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Background: Pemphigus vulgaris (PV) is characterized by autoantibodies targeting keratinocyte adhesion proteins desmoglein (Dsg) 1 and 3, and by the human leukocyte antigen (HLA) predisposition allele HLA-DRB1*0402. Treatment using rituximab (RTX) combined with short-term corticosteroids (CS) allows disease control and long-lasting remission., Objectives: The principal aim of this study was to evaluate the impact of RTX on the circulating subpopulations of Dsg3-specific T lymphocytes that specifically regulate B-cell responses: follicular helper T (Tfh) and follicular regulatory T (Tfr) lymphocytes., Methods: Using the HLA-DRB1*0402 tetramer loaded with the Dsg3 immunodominant peptide, we used flow cytometry to analyse the frequency, polarization and activation status of blood Dsg3-specific follicular T-cell populations at baseline, month (M) 6 and long-term follow-up (M60-90) from patients with PV., Results: At baseline, we observed a predominance of Tfh1* and Tfh17 subsets and an underrepresentation of the Tfh2 subset among autoreactive Dsg3-specific Tfh cells compared with nonautoreactive Tfh cells. RTX treatment induced a decrease of autoreactive Tfh cells with no effect on their polarization during follow-up. In parallel, we observed the emergence of a Dsg3-specific Tfr subpopulation with a significant overexpression of the surface activation markers PD1, ICOS and CD25 that was not observed at the surface of autoreactive Tfh and nonautoreactive Tfr cells of the same patients with PV. In contrast, very few Dsg3-specific Tfr cells were observed in patients with PV who were treated with CS alone., Conclusions: Here we show that the emergence of circulating autoreactive Dsg3-specific Tfr cells is associated with the long-term efficacy of RTX in patients with PV., Competing Interests: Conflicts of interest V.H. reports administrative support was provided by the French Society for Dermatology and Pathology of Sexually Transmitted Diseases. P.J. reports a relationship with Roche SAS that includes board membership. N.F. reports financial support provided by the European Regional Development Fund, the French Society for Dermatology and Pathology of Sexually Transmitted Diseases, the National Institute of Health and Medical Research, the Agence Nationale de la Recherche (ANR) and the Occitanie Region., (© The Author(s) 2024. Published by Oxford University Press on behalf of British Association of Dermatologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Importance: The Ritux 3 trial demonstrated the short-term efficacy and safety of first-line treatment with rituximab compared with a standard corticosteroid regimen in pemphigus. No data on the long-term follow-up of patients who received rituximab as first line are available., Objective: To assess the long-term efficacy and safety of the Ritux 3 treatment regimen., Design, Setting, and Participants: This 7-year follow-up study of the Ritux 3 trial included patients with pemphigus from 25 dermatology departments in France from January 1, 2010, to December 31, 2015., Exposure: Patients were initially randomized in the rituximab plus prednisone group or prednisone-alone group., Main Outcomes and Measures: The primary outcome was the 5- and 7-year disease-free survival (DFS) without corticosteroids, assessed by Kaplan-Meier curves. Secondary outcomes were occurrence of relapse, occurrence of severe adverse events (SAEs), and evolution of antidesmoglein (Dsg) antibody enzyme-linked immunosorbent assay values to predict long-term relapse., Results: Of the 90 patients in the Ritux 3 trial, 83 were evaluated at the end of follow-up study visit (44 in the rituximab plus prednisone group; 39 in the prednisone-alone group) with a median (IQR) follow-up of 87.3 (79.1-97.5) months. Forty-three patients (93%) from the rituximab plus prednisone and 17 patients (39%) from the prednisone-alone group had achieved complete remission without corticosteroids at any time during the follow-up. Patients from the rituximab group had much longer 5- and 7-year DFS without corticosteroids than patients from the prednisone-alone group (76.7% and 72.1% vs 35.3% and 35.3%, respectively; P < .001), and had about half the relapses (42.2% vs 83.7%; P < .001). Patients who received rituximab as second-line treatment had shorter DFS than patients treated as first line (P = .007). Fewer SAEs were reported in the rituximab plus prednisone group compared with the prednisone-alone group, 31 vs 58 respectively, corresponding to 0.67 and 1.32 SAEs per patient, respectively (P = .003). The combination of anti-Dsg1 values of 20 or more IU/mL and/or anti-Dsg3 values of 48 or more IU/mL yielded 0.83 positive predictive value and 0.94 negative predictive value to predict long-term relapse., Conclusions and Relevance: In this secondary analysis of the Ritux 3 trail, first-line treatment of patients with pemphigus with the Ritux 3 regimen was associated with long-term sustained complete remission without corticosteroid therapy without any additional maintenance infusion of rituximab.

