Synthesis and pharmacological evaluation of novel fused thiophene derivatives as 5-HT2A receptor antagonists: Molecular modeling study

Article abstract of DOI:10.1016/j.ejmech.2010.01.013

Novel derivatives of cyclopentathienopyrimidinediones 6, pyridothienopyrimidinediones 7, ethyl cycloheptathiophene-3-carboxylates 10, ethyl tetrahydrothienopyridine-3-carboxylates 11, tetrahydrocycloheptathienopyrimidin-4(3H)-ones 12, tetrahydrotriazolobenzothienopyrimidin-5(4H)-ones 17 and tetrahydro-5H-cycloheptathienopyrimidin-4(3H)-ones 21 have been synthesized and tested for their?5-HT_2A antagonist activity. Preliminary pharmacological studies showed that compounds 3-[2-[4-phenylpiperazin-1-yl]ethyl]-6,7-dihydro-5H-cyclopenta[b]thieno[2,3-d]pyrimidine-2,4(1H,3H)-dione 6a and ethyl 2-[[4-(2-methoxyphenyl)piperazin-1-yl]acetylamino]-4,5,6,7-tetrahydro-6-methylthieno[2,3-c]pyridine-3-carboxylate 11d were found to be the most active molecules as 5-HT_2A antagonists. Molecular modeling and pharmacophore prediction methodology are used to study the structural features required for 5-HT_2A antagonist properties of the active compounds compared with nonactive species by means of the molecular mechanic method. The 2-methoxy substituent in the structure of 11d seems to be necessary for its full antagonistic properties. Optimal placement of basic nitrogen relative to the plane of thiophene core was found to have a profound effect on affinity and biological activity.

Full text of DOI:10.1016/j.ejmech.2010.01.013

European Journal of Medicinal Chemistry 45 (2010) 1805–1820
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and pharmacological evaluation of novel fused thiophene derivatives as
5-HT2A receptor antagonists: Molecular modeling study
,
c
a
Mohamed M. El-Kerdawya, Eman R. El-Bendary b , Alaa A.-M. Abdel-Aziz , Dalia R. El-wasseef ,
*
Naglaa I. Abd El-Aziz a
a Department of Medicinal Chemistry, Faculty of Pharmacy, University of Mansoura, Mansoura 35516, Egypt
b Department of Pharmaceutical Chemistry, Faculty of Pharmacy, King Abdulaziz University, Jeddah 21589, Saudi Arabia
c Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, Riyadh 11451, Saudi Arabia
a r t i c l e i n f o
a b s t r a c t
Article history:
Novel derivatives of cyclopentathienopyrimidinediones 6, pyridothienopyrimidinediones 7, ethyl cyclo-
heptathiophene-3-carboxylates 10, ethyl tetrahydrothienopyridine-3-carboxylates 11, tetrahydrocyclo-
heptathienopyrimidin-4(3H)-ones 12, tetrahydrotriazolobenzothienopyrimidin-5(4H)-ones 17 and
tetrahydro-5H-cycloheptathienopyrimidin-4(3H)-ones 21 have been synthesized and tested for
their 5-HT2A antagonist activity. Preliminary pharmacological studies showed that compounds 3-[2-[4-
phenylpiperazin-1-yl]ethyl]-6,7-dihydro-5H-cyclopenta[b]thieno[2,3-d]pyrimidine-2,4(1H,3H)-dione 6a
and ethyl 2-[[4-(2-methoxyphenyl)piperazin-1-yl]acetylamino]-4,5,6,7-tetrahydro-6-methylthieno[2,3-
c]pyridine-3-carboxylate 11d were found to be the most active molecules as 5-HT2A antagonists.
Molecular modeling and pharmacophore prediction methodology are used to study the structural
features required for 5-HT2A antagonist properties of the active compounds compared with nonactive
species by means of the molecular mechanic method. The 2-methoxy substituent in the structure of 11d
seems to be necessary for its full antagonistic properties. Optimal placement of basic nitrogen relative to
the plane of thiophene core was found to have a profound effect on affinity and biological activity.
Ó 2010 Elsevier Masson SAS. All rights reserved.
Received 22 November 2008
Received in revised form
6 January 2010
Accepted 9 January 2010
Available online 20 January 2010
Keywords:
Synthesis
5-HT2A receptor
Antagonists
Molecular modeling
Flexible alignment
Pharmacophore prediction
1. Introduction
nerve systems of simple as well as complex animals. Serotonin
diffuses to serotonin-sensitive neurons, which control the animal’s
The discovery of new ligands with affinity for the family of
serotonin receptors is an area of active research in Medicinal
Chemistry due to their involvement in numerous physiological and
pathophysiological processes [1–9]. Serotonin modulates the
activity of both central nervous system and peripheral tissues. It
was realized that the action of 5-HT is not all mediated by receptors
of the same type and various pharmacological classifications have
come and gone. Serotonin is a monoamine neurotransmitter. It is
found extensively in the gastrointestinal tract of animals, and about
80–90% of the human body’s total serotonin is located in the
enterochromaffin cells in the gut, where it is used to regulate
intestinal movements [10]. The remainder is synthesized in sero-
tonergic neurons in the central nervous system (CNS) where it has
various functions, including the regulation of mood, appetite, sleep,
muscle contraction, and some cognitive functions including
memory and learning. Serotonin functions as a neurotransmitter in
perception of nutrient availability. Serotonergic signaling plays an
important role in the modulation of human mood, anger and
aggression. Serotonin has broad activities in the brain, and genetic
variation in serotonin receptors and the serotonin transporter,
which facilitates re-uptake of serotonin into presynapses, have
been implicated in neurological diseases. Drugs targeting sero-
tonin-induced pathways are being used in the treatment of many
psychiatric disorders. One focus of clinical research is the influence
of genetics on serotonin action and metabolism in psychiatric
settings. Such studies have revealed that the variation in the
promoter region of the serotonin transporter protein accounts for
nearly 10% of total variance in anxiety-related personality [11], and
the effect of this gene on depression was found to interact with the
environment [12]. Levels of serotonin in the brain show association
with aggression [13], and a mutation in the gene which codes for
the 5-HT2A receptor may double the risk of suicide for those with
that genotype [14]. Serotonin also has effects on appetite, sleep and
general metabolism. In the blood, the major storage site is platelets,
which collect serotonin from plasma. Bleeding causes serotonin
release which constricts blood vessels. Serotonin present in the
* Corresponding author. Tel.: þ966 02 6423019; fax: þ966 02 6423381.
E-mail address: emanelbendary@yahoo.com (E.R. El-Bendary).
0223-5234/$ – see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.01.013

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M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
blood then stimulates cellular growth to repair liver damage [15].
Currently, the serotonin receptors are divided into seven main
classes, 5-HT1–7, in the central and peripheral nervous systems and
in the gastrointestinal tract [16]. With particular emphasis on the
5-HT2 receptor family, the three receptor subtypes, 5-HT2A, 5-HT2B
and 5-HT2C, are G-protein coupled receptors (GPCRs) that couple
through the Gq and G11 proteins eliciting their second messenger
effects through increases in activity of phospholipase C and/or
phospholipase A [17–30]. 5-HT2A receptors are broadly distributed
in the prefrontal, parietal, and somatosensory cortex, claustrum,
pulmonary arterial rings and in platelets [24–30]. Representative
5-HT2A antagonists include ketanserin (A) [24–30], the prototype of
a large group of structurally related derivatives such as risperidone
(B) (Fig. 1) [31]. Other 5-HT2A antagonists, belonging to different
chemical classes, include the arylpiperazine derivative [32–45]. The
structure–activity data for 5-HT2A antagonists suggest that two
planar aromatic or heterocyclic ring systems separated by an
aliphatic or alicyclic chain, containing basic protonatable nitrogen,
seem to constitute a potent 5-HT2A ligand [42–45]. Additional
hydrophobic substituent in the heterocyclic ring, as demonstrated
by the 4-fluorophenyl group in sertindole and the chlorophenyl
ring in clozapine, enhances the antagonistic potency [33,39]. On the
other hand the biophore models for 5-HT1A (Fig. 2) which are
marked on the buspirone molecular structure showed four points
for fitting including aromatic centroid and three nitrogen centers
[42–45]. This three-nitrogen model may induce stabilising effect on
the ligand–receptor complexes. Recently, a series of isoindolone
derivatives as potent and selective 5-HT2C antagonists were
reported [46,47]. Additionally an in silico screening approach of the
corporate database using a 5-HT2C pharmacophore model resulted
in the identification of a related structure containing this template.
5-HT2C pharmacophore model (Fig. 3) consists of a positive ionis-
able group (blue sphere) which is mapped by the basic nitrogen, an
H-bond acceptor (red spheres), mapped by the carbonyl oxygen, an
aromatic ring (brown sphere) and 3 hydrophobic groups (orange
Fig. 2. Proposed pharmacophore models for 5-HT2A (left) and 5-HT1A (right) receptor
binding.
spheres) [46,47]. In the course of a program aimed at the discovery
of new 5-HT3 and 5-HT4 receptor agents, a comparative receptor
mapping of both serotonergic binding sites is reported [5,48,49].
This computer-aided conformational analysis has allowed to
propose a steric model for 5-HT3 and 5-HT4 receptor recognition,
which offers structural insights to aid the design of novel selective
5-HT3 and 5-HT4 antagonists (Fig. 3) [5,48,49]. This model consists
of an aromatic moiety, a coplanar carbonyl group with the oxygen
situated at ca. 3.3–3.6 Å from the centroid of the aromatic ring,
a nitrogen atom situated at ca. 6.7–8.0 Å from this centroid and ca.
5.2–5.4 Å from the oxygen of the carbonyl group, and a voluminous
substituent in the basic amino framework of the molecule (Fig. 3). It
was identifying potent and selective 5-HT6 ligands for cognitive
impairment based on novel heterocyclic scaffolds and pharmaco-
phore model shown in Fig. 4 [50]. The basis of this hypothesis is
that the basic amine and arylsulfonyl moieties of 5-HT6 ligands are
the necessary receptor pharmacophores and the indole and other
heterocyclic cores serve merely as a template to hold these phar-
macophores in the necessary orientation (Fig. 4) [50]. The phar-
macophore model for 5-HT7 antagonism was recently described
representing the first contribution to the rational design of agents
acting at this recently identified serotonin receptor (Fig. 4) [51]. The
Fig. 1. The reported and the proposed 5-HT2A antagonists.

