Dithiocarbamic acid esters as anticancer agent. Part 1: 4-Substituted-piperazine-1-carbodithioic acid 3-cyano-3,3-diphenyl-propyl esters

Article abstract of DOI:10.1016/j.bmcl.2006.05.085

A variety of 4-N atom substituted derivatives were synthesized and evaluated for their in vitro anticancer activities using 4-methylpiperazine-1-carbodithioic acid 3-cyano-3,3-diphenyl-propyl ester 4 as lead compound. Among them, compound 6a without any substituent on 4-N atom (R¹ = H) was found to be the most active anticancer agent with IC₅₀ = 5.3 μM against HL-60 and IC₅₀ = 11.5 μM against Bel-7402, respectively. Increase in the polarity and/or introduction of suitable acyl groups at the 4-N atom of the lead compound 4 are favorable for the improvement of activity.

Full text of DOI:10.1016/j.bmcl.2006.05.085

Bioorganic & Medicinal Chemistry Letters 16 (2006) 4214–4219
Dithiocarbamic acid esters as anticancer agent.
Part 1: 4-Substituted-piperazine-1-carbodithioic acid
3
a
-cyano-3,3-diphenyl-propyl esters
a
b
c
c,
*
Xueling Hou, Zemei Ge, Tingmin Wang, Wei Guo, Jingrong Cui,
a
b
a,c,
*
Tieming Cheng, Chingshan Lai and Runtao Li
a
School of Pharmaceutical Sciences, Peking University, Beijing 100083, China
b
Medinox. Inc., San Diego, CA 92121, USA
The State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences,
Peking University, Beijing 100083, China
c
Received 4 January 2006; revised 10 May 2006; accepted 29 May 2006
Available online 12 June 2006
Abstract—A variety of 4-N atom substituted derivatives were synthesized and evaluated for their in vitro anticancer activities using
-methylpiperazine-1-carbodithioic acid 3-cyano-3,3-diphenyl-propyl ester 4 as lead compound. Among them, compound 6a with-
4
1
out any substituent on 4-N atom (R = H) was found to be the most active anticancer agent with IC50 = 5.3 lM against HL-60 and
IC50 = 11.5 lM against Bel-7402, respectively. Increase in the polarity and/or introduction of suitable acyl groups at the 4-N atom of
the lead compound 4 are favorable for the improvement of activity.
ꢀ
2006 Elsevier Ltd. All rights reserved.
7
–9
Despite major breakthroughs in many areas of modern
medicine over the past 100 years, the successful treat-
ment of cancer remains a significant challenge at the
start of the 21st century. Because it is difficult to discov-
er novel agents that selectively kill tumor cells or inhibit
their proliferation without the general toxicity, the use
of traditional cancer chemotherapy is still very limited.
agents.
Another group from Italy also found that
the metal complex of dithiocarbamic acid esters exhibit-
ed anticancer activity. For example, the platinum com-
plexes have similar activity but less toxicity than the
1
0–12
cisplatin.
However, little systematic research has
been reported about anticancer activity of this class of
compounds, although compound RWJ-025856 (3) was
unexpectedly found to have attenuating effects on tumor
necrosis factor a (TNFa)-induced apoptosis in murine
Dithiocarbamic acid ester is a common class of organic
molecules. They exhibit a variety of valuable biological
1
3
fibrosarcoma WEHI 164 cells.
1
,2
effects, including antibacterial activity,
3
antifungal
4,5
activity, the ability to chelate heavy metals, and to
6
function as NO scavengers. Recently, it was found by
We have developed a convenient one-pot method for the
synthesis of dithiocarbamic acid ester and have prepared
Hirschelman’s group that (4-methanesulfinyl-butyl)-
dithiocarbamic acid methyl ester (sulforamate 1,
Fig. 1) and 5-oxohexyl dithiocarbamic acid methyl ester
O
S
S
O
C
S
R'
N
S
N
S
H
H
(oxomate 2, Fig. 1) are potent phase II enzyme inducers
which could be used as cancer chemopreventive
sulforamate (1)
oxomate (2)
S
S
NC
N N
Cl
Keywords: Dithiocarbamic acid ester; Anticancer activity; 4-Substitut-
ed-piperazine-1-carbodithioic acid 3-cyano-3,3-diphenyl-propyl ester;
Synthesis.
S
NMe2 H C N
3
N
S
*
Corresponding authors. Tel.: +86 10 8280 2467 (J.C.); tel.: +86 10
280 1504; fax: +86 10 8271 6956 (R.L.); e-mail addresses:
jrcui@bjmu.edu.cn; lirt@mail.bjmu.edu.cn
RWJ-025856(3)
990207(4)
8
Figure 1. Structures of compounds 1–4.
0
960-894X/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.05.085

