Synthesis and cytotoxic activity of N-((2-methyl-4(3H)-quinazolinon-6-yl) methyl)dithiocarbamates

Article abstract of DOI:10.1016/j.ejmech.2010.05.038

A series of N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)dithiocarbamates 5a-w were synthesized and evaluated for their cytotoxic activity against five human cancer cell lines. We found that compound 5k inhibited proliferation of A549, MCF-7, HeLa, HT29 and HCT-116 cells with IC₅₀ values of 5.44, 7.15, 12.16, 10.35 and 11.44 μM, respectively. Compound 5i was the most potent with an IC₅₀ value of 3.65 μM against proliferation of MCF-7 cells, while 5n was the most potent with an IC₅₀ value of 5.09 μM against proliferation of A549 cells. Cell cycle analysis showed that both 5i and 5k arrested A549 cells at S and G2/M phases, suggesting that these compounds act through mechanisms different from 5-Fluorouracil, which arrests cells at S phase only. Compounds 5a-w were synthesized and their cytotoxic activity against five human cancer cell lines together with the effects of three representative compounds on A549 cell cycle progression were examined.

Full text of DOI:10.1016/j.ejmech.2010.05.038

European Journal of Medicinal Chemistry 45 (2010) 3850e3857
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and cytotoxic activity of N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamates
,
a
b
b
c
c
c
Sheng-Li Cao a , Yao Wang , Lin Zhu , Ji Liao , Yan-Wen Guo , Lin-Lin Chen , Hong-Qin Liu ,
*
,
Xingzhi Xu b
**
a Department of Chemistry, Capital Normal University, Beijing 100048, PR China
b College of Life Science, Capital Normal University, Beijing 100048, PR China
c School of Science, Beijing Institute of Technology, Beijing 100081, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)dithiocarbamates 5aꢀw were synthesized and
evaluated for their cytotoxic activity against five human cancer cell lines. We found that compound 5k
inhibited proliferation of A549, MCF-7, HeLa, HT29 and HCT-116 cells with IC50 values of 5.44, 7.15, 12.16,
Received 23 February 2010
Received in revised form
16 May 2010
Accepted 19 May 2010
Available online 25 May 2010
10.35 and 11.44
m
M, respectively. Compound 5i was the most potent with an IC50 value of 3.65
m
M against
proliferation of MCF-7 cells, while 5n was the most potent with an IC50 value of 5.09
m
M against
proliferation of A549 cells. Cell cycle analysis showed that both 5i and 5k arrested A549 cells at S and G2/
M phases, suggesting that these compounds act through mechanisms different from 5-Fluorouracil,
which arrests cells at S phase only.
Keywords:
4(3H)-quinazolinone
Dithiocarbamate
Synthesis
Ó 2010 Elsevier Masson SAS. All rights reserved.
Cytotoxic activity
Cell cycle
1. Introduction
moiety inhibited the growth of A549 (human lung cancer), HCT-8
(human colorectal cancer) and Bel-7402 (human liver cancer) cell
Incorporation of lipophilic side chains into the benzene ring of
quinazoline has led to the emergence of many effective anticancer
drugs such as AG337 and Trimetrexate (Fig. 1) [1e7]. In the search
for quinazoline-based anticancer agents at our laboratory, dithio-
carbamate moiety occurring in phytoalexins Brassinin (Fig. 1),
which possesses cancer chemopreventive activity and cytotoxic
effect [8e12], had been introduced into the C6 position of 2-
methyl-4(3H)-quinazolinone. Among the resulting compounds, 1a
(Fig. 1) derived from benzylamine displayed obviously in vitro
inhibitory activity against K562 (human myelogenous leukemia)
and HeLa (human cervical carcinoma) cells [13,14]. Thereafter,
another series of derivatives of 2-methyl-4(3H)-quinazolinone had
been synthesized from benzylamines bearing different substituents
in the benzene ring or heterocyclomethylamines, of which
compound 1b (Fig. 1) containing (pyridin-3-yl)methylamine
lines with IC50 values within micromolar concentrations [15]. For 4
(3H)-quinazolinones bearing other alkyl than methyl, aryl, or het-
eroaryl group at the C2 position, however, their dithiocarbamate
derivatives hardly showed any inhibitory activity against the
growth of A549, HCT-8 and Bel-7402 cell lines [16]. Therefore, it
seems that a methyl group at the C2 position is crucial for this type
of compounds to generate cytotoxic effects.
Our previous work on the synthesis and cytotoxic evaluation
of dithiocarbamate derivatives of 2-methyl-4(3H)-quinazolinone
indicated that the connection of amino group with benzyl or
heterocyclomethyl group is favorable to the cytotoxic activity
[13e15]. As to the structure of Brassinin, it is noteworthy that
the amino group of dithiocarbamate moiety is also linked with
indole ring by methylene. In this context, the structurally
reversed analogs of compounds 1a and 1b would be synthesized,
so that 4(3H)-quinazolinone moiety previously served as ester
component would be converted into amino component. Herein,
we report the synthesis of a new series of N-((2-methyl-4(3H)-
* Corresponding author. Tel./fax: þ86 10 68902974.
** Corresponding author. Tel./fax: þ86 10 68902240.
E-mail addresses: sl_cao@sohu.com (S.-L. Cao), xingzhi_xu@mail.cnu.edu.cn
(X. Xu).
quinazolinon-6-yl)methyl)dithiocarbamates (Scheme
1
and
Table 1) and evaluation of these compounds as potential anti-
cancer agents.
0223-5234/$ e see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.05.038

S.-L. Cao et al. / European Journal of Medicinal Chemistry 45 (2010) 3850e3857
3851
Table 1
Cytotoxic activity (IC50
lines.
, mM) of compounds 5aew against five human cancer cell
Compound
R
IC50
(mM)
A549 MCF-7 HeLa HT29 HCT-116
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
5q
5r
5s
5t
5u
5v
5w
1a
CH3e
CH3CH2e
CH3CH2CH2e
(CH3)2CHe
n.t.a
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
12.78 13.22
5.49 3.65
20.66 29.80
5.44 7.15
n.t. n.t.
8.39
5.09
5.49
17.20
9.49
7.73
6.54
n.t.
15.03
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
10.35 n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
15.61
12.33
19.41
12.52
n.t.
