Design, synthesis and biological evaluation of novel diphenylthiazole-based cyclooxygenase inhibitors as potential anticancer agents

Article abstract of DOI:10.1016/j.bioorg.2014.10.001

Non-steroidal anti-inflammatory drugs (NSAIDs) are among the most widely used medications as analgesics and antipyretics. Currently, there is a growing interest in their antitumor activity and their ability to reduce the risk and mortality of several cancers. While several studies revealed the ability of NSAIDs to induce apoptosis and inhibit angiogenesis in cancer cells, their exact anticancer mechanism is not fully understood. However, both cyclooxygenase (COX)-dependent and -independent pathways were reported to have a role. In an attempt to develop new anticancer agents, a series of diphenylthiazole substituted thiazolidinone derivatives was synthesized and evaluated for their anticancer activity against a panel of cancer cell lines. Additionally, the inhibitory activity of the synthesized derivatives against COX enzymes was investigated as a potential mechanism for the anticancer activity. Cytotoxicity assay results showed that compounds 15b and 16b were the most potent anticancer agents with half maximal inhibitory concentrations (IC₅₀) between 8.88 and 19.25?μM against five different human cancer cell lines. Interestingly, COX inhibition assay results were in agreement with that of the cytotoxicity assays where the most potent anticancer compounds showed good COX-2 inhibition comparable to that of celecoxib. Further support to our results were gained by the docking studies which suggested the ability of compound 15b to bind into COX-2 enzyme with low energy scores. Collectively, these results demonstrated the promising activity of the newly designed compounds as leads for subsequent development into potential anticancer agents.

Full text of DOI:10.1016/j.bioorg.2014.10.001

Bioorganic Chemistry 57 (2014) 132–141
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Design, synthesis and biological evaluation of novel
diphenylthiazole-based cyclooxygenase inhibitors
as potential anticancer agents
Ahmed H. Abdelazeem a,d, , Ahmed M. Gouda c,d, Hany A. Omar b,e, Mai F. Tolba f
⇑
a
Department of Pharmaceutical Chemistry, College of Pharmacy, Taif University, Taif 21974, Saudi Arabia
Department of Pharmacology, College of Pharmacy, Taif University, Taif 21974, Saudi Arabia
Department of Pharmaceutical Chemistry, College of Pharmacy, Umm Al-Qura University, Makkah 21955, Saudi Arabia
Department of Medicinal Chemistry, Faculty of Pharmacy, Beni-Suef University, Beni-Suef 62514, Egypt
Department of Pharmacology, Faculty of Pharmacy, Beni-Suef University, Beni-Suef 62514, Egypt
Department of Pharmacology and Toxicology, Faculty of Pharmacy, Ain Shams University, Cairo, Egypt
b
c
d
e
f
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 19 August 2014
Available online 22 October 2014
Non-steroidal anti-inflammatory drugs (NSAIDs) are among the most widely used medications as anal-
gesics and antipyretics. Currently, there is a growing interest in their antitumor activity and their ability
to reduce the risk and mortality of several cancers. While several studies revealed the ability of NSAIDs to
induce apoptosis and inhibit angiogenesis in cancer cells, their exact anticancer mechanism is not fully
understood. However, both cyclooxygenase (COX)-dependent and -independent pathways were reported
to have a role. In an attempt to develop new anticancer agents, a series of diphenylthiazole substituted
thiazolidinone derivatives was synthesized and evaluated for their anticancer activity against a panel of
cancer cell lines. Additionally, the inhibitory activity of the synthesized derivatives against COX enzymes
was investigated as a potential mechanism for the anticancer activity. Cytotoxicity assay results showed
that compounds 15b and 16b were the most potent anticancer agents with half maximal inhibitory con-
Keywords:
NSAIDs
Diphenylthiazole
Thiazolidinone
COX
Anticancer
Docking
centrations (IC50) between 8.88 and 19.25 lM against five different human cancer cell lines. Interestingly,
COX inhibition assay results were in agreement with that of the cytotoxicity assays where the most
potent anticancer compounds showed good COX-2 inhibition comparable to that of celecoxib. Further
support to our results were gained by the docking studies which suggested the ability of compound
15b to bind into COX-2 enzyme with low energy scores. Collectively, these results demonstrated the
promising activity of the newly designed compounds as leads for subsequent development into potential
anticancer agents.
Ó 2014 Elsevier Inc. All rights reserved.
1
. Introduction
Cancer remains one of the major leading causes of mortality all
biosynthesis, were introduced as novel targets for cancer treat-
ment [1,2]. Currently, there is an increasing body of evidence stat-
ing that targeting COX enzymes, especially COX-2 isoform, is an
effective approach for the prevention or treatment of various types
of cancers. Several studies have showed that COX-2 is overexpres-
sed in solid malignancies such as colon, prostate, and breast [3]. A
significant relationship between over-expression of COX-2 and
survival of patients with various cancers has been reported in ret-
rospective studies [4]. Moreover, it was reported that the adminis-
tration of NSAIDs such as sulindac and salicylates can induce
apoptosis in cultured HT-29 human colon cancer cells [4]. It was
also proposed that these effects may occur by both COX-dependent
and COX-independent mechanisms [5]. Another mechanism by
which NSAIDs may exert their anticancer effects involves the sup-
pression of angiogenesis. In addition, COX-2 was found to induce
over the world. Although the extensive studies and the wide vari-
ety of the existing anticancer agents, the cancer treatment remains
an aggravating and challenging problem. Therefore, development
of novel anticancer drugs with better therapeutic profile and less
side effects to combat this tenacious disease is still in demand.
During the past few years, cyclooxygenases (COX-1 and COX-2),
key enzymes catalyze the rate-limiting step in prostaglandins
⇑
Corresponding author at: Department of Medicinal Chemistry, Faculty of
Pharmacy, Beni-Suef University, Beni-Suef 62514, Egypt. Fax: +20 82 2317958.
E-mail addresses: ahmed.abdelazeem@pharm.bsu.edu.eg, ahmed.pharm@yahoo.
com (A.H. Abdelazeem).
http://dx.doi.org/10.1016/j.bioorg.2014.10.001
0
045-2068/Ó 2014 Elsevier Inc. All rights reserved.

