urea anticancer quinolina

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ORIGINAL RESEARCH
Synthesis and evaluation in vitro of 1-[2-(10-
dihydroartemisininoxy) ethyl]-3-phenylurea derivatives
as potential agents against cancer
Wei Luo • Ming-yu Xia • Takashi Ikejima •
Li-hua Li • Chun Guo
Received: 4 July 2012 / Accepted: 25 October 2012 / Published online: 11 November 2012
� Springer Science+Business Media New York 2012
Abstract In order to develop potent and selective anti-
cancer agents, a series of novel artemisinin derivatives
bearing urea moiety 1a–n were facilely synthesized herein
and screened for their activities in vitro against ten human
tumor cell lines (HeLa, MCF-7, U937, K562, HL60,
HCT116, HepG2, A549, A375-S2, and HT1080). The
pharmacological results indicated that some compounds
showed excellent activity against cancer cell lines and good
selectivity, especially the compound 1c which proved to be
the most active against the cancer cells as well as dis-
tinctive patterns of selectivity.
Keywords Artemisinin derivatives � Anticancer �
Urea � Cytotoxicity
Introduction
Up to now, cancer has been the second most fatal disease
after cardiovascular disorders in the world. It is estimated
that about 7.6 million people die owing to this disease
every year, and the death may increase up to 9.0 millions
and 11.0 millions in 2015 and in 2030, respectively. Cancer
is a disease in which the control of growth is lost in one or
more cells, leading either to a solid mass of cells known as
a tumor or to a liquid cancer (Ali et al., 2011; Nussbaumer
et al., 2011). Chemotherapy has become the most impor-
tant strategy against all kinds of cancer effectively; how-
ever, the available anticancer drugs are often impaired by
the undesirable side effects and emergence of resistance
(Bondock et al., 2012; Eckford and Sharom, 2009).
Therefore, searching for novel chemotherapeutic agents
with higher selectivity and more potent anticancer activi-
ties is an imperative need.
Artemisinin, obtained from Artemisia annua, plays a
critical role successfully against resistant malarial parasites
without obvious adverse effects (Li and Wu, 2010). It not
only has well-known anti-malarial activity, but also
showed potent and broad anti-cancer properties in various
human cancer cell lines. Despite its obvious value, the
chemotherapeutic value of artemisinin is limited to a great
extent by its poor solubility either in oil or water and short
plasma half-life. In order to conquer these shortcomings,
search for novel artemisinin derivatives with more potent
anticancer activities was initiated in the world. Fortunately,
a number of artemisinin analogs have been synthesized and
showed excellent anticancer activities (Chaturvedi et al.,
2010). At the same time, urea derivatives are synthesized
largely in recent years, and it is important to note that most
of urea derivatives exhibited VEGFR kinase inhibitory
activity and anticancer profile against different human
tumor-derived cell lines (Fig. 1), followed by the suc-
cessful launching of Sorafenib, Linifanib, and Tivozanib
currently being evaluated in phase II or III clinical trials (Li
et al., 2009; Pandey et al., 2011; Fortin et al., 2010;
Asakawa et al., 2011; Luo et al., 2012a; Eskens et al.,
2011). Hence it can be reasonably speculated that urea
moiety might be an effective pharmacological moiety in
W. Luo � C. Guo (&)
Key Laboratory of Structure-Based Drug Design & Discovery,
Shenyang Pharmaceutical University, Ministry of Education,
Shenyang 110016, Liaoning, China
e-mail: chunguo63@yahoo.com.cn
W. Luo
e-mail: l198278w@yahoo.com.cn
M. Xia � T. Ikejima � L. Li
China-Japan Research Institute of Medical and Pharmaceutical
Sciences, Shenyang Pharmaceutical University,
Shenyang 110016, Liaoning, China
123
Med Chem Res (2013) 22:3170–3176
DOI 10.1007/s00044-012-0319-0
MEDICINAL
CHEMISTRY
RESEARCH
anticancer agents. On the basis of the above consideration,
we have designed a new series of artemisinin derivatives
1a–n with the urea moiety attached to C-10 position of
artemisinin affording the new hybrid scaffold which would
be effective against tumor. All the prepared compounds
were screened for their anticancer activities against HeLa,
MCF-7, U937, K562, HL60, HCT116, HepG2, A549,
A375-S2, and HT1080 cell lines in vitro. The details of
synthesis of target compounds and evaluation of their
activities against cancer cell lines were presented herein.
