Synthesis and structure-activity relationship studies of novel dual inhibitors of soluble epoxide hydrolase and 5-lipoxygenase

Article abstract of DOI:10.1021/jm301617j

Current research leads to the assumption that drugs affecting more than one target could result in a more efficient treatment of diseases and fewer safety concerns. Administration of drugs inhibiting only one branch of the arachidonic acid cascade is usua

Full text of DOI:10.1021/jm301617j

Brief Article
pubs.acs.org/jmc
Synthesis and Structure−Activity Relationship Studies of Novel Dual
Inhibitors of Soluble Epoxide Hydrolase and 5‑Lipoxygenase
Karin Meirer, Carmen B. Rodl, Joanna M. Wisniewska, Sven George, Ann-Kathrin Hafner, Estel·la Buscato,
́
̈
̈
Franca-Maria Klingler, Steffen Hahn, Dirk Berressem, Sandra K. Wittmann, Dieter Steinhilber,
Bettina Hofmann, and Ewgenij Proschak*
Institute of Pharmaceutical Chemistry, ZAFES/LiFF/OSF, Goethe-University Frankfurt, Max-von-Laue-Str. 9, D-60438
Frankfurt/Main, Germany
S
* Supporting Information
ABSTRACT: Current research leads to the assumption that drugs affecting more than one target could result in a more efficient
treatment of diseases and fewer safety concerns. Administration of drugs inhibiting only one branch of the arachidonic acid
cascade is usually accompanied by side effects. We therefore designed and synthesized a library of hybrid molecules incorporating
an imidazo[1,2-a]pyridine and an urea moiety as novel soluble epoxide hydrolase (sEH)/5-lipoxygenase (5-LO) dual inhibitors.
Evaluation of the compounds was accomplished by in vitro testing using recombinant enzyme assays.
with restrictions due to its liver toxicity and a short half-life.5
Recent research therefore focuses on the potential of dual
inhibitors such as those for prostaglandin/leukotriene synthesis.
An example for this is licofelone6 (Figure 1), which inhibits the
5-LO activating protein (FLAP), the COX-1, and the
microsomal prostaglandin E synthase-1 (mPGES-1).4,7
Besides the COX and LOX pathways, CYP enzymes also
metabolize arachidonic acid to either hydroxyeicosatetraenoic
acids (HETEs) or epoxyeicosatrienoic acids (EETs). The latter
are further hydrolyzed by the soluble epoxide hydrolase (sEH)
to their corresponding diols, the dihydroxyeicosatrienoic acids
(DHETs).8−10 EETs exhibit anti-inflammatory and antihyper-
tensive properties and have been reported to exert beneficial
effects in disease models related to cardiovascular diseases,
diabetes, inflammatory pain, and other indications.8,10
INTRODUCTION
■
The approach “one drug−one target−one disease” was the
predominant principle in drug development over decades.
However, when considering treatment of complex diseases,
interaction with one target may not be enough. Clinical studies
on approved drugs showed that we may consider a rational
design of drugs that interact with multiple targets but do not
interact with off-targets responsible for side effects. These so-
called designed multiple ligands (DMLs) have the advantage
over combination therapies that there are fewer safety concerns
because drug−drug interactions need not be considered.1
The arachidonic acid cascade is involved in many
inflammatory processes due to the production of diverse
inflammatory mediators such as prostaglandins and leuko-
trienes, products of the cyclooxygenase (COX) and the
lipoxygenase (LOX) pathways.2 The 5-lipoxygenase (5-LO) is
a dioxygenase that catalyzes two steps in the biosynthesis of
leukotrienes which regulate the innate immune response and
play a pathophysiological role in chronic inflammatory diseases
such as asthma and atherosclerosis.3 As reviewed by Pergola
and Werz,4 5-LO inhibitors have been developed for over 25
years, but the only inhibitor that entered the US market yet, is
Zileuton (Figure 1), a drug used for the treatment of asthma
Both sEH−/− mice and animals treated with sEH inhibitors
show an increased level of EETs and 5-LO products, indicating
that sEH inhibitors might synergize with COX- and 5-LO
inhibitors. Co-administration of the sEH inhibitor t-AUCB
(Figure 1) with either a COX- or 5-LO inhibitor showed a
significant enhancement of their anti-inflammatory activities.11
sEH inhibitors alone induced albuminuria in mice, probably
due to a shift toward the 5-LO branch of the arachidonic acid
cascade.12 Hwang et al. developed dual COX-2/sEH inhibitors
and showed in a lipopolysaccharide induced mouse model that
the antiallodynic activity of the dual inhibitor is more effective
than either single-target inhibitor alone or the coadministration
of a COX-2 and a sEH inhibitor.13
Therefore, the development of DMLs inhibiting both 5-LO
and sEH might lead not only to highly interesting and effective
anti-inflammatory compounds, but also to safer antihyperten-
sive drugs. We synthesized hybrid molecules out of two known
pharmacophores of sEH and 5-LO inhibitors to generate a tool
compound for the further evaluation of this hypothesis.
