I. de la Torre

Experimental evidence for the co-evolution of hominin tool-making teaching and language

Hominin reliance on Oldowan stone tools – which appear from 2.5mya and are believed to have been socially transmitted – has been hypothesised to have led to the evolution of teaching and language. Here we present an experiment investigating the efficacy of transmission of Oldowan tool-making skills along chains of adult human participants (N=184) using 5 different transmission mechanisms. Across six measures, transmission improves with teaching, and particularly with language, but not with imitation or emulation. Our results support the hypothesis that hominin reliance on stone tool-making generated selection for teaching and language and imply that (i) low-fidelity social transmission, such as imitation/emulation, may have contributed to the ~700,000 year stasis of the Oldowan technocomplex, and (ii) teaching or proto-language may have been pre-requisites for the appearance of Acheulean technology. This work supports a gradual evolution of language, with simple symbolic communication preceding behavioural modernity by hundreds of thousands of years.

Translational Mini-Review Series on B Cell-Directed Therapies: The pathogenic role of B cells in autoantibody-associated autoimmune diseases – lessons from B cell-depletion therapy

B cell depletion therapy with rituximab (BCDT) is a licensed treatment for rheumatoid arthritis and has shown promising results in the treatment of severe, refractory patients with other autoantibody‐associated autoimmune diseases (AAID). The exact role that B cells play in the pathogenesis of AAID and consequently the mechanisms by which BCDT is effective are not known. The two more widely discussed hypotheses are that BCDT is effective because it removes the precursors of plasma cells producing pathogenic autoantibody species, or because it depletes a critical mass of autoreactive B cell clones that present antigen to pathogenic autoreactive T cells. This review will focus on the effects of BCDT and whether the response of patients with AAID to BCDT could be due ultimately to its effects on autoantibodies. A better knowledge of the main role that B cells play in the pathogenesis of the different diseases and a better understanding of the most likely mechanism of relapse following an earlier response to BCDT would help to guide further developments of B cell targeting therapies and potentially increase the chance of designing a protocol that could induce a long‐term remission.

B-cell-activating factor receptor expression on naive and memory B cells: relationship with relapse in patients with rheumatoid arthritis following B-cell depletion therapy

Objectives To examine the expression of B-cell-activating factor receptor (BAFF-R) on naive CD27− and memory CD27+ B cells in normal individuals and patients with rheumatoid arthritis (RA) undergoing B-cell depletion therapy with rituximab. Patients and Methods BAFF-R expression on B-cell subsets was determined in normal controls (NC; n=11), active patients with RA pre-rituximab (pre-RX; n=15), relapsing patients either concordant for B-cell repopulation (C-R, n=13) or discordant, with relapse more than 3 months after repopulation (D-R, n=11) and patients in remission over 3 months postrepopulation (discordant non-relapsing (D-NR), n=5). Serum BAFF was measured by ELISA and analysed using Mann–Whitney. Results There was no significant difference between NC, pre-RX and D-NR patients in %BAFF-R-positive B cells or mean fluorescence intensity (MFI) in naive and memory B cells. Relapsing patients had significantly lower MFI and %BAFF-R-positive cells in both naive and memory compartments from NC and pre-RX (C-R and D-R; p<0.01). BAFF levels in pre-RX patients were within the normal range and did not correlate with BAFF-R expression in any patient group. D-NR patients had relatively lower proportions of pre and postswitch CD27+ B cells than pre-RX patients (D-NR vs pre-RX; p<0.05 for both) and also lower numbers of postswitch B cells than D-R patients (D-NR vs D-R, p<0.05). Conclusion BAFF-R expression was significantly reduced on both naive and memory B cells in patients at relapse, regardless of the relationship with B-cell repopulation or serum BAFF levels. Re-establishment of active disease was also associated with an increase in class-switch recombination. Factors responsible for lower levels of BAFF-R may relate to altered thresholds for autoreactive B-cell generation at relapse in patients with RA.

Effect of rituximab on B cell phenotype and serum B cell-activating factor levels in patients with thrombotic thrombocytopenic purpura.