Results and discussion
Chemistry
The synthesis of the target compounds was carried out
according to the procedure given in Scheme 1. Compounds
3a–n could be prepared by the reaction of corresponding
aniline with phenyl chloroformate in dichloromethane, and
substitution reaction of 3a–n in refluxing ethanol with
2-aminoethanol afforded the desired aromatic urea com-
pounds 4a–n. Artemisinin 5 was reduced to dihydroarte-
misinin (DHA) 6 using sodium borohydride in methanol ,
which was treated with compounds 4a–n at room temper-
ature to provide desired target compounds 1a–n. The
configuration at C-10 position of artemisinin derivatives
was indicated by the coupling constant between 9-H and
10-H in 1H NMR (the a-isomers, J = 9–10 Hz and the
b-isomers, J = 3–4 Hz) (Lee et al., 2000; Luo et al., 2012b).
Pharmacology and discussion
The prepared compounds 1a–n were tested against HeLa,
MCF-7, U937, K562, HL60, HCT116, HepG2, A549, A375-
S2, and HT1080 cell lines in vitro according to the MTT-
based assay. A preliminary assay is shown in Tables 1 and 2,
respectively, revealing that most synthetic derivatives of
artemisinin possessed potential cytotoxicity as well as the
selectivity for some particular cancer cell types.
As seen from Table 1, all the target compounds displayed
a significant activity against HL60 cell (IC50 \ 1 lM).
Meanwhile, with the exception of compounds 1b and 1h
(IC50 = 23.73 lM and IC50 = 25.24 lM, respectively), all
the compounds showed remarkable cytotoxicity against
U937 cell, especially compounds 1j and 1m (IC50 \ 1 lM,
respectively). Compound 1c (IC50 = 11.60 lM) appeared
to exhibit considerably higher activity against MCF-7 cell,
being fourfold more active than DHA (IC50 = 50.18 lM);
however, compound 1j turned out to be less active in the
same assay (IC50 [ 100 lM). This may be due to the reason
that the configuration at C-10 position of artemisinin might
linifanib N
O Cl
H
N
O
H
N
MeO
MeO
O
N
tivozanib
N
H
N
H
O
Cl
F3C
Cl
H
N
H
N
O
O
H
N
Osorafenib
N
HN
H
N
O
H
N
F
NH2
HO
5
O
O
O
O
O
H
H
R1=H, F, Cl, Br, Me, OMe, CF3, NO2.
R2=H, Cl, CH3.
1a-nN
H
N
H
O
R1
R2
N
NO2
NO
N O
N
HN
N N
H
N
H
O
NH2
Fig. 1 The structures of
anticancer agents and target
compounds 1a–n
Med Chem Res (2013) 22:3170–3176 3171
123
have important impact on their antitumor activity against
MCF-7 cell. In addition, as seen from Table 2, all the tested
compounds showed antitumor activities against HCT116
and A549 cells with the IC50 values of 22.67–53.78 and
20.92–63.34 lM, respectively, which were 1.7- to 4-fold and
1.2- to 3.8-fold more active than DHA (IC50 = 95.41 and
76.21 lM, respectively), suggesting that the urea moiety
introduced to artemisinin was useful for the improvement of
antitumor activities against HCT116 and A549 cells. It was
noteworthy that compounds 1b and 1j exhibited the best
potential cytotoxicity against HepG2 (IC50 = 11.21 and
11.66 lM, respectively). For A375-S2 and HT1080 cells, it
was very interesting that the compounds 1h, 1i, 1k, 1m, and
1n showed moderate cytotoxicity; however, most target
compounds displayed very weak antitumoractivity
(IC50 [ 100 lM). Furthermore, with the exception of com-
pounds 1c and 1g exhibiting antitumor activity against K562
(IC50 = 6.97 and 9.45 lM, respectively), all tested com-
pounds turned out to be less active than DHA against the
HeLa and K562 cells in the same assay, and the results
indicated that the introduction of urea structure was not
conducive to the improvement of the activities against
A375-S2, HT1080, HeLa, and K562, but could enhance the
selectivity for cell lines. Overall, it is interesting to note that
O
O
O
O
O
H
H
O
O
O
O
OH
H
H
O
O
O
O
O
H
H
R1=H, F, Cl, Br, Me, OMe, CF3, NO2.
R2=H, Cl, CH3.
