Identification of methionine aminopeptidase 2 as a molecular target of the organoselenium drug ebselen and its derivatives/analogues: Synthesis, inhibitory activity and molecular modeling study

Article abstract of DOI:10.1016/j.bmcl.2016.09.050

A collection of twenty-six organoselenium compounds, ebselen and its structural analogues, provided a novel approach for inhibiting the activity of human methionine aminopeptidase 2 (MetAP2). This metalloprotease, being responsible for the removal of the amino-terminal methionine from newly synthesized proteins, plays a key role in angiogenesis, which is essential for the progression of diseases, including solid tumor cancers. In this work, we discovered that ebselen, a synthetic organoselenium drug molecule with anti-inflammatory, anti-oxidant and cytoprotective activity, inhibits one of the main enzymes in the tumor progression pathway. Using three-step synthesis, we obtained twenty-five ebselen derivatives/analogues, ten of which are new, and tested their inhibitory activity toward three neutral aminopeptidases (MetAP2, alanine and leucine aminopeptidases). All of the tested compounds proved to be selective, slow-binding inhibitors of MetAP2. Similarly to ebselen, most of its analogues exhibited a moderate potency (IC₅₀= 1–12 μM). Moreover, we identified three strong inhibitors that bind favorably to the enzyme with the half maximal inhibitory concentration in the submicromolar range.

Full text of DOI:10.1016/j.bmcl.2016.09.050

Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Identification of methionine aminopeptidase 2 as a molecular target
of the organoselenium drug ebselen and its derivatives/analogues:
Synthesis, inhibitory activity and molecular modeling study
Ewelina We˛glarz-Tomczak a, , Małgorzata Burda-Grabowska a,b, Mirosław Giurg b, Artur Mucha a
⇑
a
_
Department of Bioorganic Chemistry, Faculty of Chemistry, Wrocław University of Technology, Wybrzeze Wyspian´skiego 27, 50-370 Wrocław, Poland
b
_
Department of Organic Chemistry, Faculty of Chemistry, Wrocław University of Technology, Wybrzeze Wyspian´skiego 27, 50-370 Wrocław, Poland
a r t i c l e i n f o
a b s t r a c t
Article history:
A collection of twenty-six organoselenium compounds, ebselen and its structural analogues, provided a
novel approach for inhibiting the activity of human methionine aminopeptidase 2 (MetAP2). This metal-
loprotease, being responsible for the removal of the amino-terminal methionine from newly synthesized
proteins, plays a key role in angiogenesis, which is essential for the progression of diseases, including
solid tumor cancers. In this work, we discovered that ebselen, a synthetic organoselenium drug molecule
with anti-inflammatory, anti-oxidant and cytoprotective activity, inhibits one of the main enzymes in the
tumor progression pathway. Using three-step synthesis, we obtained twenty-five ebselen derivatives/
analogues, ten of which are new, and tested their inhibitory activity toward three neutral aminopepti-
dases (MetAP2, alanine and leucine aminopeptidases). All of the tested compounds proved to be selective,
slow-binding inhibitors of MetAP2. Similarly to ebselen, most of its analogues exhibited a moderate
Received 1 August 2016
Revised 16 September 2016
Accepted 17 September 2016
Available online xxxx
Keywords:
Metalloaminopeptidases
Angiogenesis
Organoselenium compounds
Structure–activity relationship
Binding mode
potency (IC50 = 1–12
lM). Moreover, we identified three strong inhibitors that bind favorably to the
enzyme with the half maximal inhibitory concentration in the submicromolar range.
Ó 2016 Elsevier Ltd. All rights reserved.
