CNS Drug Discovery in Academia: Where Basic Research Meets Innovation.

The involvement of academic research in drug discovery is consistently growing. However, academic projects seldom advance to clinical trials. Here, we assess the landscape of drug discovery within the National Centre of Competence in Research (NCCR) TransCure launched by the Swiss National Science Foundation to foster basic research and early-stage drug discovery on membrane transporters. This included transporters in central nervous system (CNS) disorders, which represent a huge unmet medical need. While idea championship, sustainable funding, collaborations between disciplines at the interface of academia and industry are important for translational research, Popperian falsifiability, strong intellectual property and a motivated startup team are key elements for innovation. This is exemplified by the NCCR TransCure spin-off company Synendos Therapeutics, a clinical stage biotech company developing the first selective endocannabinoid reuptake inhibitors (SERIs) as novel treatment for neuropsychiatric disorders. We provide a perspective on the challenges related to entering an uncharted druggable space and bridging the often mentioned "valley of death". The high attrition rate of drug discovery projects in the CNS field within academia is often due to the lack of meaningful animal models that can provide pharmacological proof-of-concept for potentially disruptive technologies at the earliest stages, and the absence of solid intellectual property.

Dysfunction of homeostatic control of dopamine by astrocytes in the developing prefrontal cortex leads to cognitive impairments.

Astrocytes orchestrate neural development by powerfully coordinating synapse formation and function and, as such, may be critically involved in the pathogenesis of neurodevelopmental abnormalities and cognitive deficits commonly observed in psychiatric disorders. Here, we report the identification of a subset of cortical astrocytes that are competent for regulating dopamine (DA) homeostasis during postnatal development of the prefrontal cortex (PFC), allowing for optimal DA-mediated maturation of excitatory circuits. Such control of DA homeostasis occurs through the coordinated activity of astroglial vesicular monoamine transporter 2 (VMAT2) together with organic cation transporter 3 and monoamine oxidase type B, two key proteins for DA uptake and metabolism. Conditional deletion of VMAT2 in astrocytes postnatally produces loss of PFC DA homeostasis, leading to defective synaptic transmission and plasticity as well as impaired executive functions. Our findings show a novel role for PFC astrocytes in the DA modulation of cognitive performances with relevance to psychiatric disorders.

Δ9-cis-Tetrahydrocannabinol: Natural Occurrence, Chirality, and Pharmacology.

The cis-stereoisomers of Δ9-THC [(-)-3 and (+)-3] were identified and quantified in a series of low-THC-containing varieties of Cannabis sativa registered in Europe as fiber hemp and in research accessions of cannabis. While Δ9-cis-THC (3) occurs in cannabis fiber hemp in the concentration range of (-)-Δ9-trans-THC [(-)-1], it was undetectable in a sample of high-THC-containing medicinal cannabis. Natural Δ9-cis-THC (3) is scalemic (ca. 80-90% enantiomeric purity), and the absolute configuration of the major enantiomer was established as 6aS,10aR [(-)-3] by chiral chromatographic comparison with a sample available by asymmetric synthesis. The major enantiomer, (-)-Δ9-cis-THC [(-)-3], was characterized as a partial cannabinoid agonist in vitro and elicited a full tetrad response in mice at 50 mg/kg doses. The current legal discrimination between narcotic and non-narcotic cannabis varieties centers on the contents of "Δ9-THC and isomers" and needs therefore revision, or at least a more specific wording, to account for the presence of Δ9-cis-THCs [(+)-3 and (-)-3] in cannabis fiber hemp varieties.

Modification on the 1,2-dihydro-2-oxo-pyridine-3-carboxamide core to obtain multi-target modulators of endocannabinoid system.

Several preclinical evidence indicate that the modulation of the endocannabinoid system (ECS) represents a promising therapeutic approach for different diseases. However, only few modulators of this system have reached so far an advanced stage of clinical development, mainly due to limited efficacy and CB1 receptor-dependent side effects. Those limitations might be overcome by multi-target compounds that exert pro-cannabinoid activities through the modulation of two or more targets in the ECS. This approach can offer a safer and more effective pharmacological strategy as compared to the modulation of a single target. In this work, we report the synthesis and biological characterization of new 6-aryl-1,2-dihydro-2-oxo-pyridine-3-carboxamide derivatives. Our results identified several compounds exhibiting interesting multi-target profiles within the ECS. In particular, compound B1 showed moderate-to-high affinity for cannabinoid receptors (Ki CB1R = 304 nM, partial agonist, Ki CB2R = 3.1 nM, inverse agonist) and a potent inhibition of AEA uptake (IC50 = 62 nM) with moderate inhibition of FAAH (IC50 = 2.9 µM). The corresponding 2-alkoxypyridine analogue B14 exhibited significant inhibitor activity on both FAAH (IC50 = 69 nM) and AEA uptake (IC50 = 76 nM) without significantly binding to both cannabinoid receptor subtypes. Molecular docking analysis was carried out on the three-dimensional structures of CB1R and CB2R and of FAAH to rationalize the structure-activity relationships of this series of compounds.

