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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptHHS Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
 
J Med Chem. Author manuscript; available in PMC 2010 November 12.
Published in final edited form as:
PMCID: PMC2788771
NIHMSID: NIHMS154156

14β-Arylpropiolylamino-17-cyclopropylmethyl-7,8-dihydronormorphinones and related opioids. Further examples of pseudo-irreversible mu opioid receptor antagonists

Abstract

14β-4’-Chlorocinnamoylaminodihydronormorphinone (2a), and analogues, are selective pseudoirreversible antagonists of the mu opioid receptor (MOR). The preparation of analogues with ethynic bonds, replacing the ethenic bond of 2a, is described. The new ligands, in mouse antinociceptive assays, had pseudoirreversible MOR antagonist activity which, in the case of 8b was of longer duration than that of 2a. The related codeinone (9b) had only antagonist activity in vivo, in contrast to 2a’s codeinone equivalent 3a, which had potent antinociceptive activity.

Introduction

The 14β-hydroxy-7,8-dihydronormorphinone derivatives naloxone (1a) and naltrexone (1b) were the prototypic mu opioid receptor (MOR) antagonists and both have been introduced into clinical practice. We have been interested in derivatives of the structurally equivalent 14β-amino-17-cyclopropylmethyl-7,8-dihydronormorphinone (1c) for some time, with particular attention to the 14β-cinnamoylamino derivatives (2, 3)1 and related side-chain analogues (6–7).2 The most studied derivatives have been clocinnamox (C-CAM, 2a)11 and methcinnamox (M-CAM, 2b)10 and their codeinone precursors (3a, 3b).3 They are impressive MOR-selective pseudo-irreversible antagonists with only the codeinones (3a, 3b) having any in vivo antinociceptive activity.1 The present study was undertaken to determine the OR profile of the analogues of 14β-cinnamoylamino and 14β-cinnamylamino derivatives (2 – 5) in which the trans ethenic bond in the cinnamoyl or cinnamyl group is replaced by an ethynic bond in the arylpropiolylamino derivatives (8, 9) and arylpropargylamino derivatives (10, 11). The ethynic bond in the new ligands places the key aromatic group further from C14 than in the cinnamoylamino and cinnamylamino ligands previously studied. The data collected in the present study show that the arylpropiolylamino morphinones (8) are pseudoirreversible MOR antagonists at least the equal of their cinnamoylamino analogues.

Synthesis

While phenylpropiolic acid is commercially available, p-chlorophenylpropiolic acid (15) and p-chlorophenylpropargyl bromide (17) were obtained by preparation from the appropriate cinnamic acid (Scheme 1).4,5 Target compounds (9) were then accessed by acylation of N-cyclopropylmethyl-14β-aminodihydronorcodeinone (18b) (Scheme 2).6,7 The equivalent morphinones (8) were accessed from codeinones (9) by 3-O-demethylation with boron tribromide. Direct alkylation of N-cyclopropylmethyl-14β-aminodihydrocodeinone and N-cyclopropylmethyl-14β-aminodihydromorphinone using the arylpropargyl bromide (17) gave target compounds 10, 11 (Scheme 2).

Scheme 1
(i) EtOH, c.H2SO4, reflux, 4 h, 80%; (ii) Br2, DCM, r.t., overnight, 70%; (iii) KOH, EtOH, reflux, 6 h, 40%; (iv) DIBAL, Et2O, −78 °C to r.t., overnight, 64%; (v) PPh3, imidazole, Br2, DCM, r.t., 1.5 h, 79%.
Scheme 2
(i) R’C6H4CCCOCl, NEt3, DCM, r.t., overnight, 27 – 71% (ii) BBr3, DCM, −30 °C to r.t., 0.5 h, 72 – 75% (iii) ClC6H4CCCH2Br, K2CO3, DMF, 90 °C, 3 h, 63 – 74%.

Results

Affinity for the individual types of opioid receptors (OR) was determined by displacement binding assays using membranes prepared from Chinese hamster ovary (CHO) cells expressing recombinant human opioid receptors. The selective radioligands used were [3H]-DAMGO (MOR), [3H]U69593 (KOR) and [3H]Cl-DPDPE (DOR).8 All the new morphinones (8a, 8b, 10) had high affinity for all OR with no selectivity for any one (Table 1). The equivalent codeinones (9a, 9b, 11) had generally lower OR affinity, particularly at DOR and KOR. Overall, the affinities of the new series (8 – 11) were similar to the affinities displayed by the equivalent cinnamoylamino derivatives (2, 3) and cinnamylamino derivatives (4, 5).

Table 1
Binding affinities (Ki) of new compounds to opioid receptors and antagonist activity (Ke) in the [35S]GTPγS binding assay.

