Synthesis and biological evaluation of α- and β-6-amido derivatives of 17-cyclopropylmethyl-3,14β-dihydroxy-4, 5α- epoxymorphinan: Potential alcohol-cessation agents

Article abstract of DOI:10.1021/jm701060e

Substituted aryl and aliphatic amide analogues of 6-naltrexamine were synthesized and used to characterize the binding to and functional activity of human μ-, δ-, and κ-opioid receptors. Competition binding assays showed 11-25 and 27-31 bound to the μ (K<

Full text of DOI:10.1021/jm701060e

J. Med. Chem. 2008, 51, 1913–1924
1913
Synthesis and Biological Evaluation of r- and ꢀ-6-Amido Derivatives of
17-Cyclopropylmethyl-3, 14ꢀ-dihydroxy-4, 5r-epoxymorphinan: Potential Alcohol-Cessation
Agents
Senait Ghirmai,† Marc R. Azar,‡ Wilma E. Polgar,§ Ilona Berzetei-Gurske,§ and John R. Cashman*,†
Human BioMolecular Research Institute, 5310 Eastgate Mall, San Diego, California 92121-2804, BehaVioral Pharma, Inc., 505 Coast
BouleVard South, Suite 210, La Jolla, California 92037, and SRI International, 333 RaVenswood AVenue, Menlo Park, California 94025
ReceiVed September 18, 2007
Substituted aryl and aliphatic amide analogues of 6-naltrexamine were synthesized and used to characterize
the binding to and functional activity of human µ-, δ-, and κ-opioid receptors. Competition binding assays
showed 11–25 and 27–31 bound to the µ (Ki ) 0.05–1.2 nM) and κ (Ki ) 0.06–2.4 nM) opioid receptors.
Compounds 11–18 possessed significant binding affinity for the δ receptor (Ki ) 0.8–12.4 nM). Functional
assays showed several compounds acted as partial or full agonists of δ or κ receptors while retaining an
antagonist profile at the µ receptor. Structure–activity relationship for aryl amides showed that potent
compounds possessed lipophilic groups or substituents capable of hydrogen bonding. Metabolic stability
studies showed that 11, 12, and 14 possessed considerable stability in the presence of rat, mouse, or human
liver preparations. The ED50 of inhibition of 10% ethanol self-administration in trained rats, using operant
techniques for 11, was 0.5 mg/kg.
Introduction
Alcoholism is a serious disorder with significant social and
economic consequences. Numerous studies suggest that alcohol
interacts with endogenous opioid systems.1,2 In 1994, naltrexone
(Figure 1) was approved by the United States Food and Drug
Administration (FDA) for treatment of alcoholism. Naltrexone
is a pure µ-opioid receptor antagonist with no agonist activity
and no abuse potential. Blocking the opioid receptor inhibits
Figure 1. Structure of naltrexone 4 and nalmefene 5.
the effects of pleasure-inducing endogenous opioids released
by alcohol, resulting in an attenuation of the positive reinforcing
effects of alcohol consumption. In a laboratory study of
nonproblem drinkers, naltrexone was found to decrease the
reinforcing (i.e., stimulant) effects and increase the unpleasant
(i.e., sedative) properties of initial alcohol consumption.3 Studies
using rodent and monkey animal models have shown that the
opioid antagonists naloxone and naltrexone decrease the vol-
untary consumption and stress-induced increase in alcohol self-
administration, suggesting that these agents may prevent the
reinforcing effects of alcohol consumption.4 Naltrexone may
be most beneficial among alcoholics with higher levels of
craving and poorer cognitive functioning.5 However, in one
study, 15% of patients undergoing naltrexone treatment termi-
nated treatment early because of adverse effects including
intolerable nausea.6 Naltrexone is also associated with dose-
dependent hepatotoxic side effects that complicate use and
confound treatment of alcoholic patients with liver disease.7
Additional shortcomings include a less than desirable duration
of action, relatively low bioavailability,8 and possibly, a
relatively low affinity for δ and κ receptors thought to be
involved in diminishing the reinforcing effects of drinking
alcohol.9 Beneficial effects of naltrexone diminish gradually over
time, and an extended-release injectable suspension of naltrexone
has shown promise.10 In some cases, the effects of naltrexone
have been quite robust, but in other cases, evidence of its
efficacy has been less consistent. The naltrexone analogue,
nalmefene (Figure 1), an opioid antagonist with superior
pharmaceutical properties to naltrexone may hold greater
promise as an alcohol treatment medication.
Opioid receptor antagonists have direct effects on alcohol-
seeking behavior.4 For example, direct perfusion of naloxone
via microdialysis into the nucleus accumbens neurons inhibited
alcohol-mediated dopamine release.11,12 A decrease in alcohol
consumption by blockade of opioid receptors suggests direct
effects of naloxone in this reinforcement system.3,4 Animal
studies have provided evidence that µ-, δ-, and κ-opioid
receptors contribute to alcohol-induced reinforcement.13,14
Previously, reports of N-acyl-ꢀ-naltrexamine derivatives have
been reported in the literature. For example, a naphthalene
dialdehyde amide of naltrexamine was reported as a µ-receptor
affinity label.15 Others have reported on morphinan amide
derivatives as analgesic and diuretic agents.16 TRX-82017 and
funaltrexamine18 represent two other examples of antagonists
and agonists, respectively, of opioid receptors. The work herein
presents a description of a class of compounds that has potency
and efficacy against µ-, δ-, and κ-opioid receptors as alcohol
self-administration cessation agents and may help determine the
a Abbreviations: DETAPAC, diethylenetriaminepentaacetic acid; BOP,
benzotriazol-1-yl-oxy-tris-(dimethylamino)phosphonium hexa-fluorophos-
phate; ESI, electrospray ionization; GTP, guanosine 5′-triphosphate; FBS,
fetal bovine serum; CHO, Chinese hamster ovary; DMEM, Dulbecco’s
minimal essential medium; BAL, blood alcohol level.
* To whom correspondence should be addressed. Telephone: 858-458-
9305. Fax: 858-458-9311. E-mail: jcashman@hbri.org.
†
Human BioMolecular Research Institute.
Behavioral Pharma, Inc.
‡
§
SRI International.
10.1021/jm701060e CCC: $40.75
2008 American Chemical Society
Published on Web 02/26/2008

1914 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Ghirmai et al.
key opioid receptor(s) required for alcohol self-administration
cessation. The objective of our work was to develop metaboli-
cally stable analogues of 4 and 5. Because of the possibility
that the C-6 epoxide of nalmefene or other reactive metabolites
of naltrexone produced by metabolism contributes to hepata-
toxicity, by replacing the metabolically labile 6-keto or 6-me-
thylene groups of 4 and 5, respectively, with an amide moiety,
this may lead to more long-lived agents with potentially less
hepatatoxicity but still retain great pharmacological efficacy.
saccharides 9 and 10 were relatively selective for the µ receptor.
A 3- and 2-fold stereoselectivity for binding to the δ- and
κ-opioid receptors was observed between the R and ꢀ saccharide
isomers (i.e., 9 and 10). The benzyl derivative of saccharide 9
(i.e., compound 8) was about 4.5- and 1.6-fold less potent than
naltrexone and nalmefene, respectively, for the µ-opioid receptor
but was more potent against the κ-opioid receptor. This
suggested that lipophilic amides could retain potency for µ and
δ receptors and possibly have increased potency for the κ
receptor. Because our goal was to design agents with increased
activity for κ receptors, we synthesized lipophilic aryl amide
derivatives of 7.
Results and Discussion
The chemical synthesis of a series of substituted aryl and
aliphatic amide derivatives of 6-naltrexamine (Table 1) was
efficiently accomplished and used to characterize the structural
requirements for binding to and functional activity of human
µ-, δ-, and κ-opioid receptors.
In comparison to nalmefene, with the exception of compounds
8, 25, and 26, each of the test compounds examined possessing
a C-6 aromatic amide group showed higher affinity (Ki )
0.05–0.9 nM) for the µ-opioid receptor. Compounds 11–23
possessed 1.2-7- and 1.5-8-fold higher affinity for the µ
receptor compared to naltrexone and nalmefene, respectively.
Compound 26 contained a benzoic acid moiety and was the
least potent agent tested against the µ receptor. Compounds
27-31 showed 2–15-fold greater potency for the µ receptor
compared to nalmefene. The rank order of affinity for the most
potent aryl amides at the µ receptor was 11 > 27 > 23 ) 28 >
12–17 > 18 ) 19 > 20 > 21 > 33 > 22. In general, compounds
with substitutions on the aromatic portion of the amide (i.e.,
11–25) showed significantly more µ versus δ receptor selectivity
than µ versus κ receptor selectivity as indicated by the ratio of
δ/µ and κ/µ values (Table 2). With the exception of 27, for
compounds examined with aliphatic amide groups (i.e., 27–31),
the affinity for the µ receptor was about 1–2-fold greater
compared to naltrexone and 3–8-fold greater compared to
nalmefene. Compound 27 showed 5.5–15-fold greater potency
for the µ receptor compared to naltrexone and nalmefene,
respectively. Aliphatic amides 27–31 had considerable potency
and selectivity for the κ receptor. The values for the ratio of
κ/µ receptors for 27–31 were 15.2, 0.8, 3.5, 2.7, and 1.6,
respectively, but the ratio of δ/µ for these same compounds
was 137, 102, 122, 175, and 74, respectively. Compounds 27–31
thus possessed considerable potency and selectivity for the κ
but not the δ-opioid receptor. Of the amides examined, the
thiophene analogue (i.e., compounds 32 and 33) showed
significant stereoselectivity for binding to the δ-opioid receptor.
Thus, the R isomer 33 had 15.7-fold greater potency than the ꢀ
isomer, compound 32, for binding to the δ-opioid receptor.
Compound 33 possessed similar potency as naltrexone and
nalmefene for the µ- and κ-opioid receptor and showed higher
potency for the δ receptor compared to naltrexone or nalmefene.
The naltrexamides 8–33 were prepared in good yields
according to the synthetic method outlined in Scheme 1.
Naltrexone 4 was converted to its oxime 619 in quantitative yield
using hydroxylamine hydrochloride in the presence of sodium
acetate in refluxing ethanol. Reduction of the oxime functional
group in 6 to the corresponding amine 719b was accomplished
by heating 6 with B2H6/tetrahydrofuran (THF) for 2 days, and
subsequent aqueous workup afforded the amine 7 as a 1:9 (R/
ꢀ) mixture of diastereomers. The diastereomers were separated
by chromatography on silica gel, and the stereochemistry at C-6
was determined by the size of the coupling constant, J5,6. The
amine 7 was coupled either with a carboxylic acid in the
presence of benzotriazol-1-yl-oxy-tris-(dimethylamino)phos-
phonium hexa-fluorophosphate (BOP)a and diisopropylethylamine
or, alternatively, with an acid chloride in triethylamine. The
product was treated with K2CO3 in methanol to remove the side
product resulting from esterification of the 3-position hydroxyl
group, giving amides 8-33 in moderate to high yields. While
the BOP coupling procedure resulted in less esterification of
the 3 position than the acid chloride method, some esterification
at the 3 position could not be avoided. Thus, it was found to be
more convenient to run the reaction with an excess of the acid
derivative to aid in the purification of the intermediate amide
ester. The benzyl groups in compound 8 were removed by
hydrogenolysis in the presence of Pd-C and hydrogen gas to
give the glucose conjugate diastereomers 9 and 10. The structure
of the naltrexamides 8–33 and their respective stereochemistry
are depicted in Table 1. The amides were fully characterized
by nuclear magnetic resonance (NMR) and high-resolution mass
spectrometry (HRMS).
The IC50 values for compounds 4, 5, and 8–33 were obtained
from competition binding assays from full dose–response curves
in the presence of the µ-, δ-, and κ-opioid receptors. The IC50
values for the test compounds and reference materials were
determined using the binding assays with the following radio-
ligands: [3H]DAMGO (µ-opioid receptor agonist), [3H]DPDPE
(δ-opioid receptor agonist), and [3H]U69593 (κ-opioid receptor
agonist). The IC50 values determined by measuring the inhibition
of binding of the radioligands to the receptors by the test
compounds 4, 5, and 8–33 were converted into Ki values as
described in the Experimental Section and listed in Table 2.
In comparison to naltrexone or nalmefene, compounds 4 and
5 as well as saccharide-containing compounds 9 and 10,
generally possessed lower potency than naltrexone for the δ
and κ receptors but equal to or greater potency for the µ-opioid
receptor. The selectivity of compounds 9 and 10 for the µ versus
δ and µ versus κ receptors were greater than 50- and 3-fold,
respectively. In comparison to naltrexone but not nalmefene,
Most of the aryl and aliphatic amide compounds examined
(i.e., 8–10, 19–22, 24, and 30–32) had considerably less affinity
for the δ receptor compared to naltrexone and nalmefene and,
with the exception of a few compounds, had Ki values for the
δ receptor greater than 6 nM. The lipophilic compounds 11 and
12 had Ki values of 1.4 and 0.8 nM, respectively, and along
with 33 (i.e., Ki ) 2.1 nM) showed the greatest potency for the
δ receptor. Of the compounds tested, the polar saccharide 10
and carboxyl-containing compound 26 possessed the lowest
potency for the δ-opioid receptor.
In keeping with our initial observations for lipophilic derivatives
(e.g., compound 8), numerous lipophilic aryl and aliphatic amides
showed considerable potency for the κ receptor. The compounds
with the greatest potency against the κ-opioid receptor were
lipophilic (i.e., 28, Ki ) 0.09 nM; 12, Ki ) 0.1 nM; or 15, Ki )
0.16 nM) or possessed hydrogen-bonding capability in the meta
or para position (i.e., 13, Ki ) 0.06 nM; 20, Ki ) 0.29 nM; 22, Ki

