Design, synthesis and antiproliferative activity studies of novel dithiocarbamate–chalcone derivates

Article abstract of DOI:10.1016/j.bmcl.2016.07.012

A series of novel dithiocarbamate–chalcone derivates were designed, synthesized and evaluated for antiproliferative activity against three selected cancer cell lines (EC-109, SK-N-SH and MGC-803). Majority of the synthesized compounds exhibited moderate t

Full text of DOI:10.1016/j.bmcl.2016.07.012

Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design, synthesis and antiproliferative activity studies of novel
dithiocarbamate–chalcone derivates
Dong-Jun Fu y, Sai-Yang Zhang y, Ying-Chao Liu, Li Zhang, Jun-Ju Liu, Jian Song, Ruo-Han Zhao, Feng Li,
⇑
⇑
Hui-Hui Sun, Hong-Min Liu , Yan-Bing Zhang
Collaborative Innovation Center of New Drug Research and Safety Evaluation, School of Pharmaceutical Sciences, Zhengzhou University, Zhengzhou 450001, 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 novel dithiocarbamate–chalcone derivates were designed, synthesized and evaluated for
antiproliferative activity against three selected cancer cell lines (EC-109, SK-N-SH and MGC-803).
Majority of the synthesized compounds exhibited moderate to potent activity against all the cancer cell
lines assayed. Particularly, compounds II2 and II5 exhibited the excellent growth inhibition against SK-N-
Received 18 February 2016
Revised 2 July 2016
Accepted 5 July 2016
Available online xxxx
SH with IC50 values of 2.03 lM and 2.46 lM, respectively. Further mechanism studies revealed that com-
pound II2 could obviously inhibit the proliferation of SK-N-SH cells by inducing apoptosis and arresting
the cell cycle at G0/G1 phase.
Keywords:
Chalcone
Dithiocarbamate
Antiproliferative activity
Apoptosis
Ó 2016 Elsevier Ltd. All rights reserved.
O
Cancer, being one of the leading causes of death globally, poses
a major socioeconomic hazard to humanity at large. Although
there have been progresses in the development of treatment and
prevention of cancer, the successful treatment of cancer remains
a challenge.1 Therefore, there is still an urgent need to search for
novel antiproliferative agents that have broader spectrum of cyto-
toxicity to tumor cells.2
O
O
H
N
O
N
N
N
N
S
N
O
S
N
H
S
S
2
1
Figure 1. Structures of dithiocarbamates as antitumor agents previously reported.
Chalcones are considered to be the precursors of flavonoids and
isoflavonoids3 that have been screened for their wide range of
pharmacological activities as antibacterial,4,5 anti-tumor,6,7 antiin-
flammatory,8,9 antifungal and antioxidant agents.10,11 Dithiocarba-
mate is considered privileged scaffold in drug discovery with a
wide array of biological activities. In the literature, dithiocarba-
mate derivatives have been described as anti-fungal,12 anti-bacte-
rial,13 and carbonic anhydrase inhibitors.14 Besides, the
dithiocarbamate has always been used as a linkage to combine dif-
ferent biologically active scaffold to design new chemical entities.
Our group have reported two series of dithiocarbamates derivates
1 and 2 as antitumor agents, which can inhibit gastric cancer cell
growth, invasion and migration (Fig. 1).15,16
activity of a series of novel N-Acyl homoserine lactones (AHLs) ana-
logs 3 with the chalcone and homoserine lactone scaffold linked by
the dithiocarbamate group through molecular hybridization
approach.18 The preliminary structure–activity relationship (SAR)
studies revealed that the chalcone scaffold and dithiocarbamate
group were critical for their inhibitory activity. So in this study,
these two biologically important groups are retained and we
choose derivative 3 (namely 11a in Ref. 18) as the lead compound
due to its most potent inhibitory activity against MGC-803 cells
than other AHLs analogs. In continuation with our efforts toward
the discovery of novel anticancer agents, we herein design and
optimize chalcone–dithiocarbamate hybrids (Fig. 2).
