Design and synthesis of dual inhibitors of acetylcholinesterase and serotonin transporter targeting potential agents for Alzheimer's disease.

Article abstract of DOI:10.1021/ol026418e

Highly efficient acetylcholinesterase (AChE) and serotonin transporter (SERT) dual inhibitors, (S)-4 and (R)-13 were designed and synthesized on the basis of the hypothetical model of AChE active site. Both compounds showed potent inhibitory activities against AChE and SERT. [structure: see text]

Full text of DOI:10.1021/ol026418e

ORGANIC
LETTERS
2002
Vol. 4, No. 20
3359-3362
Design and Synthesis of Dual Inhibitors
of Acetylcholinesterase and Serotonin
Transporter Targeting Potential Agents
for Alzheimer’s Disease
Hiroshi Kogen,* Narihiro Toda, Keiko Tago, Shinji Marumoto, Kazuko Takami,
Mayuko Ori, Naho Yamada, Kazuo Koyama,† Shunji Naruto,† Kazumi Abe,‡
Reina Yamazaki,‡ Takao Hara,‡ Atsushi Aoyagi,‡ Yasuyuki Abe,‡ and
Tsugio Kaneko‡
Exploratory Chemistry Research Laboratories, Research Information Department, and
Neuroscience and Immunology Research Laboratories, Sankyo Co., Ltd., 1-2-58
Hiromachi, Shinagawa-ku, Tokyo 140-8710, Japan
hkogen@shina.sankyo.co.jp
Received June 25, 2002
ABSTRACT
Highly efficient acetylcholinesterase (AChE) and serotonin transporter (SERT) dual inhibitors, (S)-4 and (R)-13 were designed and synthesized
on the basis of the hypothetical model of AChE active site. Both compounds showed potent inhibitory activities against AChE and SERT.
Alzheimer’s disease (AD), the most common cause of
dementia, is a neurodegenerative disorder characterized by
progressive deterioration of memory and cognition.1 En-
hancement of central cholinergic function by inhibition of
acetylcholinesterase (AChE) is, so far, the only clinically
effective approach for treatment of AD.2 However, the
clinical usefulness of marketed AChE inhibitors (AChEIs)
is limited mainly by adverse effects on peripheral organs.3
AD patients often exhibit psychiatric symptoms such as
irritability, anxiety, and depression. Depression in AD
patients has been successfully treated with inhibitors of
serotonin transporter (SERT),4 antidepressants that lack
anticholinergic action. Thus, it is anticipated that combining
SERT and AChE inhibitory activity could offer greater
therapeutic benefits, because the antidepressive effect of
SERT inhibitors might reduce a demand on cholinergic
systems in the brain to ameliorate cognitive deficits. These
dual inhibitors would be a novel class of anti-AD drugs more
effective in alleviating the symptoms of AD than the known
AChE inhibitors. Two groups have studied AChEIs that
possess serotonin transporter inhibitory activity.5 The reported
† Research Information Department.
‡ Neuroscience and Immunology Research Laboratories.
(1) (a) Wurtman, R. J. Sci. Am. 1985, 252, 48. (b) Alzheimer’s Diseases
Problems, Prospects and PerspectiVes; Altman, H. J., Ed.; Plenum Press:
New York, 1987.
(2) (a) Enz, A.; Amstutz, R.; Boddeke, H.; Gmelin, G.; Malanowski, J.
Prog. Brain Res. 1993, 98, 431. (b) Millard, C. B.; Broomfield, C. A. J.
Neurochem. 1995, 64, 1909.
(3) (a) Benzi, G.; Moretti, A. Eur. J. Pharmacol. 1998, 346, 1. (b)
Giacobini, E. Jpn. J. Pharmacol. 1997, 74, 225.
(4) Lyketsos, C. G.; Sheooard, J. M.; Steele, C. D.; Kopunek, S.;
Steinberg, M.; Baker, A. S.; Brandt, J.; Rabins, P. V. Am. J. Psychiatry
2000, 157, 1686.
