CB1 Neutral Antagonists Research Articles

Effects of the cannabinoid CB1-receptor neutral antagonist AM4113 and antagonist/inverse agonist rimonabant on fentanyl discrimination in male rats

Evidence suggests the existence of a functional interaction between endogenous cannabinoid (CB) and opioid systems. Thus, targeting CB1 receptors might be a viable approach to develop new medications for opioid use disorders (OUD). The present studies were undertaken to evaluate the effects of the neutral CB1 antagonist AM4113 and the CB1 antagonist/inverse agonist rimonabant in male rats trained to discriminate 0.032 mg/kg fentanyl from saline under a 10-response fixed-ratio (FR-10) schedule of food reinforcement. Results show that the µ-opioid agonists (fentanyl, oxycodone, and morphine) substituted fully and dose-dependently for fentanyl, whereas pretreatment with the µ-opioid antagonist naltrexone antagonized fentanyl's discriminative-stimulus effects. In interaction studies, AM4113 (0.32 or 1.0 mg/kg) was more effective in blocking fentanyl discrimination at 10-fold lower doses that did not modify rates of food-maintained responding, whereas rimonabant (1.0–10 mg/kg) produced some attenuation of fentanyl's discriminative-stimulus effects at the highest dose tested which also significantly decreased response rates. These results extend our recent work showing that AM4113 can effectively block the behavioral effects of heroin without producing rimonabant-like adverse effects. Taken together, these data suggests that CB1 neutral antagonists effectively block the behavioral effects of structurally distinct morphinan (heroin) and phenylpiperidine-based (fentanyl) opioids and may provide a novel therapeutic option for the treatment of OUD.

Effects of Cannabinoid Agonists and Antagonists in Rats Discriminating Fentanyl

Extensive evidence suggests the existence of a functional interaction between endogenous cannabinoids and opioid systems. Targeting cannabinoid CB1 receptors might be a promising approach for developing new medications for opioid addiction. The present studies were undertaken to evaluate the effects of CB1 agonists and antagonists in rats (n=8) trained to discriminate, i.p., injections of 0.032 mg/kg fentanyl from saline on a 10‐response fixed‐ratio (FR‐10) schedule of food reinforcement. Results show that the µ‐opioid agonists (fentanyl, morphine, and oxycodone) substituted fully for fentanyl, whereas the muscarinic antagonist atropine did not substitute for fentanyl. The CB1 agonists Δ9‐THC, AM2201, and AM8936 all partially substituted for fentanyl’s discriminative‐stimulus effects suggesting some overlap in the discriminative‐stimulus effects of cannabinoid agonists and fentanyl. Pretreatment studies with a µ‐opioid receptor antagonist show that naltrexone antagonized fentanyl’s effects. Interestingly, pretreatment studies with CB1 antagonists show that rimonabant (inverse agonist) produced some attenuation of fentanyl’s discriminative‐stimulus effects at doses that significantly decreased response rates, whereas the neutral CB1 antagonist AM4113 blocked fentanyl discrimination at doses that did not modify rates of responding. Our findings are consistent with our recent work showing that AM4113 attenuates heroin self‐administration in rats, without producing depressive‐like effects. Collectively, these data suggests that CB1 neutral antagonists that block CB1 receptors with rimonabant‐like potency, devoid of unwanted side‐effects, may be therapeutically advantageous for countering the abuse‐related behavioral effects of opioids.

Targeting the Endocannabinoid CB1 Receptor to Treat Body Weight Disorders: A Preclinical and Clinical Review of the Therapeutic Potential of Past and Present CB1 Drugs.

Obesity rates are increasing worldwide and there is a need for novel therapeutic treatment options. The endocannabinoid system has been linked to homeostatic processes, including metabolism, food intake, and the regulation of body weight. Rimonabant, an inverse agonist for the cannabinoid CB1 receptor, was effective at producing weight loss in obese subjects. However, due to adverse psychiatric side effects, rimonabant was removed from the market. More recently, we reported an inverse relationship between cannabis use and BMI, which has now been duplicated by several groups. As those results may appear contradictory, we review here preclinical and clinical studies that have studied the impact on body weight of various cannabinoid CB1 drugs. Notably, we will review the impact of CB1 inverse agonists, agonists, partial agonists, and neutral antagonists. Those findings clearly point out the cannabinoid CB1 as a potential effective target for the treatment of obesity. Recent preclinical studies suggest that ligands targeting the CB1 may retain the therapeutic potential of rimonabant without the negative side effect profile. Such approaches should be tested in clinical trials for validation.

Cannabinoid Antagonist Drug Discrimination in Nonhuman Primates.

