Synthesis, cyclooxygenase inhibitory effects, and molecular modeling study of 4-aryl-5-(4-(methylsulfonyl)phenyl)-2-alkylthio and -2-alkylsulfonyl-1H- imidazole derivatives

Article abstract of DOI:10.1016/j.bmc.2013.01.058

A series of 4-aryl-5-(4-(methylsulfonyl)phenyl)-2-alkylthio and 2-alkylsulfonyl-1H-imidazole derivatives were synthesized. All compounds were tested in human blood assay to determine COX-1 and COX-2 inhibitory potency and selectivity. Among the synthesize

Full text of DOI:10.1016/j.bmc.2013.01.058

Bioorganic & Medicinal Chemistry 21 (2013) 2355–2362
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Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis, cyclooxygenase inhibitory effects, and molecular modeling
study of 4-aryl-5-(4-(methylsulfonyl)phenyl)-2-alkylthio
and -2-alkylsulfonyl-1H-imidazole derivatives
Amir Assadieskandar a, Amirali Amirhamzeh a, Marjan Salehi a, Keriman Ozadali b,c, Seyed Nasser Ostad d,
Abbas Shafiee e, Mohsen Amini a,
⇑
a Department of Medicinal Chemistry, Faculty of Pharmacy and Drug Design & Development Research Center, Tehran University of Medical Sciences, Tehran 14176, Iran
b Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, 2142-L Katz Group Centre for Pharmacy and Health Research, Edmonton, Alberta, Canada T6G 2E9
c Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Hacettepe, Sihhiye, 06100 Ankara, Turkey
d Department of Toxicology and Pharmacology, Faculty of Pharmacy and Rational Drug Use Research Center, Tehran University of Medical Sciences, Tehran, Iran
e Department of Medicinal Chemistry, Faculty of Pharmacy and Pharmaceutical Sciences Research Center, Tehran University of Medical Sciences, Tehran 14176, Iran
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of 4-aryl-5-(4-(methylsulfonyl)phenyl)-2-alkylthio and 2-alkylsulfonyl-1H-imidazole derivatives
were synthesized. All compounds were tested in human blood assay to determine COX-1 and COX-2
inhibitory potency and selectivity. Among the synthesized compounds, 2-alkylthio series were more
potent and selective than 2-sulfonylalkyl derivatives. In molecular modeling, interaction of 2-sul-
fonylalkyl moiety with Arg120 in COX-1 and an extra hydrogen bond with Tyr341 in COX-2 increased
the residence time of ligands in the active site in 2-sulfonylalkyl and 2-alkylthio analogs, respectively.
Ó 2013 Elsevier Ltd. All rights reserved.
Received 20 November 2012
Revised 27 January 2013
Accepted 28 January 2013
Available online 8 February 2013
Keywords:
Cyclooxygenase
Human whole blood assay
Docking
Tautomerization
2-Alkylthio-1H-imidazole
2-Alkylsulfonyl-1H-imidazole
1. Introduction
fects.6 In order to circumvent this drawback of NSAIDs, a new gen-
eration of COX-2 selective inhibitors with diaryl-heterocycle
Cyclooxygenase, the enzyme involved in the first committed
step of arachidonic acid metabolism to prostaglandin H2, has been
elucidated in 1971 to be the target of aspirin and other traditional
non-steroidal anti-inflammatory drugs (NSAIDs).1 Prostaglandins
and thromboxane have been implicated to have active roles in
the pathophysiology of some serious medical conditions including
rheumatoid arthritis,2 osteoarthritis,3 cancer4 and neurodegenera-
tive disorders.5 Since the discovery of the existence of two distinct
isoforms of cyclooxygenase (COX-1 and COX-2) in 1991, several re-
search programs have been devoted to investigate their specific
roles in aforementioned disorders and design selective inhibitors
of each isoform.
COX-1 is generally considered as a constitutional enzyme
responsible for physiologic roles. On the other hand, COX-2 is occa-
sionally an inducible enzyme, which increases during inflamma-
tory conditions. Traditional NSAIDs like indomethacin inhibit
both isoforms and are associated with gastrointestinal (GI) side ef-
scaffold containing a para sulfonylmethyl or sulfamoyl moiety on
one of vicinal aromatic rings have been developed as exemplified
by celecoxib (Celebrex) (1) (Fig. 1). Despite their better GI safety
profile,7 another class related issue has been arisen in chronic
use of COX-2 selective inhibitors. Some of the most popular mem-
bers of these drugs such as rofecoxib (Vioxx) and valdecoxib (Bex-
tra) have been voluntarily withdrawn from the market as a
consequence of fatal cardiovascular side effects.8 It has been impli-
cated that COX-2 inhibition leads to a decrease in prostacyclin
(PGI2) level, a lipid mediator with some cardioprotective effects,
while leaves COX-1 in platelets activated shifting PG metabolism
toward thromboxane A2 (TxA2) synthesis. It seems that this imbal-
ance in PGI2/TxA2 proportion is the reason why chronic use of COX-
2 selective inhibitors results in myocardial infarction (MI) in some
patients.9–11
As could be comprehended, design and synthesis of new cyclo-
oxygenase inhibitors would still merit consideration in order to in-
crease the richness of structural repertoire which contain some
safer lead compounds being discovered. Some diaryl-heterocycles
bearing a 6-alkylthio substituted lactone (pyrane-2-one) as central
⇑
Corresponding author. Tel.: +98 21 66959090; fax: +98 21 66461178.
