T cell immunoglobulin-3 as a new therapeutic target for rheumatoid arthritis

Abstract

T cell immunoglobulin-3 (Tim-3) is a surface molecule expressed on various cell types of the immune system which plays a central role in immune regulation. Recently, identification of galectin-9 (Gal-9) as a ligand for Tim-3 has established the Tim-3–Gal-9 pathway as an important regulator of Th1 immunity and induction of tolerance. The interaction of Tim-3 with Gal-9 induces cell death; the in vivo blockade of this interaction results in exacerbated autoimmunity and abrogation of tolerance in experimental models, thus establishing Tim-3 as a negative regulatory molecule. A number of previous studies have demonstrated that Tim-3 influences chronic autoimmune diseases, such as multiple sclerosis and systemic lupus erythematosus. In addition, an association between Tim-3 polymorphisms and susceptibility to several autoimmune diseases has been identified in various autoimmune diseases, including rheumatoid arthritis (RA). Recent work has focused on the role of Tim-3 in RA, and the results indicate that Tim-3 may represent a novel target for the treatment of RA. In this article we will discuss the Tim-3 pathway and the therapeutic potential of modulating the Tim-3 pathway in RA.

Keywords::

• autoimmune
• galectin-9
• rheumatoid arthritis
• T cell immunoglobulin-3

1. Introduction

Rheumatoid arthritis (RA) is a chronic systemic inflammatory disease characterized by proliferation of synovial cells and uncontrolled inflammation, resulting in irreversible joint destruction that causes loss of function, disability, and significant use of health care resources [1,2]. RA is present in 0.5 – 1% of the adult population worldwide, and is more common in women [3]. Nevertheless, the etiology of RA is incompletely understood. There is increasing evidence that genetic, environmental, hormonal, immunologic, and infectious factors may be involved in the persistent immunologic response that is central to the pathogenesis of RA [4,5]. Interestingly, the cells involved in the innate and adaptive immune systems drive the excessive immune responses which occur in RA.

Recently, it has been reported that a novel immunoregulatory molecule (T-cell immunoglobulin- and mucin-domain-containing molecule-3 [Tim-3]) expressed on T cells influences chronic autoimmune diseases, such as systemic lupus erythematosus, multiple sclerosis, and RA [6,7]. Moreover, an association between Tim-3 polymorphisms and susceptibility to RA in the Korean and Chinese Hui populations has been described [8,9]. Tim-3 is preferentially expressed on CD4⁺ T helper 1 (Th1) and T helper 17 (Th17) cells [10]. Moreover, it has been shown that galectin-9 (Gal-9), a ligand for Tim-3, inhibits the development of collagen-induced arthritis (CIA) in mice by suppressing the generation and differentiation of Th17, and promoting the induction of regulatory T cells (Tregs), which known to be suppressor T cells [11]. A recent study has reported the high expression of Tim-3 and related cytokines in the synovial tissues and peripheral mononuclear cells from patients with RA [12]. The results demonstrated increased expression of the Tim-3 ligand, Gal-9, in RA patients compared with healthy controls. In addition, Lee et al. [13] showed that Tim-3 and Gal-9 induced apoptosis of CD4⁺ T cells in RA. Collectively, these data suggest the Tim-3 and the interaction between Tim-3 and Gal-9 may play a pivotal in the pathogenesis of RA.

In the present review we provide a brief overview of the molecular properties of Tim-3, discuss the Tim-3 pathway, and the therapeutic potential of modulating the Tim-3 pathway in RA.

2. Tim-3/galectin-9 pathway

Tim-3, a member of the T cell Ig and mucin domain-containing molecule (Tim) superfamily, was first discovered in 2002 as a novel molecule specifically expressed on IFN-γ-secreting CD4⁺ Th1 and CD8⁺ T cytotoxic 1 (Tc1) cells in mice [6]. Two years later, Khademi et al. [14] showed that Tim-3 is also specifically expressed on IFN-γ-secreting T cells in human Th1 cells [14]. Moreover, the effects of targeting Tim-3 in autoimmune diseases have recently been reported [15]. Studies have revealed that administration of antibody against Tim-3 in proteolipid protein (PLP)-induced experimental autoimmune encephalomyelitis (EAE) significantly exacerbates the clinical and pathologic severity of EAE, and increases the number and level of activation of macrophages [6,16]. These observations were subsequently extended to show that in vivo treatment with Tim-3-Ig fusion protein promotes a hyperproliferative response in CD4⁺ cells and increases IFN-γ production [16]. Several similar studies complemented these findings, and showed that in vivo blockade of Tim-3 using blocking monoclonal antibodies or fusion proteins exacerbate experimental allergic encephalomyelitis and autoimmune diabetes [17,18].

