C7β-methyl analogues of the orvinols: The discovery of kappa opioid antagonists with nociceptin/orphanin FQ peptide (NOP) receptor partial agonism and low, or zero, efficacy at Mu opioid receptors

Article abstract of DOI:10.1021/acs.jmedchem.5b00130

Buprenorphine is a successful analgesic and treatment for opioid abuse, with both activities relying on its partial agonist activity at mu opioid receptors. However, there is substantial interest in its activities at the kappa opioid and nociceptin/orphan

Full text of DOI:10.1021/acs.jmedchem.5b00130

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Article
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C7β-Methyl Analogues of the Orvinols: The Discovery of Kappa
Opioid Antagonists with Nociceptin/Orphanin FQ Peptide (NOP)
Receptor Partial Agonism and Low, or Zero, Efficacy at Mu Opioid
Receptors
Juan Pablo Cueva,†,§ Christopher Roche,† Mehrnoosh Ostovar,† Vinod Kumar,†,∥ Mary J. Clark,‡
Todd M. Hillhouse,‡ John W. Lewis,† John R. Traynor,‡ and Stephen M. Husbands*,†
†Department of Pharmacy and Pharmacology, University of Bath, Bath, BA2 7AY, United Kingdom
‡Department of Pharmacology, University of Michigan, Ann Arbor, Michigan 48109, United States
S
* Supporting Information
ABSTRACT: Buprenorphine is a successful analgesic and
treatment for opioid abuse, with both activities relying on its
partial agonist activity at mu opioid receptors. However, there
is substantial interest in its activities at the kappa opioid and
nociceptin/orphanin FQ peptide receptors. This has led to an
interest in developing compounds with a buprenorphine-like
pharmacological profile but with lower efficacy at mu opioid
receptors. The present article describes aryl ring analogues of
buprenorphine in which the standard C20-methyl group has
been moved to the C7β position, resulting in ligands with the
desired profile. In particular, moving the methyl group has
resulted in far more robust kappa opioid antagonist activity
than seen in the standard orvinol series. Of the compounds synthesized, a number, including 15a, have a profile of interest for the
development of drug abuse relapse prevention therapies or antidepressants and others (e.g., 8c), as analgesics with a reduced
side-effect profile.
opioid (DOP) receptor antagonist,6 a lower potency
INTRODUCTION
■
nociceptin/orphanin FQ peptide (NOP) receptor partial
Buprenorphine (1a, Chart 1) is widely used for the treatment
of opioid abuse and as an analgesic. Its mu opioid receptor
agonist,7 and a MOP antagonist.
Encouraging results have also been observed in two small
(MOPr) partial agonist character is of primary importance for
clinical trials using a buprenorphine and naltrexone combina-
both indications, but it is apparent that activity at other
tion therapy in a ratio that should block most, or all, of
receptors, in particular, antagonism at the kappa opioid
buprenorphine’s MOPr agonist activity.8,9 The combination
receptor (KOPr), may play an important role in its clinical
utility.1
proved to be effective in reducing relapse in recovering opiate
addicts and also caused a significant reduction in cocaine use.
The beneficial effect on cocaine use is being more thoroughly
evaluated within The National Drug Abuse Treatment Clinical
Trials Network in the Cocaine Use Reduction with
Buprenorphine (CURB: CTN-0048) study.
Thus, preclinical and clinical data suggest that the
buprenorphine−naltrexone combination has significant ther-
The very high rate of relapse to drug use after a period of
abstinence is a major problem in substance abuse treatment,
with ∼70% of treated addicts relapsing within the first year
following treatment.2 The fact that many drug users use more
than one drug further complicates the situation. A variety of
factors play a role in precipitating relapse, but stress and/or a
priming dose of the drug are particular risks. In preclinical
apeutic potential for the treatment of relapse and polydrug
studies, inhibition or genetic ablation of KOPrs has been shown
addiction, but it has substantial issues that need to be resolved.
to inhibit stress-induced relapse, but this approach has not been
First, delivery of the combination is problematic; buprenor-
phine has poor oral bioavailability and is given sublingually,
whereas naltrexone is active after oral administration but much
less so after sublingual administration. The two compounds are
effective in blocking drug-prime-induced reinstatement to drug-
seeking behavior.3
In contrast to the studies using selective KOPr antagonists,
our own preclinical findings4,5 suggest that a combination of
buprenorphine and naltrexone can be effective in blocking
drug-prime-induced reinstatement to both cocaine- and opioid-
seeking behavior. The combination acts as a KOP and delta
Received: January 23, 2015
© XXXX American Chemical Society
A
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robust KOPr antagonism, enhanced NOPr activity, and low-
efficacy partial agonism, or antagonism, at MOPr. A number of
the ligands can be considered as single compound alternatives
to the buprenorphine/naltrexone combination, while others
offer potential as analgesics with a reduced side-effect profile.
