Structure-based design, synthesis, molecular docking study and biological evaluation of 1,2,4-triazine derivatives acting as COX/15-LOX inhibitors with anti-oxidant activities

Article abstract of DOI:10.3109/14756366.2016.1158713

A set of 1,2,4-triazine derivatives were designed as cyclooxygenase-2 (COX-2) inhibitors. These compounds were synthesized and screened for inhibition of cyclooxygenases (COX-1 and COX-2) based on a cellular assay using human whole blood (HWB) and lipoxygenase (LOX-15) that are key enzymes in in?ammation. The results showed that 3-(2-(benzo[d][1,3]dioxol-5-ylmethylene)hydrazinyl)-5,6-bis(4-methoxyphenyl)-1,2,4-triazine (G11) was identified as the most potent COX-2 inhibitor (78%) relative to COX-1 (50%). Ferric reducing anti-oxidant power (FRAP) assay revealed that compound G10 possesses the highest anti-oxidant activity. The compound G3 with IC₅₀ value of 124 μM was the most potent compound in LOX inhibitory assay. Molecular docking was performed and a good agreement was observed between computational and experimental results.

Full text of DOI:10.3109/14756366.2016.1158713

Journal of Enzyme Inhibition and Medicinal Chemistry
ISSN: 1475-6366 (Print) 1475-6374 (Online) Journal homepage: http://www.tandfonline.com/loi/ienz20
Structure-based design, synthesis, molecular
docking study and biological evaluation of
1,2,4-triazine derivatives acting as COX/15-LOX
inhibitors with anti-oxidant activities
Mehdi Khoshneviszadeh, Omolbanin Shahraki, Mahsima Khoshneviszadeh,
Alireza Foroumadi, Omidreza Firuzi, Najmeh Edraki, Hamid Nadri, Alireza
Moradi, Abbas Shafiee & Ramin Miri
To cite this article: Mehdi Khoshneviszadeh, Omolbanin Shahraki, Mahsima Khoshneviszadeh,
Alireza Foroumadi, Omidreza Firuzi, Najmeh Edraki, Hamid Nadri, Alireza Moradi, Abbas
Shafiee & Ramin Miri (2016): Structure-based design, synthesis, molecular docking study and
biological evaluation of 1,2,4-triazine derivatives acting as COX/15-LOX inhibitors with anti-
oxidant activities, Journal of Enzyme Inhibition and Medicinal Chemistry
To link to this article: http://dx.doi.org/10.3109/14756366.2016.1158713
Published online: 30 Mar 2016.
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Date: 30 March 2016, At: 21:10

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ISSN: 1475-6366 (print), 1475-6374 (electronic)
J Enzyme Inhib Med Chem, Early Online: 1–10
2016 Informa UK Limited, trading as Taylor & Francis Group. DOI: 10.3109/14756366.2016.1158713
!
RESEARCH ARTICLE
Structure-based design, synthesis, molecular docking study and
biological evaluation of 1,2,4-triazine derivatives acting as COX/15-LOX
inhibitors with anti-oxidant activities
Mehdi Khoshneviszadeh1,2, Omolbanin Shahraki2, Mahsima Khoshneviszadeh1, Alireza Foroumadi3,
Omidreza Firuzi1, Najmeh Edraki1, Hamid Nadri4, Alireza Moradi4, Abbas Shafiee5, and Ramin Miri1
1Medicinal and Natural Products Chemistry Research Center, Shiraz University of Medical Sciences, 2Department of Medicinal Chemistry, Faculty of
3
Pharmacy, Shiraz University of Medical Sciences, Shiraz, Iran, Drug Design and Development Research Center, Tehran University of Medical
Sciences, Tehran, Iran, 4Department of Medicinal Chemistry, Faculty of Pharmacy, Shahid Sadoughi University of Medical Sciences, Yazd, Iran, and
5Department of Medicinal Chemistry, Faculty of Pharmacy and Pharmaceutical Sciences Research Center, Tehran University of Medical Sciences,
Tehran, Iran
Abstract
Keywords
A set of 1,2,4-triazine derivatives were designed as cyclooxygenase-2 (COX-2) inhibitors. These
compounds were synthesized and screened for inhibition of cyclooxygenases (COX-1 and
COX-2) based on a cellular assay using human whole blood (HWB) and lipoxygenase (LOX-15)
that are key enzymes in inFammation. The results showed that 3-(2-(benzo[d][1,3]dioxol-5-
ylmethylene)hydrazinyl)-5,6-bis(4-methoxyphenyl)-1,2,4-triazine (G11) was identified as the
most potent COX-2 inhibitor (78%) relative to COX-1 (50%). Ferric reducing anti-oxidant power
(FRAP) assay revealed that compound G10 possesses the highest anti-oxidant activity. The
compound G3 with IC50 value of 124 mM was the most potent compound in LOX inhibitory
assay. Molecular docking was performed and a good agreement was observed between
computational and experimental results.
Cyclooxygenase, FRAP assay, 15-lipoxygen-
ase, molecular modeling, 1,2,4-triazine
History
Received 25 December 2015
Revised 13 February 2016
Accepted 15 February 2016
Published online 23 March 2016
Introduction
Non-steroidal anti-inflammatory drugs (NSAIDs) are amongst
the most widely used drugs worldwide. Traditional NSAIDs
(classical NSAIDs or nonselective COX inhibitors), such as
ibuprofen, indomethacin and naproxen inhibit both COX-1 and
COX-2. Several serious side effects, such as gastrointestinal irritation
and hemorrhage and kidney failure are caused by these agents5.
The discovery of molecules that selectively inhibit COX-2,
named coxibs (celecoxib, rofecoxib and valdecoxib) has been a
response to the necessity of lowering these side effects. Indeed,
the selectivity of this class of drugs resulted in improved
gastrointestinal tolerance. Consecutive clinical studies revealed
that long term use of several coxibs is accompanied by increased
risk level of cardiovascular problems, such as myocardial
infarction, stroke, systemic and pulmonary hypertension, and
congestive heart failure6. Hence, finding effective and safe anti-
inflammatory drugs seems to be an urgent necessity.
In previous investigations, we have reported some new
compounds as anti-inflammatory agents. As a part of our ongoing
effort to develop effective candidates for inhibition of inflamma-
tion, a set of 5,6-diaryl-1,2,4-triazine derivatives were designed7.
