Synthesis, antitumor activity, and cytotoxicity of 4-substituted 1-benzyl-5-diphenylstibano-1H-1,2,3-triazoles

Article abstract of DOI:10.1016/j.bmcl.2017.11.038

Trisubstituted 5-organostibano-1H-1,2,3-triazoles (3a–f) were synthesized by the Cu-catalyzed azide-alkyne cycloaddition of various ethynylstibanes (1) with benzylazide (2) in the presence of CuBr (5 mol%) under aerobic conditions. The reaction of 5-stiba

Full text of DOI:10.1016/j.bmcl.2017.11.038

Accepted Manuscript
Synthesis, antitumor activity, and cytotoxicity of 4-substituted 1-benzyl-5-di-
phenylstibano-1H-1,2,3-triazoles
Mizuki Yamada, Tsutomu Takahashi, Mai Hasegawa, Mio Matsumura, Kanna
Ono, Ryota Fujimoto, Yuki Kitamura, Yuki Murata, Naoki Kakusawa, Motohiro
Tanaka, Tohru Obata, Yasuyuki Fujiwara, Shuji Yasuike
PII:
S0960-894X(17)31137-X
https://doi.org/10.1016/j.bmcl.2017.11.038
BMCL 25444
DOI:
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
22 October 2017
22 November 2017
23 November 2017
Please cite this article as: Yamada, M., Takahashi, T., Hasegawa, M., Matsumura, M., Ono, K., Fujimoto, R.,
Kitamura, Y., Murata, Y., Kakusawa, N., Tanaka, M., Obata, T., Fujiwara, Y., Yasuike, S., Synthesis, antitumor
activity, and cytotoxicity of 4-substituted 1-benzyl-5-diphenylstibano-1H-1,2,3-triazoles, Bioorganic & Medicinal
Chemistry Letters (2017), doi: https://doi.org/10.1016/j.bmcl.2017.11.038
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BMCL
Synthesis,
antitumor
activity,
and
cytotoxicity
of
4-substituted
1-benzyl-5-diphenylstibano-1H-1,2,3-triazoles
a
b
a
a
Mizuki Yamada , Tsutomu Takahashi , Mai Hasegawa , Mio Matsumura , Kanna Ono b, Ryota
c
a
Fujimoto b, Yuki Kitamura a, Yuki Murata a, Naoki Kakusawa , Motohiro Tanaka , Tohru Obata a,*,
Yasuyuki Fujiwara b,*, Shuji Yasuike a,*
aSchool of Pharmaceutical Sciences, Aichi Gakuin University, 1-100 Kusumoto-cho, Chikusa-ku,
Nagoya 464-8650, Japan
bSchool of Pharmacy, Tokyo University of Pharmacy and life Sciences, 1432-1 Horinouchi, Hachioji,
Tokyo 192-0392, Japan
cFaculty of Pharmaceutical Sciences, Hokuriku University, Ho-3 Kanagawa-machi, Kanazawa
920-1181, Japan
*Corresponding authors.
Address: School of Pharmaceutical Sciences, Aichi Gakuin University, 1-100 Kusumoto-cho,
Chikusa-ku, Nagoya 464-8650, Japan
Tel: +81-52-757-6774
E-mail address: s-yasuik@dpc.agu.ac.jp (S. Yasuike)
yasuyuki@toyaku.ac.jp (Y. Fujiwara)
tobata@dpc.agu.ac.jp (T. Obata)
1

