Alexandros Makriyannis

Peripheral CB1 cannabinoid receptor blockade improves cardiometabolic risk in mouse models of obesity.

Obesity and its metabolic consequences are a major public health concern worldwide. Obesity is associated with overactivity of the endocannabinoid system, which is involved in the regulation of appetite, lipogenesis, and insulin resistance. Cannabinoid-1 receptor (CB1R) antagonists reduce body weight and improve cardiometabolic abnormalities in experimental and human obesity, but their therapeutic potential is limited by neuropsychiatric side effects. Here we have demonstrated that a CB1R neutral antagonist largely restricted to the periphery does not affect behavioral responses mediated by CB1R in the brains of mice with genetic or diet-induced obesity, but it does cause weight-independent improvements in glucose homeostasis, fatty liver, and plasma lipid profile. These effects were due to blockade of CB1R in peripheral tissues, including the liver, as verified through the use of CB1R-deficient mice with or without transgenic expression of CB1R in the liver. These results suggest that targeting peripheral CB1R has therapeutic potential for alleviating cardiometabolic risk in obese patients.

Convergent translational evidence of a role for anandamide in amygdala-mediated fear extinction, threat processing and stress-reactivity

Endocannabinoids are released ‘on-demand’ on the basis of physiological need, and can be pharmacologically augmented by inhibiting their catabolic degradation. The endocannabinoid anandamide is degraded by the catabolic enzyme fatty acid amide hydrolase (FAAH). Anandamide is implicated in the mediation of fear behaviors, including fear extinction, suggesting that selectively elevating brain anandamide could modulate plastic changes in fear. Here we first tested this hypothesis with preclinical experiments employing a novel, potent and selective FAAH inhibitor, AM3506 (5-(4-hydroxyphenyl)pentanesulfonyl fluoride). Systemic AM3506 administration before extinction decreased fear during a retrieval test in a mouse model of impaired extinction. AM3506 had no effects on fear in the absence of extinction training, or on various non-fear-related measures. Anandamide levels in the basolateral amygdala were increased by extinction training and augmented by systemic AM3506, whereas application of AM3506 to amygdala slices promoted long-term depression of inhibitory transmission, a form of synaptic plasticity linked to extinction. Further supporting the amygdala as effect-locus, the fear-reducing effects of systemic AM3506 were blocked by intra-amygdala infusion of a CB1 receptor antagonist and were fully recapitulated by intra-amygdala infusion of AM3506. On the basis of these preclinical findings, we hypothesized that variation in the human FAAH gene would predict individual differences in amygdala threat-processing and stress-coping traits. Consistent with this, carriers of a low-expressing FAAH variant (385A allele; rs324420) exhibited quicker habituation of amygdala reactivity to threat, and had lower scores on the personality trait of stress-reactivity. Our findings show that augmenting amygdala anandamide enables extinction-driven reductions in fear in mouse and may promote stress-coping in humans.

Cannabinoid receptor antagonists: pharmacological opportunities, clinical experience, and translational prognosis.

The endogenous cannabinoid (CB) (endocannabinoid) signaling system is involved in a variety of (patho)physiological processes, primarily by virtue of natural, arachidonic acid-derived lipids (endocannabinoids) that activate G protein-coupled CB1 and CB2 receptors. A hyperactive endocannabinoid system appears to contribute to the etiology of several disease states that constitute significant global threats to human health. Consequently, mounting interest surrounds the design and profiling of receptor-targeted CB antagonists as pharmacotherapeutics that attenuate endocannabinoid transmission for salutary gain. Experimental and clinical evidence supports the therapeutic potential of CB1 receptor antagonists to treat overweight/obesity, obesity-related cardiometabolic disorders, and substance abuse. Laboratory data suggest that CB2 receptor antagonists might be effective immunomodulatory and, perhaps, anti-inflammatory drugs. One CB1 receptor antagonist/inverse agonist, rimon-abant, has emerged as the first-in-class drug approved outside the United States for weight control. Select follow-on agents (taranabant, otenabant, surinabant, rosonabant, SLV-319, AVE1625, V24343) have also been studied in the clinic. However, rimonabants market withdrawal in the European Union and suspension of rimonabants, taranabants, and otenabants ongoing development programs have highlighted some adverse clinical side effects (especially nausea and psychiatric disturbances) of CB1 receptor antagonists/inverse agonists. Novel CB1 receptor ligands that are peripherally directed and/or exhibit neutral antagonism (the latter not affecting constitutive CB1 receptor signaling) may optimize the benefits of CB1 receptor antagonists while minimizing any risk. Indeed, CB1 receptor-neutral antagonists appear from preclinical data to offer efficacy comparable to or better than that of prototype CB1 receptor antagonists/inverse agonists, with less propensity to induce nausea. Continued pharmacological profiling, as the prelude to first-in-man testing of CB1 receptor antagonists with unique modes of targeting/pharmacological action, represents an exciting translational frontier in the critical path to CB receptor blockers as medicines.

