Sander W. Tas

Macrophages from patients with SLE and rheumatoid arthritis have defective adhesion in vitro, while only SLE macrophages have impaired uptake of apoptotic cells

Background: It has been suggested that defective handling of apoptotic cells by macrophages plays a key role in the development of systemic lupus erythematosus (SLE). The relative contribution of intrinsic defects and serum factors remains controversial. Objective: To compare monocytes from SLE patients, patients with rheumatoid arthritis, and healthy controls for their ability to differentiate in vitro into macrophages and to bind/engulf apoptotic cells. Methods: Peripheral blood derived monocytes from healthy donors or from patients with SLE or rheumatoid arthritis were allowed to differentiate into macrophages. The in vitro uptake of apoptotic cells by macrophages was evaluated by a flow cytometry assay that allowed discrimination between binding and internalisation. Results: Monocytes from SLE and rheumatoid patients showed a striking defect in adherence to plastic compared with healthy donors. Absence or heat inactivation of serum resulted in a reduction in the binding and engulfment of apoptotic cells by macrophages. Macrophages from rheumatoid and SLE patients had similar percentages of apoptotic cells bound to their surface compared with normal controls. However, macrophages from SLE patients showed a significant defect in the internalisation of apoptotic cells compared with those from healthy controls, even in the presence of normal human serum. Conclusions: Monocytes from patients with SLE and rheumatoid arthritis have a similar defect in their capacity to adhere to plastic. However, only macrophages from SLE patients showed an impaired ability to engulf apoptotic cells, which indicates that an intrinsic cellular defect may be responsible for this phenomenon.

Signal Transduction Pathways and Transcription Factors as Therapeutic Targets in Inflammatory Disease: Towards Innovative Antirheumatic Therapy

Many chronic inflammatory diseases are associated with deregulated intracellular signal transduction pathways. Resultant pathogenic interactions between immune and stromal cells lead to changes in cell activation, proliferation, migratory capacity, and cell survival that all contribute to inflammation. Increasing efforts are now being made in the design of novel therapeutic compounds to interfere with signaling pathways in inflammatory diseases like rheumatoid arthritis (RA). In this review we will outline the major signal transduction pathways involved in the pathogenesis of RA. We will assess advances in targeting a number of key intracellular pathways, including nuclear factor-(kappa)B (NF-(kappa)B), mitogen-associated protein kinases (MAPKs), phosphoinositide 3-kinase (PI3K)/Akt, signal transducers and activators of transcription (STATs), and reactive oxygen species (ROS) production. Finally, we will discuss recently identified lead molecules and the progress of selected compounds towards becoming new drugs for the treatment of inflammatory diseases.

Enhanced gene transfer to arthritic joints using adeno-associated virus type 5: implications for intra-articular gene therapy

Background: Gene therapy of the joint has great potential as a new therapeutic approach for the treatment of rheumatoid arthritis (RA). The vector chosen is of crucial importance for clinical success. Objective: To investigate the tropism and transduction efficiency in arthritic joints in vivo, and in synovial cells in vitro, using five different serotypes of recombinant adeno-associated virus (rAAV) encoding β-galactosidase or green fluorescent protein genes. Methods: rAAV was injected into the ankle joints of rats with adjuvant arthritis after the onset of disease. Synovial tissue was examined at different time points for β-galactosidase protein and gene expression by in situ staining and polymerase chain reaction (PCR) analysis, respectively. In addition, the ability of rAAV to transduce primary human fibroblast-like synoviocytes from patients with RA was investigated in vitro. Results: Intra-articular injection of the rAAV5 serotype resulted in the highest synovial transduction, followed by much lower expression using rAAV2. Expression of the transgene was already detectable 7 days after injection and lasted for at least 4 weeks. Only background staining was seen for serotypes 1, 3, and 4. Importantly, there was a minimal humoral immune response to rAAV5 compared with rAAV2. Additionally, it was found that both rAAV2 and rAAV5 can efficiently transduce human fibroblast-like synoviocytes obtained from patients with RA. Conclusion: Intra-articular rAAV mediated gene therapy in RA might be improved by using rAAV5 rather than other serotypes.

Gene Therapy Targeting Nuclear Factor-κB: Towards Clinical Application in Inflammatory Diseases and Cancer

Nuclear factor (NF)-κB is regarded as one of the most important transcription factors and plays an essential role in the transcriptional activation of pro-inflammatory cytokines, cell proliferation and survival. NF-κB can be activated via two distinct NF-κB signal transduction pathways, the so-called canonical and non-canonical pathways, and has been demonstrated to play a key role in a wide range of inflammatory diseases and various types of cancer. Much effort has been put in strategies to inhibit NF-κB activation, for example by the development of pharmacological compounds that selectively inhibit NF-κB activity and therefore would be beneficial for immunotherapy of transplantation, autoimmune and allergic diseases, as well as an adjuvant approach in patients treated with chemotherapy for cancer. Gene therapy targeting NF-κB is a promising new strategy with the potential of long-term effects and has been explored in a wide variety of diseases, ranging from cancer to transplantation medicine and autoimmune diseases. In this review we discuss recent progress made in the development of NF-κB targeted gene therapy and the evolution towards clinical application.