Chemical probes to potently and selectively inhibit endocannabinoid cellular reuptake.

The extracellular effects of the endocannabinoids anandamide and 2-arachidonoyl glycerol are terminated by enzymatic hydrolysis after crossing cellular membranes by facilitated diffusion. The lack of potent and selective inhibitors for endocannabinoid transport has prevented the molecular characterization of this process, thus hindering its biochemical investigation and pharmacological exploitation. Here, we report the design, chemical synthesis, and biological profiling of natural product-derived N-substituted 2,4-dodecadienamides as a selective endocannabinoid uptake inhibitor. The highly potent (IC50 = 10 nM) inhibitor N-(3,4-dimethoxyphenyl)ethyl amide (WOBE437) exerted pronounced cannabinoid receptor-dependent anxiolytic, antiinflammatory, and analgesic effects in mice by increasing endocannabinoid levels. A tailored WOBE437-derived diazirine-containing photoaffinity probe (RX-055) irreversibly blocked membrane transport of both endocannabinoids, providing mechanistic insights into this complex process. Moreover, RX-055 exerted site-specific anxiolytic effects on in situ photoactivation in the brain. This study describes suitable inhibitors to target endocannabinoid membrane trafficking and uncovers an alternative endocannabinoid pharmacology.

Synthesis, Microtubule-Binding Affinity, and Antiproliferative Activity of New Epothilone Analogs and of an EGFR-Targeted Epothilone-Peptide Conjugate.

A new simplified, epoxide-free epothilone analog was prepared incorporating an N-(2-hydroxyethyl)-benzimidazole side chain, which binds to microtubules with high affinity and inhibits cancer cell growth in vitro with nM potency. Building on this scaffold, a disulfide-linked conjugate with the purported EGFR-binding (EGFR, epidermal growth factor receptor) peptide GE11 was then prepared. The conjugate retained significant microtubule-binding affinity, in spite of the size of the peptide attached to the benzimidazole side chain. The antiproliferative activity of the conjugate was significantly lower than for the parent scaffold and, surprisingly, was independent of the EGFR expression status of cells. Our data indicate that the disulfide-based conjugation with the GE11 peptide is not a viable approach for effective tumor-targeting of highly potent epothilones and probably not for other cytotoxics.

Biological evaluation of pyridone alkaloids on the endocannabinoid system.

Naturally occurring pyridone alkaloids as well as synthetic derivatives were previously shown to induce neurite outgrowth. However, the molecular basis for this biological effect remains poorly understood. In this work, we have prepared new pyridones, and tested the effect of thirteen 4-hydroxy-2-pyridone derivatives on the components of the endocannabinoid system. Investigation of binding affinities towards CB1 and CB2 receptors led to the identification of a compound binding selectively to CB1 (12). Compound 12 and a closely related derivative (11) also inhibited anandamide (AEA) hydrolysis by fatty acid amide hydrolase. Interestingly, none of the compounds tested showed any effect on 2-AG hydrolysis by monoacylglycerol lipase at 10µM. Assessment of AEA uptake did, however, lead to the identification of four inhibitors with IC50 values in the submicromolar range and high selectivity over the other components of the endocannabinoid system.

Novel analogs of PSNCBAM-1 as allosteric modulators of cannabinoid CB1 receptor.

In this work, we explored the molecular framework of the known CB1R allosteric modulator PSNCBAM-1 with the aim to generate new bioactive analogs and to deepen the structure-activity relationships of this type of compounds. In particular, the introduction of a NH group between the pyridine ring and the phenyl nucleus generated the amino-phenyl-urea derivative SN15b that behaved as a positive allosteric modulator (PAM), increasing the CB1R binding affinity of the orthosteric ligand CP55,940. The functional activity was evaluated using serum response element (SRE) assay, which assesses the CB1R-dependent activation of the MAPK/ERK signaling pathway. SN15b and the biphenyl-urea analog SC4a significantly inhibited the response produced by CP55,940 in the low µM range, thus behaving as negative allosteric modulators (NAMs). The new derivatives presented here provide further insights about the modulation of CB1R binding and functional activity by allosteric ligands.

