Cannabinoid Analogues Research Articles

ChemInform Abstract: Identification of a Cannabinoid Analogue (I) as a New Type of Designer Drug in a Herbal Product.

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Endocannabinoids and cannabinoid analogues block cardiac hKv1.5 channels in a cannabinoid receptor-independent manner

Endocannabinoids are synthesized from lipid precursors at the plasma membranes of virtually all cell types, including cardiac myocytes. Endocannabinoids can modulate neuronal and vascular ion channels through receptor-independent actions; however, their effects on cardiac K(+) channels are unknown. This study was undertaken to determine the receptor-independent effects of endocannabinoids such as anandamide (N-arachidonoylethanolamine, AEA), 2-arachidonoylglycerol (2-AG), and endocannabinoid-related compounds such as N-palmitoylethanolamine (PEA), N-oleoylethanolamine (OEA), the endogenous lipid lysophosphatidylinositol (LPI), and the fatty acids from which some of these compounds are endogenously synthesized, on human cardiac Kv1.5 channels, which generate the ultrarapid delayed rectifier current (I(Kur)). hKv1.5 currents (I(hKv1.5)) were recorded in mouse fibroblasts (Ltk(-) cells) by using the whole-cell patch-clamp technique. Most of these compounds inhibited I(hKv1.5) in a concentration-dependent manner, the potency being determined by the number of C atoms in the fatty acyl chain. Indeed, AEA and 2-AG, which are arachidonic acid (20:4) derivatives, exhibited the highest potency (IC(50) approximately 0.9-2.5 microM), whereas PEA, a palmitic acid (PA-16:0) derivative, exhibited the lowest potency. The inhibition was independent of cannabinoid receptor engagement and of changes in the order and microviscosity of the membrane. Furthermore, blockade induced by AEA and 2-AG was abolished upon mutation of the R487 residue, which determines the external tetraethylammonium sensitivity and is located in the external entryway of the pore. AEA significantly prolonged the duration of action potentials (APs) recorded in mouse left atria. These results indicate that endocannabinoids block human cardiac Kv1.5 channels by interacting with an extracellular binding site, a mechanism by which these compounds regulate atrial AP shape.

Endocannabinoids and cannabinoid analogues block human cardiac Kv4.3 channels in a receptor-independent manner

Endocannabinoids are amides and esters of long chain fatty acids that can modulate ion channels through both receptor-dependent and receptor-independent effects. Nowadays, their effects on cardiac K + channels are unknown even when they can be synthesized within the heart. We have analyzed the direct effects of endocannabinoids, such as anandamide (AEA), 2-arachidonoylglycerol (2-AG), the endogenous lipid lysophosphatidylinositol, and cannabinoid analogues such as palmitoylethanolamide (PEA), and oleoylethanolamide, as well as the fatty acids from which they are endogenously synthesized, on human cardiac Kv4.3 channels, which generate the transient outward K + current ( I to1). Currents were recorded in Chinese hamster ovary cells, which do not express cannabinoid receptors, by using the whole-cell patch-clamp. All these compounds inhibited I Kv4.3 in a concentration-dependent manner, AEA and 2-AG being the most potent (IC 50 ∼ 0.3–0.4 µM), while PEA was the least potent. The potency of block increased as the complexity and the number of C atoms in the fatty acyl chain increased. The effects were not mediated by modifications in the lipid order and microviscosity of the membrane and were independent of the presence of MiRP2 or DPP6 subunits in the channel complex. Indeed, effects produced by AEA were reproduced in human atrial I to1 recorded in isolated myocytes. Moreover, AEA effects were exclusively apparent when it was applied to the external surface of the cell membrane. These results indicate that at low micromolar concentrations the endocannabinoids AEA and 2-AG directly block human cardiac Kv4.3 channels, which represent a novel molecular target for these compounds.

