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BackgroundIn this study, we investigated the impact of the new haemodynamic definition of pulmonary arterial hypertension (PAH) as proposed by the 6th PH World Symposium on phenotypes and survival in patients with systemic sclerosis (SSc).MethodsIn SSc patients who were prospectively and consecutively screened for PAH including right heart catheterisation in Heidelberg or Zurich, haemodynamic and clinical variables have been reassessed according to the new PAH definition. Patients have been followed for 3.7±3.7 (median 3.4) years; Kaplan-Meier survival analysis was performed. Patients with significant lung or left heart disease were excluded from comparative analyses.ResultsThe final dataset included 284 SSc patients, 146 patients (49.2%) had mean pulmonary arterial pressure (mPAP) ≤20 mm Hg, 19.3% had mPAP 21–24 mm Hg and 29.4% had mPAP ≥25 mm Hg. In the group of mildly elevated mPAP, only four patients (1.4% of the whole SSc cohort) had pulmonary vascular resistance (PVR) values ≥3 Wood Units (WU) and could be reclassified as manifest SSc-APAH. Twenty-eight (9.8%) patients with mPAP of 21–24 mm Hg and PVR ≥2 WU already presented with early pulmonary vascular disease with decreased 6 min walking distance (6MWD) (pConclusionThe data of this study show that a PVR threshold ≥3 WU is too high to enable an early diagnosis of PAH. A PVR threshold ≥2 WU was already associated with pulmonary vascular disease, significantly reduced survival and would be more appropriate in SSc patients with mild PAH.

BackgroundPatients with V600BRAF mutant metastatic melanoma have higher rates of progression-free survival (PFS) and overall survival (OS) with first-line anti-PD1 (PD1]+anti-CTLA-4 (IPI) versus PD1. Whether this is also true after BRAF/MEKi therapy is unknown. We aimed to determine the efficacy and safety of PD1 versus IPI +PD1 after BRAF/MEK inhibitors (BRAF/MEKi).MethodsPatients with V600BRAF mutant metastatic melanoma treated with BRAF/MEKi who had subsequent PD1 versus IPI+PD1 at eight centers were included. The endpoints were objective response rate (ORR), PFS, OS and safety in each group.ResultsOf 200 patients with V600E (75%) or non-V600E (25%) mutant metastatic melanoma treated with BRAF/MEKi (median time of treatment 7.6 months; treatment cessation due to progressive disease in 77%), 115 (57.5%) had subsequent PD1 and 85 (42.5%) had IPI+PD1. Differences in patient characteristics between PD1 and IPI+PD1 groups included, age (med. 63 vs 54 years), time between BRAF/MEKi and PD1±IPI (16 vs 4 days), Eastern Cooperative Oncology Group Performance Status (ECOG PS) of ≥1 (62% vs 44%), AJCC M1C/M1D stage (72% vs 94%) and progressing brain metastases at the start of PD1±IPI (34% vs 57%). Median follow-up from PD1±IPI start was 37.8 months (95% CI, 33.9 to 52.9). ORR was 36%; 34% with PD1 vs 39% with IPI+PD1 (p=0.5713). Median PFS was 3.4 months; 3.4 with PD1 vs 3.6 months with IPI+PD1 (p=0.6951). Median OS was 15.4 months; 14.4 for PD1 vs 20.5 months with IPI+PD1 (p=0.2603). The rate of grade 3 or 4 toxicities was higher with IPI+PD1 (31%) vs PD1 (7%). ORR, PFS and OS were numerically higher with IPI+PD1 vs PD1 across most subgroups except for females, those with 3 years OS (area under the curve, AUC=0.74), while ECOG PS ≥1, progressing brain metastases and presence of bone metastases predicted primary progression (AUC=0.67).ConclusionsIPI+PD1 and PD1 after BRAF/MEKi have similar outcomes despite worse baseline prognostic features in the IPI+PD1 group, however, IPI+PD1 is more toxic. A combination of clinical factors can identify long-term survivors, but less accurately those with primary resistance to immunotherapy after targeted therapy.

BackgroundTalimogene laherparepvec (T-VEC) is a licensed therapy for use in melanoma patients of stage IIIB-IVM1a with injectable, unresectable metastatic lesions in Europe. Approval was based on the Oncovex Pivotal Trial in Melanoma study, which also included patients with distant metastases and demonstrated an overall response rate (ORR) of 40.5% and a complete response (CR) rate of 16.6%.ObjectivesThe aim of this study was to assess the outcome of melanoma patients treated with T-VEC in a real-life clinical setting.MethodsBased on data from 10 melanoma centers in Austria, Switzerland and southern Germany, we conducted a retrospective chart review, which included 88 patients (44 male, 44 female) with a median age of 72 years (range 36–95 years) treated with T-VEC during the period from May 2016 to January 2020.Results88 patients fulfilled the inclusion criteria for analysis. The ORR was 63.7%. 38 patients (43.2%) showed a CR, 18 (20.5%) had a partial response, 8 (9.1%) had stable disease and 24 (27.3%) patients had a progressive disease. The median treatment period was 19 weeks (range: 1–65), an average of 11 doses (range: 1–36) were applied. 39 (45.3%) patients developed adverse events, mostly mild, grade I (64.1%).ConclusionThis real-life cohort treatment with T-VEC showed a high ORR and a large number of durable CRs.

