New thiazole derivatives as potent and selective 5-hydroxytriptamine 3 (5-HT3) receptor agonists for the treatment of constipation

Article abstract of DOI:10.1016/S0968-0896(02)00557-6

The syntheses and biological evaluation of a series of novel indeno[1,2-d]thiazole derivatives are described. Several groups reported 5-HT₃ receptor agonists which were mainly evaluated for their activities on the von Bezold-Jarisch reflex (B-J reflex). We discovered that tetrahydrothiazolopyridine derivative 1b had a contractile effect on the isolated guinea pig colon with weak B-J reflex. Our efforts to find a new type of 5-HT₃ receptor agonists on the isolated guinea pig colon focused on the synthesis of a fused thiazole derivative 1d modified from 1b and reverse-fused thiazole derivatives (7-10). In this series, 10f (YM-31636) showed high affinity and selectivity for the cloned human 5-HT₃ receptor; furthermore, it showed potent and selective 5-HT₃ receptor agonistic activity. YM-31636 was examined for its effects on defecation in animals, thus evaluating the compound as an agent against constipation.

Full text of DOI:10.1016/S0968-0896(02)00557-6

Bioorganic & Medicinal Chemistry 11 (2003) 1493–1502
New Thiazole Derivatives as Potent and Selective
-Hydroxytriptamine 3 (5-HT ) Receptor Agonists
for the Treatment of Constipation
5
3
Naoki Imanishi,* Kiyoshi Iwaoka, Hiroyuki Koshio, Shin-ya Nagashima,
Ken-ichi Kazuta, Mitsuaki Ohta, Shuichi Sakamoto, Hiroyuki Ito, Shinobu Akuzawa,
Tetsuo Kiso, Shin-ichi Tsukamoto and Toshiyasu Mase
Institute for Drug Discovery Research,Yamanouchi Pharmaceutical Co., Ltd., 21 Miyukigaoka, Tsukuba, Ibaraki 305-8585, Japan
Received 4 October 2002; accepted 23 October 2002
Abstract—The syntheses and biological evaluation of a series of novel indeno[1,2-d]thiazole derivatives are described. Several
groups reported 5-HT receptor agonists which were mainly evaluated for their activities on the von Bezold–Jarisch reflex (B–J
3
reflex). We discovered that tetrahydrothiazolopyridine derivative 1b had a contractile effect on the isolated guinea pig colon with
weak B–J reflex. Our efforts to find a new type of 5-HT receptor agonists on the isolated guinea pig colon focused on the synthesis
of a fused thiazole derivative 1d modified from 1b and reverse-fused thiazole derivatives (7–10). In this series, 10f (YM-31636)
3
3 3
showed high affinity and selectivity for the cloned human 5-HT receptor; furthermore, it showed potent and selective 5-HT
receptor agonistic activity. YM-31636 was examined for its effects on defecation in animals, thus evaluating the compound as an
agent against constipation.
#
2003 Elsevier Science Ltd. All rights reserved.
Introduction
gastrointestinal tract, 5-HT is stored in and released
from enterochromaffin (EC) cells and enteric ser-
2
a,b
Constipation is one of the most common chronic diges-
tive disorders. The definition of constipation, however,
is not sufficiently established, probably because of its
various kinds of symptoms. The principal cause of the
development of constipation is the motility disorder of
the colon; functional constipation is classified into two
main categories, that is, atonic and spastic constipa-
otonergic neurons.
A 5-HT receptor was identified
3
on enteric neurons.3a,b Selective 5-HT3 receptor
antagonists suppress stress- or 5-HT-induced defecation
4
in rats and reduce 5-HT-induced colonic motility in
5
a,b
rats and dogs. It has also been reported that 5-HT3
receptor antagonists inhibit colonic motility in healthy
6
humans, and decrease defecation frequency in women
7
1
tion. Laxatives are widely used in the treatment of
with irritable bowel syndrome. The mechanism of the
constipation, although some of them are contra-
indicated in spastic constipation. Their onset of action is
slow and dispersed, causing difficulty in control of the
time of defecation. Moreover, their main action is to
increase water secretion from the colonic mucosa. They,
therefore, tend to cause diarrhea, dehydration and elec-
trolyte disturbance.
regulation of colonic function through the 5-HT3
receptor is considered to be as follows; 5-HT is released
in compliance with stimulation of mucosal surface by
intralurminal contents, and then binds to the neuronally
located 5-HT receptors. As a result, colonic functions
3
are accelerated through the release of acetylcholine,
tachykinins, VIP and/or NO. 5-HT receptor stimulat-
3
ing agents, therefore, are expected to be new promising
candidate for the treatment of constipation.
5-Hydroxytryptamine (5-HT) is a biogenic amine that
mediates a variety of physiological actions. In the
8
À16
Several groups have reported 5-HT receptor agonists
3
identified with their activity on the 5-HT-evoked reflex
1
7
bradycardia (von Bezold–Jarisch reflex; B–J reflex). We
previously reported that thiazole derivatives, for example
*
5
Corresponding author. Tel.: +81-298-63-6712; fax: +81-298-52-
387; e-mail: imanishi@yamanouchi.co.jp
0
968-0896/03/$ - see front matter # 2003 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(02)00557-6

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N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
2
thiazolo[5,4-c] pyridine 1a, as a 5-HT receptor ago-
nist which stimulated the B–J reflex. But the B–J reflex
is an undesirable side effect against the treatment of
constipation.
- (o - methoxyanilino) - 5 - methyl - 4,5,6,7 - tetrahydro-
1
A series of indenothiazoles were prepared from 2-
bromo-1-indanone by condensation with thioamides or
thioureas in 2-propanol. The primary amine containing
compounds were generated from phthalimido deriva-
tives (Scheme 2).
8
3
We have paid attention to the contractile effects of
5
tract. We have discovered that compound 1b, the deme-
thoxy derivative of 1a, had a contractile effect on the
isolated guinea pig colon as an index for the 5-HT3
receptor agonistic activity, independent of the B–J reflex.
-HT through the 5-HT receptor in the gastrointestinal
3
Results and Discussion
The synthesized compounds were assessed for 5-HT ago-
3
nistic activities with their contractile effect on the isolated
guinea pig colon and potent compounds were assessed
for the B–J reflex in rat as an index for a side effect against
19
20
Our efforts to find a new type of 5-HT receptor ago-
3
nists on the isolated guinea pig colon focused on the
synthesis of a fused thiazole derivative 1d modified from
the cardiovascular system. 5-HT caused a dose-depen-
dent contraction within its concentration range of 0.1–
30 mM and showed the maximum response at 10–30 mM;
1
b and reverse-fused thiazole derivatives 7–10 in which
the fused ring is reversed to an aromatic ring from the
the action of 5-HT is mediated via the 5-HT receptor.
3
Activity of each compound is expressed by a relative value
in comparison with the activity of 5-HT in each specimen.
The intrinsic activity (relative efficacy compared to 5-HT)
is indicated as a percentage of the maximum response by
each compound when the maximum contraction by 5-HT
is defined as 100%. The relative potency compared to
5-HT is determined according to the following equation;
Relative potency=EC value for the 5-HT/EC value for
aliphatic amine part of 1d (Fig. 1).
