Design, synthesis, in vitro evaluation, and molecular modeling studies of N-substituted benzomorphans, analogs of LP2, as novel MOR ligands.

6,7-Benzomorphans have been investigated in medicinal chemistry for developing new drugs. This nucleus could be considered a versatile scaffold. The physicochemical properties of benzomorphan N-substituent are crucial in achieving a definite pharmacological profile at opioid receptors. Thus, the dual-target MOR/DOR ligands LP1 and LP2 were obtained through N-substituent modifications. Specifically, LP2, bearing as N-substituent the (2R/S)-2-methoxy-2- phenylethyl group, is a dual-target MOR/DOR agonist and is successful in animal models of inflammatory and neuropathic pain. To obtain new opioid ligands, we focused on the design and synthesis of LP2 analogs. First, the 2-methoxyl group of LP2 was replaced by an ester or acid functional group. Then, spacers of different lengths were introduced at N-substituent. In-vitro, their affinity profile versus opioid receptors has been performed through competition binding assays. Molecular modeling studies were conducted to deeply analyze the binding mode and the interactions between the new ligands and all opioid receptors.

LP1 and LP2: Dual-Target MOPr/DOPr Ligands as Drug Candidates for Persistent Pain Relief.

Although persistent pain is estimated to affect about 20% of the adult population, current treatments have poor results. Polypharmacology, which is the administration of more than one drug targeting on two or more different sites of action, represents a prominent therapeutic approach for the clinical management of persistent pain. Thus, in the drug discovery process the "one-molecule-multiple targets" strategy nowadays is highly recognized. Indeed, multitarget ligands displaying a better antinociceptive activity with fewer side effects, combined with favorable pharmacokinetic and pharmacodynamic characteristics, have already been shown. Multitarget ligands possessing non-opioid/opioid and opioid/opioid mechanisms of action are considered as potential drug candidates for the management of various pain conditions. In particular, dual-target MOPr (mu opioid peptide receptor)/DOPr (delta opioid peptide receptor) ligands exhibit an improved antinociceptive profile associated with a reduced tolerance-inducing capability. The benzomorphan-based compounds LP1 and LP2 belong to this class of dual-target MOPr/DOPr ligands. In the present manuscript, the structure-activity relationships and the pharmacological fingerprint of LP1 and LP2 compounds as suitable drug candidates for persistent pain relief is described.

Exploiting the Power of Stereochemistry in Drug Action: 3-[(2S,6S,11S)-8-Hydroxy-6,11-dimethyl-1,4,5,6-tetrahydro-2,6-methano-3-benzazocin-3(2H)-yl]-N-phenylpropanamide as Potent Sigma-1 Receptor Antagonist.

(+)-(2S,6S,11S)- and (-)-(2R,6R,11R)-Benzomorphan derivatives have a different binding affinity for sigma-1 (σ1R) and opioid receptors, respectively. In this study, we describe the synthesis of the (+)-enantiomer [(+)-LP1] of the benzomorphan MOR agonist/DOR antagonist LP1 [(-)-LP1]. The binding affinity of both (+)-LP1 and (-)-LP1 for σ1R and sigma-2 receptor (σ2R) was tested. Moreover, (+)-LP1 opioid receptor binding affinity was also investigated. Finally, (+)-LP1 was tested in a mouse model of inflammatory pain. Our results showed a nanomolar σ1R and binding affinity for (+)-LP1. Both (+)-LP1 and (-)-LP1 elicited a significant analgesic effect in a formalin test. Differently from (-)-LP1, the analgesic effect of (+)-LP1 was not reversed by naloxone, suggesting a σ1R antagonist profile. Furthermore, σ1R agonist PRE-084 was able to unmask the σ1R antagonistic component of the benzomorphan compound. (+)-LP1 could constitute an useful lead compound to develop new analgesics based on mechanisms of action alternative to opioid receptor activation.

Molecular modeling and biological studies show that some µ-opioid receptor agonists might elicit analgesia acting as MMP-9 inhibitors.

