Izabela Domysławska

Effect of etanercept on serum levels of soluble cell adhesion molecules (sICAM-1, sVCAM-1, and sE-selectin) and vascular endothelial growth factor in patients with rheumatoid arthritis.

Objective: Endothelium and adhesion molecules are engaged in the pathogenesis of rheumatoid arthritis (RA). This study was undertaken to analyse the effect of etanercept on the levels of soluble cell adhesion molecules (sCAMs) and vascular endothelial growth factor (VEGF) in patients with active RA. Methods: Patients were receiving 50 mg/week of subcutaneous etanercept and 10–25 mg/week of methotrexate (MTX). Serum levels of soluble intercellular adhesion molecule-1 (sICAM-1), vascular cell adhesion molecule-1 (sVCAM-1), E-selectin (sE-selectin), and VEGF were measured by enzyme-linked immunosorbent assay (ELISA) in 18 RA patients (prior to injection) at 0, 3, 6, 9, and 12 months. Results: A decrease in serum levels of sICAM-1 (p<0.001), sVCAM-1 (p<0.01), sE-selectin (p<0.01), and VEGF (p<0.001) was observed in RA patients after 3 months of treatment with etanercept. Six months of therapy with etanercept prolonged the suppression of serum sICAM-1 (p<0.01) and even more remarkably diminished sVCAM-1, sE-selectin, and VEGF (in all cases p<0.001) concentrations as compared to baseline (month 0). Treatment also effectively diminished sICAM-1, sVCAM-1, and VEGF levels at months 9 and 12 (in all cases p<0.001), and less significantly sE-selectin (p<0.05 at month 9 and p<0.01 at month 12). The Disease Activity Score including a 28-joint count (DAS28) measured at 3, 6, 9, and 12 months decreased significantly compared to baseline (in all cases p<0.001). Conclusion: Our study shows that, besides a rapid suppression of disease activity, serum sCAM and VEGF concentrations are downregulated following anti-tumour necrosis factor alpha (TNFα) therapy combined with MTX. Prolonged treatment with etanercept sustained or even more remarkably diminished the sCAM and VEGF serum concentrations.

Changes of glycosylation of IgG in rheumatoid arthritis patients treated with methotrexate.

PURPOSE In patients with active rheumatoid arthritis (RA) decrease of galactosylation is correlated with disease activity. The aim of our study was to evaluate an effect of methotrexate therapy on glycosylation disturbances of IgG in RA patients. MATERIALS/METHODS IgG glycosylation in 40 patients with active RA treated with methotrexate for 12 months prior to and after treatment were compared. The control group consisted of 20 healthy volunteers. IgG glycosylation was assessed using biotinylated lectins and immunosorbent ELISA assay. For galactose specificity Datura stramonium lectin (DSA), for sialic acid Sambucus nigra (SNA) and Maackia amurensis (MAA) and for fucose residue Areulia auranta (AAA) lectins were used. RESULTS In RA-cases N-glycan galactosylation and sialylation of IgG before treatment were significantly lower than in healthy subjects (for DSA, MAA lectins p<0.001 and SNA p<0.05). Significant increase of IgG galactosylation and sialylation in RA patients after therapy (for DSA, MAA and SNA lectin p<0.05) was detected. Moreover the glycosylation disturbances of N-glycan IgG were strongly associated with changes of disease activity based on disease activity score. For fucose residues significantly higher absorbency of AAA lectin in RA patients before treatment was observed compared to control subjects (p<0.05) and slightly, not significantly decreased after MTX therapy. CONCLUSIONS Defect of galactosylation of IgG in RA patients is a useful marker of disease activity that may be used for the assessment of therapy effectiveness. The role of IgG fucosylation and sialylation in RA pathogenesis has still to be determined.

AB0447 Regulation of Serum Matrix Metalloproteinases and Tissue Inhibitor of Metalloproteinases-1 following Rituximab Therapy in Patients with Rheumatoid Arthritis Refractory to Anti-Tumor Necrosis Factor Blockers

