O pioid receptor selective antagonists are important pharmacological probes to study the structureÀfunction relationship of each opioid receptor. 1À3 It has been demonstrated that, for many clinically available opiates, not only their analgesic function but also their side effects (such as addiction and abuse liability, respiratory depression, and tolerance) are primarily due to their interaction with the mu opioid receptor (MOR). 4À6 Yet the lack of a nonpeptidyl, highly selective, and potent MOR antagonist limits our understanding of the structureÀfunction relationship of MOR. Currently available antagonists for the MOR carry certain characteristics that limit their application (Figure 1). For example, cyprodime 7 only possesses a moderate selectivity for the MOR over the delta opioid receptor (DOR) and kappa opioid receptor (KOR) (K i value ratios are kappa/mu ≈ 45, delta/mu ≈ 40) with much lower affinity for the MOR than naloxone and naltrexone.8 β-FNA, clocinnamox, and other irreversible antagonists for the MOR 9À11 bind covalently with the receptor, which largely limits their utility. Some currently available conformation-constrained peptides, for example, CTOP and CTAP, are highly selective and reversible MOR antagonists. They are relatively metabolically stable and have been used to target the MOR in in vitro and in vivo studies, while their limited bioavailability when administered peripherally hindered their potential medical applications. 12À19 Because the utility of antagonists as pharmacological tools requires both in vitro and in vivo activity, nonpeptide ligands are still preferred due to their ability to penetrate the central nervous system (CNS) and lesser vulnerability to metabolic inactivation compared to the peptide agents. Therefore, the development of a nonpeptidyl, potent, selective, and reversible antagonist for the MOR is highly desirable.Recently, based on the "message-address concept" and molecular modeling studies, a series of 6R-and 6β-N-heterocyclic substituted naltrexamine derivatives were designed, synthesized, and characterized. 20 Among them, NAP and NAQ (Figure 1) seemed to be promising leads as MOR selective antagonists. NAP displayed high binding affinity for the MOR at K i = 0.37 nM with over 700-fold selectivity for the MOR over the DOR and more than 150-fold selectivity over the KOR. The binding affinity of NAQ to MOR was 0.55 nM with over 200-fold selectivity for the MOR over the DOR and approximately 50-fold selectivity over the KOR. Meanwhile they were both low efficacy MOR agonists compared with DAMGO in the ABSTRACT: As important pharmacological probes, highly selective opioid receptor antagonists are essential in opioid receptor structural characterization and opioid agonist functional studies. At present, a nonpeptidyl, highly selective, and reversible mu opioid receptor antagonist is still not available. Among a series of novel naltrexamine derivatives that have been designed and synthesized following molecular modeling studies, two compounds, NAP and NAQ, were ...

A 6β-N-heterocyclic substituted naltrexamine derivative, NAP, was proposed as a peripheral mu opioid receptor (MOR) selective antagonist based on the in vitro and in vivo pharmacological and pharmacokinetic studies. To further validate this notion, several functional assays were carried out to fully characterize this compound. In the charcoal gavage and intestinal motility assay in morphine-pelleted mice, when administered 0.3 mg/kg or higher doses up to 3 mg/kg subcutaneously, NAP significantly increased the intestinal motility compared to the saline treatment. The comparative opioid withdrawal precipitation study and the lower locomotor assay demonstrated that NAP showed only marginal intrinsic effect in the central nervous system either given subcutaneously or intravenously: no jumps were witnessed for the tested animals even given up to a dose of 50 mg/kg, while similar noticeable wet-dog shakes only occurred at the dose 50 times of those for naloxone or naltrexone, and significant reduction of the hyper-locomotion only happened at the dose as high as 32 mg/kg. Collectively, these results suggested that NAP may serve as a novel lead to develop peripheral MOR selective antagonist which might possess therapeutic potential for opioid-induced bowel dysfunction (OBD), such as opioid-induced constipation (OIC).