Aim: Despite the serious side effects, analgesics acting on opioid receptors are still considered the best way to get antinociception. Matrix metalloproteinases, a large family of zinc-dependent proteases implicated in many pathological conditions, such as diabetes and osteoarthritis, are also involved in inflammation and pain. Methodology & results: Looking for evidence of possible interactions of opioid pathways and inflammation mediators, molecular modeling studies of a series of recently developed µ-opioid receptor benzomorphanic agonists together with biological data on pain and inflammation molecular targets, allowed us to hypothesize a possible correlation between µ-opioid receptor system and MMP-9. Conclusion: A new compound, (-)-MML1017, emerged as a possible dual-acting agent able to interact selectively and potently with the two molecular targets.

(2S)-N-2-methoxy-2-phenylethyl-6,7-benzomorphan compound (2S-LP2): Discovery of a biased mu/delta opioid receptor agonist.

The pivotal role of the stereocenter at the N-substituent of the 6,7-benzomorphan scaffold was investigated combining synthetic and pharmacological approaches. 2R- and 2S-diastereoisomers of the multitarget MOR/DOR antinociceptive ligand LP2 (1) were synthesized and their pharmacological profile was evaluated in in vitro and vivo assays. From our results, 2S-LP2 (5) showed an improved pharmacological profile in comparison to LP2 (1) and 2R-LP2 (4). 2S-LP2 (5) elicited an antinociceptive effect with a 1.5- and 3-times higher potency than LP2 (1) and R-antipode (4), respectively. In vivo effect of 2S-LP2 (5) was consistent with the improved MOR/DOR efficacy profile assessed by radioligand binding assay, to evaluate the opioid receptor affinity, and BRET assay, to evaluate the capability to promote receptor/G-protein and receptor/ß-arrestin 2 interaction. 2S-LP2 (5) was able to activate, with different efficacy, G-protein pathway over ß-arrestin 2, behaving as biased agonist at MOR and mainly at DOR. Considering the therapeutic potential of both multitarget MOR/DOR agonism and functional selectivity over G-protein, the 2S-LP2 (5) biased multitarget MOR/DOR agonist could provide a safer treatment opportunity.

Benzomorphan skeleton, a versatile scaffold for different targets: A comprehensive review.

Despite the fact that the benzomorphan skeleton has mainly been employed in medicinal chemistry for the development of opioid analgesics, it is a versatile structure. Its stereochemistry, as well as opportune modifications at the phenolic hydroxyl group and at the basic nitrogen, play a pivotal role addressing the benzomorphan-based compounds to a specific target. In this review, we describe the structure activity-relationships (SARs) of benzomorphan-based compounds acting at sigma 1 receptor (σ1R), sigma 2 receptor (σ2R), voltage-dependent sodium channel, N-Methyl-d-Aspartate (NMDA) receptor-channel complex and other targets. Collectively, the SARs data have highlighted that the benzomorphan nucleus could be regarded as a useful template for the synthesis of drug candidates for different targets.

Synthesis and Structure-Activity Relationships of LP1 Derivatives: N-Methyl-N-phenylethylamino Analogues as Novel MOR Agonists.

The opioid pharmacological profile of cis-(-)-N-normetazocine derivatives is deeply affected by the nature of their N-substituents. Here, our efforts were focused on the synthesis and pharmacological evaluation of novel derivatives of the lead LP1, a multitarget opioid analgesic compound featuring an N-phenylpropanamido substituent. LP1 derivatives 5a-d and 6a-d were characterized by flexible groups at the N-substituent that allow them to reposition themselves relative to cis-(-)-N-normetazocine nucleus, thus producing different pharmacological profiles at the mu, delta and kappa opioid receptors (MOR, DOR and KOR) in in vitro and in vivo assays. Among the series, compound 5c, with the best in vitro and in vivo profile, resulted a MOR agonist which displays a KiMOR of 6.1 nM in a competitive binding assay, and an IC50 value of 11.5 nM and an Imax of 72% in measurement of cAMP accumulation in HEK293 cells stably expressing MOR, with a slight lower efficacy than LP1. Moreover, in a mouse model of acute thermal nociception, compound 5c, intraperitoneally administered, exhibits naloxone-reversed antinociceptive properties with an ED50 of 4.33 mg/kg. These results expand our understanding of the importance of N-substituent structural variations in the opioid receptor profile of cis-(-)-N-normetazocine derivatives and identify a new MOR agonist useful for the development of novel opioid analgesics for pain treatment.

