FTY720--the first compound of a new promising class of immunosuppressive drugs

Immunosuppressive regimens containing cyclosporin or tacrolimus have reduced acute kidney graft rejections to an incidence as low as 10–30%, but acute rejections remain one of the strongest predictors for long-term kidney graft function [1–3]. Even though an increased dosage of calcineurin inhibitors can easily suppress rejection episodes, the risk of chronic transplant nephropathy, cardiovascular diseases and diabetes mellitus rises concomitantly with such treatments [4–6]. Therefore, standard immunosuppressive regimes combine calcineurin inhibitors with azathioprin, mycophenolate mofetil or rapamycin and corticosteroids, which inhibit an ongoing immune response synergistically. FTY720 is the first compound of a new class of immunosuppressive drugs that exhibit the potency to suppress initiation and continuation of a rejection using a different mechanism in comparison with other known immunosuppressive drugs. The immunosuppressive effect of FTY720 relies on inhibition of lymphocyte recirculation. The initiation as well as the effector phase of an immune response depends on lymphocyte recirculation. Naive T cells can only be activated by antigen-presenting dendritic cells in so-called secondary lymphoid organs, such as lymph nodes, spleen and Peyer's patches [7], and probably also in the bone marrow [8]. After organ transplantation, mature host and donor antigen-loaded dendritic cells migrate from the allotransplant into the T-cell zones of secondary lymphoid organs guided by cytokines and chemokines [9]. There, dendritic cells are able to inform the immune system by activating appropriate T cells for the remaining 2–3 days of their life. Naive and central memory T cells continuously leave the blood and enter lymph nodes through high endothelial venules in a multistep adhesion cascade. This process requires certain homing molecules expressed on T cells and endothelial cells, together with various chemokines, including CCL19 (also termed ELC, EBI ligand chemokine), CCL21 (SLC, secondary lymphoid tissue chemokine) and CXCL13 (BLC, B lymphocyte chemoattractant) which are recognized by the chemokine receptors CCR7 and CXCR5 [10–12]. After entering the paracortex of lymph nodes, naive and central memory T cells encounter numerous dendritic cells in their search for a stimulating antigen. If the T cells find nothing of interest, they leave the lymph node via efferent lymphatics and join the venous blood again via subclavian vein [13]. Twice daily, naive T cells circulate between blood and secondary lymphoid organs in search of appropriate antigens. Long-lived central memory T cells also follow this constitutive homing paradigm [14]. T cells that have been activated by antigen stimulation undergo differentiation into effector cells and clonal expansion, and migrate from the lymph node. Those effector T cells travel from the lymphatics back into the blood. With the ability acquired to leave the blood through post-capillary venules, they migrate into the tissue in order to reject the transplanted organ [15]. The egress of lymphocytes from the thymus into the blood as well as from the tissue of secondary lymphoid organs into the efferent lymphatic vessels is tightly regulated. Sphingosine-1-phosphate and its receptors are substantially involved in this immune regulatory step [16] (Figure 1). FTY720 inhibits lymphocyte traffic: naive thymus-derived T cells and memory T cells circulate between blood and secondary lymphoid organs in a search for appropriate antigens presented by dendritic cells. After antigen-dependent activation, T cells leave secondary lymphoid organs to travel via the blood to the allograft. FTY720 blocks egress of lymphocytes from both thymus and lymph nodes. Sphingosine-1 is generated during metabolism of sphingomyelin, an abundant component of biological membranes. Sphingosine-1 and FTY720, which share structural homology, are phosphorylated by a sphingosine kinase, expressed in various cells. The efflux of phosphorylated FTY720 (FTY720-P) from the cell is mediated by the multidrug transporter MDR-1 [17] (Figure 2). The phosphorylation of FTY720 is reversible in blood. Therefore, plasma pharmacokinetics of FTY720 show a stable equilibrium between FTY720 and a 2- to 4-fold higher concentration of FTY720-P [18] (Figure 2). FTY720-P but not the parental drug is recognized by four of the five sphingosine-1-phosphate receptors (S1PR1, S1PR3, S1PR4 and S1PR5) in nano- or