NH2
R
N
H
R
O
OPh N
H
R
O
N
H
a b
c d
n-a4n-a3n-a2
5 6
1a-n
OH
N
H
N
H
O
R1
R21'
2'
3'
4'
5'
6'
3
4
1
2 7
9
8
10
5 6
11
Scheme 1 Synthesis of target
compounds 1a–n. Reagents and
conditions (a) phenyl
chloroformate, CH2Cl2, 0–5 �C,
2 h; (b) 2-aminoethanol, EtOH,
reflux, 7 h; (c) NaBH4, MeOH,
0–5 �C, 5 h; (d) Compound
4a–n, CH2Cl2, BF3�Et2O, rt, 2 h
Table 1 Inhibitory effects of compounds on the growth of HeLa, MCF-7, U937, K562, and HL60 cell lines in vitro
Compd. R1 R2 Configuration at C-10 IC50 (lM)
HeLa MCF-7 U937 K562 HL60
1a 40-NO2 H a 64.24 94.74 9.26 [100 \1.00
1b 40-MeO H b 61.47 [100 23.73 55.79 2.53
1c 40-CF3 H b 16.65 11.60 1.10 6.97 \1.00
1d 20-Me H b 64.69 88.90 16.21 29.44 2.11
1e H H b 25.13 43.68 6.06 24.97 3.00
1f 20-Me 50-Me b 31.01 42.2 2.51 22.96 \1.00
1g 20-Me 30-Cl b 25.65 36.84 2.59 9.45 \1.00
1h 40-F H b 35.17 46.76 25.24 38.36 3.64
1i 40-Cl H b 20.56 32.81 9.99 32.50 \1.00
1j 40-CF3 H a 31.69 [100 \1.00 26.11 \1.00
1k 40-NO2 H b 23.01 42.20 7.75 32.15 1.20
1 l 40-Br H b 18.25 25.57 5.20 27.58 \1.00
1m 40-Br H a 23.86 25.95 \1.00 34.60 \1.00
1n 40-Cl H a 19.73 25.89 2.60 37.93 \1.00
DHAa – – – 8.85 50.18 \1.00 9.81 \1.00
HeLa human cervical carcinoma cell line, MCF-7 breast carcinoma cell line, U937 human leukemic monocyte lymphoma cell line, K562 human
leukemia cell line, HL60 human promyelocytic leukemia cell line
a Dihydroartemisinin, used as a positive control
3172 Med Chem Res (2013) 22:3170–3176
123
the compound 1c was proved to be the most active against the
cancer cells as well as distinctive patterns of selectivity, and
could be considered as anticancer lead agent to be further
research.
In conclusion, we have designed and prepared a series of
artemisinin derivatives, several of which show remarkable
anticancer activity, especially the compound 1c. Further-
more, to perfect the structure–activity relationship, further
research is underway, focusing on bioisosteric exchange of
urea to thiourea and dithiocarbamate as well as exploring
whether the length of the linkage between artemisinin
nucleus and urea moiety has an effect on the activity. These
researches may provide valuable information for further
designing and developing antitumor agents.
Experimental
Chemistry
All melting points were determined (uncorrected) on a
melting point apparatus (Bu¨chi Labortechnik, Flawil,
Switzerland). 1H NMR spectra were obtained using
300 MHz spectrometer (Bruker Bioscience, Billerica, MA,
USA) with tetramethylsilane as an internal standard. Mass
spectra (MS) were taken in ESI mode on LC–MS (Agilent
1100, Palo Alto, CA, USA). Elemental analysis was
determined on a Carlo-Erba 1106 elemental analysis
instrument. Reaction progress was monitored by TLC on
silica gel precoated F254 Merck plates. Preparative flash
column chromatography was performed on the Merck 200
mesh silica gel. Unless otherwise noted, all solvents and
reagents were commercially available and used without
further purification.
General procedure for synthesis of compounds (4a–n)
Compounds 4a–n were prepared by the general synthetic
method outlined in scheme 1. To a mixture of corre-
sponding aniline (0.05 mol) and pyridine (0.10 mol) in
30 ml anhydrous methylene chloride dropwise phenyl
chloroformate (0.05 mol) was added under ice-water bath,
and stirred at room temperature for 4 h. The reaction was
quenched with saturated sodium chloride solution. The
precipitate was filtered and dried, followed by treating with
2-aminoethanol in refluxing ethanol for 7 h, and then the
solvent was removed in vacuo. The residue was purified by
crystallization from ethyl acetate.