Potential Alcohol-Cessation Agents
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1915
Table 1. Chemical Structures of Amides 8–33 and Their C-6 Diastereomeric Assignment
a Diastereomeric mixture containing R and ꢀ isomers (1:9).
) 0.4 nM; or 31, Ki ) 0.42 nM). In agreement with a previous
report,20 related negatively charged compounds (i.e., 26, Ki ) 151
nM) had considerably less affinity for the κ receptor.
To evaluate the opioid-receptor-mediated activation of its
associated G protein, test compounds were evaluated using the
[35S]GTPγS assay.21 In this assay, the potency or affinity of a
compound for the receptor is associated with its EC50 value for
stimulating [35S]GTPγS binding. Agonist activity is determined
using a selective full agonist (i.e., DAMGO for µ, DPDPE for
δ, and U69,593 for κ) as standards, compounds that are capable

1916 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Scheme 1. Synthesis of Naltrexamide Derivatives 8–33
Ghirmai et al.
Table 2. Ki Inhibition Values and Selectivity of µ-, δ-, and κ-Opioid Binding to CHO Membranes
Ki (nM) ( SEM
compound
µ
δ
κ
δ/µ
49
15
16.5
55
150
30
7.9
21
36
54
22
36
59
130
68
180
85
32
κ/µ
naltrexone, 4
nalmefene, 5
0.30 ( 0
16.31 ( 1.10
13.26 ( 0.75
24.52 ( 4.20
36.94 ( 6.20
99.00 ( 1.50
1.40 ( 0.10
0.79 ( 0.10
2.73 ( 0.50
5.82 ( 0.90
9.72 ( 1.40
4.23 ( 0.20
6.83 ( 1.00
12.40 ( 0.90
27.18 ( 5.40
17.59 ( 2.60
56.30 ( 12.50
36.52 ( 6.10
3.80 ( 0.40
25.29 ( 5.60
8.50 ( 1.00
79.23 ( 3.30
8.22 ( 0.70
12.20 ( 1.40
15.80 ( 2.00
28.00 ( 0.40
19.91 ( 0.60
32.40 ( 0.20
2.07 ( 0.20
0.81 ( 0.02
1.03 ( 0.19
0.51 ( 0.09
2.03 ( 0.16
4.50 ( 0.07
0.37 ( 0.11
0.10 ( 0.01
0.06 ( 0.02
0.72 ( 0.05
0.16 ( 0.02
0.28 ( 0.09
2.41 ( 1.00
0.78 ( 0.16
0.73 ( 0.06
0.29 ( 0.06
0.39 ( 0.07
0.40 ( 0.03
0.24 ( 0.03
0.79 ( 0.19
1.70 ( 0.40
150.61 ( 17.00
0.91 ( 0.01
0.09 ( 0.02
0.45 ( 0.09
0.43 ( 0.03
0.42 ( 0.05
1.20 ( 0.03
2.74 ( 0.93
2.5
1.1
0.34
3.0
7.0
7
1.0
0.5
4.5
0. 89
1.5
13
3.7
3.5
1.1
1.3
0.93
2.0
0.86
1.42
14.94
20
0.8
3.5
2.7
1.6
2.8
7.2
0.91 ( 0.10
1.49 ( 0.07
0.67 ( 0.12
0.64 ( 0.01
0.05 ( 0.01
0.10 ( 0.03
0.13 ( 0.04
0.16 ( 0
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
0.18 ( 0.04
0.19 ( 0.05
0.19 ( 0.02
0.21 ( 0
0.21 ( 0
0.26 ( 0.12
0.31 ( 0.05
0.43 ( 0.17
0.12 ( 0
0.92 ( 0.05
1.20 ( 0.10
10.08 ( 2.50
0.06 ( 0.05
0.12 ( 0.01
0.13 ( 0.03
0.16 ( 0.02
0.27 ( 0.05
0.43 ( 0.10
0.38 ( 0.07
28
7.08
7.860
100
100
120
180
74
75
5.5
Table 3. Stimulation of [32S]-GTP-γ-S Binding by the µ-, δ-, and κ-Opioid Receptors
µ
δ
κ
compound
EC50
Emax
EC50
Emax
EC50
Emax
naltrexone, 4
>10 000
230.00 ( 0
20.40 ( 0
2.10 ( 0
5.30 ( 0
nalmefene, 5
>10 000
31.60 ( 0
11.80 ( 0
2.86 ( 0. 70
7.30 ( 2.45
18.00 ( 5.00
7.10 ( 2.70
0.80 ( 0.20
1.53 ( 0.10
1.09 ( 0
12.60 ( 4.50
49.00 ( 2.50
11.60 ( 0.40
30.60 ( 2.60
91.40 ( 4.10
93.00 ( 11.00
104.30 ( 15.70
74.00 ( 0.80
46.30 ( 5.10
8a
9.70 ( 0.50
42.30 ( 5.40
3.90 ( 2.50
2.30 ( 1.40
1.16 ( 0.60
4.64 ( 0.90
24.80 ( 0
21.40 ( 0.50
15.80 ( 1.60
45.80 ( 3.80
84.80 ( 7.40
20.60 ( 1.50
19.30 ( 0.10
14.50 ( 3.30
16.10 ( 1.90
182.20 ( 53.30
56.00 ( 25.00
22.20 ( 7.60
1.40 ( 1.40
10.40 ( 0.60
8.51 ( 0.90
18.00 ( 1.00
17.58 ( 4.90
15.40 ( 3.60
34.20 ( 6.30
28.70 ( 7.60
84.90 ( 34.60
92.50 ( 4.40
100.20 ( 2.70
24.40 ( 2.60
81.80 ( 4.80
10
11
12
23a
25a
28
1.55 ( 0.10
94.86 ( 3.70
32a
5.20 ( 1.60
a Diastereomeric mixture containing R and ꢀ isomers in a 1:9 ratio.
of maximally stimulating [35S]GTPγS binding in the human
cloned cell line. Table 3 shows the Emax and EC50 values for
stimulation of [35S]GTPγS binding of standards and test
compounds in the cloned human cell membranes containing the
µ-, δ-, or κ-opioid receptor. With the exception of compounds
12 and 25 that showed significant agonist activity, compounds
8, 10, 23, and 28 showed significant antagonism of the µ
receptor. The [35S]GTPγS-binding data showed that compounds
23 and 25 were full δ agonists and compounds 8, 10, 11, and
28 were found to be partial δ agonists similar to naltrexone
and nalmefene. Compounds 12, 23, and 25 were observed to
be full κ agonists, whereas compounds 8, 11, 28, and 32 were
partial κ agonists. Compound 10 showed noncompetitive
antagonist activity against the κ-opioid receptor similar to

