Binding Immunoglobulin Protein (BIP) Inhibits TNF-α-Induced Osteoclast Differentiation and Systemic Bone Loss in an Erosive Arthritis Model.

OBJECTIVE: The association between inflammation and dysregulated bone remodeling is apparent in rheumatoid arthritis and is recapitulated in the human tumor necrosis factor transgenic (hTNFtg) mouse model. We investigated whether extracellular binding immunoglobulin protein (BiP) would protect the hTNFtg mouse from both inflammatory arthritis as well as extensive systemic bone loss and whether BiP had direct antiosteoclast properties in vitro. METHODS: hTNFtg mice received a single intraperitoneal administration of BiP at onset of arthritis. Clinical disease parameters were measured weekly. Bone analysis was performed by microcomputed tomography and histomorphometry. Mouse bone marrow macrophage and human peripheral blood monocyte precursors were used to study the direct effect of BiP on osteoclast differentiation and function in vitro. Monocyte and osteoclast signaling was analyzed by Western blotting, flow cytometry, and imaging flow cytometry. RESULTS: BiP-treated mice showed reduced inflammation and cartilage destruction, and histomorphometric analysis revealed a decrease in osteoclast number with protection from systemic bone loss. Abrogation of osteoclast function was also observed in an ex vivo murine calvarial model. BiP inhibited differentiation of osteoclast precursors and prevented bone resorption by mature osteoclasts in vitro. BiP also induced downregulation of CD115/c-Fms and Receptor Activator of NF-κB (RANK) messenger RNA and protein, causing reduced phosphorylation of the p38 mitogen-activated protein kinases, extracellular signal-regulated kinases 1/2 and p38, with suppression of essential osteoclast transcription factors, c-Fos and NFATc1. BiP directly inhibited TNF-α- or Receptor Activator of NF-κB Ligand (RANKL)-induced NF-κB nuclear translocation in THP-1 monocytic cells and preosteoclasts by the canonical and noncanonical pathways. CONCLUSION: BiP combines an anti-inflammatory function with antiosteoclast activity, which establishes it as a potential novel therapeutic for inflammatory disorders associated with bone loss.

Safety and patient response as indicated by biomarker changes to binding immunoglobulin protein in the phase I/IIA RAGULA clinical trial in rheumatoid arthritis.

OBJECTIVES: Binding immunoglobulin protein (BiP) is a human endoplasmic reticulum-resident stress protein. In pre-clinical studies it has anti-inflammatory properties due to the induction of regulatory cells. This randomized placebo-controlled, dose ascending double blind phase I/IIA trial of BiP in patients with active RA, who had failed accepted therapies, had the primary objective of safety. Potential efficacy was measured by DAS28-ESR and changes in biomarkers. METHODS: Twenty-four patients with active RA who had failed one or more DMARDs were sequentially assigned to three groups each of eight patients randomly allocated to receive placebo (two patients) or BiP (six patients), 1, 5 or 15 mg. Patients received a single i.v. infusion over 1 h and were observed as inpatients overnight. A 12-week follow-up for clinical, rheumatological and laboratory assessments for safety, efficacy (DAS28-ESR) and biomarker analysis was performed. RESULTS: No infusion reactions or serious adverse drug reactions were noted. Adverse events were evenly distributed between placebo and BiP groups with no BiP-related toxicities. Haematological, renal and metabolic parameters showed no drug-related toxicities. Remission was only achieved by patients in the 5 and 15 mg groups, and not patients who received placebo or 1 mg BiP. Good DAS28-ESR responses were achieved in all treatment groups. The BiP responding patients showed significantly lower serum concentrations of CRP, 2 weeks post-infusion compared with pre-infusion levels, and of VEGF and IL-8 from the placebo group. CONCLUSION: BiP (⩽15 mg) is safe in patients with active RA. Some patients had clinical and biological improvements in RA activity. BiP merits further study. TRIAL REGISTRATION: ISRCTN registry, http://isrctn.com, ISRCTN22288225 and EudraCT, https://eudract.ema.europa.eu, 2011-005831-19.

Immunoglobulin heavy-chain-binding protein (BiP): a stress protein that has the potential to be a novel therapy for rheumatoid arthritis.

Immunoglobulin heavy-chain-binding protein (BiP) or glucose-regulated protein 78 (Grp78) is a vital ubiquitous resident of the endoplasmic reticulum (ER). As an intracellular chaperone, BiP correctly folds nascent polypeptides within the ER and regulates the unfolded protein response ensuring protection of the cell from denatured protein and reinforcing its anti-apoptotic role, when the cell is under stress. Additionally, BiP is a member of the heat-shock protein (HSP) 70 family and, as a stress protein, is up-regulated by conditions of reduced oxygen and glucose. Cell stress induces surface expression and secretion of BiP. Consequently, BiP is detectable in several bodily fluids including serum, synovial fluid (SF) and oviductal fluid. However, as an extracellular protein, BiP has additional properties that are quite distinct from the intracellular functions. Extracellular BiP is immunoregulatory and anti-inflammatory causing development of tolerogenic dendritic cells (DCs), induction of regulatory T-cells, abrogation of osteoclast development and function, induction of anti-inflammatory cytokine production, including interleukin (IL)-10, IL-1 receptor antagonist and soluble tumour necrosis factor (TNF)-receptor type II, and attenuation of TNFα and IL-6. Together, these functions help drive the resolution of inflammation. Disease models of inflammatory arthritis have helped to demonstrate the novel mode of action of BiP in which the pharmacokinetics and pharmacodynamics are dissociated. The three murine models to be discussed each show BiP induced long-term therapeutic protection and therefore has potential for long-lasting drug-free therapy in rheumatoid arthritis (RA).

