New triarylpyrazoles as broad-spectrum anticancer agents: Design, synthesis, and biological evaluation

Article abstract of DOI:10.1016/j.ejmech.2013.04.067

A new series of diarylureas and diarylamides possessing 1,3,4-triarylpyrazole scaffold was designed and synthesized. Their in vitro antiproliferative activities against NCI-60 cell line panel were tested. Most of the compounds showed strong and broad-spec

Full text of DOI:10.1016/j.ejmech.2013.04.067

European Journal of Medicinal Chemistry 65 (2013) 315e322
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
New triarylpyrazoles as broad-spectrum anticancer agents: Design,
synthesis, and biological evaluation
Mohammed I. El-Gamal c, Yi Seul Park a , Dae Yoon Chi , Kyung Ho Yoo ,
Chang-Hyun Oh
a
a
,
b
,
,
d
d
e
a,b,
*
Center for Biomaterials, Korea Institute of Science and Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea
Department of Biomolecular Science, University of Science and Technology, 113 Gwahangno, Yuseong-gu, Daejeon 305-333, Republic of Korea
Department of Medicinal Chemistry, Faculty of Pharmacy, University of Mansoura, Mansoura 35516, Egypt
Department of Chemistry, Sogang University, Seoul, Republic of Korea
b
c
d
e
Chemical Kinomics Research Center, Korea Institute of Science and Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 25 January 2013
Received in revised form
A new series of diarylureas and diarylamides possessing 1,3,4-triarylpyrazole scaffold was designed and
synthesized. Their in vitro antiproliferative activities against NCI-60 cell line panel were tested. Most of
the compounds showed strong and broad-spectrum antiproliferative activities. Compound 18 exerted
sub-micromolar IC50 values over all the subpanels of nine different cancer types. Its IC50 value over MDA-
MB-435 melanoma cell line was 27 nM. Compounds 10e13, 22, and 23 possessing urea spacer exerted
lethal effect over the NCI-60 panel with mean %inhibitions more than 100% in single-dose testing.
Compounds 13 and 23 with urea linker and 3 ,5 -bis(trifluoromethyl)phenyl terminal ring showed the
highest mean %inhibition over the NCI-60 panel in single-dose testing, and showed high potencies and
broad-spectrum anticancer activities in five-dose testing.
29 April 2013
Accepted 30 April 2013
Available online 13 May 2013
0
0
Keywords:
Anticancer
Cytotoxicity
Diarylurea
Diarylamide
Ó 2013 Elsevier Masson SAS. All rights reserved.
1,3,4-Triarylpyrazole
1. Introduction
contemporary medicinal chemistry. There has been an urgent need
for development of more efficient anticancer agents with minimal
Cancer is a generic term for a large group of diseases that can
affect any part of the body. Cancer is a major leading cause of death
worldwide, and it accounted for 7.6 million death cases (around
side effects.
Diarylureas and diarylamides have been highlighted as potential
antiproliferative agents against a variety of cancer cell lines [2e17].
Ò
13% of all deaths) in 2008 according to WHO reports. Lung, stom-
Sorafenib (Nexavar ) is an example of anticancer diarylureas that
ach, liver, colon, and breast cancer cause the most cancer deaths
each year. About 30% of cancer deaths are due to the five leading
behavioral and dietary risks: high body mass index, low fruit and
vegetable intake, lack of physical activity, tobacco use, and alcohol
intake. More than 70% of all cancer deaths occurred in low- and
middle-income countries. Deaths from cancer worldwide are pro-
jected to continue to rise to over 13 million in 2030 [1]. Inspite of
the extensive efforts and investment in research, the management
of human malignancies still constitutes a major challenge for
has been approved by the U.S. Food and Drug Administration (FDA)
for treatment of advanced renal cancer [18]. It has also been
approved in Europe for treatment of hepatocellular carcinoma
(HCC) [19]. Sorafenib is currently subjected to clinical trials for
Ò
other types of cancer. Imatinib (Gleevec ) is an example of diary-
lamides which is used for treatment of chronic myeloid leukemia
(CML) with diminished side effects [20]. We have previously re-
ported a series of diarylureas and diarylamides possessing 1,3,4-
triarylpyrazole scaffold as potential antiproliferative agents
against human melanoma cell lines [12]. In the present investiga-
tion, we present positional isomers of the previously reported
compounds (Fig. 1) in order to compare their potencies and to
optimize the scaffold. The cytotoxicities of the new analogs were
tested against NCI-60 cancer cell line panel of nine different cancer
types.
*
Corresponding author. Center for Biomaterials, Korea Institute of Science and
Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea. Tel.: þ82 2
9
58 5160; fax: þ82 2 958 5308.
E-mail addresses: drmelgamal2002@yahoo.com (M.I. El-Gamal), choh@kist.re.kr
(
C.-H. Oh).
0223-5234/$ e see front matter Ó 2013 Elsevier Masson SAS. All rights reserved.
http://dx.doi.org/10.1016/j.ejmech.2013.04.067

316
M.I. El-Gamal et al. / European Journal of Medicinal Chemistry 65 (2013) 315e322
O
CF3
O
H3C
O
Cl
N
H
O
N
HN
N
N
N
N
H
N
H
HN
N
N
CH3
Sorafenib
Imatinib
A
R1
O
O
Cl
Cl
N
N
N
N
NH
O
B
O
HN
N
N
R2
1
2
A = H, CH3
B = Ar, ArNH
R
R
= H, CH3
= Ar, ArNH
The target compounds 9-28
Fig. 1. Structures of Sorafenib, Imatinib, reported 1,3,4-triarylpyrazole derivatives [12], and the target compounds 9e28.
2
. Results and discussion
4
methyl group of 2 using KMnO gave the corresponding carboxylic
acid 3 [22], which was further esterified with methanol in the
2.1. Chemistry
presence of acetyl chloride afforded the corresponding methyl ester
4. The pyridyl derivative 5 was obtained by treatment of 4 with 4-
The target triarylpyrazole compounds 9e28 were synthesized
as illustrated in Scheme 1. Methylation of the phenolic hydroxyl
group of 2-chloro-5-methylphenol (1) using dimethyl sulfate gave
-chloro-2-methoxy-4-methylbenzene (2) [21]. Oxidation of the
picoline in the presence of lithium hexamethyldisilazide (LiHMDS).
Cyclization to the pyrazole compound 6 was carried out by treat-
ment of 5 with dimethylformamide dimethyl acetal (DMF-DMA),
and subsequent treatment with hydrazine monohydrate [12,23].
