Identification of Novel Antagonists Targeting Cannabinoid Receptor 2 Using a Multi-Step Virtual Screening Strategy

The endocannabinoid system plays an essential role in the regulation of analgesia and human immunity, and Cannabinoid Receptor 2 (CB2) has been proved to be an ideal target for the treatment of liver diseases and some cancers. In this study, we identified CB2 antagonists using a three-step “deep learning–pharmacophore–molecular docking” virtual screening approach. From the ChemDiv database (1,178,506 compounds), 15 hits were selected and tested by radioligand binding assays and cAMP functional assays. A total of 7 out of the 15 hits were found to exhibit binding affinities in the radioligand binding assays against CB2 receptor, with a pKi of 5.15-6.66, among which five compounds showed antagonistic activities with pIC50 of 5.25–6.93 in the cAMP functional assays. Among these hits, Compound 8 with the 4H-pyrido[1,2-a]pyrimidin-4-one scaffold showed the best binding affinity and antagonistic activity with a pKi of 6.66 and pIC50 of 6.93, respectively. The new scaffold could serve as a lead for further development of CB2 drugs. Additionally, we hope that the model in this study could be further utilized to identify more novel CB2 receptor antagonists, and the developed approach could also be used to design potent ligands for other therapeutic targets.

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Expression of the endocannabinoid receptors in human fascial tissue

European Journal of Histochemistry ◽

10.4081/ejh.2016.2643 ◽

2016 ◽

Cited By ~ 5

Author(s):

C. Fede ◽

G. Albertin ◽

L. Petrelli ◽

M.M. Sfriso ◽

C. Biz ◽

...

Keyword(s):

Cannabinoid Receptors ◽

Cannabinoid Receptor ◽

Endocannabinoid System ◽

Trigger Points ◽

Small Samples ◽

Myofascial Trigger Points ◽

Cannabinoid Receptor 2 ◽

Culture Cells ◽

Cb1 Cannabinoid Receptor ◽

A Cell

Cannabinoid receptors have been localized in the central and peripheral nervous system as well as on cells of the immune system, but recent studies on animal tissue gave evidence for the presence of cannabinoid receptors in different types of tissues. Their presence was supposed also in myofascial tissue, suggesting that the endocannabinoid system may help resolve myofascial trigger points and relieve symptoms of fibromyalgia. However, until now the expression of CB1 (cannabinoid receptor 1) and CB2 (cannabinoid receptor 2) in fasciae has not yet been established. Small samples of fascia were collected from volunteers patients during orthopedic surgery. For each sample were done a cell isolation, immunohistochemical investigation (CB1 and CB2 antibodies) and real time RT-PCR to detect the expression of CB1 and CB2. Both cannabinoid receptors are expressed in human fascia and in human fascial fibroblasts culture cells, although to a lesser extent than the control gene. We can assume that the expression of mRNA and protein of CB1 and CB2 receptors in fascial tissue are concentrated into the fibroblasts.Â This is the first demonstration that the fibroblasts of the muscular fasciae express CB1 and CB2. The presence of these receptors could help to provide a description of cannabinoid receptors distribution and to better explain the role of fasciae as pain generator and the efficacy of some fascial treatments. Indeed the endocannabinoid receptors of fascial fibroblasts can contribute to modulate the fascial fibrosis and inflammation.

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Benzyl-1,2,4-triazoles as CB1 Cannabinoid Receptor Ligands: Preparation and In Vitro Pharmacological Evaluation

International Journal of Medicinal Chemistry ◽

10.1155/2016/1257098 ◽

2016 ◽

Vol 2016 ◽

pp. 1-9

Author(s):

Laura Hernandez-Folgado ◽

Juan Decara ◽

Fernando Rodríguez de Fonseca ◽

Pilar Goya ◽

Nadine Jagerovic

Keyword(s):

Cannabinoid Receptor ◽

Radioligand Binding ◽

Pharmacological Properties ◽

Binding Assays ◽

Receptor Ligands ◽

Cannabinoid Type 1 Receptor ◽

Cb1 Cannabinoid Receptor ◽

Nanomolar Range

In a previous study, we have identified 3-alkyl-1,5-diaryl-1H-1,2,4-triazoles to be a novel class of cannabinoid type 1 receptor (CB1R) antagonists. In order to expand the number of cannabinoid ligands with a central 1,2,4-triazole scaffold, we have synthesized a novel series of 1-benzyl-1H-1,2,4-triazoles, and some of them were evaluated by CB1R radioligand binding assays. Compound 12a showed the most interesting pharmacological properties, possessing a CB1R affinity in the nanomolar range.

