Pyridinylimidazoles as GSK3β Inhibitors: The Impact of Tautomerism on Compound Activity via Water Networks

Article abstract of DOI:10.1021/acsmedchemlett.9b00177

Glycogen synthase kinase-3β (GSK3β) is involved in many pathological conditions and represents an attractive drug target. We previously reported dual GSK3β/p38α mitogen-activated protein kinase inhibitors and identified N-(4-(4-(4-fluorophenyl)-2-methyl-1

Full text of DOI:10.1021/acsmedchemlett.9b00177

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Letter
Pyridinylimidazoles as GSK3# inhibitors: the impact of
tautomerism on compound activity via water networks
Fabian Heider, Tatu Pantsar, Mark Kudolo, Francesco Ansideri, Angela De Simone, Letizia Pruccoli, Taiane
Schneider, Marcia Inês Goettert, Andrea Tarozzi, Vincenza Andrisano, Stefan A. Laufer, and Pierre Koch
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.9b00177 • Publication Date (Web): 26 Aug 2019
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Pyridinylimidazoles as GSK3β inhibitors: the impact of tautomerism
on compound activity via water networks
Fabian Heider†#, Tatu Pantsar‡§#, Mark Kudolo†, Francesco Ansideri†, Angela De Simone∥, Letizia
Pruccoli∥, Taiane Schneider▽, Marcia Inês Goettert▽, Andrea Tarozzi∥, Vincenza Andrisano∥, Stefan
A. Laufer†, Pierre Koch†⊥∗
†Department of Pharmaceutical and Medicinal Chemistry, Institute of Pharmaceutical Sciences, Eberhard Karls
Universität Tübingen, Auf der Morgenstelle 8, 72076 Tübingen, Germany.
‡School of Pharmacy, University of Eastern Finland, P.O.BOX 1627, 70211 Kuopio, Finland.
§ Department of Internal Medicine VIII, University Hospital Tübingen, Otfried-Müller-Straße 14, 72076 Tübingen,
Germany.
∥Department for Life Quality Studies, Alma Mater Studiorum-University of Bologna, Corso D’Augusto, 237, 47921
Rimini, Italy.
▽Cell Culture Laboratory, Postgraduate Program in Biotechnology, University of Vale do Taquari (Univates), Lajeado,
Brazil
⊥Department of Pharmaceutical/Medicinal Chemistry II, Institute of Pharmacy, University of Regensburg,
Universitätsstraße 31, 93053 Regensburg, Germany.
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KEYWORDS. Protein Kinase Inhibitors; Glycogen Synthase Kinase-3β; Pyridinylimidazoles; Tautomerism; Molecular
Dynamics Simulation; Quantum Mechanics.
ABSTRACT: Glycogen synthase kinase-3β (GSK3β) is involved in many pathological conditions and represents an attractive
drug target. We previously reported dual GSK3β/p38α mitogen-activated protein kinase inhibitors and identified N-(4-(4-(4-
fluorophenyl)-2-methyl-1H-imidazol-5-yl)pyridin-2-yl)cyclopropanecarboxamide (1) as a potent dual inhibitor of both
target kinases. In this study, we aimed to design selective GSK3β inhibitors based on our pyridinylimidazole scaffold. Our
efforts resulted in several novel and potent GSK3β inhibitors with IC50 values in the low nanomolar range. 5-(2-
(Cyclopropanecarboxamido)pyridin-4-yl)-4-cyclopropyl-1H-imidazole-2-carboxamide (6g) displayed very good kinase
selectivity as well as metabolically stability and inhibits GSK3β activity in neuronal SH-SY5Y cells. Interestingly, we observed
the importance of the 2-methylimidazole’s tautomeric state for the compound activity. Finally, we reveal how this crucial
tautomerism effect is surmounted by imidazole-2-carboxamides, which are able to stabilize the binding via enhanced water
network interactions, regardless of their tautomeric state.
Glycogen Synthase Kinase 3β (GSK3β) is a ubiquitously
expressed serine/threonine kinase, which plays an
important role in a variety of different cell signaling
pathways. GSK3β plays a crucial role in almost every
pathway leading to the hallmarks of Alzheimer’s disease1-2
and is often referred to as a tau-kinase due to its capacity to
modulate tau hyperphosphorylation. Overactivity of GSK3β
has also been connected to an increased production of β-
amyloids,3 neuroinflammation and oxidative stress.4
(2), which might be located in the hydrophobic region (HR)
I of the ATP binding site, resulted in a significantly reduced
GSK3β inhibition with a complete loss of activity against
p38α MAPK. In this study, our aim was to further improve
the activity of our pyridinylimidazole scaffold while shifting
the selectivity towards GSK3β.
