G-protein-coupled receptors (GPCR) are the largest superfamily of receptors responsible for signaling between cells and tissues, and because they play important physiological roles in homeostasis, they are major drug targets. New technologies have been developed for the identification of new ligands, new GPCR functions, and for drug discovery purposes. In particular, intercellular lipid mediators, such as, lysophosphatidic acid and sphingosine 1-phosphate have attracted much attention for drug discovery and this has resulted in the development of fingolimod (FTY-720) and AM095. The discovery of new intercellular lipid mediators and their GPCRs are discussed from the perspective of drug development. Lipid GPCRs for lysophospholipids, including lysophosphatidylserine, lysophosphatidylinositol, lysophosphatidylcholine, free fatty acids, fatty acid derivatives, and other lipid mediators are reviewed.
Drug Discovery* ; Fatty Acids, Nonesterified ; Homeostasis ; Ligands ; Lysophosphatidylcholines ; Lysophospholipids ; Receptors, G-Protein-Coupled ; Sphingosine ; Fingolimod Hydrochloride

Here we report a case of pediatric multiple sclerosis treated with interferon beta-1b. Interferon beta is widely used in adult patients with multiple sclerosis (MS). However, its effects and safety in pediatric patients have not been well established. Although supporting data are limited, the use of disease modifying therapies (DMTs) such as interferon beta-1b is recommended early in treatment of children with MS. Reports of interferon beta treatment in pediatric MS patients in Korea are rare. In this report, we describe a Korean girl who was effectively treated with interferon beta-1b for three years. There were no relapses or serious side effects. Therefore, this report provides evidence supporting the use of interferon beta in pediatric MS patients in Korea and other Asian countries. We also reviewed current medical treatment of MS, including some DMTs and second-line treatment options such as natalizumab and cyclophosphamide, and several new oral agents such as fingolimod.
Adult ; Antibodies, Monoclonal, Humanized ; Asian Continental Ancestry Group ; Child ; Cyclophosphamide ; Humans ; Interferon-beta ; Interferons ; Korea ; Multiple Sclerosis ; Propylene Glycols ; Recurrence ; Sphingosine ; Fingolimod Hydrochloride ; Interferon beta-1b ; Natalizumab

Ischemia-reperfusion injury is complicated with multiple injury pathways. If a particular agent is used to restore blood flow and prevent cell death, many damaged neurons may come back to life. However, for stroke victims, there are no effective curative therapeutic approaches available other than thrombolytic treatment. The efficacy of neuroprotective agents are limited by low diffusion, narrow time window, strict dose titration, and lack of confidence at the preclinical level. NXY-059 reflects the fundamental limitation of neuroprotectant. There are recent attempts to overcome these limitations, with use of annexin A2, fingolimod, hydrogen, nitrite, and more. By covering two components, this report reviews what we have recently learned. In addition, it sheds light on some of the newer issues in clinical application.
Annexin A2 ; Benzenesulfonates ; Cell Death ; Diffusion ; Hydrogen ; Light ; Multiple Trauma ; Neurons ; Neuroprotective Agents ; Propylene Glycols ; Reperfusion Injury ; Sphingosine ; Stroke ; Fingolimod Hydrochloride

OBJECTIVETo investigate the effects of FTY720, a new immunosuppressive agent, on apoptosis and autophagy in multiple myeloma(MM) cell line U266 and to clarify its molecular mechanism.

METHODSU266 cells were treated with 0, 2.5, 5.0, 10.0 and 20.0 µmol/L FTY720 for 24 hours, and the cell viability was assayed by CCK-8 method. Then U266 cells were treated with 20.0 µmol/L FTY720 for 0, 2, 6 and 24 hours, the cell viability was tested. The apoptotic rates induced by different doses and time points of FTY720 were tested by flow cytometry separately. The expression of LC3B was detected by Western blot after U266 cells treated with different doses of FTY720 to see autophagy. U266 cells were treated with FTY720 ± Bafilomycin A1, an inhibitor of autophagy, for 24 hours, then the cell viability and apoptotic rates were tested. Meanwhile the expression of survivin, anti-apoptotic factors, were tested by Western blot.

RESULTSThe cell viability and the apoptotic rates were inhibited significantly by FTY720 (P < 0.05) in time-dependent and dose-dependent manner. The expression of LC3B-II increased significantly in a dose-dependent manner, it indicated that the autophagy was induced by FTY720. Bafilomycin A1 could rescue the cell viability and apoptotic rates in U266 cells treated with FTY720, and it could also rescue the expression of survivin decreased by FTY720.

CONCLUSIONSFTY720 can cause apoptosis and autophagy of U266 cells. The autophagy promote the apoptosis, which maybe due to the degradation of anti-apoptotic factors such as survivin or their upstream factors in lysosomes through autophagy.

