José Ramón Lamas

The Peptide Repertoires of HLA-B27 Subtypes Differentially Associated to Spondyloarthropathy (B*2704 and B*2706) Differ by Specific Changes at Three Anchor Positions

HLA-B*2704 is strongly associated with ankylosing spondylitis. B*2706, which differs from B*2704 by two amino acid changes, is not associated with this disease. A systematic comparison of the B*2704- and B*2706-bound peptide repertoires was carried out to elucidate their overlap and differential features and to correlate them with disease susceptibility. Both subtypes shared about 90% of their peptide repertoires, consisting of peptides with Arg2and C-terminal aliphatic or Phe residues. B*2706 polymorphism influenced specificity at three anchor positions: it favored basic residues at P3 and PΩ-2 and impaired binding of Tyr and Arg at PΩ. Thus, the main structural feature of peptides differentially bound to B*2704 was the presence of C-terminal Tyr or Arg, together with a strong preference for aliphatic/aromatic P3 residues. This is the only known feature of B*2704 and B*2706 that correlates to their differential association with spondyloarthropathy. The concomitant presence of basic P3 and PΩ-2 residues was observed only among peptides differentially bound to B*2706, suggesting that it impairs binding to B*2704. Similarity between peptide overlap and the degree of cross-reaction with alloreactive T lymphocytes suggested that the majority of shared ligands maintain unaltered antigenic features in the context of both subtypes.

Rheumatoid Arthritis Does Not Share Most of the Newly Identified Systemic Lupus Erythematosus Genetic Factors

OBJECTIVE Rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) share some genetic factors such as HLA, PTPN22, STAT4, and 6q23. The aim of this study was to determine whether 9 other SLE genetic factors are also implicated in RA susceptibility. METHODS A characteristic single-nucleotide polymorphism (SNP) in each of 9 genetic factors, ITGAM (rs1143679), C8orf13-BLK (rs13277113), TYK2 (rs2304256), 1q25.1 (rs10798269), PXK (rs6445975), KIAA1542 (rs4963128), MECP2 (rs17435), BANK1 (rs17266594), and LY9 (rs509749), was studied in 1,635 patients with RA and 1,906 control subjects from Spain. The rs7574865 SNP in STAT4 was also included. Analyses were conducted globally and after stratification by sex and clinical features (anti-cyclic citrullinated peptide and rheumatoid factor, shared epitope, rheumatoid nodules, radiographic changes, sicca syndrome, and pneumonitis). RESULTS No association was observed between RA and any of the 9 newly identified SLE genetic factors. A meta-analysis using previous data was consistent with these results. In addition, there were no significant differences between individuals with and those without each of the clinical features analyzed, except the frequency of the minor allele in the C8orf13-BLK locus that was decreased in patients with sicca syndrome (14.6% versus 22.4% in controls; P = 0.003). CONCLUSION None of the 9 recently identified SLE risk factors showed association with RA. Therefore, common genetic factors affecting the pathogenesis of these 2 disorders seem to be limited, revealing that the genetic component contributes to the different expression of these diseases.

Chromosomal region 16p13: further evidence of increased predisposition to immune diseases

Objective: Genome-wide studies have identified the chromosomal region 16p13 in the susceptibility to type 1 diabetes (T1D) and multiple sclerosis (MS). This region includes the CLEC16A/KIAA0350 gene and an adjacent gene, MHC2TA (MHC class II transactivator), previously associated with susceptibility to MS and rheumatoid arthritis (RA). The role of CLEC16A polymorphisms in the pathogenesis of T1D, MS and RA and its relationship with the association reported with a MHC2TA haplotype were investigated. Methods: CLEC16A (rs2903692/rs6498169/rs11074956) polymorphisms were analysed in 435 patients with MS, 316 with T1D and 600 with RA and in 550 ethnically matched controls. The MHC2TA rs3087456G/rs4774C risk haplotype was studied in an independent RA cohort. Results: rs2903692 conferred a protective effect on patients with T1D, MS and RA. The described association of rs6498169 with MS was replicated in MS and RA cohorts. The effect of the MHC2TA rs3087456G/rs4774C haplotype on RA susceptibility was confirmed, and the haplotype was found to be in negative linkage disequilibrium with the CLEC16A rs2903692A/rs6498169A haplotype. Conclusions: Associations of CLEC16A polymorphisms with T1D and MS were successfully replicated in a Spanish population. A novel association of rs6498169 with a predisposition to RA was described which is consistent with previous MHC2TA results. These data provide evidence for the influence of variants within this chromosomal region on the development of complex diseases.