E-mail address: moamini@sina.tums.ac.ir (M. Amini).
0968-0896/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2013.01.058

2356
A. Assadieskandar et al. / Bioorg. Med. Chem. 21 (2013) 2355–2362
For preparation of compounds 10–13, two different methods
H2NO2S
H3CO2S
O
were considered. In method A, benzoins were oxidized to corre-
sponding sulfoxides using OxoneÒ (potassium peroxymonosul-
fate).12 Obtained products were subjected with excess
ammonium thiocyanate in n-butanol to prepare compound 8a–c.
Subsequent alkylation of 8a–c using alkyl iodide in basic media
afforded the title compounds 10a–c and 12a–c. In method B, ben-
zoins were reacted with excess ammonium thiocyanate in n-buta-
nol to prepare compound 9a–c.20 Subsequent alkylation of 9a–c
using alkyl iodide in basic media followed by oxidation with Ox-
oneÒ, afforded the title compounds 11a–c and 13a–c (Scheme 2).
In the 1H NMR spectra of all compounds collected in DMSO-d6
at ambient temperature, two sets of signals were observed indicat-
ing a slow equilibrium between two imidazole tautomers on the
NMR time scale. This slow equilibrium in 2-(alkylthio)-1H-imidaz-
ole derveties (10a–c and 12a–c) is more distinctive. For example
when the 1H NMR spectrum of 12c was collected in DMSO-d6,
two sets of signals in nearly 3:2 ratio are detectable. However, in
CDCl3 the interconversion of the two isomers is faster on the
NMR time scale and consecutively the resonances are averaging
to a single signal. In the 13C NMR of compound 12b, most of the
aromatic carbons were duplicated. For example for C4 and C3,5 in
4-F-Phe group, two sets of signals in 162.39, 161.83 ppm with
J = 243.75 Hz and 116.39, 115.83 ppm with J = 21.25 Hz were ob-
served, respectively. However, the equilibrium in 2-(alkylsulfo-
nyl)-1H-imidazole derveties (11a–c and 13a–c) is much faster so
that hydrogen and carbon resonances in DMSO-d6 are averaging
to broad signals. This observation showed that the rate of imidaz-
ole ring tautomerization in 2-alkylthio analogs is slower than 2-
sulfonylalkyl congeners (Fig. 2).
N
N
O
CF3
SCH2CH3
H3C
1
(Celecoxib)
N
2
H3CO2S
H3CO2S
N
N
SCH3
N
N
SCH2CH3
F
Br
X
3
4
H3CO2S
N
N
CF3
SOnR
Cl
N
H
N
H
X
5
6 (X = halo, alkyl, alkoxy)
Figure 1. Representative examples of some diaryl-heterocycle COX-2 selective
inhibitors. Anti-inflammatory effect has been only reported for compound 6.
ring have been reported possessing great selectivity toward COX-2
(2).12 We have synthesized 3-alkylthio-4,5-diaryl-1,2,4-triazole
(3)13 and some 2-alkylthio substituted 1,5-diaryl-imidazole ana-
logs (4)14 exhibiting good COX-2 selectivity and anti-inflammatory
effects. Also some 4,5-diaryl-imidazoles (5) bearing a 2-fluorom-
ethyl moiety on central ring have been synthesized and shown to
have promising potency and selectivity.15 Compounds with 4,5-
diaryl-2-(substituted thio or sulfonyl)-imidazole structures lacking
the sulfonylmethyl or sulfamoyl moieties (6) have been already re-
ported to have anti-inflammatory effects in animal models. The
structure–activity relationships of these compounds were dis-
cussed in three parts: (a) the sulfur oxidation; (b) the sulfur substi-
tuent; (c) the aryl substituents. Sulfone moiety and fluorinated
alkyl substituents generally led to the most active analgesic and
antiarthritic derivatives. Although p-methoxy substituents in phe-
nyl rings were favorable in analgesic activity, the greater antiar-
thritic potency was obtained with para halogen-substitution.16,17
Considering structural features of above mentioned com-
2.2. Inhibition of cyclooxygenase
In various structure–activity relationships (SAR) studies, diaryl-
heterocycle compounds possessing two vicinal aryl moieties on the
central heterocyclic ring system represent the major class of selec-
tive COX-2 inhibitors.21 Moreover; the –SO2CH3 group at the para-
position of the aryl rings was frequently substituted and was
shown to confer optimal COX-2 selectivity and potency.22
In this study, we have prepared two classes of compounds pos-
sessing the typical –SO2CH3 COX-2 selectivity; pharmacophore in
the para-position of aryl rings attached to the central 2-alkylthio
or 2-sulfonylalkyl on imidazole ring. Synthesized compounds 10–
13 were evaluated for their ability to inhibit COX-2 and COX-1
enzymatic activity using human whole blood assay. The potency
(IC50 values) of test compounds was determined and compared
to that of the reference molecules SC-560 (selective COX-1 inhibi-
tor) and DuP-697 and celecoxib (selective COX-2 inhibitors). These
results have been summarized in Table 1. It is noteworthy to men-
tion that the functional selectivity assessed in whole blood assay is
usually considerably less than the intrinsic affinity tested by the
purified proteins.23
pounds, some 4,5-diaryl-imidazoles bearing
a sulfonylmethyl
pharmacophore on one of phenyl rings and also containing a 2-
alkylthio or 2-sulfonylalkyl on imidazole ring were synthesized
with the aim of investigating the effect of these groups on COX
selectivity and potency. A human whole blood assay was per-
formed to reveal their inhibitory effects toward COX isoforms.