Preparation of dihydroartemisinin (6)
Dihydroartemisinin 6 was prepared according to a reported
procedure (Brossi et al., 1988).
General procedure for synthesis of compounds (1a–n)
According to a reported procedure (Li et al., 2000), to a
stirred solution of DHA 6 (0.01 mol) and the corresponding
Table 2 Inhibitory effects of compounds on the growth of HCT116, HepG2, A375-S2, HT1080, and A549 cell lines in vitro
Compd. R1 R2 Configuration at C-10 IC50 (lM)
HCT116 HepG2 A549 A375-S2 HT1080
1a 40-NO2 H a 41.00 34.32 49.64 [100 [100
1b 40-MeO H b 50.70 46.19 63.22 [100 [100
1c 40-CF3 H b 31.33 11.21 29.52 [100 [100
1d 20-Me H b 53.75 53.19 41.70 [100 [100
1e H H b 53.78 34.02 63.34 [100 [100
1f 20-Me 50-Me b 47.46 23.23 33.71 [100 [100
1g 20-Me 30-Cl b 22.67 40.60 26.23 [100 [100
1h 40-F H b 43.73 26.24 43.69 33.72 26.75
1i 40-Cl H b 39.49 36.30 42.01 41.04 43.87
1j 40-CF3 H a 46.35 11.66 37.07 [100 [100
1k 40-NO2 H b 44.42 39.58 53.04 35.87 29.82
1l 40-Br H b 26.71 23.25 34.24 [100 [100
1m 40-Br H a 34.95 24.28 39.44 23.86 35.20
1n 40-Cl H a 37.76 16.62 35.51 9.25 40.26
DHAa – – – 95.41 30.13 76.21 5.48 49.14
HCT116 human colon cancer cell line, HepG2 human hepatocellular liver carcinoma cell line, A375-S2 human melanoma cell line, HT1080
human fibrosarcoma cell line, A549 human lung cancer cell line
a Dihydroartemisinin, used as a positive control
Med Chem Res (2013) 22:3170–3176 3173
123
aromatic urea 4a–n (0.01 mol) in 100 ml dry dichloro-
methane 15 drops of BF3�OEt2 was added. The mixture was
stirred at room temperature 2 h, and the solvent was removed
under reduced pressure. The residue was purified by flash
chromatography on silica gel using petroleum ether/ethyl
acetate as eluent.
1-[2-(10a-Dihydroartemisininoxy)ethyl]-3-(4-nitrophenyl)urea
(1a) Yellow solid (0.22 g, 18.61 %). m.p.: 97–98 �C. 1H
NMR (CDCl3) d: 0.91 (d, J = 7.2 Hz, 3H, 9-CH3), 0.99 (d,
J = 5.7 Hz, 3H, 6-CH3), 1.20–1.34 (m, 3H), 1.40 (s, 3H,
3-CH3), 1.47–2.10 (m, 6H), 2.41–2.42 (m, 1H), 3.36–3.86
(m, 4H), 4.52 (d, J = 9.3 Hz, 1H, 10-H), 5.30 (s, 1H, N–
H), 5.46 (s, 1H, 12-H), 7.56 (d, J = 9.0 Hz, 2H, Ar–H),
7.82 (s, 1H, N–H), 8.13 (d, J = 9.0 Hz, 2H, Ar–H). MS m/
z: 514.2 (M?Na)?, 530.1 (M?K)?, 490.0 (M-H)-, 526.0
(M?Cl)-, 536.0 (M?HCOO-)-. Anal. calcd. For
C24H33N3O8: C, 58.64; H, 6.77; N, 8.55. Found: C, 58.61;
H, 6.74; N, 8.59.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-(4-methoxyphenyl)
urea (1b) White solid (0.56 g, 38.31 %). m.p.: 66–68 �C.
