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In a previous communication, our efforts leading from 1 to the identification of spiro[cyclohexane-dihydropyrano[3,4-b]indole]-amine 2a as analgesic NOP and opioid receptor agonist were disclosed and their favorable in vitro and in vivo pharmacological properties revealed. We herein report our efforts to further optimize lead 2a, toward trans-6'-fluoro-4',9'-dihydro-N,N-dimethyl-4-phenyl-spiro[cyclohexane-1,1'(3'H)-pyrano[3,4-b]indol]-4-amine (cebranopadol, 3a), which is currently in clinical development for the treatment of severe chronic nociceptive and neuropathic pain.

We report the discovery of spiro[cyclohexane-pyrano[3,4-b]indole]-amines, as functional nociceptin/orphanin FQ peptide (NOP) and opioid receptor agonists with strong efficacy in preclinical models of acute and neuropathic pain. Utilizing 4-(dimethylamino)-4-phenylcyclo-hexanone 1 and tryptophol in an oxa-Pictet-Spengler reaction led to the formation of spiroether 2, representing a novel NOP and opioid peptide receptor agonistic chemotype. This finding initially stems from the systematic derivatization of 1, which resulted in alcohols 3-5, ethers 6 and 7, amines 8-10, 22-24, and 26-28, amides 11 and 25, and urea 12, many with low nanomolar binding affinities at the NOP and mu opioid peptide (MOP) receptors.

Cebranopadol (trans-6'-fluoro-4',9'-dihydro-N,N-dimethyl-4-phenyl-spiro[cyclohexane-1,1'(3'H)-pyrano[3,4-b]indol]-4-amine) is a novel analgesic nociceptin/orphanin FQ peptide (NOP) and opioid receptor agonist [Ki (nM)/EC50 (nM)/relative efficacy (%): human NOP receptor 0.9/13.0/89; human mu-opioid peptide (MOP) receptor 0.7/1.2/104; human kappa-opioid peptide receptor 2.6/17/67; human delta-opioid peptide receptor 18/110/105]. Cebranopadol exhibits highly potent and efficacious antinociceptive and antihypersensitive effects in several rat models of acute and chronic pain (tail-flick, rheumatoid arthritis, bone cancer, spinal nerve ligation, diabetic neuropathy) with ED50 values of 0.5-5.6 µg/kg after intravenous and 25.1 µg/kg after oral administration. In comparison with selective MOP receptor agonists, cebranopadol was more potent in models of chronic neuropathic than acute nociceptive pain. Cebranopadol's duration of action is long (up to 7 hours after intravenous 12 µg/kg; >9 hours after oral 55 µg/kg in the rat tail-flick test). The antihypersensitive activity of cebranopadol in the spinal nerve ligation model was partially reversed by pretreatment with the selective NOP receptor antagonist J-113397[1-[(3R,4R)-1-cyclooctylmethyl-3-hydroxymethyl-4-piperidyl]-3-ethyl-1,3-dihydro-2H-benzimidazol-2-one] or the opioid receptor antagonist naloxone, indicating that both NOP and opioid receptor agonism are involved in this activity. Development of analgesic tolerance in the chronic constriction injury model was clearly delayed compared with that from an equianalgesic dose of morphine (complete tolerance on day 26 versus day 11, respectively). Unlike morphine, cebranopadol did not disrupt motor coordination and respiration at doses within and exceeding the analgesic dose range. Cebranopadol, by its combination of agonism at NOP and opioid receptors, affords highly potent and efficacious analgesia in various pain models with a favorable side effect profile.

Many opioid analgesics share common structural elements; however, minor differences in structure can result in major differences in pharmacological activity, pharmacokinetic profile, and clinical efficacy and tolerability.This review compares and contrasts the chemistry, pharmacodynamics, pharmacokinetics, and CNS 'functional activity' of tapentadol and tramadol, responsible for their individual clinical utilities.The distinct properties of tapentadol and tramadol generate different CNS functional activities, making each drug the prototype of different classes of opioid/nonopioid analgesics. Tramadol's analgesia derives from relatively weak µ-opioid receptor (MOR) agonism, plus norepinephrine and serotonin reuptake inhibition, provided collectively by the enantiomers of the parent drug and a metabolite that is a stronger MOR agonist, but has lower CNS penetration. Tapentadol's MOR agonist activity is several-fold greater than tramadol's, with prominent norepinephrine reuptake inhibition and minimal serotonin effect. Accordingly, tramadol is well-suited for pain conditions for which a strong opioid component is not needed-and it has the benefit of a low abuse potential; whereas tapentadol, a schedule-II controlled substance, is well-suited for pain conditions requiring a strong opioid component-and it has the benefit of greater gastrointestinal tolerability compared to classical strong opioids. Both drugs offer distinct and complementary clinical options.

