4-Aminoethylpiperazinyl aryl ketones with 5-HT1A/5-HT 7 selectivity

Article abstract of DOI:10.1016/j.bmc.2011.11.005

The well-known 5-HT_1A/5-HT₇ selectivity issue was tackled by a new series of 4-aminoethylpiperazinyl aryl ketones (1a-1l) specifically designed to distinguish the two hydrophobic sites centered at the anchoring salt bridge. The 4-ami

Full text of DOI:10.1016/j.bmc.2011.11.005

Bioorganic & Medicinal Chemistry 20 (2012) 1139–1148
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
4-Aminoethylpiperazinyl aryl ketones with 5-HT1A/5-HT7 selectivity
Mi Kyoung Kim a, Hyo Seon Lee a,b, Sora Kim b, Suh Young Cho a, Bryan L. Roth c, Youhoon Chong a,
,
⇑
,
Hyunah Choo b, ,
⇑
a Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, 1 Hwayang-dong, Gwangjin-gu, Seoul 143-701, South Korea
b Neuro-Medicine Center, Korea Institute of Science and Technology, 39-1 Hawolgok-dong, Seoungbuk-gu, Seoul 136-791, South Korea
c National Institute of Mental Health Psychoactive Drug Screening Program, Division of Medicinal Chemistry and Natural Products, Department of Pharmacology, School of
Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
The well-known 5-HT1A/5-HT7 selectivity issue was tackled by a new series of 4-aminoethylpiperazinyl
aryl ketones (1a–1l) specifically designed to distinguish the two hydrophobic sites centered at the
anchoring salt bridge. The 4-aminoethylpiperazinyl aryl ketones showed a wide spectrum of activity
and selectivity for the 5-HT receptors depending on the type of the hydrophobic groups attached at
the aryl piperazinyl ketone scaffold. Docking study of the most active compounds against 5-HT7R and
5-HT1AR revealed that both receptors have two hydrophobic pockets around the anchoring salt bridge.
These two binding sites are perpendicular to each other in 5-HT7R but parallel in 5-HT1AR, and this obser-
vation is well matched with the previous report which claimed that 5-HT7R affinity arises from bent con-
formation of the bound ligand whereas an extended one is best suited for 5-HT1AR selectivity. Also, as
these pockets have different size and shape, inhibitory activity as well as selectivity of the 4-aminoeth-
ylpiperazinyl aryl ketones against 5-HT7R and 5-HT1AR seemed to be determined by combination of two
hydrophobic substituents attached at both ends of the title compounds.
Received 6 October 2011
Revised 2 November 2011
Accepted 3 November 2011
Available online 30 November 2011
Keywords:
Serotonergic receptor
5-HT1AR
5-HT7R
Selectivity
4-Aminoethylpiperazinyl aryl ketones
Ó 2011 Elsevier Ltd. All rights reserved.
1. Introduction
tivity issue is encountered for these two receptor subtypes. With
lack of crystallographic structure of 5-HT receptors, there have been
Serotonin (5-hydroxytryptamine, 5-HT), a monoamine neuro-
transmitter, exerts its effects on the central and peripheral nervous
system through interaction with various serotonergic receptor
subtypes, classified into seven families (5-HT1 to 5-HT7). All 5-HT
receptors with an exception of 5-HT3 receptor (a ligand-gated ion
channel) are G protein-coupled receptors (GPCRs), and dysfunction
of this system has been implicated in cardiovascular, digestive, and
psychiatric disorders.1 Due to their important physiological role,
many of the 5-HT receptors have become beneficial targets for
drug discovery, and several drug candidates have reached clinical
trial or market.2,3
5-HT1A receptor(5-HT1AR)isaGPCRinvolvedinseveralcognitive,
behavioral, and developmental functions, and selective partial ago-
nists of 5-HT1AR such as Gepirone, Tandospirone, and Zalospirone,
have been developed as anxiolytics and antidepressants.4–6 The
more recentlydiscovered5-HT7 receptor(5-HT7R) displaysa low de-
gree of homology (40%) with other serotonin GPCRs, but it shares a
similar pharmacological profile with 5-HT1AR.7,8 As a result, a selec-
attempts to address the problem of 5-HT7R versus 5-HT1AR selectiv-
ity through new sets of compounds with structural modifications at
different pharmacophoric elements present in serotonergic ligands
such as arylpiperazines (Fig. 1).8–10
In this study, based on the analysis of docking poses of the aryl-
piperazines to 5-HT1AR as well as 5-HT7R, we installed a carbonyl
as well as an aminoethyl group as a hydrogen bond acceptor and do-
nors around the piperazine ring to come up with a novel serotoner-
gic ligand, 4-aminoethylpiperazinyl aryl ketone (1, Fig. 1), with an
interesting 5-HT1A/5-HT7 selectivity profile. Herein we report a
proof-of-concept study of the 4-aminoethylpiperazinyl aryl ketones
which showed a wide spectrum of selectivity for the 5-HT receptors
depending on the type of the hydrophobic groups attached at the
O
R1
N
N
R1
N
N
R2
NHR2
⇑
Corresponding authors. Address: Life/Health Division, Korea Institute of Science
and Technology, PO Box 131, Cheongryang, Seoul 130-650, South Korea. Tel.: +82 2
958 5157; fax: +82 2 958 5189 (H.C.); fax: +82 2 454 8217 (Y.C.).
E-mail addresses: chongy@konkuk.ac.kr (Y. Chong), hchoo@kist.re.kr (H. Choo).
These two authors contributed equally to this work.
Arylpiperazine
4-Aminoethylpiperazinyl aryl ketone (1)
Figure 1. Structures of arylpiperazine and 4-aminoethylpiperazinyl aryl ketone (1).
0968-0896/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2011.11.005

1140
M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
aryl piperazinyl ketone scaffold. Also, docking study of each ligand
was performed to provide a rational explanation for the structure–
activity relationship as well as observed selectivity.
through the SwissModel server and docked the representative aryl-
piperazine ligands, MP349 (a ligand for 5-HT1AR)22 and UCM-5600
(a ligand for 5-HT7R),9 respectively, to the corresponding receptor
structures (Fig. 2).
Binding modes of the arylpiperazine ligands to 5-HT1AR and 5-
HT7R (right panels, Fig. 2) were in accordance with those previ-
ously reported8–10,14,16 in which ionic interactions between the
protonated amino groups of the ligands and Asp residues of the
receptors (Asp116 for 5-HT1AR and Asp162 for 5-HT7R) constitute
a main essential interaction. The hydrophobic groups attached at
both ends of the piperazine moiety were found in hydrophobic
pockets composed of amino acid residues with aromatic and/or ali-
phatic side chains [Thr196/Phe362 and Phe112/Val89/Val85/
Leu90/Tyr390 for 5-HT1AR (Fig. 2a); Val163/Tyr239/Phe344 and
Phe158/Trp148/Thr141 for 5-HT7R (Fig. 2b)]. Taken together, the
binding interactions of the arylpiperazine ligands with the recep-
tors of 5-HT1AR as well as 5-HT7R can commonly be characterized
as bidirectional hydrophobic interactions anchored at the salt
bridge, and the similar binding mode of the arylpiperazines to
these two serotonergic receptors was conceived to contribute to
the difficulty of designing novel arylpiperazine derivatives with
high 5-HT1A/5-HT7 selectivity.
