Synthesis and antidepressant effect of novel aralkyl piperazine and piperidine derivatives targeting SSRI/5-HT1A/5-HT7

Article abstract of DOI:10.1016/j.bmcl.2019.126703

A series of novel aralkyl piperazine and piperidine derivatives were synthesized, and evaluated for their serotonin reuptake inhibitory and 5-HT_1A/5-HT₇ receptors affinities activity. Antidepressant activities in vivo of the selective compound were screened using the forced swimming test (FST) and tail suspension test (TST). The results indicated that compound 19a exhibited high affinities for the 5-HT_1A/5-HT₇ receptors (5-HT_1A, K_i = 12 nM; 5-HT₇, K_i = 3.2 nM) coupled with potent serotonin reuptake inhibition (IC₅₀ = 14 nM) and showed a marked antidepressant-like effect in the FST and TST models.

Full text of DOI:10.1016/j.bmcl.2019.126703

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Synthesis and antidepressant effect of novel aralkyl piperazine
and piperidine derivatives targeting SSRI/5-HT1A/5-HT7
Zheng-Song Gu, Wen-Tao Wang, Hao Qian, Ai-Nan Zhou, Hong-
Bin Sun, Qing-Wei Zhang, Jian-Qi Li
PII:
S0960-894X(19)30660-2
DOI:
https://doi.org/10.1016/j.bmcl.2019.126703
BMCL 126703
Reference:
To appear in:
Received date:
Revised date:
Accepted date:
8 July 2019
28 August 2019
14 September 2019
Please cite this article as: Z.-S. Gu, W.-T. Wang, H. Qian, et al., Synthesis and
antidepressant effect of novel aralkyl piperazine and piperidine derivatives targeting SSRI/
5-HT1A/5-HT7, (2019), https://doi.org/10.1016/j.bmcl.2019.126703
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Synthesis and antidepressant effect of novel
aralkyl piperazine and piperidine derivatives
targeting SSRI/5-HT1A/5-HT7
Zheng-Song Gu a, †, Wen-Tao Wang a, b, †, Hao Qian a, Ai-Nan Zhou a, Hong-Bin Sun c,
Qing-Wei Zhang a, *, Jian-Qi Li a, *
a Shanghai Institute of Pharmaceutical Industry, China State Institute of Pharmaceutical Industry,
285 Gebaini Road, Shanghai 201203, P.R. China
b
School of Engineering, China Pharmaceutical University, 639 Longmian Avenue, Nanjing
211198, P.R. China
c
Jiangsu Key Laboratory of Drug Discovery for Metabolic Disease and State Key Laboratory of
Natural Medicines, China Pharmaceutical University, 24 Tongjia Xiang, Nanjing 210009, P.R.
China
*Corresponding author: Tel.: +86-021-20572128. E-mail: lijianqb@126.com; gzsliuyan@163.com;
† These authors contributed equally to this work.
Abstract
A series of novel aralkyl piperazine and piperidine derivatives were synthesized,
and evaluated for their serotonin reuptake inhibitory and 5-HT1A/5-HT7 receptors
affinities activity. Antidepressant activities in vivo of the selective compound were
screened using the forced swimming test (FST) and tail suspension test (TST). The
results indicated that compound 19a exhibited high affinities for the 5-HT1A/5-HT7
receptors (5-HT1A, Ki = 12 nM; 5-HT7, Ki = 3.2 nM) coupled with potent serotonin
reuptake inhibition (IC50 = 14 nM) and showed a marked antidepressant-like effect in
the FST and TST models.
Key words
Antidepressant, serotonin reuptake inhibition, 5-HT1A receptor, 5-HT7 receptor