A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
133
pro-angiogenic factors such as vascular endothelial growth factor
VEGF) and produce prostaglandins that have both autocrine and
chloride in pyridine to afford desyl chloride 8 which was readily
condensed with thiourea in absolute ethanol using a previously
reported method to give the unsubstituted starting material 10a,
4,5-diphenylthiazol-2-amine, in high yield [26]. In order to prepare
the sulfonamide derivative 10b, the desyl chloride 8 was firstly sub-
jected to a chlorosulfonation reaction using excess amount from
chlorosulfonic acid in methylene chloride at room temperature
for 24 h to form the sulfonyl chloride derivative which was subse-
(
paracrine effects on the proliferation and the migration of endothe-
lial cells in vitro [6]. Conversely, there are some other compelling
evidence suggesting COX-2 independent mechanisms that might
be involved [7,8]. Two NSAIDs, sulindac (1) and celecoxib (2),
Fig. 1, have been found to inhibit the growth of adenomatous colo-
rectal polyps and cause regression of existing polyps in patients
with familial adenomatous polyposis (FAP) [2,9,10]. Thus, cele-
coxib has been recently approved as a chemopreventive in colon
cancer and presently investigated as a chemotherapeutic for other
advanced cancers. Notably, indomethacin showed an inhibitory
effect on human colorectal cancer through apoptosis induction
and G1 arrest [11]. It was also found that it suppresses angiogene-
sis via the inhibition of mitogen activated protein kinase activity
4
quently aminated with NH OH in THF to afford the respective sul-
fonamide substituted desyl chloride 9. The cyclization of
derivative 9 was carried out as mentioned earlier with thiourea to
afford the sulfonamide derivative 10b. The intermediates 11a and
11b were obtained via reaction of the starting materials 10a and
10b with chloroacetyl chloride in methylene chloride and triethyl-
amine at 0 °C. The heterocyclization of the intermediates 11a and
11b using ammonium isothiocyanate in refluxing ethanol
efficiently furnished 2-((4,5-Diphenylthiazol-2-yl)imino)thiazoli-
din-4-one 12a and 4-(2-((4-Oxothiazolidin-2-ylidene)amino)-
(
MAPK) [12]. From a chemical point of view, diphenylheterocycle
has been recognized earlier as a promising backbone for COX
inhibitors design. It is found in several drugs such as celecoxib
and valedoxib and their relevant structural analogues which are
used as potent anti-inflammatory and analgesic agents [13].
Another example for diphenylheterocycle based-COX inhibitors is
a series of diarylthiazoles (3) which was prepared and showed high
affinity for COX enzymes, preferentially for COX-2 subtype, Fig. 1
4-phenylthiazol-5-yl)benzenesulfonamide
12b
respectively,
Scheme 1. The mechanism that has been proposed for the formation
of thiazolidine-4-one derivatives 12a and 12b was shown in Fig. 4
as previously reported [27]. Consequently, the final compounds
13–16 were obtained in excellent yields using Knoevenagel conden-
sation of the intermediates 12a and 12b with the appropriate alde-
hyde in toluene in the presence of ammonium acetate. It is
noteworthy that the methine proton mostly appeared downfield
as a singlet with chemical shift equal to more than 7.00 ppm due
to the reported deshielding effect attributed to the adjacent car-
bonyl group owing to the presence of the Knoevenagel reaction
products as only Z isomers [28]. All the newly synthesized com-
pounds (Table 1) were characterized and confirmed by elemental
[
14].
On the other hand, thiazolidinone and thiazolidinedione (TZD) are
important and frequently used scaffolds in drug design and discovery
that possess a huge variety of biological activities including an anti-
cancer effect [15–17]. Numerous compounds bearing the TZD moiety
such as the PI3K inhibitor, GSK1059615 (4) [18] and thiazolidinone-
based derivatives (5 and 6) have been recognized as potential antican-
cer agents, Fig. 2 [19]. A current paradigm proved that 4-thiazolidi-
none derivatives exhibit their anticancer activity through acting on
1
13
analysis and various spectral data ( H NMR, C NMR and HRmass).
different antitumor biotargets such as tumor necrosis factor TNF
20], antiapoptotic biocomplex Bcl-XL-BH3 [21], many types of phos-
phatases [22–24] and integrin receptor [25].
a
[
2
2
.2. Pharmacological screening
v 3
a b
Based on the aforementioned data and the existing need for
developing superior anticancer agents, we herein describe our
efforts to design and synthesize novel hybrid molecules that could
be used as potential anticancer agents with more potent pharmaco-
logical profile and less toxicity. In doing so, we combined the thia-
zolidinone skeleton with the diphenylthiazole as a COX-scaffold
using the tethering technique in drug discovery, Fig. 3. Considering
the earlier displayed reports on the biological activity of both
diphenylthiazole derivatives and thiazolidinone moiety, we could
infer that the newly designed compounds would exhibit good anti-
tumor activity via, at least in part, inhibiting COX enzymes.
.2.1. Anticancer activity
The newly synthesized compounds were screened in vitro for
their anti-proliferative activity against five cancer cell lines; HCT-
16 (human colon cancer), Caco-2 (human colon carcinoma),
1
MCF-7 (human breast carcinoma), DU-145 (human prostate carci-
noma, epithelial-like cell line) and PC-3 (human Prostate Carci-
noma). The results were expressed in terms of IC50 values (the
concentration that caused a 50% inhibition) where the well-known
anticancer agent doxorubicin was used as positive control. It was
observed from the results (Table 2) that compounds 12a was inac-
tive and 12b showed the least potency with IC50 values of more
than 140.6 lM against the all used cancer cell lines however, 12b
2
. Results and discussion
bearing a sulfonamide group exhibited slightly increased activity.
This observation confirms that the benzylidene moiety at the
5-position of the 4-thiazolidinone is essential for the biological
activity including anticancer effect based on the well-established
previous postulate [16,17,29,30]. Subsequently, extensive SAR
studies were conducted to show how the substitution at 5-position
with various substituted benzylidene or other moieties affected
2.1. Chemistry
The chemistry used for preparing the novel target compounds
was straightforward and the general synthetic pathways are
depicted in Schemes 1 and 2. Benzoin was reacted with thionyl
Fig. 1. Chemical structures of some reported NSAIDs with anticancer activity.

1
34
A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
Fig. 2. Chemical structures of some thiazolidinone and thiazolidinedione derivatives with anticancer activity.
as a molecular mechanism for the anticancer activities of the newly
synthesized compounds. In addition, an increase in the antiprolif-
erative activity was observed by substitution the 5-benzylidene
moiety with electron withdrawing groups as in compounds 15c
and 15d with exception of the 4-chloro analogue 15e. The effect
of using electron withdrawing groups on the activity of compound
1
1
5a was nearly equal to that effect of the sulfamoyl group as in
5b. It can be seen that the position of the substituent on the 5-
benzylidene affects the anticancer activity. The meta-nitro deriva-
tive 15c exerted slightly higher anticancer activity compared to
its para-analog 15d. Similarly, the substitution of 5-benzylidene
moiety with electron donating groups resulted in an increase in
the anticancer activity, but this increase in anticancer activity
due to the electron donating groups in 15f-j is slightly less than
that of the electron withdrawing groups. Finally, the replacement
of the 5-benzylidene moiety with the bulky 5-(naphthalen-1-
ylmethylene) moiety in 16a revealed an increase in the anticancer
activities. These results demonstrate that the effect of the substitu-
tion on the anticancer activity is mainly sterically more than elec-
tronically. Interestingly, compound 16b with a sulfamoyl group
was the most active compound in the series with IC50 values in
Fig. 3. Design strategy of the newly synthesized compounds.
the range 8.88–17.02 lM. Overall, the antiproliferative activity of
the antiproliferative activities. In doing so, the extension of the
chemical structure of compound 12a with furan-2-ylmethylene
moiety in 13, resulted in an observed increase in the anticancer
activity nearly 53–62%. Furthermore, the replacement of the five-
membered furan ring with pyridine ring in compound 14 revealed
an increased activity with IC50 values between 23.68 and
our compounds was essentially affected by the presence of a
substituted 5-benzylidene or more bulky moieties in addition to
a sulfonamide group on the diphenylthiazole system.