BackgroundIntegrin αvβ6 is a heterodimeric cell surface protein whose cellular expression is determined by the availability of the integrin β6 subunit (ITGB6). It is expressed at very low levels in most organs during tissue homeostasis but shows highly upregulated expression during the process of tumorigenesis in many cancers of epithelial origin. Notably, enhanced expression of integrin αvβ6 is associated with aggressive disease and poor prognosis in numerous carcinoma entities. Integrin αvβ6 is one of the major physiological activators of transforming growth factor-β (TGF-β), which has been shown to inhibit the antitumor T-cell response and cause resistance to immunotherapy in mouse models of colorectal and mammary cancer. In this study, we investigated the effect of ITGB6 expression and antibody-mediated integrin αvβ6 inhibition on the tumor immune response in colorectal cancer.MethodsUsing orthotopic and heterotopic tumor cell injection, we assessed the effect of ITGB6 on tumor growth and tumor immune response in wild type mice, mice with defective TGF-β signaling, and mice treated with anti-integrin αvβ6 antibodies. To examine the effect of ITGB6 in human colorectal cancer, we analyzed RNAseq data from the colon adenocarcinoma dataset of The Cancer Genome Atlas (TCGA-COAD).ResultsWe demonstrate that expression of ITGB6 is an immune evasion strategy in colorectal cancer, causing inhibition of the antitumor immune response and resistance to immune checkpoint blockade therapy by activating latent TGF-β. Antibody-mediated inhibition of integrin αvβ6 sparked a potent cytotoxic T-cell response and overcame resistance to programmed cell death protein 1 (PD-1) blockade therapy in ITGB6 expressing tumors, provoking a drastic increase in anti-PD-1 treatment efficacy. Further, we show that the majority of tumors in patients with colorectal cancer express sufficient ITGB6 to provoke inhibition of the cytotoxic T-cell response, indicating that most patients could benefit from integrin αvβ6 blockade therapy.ConclusionsThese findings propose inhibition of integrin αvβ6 as a promising new therapy for colorectal cancer, which blocks tumor-promoting TGF-β activation, prevents tumor exclusion of cytotoxic T-cells and enhances the efficacy of immune checkpoint blockade therapy.

ObjectivesTo integrate published single-cell RNA sequencing (scRNA-seq) data and assess the contribution of synovial fibroblast (SF) subsets to synovial pathotypes and respective clinical characteristics in treatment-naïve early arthritis.MethodsIn this in silico study, we integrated scRNA-seq data from published studies with additional unpublished in-house data. Standard Seurat, Harmony and Liger workflow was performed for integration and differential gene expression analysis. We estimated single cell type proportions in bulk RNA-seq data (deconvolution) from synovial tissue from 87 treatment-naïve early arthritis patients in the Pathobiology of Early Arthritis Cohort using MuSiC. SF proportions across synovial pathotypes (fibroid, lymphoid and myeloid) and relationship of disease activity measurements across different synovial pathotypes were assessed.ResultsWe identified four SF clusters with respective marker genes: PRG4+ SF (CD55, MMP3, PRG4, THY1neg); CXCL12+ SF (CXCL12, CCL2, ADAMTS1, THY1low); POSTN+ SF (POSTN, collagen genes, THY1); CXCL14+ SF (CXCL14, C3, CD34, ASPN, THY1) that correspond to lining (PRG4+ SF) and sublining (CXCL12+ SF, POSTN+ + and CXCL14+ SF) SF subsets. CXCL12+ SF and POSTN+ + were most prominent in the fibroid while PRG4+ SF appeared highest in the myeloid pathotype. Corresponding, lining assessed by histology (assessed by Krenn-Score) was thicker in the myeloid, but also in the lymphoid pathotype + the fibroid pathotype. PRG4+ SF correlated positively with disease severity parameters in the fibroid, POSTN+ SF in the lymphoid pathotype whereas CXCL14+ SF showed negative association with disease severity in all pathotypes.ConclusionThis study shows a so far unexplored association between distinct synovial pathologies and SF subtypes defined by scRNA-seq. The knowledge of the diverse interplay of SF with immune cells will advance opportunities for tailored targeted treatments.

ObjectivePulmonary arterial hypertension (PAH) is a severe complication of systemic sclerosis (SSc). In this longitudinal study, we aimed to identify factors associated with an unfavourable outcome in patients with SSc with early PAH (SSc-PAH) from the DETECT cohort.MethodsPatients with SSc-PAH enrolled in DETECT were observed for up to 3 years. Associations between cross-sectional variables and disease progression (defined as the occurrence of any of the following events: WHO Functional Class worsening, combination therapy for PAH, hospitalisation or death) were analysed by univariable logistic regression.ResultsOf 57 patients with PAH (median observation time 12.6 months), 25 (43.9%) had disease progression. The following factors (OR (95% CI)) were associated with disease progression: male gender (4.1 (1.2 to 14.1)), high forced vital capacity % predicted/carbon monoxide lung diffusion capacity (DLCO)% predicted ratio (3.6 (1.2 to 10.7)), high Borg Dyspnoea Index (1.7 (1.1 to 2.6)) and low DLCO% predicted (non-linear relationship).ConclusionMore than 40% of early-diagnosed patients with SSc-PAH had disease progression during a short follow-up time, with male gender, functional capacity and pulmonary function tests at PAH diagnosis being associated with progression. This suggests that even mild PAH should be considered a high-risk complication of SSc.

