Discovery and optimization of novel dual dithiocarbamates as potent anticancer agents

Article abstract of DOI:10.1016/j.ejmech.2015.02.030

A series of dual dithiocarbamates were synthesized and evaluated for their in-vitro anticancer activities on human non-small cell lung cancer cell line H460. Nine compounds exhibited significant antiproliferative activities with IC₅₀ less than 1 μM. Among them, compound 14m showed the highest inhibitory activity against H460 cell and inhibited the growth of nine types of tumor cells with IC₅₀ values less than 1 μM. It also achieved IC₅₀ of 54 nM and 23 nM against HepG2 and MCF-7 cell lines, respectively. Preliminary structure-activity relationship study indicated that: a) when the methyl group (region A) is substituted with benzene rings, ortho substitution on the benzene ring is favored for activity; b) substitution with heterocyclic structures at region A exhibited greater impact on the anti-tumor activity of compounds, in which pyridine ring, thiazole ring, coumarin and benzo[b]thiophene are favored and quinoline ring is the most favored; c) substitution with different amines (region B) also showed marked effect on the activity of compounds and dimethylamine and morpholine are preferred to other tested amines.

Full text of DOI:10.1016/j.ejmech.2015.02.030

European Journal of Medicinal Chemistry 93 (2015) 381e391
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Discovery and optimization of novel dual dithiocarbamates as potent
anticancer agents
,
Ri-Dong Li a, Hui-Ling Wang a, Ying-Bo Li a b, Zhong-Qing Wang a, Xin Wang a,
,
, *
Yi-Tao Wang b, Ze-Mei Ge a *, Run-Tao Li a
a State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, PR China
b State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, Macao 999078, 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 dual dithiocarbamates were synthesized and evaluated for their in-vitro anticancer activities
on human non-small cell lung cancer cell line H460. Nine compounds exhibited significant anti-
Received 5 October 2014
Received in revised form
27 January 2015
Accepted 19 February 2015
Available online 20 February 2015
proliferative activities with IC50 less than 1
inhibitory activity against H460 cell and inhibited the growth of nine types of tumor cells with IC50
values less than 1 M. It also achieved IC50 of 54 nM and 23 nM against HepG2 and MCF-7 cell lines,
mM. Among them, compound 14m showed the highest
m
respectively. Preliminary structureeactivity relationship study indicated that: a) when the methyl group
(region A) is substituted with benzene rings, ortho substitution on the benzene ring is favored for ac-
tivity; b) substitution with heterocyclic structures at region A exhibited greater impact on the anti-tumor
activity of compounds, in which pyridine ring, thiazole ring, coumarin and benzo[b]thiophene are
favored and quinoline ring is the most favored; c) substitution with different amines (region B) also
showed marked effect on the activity of compounds and dimethylamine and morpholine are preferred to
other tested amines.
Keywords:
Dual dithiocarbamate
Anti-tumor activity
Synthesis
Structureeactivity relationship
© 2015 Elsevier Masson SAS. All rights reserved.
1. Introduction
displayed anti-tumor activity against Ehrlich ascites carcinoma
(EAC) cell line and exhibited potent cytotoxic activity [7]. Yang et al.
Malignant cancer is one of the most life-threatening diseases.
Despite major breakthroughs in many areas of modern medicine,
there still remain significant challenges in the successful treatment
of cancer partly because of the development of drug resistance [1].
Therefore, it is still urgent to discover new anticancer agents of
novel chemical entity. Target-based approaches to drug discovery
are extensively used in the academic and pharmaceutical industry.
However, phenotypic screening is still a very important method for
drug discovery as an original screening paradigm [2].
Dithiocarbamates, a common class of molecular scaffold, have
been attracting considerable interest due to their diverse biological
activities, such as anti-fungal [3], anti-bacterial [4] and carbonic
anhydrase inhibiting activities [5,6]. In particular, some molecules
containing the dithiocarbamate groups as pivotal pharmacophores
exhibited anticancer activity. For examples, Zahran et al. have
revealed a series of novel thalidomide dithiocarbamates (1) which
have described a series of chromone derivatives bearing dithio-
carbamate moieties (2) which displayed apoptosis-inducing effects
on tumor cell lines [8]. Cao's group reported that a series of qui-
nazolinone and 2,4-diaminoquinazoline derivatives with dithio-
carbamate side chains (3,4) showed antiproliferative activity
against human cancer cell lines [9]. In addition, Liu's group
discovered a series of butenolide-containing dithiocarbamates (5),
and several compounds exhibited good anticancer activities [10].
Recently, Liu's group published a novel type of 1,2,3-triazole-
dithiocarbamate hybrids (6) which acted as selective lysine specific
demethylase 1 (LSD1) inactivators (Fig. 1) [11].
In our effort to discover new types of anticancer drugs, we have
identified several novel dithiocarbamate compounds with potent
anticancer activity (7e10, Fig. 2) [12]. To study the structur-
eeactivity relationship of compound 8, we attempted to prepare
compound 8a from the reaction of Mannich base, a precursor of the
Michael acceptor, with pyridin-3-ylmethanamine and carbon di-
sulfide (Scheme 1). However, two products (8a and 11) were ob-
tained and the dual dithiocarbamate 11 was the main product. The
mechanism for the formation of compound 11 could be stepwise
* Corresponding authors.
E-mail addresses: zmge@bjmu.edu.cn (Z.-M. Ge), lirt@bjmu.edu.cn (R.-T. Li).
http://dx.doi.org/10.1016/j.ejmech.2015.02.030
0223-5234/© 2015 Elsevier Masson SAS. All rights reserved.

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R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
Fig. 1. Structures of some dithiocarbamates with anticancer activities.
Fig. 2. Dithiocarbamate derivatives showed appreciable anticancer activities in vitro.
Scheme 1. Synthesis of compound 8a and 11. Reagents and conditions: (a) H2O-i-PrOH, reflux; (b) pyridin-3-ylmethanamine, CS2, TEA, H2O, r.t.
double Michael addition reaction, which was supported by the
isolation of key intermediate 11a (Scheme 2) [13]. To the best of our
knowledge, this type of dual dithiocarbamates has not been syn-
thesized before. More interestingly, these novel dual di-
thiocarbamates showed significant anti-tumor activities in the
preliminary anti-tumor screening. Especially, compound 11
exhibited potent anticancer activities with 92.3%, 42.9%, 87.8%,
88.2%, 88.7%, 88.8% and 95.8% inhibition rates against H460, HepG2,
The SAR study of compound 11 was carried out in two aspects
based on its structure. At first, the methyl group (called region A)
was modified with various substituted aromatic and heterocyclic
rings, and the dimethylamino group (called region B) remained
intact. Afterwards, with the most favored substitution at region A,
region B was modified with different secondary amines (Fig. 3). All
synthesized compounds were evaluated for their activities against
the human non-small cell lung cancer cell line H460.
BGC823, Hela, HCT116, MDA-MB-231 and HL-60 cell lines at 10 mM
in vitro, respectively. Furthermore, the IC50 of compound 11 against
human non-small cell lung cancer (NSCLC) cell line H460 was
2. Results and discussion
0.89
mM. These results demonstrate that compound 11 is a valuable
The modification of region A is outlined in Scheme 3. A variety of
acetyl substituted aromatic or heterocyclic compounds 12ae12t,
lead compound for the development of drugs to treat lung cancer.

R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
383
Scheme 2. Possible mechanism for the formation of compound 11.
Compd.
A
Compd.
A
Compd.
A
Fig. 3. Lead compound 11 and overall strategy for its structural modification.
14a
14h
14o
which were commercially available or prepared by literature pro-
cedures, reacted with paraformaldehyde and dimethylamine hy-
drochloride in refluxing acetic acid to afford the corresponding key
intermediates 13ae13t [13]. Without further purification, com-
pounds 13ae13t reacted with carbon disulfide and dimethylamine
hydrochloride, generating the target compounds 14ae14t. The
preparation of 14ae14j was carried out using water as solvent and
potassium carbonate as base. Since intermediates 13ke13t were
insoluble in water, N,N-dimethylformamide (DMF) was used as
solvent and triethylamine as base for the preparation of com-
pounds 14ke14t.