Potential Alcohol-Cessation Agents
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1917
Table 4. Metabolic Stability of Selected Compounds in Hepatic
nalmefene. The overall rank order of the EC50 values for the
functional assay correlated well with the Ki values derived from
the binding experiments. As described below, further kinetic
analysis was performed to characterize the properties of these
latter compounds.
Preparationsa
half-life (in min)
rat liver
microsomes
mouse liver
microsomes
human
S-9
compound
High-affinity compounds (i.e., compounds possessing low Ki
values) that showed no or low agonist activity (20% stimulation
or less in the GTPγS-binding experiment) were tested as
antagonists. A Schild analysis was conducted, using a full
dose-response curve in the presence of at least three concentra-
tions of the putative antagonist.22 If the Schild slope was
significantly different from -1.0, the antagonist activity was
judged to be noncompetitive; and in such cases, the pA2 values
were not reported, only the equilibrium dissociation constant
(Ke). Compounds 8, 10, 23, 28, and 32 were found to be full
antagonists for the µ-opioid receptor, while compound 10 was
also found to be a noncompetitive antagonist at the κ receptor.
nalmefene, 5
100
20
255
no change
49
20
11
12
14
no changeb
no change
no change
no change
346
no change
a Incubations were carried out as described in the Experimental Section.
The values are an average of two determinations, and each time point was
run in duplicate. The range of variation was less than 10%. b During the
incubation with the hepatic preparation and the NADPH-generating system,
no detectable loss of parent compound was observed.
liver S-9 (that represents both cytosolic and microsomal
enzymes), similar metabolic stability was observed. This sug-
gests that hepatic cytosolic or microsomal amidases do not
contribute in a dominant way to the metabolism of the target
compounds.
Structure–Activity Studies. The structure–activity relation-
ship (SAR) of the aromatic amide portion of the opioid
antagonists was examined. In general, meta- or para-monosub-
stituted or meta, para-disubstituted aromatic groups showed
quite potent affinity for the µ receptor (Table 1). Thus,
compounds 12–24 and 27–33 all had Ki values in the 0.1-0.9
nM range. The 4-chloro- and 3,4-dichloro-substituted aromatic
amides showed the greatest affinity for the µ receptor (i.e., Ki
) 0.05 and 0.1 nM, respectively). In contrast, of the compounds
tested, compounds 25 and 26 with a para-carboxylic acid group
or meta-N,N-dimethylamino group that should be charged at
neutral pH showed the lowest affinity for the µ receptor (i.e.,
Ki ) 1.2 and 10 nM, respectively). Compound 26 also possessed
very low potency against the δ receptor (i.e., Ki ) 79 nM) and
the κ receptor (Ki ) 151 nM). As described above, there was
some evidence that, in a few cases, compared to the ꢀ isomer,
the R diastereomer possessed greater potency for the µ and δ
receptors (i.e., 9 > 10, 17 > 20, or 33 > 32) but the
stereoselectivity was not large. In the cases examined, the ꢀ
diastereomer had greater potency than the R diastereomer for
the κ receptor (i.e., 20 > 17, 23 >14, and 32 > 33). Because
of the modest diastereoselectivity observed and the paucity of
R diastereomer available by chemical synthesis, this aspect was
not pursued further.
CYP Inhibition. As reported previously, cyclopropylmethyl-
containing amines can inhibit CYP.23,24 To understand the
metabolic stability data described above and to examine the
possible extent and selectivity of CYP inhibition, selected
compounds (i.e., 11, 12, 23, and 25) were examined for their
ability to inhibit selective functional activities of human CYP
enzymes. The observed percent inhibition for selective functional
inhibition of CYP-3A4, -2E1, -2B6, --C9, -2C19, and -2D6
were reported in Table 5. The enzyme assays were performed
using standard conditions as previously described.25 In com-
parison to nalmefene, generally, the compounds examined (i.e.,
11, 12, 23, and 25) possessed less inhibitory potency against
the CYPs examined. A possible exception was CYP2C19 that
appeared to be more sensitive to inhibition by 11, 12, 23, and
25 than nalmefene. In addition, compound 23 inhibited CYP2B6
with greater potency than nalmefene. The results suggest that
replacement of the C-6 exo-methylene group of nalmefene with
an amide substituent in this series attenuated the inhibitory
potency toward CYP. This suggests a significant contribution
of the C-6 moiety in the interaction of nalmefene with CYP
and for the C-6 substituted amides examined; it suggests a
decreased interaction with CYP. This may in part explain some
of the metabolic stability observed for the compounds in this
and related series.26 On the basis of the data from the in Vitro
metabolism studies, the compounds were sufficiently stable and
of low CYP inhibitory potency and were judged to be good
candidates to study them in ViVo in an animal model of alcohol
self-administration.
In ViWo Studies. We evaluated selected compounds for their
effect on baseline ethanol (EtOH) intake in rats trained to self-
administer a 10% (w/v) ethanol solution, using operant tech-
niques. A control group consisting of rats administered nalmefene
hydrochloride was also examined. The operant procedure
provided an animal model system to examine the effect of test
compounds on the acute positive reinforcing effects of EtOH
self-administration.27–34 Initially, dose range studies were
performed, and if active compounds were observed, more
detailed studies were performed. After administration of 0.5 mg/
kg of 10 or 28, a slight increase in alcohol consumption was
observed, and consequently, further studies were not pursued
with these compounds. Dose range studies of 23 and 25 were
attempted; however, a decrease in alcohol self-consumption only
at an elevated dose (i.e., 75% decrease at 2 mg/kg and 17%
decrease at 1.5 mg/kg for 23 and 25, respectively) was observed,
and the lack of efficacy observed precluded further study.
Compound 12 showed potent inhibition of alcohol self-
Metabolic Stability. As a prelude to studying the test
compounds in ViVo, high-performance liquid chromatography
(HPLC)-based analytical methods (see the Supporting Informa-
tion) and biochemical assays were used to assess the metabolic
stability of selected compounds in the presence of rat, mouse,
and human liver preparations and the appropriate NADPH-
generating system. These studies were performed to ascertain
the stability of the compounds toward oxidative metabolism in
advance of more detailed studies with highly purified CYPs as
well as to determine if the compounds possessed sufficient
metabolic stability for in ViVo studies. In comparison to
nalmefene, the candidate compounds 11, 12, and 14 were
metabolically quite stable (Table 4). With the exception of
compound 14, compounds 11 and 12 possessed half-life values
in excess of 255 min in the hepatic preparations used. Generally,
compounds that were metabolically stable in the presence of
mouse or human liver microsomal preparations did not afford
evidence of significant amounts of metabolite formation based
on inspection of the HPLC profiles (data not shown). It may be
that, for this class of compounds, the cyclopropyl methyl moiety
inhibited hepatic CYPs. This point is considered in greater detail
below. Rat and mouse liver microsomes are known to possess
high amounts of amidase activity, and in comparison to human

1918 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Ghirmai et al.
Table 5. Percent Inhibition of CYP3A4, CYP2B6, CYP2C9, CYP2C19, and CYP2D6 by Selected Naltrexamides and Nalmefene
percent inhibitiona
compound
CYP3A4
CYP2B6
CYP2C9
CYP2C19
CYP2D6
nalmefene, 5
60.0 ( 8.7
30.8 ( 3.3
35.5 ( 0.8
44.9 ( 7.8
39.9 ( 3.7
5.2 ( 0.5
7.0 ( 0.3
4.7 ( 0.1
49.3 ( 1.9
NDb
35.5 ( 1.7
28.1 ( 2.8
32.3 ( 1.7
NDb
17.1 ( 1.9
39.1 ( 1.9
48.3 ( 2.2
47.1 ( 5.4
70.0 ( 3.1
30.9 ( 1.9
4.7 ( 0.3
9.8 ( 0.4
31.2 ( 2.1
23.5 ( 1.7
11
12
23
25
23.1 ( 2.5
a Percent inhibition in the presence of 10 µM opioid antagonist ( standard deviation. Incubations were carried out as described in the Experimental
Section. The test compound was preincubated for 2–5 min with the enzyme and cofactor Then, the appropriate substrate (5 µM) was added, and the rate of
product was monitored and compared to the complete system without the test compound present. b ND ) no detectable inhibition was observed at the
concentration of the test compound examined.
administration at a dose of 0.025 mg/kg (i.e., 77% inhibition),
but 12 also showed potent analgesic activity and was not studied
further. Compound 11 was administered in a dose range study
and showed significant efficacy. A more detailed study employ-
ing 11 from 0.1 to 2 mg/kg showed efficacy at inhibiting alcohol
self-administration with an ED50 value of 0.5 mg/kg. This is
about the same ED50 value observed for naltrexone hydrochlo-
ride in similar experiments. In comparison, the ED50 value for
inhibition of alcohol self-administration by nalmefene was
approximately 0.04 mg/kg. We surmised that the potency of
11 for all three µ-, δ-, and κ-opioid receptors coupled with its
metabolic stability contributed to the functional activity as an
alcohol self-administration cessation agent observed in ViVo.
antagonism of the µ receptor. Compound 11 showed partial
agonism at all three opioid receptors. The addition of a meta-
chloro substituent to 11 provided a compound (i.e., 12) that
elicited considerable agonistic potency across all three receptors.
Thus, introduction of small substituent changes resulted in
dramatic biological effects in this series. The antagonism of the
κ receptor by 10 was confirmed by a Schild plot (Table 6). The
finding in the in ViVo studies that 11 was much more potent as
an alcohol-cessation agent than 10 suggests that the partial
agonist activity at all three opioid receptors might be necessary
for alcohol-cessation activity. This will be further evaluated
using other partial agonist compounds.
Hepatotoxicity of naltrexone5 and nalmefene is a concern
because, generally, the liver of individuals that abuse alcohol
is severely compromised. Although the mechanism of hepa-
toxicity of naltrexone and nalmefene is unknown, it is likely to
involve metabolic bioactivation to a reactive intermediate (i.e.,
possibly to an epoxide in the case of nalmefene or radical
intermediates in the case of naltrexone), and because a relation-
ship between metabolic bioactivation and hepatotoxicity has
been established,37 the original design in our studies was to
decrease the metabolic lability of nalmefene by synthesizing
analogues with increased bioavailability as a result of modifica-
tion of the C-6 position. As shown in Tables 4 and 5, compounds
11, 12, 14, 23, and 25 possessed greater metabolic stability and
generally less propensity to interact with hepatic CYP than
nalmefene. It may be that by decreasing the affinity of the opioid
analogues described herein for metabolic enzymes including
CYP and increasing the metabolic stability results in a class of
compounds with less potential for hepatotoxicity.
Conclusion
A series of C-6-substituted amides of naltrexamine where the
amide was appended with substituted aromatic or aliphatic
groups was prepared by the reaction of 6-naltrexamine, 7, with
aryl and aliphatic acid chlorides or carboxylic acids. Although
this is not the first report of the pharmacological properties of
N-acyl-ꢀ-naltrexamines, to our knowledge, this is the first study
examining a number of naltrexamides as alcohol-cessation
agents. The target compounds were evaluated as ligands for the
human µ-, δ-, and κ-opioid receptors, with the goal of identifying
a potent and selective alcohol self-administration cessation agent.
Originally, metabolically stable saccharide derivatives of opioids
were studied as pain medication candidates.35 It was noted that
glucuronides were less potent than glucose derivatives.35 In the
present study, we initially examined glucose derivatives as
alcohol-cessation agents but the compounds examined (e.g., 10)
did not possess significant potency in Vitro against the δ and κ
receptors and was not efficacious in ViVo. Consequently, 10 was
not studied further, in ViVo. Next, we synthesized analogues of
saccharides, such as 13, 14, 17, and 20, where the substituent
mimicked the polar functionalities of a saccharide and they
retained significant potency. On that basis, we made a number
of analogues to explore SAR. In general, meta- or para- or meta,
para-disubstituted aromatic groups showed quite potent affinity
for the µ receptor. There was not a marked difference in potency
between meta or para-substituted aryl amides. Generally, the
aliphatic amides examined all showed significant potency for
the µ receptor. With the exception of 11, generally, meta-
substituted aryl amides were somewhat more potent than para-
substituted aryl amides against the δ receptor. Similar to the µ
receptor, both meta- and para-substituted aryl amides possessed
considerable potency against the κ receptor. However, some aryl
substituents abrogated potency. For example, as described
previously,36 negatively charged carboxyl-containing com-
pounds, such as 26, had poor affinity for the µ and κ receptors.
As shown in Table 3, not all of the compounds stimulated full
[32S]GTP-γ-S binding at the µ, δ, and κ receptors. Compounds
8, 10, 23, 25, 28, and 32 showed considerable apparent
On the basis of the results of the functional assay and Ki
values and metabolic stability studies shown herein, selected
compounds were evaluated for their effects in an animal model
of alcohol self-administration. One objective was to try to
determine which opioid receptor contributed to alcohol self-
administration cessation. Compound 10 showed little agonistic
activity at all three receptors and was not effective in ViVo and,
at 0.5 mg/kg, actually stimulated a modest amount of alcohol
consumption in rats. On the other hand, compounds with
significant agonist activity at the δ and/or κ receptor (i.e., 12,
23, 25, and 32) either possessed low potency as alcohol-cessation
agents or stimulated alcohol self-administration in ViVo. Com-
pound 28 possessed significant agonistic activity against the κ
but not the µ or δ receptors. However, at 0.5 mg/kg, 28
stimulated alcohol self-administration. Of the compounds
examined, compound 11 (that was a weak agonist against all
three receptors) showed the greatest efficacy in ViVo (i.e.,
approximately ED50 of 0.5 mg/kg). On the basis of limited data
presented above, it appears that individual antagonism of the
µ, δ, or κ receptor alone is not sufficient for alcohol self-
administration cessation. It may be that potent κ antagonism
can compensate for weak δ antagonism, but what is more likely

