Synthesis and in-silico studies of some diaryltriazole derivatives as potential cyclooxygenase inhibitors

Article abstract of DOI:10.1007/s12272-013-0078-6

The synthesis of several 4-phenyl-5-pyridin-4-yl-2,3-dihydro-3H-1,2,4- triazole-3-thiones possessing N-2 Mannich bases or S-alkyl substituents, is reported. Several of them exhibited a low nanomolar COX enzyme inhibition activity. Most of the compounds sh

Full text of DOI:10.1007/s12272-013-0078-6

Arch. Pharm. Res. (2013) 36:553–563
DOI 10.1007/s12272-013-0078-6
RESEARCH ARTICLE
Synthesis and in-silico studies of some diaryltriazole derivatives
as potential cyclooxygenase inhibitors
•
Awwad A. Radwan Kamal E. H. elTahir
Received: 2 July 2012 / Accepted: 16 February 2013 / Published online: 12 March 2013
Ó The Pharmaceutical Society of Korea 2013
Abstract The synthesis of several 4-phenyl-5-pyridin-4-
yl-2,3-dihydro-3H-1,2,4-triazole-3-thiones possessing N-2
Mannich bases or S-alkyl substituents, is reported. Several
of them exhibited a low nanomolar COX enzyme inhibition
activity. Most of the compounds showed inhibition of
edema was similar to that evoked by celocoxib in animal
model. Molecular docking studies of the compounds into
the binding sites of COX-1 and COX-2 allowed us to shed
light on the binding mode of these novel COX inhibitors.
isomerases. The biosynthesis of these prostaglandins is
practically in all tissues of the human body, eliciting a variety
of pharmacological effects, some of them beneficial, as
support of renal and platelet functions, gastrointestinal pro-
tection, and other nonbeneficial as pain, fever and other
symptoms associated with the inflammatory response. Since
1987, it has been established that COX enzyme exists in two
isoforms (Tanaka et al. 1987), and are encoded by separate
genes on different chromosomes. Cyclooxygenase inhibition
formed the basis for the success of non-steroidal anti-
inflammatory drugs (NSAIDs) in treating a variety of pain
syndromes. The drawback, however, was that every year,
2–4 % of patients taking NSAIDs suffered from symptom-
atic gastrointestinal ulcers and their complications (Silver-
stein et al. 2000). As the scientific research behind the COX
enzyme progressed, it became apparent that COX-2 inhibi-
tion mediated the anti-inflammatory effects of NSAIDs,
whereas COX-1 inhibition was responsible for the adverse
effects on the gastrointestinal tract. It therefore became
reasonable to assume that inhibiting COX-2 selectively
would result in the same anti-inflammatory benefits that non-
selective NSAIDs provided but with fewer gastrointestinal
side effects. This enforced the interest of pharmaceutical
industries in manufacturing new analgesic and anti-inflam-
matory medications known as selective COX-2 inhibitors or
coxibs (Penning et al. 1997; Riendeau et al. 2001; Ranatunge
et al. 2004). A selective COX-2 inhibitor allows the desired
synthesis of cytoprotective prostaglandins, in conjunction
with a simultaneous inhibition of proinflammatory prosta-
glandin synthesis, thereby reducing dyspepsia and ulceration
(Meade et al. 1993). However, emerging evidence suggests
that adverse reactions such as gastrointestinal 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 inhibition
Keywords Triazole Á Cyclooxygenase Á
Antiinflammatory Á Surflex-Dock
Introduction
Through the path of prostaglandin–thromboxane synthesis,
the transformation of arachidonic acid to prostaglandin H2
(PGH2) is catalyzed by cytosolic prostaglandin G/H syn-
thase, more commonly known as cyclooxygenase (COX).
Prostaglandin H2 is an unstable intermediate and is further
converted to one of many prostanoids, such as prostacyclin
(PGI2) and thromboxane A2 (TXA2), by tissue-specific
A. A. Radwan
Kayyali’s Chair, College of Pharmacy, King Saud University,
P.O. Box 2457, Riyadh 11451, Saudi Arabia
A. A. Radwan (&)
Department of Pharmaceutical Organic Chemistry,
Faculty of Pharmacy, Assiut University, Assiut 71527, Egypt
e-mail: dhna_2001@hotmail.com
K. E. H. elTahir
Department of Pharmacology, College of Pharmacy, King Saud
University, P. O. Box 2457, Riyadh 11421, Saudi Arabia
123

554
A. A. Radwan, K. E. H. elTahir
occurring with long-term exposure or at higher doses (Wolfe
et al. 1999). COX-2 selective inhibitors are also known to
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 vasoconstrictor, and
promoter of platelet aggregation, which is synthesized in
platelets by COX-1 (Catella-Lawson and Crofford 2001;
Mukherjee et al. 2001). Therefore, COX-2 inhibitors
intrinsically lack anti-thrombotic activity, and some car-
diovascular liabilities have been associated preclinically
with them (deGaetano et al. 2003). Thus, there is still a need
for novel, selective, and potent COX-2 inhibitors with an
improved profile compared to current COX-2 inhibitors
(Ranatunge et al. 2004). Diaryl heterocycles, and other
central ring pharmacophore templates, have been exten-
sively studied as cyclooxygenase inhibitors. All these tri-
cyclic molecules possess 1,2-diaryl substitution on a central
four-, five-, or six-membered ring system such as cyclobu-
tenone, pyrazole, 2-(5H)-furanone, isoxazole, pyridine, or
thiazolidinedione respectively (Talley et al. 2000; Penning
et al. 1997; Riendeau et al. 2001; AbdelMoallem et al. 2011;
Zarghi et al. 2011; Ali et al. 2007). Recently, a novel class of
6-alkylthio-substituted six-membered lactone (pyrane-2-
one) rings has been designed and exhibited very good in vitro
COX-2 inhibitory potency and selectivity (Praveen et al.
2003). Furthermore, differently substituted 1,2,4-triazole-3-
thione derivatives exhibited anti-inflammatory activity
(Labanauskas et al. 2001; Tozkoparan et al. 2005). Structure
based studies have been performed to identify binding modes
and important interactions of triaryl rings at the COX-2
active site (Plount-Price and Jorgensen 2000). The triaryl
ring moiety either containing a para-sulfonyl group (Talley
et al. 2000; Penning et al. 1997; Riendeau et al. 2001;
AbdelMoallem et al. 2011; Zarghi et al. 2011; Ali et al. 2007)
or not containing a para-sulfonyl group (Sui et al. 2000;
Dannhardt and Laufer 2000; Moreau et al. 2006) has been
recognized as a pharmacophore for selective COX-2
inhibition.
and are uncorrected. IR spectra were obtained, as KBr discs,
on a Perkin-Elmer FT-IR spectrophotometer at the Research
Center, College of Pharmacy, King Saud University, Saudi
Arabia. The data are given in mmax (cm-1). 1H and 13C NMR
spectra were recorded in either DMSO-d6 or CDCl3 on a
BrukerNMR spectrophotometeroperating at500 MHzfor1H
and 125.76 MHz for 13C; the chemical shifts are expressed in
d (ppm) downfield from tetramethylsilane (TMS) used as
internal standard. Mass spectra were taken on a Varian320-
MS spectrometer at the Research Center, College of Phar-
macy, KingSaudUniversity, SaudiArabia. Massspectraldata
were given as m/z (intensity%). Elemental analysis was per-
formed on a Perkin-Elmer CHNSO analyzer, model no. 2400.