To discover novel delta-opioid receptor ligands derived from SNC80 (1), a series of 6,8-diazabicyclo[3.2.2]nonane derivatives bearing two aromatic moieties was prepared, and the affinity toward delta, mu, and kappa receptors, as well as sigma receptors, was investigated. After removal of the 4-methoxybenzyl and 2,4-dimethoxybenzyl protecting groups, the pharmacophoric N,N-diethylcarbamoylbenzyl residue was attached to the 6,8-diazabicyclo[3.2.2]nonane framework to yield the designed delta receptor ligands. In a first series of compounds the benzhydryl moiety of SNC80 was dissected, and one phenyl ring was attached to the bicyclic framework. In a second series of delta ligands the complete benzhydryl moiety was introduced into the bicyclic scaffold. The determined delta receptor affinities show that compounds based on an (R)-glutamate-derived bicyclic scaffold possess higher delta receptor affinity than their (S)-glutamate-derived counterparts. Furthermore, an intact benzhydryl moiety leads to delta receptor ligands that are more potent than compounds with two separated aromatic moieties. Compound 24, with the same spatial arrangement of substituents around the benzhydryl stereocenter as SNC80, shows the highest delta receptor affinity of this series: K(i)=24 nM. Whereas the highly potent delta ligands reveal good selectivity against mu and kappa receptors, the sigma(1) and/or sigma(2) affinities of some compounds are almost in the same range as their delta receptor affinities, such as compound 25 (sigma(2): K(i)=83 nM; delta: K(i)=75 nM). In [(35)S]GTPgammaS assays the most potent delta ligands 24 and 25 showed almost the same intrinsic activity as the full agonist SNC80, proving the agonistic activity of 24 and 25. The enantiomeric 4-benzylidene derivatives 15 and ent-15 showed selective cytotoxicity toward the 5637 (bladder) and A-427 (small-cell lung) human tumor cell lines.

Directed by molecular modeling studies the pharmacophoric benzhydryl moiety of the delta opioid receptor agonist SNC80 was separated and the two phenyl residues were attached to different positions of the conformationally constrained 6,8-diazabicyclo[3.2.2]nonane framework in order to find novel delta agonists. The crucial reaction step in the chiral pool synthesis was the establishment of the three carbon bridge by a Dieckmann analogous cyclization of the allyl and propyl derivatives 6 and 7 to yield the mixed methyl silyl acetals 8 and 9, respectively. Stereoselective Grignard reaction, dehydration, and introduction of the pharmacophoric (N,N-diethylcarbamoylbenzyl) residue led to the designed delta receptor agonists 3, ent-3, and 20 with a double bond in the bicyclic framework. Hydrogenation of the allyl derivative 14 was performed with ammonium formate and Pd/C to yield the saturated ligands 24a and 24b. Removal of the allyl substituent with RhCl(3), hydrogenation of the ring system, and re-attachment of the allyl moiety provided the allyl derivatives 4a and 4b. In receptor binding studies with the radioligand [(3)H]-deltorphine II only ent-3 showed considerable delta receptor affinity (K(i)=740 nM). Since ent-3 also interacts with mu receptors (K(i)=250 nM) it belongs to the very interesting compound class of mixed delta/mu ligands.

Clinical reports have described a long-lasting relief in neuropathic pain patients treated with NMDA receptor antagonists; it is unclear, however, what mediates this effect. In this work, we have used two NMDA antagonists of different class to investigate if the antiallodynic effects in a rat neuropathy model can outlast their in vivo NMDA antagonism. Both the uncompetitive NMDA antagonist ketamine and the glycine(B) antagonist MRZ 2/576 inhibited neuronal responses to iontophoretic NMDA in anaesthetised rats with the time course consistent with their known pharmacokinetics (t(1/2) approximately 10-12min, similar in control and nerve-injured rats). Surprisingly, the antiallodynic effects of the same doses of the NMDA antagonists in the neuropathic pain model were long-lasting (>3h with ketamine, >24h with MRZ 2/576). The effect of ketamine was further prolonged (>24h) when combined with a short-acting opioid, fentanyl, which only produced a short effect ( approximately 40min) when given alone. The duration of centrally mediated side effects of ketamine and MRZ 2/576 was short, similar to the in vivo NMDA antagonism. We speculate that NMDA receptor blockade may down-regulate the central sensitisation triggered by nerve injury, resulting in a long-lasting antiallodynic effect. Development of short-acting NMDA antagonists could represent a strategy for improving their tolerability.

