Anti-cytokine therapy in chronic destructive arthritis

Tumor necrosis factor (TNF) and interleukin-1 (IL-1) are considered to be master cytokines in chronic, destructive arthritis. Therapeutic approaches in rheumatoid arthritis (RA) patients have so far focused mainly on TNF, which is a major inflammatory mediator in RA and a potent inducer of IL-1; anti-TNF therapy shows great efficacy in RA patients. However, it is not effective in all patients, nor does it fully control the arthritic process in affected joints of good responders. Directed therapy for IL-1, with IL-1 receptor antagonist, mainly reduces erosions and is marginally anti-inflammatory. It is as yet unclear whether the limited effect is akin to the RA process or linked to suboptimal blocking of IL-1. Analysis of cytokine patterns in early synovial biopsies of RA patients reveals a marked heterogeneity, with variable staining of TNF and IL-1β, indicative of TNF-independent IL-1 production in at least some patients. Evidence for this pathway emerged from experimental arthritises in rodents, and is summarized in this review. If elements of the models apply to the arthritic process in RA patients, it is necessary to block IL-1β in addition to TNF.     

Pathogenesis of rheumatoid arthritis and c-Fos/AP-1

c-Fos/AP-1 controls the expression of inflammatory cytokines and matrix-degrading matrix metalloproteinases (MMPs) important in arthritis via promoter AP-1 binding motif. Among inflammatory cytokines, IL-1β is the most important inducer of a variety of MMPs, and mainly responsible for cartilage breakdown and osteoclastogenesis. IL-1β and c-Fos/AP-1 influence each other’s gene expression and activity, resulting in an orchestrated cross-talk that is crucial to arthritic joint destruction, where TNFα can act synergistically with them. While how to stop the degradation of bone and cartilage, i.e., to control MMP, has long been the central issue in the research of rheumatoid arthritis (RA), selective inhibition of c-Fos/AP-1 does resolve arthritic joint destruction. Thus, the blockade of IL-1β and/or c-Fos/AP-1 can be promising as an effective therapy for rheumatoid joint destruction in addition to the currently available TNFα blocking agents that act mainly on arthritis.     

The role of T-cell interleukin-17 in conducting destructive arthritis: lessons from animal models

Interleukin-17 (IL-17) is a T cell cytokine spontaneously produced by cultures of rheumatoid arthritis (RA) synovial membranes. High levels have been detected in the synovial fluid of patients with RA. The trigger for IL-17 is not fully identified; however, IL-23 promotes the production of IL-17 and a strong correlation between IL-15 and IL-17 levels in synovial fluid has been observed. IL-17 is a potent inducer of various cytokines such as tumor necrosis factor (TNF)-alpha, IL-1, and receptor activator of NF-kappaB ligand (RANKL). Additive or even synergistic effects with IL-1 and TNF-alpha in inducing cytokine expression and joint damage have been shown in vitro and in vivo. This review describes the role of IL-17 in the pathogenesis of destructive arthritis with a major focus on studies in vivo in arthritis models. From these studies in vivo it can be concluded that IL-17 becomes significant when T cells are a major element of the arthritis process. Moreover, IL-17 has the capacity to induce joint destruction in an IL-1-independent manner and can bypass TNF-dependent arthritis. Anti-IL-17 cytokine therapy is of interest as an additional new anti-rheumatic strategy for RA, in particular in situations in which elevated IL-17 might attenuate the response to anti-TNF/anti-IL-1 therapy.     