1H NMR (CDCl3) d: 0.80 (d, J = 7.5 Hz, 3H, 9-CH3), 0.92
(d, J = 5.4 Hz, 3H, 6-CH3), 1.16–1.29 (m, 3H), 1.42 (s,
3H, 3-CH3), 1.53–2.05 (m, 6H), 2.31–2.60 (m, 2H),
3.37–3.85 (m, 4H), 3.78 (s, 3H, –OCH3), 4.75 (d,
J = 3.3 Hz, 1H, 10-H), 5.23 (s, 1H, N–H), 5.38 (s, 1H,
12-H), 6.84 (d, J = 9.0 Hz, 2H, Ar–H), 7.20 (d, J =
9.0 Hz, 2H, Ar–H). MS m/z: 477.2 (M?H)?, 499.2
(M?Na)?, 515.2 (M?K)?, 475.1 (M-H)-, 511.0
(M?Cl)-, 521.1 (M?HCOO-)-. Anal. calcd. For C25H36
N2O7: C, 63.01; H, 7.61; N, 5.88. Found: C, 62.98; H, 7.65;
N, 5.85.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-[4-(trifluoromethyl)
phenyl)]urea (1c) Yellow solid (0.11 g, 7.81 %). m.p.:
74–76 �C. 1H NMR (CDCl3) d: 0.88–0.90 (m, 6H, 9-CH3,
6-CH3), 1.26–1.28 (m, 3H), 1.44 (s, 3H, 3-CH3), 1.56–2.07
(m, 6H), 2.39–2.63 (m, 1H), 3.42–3.77 (m, 4H), 4.76 (d,
J = 3.6 Hz, 1H, 10-H), 5.49 (s, 1H, 12-H),5.83 (s, 1H, N–
H), 7.50 (m, 4H, Ar–H). MS m/z: 515.4 (M?H)?, 537.5
(M?Na)?, 553.4 (M?K)?, 513.3 (M-H)-, 549.2
(M?Cl)-, 559.3 (M?HCOO-)-. Anal. calcd. For C25H33
F3N2O6: C, 58.36; H, 6.46; N, 5.44. Found: C, 58.33; H,
6.50; N, 5.40.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-(2-methylphenyl)
urea (1d) White solid (0.13 g, 10.41 %). m.p.: 57–59 �C.
1H NMR (CDCl3) d: 0.73 (d, J = 7.5 Hz, 3H, 9-CH3), 0.92
(d, J = 5.7 Hz, 3H, 6-CH3), 1.18–1.26 (m, 3H), 1.41 (s,
3H, 3-CH3), 1.56–2.05 (m, 6H), 2.28 (s, 3H, Ar–CH3),
2.31–2.29 (m, 2H), 3.34–3.90 (m, 4H), 4.72 (d,
J = 3.3 Hz, 1H, 10-H), 5.04 (t, 1H, N–H), 5.32 (s, 1H,
12-H), 6.35 (s, 1H, N–H), 7.12–7.23 (m, 3H, Ar–H), 7.37
(m, 1H, Ar–H). MS m/z: 483.4 (M?Na)?, 499.4 (M?K)?.
Anal. calcd. For C25H36N2O6: C, 65.20; H, 7.88; N, 6.08.
Found: C, 65.24; H, 7.87; N, 6.11.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-phenylurea (1e)
White solid (0.13 g, 13.23 %). m.p.: 60–62 �C. 1H NMR
(CDCl3) d: 0.84 (d, J = 7.5 Hz, 3H, 9-CH3), 0.90 (d,
J = 5.7 Hz, 3H, 6-CH3), 1.22–1.26 (m, 3H), 1.43 (s, 3H,
3-CH3), 1.58–2.05 (m, 6H), 2.33–2.62 (m, 2H), 3.44–3.82
(m, 4H), 4.76 (d, J = 3.3 Hz, 1H, 10-H), 5.42 (s, 1H,
12-H), 5.49 (s, 1H, N–H), 6.98 (s, 1H, N–H), 7.05–7.35 (m,
5H, Ar–H). MS m/z: 469.3 (M?Na)?, 485.3 (M?K)?,
481.2 (M?Cl)-, 491.3 (M?HCOO-)-. Anal. calcd. For
C24H34N2O6: C, 64.55; H, 7.67; N, 6.27. Found: C, 64.56;
H, 7.70; N, 6.29.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-(2,5-dimethylphenyl)
urea (1f) White solid (0.39 g, 20.33 %). m.p.: 64–66 �C.
