Benzylidene ketal derivatives as M2 muscarinic receptor antagonists

Article abstract of DOI:10.1016/S0960-894X(00)00553-9

Benzylidene ketal derivatives were investigated as selective M₂ receptor antagonists for the treatment of Alzheimer's disease. Compound 10 was discovered to have subnanomolar M₂ receptor affinity and 100-fold selectivity against other muscarinic receptors. Also, 10 demonstrated in vivo efficacy in rodent models of muscarinic activity and cognition. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0960-894X(00)00553-9

Bioorganic & Medicinal Chemistry Letters 10 (2000) 2727±2730
Benzylidene Ketal Derivatives as M2 Muscarinic Receptor
Antagonists
Craig D. Boyle,* Samuel Chackalamannil, Lian-Yong Chen, Sundeep Dugar,
Pradeep Pushpavanam, William Billard, Herbert Binch, III, Gordon Crosby,
Mary Cohen-Williams, Vicki L. Con, Ruth A. Du€y, Vilma Ruperto
and Jean E. Lachowicz
Schering-Plough Research Institute, 2015 Galloping Hill Road, Kenilworth, NJ 07033, USA
Received 22 June 2000; accepted 26 September 2000
AbstractÐBenzylidene ketal derivatives were investigated as selective M2 receptor antagonists for the treatment of Alzheimer's
disease. Compound 10 was discovered to have subnanomolar M2 receptor anity and 100-fold selectivity against other muscarinic
receptors. Also, 10 demonstrated in vivo ecacy in rodent models of muscarinic activity and cognition. # 2000 Elsevier Science
Ltd. All rights reserved.
Alzheimer's disease (AD) is characterized initially by a
decline in memory function that hinders normal daily
living, followed by a more severe loss of cognitive func-
tion and eventual morbidity and mortality. Cholinergic
marker loss and selective degeneration of cholinergic
neurons in the basal forebrains of AD patients have
implicated cholinergic system enhancement as a poten-
tial AD treatment.1 The current available cholinergic
pharmacotherapy for AD is in the form of acet-
ylcholinesterase inhibitors, which hinder the degrada-
tion of acetylcholine (ACh) in the synapse.1a Elevation
of synaptic ACh levels could also be achieved by selec-
tively inhibiting presynaptic muscarinic receptors of the
M2 subtype, agonist-induced stimulation of which shuts
o€ ACh release.1b,c Since inhibition of postsynaptic M1
receptors could o€set the bene®cial e€ects of pre-
synaptic M2 receptor inhibition, and M3 receptors are
associated with GI side e€ects, it is essential that such
an agent be selective for the M2 receptor.1d
the development of these compounds as clinical candi-
dates. We wish to report here the identi®cation of new
compounds which have improved upon these earlier
leads by incorporating metabolically stable spirocycles
at the benzylic position as well as raising M2 selectivity
to 100-fold over other muscarinic receptors. Addition-
ally, these new compounds demonstrate promising oral
activity in rodents.
Compounds I2 and II3 are reported M2 receptor
antagonists with a modest level of selectivity against the
M1 and M3 receptors.4 Although these compounds have
M2 selectivity, structural changes could confer increased
selectivity over M1 and M3 receptors. In addition, pre-
sence of the metabolically labile styrene moiety in II and
a chemically labile benzylic cyano group in I preclude
The syntheses of the spirocyclic targets are outlined in
Scheme 1. A diarylsul®de was formed via an arylthiol
addition/elimination reaction on the N-BOC protected
*Corresponding author. Tel.: +1-908-740-3503; fax: +1-908-740-
7152; e-mail: craig.boyle@spcorp.com
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00553-9

