FTY720: A New Dimension in Transplantation
`
`B.D. Kahan
`
`IN CONTRAST TO the subfamily of neutrophil and
`
`mononuclear chemotactic cytokines (chemokines) up-
`regulated by inflammation,1 FTY720 seems to affect the
`interactions of lymphocytes with the subfamily of homing
`chemokines, secondary lymphoid tissue chemokine (SLC)2
`and essential myosin light chain chemokine (ELC).3 These
`homing chemokines are constitutively expressed in T-cell
`zones of secondary lymphoid structures (SLS), including
`peripheral lymph nodes (PLN), Peyer’s patches (PP), ap-
`pendix, and spleen (SPL), wherein they attract lymphocytes
`and dendritic cells bearing the corresponding receptors
`CCR7 or CXCR3 (SLC) or only CCR7 (ELC).
`The chemokine receptor CCR7 is expressed on stomal
`elements within T-cell zones, on dendritic cells (DC),3 on B
`cells, on T lymphocytes bearing the Th1 as opposed to Th2
`phenotype,4 and on memory cells remaining within SLS in
`contrast to the circulating effectors that generate inflamma-
`tory mediators.5 (CCR7 is not present on granulocytes or
`monocytes.6) Following binding of CCR7 to ELC and SLC,
`lymphocytes display activation of integrin adhesion mole-
`cules,7 events necessary for incorporation into the critical
`microenvironment for immune maturation. Indeed, cells
`from CCR7 knockout (KO) mice fail to develop delayed-
`type hypersensitivity reactions or primary antibody responses.8
`
`ROLE OF CHEMOKINES IN ALLOIMMUNITY
`
`A complex pattern of inflammatory chemokines is gener-
`ated during alloimmune responses.9 During the early
`phases of acute rejection in rodent allografts, the T-lym-
`phocyte chemoattractants, lymphotactin10 and SINC/KC,
`show augmented rates of production. Thereafter, both
`RANTES (regulated on activation T cell expressed and
`secreted), a marker of T-cell infiltration, and interferon
`(IFN)-inducible protein-10 (IP-10) are increased in rodent
`renal11 and in human cardiac grafts. In human liver trans-
`plant
`rejection, macrophage
`inflammatory
`protein
`(MIP)-1␤and MIP-1␣are increased, events that potentiate
`the activating effects of IFN-␥ on mouse macrophage
`cytokine production and antagonize Th2 cytokines.12 Cor-
`respondingly, human renal allografts undergoing rejection
`display infiltrating mononuclear cells bearing the G-pro-
`tein-coupled receptors (GPCR) CXCR4 and CCR5.13 The
`pleiotropic expression of MIP-1␣/␤, monocyte chemoat-
`tractant protein (MCP)-1, RANTES, and IP-10, as well as
`their corresponding receptors, namely, CCR1 (MIP-1␣),
`
`© 2001 by Elsevier Science Inc.
`655 Avenue of the Americas, New York, NY 10010
`
`Transplantation Proceedings, 33, 3081–3083 (2001)
`
`CCR2 (MCP-1), CCR5 (MIP-1␣/␤, RANTES), and
`CXCR3 (IP-10), has been confirmed in rejecting major
`histocompatibility complex (MHC)-mismatched murine
`cardiac allografts.14 These authors14 observed that CCR1
`⫺/⫺ KO mice did not reject class II-mismatched allografts,
`an effect that was augmented by cyclosporine (CsA) treat-
`ment. In other studies, sirolimus (SRL) was reported to
`decrease KC, MIP-2, and IL-8 production, as well as
`polymorphonuclear leukocyte infiltration into rat cardiac
`allografts.15
`Chronic rejection and emergence of transplant-associ-
`ated accelerated atherosclerosis have also been associated
`with persistence of RANTES,16 as well as increased MIP-1
`and MCP-1.17 The activity of the latter two chemokines
`suggests a critical role for intragraft macrophages, a hypoth-
`esis supported by the apparent benefit produced by gamma
`lactone, which inhibits macrophage effector activation.18
`
`FTY720: A PUTATIVE CHEMOKINE RECEPTOR
`AGONIST
`
`FTY720, 2-amino-2[2-(4-octylphenyl-ethyl]-1, 3-propane-
`diol, a synthetic analogue19 of myriocin, reversibly depletes
`peripheral blood lymphocytes (PBL) but not granulocytes
`or monocytes.
`Shinomiya et al20 suggested that FTY720 induces lym-
`phocyte apoptosis. They attributed this effect to activation
`of phospholipase C (PLC), mobilization of calcium, and
`up-regulation of caspase-1 and caspase-3. However, these
`observations were made using massive exposure to the
`agent and, therefore, may not be relevant to the immuno-
`suppressive effects observed at clinical doses of 1.0 to 5.0
`mg per day. At immunosuppressive doses, FTY720- and
`vehicle-treated transplant recipients showed similar fre-
`quencies of cytotoxic ACI anti-LEW (CTL) precursors in
`limiting-dilution analyses of PLN and SPL lymphocytes and
`equal staining for apoptotic cells in allografts and PLNs
`
`From the Division of Immunology and Organ Transplantation,
`University of Texas–Houston, Houston, Texas, USA.