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THC-induced behavioral stereotypy in zebrafish as a model of psychosis-like behavior

Scientific Reports ◽

10.1038/s41598-021-95016-4 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Amelia Dahlén ◽

Mahdi Zarei ◽

Adam Melgoza ◽

Mahendra Wagle ◽

Su Guo

Keyword(s):

Nmda Receptor ◽

Cannabinoid Receptor ◽

Repetitive Behavior ◽

Endocannabinoid System ◽

Adult Zebrafish ◽

Acute Effects ◽

Cannabinoid Receptor 1 ◽

Cannabinoid Receptor 2 ◽

High Doses ◽

Underlying Mechanisms

AbstractHigh doses of the Cannabis constituent Δ9-tetrahydrocannabinol (THC) increase the risk of psychosis in humans. Highly accessible animal models are needed to address underlying mechanisms. Using zebrafish with a conserved endocannabinoid system, this study investigates the acute effects of THC on adult zebrafish behavior and the mechanisms involved. A concentration-dependent THC-induced behavioral stereotypy akin to THC’s effect in rats and the psychotropics phencyclidine and ketamine in zebrafish was established. Distinctive circular swimming during THC-exposure was measured using a novel analytical method that we developed, which detected an elevated Repetition Index (RI) compared to vehicle controls. This was reduced upon co-administration of N-methyl-D-aspartate (NMDA) receptor agonist NMDA, suggesting that THC exerts its effects via biochemical or neurobiological mechanisms associated with NMDA receptor antagonism. Co-treatment of γ‐aminobutyric acid receptor antagonist pentylenetetrazol also showed signs of reducing the RI. Since THC-induced repetitive behavior remained in co-administrations with cannabinoid receptor 1 inverse agonist AM251, the phenotype may be cannabinoid receptor 1-independent. Conversely, the inverse cannabinoid receptor 2 agonist AM630 significantly reduced THC-induced behavioral stereotypy, indicating cannabinoid receptor 2 as a possible mediator. A significant reduction of the THC-RI was also observed by the antipsychotic sulpiride. Together, these findings highlight this model’s potential for elucidating the mechanistic relationship between Cannabis and psychosis.

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Cnr2 Is Important for Ribbon Synapse Maturation and Function in Hair Cells and Photoreceptors

Frontiers in Molecular Neuroscience ◽

10.3389/fnmol.2021.624265 ◽

2021 ◽

Vol 14 ◽

Author(s):

Luis Colón-Cruz ◽

Roberto Rodriguez-Morales ◽

Alexis Santana-Cruz ◽

Juan Cantres-Velez ◽

Aranza Torrado-Tapias ◽

...

Keyword(s):

Hair Cells ◽

Cannabinoid Receptor ◽

Sensory Information ◽

Endocannabinoid System ◽

Cannabinoid Receptor 2 ◽

Ribbon Synapses ◽

Sensory Epithelia ◽

Synapse Maturation ◽

And Function

The role of the cannabinoid receptor 2 (CNR2) is still poorly described in sensory epithelia. We found strong cnr2 expression in hair cells (HCs) of the inner ear and the lateral line (LL), a superficial sensory structure in fish. Next, we demonstrated that sensory synapses in HCs were severely perturbed in larvae lacking cnr2. Appearance and distribution of presynaptic ribbons and calcium channels (Cav1.3) were profoundly altered in mutant animals. Clustering of membrane-associated guanylate kinase (MAGUK) in post-synaptic densities (PSDs) was also heavily affected, suggesting a role for cnr2 for maintaining the sensory synapse. Furthermore, vesicular trafficking in HCs was strongly perturbed suggesting a retrograde action of the endocannabinoid system (ECs) via cnr2 that was modulating HC mechanotransduction. We found similar perturbations in retinal ribbon synapses. Finally, we showed that larval swimming behaviors after sound and light stimulations were significantly different in mutant animals. Thus, we propose that cnr2 is critical for the processing of sensory information in the developing larva.