Despite a growing acceptance that withdrawal symptoms can emerge following discontinuation of cannabis products, especially in high-intake chronic users, there are no Food and Drug Administration (FDA)-approved treatment options. Drug development has been hampered by difficulties studying cannabis withdrawal in laboratory animals. One preclinical approach that has been effective in studying withdrawal from drugs in several pharmacological classes is antagonist drug discrimination. The present studies were designed to examine this paradigm in squirrel monkeys treated daily with the long-acting CB1 agonist AM2389 (0.01 mg/kg) and trained to discriminate the CB1 inverse agonist/antagonist rimonabant (0.3 mg/kg) from saline. The discriminative-stimulus effects of rimonabant were both dose and time dependent and, importantly, could be reproduced by discontinuation of agonist treatment. Antagonist substitution tests with the CB1 neutral antagonists AM4113 (0.03-0.3 mg/kg), AM6527 (0.03-1.0 mg/kg), and AM6545 (0.03-1.0 mg/kg) confirmed that the rimonabant discriminative stimulus also could be reproduced by CB1 antagonists lacking inverse agonist action. Agonist substitution tests with the phytocannabinoid ∆9-tetrahydrocannabinol (0.1-1.0 mg/kg), synthetic CB1 agonists nabilone (0.01-0.1 mg/kg), AM4054 (0.01-0.03 mg/kg), K2/Spice compound JWH-018 (0.03-0.3 mg/kg), FAAH-selective inhibitors AM3506 (0.3-5.6 mg/kg), URB597 (3.0-5.6 mg/kg), and nonselective FAAH/MGL inhibitor AM4302 (3.0-10.0 mg/kg) revealed that only agonists with CB1 affinity were able to reduce the rimonabant-like discriminative stimulus effects of withholding daily agonist treatment. Although the present studies did not document physiologic disturbances associated with withdrawal, the results are consistent with the view that the cannabinoid antagonist drug discrimination paradigm provides a useful screening procedure for examining the ability of candidate medications to attenuate the interoceptive stimuli provoked by cannabis discontinuation. SIGNIFICANCE STATEMENT: Despite a growing acceptance that withdrawal symptoms can emerge following the discontinuation of cannabis products, especially in high-intake chronic users, there are no FDA-approved pharmacotherapies to assist those seeking treatment. The present studies systematically examined cannabinoid antagonist drug discrimination, a preclinical animal model that is designed to appraise the ability of candidate medications to attenuate the interoceptive effects that accompany abrupt cannabis abstinence.

Cannabinoid-1 receptor neutral antagonist reduces binge-like alcohol consumption and alcohol-induced accumbal dopaminergic signaling

Binge alcohol (ethanol) drinking is associated with profound adverse effects on our health and society. Rimonabant (SR141716A), a CB1 receptor inverse agonist, was previously shown to be effective for nicotine cessation and obesity. However, studies using rimonabant were discontinued as it was associated with an increased risk of depression and anxiety. In the present study, we examined the pharmacokinetics and effects of AM4113, a novel CB1 receptor neutral antagonist on binge-like ethanol drinking in C57BL/6J mice using a two-bottle choice drinking-in-dark (DID) paradigm. The results indicated a slower elimination of AM4113 in the brain than in plasma. AM4113 suppressed ethanol consumption and preference without having significant effects on body weight, ambulatory activity, preference for tastants (saccharin and quinine) and ethanol metabolism. AM4113 pretreatment reduced ethanol-induced increase in dopamine release in nucleus accumbens. Collectively, these data suggest an important role of CB1 receptor-mediated regulation of binge-like ethanol consumption and mesolimbic dopaminergic signaling, and further points to the potential utility of CB1 neutral antagonists for the treatment of binge ethanol drinking.

Metabolic Profiling of CB1 Neutral Antagonists.

PIMSR is among the first neutral antagonists for the CB1R and was demonstrated pharmacologically to bind to the CB1R, yet not alter calcium flux. It was further shown computationally to be able to stabilize both the active and inactive states of CB1R revealing the molecular interactions that mechanistically afford the property of neutral antagonism. PIMSR shows dramatic positive effects in reducing weight, food intake, and adiposity as well as in improving glycemic control and lipid homeostasis in high-fat diet-induced obese mice, but also shows increased ALT and liver weight as markers of liver injury with chronic administration. Further, in a separate study, 3-day administration of PIMSR in C57BL/6J mice, hepatic steatosis from an acute administration of high of ethanol was significantly reduced. Also, it partially prevented alcohol-induced increases in ALT, AST, and LDH. The differences in ALT levels in obese and nonobese mice under different test paradigms are unlikely to be due to neutral antagonism itself since other neutral antagonists (AM6545) do not exhibit liver injury. The brain levels of low micromolar would support significant brain CB1 receptor occupancy (re: Ki=17nM), thus potentially including both CNS and peripheral influences on the observed weight loss. Overall, these studies suggest that marked improvements in aspects of metabolic disease and alcoholic steatosis can be realized with CB1R neutral antagonists and hence warrants the exploration of further members of this class of cannabinoid ligands.