CH3(CH2)2CH2e
CH3(CH2)3CH2e
CH3(CH2)4CH2e
CyclohexylCH2e
Ph2C(CN)CH2CH2e
CH2]CHCH2e
HC^CCH2e
PhCH2e
4-CH3PhCH2e
4-FPhCH2e
n.t.
n.t.
12.16 10.35 11.44
n.t.
n.t.
13.73 13.68 11.57
n.t. 12.21 n.t.
20.45 12.26 11.92
15.34 15.18 15.77
13.38 14.88 13.98
8.54 13.51 11.26
n.t.
n.t.
8.85
7.07
8.43
6.65
8.69
7.15
7.74
n.t.
9.94
6.41
12.58 15.64
Fig. 1. Structures of AG337, Trimetrexate, Brassinin, compounds 1a and 1b.
2,4-F2PhCH2e
F5PhCH2e
2. Chemistry
4-ClPhCH2e
4-BrPhCH2e
4-NO2PhCH2e
4-CNPhCH2e
4-HO2CPhCH2e
4-EtO2CPhCH2e
Ph2CHe
As shown in Scheme 1, two methods, namely Gabriel method
and Delepine method, were exploited to prepare the key inter-
mediate 6-(aminomethyl)-2-methyl-4(3H)-quinazolinone dihy-
drochloride (4) required for the synthesis of the target molecules. In
the case of Gabriel method, the reaction of 6-(bromomethyl)-2-
methyl-4(3H)-quinazolinone (2) with potassium phthalimide
gave 2-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)isoindoline-
1,3-dione (3), which was converted into 4 by hydrazinolysis, and
then acidification with dilute hydrochloric acid. Alternatively,
intermediate 4 was obtained by acid treatment of the crude product
from heating 2 with hexamethylenetetramine. In comparison with
Gabriel method, Delepine method possessed advantages such as
shorter reaction time, higher yield and no isolation and purification
of intermediate required, thus, it was adopted for the preparation of
4 in this study. Eventually, treating 4 and carbon disulfide with
different halohydrocarbon in the presence of anhydrous potassium
phosphate generated the target compounds 5aew.
n.t.
n.t.
n.t.
n.t.
n.t.
n.t.
10.69
7.25
18.00 19.16
11.70 25.06
5.58 12.65
20.07 16.19 11.56
n.t. n.t. n.t.
18.40 10.91 18.26
13.66
98.54
1b
4.71
8.46
3.65
6.00
5-Fluorouracil
6.76
7.99
a
n.t.: Cell growth inhibition was lower than 50% at the concentration of 10
and the IC50 value was not tested.
mg/mL
concentration that causes 50% of cell growth inhibition) values
were determined and summarized in Table 1. The previously
synthesized compounds 1a, 1b [13,14] and the anticancer drug 5-
Fluorouracil were used as positive controls.
Among the tested compounds, 5i, 5k and 5s presented prom-
inent activity against the growth of A549 with IC50 values of 5.49,
5.44, and 6.54
further investigation in A549 cell line. The tested compounds with
concentrations of 12.0, 6.0 and 3.0 M in medium, were incubated
mM, respectively. Therefore, they were selected for
3. Pharmacology
m
with the cells for 4, 8, 12, 24, 36 and 48 h, and cell proliferation was
monitored at each timepoint using MTT assay. Identical amount of
DMSO in medium served as the vehicle control. The growth curves
of A549 cells treated with 5i, 5k, 5s or 5-Fluorouracil were illus-
trated in Fig. 2.
3.1. Cytotoxic evaluation
The MTT [3-(4,5-dimethylthiazo-2-yl)-2,5-diphenyl-tetrazo-
lium bromide] cell proliferation assay was used to evaluate cyto-
toxic activity of the synthesized compounds against five human
cancer cell lines including A549 (lung cancer), MCF-7 (breast
adenocarcinoma), HeLa (cervical carcinoma), HT29 and HCT-116
(colorectal cancer) cell lines. The inhibition of cell proliferation was
determined 72 h after cells were exposed to the tested compounds
3.2. Cell cycle analysis
To gain further insight into the mechanisms of action of these
new and active compounds, we examined if treatment with these
compounds in A549 cells would lead to cell cycle arrest. A549 cells
were treated with 5i, 5k, 5s, or 5-Fluorouracil at a concentration
equivalent to its IC50 for 24 h. Cells were harvested for cell cycle
at a concentration of 10
mg/mL. The compounds with 50% or more
inhibition compared with vehicle-treated cells were considered
active. Inhibition of cell proliferation by these active compounds at
various concentrations was further measured, and their IC50 (the
Scheme 1. Synthetic pathway for compounds 5aew. Reagents and conditions: a. potassium phthalimide, DMF, reflux 8 h; b. (i) 80% hydrazine hydrate, ethanol, reflux 5 h; (ii) 2 mol/
L HCl, 60e90 ꢁC 3 h; c. hexamethylenetetramine, trichloromethane, reflux 4 h; d. concentrated HCl, methanol, reflux 2 h; e. CS2, K3PO4, RX, DMF, rt, 2e12 h.

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S.-L. Cao et al. / European Journal of Medicinal Chemistry 45 (2010) 3850e3857
Fig. 2. Cytotoxic effects of 5i (A), 5k (B), 5s (C) and 5-Fluorouracil (D) on A549 cells. Adherent cells proliferated in 96-well plates (7.5 ꢂ103 cells/well) were incubated with a serial of
concentrations (12.0, 6.0 and 3.0 mM) and determined by MTT assay at various time intervals.
analysis by a flow cytometry. Three independent experiments were
performed, and data from one representative experiment were
shown in Fig. 3 and Table 2.
enhance the activity (5o and 5p), in comparison to compound 5n.
Interestingly, either electron-donating group (methyl in 5m) or
electron-withdrawing group (nitro in 5s, carboxyl in 5u or ethyl-
oxycarbonyl in 5v) in the position 4 of benzyl group was favorable
to the cytotoxic activity, with the exception of cyano group
retaining the activity of 5t with the unsubstituted compound 5l.
These results suggested that the electronic effect of substituents
attached to the benzyl ring was not a critical determinant of
activity, while the steric effect would play a role in generating
cytotoxicity for this type of compounds. Consequently, by
comparing 5l with 5m, 5s with 5t, and 5u with 5v, it could be found
that the bulky groups at the 4-position of the benzene rings were
more beneficial to the cytotoxicity over the small ones.