Autoantibodies inhibiting the activity of the metalloproteinase, ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13), underlie the pathogenesis of thrombotic thrombocytopenic purpura (TTP). Rituximab (RTX) combined with plasma‐exchange (PEX) is an effective treatment in TTP. Patients can remain in remission for extended periods following PEX/RTX, and this is associated with continuing reduction in antibodies to ADAMTS13. Factors controlling B cell differentiation to autoantibody production, including stimulation through the B cell receptor and interactions with the B cell‐activating factor (BAFF), may thus impact length of remission. In this cross‐sectional study, we measured naive and memory B cell phenotypes [using CD19/immunoglobulin (Ig)D/CD27] following PEX/RTX treatment in TTP patients at B cell return (n = 6) and in 12 patients in remission 10–68 months post‐RTX. We also investigated relationships among serum BAFF, soluble CD23 (sCD23– a surrogate measure of acquiring B memory (CD27+) phenotype) and BAFF receptor (BAFF‐R) expression. At B cell return after PEX/RTX, naive B cells predominated and BAFF‐R expression was reduced compared to healthy controls (P < 0·001). In the remission group, despite numbers of CD19+ B cells within normal limits in most patients, the percentage and absolute numbers of pre‐switch and memory B cells remained low, with sCD23 levels at the lower end of the normal range. BAFF levels were correlated inversely with BAFF‐R expression and time after therapy. In conclusion, the long‐term effects of RTX therapy in patients with TTP included slow regeneration of memory B cell subsets and persistently reduced BAFF‐R expression across all B cell subpopulations. This may reflect the delay in selection and differentiation of potentially autoreactive (ADAMTS13‐specific) B cells, resulting in relatively long periods of low disease activity after therapy.

Clinical usefulness of serum level of adalimumab, in patients with rheumatoid arthritis.

Objectives To analyze the clinical relevance, in clinical practice, of adalimumab (ADA) serum levels (SL) and anti-ADA antibodies (anti-ADA-Abs). 2. To evaluate if there is correlation between SL of ADA and result of DAS28. 3. To determine the minimum appropriate SL of ADA to keep the patients in remission or in low clinical activity. Methods Serum levels of ADA and anti-ADA-Abs (ELISA kit. Promonitor®-ADA. Proteomika, Derio. Vizcaya. Spain) were analyzed in patients with rheumatoid arthritis (RA) receiving ADA >6 months. Cut-off level for serum Abs anti-ADA was >32 U/mL and for serum level of ADA <0.004 mg/L. Clinical characteristics, clinical activity index (DAS in 28 joints), using ESR, were recorded. All the patients were receiving DMARD (methotrexate, leflunomide or hydroxychloroquine). Serum samples were collected before injection of ADA (same day), and stored frozen until analysis. Patients were considered on clinical remission if they had at the same time of extraction, DAS28≤2,6, and low clinical activity if DAS28 between 2,7-3,2. The patients was distributed in tertiles groups from serum levels of ADA: <2,8 mg/L; 2,9-7,3 mg/L; >7,3 mg/L. ROC curvewas used to select optimus cut-off level of ADA to keep the patients on remission or low activity level of disease. Finally, correlation between DAS28 and SL of ADA was evaluated. Results We included 63 determinations from 56 patients with RA. 75% were women; mean age: 62 years. The average time of evolution of RA was 156±122 months, and for the treatment of ADA 32,26±18,31months. ADA was the first anti-TNF received in 80% of patients. The distribution of DMARDs: methotrexate: 65% (mean dose: 15 mg), leflunomide: 21% (18 mg) and hydroxychloroquine: 14% (200 mg). In 4 (7%) patients anti-ADA Abs was detected; all in the group of SL of ADA <2,8 mg/L. We obtained a negative relation between SL of ADA and DAS28 (r: -0.46. CI 95%: -0.66,-0.21). The cut-off of SL of ADA in ROC curves was, for DAS28≤2,6: 3,01 (AUC: 65,77%; sensitivity: 50% y specificity: 77,77%); for DAS28 2,7-3,2: 3,48 (AUC: 83,18%; sensitivity: 83,33% y specificity: 77,80%). Table show the relation of SL of ADA anti-ADA Abs and DAS28-ESR, according ADA tertiles. Conclusions The cut-off of ROC curve for serum level of ADA, to keep the patients in low activity of disease is 3,45 mg/L. 2. There is a negative correlation between the serum level of ADA and DAS28. 3. Serum level of ADA >7,3 mg/L does not increase the improvement of DAS28. In these patients, we can consider to decrease the ADA dose or its delay. 4. The prevalence of anti-ADA Abs in patients with RA treated with ADA and DMARD is 7%. Disclosure of Interest None Declared

AB0396 Infliximab and Adalimumab Levels and Antidrug Antibodies Detection in Patients with Rheumatoid Arthritis (RA): an Interlaboratory Comparison Using A Commercial ELISA Assay