Controlled downregulation of the cannabinoid CB1 receptor provides a promising approach for the treatment of obesity and obesity-derived type 2 diabetes.

Increased activity of the endocannabinoid system has emerged as a pathogenic factor in visceral obesity, which is a risk factor for type 2 diabetes mellitus (T2DM). The endocannabinoid system is composed of at least two Gprotein-coupled receptors (GPCRs), the cannabinoid receptor type 1 (CB1), and the cannabinoid receptor type 2 (CB2). Downregulation of CB1 activity in rodents and humans has proven efficacious to reduce food intake, abdominal adiposity, fasting glucose levels, and cardiometabolic risk factors. Unfortunately, downregulation of CB1 activity by universally active CB1 inverse agonists has been found to elicit psychiatric side effects, which led to the termination of using globally active CB1 inverse agonists to treat diet-induced obesity. Interestingly, preclinical studies have shown that downregulation of CB1 activity by CB1 neutral antagonists or peripherally restricted CB1 inverse agonists provided similar anorectic effects and metabolic benefits without psychiatric side effects seen in globally active CB1 inverse agonists. Furthermore, downregulation of CB1 activity may ease endoplasmic reticulum and mitochondrial stress which are contributors to obesity-induced insulin resistance and type 2 diabetes. This suggests new approaches for cannabinoid-based therapy in the management of obesity and obesity-related metabolic disorders including type 2 diabetes.

Antagonism of CB1‐mediated discriminative‐stimulus effects by CB1 neutral and inverse‐agonist antagonists

CB1 inverse‐agonist antagonists (e.g., rimonabant) have behavioral effects that limit their clinical utility. Recent data suggest that CB1 neutral antagonists may not have rimonabant‐like effects, raising the possibility of improved clinical utility. The present studies were conducted to compare how inverse‐agonist antagonists (SR141716A, AM 6538) and neutral antagonists (AM 4113, AM 6545, AM 6527) modify the discriminative‐stimulus effects of the CB1 full agonist AM 4054 (0.01 mg/kg, i.m.) in nonhuman primates (n=4). Results show that each antagonist except AM 6545 produced dose‐related rightward shifts in the AM 4054 dose‐effect curve. Also, antagonist effects of AM 6538 were evident up to 4 days following administration. Pretreatment with AM 6545 (10 mg/kg acutely or daily for 5 days) did not greatly alter AM 4054 discrimination. These results show that: 1) both CB1 neutral and inverse‐agonist antagonists, while differing in potency, can surmountably antagonize CB1 discrimination and 2) AM 6538 has long‐lasting CB1 antagonist actions. The ineffectiveness of AM 6545 agrees with previous reports of its limited brain penetration. The present data indicate that CB1 neutral antagonism is an effective strategy for blocking addiction‐related effects of CB1 agonists, and support the idea that it may be useful in managing cannabis‐related disorders. (Supported by DA023142 and DA007252)

The CB1 inverse agonist AM251, but not the CB1 antagonist AM4113, enhances retention of contextual fear conditioning in rats

The effects of CB1 antagonist/inverse agonists on the acquisition and consolidation of conditioned fear remain uncertain. Recent studies suggest that the CB1 antagonist/inverse agonist AM251 affects acquisition or consolidation of both contextual and discretely cued fear memories. AM251 is frequently referred to as a CB1 antagonist; however in vitro signal transduction assays indicate that this drug also elicits inverse agonist activity at CB1 receptors. The present studies were undertaken to compare the effects of AM251 on conditioned fear with those produced by AM4113, a novel CB1 antagonist with minimal inverse agonist activity. All drugs were administered prior to conditioning. In retention tests conducted two weeks after conditioning, both AM251 (4.0 mg/kg) and AM4113 (6.0 mg/kg)-treated animals exhibited reduced freezing during a conditioned tone cue played within a novel context. In contextual fear retention tests, animals previously treated with 4.0 or 8.0 mg/kg AM251 exhibited enhanced freezing. By contrast, no dose of AM4113 had any significant effect on contextual fear memory, which is consistent with the lower signal transduction activity of AM4113 at CB1 receptors compared to AM251. These results suggest that CB1 neutral antagonists may be less likely than CB1 inverse agonists to facilitate the acquisition or consolidation of contextual fear that may contribute to some clinical disorders.