CB2 cannabinoid receptor mediation of antinociception

Abstract Management of acute pain remains a significant clinical problem. In preclinical studies, CB2 cannabinoid receptor‐selective agonists inhibit nociception without producing central nervous system side effects. The CB2 receptor‐selective agonist AM1241 produces antinociceptive effects that are antagonized by CB2, but not CB1, receptor‐selective antagonists, suggesting that activation of CB2 receptors results in antinociception. However, it has not been possible to definitively demonstrate that these effects are mediated by CB2 receptors, because we have lacked the pharmacological tools to confirm the in vivo receptor selectivity of the antagonists used. Further, recent evidence for cannabinoid‐like receptors beyond CB1 and CB2 raises the possibility that AM1241 exerts its antinociceptive effects at uncharacterized CB2‐like receptors that are also inhibited by AM630. The experiments reported here further test the hypothesis that CB2 receptor activation inhibits nociception. They evaluated the antinociceptive actions of AM1241 and the less‐selective CB2 receptor agonist WIN55,212‐2 in wild‐type Symbol mice and in mice with genetic disruption of the CB2 receptor (Symbol mice). AM1241 inhibited thermal nociception in Symbol mice, but had no effect in Symbol littermates. WIN55,212‐2 produced equivalent antinociception in Symbol and Symbol mice, while its antinociceptive effects were reduced in Symbol compared to Symbol mice. The effects of morphine were not altered in Symbol compared to Symbol mice. These data strongly suggest that AM1241 produces antinociception in vivo by activating CB2 cannabinoid receptors. Further, they confirm the potential therapeutic relevance of CB2 cannabinoid receptors for the treatment of acute pain. Figure. No Caption available. Figure. No Caption available. Figure. No Caption available. Figure. No Caption available. Figure. No Caption available. Figure. No Caption available. Figure. No Caption available. Figure. No Caption available. Figure. No Caption available. Figure. No Caption available.

Should peripheral CB1 cannabinoid receptors be selectively targeted for therapeutic gain

Endocannabinoids, endogenous lipid ligands of cannabinoid receptors, mediate a variety of effects similar to those of marijuana. Cannabinoid CB(1) receptors are highly abundant in the brain and mediate psychotropic effects, which limits their value as a potential therapeutic target. There is growing evidence for CB(1) receptors in peripheral tissues that modulate a variety of functions, including pain sensitivity and obesity-related hormonal and metabolic abnormalities. In this review we propose that selective targeting of peripheral CB(1) receptors has potential therapeutic value because it would help to minimize addictive, psychoactive effects in the case of CB(1) agonists used as analgesics, or depression and anxiety in the case of CB(1) antagonists used in the management of cardiometabolic risk factors associated with the metabolic syndrome.

