Syntheses and opioid receptor binding properties of carboxamido-substituted opioids

Article abstract of DOI:10.1016/j.bmcl.2008.10.134

A series of 15 novel opioid derivatives were made where the prototypic phenolic-OH group of traditional opioids was replaced by a carboxamido (CONH₂) group. For 2,6-methano-3-benzazocines and morphinans similar or, in a few instances, enhanced

Full text of DOI:10.1016/j.bmcl.2008.10.134

Bioorganic & Medicinal Chemistry Letters 19 (2009) 203–208
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Syntheses and opioid receptor binding properties
of carboxamido-substituted opioids q
a
a
a
a
Mark P. Wentland a, , Rongliang Lou , Qun Lu , Yigong Bu , Melissa A. VanAlstine ,
*
Dana J. Cohen b, Jean M. Bidlack b
a Department of Chemistry and Chemical Biology, Rensselaer Polytechnic Institute, Troy, NY 12180, USA
b Department of Pharmacology and Physiology, School of Medicine and Dentistry, University of Rochester, Rochester, NY 14642, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of 15 novel opioid derivatives were made where the prototypic phenolic-OH group of traditional
opioids was replaced by a carboxamido (CONH2) group. For 2,6-methano-3-benzazocines and morphin-
Received 22 September 2008
Revised 22 October 2008
Accepted 27 October 2008
Available online 7 November 2008
ans similar or, in a few instances, enhanced affinity for
l, d and j opioid receptors was observed when the
OH ? CONH2 switch was applied. For 4,5 -epoxymorphinans, binding affinities for the corresponding
a
carboxamide derivatives were much lower than the OH partner consistent with our pharmacophore
hypothesis concerning carboxamide bioactive conformation. The active metabolite of tramadol and its
carboxamide counterpart had comparable affinities for the three receptors.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
Opioids
SAR
In 2001 we reported our observation that the prototypic pheno-
lic hydroxyl group of certain opioids can be replaced by a carbox-
amide group and retain high affinity binding to opioid receptors.1
When this OH ? CONH2 switch was applied to the cyclazocine
core structure, for example, binding affinities for cyclazocine
(1a)2 and 8-carboxamidocyclazocine (8-CAC, 1b) were comparable
there is no barrier created by H-bonding to a neighboring ether
bridge.
N
N
O
CH3
CH3
for
l
, d and
j
receptors (i.e., Ki values within 2-fold – see first entry
8
8
CH3
CH2CH3
X
X
in Table 1). A similar result was seen for the OH ? CONH2 switch in
other 2,6-methano-3-benzazocine (a.k.a. benzomorphan) core
structures (e.g., ethylketocyclazocine 2a).1 However, when the core
structure was the pentacyclic 4,5
natural products, a divergence in SAR was seen. For the morphine
(3a/3b) and naltrexone (4a/4b) pairs, binding affinity for , for
example, was reduced by 39- and 7-fold, respectively, when the
OH ? CONH2 switch was applied.3,4 We recently reported strong
evidence that in the naltrexone case, this divergent SAR is likely
a consequence of the furan O stabilizing, via a strong intramolecu-
lar H-bond, a carboxamide conformation (4c) that is not the bioac-
tive one.4 Therefore, 3-desoxy-3-carboxamido naltrexone 4b and,
by analogy, the morphine analogue 3b, must pay an energy penalty
to adopt the putative bioactive conformation depicted in 4d. For
the 2,6-methano-3-benzazocines 8-CAC (1b) and 2b, the putative
carboxamide bioactive conformation can easily be attained since
1a: X = OH (cyclazocine)
1b: X = CONH2 (8-CAC)
2a: X = OH (EKC)
2b: X = CONH2
a-epoxymorphinan derived from
l
NCH3
H
N
OH
3
3
O
O
X
O
X
OH
2
2
3a: X = OH (morphine) 4a: X = OH (naltrexone)
3b: X = CONH2
4b: X = CONH2
N
N
OH
OH
q
Presented, in part, at Wentland, M. P.; Lou, R.; Cohen, D. C.; Bidlack, J. M.
Carboxamido Analogues of Nalbuphine, Butorphanol and Nor-BNI. Proceedings of
the 66th Annual Meeting of the College on Problems of Drug Dependence, San Juan,
PR, June 12–17, 2004.
O
O
O
H2N
O
O
N
H
O
* Corresponding author. Tel.: +1 518 276 2234; fax: +1 518 276 4887.
E-mail address: wentmp@rpi.edu (M.P. Wentland).
