Synthesis and biological evaluation of novel (Thio)urea derivatives as potential antitumor agents

Article abstract of DOI:10.1002/ardp.201000267

A series of novel (thio)ureas containing the pyrimidinyl group was designed and synthesized. Their in-vitro antitumor activity against different human tumor cells was examined. Some of the compounds showed potential antitumor activity, which provided some

Full text of DOI:10.1002/ardp.201000267

Arch. Pharm. Chem. Life Sci. 2011, 344, 741–744
741
Full Paper
Synthesis and Biological Evaluation of Novel (Thio)Urea
Derivatives as Potential Antitumor Agents
Chuanfei Jin1, Yong-Ju Liang2, Hongwu He1, and Liwu Fu2
1 Key Laboratory of Pesticide & Chemical Biology, Ministry of Education, College of Chemistry,
Central China Normal University, Wuhan, P. R. China
2 State Key Laboratory of Oncology in South China, Cancer Center, Sun Yat-sen University, Guangzhou,
P. R. China
A series of novel (thio)ureas containing the pyrimidinyl group was designed and synthesized. Their
in-vitro antitumor activity against different human tumor cells was examined. Some of the compounds
showed potential antitumor activity, which provided some hints for further studies on structure
modification.
Keywords: Antitumor activity / Pyrimidinyl group / Thiourea / Urea
Received: September 15, 2010; Revised: December 19, 2010; Accepted: December 28, 2010
DOI 10.1002/ardp.201000267
Introduction
and synthesized. The preliminary in-vitro cytotoxic activity of
the new compounds was evaluated.
The urea derivatives such as ureas [1, 2], benzoylureas [3, 4],
thioureas [5–7], and diarylsulfonylureas [8, 9] represent one of Results and discussion
the generally most useful classes of anticancer agents (Fig. 1),
with a wide range of activities against various leukemia and
solid tumors. For these reasons, the synthesis of ureas and
their functionalized derivatives is a primary objective.
In recent years, many reports have been published on
topics about the synthetic and mechanistic aspects of poten-
tial anticancer drugs of the urea derivatives. For example, Dai
[10] reported a series of thienopyrimidine urea derivatives for
receptor tyrosine kinase inhibitors based on a thienopyrimi-
dine scaffold. The antitumor activity studies focused on
optimizing activity against KDR (kinase insert domain-
containing receptor tyrosine kinase), as this kinase plays
an important role in tumor angiogenesis.
Chemistry
Compound 3 could be synthesized starting from 4-amino-
phenol 1 and 4,6-dimethoxy-2-(methylsulfonyl)pyrimidine or
4,6-dichloro-2-(methylsulfonyl) pyrimidine 2 in the presence
of K2CO3 in DMF. Treatment of equimolar amounts of sub-
stituted isocyanates or isothiocyanates with compound 3 in
dry CH2Cl2 afforded the corresponding (thio)urea derivatives
(4a–4l) (Scheme 1). The preparations are summarized
in Table 1. The new compounds were fully characterized
by spectroscopic means and their purity established by
elemental analysis.
The pyrimidinyl group is a highly efficient pharmacophore
and is widely used in pesticide [11] and drug [12] molecular
design. We wanted to investigate whether there would be
some new beneficial properties if the pyrimidinyl group was
introduced in the urea derivatives. Herein, a series of novel
(thio)ureas containing the pyrimidinyl group were designed
Correspondence: Hongwu He, College of Chemistry, Central China
Normal University, 152 Luoyu Road, Wuhan, P. R. China.
E-mail: he1208@yahoo.com.cn
Fax: þ86 (0)27 67867960
Figure 1. Structures of the uera derivatives.
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742
C. Jin et al.
Arch. Pharm. Chem. Life Sci. 2011, 344, 741–744
Cytotoxic activity
In-vitro cytotoxicity of the synthesized compounds was
screened on different human tumor cell lines using MTT
assay. 5-Fluorouracil was used as the reference drug. The
results of the cytotoxicity studies are shown in Table 2.
