Sphingosine-1-phosphate Receptor Research Articles

Repurposing drugs against Alzheimer's disease: can the anti-multiple sclerosis drug fingolimod (FTY720) effectively tackle inflammation processes in AD?

Therapeutic approaches providing effective medication for Alzheimer's disease (AD) patients after disease onset are urgently needed. Previous studies in AD mouse models and in humans suggested that physical exercise or changed lifestyle can delay AD-related synaptic and memory dysfunctions when treatment started in juvenile animals or in elderly humans before onset of disease symptoms. However, a pharmacological treatment that can reverse memory deficits in AD patients was thus far not identified. Importantly, AD disease-related dysfunctions have increasingly been associated with neuro-inflammatory mechanisms and searching for anti-inflammatory medication to treat AD seems promising. Like for other diseases, repurposing of FDA-approved drugs for treatment of AD is an ideally suited strategy to reduce the time to bring such medication into clinical practice. Of note, the sphingosine-1-phosphate analogue fingolimod (FTY720) was FDA-approved in 2010 for treatment of multiple sclerosis patients. It binds to the five different isoforms of Sphingosine-1-phosphate receptors (S1PRs) that are widely distributed across human organs. Interestingly, recent studies in five different mouse models of AD suggest that FTY720 treatment, even when starting after onset of AD symptoms, can reverse synaptic deficits and memory dysfunction in these AD mouse models. Furthermore, a very recent multi-omics study identified mutations in the sphingosine/ceramide pathway as a risk factor for sporadic AD, suggesting S1PRs as promising drug target in AD patients. Therefore, progressing with FDA-approved S1PR modulators into human clinical trials might pave the way for these potential disease modifying anti-AD drugs.

Abstract 1585: Targeting acid ceramidase-1 to inhibit sphingolipid metabolism and tumor growth in pancreatic ductal carcinoma

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a leading cause of cancer-related deaths in the United States. PDAC is an aggressive disease with a poor survival rate of <10%. Hence, there is an urgent need to find novel drug targets for the clinical management of PDAC. Altered sphingolipid metabolism (SM) is a hallmark of cancer. The major sphingolipid metabolites are ceramide, sphingosine and sphingosine-1-phosphate (S1P). We found that S1P is the predominant SM metabolite in PDAC cells and tissues. The equilibrium between ceramide and S1P levels within the body is an important factor to determine cell fate. Ceramide induces apoptosis, while S1P promotes cancer growth. The IHC and ELISA assays showed that S1P levels are higher in PDAC cells and tissues compared to normal pancreatic tissue. We found that S1P induces spheroid formation in PDAC cells at nanomolar concentrations when compared to C1P and sphingosine. S1P binds to S1P receptors (SIPR1-5) to activate downstream signaling. Mechanistically, S1P treatment activates beta-catenin and notch signaling pathway proteins in PDAC cells. Our data suggest that S1P promotes tumor growth and stemness in PDAC and inhibition of S1P production can be a novel strategy for PDAC treatment. The cancer genome atlas (TCGA) data analysis indicated that genes involved in S1P production, especially acid ceramidase (ASAH1) and sphingosine kinase 1 (SPHK1) are upregulated in PDAC. ASAH1 converts ceramide to sphingosine which further gets converted to S1P by SPHK1. SPHK1 has been targeted in cancers, but kinase inhibitors are traditionally known for their non-selectivity and toxicity. Moreover, selective targeting of S1PR1-5 has been unsuccessful. Hence, there is a need to identify other targets to inhibit the production of S1P. Therefore, we decided to target Acid ceramidase 1 (ASAH1). Inhibiting ASAH1 will result in the accumulation of proapoptotic ceramide and inhibition of S1P production. Our data showed that ASAH1 is overexpressed in PDAC tissue and cells, while the ASAH1 knockdown in PDAC cells resulted in reduced proliferation, colony formation, migration and spheroid formation. Hence, targeting ASAH1 represents a novel strategy to inhibit PDAC progression by inhibiting sphingolipid metabolism and S1P production. Next, we developed a series of ASAH1 inhibitors based on Cerenib-2 (known ASAH1 inhibitor). One of the analogs, Cer-IVA inhibited the proliferation, colony and spheroid formation in PDAC cells at nanomolar concentration. Moreover, it also induced apoptosis and G2/M-cell cycle arrest in a dose and time-dependent manner. Further, it inhibited S1P production and increased ceramide production in PDAC cells. We are currently investigating the antitumor effects of Cer-IVA in an orthotopic model in C57BL/6 mice. In conclusion, targeting ASAH1 by Cer-IVA is an attractive strategy to inhibit S1P production, growth and stemness in PDAC cells. Citation Format: Krishan G. Jain, Hindole Ghosh, Sangita Bhattacharyya, Anup Kasi, Ameer Hamza, Scott J. Weir, Michael VanSaun, Stefan Bossmann, Shrikant Anant, Bernhard Biersack, Prasad R. Dandawate. Targeting acid ceramidase-1 to inhibit sphingolipid metabolism and tumor growth in pancreatic ductal carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1585.

