YM-58483, a selective CRAC channel inhibitor, prevents antigen-induced airway eosinophilia and late phase asthmatic responses via Th2 cytokine inhibition in animal models

Abstract

T cells play a regulatory role in the pathogenesis of various immune and allergic diseases, including human asthma. Recently, it was reported that a pyrazole derivative, YM-58483 (BTP2), potently inhibits Ca²⁺ release-activated Ca²⁺ (CRAC) channels and interleukin (IL)-2 production in T cells. We investigated the effects of YM-58483 on T helper type 2 (Th2) cytokine production in vitro and antigen-induced airway asthmatic responses in vivo. YM-58483 inhibited IL-4 and IL-5 production in a conalbumine-stimulated murine Th2 T cell clone (D10.G4.1), and IL-5 production in phytohemagglutinin-stimulated human whole blood cells with IC₅₀ values comparable to those reported for its CRAC channel inhibition (around 100 nM). YM-58483 inhibited antigen-induced eosinophil infiltration into airways, and decreased IL-4 and cysteinyl-leukotrienes content in inflammatory airways induced in actively sensitized Brown Norway rats. Furthermore, orally administered YM-58483 prevented antigen-induced late phase asthmatic broncoconstriction and eosinophil infiltration in actively sensitized guinea pigs. These data suggest that the inhibition of Ca²⁺ influx through CRAC channel leads to the prevention of antigen-induced airway inflammation, probably via the inhibition of Th2 cytokine production and inflammatory mediators release. YM-58483 may therefore be useful for treating airway inflammation in bronchial asthma.

Introduction

Bronchial asthma is a complex disease which is characterized by chronic airway inflammation involving the activation of immune cells and release of multiple cytokines. T cells, especially antigen-specific CD4-positive T cells, and cytokines from these cells are believed to play a critical role in the pathogenesis of bronchial asthma (Larché et al., 2003, Kay, 2006, Ngoc et al., 2005). Recent reports support the predominance of the T helper 2 (Th2) phenotype in the asthmatic lung, such as in the excessive production of type 2 cytokines including interleukin (IL)-4, IL-5 and IL-13 (Georas et al., 2005, Borish et al., 2001, Leckie et al., 2000, Izuhara et al., 2006). Thus, agents which modulate the function of T cells or these inflammatory cytokines are expected to be useful in the treatment of asthma (Yamagata and Ichinose, 2006, Ichinose and Barnes, 2004, O'Byrne et al., 2004).

The Ca²⁺ release-activated Ca²⁺ (CRAC) channel mediates store depletion-activated inward Ca²⁺ current (I_CRAC) in T cells and other inflammatory cells (Hoth and Penner, 1992, Anant, 2005). Sustained Ca²⁺ influx through CRAC channels activates several transcription factors including NFAT, which regulates the expression of cytokine genes critical for the immune response (Fanger et al., 1995, Crabtree, 2001). CRAC channels are therefore considered to play a critical role in the activation of T cells, such as cell proliferation and cytokine production. The importance of CRAC channels in the immunologic response is evidenced by the severe deficiencies in T cell activation and effector gene expression in patients lacking store-operated Ca²⁺ entry (Partiseti et al., 1994, Le Deist et al., 1995, Feske et al., 2001). Consequently, CRAC channels have attracted considerable attention as potential targets for novel therapeutics for autoimmune diseases and chronic inflammation. Recently, we and others have reported that a pyrazole derivative YM-58483 (BTP2) potently inhibit both Ca²⁺ influx through CRAC channels and NFAT-driven IL-2 production in activated T cells (Ishikawa et al., 2003, Djuric et al., 2000, Trevillyan et al., 2001). YM-58483 is the first selective and potent inhibitor of CRAC channels and subsequent Ca²⁺ signals (Ishikawa et al., 2003, Zitt et al., 2004). This compound inhibits a wide spectrum of cytokine production and T cell proliferation in concanavaline A-stimulated human peripheral blood mononuclear cells (Chen et al., 2002). Furthermore, YM-58483 has been shown to inhibit hapten-induced contact hypersensitivity in sensitized mice in vivo (Ishikawa et al., 2003). These findings raise the intriguing possibility that YM-58483 may inhibit inflammatory responses observed in various allergic diseases, such as bronchial asthma.

In the present study, we investigated the effects of YM-58483 on Th2 cytokine production in vitro, and on antigen-induced airway eosinophilia and airway responses in vivo, to determine whether YM-58483 could inhibit allergic airway inflammation.