Immunomodulatory potential of anti-IFN-beta antibodies on monocyte-derived macrophages.

Introduction: Multiple sclerosis (MS) is a disabling neurological disease with an unknown etiology, where the recombinant interferon beta (rIFNß) is the most established treatment. However, the development of anti-IFNß antibodies has posed a significant therapeutic drawback. In this study, the interaction between anti-IFNß antibodies and macrophages was investigated to assess the effects on the immune system. Methodology: Using magnetic beads, anti-IFNß antibodies were extracted from MS patients' sera positive for anti-IFNß antibodies. A negative control (antibody-negative situation) and a baseline control were obtained in parallel. Bead or extracted beadantibody complexes were then incubated vitro with monocyte-derived human macrophages. After incubation, macrophage cultures were tested for 91 immunologically relevant gene expressions by RT-PCR. Results and Discussions: A Gene expression difference between antibody positive and negative situations was hypothesized to reflect the direct effects between antibodies and macrophages. Thus, 37-39 genes were either up-regulated or downregulated due to this direct interaction. Of these, only 2-4 genes were up-regulated, and the rest were down-regulated. These observations suggest that anti-IFNß antibodies have an overall suppressive effect on immunologically relevant gene activity when antibodies interact with macrophages. Conclusion: The fate and effects of circulating anti-IFNß antibodies are mainly unknown. With the observations obtained at in vitro level, such effects, especially from an immunological point of view, are suppressive on immunocompetent cells such as macrophages. However, in vivo verification is necessary.

Validity of clinical disease activity index (CDAI) to evaluate the disease activity of rheumatoid arthritis patients in Sri Lanka: A prospective follow up study based on newly diagnosed patients.

Routine use of the Disease Activity Score-28 (DAS28) to assess the disease activity in rheumatoid arthritis (RA) is limited due to its dependency on laboratory investigations and the complex calculations involved. In contrast, the clinical disease activity index (CDAI) is simple to calculate, which makes the "treat to target" strategy for the management of RA more practical. We aimed to assess the validity of CDAI compared to DAS28 in RA patients in Sri Lanka. A total of 103 newly diagnosed RA patients were recruited, and their disease activity was calculated using DAS 28 and CDAI during the first visit to the clinic (0 months) and re-assessed at 4 and 9 months of follow-up visits. The validity of the CDAI, compared to DAS 28, was evaluated. Patients had a female preponderance (6:1) and a short symptom duration (mean = 6.33 months). Internal consistency reliability of CDAI, as assessed by Cronbach's α test, was 0.868. Convergent validity was assessed by correlation and Kappa statistics. Strong positive correlations were observed between CDAI and DAS 28 at the baseline (0 months), 4 and 9 months of evaluation (Spearman's r = 0.935, 0.935, 0.910, respectively). Moderate-good inter-rater agreements between the DAS-28 and CDAI were observed (Weighted kappa of 0.660, 0.519, and 0.741 at 0, 4, and 9 months respectively). Discriminant validity, as assessed by ROC curves at 0, 4th, and 9th months of the evaluation, showed the area under the curve (AUC) of 0.958, 0.979, and 0.910, respectively. The suggested cut-off points for different CDAI disease activity categories according to ROC curves were ≤ 4 (Remission), > 4 to ≤ 6 (low), > 6 to ≤ 18 (moderate), > 18 (high). These findings indicate that the CDAI has good concordance with DAS 28 in assessing the disease activity in RA patients, in this study sample.

Potential applicability of cytokines as biomarkers of disease activity in rheumatoid arthritis: Enzyme-linked immunosorbent spot assay-based evaluation of TNF-α, IL-1ß, IL-10 and IL-17A.