2-Aminobenzoxazole Derivatives as Potent Inhibitors of the Sphingosine-1-Phosphate Transporter Spinster Homolog 2 (Spns2).

The S1P1 receptor is the target of four marketed drugs for the treatment of multiple sclerosis and ulcerative colitis. Targeting an S1P exporter, specifically Spns2, that is "upstream" of S1P receptor engagement is an alternate strategy that might recapitulate the efficacy of S1P receptor modulators without cardiac toxicity. We recently reported the first Spns2 inhibitor SLF1081851 (16d) that has modest potency with in vivo activity. To develop more potent compounds, we initiated a structure-activity relationship study that identified 2-aminobenzoxazole as a viable scaffold. Our studies revealed SLB1122168 (33p), which is a potent inhibitor (IC50 = 94 ± 6 nM) of Spns2-mediated S1P release. Administration of 33p to mice and rats resulted in a dose-dependent decrease in circulating lymphocytes, a pharmacodynamic indication of Spns2 inhibition. 33p provides a valuable tool compound to explore both the therapeutic potential of targeting Spns2 and the physiologic consequences of selective S1P export inhibition.

MP20-17 LARD DIET PROMOTES PROSTATE CANCER PROGRESSION THROUGH MODULATING GUT MICROBIOTA AND INTRATUMORAL LIPID METABOLISM

You have accessJournal of UrologyCME1 Apr 2023MP20-17 LARD DIET PROMOTES PROSTATE CANCER PROGRESSION THROUGH MODULATING GUT MICROBIOTA AND INTRATUMORAL LIPID METABOLISM Hiromi Sato, Shintaro Narita, Yoshiko Takahashi, Masanori Ishida, Mizuki Kobayashi, Soki Kasima, Ryohei Yamamoto, Taketoshi Nara, Mingguo Huang, Kazuyuki Numakura, Mitsuru Saito, Toshiaki Yoshioka, and Tomonori Habuchi Hiromi SatoHiromi Sato More articles by this author , Shintaro NaritaShintaro Narita More articles by this author , Yoshiko TakahashiYoshiko Takahashi More articles by this author , Masanori IshidaMasanori Ishida More articles by this author , Mizuki KobayashiMizuki Kobayashi More articles by this author , Soki KasimaSoki Kasima More articles by this author , Ryohei YamamotoRyohei Yamamoto More articles by this author , Taketoshi NaraTaketoshi Nara More articles by this author , Mingguo HuangMingguo Huang More articles by this author , Kazuyuki NumakuraKazuyuki Numakura More articles by this author , Mitsuru SaitoMitsuru Saito More articles by this author , Toshiaki YoshiokaToshiaki Yoshioka More articles by this author , and Tomonori HabuchiTomonori Habuchi More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003245.17AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Lard diet (LD) is a risk factor for prostate cancer (PCa) progression. The role of dietary dysbiosis in the development of PCa is largely unknown, even though it has recently become clear that it plays a significant role in progression of many cancers. Herein, we performed a comprehensive gut microbiome and intratumoral mRNA omics analyses to analyze targetable molecules involved in LD-induced PCa progression via gut microbial alteration. METHODS: Pb-Cre; Pten (fl/fl) transgenic mice and C57BL6 mice subcutaneously inoculated with TRAMP-C2 PCa cells were fed with a LD rich in saturated fatty acid (SFA) or fish- oil diet (FOD) rich in polyunsaturated fatty acid. We obtained fecal and prostate/subcutaneous tumor samples. Using 16S rRNA gene amplicon sequencing, we compared the diversity and relative abundance of gut microbiota. We performed a cDNA microarray to assess the mRNA expression in the mouse PCa tissue. RESULTS: More weight and prostate/subcutaneous tumor volume were gained by the LD-fed mice in two mouse models compared to the FOD-fed mice. In an integrated analysis of the two models, the LD group and the FOD group formed different gut microbial clusters. According to microbiome analysis, order Lactobacillales was relatively more abundant in the FOD group than in the LD group, and order Clostridiales was enriched in the LD group compared to the FOD group. The proportion of order Lactbacillales in the gut was negatively associated with the body weight and prostate weight of the mice. In microarray analysis, we found the modulation of lipid metabolism and cholesterol biosynthesis pathways with 10 significantly modulated genes including sphingosine 1-phosphate receptor 2 (S1PR2) and low-density lipoprotein receptor (LDLR) in PCa tissues. Of these genes, S1PR2 was highly correlated with the abundance of order Lactobacillales in the mouse gut. Moreover, PCa patients with a higher Gleason score (≥8) and a higher body mass (BMI) index had significantly lower S1PR2 RNA levels in human prostatic specimens than those with a Gleason score ≤ 7 and lower BMI. CONCLUSIONS: SFA may accelerate PCa progression by altering particular gut microbial species and lipogenic genes in PCa. Therapeutics with alteration of gut microbiota and candidate genes involved in diet-induced PCa progression may be attractive in PCa. Source of Funding: GlaxoSmithKline plc © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e282 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Hiromi Sato More articles by this author Shintaro Narita More articles by this author Yoshiko Takahashi More articles by this author Masanori Ishida More articles by this author Mizuki Kobayashi More articles by this author Soki Kasima More articles by this author Ryohei Yamamoto More articles by this author Taketoshi Nara More articles by this author Mingguo Huang More articles by this author Kazuyuki Numakura More articles by this author Mitsuru Saito More articles by this author Toshiaki Yoshioka More articles by this author Tomonori Habuchi More articles by this author Expand All Advertisement PDF downloadLoading ...