14b
14c
14i
14j
14p
14q
14d
14k
14r
The synthetic routes for the starting materials 12ne12t are
depicted in Scheme 4. 1-Bromo-2-nitro-4-acetyl-benzene (15) was
treated with aniline and sodium acetate in DMF to obtain com-
pound 16. The nitro moiety of 16 was reduced under catalytic hy-
drogenation conditions to yield compound 17. Benzimidazole 12n
was obtained from the cyclization of 17 with formic acid in 4 N
hydrochloric acid solution [14]. Under basic conditions, but-3-yn-2-
one (18) and 4-phenylbut-3-yn-2-one (19) reacted with 1-
aminopyridinium iodide to provide the compound 12o and 12p,
respectively [15]. Pyridazine (20) was treated with hydroxylamine-
O-sulfonic acid (HOSA) to provide the intermediate 1-
aminopyridazinium salt (21). The subsequent addition of but-3-
yn-2-one (18) and 4-phenylbut-3-yn-2-one (19) in the presence
of a base afforded compound 12q and 12r, respectively [16]. Com-
pound 12s was synthesized by the condensation of 2-
aminopyridine (22) with dimethylformamide dimethylacetal fol-
lowed by treatment with bromoacetone. Compound 12t was ob-
tained by cyclization of 2-aminopyridine (22) with 3-
chloropentane-2,4-dione in refluxing EtOH [17].
14e
14f
14g
14l
14s
14t
14m
14n
Scheme 3. Synthesis of compounds 14ae14t. Reagents and conditions: (a) Me2NH.HCl,
(HCHO)n, AcOH, reflux; (b) Me2NH.HCl, CS2, K2CO3 or Et3N, H2O or DMF.
of activity (14a, IC50 ¼ 1.93
mM). However, the nature and position
of the substituents on the benzene ring greatly affected their ac-
tivity. Introducing an ortho substitution on the benzene ring, such
as 14c (2-F, IC50 ¼ 0.92
mM) and 14d (2-OMe, IC50 ¼ 0.81
mM),
increased the activity to that of compound 14a. On the contrary,
The activities of compounds 14ae14t modified at region A are
shown in Table 1. The results indicated that most of the compounds
showed good activity against H460 cells with IC50 less than 1 mM.
Among them, compounds 14k and 14m showed the highest
inhibitory activity. Replacing the methyl moiety of lead compound
substitution at para-position of the benzene ring led to a 5- to 15-
fold decrease in potency (14f, 4-OBn, IC50 ¼ 9.77
mM; 14g, 4-OH,
IC50 ¼ 30
m
M) comparing with 14a.
The different heterocyclic structures at region A showed great
impact on the anti-tumor activity. The compounds with pyridine
11 (IC50 ¼ 0.89
mM) with benzene ring resulted in slight reduction

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R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
Scheme 4. Synthetic routes for the starting materials 12ne12t. Reagents and conditions: (a) Aniline, AcONa, CuI, DMF; (b) H2, Pd(C), THF, rt; (c) HCOOH, HCl; (c) 1-aminopyridinium
iodide, KOH, K2CO3, DMSO; (d) HOSA, aqueous KHCO3, pH 5.0; (e)3-butyne-2-ones or 4-Phenylbut-3-yn-2-one, KOH, CH2Cl2, rt; (f) (i) Me2NCH(OMe)2, reflux; (ii) bromoacetone,
EtOH; (g) 3-chloropentane-2,4-dione, EtOH, reflux.
ring (14h, IC50 ¼ 0.87
coumarin (14k, IC50
m
¼
M), thiazole ring (14i, IC50 ¼ 0.84
0.66 M), benzo[b]thiophene (14l,
M)
m
M),
cancer), PC3 (prostate cancer), H1299 (lung cancer), A375 (mela-
noma), H460 (human non-small cell lung cancer) and COLO205
(colorectal cancer). The results (Table 3) show that compound 14m
can inhibit the growth of nine types of tumor cells with IC50 less
m
IC50 ¼ 0.85
m
M) and quinoline ring (14m, IC50 ¼ 0.40
m
exhibited similar or more potent activity comparing with the
lead compound 11. In contrast, the activities of the compounds
containing other heterocyclic structures decreased significantly
than 1 mM. It achieved IC50 of 54 nM and 23 nM against cell lines
HepG2 and MCF-7, respectively. In order to investigate whether
these compounds can selectively kill tumor cells and non-tumor
cells, we also chose compound 14m to test its cytotoxicity
against two non-tumor cell lines (human hepatic cell line LO2 and
human embryonic kidney cell line HEK293). The results indicated
that compound 14m exhibited 27-fold and 22-fold decrease of
cytotoxic activity against human hepatic cell line LO2
and some lost activity completely, such as 14n (IC50 ¼ 2.20
14s (IC50 13.03 M), 14o (IC50 31.46 M), 14j
(IC50 ¼ 97.28 M). Compounds 14k and
mM),
¼
m
¼
m
m
M), 14t (IC50 ¼ 151.4
m
14m deserve further investigation due to their excellent activity
and drug-like scaffold.
Based on the optimization results of region A, we used com-
pound 14m as lead structure to study the influence of the amino
moiety with respect to the anticancer activity. We kept 3-quinoline
as region A and varied amino moiety of region B. Compounds
25ae25g were obtained by replacing the dimethylamine moiety of
compound 14m with various secondary amines, including acyclic
and cyclic amines (Scheme 5).
(IC50 ¼ 13.13
m
M) and human embryonic kidney cell line HEK293
M) respectively, comparing with H460 cell line
M), which suggested that compound 14m has
(IC50 ¼ 10.81
(IC50 ¼ 0.476
m
m
obvious selectivity for tumor cells and non-tumor cells. Further
studies on the antitumor mechanisms of 14m are currently
underway.
Table 2 shows the biological activities of those compounds with
modified region B. Most of the amine modifications were unfa-
vorable for the activity. For example, the compounds with bulky
amines, such as diethylamine (25a), pyrrolidine (25d) and piperi-
dine (25e), showed 13-fold, 2-fold and 10-fold decrease of activity
(IC50), respectively. For the six-membered cyclic amine modifica-
tions, the heteroatoms in the cyclic amines also greatly affected the
activity. When modified with N-methyl piperazine (25b) and
morpholine (25c) amines, the activity was increased by 2-fold and
8-fold respectively, comparing with modification by piperidine
amine (25e). However, modification with thiomorpholine amine
3. Conclusion
Systematic structural modification was carried out based on
the structure of the lead compound 11. A series of novel dual
dithiocarbamates were synthesized and they were evaluated for
their in vitro anti-tumor activities. Among them, nine com-
pounds showed significant proliferation inhibition activities on
human non-small cell lung cancer cell line H460 with IC50 less
than 1
m
M. The most potent compound 14m inhibited the growth
M. And
of nine types of tumor cells with IC50 less than 1
m
(25f, IC50 ¼ 124.7
mM) led to the complete loss of activity. The
compound 14m showed IC50 of 54 nM and 23 nM against cell
lines HepG2 and MCF-7, respectively. Our investigation revealed
the following structureeactivity relationships (SAR): a) when
region A is substituted with benzene rings, the ortho substituent
on the benzene ring is favored for the activity; b) the different
heterocyclic structures at region A show great impact on the
anti-tumor activity of compounds, in which pyridine ring, thia-
zole ring, coumarin and benzo [b] thiophene are favored and
quinoline ring is the most favored substituent; c) different
amines at region B showed great effect on the activity of com-
pounds and dimethylamine and morpholine are more favored
than other examined amines. These results provide valuable
compound (25g) derived from N-methyl-1-(pyridin-3-yl)methan-
amine also lost the activity. With regard to the biological activity,
the amino moiety in the dithiocarbamate is very crucial for the
inhibitory activity. The dimethyl amine and morpholine as region B
are more favorable structural features.
To investigate the antiproliferative effect of these novel dual
dithiocarbamates on various tumor cell lines, we selected the most
potent compound 14m to evaluate its IC50 against ten human tu-
mor cell lines including HepG2 (hepatocellular carcinoma), MCF-7
(breast adenocarcinoma), MDA-MB-453 (breast adenocarcinoma),
SW480 (colorectal cancer), H522 (human non-small cell lung

R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
385
Table 1
Antiproliferative activities of compounds 14ae14t modified at region A against
H460 cell line.a
b
b
Compd.
A
IC50
M)
Compd.
A
IC50
(
m
(
m
M)
11
0.89 14k
0.66
Scheme 5. Synthesis of compounds 25ae25g. Reagents and conditions: (a) K2CO3,
acetone; (b) 13m, DMF, H2O.
14a
1.93
14l
0.85
Table 2
Antiproliferative activities of compounds 25ae25g modified at region B against
14b
14c
1.21 14m
0.92 14n
0.40
2.20
H460 cell line.a
14d
14e
0.81 14o
1.24 14p
31.46
4.09
b
b
Compd.
B
IC50
(
m
M) Compd.
