Synthesis and antitumor activity of 1,3,4-oxadiazole possessing 1,4-benzodioxan moiety as a novel class of potent methionine aminopeptidase type II inhibitors

Article abstract of DOI:10.1016/j.bmcl.2013.03.068

A series of 1,3,4-oxadiazole derivatives containing 1,4-benzodioxan moiety (7a-7q) have been designed, synthesized and evaluated for their antitumor activity. Most of the synthesized compounds were proved to have potent antitumor activity and low toxicity. Among them, compound 7a showed the most potent biological activity against Human Umbilical Vein Endothelial cells, which was comparable to the positive control. The results of apoptosis and flow cytometry (FCM) demonstrated that compound 7a induce cell apoptosis by the inhibition of MetAP2 pathway. Molecular docking was performed to position compound 7a into MetAP2 binding site in order to explore the potential target.

Full text of DOI:10.1016/j.bmcl.2013.03.068

Bioorganic & Medicinal Chemistry Letters 23 (2013) 2876–2879
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and antitumor activity of 1,3,4-oxadiazole possessing
1,4-benzodioxan moiety as a novel class of potent methionine
aminopeptidase type II inhibitors
Juan Sun a, Ming-Hui Li a, Shao-Song Qian b, Feng-Jiao Guo b, Xiao-Fang Dang b, Xiao-Ming Wang a,
,
⇑
Ya-Rong Xue a, , Hai-Liang Zhu a,b,
⇑
⇑
a State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University, Nanjing 210093, PR China
b School of Life Sciences, Shandong University of Technology, Shandong 255049, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Available online 28 March 2013
A series of 1,3,4-oxadiazole derivatives containing 1,4-benzodioxan moiety (7a–7q) have been designed,
synthesized and evaluated for their antitumor activity. Most of the synthesized compounds were proved
to have potent antitumor activity and low toxicity. Among them, compound 7a showed the most potent
biological activity against Human Umbilical Vein Endothelial cells, which was comparable to the positive
control. The results of apoptosis and flow cytometry (FCM) demonstrated that compound 7a induce cell
apoptosis by the inhibition of MetAP2 pathway. Molecular docking was performed to position compound
7a into MetAP2 binding site in order to explore the potential target.
Keywords:
Benzodioxan
Oxadiazole
Antitumor activity
MetAP2
Molecular docking
Ó 2013 Elsevier Ltd. All rights reserved.
Cancer, second cause of mortality in the world, increases with
age in both rodents and humans.1,2 Cancer continues to be a world-
wide killer, despite the enormous amount of research and rapid
developments during the past decade.3 Cancer chemotherapy has
entered a new field of molecularly targeted therapeutics, which
is highly selective and not associated with the serious toxicities
of conventional cytotoxic drugs.4 Therefore, there is an increasing
need for new therapies, especially those that are based on current
knowledge of cancer biology as well as that taking advantage of the
cancer cells phenotype, described by Hanahan and Weinberg.5
Methionine aminopeptidase 2 (MetAP2) appears to play a critical
role in cell proliferation and tumor growth. It is expressed at higher
concentrations in tumors as compared to normal cells.6 Available
reports also suggested that MetAP2 plays an important role in
the growth of different types of tumors.7
Compounds containing a 1,4-benzodioxan template have re-
ceived significant attention in chemical, medicinal and pharmaceu-
tical research as this structural scaffold is found in a variety of
drugs.8 Some 1,4-benzodioxan derivatives are anti-psychotic
agents and others are used by living species in their chemical com-
munication systems. Recently, some reports claimed that a number
of other 1,4-benzodioxan template-containing compounds also
have ability as potential anticancer drugs with excellent bioavail-
ability and low cytotoxicity.9–12 For example, (Z)-N-(5-(2,3-
dihydrobenzo[b][1,4]dioxin-6-yl)-1,3,4-thiadiazol-2-yl)-3-phenyl-
acrylamide, having a 1,4-benzodioxan structure was suggested to
have strong antitumor activity.13 Oxadiazole derivatives play a
significant role in various pharmaceutical applications.14–18 As an
important class of heterocyclic compound, 1,3,4-oxadiazoles
exhibit antitumor activities particularly.19–21 2-(2,3-dihydro-
benzo[b][1,4]dioxin-6-yl)-5-((2-methylbenzyl)thio)-1,3,4-oxadiaz-
ole, having a 1,3,4-oxadiazole structure was suggested to have
strong antitumor activity.21
A series of 1,3,4-oxadiazole derivatives containing 1,4-benzodi-
oxan template were firstly synthesized and antitumor activities
were examined. Among them, the compounds 7a displayed the
most potent antitumor activity by inhibiting Human Umbilical
Vein Endothelial Cells proliferation and had low toxicity. Prelimin-
ary study on the antitumor mechanism of the compounds found
that compounds induced cell apoptosis by the inhibition of MetAP2
pathway. From the results, we can conclude that some of the syn-
thesized compounds are potent antitumor agents.
Methyl 2,3-dihydrobenzo[b][1,4]dioxine-6-carboxylate 3 was
prepared in two steps as shown in Scheme 1. It was prepared in
two steps. Firstly, 3,4-dihydroxybenzoic acid 1 gave methyl 3,4-
dihydroxybenzoate 2, catalyzed by concentrated sulfuric acid in
methanol. Secondly, compound 3 was prepared by treatment of 2
with dibromoethane in acetone.
Seventeen 1,3,4-oxadiazole derivatives (7a–7q) were prepared
as shown in Scheme 1. Firstly, 2,3-dihydrobenzo[b][1,4]dioxine-
6-carbohydrazide was prepared by treatment of 3 with hydrazine
⇑
Corresponding authors. Tel./fax: +86 25 8359 2672.
E-mail address: zhuhl@nju.edu.cn (H.-L. Zhu).
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.03.068