PHARMACOKINETIC AND PHARMACODYNAMIC ANALYSIS OF AMISELIMOD, A SELECTIVE SPHINGOSINE 1-PHOSPHATE RECEPTOR MODULATOR, IN HEALTHY SUBJECTS: RESULTS FROM A PHASE 1 STUDY

Amiselimod is a selective sphingosine 1-phosphate receptor modulator in development for inflammatory bowel disease. It is converted to its active metabolite, amiselimod phosphate (amiselimod-P), and has a long half-life and slow accumulation to steady state. We evaluated a multiple-dose titration regimen to determine the plasma pharmacokinetic (PK) profile of amiselimod and amiselimod-P at steady state for the 0.4 mg QD therapeutic dose on day 13 and for the 0.8 mg QD supratherapeutic dose on day 26. We also investigated the pharmacodynamic (PD) effect of amiselimod on absolute lymphocyte counts (ALCs).

SMAD genes are up-regulated in brain and blood samples of individuals with schizophrenia.

Schizophrenia is a severe psychiatric disorder, with heritability around 80%, but a not fully understood pathophysiology. Signal transduction through the mothers against decapentaplegic (SMADs) are eight different proteins involved in the regulation of inflammatory processes, cell cycle, and tissue patterning. The literature is not consistent regarding the differential expression of SMAD genes among subjects with schizophrenia. In this article, we performed a systematic meta-analysis of the expression of SMAD genes in 423 brain samples (211 schizophrenia vs. 212 healthy controls), integrating 10 datasets from two public repositories, following the PRISMA guidelines. We found a statistically significant up-regulation of SMAD1, SMAD4, SMAD5, and SMAD7, and a tendency for up-regulation of SMAD3 and SMAD9 in brain samples of patients with schizophrenia. Overall, six of the eight genes showed a tendency for up-regulation, and none of them was found to have a tendency for down-regulation. SMAD1 and SMAD4 were up-regulated also in blood samples of 13 individuals with schizophrenia versus eight healthy controls, suggesting the SMAD genes' potential role as biomarkers of schizophrenia. Furthermore, SMAD genes' expression levels were significantly correlated with those of Sphingosine-1-phosphate receptor-1 (S1PR1), which is known to regulate inflammatory processes. Our meta-analysis supports the involvement of SMAD genes in the pathophysiology of schizophrenia through their role in inflammatory processes, as well as demonstrates the importance of gene expression meta-analysis for improving our understanding of psychiatric diseases.

Fingolimod Administration Following Hypoxia Induced Neonatal Seizure Can Restore Impaired Long-term Potentiation and Memory Performance in Adult Rats

Neonatal seizures commonly caused by hypoxia can lead to long-term neurological outcomes. Early inflammation plays an important role in the pathology of these outcomes. Therefore, in the current study, we explored the long-term effects of Fingolimod (FTY720), an analog of sphingosine and potent sphingosine 1-phosphate (S1P) receptors modulator, as an anti-inflammatory and neuroprotective agent in attenuating anxiety, memory impairment, and possible alterations in gene expression of hippocampal inhibitory and excitatory receptors following hypoxia-induced neonatal seizure (HINS). Seizure was induced in 24 male and female pups (6 in each experimental group) at postnatal day 10 (P10) by premixed gas (5% oxygen/ 95% nitrogen) in a hypoxic chamber for 15 minutes. Sixty minutes after the onset of hypoxia, FTY720 (0.3 mg/kg) or saline (100 µl) was administered for 12 days (from P10 up to P21). Anxiety-like behavior and hippocampal memory function were assessed at P90 by elevated plus maze (EPM) and novel object recognition (NOR), respectively. Long-term potentiation (LTP) was recorded from hippocampal dentate gyrus region (DG) following stimulation of perforant pathway (PP). In addition, the hippocampal concentration of superoxide dismutase activity (SOD), malondialdehyde (MDA), and thiol as indices of oxidative stress were evaluated. Finally, the gene expression of NR2A subunit of N-Methyl-D-aspartic acid (NMDA) receptor, GluR2 subunit of (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) AMPA receptor and γ2 subunit of γ-Aminobutyric acid (GABAA) receptor were assessed at P90 by the quantitative real-time PCR. FTY720 significantly reduced later-life anxiety-like behavior, ameliorated object recognition memory and increased the amplitude and slope of the field excitatory postsynaptic potential (fEPSP) in the rats following HINS. These effects were associated with restoration of the hippocampal thiol content to the normal values and the regulatory role of FTY720 in the expression of hippocampal GABA and glutamate receptors subunits. In conclusion, FTY720 could restore the dysregulated gene expression of excitatory and inhibitory receptors. It also increased the reduced hippocampal thiol content, which was accompanied with attenuation of HINS-induced anxiety, reduced the impaired hippocampal related memory, and prevented hippocampal LTP deficits in later life following HINS.

