Fine-mapping the MHC locus in juvenile idiopathic arthritis (JIA) reveals genetic heterogeneity corresponding to distinct adult inflammatory arthritic diseases.

OBJECTIVES: Juvenile idiopathic arthritis (JIA) is a heterogeneous group of diseases, comprising seven categories. Genetic data could potentially be used to help redefine JIA categories and improve the current classification system. The human leucocyte antigen (HLA) region is strongly associated with JIA. Fine-mapping of the region was performed to look for similarities and differences in HLA associations between the JIA categories and define correspondences with adult inflammatory arthritides. METHODS: Dense genotype data from the HLA region, from the Immunochip array for 5043 JIA cases and 14 390 controls, were used to impute single-nucleotide polymorphisms, HLA classical alleles and amino acids. Bivariate analysis was performed to investigate genetic correlation between the JIA categories. Conditional analysis was used to identify additional effects within the region. Comparison of the findings with those in adult inflammatory arthritic diseases was performed. RESULTS: We identified category-specific associations and have demonstrated for the first time that rheumatoid factor (RF)-negative polyarticular JIA and oligoarticular JIA are genetically similar in their HLA associations. We also observe that each JIA category potentially has an adult counterpart. The RF-positive polyarthritis association at HLA-DRB1 amino acid at position 13 mirrors the association in adult seropositive rheumatoid arthritis (RA). Interestingly, the combined oligoarthritis and RF-negative polyarthritis dataset shares the same association with adult seronegative RA. CONCLUSIONS: The findings suggest the value of using genetic data in helping to classify the categories of this heterogeneous disease. Mapping JIA categories to adult counterparts could enable shared knowledge of disease pathogenesis and aetiology and facilitate transition from paediatric to adult services.

CD226 (DNAM-1) is associated with susceptibility to juvenile idiopathic arthritis.

OBJECTIVES: Juvenile idiopathic arthritis (JIA) is considered a complex genetic autoimmune disease. We investigated the association of genetic variants previously implicated in JIA, autoimmunity and/or immunoregulation, with susceptibility to JIA. METHODS: A genetic association study was performed in 639 JIA patients and 1613 healthy controls of northwest European descent. Ninety-three single nucleotide polymorphisms (SNP) were genotyped in a candidate gene approach. Results of the entire JIA patient group (all subtypes) were compared with results obtained, alternatively, with a clinically homogeneous patient group including only oligoarticular and rheumatoid factor (RF) negative polyarticular JIA patients (n=493). Meta-analyses were performed for all SNPs that have been typed in other Caucasian JIA cohorts before. RESULTS: SNPs in or near PTPN22, VTCN1, the IL2-IL21 region, ANKRD55 and TNFA were confirmed to be associated with JIA (p<0.05), strengthening the evidence for involvement of these genes in JIA. In the majority of these replicated SNPs, effect sizes were larger when analysing a homogeneous patient cohort than when analysing all subtypes. We identified two novel associations with oligoarticular and RF-negative polyarticular JIA: CD226 rs763361 (OR 1.30, 95% CI 1.12 to 1.51, p=0.0006) and CD28 rs1980422 (OR 1.29, 95% CI 1.07 to 1.55, p=0.008). Meta-analyses including reported studies confirmed the association of both SNPs with susceptibility to JIA (OR 1.16, p=0.001 and OR 1.18, p=0.001, for rs763361 and rs1980422, respectively). CONCLUSIONS: The CD226 gene has been identified as novel association with JIA, and a SNP near CD28 as a suggestive association. Both genes are probable candidate risk factors, since they are involved in costimulation of T cells.

Genome-wide data reveal novel genes for methotrexate response in a large cohort of juvenile idiopathic arthritis cases.

