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Objective: To report the efficacy and safety of anti-TNF agents in patients with severe and/or refractory manifestations of Behçet's disease (BD)., Methods: We performed a multicenter study of main characteristics and outcomes of anti-TNF alpha treatments [mainly infliximab (62%), and adalimumab (30%)] in 124 BD patients [48% of men; median age of 33.5 (28-40) years]., Results: Overall response (i.e. complete and partial) rate was 90.4%. Clinical responses were observed in 96.3%, 88%, 70%, 77.8%, 92.3% and 66.7% of patients with severe and/or refractory ocular, mucocutaneous, joint, gastro-intestinal manifestations, central nervous system manifestations and cardiovascular manifestations, respectively. No significant difference was found with respect to the efficacy of anti-TNF used as monotherapy or in association with an immunosuppressive agent. The incidence of BD flares/patient/year was significantly lower during anti-TNF treatment (0.2 ± 0.5 vs 1.7 ± 2.4 before the use of anti-TNF, p < 0.0001). The prednisone dose was significantly reduced at 6 and 12 months (p < 0.0001). In multivariate analysis, retinal vasculitis was negatively associated with complete response to anti-TNF (OR = 0.33 [0.12-0.89]; p = 0.03). The efficacy and relapse free survival were similar regardless of the type of anti-TNF agent used. After a median follow-up of 21 [7-36] months, side effects were reported in 28% of patients, including infections (16.3%) and hypersensitivity reactions (4.1%). Serious adverse events were reported in 13% of cases., Conclusion: Anti-TNF alpha therapy is efficient in all severe and refractory BD manifestations. Efficacy appears to be similar regardless of the anti-TNF agent used (infliximab or adalimumab)., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Mixed cryoglobulinemia is usually considered to be a nonmalignant human B cell proliferation that produces a monoclonal IgM rheumatoid factor (RF). Important immunologic similarities and differences were described between the monoclonal B cells during mixed cryoglobulinemia and during malignant chronic lymphocytic leukemia (CLL):high frequency of the same VH and V kappa gene usage by both types of monoclonal B cells producing IgM with RF activity, apparent intraclonal homogeneity, but different expression of the pan T cell CD5 Ag. The description of an unusual CD5-negative B cell CLL case secreting a mutated IgM RF led the authors to suggest that the usage of non-mutated germline Ig genes is a property of cells derived from the CD5 lineage or stage of differentiation, rather than an intrinsic property of CLL or of IgM RF-producing cells in general. Because mixed cryoglobulinemia cells are usually CD5-negative, it was of interest to test for the existence of mutations in the VH and V kappa regions, as well as for the intraclonal homogeneity of the expressed Ig genes. In this study, we used the PCR technique to analyze the monoclonal rheumatoid factor (mRF) V genes from a patient with mixed cryoglobulinemia. We show that the CD5-negative monoclonal B cells express a slightly mutated V kappa 3 gene, but a more mutated VH1 gene whose genomic counterpart was shown to be the 51p1 germline gene. The sequence analysis of several independent clones shows some degree of intraclonal diversity, suggesting the existence of a clonal filiation. These results are discussed in terms of the origin of the monoclonal B cell during mixed cryoglobulinemia and CLL.

Polyreactive antibodies able to bind various apparently unrelated structures represent a frequent antibody population in autoimmune diseases. In this work, the structural basis of the double reactivity of such autoantibodies was investigated using as models polyclonal and monoclonal human rheumatoid factors (RF) reacting with histones. Both direct ELISA binding and competitive inhibition experiments were performed. A more precise delineation of the histone regions recognized by the RFs was made by means of 27 synthetic peptides of these proteins. Anti-idiotope (Id) murine antibodies were used to map the binding sites involved on RF in the interaction with IgG and histones. Among the 13 polyclonal and six monoclonal RFs tested, four and two respectively were found to cross-react with IgG and histones. The fragments shown to be the most frequently recognized by RFs were located in residues 1-16 and 204-218 of H1, 1-20 and 65-85 of H2A, and 1-21 of H3. The results obtained by competitive ELISA assays using IgG, histone peptides and anti-Id monoclonal antibodies led us to confirm and characterize more precisely our previous finding suggesting the existence of topographically distinct binding sites for the different targets recognized by RFs.

Different human IgM rheumatoid factor (IgM RF) idiotypes have been described defined by polyclonal rabbit anti-idiotypic antibodies. These antisera do not allow clear genetic analysis of the idiotypic determinants, be they cross-reactive or private. Therefore, we tried to obtain a set of monoclonal anti-idiotypic antibodies directed against RF idiotypes. Purified IgM RF serum from a patient with classical rheumatoid arthritis was used to immunize BALB/c mice. The spleen cells were fused with Sp 2/0 Ag 14, a nonsecreting mouse myeloma cell line, and a hybrid producing monoclonal anti-idiotypic antibody was selected. The mouse antibody, an IgG1 kappa, reacts with an identical or similar determinant located on (or close to) the binding site of all tested monoclonal or polyclonal IgM RF from totally unrelated patients with Waldenströms's macroglobulinemias or rheumatoid arthritis. The monoclonal antibody also reacts with 2 rheumatoid arthritis patients' IgG RF and with a low proportion of normal polyclonal IgM without detectable RF activity. An hypothesis is proposed to explain the existence of a such highly conserved determinant on RF idiotypes.

