Monoacylglycerol Lipase Inhibitor Research Articles

Monoacylglycerol Lipase (MAGL) Inhibition Impedes the Osteosarcoma Progression by Regulating Epithelial Mesenchymal Transition.

Osteosarcoma is a primary malignancy of mesenchymal origin, and its metastasis and multidrug resistance remain major problems affecting the therapeutic effect. This study aimed to evaluate the efficacy and underlying mechanism of monoacylglycerol lipase (MAGL) on osteosarcoma progression. MAGL expression was downregulated by shMAGL or JZL184 (an MAGL inhibitor) and upregulated through plasmid. RT-PCR and Western blot were utilized to determine the expression of target molecules. CCK-8 assay, transwell assay and ROS assay were performed to investigate the inhibitory effect of MAGL on the growth and metastasis of osteosarcoma cells. The role of JZL184 on tumor growth was examined in cisplatin-resistant MG-63 (MG-63/R) xenograft model. MAGL was highly expressed in osteosarcoma cells and tissues. MAGL knockdown significantly impeded the proliferation, clone formation, invasion and migration of MG-63 cells, whereas opposite result was observed in 143B cells with MAGL overexpression. Likewise, an MAGL inhibitor JZL184 displayed reduced viability, clone formation, invasion and migration of osteosarcoma cells. Western blot of the epithelial mesenchymal transition (EMT)-related proteins indicated that MAGL knockdown or JZL184 could upregulated E-cadherin expression and downregulated vimentin expression. In vitro and in vivo experiments indicated that JZL184 re-sensitized MG-63/R cells to cisplatin. In summary, MAGL regulated osteosarcoma by modulating EMT, and JZL184 might be a promising agent for osteosarcoma patients who are resistant to cisplatin.

Endocannabinoid metabolism and Alzheimer's disease.

Endocannabinoid metabolism and Alzheimer's disease.

A carrier free delivery system of a monoacylglycerol lipase hydrophobic inhibitor

Monoacylglycerol lipase (MAGL) is an emerging therapeutic target for cancer. It is involved in lipid metabolism and its inhibition impairs many hallmarks of cancer including cell proliferation, migration/invasion and tumor growth. For these reasons, our group has recently developed a potent reversible MAGL inhibitor (MAGL23), which showed promising anticancer activities. Here in, to improve its pharmacological properties, a nanoformulation based on nanocrystals coated with albumin was prepared for therapeutic applications. MAGL23 was solubilized by a nanocrystallization method with Pluronic F-127 as surfactant into an organic solvent and was recovered as nanocrystals in water after solvent evaporation. Finally, the solubilized nanocrystals were stabilized by human serum albumin to create a smart delivery carrier. An in-silico prediction (lipophilicity, structure at different pH and solubility in water), as well as experimental studies (solubility), have been performed to check the chemical properties of the inhibitor and nanocrystals. The solubility in water increases from less than 0.01 mg/mL (0.0008 mg/mL, predicted) up to 0.82 mg/mL in water. The formulated inhibitor maintained its potency in ovarian and colon cancer cell lines as the free drug. Furthermore, the system was thoroughly observed at each step of the solubilization process till the final formulation stage by different spectroscopic techniques and a comparative study was performed to check the effects of Pluronic F-127 and CTAB as surfactants. The formulated system is favorable to release the drug at physiological pH conditions (at pH 7.4, after 24 h, less than 20% of compound is released). In vivo studies have shown that albumin-complexed nanocrystals increase the therapeutic window of MAGL23 along with a favorable biodistribution. As per our knowledge, we are reporting the first ever nanoformulation of a MAGL inhibitor, which is promising as a therapeutic system where the MAGL enzyme is involved, especially for cancer therapeutic applications.

The Novel Monoacylglycerol Lipase Inhibitor MJN110 Suppresses Neuroinflammation, Normalizes Synaptic Composition and Improves Behavioral Performance in the Repetitive Traumatic Brain Injury Mouse Model.