The synthetic routes towards dithiocarbamate–chalcone ana-
logues (II1–II17) were shown in Scheme 1. Commercially available
substituted benzaldehydes were reacted with substituted ace-
tophenones to form chalcones by Claisen–Schmidt condensation,
which was subjected to etherification reaction with 1,3-dibromo-
propane or 1,2-dibromoethane to afford 5 and IIb. The target ana-
logues were easily obtained in high yields with the mature reaction
Molecular hybridization is a strategy of rational design of new
ligands or prototypes based on the recognition of pharmacophoric
subunits in the molecular structure of two or more known bioac-
tive derivatives.17 In the course of our search for new anticancer
agents, we recently reported the synthesis and antiproliferative
⇑
Corresponding authors.
y
These authors contributed equally.
http://dx.doi.org/10.1016/j.bmcl.2016.07.012
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Fu, D.-J.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.07.012

2
D.-J. Fu et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
O
O
Molecular
hybridization
chalcone
O
O
H
N
S
N
H
Cl
O
Previous work
O
S
O
NH2
3
IC50
:
6.90 1.08 μ M (MGC-803)
24.39 1.52 μ M ( SK-N-SH)
35.47 1.32 μ M (EC- 109)
AHLs
Optimization
This work
O
O
O
S
N
S
O
N
O
II2
IC50
:
7.37 0.96 μ M (MGC-803)
2.03 0.03 μ M (SK-N- SH)
7.43 0.70 μ M (EC-109)
Figure 2. The design and optimization of chalcone–dithiocarbamate derivates.
O
S
S
N
S
O
II14
iii
O
O
O
S
S
S
iii
ii
N
S
O
O
Br
O
O
HO
4
II17
5
n
S
i
O
ii
i
R
R
R
X
Br
X
HO
O
HO
n
IIb
IIa
iii
iii
O
O
S
S
R
N
S
O
X
N
S
O
n
n
R1
II9-II13,II16
II1-II8, II-15
II1 n = 1, R = 4-Cl,
R1 = CH3N
II9 n = 1, R = 4-Cl,
X = C
1 = CH3N
1 = CH3N
II10 n = 1, R = 3,4,5-triOMe, X = C
II2 n = 1, R = 3,4,5-triOMe,
II3 n = 1, R = 3,5-diF,
II4 n = 1, R = 3,5-diCl,
II5 n = 0, R = 3,4,5-triOMe,
II6 n = 1, R = 4-Cl,
R
R
II11 n = 1, R = 3,5-diF,
II12 n = 0, R = 4-Cl,
II13 n = 1, R = H
X = C
X = C
X = N
X = C
R1 = CH3N
R
1 = CH3N
II16 n = 0, R = 3,4,5-triOMe
R1 = CH2
R1 = CH2
II7 n = 0, R = 4-Cl,
II8 n = 0, R = 3,4,5-triOMe, R1 = CH2
II15 n = 0, R = 4-Cl,
II18 n = 1, R = 4-OCH3,
R1 = O
R1 = CH3N
Scheme 1. Synthesis of analogues II1–II17. Reagents and conditions: (i) substituted aromatic aldehyde, NaOH, EtOH, rt, 60–83% yield; (ii) 1,3-dibromopropane or 1,2-
dibromoethane, K2CO3, THF, reflux, 72–79% yield; (iii) CS2, substituted amine, Na3PO4ꢀ12H2O, acetone, rt, 63–76% yield.
conditions developed by our group.18 Finally, all the chalcone–
dithiocarbamate derivatives were fully characterized by 1H NMR,
13C NMR and HRMS.