Docking study was also accomplished to interpret selectivity pat-
tern among synthesized structures.
The most potent COX-2 inhibitors in this series were 10a
(IC50 = 0.06
ence compounds DuP-697 (IC50 = 0.03
(IC50 = 0.87 M). Biological assay results and structure–activity
l
M) and 12a (IC50 = 0.05
l
M) compared to the refer-
2. Results and discussion
2.1. Chemistry
lM) and celecoxib
l
relationship analysis revealed an interesting pattern in COX-2
selectivity of synthesized compounds. Although 2-alkylthio bear-
ing analogs (compounds in 10 & 12 series) exhibit relative COX-2
selectivity (SI nearly 3), in 2-sulfonylalkyl containing structures
(11 & 13 series) selectivity on COX-2 is almost diminished and
comparable IC50 for both isoforms was obtained with a slight pref-
erence toward COX-2. As could be seen in the Table 1, the struc-
ture–activity relationship study of all compounds indicated that
the order of COX-2 inhibitory potency according to R1 substituent
was H > F > Cl. Furthermore, increasing the size in R2 substituent
For preparing unsymmetrical benzoins, methyl phenyl sulfide
was reacted with appropriate phenyl acetic acid derivative.
Subsequently, the products were subjected to bromination
using bromine in glacial acetic acid to obtain related
bromodesoxybenzoins according to previous reports.18 Further,
-bromodesoxybenzoins were refluxed with sodium methoxide
a-
a
in methanol and finally the reaction was quenched with cold 10%
hydrochloric acid to obtain benzoin 7a–c19 (Scheme 1).

A. Assadieskandar et al. / Bioorg. Med. Chem. 21 (2013) 2355–2362
2357
Figure 2. (a) Tautomerization in compound 12c; (b) 1H NMR spectra of 12c in DMSO-d6 at room temperature; (c) 1H NMR spectra of 12c in CDCl3 at room temperature; (d)
13C NMR of 12b in DMSO-d6 at room temperature.
from methyl to ethyl group, led to an increase in COX-2 selectivity.
Amongst synthesized structures, 12c which bears a para chloro-
phenyl group and 2-ethylthio moiety on imidazole ring is the most
selective compound with SI = 3.39.
In Figure 4, the 2-sulfonylmethyl moiety of 11b oriented toward
constriction site, formed an important 2.7 Å hydrogen bond with
Arg120. As a result, it could be presumed that this interaction in
2-sulfonylalkyl derivates led to an increase in COX-1 potency.
Docking studies of synthesized compounds on COX-2 revealed
almost similar orientation of structures in the active site. However,
in 2-alkylthio series, –NH of imidazole ring oriented so that a
hydrogen bond could be formed with the Tyr341 (equivalent to
Tyr355 of COX-1) at the constriction site. It could be presumed that
this interaction made 2-alkylthio analogs more potent and selec-
tive COX-2 inhibitors than 2-sulfonylalkyl analogs. Orientation
and interactions of 12b with COX-2 enzyme have been described
in detail as an example. The SO2CH3 moiety on C-4 phenyl well fit-
ted in the side pocket of COX-2 in such a way that each of its oxy-
gens, hydrogen bonded with His75 (distance of 2.1 Å) and Gln178
(distance of 3.6 Å). The fluorophenyl group of ligand oriented to-
ward catalytic Tyr371 (equivalent to Tyr385 of COX-1) which to-
gether with side chains of Leu338, Trp373 and Phe504 constructs
a relative hydrophobic pocket. The 2-ethylthio group of 12b hosted
in a hydrophobic pocket in vicinity of constriction site which is
lined by lipophilic residues of Met99, Val102, Val335, Leu345 and
Leu517. In the central imidazole ring of 12b and other 2-alkylthio
2.3. Molecular modeling study
Docking study was performed to explain the selectivity pattern
obtained in biological assay at molecular level. 2-Alkylthio and 2-
sulfonylalkyl imidazole analogs, as exemplified by 10b and 11b,
respectively, were almost similarly docked in the COX-1 active site.
In both compounds 10b and 11b, the –SO2CH3 moiety on C-4 phe-
nyl forms two hydrogen bonds with His90 and Gln192 via each of
its oxygens; the fluorophenyl group of both ligands fitted in a
pocket containing catalytic amino acid Tyr385 and side chains of
Trp387, Met522, Phe518 and Leu352. However, differences in the
interaction of 2-alkylthio and 2-sulfonylalkyl analogs with COX-1
arise from their capability to interact with amino acids in constric-
tion site. As could be seen in Figure 3, the 2-methylthio moiety of
10b positioned in a hydrophobic pocket near the constriction site,
which also hosted carboxylate bearing aromatic ring of NSAIDs
(lined by Met113, Val116, Leu359 and Val349 lipophilic residues).