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Depression is a common mental disorder characterized by symptoms like
anhedonia, decreased energy, impaired cognitive functions and other actions, with an
estimated 350 million people affected worldwide.1 According to the WHO, the serious
depressive illness will have become the main cause of disability by 2030. 2 Although a
plethora of antidepressants are in market, targeting mainly monoamine serotonin and
norepinephrine transporters,3 there is still a significant unmet medical need for
management of this disorder, including therapeutics delayed onset, treatment
resistance and adverse effects such as sexual dysfunction.4
Currently, it has been suggested that drugs combining activities of selective
serotonin reuptake inhibitor and 5-HT receptor subtypes may form a novel strategy
for higher therapeutic efficacy of antidepressant. 5 Clinical research has demonstrated
that the antidepressant activity of selective serotonin reuptake inhibitor (SSRI) may be
augmented by coadministration of pindolol, an effect that has been attributed to
pindolol’s partial agonistic effect at the 5-HT1A receptor. 6 Meanwhile, 5-HT7 receptor
blockade has been shown to be efficacious in animal models of depression, e.g. the
mouse forced swimming test. 7 Furthermore, the combination of an SSRI with 5-HT7
receptor blockade synergistically augments the levels of 5-HT in the prefrontal cortex,
suggesting that 5-HT7 receptor antagonist in combination with serotonin transporter
(SERT) blockade has antidepressant potential. 8 Hence, it stands to reason that agents
with dual or multiple acting on SSRI/5-HT1A/5-HT7 have the potential to fill
significant clinical unmet needs in depression.
Previous reports from our laboratories have revealed two novel series (Chart. 1)
capable of modulating SSRI/5-HT1A/5-HT7 activities. 9-10 Specifically，the promising
compound 2 possessed good oral pharmacokinetic properties and exhibited significant
antidepressant activity in vivo. Our SAR studies revealed that 3-propyl indole moiety
produced 5-HT uptake inhibition pharmacophore moiety, while biphenyl piperazine
moiety served as 5-HT1A/5-HT7 pharmacophore moiety. Meanwhile, the introduction
of a fluoro group into the para-position of the piperazine ring could improve
metabolic stability significantly. In order to expand upon the type of structures that
can potentially improve SSRI/5-HT1A/5-HT7 activities and to probe the structure–

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activity relationship (SAR), we replaced the biphenyl portion of 2 with other aromatic
substituents. We herein describe the synthesis and biological evaluation of a series of
novel aralkyl piperazine and piperidine derivatives. The general structure of target
compounds I and II is shown in Chart 2.
All compounds were evaluated for binding affinities for 5-HT1A/5-HT7 receptors
and inhibiting 5-HT reuptake in vitro. The selected compounds were tested for
microsomal stabilities in vitro and antidepressant-like activities in vivo. Preliminary
results indicated several target compounds exhibited high affinities for 5-HT1A/5-HT7
receptors coupled with potent 5-HT reuptake inhibition and produced marked
antidepressant-like effects in vivo.
Chart 1. Structures of SSRI/5-HT1A/5-HT7 agents
Chart 2. Rational design of novel SSRI/5-HT1A/5-HT7 activity compounds
The synthesis of target compounds was prepared as outlined in Schemes 1-4. In
Scheme 1, a series of 5-fluoro-1H-indole piperazine and piperidine derivatives 6 were
prepared. The alcohols 3a and 3b were obtained by the Fischer indole synthesis