NMDA receptors are implicated in central sensitisation underlying chronic pain, and NMDA antagonists have a potential for the treatment of neuropathic pain. Functional NMDA receptors are also present on primary afferents, where they may play a role in pro-nociceptive plasticity. The importance of this mechanism in neuropathic pain remains unclear. In the present work, we have compared in models of chronic pain the effects of NMDA antagonists at the glycine(B) site with different central access. L-701,324 (the centrally active antagonist) and 5,7-dichlorokynurenic acid (5,7-DCK, known to have limited central access) were tested after systemic administration in rats in the formalin test and in two models of neuropathic pain. The ability of these compounds to exert central actions (sedation, ataxia) was tested in the open field locomotion test; central NMDA antagonism in vivo was tested in anaesthetised rats on responses of spinal cord neurones to iontophoretic NMDA. Both L-701,324 (2.15-21.5 mg/kg i.p.) and 5,7-DCK (10-46.4 mg/kg i.v.) dose-dependently inhibited Phase II of formalin-evoked behaviour. Likewise, both compounds reversed cold allodynia in the chronic constriction injury model and tactile allodynia in animals with spinal nerve ligation. However, only L-701,324 was able to inhibit neuronal responses to NMDA in the antihyperalgesic dose range; 5,7-DCK was inactive on NMDA responses up to 46.4 mg/kg i.v. or 68.1 mg/kg i.p. Consistent with the lack of inhibition of central NMDA-evoked activity, 5,7-DCK did not alter spontaneous behaviour in the open field test, whereas it was significantly inhibited by L-701,324. Thus, peripheral NMDA receptors may substantially contribute to the efficacy of NMDA antagonists in neuropathic pain.

The neuropeptide nociceptin/orphanin FQ (N/OFQ), the endogenous ligand of the opioid receptor-like 1 (ORL1) receptor, has been shown to play a prominent role in the regulation of several biological functions such as pain and stress. Here we describe the isolation and characterization of N/OFQ binding biostable RNA aptamers (Spiegelmers) using a mirror-image in vitro selection approach. Spiegelmers arel-enantiomeric oligonucleotide ligands that display high affinity and specificity to their targets and high resistance to enzymatic degradation compared tod-oligonucleotides. A representative Spiegelmer from the selections performed was size-minimized to two distinct sequences capable of high affinity binding to N/OFQ. The Spiegelmers were shown to antagonize binding of N/OFQ to the ORL1 receptor in a binding-competition assay. The calculated IC50values for the Spiegelmers NOX 2149 and NOX 2137a/b were 110 nM and 330 nM, respectively. The competitive antagonistic properties of these Spiegelmers were further demonstrated by their effective and specific inhibition of G-protein activation in two additional models. The Spiegelmers antagonized the N/OFQ-induced GTPγS incorporation into cell membranes of a CHO-K1 cell line expressing the human ORL1 receptor. In oocytes fromXenopus laevis,NOX 2149 showed an antagonistic effect to the N/OFQ-ORL 1 receptor system that was functionally coupled with G-protein-regulated inwardly rectifying K+channels.

5-Pyrrolidinyl substituted perhydroquinoxalines were designed as conformationally restricted κ-opioid receptor agonists restricted to the periphery. The additional N atom of the quinoxaline system located outside the ethylenediamine κ pharmacophore allows the fine-tuning of the pharmacodynamic and pharmacokinetic properties. The perhydroquinoxalines were synthesized stereoselectively using the concept of late stage diversification of the central building blocks 14. In addition to high κ-opioid receptor affinity they demonstrate high selectivity over μ, δ, σ1, σ2, and NMDA receptors. In the [35S]GTPγS assay full agonism was observed. Because of their high polarity, the secondary amines 14a (log D7.4=0.26) and 14b (log D7.4=0.21) did not penetrate an artificial blood-brain barrier. 14b was able to inhibit the spontaneous pain reaction after rectal mustard oil application to mice (ED50=2.35 mg/kg). This analgesic effect is attributed to activation of peripherally located κ receptors, since 14b did not affect centrally mediated referred allodynia and hyperalgesia.