4. Results and discussion
To search for more efficient anticancer agents and to investigate
the contribution of dithiocarbamate moiety in target molecules to
the cytotoxic activity, alkyl, substituted alkyl, alkenyl and alkynyl
groups were chosen to serve as R substituents for compounds
5aek. Meanwhile, different benzyl groups were incorporated in
compounds 5lew to examine the electronic or steric effect of
substituents attached to the benzyl ring on the cytotoxic activity.
Of the compounds listed in Table 1, 5k containing propargyl
group exhibited inhibitory activity against proliferation of all five
It also can be seen from Table 1 that the cytotoxic activity of this
type of compounds against A549 and MCF-7 cells was in general
more potent than that against HeLa, HT29 and HCT-116 cells.
Among them, compound 5i was the most effective with an IC50
cell lines with IC50 values in the range of 5.44e12.16 mM, whereas 5j
containing allyl group only inhibited proliferation of two cell lines
(A549 and MCF-7) with IC50 values of 20.66 and 29.80, respectively.
Among the compounds 5aeg in which R is a different alkyl group,
five compounds (5a and 5cef) showed moderate activity against
MCF-7 cells only. Introduction of cyano and phenyl or cyclohexyl
groups enhanced the inhibitory activity, in which 5h had IC50
value of 3.65
mM against MCF-7 cells, while 5n was the most
effective with an IC50 value of 5.09
mM against A549 cells. It is worth
noting that IC50 values of both 5n and 5i against A549 and MCF-7
cells were lower than those of the positive control 5-Fluorouracil.
Moreover, the active compounds (5k, 5n, 5pes and 5v) against
values of 12.78, 13.22 and 10.35
mM against A549, MCF-7 and HT29
HeLa cells were more potent than 5-Fluorouracil (IC50, 98.54
mM),
cells, and 5i 5.49 and 3.65 M against A549 and MCF-7, respec-
m
the most effective compound 5s had an IC50 value of 8.54
mM.
tively. These results indicated that the R substituents containing
electron-rich groups, such as allyl, propargyl or phenyl, were
favorable to the cytotoxic activity.
However, the active compounds (5k, 5mes and 5v) against HT29
and HCT-116 cells were less potent than 5-Fluorouracil which
possessed the IC50 values lower than 10 mM. In addition, these
Unlike lead compound 1a in which 4(3H)-quinazolinone moiety
served as ester component, its counterpart 5l was inactive against
all five cell lines. Introduction of a halogen atom in the position 4 of
benzyl group in 5l led to a substantial increase of cytotoxic activity,
and fluorine atom was more favorable than chlorine or bromine
atom. Thus, in the resulting compounds, 5n showed lower IC50
values against five cell lines than those of 5q or 5r. However,
introduction of the second or more fluorine atoms could not further
active compounds exhibited cytotoxicity against A549, MCF-7 and
HeLa cell lines similar to compound 1b, but were less potent than
1b against HT29 and HCT-116 cells.
The growth curves indicated that all three representative
compounds 5i, 5k and 5s can effectively suppress cell proliferation
in a dose-dependent manner (Fig. 2). Among the three tested
compounds, 5k was the most effective, which inhibited almost
100% of cell proliferation at 12 mM within 8 h of treatment, about

S.-L. Cao et al. / European Journal of Medicinal Chemistry 45 (2010) 3850e3857
3853
Fig. 3. Effects of compounds 5i, 5k and 5s on cell cycle distribution. A549 cells were treated with 5i (D), 5k (E), 5s (F), or 5-Fluorouracil (C) at a concentration equivalent to its IC50
for 24 h.
40% at 6
compared with the effect of 5-Fluorouracil at the concentration of
12 M, 5k showed more rapid and efficient inhibitory activity
against A549 cells. Particularly, 5k at a final concentration of 12
was killing almost 100% of cells within 8 h of treatment, whereas 5-
Fluorouracil at the same concentration only about 12% within the
same duration, and even after 48 h treatment 5-Fluorouracil merely
killed 55% cells. These results were also consistent with IC50 values
we determined in this study.
m
M, and 25% at 3
m
M after 48 h of treatment. Moreover, as
cells with 5-Fluorouracil almost completely arrested the cell cycle
at S phase. Treatment of A549 cells with 5i, 5k, or 5s led to
a moderate arrest at both S phase and G2/M phase (Fig. 3 and Table
2). Our results suggest that these new compounds may target both
at DNA replication and mitosis progression. Further investigation of
their mechanisms is warranted.
m
mM
5. Conclusion
5-Fluorouracil is a well known inhibitor of thymidylate syn-
thase, thus blocks DNA replication. As expected, treatment of A549
A number of dithiocarbamates in which 4(3H)-quinazolinone
moiety served as amino component were synthesized and evalu-
ated for their cytotoxic activity against five human cancer cell lines.
Our results indicated that compound 5k inhibited the growth of
five cell lines with lower IC50 values than those of lead compound
1a for each cell line. Compound 5i and 5n selectively inhibited
Table 2
Effects of compounds 5i, 5k and 5s on A549 cell cycle progression.
Compound
Cell cycle distribution (%)
proliferation of MCF-7 or A549 cells with IC50 values of 3.65
mM and
G0/G1
S
G2/M
5.09 M, respectively, and both of them were more potent than lead
m
Untreated
DMSO
5-Fluorouracil
5i
5k
5s
46.9
45.3
7.5
32.1
26.1
26.1
37.6
36.8
84.2
47.2
47.7
46.5
15.3
17.9
8.4
20.7
26.3
27.4
compound 1a and the positive control 5-Fluorouracil. These results
suggest that the structurally reversed modification of lead
compound 1a is a fairly efficient approach to enhance its cytotoxic
activity. Cell cycle profiling suggests that these compounds target
the cell cycle at both DNA replication and mitosis progression.

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S.-L. Cao et al. / European Journal of Medicinal Chemistry 45 (2010) 3850e3857
6. Experimental
1H, J ¼ 8.4 Hz, quinazolinone 8-H), 8.05 (d, 1H, J ¼ 8.4 Hz, quina-
zolinone 7-H), 8.31 (s, 1H, quinazolinone 5-H). ESI-MS m/z: 190 [M-
2HCl þ H]þ. Anal. calcd for C10H13Cl2N3O$0.3H2O: C, 44.89; H, 5.12;
N, 15.71. Found: C, 44.82; H, 4.98; N, 15.72.