Single Nucleotide Change in the Cannabinoid Receptor-1 (CNR1) Gene in Colorectal Cancer Outcome

The cannabinoid receptor-1 (CNR-1) and endogenous agonists of this receptor are present in the central and peripheral nervous systems including the gastrointestinal nervous system. The surgically rejected specimens of human colorectal cancers and paired normal tissues were studied to detect mutations in the CNR1 gene by sequencing method. The results were compared to clinicopathological parameters and correlated with overall survival time. Sixty-three colorectal cancer patients, who underwent surgical excision of colorectal carcinoma, were included in this study. The coding region of the CNR1 gene was studied: a nucleotide change (G→A) at position 1359 was identified by direct sequencing of PCR. Thirty-eight patients had the G/G genotype (wild type) in tumor areas and 25 patients had G/A heterozygous or A/A homozygous genotype. Univariate analysis revealed 2 independent variables associated with CNR1 gene mutation. The results show that the patients with Dukes stage C and D had a 2.9 times (p = 0.04) and patients that were lymph node positive had 2.8 times (p = 0.05) greater probability of nucleotide change in CNR1 gene. Genotype G/A plus A/A had a shorter overall survival time than G/G wild-type patients (p < 0.05). Indeed nontumor paired colorectal tissues showed nucleotide change. A large number of patients with mutation in the CNR1 gene were observed. These preliminary findings highlight the importance of further studies in the use of cannabinoid analogs as receptor ligands to analyze potential therapeutic effects.

The Endocannabinoid System: An Ancient Signaling Involved in the Control of Male Fertility

The effects of cannabinoids on human health have been known since the antiquities when the extract of the plant Cannabis sativa was used because of its psychoactivity. The scientific story of the cannabinoids started in the 1960s with the isolation and characterization of the active component of the plant. After the synthesis of cannabinoid analogues, the analysis of structure-effect relationships was implemented, and this had a similar effect to a positive "Pandora's box" opening. To date, numerous roles have been ascribed to the "endocannabinoid system." Here we describe its involvement in the control of male reproduction, taking into consideration possible evolutionary speculations. Indeed, the endocannabinoid system is a very ancient signaling system, being clearly present from the divergence of the protostomian/deuterostomian.

Hippocampal CB(1) receptors mediate the memory impairing effects of Delta(9)-tetrahydrocannabinol.

It is firmly established that the hippocampus, a brain region implicated in spatial learning, episodic memory, and consolidation, contains a high concentration of CB1 receptors. Moreover, systemic and intrahippocampal administration of cannabinoid agonists have been shown to impair hippocampal-dependent memory tasks. However, the degree to which CB1 receptors in the hippocampus play a specific functional role in the memory disruptive effects of marijuana or its primary psychoactive constituent Δ9-tetrahydrocannabinol (Δ9-THC) is unknown. The present study was designed to determine whether hippocampal CB1 receptors play a functional role in the memory disruptive effects of systemically administered cannabinoids, using the radial arm maze, a well characterized rodent model of working memory. Male Sprague-Dawley rats were implanted with bilateral cannulae aimed at the CA1 region of the dorsal hippocampus. The CB1 receptor antagonist, rimonabant, was delivered into the hippocampus prior to a systemic injection of either Δ9-THC or the potent cannabinoid analog, CP-55,940. Strikingly, intrahippocampal administration of rimonabant completely attenuated the memory disruptive effects of both cannabinoids in the radial arm maze task, but did not affect other pharmacological properties of cannabinoids, as assessed in the tetrad assay (i.e., hypomotility, analgesia, catalepsy, and hypothermia). Infusions of rimonabant just dorsal or ventral to the hippocampus did not prevent Δ9-THC-induced memory impairment, indicating that its effects on mnemonic function were regionally selective. These findings provide compelling evidence in support of the view that hippocampal CB1 receptors play a necessary role in the memory disruptive effects of marijuana.

Identification of a cannabimimetic indole as a designer drug in a herbal product

A cannabimimetic indole has been identified as a new adulterant in a herbal product being sold illegally in Japan for its expected narcotic effect. Liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry analyses indicated that the product contained two major compounds. One was identified as a cannabinoid analog (1RS,3SR)-3-[4-(1,1-dimethyloctyl)-2-hydroxyphenyl]cyclohexan-1-ol (1) by direct comparison with the authentic compound, which we reported previously. The other compound (2) showed a molecular weight of 341 daltons, and accurate mass spectral measurements showed its elemental composition to be C24H23NO. Both mass and nuclear magnetic resonance spectrometric data revealed that 2 was 1-pentyl-3-(1-naphthoyl)indole [or naphthalen-1-yl-(1-pentylindol-3-yl)methanone] being identical to JWH-018, which was synthesized by Wiley and coworkers in 1998. This compound was reported as a potent cannabinoid receptor agonist possessing a pharmacological cannabimimetic activity.

