ACPA-positive Rheumatoid Arthritis Patients Research Articles

Anti-modified protein antibodies can be detected in saliva, but not in intestinal secretions of seropositive rheumatoid arthritis patients - evidence of site-specific mucosal autoantibody secretion in RA.

Anti-citrullinated protein antibodies (ACPA) have been detected in sputum and saliva, indicating that anti-modified protein antibodies (AMPA) can be produced at mucosal sites in rheumatoid arthritis (RA) patients. However, the body's largest mucosal compartment, the gut, has not yet been examined. We therefore investigated the presence of several AMPA (antibodies against citrullinated (ACPA), carbamylated (anti-CarP) and acetylated (AAPA) proteins) at different mucosal sites, including the intestinal tract. Paired fecal/ileal wash, saliva and serum samples of RA patients and healthy volunteers were collected in two independent cohorts. Data involving feces was replicated in a third cohort. In these secretions AMPA were analyzed using in-house ELISA with unmodified peptides as control. In fecal samples total IgA and anti-E. coli IgA were measured. ACPA, anti-CarP and AAPA IgA were measurable in saliva of seropositive RA patients (prevalence 9-40%). No AMPA could be detected in feces. IgA was present since total IgA and anti-E. coli IgA was detectable in feces of ACPA-positive RA patients and healthy donors. Results were confirmed in another cohort using colonoscopically collected ileal wash samples. Our study shows the presence of ACPA, anti-CarP and AAPA IgA in saliva of ACPA-seropositive RA patients. However, no AMPA could be detected in feces/ileal wash samples of these patients, although our assays were able to measure other antigen-specific antibodies. These data suggest that mucosal autoantibody secretion may occur in the oral mucosa of RA patients, while no evidence could be found for this process in the lower gastro-intestinal tract.

Epigenome-wide methylation haplotype association analysis identified HLA-DRB1, HLA-DRB5 and HLA-DQB1 as risk factors for rheumatoid arthritis.

The aim of this study was to compare nonrandom associations between physically adjacent single methylation polymorphism loci among rheumatoid arthritis (RA) and normal subjects for investigating RA-risk methylation haplotypes (meplotype). With 354 ACPA-positive RA patients and 335 normal controls selected from a case-control study based on Swedish population, we conducted the first RA epigenome-wide meplotype association study using our software EWAS2.0, mainly including (i) converted the β value to methylation genotype (menotype) data, (ii) identified methylation disequilibrium (MD) block, (iii) calculated frequent of each meplotypes in MD block and performed case-control association test and (iv) screened for RA-risk meplotypes by odd ratio (OR) and p-values. Ultimately, 545 meplotypes on 334 MD blocks were identified significantly associated with RA (p-value<.05). These meplotypes were mapped to 329 candidate genes related to RA. Subsequently, combined with gene optimization, eight RA-risk meplotypes were identified on three risk genes: HLA-DRB1, HLA-DRB5 and HLA-DQB1. Our results reported the relationship between DNA methylation pattern on HLA-DQB1 and the risk of RA for the first time, demonstrating the co-demethylation of 'cg22984282' and 'cg13423887' on HLA-DQB1 gene (meplotype UU, p-value=2.90E-6, OR=1.68, 95% CI=[1.35, 2.10]) may increase the risk of RA. Our results demonstrates the potential of methylation haplotype analysis to identify RA-related genes from a new perspective and its applicability to the study of other disease.

Potential alleviation of bone mineral density loss with Janus kinase inhibitors in rheumatoid arthritis.

Rheumatoid arthritis (RA) is characterized by localized bone loss, general osteoporosis and increased fracture risks. Tumour necrosis factor inhibitors (TNFi), non-tumour necrosis factor inhibitors (non-TNFi) biologic, Janus kinase inhibitors (JAKi) had shown the suppression effects to osteoclast activation and improvement of bone mineral density (BMD). Anti-cyclic citrullinated peptide antibody (ACPA) is associated with osteoclast activation and the resultant bone loss. However, few studies have compared BMD changes among patients with RA treated with targeted therapies that have different mechanisms of action. This retrospective study recruited patients with RA who had undergone BMD testing twice. Changes in the BMD were compared using the generalized estimating equation (GEE) in treatment groups that received conventional synthetic disease-modifying anti-rheumatic drugs (csDMARDs), TNFi, non-TNFi biologics, and JAKi. In total, 362 patients with RA were enrolled (csDMARDs, n = 153, TNFi, n = 71, non-TNFi biologics, n = 108, JAKi, n = 30). We observed greater changes in femoral BMD (left, 0.06, 95% CI 0.01-0.12, p = 0.016; right, 0.09, 95% CI 0.04-0.15, p = 0.001 by GEE) following JAKi treatment as compared with other treatments. Compared to the ACPA-negative group, patients with ACPA positivity exhibited greater improvement in the femoral BMD (left, 0.09, 95% CI 0.02-0.15, p = 0.008; right, 0.11, 95% CI 0.05-0.18, p = 0.001). Compared to other targeted therapies, JAKi might exert a more potent effect to prevent BMD loss, specifically in ACPA-positive patients with RA, and could be a potential therapeutic option to mitigate generalized bone loss. Key Points •JAKi therapy inhibits systemic bone loss in patients with RA. •ACPA-positive RA patients exhibited a greater BMD improvement than ACPA-negative RA patients. •JAKi might more potently prevent BMD decline than conventional synthetic or biological DMARDs.