Melting points were determined on an X-6 microscopic melting
point apparatus and are uncorrected. IR spectra were recorded on
a Bruker Tensor 27 spectrometer. 1H NMR spectra were recorded on
a Bruker AC-200P spectrometer at 200 MHz using tetramethylsi-
lane (TMS) as internal standard. Low-resolution electrospray ioni-
zation (ESI) mass spectra were recorded on a Bruker Daltonics
Esquire-LC 00136 mass spectrometer, and high-resolution electro-
spray ionization (HR-ESI) mass spectra were recorded on an Agilent
LC/SMD TOF mass spectrometer. Elemental analyses were per-
formed by Institute of Chemistry, Chinese Academy of Science, on
a Flash EA 1112 elemental analyzer. Column chromatography was
carried out on silica gel (200e300 mesh). 6-Bromomethyl-2-
methyl-4(3H)-quinazolinone (2) was prepared according to the
reported method [15], and other commercially available reagents
were used without further purification. Compounds 5a and 5d
were prepared with iodomethane and 2-iodopropane respectively,
while the other compounds were prepared with the corresponding
bromohydrocarbons.
6.2. Procedure for the preparation of 6-(aminomethyl)-2-methyl-4
(3H)-quinazolinone dihydrochloride (4) through Delepine method
A mixture of 6-(bromomethyl)-2-methyl-4(3H)-quinazolinone
(2) (1.20 g, 4.6 mmol) and hexamethylenetetramine (0.66 g,
4.6 mmol) in trichloromethane (83 mL) was refluxed for 4 h. After
cooling to room temperature, the separated solid was collected by
filtration and dried in air. The solid was dissolved in methanol
(83 mL) under reflux, and then concentrated hydrochloric acid
(6 mL) was added. The mixture was refluxed for 2 h, and then
cooled to room temperature. The precipitate was collected by
filtration and purified by recrystallization from acetone/water (5:2)
to yield 0.85 g (71%) as a white solid, mp > 320 ꢁC. All the spec-
troscopic data were in accordance with those of compound 4
obtained through Gabriel method.
Human breast cancer cell line MCF-7, human lung cancer cell
line A549, human cervical carcinoma cell cline HeLa and human
colon carcinoma cell lines HT29 and HCT-116 were kindly provided
by Laboratory of Cancer Biology, Capital Normal University. All
cancer cells were grown in high glucose DMEM supplemented with
10% fetal bovine serum, 2 mM glutamine, 50 units/mL penicillin,
and 50 mg/mL streptomycin. Cell proliferation assay was deter-
mined using the CellTiter 96Ò AQueous One Solution Cell Prolifera-
tion Assay (Promega).
6.3. General procedure for the preparation of compounds 5aew
To a solution of 6-(aminomethyl)-2-methyl-4(3H)-quinazoli-
none dihydrochloride (4) (0.52 g, 2.0 mmol) in DMF (25 mL), finely
powered anhydrous potassium phosphate (0.51 g, 2.4 mmol) and
carbon disulfide (0.8 mL, 13.3 mmol) were added. After stirring for
1.5 h at room temperature, halohydrocarbon (3 mmol) was added
and the stirring was continued till completion of the reaction
(monitored by TLC). The reaction mixture was poured into water
(150 mL), and the crude product was obtained by filtration if there
was solid separated off. Otherwise, the solution was extracted with
dichloromethane (20 mL ꢂ 3) and the combined organic phases
were dried over anhydrous sodium sulfate, evaporated under
reduced pressure to dryness. The crude product was purified by
column chromatography (CC) on silica gel or recrystallization from
appropriate solvent to give compounds 5aew.
6.1. Synthesis of 6-(aminomethyl)-2-methyl-4(3H)-quinazolinone
dihydrochloride (4) through Gabriel method
6.1.1. Preparation of 2-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)isoindoline-1,3-dione (3)
A solution of 6-(bromomethyl)-2-methyl-4(3H)-quinazolinone
(2) (2.20 g, 8.7 mmol) and potassium phthalimide (2.09 g,
11.3 mmol) in N,N-dimethylformamide (DMF) (30 mL) was refluxed
for 8 h. After stirring at room temperature overnight, the reaction
mixture was poured into water (280 mL). The precipitate formed
was collected by filtration and recrystallized from DMF/H2O (3:2) to
6.3.1. Methyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5a)
Yield 40%, white solid, mp 224e225 ꢁC (CC, eluent: dichloro-
yield 1.93 g (70%) as a white solid, mp 297e299 ꢁC. IR (KBr, cmꢀ1
3165, 3026, 2948, 2871, 1707, 1679, 1617, 1426, 1395, 1097, 948, 845,
752, 712. 1H NMR (CDCl3)
: 2.57 (s, 3H, C2eCH3), 4.97 (s, 2H, NCH2),
)
n
:
methane/methanol ¼ 97:3). IR (KBr, cmꢀ1
) n: 3163, 3029, 2817,1719,
1655, 1568, 1509, 1421, 1395, 1304, 1096, 950, 888. 1H NMR (DMSO-
d
d6) d: 2.56 (s, 3H, CH3), 2.61 (s, 3H, CH3), 4.98 (s, 2H, CH2NH), 7.75 (d,
7.63 (d,1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.70e7.89 (m, 5H, AreH),
8.29 (s, 1H, quinazolinone 5-H), 11.76 (br s, 1H, NH). ESI-MS m/z:
342 [M þ Na]þ. Anal. calcd for C18H13N3O3$1/5H2O: C, 66.95; H,
4.18; N, 13.01. Found: C, 67.17; H, 4.21; N, 13.23.
1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.89 (d, 1H, J ¼ 8.4 Hz, quina-
zolinone 7-H), 8.04 (s, 1H, quinazolinone 5-H). ESI-MS m/z: 280
[M þ H]þ. ESI-HRMS m/z: calcd for C12H14N3OS2 ([M þ H]þ):
280.0578; found: 280.0564.