2.2.2. COX inhibition assay
6
2.16
ety showed comparable anticancer activity to the furyl derivatives
3. However, a marked increase in the anticancer activity of 15a
lM. Compound 15a with an unsubstituted benzylidene moi-
It is well known that COX enzymes, especially COX-2, are over-
expressed in many types of tumors such as colon cancer, breast
cancer, and melanoma [31–33]. This strong association existing
between COX enzymes and cancer make it a valid target for pre-
vention and treatment of cancer. In an attempt to investigate the
molecular mechanism of the new diphenylthiazole derivatives as
1
was observed on substitution with the COX-2 directing sulfamoyl
group as in compound 15b with IC50 values between 12.04 and
1
9.25 lM suggesting a possible critical role for the COX enzyme
Scheme 1. a Reaction reagents and conditions: (a) SOCl
reflux, 2 h; (e) ClCH COCl, TEA, CH Cl , rt, 5 h; (f) NH SCN, acetone.
2 2 2 4
, pyridine, 1 h; (b) chlorosulfonic acid, CH Cl , 0 °C, stirring, overnight; (c) NH OH, THF, rt, 2 h; (d) thiourea, EtOH,
2
2
2
4

A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
135
Scheme 2. a Reaction reagents and conditions: (a) Aldehyde, CH
COONH , toluene, reflux, 3 h.
3 4
Fig. 4. The proposed mechanism for the formation of thiazolidine-4-one ring and all possible tautomers.
antiproliferative agents, COX inhibition assay was performed at
concentration of 10 M and 0.5 M for COX-1 and COX-2 respec-
of the furan ring in 13 with pyridine in 14 resulted in a marked
increase in the inhibitory activity and selectivity for COX-1 without
any change in the inhibition percentage of COX-2. Moreover, com-
pounds 15g and 15i bearing electron donating groups were the
most active among the substituted benzylidene derivatives with
high COX-2 selectivity.
On the other hand, compounds 15c, 15d and 15e with electron
withdrawing groups showed good inhibitory activity against both
COX subtypes with slight preference to COX-1. It was clear from
the results that the electronic effect on the COX inhibitory activity
was nearly similar but it affects the selectivity profiles. In addition,
l
l
tively, using celecoxib as a positive control. The results were
expressed in terms of inhibition percentages and presented in
Fig. 5. Obviously, most of the compounds, except 12b and 15h,
showed moderate to high inhibitory activities compared to cele-
coxib with different selectivity profiles. Compound 12b showed a
weak inhibitory activity against both COX subtypes which is owing
to the absence of the substituted 5-benzylidene moiety while a
reverse trend of relatively increased activities was observed with
the rest of compounds except compound 15h. The replacement

1
36
A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
Table 1
List of the newly synthesized diphenylthiazole derivatives (12–16) with molecular weights, melting points and yields.
0
Comp.
R
R
Molecular formula
Mol. Wt.
M.P. (°C)
Yield (%)
12a
12b
13
H
SO
H
H
H
C
C
C
18
18
23
H
13
H
14
H
15
N
3
N
4
N
3
OS
2
351.45
430.52
429.51
240–242
146–148
277–278
67
51
81
2
NH
2
O
O
3
S
S
3
2
2
1
1
1
1
4
H
C
C
C
C
24
25
25
25
H
16
H
17
H
18
H
16
N
4
N
3
N
4
N
4
OS
2
2
440.54
439.55
518.63
484.55
218–220
253–254
112–114
261–262
77
74
59
72
5a
5b
5c
H
OS
SO
H
2
NH
2
O
O
3
S
S
3
2
3
1
1
1
1
5d
5e
5f
H
H
H
H
C
C
C
C
25
25
25
26
H
H
H
H
16
N
4
O
3
S
2
484.55
474.00
455.55
485.58
275–277
272–275
283–285
213–215
74
73
71
77
3 2
16ClN OS
17
19
N
3
O
2
S
S
2
2
5g
N
3
O
3
1
1
5h
5i
H
H
C
C
27
26
H
22
19
N
4
OS
OS
2
2
482.62
453.58
272–274
263–265
70
78
H
N
3
1
1
5j
H
H
C
C
26
29
H
19
19
N
3
O
2
S
2
469.58
489.61
231–233
241–244
76
73
6a
H
N
3
OS
2
1
6b
SO
2
NH
2
C
29
H
20
4
N O
3
S
3
568.69
165–166
54
compound 16a with the bulky naphthyl moiety exhibited good
inhibitory activity against COX-1 and COX-2 similar to compounds
compounds 12b, 13, 15a and 15h with weak COX inhibitory activ-
ities showed the lowest anticancer activity. Based on these find-
ings, it could be concluded that there is a strong correlation
between the COX inhibition and the anticancer activity which con-
firms our suggestion of COX inhibition as a possible molecular
mechanism for our compounds as anticancer agents beside other
mechanisms that should be further investigated.
1
5.
It is well established that COX-2 active site is larger than that of
COX-1 and it is characterized by the presence of side
a
pocket allowing accommodation of bulky compounds bearing
additional groups such as sulfonamide or methylsulfone [34].
Based on this fact, it was found that compounds 15b and 16b bear-
ing a sulfonamide moiety exhibited good inhibitory activities with
high selectivity towards COX-2. Generally, it can be concluded that
both free sulfamoyl and the increase in the molecular volume by
addition of aryl-methylene moiety play the major role in increas-
ing inhibitory activities against COX subtypes and they enhance
the COX-2 selectivity.
Comparing the COX inhibition results with that of the antican-
cer screening assay, there was a consistency between both results
where compounds that showed high COX inhibitory activity, in
particular with high COX-2 selectivity, as in compounds 15b and
2.3. Molecular Docking Study
In order to understand the structural basis for the COX-2 activ-
ity, selectivity and the proper binding mode of the newly synthe-
sized diphenylthiazole ligands, two compounds, 12b and 15b,
were selected to be docked into the active sites of COX-2 using
LIGANDFIT embedded in Discovery Studio software [35]. The 3D
crystal structure of COX-2 complexed with a cocrystallized inhibi-
tor (PDB code: 6COX) was selected for this study. It was reported
that the substitution of Ile523 in COX-1 with the less bulky
Val523 in COX-2 creates an additional polar side pocket and
1
6b were the most active antiproliferative agents while the

A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
137
Table 2
Arg513, replaced by a His513 in COX-1, and His90. The structure
of traditional COX-2 inhibitors exploits binding with Arg513 in
the COX-2 side-pocket, often via methylsulfone or sulfonamide
groups, to accomplish their COX-2 selectivity over COX-1.