S1PR2 is Important for Cigarette Smoke-induced Pyroptosis in Human Bronchial Epithelial Cells

Chronic obstructive pulmonary disease and other respiratory inflammatory diseases are often associated with cigarette smoke exposure. However, the underlying molecular mechanism remains unclear. This study aimed to investigate the role of sphingosine-1-phosphate receptor 2 (S1PR2) in cigarette smoke extract (CSE)-induced inflammation and pyroptosis in human bronchial epithelial (HBE) cells. CSE was administered to HBE cells and inflammation and pyroptosis were assessed. The mRNA levels of S1PR2, NLRP3, IL-1β, and IL-18 in HBE cells were detected by quantitative RT-PCR. Secreted protein levels of IL-1β and IL-18 in the culture supernatants were detected using enzyme-linked immunosorbent assay. Western blotting was used to measure the levels of S1PR2 and pyroptosis-related proteins (NLRP3, ASC, caspase-1, GSDMD, IL-1β, and IL-18). Our study revealed an upregulated expression of S1PR2, NLRP3, ASC, caspase-1, GSDMD, IL-1β, and regulated IL-18 in HBE cells after CSE exposure. Genetic blockage of S1PR2 could reverse the increased expression of these proteins related to CSE-induced pyroptosis. Conversely, S1PR2 overexpression increased CSE-induced pyroptosis by upregulating the expression of NLRP3, ASC, caspase-1, GSDMD, IL-1β, and IL-18 in HBE cells. Our results revealed that a novel S1PR2 signaling pathway may be involved in the pathogenesis of CSE-induced inflammation and pyroptosis in HBE cells. Thus, S1PR2 inhibitors could be an effective treatment for cigarette smoke-induced airway inflammation and injury.

Targeted Small Molecules for Systemic Lupus Erythematosus: Drugs in the Pipeline.

Despite the uncertainty of the pathogenesis of systemic lupus erythematosus, novel small molecules targeting specific intracellular mechanisms of immune cells are being developed to reverse the pathophysiological processes. These targeted molecules have the advantages of convenient administration, lower production costs, and the lack of immunogenicity. The Janus kinases, Bruton's tyrosine kinases, and spleen tyrosine kinases are important enzymes for activating downstream signals from various receptors on immune cells that include cytokines, growth factor, hormones, Fc, CD40, and B-cell receptors. Suppression of these kinases impairs cellular activation, differentiation, and survival, leading to diminished cytokine actions and autoantibody secretion. Intracellular protein degradation by immunoproteasomes, levered by the cereblon E3 ubiquitin ligase complex, is an essential process for the regulation of cellular functions and survival. Modulation of the immunoproteasomes and cereblon leads to depletion of long-lived plasma cells, reduced plasmablast differentiation, and production of autoantibodies and interferon-α. The sphingosine 1-phosphate/sphingosine 1-phosphate receptor-1 pathway is responsible for lymphocyte trafficking, regulatory T-cell/Th17 cell homeostasis, and vascular permeability. Sphingosine 1-phosphate receptor-1 modulators limit the trafficking of autoreactive lymphocytes across the blood-brain barrier, increase regulatory T-cell function, and decrease production of autoantibodies and type I interferons. This article summarizes the development of these targeted small molecules in the treatment of systemic lupus erythematosus, and the future prospect for precision medicine.

S1P1 Threonine 236 Phosphorylation Mediates the Invasiveness of Triple-Negative Breast Cancer and Sensitivity to FTY720.

