Non-response to rituximab therapy in rheumatoid arthritis is associated with incomplete disruption of the B cell receptor repertoire.

OBJECTIVE: To gain more insight into the dynamics of lymphocyte depletion and develop new predictors of clinical response to rituximab in rheumatoid arthritis (RA). METHODS: RNA-based next-generation sequencing was used to analyse the B cell receptor (BCR) repertoire in peripheral blood and synovial tissue samples collected from 24 seropositive patients with RA treated with rituximab. Clonal expansion, mutation load and clonal overlap were assessed in samples collected before, at week 4 and at week 16 or 24 after treatment and correlated to the patients' clinical response. RESULTS: After 4 weeks of rituximab-induced B cell depletion, the peripheral blood BCR repertoire of treated patients consisted of fewer, more dominant and more mutated BCR clones. No significant changes in the synovial tissue BCR repertoire were detected until week 16 post-treatment, when a reduced clonal overlap with baseline and an increased mutation load were observed. In patients who were non-responders at month 3 (n=5) using the European League Against Rheumatism response criteria, peripheral blood samples taken at week 4 after rituximab treatment showed more dominant clones compared with moderate responders (n=9) (median (IQR): 36 (27-52) vs 18 (16-26); p<0.01) and more clonal overlap with the baseline (median (IQR): 5% (2%-20%) vs 0% (0%-0%); p≤0.01). CONCLUSION: Significant changes in BCR clonality are observed in peripheral blood of patients 4 weeks after rituximab treatment, while changes in synovial tissue were observed at later time points. Incomplete depletion of the dominant baseline peripheral blood BCR repertoire in the first month of treatment might predict clinical non-response at 3 months.

Efficacy and safety of adalimumab for the treatment of peripheral arthritis in spondyloarthritis patients without ankylosing spondylitis or psoriatic arthritis.

OBJECTIVES: To evaluate the efficacy and safety of adalimumab in patients with peripheral spondyloarthritis (SpA) not fulfilling the criteria for ankylosing spondylitis (AS) or psoriatic arthritis (PsA). METHODS: 40 patients with active peripheral SpA fulfilling the European Spondyloarthropathy Study Group or Amor criteria but not the criteria for AS or PsA were included in a randomised, double-blind, placebo-controlled clinical trial. Patients were treated 1 : 1 with adalimumab or placebo for 12 weeks, followed by an open label extension up to week 24. Safety and efficacy measurements were performed every 6 weeks, with the patient's global assessment of disease activity at week 12 as the primary endpoint. RESULTS: At week 12, the patient's and physician's global assessment of disease activity, swollen joint count, Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), Ankylosing Spondylitis Disease Activity Score (ASDAS) and erythrocyte sedimentation rate improved significantly in the adalimumab group compared with the baseline values and compared with placebo. A similar improvement was seen upon adalimumab treatment from weeks 12 to 24 in the patients originally randomised to placebo, whereas the clinical response was maintained or even augmented at week 24 in the patients who received adalimumab from the start. ASDAS inactive disease and BASDAI50 responses were met in 42% of the adalimumab group versus 0%-5% in the placebo group at week 12 (p=0.001 and p=0.008, respectively), and were further increased at week 24. The number of adverse events was not different between the adalimumab and placebo groups. CONCLUSIONS: Adalimumab appears to be effective and well tolerated in SpA patients with peripheral arthritis, also in those patients not fulfilling the AS or PsA criteria.

Progression of structural damage is not related to rituximab serum levels in rheumatoid arthritis patients.