Identification of a Cannabinoid Analog as a New Type of Designer Drug in a Herbal Product

A new type of designer drug, a cannabinoid analog (1), was found in a herbal product distributed on the illegal drug market in Japan in expectation of its narcotic effect. The structure of 1 was identified by LC-MS, GC-MS, high-resolution MS, and NMR analyses. Compound 1 showed a molecular weight of 332, and accurate mass measurement exhibited its elemental composition to be C(22)H(36)O(2). Together, the mass and NMR spectrometric data revealed that 1 was (1RS,3SR)-3-[4-(1,1-dimethyloctyl)-2-hydroxyphenyl]cyclohexan-1-ol, which was first synthesized in 1979 by a group at Pfizer Inc. and reported as a potent cannabinoid analog possessing cannabinoid receptor binding activity and analgesic activity in the 1990 s. This is the first report to identify a cannabinoid analog in an illegal drug.

A cannabinoid analogue of Δ9‐tetrahydrocannabinol disrupts neural development in chick

Marijuana is the most commonly abused illicit drug by pregnant women. Its major psychoactive constituent, Delta(9)-THC (Delta(9)-tetrahydrocannabinol), crosses the placenta and accumulates in the foetus, potentially harming its development. In humans, marijuana use in early pregnancy is associated with miscarriage, a fetal alcohol-like syndrome, as well as learning disabilities, memory impairment, and ADHD in the offspring. Classical studies in the 1970 s have reached disparate conclusions as to the teratogenic effects of cannabinoids in animal models. Further, there is very little known about the immediate effects of Delta(9)-THC on early embryogenesis. We have used the chick embryo as a model in order to characterize the effects of a water-soluble Delta(9)-THC analogue, O-2545, on early development. Embryos were exposed to the drug (0.035 to 0.35 mg/ml) at gastrulation and assessed for morphological defects at stages equivalent to 9-14 somites. We report that O-2545 impairs the formation of brain, heart, somite, and spinal cord primordia. Shorter incubation times following exposure to the drug show that O-2545 interferes with the initial steps of head process and neural plate formation. Our results indicate that the administration of the cannabinoid O-2545 during early embryogenesis results in embryotoxic effects and serves to illuminate the risks of marijuana exposure during the second week of pregnancy, a time point at which most women are unaware of their pregnancies.

Bornyl- and Isobornyl-Δ8-tetrahydrocannabinols: A Novel Class of Cannabinergic Ligands

Structure-activity relationship studies of classical cannabinoid analogues have established that the C3 aliphatic side chain plays a pivotal role in determining cannabinergic potency. In earlier work, we provided evidence for the presence of subsites within the CB1 and CB2 cannabinoid receptor binding domains that can accommodate bulky conformationally defined substituents at the C3 alkyl side chain pharmacophore of classical cannabinoids. We have now extended this work with the synthesis of a series of Delta (8)-THC analogues in which bornyl substituents are introduced at the C3 position. Our results indicate that, for optimal interactions with both CB1 and CB2 receptors, the bornyl substituents need to be within close proximity of the tricyclic core of Delta (8)-THC and that the conformational space occupied by the C3 substituents influences CB1/CB2 receptor subtype selectivity.

A Furan Recyclization Reaction as a New Approach to Isochromenes

As part of our ongoing interest in the synthesis of benz-annelated heterocycles from O-substituted benzylfurans, recyclization of [ O-(hydroxymethyl)aryl]difurylmethanes was studied. It was shown that, under acidic conditions, the O-hydroxymethyl group in these compounds acts as a nucleophile and tetracyclic isochromene derivatives are formed via a recyclization-cyclization tandem sequence. Alternatively, these compounds can be synthesized by reduction of the corresponding isochromone derivatives with lithium aluminum hydride. The potent biologically active analogues of cannabinoids were also obtained by interaction of isochromone derivatives with excess of methylmagnesium iodide.