ObjectivesJanus kinase 2 (JAK2) has recently been described as a novel downstream mediator of the pro-fibrotic effects of transforming growth factor-β. Although JAK2 inhibitors are in clinical use for myelodysplastic syndromes, patients often rapidly develop resistance. Tumour cells can escape the therapeutic effects of selective JAK2 inhibitors by mutation-independent transactivation of JAK2 by JAK1. Here, we used selective JAK2 inhibition as a model to test the hypothesis that chronic treatment may provoke resistance by facilitating non-physiological signalling pathways in fibroblasts.MethodsThe antifibrotic effects of long-term treatment with selective JAK2 inhibitors and reactivation of JAK2 signalling by JAK1-dependent transphosphorylation was analysed in cultured fibroblasts and experimental dermal and pulmonary fibrosis. Combined JAK1/JAK2 inhibition and co-treatment with an HSP90 inhibitor were evaluated as strategies to overcome resistance.ResultsThe antifibrotic effects of selective JAK2 inhibitors on fibroblasts decreased with prolonged treatment as JAK2 signalling was reactivated by JAK1-dependent transphosphorylation of JAK2. This reactivation could be prevented by HSP90 inhibition, which destabilised JAK2 protein, or with combined JAK1/JAK2 inhibitors. Treatment with combined JAK1/JAK2 inhibitors or with JAK2 inhibitors in combination with HSP90 inhibitors was more effective than monotherapy with JAK2 inhibitors in bleomycin-induced pulmonary fibrosis and in adTBR-induced dermal fibrosis.ConclusionFibroblasts can develop resistance to chronic treatment with JAK2 inhibitors by induction of non-physiological JAK1-dependent transactivation of JAK2 and that inhibition of this compensatory signalling pathway, for example, by co-inhibition of JAK1 or HSP90 is important to maintain the antifibrotic effects of JAK2 inhibition with long-term treatment.

Objectives Smoking has been connected to citrullination of antigens and formation of anti-citrullinated peptide antibodies (ACPAs) in rheumatoid arthritis (RA). Since smoking can modify proteins by carbamylation (formation of homocitrulline), this study was conducted to investigate these effects on vimentin in animal models and RA. Methods The efficiency of enzymatic carbamylation of vimentin was characterised. B-cell response was investigated after immunisation of rabbits with different vimentin isoforms. Effects of tobacco smoke exposure on carbamylation of vimentin and formation of autoantibodies were analysed in mice. The antibody responses against isoforms of vimentin were characterised with respect to disease duration and smoking status of patients with RA. Results Enzymatic carbamylation of vimentin was efficiently achieved. Subsequent citrullination of vimentin was not disturbed by homocitrullination. Sera from rabbits immunised with carbamylated vimentin (carbVim), in addition to carbVim also recognised human IgG-Fc showing rheumatoid factor-like reactivity. Smoke-exposed mice contained detectable amounts of carbVim and developed a broad immune response against carbamylated antigens. Although the prevalence of anti-carbamylated antibodies in smokers and non-smokers was similar, the titres of carbamylated antibodies were significantly increased in sera of smoking compared with non-smoking RA. CarbVim antibodies were observed independently of ACPAs in early phases of disease and double-positive patients for anti-mutated citrullinated vimentin (MCV) and anti-carbVim antibodies showed an extended epitope recognition pattern towards MCV. Conclusions Carbamylation of vimentin is inducible by cigarette smoke exposure. The polyclonal immune response against modified antigens in patients with RA is not exclusively citrulline-specific and carbamylation of antigens could be involved in the pathogenesis of disease. Trial registration number ISRCTN36745608; EudraCT Number: 2006-003146-41.

Background: In 2010 the latest ASAS-EULAR recommendations for ankylosing spondylitis and the ASAS recommendations for the use of TNF-inhibitors (TNFi) have been published. Since then new treatments for axial Spondyloarthritis (axSpA) have become available. Aim: To update and integrate the two sets[for full text, please go to the a.m. URL], 44. Kongress der Deutschen Gesellschaft für Rheumatologie (DGRh); 30. Jahrestagung der Deutschen Gesellschaft für Orthopädische Rheumatologie (DGORh); 26. Jahrestagung der Gesellschaft für Kinder- und Jugendrheumatologie (GKJR)

During the current global outbreak of coronavirus disease 2019 (COVID-19), risk stratification of patients is of utmost importance. Currently, patients >65 years and those with pre-existing medical conditions such as cardiovascular disease, chronic respiratory disease or diabetes mellitus are considered at higher risk for severe disease.1 The Swiss Federal Office of Public Health, like similar authorities around the world, has additionally included patients on immunosuppressants in the high-risk group for developing severe COVID-19.2 However, at this moment we do not have enough evidence either to support or to reject this assumption. We report the case of a 57-year-old woman with systemic sclerosis (SSc) who developed COVID-19. Comorbidities were insulin-dependent type 2 diabetes mellitus and WHO grade I obesity. The anti-topoisomerase I antibody-positive patient was diagnosed with SSc in 2017. SSc-associated interstitial lung disease (SSc-ILD), with cough and exertion dyspnoea, was the leading organ manifestation, associated with symmetrical, non-erosive polyarthritis, and elevated acute phase reactants. Treatment with the anti-interleukin (IL) 6 receptor blocker tocilizumab, with 8 mg/kg body weight every 4 weeks intravenously, was started, leading to a good control of both …

BackgroundAnti-programmed death-ligand 1 (αPD-L1) immunotherapy is approved to treat bladder cancer (BC) but is effective in + antitumor T cells known to improve αPD-L1 efficacy. We hypothesized that the tumor microenvironment, including local immune cells in primary versus metastatic BC, differentially affects immunotherapy responses and that IL-2c effects could differ from, and thus complement αPD-L1.MethodsWe studied mechanisms of IL-2c and αPD-L1 efficacy using PD-L1+ mouse BC cell lines MB49 and MBT-2 in orthotopic (bladder) and metastatic (lung) sites.ResultsIL-2c reduced orthotopic tumor burden and extended survival in MB49 and MBT-2 BC models, similar to αPD-L1. Using antibody-mediated cell depletions and genetically T cell-deficient mice, we unexpectedly found that CD8+ T cells were not necessary for IL-2c efficacy against tumors in bladder, whereas γδ T cells, not reported to contribute to αPD-L1 efficacy, were indispensable for IL-2c efficacy there. αPD-L1 responsiveness in bladder required conventional T cells as expected, but not γδ T cells, altogether defining distinct mechanisms for IL-2c and αPD-L1 efficacy. γδ T cells did not improve IL-2c treatment of subcutaneously challenged BC or orthotopic (peritoneal) ovarian cancer, consistent with tissue-specific and/or tumor-specific γδ T cell contributions to IL-2c efficacy. IL-2c significantly altered bladder intratumoral γδ T cell content, activation status, and specific γδ T cell subsets with antitumor or protumor effector functions. Neither IL-2c nor αPD-L1 alone treated lung metastatic MB49 or MBT-2 BC, but their combination improved survival in both models. Combination treatment efficacy in lungs required CD8+ T cells but not γδ T cells.ConclusionsMechanistic insights into differential IL-2c and αPD-L1 treatment and tissue-dependent effects could help develop rational combination treatment strategies to improve treatment efficacy in distinct cancers. These studies also provide insights into γδ T cell contributions to immunotherapy in bladder and engagement of adaptive immunity by IL-2c plus αPD-L1 to treat refractory lung metastases.