H
4c
4d
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.10.134

204
M. P. Wentland et al. / Bioorg. Med. Chem. Lett. 19 (2009) 203–208
Table 1
Opioid binding data for carboxamido-substituted 2,6-methano-3-benzazocine, morphinan, 4,5a-epoxymorphinan and tramadol derivatives
2'-(S)
2'-(R)
CH3
Ph
O
O
N
N
N
N
N
H
H
(S)
(S)
(S)
(S)
(S)
CH3
CH3
CH3
CH3
CH3
(S)
CH3
CH3
CH3
CH3
CH3
X
X
X
X
X
Phenazocine core:
7a: X = OH
Mr2034 core:
Mr2034 isomer core:
9a: X = OH
Pentazocine core:
5a: X = OH
Metazocine core:
6a: X = OH
8a: X = OH
7b: X = CONH2
8b: X = CONH2
9b: X = CONH2
5b: X = CONH2
6b: X = CONH2
N
N
N
N
N
O
OH
OH
OH
OH
CH3
3
3
CH3
OH
H
O
O
O
X
X
X
X
X
O
H
OH
Ketocyclazocine core:
10a: X = OH
Naloxone core:
Butorphanol core:
11a: X = OH
Naltrexone-6-β-ol core:
13a: X = OH
Naltrexone-6-α-ol core:
14a: X = OH
12a: X = OH
10b: X = CONH2
12b: X = CONH2
11b: X = CONH2
13b: X = CONH2
14b: X = CONH2
N
H
N
Me2N
N
N
N
OH
OH
OH
HO
X
OH
CH3
C(CH3)3
OCH3
O
O
CH2
X
O
O
O
X
X
X
X
N
H
OH
Tramadol core:
19a: X = OH
19b: X = CONH2
19c: X = OCH3
Nalmefene core:
15a: X = OH
Nalorphine core:
16a: X = OH
nor-BNI (Norbinaltorphimine)core:
18a: X = OH
Buprenorphine core:
17a: X = OH
17b: X = CONH2
15b: X = CONH2
16b: X = CONH2
18b: X = CONH2
Compound
Ki ꢀ nM SEa [KBR]b
j
:
lc
j
:dd
[3H]DAMGO
[l]
[3H]Naltrindole
[d]
[3H]U69,593
[j]
Cyclazocine core
1a:X = OHe
1b: X = CONH2
0.16 0.01
0.31 0.03
[0.5]
[0.7]
[0.03]
[0.15]
[1.3]
[2.4]
[13]
2.0 0.22
5.2 0.36
[0.4]
0.07 0.01
0.06 0.001
[1.2]
[0.9]
[0.01]
[0.02]
[0.5]
[0.5]
[3.6]
[1.0]
2
5
30
90
f
EKC core
2a: X = OHg,h
0.78 0.10
1.2 0.12
3.4 0.41
9.8 0.50
[0.4]
[0.07]
[0.11]
[0.8]
[0.4]
[4.2]
[1.8]
0.62 0.11
0.70 0.08
1
2
4
14
g,h
2b: X = CONH2
Morphine core
3a: X = OHg,i
3b: X = CONH2
0.88 0.14
34 1.8
140 18
1900 81
24 2.3
2000 97
0.037
0.017
6
1
g,i
Naltrexone core
4a: X = OHj
0.11 0.006
0.71 0.058
60 3.2
550 40
0.19 0.005
0.36
0.6
0.065
3320
50
j
4b: X = CONH2
Pentazocine core
5a: X = OH
5b: X = CONH2
6.9 0.64
5.2 0.41
180 3.2
220 10
3.0 0.18
5.6 0.26
2
1
60
39
k
Metazocine core
6a: X = OH
6b: X = CONH2
3.8 0.66
1.6 0.07
140 17
320 43
9.9 0.22
19 0.54
0.4
0.08
14
17
k
Phenazocine core
7a: X = OH
7b: X = CONH2
0.20 0.07
0.015 0.0010
5.0 0.88
1.2 0.27
2.0 0.13
0.55 0.029
0.1
0.03
2.5
2
k
Mr2034 (20S) core
8a: X = OHg
0.19 0.01
0.052 0.013
[3.7]
3.6 0.40
2.0 0.15
0.09 0.01
0.089 0.004
2
0.6
40
22
g,k
8b: X = CONH2

M. P. Wentland et al. / Bioorg. Med. Chem. Lett. 19 (2009) 203–208
205
Table 1 (continued)
Compound
Ki ꢀ nM SEa [KBR]b
j
:
lc
j
:dd
[3H]DAMGO
[l]
[3H]Naltrindole
[d]
[3H]U69,593
[j]
Mr2034 diastereomer (20R) core
9a: X = OHg
9b: X = CONH2
4.0 0.54
2.9 0.17
[1.4]
[2.4]
[0.8]
[0.1]
[0.1]
[0.03]
[0.2]
[0.01]
[0.3]
[0.01]
[0.9]
67 4.3
34 0.10
[2.0]
1.5 0.07
2.8 0.24
[0.5]
3
1
45
12
g,k
Ketocyclazocine core
10a: X = OHg
3.3 0.66
1.4 0.07
20 2.7
20 2.3
[1.0]
1.0 0.24
1.8 0.10
[0.6]
3
0.8
20
11
g,k
10b: X = CONH2
Butorphanol core
11a: X = OH
11b: X = CONH2
0.12 0.058
0.15 0.019
