TRPV1 modulators: Synthesis and in vitro evaluation of 1-heteroaryl piperidinecarboxamide and piperazinylurea derivatives

Article abstract of DOI:10.1016/j.ejmech.2015.05.041

Abstract A series of new 1-heteroaryl piperidinecarboxamide and piperazinylurea derivatives was synthesized and evaluated as TRPV1 modulators in a Ca²⁺ channel assay in HEK-293 cells overexpressing the human recombinant TRPV1 channel. Structura

Full text of DOI:10.1016/j.ejmech.2015.05.041

European Journal of Medicinal Chemistry 100 (2015) 129e138
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
TRPV1 modulators: Synthesis and in vitro evaluation of 1-heteroaryl
piperidinecarboxamide and piperazinylurea derivatives
,
*
,
,
Cenzo Congiu a 1, Valentina Onnis a 1, Gianfranco Balboni a, Aniello Schiano-Moriello b,
Vincenzo Di Marzo b, Luciano De Petrocellis b
a Department of Life and Environmental Sciences, Unit of Pharmaceutical, Pharmacological and Nutraceutical Sciences, University of Cagliari, Via Ospedale
72, 09124 Cagliari, Italy
b Endocannabinoid Research Group, Institute of Biomolecular Chemistry, National Research Council, Via Campi Flegrei 34, Comprensorio Olivetti, 80078
Pozzuoli, Naples, Italy
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of new 1-heteroaryl piperidinecarboxamide and piperazinylurea derivatives was synthesized
and evaluated as TRPV1 modulators in a Ca2þ channel assay in HEK-293 cells overexpressing the human
recombinant TRPV1 channel. Structural variations in the putative key portions of the molecules afforded
several compounds endowed with agonist and/or antagonist/desensitizing activity at low micromolar
concentration. As promising examples from this series, the piperidine-3-carboxamide derivative 31
exerts agonist/desensitizing activity at low micromolar concentration, while piperazinylurea derivatives
39 and 41 act as antagonists with sub-micromolar potency.
Received 4 February 2015
Received in revised form
21 April 2015
Accepted 26 May 2015
Available online 6 June 2015
Keywords:
TRPV1 receptor
© 2015 Elsevier Masson SAS. All rights reserved.
Calcium channel assay
Piperidinecarboxamides
Piperazinylureas
1. Introduction
neuropathy [7]. Moreover, desensitization of the afferent nerves
using TRPV1 agonists such as capsaicin has been shown to give
The transient receptor potential vanilloid-1 (TRPV1) is
nonselective cation channel, with preference for calcium,
expressed predominantly on unmyelinated pain-sensing nerve fi-
bers (C-fibers) and small А fibers in the dorsal root, trigeminal, and
a
encouraging results in the treatment of bladder dysfunction asso-
ciated with spinal cord injury and multiple sclerosis [8]. On the
other hand, the identification of capsazepine (Fig. 1, B) [9], a
competitive antagonist of capsaicin binding, provided a proof of
principle for the discovery of novel analgesics based on the block-
ing of activation of TRPV1 by endogenous stimuli [10]. Given the
therapeutic potential expected for both TRPV1 agonists [11], and
antagonists [12] in the last years there has been a growing interest
in developing TRPV1 ligands that could be used for prophylaxis and
treatment of conditions and diseases including acute and chronic
nociceptive pain, ischaemia, inflammatory disorders, urinary in-
continence, and/or overactive bladder (for a recent review see
Ref. [13]).
As a continuation of our efforts to identify anti-inflammatory/
analgesic agents [14], we designed new compounds as TRPV1
modulators, based on the structural motif developed for several
series of TRPV1 agonists and antagonists. In this context, Purdue
Pharma, following an initial high throughput screening (HTS)
approach, reported the design and synthesis of a series of 4-(2-
pyridyl)piperazine-1-carboxamide analogs including N-(4-tert-
butylphenyl)-4-(3-chloropyridin-2-yl)piperazine-1-carboxamide
a
d
nodose ganglia [1]. TRPV1, identified as the receptor for the vanil-
loid compound, capsaicin (Fig. 1, A), is activated by endogenous
stimuli such as heat and low pH (<5.9) [2], and endogenous ligands
including anandamide [3], and arachidonic acid metabolites [4].
The activation of the vanilloid receptor by agonists triggers
cation influx resulting in excitation of primary sensory neurons,
and ultimately the central perception of pain. The initial excitation
is followed by a refractory state of desensitization, where the C-
fiber sensory neurons become unresponsive to TRPV1 agonists and
other inflammatory mediators [1,5]. This desensitization represents
a basis for therapeutic use of vanilloid receptor agonists in the
management of acute and chronic nociceptive pain, produced by
osteoarthritis, rheumatoid arthritis [6], and diabetic peripheral
* Corresponding author.
E-mail address: ccongiu@unica.it (C. Congiu).
These authors equally contribute to the present paper.
1
http://dx.doi.org/10.1016/j.ejmech.2015.05.041
0223-5234/© 2015 Elsevier Masson SAS. All rights reserved.

130
C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
Fig. 1. Structure of prototypical TRPV1 ligands: Capsaicin (A, agonist), Capsazepine (B, antagonist).
(BCTC). BCTC showed potent TRPV1 antagonist activity in a HEK293
cell line against capsaicin and pH 5.5 activation in vitro, and
effectively reversed the behavioral effects of inflammatory and
neuropathic pain in rats [15]. In a similar manner, through internal
high-throughput screening efforts, Swanson, et al. identified
several series of agonists and antagonists, including a series of
pyridinylpiperazine ureas [16]. In the present study, we planned to
cyano-6-(trifluoromethyl)pyridin-2-yl)piperidine-4-carboxamides
4e23 was synthesized. Heterocyclization of enaminonitrile 1 [17]
with trifluoroacetylvinyl ether 2 in boiling MeCN, followed by
alkaline cleavage afforded 1-(3-cyano-6-(trifluoromethyl)pyridin-
2-yl)piperidine-4-carboxylic acid (3). Coupling of the carboxylic
acid
phenethyl-amine mediated by 1-(3-dimethylaminopropyl)-3-
ethylcarbodiimide hydrochloride (EDC) and 1-
3 with an appropriately substituted aryl-, benzyl-, or
explore
a series of 1-heteroaryl piperidinecarboxamides and
piperazinylureas, and related structures bearing an aryl-, benzyl-,
and phenethyl-amide moiety (Fig. 2).
hydroxybenzotriazole (HOBt) in DCM as the solvent gave the am-
ides 4e23 in 46e92% yields.
In an effort to define the critical requirements for activity, and
towards the understanding of molecular determinants of novel
compounds as TRPV1 ligands, our approaches involved making
structural modifications in the putative key pharmacophoric por-
tions of the molecule. Our starting approach was focused to syn-
thesize a series of new compounds bearing a 6-CF3-3-CN-pyridine
fragment in the A region, and to explore the structureeactivity
relationships (SARs) established around the heteroaryl group
following structural changes. Then we evaluated the effect of some
variation of the cycloaliphatic linker bearing the amide function
(region B); and finally we studied the effect of the introduction of a
differently polar substituent in the aromatic ring (region C). Herein
we report the synthesis of a series of these ligands and preliminary
results of a cell-based assay utilizing the Ca2þ permeability of the
TRPV1 channel.
In a similar manner, amidation of acids 1-(3-cyanopyridin-2-yl)
piperidine-4-carboxylic (24) [18], 1-(4-(trifluoromethyl)pyrimidin-
2-yl)piperidine-4-carboxylic (25) [19], and 1-(4-(trifluoromethyl)
pyrimidin-2-yl)piperidine-3-carboxylic (26) with 4-fluoroaniline
and 4-hydroxyaniline led to analogs 27e32 in 52e89% yields
(Scheme 2). Compound 26 has been prepared upon coupling 2-
chloro-4-trifluoromethylpyrimidine (45) with ethyl piperidine-3-
carboxylate (46) in DMF solution in presence of potassium car-
bonate, followed by alkaline hydrolysis (Scheme 2).
Piperazinylurea derivatives have been prepared as shown in
Scheme 3. Coupling of 1-heteroarylpiperazine 33 [20] and 34 [21]
with a variety of phenyl N-aryl-carbamates 35a-f in DMSO solu-
tion provided piperazinylureas 36e44 in 64e93% yields. With some
modification of reported procedures [22], phenyl N-aryl-carba-
mates 35a-f were synthesized upon reacting commercially avail-
able substituted anilines with phenyl chloroformate in THF solution
in presence of diisopropyl ethylamine (DIPEA) as a base (Scheme 4).
