**5. Potential S1P-targeted therapy for RA**

Despite great efforts devoted to the development of new therapeutic targets for RA, currently used drugs have limitations in their use for the treatment of RA. As discussed in the above sections, S1P appears to be an important modulator of many aspects of the pathogenesis of RA. Manipulation of endogenous local and systemic amounts of bioactive S1P by inhibiting metabolic enzymes such as SphKs and SPL, by applying S1P-blocking agents such as FTY-720, VPC23019 and JTE-013 or anti-S1P antibodies is thus a promising approach for the treatment of RA. Studies targeting S1P metabolism and signalling for the treatment of RA are summarized in Table 1.

#### **5.1 Targeting S1P metabolism**

#### **5.1.1 Reducing S1P formation with sphingosine kinase inhibitors**

Blockade of SphK1 activity in the mouse model of CIA significantly suppressed articular inflammation and joint destruction, reduced disease severity, and down-regulated proinflammatory cytokine production and inflammatory cell recruitment into the synovium (Lai et al., 2009). Decreasing S1P production by inhibiting SphK activity is, therefore, a promising therapeutic option in chronic inflammatory arthritis. Indeed, the inhibition of S1P synthesis by blocking SphKs activity has proven useful as an anti-inflammation strategy in cancer therapy *in vitro* and in animal models (Billich et al., 2005; Bonhoure et al., 2006; French et al., 2006; Gamble et al., 2006; Leroux et al., 2007).

#### **5.1.2 Downregulation of S1P receptor activity with S1P lyase inhibitors**

SPL catalyzes the irreversible degradation of intracellular S1P to hexadecenal and phosphoethanolamine. It is the major and constitutively active S1P-degrading enzyme in cells and tissues (Fig. 1). As a result, S1P concentrations in tissues are maintained at very low levels. Inhibition of SPL leads to the accumulation of S1P in various tissues (Schwab et al., 2005). Treatment with SPL inhibitor results in accumulation of S1P in lymphoid tissues

TNF- is central to RA pathogenesis as the pro-inflammatory cytokine not only stimulates the production of other inflammatory mediators but also directly triggers the activation of synovial cells and osteoclasts leading to the irreversible damage of soft tissues and bone (Olsen and Stein, 2004). Several studies have suggested that S1P and TNF- might synergize in their regulation of synovial fibroblast functions. On the one hand, the expression of S1P3 receptor was enhanced by TNF-, leading to amplified secretion of cytokines/chemokines, including IL-6, IL-8, MCP-1, and RANTES (Zhao et al., 2008). On the other hand, S1P enhanced TNF--mediated COX-2 expression and production of PGE2 by RA synoviocytes (Kitano et al., 2006). Thus, the elevated levels of TNF- seen in RA synovium may increase S1P synthesis via activation of SphKs and make synovial fibroblasts more responsive to S1P possibly through up-regulation of S1P receptor expression. The synergy between TNF- and S1P may eventually exacerbate the clinical manifestations of RA, including enhanced synovial tissue invasion by aggressive fibroblasts (due to enhanced cell mobility), synovial hyperplasia (due to proliferation and survival of synoviocytes), and exacerbated

Despite great efforts devoted to the development of new therapeutic targets for RA, currently used drugs have limitations in their use for the treatment of RA. As discussed in the above sections, S1P appears to be an important modulator of many aspects of the pathogenesis of RA. Manipulation of endogenous local and systemic amounts of bioactive S1P by inhibiting metabolic enzymes such as SphKs and SPL, by applying S1P-blocking agents such as FTY-720, VPC23019 and JTE-013 or anti-S1P antibodies is thus a promising approach for the treatment of RA. Studies targeting S1P metabolism and signalling for the

Blockade of SphK1 activity in the mouse model of CIA significantly suppressed articular inflammation and joint destruction, reduced disease severity, and down-regulated proinflammatory cytokine production and inflammatory cell recruitment into the synovium (Lai et al., 2009). Decreasing S1P production by inhibiting SphK activity is, therefore, a promising therapeutic option in chronic inflammatory arthritis. Indeed, the inhibition of S1P synthesis by blocking SphKs activity has proven useful as an anti-inflammation strategy in cancer therapy *in vitro* and in animal models (Billich et al., 2005; Bonhoure et al., 2006;

SPL catalyzes the irreversible degradation of intracellular S1P to hexadecenal and phosphoethanolamine. It is the major and constitutively active S1P-degrading enzyme in cells and tissues (Fig. 1). As a result, S1P concentrations in tissues are maintained at very low levels. Inhibition of SPL leads to the accumulation of S1P in various tissues (Schwab et al., 2005). Treatment with SPL inhibitor results in accumulation of S1P in lymphoid tissues

**4.4.4 Synergistic action of S1P and TNF- on synovial fibroblast functions** 

inflammation (due to over production of inflammatory mediators).