1H NMR (CDCl3) d: 0.71 (d, J = 7.2 Hz, 3H, 9-CH3), 0.93
(d, J = 5.7 Hz, 3H, 6-CH3), 1.20–1.25 (m, 3H), 1.41 (s,
3H, 3-CH3), 1.51–2.04 (m, 6H), 2.23 (s, 3H, Ar–CH3), 2.30
(s, 3H, Ar–CH3), 3.33–3.88 (m, 4H), 4.72 (d, J = 3.3 Hz,
1H, 10-H), 4.87 (s, 1H, N–H), 5.31 (s, 1H, 12-H), 5.96 (s,
1H, N–H), 6.94–7.26 (m, 3H, Ar–H). MS m/z: 497.5
(M?Na)?, 513.4 (M?K)?. Anal. calcd. For C26H38N2O6:
C, 65.80; H, 8.07; N, 5.90. Found: C, 65.84; H, 8.03; N,
5.93.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-(3-chloro-2-methyl
phenyl)urea (1g) Yellow solid (0.34 g, 20.01 %). m.p.:
90–93 �C. 1H NMR (CDCl3) d: 0.75 (d, J = 7.5 Hz, 3H,
9-CH3), 0.92 (d, J = 5.7 Hz, 3H, 6-CH3), 1.21–1.34 (m,
3H), 1.39 (s, 3H, 3-CH3), 1.43–2.05 (m, 6H), 2.3 (s, 3H,
Ar–CH3), 2.39–2.60 (m, 2H), 3.34–3.86 (m, 4H), 4.73 (d,
J = 3.6 Hz, 1H, 10-H), 5.07 (m, 1H, N–H), 5.34 (s, 1H,
12-H), 6.53 (m, 1H, N–H), 7.09–7.14 (m, 1H, Ar–H),
7.22–7.25 (m, 1H, Ar–H), 7.32–7.34 (m, 1H, Ar–H). MS
m/z: 517.3 (M?Na)?, 533.2 (M?K)?, 493.2 (M-H)-,
529.2 (M?Cl)-, 539.3 (M?HCOO-)-. Anal. calcd. For
C25H35ClN2O6: C, 60.66; H, 7.13; N, 5.66. Found: C,
60.65; H, 7.10; N, 5.70.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-(4-fluorophenyl)
urea (1h) White solid (0.25 g, 13.67 %). m.p.: 70–72 �C.
1H NMR (CDCl3) d: 0.85 (d, J = 7.2 Hz, 3H, 9-CH3), 0.91
(d, J = 5.4 Hz, 3H, 6-CH3), 1.22–1.26 (m, 3H), 1.42 (s,
3H, 3-CH3), 1.53–2.05 (m, 6H), 2.32–2.63 (m, 2H),
3.41–3.82 (m, 4H), 3.41–3.82 (m, 4H), 4.75 (d,
J = 3.3 Hz, 1H, 10-H), 5.42 (s, 1H, 12-H), 6.94–7.00 (m,
2H, Ar–H), 7.14 (s, 1H, N–H), 7.29-7.31 (m, 2H, Ar–H).
3174 Med Chem Res (2013) 22:3170–3176
123
MS m/z: 487.2 (M?Na)?, 503.2 (M?K)?, 463.1 (M-H)-,
499.1 (M?Cl)-, 509.1 (M?HCOO-)-. Anal. calcd. For
C24H33FN2O6: C, 62.05; H, 7.16; N, 6.03. Found: C, 62.01;
H, 7.14; N, 6.01.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-(4-chlorophenyl)
urea (1i) White solid (0.43 g, 30.21 %). m.p.: 63–66 �C.
1H NMR (CDCl3) d: 0.87 (d, J = 7.2 Hz, 3H, 9-CH3), 0.91
(d, J = 5.4 Hz, 3H, 6-CH3), 1.19–1.26 (m, 3H), 1.43 (s,
3H, 3-CH3), 1.54–2.44 (m, 8H), 3.44–3.82 (m, 4H), 4.75
(d, J = 3.6 Hz, 1H, 10-H), 5.45 (s, 1H, 12-H), 5.61 (s, 1H,
N–H), 7.18 (s, 1H, N–H), 7.22 (d, J = 8.7 Hz, 2H, Ar–H),
7.30 (d, J = 8.7 Hz, 2H, Ar–H). MS m/z: 503.2 (M?Na)?,
519.2 (M?K)?, 479.0 (M-H)-, 515.0 (M?Cl)-, 525.0
(M?HCOO-)-. Anal. calcd. For C24H33ClN2O6: C, 59.93;
H, 6.92; N, 5.82. Found: C, 59.90; H, 6.95; N, 5.85.