`This work was supported by a grant from the National Institute
`of Diabetes and Digestive and Kidney Diseases (NIDDK 38016-
`14).
`Address reprint requests to Barry D. Kahan, PhD, MD, Division
`of Immunology and Organ Transplantation, University of Texas-
`Houston, 6431 Fannin, Suite 6.240, Houston, Texas, USA.
`
`0041-1345/01/$–see front matter
`PII S0041-1345(01)02313-2
`
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`KAHAN
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`using the TdT-mediated deoxyuridine 5⬘ triphosphate nick-
`end labeling (TUNEL) assay (data not shown).
`Alternatively, Chiba and colleagues21,22 suggest
`that
`FTY720 produces a reversible effect to sequester lympho-
`cytes to SLS. To confirm this hypothesis, we observed that
`purified lymphocytes labeled with 5-6-carboxyfluorescein
`diacetate succinimide ester (CF) upon adoptive transfer
`from FTY720-treated donors into syngeneic Balb/c hosts
`showed reduced numbers of CF䊝 cells (⫻103 ⫾ SD) in the
`blood (P ⬍ .03) and increased numbers in mesenteric lymph
`nodes (MLN), PP, and, to a lesser extent, PLN. Neither
`SPL nor thymus (TY) showed a significant increase in
`lymphocyte sequestration. Furthermore, transfers of puri-
`fied CD4⫹ or CD8⫹ T subsets as well as B cells showed
`similar migration patterns as the Pan-T cell population.23
`It has been observed that the lymphocyte sequestration
`effect of FTY720 is sensitive to Bordetella pertussis toxin and
`that the up-regulated chemotactic responses to SLC/ELC in
`vitro demand expression not only of the CCR7, but also of
`the Edg6 receptor. First, it was shown that FTY720 en-
`hanced in vitro chemotaxis. Second, the in vivo sequestra-
`tion effects produced by FTY720 were markedly reduced by
`administration of Bordetella pertussis toxin,24 an agent that
`inhibits Gi/Go ADP ribosylation of the G␣ subunit of
`heterotrimeric G-proteins. Five further observations sup-
`port the hypothesis that the lymphocyte sequestration effect
`produced by FTY720 depends upon CCR7-mediated adhe-
`sion: (1) abrogation by anti-CCR7 antibody; (2) modulated
`effect in plt-/plt- mice, which have a deletion of SLC and a
`70% to 80% reduction in ELC;25 (3) transfection of Edg6
`and CCR7 (but not one marker alone) conferred upon
`HEK293T cells the distinctive pattern of in vitro chemotac-
`tic responsiveness elicited by FTY720; (4) Clostridium toxin
`B (100 ng/mL), an agent that blocks the Rho family
`molecules, partially abrogated the chemotactic effect of
`double Edg6/CCR7 transfectants; and (5) FTY720 en-
`hanced actin polymerization by HEK239TEdg6 cells.
`
`EDG RECEPTOR SIGNALING PATHWAYS LEADING TO
`CCR7 UP-REGULATION
`
`Because FTY720 is a structural analogue of sphingosine, it
`seems reasonable to assume that it acts as a profound
`agonist for Edg6, a member of the family of receptors
`(Edg1, -3, -5, -6, and -8) that bind sphingosine-1-phosphate
`(S1P), in contradistinction to the Edg2, -4, and -7 family
`with receptors for lysophosphatidic acid (LPA, 1-acyl-
`glycerol-3-phosphate). Edg6 has limited distribution on
`lymphocytes and in lung tissue26 with the greatest degree of
`homology with Edg3, a cardiac receptor, possibly explaining
`why the only apparent clinical side effect of FTY720 is
`bradycardia. FTY720 may alter the balance of three en-
`zymes controlling S1P formation and degradation: (1) the
`sphingolipid kinase type I (SPHKI), a calcium-calmodulin-
`dependent cytosolic lipid enzyme that displays sphingosine
`substrate selectivity; (2) a specific cytosolic phosphatase
`“SHP”27 that rapidly degrades S1P, thereby dampening
`downstream events; and (3) another pathway of S1P deg-
`
`radation mediated by microsomal and/or lysosomal pyri-
`doxal phosphatase-dependent
`lyases, which cleave the
`C2-C3 bond, degrading the compound to free palmitalde-
`hyde and phosphoethanolamine.28
`
`IMMUNOSUPPRESSIVE ACTIVITY OF FTY720 IN
`TRANSPLANTATION
`Experimental Models
`
`FTY720 prolongs graft survival in a dose-dependent fash-
`ion and produces synergistic interactions with CsA and/or
`SRL. Administration of FTY720 prolongs the survival of rat
`liver and cardiac, as well as canine kidney, allografts.29
`Using the rigorous median effect model, we showed that
`FTY720 acts synergistically with CsA, with SRL, and with
`the CsA/SRL combination in rats30 and nonhuman pri-
`mates,31 findings that form the basis for ongoing clinical
`trials. In addition, a 3-day course of FTY720 combined with
