Design, synthesis, and biological evaluation of substituted 3-alkylthio-4,5-diaryl-4H-1,2,4-triazoles as selective COX-2 inhibitors

Article abstract of DOI:10.1016/j.bmc.2005.11.029

A new type of 4,5-diaryl-4H-1,2,4-triazole, possessing C-3 thio and alkylthio (SH, SMe or SEt) substituents, was designed and synthesized for evaluation as selective cyclooxygenase-2 (COX-2) inhibitors with in vivo anti-inflammatory activity. The compound

Full text of DOI:10.1016/j.bmc.2005.11.029

Bioorganic & Medicinal Chemistry 14 (2006) 2507–2517
Design, synthesis, and biological evaluation of substituted
3-alkylthio-4,5-diaryl-4H-1,2,4-triazoles as
selective COX-2 inhibitors
Latifeh Navidpour,a Hamed Shafaroodi,b Khosrou Abdi,a Mohsen Amini,a
Mohammad H. Ghahremani,c Ahmad Reza Dehpourd and Abbas Shafieea,*
aDepartment of Medicinal Chemistry and Pharmaceutical Sciences Research Center, Faculty of Pharmacy,
Tehran University of Medical Sciences, Tehran 14174, Iran
bDepartment of Pharmacology, Tehran Medical Unit, Islamic Azad University, Tehran, Iran
cDepartment of Pharmacology and Toxicology and Pharmaceutical Sciences Research Center, Faculty of Pharmacy,
Tehran University of Medical Sciences, Tehran 14174, Iran
dDepartment of Pharmacology, Faculty of Medicine, Tehran University of Medical Sciences, Tehran 14174, Iran
Received 2 October 2005; revised 14 November 2005; accepted 15 November 2005
Available online 5 December 2005
Abstract—A new type of 4,5-diaryl-4H-1,2,4-triazole, possessing C-3 thio and alkylthio (SH, SMe or SEt) substituents, was designed
and synthesized for evaluation as selective cyclooxygenase-2 (COX-2) inhibitors with in vivo anti-inflammatory activity. The com-
pound, 3-ethylthio-5-(4-fluorophenyl)-4-(4-methylsulfonylphenyl)-4H-1,2,4-triazole (10d), exhibited a high in vitro selectivity (COX-
1 IC50 = 20.5 nM; COX-2 IC50 = 1.8 nM; SI = 11.39) relative to the reference drug celecoxib (COX-1 IC50 = 3.7 nM; COX-2
IC50 = 2.2 nM; SI = 1.68) and also showed good anti-inflammatory activity compared to celecoxib in a carrageenan-induced rat
paw edema assay.
Ó 2005 Elsevier Ltd. All rights reserved.
1. Introduction
suppress synthesis of prostacyclin, a potent vasodilator,
gastroprotectant, and platelet inhibitor, via inhibition of
endothelial COX-2. COX-2 selective inhibitors do
not inhibit production of thromboxane, a vasoconstric-
tor, and promoter of platelet aggregation, which is syn-
thesized in platelets by COX-1.6,7 Therefore, COX-2
inhibitors intrinsically lack anti-thrombotic activity,
and some cardiovascular liabilities have been associated
preclinically with them.8 Thus, there is still a need for
novel, selective, and potent COX-2 inhibitors with an
improved profile compared to current COX-2
inhibitors.3
A new generation of anti-inflammatory drugs, celecoxib
(CelebrexTM)1 and etoricoxib (ArcoxiaÒ),2 is being pre-
scribed to treat acute or chronic inflammation by pro-
viding symptomatic pain relief.3
A
selective
cyclooxygenase-2 (COX-2) inhibitor allows the desired
synthesis of cytoprotective prostaglandins, in conjunc-
tion with a simultaneous inhibition of proinflammatory
prostaglandin synthesis, thereby reducing dyspepsia and
ulceration.4 However, emerging evidence suggests that
adverse reactions such as GI irritations or ulceration
and renal liabilities are associated with prolonged use
of COX-2 selective inhibitors. The adverse reactions
have been attributed, at least in part, to COX-1 inhibi-
tion occurring with long-term exposure or at higher
doses.5 COX-2 selective inhibitors are also known to
Diarylheterocycles, and other central ring pharmaco-
phore templates, have been extensively studied as cyclo-
oxygenase inhibitors. All these tricyclic molecules
possess 1,2-diaryl substitution on a central four-, five-,
or six-membered ring system such as cyclobutenone
(1), pyrazole (2), 2-(5H)-furanone (3), isoxazole (4), or
pyridine (5), respectively (Fig. 1).1,2,9–11
Keywords: Cyclooxygenase-2 inhibitor; 1,2,4-Triazoles; Alkylthio;
Celecoxib.
*
Structure–activity relationship (SAR) studies have
shown that for optimum COX-2 selectivity and
Corresponding author. Tel.: +98 21 66406757; fax: +98 21
66461178; e-mail: ashafiee@ams.ac.ir
0968-0896/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2005.11.029

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L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
inhibitory potency, a SO2Me or SO2NH2 substituent at
H3C
MeO2S
the para-position of a phenyl ring, and the presence of
a p-F substituent on a non-sulfonyl vicinal phenyl ring
improve in vivo activity.12 Recently, a novel class of 6-
alkylthio-substituted six-membered lactone (pyrane-2-
one) rings (6) has been designed and exhibited very
good in vitro COX-2 inhibitory potency and selectivi-
ty.13 Furthermore, differently substituted 1,2,4-tria-
zole-3-thione derivatives exhibited anti-inflammatory
activity.14–18 Considering these results and as part of
our ongoing program to design novel selective
COX-2 inhibitors, we describe herein the design, syn-
thesis, and biological evaluation of a novel diverse
group of alkylthio-substituted-5-(4-methylsulfonylphe-
nyl)-4-phenyl (or cyclohexyl) triazoles and alkylthio-
substituted-4-(4-methylsulfonylphenyl)-5-phenyltriazoles as
selective COX-2 inhibitors with anti-inflammatory
activity.
Me
Me
CF3
N
N
O
H2NO2S
1
2 (Celecoxib)
O
N
O
O
CH3
MeO2S
MeO2S
H2NO2S
MeO2S
4 (Valdecoxib)
3 (Rofecoxib)
O
O
Cl
N
2. Chemistry
SEt
Me
The synthetic reactions used for the synthesis of 3-alkyl-
thio-5-(4-methylsulfonylphenyl)-4-phenyl (or cyclohexyl)
triazoles (4a–n) and 3-alkylthio-4-(4-methylsulfonylphe-
nyl)-5-phenyltriazoles (10a–f) are outlined in Schemes 1
and 2.
5 (Etoricoxib)
6
Figure 1. Representative examples of selective tricyclic COX-2
inhibitors.
H3CO2S
CONHNH2
1
a
a
O
S
O
S
H3CO2S
H3CO2S
CNHNHCNH
X
CNHNHCNH
2a-f
2g
b
b
H3CO2S
H3CO2S
N
N
NH
NH
N
N
S
S
X
3a-f
3g
c
c
H3CO2S
H3CO2S
N
N
N
N
N
N
SR
SR
X
4a-l
4m-n
X: H, F, Cl, Br, OMe, Me
R: Me, Et
Scheme 1. Reagents and conditions: (a) R-N@C@S, EtOH, rt, overnight; (b) Na2CO3 aq reflux, overnight; (c) RI, KOH, rt, overnight.

L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
2509
O
C
a
H3CO2S
NH2
X
Cl
+
5
6a-c
X
O
C
H3CO2S
b
X
H3CO2S
NH
N
H
8a-c
N
N
NH2
7a-c
X
X
N
c
d
NH
N
N
N
S
SR
H3CO2S
9a-c
H3CO2S
10a-f
X: H, F, Me
R: Me, Et
Scheme 2. Reagents and conditions: (a) Et3N, THF, 0 °C to rt, 24 h; (b) PCl5, benzene, reflux, 3 h; then NH2NH2 (dry), THF, rt, overnight; (c) 1,1Õ-
thiocarbonyldiimidazole, THF, rt, 18 h; (d) RI, KOH, rt, overnight.