A Cyclotrimerization Route to Cannabinoids

Three members of the cannabinoid class, cannabinol, cannabinol methyl ether, and cannabinodiol, were synthesized using a microwave-mediated [2 + 2 + 2] cyclotrimerization reaction as the key step. This approach provides a high level of synthetic flexibility allowing for the facile synthesis of cannabinoid analogues.

Efficient one-pot synthetic approaches for cannabinoid analogues and their application to biologically interesting (−)-hexahydrocannabinol and (+)-hexahydrocannabinol

This Letter reports new and efficient synthetic approaches for biologically interesting cannabinoid analogues. The key strategies involve ethylenediamine diacetate/triethylamine-catalyzed cyclization. As an application of this methodology, one-step synthesis of biologically active natural (−)-hexahydrocannabinol and its unnatural enantiomer (+)-hexahydrocannabinol was carried out.

Nonpsychoactive Cannabidiol Prevents Prion Accumulation and Protects Neurons against Prion Toxicity

Prion diseases are transmissible neurodegenerative disorders characterized by the accumulation in the CNS of the protease-resistant prion protein (PrPres), a structurally misfolded isoform of its physiological counterpart PrPsen. Both neuropathogenesis and prion infectivity are related to PrPres formation. Here, we report that the nonpsychoactive cannabis constituent cannabidiol (CBD) inhibited PrPres accumulation in both mouse and sheep scrapie-infected cells, whereas other structurally related cannabinoid analogs were either weak inhibitors or noninhibitory. Moreover, after intraperitoneal infection with murine scrapie, peripheral injection of CBD limited cerebral accumulation of PrPres and significantly increased the survival time of infected mice. Mechanistically, CBD did not appear to inhibit PrPres accumulation via direct interactions with PrP, destabilization of PrPres aggregates, or alteration of the expression level or subcellular localization of PrPsen. However, CBD did inhibit the neurotoxic effects of PrPres and affected PrPres-induced microglial cell migration in a concentration-dependent manner. Our results suggest that CBD may protect neurons against the multiple molecular and cellular factors involved in the different steps of the neurodegenerative process, which takes place during prion infection. When combined with its ability to target the brain and its lack of toxic side effects, CBD may represent a promising new anti-prion drug.

Cannabinoids as therapeutic agents in cardiovascular disease: a tale of passions and illusions

In addition to their classical known effects, such as analgesia, impairment of cognition and learning and appetite enhancement, cannabinoids have also been related to the regulation of cardiovascular responses and implicated in cardiovascular pathology. Elevated levels of endocannabinoids have been related to the extreme hypotension associated with various forms of shock as well as to the cardiovascular abnormalities that accompany cirrhosis. In contrast, cannabinoids have also been associated with beneficial effects on the cardiovascular system, such as a protective role in atherosclerosis progression and in cerebral and myocardial ischaemia. In addition, it has also been suggested that the pharmacological manipulation of the endocannabinoid system may offer a novel approach to antihypertensive therapy. During the last decades, the tremendous increase in the understanding of the molecular basis of cannabinoid activity has encouraged many pharmaceutical companies to develop more potent synthetic cannabinoid analogues and antagonists, leading to an explosion of basic research and clinical trials. Consequently. not only the synthetic THC dronabinol (Marinol) and the synthetic THC analogue nabilone (Cesamet) have been approved in the United States, but also the standardized cannabis extract (Sativex) in Canada. At least three strategies can be foreseen in the future clinical use of cannabinoid-based drugs: (a) the use of CB(1) receptor antagonists, such as the recently approved rimonabant (b) the use of CB(2)-selective agonists, and (c) the use of inhibitors of endocannabinoid degradation. In this context, the present review examines the effects of cannabinoids and of the pharmacological manipulation of the endocannabinoid system, in cardiovascular pathophysiology.