Potential Alcohol-Cessation Agents
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1919
Table 6. pA2 Data for Selected Naltrexamide Analogues
compound
receptor
Ke (nM) ( SEM
pA2 ( SEM
slope ( SEM
naltrexone, 4
µ
δ
κ
µ
δ
κa
µ
δa
κ
µ
δ
κa
µ
δ
κ
µ
δ
κ
0.05 ( 0
2.10 ( 0.30
0.23 ( 0.02
0.34 ( 0.02
1.02 ( 0.13
0.15 ( 0.03
0.16 ( 0.02
10.35 ( 2.00
PAc
10.30 ( 0.03
8.80 ( 0.10
9.68 ( 0.03
9.53 ( 0.03
9.04 ( 0.06
-1.06 ( 0.03
-0.91 ( 0.04
-0.96 ( 0.02
-0.93 ( 0.02
-0.95 ( 0.03
nalmefene, 5
8b
9.68 ( 0.07
9.33 ( 0.05
9.78 ( 0.04
9.98 ( 0.04
-1.07 ( 0.04
-1.08 ( 0.04
-0.99 ( 0.02
-0.92 ( 0.02
10
0.37 ( 0.03
PAc
13.40 ( 1.50
0.17 ( 0.01
FAd
23b
25
FAd
0.14 ( 0.01
FAd
FAd
28
µa
δ
κ
µ
δ
κ
0.15 ( 0.02
PAc
PAc
32b
0.41 ( 0.02
9.34 ( 0.02
-1.05 ( 0.02
PAc
PAc
a Noncompetitive antagonist. b Diastereomeric mixture containing R and ꢀ isomers (1:9). c PA ) partial agonist. d FA ) full agonist.
o.d. × 250 mm) was used (condition A), the mobile phase was
is that alcohol self-administration efficacy requires antagonism
MeOH/2-propanol/HClO4 (55:45:0.01, v/v/v) at a flow rate of 1
mL/min. When a reverse-phase system was used (condition B),
HPLC was performed with a L-7100 Hitachi pump, a L-7400
UV–vis Hitachi detector (285 nM), a L-7200 Hitachi autosampler,
and a D-7000 Hitachi chromato-integrator employing a Supleco
reverse-phase column (5 µm × 4.6 mm × 250 mm). The mobile
phase was 20% 0.05 M tetrabutylammonium hydroxide and 80%
methanol using isocratic elution at a flow rate of 1 mL/min.
of all three opioid receptors. Such an agent (i.e., 11) showed
good in ViVo activity in an animal model of ethanol self-
administration in rats trained to self-administer a 10% (w/v)
ethanol solution. Another interpretation of the data is the
possibility that the compounds with µ partial agonist activity
are sufficiently rewarding in themselves that they effectively
substitute for the rewarding effects of ethanol. Compound 11
and related agents may represent an exciting lead for developing
the next generation of opioid compounds useful in the treatment
of alcohol abuse.
Microsomes from rat, mouse, and human liver expressing
functional human cytochrome P-450s were purchased from BD
Gentest (Woburn, MA) or made in house, and microsomes from
baculovirus-infected cells co-expressing cytochrome P-450s (3A4,
2B6, 2C9, 2C19, and 2D6), NADPH-cytochrome P-450 reductase,
and cytochrome b5 (BACULOSOMES) were purchased from
PanVera LLC (Madison, WI).
Experimental Section
Chemicals. Nalmefene hydrochloride and naltrexone hydrochlo-
ride were obtained from Tyco Mallinckrodt (St. Louis, MO).
NADP+, glucose-6-phosphate, glucose-6-phosphate dehydrogenase,
diethylenetriaminepentaacetic acid (DETAPAC), and MgCl2 were
all obtained from Sigma-Aldrich Chemical Co. (St. Louis, MO)
and were used as received. All of the solvents and buffers were
obtained in the highest grade commercially available from VWR
(San Diego, CA).
Naltrexone Oxime (6).19 Naltrexone (500 mg, 1.46 mmol),
NH2OH-HCl (147 mg), and NaOAc (294 mg) were dissolved in
absolute ethanol (8 mL), and the mixture was heated at reflux for
2.5 h and then concentrated to dryness. Water (20 mL) was added,
and the mixture was made basic with K2CO3 and extracted with
CHCl3. The CHCl3 extract was washed with brine, dried over
Na2SO4, filtered, and concentrated to give a white solid (463 mg,
General Procedures. All reactions were run under a positive
pressure of nitrogen with magnetic stirring at ambient temperature
using oven-dried glassware unless otherwise indicated. Air- and
moisture-sensitive liquids were transferred via syringe through
rubber septa. Silica gel (230–400 mesh) was used for column
chromatography. N,N-Dimethylformamide (DMF) was dried by
filtration through a column of neutral alumina and stored over
activated 4 Å molecular sieves under nitrogen prior to use. All other
solvents and reagents were used as received. 1H and 13C NMR
were recorded at 300.0 and 75.4 MHz, respectively, on a Varian
Mercury 300 instrument. Chemical shifts were reported in parts
per million (δ) relative to CDCl3 at 7.26 and 77 ppm, respectively.
NMR spectra were recorded in CDCl3 unless stated otherwise.
Melting points were reported uncorrected. High-resolution mass
spectra were obtained with a VG 7070 spectrometer with an Opus
V3.1 and DEC 3000 Alpha Station data system at the University
of California, Riverside, or a Waters LCT Premier instrument
operating in the ESI mode at the University of California, Irvine.
Analytical purities were determined by straight and reverse-phase
HPLC using a Hitachi D2500 Hitachi Chromato-integrator, a
L-6000 Hitachi pump, and a L-4200 UV–vis Hitachi detector (285
nM). When an AXXI-chrom silica column (5 µm i.d. × 4.6 mm
1
89%). ESI-MS m/z: 357 (MH+). H NMR (CDCl3) δ: 6.75 (d, J
) 8.2 Hz, 1H), 6.61 (d, J ) 8.2 Hz, 1H), 5.00 (s, 1H), 3.15 (m,
2H), 2.65–1.30 (m, 10H), 0.86 (m, 1H), 0.56 (m, 2H), 0.20 (m,
2H).
6-ꢀ-Naltrexamine (7a) and 6-r-Naltrexamine (7b). Naltrexone
oxime (5.83 g, 16.3 mmol) was dissolved in THF (40 mL) and
transferred by cannula over 10 min to a solution of BH3/THF (300
mL, 300 mmol, 1 M solution in THF) held at 10 °C. A white
precipitate formed and then slowly dissolved as the reaction was
heated at reflux for 48 h. The solution was cooled to room
temperature, and water (10 mL) and 1 N KOH (200 mL) were added
slowly. The solution was then reheated at reflux for 2 h. The pH
was reduced to 2.5 with 10% HCl (225 mL), and the solution was
heated at reflux for 2 h, concentrated to remove THF, and then
made basic (pH 8–9) with K2CO3. The mixture was extracted with
CHCl3 (4 × 150 mL), and the extract was dried over Na2SO4,
filtered, and concentrated. The resulting oil was purified by
chromatography on SiO2 [26 × 60 cm, elution with 25:5:1 CH3CN/
MeOH/NH4OH (v/v/v)] providing 7a (2.14 g, 38%) as a white-
1
yellow solid. Rf ) 0.2. H NMR (300 MHz, CDCl3 with 2 drops
of CD3OD) δ: 6.61 (d, J ) 8.1 Hz, 1H), 6.49 (d, J ) 8.1 Hz, 1H),