B
IC50 (mM)
14m
0.40
5.16
2.51
0.58
25d
0.79
25a
25b
25c
25e
25f
25g
4.65
14f
9.77 14q
4.55
1.00
124.7
14g
30
14r
58.20
14h
14i
0.87 14s
0.84 14t
97.28
13.03
a
H460 cell line: human non-small cell lung cancer.
Doseeresponse curves were determined at five concentrations. The IC50 values
b
are the concentrations needed to inhibit cell growth by 50% as determined from
these curves.
151.4
4. Experiment
4.1. General
14j
All reagents and solvents were purchased from commercial
sources and were used without further purification. Melting points
were determined on X4 microscope and are uncorrected. 1H NMR
and 13C NMR spectra were recorded on Bruker AVANCEⅢ 400 MHz
and 100 MHz spectrometer respectively. Elemental analyses were
performed on a Vario EL Ⅲ (Germany) instrument.
a
H460 cell line: human non-small cell lung cancer.
Doseeresponse curves were determined at five concentrations. The IC50 values
b
are the concentrations needed to inhibit cell growth by 50% as determined from
these curves.
information for the further investigation of this type of potent
anti-tumor compounds.
4.2. The procedure for synthesis of lead compound 11
A mixture of acetone (2.33 g, 40 mmol), paraformaldehyde

386
R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
Table 3
Na2SO4 and concentrated under vacuum to afford the crude prod-
uct. The crude product was purified by recrystallization from the
mixture solvents of ethyl acetate and petroleum ether to give pure
compound (14ae14j).
Antiproliferative activities of compounds 14m against various tumor cell lines and
non-tumor cell lines.a
4.3.1. 2-Benzoylpropane-1,3-diyl bis(dimethylcarbamodithioate)
(14a)
Yield 43%. Pale yellow solid. Mp: 138e139 ꢁC. 1H NMR (400 MHz,
CDCl3):
(m, 2H), 4.43e4.50 (m, 1H), 7.37e7.41 (m, 2H), 7.48e7.52 (m, 2H),
8.00e8.02 (m, 2H); 13C NMR (100 MHz, CDCl3):
d
¼ 3.23 (s, 6H), 3.45 (s, 6H), 3.57e3.62 (m, 2H), 3.67e3.73
d
¼ 38.57, 41.56,
44.49, 45.35, 128.62, 128.98, 133.51, 136.48, 196.40, 201.02; Anal.
Cald for C16H22N2OS4: C, 49.71; H, 5.74; N, 7.25; Found: C, 49.46; H,
5.69; N, 7.09.
b
b
Cell line
IC50
(
m
M)
Cell line
IC50
(m
M)
Cell line
IC50 (mM)
HepG2
MCF-7
MDA-MB-453
SW480
0.054
0.023
0.162
6.350
H522
PC3
H1299
A375
0.139
0.431
0.622
0.439
H460
COLO205
LO2
0.476b
0.325b
13.13c
10.81c
4.3.2. 2-(4-Fluorobenzoyl)propane-1,3-diyl
HEK293
bis(dimethylcarbamodithioate) (14b)
a
Yield 27%. Pale yellow solid. Mp: 133e134 ꢁC. 1H NMR (400 MHz,
HepG2: hepatocellular carcinoma; MCF-7, MDA-MB-453: breast adenocarci-
noma; SW480, COLO205: colorectal cancer; H522, H460: human non-small cell lung
cancer; PC3: prostate cancer; H1299: lung cancer; A375: melanoma; LO2: human
hepatic cell line; HEK293: human embryonic kidney cell line.
Doseeresponse curves were determined at five concentrations. The IC50 values
are the concentrations needed to inhibit cell growth by 50% as determined from
these curves.
Doseeresponse curves were determined at eight concentrations. The IC50 values
are the concentrations needed to inhibit cell growth by 50% as determined from
these curves.
CDCl3):
(m, 2H), 4.50 (s, 1H), 7.11e7.15 (m, 2H), 8.12e8.16 (m, 2H); 13C NMR
(100 MHz, CDCl3):
d
¼ 3.31 (s, 6H), 3.52 (s, 6H), 3.62e3.67 (m, 2H), 3.72e3.77
b
d
¼ 38.52, 41.60, 44.47, 45.39, 115.72 (d,
J ¼ 22 Hz), 131.77 (d, J ¼ 10 Hz), 132.94, 166.08 (d, J ¼ 254 Hz),
196.21, 199.55; Anal. Cald for C16H21FN2OS4: C, 47.50; H, 5.23; N,
6.92; Found: C, 47.75; H, 5.36; N, 7.03.
c
4.3.3. 2-(2-Fluorobenzoyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14c)
Yield 20%. Pale yellow solid. Mp: 97e98 ꢁC. 1H NMR (400 MHz,
(0.72 g, 24 mmol) and dimethylamine hydrochloride (1.63 g,
20 mmol) in H2O (5 mL) and i-PrOH (0.5 mL) was heated under
reflux for 16 h. The reaction mixture was cooled to room temper-
ature and used directly without further purification. A mixture of
dimethylamine hydrochloride (3.27 g, 40 mmol), triethylamine
(8.10 g, 80 mmol) and H2O (30 mL) was stirred for 15 min, and CS2
(3.81 g, 50 mmol) was added dropwise. After stirring 30 min, the
reaction mixture of previous reaction was added. The reaction
mixture was stirred at room temperature for 17 h. The mixture was
extracted with ethyl acetate (20 mL ꢀ 3), the combined organic
phase was washed with water (20 mL ꢀ 2), dried over anhydrous
Na2SO4 and concentrated under vacuum to afford the crude prod-
uct. The crude product was purified by recrystallization from the
mixture solvents of ethyl acetate and petroleum ether to give pure
lead compound 11.
CDCl3):
d
¼ 3.33 (s, 6H), 3.51 (s, 6H), 3.68e3.73 (m, 2H), 3.76e3.81
(m, 2H), 4.32e4.39 (m, 1H), 7.09e7.14 (m, 1H), 7.20e7.27 (m, 1H),
7.49e7.54 (m, 1H), 7.83e7.87 (m, 1H); 13C NMR (100 MHz, CDCl3):
d
¼ 37.99, 41.47, 45.32, 48.95, 116.72 (d, J ¼ 23 Hz), 124.37 (d,
J ¼ 4 Hz), 125.39 (d, J ¼ 11 Hz), 131.08, 134.65 (d, J ¼ 9 Hz), 161.31 (d,
J ¼ 254 Hz),196.26,198.69; Anal. Cald for C16H21FN2OS4: C, 47.50; H,
5.23; N, 6.92; Found: C, 47.42; H, 5.21; N, 6.90.
4.3.4. 2-(2-Methoxybenzoyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14d)
Yield 44%. White solid. Mp: 93e94 ꢁC. 1H NMR (400 MHz,
DMSO-d6):
d
¼ 3.29 (s, 6H), 3.43 (s, 6H), 3.50e3.55 (m, 2H),
3.59e3.64 (m, 2H), 3.82 (s, 3H), 4.31e4.38 (m, 1H), 7.01e7.05 (m,
1H), 7.14 (d, J ¼ 8.40 Hz, 1H), 7.50e7.56 (m, 2H); 13C NMR (100 MHz,
DMSO-d6):
d
¼ 37.70, 41.14, 44.86, 48.38, 55.78, 112.37, 120.50,
4.2.1. 2-Acetylpropane-1,3-diyl bis(dimethylcarbamodithioate) (11)
Yield 20%. While solid. Mp: 98e99 ꢁC. 1H NMR (400 MHz,
126.99, 129.95, 133.96, 157.95, 194.62, 201.55; Anal. Cald for
17H24N2O2S4: C, 49.01; H, 5.81; N, 6.72; Found: C, 49.26; H, 5.85; N,
C
CDCl3):
d
¼ 2.33 (s, 3H), 3.37 (s, 6H), 3.51e3.54 (m, 9H), 3.67e3.72
6.83.
(m, 2H); 13C NMR (100 MHz, CDCl3):
d
¼ 29.99, 37.45, 41.52, 45.43,
50.88, 196.36, 208.58; Anal. Cald for C11H20N2OS4: C, 40.71; H, 6.21;
N, 8.63; Found: C, 40.90; H, 6.34; N, 8.72.