6.1.2. Preparation of 6-(aminomethyl)-2-methyl-4(3H)-
quinazolinone dihydrochloride (4)
6.3.2. Ethyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5b)
To a heated solution of 2-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)isoindoline-1,3-dione (3) (1.65 g, 5.2 mmol) in ethanol
(75 mL), 80% hydrazine hydrate (0.48 g, 7.8 mmol) and ethanol
(8 mL) were added and the mixture was refluxed for 5 h. The
solvents were evaporated under reduced pressure to dryness and
the residue dissolved in 2 mol/L HCl (72 mL). The mixture was
heated for 2 h at 60 ꢁC, and then for an additional hour at 90 ꢁC,
followed by addition of water (28 mL). After cooling to room
temperature, the precipitate was filtered off and the filtrate was
evaporated under reduced pressure to dryness. The crude product
was purified by recrystallization from acetone/water (5:2) to yield
Yield 42%, off-white solid, mp 215e217 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 97:3). IR (KBr, cmꢀ1
) n: 3241, 3020,
2924, 1658, 1620, 1488, 1384, 1097, 805, 837. 1H NMR (DMSO-d6)
d:
1.24 (t, 3H, J ¼ 7.3 Hz, CH2CH3), 2.34 (s, 3H, C2eCH3), 3.18 (q, 2H,
J ¼ 7.3 Hz, CH2CH3), 4.94 (s, 2H, CH2NH), 7.54 (d, 1H, J ¼ 8.3 Hz,
quinazolinone 8-H), 7.68 (d, 1H, J ¼ 8.3 Hz, quinazolinone 7-H), 7.97
(s, 1H, quinazolinone 5-H). ESI-MS m/z: 294 [M þ H]þ. Anal. calcd
for C13H15N3OS2: C, 53.22; H, 5.15; N, 14.32. Found: C, 53.22; H,
5.34; N, 13.98.
6.3.3. Propyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5c)
1.02 g (75%) as a white solid, mp > 320 ꢁC. IR (KBr, cmꢀ1
2903, 2696, 1718, 1652, 1596, 1567, 1499, 1302, 1095, 895, 782. 1H
NMR (DMSO-d6) : 2.61 (s, 3H, C2eCH3), 4.21 (s, 2H, CH2N), 7.83 (d,
) n: 3424,
Yield 36%, yellowish solid, mp 200e202 ꢁC (CC, eluent:
d
dichloromethane/methanol ¼ 9:1). IR (KBr, cmꢀ1
) n: 3204, 2960,

S.-L. Cao et al. / European Journal of Medicinal Chemistry 45 (2010) 3850e3857
3855
2923, 1678, 1619, 1488, 1376, 1309, 940. 1H NMR (CDCl3)
d: 1.02 (t,
ESI-MS m/z: 362 [M þ H]þ. Anal. calcd for C18H23N3OS2: C, 59.80; H,
3H, J ¼ 7.2 Hz, CH2CH3), 1.76 (m, 2H, CH2CH2CH3), 2.56 (s, 2H,
C2eCH3), 3.30 (t, 2H, J ¼ 6.8 Hz, SCH2CH2), 5.03 (s, 2H, CH2NH), 7.51
(d, 1H, J ¼ 8.3 Hz, quinazolinone 8-H), 7.68 (d, 1H, J ¼ 8.3 Hz, qui-
nazolinone 7-H), 8.09 (s, 1H, quinazolinone 5-H). ESI-MS m/z: 308
[M þ H]þ. Anal. calcd for C14H17N3OS2: C, 54.69; H, 5.57; N, 13.67.
Found: C, 54.65; H, 5.71; N, 13.68.
6.41; N, 11.62. Found: C, 59.57; H, 6.37; N, 11.61.
6.3.9. 3-Cyano-3,3-diphenylpropyl N-((2-methyl-4(3H)-
quinazolinon-6-yl)methyl)dithiocarbamate (5i)
Yield 55%, white solid, mp 165.5e167.4 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 96:4). IR (KBr, cmꢀ1
)
n
: 3442, 3168, 2932, 1656,
1620, 1490, 1384, 1313, 1086, 933, 697. 1H NMR (DMSO-d6)
d: 2.33 (s,
6.3.4. Isopropyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5d)
3H, C2eCH3), 2.84 (m, 2H, CH2), 3.17 (m, 2H, CH2), 4.91 (s, 2H, CH2NH),
7.39 (m, 10H, PheH), 7.53 (d, 1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.67
(d,1H, J ¼ 8.4 Hz, quinazolinone 7-H), 7.95 (s,1H, quinazolinone 5-H).
ESI-MS m/z: 485 [M þ H]þ. Anal. calcd for C27H24N4OS2$1/2H2O: C,
65.69; H, 5.10; N, 11.35. Found: C, 65.74; H, 5.04; N, 11.39.
Yield 55%, white solid, mp 193.0e193.8 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 9:1). IR (KBr, cmꢀ1
)
n
: 3240, 3156, 2957, 1652,
1618, 1487, 1383, 1320, 1092, 946, 852. 1H NMR (DMSO-d6)
d: 1.32 (d,
6H, J ¼ 6.8 Hz, CH(CH3)2), 2.33 (s, 3H, C2eCH3), 3.90 (m,1H, SCH), 4.92
(s, 2H, CH2NH), 7.53 (d,1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.67 (d,1H,
J ¼ 8.4 Hz, quinazolinone 7-H), 7.95 (s, 1H, quinazolinone 5-H). ESI-
MS m/z: 308 [M þ H]þ. Anal. calcd for C14H17N3OS2: C, 54.69; H, 5.57;
N, 13.67. Found: C, 54.76; H, 5.66; N, 13.71.
6.3.10. Allyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5j)
Yield 53%, white solid, mp 189.4e189.9 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 96:4). IR (KBr, cmꢀ1
)
n
: 3425, 1680,
: 2.35 (s,
1618,1488,1384,1314,1090, 939, 833. 1H NMR (DMSO-d6)
d
6.3.5. Butyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5e)
3H, C2eCH3), 3.92 (d, 2H, J ¼ 6.8 Hz, SCH2CH), 4.94 (s, 2H, CH2NH),
5.12 (d, 1H, J ¼ 9.9 Hz, ¼ CH2), 5.29 (d, 1H, J ¼ 16.9 Hz, ¼ CH2), 5.84
(m, 1H, CH ¼ CH2), 7.55 (d, 1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.69
(d, 1H, J ¼ 8.4 Hz, quinazolinone 7-H), 7.97 (s, 1H, quinazolinone 5-
H). ESI-MS m/z: 306 [M þ H]þ. Anal. calcd for C14H15N3OS2$1/2H2O:
C, 53.48; H, 5.13; N, 13.36. Found: C, 53.42; H, 5.14; N, 13.29.