2. Results and discussion
2.1. Design of the aryl piperazinyl ketones
Arylpiperazines are efficient building blocks used for design of li-
gands for the monoamine neurotransmitter receptors and they have
been thoroughly studied as ligands for 5-HT
11
R
and 5-HT7R.9
1A
Ligand-based construction of a pharmacophore model8–10,12–14 con-
firmed that the arylpiperazines include an ideal geometry of two
essential features required for a good affinity profile: a basic amine
(protonated at physiological pH)12,15 and a hydrophobic feature.
However, due to lack of crystallographic structure, specific binding
modes of arylpiperazine ligands to 5-HT1AR and 5-HT7R have been
ambiguous. In order to overcome this problem, there have been
many efforts to generate comparative model structures of the sero-
tonergic receptors.16–21 The model structures of 5-HT1AR and
5-HT7R have also been prepared and binding modes of ligands such
as arylpiperazines were elucidated.8–10,14,16 In an attempt to get a
better understanding of the selective binding interactions involved
in 5-HT1A/5-HT7 receptors and arylpiperazine ligands, we have con-
structed homology models of 5-HT1Aas well as 5-HT7 receptors
However, it was worth to note that, at the vicinity of a charged
amine atom of the bound arylpiperazine ligands, different amino
acid residues that can serve as hydrogen bond donors/acceptors
Figure 2. Binding modes of (a) MP349 to 5-HT1A receptor and (b) UCM-5600 to 5-HT7 receptor. Right panels are two dimensional illustrations of the binding mode analyses.
Hydrogen bonds are indicated as dotted lines.

M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
1141
are located in 5-HT1AR (Asn386 and Tyr390, Fig. 2a) and 5-HT7R
(Arg367 and Thr141, Fig. 2b), respectively. Based on this observa-
tion, we reasoned that introduction of a hydrogen bond donor or
acceptor around the ionizable piperazine moiety would help differ-
entiate the bidirectional hydrophobic interactions anchored at the
salt bridge and thereby increase 5-HT1A/5-HT7 selectivity. The title
compound of this study, 4-aminoethylpiperazinyl aryl ketone (1,
Fig. 3), was thus designed via installation of the fourth pharmaco-
phoric element, carbonyl and aminoethyl group as a hydrogen
bond acceptor and donor, at both ends of the piperazine core
structure.
tert-butyl-substituted ketones (1e–1h) showed no inhibition at
all while benzimidazole-substituted ketones (1i–1l) were moder-
ately potent. On the other hand, the aromatic R1 substituent played
a critical role in determining activity of the isoxazole-substituted
ketones (1a–1d), and only the 4-chlorophenyl ketone (1a) showed
significant activity with 93.6%-inhibition.
2.3.3. 5-HT1A/5-HT7 selectivity
Regarding receptor selectivity of the title compounds, three
important features were identified. First, complete inactivity of
the tert-butyl-substituted derivatives (1e–1h) against 5-HT7R
regardless of the aromatic substituents in R1 indicates that the
tert-butyl group associated in the aryl piperazinyl ketone scaffold
cannot be accommodated by the 5-HT7R. Even though the tert-bu-
tyl-substituted derivatives (1e–1h) were only moderately potent
against 5-HT1AR, their complete inactivity toward 5-HT7R provides
a possibility to discover 5-HT1AR-selective inhibitors through
extensive structure–activity relationship study of the R1 substitu-
ent. Second, the receptor selectivity is lost upon replacement of
the tert-butyl group with a benzimidazole to result in better inhib-
itory activity of the corresponding aryl piperazinyl derivatives (1i–
1l) against 5-HT1AR and 5-HT7R. Once again, the specific role of the
tert-butyl group in discriminating the 5-HT7R is of great interest.
Third, unlike other series of the aryl piperazinyl ketones, the isox-
azole-substituted derivatives (1a–1d) showed R1-dependent inhib-
itory activity against 5-HT7R, and compound 1a was the only
example which showed significant selectivity for 5-HT7R. In other
words, this compound must have recognized the ligand binding
site of 5-HT7R more specifically than others, which indicates that
analysis of its binding mode would provide valuable information
for designing 5-HT7R-selective inhibitors.
2.2. Chemistry
A straightforward synthesis of the title compounds, aryl piper-
azinyl ketones (1a–1l), is summarized in Scheme 1. Commercially
available 1-Boc-piperazine (2) was reacted with four acyl chlorides
(R1COCl) to give the corresponding aryl piperazinyl ketones 3a–3d
in 66–98% yields. After deprotection of the Boc protecting group
under acidic conditions, ethylamino functionality was introduced
to the resulting ketones4 through substitution with N-(2-bromo-
ethyl)-phthalimide followed by N-dephthalation with hydrazine
hydrate. Reductive amination of the free amines 6, thus obtained,
with three aldehydes (R2CHO), followed by deprotection as needed,
provided the title compounds 1a–1l in 8–74% yields.
2.3. Biological activity
Newly synthesized compounds 1a–1l were assessed for in vitro
binding affinity at human serotonergic 5-HT1A and 5-HT7 receptors
by radioligand binding assays, using [3H]LSD in transfected
CHO-K1 cells and [3H]-8-OH-DPAT in transfected HEK-293 cells,
respectively.23,24 All compounds were assayed as hydrochloride
salts. The competitive inhibition assays were performed at a fixed
2.4. Binding mode analysis
dose of 10
l
M, and the %-inhibition data were shown in Table 1.
Structure–activity relationship as well as 5-HT1A/5-HT7 selectiv-
ity of the aryl piperazinyl ketones described above need to be inter-
preted by specific binding modes in order to provide information
for designing receptor-selective serotonergic ligands. For this pur-
pose, docking study of the aryl piperazinyl ketones (1a–1l) to ligand
binding sites of 5-HT1AR and 5-HT7R was performed with Glide 4.0
implemented in Maestro 7.5 (Schrödinger, Inc.). During docking, an
H-bond constraint between the charged amino group of each ligand
and Asp116 (5-HT1AR) or Asp162 (5-HT7R) was imposed in order to
avoid spurious results. Interestingly, under this constraint, the two
most active compounds against 5-HT1AR (1k and 1l, Table 1) and
the most active one against 5-HT7R (1a, Table 1) were successfully
docked to the target receptor but none of others provided docking
poses. These three derivatives maintained the same binding
modes even without the H-bond constraint, but others showed
nonspecific binding modes at the periphery of the receptor under
2.3.1. Binding affinity to 5-HT1AR
The 4-aminoethylpiperazinyl aryl ketones showed broad inhib-
itory spectrum of ligand binding to one serotonin receptor, 5-
HT1AR (Table 1). The compounds with 4-chloro and 3,4-dichloro
moieties (1a, 1b, 1e, 1f, 1i and 1j) showed only marginal activities
against 5-HT1AR. The benzimidazole-substituted derivatives 1k
and 1l were most active. It is noteworthy that these two com-
pounds have R1-aromatic substituents such as chlorine atom or tri-
fluoromethyl group at both 3 and 5 positions.
2.3.2. Binding affinity to 5-HT7R
Inhibitory activity of 4-aminoethylpiperazinyl aryl ketones
against 5-HT7R was more heavily dependent on R2 group rather
than R1 group (Table 1). Thus, regardless of R1 substituent, the
R1
R2
N
O
O
Cl
Cl
R1
N
Cl
N
R2
N
H
F3C
Cl
N
1
HN
CF3
Cl
Figure 3. Structures of the 4-aminoethylpiperazinyl aryl ketones (1) prepared in this study.