10.1021/ol026418e CCC: $22.00 © 2002 American Chemical Society
Published on Web 09/05/2002

compounds showed potent AChE inhibitory activities and
moderate SERT inhibitory activities in vitro. No compound
to date shows simultaneous activation of both the cholinergic
and serotonergic nervous systems in the brain following oral
administration, probably because of inappropriate combina-
tion of activities. We describe here the design and synthesis
of highly efficient dual inhibitors of AChE and SERT.
On the basis of the X-ray structure of the complex of
AChE and donepezil6 and molecular modeling studies,7 we
proposed a hypothetical model for inhibitor interaction at
the active site of AChE, which involves four binding sites
(Figure 1). To design the dual inhibitor, we selected one of
The type B ring-closed compound was also designed to
explore the effect of conformational restriction (Scheme 1).
Scheme 1. Design of AChE-SERT Dual Inhibitors
We synthesized and evaluated various type A derivatives
and found that most have inhibitory activities against both
AChE and SERT. Optimization of the carbamate group, the
tether-length,10 and the R′′ substituent showed 4-nitrophenoxy
ether derivative 4 (m ) 1, X ) O) to be the best type A
dual inhibitor. Both optical isomers of compound 4 were
synthesized as shown in Scheme 2. The primary amine of
Figure 1. Hypothetical model of AChE active site with proposed
pharmacophore.
Scheme 2a
the marketed AChEIs, rivastigmine,8 as a lead compound.
Rivastigmine possesses three elements that are able to bind
to the proposed binding sites of AChE: a carbamate moiety
to the catalytic triad site, an amino moiety to the ionic site,
and an aromatic ring to the hydrophobic binding site A.
Focusing on the fourth binding site of our model, we chose
fluoxetine9 as the pharmacophoric element corresponding to
the hydrophobic binding site B. Fluoxetine has potent
inhibitory activity against SERT and possesses an ethylamine
moiety that could be overlapped with that of rivastigmine.
Therefore, the type A compound designed by hybridization
of these two compounds at the common ethylamine moiety
was expected to be a dual inhibitor of AChE and SERT.
a (a) Boc2O, THF, rt, 72%; (b) Me2NCOCl, K2CO3, DMF, rt,
96%; (c) LiAlH4, -78 to 0 °C, 97%; (d) DEAD, PPh3, 4-nitro-
phenol, THF, rt; (e) NaH, MeI, DMF, 0 °C to rt, 77% (2 steps); (f)
2 N HCl-AcOEt, rt, 100%.
(5) (a) Mackenna, M. T.; Proctor, G. R.; Young, L. C.; Harvey, A. L. J.
Med. Chem. 1997, 40, 3516. (b) Hirai, K.; Kato, K.; Nakayama, T.; Hayako,
H.; Ishihara, Y.; Goto, G.; Miyamoto, M. J. Pharmocol. Exp. Ther. 1997,
280, 1261.
(6) (a) Kryger, G.; Silman, I.; Sussman, J. L. J. Physiol. Paris 1988, 92,
191. (b) Sugimoto, H. Chem. Rec. 2001, 1, 63.
(7) Kaur, J.; Zhang, M.-Q. Curr. Med. Chem. 2000, 7, 273.
(8) (a) Enz, A.; Amstutz, R.; Hofmann, A.; Gmelin, G.; Kelly, P. H.
Klin. Pharmakol. 1989, 2, 271. (b) Sramek, J. J.; Anand, R.; Wardle, T. S.;
Irwin, P.; Hartman, R. D.; Cutler, N. R. Life Sci. 1993, 58, 1201. (c) Spencer,
C. M.; Noble, S. Drugs Aging 1998, 13, 391. (d) Jann, M. W. Pharmaco-
therapy 2000, 20, 1.
known (S)-111 was Boc-protected, followed by treatment with
dimethylcarbamyl chloride to afford (S)-2. Reduction of the
ester group of (S)-2 with LiAlH4 gave primary alcohol (S)-
3. The alcohol (S)-3 was reacted under Mitsunobu conditions
(10) There is tether-length dependence to AChEI potency enhancement.