Modulation of the endocannabinoid system has emerged as an effective approach for the treatment of many neurodegenerative and neuropsychological diseases. However, the underlying mechanisms are still uncertain. Using a repetitive mild traumatic brain injury (mTBI) mouse model, we found that there was an impairment in locomotor function and working memory within two weeks post-injury, and that treatment with MJN110, a novel inhibitor of the principal 2-arachidononyl glycerol (2-AG) hydrolytic enzyme monoacylglycerol lipase dose-dependently ameliorated those behavioral changes. Spatial learning and memory deficits examined by Morris water maze between three and four weeks post-TBI were also reversed in the drug treated animals. Administration of MJN110 selectively elevated the levels of 2-AG and reduced the production of arachidonic acid (AA) and prostaglandin E2 (PGE2) in the TBI mouse brain. The increased production of proinflammatory cytokines, accumulation of astrocytes and microglia in the TBI mouse ipsilateral cerebral cortex and hippocampus were significantly reduced by MJN110 treatment. Neuronal cell death was also attenuated in the drug treated animals. MJN110 treatment normalized the expression of the NMDA receptor subunits NR2A and NR2B, the AMPA receptor subunits GluR1 and GluR2, and the GABAA receptor subunits α1, β2,3 and γ2, which were all reduced at 1, 2 and 4 weeks post-injury. The reduced inflammatory response and restored glutamate and GABA receptor expression likely contribute to the improved motor function, learning and memory in the MJN110 treated animals. The therapeutic effects of MJN110 were partially mediated by activation of CB1 and CB2 cannabinoid receptors and were eliminated when it was co-administered with DO34, a novel inhibitor of the 2-AG biosynthetic enzymes. Our results suggest that augmentation of the endogenous levels of 2-AG can be therapeutically useful in the treatment of TBI by suppressing neuroinflammation and maintaining the balance between excitatory and inhibitory neurotransmission.

Inhibition of monoacylglycerol lipase restrains proliferation, migration, invasion, tumor growth and induces apoptosis in cervical cancer.

Cervical cancer is one of common diseases among women. There are limited therapies for patients with metastatic or recurrent cervical cancer. This study sought to explore the role of monoacylglycerol lipase (MAGL), an important metabolic enzyme, in cervical cancer progression. In in vitro experiments, MAGL expression was inhibited by si-MAGL or JZL184 in cervical cancer cells. Quantitative real-time polymerase chain reaction and western blotting were performed to measure the expression of target molecules. Proliferation of cervical cancer cells was assessed by CCK-8 and colony formation assays. Apoptosis and cell cycle progression were evaluated by flow cytometry. The migration and invasion were detected by transwell assay. The in vivo tumor growth was detected in nude mice. TUNEL was utilized to observe apoptotic cells in tumor tissues. MAGL was upregulated in cervical cancer tissues and cells. Further, MAGL inhibition suppressed the growth of cervical cancer cells in vitro and in vivo. In addition, apoptosis and G1-phase cell cycle arrest were induced by MAGL knockdown. MAGL silencing-mediated upregulation of Bax and cleaved caspase-3, and downregulation of Bcl-2 was responsible for triggering apoptosis. More importantly, the migration and invasion of cervical cancer cells were restrained by MAGL depletion. MAGL drives the progression of cervical cancer, which can be a promising candidate to identify effective therapy for cervical cancer.

Glycerol is Released from a New Path in MGL Lipase Catalytic Process.

Traditionally, it is believed that the substrate and products of a monoacylglycerol lipase (MGL) share the same path to enter and exit the catalytic site. Glycerol (a product of MGL), however, was recently hypothesized to be released through a different path. In order to improve the catalytic efficacy and thermo-stability of MGL, it is important to articulate the pathways of a MGL products releasing. In this study, with structure biological approaches, biochemical experiments, and in silico methods, we prove that glycerol is released from a different path in the catalytic site indeed. The fatty acid (another product of MGL) does share the same binding path with the substrate. This discovery paves a new road to design MGL inhibitors or optimize MGL catalytic efficacy.