Hyperactive sphingosine 1-phosphate (S1P) signaling is associated with a poor prognosis of triple-negative breast cancer (TNBC). Despite recent evidence that links the S1P receptor 1 (S1P1) to TNBC cell survival, its role in TNBC invasion and the underlying mechanisms remain elusive. Combining analyses of human TNBC cells with zebrafish xenografts, we found that phosphorylation of S1P receptor 1 (S1P1) at threonine 236 (T236) is critical for TNBC dissemination. Compared to luminal breast cancer cells, TNBC cells exhibit a significant increase of phospho-S1P1 T236 but not the total S1P1 levels. Misexpression of phosphorylation-defective S1P1 T236A (alanine) decreases TNBC cell migration in vitro and disease invasion in zebrafish xenografts. Pharmacologic disruption of S1P1 T236 phosphorylation, using either a pan-AKT inhibitor (MK2206) or an S1P1 functional antagonist (FTY720, an FDA-approved drug for treating multiple sclerosis), suppresses TNBC cell migration in vitro and tumor invasion in vivo. Finally, we show that human TNBC cells with AKT activation and elevated phospho-S1P1 T236 are sensitive to FTY720-induced cytotoxic effects. These findings indicate that the AKT-enhanced phosphorylation of S1P1 T236 mediates much of the TNBC invasiveness, providing a potential biomarker to select TNBC patients for the clinical application of FTY720.

Endothelial KCa3.1 and KCa2.3 Mediate S1P (Sphingosine-1-Phosphate)-Dependent Vasodilation and Blood Pressure Homeostasis.

S1P (sphingosine-1-phosphate) has been reported to possess vasodilatory properties, but the underlying pathways are largely unknown. Isolated mouse mesenteric artery and endothelial cell models were used to determine S1P-induced vasodilation, intracellular calcium, membrane potentials, and calcium-activated potassium channels (KCa2.3 and KCa3.1 [endothelial small- and intermediate-conductance calcium-activated potassium channels]). Effect of deletion of endothelial S1PR1 (type 1 S1P receptor) on vasodilation and blood pressure was evaluated. Mesenteric arteries subjected to acute S1P stimulation displayed a dose-dependent vasodilation response, which was attenuated by blocking endothelial KCa2.3 or KCa3.1 channels. In cultured human umbilical vein endothelial cells, S1P stimulated immediate membrane potential hyperpolarization following activation of KCa2.3/KCa3.1 with elevated cytosolic Ca2+. Further, chronic S1P stimulation enhanced expression of KCa2.3 and KCa3.1 in human umbilical vein endothelial cells in dose- and time-dependent manners, which was abolished by disrupting either S1PR1-Ca2+ signaling or downstream Ca2+-activated calcineurin/NFAT (nuclear factor of activated T-cells) signaling. By combination of bioinformatics-based binding site prediction and chromatin immunoprecipitation assay, we revealed in human umbilical vein endothelial cells that chronic activation of S1P/S1PR1 promoted NFATc2 nuclear translocation and binding to promoter regions of KCa2.3 and KCa3.1 genes thus to upregulate transcription of these channels. Deletion of endothelial S1PR1 reduced expression of KCa2.3 and KCa3.1 in mesenteric arteries and exacerbated hypertension in mice with angiotensin II infusion. This study provides evidence for the mechanistic role of KCa2.3/KCa3.1-activated endothelium-dependent hyperpolarization in vasodilation and blood pressure homeostasis in response to S1P. This mechanistic demonstration would facilitate the development of new therapies for cardiovascular diseases associated with hypertension.

Beneficial Effects of Hordenine on a Model of Ulcerative Colitis.

Hordenine, a phenethylamine alkaloid, is found in a variety of plants and exhibits a broad array of biological activities and pharmacological properties, including anti-inflammatory and anti-fibrotic effects. However, the efficacy and underlying mechanisms of hordenine in treating ulcerative colitis (UC) remain unclear. To address this, we examined the therapeutic effects of hordenine on dextran sodium sulphate (DSS)-induced UC by comparing disease activity index (DAI), colon length, secretion of inflammatory factors, and degree of colonic histological lesions across diseased mice that were and were not treated with hordenine. We found that hordenine significantly reduced DAI and levels of pro-inflammatory factors, including interleukin (IL)-6, IL-1β, and tumor necrosis factor alpha (TNF-α), and also alleviated colon tissue oedema, colonic lesions, inflammatory cells infiltration and decreased the number of goblet cells. Moreover, in vitro experiments showed that hordenine protected intestinal epithelial barrier function by increasing the expression of tight junction proteins including ZO-1 and occludin, while also promoting the healing of intestinal mucosa. Using immunohistochemistry and western blotting, we demonstrated that hordenine reduced the expression of sphingosine kinase 1 (SPHK1), sphingosine-1-phosphate receptor 1 (S1PR1), and ras-related C3 botulinum toxin substrate 1 (Rac1), and it inhibited the expression of phosphorylated signal transducer and activator of transcription 3 (p-STAT3) in colon tissues. Thus, hordenine appears to be effective in UC treatment owing to pharmacological mechanisms that favor mucosal healing and the inhibition of SPHK-1/S1PR1/STAT3 signaling.

Discovery of a Promising Fluorine-18 Positron Emission Tomography Radiotracer for Imaging Sphingosine-1-Phosphate Receptor 1 in the Brain.