Chemistry
A series of tetrahydrothiazolopyidines were prepared
from thiourea or thiamide by condensation with halo-
ketones in 2-propanol in good yield (Scheme 1).
50
50
a compound. The contractile effect of each compound was
antagonized by 0.3 mM ramosetron which is a selective
5-HT3 receptor antagonist. Further, the potent com-
pounds were assessed in a [ H]-ramosetron binding study
3
2
1
on the cloned rat 5-HT receptor.
3
Compound 1a was a potent 5-HT agonist as the B–J
3
reflex inducer. Demethoxy compound 1b reduced 5-HT3
agonistic activity, but increased the contractile effect on
the isolated guinea pig colon. Replacement of the linker
nitrogen atom of 1b by a methylene group lost the
5-HT agonistic activity. Compound 1d, with the ben-
zene ring that was directly connected to the thiazole
Scheme 1. Syntheses of 5,6,7,8-tetrahydrothiazolopyridine derivatives:
(a) 3-bromo-1-methyl-4-piperidone hydrobromide, iPrOH.
3
Figure 1. Synthetic plan.
2 2
Scheme 2. Synthesis of 2-substituted indenothiazoles: (a) (EtO) P(:S)SH, 4 N HCl in EtOAc; (b) 2-bromo-1-indanone, iPrOH; (c) MeNH , MeOH.

N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
1495
Table 1. 5-HT
3
receptor agonistic activities and binding profile of the fused-thiazole derivatives
No.
RX
Fused thiazole moieties
Guinea pig colon contraction
at B–J Rr eflex
Maximum
5-HT
3
bindinge
Intrinsic
activity (%)
Relative
potency (fold)
Relative
potency (fold)
IC50(nM)f
a
b
response (%)c
d
1
a
OCH
3
NH
NH
8
—
80
3
14.4
1
b
c
H
H
H
57
0
0.3
—
55
0.25
NT
390
1
CH
2
NT
>1000
1d
bond
58
1
1
26
<0.33
113
150
5-HT
100g
100h
1
—
a
, not determined; NT, not tested.
The intrinsic activity (relative efficacy compared to 5-HT) was indicated as a percentage of the maximum response by each compound when the
maximum contraction by 5-HT is defined as 100%.
b
The relative potency compared to 5-HT was determined according to the following equation. relative potency=EC50 value for the 5-HT/EC50 value
for a compound.
c
Maximum response was indicated by the percentage of the maximum response by each compound when the maximum response by 5-HT was
defined as 100%.
d
The relative potency compared to 5-HT was determined according to the following equation. relative potency=ED50 value for the 5-HT/ED50
value for a compound.
H]-ramosetron was used as a 5-HT receptor ligand.
3
e 3
[
IC50 values determined by duplicate (n=1).
f
g
19
The EC50 value for 5-HT was 3.1 mM.
h
The ED50 value for 5-HT was 15.5 mg/kg iv.
moiety, had potent 5-HT agonistic activity for colonic
3
contraction in spite of weak B–J reflex activity (Table 1).
Further, the chain length between the aromatic ring and
the nitrogen atom were fixed at 2 or 3 carbon lengths
and the terminal amino groups were changed into cyclic
amines. Among the cyclic aliphatic amines, 3-pyrroli-
dine derivatives 8a and 8f showed similar potent con-
tractile activity and also stimulated the B–J reflex, and
they were not enough separation. Compound 8e showed
partial agonistic activity in colonic contraction without
inducing B–J reflex. Lactam compound 9 had no activ-
ity, so it was found that the basic nitrogen atom was
necessary for the activity (Table 3).
The next step, indenothiazoles, in which a fused ring as
reversed to an aromatic ring from the aliphatic amine
part of 1, were assessed for contractile effect. The dis-
tance between the indenothiazole ring and amino group
was varied; C2–C3 methylene length was favorable as a
linker group (Table 2).
Table 2. 5-HT
2-(aminoalkyl)indenothiazoles
3
receptor agonistic activities and binding profile of the
Next, aromatic amine derivatives like pyridines, imida-
zoles were assessed in Table 4. Pyridine ring containing
compound 10a showed weak activity. Changing the
substitution position was not effective with the pyridine
derivatives. However, compounds containing an imida-
zole ring showed potent colon contractile activities.
Their activities were marginally affected by the sub-
stitution position. Especially, 4-substituted compound
No.
n
RGuinea pig colon contraction
5-HT
bindinge
3
IC50 (nM)f
Intrinsic
activity (%)a
Relative
potency (fold)
b
1
0f (YM-31636) showed almost full agonistic activities
7
7
7
7
7
7
7
a
b
c
d
e
f
0
1
2
3
4
2
3
NH
NH
NH
NH
NH
2
2
2
2
2
0
0
75
52
0
—
—
0.5
1
—
3
>1000
>1000
8.2
4.3
3.5
in inducing contraction with an intrinsic activity of 0.90-
and 26-fold potency compared to 5-HT in isolated gui-
nea pig distal colon (Fig. 2). As regarding the chain
length, arylmethyl (n=1) was more favorable than an
ethyl group (10f and 10i).
NMe
NMe
2
76
47
1.2
1.4
g
2
10
We selected YM-31636 for further confirmation to
characterize as a 5-HT receptor agonist in other assays.
—, not determined; NT, not tested.
a,b,e,f
Refer to Table 1.
3

1496
N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
Table 3. 5-HT
3
receptor agonistic activities and binding profile of the aliphatic amine containing indenothiazoles
No.
RGuinea pig colon contraction
at B–J reflex
R
5-HT
Maximum
response (%)c
bindinga
3
Intrinsic
activity (%)a
Relative
potency (fold)
Relative
potency (fold)
IC50 (nM)a
b
d
8a
8b
8c
75
79
76
14
3
97
4.2
—
1.54
7.17
12.3
30
4
NT
NT
8
8
d
e
78
26
6
4
NT
NT
—
NT
NT
0.4 @1 mg/kg
8
9
f
55
10
—
75
12
NT
NT
2.4
NT
NT
—, not determined; NT, not tested.
aÀf
Refer to Table 1.
Until recently, only one subunit of the 5-HT receptor
3
5-HT agonist (Table 5). Additionally, YM-31636 did
3
had been cloned, but a distinct subunit, 5-HT , has
3
not show affinities to other receptors as shown in Table 6.
B
2
2
been discovered. Transcripts of this subunit are co-
expressed with the established 5-HT receptor subunit,
In these results, YM-31636 showed the most potent
5-HT receptor binding affinity and colon selectivity in
vitro tests.
3
5
-HT3A, in a specific brain region. YM-31636 showed
3
extremely higher affinity for the cloned human 5-HT3A
receptor compared to the other compounds known as
We examined the anticonstipation effect of YM-31636
2
3
on defecation in ferrets. YM-31636 facilitated defeca-
tion within 1 h, mostly within 30 min, after oral admin-
istration in ferrets (Fig. 3). As shown in Figure 4,
YM-31636 significantly increased the frequency of
Figure 2. The effects of 10f, 5-HT, 2-methyl-5-HT and m-chlor-
ophenylbiguanide (n=6) on contraction of isolated guinea pig distal
colon. Segments of the distal colon approximately 20 mm long were
suspended longitudinally in an organ bath containing Krebs-bicarbo-
nate solution; isometric contractions were recorded. Concentration–
response curves were constracted in a cumulative manner. Each point
represents the meanÆSEM.
