High-affinity carbamate analogues of morphinan at opioid receptors

Article abstract of DOI:10.1016/j.bmcl.2007.01.013

A series of carbamate analogues were synthesized from levorphanol (1a), cyclorphan (2a) or butorphan (3a) and evaluated in vitro for their binding affinity at μ, δ, and κ opioid receptors. Functional activities of these compounds were measured in the [³⁵S]GTPγS binding assay. Phenyl carbamate derivatives 2d and 3d showed the highest binding affinity for κ receptor (K_i = 0.046 and 0.051 nM) and for μ receptor (K_i = 0.11 and 0.12 nM). Compound 1c showed the highest μ selectivity. The preliminary assay for agonist and antagonist properties of these ligands in stimulating [³⁵S]GTPγS binding mediated by the κ opioid receptor illustrated that all of these ligands were κ agonists. At the μ receptor, compounds 1b, 1c, 2b, and 3b were agonists, while compounds 2c-e and 3c-e were μ agonists/antagonists.

Full text of DOI:10.1016/j.bmcl.2007.01.013

Bioorganic & Medicinal Chemistry Letters 17 (2007) 1508–1511
High-affinity carbamate analogues of morphinan at opioid receptors
Xuemei Peng,a Brian I. Knapp,b Jean M. Bidlackb and John L. Neumeyera,*
aAlcohol and Drug Abuse Research Center, McLean Hospital, Harvard Medical School, 115 Mill Street, Belmont, MA 02478, USA
bDepartment of Pharmacology and Physiology, School of Medicine and Dentistry, University of Rochester, Rochester, NY 14642, USA
Received 21 November 2006; revised 1 January 2007; accepted 3 January 2007
Available online 17 January 2007
Abstract—A series of carbamate analogues were synthesized from levorphanol (1a), cyclorphan (2a) or butorphan (3a) and evalu-
ated in vitro for their binding affinity at l, d, and j opioid receptors. Functional activities of these compounds were measured in the
[35S]GTPcS binding assay. Phenyl carbamate derivatives 2d and 3d showed the highest binding affinity for j receptor (Ki = 0.046 and
0.051 nM) and for l receptor (Ki = 0.11 and 0.12 nM). Compound 1c showed the highest l selectivity. The preliminary assay for
agonist and antagonist properties of these ligands in stimulating [35S]GTPcS binding mediated by the j opioid receptor illustrated
that all of these ligands were j agonists. At the l receptor, compounds 1b, 1c, 2b, and 3b were agonists, while compounds 2c–e and
3c–e were l agonists/antagonists.
Ó 2007 Elsevier Ltd. All rights reserved.
Treatment of severe pain often requires use of opioid
analgesics such as morphine, their use is limited by seri-
ous side-effects including tolerance, physical depen-
dence, respiratory depression, and addiction liability.
Numerous opiates have been synthesized in an effort
to obtain analgesics that are free of such side effects.1
which is also a potential site for metabolism, conjuga-
tion, and excretion via O-glucuronidation resulting in
low oral bioavailability and short duration of action.1
An approach to improving the pharmacological proper-
ties of analgesics such as morphine is to modify the phe-
nolic hydroxy function.
Opioids bind to specific neuronally located proteins
which initiate biologic responses. Opioid receptors were
first discovered in mammalian brain and belong to the
rhodopsin subfamily in the superfamily of over 1000 G
protein-coupled receptors.2 Three opioid receptor types,
kappa (j), delta (d), and mu (l),3–5 differ in their distri-
bution and ligand binding, and were identified through
pharmacological and physiological studies using selec-
tive ligands.
To develop additional insight into the SAR surrounding
the 3-position of morphinan, we wished to modify the
3-OH function of morphinan. The carbamate group
was introduced because it is a structure of medium
polarity, capable of forming hydrogen bonds as donor
and acceptor.7–9 Ideally, these modified compounds
would have a longer duration of action and better bio-
availability. We report a series of morphinans where
the 3-OH was modified by incorporating the carbamate
function at the 3-position of the morphinan template.
By the synthesis of a series of carbamate analogues of
morphinans and evaluation of their binding affinities
at opioid receptors, our objective is to identify long-act-
ing opioid receptor ligands useful for the treatment of
cocaine abuse. Earlier studies have shown that the basic
nitrogen and a phenol moiety were necessary for narcot-
ic analgesics to bind to its opioid receptors.6 As is the
case for morphine derivatives, the phenolic hydroxyl
group in morphinan contributes to analgesic activity,
The levorotatory morphinans (2a and 3a) were prepared
from commercially available (À)-3-hydroxy-N-meth-
ylmorphinan tartrate (levorphanol) 1a which was con-
verted to the free base and N-demethylated10–12 to
yield the normorphinan. Alkylation with cyclopropylm-
ethyl or cyclobutylmethyl bromide led to the (À)-N-
substituted morphinan characterized as their crystalline
mandelate salts (2a and 3a), respectively.
A series of carbamates were prepared using morphinans
(1a, 2a and 3a) as starting materials. The carbamates
were prepared in yields ranging from 55% to 90% by
treating the morphinans (1a, 2a and 3a) with the
Keywords: Morphinan; Carbamate; Opioid; Receptors.
*
Corresponding author. Tel.: +1 6178553388; fax: +1 6718552519;
e-mail: Neumeyer@mclean.harvard.edu
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.01.013