1142
M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
O
O
a)
66%~98%
b)
HN
R1
N
R1
N
NBoc
83%~99%
NBoc
NH
4a ~ 4d
2
3a ~ 3d
R1
c) 31%~79%
O
3a
4-Cl-Ph
3b 3,4-di-Cl-Ph
3c
3d
3,5-di-Cl-Ph
3,5-bis-CF3-Ph
R1
N
O
N
N
R1
4-Cl-Ph
R2
5a ~ 5d
1a
O
1b 3,4-di-Cl-Ph
O
N
1c
1d
3,5-di-Cl-Ph
3,5-bis-CF3-Ph
d) 79%~98%
O
1e
4-Cl-Ph
O
1f 3,4-di-Cl-Ph
e)
1g
1h
3,5-di-Cl-Ph
R1
N
R1
N
3,5-bis-CF3-Ph
N
R2
N
8%~74%
N
H
NH2
1i
4-Cl-Ph
1a ~ 1l
6a ~ 6d
N
1j 3,4-di-Cl-Ph
1k
1l
HN
3,5-di-Cl-Ph
3,5-bis-CF3-Ph
Scheme 1. Synthesis of the 4-aminoethylpiperazinyl aryl ketones (1a–1l). Reagents and conditions: (a) R1COCl, DIPEA, CH2Cl2, rt; (b) 30% TFA in CH2Cl2, rt; (c) N-(2-
bromoethyl)-phthalimide, K2CO3, CH3CN, reflux; (d) NH2NH2ꢀH2O, EtOH, reflux; (e) R2CHO, NaBH(OAc)3, MeOH, rt, then, if needed, 30% TFA in CH2Cl2, rt.
Table 1
%-Inhibition data of the 4-aminoethylpiperazinyl aryl ketones (1a–1l) against 5-HT1A and 5-HT7 receptors
O
R1
N
N
R2
N
H
1a ~ 1l
R1
R2
R1
4-Cl-Ph
R2
R1
R2
1a
1i
4-Cl-Ph
4-Cl-Ph
1e
N
1b 3,4-di-Cl-Ph
1j 3,4-di-Cl-Ph
1f 3,4-di-Cl-Ph
O
N
1c
1d
1k
1l
HN
3,5-di-Cl-Ph
3,5-di-Cl-Ph
1g
1h
3,5-di-Cl-Ph
3,5-bis-CF3-Ph
3,5-bis-CF3-Ph
3,5-bis-CF3-Ph
Compds
%-Inhibition at 10
l
M
Compds
%-Inhibition at 10
l
M
Compds
%-Inhibition at 10 lM
5-HT1A
5-HT7
5-HT1A
5-HT7
5-HT1A
5-HT7
1a
1b
1c
1d
56.5
76.4
43.2
13.0
93.6
16.1
ꢁ0.2
ꢁ0.3
1e
1f
1g
1h
36.7
65.2
33.7
ꢁ4.4
ꢁ0.6
2.0
1i
1j
1k
1l
47.5
49.5
90.1
87.7
36.8
56.2
62.7
45.4
ꢁ0.6
ꢁ0.3
the non-constrained docking conditions. In particular, due to large
hydrophobic surface and hydrogen bonding capacity, the benzimid-
azole-substituted derivatives (1i–1l) were more susceptible for
nonspecific binding, which provides a reasonable explanation for
their moderate inhibitory activity and low selectivity against the
two serotonergic receptors. Thus, the docking results were in accor-
dance with the biological activity data and analysis of the detailed
docking poses was anticipated to delineate subtle differences in the
ligand binding sites of 5-HT1AR and 5-HT7R.
receptors, 5-HT1AR and 5-HT7R. The binding mode of aryl
piperazinyl ketone 1a to 5-HT7R (Fig. 4a) shows the four key phar-
macophoric elements are all in action. Thus, in addition to the criti-
cal ionic interaction of N2-nitrogen of the piperazine with Asp162,
the keto carbonyl group forms a hydrogen bond with Arg367 (5-
HT7R) and the two substituents, 4-chlorophenyl (R1) and 3-isopro-
pylisoxazole (R2), were located in the hydrophobic pockets, R-I and
R-II composed of Leu232/Cys231/Cys155/Trp148 (for R1) and
Ile159/Phe343/Phe344/Leu370/Val163/Thr244/Ser243 (for R2),
respectively (Fig. 4).The binding site for 4-chlorophenyl R1 substitu-
ent is flat and narrow disallowing any substituent on the aromatic
As expected, hydrophobic interaction of the two substituents (R1
and R2) played the key role in distinguishing the two serotonergic

M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
1143
Figure 4. Binding mode of the compound 1a to 5-HT7R. (a) Schematic 2D plot showing intermolecular interactions. (b) Surface representation to show hydrophobic pockets
(dotted circles) perpendicular to each other. Two hydrophobic binding sites are indicated as dotted circles and the R1- and R2-binding site are noted by R-I and R-II,
respectively. The 2D schematic plots were created with the program LIGPLOT.25
ring other than at4-position (Fig. 4a), which is in matchwiththe dra-
matic decrease in the binding affinity of 1b–1d in comparison with
1a (Table 1). The R2-binding site of 5-HT7R is more globular than
the R1-binding site but not big enough to accommodate bulky or
large substituent such as tert-butyl or benzimidazole (Fig. 4a). Thus,
among the three R2-substituents introduced in this study, relatively
small 3-isopropylisoxazole fits in this pocket and the lack of inhibi-
tory activity of the tert-butyl substituted derivatives (1e–1h, Table
1) as well as peripheral binding of the benzimidazole-substituted
derivatives (1i–1l) to 5-HT7R can be associated with this feature of
the R2-binding site. Surface representation of the binding mode
(Fig. 4b) shows that these two hydrophobic binding sites (R-I and
R-II, Fig. 4b) are perpendicular to each other and adequate for bind-
ing of a bent conformation of the ligand 1a. This observation is rem-
iniscent of the report by Lepailleur et al.8 which claimed that 5-HT7R
affinity arises from bent conformation of the phenylpyrrole deriva-
tives whereas an extended one is best suited for 5-HT1AR selectivity.
The binding modes of 1k and1l to 5-HT1AR were almost identical
to each other, and the schematic 2D plot showing intermolecular
interactions of the representative ligand1k with 5-HT1AR was sim-
ilar to that of 1a (Fig. 5a). The piperazinyl N2-nitrogen of 1kwas in
ionic interaction with Asp116 and the two substituents (R1 and R2)
were located in two hydrophobic pockets, R-I0 and R-II0, composed
of Ala365/Lys191 (for R1) and Asp82/Val85/Cys119/Cys120/
Asn392/Ser123 (for R2) in Fig. 5. However, surface representation
of the binding mode of 1k (Fig. 5b) is significantly different from
that of 1a (Fig. 4b) in that the two hydrophobic binding sites (R-I0
and R-II0, Fig. 5b) are parallel rather than perpendicular. Thus, an
optimal ligand conformation for binding to 5-HT1AR is expected
to be an extended one, which, once again, is consistent with the re-
port by Lepailleur et al.8 Also, the docking pose of the ligand 1k
bound in 5-HT1AR has a flipped conformation in comparison to that
of the ligand 1a in 5-HT7R, resulting in its 3,5-dichlorophenyl ring
(R1) located at R-I0 corresponding to the R2-binding site (R-II) of
5-HT7R (Figs. 4b and 5b). Like R-II in 5-HT7R, the hydrophobic pock-
et R-I0 has a globular shape and this structural feature allows selec-
tive binding of the aromatic R1 with substituents at 3 and/or 5
position rather than 4 position. Potent inhibitory activity of 1k
and 1l against 5-HT1AR among the benzimidazole-substituted
derivatives (Table 1) might be related with the substituent-specific
binding event at R-I0.