Discovery of innovative therapies for rare immune-Mediated inflammatory Diseases via Off-Label Prescription of Biologics: The Case of IL-6 Receptor Blockade in Castleman's Disease

Biologics have revolutionized the field of clinical immunology and proven to be both effective and safe in common immune-mediated inflammatory diseases (IMIDs) such as rheumatoid arthritis, inflammatory bowel diseases, and various hematological disorders. However, in patients with rare, severe IMIDs failing on standard therapies, it is virtually impossible to conduct randomized controlled trials. Therefore, biologics are usually prescribed off-label in these often severely ill patients. Unfortunately, off-label prescription is sometimes hampered in these diseases due to a lack of reimbursement that is often based on a presumed lack of evidence for effectiveness. In the present article, we will discuss that off-label prescription of biologics can be a good way to discover new treatments for rare diseases. This will be illustrated using a case of multicentric Castleman’s disease, an immune-mediated lymphoproliferative disorder, in which off-label tocilizumab (humanized anti-IL-6 receptor blocking antibody) treatment resulted in remarkable clinical improvement. Furthermore, we will give recommendations for monitoring efficacy and safety of biologic treatment in rare IMIDs, including the use of registries. In conclusion, we put forward that innovative treatments for rare IMIDs can be discovered via off-label prescription of biologicals, provided that this is based on rational arguments including knowledge of the pathophysiology of the disease.

Nuclear Factor-κB–inducing Kinase Is Expressed in Synovial Endothelial Cells in Patients with Early Arthritis and Correlates with Markers of Inflammation: A Prospective Cohort Study

Objective. The nuclear factor-κB (NF-κB) family of transcription factors is strongly involved in synovial inflammation. We have previously shown that NF-κB–inducing kinase (NIK) is a key regulator of inflammation-induced angiogenesis in rheumatoid arthritis (RA) synovial tissue (ST). Here, we investigated synovial NIK expression in patients with early arthritis and in autoantibody-positive individuals at risk of developing RA. Methods. ST biopsies were obtained by arthroscopy from 154 patients with early arthritis (duration < 1 yr) with various diagnoses and 54 IgM rheumatoid factor–positive and/or anticitrullinated protein antibodies–positive individuals without evidence of arthritis. ST was stained for NIK and endothelial cell (EC) markers. Additionally, measures of disease activity were collected and contrast-enhanced magnetic resonance imaging (MRI) was performed in a subset of these patients. Results. In patients with early arthritis, NIK was predominantly expressed in EC of small blood vessels. Further, NIK expression correlated with erythrocyte sedimentation rate (r 0.184, p = 0.024), C-reactive protein (r 0.194, p = 0.017), joint swelling (r 0.297, p < 0.001), synovial immune cell markers (lining r 0.585, p < 0.001; sublining macrophages r 0.728, p < 0.001; T cells r 0.733, p < 0.001; and B cells r 0.264, p = 0.040), MRI effusion (r 0.665, p < 0.001), MRI synovitis (r 0.632, p < 0.001), and MRI total score (r 0.569, p < 0.001). In 18.5% of autoantibody-positive individuals, ST NIK+EC were present, but this was not predictive of the development of arthritis. Conclusion. NIK+EC are present in the earliest phase of synovial inflammation and may be indicative of high angiogenic activity in the inflamed ST. Therefore, NIK+EC may play an important role in the persistence of synovitis. Collectively, our data underscore the importance of angiogenesis in synovial inflammation and identify NIK as a potential therapeutic target in arthritis.

Castleman’s Disease

Castleman’s disease (CD) is a rare and relatively unknown lymphoproliferative disorder, with benign hyperplastic lymph nodes. The disease was first reported in 1956 by Benjamin Castleman, a pathologist from the Massachusetts General Hospital (Castleman and Towne, N Engl J Med 250:1001–1005, 1954). In this first case report, Castleman described a 60-year-old male with a mediastinal mass. Histology showed lymph node hyperplasia and follicles with small, hyalinized foci. Subsequently, in 1956, he described a series of 13 cases of localized asymptomatic mediastinal masses based on lymph node hyperplasia on X-ray (Castleman et al., Cancer 9:822–830, 1956). All of the patients described in these early papers had localized disease, which is now termed unicentric Castleman’s disease (UCD). In contrast, multicentric Castleman’s disease (MCD) is a systemic disease with generalized peripheral lymphadenopathy, hepatosplenomegaly, frequent episodes of fever and night sweats. In this chapter, we will discuss the pathophysiology, epidemiology, clinical presentation, diagnostic procedures, treatment and prognosis of both forms of CD.