In contrast with the traditional NSAIDs which belong to
different categories of chemical structures, selective COX-2
inhibitors can be classified into three groups (Figure 1): (1)
diaryl ethers with methanesulfonamide at position-2 of one ring,
Nemisulide and CGP-28237 can be considered as lead compounds
in this class8; (2) trans-stilbenoids, such as resveratrol, a natural
product found in grapes and other food products9; (3) heterocyclic
Inflammation, a process whereby a huge number of mediators are
released due to over-activation of different cascades, is associated
with many diseases, such as rheumatoid arthritis (RA), osteo-
arthritis (OA), bronchospasm and psoriasis1. Enzymatic action of
phospholipases A2 (PLA2s) on phospholipid bilayer results in
arachidonic acid release, which is a precursor in the production of
eicosanoids including prostaglandins (PGs) and leukotrienes
(LTs)2. The transformation of arachidonic acid to PGs is mediated
by cyclooxygenase (COX) enzyme family (COX-1, COX-2 and
COX-3). The most important isoforms are COX-1 and COX-2,
whereas, COX-3 is expressed in the brain and its biological role is
still unknown.
COX-1 is the constitutive form of the enzyme and is present in
the stomach, intestines, kidneys and platelets which causes
gastrointestinal cytoprotection as well as renal effects. On the
other hand, COX-2 is constitutively expressed only in low
amounts in kidney, brain and ovaries, but its expression is
considerably enhanced during inflammatory situations or mito-
genic stimulation3,4
.
Address for correspondence: Ramin Miri, Medicinal and Natural Products
Chemistry Research Center, Shiraz University of Medical Sciences,
Shiraz, Iran. Tel: +98 7132307869. Fax: +98 7132302225. E-mail:
mirir@sums.ac.ir

2
M. Khoshneviszadeh et al.
J Enzyme Inhib Med Chem, Early Online: 1–10
Figure 1. Different classes of selective COX-
2 inhibitors.
O
O
O
O
H3C
N
S
S
H2N
H2N
N
N
N
N
N
CF3
CF3
Cl
O
S
H3C
O
Br
H3C
Etoricoxib
Celecoxib
SC-558
O
O
O
O
S
S
NH
NH
H3C
H3C
O
O
NO2
HO
O
CGP-28237
nimesulide
OH
Resveratrol
OH
systems (such as 1,2,4 triazole10,11, imidazole12, pyrazol13, 3-(2-Benzylidenehydrazinyl)-5,6-bis(4-methoxyphenyl)1,2,4-tria-
quinoline14, indol15, pyrimidine16, 1,2,4-triazine17,18) with adja- zine (G1)7
cent diaryl containing sulfamoyl (SO2NH2) or methylsulfonyl
Compound G1 was prepared by the described procedure using
(SO2Me) substitution at position-4 of one ring which exist in
106 mg (1 mmol) of benzaldehyde (yield 83%), m.p.: 245–247 ꢀC;
celecoxib, SC-558 and etoricoxib.
1H NMR (DMSO-d6, 500 MHz), dH: 10.29 (s, 1H, NH), 8.24 (s, 1H,
The compounds bearing 1,2,4-triazine core have been
N¼CH), 7.81 (d, J ¼ 7.1 Hz, 2H, ArH), 7.66 (d, J ¼ 8.8 Hz, 2H,
identified to exhibit a broad spectrum of biological effects, such
ArH), 7.50 (d, J ¼ 8.7 Hz, 2H, ArH), 7.43–7.38 (m, 3H, ArH), 6.93
(d, J ¼ 8.7 Hz, 2H, ArH), 6.88 (d, J ¼ 8.7 Hz, 2H, ArH), 3.86 ppm
(s, 6H, 2 ꢁ O–CH3H); IR (KBr, ꢀ): 3440, 3211, 1605, 1571,
831 cmꢂ1; MS (EI) m/z (%): 411.2 (M+, 29), 334 (20), 308 (14), 238
(100), 223 (49), 195 (12), 152 (11). Anal. Calcd. for C24H21N5O: C,
70.06; H, 5.14; N, 17.02; found: C, 70.21; H, 4.86; N, 16.95.
as antifungal19, anti-HIV20, adenosine A2A receptor antagonist21,
kinase inhibitor antitumor22,23, analgesic17, anti-anxiety and anti-
inflammation effects24.
As a part of our research program toward the search of potent
COX-2 inhibitors with lower side effects, improved affinity and
efficacy and dual modes of action, molecular hybridization
strategy was employed to design 1,2,4-triazine derivatives. After
the synthesis of compounds, in vitro assays and molecular
docking study were also performed.
3-(2-(4-Nitrobenzylidene)hydrazinyl) 5,6-bis(4-methoxyphenyl)-
1,2,4-triazine (G2)7
The titled compound was synthesized by the described procedure
from 4-nitrobenzaldehyde (151 mg, 1 mmol) as yellow solid (yield
88%), m.p.: 265–267 ꢀC; 1H NMR (DMSO-d6, 500 MHz), dH:
12.23 (s, 1H, NH), 8.36 (s, 1H, N¼CH), 8.31 (d, J ¼ 8.6 Hz, 2H,
Experimental protocols
Chemistry
NMR spectra were recorded on Brucker 500 spectrometer or ArH), 7.99 (d, J ¼ 8.0 Hz, 2H, ArH), 7.52 (d, J ¼ 8.3 Hz, 2H,
Varian unity 400 spectrometer relative to tetramethylsilane (TMS) ArH), 7.39 (d, J ¼ 8.6 Hz, 2H, ArH), 6.98–6.96 (m, 4H, ArH),
as an internal standard using CDCl3 or dimethyl sulfoxide 3.79 ppm (s, 6H, 2 ꢁ O–CH3); IR (KBr, ꢀ): 3421, 3205, 1607,
(DMSO)-d6 as the solvents. The mass spectra were obtained on an 1585, 838 cmꢂ1; MS (EI) m/z (%): 456 (M+, 21), 308 (16), 238
Agilent 6410 instrument. The IR spectra were recorded on a FT- (100), 223 (25), 195(11). Anal. Calcd. for C24H20N6: C, 63.15; H,
IR Perkin-Elmer spectrophotometer (KBr disks). Melting points 4.42; N, 18.41; found: C, 62.78; H, 4.12; N, 18.51.
were determined on a KoFer hot stage apparatus and are
uncorrected. Elemental analyses were performed on a CHN–O– 3-(2-(4-Fluorobenzylidene)hydrazinyl)-5,6-bis(4methoxyphenyl)-
1,2,4-triazine (G3)7
rapid elemental analyzer (Heraeus, Hanau, Germany).