IC50 values for the newly synthesized compounds on five cancerous cell lines; HCT-
1
16, Caco-2, MCF-7, DU-145, PC-3 cell lines.
Comp. IC50 M)
(
l
HCT-116
Caco-2
MCF-7
DU-145
PC-3
In this regard, we firstly docked one potent and COX-2 selective
compound, 15b, within the active binding site of COX-2. Obviously,
from Fig. 6(B), compound 15b assumed binding mode and interac-
tions similar to the cocrystallized ligand, S-58701, 1-phenylsulf-
onamide-3-trifluoromethyl-5-p-bromophenylpyrazole, in which
positioning the sulfonamide moiety of 15b within the side addi-
tional pocket of COX-2 and forming H-bond with the NH of
Arg513 residue was similar to fitting of same moiety of S-58701
into the same side pocket in addition to forming another H-bond
with NHs of His90 residues, Fig. 6(C). Moreover, the thiazolidinone
ring tends to form additional two H-bonds with Arg-120. On the
other hand, the benzylidene moiety at the 5-position of the 4-thia-
zolidinone seems to protrude outside the binding pocket forming
hydrophobic aromatic stacking interactions with Tyr355 and
Tyr115 residues, Fig. 6(A). In addition, the other phenyl ring of
the Y-shaped diphenylthiazole core was hydrophobically stacked
within a hydrophobic pouch composed of the hydrophobic side
chains of Tyr345, Val349, Tyr385 and Trp387 which correlates well
with the position of the bromophenyl fragment of S-58701 into the
same pocket, Fig. 6(A).
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
2a
2b
3
NA
NA
NA
NA
NA
140.6
71.78
23.68
82.07
12.04
12.20
14.06
29.10
21.01
30.34
75.48
20.58
20.10
19.36
8.88
151.7
75.48
30.34
91.20
13.28
11.84
15.54
30.25
24.12
39.22
82.88
24.36
24.13
21.54
11.10
NA
157.62
79.18
33.3
230.14
83.62
51.06
86.21
19.25
16.51
20.72
41.29
26.3
54.76
68.82
32.54
40.25
21.98
15.90
0.89
294.52
95.46
62.16
95.14
14.36
18.30
20.9
4
5a
5b
5c
5d
5e
5f
5g
5h
5i
82.35
16.50
15.09
20.72
29.50
29.12
42.18
104.34
25.01
21.15
19.58
17.02
1.18
48.28
34.2
78.44
113.96
35.98
39.87
35.32
12.58
1.01
5j
6a
6b
Doxorubicin
0.97
Cells were treated with the test compounds or vehicle for 48 h. Data were reported
as mean ± S.D. (n = 6).
Five human cancer cell lines were used; HCT-116 (human colon cancer), Caco-2
(
human colon carcinoma), MCF-7 (human breast carcinoma), DU-145 (human
prostate carcinoma, epithelial-like cell line) and PC-3 (human Prostate Carcinoma).
Doxorubicin for was used as a positive control in the anticancer screening assay. NA
means Not Active.
On the contrary, the docking of the inactive compound 12b
within the COX-2 active pocket revealed that this compound
assumed a different binding pattern compared to the cocrystallized
ligand, S-58701, Fig. 7(C), where the 4-thiazolidinone ring was
deeply embedded inside the side pocket instead of the sulfonamide
group, Fig. 7(B). Clearly from Fig. 7(A), sulfonamide moiety was
increases the volume of the COX-2 active site that makes it accom-
modate more bulky structures [34]. The presence of such side
pocket allows additional interactions with amino acids such as
Fig. 5. The in vitro COX inhibition percentages of the newly synthesized compounds.
Fig. 6. (A) Docking and binding mode of compound 15b within COX-2 active site (PDB code: 6COX). (B) The superposition of the docked poses 15b (green) and the co-
crystallized S-58701 (cyan) within active site of COX-2. (C) Docking and binding mode of co-crystallized S-58701 into the same COX-2 binding pocket. Hydrogen bonds are
represented by dashed green lines. All hydrogens are removed for the purposes of clarity.

1
38
A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
Fig. 7. (A) Docking and binding mode of compound 12b within COX-2 active site (PDB code: 6COX). (B) The superposition of the docked poses 12b (green) and the co-
crystallized S-58701 (cyan) within active site of COX-2. (C) Docking and binding mode of co-crystallized S-58701 into the same COX-2 binding pocket. Hydrogen bonds are
represented by dashed green lines. All hydrogens are removed for the purposes of clarity.
hydrogen-bonded to Arg-120 residue and the 4-thiazolidinone ring
formed another H-bond with His90 within the additional side cor-
ridor. The different disposition of compound 12b within COX-2
active site could be the reason for the inferior bioactivity which
reflected its low antiproliferative activity, Fig. 7(B).
made on BRUKER Vector 22 (Japan), infrared spectrophotometers
ꢀ1
and were expressed in wavenumber (cm ) using potassium bro-
1
mide disc. The proton magnetic resonance H NMR spectra were
recorded on a Varian Mercury VX-300 NMR spectrometer at
400 MHz and BRUKER APX400 spectrometer at 400 MHz in the
specified solvent, chemical shifts were reported on the d scale and
3
. Conclusions
were related to that of the solvent and J values are given in Hz. 13
C
NMR spectra were obtained on a Bruker APX400 at 100 MHz at
the faculty of pharmacy, Beni-Suef University. Mass spectra were
recorded on Fennigan MAT, SSQ 7000, Mass spectrometer, at
70 eV (EI) at the microanalytical Center, Faculty of Science, Cairo
University. The high resolution mass spectra (HRMS) were recorded
on a Waters Micromass Q-Tof Micro mass spectrometer with a lock
of spray source. Thin layer chromatography, was done using Mache-
rey–Nagel Alugram Sil G/UV254 silica gel plates and methylene cho-
ride-methanol (9.5:0.5) as the eluting system.
A series of diphenylthiazole substituted thiazolidinone deriva-
tives was synthesized and evaluated as anticancer agents against
a panel of cancer cell lines. The COX inhibition assay was performed
in order to investigate the molecular mechanism of their anticancer
potential. The results showed that compounds 15b and 16b were
the most potent anticancer agents with IC50 values between 8.88
and 14.36 lM against the five human cell lines (HCT-116, Caco-2,
MCF-7, DU-145 and PC-3). Interestingly, the COX inhibition assay
results were consistent with that of the cell viability assay where
the potent anticancer compounds showed good COX-2 inhibition
percentages (up to 63%) comparable to celecoxib at a concentration
2-Chloro-1,2-diphenylethanone (8) [26]. A mixture of benzoin
(10 g, 47 mmol) and pyridine (5.7 ml) was heated until a solution
was obtained, then cooled in an ice bath until solid. The mass
obtained was coarsely ground and thionyl chloride (7.5 g, 63 mmol)
was added slowly with vigorous stirring and cooling in an ice bath.