Clinical response to methotrexate (MTX) treatment for children with juvenile idiopathic arthritis (JIA) displays considerable heterogeneity. Currently, there are no reliable predictors to identify non-responders: earlier identification could lead to a targeted treatment. We genotyped 759 JIA cases from the UK, the Netherlands and Czech Republic. Clinical variables were measured at baseline and 6 months after start of the treatment. In Phase I analysis, samples were analysed for the association with MTX response using ordinal regression of ACR-pedi categories and linear regression of change in clinical variables, and identified 31 genetic regions (P<0.001). Phase II analysis increased SNP density in the most strongly associated regions, identifying 14 regions (P<1 × 10(-5)): three contain genes of particular biological interest (ZMIZ1, TGIF1 and CFTR). These data suggest a role for novel pathways in MTX response and further investigations within associated regions will help to reach our goal of predicting response to MTX in JIA.

Association of the CCR5 gene with juvenile idiopathic arthritis.

The CC chemokine receptor 5 (CCR5) has been shown to be important in the recruitment of T-helper cells to the synovium, where they accumulate, drive the inflammatory process and the consequent synovitis and joint destruction. A 32 base-pair insertion/deletion variant (CCR5Δ32) within the gene leads to a frame shift and a nonfunctional receptor. CCR5Δ32 has been investigated for its association with juvenile idiopathic arthritis (JIA), with conflicting results. The aim of this study was to investigate whether CCR5Δ32 is associated with JIA in an UK population. CCR5Δ32 was genotyped in JIA cases (n=1054) and healthy controls (n=3129) and genotype and allele frequencies were compared. A meta-analysis of our study combined with previously published studies was performed. CCR5Δ32 was significantly associated with protection from developing JIA, in this UK data set (P(trend)=0.006, odds ratio (OR) 0.79 95% confidence interval (95% CI): 0.66-0.94). The meta-analysis of all published case-control association studies confirmed the protective association with JIA (P=0.001 OR 0.82 95% CI: 0.73-0.93). CCR5Δ32 is a functional variant determining the number of receptors on the surface of T cells, and it is hypothesized that the level of CCR5 expression could influence the migration of proinflammatory T cells into the synovium and thus susceptibility to JIA.

Association of the AFF3 gene and IL2/IL21 gene region with juvenile idiopathic arthritis.

Recent genetic studies have led to identification of numerous loci that are associated with susceptibility to autoimmune diseases. The strategy of using information from these studies has facilitated the identification of novel juvenile idiopathic arthritis (JIA) susceptibility loci, specifically, PTPN22 and IL2RA. Several novel autoimmune susceptibility loci have recently been identified, and we hypothesise that single-nucleotide polymorphisms (SNPs) within these genes may also be JIA susceptibility loci. Five SNPs within the genes AFF3, IL2/IL21, IL7R, CTLA4 and CD226, previously associated with multiple autoimmune diseases were genotyped, in a large data set of Caucasian JIA patients and controls, and tested for association with JIA. We identified two susceptibility loci for JIA, AFF3 and the IL2/IL21 region and additional weak evidence supporting an association with the CTLA4 and IL7R genes, which warrant further investigation. All results require validation in independent JIA data sets. Further characterisation of the specific causal variants will be required before functional studies can be performed.

CD226 Gly307Ser association with multiple autoimmune diseases.

Genome-wide association studies provide insight into multigenic diseases through the identification of susceptibility genes and etiological pathways. In addition, the identification of shared variants among autoimmune disorders provides insight into common disease pathways. We previously reported an association of a nonsynonymous single nucleotide polymorphism (SNP) rs763361/Gly307Ser in the immune response gene CD226 on chromosome 18q22 with type 1 diabetes (T1D) susceptibility. Here, we report efforts toward identifying the causal variant by exonic resequencing and tag SNP mapping of the 18q22 region in both T1D and multiple sclerosis (MS). In addition to the analysis of newly available samples in T1D (2088 cases and 3289 controls) and autoimmune thyroid disease (AITD) (821 cases and 1920 controls), resulting in strong support for the Ser(307) association with T1D (P=3.46 x 10(-9)) and continued potential evidence for AITD (P=0.0345), we provide evidence for association of Gly307Ser with MS (P=4.20 x 10(-4)) and rheumatoid arthritis (RA) (P=0.017). The Ser(307) allele of rs763361 in exon 7 of CD226 predisposes to T1D, MS, and possibly AITD and RA, and based on the tag SNP analysis, could be the causal variant.