Importance: The clinical relevance of antirituximab antibodies (ARAs) in patients with pemphigus who are treated with rituximab (RTX) is currently unknown., Objective: To determine the prevalence of ARAs in patients with pemphigus who are treated with RTX and their association with complete remission (CR) and relapse., Design, Setting, and Participants: This post hoc analysis of the Ritux3 trial was conducted from January 2010 to December 2015 in 25 dermatology departments in France and included 42 patients with moderate-to-severe pemphigus who were randomized to receive treatment with RTX. Five additional patients were recruited for an ancillary study. The proportions of patients who achieved CR or relapsed after an initial treatment cycle of RTX were compared depending on whether patients had ARAs., Exposures: Patients were treated with 1000 mg of RTX on days 1 and 15 and 2 maintenance infusions of 500 mg at months 12 and 18., Main Outcomes and Measures: Rates of relapse and sustained CR at month 36. Levels of ARAs, antidesmoglein 1/3 antibodies, RTX serum concentrations, and peripheral blood CD19+ B-cell frequency were measured., Results: Of 42 participants with vs without ARAs, the mean (SD) age was 55 (17) years and 56 (17) years, respectively; 25 (59.5%) were women. Antirituximab antibodies were detected in the serum samples of 13 of 42 patients (31%) during the first year. Nine patients who experienced relapse before month 12 were excluded because they received additional infusions and could not be further analyzed. Among the 33 remaining patients, 2 patients (6.1%) experienced relapse after month 12, and 31 (95.9%) maintained a sustained CR until month 36. The rate of sustained CR was not different whether patients had ARAs (11 of 13 [85%]) or not (20 of 20 [100%]) (P = .15). Both groups (ARA+ vs ARA-) also had similar CD19+ B-cell depletion and RTX levels, but patients with ARAs had higher anti-desmoglein 3 antibody (DSG3 Abs) levels compared with those without ARAs (mean [SD], 30.1 [50.9] AU/mL vs 4.0 [4.3] AU/mL; P = .03). The 2 patients with ARAs who experienced relapse after month 12 had an undetectable RTX level, incomplete B-cell depletion, and higher anti-DSG3 Abs level than the 11 patients who maintained a sustained CR with ARAs (RTX mean [SD] concentration, 0 ug/mL vs 12.5 [2.2] ug/mL; P = .03; incomplete B-cell depletion, 2 of 2 vs 4 of 11; P = .19; mean [SD] anti-DSG3 Abs levels, 103.5 [61.5] AU/mL vs 19.5 [11.0] AU/mL; P = .001) or patients without ARAs (mean [SD] RTX concentration, 0 ug/mL vs 13.5 [1.8] ug/mL; P = .02; incomplete B-cell depletion, 2 of 2 vs 5 of 20; P = .09; mean [SD] anti-DSG3 Abs level, 103.5 [61.5] AU/mL vs 4.0 [1.0] AU/mL; P < .001)., Conclusions and Relevance: The results of this cohort study suggest that ARAs are frequently detected in patients with pemphigus who are treated with RTX and generally are not associated with patient outcomes. Only a few patients with the combination of ARAs, low RTX concentration, incomplete B-cell depletion, and persistent serum anti-DSG3 Abs seem at high risk of relapse.