P.0296 Sex-dependent antinociceptive effects of fatty acid amide hydrolase but not monoacyl glycerol lipase inhibition following peripheral nerve injury in rats

P.0296 Sex-dependent antinociceptive effects of fatty acid amide hydrolase but not monoacyl glycerol lipase inhibition following peripheral nerve injury in rats

The Endocannabinoid System as a Pharmacological Target for New Cancer Therapies.

Simple SummaryCannabinoids have been shown to suppress tumour cell proliferation, tumour invasion, metastasis, angiogenesis, chemoresistance and epithelial-mesenchymal transition and to induce tumour cell apoptosis, autophagy and immune response. This review focuses on the current status of investigations on the impact of inhibitors of endocannabinoid-degrading enzymes on tumour growth and spread in preclinical oncology research.Despite the long history of cannabinoid use for medicinal and ritual purposes, an endogenous system of cannabinoid-controlled receptors, as well as their ligands and the enzymes that synthesise and degrade them, was only discovered in the 1990s. Since then, the endocannabinoid system has attracted widespread scientific interest regarding new pharmacological targets in cancer treatment among other reasons. Meanwhile, extensive preclinical studies have shown that cannabinoids have an inhibitory effect on tumour cell proliferation, tumour invasion, metastasis, angiogenesis, chemoresistance and epithelial-mesenchymal transition (EMT) and induce tumour cell apoptosis and autophagy as well as immune response. Appropriate cannabinoid compounds could moreover be useful for cancer patients as potential combination partners with other chemotherapeutic agents to increase their efficacy while reducing unwanted side effects. In addition to the direct activation of cannabinoid receptors through the exogenous application of corresponding agonists, another strategy is to activate these receptors by increasing the endocannabinoid levels at the corresponding pathological hotspots. Indeed, a number of studies accordingly showed an inhibitory effect of blockers of the endocannabinoid-degrading enzymes fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) on tumour development and spread. This review summarises the relevant preclinical studies with FAAH and MAGL inhibitors compared to studies with cannabinoids and provides an overview of the regulation of the endocannabinoid system in cancer.

On the Biomedical Properties of Endocannabinoid Degradation and Reuptake Inhibitors: Pre-clinical and Clinical Evidence.

The endocannabinoid system (ECS) is composed of endogenous cannabinoids; components involved in their synthesis, transport, and degradation; and an expansive variety of cannabinoid receptors. Hypofunction or deregulation of the ECS is related to pathological conditions. Consequently, endogenous enhancement of endocannabinoid levels and/or regulation of their metabolism represent promising therapeutic approaches. Several major strategies have been suggested for the modulation of the ECS: (1) blocking endocannabinoids degradation, (2) inhibition of endocannabinoid cellular uptake, and (3) pharmacological modulation of cannabinoid receptors as potential therapeutic targets. Here, we focused in this review on degradation/reuptake inhibitors over cannabinoid receptor modulators in order to provide an updated synopsis of contemporary evidence advancing mechanisms of endocannabinoids as pharmacological tools with therapeutic properties for the treatment of several disorders. For this purpose, we revisited the available literature and reported the latest advances regarding the biomedical properties of fatty acid amide hydrolase and monoacylglycerol lipase inhibitors in pre-clinical and clinical studies. We also highlighted anandamide and 2-arachidonoylglycerol reuptake inhibitors with promising results in pre-clinical studies using in vitro and animal models as an outlook for future research in clinical trials.