1
OH
OCH3
OCH3
OCH3
Cl
Cl
Cl
Cl
a
b
c
d
OH
O
CH3
CH3
Cl
CH3
O
O
1
2
3
4
OCH3
OCH3
Cl
OCH3
Cl
N
N
e
g
f
NH
N
NO2
Cl
O
N
N
N
5
6
7
OCH3
OH
Cl
OCH3
Cl
N
N
j
h
N
N
N
O
O
N
HN
HN
N
N
HN Ar
HN Ar
9
-13
19-23
NH2
N
i
OCH3
OH
Cl
Cl
8
N
j
N
N
N
O
O
HN
HN
N
N
Ar
Ar
14-18
24-28
ꢀ
Scheme 1. Reagents and conditions: (a) (CH
3
)
SO
2 4
, K
2
CO
3
, acetone, reflux, 1 h; (b) KMnO
4
, C
H
5 5
N, H
2
O, 50 C, 24 h, then rt, 13 h, 90%; (c) acetyl chloride, CH
3
OH, rt, 15 h, 85%; (d) 4-
picoline, LiHMDS, THF, rt, overnight, 45%; (e) (i) DMF-DMA, rt, 18 h, (ii) hydrazine monohydrate, C
2
H
5
OH, rt, overnight, 81%; f) 1-iodo-3-nitrobenzene, K CO
2
3
, CuI, L-proline, DMSO,
ꢀ
ꢀ
9
8
0
C, 8 h, 91.4%; g) H
, CH Cl
2
, Pd/C, THF, rt, 2 h, 64.8%; h) appropriate aryl isocyanate, THF, rt, 12 h, 32e85%; i) appropriate benzoic acid derivative, HOBt, EDCI, TEA, DMF, 80 C, 12 h, 35e
4%; j) BBr
3
2
2
, ꢁ78 ꢀC, 30 min; then rt, 1 h, 40e88% (19e23), 36e72% (24e28).

M.I. El-Gamal et al. / European Journal of Medicinal Chemistry 65 (2013) 315e322
Table 1 (continued )
317
1
,3,4-Triarylpyrazole derivative 8 with amino group was prepared
through N-arylation of 6 using 1-iodo-3-nitrobenzene in the
R1
R2
Compound
no.
Yield%
57
Melting
point ( C)
ꢀ
presence of anhydrous K
2
CO
3
, CuI, and
L-proline, and subsequent
reduction of the nitro group of 7 using Pd/C in hydrogen atmo-
sphere. Reaction of the amino group of 8 with the appropriate aryl
isocyanate derivatives afforded the corresponding urea derivatives
N
2
1
2
H
H
197e200
238e241
H
3
F C
9
e13. Synthesis of the methoxy compounds 14e18 with amide
linker was carried out by condensation of the amino compound 8
with the appropriate benzoic acid derivatives in the presence of
HOBt, EDCI, and triethylamine. Demethylation of the methoxy
group of 9e18 using BBr afforded the corresponding hydroxyl
3
derivatives 19e28. Table 1 illustrates structures of the final com-
Cl
N
2
50
H
3
F C
F
F
3
C
2
2
3
4
H
N
87
194e196
>280
H
pounds, their yield percentages, and their melting points.
3
C
H
H
36
39
Table 1
Structures of the target compounds, and their yield percentages and melting points.
Br
H C
R1
O
25
242e245 (dec.)
3
Cl
N
N
O
2
2
6
7
H
H
29
70
156e159
190e192
F
3
C
HN
N
R2
Cl
3
F C
Compound
no.
R1
R2
Yield%
Melting
point ( C)
ꢀ
O
N
28
H
72
148e151
3
F C
9
CH
CH
3
3
N
32
57
194e197
218e221
H
Cl
N
1
1
1
0
1
2
H
Cl
2
.2. Antiproliferative activities against 60 cell line panel at the NCI
N
CH
CH
3
3
66
68
195e198
226e228
H
F3C
Cl
2.2.1. Single-dose testing
Structures of the synthesized compounds were submitted to
National Cancer Institute (NCI), Bethesda, Maryland, USA [24], and
the 18 compounds shown in Fig. 2 were selected on the basis of
degree of structural variation and computer modeling techniques
for evaluation of their antineoplastic activity. The selected com-
pounds were subjected to in vitro anticancer assay against tumor
cells in a full panel of 60 cell lines taken from nine different tissues
N
H
F3C
F3C
F3C
13
CH
3
N
85
244e247
H
(
blood, lung, colon, CNS, skin, ovary, kidney, prostate, and breast).
The compounds were tested at a single-dose concentration of
M, and the percentages of growth inhibition over the 60 tested
1
1
4
5
CH
CH
3
3
35
60
214e215
218e221
10 m
cell lines were determined. The mean inhibition percentages for
each of the tested compounds over the full panel of cell lines are
illustrated in Fig. 2.
Br
H3C
The results showed that the methoxy derivatives 10, 11, 14,
15, 17, and 18 were more active than the corresponding hydroxyl
derivatives 20, 21, 24, 25, 27, and 28. But the hydroxyl com-
pounds 22 and 23 were exceptionally more active than the
corresponding methoxy derivatives 12 and 13. In addition, the
urea derivatives 11, 12, and 22 exerted higher activities than the
corresponding analogs with amide spacer 16, 17, and 27. This
may be attributed to that the longer linker, urea moiety, may
geometrically permit appropriate fitting of the molecule at the
receptor site. Or the terminal NH group of the urea moiety may
form additional hydrogen bond(s) at the receptor site. Any or
both of these effects would enable optimal drugereceptor
interaction, and hence higher antiproliferative activity. Com-
pounds 10e13, 22, and 23 containing urea spacer exerted lethal
effect over the NCI-60 panel with mean percentage inhibitions
more than 100%.
1
1
1
6
7
8
CH
CH
CH
3
3
3
69
41
84
171e174
232e235
167e168
F3C
Cl
F3C
O
N
F3C
1
2
9
0
H
H
N
40
88
201e204
184e187
H
Cl
N
H
Cl

318
M.I. El-Gamal et al. / European Journal of Medicinal Chemistry 65 (2013) 315e322
Fig. 2. Mean inhibition percentages observed with the final compounds in single-dose (10
mM) 60 cancer cell line screening. Mean %inhibition represents the mean inhibition
percentages over the 60 cell lines. The inhibition percentages were calculated by subtracting the growth percentages from 100.
The effects of substituents on the terminal phenyl ring were also
studied. Compounds 10e13 showed higher mean %inhibitions than
compound 9. So it can be concluded that chloro and/or tri-
fluoromethyl substituents on the terminal phenyl ring of urea de-
rivatives enhance the cytotoxic activity. Compound 16 with m-
the NCI-60 panel are illustrated in Fig. 3. At 10 mM concentration,
both compounds showed lethal effects (>100% inhibition) over 45
cell lines. Both compounds demonstrated broad-spectrum cyto-
toxicities over all the nine tested cancer types.