Pemphigus is a life-threatening auto-immune blistering disease of the skin and mucous membrane that is caused by the production of auto-antibodies (auto-Abs) directed against adhesion proteins: desmoglein 1 and 3. We demonstrated in the "Ritux3" trial, the high efficacy of rituximab, an anti-CD20 recombinant monoclonal antibody, as the first-line treatment for pemphigus. However, 25% of patients relapsed during the six-month period after rituximab treatment. These early relapses were associated with a lower decrease in anti-desmoglein auto-Abs after the initial cycle of rituximab. The N- glycosylation of immunoglobulin-G (IgG) can affect their affinity for Fc receptors and their serum half-life. We hypothesized that the extended half-life of Abs could be related to modifications of IgG N- glycans. The IgG N- glycome from pemphigus patients and its evolution under rituximab treatment were analyzed. Pemphigus patients presented a different IgG N- glycome than healthy donors, with less galactosylated, sialylated N- glycans, as well as a lower level of N- glycans bearing an additional N- acetylglucosamine. IgG N- glycome from patients who achieved clinical remission was not different to the one observed at baseline. Moreover, our study did not identify the N- glycans profile as discriminating between relapsing and non-relapsing patients. We report that pemphigus patients present a specific IgG N- glycome. The changes observed in these patients could be a biomarker of autoimmunity susceptibility rather than a sign of inflammation.

Background: Immunoglobulin G (IgG) levels are maintained by the IgG-recycling neonatal Fc-receptor (FcRn). Pemphigus vulgaris and pemphigus foliaceus are debilitating autoimmune disorders triggered by IgG autoantibodies against mucosal and epidermal desmogleins. Recently, a phase 2 clinical trial (NCT03334058; https://clinicaltrials.gov/NCT03334058) was completed in participants with pemphigus using efgartigimod, an FcRn inhibitor, in combination with prednisone. Efgartigimod demonstrated an early effect on diease activity and was well tolerated. In addition to the safety and efficacy assessment, clinical trials present an opportunity to gain more insights into the mechanism of disease, the mode of action of treatment, and potential for corticosteroid-sparing activity., Objective: The aim of our study was to assess the impact of FcRn antagonism by efgartigimod on immunological parameters known to be directly involved in pemphigus pathology, such as cellular and serological responses., Methods: We investigated total and antigen-specific IgG subclass level kinetics during and after treatment, assessed antigen-specific B-cell responses, followed T- and B-cell immunophenotypes, and analyzed how different immunophenotypes link to clinical response., Results: Treatment resulted in reduction of total IgG as well as autoreactive IgG antibody levels. Surprisingly, unlike total IgG and vaccine- or natural-infection-elicited IgG, which returned to baseline levels after stopping efgartigimod treatment, autoreactive antibody levels remained low in several study participants. Efgartigimod showed no effect on total leukocytes, neutrophils, monocytes, or lymphocytes in patients treated with extended efgartigimod therapy. Intriguingly, antigen-specific analyses revealed a loss of desmoglein-specific B cells in several participants responding to efgartigimod, in line with prolonged reduction of pathogenic IgG levels., Conclusions: Efgartigimod treatment of participants with pemphigus improved their conditions and exerted an immunomodulatory effect beyond the blockade of IgG recycling. Further studies in larger populations with an appropriate placebo control are needed to confirm these potentially important observations to establish long-term clinical responses in autoimmune diseases., Competing Interests: ZB-C reports consulting fees for Sanofi-Genzyme, Novartis, and argenx. MG has served as a consultant and on advisory boards for argenx, Biotest, GSK, Janssen, LEO Pharma, Lilly, Novartis, and UCB. MS, BB, and PV are employed by argenx. PJ has served as a consultant for Amgen, Principia Biopharma, argenx, AstraZeneca, Janssen, Thermo Fisher, Lilly, Sanofi, Akari, Chugai, Novartis, Kezar, Genentech, and Topas. GV is a paid consultant for argenx. SC, MM-V, and M-LG as employees of INSERM U1234, Normandie University have a collaborative research agreement with argenx. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. This study received funding from Argenx. The funder had the following involvement with the study: participated in the study design, research, analysis, data collection, interpretation of data, and review and approval of the publication. Argenx also funded the medical writing support of this manuscript., (Copyright © 2022 Maho-Vaillant, Sips, Golinski, Vidarsson, Goebeler, Stoevesandt, Bata-Csörgő, Balbino, Verheesen, Joly, Hertl and Calbo.)

