Anti-inflammatory Properties of Cannabidiol, a Nonpsychotropic Cannabinoid, in Experimental Allergic Contact Dermatitis

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Synthesis and binding study of certain 6-arylalkanamides as molecular probes for cannabinoid receptor subtypes

Tetrahydrocannabinol and other mixed cannabinoid (CB) receptors CB(1)/CB(2) receptor agonists are well established to elicit antinociceptive effects and psychomimetic actions, however, their potential for abuse have dampened enthusiasm for their therapeutic development. In an effort to refine a semi-rigid structural framework for CB(2) receptors binding, we designed novel compounds based on aromatic moiety and flexible linker with various amides mimicking the outlook of the endogenous anandamide which could provide as CB(2) receptor ligand. In this direction, we developed and synthesized new aryl or arylidene hexanoic acid amides and aryl alkanoic acid diamide carrying different head groups. These new compounds were tested for their affinities for human recombinant CB receptors CB(1) and CB(2) and fatty acid amide hydrolase. Although, the preliminary screening of these compounds demonstrated weak binding activity towards CB receptor subtypes at 10 µmole, yet this template still could serve up as probes for further optimization and development of affinity ligand for CB receptors

N-[1,3-Dialkyl(aryl)-2-oxoimidazolidin-4-ylidene]-aryl(alkyl)sulphonamides as Novel Selective Human Cannabinoid Type 2 Receptor (hCB2R) Ligands; Insights into the Mechanism of Receptor Activation/Deactivation

Cannabinoid type 1 (hCB1) and type 2 (hCB2) receptors are pleiotropic and crucial targets whose signaling contributes to physiological homeostasis and its restoration after injury. Being predominantly expressed in peripheral tissues, hCB2R represents a safer therapeutic target than hCB1R, which is highly expressed in the brain, where it regulates processes related to cognition, memory, and motor control. The development of hCB2R ligands represents a therapeutic opportunity for treating diseases such as pain, inflammation and cancer. Identifying new selective scaffolds for cannabinoids and determining the structural determinants responsible for agonism and antagonism are priorities in drug design. In this work, a series of N-[1,3-dialkyl(aryl)-2-oxoimidazolidin-4-ylidene]-aryl(alkyl)sulfonamides is designed and synthesized and their affinity for human hCB1R and hCB2R is determined. Starting with a scaffold selected from the NIH Psychoactive Drug Screening Program Repository, through a combination of molecular modeling and structure–activity relationship studies, we were able to identify the chemical features leading to finely tuned hCB2R selectivity. In addition, an in silico model capable of predicting the functional activity of hCB2R ligands was proposed and validated. The proposed receptor activation/deactivation model enabled the identification of four pure hCB2R-selective agonists that can be used as a starting point for the development of more potent ligands

A novel fluorophosphonate inhibitor of the biosynthesis of the endocannabinoid 2-arachidonoylglycerol with potential anti-obesity effects

Background and Purpose The development of potent and selective inhibitors of the biosynthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG) via DAG lipases (DAGL) α and β is just starting to be considered as a novel and promising source of pharmaceuticals for the treatment of disorders that might benefit from a reduction in endocannabinoid tone, such as hyperphagia in obese subjects. Experimental Approach Three new fluorophosphonate compounds O-7458, O-7459 and O-7460 were synthesized and characterized in various enzymatic assays. The effects of O-7460 on high-fat diet intake were tested in mice. Key Results Of the new compounds, O-7460 exhibited the highest potency (IC50 = 690 nM) against the human recombinant DAGLα, and selectivity (IC50 > 10 μM) towards COS-7 cell and human monoacylglycerol lipase (MAGL), and rat brain fatty acid amide hydrolase. Competitive activity-based protein profiling confirmed that O-7460 inhibits mouse brain MAGL only at concentrations ≥10 μM, and showed that this compound has only one major ‘off-target’, that is, the serine hydrolase KIAA1363. O-7460 did not exhibit measurable affinity for human recombinant CB1 or CB2 cannabinoid receptors (Ki > 10 μM). In mouse neuroblastoma N18TG2 cells stimulated with ionomycin, O-7460 (10 μM) reduced 2-AG levels. When administered to mice, O-7460 dose-dependently (0–12 mg·kg−1, i.p.) inhibited the intake of a high-fat diet over a 14 h observation period, and, subsequently, slightly but significantly reduced body weight. Conclusions and Implications O-7460 might be considered a useful pharmacological tool to investigate further the role played by 2-AG both in vitro and in vivo under physiological as well as pathological conditions

Activity-based protein profiling reveals off-target proteins of the FAAH inhibitor BIA 10-2474