X. Hou et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4214–4219
4–16
4215
1
1
many compounds using this method.
Random
(R = H) with methylsulfonyl chloride and various
substituted phenylsulfonyl chloride under basic
conditions afforded 4-methylsulfonylpiperazine-1-carbo-
dithioic acid 3-cyano-3,3-diphenyl-propyl ester 7a and
4-phenylsulfonylpiperazine-1-carbodithioic acids 3-cya-
no-3,3-diphenyl-propyl esters 7b–7i. The hydroxylethyl
screening of those compounds led to the discovery of
several compounds possessing significant antitumor
1
7,18
activity.
One of the best compounds is 4-methyl-pi-
perazine-1-carbodithioic acid 3-cyano-3,3-diphenyl-pro-
pyl ester (4, Fig. 1) with 79% and 75% inhibition rates
against HL-60 and Bel-7402 cell lines at 33 lM in vitro,
respectively. A further in vivo test of its hydrochloride
salt (4ÆHCl), which has better solubility, indicated that
the inhibition rates against tumor growth of sarcoma
180 (S180), hepatocyte carcinoma 22 (H ), and implant-
22
ed human gastric carcinoma in nude mice were 46.4–
9.6% (P < 0.01), 39.3–51.6% (P < 0.05 ꢀ 0.01), and
8.1–59.0% (P < 0.01) at different doses from 50 to
00 mg/kg, respectively. More importantly, 4ÆHCl
1
analgoue 6e (R = HOCH CH –) was reacted with thio-
2
2
nyl chloride in dichloromethane at room temperature to
give the corresponding chloride product 8. The direct
N-acylation of 6a with b-chloropropionyl chloride
provided the b-chloropropionyl derivative 9. Using b-
bromopropionyl chloride under identical conditions
gave the desired b-bromopropionyl derivative 10. The
compound 11 was synthesized from the reaction of 10
5
1
2
2
1
with silver methanesulfonate in refluxing acetonitrile
for 4.5 h in 58% yield.
showed very low toxicity. Taking it orally at a dose of
1
0 g/kg continuously for 10 days, the rats are neither
dead nor damage of organs observed by visual examina-
tion. Furthermore, the body weight of tested group is
similar to that of control group.
All of the synthesized compounds were screened for the
preliminary in vitro anticancer activity against six differ-
ent cell lines: a human promyelocyticfina leukemic cell
line (HL-60), a human hepatocellular carcinoma cell line
(Bel-7402), a human gastric carcinoma cell line (BGC-
823), a human cervical carcinoma cell line (HeLa), a hu-
man prostatic carcinoma cell line (PC 3MIE8), and a
human breast carcinoma cell line (MDA-MB-435), at
four different concentrations. Because most of com-
pounds showed inhibition activity only on HL-60 and
Bel-7402 cells at concentration of >33 lM, we herein
merely listed biological results at concentration of
1
9
To the best of our knowledge, dithiocarbamic acid ester
represents a new kind of compound with a novel struc-
4
ture, significant anticancer activity, and very low toxici-
ty. Encouraged by this result, we selected compound 4
as a lead compound to further explore the structure–
activity relationships (SAR) with the aim of optimizing
potency and anticancer activity. In this article, we focus
our attention on the optimization of substituents at the
2
2–24
4
-N atom of piperazine in the lead compound 4.
33 lM in Table 1 in order to discuss the SAR.
Meantime, the IC50 values for the most potent com-
pounds, 4, 6a, and 6e were tested.
The synthetic routes of all the target compounds are
shown in Scheme 1. According to our improved and
2
0
well-established method, a variety of 1-N-substituted
piperazines, which were commercially available or pre-
pared by standard methods, were reacted with carbon
disulfide and 3-cyano-3,3-diphenyl-propyl bromide 5 in
the presence of anhydrous potassium phosphate at room
temperature to afford the corresponding dithiocarbamic
acid esters 6a–6y in high yields. Direct sulfonation of 6a
As seen in Table 1, in order to obtain the diversity of 4-
N substituted derivatives of 4, substituents such as alkyl,
aryl, heteroaryl, sulfonyl benzyl, and benzoyl groups
were chosen to replace the 4-N methyl group of com-
pound 4. Although most of the compounds showed
weak or no activity, it is interesting to note that the com-
1
pound 6a without a substituent on 4-N atom (R = H)
S
NC
S
NC
O
O S N
N
S
N
N
S
Cl
R2
1
1
7
a-7i (from 6a, R =H)
8 (from 6e, R =CH CH OH)
2
2
b
c
S
NC
NC
a
R1
N
R1
NH + CS2 +
Br
N
N
S
5
6a-6y
d
O
S
O
S
NC
O O
S
NC
e
O
N
N
S
X
N
N
S
1
11 (from 10)
9 (X=Cl, from 6a, R =H)
1
1
0 (X=Br, from 6a, R =H)
2
Scheme 1. Reagents and conditions: (a) K
3
PO
4
, acetone, rt; (b) R SO
SO
2
Cl, Et N, CH
3
2 2 2 2 2 2 2
Cl , rt; (c) SOCl , CH Cl , reflux, 2 h, 79%; (d) XCH CH COCl,
NaHCO , CH Cl , reflux, 0.5 h, 57% for 9, 73% for 10; (e) CH
2
3
2
3
3
Ag, CH CN, reflux, 4.5 h, 58%.
3