J. Sun et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2876–2879
2877
O
O
OH
HO
HO
O
O
a
O
b
O
HOOC
OH
R2
2
1
3
1
R3
R
CHO
d
COOH
R
5a-5q
4
O
O
O
O
NH2
O
O
N
H
f
O
6
3
N
O
N
R3
R
O
R1
COOH
R2
g
O
7a-7q
a R1=F , R2=H , R3=H
b R1=H , R2= F, R3=H
c R1=H , R2=H , R3=F
d R1=Cl , R2=H , R3=H
e R1=H , R2=H , R3=Cl
f R1=Br , R2=H , R3=H
g R1=H , R2=Br , R3=H
h R1=H , R2=H , R3=Br
m R1=H , R2=NO2 , R3=H
n R1=H , R2=H , R3=NO2
o R1=H , R2=H , R3=H
i R1=CH3 , R2= H, R3=H
j R1=H , R2=CH3 , R3=H
k R1=H , R2=H , R3=CH3
l R1=NO2 , R2=H , R3=H
p R1=H , R2=CH3O , R3=H
q R1=H , R2=H , R3=CH3O
Scheme 1. Synthesis of compounds 3, 5a–5q and 7a–7q. Reagents and conditions: (a) methanol, concentrated sulfuric acid, 90 °C, 8 h; (b) dibromoethane, potassium
carbonate, acetone, 70 °C, 12 h; (d) pyridine, piperidine, 85 °C, 24 h; (f) hydrazine hydrate, ethanol, 90 °C, 4 h; (g) phosphorous oxychloride, 110 °C, 5 h.
Table 2
Table 1
Cell proliferation assay of the compounds on Human
Cytotoxicity assay of the compounds on Human
Umbilical Vein Endothelial Cells
Umbilical Vein Endothelial Cells
Compounds
IC50 SD (lM)
Compounds
CC50 SD (lM)
6
7a
7b
7c
7d
7e
7f
7g
7h
7i
7j
>50
6
200.18 12.19
402.23 21.04
295.27 20.00
325.17 31.11
367.82 18.91
312.00 21.12
287.45 12.95
321.95 22.11
491.23 19.76
356.23 11.09
200.98 11.02
265.26 21.08
323.15 21.23
309.13 12.45
298.77 22.10
209.22 18.36
332.18 24.12
312.23 18.46
1.16
3.53
4.96
7.71
8.03
8.89
13.53
22.32
40.49
40.60
45.21
25.02
30.13
32.07
>50
7a
7b
7c
7d
7e
7f
7g
7h
7i
7j
7k
7l
7m
7n
7o
7p
7q
7k
7l
7m
7n
7o
7p
7q
TNP-470
>50
>50
1.96
hydrate (85%) in ethanol. Then, compound 6 gave 1,3,4-oxadiazole
derivatives by refluxing in anhydrous phosphorus oxychloride
with different substituted cinnamic acids.
Generally speaking, the inhibitory activity of the compounds is
due to cell apoptosis or toxic effect, so we firstly performed
cytotoxicity test before detecting biological activity. The oxadiaz-
ole compounds were detected for their cyototoxicity on Human
Umbilical Vein Endothelial cells (HUVEC). The pharmacological re-
sults of these compounds were summarized in Table 1. What we