4'-O-methylhonokiol increases levels of 2-arachidonoyl glycerol in mouse brain via selective inhibition of its COX-2-mediated oxygenation.

BACKGROUND AND PURPOSE: 4'-O-methylhonokiol (MH) is a natural product showing anti-inflammatory, anti-osteoclastogenic, and neuroprotective effects. MH was reported to modulate cannabinoid CB2 receptors as an inverse agonist for cAMP production and an agonist for intracellular [Ca2+]. It was recently shown that MH inhibits cAMP formation via CB2 receptors. In this study, the exact modulation of MH on CB2 receptor activity was elucidated and its endocannabinoid substrate-specific inhibition (SSI) of cyclooxygenase-2 (COX-2) and CNS bioavailability are described for the first time. METHODS: CB2 receptor modulation ([35S]GTPγS, cAMP, and ß-arrestin) by MH was measured in hCB2-transfected CHO-K1 cells and native conditions (HL60 cells and mouse spleen). The COX-2 SSI was investigated in RAW264.7 cells and in Swiss albino mice by targeted metabolomics using LC-MS/MS. RESULTS: MH is a CB2 receptor agonist and a potent COX-2 SSI. It induced partial agonism in both the [35S]GTPγS binding and ß-arrestin recruitment assays while being a full agonist in the cAMP pathway. MH selectively inhibited PGE2 glycerol ester formation (over PGE2) in RAW264.7 cells and significantly increased the levels of 2-AG in mouse brain in a dose-dependent manner (3 to 20 mg kg(-1)) without affecting other metabolites. After 7 h from intraperitoneal (i.p.) injection, MH was quantified in significant amounts in the brain (corresponding to 200 to 300 nM). CONCLUSIONS: LC-MS/MS quantification shows that MH is bioavailable to the brain and under condition of inflammation exerts significant indirect effects on 2-AG levels. The biphenyl scaffold might serve as valuable source of dual CB2 receptor modulators and COX-2 SSIs as demonstrated by additional MH analogs that show similar effects. The combination of CB2 agonism and COX-2 SSI offers a yet unexplored polypharmacology with expected synergistic effects in neuroinflammatory diseases, thus providing a rationale for the diverse neuroprotective effects reported for MH in animal models.

Reactions of CF3-enones with arenes under superelectrophilic activation: a pathway to trans-1,3-diaryl-1-CF3-indanes, new cannabinoid receptor ligands.

4-Aryl-1,1,1-trifluorobut-3-en-2-ones ArCH[double bond, length as m-dash]CHCOCF3 (CF3-enones) react with arenes in excess of Brønsted superacids (TfOH, FSO3H) to give, stereoselectively, trans-1,3-diaryl-1-trifluoromethyl indanes in 35-85% yields. The reaction intermediates, the O-protonated ArCH[double bond, length as m-dash]CHC(OH(+))CF3 and the O,C-diprotonated ArHC(+)CH2C(OH(+))CF3 species, have been studied by means of (1)H, (13)C, (19)F NMR, and DFT calculations. Both types of the cations may participate in the reaction, depending on their electrophilicity and electron-donating properties of the arenes. The formation of CF3-indanes is a result of cascade reaction of protonated CF3-enones to form chemo-, regio- and stereoselectively three new C-C bonds. The obtained trans-1,3-diaryl-1-trifluoromethyl indanes were investigated as potential ligands for cannabinoid receptors CB1 and CB2 types. The most potent compound showed sub-micromolar affinity for both receptor subtypes with a 6-fold selectivity toward the CB2 receptor with no appreciable cytotoxicity toward SHSY5Y cells.

Elevated levels of endocannabinoids in chronic hepatitis C may modulate cellular immune response and hepatic stellate cell activation.