A recent phase 1 trial of the fatty acid amide hydrolase (FAAH) inhibitor BIA 10-2474 led to the death of one volunteer and produced mild-to-severe neurological symptoms in four others. Although the cause of the clinical neurotoxicity is unknown, it has been postulated, given the clinical safety profile of other tested FAAH inhibitors, that off-target activities of BIA 10-2474 may have played a role. Here we use activity-based proteomicmethods to determine the protein interaction landscape of BIA 10-2474 in human cells and tissues. This analysis revealed that the drug inhibits several lipases that are not targeted by PF04457845, a highly selective and clinically tested FAAH inhibitor. BIA 10-2474, but not PF04457845, produced substantial alterations in lipid networks in human cortical neurons, suggesting that promiscuous lipase inhibitors have the potential to cause metabolic dysregulation in the nervous system

2-Pentadecyl-2-Oxazoline, the Oxazoline of Pea, Modulates Carrageenan-Induced Acute Inflammation

N-acylethanolamines (NAEs) involve a family of lipid molecules existent in animal and plant, with N-palmitoylethanolamide (PEA) that arouses great attention owing to its anti-inflammatory, analgesic and neuroprotective activities. Because PEA is produced on demand and exerts pleiotropic effects, the modulation of specific amidases for NAEs (and in particular NAE-hydrolyzing acid amidase NAAA, which is more selective for PEA) could be a condition to preserve its levels. Here we investigate the effect of 2-Pentadecyl-2-oxazoline (PEA-OXA) the oxazoline of PEA, on human recombinant NAAA in vitro and in an established model of Carrageenan (CAR)-induced rat paw inflammation. PEA-OXA dose-dependently significantly inhibited recombinant NAAA and, orally administered to rats (10 mg/kg), limiting histological damage, thermal hyperalgesia and the increase of infiltrating inflammatory cells after CAR injection in the rat right hindpaw, compared to ultramicronized PEA given orally at the same dose (10 mg/kg). These effects were accompanied by elevation of paw PEA levels. Moreover, PEA-OXA markedly reduced neutrophil infiltration and pro-inflammatory cytokine release and prevented CAR-induced IκB-α degradation, nuclear translocation of NF-κB p65, the increase of inducible nitric oxide synthase, cyclooxygenase-2, intercellular adhesion molecule-1, and mast cell activation. Experiments in PPAR-α knockout mice showed that the anti-inflammatory effects of PEA-OXA were not dependent on the presence of PPAR-α receptors. In conclusion, NAAA modulators as PEA-OXA could help to maximize the tissue availability of PEA by increasing its levels and anti-inflammatory effects

Synthesis and evaluation of new pyrazole-based carboxamides as modulators of the TrPv1 channel