On the other hand, while the R1-binding site in 5-HT7R (R-I,
Fig. 4b) is formed by a shallow opening between Leu232 and
Trp148, the corresponding site in 5-HT1AR composed of Cys187
and Trp102 is closed (Fig. 5b). The lack of second hydrophobic pock-
et in 5-HT1AR is compensated by a new binding site for the R2-sub-
stituent (R-II0) near Tyr390 (Fig. 5b). This site has a structure of
narrow cleft flanked by hydrophobic side chains (Fig. 5a).Therefore,
it is unlikely for this site to accommodate bulky R1 or R2 substituent
with a branched chain and this provides a reasonable explanation
for lack of specific docking poses of the 3-isopropyloxazole- (1a–
1d) as well as tert-butyl-substituted (1e–1h) aryl piperazinyl
ketones at 5-HT1AR and thereby their moderate to low inhibitory
activity against this receptor. And the benzimidazole-substituted
aryl piperazinyl ketones (1k and 1l) with relatively long carbon
chain show potent activity with depending on the R1 substitutents.
3. Conclusion
The 4-aminoethylpiperazinyl aryl ketones showed a wide spec-
trum of activity and selectivity for the 5-HT receptors depending on

1144
M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
Figure 5. Binding mode of 1k to 5-HT1AR. (a) Schematic 2D plot showing intermolecular interactions. (b) Surface representation to show hydrophobic pockets (dotted circles)
perpendicular to each other. Two hydrophobic binding sites are indicated as dotted circles and the R1- and R2-binding site are noted by R-I0 and R-II0, respectively.
the type of the hydrophobic groups attached (R1 and R2) at the aryl
piperazinyl ketone scaffold. The title compounds with the benz-
imidazole-substituted derivatives (1k and 1l) showed the most
activity against 5-HT1AR receptor and, the compound 1a with an
isoxazole substituent showed the best binding affinity against 5-
HT7 receptor.
In order to interpret the seemingly complicated activity as
well as selectivity profile, a docking study was performed. Analy-
sis of binding modes of 1a and 1k to 5-HT7R and 5-HT1AR, respec-
tively, revealed that both receptors have two hydrophobic
pockets around the anchoring salt bridge and the hydrophobic
substituents (R1 and R2) of 1a and 1k bind to these pockets. It
is noteworthy that these two binding sites are perpendicular to
each other in 5-HT7R but parallel in 5-HT1AR, and this observation
is well matched with the previous report which claimed that 5-
HT7R affinity arises from bent conformation of the bound ligand
whereas an extended one is best suited for 5-HT1AR selectivity.
Also, as these pockets have different size and shape, appropriate
combination of R1 and R2 is crucial for selective binding to each
receptor.
4. Experimental
4.1. Chemistry
4.1.1. General remarks
Nuclear magnetic resonance spectra were recorded at 400 or
300 MHz for 1H NMR and at 100 or 75 MHz for 13C NMR with tet-
ramethylsilane as internal standard. Chemical shift are reported as
s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), or br s
(broad singlet). Coupling constants are reported in hertz (Hz). The
chemical shifts are reported as parts per million (d) relative to the
solvent peak.
4.1.2. Synthesis of tert-butyl 4-(aryloyl)piperazine-1-carboxylate (3a–3
d)
General procedure: To
a solution of 1-Boc-piperazine 2
(5.37 mmol) in CH2Cl2 was added DIPEA (10.74 mmol) and acyl
chloride (8.06 mmol). The reaction mixture was stirred for 24 h.
After reaction, the mixture was diluted with satd NaHCO3, ex-
tracted with CH2Cl2, dried MgSO4. The crude compound was puri-
fied by column chromatography (SiO2, Hexane/EtOAc = 2:1) to
afford the desired compound 3a–3d as yellow oil in 66–99% yield.
Through synthesis, biological evaluation, and docking study of
4-aminoethylpiperazinyl aryl ketones against 5-HT7R and 5-HT1AR,
we have shown that their inhibitory activity as well as selectivity
were maneuvered by interplay of two hydrophobic substituents at-
tached at both ends of the title compound. Based on this informa-
tion, extensive structure–activity relationship study is in progress
and will be published elsewhere.
4.1.2.1. Synthesis of tert-butyl 4-(4-chlorobenzoyl)piperazine-
1-carboxylate (3a).
Reaction of the compound 2with 4-chlo-
robenzyoyl chloride according to the procedure described above
afforded 3a in 66% yield as yellow oil:1H NMR (300 MHz, CDCl3)

M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
1145
d 7.42 (dd, J = 6.5, 2.1 Hz, 2H), 7.36 (dd, J = 6.5, 2.2 Hz, 2H), 3.46–
3.69 (m, 8H), 1.48 (s, 9H); 13C NMR (75 MHz, CDCl3) d 169.51,
154.51, 136.05, 133.80, 128.87, 128.61, 80.41, 43.67, 28.35.
4.1.4. Synthesis of 2-(2-(4-arylpiperazin-1-yl)ethyl)isoindoline-
1,3-dione (5a–5d)
General procedure: To a solution of piperazine 4a–4d (2.9 mmol)
obtained above in acetonitile was added K2CO3 (5.8 mmol) and N-
(2-bromoethyl)-phthalimide (4.4 mmol). The reaction mixture was
refluxed for 24 h. After reaction, the mixture was diluted with H2O
and then extracted with EtOAc three times. The combined organic
layer was dried over MgSO4. The crude product was purified by col-
umn chromatography (SiO2, Hexane/EtOAc/CH2Cl2 = 1:1:1) to af-
ford the desired compound 5a–5d as white powders in 31–79%
yield.
4.1.2.2. Synthesis of tert-butyl 4-(3,4-dichlorobenzoyl)pipera-
zine-1-carboxylate (3b).
Reaction of the compound 2 with
3,4-dichlorobenzoyl chloride according to the procedure described
above afforded 3b in 82% yield as yellow oil:1H NMR (300 MHz,
CDCl3) d 7.51–7.53 (m, 2H), 7.24–7.28 (m, 1H), 3.36–3.82 (m,
8H), 1.49 (s, 9H); 13C NMR (75 MHz, CDCl3) d 168.13, 162.91,
135.21, 134.42, 133.18, 130.73, 129.32, 126.37, 80.52, 28.35.
4.1.2.3. Synthesis of tert-butyl 4-(3,5-dichlorobenzoyl)pipera-
4.1.4.1. Synthesis of 2-(2-(4-(4-chlorobenzoyl)piperazin-1-yl)
zine-1-carboxylate (3c).
Reaction of the compound 2 with
ethyl)isoindoline-1,3-dione (5a).
Reaction of 4a with N-(2-
3,5-dichlorobenzyoyl chloride according to the procedure de-
scribed above afforded 3c in 99% yield as yellow oil:1H NMR
(300 MHz, CDCl3) d 7.44 (s, 1H), 7.29 (s, 2H), 3.45–3.75 (m, 8H),
1.48 (s, 9H); 13C NMR (75 MHz, CDCl3) d 167.55, 154.46, 138.23,
135.55, 130.02, 125.51, 80.56, 28.35.
bromoethyl)-phthalimide in the presence of K2CO3 in acetonitile
according to the procedure described above afforded 5a in 31%
yield as white powder:1H NMR (300 MHz, CDCl3) d 7.84–7.87 (m,
2H), 7.72–7.76 (m, 2H), 7.31–7.40 (m, 4H), 3.81–3.85 (m, 2H),
3.68–3.71 (m, 2H), 3.33–3.36 (m, 2H), 2.50–2.69 (m, 6H); 13C
NMR (75 MHz, CDCl3) d 169.16, 168.35, 135.74, 134.21, 133.95,
132.16, 128.73, 128.61, 123.25, 55.57, 35.09.
