Discovery of the α7 nicotinic acetylcholine receptor agonists. (R)-3′-(5-chlorothiophen-2-yl)spiro-1-azabicyclo[2.2.2]octane-3, 5′-[1′,3′]-oxazolidin-2′-one as a novel, potent, selective, and orally bioavailable ligand

Article abstract of DOI:10.1021/jm049188d

Recent advances in molecular biology suggest that neuronal nicotinic acetylcholine receptors play important roles in the central nervous system (CNS). Of these receptors, the α7 group has recently attracted interest for its CNS-related actions and is looked to as a potential new class of pharmacological targets for cognition, schizophrenia, sensory gating, and anxiety. In the course of a research program aimed at the discovery of α7 receptor agonists with high affinity, subtype selectivity, and good pharmacokinetic profile, we discovered (R)-3′-(5-chlorothiophen-2-yl) spiro-1-azabicyclo[2.2.2]octane-3,5′-[1′,3′] oxazolidin-2′-one (25). Compound 25 has potent binding affinity (K _i = 9 nmol/L) and good selectivity toward the other nicotinic subtypes (α4β2 and α1β2γδ) and has been found in pharmacokinetic evaluation to have good oral bioavailability and brain permeability.

Full text of DOI:10.1021/jm049188d

2678
J. Med. Chem. 2005, 48, 2678-2686
Discovery of the r7 Nicotinic Acetylcholine Receptor Agonists.
(R)-3′-(5-Chlorothiophen-2-yl)spiro-1-azabicyclo[2.2.2]octane-3,5′-[1′,3′]-
oxazolidin-2′-one as a Novel, Potent, Selective, and Orally Bioavailable Ligand
Ryo Tatsumi,* Masakazu Fujio, Hiroyuki Satoh, Jiro Katayama, Shin-ichi Takanashi, Kenji Hashimoto,‡ and
Hiroshi Tanaka
Pharmaceuticals Research Unit, Research & Development Division, Mitsubishi Pharma Corporation,
1000 Kamoshida-cho, Aoba-ku, Yokohama, Kanagawa 227-0033, Japan
Received October 8, 2004
Recent advances in molecular biology suggest that neuronal nicotinic acetylcholine receptors
play important roles in the central nervous system (CNS). Of these receptors, the R7 group
has recently attracted interest for its CNS-related actions and is looked to as a potential new
class of pharmacological targets for cognition, schizophrenia, sensory gating, and anxiety. In
the course of a research program aimed at the discovery of R7 receptor agonists with high
affinity, subtype selectivity, and good pharmacokinetic profile, we discovered (R)-3′-
(5-chlorothiophen-2-yl)spiro-1-azabicyclo[2.2.2]octane-3,5′-[1′,3′]oxazolidin-2′-one (25). Com-
pound 25 has potent binding affinity (Ki ) 9 nmol/L) and good selectivity toward the other
nicotinic subtypes (R4â2 and R1â2γδ) and has been found in pharmacokinetic evaluation to
have good oral bioavailability and brain permeability.
Introduction
Neuronal nicotinic acetylcholine receptors (nAChRs)
are pentameric cation-selective ion channels that are
formed from various combinations of alpha (R2-R10)
and beta (â2-â4) subunits.1,2 In heterologous expression
systems, most neuronal nAChRs are constructed from
R and â subunits, and the combinations are associated
with functional nAChRs.3-6 The contributions of R5, R6,
and â3 are less well understood.7-10 On the other hand,
only R7, R8, and R9 can form homo-oligomeric receptors
that are inhibited by R-bungarotoxin (R-BTX).11-14 The
most prevalent subtypes are associated with the R7 and
R4â2 subunits that are strongly expressed during brain
Figure 1. Structure of previously described R7 nicotinic
development15-17 and have been implicated in memory,
acetylcholine receptor agonists and compounds identified in
attention, and information processing.18,19
our laboratory.
Schizophrenia is a neuropsychiatric disorder that
brains of schizophrenic patients have been reported.24,25
affects about 1% of the population worldwide and
The R7 nicotinic acetylcholine receptor agonists are
therefore thought to be a potential pharmacological
target for CNS disorders including neurocognitive dys-
function.
frequently demonstrates a strong hereditary component.
The condition is characterized by positive symptoms
(delusions, hallucinations), negative symptoms (apathy,
anhedonia, avolition, amotivation), and neurocognitive
dysfunction.20 Abnormality in P50 auditory-evoked po-
tential gating is an endophenotype associated with
schizophrenia. Biochemical and genetic studies have
suggested that the R7 receptor is involved in this
sensory gating deficit.21 Moreover, intracerebroven-
tricular injections of R-BTX, an R7 (and R8, R9) receptor
antagonist, disrupt hippocampal auditory gating,22 while
recent studies also indicate a correlation between the
R7 receptor and several aspects of schizophrenia,23 and
decreased levels of this receptor in the postmortem
Several R7 receptor agonists have been reported in
the literature and show prospects in terms of therapeu-
tic potential (Figure 1). For example, 3-(2,4-dimethoxy-
benzylidene)anabaseine (GTS-21, 1) is a partial agonist
of the R7 receptor.26 It has been reported to be a
“functionally” selective R7 receptor agonist, it but pos-
sesses a higher affinity for the R4â2 receptor, toward
which it acts as an antagonist,27 and fails to show a
satisfactory pharmacokinetic (PK) profile in the areas
of bioavailability and brain permeability. (-)-Spiro-1-
azabicyclo[2.2.2]octane-3,5′-oxazolidin-2′-one (AR-R-
17779, 2), meanwhile, is the first subtype-selective
agonist of the R7 receptor which demonstrates a high
degree of selectivity for the R7 subtype of nAChRs.28
Compound 2, however, remains at the preclinical trial
stage. Accordingly, there continues to be interest in new
* To whom correspondence should be addressed. Phone:
+81-(0)45-963-4624. Fax: +81-(0)45-963-4211. E-mail: Tatsumi.Ryo@
mg.m-pharma.co.jp.
‡ Present address: Department of Psychiatry, Graduate School of
Medicine, Chiba University, 1-8-1 Inohana, Chuou-ku, Chiba, 260-
8670, Japan.
10.1021/jm049188d CCC: $30.25 © 2005 American Chemical Society
Published on Web 02/25/2005

R7 Nicotinic Acetylcholine Receptor Agonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 7 2679
Figure 2. Synthetic strategy to improve the pharmacokinetic profile of R7 nicotinic acetylcholine receptor agonists.
compounds for clinical development. As previously
established, (+)-3-[2-(benzo[b]thiophen-2-yl)-2-oxoethyl]-
1-azabicyclo[2.2.2]octane (3) shows high affinity for the
R7 receptor (Ki ) 65 nmol/L).29 Further modification led
to a discovery of (S)-3-(benzo[b]thiophen-2-ylmethoxy)-
1-azabicyclo[2.2.2]octane (4) (Ki ) 13 nmol/L). Although
compound 4 was not detected in plasma following oral
administration (rat, 1 mg/kg), we estimated that this
defect may be attributable to its ether part, which might
constitute a metabolic point. Furthermore, it is com-
monly suggested that one of the key factors controlling
bioavailability is molecular flexibility (the number of
rotatable bonds). On the basis of the structure of
compound 2 and this hypothesis, a strategy was devised
to fix the ether part with a cyclic structure possessing
potent binding affinity compared with compound 4 to
improve pharmacokinetic profiles (Figure 2). We report
here the design, synthesis, structure-activity relation-
ships, and PK profiles of the series of compounds leading
to the discovery of (R)-3′-(5-chlorothiophen-2-yl)spiro-
1-azabicyclo[2.2.2]octane-3,5′-[1′,3′]oxazolidin-2′-one 25,
the first potent and selective agonist of the R7 nicotinic
acetylcholine receptor to display good oral bioavailability
and brain permeability.
