Janus kinase 1/3 signaling pathways are key initiators of TH2 differentiation and lung allergic responses.

BACKGROUND: Janus kinases (JAKs) are regulators of signaling through cytokine receptors. The importance of JAK1/3 signaling on TH2 differentiation and development of lung allergic responses has not been investigated. OBJECTIVE: We sought to examine a selective JAK1/3 inhibitor (R256) on differentiation of TH subsets in vitro and on development of ovalbumin (OVA)-induced airway hyperresponsiveness (AHR) and inflammation in an experimental model of asthma. METHODS: A selective JAK1/3 inhibitor was used to assay the importance of this pathway on induction of TH1, TH2, and TH17 differentiation in vitro. In vivo, the effects of inhibiting JAK1/3 signaling were examined by administering the inhibitor during the sensitization or allergen challenge phases in the primary challenge model or just before provocative challenge in the secondary challenge model. Airway inflammation and AHR were examined after the last airway challenge. RESULTS: In vitro, R256 inhibited differentiation of TH2 but not TH1 or TH17 cells, which was associated with downregulation of signal transducer and activator of transcription (STAT) 6 and STAT5 phosphorylation. However, once polarized, TH2 cells were unaffected by the inhibitor. In vivo, R256 administered during the OVA sensitization phase prevented the development of AHR, airway eosinophilia, mucus hypersecretion, and TH2 cytokine production without changes in TH1 and TH17 cytokine levels, indicating that selective blockade of TH2 differentiation was critical. Inhibitor administration after OVA sensitization but during the challenge phases in the primary or secondary challenge models similarly suppressed AHR, airway eosinophilia, and mucus hypersecretion without any reduction in TH2 cytokine production, suggesting the inhibitory effects were downstream of TH2 cytokine receptor signaling pathways. CONCLUSIONS: Targeting the TH2-dependent JAK/STAT activation pathway represents a novel therapeutic approach for the treatment of asthma.

Suppression of skin and kidney disease by inhibition of spleen tyrosine kinase in lupus-prone mice.

OBJECTIVE: Spleen tyrosine kinase (Syk) is involved in membrane-mediated signaling in various cells, including immune cells. It is overexpressed in T cells from patients with systemic lupus erythematosus (SLE), and its inhibition has been shown to improve T cell function as well as to improve disease manifestations in (NZB x NZW)F(1) lupus-prone mice and in patients with rheumatoid arthritis. While clinical trials examining Syk inhibition in patients with SLE are being considered, the aim of our experiments was to determine whether the therapeutic effects of Syk inhibition extend to other strains of lupus-prone mice and whether they result in improvement in skin disease and modification of established disease. METHODS: Female MRL/lpr or BAK/BAX mice were studied. Starting either at age 4 weeks (before disease) or at age 16 weeks (after established disease) and continuing for up to 16 weeks, mice were fed chow containing the Syk inhibitor R788 or control chow. RESULTS: We found that inhibition of Syk in MRL/lpr and BAK/BAX mice prevented the development of skin disease and significantly reduced established skin disease. Similarly, Syk inhibition reduced the size of the spleen and lymph nodes, suppressed the development of renal disease, and suppressed established renal disease. Discontinuation of treatment resulted in extended suppression of skin disease for at least 8 weeks and suppression of renal disease for 4 weeks. CONCLUSION: Syk inhibition suppresses the development of lupus skin and kidney disease in lupus-prone mice, suppresses established disease in lupus-prone mice, and may represent a valuable treatment for patients with SLE.

JAK3 inhibition significantly attenuates psoriasiform skin inflammation in CD18 mutant PL/J mice.