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Cannabinoid Receptor 2 Alters Social Memory and Microglial Activity in an Age-Dependent Manner

Molecules ◽

10.3390/molecules26195984 ◽

2021 ◽

Vol 26 (19) ◽

pp. 5984

Author(s):

Joanna Agnieszka Komorowska-Müller ◽

Tanushka Rana ◽

Bolanle Fatimat Olabiyi ◽

Andreas Zimmer ◽

Anne-Caroline Schmöle

Keyword(s):

Cannabinoid Receptor ◽

Social Memory ◽

Brain Aging ◽

Endocannabinoid System ◽

Immunohistochemical Analysis ◽

Cellular Level ◽

Spatial Learning And Memory ◽

Dependent Manner ◽

Cannabinoid Receptor 2 ◽

Age Related

Physiological brain aging is characterized by gradual, substantial changes in cognitive ability, accompanied by chronic activation of the neural immune system. This form of inflammation, termed inflammaging, in the central nervous system is primarily enacted through microglia, the resident immune cells. The endocannabinoid system, and particularly the cannabinoid receptor 2 (CB2R), is a major regulator of the activity of microglia and is upregulated under inflammatory conditions. Here, we elucidated the role of the CB2R in physiological brain aging. We used CB2R−/− mice of progressive ages in a behavioral test battery to assess social and spatial learning and memory. This was followed by detailed immunohistochemical analysis of microglial activity and morphology, and of the expression of pro-inflammatory cytokines in the hippocampus. CB2R deletion decreased social memory in young mice, but did not affect spatial memory. In fact, old CB2R−/− mice had a slightly improved social memory, whereas in WT mice we detected an age-related cognitive decline. On a cellular level, CB2R deletion increased lipofuscin accumulation in microglia, but not in neurons. CB2R−/− microglia showed an increase of activity markers Iba1 and CD68, and minor upregulation in tnfa and il6 expression and downregulation of ccl2 with age. This was accompanied by a change in morphology as CB2R−/− microglia had smaller somas and lower polarity, with increased branching, cell volume, and tree length. We present that CB2Rs are involved in cognition and age-induced microglial activity, but may also be important for microglial activation itself.

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Discovery of Orexant and Anorexant Agents with Indazole Scaffold Endowed with Peripheral Antiedema Activity

Biomolecules ◽

10.3390/biom9090492 ◽

2019 ◽

Vol 9 (9) ◽

pp. 492 ◽

Cited By ~ 5

Author(s):

Dimmito ◽

Stefanucci ◽

Pieretti ◽

Minosi ◽

Dvorácskó ◽

...

Keyword(s):

Feeding Behavior ◽

Cannabinoid Receptors ◽

Cannabinoid Receptor ◽

Methyl Esters ◽

Endocannabinoid System ◽

Tail Flick ◽

Binding Assays ◽

Tail Flick Test

The endocannabinoid system represents an integrated neuronal network involved in the control of several organisms’ functions, such as feeding behavior. A series of hybrids of 5-(4-chlorophenyl)-1-(2,4-dichloro-phenyl)-4-methyl-N-(piperidin-1-yl)-1H-pyrazole-3-carboxamide (mimonabant), a well-known inverse agonist of the type-1 cannabinoid receptor (CB1), once used as an antiobesity drug, and the N-(2S)-substitutes of 1-[(4-fluorophenyl)methyl]indazole-3-carboxamide with 1-amino-3-methyl-1-oxobutane (AB-Fubinaca), 1-amino-3,3-dimethyl-1-oxobutane (ADB-Fubinaca), and 3-methylbutanoate (AMB-Fubinaca), endowed with potent agonistic activity towards cannabinoid receptors CB1 and CB2 were in solution as C-terminal amides, acids, methyl esters and N-methyl amides. These compounds have been studied by binding assays to cannabinoid receptors and by functional receptor assays, using rat brain membranes in vitro. The most active among them as an agonist, (S)-1-(2,4-dichlorobenzyl)-N-(3,3-dimethyl-1-(methylamino)-1-oxobutan-2-yl)-1H-indazole-3-carboxamide (LONI11), and an antagonist, (S)-2-(1-(2,4-dichlorobenzyl)-1H-indazole-3-carboxamido)-3-methylbutanoic acid (LONI4), were tested in vivo in mic, to evaluate their ability to stimulate or suppress feeding behavior after intraperitoneal (i.p.) administration. For a LONI11 formalin test and a tail flick test after an administration by the subcutaneous (s.c.) and intracerebroventricular (i.c.v.) routes, respectively, were also carried out in vivo in mice to investigate the antinociceptive property at the central and peripheral levesl. We observed a significant orexant effect for LONI11 and an intense anorexant effect for (S)-methyl 2-(1-(2,4-dichlorobenzyl)-1H-indazole-3-carboxamido)-3,3-dimethylbutanoate (LONI2) and LONI4. In zymosan-induced edema and hyperalgesia, LONI11 reduced the percent of paw volume increase and paw latency after s.c. administration, also suggesting a possible peripheral anti-inflammatory activity.