1920 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Ghirmai et al.
4.17 (d, J ) 7.5 Hz, 1H), 3.39–0.45 (20 H). ESI-MS m/z: 343
(MH+). An additional 0.64 g (12%) of material consisting of a
mixture of the R and ꢀ diastereomers was isolated. Repeated
chromatography gave an analytical sample of the R diastereomer,
amide (100 mg, 0.11 mmol) in anhydrous methanol (4 mL) was
stirred under an atmosphere of hydrogen gas in the presence of
10% activated Pd-C (100 mg) and concentrated HCl (10 µL) for
12 h. The mixture was filtered through a pad of celite (eluted with
MeOH), and the filtrate was concentrated to dryness. The resulting
residue was triturated with ether, and this afforded a white solid
(49 mg, 82%). mp ) 250 °C. Rf ) 0.80 [5:4:0.5:0.5 CHCl3/MeOH/
1
compound 7b. Rf ) 0.16. H NMR δ: 6.65 (d, J ) 8.1 Hz, 1H),
6.46 (d, J ) 8.1 Hz, 1H), 4.50 (d, J ) 3.0 Hz, 1H), 3.34 (dt, J )
3.9, 12.6 Hz, 1H), 3.04 (t, J ) 6.6 Hz, 1H), 2.95 (s, 1H), 2.63–0.29
(17H). ESI-MS m/z: 343 (MH+).
1
NH4OH/H2O (v/v/v/v)]. H NMR (CD3OD) δ: 6.73 (d, J ) 8.2
Hz, 1H), 6.65 (d, J ) 8.2 Hz, 1H), 4.60 (d, J ) 7.8 Hz, 1H),
3.92–3.53 (m, 5H), 3.28–1.53 (m, 11H), 1.08 (m, 1H), 0.81 (m,
2H), 0.47 (m, 2H). 13C NMR (CD3OD) δ: 173.6, 143.4, 142.7,
130.5, 121.7, 120.9, 119.5, 92.2, 81.5, 79.4, 77.9, 75.0, 71.9, 71.2,
64.2, 63.0, 58.8, 52.6, 47.5, 40.2, 31.1, 28.9, 24.6 (2C), 7.0, 6.4,
3.6. ESI-MS m/z: 547 (MH+). HRMS Calcd for C28H38N2O9,
547.2656; found, 547.2650.
General Procedure for the Amidation of Naltrexamine with
an Acid Chloride. Naltrexamine (104 mg, 0.3 mmol) was dissolved
in CH2Cl2 (4 mL) and NEt3 (0.13 mL, 0.93 mmol), and substituted
benzoyl chlorides (0.73 mmol) were added. The solution was stirred
for 2 h at room temperature and concentrated to dryness. The residue
was filtered through a column of SiO2 [20:1 CH2Cl2/MeOH (v/v)].
The resulting solid was dissolved in anhydrous methanol (3 mL),
and K2CO3 (300 mg) was added. The mixture was stirred at room
temperature for 12 h, concentrated, and purified by SiO2 chroma-
tography.
General Procedure for the Amidation of Naltrexamine with
a Carboxylic Acid. Naltrexamine (100 mg, 0.29 mmol), substituted
benzoic acid (0.58 mmol), and BOP (258 mg, 0.58 mmol) were
dissolved in CH2Cl2 (3 mL). To this solution, i-Pr2EtN (0.15 mL,
0.88 mmol) was added and the mixture was stirred at room
temperature for 2 h. The solution was concentrated and filtered
through a short column of SiO2 (eluted with EtOAc) providing a
white material. This product was dissolved in MeOH (3 mL), and
K2CO3 (300 mg) was then added. The mixture was stirred at room
temperature for 3 h and concentrated to dryness. The residue was
purified by SiO2 chromatography [20:1 CH2Cl2/MeOH (v/v)] to
provide the target compound.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6r-
[(2′,3′,4′,6′-tetra-O-benzyl-D-glucopyranosyl)acetamido]morphi-
nan (8). The tetra-O-benzyl amide was prepare according to the
general procedure described above; R-naltrexamine (110 mg, 0.32
mmol), 2,3,4,6-tetra-O-benzyl-D-glucopyranosyl-1-ꢀ-1-acetic acid
(283 mg, 0.49 mmol), BOP (215 mg, 0.49 mmol), and i-Pr2EtN
(0.17 mL, 0.98 mmol) were combined to produce the target
compound, and this was followed by basic hydrolysis with K2CO3
(278 mg, 2 mmol) to give the intermediate tetra-O-benzyl amide,
compound 8, as a white solid (254 mg, 87%). Rf ) 0.11 [30:1
CHCl3/MeOH (v/v)]. 1H NMR (CDCl3) δ: 7.31–7.06 (m, 2H), 6.76
(d, J ) 8.2 Hz, 1H), 6.71 (d, J ) 8.2 Hz, 1H), 4.94–4.38 (m, 10H),
3.86–3.79 (m, 3H), 3.25–1.40 (m, 20H), 1.08 (m, 1H), 0.81 (m,
2H), 0.45 (m, 2H). 13C NMR (CDCl3) δ: 170.4, 143.6, 138.1, 137.6,
128.4, 128.4, 128.3, 128.1, 127.9, 127.8, 127.7, 127.6, 118.9, 93.8,
86.8, 81.0, 78.4, 75.8, 75.7, 75.2, 75.0, 73.2, 70.1, 59.2, 50.9, 30.2,
4.1, 3.9. ESI-MS m/z 907 (MH+). HRMS Calcd for C56H62N2O9,
907.4534; found, 907.4548.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(4′-
chloro)benzamido]morphinan (11). Compound 11 was synthe-
sized according to the general procedure described above; com-
bining ꢀ-naltrexamine (100 mg, 0.29 mmol), 4-chlorobenzoic acid
(68 mg, 0.44 mmol), BOP (190 mg, 0.44 mmmol), and i-Pr2EtN
(0.16 mL, 0.87 mmol) followed by basic hydrolysis with K2CO3
gave the title compound as a white solid (28 mg, 20%). mp ) 188.8
1
°C. Rf ) 0.06 [30:1 CHCl3/MeOH (v/v)]. H NMR [9:1 CDCl3/
CD3OD (v/v)] δ: 7.77 (d, J ) 7.8 Hz, 2H), 7.40 (d, J ) 7.8 Hz,
2H), 6.70 (d, J ) 8.4 Hz,, 2H), 6.51 (d, J ) 8.4 Hz, 1H), 4.40 (d,
J ) 6.6 Hz, 1H), 4.15–4.05 (m, 1H), 3.10–1.35 (m, 11H), 0.50 (m,
2H), 0.10 (m, 2H). 13C NMR [9:1 CDCl3/CD3OD (v/v)] δ: 166.8,
142.6, 139.7, 137.6, 132.3, 130.2, 128.5, 128.4, 123.7, 118.9, 118.3,
92.7, 70.5, 62.1, 59.0, 50.8, 47.3, 43.8, 31.2, 29.4, 23.9, 22.5, 9.3,
3.9, 3.7. ESI-MS m/z: 481 (MH+). HRMS Calcd for
C27H29ClN2O4, 481.1894; found, 481.1879.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(3′,4′-
dichloro)benzamido]morphinan (12). Compound 12 was prepared
according to the general procedure described above; combining
ꢀ-naltrexamine (100 mg, 0.29 mmol), 3,4-dichlorobenzoyl chloride
(153 mg, 0.73 mmol), and Et3N (0.15 mL, 0.1 mmol) for 2 h
followed by basic hydrolysis with K2CO3 (1 g) gave the title
compound as a white solid (138 mg, 92%). mp ) 108.6 °C. Rf )
1
0.36 [10:1 CH2Cl2/MeOH (v/v)]. H NMR (CDCl3) δ: 7.93 (d, J
) 1.8 Hz, 1H), 7.67–7.64 (m, 2H), 7.42 (d, J ) 8.4 Hz, 1H), 6.60
(d, J ) 8.1 Hz, 1H), 6.51 (d, J ) 8.1 Hz, 1H), 4.71 (d, J ) 6.3 Hz,
1H), 3.99–3.93 (m, 1H). 13C NMR (CDCl3) δ: 164.7, 142.2, 139.1,
135.6, 133.8, 132.6, 130.4, 130.2, 129.2, 126.2, 124.5, 119.3, 117.5,
92.3, 70.3, 62.2, 59.3, 51.2, 47.4, 43.9, 31.6, 29.4, 23.5, 22.7, 9.5,
4.1, 4.0. ESI-MS m/z: 516 (MH+). HRMS Calcd for
C27H28Cl2N2O4, 515.1505; found, 515.1498.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(3′-
hydroxy)benzamido]morphinan (13). Compound 13 was synthe-
sized according to the general procedure described above; com-
bining ꢀ-naltrexamine (100 mg, 0.29 mmol), 3-hydroxybenzoic acid
(105 mg, 0.58 mmol), BOP (258 mg, 0.58 mmol), and i-Pr2EtN
(0.15 mL, 0.88 mmol) followed by basic hydrolysis with K2CO3
gave the title compound as a white solid (90 mg, 67%). mp ) 194.7
°C. Rf ) 0.11 [15:1 CHCl3/MeOH (v/v)]. 1H NMR (CDCl3) δ:
7.59 (d, J ) 9.0 Hz, 1H), 7.26–7.23 (m, 3H), 6.97–6.94 (m, 1H),
6.70 (d, J ) 8.1 Hz, 1H); 6.54 (d, J ) 8.1 Hz, 1H), 4.40 (d, J )
6.3 Hz, 1H), 4.15–4.10 (m, 1H). 13C NMR (CDCl3) δ: 167.6, 156.5,
142.6, 139.6, 134.7, 130.5, 129.6, 124.1, 119.4, 119.1, 118.1, 117.8,
115.3, 93.5, 70.4, 62.2, 59.2, 51.4, 47.6, 44.0, 31.2, 29.8, 24.3, 22.7,
9.5, 4.2, 3.9. ESI-MS m/z: 463 (MH+). HRMS Calcd for
C27H30N2O5, 463.2233; found, 463.2225.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6r-[(D-
glucopyranosyl)acetamido]morphinan (9). The tetra-O-benzyl
amide, 8 (100 mg, 0.11 mmol), was stirred under an atmosphere
of hydrogen gas in the presence of 10% Pd-C (100 mg) and
concentrated HCl (0.01 mL) for 12 h, filtered through a pad of
celite (eluted with MeOH, 100 mL), and concentrated to dryness.
The resulting residue was triturated with ether (3 × 2 mL),
providing a white solid (37 mg, 62%) of compound 9. Rf ) 0.06
1
[25:5:1 CH3CN/MeOH/NH4OH (v/v/v)]. mp > 300 °C (dec). H
NMR (CD3OD) δ: 6.75 (d, J ) 8.2 Hz, 1H), 6.67 (d, J ) 8.2 Hz,
1H), 4.68 (d, J ) 3.8 Hz, 1H), 3.91–3.55 (m, 5H), 3.28–2.38 (m,
7H), 1.89–1.53 (m, 4H), 1.08 (m, 1H), 0.81 (m, 2H), 0.47 (m, 2H).
HRMS Calcd for C28H38N2O9, 547.2656; found, 547.2646.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-(D-
glucopyranosyl)acetamido]morphinan (10). Compound 10 was
prepared according to the general procedure described above;
combining ꢀ-naltrexamine (66 mg, 0.19 mmol), 2,3,4,6-tetra-O-
benzyl-D-glucopyranosyl-1-ꢀ-1-acetic acid (170 mg, 0.29 mmol),
BOP (129 mg, 0.29 mmol), and i-Pr2EtN (0.1 mL, 0.58 mmol) as
above for 2 h gave the phenolic amide as a white solid (104 mg,
59%). The phenolic ester C-6 amide side product was not observed
by MS or TLC analysis of this sample. Rf ) 0.14 [30:1 CHCl3/
MeOH (v/v)]. ESI-MS m/z: 907 (MH+). A solution of the phenolic
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6r-[(3′-
methoxy)benzamido]morphinan (14). Compound 14 was prepared
according to the general procedure described above; combining
R-naltrexamine (104 mg, 0.3 mmol), 3-methoxybenzoyl chloride
(0.1 mL, 0.73 mmol), and Et3N (0.13 mL, 0.93 mmol) for 2 h
followed by basic hydrolysis with K2CO3 (300 mg) gave the title
compound as a white solid (134 mg, 92%). mp ) 249 °C. Rf )
1
0.20 [20:1 CH2Cl2/MeOH (v/v)]. H NMR (CDCl3) δ: 7.40–7.28
(m, 4H), 6.70 (d, J ) 8.2 Hz, 1H), 6.54 (d, J ) 8.2 Hz, 1H), 4.52
(d, J ) 5.7 Hz, 1H), 4.18 (m, 1H), 3.81 (s, 3H). 13C NMR δ: 166.1,