Results and Discussions
Previous efforts in our laboratories had identified two
compounds, 3 and 4, with high binding affinity
(Ki ) 65 and 13 nmol/L, respectively) for the R7 receptor,
as determined by R-bungarotoxin binding inhibition. We
first examined the effect of a spiro-carbamate bearing
various bicyclic aromatic parts on binding affinity to the
R7 receptor. The results are summarized in Table 1. The
benzo[b]thiophene derivative 7, which has the same aryl
part as compound 4, showed moderate affinity. In
contrast, the (S)-isomer 8 had no affinity for the R7
receptor. Introduction of a â-naphthyl moiety, also
known as an isostere of the benzothiophene part,
increased the affinity of compounds 7 and 8, again with
lower affinity observed for compound 10 than compound
9. A clear stereogenic preference can be seen for the (R)
versus the (S) enantiomer of compounds 7-10, which
demonstrates the necessity for the (R)-configuration to
achieve binding affinity. These observations were sup-
ported by previous finding for compound 2.28 (R)-
Compounds were therefore assessed for further evalu-
ation. Although moderate affinity was observed in
compounds 7 and 9, which have a benzo[b]thiophene
and a â-naphthyl, respectively, as the bicyclic aromatic
part, the conversion of the bicyclic part with quinoline
(11), benzothiazole (12), benzoxazole (13), and benzo-
furan (14) led to reduction or loss of the affinity for the
R7 receptor. Some monocyclic aromatic parts were also
evaluated for R7 binding affinity to evaluate the influ-
ences of steric and electric factors. Compound 15, which
has a benzene ring as the aromatic part, showed no
binding affinity. The replacement of the phenyl ring
with a pyridine (16) led to a similar result. Interestingly,
while the binding affinity was strongly enhanced com-
pared to other monocyclic derivatives by the introduc-
tion of a thiophen-2-yl moiety (17, Ki ) 80 nmol/L), the
thiophen-3-yl derivative 18 had no binding affinity. This
finding suggests that the presence of a heteroatom, such
as nitrogen or oxygen, is not conducive to R7 binding
affinity, which may be affected by some electronic
interaction with the R7 receptor. Nevertheless, the
phenyl derivative 15, which possesses no heteroatom
for the aryl moiety, showed no affinity. Moreover, potent
affinity was observed in the thiophen-2-yl derivative 17,
despite the absence of affinity in the thiophen-3-yl
derivative 18. There may thus be unknown interactions
with the R7 receptor.
Chemistry
At the beginning, our synthetic strategy was based,
as previously reported, on the preparation of compound
2 and (R)-5.28 A previous study suggested that methyl-
ation at the nitrogen atom of the carbamate moiety of
compound 2 attenuates the affinity for the R7 receptor.
We estimated, however, from the structure of compound
4 that an aryl moiety would be permissible. As shown
in Scheme 1, the coupling of 2 with various aryl halides
in the presence of potassium carbonate and copper(I)
iodide or using sodium hydride gave compounds 7, 9,
11-14, and 16-25 in moderate yields. In a similar
manner, compounds 8 and 10 were obtained from (R)-5
and 15 from (rac)-6. Compound 27 was obtained by
heating the bromide 26, which was prepared by bromin-
ation of compound 17, with Zn(CN)2 and Pd(PPh3)4.
Friedel-Crafts acylation of compound 17 in the pres-
ence of acetyl chloride and aluminum chloride afforded
compound 28. The resulting acetyl derivative 28 was
subsequently treated with NaBH4 to afford the alcohol
derivative 29 as a single isomer (absolute configuration
was not determined).

2680 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 7
Tatsumi et al.
Scheme 1. Synthesis of 1-Azabicyclo[2.2.2]octane Derivativesa
a Reagents: (a) Ar-Br, K2CO3, CuI, DMF. (b) Ar-Br (Cl), NaH, DMF. (c) NBS, DMF. (d) Zn(CN)2, Pd(PPh3)4, DMF. (e) AcCl, AlCl3,
nitrobenzene. (f) NaBH4, MeOH.
As the thiophen-2-yl derivative 17 showed higher
affinity (Ki ) 80 nmol/L) than other monocyclic deriva-
tives, the effect on R7 binding affinity of a substituent
on the thiophene ring was evaluated. Table 2 shows the
results. Introduction of a methyl moiety at the 3- or
4-position of the thiophene ring resulted in loss of the
affinity (19 and 20). Surprisingly, however, greatly
enhanced affinity was observed in the 5-methyl thiophene
derivative 21 (Ki ) 12 nmol/L). The dimethyl thiophene
derivative 22, meanwhile, had no binding affinity. These
results suggest that steric effect is important for binding
affinity, as there may be a sterically restricted binding
pocket for the receptor as observed with compounds 17-
22. A preference in relation to the position of the
substituent can also be seen, with the 5-position being
important for achieving binding affinity. Subsequently,
various substituents at the 5-position of the thiophene
ring were examined. The ethyl and isopropyl derivatives
23 and 24 had slightly enhanced affinity compared to
compound 17. To investigate the electronic effect on the
thiophene ring, introduction of a halogen atom to the
thiophene ring as an electron-withdrawing group was
investigated and found to result in an enhancement of
affinity to a level higher than that of the previously
identified compound 4 (25, Ki ) 9 nmol/L; 26, Ki ) 4
nmol/L). Substitution of the cyano group in compound
27 slightly reduced affinity relative to compound 17,
while the acetyl group in compound 28 reduced affinity
2-fold. A similar reduction of the affinity was observed
in the alcohol derivative 29. These findings suggest not
only an electric but also a steric effect on binding to the
R7 receptor. Nevertheless, the results indicate clearly
that a sulfur atom is preferred as the component for the
aromatic part to the other heteroatoms and that 5-posi-
tion is sensitive for R7 binding affinity.
activities for these compounds. Nicotine, an endogenous
ligand for the R7 receptor, and compound 2 were studied
at 1, 3, 10, 30, 100, 300, and 1000 µmol/L, while
compound 1 was studied at 1, 3, 10, 30, 100, and
300 µmol/L. Consistent with previous knowledge, the
nicotine-induced relative inward current toward
10 mmol/L choline was dose-dependent. At the concen-
trations studied, compound 2 displayed greater potency
than nicotine at all concentrations in stimulating the
relative inward current. As previously reported, com-
pound 1 showed a dose-response curve indicating a
profile as a partial agonist with lower efficacy than
compound 2, which is known as a full agonist. Our
selected compound 25 was studied at 0.1, 0.3, 1, 3, 10,
30, and 100 µmol/L. It was observed that compound 25
displayed a similar dose-response curve to that of
compound 1, also indicating it to be a partial agonist.
It should be noted that compound 25 not only showed
agonistic activity at 0.1 µmol/L but also displayed
greater efficacy than compound 1 at all doses. Although
it is unclear whether a full or partial agonist of the R7
receptor is more valid from the therapeutic viewpoint,
the partial agonist compound 1 did, as previously
reported,30 exhibit effect on some cognition-measuring
behaviors. Compound 25 would therefore also seem to
have potential in the treatment of cognitive dysfunc-
tions.