Introduction: We studied the distribution and in vitro pathogenicity of anti-DSG3 IgG subclasses during the course of pemphigus vulgaris (PV)., Methods: We longitudinally studied the distribution of anti-DSG3 IgG subclasses (before versus after treatment) in sera from PV patients, using an addressable-laser bead immunoassay (ALBIA). The in vitro pathogenicity of corresponding sera was tested using keratinocyte dissociation and immunofluorescence assays., Results: Sixty-five sera were assessed at baseline (33 from patients treated with rituximab and 32 with corticosteroids). Sixty-three percent of these baseline sera contained 2 or more anti-DSG3 IgG subclasses versus 35.7% of sera from patients in complete remission (CR) and 75.0% of sera from patients with persistent disease activity after treatment. IgG4 was the most frequently detected anti-DSG3 IgG subclass, both in patients with disease activity and in those in CR. The presence of three or more anti-DSG3 IgG subclasses was predictive of relapse, in particular when it included IgG3, with a positive predictive value of 62.5% and a negative predictive value of 92%. While anti-DSG3 IgG4 Abs from sera collected before treatment were most often pathogenic, anti-DSG3 IgG4 from sera collected after treatment were pathogenic only after adjusting their titer to the one measured before treatment. The IgG3 fraction containing anti-DSG3 Abs also had an in vitro pathogenic effect. The disappearance of the pathogenic effect of some sera after removal of anti-DSG3 IgG3 suggested an additional effect of this IgG subclass., Conclusion: The serum levels and number of anti-DSG3 IgG subclasses drive the pathogenic effect of pemphigus sera and may predict the occurrence of relapses., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Golinski, Lemieux, Maho-Vaillant, Barray, Drouot, Schapman, Petit, Hertl, Boyer, Calbo, Joly and Hébert.)

Pemphigus vulgaris is an autoimmune disease that occurs due to pathogenic autoantibodies that recognize the following epidermal adhesion proteins: desmogleins. Systemic corticosteroids usually decrease the titers of anti-desmoglein autoantibodies and improve patients' conditions. Since modifications of IgG N -glycosylation have been described in some autoimmune diseases, we hypothesized that changes in the pathogenic activity of pemphigus IgG could be related to changes in their N -glycosylation profile. The purpose of this study was to assess, longitudinally, the pathogenicity of pemphigus serum IgG and their N -glycosylation profile during phases of disease activity and clinical remission. The pathogenic activity of serum IgG was measured in vitro on immortalized keratinocytes, by immunofluorescence and dissociation assays, and IgG N -glycans were analyzed by mass spectrometry. We showed (i) a correlation between pemphigus clinical activity and the pathogenicity of serum IgG at baseline and at month 6, while the persistence of the in vitro pathogenic activity of IgG during its evolution, even in patients in clinical remission, seemed to be predictive of relapse; (ii) that modifications of the N -glycan structure were altered the in vitro pathogenicity of patients' autoantibodies; (iii) that the pathogenic properties of pemphigus IgG did not appear to be related to the disparity in IgG N -glycans during the course of pemphigus.

Pemphigus is an autoimmune blistering disease mediated by autoantibodies directed against desmogleins (DSGs). We recently showed that first-line treatment with rituximab (RTX) enables more patients to achieve long-lasting remission off therapy than corticosteroids alone. To understand the immunological mechanisms that mediate long-lasting clinical remission after RTX treatment, we analyzed the phenotype of DSG-specific memory B cells and DSG-specific T follicular helper cells by flow cytometry and measured antibody-secreting cells by enzyme-linked immune absorbent spot in patients treated with corticosteroids alone or RTX. This post hoc analysis of the RITUX3 trial showed that RTX induced a significant decrease of IgG-switched DSG-specific memory B cells. Accordingly, anti-DSG antibody-secreting cells were no longer detected in patients in complete remission after RTX. In contrast, corticosteroids did not modify the frequency or the phenotype of DSG-specific memory B cells, and anti-DSG antibody-secreting cells were still detected after treatment, even in patients in remission. Using peptide-HLADRB1∗0402 tetramer staining, we identified DSG-3-specific T follicular helper cells, which dramatically decreased after RTX, while remaining stable after corticosteroid treatment. Our findings suggest that long-lasting response to RTX in pemphigus relies on the decrease of DSG-specific circulating T follicular helper cells, which correlates with a sustained depletion of IgG-switched memory autoreactive B cells, leading to the disappearance of anti-DSG antibody-secreting cells., (Copyright © 2021 CHU CHARLES NICOLLE ROUEN. Published by Elsevier Inc. All rights reserved.)

