Synthesis of a Hybrid between SOD Mimetic and Ebselen to Target Oxidative Stress

Article abstract of DOI:10.1002/ejic.201900407

Oxidative stress emerged as target in drug discovery due to its diverse role in various diseases, such as cardiovascular, neurodegenerative, and autoimmune diseases. During the last decades, considerable progress was made in the development of compounds with the ability to reduce reactive oxygen species (ROS) like superoxide. However, the dismutation of the latter leads to formation of another harmful ROS, hydrogen peroxide, which can be depleted through peroxidase activity. The present work describes the synthesis of a hybrid, which unifies a superoxide dismutase mimetic, Mn^IIpyane, and a glutathione peroxidase mimetic, ebselen, that are connected via an amide bond. This unique hybrid is designed in order to convert superoxide into oxygen and water, i.e. as a potential biological agent for complete ROS removal and will be used in the future as mechanistic molecular tool for further elucidation of (patho) physiological consequences of ROS removal.

Full text of DOI:10.1002/ejic.201900407

A Journal of
Accepted Article
Title: Synthesis of a Hybrid between SOD Mimetic and Ebselen to
Target Oxidative Stress
Authors: Nicole Orth, Andreas Scheitler, Verena Josef, Alicja Franke,
Achim Zahl, and Ivana Ivanovic-Burmazovic
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To be cited as: Eur. J. Inorg. Chem. 10.1002/ejic.201900407
Link to VoR: http://dx.doi.org/10.1002/ejic.201900407