4.3.5. 2-(3-Methoxybenzoyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14e)
Yield 20%. White solid. Mp: 96e98 ꢁC. 1H NMR (400 MHz,
4.3. General procedure for the synthesis of compounds 14ae14j
DMSO-d6):
d
¼ 3.26 (s, 6H), 3.43 (s, 6H), 3.55e3.65 (m, 4H), 3.81 (s,
3H), 4.38e4.45 (m, 1H), 7.23e7.26 (m, 1H), 7.44e7.48 (m, 2H), 7.58
A mixture of the acetyl substituted aromatic or heterocyclic
compound (5 mmol), paraformaldehyde (0.30 g, 10 mmol) and
dimethylamine hydrochloride (0.82 g, 10 mmol) in acetic acid
(10 mL) was heated under reflux for 12 h. The reaction mixture was
concentrated under vacuum to afford the intermediate (13ae13j),
which was used directly without further purification. A mixture of
dimethylamine hydrochloride (0.82 g, 10 mmol), potassium car-
bonate (2.77 g, 20 mmol) and H2O (25 mL) was stirred for 15 min,
and CS2 (1.15 g, 15 mmol) was added dropwise. After stirring
30 min, the intermediate (13ae13j) was added. The reaction
mixture was stirred at room temperature for 12 h. The mixture was
extracted with ethyl acetate (15 mL ꢀ 3), the combined organic
phase was washed with water (15 mL ꢀ 2), dried over anhydrous
(d, J ¼ 7.60 Hz, 1H); 13C NMR (100 MHz, DMSO-d6):
¼ 38.19, 41.48,
d
44.07, 45.16, 55.54, 113.02, 120.07, 121.01, 130.17, 137.68, 159.62,
194.48, 200.39; Anal. Cald for C17H24N2O2S4: C, 49.01; H, 5.81; N,
6.72; Found: C, 48.91; H, 5.95; N, 6.84.
4.3.6. 2-(4-(Benzyloxy)benzoyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14f)
Yield 50%. Pale yellow solid. Mp: 86e88 ꢁC. 1H NMR (400 MHz,
CDCl3):
d
¼ 3.29 (s, 6H), 3.50 (s, 6H), 3.50e3.67 (m, 2H), 3.72e3.77
(m, 2H), 4.43e4.50 (m, 1H), 5.12 (s, 2H), 7.00 (d, J ¼ 8.80 Hz, 2H),
7.33e7.43 (m, 5H), 8.08 (d, J ¼ 9.20 Hz, 2H); 13C NMR (100 MHz,
CDCl3):
d
¼ 38.73, 41.59, 44.05, 45.53, 70.13, 114.65, 127.48, 128.24,
128.69, 129.70, 131.42, 136.16, 163.09, 196.44, 199.32; Anal. Cald for

R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
387
C23H28N2O2S4: C, 56.06; H, 5.73; N, 5.69; Found: C, 56.38; H, 6.07; N,
5.36.
(s, 6H), 3.38 (s, 6H), 3.65e3.75 (m, 4H), 4.42e4.48 (m, 1H),
7.41e7.49 (m, 2H), 7.74e7.78 (m, 1H), 7.95 (d, J ¼ 7.60 Hz, 1H), 8.64
(s, 1H); 13C NMR (100 MHz, DMSO- d6):
d
¼ 37.18, 41.23, 45.08,
4.3.7. 2-(4-Hydroxybenzoyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14g)
47.92, 116.13, 118.14, 123.78, 124.97, 130.73, 134.71, 147.95, 154.61,
158.19, 194.54, 196.91; Anal. Cald for C19H22N2O3S4: C, 50.19; H,
4.88; N, 6.16; Found: C, 50.14; H, 5.04; N, 5.98.
Yield 11%. White solid. Mp: 152e153 ꢁC. 1H NMR (400 MHz,
DMSO-d6):
4.27e4.33 (m, 1H), 6.88 (d, J ¼ 8.80 Hz, 2H), 7.89 (d, J ¼ 8.80 Hz, 2H);
13C NMR (100 MHz, DMSO- d6):
¼ 38.76, 41.77, 43.75, 45.46,
115.79, 128.10, 131.60, 162.09, 194.97, 198.60; Anal. Cald for
16H22N2O2S4: C, 47.73; H, 5.51; N, 6.96; Found: C, 47.87; H, 5.54; N,
d
¼ 3.27 (s, 6H), 3.44 (s, 6H), 3.49e3.62 (m, 4H),
4.4.2. 2-(Benzo[b]thiophene-3-carbonyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14l)
d
Yield 48%. Pale yellow solid. Mp: 118e120 ꢁC (eluent: petroleum
C
ether/ethyl acetate ¼ 6:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.26
6.94.
(s, 6H), 3.43 (s, 6H), 3.67e3.70 (m, 4H), 4.32e4.39 (m, 1H),
7.48e7.55 (m, 2H), 8.09 (d, J ¼ 7.88 Hz, 1H), 863 (d, J ¼ 8.00 Hz, 1H),
4.3.8. 2-Nicotinoylpropane-1,3-diyl bis(dimethylcarbamodithioate)
(14h)
8.98 (s, 1H); 13C NMR (100 MHz, DMSO-d6):
46.16, 122.90, 124.73, 125.53, 125.87, 133.60, 136.13, 139.43, 140.85,
194.36, 195.16; Anal. Cald for C18H22N2OS5: C, 48.83; H, 5.01; N,
6.33; Found: C, 48.78; H, 5.00; N, 6.34.
d
¼ 38.30, 41.30, 45.00,
Yield 25%. Yellow solid. Mp: 112e113 ꢁC. 1H NMR (400 MHz,
CDCl3):
d
¼ 3.23 (s, 6H), 3.43 (s, 6H), 3.57e3.69 (m, 4H), 4.40e4.47
(m, 1H), 7.33e7.36 (m, 1H), 8.28 (d, J ¼ 8.00 Hz, 1H), 8.69e8.70 (m,
1H), 9.15 (d, J ¼ 1.60 Hz, 1H); 13C NMR (100 MHz, CDCl3):
¼ 38.29,
d
41.54, 45.01, 45.38, 123.44, 131.93, 136.16, 150.20, 153.66, 195.75,
200.38; Anal. Cald for C15H21N3OS4: C, 46.48; H, 5.46; N, 10.84;
Found: C, 46.63; H, 5.49; N, 10.96.
4.4.3. 2-(Quinoline-3-carbonyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14m)
Yield 41%. Pale yellow solid. Mp: 130e131 ꢁC (eluent: petroleum
ether/ethyl acetate ¼ 4:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.26
4.3.9. 2-(Thiazole-2-carbonyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14i)
(s, 6H), 3.42 (s, 6H), 3.71e3.79 (m, 4H), 4.55e4.58 (m, 1H),
7.73e7.76 (m, 1H), 7.93e7.97 (m, 1H), 8.11e8.18 (m, 2H), 9.13 (s, 1H),
Yield 15%. Gray solid. Mp: 123e125 ꢁC. 1H NMR (400 MHz,
9.33 (s, 1H); 13C NMR (100 MHz, DMSO-d6):
45.04, 126.29, 127.74, 128.30, 128.76, 129.86, 132.49, 137.96, 148.79,
149.13, 194.24, 199.60; Anal. Cald for C19H23N3OS4: C, 52.14; H, 5.30;
N, 9.60; Found: C, 52.23; H, 5.34; N, 9.72.
d
¼ 37.79, 41.28, 44.89,
DMSO-d6):
4.48e4.54 (m, 1H), 8.18 (d, J ¼ 3.20 Hz, 1H), 8.25 (d, J ¼ 2.80 Hz, 1H);
13C NMR (100 MHz, DMSO-d6):
d
¼ 3.28 (s, 6H), 3.42 (s, 6H), 3.70e3.81 (m, 4H),
d
¼ 37.98, 41.77, 45.56, 46.24,
129.33, 145.76, 165.85, 193.03, 194.47; Anal. Cald for C13H19N3OS5:
C, 39.67; H, 4.87; N, 10.67; Found: C, 39.65; H, 4.91; N, 10.60.