The endocannabinoid (EC) system is implicated in many chronic liver diseases, including hepatitis C viral (HCV) infection. Cannabis consumption is associated with fibrosis progression in patients with chronic hepatitis C (CHC), however, the role of ECs in the development of CHC has never been explored. To study this question, anandamide (AEA) and 2-arachidonoyl glycerol (2-AG) were quantified in samples of HCV patients and healthy controls by gas and liquid chromatography mass spectrometry. Fatty acid amide hydrolase (FAAH) and monoaclyglycerol lipase (MAGL) activity was assessed by [3H]AEA and [3H]2-AG hydrolysis, respectively. Gene expression and cytokine release were assayed by TaqMan PCR and ELISpot, respectively. AEA and 2-AG levels were increased in plasma of HCV patients, but not in liver tissues. Hepatic FAAH and MAGL activity was not changed. In peripheral blood mononuclear cells (PBMC), ECs inhibited IFN-γ, TNF-α, and IL-2 secretion. Inhibition of IL-2 by endogenous AEA was stronger in PBMC from HCV patients. In hepatocytes, 2-AG induced the expression of IL-6, -17A, -32 and COX-2, and enhanced activation of hepatic stellate cells (HSC) co-cultivated with PBMC from subjects with CHC. In conclusion, ECs are increased in plasma of patients with CHC and might reveal immunosuppressive and profibrogenic effects.

A highly potent, orally bioavailable pyrazole-derived cannabinoid CB2 receptor-selective full agonist for in vivo studies.

The cannabinoid CB2 receptor (CB2R) is a potential therapeutic target for distinct forms of tissue injury and inflammatory diseases. To thoroughly investigate the role of CB2R in pathophysiological conditions and for target validation in vivo, optimal pharmacological tool compounds are essential. Despite the sizable progress in the generation of potent and selective CB2R ligands, pharmacokinetic parameters are often neglected for in vivo studies. Here, we report the generation and characterization of a tetra-substituted pyrazole CB2R full agonist named RNB-61 with high potency (K i 0.13-1.81 nM, depending on species) and a peripherally restricted action due to P-glycoprotein mediated efflux from the brain. 3H and 14C labelled RNB-61 showed apparent K d values < 4 nM towards human CB2R in both cell and tissue experiments. The >6000-fold selectivity over CB1 receptors and negligible off-targets in vitro, combined with high oral bioavailability and suitable systemic pharmacokinetic (PK) properties, prompted the assessment of RNB-61 in a mouse ischemia-reperfusion model of acute kidney injury (AKI) and in a rat model of chronic kidney injury/inflammation and fibrosis (CKI) induced by unilateral ureteral obstruction. RNB-61 exerted dose-dependent nephroprotective and/or antifibrotic effects in the AKI/CKI models. Thus, RNB-61 is an optimal CB2R tool compound for preclinical in vivo studies with superior biophysical and PK properties over generally used CB2R ligands.

Evidence for bidirectional endocannabinoid transport across cell membranes.

Despite extensive research on the trafficking of anandamide (AEA) across cell membranes, little is known about the membrane transport of other endocannabinoids, such as 2-arachidonoylglycerol (2-AG). Previous studies have provided data both in favor and against a cell membrane carrier-mediated transport of endocannabinoids, using different methodological approaches. Because AEA and 2-AG undergo rapid and almost complete intracellular hydrolysis, we employed a combination of radioligand assays and absolute quantification of cellular and extracellular endocannabinoid levels. In human U937 leukemia cells, 100 nm AEA and 1 µm 2-AG were taken up through a fast and saturable process, reaching a plateau after 5 min. Employing differential pharmacological blockage of endocannabinoid uptake, breakdown, and interaction with intracellular binding proteins, we show that eicosanoid endocannabinoids harboring an arachidonoyl chain compete for a common membrane target that regulates their transport, whereas other N-acylethanolamines did not interfere with AEA and 2-AG uptake. By combining fatty acid amide hydrolase or monoacyl glycerol lipase inhibitors with hydrolase-inactive concentrations of the AEA transport inhibitors UCM707 (1 µm) and OMDM-2 (5 µm), a functional synergism on cellular AEA and 2-AG uptake was observed. Intriguingly, structurally unrelated AEA uptake inhibitors also blocked the cellular release of AEA and 2-AG. We show, for the first time, that UCM707 and OMDM-2 inhibit the bidirectional movement of AEA and 2-AG across cell membranes. Our findings suggest that a putative endocannabinoid cell membrane transporter controls the cellular AEA and 2-AG trafficking and metabolism.

Identification and quantification of a new family of peptide endocannabinoids (Pepcans) showing negative allosteric modulation at CB1 receptors.