The titled compound G3 was synthesized by the general
procedure using 4-Fuorobenzaldehyde (124 mg, 1 mmol) (yield
General procedure
All 3-(2-arylidenehydrazinyl) 5,6-bis(4-methoxyphenyl)-1,2,4- 72%), m.p.: 257–259 ꢀC; 1H NMR (DMSO-d6, 500 MHz), dH:
triazine derivatives were prepared according to the procedure 11.64 (s, 1H, NH), 8.25 (s, 1H, N¼CH), 8.06–8.02 (m, 2H, ArH),
described7.
7.49 (d, J ¼ 8.5 Hz, 2H, ArH), 7.35 (d, J ¼ 8.5 Hz, 2H, ArH),

DOI: 10.3109/14756366.2016.1158713
1,2,4-Triazine derivatives
3
7.30–7.26 (m, 2H, ArH), 6.96–6.93 (m, 4H, ArH), 3.78 ppm (s, a pale yellow solid (yield 78%), m.p.: 248–251 ꢀC 1H NMR
6H, 2 ꢁ O–CH3); 13C NMR (DMSO-d6, 125 MHz), dC ¼ 163.39, (DMSO-d6, 500 MHz), ꢁH: 12.34 (s, 1H, NH), 8.74 (s, 1H,
161.45, 160.99, 159.39, 158.21, 155.45, 150.64, 141.87, 137.48, N¼CH), 7.82 (d, J ¼ 3.9 Hz, 1H, ArH), 7.49–7.47 (m, 2H, ArH),
131.19, 130.86, 130.80, 130.21, 128.63, 127.87, 122.86, 116.03, 7.39–7.37 (m, 2H, ArH), 7.21 (d, J ¼ 3.9 Hz, 1H, ArH), 6.97–6.95
115.86, 113.83, 112.42, 112.24, 55.27, 55.11 ppm; IR (KBr, ꢀ): (m, 4H, ArH), 3.79 ppm (s, 6H, 2 ꢁ O–CH3) IR (KBr, ꢀ): 3443,
3444, 3223, 2913, 1607, 1589, 1514, 1256, 836 cmꢂ1; MS (EI) 2839, 1608, 1514, 1353, 1254, 1071, 834 cmꢂ1 MS (EI) m/z (%):
m/z (%): 429 (M, 36), 334 (29), 308 (10), 238 (100), 223 (53), 195 446 (M+, 17), 400 (100), 267 (24), 238 (79), 223 (86), 195 (23),
(12), 152 (11). Anal. Calcd. for C24H20FN5O2: C, 67.12; H, 4.69; 180 (22), 152 (21), 134 (17), 119 (16), 79 (12). Anal. Calcd. for
N, 16.31; found: C, 66.85; H, 4.81; N, 15.98.
C22H18N6O5: C, 59.19; H, 4.06; N, 18.83; found: C, 59.64; H,
3.94; N, 18.11.
5,6-Bis(4-methoxyphenyl)-3-(2-(thiophen-2-ylmethylene)hydrazi-
nyl)-1,2,4-triazine (G4)7
5,6-Bis(4-methoxyphenyl)-3-(2-[(1H-indol-3-yl)methylene]hydra-
zinyl)-1,2,4-triazine (G8)7
The yellow solid product G4 was prepared from thiophene-2-
carbaldehyde (112 mg, 1 mmol) in similar manner described in The titled compound G8 was synthesized according to the general
general procedure (yield: 91%), m.p.: 250–253 ꢀC; 1H NMR described procedure using 1H-indole-3-carbaldehyde (145 mg,
(DMSO-d6, 500 MHz), dH: 11.79 (s, 1H, NH), 8.45 (s, 1H, 1 mmol) as yellow solid (yield: 74%), m.p.: 258–262 ꢀC; 1H NMR
N¼CH), 7.61 (s, 1H, ArH), 7.44–7.46 (m, 2H, ArH), 7.39–7.35 (DMSO-d6, 500 MHz), ꢁH: 11.48 (s, 1H, NH), 8.51 (d, J ¼ 7.2 Hz,
(m, 3H, ArH), 7.12 (brs, 1H, ArH), 6.96–6.95 (m, 4H, ArH), 1H, ArH), 8.45 (s, 1H, N¼CH), 7.77 (brs, 1H, NH), 7.56 (d,
3.78 ppm (s, 6H, 2 ꢁ O–CH3); 13C NMR (DMSO-d6, 125 MHz), J ¼ 7.7 Hz, 2H, ArH), 7.44 (d, J ¼ 7.7 Hz, 2H, ArH), 7.39 (d,
dC ¼ 160.94, 159.34, 158.03, 155.37, 151.23, 150.33, 139.65, J ¼ 8.4 Hz, 2H, ArH), 7.21–7.18 (m, 2H, ArH), 6.99–6.96 (m, 4H,
138.66, 131.13, 130.17, 129.28, 128.67, 127.90, 127.76, 55.27, ArH), 3.80 ppm (s, 6H, 2 ꢁ O–CH3); 13C NMR (DMSO-d6,
55.10 ppm. IR (KBr, ꢀ): 3350, 3220, 1608, 1514, 1360, 831 cmꢂ1
;
125 MHz), ꢁC ¼ 160.95, 159.24, 158.44, 155.01, 149.39, 140.71,
MS (EI) m/z (%): 417(M+, 61), 308 (10), 283 (10), 252 (11), 238 136.98, 131.82, 131.12, 130.13, 129.01, 128.92, 128.26, 124.44,
(100), 223 (57), 195(20), 180 (13), 152 (23), 39 (16). Anal. Calcd. 122.50, 122.32, 120.23, 113.83, 113.74, 112.35, 111.68, 55.32,
for C22H19N5O4S: C, 66.23; H, 4.59; N, 16.78; found: C, 65.94; H, 55.12 ppm; IR (KBr, ꢀ): 3414, 3010, 1609, 1510, 1249, 1065,
4.74; N, 16.94.
839 cmꢂ1; MS (EI) m/z (%): 450 (M+, 4), 308 (14), 238 (100),
223(47), 195 (15), 142 (18), 116 (16). Anal. Calcd. for
C26H22N6O2: C, 69.32; H, 4.92; N, 18.66; found: C, 68.97; H,
4.76; N, 18.34.