Alanine (APNs), leucine (LAPs), and methionine aminopepti-
dases (MetAPs) are three major groups of neutral aminopeptidases
containing metal ion(s) in their active site that catalyze the
removal of amino acids from the N-terminus of a peptide or pro-
tein. Biomedical significance of these exopeptidases is related to
therapeutic intervention against devastating human diseases. The
leucine and alanine aminopeptidases represent promising targets
for the development of a new generation of anti-inflammatory
drugs. The alanine and methionine aminopeptidases possess a
well-recognized potential for the design of anti-angiogenesis
agents. Furthermore, neutral aminopeptidases are involved in the
apoptosis of cancer cells, which makes them highly interesting
objects of oncological research.1–4
Human MetAP type 2 (MetAP2) is one of the three known
methionine aminopeptidases responsible for the removal of the
N-terminal translation initiator methionine from newly synthe-
sized proteins, which is a critical step in protein maturation.3,5
Protein translation mainly begins with a methionine in eukaryotes
and a formylated methionine in prokaryotes. Consequently, the
removal of methionine is indispensable for post-translational
amino group modifications, protein stability and proper localiza-
tion.6,7 MetAP2 possesses a characteristic bimetallic center acti-
vated by cobalt, manganese or iron, surrounded by residues that
form a pita-bread-shaped fold.8–10 MetAPs are expressed in many
mammalian tissues and cell lines, but only type 2 is upregulated
during cell proliferation.11 Higher expression of MetAP2 has been
observed in tumor cells compared with normal cells.12,13 To date,
increased expression of MetAP2 has been correlated with several
types of cancer, for instance, mesothelioma,14 neuroblastoma,15
and colorectal carcinoma.16 MetAP2 became a promising target
molecule after the discovery that the potent anti-angiogenic agent
fumagillin and its synthetic analogue ovalicin bind covalently and
inhibit its aminopeptidase activity without affecting the ability of
the enzyme to stabilize eukaryotic initiation factor-2.17 TNP-470,
one of the synthetic analogues of fumagillin that advanced to clin-
ical trials,18 as well as more recently published analogues with
improved pharmacological profiles,19 act similarly. Reversible inhi-
bitors are generally considered as safer due to the decreased risk of
toxicity or immunogenic responses. The non-covalent inhibitors of
MetAP2 include compounds based on fumagillin,20 bestatin,21
1,2,4-triazole,22 and most recently pyrazolo[4,3-b]indole.23 Other
types of reversible ligands of MetAP2, such as anthranilic acid sul-
fonamides24–26 and bengamides,27 have also been reported.28 In
⇑
Corresponding author. Fax: +48 71 320 2427.
E-mail address: ewelina.weglarz-tomczak@pwr.edu.pl (E. We˛glarz-Tomczak).
http://dx.doi.org/10.1016/j.bmcl.2016.09.050
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: We˛glarz-Tomczak, E.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.09.050

2
E. We˛glarz-Tomczak et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
this communication, we describe the discovery and optimization of
the nontoxic synthetic selenium-containing drug ebselen as an
inhibitor of MetAP2.
compounds against pancreatic and renal,38 liver, breast,39 and lung
cancer and cervical adenocarcinoma.40 Organoselenium com-
pounds exhibit strong electrophilic activity and are therefore cap-
able of forming selenenyl-sulfide bonds with the cysteine residues
in proteins.32,41,42 The anticancer activity is usually linked to the
inhibition of a selenocysteine-containing enzyme, thioredoxin
reductase, that is overexpressed in many types of cancer.43 All
the above examples prove not only the broad spectrum of ebselen
activity, but also convenience and safety of its use. However, no
reports are available that show the relationship between ebselen
and aminopeptidases.
Ebselen, IUPAC name 2-phenyl-1,2-benzisoselenazol-3(2H)-
one, also called PZ51, is a low-molecular-weight drug with a pleio-
tropic mode of action and with very low toxicity in humans.29 It
was first identified as an anti-inflammatory agent with glutathione
peroxidase-like activity in living cells.30 One of the most likely
modes of action of ebselen is as follows: the SeꢀN bond is readily
cleaved by the thiol group of glutathione (GSH) to produce the cor-
responding selenenyl sulfides, which undergo disproportionation
toward GSSG and 2,20-dicarbamoyldiphenyl diselenide. The dise-
lenide is oxidized to the corresponding selenenic acid in the pres-
ence of hydroperoxides, such as hydrogen peroxide (H2O2). After
water elimination, ebselen is regenerated (Scheme 1).31,32
Ebselen is a well-known agent with therapeutic activity in neu-
rological disorders, acute pancreatitis, and noise-induced hearing
loss. It also exhibits antiatherosclerotic, antithrombotic, antioxi-
dant and cytoprotective properties.29,33–35 A recent study showed
that hypoxia-induced cytotoxicity in human alveolar cells is
reduced by ebselen owing to these properties.36 An antiviral effect
on hepatitis C virus nonstructural protein 3 was also recently
demonstrated.37 Ebselen and its derivatives act as antiproliferative
Ebselen has previously been prepared by several methods,44–49
first by Lesser and Weiss in 1924.49 In this work, we synthesized
ebselen and its derivatives/analogues using a four-step literature
procedure starting from cheap and easily available reagents,
anthranilic acid and elementary selenium.47,48 The overall proce-
dure for obtaining ebselen, its derivatives/analogues and their
acyclic dimeric forms is outlined in Scheme 2. The protonation of
anthranilic acid (3), diazotization, and dilithium diselenide sele-
nenylation with nitrogen elimination gave 2,20-dicarboxydiphenyl
diselenide (4), a stable crystalline compound. The reaction of dise-
lenide with thionyl chloride in the presence of dimethylformamide
(DMF) produced 2-(chloroseleno)benzoyl chloride (5) or its acyclic
form 6 depending on the quantity of thionyl chloride used, i.e.,
7 equiv or 3.5 equiv, respectively. Acylation or tandem selenenyla-
tion/acylation reaction with appropriate amines alone or in the
presence of Et3N base in anhydrous acetonitrile (benzisoselena-
zolones 1a–r) or appropriate amines in the presence of Na2CO3 in
anhydrous dichloromethane (diselenides 2a, 2c–e and 2l–n) com-
pleted the reaction sequence. We used ammonia or eighteen struc-
turally diversified primary amines (methyl, aniline and its
derivatives, including heteroatom- and halogen-substituted, ben-
zyl, p-tert-butylbenzyl, and phenylethyl, Scheme 2). In particular,
benzyl diselenide 2p was prepared from corresponding benzisose-
lenazolone by hydrogenation with hydrazine monohydrate. The
final compounds were purified by recrystallization or standard liq-
uid column chromatography.