Stress cytokines: pivotal proteins in immune regulatory networks; Opinion.

Stress proteins have three immunological regulatory functions: within the cell, on the cell membrane as signalling receptors, and in the extracellular environment as stress cytokines. They can activate the immune system by providing danger signals or they may downregulate immune and inflammatory responses. In addition, they can modulate immune responses by acting as chaperones for antigenic peptides while they themselves are processed and presented to T cells as self-peptides. We predict that the exploitation of the downregulatory properties of stress cytokines will have therapeutic applications in the treatment of human chronic inflammatory diseases, such as rheumatoid arthritis.

Inhibition of antigen-presenting cell function and stimulation of human peripheral blood mononuclear cells to express an antiinflammatory cytokine profile by the stress protein BiP: relevance to the treatment of inflammatory arthritis.

OBJECTIVE: The stress protein and endoplasmic reticulum chaperone, immunoglobulin binding protein (BiP), is an autoantigen in rheumatoid arthritis (RA). Stress proteins, however, may have extracellular functions, mediated via cell surface receptors, that may include immunomodulatory functions. We sought to determine whether cell-free BiP is present in the synovial fluid (SF) of patients with RA and to further investigate the possible extracellular antiinflammatory and immunomodulatory properties of BiP in peripheral blood mononuclear cells (PBMCs) in vitro. METHODS: The presence of BiP in SF was established by Western blotting. PBMCs were stimulated with exogenous recombinant human BiP, and cytokine production and cell proliferation were measured in the presence and absence of cell signaling inhibitors or neutralizing anti-interleukin-10 (anti-IL-10) monoclonal antibody. Cytokine levels were quantified by enzyme-linked immunosorbent assay, cell proliferation by tritiated thymidine uptake, and cell surface molecule expression by flow cytometry. RESULTS: PBMCs responded to BiP with secretion of an antiinflammatory profile of cytokines. Although BiP stimulated the early production of tumor necrosis factor alpha (TNF alpha), the major cytokine induced was IL-10. Soluble TNF receptor II and IL-1 receptor antagonist secretion was also increased. Addition of SB203580, the MAPK p38 pathway inhibitor, partially inhibited the production of IL-10 and TNF alpha, whereas they were unaffected by the MAPK ERK-1/2 inhibitor PD98059. BiP also inhibited the recall antigen response by PBMCs to tuberculin purified protein derivative. Further investigation showed that incubation of monocytes in the presence of either BiP or IL-10 down-regulated CD86 and HLA-DR expression. The effect observed with IL-10 was transient compared with the long-lasting reduction induced by BiP. CONCLUSION: Extracellular BiP may stimulate immunomodulatory and antiinflammatory pathways, which are only partly due to the production of IL-10. These properties may be of relevance for the treatment of diseases such as RA.

Stromal cell-derived factor 1 (CXCL12) induces human cell migration into human lymph nodes transplanted into SCID mice.

Stromal cell-derived factor 1 (SDF-1; CXCL12), a CXC chemokine, has a primary role in signaling the recruitment of hemopoietic stem cell precursors to the bone marrow during embryonic development. In postnatal life, SDF-1 is widely expressed and is induced in chronically inflamed tissues such as psoriatic skin and the rheumatoid synovium, but has also been implicated in the migration of lymphocytes to lymphoid organs. To investigate the role of SDF-1 in recirculation and homing in vivo, we have developed a model in which human peripheral lymph nodes (huPLN) are transplanted into SCID mice. We have shown that huPLN transplants are viable, vascularized by the murine circulation that forms functional anastomoses with transplant vessels. In addition, grafts retain some features of the pretransplantation tissue, such as lymphoid follicles, lymphatic and high endothelial venule markers. We also show that SDF-1 is capable of inducing the migration of a SDF-1-responsive cell line (U937) and human PBLs from the murine circulation into the grafts in a dose-dependent manner, inhibitable by CXCR4 blockade. The mechanism of action of SDF-1 in this model is independent from that of TNF-alpha and does not rely on the up-regulation of adhesion molecules (such as ICAM-1) on the graft vascular endothelium. This is the first description of huPLN transplantation into SCID mice and of the functional effects of SDF-1 in regard to the migration of human cells into huPLN in vivo. This model provides a powerful tool to investigate the pathways involved in cell migration into lymphoid organs and potentially to target them for therapeutic purposes.