Recent data also suggest that Tim-3 may impact anti-tumor immunity through the promotion of CD8⁺ T cell exhaustion in chronic immune conditions [18,19]. In addition, Tim-3 is involved in establishing tolerance. Although the precise mechanism of action by which Tim-3 establishes tolerance is not known, it is proposed that Tim-3 may regulate the graft-prolonging effects of anti-CD154/donor-specific transfusion by inducing the generation of donor-specific Tregs [18,20]. Collectively, in addition to inducing autoimmune diseases by regulating Th1 cell function, Tim-3 has also been implicated in establishing tolerance.

Although Tim-3 was originally described as a surface protein that can specifically identify Th1 markers, it has recently been demonstrated that Tim-3 is not only expressed on T cells, but also on several other cell types, including cytotoxic CD8⁺ T cells, Tregs, monocytes, microglia, dendritic cells (DCs), and mast cells in mice and humans [21]. Interestingly, stimulation of Tim-3 in murine DCs and human monocytes has been reported to promote the secretion of pro-inflammatory cytokines, such as TNF, between Tim-3 and toll-like receptor (TLR) signaling [22,23]. In the initial stages of the immune response, it is likely that Tim-3 on DCs promotes inflammation by synergizing with TLRs. In the final stages of the immune response, TIM3 on differentiated Th1 cells, which also up-regulates Gal-9 expression by IFN-γ, binds Gal-9, thus leading to cell death and Th1-cell response inhibition [24,25].

It has been proposed that Tim-3 is expressed on murine Th 17 cells, which are closely associated with the etiology of RA and play a crucial role in the pathogenesis of RA [10]. How the function of Tim-3 in different cell types is regulated and which cell type predominates in different disease states are unknown.

Galectin-9 (Gal-9), a member of the S-type lectins [26], was identified as a Tim-3 ligand by immunoprecipitation of cell surface proteins that bind to Tim-3–Ig [27]. Gal-9 is expressed on a variety of cell lines and is up-regulated by IFN-γ [7,28]. In vitro experiments have shown that Gal-9 induces intracellular calcium flux, and aggregation and death of Th1 cells is Gal-9-dependent [24]. Moreover, administration of Gal-9 triggering of Tim-3 on Th1 cells during an ongoing immune response in vivo diminishes inflammation by specifically eliminating IFN-γ-producing T cells, thereby attenuating the progression of EAE [24]. More recently, evidence in support of the relationship between the Tim-3/Gal-9 pathway and immunopathogenesis has been provided by Sakai [29], who reported that Gal-9 ameliorates acute graft-versus-host-disease (aGVHD), possibly by inducing T-cell apoptosis, suggesting that Gal-9 may be an attractive candidate for the treatment of aGVHD [29]. Moreover, Veenstra et al. [30] reported that in contrast to the aGVHD effects, Tim-3/Gal-9 pathway inhibition is dependent upon the presence of donor Tregs. To date, Gal-9 is the only ligand for Tim-3 which has been identified. In contrast, Oomizu et al. [31] showed that Gal-9 can inhibit Th17 cell differentiation through an IL-2-dependent pathway, but a Tim-3-independent pathway. Moreover, recent studies have revealed potential ligands for TIM-3, including phosphatidylserine and carbohydrate [7].