Social media has profoundly changed the way we communicate and interact. Social media interaction brings new demands and challenges to our private and professional lives and has led to the development of completely new professions such as social media account managers, social media analysts, social media coordinators just to name a few.1 2 The main attraction of social media is that everyone can present themselves and their preferences, dislikes, opinions, etc., to a large audience, not least with the aim of influencing society, as large campaigns by political parties or companies impressively demonstrate. Successful social media influencers have several hundred thousand followers and can charge up to 200 Euro per Twitter post.3 What does this development mean for medicine and research? How do we wisely use social media to disseminate scientific data and interact with our key stakeholders? And does every good scientist nowadays have to be active on social media? The answer to the last question is no. Although social media presence is a great way to disseminate your research, interact professionally and stay up to date, it is certainly not a requirement to be an active member of the scientific community. People who do not feel comfortable with social media or do not want to be exposed for privacy reasons have many other ways to achieve the same results without social media. Nevertheless, it is important to understand the mechanisms and processes of social media, as it has become an important part of professional life and publishing. Most journals, including RMD Open and Annals of the Rheumatic Diseases (ARD) publish Altmetric scores alongside articles. Thus, as publishing researcher you are measured by your social media presence, regardless of whether you participate actively or not. In this editorial, we explain how the Altmetric score is composed and …

ObjectiveSystemic sclerosis (SSc) is a systemic autoimmune disease with high morbidity and significant mortality. There is a great need of predictors that would allow risk stratification of patients with SSc and ultimately initiation of treatment early enough to ensure optimal clinical results. In this study, we evaluated whether a history of digital ulcers (HDU) at presentation may be a predictor of vascular outcomes and of overall clinical worsening and death in patients with SSc.MethodsPatients from the EULAR Scleroderma Trials and Research (EUSTAR) database, satisfying at inclusion the 1980 American College of Rheumatology classification criteria for SSc, who had a follow-up of at least 3 years since baseline or who have died, were included in the analysis. HDU at presentation as a predictor of disease worsening or death was evaluated by Cox proportional hazards regression analysis.Results3196 patients matched the inclusion criteria (male sex 13.2%, 33.4% diffuse subset). At presentation, 1092/3196 patients had an HDU (34.1%). In multivariable analysis adjusting for age, gender and all parameters considered potentially significant, HDU was predictive for the presence of active digital ulcers (DUs) at prospective visits (HR (95% CI)): 2.41 (1.91 to 3.03), pConclusionsIn patients with SSc, HDU at presentation predicts the occurrence of DUs at follow-up and is associated with cardiovascular worsening and decreased survival.

BackgroundActivating transcription factor 3 (ATF3), a member of the ATF/cAMP-responsive element binding (CREB) family of transcription factors, regulates cellular response to stress including oxidative stress. The aim of this study was to analyse the role of ATF3 in fibroblast activation in systemic sclerosis (SSc).MethodsATF3 was analysed by reverse transcription quantitative PCR, western blot and immunohistochemistry. ATF3 knockout fibroblasts and mice were used to study the functional role of ATF3. Knockdown experiments, reporter assays and coimmunoprecipitation were performed to study the effects of ATF3 on Smad and activation protein 1 (AP-1) signalling. The role of c-Jun was analysed by costaining, specific inactivation and coimmunoprecipitation.ResultsTransforming growth factor-β (TGFβ) upregulates the expression of ATF3 in SSc fibroblasts. ATF3-deficient fibroblasts were less sensitive to TGFβ, whereas ectopic expression of ATF3 enhanced the profibrotic effects of TGFβ. Mechanistically, ATF3 interacts with Smad3 directly on stimulation with TGFβ and regulates Smad activity in a c-Jun-dependent manner. Knockout of ATF3 protected mice from bleomycin-induced fibrosis and fibrosis induced by overexpression of a constitutively active TGFβ receptor I. Reporter assays and analyses of the expression of Smad target genes demonstrated that binding of ATF3 regulates the transcriptional activity of Smad3.ConclusionsWe demonstrate for the first time a key role for ATF3 in fibrosis. Knockout of the ATF3 gene reduced the stimulatory effect of TGFβ on fibroblasts by interfering with canonical Smad signalling and protected the mice from experimental fibrosis in two different models. ATF3 might thus be a candidate for molecular targeted therapies for SSc.

