Cocaine addiction is a global public health and socioeconomic issue that requires pharmacological and cognitive therapies. Currently there are no FDA-approved medications to treat cocaine addiction. However, in preclinical studies, interventions ranging from herbal medicine to deep-brain stimulation have shown promise for the therapy of cocaine addiction. Recent developments in molecular biology, pharmacology, and medicinal chemistry have enabled scientists to identify novel molecular targets along the pathways involved in drug addiction. In 1994, a receptor that showed a great deal of homology to the traditional opioid receptors was characterized. However, endogenous and exogenous opioids failed to bind to this receptor, which led scientists to name it opioid receptor-like receptor, now referred to as the nociceptin receptor. The endogenous ligand of NOPr was identified a year later and named orphanin FQ/nociceptin. Nociceptin and NOPr are widely distributed throughout the CNS and are involved in many physiological responses, such as food intake, nociceptive processing, neurotransmitter release, etc. Furthermore, exogenous nociceptin has been shown to regulate the activity of mesolimbic dopaminergic neurons, glutamate, and opioid systems, and the stress circuit. Importantly, exogenous nociceptin has been shown to reduce the rewarding and addictive actions of a number of drugs of abuse, such as psychostimulants, alcohol, and opioids. This paper reviews the existing literature on the role of endogenous nociceptin in the rewarding and addictive actions of cocaine. The effect of exogenous nociceptin on these processes is also reviewed. Furthermore, the effects of novel small-molecule NOPr ligands on these actions of cocaine are discussed. Overall, a review of the literature suggests that NOPr could be an emerging target for cocaine addiction pharmacotherapy.

The cancer chemopreventive activity of green tea and its major polyphenolic constituent, epigallocatechin-3-gallate (EGCG) have been attributed to its antioxidant, antiproliferative and antiangiogenic effects. Several new molecular targets for EGCG's anticarcinogenic activity have been proposed in the recent literature. However, the understanding of the molecular mechanisms of EGCG's activity in vivo have been confounded by its low oral bioavailability and low plasma levels. Studies of EGCG would be greatly aided by the availability of synthetic analogs of EGCG designed to understand the contributions of the A, B, and D-rings and the phenolic hydroxyl groups of EGCG to its molecular mechanisms of action. We recently reported the de novo synthesis of a D-ring analog of EGCG, with the objective of using such analogs to understand the molecular mechanisms of EGCG action. We report here the first studies with a synthetic D-ring analog of EGCG. We examined the ability of the synthetic D-ring analog to inhibit tumor cell proliferation in breast carcinoma cells. We also investigated the effect of the analog on stress-induced VEGF production in breast carcinoma cells using Northern analysis and quantitative RT-PCR. We report here that the synthetic D-ring analog inhibits breast cancer cell growth in vitro with potencies equivalent to those of EGCG. Our results also show that, like EGCG, the synthetic analog inhibits hypoxia- and serum starvation-induced production of VEGF mRNA in breast cancer cells. Such synthetic analogs are valuable for understanding the structure-function relationship of EGCG and identifying relevant mechanisms of the chemopreventive action of EGCG.

Treatment of drug addiction remains an unmet medical need due to the dearth of approved pharmacotherapies. There are no approved treatments for cocaine addiction, whereas the current opioid crisis has revealed the stark reality of the limited options to treat prescription and illicit opioid abuse. Preclinical studies in rodents and nonhuman primates have shown that orphanin FQ/nociceptin (N/OFQ), the endogenous ligand for the nociceptin opioid receptor (NOP) reduces the rewarding effects of several abused substances, including opioids, psychostimulants and alcohol. A few nonpeptide small-molecule NOP agonists have also shown efficacy in attenuating the rewarding effects of various abused drugs. We previously demonstrated that a high affinity small-molecule NOP agonist AT-312 selectively reduced the rewarding effects of ethanol in the conditioned place preference paradigm in mice. In the present study, we examined if AT-312 (3 mg/kg, i.p. or s.c. respectively), would alter the rewarding action of morphine (7.5 mg/kg, s.c.) or cocaine (15 mg/kg, i.p.). The effect of AT-312 on morphine- and cocaine-induced motor stimulation was also assessed on the conditioning days. The role of the NOP receptor in the effects of AT-312 was further confirmed by conducting the place conditioning experiments in NOP knockout mice and compared to their wild-type controls. Our results showed that AT-312 significantly reduced the acquisition of morphine and cocaine CPP in wild-type mice but not in mice lacking NOP receptors. AT-312 also suppressed morphine-induced and completely abolished cocaine-induced motor stimulation in NOP wild-type mice, but not in NOP knockout mice. These results show that small-molecule NOP receptor agonists have promising efficacy for attenuating the rewarding effects of morphine and cocaine, and may have potential as pharmacotherapy for opioid and psychostimulant addiction or for treating polydrug addiction.

Background

Nociceptin/orphanin FQ, the endogenous peptide agonist for the opioid receptor-like receptor (also known as NOP or the nociceptin receptor), has been shown to block the acquisition and expression of ethanol (EtOH)-induced conditioned place preference (CPP). Here, we report the characterization of a novel small-molecule NOP ligand AT-312 (1-(1-((cis)-4-isopropylcyclohexyl)piperidin-4-yl)-1H-indol-2-yl)methanol) in receptor binding and GTPγS functional assays in vitro. We then investigated the effect of AT-312 on the rewarding action of EtOH in mice using the CPP paradigm. Further, using mice lacking the NOP receptor and their wild-type controls, we also examined the involvement of NOP in the effect of AT-312. Motivational effects of AT-312 alone were also assessed in the CPP paradigm.