M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
1807
Fig. 3. Proposed pharmacophore models for 5-HT2C (left), 5-HT3 (middle) and 5-HT4 (right) receptor antagonists.
postulated hypothesis was validated with a series of new naph-
tholactam and naphthosultam derivatives that exhibit affinity for
the 5-HT7 receptor. This study offers structural insight to aid the
development of novel 5-HT7 ligands, which are essential for the
knowledge of the (patho) physiological role of this serotonin
receptor subtype [51].
According to the previous rational, it is reported here the
synthesis of some new fused thiophene derivatives and their 5-
HT2A antagonistic activity using isolated pulmonary arterial rings of
rats [25–30]. These novel synthesized compounds were designed as
structurally related to ketanserin A and its analogues risperidone B.
Flexible alignment and pharmacophore prediction methodology
was used to identify the structural features required for 5-HT2A
antagonist properties.
cycloalkanone reacted with ethyl cyanoacetate and sulphur in
absolute ethanol in the presence of diethylamine they afforded the
key starting materials, ethyl 2-amino-5,6-dihydro-4H-cyclo-
penta[b]thiophene-3-carboxylate 1, ethyl 2-amino-5,6,7,8-tetrahy-
dro-4H-cyclohepta[b]thiophene-3-carboxylate 2, ethyl 2-amino-
4,5,6,7-tetrahydro-6-methylthieno[2,3-c]pyridine-3-carboxylate 3
and 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxylate
13 (Schemes 1 and 2).
Reaction of the o-amino esters 1 or 3 with 2-chloroethyl isocy-
anate in toluene afforded the corresponding chloroethyl ureido
derivatives 4 and
5 respectively. The structure of the latter
compounds was established on the basis of its elemental analysis
and spectral data. For example, the 1H NMR spectrum of compound 4
showed a typical quartet–triplet pattern for the ethyl ester; the tri-
plet appeared at 1.20–1.40 ppm while the quartet was observed at
4.20–4.35 ppm and a multiplet signal of (CH2CH2Cl) at 3.60–
3.72 ppm. Heating a mixture of the chloroethyl ureido derivatives 4
or 5 and the appropriate N-substituted phenylpiperazines at 140 ꢀC
afforded3-[2-[4-substitutedphenylpiperazin-1-yl]ethyl]-6,7-dihydro-
2. Results and discussion
2.1. Chemistry
5H-cyclopenta[b]thieno[2,3-d]pyrimidine-2,4(1H,3H)-diones
6a–d
2.1.1. Synthesis of compounds 1–12 (Scheme 1)
and 3-[2-[4-substituted phenylpiperazin-1-yl]ethyl]-5,6,7,8-tetrahy-
dro-7-methylpyrido[40,30:4,5]thieno[2,3-d]pyrimidine-2,4(1H,3H)-di-
ones 7a–d (Scheme 1). The structures of the isolated products were
Heterocyclic o-amino esters have attracted much attention from
the synthetic point of view as they facilitate the preparation of
numerous fused heterocyclic systems [52–54]. When an appropriate
Fig. 4. Proposed pharmacophore models for 5-HT6 (left) and 5-HT7 (right) receptor antagonists.

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M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
Scheme 1.
established on the basis of their elemental and spectral analyses.
For example, the 1H NMR spectrum showed the disappearance of
the signal of the ethyl ester group and the appearance of the protons
of the N-phenylpiperazine moiety in the expected regions of
the spectrum. Condensation of ethyl 2-amino-5,6,7,8-tetrahydro-
tetrahydro-4H-cyclohepta[b]thiophene-3-carboxylates 10a,b,d,e and
ethyl 2-[[4-(substituted phenyl)piperazin-1-yl]acetylamino]-4,5,6,7-
tetrahydro-6-methylthieno[2,3-c]pyridine-3-carboxylates 11a,b,d,e
respectively.
Cyclization of compounds 10a,b,d,e was carried out via the
reaction with hydrazine hydrate in absolute ethanol to give 3-amino-
4H-cyclohepta[b]thiophene-3-carboxylate
2 or ethyl 2-amino-
4,5,6,7-tetrahydro-6-methylthieno[2,3-c]pyridine-3-carboxylate 3
with chloroacetyl chloride in chloroform afforded the correspond-
ing chloroacetyl derivatives 8 and 9 respectively. Treatment of the
compounds 8 and 9 with the appropriate N-substituted phenyl-
piperazines in the presence of potassium carbonate in DMF afforded
ethyl 2-[[4-substituted phenylpiperazin-1-yl]acetylamino]-5,6,7,8-
2-[[4-(substituted
phenyl)piperazin-1-yl]methyl]-6,7,8,9-tetrahy-
dro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-ones 12a,b,d,e.
The IR spectrum of compound 12e revealed the NH2 and C]O
stretching absorption bands at 3280 and 1632 cmꢁ1 respectively.
Moreover, the disappearance of the quartet–triplet pattern of the
ethyl ester in the 1H NMR spectrum and the presence of a broad
COOC2H5
NH2
O
COOC2H5
NHCSNHCOC6H5
NH4SCN
C6H5COCl
N
NaOH/DMF
2. H+
S
Acetone
n
S
N
14
SH
S
n = 1 (13), 2 (2)
18
NH2NH2/DMF
KOH/C2H5OH
O
N
O
NH
S
N
15
NHNH2
S
SK
N
19
ClCOCH2Cl
CHCl3
R
ClCH2CO N
C2H5OH
N
O
O
20a,b,d,e
R
HN
N
N
N
S
S
N
N
N
N
K2CO3 / DMF
N
N
R
R
O
ClH2C
16
N
N
NH
N
N
17a-e
S
N
S
O
21a,b,d,e
R = H (a), 2-Cl (b), 4-F (c), 2-OCH3 (d) , 2-OC2H5 (e)
Scheme 2.

M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
1809
singlet peak at 6.30–6.39 ppm representing (NH2) group which is
2.2. Biological activity
a D2O exchangeable revealed its cyclized structure.
2.2.1. 5-HT2A antagonist activity using isolated pulmonary arterial
rings of rats
2.1.2. Synthesis of compounds 14–21 (Scheme 2)
Treatment of ethyl 2-amino-4,5,6,7-tetrahydrobenzo[b]thio-
phene-3-carboxylate 13 with phenyl isothiocyanate in DMF in the
presence of sodium hydroxide followed by acidification with dilute
acetic acid afforded the mercapto derivative 14. Compound 14
reacted with hydrazine hydrate in DMF to give the corresponding
hydrazino derivative 15 that condensed with chloroacetyl chloride
in chloroform to yield the cyclized 1-chloromethyl-4-phenyl-
6,7,8,9-tetrahydro[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-e]pyri-
midin-5(4H)-one 16. The structure of the latter compound was
established on the basis of its elemental analysis and spectral data,
for example, the notable feature in the 1H NMR spectrum being the
presence of a singlet peak at 4.65 ppm indicating (CH2Cl) group.
Moreover, a molecular ion peak of m/z 371 (5.63%) was clearly
identified in the mass spectrum. Compound 16 was treated with
the appropriate N-substituted phenylpiperazines in DMF to give
the corresponding 4-phenyl-1-[[4-substituted phenylpiperazin-1-
yl]methyl]-6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]benzo[b]thieno-
[3,2-e]pyrimidin-5(4H)-ones 17a–e. The 1H NMR spectrum of the
isolated products 17a–e showed the appearance of the protons of
the N-phenylpiperazine moiety in the expected regions of the
spectrum.
Selected examples of the newly synthesized compounds were
subjected to an in vitro pharmacological screening for 5-HT2A
antagonist activities using isolated pulmonary arterial rings of rats
[25–30]. Risperidone was chosen as the standard drug. Some of the
test compounds in this study inhibited the pressor response of the
isolated pulmonary arterial rings to 5-HT (Table 1) and hence
showed antagonistic activities for 5-HT2A receptors.
2.2.2. Structure–activity relationship and influence of lipophilic and
steric factors
2.2.2.1. Structure–activity relationship. Compound 6a with a phenyl-
piperazine moiety joined by an ethylene side chain to a cyclo-
pentathienopyrimidine skeleton exhibited high 5-HT2A antagonist
activities. Replacement of the cyclopentyl moiety by a pyridine
moiety showed a little effect on decreasing the activities as in
compounds 7a, 7c and 7d. Moreover, changing the position of the
arylpiperazine moiety to 2 position of pyrimidine ring caused a little
decrease in activity such as compounds 21a,b,d. The higher 5-HT2A
antagonist activity of the 2-methoxy derivative 11d may confirm the
importance of this group in the aromatic ring of this skeleton for the
activity. The presence of a triazolo moiety between the condensed
thienopyrimidine nucleus and the (un)substituted phenylpiperazine
moiety greatly decreased the activity which may explain the inac-
tivation of compounds 17a, 17b and 17d. This indicated that the
optimal structure for 5-HT2A antagonist is the presence of not more
than three fused ring system.
On the other hand, benzoyl chloride was allowed to react with
ammonium thiocyanate in anhydrous acetone at reflux for 10 min,
then
a solution of ethyl 2-amino-5,6,7,8-tetrahydro-4H-cyclo-
hepta[b]thiophene-3-carboxylate 2 in anhydrous acetone was
added and the reaction mixture was heated for another 10 min to
yield benzoylthioureido derivative 18 in 70% yield. The structure of
the latter compound was established on the basis of its elemental
analysis and spectral data. For example, the 1H NMR spectrum
2.2.2.2. Influence of lipophilic and steric factors [39,55]. The intro-
duction of spacer linear alkylic chains of increasing number of
carbons between the fused heterocyclic core and arylpiperazine
fragment of the new ligands, cycloalkane fragments fused to thio-
phene moiety and the variation of the substituents on the arylpi-
perazine fragment [5,6,35,40,41] have allowed evaluating the
influence of lipophilicity and steric parameters at the pharmaco-
phoric part of the molecules. Table 1 gathers anatgonist activities at
5-HT2A receptors as well as values of log P (lipophilic factors) and
molar refractometry (steric factors), determined by using Hyper-
Chem program [56] for each compound. Within the series of
compounds 7a–d (n ¼ NMe) we have observed little increase of the
5-HT2A antagonist activities when molar refractometry increases
from 121.2 cm3/mol (R ¼ H) to 128.4 cm3/mol (R ¼ OMe). Such
phenomenon was observed within other series such as compounds
6a and 11d with molar refractometry values 112.3 cm3/mol and
133.8 cm3/mol respectively. Although lipophilicity does not exert
a significant effect on affinity for compounds 7a–d, 6a and 11d,
showed two broad signals, one at
d 9.00–9.10 and the other
appeared at 14.50–14.60 representing the two NH groups. Cycli-
d
zation of the benzoylthioureido derivative 18 in ethanolic
potassium hydroxide afforded the potassium salt of 2-mercapto-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
one 19. The protons of the phenyl ring and ethyl ester in
compound 18 disappeared in the 1H NMR spectrum of compound
19 revealing its cyclization. 1-Chloroacetyl-4-(substituted phe-
nyl)piperazines 20a,b,d,e were treated with compound 19 to yield
2-[[4-substituted phenylpiperazin-1-yl]carbonylmethylthio]-6,7,8,
9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-ones
21a,b,d,e. The structure of the latter compounds was established
on the basis of the elemental analyses and spectral data. For
example, the 1H NMR spectrum showed the appearance of the
protons of the N-phenylpiperazine moiety in the expected regions
of the spectrum.
Table 1
Response % of the tested compounds (mmol/kg) towards inhibition of 5-HT induced pulmonary arterial ring pressor and molecular parameters of some selected compounds.
Compd. no.
Responsea [%]
Log Pb
MRc [cm3/mol]
Compd. no.
Responsea [%]
Log Pb
MRc [cm3/mol]
6a
7a
7c
7d
11a
11d
11e
86
60
63
65
62
88
65
1.5
0.7
0.9
0.5
0.8
0.6
0.9
112.3
121.2
121.6
128.4
126.5
133.8
138.6
12d
17a
17b
17d
21a
21b
21d
Risperidone
66
0
0
2.1
4.9
5.5
4.7
2.6
3.1
2.4
2.7
127.6
145.7
150.3
153.0
127.8
132.4
135.0
114.2
0
65
60
68
100
a
Response % of the tested compounds (mmol/kg) towards inhibition of 5-HT induced pulmonary arterial ring pressor.
Calculated lipophilicity.
Molar refractometry.
b
c