3-(2-(4-Hydroxy-3-methoxybenzylidene)hydrazinyl)-5,6-bis(4-
mthoxyphenyl)-1,2,4-triazine (G5)7
Compound G5 was obtained according to the described general
procedure using 4-hydroxy-3-methoxybenzaldehyde (152 mg,
1 mmol) as yellow solid (yield 88%): m.p.: 245–248 ꢀC; 1H
3-(2-(2-Chloro-6-nitrobenzylidene)hydrazinyl)-5,6-bis(4-methox-
yphenyl)-1,2,4-triazine (G9)7
NMR (DMSO-d6, 500 MHz), dH: 11.62 (s, 1H, NH), 9.46 (s, 1H, The yellow solid product G9 was prepared from 2-chloro-6-
N¼CH), 8.15 (s, 1H, ArOH), 7.49 (d, J ¼ 8.5 Hz, 2H, ArH), 7.36 nitrobenzaldehyde (185 mg, 1 mmol) according to the general
1
(d, J ¼ 8.5 Hz, 2H, ArH), 7.31 (s, 1H, ArH), 7.11 (d, J ¼ 8.1 Hz, procedure (yield: 71%), m.p.: 258–262 ꢀC; H NMR (DMSO-d6,
1H, ArH), 6.96–6.94 (m, 4H, ArH), 6.84 (d, J ¼ 8.1 Hz, 1H, ArH), 500 MHz), ꢁH: 12.33 (s, 1H, NH), 8.68 (s, 1H, N¼CH), 8.14–
3.84 (s, 3H, OCH3), 3.78 ppm (s, 6H, 2 ꢁ O–CH3); 13C NMR 8.13(m, 2H, ArH), 7.70 (dd, J ¼ 2.45 Hz, J ¼ 8.8 Hz, 1H, ArH),
(DMSO-d6, 125 MHz), dC ¼ 161.48, 159.29, 158.22, 155.28, 7.51 (d, J ¼ 8.8 Hz, 2H, ArH), 7.39 (d, J ¼ 8.7 Hz, 2H, ArH),
149.99, 148.27, 147.91, 144.08, 131.12, 130.93, 130.72, 130.14, 6.98–6.96 (m, 4H, ArH), 3.79 ppm (s, 6H, 2 ꢁ O–CH3); IR (KBr,
129.16, 128.78, 128.01, 126.29, 120.87, 115.51, 113.79, 113.70, ꢀ): 3424, 3121, 1602, 1519, 1125, 831 cmꢂ1; MS (EI) m/z (%):
113.57, 109.18, 55.50, 55.25, 55.09 ppm; IR (KBr, ꢀ): 3485, 3225, 490 (M+, 21), 492 (M+2, 6), 334 (16), 308 (15), 238 (100), 223
1607, 1508, 1257, 843 cmꢂ1; MS (EI) m/z (%): 417 (M+, 61), 323 (45), 195 (13), 152(10). Anal. Calcd. for C24H19ClN6O4: C,
(10), 308 (37), 252 (14), 238 (100), 223 (53), 195 (14), 152 (12). 58.72; H, 3.90; N, 17.12; found: C, 58.97; H, 3.72; N, 17.16.
Anal. Calcd. for C25H23N5O4: C, 65.63; H, 5.07; N, 15.31; found:
C, 65.93; H, 5.36; N, 15.51.
3-(2-(1,3-Dihydroxybenzylidene)hydrazinyl)-5,6-bis(4-methoxy-
phenyl)-1,2,4-triazine (G10)7
3-(2-(2,4-Dichlorobenzylidene)hydrazinyl)-5,6-bis(4-methoxyphe-
nyl)-1,2,4-triazine (G6)7
The titled compound G10 was synthesized according to the
general procedure from 2,4-dihydroxybenzaldehyde (138 mg,
The titled compound G6 was synthesized according to the general 1 mmol) as yellow solid (yield: 86%), m.p.: 262–266 ꢀC; 1H
procedure using 2,4-dichlorobenzaldehyde (175 mg, 1 mmol) as NMR (DMSO-d6, 500 MHz), ꢁH: 11.93 (s, 1H, NH), 11.71 (s, 1H,
pale yellow solid (yield 78%), m.p.: 248–251ꢀC; 1H NMR ArOH), 9.89 (s, 1H, ArOH) 8.31 (s, 1H, N¼CH), 7.77 (s, 1H,
(DMSO-d6, 500 MHz), dH: 12.14 (s, 1H, NH), 8.61 (s, 1H, ArH), 7.50 (d, J ¼ 8.7 Hz, 2H, ArH), 7.38 (d, J ¼ 8.7 Hz, 2H,
N¼CH), 8.06 (d, J ¼ 8.6 Hz, 1H, ArH), 7.72 (d, J ¼ 2.0 Hz, 1H, ArH), 7.25 (d, J ¼ 8.4 Hz, 1H, ArH), 6.97–6.95 (m, 4H, ArH),
ArH), 7.53 (dd, J ¼ 2.0 Hz, J ¼ 8.5 Hz, 1H, ArH), 7.50 (d, 6.37–6.35 (dd, J ¼ 2.2 Hz, J ¼ 8.3 Hz, 1H, ArH), 3.79 ppm (s, 6H,
J ¼ 8.7 Hz, 2H, ArH), 7.38 (d, J ¼ 8.7 Hz, 2H, ArH), 6.97–6.95 2 ꢁ O–CH3); IR (KBr, ꢀ): 3352, 2837, 1608, 1517, 1253,
(m, 4H, ArH), 3.79 ppm (s, 6H, 2 ꢁ O–CH3); IR (KBr, ꢀ): 3444, 834 cmꢂ1; MS (m/z, %): 443 (M+, 22), 308 (11), 238 (100), 195
3183, 1607, 1511, 1369.82 (M+2, 5), 334 (16), 308 (15), 238 (100), (11). Anal. Calcd. for C24H21N5O4: C, 65.01; H, 4.77; N, 15.79;
223 (45), 195 (13), 152 (10). Anal. Calcd. for C24H19Cl2N5O2: C, found: C, 64.91; H, 4.83; N, 16.01.
60.01; H, 3.99; N, 14.58; found: C, 59.84; H, 4.16; N, 14.41.