Besides measurement of the R7 binding affinity and
agonistic activity of compound 25, additional pharma-
cological evaluations were required to determine its
selectivity compared to that of other receptors to estab-
lish relative benefit in the treatment of chronic neuro-
psychiatric disorders such as neurocognitive dysfunc-
tion. Binding affinities were therefore determined for
the other nicotinic subtypes (R4â2 and R1â2γδ). These
are displayed in Table 3. For comparison, compounds 1
and 2 were also examined. None of the compounds
showed affinity for the R1â2γδ receptor, but compound
1 possessed strong affinity for the R4â2 receptor, greater
than that for the R7 receptor. It was observed that
We selected compound 25 for further evaluation. To
evaluate the agonistic activities of compounds 25, 1, 2,
and nicotine, we carried out electrophysiological mea-
surement of the R7 receptor-mediated response using
cultured hippocampal neurons, in which the efficacy was
assessed by measurement of the relative inward current
toward 10 mmol/L choline. Figure 3 shows the agonistic

R7 Nicotinic Acetylcholine Receptor Agonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 7 2681
Table 1. Binding Affinities of Oxazolidinone Derivatives with
Various Aryl Moieties
Figure 3. Agonistic activities of nicotine and compounds 1,
2, and 25 on R7 nicotinic receptor in cultured hippocampal
neurons. Data are represented as mean ( SEM (N ) 2-4).
Table 3. Selectivity of Compounds 25, 1, and 2 Relative to
Other Nicotinic Subtypes
binding affinity (Ki; nmol/L)a of nicotinic subtypes
compd no.
R7b
R4â2c
R1â2γδd
25
1
2
9
220
340
1800
84
>10000e
>10000e
>10000e
>10000e
a Tests were performed in two experiments. Among Ki values
in reproducibility runs, the assay showed less than 20% variability.
b R-Bungarotoxin binding. c Cytisine binding. d R-Bungarotoxin
binding. e IC50 value.
a R-Bungarotoxin binding. Tests were performed in two experi-
ments. Among Ki values in reproducibility runs, the assay showed
less than 20% variability. b IC50 value.
for compound 25 (4640 ( 570 ng h/mL) was ap-
proximately comparable with the value for compound
2. Although the rat oral peak plasma levels were not
remarkable, it was notable that compound 25 produced
longer duration of plasma concentration (mean
T1/2 ) 3.13 ( 1.37 h, iv) than compounds 1 and 2. The
oral bioavailability (BA) of compounds 25 and 2 was
very satisfactory in both cases. Most importantly,
although compounds 1 and 2 showed low or moderate
brain permeability (CB/CP ) <0.3 and 0.76 ( 0.23,
respectively), compound 25 was found to have a high
brain permeability (CB/CP ) 59 ( 9.8), which is a
desirable profile for a drug to treat CNS (central nervous
system) disorders. Compound 25 is thus a compound
with high oral bioavailability and brain penetration.
Finally, to assess the effect of compound 25 on
cognitive enhancement, auditory sensory gating was
investigated in a rat model. Auditory sensory gating, a
measurement of the ability to depress the evoked
response to the second of two auditory stimuli (condi-
tioning-testing paradigm expressed as T/C ratio), is
impaired in humans with schizophrenia, and involve-
ment of R7 receptor dysfunction in the deficit has been
suggested.31,32 In our research programs, we have used
the known NMDA antagonist MK-801 in a rat model of
auditory gating dysfunction.33 In this model, the typical
antipsychotic haloperidol had no improving effect on
auditory gating dysfunction (data not shown), but the
atypical antipsychotic clozapine showed significant ame-
lioration, as shown in Figure 4a. In patients with
schizophrenia, cognitive dysfunction is not affected by
medication with typical antipsychotics, but as previously
reported, atypical antipsychotics such as clozapine
alleviate the deficit.34,35 An MK-801-induced auditory
sensory gating model was therefore used to assess the
ability of the R7 agonist to correct cognitive dysfunction.
Table 2. Binding Affinities of (R)-Oxazolidinone Derivatives
with Various Substituents on the Thiophene Ring
compd
no.
R7a
(Ki; nmol/L)
R
17
19
20
21
22
23
24
25
26
27
28
29
80
>1000b
>1000b
12
3-methyl
4-methyl
5-methyl
4,5-dimethyl
5-ethyl
>1000b
21
5-isopropyl
5-chloro
65
9
4
5-bromo
5-cyano
85
5-acetyl
190
5-1′-hydroxyethyl
145
a R-Bungarotoxin binding. Tests were performed in two experi-
ments. Among Ki values in reproducibility runs, the assay showed
less than 20% variability. b IC50 value.
compounds 25 and 2 showed good selectivity toward the
other nicotinic subtypes, with compound 25 displaying
200-fold selectivity.
Subsequently, we selected compound 25 for further
evaluation of pharmacokinetic profiles in rats, as shown
in Table 4. Compound 25 was examined at doses of 1
and 10 mg/kg (po), while compounds 1 and 2 were
administered only at the dose of 10 mg/kg (po). Com-
pounds 1 and 2 showed AUC0-24 values of 453 ( 44 and
5390 ( 660 ng h/mL, respectively. The value for
compound 1 was only twice the AUC value for compound
25 at 1 mg/kg (po). In turn, the value at 10 mg/kg (po)

2682 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 7
Tatsumi et al.
Table 4. Pharmacokinetic Profiles of Compounds 25, 1, and 2 in Rats (N ) 4)'
compd no.
po dose (mg/kg)
AUC0-24ha,f (ng h/mL)
Cmaxb,f (ng/ mL)
T1/2c,f (h)
BAd (%)
brain permeabilitye,f (CB/CP)
25
1
10
10
10
224 ( 25
4640 ( 570
453 ( 44
56 ( 17
997 ( 87
52
108
14
3.13 ( 1.37
0.47 ( 0.08
0.75 ( 0.14
59 ( 9.8
<0.3
0.76 ( 0.23
1
2
138 ( 39
5390 ( 660
1580 ( 320
103
a Estimated area under plasma concentration time curve after oral dosing. b Maximum plasma concentration after oral dosing. c Half-
life at 1 mg/kg, iv. d Oral bioavailability. e CB/CP ) concentration in brain/concentration in plasma. Rats (N ) 3) were given a single dose
(10 mg/kg, po) of either compound 25 and compound 1 or a single dose (1 mg/kg, sc) of compound 2. The CB/CP values were calculated
2 h after administration for compounds 25 and 1 and 1 h after for compound 2. f Data are shown as mean ( SD.
Figure 4. Effects of clozapine (30 mg/kg, po) and compound 25 (10 mg/kg, po) on MK-801-induced sensory gating deficit in rats
(N ) 6).
Figure 4b shows the effect of compound 25, which was
found to reduce MK-801-induced auditory gating defi-
cit.Significant differences were observed in oral admin-
istration at doses of 10 and 30 mg/kg (data not shown
for 30 mg/kg). This finding suggests that the R7 agonists
have the potential to improve sensory gating deficit, and
indeed, it is reported that nicotine improves sensory
gating deficit in patients with schizophrenia.31 Further-
more, nicotine’s improving effect on sensory gating
deficit is inhibited by the known R7 antagonist R-bun-
garotoxin.36 It was concluded that nicotine’s effect may
be caused by stimuli acting on the R7 receptors.
agonist of the R7 nicotinic acetylcholine receptor with
good oral bioavailability and brain permeability.
Experimental Section
Chemistry.
Melting points were determined in open capillary tubes with
a Buchi 530 apparatus and are uncorrected. All 1H NMR
spectrum were recorded on a JEOL GSX-270 spectrometer
(270 MHz) and a JEOL GSX-400 spectrometer (400 MHz),
using tetramethylsilane as internal standard. The following
NMR abbreviations are used: br (broad), brs (broad singlet),
s (singlet), d (doublet), dd (double doublet), dt (double triplet),
t (triplet), q (quartet), m (multiplet). Elemental analysis was
performed on a Yanamoto CHN CORDER MT-6. Analytical
results for all compounds were within (0.4% of the theoretical
values unless otherwise noted. Column chromatography was
carried out on silica gel PSQ-100B (Fuji Silicia). Thin-layer
chromatography (TLC) was performed using plates precoated
with silica gel 60 F-254 (Merck). Abbreviations for solvents
used are DMSO, dimethyl sulfoxide; DMF, dimethylform-
amide; IPE, diisopropyl ether; AcOEt, ethyl acetate; IPA,
isopropyl alcohol. All yields were unoptimized.