The results indicated that the pyrimidinyl group intro-
duced into the (thio)urea structure was useful for the
improvement of antitumor activity. For the compound with-
out pyrimidinyl group displayed very weak antitumor
activity against the tumor cells. As could be seen from
Table 2, most of the title compounds exhibited moderate
cytotoxic activity against the tested tumor cells. Generally,
most of the title compounds tended to be more active against
KB than against other tested tumor cells. The thiourea deriva-
tives displayed higher activity than urea derivatives.
Scheme 1. General synthetic route for compounds 4a–l.
Table 1. The preparation of 1-(4-(4,6-di-substituted pyrimidin-2-
yloxy)phenyl)-3-arylurea derivatives 4a–l.
Compound
R1
X
R2
Yield (%)a
Mp (8C)
The compound 4k displayed more potent cytotoxic
activities against KB, MGC-803 and MCF-7 in comparison with
5-FU. The compound 4l displayed similar potent cytotoxic
activities against CNE2 in comparison with 5-FU. The
compound 4f displayed similar potent cytotoxic activities
against KB and MCF-7 in comparison with 5-FU. The com-
pounds 4k showed the best inhibitory activity against
KB MGC-803 and MCF-7 with IC50 9.36, 8.80 and 13.0 mM,
respectively. The compound 4l showed the best inhibitory
activity against CNE2 with IC50 6.63 mM. We noticed that the
structure of the substituent R1 had great influence on the
4a
4b
4c
4d
4e
4f
4g
4h
4i
OMe
OMe
OMe
OMe
Cl
Cl
Cl
Cl
O
O
O
O
O
O
O
O
S
H
3-Me
4-OCF3
4-F
76
84
84
88
82
85
67
86
78
88
80
82
211.0ꢀ212.3
193.6ꢀ194.5
248.8ꢀ249.7
232.2ꢀ233.1
240.1ꢀ241.5
226.5ꢀ227.8
251.8ꢀ252.6
255.5ꢀ257.6
65.3ꢀ66.7
H
3-Me
4-OCF3
4-F
OMe
OMe
Cl
H
4j
4k
4l
S
S
S
2,4-diCl
H
2,4-diCl
79.1ꢀ81.3
156.8ꢀ158.4
179.4ꢀ181.7
Cl
a
Isolated yields
Table 2. Cytotoxic activity of urea compounds against human tumor cells.
Compound
In-vitro cytotoxicity IC50 (mM)a
KBa
CNE2a
MGC-803a
MCF-7a
4a
4b
4c
4d
4e
4f
4g
4h
4i
35.7
>50
>50
18.9
13.5
12.2
14.8
20.9
>50
24.7
9.36
14.4
>50
>50
>50
34.4
>50
>50
23.6
>50
17.4
15.2
>50
>50
>50
8.80
24.0
41.7
>50
>50
43.9
>50
14.8
15.3
>50
>50
>50
13.0
46.5
b
–
14.8
13.3
15.7
27.9
>50
26.6
47.4
6.63
4j
4k
4l
O
HN
>50
>50
–
>50
HN
OMe
5-Fluorouracil
12.9
13.1
12.6
15.2
a
KB cells were the drug sensitive human oral carcinoma cells, CNE2 cells were the drug sensitive human nasopharyngeal carcinoma
cells, MGC-803 cells were the drug sensitive human gastric carcinoma cells and MCF-7 cells were the drug sensitive human breast
b
adenocarcinoma cells. –, not tested.
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Arch. Pharm. Chem. Life Sci. 2011, 344, 741–744
Synthesis and Biological Evaluation of Novel (Thio)Ureas
743
isocyanate (1 mmol) was added dropwise with constant stirring.