(
trifluoromethyl)phenyl terminal ring was more active than com-
2.2.2. Five-dose testing
pound 14 with terminal phenyl ring. Also compounds 18 and 28
Compounds 10e14, 16e18, 20e23, and 28 with promising re-
sults in single-dose testing were further tested in a five-dose
testing mode, in order to determine their IC50 values over the 60
cancer cell lines. The mean IC50 values of these 13 compounds over
the nine cancer types are shown in Table 2.
0
0
with terminal 4 -morpholino-3 -(trifluoromethyl)phenyl moiety
0
were more active than 14 and 24 with terminal phenyl ring. So 3 -
0
0
(
trifluoromethyl)phenyl and 4 -morpholino-3 -(trifluoromethyl)
phenyl terminal moieties are favorable for activity. On the other
0
0
hand, compounds 15 and 17 with 4 -bromo-3 -methylphenyl and
4
As shown in Table 2, most of the compounds exhibited high
potency (in sub-micromolar and micromolar scale) over all the nine
0
0
-chloro-3 -(trifluoromethyl)phenyl terminal rings, respectively,
were less active than compound 14. Similarly, compounds 25 and
7 showed lower activities than compound 24 with terminal
phenyl ring. So 4 -bromo-3 -methylphenyl and 4 -chloro-3 -(tri-
fluoromethyl)phenyl terminal moieties are unfavorable for anti-
cancer activity.
cancer types. Most of the mean IC50 data were less than 10
mM. Of
2
special interest, compound 18 with methoxy group, amide linker,
0
0
0
0
0
0
and 4 -morpholino-3 -(trifluoromethyl) terminal ring showed the
highest potencies over all the nine subpanels of nine different
cancer types. All its mean IC50 values were in sub-micromolar
0
0
Among all the target compounds, compounds 13 and 23 with
range. Compounds 13 and 23 with urea spacer and 3 ,5 -bis(tri-
fluoromethyl)phenyl terminal ring exerted sub-micromolar IC50
values over the nine subpanels of nine cancer types.
0
0
urea spacer and 3 ,5 -bis(trifluoromethyl)phenyl terminal ring
showed the highest mean %inhibitions. So these moieties are
favorable for anticancer activity of this series of compounds. The
steric and/or electronic effect(s) of two trifluoromethyl groups
could contribute to affinities of these compounds to the receptor
site. The %inhibitions of these two compounds over each cell line of
The IC50 values of the 13 compounds tested in five-dose mode
over the most sensitive cell line of each subpanel are summarized
in Table 3. From these data, we find that HOP-92 non-small cell lung
cancer was the most susceptible cell line to the target compounds.
Fig. 3. %inhibition expressed by compounds 13 and 23 at a single-dose concentration of 10 mM over the NCI-60 cancer cell lines.

M.I. El-Gamal et al. / European Journal of Medicinal Chemistry 65 (2013) 315e322
319
Table 2
Mean IC50 values (
m
M) of the tested compounds over in vitro subpanel cancer cell lines.a
Subpanel cancer cell linesb
I
II
III
IV
V
VI
1.31
VII
VIII
IX
1.12
Compound No.
10
1.00
0.84
1.02
0.75
0.53
1.55
2.76
0.27
1.08
1.56
0.87
0.88
2.15
1.09
1.23
1.13
0.89
1.24
0.73
0.61
1.54
4.06
0.35
1.38
2.27
0.82
0.76
2.65
0.98
0.88
0.97
0.83
0.89
4.50
2.46
0.44
1.06
2.29
0.71
0.84
2.55
1.15
1.01
1.16
0.79
1.88
1.92
6.54
0.63
1.97
2.37
1.04
0.90
8.69
1.04
1.02
1.10
0.78
0.60
1.50
3.84
0.32
1.17
1.35
0.86
0.78
3.18
1
1
1
1
1
1
1
2
2
2
2
2
1
2
3
4
6
7
8
0
1
2
3
8
0.99
1.19
0.77
2.37
2.71
5.60
0.42
1.45
3.38
0.96
0.79
15.16
0.92
1.17
0.71
0.79
1.78
4.28
0.42
2.38
4.72
1.25
0.76
11.20
1.08
1.43
0.85
2.79
2.10
8.43
0.59
1.72
4.04
1.04
0.86
18.49
1.05
1.24
0.95
1.03
5.24
3.15
0.52
1.17
3.03
0.81
0.93
11.33
a
Mean IC50 values were calculated by dividing the summation of IC50 values of the compound over cell lines of the same cancer type by the number of cell lines in the
subpanel.
b
I: Leukemia; II: Non-Small Cell Lung Cancer; III: Colon Cancer; IV: CNS Cancer; V: Melanoma; VI: Ovarian Cancer; VII: Renal Cancer; VIII: Prostate Cancer; IX: Breast Cancer.
At this cell line, all the thirteen tested compounds demonstrated
IC50 values in sub-micromolar scale. In addition, compound 18
showed 2-digit nanomolar IC50 value, 27 nM, over MDA-MB-435
melanoma cell line. Among all the target compounds, compounds
tested at a single dose of 10 mM at the NCI over 60 cell line panel,
and thirteen of them were subsequently tested in five-dose testing
mode. Compounds 10e13, 22, and 23 with urea spacer exerted
lethal effect over the NCI-60 panel with mean %inhibitions more
than 100% in single-dose testing. Among them, compounds 13 and
11, 13, 14, 18, and 23 showed the highest potencies with sub-
0
0
micromolar IC50 values over the nine different cell lines.
It is noteworthy that compounds 10, 12, 13, 17, 22, and 23 were
more potent against MDA-MB-435 melanoma cell line than the
corresponding positional isomers with para-disubstituted central
phenyl ring [12]. The IC50 values of these six compounds with meta-
disubstituted central phenyl ring were in sub-micromolar scale,
while IC50 values of the para-disubstituted phenyl isomers were in
micromolar range.
23 possessing 3 ,5 -bis(trifluoromethyl)phenyl terminal ring also
showed the highest mean %inhibitions in single-dose testing
(128.13% and 134.64%, respectively). Both compounds and com-
pound 18 demonstrated highly potent and broad-spectrum anti-
cancer activities over all the tested nine cancer types. Further
modifications of these compounds in order to optimize the scaffold
and to improve their potencies are currently in progress.