4.4.4. 1-(1-Phenyl-1H-benzo[d]imidazol-5-yl)ethanone (16)
4.3.10. 2-(1H-pyrrole-2-carbonyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14j)
To a solution of 1-(4-bromo-3-nitrophenyl)ethanone (1.23 g,
5 mmol) and aniline (0.56 g, 6 mmol) in DMF (20 mL) was added
sodium acetate (0.82 g, 10 mmol). The mixture was heated 10 h at
120 ꢁC and then cooled to room temperature. H2O (30 mL) was
added to the reaction mixture and the mixture was extracted with
ethyl acetate (15 mL ꢀ 3), the combined organic phase was washed
with brine (15 mL ꢀ 2), dried over anhydrous Na2SO4 and
concentrated under vacuum to afford the crude product. The crude
product was purified by column chromatography (eluent: petro-
leum ether/ethyl acetate ¼ 10:1) to afford 16 as a red solid (1.02 g,
79%). Mp: 104e105 ꢁC (Lit.Mp [18]: 112 ꢁC). 1H NMR (400 MHz,
Yield 34%. Colorless solid. Mp: 169e170 ꢁC. 1H NMR (400 MHz,
DMSO-d6):
3.95e4.02 (m, 1H), 6.22 (d, J ¼ 2.40 Hz, 1H), 7.04 (d, J ¼ 2.80 Hz, 1H),
7.16 (s, 1H); 13C NMR (100 MHz, DMSO-d6):
¼ 38.95, 41.76, 44.63,
45.43, 110.60, 118.34, 127.18, 131.63, 188.93, 195.02; Anal. Cald for
14H21N3OS4: C, 44.77; H, 5.64; N, 11.19; Found: C, 44.87; H, 5.67; N,
d
¼ 3.27 (s, 6H), 3.44 (s, 6H), 3.49e3.61 (m, 4H),
d
C
11.36.
4.4. General procedure for the synthesis of compounds 14ke14t
CDCl3):
d
¼ 2.57 (s, 3H), 7.17 (d, J ¼ 9.08 Hz, 1H), 7.28e7.34 (m, 3H),
A mixture of the acetyl substituted heterocyclic compound
(5 mmol), paraformaldehyde (0.60 g, 20 mmol) and dimethylamine
hydrochloride (0.82 g, 10 mmol) in acetic acid (10 mL) was heated
under reflux for 12 h. The reaction mixture was concentrated under
vacuum to afford the intermediate (13ke13t) which was used
directly without further purification. A mixture of dimethylamine
hydrochloride (0.82 g, 10 mmol), triethylamine (3.04 g, 30 mmol)
and DMF (25 mL) was stirred for 15 min, and CS2 (0.92 g, 12 mmol)
was added dropwise. After stirring 30 min, the intermediate
(13ke13t) was added. The reaction mixture was stirred at room
temperature for 24 h. Water (50 mL) was added and the mixture
was extracted with ethyl acetate (15 mL ꢀ 3), the combined organic
phase was washed with water (15 mL ꢀ 2), dried over anhydrous
Na2SO4 and concentrated under vacuum to afford the crude prod-
uct. The crude product was purified by column chromatography to
afford the pure product (14ke14t).
7.45e7.48 (m, 2H), 7.96 (d, J ¼ 9.04 Hz, 1H), 8.82 (d, J ¼ 1.60 Hz, 1H),
9.85 (s, 1H).
4.4.5. 1-(1-Phenyl-1H-benzo[d]imidazol-5-yl)ethanone (12n)
To a solution of 16 (1.02 g, 4 mmol) dissolved in THF (20 mL) was
added 5% Pd/C (0.21 g). The mixture was stirred at room temper-
ature overnight at 50 psi. The mixture was filtered and the solvent
was evaporated to afford compound 17, which was used in the next
step without further purification. Compound 17 was dissolved in
4 N hydrochloric acid solution (27.5 mL) and formic acid (2 mL), the
reaction mixture was refluxed for 8 h and then cooled to room
temperature. The pH of the reaction mixture was adjusted to 7 with
10% NaOH aqueous solution. The mixture was extracted with
dichloromethane (15 mL ꢀ 3), the combined organic phase was
washed with brine (20 mL ꢀ 2), dried over anhydrous Na2SO4 and
concentrated. The residue was purified by column chromatography
(eluent: petroleum ether/ethyl acetate ¼ 2:1) to afford 12n as a
brown solid (0.33 g, 35%). Mp: 94e96 ꢁC. 1H NMR (400 MHz,
4.4.1. 2-(2-Oxo-2H-chromene-3-carbonyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14k)
Yield 32%. Pale yellow solid. Mp: 121e123 ꢁC (eluent: petroleum
CDCl3):
8.20 (s, 1H), 8.49 (s, 1H).
d
¼ 2.70 (s, 3H), 7.51e7.63 (m, 6H), 8.02 (d, J ¼ 8.56 Hz, 1H),
ether/ethyl acetate ¼ 6:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.30

388
R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
4.4.6. 2-(1-Phenyl-1H-benzo[d]imidazole-5-carbonyl)propane-1,3-
diyl bis(dimethylcarbamodithio-ate) (14n)
4.4.11. 1-(Pyrazolo[1,5-b]pyridazin-3-yl)ethanone (12q)
To solution of hydroxylamine-O-sulfonic acid (4.2 g,
a
Yield 21%. Pale yellow solid. Mp: 73e75 ꢁC (eluent: petroleum
37.5 mmol) neutralized with 2.5 M potassium bicarbonate to pH 5
was added pyridazine (2.0 g, 25 mmol) at 70 ꢁC over 15 min. The
reaction mixture was stirred at 70 ꢁC for 2 h followed by cooling to
room temperature. After neutralization of the reaction mixture to
pH 8, but-3-yn-2-one (0.85 g, 12.5 mmol) in 50 mL of dichloro-
methane and potassium hydroxide (2.6 g, 46.8 mmol) were added,
and the mixture was stirred at room temperature for 24 h. The
reaction mixture was extracted with dichloromethane (20 mL ꢀ 3),
and the combined organic layer was washed with brine
(15 mL ꢀ 2), dried over anhydrous Na2SO4 and concentrated. The
residue was purified by column chromatography (eluent: petro-
leum ether/ethyl acetate ¼ 10:1) to afford 12q as a brown solid
ether/ethyl acetate ¼ 2:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.26
(s, 6H), 3.44 (s, 6H), 3.65e3.66 (m, 4H), 4.54e4.57 (m, 1H),
7.53e7.56 (m, 1H), 7.64e7.73 (m, 5H), 7.79 (d, J ¼ 8.48 Hz, 1H), 8.48
(s, 1H), 8.75 (s, 1H); 13C NMR (100 MHz, DMSO-d6):
d
¼ 38.31, 41.30,
43.76, 44.98, 111.09, 121.54, 123.84, 123.95, 128.22, 130.10, 131.10,
135.38, 136.71, 143.43, 145.65, 194.39, 199.69; Anal. Cald for
C
23H26N4OS4: C, 54.95; H, 5.21; N, 11.14; Found: C, 54.73; H, 5.24; N,
11.04.
4.4.7. 1-(Pyrazolo[1,5-a]pyridin-3-yl)ethanone (12o)
but-3-yn-2-one (0.71 g, 10.4 mmol) and 1-aminopyridinium
iodide (2.69 g, 12 mmol) were dissolved in DMSO (21 mL) and
treated with K2CO3 (1.30 g, 9.36 mmol) and KOH (1.17 g,
20.8 mmol). The mixture was stirred for 6 h at room temperature.
Then the reaction mixture was poured in water (50 mL) and
extracted with ethyl acetate (15 mL ꢀ 3), the combined organic
phase was washed with brine (20 mL ꢀ 2), dried over anhydrous
Na2SO4 and concentrated. The residue was purified by column
chromatography (eluent: petroleum ether/ethyl acetate ¼ 4:1) to
provide 12o as a pale yellow solid (0.53 g, 31%). Mp: 94e96 ꢁC
(1.06 g, 52%). Mp: 171e172 ꢁC. 1H NMR (400 MHz, CDCl3):
d
¼ 2.59
(s, 3H), 7.27e7.33 (m, 1H), 8.43e8.47 (m, 2H), 8.75 (d, J ¼ 8.0 Hz,
1H).
4.4.12. 2-(Pyrazolo[1,5-b]pyridazine-3-carbonyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14q)
Yield 55%. Pale yellow solid. Mp: 168e170 ꢁC (eluent: petroleum
ether/ethyl acetate ¼ 3:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.28
(s, 6H), 3.44 (s, 6H), 3.65e3.69 (m, 4H), 4.14e4.19 (m, 1H), 7.61e7.65
(m, 1H), 8.70e8.74 (m, 2H), 8.80 (s, 1H); 13C NMR (100 MHz, DMSO-
(Lit.Mp [19]: 102e103 ꢁC). 1H NMR (400 MHz, CDCl3):
d
¼ 2.56 (s,
d6):
d
¼ 38.00, 41.31, 45.02, 46.25, 111.76, 121.84, 128.42, 134.41,
3H), 7.01 (t, J ¼ 8.0 Hz, 1H), 7.47 (t, J ¼ 8.0 Hz, 1H), 8.34e8.40 (m,
142.39, 145.21, 193.79, 194.41; Anal. Cald for C16H21N5OS4: C, 44.94;
H, 4.95; N, 16.38; Found: C, 44.97; H, 4.99; N, 16.49.