Chart 1
SYNTHESIS
■
In theory, a C7β-methyl group could be introduced into orvinol
type compounds by methylation of the keto-intermediate (2,
Chart 1). In practice, this leads to a number of rearrangement
products resulting from the instability of the C7-anion.16,17 The
alternative approach is to introduce the methyl group during
the Diels−Alder reaction. Methacrylonitrile is known to
undergo the Diels−Alder reaction with thebaine and N-
cyclopropylnorthebaine, but it gives the C7α-methyl adduct
(3, Chart 1), the opposite to that desired and a result of the
methyl group being bulkier than the nitrile.18 Methacrolein
should give a more favorable distribution of product due to the
increased bulk of the aldehyde, but it has been reported that
methacrolein does not undergo Diels−Alder reaction with
thebaine.19 We have previously shown that lithium tetrafluor-
oborate catalyzes Diels−Alder reactions between N-cyclo-
propylcarbonylnorthebaine and cycloalkenones20 and have
now applied this method to the reaction with methacrolein
(Scheme 1). The reaction was successful, giving a 1.4:1 mixture
of the C7α-methyl (4a) and C7β-methyl (4b) adducts that
could be separated by silica gel chromatography. Addition of
phenyl Grignard, in the presence of tetrabutylammonium
bromide, to 4b gave the secondary alcohol 5a, with opposite
stereochemistry at C20 to that desired. This bias toward the S-
isomer (5a) parallels our findings on Grignard addition to the
aldehyde of the related C7β-H series14 and is opposite to that
observed on addition of Grignard reagent to the methyl ketone
in the C7β-H series.14,15 Treatment of 5a with LiAlH4 to
reduce the amide before 3-O-demethylation gave 10a. To
obtain the desired diastereoisomer, 5a was oxidized to the
phenyl ketone (6a) before LiAlH4 reduction of both the
cyclopropylcarbonyl group and the ketone, the latter occurring
stereoselectively, to give 7a. 3-O-Demethylation yielded 8a.
The analogues (8b−8e; 10b−10e) were prepared in an
identical manner. The 4-fluoro analogues (7e, 9e) were not
fully stable to propanethiolate-mediated 3-O-demethylation,
which, in the case of 7e, resulted in the formation of isolable
amounts of 8f where the fluoro substituent was substituted by
propanethiol.
therefore not amenable to formulation as a single tablet, and it
is not desirable to use the two drugs separately due to potential
compliance issues and the possibility of buprenorphine being
diverted for illicit, nonmedical use. Second, the exact
pharmacological profile provided by the combination is not
clear, particularly whether there is a “pure” antagonist or a low-
efficacy MOPr component. Provision of single chemical entities
that provide buprenorphine-like activity at KOP, DOP, and
NOP receptors but with substantially reduced efficacy at MOPr
will allow these issues to be resolved and will provide
compounds with potential therapeutic utility for relapse
prevention and poly drug abuse.
Buprenorphine has also found widespread use as an analgesic
and appears to be effective in neuropathic pain conditions that
are not sensitive to other opioids, although the mechanism for
this difference is not clear.10 There has been speculation that
buprenorphine’s unique clinical profile might be linked to its
ability to act as a low-efficacy partial agonist at NOPr in
addition to its MOPr partial agonism,11 although Cremeans et
al. found no evidence for activation of NOPr being important
for buprenorphine’s analgesic activity in primates.12 On the
other hand, they did find that coactivation of MOPr and NOPr
produced synergistic antinociception. This suggests that
bifunctional ligands with low efficacy at MOPr and moderate
efficacy at NOPr might have utility as analgesics with a reduced
side-effect profile. Efforts to develop such bifunctional ligands
are being made.13
The etheno-bridged aldehyde 4b was also hydrogenated to
11, and the same sequence of steps was carried out to provide
the ethano-bridged analogues 15 and 17 (Scheme 2).
RESULTS
■
In order to streamline the development process and avoid
unnecessary full evaluation of ligands of limited interest to this
project, the compounds were screened in a [35S]GTPγS assay
for MOPr, KOPr, and NOPr efficacy at a very high
concentration (10 μM) to determine peak efficacy at each
receptor (Table 1). Potencies as agonists (EC50) or antagonists
(Ke) were then determined for the most interesting compounds
(chosen based on efficacies at the receptors and their fit with
the target profiles) and reported in Table 1. A range of
compounds were also evaluated for affinity at MOP, DOP, and
KOP receptors by measuring displacement of [3H]-
diprenorphine binding from membranes of C6-rat glioma
Orvinols, the series of compounds to which buprenorphine
belongs, typically display high efficacy at one, or both, of the
MOPr and KOPr.14 However, we recently reported on a series
of aryl analogues of buprenorphine, some of which displayed
substantially lower efficacy at the opioid receptors while
retaining high affinity for opioid receptors and moderate
(equivalent to buprenorphine) affinity for NOPr.15 We now
report on a series of orvinol analogues in which the C20-methyl
group has been moved to the C7-β position, resulting in more
B
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a
Scheme 1
a
(i) Methacrolein, LiBF4; (ii) ArMgBr, Bu4NBr, THF, reflux; (iii) LiAlH4, THF; (iv) PrSNa, HMPA; (v) DMSO, oxalyl chloride, CH2Cl2.