Monitoring of reactions and checking of purity of the final
products were carried out by thin layer chromatography
(TLC) using silica gel precoated aluminum sheets (60 F254,
Merck) and visualization with ultraviolet light (UV) at
365 and 254 nm. Isonicotinic acid hydrazide (1) and phen-
ylisothiocyanate (2) were purchased from Sigma-Aldrich
Company (St. Louis, MO, USA). 2-Isonicotinoyl-N-phe-
nylhydrazinecarbothioamide (3) and 4-phenyl-5-pyridin-4-
yl-2,4-dihydro-3H-1,2,4-triazole-3-thione (4) were synthe-
sized according reported procedures (Bayrak et al. 2009).
Molecular modeling was carried out using Surflex-Dock
module within Sybyl 8.1.1 package (2006) (Tripos Inc., St.
Louis, USA). The program operated under ‘‘Linux_OS2x’’
operating system installed on HPxw8400 workstation at lab-
oratory of drug design, School of Pharmaceutical Sciences,
Kitasato University, Tokyo, Japan.
Ethyl[4-phenyl-5-pyridin-4-yl-3-thioxo-2,4-dihydro-3H-
1,2,4-triazol-2-yl]carboxylate 5
Amixtureofcompound4 (2.54 g,10 mmol),0.9 ganhydrous
sodium carbonate and the ethyl chloroformate (10 mmol) in
DMF (5 mL) was refluxed for 7 h. The reaction mixture was
cooled then poured into ice-cold water. The solid formed was
filtered off and recrystallized from acetic acid to give com-
pounds 5. Yield 2.7 g, 82 %, m.p. 231–232 °C; IR (KBr)
3057, 1770, 1654, 1636, 1298, 828, 741, 697 cm-1; 1H NMR
(DMSO-d6, d ppm): 1.6 (3H, t, CH3), 4.5 (2H, q, CH2–O),
7.0–7.1 (3H, m, C2H, C4H and C6H of phenyl), 7.1–7.25 (2H,
m, C3H and C5H of phenyl), 7.4 (2H, d, C3H and C5H of
pyridine), 8.5 (2H, d, C2H and C6H of pyridine). Anal. calcd.
for C16H14N4O2S: C, 58.88; H, 4.32; N, 17.17; S, 9.82. Found:
C, 58.65; H, 4.53; N, 17.08; S, 9.61.
Considering these results and as part of our ongoing pro-
gram to design novel selective COX-2 inhibitors, we describe
herein the design, synthesis, and biological evaluation of a
novel diverse group of 4-phenyl-5-pyridin-4-yl-4H-1,2,4-tri-
azole derivatives incorporating 2-alkyl(aryl)aminomethyl or
3-alkkylthio moiety at the central triazole ring.
Materials and methods
2-Hydroxymethyl-4-phenyl-5-pyridin-4-yl-2,4-dihydro-3H-
1,2,4-triazole-3-thione 6
Chemistry
A mixture of 4 (2.54 g, 10 mmol) and formalin (2 mL,
25 mmol) in water (10 mL) was heated under reflux for
5 min, (Wujec et al. 2003; Wujec and Paneth 2008) cooled
Melting points were determined on Barnstead 9001 Electro-
thermal melting point apparatus using open capillary tubes
123

Triazoles: synthesis and docking studies
555
to room temperature and filtered. The solid was washed
with cold water and dried to produce 6. The crude product
was recrystallized from ethanol. Yield 2.2 g, 78 %, m.p.
277–278 °C; IR (KBr) 3450, 3116, 2851, 1654, 1270, 830,
741, 695 cm-1; 1H NMR (DMSO-d6, d ppm): 3.3–3.8 (1H,
broad s, OH), 5.7 (2H, s, NCH2O), 7.05–7.25 (3H, m, C2H,
C4H and C6H of phenyl), 7.25–7.4 (2H, m, C3H and C5H of
phenyl), 7.5 (2H, d, C3H and C5H of pyridine), 8.5 (2H, d,
C2H and C6H of pyridine). Anal. calcd. for C14H12N4OS:
C, 59.14; H, 4.25; N, 19.70; S, 11.28. Found: C, 58.92; H,
4.05; N, 19.40; S, 11.35.
d6, d ppm): 1.8 (4H, t, CH2CH2), 3.0 (4H, t, CH2N CH2),
5.7 (2H, s, NCH2N), 7.0–7.2 (3H, m, C2H, C4H and C6H of
phenyl), 7.2–7.4 (2H, m, C3H and C5H of phenyl), 7.6 (2H,
d, C3H and C5H of pyridine), 8.6 (2H, d, C2H and C6H of
pyridine). Anal. calcd. for C18H19N5S: C, 64.07; H, 5.68;
N, 20.75; S, 9.50. Found: C, 64.21; H, 5.82; N, 20.58; S,
9.34.
N-Methyl-N-[(4-phenyl-5-pyridin-4-yl-3-thioxo-2,
4-dihydro-3H-1,2,4-triazol-2-y)methyl]-2-aminoethanol 10
Yield 2.52 g, 73 %, m.p. 173–174 °C; IR (KBr) 3385, 3104,
1
2-Substituted-4-phenyl-5-pyridin-4-yl-2,4-dihydro-3H-
1,2,4-triazole-3-thiones 7–12
2850, 1654, 1270, 868, 774, 694 cm-1; H NMR, (CDCl3,
d ppm): 1.8 (1H, broad s, OH), 2.8 (3H, s, NCH3), 3.0 (2H, t,
NCH2), 4.0 (2H, t, CH2O), 5.8 (2H, s, NCH2N), 7.20–7.25
(3H, m, C2H, C4H and C6H of phenyl), 7.27–7.38 (2H, m,
C3H and C5H of phenyl), 7.6 (2H, d, C3H and C5H of pyri-
dine), 8.6 (2H, d, C2H and C6H of pyridine). Anal. calcd. for
C17H19N5OS: C, 59.80; H, 5.61; N, 20.51; S, 9.39. Found: C,
60.01; H, 5.50; N, 20.32; S, 9.25.
To a solution of compound 4 (2.54 g, 10 mmol) in DMF
(10 mL), formaldehyde (37 %, 1.55 mL) and an appro-
priate amine (10 mmol) were added and the mixture was
stirred at room temperature for 3 h. Then, excess amount of
pure water was added to this solution and the mixture was
kept overnight in cold. The resulting solid separated was
collected by filtration, washed with water, recrystallized
from ethanol to yield the title compounds.
2-[(4-Bromophenylamino)methyl]-4-phenyl-5-pyridin-4-yl-
2,4-dihydro-3H-1,2,4-triazole-3-thione 11
2-Dimethylaminomethyl-4-phenyl-5-pyridin-4-yl-2,
4-dihydro-3H-1,2,4-triazole-3-thione 7
Yield 3.19 g, 75 %, m.p. 208–209 °C; IR (KBr) 3032, 2944,
1
1654, 1636, 1276, 833, 807, 740 cm-1; H NMR, (CDCl3,
d ppm): 5.5 (1H, broad s, NH), 5.7 (2H, s, NCH2N), 6.9–7.0
(3H, m, C2H, C4H and C6H of phenyl), 7.15–7.3 (2H, m, C3H
and C5H of phenyl), 7.3 (2H, d, C2H and C6H of p-bromo-
phenyl), 7.4 (2H, d, C3H and C5H of p-bromophenyl), 7.6
(2H, d, C3H and C5H of pyridine), 8.6 (2H, d, C2H and C6H of
pyridine). 13C NMR, (CDCl3, d ppm): 65.44 (NCH2N),
111.52 (C), 115.94 (2CH), 121.69 (2CH), 128.06 (CH),
130.12 (2CH), 130.46 (2CH), 132.19 (2CH), 132.86 (C),
134.10 (C), 143.73 (C), 146.86 (2CH), 150.10 (triazole C-3),
169.51 (triazole C-5). Anal. calcd. for C20H16BrN5S: C,
54.80; H, 3.68; N, 15.98; S, 7.32. Found: C, 54.69; H, 3.51;
N, 15.83; S, 7.12.