Monoacylglycerol Lipase Inhibition: A Strategy to Treat Chronic Pain in a Humanized Sickle Cell Mouse Model

Introduction: Sickle Cell Disease (SCD) is a major cause of morbidity and mortality worldwide, and affects more than 100,000 people in the US. Acute, vaso-occlusive pain crises are the hallmark of the disease and the primary cause of hospitalization. Recurrent, acute pain episodes compound and result in chronic pain for many patients, though the mechanisms of this transition are poorly understood. Although opioids remain the standard of care to treat SCD chronic pain, their myriad adverse side effects (e.g., constipation, respiratory depression, abuse liability, dependence) as well as the fact that SCD chronic pain requires prolonged opioid treatment that results in tolerance, severely limit their therapeutic utility. Thus, a pressing need exists to identify effective non-opioid analgesic strategies to reduce SCD chronic pain. Humanized mouse models of SCD, such as the Berkeley (BERK) model, provide a useful tool to investigate disease pathophysiology and evaluate novel therapeutic strategies. Dorsal Root Ganglion neurons are peripheral sensory neurons essential for pain preclinical investigations. DRG neurons from rodents in neuropathic pain models show hyperexcitability. Inhibitors of the major degradative enzyme of 2-arachidonoylglycerol, monoacylglycerol lipase (MAGL), reduce nociceptive behavior in neuropathic and inflammatory preclinical models of pain through cannabinoid receptor-dependent and -independent mechanisms. MAGL inhibitors have yet to be tested in BERK mice and we hypothesize that they will ameliorate pain-like behavior and neuronal hyperexcitability. Here we evaluate the effects of MJN110 in a series of pain-related behaviors in BERK mice.Methods: Male and female HbSS-BERK (sickle) and HbAA-BERK (control) mice were used as subjects for these experiments. Pain-like behaviors were assessed using the Von Frey, Hot Plate, and Grip Strength tests. Neuronal hyperexcitability was assessed using whole cell patch clamp electrophysiology of L4-S1 dorsal root ganglia (DRG) neurons. For the dose-response experiments, MJN-110 (1.25, 2.5, 5, and 10 mg/kg, i.p.) was administered one hour prior to testing. Data were analyzed as Student's t-test, one- and two-way ANOVAs followed by Tukey or Sidak post-hoc analysis when appropriate (p < 0.05 considered significant).Results: HbSS-BERK mice displayed profound mechanical allodynia, as well as thermal and deep tissues/musculoskeletal hyperalgesia. HbSS-BERK mice possess extremely hyperexcitable DRG neurons. MJN-110 dose-dependently reduced mechanical allodynia and thermal hyperalgesia in HbSS-BERK mice. Importantly, BERK mice given seven days of daily injections of MJN-110 (5 mg/kg) displayed sustained antinociception that did not undergo tolerance. Ongoing studies are aimed to determine whether MJN110 ameliorates the hyperexcitability of DRG neurons.Conclusion: These initial findings, not only validate that BERK mice show hypersensitive responses to mechanical and heat stimuli, but also demonstrate that neurons innervating the distal parts of BERK mice are hyperexcitable. Additionally, MJN110 reduces these hyper-nociceptive behaviors suggesting that MAGL inhibition represents a viable strategy to reduce chronic pain behaviors in the BERK mouse model. DisclosuresGupta: Tautona Group: Consultancy, Honoraria; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; 1910 Genetics: Other: Grantee; Grifols: Other: Grantee; SCDAA: Membership on an entity's Board of Directors or advisory committees; CSL Behring LLC: Honoraria; NIH: Other: Grantee; University of Minnestoa Foundation: Other: Philanthropic Funding; Southern California Institute for Research and Education Foundation: Other: Philanthropic Funding; Cyclerion: Research Funding; UCI Foundation: Other: Philanthropic Funding. Smith: Emmaus Pharmaceuticals: Consultancy; Global Blood Therapeutics: Consultancy, Honoraria, Research Funding, Speakers Bureau; Novartis Pharmaceuticsl: Consultancy, Honoraria; Novo-Nordisk: Other: DSMB; Pfizer: Consultancy, Research Funding; Imara: Research Funding; Forma Therapeutics: Consultancy, Research Funding; Agios: Research Funding.

Discovery of Isatin-Based Carbohydrazones as Potential Dual Inhibitors of Fatty Acid Amide Hydrolase and Monoacylglycerol Lipase.