cells expressing recombinant rat MOPr and DOPr and CHO
cells expressing recombinant human KOPr (Table 2). NOPr
binding affinity was measured by displacement of [3H]-
nociceptin from membranes of HEK cells expressing
recombinant NOPr. Details of these assays have been described
previously.6,21−24
very low, or no, efficacy at KOPr. A number (15a−e) were
evaluated as antagonists at this receptor and were found to be
as potent as buprenorphine (all having Ke < 1 nM). As with
series 8, they were also antagonists (e.g., 15a) or low-efficacy
partial agonists at MOPr (≤17% of DAMGO). While SAR at
this receptor was not absolutely clear cut, 4′-substituents again
gave rise to higher efficacy than 3′-substituents (e.g., 15e vs
15h, 15d vs 15c). This series was extended to include
heterocyclic analogues 15i−15m. Those having a thiophene
ring (15i−15l) had partial agonist activity at KOPr (10−28%)
and at MOPr (10−61%), whereas 15m, having a 3-furanyl
group, was devoid of efficacy at KOPr and MOPr. Only 15m
had appreciable efficacy at NOPr within the series of
heterocyclic analogues. 17, the C20-diastereoisomer of 15a,
was a full agonist at KOPr (97% stim) and a partial agonist at
MOPr, with little efficacy at NOPr.
Binding affinities were determined for a range of compounds,
including the more interesting analogues from series 8 and 15.
As would be expected, all had high affinity for MOPr and KOPr
(and DOPr when tested) with subnanomolar Ki values. Of
particular note was the range of affinities at NOPr, where a
number of the compounds had substantially higher affinity than
that of buprenorphine (e.g 8e, 15d, 15e, 15f, 15k), whereas
All ligands from series 8 are KOPr antagonists with maximal
percent stimulation <10%. Efficacy at MOPr ranges from zero
(8a) to moderate efficacy partial agonism (8d, 8f: 35 and 49%
of DAMGO, respectively) with clear SAR relating to the
location of the phenyl ring substituent (4′ > 3′ > 2′). At NOPr,
the compounds were partial agonists somewhat similar in
efficacy to buprenorphine, with the highest efficacy demon-
strated by the unsubstituted (8a; 56%), 3′-methyl substituted
(8c; 43%), and 4′fluoro (8e, 51%) analogues. Potency at NOPr
was higher than seen with buprenorphine. Series 10, the
diastereomers of 8, were consistently of higher efficacy at the
KOPr with only the 4′-Me (10d) substituted analogue profiling
as a KOPr antagonist. The others in the series were moderate
to high efficacy KOPr agonists (45−92% of U69,593). At
MOPr, they were all of moderate efficacy (33−55%) and very
low efficacy at NOPr.
The phenyl and substituted phenyl, ethano-bridge analogues
(15a−h) had very similar SAR to series 8 in that they also had
C
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Article
a
Scheme 2
a
(i) H2, Pd/C, EtOH; (ii) ArMgBr, Bu4NBr, THF, reflux; (iii) LiAlH4, THF; (iv) PrSNa, HMPA; (v) DMSO, oxalyl chloride, CH2Cl2.
others had roughly equivalent affinity to that of buprenorphine
(e.g., 8a, 8c, 15a, 15b, 15c).
of efficacy at KOPr being attributed to the significant steric bulk
adjacent to C20.14 Initially, it was believed that buprenorphine
was also unusual in having measurable affinity for NOPr,28 an
activity that has been postulated to explain some of
buprenorphine’s in vivo pharmacology.29,30 However, in recent
years, it has become apparent that some other members of the
orvinol family also interact with NOPr, particularly those with a
bulky group attached to C20 (the t-butyl group of
buprenorphine fulfils this role).31−33 While it has been possible
to introduce good affinity for NOPr, it has proven to be difficult
to couple this with moderate efficacy for this receptor without
also introducing efficacy for classical opioid receptors, in
particular KOPr. The orvinol TH-030418 (18; Chart 1) is
reported as having affinity for NOPr virtually equivalent to its
affinity for MOPr, KOPr, and DOPr; however, it also displays
very potent and efficacious antinociceptive activity in vivo. This
activity is blocked by the opioid antagonist naloxone, indicating
high efficacy at MOPr and/or KOPr.33 Even more closely
related to the current series are analogues of 1a having
alternative bulky groups at C20.31 In that series, moderate
affinity and efficacy for NOPr was always associated with partial
agonism at MOPr and/or KOPr. The robust KOPr antagonist
activity of series 8 and 15, especially when coupled with
In order to help explain the unusually low efficacy at KOPr of
this series, the docking of 15a to the crystal structure of the
KOPr was examined. The crystal structure of the KOPr was
determined in the presence of the selective KOPr antagonist
JDTic and presumably represents the inactive conformation of
the receptor and, importantly, the conformation favored when
binding 15a.25 15a sits in the hydrophobic binding pocket with
the expected interaction between the protonated nitrogen and
the side chain of Asp138, while the C7β-methyl projects toward
Tyr139 in TM3 (Figure 1).