The N-methyl-D-aspartate (NMDA) antagonist ifenprodil and several structurally related compounds are highly selective for the NR2B-containing receptor subtype. This selectivity could provide an explanation for the reported difference of the analgesic and side-effect profile of ifenprodil-like compounds from other NMDA antagonists. In this work, we have queried if the ifenprodil-induced antinociception can be attributed to the block of NMDA receptors in the spinal cord. Ifenprodil and some other NMDA antagonists (MK-801, memantine) were tested in a model of inflammatory pain (Randall-Selitto) in rats. The in vivo NMDA antagonism was assessed in anaesthetised rats on responses of spinal dorsal horn (DH) neurones to iontophoretic NMDA and in the model of single motor unit (SMU) wind-up. Ifenprodil, MK-801 and memantine dose-dependently increased nociceptive thresholds in the Randall-Selitto model. Antinociceptive doses of the channel blockers selectively antagonised NMDA responses of DH neurones and inhibited wind-up. In contrast, antinociceptive doses of ifenprodil did not show any NMDA antagonism in electrophysiological tests. Although ifenprodil did not inhibit the SMU responses to noxious stimuli in spinalised rats, it markedly and dose-dependently inhibited nociceptive SMU responses in sham-spinalised rats. These results argue against the spinal cord being the principal site of antinociceptive action of ifenprodil; supraspinal structures seem to be involved in this effect.

The d-enantiomer of the opioid methadone is a weak opioid with low micromolar affinity to the N-methyl-D-aspartate (NMDA) receptor. We have investigated the antinociception and in vivo NMDA antagonism after systemic administration of d-methadone in the rat spinal cord. d-Methadone caused antinociception in the Randall-Selitto model of inflammatory pain and inhibited the responses of hindlimb single motor units to noxious electrical and mechanical stimulation (ED(50) 6.6, 6.8 and 7.2 mg/kg intravenous (i.v.), respectively); the wind-up of these responses was only inhibited at the dose almost completely abolishing the baseline responses. d-Methadone inhibited the activity of spinal dorsal horn neurones evoked by both iontophoretic NMDA and (R, S)-alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA, ED(50) 5.7 and 8.2 mg/kg i.v., respectively). After pre-treatment with naloxone, d-methadone was unable to inhibit nociception in the Randall-Selitto model, the NMDA- or AMPA-evoked neuronal activity or the motoneurone wind-up. Thus, in the antinociceptive dose range, the NMDA antagonism does not appear to contribute to the mechanism of d-methadone antinociception.

The role of the opioid-like receptor 1 (ORL1) and its endogenous ligand, nociceptin/orphanin FQ (N/OFQ), in nociception, anxiety, and learning remains to be defined. To allow the rapid identification of agonists and antagonists, a reporter gene assay has been established in which the ORL1 receptor is functionally linked to the cyclic AMP-dependent expression of luciferase. N/OFQ and N/OFQ(1-13)NH(2) inhibited the forskolin-induced luciferase gene expression with IC(50) values of 0.81 +/- 0.5 and 0.87 +/- 0.16 nM, respectively. Buprenorphine was identified as a full agonist at the ORL1 receptor with an IC(50) value of 8.4 +/- 2.8 nM. Fentanyl and 7-benzylidenenaltrexone displayed a weak agonistic activity. The ORL1 antagonist [Phe(1)Psi(CH(2)-NH)Gly(2)]N/OFQ((1-13))NH(2) clearly behaved as an agonist in this assay with an IC(50) value of 85 +/- 47 nM. Thus, there is still a need for antagonistic tool compounds that might help to elucidate the neurophysiological role of N/OFQ.