Apoptosis ; drug effects ; Autophagy ; drug effects ; Cell Line, Tumor ; Fingolimod Hydrochloride ; Humans ; Multiple Myeloma ; pathology ; Propylene Glycols ; pharmacology ; Sphingosine ; analogs & derivatives ; pharmacology

Sphingosine 1-phosphate (S1P) is an important biologically active lysophospholipid that transmits signals through a family of G-protein-coupled receptors (GPCRs) to regulate the vital functions of several types of immune cells. The S1P GPCRs suppress both generation of specialized functional cytokines, such as IFN-gamma and IL-4, and proliferation of T-cells. Although S1P is chemotactic to T cells, B cells, dendritic cells, and natural killer cells, the major effect of S1P on the immune system is the regulation of lymphocyte recirculation and tissue distribution by S1P and S1P1. Chemotactic response of CD4+CD25+ regulatory T cells to S1P is reduced, but its optimal suppressive activities require S1P. FTY720, a new class of immunomodulator, is rapidly phosphorylated by sphingosine kinase 2 in vivo to form the biologically active phosphorylated-FTY720 (FTY720-P), which closely resembles S1P. The FTY720-P is a true agonist for S1P1, S1P3, S1P4, and S1P5, it affects the tissue distribution and functional activity of T cells, B cells, dendritic cells and regulatory T cells. FTY720 were demonstrated to be a hypotoxic, great effective and reversible immunosuppressive efficacy to prevent allograft rejection and treat some autoimmune diseases. In this article, the synthesis and metabolism of S1P, the expression of S1P GPCRs in immune cells, the effect of S1P on immune cells, the drugs targeted to S1P GPCRs and their clinical implications are reviewed.
Fingolimod Hydrochloride ; Humans ; Immunomodulation ; physiology ; Lysophospholipids ; immunology ; physiology ; Propylene Glycols ; metabolism ; Receptors, G-Protein-Coupled ; immunology ; physiology ; Sphingosine ; analogs & derivatives ; immunology ; metabolism ; physiology

Initial discovery on sphingosine 1-phosphate (S1P) as an intracellular second messenger was faced unexpectedly with roles of S1P as a first messenger, which subsequently resulted in cloning of its G protein-coupled receptors, S1P₁₋₅. The molecular identification of S1P receptors opened up a new avenue for pathophysiological research on this lipid mediator. Cellular and molecular in vitro studies and in vivo studies on gene deficient mice have elucidated cellular signaling pathways and the pathophysiological meanings of S1P receptors. Another unexpected finding that fingolimod (FTY720) modulates S1P receptors accelerated drug discovery in this field. Fingolimod was approved as a first-in-class, orally active drug for relapsing multiple sclerosis in 2010, and its applications in other disease conditions are currently under clinical trials. In addition, more selective S1P receptor modulators with better pharmacokinetic profiles and fewer side effects are under development. Some of them are being clinically tested in the contexts of multiple sclerosis and other autoimmune and inflammatory disorders, such as, psoriasis, Crohn’s disease, ulcerative colitis, polymyositis, dermatomyositis, liver failure, renal failure, acute stroke, and transplant rejection. In this review, the authors discuss the state of the art regarding the status of drug discovery efforts targeting S1P receptors and place emphasis on potential clinical applications.
Acute Kidney Injury ; Animals ; Clone Cells ; Cloning, Organism ; Colitis, Ulcerative ; Dermatomyositis ; Drug Discovery* ; Fingolimod Hydrochloride ; Graft Rejection ; In Vitro Techniques ; Liver Failure ; Mice ; Multiple Sclerosis ; Polymyositis ; Psoriasis ; Receptors, Lysosphingolipid* ; Second Messenger Systems ; Sphingosine* ; Stroke

This study was purpose to investigate the role of reactive oxygen species (ROS) in apoptosis and autophagy induced by FTY720 in multiple myeloma cell line U266. U266 cells were treated by different concentrations of FTY720 for 24 h, the apoptotic rates were detected by flow cytometry, and the expression of LC3B was detected by Western blot. The results indicated that apoptosis and autophagy were induced by FTY720 in U266 cells. Autophagy induced by FTY720 could lead to cell death. Bafilomycin A1, the inhibitor of autophagy, could enhance the cell viability in U266 cells treated with FTY720. NAC or Tiron, ROS scavenger, could decrease the FTY720 induced apoptosis and the expression of LC3B-II was reduced in combination of FTY720 with NAC or Tiron as compared with treatment with FTY720 only. It is concluded that FTY720 can induce U266 cell apoptosis and autophagy. ROS is the mediator that regulates both the apoptosis and autophagy in multiple myeloma cells.
1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt ; Apoptosis ; drug effects ; Autophagy ; drug effects ; Cell Line, Tumor ; Fingolimod Hydrochloride ; Humans ; Macrolides ; Microtubule-Associated Proteins ; metabolism ; Multiple Myeloma ; metabolism ; pathology ; Propylene Glycols ; pharmacology ; Reactive Oxygen Species ; metabolism ; Sphingosine ; analogs & derivatives ; pharmacology