Figure 3. 10f facilitated defecation after oral administration in ferrets.
The between-subjects and within-subjects effects and the interaction
were significantly different by two-way repeated measures ANOVA.

N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
1497
Table 4. 5-HT
3
receptor agonistic activities and binding profile of the aromatic amine containing indenothiazoles
No.
n
Ar
Guinea pig colon contraction
Rat B–J reflex
5-HT
3
bindinga
Intrinsic
activity (%)a
Relative
potency (fold)
Maximum
response (%)c
Relative
potency (fold)
IC50(nM)a
b
d
10a
1
13
1/6
NT
NT
185
1
1
1
1
1
1
1
0b
0c
0d
0e
0f
1
1
1
1
1
2
2
2
0
0
—
—
—
3
NT
NT
NT
NT
41.6
83.6
NT
0
NT
NT
61
90
0
NT
NT
11.7
0.40
457
26
—
4
28 @1 mg/kg
NT
<0.01
NT
0g
0h
0i
99
64
NT
NT
12.8
0.70
1
12
NT
NT
—, not determined; NT, not tested.
aÀf
Refer to Table 1.
defecation within 1 h from 0.03 mg/kg po administration
and this effect was inhibited by ramosetron, a selective
constipation of both atonic and spastic types, such as
atonic constipation, irritable bowel syndrome (con-
stipation-predominant), morphine-induced constipa-
tion, post-operative ileus, symptomatic constipation
(diabetes mellitus, Parkinson’s disease, etc.), chronic
colonic obstruction and colonic inertia.
5
-HT receptor antagonist. These results suggest that
3
YM-31636 is expected to be a new type of agent against
constipation which has high potency and efficacy. The
results also indicate that YM-31636 has an early and
restricted onset of action compared to existing laxatives.
In conclusion, we found that YM-31636 (10f) was a
potent and selective 5-HT receptor agonist. YM-31636
facilitated defecation with high potency and efficacy.
YM-31636 would be promising in the treatment of
Table 6. Affinity of YM-31636 for other receptors21
3
Receptor
[3H] ligand
Tissue
i
K (nM)
a
a
1
-Adrenergic
-Adrenergic
Prazosin
Rauwolsine
DHA
Rat brain
Rat brain
Rat brain
>10,000
1300Æ80
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
0.20Æ0.02
2
Table 5. Affinity of 5-HT
-HT3A receptors21
3
receptor agonists for cloned human
b-Adrenergic
5
Benzodiazepine
Flunitrazepam
SCH23390
Racropride
Pyrilamine
QNB
Naloxone
5-HT
Ketanserin
GR65630
Rat brain
Dopamine D
Dopamine D
1
2
Rat striatum
Rat striatum
g.p. brain
Rat brain
Rat brain
Rat brain
Rat brain
Rabbit illeum
Compd
pK
i
(nM)
nH
Histamine H
Muscarinic
Opiate
1
5
2
-HT
-Methyl-5-HT
6.47Æ0.17
6.18Æ0.09
6.65Æ0.07
9.67Æ0.05
0.81Æ0.07
1.10Æ0.13
0.91Æ0.09
0.89Æ0.19
m-Chlorophenylbiguanide
YM-31636
5-HT1
5-HT2
5
-HT
3
Each value represents meanÆSEM (n=3). Radioligand used was
3
[
H]ramosetron.
Each value represents meanÆSEM (n=3).

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N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
Figure 4. The effect of 10f on defecation in normal ferrets. 10f (0.01–3 mg/kg, n=10–30) was administered orally and the frequency of defecation
was observed. Each value represents the meanÆSEM for 1 h after administration. (a) Pretreatment with ramosetron, 1 mg/kg sc, 0.5 h before
#
administration of YM-31636. *p<0.05 compared with control group with Dunnett’s test. p<0.05 compared with YM-31636 (1 mg/kg) treatment
group with Student’s t-test.
Experimental
under reflux for 10 h and concentrated in vacuo. The
Chemistry
residue was dissolved in ethyl acetate, and the organic
layer was extracted with aqueous 1 N HCl. The organic
layer was separated and discarded. The aqueous layer
was basified with aqueous saturated NaHCO3 and
All melting points were determined on a Yanaco MP-
1
5
00D melting point apparatus without correction. H
NMRspectra were measured with a JEOL FX90Q, a
FX100, a FX270 or FX400 spectrometer; chemical
shifts are reported in d units using tetramethylsilane as
internal standard and the following abbreviations are
used: s=singlet, d=doublet, t=triplet, q=quartet,
m=multiplet, dd=double doublet, dt=double triplet,
br=broad. Mass spectra were recorded with a Hitachi
M-80 electron impact (EI), JEOL JMS-DX300 (FAB)
spectrometer or Hewlett Packard 5970 MSD (GC)
spectrometer. Elemental analyses were performed with a
Yanaco MT-5. All organic extracts were dried over
anhydrous magnesium sulfate and concentrated with a
rotary evaporator under reduced pressure.
extracted with CHCl . The organic layer was washed
3
with H O, brine and dried over MgSO , then con-
2
4
centrated in vacuo. The residue was purified by column
chromatography on silica gel eluting with 50:1:0.1
CHCl /methanol/29% aqueous ammonia. The obtained
3
free amine was dissolved in methanol and treated with
fumaric acid. 1d was obtained as a fumarate (0.24 g
1
6.8% yield): H NMR(DMSO- d ) d 2.42 (3H, s), 2.77–
6
2.85 (4H, m), 3.68 (2H, s), 6.62 (2H, s), 7.45–7.50 (3H,
+
m), 7.86–7.88 (2H, m); EI-MS m/e (M ) 230; Anal.
.
calcd for C H N S C H O 0.5C H OH 0.2H O: C,
.
.
1
3
14
2
4
4
4
2
5
2
58.34; H, 5.30; N, 8.00; S, 9.16. Found: C, 58.18; H,
ꢀ
5.23; N, 7.84; S, 9.26; mp 185–188 C.
2
-Anilino-5-methyl-4,5,6,7-tetrahydrothiazolo[5,4-c]pyri-
dine (1b). A solution of phenylthiourea 2b (0.30 g,
.97 mmol) and 3-bromo-1-methylpiperidin-4-one
2-Benzyl-5-methyl-4,5,6,7-tetrahydrothiazolo[5,4-c]pyri-
dine (1c). Compound 1c was synthesized from phe-
nylthioacetoamide 3 and 3-bromo-1-methylpiperidin-4-
one hydrobromide according to the same procedure as
1
hydrobromide (0.54 g 1.98 mmol) in 2-propanol (5 mL)
solution was stirred under reflux for 1 h and con-
centrated in vacuo. The residue was dissolved in ethyl
acetate, which was washed with saturated aqueous
NaHCO , H O, brine and dried over MgSO , then
concentrated in vacuo to afford the crude aminothia-
zole. This oil was dissolved in ethanol and treated with
fumaric acid. 1b was obtained as a hemifumarate (0.12 g
that for 1d. Compound 1c was obtained as a fumarate
1
(yield 9.3%): H NMR(DMSO- d ) d 2.38 (3H, s), 2.74
6
(4H, s), 3.57 (2H, s), 4.24 (2H, s), 6.61 (2H, s), 7.27–7.37
+
(5H, m); EI-MS m/e (M ) 244; Anal. calcd for
3
2
4
.