ABSTRACT
Trisubstituted 5-organostibano-1H-1,2,3-triazoles (3a-f) were synthesized by the Cu-catalyzed
azide-alkyne cycloaddition of various ethynylstibanes (1) with benzylazide (2) in the presence of
CuBr (5 mol%) under aerobic conditions. The reaction of 5-stibanotriazoles with HCl afforded
C5-unsubstituted
1,2,3-triazoles
(4a-f).
The
antitumor
activity
of
trisubstituted
5-organostibano-1H-1,2,3-triazoles (3a-f) and their 5-unsubstituted 1,2,3-triazoles (4a-f) were
evaluated in several tumor cell lines. All 5-stibanotriazoles (3a-f) exerted an excellent antitumor
activity. On the contrary, 5-unsubstituted 1,2,3-triazoles (4a-f) without a diphenylantimony group in
the molecule exhibited very low antitumor activity compared with 5-stibanotriazoles (3a-f). In
compounds of both the series, the substituted 4-butyl group appeared to decrease antitumor activity.
However, results suggested that organometal (antimony) in the molecule was required for greater
antitumor activity. In addition, all 5-stibanotriazoles (3a-f), but not all 5-unsubstituted 1,2,3-triazoles
(4a-f), exhibited cytotoxicity in normal vascular endothelial cells derived from bovine aorta. Among
the compounds (3b-e) that exhibited excellent antitumor activity, those with 4-methylphenyl (3b) and
1-cyclohexenyl (3e) showed relatively low cytotoxicity to vascular endothelial cells. Together, these
results suggest that trisubstituted 5-organostibano-1H-1,2,3-triazoles, including compounds 3b and
3e, may serve as potential anticancer therapeutic drugs in the future.
Keywords:
Cu-catalyzed azide-alkyne cycloaddition
5-Stibanotriazole
Antimony
Antitumor
Cytotoxicity
1,2,3-Triazoles are an important class of nitrogen-containing heterocyclic ring, and are key in the
design
and
synthesis
of
novel
biologically
active
agents.1,2
Among
these,
2

1,4,5-trisubstituted-1,2,3-triazoles have attracted interest as target molecules because of their use as
reagents in biological activities. For instance, SST0287CL1 I3 exhibits heat shock protein 90
inhibition, sulfur-containing triazole II4 is a potential herbicide with antifungal activity, triazole III5
behaves as a potent inhibitor of human aromatase, and triazole derivative IV6 having isoxazolyl is a
potent non-pseudo-substrate inhibitor of O6-alkylguanine-DNA-methyltransferase (Fig. 1).
Additionally, antimony compounds have attracted much interest because of their importance as
reagents in biological activities.7-9 Interestingly, sodium stibogluconate, of the pentavalent antimony
compound class, has been used for more than half a century in the therapy of the parasitic disease
leishmaniasis.10-12 However, there are few reports on the biological activities of trivalent
organoantimony compounds9 and we have recently reported on the synthesis and biological activity
of both trivalent and pentavalent organoantimony compounds.13-18 Among these, organoantimony (III)
compounds
such
as
1-[(2-di-p-tolylstibanophenyl)diazenyl]pyrrolidine15
and
2-(di-p-tolylstibano)-N-p-tolylbenzamide16 showed potent antiproliferative activity on human tumor
cell lines such as NB4, HeLa, L1210, Mm1, and DLD-1. For instance,
2-(di-p-tolylstibano)-N-p-tolylbenzamide showed moderate antitumor activities [IC50 = 4.6–10.8 µM]
in several tumor cell lines; however, N-p-tolylbenzamide without antimony group was inactive [IC50
3

> 200 µM].16 Since these results indicate that organoantimony (III) compounds affect antitumor
activity, we are interested in the biological activity of fully substituted 1,2,3-triazoles with antimony
as
a
substituent.
We
present
here
the
synthesis
of
novel
4-substituted
1-benzyl-5-diphenylstibano-1,2,3-triazoles using the Cu-catalyzed azide-alkyne cycloaddition of
ethynyldiphenylstibane with benzyl azide and their antitumor activity and cytotoxicity, including
comparisons of the obtained organoantimony compounds and 5-unsubstituted 1,2,3-triazoles.
Recently, we developed the regioselective Cu-catalyzed azide-alkyne cycloaddition of
ethynylstibanes with organic azides to form fully substituted 5-organostibano-1,2,3-triazoles.19,20
However, the ethynylstibanes used required p-tolyl group on antimony for determining
regioselectivity of the reaction, and their synthesis has been complicated for general synthesis.
Therefore, ethynylstibanes (1a-f) were selected as the key starting material, as they could be easily
prepared according to the general method used previously.21 The terminal alkynes were treated with
n-BuLi in dry ether under an argon atmosphere at 0 °C, and followed by trapping with
diphenylantimony bromide to afford arylethynyl, vinylethynyl, and alkylethynyl compounds (1a-f) in
44–83% yields. The Cu-catalyzed azide-alkyne cycloaddition of ethynylstibanes with aryl (1a-d),
vinyl (1e), and alkyl (1f) group with benzylazide (2) in the presence of CuBr2 (5 mol%) led to the
formation of 5-stibanotriazoles (3a-f) in good-to-excellent yields under the optimal conditions, as
reported previously (Scheme 1).19,20,22 The diphenylantimony group in 3 was readily removed by
treatment with 10% HCl in THF to obtain the desired C5-unsubstituted 1,2,3-triazoles (4) in
satisfactory yields. The structures of 4 were confirmed by comparisons of NMR and MS spectra with
authentic samples in the literature. Moreover, a nuclear Overhauser effect (NOE) was observed in
benzyl protons and C-5 proton on triazole ring in 4. These results show that Cu-catalyzed
azide-alkyne cycloaddition of 1 with 2 proceeds regioselectively to yield 5-stibanotriazole (3).
4