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J. Sun et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2876–2879
Table 3
MetAP2 inhibitory activity of the selected compounds
The MetAP2 inhibitory potency of the oxadiazole derivatives
containing 1,4-benzodioxan was examined and the results were
summarized in Table 3. Most of the tested compounds displayed
potent MetAP2 inhibiting activity. Among them, compound 7a
Compounds
MetAP2 (IC50, lM)
7a
7b
2.08
4.79
showed the most potent inhibitory with IC50 of 2.08 lM. The
7c
6.33
results of MetAP2 inhibitory activity of the tested compounds were
in agreement to the structure relationships (SAR) of their antitu-
mor activities. This agreement suggested that antitumor activities
of the synthesized compounds may be derived from the inhibition
of MetAP2 enzymatic activities.
In order to study the preliminary antitumor mechanism of the
compounds, we performed flow cytometry (FCM) (Fig. 1). As
shown in Figure 1, HUVEC were treated with 0, 0.5, 1, 2 and
7d
7e
10.04
7.92
7f
8.41
7g
7h
7l
7m
7n
10.71
12.66
22.03
18.24
13.95
1.32
TNP-470
4 lM of compound 7a for 24 h. The compound can increase the
percentage of apoptosis in a dose-independent manner. The results
indicated that compound 7a can induce apoptosis of HUVEC.
In an effort to elucidate the possible mechanism by which the
title compounds can induce anticancer activity and guide further
SAR studies, molecular docking of the potent inhibitor 7a into
ATP binding site of MetAP2 was performed on the binding model
based on the MetAP2 (2EA4.pdb). The binding models of
compound 7a and MetAP2 were depicted in Figure 2. In the bind-
can see from the data is that most of the compounds were low
toxic.
All the synthesized derivatives (7a–7q) were tested in vitro for
the inhibition activity on HUVEC. The results were summarized in
Table 2. The table showed that most of the synthesized compounds
exhibited potent inhibitory activity with the IC50 value at low
micromolar.
ing model, compound 7a was nicely bound to the MetAP2 via p–p
Structure–activity relationship (SAR) analysis indicated that
compounds with electron-withdrawing group showed stronger
activity than that with electron-donating group, with all the IC50
interaction (between benzene ring and HIS 231) and one hydrogen
bond (F atom on the benzene ring and amino hydrogen of ASN 329
form hydrogen bond: H–FÁ Á ÁH).
values below 50
l
M. In further study of compounds with elec-
In conclusion, a series of 1,3,4-oxadiazole derivatives containing
1,4-benzodioxan have been synthesized and evaluated for their
antitumor activities. Compound 7a demonstrated the most potent
inhibitory activity that inhibited the growth of HUVEC cells with
tron-withdrawing group, different group led to different antitumor
activity, and the potency order was F (fluorine) > Cl (chlorine) > Br
(bromine) > NO2 (nitro-group) > CH3 > CH3O. Among these com-
pounds, substituent at different positions led to different antitu-
mor activity, and the potency order was ortho- > meta- > para-.
IC50 of 1.16
lM and inhibited the activity of MetAP2 with IC50 of
2.08 M, which was comparable to the positive control TNP-470.
l
Figure 1. HUVEC were cultured with anticancer and various concentrations of 7a for 24 h. Cells were stained by Annexin VeFITC/PI and apoptosis was analyzed by flow
cytometry. Inhibition including early and late apoptosis.

J. Sun et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2876–2879
2879
Figure 2. Molecular docking modeling of compound 7a with MetAP2: for clarity, only interacting residues are displayed. Left: 3D model of the interaction between
compound 7a and the MetAP2 binding site. Right: 2D model of the interaction between compound 7a and the MetAP2 binding site.
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This work was supported by Jiangsu National Science Founda-
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