C H N S C H O : C, 59.98; H, 5.59; N, 7.77; S, 8.90.
14 16 2 4 4 4
ꢀ
Found: C, 59.74; H, 5.63; N, 7.65; S, 8.82; mp 160–161 C.
1
1
7% yield): H NMR(DMSO- d ) d 2.48 (3H, s), 2.65
Phthalimido-thioacetamide (5b). A solution of aminoace-
tonitrile hydrochloride (2.30 g 24.9 mmol), triethylamine
(3.46 mL, 24.9 mmol), and phthalic anhydride (3.70 g,
24.9 mmol) in CHCl (30 mL) was stirred under reflux
3
for 6 h. The reaction mixture was poured into H O; then
2
6
(
(
2H, br), 2.85 (2H, br), 3.60 (2H, s), 6.62 (1H, s), 6.91
1H, t, J=7.3 Hz), 7.28 (2H, t, J=7.9 Hz), 7.59 (2H, d,
+
J=8.0 Hz), 10.04 (1H, br); EI-MS m/e (M ) 245; Anal.
.
calcd for C H N S 0.5C H O 0.5C H OH H O: C,
.
.
1
3
15
3
4
4
4
2
5
2
5
6
5.80; H, 6.44; N, 12.20; S, 9.31. Found: C, 55.71; H,
ꢀ
the organic layer was separated. The organic layer was
washed with H O, brine and dried over MgSO , and
.05; N, 11.89; S, 9.52; mp 214–216 C.
2
4
then concentrated in vacuo to afford phthalimideaceto-
nitrile (3.07 g, quantitative). The acetonitrile derivative
was dissolved in 4 N HCl in ethyl acetate solution
5
-Methyl-2-phenyl-4,5,6,7-tetrahydrothiazolo[5,4-c]pyri-
dine (1d). A solution of thiobenzamide 4 (1.40 g,
0.2 mmol), 3-bromo-1-methylpiperidin-4-one hydro-
bromide (3.00 g 11.0 mmol), and calcium carbonate
6.00 g) in 2-propanol (30 mL) solution was stirred
1
(20 mL),
and
diethyldithiophosphate
(3.17 mL,
18.9 mmol) was added to this solution. The reaction
mixture was stirred for 6 h at room temperature. The
(

N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
1499
obtained precipitate was collected and washed with
ethyl acetate and ethyl ether. Phthalimido-thioaceta-
aqueous NaHCO , H O, brine and dried over MgSO ,
3 2 4
and then concentrated in vacuo to afford the crude
aminoindenothiazole. This oil was dissolved in metha-
nol and treated with fumaric acid; 6a was obtained as a
1
mide 5b was obtained (2.29 g, 54%): H NMR(DMSO-
d ) d 4.46 (2H, s), 7.85–7.98 (4H, m), 9.44 (1H, brs), 9.76
6
+
1
(
1H, brs); EI-MS m/e (M ) 220.
hemifumarate (0.10 g, 4.4% yield): H NMR(DMSO-
d6) d 3.68 (2H, s), 6.64 (1H, s), 7.14 (1H, t, J=7.8 Hz),
2-Phthalimidopropanethioamide (5c). Compound 5c was
synthesized from 4-phthalimidopropionitrile according
7.28 (1H, t, J=7.3 Hz), 7.46 (1H, d, J=7.3 Hz); EI-MS
+
.
m/e (M ) 188; Anal. calcd for C H N S 0.5C H O :
10 8 2 4 4 4
1
to the same procedure as that for 5b (yield quant): H
NMR(DMSO- d ) d 2.82 (2H, t, J=7.6 Hz), 3.92 (2H, t,
C,58.52; H, 4.09; N, 11.37; S, 13.02. Found: C, 58.38; H,
ꢀ
4.06; N, 11.28; S, 13.10; mp 220–223 C.
6
J=6.8 Hz), 7.85 (4H, s), 9.37 (2H, brs); EI-MS m/e
+
(
M ) 234.
2-Aminomethyl-8H-indeno[1,2-d]thiazole (7b). To
a
solution of 6b (1.30 g, 3.90 mmol) in methanol (10 mL)
was added 40% methylamine in methanol (20 mL). The
reaction mixture was stirred for 16 h at room
temperature. The reaction mixture was concentrated in
3
-Phthalimidobutylthioamide (5d). Compound 5d was
synthesized from 3-phthalimidobutyronitrile according
to the same procedure as that for 5b (yield quantitative):
1
H NMR(DMSO- d ) d 1.97–2.06 (2H, m), 2.42–2.55
vacuo; then the residue was dissolved in CHCl and the
3
6
(
(
2H, m), 3.60 (2H, t, J=7.1 Hz), 7.86 (4H, s), 9.20–9.40
+
organic layer was extracted with aqueous 1 N HCl.
The organic layer was separated and discarded. The
aqueous layer was basified with aqueous 1 N NaOH
2H, br); EI-MS m/e (M ) 248.
4-Phthalimidovarerothioamide (5e). Compound 5e was
synthesized from 4-phthalimidovareronitrile according
and extracted with CHCl . The organic layer was
3
washed with H O, brine and dried over Na SO ,
2
2
4
to the same procedure as that for 5b (yield quantitative):
1
and then concentrated in vacuo to afford the crude
2-aminomethyl-8H-indeno[1,2-d]thiazole. This oil was
dissolved in methanol and treated with fumaric acid. 7b
H NMR(DMSO- d ) d 1.62 (4H, m), 2.50 (2H, t,
6
J=6.5 Hz), 3.58 (2H, t, J=6.5 Hz), 7.85 (4H, m), 9.30
+
1
(
2H, br); EI-MS m/e (M ) 262.
was obtained as a fumarate (730 mg, yield 59%): H
NMR(DMSO- d ) d 3.93 (2H, s), 4.28 (2H, s), 6.55
6
2
-Phthalimidomethyl-8H-indeno[1,2-d]thiazole (6b).
A
(2H, s), 7.26 (1H, t, J=6.7 Hz), 7.37 (1H, t, J=7.3 Hz),
7.57 (1H, d, J=7.3 Hz), 7.63 (1H, d, J=7.3 Hz); EI-
solution of 5b (2.00 g, 7.80 mmol) and 2-bromo-1-inda-
none (1.64 g, 7.80 mmol) in 2-propanol (100 mL) was
stirred under reflux for 6 h. The precipitate was collected
MS
m/e
(M+)
.
202;
Anal.
calcd
for
C H N S C H O 0.1H O: C,56.27; H, 4.47; N, 8.75;
.
1
1
10
2
4
4
4
2
and dissolved in CHCl , which was washed with satu-
3
S, 10.02. Found: C, 56.12; H, 4.45; N, 8.62; S, 10.12; mp
ꢀ
rated aqueous NaHCO , H O, brine and dried over
3
184–186 C.