picomolar concentrations [18]. These G-protein-coupled receptors are expressed on T cells (S1PR4 and S1PR5), endothelial cells (S1PR1 and S1PR3), vascular smooth muscle cells and myocardium (S1PR1 and S1PR2) [16], correlating with immunosuppressive and cardiovascular effects. Interestingly, FTY720 inhibits signalling through the S1PR1, S1PR2 and S1PR5 by inducing S1PR internalization at nanomolar concentrations, without activating or inducing recycling of the receptor [19]. The immunosuppressive effect of FTY720 correlates with the degree of blood lymphopoenia. Intravenous administration of FTY720 induces lymphopoenia in blood with a nadir within 4 h. At the same time, the number of lymphocytes markedly increases in peripheral lymph nodes, mesenteric lymph nodes and Peyer's patches, but not in spleen [20]. Myelo-monocytic cells are not affected [20]. It has been suggested that FTY720 induces lymphopoenia by increased influx of lymphocytes into lymph nodes and Peyer's patches due to an accelerated chemotactic response. Indeed, FTY720-P promotes homing of lymphocytes due to an enhanced leukotriene-dependent chemokine receptor CCR7-mediated chemotaxis [17]. However, experiments in CXCR5 and CCR7 knockout mice demonstrated that FTY720-induced lymphocyte sequestration in lymph nodes appears to involve additional mechanisms [21,22]. A single dose of FTY720 induces trafficking of lymphocytes to the abluminal side of the sinus lymphaticus-lining endothelium and the disappearance of lymphocytes from subcapsular and medullary areas rich in dendritic cells [18]. However, lymphocytes do not overcome the endothial barrier to enter the sinus lymphaticus. Egress of T and B cells into the efferent lymphatics is inhibited by FTY720. In accordance with this, lymphocytes are not detectable in the thoracic duct of FTY720-treated rats [20]. The mechanism of FTY720-regulated inhibition of cell egress remains uncertain at the cellular level. S1PRs are involved in the assembly of vascular endothelial cell–cell junctions [23]. FTY720 inhibits vascular permeability [24]. Therefore, the sinus-lining endothelium might contribute to the inhibition of lymphocyte egress under the influence of FTY720. FTY720 also inhibits the migration of naive T cells from thymus into the blood, resulting in a declining supply of naive T cells to the organism [25]. FTY720 is phosphorylated intracellulary before binding to sphingosine-1-phosphate receptors. In summary, the immunosuppressive effect of FTY720 reflects the induction of ignorance of peripheral antigens by various mechanisms: naive T cells are directed to the wrong place in the lymph node, which limits their ability to recognize antigens. The interruption of recirculation of naive T cells bearing a specific antigen receptor further reduces the probability of T cells encountering the appropriate antigen-presenting dendritic cell. Moreover, the number of naive T cells is reduced, since the organism is cut off from supply from the thymus. However, the effector phase is also inhibited by FTY720, since already activated CD4+ and CD8+ effector T cells are trapped in the draining lymph nodes and prevented from travelling to the lymph and the blood [26]. These immunosuppressive mechanisms, which are completely different from those alternative immunosuppressive agents, make this drug attractive for immunosuppressive protocols in transplantation and autoimmune disorders. Treatment of rats, mice, dogs or monkeys with FTY720 alone or in combination with cyclosporin prolonged survival of various allotransplants including skin, heart, liver, kidney and intestine [27]. In such experimental models, FTY720 treatment results in a marked reduction of graft-infiltrating lymphocytes [28]. The effectiveness of FTY720 in suppressing a pathological immune response has also been proven in experimental animal models of autoimmune encephalomyelitis, certain models of systemic lupus erythematosus and autoimmune myocarditis [29]. These promising results justified trials to test the immune-modulatory effect of FTY720 in patients. Pharmacokinetic data were evaluated in multicentre studies in stable kidney-transplanted patients maintained on cyclosporin and corticosteroids after single or multiple dosages of FTY720 in a range of 0.125–5.0 mg/day [30,31]. FTY720 displayed dose linearity with drug absorption independently of formulation and of interaction with food, and a high oral bioavailability of 37–71%. The maximum concentration (Cmax) is reached after 12–36 h due to a long