Using ligand-based design strategy, a set of isatin-3-carbohydrazones was designed, synthesized and evaluated for dual fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) inhibition properties. Compound 5-chloro-N'-(5-chloro-2-oxoindolin-3-ylidene)-2-hydroxybenzohydrazide (13 b) emerged as a potent MAGL inhibitor with nanomolar activity (IC50 =3.33 nM), while compound 5-chloro-N'-(1-(4-fluorobenzyl)-2-oxoindolin-3-ylidene)-2-hydroxybenzohydrazide (13 j) was the most potent selective FAAH inhibitor (IC50 =37 nM). Compound 5-chloro-N'-(6-chloro-2-oxoindolin-3-ylidene)-2-hydroxybenzohydrazide (13 c) showed dual FAAH-MAGL inhibitory activity with an IC50 of 31 and 29 nM respectively. Enzyme kinetics studies revealed that the isatin-based carbohydrazones are reversible inhibitors for both FAAH and MAGL. Further, blood-brain permeability assay confirmed that the lead compounds (13 b, 13 c, 13 g, 13 m and 13 q) are suitable as CNS candidates. Molecular dynamics simulation studies revealed the putative binding modes and key interactions of lead inhibitors within the enzyme active sites. The lead dual FAAH-MAGL inhibitor 13 c showed significant antioxidant activity and neuroprotection in the cell-based cytotoxicity assay. In summary, the study yielded three potent FAAH/MAGL inhibitor compounds (13 b, 13 c and 13 j) with acceptable pharmacokinetic profile and thus can be considered as promising candidates for treating neurological and mood disorders.

Monoacylglycerol lipase (MAGL) inhibitors based on a diphenylsulfide-benzoylpiperidine scaffold.

Monoacylglycerol lipase (MAGL) is an enzyme belonging to the endocannabinoid system that mainly metabolizes the endocannabinoid 2-arachidonoylglycerol (2-AG). Numerous studies have shown the involvement of this enzyme in various pathological conditions such as pain, cancer progression, Parkinson's and Alzheimer's disease, thus encouraging the development of new MAGL modulators. In this context, we developed new diphenylsulfide-benzoylpiperidine derivatives characterized by a high enzymatic MAGL inhibition activity in the low nanomolar range, a reversible mechanism of action and selectivity. The three most active compounds (15-17) induced an appreciable inhibition of cell viability in a panel of nine cancer cell lines, with IC50 values ranging between 0.32 and 10μM, thus highlighting their potential as novel anticancer agents.

ABHD6 Controls Amphetamine-Stimulated Hyperlocomotion: Involvement of CB1 Receptors.

Introduction: Activation of cannabinoid 1 receptors (CB1Rs) by endocannabinoids (eCBs) is controlled by both eCB production and eCB inactivation. Accordingly, inhibition of eCB hydrolyzing enzymes, monoacylglycerol lipase (MAGL) and α/β-hydrolase domain containing 6 (ABHD6), enhances eCB accumulation and CB1R activation. It is known that inhibition of MAGL regulates select CB1R-dependent behaviors in mice, including locomotor behaviors and their modulation by psychostimulants, but much less is known about the effect of inhibiting ABHD6 activity on such behaviors. Methods: We report a new mouse line that carries a genetic deletion of Abhd6 and evaluated its effect on spontaneous locomotion measured in a home cage monitoring system, motor coordination measured on a Rotarod, and amphetamine-stimulated hyperlocomotion and amphetamine sensitization (AS) measured in an open-field chamber. Results: ABHD6 knockout (KO) mice reached adulthood without exhibiting overt behavioral impairment, and we measured only mild reduction in spontaneous locomotion and motor coordination in adult ABHD6 KO mice compared to wild-type (WT) mice. Significantly, amphetamine-stimulated hyperlocomotion was enhanced by twofold in ABHD6 KO mice compared to WT mice and yet ABHD6 KO mice expressed AS to the same extent as WT mice. A twofold increase in amphetamine-stimulated hyperlocomotion was also measured in ABHD6 heterozygote mice and in WT mice treated with the ABHD6 inhibitor KT-182. It is known that amphetamine-stimulated hyperlocomotion is not affected by the CB1R antagonist, SR141617, and we discovered that the enhanced amphetamine-stimulated hyperlocomotion resulting from ABHD6 inhibition is blocked by SR141617. Conclusions: Our study suggests that ABHD6 controls amphetamine-stimulated hyperlocomotion by a mechanistic switch to a CB1R-dependent mechanism.

