An Update on Potential Antidepressants Derived from Marine Natural Products.

INTRODUCTION: Depression is one of the most frequently occurring psychiatric disorders worldwide, affecting 121 million worldwide. World Health Organization (WHO) estimates that it is the leading cause of disability and the fourth leading contributor to the "global burden of diseases". OBJECTIVE: Investigating and developing a drug with a novel benefit-risk profile is critical. Marine sources have been explored for their benefits as an alternative therapy for depression treatment. Numerous studies have shown that natural compounds containing peptides, alkaloids, polyphenols, diterpenes, glycosides, vitamins, and minerals from marine sources can potentially treat a wide range of disorders, including depression. Such phytoconstituents are known to reduce oxidative stress and neuroinflammation, regulate the synthesis or function of neurotransmitters such as glutamate and acetylcholinesterase, and aid in enhancing serotonin levels and nerve development. METHODS: In this review study, a literature search was conducted using terms often used, including animal models of depression and their precise phases, marine sources, algae, sponges, and indole alkaloids. Additionally, databases were examined, including Scopus, Wiley, Elsevier, Google Scholar, and Web of Science. The Snowball technique was used to identify several articles about depression but correlated to marine sources in addition to database searches. RESULTS: Current antidepressant medications have several negative side effects on the human body, including dry mouth, cardiovascular interference, gastrointestinal symptoms, genitourinary symptoms, hepatotoxicity, convulsions, and obesity. As a result, researchers can identify a wide range of potential targets for medications derived from marine sources. A combination of marinederived drugs and available treatments can be estimated to minimize the negative effects. So that these resources can be used as efficiently as possible, and various marine-derived substances can be studied for therapeutic efficacy. CONCLUSION: This review focuses on the preclinical and clinical findings of marine-derived compounds with antidepressant properties that alter behavioural parameters and biochemical abnormalities, as well as their mechanism of action and in-vivo potential.

Defective Induction of IL-27-Mediated Immunoregulation by Myeloid DCs in Multiple Sclerosis.

The purpose of this study was to examine whether myeloid dendritic cells (mDCs) from patients with multiple sclerosis (MS) and healthy controls (HCs) become similarly tolerogenic when exposed to IL-27 as this may represent a potential mechanism of autoimmune dysregulation. Our study focused on natural mDCs that were isolated from HCs and MS patient peripheral blood mononuclear cells (PBMCs). After a 24-h treatment with IL-27 ± lipopolysaccharide (LPS), the mDCs were either harvested to identify IL-27-regulated gene expression or co-cultured with naive T-cells to measure how the treated DC affected T-cell proliferation and cytokine secretion. mDCs isolated from HCs but not untreated MS patients became functionally tolerogenic after IL-27 treatment. Although IL-27 induced both HC and untreated MS mDCs to produce similar amounts of IL-10, the tolerogenic HC mDCs expressed PD-L2, IDO1, and SOCS1, while the non-tolerogenic untreated MS mDCs expressed IDO1 and IL-6R. Cytokine and RNA analyses identified two signature blocks: the first identified genes associated with mDC tolerizing responses to IL-27, while the second was associated with the presence of MS. In contrast to mDCs from untreated MS patients, mDCs from HCs and IFNb-treated MS patients became tolerogenic in response to IL-27. The genes differentially expressed in the different donor IL-27-treated mDCs may contain targets that regulate mDC tolerogenic responses.

Biological activities of meroterpenoids isolated from different sources.