3-(2-(Benzo[d][1,3]dioxol-5-ylmethylene)hydrazinyl)-5,6-bis(4-
methoxyphenyl)-1,2,4-triazine (G11)7
5,6-Bis(4-methoxyphenyl)-3-(2-((5-nitrofuran-2-yl)methylene)
hydrazinyl)-1,2,4-triazine (G7)
The titled compound G11 was synthesized by the described
The titled compound G7 was synthesized according to the general general procedure using 1,3-benzodioxole-5-carbaldehyde (piper-
1
procedure using 5-nitrofuran-2-carbaldehyde (141 mg, 1 mmol) as onal) (150 mg, 1 mmol) (yield 88%), m.p.: 268–272 ꢀC; H NMR

4
M. Khoshneviszadeh et al.
J Enzyme Inhib Med Chem, Early Online: 1–10
(DMSO-d6, 400 MHz), ꢁH: 11.62 (s, 1H, NH), 8.16 (s, 1H, (m/z, %): 489.1 (M+, 11.5), 491.1 (M+2, 11.4). Anal. Calcd. for
N¼CH), 7.49 (d, J ¼ 7.6 Hz, 2H, ArH), 7.40–7.34 (m, 2H, ArH), C24H20BrN5O2: C, 58.79; H, 4.11; N, 14.28; found: C, 59.35; H,
7.31 (s, 1H, ArH), 14 (d, J ¼ 7.6 Hz, 2H, ArH), 6.98–6.94 (m, 4H, 4.91; N, 13.24.
ArH), 6.08 (s, 2H, ArH), 3.78 ppm (s, 6H, 2 ꢁ O–CH3); IR (KBr,
455 (M+, 18), 334 (10), 308 (16), 238 (100), 223 (38), 195 (11).
Anal. Calcd. for C25H21N5O4: C, 65.93; H, 4.65; N, 15.38; found:
C, 65.71; H, 5.01; N, 15.65.
ꢀ): 3429, 2962, 1606, 1515, 1175, 836 cmꢂ1; MS (EI) m/z (%):
Biological activities
In vitro COX inhibition assays
In vitro bioactivities of all the synthesized compounds were
evaluated using celecoxib and indomethacin as the reference
drugs. The HWB assay was employed to determine the activity of
COX isoenzymes36. Five-hundred microliters of fresh blood
samples obtained from human volunteers were incubated with 1 ml
of DMSO as a vehicle or 1 ml of 10 mM solution of synthesized
compounds or indomethacin and celecoxib as the reference drugs.
The plasma was separated after 1 h incubation at 37 ꢀC. COX-1
level was measured by the production of thromboxane B2 (TxB2)
using a commercially available enzyme immunoassay (EIA) kit.
In order to determine COX-2 activity, 500 ml of fresh blood
samples obtained from human volunteers were collected in
EDTA-containing tubes, incubated with 1 ml of DMSO as a
vehicle or 1 ml of 10 mM solution of synthesized compounds or
indomethacin and celecoxib as the reference drugs. The samples
were incubated at 37 ꢀC in the presence of 10 mg/ml LPS for 24 h,
after which the plasma was separated. COX-2 level was measured
by the production of PGE2 using a commercially available EIA
kit.
3-(2-(Furan-2-ylmethylene)hydrazinyl)-5,6-bis(4-methoxyphenyl)-
1,2,4-triazine (G12)7
The yellow solid product G12 was prepared from furan-2-
carbaldehyde (96 mg, 1 mmol) according to the general procedure
1
(yield: 71%), m.p.: 258–262 ꢀC; H NMR (DMSO-d6, 500 MHz),
ꢁH: 11.79 (s, 1H, NH), 8.15 (s, 1H, N¼CH), 7.82 (s, 1H, ArH),
7.47 (d, J ¼ 8.6 Hz, 2H, ArH), 7.36 (d, J ¼ 8.6 Hz, 2H, ArH),
6.96–6.94 (m, 4H, ArH), 6.85 (d, J ¼ 3.2, 1H, ArH), 6.63 (brs, 1H,
ArH), 3.78 ppm (s, 6H, 2 ꢁ O–CH3); 13C NMR (DMSO-d6,
125 MHz), ꢁC ¼ 161.57, 161.42, 159.99, 158.72, 156.17, 151.82,
150.92, 150.33, 145.73, 144.97, 134.10, 131.66, 130.71, 129.16,
128.44, 116.15, 114.30, 114.22, 112.57, 55.77, 55.61 ppm; IR
(KBr, ꢀ): 3443, 2839, 1608, 1254, 834 cmꢂ1; MS (EI) m/z (%):
401 (M+, 66), 372 (23), 308 (15), 267 (47), 252 (14), 238 (100),
223 (89), 195 (22), 152 (21), 135 (37). Anal. Calcd. for
C22H19N5O3: C, 65.83; H, 4.77; N, 17.45; found: C, 65.51; H,
5.03; N, 17.12.
Inhibition of 15-LOX
3-(2-(4-Methoxybenzylidene)hydrazinyl)-5,6-bis(4-methoxyphe-
nyl)-1,2,4-triazine (G13)7
The assay was performed according to the previously reported
method37. The compounds were dissolved in DMSO and
phosphate buffer. The solution was added to the assay mixture
that contained 9 ml of 0.1 M sodium phosphate buffer, pH 8 and
enzyme (Enal concentration: 167 U/ml). The mixture was
incubated for four minutes and then linoleic acid as substrate
was added to reach the Enal concentration of 134 mM. Quercetin,
a well-known inhibitor of 15-LOX was used as a positive
control. Absorbance was recorded by spectrophotometer at
234 nm during the measurement period. Enzyme activity
experiments were tested in triplicate and the values are
expressed as means SD.
The titled compound G13 was synthesized according to the
general procedure from 4-methoxybenzaldehyde (136 mg,
1 mmol) as yellow solid (yield: 91%), m.p.: 254–256 ꢀC; 1H
NMR (DMSO-d6, 500 MHz), ꢁH: 11.72 (s, 1H, NH), 8.21 (s, 1H,
N¼CH), 7.67 (d, J ¼ 8.0 Hz, 2H, ArH), 7.49 (d, J ¼ 8.0 Hz, 2H,
ArH), 7.36 (d, J ¼ 8.0 Hz, 2H, ArH), 7.02–6.94 (m, 6H, ArH),
3.80 (s, 3H, OCH3), 3.78 ppm (s, 6H, 2 ꢁ O–CH3); IR (KBr, ꢀ):
3444, 3215, 1608, 1577, 1248, 834 cmꢂ1; MS (EI) m/z (%): 441
(M+, 14), 308 (31), 238 (100), 223 (49), 195 (12), 152 (10). Anal.
Calcd. for C25H23N5O3: C, 68.01; H, 5.25; N, 15.86; found: C,
68.28; H, 4.93; N, 15.67.