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C. D. Boyle et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2727±2730
Scheme 1. General synthesis of ketal analogues. Reagents and conditions: (a) (BOC)2O, 10% NaOH, Et2O; (b) ArSH, NaH, DMF; (c) TFA,
CH2Cl2; (d) Ti(O-i-Pr)4, N-carbethoxy-4-piperidone, CH2Cl2, then NaBH3CN, MeOH; (e) NaBO3ꢀ 4H2O (1 or 2 equiv), AcOH; (f) HO(CH2)mOH,
p-TsOH, HC(OEt)3, PhCH3, Á; (g) HS(CH2)mSH, BF3ꢀ OEt2, CH2Cl2.
analogue of A. Subsequent deprotection followed by
reductive amination a€orded the bis-piperidine B0. Sul-
®de oxidation provided the desired sulfoxide and sul-
fone intermediates B1 and B2. From B1 and B2, simple
ketalization provided the benzylidene ketals C.5
other muscarinic receptors by 2-fold. However, the ®ve-
membered spirocyclic ketals 2 and 4 had M2 anity
comparable to that of II. This phenomenon could be
due to the ketone of 1 deactivating the aromatic ring to
a greater degree than the ketal or the ole®n. Also, the
ketal heteroatoms could be in a more favorable position
for M2 receptor binding than the ketone oxygen. Note
that only a limited amount of steric bulk was tolerated
as the dioxane and dithiane compounds had decreased
M2 anity over their ®ve-membered ring counterparts.
Table 1 summarizes initial studies with analogues of II.6
Replacement of the methylene unit of II with oxygen
decreased M2 anity by 40-fold and selectivity over the
Table 1. Initial analogues of II
Having shown promise, this ketal series was synthesized
with a methylenedioxyphenyl replacement for the
p-methoxyphenyl moiety in order to avoid the meta-
bolic demethylation often associated with methoxy-
phenyl groups.7 The ketone intermediate 6 had good M2
anity and selectivity. As with the p-methoxyphenyl
derivatives, the ketals generally improved M2 anity
over the ketone, but su€ered from a lack of M2 selec-
tivity. However, compound 8 demonstrated that mod-
i®cations to the ketal structure could enhance M2
selectivity without adversely a€ecting the M2 Ki value.
Only speci®c positioning of steric bulk could be toler-
ated, as there was a 60-fold di€erence in M2 anity
between 8 and its enantiomer 9.
Compound
Ar
X
M2 Ki
(nM)
M1/M2 M3/M2
II
1
2
3
4
5
0.07
2.91
0.16
0.72
0.11
14.3
68.3
26.8
28.3
21.0
22.5
4.4
21.9
10.2
7.1
After displaying the potential of the ketal to positively
e€ect M2 potency without hurting selectivity, we
replaced the ethyl carbamate with an n-propylsulfon-
amide group in order to avoid the decarboxylation that
could occur with II. To this end, ketal derivatives of
compound 7 were treated with KOH in re¯uxing ethyl-
ene glycol to remove the ethyl carbamate. The resulting
intermediate was exposed to n-propylsulfonyl chloride
to provide the ®nal ketal compounds 10±19. Both the
ketal moiety and the oxidation state of the biaryl sulfur
linkage were modi®ed.
14.4
3.0
2.9
6
7
8
0.13
47.6
4.1
7.5
1.3
0.073
0.030
In the sulfone series (Table 2: even numbers), the non-
substituted dioxolane 10 exhibited the most desirable
M2 anity/selectivity pro®le. The tetrahydrofuran deri-
vative 168 demonstrated that both heteroatoms of the
ketal were required for M2 potency, as 16 had reduced
M2 anity of 500- and 160-fold versus the dioxolane 10
and dithiolane 14, respectively. Also, the extra hetero-
atom present in 10 and 14 imparted signi®cant selectivity
21.7
46.0
9
1.82
10.2
6.1