Starting from 4-methylsulfonylbenzoic acid hydrazide 1,
3-alkylthio-5-(4-methylsulfonylphenyl)-4-aryl (or cyclo-
hexyl)-4H-1,2,4-triazole (4a–n) were prepared. The reac-
tion of corresponding hydrazide 1 with respective aryl
(or cyclohexyl) isothiocyanates yielded 2a–g, which were
cyclized in saturated sodium carbonate solution to 3a–g.
Subsequent alkylation of 3a–g using alkyl iodide in ba-
sic media afforded the title compounds 4a–n (see
Scheme 1).19
eral, for these compounds, COX-2 selectivity and poten-
cy were dependent upon steric properties of C-3 thio or
alkylthio substituent on the central triazole ring and
electronic properties of the C-4 phenyl ring substituent.
For bulkier C-4-substituted compounds 3c–f and 4e–l,
the size of the C-3 substituent had a significant effect
on COX-2 inhibitory potency (SH > SMe > SEt) and
COX-2 selectivity. Decreasing the size led to an increase
in COX-2 potency and selectivity (see Table 1).
The target 3-alkylthio-4-(4-methylsulfonylphenyl)-5-
phenyl-4H-1,2,4-triazoles (10a–f) were prepared start-
ing with the condensation of 4-methylsulfonylaniline
6 with respective benzoyl chloride 6 in the presence
of Et3N to produce N-(4-methylsulfonylphenyl)-4-
substituted benzamides (7a–c). Subsequent reaction of
7a–c with PCl5, followed by reaction with dry hydra-
zine, yielded N-(4-methylsulfonylphenyl)aryl carbohy-
drazonamides (8a–c) that undergoes a ring closure
using 1,1Õ-thiocarbonyldiimidazole and subsequent
alkylation to afford 10a–f (see Scheme 2).20,21 3-Alkyl-
thio-1,2,4-triazoles (11b–c) were prepared according to
the literature.22
In addition, among the different substituents on C-4 of
the phenyl ring, the best substituent was C-4 methyl.
Thus, the most potent and selective COX-2 inhibitor
was 3f, (COX-2 IC50 = 0.6 nM; COX-1 IC50 = 3.1 nM;
S.I. = 5.17) relative to reference drug celecoxib (COX-2
IC50 = 2.2 nM; COX-1 IC50 = 3.7 nM; S.I. = 1.68). This
phenomenon is not particularly surprising or unexpect-
ed since it is also found in celecoxib at the 5-position
of the central pyrazole ring.
In the cyclohexyl subgroup (3g, 4m, and 4n) also
decreasing the size of the thio group significantly in-
creased COX-2 selectivity with 3g, showing a potent
inhibition of COX-2 and good selectivity index (COX-
2 IC50 = 0.6 nM; COX-1 IC50 = 2.4 nM; S.I. = 4.0) rela-
tive to reference compound celecoxib (COX-2
IC50 = 2.2 nM; COX-1 IC50 = 3.7 nM; S.I. = 1.68).
3. Results and discussion
The effect of thio and alkylthio substituents on the central
five-membered triazole ring on COX-2 selectivity and
potency was determined by colorimetric COX (ovine)
inhibitor screening assay (Tables 1–3).23–25 As an initial
in vitro screening, compounds were assayed against
COX-2 at 5 and 10 nM, and COX-1 at 10 and 50 nM
(Tables 1 and 3). For the more potent and selective
COX-2 inhibitors (relative to reference compound, cele-
coxib), the IC50 values were then determined (Table 2).
3-Alkylthio-4-(4-methylsulfonylphenyl)-5-phenyl-4H-1,2,
4-triazoles (10b–e) exhibited weak COX-1 inhibition (3.8–
20.5 nM range) with good COX-2 inhibition in the 1.6–
2.5 nM range. In this series, 10d was the most potent
and selective COX-2 inhibitor (COX-2 IC50 = 1.8 nM;
COX-1 IC50 = 20.5 nM; S.I. = 11.39) relative to reference
compound celecoxib (COX-2 IC50 = 2.2 nM; COX-1
IC50 = 3.7 nM; S.I. = 1.68).
In vitro enzyme inhibition studies for 3a–f and 4a–l
showed weak to good COX-2 inhibitory activity. In gen-
Further studies for 3-alkylthio-1,2,4-triazole subgroup
(11a–c) showed good COX-1 inhibition with moderate

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L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
Table 1. In vitro inhibition of COX-1 and COX-2 by 3a–g, 4a–n, and 10a–f
N
N
SR3
R1
N
R2
Compound
R1
R2
R3
Inhibition
of COX-2
at 5 nma
Inhibition
of COX-2
at 10 nma
Inhibition
of COX-1
at 10 nma
Inhibition
of COX-1
at 50 nma
Ratio
COX-2/COX-1
at 10 nm
3a
3b
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
4-CH3SO2
H
Phenyl
4-F-phenyl
H
0.74
0.77
0.99
0.91
1.12
0.81
0.90
0.82
0.86
0.79
0.91
0.85
0.48
0.99
0.54
1.00
1.47
1.24
0.43
1.27
1.11
1.11
1.14
0.93
0.98
1.37
1.17
0.91
1.18
1.11
1.09
0.99
1.20
1.15
1.05
0.88
0.98
0.85
1.00
0.93
0.65
0.99
0.64
1.02
0.83
1.37
0.65
1.38
1.35
1.17
1.17
1.10
1.01
1.24
0.91
1.00
0.82
0.81
0.99
0.87
0.86
0.78
0.94
0.95
0.83
0.83
0.98
0.91
0.92
0.93
0.97
0.80
0.95
0.96
0.96
0.96
0.79
1.04
0.98
0.85
0.62
0.88
0.86
0.90
0.87
1.06
1.15
0.90
0.93
0.96
0.98
0.99
0.87
0.97
0.97
0.98
1.13
0.97
0.91
1.00
0.97
1.05
0.89
1.08
1.14
1.11
1.11
1.06
0.86
0.87
0.98
1.01
1.44
1.36
1.09
1.13
1.40
1.58
1.12
0.92
1.17
1.03
1.02
1.02
0.71
1.06
0.66
1.27
0.87
1.43
0.68
1.44
1.71
1.13
1.19
1.30
1.64
1.40
1.06
1.11
H
3c
3d
4-Cl-phenyl
4-Br-phenyl
H
H
3e
4-OCH3-phenyl
4-CH3-phenyl
Cyclohexyl
H
3f
3g
H
H
CH3
4a
4b
Phenyl
Phenyl
CH2CH3
CH3
4c
4d
4-F-phenyl
4-F-phenyl
CH2CH3
CH3
4e
4-Cl-phenyl
4-Cl-phenyl
4f
4g
CH2CH3
CH3
4-Br-phenyl
4-Br-phenyl
4h
4i
CH2CH3
CH3
4-OCH3-phenyl
4-OCH3-phenyl
4-CH3-phenyl
4-CH3-phenyl
Cyclohexyl
4j
4k
CH2CH3
CH3
4l
CH2CH3
CH3
4m
4n
Cyclohexyl
CH2CH3
CH3
10a
10b
10c
10d
10e
10f
Celecoxib
4-CH3SO2-phenyl
4-CH3SO2-phenyl
4-CH3SO2-phenyl
4-CH3SO2-phenyl
4-CH3SO2-phenyl
4-CH3SO2-phenyl
H
CH2CH3
CH3
F
F
CH3
CH2CH3
CH3
CH2CH3
CH3
a Values are means of two determinations and deviation from the mean is <10% of the mean value.
Table 2. IC50 values and anti-inflammatory activities of some of 3, 4, and 10
Compound
IC50 (nM)
Selectivity index (COX-1/COX-2)
AI activity
COX-1a
COX-2a
% inhibition at 3 hb
% inhibition at 5 hb
3a
3b
3.5
3.7
3.1
2.4
3.7
3.3
4.7
6.4
3.7
3.8
12.1
20.5
4.1
3.7
1.6
2.0
0.6
0.6
2.3
2.2
2.1
2.3
1.9
1.6
2.0
1.8
2.5
2.2
2.19
1.85
5.17
4.0
59.1 2.7
38.7 2.8
25.8 10.6
51.5 4.6
70.4 10.4
52.8 6.2
64.0 4.6
32.4 4.6
44.0 8.8
76.3 8.2
53.8 9.3
66.6 5.8
66.1 3.3
70.5 4.7
13.3 6.2
28.8 4.4
20.8 7.9
36.7 2.9
63.3 3.3
51.7 5.5
34.0 6.6
32.4 4.6
39.7 7.8
71.1 10.6
52.2 7.7
43.4 4.9
53.3 3.3
50.0 2.5
3f
3g
4c
1.61
1.50
2.24
2.78
1.95
2.37
6.05
11.39
1.64
1.68
4i
4k
4m
4n
10b
10c
10d
10e
Celecoxib
a Values are means of two determinations and deviation from the mean is <10% of the mean value.
b Inhibitory activity on carrageenan-induced rat paw edema. The results are expressed as means SEM (n = 4–6) following a 50 mg/kg oral dose of
the test compound.