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M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
a remarkable decrease in potency was observed for 17a–d, with
values of log P higher than 4.7. Moreover, a spacer carbon chain
with one carbon atom between the arylpiperazine and heterocyclic
core diminishes the activity as in compound 17, if compared with
compounds 6, 7, 11, 12, and 21. Variations of lipophilicity produced
by cycloalkanone (n ¼ 0, 2, NMe) or aromatic substituents (R ¼ H, Cl
and OMe) did not notably alter 5-HT2A antagonist activities. Simi-
larly, for analogues 21a–d no significant variations in 5-HT2A
antagonist activity were found, pointing out that lipophilicity of the
molecules is not a crucial factor for the activity. Regarding steric
parameters, from the data gathered in Table 1 there is a clear
influence of referactometry on 5-HT2A antagonist activity
compared with lipophilicity for either series of all compounds. The
optimal lipophilicity and steric parameters for the active
compounds was in range of 0.5–3.1 and 112.3–138.6 cm3/mol,
respectively, higher values diminished the activity.
point by the absence of negative eigen value of the Hessian through
frequency calculation [62–65]. Two main conformers were
observed for all compounds under investigation including
extended- and folded-conformers. At this stage, calculations at the
AM1 level were considered in order to determine relative energies
of the extended- and folded-conformers of compounds under
studies. The results show that extended-conformers were more
stable than folded-conformers by 3–4 kcal/mol (Fig. 5).
2.3.2. Flexible alignment and RMS fit overlay
2.3.2.1. Flexible alignment. Flexible alignment is an application for
flexibly aligning small molecules. The method accepts as input
a collection of small molecules with 3D coordinates and computes
a collection of alignments. Each alignment is given a score that
quantifies the quality of the alignment in terms of both internal
strain and overlap of molecular features. Often, atomic-level details
of the structures of pharmaceutically relevant receptors are not
available. In such cases, 3D alignment (or superposition) of putative
ligands can be used to deduce structural requirements for biolog-
ical activity. Methodologies based upon 3D alignment for finding
biologically active ligands generally make use of the qualitative
assumption that if two ligands have similar biological activity and
bind in similar modes, then the bound conformations of the two
ligands align well and inferences can be made about the nature of
the receptor [66,67].
To probe similarity between the three-dimensional structures of
active compounds 6a and 11d, flexible alignments were employed.
Our initial approach was to employ MOE/flexible alignment [68] to
automatically generate superpositions of the compounds under
investigation with minimal user bias. Using MOE/MMFF94, 150
conformers of each compound were generated and minimized with
a distance-dependent dielectric model. A low energy set of 50 was
selected for further analysis. Conformations of compound 6a were
generated using distance geometry and optimized with MMFF94
[69]. Nine low energy, maximally dissimilar structures were
selected for comparison to the other compound 11d. After assigning
MMFF94 charges to all molecules, flexible alignment was employed
to scan and rank overlays of compounds 6a and 11d based on steric,
electrostatic field, hydrophobic areas overlap, hydrogen bond
donors and acceptors overlap. From the top scoring superposition,
several sets most consistent with the structure–activity relation-
ships of the two 5-HT2A antagonist compounds were selected and
subjected to more refined searching using MOE/flexible alignment
module [68], the highest scoring aligned pair is shown in Fig. 6a.
Since both molecules are highly flexible, the limited set of
conformers used in the analysis was not capable of achieving
2.3. Molecular modeling study
Modeling studies are required in order to construct molecular
models that incorporate all experimental evidence reported. These
models are necessary to obtain a consistent, more precise picture of
the biologically active molecules at the atomic level and further-
more, provide new insights that can be used to design novel thera-
peutic agents. Considering ligand–receptor interaction, one usually
takes into account two points: strength of the interaction (what we
refer to as ‘affinity’) and type of an effect at the biochemical, elec-
trophysiological and behavioural level triggered by the ligand (what
we refer to as ‘intrinsic activity’). To get an insight into the type of
chemical forces involved in the interaction, one either builds models
of a specific pharmacophore or constructs models of a receptor
protein and examines the binding forces. Interaction of a ligand with
a receptor may result in receptor activation (agonists), blockade
(antagonists) or inactivation (inverse agonists) [5,6,49,57,58].
2.3.1. Conformational analysis
An attempt to gain a better insight into the molecular structures
of the active compounds 6a, 11d and 21d and inactive compound
17a as a representative example, conformational analysis of the
target compound has been performed by use of the molecular
mechanics (MMþ) [59,60] force-field (calculations in vacuo, bond
dipole option for electrostatics, Polak–Ribiere algorithm and root
mean square (RMS) gradient of 0.01 kcal/Å mol) as implemented in
HyperChem 5.1 [56]. The most stable conformer was fully opti-
mized with AM1 semi-empirical molecular orbital calculation [61].
The global minimum was confirmed as true minimum, not saddle
Fig. 5. Energy minimization of compound 6a as a representative example showing the extended-conformer (left panel) and the folded-conformer (right panel).

M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
1811
Fig. 6. (a, Left) Superposition of the most active compounds 6a (in green) and 11d (in red) using flexible alignments. (b Right) Further refinement of the overlay subjected to the
constraints described in the text.
complete atom to atom superposition. A further refinement was
generated (Fig. 6b) by constraining the functional group mapping
suggested from the initial alignment. The calculated energy
difference between the native conformers derived from the flexible
alignment (Fig. 6a) and the final conformers in Fig. 6b is minimal. In
fact, the refined structure of compound 6a was 0.5 kcal/mol lower
in energy than the starting structure, and compound 11d was
1.0 kcal/mol lower. A common feature of the MOE-generated
alignments is the superposition of the carbonyl group of both
compounds. As shown in Fig. 6b, the carbonyl unit of compound 6a
maps to the carbonyl of the compound 11d (0.8 Å). The aromatic
ring and basic nitrogen of piperazine nucleus align fairly well for
both compounds and almost complete superposition of the thio-
phene rings of compounds 6a and 11d (0.1 Å).
By the same way, 5-HT2A antagonist compounds 6a,11d, 21a and
inactive compound 17a were subjected to flexible alignments. As
shown in Fig. 7, all active compounds have the same feature of
alignment as we described above showing four points of similari-
ties; aromatic thiophene core, carbonyl group directly attached to
thiophene moiety, basic nitrogen of piperazine fragment and
aromatic ring attached to such basic nitrogen. This alignment
suggests the possible existence of a region on the receptor suited
for specific recognition of such features. This hypothesis is also
supported by the known preference for serotonin receptor recog-
nition [45,51]. Analysis of inactive molecule is an important step to
understand the essential features for a given activity. It is clear that
compound 17a was flexibly aligned in a different manner when
compared to the active compounds explaining why such compound
was void of 5-HT2A antagonist activity (Fig. 7).
2.3.2.2. RMS fit overlay (rigid alignment). It is obvious from the
flexible alignment, the occurrence of four points of similarities
among the active compounds. However, from these results alter-
native mappings for all compounds under investigation were also
identified using rigid overlays methodology. The stable extended-
conformers resulting from computational chemistry analysis using
MMþ force-field [59,60] as implemented in HyperChem 5.1 [56]
were superimposed in order to reveal the similarities and differ-
ences in structure (Fig. 8). The strategy of overlay fit was to match
thiophene rings and to examine any spatial differences between the
atoms of the arylpiperazine moieties. As shown in Fig. 8, the
carbonyl unit and thiophene core of all compounds completely
align (0.1–0.3 Å). The aromatic ring and basic nitrogen of piperazine
nucleus for active compounds 6a, 11d and 21a align fairly well (1.2–
2.3 Å). More interestingly, the aromatic ring and the basic nitrogen
of the inactive compound 17a completely deviated from the active
analogues by 9.7 Å and 7.9 Å respectively (Fig. 8). Examination of
Fig. 8 reveals that no element of compound 17a correlates with the
5-HT2A antagonist activity and this is not surprising in the context
of the structure–activity relationships.
2.3.3. Electrostatic and hydrophobic mappings
In an attempt to understand the inactivity of compound 17a and
5-HT2A antagonist activity of 6a, 11d and 21a, electrostatic and
hydrophobic mappings have been carried out for the lowest energy
conformers, to examine the similarity and dissimilarity in elec-
tronic, electrostatic binding characteristics of the surface of the
Fig. 7. Flexible alignment of the active compounds 6a (green), 11d (blue), 21a (red) and
the inactive compound 17a (violet).
Fig. 8. RMS fit overlay (rigid alignment) of the active compounds 6a (green), 11d
(blue), 21a (red) and the inactive compound 17a (violet).