Modulation at multiple anchor positions of the peptide specificity of HLA–B27 subtypes differentially associated with ankylosing spondylitis

OBJECTIVE To investigate the rules governing peptide binding to HLA-B*2705, and to B*2704 and B*2706, which are 2 subtypes differentially associated with ankylosing spondylitis. METHODS Poly-Ala analogs carrying the HLA-B27 motif Arg-2, and substitutions at anchor positions P1, P3, or Pomega, were used to determine a binding score for each residue at each position. Binding was assessed in a quantitative epitope stabilization assay, where the cell surface expression of HLA-B27 was measured by flow cytometry as a function of peptide concentration. RESULTS Peptide anchor residues contributed additively to B*2705 binding. About 15% of the natural B*2705 ligands used a deficient P3 or Pomega anchor, but never both, indicating that detrimental anchoring at one of these positions is always compensated by a good anchor at the other one. About 50% of the B*2705 ligands used suboptimal P1 residues. However, this was compensated with optimal P3 and/or Pomega anchoring. Peptides that were longer than decamers used good anchor residues at the 3 positions, suggesting more stringent binding requirements. B*2704 and B*2706 differed in their residue specificity at P1, P3, and Pomega. The rules derived for B*2705 also applied to the known ligands of these 2 subtypes. CONCLUSION The B*2705, B*2704, and B*2706 peptide repertoires are limited by the allowed residue combinations described in this study. The differential association of B*2704 and B*2706 with spondylarthropathy correlates with differences in their peptide specificity at multiple anchor positions. However, it is now possible to predict the peptide features that determine this differential binding to both subtypes.

Fine specificity of antigen binding to two class I major histocompatibility proteins (B*2705 and B*2703) differing in a single amino acid residue.

Starting from the X-ray structure of a class I majorhistocompatibility complex (MHC)-encoded protein (HLA-B*2705), a naturallypresented self-nonapeptide and two synthetic analogues were simulated in thebinding groove of two human leukocyte antigen (HLA) alleles (B*2703 andB*2705) differing in a single amino acid residue. After 200 ps moleculardynamics simulations of the solvated HLA–peptide pairs, some molecularproperties of the complexes (distances between ligand and protein center ofmasses, atomic fluctuations, buried versus accessible surface areas,hydrogen-bond frequencies) allow a clear discrimination of potent from weakMHC binders. The binding specificity of the three nonapeptides for the twoHLA alleles could be explained by the disruption of one hydrogen-bondingnetwork in the binding pocket of the HLA-B*2705 protein where the singlemutation occurs. Rearrangements of interactions in the B pocket, which bindsthe side chain of peptidic residue 2, and a weakening of interactionsinvolving the C-terminal end of the peptide also took place. In addition,extension of the peptide backbone using a β-Ala analogue did notabolish binding to any of the two HLA-B27 subtypes, but increased theselectivity for B*2703, as expected from the larger peptide binding groovein this subtype. A better understanding of the atomic details involved inpeptide selection by closely related HLA alleles is of crucial importancefor unraveling the molecular features linking particular HLA alleles toautoimmune diseases, and for the identification of antigenic peptidestriggering such pathologies.

Association of the IFIH1-GCA-KCNH7 chromosomal region with rheumatoid arthritis

The cause of rheumatoid arthritis (RA) remains unknown, although both genetic and environmental factors are involved. A genome-wide association study of non-synonymous single nucleotide polymorphisms (SNPs) showed the involvement of rs1990760, a change Ala946Thr in the IFIH1 interferon induced helicase gene, in type 1 diabetes risk.1 Further analyses of the locus surrounding this marker obtained compelling statistical support of the genetic equivalence of some other SNPs in strong linkage disequilibrium with rs1990760, making it impossible to ascertain the aetiological variant. The associated chromosomal region in 2q24.3 includes three genes: IFIH1 , also known as helicard or melanoma differentiation associated gene-5 ( mda-5 )2; grancalcin ( GCA ); and a potassium voltage gated channel ( KCNH7 ). We analysed three variants along this region to investigate whether this locus is involved in RA, another autoimmune disease. We replicated the effect …

Large-scale gene expression in bone marrow mesenchymal stem cells: a putative role for COL10A1 in osteoarthritis

Objectives To elucidate disease-specific molecular changes occurring in osteoarthritis (OA) by analysing the differential gene expression profiles of bone marrow mesenchymal stem cells (BM-MSCs) from patients with OA compared with those without OA. Methods Expression profiles of BM-MSCs from eight paired patients with OA and patients with hip fracture without signs of OA were compared by DNA microarray expression analysis and significant differences were evaluated by computational Gene Set Enrichment Analysis. To validate the involvement of COL10A1 as part of the most downregulated gene set in OA, three tagging single nucleotide polymorphisms were genotyped in 191 patients with OA and 283 controls. COL10A1 expression was also assessed by quantitative RT-PCR in additional subjects. Results Expression levels in 9% (1967) of the overall transcripts were significantly different (p<0.05) between MSCs from patients with OA and controls (532 genes reached twofold differences: 240 were upregulated and 292 were downregulated). Cell development and differentiation were the functional categories accounting for most genes with altered expression. Interestingly, several genes related to the Wnt/-catenin pathway and collagen genes were downregulated in MSCs from patients with OA. The collagen gene set was clearly downregulated in OA. Furthermore, the expression of COL10A1 was significantly reduced in patients with OA. A genetic association between the COL10A1 rs11965969 polymorphism and OA was also found. Conclusion COL10A1 downregulation seems to have a role in the establishment of a defective and/or unstable subchondral cartilage matrix in OA disease. It is proposed that OA may be linked to the intrinsic defective regenerative potential of BM-MSCs resulting from its reduced expression of fate commitment-related genes.