Autoantibodies against complement factor B in rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disorder affecting the joints. Many patients carry anti-citrullinated protein autoantibodies (ACPA). Overactivation of the complement system seems to be part of the pathogenesis of RA, and autoantibodies against the pathway initiators C1q and MBL, and the regulator of the complement alternative pathway, factor H (FH), were previously reported. Our aim was to analyze the presence and role of autoantibodies against complement proteins in a Hungarian RA cohort. To this end, serum samples of 97 ACPA-positive RA patients and 117 healthy controls were analyzed for autoantibodies against FH, factor B (FB), C3b, C3-convertase (C3bBbP), C1q, MBL and factor I. In this cohort, we did not detect any patient with FH autoantibodies but detected C1q autoantibodies in four patients, MBL autoantibodies in two patients and FB autoantibodies in five patients. Since the latter autoantibodies were previously reported in patients with kidney diseases but not in RA, we set out to further characterize such FB autoantibodies. The isotypes of the analyzed autoantibodies were IgG2, IgG3, IgGκ, IgGλ and their binding site was localized in the Bb part of FB. We detected in vivo formed FB-autoanti-FB complexes by Western blot. The effect of the autoantibodies on the formation, activity and FH-mediated decay of the C3 convertase in solid phase convertase assays was determined. In order to investigate the effect of the autoantibodies on complement functions, hemolysis assays and fluid phase complement activation assays were performed. The autoantibodies partially inhibited the complement-mediated hemolysis of rabbit red blood cells, inhibited the activity of the solid phase C3-convertase and C3 and C5b-9 deposition on complement activating surfaces. In summary, in ACPA-positive RA patients we identified FB autoantibodies. The characterized FB autoantibodies did not enhance complement activation, rather, they had inhibitory effect on complement. These results support the involvement of the complement system in the pathomechanism of RA and raise the possibility that protective autoantibodies may be generated in some patients against the alternative pathway C3 convertase. However, further analyses are needed to assess the exact role of such autoantibodies.

Genetic variant in SPAG16 is associated with the susceptibility of ACPA-positive rheumatoid arthritis possibly via regulation of MMP-3

ObjectivesIn two previously published genome-wide association studies, a cluster of variants of sperm-associated antigen16 (SPAG16) were reported to be associated with the radiological progression rate of ACPA-positive rheumatoid arthritis (RA) patients from North American and Southern European ancestry. In this study, we aimed to investigate whether the reported RA-risk loci in SPAG16 are associated with the disease in the Chinese population and to further validate the functional role of the susceptible locus in RA tissues.MethodsA total of 500 ACPA-positive RA patients and 1000 age-matched healthy subjects were recruited. Two SNPs of SPAG16, including rs7607479 (C/T) and rs6435818 (A/C), were genotyped, and the genotyping data were compared with chi-square test. Gene expression analysis was performed in synovial tissues obtained from 40 RA patients and 30 non-RA controls surgically treated for bone fracture. The tissue expression of SPAG16 and matrix metalloproteinase 3 (MMP-3) was compared between the two groups by the Student’s t test. The relationship between serum indexes and mRNA expression of SPAG16 and MMP-3 were evaluated by Spearman’s correlation analysis.ResultFor rs7607479, the frequency of genotype TT was significantly higher in RA patients than in the controls (49.0% vs. 40.4%, p = 0.002). The RA patients were found to have significantly lower frequency of allele C than the controls (30.9% vs. 36.8%, p = 0.001). As for rs6435818, there was no significant difference of genotype or allele frequency between the two groups. The mRNA expression of MMP-3 was 1.63-fold higher in the RA patients than in the controls (p < 0.001). The expression of SPAG16 was comparable between the two groups (p = 0.43). The mRNA expression of MMP-3 was 1.39-fold higher in patients with genotype TT than in the patients with genotype CC (p = 0.006). The mRNA expression level of MMP-3 was significantly correlated with serum rheumatoid factor (r = 0.498, p < 0.001) and C-reactive protein (r = 0.272, p = 0.01), weakly correlated with erythrocyte sedimentation rate (r = 0.236, p = 0.09).ConclusionsWe validated a common genetic risk factor in ACPA-positive patients with RA, which is associated with the tissue production of MMP-3 and disease progression. Further functional analysis into the role of rs7607479 in MMP-3 expression can shed new light on the genetic architecture of ACPA-positive RA.

Efficacy of Abatacept Versus Tumor Necrosis Factor Inhibitors in Anti-citrullinated Protein Antibody-Positive Patients with Rheumatoid Arthritis: Results from a Korean Nationwide Biologics Registry.