C. D. Boyle et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2727±2730
2729
Table 2. n-Propylsulfonamide series
Compound
X
M2 Ki (nM)
M1/M2
M3/M2
10 (n=2)
11 (n=1)
0.010
0.823
101.5
10.8
93.0
10.0
12 (n=2)
13 (n=1)
1.22
9.09
8.7
7.5
46.1
12.9
Figure 1. Microdialysis data. Compound 10, 10 mpk dose, po, methyl
cellulose in water.
14 (n=2)
15 (n=1)
0.031
0.82
62.6
10.7
57.4
12.8
needed to simply stay in the light chamber and not enter
the darkened area, hence the name passive-avoidance
response. In a test session given 24 h later, the latency
time (length of time the rats avoided the dark chamber)
was measured, with longer latency denoting better
reference memory. Previous work has indicated that
cholinergic enhancing drugs improve learning as mea-
sured in this PAR experiment, and that these conditions
served as a useful model of impaired memory caused by
cholinergic hypofunction. When 10 was dosed to young
rats 2 h before training followed by testing 24 h later, a
signi®cant increase in latency time was observed with
doses of 0.1 mg/kg or greater (Fig. 2). This enhancement
of reference memory in vivo demonstrated the utility of 10
to improve cognition via cholinergic system stimulation.
16 (n=2)
17 (n=1)
5.07
12.1
0.22
0.49
0.24
0.50
18 (n=2)
19 (n=1)
1.02
34.8
42.6
4.6
93.4
11.8
over the other muscarinic receptors, as the tetra-
hydrofuran 16 displayed a lower Ki value for the other
muscarinic receptors when compared to M2. In com-
parison to the sulfone series, the sulfoxide series (Table
2: odd numbers) was disappointing. The compounds
were all less potent and less selective than their sulfone
analogues.
Before undergoing in vivo studies, both the chemical
and metabolic stability of 10 were examined. The
hydrochloride salt of 10 was treated with 0.1 N HCl (pH
1), and after several days there was no evidence of ketal
hydrolysis. This ketal stability to acid was likely due to
the electron withdrawing p-sulfone substituent. In addi-
tion, 10 showed stability in rat and human microsomes.9
In vivo, 10 (po dosing) was studied in microdialysis and
passive-avoidance response (PAR) assays. In the micro-
dialysis experiment, ACh levels in the rat striatum were
sampled via a microdialysis probe and measured by
HPLC/ECD.10 A detectable level of ACh was achieved
by addition of neostigmine, an acetylcholinesterase
inhibitor. After attaining a stable baseline, 10 was dosed
orally, and ACh levels were monitored over a period of
approximately 2 h. As shown in Figure 1, compound 10
promoted a prolonged release of ACh in the rat brain,
demonstrating blood±brain barrier penetration and M2
potency. The ACh levels of 10 surpass that of II in a
similar study,3 which implies that the ketal derivative
has improved metabolic stability, CNS levels, and/or
receptor binding kinetics over the original lead.
Figure 2. PAR data. Compound 10, po, methyl cellulose in water.
In conclusion, 10 improved upon leads such as I and II
not only by improving M2 selectivity versus other mus-
carinic receptors, but also by replacing metabolically
unstable benzylic moieties with the chemically and
metabolically stable ketal. Furthermore, 10 was shown
to improve cholinergic system activity and memory in
orally dosed in vivo microdialysis and PAR studies.
Ongoing studies in this promising ketal series for the
treatment of AD will be described in future publications.
For the PAR experiment, 10 was dosed orally to a
young rat before testing in a reference memory para-
digm (very young rats exhibit memory de®cits when
tested in such an assay).11 The rat was given a training
session in a dark/light box, in which the rat had a nat-
ural aversion to the light chamber. Entry to the dar-
kened area was followed by a mild electrical shock to
the feet. In order to avoid the electrical shock, the rat
Acknowledgements
The authors wish to thank Drs. John Clader, William
Greenlee, Michael Green, and Yuguang Wang for

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C. D. Boyle et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2727±2730
helpful discussions. We also thank Dr. Pradip Das for
obtaining analytical data.
ity. For further information, see the following articles and
references cited therein: Doller, D.; Chackalamannil, S.;
Czarniecki, M.; McQuade, R.; Ruperto, V. Bioorg. Med.
Chem. Lett. 1999, 9, 901, and Kozikowski, A. P.; Fauq, A. H.;
Miller, J. H.; McKinney, M. Bioorg. Med. Chem. Lett. 1992, 2,
797.
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