A Novel Immunomodulator, FTY720, Prevents Development of Experimental Autoimmune Myasthenia Gravis in C57BL/6 Mice

Abstract

It has been well established that myasthenia gravis (MG) is caused by autoimmune responses against nicotinic acetylcholine receptor (AChR) on postsynaptic membrane in neuromuscular junctions.1) Experimental autoimmune myasthenia gravis (EAMG) has been used as an animal model for human MG. This model is prepared by immunizing mice or rats with AChR from Torpedo californica; the animals develop MG-like symptoms, including muscle weakness, with production of autoantibody against AChR.2)The novel immunomodulator (immunosuppressant) FTY720, which was discovered by Fujita et al. 3,4) of our group, is a synthetic structural analogue of myriocin (ISP-I), a metabolite of Isaria cinclairii.5) The efficacy of FTY720 has been well established in preclinical transplantation models, 6) and also recently in renal transplantation in humans. [7][8][9][10] Although the mechanisms of pharmacological action of FTY720 remain to be fully clarified, 11,12) it has been proposed that FTY720 is phosphorylated by sphingosine kinase, and the product, FTY720 monophosphate, suppresses immune response by sequestering lymphocytes from blood and peripheral tissues to the secondary lymphoid tissues.13-16) FTY720 has no inhibitory effect on cytokine production in vitro, in contrast to established immunosuppressants (cyclosporin and tacrolimus hydrate).16) Its mechanism of action is unique and differs from that of the established immunosuppressants. The development of FTY720 was described in our previous report. 17)In this study, we examined the efficacy of FTY720 for preventing the development of EAMG, an animal model of human MG. MATERIALS AND METHODSAnimals Specific pathogen-free (SPF) C57BL/6 mice (7 weeks of age, females) were purchased from Japan SLC Inc., Shizuoka, Japan. The mice were given g-ray-irradiated food (CRF-1, Oriental Bio Co., Kyoto, Japan) and distilled water for injection (Otsuka Pharmaceutical Co., Ltd., Tokyo, Japan).FTY720 2-Amino-2-[2-(4-octylphenyl)ethyl]propane-1,3-diol hydrochloride (FTY720) was kindly provided by Yoshitomi Pharmaceutical Industries, Ltd., Japan.Acetylcholine Receptor from Torpedo californica Electric organ of Torpedo californica was purchased from Aquatic Research Consultants (San Pedro, CA, U.S.A.), and acetylcholine receptor (tAChR) was purified by the method of Froehner and Rafto using cobrotoxin-coupled Sepharose.18) AChR concentration was quantified by the Lowry method.Evaluation of Clinical Symptoms of EAMG Muscle strength was evaluated by the inverted hanging technique as described by Karachunski et al. 19,20) Mice with a holding time of 10 min or more were considered normal, and those with a holding time of less than 10 min were considered as having EAMG.Study Protocol Twelve C57BL/6 mice were immunized intracutaneously on the back with tAChR (5 mg) in the presence of Freund's complete adjuvant at 8, 10 and 11 weeks of age. Six mice (treated group, Nos. 1-6) out of the twelve were orally given FTY720 in water (1.0 mg/kg) three times a week from the day before the first immunization. The other...

FTY720, a Synthetic Sphingosine 1 Phosphate Analogue, Inhibits Development of Atherosclerosis in Low-Density Lipoprotein Receptor–Deficient Mice

“…[21][22][23] Consistent with these pharmacokinetic properties, twice-or thrice-weekly administration of FTY720 is sufficient to suppress inflammatory processes in various mouse models. 24,25 In the present study, we found that repeated intraperitoneal injection of the compound (20 g/LDL-R Ϫ/Ϫ mouse, thrice weekly) produces FTY720 plasma concentrations of 67.5Ϯ8.7 ng/mL, which is within the known therapeutic concentration range. 23 Examination of FTY720 distribution among lipoprotein fractions isolated from LDL-R Ϫ/Ϫ mouse plasma spiked with 50 ng/mL of the compound revealed that 32%, 24%, and 44% were present in very-low-density lipoprotien, LDL, and HDL, respectively.…”

Relapse of Experimental Autoimmune Encephalomyelitis after Discontinuation of FTY720 (Fingolimod) Treatment, but Not after Combination of FTY720 and Pathogenic Autoantigen

“…4) Also, FTY720 has been reported to be effective in preventing the development of several immunological disease models, including rheumatoid arthritis, 5) myasthenia gravis, 6) and atopic dermatitis. 7) The mechanism of action of FTY720 differs from that of established immunosuppressants, such as tacrolimus hydrate and cyclosporine.…”

Therapy of autoimmune diseases by novel immunosuppressant FTY720

“…FTY suppressed the OVA-induced Th1-mediated airway inflammation characterized by increased numbers of lymphocytes and neutrophils in bronchoalveolar lavage fluid 35) . The efficacy of FTY in autoimmune diabetes 36) , autoimmune myocarditis 37) , and myasthenia gravis 38) have also been reported. …”