(R)-3′-(Benzo[b]thiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one Hydrochloride (7). A
mixture of (S)-spiro-1-azabicyclo[2,2,2]octane-3,5′-oxazolidin-
2′-one (2) (1.0 g, 5.49 mmol), 2-bromobenzo[b]thiophene
(2.6 g, 12.20 mmol), CuI (0.1 g, 0.53 mmol), and K2CO3
(0.69 g, 4.99 mmol) was heated at 130 °C for 12 h. The reaction
mixture was cooled and diluted with chloroform and purified
by silica gel column chromatography (eluent, CHCl3:
MeOH ) 10:1) to give compound 7, which was then converted
to a HCl salt (0.40 g, 21%) to give pale yellow crystals: mp
>270 °C; 1H NMR (DMSO-d6) δ 1.85-2.10 (m, 4H), 3.19-3.23
(m, 3H), 3.24-3.40 (m, 2H), 3.68 (dd, J ) 15 Hz, 22 Hz, 2H),
4.19 (d, J ) 10 Hz, 1H), 4.40 (d, J ) 10 Hz, 1H), 6.81 (s, 1H),
7.25 (t, J ) 7 Hz, 1H), 7.34 (t, J ) 7 Hz, 1H), 7.71 (d,
J ) 8 Hz, 1H), 7.87 (d, J ) 8 Hz, 1H), 10.64 (brs, 1H). Anal.
(C17H18N2O2S‚HCl) C, H, N.
Conclusion
A series of substituted-spiro-1-azabicyclo[2.2.2]octane-
3,5′-[1′,3′]oxazolidin-2′-ones was synthesized and evalu-
ated for R7 binding affinity. Preliminary analysis of
structure-activity relationships using various aryl moi-
eties identified the thiophene derivative 25 as having
potent binding affinity (Ki ) 9 nmol/L) and agonistic
activity. The agonistic activity profile of compound 25
is different from that of compound 2, which is known
as the first selective agonist of the R7 receptors. It also
showed satisfactory selectivity for the other nicotinic
subtypes (R4â2 and R1â2γδ). Furthermore, pharmaco-
kinetic screening demonstrated that compound 25 had
a PK profile improved relative to the earlier compounds
and also more favorable than that of compounds 1 and
2, which thus supports our original hypothesis. In an
MK-801-induced sensory gating model, compound 25
had an improving effect on cognitive dysfunction. Com-
pound 25 thus shows promise as a potential candidate
for further development as a novel, potent, and selective

R7 Nicotinic Acetylcholine Receptor Agonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 7 2683
(S)-3′-(Benzo[b]thiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one Hydrochloride (8) This
compound was prepared from (R)-spiro-1-azabicyclo[2,2,2]-
octane-3,5′-oxazolidin-2′-one (5) (0.9 g, 4.94 mmol) and
2-bromobenzo[b]thiophene (2.7 g, 12.67 mmol) using the same
procedure as described in the preparation of compound 7 and
then converted to a HCl salt (1.03 g, 59%) to give pale yellow
crystals: mp >270 °C; 1H NMR (DMSO-d6) δ 1.85-2.10
(m, 4H), 3.19-3.23 (m, 3H), 3.24-3.40 (m, 2H), 3.68 (dd,
J ) 15 Hz, 22 Hz, 2H), 4.19 (d, J ) 10 Hz, 1H), 4.40 (d, J ) 10
Hz 1H), 6.81 (s, 1H), 7.25 (t, J ) 7 Hz, 1H), 7.34 (t, J ) 7 Hz,
1H), 7.71(d, J ) 8 Hz, 1H), 7.87 (d, J ) 8 Hz, 1H), 10.64 (brs,
1H). Anal. (C17H18N2O2S‚HCl) C, H, N.
(R)-3′-(2-Naphthyl)spiro-1-azabicyclo[2.2.2]octane-3,5′-
[1′,3′]oxazolidin-2′-one (9). This compound was prepared as
pale yellow crystals (1.2 g, 71%) from (S)-spiro-1-azabicyclo-
[2,2,2]octane-3,5′-oxazolidin-2′-one (2) (1.0 g, 5.49 mmol) and
2-bromonaphthalene (2.6 g, 12.56 mmol) using the same
procedure as described in the preparation of compound 7: mp
193-195 °C; 1H NMR (CDCl3) δ 1.51-1.90 (m, 3H), 2.10-2.25
(m, 2H), 2.76-3.08 (m, 4H), 3.04 (d, J ) 15 Hz, 1H), 3.39 (d,
J ) 15 Hz, 1H), 3.92 (d, J ) 9 Hz, 1H), 4.23 (d, J ) 9 Hz, 1H),
7.43-7.51 (m, 2H), 7.70 (d, J ) 2 Hz, 1H), 7.78-7.87 (m, 3H),
7.99 (dd, J ) 2 Hz, 9 Hz, 1H). Anal. (C19H20N2O2) C, H, N.
(S)-3′-(2-Naphthyl)spiro-1-azabicyclo[2.2.2]octane-3,5′-
[1′,3′]oxazolidin-2′-one (10). This compound was prepared
as pale yellow crystals (0.75 g, 49%) from (R)-spiro-1-azabicyclo-
[2,2,2]octane-3,5′-oxazolidin-2′-one (5) (0.9 g, 4.94 mmol) and
2-bromonaphthalene (2.6 g, 12.56 mmol) using the same
procedure as described in the preparation of compound 7: mp
193-194 °C; 1H NMR (CDCl3) δ 1.51-1.90 (m, 3H), 2.10-2.25
(m, 2H), 2.76-3.08 (m, 4H), 3.04 (d, J ) 15 Hz, 1H), 3.39 (d,
J ) 15 Hz, 1H), 3.92 (d, J ) 9 Hz, 1H), 4.23 (d, J ) 9 Hz, 1H),
7.43-7.51 (m, 2H), 7.70 (d, J ) 2 Hz, 1H), 7.78-7.87 (m, 3H),
7.99 (dd, J ) 2 Hz, 9 Hz, 1H). Anal. (C19H20N2O2) C, H, N.
(R)-3′-(Quinolin-2-yl)spiro-1-azabicyclo[2.2.2]octane-
3,5′-[1′,3′]oxazolidin-2′-one (11). A solution of (S)-spiro-1-
azabicyclo[2,2,2]octane-3,5′-oxazolidin-2′-one (2) (0.55 g,
3.02 mmol) and NaH (60%, 132 mg, 9.17 mmol) was stirred in
DMF (5 mL) at room temperature for 0.5 h. After addition of
2-chloroquinoline (0.55 g, 3.36 mmol) at room temperature,
the reaction mixture was stirred at 100 °C for 1.5 h. Water
was added to the resulting reaction mixture, which was then
extracted with CHCl3. The combined CHCl3 solution was
washed with brine, dried over anhydrous magnesium sulfate,
and evaporated to afford the residue, which was crystallized
from AcOEt/IPE and washed with IPE to give compound 11
(0.14 g, 15%) as pale yellow crystals: mp 170-172 °C; 1H NMR
(CDCl3) δ 1.49-1.57 (m, 1H), 1.72-1.82 (m, 2H), 2.13-2.20
(m, 2H), 2.79-3.02 (m, 4H), 3.07 (dd, J ) 2 Hz, 15 Hz, 1H),
3.36 (d, J ) 15 Hz, 1H), 4.15 (d, J ) 11 Hz, 1H), 4.55 (d,
J ) 11 Hz, 1H), 7.46 (dt, J ) 1 Hz, 8 Hz, 1H), 7.67 (dt, J ) 1
Hz, 7 Hz, 1H), 7.78 (d, J ) 8 Hz, 1H), 7.88 (d, J ) 9 Hz, 1H),
8.14 (d, J ) 9 Hz, 1H), 8.44 (d, J ) 9 Hz, 1H). Anal.
