Design and synthesis of ether analogues as potent and selective M2 muscarinic receptor antagonists

Article abstract of DOI:10.1016/S0960-894X(01)00100-7

Novel, selective M₂ muscarinic antagonists, which replace the metabolically labile styrenyl moiety of the prototypical M₂ antagonist 1 with an ether linkage, were synthesized. A detailed SAR study in this class of compounds has yielded highly active compounds that showed M₂ K_i values of < 1.0 nM and > 100-fold selectivity against M₁, M₃, and M₅ receptors.

Full text of DOI:10.1016/S0960-894X(01)00100-7

Bioorganic & Medicinal Chemistry Letters 11 (2001) 891–894
Design and Synthesis of Ether Analogues as Potent and Selective
M2 Muscarinic Receptor Antagonists
Yuguang Wang,* Samuel Chackalamannil, Wei Chang, William Greenlee,
Vilma Ruperto, Ruth A. Duffy, Robert McQuade and Jean E. Lachowicz
Schering-Plough Research Institute, 2015 Galloping Hill Road, Kenilworth, NJ 07033, USA
Received 27 November 2000; accepted 1 February 2001
Abstract—Novel, selective M2 muscarinic antagonists, which replace the metabolically labile styrenyl moiety of the prototypical M2
antagonist 1 with an ether linkage, were synthesized. A detailed SAR study in this class of compounds has yielded highly active
compounds that showed M2 Ki values of <1.0 nM and >100-fold selectivity against M1, M3, and M5 receptors. # 2001 Elsevier
Science Ltd. All rights reserved.
Acetylcholine is an important neurotransmitter that
plays a key role in learning and memory.1 The cognitive
effects of acetylcholine are mediated by agonist-induced
stimulation of post-synaptic nicotinic and muscarinic
M1 receptors.2 Under normal physiological conditions,
synaptic acetylcholine levels are modulated by a feed-
back shut off mechanism initiated by acetylcholine-
induced stimulation of presynaptic M2 receptors. The
senile dementia associated with Alzheimer’s disease is
directly correlated with depleted cholinergic activity in
the cortical and hippocampal areas of brain and the
current form of therapy addresses this issue by inhibit-
ing cholinesterase, which breaks down acetylcholine.
There have also been considerable efforts to develop an
M1 agonist for the treatment of Alzheimer’s disease.
Several M1 agonists have been demonstrated to improve
cognitive functions in clinical trials.3 Alternatively,
selective inhibition of presynaptic M2 muscarinic recep-
tors has been shown to enhance acetylcholine levels in
vitro as well as in vivo.4 The success of this approach
would greatly depend on selectively achieving M2
receptor inhibition, since inhibition of post-synaptic M1
receptors and peripheral M1 and M3 receptors carries
the potential liabilities of serious side effects.
In a previous communication,5 we have described the
design and synthesis of highly potent and selective M2
antagonists represented by structure 1 (Fig. 1). In vitro
metabolism studies on these compounds using rat liver
microsomes gave the aldehyde 36 as the principal meta-
bolite which presumably arises through the rearrange-
ment of the putative intermediate epoxide 2. Since the
epoxide 2 as well as the aldehyde 3 has potential toxic
liabilities, we redirected our efforts toward identifying
an ethylidine replacement. Herein we wish to report the
successful outcome of our efforts.
An ether linkage was explored as a surrogate for the
ethylidine unit (Fig. 2). The CꢀOꢀC bond angle of
ethers ranges from 124ꢁ (diphenyl ether) to 111.5ꢁ
Figure 1. Metabolism of 1.
*Corresponding author. Tel.: +1-908-740-3508; fax: +1-908-740-4563; e-mail: yuguang.wang@spcorp.com
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00100-7