DISCUSSION
■
Members of the orvinol series of compounds are mostly known
for their nonselective binding to MOPr, KOPr, and DOPr and
potent agonist activity at one or more of these receptors.26
Even those with a cyclopropylmethyl (CPM) group attached to
the basic nitrogen, which has the effect of reducing efficacy
particularly at MOPr,27 tend to be efficacious and potent
agonists, primarily at KOPr.14 Buprenorphine, being a KOPr
antagonist, is a very distinct member of the series, with the lack
D
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Table 1. Maximal Stimulation of [35S]GTPγS Binding of the New C7β-Methyl Orvinol Analogues to Opioid and NOP
Receptors
a
b
[35S]GTPγS, % stimulation, and, in brackets, agonist EC50/nM or antagonist Ke/nM
MOPr
KOPr
NOPr
39 12% (EC50: 1480 980 nM)
14
DOPr
3%
1a
1b
8a
8b
8c
8d
8e
8f
20 6% (EC50: 0.7 0.3nM)
0
6% (Ke = 0.14 0.06 nM)
7
7
6.0
4
1
19
3
4
4
6%
7%
56 2.5% (EC50: 416 74 nM)
17 1%
3%
12 5%
−4 1%
2%
−6 1%
22 5% (EC50: 1.4 0.4 nM)
35 11% (EC50: 0.24 0.04 nM)
26 8% (EC50: 0.22 0.06 nM)
49 2%
6
54 11% (EC50: 14 5 nM)
14 2%
15 3%
9
1%
51 20% (EC50: 7.3 3 nM)
20 4%
4
10%
10a
10b
10c
10d
10e
33 6%
48 1%
40 13%
49 3%
55 7%
62 4%
72 4%
92 4%
5
6%
12 2%
11 3%
8
5%
4
7
5%
3%
45 6%
15a
15b
15c
15d
15e
15f
2
4% (Ke = 0.28 0.04 nM)
15 7%
1%
−2 1% (Ke = 0.09 0.04 nM)
−4 4% (Ke = 0.13 0.05 nM)
−1 1% (Ke = 0.12 0.08 nM)
−5 4% (Ke = 0.10 0.05nM)
−5 3% (Ke = 0.10 0.05 nM)
56 1% (EC50: 147 33 nM)
15 4%
0
6
4%
3%
7
61 15% (EC50: 331 223 nM)
14 4%
17 2%
15 1%
4
3%
13 5%
1%
2
1%
3
15g
15h
15i
7
2
1%
1%
7
4%
42 3% (EC50: 230 37 nM)
15 4%
28 + 6
31 2% (EC50: 160 70 nM)
10 + 1
2 + 1
15j
31 + 4
26 + 2
15 + 1
16 3%
15k
15l
61 2%
10 4%
10 2%
20 2%
6
3%
15m
1
1%
2
4%
29 3% (EC50: 230 70 nM)
17
33 7%
97 5%
10 4%
a
Percent maximal stimulation (% stim) at a single high concentration (10 μM) with respect to the standard agonists DAMGO (MOPr) and U69,593
b
(KOPr) and nociceptin (NOPr); values are an average SEM from three separate experiments. In brackets for selected compounds, agonist EC50/
nM or antagonist Ke (the antagonist dissociation constant determined against the standard agonists listed above) are given.
improved affinity for NOPr, is therefore unprecedented within
the orvinols and close analogues. That they are 2° alcohols,
which typically have very much higher efficacy than their 3°
analogues,14 confirms the very substantial effect of moving the
methyl group from C20 to the C7β-position. A comparison
with the recently reported 1b and aryl substituted analogues
confirms the importance of this methyl group. 1b and
analogues differ from 15 only in the location of this single
methyl, yet in the 1b series, only the parent compound (R =
Ph) and the 3′-chloro analogue (1c: R = 3-ClPh) had low
efficacy at KOPr (19 and 26%, respectively), with other
substituents (2′-, 3′-, and 4′-methyl, 3′- and 4′-F) all resulting
in efficacy in the range 77−102% at this receptor.15
Docking of 15a to the published crystal structure of the
KOPr25 allows a hypothesis to be formulated to explain the
very low efficacy of this series to the KOPr. In the model of the
KOPr active and inactive states,23 the methyl group of 15a
projects toward Tyr139 in TM3. In the inactive conformation,
Tyr139 and the whole of TM3 move away from this position,
whereas in the active conformation, they are much closer,
possibly too close to allow the active conformation to be taken
up; i.e., through an interaction between the C7β-methyl group
and Tyr139, 15a and analogues may disfavor the active state of
the receptor.
