Adrian Cortes

Identification of multiple risk variants for ankylosing spondylitis through high-density genotyping of immune-related loci

Ankylosing spondylitis is a common, highly heritable inflammatory arthritis affecting primarily the spine and pelvis. In addition to HLA-B*27 alleles, 12 loci have previously been identified that are associated with ankylosing spondylitis in populations of European ancestry, and 2 associated loci have been identified in Asians. In this study, we used the Illumina Immunochip microarray to perform a case-control association study involving 10,619 individuals with ankylosing spondylitis (cases) and 15,145 controls. We identified 13 new risk loci and 12 additional ankylosing spondylitis–associated haplotypes at 11 loci. Two ankylosing spondylitis–associated regions have now been identified encoding four aminopeptidases that are involved in peptide processing before major histocompatibility complex (MHC) class I presentation. Protective variants at two of these loci are associated both with reduced aminopeptidase function and with MHC class I cell surface expression.

Major histocompatibility complex associations of ankylosing spondylitis are complex and involve further epistasis with ERAP1.

Ankylosing spondylitis (AS) is a common, highly heritable, inflammatory arthritis for which HLA-B*27 is the major genetic risk factor, although its role in the aetiology of AS remains elusive. To better understand the genetic basis of the MHC susceptibility loci, we genotyped 7,264 MHC SNPs in 22,647 AS cases and controls of European descent. We impute SNPs, classical HLA alleles and amino-acid residues within HLA proteins, and tested these for association to AS status. Here we show that in addition to effects due to HLA-B*27 alleles, several other HLA-B alleles also affect susceptibility. After controlling for the associated haplotypes in HLA-B, we observe independent associations with variants in the HLA-A, HLA-DPB1 and HLA-DRB1 loci. We also demonstrate that the ERAP1 SNP rs30187 association is not restricted only to carriers of HLA-B*27 but also found in HLA-B*40:01 carriers independently of HLA-B*27 genotype.

Genome-wide genetic data on ~500,000 UK Biobank participants

The UK Biobank project is a large prospective cohort study of ~500,000 individuals from across the United Kingdom, aged between 40-69 at recruitment. A rich variety of phenotypic and health-related information is available on each participant, making the resource unprecedented in its size and scope. Here we describe the genome-wide genotype data (~805,000 markers) collected on all individuals in the cohort and its quality control procedures. Genotype data on this scale offers novel opportunities for assessing quality issues, although the wide range of ancestries of the individuals in the cohort also creates particular challenges. We also conducted a set of analyses that reveal properties of the genetic data – such as population structure and relatedness – that can be important for downstream analyses. In addition, we phased and imputed genotypes into the dataset, using computationally efficient methods combined with the Haplotype Reference Consortium (HRC) and UK10K haplotype resource. This increases the number of testable variants by over 100-fold to ~96 million variants. We also imputed classical allelic variation at 11 human leukocyte antigen (HLA) genes, and as a quality control check of this imputation, we replicate signals of known associations between HLA alleles and many common diseases. We describe tools that allow efficient genome-wide association studies (GWAS) of multiple traits and fast phenome-wide association studies (PheWAS), which work together with a new compressed file format that has been used to distribute the dataset. As a further check of the genotyped and imputed datasets, we performed a test-case genome-wide association scan on a well-studied human trait, standing height.

Fine-mapping inflammatory bowel disease loci to single-variant resolution

Inflammatory bowel diseases are chronic gastrointestinal inflammatory disorders that affect millions of people worldwide. Genome-wide association studies have identified 200 inflammatory bowel disease-associated loci, but few have been conclusively resolved to specific functional variants. Here we report fine-mapping of 94 inflammatory bowel disease loci using high-density genotyping in 67,852 individuals. We pinpoint 18 associations to a single causal variant with greater than 95% certainty, and an additional 27 associations to a single variant with greater than 50% certainty. These 45 variants are significantly enriched for protein-coding changes (n = 13), direct disruption of transcription-factor binding sites (n = 3), and tissue-specific epigenetic marks (n = 10), with the last category showing enrichment in specific immune cells among associations stronger in Crohn’s disease and in gut mucosa among associations stronger in ulcerative colitis. The results of this study suggest that high-resolution fine-mapping in large samples can convert many discoveries from genome-wide association studies into statistically convincing causal variants, providing a powerful substrate for experimental elucidation of disease mechanisms.

Genetic Dissection of Acute Anterior Uveitis Reveals Similarities and Differences in Associations observed with Ankylosing Spondylitis

To use high‐density genotyping to investigate the genetic associations of acute anterior uveitis (AAU) in patients with and those without ankylosing spondylitis (AS).