Received: November 2, 2012
Published: January 28, 2013
Figure 1. Known inhibitors of sEH and 5-LO.
© 2013 American Chemical Society
1777
dx.doi.org/10.1021/jm301617j | J. Med. Chem. 2013, 56, 1777−1781

Journal of Medicinal Chemistry
Brief Article
inhibitors.10 Besides adamantyl (F) and cyclohexyl (E)
residues, known from AUDA10 and CDU10 (Figure 1), we
also introduced para-substituted phenyl groups (A−D) that
showed improved pharmacokinetic properties in animal
studies.15 The linkage between these two pharmacophores
was achieved with either a piperidine ring or a propyl linker
(Figure 2).10,13,16
RESULTS AND DISCUSSION
■
Chemistry. The recently published imidazo[1,2-a]pyridine
EP6 (Figure 1) was used as backbone for the 5-LO
pharmacophore, with variations on four moieties (Figure
2).14 Urea compounds are a well-known class of sEH
In both cases, formation of the urea (2, 6a−e) was achieved
by reacting an amine with an isocyanate.17 The secondary urea
incorporating the piperidine ring 2 underwent a Mitsunobu
reaction18 with 4-hydroxybenzaldehyde to form the desired
intermediate 3. The last step was a Groebke−Blackburn−
Bienayme
́
multicomponent reaction19 with an isonitrile and 6-
amino-3-picoline under catalytic acidic conditions to form the
desired DMLs 4a−b (Table 1). In contrast to the Mitsunobu
reaction, ureas 6a−e reacted with a hydroxybenzaldehyde
derivative in a Williamson ether synthesis20 to form
intermediates 7a−g. Formation of DMLs 8a−t was accom-
plished in the same way as for compounds 4a−b with the
difference of using diverse amino pyridine derivatives (Scheme
1).
Variation of the sEH Backbone Introducing a Differ-
ent Linker. We therefore implemented a propyl linker
between the two pharmacophores, combined with variations
on the terminal hydrophobic moiety, as better inhibitory
potential and better pharmacokinetics have been postulated for
para-substituted aryl groups with either a trifluoromethyl (C)
or a trifluoromethoxy (D) group.13,15 The new linker increased
Figure 2. Combinatorial strategy for dual inhibitors of sEH/5-LO. R1
= −Me, −F, −Cl; R2 = −ipropyl, −cyclopentyl, −cyclohexyl; R3 = −H,
−OMe, −Cl, C, D; R4 = −OMe, A, B, C, D, E, F.
Table 1. Inhibitory Evaluation of Compounds 4a−b and 8a−t in Vitro with Recombinant Human Enzymes
a
IC50 [nM] SD
entry
EP6
R1
R2
R3
R4
5-LO
sEH
>10000
59 22.9
Zileuton
AUDA
2
591 63.5
107 14.7
>10000
2237 495.4
4a
4b
Me
Me
cyclohexyl
isopropyl
H
A
106 24.9
65 15.9
1467 153.7
1540 155.8
H
A
8a
8b
8c
8d
8e
8f
Me
Me
Me
Cl
cyclohexyl
cyclohexyl
cyclohexyl
cyclohexyl
cyclohexyl
cyclohexyl
cyclohexyl
cyclopentyl
cyclopentyl
isopropyl
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
D
B
70 16.3
44 7.0
842 20.0
439 97.3
116 14.7
191 26.7
336 26.1
696 112.7
649 140.4
473 47.2
614 51.9
558 91.1
620 54.7
459 91.8
480 156.5
439 52.1
1107 102.1
1593 177.5
253 57.0
192 32.7
189 60.6
190 41.6
C
D
63 15.8
32 4.2
D
Cl
C
30 7.6
F
D
82 20.1
52 10.1
71 11.1
70 18.9
147 15.2
201 73.5
92 47.6
42 6.5
8g
8h
8i
F
C
Me
Me
Me
Me
Cl
D
C
8j
D
8k
8l
isopropyl
C
cyclopentyl
cyclopentyl
isopropyl
C
8m
8n
8o
8p
8q
8r
8s
8t
F
C
Cl
C
104 83.8
38 8.7
Me
Me
Me
Me
Me
Me
cyclohexyl
cyclohexyl
cyclohexyl
cyclohexyl
cyclohexyl
cyclohexyl
OMe
OMe
C
C
38 16.5
62 8.5
H
Cl
C
189 116.7
100 34.5
242 81.3
OMe
OMe
F
E
a
IC50 values are means with SD from at least three independent experiments.