Increased 2-arachidonoyl-sn-glycerol levels normalize cortical responses to sound and improve behaviors in Fmr1 KO mice

BackgroundIndividuals with Fragile X syndrome (FXS) and autism spectrum disorder (ASD) exhibit an array of symptoms, including sociability deficits, increased anxiety, hyperactivity, and sensory hyperexcitability. It is unclear how endocannabinoid (eCB) modulation can be targeted to alleviate neurophysiological abnormalities in FXS as behavioral research reveals benefits to inhibiting cannabinoid (CB) receptor activation and increasing endocannabinoid ligand levels. Here, we hypothesize that enhancement of 2-arachidonoyl-sn-glycerol (2-AG) in Fragile X mental retardation 1 gene knock-out (Fmr1 KO) mice may reduce cortical hyperexcitability and behavioral abnormalities observed in FXS.MethodsTo test whether an increase in 2-AG levels normalized cortical responses in a mouse model of FXS, animals were subjected to electroencephalography (EEG) recording and behavioral assessment following treatment with JZL-184, an irreversible inhibitor of monoacylglycerol lipase (MAGL). Assessment of 2-AG was performed using lipidomic analysis in conjunction with various doses and time points post-administration of JZL-184. Baseline electrocortical activity and evoked responses to sound stimuli were measured using a 30-channel multielectrode array (MEA) in adult male mice before, 4 h, and 1 day post-intraperitoneal injection of JZL-184 or vehicle. Behavior assessment was done using the open field and elevated plus maze 4 h post-treatment.ResultsLipidomic analysis showed that 8 mg/kg JZL-184 significantly increased the levels of 2-AG in the auditory cortex of both Fmr1 KO and WT mice 4 h post-treatment compared to vehicle controls. EEG recordings revealed a reduction in the abnormally enhanced baseline gamma-band power in Fmr1 KO mice and significantly improved evoked synchronization to auditory stimuli in the gamma-band range post-JZL-184 treatment. JZL-184 treatment also ameliorated anxiety-like and hyperactivity phenotypes in Fmr1 KO mice.ConclusionsOverall, these results indicate that increasing 2-AG levels may serve as a potential therapeutic approach to normalize cortical responses and improve behavioral outcomes in FXS and possibly other ASDs.

Novel Reversible-Binding PET Ligands for Imaging Monoacylglycerol Lipase Based on the Piperazinyl Azetidine Scaffold.

Monoacylglycerol lipase (MAGL) is a 33 kDa serine protease primarily responsible for hydrolyzing 2-arachidonoylglycerol into the proinflammatory eicosanoid precursor arachidonic acid in the central nervous system. Inhibition of MAGL constitutes an attractive therapeutic concept for treating psychiatric disorders and neurodegenerative diseases. Herein, we present the design and synthesis of multiple reversible MAGL inhibitor candidates based on a piperazinyl azetidine scaffold. Compounds 10 and 15 were identified as the best-performing reversible MAGL inhibitors by pharmacological evaluations, thus channeling their radiolabeling with fluorine-18 in high radiochemical yields and favorable molar activity. Furthermore, evaluation of [18F]10 and [18F]15 ([18F]MAGL-2102) by autoradiography and positron emission tomography (PET) imaging in rodents and nonhuman primates demonstrated favorable brain uptakes, heterogeneous radioactivity distribution, good specific binding, and adequate brain kinetics, and [18F]15 demonstrated a better performance. In conclusion, [18F]15 was found to be a suitable PET radioligand for the visualization of MAGL, harboring potential for the successful translation into humans.