Cytokine production of stimulated whole blood cultures in rheumatoid arthritis patients receiving short-term infliximab therapy

Patients with rheumatoid arthritis (RA) treated with anti-tumor necrosis factor (TNF) strategies have an increased susceptibility to infections, especially those caused by intracellular pathogens. In this study we assessed the cytokine production capacity in patients with RA and we further investigated whether anti-TNF therapy modulates the production of pro-inflammatory cytokines involved in the resistance against infections. Whole blood cultures from 10 RA patients and 10 healthy controls were stimulated with heat-killed Candida albicans, Salmonella typhimurium, Staphyloccocus aureus, Aspergillus fumigatus or Mycobacterium tuberculosis and production of interleukin (IL)-1beta, IL-6, IL-10, interferon (IFN)-gamma and TNF-alpha was measured. Before anti-TNF therapy, whole blood cultures from RA patients released significantly less IFN-gamma than healthy controls after stimulation with all tested microorganisms. Short-term anti-TNF therapy did not have an inhibitory effect on the release of the cytokines tested. We conclude that cells of patients with RA have a strongly reduced production capacity of IFN-gamma after bacterial challenge. Although short-term therapy with anti-TNF agents did not further decrease the release of other proinflammatory cytokines, the combination of defective IFN-gamma production in basal conditions and TNF neutralization during anti-TNF therapy is likely to be responsible for the higher susceptibility to infections in patients with RA.     

Positive regulators of osteoclastogenesis and bone resorption in rheumatoid arthritis

Bone destruction is a frequent and clinically serious event in patients with rheumatoid arthritis (RA). Local joint destruction can cause joint instability and often necessitates reconstructive or replacement surgery. Moreover, inflammation-induced systemic bone loss is associated with an increased fracture risk. Bone resorption is a well-controlled process that is dependent on the differentiation of monocytes to bone-resorbing osteoclasts. Infiltrating as well as resident synovial cells, such as T cells, monocytes and synovial fibroblasts, have been identified as sources of osteoclast differentiation signals in RA patients. Pro-inflammatory cytokines are amongst the most important mechanisms driving this process. In particular, macrophage colony-stimulating factor, RANKL, TNF, IL-1 and IL-17 may play dominant roles in the pathogenesis of arthritis-associated bone loss. These cytokines activate different intracellular pathways to initiate osteoclast differentiation. Thus, over the past years several promising targets for the treatment of arthritic bone destruction have been defined.     

Increased mRNA expression and protein secretion of interleukin-6 in primary human osteoblasts differentiated in vitro from rheumatoid and osteoarthritic bone

We have investigated the expression and synthesis of potential bone-resorbing cytokines, interleukin-6 (IL-6), interleukin-1 (IL-1), and tumor necrosis factor (TNF) in rheumatoid arthritic (RA) and osteoarthritic (OA) bone, two common diseases which are associated with bone loss. Primary human osteoblast (hOB) cultures were established to determine the temporal mRNA expression of IL-6, IL-1 (alpha and beta), and TNF (alpha and beta) in relation to osteoblast growth and phenotypic genes. IL-6 mRNA levels were found to be significantly higher (P < 0.04) in both OA hOB (17 patients) and RA hOB (10 patients) compared to normal (NO) hOB (9 patients) and reached five-fold increases in OA hOB and 13-fold increases in RA hOB. Maximal levels of IL-6 are expressed at Day 21 which corresponds to the mineralization stage reflected by decreasing collagen I (alpha(1)), osteopontin, bone sialoprotein, alkaline phosphatase mRNA levels, while osteocalcin (OC) mRNA levels increased. IL-6 protein levels also were significantly higher (P < 0.05) in OA hOB and RA hOB compared to NO hOB. These increases were not attributable to sex or age of the donor bone. Neither the mRNA encoding IL-1(alpha and beta) and TNF(alpha and beta) nor the related proteins were detectable. These results indicate that differentiated OA hOB and RA hOB within a bone tissue-like matrix constitutively express and secrete high levels of IL-6. This inherent property suggests that these osteoblasts, independent of local inflammatory parameters, can contribute to enhanced recruitment of osteoclast progenitors and thereby bone resorption.     