12 3.8
14 2.1
[0.9]
0.22 0.023
0.39 0.057
[0.6]
0.55
0.38
50
40
k
Naloxone core
12a: X = OH
12b: X = CONH2
0.66 0.11
4.5 0.18
120 13
1000 59
[0.1]
1.2 0.072
46 1.3
[0.03]
[0.02]
[0.01]
[0.1]
0.55
0.1
100
22
k
Naltrexone-6-b-ol core
13a: X = OH
13b: X = CONH2
0.19 0.016
2.5 0.95
53 3.5
[<0.01]
[<0.01]
[0.1]
0.48 0.010
30 0.86
0.4
0.08
110
—
k
>10
lM
Naltrexone-6-
14a: X = OH
14b: X = CONH2
a
-ol core
0.21 0.09
7.1 0.88
56 5.1
>10
0.56 0.11
100 2.7
0.4
0.07
100
—
k
lM
Nalmefene core
15a: X = OH
15b: X = CONH2
0.44 0.083
2.6 0.33
9.3 1.5
67 2.6
0.12 0.0042
2.1 0.29
4
1
78
32
k
Nalorphine core
16a: X = OH
16b: X = CONH2
0.19 0.01
18 2.2
120 17
>10 lM
[<.01]
[0.1]
0.38 0.05
100 7.8
[0.004]
[0.3]
0.5
0.2
320
—
k
Buprenorphine core
17a: X = OH
17b: X = CONH2
0.21 0.024
0.77 0.065
2.9 0.49
54 8.4
0.62 0.073
2.1 0.12
0.3
0.4
5
26
k
Nor-BNI core
18a: X = OH
18b: X = CONH2
2.7 0.33
400 32
24 1.6
1000 241
[0.02]
[1.2]
0.027 0.008
21 4.7
[0.001]
[1.3]
100
19
900
50
k
Tramadol core
19a: X = OH
19b: X = CONH2
19c: X = OCH3 (tramadol)
8.6 0.37
9.5 0.25
1600 241
2900 66
2400 200
9.4 0.79
450 43
350 10
14 0.44
0.02
0.03
114
6
7
0.7
k
a
Binding assays used to screen compounds are similar to those previously reported (see Refs. 26 and 27). Membrane protein from CHO cells that stably expressed one type
of the human opioid receptor or (as indicated) from guinea pig brain were incubated with 12 different concentrations of the compound in the presence of either 1 nM
[3H]U69,593 (
l
), 0.25 nM [3H]DAMGO (d) or 0.2 nM [3H]naltrindole (
j
) in a final volume of 1 mL of 50 mM Tris–HCl, pH 7.5, at 25 °C. Incubation times of 60 min were used for
[3H]U69,593 and [3H]DAMGO. Because of a slower association of [3H]naltrindole with the receptor, a 3 h incubation was used with this radioligand. Samples incubated with
[3H]naltrindole also contained 10 mM MgCl2 and 0.5 mM phenylmethylsulfonyl fluoride. Nonspecific binding was measured by inclusion of 10
M naloxone. The binding was
l
terminated by filtering the samples through Schleicher and Schuell No. 32 glass fiber filters using a Brandel 48-well cell harvester. The filters were subsequently washed three
times with 3 mL of cold 50 mM Tris–HCl, pH 7.5, and were counted in 2 mL Ecoscint A scintillation fluid. For [3H]naltrindole and [3H]U69,593 binding, the filters were soaked
in 0.1% polyethyleneimine for at least 60 min before use. IC50 values will be calculated by least squares fit to a logarithm-probit analysis. Ki values of unlabeled compounds
were calculated from the equation Ki = (IC50)/1 + S where S = (concentration of radioligand)/(Kd of radioligand) – see Ref. 28 The Kd values for [3H]DAMGO, [3H]U69,593, and
[3H]naltrindole were 0.56 nM, 0.34 nM, and 0.10 nM, respectively. Data are means SEM from at least three experiments performed in triplicate.
b
KBR (Ki binding ratio) = Ki (OH)/Ki (CONH2) for l, d or j opioid receptors.
c
d
e
f
j:
l
= Ki (
l)/Ki (
j
).
j
:d = Ki (d)/Ki (
j).