Targeting Monoacylglycerol Lipase in Pursuit of Therapies for Neurological and Neurodegenerative Diseases.

Neurological and neurodegenerative diseases are debilitating conditions, and frequently lack an effective treatment. Monoacylglycerol lipase (MAGL) is a key enzyme involved in the metabolism of 2-AG (2-arachidonoylglycerol), a neuroprotective endocannabinoid intimately linked to the generation of pro- and anti-inflammatory molecules. Consequently, synthesizing selective MAGL inhibitors has become a focus point in drug design and development. The purpose of this review was to summarize the diverse synthetic scaffolds of MAGL inhibitors concerning their potency, mechanisms of action and potential therapeutic applications, focusing on the results of studies published in the past five years. The main irreversible inhibitors identified were derivatives of hexafluoroisopropyl alcohol carbamates, glycol carbamates, azetidone triazole ureas and benzisothiazolinone, whereas the most promising reversible inhibitors were derivatives of salicylketoxime, piperidine, pyrrolidone and azetidinyl amides. We reviewed the results of in-depth chemical, mechanistic and computational studies on MAGL inhibitors, in addition to the results of in vitro findings concerning selectivity and potency of inhibitors, using the half maximal inhibitory concentration (IC50) as an indicator of their effect on MAGL. Further, for highlighting the potential usefulness of highly selective and effective inhibitors, we examined the preclinical in vivo reports regarding the promising therapeutic applications of MAGL pharmacological inhibition.

The effects of systemic and local fatty acid amide hydrolase and monoacylglycerol lipase inhibitor treatments on the metabolomic profile of lungs.

The contribution of the endocannabinoid system to both physiology and pathological processes in the respiratory system makes it a promising target for inflammatory airway diseases. Previously, we have shown that increasing the tissue endocannabinoid levels by fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) inhibitors can prevent airway inflammation and hyperreactivity. In this study, the changes in the levels of major metabolites of endocannabinoids by systemic and local FAAH or MAGL inhibitor treatments were evaluated. Mice were treated with either the FAAH inhibitor URB597 or the MAGL inhibitor JZL184 by local (intranasal) or systemic (intraperitoneal) application. Bronchoalveolar lavage (BAL) fluids and lungs were isolated afterward in order to perform histopathological and metabolomic analyses. There were no significant histopathological changes in the lungs and neutrophil, and macrophage and lymphocyte numbers in BAL fluid were not altered after local and systemic treatments. However, GC-MS-based metabolomics profile allowed us to identify 102 metabolites in lung samples, among which levels of 75 metabolites were significantly different from the control. The metabolites whose levels were changed by treatments were mostly related to the endocannabinoid system and energy metabolism. Therefore, these changes may contribute to the anti-inflammatory effects of URB597 and JZL184 treatments in mice.

Oxidative Stress and Multi-Organel Damage Induced by Two Novel Phytocannabinoids, CBDB and CBDP, in Breast Cancer Cells.

Over the last few years, much attention has been paid to phytocannabinoids derived from Cannabis for their therapeutic potential. Δ9-tetrahydrocannabinol (Δ9-THC) and cannabidiol (CBD) are the most abundant compounds of the Cannabis sativa L. plant. Recently, novel phytocannabinoids, such as cannabidibutol (CBDB) and cannabidiphorol (CBDP), have been discovered. These new molecules exhibit the same terpenophenolic core of CBD and differ only for the length of the alkyl side chain. Roles of CBD homologs in physiological and pathological processes are emerging but the exact molecular mechanisms remain to be fully elucidated. Here, we investigated the biological effects of the newly discovered CBDB or CBDP, compared to the well-known natural and synthetic CBD (nat CBD and syn CBD) in human breast carcinoma cells that express CB receptors. In detail, our data demonstrated that the treatment of cells with the novel phytocannabinoids affects cell viability, increases the production of reactive oxygen species (ROS) and activates cellular pathways related to ROS signaling, as already demonstrated for natural CBD. Moreover, we observed that the biological activity is significantly increased upon combining CBD homologs with drugs that inhibit the activity of enzymes involved in the metabolism of endocannabinoids, such as the monoacylglycerol lipase (MAGL) inhibitor, or with drugs that induces the activation of cellular stress pathways, such as the phorbol ester 12-myristate 13-acetate (PMA).