1778
dx.doi.org/10.1021/jm301617j | J. Med. Chem. 2013, 56, 1777−1781

Journal of Medicinal Chemistry
Brief Article
a
Scheme 1. Synthesis of Compounds 4a−b and 8a−t
a
(a) Chlorophenylisocyanate, DCM, 0 °C to RT, 20 min, quant; (b) 4-hydroxybenzaldehyde, TPP, DIAD, THF, argon, 0 °C to RT, overnight, 60%;
(c) R2-isonitrile, 6-amino-3-picoline, AcOH, MeOH, RT, overnight, 19−22%; (d) R-amine, DCM, 0 °C to RT, overnight, 64−99%; (e) 3-R3-4-
hydroxybenzaldehyde or 3-hydroxy-4-methoxybenzaldehyde, Cs2CO3, DMF, 70 °C, overnight, 8−56%; (f) R2-isonitrile, 2-amino-5-R1-pyridine,
AcOH, MeOH, overnight, 7−64%. R = p-Cl-phenyl, p-CF3-phenyl, p-OCF3-phenyl, −adamantyl, −cyclohexyl; R1 = −Me, −F, −Cl; R2 = −ipropyl,
−cyclopentyl, −cyclohexyl; R3 = −H, −OMe, −Cl, C, D; R4 = −OMe, A, B, C, D, E, F.
sEH inhibition 2-fold (8a, IC50 = 842 nM) compared to 4a and
4b, whereas inhibition of 5-LO was maintained. Introduction of
the trifluoromethyl group (8b) raised the sEH inhibition again
2-fold, while incorporation of the trifluoromethoxy group into
the molecule (8c) gave a further advance to sEH inhibition,
leading to the best IC50 value of 116 nM. 5-LO inhibition
Modifications of R4 with Nonaromatic Residues (E,F).
Compounds such as AUDA and CDU showed that an aromatic
moiety is not necessary for the inhibition of sEH.10 We
therefore combined our 5-LO backbone with an urea
connected with a propyl linker to either an adamantyl (F, 8s)
or a cyclohexyl (E, 8t) moiety. While IC50 values for sEH
inhibition were constant at 189 and 190 nM, slightly lower than
the best compound 8c, we obtained a decrease in 5-LO
inhibition (8s, IC50 = 100 nM; 8t, IC50 = 242 nM) compared to
8a−c.
remained constant throughout these modifications (8b, IC50
=
44 nM; 8c, IC50 = 63 nM). Therefore, a propyl spacer is
favorable for the continuing design of inhibitors, in analogy to
the studies by Hwang et al.13
Taken together, we developed dual inhibitors with adequate
inhibitory potential toward both target enzymes. Optimization
on the target 5-LO led to IC50 values in a nanomolar range, 30
nM being the best value (8e). If we compare the dual inhibitor
8e with the 5-LO inhibitor EP6 (59 nM), the values are in the
same range, showing that the sEH inhibitor part of 8e is
tolerated by the 5-LO very well. Inhibition of the sEH reached
values in the range of 116−192 nM (8c, 8d, 8r, 8s, 8t), which
are also comparable to AUDA (107 nM). Compounds 8r and
8t are equipotent on the two targets, whereas 8c and 8d
combine the best sEH inhibitory potency with one of the best
potentials toward 5-LO.
SAR of Dual Inhibitors Incorporating a Propyl Linker.
From this starting point (8a−8c), we developed further
variations of inhibitors, modifying each moiety in order to
enhance inhibitory potential. The methyl group in the position
R1 was exchanged by fluorine or chlorine. While sEH inhibition
was mostly unaffected by chlorine (8d, IC50 = 191 nM; 8e, IC50
= 336 nM), introduction of fluorine (8f, IC50 = 696 nM; 8g,
IC50 = 649 nM) diminished the inhibitory potential of the
compounds. Also, for 5-LO inhibition, introduction of chlorine
(8d, IC50 = 32 nM; 8e, IC50 = 30 nM) was preferred over
fluorine (8f, IC50 = 82 nM; 8g, IC50 = 52 nM).