After 1 h, formation of white solid occurred and the resultant
solid product was filtered. Recrystallization with ethanol
afforded the desired solid urea.
antitumor activity. For some compounds with OMe group
(4b, 4c and 4i) displayed very weak antitumor activity against
the tumor cells, whereas some compounds with Cl group (4f,
4g and 4k) displayed potential antitumor activity against the
tumor cells. Still, the change of R2 group also had a little
influence on their activity.
1-(4-(4,6-Dimethoxypyrimidin-2-yloxy)phenyl)-3-
phenylurea 4a
White solid; 1H-NMR (DMSO-d6, 600 MHz): 3.78 (s, 6H, OCH3), 5.98
(s, 1H, CH), 6.97 (m, 1H, ArH), 7.14 (d, 2H, J ¼ 7.8 Hz, ArH),
7.27ꢀ7.30 (m, 2H, ArH), 7.45ꢀ7.49 (m, 4H, ArH), 8.67 (s, 1H,
NH), 8.72 (s, 1H, NH); 13C-NMR (DMSO-d6, 125 MHz): 54.2, 83.7,
118.2, 118.9, 121.9, 122.1, 128.8, 136.8, 139.7, 146.9, 152.6, 163.0,
172.7; Anal. calcd. for C19H18N4O4 (366.4): C, 62.29; H, 4.95; N,
15.29. Found: C, 62.01; H, 4.75; N, 15.26.
Conclusion
In conclusion, a series of novel (thio)urea derivatives was
designed and synthesized. The in-vitro preliminary bioassay
data showed that some of the synthesized compounds have
potential antitumor activities. The compound 4k was found
to have more potent cytotoxic activities against some human
tumor cells than 5-FU. Further investigations on structural
optimization and biological activities about these derivatives
are still underway in our laboratory.
1-(4-(4,6-Dimethoxypyrimidin-2-yloxy)phenyl)-3-
m-tolylurea 4b
White solid; 1H-NMR (DMSO-d6, 600 MHz): 2.27 (s, 3H, CH3), 3.78
(s, 6H, OCH3), 5.98 (s, 1H, CH), 6.79 (d, 1H, J ¼ 7.2 Hz, ArH),
7.13ꢀ7.29 (m, 5H, ArH), 7.47 (d, 2H, J ¼ 9.0 Hz, ArH), 8.60
(s, 1H, NH), 8.71 (s, 1H, NH); 13C-NMR (DMSO-d6, 125 MHz):
20.4, 54.2, 83.8, 115.4, 118.7, 118.9, 121.9, 122.0, 128.6, 136.8,
138.0, 139.6, 146.9, 152.5, 164.0, 172.7; Anal. calcd. for
C20H20N4O4 (380.4): C, 63.15; H, 5.30; N, 14.73. Found: C,
63.38; H, 5.02; N, 14.35.
Experimental
General
1H-NMR spectra were recorded at 600 MHz, in CDCl3 or DMSO-d6
solution on a Varian Mercury-Plus 600 spectrometer and chemi-
cal shifts were recorded in parts per million (ppm) with TMS as
the internal reference. Melting points were taken on a Buechi
B-545 melting point apparatus. Element analysis (EA) was
measured on a Vario ELIII CHNSO elemental analyzer. All com-
mercially available solvents and reagents were used as supplied
by Acros Organics unless otherwise stated.
1-(4-(4,6-Dimethoxypyrimidin-2-yloxy)phenyl)-3-
(4-(trifluoromethoxy)phenyl)urea 4c
White solid; 1H-NMR (DMSO-d6, 600 MHz): 3.75 (s, 6H, OCH3), 5.95
(s, H, CH), 7.12 (d, 2H, J ¼ 8.4 Hz, ArH), 7.26 (d, 2H, J ¼ 8.4 Hz,
ArH), 7.46 (d, 2H, J ¼ 9.6 Hz, ArH), 7.53 (d, 2H, J ¼ 8.4 Hz, ArH),
8.75 (s, 1H, NH), 8.86 (s, 1H, NH); 13C-NMR (DMSO-d6, 125 MHz):
54.2, 83.8, 119.1, 119.4, 121.8, 121.9, 136.6, 139.1, 142.6, 147.1,
152.6, 164.0, 172.7; Anal. calcd. for C20H17F3N4O5 (450.4): C,
53.34; H, 3.80; N, 12.44. Found: C, 52.91; H, 3.46; N, 12.02.
