The dual S1PR1/S1PR5 drug BAF312 (Siponimod) attenuates demyelination in organotypic slice cultures.

Catherine O’Sullivan,Kumlesh K. Dev,Anna Schubart,Anis K. Mir

doi:10.1186/s12974-016-0494-x

Catherine O’Sullivan, Kumlesh K. Dev + Show 2 more

Open Access

https://doi.org/10.1186/s12974-016-0494-x

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Abstract

BackgroundBAF312 (Siponimod) is a dual agonist at the sphingosine-1 phosphate receptors, S1PR1 and S1PR5. This drug is currently undergoing clinical trials for the treatment of secondary progressive multiple sclerosis (MS). Here, we investigated the effects of BAF312 on isolated astrocyte and microglia cultures as well as in slice culture models of demyelination.MethodsMouse and human astrocytes were treated with S1PR modulators and changes in the levels of pERK, pAkt, and calcium signalling as well as S1PR1 internalization and cytokine levels was investigated using Western blotting, immunochemistry, ELISA and confocal microscopy. Organotypic slice cultures were prepared from the cerebellum of 10-day-old mice and treated with lysophosphatidylcholine (LPC), psychosine and/or S1PR modulators, and changes in myelination states were measured by fluorescence of myelin basic protein and neurofilament H.ResultsBAF312 treatment of human and mouse astrocytes activated pERK, pAKT and Ca2+ signalling as well as inducing S1PR1 internalization. Notably, activation of S1PR1 increased pERK and pAKT in mouse astrocytes while both S1PR1 and S1PR3 equally increased pERK and pAKT in human astrocytes, suggesting that the coupling of S1PR1 and S1PR3 to pERK and pAKT differ in mouse and human astrocytes. We also observed that BAF312 moderately attenuated lipopolysaccharide (LPS)- or TNFα/IL17-induced levels of IL6 in both astrocyte and microglia cell cultures. In organotypic slice cultures, BAF312 reduced LPC-induced levels of IL6 and attenuated LPC-mediated demyelination. We have shown previously that the toxic lipid metabolite psychosine induces demyelination in organotypic slice cultures, without altering the levels of cytokines, such as IL6. Importantly, psychosine-induced demyelination was also attenuated by BAF312.ConclusionsOverall, this study suggests that BAF312 can modulate glial cell function and attenuate demyelination, highlighting this drug as a further potential therapy in demyelinating disorders, beyond MS.

Highlights

BAF312 (Siponimod) is a dual agonist at the sphingosine-1 phosphate receptors, S1PR1 and S1PR5
Activation of S1PR1/S1PR5 promotes pERK and pAKT signalling in mouse and human astrocytes The Ras/Raf/MEK/ERK and PI3K/PTEN/AKT signalling cascades are key signalling pathways involved in the regulation of cell survival and proliferation of numerous cell types
Cultured human and mouse astrocytes were serum starved for 4 h and treated with increasing concentrations of BAF312 (1 nM, 10 nM, 100 nM and 1 μM) for 10 min or 30 min and the samples prepared for Western blotting

Summary

Introduction

BAF312 (Siponimod) is a dual agonist at the sphingosine-1 phosphate receptors, S1PR1 and S1PR5. This drug is currently undergoing clinical trials for the treatment of secondary progressive multiple sclerosis (MS). The family of spingosine-1-phosphate receptors (S1PR) have been rapidly gaining attention as important mediators of many cellular processes, including cell differentiation, migration, survival, angiogenesis, calcium homeostasis, inflammation and immunity [1]. These receptors are Gprotein coupled and are known targets for the drug Gilenya® (pFTY720), an oral therapy used in patients with multiple sclerosis (MS) [2]. S1PR expression is cell-type dependent and these receptors are known to play a role in, for example, astrocyte migration, oligodendrocyte myelination state and neurite outgrowth and neurogenesis [5–9].

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