The compounds possessing a longer tether showed stronger AChEI activities;
however, only the type A compounds possessing adequate tether length (m
) 1) showed SERT inhibitory activities.
(9) Golstein, J.; Schreiber, S.; Velkeniers, B.; Vanhaelst, L. Eur. J.
Pharmacol. 1983, 91, 239.
(11) Davies, S. G.; Ichihara, O. Tetrahedron: Asymmetry 1991, 2, 183.
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Org. Lett., Vol. 4, No. 20, 2002

with p-nitrophenol, followed by methylation of the Boc-
protected amine with iodomethane. Finally, deprotection of
the Boc group with 2 N HCl in ethyl acetate provided (S)-4.
Enantiomer (R)-4 was prepared by the same method starting
from (R)-1.11
and the resulting amino group was subsequently protected
with Boc to yield (R)-8 (99% ee).16 Compound (R)-8 was
treated with dimethylcarbamyl chloride and reduced with
LiAlH4 to provide alcohol (R)-9. The alcohol (R)-9 was
reacted with p-nitrophenol under Mitsunobu conditions.
Treatment with concentrated HCl and Boc protection of the
amino group afforded phenol (R)-10, which was converted
to the aryl triflate by treatment with trifluoromethanesulfonic
anhydride. The resulting aryl triflate was coupled with
tributylvinylstannane by a palladium-catalyzed Stille reac-
tion17 to afford the corresponding styrene compound. N-
Allylation of the obtained styrene compound was accom-
plished by treating with allyl bromide and NaH to give (R)-
11. Ring-closing olefin metathesis of (R)-11 with 10 mol %
of Grubbs catalyst 1218 gave the desired seven-membered
ring product in high yield. Deprotection of the Boc group in
the resulting cyclic compound by 2 N HCl in ethyl acetate,
followed by Eshweiler-Clarke methylation, provided (R)-13.
Enantiomer (S)-13 was prepared by the same method by
using chiral amine (R)-7.15
Compound (S)-4 showed potent inhibitory activities against
both AChE (IC50 ) 101 nM) and SERT (IC50 ) 42 nM).
However, though enantiomer (R)-4 showed similar potency
against AChE (IC50 ) 143 nM), SERT inhibition activity
(IC50 ) 135 nM) was slightly diminished compared to that
of (S)-4.12
Type B compound (R)-13 (n ) 1)13 was synthesized as
shown in Scheme 3. The phenol group of 514 was protected
Scheme 3a
Surprisingly, the inhibitory activities of (R)-13 against both
AChE and SERT (IC50 ) 14 and 6 nM, respectively) were
extremely enhanced as compared to those of 4 (Table 1).
Table 1. Inhibitory Actibities of 4 and 13 against AChE
a (a) MOMCl, i-Pr2NEt, CH2Cl2, 0 °C, 79%; (b) (EtO)2P(O)CH2-
CO2Et, NaH, THF, 0 °C, 99%; (c) (i) (S)-7, n-BuLi, THF, -78
°C; (ii) H2, 20% Pd(OH)2/C, MeOH-H2O-AcOH (20:2:1), rt, 67%
(2 steps); (d) Boc2O, Et3N, MeOH, rt, 97%; (e) Me2NCOCl, K2CO3,
DMF, rt, 97%; (f) LiAlH4, THF, -50 to 0 °C, 82%; (g)
4-nitrophenol, DEAD, PPh3, THF, rt; (h) concentrated HCl, MeOH,
rt, 68% (2 steps); (i) Boc2O, Et3N, MeOH, rt, 85%; (j) (i) Tf2O,
pyridine, CH2Cl2, 0 °C; (ii) (CH2dCH)SnBu3, cat. Pd(PPh3)4, LiCl,
dioxane, 100 °C, 86% (2 steps); (k) allyl bromide, NaH, DMF, 0
°C to rt, 86%; (l) 12 (10 mol %), CH2Cl2 (15mM), 45 °C, 96%;
(m) 2 N HCl-AcOEt, rt, 96%; (n) (i) 37% HCHO aq., HCO2H,
80 °C; (ii) 2 N HCl-AcOEt, rt, 78% (2 steps).