4216
X. Hou et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4214–4219
À5
Table 1. The biological activities of compounds 6a–y, 7a–i, and 8–11 at a concentration of 3.3 · 10 lM
S
NC
S
NC
O
R1
N
N
S
O S N
N
S
R2
6
a-6y, 8-11
7a-7i
1
R or R
2
Compound
Inhibition rate against HL-60 (%)
Inhibition rate against Bel-7402 (%)
a
79.0 (9.9 )
a
75.2 (27.1 )
4
CH
H–
3
–
a
98.7 (5.3 )
a
95.5 (11.5 )
6a
6b
6c
6d
6e
C
2
H
5
–
À7.81
À13.5
2.57
4.97
11.0
12.7
n-C
n-C
4
H
9
–
6
H
13
–
a
a
HO(CH
2
)
2
–
92.5 (24.5 )
57.7 (11.7 )
6
6
6
6
6
6
6
6
6
6
6
6
6
6
6
6
6
f
10.5
0
0
g
h
i
4.3
15.9
26.3
0
F
F
0
F3C
8.5
0
j
Cl
k
l
4.2
0.7
0
0
Cl
Cl
Cl
23.3
14.6
0
Me
m
n
o
p
q
r
Cl
4.8
8.5
9.8
0
OMe
MeO
MeO
14.9
15.5
0
Me
O2N
7.8
31.8
24.2
12.5
0
2.2
22.6
33.2
4.7
16.6
s
N
N
N
t
u
v
C
6
H
5 2
CH –
O
O
(
continued on next page)

X. Hou et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4214–4219
4217
Table 1 (continued)
1
R or R
2
Compound
Inhibition rate against HL-60 (%)
2.7
Inhibition rate against Bel-7402 (%)
6
6
6
7
7
7
7
7
7
7
7
7
8
9
1
1
w
x
y
a
b
c
d
e
f
18.0
Cl
O
O
30.4
0
17.7
35.0
13.4
0
O
O
H3C
24.9
0
MeO
O2N
1.5
0
5.4
0.2
5.5
0
0
O2N
0
NO2
Cl
g
h
i
0
0
Cl
0
0
F3C
F
0
9.7
28.7
22.0
23.4
1.09
ClCH
2
CH
2
–
53.6
43.3
82.2
2 2
Cl(CH ) CO–
0
1
2 2
Br(CH ) CO–
MeSO CH
3
2
CH
2
CO–
0
a
IC50 value.
exhibited better activities (98.7% against HL-60,
IC50 = 5.3 lM; 95.5% against Bel-7402, IC50 = 11.5 lM)
than those of the lead compound 4 (R = Me, 79.0%
against HL-60, IC50 = 9.9 lM; 75.2% against Bel-7402,
IC50 = 27.1 lM). This result demonstrates that the
introduction of bulky substituent is unfavorable for
the improvement of activity.
differences between their structures, their activities were
completely different. Compound 6b showed no activity,
while 6e showed inhibition rates of 92.5% against HL-60
and 57.7% against Bel-7402. Based on the above results
and the activity data of compound 6a, we consider that
the polarity of the molecule might be a very important
factor affecting the activity.
1
1
Comparing compound 6b (R = –C H ) with 6e (R =
1
1
Introduction of heteroaryl groups (6s, R = pyridyl; 6t,
2
5
1
R = pyrimidinyl) led to a significant decrease of activity
–
CH CH OH), it was found that despite only slight
2
2