4. Experimental
3
. Conclusions
4.1. General
A new series of diarylurea and diarylamide derivatives pos-
sessing 1,3,4-triarylpyrazole scaffold was synthesized based on our
previous literature studies, and as a continuation of our ongoing
anticancer development program. Eighteen final compounds were
All melting points were obtained on a Walden Precision Appa-
ratus Electrothermal 9300 apparatus and are uncorrected. Mass
spectra (MS) were taken in ESI mode on a Waters 3100 Mass
Table 3
IC50 values (
m
M) of the tested compounds over the most sensitive cell line of each subpanel.
Cancer cell lines
RPMI-8226a
HOP-92b
KM12c
SF-295d
MDA-MB-435e
OVCAR-3f
A498g
PC-3h
MDA-MB-468i
Compound No.
10
0.39
0.34
0.87
0.31
0.85
1.57
1.62
0.34
1.03
1.34
0.58
0.48
1.40
0.15
0.56
0.50
0.14
0.32
0.56
0.67
0.12
0.68
0.51
0.38
0.34
0.56
0.81
0.57
1.06
0.53
0.56
1.66
2.06
0.25
1.01
1.33
1.00
0.51
1.94
0.69
0.31
0.84
0.40
0.34
1.02
1.38
0.15
Not tested
0.96
0.74
0.62
1.71
0.94
0.87
0.68
0.89
0.19
0.71
0.66
0.027
1.10
1.49
0.66
0.91
2.04
0.70
0.58
0.89
0.41
0.33
1.20
2.42
0.22
1.05
1.06
0.83
0.40
4.81
1.14
0.96
0.84
0.61
0.36
0.69
2.70
0.23
0.94
2.24
1.40
0.79
15.50
0.55
0.37
0.69
0.25
0.43
1.23
1.29
0.25
0.83
0.95
0.59
0.32
2.15
0.57
0.60
0.83
0.31
0.35
1.32
3.69
0.18
0.73
0.98
0.54
0.27
7.04
1
1
1
1
1
1
1
2
2
2
2
2
1
2
3
4
6
7
8
0
1
2
3
8
a
Leukemia cell line.
b
Non-small cell lung cancer cell line.
Colon cancer cell line.
CNS cancer cell line.
c
d
e
f
Melanoma cell line.
Ovarian cancer cell line.
Renal cancer cell line.
Prostate cancer cell line.
Breast cancer cell line.
g
h
i

320
M.I. El-Gamal et al. / European Journal of Medicinal Chemistry 65 (2013) 315e322
Detector (Waters, Milford, MA, USA). Nuclear magnetic resonance
NMR) spectroscopy was performed using a Bruker ARX-300,
00 MHz (Bruker Bioscience, Billerica, MA, USA) and a Bruker
The reaction mixture was stirred at room temperature for 12 h. The
mixture was evaporated under reduced pressure, and the residue
was purified by column chromatography (silica gel, hexaneeethyl
acetate 1:5 v/v) to yield the target compounds 9e13.
(
3
ARX-400, 400 MHz (Bruker Bioscience, Billerica, MA, USA) with
TMS as an internal standard. Purities of the target compounds 9e28
(>95%) were determined by LC-MS analysis using the following
4.5.1. 1-(3-(3-(4-Chloro-3-methoxyphenyl)-4-(pyridin-4-yl)-1H-
system: Waters 2998 photodiode array detector, Waters 3100 mass
detector, Waters SFO system fluidics organizer, Waters 2545 binary
gradient module, Waters reagent manager, Waters 2767 sample
pyrazol-1-yl)phenyl)-3-phenylurea (9)
1
3
H NMR (400 MHz, CDCl ) d 10.37 (brs, 1H), 8.98 (brs, 1H), 8.56
(s, 2H), 8.14 (s, 1H), 7.64e7.57 (m, 2H), 7.48 (dd, 1H, J ¼ 1.4 Hz,
manager, SunfireÔ C18 column (4.6 ꢂ 50 mm, 5
Solvent gradient ¼ 95% A at 0 min, 1% A at 5 min; solvent A: 0.035%
trifluoroacetic acid (TFA) in water; solvent B: 0.035% TFA in CH OH;
mm particle size);
8.3 Hz), 7.43e7.40 (m, 2H), 7.36e7.25 (m, 6H), 7.16e7.13 (m, 2H),
13
3
7.07e7.03 (m, 1H), 3.81 (s, 3H); C NMR (75 MHz, CDCl ) d 155.0,
3
153.2, 153.0, 150.1, 140.0, 139.0, 137.0, 136.2, 130.8, 130.3, 130.2,
129.8, 129.2, 127.5, 125.1, 123.0, 121.6, 121.1, 120.8, 118.6, 114.8, 112.2,
flow rate ¼ 3.0 mL/min; the area under curve (AUC) was calculated
using Waters MassLynx 4.1 software. Unless otherwise noted, all
solvents and reagents were commercially available and used
without further purification.
þ
110.7, 56.2; MS m/z: 497.15 [M þ H] .
4.5.2. 1-(3-(3-(4-Chloro-3-methoxyphenyl)-4-(pyridin-4-yl)-1H-
pyrazol-1-yl)phenyl)-3-(3,4-dichlorophenyl)urea (10)
1
4.2. Synthesis of 4-(3-(4-chloro-3-methoxyphenyl)-1H-pyrazol-4-yl)
6
H NMR (300 MHz, DMSO-d ) d 9.15 (brs,1H), 9.08 (brs,1H), 8.99
pyridine (6)
(s, 1H), 8.55 (d, 2H, J ¼ 4.4 Hz), 8.16 (s, 1H), 7.92 (s, 1H), 7.58e7.47
(
m, 3H), 7.44 (d, 1H, J ¼ 5.5 Hz), 7.38e7.35 (m, 4H), 7.23 (s, 1H), 7.07
13
It was synthesized by the previously reported 5-step method
(d,1H, J ¼ 8.0 Hz), 3.78 (s, 3H); C NMR (75 MHz, DMSO-d
6
) d 154.4,
ꢀ
1
[12,23]. mp: 248e251 C; H NMR (400 MHz, DMSO-d
6
)
d
13.39
152.2,149.9,140.5,139.7,139.5,132.7, 131.0,130.5,130.1,130.0,129.1,
(
brs, 1H), 8.48 (d, 2H, J ¼ 6.0 Hz), 8.14 (brs, 1H), 7.45 (d, 1H,
122.6, 121.3, 119.6, 119.4, 118.5, 116.9, 112.5, 112.3, 108.9, 55.9; MS m/
z: 565.13 [M þ H] .
þ
J ¼ 8.0 Hz), 7.28 (d, 2H, J ¼ 5.9 Hz), 7.18 (s, 1H), 6.97 (dd, 1H,
þ
J ¼ 1.5 Hz, J ¼ 8.2 Hz), 3.77 (s, 3H); MS m/z: 287.0 [M þ H] .