COX-2 inhibition relative to the reference drug celecox-
ib (Table 3). These compounds similar to classical NSA-
IDs inhibit both isoforms non-selectively.
The orientation of the highly potent and selective COX-
inhibitors, 4-(4-methylphenyl)-5-(4-methylsulfonyl-
phenyl)-2,4-dihydro-3H-1,2,4-triazole-3-thione (3f) and
2

L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
2511
Table 3. In vitro inhibition of COX-1 and COX-2, and anti-inflammatory activity by 11a–c
N
N
SR
N
H
Compound
R
Inhibition of Inhibition of Inhibition of Inhibition of Ratio
COX-2 at
5 nma
AI activity
% inhibition at 3 hb % inhibition at 5 hb
COX-2 at
10 nma
COX-1 at
10 nma
COX-1 at
50 nma
COX-2/COX-1
at 10 nm
11a
11b
H
CH3
0.80
0.72
0.81
0.76
0.80
1.00
0.91
0.85
0.93
0.90
0.92
0.89
0.95
1.01
0.89
0.89
0.86
1.11
57.0 6.9
46.5 5.6
59.3 8.4
70.5 4.7
34.6 11.8
24.1 15.2
49.3 9.5
50.0 2.5
11c
Celecoxib
CH2CH3 0.76
0.91
a Values are means of two determinations and deviation from the mean is <10% of the mean value.
b Inhibitory activity on carrageenan-induced rat paw edema. The results are expressed as means SEM (n = 4–6) following a 50 mg/kg oral dose of
the test compound.
3-ethylthio-5-(4-fluorophenyl)-4-(4-methylsulfonylphe-
nyl)-4H-1,2,4-triazole (10d), in the COX-2 active site
was examined by a docking experiment.26 This study
showed that 3f binds in the center of the primary bind-
ing site of COX-2 with the SO2Me moiety interacting
with the secondary pocket amino acid residues Phe518
,
Gln192, Arg513, Leu352, Ser353, and Val523. One of the
O-atoms of the SO2Me substituent forms hydrogen
bonds with the amine hydrogen (guanidine group) of
90
Arg513 (2.5 A) and His (2.7 A), and the other oxygen
˚
˚
forms hydrogen bond with Phe518 (2.9 A) as shown in
˚
Figure 2. Interestingly, the C-3 C@S sulfur atom is ori-
ented toward the hydrophobic region formed by Ala527
,
Leu359, Leu531, Met113, Val116, and Val349. This C@S
sulfur atom is positioned about 4.5 A from the NH2
˚
(guanidino) group of the polar amino acid Arg120 and
5.9 A from the OH group of Tyr355. These interactions
˚
Figure 3. Docking of 10d (ball and stick) in the active site of murine
COX-2 (line and stick).
may disrupt the salt bridge between His90, Arg120
,
Tyr355, and Glu524 at the mouth of the COX-2 active
site. A similar docking study of 10d showed that it binds
in the primary binding site such that one of the O-atoms
is located in a hydrophobic region formed by Ala527
,
of the p-SO2Me substituent forms hydrogen bonds with
Leu359, Leu531, Met113, Val116, and Val349, with the S-
atom forming a weak hydrogen bond with the NH2
the amine hydrogen of Arg513 (1.8 A) and His (2.5 A),
90
˚
˚
and the other oxygen forms hydrogen bond with Phe518
˚
(3.6 A) as shown in Figure 3. The C-3 EtS-substituent
(guanidino) group of the polar amino acid Arg120
355
(3.3 A) and with the OH group of Tyr (2.8 A).
˚
˚
These observations provide a good explanation for the
potent and selective inhibitory activity of 3f and 10d.
In vivo pharmacological evaluation of 3, 4, and 10 was
carried out to assess their potential anti-inflammatory
activity.27 Initial compound selection was based on the
in vitro COX-1/COX-2 enzyme inhibition data
obtained. Qualitative structure–activity relationship
data, acquired using the anti-inflammatory rat paw
edema assay, showed that this group of C-3 thio- and
alkylthio-substituted
4,5-diaryl-4H-1,2,4-triazoles
exhibits anti-inflammatory activity with moderate to
good activity range (13–76% inhibition) (Table 2).
In the C-3 thio series (3a, 3b, 3f, and 3g), 4-cyclohexyl-5-
(4-methylsulfonylphenyl)-2,4-dihydro-3H-1,2,4-triazole-
3-thione (3g) was the most active anti-inflammatory
agent (51 and 37% reduction in inflammation at 3
and 5 h postdrug administration, respectively) for a
Figure 2. Docking of 3f (ball and stick) in the active site of murine
COX-2 (line and stick).

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L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
50 mg/kg oral dose as compared to celecoxib (70 and
50% reduction in inflammation at 3 and 5 h postdrug
administration, respectively). In this series of com-
pounds, 3b and 3f exhibit moderate anti-inflammatory
activity. It is possible that these compounds are more
susceptible to metabolic inactivation when administered
by the oral route which may result in a lower amount of
the drug localization at the inflammation site.
anti-inflammatory screens, were purchased from Pasteur
Institute (Karaj, Iran), and experiments were carried out
using protocols approved by the Ethics Committee of
Tehran University of Medical Sciences.
5.1. General procedure for the preparation of 1-(4-
methylsulfonylbenzoyl)-4-aryl (or cyclohexyl)
thiosemicarbazides (2a–g)
The C-3 alkylthio-substituted subgroup of 4 (4c, 4i, 4k,
4m, and 4n) reduced inflammation by 32–70% at
different time intervals as shown in Table 2. 4-(4-Fluor-
ophenyl)-5-(4-methylsulfonylphenyl)-3-methylthio-4H-1,2,
4-triazole (4c) was the most potent anti-inflammatory
agent in this series, producing a 70 and 63% reduction
in inflammation at 3 and 5 h postdrug administration,
respectively.
To a solution of 4-methylsulfonylbenzoic acid hydrazide
1 (320 mg, 1.5 mmol) in ethanol (5 mL) was added
respective isothiocyanate (1.5 mmol) and the mixture
was stirred at room temperature for 24 h. The precipi-
tate was filtered and crystallized from ethanol to give
the title compounds 2a–g as white solids.
5.1.1. 4-Phenyl-1-(4-methylsulfonylbenzoyl)thiosemicar-
bazide (2a). Yield, 86%; mp 201–203 °C; IR (KBr):
3-Alkylthio-4-(4-methylsulfonylphenyl)-5-phenyl-4H-1,2,
4-triazoles (10b–e) exhibit good anti-inflammatory activ-
ity. In this series, 10b was the most active anti-inflamma-
tory agent (76 and 71% reduction in inflammation at 3
and 5 h postdrug administration, respectively).
3307, 3261 (NH), 1681 (CO), 1293, 1146 (SO2) cmꢀ1
;
1H NMR (CDCl3 + DMSO-d6) d 10.52 (br s, 1H), 9.58
(br s, 1H), 8.22 (d, J = 8.5 Hz, 2H), 7.90 (d,
J = 8.5 Hz, 2H), 7.75–7.12 (m, 5H), 3.11 (s, 3H); MS
m/z (%) 349 (27), 331 (29), 313 (35), 298 (47), 242 (45),
202 (25), 173 (100), 129 (19), 75 (10), 49 (14). Anal.
Calcd for C15H15N3O3S2: C, 51.56; H, 4.33; N, 12.03.
Found: C, 51.41; H, 4.45; N, 11.90.
The 3-thio and alkylthio-1,2,4-triazole group of com-
pounds (11a–c) reduced inflammation by 24–59% at
different time intervals as shown in Table 3.