Meroterpenoids are natural products synthesized by unicellular organisms such as bacteria and multicellular organisms such as fungi, plants, and animals, including those of marine origin. Structurally, these compounds exhibit a wide diversity depending upon the origin and the biosynthetic pathway they emerge from. This diversity in structural features imparts a wide spectrum of biological activity to meroterpenoids. Based on the biosynthetic pathway of origin, these compounds are either polyketide-terpenoids or non-polyketide terpenoids. The recent surge of interest in meroterpenoids has led to a systematic screening of these compounds for many biological actions. Different meroterpenoids have been recorded for a broad range of operations, such as anti-cholinesterase, COX-2 inhibitory, anti-leishmanial, anti-diabetic, anti-oxidative, anti-inflammatory, anti-neoplastic, anti-bacterial, antimalarial, anti-viral, anti-obesity, and insecticidal activity. Meroterpenoids also possess inhibitory activity against the expression of nitric oxide, TNF- α, and other inflammatory mediators. These compounds also show renal protective, cardioprotective, and neuroprotective activities. The present review includes literature from 1999 to date and discusses 590 biologically active meroterpenoids, of which 231 are from fungal sources, 212 are from various species of plants, and 147 are from marine sources such as algae and sponges.

Therapeutic Peptides: Unravelling Conformational Dynamics by Systematic Application of Biophysical Techniques.

Peptide therapeutics represents one of the fastest-growing sectors in the pharmaceutical drugs pipeline, with an increasing number of regulatory approvals every year. Their pharmacological diversity, biocompatibility, high degree of potency and selectivity make them an attractive choice in several therapeutic areas, such as diabetes, cancer, immune, metabolic, cardiovascular and infectious diseases. However, the development of peptides as drugs presents its own set of challenges, necessitating extensive property optimization aimed at improving their drug-like properties and stability in biological environments. The discovery and development of innovative peptide therapeutic platforms entail the employment of several biophysical techniques, which monitor the structural as well as the functional integrity of peptides. Small structural changes of the bioactive peptides in response to the presence of various excipients can have a major impact on their pharmaceutical prowess, necessitating the use of analytical techniques for efficient quality control during development. Here we present some widely used methods, such as circular dichroism, fluorescence spectroscopy and multi-dimensional homo and heteronuclear nuclear magnetic resonance spectroscopy that form an integral part of therapeutic peptides development. The application of combination biophysical platforms ensures the maintenance of the appropriate folded structure, which is a prerequisite for the safety and efficacy of peptide pharmaceuticals.

microRNA-92a promotes CNS autoimmunity by modulating the regulatory and inflammatory T cell balance.

A disequilibrium between immunosuppressive Tregs and inflammatory IL-17-producing Th17 cells is a hallmark of autoimmune diseases, including multiple sclerosis (MS). However, the molecular mechanisms underlying the Treg and Th17 imbalance in CNS autoimmunity remain largely unclear. Identifying the factors that drive this imbalance is of high clinical interest. Here, we report a major disease-promoting role for microRNA-92a (miR-92a) in CNS autoimmunity. miR-92a was elevated in experimental autoimmune encephalomyelitis (EAE), and its loss attenuated EAE. Mechanistically, miR-92a mediated EAE susceptibility in a T cell-intrinsic manner by restricting Treg induction and suppressive capacity, while supporting Th17 responses, by directly repressing the transcription factor Foxo1. Although miR-92a did not directly alter Th1 differentiation, it appeared to indirectly promote Th1 cells by inhibiting Treg responses. Correspondingly, miR-92a inhibitor therapy ameliorated EAE by concomitantly boosting Treg responses and dampening inflammatory T cell responses. Analogous to our findings in mice, miR-92a was elevated in CD4+ T cells from patients with MS, and miR-92a silencing in patients' T cells promoted Treg development but limited Th17 differentiation. Together, our results demonstrate that miR-92a drives CNS autoimmunity by sustaining the Treg/Th17 imbalance and implicate miR-92a as a potential therapeutic target for MS.

Regulation of splenic monocyte homeostasis and function by gut microbial products.

Splenic Ly6Chigh monocytes are innate immune cells involved in the regulation of central nervous system-related diseases. Recent studies have reported the shaping of peripheral immune responses by the gut microbiome via mostly unexplored pathways. In this study, we report that a 4-day antibiotic treatment eliminates certain families of the Bacteroidetes, Firmicutes, Tenericutes, and Actinobacteria phyla in the gut and reduces the levels of multiple pattern recognition receptor (PRR) ligands in the serum. Reduction of PRR ligands was associated with reduced numbers and perturbed function of splenic Ly6Chigh monocytes, which acquired an immature phenotype producing decreased levels of inflammatory cytokines and exhibiting increased phagocytic and anti-microbial abilities. Addition of PRR ligands in antibiotic-treated mice restored the number and functions of splenic Ly6Chigh monocytes. Our data identify circulating PRR ligands as critical regulators of the splenic Ly6Chigh monocyte behavior and suggest possible intervention pathways to manipulate this crucial immune cell subset.