Cannabinoid CB1 receptor inverse agonists and neutral antagonists: Effects on food intake, food-reinforced behavior and food aversions

Drugs that interfere with cannabinoid CB1 receptor transmission suppress a number of food-related behaviors, and these compounds are currently being assessed for their potential utility as appetite suppressants. In addition to rimonabant (SR141716A), several other compounds have been evaluated, including AM251 and AM1387. Biochemical studies indicate that most of the drugs assessed thus far have been CB1 inverse agonists, and these drugs all act to suppress food intake and disrupt food-reinforced behavior. Behavioral tests involving intake of different diets (i.e., high fat, high carbohydrate, laboratory chow) indicate that consumption of all three food types is disrupted by CB1 inverse agonists, and that, expressed as a percent of baseline intake, the effect is roughly comparable across different diets. Although CB1 inverse agonists do not appear to produce severe motor impairments that disrupt feeding behavior, there is evidence that they can induce nausea and malaise. Recent studies have been undertaken to characterize the behavioral effects of CB1 receptor neutral antagonists such as AM4113 to determine if these drugs can reduce feeding and food-reinforced behaviors. Across a variety of different tests, AM4113 produces effects on food-motivated behavior that are very similar to those produced by CB1 inverse agonists. Moreover, this drug did not induce conditioned gaping in rats or vomiting in ferrets. These results suggest that CB1 receptor neutral antagonists may decrease appetite by blocking endogenous cannabinoid tone, and that these drugs may be less associated with nausea than is the case for CB1 inverse agonists.

The Endogenous Cannabinoid Anandamide Produces δ-9-Tetrahydrocannabinol-Like Discriminative and Neurochemical Effects That Are Enhanced by Inhibition of Fatty Acid Amide Hydrolase but Not by Inhibition of Anandamide Transport

Anandamide is an endogenous ligand for brain cannabinoid CB1 receptors, but its behavioral effects are difficult to measure due to rapid inactivation. Here we used a drug-discrimination procedure to test the hypothesis that anandamide, given i.v. or i.p., would produce discriminative effects like those of δ-9-tetrahydrocannabinol (THC) in rats when its metabolic inactivation was inhibited. We also used an in vivo microdialysis procedure to investigate the effects of anandamide, given i.v. or i.p., on dopamine levels in the nucleus accumbens shell in rats. When injected i.v., methanandamide (AM-356), a metabolically stable anandamide analog, produced clear dose-related THC-like discriminative effects, but anandamide produced THC-like discriminative effects only at a high 10-mg/kg dose that almost eliminated lever-press responding. Cyclohexyl carbamic acid 3′-carbamoyl-biphenyl-3-yl ester (URB-597), an inhibitor of fatty acid amide hydrolase (FAAH), the main enzyme responsible for metabolic inactivation of anandamide, produced no THC-like discriminative effects alone but dramatically potentiated discriminative effects of anandamide, with 3 mg/kg anandamide completely substituting for the THC training dose. URB-597 also potentiated the ability of anandamide to increase dopamine levels in the accumbens shell. The THC-like discriminative-stimulus effects of anandamide after URB-597 and methanandamide were blocked by the CB1 receptor antagonist rimonabant, but not the vanilloid VR1 receptor antagonist capsazepine. Surprisingly, the anandamide transport inhibitors N-(4-hydroxyphenyl)-eicosa-5,8,11,14-tetraenamide (AM-404) and N-(3-furylmethyl)eicosa-5,8,11,14-tetraenamide (UCM-707) did not potentiate THC-like discriminative effects of anandamide or its dopamine-elevating effects. Thus, anandamide has THC-like discriminative and neurochemical effects that are enhanced after treatment with a FAAH inhibitor but not after treatment with transport inhibitors, suggesting brain area specificity for FAAH versus transport/FAAH inactivation of anandamide.

Dual Modulation of Endocannabinoid Transport and Fatty Acid Amide Hydrolase Protects against Excitotoxicity