5.1.2. 4-(4-Fluorophenyl)-1-(4-methylsulfonylbenzoyl)thi-
osemicarbazide (2b). Yield, 91%; mp 200–202 °C; IR
(KBr): 3276, 3257 (NH), 1668 (CO), 1299, 1144 (SO2)
4. Conclusions
cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 10.55 (br s,
;
A new class of 3-thio and alkylthio-4,5-diaryl-4H-1,2,4-
triazoles was designed to develop further structure–ac-
tivity relationship data. The results of this investigation
show: (i) a C-3 SR substituent (10d) in this class of
diarylheterocycles provides potent and selective inhibi-
tion of the COX-2 isozyme, (ii) molecular modeling
studies indicate that the SO2Me moiety inserts deep into
the COX-2 secondary pocket and the C-3 SR sulfur
atom forms a weak hydrogen bond with NH2 atom of
Arg120, and (iii) these C-3 alkylthio compounds could
serve as useful probes to study the function and catalytic
activity of the COX-2 isozyme.
1H), 9.56 (br s, 1H), 8.22 (d, J = 8.5 Hz, 2H), 7.90 (d,
J = 8.5 Hz, 2H), 7.52 (dd, J = 5.1, 9.0 Hz, 2H), 7.02 (t,
J = 8.4 Hz, 2H), 3.11 (s, 3H); MS m/z (%) 367 (5), 348
(100), 333 (75), 304 (16), 256 (33), 215 (15), 183 (11).
Anal. Calcd for C15H14FN3O3S2: C, 49.03; H, 3.84; N,
11.44. Found: C, 48.88; H, 3.95; N, 11.52.
5.1.3.
4-(4-Chlorophenyl)-1-(4-methylsulfonylbenzoyl)-
thiosemicarbazide (2c). Yield, 87%; mp 196–198 °C; IR
(KBr): 3310, 3239, 3165 (NH), 1663 (CO), 1298, 1145
(SO2) cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 10.72
;
(br s, 1H), 9.65 (br s, 1H), 8.20 (d, J = 8.5 Hz, 2H),
8.01 (d, J = 8.5 Hz, 2H), 7.59 (d, J = 8.7 Hz, 2H), 7.29
(d, J = 8.7 Hz, 2H), 3.11 (s, 1H); MS m/z (%) 383 (10),
365 (78), 349 (100), 336 (35), 256 (42), 214 (18), 183
(10), 169 (20). Anal. Calcd for C15H14ClN3O3S2: C,
46.93; H, 3.68; N, 10.95. Found: C, 46.85; H, 3.82; N,
11.08.
5. Experimental
Melting points were determined with a Reichert-Jung
hot-stage microscope and are uncorrected. Infrared
spectra were recorded on a Nicolet Magna 550-FT spec-
1
trometer. H NMR (400 MHz) spectra were measured
5.1.4.
4-(4-Bromophenyl)-1-(4-methylsulfonylbenzoyl)-
on a Varian Unity plus 400 spectrometer in CDCl3 or
DMSO-d6 with TMS as the internal standard, where J
(coupling constant) values are estimated in Hertz. Spin
multiples are given as s (singlet), d (doublet), dd (dou-
blet of doublets), t (triplet), q (quartet), m (multiplet),
and br (broad). Mass spectra were obtained on a Finn-
igan Mat TSQ-70 spectrometer. Elemental microanaly-
ses were carried out with a Perkin-Elmer 240-C
apparatus and were within 0.4% of the theoretical val-
ues for C, H, and N. All solvents and reagents were pur-
chased from the Fluka, Aldrich or Merck Chemical
Company. Male Sprague–Dawley rats, used in the
thiosemicarbazide (2d). Yield, 88%; mp 185–187 °C; IR
(KBr): 3320, 3309 (NH), 1685 (CO), 1292, 1148 (SO2)
cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 10.22 (br s,
;
1H), 9.35 (br s, 1H), 8.18 (d, J = 8.5 Hz, 2H), 8.05 (d,
J = 8.5 Hz, 2H), 7.45 (d, J = 8.7 Hz, 2H), 7.12 (d,
J = 8.7 Hz, 2H), 3.12 (s, 1H). Anal. Calcd for
C15H14BrN3O3S2: C, 42.06; H, 3.29; N, 9.81. Found:
C, 41.95; H, 3.39; N, 9.96.
5.1.5. 4-(4-Methoxyphenyl)-1-(4-methylsulfonylbenzoyl)thi-
osemicarbazide (2e). Yield, 95%; mp 195–197 °C; IR
(KBr): 3328, 3247, 3177 (NH), 1664 (CO), 1297, 1144

L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
2513
(SO2) cmꢀ1; 1H NMR (CDCl3 + DMSO-d6) d 10.72 (br s,
1H), 9.62 (br s, 1H), 8.20 (d, J = 8.4 Hz, 2H), 8.01 (d,
J = 8.4 Hz, 2H), 7.39 (d, J = 8.8 Hz, 2H), 6.86 (d,
J = 8.8 Hz, 2H), 3.80 (s, 3H), 3.11 (s, 3H); MS m/z (%)
379 (12), 361 (38), 328 (55), 313 (10), 256 (32), 239 (52),
213 (42), 183 (100), 165 (62), 150 (41), 122 (10). Anal.
Calcd for C16H17N3O4S2: C, 50.64; H, 4.52; N, 11.0.
Found: C, 50.52; H, 4.60; N, 11.12.
1H), 7.89 (d, J = 8.2 Hz, 2H), 7.82–7.25 (m, 6H), 3.08
(s, 3H). Anal. Cacld for C15H12ClN3O2S2: C, 49.24; H,
3.31; N, 11.49. Found: C, 49.12; H, 3.42; N, 11.66.
5.2.4. 4-(4-Bromophenyl)-5-(4-methylsulfonylphenyl)-2,4-
dihydro-3H-1,2,4-triazole-3-thione (3d). Yield, 90%; mp
266–268 °C; IR (KBr): 3365 (NH), 1312, 1148 (SO2)
cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 14.15 (br s,
;
1H), 7.89 (d, J = 8.2 Hz, 2H), 7.65 (d, J = 8.2 Hz, 2H),
7.51 (d, J = 8.2 Hz, 2H), 7.25 (d, J = 8.2 Hz, 2H), 3.07
(s, 3H). Anal. Calcd for C15H12BrN3O2S2: C, 43.91; H,
2.95; N, 10.24. Found: C, 43.80; H, 3.06; N, 10.36.
5.1.6. 4-(4-Methylphenyl)-1-(4-methylsulfonylbenzoyl)-
thiosemicarbazide (2f). Yield, 92%; mp 193–195 °C; IR
(KBr): 3306, 3241 (NH), 1682 (CO), 1294, 1143 (SO2)
cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 10.58 (br s,
;
1H), 9.65 (br s, 1H), 8.21 (d, J = 8.4 Hz, 2H), 8.01 (d,
J = 8.4 Hz, 2H), 7.41 (d, J = 8.2 Hz, 2H), 7.13 (d,
J = 8.2 Hz, 2H), 3.11 (s, 3H), 2.33 (s, 3H); MS m/z (%)
363 (5), 338 (20), 321 (60), 306 (52), 276 (78), 248 (48),
208 (32), 199 (42), 170 (100), 140 (45). Anal. Calcd for
C16H17N3O3S2: C, 52.87; H, 4.71; N, 11.56. Found: C,
52.99; H, 4.55; N, 11.44.
5.2.5. 4-(4-Methoxyphenyl)-5-(4-methylsulfonylphenyl)-
2,4-dihydro-3H-1,2,4-triazole-3-thione (3e). Yield, 85%;
mp 296–298 °C; IR (KBr): 3328 (NH), 1328, 1171
(SO2) cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 14.25
;
(br s, 1H), 7.83 (d, J = 8.5 Hz, 2H), 7.56 (d,
J = 8.5 Hz, 2H), 7.22 (d, J = 8.4 Hz, 2H), 7.01 (d,
J = 8.4 Hz, 2H), 3.87 (s, 3H), 3.04 (s, 3H). Anal. Calcd
for C16H15N3O3S2: C, 53.17; H, 4.18; N, 11.63. Found:
C, 53.10; H, 4.29; N, 11.80.
5.1.7. 4-Cyclohexyl-1-(4-methylsulfonylbenzoyl)thiosemi-
carbazide (2g). Yield, 65%; mp 200–202 °C; IR (KBr):
3419, 3320, 3244 (NH), 1692 (CO), 1293, 1149 (SO2)
5.2.6.