Since the cloning of the vanilloid receptor protein (TRPV1) by Caterina et al.,[1] this ligand gated nonselective cation channel has attracted growing interest due to its function associated with nociception signaling pathway. TRPV1 is activated by vanilloid ligands such as capsaicin and resiniferatoxin, noxious heat (> 42 °C), protons (extracellular pH < 6), and modulated by a variety of endogenous ligands including cannabinoid anandamide and arachidonic acid metabolites. TRPV1 activation enhances Ca2+ permeability that leads to an increase in intracellular Ca2+, resulting in excitation of primary sensory neurons and the central perception of pain. Activation is followed by the desensitization making the C-fiber sensory neurons unresponsive to TRPV1 agonists and other inflammatory mediators. In addition, capsazepine, the first reported antagonist of TRPV1, blocks the capsaicin-induced uptake of Ca2+ in cell-based assays. In vivo, capsazepine demonstrates species-dependent efficacies in various models of inflammatory hyperalgesia and chronic pain. [2] Therefore, both desensitization by an agonist and the direct blockade by an antagonist of TRPV1 might have therapeutic utility in the management of acute and chronic nociceptive pain. [3] Recently, a variety of TRPV1 ligands bearing a heterocyclic linker has been reported to show very interesting both in vitro and in vivo biological profiles. Based on these findings, we designed a series of pyrazole-based carboxamide analogues (Figure 1), assuming that the rigid heterocyclic linkage can induce more appropriate conformation for TRPV1 binding. figure 1 In this communication, we describe the synthesis and activities of a series of these ligands, which were screened by a cell-based assay utilizing the Ca2+ permeability of the TRPV1 channel. references 1. Caterina, M. J.; Schumacher, M. A.; Tominaga, M.; Rosen, T. A.; Levine, J. D.; Julius, D. Nature 1997, 387, 816–824. 2. Walker, K. M.; Urban, L.; Medhurst, S. J.; Patel, S.; Panesar, M.; Fox, A. J. J. Pharmacol. Exp. Ther. 2003, 304, 56-62. 3. De Petrocellis, L.; Schiano-Moriello A. Recent Pat. CNS Drug Discov. 2013, 8, 180-204 Synthesis and evaluation of new pyrazole-based carboxamides as modulators of the TRPV1 channel Cenzo Congiu,1* Valentina Onnis,1 Marco Allarà,2 Aniello Schiano-Moriello,2 Vincenzo Di Marzo,2 and Luciano De Petrocellis.2 1University of Cagliari, Department of Life and Environment Sciences, Via Ospedale 72, I-09124 – Cagliari, Italy; 2Endocannabinoid Research Group (ERG), Institute of Biomolecular Chemistry, Consiglio Nazionale delle Ricerche, Via Campi Flegrei 34, I-80078-Pozzuoli (NA), Italy Since the cloning of the vanilloid receptor protein (TRPV1) by Caterina et al.,[1] this ligand gated nonselective cation channel has attracted growing interest due to its function associated with nociception signaling pathway. TRPV1 is activated by vanilloid ligands such as capsaicin and resiniferatoxin, noxious heat (> 42 °C), protons (extracellular pH < 6), and modulated by a variety of endogenous ligands including cannabinoid anandamide and arachidonic acid metabolites. TRPV1 activation enhances Ca2+ permeability that leads to an increase in intracellular Ca2+, resulting in excitation of primary sensory neurons and the central perception of pain. Activation is followed by the desensitization making the C-fiber sensory neurons unresponsive to TRPV1 agonists and other inflammatory mediators. In addition, capsazepine, the first reported antagonist of TRPV1, blocks the capsaicin-induced uptake of Ca2+ in cell-based assays. In vivo, capsazepine demonstrates species-dependent efficacies in various models of inflammatory hyperalgesia and chronic pain.[2] Therefore, both desensitization by an agonist and the direct blockade by an antagonist of TRPV1 might have therapeutic utility in the management of acute and chronic nociceptive pain.[3] Recently, a variety of TRPV1 ligands bearing a heterocyclic linker has been reported to show very interesting both in vitro and in vivo biological profiles. Based on these findings, we designed a series of pyrazole-based carboxamide analogues (Figure 1), assuming that the rigid heterocyclic linkage can induce more appropriate conformation fo

Investigations on the 4-Quinolone-3-carboxylic Acid Motif. 7. Synthesis and Pharmacological Evaluation of 4-Quinolone-3-carboxamides and 4-Hydroxy-2-quinolone-3-carboxamides as High Affinity Cannabinoid Receptor 2 (CB2R) Ligands with Improved Aqueous Solubility

4-Quinolone-3-carboxamide derivatives have long been recognized as potent and selective cannabinoid type-2 receptor (CB2R) ligands. With the aim to improve their physicochemical properties, basically aqueous solubility, two different approaches were followed, entailing the substitution of the alkyl chain with a basic replacement or scaffold modification to 4-hydroxy-2-quinolone structure. According to the first approach, compound 6d was obtained, showing slightly reduced receptor affinity (Ki = 60 nM) compared to the lead compound 4 (0.8 nM) but greatly enhanced solubility (400-3400 times depending on the pH of the medium). On the other hand, shifting from 4-quinolone to 4-hydroxy-2-quinolone structure enabled the discovery of a novel class of CB2R ligands, such as 7b and 7c, characterized by Ki 1300. At pH 7.4, compound 7c resulted by 100-fold more soluble than 4

Prostamide F2α receptor antagonism combined with inhibition of FAAH may block the pro-inflammatory mediators formed following selective FAAH inhibition