Distinct SAR for affinity and efficacy at NOPr is found in
series 8 and 15. In both cases, the unsubstituted parent or a 3′-
substituent gives highest efficacy at NOPr, whereas highest
affinity is associated with a 4′-substituent, although this latter
effect is not large.
CONCLUSIONS
■
Moving the methyl group from C20 to the C7β position has
resulted in one of the most striking pieces of SAR in the orvinol
series. In particular, the robust reduction in efficacy at KOPr
coupled with a retention, or increase, in NOPr affinity and
efficacy has resulted in a series of compounds worthy of
consideration as (i) relapse prevention agents and (ii) low
abuse liability analgesics. 15a is currently undergoing in vivo
evaluation as part of its preclinical development.
EXPERIMENTAL SECTION
■
General Procedures. Reagents and solvents were purchased from
Sigma-Aldrich or Alfa Aesar and used as received. Buprenorphine (1a)
was supplied by the National Institute on Drug Abuse, Bethesda,
Maryland. 1H and 13C NMR spectra were obtained with a Bruker 400
MHz instrument (1H at 400 MHz, 13C at 100 MHz); δ is given in
ppm, J, in Hz, with TMS as an internal standard. ESIMS: microTOF
(BRUKER), EIMS: Fisons Autosampler. Microanalysis: PerkinElmer
E
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induce crystallization. The crystals were collected by filtration and
dried under vacuum.
Table 2. Binding Affinities (Ki/nM) of of the New C7β-
Methyl Orvinol Analogues to Opioid and NOP Receptors
Swern Oxidation/LiAlH4 Reduction of Secondary Alcohols
(General Procedure B). A solution of oxalyl chloride (1.25 equiv) in
CH2Cl2 (3 mL/mmol) was cooled to −78 °C in a one-necked flask.
Into this flask was added dropwise a solution of dry DMSO (2.6 equiv)
in CH2Cl2 (3 mL/mmol). The solution stirred for 5 min, and then a
solution of thevinol 5 or 12 in CH2Cl2 (2 mL/mmol) was added. The
mixture stirred for 20 min, and then Et3N (5 equiv) was added. The
reaction was removed from the cold bath and stirred for 1 h, and water
was added. The mixture was shaken, and the organic layer was
separated and washed with a saturated solution of NH4Cl and then
with a concentrated solution of NaHCO3. The solution was washed
once more with brine, dried over magnesium sulfate, and filtered, and
the solvents were removed under reduced pressure to yield crude 6 or
13 as a clear solid.
This residue was dissolved in dry THF (10 mL/mmol) and added
to a stirring suspension of LiAlH4 (4 equiv) in dry THF (5 mL/mmol)
at 0 °C. The suspension was allowed to warm to RT and was stirred
for 24 h. The reaction was cooled to 0 °C and quenched with water in
THF. The mixture was filtered, rinsing the solids with hot THF. The
solution was subjected to rotary evaporation to yield an oil that was
subjected to silica gel column chromatography, eluting with 15%
EtOAc in petroleum ether to yield two constituents, 7 or 14 (as the
major product), as a high Rf component, and 9 or 16 (as the minor
product), with lower Rf.
O-Demethylation Using NaSPr/HMPA (General Procedure
C). A solution of thevinol 7, 9, 14, or 16 in dry HMPA (6 mL/mmol)
was added sodium propanethiolate (6 equiv). The reaction was stirred
for 3 h at 115 °C and then cooled to RT and quenched with 7 mL/
mmol of a concentrated solution of NH4Cl. The mixture was extracted
three times with Et2O. The organic layer was then extracted five times
with water and once with brine, dried over MgSO4, and filtered, and
the solvents were removed under reduced pressure. The residue was
then subjected to silica gel flash column chromatography, eluting with
a gradient of EtOAc in petroleum ether. The fractions containing the
compound of interest were then evaporated to dryness, dissolved in a
2 M solution of HCl in EtOH, and then induced to crystallize upon
addition of EtOAc. The crystals were collected by filtration and dried
under vacuum.