Two mouse monoclonal antibodies (mAb A216-5 and L 49-3) with antiidiotypic activity against two human monoclonal IgM rheumatoid factors (IgM RFs) were defined. Each of these monoclonal antibodies (two mouse IgG 1K) reacted with an idiotope located on the heavy chain of the immunizing monoclonal IgM RF and was able to inhibit RF fixation to the antigen. These monoclonal antibodies did not react with other monoclonal IgM RFs from patients with macroglobulinemias or cryoglobulinemias and, therefore, did not recognize the known cross-reactive idiotopes of human monoclonal RFs. The presence of both 216-5 and 49-3 idiotopes on polyclonal IgM RFs from unrelated patients was undetectable by the inhibition assays. However, using a four-stage solid-phase radioimmunoassay, the 216-5 idiotope (minor), but not the 49-3 idiotope (private), was frequently present at a low concentration on polyclonal IgM RFs from patients suffering from rheumatoid arthritis, primary Sjögren syndromes, various infectious diseases, systemic vasculitis, and sarcoidosis and during aging. Interestingly, the 216-5 idiotope was undetectable among polyclonal IgM RFs of 12 normal adults. The main conclusions of these data are the following. The definition of minor and private idiotopes of human RFs requires the use of assays able to detect low amounts of antibodies among polyclonal Ig. The anti-IgG B cells which are sometimes clonally expanded during Waldenström diseases and cryoglobulinemias can also be activated during nonneoplastic diseases, among the other RF-secreting B cells.(ABSTRACT TRUNCATED AT 250 WORDS)

The potential therapeutic value of anti-idiotypic antibodies during B cell proliferations largely depends on the stability of the target Ig idiotopes. We investigated this stability in a clinical condition of so called nonmalignant monoclonal B cell proliferation, mixed cryoglobulinemia. The idiotypic profile of a single IgM kappa monoclonal auto-antibody with anti-IgG activity (rheumatoid factor (RF] which originated from a patient suffering from a nonmalignant mixed cryoglobulinemia was followed over a period of 3 yr. As judged from the reactivity of a panel of five different mouse monoclonal anti-idiotypic antibodies mapping the RF variable regions, there was no idiotypic change in the serum IgM RF. At a cellular level, in vitro stimulation of the patient's PBL gives rise to IgM kappa auto-antibodies that were shown to bear the same idiotypic determinants as the serum IgM kappa. We then investigated the effects of the anti-idiotypic antibodies on the in vitro IgM kappa production. When stimulated with PWM and in the presence of anti-idiotypic antibodies (10 micrograms/ml), the patient's PBL produced less IgM RF (18 to 62% inhibition). The same inhibition of IgM RF production was observed after EBV infection of the patient's PBL (from 19 to 90% inhibition). In both cases, the remaining IgM RF production was idiotypically indistinguishable from the serum IgM RF. The implications of the idiotypic stability and of the results of in vitro idiotypic manipulation could be important in view of both the understanding of nonmalignant cryoglobulinemia and of the possible therapeutic use of anti-idiotypic antibodies in diseases.

In an effort to analyze both IgM rheumatoid factor (RF) repertoire and regulation of RF production in humans, we developed a panel of four mouse monoclonal antibodies (mAb) defining distinct K light chain-associated idiotopes (id) of a human monoclonal IgM RF (Alt). These mAb (A75, AM1, AM2, AM3) had equivalent reactivities with the immunizing RF during classic inhibition of antigen-binding assays. These anti-id reagents were reacting to neither other tested monoclonal IgM RF nor normal polyclonal IgM. It was possible to distinguish the id defined by the mAb from the results of four sets of experiments: dissociation of Alt RF heavy (H) and light (L) chains showed that A75, AM1, and AM2 reacted to id located on the L chain, whereas AM3 defined a conformational RF id; recombination experiments of H and L chains showed that A75 and AM2 reacted well with both homologous (Alt H + Alt L) and heterologous (Alt L + unrelated H) recombinants, whereas AM1 reacted better with the homologous recombinant than with the heterologous one; the relative affinities of the mAb were drawn from their ability to shift already bound labeled Alt RF from solid phase IgG; and radiolabeling of two mAb (A75 and AM3) and experiments of inhibition of id binding with cold anti-id and cold anti-CK showed that A75 recognized a proximal id (close to the K constant region), whereas AM3 defined a more distal id, AM2 and AM1 being located between A75- and AM3-defined sites. This topographic mapping of K light chain-associated id of a human RF with anti-id of known relative affinities could help in studying idiotypic regulation in humans.

The structural basis of the double reactivity of a human monoclonal rheumatoid factor (RF) with both human IgG and histones H1 and H3 was investigated by means of competitive inhibition experiments. The monoclonal RF binding to solid-phase histones was inhibited by increasing concentrations of heat-aggregated IgG. However, increasing concentrations of purified histones were almost unable to reduce the RF binding to solid-phase IgG. Inhibition of antigen binding with two murine monoclonal anti-idiotopes reacting with distinct idiotopes on the monoclonal RF indicated that the fixation to the different antigens was mediated by distinct binding sites. This result was confirmed by showing the selective sensitivity to mild acid treatment of the histone binding site but not of the IgG binding site. This report provides a structural basis for the existence of polyfunctional combining regions on a human autoantibody.