4-(4-Methylphenyl)-5-(4-methylsulfonylphenyl)-
cmꢀ1
;
1H NMR (CDCl3 + DMSO-d6) d 10.55 (br s,
2,4-dihydro-3H-1,2,4-triazole-3-thione (3f). Yield, 90%;
mp 291–292 °C; IR (KBr): 3393 (NH), 1316, 1150
1H), 9.85 (br s, 1H), 8.18 (d, J = 8.5 Hz, 2H), 8.02 (d,
J = 8.5 Hz, 2H), 4.25 (m, 1H), 3.11 (s, 3H), 2.60–1.20
(m, 10H). Anal. Calcd for C15H21N3O3S2: C, 50.68; H,
5.95; N, 11.82. Found: C, 50.82; H, 6.05; N, 11.69.
(SO2) cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 14.05
;
(br s, 1H), 7.85 (d, J = 8.4 Hz, 2H), 7.54 (d,
J = 8.4 Hz, 2H), 7.26 (d, J = 8.2 Hz, 2H), 7.10 (d,
J = 8.2 Hz, 2H), 3.05 (s, 3H), 2.44 (s, 3H). Anal. Calcd
for C16H15N3O2S2: C, 55.63; H, 4.38; N, 12.16. Found:
C, 55.85; H, 4.25; N, 12.26.
5.2. General procedure for the preparation of 5-(4-meth-
ylsulfonylphenyl)-4-aryl (or cyclohexyl)-2,4-dihydro-3H-
1,2,4-triazol-3-thiones (3a–g)
5.2.7.
4-Cyclohexyl-5-(4-methylsulfonylphenyl)-2,4-di-
A stirring mixture of compounds 2a–g (1.5 mmol) and
saturated aqueous sodium carbonate solution (15 mL)
was refluxed overnight. After cooling, the solution
was acidified with hydrochloric acid and the precipi-
tate was filtered and rinsed with water, and then crys-
tallized from methanol to give the title compounds
3a–g.
hydro-3H-1,2,4-triazole-3-thione (3g). Yield, 76%; mp
301–303 °C; IR (KBr): 3434 (NH), 1315, 1152 (SO2)
cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 13.75 (br s,
;
1H), 8.09 (d, J = 8.3 Hz, 2H), 7.70 (d, J = 8.4 Hz, 2H),
4.25 (m, 1H), 3.15 (s, 3H), 2.55–1.25 (m, 10H). Anal.
Calcd for C15H19N3O2S2: C, 53.39; H, 5.67; N, 12.45.
Found: C, 53.48; H, 5.82; N, 12.53.
5.2.1. 5-(4-Methylsulfonylphenyl)-4-phenyl-2,4-dihydro-
3H-1,2,4-triazole-3-thione (3a). Yield, 97%; mp 290–
5.3. General procedure for the preparation of 3-alkylthio-
5-(4-methylsulfonylphenyl)-4-aryl (or cyclohexyl)-4H-
1,2,4-triazoles (4a–n)
292 °C; IR (KBr): 3381 (NH), 1317, 1148 (SO2) cmꢀ1
;
1H NMR (CDCl3 + DMSO-d6) d 13.98 (br s, 1H), 7.85
(d, J = 8.2 Hz, 2H), 7.78–7.18 (m, 7H), 3.05 (s, 3H).
Anal. Calcd for C15H13N3O2S2: C, 54.36; H, 3.95; N,
12.68. Found: C, 54.22; H, 4.09; N, 12.61.
To a stirring solution of compounds3a–g (0.5 mmol)
and sodium hydroxide (0.5 mmol) in ethanol (5 mL)
was added alkyl iodide (0.5 mL) and the mixture
was stirred overnight. The volatiles were evaporated.
Flash chromatography (MeOH/CHCl3, 1:20) and
crystallization from n-butanol gave the title com-
pounds 4a–n.
5.2.2. 4-(4-Fluorophenyl)-5-(4-methylsulfonylphenyl)-2,4-
dihydro-3H-1,2,4-triazole-3-thione (3b). Yield, 94%; mp
296–298 °C; IR (KBr): 3323 (NH), 1327, 1153 (SO2)
cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 14.05 (br s,
;
1H), 7.87 (d, J = 8.2 Hz, 2H), 7.82–7.15 (m, 6H), 3.08
(s, 3H). Anal. Calcd for C15H12FN3O2S2: C, 51.56; H,
3.46; F, 5.44; N, 12.03. Found: C, 51.44; H, 5.29; N,
11.95.
5.3.1. 5-(4-Methylsulfonylphenyl)-3-methylthio-4-phenyl-
4H-1,2,4-triazole (4a). Yield, 83%; mp 215–216 °C; IR
(KBr): 1311, 1150 (SO2) cmꢀ1 1H NMR (CDCl3) d
;
7.85 (d, J = 8.4 Hz, 2H), 7.63 (d, J = 8.4 Hz, 2H),
7.59–7.51 (m, 3H), 7.22–7.29 (m, 2H), 3.04 (s, 3H),
2.75 (s, 3H); MS m/z 345 (28), 330 (58), 266 (5), 251
(100), 218 (9), 192 (18), 164 (22), 135 (12), 108 (27).
Anal. Calcd for C16H15N3O2S2: C, 55.63; H, 4.38; N,
12.16. Found: C, 55.78; H, 4.49; N, 12.03.
5.2.3. 4-(4-Chlorophenyl)-5-(4-methylsulfonylphenyl)-2,4-
dihydro-3H-1,2,4-triazole-3-thione (3c). Yield, 95%; mp
263–265 °C; IR (KBr): 3398 (NH), 1317, 1153 (SO2)
cmꢀ1 1H NMR (CDCl3 + DMSO-d6) d 14.15 (br s,
;

2514
L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
5.3.2. 3-Ethylthio-5-(4-methylsulfonylphenyl)-4-phenyl-
4H-1,2,4-triazole (4b). Yield, 86%; mp 175–176 °C; IR
5.3.8. 4-(4-Bromophenyl)-3-ethylthio-5-(4-methylsulfo-
nylphenyl)-4H-1,2,4-triazole (4h). Yield, 83%; mp 172–
1
(KBr): 1302, 1150 (SO2) cmꢀ1
;
1H NMR (CDCl3) d
174 °C; IR (KBr): 1308, 1150 (SO2) cmꢀ1; H NMR
7.85 (d, J = 8.8 Hz, 2H), 7.62 (d, J = 8.8 Hz, 2H),
7.59–7.51 (m, 3H), 7.29–7.22 (m, 2H), 3.37 (q,
J = 7.2 Hz, 2H), 3.04 (s, 3H), 1.45 (t, J = 7.2 Hz, 3H);
MS m/z (%) 359 (68), 330 (100), 296 (4), 279 (4), 256
(7). Anal. Calcd for C17H17N3O2S2: C, 56.80; H, 4.77;
N, 11.69. Found: C, 56.63; H, 4.59; N, 11.60.
(CDCl3) d 7.89 (d, J = 8.4 Hz, 2H), 7.67 (d, J = 8.8 Hz,
2H), 7.62 (d, J = 8.4 Hz, 2H), 7.12 (d, J = 8.8 Hz, 2H),
3.33 (q, J = 7.2 Hz, 2H), 3.05 (s, 3H), 1.46 (t,
J = 7.2 Hz, 3H); MS m/z (%) 439 (52), 437 (52), 412
(81), 410 (81), 330 (11), 328 (11), 257 (17), 255 (17),
250 (72), 198 (16), 196 (16), 185 (100), 183 (100), 171
(25), 157 (13), 104 (33), 90 (32), 78 (20). Anal. Calcd
for C17H16BrN3O2S2: C, 46.58; H, 3.68; N, 9.59. Found:
C, 46.42; H, 3.49; N, 9.73.
5.3.3. 4-(4-Fluorophenyl)-5-(4-methylsulfonylphenyl)-3-
methylthio-4H-1,2,4-triazole (4c). Yield, 90%; mp 201–
203 °C; IR (KBr): 1307, 1157 (SO2) cmꢀ1
;
1H NMR
(CDCl3) d 7.87 (d, J = 8.4 Hz, 2H), 7.62 (d, J = 8.4 Hz,
2H), 7.15–7.31 (m, 4H), 3.05 (s, 3H), 2.76 (s, 3H); MS
m/z (%) 363 (85), 283 (20), 281 (35), 249 (15), 236 (45),
207 (100), 197 (42), 176 (23), 145 (30), 135 (58), 109
(78), 91 (16). Anal. Calcd for C16H14FN3O2S2: C,
52.88; H, 3.88; N, 11.56. Found: C, 52.69; H, 3.76; N,
11.62.