Introduction To compare the efficacy of abatacept and tumor necrosis factor inhibitor (TNFi) in patients with anti-citrullinated protein antibody (ACPA)-positive rheumatoid arthritis (RA) and identify those who benefit most from abatacept over TNFi.MethodsThis observational study identified RA patients who were ACPA-positive and initiated abatacept or TNFi from the Korean College of Rheumatology Biologics and Targeted therapy registry. Propensity score (PS) matching was performed to balance baseline confounding in abatacept- or TNFi-treated patients. The major endpoints were changes in Clinical Disease Activity Index (CDAI) and achievement of CDAI remission/low disease activity after 1 year of treatment. Subgroup analysis was mainly performed stratified by prior biologics use.ResultsA total of 291 PS-matched, ACPA-positive RA patients who initiated abatacept (n = 97) and TNFi (n = 194) were included. From baseline CDAI scores of 26.52 in the abatacept group and 26.38 in the TNFi group, the mean changes after 1 year were − 16.78 and − 13.61, respectively (difference − 3.17, p = 0.020). The proportion of patients achieving CDAI remission/low disease activity was 68.0% with abatacept and 52.6% with TNFi (p = 0.013). In the subgroup analysis, patients that were biologics-naïve had better improvement in CDAI after treatment with abatacept than TNFi (difference − 3.35, p = 0.021).ConclusionsThis real-world study suggests that abatacept may have better clinical response compared to TNFi in patients with established ACPA-positive RA, especially in those that were biologics-naïve.Supplementary InformationThe online version contains supplementary material available at 10.1007/s40744-022-00467-4.

Personalized Therapeutic Strategies in the Management of Osteoporosis in Patients with Autoantibody-Positive Rheumatoid Arthritis.

Bone mineral density (BMD) reduction and fragility fractures still represent a major source of morbidity in rheumatoid arthritis (RA) patients, despite adequate control of the disease. An increasing number of clinical and experimental evidence supports the role of autoantibodies, especially anti-citrullinated protein antibodies (ACPAs), in causing localized and generalised bone loss in ways that are both dependent on and independent of inflammation and disease activity. The human receptor activator of nuclear factor kappa B and its ligand—the so-called RANK-RANKL pathway—is known to play a key role in promoting osteoclasts’ activation and bone depletion, and RANKL levels were shown to be higher in ACPA-positive early untreated RA patients. Thus, ACPA-positivity can be considered a specific risk factor for systemic and periarticular bone loss. Through the inhibition of the RANK-RANKL system, denosumab is the only antiresorptive drug currently available that exhibits both a systemic anti-osteoporotic activity and a disease-modifying effect when combined with conventional synthetic or biologic disease-modifying anti-rheumatic drugs (DMARDs). Thus, the combination of DMARD and anti-RANKL therapy could be beneficial in the prevention of fragility fractures and structural damage in the subset of RA patients at risk of radiographic progression, as in the presence of ACPAs.

Exhaled nitric oxide in early rheumatoid arthritis and effects of methotrexate treatment

Patients with established rheumatoid arthritis (RA) and disease modifying treatments have lower nitric oxide (NO) levels in the alveolar compartment (CANO) and in the airway wall (CawNO), but also higher diffusion capacities for NO in the airways (DawNO) compared to matched controls. The aim of the present study was to investigate the NO lung dynamics in patients with recent onset RA before and after immune suppression with methotrexate therapy. Patients with early RA and antibodies against anticitrullinated peptides (ACPA) were recruited. Measurement of exhaled NO and inflammatory markers in serum were performed. Clinical disease activity was evaluated with Disease Activity Score for 28 joints. Healthy individuals were used as matched controls. Data are presented as median (lower quartile, upper quartile) values. RA patients (n = 44) had lower exhaled NO (FENO50) 16 (10–24) ppb compared to controls 21 (15, 29) ppb, p = 0.013. In NO-dynamics, CANO was lower in RA patients 1.6 (1.0, 2.2) ppb compared to the control subjects 2.3 (1.3, 3.1) ppb, p = 0.007. CawNO was also lower in the RA patients 55 (24, 106) ppb compared to control subjects 124 (110, 170) ppb, p < 0.001, but DawNO was higher 17 (8, 30) mL/s and 9 (5, 11) mL/s respectively, p < 0.001. Methotrexate treatment for three months reduced disease activity, but did not change the NO dynamics. In conclusion, the altered NO dynamics of the lung in ACPA-positive RA patients are already present in the early stages of the disease before any treatments and do not change after methotrexate therapy suggesting a role in the pathogenesis.