4.5.3. 1-(3-(3-(4-Chloro-3-methoxyphenyl)-4-(pyridin-4-yl)-1H-
4
.3. 4-[3-(4-Chloro-3-methoxyphenyl)-1-(3-nitrophenyl)-1H-
pyrazol-1-yl)phenyl)-3-(3-(trifluoromethyl)phenyl)urea (11)
1
pyrazol-4-yl]pyridine (7)
6
H NMR (300 MHz, DMSO-d ) d 9.12 (s, 2H), 8.98 (s, 1H), 8.56 (d,
2
H, J ¼ 5.3 Hz), 8.17 (s, 1H), 8.05 (s, 1H), 7.60 (d, 1H, J ¼ 4.7 Hz), 7.56
A mixture of compound 6 (0.5 g, 1.7 mmol), 1-iodo-3-
(d,1H, J ¼ 1.8 Hz), 7.52 (t, 2H, J ¼ 3.7 Hz), 7.48 (d, 1H, J ¼ 3.3 Hz), 7.45
(s, 1H), 7.39 (d, 2H, J ¼ 5.9 Hz), 7.33 (d, 1H, J ¼ 7.5 Hz), 7.24 (d, 1H,
nitrobenzene (0.9 g, 3.5 mmol), K
2
3
CO (0.7 g, 5.2 mmol), CuI
13
(
(
0.033 g, 0.2 mmol), and
7 mL) was heated at 90 C under nitrogen atmosphere for 12 h. The
L
-proline (0.04 g, 0.2 mmol) in DMSO
J ¼ 1.6 Hz), 7.08 (dd, 1H, J ¼ 1.7 Hz, 8.1 Hz), 3.78 (s, 3H); C NMR
ꢀ
6
(75 MHz, DMSO-d ) d 154.9, 153.0, 150.3, 149.5, 141.1, 140.9, 140.3,
cooled reaction mixture was partitioned between water and ethyl
acetate. The organic phase was washed with brine (3 times) and
140.0, 133.2, 130.6, 130.4, 129.8, 129.6, 123.1, 122.9, 122.5, 121.8,
121.7, 120.1, 117.4, 114.8, 113.1, 112.8, 109.4, 56.4; MS m/z: 565.07
þ
dried over anhydrous Na
2
SO
4
. After evaporation of the organic
[M þ H] .
solvent, the residue was purified by column chromatography (silica
gel, hexaneeethyl acetate 1:5 v/v) to yield compound 7 (0.85 g,
4.5.4. 1-(4-Chloro-3-(trifluoromethyl)phenyl)-3-(3-(3-(4-chloro-3-
1
91.4%). H NMR (400 MHz, DMSO-d
6
)
d
9.34e9.29 (m, 1H), 8.82e
methoxyphenyl)-4-(pyridin-4-yl)-1H-pyrazol-1-yl)phenyl)urea (12)
1
8
.79 (m, 1H), 8.63 (s, 2H), 8.48 (d, 1H, J ¼ 7.6 Hz), 8.25 (d, 1H,
H NMR (300 MHz, CD
3
OD)
d
8.62 (s, 1H), 8.39 (d, 2H, J ¼ 4.5 Hz),
J ¼ 6.8 Hz), 7.94e7.89 (m, 1H), 7.58e7.51 (m, 1H), 7.44e7.41 (m, 2H),
8.08 (s, 1H), 7.95 (s, 1H), 7.56 (d, 1H, J ¼ 8.6 Hz), 7.45e7.35 (m, 6H),
13
7
(
1
.30 (t, 1H, J ¼ 8.0 Hz), 7.12 (d, 1H, J ¼ 6.8 Hz), 3.83 (s, 3H); C NMR
100 MHz, DMSO-d 162.3, 154.9, 150.4, 149.1, 140.2, 139.9, 132.6,
31.7, 130.6,130.4,124.9, 123.0,122.0,121.8, 121.7,120.8,113.4, 113.0,
7.33e7.30 (m, 2H), 7.15 (d, 1H, J ¼ 5.7 Hz), 6.98 (dd, 1H, J ¼ 1.8 Hz,
þ
6
)
d
8.0 Hz), 3.72 (s, 3H); MS m/z: 599.0 [M þ H] .
5
6.5.
4.5.5. 1-(3,5-bis(trifluoromethyl)phenyl)-3-(3-(3-(4-chloro-3-
methoxyphenyl)-4-(pyridin-4-yl)-1H-pyrazol-1-yl)phenyl)urea (13)
1
4.4. 4-[3-(4-Chloro-3-methoxyphenyl)-1-(3-aminophenyl)-1H-
6
H NMR (300 MHz, DMSO-d ) d 9.51 (brs,1H), 9.37 (brs,1H), 9.03
pyrazol-4-yl]pyridine (8)
(s, 1H), 8.59 (d, 2H, J ¼ 4.4 Hz), 8.21 (brs, 3H), 7.70e7.63 (m, 2H),
7
.56e7.51 (m, 2H), 7.42 (d, 3H, J ¼ 4.6 Hz), 7.28 (s, 1H), 7.11 (d, 1H,
1
3
A mixture of compound 7 (0.5 g, 1.2 mmol) and Pd/C (0.5 g) in
THF (5 mL) was stirred at room temperature under hydrogen at-
mosphere for 2 h. The mixture was filtered through celite and the
J ¼ 8.1 Hz), 3.82 (s, 3H); C NMR (75 MHz, DMSO-d
6
) d 154.9, 152.9,
150.3, 149.5, 142.2, 140.8, 140.3, 140.0, 133.2, 131.4, 131.0, 130.6,
129.7, 123.1, 121.8, 120.1, 118.6, 117.8, 115.2, 113.0, 109.8, 56.4; MS m/
þ
filtrate was evaporated under reduced pressure to give compound 8
z: 633.1 [M þ H] .
1
(
0.3 g, 64.8%). H NMR (400 MHz, CDCl
3
)
d
8.55 (d, 2H, J ¼ 5.6 Hz),
8
.08 (s, 1H), 7.34 (d, 1H, J ¼ 8.0 Hz), 7.27e7.22 (m, 4H), 7.17e7.14 (m,
4.6. General procedure for synthesis of diarylamide derivatives 14e18
1
3
2
H), 7.06 (d, 2H, J ¼ 8.0 Hz), 6.65e6.63 (m, 2H), 3.80 (s, 3H);
155.0, 150.1, 149.6, 147.8, 140.7, 140.5,
32.4, 130.4, 130.3, 127.4, 123.0, 122.6, 121.5, 120.0, 113.8, 112.2,
C
NMR (100 MHz, CDCl
3
)
d
A mixture of compound 8 (50 mg, 0.1 mmol), the appropriate
1
benzoic acid derivative (0.2 mmol), HOBt (36 mg, 0.3 mmol), and
ꢀ
108.8, 105.9, 56.1.