Yield 2.52 g, 78 %, m.p. 173–174 °C; IR (KBr) 3065,
1
2973, 1654, 1279, 832, 752, 697 cm-1; H NMR, (CDCl3,
d ppm): 2.7 (6H, s, (CH3)2N), 5.3 (2H, s, NCH2N),
7.2–7.35 (3H, m, C2H and C6H of phenyl), 7.35–7.55 (3H,
m, C3H, C4H and C5H of phenyl), 7.6 (2H, d, C3H and C5H
of pyridine), 8.6 (2H, d, C2H and C6H of pyridine). Anal.
calcd. for C16H17N5S: C, 61.71; H, 5.50; N, 22.49; S,
10.30. Found: C, 61.53; H, 5.74; N, 22.21; S, 10.62.
2-Diethylaminomethyl-4-phenyl-5-pyridin-4-yl-2,
4-dihydro-3H-1,2,4-triazole-3-thione 8
Yield 2.79 g, 83 %, m.p. 216–217 °C; IR (KBr) 3076,
2-[(4-Chlorobenzylamino)methyl]-4-phenyl-5-pyridin-
4-yl-2,4-dihydro-3H-1,2,4-triazole-3-thione 12
1
2965, 1654, 1279, 835, 748, 695 cm-1; H NMR (DMSO-
d6, d ppm): 1.7 (6H, t, 2CH3), 2.6 (4H, q, 2CH2N), 5.7 (2H,
s, NCH2N), 7.0–7.2 (3H, m, C2H, C4H and C6H of phenyl),
7.2–7.4 (2H, m, C3H and C5H of phenyl), 7.6 (2H, d, C3H
and C5H of pyridine), 8.6 (2H, d, C2H and C6H of pyri-
dine). Anal. calcd. for C18H21N5S: C, 63.69; H, 6.24; N,
20.63; S, 9.45. Found: C, 63.85; H, 6.33; N, 20.42; S, 9.11.
Yield 2.89 g, 72 %, m.p. 222–223 °C; IR (KBr) 3052,
1
2928, 1654, 1636, 1274, 828, 798, 697 cm-1; H NMR,
(CDCl3, d ppm): 1.9 (1H, broad s, NH), 4.5 (2H, s, ben-
zylic CH2), 5.8 (2H, s, NCH2N), 7.12–7.17 (3H, m, C2H,
C4H and C6H of phenyl), 7.21–7.26 (2H, m, C3H and C5H
of phenyl), 7.29 (2H, d, C2H and C6H of p-chlorophenyl),
7.38 (2H, d, C3H and C5H of p-chlorophenyl), 7.55 (2H, d,
C3H and C5H of pyridine), 8.59 (2H, d, C2H and C6H of
pyridine). 13C NMR, (CDCl3, d ppm): 55.23 (benzylic
CH2N), 68.11 (NCH2N), 121.56 (2CH), 128.06 (CH),
128.20 (2CH), 129.63 (2CH), 130.05 (2CH), 130.34 (2CH),
4-Phenyl-5-pyridin-4-yl-2-pyrrolidinylmethyl-2,
4-dihydro-3H-1,2,4-triazole-3-thione 9
Yield 2.96 g, 90 %, m.p. 139 °C; IR (KBr) 3116, 2836,
1654, 1636, 1270, 832, 742, 695 cm-1; 1H NMR, (DMSO-
123

556
A. A. Radwan, K. E. H. elTahir
132.77 (C), 132.86 (C), 134.38 (C), 136.86 (C), 146.99
(2CH), 150.41 (triazole C-3), 170.09 (triazole C-5). Anal.
calcd. for C21H18ClN5S: C, 61.83; H, 4.45; N, 17.17; S,
7.86. Found: C, 61.60; H, 4.57; N, 17.01; S, 8.03.
d ppm): 0.85 (3H, t, CH3), 1.3–1.4 (2H, m, CH2), 1.4–1.7
(5H, m, CH3 and CH2), 3.5–4.1 (1H, m, CH), 7.0–7.15 (3H,
m, C2H, C4H and C6H of phenyl), 7.15–7.30 (2H, m, C3H
and C5H of phenyl), 7.5 (2H, d, C3H and C5H of pyridine),
8.4 (2H, d, C2H and C6H of pyridine). Anal. calcd. for
C18H20N4S: C, 66.63; H, 6.21; N, 17.27; S, 9.88. Found: C,
66.81; H, 6.35; N, 16.98; S, 10.03.
3-Substituted-4-phenyl-5-pyridin-4-yl-2,4-dihydro-3H-
1,2,4-triazoles 13-16
To a solution of compound 4 (2.54 g, 10 mmol) in absolute
ethanol, 1 equiv. of sodium was added and the mixture was
stirred at room temperature for 30 min. Then, the appro-
priate alkyl halide (20 mmol) was added and refluxed for
4 h. After evaporating the solvent under reduced pressure a
solid appeared. The solid was recrystallized from ethanol/
water (1:1) to obtain target compound.
3-[(Pent-4-enyl)thio]-4-phenyl-5-pyridin-4-yl-2,4-dihydro-
3H-1,2,4-triazole 16
Yield 2.29 g, 73 %, m.p. 132–133 °C; IR (KBr) 3037, 2932,
1654, 1640, 991, 914, 837, 776, 700 cm-1 1H NMR,
;
(CDCl3, d ppm): 1.6–2.4 (4H, m, =CCH2 CH2), 4.7 (2H, d,
=CH2), 5.1 (1H, d, =CH), 7.0–7.15 (3H, m, C2H, C4H and
C6H of phenyl), 7.15–7.30 (2H, m, C3H and C5H of phenyl),
7.5 (2H, d, C3H and C5H of pyridine), 8.4 (2H, d, C2H and
C6H of pyridine). Anal. calcd. for C18H18N4S: C, 67.05; H,
5.63; N, 17.38; S, 9.94. Found: C, 66.92; H, 5.75; N, 17.45; S,
10.06.
3-Methylthio-4-phenyl-5-pyridin-4-yl-2,4-dihydro-3H-
1,2,4-triazole 13
Yield 1.52 g, 61 %, m.p. 145–146 °C (reported 168–170 °C)
(Bayrak et al. 2009); IR (KBr) 3035, 2957, 1654, 1637, 1265,
836, 775, 693 cm-1; 1H NMR, (DMSO-d6, d ppm): 2.75 (1H,
s, SCH3), 7.27–7.29 (3H, m, C2H, C4H and C6H of phenyl),
7.32–7.34 (2H, m, C3H and C5H of phenyl), 7.58 (2H, d, C3H
and C5H of pyridine), 8.58 (2H, d, C2H and C6H of pyridine).