Chemistry
General procedure for the synthesis of compounds 3a,3b
A solution of 4,6-dimethoxy-2-(methylsulfonyl)pyrimidine or
4,6-dichloro-2-(methylsulfonyl)
(18 mmol),
4-aminophenol
1-(4-(4,6-Dimethoxypyrimidin-2-yloxy)phenyl)-3-
(18 mmol) and K2CO3 (36 mmol) in DMF (50 mL) was stirred at
1008C for 2 h. The reaction mixture was poured into water and
extracted with EtOAc. The organic layer was washed with water
and saturated brine, dried over Na2SO4 and evaporated. The
residue was purified by column chromatography on silica gel
(PE/EA, 2:1) to give the compound 3a,3b.
(4-fluorophenyl)urea 4d
White solid; 1H-NMR (DMSO-d6, 600 MHz): 3.78 (s, 6H, OCH3), 5.98
(s, 1H, CH), 7.12ꢀ7.14 (m, 3H, ArH), 7.46ꢀ7.48 (m, 4H, ArH), 8.71
(s, 1H, NH), 8.72 (s, 1H, NH); 13C-NMR (DMSO-d6, 125 MHz): 54.2,
83.8, 115.4, 119.0, 119.9, 121.9, 136.0, 136.8, 146.9, 152.6, 158.6,
164.0, 172.7; Anal. calcd. for C19H17FN4O4 (384.4): C, 59.37; H,
4.46; N, 14.58. Found: C, 59.25; H, 4.30; N, 14.73.
4-(4,6-Dimethoxypyrimidin-2-yloxy)benzenamine 3a
Yellow solid; yield 85%; mp 84.6ꢀ85.48C; 1H-NMR (CDCl3,
600 MHz): 3.64 (s, 2H, NH2), 3.84 (s, 6H, OMe), 5.75 (s, H, CH),
6.68 (d, 2H, J ¼ 12 Hz, ArH), 7.00 (d, 2H, J ¼ 12 Hz, ArH). MS (EI):
m/z 247 (Mþ, 100), 216 (36), 108 (34), 80 (31).
1-(4-(4,6-Dichloropyrimidin-2-yloxy)phenyl)-3-
phenylurea 4e
White solid; 1H-NMR (DMSO-d6, 600 MHz): 6.95ꢀ6.97 (m, 1H,
ArH), 7.16ꢀ7.28 (m, 4H, ArH), 7.44 (d, 2H, J ¼ 7.8 Hz, ArH),
7.50 (d, 2H, J ¼ 9.0 Hz, ArH), 7.74 (s, 1H, CH), 8.70 (s, 1H, NH),
8.76 (s, 1H, NH); 13C-NMR (DMSO-d6, 125 MHz): 115.9, 118.2,
119.3, 119.4, 121.8, 128.8, 137.6, 139.7, 146.4, 152.6, 162.6,
163.8; Anal. calcd. for C17H12Cl2N4O2 (375.2): C 54.42, H 3.22,
N 14.93. Found: C 54.42, H 3.26, N 14.68.
4-(4,6-Dichloropyrimidin-2-yloxy)benzenamine 3b
White solid; yield 45%; mp 258.9ꢀ260.28C; 1H-NMR (CDCl3,
600 MHz): 3.68 (s, 2H, NH2), 6.72 (d, 2H, J ¼ 12 Hz, ArH), 6.98
(d, 2H, J ¼ 12 Hz, ArH), 7.10 (s, H, CH). MS (EI): m/z 259 (Mþ4, 3),
257 (Mþ2, 10), 255 (Mþ, 28), 220 (100), 157 (19), 108 (24), 92 (30),
65 (41).