2
MgSO , then concentrated in vacuo to afford 6b (1.30 g,
4
1
6
7
(
1%): H NMR(DMSO- d ) d 3.79 (2H, s), 5.29 (2H, s),
2-(2-Aminoethyl)-8H-indeno[1,2-d]thiazole (7c). Com-
pound 7c was synthesized from 6c according to the
same procedure as that for 7b. Compound 7c was
6
.20–7.51 (4H, m), 7.70–7.98 (4H, m); FAB-MS m/e
+
M +1) 333.
1
obtained as a fumarate (yield 64%): H NMR(DMSO-
2
Compound 6c was synthesized from 5c according to
the same procedure as that for 6b (yield 70%): H NMR
-(2-Phthalimidoethyl)-8H-indeno[1,2-d]thiazole
(6c).
d6) d 3.23 (2H, t, J=7.3 Hz), 3.36 (2H, t, J=7.3 Hz),
3.92 (2H, s), 6.45 (2H, s), 7.27 (1H, t, J=7.3 Hz), 7.37
(1H, t, J=7.3 Hz), 7.57 (1H, d, J=7.3 Hz), 7.66 (1H, d,
1
+
(
t, J=6.6 Hz), 7.19–7.90 (8H, m); EI-MS m/e (M ) 346.
CDCl ) d 3.49(2H, t, J=7.4 Hz), 3.78 (2H, s), 4.18 (2H,
J=7.3 Hz); EI-MS m/e (M ) 216; Anal. calcd for
.
C H N S C H O : C,57.82; H, 4.85; N, 8.43; S, 9.65.
3
+
1
2
12
2
4
4
4
Found: C, 57.48; H, 4.91; N, 8.38; S, 9.62; mp 208–
ꢀ
2-(3-Phthalimidopropyl)-8H-indeno[1,2-d]thiazole (6d).
Compound 6d was synthesized from 5d according to
210 C.
1
the same procedure as that for 6b (yield 49%): H NMR
CDCl ) d 2.21–2.45 (2H, m), 3.19 (2H, t, J=7.4 Hz),
2-(3-Aminopropyl)-8H-indeno[1,2-d]thiazole (7d). Com-
pound 7d was synthesized from 6d according to the
same procedure as that for 7b. Compound 7d was
(
3
3
7
.78 (2H, s), 3.87 (2H, t, J=7.1 Hz), 7.19–7.51 (4H, m),
+
1
.60–7.87 (4H, m); EI-MS m/e (M ) 360.
obtained as a fumarate (yield quantitative): H NMR
(DMSO-d6) d 2.03–2.11 (2H, m), 2.90 (2H, t,
2
-(3-Phthalimidobutyl)-8H-indeno[1,2-d]thiazole
(6e).
Compound 6e was synthesized from 5e according to
J=6.8 Hz), 3.17 (2H, t, J=7.3 Hz), 3.91 (2H, s), 6.42
(2H, s), 7.26 (1H, t, J=7.3 Hz), 7.37 (1H, t, J=7.3 Hz),
7.56 (1H, d, J=7.3 Hz), 7.63 (1H, d, J=7.3 Hz); EI-MS
1
the same procedure as that for 6b (yield 28%): H NMR
(
+
.
CDCl ) d 1.74–1.83 (4H, m), 3.05–3.28 (2H, m), 3.70–
m/e (M ) 230; Anal. calcd for C H N S C H O :
4
3
13 14
2
4
4
3.77 (2H, m), 3.78 (2H, s), 7.69–7.80 (8H, m); EI-MS m/
+
e (M ) 374.
C,58.94; H, 5.24; N, 8.09; S, 9.26. Found: C, 58.73; H,
ꢀ
5.17; N, 8.05; S, 9.20; mp 178–180 C.
2
-Amino-8H-indeno[1,2-d]thiazole (7a). A solution of
2-(4-Aminobutyl)-8H-indeno[1,2-d]thiazole (7e). Com-
pound 7e was synthesized from 6e according to the
same procedure as that for 7b. Compound 7e was
obtained as a fumarate (yield 32%): H NMR(DMSO-
d6) d 1.62–1.70 (2H, m), 1.80–1.87 (2H, m), 2.82 (2H, t,
2
-bromo-1-indenone (1.93 g, 9.14 mmol) and thiourea
696 mg, 9.14 mmol) in ethanol (20 mL) was stirred
(
under reflux for 4 h. The precipitate was collected and
1
dissolved in CHCl , which was washed with saturated
3

1500
N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
J=7.3 Hz), 3.10 (2H, t, J=7.3 Hz), 3.90 (2H, s), 6.41
2H, s), 7.25 (1H, t, J=7.3 Hz), 7.36 (1H, t, J=7.3 Hz),
.56 (1H, d, J=7.3 Hz), 7.62 (1H, d, J=7.3 Hz); EI-MS
2-[2-(1-Methyl-2-pyrrolidinyl)ethyl]-8H-indeno[1,2-d]thia-
zole (8d). Compound 8d was obtained as a fumarate
(
7
1
(yield 30%): H NMR(DMSO- d ) d 1.62–1.71 (1H, m),
6
+
.
m/e (M ) 244; Anal. calcd for C H N S C H O :
1
1.81–1.89 (2H, m), 1.91–1.98 (2H, m), 2.11–2.19 (1H,
m), 2.30–2.36 (1H, m), 2.53 (3H, s), 2.65–2.69 (1H, m),
2.87 (1H, br), 3.09–3.24 (2H, m), 3.34–3.37 (1H, m),
4
16
2
4
4
4
C,59.68; H, 5.62; N, 7.73; S, 8.85. Found: C, 59.50; H,
ꢀ
5
.73; N, 7.62; S, 8.87; mp 183–186 C.
3
(
.93 (2H, s), 6.57 (2H, s), 7.28 (1H, t, J=7.3 Hz), 7.39
1H, t, J=7.3 Hz), 7.58 (1H, d, J=7.8 Hz), 7.66 (1H, d,
J=7.3 Hz); EI-MS m/e (M+) 284; Anal. calcd for
General procedure for synthesis of the 2-substituted-
indeno[1,2-d]thiazoles
.
.
C H N S C H O 0.4H O: C, 61.87; H, 6.13; N, 6.87;
4
1
7
20
2
4
4
2
2
A
2
-(2-Dimethylaminoethyl)-8H-indeno[1,2-d]thiazole (7f).
solution of 2-bromo-1-indenone (630 mg,
.98 mmol), 3-dimethylaminopropanethioamide hydro-
S, 7.87. Found: C, 61.84; H, 6.08; N, 6.82; S, 7.84; mp
ꢀ
138–139 C.
chloride (500 mg, 2.98 mmol) in 2-propanol (10 mL) was
stirred under reflux for 1.5 h. The precipitate was col-
lected and dissolved in CHCl , which was washed with
2-(3-Quinuclidinyl)-8H-indeno[1,2-d]thiazole (8e). Com-
1
pound 8e was obtained as a fumarate (yield 26%): H
NMR(DMSO- d ) d 1.62–1.99 (4H, m), 2.29–2.30 (1H,
3
6
saturated aqueous NaHCO , H O, brine and dried over
3
MgSO , and then concentrated in vacuo to afford the
4
m), 3.02–3.17 (4H, m), 3.53–3.75 (3H, m), 3.94 (2H, s),
6.52 (2H, s), 7.27 (1H, t, J=7.3 Hz), 7.35 (1H, t,
J=7.3 Hz), 7.51 (1H, d, J=7.8 Hz), 7.69 (1H, d,
2
crude aminoindenothiazole. This oil was dissolved in
methanol and treated with fumaric acid. 7f was
+
J=7.3 Hz); FAB-MS m/e (M +1) 283; Anal. calcd for
.