13C NMR, (DMSO-d6, d ppm): 14.59 (CH3–S), 121.50
(2CH), 127.11 (CH), 130.38 (2CH), 130.55 (2CH), 133.68
(1CH), 134.32 (1CH), 149.96 (triazole C-3), 152.51 (2CH),
155.47 (triazole C-5). Anal. calcd. for C14H12N4S: C, 62.66;
H, 4.51; N, 20.88; S, 11.95. Found: C, 62.51; H, 4.64; N,
20.72; S, 12.10.
In vivo carrageenan induced rat paw edema assay
Anti-inflammatory activity was determined by the carra-
geenan-induced rat paw edema method described by Winter
et al. (Winter et al. 1962). Male Sprague–Dawley rats
weighing 150–200 g (6–8 weeks old) were used in groups of
six animals per group for the experiments. The animals were
housed in a room with temperature of 22 2 °C under a 12 h
light/dark cycle. They were allowed free access to food and
water ad libitum. The protocol for the animal experiments
performed was approved by the Research Ethics Committee
and Animal Care and Use Committee, Govt. of Saudi Arabia.
Compounds were administered intravenously in dimethyl
sulfoxide solution. Paw edema was induced by intradermal
injection of 50 lL of 1 % k-carrageenan (Sigma, USA) into
the subplantar region of the right hind paw, after 1 h of
compound administration. The paw volume was measured
immediately after injection and after 2 h using a plethys-
mometer (UGO-Basile, Italy). The control group received
only the vehicle. Increase in paw volume was compared with
that in the control group and percent inhibition was calculated
taking the values in the control group as 0 % inhibition.
4-Phenyl-3-(prop-2-ynylthio)-5-pyridin-4-yl-2,4-dihydro-
3H-1,2,4-triazole 14
Yield 2.26 g, 79 %, m.p. 151–152 °C; IR (KBr) 3321,
2974, 1654, 1637, 826, 783, 698 cm-1; 1H NMR, (DMSO-
d6, d ppm): 1.75 (1H, s, HC:C), 4.1 (2H, s, :C–CH2S),
7.25–7.30 (3H, m, C2H, C4H and C6H of phenyl),
7.30–7.35 (2H, m, C3H and C5H of phenyl), 7.6 (2H, d,
C3H and C5H of pyridine), 8.57 (2H, d, C2H and C6H of
pyridine). 13C NMR, (DMSO-d6, d ppm): 22.12 (–CH2–S),
72.78 (CH:), 86.73 (:C), 121.49 (2CH), 127.18 (CH),
130.36 (2CH), 130.59 (2CH), 133.58 (1CH), 133.99 (1CH),
150.25 (triazole C-3), 152.63 (2CH), 153.52 (triazole C-5).
Anal. calcd. for C16H12N4S: C, 65.73; H, 4.14; N, 19.16; S,
10.97. Found: C, 65.56; H, 3.92; N, 19.25; S, 11.09.
In vitro COX inhibition assay
The final compounds were evaluated for their ability to
inhibit ovine COX-1 and COX-2 enzymes [50 % percent
inhibition is expressed in molar concentration (nmol)]
(Sano et al. 2005). Inhibition of the enzymes was deter-
mined with the colorimetric COX (ovine) inhibitor
screening assay kit (Cayman Chemicals, USA) using
ELISA reader.
3-[(Pent-2-yl)thio]-4-phenyl-5-pyridin-4-yl-2,4-dihydro-
3H-1,2,4-triazole 15
Yield 2.61 g, 77 %, m.p. 96–97 °C; IR (KBr) 3036, 2957,
1654, 1637, 836, 776, 694 cm-1 1H NMR, (CDCl3,
;
123

Triazoles: synthesis and docking studies
557
Molecular docking
Results and discussion
Molecular docking study was done using Surflex-Dock
within Sybyl 8.1.1 on HPxw8400 workstation, linux_OS2x
at laboratory of drug design, School of Pharmaceutical
Sciences, Kitasato University, Tokyo, Japan. The crystal
structures of the two isozymes COX-1 with flurbiprofen
(PDB entry code: 1cqe) (Picot et al. 1994) and the COX-2
with sc-558 (PDB entry code: 1cx2) (Kurumbail et al. 1996)
were retrieved from the RCSB Protein Data Bank. Surflex-
Dock uses an empirical scoring function and a patented
search engine to dock ligands into a protein’s binding site
(Jain 2003). A Protomol, which was used to guide molec-
ular docking, is a computational representation of the
intended binding site to which putative ligands are aligned.
Protomols can be produced by one of three routes: (Ruppert
et al. 1997) (1) automatic: Surflex-Dock finds the largest
cavity in the receptor protein; (2) ligand-based: a ligand in
the same coordinate space as the receptor; (3) residue-
based: specified residues in the receptor. Thus, a Protomol
can be generated automatically or defined based on a cog-
nate ligand or known active site. In the current paper, a
Protomol was generated automatically. Two parameters
determining the extent of the Protomol—a threshold
Chemistry
In Scheme 1, 1-isonicotinoyl-4-phenyl thiosemicarbazide
(3) and 4-phenyl-5-pyridin-4-yl-4H-1,2,4-triazole-3-thiol
(4) were prepared using a previously published procedure
(Pomarnacka and Kornicka 2001; Bayrak et al. 2009; Vera
et al. 2004). The reaction of 4-phenyl-5-pyridin-4-yl-4H-
1,2,4-triazole-3-thiol (4) with ethyl chloroformate in pres-
ence of anhydrous sodium carbonate afforded compound 5.
The reaction of compound 4 with formaline solution resulted
in 2-hydroxymethyl derivative (6). The reaction of 4-phenyl-
5-pyridin-4-yl-4H-1,2,4-triazole-3-thiol (4) with the appro-
priate amine in the presence of formaldehyde solution
afforded the corresponding Mannich base derivatives incor-
porating dimethylamine, diethylamine, pyrrolidine, N-methy-
lethanolamine, p-bromoaniline, p-chlorobenzylamine (7–12).
S-alkylation of 4-phenyl-5-pyridin-4-yl-4H-1,2,4-triazole-3-
thiol (4) was performed by its reaction with several alkyl
halides in basic media results in compounds (13–16) incor-
porating methyl, 2-pentyl, pent-4-enyl, 3-propynyl.
Anti-inflammatory activity
˚
parameter of 0.46 and a bloat parameter of 1 A—were
established. All the water molecules in 1cqe or 1cx2
(receptor) were deleted, and hydrogen atoms were added to
them (Muthas et al. 2008; Clark 2008). The protein struc-
ture was utilized in subsequent docking experiments with-
out energy minimization. In addition, treatment of docking
small molecules (ligands) was as follows: preparation of 3D
structures of ligands using Ligprep software module
(Schrodinger, Inc., New York, NY). During this preparation
step, hydrogen atoms were explicitly added, all possible
ionization states were generated between pH 6.0 and pH 8.0
using the apic module, and the 3D molecular structures
were minimized with OPLS 2005 force-field in Schrodinger
software suite. Tautomers were also generated in this step.
This preparation step was done on dell workstation under
linux x86-32 at laboratory of drug design, School of phar-
maceutical Sciences, Kitasato University, Tokyo, Japan.
Charge calculation with energy minimization method:
Powell; force field: tripos; charge: MMFF94; max itera-
The triazoles (5–16) were screened for their anti-inflamma-
tory activity using rat hind paw method of Winter et al. 1962
modified by Srimal and Dhawan 1974. Among the triazole
series compounds 6–9 and 13, 15 showed promising antiin-
flammatory activity compared with celecoxib as reference
drug (Table 1). Compounds with aromatic or unsaturated
moiety at position 2 or 3 respectively (11, 12, 14 and 16) are
either moderately active. Compounds 5 and 10 showed no
anti-inflammatory activity. In the literature it was shown that
the triazole-scaffold containing compounds exhibit anti-
inflammatory activity mediated through inhibition of COX-1
and 2 enzymes depending on the position and kind of sub-
stituent on the 1,2,4-triazole system (Navidpour et al. 2006).