1-(4-(4,6-Dichloropyrimidin-2-yloxy)phenyl)-3-
m-tolylurea 4f
General procedure for the synthesis of compounds 4a–4l
Compound 3 (1 mmol) was dissolved in 15 mL dichloromethane.
Temperature was maintained at 08C, then substituted phenyl
1
White solid; H-NMR (DMSO-d6, 600 MHz): 2.28 (s, 3H, Me), 6.79
(d, 2H, J ¼ 7.2 Hz, ArH), 7.14ꢀ7.30 (m, 4H, ArH), 7.51 (d, 2H,
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744
C. Jin et al.
Arch. Pharm. Chem. Life Sci. 2011, 344, 741–744
J ¼ 9.0 Hz, ArH), 7.76 (s,1H, CH), 8.63 (s, 1H, NH), 8.76 (s, 1H, NH);
13C-NMR (DMSO-d6, 125 MHz): 21.2, 115.4, 116.0, 118.7, 119.3,
121.7, 122.6, 128.7, 137.6, 138.0, 139.5, 146.3, 152.5, 162.5, 163.8;
Anal. calcd. for C18H14Cl2N4O2 (389.2): C 55.54, H 3.63, N 14.39.
Found: C 55.18, H 3.31 N 14.12.
Pharmacology
The in-vitro cytotoxicity measurement of the synthesized com-
pounds against different cancer cell lines was performed with
the MTT assay according to the Mosmann’s method [13]. The MTT
assay is based on the reduction of the soluble 3-(4,5-dimethyl-2-
thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) into a blue-
purple formazan product, mainly by mitochondrial reductase
activity inside living cells. The cells used in the cytotoxicity assay
were cultured in RPMI 1640 medium supplemented with 10%
fetal calf serum. Cells suspended in the medium (2Y0 104/mL) were
plated in 96-well culture plates and incubated at 378C in a 5% CO2
incubator. After 12 h, the test sample (2 mL) was added to the
cells (2Y0 104) in 96-well plates and cultured at 378C for 3 days. The
cultured cells were mixed with 20 mL of MTT solution and
incubated for 4 h at 378C. The supernatant was carefully removed
from each well and 100 mL of DMSO was added to each well to
dissolve the formazan crystals which were formed by the cellular
reduction of MTT. After mixing with a mechanical plate mixer,
the absorbance of each well was measured by a microplate reader
using a test wavelength of 570 nm. The results were expressed as
the IC50, which is the concentration of the drugs inducing a 50%
inhibition of cell growth of treated cells when compared to the
growth of control cells. Each experiment was performed at least
3 times. There was a good reproducibility between replicate wells
with standard errors below 10%.
1-(4-(4,6-Dichloropyrimidin-2-yloxy)phenyl)-3-
(4-(trifluoromethoxy)phenyl)urea 4g
White solid; 1H-NMR (DMSO-d6, 600 MHz): 7.17 (d, 2H, J ¼ 8.4 Hz,
ArH), 7.27 (d, 2H, J ¼ 8.4 Hz, ArH), 7.49 (d, 2H, J ¼ 8.4 Hz, ArH),
7.54 (d, 2H, J ¼ 8.4 Hz, ArH), 7.73 (s,1H, CH), 8.81 (s, 1H, NH), 8.90
(s, 1H, NH); 13C-NMR (DMSO-d6, 125 MHz): 116.0, 119.4, 119.5,
120.6, 121.1, 121.8, 137.4, 139.0, 142.6, 146.5, 152.6, 162.6, 163.8;
Anal. calcd. for C18H11Cl2F3N4O3 (459.2): C 47.08, H 2.41, N 12.20.
Found: C 47.09, H 2.33, N 12.43.