Background and Purpose Prostamides are lipid mediators formed by COX-2-catalysed oxidation of the endocannabinoid anandamide and eliciting effects often opposed to those caused by anandamide. Prostamides may be formed when hydrolysis of anandamide by fatty acid amide hydrolase (FAAH) is physiologically, pathologically or pharmacologically decreased. Thus, therapeutic benefits of FAAH inhibitors might be attenuated by concomitant production of prostamide F2α. This loss of benefit might be minimized by compounds designed to selectively antagonize prostamide receptors and also inhibiting FAAH. Experimental Approach Inhibition of FAAH by a series of selective antagonists of prostamide receptors, including AGN 204396, AGN 211335 and AGN 211336, was assessed using rat, mouse and human FAAH in vitro, together with affinity for human recombinant CB1 and CB 2 receptors. Effects in vivo were measured in a model of formalin-induced inflammatory pain in mice. Key Results The prostamide F 2α receptor antagonists were active against mouse and rat FAAH in the low μM range and behaved as non-competitive and plasma membrane-permeant inhibitors. AGN 211335, the most potent inhibitor of rat FAAH (IC50 = 1.2 μM), raised exogenous anandamide levels in intact cells and also bound to cannabinoid CB1 receptors. Both AGN 211335 and AGN 211336 (0.25-1 mg·kg-1, i.p.) inhibited the formalin-induced nociceptive response in mice. Conclusions and Implications Synthetic compounds with indirect agonist activity at cannabinoid receptors and antagonist activity at prostamide receptors can be developed. Such compounds could be used as alternatives to selective FAAH inhibitors to prevent the possibility of prostamide F2α-induced inflammation and pain. Linked Articles This article is part of a themed section on Cannabinoids 2013. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2014.171. © 2013 The British Pharmacological Society.Background and Purpose Prostamides are lipid mediators formed by COX-2-catalysed oxidation of the endocannabinoid anandamide and eliciting effects often opposed to those caused by anandamide. Prostamides may be formed when hydrolysis of anandamide by fatty acid amide hydrolase (FAAH) is physiologically, pathologically or pharmacologically decreased. Thus, therapeutic benefits of FAAH inhibitors might be attenuated by concomitant production of prostamide F2α. This loss of benefit might be minimized by compounds designed to selectively antagonize prostamide receptors and also inhibiting FAAH. Experimental Approach Inhibition of FAAH by a series of selective antagonists of prostamide receptors, including AGN 204396, AGN 211335 and AGN 211336, was assessed using rat, mouse and human FAAH in vitro, together with affinity for human recombinant CB1 and CB 2 receptors. Effects in vivo were measured in a model of formalin-induced inflammatory pain in mice. Key Results The prostamide F 2α receptor antagonists were active against mouse and rat FAAH in the low μM range and behaved as non-competitive and plasma membrane-permeant inhibitors. AGN 211335, the most potent inhibitor of rat FAAH (IC50 = 1.2 μM), raised exogenous anandamide levels in intact cells and also bound to cannabinoid CB1 receptors. Both AGN 211335 and AGN 211336 (0.25-1 mg·kg-1, i.p.) inhibited the formalin-induced nociceptive response in mice. Conclusions and Implications Synthetic compounds with indirect agonist activity at cannabinoid receptors and antagonist activity at prostamide receptors can be developed. Such compounds could be used as alternatives to selective FAAH inhibitors to prevent the possibility of prostamide F2α-induced inflammation and pain. Linked Articles This article is part of a themed section on Cannabinoids 2013. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2014.171. © 2013 The British Pharmacological Society

A Two-Step Feature Selection Radiomic Approach to Predict Molecular Outcomes in Breast Cancer

Breast Cancer (BC) is the most common cancer among women worldwide and is characterized by intra- and inter-tumor heterogeneity that strongly contributes towards its poor prognosis. The Estrogen Receptor (ER), Progesterone Receptor (PR), Human Epidermal Growth Factor Receptor 2 (HER2), and Ki67 antigen are the most examined markers depicting BC heterogeneity and have been shown to have a strong impact on BC prognosis. Radiomics can noninvasively predict BC heterogeneity through the quantitative evaluation of medical images, such as Magnetic Resonance Imaging (MRI), which has become increasingly important in the detection and characterization of BC. However, the lack of comprehensive BC datasets in terms of molecular outcomes and MRI modalities, and the absence of a general methodology to build and compare feature selection approaches and predictive models, limit the routine use of radiomics in the BC clinical practice. In this work, a new radiomic approach based on a two-step feature selection process was proposed to build predictors for ER, PR, HER2, and Ki67 markers. An in-house dataset was used, containing 92 multiparametric MRIs of patients with histologically proven BC and all four relevant biomarkers available. Thousands of radiomic features were extracted from post-contrast and subtracted Dynamic Contrast-Enanched (DCE) MRI images, Apparent Diffusion Coefficient (ADC) maps, and T2-weighted (T2) images. The two-step feature selection approach was used to identify significant radiomic features properly and then to build the final prediction models. They showed remarkable results in terms of F1-score for all the biomarkers: 84%, 63%, 90%, and 72% for ER, HER2, Ki67, and PR, respectively. When possible, the models were validated on the TCGA/TCIA Breast Cancer dataset, returning promising results (F1-score = 88% for the ER+/ER&minus; classification task). The developed approach efficiently characterized BC heterogeneity according to the examined molecular biomarkers