See text for references to known phenolic-OH opioids.
See Ref. 29.
Guinea pig membranes.
See Ref. 1.
See Ref. 3.
g
h
i
j
See Ref. 4.
k
Proton NMR, IR and MS were consistent with the assigned structures of all new compounds. C, H, and N elemental analyses were obtained for all new targets and most
intermediates and were within 0.4% of theoretical values.
within each pair to determine if the SAR is consistent with that
seen with the limited number of pairs previously reported. Intrin-
sic opioid-receptor mediated activity for a number of high affinity
carboxamide-containing ligands compared to the corresponding
Subsequent to these findings, other researchers have published
studies where the OH ? CONH2 switch was applied to a number of
other well-known opioid core structures; these include the mor-
phinan class (e.g., levorphanol, cyclorphan, MCL-101),5 phenylpi-
peridine class6,7 and opioid peptides.8,9 We now wish to report
additional examples of carboxamido-substituted opioids. The
objective of this study was to take 15 well-known phenolic-OH-
containing opioids (structures shown in Table 1) having divergence
in core structures to use as substrates for the OH ? CONH2 switch
phenolic-OH prototype was also determined using [35S]GTP
binding assays.
cS
Carboxamide targets 5b–11b, 13b, 14b, 16b, 17b and 19b were
made directly from their corresponding known phenols using
methodology previously reported (Scheme 1).1,3,4 Triflate ester for-
mation was accomplished by treating the phenol with (CF3SO2)2O
and compare affinity and selectivity for l, d and j opioid receptors

206
M. P. Wentland et al. / Bioorg. Med. Chem. Lett. 19 (2009) 203–208
R'
R'
R'
R'
i
ii
iii
R''
R''
R''
R''
HO
R'''
CF3SO2O
R'''
NC
R'''
H2N
R'''
O
5a-11a, 13a,14a,
16a, 17a, 19a
5b-11b, 13b,14b,
16b, 17b, 19b
iv
Scheme 1. Reagents and conditions: (i) (CF3SO2)2O, pyr, CH2Cl2, 25 °C (Method A) or PhN(Tf)2, Et3N, CH2Cl2, 25 °C (Method B); (ii) Zn(CN)2, Pd(PPh3)4, DMF, 120 °C, 20 h or
under microwave radiation 150 °C for 15 min; (iii) t-BuOH, KOH, 82 °C (combined ii + iii = Method C); (iv) CO/NH3/Pd(OAc)2/DPPF in DMSO (Method D) or CO, HN(SiMe3)2,
PdCl2, PPh3, DMF followed by 2 N H2SO4 (Method E).
Carboxamide target 12b corresponding to naloxone (12a)20 was
prepared from the known 6-ethylene ketal derivative21 of nalox-
N
N
one. The OH ? CONH2 conversion was accomplished using Meth-
OH
OH
ods
B (74%) and E (69%); deketalization using 6 N HCl in
ii
refluxing acetone for 4 h (87%) gave 12b. As shown in Scheme 2,
carboxamide target 15b corresponding to nalmefene (15a)22 was
prepared from the known 3-desoxy-3-cyanonaltrexone 2123 by
first using standard Wittig olefination conditions to provide inter-
mediate 22 in 78% yield. Conversion of 22 to target 15b was
accomplished using K2CO3, H2O2 in DMSO in 78% yield. Lastly,
the carboxamide analogue 18b of nor-BNI (18a)24 was made
(Scheme 3) in 51% yield by treating 4b with hydrazine hydrochlo-
ride using conditions very similar to those used to make 18a.24
Target compounds were evaluated for their affinity and selec-
O
O
NC
Y
H2N
CH2
O
15b
21: Y = O
i
22: Y = CH2
Scheme 2. Reagents and conditions: (i) Ph3PCH3Br, NaNH2, THF, 25 °C, 20 h; (ii)
K2CO3, H2O2, DMSO, 25 °C, 3 h.