Sphingosine-1-phosphate receptor 1 (S1PR1) is recognized as a novel therapeutic and diagnostic target in neurological disorders. We recently transferred the S1PR1 radioligand [11C]CS1P1 into clinical investigation for multiple sclerosis. Herein, we reported the design, synthesis and evaluation of novel F-18 S1PR1 radioligands. We combined the structural advantages of our two lead S1PR1 radioligands and synthesized 14 new S1PR1 compounds, then performed F-18 radiochemistry on the most promising compounds. Compound 6h is potent (IC50 = 8.7 nM) and selective for S1PR1. [18F]6h exhibited a high uptake in macaque brain (SUV > 3.0) and favorable brain washout pharmacokinetics in positron emission tomography (PET) study. PET blocking and displacement studies confirmed the specificity of [18F]6h in vivo. Radiometabolite analysis confirmed no radiometabolite of [18F]6h entered into the brain to confound the PET measurement. In summary, [18F]6h is a promising radioligand to image S1PR1 and worth translational clinical investigation for humans with brain disorders.

Fingolimod ameliorates chronic experimental autoimmune neuritis by modulating inflammatory cytokines and Akt/mTOR/NF-κB signaling.

Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is an immune-mediated disease that targets the myelin sheaths of the peripheral nerves. Fingolimod is a sphingosine 1 phosphate (S1P) receptor antagonist with a high affinity for S1P receptors through the Akt-mTOR pathway, and prior research has suggested that it might be helpful in autoimmune illnesses. Chronic experimental autoimmune neuritis (c-EAN) was induced by immunizing Lewis rats with the S-palm P0(180-199) peptide, and then the treatment group was intraperitoneally injected with fingolimod (1mg/kg) daily. Hematoxylin and eosin staining was used to assess the severity of nerve injury. Immunohistochemistry staining showed that fingolimod's anti-inflammatory effects on c-EAN rats might be realized through the NF-κB signaling pathway. Tumor necrosis factor-α (TNF-α), interferon-γ (INF-γ), interleukin-1beta (IL-1β), interleukin 6 (IL-6), inducible nitric oxide synthase (iNOS), and intercellular adhesion molecule-1 (ICAM-1) were measured to evaluate the inflammation levels, and pAkt, p-S6, and p-p65 were used to measure the abundance of downstream activation markers to determine whether the Akt/mTOR/NF-κB signaling pathway was activated in the c-EAN model. Fingolimod treatment reduced the inflammatory reaction and the expression of NF-κB in sciatic nerves. It also decreased the mRNA levels of the proinflammatory cytokines TNF-α, IFN-γ, IL-1β, IL-6, iNOS, and ICAM-1 and pAkt, p-S6, and p-p65, representing the Akt/mTOR/NF-κB signaling pathway. Our data showed that fingolimod could improve the disease course, alleviate the decrease in inflammation, and reduce proinflammatory cytokines through the Akt/mTOR/NF-κB axis in c-EAN rats, which could be beneficial for the development of CIDP-related research.

Pericytes protect rats and mice from sepsis-induced injuries by maintaining vascular reactivity and barrier function: implication of miRNAs and microvesicles

BackgroundVascular hyporeactivity and leakage are key pathophysiologic features that produce multi-organ damage upon sepsis. We hypothesized that pericytes, a group of pluripotent cells that maintain vascular integrity and tension, are protective against sepsis via regulating vascular reactivity and permeability.MethodsWe conducted a series of in vivo experiments using wild-type (WT), platelet-derived growth factor receptor beta (PDGFR-β)-Cre + mT/mG transgenic mice and Tie2-Cre + Cx43flox/flox mice to examine the relative contribution of pericytes in sepsis, either induced by cecal ligation and puncture (CLP) or lipopolysaccharide (LPS) challenge. In a separate set of experiments with Sprague–Dawley (SD) rats, pericytes were depleted using CP-673451, a selective PDGFR-β inhibitor, at a dosage of 40 mg/(kg·d) for 7 consecutive days. Cultured pericytes, vascular endothelial cells (VECs) and vascular smooth muscle cells (VSMCs) were used for mechanistic investigations. The effects of pericytes and pericyte-derived microvesicles (PCMVs) and candidate miRNAs on vascular reactivity and barrier function were also examined.ResultsCLP and LPS induced severe injury/loss of pericytes, vascular hyporeactivity and leakage (P < 0.05). Transplantation with exogenous pericytes protected vascular reactivity and barrier function via microvessel colonization (P < 0.05). Cx43 knockout in either pericytes or VECs reduced pericyte colonization in microvessels (P < 0.05). Additionally, PCMVs transferred miR-145 and miR-132 to VSMCs and VECs, respectively, exerting a protective effect on vascular reactivity and barrier function after sepsis (P < 0.05). miR-145 primarily improved the contractile response of VSMCs by activating the sphingosine kinase 2 (Sphk2)/sphingosine-1-phosphate receptor (S1PR)1/phosphorylation of myosin light chain 20 pathway, whereas miR-132 effectively improved the barrier function of VECs by activating the Sphk2/S1PR2/zonula occludens-1 and vascular endothelial-cadherin pathways.ConclusionsPericytes are protective against sepsis through regulating vascular reactivity and barrier function. Possible mechanisms include both direct colonization of microvasculature and secretion of PCMVs.