EDCI (38 mg, 0.2 mmol) in DMF (1.0 mL) was cooled to 0 C under
nitrogen atmosphere. Triethylamine (0.03 mL, 0.2 mmol) was
added thereto at the same temperature. The mixture was then
stirred at 80 C for 12 h. The reaction mixture was cooled and then
4.5. General procedure for synthesis of diarylurea derivatives 9e13
ꢀ
To a solution of compound 8 (50 mg, 0.1 mmol) in anhydrous
2
partitioned between H O and ethyl acetate. The organic layer was
THF (1 mL), a solution of the appropriate aryl isocyanate (0.1 mmol)
in THF (1 mL) was added dropwise at room temperature under N
separated and the aqueous layer was extracted with ethyl acetate
(3 ꢂ 5 mL). The combined organic layer extracts were washed with
2
.

M.I. El-Gamal et al. / European Journal of Medicinal Chemistry 65 (2013) 315e322
321
brine and dried over anhydrous Na
2
SO
4
. After evaporation of the
(silica gel, hexaneeethyl acetate 1:5 v/v) to yield the target com-
pounds 19e28.
organic solvent, the residue was purified by column chromatog-
raphy (silica gel, hexaneeethyl acetate 1:1 v/v) to yield the target
compounds 14e18.
4.7.1. 1-(3-(3-(4-Chloro-3-hydroxyphenyl)-4-(pyridin-4-yl)-1H-
pyrazol-1-yl)phenyl)-3-phenylurea (19)
1
4.6.1. N-(3-(3-(4-chloro-3-methoxyphenyl)-4-(pyridin-4-yl)-1H-
6
H NMR (400 MHz, DMSO-d ) d 10.36 (brs, 1H), 10.34 (brs, 1H),
pyrazol-1-yl)phenyl)benzamide (14)
8.90 (s, 1H), 8.72 (s, 2H), 8.13 (s, 1H), 7.65e7.59 (m, 2H), 7.48e7.40
(m, 3H), 7.33e7.24 (m, 6H), 7.22e7.19 (m, 2H), 7.04e6.94 (m, 1H);
MS m/z: 483.1 [M þ H] .
1
H NMR (300 MHz, CDCl
.25 (s, 1H), 8.06 (s, 1H), 7.81 (d, 2H, J ¼ 7.6 Hz), 7.49e7.44 (m, 3H),
.40e7.32 (m, 3H), 7.24 (d, 1H, J ¼ 8.0 Hz), 7.15 (d, 2H, J ¼ 4.7 Hz),
3
)
d
8.49 (s, 1H), 8.45 (d, 2H, J ¼ 4.6 Hz),
þ
8
7
13
7
.04 (s, 1H), 6.96 (d, 1H, J ¼ 8.1 Hz), 3.69 (s, 3H); C NMR (75 MHz,
CDCl 166.2, 155.0, 150.0, 140.5, 140.0, 139.5, 134.5, 132.2, 130.3,
30.2, 128.9, 127.5, 127.2, 123.0, 122.8, 121.5, 120.3, 118.5, 114.9,
4.7.2. 1-(3-(3-(4-Chloro-3-hydroxyphenyl)-4-(pyridin-4-yl)-1H-
3
)
d
pyrazol-1-yl)phenyl)-3-(3,4-dichlorophenyl)urea (20)
1
1
6
H NMR (400 MHz, DMSO-d ) d 10.36 (brs, 1H), 9.27 (s, 1H), 9.19
þ
112.2, 110.9, 56.1; MS m/z: 482.11 [M þ H] .
(s, 1H), 8.63 (s, 1H), 8.55 (s, 2H), 7.98 (s, 1H), 7.89 (s, 1H), 7.78 (d, 1H,
J ¼ 7.4 Hz), 7.54 (d, 1H, J ¼ 7.3 Hz), 7.45 (d, 1H, J ¼ 7.1 Hz), 7.35 (brs,
13
4
4
.6.2. 4-Bromo-N-(3-(3-(4-chloro-3-methoxyphenyl)-4-(pyridin-
5H), 7.16 (s, 1H), 6.91 (d, 1H, J ¼ 8.0 Hz); C NMR (75 MHz, DMSO-
-yl)-1H-pyrazol-1-yl) phenyl)-3-methylbenzamide (15)
6
d ) d 153.6, 152.7, 150.4, 149.2, 140.3, 140.0, 139.4, 134.2, 133.8,132.6,
1
H NMR (400 MHz, CDCl
3
)
d
8.56 (brs, 2H), 8.30 (s, 2H), 8.17 (s,
131.5, 131.1, 130.5, 124.0, 123.1, 120.7, 120.4, 120.3, 120.1, 119.1, 118.7,
þ
1
1
1
H), 7.76 (s, 1H), 7.63 (d, 1H, J ¼ 8.1 Hz), 7.58e7.53 (m, 3H), 7.47 (t,
118.2, 116.7, 110.0; MS m/z: 551.03 [M þ H] .
H, J ¼ 7.9 Hz), 7.34 (d,1H, J ¼ 8.0 Hz), 7.25 (d, 2H, J ¼ 3.8 Hz), 7.13 (s,
13
H), 7.07 (d, 1H, J ¼ 8.1 Hz), 3.79 (s, 3H), 2.46 (s, 3H); C NMR
4.7.3. 1-(3-(3-(4-Chloro-3-hydroxyphenyl)-4-(pyridin-4-yl)-1H-
(
75 MHz, CDCl
3
)
d
165.3, 162.3, 155.0, 150.2, 140.4, 140.0, 139.2,
pyrazol-1-yl)phenyl)-3-(3-(trifluoromethyl)phenyl)urea (21)
1
139.0, 133.5, 132.9, 132.2, 130.4, 129.5, 127.4, 125.7, 123.0, 121.5,
6
H NMR (400 MHz, DMSO-d ) d 10.34 (brs, 1H), 9.24e9.20 (m,
þ
120.4, 118.4, 115.0, 112.2, 110.8, 56.1, 23.1; MS m/z: 575.06 [M þ H] .