4.1.2.4. Synthesis of tert-butyl 4-(3,5-bis-trifluoromethylben-
zoyl)piperazine-1-carboxylate (3d).
Reaction of the com-
4.1.4.2. Synthesis of 2-(2-(4-(3,4-dichlorobenzoyl)piperazin-
pound 2with 3,5-bis-trifluoromethylbenzoyl chloride according
to the procedure described above afforded 3d in 99% yield as yel-
low oil: 1H NMR (300 MHz, CDCl3) d 7.94 (s, 1H), 7.87 (s, 2H),
3.30–3.80 (m, 4H), 2.80–3.00 (m, 4H), 2.01 (s, 1H).
1-yl)ethyl)isoindoline-1,3-dione (5b).
Reaction of 4b
with N-(2-bromoethyl)-phthalimide in the presence of K2CO3
in acetonitile according to the procedure described above affor-
ded 5b in 69% yield as white powder:1H NMR (300 MHz, DMSO)
d: 7.82–7.90 (m, 4H), 7.65–7.71 (m, 2H), 7.37 (dd, J = 8.2, 1.9 Hz,
1H), 3.71 (t, J = 6.4 Hz, 2H), 3.45–3.55 (m, 2H), 3.13–3.24 (m,
2H), 2.57 (t, J = 6.4 Hz, 2H), 2.41–2.51 (m, 4H).
4.1.3. Synthesis of aryl(piperazin-1-yl)methanone (4a–4d)
General procedure: To a solution of 3a–3d (3.5 mmol) in CH2Cl2
was added 30% TFA, and the resulting mixture was stirred at rt for
5 h. After reaction, the mixture was made basic with saturated NaH-
CO3 solution. This mixture was extracted with CH2Cl2, dried over
MgSO4, and concentrated under reduced pressure to give the desired
compounds 4a–4d as white solids. These compounds were used for
the next reaction without further purification (83–99% yield).
4.1.4.3. Synthesis of 2-(2-(4-(3,5-dichlorobenzoyl)piperazin-1-yl)
ethyl)isoindoline-1,3-dione (5c).
Reaction of 4c with N-(2-bromo-
ethyl)-phthalimide in the presence of K2CO3 in acetonitile according to
the procedure described above afforded 5c in 65% yield as white powder:
1H NMR (300 MHz, DMSO) d 7.87 (dd, J = 5.4 Hz, 3.0 Hz, 2H), 7.74 (dd,
J = 5.4 Hz, 3.0 Hz, 2H), 7.41 (d, J = 2.1 Hz, 1H), 7.25–7.28 (m, 2H), 3.84 (t,
J = 6.3 Hz, 2H), 3.69 (s, 2H), 3.34 (s, 2H), 2.51–2.71 (m, 6H); 13C NMR
(75 MHz, CDCl3) d: 168.35, 138.65, 135.37, 133.97, 132.15, 129.73,
125.50, 123.26, 55.53, 35.07.
4.1.3.1. Synthesis of (4-chlorophenyl)(piperazin-1-yl)metha-
none (4a).
Reaction of 3a with 30% TFA in CH2Cl2according
to the procedure described above afforded 4a in 83% yield as white
solid: 1H NMR (300 MHz, CDCl3) d 7.33–7.41 (m, 4H), 3.40–3.71 (m,
4H), 2.85–2.90 (m, 4H), 1.79 (s, 1H); 13C NMR (75 MHz, CDCl3) d
169.31, 135.70, 134.26, 128.75, 128.57, 46.20.
4.1.4.4. Synthesis of 2-(2-(4-(3,5-bis-trifluoromethylbenzoyl)
piperazin-1-yl)ethyl)isoindoline-1,3-dione (5d).
Reaction
of 4d with N-(2-bromoethyl)-phthalimide in the presence of
K2CO3 in acetonitile according to the procedure described above
afforded 5d in79% yield as white powder: 1H NMR (300 MHz,
DMSO) d: 7.94 (s, 1H), 7.85–7.88 (m, 4H), 7.72–7.77 (m, 2H), 3.84
(t, J = 6.2 Hz, 2H), 3.71–3.76 (m, 2H), 3.30–3.35 (m, 2H), 2.70 (t,
J = 6.2 Hz, 2H), 2.50–2.65 (m, 4H).
4.1.3.2. Synthesis of (3,4-dichlorophenyl)(piperazin-1-yl)meth-
anone (4b).
Reaction of 3b with 30% TFA in CH2Cl2according
to the procedure described above afforded 4b in 97% yield as white
solid: 1H NMR (300 MHz, CDCl3) d 7.48–7.52 (m, 2H), 7.23–7.28 (m,
1H), 3.40–3.73 (m, 4H), 2.85–2.90 (m, 4H), 2.02 (s, 1H); 13C NMR
(75 MHz, CDCl3) d 167.90, 135.63, 134.04, 133.01, 130.61, 129.26,
126.36, 46.12.
4.1.5. Synthesis of (4-(2-aminoethyl)piperazin-1-yl)arylmetha none
(6a–6d)
General procedure: To a solution of the phthalimide obtained
above (0.91 mmol) in EtOH was added hydrazine monohydrate
(2.73 mmol). The mixture was refluxed at 60 °C for 2 h and then
cooled at rt. After reaction, EtOH was removed, and then filtered.
The organic solvent was evaporated to give the desired compound
6a–6d as yellow oil in 79–91% yield, which was used for the next
step without further purification.
4.1.3.3. Synthesis of (3,5-dichlorophenyl)(piperazin-1-yl)meth-
anone (4c).
Reaction of 3c with 30% TFA in CH2Cl2according
to the procedure described above afforded 4c in 97% yield as white
solid: 1H NMR (300 MHz, CDCl3) d 7.43 (s, 1H), 7.28 (s, 2H), 3.35–
3.95 (m, 6H), 2.90–3.00 (m, 4H); 13C NMR (75 MHz, CDCl3) d:
167.37, 138.33, 135.50, 129.90, 125.48, 45.50.
4.1.5.1. Synthesis of (4-(2-aminoethyl)piperazin-1-yl)(4-chloro-
4.1.3.4. Synthesis of (3,5-bis-trifluoromethylphenyl)(piperazin-
phenyl)methanone (6a).
Reaction of 5a with hydrazine
1-yl)methanone (4d).
Reaction of 3d with 30% TFA in CH2Cl2
monohydrate in EtOH according to the procedure described above
afforded 6a in 92% yield as yellow oil: 1H NMR (300 MHz, CDCl3) d
7.33–7.41 (m, 4H), 3.76–3.78 (m, 2H), 3.41–3.43 (m, 2H), 2.79–2.83
according to the procedure described above afforded 4d in 99%
yield as white solid: 1H NMR (300 MHz, CDCl3) d 7.94 (s, 1H),
7.87 (s, 2H), 3.30–3.80 (m, 4H), 2.80–3.00 (m, 4H), 2.01 (s, 1H).

1146
M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
(m, 2H), 2.44–2.51 (m, 6H), 1.67 (s, 2H); 13C NMR (75 MHz, CDCl3)
(1c).
Reaction of 6c with 3-isopropylisoxazole-5-carbalde-
d 169.19, 135.76, 134.15, 128.75, 128.61, 60.91, 53.48, 38.64.
hyde according to the procedure described above afforded 1c in
33% yield as yellow oil after column chromatography (SiO2,
CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d: 7.41 (s, 1H),
7.27 (s, 2H), 6.03 (s, 1H), 3.90 (s, 2H), 3.73–3.77 (m, 2H), 3.38–
3.42 (m, 2H), 3.02–3.07 (m, 1H), 2.73–2.77 (m, 2H), 2.38–2.57
(m, 6H), 1.28 (d, J = 7.0 Hz, 6H); 13C NMR (75 MHz, CDCl3) d:
170.92, 169.35, 167.18, 138.57, 135.40, 129.77, 125.50, 99.87,
57.49, 45.45, 45.00, 26.48, 21.76.