2358
A. Assadieskandar et al. / Bioorg. Med. Chem. 21 (2013) 2355–2362
H3CS
a
H3CS
b
H3CS
c
HO
O
O
O
O
+
Br
OH
SCH3
R1
R1
R1
R1
7a, R1=H
7b, R1=F
7c, R1=Cl
Scheme 1. Reagents and conditions: (a) H3PO4, (CF3CO)2O, 25 °C; (b) Br2, glacial AcOH, rt; (c) CH3ONa, CH3OH, reflux, 10% HCl.
H3CO2S
H3CO2S
H
N
N
c
a
A
S
SR2
N
N
H
b
H
10a, R1=H, R2=CH3
H3CS
10b, R1=F, R2=CH3
8a, R1=H
8b, R1=F
8c, R1=Cl
R1
R1
10c, R1=Cl, R2=CH3
O
12a, R1=H, R2=CH2CH3
12b, R1=F, R2=CH2CH3
12c, R1=Cl, R2=CH2CH3
OH
H3CS
H3CO2S
R1
H
N
N
c
d
S
SO2R2
B
b
N
H
N
H
11a, R1=H, R2=CH3
11b, R1=F, R2=CH3
R1
R1
9a, R1=H
9b, R1=F
9c, R1=Cl
11c, R1=Cl, R2=CH3
13a, R1=H, R2=CH2CH3
13b, R1=F, R2=CH2CH3
13c, R1=Cl, R2=CH2CH3
Scheme 2. Reagents and conditions: (a) 2 mmol OxoneÒ (potassium peroxymonosulfate), CH3OH/THF/H2O, 0 °C, 18 h; (b) NH4SCN, n-butanol, reflux; (c) alkyl iodide, CH3OH,
reflux; (d) 4 mmol OxoneÒ (potassium peroxymonosulfate), CH3OH/THF/H2O, 0 °C, 18 h.
Table 1
IC50 values for the inhibition of COX-1 and COX-2 in the human whole blood assays, COX-2 selectivity index (SI), and
DGintermolecular data
H3CO2S
N
SOnR2
N
H
R1
SIb
D
Gintermolecular (kJ/mol)
a
c
Compounds
n
R1
R2
IC50
(
lM)
COX-1
COX-2
COX-1
COX-2
10a
10b
10c
11a
11b
11c
11a
12b
12c
13a
13b
13c
Celecoxib
SC-560
DuP-697
0
0
0
2
2
2
0
0
0
2
2
2
H
F
Cl
H
F
Cl
H
F
Cl
H
F
CH3
CH3
CH3
CH3
CH3
CH3
CH2CH3
CH2CH3
CH2CH3
CH2CH3
CH2CH3
CH2CH3
0.16
0.21
0.32
0.34
0.39
0.42
0.17
0.24
0.39
0.36
0.40
0.44
6.65
0.0089
—
0.06
0.07
0.10
0.28
0.28
0.31
0.05
0.08
0.11
0.24
0.31
0.32
0.87
—
2.69
3.07
3.18
1.22
1.41
1.35
3.30
2.94
3.39
1.52
1.30
1.35
7.64
À37
À39
À35
À33
À39
À32
À37
À34
À39
À27
À30
À30
À46
À46
À47
À38
À38
À42
À52
À53
À56
À39
À39
À38
Cl
0.03
a
The in vitro test compound concentration required to produce 50% inhibition of enzymatic activity. The result (IC50, l M) is the mean of three determinations acquired
using the human whole blood assay and the deviation from the mean is <10% of the mean value.
b
In vitro COX-2 selectivity index (COX-1 IC50/COX-2 IC50).
HYDE predicted intermolecular free energy.
c

A. Assadieskandar et al. / Bioorg. Med. Chem. 21 (2013) 2355–2362
2359
Figure 3. Compound 10b (carbons in purple) in COX-1. (a) The sulfonylmethyl
group of 4b forms two hydrogen bonds, a 2.1 Å with Gln192 and a 3.6 Å with His90
(pictures are prepared by Molsoft ICM-browser). (b) Pose view image of 10b
orientation in COX-1.
Figure 4. Compound 11b (carbons in purple) in COX-1. (a) The sulfonylmethyl
moiety on phenyl ring of 11b forms two hydrogen bonds, a 1.8 Å with Gln192 and a
4.5 Å with His90. As could be seen one of oxygens of sulfonylmethyl moiety on
imidazole participates in a 2.7 Å hydrogen bond with guadinium group of Arg120 at
the constriction site. (b) Pose view image of 5b orientation in COX-1.
analougs, –NH group forms hydrogen bond with phenolic oxygen
of Tyr341 (Fig. 5).
For additional support, HYDE estimated energy of hydrogen
bond between –NH imidazole of ligand and COX-2 Tyr341 was
studied for all synthesized compounds in Table 2. The HYDE esti-
mated free energy of this interaction for 2-alkylthio analogs was
in the range of À4.5 to À2.0 kJ/mol. However, in 2-sulfonylalkyl
congeners HYDE analysis predicted a weak interaction of this type
just for 13c. According to the HYDE data and the observation from
imidazole ring tautomerization in NMR spectra, it could be postu-
lated that hydrogen bond formation in 2-alkylthio series via –NH of
imidazole ring with Tyr341 might be thermodynamically and
kinetically more favorable.