The Application of 3D-QSAR Studies for Novel Cannabinoid Ligands Substituted at the C1‘ Position of the Alkyl Side Chain on the Structural Requirements for Binding to Cannabinoid Receptors CB1 and CB2

A set of 30 novel Delta8-tetrahydrocannabinol and cannabidiol analogues were subjected to three-dimensional quantitative structure-activity relationship studies using the comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) approaches. Using a combination of molecular modeling techniques and NMR spectroscopy, the putative bioactive conformation of the most potent cannabinoid (CB) ligand in the training set was determined. This conformer was used as the template and CB1 and CB2 pharmacophore models were developed. These models were fitted with experimental binding data and gave high correlation coefficients. Contour maps of the CB1 and CB2 models of CoMFA and CoMSIA approaches show that steric effects dominantly determine the binding affinities. The CoMFA and CoMSIA analyses based on the binding affinity data of CB ligands at the CB1 and CB2 receptors allowed us to deduce the possible optimal binding positions. This information can be used for the design of new CB analogues with enhanced activity and other tailored properties.

Cannabinoids inhibit sodium-dependent, high-affinity excitatory amino acid transport in cultured rat cortical astrocytes

Cannabinoids have been shown to increase the extracellular levels of glutamate in vivo and in vitro, but no studies have evaluated the possible involvement of glial glutamate reuptake system. The present study investigates whether cannabinoids and endocannabinoid, anandamide have an effect on astroglial excitatory amino acid (EAA) transport. The kinetics of glutamate transport was studied in rat cortical astrocytes, using the radiolabeled, non-metabolized amino acid, d-[ 3H] aspartate in the absence or presence of cannabinoid receptor agonists. The results show that in vehicle controls the uptake of d-aspartate was rapid, sodium-dependent and saturated within the first 5 min, resulting in a K m 7.365 ± 1.16 μmol/L ( n = 5) and the maximum velocity ( V max) 1207 ± 51 nmol/mg protein/min. Addition of the synthetic cannabinoid analog R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolol][1,2,3de]-1,4-benzoxazinyl]-(1-naphthalenyl)methanone (WIN 55,212-2; 3 μmol/L) increased the K m (26.25 ± 4.84 μmol/L) without affecting the V max (1122 ± 77 nmol/mg protein/min), suggesting the inhibition was competitive and reversible. Various other cannabinoid agonists also inhibited d-aspartate uptake in a dose-dependent and stereospecific manner. The cannabinoid inhibition of EAA transport was partially blocked by the cannabinoid type-1 (CB1) receptor antagonist N-(piperidin-1-yl-5(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl- 1H-pyrazole-3-carboxamidehydrochloride (SR141716A; 100 nmol/L). The inhibitory effects of WIN 55,212-2, or its endogenous counterpart anandamide were reversed by 98,059, an inhibitor of mitogen-activated kinase (MAPK) kinase (MEK). These results suggest that cannabinoids and endocannabinoids may constitute a novel class of inhibitors of astroglial glutamate transport system.

P-402: Fertility loss in the sperm of tobacco smokers may be reversed after washing with a cannabinoid agonist