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reaction of 4-Fluorophenylhydrazine hydrochloride 2 with 2,3-dihydrofuran 1a and
3,4-dihydro-2H-pyran 1b, respectively. Bromination of compound 3a with
tetrabromomethane and triphenylphosphine provided compound 4a. Activation of
alcohol 3b with 4-toluenesulfonyl chloride in the presence of triethylamine at room
temperature provided compound 4b. The desired compounds 6 were offered by
nucleophilic substitution reaction of intermediates 4a-b with aryl piperazine and
piperidine derivatives 5. The four carbon linker compound 11 was prepared (Scheme
2). The acylation of the aromatic ring 7 with 4-chlorobutanoyl chloride provided
compound 8. Then intermediate 8 was reduced with triethylsilane and trifluoroacetic
acid to yield compound 9. The compound 10 was obtained by substitution of
intermediate 9 with 3-(piperazin-1-yl)benzo[d]isothiazole. The deprotection of
intermediate 10 with 4N aqueous NaOH solution gave the desired compound 11.
According to Scheme 3, the 5-fluoro-1H-indole piperazine analogs 15 were
prepared. 1-(4-fluorophenyl)piperazine 12 with N-bromosuccinimide in the presence
of pyridine by electrophilic substitution reaction afforded intermediate 13. The
resulting compound 13 was then treated with 3-(5-fluoro-1H-indol-3-yl)propyl
4-methylbenzenesulfonate 4b via SN2 mechanism to obtain intermediate 14. The
target compounds 15 were obtained via the Suzuki cross coupling reaction of
intermediate 14 with arylboronic acids.
The preparation of the desired 5-fluoro-1H-indole piperidine derivatives 19 was
depicted in Scheme 4. 4-(2-bromo-4-fluorophenyl)pyridine 16 was treated with
arylboronic acids to yield intermediate 17 by Suzuki cross coupling reaction. The
target compounds 19 was obtained by hydrogenation and substitution.
Scheme 1. Reagents and conditions: (a) H2SO4 (4%), DMAC, 100 oC; (b) for 4a: CBr4, PPh3,

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DCM, r.t.; for 4b: TsCl, Et3N, DMAP, DCM, r.t.; (c) for 6a: K2CO3, KI, DMF, 100 oC; for 6b and
6c : K2CO3, CH3CN, 80 oC.
o
Scheme 2. Reagents and conditions: (a) 4-chlorobutanoyl chloride, AlCl3, DCM, 0 C→r.t.; (b)
HSiEt3, TFA, 50 oC; (c) K2CO3, KI, DMF, 100 oC; (d) 4N NaOH, EtOH/THF, 80 oC.
Scheme 3. Reagents and conditions: (a) NBS, Pyridine, DCM, 0°C→r.t.; (b) K2CO3, CH3CN, 80
oC; (c) Pd(dppf)Cl2, Na2CO3, Dioxane/H2O (3:1), 80 oC.
Scheme 4. Reagents and conditions: (a) Pd(dppf)Cl2, Na2CO3, Dioxane/H2O (3:1), 80 oC; (b) PtO2,
H2, AcOH, r.t.; (c) K2CO3, CH3CN, 80 oC.
All compounds were tested for their inhibition of 5-HT reuptake and binding
affinities for the 5-HT1A and 5-HT7 receptors. The binding affinity for 5-HT1A
receptor was determined by investigating the displacement of [3H]-8-OH-DPAT to