Yield 34%, yellow solid, mp 198e199 ꢁC (from DMF/H2O ¼ 1:1.2).
IR (KBr, cmꢀ1
) n: 3211, 3003, 2957, 2927, 1679, 1620, 1488, 1376,
1308, 1080, 939, 830. 1H NMR (DMSO-d6)
d
: 0.88 (t, 3H, J ¼ 7.2 Hz,
CH2CH3), 1.36 (m, 2H, CH2CH2CH3), 1.59 (m, 2H, CH2CH2CH3), 2.34
(s, 3H, C2eCH3), 3.20 (t, 2H, J ¼ 7.2 Hz, SCH2CH2), 4.94 (s, 2H,
CH2NH), 7.54 (d, 1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.68 (d, 1H,
J ¼ 8.4 Hz, quinazolinone 7-H), 7.96 (s, 1H, quinazolinone 5-H). ESI-
MS m/z: 322 [M þ H]þ. Anal. calcd for C15H19N3OS2: C, 56.04; H,
5.96; N, 13.07. Found: C, 55.91; H, 5.97; N, 13.07.
6.3.11. Prop-2-ynyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5k)
Yield 79%, white solid, mp 188.7e189.7 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 95:5). IR (KBr, cmꢀ1
)
n
: 3443, 3294, 3240, 2922,
1660, 1619, 1487, 1384, 1093, 944, 854. 1H NMR (DMSO-d6)
d: 2.35 (s,
6.3.6. Pentyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5f)
3H, C2eCH3), 3.17 (t, 2H, J ¼ 2.6 Hz, C^CH), 4.09 (d, 2H, J ¼ 2.6 Hz,
CH2C^), 4.95 (s, 2H, CH2NH), 7.56 (d,1H, J ¼ 8.4 Hz, quinazolinone 8-
H), 7.71 (d, 1H, J ¼ 8.4 Hz, quinazolinone 7-H), 7.99 (s, 1H, quinazoli-
none 5-H). ESI-MS m/z: 304 [M þ H]þ. Anal. calcd for C14H13N3OS2: C,
55.42; H, 4.32; N, 13.85. Found: C, 55.02; H, 4.38; N, 13.59.
Yield 64%, white solid, mp 180.5e182.1 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 96:4). IR (KBr, cmꢀ1
) n: 3216, 2921,
1679, 1621, 1526, 1486, 1375, 1308, 1074, 938, 849. 1H NMR (DMSO-
d6)
d
: 0.86 (t, 3H, J ¼ 6.6 Hz, CH2CH3), 1.30 (m, 4H, CH2CH2CH3), 1.60
(m, 2H, SCH2CH2CH2), 2.34 (s, 3H C2eCH3), 3.19 (t, 2H, J ¼ 7.3 Hz,
SCH2CH2), 4.94 (s, 2H, CH2NH), 7.54 (d,1H, J ¼ 8.4 Hz, quinazolinone
8-H), 7.69 (dd, 1H, J ¼ 8.4 and 2.0 Hz, quinazolinone 7-H), 7.96 (d,
1H, J ¼ 2.0 Hz, quinazolinone 5-H). ESI-MS m/z: 336 [M þ H]þ. Anal.
calcd for C16H21N3OS2: C, 57.28; H, 6.31; N,12.53. Found: C, 57.09; H,
6.40; N, 12.32.
6.3.12. Benzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5l)
Yield 32%, white solid, mp 186.6e188 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 95:5). IR (KBr, cmꢀ1
) n: 3441, 3229, 3024, 2925,
1661, 1618,1489, 1383,1322,1088, 946, 854, 702. 1H NMR (DMSO-d6)
d: 2.35 (s, 3H, C2eCH3), 4.53(s, 2H, SCH2), 4.96(s, 2H, CH2NH), 7.29 (m,
5H, PheH), 7.55 (d, 1H, J ¼ 8.3 Hz, quinazolinone 8-H), 7.70 (d, 1H,
J ¼ 8.3 Hz, quinazolinone 7-H), 7.99 (s, 1H, quinazolinone 5-H). ESI-
MS m/z: 356 [M þ H]þ. Anal. calcd for C18H17N3OS2$H2O: C, 57.88; H,
5.13; N, 11.25. Found: C, 58.09; H, 4.87; N, 10.90.
6.3.7. Hexyl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5g)
Yield 67%, yellow solid, mp 195e197 ꢁC (from DMF/H2O ¼ 0.8:1).
IR (KBr, cmꢀ1
) n: 3216, 2955, 2926, 2854, 1680, 1621, 1487, 1376,
1309, 1075, 939, 830. 1H NMR (DMSO-d6)
d
: 0.84 (t, 3H, J ¼ 6.6 Hz,
6.3.13. 4-Methylbenzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5m)
CH2CH3), 1.27 (m, 6H, CH2 ꢂ 3), 1.58 (m, 2H, CH2), 2.33 (s, 3H,
C2eCH3), 3.18 (t, 2H, J ¼ 6.1 Hz, SCH2CH2), 4.92 (s, 2H, CH2NH), 7.53
(d, 1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.67 (d, 1H, J ¼ 8.4 Hz, qui-
nazolinone 7-H), 7.95 (s, 1H, quinazolinone 5-H). ESI-MS m/z: 350
[M þ H]þ. Anal. calcd for C17H23N3OS2: C, 58.42; H, 6.63; N, 12.02.
Found: C, 58.34; H, 6.62; N, 12.08.
Yield 72%, white solid, mp 199.2e200.2 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 95:5). IR (KBr, cmꢀ1
) n: 3449, 3234,
3022, 2927, 1654, 1618, 1487, 1317, 1090, 955, 853. 1H NMR (DMSO-
d6) d: 2.27 (s, 3H, PheCH3), 2.35 (s, 3H, C2eCH3), 4.47 (s, 2H, SCH2),
4.95 (s, 2H, CH2NH), 7.12 (d, 1H, J ¼ 7.9 Hz, PheH), 7.26 (d, 1H,
J ¼ 7.9 Hz, PheH), 7.55 (d, 1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.69
(dd, 1H, J ¼ 8.4, 2.0 Hz, quinazolinone 7-H), 7.98 (d, 1H, J ¼ 2.0 Hz,
quinazolinone 5-H). ESI-MS m/z: 370 [M þ H]þ. Anal. calcd for
C19H19N3OS2$1/2CH3OH: C, 60.75; H, 5.49; N, 10.90. Found: C,
60.71; H, 5.14; N, 10.82.