The compounds under study are speculated to show anti-
inflammatory activity through COX-1 and/or COX-2.
In vitro cyclooxygenase inhibition
˚
tions: 1,000; termination: 0.001 kcal/(mol*A); root mean
˚
square (RMS) displacement: 0.001 A; other parameters:
The effect of incorporation of substituent at position 2 and/or
3 of the central triazole ring on COX-2 selectivity and
potency was determined by the IC50 values relative to the
reference compound, celecoxib, using colorimetric COX
(ovine) inhibitor screening assay (Table 1) (Kulmacz and
Lands 1983). In vitro enzyme inhibition studies for com-
pounds 5–16 showed variable COX-2 inhibitory activity. In
general, for these compounds, COX-2 selectivity and
potency were dependent upon steric properties of the sub-
stituent at position 2 and at position 3 on the central triazole
treated by default. In the docking procedure, five additional
starting conformations are used and ten binding poses per
ligand were obtained, and the binding pose with the highest
total score was taken into consideration for ligand–receptor
interactions. Docking results were validated by finding the
root mean squared deviation (RMSD) between the docking
position calculated for sc-588 and flurbiprofen and that
observed in their crystal structures.
123

558
A. A. Radwan, K. E. H. elTahir
Scheme 1 General synthesis of
compounds 5–16. Reagents and
conditions: i Ethanol, rt; ii
NaOH; iii ethyl chloroformate,
anhydrous sodium carbonate,
DMF, reflux 7 h; iv formaline
water, reflux 5 min; v amine,
dimethyl formamide,
S
NH
SCN
NH2
HN
O
HN NH
i
N
N
+
O
3
1
2
formaldehyde, rt 3 h; vi alkyl
halide, sodium ethoxide,
ethanol, reflux 4 h
ii
O
OEt
OH
S
H
N
N
N
N
iii
N
iv
S
N
S
N
N
N
N
N
N
5
4
6
vi
v
R
N
N
N
N
N
SR
N
S
N
N
13-16
7-12
CH3
13; R=
14; R=
CH
N
N
N
N
9; R=
7; R=
8; R=
10; R=
CH3
OH
15; R=
16; R=
HN
CH3
CH2
HN
Br 12; R=
Cl
11; R=
ring. Compounds (6–9, 13, 15) showed high in vitro potency
and selectivity (6; COX-2 IC50 = 0.8 nM; COX-1
IC50 = 3.5 nM; S.I. = 4.38, 7; COX-2 IC50 = 0.9 nM;
COX-1 IC50 = 3.1 nM; S.I. = 3.44, 8; COX-2 IC50 =
1.1 nM; COX-1 IC50 = 3.8 nM; S.I. = 3.46, 9; COX-2
IC50 = 0.9 nM; COX-1 IC50 = 3.7 nM; S.I. = 4.11, 13;
COX-2 IC50 = 2.1 nM; COX-1 IC50 = 5.3 nM; S.I. =
2.52, 15; COX-2 IC50 = 1.4 nM; COX-1 IC50 = 3.7 nM;
S.I. = 2.64) relative to the reference drug celecoxib (COX-2
IC50 = 1.9 nM; COX-1 IC50 = 4.1 nM; S.I. = 2.16).
Compounds 10, 16 explored almost equal or slight lower
potency and selectivity relative to the reference drug cele-
coxib (10; COX-2 IC50 = 2.1 nM; COX-1 IC50 = 4.5 nM;
S.I. = 2.14, 16; COX-2 IC50 = 2.6 nM; COX-1 IC50 =
4.4 nM; S.I. = 1.69). Compounds with ethoxycarbonyl (5)
or aromatic (11, 12) moiety at position 2 or alkyne (14)
moiety at position 3 showed lower inhibition of COX-1 and
COX-2 with lower selectivity value than celecoxib (5; COX-
2 IC50 = 7.8 nM; COX-1 IC50 = 9.2 nM; S.I. = 1.18, 11;
COX-2 IC50 = 3.9 nM; COX-1 IC50 = 6.4 nM; S.I. =
1.64, 12; COX-2 IC50 = 6.8 nM; COX-1 IC50 = 8.7 nM;
S.I. = 1.38, 14; COX-2 IC50 = 5.3 nM; COX-1 IC50 =
7.5 nM; S.I. = 1.42).
Molecular docking
˚
COX-1 and 2 monomers each contain a 25-A hydrophobic
channel that originates at the membrane binding domain
(MBD) and extends into the core of the globular domain
(Picot et al. 1994; Kurumbail et al. 1996; Luong et al.
1996). The MBD forms the mouth and the first half of the
channel. The NSAID binding site involves the upper half of
this channel from Arg120 to near Tyr385. It is clear that the
NSAID binding site corresponds to the cyclooxygenase
active site (Rome and Lands 1975; Smith and DeWitt
1996). Several amino acids composing the upper half of
the channel are uniquely important in cyclooxygenase
catalysis. Twenty-four residues line the hydrophobic
123

Triazoles: synthesis and docking studies
559
Table 1 Percent inhibition of
Compounds
AI activity
IC50 (nM)
COX-1
Selectivity
index
(COX-1/COX-2)
carrageenan paw edema (CPE)
and COX-1 and COX-2
inhibitor activity of the
compounds 5–16
% inhibition at 1 h
% inhibition at 2 h
1.8
COX-2
5
0.0
5
9.2
3.5
3.1
3.8
3.7
4.5
6.4
8.7
5.3
7.5
3.7
4.4
4.1
7.8
0.8
0.9
1.1
0.9
2.1
3.9
6.8
2.1
5.3
1.4
2.6
1.9
1.18
4.38
3.44
3.46
4.11
2.14
1.64
1.38
2.52
1.42
2.64
1.69
2.16
6
25 1.8
51 1.5
45 1.1
62 3.4
0
66 2.6
76 2.8
66 1.2
66 1.5
16 5.9
29 3.8
19 6.3
61 4.1
12 4.9
50 2.5
38 1.8
71.5 3.7
7
8
9
10
11
12.5 1.6
16 5.7
46 2.9
10 5.4
36 2.1
12
13
14
15
16
9
1.3
Celecoxib
50 2.1
cyclooxygenase active site with only one difference
between the isozymes—Ile at position 523 in COX-1 and
Val at position 523 in COX-2. (Amino acids lining the
hydrophobic cyclooxygenase active site channel include
Leu117, Arg120, Phe205, Phe209, Val344, Ile345, Tyr348,
Val349, Leu352, Ser353, Tyr355, Leu359, Phe381,
Leu384, Tyr385, Trp387, Phe518, Ile/Val523, Gly526,
Ala527, Ser530, Leu531, Gly533, Leu534.) Only three
of the channel residues are polar (Arg120, Ser353, and
Ser530). Ser530 is the site of acetylation by aspirin,
(DeWitt et al. 1990; Lecomte et al. 1994; Loll et al. 1995)
and Arg120 binds to the carboxylate groups of fatty acids
and many NSAIDs (Greig et al. 1997; Kulmacz 1987;
Mancini et al. 1995; Rouzer and Marnett 2003). The
mechanism for a differential inhibition by classical and
COX-2 NSAIDs can be rationalized to some extent based on
differences between the cyclooxygenase active sites of
COX-1 and 2. Substitution of Ile523 in COX-1 with Val523
in COX-2 results in the presence of a small side pocket
adjacent to the active site channel, appreciably increasing the
volume of the COX-2 active site (Luong et al. 1996). This
change is compounded by the substitution of Ile434 in COX-
1 with Val434 in COX-2, within the second shell of amino
acids surrounding the cyclooxygenase active site. The Ile to
Val substitution at position 434 outside the COX-2 catalytic
centre further increases the effective size of the active site
channel by enhancing the local mobility of side chains within
the side pocket. The combination of these two differences at
positions 523 and 434 in COX-2 causes a movement of
Phe518 that further increases the size of the side pocket. The
larger main channel combined with the extra nook increases
the volume of the COX-2 NSAID binding site by about 20 %
over that in COX-1 (Luong et al. 1996). This extra size is a
structural feature exploited by COX-2 inhibitors. Finally, the
substitution of His513 in COX-1 with Arg513 in COX-2
results in a stable positive charge being placed at the centre of
this pocket, which can interact with polar moieties entering
the pocket (Kurumbail et al. 1996). For example, Arg513
appears to interact with the 4-methysulfonyl or 4-sulfona-
moylphenyl substituents of diaryl heterocyclic COX-2
inhibitors.