O
H2O
GSH
N
Se
ebselen
H
H
N
N
O
O
SeOH
SeSG
H
N
O
H2O2
Twenty-six compounds, eighteen benzisoselenazol-3(2H)-ones
(1a–r) and eight 2,20-dicarbamoyldiaryl diselenides (2a, 2c–e, 2l–
n and 2p), ten of which are new (1f, 1i–m, 1q, 1r, 2l and 2m), were
obtained. The products were characterized by 1H, 13C, and 77Se
NMR spectroscopy, mass spectrometry and melting point (see
Supplementary data).
Se
Se
GSSG
N
H
O
Bis(2-phenylcarbamoyl)phenyl diselenide
Scheme 1. Plausible mechanism for the GSH activity of ebselen.31,32
O
COCl
d
N
Se
R
SeCl
b
c
5
6
1a-r
COOH
NH2
COOH
a
f
Se
2
H
R
N
3
4
COCl
e
O
Se
2
Se
2
R = Ph (1a, 2a),
4-methylphenyl (1b)
1c, 2c)
2a 2c-e 2l-n 2p
,
,
,
4-fluorophenyl (1h)
1i
(
1n, 2n
)
2-hydroxyphenyl (
2-methoxy-5-methylphenyl
)
H (
2-methoxyphenyl (1d, 2d) 4-methoxy-2-methylphenyl (1j)
4-methoxyphenyl (1e, 2e) 5-tert-butyl-2-methoxyphenyl (1k)
Me (1o)
Bz (1p, 2p)
1f
4-n-butoxyphenyl (
1l, 2l
(
1q
)
)
2-chloro-4-methylphenyl
)
4-tert-butylbenzyl (
3-fluorophenyl (1g)
5-chloro-2-methoxyphenyl (1m, 2m)
2-phenylethyl (1r)
Scheme 2. Synthesis of the benzisoselenazol-3(2H)-ones 1a–r and bis(2-carbamoyl)phenyl diselenides 2a, 2c–e, 2l–n and 2p. Reagents and conditions: (a) (i) HCl, (ii) NaNO2,
0 °C, (iii) Li2Se2, 0 °C; (b) 7 equiv SOCl2, DMF, benzene, reflux; (c) 3.5 equiv SOCl2, DMF, benzene, reflux; (d) RNH2, Et3N, MeCN, or RNH2, MeCN, (e) RNH2, Na2CO3, CH2Cl2, (f)
H2NNH2 ꢁ H2O, MeOH, 25 °C.
Please cite this article in press as: We˛glarz-Tomczak, E.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.09.050

E. We˛glarz-Tomczak et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
3
Table 1
Table 2
Inhibitory activity of ebselen and its derivatives obtained by substitution/function-
alization of the phenyl ring toward MetAP2 aminopeptidase (measured after 30 min
of incubation). The most significant inhibition is highlighted in bold. The activity of
the reference compound (A832234), a transition state analogue inhibitor, is given
Inhibitory activity of ebselen analogues possessing substituents other than phenyl in
position 2 toward human MetAP2 (measured after 30 min of incubation). The most
significant inhibition is highlighted in bold
O
O
N
Se
R
N
Se
R
Entry
R
IC50 [lM]
Entry
R
IC50 [lM]
1n
1o
H
Me
4.97 0.25
10.4 1.2
1a, ebselen
1b
2.43 0.35
24.3 2.2
1p
0.32 0.09
HO
1c
10.2 1.3
1q
1r
8.85 1.32
7.57 0.42
MeO
t-Bu
1d
9.98 0.51
OMe
1e
1f
2.07 0.19
6.27 0.85
O-n-Bu
F
with glutathione in physiological conditions. Thus, measuring the
activity of 2,20-dicarbamoyldiaryl diselenides is of fundamental
value in the context of in vivo studies.