Serum Antigenome Profiling Reveals Diagnostic Models for Rheumatoid Arthritis

ObjectiveThe study aimed to investigate the serum antigenomic profiling in rheumatoid arthritis (RA) and determine potential diagnostic biomarkers using label-free proteomic technology implemented with machine-learning algorithm.MethodSerum antigens were captured from a cohort consisting of 60 RA patients (45 ACPA-positive RA patients and 15 ACPA-negative RA patients), together with sex- and age-matched 30 osteoarthritis (OA) patients and 30 healthy controls. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was then performed. The significantly upregulated and downregulated proteins with fold change > 1.5 (p < 0.05) were selected. Based on these differentially expressed proteins (DEPs), a machine learning model was trained and validated to classify RA, ACPA-positive RA, and ACPA-negative RA.ResultsWe identified 62, 71, and 49 DEPs in RA, ACPA-positive RA, and ACPA-negative RA, respectively, as compared to OA and healthy controls. Typical pathway enrichment and protein–protein interaction networks were shown among these DEPs. Three panels were constructed to classify RA, ACPA-positive RA, and ACPA-negative RA using random forest models algorithm based on the molecular signature of DEPs, whose area under curve (AUC) were calculated as 0.9949 (95% CI = 0.9792–1), 0.9913 (95% CI = 0.9653–1), and 1.0 (95% CI = 1–1).ConclusionThis study illustrated the serum auto-antigen profiling of RA. Among them, three panels of antigens were identified as diagnostic biomarkers to classify RA, ACPA-positive, and ACPA-negative RA patients.

Внеклеточные ловушки нейтрофилов: динамика образования, ассоциированная с переходом от ремиссии к активному ревматоидному артриту

Tne aim of the research. To evaluate the main parameters of neutrophil extracellular traps formation (NETosis) and their dynamics in rheumatoid arthritis (RA) patients associated with the transition from remission to active autoimmune infl ammation. Material and methods. A total of 37 patients (6 males and 31 females) with verifi ed RA according to the ACR/EULAR 2010 criteria were enrolled in the study and formed the main group. Th e comparison group was constituted by 30 healthy volunteers. Circulating neutrophils were isolated with one-step density gradient centrifugation using double layers of iohexol gradient in the original modifi cation. Th e quantitative composition of neutrophils was assessed through microscopy of smears stained according to May-Grunwald. Cell viability as well as non-specifi c activation were evaluated microscopically using trypan blue exclusion assay and nitroblue tetrazolium test, respectively. Neutrophil extracellular trap (NETs) formation was induced by 50 nM phorbol-12-myristate-13- acetate and assessed using fl uorescence microscopy. Results. Neutrophils isolated from RA patients in clinical remission have been revealed to have a higher ability for spontaneous and induced NETs formation than neutrophils from the comparison group. Th e transition from remission to active RA was characterised by a signifi cant increase in induced and, especially, spontaneous formation of NETs. Th e growth rate of spontaneous NET formation was 3.9-fold higher than the induced NETs formation. Spontaneous formation of NETs above 16.4 % may serve as a possible RA activation indicator. Neutrophils from ACPA-positive RA patients were found to have increased spontaneous and induced NETs formation compared to ACPA-negative RA patients. Conclusion. We have studied the main parameters of NETs formation and their dynamics associated with the transition from remission to active autoimmune infl ammation in RA patients. Th e activity of RA has a more pronounced eff ect on the intensity of NETs formation than the presence of ACPA. NETs may be considered a new potential diagnostic and prognostic biomarker

Impact of Anti-Citrullinated Protein Antibodies on Progressive Systemic Bone Mineral Density Loss in Patients With Early Rheumatoid Arthritis After Two Years of Treat-to-Target.

ObjectivesTo investigate the association of anti-citrullinated protein antibodies (ACPA) with changes in systemic bone mineral density (BMD) in patients with early rheumatoid arthritis (RA) after two years of treat-to-target.MethodsBMD was measured at the lumbar spine (LS) and femoral neck (FN) in 100 patients with recent onset RA at baseline and after 24 months of treatment aimed at low disease activity (LDA) according to the 28-joints disease activity score (DAS28 <3.2). Multivariable regression analyses were performed to determine independent associations between autoantibodies and other disease and treatment-related parameters with BMD loss.ResultsAfter 24 months, the majority of the patients were at least in LDA (78%), with slightly more ACPA-positive subjects achieving the target. The BMD had significantly decreased at both the LS (mean [SD] percent loss -1.8 [6.2], p=0.03) and the FN (-2.4 [7.3], p=0.03) in ACPA-positive but not in ACPA-negative patients. Consequently, the proportion of patients with reduced BMD (Z score ≤-1) after 24 months was significantly higher among ACPA-positive patients at both the spine (39.5% vs 19.3%, p=0.05) and the hip (37.2% vs 12.2%, p=0.007). The association between ACPA and BMD loss was independent of other variables including age, gender, disease activity, cumulative dose of glucocorticoids and duration of therapy with bisphosphonates at the LS but not the FN.ConclusionsACPA are associated with ongoing BMD loss at the spine despite suppression of inflammation and adoption of prophylactic measures. ACPA-positive RA patients should be therefore strictly monitored for the development of osteoporosis.

Periodontal disease in seropositive rheumatoid arthritis: scoping review of the epidemiological evidence.