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HEK-293 cell membrane homogenates, 11 and the affinity for the 5-HT7 receptor was
determined by displacement binding to CHO cell membrane homogenates using
[3H]-LSD to previously procedures reported. 12 The inhibition of uptake of [3H]-5-HT
into rat brain synaptosomes through the serotonin transporter was also evaluated. 13
All compounds were initially screened at 10μM concentration, and then potent
compounds (inhibition >90%) were assayed to obtain their IC50 values. The detail
results were summarized in Table 1. The metabolic stability of the test compounds
was measured in vitro using rat and human liver microsomes, a system widely used to
evaluate the susceptibility to first-pass oxidative metabolism (Table 2-3).14 The
agonist property of preferred compound on 5-HT1AR was evaluated in the
[35S]guanosine 5’-O-(3-thio)- triphosphate ([35S]GTPγS) binding assay.15 Additionally,
the antagonist property of preferred compound against 5-HT7R was tested in the
cyclic adenosine 3', 5'-monophosphate (cAMP) functional cellular assays (Table 4). 16
The influence of the length of the linker on 5-HT reuptake inhibition in relation
to 5-HT1A and 5-HT7 receptor affinity was firstly explored in series I (Chart 2). The
three-carbon chain analog 6b displayed outstanding affinity for 5-HT1A/5-HT7
receptors (5-HT1A, Ki = 1.6 nM; 5-HT7, Ki = 2.2 nM) and better 5-HT reuptake
inhibitory activity (IC50 = 1.1 nM) (6b vs 6a). Similarly, three-carbon chain analog 6c
exhibited higher inhibitory for 5-HT reuptake (IC50 = 0.92 nM) and showed more than
90% inhibition for 5-HT1A/5-HT7 receptors at a concentration of 10 μM (6c vs 6a).
However, the compound 11 showed dramatically decreased inhibition of 5-HT
reuptake when the linker was elongated to four carbons (IC50 = 7.08 nM) (11 v 6b).
In our next series, we investigated the effects of replacing the biphenyl moiety
with other aromatic substituents (Chart 2). As shown in Table 1, the six-membered
aromatic heterocycle derivatives 15a-15b showed more than 90% inhibition for
5-HT1A/5-HT7 receptors and 5-HT reuptake at 10μM. Meanwhile, the role of various
substituted five-membered aromatic heterocycle groups was also explored.
Surprisingly, the thiophen-3-yl analog 15c displayed higher potency for the
5-HT1A/5-HT7 receptors and the 5-HT reuptake inhibition compared to 2 [RUI, IC50 =
4.0 nM; 5-HT1A, Ki = 8.4 nM; 5-HT7, Ki = 2.0 nM]. However, the thiophen-2-yl

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analog 15d showed dramatically decreased inhibition of 5-HT reuptake and reduced
affinities for the 5-HT1A and 5-HT7 receptors (15d vs 15c). Those results suggested
that the position of aromatic heterocycle groups had a great influence on 5-HT
reuptake/5-HT1A/5-HT7 activities. Similarly, the furan-3-yl analog 15e and
pyrazol-4-yl analog 15g-15h also exhibited high inhibition for 5-HT reuptake and
displayed excellent affinities for 5-HT1A/5-HT7 receptors, whereas pyrrol-3-yl analog
15f showed selective inhibition for 5-HT1A/5-HT7 receptors (inhibition ratio > 90%).
Finally, the effect of the piperazine ring was explored. When the biphenyl feature
remained, the replacement of the piperazine moiety with the piperidine moiety
increased inhibition of 5-HT reuptake (IC50 = 14 nM) and binding affinities for
5-HT1A (Ki = 12 nM) (19a vs 2). Meanwhile, the thiophen-3-yl analog 19b showed a
increase in inhibition of 5-HT1A (Ki = 2.6 nM) and a reduction in affinities for 5-HT7
(Ki = 11 nM) (19b vs 19a).
With the above information in hand, the potent compounds 6a-6c, 11, 15a-15c,
15e, 15g-15h and 19a-19b were selected for rat liver microsomal metabolic stability
evaluation in vitro. The results are shown in Table 2. The data indicated that
compound 19a displayed a better metabolic stability (t1/2 values was 48.6 min).
Moreover, compound 19a was tested for human liver microsomal metabolic stability
and the results indicated that 19a showed an acceptable profile (t1/2 values was 49.93
min) (Table 3).
The functional properties of compound 19a on 5-HT1AR and 5-HT7R were also
evaluated. As shown in Table 4, compound 19a showed agonist property (EC50=1180
nM) at 5-HT1AR and antagonist effect (IC50=650 nM) at 5-HT7R in vitro functional
activity assays.
Compounds 19a was further selected for profiling in the mouse forced swim test
(FST). 17 The compound was administered orally once daily to ICR male mice for 7
days at doses of 10, 20 and 40 mg/kg/day (PO), respectively. For comparative
purposes, Vortioxetine was acted as a positive control (40 mg/kg/day, PO). The results
are shown in Figure 1. Compared with vehicle, the selected compound 19a reduced
immobility times in the FST in a dose-dependent manner that was statistically