Antimicrobial Hexaaquacopper(II) Complexes with Novel Polyiodide Chains.

The non-toxic inorganic antimicrobial agents iodine (I2) and copper (Cu) are interesting alternatives for biocidal applications. Iodine is broad-spectrum antimicrobial agent but its use is overshadowed by compound instability, uncontrolled iodine release and short-term effectiveness. These disadvantages can be reduced by forming complex-stabilized, polymeric polyiodides. In a facile, in-vitro synthesis we prepared the copper-pentaiodide complex [Cu(H2O)6(12-crown-4)5]I6 · 2I2, investigated its structure and antimicrobial properties. The chemical structure of the compound has been verified. We used agar well and disc-diffusion method assays against nine microbial reference strains in comparison to common antibiotics. The stable complex revealed excellent inhibition zones against C. albicans WDCM 00054, and strong antibacterial activities against several pathogens. [Cu(H2O)6(12-crown-4)5]I6 · 2I2 is a strong antimicrobial agent with an interesting crystal structure consisting of complexes located on an inversion center and surrounded by six 12-crown-4 molecules forming a cationic substructure. The six 12-crown-4 molecules form hydrogen bonds with the central Cu(H2O)6. The anionic substructure is a halogen bonded polymer which is formed by formal I5- repetition units. The topology of this chain-type polyiodide is unique. The I5- repetition units can be understood as a triodide anion connected to two iodine molecules.

MicroRNA-146a limits tumorigenic inflammation in colorectal cancer.

Chronic inflammation can drive tumor development. Here, we have identified microRNA-146a (miR-146a) as a major negative regulator of colonic inflammation and associated tumorigenesis by modulating IL-17 responses. MiR-146a-deficient mice are susceptible to both colitis-associated and sporadic colorectal cancer (CRC), presenting with enhanced tumorigenic IL-17 signaling. Within myeloid cells, miR-146a targets RIPK2, a NOD2 signaling intermediate, to limit myeloid cell-derived IL-17-inducing cytokines and restrict colonic IL-17. Accordingly, myeloid-specific miR-146a deletion promotes CRC. Moreover, within intestinal epithelial cells (IECs), miR-146a targets TRAF6, an IL-17R signaling intermediate, to restrict IEC responsiveness to IL-17. MiR-146a within IECs further suppresses CRC by targeting PTGES2, a PGE2 synthesis enzyme. IEC-specific miR-146a deletion therefore promotes CRC. Importantly, preclinical administration of miR-146a mimic, or small molecule inhibition of the miR-146a targets, TRAF6 and RIPK2, ameliorates colonic inflammation and CRC. MiR-146a overexpression or miR-146a target inhibition represent therapeutic approaches that limit pathways converging on tumorigenic IL-17 signaling in CRC.

Identification of TNFAIP3 as relapse biomarker and potential therapeutic target for MOG antibody associated diseases.

MOG-antibody associated disease (MOG-AAD) is a recently recognized demyelinating disorder predominantly affecting children but also occurs in adults, with a relapsing course in approximately 50% of patients. We evaluated peripheral blood mononuclear cells from MOG-AAD patients by flow cytometry and found a strong antigen specific central memory cell (CMC) response with increased Th1 and Th17 cells at the time of a relapse. Transcriptomic analysis of CMCs by three independent sequencing platforms revealed TNFAIP3 as a relapse biomarker, whose expression was down regulated at a relapse compared to remission in MOG-AAD patients. Serum in an additional cohort of patients showed decreased TNFAIP3 levels at relapse compared to remission state in MOG-AAD patients. Our studies suggest that alterations in TNFAIP3 levels are associated with relapses in MOG-AAD patients, which may have clinical utility as a disease course biomarker and therapeutic target.