The link between periodontal disease (PD) and rheumatoid arthritis (RA) has been hypothesized to lie in the anti-cyclic citrullinated protein antibody (ACPA) molecules present in seropositive RA. This review aimed to discuss how RA and specifically ACPA-positive RA link to PD, and appraise the epidemiological evidence on the relationship between ACPA-positive RA and PD. Articles were searched following the PRISMA guideline across the MEDLINE, Web of Science, Scopus and Cochrane Library databases. A total of 21 articles met the inclusion criteria of reporting the epidemiological data on the different ACPA status of the subjects with RA and PD (or periodontitis) parameters. A discrepancy is noted in the epidemiological evidence on the difference in the prevalence and severity of PD between ACPA-positive and ACPA-negative RA patients. Although the link between RA and PD is mostly discussed in terms of ACPA, reports on the different manifestations of PD between the two RA subsets remains inconclusive.

AB0150 ACPA-STATUS AND RHEUMATOID ARTHRITIS: MUCH MORE TO BE REVEALED!

Background:Anti-citrullinated protein antibodies (ACPA) are commonly associated with Rheumatoid arthritis (RA). RA is, therefore, classified as immunopositive or immunonegative, with disparate mechanisms in predisposition.Objectives:Our study aimed to determine the baseline characteristics and differences of ACPA-positive and ACPA-negative RA.Methods:We conducted a cross-sectional study including 224 patients with RA. All patients fulfilled the 2010 American College of Rheumatology/European League Against Rheumatism RA classification criteria. The patients were divided according to their ACPA status into two groups: ACPA-positive group (G1) and ACPA-negative group (G2). We compared clinical, radiological, and laboratory findings between the two groups, as well as extra-articular manifestations, comorbidities including fractures and osteoporosis. The Fracture Risk Assessment Tool (FRAX) was used to estimate the 10-year probability of major osteoporotic fracture (MOF) and also hip fracture (FH).Results:Of the 224 patients, 31.6% were negative for ACPA (n=71). Female predominance was found in both groups with a sex ratio of 0.25 (p=0.203). ACPA-negative subjects were younger (57±11 versus 59±12 years) (p= 0.305).The initial presentation of RA was different between the two groups without reaching statistical significance. In the ACPA-negative group, alteration of general condition was more frequent (16.9% in G2 versus 13.7% in G1) (p=0.533), with a tendency to oligo-articular onset (18.5% in G2 versus 6.7% in G1) (p=0.737). ACPA-positivity was more associated with an acute start of symptoms (10.4% in G1 versus 8.4% in G2) (p=0.639)There was no significant difference in the mean DAS28-VS (5.2±1.1 in G2 versus 5.5±1.3 in G1) and DAS28-CRP levels (5±1 in G2 versus 5.3±1.2 in G1) (p=0.069 and p=0.098 respectively).ACPA-positive RA was, however, significantly associated with more structural joint damage: erosions (55.9±53 in G1 versus 78±36 in G2, p=0.01), joint space narrowing (50.4±45.5 in G1 versus 33.1±36.6 in G2, p=0.003), Sharp/van der Heijde radiographic score (126.6 ±103.2 in G1 versus 88.8±81.5 in G2, p=0.004). ACPA-positive RA patients had more atlantoaxial dislocation: 20.2% in G1 versus 7% in G2 (p=0.012). There was no significant difference in hip involvement (9.8% in G1 versus 14% G2) (p=0.344).There were no significant differences in extra-articular manifestations between the two groups: Rheumatoid nodules (10.4% in G1 versus 18.3% in G2) (p=0.891), Sjögren’s syndrome (16.3% in G1 versus 16.9% in G2)(p=0.715), amyloidosis (0.6% in G1) (p=1), pulmonary fibrosis (5.8% in G1 versus 4.2% in G2) (p=0.757), neurological signs (4.5% in in G1 versus 5.6% in G2) (p=0.733), anaemia (5.8% in G1 versus 1.4% in G2) (p=0.175).When analyzing comorbidities, no significant differences were found: diabetes (10.4% in G1 versus 18.3% in G2) (p=0.103), cardiovascular diseases (19.6% in G1) (p=1), neurological diseases (0.06% in G1 versus 1.4% in G2) (p=0.534), dysthyroidism (2.6% in G1 versus 5.6% in G2) (p=0.267), dyslipidemia (3.2% in G1 versus 4.2% in G2) (p=0.711), cancer (1.3% in G1) (p=1). There were neither significant differences in the prevalence of fracture (21.5% in G1 versus 18.3% in G2) (p=0.574), and osteoporosis (23.6% in G1 versus 29.5% in G2) (p=0.347), between the two groups. However, ACPA-positive patients presented with a significantly higher FRAX score of MOF (2±2.8 in G1 versus 1.2±1 in G2) (p=0.006), and FRAX score of FH (0,9±1.8 in G1 versus 0.3±0.5 in G2) (p=0.003).Conclusion:ACPA status appears to influence both the clinical presentation and radiological progression of RA patients. ACPA-positive patients present with an acute start of symptoms, with more structural damages and atlantoaxial dislocation. Comorbidities, including osteoporosis, does not seem affected by ACPA status. However, regardless of osteoporosis, ACPA-positivity is associated with a higher probability of major osteoporotic and hip fractures. Further research is needed to clarify this relationship.Disclosure of Interests:None declared

HLA-DRB1*07:01 and *08:02 Alleles Confer a Protective Effect Against ACPA-Positive Rheumatoid Arthritis in a Latin American Admixed Population.