2H), 8.63 (s, 1H), 8.55e8.54 (m, 2H), 8.03e7.99 (m, 2H), 7.77 (d, 1H,
J ¼ 8.8 Hz), 7.61e7.51 (m, 4H), 7.48e7.39 (m, 4H), 7.15 (d, 1H,
13
4.6.3. N-(3-(3-(4-Chloro-3-methoxyphenyl)-4-(pyridin-4-yl)-1H-
J ¼ 1.8 Hz), 6.91 (dd, 1H, J ¼ 1.9 Hz, 8.2 Hz); C NMR (75 MHz,
pyrazol-1-yl)phenyl)-3-(trifluoromethyl)benzamide (16)
6
DMSO-d ) d 153.6, 152.8, 150.2, 149.8, 141.1, 140.7, 39.5, 134.2, 132.7,
1
H NMR (300 MHz, CDCl
3
)
d
8.57 (d, 2H, J ¼ 4.7 Hz), 8.31 (s, 1H),
132.6, 132.1, 132.0, 130.6, 123.5, 123.2, 121.0, 120.9, 120.8, 120.5,
þ
8
1
.23 (s, 1H), 8.18 (d, 2H, J ¼ 7.7 Hz), 8.10 (d, 1H, J ¼ 7.7 Hz), 7.85 (d,
119.1, 118.6, 118.2, 116.7, 109.8; MS m/z: 551.0 [M þ H] .
H, J ¼ 7.7 Hz), 7.69e7.60 (m, 4H), 7.52 (d,1H, J ¼ 8.1 Hz), 7.36 (d,1H,
J ¼ 8.1 Hz), 7.27 (d, 1H, J ¼ 1.4 Hz), 7.14 (s, 1H), 7.08 (dd, 1H,
4.7.4. 1-(4-Chloro-3-(trifluoromethyl)phenyl)-3-(3-(3-(4-chloro-3-
hydroxyphenyl)-4-(pyridin-4-yl)-1H-pyrazol-1-yl)phenyl)urea (22)
þ
J ¼ 1.7 Hz, 8.1 Hz), 3.81 (s, 3H); MS m/z: 550.1 [M þ H] .
1
H NMR (400 MHz, CD
3
OD)
d
8.56 (s, 1H), 8.37 (d, 2H, J ¼ 5.5 Hz),
4
4
.6.4. 4-Chloro-N-(3-(3-(4-chloro-3-methoxyphenyl)-4-(pyridin-
8.05 (d, 1H, J ¼ 1.9 Hz), 7.92 (d, 1H, J ¼ 2.4 Hz), 7.55 (dd, 1H,
J ¼ 2.4 Hz, 8.6 Hz), 7.45e7.40 (m, 2H), 7.35e7.32 (m, 3H), 7.26e7.23
(m, 2H), 6.99 (d, 1H, J ¼ 2.0 Hz), 6.88 (dd, 1H, J ¼ 1.9 Hz, 8.2 Hz); MS
-yl)-1H-pyrazol-1-yl)phenyl)-3-(trifluoromethyl)benzamide (17)
1
H NMR (300 MHz, CDCl
3
)
d
8.57 (d, 2H, J ¼ 4.7 Hz), 8.27e8.16
þ
(
m, 4H), 8.03 (d, 1H, J ¼ 8.1 Hz), 7.68e7.61 (m, 3H), 7.51 (t, 1H,
m/z: 585.1 [M þ H] .
J ¼ 7.8 Hz), 7.36 (d, 1H, J ¼ 8.2 Hz), 7.27e7.24 (m, 2H), 7.13 (s, 1H),
þ
7.07 (d, 1H, J ¼ 8.0 Hz), 3.81 (s, 3H); MS m/z: 584.1 [M þ H] .
4.7.5. 1-(3,5-bis(Trifluoromethyl)phenyl)-3-(3-(3-(4-chloro-3-
hydroxyphenyl)-4-(pyridin-4-yl)-1H-pyrazol-1-yl)phenyl)urea (23)
1
4.6.5. N-(3-(3-(4-Chloro-3-methoxyphenyl)-4-(pyridin-4-yl)-1H-
6
H NMR (400 MHz, DMSO-d ) d 10.34 (brs, 1H), 9.59 (s, 1H), 9.46
pyrazol-1-yl)phenyl)-4-morpholino-3-(trifluoromethyl)benzamide (18)
(s, 1H), 8.63 (s, 1H), 8.54 (d, 2H, J ¼ 6.0 Hz), 8.16 (s, 1H), 7.99 (d, 1H,
J ¼ 2.5 Hz), 7.79 (d, 1H, J ¼ 8.8 Hz), 7.68 (s, 1H), 7.52 (dd, 1H,
J ¼ 2.5 Hz, 8.8 Hz), 7.39e7.35(m, 3H), 7.15 (d, 1H, J ¼ 1.9 Hz), 6.91
1
3
H NMR (300 MHz, CDCl ) d 8.74 (brs, 1H), 8.56 (s, 2H) 8.31 (s,
1
H), 8.18 (d, 1H, J ¼ 6.8 Hz), 8.11 (s, 1H), 7.59 (d, 1H, J ¼ 7.2 Hz), 7.48
13
(
(
(
d, 1H, J ¼ 7.8 Hz), 7.35 (t, 2H, J ¼ 6.2 Hz), 7.25 (t, 2H, J ¼ 3.6 Hz), 7.18
(dd, 1H, J ¼ 1.9 Hz, 8.2 Hz); C NMR (75 MHz, DMSO-d
6
) d 153.6,
s, 1H), 7.13 (d, 1H, J ¼ 7.1 Hz), 7.07 (brs, 2H), 3.93e3.78 (m, 7H), 3.02
152.8, 150.4, 149.2, 142.0, 140.3, 140.1, 139.4, 134.2, 133.8, 132.6,
131.9, 131.4, 131.0, 130.5, 125.5, 123.1, 121.1, 120.4, 120.3, 118.8, 118.6,
1
3
t, 4H, J ¼ 2.7 Hz); C NMR (75 MHz, CDCl
3
) d 164.5, 154.9, 150.1,
þ
147.8,140.7,140.4,140.0,139.3,132.3,132.1,132.0,130.4,130.3,130.1,
116.7, 110.5; MS m/z: 618.1 [M þ H] .
1
27.4, 123.3, 123.0, 121.5, 119.9, 118.6, 115.0, 113.8, 112.2, 111.0, 108.8,
þ
105.9, 67.1, 56.1, 53.4; MS m/z: 635.17 [M þ H] .