5.3.9. 4-(4-Methoxyphenyl)-5-(4-methylsulfonylphenyl)-
3-methylthio-4H-1,2,4-triazole (4i). Yield, 76%; mp
176–177 °C; IR (KBr): 1301, 1151 (SO2) cmꢀ1
;
1H
NMR (CDCl3) d 7.86 (d, J = 8.8 Hz, 2H), 7.65 (d,
J = 8.8 Hz, 2H), 7.16 (d, J = 8.8 Hz, 2H), 7.02 (d,
J = 8.8 Hz, 2H), 3.89 (s, 3H), 3.05 (s, 3H), 2.75 (s,
3H); MS m/z (%) 375 (30), 376 (100), 342 (10), 312
(15), 288 (17), 209 (27), 165 (12), 147 (37), 121 (12),
102 (15), 89 (16), 63 (36). Anal. Calcd for
C17H17N3O3S2: C, 54.38; H, 4.56; N, 11.19. Found: C,
54.52; H, 4.77; N, 11.31.
5.3.4. 3-Ethylthio-4-(4-fluorophenyl)-5-(4-methylsulfonyl-
phenyl)-4H-1,2,4-triazole (4d). Yield, 84%; mp 191–
192 °C; IR (KBr): 1307, 1153 (SO2) cmꢀ1
;
1H NMR
(CDCl3) d 7.88 (d, J = 8.4 Hz, 2H), 7.63 (d, J = 8.4 Hz,
2H), 7.21–7.29 (m, 4H), 3.33 (q, J = 7.2 Hz, 2H), 3.05
(s, 3H), 1.44 (t, J = 7.2 Hz, 3H); MS m/z (%) 377 (60),
349 (58), 348 (100), 276 (5), 250 (18), 197 (5), 78 (12), 52
(34), 45 (35). Anal. Calcd for C17H16FN3O2S2: C, 54.09;
H, 4.27; N, 11.13. Found: C, 54.22; H, 4.18; N, 11.05.
5.3.10.
ylsulfonylphenyl)-4H-1,2,4-triazole (4j). Yield, 80%; mp
150–152 °C; IR (KBr): 1308, 1149 (SO2) cmꢀ1 1H
NMR (CDCl3) d 7.86 (d, J = 8.4 Hz, 2H), 7.65 (d,
J = 8.4 Hz, 2H), 7.15 (d, J = 8.8 Hz, 2H), 7.01 (d,
J = 8.8 Hz, 2H), 3.89 (s, 3H), 3.31 (q, J = 7.6 Hz, 2H),
3.05 (s, 3H), 1.45 (t, J = 7.6 Hz, 3H); MS m/z (%) 389
(40), 373 (56), 360 (59), 346 (50), 344 (100), 288 (5),
250 (12), 209 (3), 135 (6), 119 (5), 65 (3). Anal. Calcd
for C18H19N3O3S2: C, 55.51; H, 4.92; N, 10.79. Found:
C, 55.66; H, 5.12; N, 10.86.
3-Ethylthio-4-(4-methoxyphenyl)-5-(4-meth-
;
5.3.5. 4-(4-Chlorophenyl)-5-(4-methylsulfonylphenyl)-3-
methylthio-4H-1,2,4-triazole (4e). Yield, 77%; mp 131–
132 °C; IR (KBr): 1305, 1148 (SO2) cmꢀ1
;
1H NMR
(CDCl3) d 7.88 (d, J = 8.8 Hz, 2H), 7.62 (d, J = 8.8 Hz,
2H), 7.52 (d, J = 8.8 Hz, 2H), 7.20 (d, J = 8.8 Hz, 2H),
3.05 (s, 3H), 2.76 (s, 3H); MS m/z (%) 379 (100), 365
(65), 346 (100), 317 (40), 292 (88), 236 (68), 213 (48),
208 (12), 151 (28), 125 (90), 75 (12), 63 (13). Anal. Calcd
for C16H14ClN3O2S2: C, 50.59; H, 3.71; N, 11.06.
Found: C, 50.48; H, 3.58; N, 10.95.
5.3.11. 4-(4-Methylphenyl)-5-(4-methylsulfonylphenyl)-3-
methylthio-4H-1,2,4-triazole (4k). Yield, 82%; mp 109–
1
110 °C; IR (KBr): 1295, 1149 (SO2) cmꢀ1; H NMR
(CDCl3) d 7.85 (d, J = 8.4 Hz, 2H), 7.64 (d, J = 8.4 Hz,
2H), 7.33 (d, J = 8.4 Hz, 2H), 7.12 (d, J = 8.4 Hz, 2H),
3.05 (s, 3H), 2.75 (s, 3H), 2.46 (s, 3H); MS m/z (%)
359 (100), 345 (19), 326 (45), 297 (10), 279 (16), 273
(14), 235 (8), 131 (11), 105 (15). Anal. Calcd for
C17H17N3O2S2: C, 56.80; H, 4.77; N, 11.69. Found: C,
56.92; H, 4.89; N, 11.56.
5.3.6. 4-(4-Chlorophenyl)-3-ethylthio-5-(4-methylsulfo-
nylphenyl)-4H-1,2,4-triazole (4f). Yield, 82%; mp 175–
176 °C; IR (KBr): 1317, 1152 (SO2) cmꢀ1
;
1H NMR
(CDCl3) d 7.87 (d, J = 8.8 Hz, 2H), 7.62 (d, J = 8.8 Hz,
2H), 7.52 (d, J = 8.8 Hz, 2H), 7.19 (d, J = 8.8 Hz, 2H),
3.33 (q, J = 7.2 Hz, 2H), 3.05 (s, 3H), 1.46 (t,
J = 7.2 Hz, 3H); MS m/z (%) 394 (55), 366 (33), 364
(100), 330 (10), 292 (8), 250 (13), 213 (12), 139 (9), 63
(15). Anal. Caclcd for C17H16ClN3O2S2: C, 51.83; H,
4.09; N, 10.67. Found: C, 51.90; H, 4.01; N, 10.88.
5.3.12. 3-Ethylthio-4-(4-methylphenyl)-5-(4-methylsulfo-
nylphenyl)-4H-1,2,4-triazole (4l). Yield, 76%; mp 147–
1
148 °C; IR (KBr): 1298, 1148 (SO2) cmꢀ1; H NMR
(CDCl3) d 7.85 (d, J = 8.8 Hz, 2H), 7.64 (d, J = 8.8 Hz,
2H), 7.32 (d, J = 8.4 Hz, 2H), 7.11 (d, J = 8.4 Hz, 2H),
3.31 (q, J = 7.2 Hz, 2H), 3.04 (s, 3H), 2.46 (s, 3H),
1.45 (t, J = 7.2 Hz, 3H); MS m/z (%) 373 (26), 358
(68), 344 (45), 330 (100), 313 (4), 250 (16), 213 (5), 139
(10), 63 (5). Anal. Calcd for C18H19N3O2S2: C, 57.88;
H, 5.13; N, 11.25. Found: C, 57.69; H, 5.03; N, 11.16.
5.3.7. 4-(4-Bromophenyl)-5-(4-methylsulfonylphenyl)-3-
methylthio-4H-1,2,4-triazole (4g). Yield, 80%; mp 146–
147 °C; IR (KBr): 1304, 1149 (SO2) cmꢀ1
;
1H NMR
(CDCl3) d 7.89 (d, J = 8.8 Hz, 2H), 7.68 (d, J = 8.8 Hz,
2H), 7.62 (d, J = 8.8 Hz, 2H), 7.13 (d, J = 8.8 Hz, 2H),
3.05 (s, 3H), 2.76 (s, 3H); MS m/z (%) 425 (100), 423
(100), 390 (55), 360 (30), 338 (50), 311 (42), 259, 257
(25), 236 (45), 197, 195 (27), 171, 169 (75), 91 (13), 76
(14), 63 (12). Anal. Calcd for C16H14BrN3O2S2: C,
45.29; H, 3.33; N, 9.90. Found: C, 45.45; H, 3.52; N, 9.81.