2. Results and discussion
2.2. Biological activity
2.1. Chemistry
The activity of the new compounds on TRPV1 channel was
determined by measuring the effect on intracellular Ca2þ ([Ca2þ]i)
elevation in HEK-293 cells overexpressing the human recombinant
TRPV1 receptor (hTRPV1), with or without the subsequent (after
The synthetic routes employed to obtain the target compounds
are shown in Schemes 1e4. According to Scheme 1, a series of 1-(3-
5 min) stimulation of cells with capsaicin (0.1 mM), as previously
reported [23]. The efficacy of TRPV1 agonism was determined by
normalizing the effect of the compounds to the maximum effect on
[Ca2þ]i increase observed with application of 4
m
M ionomycin, i.e.
the efficacy was expressed as percent of the effect obtained with
M ionomycin. Potency was instead expressed as the concen-
4
m
tration of test substance exerting a half-maximal agonist effect
(EC50). Antagonist/desensitizing behavior was evaluated against
0.1 mM capsaicin. Data were expressed as the concentration exert-
ing a half-maximal inhibition of capsaicin-induced [Ca2þ]i elevation
(IC50). The effect on intracellular Ca2þ exerted by capsaicin alone
was taken as 100%. All determinations were performed at least in
triplicate. The effects on functional activity on [Ca2þ]i elevation in
HEK-293 cells mediated by the TRPV1 channel of the compounds
herein studied are summarized in Table 1. Data for capsazepine
(CPZ) [24] and BCTC [15a] as standard compounds are also included
Fig. 2. Structural modifications of new modulators of the TRPV1 channel.

C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
131
Scheme 1. Reagents and conditions: (i) MeCN, reflux, 6 h; (ii) MeOH, 2 N aq NaOH, rt, 16 h, then HCl; (iii) EDC, HOBt, DCM, rt, overnight.
Scheme 2. Reagents and conditions: (i) 4-Fluoro- or 4-hydroxyaniline, EDC, HOBt, DCM, rt, overnight; (ii) DMF, K2CO3, 100 ꢀC, overnight; (iii) MeOH, 2 N aq NaOH, rt, overnight, then
HCl.
Scheme 3. Reagents and conditions: DMSO, rt, 18 h.
in the Table. This assay led to the identification of various hTRPV1
modulators that exhibit agonist and/or antagonist/desensitizing
activity.
When exploring the SAR for the series of piperidine-4-
carboxamides bearing a 3-cyano-6-(trifluoromethyl)pyridine moi-
ety 4e23, the 4-fluorophenylamide 4 exhibits agonist/desensitizing
activity at low micromolar concentration. The substitution of the 4-
fluorine atom of 4 with either a 4-chlorine (5), 4-tert-butyl (8), 3-
CF3 (9), or 4-CF3 (10) group affords analogs showing antagonist
behavior with moderate inhibitory activity. In contrast, the sub-
stitution with oxygen-bearing groups to produce compounds 7,
11e14, results in loss of TRPV1 ligand property. However, the
phenol derivative 6 maintains a low activity as TRPV1 modulator.
Introduction of a benzyl or phenethyl substituent on the amide
function results in general loss of activity, probably due to a steric
hindrance, even though compounds 15 and 19, bearing a 4-fluoro-

132
C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
Scheme 4. Reagents and conditions: THF, DIPEA, 0 ꢀC to rt, 4 h.
and 4-trifluoromethylbenzylamide group respectively, retain
micromolar agonist/desensitizing activity in the TRPV1 Ca2þ assay.
Introduction of a 4-trifluoromethylpyrimidine group for the het-
eroaryl fragment (region A) of 4 leads to the analog 29 that exhibits
mixed functional activity with mild efficacy in the calcium channel
influx assay. However, the substitution of the piperidine-4-
identify a series of compounds endowed with agonist and/or
antagonist/desensitizing activity with moderate to good potency.
As promising examples from this series, the piperidine-3-
carboxamide derivative 31 exerts agonist/desensitizing activity at
low micromolar concentration, while piperazinylurea derivatives
39 and 41 act as antagonists with sub-micromolar potency. Across
the series of piperazinylurea derivatives, the presence of a 4-CF3-
pyrimidine moiety in the A region results in partial agonists of the
TRPV1 receptor; whereas, a 3-CN-pyridine moiety, preferably
combined with a bulky lipophilic substituent such as a tert-butyl or
CF3 group in the para position of the phenyl ring (region C), seems
to favor antagonistic activity. Prompted by herein reported pre-
liminary results, further developments across the putative key
portions of the molecules are now in progress, and the results will
be presented in due course.
carboxamide linker (region B) of 29 with
a piperidine-3-
carboxamide fragment affords compound 31, which acts as
agonist/desensitizer at low micromolar concentration, with
increased efficacy on [Ca2þ]i elevation when compared to both
analogs 4 and 29. In contrast, removal of the CF3 on the heteroaryl
fragment of 4 to give compound 27 results in loss of activity at the
TRPV1 receptor. Among piperazinylureas, compounds 39 and 41,
bearing a 3-cyanopyridine group in the A region, exhibit antagonist
activity with sub-micromolar IC50 inhibitory values comparable to
that of the prototypical competitive antagonist capsazepine [24].
Noteworthy, compound 39, bearing a 4-tert-butylphenyl substitu-
ent in the aromatic region, shows an IC50 inhibitory value of
4. Experimental section
0.16
m
M, lower than that reported for the analog bearing a 4-iso-
4.1. Chemistry
propylphenyl substituent in the aromatic region (IC50 ¼ 0.287
mM)
[15a]. On the other hand, the 4-trifluoromethylpyrimidine de-
rivatives 42, 43 retain mixed functional activity at micromolar
concentrations, whereas 44 acts as agonist/desensitizing with re-
sidual functional efficacy at the channel.
4.1.1. General methods
Unless otherwise noted, all solvents, including anhydrous sol-
vents and chemicals, were purchased from Aldrich Co. and/or Alfa
Aesar, and used without further purification. Melting points were
recorded on a Stuart Scientific melting point SMP1 apparatus and
are uncorrected. Positive-ion electrospray ionization (ESI) mass
spectra were recorded on a double-focusing Finnigan MAT 95 in-
strument with BE geometry. Proton nuclear magnetic resonance
spectra were recorded on a Varian Inova 500 spectrometer at
500 MHz, in DMSO-d6 as the solvent, and TMS as the internal
standard. Chemical shifts are expressed in ppm relative to tetra-
methylsilane. Splitting patterns are designated as singlet (s),
doublet (d), triplet (t), quartet (q), and multiplet (m). Infrared
spectra were run on Bruker Vector 22 spectrophotometer. Reaction
courses and product mixtures were routinely monitored by TLC on
silica gel (precoated F254 Merck plates). Developed plates were
visualized by a Spectroline ENF 260C/F UV apparatus. Concentra-
tion and evaporation of the solvent after reaction or extraction were
carried out on a rotary evaporator (Büchi Rotavapor) operating at
reduced pressure. Elemental analyses were carried out with a Carlo
Erba model 1106 elemental analyzer, and all values were within
0.4% of the calculated values, which indicates >95% purity of the
tested compounds.
Thus, in this series of compounds, the presence of a 4-CF3-py-
rimidine moiety in the A region leads to partial agonists with
moderate efficacy at the TRPV1 channel; whereas, a 3-CN-pyridine
moiety, preferably combined with a bulky lipophilic substituent
such as a tert-butyl or CF3 group in the para position of the phenyl
ring (region C), seems to favor antagonist activity. Moreover,
comparing compounds 8 and 10 with analogs 39 and 41, respec-
tively, removal of the CF3 from heteroaryl fragment along with
introduction of a piperazine for the piperidine linker in the B region
leads to increased inhibitory effects (Table 1). Such effects seem to
be correlated at least in part with enhanced hydrophilicity, as
assessed by the calculated logP values [25]. In fact, with respect to
compound 8 (logP ¼ 5.10), the above described structural changes
to give the analog 39 (logP ¼ 3.79) result in about 100-fold increase
in antagonistic effect; in a similar manner, passing from 10
(logP ¼ 4.29) to the analog 41 (logP ¼ 2.98) produces a ~10-fold
enhancement of activity. Probably, differences in hydrophilicity
combined with electronic and/or steric effects across this series of
compounds affect significantly their ability to interact with the
TRPV1 binding site, and influence both the kinetic and inhibitory
activity of antagonists at the channel [1,12c,26].