Determination of the anti-oxidant activity by FRAP assay
5,6-Bis(4-methoxyphenyl)-3-(2-(pyridin-4-ylmethylene)-hydrazi-
nyl)-1,2,4-triazine (G14)7
FRAP assay33 was employed to evaluate the anti-oxidant activities
of the target compounds. For this purpose, FRAP solution was
prepared by adding 10 ml acetate buffer 300 mM at pH 3.6 to 1 ml
of aqueous solution of FeCl3ꢃ6H2O and 1 ml (10 mM) acidic
solution of 2,4,6-tris(2-pyridyl)-s-triazine (TPTZ). Twenty five
microliters of 100 mM of compounds was added to FRAP solution.
After incubation at 37 ꢀC for 15 min, the absorbance at 585 nm
was monitored relative to ferrous sulfate (FeSO4) as the standard.
Results are reported as mmol Fe(II) per gram of dry mass of
compounds. Ascorbic acid was selected as a reference. The data
are mean of three independent experiments.
The yellow solid product G14 was prepared from pyridine-4-
carbaldehyde (107 mg, 1 mmol) according to the general proced-
ure (yield: 81%), m.p.: 255–258 ꢀC; 1H NMR (DMSO-d6,
500 MHz), ꢁH: 12.17 (s, 1H, NH), 8.63 (d, J ¼ 4.4 Hz, 2H,
ArH), 8.23 (s, 1H, N¼CH), 7.67 (d, J ¼ 5.7 Hz, 2H, ArH), 7.52 (d,
J ¼ 8.8 Hz, 2H, ArH), 7.39 (d, J ¼ 8.6 Hz, 2H, ArH), 6.98–6.96
(m, 4H, ArH), 3.79 ppm (s, 6H, 2 ꢁ O–CH3); IR (KBr, ꢀ): 3434,
3146, 1609, 1569, 1507, 1237, 841 cmꢂ1; MS (EI) m/z (%): 412
(M+, 32), 334 (16), 308 (12), 238 (100), 223 (25), 195 (10). Anal.
Calcd. for C23H20N6O2: C, 66.98; H, 4.89; N, 20.38; found: C,
66.75; H, 5.71;N, 19.93.
Molecular docking study
3-(2-(4-Bromobenzylidene)hydrazinyl)-5,6-bis(4-methoxyphenyl)-
1,2,4-triazine (G15)
Molecular docking study was performed using Autodock 4.2 and
Auto Dock Tools 1.5.4 (ADT). For this purpose, the X-ray crystal
structure of COX-1 carrying diclofenac as cognate ligand in
binding site (PDB code: 3N8Y) and X-ray crystal structure of
COX-2 in complex with SC-558 (PDB code: 1CX2) were
downloaded from protein data bank (http://www.rcsb.org).
Water molecules and co-crystalized ligand were excluded. The
PDB file of protein was regenerated by adding polar hydrogens
and computing Gasteiger charges.
The yellow solid product G15 was prepared from 4-bromoben-
zaldehyde (185 mg, 1 mmol) according to the general procedure
1
(yield: 82%), m.p.: 254–256 ꢀC. H NMR (DMSO-d6, 500 MHz),
ꢁH: 11.92 (s, 1H, NH), 8.24 (s, 1H, N¼CH), 7.69–7.64 (m, 4H,
ArH), 7.50 (d, J ¼ 8.2, 2H, ArH), 7.37 (d, J ¼ 8.2 Hz, 2H, ArH),
6.97–6.95 (m, 4H, ArH), 3.79 ppm (s, 6H, 2 ꢁ O–CH3). IR (KBr,
ꢀ): 3422, 3223, 2907, 1607, 1592, 1510, 1254, 837 cmꢂ1; MS

DOI: 10.3109/14756366.2016.1158713
1,2,4-Triazine derivatives
5
OH
OH
O
S
H
HO
N
H2N
N
N
OH
O
4
5
COOH
6
MeO
N
S
NH
N
CH3
N
N
N
S
N
Ar
7
N
H
MeO
H3C
N
N
O
Cl
O
S
H3C
S
NH
O
H3C
1
N
O
N
CH3
S
3
N
MeO
2
Figure 2. Design of COX-2 inhibitors using molecular hybridization method.
O
OH
O
O
MeO
MeO
CHO
(a)
(b)
N
N
(c)
N
MeO
SH
MeO
OMe
MeO
OMe
A
B
C
D
MeO
MeO
MeO
MeO
MeO
MeO
N
N
N
N
N
N
N
(d)
N
(e)
(f)
N
NH2
S
N
Ar
N
H
N
H
E
F
G1-G15
Scheme 1. Synthesis pathway for the preparation of triazine derivatives. Reagents and conditions: (a) thiamine hydrochloride, NaOH, EtOH, reFux,
3 h; (b) HNO3, 60 ꢀC; (c) thiosemicarbazide, CH3COOH, reFux, 3 h; (d) CH3I, NaOH, EtOH, 2 h, rt; (e) NH2NH2ꢃH2O, EtOH, reFux, 12 h; (f) ArCHO,
EtOH, 1–10 h, rt.
The 3D structures of ligands were created using Avogadro Results and discussion
software. Each ligand structure was separately submitted to a
Fragment-based design
molecular mechanics conformational search (MM+ force Eeld)
and the geometry of the most stable conformer was then The target 1,2,4-triazine derivatives were designed by the molecu-
optimized by semi-empirical method (AM1) in Hyperchem lar hybridization approach based on selective COX-2 inhibitor
software. The Gasteiger charges were added and the degrees of etoricoxib (1) and a number of potent anti-inflammatory agents that
torsions were defined to generate PDBQT format of the ligands. are reported in the literature (compounds 2–7) (Figure 2).
The grid parameter file was generated using AutoGrid. Etoricoxib (1), commonly used for pain relief after surgery, is
Numbers of points in x, y,
z
dimensions were set to approved in more than 80 countries worldwide which is a selective
60 ꢁ 60 ꢁ 60 with 0.375 A grid spacing. Rigid macromolecule COX-2 inhibitor, containing pyridine with bearing adjacent diaryls.
was chosen to define the docking parameter file using Lamarckian It has been reported that 1,2,4-triazine heterocyclic compounds
Genetic Algorithm (LGA), with 200 GA runs and 25 000 000 with diaryl substituent (2) are effective COX-2 inhibitors17,18
˚
.
energy evaluations while the other parameters were left as default. Some experiments have reported structures in which the sulfonyl
The results revealed that all tested compounds fit much well in the moiety on the aromatic ring is missing (3)25–27. Niedballa et al.
COX-2 cavity rather than COX-1 cavity.
showed that sulfonyl moiety can be replaced with alkoxy groups28.