An LP1 analogue, selective MOR agonist with a peculiar pharmacological profile, used to scrutiny the ligand binding domain.

The hypothesis that central analgesia with reduced side effects is obtainable by occupying an 'allosteric' site in the MOR ligand binding domain requires the development of new ligands with peculiar pharmacological profile to be used as tools. New benzomorphan derivatives, analogues of LP1, a multitarget MOR agonist/DOR antagonist, were designed to examine in depth MOR ligand binding domain. Compound 5, bearing a diphenylic N-substituent on the benzomorphan nucleus, showed an affinity (Kiµ=0.5±0.2nM) comparable to that of LP1 and a better selectivity versus DOR and KOR. It elicits antinociceptive effects in ex vivo (GPI) and in vivo. This new compound engages receptor amino acidic residues not reached by LP1 and by other established MOR ligands. Molecular modeling studies, conducted on 5 and on several reference compounds, allowed us to propose possible residues in the MOR ligand binding domain essential for their interactions with 'orthosteric' and 'allosteric' binding sites.

Evaluation of N-substituent structural variations in opioid receptor profile of LP1.

The benzomorphan scaffold has great potential as lead structure and the nature of the N-substituent is able to influence affinity, potency, and efficacy at all three opioid receptors. Building upon these considerations, we synthesized a new series of LP1 analogues by introducing naphthyl or heteroaromatic rings in propanamide side chain of its N-substituent (9-15). In vitro competition-binding assays in HEK293 cells stably expressing MOR, DOR or KOR showed that in compound 9 the 1-naphthyl ring led to the retention of MOR affinity (Ki(MOR)=38±4nM) displaying good selectivity versus DOR and KOR. In the electrically stimulated GPI, compound 9 was inactive as agonist but produced an antagonist potency value (pA2) of 8.6 in presence of MOR agonist DAMGO. Moreover, subcutaneously administered it antagonized the antinociceptive effects of morphine with an AD50=2.0mg/kg in mouse-tail flick test. Modeling studies on MOR revealed that compound 9 fit very well in the binding pocket but in a different way in respect to the agonist LP1. Probably the replacement of its N-substituent on the III, IV and V TM domains reflects an antagonist behavior. Therefore, compound 9 could represent a potential lead to further develop antagonists as valid therapeutic agents and useful pharmacological tools to study opioid receptor function.

Norbenzomorphan Framework as a Novel Scaffold for Generating Sigma†2 Receptor/PGRMC1 Subtype-Selective Ligands.

A novel structural class with high affinity and subtype selectivity for the sigma 2 receptor has been discovered. Preliminary structure-affinity relationship data are presented showing that 8-substituted 1,3,4,5-tetrahydro-1,5-methanobenzazepine (norbenzomorphan) derivatives elicit modest to high selectivity for the sigma 2 over the sigma 1 receptor subtype. Indeed, piperazine analogue 8-(4-(3-ethoxy-3-oxopropyl)piperazin-1-yl)-1,3,4,5-tetrahydro-1,5-methanobenzazepine-2-carboxylate (SAS-1121) is 574-fold selective for the sigma 2 over the sigma 1 receptor, thereby establishing it as one of the more subtype-selective sigma 2 binding ligands reported to date. Emerging evidence has implicated the sigma 2 receptor in multiple health disorders, so the drug-like characteristics of many of the selective sigma 2 receptor ligands disclosed herein, coupled with their structural similarity to frameworks found in known drugs, suggest that norbenzomorphan analogues may be promising candidates for further development into drug leads.