4.1.2. 1-(3-Cyano-6-(trifluoromethyl)pyridin-2-yl)piperidine-4-
carboxylic acid (3)
3. Conclusions
A solution of 3-amino-3-(4-(ethoxycarbonyl)piperidin-1-yl)
propenenitrile (1, 2.23 g, 10 mmol) and 4-ethoxy-1,1,1-trifluoro-3-
buten-2-one (2, 1.7 g, 10 mmol) in MeCN (50 ml) was refluxed
under stirring for 6 h. After evaporation of the solvent in vacuo, the
residue was dissolved in methanol (100 ml), and 2 N NaOH (25 ml)
was added. The mixture was stirred overnight at room tempera-
ture, and after evaporation of the solvent, the residue was dissolved
A
series of new 1-heteroaryl piperidinecarboxamide and
piperazinylurea derivatives was synthesized and evaluated as
TRPV1 modulators in a Ca2þ channel assay in HEK-293 cells over-
expressing the human recombinant TRPV1 receptor. Structural
variations in the putative key portions of the molecules allowed to

C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
133
Table 1
Results of TRPV1 Ca2þ channel assays of piperidinecarboxamides 4e23, 27e32, and piperazinylureas 36e44.a
Compd
HetAr
Linker
n
0
R
Antagonism/desensitization
IC50
M) b
Efficacy at 10
m
M c
Potency EC50
1.1 0.1
(mM)
(m
4
4-F
1.6 0.1
8.8 0.7
5.4 0.1
>10
44.3 2.2
5
0
0
0
0
0
0
0
0
0
0
1
1
1
1
1
2
2
4-Cl
<10
Nd d
6
4-OH
48.0 1.3
<10
3.8 0.5
Nd
7
4-OMe
4-t-Butyl
3-CF3
8
15.7 1.8
6.9 0.1
7.4 0.3
>10
<10
Nd
9
<10
Nd
10
11
12
13
14
15
16
17
18
19
20
21
4-CF3
<10
Nd
4-CH2OH
4-CH(CH3)OH
3,4-(OCH2O)
3,4-(O(CH2)2O)
4-F
<10
Nd
>10
<10
Nd
>10
<10
Nd
>10
<10
Nd
7.0 0.3
9.2 0.1
>10
42.2 2.9
26.6 0.2
<10
4.4 1.2
2.5 0.1
Nd
4-Cl
4-OMe
4-t-Butyl
4-CF3
>10
<10
Nd
8.1 0.4
>10
48.0 0.7
18.0 0.1
24.0 0.5
5.5 2.6
2.1 0.1
5.1 0.2
4-F
4-Cl
>10
(continued on next page)

134
C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
Table 1 (continued )
c
Compd
HetAr
Linker
n
2
R
Antagonism/desensitization
Efficacy at 10
<10
mM
Potency EC50
Nd
(mM)
IC50 (m
M) b
22
4-OH
>10
23
2
4-OMe
>10
18.5 0.1
2.8 0.1
27
28
29
30
31
32
36
37
38
39
40
41
42
43
44
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
4-F
>10
>10
<10
Nd
4-OH
4-F
<10
Nd
7.5 1.5
>10
23.5 1.8
19.7 2.1
61.8 1.1
42.7 0.6
<10
2.1 0.9
3.3 1.6
1.1 0.1
2.8 0.2
Nd
4-OH
4-F
1.1 0.05
5.1 0.3
>10
4-OH
4-F
4-Cl
10.2 0.1
>10
<10
Nd
4-OH
4-t-Butyl
3-CF3
4-CF3
4-F
<10
Nd
0.16 0.01
8.7 0.2
0.72 0.01
2.3 0.1
4.2 0.1
0.88 0.04
<10
Nd
<10
Nd
<10
Nd
40.2 1.0
37.7 0.3
10.6 0.1
2.5 0.3
2.0 0.1
5.0 0.1
4-OH
4-t-Butyl
CPZ
0.105 0.01
BCTC
0.0349 0.019
a
Data are means SEM of N ¼ 3 determinations.
b
c
Determined against the effect of 0.1
As percent of 4 M ionomycin.
Not determined, when efficacy is lower than 10.
mM capsaicin.
m
d
in water. 2 N hydrochloric acid was added to pH 3, the solid formed
was filtered off and dried in vacuo to yield 1-[3-cyano-6-(tri-
fluoromethyl)pyridin-2-yl]piperidine-3-carboxylic acid (3) (2.18 g,
73% yield), which was used without further purification in the next
(m/z): 300 (M þ H)þ.
4.1.3. 1-[4-(Trifluoromethyl)pyrimidin-2-yl]piperidine-3-carboxylic
acid (26)
step. 1HNMR (DMSO-d6)
d 1.61 (m, 2H), 1.94 (m, 2H), 2.57 (m, 1H),
To
a solution of 2-chloro-4-trifluoromethylpyrimidine (45,
3.18 (m, 2H), 4.21 (m, 2H), 7.24 (d, J ¼ 7.8 Hz, 1H), 8.66 (d, J ¼ 7.8 Hz,
1.83 g, 10 mmol) and ethyl piperidine-3-carboxylate (46, 1.90 g,
12 mmol) in DMF (100 ml) potassium carbonate (2.8 g, 20 mmol)
1H), 11.94 (s, 1H). IR (Nujol) 2880, 2208, 1708, 1595 cmꢁ1. ESI-MS

C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
135
was added, and the mixture was stirred overnight at 100 ꢀC. The
reaction mixture was taken up with brine and extracted with
AcOEt. The organic layer was washed with 2 N hydrochloric acid,
saturated NaHCO3, and evaporated in vacuo. The residue was dis-
solved in methanol (100 ml) and 2 N NaOH (25 ml) was added. The
mixture was stirred overnight at room temperature, and after
evaporation of the solvent, the residue was dissolved in water. 2 N
hydrochloric acid was added to pH 3, the solid formed was filtered
off and dried in vacuo to yield 1-[4-(trifluoromethyl)pyrimidin-2-
yl]piperidine-3-carboxylic acid (26) (1.71 g, 62% yield), which was
used in the next step without further purification. 1HNMR (DMSO-
(M þ H)þ. 1HNMR:
d 1.75 (m, 2H),1.93 (m, 2H), 2.65 (m,1H), 3.18 (m,
2H), 3.71 (s, 3H), 4.39 (m, 2H), 6.87 (d, J ¼ 8.4 Hz, 2H), 7.28 (d,
J ¼ 7.5 Hz,1H), 7.51 (d, J ¼ 8.4 Hz, 2H), 8.34 (d, J ¼ 7.5 Hz,1H), 9.80 (s,
1H); IR (Nujol) 3262, 2220, 1640, 1589 cmꢁ1. Anal. Calcd for
C20H19F3N4O2: C, 59.40; H, 4.74; N, 13.85. Found: C, 59.33; H, 4.70;
N, 13.78.
4.1.4.5. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-tert-butyl-
phenyl)piperidine-4-carboxamide (8). Following the general proce-
dure, the title compound was obtained from 3 and 4-tert-
butylaniline in 85% yield; mp 202e204 ꢀC. ESI-MS (m/z): 431
d6)
d
1.43 (m, 1H), 1.79 (m, 2H), 1.99 (m, 1H), 3.08 (m, 1H), 3.21 (m,
(M þ H)þ. 1HNMR:
d 1.23 (s, 9H), 1.78 (m, 2H), 1.91 (m, 2H), 2.66 (m,
2H), 4.50 (m, 2H), 6.98 (d, J ¼ 3.0 Hz, 1H), 8.66 (d, J ¼ 3.0 Hz, 1H),
12.36 (s, 1H). IR (Nujol) 2856, 1713, 1597 cmꢁ1. ESI-MS (m/z): 276
(M þ H)þ.
1H), 3.16 (m, 2H), 4.38 (m, 2H), 7.25 (m, 3H), 7.49 (d, J ¼ 8.5 Hz, 2H),
8.31 (d, J ¼ 7.5 Hz, 1H), 9.87 (s, 1H); IR (Nujol) 3349, 2220, 1652,
1590 cmꢁ1. Anal. Calcd for C23H25F3N4O: C, 64.17; H, 5.85; N, 13.02.
Found: C, 64.09; H, 5.80; N, 12.90.
4.1.4. General procedure for the preparation of 1-
heteroarylpiperidinecarboxamides 4e23, 27e32
4.1.4.6. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(3-
trifluoromethtylphenyl)piperidine-4-carboxamide (9). Following the
general procedure, the title compound was obtained from 3 and 3-
(trifluoromethyl)aniline in 76% yield; mp 178e180 ꢀC. ESI-MS (m/
To a stirred solution of 3, or 24e26 (0.50 mmol) in DCM (5 mL)
were added HOBt (102 mg, 0.60 mmol) and EDC (116 mg,
0.60 mmol). The mixture was stirred for 30 min at room temper-
ature. Then
a
substituted aryl, benzyl, or phenethylamine
z): 443 (M þ H)þ. 1HNMR:
d 1.74 (m, 2H), 1.96 (m, 2H), 2.70 (m, 1H),
(0.60 mmol) was added, and the mixture was stirred 24 h at room
temperature. After evaporation of the solvent, the residue was
taken up with brine and extracted with AcOEt. The organic phase
was washed with 0.5 N hydrochloric acid, saturated NaHCO3, and
brine, dried (Na2SO4), and evaporated under vacuum. The residue
was crystallized with i-Pr2O/MeOH to give piperidinecarboxamides
4e23, 27e32 in 46e92% yields.