1-(4-(4,6-Dichloropyrimidin-2-yloxy)phenyl)-3-
(4-fluorophenyl)urea 4h
White solid; 1H-NMR (DMSO-d6, 600 MHz): 7.10 (d, 2H, J ¼ 8.4 Hz,
ArH), 7.17 (d, 2H, J ¼ 8.4 Hz, ArH), 7.45 (d, 2H, J ¼ 7.4 Hz, ArH),
7.49 (d, 2H, J ¼ 8.4 Hz, ArH), 7.74 (s, 1H, CH), 8.74 (s, 1H, NH), 8.76
(s, 1H, NH); 13C-NMR (DMSO-d6, 125 MHz): 115.2, 116.0, 119.5,
121.7, 136.0, 137.6, 146.4, 152.7, 156.6, 158.2, 162.6, 163.8; Anal.
calcd. for C17H11Cl2F N4O2 (393.2): C 51.93, H 2.82, N 14.25.
Found: C 51.91, H 2.66, N 13.95.
This work was supported by the National Natural Science Foundation of
China (20772042).
1-(4-(4,6-Dimethoxypyrimidin-2-yloxy)phenyl)-3-
phenylthiourea 4i
White solid; 1H-NMR (DMSO-d6, 600 MHz): 3.78 (s, 6H, OCH3), 5.99
(s, H, CH), 7.12 (s, 1H, ArH), 7.18 (d, 2H, J ¼ 7.2 Hz, ArH),
7.31ꢀ7.33 (m, 2H, ArH), 7.46 (d, 2H, J ¼ 7.2 Hz, ArH), 7.50
(d, 2H, J ¼ 7.2 Hz, ArH), 9.80 (s, H, NH), 9.82 (s, H, NH); Anal.
calcd. for C19H18N4O3S (382.4): C 59.67, H 4.74, N 14.65. Found: C
59.65, H 4.62, N 14.65.
The authors have declared no conflict of interest.
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White solid; 1H-NMR (DMSO-d6, 600 MHz): 3.77 (s, 6H, OCH3, 5.99
(s, H, CH), 7.20 (d, 2H, J ¼ 8.4 Hz, ArH), 7.42 (d, 1H, J ¼ 9.0 Hz,
ArH), 7.52 (d, 2H, J ¼ 7.8 Hz, ArH), 7.60 (d, 1H, J ¼ 9.0 Hz, ArH),
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phenylthiourea 4k
White solid; 1H-NMR (DMSO-d6, 600 MHz): 7.11 (d, 1H, J ¼ 7.8 Hz,
ArH), 7.21 (d, 2H, J ¼ 9.0 Hz, ArH), 7.30ꢀ7.33 (m, 2H, ArH), 7.45
(d, 2H, J ¼ 7.2 Hz, ArH), 7.21 (d, 2H, J ¼ 9.0 Hz, ArH), 7.76 (s, 1H,
ArH), 9.83 (s, 1H, NH), 9.88 (s, 1H, NH); Anal. calcd.
for C17H12Cl2N4OS (391.3): C 52.18, H 3.09, N 14.32. Found: C
52.22, H 3.01, N 14.65.
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1-(2,4-Dichlorophenyl)-3-(4-(4,6-dichloropyrimidin-2-
yloxy)phenyl)thiourea 4l
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Chem. 1997, 45, 2777–2783.
White solid; 1H-NMR (DMSO-d6, 600 MHz): 7.27 (d, 2H, J ¼ 8.4 Hz,
ArH), 7.44 (d, 1H, J ¼ 9.0 Hz, ArH), 7.67ꢀ7.62 (m, 3H, ArH), 7.70
(s, 1H, CH), 7.79 (s, 1H, ArH), 9.58 (s, H, NH), 10.09 (s, H, NH); Anal.
calcd. for C17H10Cl4N4OS (460.2): C 44.37, H 2.19, N 12.18. Found:
C 44.70, H 2.56, N 12.12.
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