2H), 8.54 (d, J ¼ 8.0 Hz, 1H).
4.4.8. 2-(Pyrazolo[1,5-a]pyridine-3-carbonyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14o)
4.4.13. 1-(2-Phenylpyrazolo[1,5-b]pyridazin-3-yl)ethanone (12r)
To
a solution of hydroxylamine-O-sulfonic acid (2.72 g,
Yield 57%. Pale yellow solid. Mp: 159e160 ꢁC (eluent: petroleum
24 mmol) neutralized with 2.5 M potassium bicarbonate to pH 5
was added pyridazine (1.28 g, 16 mmol) at 70 ꢁC over 15 min. The
reaction mixture was stirred at 70 ꢁC for 3 h followed by cooling to
room temperature. After neutralization of the reaction mixture to
pH 8, 4-phenylbut-3-yn-2-one (1.15 g, 8 mmol) in 32 mL of
dichloromethane and potassium hydroxide (1.68 g, 30 mmol) were
added, and the mixture was stirred at room temperature for 24 h.
The reaction mixture was extracted with dichloromethane
(15 mL ꢀ 3), and the combined organic layer was washed with
brine (15 mL ꢀ 2), dried over anhydrous Na2SO4 and concentrated.
The residue was purified by column chromatography (eluent: pe-
troleum ether/ethyl acetate ¼ 6:1) to afford 12r as a yellow solid
ether/ethyl acetate ¼ 3:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.29
(s, 6H), 3.46 (s, 6H), 3.61e3.72 (m, 4H), 4.10e4.17 (m, 1H), 7.26 (t,
J ¼ 6.76 Hz, 1H), 7.69e7.31 (m, 1H), 8.31 (d, J ¼ 8.72 Hz, 1H), 8.71 (s,
1H), 8.93 (d, J ¼ 6.80 Hz, 1H); 13C NMR (100 MHz, DMSO- d6):
d
¼ 38.33, 41.29, 44.98, 46.05, 111.69, 115.56, 118.87, 130.00, 130.07,
139.77, 145.06, 192.89, 194.60; Anal. Cald for C17H22N4OS4: C, 47.86;
H, 5.20; N, 13.13; Found: C, 47.81; H, 5.20; N, 13.14.
4.4.9. 1-(2-Phenylpyrazolo[1,5-a]pyridin-3-yl)ethanone (12p)
4-Phenylbut-3-yn-2-one (0.75 g, 5.2 mmol) and 1-
aminopyridinium iodide (1.34 g, 6.04 mmol) were dissolved in
DMSO (10.5 mL) and treated with K2CO3 (0.65 g, 4.68 mmol) and
KOH (0.58 g, 10.4 mmol). The mixture was stirred for 3 h at room
temperature. Then the reaction mixture was poured in water
(50 mL) and extracted with ethyl acetate (15 mL ꢀ 3), the combined
organic phase was washed with brine (15 mL ꢀ 2), dried over
anhydrous Na2SO4 and concentrated. The residue was purified by
column chromatography (eluent: petroleum ether/ethyl
acetate ¼ 10:1) to provide 12p as a yellow solid (0.77 g, 59%). Mp:
90e91 ꢁC (Lit.Mp [19]: 94e95 ꢁC). 1H NMR (400 MHz, CDCl3):
(1.50 g, 79%). Mp: 162e164 ꢁC. 1H NMR (400 MHz, CDCl3):
(s, 3H), 7.29e7.33 (m, 1H), 7.52 (d, J ¼ 2.68 Hz, 3H), 7.63 (s, 2H), 8.47
(d, J ¼ 2.56 Hz, 1H), 8.75e8.77 (m, 1H).
d
¼ 2.19
4.4.14. 2-(2-Phenylpyrazolo[1,5-b]pyridazine-3-carbonyl)propane-
1,3-diyl bis(dimethylcarbamodi-thioate) (14r)
Yield 46%. Off-white solid. Mp: 63e64 ꢁC (eluent: petroleum
ether/ethyl acetate ¼ 2:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.25
(s, 6H), 3.38 (s, 6H), 3.46e3.53 (m, 4H), 3.69e3.74 (m, 1H),
7.43e7.52 (m, 1H), 7.56e7.61 (m, 1H), 8.56e8.58 (m, 1H), 8.72 (s,
d
¼ 2.14 (s, 3H), 7.02 (t, J ¼ 8.0 Hz, 1H), 7.46e7.50 (m, 4H), 7.57e7.60
1H); 13C NMR (100 MHz, DMSO-d6):
d
¼ 37.91, 41.16, 44.99, 47.12,
(m, 2H), 8.44 (d, J ¼ 8.0 Hz, 1H), 8.52 (d, J ¼ 8.0 Hz, 1H).
109.91, 121.17, 128.34, 128.41, 129.20, 129.65, 131.83, 135.90, 144.96,
152.98,194.14,195.10; Anal. Cald for C22H25N5OS4: C, 52.46; H, 5.00;
N, 13.90; Found: C, 52.43; H, 5.03; N, 13.93.
4.4.10. 2-(2-Phenylpyrazolo[1,5-a]pyridine-3-carbonyl)propane-
1,3-diyl bis(dimethylcarbamodith-ioate) (14p)
Yield 61%. Off-white solid. Mp: 118e120 ꢁC (eluent: petroleum
ether/ethyl acetate ¼ 5:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.25
4.4.15. 1-(Imidazo[1,2-a]pyridin-3-yl)ethanone (12s)
(s, 6H), 3.40 (s, 6H), 3.45e3.47 (m, 4H), 3.64e3.70 (m, 1H),
7.22e7.26 (m, 1H), 7.41e7.44 (m, 3H), 7.47e7.55 (m, 2H), 7.65e7.69
(m, 1H), 8.23 (d, J ¼ 8.84 Hz, 1H), 8.89 (d, J ¼ 6.80 Hz, 1H); 13C NMR
2-Aminopyridine (1.88 g, 20 mmol) in 1,1-dimethoxyN,N-
dimethylmethanamine (3.82 g, 32 mmol) was refluxed for 23 h and
then evaporated. The resulting residue was dissolved in EtOH
(35 mL), 1-bromoacetone (3.12 g, 22.75 mmol) was added and the
reaction mixture was stirred at room temperature for 24 h. The
reaction mixture was concentrated and the residue was purified by
(100 MHz, DMSO-d6):
d
¼ 38.33, 41.14, 44.94, 46.43, 109.75, 115.41,
118.90, 128.25, 128.94, 129.48, 129.61, 132.55, 141.47, 155.78, 194.30,
194.38; Anal. Cald for C23H26N4OS4: C, 54.95; H, 5.21; N, 11.14;
Found: C, 54.97; H, 5.30; N, 10.98.
column
chromatography
(eluent:
dichloromethane/

R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
389
methanol ¼ 40:1) to afford 12s as a pale yellow solid (2.06 g, 64%).
Mp: 95e96 ꢁC (Lit.Mp [20]: 98e99 ꢁC). 1H NMR (400 MHz, CDCl3):
9.08 (s, 1H), 9.32 (s, 1H); 13C NMR (100 MHz, DMSO-d6):
12.16, 37.29, 44.62, 46.52, 49.20, 126.30, 127.70, 128.70, 128.76,
129.79, 132.42, 137.80, 148.69, 149.16, 192.84, 200.15; Anal. Cald for
C23H31N3OS4: C, 55.95; H, 6.33; N, 8.51; Found: C, 55.93; H, 6.33; N,
8.52.
d
¼ 11.20,
d
¼ 2.61 (s, 3H), 7.08 (t, J ¼ 6.80 Hz, 1H), 7.50 (t, J ¼ 8.00 Hz,1H), 7.76
(d, J ¼ 9.00 Hz, 1H), 8.34 (s, 1H), 9.65 (d, J ¼ 6.84 Hz, 1H).