X. Peng et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1508–1511
1509
N
N
N
RO
RO
RO
(–)1
(–)2
(–)3
2a, R=H, cyclorphan
2b, R=CONHCH3
2c, R=CONHCH2CH3
2d, R=CONHC6H5
2e, R=CONHCH2C6H5
3a, R=H, butorphan
3b, R=CONHCH3
3c, R=CONHCH2CH3
3d, R=CONHC6H5
3e, R=CONHCH2C6H5
1a, R=H, levorphanol
1b, R=CONHCH3
1c, R=CONHCH2CH3
Figure 1. Structures of morphinans levorphanol, cyclorphan, butorphan and carbamate analogues.
corresponding commercially available aryl or alkyl iso-
cyanates (Fig. 1).
slightly lower affinity (4-fold) at j, l, and d receptors.
All N-cyclopropylmethyl derivatives retained j/l
selectivity, while all N-cyclobutylmethyl derivatives
showed higher l selectivity over j receptors compared
to butorphan (3a).
Synthesized ligands were evaluated for their affinity at
and selectivity for l, d, and j opioid receptors with Chi-
nese hamster ovary (CHO) cell membranes stably
expressing one type of the human opioid receptors.
The data are summarized in Table 1. For comparison
purposes, opioid-binding affinity data for levorphanol
(1a), cyclorphan (2a), and butorphan (3a) are also
included.11
It is interesting to note that compound 2e (benzyl car-
bamate of cyclorphan 2a) displayed almost identical
affinities at all three opioid receptors to the analogue
ethyl carbamate of cyclorphan 2c, while the benzyl car-
bamate of butorphan 3e displayed higher affinities than
the ethyl carbamate of butorphan 3c at all three
receptors.
From the data shown in Table 1, the binding affinities of
the carbamate-derived opioids were generally lower than
the binding affinities of the phenol precursors. It is note-
worthy that the phenyl carbamate analogues (2d and 3d)
retained the same high affinity (Ki = 0.046 –0.051 nM) at
the j receptor, and a 2-fold decrease at l for 2d and a
2-fold increase at l for 3d were observed when
compared with cyclorphan (2a) and butorphan (3a).
The relative efficacy of these morphinan ligands, 1b, 1c,
2b–2e and 3b–3e, compared with levorphanol (1a),
cyclorphan (2a), and butorphan (3a) was selected
for the [35S]GTPcS assay. Table 2 shows the agonist
and antagonist properties of these ligands in stimulating
[35S]GTPcS binding mediated by the j opioid receptor.
The methyl carbamate analogues (1b, 2b and 3b), com-
pared to the phenolic precursors, showed 1.4–100-fold
decrease in affinity at j, 3–22-fold decrease at d and
5–85-fold decrease at the l receptor and compound 3b
displayed a good selectivity for j versus d receptors.
Similarly, appreciable decreases in affinity were observed
in the ethyl carbamate analogues (1c, 2c and 3c) com-
pared to the parent compounds. Compared to phenol
2a and 3a, the benzyl carbamate a 2e and 3e displayed
Ligands 2c–e and 3e produced high maximal stimulation
of [35S]GTPcS binding (Emax) comparable to that of
selective agonist U50,488, while ligands 1b, 2b, 3b and
3d produced similar maximal stimulation to that of the
compounds 2a and 3a, but less than that of selective
agonist U50,488.
The EC50 values of these ligands are similar which sub-
stantially correlate with the Ki values obtained for the
Table 1. Ki values for the inhibition of l, d, and j opioid binding to Chinese hamster ovary membrane by carbamate opioids
Compound
Ki SEM (nM)
[3H]U69,593 (j)
Selectivity
[3H]DAMGO (l)
[3H]Naltrindole (d)
l/j
d/j
(À)1a (levorphanol)
(À)1b (MCL-433)
(À)1c (MCL-431)
(À)2a (cyclorphan)
(À)2b (MCL-434)
(À)2c (MCL-449)
(À)2d (MCL-429)
(À)2e (MCL-444)
(À)3a (butorphan)
(À)3b (MCL-435)
(À)3c (MCL-432)
(À)3d (MCL-428)
(À)3e (MCL-443)
0.21 0.02
1.4 0.0087
4.8 0.20
2.3 0.3
3.3 0.40
36 8.5
4.2 2.3
53 4.5
78 4.6
0.09
0.4
0.1
2
2
16
2
0.062 0.003
5.3 0.62
0.034 0.002
3.4 0.63
1.9 0.072
42 6.3
60
12
20
35
20
70
37
6
1.6
2
0.29 0.025
0.11 0.022
0.30 0.030
0.23 0.01
1.2 0.12
0.14 0.015
0.046 0.005
0.14 0.017
0.079 0.003
0.81 0.066
4.3 0.30
2.8 0.30
1.6 0.15
2.7 0.27
5.9 0.55
30 0.67
27 2.5
2.4
2
3
1.5
0.9
2
3.7 0.69
0.12 0.01
0.70 0.063
0.051 0.002
0.30 0.045
3.9 0.26
15 1.5
76
50
0.4