892
Y. Wang et al. / Bioorg. Med. Chem. Lett. 11 (2001) 891–894
containing the methelenedioxyphenyl moiety for further
SAR studies by varying the substituents on the piper-
idine nitrogen.
A representative SAR of the sulfonamide series is pre-
sented in Table 2. It is evident that the binding affinity is
affected by the size of the alkyl sulfonyl group. Com-
pounds with smaller and unbranched sulfonyl alkyl
groups (14 and 15) demonstrated higher M2 binding
affinity. However, their selectivity versus M1, M3, and
M5 are low. Aromatic 1-naphthyl sulfonamide 19
showed low M2 binding affinity and selectivity.
Figure 2. Designed targets.
(dimethyl ether),7 roughly corresponding to an sp2 car-
bon and the lone pair of electrons somewhat emulating
the potential role, if any, of the p electrons of the ethyl-
idine unit. Additionally, we decided to replace the p-
methoxy substitute of the phenyl ring by a metabolically
less labile methylenedioxy group.8 Based on this rationale,
the following two types of targets were explored.
The data for carbamate derivatives are presented in
Table 3. Since most of the carbamates reported in the
previous communication had high M2 binding affinity
with low selectivity,5 a few carbamates were prepared in
the ether series also. The results in Table 3 are con-
sistent with the results previously reported. In spite of
the excellent M2 binding affinity, we have so far been
unable to achieve acceptable selectivity in the carbamate
series.
The synthesis of these compounds from commercially
available starting materials is described in Scheme 1.
Reduction of ketone 4 was followed by coupling with 4-
iodophenol to produce ether 5. Displacement of the
iodo group of 5 with suitably substituted benzenethiol
gave intermediate 6,9 deprotection of which with tri-
fluoroacetic acid followed by reductive amination affor-
ded 8. Oxidation of 8 with mCPBA followed by
deprotection gave sulfone 10, which was transformed to
the final targets by functionalization of the piperidine
nitrogen as shown in Scheme 1.10
The most promising results were obtained from the
amide series (Table 4). The selection of the R groups for
the amide series greatly affects M2 binding affinity and
selectivity. Substituted alkyl amides 23 and 24 are less
potent and selective. Aromatic amide 25 improved M2
binding affinity by 10-fold, but it has low M2 selectivity
versus other receptor subtypes. Unlike the sulfonamide
series (19), introduction of a 1-naphthyl group (26) in
the amide series dramatically increased both the M2
binding affinity and selectivity. This result provided a
new direction for SAR modification.
The binding affinity of the newly synthesized targets
against cloned human muscarinic receptors, assayed
according to the reported protocol,11 is presented in
Table 1.
As shown in Table 1, the direct replacement of the
ethylidine moiety of 12 with an oxygen atom to generate
compound 13 resulted in reduced M2 binding affinity by
about 1000-fold. However, the lost binding affinity of 13
could be restored to a reasonable level by replacement
of the 4-methoxy group with a 3,4-methylenedioxy
group (14). These results indicated that the decreased
binding affinity of ether derivatives could be compen-
sated for by appropriate structural modification in other
parts of the molecule. Due to the excellent M2 binding
affinity of compound 14, we selected the type 2 targets
Since the amide series offered superior M2 binding and
selectivity profile compared with the sulfonamide series,
we focused further modifications on the amide series. A
series of amides with 1-naphthyl bioisosteres was pre-
pared. The results are shown in Table 5. Among the
quinoline analogues, the position of the nitrogen atom
plays an important role for M2 binding and selectivity.
For example, 4-quinoline analogue 27 is highly potent
with 100-fold selectivity versus M1, M3, and M5 while
the 8-quinoline analogue (29) showed low M2 potency
and selectivity. Introduction of 2,3-dimethyl phenyl
Scheme 1. (a) NaBH4, EtOH, 100%; (b) Ph3P, DEAD, THF, 4-iodophenol, 64%; (c) 4-methoxybenzenethiol or 3,4-methylenedioxybenzenethiol,
DMPU, CuI, K2CO3, 50–70%; (d) 30% TFA/CH2Cl2, 100%; (e) NaBH(AcO)3, 1,2-dichloroethane, 4, 70–85%; (f) MeSO3H/mCPBA, 25%TFA/
CH2Cl2, 50–70%; (g) sulfonyl chloride, chloroformate, or acyl chloride, Et3N, CH2Cl2, 85–98%.