4.7.6. N-(3-(3-(4-Chloro-3-hydroxyphenyl)-4-(pyridin-4-yl)-1H-
pyrazol-1-yl)phenyl) benzamide (24)
1
4.7. General procedure for demethylation to the target hydroxyl
6
H NMR (300 MHz, DMSO-d ) d 10.52 (s, 1H), 10.39 (s, 1H), 8.98
derivatives 19e28
(s, 1H), 8.56 (d, 2H, J ¼ 5.9 Hz), 8.50 (s, 1H), 8.01 (d, 2H, J ¼ 6.8 Hz),
7
.80 (d, 1H, J ¼ 8.1 Hz), 7.67e7.51 (m, 5H), 7.42e7.37 (m, 3H), 7.16 (d,
13
To a solution of compounds 9e18 (0.1 mmol) in methylene
chloride (1 mL), BBr
1H, J ¼ 1.8 Hz), 6.93 (dd, 1H, J ¼ 1.8 Hz, 8.2 Hz); C NMR (75 MHz,
3
(0.08 mL of a 1 M solution in methylene
DMSO-d 166.1, 153.1, 149.9, 140.4, 139.8, 139.3, 134.6, 132.3,
6
) d
ꢀ
chloride, 1.2 mmol) was added dropwise at ꢁ78 C under N
2
, and
130.0,128.4,127.7, 126.9,123.3,122.5,121.9,120.0,118.6, 114.9,111.9,
þ
the reaction mixture was stirred at the same temperature for
0 min. The mixture was allowed to warm to room temperature
and stirred for 1 h. The mixture was quenched with saturated
aqueous NaHCO . Ethyl acetate was added and the organic layer
110.7; MS m/z: 468.1 [M þ H] .
3
4.7.7. 4-Bromo-N-(3-(3-(4-chloro-3-hydroxyphenyl)-4-(pyridin-4-
3
yl)-1H-pyrazol-1-yl)phenyl)-3-methylbenzamide (25)
1
was separated. The aqueous layer was extracted with ethyl acetate.
The combined organic layer extracts were washed with brine, and
then dried over anhydrous Na
6
H NMR (400 MHz, DMSO-d ) d 10.54 (brs, 1H), 10.38 (brs, 1H),
8.99 (s,1H), 8.56 (brs, 2H), 8.47 (s,1H), 8.00 (s,1H), 7.82e7.77 (m, 3H),
7.68 (d,1H, J ¼ 7.2 Hz), 7.55 (t,1H, J ¼ 8.0 Hz), 7.42e7.37 (m, 3H), 7.16
2 4
SO . After evaporation of the organic
þ
solvent, the residue was purified by short column chromatography
(s,1H), 6.94 (d,1H, J ¼ 7.8 Hz), 2.47 (s, 3H); MS m/z: 561.05 [M þ H] .

322
M.I. El-Gamal et al. / European Journal of Medicinal Chemistry 65 (2013) 315e322
4
.7.8. N-(3-(3-(4-Chloro-3-hydroxyphenyl)-4-(pyridin-4-yl)-1H-
Using the seven absorbance measurements [time zero, (Tz), control
growth, (C), and test growth in the presence of drug at the five
concentration levels (Ti)], the percentage growth is calculated at
each of the drug concentrations levels. Percentage growth inhibi-
tion is calculated as:
pyrazol-1-yl)phenyl)-3-(trifluoromethyl)benzamide (26)
1
H NMR (400 MHz, DMSO-d
6
)
d
8.72 (d, 2H, J ¼ 4.7 Hz), 8.34e
8
1
.30 (m, 2H), 8.19 (d, 2H, J ¼ 7.8 Hz), 8.14 (d, 1H, J ¼ 7.9 Hz), 7.89 (d,
H, J ¼ 7.7 Hz), 7.68e7.61 (m, 4H), 7.58 (d,1H, J ¼ 8.0 Hz), 7.37 (d,1H,
J ¼ 8.1 Hz), 7.36 (d, 1H, J ¼ 7.9 Hz), 7.14 (s, 1H), 7.09 (dd, 1H,
þ
J ¼ 1.7 Hz, 8.1 Hz); MS m/z: 536.1 [M þ H] .
ꢃ [(Ti ꢁ Tz)/(C ꢁ Tz)] ꢂ 100 for concentrations for which Ti ꢄ Tz
ꢃ [(Ti ꢁ Tz)/Tz] ꢂ 100 for concentrations for which Ti < Tz.
4.7.9. 4-Chloro-N-(3-(3-(4-chloro-3-hydroxyphenyl)-4-(pyridin-4-
yl)-1H-pyrazol-1-yl)phenyl)-3-(trifluoromethyl)benzamide (27)
Growth inhibition of 50% (IC50) is calculated from [(Ti ꢁ Tz)/
(C ꢁ Tz)] ꢂ 100 ¼ 50, which is the drug concentration resulting in a
50% reduction in the net protein increase (as measured by SRB
staining) in control cells during the drug incubation.
1
H NMR (400 MHz, DMSO-d
.55e8.54 (m, 2H), 8.44 (d, 2H, J ¼ 7.3 Hz), 8.31 (d, 1H, J ¼ 7.9 Hz),
.95 (d,1H, J ¼ 8.5 Hz), 7.82 (d,1H, J ¼ 7.9 Hz), 7.69 (d,1H, J ¼ 7.4 Hz),
6
) d 10.78 (brs, 1H), 8.99 (s, 1H),
8
7
7
.56 (t, 1H, J ¼ 8.1 Hz), 7.39e7.36 (m, 4H), 7.13 (s, 1H), 6.86 (d, 1H,
þ
J ¼ 7.8 Hz); MS m/z: 570.1 [M þ H] .
Acknowledgments
4
.7.10. N-(3-(3-(4-Chloro-3-hydroxyphenyl)-4-(pyridin-4-yl)-1H-
pyrazol-1-yl)phenyl)-4-morpholino-3-(trifluoromethyl)benzamide
28)
1H NMR (300 MHz, CDCl
) d 9.73 (brs, 1H), 8.34 (d, 2H,
This work was supported by Korea Institute of Science and
Technology (KIST), KIST Project (2E22360). We would like to thank
the National Cancer Institute (NCI), Bethesda, Maryland, USA, for
performing the anticancer testing of the target compounds over 60
cancer cell lines.
(
3
J ¼ 4.8 Hz), 8.27 (s, 1H), 8.15 (d, 3H, J ¼ 6.9 Hz), 8.06 (d, 1H,
J ¼ 4.0 Hz), 7.59 (t, 2H, J ¼ 6.9 Hz), 7.49 (d, 1H, J ¼ 7.7 Hz), 7.43e7.37
(
(
m, 3H), 7.29 (s, 1H), 7.10e7.03 (m, 2H), 3.86 (t, 4H, J ¼ 4.3 Hz), 3.03
þ
Appendix A. Supplementary data
t, 4H, J ¼ 3.9 Hz); MS m/z: 620.16 [M þ H] .
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2013.04.067.
4.8. 60 Cancer cell line screening at the NCI
Screening against a panel of 60 cancer cell lines was applied at
the National Cancer Institute (NCI), Bethesda, Maryland, USA [24],
applying the following procedure. The human cell lines are grown
in RPMI 1640 medium containing 5% fetal bovine serum and 2 mM
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