tivity for
l, d and j opioid receptors. Membrane protein from
CHO cells that stably expressed one type of the human opioid
receptor was used.26 In three instances where indicated, mem-
brane protein from guinea pig brain was used. Opioid receptors
from two species were used due to a change in our primary assay
midway through this study, however, where data are available,
absolute and relative affinities, using human or guinea pig recep-
tors were quite similar. Binding data for all new carboxamide tar-
gets compared to their phenolic-OH counterparts are detailed in
Table 1. Also included in Table 1 are columns that summarize
the Ki Binding Ratio (‘KBR’) for each core structure against the three
receptors [KBR = Ki (OH)/Ki (CONH2)]. A KBR that is P0.5 and 62
means the Ki values for the pair of compounds are within 2-fold
and have comparable binding affinity for that receptor. A KBR of
>2 indicates the carboxamide has higher affinity for the receptor
than does its OH counterpart; if it is <0.5, the OH partner has
higher affinity.
and pyridine in CH2Cl2 (Method A) or PhN(SO2CF3)2 and triethyl-
amine in CH2Cl2 (Method B). For targets 6b–11b, 13b, 14b and
16b, preparation of the corresponding carboxamide was enabled
via a two-step procedure where the triflate was first converted to
a nitrile using Zn(CN)2, Pd(PPh3)4, in DMF followed by partial
hydrolysis of the nitrile using KOH/t-BuOH (Method C). Alterna-
tively, triflate esters were directly converted to the carboxamide
targets 5b and 17b via palladium catalyzed procedures using CO/
NH3/Pd(OAc)2/DPPF in DMSO (Method D) or CO/HN(SiMe3)2/
PdCl2/PPh3 in DMF followed by workup with dilute sulfuric acid
(Method E).1,3
Using the methodology described in Scheme 1, the following
known phenolic-OH-containing opioids were converted to the cor-
responding novel carboxamide targets as follows: pentazocine
(5a)10 ? 5b, Methods
A
(93%) and
D
(18%); metazocine
(6a)11 ? 6b, Methods A (75%) and C (26% combined); phenazocine
(7a)12 ? 7b, Methods B (96%) and C (54%); Mr2034 (8a)13 ? 8b,
Methods A (95%) and C (98%); Mr2034 diastereomer (9a)13 ? 9b,
Methods A (96%) and C (88%); ketocyclazocine (10a)14 ? 10b, Meth-
ods A (98%) and C (93%); butorphanol (11a)15 ? 11b, Methods A
(77%) and C (89%); naltrexone-6-b-ol (13a)16 ? 13b, Methods B
For 2,6-methano-3-benzazocine core structures (5–10), high
affinity for
affinity for d is observed. The KBRs for
13 (for 7) indicating the carboxamide partners have, in several in-
stances, much higher potency than the OH counterpart. For d and
l
and
j
receptors is seen in all CONH2 targets; lower
l
range from 1.3 (for 5) to
j
(99%) and C (65%); naltrexone-6-a
-ol (14a)17 ? 14b, Methods B
receptors, within each pair, both partners have comparable affinity
(KBRs P 0.5 and 62) except for the phenazocine core (7) where the
carboxamide has considerably higher affinity (KBR = 4.2 and 3.6,
respectively). For the butorphanol pair 11, an example of the mor-
phinan class, a very similar trend in KBRs (i.e., near unity) and
(98%) and C (86%); nalorphine (16a)10 ? 16b, Methods B (97%)
and C (11%); buprenorphine (17a)18 ? 17b, Methods B (95%) and E
(48%); and tramadol active metabolite (19a)19 ? 19b, Methods A
(91%) and C (62%).
N
N
N
OH
OH
HO
(i)
O
H2N
O
O
O
O
H2N
N
H
NH2
4b
O
18b
O
Scheme 3. Reagents and conditions: (i) H2NNH2ꢁHCl, DMF, 150 °C, 1 h.

M. P. Wentland et al. / Bioorg. Med. Chem. Lett. 19 (2009) 203–208
207
receptor selectivity (i.e., higher affinity for
l
and
j
receptors than
were 0.03 or lower. For the seven known phenolic-OH 4,5
a-epoxy-
for d) was seen compared to published data for the morphinans
levorphanol, cyclorphan and MCL-101.5 Comparing receptor selec-
tivities of carboxamides 5b–11b to their phenolic-OH counterparts
morphinans 12a–18a studied, six have comparable affinity for
l
and and much less affinity for d; these mixed compounds
j
l/j
are 12a–17a. Comparing this receptor selectivity pattern to that
5a–11a, the
j:l and j:d selectivity ratios are similar within each
of corresponding carboxamides 12b–17b, little correlation is ob-
pair with two exceptions. The exceptions are the metazocine (6)
and phenazocine (7) examples where a significant divergence in
served since
for ; the
j
affinity is very low for 12b–17b compared to affinity
[Ki ( )/Ki ( )] ratios ranged from 1 for nalmefene
l
j
:l
l
j
the
tively, are much lower (i.e., the carboxamide has much lower affin-
ity for than ) than seen in the other core structures.