`PV injection of modified class I MHC proteins induced
`donor-specific transplantation tolerance.32
`In a nonhuman primate model, cynomolgus monkeys
`transplanted with renal allografts were treated with concen-
`tration-controlled doses of CsA (intramuscular, selected to
`achieve target whole blood concentrations 40 to 200 ng/
`mL), and 0, 0.1, 0.3, or 1.0 mg/kg/d FTY720 (IV). Addition
`of 0.1 or 0.3 mg/kg/d FTY720 to the CsA regimen increased
`graft mean survival time (MST) from 8.5 to 71 or 63 days
`(P ⬍ .04 or P ⬍ .05, respectively). Histopathologic evalua-
`tion confirmed the therapeutic benefits of the agent; trans-
`plants in FTY720-treated hosts showed only minimal in-
`flammatory infiltrates upon protocol biopsies.31
`
`Clinical Development
`
`The preclinical results on lymphocyte homing were con-
`firmed by a multiple-dose Phase I clinical study reported by
`the applicant group.33 Ascending amounts of FTY720
`added to the CsA-Pred regimen of cohorts of stable renal
`transplant patients produced a prompt, dose-dependent,
`and reversible decrease in the number of circulating PBLs
`without effects on monocytes or neutrophils. Another mul-
`ticenter, randomized, open-label study evaluated the safety,
`tolerability, and efficacy of FTY720 versus mycophenolate
`mofetil (MMF) with CsA and steroids in de novo transplan-
`tation. Episodes of transient bradycardia without symptoms
`or sequelae, most of which occurred within the first 24
`hours posttransplant, were reported in 11/124 (8.9%) of
`FTY720-treated patients versus 2/35 (5.7%) of MMF-
`treated patients. Graft survival was 99% (one graft loss in
`the MMF group) and patient survival was 100%. FTY720
`was well tolerated and appeared to be effective in the
`prevention of acute rejection in combination with CsA and
`steroids.
`In addition, immunosuppressive regimens that do not
`include a calcineurin antagonist are of especial interest for
`patients at increased risk of delayed graft function (DGF).
`Early results in an ongoing study suggest the potential
`utility of an FTY720/RAD regimen in the setting of such
`
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`FTY720: A NEW DIMENSION IN TRANSPLANTATION
`
`3083
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`high-risk renal transplant patients. Entry criteria included a
`constellation of risk factors for DGF defined as the need for
`a dialysis treatment, namely, retransplantation status, Afri-
`can-American ethnicity, prolonged cold ischemia time, ad-
`vanced donor age, and cause of donor death due to
`cerebrovascular accident. The immunosuppressive regimen
`included FTY720 (loading dose 4 mg, maintenance dose 2.5
`mg/d), everolimus according to a concentration-controlled
`regimen (6 to 8 ng/mL trough), and tapering doses of
`steroids. Follow-up data are available for 11 to 93 days
`posttransplant (mean 96 days). As predicted, virtually every
`patient experienced DGF. During the follow-up period, six
`(22%) patients experienced an acute rejection episode;
`three (11%), graft loss; and one (3%), death.
`
`SUMMARY
`
`FTY720 is a myriocin derivative currently undergoing mul-
`ticenter trials for the prophylaxis of renal transplant rejec-
`tion. The drug is a novel immunomodulator demonstrating
`unique and potent pharmacologic activity by selectively
`altering lymphocyte homing from peripheral blood to SLS.
`Initial clinical trials indicate that FTY720 absorption and
`disposition is predictable and dose-linear, with minimal
`circadian variation. The time needed to reach steady state is
`long, suggesting the benefit of a loading dose. Doses up to
`5.0 mg/d for 28 days were well tolerated in stable renal
`transplant patients maintained on cyclosporine (CsA) and
`steroids, with no evidence of a pharmacokinetic interaction
`with CsA. A combination regimen (FTY720, CsA, and
`steroids) reduced the occurrence of acute rejection epi-
`sodes to less than 10% in a pilot study of de novo renal
`transplant patients. Preliminary data from another pilot
`study using the alternate combination (FTY720, RAD, and
`steroids) also shows a marked decrease in these episodes in
`high-risk patients. The unique mode of action, indepen-
`dence from cytochrome P450 3A4 metabolism, and sugges-
`tions of immunosuppressive potency together suggest that
`FTY720 will be a valuable addition to the immunosuppres-
`sive armamentarium.
`
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