After about an hour, water was added and the solid is coarsely
ground, filtered and washed several times with water. The crude
product was dried by suction and left overnight over calcium
chloride until full dryness. The white powder obtained was recrys-
tallized from ethanol to give colorless crystals of 2-chloro-1,2-
of 0.5 lM. Combining the results of the cell viability assay and COX
inhibition, we can conclude that a strong relationship exists
between the anticancer activity and the COX inhibitory activity of
these compounds, and we can suggest that COX inhibition may con-
tribute, at least in part, to the mechanism of their anticancer activ-
ity. The docking studies also support the results concluded from the
COX assay and the anti-proliferation screening. It was clear that our
ligands that bearing a benzylidene moiety at the 5-position of the 4-
thiazolidinone in addition to a sulfonamide group have high bind-
ing affinities for COX-2 enzyme, which was reflected in their
in vitro activity. In conclusion, these results demonstrate that our
compounds are good leads for subsequent development into poten-
tial anticancer agents. Further experiments are needed to unveil
additional possible molecular mechanisms involved in the antican-
cer activity of our compounds and to evaluate their chemothera-
peutic potential in more details.
+
diphenylethanone 8 (79%). MS (EI) m/z 231.15 (M +1).
4-(1-Chloro-2-oxo-2-phenylethyl)benzenesulfonamide (9). Chlo-
rosulfonic acid (4.0 mL, 60 mmol) was carefully added to a solution
of 2-chloro-1,2-diphenylethanone 8 (2.3 g, 10 mmol) in CH Cl
2 2
(20 ml) at 0 °C. After about 30 min, the ice bath was removed
and the resulting solution was stirred at room temperature for
24 h. The mixture was thereafter poured on ice/water, and
extracted with CH
were washed with saturated aqueous NaHCO
over Na SO . The solvent was removed in vacuo and the obtained
2
Cl
2
(3 ꢁ 40 ml). The combined organic extracts
3
, brine and dried
2
4
4
. Experimental protocols
crude sulfonyl chloride product was used immediately for the next
step. To a solution of the crude sulfonyl chloride (1 g, 3 mmol) in
4.1. Chemistry
THF (20 mL) at room temperature, an excess of NH
4
OH (5 ml of
3
0% w/v) was added dropwise with stirring. The reaction was
Chemical reagents and solvents were obtained from commercial
allowed to proceed for 2 h. The reaction mixture was quenched
sources. Solvents are dried by standard methods when necessary.
Melting points (m.p.) were uncorrected and were carried out by
open capillary tube method using IA 9100MK-Digital Melting Point
Apparatus. Microanalyses were carried out at the microanalytical
Center, Faculty of Science, Cairo University. Infrared spectra were
with water (100 ml) and extracted with ethyl acetate (3 ꢁ 40 ml).
2 4
The organic extract was washed with water and dried over Na SO .
The solvent was removed in vacuo, and the residue was purified by
flash column chromatography (SiO using dichloromethane/
methanol (9.5: 0.5) as eluent to give compound 9 (43%) as a yellow
2
)

A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
139
1
solid. H NMR (400 MHz, DMSO-d
1
6
): d 3.60 (s, 2H, SO
2
NH
2
), 6.24 (s,
167.23 (C@O), 166.66, 157.76, 157.71, 150.23, 147.91, 146.44,
134.63, 131.89, 129.89, 129.52, 129.01, 128.97, 128.77, 128.51,
+
H), 7.23–8.14 (m, 9H). MS (EI) m/z 310.32 (M +1).
,5-Diphenylthiazol-2-amine (10a). [26]. A solution of 2-Chloro-
,2-diphenylethanone, 8 (10 g, 41.5 mmol) in ethanol (40 ml) and
4
122.11, 119.04, 118.93, 114.10. HRMS calcd. for C23
H
15
N
3
O
2
S
2
+
1
[M+H] 429.0606, found 430.0760. Anal. Calcd for: C23
H
15
N
3
O
2
S
2
:
thiourea (3.5 g, 45 mmol) were refluxed for 2 h. The solvent was
removed under vacuum and the solid product was soaked in
C, 64.32; H, 3.52; N, 9.78. Found: C, 64.51; H, 3.36; N, 10.07.
2-((4,5-Diphenylthiazol-2-yl)imino)-5-(pyridin-4-ylmethylene)
1
1
00 mL of 30% NaOH at 50 °C for 5 h. The crude product was
thiazolidin-4-one (14). General Procedure A, orange solid. H NMR
filtered and washed with water to neutrality and then was
recrystallized from 95% ethanol to obtain a yellow solid (75%) of
compound 10a. MS (EI) m/z 252.00 (M ).
(400 MHz, DMSO-d
(d, J = 6.49 Hz, 2H), 7.54 (d, J = 4.01 Hz, 1H), 8.02 (d, J = 4.29 Hz,
1H), 8.71 (d, J = 8.20 Hz, 1H), 12.17 (s, 1H, NH).
6
): d 7.28–7.33 (m, 4H), 7.34–7.41 (m, 6H), 7.48
+
13
C NMR
4
-(2-amino-4-phenylthiazol-5-yl)benzenesulfonamide (10b). Yel-
(101 MHz, DMSO): 174.62 (C@O), 167.23, 163.93, 150.84,
146.07, 145.30, 136.79, 134.75, 132.01, 129.87, 129.82, 129.48,
d
1
low solid, yield 41%. H NMR (400 MHz, DMSO-d
6
): d 1.98 (s, 2H,
C NMR (101 MHz, DMSO): d
13
SO
1
1
2
NH
2
), 7.27–7.90 (m, 9H).
129.35, 128.85, 128.79, 128.75, 128.36, 119.87. HRMS calcd. for
+
70.83, 145.07, 132.44, 129.80, 129.30, 129.11, 128.93, 128.72,
26.53, 124.42, 117.70. MS (EI) m/z 332.27 (M +1).
C
C
24
H
H
16
N
N
4
OS
OS
2
[M+H] 440.0766, found 441.0809. Anal. Calcd for:
: C, 65.43; H, 3.66; N, 12.72. Found: C, 65.62; H,
+
24
16
4
2
2
-Chloro-N-(4,5-diphenylthiazol-2-yl)acetamide (11a). [36]. To a
solution of compound 10a (5 g, 19.8 mmol) and triethylamine
6 g, 59.5 mmol) in CH Cl at 0 °C, chloroacetyl chloride (2.9 g,
5.7 mmol) was added. The reaction mixture was stirred for 1 h
3.57; N, 12.49.
5-Benzylidene-2-((4,5-diphenylthiazol-2-yl)imino)thiazolidin-4-
one (15a). General Procedure A, yellow solid. H NMR (400 MHz,
1
(
2
2
2
DMSO-d
J = 7.58 Hz, 2H), 7.76 (s, 1H), 12.80 (s, 1H, NH).