C H N S C H O : C,63.30; H, 5.56; N, 7.03; S, 8.05.
1
obtained as a fumarate (226 mg, 21% yield): H NMR
(
1
7
18
2
4
4
4
DMSO-d ) d 2.35 (6H, s), 2.86 (2H, t), 3.23 (2H, t),
Found: C, 63.50; H, 5.67; N, 6.95; S, 7.95; mp 199–
ꢀ
6
3
.90 (2H, s), 6.59 (2H, s), 7.25 (1H, t, J=7.3 Hz), 7.36
1H, t, J=7.8 Hz), 7.55 (1H, d, J=7.3 Hz), 7.62 (1H, d,
202 C.
(
J=7.3 Hz); FAB-MS m/e (M +1) 245; mp 142–144 C.
+
ꢀ
5-[(8H-Indeno[1,2-d]thiazol-2-yl)methyl]-1-azabici-
clo[3,3,0]octane (8f). Compound 8f was obtained as a
1
The following compounds were synthesized using the
above general procedure.
fumarate (yield 22%): H NMR(DMSO- d ) d 1.79–1.85
6
(2H, m), 2.00–2.03 (4H, m), 2.31–2.36 (2H, m), 2.87–
2.92 (2H, m), 3.52 (2H, s), 3.57–3.62 (2H, m), 3.84 (2H,
2-(3-Dimethylaminopropyl)-8H-indeno[1,2-d]thiazole (7g).
Compound 7g was obtained as a sesqui fumarate (yield
s), 6.69 (2H, s), 7.24 (1H, t, J=7.3Hz), 7.35 (1H, t,
J=7.3Hz), 7.51 (1H, d, J=7.8 Hz), 7.69 (1H, d,
1
+
4
7%): H NMR(DMSO- d ) d 2.10 (2H, m), 2.59 (6H,
J=7.3Hz); FAB-MS m/e (M +1) 297; Anal. calcd for
6
.
C H N S C H O : C,64.06; H, 5.86; N, 6.79; S, 7.77.
18 20 2 4 4 4
s), 2.91 (2H, t, J=7.8 Hz), 3.13 (2H, t, J=7.8 Hz), 3.91
2H, s), 6.56 (3H, s), 7.14 (1H, t, J=7.8 Hz), 7.28 (1H, t,
J=7.3 Hz), 7.34 (1H, d, J=7.3 Hz), 7.46 (1H, d,
ꢀ
(
Found: C, 63.79; H, 5.89; N, 6.74; S, 7.79; mp 201–202 C.
+
J=7.3 Hz); EI-MS m/e (M ) 258; Anal. calcd for
.
C H N S 1.5C H O : C,57.84; H, 5.64; N, 6.42; S,
2-(1-Methyl-2-oxo-4-pyrrolidinyl)-8H-indeno[1,2-d]thiazole
(9). Compound 9 was obtained as a white powder (yield
1
5
18
2
4
4
4
1
7
1
.35. Found: C, 57.71; H, 5.55; N, 6.41; S, 7.59; mp
ꢀ
36%): H NMR(DMSO- d ) d 2.64 (1H, dd, J=6.7 Hz,
6
23–125 C.
16.5 Hz), 2.79 (3H, s), 2.84 (1H, dd, J=6.7 Hz, 16.5 Hz),
3.64 (1H, dd, J=6.1 Hz, 9.7 Hz), 3.87 (1H, dd,
2-[(1-Methyl-3-pyrrolidinyl)methyl]-8H-indeno[1,2-d]thia-
zole (8b). Compound 8b was obtained as a fumarate
J=8.5 Hz, 9.7 Hz), 3.94 (2H, s), 4.09–4.15 (1H, m), 7.26
(1H, t, J=7.3 Hz), 7.38 (1H, t, J=7.3 Hz), 7.59 (1H, d,
+
J=7.8 Hz), 7.66 (1H, d, J=7.3 Hz); EI-MS m/e (M )
1
(
yield 59%): H NMR(DMSO- d ) d 1.65–1.74 (1H, m),
6
ꢀ
2
2
.06–2.14 (1H, m), 2.56 (3H, s), 2.74–2.82 (2H, m),
.91–3.03 (2H, m), 3.09–3.15 (1H, m), 3.19–3.23 (2H,
270; mp 114–116 C.
m), 3.91 (2H, s), 6.52 (2H, s), 7.24 (1H, t, J=7.3 Hz),
.37 (1H, t, J=7.3 Hz), 7.56 (1H, d, J=7.3 Hz), 7.64
2-(2-Pyridylmethyl)-8H-indeno[1,2-d]thiazole (10a).
Compound 10a was obtained as a fumarate (yield
7
+
1
(
calcd for C H N S C H O : C, 62.16; H, 5.74; N,
1H, d, J=7.3 Hz); FAB-MS m/e (M +1) 271; Anal.
.
6.5%): H NMR(DMSO- d ) d 3.89 (2H, s), 4.59 (2H,
6
s), 6.64 (2H, s), 7.24 (1H, dd, J=6.3 Hz, 7.2 Hz), 7.26–
7.38 (2H, m), 7.46 (1H, d, J=7.8 Hz), 7.55 (1H, d,
J=7.8 Hz), 7.63 (1H, d, J=7.8 Hz), 7.79 (1H, dt,
J=2.0Hz,7.8 Hz), 8.54 (1H, d, J=4.8Hz), 13.15 (2H,
brs); EI-MS m/e (M+) 264; Anal. calcd for
1
6
18
2
4
4
4
7.25; S, 8.30. Found: C, 62.11; H, 5.69; N, 7.22; S, 8.31;
ꢀ
mp 142–143 C.
2-[(1-Methyl-2-pyrrolidinyl)methyl]-8H-indeno[1,2-d]thia-
zole (8c). Compound 8c was obtained as a fumarate
.
C H N S C H O : C,63.15; H, 4.24; N, 7.36; S, 8.43.
4
1
6
12
2
4
4
1
ꢀ
(
yield 72%): H NMR(DMSO- d ) d 1.59–1.77 (3H, m),
Found: C, 63.09; H, 4.18; N, 7.33; S, 8.50; mp 158–160 C.
6
1
.92–1.99 (1H, m), 2.44–2.54 (4H, m), 2.97–3.05 (1H,
m), 3.15–3.24 (2H, m), 3.48–3.53 (1H, m), 3.91 (2H, s),
.58 (2H, s), 7.23–7.27 (1H, m), 7.34–7.38 (1H, m), 7.56
1H, d, J=7.3 Hz), 7.64 (1H, d, J=7.3 Hz); FAB-MS
2-(3-Pyridylmethyl)-8H-indeno[1,2-d]thiazole (10b).