The next step was the implementation of smaller aliphatic
residues in moiety R2. We observed the tendency for sEH that
cyclohexyl (8b, 8c) is preferred over cyclopentyl (8h, 8i) or
isopropyl (8j, 8k), indicating that a larger moiety improves the
inhibitory potential. Cyclohexyl and cyclopentyl, with com-
parable values, were also preferred on 5-LO over an isopropyl
residue. Combination of fluorine or chlorine in moiety R1 with
cyclopentyl (8l, IC50 = 459 nM, 8m, IC50 = 480 nM) or
isopropyl (8n, IC50 = 439 nM) in moiety R2 led to a decrease in
sEH inhibition relative to the best inhibitor 8c. But when
comparing 8l, 8m, and 8n to compounds with a single
modification on either R1 or R2 (8d, 8g, 8i, 8k), sEH inhibition
was maintained or increased. For 5-LO, combination of fluorine
with cyclopentyl 8m is comparable to other potent inhibitors
(8b, 8d, 8e, 8g), showing an IC50 value of 42 nM.
CONCLUSION
■
We report the rational design and synthesis of dual inhibitors of
sEH and 5-LO. Compounds synthesized for a SAR showed
good inhibitory potential on recombinant enzyme for both
targets. In analogy to previous studies,13,21 it was possible to
incorporate the sEH pharmacophore into a dual-target
inhibitor. These compounds can be used as tools to further
assess the idea of adopting dual sEH/5-LO inhibitors as drugs
targeting several disease models. Whether an equipotent
inhibition (8r, 8t) is more favorable over a higher inhibition
on one of the two targets (8c, 8d) needs to be evaluated.
EXPERIMENTAL SECTION
■
A connection between the two pharmacophores in the meta
position (8o, 8p) led to equivalent values in terms of 5-LO
inhibition, but was not tolerated by sEH, possibly due to
sterical hindrance. Besides meta substitution, modifications on
moiety R3 showed that either a methoxy group (8b, IC50 = 44
nM) or a hydrogen atom (8q, IC50 = 62 nM) was preferred
over a chlorine (8r, IC50 = 189 nM) for 5-LO, whereas for sEH,
the chlorine (8r, IC50 = 192 nM) and the hydrogen atom (8q,
IC50 = 253 nM) were favored over the methoxy group (8b, IC50
= 439 nM).
General. All reagents and solvents were purchased from the
suppliers Sigma-Aldrich Chemistry GmbH (Hannover, Germany) or
Alfa-Aesar GmbH & Co. KG (Karlsruhe, Germany) and were used
without further purification. Flash chromatography was performed on
packed silica columns (particle size 50 μm) from Varian Medical
Systems GmbH (Darmstadt, Germany). NMR spectra were measured
on AV 250 and AV 300 nuclear magnetic resonance spectrometers
from Bruker. Chemical shifts are reported in parts per million (ppm)
using TMS as internal standard: 1H (250/300 MHz), 13C (63/75
MHz), 19F (282 MHz). Mass spectra were measured using ESI with a
VG Platform II spectrometer by Fisons or a Mariner Biospectrometry
1779
dx.doi.org/10.1021/jm301617j | J. Med. Chem. 2013, 56, 1777−1781

Journal of Medicinal Chemistry
Brief Article
workstation by Perspective Biosystems. HRMS spectra were measured
by a MALDI LTQ Orbitrap XL spectrometer from Thermo Scientific.