As predicted from our previous results with morphine and nal-
trexone,3,4 there is an overwhelming trend in the seven 4,5
-epox-
ymorphinan pairs (12a/b–18a/b) studied that the carboxamide
partner has lower receptor affinity than the corresponding OH-
containing opioid. KBRs versus all three receptors were <0.5. For
j
:
l
selectivity is seen; the
j
:
l
ratios of 0.08 and 0.03, respec-
analogue 15b to 0.07 for the naltrexone-6-b-ol analogue 14b.
Receptor selectivity for 18b showed a similar trend to 18a, the
j
l
well-known
-selective. However, the carboxamide partner displayed very
low affinity for all receptors, including . The carboxamide partner
j
selective ligand nor-BNI,24 in that both were
j
a
j
19b of the active metabolite 19a19 of tramadol 19c had comparable
binding affinities (KBRs = 0.9–1.3) and a similar receptor selectivity
pattern to 19a for all three receptors. Following characterization of
these compounds in our laboratories, an independent study
describing their syntheses and biological properties appeared.25
Intrinsic opioid-receptor mediated activity for a number of high
affinity carboxamide-containing ligands compared to the correspond-
l, KBRs ranged from 0.3 for the buprenorphine core 17a/b to
0.01 for the nalorphine and nor-BNI cores, 16a/b and 18a/b, respec-
tively. Against the d receptor, binding affinity for the carboxamide
partner was also very low compared to the traditional OH partner
and KBRs ranged from 0.1 (12a/b, 15a/b and 17a/b) to <0.01 for
13a/b and 14a/b. Carboxamide targets 12b–18b displayed rela-
ing phenolic-OH prototype was determined using [35S]GTP
ing assays at and receptors; results are shown in Table 2. Due
to the relatively poor binding affinity to receptors, these
cS bind-
l
j
tively low affinity for
j
and KBRs of all pairs (12a/b–18a/b) for
j
d
were low and ranged from 0.3 (buprenorphine core 17a/b) to as
low as 0.001 (nor-BNI core 18a/b); more than half of these values
compounds were not evaluated for functional activity at d. Proce-
dures similar to those previously reported were used.29 For the
Table 2
EC50 and Emax values for the stimulation of [35S]GTP S binding and IC50 and Imax values for the inhibition of agonist-stimulated [35S]GTP
c cS binding to the human l and j opioid
receptorsa
Compound
Functional description
EC50 (nM)
Emax (% maximal stimulation)
IC50 (nM)
Imax (% maximal inhibition)
Mu opioid receptor
DAMGO
1a
1b
6a
6b
7a
7b
8a
8b
9a
9b
10a
10b
11a
11b
17a
17b
Agonist
55
7
116
4
NIb
13 2.2
15 4.0
NI
25 15
NI
NI
17 3.3
58 14
NI
250 36
130 24
150 57
14 3.3
30 5.6
NI
Agonist/antagonist
Weak agonist/antagonist
Agonist
Agonist/antagonist
Agonist
4.0 1.3
1.0 0.70
40 4.6
24 7.3
27 3.1
5.3 0.87
2.7 0.72
NA
24 2.7
20 2.4
78 4.6
23 1.5
79 0.82
110 2.9
31 4.9
ꢂ20%
67 3.1
76 3.6
NI
70 2.6
NI
Agonist
NI
Agonist/antagonist
Weak agonist/antagonist
Weak agonist
Agonist/antagonist
Agonist/antagonist
Agonist/antagonist
Weak agonist/antagonist
Weak agonist/antagonist
Agonist/antagonist
Antagonist
61 3.9
89 3.9
NI
68 2.9
63 5.2
73 3.1
54 2.7
86 6.9
48 0.87
57 8.4
NA
ꢂ20%
101 50
21 3.2
4.2 1.4
NA
NA
0.11 0.021
NA
42 2.0
39 0.21
23 1.5
22 5.8
11 2.6
32 6.2
9.0 7.0
0.59 0.21
4.0 0.80
Kappa opioid receptor
U50,488
1a
1b
6a
6b
Agonist
Agonist
Agonist
Agonist
36 5.0
1.3 0.20
2.4 0.65
120 26
NT
77 11
57 3.8
59 2.4
50 2.3
NT
NI
NI
NI
NI
NI
NI
NI
NI
NT
NT
NT
7a
7b
8a
8b
9a
9b
10a
10b
11a
11b
17a
17b
Agonist
Agonist/antagonist
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
Agonist
8.1 0.93
4.4 1.2
1.0 0.13
30 9.9
41 11
170 29
3.3 0.17
9.6 4.1
2.9 1.0
3.8 0.42
0.18 0.014
NA
95 4.5
60 2.6
60 2.0
75 3.6
83 10
65 3.3
84 1.2
60 0.97
73 8.8
55 4.1
35 2.5
ꢀ1.3 2.5
NI
NI
49 4.4
NI
NT
NT
NT
NI
NI
NI
640 137
NI
NT
NT
NT
NI
NI
NI
Agonist
Agonist/antagonist
Antagonist
NI
15 6.8
140 18
NI
24 1.7
59 5.1
NA, not applicable.