As discussed in the literature, diaryl-heterocycles like cele-
coxib inhibit COX-2 with time dependent kinetics while COX-1
inhibition takes place time independently and this is the reason
why these structures inhibit COX-2 selectively. It seems that
interaction with Arg120 (Arg106 in COX-2) and Tyr355 (Tyr341
in COX-2) in the constriction site is important for time depen-
dent inhibition.24–27 Limongelli et al. through a metadynamics-
based approach explained time dependent inhibition mechanism
of COX-2 by SC-558, a celecoxib analog. They have revealed that
this ligand takes an alternative mode of orientation from the ori-
ginal one through interaction with the active site. While SC-558
firstly interacts with Arg499 in the side pocket of COX-2 through
its sulfamoyl moiety then reorients in the active site so that the
sulfamoyl moiety forms hydrogen bond with Arg106 and Tyr341
in the constriction site. Because of the smaller space available in
COX-1 active site, this reorientation for SC-558 is not possible.
This extra interaction has been proposed to be the reason for
this observation that SC-558 and its analogs inhibit COX-2 time
dependently while COX-1 inhibition takes place through time
independent kinetics.28 After all it has been speculated that
our molecular modeling study reveals some similar results with
aforementioned study. For 2-alkylthio analogs, which exhibit
COX-2 selectivity, in addition to possibility of reorientation in
Figure 5. Compound 12b (carbons in purple) in COX-2. (a) The sulfonylmethyl
group of 12b forms two hydrogen bonds, a 3.6 Å with Gln178 and a 2.1 Å with His75
at the side pocket of COX-2. As could be seen another hydrogen bonding via –NH of
imidazole ring distanced 2.1 Å to the Tyr341 (equivalent to Tyr355 of COX-1) at the
constriction site has occurred. (b) Pose view image of 12b orientation in COX-2.

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A. Assadieskandar et al. / Bioorg. Med. Chem. 21 (2013) 2355–2362
Table 2
HYDE estimated energy of hydrogen bond between –NH imidazole of ligands and COX-2 Tyr341
Compounds
10a
10b
10c
11a
11b
11c
12a
12b
12c
13a
13b
13c
D
G (kJ/mol)
À3.8
À4.5
À2.0
—
—
—
À4.0
À4.0
À3.2
—
—
À1.5
the COX-2 active site, an extra hydrogen bond with Tyr341 has
been predicted that could increase the residence time of ligands
in the active site. 2-Sulfonylalkyl analogs inhibit both isoforms
almost similarly. The reason for this pattern of selectivity for
11 and 13 series could be explained through molecular model-
ing. While these compounds have structural determinants to in-
hibit COX-2 time dependently, the extra sulfonyl group on
imidazole ring makes the difference through COX-1 inhibition.
Molecular modeling showed that 2-sulfonylalkyl moiety of these
series interacts with Arg120 in COX-1 and this interaction could
increase the residence time of ligand in the active site. Because
of this extra hydrogen bonding, time dependent kinetic of
COX-1 inhibition by 2-sulfonylalkyl analogs would be a strong
possibility. As proposed in literature, NSAIDs like naproxen
which inhibit COX-1 time dependently29 have safer cardiovascu-
lar profile and even reveal some cardioprotective effects.10 All in
all it demands further investigation to reveal COX inhibition
kinetics for synthesized structures and also their cardiovascular
safety profile.
4.1.2. 4-(4-Fluorophenyl)-5-(4-(methylsulfonyl)phenyl)-2-
(methylthio)-1H-imidazole (10b)
Yield, 74%; mp 131–133 °C; IR (KBr, cmÀ1): v 3278 (NH), 1280,
1142 (SO2); 1H NMR (500 MHz, DMSO-d6): d 2.63 (s, 3H, SCH3),
3.19 and 3.25 (two br s, 3H, SO2CH3), 7.19 and 7.31(two dd,
J = 8.5 Hz, J = 8.5 Hz, 2H, H3,5-fluorophenyl), 7.43–7.54 (m, 2H,
H2,6-fluorophenyl), 7.61 and 7.69 (two d, J = 8 Hz, 2H, H2,6-meth-
ylsulfonylphenyl), 7.82 and 7.91 (two d, J = 8 Hz, 2H, H3,5-meth-
ylsulfonylphenyl), 12.70 and 12.73 (two br s, 1H, NH). Anal.
Calcd for C17H15FN2O2S2: C, 56.34; H, 4.17; N, 7.73. Found: C,
56.52; H, 4.35; N, 7.43.
4.1.3. 4-(4-Chlorophenyl)-5-(4-(methylsulfonyl)phenyl)-2-
(methylthio)-1H-imidazole (10c)
Yield, 78%; mp 121–123 °C; IR (KBr, cmÀ1): v 3308 (NH), 1285,
1137 (SO2); 1H NMR (500 MHz, DMSO-d6): d 2.63 (s, 3H, SCH3),
3.20 and 3.25 (two br s, 3H, SO2CH3), 7.36–7.56 (m, 4H, chloro-
phenyl), 7.63 and 7.70 (two d, J = 7.5 Hz, 2H, H2,6-methylsulfonyl-
phenyl), 7.84 and 7.93 (two d, J = 7.5 Hz, 2H, H3,5
methylsulfonylphenyl), 12.77 (br s, 1H, NH). Anal. Calcd for
17H15ClN2O2S2: C, 53.89; H, 3.99; N, 7.39. Found: C, 53.64; H,
-
C
3. Conclusions
3.71; N, 7.62.
A series of 4-aryl-5-(4-(methylsulfonyl)phenyl)-2-alkylthio and
2-alkylsulfonyl-1H-imidazole derivatives were synthesized and
their inhibitory potency against cycloxygenase was determined.