OBJECTIVE: The most cost-effective dosing regimen for rituximab treatment in RA is currently unknown. The objective of this study is to determine whether low rituximab serum levels are associated with progression of structural damage in RA patients. METHODS: Sixty-two RA patients were treated with rituximab in three different centres. Structural damage was assessed on radiographs of hands and feet before and 1 year after therapy using the Sharp-van der Heijde scoring method (SHS). Patients were divided into progressors vs non-progressors based on different cut-off values. Rituximab serum levels were measured by sandwich ELISA after 4 and 12 weeks (Leiden University Medical Center and University Medical Centre, Utrecht cohorts) or 4 and 16 weeks (Academic Medical Center/University of Amsterdam cohort). RESULTS: There was no difference in rituximab levels between progressors and non-progressors 4 weeks and 12 or 16 weeks after initiation of treatment in the different cohorts. There was also no correlation between rituximab levels at week 4 or week 12 or 16 and change in SHS score after 1 year. CONCLUSION: Low rituximab serum levels are not associated with progression of structural damage in RA patients. The results do not support the use of dosages higher than 2 × 1000 mg rituximab to inhibit progression of joint destruction.

Rice availability and stability in Africa under future socio-economic development and climatic change.

As Africa is facing multiple challenges related to food security, frameworks integrating production and availability are urgent for policymaking. Attention should be given not only to gradual socio-economic and climatic changes but also to their temporal variability. Here we present an integrated framework that allows one to assess the impacts of socio-economic development, gradual climate change and climate anomalies. We apply this framework to rice production and consumption in Africa whereby we explicitly account for the continent's dependency on imported rice. We show that socio-economic development dictates rice availability, whereas climate change has only minor effects in the long term and is predicted not to amplify supply shocks. Still, rainfed-dominated or self-producing regions are sensitive to local climatic anomalies, while trade dominates stability in import-dependent regions. Our study suggests that facilitating agricultural development and limiting trade barriers are key in relieving future challenges to rice availability and stability.

Rituximab abrogates joint destruction in rheumatoid arthritis by inhibiting osteoclastogenesis.

OBJECTIVES: To examine how rituximab may result in the inhibition of joint destruction in rheumatoid arthritis (RA) patients. METHODS: Twenty-eight patients with active RA were treated with rituximab. Radiographs of hands and feet before and 1 year after therapy were assessed using the Sharp-van der Heijde score (SHS). Expression of bone destruction markers was evaluated by immunohistochemistry and immunofluorescence of synovial biopsies obtained before and 16 weeks after the initiation of treatment. Serum levels of osteoprotegerin, receptor activator of nuclear factor κB ligand (RANKL), osteocalcin and cross-linked N-telopeptides of type I collagen (NTx) were measured by ELISA before and 16 weeks post-treatment. RESULTS: After 1 year, the mean (SD) change in total SHS was 1.4 (10.0). Sixteen weeks after treatment there was a decrease of 99% in receptor activator of nuclear factor κB-positive osteoclast precursors (p=0.02) and a decrease of 37% (p=0.016) in RANKL expression in the synovium and a trend towards reduced synovial osteoprotegerin expression (25%, p=0.07). In serum, both osteoprotegerin (20%, p=0.001) and RANKL (40%, p<0.0001) levels were significantly reduced 16 weeks after treatment, but the osteoprotegerin/RANKL ratio increased (157%, p=0.006). A trend was found towards an increase of osteocalcin levels (p=0.053), while NTx concentrations did not change. CONCLUSIONS: Rituximab treatment is associated with a decrease in synovial osteoclast precursors and RANKL expression and an increase in the osteoprotegerin/RANKL ratio in serum. These observations may partly explain the protective effect of rituximab on the progression of joint destruction in RA.

Relationship between the type I interferon signature and the response to rituximab in rheumatoid arthritis patients.