Background The assessment of biological drug levels and immunogenicity might be essential in terms of a more effective and rational use of biological therapies and it is dependent upon the establishment of efficient standardized assays or a consensus that could allow a direct comparison of drug levels and anti-drug antibodies (ADA) data with clinical outcome. Objectives To determine whether an enzyme-linked immunosorbent assay (ELISA) performed with two versions of a commercial kit to assess IFX and ADL levels and ADA yield similar results. Methods The diagnostic capability of two ELISA versions [Promonitor® IFX R1 and R2 (V.1), Promonitor® IFX and Anti-IFX (V.2); Promonitor® ADL R1 and R2 (V.1), Promonitor® ADL and Anti-ADL (V.2) kits (Progenika Biopharma, Spain)] was evaluated in patients with RA treated either with infliximab (IFX; n=24) or adalimumab (ADL; n=24) by three different laboratories. The reliability was determined using the Cohens Kappa coefficient (K), the Pearsons r, the intraclass correlation coefficient (ICC) and the Lins concordance correlation coefficient (CCC). The Bland-Altman plots of differences between V.1 and V.2 were drawn to compare values of each assay. Results The qualitative discordant results for IFX levels V.1 were 9/24 samples (K= good) and 2/23 V.2 (K= very good), for ADL levels V.1 were 7/24 (K= moderate) and for V.2 were 0/24 (K= very good). For IFX-ADA were 0/24 in V.1 and V.2 (K= excellent), while for ADL-ADA were 1/24 in V.1 and 4/24 in V.2 (K= from very good to good). The quantitative agreement is shown in table 1, we found a good linear association using the Pearsons r, the ICC was questionable to excellent for both versions in IFX and ADL levels. The CCC results were poor in all determinations. Bland-Altman plots for both, IFX and ADL levels demonstrate the differences between both versions (Fig. 1 and 2, respectively). Conclusions We observed a better agreement in the qualitative than in the quantitative results in both, for IFX and ADL levels and for IFX-ADA and ADL-ADA. We have found a good linear association (Pearsons r) but a low agreement (K, ICC and CCC) comparing results for V.1 and V.2. Further interlaboratory investigations are necessary to improve results and determine their possible clinical application. Disclosure of Interest L. Valor: None declared, D. Hernández Flόrez: None declared, I. de la Torre: None declared, F. Llinares: None declared, J. Rosas: None declared, J. Yaque: None declared, E. Naredo Grant/research support: UCB and MSD, Consultant for: Abbvie, Roche Farma, Bristol-Myers Squibb, Pfizer, UCB, General Electric Healthcare, and Esaote, C. Gonzalez: None declared, J. Lόpez-Longo: None declared, I. Monteagudo Consultant for: Abbvie, Roche Farma, Bristol-Myers Squibb, Pfizer, UCB, General Electric Healthcare, MSD and Esaote, M. Montoro: None declared, L. Carreño Perez: None declared DOI 10.1136/annrheumdis-2014-eular.3290

SAT0340 Infliximab Levels and Anti-Infliximab Antibodies Comparison between Two Comercial ELISA Versions in Patients with Ankylosing Spondylitis