The
antitumor
activities
of
the
novel
synthesized
4-substituted
1-benzyl-5-diphenylstibano-1,2,3-triazoles and their 5-unsubstituted 1,2,3-triazoles were evaluated in
eight cultured tumor cell lines including those of mouse and human origin (Table 1). All
5-stibanotriazoles (3a-f) exerted excellent antitumor activity in all tested tumor cell lines, even
human solid tumor cell lines such as breast, colon, and gastric tumors. The range of IC50 value was
indicated at 0.25–11.9 µM and displayed almost equal IC50 values as those of cisplatin (CDDP), a
well-known commercial antitumor drug containing a metal in its molecular structure. Among them,
the compounds with 4-methylphenyl (3b), phenyl (3c), 4-trifluoromethylphenyl (3d), and
1-cyclohexenyl (3e) groups had the lowest IC50 values, and those with 4-methoxylphenyl (3a)
indicated sequential activity. The average IC50 values in those cell lines for the compound with
4-n-butyl group (3f) was the highest recorded value (7.16 µM) but the antitumor activity of it was not
disappearance. Conversely, 5-unsubstituted 1,2,3-triazoles (4a-f) without diphenylantimony group in
the molecule exhibited very low antitumor activity compared with 5-stibanotriazoles (3a-f). All IC50
values of the compounds without diphenylantimony groups increased compared to that of their
corresponding compounds. Among them, 1-benzyl-4-(p-trifluoromethylphenyl)-1H-1,2,3-triazole
(4d) exhibited middling antitumor activity with an average IC50 value of 82 µM. In compounds of
both series, the substituted alkyl chain, such as 4-butyl group, appeared to decrease antitumor activity
but the effect of the substituted functional group such as electron-withdrawing and -donating groups
at the 4-position was less. However, it appeared that the existence of organometal (antimony) in the
molecule was required for greater antitumor activity.
5

Table 1. Comparison of antitumor activity between 5-stibanotriazoles (3a-f) and 5-unsubstituted triazoles (4a-f) in tumor
cell lines.
P388
B16-F10
HL-60
KB
HT-1080
MCF-7
DLD-1
NUGC-3
3a
3b
0.54 0.20 0.47 0.04 0.87 0.45 0.86 0.24 0.50 0.12 0.66 0.27 0.80 0.18 1.38 0.59
0.61 0.33 0.32 0.02 0.47 0.07 0.49 0.14 0.35 0.09 0.47 0.10 0.37 0.06 0.84 0.10
0.31 0.04 0.44 0.12 0.93 0.60 0.52 0.09 0.38 0.04 0.59 0.35 0.43 0.12 0.87 0.48
0.25 0.06 0.39 0.05 0.50 0.31 0.50 0.10 0.40 0.11 0.75 0.43 0.38 0.04 0.72 0.28
0.77 0.40 0.34 0.01 0.72 0.28 0.47 0.10 0.41 0.16 0.53 0.07 0.40 0.05 0.71 0.15
3c
3d
3e
3f
6.11 1.73 1.61 0.36
83.8 13.9 88.5 24.5
94.1 31.5 66.4 24.0
99.1 37.8 96.1 19.1
11.9 7.9 6.54 1.06 9.02 3.12 7.73 2.84 4.90 1.39 9.50 3.88
>100
>100
>100
86.6 7.9
79.5 11.0 84.4 23.8
>100 651 559
146 106
231 100
253 190
490 157
162 97
>100
139 69
102 31
>100
158 29
129 23
836 429
4a
4b
4c
4d
4e
4f
17.5 7.6
49.6 12.6 45.5 18.7 171 47.0
217 135 421 128 >100
35.4 17.1 171 64.2 60.2 11.6 57.7 10.5
74.3 52.2 81.0 25.9
>100
>100
>100
>100
219 145
>100
>100
>100
>100
Each value was represented IC50 SD (µM).
In
order
to
test
the
cytotoxicity
of
the
novel
synthesized
4-substituted
1-benzyl-5-diphenylstibano-1,2,3-triazoles and their 5-unsubstituted 1,2,3-triazoles in normal cells,
the non-tumor vascular endothelial cells derived from bovine aorta were used. All 5-stibanotriazoles
(3a-f) exhibited higher antitumor activity (Table 1) and cytotoxicity (Table 2) than the
5-unsubstituted 1,2,3-triazoles (4a-f). The highest cytotoxic effect to vascular endothelial cells was
observed for compounds with 4-methoxylphenyl (3a, CC50 = 1.9 0.45 µM), phenyl (3c, CC50 = 2.1
0.22 µM), and 4-trifluoromethylphenyl (3d, CC50 = 1.5 0.07 µM) group. Although the compounds
with 4-methylphenyl (3b) and 1-cyclohexenyl (3e) group, together with the compounds with phenyl
(3c) and 4-trifluoromethylphenyl (3d) group, had the lowest IC values on the tumor cell lines (Table
50
1), they did not have the lowest CC50 values on the vascular endothelial cells (3b, CC50 = 5.4 1.61
µM; 3e, CC50 = 7.4 0.71 µM). The compound with 4-n-butyl (3f) showed a lower toxicity on the
vascular endothelial cells as well as lower antitumor activity among all 5-stibanotriazoles (3a-f).
Moreover, the treatment of compounds with 4-methylphenyl (3b) and 1-cyclohexenyl (3e) at a
concentration of 0.5 µM supported antitumor activity in tumor cell lines, but did not exhibit a
cytotoxic effect in non-tumor vascular endothelial cells (data not shown). These results indicate that
tumor cell lines are more sensitive to compounds with 4-methylphenyl (3b) and 1-cyclohexenyl (3e)
than non-tumor vascular endothelial cells. Moreover, since compounds 3b and 3e exhibited excellent
6