Toward the design of cannabinoid CB1 receptor inverse agonists and neutral antagonists

AbstractThe cannabinoid CB1 receptor belongs to Class A of the G‐protein‐coupled receptor (GPCR) family. The high constitutive activity of CB1 facilitates inverse agonism at this receptor, and CB1 inverse agonists/antagonists have recently been considered for the treatment of obesity and metabolic syndrome. GPCRs are assumed to have a common topology and to share a common molecular activation mechanism involving their intracellular domains. However, each individual receptor will also have a molecular switch within the ligand binding pocket that is a noncovalent intramolecular interaction in the basal state of the GPCR that must be disrupted to achieve an active state or stabilized to maintain the inactive state. Knowledge of the molecular switch within the ligand binding pocket can greatly facilitate the rational design of inverse agonists and neutral antagonists. This review begins with a brief review on the CB1 receptor and its ligands. The review then focuses on the experimental literature on GPCR structure and activation in Class A receptors. The identification of the molecular switch region in the ligand binding pocket of CB1 (F3.36/W6.48) is discussed and the combined mutation and modeling studies that have led to the identification of interactions key to the inverse agonism of SR141716A are presented. Finally, the development of the first CB1 neutral antagonists based on these modeling/mutation results is discussed. Drug Dev Res 70:585–600, 2009. © 2009 Wiley‐Liss, Inc.

Intracerebroventricular administration of cannabinoid CB1 receptor antagonists AM251 and AM4113 fails to alter food-reinforced behavior in rats

Drugs that interfere with cannabinoid CB1 transmission suppress food-motivated behaviors and may be useful as appetite suppressants, but there is uncertainty about the locus of action for the feeding-suppression effects of these drugs. The present work was conducted to determine if two drugs that interfere with cannabinoid receptor transmission, AM251 and AM4113, have effects on food-reinforced behavior after administration into the lateral ventricle (intracerebroventricular (ICV)). Although systemic administration of both drugs can suppress food-reinforced behavior, neither AM251 (40, 80, and 160 microg) nor AM4113 (60, 120, and 240 microg) administered at various times prior to testing produced any suppression of food-reinforced operant responding on a fixed-ratio 5 schedule. Because the modulation of locomotion by drugs that act on CB1 receptors is hypothesized to be a forebrain effect, these drugs also were assessed for their ability to reverse the locomotor suppression produced by the CB1 agonist AM411. ICV administration of either AM251 or AM4113 reversed the locomotor suppression induced by the CB1 agonist AM411 in the same dose range that failed to produce any effects on feeding. This indicates that both AM4113 and AM251, when administered ICV, can interact with forebrain CB1 receptors and are efficacious on forebrain-mediated functions unrelated to feeding. These results suggest that CB1 neutral antagonists or inverse agonists may not be affecting food-reinforced behavior via interactions with forebrain CB1 receptors located in nucleus accumbens or hypothalamus and that lower brainstem or peripheral receptors may be involved.

The Endocannabinoid System: A Promising Target for the Management of Type 2 Diabetes

Type 2 diabetes is closely related to abdominal obesity and is generally associated with other cardiometabolic risk factors, resulting in a high incidence of cardiovascular complications. Several animal and human observations suggest that the endocannabinoid (EC) system is overactivated in presence of abdominal obesity and/or diabetes, and contributes to disturbances of energy balance and metabolism. Not only it regulates the intake of nutrients through central mechanisms located within the hypothalamus and limbic area, but it also intervenes in transport, metabolism and deposit of the nutrients in the digestive tract, liver, adipose tissue, skeletal muscle, and possibly pancreas. Activation of both central and peripheral CB1 receptors promotes weight gain and associated metabolic changes. Conversely, rimonabant, the first selective CB(1) receptor antagonist in clinical use, has been shown to reduce body weight, waist circumference, triglycerides, blood pressure, insulin resistance and C-reactive protein levels, and to increase HDL cholesterol and adiponectin concentrations in both non-diabetic and diabetic overweight/obese patients. In addition, a 0.5-0.7% reduction in glycated hemoglobin (HbA1c) levels was observed in metformin- or sulfonylurea-treated patients with type 2 diabetes and in drug-naive or insulin-treated diabetic patients. Almost half of metabolic changes occurred beyond weight loss, in agreement with direct peripheral effects. Rimonabant was generally well-tolerated, but with a slightly higher incidence of depressed mood disorders, anxiety, nausea and dizziness compared to placebo. New trials are supposed to confirm the potential role of rimonabant (and other CB1 neutral antagonists or inverse agonists) in overweight/obese patients with type 2 diabetes and high risk cardiovascular disease.