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M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
molecules and conformational properties (Figs.
9
and 10).
contrast, the inactivity of compound 17a may be attributed to the
difference in electrostatic mapping in which the red hydrogen bond
acceptor area was located on the heterocyclic triazole ring (Fig. 10).
Similarly, hydrophobic mappings of active compounds showed
that the hydrophobic region in green was distributed on both sides
of aromatic ring of arylpiperazine moieties and thiophene core
while the hydrophilic region in red was located mainly on carbonyl
groups. On the other hand the hydrophobic distributions of the
Comparison of the electrostatic mappings of 6a, 11d and 21a shows
that increased negative charge regions located on the carbonyl
groups representing hydrogen bond acceptor regions (in red) and
medium polar heterocyclic nitrogen atoms located on the basic
nitrogen of piperazine moieties (in cyan). Such results indicated the
structural similarity among these analogues and so similar bio-
logical activity as evident from the experimental data (Fig. 9). In
Fig. 9. Electrostatic maps (left panels) and hydrophobic maps (right panels) of the lowest energy conformers for the active compounds 6a, 11d and 21a, maps are color coded: red
for a hydrogen bond and a hydrophilic region, cyan for a medium polar region, and green for a hydrophobic region.

M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
1813
Fig. 10. Electrostatic maps (left panels) and hydrophobic maps (right panels) of the lowest energy conformer for the inactive compound 17a, maps are color coded: red for
a hydrogen bond and a hydrophilic region, cyan for a medium polar region, and green for a hydrophobic region.
inactive compound 17a occupy an additional aromatic ring indi-
cating the higher lipohpilicity of such compound while the
hydrophilic red region was located on the triazole ring.
2.3.4. Pharmacophore prediction [45,51,70]
For pharmacophore generation and prediction, compound 11d
(Fig. 11) served as the reference to which all conformations of each
analogue were aligned. All structures were built de novo using 2D/
3D editor sketcher in MOE [68]. Conformational models were
calculated using a 15 kcal energy cut off (minimization convergence
It is clear that the related charge distribution, electrostatic and
hydrophobic mappings suggest a similar interaction of the mole-
cules with a potential protein-binding site. As a result, we find that
compounds 6a, 11d and 21a all can adopt conformations that have
identical distances and orientations of the protein-binding site and
that show sufficient agreement of the overall structure and of the
electrostatic potential pattern. Consequently these conformations
permit approach to a protein surface in much the same way con-
necting to a binding site that fits all three ligands (Table 2). They can
thus be regarded as the active binding conformations. Further
reorientation at the site will lead to an induced fit different for the
specific ligands and responsible for variations in their binding
affinity.
criteria
during
conformational
analysis:
energy
con-
vergence ¼ 0.01 kcal/mol, gradient convergence ¼ 0.01 kcal/mol).
The number of conformers generated for each substrate was
limited to a maximum of 150. All molecules with their associated
conformations were regrouped including the biological data.
Hypothesis generation was performed using low energy
conformers of the molecules. After assignment of possible phar-
macophore elements for each analogue the calculation and analysis
were carried out using the MOE program [68] and a superposition
of the molecules, including the assigned elements, was attempted
(Figs. 11 and 12). Several runs of calculation were repeated to
generate pharmacophore maps. For each run a distinct number of
specified pharmacophore elements were adapted. All adapted
models showed that the acceptor atoms of the carbonyl and basic
nitrogen fragments as hydrophilic element and the aromatic rings
as hydrophobic element were well superimposed within the set
distance tolerance. This confirms the important role of the hydro-
philic and hydrophobic moieties for recognition and binding to
receptor sites.
Table 2
Proposed pharmacophore model for 5-HT2A receptor antagonists and structural
parameters of the lowest energy extended-conformers of analyzed compounds.
Basic nitrogen
HB Acceptor oxygen
d
Hydrophobic Aromatic
N
d
O
Models for 11d and its analogues 6a and 21a (Fig. 11, Table 2)
possess pharmacophore elements in the piperazine basic nitrogen
which is out of the thiophene plane as well as carbonyl moiety in
the same plane of thiophene core. In contrast, model for 17a
(Fig. 12, Table 2) showed common elements only in carbonyl
oxygen and thiophene core fused to non-favourable triazole frag-
ment. Since the arylpiperazine part of compound 11d plays an
important role in binding, model 17a was excluded from further
considerations. Therefore, model 11d was considered as the
representative pharmacophore map of the antagonism fitting the
binding site.
According to the pharmacophore generated by MOE [68] and
DSV [71], the minimal structural requirements for 5-HT2A antago-
nism consist of an aromatic ring, a basic nitrogen atom, an H-
bonding acceptor group and a hydrophobic region directly attached
to the basic center (Table 2). For all the active molecules, reasonable
nonbonded distances among the basic nitrogen, carbonyl group
and the thiophene centroid as well as bond angles between the
plane of the thiophene core and the basic nitrogen aligned on the
d
t
(P
la
n
e
a
n
g
e
l
)
Ring Aromatic
a
b
c
Compd.
d1
(Å)
7.0
d2
(Å)
d3
Plane angled
(tꢀ)
(Å)
6a
3.5
3.4
3.1
3.8
8.8
7.4
8.2
8.2
142.4
152.9
123.3
141.0
11d
17a
21a
6.6
10.3
7.2
a
Distance between the basic nitrogen and the oxygen of the carbonyl group.
Distance between the centroid of the ring aromatic and the oxygen of the
b
carbonyl group.
c
Distance between the basic nitrogen and the centroid of the hydrophobic
aromatic.
d
Angle between the basic nitrogen and the plane of the ring aromatic.

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M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
Fig. 11. The active compounds 6a, 11d and 21a, mapped to the pharmacophore model for 5-HT2A receptor antagonists. Pharmacophore features are color coded: brown and yellow
for hydrophobics aromatic, cyan for a hydrogen bond acceptor, and blue for a positive ionisable feature.
predicted pharmacophore maps were found (Table 2). This phar-
macophoric assumption was in consistence with reported results
for 5-HT2A antagonist activity [39–41,45,49,51,57,59].
[2,3-d]pyrimidine-2,4(1H,3H)-dione 6a and ethyl 2-[[4-(2-methoxy-
phenyl)piperazin-1-yl]acetylamino]-4,5,6,7-tetrahydro-6-methylthi-
eno[2,3-c]pyridine-3-carboxylate 11d, which have shown higher
antiserotonergic activities than all the other tested compounds.
Flexible alignment was conducted on the basis of experimental
data from which four featured pharmacophore model was devel-
oped. The model was mapped onto the 5-HT2A antagonist mole-
cules and compared with nonactive compounds. This model
justifies the importance of the main pharmacophore groups
3. Conclusion
The present work led to the development of novel 5-HT2A
antagonist molecules and two of the novel agents namely, 3-[2-
[4-phenylpiperazin-1-yl]ethyl]-6,7-dihydro-5H-cyclopenta[b]thieno-