1g
1h
0.935 0.13
10.9 1.9
F
The collection of compounds was screened for the inhibition of
three neutral metallo-aminopeptidases, APN, LAP and MetAP2 (see
Supplementary data). Organoselenium products were found to be
good, generally low-micromolar-range, slow-binding inhibitors of
exclusively MetAP2. They were inactive toward APN and LAP
below inhibitor concentration 2 mM, apparently due to the lack
of specific interactions within their binding sites. Steady-state
binding of the inhibitors to the active site of MetAP2 was achieved
over time. The slow binding mechanism was not defined, the
organoselenium compounds showed a mixed type A and B mech-
anism. The plot of the first-order rate constant (kapp) versus inhibi-
tor concentration ([I]) was neither linear (type A) nor hyperbolic
(type B, see Supplementary data). Therefore, it was not possible
to distinguish between mechanism type A (reversible slow-bind-
ing) and type B (enzyme isomerization).50 The non-covalent, com-
petitive mode of inhibition clearly predominates. This was
confirmed by regain of the enzymatic activity after dilution exper-
iments and dialysis. However, we do not exclude conformational
changes upon covalent modifications of solvent-exposed cysteine
residues. To avoid uncertainties we decided to present the inhibi-
MeO
MeO
1i
1j
0.92 0.16
1.35 0.45
OMe
1k
1l
2.67 0.31
0.121 0.066
2.35 0.16
t-Bu
Cl
MeO
1m
Cl
F
CO2H
H
N
A832234, the reference
compound
S
0.01126
O
O
N
tory potency as the half maximal inhibitory concentration (IC50
)
determined after 30 min incubation of enzyme with an inhibitor
(Tables 1–3).
Selenazolones of the first group appeared moderate to good
inhibitors of methionine aminopeptidase 2, with nearly all of the
IC50 constants in the low micromolar range, e.g. 2.43 lM for the
The compounds employed for inhibitory studies were formally
divided into three groups. The first group included ebselen and 2-
phenylbenzisoselenazol-3(2H)-ones with the phenyl rings mono-
or disubstituted with different residues and functional groups,
such as Me, t-Bu, OH, OMe, n-BuO, and halogens (1a–m). The sec-
ond group (compounds 1n–r) comprised analogues of ebselen
based on the benzisoselenazol-3(2H)-one core modified in position
2 (the nitrogen atom). Instead of phenyl, the benzisoselenazolone
system remained unsubstituted (1n) or it was substituted by
methyl (1o), benzyl (1p), p-tert-butylbenzyl (1q) or phenylethyl
(1r). The structure of the third group of the studied compounds
(2a, 2c–e, 2l–n) was based on diselenide, the acyclic form of ebse-
len. Seven derivatives were chosen to investigate the mechanism
by which this modification influences the activity. As mentioned
in the introductory part the dimeric Se–Se form of ebselen has
been assigned as the product of benzisoselenazolone interference
lead ebselen (Table 1). The inhibitory potency varied within an
order of magnitude up and down to ebselen depending on charac-
ter and position of substitution. For example the affinity of com-
pound 1b with methyl substitution in the para position is
diminished ten-fold compared to 1a. In the case of substitution
by hydroxy (1c) or methoxy group (1d) at the ortho position, the
activity also decreased (four-fold). A methoxy group in the para
position (1e) did not influence the inhibitory potency, whereas a
butoxy group in the para position (1f) reduced the activity. There-
fore, electron-donating functional groups were generally not well
tolerated. The without strongly electron-withdrawing fluoride sub-
stitution exerted an ambiguous influence on the affinity of ligand.
Fluoride in the meta position (1g) increased the inhibitory potency
to the submicromolar range. However, the opposite effect was
Please cite this article in press as: We˛glarz-Tomczak, E.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.09.050

4
E. We˛glarz-Tomczak et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
Table 3
tem can be proposed as the principal hydrophobic contacts. The
oxygen atom of the selenazole ring coordinates well with one of
the active site metal ions. The close Oꢂ ꢂ ꢂCo distance is 1.13 Å while
the other contact is not tight (3.43 Å).
Indiscriminate docking results were obtained upon modeling
interactions of the most potent inhibitor 1l with the MetAP2 active
site. In this case more than one conformation showed comparable
thermodynamic stability. The binding mode corresponding to that
described for ebselen was one of those possibilities. A conforma-
tion characterized by the rotated plane of benzoisoselenazolone
Inhibitory activity for diselenides, the acyclic forms of ebselen and analogues, toward
human MetAP2 (measured after 30 min of incubation)
O
R
N
H
Se
2
Entry
R
IC50 [lM]
2a
0.23 0.04
HO
2c
5.44 0.71
MeO
2d
2e
2l
6.79 0.23
13.9 0.23
119.2 4.8
OMe
Cl
MeO
2m
4.47 0.38
Cl
2n
2p
H
9.13 0.96
3.43 0.63
measured for 1h with F in the para position. The combination of a
methyl in the meta position with a methoxy group in the non-adja-
cent ortho position in compound 1i position resulted in a three-fold
improvement in the IC50 value to 920 nM. The substitution of the
compound 1d with chlorine (1m) improved the inhibitory potency
by four-fold. A similar modification in the case of compound 1b
resulted in the best inhibitor in this study (1l, IC50 = 121 nM).