Y. Wang et al. / Bioorg. Med. Chem. Lett. 11 (2001) 891–894
893
Table 1. Comparison of M2 binding affinity among different series
Table 3. Results of M2 binding affinity and selectivity of the carba-
mate series
Compound
X
Y
R
M2 (Ki, nM)
1
4-Methoxy
4-Methoxy
4-Methoxy
CH¼CH2
CO2Et
0.11
0.29
138.8
1.6
Compound
R
M2, Ki M1/M2 M3/M2 M4/M2 M5/M2
(nM)
12
13
14
CH¼CH2 SO2-n-C3H7
O
O
SO2-n-C3H7
SO2-n-C3H7
3,4-Methylenedioxy
20
21
22
Et
i-C3H7
Ph
6.0
3.2
1.1
4
59
48
2
19
61
1
7
11
14
10
69
Table 2. Results of M2 binding affinity and selectivity of the sulfon-
amide series
Table 4. Results of M2 binding affinity and selectivity of the amide
series
Compound
R
M2, Ki M1/M2 M3/M2 M4/M2 M5/M2
(nM)
Compound
R
M2, Ki M1/M2 M3/M2 M4/M2 M5/M2
(nM)
15
14
16
17
18
19
Et
n-C3H7
i-C3H7
n-C4H9
CH2Ph
1-Naphthyl
2.0
1.6
14.0
6.9
15.0
34.7
49
58
10
35
19
2
122
46
2
50
12
8
12
25
2
4
4
46
39
16
35
1
23
24
25
26
CH2OC2H3 17.9
6
8
5
5
2
3
13
13
15
50
CH2OPh
Ph
1-Naphthyl
59.5
1.6
18
357
6
216
6
28
1
4
0.5
Table 5. Results of M2 binding affinity and selectivity of 1-naphthyl bioisosteres
Compound
R
M2, Ki
(nM)
M1/M2
M3/M2
M4/M2
M5/M2
26
27
28
29
30
31
1-Naphthyl
4-Quinoline
5-Quinoline
0.5
1.0
0.5
12
0.4
0.2
357
140
233
9
155
234
216
117
124
10
132
109
28
18
21
2
31
23
50
1485
64
4
56
8-Quinoline
2,3-DiMePh
2-(3-Methylthiophenyl)
196
group (30) also resulted in excellent M2 binding affinity
and selectivity. Compounds 27 and 31 have the best
overall M2 binding and selectivity profile among the
ether analogues.
Acknowledgements
The authors would like to thank Mr. Yao H. Ing and
Dr. P. Das for mass spectral data, Dr. N. J. Murgolo
for molecular modeling studies, and Dr. J. W. Clader
for helpful discussion.
In summary, the metabolically labile alkene group has
been successfully replaced with an ether linker. The
results of SAR studies showed that the 1-naphthyl
group and its bioisosteres contribute to high M2 binding
affinity and selectivity in the amide series. Complete
results of SAR studies, as well as the in vivo efficacy
studies for the amide series, will be reported in due
course of time.
References and Notes
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Y. Wang et al. / Bioorg. Med. Chem. Lett. 11 (2001) 891–894
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11. For radioligand binding analysis, each muscarinic recep-
tor subtype was stably expressed in CHO-K1 cells. Clonal cell
lines were selected which expressed receptors at levels between
1 and 9 pmol/mg protein. The Kd of QNB (l-quinuclidinyl
benzilate) at each receptor subtype was determined by satura-
tion binding using 5–2500 pM [3H]QNB in 10 mM potassium
phosphate buffer, pH 7.4. Protein concentrations were adjus-
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methylcellulose. Non-specific binding was defined by 0.5 mM
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at rt), bound and free radioactivity were separated by filtration
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antagonist activity was performed on related compounds by
measuring the effects of the compound’s adenylyl cyclase
inhibition mediated by oxotremorine M. Ki was expressed as
mean of duplicate value (SEM<15%). All determinations
were performed at least twice.
6. Metabolite 3 was confirmed by analysis with an authentic
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