A post-transcriptional program of chemoresistance by AU-rich elements and TTP in quiescent leukemic cells.

BACKGROUND: Quiescence (G0) is a transient, cell cycle-arrested state. By entering G0, cancer cells survive unfavorable conditions such as chemotherapy and cause relapse. While G0 cells have been studied at the transcriptome level, how post-transcriptional regulation contributes to their chemoresistance remains unknown. RESULTS: We induce chemoresistant and G0 leukemic cells by serum starvation or chemotherapy treatment. To study post-transcriptional regulation in G0 leukemic cells, we systematically analyzed their transcriptome, translatome, and proteome. We find that our resistant G0 cells recapitulate gene expression profiles of in vivo chemoresistant leukemic and G0 models. In G0 cells, canonical translation initiation is inhibited; yet we find that inflammatory genes are highly translated, indicating alternative post-transcriptional regulation. Importantly, AU-rich elements (AREs) are significantly enriched in the upregulated G0 translatome and transcriptome. Mechanistically, we find the stress-responsive p38 MAPK-MK2 signaling pathway stabilizes ARE mRNAs by phosphorylation and inactivation of mRNA decay factor, Tristetraprolin (TTP) in G0. This permits expression of ARE mRNAs that promote chemoresistance. Conversely, inhibition of TTP phosphorylation by p38 MAPK inhibitors and non-phosphorylatable TTP mutant decreases ARE-bearing TNFα and DUSP1 mRNAs and sensitizes leukemic cells to chemotherapy. Furthermore, co-inhibiting p38 MAPK and TNFα prior to or along with chemotherapy substantially reduces chemoresistance in primary leukemic cells ex vivo and in vivo. CONCLUSIONS: These studies uncover post-transcriptional regulation underlying chemoresistance in leukemia. Our data reveal the p38 MAPK-MK2-TTP axis as a key regulator of expression of ARE-bearing mRNAs that promote chemoresistance. By disrupting this pathway, we develop an effective combination therapy against chemosurvival.

Smad7 Controls Immunoregulatory PDL2/1-PD1 Signaling in Intestinal Inflammation and Autoimmunity.

Smad7, a negative regulator of TGF-ß signaling, has been implicated in the pathogenesis and treatment of inflammatory bowel diseases (IBDs), including Crohn's disease (CD) and ulcerative colitis (UC). Here, we found that Smad7 mediates intestinal inflammation by limiting the PDL2/1-PD1 axis in dendritic cells (DCs) and CD4+T cells. Smad7 deficiency in DCs promotes TGF-ß responsiveness and the co-inhibitory molecules PDL2/1 on DCs, and it further imprints T cell-PD1 signaling to promote Treg differentiation. DC-specific Smad7 deletion mitigates DSS-induced colitis by inducing CD103+PDL2/1+DCs and Tregs. In addition, Smad7 deficiency in CD4+T cells promotes PD1 and PD1-induced Tregs in vitro. The transfer of Smad7-deficient CD4+T cells enhances Tregs in vivo and protects against T cell-mediated colitis. Furthermore, Smad7 antisense ameliorates DSS-induced UC, increasing TGF-ß and PDL2/1-PD1 signaling. Enhancing PD1 signaling directly via Fc-fused PDL2/1 is also beneficial. Our results identify how Smad7 mediates intestinal inflammation and leverages these pathways therapeutically, providing additional strategies for IBD intervention.

Monomethyl fumarate treatment impairs maturation of human myeloid dendritic cells and their ability to activate T cells.