Replacing phenyl in position 2 with less hydrophobic sub-
stituents, hydrogen (1n) or methyl (1o), was not beneficial
(Table 2). The inhibition constants indicated two- and four-fold
drop in activity, respectively. In contrast, benzyl substitution (1p)
resulted in a very active inhibitor with IC50 = 320 nM, improved
by almost one order of magnitude compared with ebselen.
Interestingly, there is an increase in affinity only in case of the
compound with benzyl substitution. Further extension of the
hydrophobic fragment to p-tert-butylbenzyl (1q) or phenylethyl
(1r) was no more profitable.
The impact of the opening of isoselenazolones on the inhibitory
activity was also tested. Interestingly, the acyclic diselenide form
of ebselen (2a) was found to be one order of magnitude more
potent than 1a. For other cases there is no essential difference in
affinity between compounds 1c–e, 1m, 1n and 1p and correspond-
ing diselenides. Only for the o-chloro-p-methylphenyl substituent,
the acyclic counterpart (2l) proved to be three orders of magnitude
less active than 1l.
The SAR discussion was further illustrated by molecular model-
ing study on the binding mode of the most significant ligands with
human MetAP2. This study confirmed the binding to the active site
and thus, competitive mechanism of inhibition. The modeled inter-
actions of ebselen (1a) shows that the molecule occupies an inter-
section between putative S1 and S10 regions, with the N-phenyl
ring buried deeper, and adopts a bent-shaped conformation which
fits well to the hydrophobic environment (Fig. 2A and Graphical
Abstract). Edge-to-face interactions between surrounding imida-
zoles and aromatic rings of the inhibitor, namely His231 with
N-phenyl and both His231 and His331 with the ligand bicyclic sys-
Figure 1. The privilege conformation of ebselen (A) and alternative conformations
in the model of the complex of ebselen derivative 1l (B and C) with human
MetAP219 (PDB ID code 5D6E). Metal–oxygen interactions are marked in gray.
Please cite this article in press as: We˛glarz-Tomczak, E.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.09.050

E. We˛glarz-Tomczak et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
5
micromolar and submicromolar range. The structure optimization
of the lead compound resulted in an improvement of the inhibitory
potency from 2.43 lM to 0.121 lM. Increasing the size of the
N-phenyl fragment of ebselen by disubstitution with a 2-chloro
and a 4-methyl group was so beneficial. Extension of N-phenyl
group to N-benzyl also resulted in a very potent inhibitor with
an inhibition constant of 0.32 lM. Among the group of diselenides,
the acyclic forms of the benzisoselenazolones, the opened form of
ebselen was the most active: the IC50 value decreased by one order
of magnitude compared to the lead, reaching 0.23 lM. Our discov-
ery may result in a widening range of applications of the antioxi-
dant drug ebselen as an antiangiogenic agent. Ebselen has the
following two undoubted advantages: (1) it has passed clinical tri-
als, and there are no barriers to using it in humans; (2) it is selec-
tive toward MetAP2 versus other neutral aminopeptidases such as
APNs and LAP. It must be remembered that in the human organ-
ism, benzisoselenazolones, glutathione peroxidase mimetics, are
transformed to the corresponding diselenides31,32 which, as we
showed here, can also bind to MetAP2 even with higher affinity.
Thus, ebselen can be considered as the drug and a prodrug at the
same time. The identified active organoselenium compounds may
be useful tools in basic studies on the function of MetAP2 and prof-
itable in the development of new generations of therapeutic
agents.
Figure 2. A model of the complex of compound 2a with human MetAP219 (PDB ID
code 5D6E). The hydrophobic active site surface is marked in blue (hydrophilic in
brown), metal–oxygen interactions are marked in gray, intermolecular hydrogen
bond ligand–NHꢂ ꢂ ꢂAsp262 carboxylate is marked in green.
ring was identified as the second, almost equally favorable option
(Fig. 1B). In this case the selenium side of the bicyclic system is
pointed toward cobalt ions. However, the minimal energy was cal-
culated for a reversed arrangement, with the selenazolone frag-
ment buried deeper in the binding pocket (Fig. 1C). This
conformation does not demand any fold of the structure, the rings
are only slightly twisted to each other. The key hydrophobic con-
tacts with His231 and His331 are well conserved in both the
rotated and inversed version. An increased lipophilic surface
(2-chloride and 4-methyl groups) of the inhibitor gives rise to fur-
ther stacking involving mainly Ile338, and additionally with
Phe219 for the most privileged conformation depicted in Figure 1C.