Simple SummaryThe focus of the genetic studies on rheumatoid arthritis has been on Europeans or European-derived populations, with limited information available on other populations, especially in Latin America. The aim of the present study was to test previously reported HLA markers, the most important genetic contributors for the risk of developing rheumatoid arthritis, associated with anti-citrullinated protein antibodies-positive rheumatoid arthritis in a Latin American population with admixed ancestry. Our study shows for the first time an association between HLA-DRB1*07:01 and *08:02 alleles and protection against anti-citrullinated protein antibodies-positive rheumatoid arthritis in the Chilean population. In addition, our results seem to support the existence of differentiated genomic patterns in Chilean, and probably other Latin American populations, that are not the same that the found in Europeans regarding to loci repeatedly involved in rheumatoid arthritis. Identifying relationships between HLA-DRB1 alleles and rheumatoid arthritis is important for identifying disease associations in different ethnic groups in order to reach a better understanding of rheumatoid arthritis worldwide.HLA-DRB1 shared epitope (SE) alleles are important genetic contributors for the risk of developing anti-citrullinated protein antibodies (ACPA)-positive rheumatoid arthritis (RA), particularly in Caucasians. We aimed to analyze the contribution of HLA-DRB1 alleles and single nucleotide polymorphisms (SNPs) within the major histocompatibility complex (MHC) region to the susceptibility to develop ACPA-positive RA in a Latin American (LA) population with admixed ancestry. A total of 289 ACPA-positive RA patients and 510 controls were enrolled in this study. The presence of HLA-DRB1*04:01, *09:01 and *10:01 was increased in ACPA-positive RA patients compared with healthy controls (p < 0.0001, p < 0.001 and p < 0.01, respectively), whereas DRB1*07:01 and *08:02 was associated with a decreased risk of ACPA-positive RA (p < 0.001 and p < 0.01, respectively). These results showed a strong correlation with estimates from studies in Asians but not in Caucasian populations. The present study describes the protective effects of the HLA-DRB1*07:01 and *08:02 alleles in ACPA-positive RA patients in a LA population for the first time. Identifying relationships between HLA-DRB1 alleles and RA is important for identifying disease associations in different ethnic groups in order to reach a better understanding of RA worldwide.

FRI0107 EFFECTIVENESS OF ABATACEPT ON CLINICAL DISEASE ACTIVITY AND RADIOGRAPHIC PROGRESSION IN RHEUMATOID ARTHRITIS PATIENTS IN DAILY CLINICAL PRACTICE IN JAPAN: COMPARISONS ACCORDING TO ACPA STATUS