Objective To investigate the effects of BET bromodomain protein inhibition on inflammatory activation and functional properties of rheumatoid arthritis synovial fibroblasts (RASF). Methods The expression of the BET bromodomain proteins BRD2, BRD3 and BRD4 was analysed in synovial tissue by immunohistochemistry. RASE were stimulated with tumour necrosis factor (TNF)-alpha, interleukin (IL)-1 beta and toll-like receptor (TLR) ligands (Pam3, pIC and lipopolysaccharide (LPS)) in the presence or absence of the BET inhibitor I-BET151, or siRNA targeting BRD2, BRD3 and BRD4. RASF expression of inflammatory mediators, including MMP1, MMP3, IL-6 and IL-8, was measured by q-PCR, q-PCR array and ELISA. Cellular viability, apoptosis, proliferation and chemoattractive properties of RASF were investigated using MTT, cell apoptosis ELISA, BrdU-based proliferation and transwell migration assays. Results BRD2, BRD3 and BRD4 proteins were detected in rheumatoid arthritis (RA) synovial tissue, expressed in both RASF and macrophages. I-BET151 suppressed cytokine and TLR ligand-induced secretion of MMP1, MMP3, IL-6 and IL-8, and mRNA expression of more than 70% of genes induced by TNF-alpha and IL-1 beta. Combined silencing of BRD2, BRD3 and BRD4 significantly reduced cytokine and TLR ligand-induced expression of a subset of gene products targeted by I-BET151, including MMP1, CXCL10 and CXCL11. I-BET151 treatment of RASF reduced RASF proliferation, and the chemotactic potential for peripheral blood leucocytes of RASF conditioned medium. Conclusions Inhibition of BET family proteins suppresses the inflammatory, matrix-degrading, proliferative and chemoattractive properties of RASF and suggests a therapeutic potential in the targeting of epigenetic reader proteins in RA

Objectives S100A4 is a calcium binding protein with regulatory functions in cell homeostasis, proliferation and differentiation that has been shown to promote cancer progression and metastasis. In the present study, we evaluated the role of S100A4 in fibroblast activation in systemic sclerosis (SSc). Methods The expression of S100A4 was analysed in human samples, murine models of SSc and in cultured fibroblasts by real-time PCR, immunohistochemistry and western blot. The functional role of S100A4 was evaluated using siRNA, overexpression, recombinant protein and S100A4 knockout (S100A4 −/− ) mice. Transforming growth factor β (TGF-β) signalling was assessed by reporter assays, staining for phosphorylated Smad2/3 and analyses of target genes. Results The expression of S100A4 was increased in SSc skin and in experimental fibrosis in a TGF-β/Smad-dependent manner. Overexpression of S100A4 or stimulation with recombinant S100A4 induced an activated phenotype in resting normal fibroblasts. In contrast, knockdown of S100A4 reduced the pro-fibrotic effects of TGF-β and decreased the release of collagen. S100A4 −/− mice were protected from bleomycin-induced skin fibrosis with reduced dermal thickening, decreased hydroxyproline content and lower myofibroblast counts. Deficiency of S100A4 also ameliorated fibrosis in the tight-skin-1 (Tsk-1) mouse model. Conclusions We characterised S100A4 as a downstream mediator of the stimulatory effects of TGF-β on fibroblasts in SSc. TGF-β induces the expression of S100A4 to stimulate the release of collagen in SSc fibroblasts and induce fibrosis. Since S100A4 is essentially required for the pro-fibrotic effects of TGF-β and neutralising antibodies against S100A4 are currently evaluated, S100A4 might be a candidate for novel antifibrotic therapies.

Objectives: Casein kinase II (CK2) is a constitutively active serine/threonine protein kinase that plays a key role in cellular transformation and tumorigenesis. The purpose of the study was to characterise whether CK2 contributes to the pathologic activation of fibroblasts in patients with SSc and to evaluate the antifibrotic potential of CK2 inhibition. Methods: Activation of CK2, JAK2 and STAT3 in human skin and in experimental fibrosis was analysed by immunohistochemistry. CK2 signalling was inhibited by the selective CK2 inhibitor 4, 5, 6, 7-Tetrabromobenzotriazole (TBB). The mouse models of bleomycin-induced and TGFβ receptor I (TBR)-induced dermal fibrosis were used to evaluate the antifibrotic potential of specific CK2 inhibition in vivo. Result: Increased expression of CK2 was detected in skin fibroblasts of SSc patients. Inhibition of CK2 by TBB abrogated the TGFβ-induced activation of JAK2/STAT3 signalling and prevented the stimulatory effects of TGFβ on collagen release and myofibroblasts differentiation in cultured fibroblasts. Inhibition of CK2 prevented bleomycin-induced and TBR-induced skin fibrosis with decreased dermal thickening, lower myofibroblast counts and reduced accumulation of collagen. Treatment with TBB also induced regression of pre-established fibrosis. The antifibrotic effects of TBB were accompanied by reduced activation of JAK2/STAT3 signalling in vivo. Conclusions: We provide evidence that CK2 is activated in SSc and contributes to fibroblast activation by regulating JAK2/STAT3 signalling. Inhibition of CK2 reduced the pro-fibrotic effects of TGFβ and inhibited experimental fibrosis. Targeting of CK2 may thus be a novel therapeutic approach for SSc and other fibrotic diseases.

BackgroundSystemic sclerosis (SSc) is a severe, progressive multiorgan disease but to date, there are no established standardised international guidelines for follow-up of patients with SSc. The goal of this project was to develop an expert consensus for annual systematic investigations in patients with SSc to enhance their standard-of-care.Material and methodsThe Delphi method was applied. All SSc experts from the European Scleroderma Trials and Research group network and the Scleroderma Clinical Trial Consortium were invited to participate. All experts were asked to answer questionnaires in five Delphi steps to determine the domains of interest and tools for each domain for an annual systematic assessment of patients with SSc. Each item was rated on a scale between 0% and 100% (not and very important), and parameters rated >80% by more than 75% of the experts were regarded as acceptable.ResultsIn total, 157 experts worldwide participated with 71.3% experts seeing >50 patients with SSc annually. In the first round, 23 domains and 204 tools were suggested. After five Delphi steps, experts agreed on 10 domains including (1) Raynaud’s phenomenon; (2) Digital ulcers; (3) Skin and mucosa; (4) Lung; (5); Heart; (6) GI domain, (7) Renal; (8) Musculoskeletal; (9) Laboratory and (10) Treatment. Overall, 55 tools were identified including clinical assessments, laboratory measurements and imaging or functional investigations.ConclusionThrough five Delphi steps with world leading experts, a consensus was established on strongly suggested tools for a minimum annual systemic assessment of organ involvement in SSc. This work should enhance the standardisation and homogenisation of the practices.