Combustion analysis was performed on an Elementar Vario Micro
Tube CHNO rapid elemental analyzer. HPLC was performed on a
LC2020 (Shimadzu, Duisburg, Germany), conditions: water/methanol
gradient run 5−95% using a Kinetex 2.6 μm C18 100 Å, 100 mm × 2.1
mm (Phenomenex, Aschaffenburg, Germany) at room temperature,
UV detection at 254 and 280 nm. All compounds were characterized
(0.08 g, 0.21 mmol) were stirred with glacial acetic acid (0.01 mL, 0.21
mmol) in 20 mL of MeOH, and then N-cyclohexylisonitrile (0.02 g,
0.21 mmol) was added. The mixture was stirred overnight at room
temperature. The reaction was quenched with 2N HCl. The solvent
was removed under reduced pressure. The residue was solved in 20
mL of EtOAc and washed (3 × 20 mL) with saturated NaHCO3. The
organic phase was dried over MgSO4, the solvent was removed under
reduced pressure, and the crude product was recrystallized from
EtOH. The product was isolated as a yellow solid (26 mg, 0.05 mmol,
22%). 1H NMR (250 MHz, DMSO-d6): δ 8.6 (s, 1H), 8.15−8.06 (m,
2H), 7.51 (d, J = 16.7 Hz, 2H), 7.35−7.27 (m, 3H), 7.01−6.96 (m,
3H), 4.15 (d, J = 21.5 Hz, 2H), 3.89 (d, J = 6.4 Hz, 2H), 2.9−2.78 (m,
1H), 2.31 (s, 3H), 2.02−1.92 (m, 1H), 1.84−1.48 (m, 9H), 1.27−1.02
(m, 9H). 13C NMR (75 MHz, DMSO-d6): 157.6, 154.6, 139.7, 139.5,
134.9, 128.0, 127.6, 127.4, 126.4, 125.1, 124.3, 120.8, 120.5, 120.1,
115.9, 114.1, 71.7, 56.2, 43.6, 35.6, 33.4, 28.5, 25.4, 24.5, 17.8. MS
(ESI, 70 eV) m/z (%): 573.0 (100) (M + H). Anal. Calcd for
C33H38ClN5O2·0.25HCl: C, 68.19, H, 6.63, N, 12.05. Found: C, 68.44,
H, 6.54, N, 12.06.
Synthesis and analytical data for compounds 4a−b and 8a−t and
their intermediates are detailed in the Supporting Information.
sEH Activity Assay. For the determination of inhibitory potential
of our compounds, we used recombinant, affinity-purified human sEH
from Escherichia coli. The assay setup was as previously described,22
using a fluorescence-based assay with the substrate 3-phenyl-cyano(6-
methoxy-2-naphthalenyl)methylester-2-oxiraneacetic acid (PHOME).
IC50 values were determined in a 96-well format utilizing 2 μg/well
recombinant sEH. After incubation with inhibitors for 30 min at room
temperature in 25 mM Bis-Tris/HCl, 0.1 mg/mL BSA, and 0.01%
TritonX 100 buffer (110 μL, pH 7.0), the substrate (PHOME) was
added ([S]final = 50 μM). Activity was evaluated by measuring the
appearance of the fluorescent product 6-methoxy-2-naphthaldehyde
(λem = 330 nm, λex = 465 nm).
1
by H NMR and MS, final compounds additionally by 13C NMR, 19F
NMR, and HRMS, and combustion analysis or HPLC to determine
purity >95%. Activity assays for sEH22 and 5-LO23 were performed
according to earlier publications and are detailed in the Supporting
Information. All IC50 values are means with SD obtained from
measurements at six−eight different concentrations of the compounds
from at least three separate experiments. IC50 values were determined
using a four-parameter sigmoidal dose response equation (in the case
of Zileuton, five different concentrations from three separate
experiments were used to fit a three-parameter sigmoidal dose
response equation) from GraphPad Prism software (GraphPad
Software, LaJolla, CA, USA) and constitute relative IC50 values.
General Procedure d for Compounds 6a−e. 3-Chloropropyliso-
cyanate (5) (1 equiv) was added to a solution of trifluoromethylaniline
(or other aniline derivative, 1 equiv) in DCM at 0 °C. The mixture was
allowed to warm to room temperature and was stirred overnight. A
yellow solution was obtained. The solvent was removed under reduced
pressure and the white solid was triturated with hexane to form the
pure product.
General Procedure e for Compounds 7a−i. Vanillin (or derivative,
1 equiv) and cesium carbonate (1 equiv) in 40 mL of DMF were
heated to 70 °C for 30 min. Compound 6a−e (1 equiv), dissolved in
DMF, was added, and the mixture was stirred overnight at 70 °C. The
solvent was removed, and the residue was diluted in 20 mL of EtOAc.
The organic phase was extracted with 20 mL of water, twice with 20
mL of NaOH solution (1M), and again with 20 mL of water. After
drying the organic layer over MgSO4, the solvent was removed under
reduced pressure. The product was recrystallized from hexane/EtOAc.
If necessary, a further recrystallization from EtOAC/EtOH or flash
column chromatography with the same solvents was performed in
order to obtain the pure products.