(101 MHz, DMSO): 170.03 (C@O), 166.49, 157.68, 156.80,
6
): d 7.34–7.41 (m, 8H), 7.51–7.61 (m, 5H), 7.71 (d,
13
at 0 °C then for 5 h at room temperature. The organic layer was
then separated and evaporated in vacuo. The residue obtained
C NMR
d
was purified by flash column chromatography (SiO
2
) using dichlo-
154.12, 146.36, 143.80, 134.59, 134.02, 132.54, 130.79, 130.59,
romethane/methanol (9.5: 0.5) as eluent to give brown solid (62%)
of 2-chloro-N-(4,5-diphenylthiazol-2-yl)acetamide 11a. H NMR
129.88, 129.75, 129.55, 128.92, 128.74, 128.60, 125.44. HRMS calcd
1
+
for C25
17
H N
3
OS
2
[M+H] 439.0813, found 440.0977. Anal. Calcd for:
(
1
400 MHz, DMSO-d
0.48 (s, 1H, NH). MS (EI) m/z 328.05 (M ).
-Chloro-N-(4-phenyl-5-(4-sulfamoylphenyl)thiazol-2-yl)acetam-
6
): d 4.45 (s, 2H, CH
2
), 7.29–7.42 (m, 10H),
25 17 3 2
C H N OS : C, 68.31; H, 3.90; N, 9.56. Found: C, 68.11; H, 3.59; N,
9.46.
+
2
4-(2-((E)-((Z)-5-Benzylidene-4-oxothiazolidin-2-ylidene)amino)-
1
ide (11b). Brown solid, yield 53%. H NMR (400 MHz, DMSO-d
6
): d
4-phenylthiazol-5-yl)benzenesulfonamide (15b). General Procedure
1
1
.99 (s, 2H, SO
H, NH). MS (EI) m/z 407.15 (M ).
-((4,5-Diphenylthiazol-2-yl)imino)thiazolidin-4-one (12a).
mixture of 2-chloro-N-(4,5-diphenylthiazol-2-yl)acetamide, 11a
3 g, 9.1 mmol) and ammonium thiocyanate (1.4 g, 18.2 mmol)
2
NH
2
), 4.96 (s, 2H, CH
2
), 7.34–7.94 (m, 9H), 12.12 (s,
A, brown solid. H NMR (400 MHz, DMSO-d
6
): d 4.20 (s, 2H, SO
2
+
13
1
NH
2
), 7.27–7.60 (m, 14H), 8.05 (s, 1H), 10.02 (s, 1H, NH).
C
2
A
NMR (101 MHz, DMSO): d 172.53 (C@O), 163.13, 161.26, 137.73,
132.89, 130.00, 129.84, 129.65, 129.45, 129.35, 129.14, 129.07,
(
128.93, 128.66, 128.29, 128.07, 127.07, 126.72, 125.77. HRMS
+
was refluxed for 5 h in acetone (40 ml). After the reaction was com-
pleted, the mixture was cooled. The precipitate was collected by
filtration, washed with water and recrystallized from ethanol/
calcd. for C25
Calcd for: C25
57.61; H, 3.26; N, 11.07.
H
H
18
N
4
O
3
S
3
[M+H] 518.0541, found 519.0609. Anal.
: C, 57.90; H, 3.50; N, 10.80. Found: C,
18
N
4
O
3
S
3
acetone mixture to afford compound 6a as a light brown solid.
2-((4,5-Diphenylthiazol-2-yl)imino)-5-(3-nitrobenzylidene)thiaz-
1
olidin-4-one (15c). General Procedure A, orange solid. 1H NMR
H NMR (400 MHz, DMSO-d
0H), 12.17 (s, 1H, NH). 13C NMR (101 MHz, DMSO): d 174.57
C@O), 167.25, 163.87, 146.08, 134.76, 132.01, 129.82, 129.48,
6 2
): d 4.03 (s, 2H, CH ), 7.29–7.49 (m,
1
(400 MHz, DMSO-d
6
): d 7.34–7.60 (m, 9H), 7.85–7.93 (m, 3H),
13
(
8.15–8.34 (m, 2H), 8.53 (d, J = 12.07 Hz, 1H), 8.96 (s, 1H, NH).
C
1
C
C
3
28.86, 128.79, 128.74, 128.36, 35.53 (CH
2
). HRMS calcd. for
NMR (101 MHz, DMSO): d 167.22 (C@O), 148.71, 148.42, 147.68,
135.56, 134.63, 134.63, 131.83, 131.32, 131.07, 129.91, 129.86,
H
H
13
N
N
3
OS
OS
2
[M+H]+ 351.0500, found 352.0615. Anal. Calcd for:
: C, 61.52; H, 3.73; N, 11.96. Found: C, 61.15; H,
18
18
13
3
2
129.58, 129.12, 128.96, 128.75, 124.87, 123.45, 122.62, 118.49,
+
.48; N, 12.03.
-(2-((4-Oxothiazolidin-2-ylidene)amino)-4-phenylthiazol-5-yl)
113.04. HRMS calcd. for C25
485.0715. Anal. Calcd for: C25
11.56. Found: C, 62.15; H, 3.61; N, 11.29.
H
16
N
4
O
3
S
2
[M+H] 484.0664, found
4
16 4 3 2
H N O S : C, 61.97; H, 3.33; N,
1
benzenesulfonamide (12b). Dark orange solid, H NMR (400 MHz,
DMSO-d ): d 4.05 (s, 2H, SO NH ), 4.20 (s, 2H, CH ), 7.24–7.90
m, 9H), 12.12 (s, 1H, NH). C NMR (101 MHz, DMSO): d 174.29
C@O), 168.14, 162.49, 145.65, 137.00, 136.47, 130.11, 129.11,
29.00, 128.94, 127.51, 126.73, 126.54, 34.83 (CH ). HRMS calcd.
6
2
3
2
2
2-((4,5-Diphenylthiazol-2-yl)imino)-5-(4-nitrobenzylidene)thiaz-
1
1
(
(
1
olidin-4-one (15d). General Procedure A, dark orange solid. H NMR
6
(400 MHz, DMSO-d ): d 7.31–7.49 (m, 11H), 7.77–7.86 (m, 2H),
1
3
2
8.28–8.35 (m, 2H), 10.15 (s, 1H, NH). C NMR (101 MHz, DMSO):
d 170.58 (C@O), 156.04, 148.50, 144.31, 135.13, 132.31, 129.75,
129.39, 129.05, 128.96, 128.92, 128.89, 128.72, 128.49, 128.34,
+
for C18
for: C18
H
14
N
4
O
3
S
3
[M+H] 430.0228, found 431.0377. Anal. Calcd
H
14
4 3 3
N O S : C, 50.22; H, 3.28; N, 13.01. Found: C, 50.15; H,
3
.65; N, 13.27.