absorption phase. FTY720 is metabolized extensively in the liver by cytochrome P450 4F2/3 enzymes. None of the identified metabolites exhibited immunosuppressive activity [32]. The metabolites could be detected in urine (40–50%) and in faeces (20–50%). Although FTY720 utilizes the common cytochrome P450 metabolic pathway, no interaction with cyclosporin or rapamycin has been described, since the P450 isoenzymes used are different. FTY720 displays an elimination half-life of 89–157 h. Impaired kidney function after renal transplantation was not associated with accumulation of FTY720. Recently, two phase II trials have analysed the safety and efficacy of FTY720 after kidney transplantation [33,34]. The administration of 5 mg FTY720 in combination with steroids, and a cyclosporin dose adjusted to blood levels at 2 h post-dose (C2) to achieve a difference in exposure from 50% of a full cyclosporin dose, reduced the incidence of acute biopsy-proven kidney graft rejection significantly when compared with a standard immunosuppressive regimen containing 2–3 g/day mycophenolate mofetil in combination with a full dose of cyclosporin and corticosteroids; 12.5 vs 17.9% [34]. FTY720 treatment was associated with a drop in lymphocyte count, with a nadir 4 h after initiation of the therapy. The drug was tolerated very well. The main adverse advent was bradycardia, which was reported in ∼25% of all FTY720-treated patients. Bradycardia was observed most often during the first 2 days after initiation of FTY720 treatment [34,35]. Although bradycardia was usually self-limited and modest, with a mean decrease in pulse rate of 13–15 b.p.m., bradycardia was managed using atropine or β-mimetics in ∼12% of FTY720-treated patients. The surprising negative chronotropic effect of FTY720 may be explained by the fact that it binds to S1PRs (S1PR1 and S1PR2) in the heart. Activation of S1PRs reduces the activity of adenylate cyclase and the intracellular level of cAMP in a β-adrenergic agonist-sensitive manner [36]. It is likely that FTY720-P uses the same pathway. Hypertensive kidney graft recipients pre-treated with β-blockers may therefore be at considerable risk of a further reduction of the heart rate after initiation of FTY720 therapy. However, the incidence of cardiac failure, arrhythmia, myocardial infarction and cardiac arrest in FTY720-treated patients was similar to the incidence reported in >2500 kidney graft recipients treated with cyclosporin, azathioprin and prednisolone [37]. The overall incidence and severity of infections did not differ between FTY720 and mycophenolate mofetil treatments. In patients receiving a single oral dose of FTY270, the peripheral blood lymphocyte counts were reduced by 30–70% and returned to baseline levels within 24 h [35]. Lymphopoenia persisted for >96 h only in patients treated with the highest dose (3.5 mg) of FTY270. In animal experiments, FTY720 impaired a delayed-type hypersensitivity reaction (DTH), the induction of T cell-mediated autoimmune responses and allograft rejections, while an immune response against systemic viral infections remained unaffected [39]. An antibody-mediated immune response against systemic viral infections is generated in secondary lymphoid organs including spleen, and does not require lymphocyte recirculation and migration of B and T cells to the periphery. In contrast, in order to combat a transplanted solid organ, T cells have to migrate from lymph nodes. Since FTY720 inhibits egress of lymphocytes from lymph nodes, the mode of action is in accordance with the results obtained from virus-infected mice. Indeed, the incidence of cytomegalovirus (CMV) infections in kidney graft recipients was lower in the FTY720 groups in comparison with the mycophenolate mofetil group [34]. However, CMV infections after transplantation are affected by several factors including immunosuppressive co-medication, donor and recipient immunity against CMV prior to transplantation, and others. In summary, FTY720 is a potent immunosuppressive agent that is well tolerated in patients. Due to its mode of action, which differs substantially from that of other known immunosuppressive drugs, and due to its different side effect profile, FTY720 is a promising partner in immunosuppressive regimens and will substantially extend the standard immunosuppressive repertoire. Conflict of interest statement. None declared. 1Division of Nephrology and Hypertension and 2Institute of Immunology, University Hospital of Schleswig-Holstein, Campus Kiel, Germany