Animals ; Apoptosis ; drug effects ; Capillaries ; pathology ; Fibrosis ; Fingolimod Hydrochloride ; Kidney Diseases ; metabolism ; pathology ; Kidney Tubules ; blood supply ; Male ; Propylene Glycols ; pharmacology ; therapeutic use ; Rats ; Rats, Sprague-Dawley ; Sphingosine ; analogs & derivatives ; pharmacology ; therapeutic use ; Vascular Diseases ; prevention & control

OBJECTIVETo determine the role of sphingosine 1-phosphate receptor (S1PRs ) signaling in CD34+ hematopoietic stem/progenitor cell transmigration.

METHODSCD34(+) cells were separated by Ficoll density gradient centrifugation and incubated in DMEM medium with 10% fetal calf serum. The cells were pretreated by FTY720, with or without pertussis toxin (PTX) and antiCXCR4 mAb in the medium, followed by addition of 100 ng/ml SDF-1 into the lower chamber of a Costar 24-well transwell. The migrated cells were counted using FACS and the migrating rates were determined. The expressions of sphingosine 1-phosphate receptors were analyzed in CD34(+) cells before and after the transmigration by reverse transcriptase- polymerase chain reaction (RT-PCR). Cord blood CD34(+) cells were treated with or without FTY720 (10(+) mol/L), and the expressions of CD49d (VLA-4), CD11a (LFA-1), and CD62L (L-selectin) were analyzed at 1, 8, and 16 h after the treatment.

RESULTSWhile FTY720 did not affect spontaneous migration, a substantial increase of SDF-1-induced transmigration was observed in the presence of FTY720 (15.26 2.14 to 28.64 2.37). The FTY720-enhanced transmigration was completely blocked by addition of PTX or antiCXCR4 mAb. S1p1-5 was expressed in fresh isolated cord blood CD34(+) cells. The migrating cells stimulated by FTY720 and SDF-1 only expressed S1P1, S1P3, and S1P4. The expressions of CD49d, CD11a and CD62L on CD34(+) cells treated with FTY720 remained unchanged at the selected time points as compared with the control.

CONCLUSIONSS1PRs are involved the transmigration of CD34(+) cells. The activation of S1PRs results in increased chemotactic response of CD34(+) to SDF-1. These effects are mediated through CXCR4 and PTX-sensitive Gi proteins. Only the CD34(+) cells expressing the specific receptors can rapidly transmigrate. The activation of the S1PRs does not affect the expressions of the adhesion molecules on cord blood CD34(+) cells.

Antigens, CD34 ; metabolism ; Cell Movement ; Cells, Cultured ; Chemokine CXCL12 ; pharmacology ; Fetal Blood ; cytology ; Fingolimod Hydrochloride ; Hematopoietic Stem Cell Mobilization ; Hematopoietic Stem Cells ; cytology ; drug effects ; Humans ; Propylene Glycols ; pharmacology ; Receptors, Lysosphingolipid ; metabolism ; physiology ; Signal Transduction ; Sphingosine ; analogs & derivatives ; pharmacology

Syl948 is a synthesized selective S1P1 agonist with novel structure. HTRF-IP1 test indicated that Syl948-P, the active form of Syl948 in vitro, has strong activity against S1P1 (EC50: 83 +/- 16 nmol x L(-1)), but its effect on S1P3 was very weak (EC50: 1 026 +/- 90 nmol x L(-1)). In SD rats, oral administration of Syl948 10 mg x kg(-1) significantly decreased the peripheral blood lymphocytes (PBL), with the maximal PBL inhibition rate of 63%, which was as similar as equal dose of fingolimod (FTY720). Oral administration of Syl948 10 mg x kg(-1) had no effect on heart rate of SD rats, which was better than FTY720. Daily oral administration with Syl948 (2 or 4 mg x kg(-1)) significantly prolonged the survival time of the allografts of skin slice on mice. In summary, the above results demonstrated that Syl948 has great selectivity in vitro and good activity in vivo, which indicated its potential use as an anti-rejection drug in skin transplantation.
Animals ; Fingolimod Hydrochloride ; Graft Survival ; drug effects ; Immunosuppressive Agents ; pharmacology ; Lymphocytes ; drug effects ; Mice ; Propylene Glycols ; pharmacology ; Rats ; Receptors, Lysosphingolipid ; agonists ; Skin Transplantation ; Sphingosine ; analogs & derivatives ; pharmacology ; Transplantation, Homologous