OBJECTIVE: To analyze the relationship between the type I interferon (IFN) signature and clinical response to rituximab in rheumatoid arthritis (RA) patients. METHODS: Twenty RA patients were treated with rituximab (cohort 1). Clinical response was defined as a decrease in the Disease Activity Score evaluated in 28 joints (DAS28) and as a response according to the European League Against Rheumatism (EULAR) criteria at week 12 and week 24. The presence of an IFN signature was analyzed in peripheral blood mononuclear cells by measuring the expression levels of 3 IFN response genes by quantitative polymerase chain reaction analysis. After comparison with the findings in healthy controls, patients were classified as having an IFN high or an IFN low signature. The data were confirmed in a second independent cohort (n = 31). Serum IFNα bioactivity was analyzed using a reporter assay. RESULTS: In cohort 1, there was a better clinical response to rituximab in the IFN low signature group. Consistent with these findings, patients with an IFN low signature had a significantly greater reduction in the DAS28 and more often achieved a EULAR response at weeks 12 and 24 as compared with the patients with an IFN high signature in cohort 2 versus cohort 1. The pooled data showed a significantly stronger decrease in the DAS28 in IFN low signature patients at weeks 12 and 24 as compared with the IFN high signature group and a more frequent EULAR response at week 12. Accordingly, serum IFNα bioactivity at baseline was inversely associated with the clinical response, although this result did not reach statistical significance. CONCLUSION: The type I IFN signature negatively predicts the clinical response to rituximab treatment in patients with RA. This finding supports the notion that IFN signaling plays a role in the immunopathology of RA.

Intra-articular etanercept treatment in inflammatory arthritis: A randomized double-blind placebo-controlled proof of mechanism clinical trial validating TNF as a potential therapeutic target for local treatment.

OBJECTIVE: There is an increased interest in developing gene therapy approaches for local delivery of therapeutic genes in patients with arthritis. Intra-articular (i.a.) gene delivery, using an adeno-associated virus encoding a TNF soluble receptor, resulted in reduced paw swelling in an arthritis animal model, but i.a. treatment with a similar vector did not induce robust clinical improvement in patients. It is unclear whether this can be explained by for instance insufficient transduction efficiency or the fact that TNF is not a good therapeutic target for i.a treatment. The objective of this study was to explore the effects of i.a TNF blockade. METHODS: Thirty-one patients with rheumatoid or psoriatic arthritis were assigned to a single i.a. injection of 25mg etanercept or placebo in a double-blind randomised controlled clinical trial. The primary end point was target joint improvement, determined by a composite change index. RESULTS: Twenty-two patients received etanercept and 9 received placebo. Treatment was generally well tolerated. Treatment with etanercept resulted in a prompt and statistically significant improvement of the index (P<0.001) in comparison with placebo. As expected in light of the half-life of etanercept, the beneficial effect was transient and only statistically significant at week 1 and 2 after i.a. injection. CONCLUSION: The results support the development of novel approaches for long-term inhibition of TNF at the site of inflammation, such as gene therapy. TRIAL REGISTRATION: The Netherlands National Trial Register (NTR), www.trialregister.nl, NTR-1210.

Pro-atherogenic lipid changes and decreased hepatic LDL receptor expression by tocilizumab in rheumatoid arthritis.

OBJECTIVES: Blocking the interleukin-6 pathway by tocilizumab (TCZ) has been associated with changes in the lipoprotein profile, which could adversely impact cardiovascular (CV) risk in patients with rheumatoid arthritis (RA). In the present study, we addressed the effect of TCZ on lipoproteins in both fasting and non-fasting state in RA patients and tested the effect of TCZ on LDL receptor (LDLr) expression in vitro. METHODS: Twenty patients with active RA and an inadequate response to TNF blockers received monthly TCZ intravenously. On week 0, 1 and 6 blood was drawn before and after an oral fat load, the lipid profiles and HDL antioxidative capacity were measured. Effects of TCZ on LDLr expression in transfected HepG2 cells were subjected. RESULTS: After 6 weeks of TCZ, total cholesterol increased by 22% (4.8 ± 0.9 to 5.9 ± 1.3 mmol/L; p < 0.001), LDLc by 22% (3.0 ± 0.6 to 3.6 ± 0.8 mmol/L; p < 0.001) and HDLc by 17% (1.4 ± 0.4 to 1.7 ± 0.7 mmol/L; p < 0.016). Fasting triglycerides (TG) increased by 48% (1.0 ± 0.4 to 1.4 ± 0.8 mmol/L; p = 0.011), whereas postprandial incremental area under the curve TG increased by 62% (p = 0.002). Lipid changes were unrelated to the change in disease activity or inflammatory markers. No difference in HDL antioxidative capacity was found. In vitro, LDLr expression in cultured liver cells was significantly decreased following TCZ incubation (P < 0.001). CONCLUSIONS: TCZ adversely impacts on both LDLc as well as fasting and postprandial TG in patients with RA. The changes in hepatic LDLr expression following TCZ imply that adverse lipid changes may be a direct hepatic effect of TCZ. The net effect of TCZ on CV-morbidity has to be confirmed in future clinical trials.