6.3.8. Cyclohexylmethyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5h)
Yield 65%, white solid, mp 191.4e192.1 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 95:5). IR (KBr, cmꢀ1
) n: 3445, 3227,
2921, 2851, 1661, 1617, 1485, 1371, 1322, 1099, 945, 854. 1H NMR
(DMSO-d6)
d
: 1.10 (m, 5H, cyclohexane-H), 1.72 (m, 6H, cyclo-
6.3.14. 4-Fluorobenzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5n)
hexane-H), 2.35 (s, 3H, C2eCH3), 3.13 (d, 2H, J ¼ 6.7 Hz, SCH2), 4.94
(s, 2H, CH2NH), 7.55 (d, 1H, J ¼ 8.3 Hz, quinazolinone 8-H), 7.69 (d,
1H, J ¼ 8.3 Hz, quinazolinone 7-H), 7.96 (s, 1H, quinazolinone 5-H).
Yield 78%, white solid, mp 198.7e199.2 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 95:5). IR (KBr, cmꢀ1
) n: 3442, 3228,

3856
S.-L. Cao et al. / European Journal of Medicinal Chemistry 45 (2010) 3850e3857
3022, 2958, 1663, 1621, 1508, 1384, 1227, 1085, 947, 842 1H NMR
(DMSO-d6) : 2.35 (s, 3H, C2eCH3), 4.53 (s, 2H, SCH2), 4.95 (s, 2H,
CH2NH), 7.14 (m, 2H, PheH), 7.42 (m, 2H, PheH), 7.55 (d, 1H,
J ¼ 8.4 Hz, quinazolinone 8-H), 7.69 (d, 1H, J ¼ 8.4 Hz, quinazolinone
7-H), 7.98 (s, 1H, quinazolinone 5-H). ESI-MS m/z: 374 [M þ H]þ.
Anal. calcd for C18H16FN3OS2$2/3H2O: C, 56.08; H, 4.53; N, 10.90.
Found: C, 55.99; H, 4.38; N, 10.90.
6.3.20. 4-Cyanobenzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5t)
d
Yield 22%, white solid, mp 179.4e180.4 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 95:5). IR (KBr, cmꢀ1
) n: 3440, 3157,
2877, 2232, 1671, 1619, 1488, 1384, 1313, 1074, 933, 830 1H NMR
(DMSO-d6) d: 2.34 (s, 3H, C2eCH3), 4.63 (s, 2H, SCH2), 4.95 (s, 2H,
CH2NH), 7.56 (m, 3H, quinazolinone 8-H and PheH), 7.73 (m, 3H,
quinazolinone 7-H and PheH), 7.96 (s, 1H, quinazolinone 5-H). ESI-
MS m/z: 381 [M þ H]þ. Anal. calcd for C19H16N4OS2$1/2H2O: C,
58.59; H, 4.40; N, 14.38. Found: C, 58.78; H, 4.37; N, 14.34.
6.3.15. 2,4-Difluorobenzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5o)
Yield 63%, white solid, mp 180.9e182.0 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 95:5). IR (KBr, cmꢀ1
)
n
: 3422, 2925, 1657, 1618,
6.3.21. 4-Carboxybenzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5u)
1503,1384,1088, 967, 847.1H NMR (DMSO-d6)
d: 2.35 (s, 3H, C2eCH3),
4.52(s, 2H, SCH2), 4.94(s, 2H, CH2NH), 7.06(m,1H, PheH), 7.23(m,1H,
PheH), 7.53 (s,1H, PheH), 7.55 (d,1H, J ¼ 8.4 Hz, quinazolinone 8-H),
7.69 (d, 1H, J ¼ 8.4 Hz, quinazolinone 7-H), 7.97 (s, 1H, quinazolinone
5-H). ESI-MS m/z: 392 [M þ H]þ. Anal. calcd for C18H15F2N3OS2: C,
55.23; H, 3.86; N, 10.73. Found: C, 55.30; H, 3.87; N, 10.55.
Yield 49%, white solid, mp 221.9e223.5 ꢁC (from dichloro-
methane/methanol ¼ 3:1). IR (KBr, cmꢀ1
)
n
: 3429, 2924, 1680, 1617,
: 2.35 (s, 3H,
1493, 1384, 1315, 1088, 939, 834. 1H NMR (DMSO-d6)
d
C2eCH3), 4.62 (s, 2H, SCH2), 4.96 (s, 2H, CH2NH), 7.52 (m, 3H, qui-
nazolinone 8-H and PheH), 7.69 (d, 1H, J ¼ 8.4 Hz, quinazolinone 7-
H), 7.89 (d, 2H, J ¼ 8.2 Hz, PheH), 7.98 (s, 1H, quinazolinone 5-H).
ESI-MS m/z: 400 [M þ H]þ. Anal. calcd for C19H17N3O3S2$1/2CH2Cl2:
C, 52.99; H, 4.11; N, 9.51. Found: C, 52.90; H, 4.32; N, 9.51.
6.3.16. 2,3,4,5,6-Pentafluorobenzyl N-((2-methyl-4(3H)-
quinazolinon-6-yl)methyl)dithiocarbamate (5p)
Yield 85%, white solid, mp 214.3e215.5 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 95:5). IR (KBr, cmꢀ1
)
n
: 3435, 3231,
6.3.22. 4-Ethoxycarbonylbenzyl N-((2-methyl-4(3H)-quinazolinon-
6-yl)methyl)dithiocarbamate (5v)
2958, 1623, 1619, 1503, 1384, 1127, 988, 855. 1H NMR (DMSO-d6)
d:
2.34 (s, 3H, C2eCH3), 4.65 (s, 2H, SCH2), 4.94 (s, 2H, CH2NH), 7.55 (d,
1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.69 (d, 1H, J ¼ 8.4 Hz, quina-
zolinone 7-H), 7.95 (s, 1H, quinazolinone 5-H). ESI-MS m/z: 466
[M þ H]þ. Anal. calcd for C18H12F5N3OS2$1/2H2O: C, 47.57; H, 2.88;
N, 9.25. Found: C, 47.83; H, 3.02; N, 9.03.