Compound 10b was obtained as a pale yellow crystal
6
(
1
(yield 25%): H NMR(CDCl ) d 3.79 (2H, s), 4.44 (2H,
3
+
m/e calcd
C H N S C H O 0.2H O: C,61.58; H, 5.79; N, 7.18;
(M +1)
.
271;
Anal.
for
s), 7.25–7.28 (2H, m), 7.39 (1H, d, J=7.8 Hz), 7.48 (1H,
d, J=7.8 Hz), 7.69 (1H, d, J=7.8 Hz), 7.74 (1H, d,
J=7.8 Hz), 8.54 (1H, dd, J=1.5 Hz, 4.9 Hz), 8.65 (1H,
J=2.0 Hz); EI-MS m/e (M+) 264; Anal. calcd for
.
1
6
18
2
4
4
4
2
S, 8.22. Found: C, 61.59; H, 5.69; N, 7.25; S, 8.17; mp.
ꢀ
1
72 C(dec).

N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
1501
C H N S: C,72.70; H, 4.58; N, 10.60; S, 12.13. Found:
1
C, 72.71; H, 4.59; N, 10.61; S, 12.31; mp 165–167 C.
2-[2-(1H-Imidazol-4-yl)ethyl]-8H-indeno[1,2-d]thiazole
(10i). Compound 10i was obtained as a fumarate (yield
6
12
2
ꢀ
1
2
3%): H NMR(DMSO- d ) d 3.03 (2H, t, J=7.3 Hz),
6
2
Compound 10c was obtained as a pale yellow crystal
-(4-Pyridylmethyl)-8H-indeno[1,2-d]thiazole
(10c).
3.39 (2H, t, J=7.4 Hz), 3.89 (2H, s), 6.62 (3H, s), 6.88
(1H, s), 7.25 (1H, t, J=6.4 Hz), 7.36 (1H, t, J=6.8 Hz),
7.56 (1H, d, J=7.3 Hz), 7.62 (1H, d, J=7.8 Hz), 7.70
1
(
yield 20%): H NMR(CDCl ) d 3.81 (2H, s), 4.43 (2H,
3
ꢀ
s), 7.23–7.29 (3H, m), 7.38 (1H, t, J=7.3 Hz), 7.49 (1H,
d, J=7.8 Hz), 7.78 (1H, d, J=7.3 Hz), 8.57 (2H, d,
J=5.5 Hz); EI-MS m/e (M+) 264; Anal. calcd for
C H N S: C,72.70; H, 4.58; N, 10.60; S, 12.13. Found:
(1H, s); EI-MS m/e (M+) 267; mp 143–147 C.
2-(1-Methyl-3-pyrrolidinyl)-8H-indeno[1,2-d]thiazole (8a).
A solution of compound 9 (1.54 g, 5.70 mmol) in tolu-
1
6
12
2
ꢀ
1
C, 72.63; H, 4.59; N, 10.43; S, 11.96; mp 122–123 C.
ene (70 mL) was added a Red-Al (4.1 mL) was added
with dropwise at 0 C. This solution was warmed up to
ꢀ
room temperature in 30 min, then stirred under reflux
for 2 h. The reaction mixture was cooled to 0 C, and
2-(1H-Imidazol-2-ylmethyl)-8H-indeno[1,2-d]thiazole (10d).
Compound 10d was obtained as a fumarate (yield
ꢀ
1
4
.1%): H NMR(DMSO- d ) d 3.89 (2H, s), 4.47 (2H,
added aqueous 1 N NaOH solution (30 mL) then stirred
for 30 min at room temperature. Organic layer was
separated, then washed with brine, dried over MgSO4
then, concentrated in vacuo. The residue was purified by
column chromatography on silica gel eluting with
6
s), 6.63 (2H, s), 7.25 (1H, t, J=7.3 Hz), 7.36 (1H, t,
J=7.3 Hz), 7.56 (1H, d, J=7.8 Hz), 7.63 (1H, d,
+
J=7.3 Hz); FAB-MS m/e (M +1) 254; Anal. calcd for
.
C H N S C H O : C,62.98; H, 6.04; N, 6.99; S, 8.01.
1
7
20
2
4
4
4
Found: C, 62.88; H, 6.08; N, 7.04; S, 8.16; mp 201–
ꢀ
50:1:0.1 CHCl /methanol/29% aqueous ammonia.
3
2
04 C.
Obtained free amine was dissolved in methanol and
treated with fumaric acid. 8a was obtained as a fumarate
1
2-(1-Imidazolylmethyl)-8H-indeno[1,2-d]thiazole (10e).
Compound 10e was obtained as a fumarate (yield
(226 mg, 21% yield): H NMR(DMSO- d ) d 2.09–2.13
6
(1H, m), 2.38–2.47 (4H, m), 2.74 (1H, m), 3.10 (1H, m),
3.87–3.93 (3H, m), 6.59 (2H, s), 7.25 (1H, t, J=6.4 Hz),
7.36 (1H, t, J=6.8 Hz), 7.56 (1H, d, J=7.3Hz), 7.63 (1H,
1
2
6
0%): H NMR(DMSO- d ) d 3.93 (2H, s), 5.69 (2H, s),
6
.63 (2H, s), 6.98 (1H, s), 7.28 (1H, t, J=7.8 Hz), 7.33
+
(
7
2
1H, s), 7.38 (1H, t, J=7.3 Hz), 7.57 (1H, d, J=7.3 Hz),
d, J=7.8Hz); EI-MS m/e (M ) 256; Anal. calcd for
+
.
C H N S C H O H O: C,60.98; H, 5.44; N, 7.49; S,
15 16 2 4 4 4
.
.66 (1H, d, J=7.3 Hz), 7.89 (1H, s); EI-MS m/e (M )
2
.
53; Anal. calcd for C H N S C H O 0.1H O:
.
8.57. Found: C, 60.80; H, 5.35; N, 7.48; S, 8.30; mp 129–
ꢀ
1
4
11
3
4
4
4
2
C,58.24; H, 4.13; N, 11.32; S, 8.64. Found: C, 58.21; H,
ꢀ
131 C.
4
.09; N, 11.05; S, 8.74; mp 155–159 C.
Biology
2-(1H-imidazol-4-ylmethyl)-8H-indeno[1,2-d]thiazole (10f).
Compound 10f was obtained as a fumarate (yield
Contractile effects in the isolated guinea pig colon.19 The
distal portion of the colon was removed from a Hartley
guinea pig (300–500 g). The colon was cleaned in fresh
Krebs-bicarbonate buffer at room temperature and cut
into approximately 20-mm segments. The segments
were suspended longitudinally in an organ bath con-
1
3
6
2%): H NMR(DMSO- d ) d 3.87 (2H, s), 4.34 (2H, s),
.63 (2H, s), 7.07 (1H, s), 7.24 (1H, t, J=7.8 Hz), 7.36
6
(
J=7.3 Hz), 7.69 (1H, s); EI-MS m/e (M+) 253; Anal.
