Sphingosine kinase type 2 is essential for lymphopenia induced by the immunomodulatory drug FTY720

Abstract

FTY720, a potent immunomodulatory drug in phase 2/3 clinical trials, induces rapid and reversible sequestration of lymphocytes into secondary lymphoid organs, thereby preventing their migration to sites of inflammation. As prerequisite for its function, phosphorylation of FTY720 to yield a potent agonist of the sphingosine-1-phosphate receptor S1P 1 is required in vivo, catalyzed by an as-yet-unknown kinase. Here, we report on the generation of sphingosine kinase 2 (SPHK2) knockout mice and demonstrate that this enzyme is essential for FTY720 phosphate formation in vivo. Consequently, administration of FTY720 does not induce lymphopenia in SPHK2-deficient mice. After direct dosage of FTY720 phosphate, lymphopenia is only transient in this strain, indicating that SPHK2 is constantly required to maintain FTY720 phosphate levels in vivo. IntroductionNaive T cells regularly circulate between the bloodstream and lymphatic tissue in search for foreign antigen, as well as for tumor and autoantigen. Their activation in secondary lymphoid organs followed by regulated egress back into the circulation to reach sites of inflammation is a prerequisite for any adaptive immune response in the T-cell compartment. Recently, one of the G protein-coupled receptors for sphingosine-1-phosphate (S1P), namely the S1P 1 receptor, was shown to be crucial for the tempo-spatial trafficking of T cells into and out of the secondary peripheral lymphoid organs. 1 The importance of S1P 1 in lymphocyte trafficking became clear through studies with FTY720, an analog of sphingosine. FTY720, after phosphorylation in vivo to FTY720 phosphate (FTY720-P), induces a reversible sequestration of lymphocytes into lymph nodes and Peyer patches. 2,3 FTY720-P thereby acts as a functional antagonist of the S1P 1 receptor, thus inducing aberrant internalization and consequently rendering T cells unresponsive to the obligatory egress signal provided by S1P. 1,[4][5][6] FTY720 has emerged as a potent immunomodulatory agent with usefulness in the control of organ transplant rejection and for treatment of autoimmune diseases. In animals, FTY720 is efficacious in prolonging graft survival, as well as in models of multiple sclerosis, acute lung injury, autoimmune diabetes, atherosclerosis, and renal ischemia-reperfusion injury. 7 Promising results have been obtained from human trials on FTY720 for indications in renal transplantation and multiple sclerosis. 7,8 Since FTY720 prodrug activation is essential for its action on T (and B) cells, understanding how the drug gets phosphorylated in vivo is of high interest, in particular for the design of novel analogs with altered pharmacologic properties.FTY720 is known to be phosphorylated in vitro by the 2 mammalian sphingosine kinases SPHK1 and 2, with SPHK2 being considerably more efficient. [9][10][11] As shown by a recent study in SPHK1-deficient mice, 12 this enzyme appears to be dispensable for the action of FTY720 in vivo, as Sphk1-null mice are still rendered lymphopenic by the drug.In this study, we describe th...

Design and Synthesis of Selective and Potent Orally Active S1P5 Agonists

“…[1,2] FTY720-P activates sphingosine-1-phosphate (S1P) receptors S1P1, S1P3, S1P4, and S1P5 at low nanomolar concentrations and is inactive toward the S1P2 receptor.[1] The FTY720-P-mediated activation of the S1P1 receptor on lymphocytes induces receptor internalization, which attenuates T-cell response to S1P gradients, preventing their egress from secondary lymphoid tissues. [3] In addition to playing a role in the immune system, all S1P receptors except S1P4 are also found differentially expressed in the central nervous system [4] and on various tumor cell types.…”

“…[1,2] FTY720-P activates sphingosine-1-phosphate (S1P) receptors S1P1, S1P3, S1P4, and S1P5 at low nanomolar concentrations and is inactive toward the S1P2 receptor.…”

“…[1,2] FTY720-P activates sphingosine-1-phosphate (S1P) receptors S1P1, S1P3, S1P4, and S1P5 at low nanomolar concentrations and is inactive toward the S1P2 receptor. [1] The FTY720-P-mediated activation of the S1P1 receptor on lymphocytes induces receptor internalization, which attenuates T-cell response to S1P gradients, preventing their egress from secondary lymphoid tissues.…”

Sphingosine kinase 1 and 2 regulate the capacity of mesangial cells to resist apoptotic stimuli in an opposing manner

“…Previous studies by other groups also revealed unexpected results regarding sphingolipid levels in SK-1-and SK-2-deficient mice and tissues. In this context, Zemann et al (2006) reported that serum S1P levels in SK-2-deficient mice increased, but decreased by approximately 50% in SK-1-deficient mice. Moreover, Michaud et al (2006) measured S1P levels in forepaw tissue and found only slight decreases in SK-1-deficient mice.…”

Sphingolipids and membrane biology as determined from genetic models

“…SphK2 -/-mice are viable and sub-fertile [188]. Two recent reports have shown that SPHK2 is involved in the phosphorylation of the immune modulator FTY720 (a sphingosine analog) and that SPHK2 null mice do not respond to this drug by developing lymphopenia, as normal mice do [188,189]. In addition, circulating S1P (generated mostly by SphK1) and intrinsic mast cell SPHK2 together determine the anaphylactic response, a phenomenon that involves degranulation, which is a specialized form of exocytosis.…”