Importantly, in the last-mentioned arrangement, chloride coordi-
nates with the cobalt ions (2.26 Å and 3.01 Å). These favorable
structural features seems to be responsible for an improved activ-
Acknowledgments
This work was financed by a statutory activity subsidy from the
Polish Ministry of Science and Higher Education for the Faculty of
Chemistry of Wrocław University of Technology, and by Wrocław
Centre of Biotechnology, program The Leading National Research
Centre (KNOW) for years 2014–2018. The Biovia Discovery Studio
package was used under a Polish country-wide license. The use of
software resources (Biovia Discovery Studio program package) of
the Wrocław Centre for Networking and Supercomputing is
acknowledged. We also gratefully acknowledge Michał Talma for
his assistance with molecular modeling.
Supplementary data
ity of 1l compared with ebselen (IC50 = 0.121 vs 2.43 lM).
The opening of the selenazole ring at the selenium-nitrogen
bond and dimerization increased the inhibitory potency of ebselen
by one order of magnitude (IC50 = 0.23 lM for diselenide 2a). In the
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2016.09.
050.
arrangement predicted by molecular modeling, the typical coordi-
nation between the cobalt ions and the oxygen of the carbonyl
group are well reproduced (compare Fig. 1A). The distances are
shorter than for 1a, 1.14 Å and 3.01 Å, respectively. Moreover, a
hydrogen bond was identified between the nitrogen atom of the
ligand amide group and the carboxyl of Asp262 (2.10 Å). Aromatic
rings interact tightly with the surrounding residues (Ala141,
Phe219, His231, Ala230, Asp251, Leu328, Ile338 and Tyr444)
References and notes
1. Matsui, M.; Fowler, J. H.; Walling, L. L. Biol. Chem. 2006, 387, 1535.
2. Sato, Y. Biol. Pharm. Bull. 2004, 27, 772.
3. Selvakumar, P.; Lakshmikuttyamma, A.; Dimmock, J. R.; Sharma, R. K. Biochim.
Biophys. Acta. 2006, 1765, 148.
4. Mina-Osorio, P. Trends Mol. Med. 2008, 14, 361.
5. Leszczyniecka, M.; Bhatia, U.; Cueto, M.; Nirmala, N. R.; Towbin, H.; Vattay, A.;
Wang, B.; Zabludoff, S.; Phillips, P. E. Oncogene 2006, 25, 3471.
6. Wiltschi, B.; Merkel, L.; Budisa, N. Chembiochem 2009, 10, 217.
7. Frottin, F.; Martinez, A.; Peynot, P.; Mitra, S.; Holz, R. C.; Giglione, C.; Meinnel, T.
Mol. Cell Proteomics 2006, 5, 2336.
8. Yang, G.; Kirkpatrick, R. B.; Ho, T.; Zhang, G.-F.; Liang, P. H.; Johanson, K. O.;
Casper, D. J.; Doyle, M. L.; Marino, J. P., Jr.; Thompson, S. K.; Chen, W.; Tew, D.
G.; Meek, T. D. Biochemistry 2001, 40, 10645.
9. Wang, J.; Sheppard, G. S.; Lou, P.; Kawai, M.; Park, C.; Egan, D. A.; Schneider, A.;
Bouska, J.; Lesniewski, R.; Henkin, J. Biochemistry 2003, 42, 5035.
10. Lowther, W. T.; Zhang, Y.; Sampson, P. B.; Honek, J. F.; Matthews, B. W.
Biochemistry 1999, 38, 14810.
11. Wang, J.; Lou, P.; Henkin, J. J. Cell Biochem. 2000, 77, 465.
12. Kanno, T.; Endo, H.; Takeuchi, K.; Morishita, Y.; Fukayama, M.; Mori, S. Lab.
Invest. 2002, 82, 839.
employing the whole range of p–p, p–alkyl and p–anion contacts
(outlined in Fig. 2 as a fit to the hydrophobic surface, see also Graph-
ical Abstracts). For diselenides even the simplest analogue 2a fits
extremely tightly to the MetAP2 binding site, thus, any further
extension can alter the complementarity. Indeed, only 2c and 2d,
containing an oxygen atom in the ortho position, slightly improved
the potency. The improvement was correlated with hydrogen bond-
ing formation, e.g. with Asn329 (2.72 Å) for 2c, however, to achieve
this contact the diselenide backbone is forced to adopt a sterically
demanded arrangement.