5.3.13. 4-Cyclohexyl-5-(4-methylsulfonylphenyl)-3-meth-
ylthio-4H-1,2,4-triazole (4m). Yield, 74%; mp 197–
199 °C; IR (KBr) 1309, 1152 (SO2) cmꢀ1
;
1H NMR
(CDCl3) d 8.09 (d, J = 8.4 Hz, 2H), 7.72 (d, J = 8.4 Hz,

L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
2515
2H), 3.98 (m, 1H), 3.13 (s, 3H), 2.82 (s, 3H), 2.26–2.10
(m, 3H), 1.96–1.70 (m, 5H), 1.74–1.64 (m, 2H); MS m/
z (%) 351(37), 336 (3), 269 (100), 242 (15), 223 (12),
207 (4), 190 (13), 182 (13), 161 (15), 83 (20), 65 (16).
Anal. Calcd for C16H21N3O2S2: C, 54.67; H, 6.02; N,
11.95. Found: C, 54.52; H, 6.18; N, 11.82.
and extracted with ethyl acetate. The organic phase
was washed with brine, dried (Na2SO4) and evaporated.
Crystallization of the residue gave the title compounds
8a–c.
5.5.1. N-(4-Methylsulfonylphenyl)phenylcarbohydrazon-
amide (8a). Compound 8a was reported previously.20
5.3.14. 4-Cyclohexyl-3-ethylthio-5-(4-methylsulfonylphe-
nyl)-4H-1,2,4-triazole (4n). Yield, 70%; mp 221–223 °C;
5.5.2. 4-Fluorophenyl-N-(4-methylsulfonylphenyl)carbo-
hydrazonamide (8b). Yield, 83%; mp 199–201 °C; IR
(KBr): 3416, 3309 (NH, NH2), 1588 (C@N), 1286,
1
IR (KBr): 1311, 1159 (SO2) cmꢀ1; H NMR (CDCl3) d
8.09 (d, J = 8.4 Hz, 2H), 7.73 (d, J = 8.4 Hz, 2H), 3.98
(m, 1H), 3.41 (q, J = 7.6 Hz, 2H), 3.13 (s, 3H), 2.23–
2.15 (m, 7H), 1.49 (t, J = 7.6 Hz, 3H); MS m/z (%) 366
(M+1, 17), 365 (33), 336 (16), 284 (82), 283 (100), 256
(93), 255 (78), 251 (63), 203 (32), 182 (34),118 (14), 102
(17), 81 (15). Anal. Calcd for C17H23N3O2S2: C, 55.86;
H, 6.34; N, 11.50. Found: C, 55.69; H, 6.47; N, 11.66.
1
1132 (SO2) cmꢀ1; H NMR (CDCl3) d 7.78–7.35 (m,
4H), 7.05 (t, J = 8.36 Hz, 2H), 6.75 (d, J = 8.6 Hz,
2H), 5.95 (br s, 2H), 5.40 (br s, 1H), 2.99 (s, 3H). Anal.
Calcd for C14H14FN3O2S: C, 54.71; H, 4.59; N, 13.67.
Found: C, 54.88; H, 4.75; N, 13.49.
5.5.3. 4-Methylphenyl-N-(4-methylsulfonylphenyl)carbo-
hydrazonamide (8c). Yield, 80%; mp 211–213 °C; IR
(KBr): 3489 (NH), 3380, 3299 (NH2), 1555 (C@N),
5.4. General procedure for the preparation of N-(4-
methylsulfonylphenyl)-4-substituted benzamides (7a–c)
1
1298, 1132 (SO2) cmꢀ1; H NMR (CDCl3) d 7.65 (d,
To a solution of 4-methylsulfonylaniline 5 (300 mg,
1.75 mmol) in THF (20 mL) was added dropwise 4-
substituted benzoyl chloride 6a–c (1.78 mmol) dissolved
in THF (5 mL) under N2 at 0 °C followed by addition of
triethylamine (0.25 mL, 1.8 mmol). The reaction mix-
ture was stirred overnight at 24 °C, filtered, and the pre-
cipitate was washed with THF. The filtrate was
concentrated and crystallized from methanol to give
the title compounds 7a–c.
J = 8.6 Hz, 2H), 7.40 (d, J = 8.4 Hz, 2H), 7.18 (d,
J = 8.4 Hz, 2H), 6.75 (d, J = 8.6 Hz, 2H), 6.10 (br s,
2H), 5.52 (br s, 1H), 2.99 (s, 3H), 2.38 (s, 3H). Anal.
Calcd for C15H17N3O2S: C, 59.38; H, 5.65; N, 13.85.
Found: C, 59.56; H, 5.81; N, 13.66.
5.6. General procedure for the preparation of 4-(4-methyl
sulfonylphenyl-5-aryl-2,4-dihydro-3H-1,2,4-triazol-3-thiones
(9a–c)
5.4.1. N-(4-Methylsulfonylphenyl)benzamide (7a). Com-
pound 7a was reported previously.20
N-(4-Methylsulfonylphenyl)-4-substituted benzenecar-
bohydrazonamides 8a–c (1.73 mmol) were dissolved in
THF (100 mL) under N2, and 1,10-thiocarbonyldiimi-
dazole (340 mg, 2 mmol) was added. The solution was
stirred overnight at room temperature. The solvent
was removed under reduced pressure. The residue was
taken up in ethyl acetate and washed with 0.1 N HCl
solution, water, and brine prior to drying (Na2SO4).
The solvent was evaporated and the residue was crystal-
lized from methanol to give the title compounds 9a–c.
5.4.2. N-(4-Methylsulfonylphenyl)-4-fluorobenzamide (7b).
Yield, 93%; mp 218–219 °C; IR (KBr): 3375 (NH), 1680
(CO), 1291, 1143 (SO2) cmꢀ1; 1H NMR (CDCl3) d 10.18
(s, 1H), 8.08–7.90 (m, 4H), 7.90 (d, J = 8.8 Hz, 2H), 7.18
(t, J = 8.4 Hz, 2H), 3.06 (s, 3H). Anal. Calcd for
C14H12FNO3S: C, 57.33; H, 4.12; N, 4.78. Found: C,
57.50; H, 4.29; N, 4.90.
5.4.3. N-(4-Methylsulfonylphenyl)-4-methylbenzamide (7c).
Yield, 96%; mp 258–259 °C; IR (KBr): 3365 (NH), 1674
5.6.1. 4-(4-Methylsulfonylphenyl)-5-phenyl-2,4-dihydro-3H-
1,2,4-triazol-3-thione (9a). Compound 9a was reported
previously.20
1
(CO), 1294, 1142 (SO2) cmꢀ1; H NMR (CDCl3) d 9.81
(s, 1H), 7.96 (d, J = 8.8 Hz, 2H), 7.90 (d, J = 8.8 Hz,
2H), 7.79 (d, J = 8.4 Hz, 2H), 7.32 (d, J = 8.4 Hz, 2H),
3.06 (s, 3H), 2.45 (s, 3H). Anal. Calcd for C15H15NO3S:
C, 62.26; H, 5.23; N, 4.84. Found: C, 62.36; H, 5.36; N,
4.96.
5.6.2. 5-(4-Fluorophenyl)-4-(4-methylsulfonylphenyl)-2,4-
dihydro-3H-1,2,4-triazol-3-thione (9b). Yield, 48%; mp
227–229 °C; IR (KBr): 3396 (NH), 1367, 1114 (SO2)
1
cmꢀ1; H NMR (CDCl3 + DMSO-d6) d 13.90 (s, 1H),
8.07 (d, J = 7.8 Hz, 2H), 7.52–7.18 (m, 6H), 3.14 (s,
3H). Anal. Calcd for C15H12FN3O2S2: C, 51.56; H,
3.46; N, 12.03. Found: C, 51.65; H, 3.59; N, 11.89.
5.5. General procedure for the preparation of N-(4-meth-
ylsulfonylphenyl)arylcarbohydrazonamides (8a–c)
N-(4-Methylsulfonylphenyl)-4-substituted benzamides
7a–c (1.8 mmol) were dissolved in benzene (10 mL) un-
der N2, and phosphorus pentachloride (416 mg, 2 mmol)
was added. The solution was heated at reflux for 3 h. It
was concentrated to remove POCl3, the residue was tak-
en up in THF (15 mL) and added dropwise into a stirred
THF (10 mL) solution of anhydrous hydrazine (0.6 mL)
5.6.3.