In vitro OR functional activity of the new ligands was determined in assays in which stimulation of [35S]GTPγS binding is measured for individual recombinant human OR transfected into CHO cells.8,9 The morphinones (8a, 8b, 10) were potent antagonists for all three OR, except that 8a showed low efficacy agonism at KOR (Table 1). The equivalent codeinones (9a, 9b, 11) had MOR antagonist activity of lower potency than their morphinone equivalents (8a, 8b, 10). The only codeinone with both KOR and DOR antagonist activity was the arylpropargyl derivative (11), but potency was low in each case so that 11 profiled as a MOR antagonist of some selectivity. The arylpropiolylamino codeinones (9a, 9b) were KOR partial agonists; 9a also had DOR partial agonist activity whereas 9b was a DOR antagonist (Table 1).

The arylpropiolylamino derivatives (8a, 8b, 9a, 9b) were evaluated in vivo in mouse antinociceptive assays using thermal (50 °C water tail withdrawal; TW) and chemical (acetic acid induced stretching; AS) nociceptors.10 Only the phenylpropiolylamino codeinone (9a) had any antinociceptive activity in these assays. In TW it was a partial agonist with peak effect (at 3.2 mg/kg) reaching about two thirds of the maximum possible (Fig 1). In AS 9a at a dose of 0.32mg/kg totally inhibited the stretching response. This effect was reversed by the selective MOR antagonist methcinnamox (M-CAM, 2b) but not by naltrindole (DOR) and partially by norBNI (KOR) (Table 2).

Figure 1
Antinociceptive activity of 9a in the mouse warm water tail withdrawal assay.
Table 2
Agonist selectivity of 9a (0.32 mg/kg, s.c.) in the mouse AS assay

All of the arylpropiolylamino ligands were able to antagonise the antinociceptive effects of morphine in TW (Table 3). When a high dose of the test ligand (32 mg/kg) was administered 30 min before morphine, 8a, 8b and 9b all flattened the morphine dose-response curve and were shown to retain antagonist activity beyond 48h, 144h and 24h respectively. 9a, which had agonist activity in TW, was only tested at 24h and beyond; it was active as an MOR antagonist at 24h (Table 3).

Table 3
Antagonist activity in the mouse tail-withdrawal (50 °C) test.

The most impressive in vivo morphine antagonist among the new ligands was thus the p-chlorophenylpropiolylamino morphinone (8b). It’s OR antagonist selectivity was investigated in AS against ED100 doses of the selective agonists morphine (MOR), bremazocine (KOR) and BW373U86 (DOR). At a dose of 3.2 mg/kg of 8b, the inhibitory effect of each of the selective agonists was fully reversed; 1mg/kg 8b was also substantially effective with the MOR and DOR activity showing superiority over KOR activity (Fig 2). With 24h pre-treatment 3.2 mg/kg 8b still showed substantial OR antagonist activity; this activity was significantly MOR selective (Fig. 3).

Figure 2
Antagonist activity of 8b in the mouse abdominal stretch (AS) assay versus fully effective doses of standard opioid receptor agonists.
Figure 3
Antagonist selectivity of 8b (3.2 mg/kg) in the mouse abdominal stretch (AS) assay after 24 h pretreatment. ** p < 0.01 compared to morphine. ≠≠ p < 0.01 compared to control (8b alone), which was not different from vehicle ...

Discussion

The data obtained from the evaluation of the new ligands synthesised in the present study conform to the pattern of structure-activity relationships established for the analogous 14-cinnamoylamino derivatives.1 That is, the arylpropiolylamino morphinones (8a, 8b) behave as pseudoirreversible MOR antagonists of long duration and that p-chloro substitution in the side-chain aromatic ring eliminates agonist activity, enhances MOR antagonist activity and increases the duration of the latter effect. Comparison with the equivalent cinnamoylaminomorphinones (2a, 2c) reveals substantial similarity between 2a and 8b in binding and in vitro assays (Table 1). Comparison of the binding of the 14-propiolylamino codeinones (9) with the corresponding morphinones (8) shows little difference in MOR binding affinity but greater difference in KOR and particularly DOR binding. Similar effects were seen in comparison of the 14-cinnamoylamino morphinones (2) with codeinones (3). This confirms that for MOR binding the lipophilic C14 side chain is the dominant structural motif whereas for KOR and DOR binding the C3 phenolic group is also required. In vivo it appears that the arylpropiolylamino derivatives (8a, 8b) have somewhat longer duration of morphine antagonist activity than the cinnamoylamino equivalents (2a, 2c).1 When the morphine dose-response curve was determined 4 days after administration of a 32 mg/kg dose of the test antagonist, 2a shifted the dose-response curve about 5-fold to the right11 whereas in the present study the same dose of 8b in the same protocol resulted in a 12-fold shift (Table 3).