Association study of genes related to bone formation and resorption and the extent of radiographic change in ankylosing spondylitis

Objective To identify genetic associations with severity of radiographic damage in ankylosing spondylitis (AS). Method We studied 1537 AS cases of European descent; all fulfilled the modified New York Criteria. Radiographic severity was assessed from digitised lateral radiographs of the cervical and lumbar spine using the modified Stoke Ankylosing Spondylitis Spinal Score (mSASSS). A two-phase genotyping design was used. In phase 1, 498 single nucleotide polymorphisms (SNPs) were genotyped in 688 cases; these were selected to capture >90% of the common haplotypic variation in the exons, exon–intron boundaries, and 5 kb flanking DNA in the 5′ and 3′ UTR of 74 genes involved in anabolic or catabolic bone pathways. In phase 2, 15 SNPs exhibiting p<0.05 were genotyped in a further cohort of 830 AS cases; results were analysed both separately and in combination with the discovery phase data. Association was tested by contingency tables after separating the samples into ‘mild’ and ‘severe’ groups, defined as the bottom and top 40% by mSASSS, adjusted for gender and disease duration. Results Experiment-wise association was observed with the SNP rs8092336 (combined OR 0.32, p=1.2×10−5), which lies within RANK (receptor activator of NFκB), a gene involved in osteoclastogenesis, and in the interaction between T cells and dendritic cells. Association was also found with the SNP rs1236913 in PTGS1 (prostaglandin-endoperoxide synthase 1, cyclooxygenase 1), giving an OR of 0.53 (p=2.6×10−3). There was no observed association between radiographic severity and HLA-B*27. Conclusions These findings support roles for bone resorption and prostaglandins pathways in the osteoproliferative changes in AS.

Current Smoking is Associated with Incident Ankylosing Spondylitis — The HUNT Population-based Norwegian Health Study

Objective. Smoking contributes to progression of ankylosing spondylitis (AS). Because smoking is also a risk factor for incident rheumatoid arthritis (RA) and psoriatic arthritis, our aim was to test whether smoking habits are associated with incident AS. Methods. Using data from the HUNT health study of the entire adult population of Nord-Trøndelag, Norway, participants in HUNT2 (1995–1997) and HUNT3 (2006–2008) were identified who reported a diagnosis of AS in HUNT3 but not in HUNT2 (n = 107). Incident AS cases were compared with AS-unaffected individuals (n = 35,278) in a case-control design. Participants with RA were excluded. Results. Present smoking was significantly associated with incident self-reported AS in logistic regression adjusted for potential confounders (OR 1.99, 95% CI 1.28–3.11, p = 0.002). Previous smoking (OR 1.15, 95% CI 0.66–2.02, p = 0.62) or total pack-years at HUNT2 (OR 1.01, 95% CI 0.99–1.04, p = 0.21) were not significant. The association with present smoking remained significant in various sensitivity analyses: including only cases with high probability of true AS diagnosis (OR 1.82, 95% CI 1.03–3.19, p = 0.04); including only cases with AS reported more than 3–5 years after HUNT2 (OR 2.34, 95% CI 1.09–5.03, p = 0.029), or including only participants genotyped for HLA-B27 (94 cases and 859 controls) adjusting for genotype (OR 1.79, 95% CI 1.04–2.85, p = 0.033). Hypertension was also significantly associated with incident AS (OR from 1.65 to 2.81). Conclusion. In the HUNT population-based study, incident AS was associated with current smoking and hypertension. If verified in further studies, this suggests that smoking should be discouraged in those at a higher AS risk, e.g., with a family history or carrying HLA-B27.

Resolving TYK2 locus genotype-to-phenotype differences in autoimmunity

Resolving TYK2 locus genotype-to-phenotype differences reveals an immune signaling optimum that may be exploited therapeutically for treating autoimmune diseases. TYK2’s balancing act Determining the biological consequences of the thousands of genetic variants that contribute to common diseases for the purpose of improving health care is challenging. Genetic variants that influence autoimmune diseases have been identified in the tyrosine kinase 2 (TYK2) gene, but conflicting evidence regarding their biological impact obscures the therapeutic potential of TYK2. By resolving this conflict, Dendrou et al. have revealed a genetic effect that drives an optimal degree of immune signaling: low enough to be protective against autoimmunity but high enough to prevent immunodeficiency. These findings indicate that TYK2 may be a potential drug target in a number of autoimmune conditions. Thousands of genetic variants have been identified, which contribute to the development of complex diseases, but determining how to elucidate their biological consequences for translation into clinical benefit is challenging. Conflicting evidence regarding the functional impact of genetic variants in the tyrosine kinase 2 (TYK2) gene, which is differentially associated with common autoimmune diseases, currently obscures the potential of TYK2 as a therapeutic target. We aimed to resolve this conflict by performing genetic meta-analysis across disorders; subsequent molecular, cellular, in vivo, and structural functional follow-up; and epidemiological studies. Our data revealed a protective homozygous effect that defined a signaling optimum between autoimmunity and immunodeficiency and identified TYK2 as a potential drug target for certain common autoimmune disorders.

Using PLINK for genome-wide association studies (GWAS) and data analysis

Within this chapter we introduce the basic PLINK functions for reading in data, applying quality control, and running association analyses. Three worked examples are provided to illustrate: data management and assessment of population substructure, association analysis of a quantitative trait, and qualitative or case-control association analyses.

Structural and regulatory diversity shape HLA-C protein expression levels.

Expression of HLA-C varies widely across individuals in an allele-specific manner. This variation in expression can influence efficacy of the immune response, as shown for infectious and autoimmune diseases. MicroRNA binding partially influences differential HLA-C expression, but the additional contributing factors have remained undetermined. Here we use functional and structural analyses to demonstrate that HLA-C expression is modulated not just at the RNA level, but also at the protein level. Specifically, we show that variation in exons 2 and 3, which encode the α1/α2 domains, drives differential expression of HLA-C allomorphs at the cell surface by influencing the structure of the peptide-binding cleft and the diversity of peptides bound by the HLA-C molecules. Together with a phylogenetic analysis, these results highlight the diversity and long-term balancing selection of regulatory factors that modulate HLA-C expression.