a
binding/nM
MOPr
KOPr
NOPr
DOPr
1a
0.13 0.02
0.17 0.05
0.08 0.02
NT
0.089 0.02
0.044 0.015
0.08 0.03
NT
212
7
0.48 0.26
NT
1b
43.2 13.4
97 12
8a
0.48 0.05
NT
8b
1270 170
8c
0.17 0.11
0.16 0.12
0.10 0.02
0.20 0.07
0.098 0.022
0.14 0.06
0.071 0.014
NT
0.04 0.01
0.05 0.01
0.04 0.01
0.12 0.03
0.11 0.04
0.16 0.05
0.10 0.03
NT
79
34
8
6
0.40 0.16
NT
8e
15a
15b
15c
15d
15e
15f
15g
15h
15k
15l
15m
80 10
820 60
240 50
0.25 0.18
NT
NT
56
56
26
5
3
6
NT
0.47 0.38
NT
0.052 0.007
0.09 0.03
NT
0.094 0.04
0.13 0.05
NT
47.8 17
62.2 31
44 11
NT
NT
NT
NT
NT
571 77
1455 469
NT
0.11 0.011
0.14 0.10
NT
a
Ki (nM) versus [3H]diprenorphine (for MOPr, KOPr, and DOPr)
and [3H]nociceptin (for NOPr); values are an average SEM from
three separate experiments. NT: not tested
240C analyzer. Column chromatography was performed using
RediSep prepacked columns with a Teledyne Isco CombiFlash
instrument. Most ligands were tested as their hydrochloride salts,
prepared by adding 5 equiv of HCl (1 N solution in diethyl ether) to a
solution of compound in anhydrous methanol. Alternatively, the
oxalate salt was formed by adding 1 equiv of oxalic acid in EtOH to the
ligand in EtOH. All reactions were carried out under an inert
atmosphere of nitrogen unless otherwise indicated. All compounds
were >95% pure, as determined by microanalysis. A representative
synthesis is reported here.
Arylmagnesium Halide Addition (General Procedure A). To
a solution of aldehyde 4b or 11 in dry THF (10 mL/mmol of
aldehyde) were added 3 equiv of Bu4NBr followed by 2 equiv of
arylmagnesium halide as solution in THF. The solution was then
heated at reflux for 48 h, cooled to RT, and quenched with 0.05 mL of
water. The mixture was allowed to stir for 5 min and then filtered over
Celite. The solids were washed with hot THF, and the solution was
removed of its solvent by rotary evaporation. The remaining residue
was partitioned between EtOAc (20 mL) and water (10 mL). The
aqueous layer was extracted twice with 5 mL of EtOAc. The pooled
organic solvent was washed twice with water (5 mL) and once with
brine, dried over MgSO4, filtered, and evaporated under reduced
pressure. The residue was dissolved in a minimum amount of Et2O to
N-Cyclopropylcarbonyl-7α-formyl-7β-methyl-6,14-endoe-
thenotetrahydronorthebaine (4b). To a suspension of 13.61 g
(37.29 mmol) N-CPCnorthebaine in 20 mL of methacrolein was
added 3.49 g of LiBF4. The resulting solution was stirred for 16 h at
RT. Into this solution was added 30 mL of CH2Cl2, and the mixture
was extracted with water (10 mL × 3) and brine (5 mL). The solution
was dried, filtered, and removed of solvent on a rotary evaporator to
afford a dark red syrup. This material was subjected to silica gel flash
column chromatography, eluting with 50% EtOAc in petroleum ether
to afford 5.91 g of the faster running component 4a (N-cyclo-
propylcarbonyl-7β-formyl-7α-methyl-6,14-endo-ethenotetrahydronor-
1
thebaine) as a white solid. H NMR (400 MHz, CDCl3) δ 9.85 (s,
1H), 6.67 (d, 1H, J = 8.0 Hz), 6.57 (d, 1H, J = 8.0 Hz), 6.14−6.07 (2d,
Figure 1. Proposed binding modes for 15a (purple) in the models of inactive (A) and active (B) conformations of human KOPr. The C7β-methyl-
group of 15a negatively interacts with Tyr139 in the active receptor conformation but not in the inactive one.
F
DOI: 10.1021/acs.jmedchem.5b00130
J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
1
1H), 5.57 (d, 1H, J = 12.0 Hz), 5.33 (d, 1H, J = 8.0 Hz), 4.76 (d, 1H,
0.4H), 4.57−4.53 (m, 1.6H), 4.15−4.10 (m, 1H), 3.85 (s, 3H), 3.72 (s,
3H), 3.42 (dt, 1H, Ja = 12.0 Hz, Jb = 4.0 Hz), 3.11−3.05 (dd, 1H, Ja =
8.0 Hz, Jb = 4.0 Hz), 2.88−2.83 (m, 1H), 2.43−2.35 (dt, 1H, Ja = 12.0
Hz, Jb = 8.0 Hz), 1.85−1.76 (m, 1H), 1.69−1.65 (m, 1H), 1.09−1.03
(m, 5H), 0.91−0.76 (m, 2H). ESIMS: m/z 436 (M + H+, 100). In
addition, 4.11 g of a slower running component, N-cyclopropylcar-
bonyl-7α-formyl-7β-methyl-6,14-endoethenotetrahydronorthebaine
solid. H NMR (400 MHz, CDCl3) δ 7.49−7.42 (m, 2H), 7.35−7.28
(m, 2H), 7.28−7.20 (m, 1H), 6.61 (d, 1H, J = 8.0 Hz), 6.47 (d, 1H, J =
8.0 Hz), 5.98 (s, 1H), 5.05 (s, 1H), 4.95 (d, 1H, J = 1.6 Hz), 3.59 (s,
3H), 2.98 (d, 1H, J = 6.4 Hz), 2.93 (d, 1H, J = 18.3 Hz), 2.63−2.50
(m, 1H), 2.32−2.13 (m, 5H), 1.95 (dd, 1H, Ja = 14.2 Hz, Jb = 3.9 Hz),
1.89−1.71 (m, 2H), 1.59−1.44 (m, 2H), 1.35−1.21 (m, 4H), 0.97−
0.81 (m, 1H,), 0.75−0.60 (m, 1H), 0.52−0.36 (m, 2H), 0.10−0.03 (m,
2H). ESIMS: m/z 488 (M + H+, 100).