(C18H19N3O2‚0.2H2O) C, H, N.
(m, 3H), 2.10-2.15 (m, 2H), 2.76-2.94 (m, 4H), 3.04 (d,
J ) 15 Hz, 1H), 3.36 (d, J ) 15 Hz, 1H), 4.05 (d, J ) 10 Hz,
1H), 4.41 (d, J ) 10 Hz, 1H), 7.25-7.35 (m, 2H), 7.54
(d, J ) 8 Hz, 1H), 7.59 (d, J ) 9 Hz, 1H). Anal. (C16H17N3O3‚
0.1H2O) C, H, N.
(R)-3′-(Benzo[b]furan-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one (14). This compound was
prepared as brown crystals (6.0 mg, 7%) from (S)-spiro-1-
azabicyclo[2,2,2]octane-3,5′-oxazolidin-2′-one (2) (0.54 g, 2.96
mmol) and 2-bromobenzo[b]furan (1.50 g, 7.61 mmol) using
the same procedure as described in the preparation of com-
pound 7: mp 96-98 °C; 1H NMR (CDCl3) δ 1.60-1.92 (m, 3H),
2.15-2.30 (m, 2H), 2.90-3.14 (m, 4H), 3.15-3.32 (m, 1H),
3.40-3.53 (m, 1H), 3.97-4.06 (m, 1H), 4.35 (d, J ) 10 Hz, 1H),
6.67 (s, 1H), 7.16-7.32 (m, 2H), 7.37 (d, J ) 7 Hz, 1H), 7.49
(d, J ) 7 Hz, 1H). Anal. (C17H18N2O3‚0.2H2O) C, H, N.
3′-Phenylspiro-1-azabicyclo[2.2.2]octane-3,5′-[1′,3′]ox-
azolidin-2′-one (15). This compound was prepared as brown
crystals (506 mg, 36%) from (rac)-spiro-1-azabicyclo[2,2,2]-
octane-3,5′-oxazolidin-2′-one (6) (1.00 g, 5.49 mmol) and
bromobenzene (1.96 g, 12.5 mmol) using the same proce-
dure as described in the preparation of compound 7: mp
153-154 °C; 1H NMR (CDCl3) δ 1.49-1.80 (m, 3H), 2.08-2.21
(m, 2H), 2.76-3.06 (m, 5H), 3.36 (dd, J ) 2 Hz, 15 Hz, 1H),
3.79 (d, J ) 9 Hz, 1H), 4.11(d, J ) 10 Hz, 1H), 7.15 (t, J ) 6
Hz, 1H), 7.39 (t, J ) 8 Hz, 2H), 7.54 (d, J ) 9 Hz, 2H). Anal.
(C15H18N2O2‚0.2H2O) C, H, N.
(R)-3′-(2-Pyridyl)spiro-1-azabicyclo[2.2.2]octane-3,5′-
[1′,3′]oxazolidin-2′-one (16). This compound was prepared
as colorless crystals (23.6 mg, 3%) from (S)-spiro-1-azabicyclo-
[2,2,2]octane-3,5′-oxazolidin-2′-one (2) (0.54 g, 2.96 mmol) and
2-bromopyridine (0.32 mL, 3.30 mmol) using the same proce-
dure as described in the preparation of compound 11: mp 124-
125 °C; 1H NMR (CDCl3) δ 1.50-1.65 (m, 1H), 1.66-1.84
(m, 2H), 2.08-2.16 (m, 2H), 2.78-3.12 (m, 5H), 3.31-3.45
(m, 1H), 3.94 (d, J ) 11 Hz, 1H), 4.38 (d, J ) 11 Hz, 1H), 7.03
(t, J ) 6 Hz, 1H), 7.70 (t, J ) 8 Hz, 1H), 8.20 (t, J ) 9 Hz,
1H), 8.30 (t, J ) 3 Hz, 1H). Anal. (C14H17N3O2) C, H, N.
(R)-3′-(2-Thienyl)spiro-1-azabicyclo[2.2.2]octane-3,5′-
[1′,3′]oxazolidin-2′-one Hydrochloride (17). This compound
(3.3 g, 76%) was prepared from (S)-spiro-1-azabicyclo[2,2,2]-
octane-3,5′-oxazolidin-2′-one (2) (3.0 g, 16.5 mmol) and
2-bromothiophene (6.7 g, 41.1 mmol) using the same procedure
as described in the preparation of compound 5. Part of
compound 17 was then converted to a HCl salt to give colorless
crystals: mp >270 °C; 1H NMR (DMSO-d6) δ 1.78-1.93
(m, 3H), 2.02-2.12 (m, 1H), 2.42 (brs, 1H), 3.12-3.21 (m, 3H),
3.25-3.36 (m, 1H), 3.59 (d, J ) 15 Hz, 1H), 3.66 (dd, J ) 2
Hz, 15 Hz, 1H), 4.10 (d, J ) 9 Hz, 1H), 4.27 (d, J ) 10 Hz,
1H), 6.56 (dd, J ) 1 Hz, 4 Hz, 1H), 6.92 (dd, J ) 1 Hz, 4 Hz,
1H), 7.10 (dd, J ) 1 Hz, 5 Hz, 1H), 10.72 (brs, 1H). Anal.
(C13H16N2O2S‚HCl) C, H, N.
(R)-3′-(3-Thienyl)spiro-1-azabicyclo[2.2.2]octane-3,5′-
[1′,3′]oxazolidin-2′-one hydrochloride (18). This compound
was prepared from (S)-spiro-1-azabicyclo[2,2,2]octane-3,5′-
oxazolidin-2′-one (2) (0.54 g, 2.96 mmol) and 3-bromothiophene
(1.2 g, 7.36 mmol) using the same procedure as described in
the preparation of compound 7 and was then converted to a
HCl salt (0.31 g, 35%) to give brown crystals: mp >270 °C;
1H NMR (DMSO-d6) δ 1.80-1.98 (m, 3H), 2.05-2.13 (m, 1H),
2.42 (brs, 1H), 3.14-3.25 (m, 3H), 3.26-3.40 (m, 1H), 3.64
(dd, J ) 4 Hz, 20 Hz, 2H), 4.07 (d, J ) 10 Hz, 1H), 4.26
(d, J ) 10 Hz, 1H), 7.19 (dd, J ) 1 Hz, 3 Hz, 1H), 7.42
(dd, J ) 1 Hz, 5 Hz, 1H), 7.61 (dd, J ) 3 Hz, 5 Hz, 1H), 10.57
(brs, 1H). Anal. (C13H16N2O2S‚HCl‚0.1H2O) C, H, N.