Synovial tissue sublining CD68 expression is a biomarker of therapeutic response in rheumatoid arthritis clinical trials: consistency across centers.

OBJECTIVE: To determine whether the correlation between the mean change in disease activity and the mean change in synovial sublining (sl) CD68 expression could be demonstrated across different academic centers. METHODS: Synovial biopsies obtained at arthroscopy from patients with rheumatoid arthritis before and 160 days after rituximab therapy were selected and coded. Paired sections were processed independently at Amsterdam Medical Center (AMC) and at St. Vincent's University Hospital (SVUH), Dublin. Digital image analysis (DIA) was employed at both centers to quantify sublining CD68 expression. RESULTS: After analysis of CD68sl expression at centers in 2 different countries, high levels of intracenter and intercenter agreement were observed. For the pooled sections stained at AMC, the correlation between 2 investigators was R = 0.942, p = 0.000, and for sections stained at SVUH, R = 0.899, p = 0.001. Similarly, the intracenter correlations for DeltaCD68sl expression after treatment were R = 0.998, p = 0.000, for sections stained at AMC and R = 0.880, p = 0.000, for sections stained at SVUH. The intercenter correlation for the pooled scores of sections stained at AMC was R = 0.85, p = 0.000, and for the sections stained at SVUH, R = 0.62, p = 0.001. The consistent correlation between DeltaDAS (Disease Activity Score) and DeltaCD68sl expression across different studies (Pearson correlation = 0.895, p < 0.001) was confirmed. The standardized response mean values for DeltaCD68sl, calculated from analyses at both AMC and SVUH, were consistently 0.5 or greater, indicating a moderate to high potential to detect change. CONCLUSION: The correlation between mean DeltaDAS and mean DeltaCD68sl expression was confirmed across 2 centers. Examination of serial biopsy samples can be used reliably to screen for interesting biological effects at the site of inflammation at an early stage of drug development.

Disease activity-guided rituximab therapy in rheumatoid arthritis: the effects of re-treatment in initial nonresponders versus initial responders.

OBJECTIVE: To explore the efficacy of re-treatment with rituximab in patients with rheumatoid arthritis (RA) who were initial nonresponders to treatment, and to evaluate the effects of rituximab in RA when re-treatment is given in a standardized way based on the Disease Activity Score in 28 joints (DAS28), according to the international consensus statement. METHODS: Patients with RA who had a DAS28 of > or =3.2 received up to 3 courses of rituximab at intervals of at least 6 months, regardless of whether the patient had responded to the first course of treatment with rituximab. RESULTS: Of the 30 patients with RA who were included in the study, 26 could be evaluated for the efficacy of treatment after 6 months. Eighteen patients qualified for re-treatment at 6 months, 6 patients were re-treated at a later time point because of a disease relapse, and 2 other patients were not re-treated because they had low disease activity. Seven of the 24 patients who qualified for re-treatment had not exhibited clinical improvement after the first treatment course. These patients typically did not respond to subsequent courses of rituximab. Of interest, in the 17 patients who had exhibited a clinical response to the first course of rituximab, the second and third treatment courses resulted in European League Against Rheumatism responses similar to those observed after the first course, and no major relapses occurred before re-treatment. CONCLUSION: Rituximab re-treatment is not effective in patients who do not exhibit clinical improvement after the first treatment course, which is consistent with the notion that such patients represent a different pathogenetic subset of RA. Patients who initially responded to rituximab treatment experienced sustained benefit from DAS28-based systematic re-treatment according to the international consensus statement.