1
Molecular Modeling. We used the crystal structure of the human
KOR (PDB ID: 4djh) together with our previous active state
models.23 Compound 15a was built, and the low-energy conformation
was then subjected to automated rigid docking implemented in
QUANTA (Accelrys Inc.).
(4b), was afforded as a white solid. H NMR (400 MHz, CDCl3) δ
9.54 (s, 0.5H), 9.45 (s, 0.5H), 6.69 (d, 1H, J = 8.0 Hz), 6.59 (d, 1H, J
= 8.0 Hz), 6.14 (t, 1H), 5.57 (dd, 1H, Ja = 12.0 Hz, Jb = 8.0 Hz), 5.35
(d, 0.5H, J = 4.0 Hz), 4.93 (s, 1H), 4.80 (d, 0.5H, J = 8.0 Hz), 4.64
(dd, 1H, Ja = 8.0 Hz, Jb = 4.0 Hz), 4.15 (dd, 1H, Ja = 8.0 Hz, Jb = 4.0
Hz), 3.85 (s, 3H), 3.72 (2s, 3H), 3.49 (dt, 1H), 3.31−3.21 (m, 2H),
2.37−2.26 (dt, 0.5H), 2.27−2.15 (dt, 0.5H), 2.06−1.72 (m, 3H), 1.35
ASSOCIATED CONTENT
■
1
(s, 3H), 1.08 (m, 2H), 0.82 (m, 2H). At RT, the H NMR spectra of
S
* Supporting Information
this compound in DMSO-d6 has two signals at δ 9.408 (s, 0.5H) and δ
1
Full characterization of final compounds; microanalysis and
HPLC; pharmacological methods. The Supporting Information
is available free of charge on the ACS Publications website at
DOI: 10.1021/acs.jmedchem.5b00130.
9.375 (s, 0.5H) that coalesce when running the H NMR experiment
at 360 K. ESIMS: m/z 436 (M + H+, 100).
N-Cyclopropylcarbonyl-7α-formyl-7β-methyl-6,14-endoe-
thanotetrahydronorthebaine (11). Aldehyde 4b (500 mg) was
dissolved in 15 mL of EtOH. Into this solution was added 30 mg of
10% Pd on carbon. The mixture was shaken in a Parr hydrogenator
under 100 psi of H2 for 12 h. The mixture was filtered, and the
solvents were removed under reduced pressure to yield 510 mg of 11
AUTHOR INFORMATION
■
Corresponding Author
*Phone: 44 1225 383103. Fax: 44 1225 386114. E-mail: s.m.
husbands@bath.ac.uk.
1
as a white solid. H NMR (400 MHz, CDCl3) δ 9.70 (s, 0.6H), 9.69
(s, 0.4H), 6.76 (d, 1H, J = 8.1 Hz), 6.61 (d, 1H, J = 8.1 Hz), 4.9 (d,
0.4H, J = 6.9 Hz), 4.82 (s, 0.6H), 4.81 (s, 0.4H), 4.52 (dd, 0.6H, Ja =
14 Hz, Jb = 5.7 Hz), 4.34 (d, 0.6H, J = 6.8 Hz), 4.04 (dd, 0.4H, Ja = 15,
Jb = 6 Hz), 3.89 (s, 3H), 3.48 (s, 1.8H), 3.46 (s, 1.2H), 3.34 (td, 0.4H,
Ja = 13.4, Jb = 3.2 Hz), 3.05 (dd, 0.6H, Ja = 18.5 Hz, Jb = 6.9 Hz), 2.97
(dd, 0.4H, Ja = 18.7 Hz, Jb = 7.2 Hz), 2.87 (d, 0.6H, J = 18.7 Hz), 2.81
(td, 0.6H, Ja = 13.7, Jb = 4.1 Hz), 2.71 (d, 0.4H, J = 18.8 Hz), 2.42−
2.37 (m, 1H), 2.27 (td, 0.4H, Ja = 13.1 Hz, Jb = 5.7 Hz), 2.16 (td, 0.6H,
Ja = 13.2 Hz, Jb = 5.8 Hz), 1.80−1.60 (m, 3.6H), 1.5 (dd, 0.4H, Ja =
13.7 Hz, Jb = 3.7 Hz), 1.33−1.18 (m, 5H), 1.15−1.06 (m, 1H), 1.03−
0.94 (m, 2H), 0.84−0.71 (m, 2H). ESIMS: m/z 438 (M + H+, 100).