(R)-3′-(Benzothiazol-2-yl)spiro-1-azabicyclo[2.2.2]octane-
3,5′-[1′,3′]oxazolidin-2′-one (12). This compound was pre-
pared as colorless crystals (0.17 g, 18%) from (S)-spiro-1-
azabicyclo[2,2,2]octane-3,5′-oxazolidin-2′-one (2) (0.55 g,
3.02 mmol) and 2-chlorobenzothiazole (0.56 g, 3.30 mmol)
using the same procedure as described in the preparation of
compound 11: mp 195-197 °C; 1H NMR (CDCl3) δ 1.52-1.59
(m, 1H), 1.70-1.77 (m, 2H), 2.11-2.15 (m, 2H), 2.79-2.98
(m, 4H), 3.05 (d, J ) 15 Hz, 1H), 3.37 (d, J ) 15 Hz, 1H), 4.13
(d, J ) 11 Hz, 1H), 4.53 (d, J ) 10 Hz, 1H), 7.32 (t, J ) 7 Hz,
1H), 7.44 (dt, J ) 1 Hz, 8 Hz, 1H), 7.80 (d, J ) 8 Hz, 2H).
Anal. (C16H17N3O2S) C, H, N.
(R)-3′-(Benzoxazol-2-yl)spiro-1-azabicyclo[2.2.2]octane-
3,5′-[1′,3′]oxazolidin-2′-one (13). This compound was pre-
pared as colorless crystals (0.12 g, 13%) from (S)-spiro-1-
azabicyclo[2,2,2]octane-3,5′-oxazolidin-2′-one (2) (0.55 g,
3.02 mmol) and 2-chlorobenzoxazole (0.51 g, 3.32 mmol) using
the same procedure as described in the preparation of com-
pound 11: mp 201-203 °C; 1H NMR (CDCl3) δ 1.51-1.81
(R)-3′-(3-Methylthiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one Hydrochloride (19).
This compound was prepared from (S)-spiro-1-azabicyclo[2,2,2]-
octane-3,5′-oxazolidin-2′-one (2) (0.54 g, 2.96 mmol) and 2-bromo-
3-methylthiophene (1.33 g, 7.50 mmol) using the same proce-
dure as described in the preparation of compound 7 and was
then converted to a HCl salt (48.5 mg, 5%) to give brown
crystals: mp 269-271 °C; 1H NMR (DMSO-d6) δ 1.75-1.90
(m, 3H), 2.00-2.20 (m, 1H), 2.06 (s, 3H), 2.42 (brs, 1H), 3.10-

2684 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 7
Tatsumi et al.
3.60 (m, 4H), 3.50-3.70 (m, 2H), 3.95 (d, J ) 9 Hz, 1H), 4.02
(d, J ) 9 Hz, 1H), 6.84 (d, J ) 5 Hz, 1H), 7.33 (d, J ) 5 Hz,
1H), 10.78 (brs, 1H). Anal. (C14H18N2O2S‚HCl‚0.5H2O) C,
H, N.
octane-3,5′-oxazolidin-2′-one (2) (0.54 g, 2.96 mmol) and 2-bromo-
5-chlorothiophene (1.48 g, 7.49 mmol) using the same proce-
dure as described in the preparation of compound 7 and was
then converted to a HCl salt (0.16 g, 16%) to give pale yellow
crystals: mp >270 °C; 1H NMR (DMSO-d6) δ 1.78-1.94
(m, 3H), 2.00-2.10 (m, 1H), 2.44 (brs, 1H), 3.10-3.24 (m, 3H),
3.25-3.36 (m, 1H), 3.62 (dd, J ) 15 Hz, 25 Hz, 2H), 4.04 (d, J
) 10 Hz, 1H), 4.26 (d, J ) 10 Hz, 1H), 6.37 (d, J ) 4 Hz, 1H),
6.97 (d, J ) 4 Hz, 1H), 10.91 (brs, 1H). Anal. (C13H15ClN2O2S‚
HCl) C, H, N.
(R)-3′-(5-Bromothiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one (26). (R)-3′-(2-Thienyl)-
spiro-1-azabicyclo[2.2.2]octane-3,5′-[1′,3′]oxazolidin-2′-one (17)
(1.3 g, 4.92 mmol) was dissolved in DMF (20 mL) and treated
with N-bromosuccinimide (NBS) (0.87 g, 4.89 mmol). The
mixture was stirred at 80 °C for 3 h. Most of the DMF was
then removed under reduced pressure. The resultant mixture
was diluted with CHCl3, washed with 10% aqueous K2CO3
solution, and dried over K2CO3. The organic solution was then
evaporated and purified by silica gel column chromatography
(eluent, CHCl3:MeOH ) 50:1) and the obtained crystals were
recrystallized from EtOH to give compound 26 (0.86 g, 51%)
as colorless crystals: mp 180-182 °C; 1H NMR (CDCl3)
δ 1.50-1.71 (m, 2H), 1.73-1.83 (m, 1H), 2.10-2.19 (m, 2H),
2.81-3.00 (m, 4H), 3.07 (d, J ) 15 Hz, 1H), 3.37 (d, J ) 15
Hz, 1H), 3.74 (d, J ) 9 Hz, 1H), 4.05 (d, J ) 10 Hz, 1H), 6.17
(d, J ) 4 Hz, 1H), 6.83 (d, J ) 4 Hz, 1H). Anal. (C13H15-
BrN2O2S) C, H, N.
(R)-3′-(5-Cyanothiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one (27). (R)-3′-(5-Bromo-
thiophen-2-yl)spiro-1-azabicyclo[2.2.2]octane-3,5′-[1′,3′]oxazolidin-
2′-one (26) (0.6 g, 1.75 mmol) was dissolved in DMF (10 mL),
after which Zn(CN)2 (0.2 g, 1.70 mmol) and the catalyst
Pd(PPh3)4 (1.0 g, 0.87 mmol) were added. The reaction mixture
was heated at 100 °C for 1 h, cooled to ambient temperature,
and subjected to addition of 5% Na2S2O3 aqueous solution, 10%
K2CO3 aqueous solution, and AcOEt. The unsoluble solids were
filtered off, and the solution was washed with H2O and brine.
The organic solution was then dried over Na2SO4, evaporated,
and purified by silica gel column chromatography (eluent,
CHCl3:MeOH ) 10:1). The obtained crystals were recrystal-
lized from EtOH to give compound 27 (96.9 mg, 19%) as
colorless crystals: mp 230-232 °C; 1H NMR (CDCl3) δ 1.50-
1.67 (m, 2H), 1.70-1.82 (m, 1H), 2.08-2.17 (m, 2H), 2.76-
2.97 (m, 4H), 3.02 (d, J ) 15 Hz, 1H), 3.37 (d, J ) 15 Hz, 1H),
3.80 (d, J ) 9 Hz, 1H), 4.15 (d, J ) 9 Hz, 1H), 6.38 (d, J ) 4
Hz, 1H), 7.44 (d, J ) 4 Hz, 1H). Anal. (C14H15N3O2S) C, H, N.
(R)-3′-(5-Acetylthiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one (28). Acetyl chloride
(0.3 mL, 4.22 mmol) was added to a solution of (R)-3′-(2-
Thienyl)spiro-1-azabicyclo[2.2.2]octane-3,5′-[1′,3′]oxazolidin-2′-
one (17) (1.0 g, 3.78 mmol) in nitrobenzene (10 mL). The
mixture was cooled to 0 °C, and aluminum chloride (1.1 g, 8.25
mmol) was added in small portions. The reaction mixture was
stirred at 85 °C for 4 h and added to an ice-water solution.