1510
X. Peng et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1508–1511
Table 2. Agonist and antagonist properties of compounds in stimulating [35S]GTPcS binding mediated by the j opioid receptora
Compound
Pharmacological properties
Emax (%maximal stimulation)
EC50 (nM)
Imax (% maximal inhibition)
(À)U50,488
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
110 2.0
90 10
46 16
0.19 0.04
1.3 0.4
29 5.7
—
—
(À)2a (cyclorphan)
(À)3a (butorphan)
(À)1b (MCL-433)
(À)2b (MCL-434)
(À)2c (MCL-449)
(À)2d (MCL-429)
(À)2e (MCL-444)
(À)3b (MCL-435)
(À)3c (MCL-432)
(À)3d (MCL-428)
(À)3e (MCL-443)
80 6.8
96 10
—
No effect
No effect
No effect
No effect
No effect
No effect
No effect
No effect
No effect
78 6.8
140 4.7
120 7.5
130 10
88 8.5
61 2.5
74 4.7
110 0.88
16 0.19
1.2 0.41
0.38 0.10
1.1 0.12
9.6 0.4
23 3.6
2.2 0.91
4.7 0.49
a Membranes from CHO cells that stably expressed only one type of the opioid receptor were incubated with varying concentrations of the
compounds. The stimulation of [35S]GTPcS binding was measured as described previously.13 To determine the antagonist properties of a com-
pound, membranes were incubated with 100 nM of the j agonist U50,488 in the presence of varying concentrations of the compound. The Imax
value is the maximal percent inhibition obtained with the compound. The IC50 value is the concentration of compound needed to produce half-
maximal inhibition. Dashed lines indicate that the compound was not tested for antagonist properties because of its high Emax value.
Table 3. Agonist and antagonist properties of compounds in stimulating [35S]GTPcS binding mediated by the l opioid receptora
Compound
Pharmacological properties Emax (% maximal stimulation) EC50 (nM) Imax (% maximal inhibition) IC50 (nM)
DAMGO
Agonist
120 12
40 2.9
50 2.5
71 4.1
44 2.9
40 2.5
39 3.0
39 1.9
60 5.8
3 2 0.88
80 10
110 9.0 —
0.80 0.06 50
—
1.7 0.4
(À)2a (cyclorphan) Agonist/antagonist
1
3
(À)3a (butorphan)
(À)1b (MCL-433)
(À)2b (MCL-434)
(À)2c (MCL-449)
(À)2d (MCL-429)
(À)2e (MCL-444)
(À)3b (MCL-435)
(À)3c (MCL-432)
(À)3d (MCL-428)
(À)3e (MCL-443)
Agonist/antagonist
Agonist
Agonist
1.6 0.2
50 4.0
14 2.3
50
20
3
No effect
No effect
No effect
No effect
10 1.9
6.4 2.0
16 2.9
No effect
NA
Agonist/antagonist
Agonist/antagonist
Agonist/antagonist
Agonist
2.4 0.24 68 2.7
0.44 0.19 70 3.5
2.2 0.18 71 1.8
11 1.1
39 7.5
No effect
33 3.1
Agonist/antagonist
Agonist/antagonist
Agonist/antagonist
1.3 0.16 37 1.2
7.1 0.48 50 2.5
22 1.9
260 88
54 1.2
a Membranes from CHO cells that stably expressed only the opioid receptor were incubated with varying concentrations of the compounds. The
stimulation of [35S]GTPcS binding was measured as described previously.13 EC50 values were the concentration of compound needed to produce
50% of the Emax value. When the Emax value was 30% or lower, it was not possible to calculate an EC50 value. To determine the antagonist
properties of a compound, membranes were incubated with 200 nM of the l agonist DAMGO in the presence of varying concentrations of the