**5.1.1 Reducing S1P formation with sphingosine kinase inhibitors** 

**5.1.2 Downregulation of S1P receptor activity with S1P lyase inhibitors** 

French et al., 2006; Gamble et al., 2006; Leroux et al., 2007).

**5. Potential S1P-targeted therapy for RA** 

treatment of RA are summarized in Table 1.

**5.1 Targeting S1P metabolism** 


Table 1. Targeting the SphK/S1P/S1PR signalling pathway in RA.

and induces premature internalization of the exit-signal-sensing S1P1 receptor on lymphocytes (Schwab et al., 2005). S1P1 receptor internalization renders lymphocytes unresponsive to S1P, preventing their egress from the thymus and lymph nodes (Lo et al., 2005). One physiological outcome of this systemic redistribution of lymphocytes is potent immunosuppression, which offers new opportunities for developing immunoregulatory agents to treat autoimmune and inflammatory diseases (Gardell et al., 2006; Huwiler and Pfeilschifter, 2008; Mandala et al., 2002; Rosen et al., 2003; Zhang and Schluesener, 2007). Indeed, SPL-deficient mice showed resistance to various inflammatory and autoimmune challenges (Bagdanoff et al., 2009; Bandhuvula et al., 2007; Vogel et al., 2009). The evaluation of a synthetic SPL inhibitor, LX2931, is currently underway in phase II clinical trials in RA patients (Bagdanoff et al., 2010).

Targeting the Metabolism and Receptors of

**5.2.2 S1P receptor antagonists** 

**6. Conclusion** 

the treatment of RA.

**7. Acknowledgment** 

of cancer progression (reviewed in Peyruchaud, 2009).

Sphingosine-1-Phosphate for the Treatment of Rheumatoid Arthritis 205

FTY-720 treatment clearly reduced these pathological parameters. Similarly, Matsuura et al. (2000) compared FTY-720 with two other anti-rheumatic compounds, mizoribine and prednisolone, in the rat models of CIA and AIA. FTY-720 completely suppressed the increase in hind paw volume and bone destruction to normal control levels by inhibiting leukocyte accumulation in the arthritic joint. Moreover, FTY-720 was shown to possess antiarthritic activity with a wider margin of safety in AIA and CIA models as compared to mizoribine and prednisolone. A recent study by Tsunmi et al. (2010) revealed that FTY-720 administration suppressed the progression of laminarin-induced arthritis in SKG mice. FTY-720 treatment decreased IL-6 and TNF- expression by synovial fibroblasts, diminished the number of inflammatory cells migrating into the�joints, and suppressed bone destruction.

Blocking S1P receptor activity by using S1P receptor antagonists may lead to promising therapeutic strategies for patients suffering from RA. Despite the lack of more comprehensive *in vivo* data, considerable progress has been made in the identification of the S1P receptors regulating synovial fibroblast migration, production of cytokines/chemokines and PGE2, proliferation, and survival. Several compounds targeting S1P receptors have also been used to decipher the biological roles of S1P receptors. The S1P1/3 antagonist VPC23019 and S1P3 antagonist CAY10444 blocked S1Pmediated synoviocyte migration and cytokine secretion, whereas the S1P2 antagonist JTE-013 attenuated S1P-mediated cytokine secretion (Kitano et al., 2006; Zhao et al., 2008). The pharmacokinetics, bioavailability and metabolic characteristics of these S1P antagonists are essential to advance *in vivo* studies and for therapeutic intervention. Indeed, pharmacological approaches have been developed to block the action of S1P in the context

In summary, SphKs and S1P/S1PR signalling appear to play essential roles in modulating RA pathogenesis. The SphK1 pathway is activated and likely plays a pro-inflammatory role in mouse models of inflammatory arthritis. It is fascinating that the blockade of SphK1 activity results in the simultaneous reduction of several inflammatory responses such as pro-inflammatory cytokines and inflammatory cell infiltration into the synovium. Excessive S1P and enhanced S1P receptor expression are detected in the synovium of RA patients. S1P and signalling through S1P receptors induce expression of inflammatory cytokines and suppress apoptosis of B lymphoblastoid cells and fibroblast-like synoviocytes. Although the possible mechanism by which S1P exerts its activity in RA remains to be fully characterized, further understanding of S1P metabolism and S1P receptor expression by synovial tissues represents an exploitable objective for the development of novel chemotherapeutic agents in

The authors would like to thank Ms. Lynn Davis for her editorial assistance. The works cited in the review from the authors' laboratory were supported by research grants from the

Canadian Arthritis Network and the Arthritis Society of Canada.