Autoinflammatory genes and susceptibility to psoriatic juvenile idiopathic arthritis.

OBJECTIVE: To investigate the association of NLRP3, NOD2, MEFV, and PSTPIP1, genes that cause 4 of the autoinflammatory hereditary periodic fever syndromes (HPFS), with juvenile idiopathic arthritis (JIA). METHODS: Fifty-one single-nucleotide polymorphisms (SNPs) across the 4 loci were investigated using MassArray genotyping in 950 Caucasian patients with JIA living in the UK and 728 ethnically matched healthy controls. RESULTS: Prior to Bonferroni correction for multiple testing, significant genotype associations between 6 SNPs in MEFV and JIA were observed and, in subgroup analysis, associations between 12 SNPs across all 4 loci and the subgroup of patients with psoriatic JIA were found. After Bonferroni correction for multiple testing, 2 genotype associations remained significant in the subgroup of patients with psoriatic JIA (MEFV SNP rs224204 [corrected P = 0.025] and NLRP3 SNP rs3806265 [corrected P = 0.04]). CONCLUSION: These findings support the use of monogenic loci as candidates for investigating the genetic component of complex disease and provide preliminary evidence of association between SNPs in autoinflammatory genes and psoriatic JIA. Our findings raise the interesting possibility of a shared disease mechanism between the HPFS and psoriatic JIA, potentially involving abnormal production of interleukin-1beta.

SNPs in the FOXP3 gene region show no association with Juvenile Idiopathic Arthritis in a UK Caucasian population.

OBJECTIVE: A region on the short arm of the X-chromosome, Xp11, has previously been linked to childhood-onset polyarthritis. Mapping to the linked region is FOXP3, a transcription factor that regulates regulatory T cell (T(reg)) development and function. The objective of this study was to determine whether single nucleotide polymorphisms (SNPs) in the FOXP3 gene region contribute to JIA susceptibility. METHOD: Nine FOXP3 SNPs were genotyped in 761 JIA cases and 402 controls using the Sequenom MassARRAY system. Association was measured using either chi(2) or Fisher's exact test at the allelic and genotypic level. Furthermore, cases and controls were stratified by gender and association measured for each stratum. RESULTS: None of the SNPs showed an association with JIA. Similarly, the lack of association was also evident in both the female and male cohorts. CONCLUSION: Although FOXP3 presents itself as a good candidate for contributing to JIA susceptibility, this study, which was powered to detect associations with genotypic relative risk >2 in the female cohort, has failed to find an association between SNPs in the FOXP3 gene region and JIA.

The type 1 diabetes susceptibility gene SUMO4 at IDDM5 is not associated with susceptibility to rheumatoid arthritis or juvenile idiopathic arthritis.

OBJECTIVES: Linkage and association of rheumatoid arthritis (RA) and rheumatoid factor (RF)-negative juvenile idiopathic arthritis (JIA) has previously been demonstrated to the type 1 diabetes (T1D) locus, IDDM5, on chromosome 6q25. An association of a methionine-to-valine polymorphism (rs237025, 163A --> G, M55V) in the SUMO4 gene within IDDM5 has recently been described in T1D. The objective of this study was to test the hypothesis that SUMO4 is a general autoimmune susceptibility gene by investigating whether the SUMO4 polymorphism is associated with RA and/or JIA. METHODS: The SUMO4 SNP was genotyped in 875 RA patients, 668 JIA patients and 484 healthy controls using a TaqMan allelic discrimination assay. Allele and genotype frequencies were compared between cases and controls using the chi2 test. Analyses were also carried out with RA patients stratified by gender, age at onset, RF status, the presence of erosive disease and shared epitope status, while JIA patients were stratified by their International League of Associations for Rheumatology (ILAR) subgroup. RESULTS: No deviation from Hardy-Weinberg equilibrium was detected in either set of cases or controls. No association was observed between rs237025 and RA (chi2 = 0.17, P = 0.93), or with any RA subset. Similarly, there was no association between this SNP and JIA (chi2 = 0.21, P = 0.90), or with any ILAR subgroup. CONCLUSIONS: The M55V substitution in the SUMO4 gene is not associated with susceptibility to RA or JIA in the UK population studied. However, other candidate genes mapping within IDDM5 remain to be investigated.