European Journal of Inorganic Chemistry
10.1002/ejic.201900407
COMMUNICATION
Synthesis of a Hybrid between SOD Mimetic and Ebselen to
Target Oxidative Stress
Nicole Orth, Andreas Scheitler, Verena Josef, Alicja Franke, Achim Zahl and Ivana Ivanović-
Burmazović*[a]
Abstract: Oxidative stress emerged as target in drug discovery due
to its diverse role in various diseases, such as cardiovascular,
neurodegenerative and autoimmune diseases. During the last
decades, considerable progress was made in the development of
compounds with the ability to reduce reactive oxygen species (ROS)
like superoxide. However, the dismutation of the latter leads to
formation of another harmful ROS, hydrogen peroxide, which can be
depleted through peroxidase activity. The present work describes the
synthesis of a hybrid, which unifies a superoxide dismutase mimetic
Hydrogen peroxide in general can damage biomolecules. It can
impair the function of enzymes involved in energy metabolism and
is capable of DNA damage by its reaction with transition metal
ions or by radiation with UV light to form hydroxyl radical.[
Hence, the presence of peroxide cleaving enzymes like
glutathione peroxidase (GPx) utilizing glutathione (GSH) as
reductant is crucial for maintaining healthy ROS levels in the body
(Scheme 2).[5]
1, 4]
(Mn(II)pyane) and a glutathione peroxidase mimetic (ebselen) that are
connected via an amide bond. This unique hybrid is designed in order
to convert superoxide into oxygen and water, i.e. as a potential
biological agent for complete ROS removal, and will be used in the
future as mechanistic molecular tool for further elucidation of (patho)
physiological consequences of ROS removal.
Scheme 2. GPx catalyzed reaction of peroxide with glutathione GSH.[5]
The organoselenenium compound ebselen (Figure 1) has gained
large interest, not merely due to its GPx mimicking abilities, but
furthermore, since it is widely applicable as biological antioxidant
in the treatment of neurodegeneration, bipolar disease, diabetes,
cancer and due to its antibacterial, antimycotic and antiviral
effects.[6] The only drawback is the sparse solubility in aqueous
media causing difficulties in administration as a drug.[6e]
The superoxide anion radical is considered to be a reactive
oxygen species (ROS).[1] Although it naturally occurs in the body
as a product of autoxidation or enzyme catalyzed oxidative
processes, such as those driven by xanthine oxidase,
lipoxygenase, cyclooxygenase or NADPH dependent oxidase,
and it is involved in signaling pathways of synaptic plasticity and
memory formation, an imbalance of superoxide is linked to aging
Among the superoxide dismutase mimetics (SODm),
manganese(II) pentaazamacrocycles (MnPAMs) were observed
to have remarkable SOD activity and beneficial biological
effects.[7] Mn(II)pyane is a prototype of MnPAMs, which has been
processes, cancer, cardiovascular and neurodegenerative
diseases.[1-2] The enzyme superoxide dismutase (SOD) should
balance its steady-state concentration in living organisms by
catalyzing superoxide decomposition to hydrogen peroxide and
oxygen (Scheme 1).[3]
extensively studied in chemical settings as well as in cellular
milieu and in vivo.[
7e, 8]
It has also been functionalized by us for the
purposes of different studies.[8b, 8c, 9] One of these derivatives
includes a primary amine residue suitable as a linker for
hybridization with another moiety.[8b, 9a]
Scheme 1. Disproportion of superoxide to hydrogen peroxide and oxygen via
superoxide dismutase (SOD).[3]
[
a]
N. Orth, A. Scheitler, V. Josef, Dr. Alicja Franke, Dr. Achim Zahl,
Prof. Dr. I. Ivanović-Burmazović
Department of Chemistry and Pharmacy
University of Erlangen-Nuremberg
Egerlandstrasse 1, 91058 Erlangen (Germany)
E-mail: ivana.ivanovic-burmazovic@fau.de
Figure 1. Structures of ebselen (left) Mn(II)pyane (middle) and Mn(II)pyane-
2 2
CH NH
(right).[
6a, 8b]
Supporting information for this article can be found under:
The combination of two or more active agents covalently bonded
is long known as the concept of hybridization and represents a
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European Journal of Inorganic Chemistry
10.1002/ejic.201900407
COMMUNICATION
sophisticated approach aiming at several targets.[10] In the past,
several examples have demonstrated the advantages of
hybridization regarding improved biological activity with remaining
minimal toxicity and potential to overcome drug resistances.[10-11]
To eliminate oxidative stress and its accompanying damage, we
as the one for its mother-compound Mn(II)pyane (Figure S20).
The GPx activity of the hybrid was determined using the
reductase coupled assay, which resulted in a catalytic rate of
-1
-1
k
cat = 0.022 ± 0.003 M s (Figures S21). By way of comparison,
for ebselen alone we obtained a catalytic rate of kcat = 0.02 ± 0.01
-1 -1
designed a hybrid molecule of Mn(II)pyane-CH
2
NH
2
and ebselen
M s (Figure S22), being in accordance with the literature value
-
1 -1 [17]
(
Figure 2) to target both superoxide, with the former, and
(kcat = 0.0216 M s ).
Thus, as we could experimentally
hydrogen peroxide, with the latter, potentially resulting in water
and oxygen as products, i.e. complete ROS removal. Compounds