Background:The clinical effectiveness of abatacept (ABA) in rheumatoid arthritis (RA) patients has been reported to be higher when the patients’ anti-cyclic citrullinated peptide antibody (ACPA) status is positive. The report from the ORA registry demonstrated that the ACPA positivity was associated with a better response to ABA [1]. In a sub-analysis of the AMPLE trial, patients with very high ACPA titers who were treated with ABA had a statistically significant response compared to patients with lower titers [2]. However, these studies did not demonstrate the data regarding the structural progression.Objectives:This study aimed to evaluate the effectiveness of ABA on the clinical disease activity as well as the radiographic progression in patients with RA in the clinical settings.Methods:All eligible patients were registered in the TBCR, a Japanese multicenter registry system for RA patients treated with biologics [3]. The present study included 553 consecutive patients whose ACPA data were obtained, treated with ABA and observed for longer than 52 weeks. We primarily compared the status of disease activity (SDAI) and radiographic progression (van der Heijde modified total Sharp score: mTSS) between ACPA-positive [ACPA (+)] and ACPA-negative [ACPA (-)] RA patients. The ACPA positive was defined as ≥13.5 U/ml of anti-CCP antibody.Results:Number of cases was 446/ 107 [ACPA (+)/ ACPA (-)], respectively. Baseline characteristics between groups were quite similar; mean age was 68.0/ 67.3 years, rate of methotrexate (MTX) use rate was 41.2/ 50.0%, rate of bio-naive was 28.0/ 31.8%, and mean SDAI score was 22.2/ 20.8. Significant difference was observed in mean change in SDAI score from baseline to 52 weeks between the ACPA (+) and ACPA (-) group (-13.4 vs -9.9, p = 0.027) (Figure 1A). Proportion of patents that achieved low disease activity (LDA; SDAI ≤11) at 52 weeks was significantly higher in the ACPA (+) group compared to the ACPA (-) group (72.1 vs 56.0%, p &lt; 0.01) (Figure 1B). In univariate and multivariate logistic regression analysis, ACPA positivity was an independent predictor for achievement of LDA at 52 weeks (Table). There observed no significant difference between ACPA (+) and ACPA (-) group in the proportion of patients that achieved structural remission (ΔmTSS ≤0.5) at 52 weeks (66.2 vs 62.1%) (Figure 2A) as well as mean change in mTSS (1.66 vs 1.17), erosion score (0.60 vs 0.53), and joint narrowing (JSN) score (1.06 vs 0.64) (Figure 2B).Table.UnivariateMultivariateVariablesOR (95%CI)p-valueadjusted OR (95%CI)p-valueAge0.99 (0.98-1.01)0.4391.00 (0.97-1.02)0.749male (vs female)1.12 (0.70-1.80)0.6340.79 (0.40-1.58)0.511disease duration0.99 (0.97-1.00)0.0530.99 (0.97-1.01)0.468Biologics-naïve1.23 (0.81-1.85)0.3351.18 (0.67-2.08)0.575Concomitant MTX use1.12 (0.75-1.69)0.5851.14 (0.66-1.95)0.649Concomitant PSL use0.82 (0.55-1.23)0.3290.97 (0.58-1.64)0.923SDAI @baseline0.96 (0.94-0.97)&lt;0.0010.96 (0.94-0.98)&lt;0.001mHAQ @baseline0.50 (0.36-0.69)&lt;0.0010.57 (0.38-0.86)0.008ACPA positive2.03 (1.29-3.17)0.0022.61 (1.36-5.00)0.004Bold italic, p&lt;0.05Conclusion:Consistent with previous reports, the ACPA-positive group demonstrated significantly higher LDA achievement rate at 52 weeks and indeed the ACPA positivity was significantly associated with LDA achievement in multivariate analysis. However, the ACPA-negative group demonstrated quite similar transition of SDAI score and LDA achievement rate except at 52 weeks compared with the ACPA-positive group. Additionally, there was no significant difference in the structural progression at 52 weeks between the groups. ABA treatment may be considered not only in the ACPA-positive RA patients but also in the ACPA-negative patients in the clinical practice.

Efficacy and safety of NI-0101, an anti-toll-like receptor 4 monoclonal antibody, in patients with rheumatoid arthritis after inadequate response to methotrexate: a phase II study

ObjectivesAnti-citrullinated protein antibodies (ACPAs) form immune complexes with citrullinated proteins binding toll-like receptor (TLR) 4, which has been proposed as a mediator of rheumatoid arthritis (RA). NI-0101 is a first-in-class...

N-Glycosylation Site Analysis of Citrullinated Antigen-Specific B-Cell Receptors Indicates Alternative Selection Pathways During Autoreactive B-Cell Development.

Many autoimmune diseases are hallmarked by autoreactive B and plasma cell responses that are directly or indirectly involved in disease pathogenesis. These B-cell responses show large variability between diseases, both in terms of the secreted autoantibody repertoire and the dynamics and characteristics of the underlying B-cell responses. Hence, different mechanisms have been proposed to explain the emergence of autoreactive B cells in an otherwise self-tolerant immune system. Notably, most mechanistic insights have been obtained from murine studies using models harboring genetic modifications of B and T cells. Given recent technological advances that have rendered autoreactive human B cells accessible for analysis, we here discuss the phenomenon of extensive N-glycosylation of the B-cell receptor (BCR) variable domain of a prototypic human autoreactive B-cell response and its potential role in the generation of autoimmunity. Anti-citrullinated protein antibodies (ACPA) hallmark the most disease-specific autoimmune response in Rheumatoid Arthritis (RA). Remarkably, ACPA-IgG are heavily N-glycosylated in the variable domain due to somatic mutations that generate abundant N-glycosylation consensus sequences. These sites, obtained from full-length BCR sequences of ACPA-expressing B cells from 12 ACPA-positive RA patients, were here analyzed in detail. Sites that required a single nucleotide mutation to be generated were defined as single somatic hypermutation (s-SHM) sites, whereas sites requiring multiple mutations were defined as m-SHM sites. IgG sequences of 12 healthy donors were used as control. Computational modeling of the germinal center reaction (CLONE algorithm) was used with the germline counterparts of ACPA-IgG heavy chain (HC) sequences to simulate the germinal center response. Our analyses revealed an abundance of N-glycosylation sites in ACPA-IgG HC that frequently required multiple mutations and predominated in specific positions. Based on these data, and taking into account recent insights into the dynamics of the ACPA-response during disease development, we here discuss the hypothesis that N-glycosylation sites in ACPA-IgG variable domains could lead to alternative, possibly antibody affinity-independent selection forces. Presumably, this occurs during germinal center responses allowing these B cells to escape from putative tolerance checkpoints, thereby driving autoreactive B cell development in the pathogenesis of RA.

Does immunological remission, defined as disappearance of autoantibodies, occur with current treatment strategies? A long-term follow-up study in rheumatoid arthritis patients who achieved sustained DMARD-free status

ObjectivesSustained disease-modifying antirheumatic drug (DMARD)-free status, the sustained absence of synovitis after cessation of DMARD therapy, is infrequent in autoantibody-positive rheumatoid arthritis (RA), but approximates cure (ie, disappearance of signs...