(1′S,5α,6R,7R,14α)-1′-Phenyl-1′-(4,5-epoxy-7,8-dihydro-3,6-
dimethoxy-7β-methyl 17-cyclopropylcarbonyl-6,14-ethano-
morphinan-7-yl)-methan-1′-ol (12a). General procedure A was
followed using 515 mg of 11 to yield 336 mg of 12a as white crystals.
1H NMR (400 MHz, CDCl3) δ 7.36−7.23 (m, 5H), 6.78−6.75 (2d,
1H), 6.61−6.58 (2d, 1H), 5.06 (d, 0.55H, J = 2.6 Hz), 4.99 (d, 0.45H,
J = 2.6 Hz), 4.91−4.87 (m, 1H), 4.51−4.46 (dd, 0.55H, Ja = 5.4 Hz, Jb
= 13.7 Hz), 4.35−4.34 (d, 0.55H, J = 6.8 Hz), 4.03−3.98 (d, 0.45H, Ja
= 4.9 Hz, Jb = 13.5 Hz), 3.92−3.914 (2s, 3H), 3.53 (s, 1.65H), 3.48 (s,
1.35H), 3.38−3.30 (dt, 0.55H, Ja = 3.8 Hz, Jb = 8.6 Hz), 3.11−3.04
(dd, 0.55H, Ja = 6.9 Hz, Jb = 18.3 Hz), 3.02−2.96 (dd, 0.45H, Ja = 6.9
Hz, Jb = 18.5 Hz), 2.92−2.88 (d, 0.55H, J = 18.3 Hz), 2.49−2.44 (m,
1H), 2.36−2.29 (dt, 0.45 Hz, Ja = 5.6 Hz, Jb = 12.9 Hz), 2.25−2.17 (dt,
0.55 Hz, Ja = 6.0 Hz, Jb = 13.2 Hz), 2.03−1.90 (m, 2H), 1.86−1.80 (m,
0.45 Hz), 1.76−1.51 (m, 6H), 1.10−0.69 (m, 7H). ESIMS: m/z 516
(M + H+, 100).
(1′R,5α,6R,7R,14α)-1′-Phenyl-1′-(4,5-epoxy-7,8-dihydro-3,6-
dimethoxy-7β-methyl 17-cyclopropylmethyl-6,14-ethano-
morphinan-7-yl)-methan-1′-ol (14a). General procedure B was
followed using 473 mg of 12a to yield a clear residue after extraction.
Crystallization from CH2Cl2/Et2O gave 133 mg of 14a as white
crystals. 1H NMR (400 MHz, CDCl3) δ 7.45 (m, 2H), 7.34−7.30 (m,
2H), 7.27−7.24 (m, 1H), 6.73 (d, 1H, J = 8.4 Hz), 6.57 (d, 1H, J = 8.0
Hz), 5.76 (s, 1H), 5.03 (s, 1H), 4.96 (s, 1H), 3.91 (s, 3H), 3.64 (s,
3H), 3.01−2.94 (m, 2H), 2.57 (d, 1H, J = 6.8 Hz), 2.29 (dd, 1H, Ja =
6.8 Hz, Jb = 18.3 Hz), 2.24−2.20 (m, 3H), 1.95 (dd, 1H, Ja = 4 Hz, Jb =
14.3 Hz), 1.90−1.77 (m, 2H), 1.60−1.52 (m, 2H), 1.40−1.28 (m,
4H), 0.95 (m, 1H), 0.68 (m, 1H), 0.51−0.39 (m, 2H), 0.08−0.01 (m,
2H). ESIMS: m/z 502 (M + H+, 100).
Present Addresses
§(J.P.C.) Neurocrine Biosciences, Inc., San Diego, California
92130, United States.
∥(V.K.) Centre for Chemical and Pharmaceutical Sciences,
Central University of Punjab, Bathinda, 151001, India.
Notes
The authors declare the following competing financial
interest(s): Subsequent to preparation of the manuscript the
compounds disclosed have been licensed to a biopharmaceut-
ical company.
ACKNOWLEDGMENTS
■
This work was funded by the National Institutes of Health,
National Institute on Drug Abuse grant DA07315 (to S.M.H.).
T.M.H. was supported by T32 DA07268. Irina Pogozheva
(University of Michigan) is thanked for docking studies
between 15a and the KOPr.
ABBREVIATIONS USED
■
MOP receptor, mu opioid receptor; DOP receptor, delta opioid
receptor; KOP receptor, kappa opioid receptor; NOP receptor,
nociceptin opioid peptide receptor; EKC, ethylketocyclazocine;
N/OFQ, nociceptin/orphanin FQ
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