4.1.5.2. Synthesis of (4-(2-aminoethyl)piperazin-1-yl)(3,4-dichlo
rophenyl)methanone (6b).
Reaction of 5b with hydrazine
monohydrate in EtOH according to the procedure described above
afforded 6b in 91% yield as yellow oil:1H NMR (300 MHz, CDCl3) d
7.47–7.50 (m, 2H), 7.23 (d, J = 8.2, 1.9 Hz, 1H), 3.35–3.75 (m, 4H),
2.81 (t, J = 5.8 Hz, 2H), 2.40–2.50 (m, 6H), 2.24 (s, 2H); 13C NMR
(75 MHz, CDCl3) d 167.78, 135.53, 134.08, 132.98, 130.60, 129.26,
126.38, 60.63, 53.38, 42.36, 38.48.
4.1.6.4. Synthesis of (3,5-bis(trifluoromethyl)phenyl)(4-(2-
((3-isopropylisoxazol-5-yl)methylamino)ethyl)piperazin-1-yl)
4.1.5.3. Synthesis of (4-(2-aminoethyl)piperazin-1-yl)(3,5-dichlo
methanone (1d).
Reaction of 6d with 3-isopropylisoxazole-
rophenyl)methanone (6c).
Reaction of 5c with hydrazine
5-carbaldehyde according to the procedure described above affor-
ded 1d in10% yield as yellow oil after column chromatography
(SiO2, CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d 7.94 (s,
1H), 7.87 (s, 2H), 6.03 (s, 1H), 3.91 (s, 2H), 3.80–3.82 (m, 2H),
3.40–3.42 (m, 2H), 3.01–3.07 (m, 1H), 2.74–2.77 (m, 2H), 2.40–
2.58 (m, 6H), 1.28 (d, J = 7.0 Hz, 6H); 13C NMR (75 MHz, CDCl3) d
170.80, 169.35, 166.98, 137.82, 132.15 (q, J = 33.5 Hz), 127.45,
123.55, 122.86 (q, J = 271.4 Hz), 99.88, 57.45, 52.72, 45.43, 44.97,
26.47, 21.93.
monohydrate in EtOH according to the procedure described above
afforded 6c in 79% yield as yellow oil: 1H NMR (300 MHz, CDCl3) d
7.42 (s, 1H), 7.28 (s, 2H), 3.35–3.80 (m, 4H), 2.82 (t, J = 5.9 Hz, 2H),
2.45–2.55 (m, 6H), 1.86 (s, 2H); 13C NMR (75 MHz, CDCl3) d:
167.20, 138.61, 135.39, 129.76, 125.50, 60.65, 38.56.
4.1.5.4. Synthesis of (4-(2-aminoethyl)piperazin-1-yl)(3,5-bis
(trifluoromethyl)phenyl)methanone (6d).
Reaction of 5d
with hydrazine monohydrate in EtOH according to the procedure
described above afforded 6d in 81% yield as yellow oil:1H NMR
(300 MHz, CDCl3) d 7.94 (s, 1H), 7.87 (s, 2H), 3.80–3.85 (m, 2H),
3.40–3.50 (m, 2H), 2.81 (t, J = 5.9 Hz, 2H), 2.47–2.62 (m, 6H), 1.65
(s, 2H).
4.1.6.5. Synthesis of (4-chlorophenyl)(4-(2-(3,3-dimethylbutyla-
mino)ethyl)piperazin-1-yl)methanone (1e).
Reaction of 6a
with 3,3-dimethylbutanalaccording to the procedure described
above afforded 1e in 8% yield as yellow oil after column chroma-
tography (SiO2, CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3)
d: 7.33–7.41 (m, 4H), 3.34–3.77 (m, 4H), 2.50–2.61 (m, 10H),
1.35–1.40 (m, 2H), 0.90 (s, 9H); 13C NMR (75 MHz, CDCl3) d:
169.21, 135.79, 134.09, 128.81, 128.76, 56.68, 51.10, 50.07, 39.73,
29.79, 29.49.
4.1.6. Synthesis of aryl(4-alkylaminoethyl)piperazin-1-yl)meth anone
(1a–1l)
General procedure: To a solution of an aldehyde (0.76 mmol) in
MeOH was added the amine 6a–6d prepared above (0.84 mmol).
The mixture was stirred for 2 h, and then NaBH(OAc)3 (2.28 mmol)
was added. After 6 h of stirring, the solvent was removed. The reac-
tion mixture was diluted with H2O, added saturated NaHCO3 solu-
tion, extracted with CH2Cl2, dried MgSO4. The crude compound
was purified by column chromatography to afford the desired com-
pound 1a–1h as an oil in 8–74% yield.
4.1.6.6. Synthesis of (3,4-dichlorophenyl)(4-(2-(3,3-dimethylbu-
tylamino)ethyl)piperazin-1-yl)methanone (1f).
Reaction of
6b with 3,3-dimethylbutanalaccording to the procedure described
above afforded 1f in 61% yield as yellow oil after column chroma-
tography (SiO2, CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d
7.48–7.51 (m, 2H), 7.24 (d, J = 8.1 Hz, 1H), 3.70–3.75 (m, 2H), 3.40–
3.45 (m, 2H), 2.73 (t, J = 6.0 Hz, 2H), 2.59–2.65 (m, 2H), 2.44–2.56
(m, 6H), 1.39–1.44 (m, 2H), 0.91 (s, 9H); 13C NMR (75 MHz, CDCl3)
d 167.75, 135.59, 134.07, 132.98, 130.59, 129.28, 126.39, 57.89,
53.47, 46.51, 46.18, 44.11, 29.82, 29.60.
4.1.6.1. Synthesis of
xazol-5-yl)methylamino)ethyl)piperazin-1-yl)methanone
(1a) . Reaction of 6a with 3-isopropylisoxazole-5-carbalde-
(4-chlorophenyl)(4-(2-((3-isopropyliso
hyde according to the procedure described above afforded 1a in
74% yield as yellow oil after column chromatography (SiO2,
CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d 7.34–7.41 (m,
4H), 6.03 (s, 1H), 3.91 (s, 2H), 3.76–3.78 (m, 2H), 3.42–3.43 (m,
2H), 3.03–3.07 (m, 1H), 2.73–2.77 (m, 2H), 2.40–2.56 (m, 4H),
1.90 (s, 1H), 1.28 (d, J = 6.9 Hz, 6H); 13C NMR (75 MHz, CDCl3) d
170.94, 169.35, 169.19, 135.79, 134.11, 128.77, 128.62, 99.86,
57.44, 45.49, 45.02, 26.48, 21.78.
4.1.6.7. Synthesis of (3,5-dichlorophenyl)(4-(2-(3,3-dimethylbu-
tylamino)ethyl)piperazin-1-yl)methanone (1g).
Reaction of
6c with 3,3-dimethylbutanalaccording to the procedure described
above afforded 1g in 55% yield as yellow oil after column chroma-
tography (SiO2, CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d
7.42 (s, 1H), 7.28 (s, 2H), 3.75–3.80 (m, 2H), 3.38–3.42 (m, 2H), 2.73
(t, J = 5.9 Hz, 2H), 2.42–2.65 (m, 8H), 1.39–1.44 (m, 2H), 0.92 (s,
9H); 13C NMR (75 MHz, CDCl3) d 167.18, 138.63, 135.39, 129.74,
125.50, 57.91, 46.53, 46.20, 44.15, 29.82, 29.60.