4.4.16. 2-(Imidazo[1,2-a]pyridine-3-carbonyl)propane-1,3-diyl
bis(dimethylcarbamodithioate) (14s)
4.5.2. 2-(Quinoline-3-carbonyl)propane-1,3-diyl bis(4-
methylpiperazine-1-carbodithioate) (25b)
Yield 41%. White solid. Mp: 166e167 ꢁC (eluent: petroleum
ether/ethyl acetate ¼ 4:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.27
Yield 17%. White solid. Mp: 133e134 ꢁC (eluent: ethyl acetate/
(s, 6H), 3.43 (s, 6H), 3.67e3.69 (m, 4H), 4.18e4.21 (m, 1H), 7.31e7.35
(m,1H), 7.68e7.72 (m,1H), 7.88 (d, J ¼ 8.44 Hz,1H), 8.59 (s,1H), 9.58
methanol ¼ 20:1). 1H NMR (400 MHz, CDCl3):
d
¼ 2.23 (s, 8H),
2.39e2.40 (m, 6H), 3.78e3.83 (m, 8H), 4.31 (s, 4H), 4.74e4.77 (m,
1H), 7.61e7.64 (m, 1H), 7.82e7.86 (m, 1H), 8.00 (d, J ¼ 7.88 Hz, 1H),
8.15 (d, J ¼ 8.32 Hz,1H), 9.03 (s,1H), 9.47 (s,1H); 13C NMR (100 MHz,
(d, J ¼ 6.72 Hz, 1H); 13C NMR (100 MHz, DMSO-d6):
¼ 38.29, 41.32,
d
44.99, 45.66, 115.94, 117.48, 123.44, 128.29, 130.23, 144.31, 148.67,
188.61, 194.36; Anal. Cald for C17H22N4OS4: C, 47.86; H, 5.20; N,
13.13; Found: C, 47.81; H, 5.12; N, 13.06.
CDCl3):
d
¼ 37.84, 44.69, 45.42, 54.22,126.72,127.40,129.00,129.40,
129.76, 132.12, 138.39, 149.48, 149.88, 195.41, 200.54; Anal. Cald for
25H33N5OS4: C, 54.81; H, 6.07; N, 12.78; Found: C, 54.85; H, 6.09;
C
4.4.17. 1-(2-Methylimidazo[1,2-a]pyridin-3-yl)ethanone (12t)
A mixture of 2-aminopyridine (0.94 g, 10 mmol) and 3-
chloropentane-2,4-dione (1.35 g, 10 mmol) in ethanol (6 mL) was
refluxed for 10 h. After evaporation, the residue was purified by
N, 12.73.
4.5.3. 2-(Quinoline-3-carbonyl)propane-1,3-diyl dimorpholine-4-
carbodithioate (25c)
column
chromatography
(eluent:
dichloromethane/
Yield 15%. White solid. Mp: 124e125 ꢁC (eluent: petroleum
methanol ¼ 50:1) to afford 12t as a pale yellow solid (0.58 g, 33%).
ether/ethyl acetate
¼ 3.31e3.57 (m, 8H), 3.77e3.79 (m, 8H), 4.17 (s, 4H), 4.63e4.66
(m, 1H), 7.73e7.76 (m, 1H), 7.93e7.97 (m, 1H), 8.11e8.17 (m, 2H),
9.09 (s, 1H), 9.33 (s, 1H); 13C NMR (100 MHz, DMSO-d6):
¼
2:1). 1H NMR (400 MHz, DMSO-d6):
Mp: 107e108 ꢁC (Lit.Mp [21]: 108e110 ꢁC). 1H NMR (400 MHz,
d
CDCl3):
d
¼ 2.62 (s, 3H), 2.80 (s, 3H), 7.01 (t, J ¼ 6.68 Hz, 1H), 7.45 (t,
J ¼ 8.00 Hz, 1H), 7.63 (d, J ¼ 8.84 Hz, 1H), 9.74 (d, J ¼ 6.92 Hz, 1H).
d
¼ 37.11,
44.60, 51.32, 65.42, 126.28, 127.78, 128.56, 128.78, 129.87, 132.53,
137.88, 148.79, 149.17, 194.51, 199.76; Anal. Cald for C23H27N3O3S4:
C, 52.95; H, 5.22; N, 8.05; Found: C, 52.98; H, 5.20; N, 8.01.
4.4.18. 2-(2-Methylimidazo[1,2-a]pyridine-3-carbonyl)propane-
1,3-diyl bis(dimethylcarbamodithi-oate) (14t)
Yield 22%. Pale yellow solid. Mp: 128e130 ꢁC (eluent: petroleum
ether/ethyl acetate ¼ 2:1).
1H NMR (400 MHz, DMSO-d6):
d
¼ 2.69 (s, 3H), 3.27 (s, 6H), 3.43
4.5.4. 2-(Quinoline-3-carbonyl)propane-1,3-diyl dipyrrolidine-1-
carbodithioate (25d)
(s, 6H), 3.63e3.73 (m, 4H), 4.23e4.26 (m, 1H), 7.23e7.26 (m, 1H),
7.63e7.67 (m, 1H), 7.74 (d, J ¼ 8.64 Hz, 1H), 9.66 (d, J ¼ 6.64 Hz, 1H);
Yield 35%. Pale yellow solid. Mp: 162e163 ꢁC (eluent: petroleum
13C NMR (100 MHz, DMSO-d6):
115.26, 116.31, 121.21, 128.58, 130.20, 146.48, 153.06, 189.47, 194.36;
Anal. Cald for C18H24N4OS4: C, 49.06; H, 5.49; N, 12.71; Found: C,
48.82; H, 5.55; N, 12.50.
d
¼ 18.32, 38.64, 41.29, 45.02, 45.79,
ether/ethyl acetate ¼ 4:1). 1H NMR (400 MHz, CDCl3):
d
¼ 1.84 (s,
8H), 3.40e3.80 (m, 12H), 4.58 (s, 1H), 7.53 (s, 1H), 7.75 (s, 1H), 7.92
(d, J ¼ 6.92 Hz,1H), 8.05 (d, J ¼ 7.44 Hz,1H), 9.04 (s,1H), 9.38 (s,1H);
13C NMR (100 MHz, CDCl3):
54.95, 126.63, 127.28, 128.66, 129.24, 129.62, 131.98, 138.48, 149.45,
149.74, 191.39, 200.22; Anal. Cald for C23H27N3OS4: C, 56.41; H,
5.56; N, 8.58; Found: C, 56.30; H, 5.51; N, 8.61.
d
¼ 24.08, 25.81, 37.18, 45.44, 50.65,
4.5. General procedure for the synthesis of compounds 25ae25g
A
mixture of 3-acetylquinoline (0.34 g, 2 mmol), para-
formaldehyde (0.24 g, 8 mmol) and dimethylamine hydrochloride
(0.33 g, 4 mmol) in acetic acid (5 mL) was heated under reflux
for12 h. The reaction mixture was concentrated under vacuum to
afford the intermediate 13m which was used directly without
further purification. A mixture of secondary amine (4 mmol), po-
tassium carbonate (1.11 g, 8 mmol) and acetone (10 mL) was stirred
for 15 min, and CS2 (0.37 g, 4.8 mmol) was added dropwise. After
stirring 1 h, the reaction mixture was concentrated to give the in-
termediate (24ae24g). The intermediate (24ae24g) was dissolved
in water (5 ml) and the DMF (5 ml) solution of 13m was added. The
reaction mixture was stirred at room temperature for 12 h. Water
(50 mL) was added and the mixture was extracted with ethyl ac-
etate (15 mL ꢀ 3), the combined organic phase was washed with
water (20 mL ꢀ 2), dried over anhydrous Na2SO4 and concentrated
under vacuum to afford the crude product. The crude product was
purified by column chromatography to obtain the pure product
(25ae25g).
4.5.5. 2-(Quinoline-3-carbonyl)propane-1,3-diyl dipiperidine-1-
carbodithioate (25e)
Yield 15%. Off-white solid. Mp: 110e111 ꢁC (eluent: petroleum
ether/ethyl acetate
¼ 1.48e1.50 (m, 12H), 3.67e3.76 (m, 8H), 4.14 (s, 4H), 4.65e4.68
(m, 1H), 7.72e7.75 (m, 1H), 7.92e7.96 (m, 1H), 8.09e8.15 (m, 2H),
9.05 (s, 1H), 9.31 (s, 1H); 13C NMR (100 MHz, DMSO-d6):
¼
6:1). 1H NMR (400 MHz, DMSO-d6):
d
d
¼ 23.38,
37.35, 44.44, 51.02, 52.52, 126.30, 127.74, 128.75, 128.86, 129.85,
132.46, 137.79, 148.76, 149.16, 192.53, 200.18; Anal. Cald for
C25H31N3OS4: C, 57.99; H, 6.03; N, 8.12; Found: C, 57.96; H, 6.01;
N, 8.13.