The efficacy of the B-cell-depleting agent rituximab has been reported in immune diseases but relapses are frequent, suggesting the need for repeated infusions. The B-cell activating factor (BAFF) is an important factor for B cell survival, class switch recombination and selection of autoreactive B cells, as well as maintaining long-lived plasma cells. It has been hypothesized that relapses after rituximab might be due to the increase of serum BAFF levels. From the Ritux3 trial, we showed that baseline serum BAFF levels were higher in pemphigus patients than in healthy donors (308 ± 13 pg/mL versus 252 ± 28 pg/mL, p=0.037) and in patients with early relapse compared who didn't (368 ± 92 vs 297 ± 118 pg/mL, p=0.036). Rituximab and high doses of CS alone have different effects on the BAFF/BAFF-R axis. Rituximab led to an increase of BAFF levels associated to a decreased mRNA (Day 0: 12.3 ± 7.6 AU vs Month 36: 3.3 ± 4.3 AU, p=0.01) and mean fluorescence intensity of BAFF-R in non-autoreactive (Day 0: 3232 vs Month 36: 1527, mean difference: 1705, 95%CI: 624 to 2786; p=0.002) as well as on reappearing autoreactive DSG-specific B cells (Day 0: 3873 vs Month 36: 2688, mean difference: 1185, 95%CI: -380 to 2750; p=0.20). Starting high doses of corticosteroids allowed a transitory decrease of serum BAFF levels that re-increased after doses tapering whereas it did not modify BAFF-R expression in autoreactive and non-autoreactive B cells. Our results suggest that the activation of autoreactive B cells at the onset of pemphigus is likely to be related to the presence of high BAFF serum levels and that the decreased BAFF-R expression after rituximab might be responsible for the delayed generation of memory B cells, resulting in a rather long period of mild pemphigus activity after rituximab therapy. Conversely, the incomplete B cell depletion and persistent BAFF-R expression associated with high BAFF serum levels might explain the high number of relapses in patients treated with CS alone., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Hébert, Maho-Vaillant, Golinski, Petit, Riou, Boyer, Musette, Calbo and Joly.)

Importance: Rituximab and short-term corticosteroid therapy are the criterion standard treatments for patients with newly diagnosed moderate to severe pemphigus., Objective: To examine factors associated with short-term relapse in patients with pemphigus treated with rituximab., Design, Setting, and Participants: This post hoc analysis of a randomized clinical trial (Comparison Between Rituximab Treatment and Oral Corticosteroid Treatment in Patients With Pemphigus [RITUX 3]) conducted from January 1, 2010, to December 31, 2015, included patients from 20 dermatology departments of tertiary care centers in France from the RITUX 3 trial and 3 newly diagnosed patients treated according to the trial protocol. Data analysis was performed from February 1 to June 30, 2019., Exposure: Patients randomly assigned to the rituximab group in the RITUX 3 trial and the 3 additional patients were treated with 1000 mg of intravenous rituximab on days 0 and 14 and 500 mg at months 12 and 18 combined with a short-term prednisone regimen., Main Outcomes and Measures: Baseline (pretreatment) clinical and biological characteristics (Pemphigus Disease Area Index [PDAI] score, ranging from 0-250 points, with higher values indicating more severe disease) and changes in anti-desmoglein (DSG) 1 and anti-DSG3 values as measured by enzyme-linked immunosorbent assay during the 3 months after rituximab treatment were compared between patients with disease relapse and those who maintained clinical remission during the first 12 months after treatment. The positive and negative predictive values of these factors were calculated., Results: Among 47 patients (mean [SD] age, 54.3 [17.0] years; 17 [36%] male and 30 [64%] female) included in the study, the mean (SD) baseline PDAI score for patients with relapsing disease was higher than that of the patients with nonrelapsing disease (54 [33] vs 28 [24]; P = .03). At month 3, 7 of 11 patients with relapsing disease (64%) vs 7 of 36 patients with nonrelapsing disease (19%) had persistent anti-DSG1 antibody values of 20 IU/mL or higher and/or anti-DSG3 antibody values of 130 IU/mL or higher (P = .01). A PDAI score of 45 or higher defining severe pemphigus and/or persistent anti-DSG1 antibody values of 20 IU/mL or higher and/or anti-DSG3 antibody values of 130 IU/mL or higher at month 3 provided a positive predictive value of 50% (95% CI, 27%-73%) and a negative predictive value of 94% (95% CI, 73%-100%) for the occurrence of relapse after rituximab., Conclusions and Relevance: The findings suggest that initial PDAI score and changes in anti-DSG antibody values after the initial cycle of rituximab might help differentiate a subgroup of patients with high risk of relapse who might benefit from maintenance rituximab infusion at month 6 from a subgroup of patients with low risk of relapse who do not need early maintenance therapy., Trial Registration: NCT00784589.