4.1.6.2. Synthesis of (3,4-dichlorophenyl)(4-(2-((3-isopro-
pyliso xazol-5-yl)methylamino)ethyl)piperazin-1-yl)metha-
none (1b) .
Reaction of 6b with 3-isopropylisoxazole-5-
carbaldehyde according to the procedure described above afforded
1b in 55% yield as yellow oil after column chromatography (SiO2,
CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d 7.47–7.50 (m,
2H), 7.23 (dd, J = 8.2, 1.9 Hz, 1H), 6.02 (s, 1H), 3.88 (s, 2H), 3.70–
3.75 (m, 2H), 3.40–3.45 (m, 2H), 2.99–3.06 (m, 1H), 2.72–2.76
(m, 2H), 2.40–2.56 (m, 6H), 1.88 (s, 1H), 1.27 (d, J = 7.0 Hz, 6H);
13C NMR (75 MHz, CDCl3) d 170.93, 169.33, 167.74, 135.55,
134.08, 132.99, 130.59, 129.28, 126.39, 99.85, 57.51, 45.48, 45.00,
26.47, 21.75.
4.1.6.8. Synthesis of (3,5-bis-trifluoromethylphenyl)(4-(2-
(3,3-dimethylbutylamino)ethyl)piperazin-1-yl)methanone
(1h) .
Reaction of 6d with 3,3-dimethylbutanalaccording to
the procedure described above afforded 1h in63% yield as yellow
oil after column chromatography (SiO2, CH2Cl2/MeOH = 20:1): 1H
NMR (300 MHz, CDCl3) d 7.94 (s, 1H), 7.87 (s, 2H), 3.80–3.83 (m,
2H), 3.38–3.42 (m, 2H), 2.65–2.75 (m, 2H), 2.50–2.63 (m, 8H),
1.39–1.44 (m, 2H), 0.91 (s, 9H); 13C NMR (75 MHz, CDCl3) d
166.97, 137.86, 132.13 (q, J = 33.8 Hz), 127.45, 123.50, 122.86 (q,
J = 271.3 Hz), 57.87, 46.51, 46.19, 44.14, 29.81, 29.58.
4.1.6.3. Synthesis of (3,5-dichlorophenyl)(4-(2-((3-isopro-
pylisoxazol-5-yl)methylamino)ethyl)piperazin-1-yl)methanone

M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
1147
4.1.6.9. Synthesis of (4-(2-(3-(1H-benzo[d]imidazol-2-yl)pro-
pylamino)ethyl)piperazin-1-yl)(4-chlorophenyl)methanone
TFA, and the resulting solution was stirred at rt for 5 h. After reac-
tion, the mixture was made basic with saturated NaHCO3 solution.
This mixture was extracted with CH2Cl2 three times and the com-
bined organic layer was dried over MgSO4.The crude compound
was purified by column chromatography (SiO2,CH2Cl2/
MeOH = 10:1?CHCl3/MeOH/H2O/NH4OH = 70:30:3:3) to give the
desired product (40.2 mg, 0.087 mmol, 62% yield): 1H NMR
(300 MHz, CDCl3) d 7.53–7.57 (m, 3H), 7.41–7.42 (m, 1H), 7.19–
7.24 (m, 4H), 3.66 (s, 2H), 3.28 (s, 2H), 3.05 (t, J = 6.8 Hz, 3H),
2.31–2.57 (m, 10H), 1.92–1.96 (m, 3H); 13C NMR (75 MHz, CDCl3)
d 167.22, 155.70, 138.45, 137.96, 135.45, 129.87, 125.44, 122.78,
115.11m 56.23, 53.54, 51.71, 50.41, 27.01, 25.95.
(1i).
Reaction of 6a with tert-butyl 2-(3-oxopropyl)-1H-
benzo[d]imidazole-1-carboxylate according to the procedure
described above afforded tert-butyl 2-(3-(2-(4-(4-chloroben-
zoyl)piperazin-1-yl)ethylamino)propyl)-1H-benzo[d]imidazole-1-
carboxyl atein 28% yield after column chromatography (SiO2,
CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d 7.29–7.37 (m,
5H), 6.83–6.91 (m, 2H), 6.65–6.67 (m, 1H), 3.21–3.62 (m, 10H),
2.46–2.65 (m, 6H), 1.96–2.11 (m, 2H), 1.47 (s, 9H).
To a solution of the tert-butyl 2-(3-(2-(4-(4-chlorobenzoyl)pip-
erazin-1-yl)ethylamino)propyl)-1H-benzo[d]imidazole-1-carbox-
ylate obtained above (105.9 mg, 0.2 mmol) in CH2Cl2 was added
30% TFA, and the resulting solution was stirred at rt for 5 h. After
reaction, the mixture was made basic with saturated NaHCO3 solu-
tion. This mixture was extracted with CH2Cl2 three times and the
combined organic layer was dried over MgSO4. The crude com-
pound was purified by column chromatography (SiO2, CH2Cl2/
MeOH = 10:1?CHCl3/MeOH/H2O/NH4OH = 70:30:3:3) to give the
desired product (37 mg, 0.087 mmol, 44% yield): 1H NMR
(300 MHz, CDCl3) d 7.45–7.50 (m, 2H), 7.31–7.40 (m, 4H), 7.15–
7.20 (m, 2H), 3.40–3.73 (m, 4H), 3.03 (t, J = 6.4 Hz, 2H), 2.79–2.88
(m, 4H), 2.55–2.62 (m, 6H), 1.98–2.06 (m, 2H); 13C NMR
(75 MHz, CDCl3) d 169.24, 155.06, 140.40, 138.64, 135.91, 135.84,
128.82, 128.58, 122.18, 115.00, 56.75, 49.22, 45.86, 28.03, 26.29.
4.1.6.12. Synthesis of (4-(2-(3-(1H-benzo[d]imidazol-2-yl)pro-
pylamino)ethyl)piperazin-1-yl)(3,5-bis(trifluoro-
methyl)phenyl)methanone (1l).
Reaction of 6d with tert-butyl
2-(3-oxopropyl)-1H-benzo[d]imidazole-1-carboxylate according to
the procedure described above afforded tert-butyl 2-(3-(2-(4-(3,5-
bis-trifluoromethylbenzoyl)piperazin-1-yl)ethylamino)propyl)-1H-be
nzo[d]imidazole-1-carboxylatein 23% yield aftercolumn chromatogra-
phy (SiO2, CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d 7.85–
7.98 (m, 5H), 7.38–7.67 (m, 2H), 3.44–3.79 (m, 4H), 3.31–3.63 (m,
6H), 2.89 (t, J = 6.4 Hz, 2H), 2.49–2.67 (m, 4H), 1.99–2.16 (m, 2H),
1.49 (s, 9H).