Emerging therapies for eosinophilic esophagitis.

Eosinophilic esophagitis (EoE) is an inflammatory condition of the esophagus that is quickly becoming a more recognized cause of esophageal dysfunction and dysphagia. The cause of EoE is thought to be due to an inflammatory response triggered by exposure to dietary or environmental antigens. Symptoms may be progressive and include reflux, nausea, vomiting, dysphagia, and reduction in quality of life. Both children and adults may be affected. Initial treatments for EoE have focused on elimination diets to potentially identify and remove dietary triggers, acid suppression with proton pump inhibitors, and topical delivery of orally administered swallowed corticosteroids. These therapies are often ineffective in a large proportion of patients, leading to exploration of other approaches by focusing on cytokines involved in the inflammatory response. Most recently, dupilumab, a monoclonal antibody targeting IL-4 received approval for use in patients aged 12 years or older with EoE. Use of dupilumab has seen favorable improvements in disease progression and symptoms with chronic use. Other therapies targeting IL-5, such as mepolizumab, reslizumab, and benralizumab are currently being studied for use in EoE. Therapies targeting other key inflammatory mediators in EoE, such as Siglec-8 (lirentelimab), IL-13 (cendakimab), and the sphingosine 1-phosphate receptor (etrasimod) are currently being evaluated in clinical trials.

Global sphingosine-1-phosphate receptor 2 deficiency attenuates neuroinflammation and ischemic-reperfusion injury after neonatal stroke

Arterial ischemic stroke is common in neonates-1 per 2,300-5,000 births-and therapeutic targets remain insufficiently defined. Sphingosine-1-phosphate receptor 2 (S1PR2), a major regulator of the CNS and immune systems, is injurious in adult stroke. Here, we assessed whether S1PR2 contributes to stroke induced by 3h transient middle cerebral artery occlusion (tMCAO) in S1PR2 heterozygous (HET), knockout (KO), and wild type (WT) postnatal day 9 pups. HET and WT of both sexes displayed functional deficits in Open Field test whereas injured KO at 24h reperfusion performed similarly to naives. S1PR2 deficiency protected neurons, attenuated infiltration of inflammatory monocytes, and altered vessel-microglia interactions without reducing increased cytokine levels in injured regions at 72 h. Pharmacologic inhibition of S1PR2 after tMCAO by JTE-013 attenuated injury 72h after tMCAO. Importantly, the lack of S1PR2 alleviated anxiety and brain atrophy during chronic injury. Altogether, we identify S1PR2 as a potential new target for mitigating neonatal stroke.

Etrasimod as induction and maintenance therapy for ulcerative colitis (ELEVATE): two randomised, double-blind, placebo-controlled, phase 3 studies