There is perhaps greater difference in in vivo activity between the arylpropiolylamino codeinones (9a, 9b) and the corresponding cinnamoylamino codeinones (3a, 3c). Whereas 9b had no antinociceptive activity in TW or AS, 3a, though having no agonist activity in TW, has potent antinociceptive activity in AS, being fully effective at 0.2mg/kg.1,12,13 Similarly the MOR partial agonism shown in vivo by 3c is of higher efficacy than that of 9a in the present study.1 Thus it appears that the conformationally linear side chains of the arylpropiolylamino derivatives (8, 9) are associated with lower MOR efficacy and more profound MOR antagonism than the equivalent cinnamoylamino derivatives (2, 3). This suggests that for antagonism, the optimum position for the lipophilic aryl moiety is further from C14 than is achieved in the cinnamoyl derivatives. Rennison et al2 showed that the 14β-phenylbutylamidomorphinone (12), with a longer spacer between C14 and the aryl group, was more effective than C-CAM in flattening the dose-response curve of DAMGO in a [35S]GTPγS binding assay.

In the present study there was pronounced similarity in OR binding and in vitro profiles between the p-chlorophenylpropiolylamino derivatives (8b, 9b) and the p-chlorophenylpropargylamino derivatives (10, 11). In this respect the SAR followed that established by Rennison et al2 for a range of 14-acylamino- (6) and 14-alkylaminomorphinones and codeinones (7).

Conclusions

The 14β-arylpropiolylamino morphinones and codeinones of the present study provide further examples of pseudoirreversible MOR antagonists comparable to the previously reported clocinnamox and methcinnamox. The SAR of the new series is very similar to that of the equivalent cinnamoylamino series but there are trends to lower MOR efficacy and more profound antagonism.

Experimental

Reagents and solvents were purchased from Aldrich or Lancaster and used as received. NMR Spectra: Jeol Lambda-270-MHz instrument: 1H at 270 MHz, with TMS as an internal standard. Only representative examples of the synthesis are shown. Oxalate salts were formed prior to pharmacological evaluation. Tested compounds had purity ≥95%.

N-Cyclopropylmethyl-14β-[phenylpropioloylamino]-7,8-dihydronorcodeinone (9a)

Oxalyl chloride (8.8 eqv), phenylpropiolic acid (1.1 eqv) in anhydrous toluene were heated at reflux for 1 h. The solvent was removed, the residue dissolved in anhydrous CH2Cl2, added dropwise to a solution of 18b (1 eqv) and triethylamine (1.1 eqv) in anhydrous CH2Cl2, and stirred at r.t overnight. The solvent was removed and the crude residue purified by column chromatography to yield a white solid (71%); Rf (CH2Cl2:MeOH, 50:1) 0.26; 1H NMR (CDCl3) 0.21 (2H, m), 0.60 (2H, m), 0.89 (1H, m), 2.32–2.52 (2H, m), 3.10 (1H, d), 3.88 (3H, s), 4.95 (1H, s), 6.63 (1H, d), 6.74 (1H, d), 7.30 (1H), 7.35–7.61 (5H, m).

N-Cyclopropylmethyl-14β-[phenylpropioloylamino]-7,8-dihydronormorphinone (8a)

To the codeinone (9a) in anhydrous CH2Cl2 at −30°C under N2, was added BBr3 (6 eqv, 1M in CH2Cl2) slowly. The reaction was allowed to reach r.t. over 1 h before adding a 1:1 mixture of ice:ammonia (conc’). The organic phase was isolated, the aqueous layer washed (x 3) with CHCl3:MeOH (3:1), the combined organic fractions washed with brine, dried and evaporated to dryness. Column chromatography gave 8a as a white solid (72%). Rf (CH2Cl2:MeOH, 20:1) 0.47; 1H NMR (DMSO) 0.25 (2H, m), 0.62 (2H, m), 0.92 (1H, m), 3.10 (1H, d), 5.01 (1H, s), 6.61 (1H, d), 6.80 (1H, d), 7.38 (5H, m).

N-Cyclopropylmethyl-14β-[3’-(4”-chlorophenyl)-propargylamino]-7,8-dihydronormorphinone (10)

18a was treated with 3-(4’-chlorophenyl)propargyl bromide (17, 1.1 eq.) in the presence of potassium carbonate (5 eq.) in dimethylformamide at 90 °C for 12 h. The solvent removed and the crude residue purified by column chromatography to afford 10 as a white solid (63%). Rf 0.42 (CH2Cl2:MeOH, 20:1); 1H NMR (CDCl3) δ 0.18 (2H, m), 0.54 (2H, m), 0.89 (1H, m), 3.06 (1H, d), 3.57 (2H, s), 4.70 (1H, s), 6.60 (1H, d), 6.71 (1H, d), 7.29 (2H, d), 7.37 (2H, d).

Chart 1
14-Substituted 7,8-dihydromorphinones and codeinones.

Supplementary Material

1_si_001

Acknowledgement

NIDA grants DA00254 & DA07315. In vitro evaluation through the NIDA Abuse Treatment Discovery Program (ATDP).

Abbreviations

OR
opioid receptors
C-CAM
14β-4’-Chlorocinnamoylaminodihydronormorphinone
M-CAM
14β-4’-Methylcinnamoylaminodihydronormorphinone
norBNI
nor-binaltorphimine
TW
tail withdrawal
AS
abdominal stretch

Footnotes

Supporting Information Available: Full experimental details, including biological assay methods. This material is available free of charge via the Internet at http://pubs.acs.org

References

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