Background Ankylosing spondylitis (AS) is a chronic inflammatory disease which can result in invalidating deformities of the joints and spine at an early age. The introduction of tumor necrosis factor (TNF) blocking agents has changed the treatment options in AS. Nevertheless, the reasons for lack or loss of response to infliximab (IFX) are unclear. So far determinations of both, IFX serum levels and the presence of anti-drug antibodies (ADA) anti-IFX have not been standardized for clinical use. Objectives To assess the correlation between two available versions (V.1 and V.2) of a commercial enzyme-linked immunosorbent assay (ELISA) for serum levels of IFX and IFX-ADA in patients with AS. Methods In this cross sectional study we assessed 40 serum samples taken prior to infusion from patients diagnosed with AS treated with IFX for more than 12 months (1st line of biological treatment). IFX levels and IFX-ADA were measured using two different ELISA assays [Promonitor® IFX R1 and R2 (V.1), Promonitor® IFX and Anti-IFX (V.2) (Progenika Biopharma, Spain)] according to manufacturers specifications. The relation comparing V.1 and V.2 for IFX levels and IFX-ADA concentrations was performed using the coefficient of variation (CV), the Cohens Kappa coefficient, the Pearsons r, the intraclass correlation coefficient (ICC) and the Lins concordance correlation coefficient (CCC). Bland-Altman plots were drawn to compare both versions of the assays. Results As shown in the table below, we found a greater CV for IFX levels than for IFX-ADA. Regarding the qualitative results the Cohens Kappa was from moderate to fair and considering the quantitative results the Pearsons r was low for IFX levels and high for IFX-ADA, the ICC was questionable for both versions and both determinations. We also determined the CCC and the results showed a poor agreement. Bland-Altman plots showed the difference between both versions for IFX levels (Fig. 1). Conclusions A low reliability for IFX levels and IFX-ADA was obtained for both versions. There is a need to standardize laboratory techniques (variability inter/intra-assay and inter/intra-laboratory) in order to validate this information and its possible clinical application. Disclosure of Interest D. Hernández Flόrez: None declared, L. Valor: None declared, J. C. Nieto: None declared, L. Martinez: None declared, I. de la Torre: None declared, T. del Río: None declared, C. Gonzalez: None declared, J. Lopez-Longo: None declared, I. Monteagudo Consultant for: Abbvie, Roche Farma, Bristol-Myers Squibb, Pfizer, UCB, General Electric Healthcare, and Esaote, E. Naredo Grant/research support: UCB and MSD, Consultant for: Abbvie, Roche Farma, Bristol-Myers Squibb, Pfizer, UCB, General Electric Healthcare, and Esaote, M. Montoro: None declared, L. Carreño Perez: None declared DOI 10.1136/annrheumdis-2014-eular.3274

FRI0042 BAFF-R expression in naÏve CD5+IGM+ B cells in rheumatoid arthritis patients repopulating after rituximab

Background Serum levels of B cell activating factor (BAFF) rise following Rituximab (RTX) therapy in patients with Rheumatoid arthritis (RA). CD5+IgM+B cells are present within transitional and naïve B cell subsets and their increased production or accumulation is associated with some autoimmune diseases. Previous studies have shown that BAFF does not enhance their survival compared with CD5- naïve B cells, suggesting that signalling pathways are important in promoting their survival. Objectives To determine serum BAFF levels and BAFF-receptor (BAFF-R) expression in CD5+IgM+B cells in healthy controls (HC), RTX-naïve RA patients (pre-RTX), and relapsing at different time points after peripheral B cell repopulation post-RTX treatment, divided into 2 groups: early relapsers (0–3 months post-B cell return) and later relapsers (>4 months post-B cell return). Methods Immunophenotyping of peripheral blood mononuclear cells was used to determine %CD5+IgM+B cells and BAFF-R (% and expression, mean fluorescence intensity (MFI)) in 5 HC, 13 pre-RTX and 12 post-RTX RA patients. Results were analyzed with respect to timing of relapse after peripheral B cell return (≥5 B cells/μL) and serum BAFF levels. Results %CD5+IgM+B cells, but not absolute numbers, were significantly higher in post-RTX early relapsers compared to HC (p<0.01), pre-RTX patients (p<0.001) and post-RTX later relapsers (p<0.01). There was a strong inverse correlation between %CD5+IgM+B cells and time after B cell return (r2=0.88, p<0.0001). BAFF-R+ expression was significantly lower in both post-RTX groups compared to HC and pre-RTX patients; early relapsers showed the lowest % and MFI BAFF-R+ expression, compared with later relapsers (p<0.01). BAFF-R+ expression increased with time after B cell return, both % (r2=0.47, p<0.002) and MFI (r2=0.76, p<0.0004). BAFF levels were significantly higher in both post-RTX groups compared to HC and pre-RTX patients, with the highest BAFF levels in early relapsers (p<0.05 compared to later relapsers). There was a significant inverse correlation between BAFF levels and % (r2=0.51, p<0.01) and MFI (r2=0.4, p<0.05) BAFF-R+ expression.Table 1 %CD5+IgM+ (median;range) %BAFF-R+ve BAFF-R+ve (MFI) BAFF levels (ng/ml) (median;range) HC 18.5 (9.1–26.8) 98.7 (97.9–99.5) 62.5 (50.5–82.4) 1.1 (0.9–1.2) Pre-RTX 5.4 (1.9–21.4) 97.3 (85.8–100) 58 (34.2–139) 1.4 (0.9–1.7) Post-RTX early relapse 44.4 (39.6–55.8) **###

FRI0565 Influence of Route of Administration/drug Formulation and Other Factors on Compliance with Treatment in Rheumatoid Arthritis and Dyslipidaemia

60.6 (12.9–71.2)**###