Potential Alcohol-Cessation Agents
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1921
159.6, 143.0, 139.1, 135.8, 130.6, 129.3, 124.6, 119.0, 118.8, 117.6,
117.5, 112.3, 92.8, 70.0, 62.2, 59.3, 55.4, 50.3, 47.3, 43.9, 31.8,
29.1, 23.3, 22.7, 9.5, 4.1, 3.9. ESI-MS m/z: 477 (MH+). HRMS
Calcd for C28H32N2O5, 477.2390; found, 477.2392.
J ) 7.8 Hz, 1H), 4.37 (d, J ) 7.2 Hz, 1H), 3.92 (m, 1H), 3.81 (s,
3H). 13C NMR (9:1 CDCl3/CD3OD) δ: 166.9, 166.3, 142.5, 139.7,
138.0, 132.4, 130.2, 129.4, 127.1, 123.7, 118.9, 118.2, 92.6, 70.5,
62.0, 59.0, 52.3, 50.9, 47.3, 43.8, 31.1, 29.4, 23.9, 22.5, 9.3, 3.9,
3.7. ESI-MS m/z: 505 (MH+). HRMS Calcd for C29H32N2O6,
505.2339; found, 505.2330.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-(ben-
zamido)morphinan (15). Compound 15 was synthesized according
to the general procedure described above; ꢀ-naltrexamine (100 mg,
0.29 mmol), benzoic acid (71 mg, 0.58 mmol), BOP (258 mg, 0.58
mmol), and i-Pr2EtN (0.15 mL, 0.88 mmol) were combined, and
basic hydrolysis with K2CO3 (1 g) provided the title compound as
a white solid (55 mg, 42%). mp ) 139.2 °C. Rf ) 0.12 [20:1
CH2Cl2/MeOH (v/v)]. 1H NMR (CDCl3) δ: 7.77–7.74 (m, 2H),
7.42–7.32 (m, 3H), 6.62 (d, J ) 8.1 Hz, 1H), 6.47 (d, J ) 8.1 Hz,
1H), 4.37 (d, J ) 6.6 Hz, 1H), 4.05 (m, 1H). 13C (CDCl3) δ: 166.9,
143.1, 139.6, 134.2, 131.4, 130.3, 128.4, 127.0, 123.8, 119.1, 117.9,
92.8, 70.1, 62.2, 59.2, 50.2, 47.1, 44.4, 31.2, 29.3, 23.3, 22.9, 8.9,
4.3, 3.9. ESI-MS m/z: 447 (MH+). HRMS Calcd for C27H30N2O4,
447.2284; found, 447.2284.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(3′,5′-
dimethoxy)benzamido]morphinan (20). Compound 20 was syn-
thesized according to the general procedure described above;
ꢀ-naltrexamine (100 mg, 0.29 mmol), 3,5-dimethoxybenzoic acid
(106 mg, 0.58 mmol), BOP (258 mg, 0.58 mmol), and i-Pr2EtN
(0.15 mL, 0.88 mmol) were combined and basic hydrolysis with
K2CO3 (1 g) provided the title compound (140 mg, 95%) as a white
1
solid. mp ) 146.7 °C. Rf ) 0.27 [20:1 CH2Cl2/MeOH (v/v)]. H
NMR (CDCl3) δ: 6.95 (s, 1H), 6.94 (s, 1H), 6.70 (d, J ) 8.1 Hz,
1H), 6.55–6.53 (m, 2H), 4.53 (d, J ) 5.4 Hz, 1H), 4.20–4.11 (m,
1H), 3.80 (s, 6H). 13C NMR (CDCl3) δ: 166.7, 160.6, 142.9, 139.1,
136.5, 130.6, 124.6, 119.1, 117.5, 104.9, 103.5, 92.9, 70.1, 62.2,
59.3, 55.6, 50.4, 47.3, 43.9, 31.7, 29.1, 23.4, 22.7, 9.5, 4.1, 3.9.
ESI-MS m/z: 507 (MH+). HRMS Calcd for C29H34N2O6, 507.2495;
found, 507.2485.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(3′-
nitro)benzamido]morphinan (16). Compound 16 was synthesized
according to the general procedure described above; combining
ꢀ-naltrexamine (100 mg, 0.29 mmol), 3-nitrobenzoyl chloride (135
mg, 0.73 mmol), and Et3N (0.15 mL, 1.1 mmol) for 2 h followed
by basic hydrolysis with K2CO3 (1 g) gave the title compound as
a white solid (72 mg, 50%). mp ) 291 °C. Rf ) 0.36 [10:1 CH2Cl2/
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(2′-
pyridyl)acetamido]morphinan (21). Compound 21 was synthe-
sized according to the general procedure described above; com-
bining ꢀ-naltrexamine (100 mg, 0.29 mmol), nicotinoyl chloride
hydrochloride (130 mg, 0.73 mmol), and Et3N (0.15 mL, 1.1 mmol)
for 2 h followed by basic hydrolysis with K2CO3 (1 g) gave the
title compound as a white solid (81 mg, 61%). mp ) 197.7 °C. Rf
1
MeOH (v/v)]. H NMR (CDCl3) δ: 8.67 (m, 1H), 8.30–8.24 (m,
2H), 7.94 (d, J ) 8.7 Hz, 1H), 7.59 (d, J ) 9 Hz, 1H), 6.52 (d, J
) 8.1 Hz, 1H), 6.46 (d, J ) 8.1 Hz, 1H), 4.81 (d, J ) 6 Hz, 1H),
4.06–3.97 (m, 1H). 13C NMR (CDCl3) δ: 164.4, 148.0, 142.0, 139.0,
135.7, 133.4, 130.5, 129.4, 125.8, 124.7, 121.9, 119.3, 117.2, 92.3,
70.3, 62.1, 59.4, 51.3, 47.4, 43.8, 31.6, 29.4, 23.4, 22.7, 9.5, 4.1,
4.0. ESI-MS m/z: 492 (MH+). HRMS Calcd for C27H29N3O6,
492.2135; found, 492.2124.
1
) 0.23 [15:1 CH2Cl2/MeOH (v/v)]. H NMR (CDCl3) δ: 9.01 (d,
J ) 1.5 Hz, 1H), 8.65 (dd, J ) 1.5, 5.1 Hz, 1H), 8.09–8.05 (m,
1H), 7.80 (d, J ) 5.1 Hz, 1H), 7.32–7.28 (m, 1H), 6.73 (d, J ) 8.1
Hz, 1H), 6.56 (d, J ) 8.1 Hz, 1H), 4.59 (d, J ) 5.7 Hz, 1H), 4.14
(m, 1H). 13C (CDCl3) δ: 164.9, 151.5, 147.7, 143.0, 139.5, 135.5,
130.5, 130.1, 124.3, 123.4, 119.2, 117.9, 92.3, 70.2, 62.2, 59.3,
50.7, 47.4, 44.0, 31.7, 29.2, 23.5, 22.7, 9.5, 4.2, 3.9. ESI-MS m/z:
448 (MH+). HRMS Calcd for C26H29N3O4, 448.2236; found,
448.2233.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6r-[(3′, 5′-
dimethoxy)benzamido]morphinan (17). Compound 17 was syn-
thesized according to the general procedure described above;
combining R-naltrexamine (75 mg, 0.22 mmol), 3,5-dimethoxy-
benzoic acid (76 mg, 0.42 mmol), BOP (193 mg, 0.43 mmol), and
i-Pr2EtN (0.11 mL, 0.63 mmol) for 2 h followed by hydrolysis with
K2CO3 (300 mg) gave the title compound as a white solid (101
mg, 91%). Rf ) 0.20 [10:1 CH2Cl2/MeOH (v/v)]. mp ) 221 °C.
1H NMR (CDCl3) δ: 6.84 (s, 1H), 6.83 (s, 1H), 6.69 (d, J ) 8.4
Hz, 1H), 6.55–6.53 (m, 1H), 6.46 (d, J ) 8.4 Hz, 1H), 4.77–4.70
(m, 2H). 13C NMR (CDCl3) δ: 166.7, 160.5, 144.9, 137.0, 136.8,
130.9, 126.0, 119.2, 117.0, 104.9, 103.3, 90.3, 69.5, 62.1, 59.7,
55.6, 47.3, 46.7, 43.2, 33.6, 29.1, 22.9, 21.1, 9.5, 4.1, 4.0. ESI-MS
m/z: 507 (MH+). HRMS Calcd for C29H34N2O6, 507.2495; found,
507.2481.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(4′-
amino)benzamido]morphinan (22). 17-Cyclopropylmethyl-3, 14ꢀ-
dihydroxy-4, 5R-epoxy-6ꢀ-[(4′-tert-butoxycarbonylamino)benza-
mido]morphinan (24) (71 mg, 0.13 mmol) was dissolved in CH2Cl2
(2 mL), and TFA (2 mL) was added. The solution was stirred at
room temperature for 1 h before the liquids were removed under a
stream of nitrogen. The residue was filtered through a pipet column
of SiO2 [10:1:0.2 CHCl3/MeOH/NH4OH (v/v/v)] and concentrated.
The residue was triturated with ether to provide the title compound
as a white solid (30 mg, 52%). mp ) 215.8 °C. Rf ) 0.24 [10:1
1
CHCl3/MeOH (v/v)]. H NMR (9:1 CDCl3/MeOH) δ: 8.05 (d, J
) 8.7 Hz, 1H); 7.72 (d, J ) 6.9 Hz, 2H), 6.82–6.63 (m, 4H), 4.59
(d, J ) 7.2 Hz, 1H), 4.10 (m, 1H). 13C NMR (CD3OD) δ: 170.0,
153.2, 143.7, 142.8, 130.7, 129.9, 122.7, 121.7, 120.7, 119.6, 114.5,
92.2, 79.4, 71.3, 64.3, 58.7, 53.0, 52.9, 31.2, 28.9, 24.9, 24.5, 6.9,
6.2, 3.4. ESI-MS m/z: 461 (MH+). HRMS Calcd for C27H31N3O4,
461.2440; found, 461.2439.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6r-(ben-
zamido)morphinan (18). Compound 18 was synthesized according
to the general procedure described above; combining R-naltrex-
amine (30 mg, 0.09 mmol), benzoic acid (21 mg, 0.18 mmol), BOP
(77 mg, 0.18 mmol), and i-Pr2EtN (0.05 mL, 0.26 mmol) for 2 h
followed by basic hydrolysis with K2CO3 (300 mg) gave the title
compound as a white solid (17 mg, 43%). mp ) 121.5 °C. Rf )
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(3′-
methoxy)benzamido]morphinan (23). Compound 23 was syn-
thesized according to the procedure described above; combining
ꢀ-naltrexamine (104 mg, 0.3 mmol), NEt3 (0.13 mL, 0.93 mmol),
and 3-methoxybenzoyl chloride (0.1 mL, 0.73 mmol) for 2 h
followed by basic hydrolysis with K2CO3 afforded a white solid
(134 mg, 92%). mp ) 249 °C. Rf ) 0.20 [20:1 CH2Cl2/MeOH
1
0.17 [10:1 CH2Cl2/MeOH (v/v)]. H NMR (CDCl3) δ: 7.77–7.74
(m, 2H), 7.48–7.34 (m, 3H), 6.64 (d, J ) 8.4 Hz, 1H), 6.50 (d, J
) 8.4 Hz, 1H), 4.71–4.63 (m, 2H). 13C NMR (CDCl3) δ: 167.6,
145.2, 137.5, 134.0, 131.2, 130.5, 128.1, 127.0, 125.0, 119.0, 117.0,
89.6, 69.5, 61.9, 59.4, 47.0, 46.5, 43.1, 33.3, 29.1, 22.7, 20.7, 9.1,
4.0, 3.6. HRMS Calcd for C27H30N2O4, 447.2284; found, 447.2276.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(4′-
carbomethoxy)benzamido]morphinan (19). Compound 19 was
synthesized according to the general procedure described above;
combining ꢀ-naltrexamine (100 mg, 0.29 mmol), 4-carbomethoxy-
benzoic acid (105 mg, 0.58 mmol), BOP (258 mg, 0.58 mmol),
and i-Pr2EtN (0.15 mL, 0.88 mmol) for 2 h followed by basic
hydrolysis with K2CO3 (1 g) gave the title compound as a white
solid (118 mg, 80%). Rf ) 0.12 [10:1 CH2Cl2/MeOH (v/v)]. mp )
1
(v/v)]. H NMR (CDCl3) δ: 7.40–7.28 (m, 4H), 6.70 (d, J ) 8.2
Hz, 1H), 6.54 (d, J ) 8.2 Hz, 1H), 4.52 (d, J ) 5.7 Hz, 1H), 4.18
(m, 1H), 3.81 (s, 3H). 13C NMR δ: 166.1, 159.6, 143.0, 139.1,
135.8, 130.6, 129.3, 124.6, 119.0, 118.8, 117.6, 117.5, 112.3, 92.8,
70.0, 62.2, 59.3, 55.4, 50.3, 47.3, 43.9, 31.8, 29.1, 23.3, 22.7, 9.5,
4.1, 3.9. ESI-MS m/z: 477 (MH+). HRMS Calcd for C28H32N2O5,
477.2390; found, 477.2385.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(4′-
tert-butoxycarbonylamino)benzamido]morphinan (24). Com-
pound 24 was prepared according to the general procedure described
1
156.2 °C. H NMR (CDCl3/CD3OD, 9:1) δ: 7.96 (d, J ) 8.8 Hz,
1H), 7.79 (d, J ) 8.8 Hz, 1H), 6.57 (d, J ) 7.8 Hz, 1H), 6.44 (d,