4218
X. Hou et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4214–4219
compared with the lead compound 4. However, their
activities are obviously higher than those of the aryl
substituted derivatives (6f–6r). This result suggested that
it is possible to improve the activity by introducing suit-
able heteroaryl groups into the 4-N atom of the lead
compound 4. Furthermore, the activity improvement
of 6s and 6t may be related to the increase in polarity
of molecule caused by introduction of heteroaryl group.
References and notes
1. Aboul-Fadl, T.; El-Shorbagi, A. Eur. J. Med. Chem. 1996,
3
. Imamura, H.; Ohtake, N.; Jona, H.; Shimizu, A.; Moriya,
M.; Sato, H.; Sugimoto, Y.; Ikeura, C.; Kiyonaga, H.;
Nakano, M.; Hagano, R.; Abe, S.; Yamada, K.; Hashiz-
ume, T.; Morishima, H. Bioorg. Med. Chem. Lett. 2000,
1, 165.
2
1
0, 109.
1
The activity results of compounds 6x (R = furoyl,
3
. Len, C.; Boulognemerlot, A. S.; Posted, D. J. Agric. Food
Chem. 1996, 44, 2856.
4. Hidaka, S.; Funakoshi, T.; Shimada, H.; Tsuruoka, M.;
1
0.4% against HL-60) and 6y (R = ethoxycarbonyl,
5.0% against Bel-7402) also support the above
3
3
suggestions.
Kojima, S. J. Appl. Toxicol. 1995, 15, 267.
. Tandon, S. K.; Singh, S.; Jain, V. K.; Prasad, S. Fundam.
Appl. Toxicol. 1996, 31, 141.
5
Because the furoyl substituted analogue 6x showed
better activity than other substituted analogues, we
speculate that replacement of the 4-N-Me of the lead
compound 4 by acyl groups with anticancer action
would improve the pharmacological properties. There-
fore, 3-chloropropionyl, 3-bromopropionyl, and 3-
methanesulfonyloxy-propionyl were selected as substit-
uents to obtain the compounds 9–11. At the same
time, as the analogue of compound 6e, the chloroethyl
substituted compound 8 was also prepared. As shown
in Table 1, except for compound 11, all of the com-
pounds 8–10 exhibited higher activities than that of
the compound 6x. Most importantly, the 3-bromopro-
pionyl derivative 10 showed 82.2% inhibition rate
against HL-60. This result demonstrates that it is pos-
sible to improve the anticancer activity by introducing
suitable acyl groups at the 4-N atom of the lead com-
pound 4.
6
7
. Lai, C. S.; Vassilev, V. P.; Wang, T. M. US 5847004.
. Hirschelman, W. H.; Song, L. S.; Park, E. J.; Tan, Y.; Yu,
R.; Hawthorne, M.; Mehta, R. G.; Grubbs, C. J.; Lubet,
R. A.; Moriarty, R. M.; Pezzuto, J. M. 224th ACS
National Meeting: Division of Medicinal Chemistry, 2002,
9
8.
8
9
. Hirschelman, W. H.; Kosmeder, II J. W.; Song, L. S.;
Park, E. J.; Moriarty, R. M.; Pezzuto, J. M. 224th ACS
National Meeting: Division of Medicinal Chemistry, 2002,
178.
. Hirschelman, W. H. 225th ACS National Meeting: Divi-
sion of Medicinal Chemistry, 2003, 86.
1
0. Fregona, D.; Giovagnini, L.; Ronconi, L.; Marzano, C.;
Trevisan, A.; Sitran, S.; Biondi, B.; Bordin, F. J. Inorg.
Biochem. 2003, 93, 181.
1. Marzano, C.; Trevisan, A.; Giovagnini, L.; Fregona, D.
Toxicol. Vitro 2002, 16, 413.
2. Faraglia, G.; Fregona, D.; Sitran, S.; Giovagnini, L.;
Marzano, C.; Baccicchetti, F.; Casellato, U.; Graziani, R.
J. Inorg. Biochem. 2001, 83, 31.
1
1
In summary, we have synthesized various 4-N atom
substituted derivatives of the lead compound 4 and
tested their preliminary in vitro anticancer activities.