3.16 (m, 2H), 4.38 (m, 2H), 7.26 (d, J ¼ 7.8 Hz, 1H), 7.37 (m, 1H), 7.53
(m, 1H), 7.78 (m, 1H), 8.10 (s, 1H), 8.32 (d, J ¼ 7.8 Hz, 1H), 10.30 (s,
1H); IR (Nujol) 3282, 2224, 1662, 1589 cmꢁ1. Anal. Calcd for
C20H16F6N4O: C, 54.30; H, 3.65; N, 12.67. Found: C, 54.22; H, 3.60; N,
12.58.
4.1.4.7. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
trifluoromethtylphenyl)piperidine-4-carboxamide
(10).
4.1.4.1. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
fluorophenyl)piperidine-4-carboxamide (4). Following the general
procedure, the title compound was obtained from 3 and 4-
fluoroaniline in 68% yield; mp 172e174 ꢀC. ESI-MS (m/z): 393
Following the general procedure, the title compound was obtained
from 3 and 4-(trifluoromethyl)aniline in 88% yield; mp 226e228 ꢀC.
ESI-MS (m/z): 443 (M þ H)þ. 1HNMR:
d 1.75 (m, 2H), 1.95 (m, 2H),
2.72 (m, 1H), 3.20 (m, 2H), 4.38 (m, 2H), 7.27 (d, J ¼ 7.6 Hz, 1H), 7.65
(d, J ¼ 8.4 Hz, 2H), 7.80 (d, J ¼ 8.4 Hz, 2H), 8.32 (d, J ¼ 7.6 Hz, 1H),
10.33 (s, 1H); IR (Nujol) 3281, 2221, 1664, 1592 cmꢁ1. Anal. Calcd for
(M þ H)þ. 1HNMR:
d 1.74 (m, 2H),1.92 (m, 2H), 2.65 (m,1H), 3.15 (m,
2H), 4.37 (m, 2H), 7.10 (m, 2H), 7.27 (d, J ¼ 7.5 Hz, 1H), 7.60 (m, 2H),
8.32 (d, J ¼ 7.5 Hz, 1H), 10.03 (s, 1H); IR (Nujol) 3272, 2220,1661,
1590 cmꢁ1. Anal. Calcd for C19H16F4N4O: C, 58.16; H, 4.11; N, 14.28.
Found: C, 58.08; H, 4.06; N, 14.21.
C20H16F6N4O: C, 54.30; H, 3.65; N, 12.67. Found: C, 54.24; H, 3.59; N,
12.60.
4.1.4.8. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-(hydrox-
ymethyl)phenyl)piperidine-4-carboxamide (11). Following the gen-
eral procedure, the title compound was obtained from 3 and 4-
(hydroxymethyl)aniline in 54% yield; mp 155e157 ꢀC. ESI-MS (m/z):
4.1.4.2. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
chlorophenyl)piperidine-4-carboxamide (5). Following the general
procedure, the title compound was obtained from 3 and 4-
chloroaniline in 92% yield; mp 197e199 ꢀC. ESI-MS (m/z): 410
405 (M þ H)þ. 1HNMR:
d 1.74 (m, 2H), 1.93 (m, 2H), 2.67 (m, 1H),
(M þ H)þ. 1HNMR:
d
1.74 (m, 2H),1.93 (m, 2H), 2.67 (m,1H), 3.19 (m,
3.14 (m, 2H), 4.40 (m, 4H), 5.06 (s, 1H), 7.22 (d, J ¼ 6.0 Hz, 2H), 7.27
(d, J ¼ 7.2 Hz, 1H), 7.54 (d, J ¼ 6.0 Hz, 2H), 8.35 (d, J ¼ 7.2 Hz, 1H),
9.90 (s, 1H); IR (Nujol) 3340, 3296, 2221, 1659, 1591 cmꢁ1. Anal.
Calcd for C20H19F3N4O2: C, 59.40; H, 4.74; N, 13.85. Found: C, 59.32;
H, 4.71; N, 13.76.
2H), 4.38 (m, 2H), 7.27 (d, J ¼ 7.8 Hz,1H), 7.33 (d, J ¼ 8.4 Hz, 2H), 7.63
(d, J ¼ 8.4 Hz, 2H), 8.33 (d, J ¼ 7.8 Hz, 1H), 10.04 (s, 1H); IR (Nujol)
3279, 2218,1660, 1592 cmꢁ1. Anal. Calcd for C19H16ClF3N4O: C,
55.82; H, 3.94; N, 13.71. Found: C, 55.73; H, 3.90; N, 13.68.
4.1.4.3. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
hydroxyphenyl)piperidine-4-carboxamide (6). Following the general
procedure, the title compound was obtained from 3 and 4-
hydroxyaniline in 50% yield; mp 218e220 ꢀC. ESI-MS (m/z): 391
4.1.4.9. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-(1-
hydroxyethyl)phenyl)piperidine-4-carboxamide (12). Following the
general procedure, the title compound was obtained from 3 and 4-
(1-hydroxyethyl)aniline in 46% yield; mp 112e114 ꢀC. ESI-MS (m/z):
(M þ H)þ. 1HNMR:
d
1.72 (m, 2H),1.88 (m, 2H), 2.58 (m,1H), 3.14 (m,
419 (M þ H)þ. 1HNMR:
d
1.25 (d, J ¼ 6.0 Hz, 3H), 1.69 (m, 2H), 1.90
2H), 4.37 (m, 2H), 6.66 (d, J ¼ 8.0 Hz, 2H), 7.25 (d, J ¼ 7.6 Hz, 1H),
7.34 (d, J ¼ 8.0 Hz, 2H), 8.29 (d, J ¼ 7.6 Hz, 1H), 9.24 (s, 1H), 9.69 (s,
1H); IR (Nujol) 3311, 2225, 1660, 1592 cmꢁ1. Anal. Calcd for
(m, 2H), 2.65 (m, 1H), 3.15 (m, 2H), 4.37 (m, 2H), 4.64 (m, 1H), 5.12
(m, 1H), 7.24 (m, 3H), 7.50 (d, J ¼ 8.0 Hz, 2H), 8.29 (d, J ¼ 7.8 Hz, 1H),
9.96 (s, 1H); IR (Nujol) 3338, 3292, 2220, 1660, 1592 cmꢁ1. Anal.
Calcd for C20H19F3N4O2: C, 60.28; H, 5.06; N, 13.39. Found: C, 60.20;
H, 5.00; N, 13.31.
C
19H17F3N4O2: C, 58.46; H, 4.39; N, 14.35. Found: C, 58.40; H, 4.35;
N, 14.28.
4.1.4.4. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
methoxyphenyl)piperidine-4-carboxamide (7). Following the gen-
eral procedure, the title compound was obtained from 3 and 4-
methoxyaniline in 74% yield; mp 193e195 ꢀC. ESI-MS (m/z): 405
4.1.4.10. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(3,4-
methylenedioxyphenyl)piperidine-4-carboxamide
(13).
Following the general procedure, the title compound was obtained
from 3 and 3,4-methylendioxyaniline in 74% yield; mp 196e198 ꢀC.

136
C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
ESI-MS (m/z): 419 (M þ H)þ. 1HNMR:
d
1.73 (m, 2H), 1.92 (m, 2H),
1H), 3.21 (m, 2H), 4.34 (m, 4H), 7.22 (d, J ¼ 7.5 Hz, 1H), 7.43 (d,
2.63 (m, 1H), 3.16 (m, 2H), 4.38 (m, 2H), 5.96 (s, 2H), 6.83 (d,
J ¼ 7.8 Hz, 1H), 6.97 (d, J ¼ 7.8 Hz, 1H), 7.27 (d, J ¼ 7.2 Hz, 1H), 7.30 (s,
1H), 8.33 (d, J ¼ 7.2 Hz, 1H), 9.85 (s, 1H); IR (Nujol) 3261, 2222, 1648,
1592 cmꢁ1. Anal. Calcd for C20H17F3N4O3: C, 57.42; H, 4.10; N, 13.39.
Found: C, 57.35; H, 4.06; N, 13.31.