CD11c + B cells have been reported to be increased in autoimmune diseases, but they are detected in the blood of healthy individuals as well. We aimed to characterize CD11c + B cells from healthy donors by flow cytometry, microarray analysis, and in vitro functional assays. Here, we report that CD11c + B cells are a distinct subpopulation of B cells, enriched in the memory subpopulation even if their phenotype is heterogeneous, with overexpression of genes involved in B-cell activation and differentiation as well as in antigen presentation. Upon activation, CD11c + B cells can differentiate into antibody-secreting cells, and CD11c could be upregulated in CD11c - B cells by B-cell receptor activation. Finally, we show that patients with pemphigus, an autoimmune disease mediated by B cells, have a decreased frequency of CD11c + B cell after treatment, relative to baseline. Our findings show that CD11c + B cells are mainly memory B cells prone to differentiate into antibody secreting cells that accumulate with age, independently of gender., (Copyright © 2020 Golinski, Demeules, Derambure, Riou, Maho-Vaillant, Boyer, Joly and Calbo.)

Pemphigus Vulgaris is an autoimmune disease of the skin and mucous membranes, which is due to the production of pathogenic autoantibodies targeting desmoglein (DSG) 1 and 3, which are adhesion proteins of the keratinocytes. Rituximab is an anti-CD20 mAb which induces a prolonged depletion of blood B cells. We recently showed that rituximab was more effective than a standard oral corticosteroid (CS) treatment, allowing 90% of patients to achieve complete remission (CR). Additionally, we showed that DSG-specific-B (DSG positive) cells were still detectable during the B cell recovery which follows the initial rituximab-induced B cell depletion, even in patients in CR. In order to characterize DSG positive B cells in patients in CR after rituximab or CS treatment relative to those detectable at baseline in patients with an active pemphigus, we studied the expression profile of 31 genes of interest related to inflammatory cytokines, TNF receptors and activation markers. Using quantitative Polymerase Chain Reaction performed on one cell with a microfluidic technique, we found that patients' autoreactive B cells collected at baseline had a significantly higher expression of genes encoding for IL-1β, IL-23p19, and IL-12p35 pro-inflammatory cytokines and the IRF5 transcription factor, than non-autoreactive B cells. Surprisingly, the gene expression profile of DSG positive B cells collected after rituximab treatment in patients in CR was close to that of DSG positive B cells at baseline in patients with active pemphigus, except for the IL-1β and the CD27 memory marker genes, which were under-expressed after rituximab compared to baseline. Conversely, we observed a decreased expression of genes encoding for IL-1β and IL-23p19 in patients treated with CS relative to baseline. This study showed that: (i) DSG positive autoreactive B cells have a different gene expression profile than non-autoreactive B cells; (ii) rituximab and CS have different effects on the genes' expression in autoreactive DSG positive B cells from pemphigus patients.