1922 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Ghirmai et al.
above; combining ꢀ-naltrexamine (100 mg, 0.29 mmol), 4-(tert-
butoxycarbonylamino)benzoic acid (138 mg, 0.58 mmol), BOP (258
mg, 0.58 mmol), and i-Pr2EtN (0.15 mL, 0.88 mmol) for 2 h
followed by basic hydrolysis with K2CO3 (1 g) gave the title
compound as a white foam (154 mg, 94%). mp ) 193.5 °C. Rf )
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(3′-
methoxy)phenylacetamido]morphinan (29). Compound 29 was
synthesized according to the general procedure described above;
combining ꢀ-naltrexamine (100 mg, 0.29 mmol), 3-methoxyphe-
nylacetyl chloride (0.11 mL, 0.29 mmol), and Et3N (0.15 mL, 1.1
mmol) for 2 h followed by basic hydrolysis with K2CO3 (1 g) gave
the title compound as a white solid (82 mg, 57%). mp ) 118.1 °C.
Rf ) 0.38 [10:1 CH2Cl2/MeOH (v/v)]. 1H NMR (CDCl3) δ:
7.29–7.24 (m, 1H), 6.86–6.83 (m, 2H), 6.71 (d, J ) 8.1 Hz, 1H),
6.54 (d, J ) 8.1 Hz, 1H), 6.06 (d, J ) 9.3, 1H), 4.25 (d, J ) 6.9
Hz, 1H), 3.90 (m, 1H), 3.88 (s, 3H), 3.55 (s, 2H). 13C NMR (CDCl3)
δ: 170.8, 159.8, 142.5, 139.6, 136.2, 130.7, 129.9, 124.4, 121.7,
119.0, 117.5, 115.1, 112.6, 93.2, 70.1, 62.2, 59.2, 55.3, 50.8, 47.5,
44.0, 31.0, 29.7, 24.0, 22.7, 9.5, 4.1, 3.9. ESI-MS m/z: 491 (MH+).
HRMS Calcd for C29H34N2O5, 491.2546; found, 491.2531.
1
0.10 [20:1 CH2Cl2/MeOH (v/v)]. H NMR (CDCl3) δ: 7.76 (d, J
) 8.6 Hz, 2H), 7.41 (d, J ) 8.6 Hz, 2H), 7.29 (d, J ) 9 Hz, 1H),
6.89 (s, 1H), 6.72 (d, J ) 8.2 Hz, 1H), 6.55 (d, J ) 8.2 Hz, 1H),
4.53 (d, J ) 6 Hz, 1H), 4.20 (m, 1H). 13C NMR (CDCl3) δ: 166.3,
152.1, 143.1, 141.3, 139.1, 137.2, 130.5, 128.4, 128.1, 124.6, 119.0,
117.6, 92.9, 81.0, 70.2, 62.2, 59.3, 50.2, 47.2, 44.0, 31.8, 29.1, 28.4,
23.4, 22.7, 9.5, 4.2, 3.9. ESI-MS m/z: 562 (MH+). HRMS Calcd
for C32H39N3O6, 562.2917; found, 562.2900.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6-[(3′-
dimethylamino)benzamido]morphinan (25). Compound 25 was
synthesized according to the general procedure described above;
combining ꢀ-naltrexamine (50 mg, 0.15 mmol), 3-(dimethylami-
no)benzoic acid (27 mg, 0.16 mmol), BOP (71 mg, 0.16 mmol),
and i-Pr2EtN (0.09 mL, 0.51 mmol) followed by basic hydrolysis
with K2CO3 provided the desired product as a white solid (57 mg,
80%) after flash chromatography [10:1 CHCl3/MeOH (v/v)]. Rf )
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-(ben-
zylacetamido)morphinan (30). Compound 30 was synthesized
according to the general procedure described above; combining
ꢀ-naltrexamine (100 mg, 0.29 mmol), phenylacetyl chloride (0.98
mL, 0.73 mmol), and Et3N (0.15 mL, 1.1 mmol) for 2 h followed
by basic hydrolysis with K2CO3 (1 g) gave the title compound as
a white solid (48 mg, 36%). mp ) 154.3 °C. Rf ) 0.33 [10:1
1
0.16. ESI-MS m/z 490 (MH+). H NMR (D2O) δ: 7.99 (s, 1H),
1
CH2Cl2/MeOH (v/v)]. H NMR δ: 7.39–7.25 (m, 5H), 6.72 (d, J
7.94 (d, J ) 7.9 Hz, 1H), 7.81 (d, J ) 8.3 Hz, 1H), 7.74 (m, 1H),
6.92 (d, J ) 8.3 Hz, 1H), 6.82 (d, J ) 8.3 Hz, 1H), 4.10 (d, J )
5.7 Hz, 1H), 3.94–2.76 (m, 9H), 2.61 (d, J ) 9.5, 6H), 2.00–1.67
(m, 5H), 1.11 (m, 2H), 0.75 (m, 2H), 0.47 (m, 2H). HRMS Calcd
for C29H36N3O4, 490.2706; found, 490.2694.
) 8.1 Hz, 1H), 6.55 (d, J ) 8.1 Hz, 1H), 6.06 (d, J ) 9.0 Hz, 1H),
4.25 (d, J ) 6.9 Hz, 1H), 3.93–3.86 (m, 1H), 3.59 (s, 2H). 13C
NMR (CDCl3) δ: 171.0, 142.6, 139.6, 134.7, 130.7, 129.4, 128.9,
127.2, 124.3, 119.0, 117.6, 93.2, 70.1, 62.2, 59.2, 50.7, 47.5, 44.0,
43.9, 31.0, 29.6, 23.9, 22.7, 9.4, 4.1, 3.9. ESI-MS m/z: 461 (MH+).
HRMS Calcd for C28H33N2O4, 461.2440; found, 461.2433.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(4′-
methoxy)benzylacetamido]morphinan (31). Compound 31 was
synthesized according to the general procedure described above;
combining ꢀ-naltrexamine (100 mg, 0.29 mmol), 4-methoxyphe-
nylacetyl chloride (0.11 mL, 0.73 mmol), and Et3N (0.15 mL, 1.1
mmol) for 2 h followed by basic hydrolysis with K2CO3 (500 mg)
gave the title compound as a white solid (22 mg, 15%). mp ) 189
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-[(4′-
carboxy)benzamido]morphinan (26). The methyl ester 19 (74 mg,
0.15 mmol) was dissolved in 1:1 THF/H2O (4 mL), and LiOH (62
mg, 1.47 mmol) was added. The solution was stirred at room
temperature for 18 h. The pH was reduced to 5 with 10% HCl,
and the solution was concentrated. The residue was purified by SiO2
chromatography providing the title compound as a white solid (49
mg, 72%). Rf ) 0.28 [2:1 CHCl3/MeOH (v/v)]. mp >300 °C (dec).
1H NMR (CD3OD) δ: 8.00 (d, J ) 8.1 Hz, 2H), 7.82 (d, J ) 8.1
Hz, 2H), 6.65 (d, J ) 8.1 Hz, 1H), 6.59 (d, J ) 8.1 Hz, 1H), 4.65
(d, J ) 7.5 Hz, 1H), 4.60 (m, 1H). ESI-MS m/z: 491 (MH+).
HRMS Calcd for C28H31N2O6, 491.2182; found, 491.2178.
1
°C. Rf ) 0.45 [10:1 CH2Cl2/MeOH (v/v)]. H NMR (CDCl3) δ:
7.17 (d, J ) 9.0 Hz, 2H), 6.88 (d, J ) 9.0 Hz, 2H), 6.71 (d, J )
8.1 Hz, 1H), 6.54 (d, J ) 8.1 Hz, 1H), 6.02 (d, J ) 9.3 Hz, 1H),
4.23 (d, J ) 6.9 Hz, 1H), 3.90 (m, 1H), 3.82 (s, 3H), 3.52 (s, 2H).
13C NMR (CDCl3) δ: 171.3, 158.7, 142.5, 139.6, 130.5, 126.5,
124.5, 119.1, 117.6, 114.4, 93.6, 70.0, 62.2, 59.2, 55.3, 50.7, 47.6,
44.0, 43.1, 30.9, 29.8, 24.2, 22.7, 9.5, 4.1, 3.9. ESI-MS (ESI) m/z:
491 (MH+). HRMS Calcd for C29H34N2O5, 491.2546; found,
491.2547.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-(cin-
namoylacetamido)morphinan (27). Compound 27 was synthesized
according to the general procedure described above; ꢀ-naltrexamine
(100 mg, 0.29 mmol), trans-cinnamoyl chloride (124 mg, 0.73
mmol), and Et3N (0.15 mL, 1.1 mmol) were combined for 2 h
followed by basic hydrolysis with K2CO3 (1 g) gave the title
compound as a white solid (91 mg, 66%). mp ) 248 °C (dec). Rf
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-
[(thiophen-2′-yl)acetamido]morphinan (32). Compound 32 was
synthesized according to the general procedure described above;
combining ꢀ-naltrexamine (100 mg, 0.29 mmol), 2-thiopheneactyl
chloride (0.07 mL, 0.58 mmol), and Et3N (0.12 mL, 0.88 mmol)
for 2 h followed by basic hydrolysis with K2CO3 (1 g) gave the
title compound as a white solid (30 mg, 22%). mp ) 188.8 °C. Rf
) 0.35 [20:1 CH2Cl2/MeOH (v/v)]. 1H NMR (CDCl3) δ: 7.25–7.21
(m, 1H), 6.99–6.92 (m, 1H), 6.70 (d, J ) 7.8 Hz, 1H), 6.53 (d, J
) 8.5 Hz, 1H), 6.40 (d, J ) 8.5 Hz, 1H), 4.30 (d, J ) 6.6 Hz, 1H),
3.93–3.82 (m, 1H), 3.77 (s, 2H). 13C NMR (CDCl3) δ: 169.7, 142.5,
139.6, 135.9, 130.7, 127.4, 127.3, 125.6, 124.5, 119.1, 117.5, 93.2,
70.0, 62.2, 59.2, 50.7, 47.5, 44.0, 37.8, 31.0, 29.7, 24.0, 22.7, 9.5,
4.1, 3.9. ESI-MS m/z: 467 (MH+). HRMS Calcd for C26H30N2O4S,
467.2004; found, 467.1994.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6r-
[(thiophen-2′-yl)acetamido]morphinan (33). Compound 33 was
synthesized according to the general procedure described above;
combining R-naltrexamine (40 mg, 0.18 mmol), 2-thiopheneacetyl
chloride (0.03 mL, 0.23 mmol), and Et3N (0.05 mL, 0.35 mmol)
for 2 h followed by basic hydrolysis with K2CO3 (300 mg) gave
the title compound as a white-yellow solid (30 mg, 55%). mp )
79.8 °C; Rf ) 0.17 [20:1 CH2Cl2/MeOH (v/v)]. 1H NMR δ:
7.29–7.25 (m, 1H), 7.00 (dd, J ) 3.6, 5.1 Hz, 1H), 6.93 (d, J )
3.6 Hz, 1H), 6.68 (d, J ) 8.4 Hz, 1H), 6.52 (d, J ) 8.4 Hz, 1H),
4.61 (d, J ) 3.9 Hz, 1H), 4.60–4.47 (m, 1H), 3.78 (s, 2H). 13C
1
) 0.33 [15:1 CH2Cl2/MeOH (v/v)]. H NMR (CDCl3) δ: 7.60 (d,
J ) 15.6 Hz, 1H), 7.47–7.31 (5H), 6.81 (d, J ) 7.8 Hz, 1H), 6.73
(d, J ) 7.8 Hz, 1H), 6.53 (d, J ) 8.1 Hz, 1H), 6.41 (d, J ) 15.6
Hz, 1H), 4.54 (d, J ) 6.6 Hz, 1H), 4.06 (m, 1H). 13C NMR (CDCl3)
δ: 165.6, 142.7, 140.9, 139.4, 134.7, 130.7, 129.5, 128.6, 127.7,
124.6, 120.9, 119.1, 117.5, 93.0, 70.2, 62.2, 59.3, 50.6, 47.5, 44.0,
31.4, 29.5, 23.7, 22.7, 9.5, 4.1, 3.9. ESI-MS m/z: 473 (MH+).
HRMS Calcd for C29H32N2O4, 473.2440; found, 473.2433.
17-Cyclopropylmethyl-3,14ꢀ-dihydroxy-4,5r-epoxy-6ꢀ-(hy-
droxycinnamoylacetamido)morphinan (28). Compound 28 was
synthesized according to the general procedure described above;
combination of ꢀ-naltrexamine (100 mg, 0.29 mmol), hydrocin-
namoyl chloride (0.11 mL, 0.73 mmol), and Et3N (0.15 mL, 1.1
mmol) for 2 h followed by basic hydrolysis with K2CO3 (1 g)
provided the title compound as a white solid (36 mg, 26%). mp )
120.2 °C. Rf ) 0.36 [10:1 CH2Cl2/MeOH (v/v)]. 1H NMR (CDCl3)
δ: 7.31–7.17 (m, 5H), 6.72 (d, J ) 8.4 Hz, 1H), 6.55 (d, J ) 8.4
Hz, 1H), 6.08 (d, J ) 9.0 Hz, 1H), 4.28 (d, J ) 6.6 Hz, 1H),
3.95–3.85 (m, 1H). 13C NMR (CDCl3) δ: 172.0, 142.7, 140.6, 139.5,
130.7, 128.4, 128.3, 126.1, 124.4, 119.0, 117.7, 93.3, 70.1, 62.2,
59.2, 50.3, 47.4, 44.0, 38.8, 31.8, 31.2, 29.5, 23.9, 22.7, 9.5, 4.1,
3.9. ESI-MS m/z: 475 (MH+). HRMS Calcd for C29H34N2O4,
475.2597; found, 475.2589.

