiple testing. We applied this approach to 2 published asthma GWASs (combined n = 46,044) and used mouse studies to provide initial functional insights into 2 genes with novel genetic associations. Results: We tested the association between asthma and 17,190 genes that were found to have cis‐ and/or trans‐eQTLs across 16 published eQTL studies. At an empirical FDR of 5%, 48 genes were associated with asthma risk. Of these, for 37, the association was driven by eQTLs located in established risk loci for allergic disease, including 6 genes not previously implicated in disease cause (eg, LIMS1, TINF2, and SAFB). The remaining 11 significant genes represent potential novel genetic associations with asthma. The association with 4 of these replicated in an independent GWAS: B4GALT3, USMG5, P2RY13, and P2RY14, which are genes involved in nucleotide synthesis or nucleotide‐dependent cell activation. In mouse studies, P2ry13 and P2ry14—purinergic receptors activated by adenosine 5‐diphosphate and UDP‐sugars, respectively—were upregulated after allergen challenge, notably in airway epithelial cells, eosinophils, and neutrophils. Intranasal exposure with receptor agonists induced the release of IL‐33 and subsequent eosinophil infiltration into the lungs. Conclusion: We identified novel associations between asthma and eQTLs for 4 genes related to nucleotide synthesis/signaling and demonstrated the power of gene‐based analyses of GWASs.

Mutation analysis of interleukin-5 in an asthmatic cohort

Interleukin‐5 (IL‐5) is a potential candidate gene in the pathogenesis of asthma, as it is the main cytokine controlling eosinophil activity and eosinophils are pivotal in the development of airway inflammation. Mutation detection studies were performed on the IL‐5 gene and the α‐chain of its receptor in 30 asthmatic and 30 nonasthmatic subjects. Single‐strand conformational polymorphism (SSCP) and heteroduplex analysis (HA) did not reveal any change from the reported normal sequence in all 4 exons of IL‐5 as well as the promoter and 3′‐untranslated regions of the gene. No SSCP variations were seen within the complete coding sequence of the IL‐5 receptor α‐chain. Mutations of the IL‐5 gene coding region, its promoter and receptor are unlikely to be common causes of an inherited predisposition to asthma. Hum Mutat 11:51–54, 1998.

Asthma education material for children and their families; a global survey of current resources

One of the keys to high quality paediatric asthma management is the provision of age appropriate information regarding the disease and its management. In order to determine whether the generation of a minimum dataset of information which can be translated into a wide range of languages might be used to assist children and their parents around the world, we undertook a survey of national Member Societies of the World Allergy Organization (WAO) to determine what educational material on asthma for children and their families already exists.A questionnaire was developed using Survey Monkey and distributed in 2014 to 263 representatives of the WAO member Societies from 95 countries.Thirty-three replies were received from thirty-one countries. The survey highlighted a considerable disparity in availability of material among the responding countries, with some countries reporting that information was freely available in hard copy and online and others reporting a lack of suitable material locally.The results highlight the need to develop a core set of simple, clear and consistent age appropriate information that can be easily translated and delivered in a cultural and educationally effective format.