General Procedure c for Compound 4b and f for Compounds
8a−t. 6-Amino-3-picoline (or other pyridine amine, 1.2 equiv) and 3
or 7a−i (1 equiv) were stirred with glacial acetic acid in 20 mL of
MeOH, and then N-cyclohexylisonitrile (or other isonitrile, 1 equiv)
was added at 0 °C. After stirring overnight at room temperature, the
reaction mixture was quenched with 2N HCl. The solvent was
removed under reduced pressure. The residue was solved in 20 mL of
EtOAc and washed (3 × 20 mL) with saturated NaHCO3. The organic
phase was dried over MgSO4, the solvent removed under reduced
pressure, and the crude product was recrystallized from hexane/EtOAc
or by flash column chromatography using hexane/EtOAc or EtOAc/
EtOH.
Synthesis of N-(4-Chlorophenyl)-4-(hydroxymethyl)piperidine-1-
carboxamide (2). 4-Chlorophenylisocyanate (0.33 g, 2.15 mmol) was
slowly added to a solution of 4-piperidinemethanol (1) (0.25 g, 2.17
mmol) in DCM at 0 °C. The mixture was allowed to warm to room
temperature and was stirred for further 20 min. The solvent was
removed and the crude product was triturated with hexane to form the
pure product as a white solid (0.55 g, 2.03 mmol, 94%).
5-LO Activity Assay. The evaluation of the activity on purified 5-
LO was accomplished with the method published by Brungs et al.23
Therefore, 2−3 μg of partially purified human 5-LO from E. coli were
diluted in 1 mL of 1 mM PBS/EDTA solution. Preincubation with the
test compounds or vehicle (DMSO) for 15 min at 4 °C was followed
by prewarming for 30 s at 37 °C. Afterward, the reaction was started
by the addition of 2 mM CaCl2 and 20 μM AA at 37 °C. Ice-cold
MeOH (1 mL) was added to stop the reaction after 10 min.
Metabolites formed during the reaction were analyzed by HPLC.
ASSOCIATED CONTENT
* Supporting Information
■
S
Synthesis and analytical data of all compounds. This material is
available free of charge via the Internet at http://pubs.acs.org.
AUTHOR INFORMATION
Corresponding Author
*E-mail: proschak@pharmchem.uni-frankfurt.de.
■
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
This work was supported by the Deutsche Forschungsgemein-
schaft (DFG, Sachbeihilfe PR 1405/2-1), LOEWE Lipid
Signaling Forschungszentrum Frankfurt (LiFF), LOEWE-
Schwerpunkt: Anwendungsorientierte Arzneimittelforschung
(Fraunhofer-TMP), the Oncogenic Signaling Frankfurt
(OSF), Fonds der Chemischen Industrie. K.M. thanks
FIRST, E.B. thanks DAAD-La Caixa.
Synthesis of N-(4-Chlorophenyl)-4-((4-formylphenoxy)methyl)-
piperidine-1-carboxamide (3). 2 (0.25 g, 0.93 mmol), 4-hydroxy-
benzaldehyde (0.1 g, 0.78 mmol), and TPP (0.25 g, 0.97 mmol) were
stirred in 20 mL of anhydrous DMF under argon atmosphere in an ice
bath. DIAD (0.19 mL, 0.97 mmol) was added dropwise in 10 mL of
DMF. After 24 h, the solvent was removed under reduced pressure
and the crude mixture was purified by flash column chromatography
using hexane:EtOAc 2:3.The product was isolated as an off-white solid
(0.17g, 0.47 mmol, 60.2%).
ABBREVIATIONS USED
t-AUCB, trans-4-[4-(3-adamantan-1-yl-ureido)-cyclohexyl-oxy]-
benzoic acid; AUDA, 12-(3-adamantan-1-yl-ureido)dodecanoic
■
Synthesis of N-(4-Chlorophenyl)-4-((4-(3-(cyclohexyl-amino)-6-
methylimidazo[1,2-a]pyridin-2-yl)phenoxy)-methyl)piperidine-1-
carboxamide (4a). 6-Amino-3-picoline (0.03 g, 0.25 mmol) and 4
1780
dx.doi.org/10.1021/jm301617j | J. Med. Chem. 2013, 56, 1777−1781

Journal of Medicinal Chemistry
Brief Article
(15) Tsai, H.-J.; Hwang, S. H.; Morisseau, C.; Yang, J.; Jones, P. D.;
Kasagami, T.; Kim, I.-H.; Hammock, B. D. Pharmacokinetic screening
of soluble epoxide hydrolase inhibitors in dogs. Eur. J. Pharm. Sci.
2010, 40, 222−238.