6
M. Khoshneviszadeh et al.
J Enzyme Inhib Med Chem, Early Online: 1–10
Table 1. COX-1 and COX-2 inhibitory activities of synthesized 1,2,4-triazine compounds.
O
N
N
N
Ar
N
N
H
O
% Inhibition
COX-2/COX-1
Compound
G1
Ar
COX-2 (10 mM)
COX-1 (10 mM)
71.85 8.039
67.3 6.057
1.07
0.94
G2
72.32 9.023
76.86 13.098
NO2
G3
G4
72.09 11.027
87.45 11.008
69.1 6.088
72.03 7.109
1.04
1.21
F
S
G5
63.58 8.004
59 6.098
1.08
OMe
OH
Cl
G6
G7
G8
61.2 7.006
65.85 4.010
60.5 9.006
57.2 3.073
59.8 5.099
38.76 6.054
1.07
1.10
1.56
Cl
O
NO2
H
N
Cl
G9
65.7 7.009
50.76 10.108
1.29
O2N
G10
G11
68.9 5.007
78.00 .8.012
63.73 2.119
50.33 3.085
1.08
1.55
HO
OH
O
O
G12
G13
78.71 9.023
71.85 10.012
66.43 4.204
53.66 6.102
1.18
1.34
O
O
G14
G15
73.74 9.011
73.27 12.006
64.26 7.123
59.36 6.097
1.15
1.23
N
Br
Indomethacin at 500 nM
Celecoxib at 500 nM
–
–
84.32 0.018
70.91 0.031
73.96 0.032
32.4 0.024
1.14
2.19
The results are summarized for inhibition% of COX isoenzymes. The data are expressed as the mean SEM from three different experiments (n ¼ 3).

DOI: 10.3109/14756366.2016.1158713
1,2,4-Triazine derivatives
7
Table 2. The 15-lipoxygenase inhibitory and anti-oxidant
activities of the synthesized 1,2,4-triazine compounds.
The natural product (4) resveratrol, has been introduced
a
COX-2 inhibitor29. Lin et al. demonstrated that
as
bioisosteric replacement of the central –N¼N– linkage of (5)
with –N¼C–double bond leads to 114-fold more COX-2 inhibi-
tory effect than COX-130.
2-Benzylidene-1-phenylhydrazine derivative (6) has been
proposed as a COX-2 inhibitor and hydrazine derivative (7) is
described as a dual COX/5-LOX inhibitor31. With respect to these
structural features, we designed a set of COX-2 inhibitors. The
target compounds were synthesized via the same route that we
had previously reported7.
Compound
SLOa IC50 (mM)b
FRAP value
G1
G2
G3
G4
G5
G6
G7
G8
143
225
124
420
NAc
NA
NA
NA
370
205
NA
NA
NA
190
NA
16
0.309
0.29
0.309
60.7
269.9
0.264
0.1639
147.38
0.198
487
0.2781
0.3704
0.327
42.8
G9
G10
G11
G12
G13
G14
G15
Quercetin
Vitamin C
Chemistry
The synthetic reactions used for the synthesis of the designed
fifteen 1,2,4-triazine derivatives (G1–G15) are outlined in
Scheme 1. 2-Hydroxy-1,2-bis(4-methoxyphenyl) ethanone (B)
was prepared by thiamine hydrochloride-mediated coupling of 4-
methoxy benzaldehyde (A). Oxidation of compound B afforded
the corresponding 1,2-bis(4-methoxyphenyl)ethane-1,2-dione (C).
5,6-Diaryl-1,2,4-triazine-3-thiol (D) was acquired through cyc-
lization of compound (C) with thiosemicarbazide. Compound (D)
was treated with methyl iodide to afford corresponding methylthio
compound (E). In the presence of excess amount of hydrazine
hydrate, compound (F) was prepared. The final compounds (G1–
G15) were prepared by treatment of (F) with equimolar amounts
of the appropriate aldehydes. All the steps were carried out
according to our previously reported procedures7.
0.293
1903.6
674.78
–
aSoybean lipoxygenase.
bData are mean of three independent experiments.
cNo activity.
The compound G3 containing flourophenyl moiety with IC50
value of 124 mM was the most potent one. As reported in Table 2,
the phenyl substituted compound G1 showed IC50 value of
143 mM and insertion of different substituents resulted in
decreased activity. Compounds G1, G14 and G10 (phenyl, pyridyl
and di-hydroxy derivatives, respectively) were active against 15-
LOX to some extent and compounds G2, G9, G4 (nitrophenyl, 2-
chloro-6-nitrophenyl and thiophenyl, respectively) showed less
activity while the other compounds were inactive.
Biological activity
In vitro COX inhibition assay
The human whole blood (HWB) assay, initially developed by
Patrignani et al.32, was performed to assess the ability of
synthesized compounds to inhibit COX-2 and COX-1 isoenzymes.
Celecoxib and indomethacin were used as the reference drugs.
The results are summarized in Table 1. Celecoxib as a selective
COX-2 inhibitor exhibited 70.9%, 32.4% inhibition for COX-2
and COX-1, respectively, while in the case of indomethacin which
is a nonselective COX inhibitor exhibited 84.3% inhibition for
COX-2 and 73.9% inhibition for COX-1. These 15 compounds
inhibited COX isoenzymes to various degrees, in a range of 60–
78% at 10 mM. Compounds G9, G4, G13, G8 and G11 represented
the highest selectivity for COX-2 in comparison with COX-1. The
compound G11, namely 1,3-benzodioxole-substituted compound,
showed the highest selectivity for COX-2 at 10 mM. A brief
structure–activity relationship (SAR) can be distinguished as
follows according to the data presented in Table 1:
ꢄ The introduction of electron-withdrawing moieties, such as
nitro or Fuoro on phenyl ring resulted in compounds with the
lowest activities against COX-2 in this set. This effect can be
seen in compounds G2 and G3.
ꢄ Para-substituted phenyl rings by both electron-donating and
electron-withdrawing groups represented reduced activities.
G13 and G15 showed a lower affinity toward COX-1 and
hence represented higher COX-2/COX-1 ratios.
Ferric reducing anti-oxidant power (FRAP) assay
The anti-oxidant potential of target compounds were determined
using FRAP assay33. This method is based on the ability of
compounds in the reduction of ferric tripyridyl triazine complex
to ferrous (Fe2+) form. The result is production of an intense blue
color. The change in the absorbance at 585 nm is a tool for
monitoring the reaction progress. Higher increase in the absorb-
ance indicates that the potential of reduction is higher. The results
are presented in Table 2. Ascorbic acid (vitamin C) was
considered as a reference compound. Compound G10 showed
the highest anti-oxidant activity.