F
N
N
GSK3β has also been associated with a plethora of other
pathological conditions such as diabetes,5 cancer,6-8
schizophrenia,9 bipolar disorders10 and osteoporosis.11
Thus, GSK3β is considered to be an attractive drug target.
CH3
CH3
N
N
H
H
N
N
HN
HN
We recently reported a series of pyridinylimidazoles as
dual GSK3β/ p38α MAP kinase (MAPK) inhibitors and
identified trisubstituted imidazole 1 as a potent balanced
inhibitor of both target enzymes (Figure 1).12 Furthermore,
we observed that the removal of the para-fluorophenyl ring
1
2
O
O
IC50 (GSK3) = 0.053 µM
IC50 (p38) = 0.019 µM
IC50 (GSK3) = 1.68 µM
IC50 (p38) > 10 µM
Figure 1. Pyridinylimidazole-based lead compounds 1 and 2.
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Recently, employing quantum mechanics (QM) to drug
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compounds (Figure 2, see SI for details). Indeed, QM results
indicated that the active compounds generally prefer the
tautomer A or at least represent a reasonable population of
this tautomeric state (Table S3). For instance, the low
nanomolar inhibitors 3a, 3b, 3d and 3e, display a clear
preference for tautomer A (>71.5%). In turn, the Less active
compounds 3c and 3j-m, display a clearly diminished
population of tautomer A (<17%).
1
2
3
4
5
6
7
8
design and development has become increasingly popular.
For instance, QM can be utilized to improve docking and
scoring, determining protonation states and optimizing
structures as well as ligand binding energies.13-14 Also, the
importance of water in drug design is gaining more and
more emphasis.15 The effect of water network stabilization
for ligand binding has been demonstrated e.g. by Klebe and
coworkers.16 In addition, molecular dynamics (MD)
simulations offer valuable insights into ligand binding
interactions.17 By utilizing QM calculations with MD
simulations we disclosed the importance of tautomerism
and water networks for the activity of our
pyridinylimidazole compounds. In this case, the observed
SAR could not have been clarified by simplified
computational tools, such as docking, which has major
caveats especially related to the solvent effects and
dynamics of the system18 that were found determining for
the activity differences among imidazoles.
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Results and discussion
Detailed descriptions of the synthetic sequences are
reported in Schemes S1-S14 (Supporting Information, SI).
To address the alarming diffuse trend of increasing the
inhibitor logP value in the lead optimization,19 we
monitored the lipophilic ligand efficacy (LLE) of the
synthesized compounds.20 The LLE of our lead compounds
1 and 2 was already high with values of 5.10 and 4.40,
respectively (Table 1).
Figure 2. (A) The imidazole ring has two potential tautomeric
forms: in tautomer A, the nitrogen next to the R1-group is
unprotonated and can act as a H-bond acceptor, whereas in
tautomer B it is protonated and can act as a H-bond donor. (B)
The R1-group influences the preferred tautomeric state and
conformation. As an example, the lowest energy conformations
(in solution) of the highly active compound 3d and of the
poorly active compound 3l are shown here. Compound 3d
prefers the active conformation with tautomer A and 3l exists
in the inactive conformation with tautomer B.
Initially, we examined the influence of different
substituents reaching into the HR I of GSK3β. To this end,
we synthesized a series of 2-methylpyridinylimidazoles
with different (cyclo)aliphatic and aromatic moieties
attached to the imidazole-C4 position.
Replacing the aromatic ring with cycloalkyl moieties at
the imidazole-C4 position (3j-m) resulted in a substantial
loss of activity, leading to modest inhibitors of GSK3β in the
micromolar range displaying complete inactivity against
p38α MAPK.
Obviously, the preference of a specific tautomeric state
does not fully determine the compound activity. For
example, the inactive compound 3g, clearly prefers
tautomer A (86.9%), but the pyrimidine group is
suboptimal for the hydrophobic region (solvent
preference). On the contrary, the highly lipophilic para-
fluorophenyl substituent in compound 1 clearly increases
potency, despite its preference for tautomer B.
Interestingly, the inactive 2-methoxyphenyl derivative 3f
appears only in a specific conformation as a tautomer A,
wherein the methoxy-group folds on top of the pyridinyl
ring (Table S3), which most likely impedes the binding.
Overall, the tautomeric state preference partially, but not
solely, determines the 2-methylimidazole activity.
Compounds with bulky moieties, such as 2-naphtyl (3h),
were inactive, probably because of a steric clash in the HR I.