(16) Eldrup, A. B.; Soleymanzadeh, F.; Farrow, N. A.; Kukulka, A.;
De Lombaert, S. Optimization of piperidyl-ureas as inhibitors of
soluble epoxide hydrolase. Bioorg. Med. Chem. Lett. 2010, 20 (2), 571−
575.
acid; CDU, 1-cyclohexyl-3-dodecyl urea; DHET, dihydroxyei-
cosatrienoic acid; DML, designed multiple ligand; EET,
epoxyeicosatrienoic acid; HETE, hydroxyeicosatetranoic acid;
5-LO, 5-lipoxygenase; 5-LO-p, 5-LO pharmacophore; FLAP, 5-
LO activating protein; mPGES-1, microsomal prostaglandin E
synthase-1; sEH, soluble epoxide hydrolase
(17) (a) Aicher, T. D.; Anderson, R. C.; Gao, J.; Shetty, S. S.;
Coppola, G. M.; Stanton, J. L.; Knorr, D. C.; Sperbeck, D. M.; Brand,
L. J.; Vinluan, C. C.; Kaplan, E. L.; Dragland, C. J.; Tomaselli, H. C.;
Islam, A.; Lozito, R. J.; Liu, X.; Maniara, W. M.; Fillers, W. S.;
DelGrande, D.; Walter, R. E.; Mann, W. R. Secondary amides of (R)-
3,3,3-trifluoro-2-hydroxy-2-methylpropionic acid as inhibitors of
pyruvate dehydrogenase kinase. J. Med. Chem. 2000, 43 (2), 236−
249. (b) Bernstein, P.; Hill, D.; Schooler, M.; Shenvi, A.; Jacobs, R.
Chroman Compounds as 5 -HTlB Antagonists. (Astrazeneca AB)
WO/2007/053094 A1; US SE2006/001253, May 10, 2007.
(18) Mitsunobu, O. The Use of Diethyl Azodicarboxylate and
Triphenylphosphine in Synthesis and Transformation of Natural
Products. Synthesis 1981, 1−28.
REFERENCES
■
(1) (a) Morphy, R.; Rankovic, Z. Designed multiple ligands. An
emerging drug discovery paradigm. J. Med. Chem. 2005, 48, 6523−
6543. (b) Morphy, J. R.; Kay, C.; Rankovic, Z. From magic bullets to
designed multiple ligands. Drug Discovery Today 2004, 9, 641−651.
(2) Hyde, C. A.; Missailidis, S. Inhibition of arachidonic acid
metabolism and its implication on cell proliferation and tumour-
angiogenesis. Int. Immunopharmacol. 2009, 9 (6), 701−715.
(3) Radmark, O.; Werz, O.; Steinhilber, D.; Samuelsson, B. 5-
̊
Lipoxygenase: regulation of expression and enzyme activity. Trends
Biochem. Sci. 2007, 32, 332−341.
(4) Pergola, C.; Werz, O. 5-Lipoxygenase inhibitors: a review of
recent developments and patents. Expert Opin. Ther. Pat. 2010, 20 (3),
355−375.
(19) Chartres, J. D.; Groth, A. M.; Lindoy, L. F.; Meehan, G. V.
Metal ion recognition. Selective interaction of silver(I) with trilinked
N2S2-donor macrocycles and their single-ring analogues. J. Chem. Soc.,
Dalton Trans. 2002, 371−376.
(5) (a) Sorkness, C. A. The use of 5-lipoxygenase inhibitors and
leukotriene receptor antagonists in the treatment of chronic asthma.
Pharmacotherapy. 1997, 17, 50S−54S. (b) Fischer, A. R.; McFadden,
C. A.; Frantz, R.; Awni, W. M.; Cohn, J.; Drazen, J. M.; Israel, E. Effect
of chronic 5-lipoxygenase inhibition on airway hyperresponsiveness in
asthmatic subjects. Am. J. Respir. Crit. Care Med. 1995, 152, 1203−
1207.
(6) Laufer, S.; Tries, S.; Augustin, J.; Dannhardt, G. Pharmacological
profile of a new pyrrolizine derivative inhibiting the enzymes
cyclooxygenase and 5-lipoxygenase. Arzneim. Forsch. 1994, 44 (5),
629−636.
(20) Groebke, K.; Weber, L.; Mehlin, F. Synthesis of Imidazo[1,2-a]
Annulated Pyridines, Pyrazines, and Pyrimidines by a Novel Three-
Component Condensation. Synlett 1998, 29 (39), 661−663.