ObjectivesTo assess whether the discrepancy between the strong antifibrotic effects of tyrosine kinase inhibitors (TKIs) in animal models and the inconsistent results in clinical studies might be related to the activation levels of drug targets.MethodsSkin sections of bleomycin, TSK1, Fra-2 transgenic mice, SSc patients and controls were analysed by histology and immunohistochemistry. Subgroups of mice were treated with the TKIs nilotinib or imatinib. Differences in the activation levels of the TKI targets p-PDGFRβ (platelet derived growth factor β) and p-c-abl were assessed.ResultsIn bleomycin and TSK1 mice, expression of activated p-PDGFRβ (platelet derived growth factor receptor β) and p-c-abl was ubiquitous with strong upregulation compared with controls. Treatment with TKIs resulted in successful target inhibition and consequently reduced dermal fibrosis. In the Fra-2 model, the activation levels of p-PDGFRβ and p-c-abl were much lower than in the bleomycin and the TSK1 models. Accordingly, nilotinib did not prevent dermal fibrosis and target inhibition was unsuccessful. Notably, in skin biopsies of SSc patients, the mean activation levels of TKI targets were only moderate and in the majority of patients resembled those of the non-responsive Fra-2 model.ConclusionsAnimal models for proof-of-concept studies should be selected based on a similar activation level and expression pattern of drug targets as in human SSc.

Objective Based on previous data that have linked the small ubiquitin-like modifier-1 (SUMO-1) to the pathogenesis of rheumatoid arthritis (RA), we have investigated the expression of the highly homologous SUMO family members SUMO-2/3 in human RA and in the human tumour necrosis factor α transgenic (hTNFtg) mouse model of RA and studied their role in regulating disease specific matrixmetalloproteinases (MMPs). Methods Synovial tissue was obtained from RA and osteoarthritis (OA) patients and used for histological analyses as well as for the isolation of synovial fibroblasts (SFs). The expression of SUMO-2/3 in RA and OA patients as well as in hTNFtg and wild type mice was studied by PCR, western blot and immunostaining. SUMO-2/3 was knocked down using small interfering RNA in SFs, and TNF-α induced MMP production was determined by ELISA. Activation of nuclear factor-κB (NF-κB) was determined by a luciferase activity assay and a transcription factor assay in the presence of the NF-κB inhibitor BAY 11-7082. Results Expression of SUMO-2 and to a lesser extent of SUMO-3 was higher in RA tissues and RASFs compared with OA controls. Similarly, there was increased expression of SUMO-2 in the synovium and in SFs of hTNFtg mice compared with wild type animals. In vitro, the expression of SUMO-2 but not of SUMO-3 was induced by TNF-α. The knockdown of SUMO-2/3 significantly increased the TNF-α and interleukin (IL)-1β induced expression of MMP-3 and MMP-13, accompanied by increased NF-κB activity. Induction of MMP-3 and MMP-13 was inhibited by blockade of the NF-κB pathway. TNF-α and IL-1β mediated MMP-1 expression was not regulated by SUMO-2/3. Conclusions Collectively, we show that despite their high homology, SUMO-2/3 are differentially regulated by TNF-α and selectively control TNF-α mediated MMP expression via the NF-κB pathway. Therefore, we hypothesise that SUMO-2 contributes to the specific activation of RASF.

Objective To establish an expert consensus on which criteria are the most appropriate in clinical practice to refer patients with systemic sclerosis (SSc) for right heart catheterisation (RHC) when pulmonary hypertension (PH) is suspected. Methods A three stage internet based Delphi consensus exercise involving worldwide PH experts was designed. In the first stage, a comprehensive list of domains and items combining evidence based indications and expert opinions were obtained. In the second and third stages, experts were asked to rate each item selected in the list. After each of stages 2 and 3, the number of items and criteria were reduced according to a cluster analysis. Results A literature search and the opinions of 47 experts participating in Delphi stage 1 provided a list of seven domains containing 142 criteria. After stages 2 and 3, these domains and tools were reduced to three domains containing eight tools: clinical (progressive dyspnoea over the past 3 months, unexplained dyspnoea, worsening of WHO dyspnoea functional class, any finding on physical examination suggestive of elevated right heart pressures and any sign of right heart failure), echocardiography (systolic pulmonary artery pressure >45 mm Hg and right ventricle dilation) and pulmonary function tests (diffusion lung capacity for carbon monoxide Conclusions Among experts in pulmonary arterial hypertension–SSc, a core set of criteria for clinical practice to refer SSc patients for RHC has been defined by Delphi consensus methods. Although these indications are recommended by this expert group to be used as an interim tool, it will be necessary to formally validate the present tools in further studies.