J ¼ 8.0 Hz, 2H), 7.64 (d, J ¼ 8.0 Hz, 2H), 8.27 (d, J ¼ 7.5 Hz, 1H), 8.50
(m, 1H); IR (Nujol) 3286, 2218, 1646, 1588 cmꢁ1. Anal. Calcd for
C21H18F6N4O: C, 55.27; H, 3.98; N, 12.28. Found: C, 55.20; H, 3.94; N,
12.21.
4.1.4.17. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
fluorophenethyl)piperidine-4-carboxamide (20). Following the gen-
eral procedure, the title compound was obtained from 3 and 4-
fluorophenethylamine in 68% yield; mp 163e165 ꢀC. ESI-MS (m/
4.1.4.11. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(3,4-
ethylenedioxyphenyl)piperidine-4-carboxamide (14). Following the
general procedure, the title compound was obtained from 3 and
3,4-ethylendioxyaniline in 86% yield; mp 203e205 ꢀC. ESI-MS (m/
z): 421 (M þ H)þ. 1HNMR:
d 1.63 (m, 2H), 1.76 (m, 2H), 2.41 (m, 1H),
z): 433 (M þ H)þ. 1HNMR:
d
1.76 (m, 2H), 1.91 (m, 2H), 2.61 (m, 1H),
2.70 (m, 2H), 3.11 (m, 2H), 3.26 (m, 2H), 4.31 (m, 2H), 7.08 (m, 2H),
7.24 (m, 3H), 7.89 (br s, 1H), 8.32 (d, J ¼ 7.0 Hz, 1H); IR (Nujol) 3304,
2224, 1641, 1589 cmꢁ1. Anal. Calcd for C21H20F4N4O: C, 60.00; H,
4.80; N, 13.33. Found: C, 60.02; H, 4.74; N, 13.26.
3.16 (m, 2H), 4.19 (m, 4H), 4.38 (m, 2H), 6.75 (d, J ¼ 8.5 Hz, 1H), 6.98
(d, J ¼ 8.5 Hz, 1H), 7.23 (s, 1H), 7.27 (d, J ¼ 7.5 Hz, 1H), 8.33 (d,
J ¼ 7.5 Hz, 1H), 9.76 (s, 1H); IR (Nujol) 3264, 2221, 1645, 1594 cmꢁ1
.
Anal. Calcd for C21H19F3N4O3: C, 58.33; H, 4.43; N, 12.96. Found: C,
58.30; H, 4.38; N, 12.88.
4.1.4.18. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
chlorophenethyl)piperidine-4-carboxamide (21). Following the
general procedure, the title compound was obtained from 3 and 4-
chlorophenethylamine in 70% yield; mp 171e173 ꢀC. ESI-MS (m/z):
4.1.4.12. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
fluorobenzyl)piperidine-4-carboxamide (15). Following the general
procedure, the title compound was obtained from 3 and 4-
fluorobenzylamine in 56% yield; mp 170e172 ꢀC. ESI-MS (m/z):
438 (M þ H)þ. 1HNMR:
d 1.60 (m, 2H), 1.75 (m, 2H), 2.40 (m, 1H),
2.69 (m, 2H), 3.09 (m, 2H), 3.25 (m, 2H), 4.30 (m, 2H), 7.20 (d,
J ¼ 8.0 Hz, 2H), 7.24 (d, J ¼ 7.0 Hz, 1H), 7.30 (d, J ¼ 8.0 Hz, 2H), 7.90
(m, 1H), 8.29 (d, J ¼ 7.0 Hz, 1H); IR (Nujol) 3299, 2223, 1640,
1589 cmꢁ1. Anal. Calcd for C21H20ClF3N4O: C, 57.74; H, 4.61; N,
12.82. Found: C, 57.69; H, 4.59; N, 12.74.
407 (M þ H)þ. 1HNMR:
d 1.68 (m, 2H), 1.85 (m, 2H), 2.54 (m, 1H),
3.12 (m, 2H), 4.24 (d, J ¼ 6.0 Hz, 2H), 4.34 (m, 2H), 7.21 (m, 2H), 7.25
(m, 3H), 8.29 (d, J ¼ 7.6 Hz,1H), 8.39 (br s,1H); IR (Nujol) 3290, 2221,
1638, 1589 cmꢁ1. Anal. Calcd for C20H18F4N4O: C, 59.11; H, 4.46; N,
13.79. Found: C, 59.05; H, 4.41; N, 13.70.
4.1.4.19. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
hydroxyphenethyl)piperidine-4-carboxamide (22). Following the
general procedure, the title compound was obtained from 3 and 4-
hydroxyphenethylamine in 48% yield; mp 134e136 ꢀC. ESI-MS (m/
4.1.4.13. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
chlorobenzyl)piperidine-4-carboxamide (16). Following the general
procedure, the title compound was obtained from 3 and 4-
chlorobenzylamine in 90% yield; mp 179e181 ꢀC. ESI-MS (m/z):
z): 419 (M þ H)þ. 1HNMR:
d 1.61 (m, 2H), 1.76 (m, 2H), 2.40 (m, 1H),
424 (M þ H)þ. 1HNMR:
d
1.67 (m, 2H), 1.85 (m, 2H), 2.52 (m, 1H),
2.56 (m, 2H), 3.08 (m, 2H), 3.18 (m, 2H), 4.30 (m, 2H), 6.65 (d,
J ¼ 8.0 Hz, 2H), 6.96 (d, J ¼ 8.0 Hz, 2H), 7.23 (d, J ¼ 7.5 Hz, 1H), 7.86
(m, 1H), 8.28 (d, J ¼ 7.5 Hz, 1H), 9.19 (s, 1H); IR (Nujol) 3321, 2221,
1641, 1588 cmꢁ1. Anal. Calcd for C21H21F3N4O2: C, 60.28; H, 5.06; N,
13.39. Found: C, 60.20; H, 5.01; N, 13.30.
3.12 (m, 2H), 4.24 (d, J ¼ 6.0 Hz, 2H), 4.33 (m, 2H), 7.24 (m, 3H), 7.35
(d, J ¼ 8.0 Hz, 2H), 8.30 (d, J ¼ 7.8 Hz, 1H), 8.41 (m, 1H); IR (Nujol)
3279, 2219, 1643, 1587 cmꢁ1. Anal. Calcd for C20H18ClF3N4O: C,
56.81; H, 4.29; N, 13.25. Found: C, 56.79; H, 4.25; N, 13.20.
4.1.4.14. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
methoxybenzyl)piperidine-4-carboxamide (17). Following the gen-
eral procedure, the title compound was obtained from 3 and 4-
methoxybenzylamine in 84% yield; mp 171e173 ꢀC. ESI-MS (m/z):
4.1.4.20. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-
methoxyphenethyl)piperidine-4-carboxamide (23). Following the
general procedure, the title compound was obtained from 3 and 4-
methoxyphenethylamine in 65% yield; mp 164e166 ꢀC. ESI-MS (m/
419 (M þ H)þ. 1HNMR:
d
1.69 (m, 2H), 1.85 (m, 2H), 2.54 (m, 1H),
z): 433 (M þ H)þ. 1HNMR:
d 1.64 (m, 2H), 1.77 (m, 2H), 2.40 (m, 1H),
3.13 (m, 2H), 3.73 (s, 3H), 4.20 (d, J ¼ 5.5 Hz, 2H), 4.35 (m, 2H), 6.88
(d, J ¼ 8.5 Hz, 2H), 7.16 (d, J ¼ 8.5 Hz, 2H), 7.26 (d, J ¼ 7.5 Hz, 1H),
8.34 (m, 2H); IR (Nujol) 3286, 2220,1636, 1588 cmꢁ1. Anal. Calcd for
2.64 (m, 2H), 3.11 (m, 2H), 3.23 (m, 2H), 3.71 (s, 3H), 4.31 (m, 2H),
6.84 (d, J ¼ 8.8 Hz, 2H), 7.10 (d, J ¼ 8.8 Hz, 2H), 7.25 (d, J ¼ 7.6 Hz,
1H), 7.87 (m, 1H), 8.32 (d, J ¼ 7.6 Hz, 1H); IR (Nujol) 3296, 2224,
1639, 1591 cmꢁ1. Anal. Calcd for C22H23F3N4O2: C, 61.10; H, 5.36; N,
12.96. Found: C, 61.04; H, 5.30; N, 12.87.
C
21H21F3N4O2: C, 60.28; H, 5.06; N, 13.39. Found: C, 60.20; H, 4.98;
N, 13.32.