ACPA-negative RA consists of subgroups: patients with high likelihood of achieving sustained DMARD-free remission can be identified by serological markers at disease presentation

BackgroundDisease-modifying antirheumatic drug (DMARD)-free remission, the sustained absence of synovitis after DMARD cessation, is increasingly achievable, especially in autoantibody-negative rheumatoid arthritis (RA). However, underlying mechanisms are unknown and patient subgroups that achieve this outcome are insufficiently characterized. We evaluated whether serological biomarkers at disease onset, as measured within the multi-biomarker disease activity (MBDA) score, are differently expressed in RA patients who achieve sustained DMARD-free remission.MethodsTwo hundred ninety-nine RA patients were evaluated for achievement of sustained DMARD-free remission during a median follow-up of 4.3 years. Twelve biomarkers, as included in the MBDA score, were determined from the serum obtained at disease onset. Patients were categorized as having a low (< 30), moderate (30–44) or high (> 44) score. Analyses were stratified for anti-citrullinated protein antibodies (ACPA) based under the assumption that ACPA-positive and ACPA-negative RA are different disease entities.ResultsTwenty percent achieved sustained DMARD-free remission. Overall, high MBDA scores were associated with achieving DMARD-free remission (high vs. low HR 3.8, 95% CI 1.2–12.2). Among ACPA-negative RA patients, moderate or high scores associated strongly with DMARD-free remission (moderate vs. low HR 9.4, 95% CI 1.2–72.9; high vs. low HR 9.7, 95% CI 1.3–71.1). This association was independent of age and other clinical factors (high vs. low HR 8.2, 95% CI 1.1–61.8). For ACPA-negative RA patients, the biomarkers C-reactive protein, serum amyloid A and matrix metalloproteinase-3 were individually associated with sustained DMARD-free remission. Among ACPA-positive RA patients, scores were not associated with DMARD-free remission.ConclusionsACPA-negative RA patients who achieved sustained DMARD-free remission after treatment withdrawal were characterized by moderate to high MBDA scores at diagnosis. This is the first evidence that ACPA-negative RA can be subdivided in clinically relevant subsets at disease onset using a protein profile.

Neutrophil extracellular traps exert both pro- and anti-inflammatory actions in rheumatoid arthritis that are modulated by C1q and LL-37

ObjectiveNeutrophil extracellular traps (NET), produced by activated polymorphonuclear neutrophils (PMN), are supposed to play a role in the pathogenesis of rheumatoid arthritis (RA), a chronic inflammatory autoimmune disease characterized by anti-citrullinated protein antibodies (ACPA). Indeed, NET contain citrullinated autoantigens and some RA autoantibodies recognize NET. However, the mechanisms by which NET trigger or perpetuate the inflammatory process in RA are hitherto not elucidated. We hypothesized that, in addition to citrullination, NET might also contain stimulatory proteins and directly activate inflammatory target cells, as PMN and macrophages. MethodsNET antigenic and inflammatory properties were analyzed in 157 healthy donors (HD) and RA patients, the largest analysis reported so far. Primary PMN and monocyte-derived macrophages were isolated and immunoglobulin G (IgG) purified. NET were induced (NETosis), isolated and quantified. NET antigenicity was analyzed by fluorescence microscopy. PMN and macrophages were stimulated with NET with/without ACPA, C1q, LL-37 or lipopolysaccharide (LPS) and cell activation was estimated by flow cytometry and ELISA. ResultsPMN from RA patients produced more NET than HD PMN. We next dissected how NET mechanistically affect inflammatory cells. Particularly, we show for the first time that RA and HD NET activated both resting macrophages and PMN, but importantly RA NET were more stimulatory, leading to secretion of inflammatory cytokines and up-regulation of HLA/CD86/CD11b. IgG from ACPA-positive RA patients specifically recognized RA and even HD NET. Nevertheless, NET-induced cell activation occurs independently of immune complex formation with ACPA. Likewise, endosomal acidification was not required. Notably, we also report that complement C1q increased the NET stimulatory activity on macrophages only, due to higher expression of C1q receptors, which was further supported by the LL-37 antimicrobial peptide. In contrast, NET specifically inhibited interleukin (IL)-6 secretion by LPS-activated macrophages and not PMN, especially with C1q/LL-37. This inhibition was not mediated by NET-derived proteases or LPS neutralization and was associated with the simultaneous induction of IL-10 secretion. ConclusionWe show that NET possess both pro- and anti-inflammatory properties depending on target cells, their activation levels and C1q/LL-37. Thus, independently of ACPA, NET modulate RA chronic inflammation via this new dual activity we identified. In addition, NET may trigger autoimmunity in RA as ACPA recognize NET antigens but not non-activated PMN. Therefore, we conclude that excess of NETosis together with enhanced NET activity participate to RA pathogenesis at different levels.