Background Immune checkpoint inhibitors targeting cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and programmed cell death protein 1 (PD-1) have been recently approved for treatment of patients with metastatic melanoma and non-small cell lung cancer (NSCLC). Despite important clinical benefits, these therapies are associated with a diverse spectrum of immune-related adverse events (irAEs) that are typically transient, but occasionally severe or even fatal. Case presentation This autopsy case illustrates that clinically overt irAEs may represent only a fraction of the total spectrum of immune-related organ pathology in patients treated with immune checkpoint inhibitors. We report a comprehensive analysis of systemic irAE pathology based on the autopsy of a 35-year-old female patient with metastatic melanoma treated first with ipilimumab and then nivolumab. The clinical course was characterized by a mixed tumor response with regression of skin and lung metastases and fatal progression of metastatic disease in the small bowel, peritoneum and brain. During therapy with ipilimumab, radiographic features of immune-related pneumonitis were noted. The autopsy examination established a sarcoid-like granulomatous reaction of the lung, pulmonary fibrosis and diffuse alveolar damage. Importantly, a clinically unapparent but histologically striking systemic inflammation involving the heart, central nervous system, liver and bone marrow was identified. Severe immune-related end-organ damage due to lymphocytic myocarditis was found. Conclusions Autopsy studies are an important measure of quality control and may identify clinically unapparent irAEs in patients treated with immunotherapy. Pathologists and clinicians need to be aware of the broad spectrum of irAEs for timely management of treatment-related morbidity. Electronic supplementary material The online version of this article (doi:10.1186/s40425-016-0117-1) contains supplementary material, which is available to authorized users.

ObjectivesFibrosis is a predominant cause of death in systemic sclerosis (SSc). First epigenetic modifications have recently been shown to contribute to activation of SSc fibroblasts. Here, we investigated inhibition of sumoylation as a novel antifibrotic approach.MethodsSumoylation was inhibited by siRNA-mediated knockdown of the Small Ubiquitin-like MOdifiers (SUMO) E2-conjugating enzyme Ubc9, which is essential for sumoylation. The effects of knockdown of Ubc9 were analysed in bleomycin-induced dermal fibrosis, and in the model of fibrosis induced by overexpression of a constitutively active TGF-beta receptor type I (TBR). SUMO-1 and phosphorylated Smad3 were detected by immunohistochemistry.ResultsIncreased staining for SUMO-1 was detected in patients with SSc and in experimental fibrosis. Inhibition of sumoylation exerted potent antifibrotic effects and prevented dermal thickening, myofibroblast differentiation and accumulation of collagen induced by bleomycin, or by overexpression of constitutively active TBR. Moreover, knockdown of Ubc9 reduced the accumulation of phosphorylated Smad3 in experimental fibrosis indicating that inhibition of sumoylation may normalise canonical TGF-β signalling in vivo.ConclusionsWe demonstrate that inhibition of sumoylation reduces canonical TGF-β signalling and prevents experimental fibrosis in different preclinical models. These data provide first evidence that targeting of aberrant sumoylation may be a novel therapeutic approach for fibrotic diseases.

ObjectivesTissue fibrosis is a leading cause of death in patients with systemic sclerosis (SSc). Effective antifibrotic treatments are not available. Here, the authors investigated inhibition of hedgehog signalling by targeting Smoothened (Smo) as a novel antifibrotic approach.MethodsThe activation status of the hedgehog pathway was assessed by immunohistochemistry for Gli transcription factors and by quantification of hedgehog target genes. Hedgehog signalling was inhibited by the selective inhibitor LDE223 and by small interfering RNA against Smo in the models of bleomycin-induced dermal fibrosis and in tight-skin-1 mice.ResultsHedgehog signalling is activated in SSc and in murine models of SSc. Inhibition of Smo either by LDE223 or by small interfering RNA prevented dermal thickening, myofibroblast differentiation and accumulation of collagen upon challenge with bleomycin. Targeting Smo also exerted potent antifibrotic effects in tight-skin-1 mice and did prevent progression of fibrosis and induced regression of pre-established fibrosis.ConclusionsInhibition of hedgehog signalling exerted potent antifibrotic effects in preclinical models of SSc in both preventive and therapeutic settings. These findings might have direct translational implications because inhibitors of Smo are already available and yielded promising results in initial clinical trials.

The objective of this study is to assess diagnostic accuracy for the detection of interstitial lung disease (ILD) in image series with high increment and reduced number of slices in patients with systemic sclerosis (SSc).45 patients with SSc underwent high-resolution CT (HRCT). Three series of secondary captures were reconstructed as follows: series 1, series with 10 mm increment and 1 mm slices; series 2, seven axial images with baso-apical gradient; series 3, three axial images were obtained at the apical, at the level of the carina and basal. The presence and extent of ILD, and the degree of diagnostic confidence were recorded. The effective dose for each image series was estimated. Standard HRCT was the standard of reference.The prevalence of ILD was 55% (25/45). Diagnostic sensitivity and accuracy of series 1, series 2 and series 3 were 100% and 94.4%, 94% and 97.8%, 92% and 97.8%, respectively. The extent of ILD was underestimated in series 3 (p0.05) and was comparable to the standard HRCT in series 1 and 2 (p0.05). Estimated dose reduction was more than 90% in all image series.HRCT image series with low sampling rate allow an accurate detection of ILD with very-low-radiation dose, making this approach potentially valuable for screening in patients with SSc.