Bullous Pemphigoid is the most common auto-immune bullous skin disease. It is characterized by the production of auto-antibodies directed against 2 proteins of the hemi-desmosome (BP180 and BP230). We assessed the efficacy and mechanisms of action of rituximab, an anti-CD20 monoclonal antibody, in 17 patients with severe and relapsing type of bullous pemphigoid. The phenotype, cytokine gene expression, and rearrangement of BP180-specific B-cell receptor genes were performed over 2 years following treatment. At the end of the study, 5 patients had died, 3 had withdrawn from the study, and 9 patients were in complete remission. The one- and two-year relapse rates were 44.1% (95% Confidence Interval (CI): 21.0-76.0%) and 66.5%, (95% CI: 38.4-91.4%), respectively. Phenotypic analyses confirmed dramatic B-cell depletion, which lasted for 9 to 12 months. The ELISA values of serum anti-BP180 antibodies and the frequency of BP180-specific circulating B cells decreased dramatically following treatment, which paralleled the improvement of skin lesions. During B-cell reconstitution, a polyclonal IgM repertoire appeared and a shift in the rearrangement of the B-cell receptor genes of BP180-specific circulating B cells was observed. Concurrently, we observed a decrease of IL-15, IL-6 and TNFα expressing BP180-specific B cells, and the emergence of IL-10 and IL-1RA-expressing BP180-specific IgM+ B cells in patients in complete remission off therapy, suggesting the functional plasticity of BP180-specific auto-immune B cells after rituximab treatment.

Background: High doses of corticosteroids are considered the standard treatment for pemphigus. Because long-term corticosteroid treatment can cause severe and even life-threatening side-effects in patients with this disease, we assessed whether first-line use of rituximab as adjuvant therapy could improve the proportion of patients achieving complete remission off-therapy, compared with corticosteroid treatment alone, while decreasing treatment side-effects of corticosteroids., Methods: We did a prospective, multicentre, parallel-group, open-label, randomised trial in 25 dermatology hospital departments in France (Ritux 3). Eligible participants were patients with newly diagnosed pemphigus aged 18-80 years being treated for the first time (not at the time of a relapse). We randomly assigned participants (1:1) to receive either oral prednisone alone, 1·0 or 1·5 mg/kg per day tapered over 12 or 18 months (prednisone alone group), or 1000 mg of intravenous rituximab on days 0 and 14, and 500 mg at months 12 and 18, combined with a short-term prednisone regimen, 0·5 or 1·0 mg/kg per day tapered over 3 or 6 months (rituximab plus short-term prednisone group). Follow-up was for 3 years (study visits were scheduled weekly during the first month of the study, then monthly until month 24, then an additional visit at month 36). Treatment was assigned through central computer-generated randomisation, with stratification according to disease-severity (severe or moderate, based on Harman's criteria). The primary endpoint was the proportion of patients who achieved complete remission off-therapy at month 24 (intention-to-treat analysis). This study is registered with ClinicalTrials.gov, number NCT00784589., Findings: Between May 10, 2010, and Dec 7, 2012, we enrolled 91 patients and randomly assigned 90 to treatment (90 were analysed; 1 patient withdrew consent before the random assignment). At month 24, 41 (89%) of 46 patients assigned to rituximab plus short-term prednisone were in complete remission off-therapy versus 15 (34%) of 44 assigned to prednisone alone (absolute difference 55 percentage points, 95% CI 38·4-71·7; p<0·0001. This difference corresponded to a relative risk of success of 2·61 (95% CI 1·71-3·99, p<0·0001), corresponding to 1·82 patients (95% CI 1·39-2·60) who would need to be treated with rituximab plus prednisone (rather than prednisone alone) for one additional success. No patient died during the study. More severe adverse events of grade 3-4 were reported in the prednisone-alone group (53 events in 29 patients; mean 1·20 [SD 1·25]) than in the rituximab plus prednisone group (27 events in 16 patients; mean 0·59 [1·15]; p=0·0021). The most common of these events in both groups were diabetes and endocrine disorder (11 [21%] with prednisone alone vs six [22%] with rituximab plus prednisone), myopathy (ten [19%] vs three [11%]), and bone disorders (five [9%] vs five [19%])., Interpretation: Data from our trial suggest that first-line use of rituximab plus short-term prednisone for patients with pemphigus is more effective than using prednisone alone, with fewer adverse events., Funding: French Ministry of Health, French Society of Dermatology, Roche., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Rheumatoid arthritis (RA) is the most common form of chronic inflammatory rheumatism. Identifying auto-antigens targeted by RA auto-antibodies is of major interest. Alpha-enolase (ENO1) is considered to be a pivotal auto-antigen in early RA but its pathophysiologic role remains unknown. The main objective of this study was to investigate the in vitro effects of soluble ENO1 on peripheral blood mononuclear cells (PBMC) from healthy donors and RA patients in order to determine the potential pathogenic role of ENO1. ELISA, transcriptomic analysis, experiments of receptor inhibition and flow cytometry analysis were performed to determine the effect, the target cell population and the receptor of ENO1. We showed that ENO1 has the ability to induce early production of pro-inflammatory cytokines and chemokines with delayed production of IL-10 and to activate the innate immune system. We demonstrated that ENO1 binds mainly to monocytes and activates the CD14-dependent TLR4 pathway both in healthy subjects and in RA patients. Our results establish for the first time that ENO1 is able to activate in vitro the CD14-dependent TLR4 pathway on monocytes involving a dual mechanism firstly pro-inflammatory and secondly anti-inflammatory. These results contribute to elucidating the role of this auto-antigen in the pathophysiologic mechanisms of RA.