General procedure A. Synthesis of compounds (13, 14, 15a-j and
6a-b). 2-((4,5-diphenylthiazol-2-yl)imino)-5-(furan-2-ylmethyl-
128.17, 125.94, 117.00, 112.73. HRMS calcd. for C25
H
16
N
4
O
3
S
2
+
[M+H] 484.0664, found 485.0505. Anal. Calcd for: C25
H
16
N
4
O
3
S
2
:
1
C, 61.97; H, 3.33; N, 11.56. Found: C, 62.13; H, 3.51; N, 11.75.
ene)thiazolidin-4-one (13). A mixture of compound 12a (0.5 g,
.57 mmol) and the 2-furaldehyde (0.15 g, 1.57 mmol) was
5-(4-Chlorobenzylidene)-2-((4,5-diphenylthiazol-2-yl)imino)thiaz-
1
olidin-4-one (15e). General Procedure A, yellow solid. 1H NMR
refluxed with ammonium acetate (0.25 g, 3.20 mmol) in 20 mL
anhydrous toluene. The reaction mixture was refluxed for 3 h
and then the solvent was evaporated. The residue was dissolved
in water with acidified with conc. HCl. The product was extracted
into ethyl acetate, dried over sodium sulfate and evaporated under
(400 MHz, DMSO-d
C NMR (101 MHz, DMSO): d 182.87 (C@O), 156.75, 154.77,
132.47, 132.16, 131.39, 130.90, 130.61, 130.17, 129.91, 129.60,
129.01, 128.62, 128.23, 127.77, 127.65, 127.45, 127.35, 126.25.
HRMS calcd. for
474.0557. Anal. Calcd for: C25
8.87. Found: C, 63.47; H, 3.73; N, 8.96.
6
): d 7.29–7.93 (m, 15H), 12.16 (s, 1H, NH).
1
3
+
25
C H16CIN
3
OS
2
[M+H] 473.0423, found
vacuum to obtain the purified target compound 13 as greenish yel-
H16CIN
3
OS
2
,C, 63.35; H, 3.40; N,
1
low solid. H NMR (400 MHz, DMSO-d
6
): d 6.79 (d, J = 3.50 Hz, 1H),
7
8
.11 (t, J = 4.26, 1H), 7.33–7.43 (m, 8H), 7.57 (d, J = 6.88 Hz, 3H),
2-((4,5-Diphenylthiazol-2-yl)imino)-5-(4-hydroxybenzylidene)
thiazolidin-4-one (15f). General Procedure A, dark yellow solid. H
.11 (s, 1H), 12.66 (s, 1H, NH). 13C NMR (101 MHz, DMSO): d
1

1
40
A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
NMR (400 MHz, DMSO-d
6
): d 4.01 (s, 1H, OH), 6.93 (d, J = 8.23 Hz,
13C NMR (101 MHz, DMSO): d 167.83 (C@O), 165.13, 150.71,
2
1
1
1
1
H), 7.32–7.37 (m, 8H), 7.52 (d, J = 9.14 Hz, 4H), 7.63 (s, 1H),
149.75, 138.03, 133.48, 131.69, 129.91, 129.55, 129.53, 129.20,
129.18, 129.03, 128.74, 128.57, 128.56, 128.52, 128.49, 128.43,
2.62 (s, 1H, NH). 1 C NMR (101 MHz, DMSO): d 167.57 (C@O),
3
66.57, 160.22, 157.08, 146.20, 134.70, 133.03, 132.93, 131.90,
128.38, 128.15, 128.06, 126.99, 125.57, 122.54. HRMS calcd. for
+
29.84, 129.43, 129.35, 128.99, 128.91, 128.76, 128.44, 124.97,
C
C
29
H
H
20
N
N
4
O
O
3
S
S
3
[M+H] 568.0698, found 569.0667. Anal. Calcd for:
: C, 61.25; H, 3.54; N, 9.85. Found: C, 61.30; H,
+
21.02, 116.67. HRMS calcd. for C25
H
17
N
3
O
S
2 2
[M+H] 455.0762,
29
20
4
3
3
found 456.0769. Anal. Calcd for: C25
N, 9.22. Found: C, 56.64; H, 3.53; N, 8.97.
-((4,5-Diphenylthiazol-2-yl)imino)-5-(4-hydroxy-3-methoxyben-
zylidene)thiazolidin-4-one (15g). General Procedure A, dark Yellow
H
17
N
3
O
2
S
2
: C, 56.91; H, 3.76;
3.71; N, 10.12.
2
4.2. Pharmacological screening
1
solid. H NMR (400 MHz, DMSO-d
H, OH), 6.93 (s, 1H), 7.18–7.65 (m, 13H), 12.64 (s, 1H, NH).
NMR (101 MHz, DMSO): d 167.56 (C@O), 166.65, 157.39, 156.77,
6
): d 3.78 (s, 3H, CH
3
), 4.02 (s,
4.2.1. Anticancer activity
1
3
1
C
4.2.1.1. Cell culture. The human cancer cell lines HCT-116, Caco-2,
MCF-7, DU-145 and PC-3 were obtained from the American Type
Culture Collection (ATCC, Manassas, VA) and cultured in Dulbecco’s
modified Eagle’s medium/F12 medium (DMEM/F-12), DMEM or
RPMI-1640 media (Gibco, Grand Island, NY) supplemented with
10% fetal bovine serum (FBS; Gibco) according to ATCC recommen-
dation. All the cell lines were cultured at 37 °C in a humidified
1
1
9
49.86, 148.45, 146.52, 134.77, 133.43, 131.88, 129.82, 129.47,
28.92, 128.77, 128.48, 126.29, 125.22, 120.74, 116.52, 113.15,
+
9.98, 55.88. HRMS calcd. for C26
H
H
19
N
3
O
3
S
2
[M+H] 485.0868,
: C, 64.31; H, 3.94;
found 486.0891. Anal. Calcd for: C26
N, 8.65. Found: C, 64.67; H, 4.12; N, 8.97.
-(4-(Dimethylamino)benzylidene)-2-((4,5-diphenylthiazol-2-yl)
imino)thiazolidin-4-one (15h). General Procedure A, orange solid.
19
N
3
O
3
S
2
5
2
incubator containing 5% CO .
1
H NMR (400 MHz, DMSO-d
J = 8.52 Hz, 2H), 7.29–7.61 (m, 13H), 12.54 (s, 1H, NH). C NMR
101 MHz, DMSO): 166.98 (C@O), 161.41, 156.49, 147.63,
34.61, 132.66, 129.87, 129.56, 129.53, 129.03, 128.96, 128.80,
28.74, 128.56, 127.50, 127.49, 124.59, 119.86, 115.36, 46.25
6
): d 3.03 (s, 6H, 2CH
3
), 6.82 (d,
4.2.1.2. Cell viability assay. All the synthesized diphenylthiazole
substituted thiazolidinone derivatives were evaluated in vitro for
their cytotoxic activity against five different cancer cell lines,
HCT-116, Caco-2, MCF-7, DU-145 and PC-3, using the 3-(4,5-
13
(
1
1
d
dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium
bromide
+
(
4
1
2CH
3
). HRMS calcd. for C27
83.1408. Anal. Calcd for: C27
1.61. Found: C, 67.34; H, 4.79; N, 11.36.
-((4,5-Diphenylthiazol-2-yl)imino)-5-(4-methylbenzylidene)thiaz-
H
22
N
4
OS
2
[M+H] 482.1235, found
(MTT) assay as mentioned before [37,38]. Briefly, cancer cells were
seeded in 96-well plates for 24 h, and treated with test compounds
in 5% FBS-supplemented media for 72 h. Controls received DMSO
(vehicle) at a concentration equal to that in drug-treated cells.