The α-hemoglobin-derived dodecapeptide RVD-hemopressin (RVDPVNFKLLSH) has been proposed to be an endogenous agonist for the cannabinoid receptor type 1 (CB(1)). To study this peptide, we have raised mAbs against its C-terminal part. Using an immunoaffinity mass spectrometry approach, a whole family of N-terminally extended peptides in addition to RVD-Hpα were identified in rodent brain extracts and human and mouse plasma. We designated these peptides Pepcan-12 (RVDPVNFKLLSH) to Pepcan-23 (SALSDLHAHKLRVDPVNFKLLSH), referring to peptide length. The most abundant Pepcans found in the brain were tested for CB(1) receptor binding. In the classical radioligand displacement assay, Pepcan-12 was the most efficacious ligand but only partially displaced both [(3)H]CP55,940 and [(3)H]WIN55,212-2. The data were fitted with the allosteric ternary complex model, revealing a cooperativity factor value α < 1, thus indicating a negative allosteric modulation. Dissociation kinetic studies of [(3)H]CP55,940 in the absence and presence of Pepcan-12 confirmed these results by showing increased dissociation rate constants induced by Pepcan-12. A fluorescently labeled Pepcan-12 analog was synthesized to investigate the binding to CB(1) receptors. Competition binding studies revealed K(i) values of several Pepcans in the nanomolar range. Accordingly, using competitive ELISA, we found low nanomolar concentrations of Pepcans in human plasma and ∼100 pmol/g in mouse brain. Surprisingly, Pepcan-12 exhibited potent negative allosteric modulation of the orthosteric agonist-induced cAMP accumulation, [(35)S]GTPγS binding, and CB(1) receptor internalization. Pepcans are the first endogenous allosteric modulators identified for CB(1) receptors. Given their abundance in the brain, Pepcans could play an important physiological role in modulating endocannabinoid signaling.

Assay of Endocannabinoid Uptake.

Endocannabinoids at nanomolar physiological concentrations cross cellular membranes by facilitated diffusion, a process that can be studied by measuring transport kinetics and endocannabinoid trafficking employing radioligands and mass spectrometry. Here, we describe radiosubstrate-based assays using arachidonoyl[1-3H]ethanolamine and 2-arachidonoyl[1,2,3-3H]glycerol to measure cellular endocannabinoid uptake in a three-phase assay with human U937 cells. Liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS/MS)-based lipidomics was used to interrogate the roles of serum and albumin for endocannabinoid trafficking in U937 cells.

Analysis of reptilian APOBEC1 suggests that RNA editing may not be its ancestral function.

The Activation Induced Deaminase (AID)/APOBEC family of deaminases targeting nucleic acids arose at the beginning of the vertebrate radiation and further expanded in mammals. Following an analysis of the available genomic data, we report the identification of the APOBEC5, a novel group of paralogues in tetrapods. Moreover, we find bona fide homologues of Apolipoprotein B Editing Complex 1 (APOBEC1) in the genomes of anole lizard and zebra finch, thus implying its appearance prior to the divergence of the amniotes. apolipoprotein B editing complex 1 (APOBEC1), in contrast with other AID/APOBECs acting on DNA, is an RNA-editing enzyme that targets the transcript of Apolipoprotein B (ApoB), thereby causing the translation of a truncated form of the protein. 3'RACE experiments reveal a lizard APOBEC1-like molecule lacking a C-terminal region important for mammalian ApoB RNA editing. This observation pairs with the finding that lizard ApoB is not deaminated at the region corresponding to the mammalian site of editing. Similar to mammalian APOBEC1, the lizard protein is able to deaminate DNA in bacteria and shows a conserved mutational context. Although not precluding the possibility that lizard APOBEC1 acts on unknown mRNA targets, these findings suggest that its ability to target DNA predates its role in RNA editing.

Synthesis and cytotoxic activity of non-naturally substituted 4-oxycoumarin derivatives.

Coumarins are a large family of natural and synthetic compounds exerting different pharmacological effects, including cytotoxic, anti-inflammatory or antimicrobial. In the present communication we report the synthesis of a series of 12 diversely substituted 4-oxycoumarin derivatives including methoxy substituted 4-hydroxycoumarins, methyl, methoxy or unsubstituted 3-aryl-4-hydroxycoumarins and 4-benzyloxycoumarins and their anti-proliferative effects on breast adenocarcinoma cells (MCF-7), human promyelocytic leukemia cells (HL-60), human histiocytic lymphoma cells (U937) and mouse neuroblastoma cells (Neuro2a). The most potent bioactive molecule was the 4-hydroxy-5,7-dimethoxycoumarin (compound 1) which showed similar potency (IC(50) 0.2-2 µM) in all cancer cell lines tested. This non-natural product reveals a simple bioactive scaffold which may be exploited in further studies.