́
(21) Buscato, E.; Blocher, R.; Lamers, C.; Klingler, F. M.; Hahn, S.;
̈
Steinhilber, D.; Schubert-Zsilavecz, M.; Proschak, E. Design and
synthesis of dual modulators of soluble epoxide hydrolase and
peroxisome proliferator-activated receptors. J. Med. Chem. 2012, 55
(23), 10771−10775.
́
(22) Hahn, S.; Achenbach, J.; Buscato, E.; Klingler, F.-M.; Schroeder,
(7) (a) Fischer, L.; Hornig, M.; Pergola, C.; Meindl, N.; Franke, L.;
Tanrikulu, Y.; Dodt, G.; Schneider, G.; Steinhilber, D.; Werz, O. The
molecular mechanism of the inhibition by licofelone of the
biosynthesis of 5-lipoxygenase products. Br. J. Pharmacol. 2007, 152,
M.; Meirer, K.; Hieke, M.; Heering, J.; Barbosa-Sicard, E.; Loehr, F.;
Fleming, I.; Doetsch, V.; Schubert-Zsilavecz, M.; Steinhilber, D.;
Proschak, E. Complementary screening techniques yielded fragments
that inhibit the phosphatase activity of soluble epoxide hydrolase.
ChemMedChem 2011, 6, 2146−2149.
471−480. (b) Koeberle, A.; Siemoneit, U.; Buhring, U.; Northoff, H.;
̈
Laufer, S.; Albrecht, W.; Werz, O. Licofelone suppresses prostaglandin
E2 formation by interference with the inducible microsomal
prostaglandin E2 synthase-1. J. Pharmacol. Exp. Ther. 2008, 326,
975−982.
(23) Brungs, M.; Radmark, O.; Samuelsson, B.; Steinhilber, D.
̊
Sequential induction of 5-lipoxygenase gene expression and activity in
Mono Mac 6 cells by transforming growth factor beta and 1,25-
dihydroxyvitamin D3. Proc. Natl. Acad. Sci. U. S. A. 1995, 92, 107−111.
(8) Imig, J. D. Epoxides and soluble epoxide hydrolase in
cardiovascular physiology. Physiol. Rev. 2012, 92, 101−130.
(9) Morisseau, C.; Hammock, B. D. Impact of soluble epoxide
hydrolase and epoxyeicosanoids on human health. Annu. Rev.
Pharmacol. Toxicol. 2013, 53, 3.11−3.22.
(10) Shen, C. H. Soluble epoxide hydrolase inhibitors: a patent
review. Expert Opin. Ther. Pate. 2010, 20 (7), 941−956.
(11) Liu, J.; Yang, J.; Inceoglu, B.; Qiu, H.; Ulu, A.; Hwang, S.;
Chiamvimonvat, N.; Hammock, B. D. Inhibition of soluble epoxide
hydrolase enhances the anti-inflammatory effects of aspirin and 5-
lipoxygenase activation protein inhibitor in a murine model. Biochem.
Pharmacol. 2010, 79, 880−887.
(12) Jung, O.; Jansen, F.; Mieth, A.; Barbosa-Sicard, E.; Pliquett, R.
U.; Babelova, A.; Morisseau, C.; Hwang, S. H.; Tsai, C.; Hammock, B.
D.; Schaefer, L.; Geisslinger, G.; Amann, K.; Brandes, R. P. Inhibition
of the soluble epoxide hydrolase promotes albuminuria in mice with
progressive renal disease. PLoS One 2010, 5, e11979.
(13) Hwang, S. H.; Wagner, K. M.; Morisseau, C.; Liu, J. Y.; Dong,
H.; Wecksler, A. T.; Hammock, B. D. Synthesis and structure−activity
relationship studies of urea-containing pyrazoles as dual inhibitors of
cyclooxygenase-2 and soluble epoxide hydrolase. J. Med. Chem. 2011,
54 (8), 3037−3050.
(14) Wisniewska, J. M.; Rodl, C. B.; Kahnt, A. S.; Buscato,
́
E.; Ulrich,
̈
S.; Tanrikulu, Y.; Achenbach, J.; Rorsch, F.; Grosch, S.; Schneider, G.;
̈
̈
Cinatl, J., Jr; Proschak, E.; Steinhilber, D.; Hofmann, B. Molecular
characterization of EP6a novel imidazo[1,2-a]pyridine based direct
5-lipoxygenase inhibitor. Biochem. Pharmacol. 2012, 83 (2), 228−240.
1781
dx.doi.org/10.1021/jm301617j | J. Med. Chem. 2013, 56, 1777−1781