After treatment, cells were incubated again in the same media con-
taining 0.5 mg/mL MTT at 37 °C for 2 h. Reduced MTT was solubi-
H
22
N
4
OS
2
: C, 67.19; H, 4.59; N,
2
1
olidin-4-one (15i). General Procedure A, shiny yellow solid. H NMR
400 MHz, DMSO-d ): d 1.55 (s, 3H, CH ), 7.14 (d, J = 12.42 Hz, 2H),
.34–7.66 (m, 12H), 7.71 (s, 1H), 12.70 (s, 1H, NH). C NMR
101 MHz, DMSO): 174.41 (C@O), 167.02, 163.71, 150.62,
45.85, 145.08, 136.58, 134.53, 131.80, 129.61, 129.26, 129.14,
28.64, 128.58, 128.54, 128.44, 128.14, 123.02, 119.65, 25.31.
(
7
6
3
13
lized in DMSO (200 lL/well) for determination of absorbance at
570 nm using a microplate reader, Table 2.
(
d
1
1
4.2.2. Colorimetric COX-1/COX-2 inhibition assay
+
HRMS calcd. for C26
Anal. Calcd for: C26
8.71; H, 3.99 N, 9.67.
-((4,5-Diphenylthiazol-2-yl)imino)-5-(4-methoxybenzylidene)
H
19
N
3
OS
2
[M+H] 453.0970, found 454.1008.
The in vitro inhibition of COX-1/COX-2 was measured using col-
orimetric COX (ovine) Inhibitor Screening Assay Kit (Cayman
Chemical, Ann Arbor, MI, USA) according to the manufacturer’s
instructions and as mentioned before [39]. For COX-1 reaction,
H
19
N
3
OS : C, 68.85; H, 4.22, N, 9.26. Found: C,
2
6
2
1
thiazolidin-4-one (15j). General Procedure A, yellow solid. H NMR
400 MHz, DMSO-d ): d 3.85 (s, 3H, OCH ), 7.14 (d, J = 8.42 Hz, 2H),
10
l
M of celecoxib or test compounds were used while for COX-
M of celecoxib or test compounds were used. After
(
6
3
2, only 0.5 l
7
7
1
1
1
.32–7.41 (m, 8H), 7.56 (d, J = 7.56 Hz, 2H), 7.65 (d, J = 8.42 Hz, 2H),
the addition of arachidonic acid and incubation, the absorbance
was measured at 590 nm using plate reader, Fig. 5.
.71 (s, 1H), 12.60–12.71 (s, 1H). 13C NMR (101 MHz, DMSO): d
70.58 (C@O), 156.04, 148.50, 144.31, 135.13, 132.31, 129.75,
29.39, 129.14, 129.05, 128.96, 128.92, 128.89, 128.72, 128.49,
4.3. Molecular Docking Study
28.34, 125.94, 117.00, 112.73, 53.50 (OCH
3
). HRMS calcd. for C26-
[M+H] 469.1954, found 470.1141. Anal. Calcd for: C26-
: C, 66.50; H, 4.08; N, 8.95. Found: C, 66.64; H, 4.38; N,
+
H
H
19
N
N
3
O
O
2
S
S
2
The binding sites were generated from the co-crystallized
ligand, S-58701, within COX-2 protein structure (PDB code:
6COX). Selected two ligands, 12b and 15b, were energy minimized
using CHARMm ForceField and then docked into the former pre-
pared proteins active sites using LIGANDFIT imbedded into Discov-
ery Studio Software [35] with the following docking protocol: (i)
number of Monte Carlo search trials = 30,000, search step for tor-
sions with polar hydrogens = 30°. (ii) The Root Mean Square Differ-
ence (RMS) threshold for ligand-to-binding site shape match was
set to 2.0 employing a maximum of 1.0 binding site partitions
and 1.0 site partition seed. (iii) The interaction energies were
assessed employing Consistent Force Field (CFF) force field with a
non-bonded cutoff distance of 10.0 Å and distance-dependent
dielectric. An energy grid extending 3.0 Å from the binding site
was implemented. (iv) Rigid body ligand minimization parameters
were: 10 iterations of steepest descend (SD) minimization fol-
lowed by 20 Broyden–Fletcher–Goldfarb–Shanno (BFGS) iterations
applied to every successful orientation of the docked ligand. (v) A
maximum of 10 diverse docked conformations/poses of optimal
interaction energies were saved. (vi) The saved conformers/ poses
19
3
2
2
9
.09.
-((4,5-Diphenylthiazol-2-yl)imino)-5-(naphthalen-1-ylmethyl-
ene)thiazolidin-4-one (16a). General Procedure A, dark orange
2
1
solid. H NMR (400 MHz, DMSO-d
6
): d 7.23–7.49 (m, 10H), 7.60–
7
.83 (m, 4H), 7.98–8.13 (m, 3H), 8.35 (d, J = 19.74 Hz, 1H), 12.77
1
3
(
s, 1H, NH). C NMR (101 MHz, DMSO): d 168.14 (C@O), 165.45,
1
1
1
51.03, 150.06, 138.35, 133.80, 132.00, 129.86, 129.84, 129.52,
29.49, 129.34, 129.05, 128.88, 128.87, 128.83, 128.80, 128.74,
28.69, 128.47, 128.37, 127.30, 125.88, 122.85, 100.00. HRMS
Calcd for
C
C
[M+H]+ 489.0970, found 490.1122. Anal. Calcd for:
: C, 71.14; H, 3.91; N, 8.58. Found: C, 71.39; H, 3.78;
29
H
H
19
N
N
3
OS
OS
2
29
19
3
2
N, 8.76.
-(2-((E)-((Z)-5-(Naphthalen-1-ylmethylene)-4-oxothiazolidin-2-
ylidene)amino)-4-phenylthiazol-5-yl)benzenesulfonamide (16b).
General Procedure A, light brown solid. H NMR (400 MHz,
DMSO-d ): d 4.12 (s, 2H, SO NH ), 7.23–7.49 (m, 10H), 7.60–7.83
m, 2H), 7.98–8.12 (m, 3H), 8.13–8.38 (m, 2H), 12.71 (s, 1H, NH).
4
1
6
2
2
(

A.H. Abdelazeem et al. / Bioorganic Chemistry 57 (2014) 132–141
141
were further energy-minimized within the binding site for a max-
imum of 1000 rigid-body iterations.
L. Luo, K. Raha, C.S. Sherk, C.-M. Sung, D. Sutton, P.J. Tummino, R.J. Wegrzyn,
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Acknowledgment
This work was supported by grant Number (1-434-2773) from
The Deanship of Scientific Research, Taif University, Saudi Arabia.
(
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