5-(4-Methylphenyl)-4-(4-methylsulfonylphenyl)-
2,4-dihydro-3H-1,2,4-triazol-3-thione (9c). Yield, 55%;
mp 239–241 °C; IR (KBr): 3412 (NH), 1319, 1149
(SO2) cmꢀ1; H NMR (CDCl3 + DMSO-d6) d 13.68 (s,
1
1H), 8.02 (d, J = 8 Hz, 2H), 7.55 (d, J = 8 Hz, 2H),
7.28 (d, J = 8.4 Hz, 2H), 7.10 (d, J = 8.4 Hz, 2H), 3.13
(s, 3H), 2.44 (s, 3H). Anal. Calcd for C16H15N3O2S2:
C, 55.63; H, 4.38; N, 12.16. Found: C, 55.81; H, 4.45;
N, 12.02.
at 0 °C under N . The reaction mixture was stirred for
2
1 h at room temperature, poured into water (20 mL),

2516
L. Navidpour et al. / Bioorg. Med. Chem. 14 (2006) 2507–2517
5.7. General procedure for the preparation of 3-alkylthio-
4-(4-methylsulfonylphenyl)-5-phenyl-4H-1,2,4-triazoles
(10a–f)
(CDCl3) d 8.06 (d, J = 8.8 Hz, 2H), 7.43 (d, J = 8.8 Hz,
2H), 7.22 (d, J = 8.4 Hz, 2H), 7.11 (d, J = 8.4 Hz, 2H),
3.32 (q, J = 7.2 Hz, 2H), 3.14 (s, 3H), 1.45 (t,
J = 7.2 Hz, 3H); MS m/z (%) 374 (26), 373 (94), 346
(92), 344 (100), 272 (22), 265 (15), 193 (58), 191 (90),
186 (92), 163 (19), 149 (17), 134 (18), 121 (37), 91 (73),
76 (48). Anal. Calcd for C18H19N3O2S2: C, 57.88; H,
5.13; N, 11.25. Found: C, 57.69; H, 5.02; N, 11.15.
To a stirring solution of compounds9a–c (1 mmol) and
sodium hydroxide (1 mmol) in ethanol (5 mL) was add-
ed alkyl iodide (0.5 mL) and the mixture was stirred
overnight. The volatiles were evaporated. Flash chroma-
tography (MeOH/CHCl3, 1:20) gave the title com-
pounds 10a–f.
5.8. In vitro cyclooxygenase (COX) inhibition assay
5.7.1. 4-(4-Methylsulfonylphenyl)-3-methylthio-5-phenyl-
4H-1,2,4-triazole (10a). Compound 10a was reported
previously.20
The ability of the test compounds listed in Tables 1–3 to
inhibit ovine COX-1 and COX-2 was determined using a
colorimetric COX (ovine) inhibitor screening assay
which utilizes the peroxidase component of cyclooxy-
genase. The peroxidase activity is assayed colorimetri-
cally by monitoring the appearance of oxidized
N,N,N0,N0-tetramethyl-p-phenylenediamine (TMPD) at
590 nm.23–25
5.7.2. 3-Ethylthio-4-(4-methylsulfonylphenyl)-5-phenyl-
4H-1,2,4-triazole (10b). Yield, 82%; mp 173–175 °C; IR
(KBr): 1314, 1155 (SO2) cmꢀ1 1H NMR (CDCl3) d
;
8.05 (d, J = 8.4 Hz, 2H), 7.43 (d, J = 8.4 Hz, 2H), 7.33
(s, 5H), 3.32 (q, J = 7.2 Hz, 2H), 3.12 (s, 3H), 1.49 (t,
J = 7.2 Hz, 3H); MS m/z (%) 359 (78), 331 (100), 326
(10), 258 (22), 251 (14), 183 (32), 177 (65), 172 (73), 111
(9), 103 (10). Anal. Calcd for C17H17N3O2S2: C, 56.80;
H, 4.77; N, 11.69. Found: C, 56.92; H, 4.95; N, 11.56.
5.9. Anti-inflammatory assay
The test compounds were evaluated using in vivo rat
carrageenan-induced foot paw edema model reported
previously.27
5.7.3. 5-(4-Fluorophenyl)-4-(4-methylsulfonylphenyl)-3-
methylthio-4H-1,2,4-triazole (10c). Yield, 85%; mp
5.10. Molecular modeling (docking) studies
212–214 °C; IR (KBr): 1322, 1156 (SO2) cmꢀ1
;
1H
NMR (CDCl3) d 8.08 (d, J = 8 Hz, 2H), 7.42 (d,
J = 8 Hz, 2H), 7.32 (dd, J = 8.4, 5.6 Hz, 2H), 7.01 (t,
J = 8.4 Hz, 2H), 3.13 (s, 3H), 2.75 (s, 3H); MS m/z (%)
363 (100), 330 (10), 276 (12), 251 (12), 197 (22), 176
(47), 152 (15), 122 (17), 107 (92), 91 (25), 76 (27), 63
(20). Anal. Calcd for C16H14FN3O2S2: C, 52.88; H,
3.88; N, 11.56. Found: C, 52.78; H, 3.72; N, 11.44.
Docking studies were performed using MOE software
version 2003.02 (CCG Inc.).26 The coordinates of the
X-ray crystal structure of the selective COX-2 inhibitor
SC-558 bound to the murine COX-2 enzyme were ob-
tained from the RCSB Protein Data Bank (1cx2). The
ligand molecules were constructed using the Builder
module and were energy optimized. The purpose of
docking is to search for favorable binding configuration
between the small flexible ligands and the rigid protein.
5.7.4. 3-Ethylthio-5-(4-fluorophenyl)-4-(4-methylsulfonyl-
phenyl)-4H-1,2,4-triazole (10d). Yield, 87%; mp 168–
˚
Protein residues with atoms greater than 7.5 A from the
170 °C; IR (KBr): 1314, 1156 (SO2) cmꢀ1
;
1H NMR
docking box were removed for efficiency. Searching is
conducted within a specified 3D docking box using sim-
ulated annealing based on the Monte Carlo method and
MMFF94 molecular mechanics force fields for 8000
iterations.
(CDCl3) d 8.08 (d, J = 8 Hz, 2H), 7.44 (d, J = 8 Hz, 2H),
7.34 (dd, J = 8.4, 5.6 Hz, 2H), 7.02 (t, J = 8.4 Hz, 2H),
3.34 (q, J = 7.2 Hz, 2H), 3.15 (s, 3H), 1.46 (t, J = 7.2 Hz,
3H); MS m/z (%) 377 (64), 350 (80), 348 (100), 304 (23),
302 (95), 184 (22), 270 (12), 245 (17), 229 (20), 211 (45),
190 (92), 149 (62), 121 (37), 91 (57), 79 (38), 57 (45). Anal.
Calcd for C17H16FN3O2S2: C, 54.09; H, 4.27; N, 11.13.
Found: C, 54.22; H, 4.10; N, 11.02.
Acknowledgments
This work was supported by grants from the research
council of Tehran University of Medical Sciences and
Iranian National Science Foundation. We also thank
Dr. H. Shadnia for helpful discussions.
5.7.5. 5-(4-Methylphenyl)-4-(4-methylsulfonylphenyl)-3-
methylthio-4H-1,2,4-triazole (10e). Yield, 80%; mp
218–220 °C; IR (KBr): 1308, 1154 (SO2) cmꢀ1
;
1H
NMR (CDCl3) d 8.06 (d, J = 8.8 Hz, 2H), 7.44 (d,
J = 8.8 Hz, 2H), 7.23 (d, J = 8.4 Hz, 2H), 7.11 (d,
J = 8.4 Hz, 2H), 3.14 (s, 3H), 2.76 (s, 3H); MS m/z (%)
360 (66), 359 (100), 326 (28), 280 (17), 271 (26), 247
(35), 204 (24), 193 (39), 172 (64), 162 (37), 118 (35),
107 (69), 90 (43), 76 (31). Anal. Calcd for
C17H17N3O2S2: C, 56.80; H, 4.77; N, 11.69. Found: C,
56.69; H, 4.66; N, 11.55.
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