The involvement of endogenous galanin to antinociception elicited by intrathecally (i.t.) or systemically administered drugs from different chemical and therapeutic classes was investigated using the rat Randall-Selitto or the rat tail-flick test, in the absence or presence of the i.t. administered galanin receptor antagonists galantide and M-35. Antinociception elicited by i.t. tramadol (24 micrograms), DAMGO (1 microgram), clonidine (48 micrograms), desipramine (6 micrograms) or fenfluramine (60 micrograms) was attenuated by i.t. galantide (2 micrograms); the attenuation reached significance at least at one time point. A partial antagonism by i.t. galantide was also observed against the antinociception of i.p. tramadol (10 mg/kg), i.v. clonidine (1 mg/kg), i.p. desipramine (1 mg/kg), or i.p. dipyrone (1000 mg/kg), but antinociception by i.p. fenfluramine (30 mg/kg) was not affected. Using M-35 (2 micrograms i.t.), the antinociception of i.t. tramadol or DAMGO was attenuated, but no inhibition was observed when clonidine, desipramine or fenfluramine were used i.t. If drugs were administered systemically, only antinociception of i.p. fenfluramine but not that of i.p. tramadol, or i.v. clonidine, or i.p. desipramine or i.p dipyrone was attenuated. In the rat tail flick test, co-injection of either 2 micrograms i.t. galantide or M-35 with i.t. tramadol (12 micrograms) almost abolished the antinociceptive effect, whereas the antinociception of systemically administered tramadol (4.6 mg/kg i.p.) was only partially attenuated by i.t. galantide and not affected by i.t. M-35. Binding studies in dorsal spinal cord tissue showed no affinity of galantide or M-35 to spinal mu-, or delta-, or kappa-opioid receptors and none of the other drugs interfered with the spinal galanin binding site. These data give further support of at least a partial galanin link in spinal processes of antinociception.

The stereochemistry the 2,4-di-arene substituted 3,7-diazabicyclo[3.3.1]nonan-9-one 1,5-dicarboxylate skeleton was found to be regulated by the kind of substituents attached to the arene rings as well as to the nitrogens N3 and N7. Conformational isomers, i.e., chair/chair, boat/chair and chair/boat, in addition to cis/trans conflgurational isomerism with respect to the arene rings were reported. Since the analgesic potency of the diazabicyclononanones, which is related to their affinity toward the κ-opioid receptor, is governed by the stereochemistry of the molecules, the influence of the substituents at nitrogen N7 was studied herein. The various differently N7 substituted diazabicyclononanones were found to crystallise in a highly symmetrical chair/chair conformation. However, beside HZ2 none of the compounds exhibits high affinity to the κ receptor. In contrast, some compounds with affinity to the μ receptor could be identified. In addition, the N7-(4-carboxybenzyl) substituted compound was found to have affinity to the δ receptor in the submi-cromolar range of concentration.

Three sets of stereoisomeric bicyclic kappa agonists with defined orientation of the pharmacophoric elements pyrrolidine and dichlorophenylacetamide were stereoselectively prepared and pharmacologically evaluated. Stereoselective reduction, reductive amination, and Mitsunobu inversions were the key steps for the establishment of the desired stereochemistry. The kappa affinity decreased in the following order depending on the N-substituent: CO(2)CH(3)benzylCOCH(2)CH(3). Bicyclic derivatives with (1S,2R,5R)-configuration showed the highest kappa receptor affinity, which led to dihedral angles of 97 degrees and 45 degrees for the N(pyrrolidine)-C-C-N(phenylacetamide) structural element. The most potent kappa agonist of this series was (+)-methyl (1S,2R,5R)-8-[2-(3,4-dichlorophenyl)acetyl]-2-(pyrrolidin-1-yl)-6,8-diazabicyclo[3.2.2]nonane-6-carboxylate (ent-23, WMS-0121) with an K(i) value of 1.0 nM. ent-23 revealed high selectivity against the other classical opioid receptors and related receptor systems. In the [(35)S]GTPgammaS-binding assay at human kappa-opioid receptors, ent-23 was proved to be a full agonist with the same EC(50) value (87 nM) as the prototypical full agonist U-69,593 (EC(50) = 80 nM).

Tapentadol exerts its analgesic effects through micro opioid receptor agonism and noradrenaline reuptake inhibition in the central nervous system. Preclinical studies demonstrated that tapentadol is effective in a broad range of pain models, including nociceptive, inflammatory, visceral, mono- and polyneuropathic models. Moreover, clinical studies showed that tapentadol effectively relieves moderate to severe pain in various pain care settings. In addition, it was reported to be associated with significantly fewer treatment discontinuations due to a significantly lower incidence of gastrointestinal-related adverse events compared with equivalent doses of oxycodone. The combination of these reduced treatment discontinuation rates and tapentadol efficacy for the relief of moderate to severe nociceptive and neuropathic pain may offer an improvement in pain therapy by increasing patient compliance with their treatment regimen.