After addition of 10% K2CO3 aqueous solution and CHCl3, the
resultant mixture was filtered and the solvent extracted with
CHCl3 and dried over K2CO3. The organic solution was then
evaporated and purified by silica gel column chromatography
(eluent, CHCl3:MeOH ) 10:1), and the obtained crystals were
washed with IPE to give compound 28 (0.62 g, 54%) as
colorless crystals: mp >270 °C; 1H NMR (CDCl3) δ 1.47-1.81
(m, 3H), 2.06-2.17 (m, 2H), 2.50 (s, 3H), 2.73-2.93 (m, 4H),
3.00 (d, J ) 15 Hz, 1H), 3.35 (d, J ) 13 Hz, 1H), 3.77 (d,
J ) 9 Hz, 1H), 4.12 (d, J ) 9 Hz, 1H), 6.58 (d, J ) 4 Hz, 1H),
7.52 (d, J ) 4 Hz, 1H). Anal. (C15H18N2O3S) C, H, N.
(R)-3′-(4-Methylthiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one Hydrochloride (20).
This compound was prepared from (S)-spiro-1-azabicyclo[2,2,2]-
octane-3,5′-oxazolidin-2′-one (2) (1.1 g, 6.04 mmol) and a
mixture of 2-bromo-4-methylthiophene and 2-bromo-3-methyl-
thiophene (2.6 g, ratio ) 3:2), which was obtained by a
previously reported method,37 using the same procedure as
described in the preparation of compound 7 and was then
converted to a HCl salt (0.76 g, 39%) to give pale gray
crystals: mp >270 °C; 1H NMR (DMSO-d6) δ 1.77-1.93
(m, 3H), 2.00-2.10 (m, 1H), 2.15 (s, 3H), 2.40 (brs, 1H), 3.08-
3.43 (m, 4H), 3.57 (d, J ) 14 Hz, 1H), 3.65 (d, J ) 16 Hz, 1H),
4.07 (d, J ) 10 Hz, 1H), 4.24 (d, J ) 10 Hz, 1H), 6.40 (s, 1H),
6.68 (dd, J ) 1 Hz, 2 Hz, 1H), 11.11 (brs, 1H). Anal.
(C14H18N2O2S‚HCl‚0.25H2O) C, H, N.
(R)-3′-(5-Methylthiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one Hydrochloride (21).
This compound was prepared from (S)-spiro-1-azabicyclo[2,2,2]-
octane-3,5′-oxazolidin-2′-one (2) (0.54 g, 2.96 mmol) and 2-bromo-
5-methylthiophene (1.3 g, 7.34 mmol) using the same proce-
dure as described in the preparation of compound 7 and was
then converted to a HCl salt (0.35 g, 37%) to give brown
crystals: mp >270 °C; 1H NMR (DMSO-d6) δ 1.80-1.94
(m, 3H), 2.04-2.13 (m, 1H), 2.37 (s, 3H), 2.43 (brs, 1H), 3.14-
3.24 (m, 3H), 3.28-3.40 (m, 1H), 3.64 (dd, J ) 15 Hz, 20 Hz,
2H), 4.05 (d, J ) 10 Hz, 1H), 4.24 (d, J ) 10 Hz, 1H), 6.36
(d, J ) 3 Hz, 1H), 6.61 (d, J ) 3 Hz, 1H), 10.70 (brs, 1H). Anal.
(C14H18N2O2S‚HCl‚0.25H2O) C, H, N.
(R)-3′-(4,5-Dimethylthiophen-2-yl)spiro-1-azabicyclo-
[2.2.2]octane-3,5′-[1′,3′ ]oxazolidin-2′-one Hydrochloride
(22). This compound was prepared from (S)-spiro-1-azabicyclo-
[2,2,2]octane-3,5′-oxazolidin-2′-one (2) (0.54 g, 2.96 mmol) and
2-bromo-4,5-dimethylthiophene (1.8 g, 9.42 mmol) using the
same procedure as described in the preparation of compound
7 and was then converted to a HCl salt (0.16 g, 16%) to give
brown crystals: mp >270 °C; 1H NMR (DMSO-d6) δ 1.74-
1.93 (m, 3H), 1.96-2.11 (m, 1H), 2.03 (s, 3H), 2.22 (s, 3H), 2.39
(brs, 1H), 3.12-3.23 (m, 3H), 3.25-3.39 (m, 1H), 3.62 (dd, J
) 15 Hz, 20 Hz, 2H), 4.02 (d, J ) 9 Hz, 1H), 4.19 (d, J ) 10
Hz, 1H), 6.27 (s, 1H), 10.72 (brs, 1H). Anal. (C15H20N2O2S‚HCl‚
0.1H2O) C, H, N.
(R)-3′-(5-Ethylthiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one Hydrochloride (23).
This compound was prepared from (S)-spiro-1-azabicyclo[2,2,2]-
octane-3,5′-oxazolidin-2′-one (2) (0.54 g, 2.96 mmol) and 2-bromo-
5-ethylthiophene (1.43 g, 7.48 mmol) using the same procedure
as described in the preparation of compound 7 and was then
converted to a HCl salt (0.19 g, 20%) to give pale yellow
1
crystals: mp >270 °C; H NMR (DMSO-d6) δ 1.19 (t, J ) 8
Hz, 3H), 1.77-1.95 (m, 3H), 2.01-2.12 (m, 1H), 2.41 (brs, 1H),
2.72 (q, J ) 8 Hz, 2H), 3.10-3.22 (m, 3H), 3.25-3.37 (m, 1H),
3.62 (dd, J ) 15 Hz, 22 Hz, 2H), 4.04 (d, J ) 9 Hz, 1H), 4.22
(d, J ) 9 Hz, 1H), 6.36 (d, J ) 4 Hz, 1H), 6.62 (d, J ) 3 Hz,
1H), 10.73 (brs, 1H). Anal. (C15H20N2O2S‚HCl) C, H, N.
(R)-3′-(5-Isopropylthiophen-2-yl)spiro-1-azabicyclo-
[2.2.2]octane-3,5′-[1′,3′]oxazolidin-2′-one Hydrochloride
(24). This compound was prepared from (S)-spiro-1-azabicyclo-
[2,2,2]octane-3,5′-oxazolidin-2′-one (2) (0.54 g, 2.96 mmol) and
2-bromo-5-isopropylthiophene (1.54 g, 7.51 mmol) using the
same procedure as described in the preparation of compound
7 and was then converted to a HCl salt (29.8 mg, 3%) to give
yellow crystals: mp >270 °C; 1H NMR (DMSO-d6) δ 1.23 (d, J
) 7 Hz, 6H), 1.75-1.95 (m, 3H), 2.00-2.10 (m, 1H), 2.40 (brs,
1H), 3.00-3.25 (m, 3H), 3.26-3.41 (m, 2H), 3.56-3.70 (m, 2H),
4.04 (d, J ) 10 Hz, 1H), 4.22 (d, J ) 9 Hz, 1H), 6.36 (d, J ) 4
Hz, 1H), 6.63 (d, J ) 4 Hz, 1H), 10.67 (brs, 1H). Anal.
(C16H22N2O2S‚HCl) C, H, N.
3′-[5-(1-Hydroxyethyl)thiophen-2-yl]spiro-1-azabicyclo-
[2.2.2]octane-3,5′-[1′ ,3′]oxazolidin-2′-one (29). To a solu-
tion of (R)-3′-(5-Acetylthiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one (28) (0.34 g, 1.10 mmol) in
MeOH (10 mL) was added NaBH4 (127 mg, 3.30 mmol) at
0 °C, and the reaction mixture was stirred at room tempera-
ture for 5 h. The resultant mixture was evaporated and was
subjected to addition of 10% K2CO3 aqueous solution, extracted
(R)-3′-(5-Chlorothiophen-2-yl)spiro-1-azabicyclo[2.2.2]-
octane-3,5′-[1′,3′]oxazolidin-2′-one Hydrochloride (25).