Opiate abuse reportedly can exaggerate complications of human immunodeficiency virus type-1 (HIV-1) infection in the central nervous system (CNS), while opiate drugs are often indicated in the treatment of HIV-1-related neuropathic pain. Despite this quandary, few studies have assessed the relationship between the duration or extent of HIV-1 infection and the intrinsic neurobehavioral responsiveness to opioids. To address this problem, doxycycline (DOX)-inducible HIV-Tat1-86 transgenic mice were used as a model for HIV-1-associated neurocognitive disorders, which permitted the regulation of Tat exposure and duration. The effects of continuous Tat induction on the activity of morphine were examined at weekly intervals using standard behavioral assays for nociception and motor function. In the spinal cord, Tat mRNA levels did not increase until the second and third weeks following induction, which corresponded to a significant loss of morphine antinociception as assessed in the tail-flick test. Alternatively, in the striatum, sustained increases in Tat mRNA expression during the second week of induction coincided with significant decreases in rotarod performance and interactions with morphine. Importantly, the behavioral effects of morphine differed depending on the timing and location of Tat expression; with increases in Tat transcript levels in the spinal cord and striatum corresponding to significant alterations in morphine-dependent nociception and rotarod performance, respectively. Assuming Tat levels contribute to the clinical manifestations of HIV-1, the results suggest that regional differences in viral load and opioid phenotype might influence the nature and degree that opiate responsiveness is altered in HIV-1 infected individuals.

Background and purpose: Chronic morphine administration produces tolerance in vivo and attenuation of m opioid receptor (MOR)-mediated G-protein activation measured in vitro, but the relationship between these adaptations is not clear. The present study examined MOR-mediated G-protein activation in the CNS of mice with different levels of morphine tolerance. Experimental approach: Mice were implanted with morphine pellets, with or without supplemental morphine injections, to induce differing levels of tolerance as determined by a range of MOR-mediated behaviours. MOR function was measured using agonist-stimulatedand receptor binding throughout the CNS. Key results: Morphine pellet implantation produced 6-12-fold tolerance in antinociceptive assays, hypothermia and Straub tail, as measured by the ratio of morphine ED 50 values between morphine-treated and control groups. Pellet implantation plus supplemental injections produced 25-50-fold tolerance in these tests. In morphine pellet-implanted mice, MOR-stimulated [ 35 S]GTPgS binding was significantly reduced only in the nucleus tractus solitarius (NTS) and spinal cord dorsal horn in tissue sections from morphine pellet-implanted mice. In contrast, MOR-stimulated [35 S]GTPgS binding was significantly decreased in most regions examined in morphine pellet þ morphine injected mice, including nucleus accumbens, caudate-putamen, periaqueductal gray, parabrachial nucleus, NTS and spinal cord. Conclusions and implications: Tolerance and the regional pattern of apparent MOR desensitization were influenced positively by the level of morphine exposure. These results indicate that desensitization of MOR-mediated G-protein activity is more regionally widespread upon induction of high levels of tolerance, suggesting that this response contributes more to high than low levels of tolerance to CNS-mediated effects of morphine.

Peripheral selective mu opioid receptor (MOR) antagonists could alleviate the symptoms of opioid-induced constipation (OIC) without compromising the analgesic effect of opioids. However, a variety of adverse effects were associated with them, partially due to their relatively low MOR selectivity. NAP, a 6β-N-4'-pyridyl substituted naltrexamine derivative, was identified previously as a potent and highly selective MOR antagonist mainly acting within the peripheral nervous system. The noticeable diarrhea associated with it prompted the design and synthesis of its analogues in order to study its structure activity relationship. Among them, compound 8 showed improved pharmacological profiles compared to the original lead, acting mainly at peripheral while increasing the intestinal motility in morphine-pelleted mice (ED50=0.03 mg/kg). The slight decrease of the ED50 compared to the original lead was well compensated by the unobserved adverse effect. Hence, this compound seems to be a more promising lead to develop novel therapeutic agents toward OIC.