BD750, a benzothiazole derivative, inhibits T cell proliferation by affecting the JAK3/STAT5 signalling pathway

Abstract

Background and Purpose A series of benzothiazole derivatives were screened for immunosuppressive activity; of these compounds BD750 was found to be the most effective immunosuppressant. The purpose of the current study was to determine the immunosuppressive activity of BD750 on T cell proliferation and its potential mode of action. Experimental Approach T cell proliferation, CD25 and CD69 expression and cell cycle distribution were measured in vitro by flow cytometry. Cell viability was determined by CCK‐8 assay. Cytokine levels were measured by elisa. The activation of signal‐regulated molecules was assessed by Western blot analysis. The effects of BD750 were evaluated in vivo in a mouse model of delayed‐type hypersensitivity. Key Results BD750 significantly inhibited mouse and human T cell proliferation, stimulated either by anti‐CD3/anti‐CD28 monoclonal antibodies or by an alloantigen, in a dose‐dependent manner in vitro. No obvious cytotoxic effects of BD750 were observed in our experimental conditions. Furthermore, BD750 did not inhibit CD25 and CD69 expression or IL‐2 and IL‐4 secretion, but induced cell cycle arrest at the G0/G1 phase in activated T cells. In IL‐2‐stimulated CTLL‐2 cells and primary activated T cells, BD750 inhibited cell proliferation and STAT5 phosphorylation, but not Akt or p70S6K phosphorylation. BD750 also reduced the T cell‐mediated delayed‐type hypersensitivity response in mice in a dose‐dependent manner. Conclusion and Implications These data indicate that BD750 inhibits IL‐2‐induced JAK3/STAT5‐dependent T cell proliferation. BD750 has the potential to be used as a lead compound for the design and development of new immunosuppressants for preventing graft rejection and treating autoimmune diseases.

Tolerogenic Dendritic Cells Induced by BD750 Ameliorate Proinflammatory T cell Responses and Experimental Autoimmune Encephalitis in Mice

“…Although it might be difficult for tolDC to pass the CNS (26), they did inhibit splenic Th1 and Th17 responses in EAE mice, similar to a previous report (26). In addition, although BD750 did not affect IL-10 production by LPS-stimulated DCs, BD750-DC secreted a low level of but not AKT phosphorylation, consistent with our previous findings (15). This observation suggests that BD750 may be a specific inhibitor of STAT5.…”

“…Inhibition of STAT5 can impair the maturation of monocyte-derived DCs with downregulated expression of costimulatory molecules and reduced secretion of IL-12 (27,28). Our previous study identified that BD750 is an inhibitor of JAK3/STAT5 and can inhibit T cell proliferation (15). In this study, we found that BD750 induced tolDC.…”

“…Our previous study identified that a benzothiazole derivative, BD750 [2-(2-benzothiazoleyl)-4,5,6,7-tetrahydro-2H-indazol-3-ol, C 14 H 13 N 3 OS, MW: 271.3], is an inhibitor of JAK3/STAT5 signaling and can inhibit T cell proliferation (15). JAK3 is crucial for the maturation of DCs, and JAK3 -/-DCs fail to induce T cell proliferation (16,17).…”

PO‐322 exerts potent immunosuppressive effects in vitro and in vivo by selectively inhibiting SGK1 activity

“…T cells (10 6 cells·ml −1 ) were treated with rapamycin (0.1 μM, Sigma‐Aldrich, St. Louis, MO, USA), FK506 (0.1 μM, Sigma‐Aldrich), GSK650394 (10 μM, MedChem Express, Monmouth Junction, NJ, USA), AG‐490 (50 μM, Sigma‐Aldrich), LY‐294002 (50 μM, Promega, Madison, WI, USA), PD184352 (2 μM, Sigma‐Aldrich), or different concentrations of PO‐322 and activated by plate‐bound anti‐CD3 (2 μg·ml −1 , HIT3a clone, BD Pharmingen, San Diego, CA, USA) and soluble anti‐CD28 (1 μg·ml −1 , CD28.2 clone, BD Pharmingen) as previously described (Liu et al, ). Cells were randomly assigned to treatment groups, and the experimenter was blinded to drug treatment until data analysis has been performed.…”

Activation of Toll-Like Receptor 3 Induces Interleukin-1 Receptor Antagonist Expression by Activating the Interferon Regulatory Factor 3

“…Human bone marrow mesenchymal stem cells (BMSC) were generated from marrow aspirates, as described previously [38]. Human T cells and B cells were isolated from human peripheral blood mononuclear cells by negative selection using the Pan T cell Isolation Kit II Human or the B cell Isolation Kit II Human (Miltenyi Biotec, Bergisch Gladbach, Germany), as previously described [39]. Mouse FLS were derived from the ankle joints of C57BL/6J mice at 10-12 weeks old.…”