4.1.4.15. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-tert-
butylbenzyl)piperidine-4-carboxamide (18). Following the general
procedure, the title compound was obtained from 3 and 4-tert-
butylbenzylamine in 78% yield; mp 138e140 ꢀC. ESI-MS (m/z): 445
4.1.4.21. 1-(3-cyanopyridin-2-yl)-N-(4-fluorophenyl)piperidine-4-
carboxamide (27). Following the general procedure, the title com-
pound was obtained from 24 and 4-fluoroaniline in 84% yield; mp
204e206 ꢀC. ESI-MS (m/z): 325 (M þ H)þ. 1HNMR:
d 1.74 (m, 2H),
(M þ H)þ. 1HNMR:
d
1.25 (s, 9H), 1.70 (m, 2H), 1.85 (m, 2H), 2.52 (m,
1.87 (m, 2H), 2.58 (m, 1H), 3.06 (m, 2H), 4.28 (m, 2H), 6.67 (d,
1H), 3.12 (m, 2H), 4.22 (d, J ¼ 5.5 Hz, 2H), 4.34 (m, 2H), 7.15 (d,
J ¼ 8.5 Hz, 2H), 7.25 (d, J ¼ 7.6 Hz, 1H), 7.32 (d, J ¼ 8.5 Hz, 2H), 8.32
(m, 2H); IR (Nujol) 3266, 2220, 1637, 1588 cmꢁ1. Anal. Calcd for
J ¼ 8.8 Hz, 2H), 6.90 (m, 1H), 7.37 (d, J ¼ 8.8 Hz, 2H), 8.05 (m, 1H),
8.40 (m, 1H), 9.65 (s, 1H); IR (Nujol) 3305, 2223,1660, 1585 cmꢁ1
.
Anal. Calcd for C18H17FN4O: C, 66.65; H, 5.28; N, 17.27. Found: C,
66.59; H, 5.22; N, 17.20.
C
24H27F3N4O: C, 64.85; H, 6.12; N, 12.60. Found: C, 64.78; H, 6.09; N,
12.52.
4.1.4.22. 1-(3-cyanopyridin-2-yl)-N-(4-hydroxyphenyl)piperidine-4-
carboxamide (28). Following the general procedure, the title com-
pound was obtained from 24 and 4-hydroxyaniline in 64% yield;
4.1.4.16. 1-(3-cyano-6-(trifluoromethyl)pyridin-2-yl)-N-(4-(tri-
fluoromethyl)benzyl)piperidine-4-carboxamide (19). Following the
general procedure, the title compound was obtained from 3 and 4-
(trifluoromethyl)benzylamine in 62% yield; mp 189e191 ꢀC. ESI-MS
mp 225e227 ꢀC. ESI-MS (m/z): 323 (M þ H)þ. 1HNMR:
d 1.74 (m,
2H), 1.90 (m, 2H), 2.61 (m, 1H), 3.09 (m, 2H), 4.28 (m, 2H), 6.90 (m,
1H), 7.11 (d, J ¼ 8.5 Hz, 2H), 7.62 (d, J ¼ 8.5 Hz, 2H), 8.04 (m,1H), 8.40
(m/z): 457 (M þ H)þ. 1HNMR:
d
1.67 (m, 2H), 1.86 (m, 2H), 2.54 (m,

C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
137
(m, 1H), 9.99 (s, 1H), 10.52 (s, 1H); IR (Nujol) 3427, 3299, 2220,1652,
1585 cmꢁ1. Anal. Calcd for C18H18N4O2: C, 66.65; H, 5.28; N, 17.27.
Found: C, 66.59; H, 5.22; N, 17.20.
4.1.6.1. 4-(3-cyanopyridin-2-yl)-N-(4-fluorophenyl)piperazine-1-
carboxamide (36). Following the general procedure, the title com-
pound was obtained from 33 and 35a in 72% yield; mp 189e191 ꢀC.
ESI-MS (m/z): 326 (M þ H)þ. 1HNMR:
d 3.61 (m, 4H), 3.68 (m, 4H),
6.95 (m, 1H), 7.08 (m, 2H), 7.48 (m, 2H), 8.09 (m, 1H), 8.44 (m, 1H),
4.1.4.23. N-(4-fluorophenyl)-1-(4-(trifluoromethyl)pyrimidin-2-yl)
piperidine-4-carboxamide (29). Following the general procedure,
the title compound was obtained from 25 and 4-fluoroaniline in
89% yield; mp 191e193 ꢀC. ESI-MS (m/z): 369 (M þ H)þ. 1HNMR:
8.61 (s, 1H); IR (Nujol) 3338, 2222, 1637, 1584 cmꢁ1. Anal. Calcd for
C17H16FN5O: C, 62.76; H, 4.96; N, 21.53. Found: C, 62.80; H, 4.90; N,
21.47.
d
1.60 (m, 2H), 1.90 (m, 2H), 2.66 (m, 1H), 3.04 (m, 2H), 4.68 (m, 2H),
4.1.6.2. 4-(3-Cyanopyridin-2-yl)-N-(4-chlorophenyl)piperazine-1-
carboxamide (37). Following the general procedure, the title com-
pound was obtained from 33 and 35b in 81% yield; mp 178e180 ꢀC.
6.99 (d, J ¼ 5.0 Hz, 1H), 7.12 (m, 2H), 7.61 (m, 2H), 8.68 (d, J ¼ 5.0 Hz,
1H), 9.98 (s, 1H); IR (Nujol) 3277, 1652, 1595 cmꢁ1. Anal. Calcd for
C
17H16F4N4O: C, 55.43; H, 4.38; N, 15.21. Found: C, 55.36; H, 4.35; N,
ESI-MS (m/z): 343 (M þ H)þ. 1HNMR:
d 3.62 (m, 4H), 3.67 (m, 4H),
15.15.
6.95 (m, 1H), 7.28 (d, J ¼ 8.4 Hz, 2H), 7.52 (d, J ¼ 8.4 Hz, 2H), 8.10 (m,
1H), 8.44 (m, 1H), 8.70 (s, 1H); IR (Nujol) 3347, 2221, 1638,
1583 cmꢁ1. Anal. Calcd for C17H16ClN5O: C, 59.74; H, 4.72; N, 20.49.
Found: C, 59.69; H, 4.68; N, 20.42.
4.1.4.24. N-(4-hydroxyphenyl)-1-(4-(trifluoromethyl)pyrimidin-2-yl)
piperidine-4-carboxamide (30). Following the general procedure,
the title compound was obtained from 25 and 4-hydroxþyaniline in
52% yield; mp 232e234 ꢀC. ESI-MS (m/z): 367 (M þ H) . 1HNMR:
4.1.6.3. 4-(3-cyanopyridin-2-yl)-N-(4-hydroxyphenyl)piperazine-1-
carboxamide (38). Following the general procedure, the title com-
pound was obtained from 33 and 35c in 66% yield; mp 203e205 ꢀC.
d
1.57 (m, 2H), 1.87 (m, 2H), 2.61 (m, 1H), 3.04 (m, 2H), 4.68 (m, 2H),
6.67 (d, J ¼ 8.4 Hz, 2H), 6.99 (d, J ¼ 4.8 Hz, 1H), 7.36 (d, J ¼ 8.4 Hz,
2H), 8.68 (d, J ¼ 4.8 Hz,1H), 9.13 (s,1H), 9.66 (s, 1H); IR (Nujol) 3296,
1653, 1595 cmꢁ1. Anal. Calcd for C19H17F3N4O2: C, 55.74; H, 4.68; N,
15.29. Found: C, 55.67; H, 4.64; N, 15.20.
ESI-MS (m/z): 324 (M þ H)þ. 1HNMR:
d 3.58 (m, 4H), 3.65 (m, 4H),
6.65 (d, J ¼ 8.6 Hz, 2H), 6.95 (m, 1H), 7.20 (d, J ¼ 8.6 Hz, 2H), 8.09 (m,
1H), 8.30 (s, 1H), 8.44 (m, 1H), 9.00 (s, 1H); IR (Nujol) 3365, 3146,
2220,1640, 1582 cmꢁ1. Anal. Calcd for C17H17N5O2: C, 63.15; H, 5.30;
N, 21.66. Found: C, 63.08; H, 5.24; N, 21.60.
4.1.4.25. N-(4-fluorophenyl)-1-(4-(trifluoromethyl)pyrimidin-2-yl)
piperidine-3-carboxamide (31). Following the general procedure,
the title compound was obtained from 26 and 4-fluoroaniline in
60% yield; mp 202e204 ꢀC. ESI-MS (m/z): 369 (M þ H)þ. 1HNMR:
4.1.6.4. 4-(3-cyanopyridin-2-yl)-N-(4-t-butylphenyl)piperazine-1-
carboxamide (39). Following the general procedure, the title com-
pound was obtained from 33 and 35d in 82% yield; mp 135e137 ꢀC.
d
1.47 (m, 1H),1.75 (m, 2H), 2.03 (m, 1H), 3.02 (m, 1H), 3.17 (m, 2H),
4.64 (m, 2H), 6.99 (d, J ¼ 4.8 Hz, 1H), 7.13 (m, 2H), 7.61 (m, 2H), 8.68
ESI-MS (m/z): 364 (M þ H)þ. 1HNMR:
d 1.24 (s, 9H), 3.60 (m, 4H),
(d, J ¼ 4.8 Hz, 1H), 10.04 (s, 1H); IR (Nujol) 3287, 1658, 1593 cmꢁ1
.