NT, not tested.
a
See Ref. 29 for experimental details. Data are mean values SEM from at least three separate experiments, performed in triplicate. For calculation of the Emax values, the
S binding was set at 0%. For inhibition studies, 200 nM DAMGO was used as the agonist for the receptor, U50,488 at final concentration of 100 nM was used
receptor.
NI, no inhibition.
basal [35S]GTP
for the
c
l
j
b

208
M. P. Wentland et al. / Bioorg. Med. Chem. Lett. 19 (2009) 203–208
benzomorphan pair cyclazocine (1a) and 8-CAC (1b), both are
mixed agonists/antagonists at the receptor and agonists at
Both displayed similar potencies that correlated well with binding
affinities. In the case of the 6a/6b pair (metazocine core) against
the carboxamide partner 6a is an agonist/antagonist while the OH
partner is an agonist. Because the affinity of 6b for receptors was
relatively weak, it was not studied in the [35S]GTP
S binding assay.
For 7a/7b having a phenazocine core, both were agonists at hav-
ing qualitatively similar potencies. Against , 7b was a mixed ago-
nist/antagonist and 7a was an agonist. The agonist potencies of the
two were similar. A divergence in functional activity at the
receptor was noted for 8a (agonist/antagonist) and 8b (weak ago-
nist/antagonist). At the receptor, both were agonists, although 8a
had much higher potency. Divergence was also seen at the recep-
tor for the 9a (weak agonist) and 9b (agonist/antagonist) pair.
Against , both were agonists, however, 9a had much higher po-
tency. For the 10a/10b, another benzomorphan pair, both were
mixed agonists/antagonists at and agonists at ; agonist poten-
cies at were similar. For the butorphanol core, an example of a
tetracyclic morphinan, the OH partner 11a exhibited a weak ago-
nist/antagonist profile at , whereas the carboxamide 11b was an
antagonist. At , both were agonists having similar potencies. At
and receptors, the carboxamide derivative 17b of buprenor-
phine displayed a somewhat different profile (antagonist) than
buprenorphine (17a) (agonist/antagonist). As and antagonists,
17a was more potent than 17b. In the [35S]GTP
S binding assay
mediated by the opioid receptor, some compounds showed a less
than maximal activation of [35S]GTP
S binding, but did not have
12a–18a. These observations were, in fact, predicted from our ear-
lier studies and are consistent with our pharmacophore hypothesis
concerning the bioactive conformation of the carboxamide group.4
l
j.
l,
Receptor selectivity of carboxamides 12b–18b for l, d and j recep-
tors was, in general, similar to that seen for corresponding OH part-
ners 12a–18a. SAR findings that we now report are consistent with
our previous OH ? CONH2 switch studies1,3–5 and as well as those
from other laboratories.5–9 For those OH/CONH2 pairs studied in
j
c
l
j
[
35S]GTP
c
S binding assays, within each pair, similarities in their
function and potency profiles were frequently observed especially
at the receptor. At the receptor, more divergence was observed
l
j
l
with a trend towards the carboxamide partner displaying less ago-
nist activity.
j
l
Acknowledgments
j
We gratefully acknowledge the contributions of Rensselaer’s
mass spectroscopist Dr. Dmitri Zagorevski and the technical assis-
tance provided by Brian I. Knapp of the University of Rochester.
Funding of this research was from NIDA (DA12180 and KO5-
DA00360) and the NSF (Agilent 1100 series LC/MSD system).
l
j
j
l
j
l
j
References and notes
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Bioorg. Med. Chem. Lett. 2001, 11, 623.