The endocannabinoid system has been suggested to elicit signals that defend against several disease states including excitotoxic brain damage. Besides direct activation with CB1 receptor agonists, cannabinergic signaling can be modulated through inhibition of endocannabinoid transport and fatty acid amide hydrolase (FAAH), two mechanisms of endocannabinoid inactivation. To test whether the transporter and FAAH can be targeted pharmacologically to modulate survival/repair responses, the transport inhibitor N-(4-hydroxyphenyl)-arachidonamide (AM404) and the FAAH inhibitor palmitylsulfonyl fluoride (AM374) were assessed for protection against excitotoxicity in vitro and in vivo. AM374 and AM404 both enhanced mitogen-activated protein kinase (MAPK) activation in cultured hippocampal slices. Interestingly, combining the distinct inhibitors produced additive effects on CB1 signaling and associated neuroprotection. After an excitotoxic insult in the slices, infusing the AM374/AM404 combination protected against cytoskeletal damage and synaptic decline, and the protection was similar to that produced by the stable CB1 agonist AM356 (R-methanandamide). AM374/AM404 and the agonist also elicited cytoskeletal and synaptic protection in vivo when coinjected with excitotoxin into the dorsal hippocampus. Correspondingly, potentiating endocannabinoid responses with the AM374/AM404 combination prevented behavioral alterations and memory impairment that are characteristic of excitotoxic damage. The protective effects mediated by AM374/AM404 were (1) evident 7 d after insult, (2) correlated with the preservation of CB1-linked MAPK signaling, and (3) were blocked by a selective CB1 antagonist. These results indicate that dual modulation of the endocannabinoid system with AM374/AM404 elicits neuroprotection through the CB1 receptor. The transporter and FAAH are modulatory sites that may be exploited to enhance cannabinergic signaling for therapeutic purposes.

Dietary docosahexaenoic acid supplementation alters select physiological endocannabinoid-system metabolites in brain and plasma

The endocannabinoid metabolome consists of a growing, (patho)physiologically important family of fatty-acid derived signaling lipids. Diet is a major source of fatty acid substrate for mammalian endocannabinoid biosynthesis. The principal long-chain PUFA found in mammalian brain, docosahexaenoic acid (DHA), supports neurological function, retinal development, and overall health. The extent to which dietary DHA supplementation influences endocannabinoid-related metabolites in brain, within the context of the circulating endocannabinoid profile, is currently unknown. We report the first lipidomic analysis of acute 2-week DHA dietary supplementation effects on the physiological state of 15 fatty-acid, N-acylethanolamine, and glycerol-ester endocannabinoid metabolome constituents in murine plasma and brain. The DHA-rich diet markedly elevated DHA, eicosapentaenoic acid, 2-eicosapentanoylglycerol (EPG), and docosahexanoylethanolamine in both compartments. Dietary DHA enhancement generally affected the synthesis of the N-acyl-ethanolamine and glycerol-ester metabolites to favor the docosahexaenoic and eicosapentaenoic vs. arachidonoyl and oleoyl homologs in both brain and plasma. The greater overall responsiveness of the endocannabinoid metabolome in plasma versus brain may reflect a more circumscribed homeostatic response range of brain lipids to dietary DHA supplementation. The ability of short-term DHA enhancement to modulate select constituents of the physiological brain and plasma endocannabinoid metabolomes carries metabolic and therapeutic implications.

Latest advances in cannabinoid receptor agonists.

Background: Since the discovery of cannabinoid receptors and their endogenous ligands in early 1990s, the endocannabinoid system has been shown to play a vital role in several pathophysiological processes. It has been targeted for the treatment of several diseases including neurodegenerative diseases (Parkinsons disease, Alzheimers disease, Huntingtons disease and MS), cancer, obesity, inflammatory bowel disease, neuropathic and inflammatory pain. The last decade has witnessed remarkable advances in the development of cannabinergic ligands displaying high selectivity and potency towards two subtypes of cannabinoid receptors, namely CB1 and CB2. Objective: In this review, we highlight the latest advances made in the development of cannabinoid agonists and summarize recently disclosed, novel chemical scaffolds as CB-selective agonists in patents that appeared during January 2008 – June 2009. Methods: Data presented here are obtained through the search of PubMed for research articles and reviews, and the website of European patents (http://ep.espacenet.com), SciFinder Scholar™ and US patents (www.uspto.gov). Conclusions: Our analysis reveals prolific patenting activity mainly in the CB2 selective agonist area. Limiting the BBB penetrability, thereby, leading to peripherally restricted CB1/CB2 agonists and enhancing CB2-selectivity emerge as likely prerequisites for avoidance of adverse central CB1 mediated side effects.