Predicted structures for kappa opioid G-protein coupled receptor bound to selective agonists.

Human kappa opioid receptor (κ-OR), a G protein-coupled receptor (GPCR), has been identified as a drug target for treatment of such human disorders as pain perception, neuroendocrine physiology, affective behavior, and cognition. In order to find more selective and active agonists, one would like to do structure based drug design. Indeed, there is an X-ray structure for an antagonist bound to κ-OR, but structures for activated GPCRs are quite different from those for the inactive GPCRs. Here we predict the ensemble of 24 low-energy structures of human kappa opioid receptor (κ-OR), obtained by application of the GEnSeMBLE (GPCR Ensemble of Structures in Membrane Bilayer Environment) complete sampling method, which evaluates 13 trillion combinations of tilt and rotation angles for κ-OR to select the best 24. To validate these structures, we used the DarwinDock complete sampling method to predict the binding sites for five known agonists (ethylketocyclazocine, bremazocine, pentazocine, nalorphine, and morphine) bound to all 24 κ-OR conformations. We find that some agonists bind selectively to receptor conformations that lack the salt bridge between transmembrane domains 3 and 6 as expected for active conformations. These 3D structures for κ-OR provide a structural basis for understanding ligand binding and activation of κ-OR, which should be useful for guiding subtype specific drug design.

The multitarget opioid ligand LP1's effects in persistent pain and in primary cell neuronal cultures.

Persistent pain states, such as those caused by nerve injury or inflammation, are associated with altered sensations, allodynia and hyperalgesia, that are resistant to traditional analgesics. A contribution to development and maintenance in altered pain perception comes from nociceptive processing and descending modulation from supraspinal sites. A multitarget ligand seems to be useful for pain relief with a decreased risk of adverse events and a considerable analgesic efficacy. The multitarget MOR agonist-DOR antagonist LP1, (3-[(2R,6R,11R)-8-hydroxy-6,11-dimethyl-1,4,5,6-tetrahydro-2,6-methano-3-benazocin-3(2H)-yl]-N-phenylpropanamide, is a central acting antinociceptive agent with low potential to induce tolerance. LP1 was tested in models of neuropathic pain - induced by chronic constriction injury (CCI) of the left sciatic nerve - and inflammatory pain - produced by intraplantar injection of carrageenan. In CCI rats, subcutaneous (s.c.) LP1 (3 mg/kg) showed a significant antiallodynic effect, measured with von Frey filaments, and antihyperalgesic effect, evoked in response to a radiant heat stimulus with plantar test. Analogously, LP1 significantly reduced allodynic and hyperalgesic thresholds in a model of inflammatory pain induced by carrageenan. To evaluate the contribution of opioid receptor subtypes in LP1 antinociceptive effects, the multitarget LP1 profile was assessed using selective opioid antagonists. Moreover, functional electrophysiological in vitro assays, using primary cortical and spinal cord networks, allowed to define the "pharmacological fingerprint" of LP1. The EC50 values in this functional screening seem to confirm LP1 as a potent opioid ligand (EC50 = 0.35 fM and EC50 = 44 pM in spinal cord and frontal cortex, respectively). Using a NeuroProof data-base of well characterised reference compounds, a similarity profile of LP1 to opioid and non-opioid drugs involved in pain modulation was detected. Our studies seem to support that multitarget ligand approach should be useful for persistent pain conditions in which mechanical allodynia and thermal hyperalgesia are significant components of the nociceptive response.

Expedient synthesis of norbenzomorphan library via multicomponent assembly process coupled with ring-closing reactions.

A 124-member norbenzomorphan library has been prepared utilizing a novel multicomponent assembly process (MCAP) followed by a variety of ring-closing reactions to generate norbenzomorphan scaffolds that were readily derivatized via a series of aryl halide cross-coupling and nitrogen functionalization reactions. Biological screening has revealed some novel activities that have not been previously associated with this class of compounds.