This compound was prepared from (S)-spiro-1-azabicyclo[2,2,2]-

R7 Nicotinic Acetylcholine Receptor Agonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 7 2685
with CHCl3, and dried over K2CO3. The organic solution was
then evaporated and recrystallized from IPA/IPE to give
compound 29 (230.9 mg, 68%) as colorless crystals: mp 207-
209 °C; 1H NMR (CDCl3) δ 1.42-2.25 (m, 9H), 2.70-3.15
(m, 5H), 3.36 (d, J ) 15 Hz, 1H), 3.76 (d, J ) 9 Hz, 1H), 4.07
(d, J ) 9 Hz, 1H), 5.00-5.07 (m. 1H), 6.35 (d, J ) 4 Hz, 1H),
6.75 (d, J ) 4 Hz, 1H). Anal. (C15H20N2O3S‚H2O) C, H, N.
(mmol/L) of the external solution was as follows: NaCl, 135;
KCl, 2; MgCl2, 1; CaCl2, 5; D-glucose, 10; and HEPES, 12.
During the recordings, 0.3 µmol/L tetrodotoxin, 10 µmol/L
bicuculline, 1 µmol/L atropine, and 0.01 µmol/L dihydro-b-
erythroidine were added to the external solution to block action
potential generation, GABAA-receptor-mediated miniature
IPSCs, muscarinic receptor-mediated responses, and
R4â2 nAChR-mediated responses, respectively. The composi-
tion (mmol/L) of the internal solution was as follows: CsCl,
120; EGTA, 10; MgATP, 5; HEPES, 10; and diTris phospho-
creatine, 14. The pHs of the external solution and the internal
solution were adjusted with Tris-base to 7.4 and 7.2, respec-
tively. Application of choline (10 mmol/L), a full agonist at the
R7 nicotinic receptor, induced a rapidly desensitizing inward
current in over 50% of the neurons tested (N ) 200). The
choline-induced rapidly desensitizing current was blocked by
1 nmol/L of methylcaconitine, an antagonist at the R7 nicotinic
receptor, suggesting that the choline-induced inward current
is mediated by this receptor. In all neurons tested, the inward
currents induced by the test compounds were compared with
that induced by 10 mmol/L choline.
Effects of Compound 25 on Paired Auditory-Evoked
Potential Parameters. This examination was performed
using a procedure similar to that previously reported.31 Rats
(SLC Wistar) weighing 250-300 g were equipped with a skull-
screw electrode for the recording of auditory-evoked potentials,
which was permanently placed on the brain surface at the
“vertex” (4.0 mm posterior to the bregma on the midline) under
pentobarbital anesthesia (50 mg/kg, intraperitoneally). Refer-
ence electrodes were placed on the dura 3.0 mm anterior to
the bregma on either side of the midline. Three screws and
dental acrylic cement were applied to secure the electrodes to
the skull surface. After 1-week recovery from surgery, the rats
were connected to the recording electronics via a cable attached
to the headpiece and a commutator on top of the recording
chamber. The rat was allowed free movement within the
recording chamber. A speaker emitted the auditory stimuli at
a sound level of 90 dB as measured by a sound meter
(Neuropack2, Nihon Kohden). These auditory stimuli consisted
of paired clicks (condition and test clicks) of 0.5-ms duration,
0.5 s apart, at 15 s intervals. Normally, 30 trials in duplicate
were accumulated for each recording to determine the T/C
ratio (for N40). The following compounds were used for the
study: MK-801 (3 mg/kg, sc), clozapine (30 mg/kg, po), and
compound 25 (10 mg/kg, po). To avoid drug-induced alterations
in sensitivity, a minimum of 1 week was left between the
compound testing sessions. MK-801 was administered after
measurement of the T/C ratio for control. The increase in the
T/C ratio, indicating MK-801-induced gating deficit, was
measured 22 h later. Thirty minutes later, each compound was
administered and the T/C ratio was measured 30-60 min
later.
Receptor Binding Assay.
r7 nACh Receptor Affinity. [125I]-R-Bungarotoxin binding
to membranes prepared from rat hippocampi was deternimed
using a modification of the method of Briggs et al.38 Hip-
pocampi excised from Wistar rats were homogenized in 15
volumes of 0.32 M sucrose and centrifuged at 1000g for 10 min.
The supernatant was centrifuged at 20 000g for 20 min and
the resulting pellet suspended in pure water and centrifuged
at 8000g for 20 min. This supernatant was centrifuged at
40 000g for 20 min and the pellet washed with pure water
again. The final pellet was resuspended in a buffer solution
(118 mM NaCl, 4.8 mM KCl, 2.5 mM CaCl2, 1.2 mM MgCl2,
and 20 mM Na-HEPES, pH 7.5) and used for binding assay.
The suspended membranes were incubated with [125I]-R-
bungarotoxin (Amersham) and test compounds at 37 °C for
3 h. Bound radioactivity was counted after rapid vacuum
filtration on a GF/B glass filter. Nonspecific binding was
determined using 100 µM (-)-nicotine (Research Biochemi-
cals).
r4â2 nACh Receptor Affinity. [3H]-Cytisine binding to
membranes prepared from rat cerebral cortex was determined
using a modification of the method of Briggs et al.38 Cerebral
cortex excised from Wistar rats was homogenized in 15
volumes of 0.32 M sucrose and centrifuged at 1000g for 10 min.
The supernatant was centrifuged at 20 000g for 20 min and
the resulting pellet suspended in pure water and centrifuged
at 8000g for 20 min. This supernatant was centrifuged at
40 000g for 20 min and the pellet washed with pure water
again. The final pellet was resuspended in a buffer solution
(120 mM NaCl, 2.5 mM KCl, 1 mM CaCl2, 1 mM MgCl2, and
50 mM Tris-HCl, pH 7.4) and used for the binding assay. The
suspended membranes were incubated with [3H]cytisine
(NEN Life Science Products) and test compounds at 4 °C for
75 min. After rapid vacuum filtration on a GF/B glass filter,
the radioactivity was quantified by liquid scintillation spec-
troscopy. Nonspecific binding was determined using 100 µM
(-)-nicotine (Research Biochemicals).
Muscle-type (r1â2γδ) nACh Receptor Affinity. [125I]-R-
Bungarotoxin binding to membranes prepared from BC3H1
cells, a muscle cell line, was determined in order to investigate
the affinity to muscle-type nAChRs. BC3H1 cells were pur-
chased from American Type Culture Collection. BC3H1 cells
were cultured in Dulbecco’s modified Eagle’s minimal essential
medium (DMEM) supplemented with 4 mM l-glutamine and
20% fetal bovine serum. Confluent BC3H1 cells were har-
vested, homogenized in a buffer solution (118 mM NaCl,
4.8 mM KCl, 2.5 mM CaCl2, 1.2 mM MgCl2 and 2 0mM Na-
HEPES, pH 7.5), and centrifuged at 40 000g for 20 min. The
resulting pellet was resuspended in a buffer solution and
centrifuged at 40 000g for 20 min. The final pellet was
resuspended in a buffer solution (118 mM NaCl, 4.8 mM KCl,
2.5 mM CaCl2, 1.2 mM MgCl2, and 20 mM Na-HEPES,
pH 7.5) and used for the binding assay. The suspended
membranes were incubated with [125I]-R-bungarotoxin (Am-
ersham) and test compounds at 37 °C for 3 h. Bound radio-
activity was counted after rapid vacuum filtration on a GF/B
glass filter. Nonspecific binding was determined using 100 µM
R-bungarotoxin (Sigma).
Acknowledgment. We thank Dr. Masafumi Arita
and Dr. Takashi Futamura for their encouragement and
support in these studies.
Supporting Information Available: Table of elemental
analysis data. This material is available free of charge via the
Internet at http://pubs.acs.org.
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