Human sperm carry a nicotinic acetylcholine receptor and respond to nicotine from tobacco. They also have cannabinoid (CB) receptors and respond to marijuana and endocannabinoids or synthetic CB chemicals. We previously reported that sperm from 2/3 of tobacco smokers showed a significant drop in fertilizing capacity, using the Hemizona Assay (HZA). Their sperm showed reduced binding to the zona pellucida (cover) of the human egg. In Part II, we hypothesized that treatment of the smoker’s sperm with a CB agonist would improve sperm binding capacity. We have studied the effects of a CB agonist (AM-1346; analog of the endocannabinoid anandamide). After obtaining a positive response when donor sperm were treated with nicotine plus a CB agonist, we studied the sperm from 8 chronic tobacco smokers in 24 in vitro HZA tests. Each smoker was previously tested in the HZA, where his sperm fertilizing function was compared to a non-smoking donor. Four of 8 smokers (Group I) had shown normal sperm fertilizing function (HZI > 65). The other 4 smokers (Group II) had failed the zona assay. In Part II, each new test used the sperm from one tobacco smoker. His sperm were washed with and without a CB ligand (Control had no ligand present). We tested the effects of a potent CB1 agonist, AM-1346 (n = 17; 100 pM to 100 nM). After a “swim-up,” the motile sperm fraction was used for the HZA. Here, a nonviable human oocyte was cut in half. One half was incubated with smoker’s sperm that had been treated with a CB ligand. The matching Hemizona (HZ) was placed with that smoker’s Control sperm. Post-incubation, each HZ was rinsed and counted for the number of sperm tightly bound to the outside. The ratio of [# bound for the treated sperm / # bound for the Control sperm] was calculated as the Hemizona Index (HZI). HZI = 100 indicates that the treated sperm bound to the zona as well as the Control sperm. We hypothesized that the HZI would improve with in vitro CB treatment. After exposing the sperm to AM-1346, at any concentration, we found that Group II (low fertility) responded with markedly improved zona binding, compared to the Control sperm (4 out of 8 experiments). Specifically, when semen quality was poor, Group II showed a mean HZI of 177% (77% improvement compared to the untreated sperm). However, the sperm from smokers with normal fertility (Group I) behaved differently. Here, only 1 of 9 tests showed a zona binding enhancement (p < 0.05). In Groups I & II, there were 7 experiments where the smoker was tested with a low concentration of AM-1346 (one assay) and also tested at a higher 1346 level (second assay). In 6 of 7 cases, the higher 1346 dose produced a better HZI score (improved binding). We conjecture that most smoking men would show a stimulation of zona binding if the dose of AM-1346 is high enough. Based on our previous data, and published literature, it is clear that a majority of tobacco smokers will exhibit a small or significant decline in fertility potential. Infertility clinics worldwide are trying to meet the challenge of male and female tobacco smokers who have only borderline fertility. Smoking cessation is the best solution. However, the in vitro use of synthetic cannabinoid analogs may significantly improve the fertilizing capacity of tobacco smokers who cannot quit.

Cannabinoids and pain management

controlled, parallel-group trial studying the analgesic effects of two different doses of nabilone, a synthetic analogue of THC, in managing acute postoperative pain following major surgery. 5 Unfortunately, the study was prematurely terminated after 18 months. The results of this study demonstrated that nabilone administration was not associated with a decrease in morphine consumption in patients following major surgery. In addition, nabilone at high doses significantly increased pain scores at rest and upon movement. The investigator concluded that, although cannabinoids may be effective in relieving chronic pain in humans, their role in the treatment of acute postoperative pain is uncertain, and requires further clinical evaluation. Small sample size, early termination of the study, and the lack of standardization of surgical procedures with possible variation in intensity of postoperative pain (e.g., total hip replacement vs total knee replacement) may limit the conclusions that can be drawn from this study. Previous studies examining cannabinoids in treating postoperative pain have also demonstrated only a modest analgesic effect. 6 A systematic review concluded that cannabinoids are no more effective than codeine in controlling acute pain, although this was a conclusion based upon only one Phase II study. 7 While the current study suggests a very limited role for cannabinoids in postoperative pain, it is important to recognize that this may change as novel, more potent and more specific synthetic cannabinoid analogs, with limited central nervous system effects, are developed. The role of cannabinoids in managing chronic pain is more promising. A systematic review of studies available prior to 2002 identified seven randomized controlled trials examining cannabinoids in chronic pain. These studies demonstrated that oral THC preparations exhibit a moderate analgesic effect, equivalent

Structural modifications of the cannabinoid side chain towards C3-aryl and 1′,1′-cycloalkyl-1′-cyano cannabinoids

The compounds reported in this study are Δ8-THC analogues in which the C3 five-carbon linear side chain of Δ8-THC was replaced with aryl and 1′,1′-cycloalkyl substituents. Of the compounds described here analogues 2d (CB1, Ki=11.7nM. CB2, Ki=9.39 nM) and 2f (CB1, Ki=8.26nM. CB2, Ki=3.86nM) exhibited enhanced binding affinities for CB1 and CB2, exceeding that of Δ8-THC. Efficient procedures for the synthesis of these novel cannabinoid analogues are described.