Background: B and T cells play a key role in rheumatoid arthritis (RA) pathophysiology. RasGRP1 and RasGRP3 are involved in T and B cell receptors signaling, and belong to gene combination able to predict infliximab responsiveness, leading to the question of RasGRP1 and RasGRP3 involvement in RA., Methods: RasGRP1 and RasGRP3 expression levels were measured by qRT-PCR and/or western-blot in peripheral blood mononuclear cells (PBMCs), in T and B cells from untreated RA patients and in RA patients treated by TNFα inhibitors. T and B cells from healthy controls (HC) were cultured with TNFα, and TNFα receptors neutralizing antibodies to highlight the TNFα effects on RasGRP1 and RasGRP3 pathways. MAPK pathways and apoptosis were respectively analyzed using the Proteome Profiler arrays and flow cytometry., Results: In PBMCs from RA patients, gene expression levels of RasGRP1 were invariant while RasGRP3 was downregulated under TNFα inhibitors and upregulated under TNFα. In T cells from RA patients, RasGRP1 was decreased and its gene expression level was correlated with disease activity. In T cells from HC, TNFα stimulation increased RasGRP1 gene expression level while it reduced RasGRP1 protein expression level. Bryostatin-1 experiments have confirmed that the TNFα effect observed on T cells proliferation was due to the decrease of RasGRP1 expression. Besides, RasGRP3 expression level increased in PBMCs from RA patients under TNFα and in B cells from HC leading us to conclude that RasGRP3 in B cells was modulated by TNFα., Conclusion: This study demonstrates RasGRP1 dysregulation in RA patients while RasGRP3 is characterized as a biomarker linked to TNFα inhibitors. After binding to TNFR1, TNFα reduced RasGRP1 protein expression resulting in inhibition of T cell activation., Trial Registration: Clinicaltrials.gov NCT00234234 , registered 04 November 2008; NCT00767325 , registered 05 October 2005.

Objective: To evaluate the ability of the glycolytic enzyme alpha-enolase (ENO1) or its immunodominant peptide (pEP1) to reduce the severity of CIA in DBA/1 mice when injected in a prophylactic way., Methods: Mice were treated with mouse ENO1 or pEP1 one day prior to collagen II immunization. Clinical assessment was evaluated using 4 parameters (global and articular scores, ankle thickness and weight). Titers of serum anti-ENO1, anti-cyclic citrullinated peptides (anti-CCP) and anti-CII (total IgG and IgG1/IgG2a isotypes) antibodies were measured by ELISA at different time-points. Disease activity was assessed by histological analysis of both anterior and hind paws at the end of experimentation., Results: Prophylactic injection of 100 μg of ENO1 reduced severity of CIA. Serum levels of anti-CII antibodies were reduced in ENO1-treated mice. Concordantly, ENO1-treated mice joints presented less severe histological signs of arthritis. ENO1 did not induce a shift toward a Th2 response since IgG1/IgG2a ratio of anti-CII antibodies remained unchanged and IL-4 serum levels were similar to those measured in the control group., Conclusions: Pre-immunization with ENO1 or its immunodominant peptide pEP1 reduces CIA severity at the clinical, immunological and histological levels. Effects of pEP1 were less pronounced. This immunomodulatory effect is associated with a reduction in anti-CII antibodies production but is not due to a Th1/Th2 shift.