Docking studies of the synthesized compounds 5–16 in
the active sites of both COX-1 (PDB code: 1CQE) (Picot
et al. 1994) and COX-2 (PDB code: 1CX2) (Kurumbail
et al. 1996) were performed in order to get further insight
into the nature of interactions between the compounds and
the active site amino acids to rationalize the obtained
biological results. For validation of our docking procedure
Fig. 1 COX-1: the docked co-crystallized flurobiprofen (from
1CQE.pdb, coloured cyan). Hydrogen bonds are displayed in magenta
123

560
A. A. Radwan, K. E. H. elTahir
their in vitro screening results against the enzymes. While
compound 5 showed similar binding mode and hydrogen
bonding with high docking score at COX-1 and COX-2 as
well as flurbiprofen and sc-558. This was not in agreement
with its lower in vitro activity against both COX-1 and
COX-2.
Whether this binding results or not into a functional
inhibition is more complicated to predict since it may be
related to kinetic of dissociation rather than to the stability
of the complex. The failure of static docking experiments
in predicting the inactivity of analogues, closely related to
COX-2 active diaryl heterocyclic derivatives can be com-
mented on the light of a recent paper by Limongell et al.
2010 who put forward not only the importance of the sta-
bility of the binding mode, but also the importance of the
dynamic path of ligand(s) to reach the binding pose. In this
context, Limongell et al. 2010 proposed the existence of
alternative binding modes of diaryl heterocyclic derivatives
to COX-2. It can be speculated that the reason for the
inactivity of some compounds may reside in their inability
to achieve one or more alternative binding modes possibly
relevant to functional inhibition (Limongell et al. 2010).
As examples the docking solution obtained for com-
pounds 9 and 15 at COX-2 are shown in Figs. 3 and 4. The
docked compounds fit well into the binding pocket and show
hydrogen bonds with Arg513. The pyridine moiety at the C-5
position of the triazole ring fits into the COX-2 secondary
pocket and makes hydrogen bond between its ring-nitrogen
and Arg513, showing a similar interaction as the sulfon-
amide group of SC-588. The phenyl ring at C-4 position
of the triazole ring occupies the same position of
Fig. 2 COX-2: the docked co-crystallized SC-558 (from 1CX2.pdb,
coloured cyan). Hydrogen bonds are displayed in yellow
Table 2 Calculated docking score for COX-1 and COX-2
Compounds
Total docking score
COX-1
COX-2
5
5.42
4.59
5.56
6.21
4.89
6.61
4.25
4.31
5.63
6.13
7.56
7.23
4.31
6.8
7.87
6.63
8.60
9.37
8.41
8.10
4.64
5.74
6.51
6.31
8.36
9.15
7.99
6.19
6
7
8
9
10
11
12
13
14
15
16
Celecoxib
Flurbiprofen
flurbiprofen and sc-558 were docked inside 1CQE and
1CX2 respectively. The docking results of 1CQE and
1CX2 were able to reproduce the experimentally observed
interaction mode (Limongell et al. 2010) with rmsd values
between experimentally used and docked ligand structure
of 0.31 and 0.42 respectively (Figs. 1, 2). Table 2 shows
that compounds 6–10 and 13, 15, 16 explored comparable
interactions, with high docking score at COX-1 and COX-
2, with that of the co-crystallized flurbiprofen and sc-558
while compounds 11, 12, 14 explored lower docking score
value than that of flurbiprofen and sc-558. Docking score
values of compounds 6–16 were found in agreement with
Fig. 3 COX-2: the docked compound 9 (coloured orange red).
Hydrogen bond is displayed in yellow
123

Triazoles: synthesis and docking studies
561
Fig. 6 COX-1: the docked compound 15 (coloured magenta)
pyrrolidinylmethyl substituent occupied the same position
of the terminal aromatic ring of flurbiprofen.
Fig. 4 COX-2: the docked compound 15 (coloured magenta).
Hydrogen bond is displayed in yellow
Conclusions
In summary, a series of triazole compounds were synthe-
sized and biologically evaluated. Some of the synthesized
derivatives were more potent and more selective on COX-2,
compared to celecoxib. In animal model, compound 7 was
slightly more active and compounds 6, 8, 9 were slightly
less active than celecoxib, reference drug. However,
compounds 5 and 10 almost were biologically inactive in
animal model. Compounds 6–9 and 15 explored higher
anti-COX-2 activity and selectivity than that of celecoxib.
Compounds 10 and 13 explored same anti-COX potency
and COX-2 selectivity, as well as celecoxib. Compounds 5,
11, 12 and 14 showed lower anti-COX activity with lower
COX-2 selectivity.
Molecular docking was further performed to study the
inhibitor-COX protein interactions. After analysis of the
binding model of compounds 9 and 15 with COX-2, it was
found that these compounds occupied same orientation as
well as sc-558 and showed hydrogen with Arg513 in COX-2
binding site. The analysis of their binding model with
COX-1, it was found that these compounds fitted well to
the same binding sites of flurbiprofen with an additional
hydrophobic interaction with Ile523 while did not show
any hydrogen interaction. Among these compounds, it
could be concluded that compound 9 had been demon-
strated to show significant selective COX-2 inhibitory
activity as a potential antiinflammatory agent. The result of
this work might be helpful for the design and synthesis of
more selective COX-2 inhibitors with stronger activity.
Fig. 5 COX-1: the docked compound 9 (coloured orange red)
trifluoromethyl of sc-558, the N-2 or C-3 substituent could
occupy the same position of the the bromophenyl ring of
sc-558. As examples the docking solution obtained for
compounds 9 and 15 at COX-1 are shown in Figs. 5 and 6.
The docked compounds fitted well into the binding pocket
while did not show hydrogen bonding with Arg120.
The central triazole ring adopts the same position as the
central aromatic ring of flurbiprofen. The S-pent-2-yl or
123

562
A. A. Radwan, K. E. H. elTahir
Acknowledgments The authors are grateful to the sponsorship of
the Research Centre, College of Pharmacy and the Deanship of Sci-
entific Research, King Saud University, Riyadh 11451, Saudi Arabia.