Etrasimod, a once-daily, oral, sphingosine 1-phosphate (S1P) receptor modulator that selectively activates S1P receptor subtypes 1, 4, and 5, with no detectable activity on S1P2,3, is in development for the treatment of immune-mediated diseases, including ulcerative colitis. In these two phase 3 trials, we aimed to evaluate the safety and efficacy of etrasimod in adult patients with moderately to severely active ulcerative colitis. In two independent randomised, multicentre, double-blind, placebo-controlled, phase 3 trials, ELEVATE UC 52 and ELEVATE UC 12, adults with active moderate-to-severe ulcerative colitis and an inadequate or loss of response or intolerance to at least one approved ulcerative colitis therapy were randomly assigned (2:1) to once-daily oral etrasimod 2 mg or placebo. Patients in ELEVATE UC 52 were enrolled from 315 centres in 40 countries. Patients in ELEVATE UC 12 were enrolled from 407 centres in 37 countries. Randomisation was stratified by previous exposure to biologicals or Janus kinase inhibitor therapy (yes vs no), baseline corticosteroid use (yes vs no), and baseline disease activity (modified Mayo score [MMS]; 4-6 vs 7-9). ELEVATE UC 52 comprised a 12-week induction period followed by a 40-week maintenance period with a treat-through design. ELEVATE UC 12 independently assessed induction at week 12. The primary efficacy endpoints were the proportion of patients with clinical remission at weeks 12 and 52 in ELEVATE UC 52 and week 12 in ELEVATE UC 12. Safety was evaluated in both trials. ELEVATE UC 52 and ELEVATE UC 12 were registered with ClinicalTrials.gov, NCT03945188 and NCT03996369, respectively. Patients in ELEVATE UC 52 were enrolled between June 13, 2019, and Jan 28, 2021. Patients in ELEVATE UC 12 were enrolled between Sept 15, 2020, and Aug 12, 2021. ELEVATE UC 52 and ELEVATE UC 12 screened 821 patients and 606 patients, respectively, with 433 and 354 subsequently undergoing random assignment. The full analysis set of ELEVATE UC 52 comprised 289 patients assigned to etrasimod and 144 to placebo. In ELEVATE UC 12, 238 patients were assigned to etrasimod and 116 to placebo. In ELEVATE UC 52, a significantly greater proportion of patients in the etrasimod group achieved clinical remission compared with patients in the placebo group at completion of the 12-week induction period (74 [27%] of 274 patients vs ten [7%] of 135 patients; p<0·0001) and at week 52 (88 [32%] of 274 patients vs nine [7%] of 135 patients; p<0·0001). In ELEVATE UC 12, 55 (25%) of 222 patients in the etrasimod group had clinical remission compared with 17 (15%) of 112 patients in the placebo group at the end of the 12-week induction period (p=0·026). Adverse events were reported in 206 (71%) of 289 patients in the etrasimod group and 81 (56%) of 144 patients in the placebo group in ELEVATE UC 52 and 112 (47%) of 238 patients in the etrasimod group and 54 (47%) of 116 patients in the placebo group in ELEVATE UC 12. No deaths or malignancies were reported. Etrasimod was effective and well tolerated as an induction and maintenance therapy in patients with moderately to severely active ulcerative colitis. Etrasimod is a treatment option with a unique combination of attributes that might address the persistent unmet needs of patients with ulcerative colitis. Arena Pharmaceuticals.

S1PR1 serves as a viable drug target against pulmonary fibrosis by increasing the integrity of the endothelial barrier of the lung.

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with unclear etiology and limited treatment options. The median survival time for IPF patients is approximately 2-3 years and there is no effective intervention to treat IPF other than lung transplantation. As important components of lung tissue, endothelial cells (ECs) are associated with pulmonary diseases. However, the role of endothelial dysfunction in pulmonary fibrosis (PF) is incompletely understood. Sphingosine-1-phosphate receptor 1 (S1PR1) is a G protein-coupled receptor highly expressed in lung ECs. Its expression is markedly reduced in patients with IPF. Herein, we generated an endothelial-conditional S1pr1 knockout mouse model which exhibited inflammation and fibrosis with or without bleomycin (BLM) challenge. Selective activation of S1PR1 with an S1PR1 agonist, IMMH002, exerted a potent therapeutic effect in mice with bleomycin-induced fibrosis by protecting the integrity of the endothelial barrier. These results suggest that S1PR1 might be a promising drug target for IPF therapy.

Necroptosis of macrophage is a key pathological feature in biliary atresia via GDCA/S1PR2/ZBP1/p-MLKL axis

Biliary atresia (BA) is a severe inflammatory and fibrosing neonatal cholangiopathy disease characterized by progressive obstruction of extrahepatic bile ducts, resulting in cholestasis and progressive hepatic failure. Cholestasis may play an important role in the inflammatory and fibrotic pathological processes, but its specific mechanism is still unclear. Necroptosis mediated by Z-DNA-binding protein 1 (ZBP1)/phosphorylated-mixed lineage kinase domain-like pseudokinase (p-MLKL) is a prominent pathogenic factor in inflammatory and fibrotic diseases, but its function in BA remains unclear. Here, we aim to determine the effect of macrophage necroptosis in the BA pathology, and to explore the specific molecular mechanism. We found that necroptosis existed in BA livers, which was occurred in liver macrophages. Furthermore, this process was mediated by ZBP1/p-MLKL, and the upregulated expression of ZBP1 in BA livers was correlated with liver fibrosis and prognosis. Similarly, in the bile duct ligation (BDL) induced mouse cholestatic liver injury model, macrophage necroptosis mediated by ZBP1/p-MLKL was also observed. In vitro, conjugated bile acid-glycodeoxycholate (GDCA) upregulated ZBP1 expression in mouse bone marrow-derived monocyte/macrophages (BMDMs) through sphingosine 1-phosphate receptor 2 (S1PR2), and the induction of ZBP1 was a prerequisite for the enhanced necroptosis. Finally, after selectively knocking down of macrophage S1pr2 in vivo, ZBP1/p-MLKL-mediated necroptosis was decreased, and further collagen deposition was markedly attenuated in BDL mice. Furthermore, macrophage Zbp1 or Mlkl specific knockdown also alleviated BDL-induced liver injury/fibrosis. In conclusion, GDCA/S1PR2/ZBP1/p-MLKL mediated macrophage necroptosis plays vital role in the pathogenesis of BA liver fibrosis, and targeting this process may represent a potential therapeutic strategy for BA.