Inhibitors of the JAK family of non-receptor tyrosine kinases have demonstrated clinical efficacy in rheumatoid arthritis and other inflammatory disorders; however, the precise mechanisms by which JAK inhibition improves inflammatory immune responses remain unclear. Here we examined the mode of action of tofacitinib (CP-690,550) on JAK/STAT signaling pathways involved in adaptive and innate immune responses. To determine the extent of inhibition of specific JAK/STAT-dependent pathways, we analyzed cytokine stimulation of mouse and human T cells in vitro. We also investigated the consequences of CP-690,550 treatment on Th cell differentiation of naïve murine CD4+ T cells. CP-690,550 inhibited IL-4-dependent Th2 cell differentiation, and interestingly also interfered with Th17 cell differentiation. Expression of IL-23 receptor and of the Th17 cytokines IL-17A, IL-17F and IL-22 were blocked when naïve Th cells were stimulated with IL-6 and IL-23. In contrast, IL-17A-production was enhanced when Th17 cells were differentiated in the presence of TGF-β. Moreover, CP-690,550 also prevented activation of STAT1, induction of T-bet and subsequent generation of Th1 cells. In a model of established arthritis, CP-690,550 rapidly improved disease by inhibiting production of inflammatory mediators and suppressing STAT1-dependent genes in joint tissue. Furthermore, efficacy in this disease model correlated with inhibition of both JAK1 and JAK3 signaling pathways. CP-690,550 also modulated innate responses to LPS in vivo through a mechanism likely involving inhibition of STAT1 signaling. Thus, CP-690,550 may improve autoimmune diseases and prevent transplant rejection by suppressing the differentiation of pathogenic Th1 and Th17 cells, as well as innate immune cell signaling.
The title compound {systematic name: bis[2-(1,3-dioxoisoindol-2-yl)ethyl]azanium chloride dihydrate}, C20H18N3O4
+·Cl−·2H2O, is a phthalimide-protected polyamine that was synthesized by a previous method. It was characterized by ESI–MS, 1H NMR, and FT–IR. Crystals were grown from a solution of H2O and 0.1 M HCl. The central nitrogen atom is protonated and forms hydrogen bonds with the chloride ion and a water molecule. The two phthalimide units make a dihedral angle of 22.07 (3)°. The crystal packing features a hydrogen-bond network, two-coordinated chloride, and off-set π–π stacking.
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