Molecular modeling
In order to clarify the mechanism of COX inhibition and virtual
interactions between synthetic compounds and active sites of
COX-1 (PDB code: 3N8Y)34 and COX-2 (PDB code: 1CX2)35,
the molecular docking was performed. Substitution of Ile523 in
COX-1 relative to Val523 in COX-2 which is a less bulky amino
acid and hence the conformational change at Tyr355 results in the
presence of an accessible 2ꢀ-pocket in COX-2 active site. While
the similar pocket in COX-1 is inaccessible. The other reason for
effective COX-2 inhibition is the replacement of His513 in COX-
1 by Arg513 in COX-2 which enables the ligands to form more H-
bonds in the COX-2 active site. Therefore, because of the
differences between the active sites of enzymes, it enables us to
design selective COX-2 inhibitors.
ꢄ Compounds with five membered heterocyclic rings, such as
G4 and G12 displayed COX-2 selectivity. G11 and G8 with
heterocyclic fused ring aldehydes were the most selective ones
among the tested compounds.
The docking study was performed using Autodock4.2. All the
ligands showed a similar orientation in COX-2 active site (Figure
3). Compound G11 docked in COX-2 is shown (Figure 4). In
general, it is observed that the oxygen atom of methoxy phenyl
moiety is placed close to the side chain of Arg-120 (guanidine
In vitro 15-lipoxygenase (LOX) inhibitory activity
In vitro 15-LOX inhibitory activities of synthesized compounds
were tested. The results are reported as IC50 values in Table 2.

8
M. Khoshneviszadeh et al.
J Enzyme Inhib Med Chem, Early Online: 1–10
Figure 3. Superimposition of ligands in the
COX-2 binding pocket.
Figure 4. 3D orientation model of G11 (ball
and stick) in COX-2 active pocket (blue:
hydrophobic pocket, red: Tyr353, yellow:
responsible for the COX-2 selectivity, purple:
selectivity pocket).
˚
H2N. . .OCH3, 2.5 A) while the other methoxy phenyl is
The compound G11 as a representative example is shown in
accommodated in hydrophobic pocket formed by Tyr385, Figure 5. As it was mentioned before, the presence of an
Gly526, Ser530, Ala527. 1,2,4-Triazine and hydrazine moieties inaccessible pocket in COX-1 resulted in different conformation
were inserted in the selectivity pocket that was created as a result and hence different H-bonds with Ser530 (triazine N. . .C¼O,
˚
˚
˚
of Val523 presence in COX-2 by Ile523 in COX-1, then they got 2.8 A, 2.9 A), Tyr385 (triazine N. . .OH, 2.8 A) and Tyr355 (CH3-
˚
˚
involved in H-bond with Leu352 (N¼N. . .C¼O, 2.2 A), (triazine O. . .OH, 3.5 A).
˚
˚
N. . .C¼O, 2.9 A), Ser353 (triazine N. . .C¼O, 3.1 A), Tyr355
˚
(triazine N. . .O–H, 2.8 A). The other hydrogen bond was formed
˚
by benzo[1,3]dioxole fragment (O. . .N, 3.2 A).
Conclusion
A similar method was used to compare ligand interactions in
COX-1 (PDB code: 3N8Y) enzyme active site. All compounds
showed a different conformation in COX-1 active site from their
conformation in COX-2 active site.
In summary, fragment based design strategy was utilized to design
1,2,4-triazine derivatives as selective COX-2 inhibitors. The
designed compounds were synthesized and their biological

DOI: 10.3109/14756366.2016.1158713
1,2,4-Triazine derivatives
9
Figure 5. 3D model of the orientation of G11
(ball and stick) in COX-1 active site (blue:
hydrophobic pocket, red: Tyr353, yellow:
responsible for the COX-2 selectivity, purple:
selectivity pocket).
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7. Khoshneviszadeh M, Ghahremani MH, Foroumadi A, et al. Design,
synthesis and biological evaluation of novel anti-cytokine 1,2,4-
triazine derivatives. Bioorg Med Chem 2013;21:6708–17.
8. Cignarella G, Vianello P, Berti F, et al. Synthesis and pharmaco-
logical evaluation of derivatives structurally related to nimesulide.
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activities were evaluated. The results demonstrated that com-
pounds G9, G4, G13, G8 and G11 exhibited stronger effect
against COX-2 in comparison with their activity against COX-1 at
10 mM. In vitro 15-LOX inhibitory activity of synthesized
compounds was also tested. Compound G3 with an IC50 value
of 124 mM was the most potent derivative. The anti-oxidant
potential of synthesized compounds was determined by FRAP
assay and compound G10 showed the highest anti-oxidant
activity. The binding patterns of compounds inside the COX-2/
COX-1 active sites were predicted using a docking technique.
Considering the docking results, it can be suggested that 1,2,4-
triazine and hydrazine moieties are positioned in the selectivity
pocket in COX-2 active site, which provides a useful scaffold for
novel compounds with improved inhibitory activities.
Declaration of interest
The authors report no conflicts of interest. The authors alone are
responsible for the content and writing of this article. The authors
acknowledge the financial support of Shiraz University of
Medical Sciences, Vice Chancellor of Research (Grant: 7442).
12. Navidpour L, Shadnia H, Shafaroodi H, et al. Design, synthesis,
and biological evaluation of substituted 2-alkylthio-1,5-diarylimi-
dazoles as selective COX-2 inhibitors. Bioorg Med Chem 2007;15:
1976–82.
13. Qiu K-M, Yan R, Xing M, et al. Synthesis, biological evaluation and
molecular modeling of dihydro-pyrazolyl-thiazolinone derivatives as
potential COX-2 inhibitors. Bioorg Med Chem 2012;20:6648–54.
14. Ghodsi R, Zarghi A, Daraei B, et al. Design, synthesis and biological
evaluation of new 2,3-diarylquinoline derivatives as selective
cyclooxygenase-2 inhibitors. Bioorg Med Chem 2010;18:1029–33.
15. Reddy M, Billa VK, Pallela VR, et al. Design, synthesis, and
biological evaluation of 1-(4-sulfamylphenyl)-3-trifluoromethyl-5-
indolyl pyrazolines as cyclooxygenase-2 (COX-2) and lipoxygenase
(LOX) inhibitors. Bioorg Med Chem 2008;16:3907–16.
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