The role of T cell interleukin-17 in conducting destructive arthritis: lessons from animal models

Interleukin-17 (IL-17) is a T cell cytokine spontaneously produced by cultures of rheumatoid arthritis (RA) synovial membranes. High levels have been detected in the synovial fluid of patients with RA. The trigger for IL-17 is not fully identified; however, IL-23 promotes the production of IL-17 and a strong correlation between IL-15 and IL-17 levels in synovial fluid has been observed. IL-17 is a potent inducer of various cytokines such as tumor necrosis factor (TNF)-α, IL-1, and receptor activator of NF-κB ligand (RANKL). Additive or even synergistic effects with IL-1 and TNF-α in inducing cytokine expression and joint damage have been shown in vitro and in vivo. This review describes the role of IL-17 in the pathogenesis of destructive arthritis with a major focus on studies in vivo in arthritis models. From these studies in vivo it can be concluded that IL-17 becomes significant when T cells are a major element of the arthritis process. Moreover, IL-17 has the capacity to induce joint destruction in an IL-1-independent manner and can bypass TNF-dependent arthritis. Anti-IL-17 cytokine therapy is of interest as an additional new anti-rheumatic strategy for RA, in particular in situations in which elevated IL-17 might attenuate the response to anti-TNF/anti-IL-1 therapy.     

The combined effects of anti-TNFα antibody and IL-1 receptor antagonist in human rheumatoid arthritis synovial membrane

TNFα and IL-1 are the pivotal cytokines involved in rheumatoid arthritis (RA). More recently, the biological therapy targeting TNFα or IL-1 has been impressively effective for many RA patients, however, it remains insufficient in some patients. In the present study, we examined the combined effects of two agents against TNFα and IL-1 in human RA synovial membrane. Synovial explants (an ex vivo model) and synovial fibroblasts (an in vitro model) were prepared from 11 RA patients, and then anti-TNFα antibody (Anti-TNFα) and IL-1 receptor antagonist (IL-1Ra), either alone or in combination, were added to the synovial explants and fibroblasts. IL-6 and MMP-3 production were measured after incubation. As a result, their production significantly decreased by the combination of agents compared with the control group in both the synovial explants and fibroblasts. The efficacy of this combination was also observed for IL-6 production compared with each agent alone in the synovial explants, and for IL-6 and MMP-3 production compared with each agent alone in the synovial fibroblasts. Therefore, the combination of Anti-TNFα and IL-1Ra appears more beneficial in synovial membrane, particularly when compared with a single agent alone.     

Synovial expression of IL-15 in rheumatoid arthritis is not influenced by blockade of tumour necrosis factor

Blockade of tumour necrosis factor (TNF) is an effective treatment in rheumatoid arthritis (RA), but both non-responders and partial responders are quite frequent. This suggests that other pro-inflammatory cytokines may be of importance in the pathogenesis of RA and as possible targets for therapy. In this study we investigated the effect of TNF blockade (infliximab) on the synovial expression of IL-15 in RA in relation to different cell types and expression of other cytokines, to elucidate whether or not IL-15 is a possible target for therapy, independently of TNF blockade. Two arthroscopies with multiple biopsies were performed on nine patients with RA and knee-joint synovitis before and after three infusions of infliximab (3 mg/kg). Synovial biopsies were analysed with immunohistochemistry for expression of IL-15, TNF, IL-1α, IL-1ß and IFN-γ, and for the cell surface markers CD3, CD68 and CD163. Stained synovial biopsy sections were evaluated by computerized image analysis. IL-15 expression was detected in all synovial biopsies taken at baseline. After infliximab therapy, the expression of IL-15 was increased in four patients and reduced in five. Synovial expression of IL-15 was not correlated with any CD marker or with the presence of any other cytokine. Synovial cellularity was decreased after 8 to 10 weeks of treatment with a significant reduction of the CD68-positive synovial cells, whereas no significant change was seen in the number of CD3-positive T cells and CD163-expressing macrophages. The number of TNF-producing cells in the synovial tissue at baseline was correlated with a good response to therapy. Thus, in this study the synovial expression of IL-15 in RA was not consistently influenced by TNF blockade, being apparently independent of TNF expression in the synovium. Consequently, we propose that IL-15 should remain as a therapeutic target in RA, regardless of the response to TNF blockade.     