In synthesized compounds, 2-alkylthio derivatives are more potent
and selective than 2-sulfonylalkyl derivatives. Molecular modeling
showed that 2-sulfonylalkyl series interact with Arg120 in COX-1
active site and this interaction could increase the residence time
of ligand in the active site. In 2-alkylthio series, which exhibit
COX-2 selectivity, in addition to possibility of reorientation in the
COX-2 active site, an extra hydrogen bond with Tyr341 could in-
crease the residence time of ligands in the active site and conse-
quently augments COX-2 selectivity.
4.1.4. 2-(Methylsulfonyl)-5-(4-(methylsulfonyl)phenyl)-4-
phenyl-1H-imidazole (11a)
Yield, 68%; mp 218–221 °C; IR (KBr, cmÀ1): v 3230 (NH), 1285,
1137 (SO2); 1H NMR (500 MHz, DMSO-d6): d 3.22 (br s, 3H,
SO2CH3), 3.45 (s, 3H, SO2CH3), 7.31–7.57 (m, 5H, phenyl), 7.71 (d,
J = 8 Hz , 2H, H2,6-methylsulfonylphenyl), 7.87 and 7.97 (two d,
J = 8 Hz, 2H, H3,5-methylsulfonylphenyl), 14.25 and 14.27 (two br
s, 1H, NH). Anal. Calcd for C17H16N2O4S2: C, 54.24; H, 4.28; N,
7.44. Found: C, 54.46; H, 4.42; N, 7.24.
4.1.5. 4-(4-Fluorophenyl)-2-(methylsulfonyl)-5-(4-
(methylsulfonyl)phenyl)-1H-imidazole (11b)
Yield, 67%; mp 227–229 °C; IR (KBr, cmÀ1): v 3211 (NH), 1280,
1142 (SO2); 1H NMR (500 MHz, DMSO-d6): d 3.23 (br s, 3H,
SO2CH3), 3.44 (s, 3H, SO2CH3), 7.17–7.40 (m, 2H, H3,5-fluoro-
4. Experimental section
4.1. Chemistry
phenyl), 7.41–7.61 (br s, 2H,
J = 8.5 Hz, 2H, 2,6-methylsulfonylphenyl), 7.80–7.99 (m, 2H,
3,5-methylsulfonylphenyl), 14.28 (br s, 1H, NH). Anal. Calcd for
17H15FN2O4S2: C, 51.76; H, 3.83; N, 7.10. Found: C, 51.39; H,
H2,6-fluorophenyl), 7.70 (d,
H
1H NMR spectra were recorded on a 500 MHz Bruker spectrom-
eter using CDCl3 or DMSO-d6 as solvent. Chemical shifts (d) are re-
ported in ppm relative to tetramethylsilane (TMS) as internal
standard. Infrared spectra were acquired on a Nicolet Magna
550-FT spectrometer. IR spectra of solids were recorded in KBr
H
C
3.61; N, 7.34.
4.1.6. 4-(4-Chlorophenyl)-2-(methylsulfonyl)-5-(4-
(methylsulfonyl)phenyl)-1H-imidazole (11c)
and the absorption band was given in wave numbers
m .
in cmÀ1
Yield, 68%; mp 233–235 °C; IR (KBr, cmÀ1): v 3196 (NH), 1285,
1132 (SO2); 1H NMR (500 MHz, DMSO-d6): d 3.25 (br s, 3H,
SO2CH3), 3.44 (s, 3H, SO2CH3), 7.41–7.61 (m, 4H, chlorophenyl),
7.71 (d, J = 8 Hz, 2H, H2,6-methylsulfonylphenyl), 7.93 (d, J = 8 Hz,
2H, H3,5-methylsulfonylphenyl), 14.33 (br s, 1H, NH). Anal. Calcd
for C17H15ClN2O4S2: C, 49.69; H, 3.68; N, 6.82. Found: C, 49.83; H,
3.46; N, 6.69.
Elemental microanalyses were within 0.4 of the theoretical values
for C, H and N.
4.1.1. 5-(4-(Methylsulfonyl)phenyl)-2-(methylthio)-4-phenyl-
1H-imidazole (10a)
Yield, 78%; mp 121–124 °C; IR (KBr, cmÀ1): v 3298 (NH), 1280,
1137 (SO2); 1H NMR (500 MHz, DMSO-d6): d 2.63 (s, 3H, SCH3),
3.20 and 3.25 (two br s, 3H, SO2CH3), 7.26–7.52 (m, 5H, phenyl),
7.62 and 7.71 (two d, J = 8 Hz, 2H, H2,6-methylsulfonylphenyl),
7.81 and 7.90 (two d, J = 8 Hz, 2H, H3,5-methylsulfonylphenyl),
12.70 and 12.73 (two br s, 1H, NH). Anal. Calcd for C17H16N2O2S2:
C, 59.28; H, 4.68; N, 8.13. Found: C, 59.46; H, 4.41; N, 8.32.