All synthesized compounds were evaluated for their antiprolif-
erative activity against three human cancer cell lines (MGC-803,
EC-109, SK-N-SH) using MTT assay method and compared with
the well-known anticancer drug 5-fluorouracil and lead compound
AHLs derivative 3 (Table 1). The majority of the synthesized com-
pounds exhibited moderate to potent activity against all the cancer
cell lines. Particularly, compounds II2 and II5 exhibited excellent
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D.-J. Fu et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
3
Table 1
Antiproliferative activity of chalcone–dithiocarbamate derivatives
a
Compound
IC50
(lM)
EC-109
SK-N-SH
MGC-803
II1
II2
II3
II4
II5
II6
II7
II8
34.19 0.68
7.43 0.70
14.50 0.56
12.43 1.07
10.02 0.52
42.61 1.26
26.12 1.14
38.90 1.31
18.61 0.36
11.12 1.27
28.78 0.30
27.81 0.98
29.82 0.28
36.38 1.22
15.42 0.96
10.71 0.81
18.25 1.08
>100
31.88 1.31
2.03 0.03
13.20 0.72
24.62 0.92
2.46 0.02
29.36 0.72
7.37 0.96
16.26 0.78
19.36 1.27
8.28 0.36
56.82 1.02
12.80 1.69
41.33 0.78
34. 54 0.77
14.45 0.69
28.37 0.81
47.93 1.29
19.48 0.48
41.32 1.99
11.46 0.30
31.66 0.58
32.36 0.35
19.44 1.09
6.90 1.08
32.23 2.13
27.38 0.48
48.36 0.91
19.05 1.17
12.78 0.91
36.12 0.90
18.34 1.37
24.46 0.52
38.17 1.34
11.74 0.42
12.12 1.62
19.08 0.67
29.54 1.53
24.39 1.52
9.84 0.75
II9
II10
II11
II12
II13
II14
II15
II16
II17
II18
318
35.47 1.32
10.30 0.83
5-FU
7.21 1.04
a
Antiproliferative activity was assayed by exposure for 72 h to substances and
expressed as concentration required to inhibit tumor cell proliferation by 50% (IC50).
Data are presented as the means SDs from the dose–response curves of three
independent experiments.
Reduction of the carbon linker length by one
carbon without obvious change for activity
3,4,5-trimethoxy is better
than p-Cl or 3,5-diCl
O
O
O
S
N
N
S
O
N
O
3,4,5-trimethoxyphenyl ring
is better than thiofuran
N
N
or
>
N
Good selectivity between cancer and normal cells
Scheme 2. Summary of SAR of chalcone–dithiocarbamate derivatives.
growth inhibition activity against SK-N-SH with IC50 values of
2.03 lM and 2.46 lM, respectively.
Compounds II1–II4, II9–II11 and II18 with different sub-
stituents on the aryl ring were synthesized and evaluated for their
cytotoxicity. Compound II2 with a 3,4,5-trimethoxy group showed
the most potent activity against SK-N-SH cells, with an IC50 of
2.03 lM, which is 15 times higher than compound II1 with a halo-
Figure 3. Compound II2 induced apoptosis in SK-N-SH cells. (A) Apoptosis analysis
with Hoechst-33258 staining after 24 h, 48 h treatment of II2 in SK-N-SH cells. (B
and C) Quantitative analysis of apoptotic cells using annexin V-FITC/PI double
staining and flow-cytometry calculation. (*) P <0.05 was considered statistically
highly significant. (##) P <0.01 was considered statistically highly significant. Data
are the mean SD. All experiments were carried out at least three times.
gen atom Cl on the phenyl ring. In addition, compound II10 with a
3,4,5-trimethoxy group was better than compound II9 with a 4-Cl
and compound II11 with a 3,5-diF for the inhibitory activity
against SK-N-SH cells. For SK-N-SH cells and MGC-803 cells, ana-
logs II3 substituted by 3,5-diF exhibited more potent antiprolifer-
ative activity than II4 substituted by 3,5-diCl. Meanwhile,
replacing 3,4,5-trimethoxy group with electron-donating group
4-methoxy led to a complete loss of potency against EC-109 cells
(II2 vs II18). From the biological data of compounds II1–II4, II9–
II11, and II18, we can conclude that the substituent on the phenyl
group of chalcone had a remarkable effect on their cytotoxic
activity.
activity. However, replacing the phenyl scaffold of compound
II11 with a pyridine ring of II13 led to an increase of activity.
