TY - JOUR A1 - Stepanovska, Bisera A1 - Zivkovic, Aleksandra A1 - Enzmann, Gaby A1 - Tietz, Silvia A1 - Homann, Thomas A1 - Kleuser, Burkhard A1 - Engelhardt, Britta A1 - Stark, Holger A1 - Huwiler, Andrea T1 - Morpholino analogues of fingolimod as novel and selective S1P1 ligands with in vivo efficacy in a mouse model of experimental antigen-induced encephalomyelitis JF - International journal of molecular sciences N2 - Multiple sclerosis (MS) is a chronic, inflammatory, autoimmune disease of the central nervous system (CNS) which is associated with lower life expectancy and disability. The experimental antigen-induced encephalomyelitis (EAE) in mice is a useful animal model of MS, which allows exploring the etiopathogenetic mechanisms and testing novel potential therapeutic drugs. A new therapeutic paradigm for the treatment of MS was introduced in 2010 through the sphingosine 1-phosphate (S1P) analogue fingolimod (FTY720, Gilenya(R)), which acts as a functional S1P(1) antagonist on T lymphocytes to deplete these cells from the blood. In this study, we synthesized two novel structures, ST-1893 and ST-1894, which are derived from fingolimod and chemically feature a morpholine ring in the polar head group. These compounds showed a selective S1P(1) activation profile and a sustained S1P(1) internalization in cultures of S1P(1)-overexpressing Chinese hamster ovary (CHO)-K1 cells, consistent with a functional antagonism. In vivo, both compounds induced a profound lymphopenia in mice. Finally, these substances showed efficacy in the EAE model, where they reduced clinical symptoms of the disease, and, on the molecular level, they reduced the T-cell infiltration and several inflammatory mediators in the brain and spinal cord. In summary, these data suggest that S1P(1)-selective compounds may have an advantage over fingolimod and siponimod, not only in MS but also in other autoimmune diseases. KW - ST-1893 KW - ST-1894 KW - morpholino analogues of fingolimod KW - sphingosine KW - 1-phosphate KW - immunomodulator KW - lymphopenia KW - multiple sclerosis KW - experimental antigen-induced encephalomyelitis Y1 - 2020 U6 - https://doi.org/10.3390/ijms21186463 SN - 1422-0067 VL - 21 IS - 18 PB - MDPI CY - Basel ER - TY - JOUR A1 - Imeri, Faik A1 - Fallegger, Daniel A1 - Zivkovic, Aleksandra A1 - Schwalm, Stephanie A1 - Enzmann, Gaby A1 - Blankenbach, Kira A1 - Heringdorf, Dagmar Meyer Zu A1 - Homann, Thomas A1 - Kleuser, Burkhard A1 - Pfeilschifter, Josef A1 - Engelhardt, Britta A1 - Stark, Holger A1 - Huwiler, Andrea T1 - Novel oxazolo-oxazole derivatives of FTY720 reduce endothelial cell permeability, immune cell chemotaxis and symptoms of experimental autoimmune encephalomyelitis in mice JF - Neuropharmacology N2 - The immunomodulatory FTY720 (fingolimod) is presently approved for the treatment of relapsing-remitting multiple sclerosis. It is a prodrug that acts by modulating sphingosine 1-phosphate (S1P) receptor signaling. In this study, we have developed and characterized two novel oxazolo-oxazole derivatives of FTY720, ST-968 and the oxy analog ST-1071, which require no preceding activating phosphorylation, and proved to be active in intact cells and triggered S1P(1) and S1P(3), but not S1P(2), receptor internalization as a result of receptor activation. Functionally, ST-968 and ST-1071 acted similar to FTY720 to abrogate S1P-triggered chemotaxis of mouse splenocytes, mouse T cells and human U937 cells, and reduced TNFa- and LPS-stimulated endothelial cell permeability. The compounds also reduced TNF alpha-induced ICAM-1 and VCAM-1 mRNA expression, but restored TNF alpha-mediated downregulation of PECAM-1 mRNA expression. In an in vivo setting, the application of ST-968 or ST-1071 to mice resulted in a reduction of blood lymphocytes and significantly reduced the clinical symptoms of experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice comparable to FTY720 either by prophylactic or therapeutic treatment. In parallel to the reduced clinical symptoms, infiltration of immune cells in the brain was strongly reduced, and in isolated tissues of brain and spinal cord, the mRNA and protein expressions of ICAM-1 and VCAM-1, as well as of matrix metalloproteinase-9 were reduced by all compounds, whereas PECAM-1 and tissue inhibitor of metalloproteinase TIMP-1 were upregulated. In summary, the data suggest that these novel butterfly derivatives of FTY720 could have considerable implication for future therapies of multiple sclerosis and other autoimmune diseases. (C) 2014 Elsevier Ltd. All rights reserved. KW - Fingolimod KW - ST-968 KW - ST-1071 KW - Sphingosine 1-phosphate KW - Endothelial cells KW - Permeability KW - Multiple sclerosis Y1 - 2014 U6 - https://doi.org/10.1016/j.neuropharm.2014.05.012 SN - 0028-3908 SN - 1873-7064 VL - 85 SP - 314 EP - 327 PB - Elsevier CY - Oxford ER - TY - JOUR A1 - Pastukhov, Oleksandr A1 - Schwalm, Stephanie A1 - Zangemeister-Wittke, Uwe A1 - Fabbro, Doriano A1 - Bornancin, Frederic A1 - Japtok, Lukasz A1 - Kleuser, Burkhard A1 - Pfeilschifter, Josef A1 - Huwiler, Andrea T1 - The ceramide kinase inhibitor NVP-231 inhibits breast and lung cancer cell proliferation by inducing M phase arrest and subsequent cell death JF - British journal of pharmacology : journal of The British Pharmacological Society N2 - Background and PurposeCeramide kinase (CerK) catalyzes the generation of ceramide-1-phosphate which may regulate various cellular functions, including inflammatory reactions and cell growth. Here, we studied the effect of a recently developed CerK inhibitor, NVP-231, on cancer cell proliferation and viability and investigated the role of cell cycle regulators implicated in these responses. Experimental ApproachThe breast and lung cancer cell lines MCF-7 and NCI-H358 were treated with increasing concentrations of NVP-231 and DNA synthesis, colony formation and cell death were determined. Flow cytometry was performed to analyse cell cycle distribution of cells and Western blot analysis was used to detect changes in cell cycle regulator expression and activation. Key ResultsIn both cell lines, NVP-231 concentration-dependently reduced cell viability, DNA synthesis and colony formation. Moreover it induced apoptosis, as measured by increased DNA fragmentation and caspase-3 and caspase-9 cleavage. Cell cycle analysis revealed that NVP-231 decreased the number of cells in S phase and induced M phase arrest with an increased mitotic index, as determined by increased histone H3 phosphorylation. The effect on the cell cycle was even more pronounced when NVP-231 treatment was combined with staurosporine. Finally, overexpression of CerK protected, whereas down-regulation of CerK with siRNA sensitized, cells for staurosporine-induced apoptosis. Conclusions and ImplicationsOur data demonstrate for the first time a crucial role for CerK in the M phase control in cancer cells and suggest its targeted inhibition, using drugs such as NVP-231, in combination with conventional pro-apoptotic chemotherapy. Y1 - 2014 U6 - https://doi.org/10.1111/bph.12886 SN - 0007-1188 SN - 1476-5381 VL - 171 IS - 24 SP - 5829 EP - 5844 PB - Wiley-Blackwell CY - Hoboken ER - TY - JOUR A1 - Bhabak, Krishna P. A1 - Kleuser, Burkhard A1 - Huwiler, Andrea A1 - Arenz, Christoph T1 - Effective inhibition of acid and neutral ceramidases by novel B-13 and LCL-464 analogues JF - Bioorganic & medicinal chemistry : a Tetrahedron publication for the rapid dissemination of full original research papers and critical reviews on biomolecular chemistry, medicinal chemistry and related disciplines N2 - Induction of apoptosis mediated by the inhibition of ceramidases has been shown to enhance the efficacy of conventional chemotherapy in several cancer models. Among the inhibitors of ceramidases reported in the literature, B-13 is considered as a lead compound having good in vitro potency towards acid ceramidase. Furthermore, owing to the poor activity of B-13 on lysosoamal acid ceramidase in living cells, LCL-464 a modified derivative of B-13 containing a basic omega-amino group at the fatty acid was reported to have higher potency towards lysosomal acid ceramidase in living cells. In a search for more potent inhibitors of ceramidases, we have designed a series of compounds with structural modifications of B-13 and LCL-464. In this study, we show that the efficacy of B-13 in vitro as well as in intact cells can be enhanced by suitable modification of functional groups. Furthermore, a detailed SAR investigation on LCL-464 analogues revealed novel promising inhibitors of aCDase and nCDase. In cell culture studies using the breast cancer cell line MDA-MB-231, some of the newly developed compounds elevated endogenous ceramide levels and in parallel, also induced apoptotic cell death. In summary, this study shows that structural modification of the known ceramidase inhibitors B-13 and LCL-464 generates more potent ceramidase inhibitors that are active in intact cells and not only elevates the cellular ceramide levels, but also enhances cell death. KW - Sphingolipids KW - Ceramide KW - Ceramidase inhibitors KW - Structure-activity-relationship Y1 - 2013 U6 - https://doi.org/10.1016/j.bmc.2012.12.014 SN - 0968-0896 VL - 21 IS - 4 SP - 874 EP - 882 PB - Elsevier CY - Oxford ER -