Synovial expression of IL-15 in rheumatoid arthritis is not influenced by blockade of tumour necrosis factor

Blockade of tumour necrosis factor (TNF) is an effective treatment in rheumatoid arthritis (RA), but both non-responders and partial responders are quite frequent. This suggests that other pro-inflammatory cytokines may be of importance in the pathogenesis of RA and as possible targets for therapy. In this study we investigated the effect of TNF blockade (infliximab) on the synovial expression of IL-15 in RA in relation to different cell types and expression of other cytokines, to elucidate whether or not IL-15 is a possible target for therapy, independently of TNF blockade. Two arthroscopies with multiple biopsies were performed on nine patients with RA and knee-joint synovitis before and after three infusions of infliximab (3 mg/kg). Synovial biopsies were analysed with immunohistochemistry for expression of IL-15, TNF, IL-1alpha, IL-1ss and IFN-gamma, and for the cell surface markers CD3, CD68 and CD163. Stained synovial biopsy sections were evaluated by computerized image analysis. IL-15 expression was detected in all synovial biopsies taken at baseline. After infliximab therapy, the expression of IL-15 was increased in four patients and reduced in five. Synovial expression of IL-15 was not correlated with any CD marker or with the presence of any other cytokine. Synovial cellularity was decreased after 8 to 10 weeks of treatment with a significant reduction of the CD68-positive synovial cells, whereas no significant change was seen in the number of CD3-positive T cells and CD163-expressing macrophages. The number of TNF-producing cells in the synovial tissue at baseline was correlated with a good response to therapy. Thus, in this study the synovial expression of IL-15 in RA was not consistently influenced by TNF blockade, being apparently independent of TNF expression in the synovium. Consequently, we propose that IL-15 should remain as a therapeutic target in RA, regardless of the response to TNF blockade.     

Interleukin-17 acts independently of TNF-alpha under arthritic conditions

The proinflammatory T cell cytokine IL-17 is a potent inducer of other cytokines such as IL-1 and TNF-alpha. The contribution of TNF in IL-17-induced joint inflammation is unclear. In this work we demonstrate using TNF-alpha-deficient mice that TNF-alpha is required in IL-17-induced joint pathology under naive conditions in vivo. However, overexpression of IL-17 aggravated K/BxN serum transfer arthritis to a similar degree in TNF-alpha-deficient mice and their wild-type counterparts, indicating that the TNF dependency of IL-17-induced pathology is lost under arthritic conditions. Also, during the course of the streptococcal cell wall-induced arthritis model, IL-17 was able to enhance inflammation and cartilage damage in the absence of TNF. Additional blocking of IL-1 during IL-17-enhanced streptococcal cell wall-induced arthritis did not reduce joint pathology in TNF-deficient mice, indicating that IL-1 is not responsible for this loss of TNF dependency. These data provide further understanding of the cytokine interplay during inflammation and demonstrate that, despite a strong TNF dependency under naive conditions, IL-17 acts independently of TNF under arthritic conditions.     

Antibody-mediated delivery of IL-10 inhibits the progression of established collagen-induced arthritis

The antibody-mediated targeted delivery of cytokines to sites of disease is a promising avenue for cancer therapy, but it is largely unexplored for the treatment of chronic inflammatory conditions. Using both radioactive and fluorescent techniques, the human monoclonal antibodies L19 and G11 (specific to two markers of angiogenesis that are virtually undetectable in normal adult tissues) were found to selectively localize at arthritic sites in the murine collagen-induced model of rheumatoid arthritis following intravenous (i.v.) administration. The same animal model was used to study the therapeutic action of the L19 antibody fused to the cytokines IL-2, tumour necrosis factor (TNF) and IL-10. Whereas L19–IL-2 and L19–TNF treatment led to increased arthritic scores and paw swellings, the fusion protein L19–IL-10 displayed a therapeutic activity, which was superior to the activity of IL-10 fused to an antibody of irrelevant specificity in the mouse. The anti-inflammatory cytokine IL-10 has been investigated for the treatment of patients with rheumatoid arthritis, but clinical development plans have been discontinued because of a lack of efficacy. Because the antigen recognised by L19 is strongly expressed at sites of arthritis in humans and identical in both mice and humans, it suggests that the fusion protein L19–IL-10 might help overcome some of the clinical limitations of IL-10 and provide a therapeutic benefit to patients with chronic inflammatory disorders, including arthritis.     