BACKGROUND: Dimethyl fumarate (DMF) and its active metabolite monomethyl fumarate (MMF) effectively lead to reduction in disease relapses and active magnetic resonance imaging (MRI) lesions. DMF and MMF are known to be effective in modulating T- and B-cell responses; however, their effect on the phenotype and function of human myeloid dendritic cells (mDCs) is not fully understood. OBJECTIVE: To investigate the role of MMF on human mDCs maturation and function. METHODS: mDCs from healthy controls were isolated and cultured in vitro with MMF. The effect of MMF on mDC gene expression was determined by polymerase chain reaction (PCR) array after in vitro MMF treatment. The ability of mDCs to activate T cells was assessed by in vitro co-culture system. mDCs from DMF-treated multiple sclerosis (MS) patients were analyzed by flow cytometry and PCR. RESULTS: MMF treatment induced a less mature phenotype of mDCs with reduced expression of major histocompatibility complex class II (MHC-II), co-stimulatory molecules CD86, CD40, CD83, and expression of nuclear factor κB (NF-κB) subunits RELA and RELB. mDCs from DMF-treated MS patients also showed the same immature phenotype. T cells co-cultured with MMF-treated mDCs showed reduced proliferation with decreased production of interferon gamma (IFN-γ), interleukin-17 (IL-17), and granulocyte-macrophage colony-stimulating factor (GM-CSF) compared to untreated cells. CONCLUSION: We report that MMF can modulate immune response by affecting human mDC function.

Identification of MS-specific serum miRNAs in an international multicenter study.

OBJECTIVE: To identify circulating microRNAs (miRNAs) linked to disease, disease stage, and disability in MS across cohorts. METHODS: Samples were obtained from the Comprehensive Longitudinal Investigation of Multiple Sclerosis (CLIMB, Boston, MA), EPIC (San Francisco, CA), AMIR (Beirut, Lebanon) as part of the SUMMIT consortium, and Stockholm Prospective Assessment of Multiple Sclerosis (Stockholm, Sweden) cohorts. Serum miRNA expression was measured using locked nucleic acid-based quantitative PCR. Four groups were compared: (1) MS vs healthy control (HC), (2) relapsing-remitting (RR) vs HC, (3) secondary progressive (SP) vs HC, and (4) RR vs SP. A Wilcoxon rank-sum test was used for the comparisons. The association between each miRNA and the Expanded Disability Status Scale (EDSS) score was assessed using the Spearman correlation coefficient. For each comparison, the p values were corrected for multiple comparisons using the approach of Benjamini and Hochberg to control the false discovery rate. RESULTS: In the CLIMB cohort, 5 miRNAs (hsa-miR-484, hsa-miR-140-5p, hsa-miR-320a, hsa-miR-486-5p, and hsa-miR-320c) showed a significant difference between patients with MS and healthy individuals; among these, miR-484 remained significant after accounting for multiple comparisons (p = 0.01). When comparing RRMS with HCs, hsa-miR-484 showed a significant difference (p = 0.004) between the groups after accounting for multiple group comparisons. When SP and HC were compared, 6 miRNAs (hsa-miR-484, hsa-miR-140-5p, hsa-miR-142-5p, hsa-miR-320a, hsa-miR-320b, and hsa-miR-320c) remained significantly different after accounting for multiple comparisons. Disability correlation analysis with miRNA provided 4 miRNAs (hsa-miR-320a, hsa-miR-337-3p, hsa-miR-199a-5p, and hsa-miR-142-5p) that correlated with the EDSS during the internal reproducibility phase. Among these, hsa-miR-337-3p was the most statistically significant miRNA that negatively correlated with the EDSS in three of the MS cohorts tested. CONCLUSIONS: These findings further confirm the use of circulating serum miRNAs as biomarkers to diagnose and monitor disease status in MS. CLASSIFICATION OF EVIDENCE: This study provides Class III evidence that levels of circulating miRNAs identify patients with MS.

Correlating serum micrornas and clinical parameters in amyotrophic lateral sclerosis.