M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
1815
Fig. 12. The inactive compound 17a, mapped to the pharmacophore model for 5-HT2A receptor antagonists. Pharmacophore features are color coded: brown and yellow for
hydrophobic aromatics, cyan for a hydrogen bond acceptor, and blue for a positive ionisable feature.
(particularly, the carbonyl fragment and the basic nitrogen atom) as
well as of their relative distance. In fact, while the substitution
pattern on the phenyl ring considerably affects the activity (with
the o-methoxy substituent associated with the best activity),
spatial considerations are particularly important for the basic
nitrogen atom, since its spatial locations in regard to the plane of
thiophene core are critical for the biological activity. Therefore, our
pharmacophore model could be very useful for the virtual
screening in the development of new 5-HT2A antagonists.
2.90 (m, 4H, 2CH2), 3.60–3.72 (m, 4H, CH2CH2Cl), 4.20–4.35 (q, 2H,
OCH2CH3), 5.50–5.70 (br s,1H, NH, D2O exchang.),10.25–10.30 (br s,
1H, NH, D2O exchang.). Anal. Calcd. for C13H17ClN2O3S: C, 49.29; H,
5.41; N, 8.84. Found: C, 49.60; H, 5.38; N, 9.25.
4.2.2. Ethyl 2-[3-(2-chloroethyl)ureido]-4,5,6,7-tetrahydro-6-
methylthieno[2,3-c]pyridine-3-carboxylate 5
Brown powder (2.07 g, 60%) m.p. 219–220 ꢀC. 1H NMR (DMSO-
d6);
d 1.30–1.40 (t, 3H, OCH2CH3), 2.70–3.00 (m, 7H, 2CH2, NCH3),
3.40 (s, 2H, CH2), 3.60–3.80 (m, 4H, CH2CH2Cl), 4.50–4.60 (q, 2H,
OCH2CH3), 6.30 (s, 1H, NH, D2O exchang.),10.30–10.50 (br s,1H, NH,
D2O exchang.). MS m/z (%) 347, (Mþ þ 1, 20.99), 346 (Mþ, 30.09),
308 (18.51), 240 (15.84), 239 (64.16), 125 (100), 122 (17.53). Anal.
Calcd. for C14H20ClN3O3S: C, 48.62; H, 5.83; N, 12.15. Found: C,
48.94; H, 5.45; N, 12.48.
4. Experimental
4.1. General
Melting points were measured with a Fisher–Johns melting point
apparatus and are uncorrected. IR spectra were recorded on Matt-
son 5000 FT-IR spectrometer. 1H NMR spectra were determined in
CDCl3 or DMSO-d6 on FT-NMR spectrometer (200 MHz) Gemini
Varian using TMS as internal standard. Mass spectra were
measured on JEOL JMS-600H spectrometer. Elemental analysis was
carried out at the Microanalytical Center of Cairo University.
o-Amino esters 1–3,13 [52–54], ethyl 2-[(chloroacetyl)amino]-
4.3. 3-[2-[4-Substituted phenylpiperazin-1-yl]ethyl]-6,7-dihydro-
5H-cyclopenta[b]thieno[2,3-d]pyrimidine-2,4-(1H,3H)-diones and
3-[2-[4-substituted phenylpiperazin-1-yl]ethyl]-5,6,7,8-tetrahydro-
7-methylpyrido[40,30:4,5]thieno[2,3-d]pyrimidine-2,4-(1H,3H)-
diones 6,7a–d (Scheme 1)
5,6,7,8-tetrahydro-4H-cyclohepta[b]thiophene-3-carboxylate
8
A mixture of the ureido derivative 4 or 5 (5 mmol) and the
appropriate N-substituted phenylpiperazine (25 mmol) was
heated in an oil bath for 6–8 h at 140 ꢀC. After cooling, the
reaction mixture was treated with a small amount of ethanol and
the solid obtained was filtered off. The filtrate was evaporated
under reduced pressure and the solid obtained was crystallized
from DMF. The sticky product, compound 6b was chromato-
graphed on silica gel and eluted with ethyl acetate:petroleum
ether (1:4).
[54], 2-mercapto-3-phenyl-5,6,7,8-tetrahydrobenzo[b]thieno[2,3-
d]pyrimidin-4(3H)-one 14 [72,73], 2-hydrazino-3-phenyl-5,6,7,8-
tetrahydrobenzo[b]thieno[2,3-d]pyrimidin-4(3H)-one 15 [74] and
1-chloroacetyl-4-substituted phenylpiperazines 20 [75–77] were
prepared following the procedures reported in the literature.
4.2. Ethyl 2-[3-(2-chloroethyl)ureido]-3-carboxylates 4,5 (Scheme 1)
2-Chloroethyl isocyanate (1.58 g, 15 mmol) was added to
a suspension of 1 or 3 (10 mmol) in hot toluene (20 ml) and the
reaction mixture was heated under reflux for 6 h. The solvent was
then evaporated under reduced pressure. The residue of compound
4 was chromatographed on silica gel and eluted with ethyl aceta-
te:petroleum ether (1:4) to give the ureido derivative 4. The residue
of compound 5 was triturated with a little amount of ethanol and
the separated solid was filtered, dried and crystallized from water.
4.3.1. 3-[2-[4-Phenylpiperazin-1-yl]ethyl]-6,7-dihydro-5H-
cyclopenta[b]thieno[2,3-d]pyrimidine-2,4-(1H,3H)-dione 6a
Yellow crystal (0.79 g, 40%) m.p. 132–133 ꢀC. IR (KBr) nmax/cmꢁ1
3210, 3150 (NH) and 1647, 1640 (C]O). MS m/z (%); 396 (Mþ, 0.54),
335 (0.57), 207 (0.37), 189 (3.26), 162 (24.74), 120 (100), 56 (22.38).
Anal. Calcd. for C21H24N4O2S: C, 63.61; H, 6.10; N, 14.13. Found: C,
63.35; H, 6.45; N, 14.54.
4.2.1. Ethyl 2-[3-(2-chloroethyl)ureido]-5,6-dihydro-4H-
4.3.2. 3-[2-[4-(2-Chlorophenyl)piperazin-1-yl]ethyl]-6,7-dihydro-
5H-cyclopenta[b]thieno[2,3-d]pyrimidine-2,4-(1H,3H)-dione 6b
Yellow powder (0.71 g, 33%) m.p. 120–122 ꢀC. 1H NMR (CDCl3);
cyclopenta[b]thiophene-3-carboxylate 4
White shinny needles (1.26 g, 40%) mp 118–119 ꢀC. IR (KBr) nmax
/
cmꢁ1 3220, 3145 (NH), 1740 (COOC2H5), 1640 (C]O). 1H NMR
(CDCl3); 1.20–1.40 (t, 3H, OCH2CH3), 2.25–2.43 (m, 2H, CH2), 2.70–
d
2.30–2.40 (m, 2H, CH2), 2.80–2.92 (m, 4H, 2CH2), 3.20–3.80 (m,
d
12H, 3NCH2, 2Ar-NCH2, CONCH2), 6.90–7.50 (m, 4H, Ar-H), 10.41 (s,