3. Tim-3 in the pathogenesis of RA

Tim-3 is a Th1–associated cell surface protein that inhibits aggressive Th1-mediated autoimmune responses and promotes immune tolerance in mice [20,32].Tim-3 is also reported to promote or terminate Th1 immune responses and may influence a series of inflammatory conditions. The relative levels of expression of Tim-3 have previously been reported in peripheral blood mononuclear cells (PBMC) from patients with RA [33]. Interestingly, it was also shown that the percentage of Tim-3 positive cells is higher in PBMC from patients with RA than healthy individuals based on flow cytometric analyses [33]. Moreover, it was further demonstrated that the expression of Tim-3 is up-regulated on various immune cells, including CD4⁺ T, CD8⁺ T cells and NKT cells from PBMCs in patients with RA, which was inversely correlated with disease activity. Expression of Tim-3 on CD8⁺ T and NKT cells negatively correlates with disease activity score 28 (DAS28) and plasma TNF-α in patients with RA. Recently, another study showed that the relative levels of mRNA expression of Tim-3 is high in synovial tissues (STs) from RA patients than osteoarthritis (OA) controls [12]. Similarly, Gal-9 mRNA is highly expressed in PBMC of RA than healthy controls, which is positively correlated with the expression of FoxP3, the master Treg [12]. Gal-9 is expressed in the STs and fibroblast-like synoviocytes and is clearly more highly expressed in patients with RA than patients with OA [34]. Moreover, Lee et al. [12]showed that the lower level of Tim-3 mRNA on CD4⁺ T cells in human RA compared to CD4⁺ T-cells from normal subjects causes blunted Gal-9-mediated apoptosis of CD4⁺ T cells that negatively regulates the Th1 response.

Emerging evidence suggests that Tim-3 has a protective role in RA. Therefore, Gal-9 administration induces decreased levels of pro-inflammatory Th1 and Th17 cytokine levels in joints, thus causing suppressed CIA arthritic symptoms in mice. Reports from a number of investigators have demonstrated a genetic association between Tim-3 and patients with RA, supporting a potential role for Tim-3 in human RA [8,9]. Chae et al. [8] reported that the −574T > G and 4259G > T polymorphisms of Tim-3 are more prevalent in RA patients than normal controls within the Korean population, suggesting that genetic polymorphisms might be associated with susceptibility to RA. Also, the -1541 C > T and +4259 G > T polymorphisms of Tim-3 are associated with RA susceptibility in the Chinese Hui population [9].

4. Therapeutic implications for targeting Tim-3 in RA

Few reports have focused on Tim-3 targeting therapy in RA. Because the Tim-3–Gal-9 pathway may play a crucial role in the induction and development of systemic autoimmune diseases, inhibition of the pathway in RA appears to be a reasonable proposal at this time. Indeed, the recent demonstration of the efficacy of targeting the Tim-3 pathway in animal models of RA positions Tim-3 as an attractive candidate for clinical development.

Targeting Tim-3 by Gal-9 in a mouse CIA model, first described by Seki et al. [11], effectively inhibited the severity of CIA, including pannus tissue formation, marked inflammatory cell infiltration, and bone and cartilage destruction. Moreover, treatment with Gal-9 also down-regulated pro-inflammatory cytokines (IL-17, IL-12, and IFN-γ) in mouse joints, and decreased the number of CD4⁺ Tim-3+ T cells in peripheral blood. Interestingly, Gal-9-deficient mice become susceptible to CIA, which may be linked to the increased number of CD4⁺ Tim-3+ T cells and decreased number of Tregs [11]. When evaluated in vitro, the administration of Gal-9 induced differentiation of naive T cells to Tregs, and suppressed the activity and differentiation of Th17 cells [11]. Gal-9 has been shown to prohibit the production of TNF-α and IL-1β, but increase the production of IL-10 by macrophages stimulated with immune complexes [35]. Similarly, treatment of Gal-9 also suppressed the levels of pro-inflammatory cytokines in the joints and serum C5a in the anti-CII monoclonal antibody (mAb)-induced arthritis (CAIA) murine model [35]. Moreover, Fcγ receptor expression in Gal-9–deficient macrophages was down-regulated, while Fcγ receptor expression was up-regulated from Gal-9-treated mice, thus indicating that Gal-9 suppresses immune complex-induced arthritis partly by regulating Fcγ receptor expression.

5. Conclusions and future perspectives

Since the original discovery of Tim-3, considerable progress has been made in characterizing Tim-3 ligands, and Tim-3 expression and function in autoimmune diseases. It is now well-known that in addition to Th1 and Tc1 cells, Tim-3 is also expressed on several other cell types during RA development, suggesting that the Tim-3/Gal-9 pathway may play an important role in the etiology of RA. However, limited data about the functions of Tim-3 modulation in RA has largely been derived from murine models. One must be cognizant of the fact that animal models may not mimic the complex human situation in all circumstances. Therefore, additional studies are required, not only in animals, but in humans to further illustrate the clear relationship between Tim-3 and RA.

Declaration of interest

This work was supported by the China National Science Foundation grants (30901526, 81101372, and 81270011).