4.1.7. 2-(Ethylthio)-5-(4-(methylsulfonyl)phenyl)-4-phenyl-1H-
imidazole (12a)
Yield, 78%; mp 157–158 °C; IR (KBr, cmÀ1): v 3247 (NH), 1296,
1142 (SO2); 1H NMR (500 MHz, DMSO-d6): d 1.33 (t, J = 7.5 Hz, 3H,

A. Assadieskandar et al. / Bioorg. Med. Chem. 21 (2013) 2355–2362
2361
SCH2CH3), 3.14 (q, J = 7.5 Hz, 2H, SCH2CH3), 3.20 and 3.25 (two br s,
3H, SO2CH3), 7.26–7.52 (m, 5H, phenyl), 7.63 and 7.71 (two d,
J = 8 Hz, 2H, H2,6-methylsulfonylphenyl), 7.81 and 7.91 (two d,
J = 8 Hz, 2H, H3,5-methylsulfonylphenyl), 12.74 and 12.76 (two br
s, 1H, NH). Anal. Calcd for C18H18N2O2S2: C, 60.31; H, 5.06; N,
7.81. Found: C, 60.58; H, 5.27; N, 7.67.
C18H17ClN2O4S2: C, 50.88; H, 4.03; N, 6.59. Found: C, 50.65; H,
4.21; N, 6.39.
4.2. Molecular modeling
The molecular geometry of all compounds was fully minimized
by MMFF94 force field using ChemBio3D Ultra 12.0 (Cambridge-
soft), setting the terminal condition as the RMS of potential energy
smaller than 0.0001 kcal ÅÀ1 molÀ1. The crystal structures of cyclo-
oxygenase-2 in complex with celecoxib (entery code 3LN1) and
cyclooxygenase-1 also complexed with celecoxib (entery code
3KK6) were retrieved from the Brookhaven Protein Data Bank
(http://www.rcsb.org/pdb/home/home.do). The FlexX program
interfaced with LeadIT 2.1.2 (BioSolveIT GmbH, Sankt Augustin,
Germany) was used to dock all compounds. FlexX is an automated
docking program, which considers ligand conformational flexibility
by an incremental fragment placing method.30 The active site for
docking was determined as all atoms within 7 Å radius of the
cocrystallized ligand. Validation of the docking procedure was
accomplished by energy minimization of celecoxib structure by
the above mentioned method and the optimized geometry struc-
ture was redocked in the active site of COX-2 and COX-1 which re-
sulted in predicted docking poses with RMSD 0.723 and 0.975,
respectively, within the best scored poses.
4.1.8. 2-(Ethylthio)-4-(4-fluorophenyl)-5-(4-(methylsulfonyl)
phenyl)-1H-imidazole (12b)
Yield, 73%; mp 82–84 °C; IR (KBr, cmÀ1): v 3277 (NH), 1306,
1137 (SO2); 1H NMR (500 MHz, DMSO-d6): d 1.33 (t, J = 7.5 Hz,
3H, SCH2CH3), 3.14 (q, J = 7.5 Hz, 2H, SCH2CH3), 3.19 and 3.25
(two s, 3H, SO2CH3), 7.19 and 7.30(two dd, J = 9 Hz, J = 9 Hz, 2H,
H3,5-fluorophenyl), 7.44–7.54 (m, 2H, H2,6-fluorophenyl), 7.61 and
7.69 (two d, J = 8.5 Hz, 2H, H2,6-methylsulfonylphenyl), 7.82 and
7.91 (two d, J = 8.5 Hz, 2H, H3,5-methylsulfonylphenyl), 12.70 and
12.73 (two br s, 1H, NH). Anal. Calcd for C18H17FN2O2S2: C, 57.43;
H, 4.55; N, 7.44. Found: C, 57.67; H, 4.36; N, 7.61.
4.1.9. 4-(4-Chlorophenyl)-2-(ethylthio)-5-(4-(methylsulfonyl)
phenyl)-1H-imidazole (12c)
Yield, 75%; mp 92–94 °C; IR (KBr, cmÀ1): v 3262 (NH), 1311,
1147 (SO2); 1H NMR (500 MHz, DMSO-d6): d 1.33 (t, J = 7 Hz, 3H,
SCH2CH3), 3.14 (q, J = 7 Hz, 2H, SCH2CH3), 3.20 and 3.26 (two s,
3H, SO2CH3), 7.41 and 7.44 (two d, J = 8.5 Hz, 2H, chlorophenyl),
7.48 and 7.52 (two d, J = 8.5 Hz, 2H, chlorophenyl), 7.63 and 7.70
(two d, J = 8.5 Hz, 2H, H2,6-methylsulfonylphenyl), 7.84 and 7.93
For evaluation of the ligands affinity toward docked receptor,
the HYDE assessment facility of LeadIT software was implemented
to report the free energy of binding (DG) and ligand efficiency of
(two d, J = 8.5 Hz, 2H,
H
3,5-methylsulfonylphenyl), 12.76 and
some of the best dock scored poses. HYDE is an empirical scoring
function, which assesses protein–ligand complex by considering
hydrogen bond interactions and also hydrophobic and desolvation
effects and provides estimation for the binding affinity.31,32
12.78 (two br s, 1H, NH). 1H NMR (500 MHz, CDCl3): d 1.43 (t,
J = 7 Hz, 3H, SCH2CH3), 3.08 (s, 3H, SO2CH3), 3.20 (q, J = 7 Hz, 2H,
SCH2CH3), 7.38 (br s, 4H, chlorophenyl), 7.73 (d, J = 7.5 Hz, 2H,
H2,6-methylsulfonylphenyl), 7.86 (d, J = 7.5 Hz, 2H, H3,5-meth-
ylsulfonylphenyl), 12.76 and 12.78 (two br s, 1H, NH) Anal. Calcd
for C18H17ClN2O2S2: C, 55.02; H, 4.36; N, 7.13. Found: C, 55.28; H,
4.43; N, 7.29.