δ-Opioid receptor agonist SNC80 induces central antinociception mediated by Ca2+ -activated Cl- channels.

OBJECTIVES: The aim of this study was to determine whether Ca(2+)-activated Cl(-) channels (CaCCs) are involved in central antinociception induced by the activation of µ-, δ- and κ-opioid receptors. METHODS: The nociceptive threshold for thermal stimulation was measured using the tail-flick test in Swiss mice. The drugs were administered via the intracerebroventricular route. Probabilities values of P < 0.05 were considered to be statistically significant (analysis of variance/Bonferroni test). KEY FINDINGS: The results demonstrate that exposure to the CaCC blocker niflumic acid (2, 4 and 8 µg) partially reverses the central antinociception induced by the δ-opioid receptor agonist SNC80 ((+)-4-[(αR)-α-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide; 4 µg). In contrast, niflumic acid did not modify the antinociceptive effect of the µ-opioid receptor agonist [D-Ala(2), N-Me-Phe(4), Gly(5)-ol]-enkephalin (0.5 µg) or κ-opioid receptor agonist bremazocine (4 µg). CONCLUSIONS: These data provide evidence for the involvement of CaCCs in δ-opioid receptor-induced central antinociception resulting from receptor activation by the agonist SNC80. CaCC activation does not appear to be involved when µ- and κ-opioid receptors are activated.

The benzomorphan-based LP1 ligand is a suitable MOR/DOR agonist for chronic pain treatment.

AIMS: Powerful analgesics relieve pain primarily through activating mu opioid receptor (MOR), but the long-term use of MOR agonists, such as morphine, is limited by the rapid development of tolerance. Recently, it has been observed that simultaneous stimulation of the delta opioid receptor (DOR) and MOR limits the incidence of tolerance induced by MOR agonists. 3-[(2R,6R,11R)-8-hydroxy-6,11-dimethyl-1,4,5,6-tetrahydro-2,6-methano-3-benzazocin-3(2H)-yl]-N-phenylpropanamide (LP1) is a centrally acting agent with antinociceptive activity comparable to morphine and is able to bind and activate MOR and DOR. The aim of this work was to evaluate and compare the induction of tolerance to antinociceptive effects from treatment with LP1 and morphine. MAIN METHODS: Here, we evaluated the pharmacological effects of LP1 administered at a dose of 4 mg/kg subcutaneously (s.c.) twice per day for 9 days to male Sprague-Dawley rats. In addition, the LP1 mechanism of action was assessed by measurement of LP1-induced [(35)S]GTPγS binding to the MOR and DOR. KEY FINDINGS: Data obtained from the radiant heat tail flick test showed that LP1 maintained its antinociceptive profile until the ninth day, while tolerance to morphine (10mg/kg s.c. twice per day) was observed on day 3. Moreover, LP1 significantly enhanced [(35)S]GTPγS binding in the membranes of HEK293 cells expressing either the MOR or the DOR. SIGNIFICANCE: LP1 is a novel analgesic agent for chronic pain treatment, and its low tolerance-inducing capability may be correlated with its ability to bind both the MOR and DOR.

Evaluation of N-substitution in 6,7-benzomorphan compounds.