Biomarkers play a pivotal role in the management of rheumatoid arthritis (RA) by facilitating early diagnosis and 'treat to the target.' However, no gold standard biomarker has been identified for monitoring the disease activity in RA. Cytokines, a diverse group of small protein molecules secreted by peripheral blood mononuclear cells (PBMCs), play a pivotal role in pathogenesis and disease progression in RA. Research is currently underway to find out the applicability of cytokines as biomarkers in RA. This study aimed to quantify the PBMCs that secrete four types of cytokines; TNF-α, IL-1ß, IL-10 and IL-17A in two cohorts of active RA patients (early RA patients and established RA patients), compared to healthy controls (HC), using the enzyme-linked immunosorbent spot (ELISPOT) assay, and to assess their association with measures of disease activity of RA. Patients were recruited from outpatient rheumatology clinics, and the disease activity was assessed using single and composite measures of disease activity. The cytokine expression was evaluated using freshly separated PBMCs from whole blood of RA patients using the ELISPOT assay. The number of PBMCs (counted as spot-forming cells (SFCs) per 105 PBMCs) that secreted the cytokine of interest were statistically significantly higher in early RA patients, compared to HC, for IL-17A (P<0.05). Such an increased number of SFCs was not observed in the established RA group, compared to controls, for any of the cytokines tested. The correlation analysis showed that IL-17A is having a moderate correlation (Spearman`s ρ, p <0.05) with five clinical measures of disease activity, including disease activity score 28 (DAS28). According to the multivariable linear regression models, IL17A was a good predictor of both the disease activity score 28 (DAS28) and clinical disease activity index (CDAI). In conclusion, IL-17A has potential applicability as a biomarker of disease activity of RA.

Prevalence of anti-drug antibodies against interferon beta has decreased since routine analysis of neutralizing antibodies became clinical practice.

BACKGROUND: Neutralizing antibodies (NAbs) against interferon beta (IFNß) lead to loss of treatment efficacy in multiple sclerosis patients. The seroprevalence of NAbs in multiple sclerosis patients treated with IFNß during 2003-2004 was 32% in a cross-sectional analysis of routine data. OBJECTIVES: The aim of this study was to investigate whether the seroprevalence of NAbs, the levels of NAb titres and the IFNß preparations used for treatment of multiple sclerosis patients had changed in 2009-2010. METHODS: This study included 1296 patients, analysed for NAbs with the myxovirus resistance protein A gene expression assay in 2009-2010. RESULTS: The seroprevalence of NAbs had decreased to 19% in 2009-2010, which is significantly lower compared with the previous study in 2003-2004 (p<0.0001). This decrease was attributed to the IFNß-1a preparations only, not to IFNß-1b. The frequency of patients with high positive titres decreased the most, from 16% to 7% (p<0.0001). CONCLUSIONS: NAb seroprevalence has decreased since NAb monitoring became clinical practice in 2003, especially for patients with high NAb titres. This might be due to the stricter monitoring of NAb titres that prompt NAb positive patients to stop treatment, to preferential use of less immunogenic drugs and to alteration of drug formulations.

Inhibition of endogenous interferon beta by neutralizing antibodies against recombinant interferon beta.

OBJECTIVE: To determine if neutralizing antibodies (NAbs) against interferon beta from patients with multiple sclerosis (MS) cross-react with other type 1 interferons, especially endogenous interferon beta, and thus might impede the immune systems of affected patients. DESIGN: Masked serum samples from MS patients were challenged in vitro against recombinant interferon beta-1a and interferon beta-1b, as well as human leukocyte interferon and fibroblast interferon, the latter representing endogenous interferon. The neutralizing capacity of serum samples on these type 1 interferons was assessed using a luciferase reporter gene assay. Randomly selected samples were titrated to further delineate the cross-reactivity of antibodies. SETTING: University medical center in Düsseldorf, Germany. PATIENTS: We randomly selected 150 samples from interferon beta-treated MS patients who had previously been tested for the presence of binding antibodies and NAbs. MAIN OUTCOME MEASURES: Neutralization of interferon beta bioactivity and cross-reactivity of anti-interferon beta antibodies. RESULTS: Antibody-mediated neutralization of interferon beta bioactivity in vitro against recombinant interferon beta was observed in all serum samples that had previously tested positive for binding antibodies and NAbs. A neutralizing pattern comparable to that of recombinant interferon beta was observed when endogenous interferon was assessed, reflecting cross-reactivity of NAbs. No differences in neutralization between recombinant and endogenous interferon were observed with respect to the interferon beta preparation used for treatment. Furthermore, no neutralization of other type 1interferons by NAbs could be detected. CONCLUSIONS: A proportion of MS patients who are treated with recombinant interferon beta develop NAbs that also neutralize endogenous interferon. Because NAbs at high titers can persist for years, these antibodies may impede the immune system in affected MS patients regardless of their current treatment regimen.