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significant at 20 and 40 mg/kg (PO). The positive control, Vortioxetine, also produced
a statistically significant reduction of immobility time at 40 mg/kg dose.
Compound 19a was further characterized in the mouse tail suspension test
(TST).18 The results are shown in Figure 2. Vilazodone was used as a reference
compound (40 mg/kg/day, PO). The compound 19a was administered orally once
daily to ICR male mice for 7 days at doses of 10, 20 and 40 mg/kg/day (PO).
Compared to the control group, compound 19a was dose dependently reduced the
immobility time in the TST, which was statistically significant at 20 and 40 mg/kg
doses.
In summary, a novel series of aralkyl piperazine and piperidine derivatives
were designed and synthesized. Several compounds showed outstanding binding
affinities at the 5-HT1A/5-HT7 receptors and excellent inhibition of 5-HT reuptake.
Especially, compound 19a exhibited a good metabolic stability and showed potent
efficacy in vivo in the animal test. In addition, compound 19a was found to be an
agonist at 5-HT1A receptor and antagonist at 5-HT7 receptor. It was suggested that
compound 19a could act as a potential candidate for a promising antidepressant. The
preliminary results lay a foundation for the future development of novel derivatives
targeting SSRI/5-HT1A/5-HT7 as potent multi-model antidepressant agents.

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Table 1
5-HT1A and 5-HT7 Receptor Binding and 5-HT Reuptake Inhibition of target compounds
compd
RUI (IC50, nM)
23 b
5-HT1A(Ki, nM)
1.6 b
5-HT7 (Ki, nM)
0.99 b
2.2 b
6a
6b
1.1 b
1.6 b
0.92 b
7.08 b
102.0% a
104.5% a
4.0 b
100.7% a
100.7% a
98.8% a
99.1% a
8.4 b
99.8% a
98.5% a
90.9% a
100.2% a
2.0 b
6c
11
15a
15b
15c
21 b
38 b
11 b
15d
3.0 b
78.9% a
1.3 b
15 b
97.4% a
6.3 b
11 b
99.1% a
0.9 b
15e
15f
15g
2.7 b
14 b
8.4 b
12 b
1.0 b
3.2 b
15h
19a
14 b
2.9 b
0.5c
2.6 b
9.5 b
0.2c
11 b
26 b
3900c
19b
Vortioxetine
Vilazodone
a Percent inhibition measured at a concentration of 10μM.
b IC50 and Ki values were obtained from 8 concentrations of the compound, each in duplicate.
(binding assays were conducted by Eurofins Cerep SA, Celle L’Evescault, France).
c Data for Vilazodone was reported by Heinrich et al. 19
Table 2
Rat Liver Microsomal Metabolic Stability Assay
Compd.
6a
T1/2 (min)
3.1
CL (μL/min/mg)
452.1
3.5
396.9
6b
4.6
300.9
6c
3.4
402.8
11
6.6
209.3
15a
15b
15c
15e
5.8
239.7
24.5
10.3
56.5
134.0

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7.2
9.5
191.6
146.4
28.5
15g
15h
48.6
18.33
9
19a
75.6
19b
153.2
48.59
162.6
Vortioxetine
Vilazodone
Omeprazole
28.52
8.5
Table 3
Human Liver Microsomal Metabolic Stability Assay
Compd.
t1/2 (min)
CL (μL/min/mg)
19a
49.93
214.32
53.32
27.76
6.47
Vortioxetine
26.00
31.03
Vilazodone
44.67
Dextromethorphan
Table 4
5-HT1A and 5-HT7 Receptor functional activity assays
5-HT1A
5-HT7
Compd.
EC50, nM
IC50, nM
EC50, nM
IC50, nM
650
1180
18
-
-
-
-
-
-
-
19a
Serotonin
Methiothepin
-
0.82
(Functional assays were conducted by Eurofins Cerep SA, Celle L'Evescault, France).
compound 19a (mg/kg, PO)
Fig. 1. Effect of treatment of mice with compound 19a at graded doses on the immobility time in