6,7-benzomorphan derivatives, exhibiting different mu, delta, and kappa receptor selectivity profiles depending on the N-substituent, represent a useful skeleton for the synthesis of new and better analgesic agents. In this work, an aromatic ring and/or alkyl residues have been used with an N-propanamide or N-acetamide spacer for the synthesis of a new series of 5,9-dimethyl-2'-hydroxy-6,7-benzomorphan derivatives (12-22). Data obtained by competition binding assays showed that the mu opioid receptor seems to prefer an interaction with the 6,7-benzomorphan ligands having an N-substituent with a propanamide spacer and less hindered amide. Highly stringent features are required for delta receptor interaction, while an N-acetamide spacer and/or bulkier amide could preferentially lead to kappa receptor selectivity. In the propanamide series, compound 12 (named LP1) displayed high mu affinity (Ki=0.83 nM), good delta affinity (Ki=29 nM) and low affinity for the kappa receptor (Ki=110 nM), with a selectivity ratio delta/mu and kappa/mu of 35.1 and 132.5, respectively. Further, in the adenylyl cyclase assay, LP1 displayed a mu/delta agonist profile, with IC50 values of 4.8 and 12 nM at the mu and delta receptors, respectively. The antinociceptive potency of LP1 in the tail-flick test after sc administration in rat was comparable with the potency of morphine (ED50=2.03 and 2.7 mg/kg, respectively), and was totally reversed by naloxone. LP1, possessing a mu/delta agonist profile, could represent a lead in further developing benzomorphan-based ligands with potent in vivo analgesic activity and a reduced tendency to induce side effects.

Effects of atypical kappa-opioid receptor agonists on intrathecal morphine-induced itch and analgesia in primates.

Itch/pruritus is the most common side effect associated with spinal administration of morphine given to humans for analgesia. The aim of this study was to investigate the effectiveness of kappa-opioid receptor (KOR) agonists with diverse chemical structures as antipruritics and to elucidate the receptor mechanism underlying the antipruritic effect in monkeys. In particular, previously proposed non-KOR-1 agonists, including nalfurafine [TRK-820, 17-cyclopropylmethyl-3,14 beta-dihydroxy-4,5 alpha-epoxy-6 beta-[N-methyl-trans-3-(3-furyl)acrylamido]morphinan], bremazocine [(+/-)-6-ethyl-1,2,3,4,5,6-hexahydro-3-[(1-hydroxycyclopropy)-methyl]-11,11-dimethyl-2,6-methano-3-benzazocin-8-ol], and GR 89696 [4-[(3,4-dichlorophenyl)acetyl]-3-(1-pyrrolidinylmethyl)-1-piperazinecarboxylic acid methyl ester] were studied in various behavioral assays for measuring itch/scratching, analgesia, and respiratory depression. Systemic administration of nalfurafine (0.1-1 microg/kg), bremazocine (0.1-1 microg/kg), or GR 89696 (0.01-0.1 microg/kg) dose-dependently attenuated intrathecal morphine (0.03 mg)-induced scratching responses without affecting morphine antinociception. The combination of intrathecal morphine with these KOR agonists did not cause sedation. In addition, pretreatment with effective antiscratching doses of nalfurafine, bremazocine, or GR 89696 did not antagonize systemic morphine-induced antinociception and respiratory depression. The dose-addition analysis revealed that there is no subadditivity for nalfurafine in combination with morphine in the antinociceptive effect. Furthermore, the KOR antagonist study revealed that antiscratching effects of both nalfurafine and a prototypical KOR-1 agonist, U-50488H [trans-(+/-)-3,4-dichloro-N-methyl-N-(2-[1-pyrrolidinyl]-cyclohexyl)-benzeneacetamide], could be blocked completely by a selective KOR antagonist, nor-binaltorphimine (3 mg/kg). These findings suggest that the agonist action on KOR mainly contributes to the effectiveness of these atypical KOR agonists as antipruritics, and there is no evidence for KOR subtypes or mu-opioid antagonist action underlying the effects of these KOR agonists. This mechanism-based study further supports the clinical potential of KOR agonists as antipruritics under the context of spinal opioid analgesia.

Acute delta- and kappa-opioid agonist pretreatment potentiates opioid antagonist-induced suppression of water consumption.