Re-audit of radiotherapy waiting times 2003.

AIMS: There has been increasing concern, by clinical oncologists in the UK, about the continuing long waiting times for radiotherapy treatment. An audit of patients starting treatment in 1 week in 2003 was carried out to determine waiting times and to compare current results with those obtained from a similar audit in 1998. MATERIALS AND METHODS: All centres in the UK completed the audit, which recorded waiting-list status and treatment intent for all new treatments started in 1 week, date of booking form completion and fractionation scheme used. RESULTS: Waiting times have lengthened in most centres for all categories of patients although, for palliative treatments, there was no additional increase in median waiting time compared with the last audit. Fewer patients in all categories are being treated within the Joint Collegiate Council for Oncology (JCCO) guidelines. Single-fraction treatments are now common for palliation. Most adjuvant treatment uses one of three prescriptions. In each Strategic Health Authority (StHA), the same proportion of the total number of patients with cancer seemed to be given radiotherapy. There was no obvious simple correlation of radiographer, physicist or treatment machine numbers with waiting times. CONCLUSION: The results of this survey suggest a continuing mismatch between capacity and demand. Increased complexity of radical treatments, and possibly more patients being referred for treatment, may have been offset in part by reduced fractionation for palliation.

Association of protein kinase C alpha (PRKCA) gene with multiple sclerosis in a UK population.

Twin, family and adoption studies suggest that susceptibility to multiple sclerosis is substantially mediated by genetic factors. Linkage to human chromosome 17q, homologous to a locus linked to experimental animal models of multiple sclerosis, has been widely replicated and the region likely to harbour a multiple sclerosis susceptibility gene has recently been refined to a 2.5 Mb region of 17q22-24. The candidate multiple sclerosis susceptibility gene, protein kinase C alpha (PRKCA), maps within this interval and association with 35 single-nucleotide polymorphism (SNP) markers, spanning the gene with a median spacing of 7.8 kb, was tested using a case-control approach. Single-marker genotype and estimated haplotype frequencies were compared in UK unrelated cases with multiple sclerosis (n = 184) and healthy controls (n = 340) in order to investigate association with susceptibility to disease. A haplotype of two SNPs mapping to the proximal region of the gene showed evidence for association with susceptibility (Bonferroni-corrected P value = 1.1 x 10(-5)). These findings suggest that further investigation of the PRKCA gene is warranted, particularly in cohorts with evidence of linkage to 17q22. Most of the SNPs investigated in this study were intronic and screening to identify disease-associated functional mutations is now required. Our results suggest that the promoter and proximal gene region should be not only included but prioritized in any screening strategy.

Polymorphisms in the tumour necrosis factor gene are not associated with severity of inflammatory polyarthritis.

BACKGROUND: Tumour necrosis factor alpha (TNFalpha) is a powerful inflammatory mediator in rheumatoid and other types of inflammatory arthritis. Polymorphisms within the TNFalpha gene have previously been investigated to determine their role in the aetiopathogenesis of rheumatoid arthritis (RA), but it is unclear whether reported associations are with susceptibility to, or severity of, disease. OBJECTIVE: To examine the association between both individual TNFalpha single nucleotide polymorphisms (SNPs) and haplotypes with the development and severity of erosions by 5 years in patients with inflammatory polyarthritis (IP). METHODS: 438 patients from the Norfolk Arthritis Register observational inception cohort of patients with IP were x rayed 5 years after disease onset. They were genotyped for nine SNPs mapping to the TNFalpha gene, using a SNaPshot primer extension assay. Haplotypes were constructed in patients with IP, who were compared for the presence and extent of erosions at 5 years. RESULTS: No association between individual TNFalpha SNPs or haplotypes in the patients who developed erosions at 5 years compared with those who remained non-erosive was found. Restricting analysis to patients who satisfied ACR criteria for RA by 5 years did not affect the conclusions. CONCLUSION: The TNFalpha gene does not seem to be associated with severity as assessed by erosive outcome at 5 years in patients with IP.