compound. The Imax value is the maximal percent inhibition obtained with the compound. The IC50 value is the concentration of compound needed
to produce half-maximal inhibition. Dashed lines indicate that the compound was not tested.
compounds in the binding assays with [3H]U69,593.
Similar to cyclorphan (2a) and butorphan (3a),
these ligands did not inhibit U50,488-stimulated
[35S]GTPcS, which suggested that all of these ligands
were full j agonists.
inhibition (Imax) of the DAMGO-stimulated [35S]GTPcS
binding than the carbamates of the cyclorphan (ligands
2c–e). These data indicated that ligands 3c–e are weak l
agonists and antagonists.
The preliminary assay for agonist and antagonist prop-
erties of these ligands in stimulating [35S]GTPcS binding
mediated by the j opioid receptor illustrated that all of
these ligands were j agonists, however compounds 1b,
2b and 3b were agonists at the l receptor, whereas com-
pounds 2c–e and 3c–e were both agonists and antago-
nists at the l receptor.
The properties of these ligands in stimulating
[35S]GTPcS binding mediated by the l opioid receptor
are shown in Table 3.
Ligands 1b, 2b and 3b produced maximal stimulation of
[35S]GTPcS binding mediated by l receptor comparable
to that of the parent compound cyclorphan (2a) and
butorphan (3a) and no inhibition of DAMGO-stimu-
lated [35S]GTPcS binding. These data indicated that
1b, 2b and 3b were l agonists. Ligands 2c–e had the low-
est maximal stimulation of [35S]GTPcS binding and the
highest maximal inhibition (Imax) of the DAMGO-stim-
ulated [35S]GTPcS binding indicating that ligands 2c–e
are l antagonists with weak agonistic activities at the
l receptors. Compounds 3c–e produced lower maximal
A series of carbamate analogues were synthesized from
levorphanol (1a), cyclorphan (2a) or butorphan (3a)
and evaluated in vitro for their binding affinity at l, d,
and j opioid receptors. The binding affinities of the car-
bamate-derived opioids were generally lower than the
binding affinities of the phenol precursors. Phenyl carba-
mate derivatives 2d and 3d showed the highest binding
affinity for j receptor and increased affinity at l for 3d.

X. Peng et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1508–1511
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Functional activities of these compounds were measured
in the [35S]GTPcS binding assay, indicating that all of
these ligands were j agonists while, compounds 1b, 1c,
2b and 3b were l agonists, and compounds 2c–e and
3c–e were l agonists/antagonists. Such carbamate deriv-
atives may be useful for the development of longer-act-
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This work was supported by the National Institute
on Drug Abuse: RO1-DA14251 (J.L.N.) and
KO5DA00360 (J.M.B.). Levorphanol tartrate was gen-
erously donated by Mallinckrodt Inc.
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