4.3. Human whole blood assays for COX-2 and COX-1
The assays were carried out using previously described proce-
dures.33,34 Fresh blood was taken from human volunteers who
had no apparent inflammatory conditions and had not taken any
NSAIDs for at least 7 days prior to blood collection. For the COX-
4.1.10. 2-(Ethylsulfonyl)-5-(4-(methylsulfonyl)phenyl)-4-
phenyl-1H-imidazole (13a)
Yield, 67%; mp 248–250 °C; IR (KBr, cmÀ1): v 3180 (NH), 1291,
1142 (SO2); 1H NMR (500 MHz, DMSO-d6): d 1.27 (t, J = 7.5 Hz, 3H,
SO2CH2CH3), 3.22 (br s, 3H, SO2CH3), 3.52 (q, J = 7.5 Hz, 2H,
SO2CH2CH3), 7.26–7.59 (m, 5H, phenyl), 7.71 (d, J = 8 Hz, 2H,
H2,6-methylsulfonylphenyl), 7.82–7.03 (m, 2H, H3,5-methylsulfo-
nylphenyl), 14.27 (br s, 1H, NH). Anal. Calcd for C18H18N2O4S2: C,
55.37; H, 4.65; N, 7.17. Found: C, 55.62; H, 4.87; N, 7.34.
2 assay, heparinized blood samples (500
vehicle (2 L of DMSO) or test compound (2
at 37 °C for 15 min; then, blood samples were incubated in the
presence of 10 L of lipopolysaccharide (LPS; 100 g/mL in PBS)
at 37 °C for 24 h. After incubation, all blood samples were centri-
fuged (12,000g for 5 min); 100 L of plasma were mixed with
400 L of methanol to precipitate plasma proteins. The obtained
l
L) were incubated with
l
lL of a DMSO solution)
l
l
l
l
supernatants were assayed for PGE2 levels by an enzyme immuno-
assay kit according to the instructions of the manufacturer (Cay-
man Chemicals, Ann Harbor, MI, #500141). For the COX-1 assay,
4.1.11. 2-(Ethylsulfonyl)-4-(4-fluorophenyl)-5-(4-
(methylsulfonyl)phenyl)-1H-imidazole (13b)
Yield, 63%; mp 225–228 °C; IR (KBr, cmÀ1): v 3211 (NH), 1285,
1137 (SO2); 1H NMR (500 MHz, DMSO-d6): 1.26 (t, J = 7 Hz, 3H,
SO2CH2CH3), 3.28 (br s, 3H, SO2CH3), 3.52 (q, J = 7 Hz, 2H,
SO2CH2CH3), 7.17–7.41 (m, 2H, H3,5-fluorophenyl), 7.43–7.62 (m,
2H, H2,6-fluorophenyl), 7.70 (d, J = 8 Hz, 2H, H2,6-methylsulfonyl-
phenyl), 7.81–8.03 (m, 2H, H3,5-methylsulfonylphenyl), 14.29 (br
s, 1H, NH). Anal. Calcd for C18H17FN2O4S2: C, 52.93; H, 4.19; N,
6.86. Found: C, 52.76; H, 4.31; N, 6.58.
the aliquots of blood samples (500
ferred to siliconized microcentrifuge tubes and mixed with either
L of DMSO or a test compound. Following vortexing at 37 °C
for 1 h, the samples were centrifuged (12,000Âg, 5 min) and
100 L of serum supernatants were mixed with 400 L of metha-
lL) were immediately trans-
2
l
l
l
nol for protein precipitation. The obtained supernatants were as-
sayed for TxB2 levels by an enzyme immunoassay kit according
to the instructions of the manufacturer (Cayman, Ann Arbor, MI,
#519031). The reference molecules SC-560, celecoxib and Dup-
697 were analyzed under the same experimental conditions. The
IC50 values (the concentration of the test compound causing 50%
inhibition) were calculated from the concentration inhibition re-
sponse curves (duplicate determinations).
4.1.12. 4-(4-Chlorophenyl)-2-(ethylsulfonyl)-5-(4-
(methylsulfonyl)phenyl)-1H-imidazoleimidazole (13c)
Yield, 76%; mp 243–246 °C; IR (KBr, cmÀ1): v 3206 (NH), 1275,
1132 (SO2); 1H NMR (500 MHz, DMSO-d6): d 1.26 (t, J = 7.5 Hz, 3H,
SO2CH2CH3), 3.25 (br s, 3H, SO2CH3), 3.52 (q, J = 7.5 Hz, 2H,
SO2CH2CH3), 7.41–7.63 (m, 4H, chlorophenyl), 7.71 (d, J = 8 Hz
Acknowledgments
2 Hz, 2H, H2,6-methylsulfonylphenyl), 7.84–8.03 (m, 2H, H3,5
methylsulfonylphenyl), 14.35 (br s, 1H, NH). Anal. Calcd for
-
This research has been supported by Tehran University of Med-
ical Sciences & Health Services Grant No. 15098. The authors are

2362
A. Assadieskandar et al. / Bioorg. Med. Chem. 21 (2013) 2355–2362
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