The impact of human leukocyte antigen (HLA)-B27 on the age at disease onset in patients with ankylosing spondylitis (AS) has consistently been shown in several cohorts.1–4 A potential gender difference with respect to age at symptom onset remains, however, controversial5–8 as the distribution of HLA-B27 within these populations is not known in all studies. While no sex differences with regard to disease onset were found in several cohorts from Europe,5 ,6 a later disease onset in men was reported in a North American investigation.7 We analysed this issue within the ongoing Swiss Clinical Quality Management cohort of patients with axial spondyloarthritis (axSpA).9 Ethics approval was given by the regional review boards. Informed consent was obtained from all patients. From a total of 3046 spondyloarthritis patients recruited until June 2014, 2098 (1294 men, 804 women) were included in this analysis, as they fulfilled the Assessment in SpondyloArthritis International Society (ASAS) classification criteria, with the following minor modifications, as the cohort was initiated before the publication of these criteria.9 First, …

Systemic sclerosis (SSc) is characterised by a progressive microangiopathy that contributes significantly to the morbidity of patients with SSc. Besides insufficient angiogenesis, defective vasculogenesis with altered numbers of endothelial precursor cells (EPCs) might also contribute to the vascular pathogenesis of SSc. However, different protocols for isolation, enrichment, culture and quantification of EPCs are currently used, which complicate comparison and interpretation of the results from different studies.The aim of the European League Against Rheumatism Scleroderma Trials and Research (EUSTAR) group expert panel was to provide recommendations for standardisation of future research on EPCs. Consensus statements and recommendations were developed in a face to face meeting by an expert panel of the basic science working group of EUSTAR.The findings were: cardiovascular risk factors and medications such as statins should be described in detail. A detailed description of methods considering isolation, culture, enrichment and detection of EPCs should be given. For in vitro culture of EPCs, no protocol has been shown to be superior to another, but coating with laminin and type IV collagen would resemble most closely the situation in vivo. The endothelial phenotype should be confirmed in all in vitro cultures at the end of the culture period. We recommend using CD133, vascular endothelial growth factor type 2 receptor (VEGFR2) and CD34 in combination with a viability marker for quantification of EPCs in the blood. Finally, exact standard operating procedures for fluorescence-activated cell sorting (FACS) analysis are given that should be strictly followed.In summary, the EUSTAR recommendations will help to unify EPC research and allow better comparison between the results of different studies.

Background:Randomised trials have demonstrated that the efficacy of anti-tumour necrosis factor (TNF) agents is significantly increased by concomitant methotrexate (MTX) in rheumatoid arthritis (RA). In clinical routine, anti-TNF agents are commonly prescribed with other disease-modifying antirheumatic drugs (DMARDs) than MTX, however their effectiveness in combination with anti-TNF agents is not well established.Objective:To compare the effectiveness of leflunomide (LEF) and other conventional DMARDs with MTX as co-therapy to anti-TNF agents in RA.Methods:All patients on anti-TNF agents and conventional DMARDs within the Swiss Clinical Quality Management (SCQM)-RA database were included (n = 1218) and categorised according to the type of co-therapy into anti-TNF+MTX (n = 842), anti-TNF+LEF (n = 260) and anti-TNF+other DMARDs (n = 116). Drug discontinuation rates and incidence of toxic side effects were analysed using Cox proportional hazard models. Progression of radiographic damage, the evolution of functional disability and the improvement of RA disease activity were analysed using longitudinal regression models, adjusting for potential confounders.Results:The overall discontinuation rates of anti-TNF and conventional DMARD combination therapies were relatively high with a median survival of only 16 months (interquartile range (IQR): 10–37), but they did not differ between the three regimens (p = 0.69). The progression of radiographic damage (p = 0.77), functional disability (p = 0.09) and RA disease activity (p = 0.33) were also similar between the different regimen. In addition, no significant difference in the frequency of adverse events emerged.Conclusion:Overall these results suggest that LEF and potentially other conventional DMARDs offer an effective and safe alternative to MTX as co-therapy in combination with anti-TNF agents.

The metastasis-associated protein S100A4 belongs to the large family of S100 calcium-binding proteins that appear to play regulatory roles in diverse biological activities. Moreover, a prognostic role of S100A4 has been suggested for patients with several types of cancer. Cancer promoting properties for S100A4 have been demonstrated, particularly through its regulation of cell motility, proliferation and apoptosis, as well as by stimulation of angiogenesis and remodelling of the extracellular matrix. Increased expression of S100A4 mRNA has been detected in proliferating synovial fibroblasts in rheumatoid arthritis. Furthermore, strong upregulation of the S100A4 protein in rheumatoid arthritis synovial tissue compared with osteoarthritis and control tissues has been demonstrated recently, especially at sites of joint invasion. Several immune and vascular cells were also identified to be producing S100A4 within the synovium. The local upregulation of S100A4 was accompanied by high plasma and synovial fluid concentrations of the S100A4 protein existing in the bioactive oligomeric form in patients with rheumatoid arthritis. Consistent with data from cancer studies, the extracellular S100A4 oligomer appears to be involved in regulation of several matrix-degrading enzymes and modulation of the transcriptional activation function of the tumour suppressor protein p53 in rheumatoid arthritis synovial fibroblasts. Taken together, one can speculate that increased S100A4 protein in circulation and locally at sites of inflammation, particularly at sites of joint destruction, might be linked to the process of aggressive fibroblast behaviour contributing to the pathogenesis of chronic autoinflammatory diseases such as rheumatoid arthritis.

Stromal cells like synovial fibroblasts gained great interest over the years, since it has become clear that they strongly influence their environment and neighbouring cells. The current review describes the role of synovial fibroblasts as cells of the innate immune system and expands on their involvement in inflammation and cartilage destruction in rheumatoid arthritis (RA). Furthermore, epigenetic changes in RA synovial fibroblasts and studies that focused on the identification of different subsets of synovial fibroblasts are discussed.

More than a decade of use of the anti-tumour necrosis factor (anti-TNF) agents in rheumatology has confirmed their acceptable safety profile and very few new rare side effects have emerged. However, TNFα, a pleiotropic cytokine, is important to many physiological functions and its suppression may lead to unexpected consequences. We describe here a rare case of reversible oligoasthenozoospermia in a young patient. This case concerns a 35-year-old father of a healthy 4-year-old child who was successfully treated for 3 years with adalimumab, a monoclonal human antibody against TNFα, 40 mg subcutaneously every 2 weeks for ankylosing spondylitis (AS). The therapy …