Potential Alcohol-Cessation Agents
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1923
NMR (CDCl3) δ: 169.1, 144.8, 137.0, 136.4, 130.8, 127.2, 127.1,
125.9, 125.4, 119.2, 116.9, 90.1, 69.4, 62.1, 59.6, 47.2, 46.2, 43.2,
37.7, 33.4, 29.0, 22.9, 21.1, 9.4, 4.1, 3.9. ESI-MS m/z: 467 (MH+).
HRMS Calcd for C26H30N2O4S, 467.2004; found, 467.1991.
Receptor Binding and Functional Experiments. Receptor-
binding studies were conducted on human opioid receptors trans-
fected into Chinese hamster ovary (CHO) cells. The µ cell line
was maintained in Ham’s F-12 medium supplemented with 10%
fetal bovine serum (FBS) and 400 µg/mL Geneticin (G418). The
δ and the κ cell lines were maintained in Dulbecco’s minimal
essential medium (DMEM) supplemented with 10% FBS, 400 µg/
mL G418, and 0.1% penicillin/streptomycin. All cell lines were
grown to confluence and then harvested for membrane preparation.
The membranes for functional assays were prepared in buffer A
(20 mM HEPES, 10 mM MgCl2, and 100 mM NaCl at pH 7.4),
and the membranes for binding assays were prepared in 50 mM
Tris buffer (pH 7.7). Cells were scraped from the plates and
centrifuged at 500g for 10 min. The cell pellet was homogenized
in buffer with a polytron, centrifuged at 20000g for 20 min, washed,
recentrifuged, and finally resuspended at 3 mg of protein/mL in
buffer to determine the protein content. The homogenate was then
stored at -70 °C in 1 mL aliquots.
NADP+, 0.5 mM glucose-6-phosphate, 5 IU/mL glucose-6-
phosphate dehydrogenase, 1 mg/mL diethylenetriaminepentaacetic
acid (DETAPAC), and 7 mM MgCl2 for a final incubation volume
of 0.1 mL. Incubations were run for 0, 10, 25, 40, and 60 min in
air with shaking at 37 °C in a water bath and were terminated by
the addition of 1 mL CH2Cl2/2-propanol (3:1, v/v). After centrifu-
gation at 13 000 rpm for 5 min, the organic fraction was collected
and the solvent was removed with a stream of argon. The residue
was reconstituted in methanol (200 µL) and centrifuged at 13 000
rpm for 5 min, and the supernatant was analyzed by HPLC with
an Axxi-chrom (straight-phase) silica column (4.6 × 250 mm, 5
µm) or with a Supelco (reverse-phase) HS F5 pentafluorophenyl
column (4.6 × 250 mm, 5 µm) as described above. Standard
conditions used an isocratic, ternary-solvent system consisting of
solvents A (methanol), B (isopropanol), and C (aqueous 70%
HClO4) set at a flow rate of 1.5 mL/min (straight phase) or A and
D (water) and E (HCO2H) set at a flow rate of 1.0 mL/min (reverse
phase), λ ) 254 nm with retention times (tR) evaluated in minutes.
The specific conditions and retention times of each individual
compound are specified in the Supporting Information.
CYP Inhibition Assays. To measure CYP3A4 activity, test-
osterone 6-hydroxylation, was determined by a HPLC method as
previously described.25 To measure CYP2C9, diclofenac hydroxy-
lase activity was measured by a HPLC method.38 For determination
of CYP2B6, CYP2C19, and CYP2D6 activity, isozyme-specific
Vivid Blue substrate O-dealkylation was determined via a modified
Panvera Vivid Assay Protocol as previously described.25 Briefly,
for CYP2B6, -2C19, and -2D6, microsomes containing 1 pmol of
CYP were added to 0.05 mM Tris buffer (pH 7.4) containing a
NADPH-generating system (i.e., 0.5 mM NADP+, 0.5 mM glucose-
6-phosphate dehydrogenase, 1 mg/mL DETAPAC, and 7 mM
MgCl2) in a total volume of 100 µL. Test compounds (10 µM)
were added, and the substrate (5 µM PanVera Vivid Assay
substrate) was added to initiate the incubation after a brief but
thorough mixing. Incubations were run in a 96-well plate (BD
Falcon Microtest, Black Flat Bottom) for up to 60 min and
monitored continuously to follow the linear portion of the fluores-
cent product versus the time profile using a Wallac Victor2
Multilabel Counter. The inhibition of the amount of product formed
was determined by interpolation from a standard curve and a
comparison of the complete system without inhibitor. The average
percent inhibition ( standard deviation was calculated from three
separate experiments.
Binding assays were conducted using [3H]DAMGO, [3H]C1-
DPDPE, and [3H]U69,593 at the µ, δ, and κ receptors, respectively.
The assay was performed in triplicate in a 96-well plate. Nonspecific
binding was determined with 1.0 µM of the unlabeled counterpart
of each radioligand. Cell membranes were incubated with the
appropriate radioligand and test compound at 25 °C for 60 min.
The incubation was terminated by rapid filtration through glass fiber
filter paper on a Tomtec cell harvester. The filters were dried
overnight and bagged with 10 mL scintillation cocktail before
counting for 2 min on a Wallac Betaplate 1205 liquid scintillation
counter. Full characterization of compounds included analysis of
the data for IC50 values and Hill coefficients using PRISM. Ki values
were calculated using the Cheng Prusoff transformation
IC50
Ki )
1 + L/Kd
where L is the radioligand concentration and Kd is the binding
affinity of the radioligand, as determined previously by saturation
analysis.
[35S]GTPγS Binding for Functional (Agonist/Antagonist)
Determinations. Membranes prepared as described above were
incubated with [35S]GTPγS (50 pM), GDP (usually 10 µM), and
the test compound, in a total volume of 1 mL, for 60 min at 25
°C.21 Samples were filtered over glass fiber filters and counted as
described for the binding assays. A dose-response curve with a
prototypical full agonist (DAMGO, DPDPE, and U69,593 for µ,
δ, and κ receptors, respectively) was conducted in each experiment
to identify full and partial agonist compounds.
In ViWo Studies. Briefly, animals (n ) 5) were trained to
voluntarily self-administer EtOH by the oral route using the
saccharin fadeout method39 and were tested for their response for
10% (w/v) EtOH solution in a two-lever free-choice situation. Once
baseline EtOH intake was achieved, blood alcohol levels (BALs)
were measured to ensure animals were consuming pharmacologi-
cally relevant amounts of EtOH during operant sessions. After
positive verification of BALs and an indication of potency with an
initial screen, a dose-response profile for each compound was
established. To allow for a complete dissipation of any carryover
effects, a 1 week washout period, where rats were rebaselined during
daily 30 min operant sessions, occurred between testing of each
compound.
High-affinity compounds (Ki value is 100 nM or less) that showed
no agonist activity were tested as antagonists. A Schild analysis22
was conducted using a full agonist dose-response curve in the
presence of at least three concentrations of the antagonist. pA2
values and Schild slopes were determined using a statistical program
designed for these experiments. If the Schild slope was significantly
different from -1.00, the antagonist activity was noncompetitive;
in such cases, the pA2 value was not reported. Equilibrium
dissociation constants (Ke values) were calculated as follows:
a
Ke )
Acknowledgment. The authors thank James MacDougll for
some synthetic work and Luke Guo and John Buza for their
help with some of the analytical work (supported by a grant
from the San Diego Foundation and The David Copley
Foundation to J.R.C.). We thank Selena Bartlett and Carsten
Nielsen (Ernest Gallo Clinic and Research Center, Emeryville,
CA) for the in Vitro data of compounds 11 and 12 (Table 3).
This work was financially supported by the National Institute
of Health (NIH) Grant AA16029 awarded to M.R.A.
DR - 1
where a was the nanomolar concentration of the antagonist and
DR was the shift of the agonist concentration–response curve to
the right in the presence of a given concentration of antagonist.
Rat and Mouse Liver Microsome and Human Liver S-9
Stability Assays. A typical assay mixture contained rat or mouse
liver microsomes or human liver S-9 (0.4–0.5 mg of protein), 100
µM potassium phosphate buffer (pH 7.4), 40 µM test compound
oxalate salt, an NADPH-generating system consisting of 0.5 mM

1924 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
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Supporting Information Available: Table listing the HPLC
retention times and the mobile phase and method for analysis. This
material is available free of charge via the Internet at http://
pubs.acs.org.
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