The inhibition of human methionine aminopeptidase 2 has been
identified as a crucial way to inhibit angiogenesis during the
growth and metastasis of solid tumors.9,15 Here, we reported a
well-known antioxidant drug ebselen21 and its derivatives/
analogues as inhibitors of MetAP2 with IC50 values in the low
13. Wu, S.; Gupta, S.; Chatterjee, N.; Hileman, R. E.; Kinzy, T. G.; Denslow, N. D.;
Merrick, W. C.; Chakrabarti, D.; Osterman, J. C.; Gupta, N. K. J. Biol. Chem. 1993,
268, 10796.
14. Catalano, A.; Romano, M.; Robuffo, I.; Strizzi, L.; Procopio, A. Am. J. Pathol. 2001,
159, 721.
15. Shusterman, S.; Maris, J. M. Cancer Lett. 2005, 228, 171.
Please cite this article in press as: We˛glarz-Tomczak, E.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.09.050

6
E. We˛glarz-Tomczak et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
16. Selvakumar, P.; Lakshmikuttyamma, A.; Kanthan, R.; Kanthan, S. C.; Dimmock,
J. R.; Sharma, R. K. Clin. Cancer Res. 2004, 10, 2771.
28. Yin, S. Q.; Wang, J. J.; Zhang, C. M.; Liu, Z. P. Curr. Med. Chem. 2012, 19, 1021.
29. Schewe, T. Gen. Pharmacol. 1995, 26, 1153.
17. Griffith, E. C.; Su, Z.; Turk, B. E.; Chen, S.; Chang, Y.-H.; Wu, Z.; Biemann, K.; Liu,
J. O. Chem. Biol. 1997, 4, 461.
18. Logothetis, C. J.; Wu, K. K.; Finn, L. D.; Daliani, D.; Figg, W.; Ghaddar, H.;
Gutterman, J. U. Clin. Cancer Res. 2001, 7, 1198.
19. Morgen, M.; Joest, C.; Malz, M.; Janowski, R.; Niessing, D.; Klein, C. D.; Gunkel,
N.; Miller, A. K. ACS Chem. Biol. 2016, 11, 1001.
20. Zhou, G.; Tsai, C. W.; Liu, J. O. J. Med. Chem. 2003, 46, 3452.
21. Sheppard, G. S.; Wang, J.; Kawai, M.; BaMaung, N. Y.; Craig, R. A.; Erickson, S. A.;
Lynch, L.; Patel, J.; Yang, F.; Searle, X. B.; Lou, P.; Park, C.; Kim, K. H.; Henkin, J.;
Lesniewski, R. Bioorg. Med. Chem. Lett. 2004, 14, 865.
22. Kallander, L. S.; Lu, Q.; Chen, W.; Tomaszek, T.; Yang, G.; Tew, D.; Meek, T. D.;
Hofmann, G. A.; Schulz-Pritchard, C. K.; Smith, W. W.; Janson, C. A.; Ryan, M. D.;
Zhang, G. F.; Johanson, K. O.; Kirkpatrick, R. B.; Ho, T. F.; Fisher, P. W.; Mattern,
M. R.; Johnson, R. K.; Hansbury, M. J.; Winkler, J. D.; Ward, K. W.; Veber, D. F.;
Thompson, S. K. J. Med. Chem. 2005, 48, 5644.
23. McBride, C.; Cheruvallath, Z.; Komandla, M.; Tang, M.; Farrell, P.; Lawson, J. D.;
Vanderpool, D.; Wu, Y.; Dougan, D. R.; Plonowski, A.; Holub, C.; Larson, C.
Bioorg. Med. Chem. Lett. 2016, 26, 2779.
30. Sies, H.; Masumoto, H. Adv. Pharmacol. 1996, 38, 229.
31. Mugesh, G.; Singh, H. B. Chem. Soc. Rev. 2000, 29, 347.
32. Bhabak, K. P.; Mugesh, G. Acc. Chem. Res. 2010, 43, 1408.
33. Batna, A.; Fuchs, C.; Spiteller, G. Chem. Phys. Lipids 1997, 87, 149.
34. Chew, P.; Yuen, D. Y.; Stefanovic, N.; Pete, J.; Coughlan, M. T.; Jandeleit-Dahm,
K. A.; Thomas, M. C.; Rosenfeldt, F.; Cooper, M. E.; de Haan, J. B. Diabetes 2010,
59, 3198.
35. Chew, P.; Yuen, D. Y.; Koh, P.; Stefanovic, N.; Febbraio, M. A.; Kola, I.; Cooper, M.
E.; de Haan, J. B. Arterioscler. Thromb. Vasc. Biol. 2009, 29, 823.
36. Kushwah, N.; Prabhakar, A.; Prasad, D.; Ravi, K.; Singh, S. B.; Khan, N. BMRJ
2015, 8, 644.