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the mouse forced swim test. Results are represented as mean ± SEM. with n = 10 in each group.
Values are significant at *P < 0.05, **P < 0.01 when compared with vehicle group.
compound 19a (mg/kg, PO)
Fig. 2. Effect of treatment of mice with compound 19a at graded doses on the immobility time in
the mouse tail suspension test. Results are represented as mean ± SEM. with n = 10 in each group.
Values are significant at *P < 0.05, **P < 0.01 when compared with vehicle group.
Acknowledgements
The authors gratefully acknowledge the financial support from the National
Science and Technology Major Project (Grant No. 2018ZX09711002-002-009), the
National Natural Science Foundation of China (Grant No. 81703358), Shanghai
Science and Technology Committee(Grant No. 18QB1404200 ， 16431903400,
14431901500, 17431903900) . We thank M.S. Feng-hua Gu (Department of
pharmacology, Shanghai Institute of Pharmaceutical Industry, China) for in vivo
studies.
References and notes
1. Depression Fact Sheet; World Health Organization (WHO): Geneva, 2017;
https://www.who.int/en/news-room/fact-sheets/detail/depression (accessed March 22, 2018).
2. Sheehan DV, Nakagome K, Asami Y, et al. J Affect Disord. 2017; 215: 299-313.
3. Gelenberg AJ, Hopkins HS. Am J Med. 2007;120: 105-108.

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4. Fournier JC, DeRubeis RJ, Hollon SD, et al. J Am Med Assoc. 2010; 303: 47-53.
5. Millan M J. Neurotherapeutics. 2009; 6: 53-77.
6. Artigas F, Adell A, Celada P. Curr Drug Targets. 2006; 7: 139-147.
7. Wesołowska A, Tatarczyńska E, Nikiforuk A, et al. Eur J Pharmacol. 2007; 555: 43-47.
8. Bonaventure P, Kelly L, Aluisio L, et al. J Pharmacol Exp Ther. 2007; 321: 690-698.
9. Gu ZS, Xiao Y, Zhang QW, Li JQ. Bioorg Med Chem Lett. 2017; 27: 5420-5423.
10. Gu ZS, Zhou AN, Xiao Y, et al. Eur J Med Chem. 2018;144: 701-715.
11. Mulheron JG, Casanas, SJ, Arthur JM, et al. J Biol Chem. 1994; 269: 12954-12962.
12. Shen Y, Monsma FJ, Metcalf MA, et al. J Biol Chem.1993; 268: 18200-18204.
13. Perovic S, Muller WEG, Forsch A. Drug Res. 1995; 45: 1145-1148.
14. Obach RS, Baxter JG, Liston TE, et al. J Pharmacol Exp Ther. 1997; 283: 46-58.
15.Adlersberg M, Arango V, Hsiung S, et al. J Neurosci Res. 2000; 61: 674 - 685.
16. Adham N, Zgombick JM, Bard J, et al. J Pharmacol Exp Ther. 1998; 287: 508-514.
17. Porsolt RD, Bertin A, Jalfre M. Arch Int Pharmacodyn Ther. 1977; 229: 327-336.
18. Steru L, Chermat R, Thierry B, Simon P. Psychopharmacology (Berl). 1985; 85: 367-370.
19. Heinrich, T, Bottcher, H, Gericke, R, et al. J. Med. Chem. 2004; 47, 4684-4692.

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Highlights
 A series of aralkyl piperazine and piperidine derivatives have been synthesized.
 All compounds were evaluated for SSRI/5-HT1A/5-HT7 activities in vitro.
 The potent compounds were screened using the forced swimming test and the tail
suspension test in vivo.
 Compound 19a displayed
a
good metabolic stability and potent
antidepressant-like profiles in vitro and in vivo.

Figure 1

Figure 2

Figure 3

Figure 4