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Ankylosing spondylitis (AS) susceptibility is strongly correlated with genetic variation within the major histocompatibility complex (MHC), and the class I HLA-B*27 allele confers the major genetic risk factor to AS.1 Furthermore, strong evidence for additional alleles in the MHC has been observed which affect susceptibility independently from the HLA-B*27 allele, either by direct genotyping in candidate gene studies2 or through large-scale imputation-driven association studies.3 4 We have previously identified variants in both class I and II which affect susceptibility to AS through imputation of classical HLA alleles.4
The MICA gene is encoded 46 kbps from HLA-B and previous studies have reported genetic variants within this locus to affect susceptibility to AS and other inflammatory diseases.5 In particular, in Annals of Rheumatic Diseases, Zhou et al 6 evaluated the association of MICA variants with susceptibility to AS in cohorts from North America and China. This study found that the allele MICA*007:01 was strongly associated with risk of AS independently of HLA-B*27, with an association observed in the HLA-B27-negative cohort (US cohort OR=9.12, p=4.28×10–8; Chinese cohort OR=42.2, p=9.35×10–7). Weak protective associations with other MICA alleles were noted, likely because of the over-representation of the risk allele MICA*007:01.
Having previously observed extremely strong linkage disequilibrium (LD) between HLA-B and MICA single nucleotide polymorphisms (SNPs),1 we sought to independently test this report. We performed imputation of alleles in the MICA gene using a large reference panel of 1046 psoriatic arthritis individuals of European ancestry7 and assessed the evidence of association of MICA alleles controlling for susceptibility HLA-B alleles. We successfully imputed HLA-B and MICA alleles in 9429 AS cases and 13 459 controls of European ancestry from the IGAS cohort3 genotyped using the Illumina Immunochip SNP microarray.8 Within the MICA locus, we observed 14 distinct alleles where MICA*008:01 was observed to be the most common with an allele frequency of 50%. We performed association analysis at HLA-B and MICA imputed alleles with logistic regression controlling for the HLA-B*27 allele and 10 principal components from a genome-wide analysis to control for population structure.
In our study, we imputed the MICA*007 allele with high confidence (imputation performed in batches, all r2>0.85, mean r2=0.90) and genotypes were confirmed by Sanger sequencing of exons 2, 3, 4 and 5 of the MICA gene with perfect accuracy in 7 MICA*007 homozygote individuals and 5 non-carriers of the MICA*007 allele. We observed no association with MICA*007 after controlling for the effect of HLA-B*27 (p>0.05). We also observed that the HLA-B*27 and the MICA*007 alleles are in high LD (r2=0.66 in controls, r2=0.52 in cases, r2=0.53 in the reference imputation panel).
No association was observed between MICA*007 and AS in either HLA-B*27 negative subjects (1669 cases, 12 263 controls) (OR=1.32, p=0.07) or HLA-B*27 positive subjects (7760 cases, 1196 controls) (OR=1.04, p=0.34), and our study had over 80% power to detect an association with MICA*007 in both HLA-B*27 negative and positive analyses given the study sample size and the effect size previously reported for MICA*007 (OR=9). To assess whether heterogeneity of the different populations could have affected these findings, the analysis was repeated in the relatively homogenous UK population alone (n=4198 cases, 9611 controls). The findings were similar in HLA-B*27 positive subjects (p=0.89) and in HLA-B*27 negative subjects (p=0.53).
In this study, we found no evidence of association between the MICA*007 allele and AS susceptibility in a large cohort of European ancestry. Given the sample size in the HLA-B*27 negative cohort, we had 80% power to identify an independent effect of MICA*007 with an effect size of at least 1.55. We identified strong LD between HLA-B*27 and MICA*007 potentially explaining previously reported associations with this allele.
We thank all participating subjects with ankylosing spondylitis and healthy individuals who provided the DNA and clinical information necessary for this study.
Handling editor Josef S Smolen
Contributors All authors contributed to the design and/or acquisition and analysis of data. All authors drafted or reviewed the manuscript critically and gave final approval for the version submitted. All authors agree to be accountable for all aspects of the submitted work.
Funding This research received no specific grant from any funding agency in the public, commercial or not-for-profit sectors.
Competing interests None declared.
Patient consent Not required.
Provenance and peer review Not commissioned; externally peer reviewed.
Data sharing statement Data will be made available on reasonable request.
Collaborators International Genetics of Ankylosing Spondylitis (IGAS) Consortium: Paul Bowness, Paul Wordsworth: NIHR Oxford Musculoskeletal Biomedical Research Unit, Nuffield Orthopaedic Centre, Headington, Oxford, UK; Maxime Breban: INSERM UMR 1173, Université de Versailles Saint Quentin en Yvelines, Laboratoire d’excellence Inflamex, Saint-Quentn-En-Yvelines, France; Matthew Brown: School of Biomedical Sciences, Queensland University of Technology, Brisbane, Australia; Robert Colbert: National Institute of Arthritis and Musculoskeletal and Skin Diseases, NIH, Bethesda, Maryland, USA; Adrian Cortes: Nuffield Department of Clinical Neurosciences, University of Oxford, Oxford, UK; Bart Crusius: Department of Medical Microbiology and Infection Control, Laboratory of Immunogenetics, VU University Medical Center, Amsterdam, The Netherlands; Jing Cui, Soumya Raychaudhuri: Department of Medicine, Brigham and Women’s Hospital, Boston, MA, USA; Dirk Elewaut: Department of Rheumatology, Gent University Hospital, Gent, Belgium; David Evans: University of Queensland Diamantina Institute, Princess Alexandra Hospital, Brisbane, Australia; Øystein Førre: Department of Rheumatology, Oslo University Hospital, and University of Oslo, Oslo, Norway; Dafna Gladman: Division of Rheumatology, University of Toronto, Toronto, Canada; Nigil Haroon, Robert Inman: Division of Rheumatology, Toronto Western Hospital, University of Toronto, Toronto, Canada; Benedicte Lie: Department of Medical Genetics, University of Oslo and Oslo University Hospital, Oslo, Norway; Carlos Lopez-Larrea: Department of Immunology, Hospital Universitario Central de Asturias, Oviedo, Spain; Walter Maksymowych: Department of Medicine, University of Alberta, Alberta, Canada; Javier Martin: Instituto de Parasitología y Biomedicina López-Neyra, Consejo Superior de Investigaciones-Científicas, Granada, Spain; Hans Nossent: School of Medicine, University of Western Australia, Perth, Australia Proton Rahman: Memorial University of Newfoundland, Newfoundland, Canada; John Reveille: Department of Rheumatology and Clinical Immunogenetics, University of Texas Health Science Center at Houston, Houston, Texas, USA; Fernando Santos: Chronic Diseases Research Centre (CEDOC), Faculdade de Ciências Médicas, Universidade Nova de Lisboa, Lisboa, Portugal; Simon Stebbings: Department of Medicine, Dunedin School of Medicine, University of Otago, Dunedin, New Zealand; Jaakko Tuomilehto: Department of Chronic Disease Prevention, National Institute for Health and Welfare, Helsinki, Finland; Rafael Valle-Oñate: Spondyloarthropaty Group-Division of Rheumatology, Hospital Militar Central/Universidad de La Sabana, Bogotá, NA, Colombia; Irene van der Horst-Bruinsma: Department of Rheumatology, VU University Medical Centre, Amsterdam, Netherlands; Michael Weisman: Department of Medicine/Rheumatology, Cedars-Sinai Medical Center, Los Angeles, California, USA.
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