37. Mukherjee, S.; Weiner, W. S.; Schroeder, C. E.; Simpson, D. S.; Hanson, A. M.;
Sweeney, N. L.; Marvin, R. K.; Ndjomou, J.; Kolli, R.; Isailovic, D.; Schoenen, F. J.;
Frick, D. N. ACS Chem. Biol. 2014, 9, 2393.
38. Hanavan, P. D.; Borges, C. R.; Katchman, B. A.; Faigel, D. O.; Ho, T. H.; Ma, C.-T.;
Sergienko, E. A.; Meurice, N.; Petit, J. L.; Lake, D. F. Oncotarget 2015, 6, 18418.
39. Yan, J.; Guo, Y.; Wang, Y.; Mao, F.; Huang, L.; Li, X. Eur. J. Med. Chem. 2015, 95,
220.
24. Kawai, M.; BaMaung, N. Y.; Fidanze, S. D.; Erickson, S. A.; Tedrow, J. S.; Sanders,
W. J.; Vasudevan, A.; Park, C.; Hutchins, C.; Comess, K. M.; Kalvin, D.; Wang, J.;
Zhang, Q.; Lou, P.; Tucker-Garcia, L.; Bouska, J.; Bell, R. L.; Lesniewski, R.;
Henkin, J.; Sheppard, G. S. Bioorg. Med. Chem. Lett. 2006, 16, 3574.
25. Wang, G. T.; Mantei, R. A.; Kawai, M.; Tedrow, J. S.; Barnes, D. M.; Wang, J.;
Zhang, Q.; Lou, P.; Garcia, L. A.; Bouska, J.; Yates, M.; Park, C.; Judge, R. A.;
Lesniewski, R.; Sheppard, G. S.; Bell, R. L. Bioorg. Med. Chem. Lett. 2007, 17, 2817.
26. Sheppard, G. S.; Wang, J.; Kawai, M.; Fidanze, S. D.; BaMaung, N. Y.; Erickson, S.
A.; Barnes, D. M.; Tedrow, J. S.; Kolaczkowski, L.; Vasudevan, A.; Park, D. C.;
Wang, G. T.; Sanders, W. J.; Mantei, R. A.; Palazzo, F.; Tucker-Garcia, L.; Lou, P.;
Zhang, Q.; Park, C. H.; Kim, K. H.; Petros, A.; Olejniczak, E.; Nettesheim, D.;
Hajduk, P.; Henkin, J.; Lesniewski, R.; Davidsen, S. K.; Bell, R. L. J. Med. Chem.
2006, 49, 3832.
40. Li, B. L.; Li, B.; Zhang, R. L.; Zhao, J. J.; Wang, X. F.; Liu, Y. M.; Shi, Y. P.; Liu, J. B.;
Chen, B. Q. Bioorg. Med. Chem. Lett. 2016, 26, 1279.
41. Kim, I. Y.; Stadtman, T. C. Proc. Natl. Acad. Sci. U.S.A. 1997, 94, 12904.
42. Gopalakrishna, R.; Gundimeda, U.; Chen, Z. H. Arch. Biochem. Biophys. 1997, 348,
25.
43. Koharyova, M.; Kollarova, M. Gen. Physiol. Biophys. 2008, 27, 71.
44. Engman, L.; Hallberg, A. J. Org. Chem. 1989, 54, 2964.
45. Palus, J.; Młochowski, J.; Juchniewicz, L. Pol. J. Chem. 1998, 72, 1931.
46. Młochowski, J.; Gryglewski, G. J.; Inglot, A. D.; Jakubowski, A.; Juchniewicz, L.;
Kloc, K. Liebigs Ann. Chem. 1996, 1751.
47. Młochowski, J.; Kloc, K.; Syper, L.; Inglot, A. D.; Piasecki, E. Liebigs Ann. Chem.
1993, 1239.
48. Młochowski, J.; Brza˛szcz, M.; Chojnacka, M.; Giurg, M.; Wójtowicz, H. Arkivoc
2004, 3, 226.
49. Lesser, R.; Weiss, R. Ber. Dtsch. Chem. Ges. 1924, 57, 1077.
50. Pawełczak, M.; Hurek, J. Chemik 2014, 68, 377.
27. Towbin, H.; Bair, K. W.; DeCaprio, J. A.; Eck, M. J.; Kim, S.; Kinder, F. R.; Morollo,
A.; Mueller, D. R.; Schindler, P.; Song, H. K.; van Oostrum, J.; Versace, R. W.;
Voshol, H.; Wood, J.; Zabludoff, S.; Phillips, P. E. J. Biol. Chem. 2003, 278, 52964.
Please cite this article in press as: We˛glarz-Tomczak, E.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.09.050