with this synergistic mode of action include manganese salen
observe the SOD and GPx activity of the two bioactive units
remains unchanged upon their chemical coupling. This is an
important property required for the future studies, in order to
correctly evaluate a possible differences between biological
effects of the hybrid and its separate components (Mn(II)pyane
and ebselen), as well as their mixtures.
complexes
(EUK
series),
porphyrins,
peptides
and
organoselenium-modified β-cyclodextrins to name a few and
represent a promising class of ROS scavengers.[12]
Figure 2. Newly synthesized Mn(II)pyane/ebselen hybrid (6).
All synthetic steps for the hybrid are described in Figure 3.
Starting from benzocaine, which represents
available starting material, ethyl 4-(2-iodobenzamido)benzoate
1) is formed in reaction with the activated acid chloride 2-
a cheap and
(
[
13]
iodobenzoyl chloride in yields of 90 %. The ring closure to form
the isoselenazolone 2 was achieved in good yields by preparation
t
of KSeO Bu with selenium powder and potassium tert-butoxide in
dimethylformamide to react with 1.[14] Quantitative hydrolysis of
the ethyl ester was achieved with 5 equivalents of lithium
hydroxide to yield ebselen acid (3) in a mixture of tetrahydrofuran,
methanol and water.[15] To activate the carboxylic acid function,
N-hydroxysuccinimide in combination with (1-Ethyl-3-(3-
dimethylaminopropyl)carbodiimide hydrochloride (EDCxHCl) was
used and extraction with water facilitated the removal of the urea
byproduct.[16] After purification by column chromatography the N-
hydroxysuccinimide ester can finally be coupled to the amine of
2 2
the Mn(II)pyane-CH NH (5), which was synthesized by our group
earlier.[8b] The formed Mn(II)pyane/ebselen hybrid 6 is extracted
with water from the organic phase, which reveals that ebselen’s
insolubility in polar solvents is eradicated by coupling with the
Figure 3. Synthesis of Mn(II)pyane/ebselen hybrid 6. Reaction conditions: a)
[
6e]
[13]
t
Mn(II)pyane moiety.
All compounds were characterized by
NEt
3
(1.0 eq) in DCM, RT, 24 h;
b) KO Bu (1.6 eq) and selenium powder
c) LiOH x H O (5.0 eq) in THF/MeOH/H
[
14]
elemental analysis, UHR ESI-MS and 1H and 13C NMR,
(1.3 eq) in DMF, 60 °C, 48 h;
2
2
O
15]
(
2:1:1), RT, 4 days, acidic workup; [ d) N-Hydroxysuccinimide (1.5 eq) and
respectively. The spectra are included in the supporting
information.
EDCxHCl (1.5 eq) in DCM/THF (1:1), 67 °C, 17 h, washed with water; [18] e) in
DCM and MeOH (2:1), RT, 72 h, extraction with water.[
8b, 16, 18]
We have also studied the SOD and GPx activity of the hybrid. The
obtained SOD activity, with a catalytic rate of kcat = 1.35(1) x 107
-1 -1
M s (MOPS buffer, pH 7.4), is - within the error limit- the same
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European Journal of Inorganic Chemistry
10.1002/ejic.201900407
COMMUNICATION
In conclusion, we describe the synthesis of a Mn(II)pyane/ebselen
hybrid with a potential synergic SOD and GPx activity combined
in one molecule. The two separate functionalities Mn(II)pyane and
ebselen were studied for years and are promising candidates in
balancing ROS: the former catalyzes the conversion of
superoxide to water and hydrogen peroxide, the latter, in
combination with the cellular widely available tripeptide
glutathione, is capable of cleaving the peroxide, protecting
against cell damage. Solubility in water of the hybrid 6 is a major
advantage regarding future in vitro and in vivo testing and is an
enormous improvement when compared to ebselen. The
molecule delivers new opportunities as a potential agent against
two significant reactive oxygen species, superoxide and hydrogen
peroxide and as such, it could be applied as mechanistic tool to
further decipher the role of these two ROS entities in physiological
and pathophysiological processes. Knowing that the applicability
of both components ranges from activity against cancer,
Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral
sclerosis (ALS), bipolar disorder, diabetes over the process of
aging to antibacterial, antimycotic and antiviral effects, it is
expected that the obtained hybrid will become an attractive object
for diverse studies.[2b, 6] In future investigations, its activity will be
compared with that of its pure components as well as with the
activity of their mixture. In that way we will be able to judge
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European Journal of Inorganic Chemistry
10.1002/ejic.201900407
COMMUNICATION
Suggestion for the Entry for the Table of Contents
COMMUNICATION
Newly synthesized tool against oxidative
stress with dual functionality.
N. Orth, A. Scheitler, Verena Josef,
Alicja Franke, A. Zahl, I. Ivanović-
Burmazović*
The synthesis of a molecule that unifies
moieties
mimicking
superoxide
Page No. – Page No.
dismutase and glutathione peroxidase is
described. The hybrid delivers new
opportunities as a potential agent against
two significant reactive oxygen species,
superoxide and hydrogen peroxide,
playing a crucial role in various diseases.
Synthesis of a Hybrid between SOD
Mimetic and Ebselen to Target
Oxidative Stress
This article is protected by copyright. All rights reserved.