3.66 (m, 4H), 6.94 (m, 1H), 7.24 (d, J ¼ 8.4 Hz, 2H), 7.36 (d, J ¼ 8.4 Hz,
2H), 8.08 (m, 1H), 8.43 (m, 1H), 8.48 (s, 1H); IR (Nujol) 3357, 2214,
1633, 1584 cmꢁ1. Anal. Calcd for C21H25N5O: C, 69.40; H, 6.93; N,
19.27. Found: C, 69.34; H, 6.88; N, 19.19.
Anal. Calcd for C17H16F4N4O: C, 55.43; H, 4.38; N, 15.21. Found: C,
55.35; H, 4.32; N, 15.17.
4.1.4.26. N-(4-hydroxyphenyl)-1-(4-(trifluoromethyl)pyrimidin-2-yl)
piperidine-3-carboxamide (32). Following the general procedure,
the title compound was obtained from 26 and 4-hydroxþyaniline in
56% yield; mp 222e224 ꢀC. ESI-MS (m/z): 367 (M þ H) . 1HNMR:
4.1.6.5. 4-(3-cyanopyridin-2-yl)-N-(3-(trifluoromethyl)phenyl)piper-
azine-1-carboxamide (40). Following the general procedure, the
title compound was obtained from 33 and 35e in 70% yield; mp
116e118 ꢀC. ESI-MS (m/z): 376 (M þ H)þ. 1HNMR:
d 3.66 (m, 8H),
d
1.44 (m, 1H),1.78 (m, 2H), 1.99 (m, 1H), 2.99 (m, 1H), 3.13 (m, 2H),
6.95 (m, 1H), 7.27 (d, J ¼ 7.6 Hz, 1H), 7.47 (m, 1H), 7.77 (m, 1H), 7.95
(s, 1H), 8.09 (d, J ¼ 7.6 Hz, 1H), 8.43 (m, 1H), 8.92 (s, 1H); IR (Nujol)
3309, 2216, 1665, 1584 cmꢁ1. Anal. Calcd for C20H16F6N4O: C, 54.30;
H, 3.65; N, 12.67. Found: C, 54.23; H, 3.58; N, 12.60.
4.64 (m, 2H),6.68 (d, J ¼ 8.0 Hz, 2H), 6.98 (d, J ¼ 4.0 Hz, 1H), 7.35 (d,
J ¼ 8.0 Hz, 2H), 8.67 (d, J ¼ 4.0 Hz, 1H), 9.15 (s, 1H), 9.70 (s, 1H); IR
(Nujol) 3297, 3208, 1644, 1597 cmꢁ1. Anal. Calcd for C19H17F3N4O2:
C, 55.74; H, 4.68; N, 15.29. Found: C, 55.69; H, 4.63; N, 15.22.
4.1.6.6. 4-(3-cyanopyridin-2-yl)-N-(4-(trifluoromethyl)phenyl)piper-
azine-1-carboxamide (41). Following the general procedure, the
title compound was obtained from 33 þand 35f in 93% yield; mp
4.1.5. General procedure for the preparation of phenyl N-
arylcarbamates 35aef
174e176 ꢀC. ESI-MS (m/z): 376 (M þ H) . 1HNMR:
d 3.66 (m, 8H),
To a stirred solution of a substituted aniline 47aef (1 mmol) and
DIPEA (0.194 g, 1.5 mmol) in anhydrous THF (10 mL) at 0 ꢀC phenyl
chloroformate (0.189 g, 1.2 mmol) in THF (2.5 mL) was added
dropwise. The mixture was stirred at room temperature for 4 h.
After evaporation of the solvent, water (10 mL) was added; the
formed solid was filtered off, washed with hexane and dried in
vacuo to give 35aef, which were used without further purification
in the next step.
6.96 (m, 1H), 7.59 (d, J ¼ 8.5 Hz, 2H), 7.71 (d, J ¼ 8.5 Hz, 2H), 8.10 (m,
1H), 8.44 (m, 1H), 8.97 (s, 1H); IR (Nujol) 3361, 2218, 1668,
1585 cmꢁ1. Anal. Calcd for C18H16F3N5O: C, 57.60; H, 4.30; N, 18.66.
Found: C, 57.53; H, 4.28; N, 18.58.
4.1.6.7. N-(4-fluorophenyl)-4-(4-(trifluoromethyl)pyrimidin-2-yl)
piperazine-1-carboxamide (42). Following the general procedure,
the title compound was obtained from 34 and 35a in 64% yield; mp
4.1.6. General procedure for the preparation of 4-
196e198 ꢀC. ESI-MS (m/z): 370 (M þ H)þ. 1HNMR:
d 3.57 (m, 4H),
heteroarylpiperazine-1-carboxamide derivatives 36e44
A mixture of 1-substituted piperazine 33 or 34 (0.5 mmol) and
phenyl arylcarbamate 35 (0.5 mmol) in anhydrous DMSO (5 mL)
was stirred at room temperature for 18 h. The reaction mixture was
then diluted with water (10 mL); the formed solid was collected
with water, filtered off, air dried, and crystallized with MeOH to
give 36e44 in 64e93% yields.
3.82 (m, 4H), 7.08 (m, 3H), 7.47 (m, 2H), 8.65 (s, 1H), 8.72 (d,
J ¼ 4.5 Hz, 1H); IR (Nujol) 3278, 1632, 1592 cmꢁ1. Anal. Calcd for
C16H15F4N5O: C, 52.03; H, 4.09; N, 18.96. Found: C, 51.96; H, 4.06; N,
18.88.
4.1.6.8. N-(4-hydroxyphenyl)-4-(4-(trifluoromethyl)pyrimidin-2-yl)
piperazine-1-carboxamide (43). Following the general procedure,

138
C. Congiu et al. / European Journal of Medicinal Chemistry 100 (2015) 129e138
[12] (a) M. Pal, S. Angaru, A. Kodimuthali, N. Dhingra, Curr. Pharm. Des. 15 (2009)
1008;
the title compound was obtained from 34 and 35c in 83% yield; mp
218e220 ꢀC. ESI-MS (m/z): 368 (M þ H)þ. 1HNMR:
d 3.54 (m, 4H),
(b) L.M. Broad, S.J. Keding, M.J. Blanco, Curr. Top. Med. Chem. 8 (2008) 1431;
(c) P.R. Kym, M.E. Kort, C.W. Hutchins, Biochem. Pharmacol. 78 (2009) 211;
(d) A. Szallasi, P.M. Blumberg, Pharmacol. Rev. 51 (1999) 159;
(e) A. Szallasi, G. Appendino, J. Med. Chem. 47 (2004) 2717;
(f) R.L. McLeod, C.C. Correll, Y. Jia, J.C. Anthes, Lung 186 (2008) S59;
(g) M.J. Gunthorpe, B.A. Chizh, Drug Discov. Today 14 (2009) 56;
(h) G.Y. Wong, N.R. Gavva, Brain Res. Rev. 60 (2009) 267.
[13] L. De Petrocellis, A. Schiano-Moriello, Recent Pat. CNS Drug Discov. 8 (2013)
180.
3.81 (m, 4H), 6.65 (d, J ¼ 8.0 Hz, 2H), 7.05 (d, J ¼ 5.0 Hz, 1H), 7.20 (d,
J ¼ 8.0 Hz, 2H), 8.33 (s, 1H), 8.71 (d, J ¼ 5.0 Hz, 1H), 9.00 (s, 1H); IR
(Nujol) 3407, 3276, 1634, 1595 cmꢁ1. Anal. Calcd for C16H16F3N5O2:
C, 52.32; H, 4.39; N, 19.07. Found: C, 52.27; H, 4.34; N, 19.00.