The results revealed that the aromatic rings of chalcones had a pro-
found effect on the activity. Structure activity relationship analysis
showed that different dithiocarbamates effected the antitumor
activity, for example, dithiocarbamate contained 1-methylpiper-
azine of compound II2 was more active than dithiocarbamate con-
tained pyrrolidine of compound II10 against SK-N-SH cells. Based
on the above findings, we further tested whether the length of
The importance of the aromatic ring of chalcone scaffold in the
anticancer activity was also explored. Replacing the phenyl scaf-
fold of compound II16 with thiophene ring of II17 led to a loss of
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D.-J. Fu et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
Figure 4. Effect of compound II2 on the cell cycle distribution of SK-N-SH cells. Cells were treated with different concentrations (2.5 lM, 5 lM) for 24 h or 48 h. Then the cells
were fixed and stained with PI to analyze DNA content by flow cytometry. (A) Incubated for 24 h; (B) incubated for 48 h. The experiments were performed three times, and a
representative experiment is shown.
cer cell line (MGC-803) and a normal cell line (GES-1). The detailed
illustration for structure activity relationship of target derivatives
was showed in Scheme 2.
To explore cytotoxicity of II2 in SK-N-SH, cell apoptosis was
investigated with Hoechst 33258 staining. After 24 h and 48 h
incubation with II2 at indicated concentration, characteristic apop-
totic morphological changes were observed by fluorescence micro-
scope, including cell rounding, chromatin shrinkage, and formation
of apoptotic bodies (Fig. 3A). The apoptotic analysis was also per-
formed with annexin V-FITC/PI double staining and quantitated
by flow cytometry. Compound II2 treatment of SK-N-SH cells time
dependently increased the percentage of the apoptotic population
up to 20.6% and 36.8%, respectively, compared to control
(Fig. 3B and C).
To have a better understanding of the mechanism of action of
cytotoxic activity of compound II2, a cell-cycle cytotoxicity assay
was performed by treating SK-N-SH cells with different concentra-
tions with compound II2 (2.5 lM, 5 lM). After treatment SK-N-SH
Figure 5. Expression of p53, pro-caspases 3 and Active-caspase 3 in SK-N-SH cells
after treatment by compound II2 for 48 h.
cells for 24 h, it was observed that the percentage of cells in G0/G1
phase at different concentrations were 50.03% and 68.19%, respec-
tively (Fig. 4A), whereas treatment for 48 h, the percentage of cells
in G0/G1 phase were 59.64% and 74.85%, respectively (Fig. 4B). The
results suggested that II2 caused an obvious G0/G1 arrest pattern
in a concentration- and time-dependent manner with a concomi-
tant decrease in terms of the number of cells in other phases of
the cell cycle. Detailed activity evaluation and apoptosis mecha-
nism studies are currently underway in our laboratory.
carbon linker between the dithiocarbamates and chalcones affect
the anticancer activity. With reduction of the carbon tether length
by one carbon (II2 vs II5, II6 vs II7, II9 vs II12, II10 vs II16), an obvi-
ous increase of potency was not observed.
Compounds II2, II5 were further examined for possible cytotox-
icity against GES-1 (normal human gastric epithelial cell line). We
found that compounds II2, II5 exhibited no cytotoxicity against
At the same time, we also evaluated the level of p53, caspases 3
and pro-caspase 3 (inactive form of apoptotic effector caspase 3) in
SK-N-SH cells (Fig. 5). Cleavage of pro-caspases 3 results in the pro-
duction of an active effector (caspase 3), which can cleave essential
GES-1 (>64
II5 exhibited potent cytotoxicity against the selected MGC-803 cell
line (2.03 M, 2.46 M, respectively). The results indicated that
compounds II2, II5 had good selectivity between the selected can-
lM, >64 lM, respectively). However, compounds II2,
l
l
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D.-J. Fu et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
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This work was supported by the National Natural Sciences
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Please cite this article in press as: Fu, D.-J.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.07.012