Compound 6a without substituent on the 4-N atom
was found to be more potent than the lead compound
13. Lee, D. H. S.; Macintyre, J. P.; Taylor, G. R.; Wang, E.;
Plante, R. K.; Tam, S. S. C.; Pope, B. L.; Lau, C. Y.
J. Pharmacol. Exp. Ther. 1999, 289, 1465.
1
4. Ge, Z. M.; Li, R. T.; Cheng, T. M. Synth. Commun. 1999,
9, 3191.
2
1
5. Guo, B. G.; Ge, Z. M.; Cheng, T. M.; Li, R. T. Synth.
Commun. 2001, 31, 3021.
4
. Increase in the polarity and/or introduction of suit-
able acyl groups at 4-N atom of the lead compound 4
are favorable to improve the activity. These results
provide promising information for further develop-
ment of potent inhibitors. Further optimization results
for the lead compound 4 will be reported in due
course.
1
1
6. Li, R. T.; Cai, M. S. Synth. Commun. 1999, 29, 65.
7. Guo, B. G.; Ge, Z. M.; Cheng, T. M.; Li, R. T. Acta
Pharm. Sin. 2001, 36, 185.
18. Ge, Z. M.; Li, R. T.; Cheng, T. M.; Cui, J. R. Arch.
Pharm. Pharm. Med. Chem. 2001, 334, 173.
1
9. Guo, W.; Ran, F. X.; Wang, R. Q.; Cui, J. R.; Li, R. T.;
Cheng, T. M.; Ge, Z. M. Chin. J. Clin. Pharmacol. Ther.
2
004, 9, 59.
2
2
2
0. Li, R. T.; Ding, P. Y.; Han, M.; Cai, M. S. Synth.
Commun. 1998, 28, 295.
1. Crookes, J. C.; Woolf, A. A. J. Chem. Soc., Dalton Trans.
Acknowledgments
This research was supported by the funds of National
Natural Science Foundation of China (No. 20172006)
and the Major State Basic Research Development Pro-
gram (Grant No. 2004CB719900). Biological activities
were completed by the State Key Laboratory of Natural
and Biomimetic Drugs, Beijing, and National Center
For Drug Screening, Shanghai Institute of Medica,
Chinese Academy of Sciences.
1
973, 12, 1241.
2. Human promyelocytic leukemic cell line (HL-60), human
hepatocellular carcinoma cell line (Bel-7402), human
gastric carcinoma cell line (BGC-823), human cervical
carcinoma cell line (HeLa), human prostatic carcinoma
cell line (PC 3MIE8), and human breast carcinoma cell
line (MDA-MB-435) were grown and maintained in
RPMI-1640 medium supplemented with 10% fetal bovine
serum, penicillin (100 U/mL), and streptomycin (100 lg/
mL) at 37 ꢁC in humidified incubators in an atmosphere of
5
2
% CO . All of experiments were performed on exponen-
Supplementary data
tially growing cancer cells. Numbers of viable cell of
cancer were determined by MTT and SRB assays. Briefly,
4
À1
Supplementary data associated with this article can be
found, in the online version, at doi:10.1016/j.bmcl.
cancer cells (1–2.5 · 10 cells mL ) were inoculated in 96-
well culture plates (180 lL/well). After 24 h, 20 lL of
culture medium containing compounds of various con-
2
006.05.085.

X. Hou et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4214–4219
4219
centrations was added to wells, and RPMI-1640 medium
in control cells, then cells were incubated for 48 h. HL-60
cells were assayed by MTT, and the Bel-7402 cells were
assayed by SRB. The absorbance of each well was
measured using a microculture plate reader at 570 nm
23. Skehan, P.; Storeng, R.; Scudiero, D.; Monks, A.;
McMahon, J.; Vistica, D.; Warren, J. T.; Bokesch, H.;
Kenney, S.; Boyd, M. R. J. Natl. Cancer Inst. 1990, 82,
1170.
24. Carmichael, J.; DeGraff, W. G.; Gazdar, A. F.; Minna, J.
D.; Mitchell, J. B. Cancer Res. 1987, 47, 936.
(
MTT) and 540 nm (SRB).