Background Serum matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) play important role in the pathogenesis of rheumatoid arthritis (RA). Objectives In our article we evaluated the regulatory effects of the infusions of rituximab, a monoclonal antibody directed against CD20+ B cells, on the serum MMPs and TIMP-1 levels in patients with active RA not responding to anti-tumor necrosis factor (anti-TNF) therapy. Methods Twelve RA patients were planned to receive 4 infusions of 1000mg of rituximab at weeks 0, 2, 24, and 26. The therapy was combined with methotrexate (MTX) (20-30mg/week). Seven patients were refractory to previously received infliximab, and 5 to etanercept. Serum concentrations of interstitial collagenase (MMP-1), stromelysin-1 (MMP-3), gelatinase B (MMP-9) and TIMP-1 were measured by ELISA on weeks 0, 2, 12, 24, 36 and 52. Results Initial infusion of rituximab down-regulated serum MMP-1 (p<0.01), MMP-3 (p<0.001), MMP-9 (p<0.001) and TIMP-1 (p<0.05) levels. Second drug administration caused even more remarkable reduction of measured MMPs (p<0.001 in all cases) and TIMP-1 level (p<0.01). These findings were accompanied by significantly decreased ratios of measured MMPs to TIMP-1. Next rituximab infusions on weeks 24 and 26 sustained the suppression of serum MMPs levels. Prior to the initial rituximab infusion serum concentrations of studied MMPs and TIMP-1 significantly correlated with markers of RA activity such as disease activity score (DAS28) and CRP levels. Conclusions Rituximab therapy, beside a rapid clinical improvement, reduced serum MMPs concentrations in RA patients refractory to anti-TNF treatment. Repeated infusions of rituximab maintained initial serum MMPs suppression. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.1496

AB0121 Changes of Glycosylation of Immunoglobulin G in Rheumatoid Arthritis Patients Treated with Methotrexate

Background In patients with active rheumatoid arthritis (RA) decrease of galactosylation of IgG is observed. It has been shown that agalactosylation is correlated with disease activity (1, 3, 4). Objectives The aim of our study was to evaluate an effect of methotrexate (MTX) therapy on glycosylation disturbances of IgG in RA patients. Methods 40 patients with active RA was treated with MTX for 12 months in dosage 15-25 mg per week (2). All patients were in remission based on DAS 28 (≤2.6) at the end of the study. The control group was consisted of 20 healthy volunteers in the same age. IgG was isolated from patients sera and analysis of IgG galactosylation by biotinylated lectins was performed. Immunosorbent assay ELISA was used for this purpose. For galactose specifity Datura stramonium lectin (DSA), for sialic acid Sambucus nigra (SNA) and Maackia amurensis (MAA) and for fucose residue (Areulia auranta) AAA lectin was used (5). Results Our data demonstrated that N-glycan galactosylation and sialylation of IgG in untreated (naive) RA patients was significantly lower than in control group (for DSA, MAA lectins p<0.001 and SNA p<0.05). Furthermore we showed significant increase of IgG galactosylation and sialylation in RA patients after twelve months of MTX therapy (for DSA,MAA and SNA lectin p<0.05) compared to those before treatment. Moreover the glycosylation disturbances of N-glycan IgG were strongly associated with changes of disease activity based on DAS 28 (p<0.001). For fucose residues significantly higher absorbancy of AAA lectin in RA patients before MTX treatment was observed compared to control group (p<0.05). There were no correlation between absorbancy of this lectins before and after treatment but in RA treated patients still higher level of fucosylation was observed. There were no correlation between lectins absorbancy and ACPA and RF titer. Conclusions Defect of glycosylation of IgG in RA can be detected by simple lectin method. Assessment of disturbances of glycosylation of IgG might be a useful marker of disease activity and effectiveness of treatment. References Watson M, Rudd PM, Bland M et al. Sugar printing rheumatic diseases. Arthritis and Rheum. Vol.42, No 8, Aug 1999, pp 1682-1690 Arnett FC, Edworthy SM, Bloch DA et al. The American Rheumatism Association 1987 revised criteria for the classification of rheumatoid arthritis. Arthritis Rheum. 1998, 31: 315-324 Axford JS, Sumar N, Alavi A et al. Changes in normal glycosylation mechanisms in autoimmune rheumatic disease. J. Clin. Invest., 1992; 89: 1021-1031 Gindzienska-Sieskiewicz E, Klimiuk PA, Kisiel DG et al. The changes In monosaccharide composition of immunoglobulin G in the course of rheumatoid arthritis. Clin Rheumatol. 2007 May;26(5):685-90. Radziejewska I, Borzym-Kluczyk M, Namiot Z et al. Glycosylation of mucins present In gastrin juice: the effect of Helicobacter pylori eradication treatment. Clin Exp Med. 2011, 11:81-88 Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.2100