Methods

Female mice lacking NOP and/or their wild-type controls received conditioning in the presence or absence of the NOP agonist [AT-312 (1, 3, and 10 mg/kg) or the control NOP agonist SCH221510 (10 mg/kg)] followed by saline/EtOH for 3 consecutive days (twice daily) and tested for CPP in a drug-free state on the next day.

Results

Our in vitro data showed that AT-312 is a high-affinity, selective NOP full agonist with 17-fold selectivity over the mu opioid receptor and >200-fold selectivity over the kappa opioid receptor. The results of our in vivo studies showed that AT-312 reduced EtOH CPP at the lowest dose (1 mg/kg) tested but completely abolished EtOH CPP at higher doses (3 or 10 mg/kg) compared to their vehicle-treated control group. AT-312 (3 mg/kg) did not alter EtOH-induced CPP in mice lacking NOP, confirming that AT-312 reduced EtOH CPP through its action at the NOP receptor. AT-312 (3 mg/kg) did not induce reward or aversion when administered alone, showing that the novel small-molecule NOP agonist shows efficacy in blocking EtOH-induced CPP via the NOP receptor.

Conclusions

Together, these data suggest that small-molecule NOP agonists have the potential to reduce alcohol reward and may be promising as medications to treat alcohol addiction.

The strong reinforcing effects of nicotine and the negative symptoms such as anxiety experienced during a quit attempt often lead to relapse and low success rates for smoking cessation. Treatments that not only block the reinforcing effects of nicotine but also attenuate the motivation to relapse are needed to improve cessation rates. Recent genetic and preclinical studies have highlighted the involvement of the α3, β4, and α5 nicotinic acetylcholine receptor (nAChR) subunits and the α3β4 nAChR subtype in nicotine dependence and withdrawal. However, the involvement of these nAChR in relapse is not fully understood. We previously reported that the α3β4 nAChR partial agonist AT-1001 selectively decreases nicotine self-administration in rats without affecting food responding. In the present experiments, we examined the efficacy of AT-1001 in attenuating reinstatement of nicotine-seeking behavior in a model of stress-induced relapse. Rats extinguished from nicotine self-administration were treated with the pharmacological stressor yohimbine prior to AT-1001 treatment and reinstatement testing. We also examined whether AT-1001 produced any withdrawal-related effects when administered to nicotine-dependent rats. We found that AT-1001 dose-dependently reduced yohimbine stress-induced reinstatement of nicotine seeking. When administered to nicotine-dependent rats at the dose that significantly blocked nicotine reinstatement, AT-1001 elicited minimal somatic withdrawal signs in comparison to the nicotinic antagonist mecamylamine, which is known to produce robust withdrawal. Our data suggest that α3β4 nAChR-targeted compounds may be a promising approach for nicotine addiction treatment because they can not only block nicotine's reinforcing effects, but also decrease motivation to relapse without producing significant withdrawal effects.

Tobacco dependence is difficult to treat, with the vast majority of those who try to quit relapsing within the first year. Improvements in smoking cessation therapies may be achieved by improving current preclinical research methods. However, most experimental tests in animals use nicotine alone, ignoring the 8000 other constituents found in tobacco smoke. To improve on this model, we have used self-administration to test the reinforcing properties of aqueous cigarette smoke extract (CSE) in rats, made by bubbling cigarette smoke through a saline solution. CSE is more potent than nicotine alone in both the acquisition and maintenance of self-administration, but did not exhibit higher progressive ratio responding. Mecamylamine and varenicline had similar potencies to block nicotine and CSE self-administration, indicating the involvement of nicotinic receptors in CSE reinforcement. Following extinction of responding, reinstatement was triggered by exposing animals to a pharmacological stressor, yohimbine (2.5 mg/kg, i.p.), alone and in combination with cues. Animals that self-administered CSE were significantly more sensitive to stress-induced reinstatement than those that self-administered nicotine. Ligand binding autoradiography studies showed nicotine and CSE to have similar affinities for different nicotinic receptor types. CSE significantly reduced MAO-A and MAO-B activities in vitro, whereas nicotine did not. Although CSE inhibition of MAO-A activity in vitro was found to be partially irreversible, irreversible inhibition was not observed in vivo. These experiments show that CSE is an effective reinforcer acting via nicotinic receptors. Furthermore, it better models MAO inhibition and is more sensitive to stress-induced reinstatement than nicotine alone, which is a potent trigger for relapse in smokers.

Despite extensive research, current therapies for smoking cessation are largely ineffective at maintaining abstinence for more than a year. Whereas most preclinical studies use nicotine alone, the goal of the present study was to evaluate whether inclusion of non-nicotine tobacco constituents provides better face validity for the development of new pharmacological therapies for smoking cessation. Here, we trained adult male rats to self-administer nicotine alone or cigarette smoke extract (CSE), which contains nicotine and other aqueous constituents of cigarette smoke. After stable self-administration behavior was established, animals underwent extinction training followed by drug and cue primed reinstatement testing. We show that animals that self-administered CSE had significant reinstatement in all drug and drug + cue stimulus conditions whereas animals that self-administered nicotine only showed significant reinstatement in the drug + cue conditions. AT-1001, an α3β4 nicotinic acetylcholine receptor (nAChR) functional antagonist, attenuated drug + cue-primed reinstatement of both CSE- and nicotine-seeking behavior. However, AT-1001 was less potent in blocking drug-primed reinstatement in animals that had self-administered CSE than in those that had self-administered nicotine alone. This was the case even when nicotine was used to prime reinstatement in animals that had self-administered CSE, suggesting that prior CSE exposure had altered the functional role of α3β4-containing nAChRs in drug-seeking behavior. These findings confirm the importance of non-nicotine tobacco constituents and α3β4* nAChRs in cue- and nicotine-primed craving. They also suggest that tests using CSE may be more valid models to study tobacco dependence than use of nicotine alone.