Silencing NIK potentiates anti-VEGF therapy in a novel 3D model of colorectal cancer angiogenesis

Angiogenesis is essential for colorectal cancer (CRC) progression, as demonstrated by the beneficial clinical effects of therapeutics inhibiting VEGF signaling. However, alternative mechanisms of neovascularization can develop, resulting in treatment failure. Previously we demonstrated NF-κB-inducing kinase (NIK) contributes to pathological angiogenesis. Here, we investigate NIK as a therapeutic target in endothelial cells (EC) in CRC. To determine NIK expression levels in CRC tissues, we immunostained both primary colorectal tumors and tumors metastasized to the liver. Additionally, a 3D tumor-stromal cell interaction model was developed including EC, fibroblasts and CRC cells to study tumor angiogenesis. This model tested efficacy of NIK-targeting siRNA (siNIK) in EC alone or in combination with the anti-VEGF antibody, bevacizumab. Both primary CRC and liver metastases contained blood vessels expressing NIK. In patients receiving chemotherapy plus bevacizumab, immature NIK+ vessels (p < 0.05) were increased as compared to chemotherapy alone. Activation of NIK by lymphotoxin-beta receptor (LTβR) induced increases in pro-angiogenic mediators, including interleukin (IL)-6, IL-8, chemokine (C-X-C motif) ligand (CXCL)1 and CXCL5 in EC and fibroblasts, accompanied by sprouting in the 3D model, which was blocked by siNIK in EC. Treatment with bevacizumab plus siNIK in EC resulted in a synergistic effect and reduced VEGF and bFGF-induced sprouting (p < 0.05). Here, we demonstrate a role for NIK in CRC-associated angiogenesis. Targeting NIK in EC in combination with anti-VEGF antibody bevacizumab may hold therapeutic potential to increase efficiency in blocking tumor neovascularization, either to prevent treatment failure due to activation of accessory pathways such as NF-κB signaling or as a rescue treatment.

Corrigendum: Synovial tissue research: a state-of-the-art review

This corrects the article DOI: 10.1038/nrrheum.2017.115

03.18 Cd40-mediated activation of noncanonical NF-κB signalling in dendritic cells induces extrathymic autoimmune regulator (aire) expression

Background Nuclear factor (NF)-κB signalling plays an important role in the regulation of immune responses. We have previously demonstrated that CD40-mediated noncanonical NF-κB signalling in dendritic cells (DC) induces the immunoregulatory enzyme indoleamine 2,3-dioxygenase (IDO). Noncanonical NF-κB signalling is also important for AIRE expression in the thymus, which is essential for establishment of central tolerance. In the periphery, extrathymic AIRE-expressing antigen presenting cells (including DC) can induce or maintain peripheral tolerance. However, stimuli that induce extrathymic AIRE expression in DC are hitherto unknown. Materials and methods AIRE expressing cells were characterised by confocal microscopy in tonsil tissue and inflamed tissues from patients with rheumatoid arthritis (RA) and primary Sjögren’s Syndrome (pSS). Monocyte-derived DC (moDC), peripheral blood conventional DC (cDC) and plasmacytoid DC (pDC) were stimulated by anti-CD40 to induce NF-κB activation. Gene transcription and protein expression levels of AIRE, IDO and noncanonical NF-κB signalling components were detected by real time-PCR, Western blot and confocal microscopy. Expression of NF-κB-inducing kinase (NIK), the main activating kinase of the noncanonical pathway, was modulated by siRNA-mediated silencing. Results First, we identified AIRE expressing cells in inflamed tonsil tissue, RA synovial tissue and pSS glandular tissue and could demonstrate that these cells have DC characteristics, including MHCII, CD40 and CD11c expression. Next, we demonstrated that CD40-stimulation of moDC and pDC, but not cDC was accompanied by upregulation of AIRE expression. This upregulation was crucially dependent on noncanonical NF-κB signalling, since AIRE expression was abrogated by NIK siRNA-mediated silencing and was not affected by either inhibition or induction of canonical NF-κB signalling. Furthermore, preliminary data show that CD40-stimulated moDC from APECED (AIRE deficient) patients exhibited increased T cell proliferation in a mixed lymphocyte reaction compared to healthy individuals. Conclusions Collectively, our data demonstrate that AIRE expression in DC is critically dependent on CD40-mediated noncanonical NF-κB signalling. These findings suggest a role for peripheral AIRE expression in controlling T cell proliferation during adaptive immune responses, including the inflamed target tissues of patients with RA and pSS. These findings could potentially be exploited as a novel approach to boost peripheral tolerance and induce immune regulation.