SAT0024 Increased production of leukotrienes by peripheral blood mononuclear cells is associated with more severe disease and worse prognosis in patients with systemic sclerosis

Background Leukotrienes (LTs) are a group of pro-inflammatory and pro-fibrotic, arachidonic acid-derived lipid mediators. We have shown that LTs are increased in the lungs of patients with interstitial lung disease (ILD) associated with systemic sclerosis (SSc). Objectives Since leukocytes are considered a major source of LTs in humans and since inflammatory infiltrates consisting of mononuclear cells are found in the skin and internal organs in early SSc, we undertook this study to investigate the relationship between the synthesis of LTs by peripheral blood mononuclear cells (PBMC) and clinical features of SSc. Methods Leukotriene B4 (LTB4) and cysteinyl leukotrienes (CysLTs) were measured by ELISA in the supernatants from ionophore-stimulated PBMC of 39 patients with SSc and 24 age- and sex-matched healthy controls (HC). Only patients, who had not received immunosuppressive drugs, aspirin or other NSAIDs for at least 6 months, 1 month and 7 days respectively before the study, were included. Follow-up data were available in 25 SSc patients (mean ± SD follow-up time: 34±18 months). Disease progression was defined as death due to SSc-related organ complication, development of a new or progression of pre-existing SSc-related organ involvement (>10% decrease in FVC, increase by at least one WHO class, increase by ≥120% of upper normal limit in serum creatinine and/or loss of >10% of weight after excluding other causes). Results Concentration of LTB4 was significantly higher in PBMC cultures from patients with SSc (640±518 pg/mL/105cells) as compared with HC (353±216 pg/mL/105cells, p<0.05). Higher LTB4 levels were associated with the presence of diffuse SSc, restrictive ILD (FVC<80% predictive and FEV1/FVC >0.8), pulmonary hypertension (PASP>45 mmHg by echo), and more severe microangiopathy in capillaroscpy. Mean concentration of LTB4 was higher in 7/25 SSc patients who experienced subsequent progression of the disease (918±665 pg/mL/105cells) compared with the remaining 18/25 with stable disease (569±549 pg/mL/105cells), but the difference was not significant (p=0.1). We did not find significant differences in the synthesis of CysLTs between SSc patients (313±162 pg/mL/105cells) and HC (266±138 pg/mL/105cells, p>0.05), nor significant associations of CysTLs concentration and SSc-related organ involvement. However, concentration of CysLTs was significantly higher in PBMC cultures from SSc patients with subsequent progression of the disease (452.0±197.3 pg/mL/105cells) as compared with those with stable SSc (275.6±138.9 pg/mL/105cells, p<0.05). Conclusions The results of our study indicate that increased synthesis of LTs might be involved in the pathogenesis and progression of SSc. Consequently, inhibition of LTs synthesis or action might represent a new, promising target in the treatment of SSc. Disclosure of Interest A. Lapinska: None Declared, M. Bielecki: None Declared, O. Distler Grant/Research support from: PD Dr. O. Distler has consultancy relationship and/or has received research funding from Actelion, Pfizer, Ergonex, BMS, Sanofi-Aventis, United BioSource Corporation, medac, 4D Science, Boehringer-Ingelheim, Active Biotech and Roche in the area of potential treatments of scleroderma and its complications. Speakers Bureau: PD Dr O. Distler has received lecture honoraria from Actelion, Pfizer, Encysive and Ergonex., I. Domyslawska: None Declared, L. Chyczewski: None Declared, S. Sierakowski: None Declared, S. Gay: None Declared, K. Kowal: None Declared, O. Kowal-Bielecka Grant/Research support from: Dr O Kowal-Bielecka work was supported by research grant (2-PO5B-050-27) from the Polish State Committee for Scientific Research.

[Coexistence of scleroderma-like syndrome and idiopathic myelofibrosis in a 54-year-old female patient: case report].

Systemic sclerosis (SSc) is characterized by immunological disturbances, vascular damage and overproduction of extracellular matrix by stimulated fibroblasts. It has been postulated that immunological reactions involved in the pathogenesis of SSc may promote the development of malignancies. Coexistence of this disease with neoplasmatic processes is relatively frequent. In our report we describe a case a 54-year-old woman with scleroderma-like syndrome, which has preceded the occurrence of idiopathic myelofibrosis by many years. Owing to multiple repeated diagnostic tests we managed to diagnose this disease at the early stage, which enabled effective therapy with remission of blood dyscrasia as well as inhibition of skin lesions and lung fibrosis.