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M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
1H, NH, D2O exchang.). Anal. Calcd. for C21H23ClN4O2S: C, 58.53; H,
5.38; N, 13.00. Found: C, 58.21; H, 5.40; N, 12.60.
crystallized from ethanol to afford yellow crystal of compound 9 in
3.17 g (50%) yield; m.p. 230–232 ꢀC; 1H NMR (DMSO-d6);
1.20–
d
1.30 (t, 3H, OCH2CH3), 2.50–2.80 (m, 7H, 2CH2, NCH3), 3.20 (s, 2H,
CH2), 4.20–4.40 (q, 2H, OCH2CH3), 4.65 (s, 2H, CH2Cl), 11.85–11.90
(br s, 1H, NH, D2O exchang.). MS m/z (%); 319 (Mþ þ 2, 2.30), 318
(Mþþ, 16.20), 317 (Mþ, 7.80), 282 (0.71), 269 (100), 240 (6.60), 224
(0.90). Anal. Calcd. for C13H17ClN2O3S: C, 49.29; H, 5.41; N, 8.84.
Found: C, 48.98; H, 5.80; N, 9.00.
4.3.3. 3-[2-[4-(4-Fluorophenyl)piperazin-1-yl]ethyl]-6,7-dihydro-
5H-cyclopenta[b]thieno[2,3-d]pyrimidine-2,4-(1H,3H)-dione 6c
Brown crystal (0.76 g, 37%) m.p. 133–135 ꢀC. 1H NMR (DMSO-d6);
d
2.35–2.40 (m, 2H, CH2), 2.70–2.90 (m, 4H, 2CH2), 3.00–3.70 (m,10H,
3NCH2, 2Ar-NCH2), 3.90–4.10 (t, 2H, CONCH2), 6.90–7.20 (m, 4H, Ar-
H), 10.70 (s, 1H, NH, D2O exchang.). Anal. Calcd. For C21H23FN4O2S: C,
60.85; H, 5. 59; N, 13.52. Found: C, 61.25; H, 5.41; N, 13.92.
4.5. Ethyl 2-[[4-substituted phenylpiperazin-1-yl]acetylamino]-
5,6,7,8-tetrahydro-4H-cyclohepta[b]thiophene-3-carboxylates
10a,b,d,e and ethyl 2-[[4-substituted phenylpiperazin-1-
yl]acetylamino]-4,5,6,7-tetrahydro-6-methylthieno[2,3-c]pyridine-
3-carboxylates 11a,b,d,e (Scheme 1)
4.3.4. 3-[2-[4-(2-Methoxyphenyl)piperazin-1-yl]ethyl]-6,7-
dihydro-5H-cyclopenta[b]thieno[2,3-d]pyrimidine-2,4-(1H,3H)-
dione 6d
White powder (0.63 g, 30%) m.p. 130–132 ꢀC. 1H NMR (CDCl3);
d
2.30–2.40 (m, 2H, CH2), 2.75–2.90 (m, 4H, 2CH2), 3.00–3.15 (m,
A mixture of 8 or 9 (3.9 mmol), the appropriate N-substituted
phenylpiperazine (3.9 mmol) and potassium carbonate (0.539 g,
3.9 mmol) in DMF (10 ml) was heated under reflux for 5 h. The
solvent was evaporated under reduced pressure and the residue
thus obtained was triturated with water, filtered and dried to give
compounds 10 which crystallized from DMF/water and compounds
11 which crystallized from ethanol.
6H, 3NCH2), 3.30–3.50 (m, 4H, 2Ar-NCH2), 3.55–3.65 (t, 2H,
CONCH2), 3.80 (s, 3H, OCH3), 6.88–7.20 (m, 4H, Ar-H), 10.30 (s, 1H,
NH, D2O exchang.). Anal. Calcd. for C22H26N4O3S: C, 61.95; H, 6.14;
N, 13.41. Found: C, 62.35; H, 5.80; N, 13.50.
4.3.5. 3-[2-[4-Phenylpiperazin-1-yl]ethyl]-5,6,7,8-tetrahydro-7-
methylpyrido[40,30:4,5]thieno[2,3-d]pyrimidine-2,4-(1H,3H)-dione 7a
White powder (0.74 g, 35%) m.p. 160–161 ꢀC. IR (KBr) nmax/cmꢁ1
4.5.1. Ethyl 2-[[4-phenylpiperazin-1-yl]acetylamino]-5,6,7,8-
tetrahydro-4H-cyclohepta[b]thiophene-3-carboxylate 10a
White needles (1.19 g, 70%) m.p. 160–162 ꢀC. 1H NMR (CDCl3);
3220, 3160 (NH), 1648, 1635 (C]O). 1H NMR (CDCl3);
d 2.30–3.00
(m, 13H, 2CH2, NCH3, 3NCH2), 3.20–3.55 (m, 6H, CH2, 2Ar-NCH2),
3.70–3.80 (t, 2H, CONCH2), 6.70–7.30 (m, 5H, Ar-H),10.90 (s,1H, NH,
D2O exchang.). Anal. Calcd. for C22H27N5O2S: C, 62.09; H, 6.40; N,
16.46. Found: C, 61.70; H, 6.05; N, 16.96.
d
1.33–1.36 (t, 3H, OCH2CH3), 1.58–1.66 (m, 4H, 2CH2), 1.80–1.87 (m,
2H, CH2), 2.70–2.80 (m, 6H, CH2, 2NCH2), 3.00–3.10 (t, 2H, CH2),
3.29–3.40 (m, 6H, COCH2, 2Ar-NCH2), 4.30–4.39 (q, 2H, OCH2CH3),
6.80–7.30 (m, 5H, Ar-H),12.10 (s,1H, NH, D2O exchang.). Anal. Calcd.
for C24H31N3O3S: C, 65.28; H, 7.08; N, 9.52. Found: C, 65.15; H, 6.80;
N, 9.99.
4.3.6. 3-[2-[4-(2-Chlorophenyl)piperazin-1-yl]ethyl]-5,6,7,8-
tetrahydro-7-methylpyrido[40,30:4,5]thieno[2,3-d]pyrimidine-2,4-
(1H,3H)-dione 7b
Yellow crystal (0.75 g, 33%) m.p.118–119 ꢀC. 1H NMR (DMSO-d6);
2.60–3.25 (m, 13H, 2CH2, NCH3, 3NCH2), 3.30–3.65 (m, 6H, CH2,
4.5.2. Ethyl 2-[[4-(2-chlorophenyl)piperazin-1-yl]acetylamino]-
5,6,7,8-tetrahydro-4H-cyclohepta[b]thiophene-3-carboxylate 10b
White needles (1.38 g, 75%) m.p. 115–116 ꢀC. 1H NMR (CDCl3);
d
2Ar-NCH2), 3.90–4.00 (t, 2H, CONCH2), 6.80–7.40 (m, 4H, Ar-H),
11.20 (s, 1H, NH, D2O exchang.). Anal. Calcd. for C22H26ClN5O2S: C,
57.44; H, 5.70; N, 15.22. Found: C, 57.00; H, 5.50; N, 15.65.
d
1.33–1.35 (t, 3H, OCH2CH3), 1.56–1.62 (m, 4H, 2CH2), 1.80–1.87 (m,
2H, CH2), 2.69–2.75 (m, 6H, CH2, 2NCH2), 3.00–3.10 (t, 2H, CH2),
3.27–3.41 (m, 6H, COCH2, 2Ar-NCH2), 4.20–4.40 (q, 2H, OCH2CH3),
6.88–7.40 (m, 4H, Ar-H), 12.30 (s, 1H, NH, D2O exchang.). Anal.
Calcd. for C24H30ClN3O3S: C, 60.55; H, 6.35; N, 8.83. Found: C, 60.91;
H, 6.00; N, 9.30.
4.3.7. 3-[2-[4-(4-Fluorophenyl)piperazin-1-yl]ethyl]-5,6,7,8-
tetrahydro-7-methylpyrido[40,30:4,5]thieno[2,3-d]pyrimidine-2,4-
(1H,3H)-dione 7c
Yellowish white crystal (0.88 g, 40%) m.p. 120–122 ꢀC. MS m/z
(%); 446 (Mþ þ 2, 1.43), 264.5 (14.00), 237 (3.56), 236 (6.66), 192
(100), 179 (5.95), 151 (9.90). Anal. Calcd. for C22H26FN5O2S: C, 59.57;
H, 5.91; N, 15.79. Found: C, 59.80; H, 6.23; N, 15.30.
4.5.3. Ethyl 2-[[4-(2-methoxyphenyl)piperazin-1-yl]acetylamino]-
5,6,7,8-tetrahydro-4H-cyclohepta[b]thiophene-3-carboxylate 10d
White crystal (1.38 g, 75%) m.p. 133–135 ꢀC. IR (KBr) nmax/cmꢁ1
3240, 3210 (NH), 1750 (COOC2H5), 1630 (C]O). MS m/z (%); 473
(Mþ þ 1, 3.65), 472 (Mþ, 7.70), 282 (1.80), 233 (2.00), 225 (2.70), 205
(31.30), 150 (100). Anal. Calcd. for C25H33N3O4S: C, 63.67; H, 7.05; N,
8.91. Found: C, 63.45; H, 6.60; N, 9.15.
4.3.8. 3-[2-[4-(2-Methoxyphenyl)piperazin-1-yl]ethyl]-5,6,7,8-
tetrahydro-7-methylpyrido[40,30:4,5]thieno[2,3-d]pyrimidine-2,4-
(1H,3H)-dione 7d
Yellow crystal (0.67 g, 30%) m.p. 130–132 ꢀC. 1H NMR (DMSO-
d6);
d
2.30–2.90 (m, 13H, 2CH2, NCH3, 3NCH2), 3.10–3.40 (m, 6H,
4.5.4. Ethyl 2-[[4-(2-ethoxyphenyl)piperazin-1-yl]acetylamino]-
5,6,7,8-tetrahydro-4H-cyclohepta[b]thiophene-3-carboxylate 10e
Yellowish white needles (1.36 g, 73%) m.p. 130–132 ꢀC. 1H NMR
CH2, 2Ar-NCH2), 3.60–3.70 (t, 2H, CONCH2), 3.80 (s, 3H, OCH3),
6.80–7.40 (m, 4H, Ar-H), 10.90–11.00 (br.s, 1H, NH, D2O exchang.).
Anal. Calcd. for C23H29N5O3S: C, 60.64; H, 6.42; N, 15.73. Found: C,
60.95; H, 6.10; N, 15.87.
(CDCl3);
d
1.33–1.38 (m, 6H, 2ꢀ CH2CH3), 1.55–1.69 (m, 4H, 2CH2),
1.80–1.87 (m, 2H, CH2), 2.70–2.79 (m, 6H, CH2, 2NCH2), 3.00–3.09 (t,
2H, CH2), 3.28–3.36 (m, 6H, COCH2, 2Ar-NCH2), 4.30–4.38 (m, 4H, 2ꢀ
CH2CH3), 6.89–7.42 (m, 4H, Ar-H), 12.32 (s, 1H, NH, D2O exchang.).
Anal. Calcd. for C26H35N3O4S: C, 64.30; H, 7.26; N, 8.62. Found: C,
64.33; H, 7.11; N, 9.00.
4.4. Ethyl 2-(chloroacetylamino)-4,5,6,7-tetrahydro-6-
methylthieno[2,3-c]pyridine-3-carboxylate 9 (Scheme 1)
Chloroacetyl chloride (2.25 g, 20 mmol) was added dropwise
under cooling to a solution of ethyl 2-amino-4,5,6,7-tetrahydro-6-
methylthieno[2,3-c]pyridine-3-carboxylate 3 (4.80 g, 20 mmol) in
chloroform (40 ml). The reaction mixture was heated under reflux
for 6 h. The separated solid was filtered while hot dried and
4.5.5. Ethyl 2-[[4-phenylpiperazin-1-yl]acetylamino]-4,5,6,7-
tetrahydro-6-methylthieno[2,3-c]pyridine-3-carboxylate 11a
White powder (0.68 g, 40%) m.p. 117–118 ꢀC. MS m/z (%); 444
(Mþ þ 1, 1.58), 443 (Mþ, 5.05), 442 (Mþ ꢁ 1, 17.52), 282 (3.13), 241