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5. Zhang, A.; Xiong, W.; Bidlack, J. M.; Hilbert, J. E.; Knapp, B. I.; Wentland, M. P.;
Neumeyer, J. L. J. Med. Chem. 2004, 47, 165.
l
j
c
j
c
antagonist properties. While most partial agonists have antagonist
properties, not all compounds that produce less than a maximal ef-
fect in an assay have antagonist effects. Some compounds are less
efficacious than an agonist that produces a maximal effect. While
many of these compounds have antagonist properties, too, making
them partial agonists, there are some compounds that are less effi-
cacious than full agonists but do not have antagonist properties.
Compounds 1a, 1b, 6a, 8a and 10b are examples of compounds
6. Le Bourdonnec, B.; Belanger, S.; Cassel, J. A.; Stabley, G. J.; DeHaven, R. N.; Dolle,
R. E. Bioorg. Med. Chem. Lett. 2003, 13, 4459.
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E.; Woodland, J.; Surface, P.; Mitch, C. Bioorg. Med. Chem. Lett. 2005, 15, 3844.
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G. J.; Graczyk, T. M.; DeHaven, R. N. Bioorg. Med. Chem. Lett. 2004, 14, 3545.
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that have a lower efficacy in the [35S]GTP
ated by the receptor, but these compounds do not antagonize
35S]GTP
S binding that was induced by the agonist U50,488.
A series of novel opioids 5b–19b have been prepared where the
cS binding assay medi-
j
[
c
j
phenolic-OH group of traditional and well-studied opioids 5a–19a
was replaced by a carboxamide (CONH2) group. Characterization
of target and known compounds in opioid receptor binding assays
revealed that carboxamide targets 5b–11b and 19b derived from
2,6-methano-3-benzazocine (a.k.a. benzomorphans), morphinan
or tramadol-based core structures have high affinity to
l and j
receptors and relatively low affinity for d receptors. Compared to
their phenolic-OH counterparts 5a–11a and 19a, comparable or en-
hanced affinity was seen for all three receptors. Receptor selectivity
for these carboxamides was, in general, similar to the OH partners.
It is interesting to note that carboxamide 7b having the phenazo-
cine core, has KBRs for all three receptors considerably higher than
the other 2,6-methano-3-benzazocine cores and has the least
18. Lewis, J. W. Drug Alcohol Depend. 1985, 14, 363.
19. Subrahmanyam, V.; Renwick, A. B.; Walters, D. G.; Young, P. J.; Price, R. J.;
Tonelli, A. P.; Lake, B. G. Drug Metab. Dispos. 2001, 29, 1146.
20. Linder, C.; Fishman, J. J. Med. Chem. 1973, 16, 553.
21. Schmidhammer, H.; Spetea, M.; Schuetz, J.; Greiner, E.; Schuellner, F.; Sailer, B.;
Stuebegger, K. WO 2004005294, 2004.
22. Hahn, E. F.; Fishman, J.; Heilman, R. D. J. Med. Chem. 1975, 18, 259.
23. Kubota, H.; Rice, K. C. Tetrahedron Lett. 1998, 39, 2907.
24. Portoghese, P. S.; Lipkowski, A. W.; Takemori, A. E. J. Med. Chem. 1987, 30, 238.
25. Carson, J. R.; Pitis, J. M. US Patent Appl. US2005/0256203.
26. Neumeyer, J. L.; Zhang, A.; Xiong, W.; Gu, X.; Hilbert, J. E.; Knapp, B. I.; Negus, S.
S.; Mello, N. K.; Bidlack, J. M. J. Med. Chem. 2003, 46, 5162.
27. Archer, S.; Seyed-Mozaffari, A.; Jiang, Q.; Bidlack, J. M. J. Med. Chem. 1994, 37,
1578.
degree of selectivity between
is the observation that when a divergence in selectivity ratios is
seen, there is a trend toward higher selectivity for than
A divergent SAR was seen for carboxamides 12b–18b having the
-epoxymorphinan core. Without exception, these carboxam-
ides had lower and in many instances, much lower affinity for
d and opioid receptors than their phenolic-OH counterparts
l, d and j. Another note of interest
l
j.
4,5a
28. Cheng, Y. C.; Prusoff, W. H. Biochem. Pharmacol. 1973, 22, 3099.
29. Wentland, M. P.; VanAlstine, M.; Kucejko, R.; Lou, R.; Cohen, D. J.; Parkhill, A. L.;
Bidlack, J. M. J. Med. Chem. 2006, 49, 5635.
l,
j