INTRODUCTION: Amyotrophic lateral sclerosis (ALS) is a debilitating neurologic disorder with poor survival rates and no clear biomarkers for disease diagnosis and prognosis. METHODS: We compared serum microRNA (miRNA) expression from patients with ALS with healthy controls and patients with multiple sclerosis and Alzheimer disease. We also correlated miRNA expression in cross-sectional and longitudinal cohorts of ALS patients with clinical parameters. RESULTS: We identified 7 miRNAs (miR-192-5p, miR-192-3p, miR-1, miR-133a-3p, miR-133b, miR-144-5p, miR-19a-3p) that were upregulated and 6 miRNAs (miR-320c, miR-320a, let-7d-3p, miR-425-5p, miR-320b, miR-139-5p) that were downregulated in patients with ALS compared with healthy controls, patients with Alzheimer disease, and patients with multiple sclerosis. Changes in 4 miRNAs (miR-136-3p, miR-30b-5p, miR-331-3p, miR-496) correlated positively and change in 1 miRNA (miR-2110) correlated negatively with changes in clinical parameters in longitudinal analysis. DISCUSSION: Our findings identified serum miRNAs that can serve as biomarkers for ALS diagnosis and progression. Muscle Nerve 58: 261-269, 2018.

Association Between Serum MicroRNAs and Magnetic Resonance Imaging Measures of Multiple Sclerosis Severity.

Importance: MicroRNAs (miRNAs) are promising multiple sclerosis (MS) biomarkers. Establishing the association between miRNAs and magnetic resonance imaging (MRI) measures of disease severity will help define their significance and potential impact. Objective: To correlate circulating miRNAs in the serum of patients with MS to brain and spinal MRI. Design, Setting, and Participants: A cross-sectional study comparing serum miRNA samples with MRI metrics was conducted at a tertiary MS referral center. Two independent cohorts (41 and 79 patients) were retrospectively identified from the Comprehensive Longitudinal Investigation of Multiple Sclerosis at the Brigham and Women's Hospital. Expression of miRNA was determined by locked nucleic acid-based quantitative real-time polymerase chain reaction. Spearman correlation coefficients were used to test the association between miRNA and brain lesions (T2 hyperintense lesion volume [T2LV]), the ratio of T1 hypointense lesion volume [T1LV] to T2LV [T1:T2]), brain atrophy (whole brain and gray matter), and cervical spinal cord lesions (T2LV) and atrophy. The study was conducted from December 2013 to April 2016. Main Outcomes and Measures: miRNA expression. Results: Of the 120 patients included in the study, cohort 1 included 41 participants (7 [17.1%] men), with mean (SD) age of 47.7 (9.5) years; cohort 2 had 79 participants (26 [32.9%] men) with a mean (SD) age of 43.0 (7.5) years. Associations between miRNAs and MRIs were both protective and pathogenic. Regarding miRNA signatures, a topographic specificity differed for the brain vs the spinal cord, and the signature differed between T2LV and atrophy/destructive measures. Four miRNAs showed similar significant protective correlations with T1:T2 in both cohorts, with the highest for hsa.miR.143.3p (cohort 1: Spearman correlation coefficient rs = -0.452, P = .003; cohort 2: rs = -0.225, P = .046); the others included hsa.miR.142.5p (cohort 1: rs = -0.424, P = .006; cohort 2: rs = -0.226, P = .045), hsa.miR.181c.3p (cohort 1: rs = -0.383, P = .01; cohort 2: rs = -0.222, P = .049), and hsa.miR.181c.5p (cohort 1: rs = -0.433, P = .005; cohort 2: rs = -0.231, P = .04). In the 2 cohorts, hsa.miR.486.5p (cohort 1: rs = 0.348, P = .03; cohort 2: rs = 0.254, P = .02) and hsa.miR.92a.3p (cohort 1: rs = 0.392, P = .01; cohort 2: rs = 0.222, P = .049) showed similar significant pathogenic correlations with T1:T2; hsa.miR.375 (cohort 1: rs = -0.345, P = .03; cohort 2: rs = -0.257, P = .022) and hsa.miR.629.5p (cohort 1: rs = -0.350, P = .03; cohort 2: rs = -0.269, P = .02) showed significant pathogenic correlations with brain atrophy. Although we found several miRNAs associated with MRI outcomes, none of these associations remained significant when correcting for multiple comparisons, suggesting that further validation of our findings is needed. Conclusions and Relevance: Serum miRNAs may serve as MS biomarkers for monitoring disease progression and act as surrogate markers to identify underlying disease processes.