To a solution of tert-butyl 2-(3-(2-(4-(3,5-bis-trifluoromethylben-
zoyl)piperazin-1-yl)ethylamino)propyl)-1H-benzo[d]imidazole-1-car
boxylate obtained above (102.2 mg, 0.16 mmol) in CH2Cl2 was added
30% TFA, and the resulting solution was stirred at rt for 5 h. After
reaction, the mixture was made basic with saturated NaHCO3 solu-
tion. This mixture was extracted with CH2Cl2 three times and the
combined organic layer was dried over MgSO4.The crude compound
was purified by column chromatography (SiO2, CH2Cl2/
MeOH = 10:1?CHCl3/MeOH/H2O/NH4OH = 70:30:3:3) to give the de-
sired product (40.7 mg, 0.077 mmol, 48% yield): 1H NMR (300 MHz,
CDCl3) d 7.95 (s, 1H), 7.85 (s, 2H), 7.47–7.51 (m, 2H), 7.16–7.20 (m,
2H), 3.76 (s, 2H), 3.32–3.35 (m, 2H), 3.06 (t, J = 6.3 Hz, 2H), 2.81–
2.90 (m, 4H), 2.47–2.66 (m, 6H), 2.01–2.06 (m, 2H); 13C NMR
4.1.6.10. Synthesis of
yl)propylamino)ethyl)piperazin-1-yl)(4-chlorophenyl)metha-
none (1j) . Reaction of 6b with tert-butyl 2-(3-oxopropyl)-1H-
(4-(2-(3-(1H-benzo[d]imidazol-2-
benzo[d]imidazole-1-carboxylate according to the procedure described
above afforded tert-butyl 2-(3-(2-(4-(3,4-dichlorobenzoyl)piperazin-1-
yl)ethylamino) propyl)-1H-benzo[d]imidazole-1-carboxylatein 25% yield
after column chromatography (SiO2, CH2Cl2/MeOH = 20:1): 1H NMR
(300 MHz, CDCl3) d: 7.48–7.51 (m, 2H), 7.39 (s, 1H), 7.23 (dd, J = 8.2,
2.0 Hz, 1H), 6.87–6.96 (m, 2H), 6.69 (dd, J = 7.7, 1.5 Hz, 1H), 4.10–4.14
(m, 2H), 3.65 (t, J = 6.7 Hz, 2H), 3.40–3.50 (m, 4H), 2.48–2.69 (m, 8H),
1.95–2.01 (m, 2H), 1.51 (s, 9H).
To a solution of tert-butyl 2-(3-(2-(4-(3,4-dichlorobenzoyl)pip-
erazin-1-yl)ethylamino)propyl)-1H-benzo[d]imidazole-1-carbox-
ylate (100 mg, 0.18 mmol) obtained above in CH2Cl2 was added
30% TFA, and the resulting solution was stirred at rt for 5 h. After
reaction, the mixture was made basic with saturated NaHCO3 solu-
tion. This mixture was extracted with CH2Cl2 three times and the
combined organic layer was dried over MgSO4.The filtrate was con-
centrated under reduced pressure to give white solid (58 mg,
0.13 mmol, 70% yield): 1H NMR (300 MHz, CDCl3) d: 7.48–7.52
(m, 4H), 7.18–7.24 (m, 3H), 3.40–3.76 (m, 4H), 3.08 (t, J = 6.3 Hz,
2H), 2.79–2.89 (m, 4H), 2.47–2.64 (m, 6H), 1.97–2.05 (m, 2H);
13C NMR (75 MHz, CDCl3) d: 167.91, 155.25, 138.85, 135.50,
134.32, 133.15, 130.77, 129.38, 126.50, 122.06, 114.73, 57.25,
49.54, 45.86, 28.39, 26.72.
(75 MHz, CDCl3) d 166.98, 155.04, 138.60, 137.67, 132.16 (q,
J = 33.8 Hz), 127.42, 123.59, 123.58 (q, J = 270.8 Hz), 122.05, 114.58,
56.69, 49.25, 45.54, 28.10, 26.25.
4.2. Biological assay for synthesized compounds against 5-
HT1AR and 5-HT7R
This screen was performed by the National Institute of Mental
Health Psychoactive Drug Screening Program (PDSP). Radioligands
were purchased by PDSP from Perkin–Elmer or GE Healthcare.
Competition binding assays were performed using transfected or
stably expressing cell membrane preparations as previously de-
scribed23,24 and are available online (http://pdsp.med.unc.edu).
All experimental details are available online (http://pdsp.med.un-
c.edu/UNC-CH%20Protocol%20Book.pdf).
4.1.6.11. Synthesis of(4-(2-(3-(1H-benzo[d]imidazol-2-yl)pro-
pylamino)ethyl)piperazin-1-yl)(3,5-dichlorophenyl)methanone
4.3. Docking study
(1k).
1H-benzo[d]imidazole-1-carboxylate according to the procedure
described above afforded tert-butyl 2-(3-(2-(4-(3,5-dic-
hlorobenzoyl)piperazin-1-yl)ethylamino)propyl)-1H-benzo[d]imid-
azole-1-carboxylatein 13%yield after column chromatography(SiO2,
CH2Cl2/MeOH = 20:1): 1H NMR (300 MHz, CDCl3) d 7.87–7.90 (m,
1H), 7.64–7.67 (m, 1H), 7.41 (s, 1H), 7.21–7.31 (m, 4H), 3.72 (s,
2H), 3.19–3.38 (m, 4H), 2.31–2.69 (m, 10H), 2.06 (s, 2H), 1.69 (s,
9H), 1.49–1.65 (m, 1H).
Reaction of 6c with tert-butyl 2-(3-oxopropyl)-
4.3.1. Preparation of model structures
For homology modeling of human 5-HT1Aand 5-HT7 serotonin
receptors, the crystal structure of the seven helix bundle of b2-
adrenergic receptor (PDB code 2rh1) was used as a template.26 Ami-
noacid sequences of the receptors (P08908 for 5-HT1AR andP34969
for 5-HT7R) were downloaded from the Uniprot database (http://
www.uniprot.org) and the starting homology models of each recep-
tor were generated using the SwissModel server (accessible from the
program DeepView/SwissPdb-Viewer). Further modifications of the
models were performed using components of Schrödinger Suite
2008. The models were subjected to energy minimization and
To a solution of tert-butyl 2-(3-(2-(4-(3,5-dichlorobenzoyl)pip-
erazin-1-yl)ethylamino)propyl)-1H-benzo[d]imidazole-1-carboxy-
lateobtained above (80.4 mg, 0.14 mmol) in CH2Cl2 was added 30%

1148
M. K. Kim et al. / Bioorg. Med. Chem. 20 (2012) 1139–1148
molecular dynamics simulations (MD) in order to obtain a stable,
low-energy conformation. Energy minimization was performed
using a conjugate gradient minimize (0.05 convergence criteria),
the OPLS-AA force field, and GB/SA continuum water model. MD
simulations were performed by pre-equilibration for 100 ps and
simulation for 1 ns at 300 K with a 1-fs time step and SHAKE applied
to all bonds to hydrogen. In the next step, induced fit docking (IFD)
was involved for ligand-based optimization of the receptors.27,28
For this purpose, the representative arylpiperazine ligands with high
affinity for given receptors, MP34922 for 5-HT1AR and UCM-56009 for
5-HT7R, were chosen. Receptor models found in the top-scored com-
plexes were selected to serve as molecular targets in further docking
studies.
References and notes
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ligands were then docked into the homology models using GLIDE
4.0 (http://www.schrodinger.com) with H-bond constraint be-
tween Asp116 (5-HT1AR) or Asp162 (5-HT7R) and the protonated
nitrogens of the ligands. The default setting of the extreme preci-
sion mode of GLIDE was employed for the docking, and up to ten
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This research was supported by Basic Science Research Program
through the National Research Foundation of Korea (NRF) funded
by the Ministry of Education, Science and Technology
(2011-0005041) and Priority Research Centers Program through
the National Research Foundation of Korea (NRF) funded by the
Ministry of Education, Science and Technology (2009-0093824),
and by a grant from Next Generation Bio-Green 21 PJ007982 (Kor-
ea Rural Development Administration).
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