4.1.6.9. N-(4-t-butylphenyl)-4-(4-(trifluoromethyl)pyrimidin-2-yl)
piperazine-1-carboxamide (44). Following the general procedure,
the title compound was obtained from 34þand 35d in 69% yield; mp
€
€
€
[14] (a) V. Onnis, C. Congiu, E. Bjorklund, F. Hempel, E. Soderstrom, C.J. Fowler,
J. Med. Chem. 53 (2010) 2286;
163e165 ꢀC. ESI-MS (m/z): 408 (M þ H) . 1HNMR:
d 1.26 (s, 9H),
€
(b) C.J. Fowler, E. Bjorklund, A.H. Lichtman, P.S. Naidu, C. Congiu, V. Onnis,
J. Enzym. Inhib. Med. Chem. 28 (2013) 172;
3.56 (m, 4H), 3.82 (m, 4H), 7.06 (d, J ¼ 4.5 Hz, 1H), 7.26 (d, J ¼ 8.5 Hz,
2H), 7.37 (d, J ¼ 8.5 Hz, 2H), 8.54 (s, 1H), 8.77 (d, J ¼ 4.5 Hz, 1H); IR
(Nujol) 3267, 1645, 1593 cmꢁ1. Anal. Calcd for C20H24F3N5O2: C,
58.96; H, 5.94; N, 17.19. Found: C, 58.87; H, 5.89; N, 17.15.
€
(c) M. Cipriano, E. Bjorklund, A.A. Wilson, C. Congiu, V. Onnis, C.J. Fowler, Eur.
J. Pharmacol. 720 (2013) 383.
[15] (a) Q. Sun, L. Tafesse, K. Islam, X. Zhou, S. Victory, C. Zhang, M. Hachicha,
L. Schmid, A. Patel, Y. Rotshteyn, K.J. Valenzano, D.J. Kyle, Bioorg. Med. Chem.
Lett. 13 (2003) 3611;
(b) K.J.Valenzano, E. R.Grant, G.Wu, M.Hachicha, L.Schmid, L.Tafesse, Q.Sun,
Y.Rotshteyn, J.Francis, J.Limberis, S.Malik, E.R.Whittemore, D.J.Hodges, Phar-
macol. Exp. Ther.. 306 (2003) 377. (c) J.D. Pomonis, J.E. Harrison, L. Mark,
D.R. Bristol, K.J. Valenzano, K.J. Walker, Pharmacol. Exp. Ther. 306 (2003) 387.
[16] D.M. Swanson, A.E. Dubin, C. Shah, N. Nasser, L. Chang, S.L. Dax, M. Jetter,
J.G. Breitenbucher, C. Liu, C. Mazur, B. Lord, L. Gonzales, K. Hoey, M. Rizzolio,
M. Bogenstaetter, E.E. Codd, D.H. Lee, S.-P. Zhang, S.R. Chaplan, N.I. Carruthers,
J. Med. Chem. 48 (2005) 1857.
[17] M.T. Cocco, C. Congiu, V. Onnis, A. Maccioni, Synthesis 7 (1991) 529.
[18] T. Nakagawa, T. Suzuki, K. Takenaka, S. Fujita, Y. Yamada, Y. Shima, T. Oku-
zumi, T. Yoshimura, M. Yoshida, M. Murata, Patent US2009/0258900 A1, 2009.
[19] C.M.S. Heeney, Patent WO2010/097576 A1, 2010.
Acknowledgments
This study was supported in part by the Italian Ministero del-
l’Istruzione, Universita e della Ricerca (PRIN 2010e2011, Prot. no.
120105YY2HL_002), the Research Funds of the University of
Cagliari (to C.C., V.O., G.B.), and Regione Autonoma Sardegna
(project L.R. 7/2007, CRP-59473/2012, to C.C., V.O., G.B., V.D.M.).
ꢀ
References
[20] K. Ishizumi, A. Kojima, F. Antoku, Chem. Pharm. Bull. 39 (1991) 2288.
[21] C.-B. Xue, C. Zheng, H. Feng, M. Xia, J. Glenn, G. Cao, B.W. Metcalf, Patent
WO2006/004741 A2, 2006.
[1] M.J. Caterina, D. Julius, Annu. Rev. Neurosci. 24 (2001) 487.
[2] M. Tominaga, M.J. Caterina, A.B. Malmberg, T.A. Rosen, H. Gilbert, K. Skinner,
B.E. Raumann, A.I. Basbaum, D. Julius, Neuron 21 (1998) 531.
[22] (a) C. Sheng, X. Che, W. Wang, S. Wang, Y. Cao, Z. Miao, J. Yao, W. Zhang, Eur. J.
Med. Chem. 46 (2011) 5276;
[3] (a) D. Smart, M.J. Gunthorpe, J.C. Jerman, S. Nasir, J. Gray, A.I. Muir,
J.K. Chambers, A.D. Randall, J.B. Davis, Br. J. Pharmacol. 129 (2000) 227;
(b) C.H. Mack, H.H. McGregor Jr., S.R. Hobart, J. Chem. Eng. Data 14 (1969) 258;
(c) P.F. Cirillo, S. Breitfelder, U. R. Patel, J. Proudfoot, A. Swinamer, Patent
US2003/0225077 A1, 2003.
(d) L.A. Gharat, L. Narayana, N. Khairatkar-Joshi, V.G. Kattige, Patent WO2009/
090548 A2, 2009.
ꢀ
(b) L. De Petrocellis, T. Bisogno, M. Maccarrone, J.B. Davis, A. Finazzi-Agro, V. Di
Marzo, J. Biol. Chem. 276 (2001) 12856.
[4] (a) P.M. Zygmunt, J. Petersson, D.A. Andersson, H.-h. Chuang, M. Sørgård, V. Di
€
€
Marzo, D. David Julius, E.D. Hogestatt, Nature 400 (1999) 452;
(b) S.W. Hwang, H. Cho, J. Kwak, S.Y. Lee, C.-J. Kang, J. Jung, S. Cho, K.H. Min,
Y.G. Suh, D. Kim, U. Oh, Proc. Natl. Acad. Sci. U. S. A. 97 (2000) 6155.
[5] P. Holzer, Pharmacol. Rev. 43 (1991) 143.
[23] (a) L. De Petrocellis, T. Bisogno, J.B. Davis, R.G. Pertweed, V. Di Marzo, FEBS
Lett. 483 (2000) 52;
(b) G. Ortar, M.G. Cascio, L. De Petrocellis, E. Morera, F. Rossi, A. Schiano
Moriello, M. Nalli, V. de Novellis, D.F. Woodward, S. Maione, V. Di Marzo,
J. Med. Chem. 50 (2007) 6554.
[6] M.S. Chong, J. Hester, Drugs 67 (2007) 569.
[7] P.G. Jensen, J.R. Larson, Drugs Aging 18 (2001) 737.
[8] (a) C.A. Maggi, Life Sci. 51 (1992) 1777;
[24] S. Maione, L. De Petrocellis, V. de Novellis, A. Schiano Moriello, S. Petrosino,
E. Palazzo, F.S. Rossi, D.F. Woodward, V. Di Marzo, Br. J. Pharmacol. 150 (2007)
766.
(b) D. DeRidder, V. Chandiramani, P. Dasgupta, H. VanPoppel, L. Baert,
C.J. Fowler, J. Urol. 158 (1997) 2087.
[25] http://www.molinspiration.com/cgi-bin/properties.
[9] (a) C.S.J. Walpole, S. Bevan, G. Bovermann, J.J. Boelsterli, R. Breckenridge,
J.W. Davies, G.A. Hughes, I. James, L. Oberer, J. Med. Chem. 37 (1994) 1942;
(b) K.M. Walker, L. Urban, S.J. Medhurst, S. Patel, M. Panesar, A.J. Fox,
P. McIntyre, J. Pharmacol. Exp. Ther. 304 (2003) 56.
[10] A. Szallasi, D.N. Cortright, C.A. Blum, S.R. Eid, Nat. Rev. Drug Discov. 6 (2007)
357.
[26] (a) Y.G. Suh, U. Oh, Curr. Pharm. Des. 11 (2005) 474. J. Lazar, D.C. Braun, A.
ꢁ
Toth, Y. Wang, L.V. Pearce, V.A. Pavlyukovets, P.M. Blumberg, S.H. Garfield, S.
Wincovitch, H.-K. Choi, J. Lee, Mol. Pharmacol. 69 (2006) 1166;
(b) J. Vriens, G. Appendino, B. Nilius, Mol. Pharmacol. 75 (2009) 1262;
(c) P. Ambrosino, M.V. Soldovieri, C. Russo, M. Taglialatela, Br. J. Pharmacol.
168 (2013) 1430;
[11] (a) K.R. Bley, Expert. Opin. Investig. Drugs 13 (2004) 1445;
(b) H. Knotkova, M. Pappagallo, A. Szallasi, Clin. J. Pain 24 (2008) 142;
(c) S.J. Conway, Chem. Soc. Rev. 37 (2008) 1530.
(d) L. De Petrocellis, A. Schiano Moriello, G. Fontana, A. Sacchetti, D. Passarella,
G. Appendino, V. Di Marzo, Br. J. Pharmacol. 171 (2014) 2608.