1-[2-(10a-Dihydroartemisininoxy)ethyl]-3-[4-(trifluoromethyl)
phenyl]urea (1j) Yellow solid (0.11 g, 8.61 %). m.p.:
87–90 �C. 1H NMR (CDCl3) d: 0.90 (d, J = 6.9 Hz, 3H,
9-CH3), 0.98 (d, J = 5.1 Hz, 3H, 6-CH3), 1.23–1.32 (m,
3H), 1.42 (s, 3H, 3-CH3), 1.57–2.46 (m, 8H), 3.37–3.82 (m,
4H), 4.49 (d, J = 9.3 Hz, 1H, 10-H), 5.42 (s, 1H, 12-H),
6.07 (s, 1H, N–H), 7.47 (d, J = 9.0 Hz, 2H, Ar–H), 7.52
(d, J = 9.0 Hz, 2H, Ar–H), 7.56 (s, 1H, N–H). MS m/z:
515.5 (M?H)?, 537.5 (M?Na)?, 553.4 (M?K)?, 549.2
(M?Cl)-, 559.3 (M?HCOO-)-. Anal. calcd. For
C25H33F3N2O6: C, 58.36; H, 6.46; N, 5.44. Found: C,
58.35; H, 6.49; N, 5.40.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-(4-nitrophenyl)urea
(1k) Yellow solid (0.26 g, 19.64 %). m.p.: 79–81 �C. 1H
NMR (CDCl3) d: 0.91 (d, J = 6.9 Hz, 3H, 9-CH3), 0.93 (d,
J = 5.7 Hz, 3H, 6-CH3), 1.26–1.35 (m, 3H), 1.43 (s, 3H,
3-CH3), 1.51–2.63 (m, 8H), 3.37–3.80 (m, 4H), 4.77 (d,
J = 3.3 Hz, 1H, 10-H), 5.53 (s, 1H, 12-H), 6.04 (m, 1H,
N–H), 7.62 (d, J = 9.3 Hz, 2H, Ar–H), 7.89 (s, 1H, N–H),
8.16 (d, J = 9.3 Hz, 2H, Ar–H). MS m/z: 514.2 (M?Na)?,
530.1 (M?K)?, 490.0 (M-H)-, 526.0 (M?Cl)-, 536.0
(M?HCOO-)-. Anal. calcd. For C24H33N3O8: C, 58.64;
H, 6.77; N, 8.55. Found: C, 58.60; H, 6.80; N, 8.56.
1-[2-(10b-Dihydroartemisininoxy)ethyl]-3-(4-bromophenyl)
urea (1l) White solid (0.26 g, 14.23 %). m.p.: 82–83 �C.
1H NMR (CDCl3) d: 0.88 (d, J = 7.5 Hz, 3H, 9-CH3), 0.91
(d, J = 5.7 Hz, 3H, 6-CH3), 1.25–1.30 (m, 3H), 1.44 (s,
3H, 3-CH3), 1.56–2.06 (m, 6H), 2.38–2.63 (m, 2H),
3.43–3.76 (m, 4H), 4.76 (d, J = 3.3 Hz, 1H, 10-H), 5.44 (s,
1H, 12-H), 5.5 (t, 1H, N–H), 7.02 (s, 1H, N–H), 7.25 (d,
J = 9.0 Hz, 2H, Ar–H), 7.38 (d, J = 9.0 Hz, 2H, Ar–H).
MS m/z: 547.4 (M?Na)?, 565.4 (M?K)?, 559.2 (M?Cl)-,
569.2 (M?HCOO-)-. Anal. calcd. For C24H33BrN2O6: C,
54.86; H, 6.33; N, 5.33. Found: C, 54.85; H, 6.30; N, 5.37.
1-[2-(10a-Dihydroartemisininoxy)ethyl]-3-(4-bromophenyl)
urea (1m) White solid (0.36 g, 17.66 %). m.p.:
77–78 �C. 1H NMR (CDCl3) d: 0.88 (d, J = 7.2 Hz, 3H,
9-CH3), 0.98 (d, J = 5.7 Hz, 3H, 6-CH3), 1.22–1.32 (m,
3H), 1.43 (s, 3H, 3-CH3), 1.54–2.35 (m, 6H), 2.37–2.44 (m,
2H), 3.34–3.54 (m, 4H), 4.46 (d, J = 9.3 Hz, 1H, 10-H),
5.40 (s, 1H, 12-H), 5.93 (s, 1H, N–H), 7.28–7.36 (m, 4H,
Ar–H). MS m/z: 547.4 (M?Na)?, 565.4 (M?K)?, 559.2
(M?Cl)-, 569.2 (M?HCOO-)-. Anal. calcd. For
C24H33BrN2O6: C, 54.86; H, 6.33; N, 5.33. Found: C,
54.88; H, 6.30; N, 5.32.