Comprehensive evaluation of serum microRNAs as biomarkers in multiple sclerosis.

OBJECTIVE: To identify circulating microRNAs (miRNAs) linked to disease stage and disability in multiple sclerosis (MS). METHODS: Sera from 296 participants including patients with MS, other neurologic diseases (Alzheimer disease and amyotrophic lateral sclerosis), and inflammatory diseases (rheumatoid arthritis and asthma) and healthy controls (HCs) were tested. miRNA profiles were determined using LNA (locked nucleic acid)-based quantitative PCR. Patients with MS were categorized according to disease stage and disability. In the discovery phase, 652 miRNAs were measured in sera from 26 patients with MS and 20 HCs. Following this, significant miRNAs (p < 0.05) from the discovery set were validated using quantitative PCR in 58 patients with MS, 30 HCs, and in 74 samples from other disease controls (Alzheimer disease, amyotrophic lateral sclerosis, asthma, and rheumatoid arthritis). RESULTS: We validated 7 miRNAs that differentiate patients with MS from HCs (p < 0.05 in both the discovery and validation phase); miR-320a upregulation was the most significantly changing serum miRNA in patients with MS. We also identified 2 miRNAs linked to disease progression, with miR-27a-3p being the most significant. Ten miRNAs correlated with the Expanded Disability Status Scale of which miR.199a.5p had the strongest correlation with disability. Of the 15 unique miRNAs we identified in the different group comparisons, 12 have previously been reported to be associated with MS but not in serum. CONCLUSIONS: Our findings identify circulating serum miRNAs as potential biomarkers to diagnose and monitor disease status in MS. CLASSIFICATION OF EVIDENCE: This study provides Class III evidence that circulating serum miRNAs can be used as biomarker for MS.

Identification of a novel mechanism of action of fingolimod (FTY720) on human effector T cell function through TCF-1 upregulation.

BACKGROUND: Fingolimod (FTY720), the first oral treatment for multiple sclerosis (MS), blocks immune cell trafficking and prevents disease relapses by downregulation of sphingosine-1-phosphate receptor. We determined the effect of FTY720 on human T cell activation and effector function. METHODS: T cells from MS patients and healthy controls were isolated to measure gene expression profiles in the presence or absence of FTY720 using nanostring and quantitative real-time polymerase chain reaction (qPCR). Cytokine protein expression was measured using luminex assay and flow cytometry analysis. Lentivirus vector carrying short hairpin RNA (shRNA) was used to knock down the expression of specific genes in CD4+ T cells. Chromatin immunoprecipitation was performed to assess T cell factor 1 (TCF-1) binding to promoter regions. Luciferase assays were performed to test the direct regulation of interferon gamma (IFN-γ) and granzyme B (GZMB) by TCF-1. Western blot analysis was used to assess the phosphorylation status of Akt and GSK3ß. RESULTS: We showed that FTY720 treatment not only affects T cell trafficking but also T cell activation. Patients treated with FTY720 showed a significant reduction in circulating CD4 T cells. Activation of T cells in presence of FTY720 showed a less inflammatory phenotype with reduced production of IFN-γ and GZMB. This decreased effector phenotype of FTY720-treated T cells was dependent on the upregulation of TCF-1. FTY720-induced TCF-1 downregulated the pathogenic cytokines IFN-γ and GZMB by binding to their promoter/enhancer regions and mediating epigenetic modifications. Furthermore, we observed that TCF-1 expression was lower in T cells from multiple sclerosis patients than in those from healthy individuals, and FTY720 treatment increased TCF-1 expression in multiple sclerosis patients. CONCLUSIONS: These results reveal a previously unknown mechanism of the effect of FTY720 on human CD4+ T cell modulation in multiple sclerosis and demonstrate the role of TCF-1 in human T cell activation and effector function.