M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
1817
(1.27), 175 (100), 161 (2.20). Anal. Calcd. for C23H30N4O3S: C, 62.42;
H, 6.83; N, 12.66. Found: C, 62.10; H, 6.65; N, 13.00.
193 (31.02). Anal. Calcd. For C23H29N5O2S: C, 62.84; H, 6.65; N,
15.93. Found: C, 62.65; H, 6.40; N, 16.30.
4.5.6. Ethyl 2-[[4-(2-chlorophenyl)piperazin-1-yl]acetylamino]-
4,5,6,7-tetrahydro-6-methylthieno[2,3-c]pyridine-3-carboxylate 11b
Yellowish white crystal (0.72 g, 39%) m.p. 120–121 ꢀC. 1H NMR
4.6.4. 3-Amino-2-[[4-(2-ethoxyphenyl)piperazin-1-yl]methyl]-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
one 12e
(CDCl3);
d
1.20–1.30 (t, 3H, OCH2CH3), 2.40–2.90 (m, 11H, 2CH2,
Brown powder (0.49 g, 45%) m.p. 150–152 ꢀC. 1H NMR (CDCl3);
NCH3, 2NCH2), 3.00–3.65 (m, 8H, CH2, COCH2, 2Ar-NCH2), 4.20–4.40
(q, 2H, OCH2CH3), 6.90–7.40 (m, 4H, Ar-H), 12.20–12.40 (br s, 1H,
NH, D2O exchang.). Anal. Calcd. for C23H29ClN4O3S: C, 57.91; H, 6.13;
N, 11.75. Found: C, 58.22; H, 6.44; N, 12.00.
d 1.42–1.48 (t, 3H, OCH2CH3), 1.66–1.74 (m, 4H, 2CH2), 1.86–1.93
(m, 2H, CH2), 2.71–2.88 (m, 6H, CH2, 2NCH2), 3.00–3.20 (t, 2H,
CH2), 3.26–3.34 (m, 4H, 2Ar-NCH2), 3.81 (s, 2H, CH2 attached to
piperazine moiety), 4.00–4.10 (q, 2H, OCH2CH3), 6.40–6.49 (br s,
2H, NH2, D2O exchang.), 6.83–7.0 (m, 4H, Ar-H). Anal. Calcd. For
C24H31N5O2S: C, 63.55; H, 6.89; N, 15.44. Found: C, 63.10; H, 7.00;
N, 15.86.
4.5.7. Ethyl 2-[[4-(2-methoxyphenyl)piperazin-1-yl]acetylamino]-
4,5,6,7-tetrahydro-6-methylthieno[2,3-c]pyridine-3-carboxylate 11d
White crystal (0.65 g, 36%) m.p. 125–127 ꢀC. IR (KBr) nmax/cmꢁ1
3210, 3190 (NH), 1745 (COOC2H5), 1640 (C]O). MS m/z (%); 473
(Mþ, 0.53), 472 (Mþ ꢁ 1, 1.89), 367 (1.07), 282 (0.40), 240 (1.52), 233
(0.71), 150 (100). Anal. Calcd. for C24H32N4O4S: C, 60.99; H, 6.82; N,
11.86. Found: C, 60.65; H, 6.59; N, 12.10.
4.7. 1-Chloromethyl-4-phenyl-6,7,8,9-tetrahydro-
[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-e]pyrimidin-5(4H)-one 16
(Scheme 2)
4.5.8. Ethyl 2-[[4-(2-ethoxyphenyl)piperazin-1-yl]acetylamino]-
4,5,6,7-tetrahydro-6-methylthieno[2,3-c]pyridine-3-carboxylate 11e
White crystal (0.75 g, 40%) m.p. 110–111 ꢀC. 1H NMR (CDCl3);
Chloroacetyl chloride (5.64 g, 0.05 mol) was added dropwise
under cooling to a solution of 15 (7.80 g, 0.025 mol) in chloroform
(30 ml). The reaction mixture was heated under reflux for 4 h.
The separated solid was filtered, dried and crystallized from
ethanol to give red crystal of compound 16 in 4.16 g (45%); m.p.
253–255 ꢀC. IR (KBr) nmax/cmꢁ1 1640 (C]O), 1552, 1503 (C]N).
d
1.30–1.42 (m, 6H, 2ꢀ CH2CH3), 2.20–3.00 (m, 11H, 2CH2, NCH3,
2NCH2), 3.20–3.70 (m, 8H, CH2, COCH2, 2Ar-NCH2), 4.30–4.48 (m,
4H, 2ꢀ CH2CH3), 6.99–7.55 (m, 4H, Ar-H), 12.21–12.30 (br s, 1H, NH,
D2O exchang.). Anal. Calcd. for C25H34N4O4S: C, 61.70; H, 7.04; N,
11.51. Found: C, 61.49; H, 7.11; N, 11.90.
1H NMR (DMSO-d6);
d 1.75–1.82 (m, 4H, 2CH2), 2.80–2.90 (m, 4H,
2CH2), 4.65 (s, 2H, CH2Cl), 7.50–7.70 (m, 5H, Ar-H). MS m/z (%);
372 (Mþ þ 1, 6.87), 371 (Mþ, 5.63), 322 (2.53), 295 (1.12), 283
(7.06), 206 (0.15), 57 (100). Anal. Calcd. for C18H15ClN4OS
(370.86): C, 58.30; H, 4.08; N, 15.11. Found: C, 58.00; H, 4.54; N,
15.60.
4.6. 3-Amino-2-[[4-substituted phenylpiperazin-1-yl]methyl]-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
ones 12a,b,d,e (Scheme 1)
A mixture of ethyl 2-[[[4-(substituted phenyl)piperazin-1-yl]a-
cetyl]amino]-5,6,7,8-tetrahydro-4H-cyclohepta[b]thiophene-3-
carboxylate 10 (2.1 mmol) and 99% hydrazine hydrate (4.0 g,
80 mmol) in absolute ethanol (10 ml) was heated under reflux for
12 h. The separated solid was filtered while hot, dried and crys-
tallized from DMF/water to afford compounds 12.
4.8. 1-[[4-Substituted phenylpiperazin-1-yl]methyl]-4-phenyl-
6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-
e]pyrimidin-5(4H)-ones 17a–e (Scheme 2)
A
mixture of 1-chloromethyl-4-phenyl-6,7,8,9-tetrahydro-
[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-e]pyrimidin-5(4H)-one 16
(1.44 g, 3.9 mmol), the appropriate N-substituted phenylpiperazine
(3.9 mmol) and potassium carbonate (0.539 g, 3.9 mmol) in DMF
(10 ml) was heated under reflux for 6 h. The solvent was evaporated
under reduced pressure. The residue thus formed was triturated
with water, filtered, dried and crystallized from aqueous ethanol to
afford compounds 17a,c,d or crystallized from DMF to afford
compounds 17b,e.
4.6.1. 3-Amino-2-[[4-phenylpiperazin-1-yl]methyl]-6,7,8,9-
tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-one 12a
Yellowish white needles (0.38 g, 50%) m.p. 170–172 ꢀC. IR (KBr)
nmax/cmꢁ1 3280 (NH2), 1632 (C]O). 1H NMR (CDCl3);
d 1.60–1.75
(m, 4H, 2CH2), 1.83–1.94 (m, 2H, CH2), 2.70–2.90 (m, 6H, CH2,
2NCH2), 3.10–3.36 (m, 6H, CH2, 2Ar-NCH2), 3.80 (s, 2H, CH2 attached
to piperazine moiety), 6.30–6.39 (br s, 2H, NH2, D2O exchang.),
6.90–7.30 (m, 5H, Ar-H). Anal. Calcd. for C22H27N5OS: C, 64.52; H,
6.64; N, 17.10. Found: C, 64.92; H, 6.50; N, 17.60.
4.8.1. 1-[[4-Phenylpiperazin-1-yl]methyl]-4-phenyl-6,7,8,9-
tetrahydro-[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-e]pyrimidin-
5(4H)-one 17a
Brown powder (0.77 g, 40%) m.p. 133–135 ꢀC. MS m/z (%); 499
(Mþ þ 2, 0.59), 498 (Mþ þ 1, 1.41), 421 (0.44), 336 (10.69), 161
(14.03), 145 (100), 77 (20.49). Anal. Calcd. for C28H28N6OS: C, 67.72;
H, 5.68; N, 16.92. Found: C, 67.23; H, 5.98; N, 17.30.
4.6.2. 3-Amino-2-[[4-(2-chlorophenyl)piperazin-1-yl]methyl]-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
one 12b
White powder (0.41 g, 45%) m.p. 140–143 ꢀC. 1H NMR (CDCl3);
d
1.65–1.75 (m, 4H, 2CH2), 1.87–1.93 (m, 2H, CH2), 2.75–2.87 (m, 6H,
CH2, 2NCH2), 3.00–3.12(t, 2H, CH2), 3.24–3.35(m, 4H, 2Ar-NCH2), 3.81
(s, 2H, CH2 attached to piperazine moiety), 6.42–6.48 (br s, 2H, NH2,
D2Oexchang.), 6.84–7.00(m, 4H, Ar-H).Anal. Calcd.forC22H26ClN5OS:
C, 59.51; H, 5.90; N, 15.77. Found: C, 59.80; H, 6.25; N, 16.15.
4.8.2. 1-[[4-(2-Chlorophenyl)piperazin-1-yl]methyl]-4-phenyl-
6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-
e]pyrimidin-5(4H)-one 17b
Yellow crystal (1.13 g, 55%) m.p. 311–312 ꢀC. 1H NMR (CDCl3);
4.6.3. 3-Amino-2-[[4-(2-methoxyphenyl)piperazin-1-yl]methyl]-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
one 12d
White powder (0.38 g, 42%) m.p. 160–162 ꢀC. MS m/z (%); 439.45
(Mþ, 6.53), 411 (100), 250 (2.45), 237 (18.57), 221 (8.16), 209 (1.63),
d 1.75–1.90 (m, 4H, 2CH2), 2.70–3.00 (m, 8H, 2CH2 and 2NCH2),
3.20–3.40 (m, 4H, 2Ar-NCH2), 4.15 (s, 2H, CH2 attached to pipera-
zine moiety), 6.90–7.90 (m, 9H, Ar-H). Anal. Calcd. for
C28H27ClN6OS: C, 63.30; H, 5.12; N, 15.82. Found: C, 63.00; H, 5.50;
N, 16.20.