4.5.6. 2-(Quinoline-3-carbonyl)propane-1,3-diyl dithiomorpholine-
4-carbodithioate(25f)
Yield 26%. Pale green solid. Mp: 136e137 ꢁC (eluent: petroleum
ether/ethyl acetate
¼
5:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 2.50e2.63 (m, 8H), 3.77 (s, 4H), 4.11e4.12 (m, 4H), 4.45 (s, 4H),
4.5.1. 2-(Quinoline-3-carbonyl)propane-1,3-diyl
bis(diethylcarbamodithioate) (25a)
4.66e4.68 (m, 1H), 7.75 (d, J ¼ 7.04 Hz, 1H), 7.92e7.96 (m, 1H),
8.10e8.16 (m, 2H), 9.07 (s, 1H), 9.32 (s, 1H); 13C NMR (100 MHz,
Yield 24%. White solid. Mp: 103e104 ꢁC (eluent: petroleum
DMSO-d6):
d
¼ 26.46, 37.28, 44.37, 54.12, 126.29, 127.83, 128.65,
ether/ethyl acetate
¼ 1.06e1.12 (m, 12H), 3.63e3.78 (m, 8H), 3.92 (s, 4H), 4.61e4.64
(m, 1H), 7.72e7.75 (m, 1H), 7.72e7.96 (m, 1H), 8.10e8.14 (m, 2H),
¼
8:1). 1H NMR (400 MHz, DMSO-d6):
128.82, 129.84, 132.53, 137.85, 148.76, 149.19, 194.08, 199.91; Anal.
Cald for C23H27N3OS6: C, 49.88; H, 4.91; N, 7.59; Found: C, 49.72; H,
4.89; N, 7.64.
d

390
R.-D. Li et al. / European Journal of Medicinal Chemistry 93 (2015) 381e391
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4.5.7. 2-(Quinoline-3-carbonyl)propane-1,3-diyl
bis(methyl(pyridin-3-ylmethyl)carbamodithioate) (25g)
Yield 18%. Pale yellow solid. Mp: 55e56 ꢁC (eluent: ethyl ace-
tate/methanol
¼
100:1). 1H NMR (400 MHz, DMSO-d6):
d
¼ 3.27e3.44 (m, 7H), 3.80 (s, 4H), 4.68 (s, 1H), 5.03 (s, 1H), 5.32 (s,
2H), 7.16e7.47 (m, 3H), 7.63e7.65 (m,1H), 7.72e7.76 (m,1H), 7.95 (s,
1H), 8.12e8.14 (m, 2H), 8.40e8.53 (m, 4H), 9.10e9.13 (m, 1H),
9.33e9.36 (m,1H); 13C NMR (100 MHz, DMSO-d6):
d
¼ 38.00, 44.61,
54.54, 56.52, 123.54, 126.32, 127.76, 128.37, 128.80, 129.88, 131.39,
132.51, 134.46, 135.00, 137.97, 148.69, 148.83, 149.19, 196.63, 199.64;
Anal. Cald for C29H29N5OS4: C, 58.85; H, 4.94; N, 11.83; Found: C,
58.84; H, 4.97; N, 11.75.
4.6. Antiproliferative activity assays
4.6.1. Antiproliferative activity against tumor cells
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determined by MTT assay on human non-small cell lung cancer cell
line H460 and the antiproliferative activity of the compound 14m
was determined by MTT assay on the following cell lines: HepG2,
MCF-7, MDA-MB-453, SW480, H522, PC3, H1299, A375, H460 and
COLO205. All cells used in the research were prepared at 3.5 ꢀ 103
(b) S.L. Cao, Y.P. Feng, X.L. Zheng, Y.Y. Jiang, M. Zhang, Y. Wang, M. Xu, Syn-
thesis
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substituted-benzylamino-
and
heterocyclylmethylamino-
carbodithioate derivatives of 4(3H)-quinazolinone and their cytotoxic activ-
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thesis and cytotoxic activity of N-((2-methyl-4(3H)-quinazolinon-6-yl)
methyl)dithiocarbamates, Eur. J. Med. Chem. 45 (2010) 3850e3857;
(d) S.L. Cao, Y. Han, C.Z. Yuan, Y. Wang, Z.-K. Xiahou, J. Liao, R.T. Gao, B.B. Mao,
B.L. Zhao, Z.F. Li, X. Xu, Synthesis and antiproliferative activity of 4-
cells/mL concentration and each 100 mL cell suspension was seeded
onto 96-well microtiter plates for 24 h (37 ꢁC, 5% CO2). Then various
appropriate dilutions of tested compounds were added and incu-
bated for 72 h. For the control group, equivalent concentration of
DMSO (final concentration 0.5%) was added. MTT (3-[4,5-
dimethylthiazol-2yl]-diphenyl tetrazolium bromide) method was
used to measure the number of surviving cells and recorded the OD
value at 492/620 nm. The IC50 values were calculated using Prism
Graphpad software of the triplicate experiment.
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derivatives
of
2,4-
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(b) Y.C. Duan, Y.C. Zheng, X.C. Li, M.M. Wang, X.W. Ye, Y.Y. Guan, G.Z. Liu,
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novel 1,2,3-triazole-dithiocarbamate-urea hybrids, Eur. J. Med. Chem. 64
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4.6.2. Antiproliferative activity against non-tumor cells
The antiproliferative activity of compound 14m was determined
by Alamar Blue assay on human hepatic cell line LO2 and human
embryonic kidney cell line HEK293. Two cells used in the research
(c) Y.C. Zheng, Y.C. Duan, J.L. Ma, R.M. Xu, X.L. Zi, W.L. Lv, M.M. Wang, X.W. Ye,
S. Zhu, D. Mobley, Y.Y. Zhu, J.W. Wang, J.F. Li, Z.R. Wang, W. Zhao, H.M. Liu,
Triazoleꢂdithiocarbamate based selective lysine specific demethylase
1
(LSD1) inactivators inhibit gastric cancer cell growth, invasion, and migration,
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mL
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macol. Sin. 36 (2001) 185e187;
cell suspension was seeded onto 96-well microtiter plates for 24 h
(37 ꢁC, 5% CO2). Then various appropriate dilutions of compound
14m were added and incubated for 72 h. For the control group,
equivalent concentration of DMSO (final concentration 0.5%) was
added. Alamar Blue method was used to measure the number of
surviving cells and recorded the OD value at 492/620 nm. The IC50
values were calculated using Prism Graphpad software of the
triplicate experiment.
(b) Z.M. Ge, R.T. Li, T.M. Cheng, J.R. Cui, Synthesis and biological activities of
dipiperazinium salts containing dithiocarboxyl, Arch. Pharm. Pharm. Med.
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zine-1-carbodithioic acid 3-cyano-3,3-diphenyl-propyl esters, Bioorg. Med.
Chem. Lett. 16 (2006) 4214e4219;
(d) X.L. Hou, Z.M. Ge, T.M. Wang, W. Guo, J. Wu, J.R. Cui, C.S. Lai, R.T. Li,
Synthesis and structure-activity relationships of a novel class of dithiocarba-
mic acid esters as anticancer agent, Arch. Pharm. Chem. Life Sci. 11 (2011)
320e332;
Acknowledgments
(e) R.D. Li, X. Zhang, Q.Y. Li, Z.M. Ge, R.T. Li, Novel EGFR inhibitors prepared by
combination of dithiocarbamic acid esters and 4-anilinoquinazolines, Bioorg.
Med. Chem. Lett. 21 (2011) 3637e3640;
The authors thank the National Natural Science Foundation of
China (No. 21172011 and 21372019) and the Macao Science and
Technology Development Fund (077/2011/A3) for financial support.
(f) X. Zhang, R.D. Li, K. Qiao, Z.M. Ge, L.R. Zhang, T.M. Cheng, R.T. Li, Novel
dithiocarbamic
acid
esters
derived
from
6-aminomethyl-
4-
anilinoquinazolines and 6-aminomethyl-4-anilino-3-cyanoquinolines as
potent EGFR inhibitors, Arch. Pharm. Chem. Life Sci. 346 (2013) 44e52;
(g) N. Zhang, W. Guo, L. Wang, W. Huang, B. Xu, Z.M. Ge, M. Li, R.T. Li, J.R. Cui,
Effect of TM208 on QGY-7703 xenograft tumor growth, Anti-Cancer Drugs 19
(2008) 593e598;
(h) Y.B. Li, Z.Q. Wang, X. Yan, M.W. Chen, J.L. Bao, G.S. Wu, Z.M. Ge, D.M. Zhou,
Y.T. Wang, R.T. Li, IC-4, a new irreversible EGFR inhibitor, exhibits prominent
anti-tumor and anti-angiogenesis activities, Cancer Lett. 340 (2013) 88e96.
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2015.02.030.
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