JAK3, a member of the Janus kinase family, is predominantly expressed in hemopoietic cells and binds specifically to the common gamma chain of a subfamily of cytokine receptors that includes IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21. Previous studies suggest that this tyrosine kinase plays key roles in mediating T cell functions, and inhibition of JAK3 has been shown to prevent graft rejection and decrease the severity of arthritis in rodent models. However, the functions of JAK3 in the development of skin immune responses and diseases such as psoriasis have not been determined. CD18 mutant PL/J mice develop spontaneous T cell-dependent psoriasiform skin disease with several similarities to human psoriasis. In this study, we treated mice with established skin disease with R348, a small molecule inhibitor of JAK3, and observed a marked attenuation of skin lesions following 6 wk of treatment. Histological analyses revealed major reductions of both epidermal and dermal lesion severity scores in R348-treated CD18-deficient PL/J mice compared with vehicle controls, which was associated with decreased CD4(+) T cell infiltration. In addition, systemic levels of IL-17, IL-22, IL-23, and TNF-alpha were significantly lower in mice receiving the compound, and T cells isolated from R348-treated mice also showed reduced phosphorylation of Stat5 after stimulation with IL-2. These findings suggest that small-molecule inhibitors of JAK3 may be useful in the treatment of inflammatory skin diseases such as psoriasis and strongly implicate JAK signaling events as important in the pathogenesis of this disease.

An orally bioavailable spleen tyrosine kinase inhibitor delays disease progression and prolongs survival in murine lupus.

OBJECTIVE: To assess whether R788, an orally bioavailable small molecule inhibitor of spleen tyrosine kinase (Syk)-dependent signaling, could modulate disease in lupus-prone (NZB x NZW)F1 (NZB/NZW) mice via inhibition of Fc receptor (FcR) and B cell receptor signaling. METHODS: R788 was administered to NZB/NZW mice before and after disease onset. Proteinuria, blood urea nitrogen levels, and autoantibody titers were examined periodically, and overall survival and renal pathologic features were assessed following long-term treatment (24-34 weeks). The distribution and immunophenotype of various splenic T cell and B cell subpopulations were evaluated at the time of study termination. Arthus responses in NZB/NZW mice pretreated with R788 or Fc-blocking antibody (anti-CD16/32) were also examined. RESULTS: When R788 was administered prior to or after disease onset, it delayed the onset of proteinuria and azotemia, reduced renal pathology and kidney infiltrates, and significantly prolonged survival of lupus-prone NZB/NZW mice; autoantibody titers were minimally affected throughout the study. Dose-dependent reductions in the numbers of CD4+ activated T cells expressing high levels of CD44 or CD69 were apparent in spleens from R788-treated mice. Minimal effects on the numbers of naive T cells expressing CD62 ligand and total CD8+ T cells per spleen were observed following long-term drug treatment. R788 pretreatment resulted in reduced Arthus responses in NZB/NZW mice, similar to results obtained in mice pretreated with FcR-blocking antibody. CONCLUSION: We demonstrate that a novel Syk-selective inhibitor prevents the development of renal disease and treats established murine lupus nephritis. These data suggest that Syk inhibitors may be of therapeutic benefit in human lupus and related disorders.

Inflammation and bone erosion are suppressed in models of rheumatoid arthritis following treatment with a novel Syk inhibitor.

Spleen tyrosine kinase (Syk), a key mediator of immunoreceptor signaling in inflammatory cells, is essential for immune complex-mediated signal transduction initiated by activated receptors for immunoglobulin G. In collagen-induced arthritis, R788/R406, a novel and potent small molecule Syk inhibitor suppressed clinical arthritis, bone erosions, pannus formation, and synovitis. Serum anti-collagen type II antibody levels were unaltered, while the half-life of exogenous antibody was extended when co-administered with R406. Expression of the targeted kinase (Syk) in synovial tissue correlated with the joint level of inflammatory cell infiltrates and was virtually undetectable in treated rats. Syk inhibition suppressed synovial cytokines and cartilage oligomeric matrix protein (COMP) in serum, suggesting a sensitive and reliable biomarker for R406 activity. These results highlight the role of activating Fcgamma receptors in inflammatory synovitis and suggest that interruption of the signaling cascade with a novel Syk inhibitor may be a useful addition to immunosuppressive disease-modifying anti-rheumatic drugs currently used in the treatment of human autoimmune diseases such as rheumatoid arthritis.