antitumor activity in all tested tumor cell lines and relatively low cytotoxicity in non-tumor vascular
endothelial cells, it is possible that these compounds can effectively suppress tumor growth without
damage to normal cells.
Table 2. Comparison of cytotoxicity between 3
and 4 in non-tumor vascular endothelial cells.
Compounds
CC50 (mean SD, µM) a
1.9 0.45
5.4 1.61
2.1 0.22
1.5 0.07
7.4 0.71
22.9 6.87
> 100
3a
3b
3c
3d
3e
3f
4a
4b
4c
4d
4e
4f
> 100
> 100
> 100
> 100
> 100
a CC50: 50% cytotoxic concentration.
In conclusion, the Cu-catalyzed azide-alkyne cycloaddition of various ethynylstibanes with
benzylazide,
under
mild
reaction
conditions,
afforded
4-substituted
1-benzyl-5-diphenylstibano-1H-1,2,3-triazoles. The 5-stibanotriazoles exerted greater antitumor
activity in eight cultured tumor cell lines, including the human solid tumor cell line, than the
corresponding 5-unsubstituted triazoles. Among the compounds 3b-e that exhibited excellent
antitumor activity, compounds with 4-methylphenyl (3b) and 1-cyclohexenyl (3e) showed relatively
low cytotoxicity to normal cells. These results suggest that compounds 3b and 3e may serve as
potential anticancer therapeutic drugs in future.
Acknowledgements
This research was supported by a research grant from Nagai Memorial Research Scholarship from
the Pharmaceutical Society of Japan (to M. Y.). The authors also thank Institute of Pharmaceutical
Life Sciences, Aichi Gakuin University, Hokuriku University, and Tokyo University of Pharmacy and
Life Sciences for generous financial support.
7

A. Supplementary data
Supplementary data associated with this article can be found, in the online version, at http://
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9

Highlights:
Trisubstituted 5-stibanotriazoles were prepared by the Cu-catalyzed cycloaddition.
5-Substituted triazoles exerted great antitumor activity in several tumor cell lines.
Two 5-stibanotriazoles showed low cytotoxicity to normal vascular endothelial cells.
10

Vascular endothelial cells
1) A novel 3a-f with excellent antitumor activity
were designed and synthesized
(µM)
2) Antitumor activity of 3a-f against DLD-1 with
IC50 = 0.37 – 4.9 µM
3) 3b (4-tolyl) and 3e (4-cyclohexenyl) showed
low cytotoxicity to non-tumor cells
IC50 CC50
3d 0.38
1.5
5.4
7.4
3b 0.37
3e 0.40
Control
3d
3b
3e