The primary objective of this study was to determine whether pretreatment with kappa- and delta-opioid agonists potentiates naltrexone-induced suppression of water consumption following 24h of deprivation. This study also examined the temporal effects of agonist-induced antinociception using the tail-flick and hot-plate tests. Adult male Sprague-Dawley rats were water deprived 20 h and then given an injection (s.c. or i.c.) of an opioid agonist or saline. Drugs included the mu-opioid agonists morphine and DAMGO ([d-Ala2,NMePhe4,Gly-ol5]-enkephalin), the kappa-opioid agonists spiradoline, bremazocine, and U69,593, and the delta-opioid agonists BW 373U86 and DPDPE ([D-Pen2, D-Pen5]-enkephalin). Three hours and forty-five minutes later, animals received a single dose of naltrexone (0.1-30 mg/kg, s.c.) or saline. Fifteen minutes later, animals were allowed free access to water for 30 min. For the tail-flick and hot-plate tests, animals were given a single injection of agonist and tested in both procedures every 30 min for up to 2h, then hourly up to 6h post-injection. Naltrexone dose-dependently suppressed fluid consumption 24h after deprivation. The effects of naltrexone on drinking were potentiated following pretreatment with at least one dose of the agonists tested except BW 373U86. With the exception of BW 373U86, DAMGO, and DPDPE, all of the opioid agonists produced significant antinociception in the hot-plate test. Only BW 373U86 failed to have an antinociceptive effect in the tail-flick test. By 4h after treatment, drug-induced antinociception had largely waned, suggesting the potentiation of naltrexone-induced drinking suppression was not a result of a direct interaction with the agonists. In conclusion, kappa-opioid and delta-opioid receptors appear to contribute to the manifestation of acute opioid dependence, albeit to a lesser degree than mu-opioid receptors.

The mu-opioid receptor agonist morphine, but not agonists at delta- or kappa-opioid receptors, induces peripheral antinociception mediated by cannabinoid receptors.

BACKGROUND AND PURPOSE: Although participation of opioids in antinociception induced by cannabinoids has been documented, there is little information regarding the participation of cannabinoids in the antinociceptive mechanisms of opioids. The aim of the present study was to determine whether endocannabinoids could be involved in peripheral antinociception induced by activation of mu-, delta- and kappa-opioid receptors. EXPERIMENTAL APPROACH: Nociceptive thresholds to mechanical stimulation of rat paws treated with intraplantar prostaglandin E2 (PGE2, 2 microg) to induce hyperalgesia were measured 3 h after injection using an algesimetric apparatus. Opioid agonists morphine (200 microg), (+)-4-[(alphaR)-alpha-((2S,5R)-4-Allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80) (80 microg), bremazocine (50 microg); cannabinoid receptor antagonists N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251) (20-80 microg), 6-iodo-2-methyl-1-[2-(4-morpholinyl)ethyl]-1H-indol-3-yl(4-methoxyphenyl) methanone (AM630) (12.5-100 microg); and an inhibitor of methyl arachidonyl fluorophosphonate (MAFP) (1-4 microg) were also injected in the paw. KEY RESULTS: The CB1-selective cannabinoid receptor antagonist AM251 completely reversed the peripheral antinociception induced by morphine in a dose-dependent manner. In contrast, the CB2-selective cannabinoid receptor antagonist AM630 elicited partial antagonism of this effect. In addition, the administration of the fatty acid amide hydrolase inhibitor, MAFP, enhanced the antinociception induced by morphine. The cannabinoid receptor antagonists AM251 and AM630 did not modify the antinociceptive effect of SNC80 or bremazocine. The antagonists alone did not cause any hyperalgesic or antinociceptive effect. CONCLUSIONS AND IMPLICATIONS: Our results provide evidence for the involvement of endocannabinoids, in the peripheral antinociception induced by the mu-opioid receptor agonist morphine. The release of cannabinoids appears not to be involved in the peripheral antinociceptive effect induced by kappa- and delta-opioid receptor agonists.

The influence of esters and carboxylic acids as the N-substituent of opioids. Part 1: Benzomorphans.

To investigate the effects of carboxylic ester and acid moieties as the N-substituent of opioids, a short series of racemic N-substituted normetazocines was prepared. The introduction of both groups as the normetazocine N-substituent produced compounds which displayed low potency in vitro and in vivo, with the esters displaying the greater activity. The pharmacology of the compounds is discussed with implications resulting from potential in vivo metabolic hydrolysis.