Levels of gastrin-releasing peptide and substance P in synovial fluid and serum correlate with levels of cytokines in rheumatoid arthritis

It is well known that cytokines are highly involved in the disease process of rheumatoid arthritis (RA). Recently, targeting of neuropeptides has been suggested to have potential therapeutic effects in RA. The aim of this study was to investigate possible interrelations between five neuropeptides (bombesin/gastrin-releasing peptide (BN/GRP), substance P (SP), vasoactive intestinal peptide, calcitonin-gene-related peptide, and neuropeptide Y) and the three cytokines tumour necrosis factor (TNF)-α, IL-6, and monocyte chemoattractant protein-1 in synovial fluid of patients with RA. We also investigated possible interrelations between these neuropeptides and soluble TNF receptor 1 in serum from RA patients. Synovial fluid and sera were collected and assayed with ELISA or RIA. The most interesting findings were correlations between BN/GRP and SP and the cytokines. Thus, in synovial fluid, the concentrations of BN/GRP and SP grouped together with IL-6, and SP also grouped together with TNF-α and monocyte chemoattractant protein-1. BN/GRP and SP concentrations in synovial fluid also grouped together with the erythrocyte sedimentation rate. In the sera, BN/GRP concentrations and soluble TNF receptor 1 concentrations were correlated. These results are of interest because blocking of SP effects has long been discussed in relation to RA treatment and because BN/GRP is known to have trophic and growth-promoting effects and to play a role in inflammation and wound healing. Furthermore, the observations strengthen a suggestion that combination treatment with agents interfering with neuropeptides and cytokines would be efficacious in the treatment of RA. In conclusion, BN/GRP and SP are involved together with cytokines in the neuroimmunomodulation that occurs in the arthritic joint.     

Expression of Cell Adhesion Molecules and Cytokines in Murine Antigen-Induced Arthritis

Adhesion molecules and cytokines are important in chronic inflammatory conditions such as rheumatoid arthritis (RA) by virtue of their role in cell activation and emigration. Using immunohistochemical techniques we studied the expression of adhesion molecules and cytokines in cryopreserved sections of murine knee joint in the course of antigen-induced arthritis, an animal model of human RA. Various adhesion molecules and cytokines are expressed in the arthritic joint tissue. LFA-I, Mac-1, CD44, ICAM-I and P-selectin were strongly expressed in the acute phase and to a lesser degree in the chronic phase of arthritis. VLA-4 and VCAM-I appeared to be moderately expressed on day 1, L-selectin between days 1 and 3. LFA-I, Mac-I, CD44, a4-integrin, ICAM-I and the selectins were found expressed on cells of the synovial infiltrate, LFA-1, Mac-1 and ICAM-I on the synovial lining layer, and VCAM-I and P-selectin on endothelial cells. Expression of E-selectin could be demonstrated throughout the experiment at a low level in cells of the acute cell infiltrate. Cytokines, especially IL-2, IL-4, IL-6, TNF, and IFN-7, were heavily expressed during the acute phase of arthritis in cellular infiltrate. Taken together these data demonstrate that cytokines and their activation of adhesion molecules contribute to cell infiltration and activation during the initial phase of arthritis and to the induction and progression of tissue destruction in arthritic joints. These molecules might be potential targets for novel therapeutic strategies in inflammatory and arthritic disorders.     