Association of endocannabinoids with pain in endometriosis.

ABSTRACT: Endocannabinoid (eCB) levels fluctuate in inflammatory conditions and as such may take part in endometriosis-associated pain or even in endometriosis pathogenesis. In this case-control (23 cases and 19 controls) study, targeted lipids were measured in the serum and peritoneal fluid collected during laparoscopy. Endometriosis was confirmed histologically. Dysmenorrhea, abdominal pain, and dyspareunia were assessed using the Numeric Rating Scale for pain. Steroids, eCBs, and related lipids were quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Tumor necrosis factor alpha, IL-8, PAPP-A, PP14, RANTES, OPG, MIDKINE, MCP-1, VEGF, leptin, and defensins were quantified by ELISA. We found that eCB levels were significantly influenced by both noncyclic and cyclic abdominal pain. Specifically, women suffering from noncyclic abdominal pain were characterized by a higher 2-AG level in the peritoneal fluid throughout the menstrual cycle, whereas women suffering from dysmenorrhea had higher 2-AG levels and lower AEA levels during the proliferative phase alone. In addition, 2-AG positively correlated with prostaglandin E2 (PGE2), and the ratio AEA/2-AG positively correlated with defensins, suggesting a possible link between endocannabinoids system and inflammatory pain. The results of the current study indicate that the eCB system may play a role in endometriosis-associated pain, but additional studies are needed to investigate the causal relationship.

Reversible Monoacylglycerol Lipase Inhibitors: Discovery of a New Class of Benzylpiperidine Derivatives.

Monoacylglycerol lipase (MAGL) is the enzyme responsible for the metabolism of 2-arachidonoylglycerol in the brain and the hydrolysis of peripheral monoacylglycerols. Many studies demonstrated beneficial effects deriving from MAGL inhibition for neurodegenerative diseases, inflammatory pathologies, and cancer. MAGL expression is increased in invasive tumors, furnishing free fatty acids as pro-tumorigenic signals and for tumor cell growth. Here, a new class of benzylpiperidine-based MAGL inhibitors was synthesized, leading to the identification of 13, which showed potent reversible and selective MAGL inhibition. Associated with MAGL overexpression and the prognostic role in pancreatic cancer, derivative 13 showed antiproliferative activity and apoptosis induction, as well as the ability to reduce cell migration in primary pancreatic cancer cultures, and displayed a synergistic interaction with the chemotherapeutic drug gemcitabine. These results suggest that the class of benzylpiperidine-based MAGL inhibitors have potential as a new class of therapeutic agents and MAGL could play a role in pancreatic cancer.

Selective Endocannabinoid Reuptake Inhibitor WOBE437 Reduces Disease Progression in a Mouse Model of Multiple Sclerosis.

The modulation of the endocannabinoid system (ECS) has shown positive results in animal models of multiple sclerosis (MS) and immune and inflammatory disorders. However, chronic administration of CB1 receptor agonists and degrading enzyme inhibitors can lead to CB1 receptor desensitization and sedation. WOBE437 is the prototype of a new class of ECS modulators named selective endocannabinoid reuptake inhibitors (SERIs), which mildly and selectively increase central endocannabinoid levels with a self-limiting mode of action. In previous studies, WOBE437 demonstrated analgesic, anxiolytic, and anti-inflammatory effects. Here, we tested the therapeutic potential of WOBE437 in a clinically relevant mouse model of MS (experimental autoimmune encephalomyelitis). C57BL/6 mice were administered WOBE437 (10 mg/kg, 20 days) or vehicle using two therapeutic options: (1) starting the treatment at the disease onset or (2) before reaching the peak of the disease. In both strategies, WOBE437 significantly reduced disease severity and accelerated recovery through CB1 and CB2 receptor-dependent mechanisms. At the peak of the disease, WOBE437 increased endocannabinoid levels in the cerebellum, concurring with a reduction of central nervous system (CNS)-infiltrating immune cells and lower microglial proliferation. At the end of treatment, endocannabinoid levels were mildly increased in brain, cerebellum, and plasma of WOBE437-treated mice, without desensitization of CB1 receptor in the brain and cerebellum. In a mouse model of spasticity (Straub test), WOBE437 (10 mg/kg) induced significant muscle relaxation without eliciting the typical sedative effects associated with muscle relaxants or CB1 receptor agonists. Collectively, our results show that WOBE437 (and SERIs) may represent a novel therapeutic strategy for slowing MS progression and control major symptoms.