1818
M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
4.8.3. 1-[[4-(4-Fluorophenyl)piperazin-1-yl]methyl]-4-phenyl-
6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-
e]pyrimidin-5(4H)-one 17c
4.11. 2-[[4-Substituted phenylpiperazin-1-yl]carbonylmethylthio]-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
ones 21a,b,d,e (Scheme 2)
Yellow powder (0.78 g, 39%) m.p. 290–292 ꢀC. 1H NMR
(CDCl3);
d
1.75–1.90 (m, 4H, 2CH2), 2.65–2.90 (m, 8H, 2CH2 and
1-Chloroacetyl-4-(substituted phenyl)piperazine derivative 20
(3.6 mmol) was added to a suspension of compound 19 (0.87 g,
3 mmol) in DMF (15 ml) and the reaction mixture was heated under
reflux for 10 h. After cooling, the separated solid was collected by
filtration, dried and crystallized from DMF.
2NCH2), 3.20–3.30 (m, 4H, 2Ar-NCH2), 3.75 (s, 2H, CH2 attached
to piperazine moiety), 7.00–7.80 (m, 9H, Ar-H). Anal. Calcd. for
C28H27 FN6OS: C, 65.35; H, 5.29; N, 16.33. Found: C, 65.20; H,
5.59; N, 16.70.
4.8.4. 1-[[4-(2-Methoxyphenyl)piperazin-1-yl]methyl]-4-phenyl-
6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-
e]pyrimidin-5(4H)-one 17d
4.11.1. 2-[[4-Phenylpiperazin-1-yl]carbonylmethylthio]-6,7,8,9-
tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-ones
21a
Brown crystal (0.86 g, 42%) m.p. 301–303 ꢀC. 1H NMR (DMSO-
Yellow powder (0.65 g, 40%) m.p. 245–246 ꢀC. 1H NMR (DMSO-
d6);
d
1.50–1.80 (m, 4H, 2CH2), 2.70–2.80 (m, 8H, 2CH2 and 2NCH2),
d6); d 1.47–1.60 (m, 4H, 2CH2), 1.75–1.82 (m, 2H, CH2), 2.67–2.75 (t,
2.95–3.10 (m, 4H, 2Ar-NCH2), 3.80 (s, 2H, CH2 attached to pipera-
zine moiety) 3.90 (s, 3H, OCH3), 6.70–7.60 (m, 9H, Ar-H). Anal.
Calcd. For C29H30N6O2S: C, 66.14; H, 5. 74; N, 15.96. Found: C, 66.44;
H, 5.54; N, 16.40.
2H, CH2), 3.05–3.25 (m, 6H, CH2, 2NCH2), 3.56–3.71 (m, 4H, 2Ar-
NCH2), 4.23 (s, 2H, CH2CO), 6.74–7.30 (m, 5H, Ar-H), 12.59–12.63 (br
s, 1H, NH, D2O exchang.). Anal. Calcd. for C23H26N4O2S2: C, 60.77; H,
5.76; N, 12.32. Found: C, 61.10; H, 5.99; N, 12.00.
4.8.5. 1-[[4-(2-Ethoxyphenyl)piperazin-1-yl]methyl]-4-phenyl-
6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]benzo[b]thieno[3,2-
e]pyrimidin-5(4H)-one 17e
4.11.2. 2-[[4-(2-Chlorophenyl)piperazin-1-yl]carbonylmethylthio]-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
ones 21b
White powder (1.05 g, 50%) m.p. 150–152 ꢀC. 1H NMR (CDCl3);
White powder (0.61 g, 35%) m.p. 245–246 ꢀC. 1H NMR (CDCl3);
d
1.35–1.50 (t, 3H, OCH2CH3), 1.70–1.88 (m, 4H, 2CH2), 2.70–2.96 (m,
d 1.52–1.73 (m, 4H, 2CH2), 1.82–1.92 (m, 2H, CH2), 2.75–2.82 (t, 2H,
8H, 2CH2 and 2NCH2), 3.08–3.21 (m, 4H, 2Ar-NCH2), 3.72 (s, 2H, CH2
attached to piperazine moiety), 4.00–4.15 (q, 2H, OCH2CH3), 6.87–
7.40 (m, 9H, Ar-H). Anal. Calcd. for C30H32N6O2S: C, 66.64; H, 5.97;
N, 15.54. Found: C, 67.00; H, 5.51; N, 15.90.
CH2), 3.02–3.30 (m, 6H, CH2, 2NCH2), 3.65–3.83 (m, 4H, 2Ar-NCH2),
4.15 (s, 2H, CH2CO), 6.76–7.33 (m, 4H, Ar-H), 12.60–12.68 (br s, 1H,
NH, D2O exchang.). Anal. Calcd. for C23H25ClN4O2S2: C, 56.49; H,
5.15; N, 11.46. Found: C, 56.80; H, 4.80; N, 11.95.
4.11.3. 2-[[4-(2-Methoxyphenyl)piperazin-1-yl]carbonylmethylthio]-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
ones 21d
4.9. Ethyl 2-(benzoylthioureido)-5,6,7,8-tetrahydro-4H-
cyclohepta[b]thiophene-3-carboxylate 18 (Scheme 2)
Brown crystal (0.78 g, 45%) m.p. 230–232 ꢀC. IR (KBr) nmax/cmꢁ1
3310 (NH), 1670, 1643 (C]O), 1150 (C–S–C). MS m/z (%); 485 (Mþ,
1.87), 378 (1.75), 294 (5.02), 234 (7.58), 220 (1.98), 191 (5.25), 133
(100). Anal. Calcd. for C24H28N4O3S2: C, 59.48; H, 5. 82; N, 11.56.
Found: C, 59.70; H, 6.10; N, 11.71.
Benzoyl chloride (3.37 g, 24 mmol) was added to a solution of
ammonium thiocyanate (2.58 g, 34 mmol) in anhydrous acetone
(15 ml) and the resulting suspension was heated under reflux for
10 min. A solution of ethyl 2-amino-5,6,7,8-tetrahydro-4H-cyclo-
hepta[b]thiophene-3-carboxylate 2 (5.74 g, 24 mmol) in anhydrous
acetone (60 ml) was added to the previous suspension and the
reaction mixture was heated under reflux for another 10 min. After
cooling, the separated solid was collected by filtration, poured into
water and filtered. The solid obtained was dried and crystallized
from acetone to afford compound 18 in 6.76 g (70%); m.p. 152–3 ꢀC.
4.11.4. 2-[[4-(2-Ethoxyphenyl)piperazin-1-yl]carbonylmethylthio]-
6,7,8,9-tetrahydro-5H-cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-
ones 21e
Brown powder (0.71 g, 40%) m.p. 220–221 ꢀC. 1H NMR (CDCl3);
d
1.28–1.34 (t, 3H, OCH2CH3), 1.54–1.76 (m, 4H, 2CH2), 1.84–1.93 (m,
1H NMR (CDCl3);
d 1.40–1.48 (t, 3H, OCH2CH3), 1.60–1.70 (m, 4H,
2H, CH2), 2.74–2.80 (t, 2H, CH2), 2.90–3.30 (m, 6H, CH2, 2NCH2),
3.69–3.88 (m, 4H, 2Ar-NCH2), 4.20 (s, 2H, CH2CO), 4.25–4.30 (q, 2H,
OCH2CH3), 6.70–7.32 (m, 4H, Ar-H), 12.40–12.45 (br s, 1H, NH, D2O
exchang.). Anal. Calcd. for C25H30N4O3S2: C, 60.21; H, 6.06; N, 11.24.
Found: C, 60.55; H, 6.40; N, 11.65.
2CH2), 1.82–1.89 (m, 2H, CH2), 2.77–2.80 (t, 2H, CH2), 3.00–3.10 (t,
2H, CH2), 4.45–4.58 (q, 2H, OCH2CH3), 7.50–7.90 (m, 5H, Ar-H),
9.00–9.10 (br s, 1H, NH, D2O exchang.), 14.50–14.60 (br s, 1H, NH,
D2O exchang.). MS m/z (%); 405 (Mþ þ 2, 0.99), 404 (Mþ þ 1, 2.04),
403 (Mþ, 2.95), 325 (0.99), 297 (1.36), 239 (27.77), 105 (100). Anal.
Calcd. for C20H22N2O3S2: C, 59.68; H, 5.51; N, 6.96. Found: C, 59.23;
H, 5.32; N, 7.40.
5. Pharmacology
5.1. 5-HT2A antagonist activity
4.10. Potassium salt of 2-mercapto-6,7,8,9-tetrahydro-5H-
cyclohepta[b]thieno[2,3-d]pyrimidin-4(3H)-one 19 (Scheme 2)
5.1.1. Materials and animals
Some of the prepared compounds were screened for 5-HT2A
antagonist activity using isolated pulmonary arterial rings of rats
[28–30]. Adult Sprague-Dawley rats of both sexes weighing 200–
250 g were used in this experiment. They were purchased from
local source. Animals were maintained under standard conditions
of temperature with regular 12 h light/12 h dark cycle and allowed
free access to standard laboratory food and water. Diethylether was
used to anesthetize the animals. A physiological salt solution (PSS)
in mM/l composed of: NaCl, 118; KCl, 4.7; CaCl2$2H2O, 2.5; KH2PO4,
1.2; MgSO4$7H2O, 1.2; NaHCO3, 25; and glucose, 10 was used to
A mixture of the thioureido derivative 18 (3.46 g, 8.6 mmol) and
potassium hydroxide (0.48 g, 8.6 mmol) in absolute ethanol (35 ml)
was heated under reflux for 3 h. The separated solid was filtered
while hot, dried and crystallized from n-butanol to give compound
19 in 1.74 g (70%); m.p. 315–317 ꢀC. 1H NMR (DMSO-d6);
d 1.46–1.60
(m, 4H, 2CH2), 1.70–1.80 (m, 2H, CH2), 2.59–2.61 (t, 2H, CH2), 3.08–
3.10 (t, 2H, CH2), 10.15–10.22 (br s, 1H, NH, D2O exchang.). Anal.
Calcd. for C11H11KN2OS2C, 45.49; H, 3.82; N, 9.64. Found: C, 45.79;
H, 3.50; N, 10.15.

M.M. El-Kerdawy et al. / European Journal of Medicinal Chemistry 45 (2010) 1805–1820
1819
maintain the arterial rings of rats in it [28–30]. Serotonin creatinine
Acknowledgments
sulfate monohydrate (5-HT), as white powder (Fluka Chemie,
Steinheim, Switzerland) was used an agonist. risperidone was used
as the standard drug. 5-HT, risperidone and the tested compounds
were freshly prepared by dissolving in water just before use in 10ꢁ4
molar concentration.
Our sincere acknowledgments to Chemical Computing Group
Inc, 1010 Sherbrooke Street West, Suite 910, Montreal H3A 2R7,
Canada, for using the package of MOE 2008.10 software. Also, our
deep thanks are due to all members of Pharmacology Department,
Faculty of Pharmacy, Mansoura University, for their aid in the in
vitro biological evaluations.
5.1.2. Methods
Rats were killed by an overdose of diethylether. The chest was
opened along the right side of the sternum after cutting the dia-
phragm. The pulmonary artery was tied gently, then separated and
rapidly placed in warm physiological salt solution (PSS). The
pulmonary artery was trimmed and cut into rings. The rings were
cleaned of connective tissue with care taken not to touch the
luminal surface. Each ring was suspended horizontally between
two stainless steel parallel hooks, one of which was fixed and the
other attached to isometric tension transducer for the measure-
ment of isometric tension in an organ bath filled with 50 ml of PSS
at fixed temperature of 37 ꢀC and continuously bubbled with 100%
pure O2. The suspended pulmonary arterial ring was allowed to
equilibrate for 30 min under a resting load of 3 g. Isometric tension
generated by the vascular smooth muscles was measured using
a displacement transducer (model 50-7905, Harvard apparatus
LTD, South Natick, MA, USA) and recorded on a two-channel
oscillograph (model 50-8622, Harvard Apparatus Ltd, USA).
A dose of 5-HT, 10ꢁ4 M, was applied and allowed to act till the
pressor response was fully developed. The pulmonary arterial ring
was washed with PSS. A concentration of 10ꢁ4 M of the test
compounds or risperidone was applied and incubated for 5 min in
the organ bath to evaluate its activity on the 5-HT2A receptors. 5-HT
was added in the organ bath in the presence of the test compounds
in order to evaluate the antagonistic activity on the 5-HT2A recep-
tors. The activities of the tested compounds are shown in Table 1.
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