A follow-up to "Anti-cytokine therapy in chronic destructive arthritis" by Wim B van den Berg

In recent years, the effectiveness of anti-TNF therapy in treating rheumatoid arthritis (RA) has become apparent. While trials of IL-1 receptor antagonist in RA have been encouraging, it clearly is more difficult to target two molecules (IL-1 α and β) than one (TNF-α). In his review article, Professor Wim van den Berg argues that both TNF-α and IL-1 must be blocked in RA and that although TNF is clearly a potent inflammatory molecule, the dominant cytokine in the subsequent degradation of the joint tissue is IL-1. This commentary discusses his hypothesis in light of animal studies and the limitations of the conclusions that can be drawn from them. More broadly, it discusses the biology of TNF-α and IL-1 and suggests explanations of why TNF-α is a pivotal cytokine in this disease.     

Production of interleukin (IL)-1beta, IL-1 receptor antagonist and IL-10 by blood mononuclear cells in chronic arthritis

Joint erosion is a prevalent feature of rheumatoid arthritis (RA) but not of many other chronic inflammatory arthritides (non-RA). Joint destruction is mediated by cytokines, primarily interleukin (IL)-1 and tumour necrosis factor. Less erosive activity in patients with non-RA compared to RA might be related to factors that inhibit production and/or function of IL-1. Release of IL-1beta, and the two antagonists, IL-1 receptor antagonist (IL-1ra) and IL-10 from blood mononuclear cells were therefore quantitated by ELISA in 22 patients with RA, 11 with non-RA and 15 healthy age-matched controls. Release of IL-1beta was comparable between the three groups but only detectable in cultures stimulated with lipopolysaccharide; it decreased in patients treated with prednisolone: 3.8 ng/10(6)monocytes (median) vs 11.7 (P=0.045). Release of IL-1ra was in all but IgG-stimulated cultures comparable between groups. The ratio of IL-1ra/IL-1beta was elevated in LPS-stimulated cells from RA patients only: 2.0 versus 1.3 (P=0.02). In contrast, IgG-induced IL-1ra release was significantly elevated only in non-RA patients: 95 ng/10(6)monocytes vs 40 (P=0.014), and the levels correlated positively to those of blood CRP (P=0.02). Though stimulated release of IL-10 was similar between the three groups, the levels were lower in non-erosive than erosive arthritis patients, and controls (P=0. 05). In conclusion, increased IgG-stimulated IL-1ra release and elevated IL-1ra/IL-1beta ratio may protect against actions of IL-1 in vivo, and decreased release of IL-10 might be related to features of non-erosive arthritis.     

IL-23 Dependent and Independent Stages of Experimental Arthritis: No Clinical Effect of Therapeutic IL-23p19 Inhibition in Collagen-induced Arthritis

IL-23p19 deficient mice have revealed a critical role of IL-23 in the development of experimental autoimmune diseases, such as collagen-induced arthritis (CIA). Neutralizing IL-23 after onset of CIA in rats has been shown to reduce paw volume, but the effect on synovial inflammation and the immunological autoimmune response is not clear. In this study, we examined the role of IL-23 at different stages of CIA and during T cell memory mediated flare-up arthritis with focus on changes in B cell activity and Th1/Th17 modulation. Anti-IL-23p19 antibody (anti-IL23p19) treatment, starting 15 days after the type II collagen (CII)-immunization but before clinical signs of disease onset, significantly suppressed the severity of CIA. This was accompanied with significantly lower CII-specific IgG1 levels and lower IgG2a levels in the anti-IL-23p19 treated mice compared to the control group. Importantly, neutralizing IL-23 after the first signs of CIA did not ameliorate the disease. This was in contrast to arthritic mice that underwent an arthritis flare-up since a significantly lower disease score was observed in the IL-23p19 treated mice compared to the control group, accompanied by lower synovial IL-17A and IL-22 expression in the knee joints of these mice. These data show IL-23-dependent and IL-23-independent stages during autoimmune CIA. Furthermore, the memory T cell mediated flare-up arthritis is IL-23-mediated. These data suggest that specific neutralization of IL-23p19 after onset of autoimmune arthritis may not be beneficial as a therapeutic therapy for patients with rheumatoid arthritis (RA). However, T cell mediated arthritis relapses in patients with RA might be controlled by anti-IL-23p19 treatment.