Role of the human endogenous retrovirus HERV-K18 in autoimmune disease susceptibility: study in the Spanish population and meta-analysis.

Background Human endogenous retroviruses (HERVs) are genomic sequences that resulted from ancestral germ-line infections by exogenous retroviruses and therefore are transmitted in a Mendelian fashion. Increased HERV expression and antibodies to HERV antigens have been found in various autoimmune diseases. HERV-K18 in chromosome 1 was previously associated with type one diabetes and multiple sclerosis (MS). The etiology of these complex conditions has not been completely elucidated even after the powerful genome wide association studies (GWAS) performed. Nonetheless, this approach does not scrutinize the repetitive sequences within the genome, and part of the missing heritability could lie behind these sequences. We aimed at evaluating the role of HERV-K18 in chromosome 1 on autoimmune disease susceptibility. Methods Two HERV-K18 SNPs (97Y/C and 154W/Stop substitutions) conforming three haplotypes were genotyped in Spanish cohorts of multiple sclerosis (n = 942), rheumatoid arthritis (n = 462) and ethnically matched controls (n = 601). Our findings were pooled in a meta-analysis including 5312 autoimmune patients and 4032 controls. Results Significant associations of both HERV-K18 polymorphisms in chromosome 1 with MS patients stratified by HLA-DRB1*15∶01 were observed [97Y/C p = 0.02; OR (95% CI) = 1.5 (1.04–2.17) and 154W/Stop: p = 0.001; OR (95% CI) = 1.6 (1.19–2.16)]. Combined meta-analysis of the previously published association studies of HERV-K18 with different autoimmune diseases, together with data derived from Spanish cohorts, yielded a significant association of the HERV-K18.3 haplotype [97Y–154W: pM-H = 0.0008; ORM-H (95% CI) = 1.22 (1.09–1.38)]. Conclusion Association of the HERV-K18.3 haplotype in chromosome 1 with autoimmune-disease susceptibility was confirmed through meta-analysis.

Orthopedic Surgery in Rheumatoid Arthritis in the Era of Biologic Therapy

Objective. To analyze sociodemographic and clinic-related factors associated with the use of orthopedic surgical procedures in rheumatoid arthritis (RA), focusing on the potential role of new biologic therapies. Methods. A retrospective medical record review was performed in a probability sample of 1272 patients with RA from 47 units distributed in 19 Spanish regions. Sociodemographic and clinical features, use of drugs, and arthritis-related joint surgeries were recorded following a standardized protocol. Results. A total of 94 patients (7.4%) underwent any orthopedic surgery during their disease course, with a total of 114 surgeries; 47 (41.2%) of these surgeries were total joint replacement (TJR). The median time to first orthopedic procedure was 7.9 years from the onset of RA symptoms, and the rate of orthopedic surgery (excluding TJR) was 4.5 procedures per 100 person-years from the beginning of RA, while the rate of TJR was 2.25 interventions per 100 person-years. A higher risk of undergoing an orthopedic surgical procedure was associated with taking nonsteroidal antiinflammatory drugs (NSAID) in the previous 2 years, female sex, longterm disease, and the presence of extraarticular complications. The risk factors for undergoing a TJR were being old, having a longterm disease, and taking biologic therapies. Conclusion. In the era of biologics, our national audit found a low percentage of patients who underwent orthopedic surgery, probably reflecting a thorough management of the RA. Sociodemographic factors, longterm RA, extraarticular complications, and NSAID were associated with orthopedic surgery.

Influence of IL6R rs8192284 Polymorphism Status in Disease Activity in Rheumatoid Arthritis

Objective. To analyze the influence of IL6R rs8192284 polymorphism on the disease activity of rheumatoid arthritis (RA). Methods. Patients with RA (n = 281) were followed for a median of 4.2 years. A total of 1143 disease activity measurements using the 28-joint count Disease Activity Score (DAS28) were performed. A mixed-effect model was used to analyze the measurements. Results. A statistically significant interaction was observed between IL6R rs8192284 polymorphism and the presence of anticyclic citrullinated peptide (anti-CCP) antibodies (p = 0.008). An inverse relationship between the polymorphism and DAS28 was observed depending on anti-CCP status. Conclusion. The anti-CCP status in patients with RA determines the association between the IL6R rs8192284 polymorphism and disease activity.