Early effects of rituximab on the synovial cell infiltrate in patients with rheumatoid arthritis.

OBJECTIVE: To study the specific effects of rituximab treatment on the synovium in patients with rheumatoid arthritis (RA) early after initiation of treatment. METHODS: Seventeen RA patients underwent an arthroscopic synovial biopsy procedure directly before and 1 month after receiving 2 infusions of the chimeric anti-CD20 monoclonal antibody rituximab (1,000 mg on days 1 and 15; both without methylprednisolone premedication). Immunohistochemical analysis was performed to characterize the cell infiltrate. Stained tissue sections were analyzed by digital image analysis. Statistical analysis was performed using Wilcoxon's signed rank test. RESULTS: No significant change in the Disease Activity Score 28-joint assessment was found at 4 weeks after the first rituximab infusion. At 2 and 4 weeks after infusion, B cells in peripheral blood were almost completely depleted. Most B cells in the synovium were found in large lymphocyte aggregates. Interestingly, a significant reduction in B cell numbers at sites of inflammation was observed 4 weeks after treatment (median 26 cells/mm(2) [interquartile range 4-150] before treatment and 11 cells/mm(2) [interquartile range 0-29] after treatment; P < 0.02). B cells disappeared completely in 3 patients, whereas there was partial depletion in 11 patients. In the other 3 patients, no B cells were present in biopsy tissues obtained either pretreatment or posttreatment. No reductions in other synovial cell populations were observed at 4 weeks. CONCLUSION: Rituximab treatment leads to a rapid and significant decrease in synovial B cell numbers, but not in all patients. Whether the variable tissue response is related to the clinical response over time remains to be clarified.

How to diagnose rheumatoid arthritis early: a prediction model for persistent (erosive) arthritis.

OBJECTIVE: To develop a clinical model for the prediction, at the first visit, of 3 forms of arthritis outcome: self-limiting, persistent nonerosive, and persistent erosive arthritis. METHODS: A standardized diagnostic evaluation was performed on 524 consecutive, newly referred patients with early arthritis. Potentially diagnostic determinants obtained at the first visit from the patient's history, physical examination, and blood and imaging testing were entered in a logistic regression analysis. Arthritis outcome was recorded at 2 years' followup. The discriminative ability of the model was expressed as a receiver operating characteristic (ROC) area under the curve (AUC). RESULTS: The developed prediction model consisted of 7 variables: symptom duration at first visit, morning stiffness for > or =1 hour, arthritis in > or =3 joints, bilateral compression pain in the metatarsophalangeal joints, rheumatoid factor positivity, anti-cyclic citrullinated peptide antibody positivity, and the presence of erosions (hands/feet). Application of the model to an individual patient resulted in 3 clinically relevant predictive values: one for self-limiting arthritis, one for persistent nonerosive arthritis, and one for persistent erosive arthritis. The ROC AUC of the model was 0.84 (SE 0.02) for discrimination between self-limiting and persistent arthritis, and 0.91 (SE 0.02) for discrimination between persistent nonerosive and persistent erosive arthritis, whereas the discriminative ability of the American College of Rheumatology 1987 classification criteria for rheumatoid arthritis was significantly lower, with ROC AUC values of 0.78 (SE 0.02) and 0.79 (SE 0.03), respectively. CONCLUSION: A clinical prediction model was developed with an excellent ability to discriminate, at the first visit, between 3 forms of arthritis outcome. Validation in other early arthritis clinics is necessary.