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Virtual Screening of C. Sativa Constituents for the Identification of Selective Ligands for Cannabinoid Receptor 2

International Journal of Molecular Sciences ◽

10.3390/ijms21155308 ◽

2020 ◽

Vol 21 (15) ◽

pp. 5308

Author(s):

Mikołaj Mizera ◽

Dorota Latek ◽

Judyta Cielecka-Piontek

Keyword(s):

Virtual Screening ◽

Cannabis Sativa ◽

Cannabinoid Receptor ◽

Data Bank ◽

Qsar Model ◽

Molecular Structures ◽

Computer Assisted ◽

Cannabinoid Receptor 2 ◽

Selective Ligands ◽

Qsar Models

The selective targeting of the cannabinoid receptor 2 (CB2) is crucial for the development of peripheral system-acting cannabinoid analgesics. This work aimed at computer-assisted identification of prospective CB2-selective compounds among the constituents of Cannabis Sativa. The molecular structures and corresponding binding affinities to CB1 and CB2 receptors were collected from ChEMBL. The molecular structures of Cannabis Sativa constituents were collected from a phytochemical database. The collected records were curated and applied for the development of quantitative structure-activity relationship (QSAR) models with a machine learning approach. The validated models predicted the affinities of Cannabis Sativa constituents. Four structures of CB2 were acquired from the Protein Data Bank (PDB) and the discriminatory ability of CB2-selective ligands and two sets of decoys were tested. We succeeded in developing the QSAR model by achieving Q2 5-CV > 0.62. The QSAR models helped to identify three prospective CB2-selective molecules that are dissimilar to already tested compounds. In a complementary structure-based virtual screening study that used available PDB structures of CB2, the agonist-bound, Cryogenic Electron Microscopy structure of CB2 showed the best statistical performance in discriminating between CB2-active and non-active ligands. The same structure also performed best in discriminating between CB2-selective ligands from non-selective ligands.

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The Endocannabinoid System and Alcohol Dependence: Will Cannabinoid Receptor 2 Agonism be More Fruitful than Cannabinoid Receptor 1 Antagonism?

CNS & Neurological Disorders - Drug Targets ◽

10.2174/1871527320666210211115007 ◽

2021 ◽

Vol 20 ◽

Author(s):

Aboagyewaah Oppong-Damoah ◽

Brenda Marie Gannon ◽

Kevin Sean Murnane

Keyword(s):

Cannabinoid Receptor ◽

Endocannabinoid System ◽

Health Concern ◽

Cb2 Receptor ◽

Neuroprotective Effects ◽

Receptor Agonists ◽

Clinical Phase ◽

Anti Inflammatory ◽

Cb1 Antagonists ◽

Psychiatric Side Effects

: Alcohol-use disorder (AUD) remains a major public health concern. In recent years, there has been a heightened interest in components of the endocannabinoid system for the treatment of AUD. Cannabinoid type 1 (CB1) receptors have been shown to modulate the rewarding effects of alcohol, reduce the abuse-related effects of alcohol, improve cognition, exhibit anti-inflammatory, and neuroprotective effects, which are all favorable properties of potential therapeutic candidates for the treatment of AUD. However, CB1 agonists have not been investigated for the treatment of AUD because they stimulate the motivational properties of alcohol, increase alcohol intake, and have the tendency to be abused. Preclinical data suggest significant potential for the use of CB1 antagonists to treat AUD; however, a clinical phase I/II trial with SR14716A (rimonabant), a CB1 receptor antagonist/inverse agonist showed that it produced serious neuropsychiatric adverse events such as anxiety, depression, and even suicidal ideation. This has redirected the field to focus on alternative components of the endocannabinoid system, including cannabinoid type 2 (CB2) receptor agonists as a potential therapeutic target for AUD. CB2 receptor agonists are of particular interest because they can modulate the reward pathway, reduce abuse-related effects of alcohol, reverse neuroinflammation, improve cognition, and exhibit anti-inflammatory and neuroprotective effects, without exhibiting the psychiatric side effects seen with CB1 antagonists. Accordingly, this article presents an overview of the studies reported in the literature that have investigated CB2 receptor agonists with regards to AUD and provides commentary as to whether this receptor is a worthy target for continued investigation.

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