Limongell, V., M. Bonomi, L. Marinelli, F.L. Gervasio, A. Cavalli,
E. Vovellino, and M. Parrinello. 2010. Molecular basis of
cyclooxygenase enzymes (COXs) selective inhibition. PNAS
107: 5411–5416.
Loll, P.J., D. Picot, and R.M. Garavito. 1995. The structural basis of
aspirin activity inferred from the crystal structure of inactivated
prostaglandin H2 synthase. Natural Structural Biology 2:
637–643.
References
Luong, C., A. Miller, J. Barnett, J. Chow, C. Ramesha, and M.F.
Browner. 1996. Flexibility of the NSAID binding site in the
structure of human cyclooxygenase-2. Natural Structural Biol-
ogy 3: 927–933.
Mancini, J.A., D. Riendeau, J.P. Falgueyret, P.J. Vickers, and G.P.
O’Neill. 1995. Arginine 120 of prostaglandin G/H synthase-1 is
required for the inhibition by nonsteroidal anti-inflammatory
drugs containing a carboxylic acid moiety. Journal of Biological
Chemistry 270: 29372–29377.
AbdelMoallem, S., F. Hadizadeh, and I. Yavar. 2011. New N-aryl-4-
(methysulfony)aminobenzenesulfonamides as selective COX-2
inhibitors. Journal of Biological Sciences 11: 496–499.
Ali, A.M., G.E. Saber, N.M. Mahfouz, M.A. EI-Gendy, A.A. Radwan,
and M.A. Hamid. 2007. Synthesis and three-dimensional qual-
itative structure selectivity relationship of 3,5-disubstituted-2,4-
thiazolidinedione derivatives as COX-2 inhibitors. Archives of
Pharmacal Research 30: 1186–1204.
Bayrak, H., A. Demirbas, S.A. Karaoglu, and N. Demirbas. 2009.
Synthesis of some new 1,2,4-triazoles, their Mannich and Schiff
bases and evaluation of their antimicrobial activities. European
Journal of Medicinal Chemistry 44: 1057–1066.
Catella-Lawson, F., and L.J. Crofford. 2001. Cyclooxygenase inhi-
bition and thrombogenicity. American Journal of Medicine 110:
28S–32S.
Meade, E.A., W.L. Smith, and D.L. DeWitt. 1993. Differential
inhibition of prostaglandin endoperoxide synthase (cyclooxy-
genase) isozymes by aspirin and other non-steroidal anti-
inflammatory drugs. Journal of Biological Chemistry 268:
6610–6614.
Moreau, A., P.N.P. Rao, and E.E. Knaus. 2006. Synthesis and
biological evaluation of acyclic triaryl (Z)-olefins possessing a
3,5-di-tert-butyl-4-hydroxyphenyl pharmacophore: dual inhibi-
tors of cyclooxygenases and lipoxygenases. Bioorganic &
Medicinal Chemistry 14: 5340–5350.
Clark, R.D. 2008. A ligand’s-eye view of protein binding. Journal of
Computer-Aided Molecular Design 22: 507–521.
Dannhardt, G., and S. Laufer. 2000. Structural approaches to explain
the selectivity of COX-2 inhibitors: is there a common pharma-
cophore? Current Medicinal Chemistry 7: 1101–1112.
DeGaetano, G., M.B. Donati, and C. Cerletti. 2003. Prevention of
thrombosis and vascular inflammation: benefits and limitations
of selective or combined COX-1, COX-2 and 5-LOX inhibitors.
Trends in Pharmacological Sciences 24: 245–252.
DeWitt, D.L., E.A. el-Harith, S.A. Kraemer, M.J. Andrews, E.F. Yao,
R.L. Armstrong, and W.L. Smith. 1990. The aspirin and heme-
binding sites of ovine and murine prostaglandin endoperoxide
synthases. Journal of Biological Chemistry 265: 5192–5198.
Greig, G.M., D.A. Francis, J.P. Falgueyret, M. Ouellet, M.D. Percival,
P. Roy, C. Bayly, J.A. Mancini, and G.P. O’neill. 1997. The
interaction of arginine 106 of human prostaglandin G/H
synthase-2 with inhibitors is not a universal component of
inhibition mediated by nonsteroidal anti-inflammatory drugs.
Molecular Pharmacology 52: 829–838.
Jain, A.N. 2003. Surflex: fully automatic flexible molecular docking
using a molecular similarity-based search engine. Journal of
Medicinal Chemistry 46: 499–511.
Kulmacz, R.J., and W.E.M. Lands. 1983. Requirements for hydro-
peroxide by the cyclooxygenase and peroxidase activities of
prostaglandin H synthase. Prostaglandins 25: 531–540.
Kulmacz, R.J. 1987. Prostaglandin G2 levels during reaction of
prostaglandin H synthase with arachidonic acid. Prostaglandins
34: 225–240.
Mukherjee, D., S.E. Nissen, and E.J. Topol. 2001. Risk of cardio-
vascular events associated with selective COX-2 inhibitors.
JAMA 286: 954–959.
´
Muthas, D., Y.A. Sabnis, M. Lundborg, and A. Karlen. 2008. Is it
possible to increase hit rates in structure-based virtual screening
by pharmacophore filtering? An investigation of the advantages
and pitfalls of post-filtering. Journal of Molecular Graphics and
Modelling 26: 1237–1251.
Navidpour, L., H. Shafaroodi, K. Abdi, M. Amini, M.H. Ghahremani,
A.R. Dehpour, and A. Shafiee. 2006. Design, synthesis, and
biological evaluation of substituted 3-alkylthio-4,5-diaryl-4H-
1,2,4-triazoles as selective COX-2 inhibitors. Bioorganic &
Medicinal Chemistry 14: 2507–2517.
Penning, T.D., J.J. Tally, S.R. Bertenshaw, J.S. Carter, P.W. Collins,
S. Docter, M.J. Graneto, L.F. Lee, J.W. Malecha, J.M. Miyashiro,
D.J. Rogers, S.S. Rogier, G.D. Yu, E.G. Anderson, J.N. Burton,
S.A. Cogburn, C.M. Gregory, R.S. Koboldt, W.E. Perkins,
K. Seibert, A.W. Veenhuizen, Y.Y. Zhang, and P.C. Isakson.
1997. Synthesis and biological evaluation of the 1,5-diarylpyraz-
ole class of cyclooxygenase-2 inhibitors: identification of 4-[5-(4-
methylphenyl)-3-(trifluoromethyl)-1H-pyrazol-1-yl]benze nesulf-
onamide (SC-58635, celecoxib). Journal of Medicinal Chemistry
40: 1347–1365.
Picot, D., P.J. Loll, and R.M. Garavito. 1994. The X-ray crystal
structure of the membrane protein prostaglandin H2 synthase-1.
Nature 367: 243–249.
Plount-Price, M.L., and W.L. Jorgensen. 2000. Analysis of binding
affinities for celecoxib analogues with COX-1 and COX-2 from
combined docking and Monte Carlo simulations and insight into
the COX-2/COX-1 selectivity. Journal of the American Chem-
ical Society 122: 9455–9466.
Kurumbail, R.G., A.M. Stevens, J.K. Gierse, J.J. McDonald, R.A.
Stegeman, J.Y. Pak, D. Gildehaus, J.M. Miyashiro, T.D.
Penning, K. Seibert, P.C. Isakson, and W.C. Stallings. 1996.
Structural basis for selective inhibition of cyclooxygenase-2 by
anti-inflammatory agents. Nature 384: 644–648.