Although both enantiomers of 4 showed similar AChE
inhibitory activities, (R)-13 showed inhibitory activity against
AChE that was extremely stronger than that of (S)-13. We
supposed that in the case of the ring closure compound, the
restriction of the conformational flexibility of the amine
moiety induced the appropriate binding to AChE.19
These compounds hardly inhibited butyrylcholinesterase
and choline acetyltransferase (IC50’s > 100 µM) and showed
weak affinities for transporters of norepinephrine and
with a methoxymethyl group (MOM), and a Horner-
Emmons reaction provided 6. The ester 6 was subjected to
the chiral amination method that was reported by Davies,15
(12) By our assay procedures, rivastigmine showed weak inhibitory
activity against AChE (IC50 ) 11 µM). Donepezil showed potent inhibitory
activity only against AChE (IC50 ) 10 nM), and fluoxetine showed
inhibitory activities only against SERT (IC50 ) 180 nM).
(13) Seven-membered ring compound (m ) 1, n ) 1, X ) O) showed
the most potent AChE and SERT inhibitory activities.
(14) Mendelson, W. L.; Holmes, M.; Dougherty, J. Synth. Commun. 1996,
26, 593.
(16) The determination of optical purity and absolute configuration of
(R)-8 are included in Supporting Information.
(17) For a recent review, see: Farina, V.; Krishnamurthy, V.; Scott, W.
J. Org. React. 1997, 50, 1.
(18) For a recent review, see: (a) Grubbs, R. H.; Chang, S. Tetrahedron
1998, 54, 4413. (b) Fu¨rstner, A. Angew. Chem., Int. Ed. 2000, 39, 3013.
(c) Yet, L. Chem. ReV. 2000, 100, 2963.
(19) Since there is no information on the structure of SERT, the reason
of the opposite stereochemical preference is unclear.
(15) Davies, S. G.; Walters, A. S. J. Chem. Soc., Perkin Trans. 1 1994,
1129.
Org. Lett., Vol. 4, No. 20, 2002
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dopamine (IC50’s > 10 µM). In addition, (S)-4 and (R)-13
had weak affinities for existing neurotransmitter receptors
such as serotonin, norepinephrine, and dopamine (IC50’s >
10 µM). Compound (S)-4 shows more potent inhibitory
activities than (R)-13 against AChE in the brain following
an oral administration (Figure 2).20,21 The difference in
efficiency between these two compounds could be attributed
to the difference in absorption from the intestine and/or
penetration into the brain.
In summary, we designed and synthesized AChE-SERT
dual inhibitors 4 and 13 on the basis of a hypothetical model
of the AChE active site. Additionally, ring-closing compound
(R)-13 showed extremely potent inhibitory activities against
both AChE and SERT. Compound (S)-4 is the first com-
pound that can facilitate both the cholinergic and serotonergic
transmission. Further pharmacological evaluation of these
compounds targeting Alzheimer’s disease is underway.
Figure 2. Inhibition of AChE and SERT in the brain following
an oral administration in mice (ex vivo assay).
(20) The brain of mouse was isolated 1 h after an oral administration of
compounds, and AChE and/or SERT activity were measured. Each symbol
represents the mean of five mice. The doses (95% confidence interval) of
(S)-4 necessary for 50% inhibition of AChE and SERT in the brain were
determined as 13 (11-15) and 2.4 (1.9-2.9) mg/kg, p.o., respectively.
Details will be reported soon.
(21) The SERT inhibition of R-13 was not determined because a low
penetration of the compound into the brain was predicted from the ex vivo
study of AChE inhibition.
Supporting Information Available: Synthetic procedures
and characterization of new compounds, in vitro assay of
(S)-4 and (R)-13, and the determination of optical purity and
absolute configuration of (R)-8. This material is available
free of charge via the Internet at http://pubs.acs.org.
OL026418E
3362
Org. Lett., Vol. 4, No. 20, 2002