Yield 48%, white solid, mp 202.9e203.2 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 97:3). IR (KBr, cmꢀ1
)
n
: 3450, 3244, 2985, 1717,
16593,1616,1383,1279,1102, 948, 853,711.1HNMR(DMSO-d6)
d
:1.31
(t, 3H, J ¼ 7.1 Hz, CH2CH3), 2.35 (s, 3H, C2eCH3), 4.30 (q, 2H, J ¼ 7.1 Hz,
OCH2CH3), 4.62 (s, 2H, SCH2), 4.95 (s, 2H, CH2NH), 7.53 (m, 3H, qui-
nazolinone 8-H and PheH), 7.68 (d, 1H, J ¼ 8.4 Hz, quinazolinone 7-
H), 7.90 (d, 2H, J ¼ 8.2 Hz, PheH), 7.98 (s,1H, quinazolinone 5-H). ESI-
MS m/z: 428 [M þ H]þ. Anal. calcd for C21H21N3O3S2: C, 58.99; H, 4.95;
N, 9.83. Found: C, 59.42; H, 4.93; N, 9.86.
6.3.17. 4-Chlorobenzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5q)
Yield 78%, white solid, mp 185.7e186.6 ꢁC (CC, eluent:
dichloromethane/methanol ¼ 95:5). IR (KBr, cmꢀ1
) n: 3440, 3230,
2953, 1662, 1619, 1490, 1384, 1323, 1086, 946, 855. 1H NMR (DMSO-
6.3.23. Benzhydryl N-((2-methyl-4(3H)-quinazolinon-6-yl)methyl)
dithiocarbamate (5w)
d6) d: 2.35 (s, 3H, C2eCH3), 4.54 (s, 2H, SCH2), 4.96 (s, 2H, CH2NH),
7.40 (m, 4H, PheH), 7.55 (d, 1H, J ¼ 8.3 Hz, quinazolinone 8-H), 7.69
(d, 1H, J ¼ 8.3 Hz, quinazolinone 7-H), 7.98 (s, 1H, quinazolinone 5-
H). ESI-MS m/z: 390 [M þ H]þ. Anal. calcd for C18H16ClN3OS2$H2O: C,
53.00; H, 4.45; N, 10.30. Found: C, 52.87; H, 4.19; N, 9.99.
Yield 47%, white solid, mp 207.9e208.2 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 95:5). IR (KBr, cmꢀ1
)
n
: 3445, 2928, 1674, 1620,
1509,1387,1324, 934, 704.1H NMR (DMSO-d6)
d: 2.33 (s, 3H, C2eCH3),
4.89(s, 3H, CH2NH), 5.41 (s,1H, SCH), 7.31 (m,10H, PheH), 7.51 (m, 2H,
quinazolinone 8-H and 7-H), 7.93 (s, 1H, quinazolinone 5-H). ESI-MS
m/z: 432 [M þ H]þ. Anal. calcd for C24H21N3OS2: C, 66.79; H, 4.90; N,
9.74. Found: C, 66.64; H, 4.97; N, 9.78.
6.3.18. 4-Bromobenzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5r)
Yield 58%, white solid, mp 186.7e188.5 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 95:5). IR (KBr, cmꢀ1
)
n
: 3442, 3165, 2904, 1673,
6.4. MTT assay
1620, 1486, 1383, 1309, 1071, 933, 831. 1H NMR (DMSO-d6)
d: 2.35 (s,
3H, C2eCH3), 4.52(s, 2H, SCH2), 4.95(s, 2H, CH2NH), 7.36e7.66(m, 6H,
PheH, quinazolinone 8-H and 7-H), 7.97 (s, 1H, quinazolinone 5-H).
ESI-MS m/z: 434 [M þ H]þ. Anal. calcd for C18H16BrN3OS2: C, 49.77; H,
3.71; N, 9.67. Found: C, 50.03; H, 3.86; N, 9.60.
Cytotoxic activity of the synthesized compounds was determined
by MTTassayon the followingcelllines:A549, MCF-7, HeLa, HT29 and
HCT-116. The tested compounds were prepared in DMSO as stock
solutions and stored at ꢀ20 ꢁC. After thawing, the stock solutions
were freshly diluted to a series of concentrationsin medium just prior
to the assay. Final concentration of DMSO in medium was 0.2%.
Cells in logarithmic growth were plated in 96-well plate at
following densities: 7.5 ꢂ103 cells/well for A549, 5 ꢂ103 cells/well
for MCF-7, 1 ꢂ104 cells/well for HeLa, 5 ꢂ103 cells/well for HT29,
6.3.19. 4-Nitrobenzyl N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamate (5s)
Yield 57%, white solid, mp 190e192 ꢁC (CC, eluent: dichloro-
methane/methanol ¼ 98:2). IR (KBr, cmꢀ1
) n: 3445, 3157, 2854,
1668, 1619, 1515, 1382, 1342, 1074, 932, 830. 1H NMR (DMSO-d6)
d:
and 5 ꢂ103 cells/well for HCT-116. 24 h after seeding, 10
mL of 2%
2.33 (s, 3H, C2eCH3), 4.68 (s, 2H, SCH2), 4.94 (s, 2H, CH2NH), 7.53(d,
1H, J ¼ 8.4 Hz, quinazolinone 8-H), 7.65 (m, 3H, quinazolinone 7-H
and PheH), 7.95 (s, 1H, quinazolinone 5-H), 8.16 (d, 2H, J ¼ 8.7 Hz,
PheH). ESI-MS m/z: 401 [M þ H]þ. Anal. calcd for C18H16N4O3S2$1/
2H2O: C, 52.80; H, 4.18; N, 13.68. Found: C, 52.92; H, 4.17; N, 13.44.
DMSO (vehicle control) or solution of the tested compounds was
added into each well and incubated for 72 h, 20 mL of MTT solution
was then added into each well and incubated for an additional 2 h,
and absorbance at 492 nm was measured on a microplate reader.
Cell proliferation inhibition was given by the expression:
h
ꢀ
ꢁ.
i
Cell proliferation inhibition ð%Þ ¼ 1 ꢀ ODcellsþtest compound ꢀ ODblank ðODcellsþDMSO ꢀ ODblankÞ ꢂ 100

S.-L. Cao et al. / European Journal of Medicinal Chemistry 45 (2010) 3850e3857
3857
where OD means the absorbance at 492 nm in the test (ODcells þ test
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