1-[2-(10a-Dihydroartemisininoxy)ethyl]-3-(4-chlorophenyl)
urea (1n) White solid (0.43 g, 30.21 %). m.p.: 85–87 �C.
1H NMR (CDCl3) d: 0.89 (d, J = 6.9 Hz, 3H, 9-CH3), 0.98
(d, J = 5.4 Hz, 3H, 6-CH3), 1.22–1.32 (m, 3H), 1.43 (s,
3H, 3-CH3), 1.56–2.09 (m, 6H), 2.36–2.45 (m, 2H),
3.31–3.84 (m, 4H), 4.46 (d, J = 9.3 Hz, 1H, 10-H), 5.41 (s,
1H, 12-H), 7.03 (s, 1H, N–H), 7.21 (d, J = 9.0 Hz, 2H,
Ar–H), 7.34 (d, J = 9.0 Hz, 2H, Ar–H). MS m/z: 503.1
(M?Na)?, 519.1 (M?K)?, 479.0 (M-H)-, 514.9
(M?Cl)-, 525.0 (M?HCOO-)-. Anal. calcd. For
C24H33ClN2O6: C, 59.93; H, 6.92; N, 5.82. Found: C,
59.91; H, 6.89; N, 5.85.
Biological assay
The cytotoxic effect of target compounds 1a–n was mea-
sured by the standard MTT assay in vitro (Bonomi, 2002).
The cells were dispensed in 96-well cell culture plates
(NUNC, Roskilde, Denmark) at a density of 5 9 103 cells
per well. After 24 h incubation, the cells were treated with
target compounds 1a–n for 48 h. All wells were washed
three times with PBS, and then RPMI 1640 medium con-
taining 10 % FBS and 0.5 mg/ml MTT were added with
further 4 h incubation at 37 �C. For adherent cells, the
supernatants were gently removed and 150 ll DMSO was
added into each well. The plates were gently shaked for
10 min and were read with a plate reader at 492 nm. For
suspension cells, 100 ll solution (SDS 10 g, 10 M HCl
0.1 ml, isobutanol 5 ml, diluted with distilled waterto
100 ml) was added and the plate was incubated at 37 �C
overnight and read with a plate reader at 492 nm. The
percentage of cell growth inhibition was calculated as
follows:
Med Chem Res (2013) 22:3170–3176 3175
123
Cell growth inhibition %ð Þ ¼
A492;control�A492;experiment
� �
= A492;control�A492;blank
� �
� 100%:
Acknowledgments The authors are grateful to financial support by
the National Science and Technology major projects (Number:
2009ZX09301-012).
Conflict of interest The authors declare that they have no conflict
of interest.
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123
	Synthesis and evaluation in vitro of 1-[2-(10-dihydroartemisininoxy) ethyl]-3-phenylurea derivatives as potential agents against cancer
	Abstract
	Introduction
	Results and discussion
	Chemistry
	Pharmacology and discussion
	Experimental
	Chemistry
	General procedure for synthesis of compounds (4a--n)
	Preparation of dihydroartemisinin (6)
	General procedure for synthesis of compounds (1a--n)
	1-[2-(10 alpha -Dihydroartemisininoxy)ethyl]-3-(4-nitrophenyl)urea (1a)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-(4-methoxyphenyl)nullurea (1b)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-[4-(trifluoromethyl)nullphenyl)]urea (1c)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-(2-methylphenyl)nullurea (1d)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-phenylurea (1e)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-(2,5-dimethylphenyl)nullurea (1f)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-(3-chloro-2-methylnullphenyl)urea (1g)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-(4-fluorophenyl)nullurea (1h)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-(4-chlorophenyl)nullurea (1i)
	1-[2-(10 alpha -Dihydroartemisininoxy)ethyl]-3-[4-(trifluoromethyl)nullphenyl]urea (1j)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-(4-nitrophenyl)urea (1k)
	1-[2-(10 beta -Dihydroartemisininoxy)ethyl]-3-(4-bromophenyl)nullurea (1l)
	1-[2-(10 alpha -Dihydroartemisininoxy)ethyl]-3-(4-bromophenyl)nullurea (1m)
	1-[2-(10 alpha -Dihydroartemisininoxy)ethyl]-3-(4-chlorophenyl)nullurea (1n)
	Biological assay
	Acknowledgments
	References