(–)-(1 R ,2 R )-3-(3-Dimethylamino-1-ethyl-2-methyl-propyl)-phenol hydrochloride (tapentadol HCl) is a novel μ-opioid receptor (MOR) agonist ( K i = 0.1 μM; relative efficacy compared with morphine 88% in a [ 35 S]guanosine 5′-3- O -(thio)triphosphate binding assay) and NE reuptake inhibitor ( K i = 0.5 μM for synaptosomal reuptake inhibition). In vivo intracerebral microdialysis showed that tapentadol, in contrast to morphine, produces large increases in extracellular levels of NE (+450% at 10 mg/kg i.p.). Tapentadol exhibited analgesic effects in a wide range of animal models of acute and chronic pain [hot plate, tail-flick, writhing, Randall-Selitto, mustard oil colitis, chronic constriction injury (CCI), and spinal nerve ligation (SNL)], with ED 50 values ranging from 8.2 to 13 mg/kg after i.p. administration in rats. Despite a 50-fold lower binding affinity to MOR, the analgesic potency of tapentadol was only two to three times lower than that of morphine, suggesting that the dual mode of action of tapentadol may result in an opiate-sparing effect. A role of NE in the analgesic efficacy of tapentadol was directly demonstrated in the SNL model, where the analgesic effect of tapentadol was strongly reduced by the α 2 -adrenoceptor antagonist yohimbine but only moderately attenuated by the MOR antagonist naloxone, whereas the opposite was seen for morphine. Tolerance development to the analgesic effect of tapentadol in the CCI model was twice as slow as that of morphine. It is suggested that the broad analgesic profile of tapentadol and its relative resistance to tolerance development may be due to a dual mode of action consisting of both MOR activation and NE reuptake inhibition.

The function of the transient receptor potential vanilloid 1 (TRPV1) cation channel was analyzed with RNA interference technologies and compared to TRPV1 knockout mice. Expression of shRNAs targeting TRPV1 in transgenic (tg) mice was proven by RNase protection assays, and TRPV1 downregulation was confirmed by reduced expression of TRPV1 mRNA and lack of receptor agonist binding in spinal cord membranes. Unexpectedly, TRPV3 mRNA expression was upregulated in shRNAtg but downregulated in knockout mice. Capsaicin-induced [Ca(2+)](i) changes in small diameter DRG neurons were significantly diminished in TRPV1 shRNAtg mice, and administration of capsaicin hardly induced hypothermia or nocifensive behaviour in vivo. Likewise, sensitivity towards noxious heat was reduced. Interestingly, spinal nerve injured TRPV1 knockout but not shRNAtg animals developed mechanical allodynia and hypersensitivity. The present study provides further evidence for the relevance of TRPV1 in neuropathic pain and characterizes RNA interference as valuable technique for drug target validation in pain research.

In order to find novel delta opioid receptor agonists, the pharmacophoric benzhydryl moiety of the lead compound SNC 80 (1) was dissected and the phenyl residues were attached to different positions of the 6,8-diazabicyclo[3.2.2]nonane core system (4). The position of the carboxamido group, the stereochemistry, the C3/C4 bond order and the kind and length of the spacer X were considered. The resulting compounds were compared with the four energetically most favourable conformations of SNC 80 by a multifit analysis. These calculations led to the structures 5-10, which fit best to SNC 80. Herein the synthesis of one of these compounds (9) is described. Starting from (S)-glutamate two alternative routes are detailed to obtain the key intermediate 14. A variation of the Dieckmann cyclization, which uses trapping of the first cyclization product with ClSiMe(3) provided the mixed acetal 20, which was carefully hydrolyzed to yield the bicyclic ketone 17. Stereoselective addition of phenylmagnesium bromide, dehydration, LiAlH(4) reduction and exchange of the N-6 residue afforded the designed compound 9. The affinities of 9 towards delta, mu, kappa and ORL1 receptors were determined in receptor binding studies with radioligands. Only moderate receptor affinity was found.

3,7-Diazabicyclo[3.3.1]nonan-9-ones having aryl rings in positions 2 and 4 with systematically varied substituents were synthesized using a double Mannich procedure. Radioligand binding assays were performed to measure the affinity of the compounds to the mu-, delta-, and kappa-opioid receptors. The affinity of all 2, 4-diphenyl-substituted 3,7-diazabicyclo[3.3.1]nonan-9-ones to the mu- and delta-receptors was found to be low. In contrast, with exception of the nitro- and cyanophenyl-substituted compounds, most of the diazabicycles showed considerable affinity for the kappa-receptor. In particular, the m-fluoro-, p-methoxy-, and m-hydroxy-substituted compounds have an affinity in the submicromolar range. Due to solubility problems in aqueous media, salts of HZ2 were synthesized. The methiodide shows high kappa-affinity and may, thus, be a promising candidate for development of a peripheral kappa-agonist, e.g. for use in the case of rheumatoid arthritis.