Labanauskas, L., V. Kalcas, E. Udrenaite, P. Gaidelis, A. Brukstus,
and U. Dauksas. 2001. Synthesis of 3-(3,4-dimethoxyphenyl)-1H-
1,2,4-triazole-5-thiol and 2-amino-5-(3,4-dimethoxyphenyl)-1,3,
4-thiadiazole derivatives exhibiting anti-inflammatory activity.
Pharmazie 56: 617–619.
Pomarnacka, E., and A. Kornicka. 2001. Synthesis and in vitro
anticancer and anti-HIV evaluation of new 2-mercaptobenzene-
sulfonamides. Il Farmaco 56: 571–577.
Praveen, R.P.N., A. Mohsen, L. Huiying, G.H. Amgad, and
E.K. Edward. 2003. Design, synthesis and biological evaluation
of 6-substituted-3-(4-methanesulfonylphenyl)-4-phenylpyran-2-
ones: a novel class of diarylheterocyclic selective cyclooxygen-
ase-2 inhibitors. Journal of Medicinal Chemistry 46: 4872–4882.
Lecomte, M., O. Laneuville, C. Ji, D.L. DeWitt, and W.L. Smith.
1994. Acetylation of human prostaglandin endoperoxide syn-
thase-2 (cyclooxygenase-2) by aspirin. Journal of Biological
Chemistry 269: 13207–13215.
123

Triazoles: synthesis and docking studies
563
Ranatunge, R.R., R.A. Earl, D.S. Garvey, D.R. Janero, L.G. Letts,
A.M. Martino, M.G. Murty, S.K. Richardson, D.J. Schwalb,
D.V. Young, and I.S. Zemtseva. 2004. 3-(2-Methoxytetrahydro-
furan-2-yl)pyrazoles: a novel class of potent, selective cycloox-
1,3-Diarylcycloalkanopyrazoles and diphenyl hydrazides as
selective inhibitors of cyclooxygenase-2. Bioorganic & Medici-
nal Chemistry Letters 10: 601–614.
SYBYL Molecular Modelling Software. 2006. Tripos Inc., St. Louis,
http://www.tripos.com. Accessed July 2010.
ygenase-2 (COX-2) inhibitors. Bioorganic
Chemistry Letters 14: 6049–6052.
& Medicinal
Talley, J.J., D.L. Brown, J.S. Carter, M.J. Graneto, C.M. Koboldt, J.L.
Masferrer, W.E. Perkins, R.S. Rogers, A.F. Shaffer, Y.Y. Zhang,
B.S. Zweifel, and K. Seibert. 2000. 4-[5-Methyl-3-phenylisox-
azol-4-yl]-benzenesulfonamide, valdecoxib: a potent and selec-
tive inhibitor of COX-2. Journal of Medicinal Chemistry 43:
775–777.
Tanaka, Y., S.L. xWard, and W.L. Smith. 1987. Immunohistochem-
ical and kinetic evidence for two different prostaglandin
H-prostaglandin E isomerases in sheep vesicular gland micro-
somes. Journal of Biological Chemistry 262: 1374–1381.
Tozkoparan, B., E. Kupeli, E. Yesilada, S. Isik, M. Ozalp, and M.
Ertan. 2005. Synthesis and evaluation of analgesic/anti-inflam-
matory and antimicrobial activities of 3-substituted-1,2,4-tria-
zole-5-thiones. Arzneimittelforschung 55: 533–540.
Vera, K., Z. Lenka, W. Karel, K. Jamila, and M. Ute. 2004. Synthesis
and antimycobacterial activity of 1,2,4-triazole 3-benzylsulfanyl
derivatives. Il Farmaco 59: 279–288.
Winter, C.A., E.A. Risley, and G.W. Nuss. 1962. Carrageenin-induced
edema in hind paw of the rat as an assay for antiinflammatory
drugs. Proceedings of the Society for Experimental Biology and
Medicine 111: 544–547.
Wolfe, M.M., D.R. Lichtenstein, and G. Singh. 1999. Gastrointestinal
toxicity of nonsteroidal antiinflammatory drugs. New England
Journal of Medicine 340: 1888–1899.
Wujec, M., and P. Paneth. 2008. Mechanism of 4-methyl-1,2,4-triazol-
3-thione reaction with formaldehyde. Journal of Physical Organic
Chemistry 21: 345–348.
Wujec, M., M. Pitucha, M. Dobosz, U. Kosikowska, and A. Malm.
2003. Synthesis and biological action of 1-substituted-3-R-4-
phenyl-delta 2–1,2,4-triazoline-5-thion. Acta Poloniae Pharma-
ceutica 60: 451–456.
Zarghi, A., F.S. Javid, R. Ghodsi, O.G. Dadrass, B. Daraei, and
M. Hedayati. 2011. Design, synthesis and biological evaluation
of new 5,5-diarylhydantoin derivatives as selective cyclooxy-
genase-2 inhibitors. Scientia Pharmaceutica 79: 449–460.
Riendeau, D., M.D. Percival, C. Brideau, C.S. Dube, D. Ethier,
J.P. Falgueyret, R.W. Friesen, R. Gordon, G. Greig, J. Guay,
Y. Girard, P. Prasit, R. Zamboni, I.W. Rodger, M. Gresser,
A.W. Ford-Hutchinson, R.N. Young, and C.C. Chan. 2001.
Etoricoxib (MK-0663): preclinical profile and comparison with
other agents that selectively inhibit cyclooxygenase-2. Journal of
Pharmacology and Experimental Therapeutics 296: 558–566.
Rome, L.H., and W.E. Lands. 1975. Structural requirements for time-
dependent inhibition of prostaglandin biosynthesis by anti-
inflammatory drugs. Proceedings of the National Academy of
Sciences of the United States of America 72: 4863–4865.
Rouzer, C.A., and L.J. Marnett. 2003. Mechanism of free radical
oxygenation of polyunsaturated fatty acids by cyclooxygenases.
Chemical Reviews 103: 2239–2304.
Ruppert, J., W. Welch, and A.N. Jain. 1997. Automatic identification
and representation of protein binding sites for molecular
docking. Protein Science 6: 524–533.
Sano, H., T. Noguchi, A. Tanatani, Y. Hashimoto, and H. Miyachi.
2005. Design and synthesis of subtype-selective cyclooxygenase
(COX) inhibitors derived from thalidomide. Bioorganic &
Medicinal Chemistry 13: 3079–3091.
Silverstein, F.E., G. Faich, J.L. Goldstein, L.S. Simon, T. Pincus,
A. Whelton, R. Makuch, G. Eisen, N.M. Agrawal, W.F. Stenson,
A.M. Burr, W.W. Zhao, J.D. Kent, J.B. Lefkowith, K.M.
Verburg, and G.S. Geis. 2000. Gastrointestinal toxicity with
celecoxib vs nonsteroidal anti-inflammatory drugs for osteoar-
thritis and rheumatoid arthritis: the CLASS Study: a randomized
controlled trial. JAMA 284: 1247–1255.
Smith, W.L., and D.L. DeWitt. 1996. Prostaglandin endoperoxide H
synthases-1 and -2. Advances in Immunology 62: 167–215.
Srimal, R.C., and B.N. Dhawan. 1974. Biological, toxicological and
clinical evaluation. Indian Journal of Pharmacology 3: 4–13.
Sui, Z., J. Guan, M.P. Ferro, K. McCoy, M.P. Wachter, W.W. Murray,
M. Singer, M. Steber, D.M. Ritchie, and D.C. Argentieri. 2000.
123