1H, t, J=7.3 Hz), 7.54 (1H, d, J=7.8 Hz), 7.63 (1H, d,
.
calcd for C H N S C H O : C, 58.53; H, 4.09; N,
1
4
11
3
4
4
4
ꢀ
1
8
1.38; S, 8.68. Found: C, 58.37; H, 4.21; N, 11.25; S,
ꢀ
taining Krebs-bicarbonate solution warmed to 37 C
and equilibrated with 95% O /5% CO . Isometric con-
.69; mp 202–203 C.
2
2
traction under a loading tension of 1 g was recorded.
The agonists were applied cumulatively to the bath. For
antagonist studies, antagonists were added to the bath
15 min before the application of the agonists. For the
desensitization of 5-HT3 receptors, 2-methyl-5-HT
(30 mM) was added to the bath 30 min before the appli-
cation of the agonists. Activity of each compound is
expressed by a relative value in comparison with the
activity of 5-HT in each specimen. The intrinsic activity
(relative efficacy compared to 5-HT) is indicated as per-
centage of the maximum response by each compound
when the maximum contraction by 5-HT is defined as
100%. The relative potency compared to 5-HT is deter-
mined according to the following equation; relative
potency=EC50 value for the 5-HT/EC50 value for a
compound. The contractile effect of each compound
was antagonized by 0.3 mM ramosetron which is a
2
(
5
3
-[2-(1H-imidazol-2-yl)ethyl]-8H-indeno[1,2-d]thiazole
10g). Compound 10g was obtained as a fumarate (yield
1
.9%): H NMR(DMSO- d ) d 3.15 (2H, t, J=7.8 Hz),
6
.48 (2H, t, J=7.8 Hz), 3.89 (2H, s), 6.62 (2H, s), 6.94
(
7
2H, s), 7.25 (1H, t, J=7.8 Hz), 7.36 (1H, t, J=7.3 Hz),
.54(1H, d, J=7.8 Hz), 7.63 (1H, d, J=7.3 Hz); EI-MS
m/e (M+) 267; Anal. calcd for
C H N S C H O 0.1H O: C,59.24; H, 4.50; N, 10.91;
.
.
1
5
13
3
4
4
4
2
S, 8.31. Found: C, 58.00; H, 4.21; N, 10.31; S, 7.80; mp
ꢀ
1
77–180 C.
2
(
-[2-(1H-Imidazol-1-yl)ethyl]-8H-indeno[1,2-d]thiazole
10h). Compound 10h was obtained as a white powder
1
(yield 24%): H NMR(DMSO- d ) d 3.57 (2H, t,
J=7.3 Hz), 3.89 (2H, s), 4.49 (2H, t, J=7.3 Hz), 7.26
6
(
1H, t, J=3.9 Hz), 7.27 (1H, t, J=7.8 Hz), 7.37 (1H, t,
J=7.3 Hz), 7.56 (1H, d, J=7.3 Hz), 7.64 (1H, d,
J=7.3 Hz), 7.71 (s, 1H); EI-MS m/e (M+) 267; Anal.
calcd for C H N S 0.4H O: C,65.62; H, 5.07; N,
1
1
1
selective 5-HT receptor antagonist.
3
Radioligand 5-HT3 receptor binding studies.21 The
cloned rat 5-HT receptor was expressed on COS-1 cells
by transfection of a plasmid. The transfected cells were
.
5
13
3
2
5.30; S, 11.68. Found: C, 65.74; H, 4.90; N, 15.13; S,
ꢀ
3
1.60; mp 118–121 C.

1502
N. Imanishi et al. / Bioorg. Med. Chem. 11 (2003) 1493–1502
homogenized in HEPES buffer and centrifuged at
Tract. 3rd ed.; Johnson L. R., Ed.; Raven: New York, 1994;
Vol. I, p 423.
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Eur. J. Pharmacol. 1995, 283, 107.
4
[
8,000 g for 10 min. Membranes were incubated with
ꢀ
3
H]ramosetron and inhibitor for 30 min at 25 C. The
incubation was terminated by filtration and washing
with HEPES buffer through GF/B filters. Radioactivity
of the filter was counted with Top Count. IC50 values
were calculated by logit-log analysis.
4
. Miyata, K.; Kamato, T.; Nishida, A.; Ito, H.; Yuki, H.;
Yamano, M.; Tsutsumi, R.; Katsuyama, Y.; Honda, K. J.
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. (a) Kamato, T.; Ito, H.; Nagakura, Y.; Yamano, M.;
Defecation in normal ferrets.23 Male ferrets weighing
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7. Camilleri, M.; Mayer, E. A.; Drossman, D. A.; Heath, A.;
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0
.8–1.5 kg (n=60) were used. The animals were fed an
ordinary laboratory chow and allowed free access to
water under a constant 12 h light-dark cycle. The food
was given once daily in the middle of the light cycle
(
1:00 pm). The animals were used repeatedly with 1
week washout periods between the different tests. All
experiments were performed in compliance with the
regulations of the Animal Ethical Committee of Yama-
nouchi Pharmaceutical. 10f was administered orally at
8
. Richardson, B. P.; Engrl, G.; Donatsch, P.; Stadler, P. A.
Nature 1985, 316, 126.
. Kilpatrick, G. H.; Butler, A.; Burridge, J.; Oxford, A. W.
1
0:00 am and defecation, emetic episodes including
9
vomiting and retching, and some other behavioral
changes were observed. In the preliminary observations
on the duration of the effects of these compounds on
defecation, the effect of 10f finished within 2 h after
administration. We observed, therefore, the effects of
Eur. J. Pharmacol. 1990, 182, 193.
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1
1
990, 33, 13.
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1
0f for 2 h, respectively. No food was supplied during
the observation. In the antagonist study, ramosetron
.1 mg/kg was administered subcutaneously 0.5 h before
lor, A.; Rhodes, K. F. J. Med. Chem. 1992, 35, 1515.
2. Bermudez, J.; Boyland, P.; King, F. D.; Summersell, R. J.
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Gozlan, H.; Hamon, M.; Keane, P. E.; Soubrie, P.; Fur, G. L.
Eur. J. Pharmacol. 1993, 237, 299.
1
0
the administration of 10f. The stools were collected and
1
´
ꢀ
dried (110 C, 24 h). The water content was estimated by
measuring both wet and dry weights of stools. In the
case of repeated dosing, 10f was orally administered
once daily at 10:00 am for 5 days, and the frequency of
defecation for 1 h after administration was observed
each day.
´
14. Perrone, R.; Berardi, F.; Colabufo, N. A.; Tortorella, V.;
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Acknowledgements
1
6. Sato, Y.; Yamada, M.; Yoshida, S.; Soneda, T.; Ishikawa,
M.; Nizato, T.; Suzuki, K.; Konno, F. J. Med. Chem. 1998,
1, 3015.
We thank Dr. T. Shibanuma and K. Miyata for their
support during the course of this work, Drs. Y. Kat-
suyama, T. Kamato and T. Suzuki for helpful discus-
sions, and Messrs H. Kaniwa, M. Shimizu, and the staff
of the Structure Analysis Department for special mea-
surements and elemental analyses. We are also grateful
to Drs. H. Yuki, Y. Nagakura and M. Yamano for the
biological results.
4
17. Fozord, J. R.; Host, M. Br. J. Pharmacol. 1982, 77, 520.
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Application WO92/07849, May 14, 1992.
1
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1