Galanin, a 29 amino acid peptide, has been reported to possess antinociceptive properties at the spinal site and to potentiate opioid-induced antinociception. Our aim was to investigate whether also endogenous galanin interacts with an exogenously administered opioid, morphine, in the rat spinal cord. This question was investigated by use of the recently developed galanin receptor antagonists galantide [M-15, galanin-(1-13)-substance P-(5-11) amide] and M-35 [galanin-(1-13)-bradykinin-(2-9) amide]. Nociception was assessed in the rat tail-flick test using radiant heat and the rat Randall-Selitto model of inflammatory pain using vocalization as the nociceptive criterion. Intrathecal (i.t.) injections were performed in rats under ether anaesthesia. Morphine was administered either i.t. or intraperitoneally (i.p.), and the antagonists were injected i.t. [I-125]Galanin binding experiments were performed on crude synaptosomal membranes of the rat spinal cord. In the rat tail-flick test, i.t. injection of 3 mu g morphine evoked antinociception of about 75% of the maximal possible effect (% MPE). Co-injection of either 2 mu g galantide or 2 mu g M-35 with morphine almost completely abolished the antinociceptive effect of morphine. I.p. injection of 2.15 mg/kg morphine elicited about 80% MPE when given 10 min prior to i.t. saline injection. Injection of the antagonists instead of saline antagonised the antinociceptive effect of morphine partially thus showing the spinal proportion of the overall antinociceptive effect. In the rat Randall-Selitto test, 3 mu g morphine, injected i.t., produced antinociception of almost 100% MPE. Co-injection of the antagonists reduced the maximum effect partially by about 25-35%. I.p. injection of 7.5 mg/kg morphine 10 min prior to i.t. injection of saline elicited an antinociceptive effect of 90-100% MPE; injection of the antagonists instead of saline reduced the peak effect to a similar degree as after i.t. injection of 3 mu g morphine. To exclude a direct interference by morphine with the galanin receptor, in vitro binding of [I-125]galanin to a spinal synaptosomal fraction was assessed. Morphine, 10 mu M, did not interfere with the specific [I-125]galanin binding. These results provide further evidence that galanin is involved in spinal nociceptive processing. It seems to be involved in the mediation of the effects of morphine at this site, either as a co-transmitter, or subsequent to mu-receptor activation on nerve terminals or on interneurones.

Previously 2,4-dipyridine substituted 3,7-diazabicyclo[3.3.1]nonanone diesters were found to have a high affinity and selectivity towards the kappa-opioid receptor. The purpose of this study was to check the influence of substituents at position N3 on the affinity to the mu-, delta-, and kappa-receptors. Whereas a phenylethyl group is able to create affinity to the mu-receptor, small substituents such as a hydrogen or a methyl group are responsible for a high affinity to the kappa-receptor. In addition, a dimeric compound was found to have affinity to the kappa-receptor. Although all compounds will bear at least one positive charge under physiological conditions they show a considerable lipophilicity, indicating the possibility of passing the blood-brain barrier.

The stereochemistry the 2,4-di-arene substituted 3,7-diazabicyclo[3.3.1]nonan-9-one 1,5-dicarboxylate skeleton was found to be regulated by the kind of substituents attached to the arene rings as well as to the nitrogens N3 and N7. Conformational isomers, i.e., chair/chair, boat/chair and chair/boat, in addition to cis/trans conflgurational isomerism with respect to the arene rings were reported. Since the analgesic potency of the diazabicyclononanones, which is related to their affinity toward the κ-opioid receptor, is governed by the stereochemistry of the molecules, the influence of the substituents at nitrogen N7 was studied herein. The various differently N7 substituted diazabicyclononanones were found to crystallise in a highly symmetrical chair/chair conformation. However, beside HZ2 none of the compounds exhibits high affinity to the κ receptor. In contrast, some compounds with affinity to the μ receptor could be identified. In addition, the N7-(4-carboxybenzyl) substituted compound was found to have affinity to the δ receptor in the submi-cromolar range of concentration.