Monoacylglycerol Lipase Inhibitor MJN110 Reduces Neuronal Hyperexcitability, Restores Dendritic Arborization Complexity, and Regulates Reward-Related Behavior in Presence of HIV-1 Tat.

While current therapeutic strategies for people living with human immunodeficiency virus type 1 (HIV-1) suppress virus replication peripherally, viral proteins such as transactivator of transcription (Tat) enter the central nervous system early upon infection and contribute to chronic inflammatory conditions even alongside antiretroviral treatment. As demand grows for supplemental strategies to combat virus-associated pathology presenting frequently as HIV-associated neurocognitive disorders (HAND), the present study aimed to characterize the potential utility of inhibiting monoacylglycerol lipase (MAGL) activity to increase inhibitory activity at cannabinoid receptor-type 1 receptors through upregulation of 2-arachidonoylglycerol (2-AG) and downregulation of its degradation into proinflammatory metabolite arachidonic acid (AA). The MAGL inhibitor MJN110 significantly reduced intracellular calcium and increased dendritic branching complexity in Tat-treated primary frontal cortex neuron cultures. Chronic MJN110 administration in vivo increased 2-AG levels in the prefrontal cortex (PFC) and striatum across Tat(+) and Tat(–) groups and restored PFC N-arachidonoylethanolamine (AEA) levels in Tat(+) subjects. While Tat expression significantly increased rate of reward-related behavioral task acquisition in a novel discriminative stimulus learning and cognitive flexibility assay, MJN110 altered reversal acquisition specifically in Tat(+) mice to rates indistinguishable from Tat(–) controls. Collectively, our results suggest a neuroprotective role of MAGL inhibition in reducing neuronal hyperexcitability, restoring dendritic arborization complexity, and mitigating neurocognitive alterations driven by viral proteins associated with latent HIV-1 infection.

Early suppression of the endocannabinoid degrading enzymes FAAH and MAGL alters locomotor development in zebrafish.

The fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) enzymes are the predominant catabolic regulators of the major endocannabinoids (eCBs) anadamide (AEA) and 2-arachidonoylglycerol (2-AG), respectively. The expression and roles of eCBs during early embryogenesis remain to be fully investigated. Here, we inhibited FAAH and MAGL in zebrafish embryos during the first 24 h of life and examined motor neuron and locomotor development at 2 and 5 days post fertilization (dpf). Application of the dual FAAH/MAGL inhibitor, JZL195 (2 µmoll-1), resulted in a reduction in primary and secondary motor neuron axonal branching. JZL195 also reduced nicotinic acetylcholine receptor (nAChR) expression at neuromuscular junctions. Application of URB597 (5 µmoll-1), a specific inhibitor of the FAAH enzyme, also decreased primary motor neuron branching but did not affect secondary motor neuron branching and nAChR expression. Interestingly, JZL184 (5 µmoll-1), a specific inhibitor of MAGL, showed no effects on motor neuron branching or nAChR expression. Co-treatment of the enzyme inhibitors with the CB1R inhibitor AM251 confirmed the involvement of CB1R in motor neuron branching. Disruption of FAAH or MAGL reduced larval swimming activity, and AM251 attenuated the JZL195- and URB597-induced locomotor changes, but not the effects of JZL184. Together, these findings indicate that inhibition of FAAH, or augmentation of AEA acting through CB1R during early development, may be responsible for locomotor deficiencies.