Replacement of the para-fluorophenyl ring with the 5-
membered heteroaromatic rings thiophene (3d) or furan
(3e) as well as other minor changes on the para-
fluorophenyl ring, such as addition of a methyl (3o) or a
second fluorine atom (3i), led to inhibitors with slightly
increased IC50 values compared to 1. Introduction of a
pyrimidine (3g) at the imidazole-C4 led to a completely
inactive derivative. The less lipophilic ortho- and meta-
hydroxyphenyl derivatives (3a and 3b, respectively) turned
out to be potent GSK3β inhibitors with sound LLE values,
while the para-hydroxyphenyl compound (3c) displayed
substantially diminished inhibition against both kinases. All
potent GSK3β inhibitors bearing an aromatic ring in the HR
I, however, remained potent inhibitors of p38α MAPK,
except for 3e with 10-fold selectivity for GSK3β.
Next, we attempted to improve the binding affinity of 1 via
enhancing interactions at the solvent interface in the HR II.
To this end, MD simulations (200 ns) demonstrated the
potential suitability of compounds bearing N-(pyridin-2-
yl)tri- or tetrazolepropanamide moieties (4a,b; Figures S2–
S4, SI). Both displayed cation-π interactions with the
Arg141 and improved solvent interactions combined with
significantly lower log P values (Table 2) (see SI for synthetic
details). The simulation of 4a highlighted an identical
binding mode for the triazole ring as observed in a crystal
structure (PDB ID: 5K5N).21
To elucidate the observed dramatic loss of activity of
certain compounds, we investigated the influence of the R1-
substituent on the imidazole ring’s tautomeric state. To this
end, we conducted QM calculations to assess the probability
of different tautomeric states and conformations for the
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Table 1. Activity and Physicochemical Parameters of 2-Methylimidazoles 3a-o.
2
3
R1
N
4
5
6
CH3
N
H
N
7
8
9
HN
O
Cpd
R1
IC50 ± SEM [µM]
IC50 ± SEM [µM]
p38α MAPK
Alog Pb
LLE
Tautomer A
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11
12
13
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60
GSK3βa
population (%)c d
1
2
F
0.053 ± 0.012d
1.68 ± 0.12d
0.019d,e
>10d,e
2.88
0.72
4.40
5.10
29.386
n.d.f
H
OH
3a
3b
0.011 ± 0.001
0.043 ± 0.005
0.050
0.024
2.40
2.40
5.56
4.97
85.508
HO
2.834 (93.885g)
3c
3d
3e
HO
0.893 ± 0.001
0.069 ± 0.000
0.099 ± 0.033
1.851
0.048
0.987
2.40
2.40
1.84
3.65
4.77
5.17
12.687
81.318
71.467
S
O
O CH3
3f
>10
0.504
2.65
-
31.19
N
N
3g
3h
>10
>10
>10
0.89
3.58
-
-
85.892
20.289
0.081
F
3i
0.059 ± 0.007
0.019
3.08
4.15
n.d.f
F
3j
3k
3l
3.09 ± 0.30
4.11 ± 0.19
5.46 ± 0.01
4.64 ± 1.16
>10
>10
>10
>10
1.76
2.22
2.68
3.13
3.75
3.17
2.59
2.20
6.014
10.229
16.734
16.552
3m
O
3n
0.467 ± 0.006
0.030
2.44
3.89
n.d.
O
H3C
3o
0.117 ± 0.015
0.007
3.36
3.57
n.d.
F
an=2; bcalculated with Canvas (Schrödinger LLC)22; caccording to QM Conformer & Tautomer Predictor of Maestro (Schrödinger,
LLC, New York, NY, 2018) (see SI and Table S3 for details); dvalues taken from Heider et al.12; determined by ELISA activity assay23;
e
gthe intramolecular H-bond to amide conformation excluded (see Table S3); fn.d. = not determined.
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Table 2. Activity and Physicochemical Parameters of N-(pyridin-2-yl)tri- or tetrazolepropanamide bearing 2-
Methylimidazoles 4a-d.
1
2
3
R1
N
4
5
6
7
8
9
CH3
N
H
N
HN
R2
O
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IC50 ± SEM [µM]
IC50 ± SEM [µM]
p38
Cpd
4a
R1
R2
N
N
GSK3βa
Alog Pb
LLE
F
F
0.082 ± 0.007
0.072 ± 0.008
5.18 ± 0.10
0.041
0.038
>10
1.16
5.92
N
N
N
N
4b
4c
4d
N
N
1.83
-0.99
-0.32
5.32
6.27
5.65
N
N
N
H
N
N
N
H
4.42 ± 0.29
>10
an=2; bcalculated with Canvas (Schrödinger LLC)22.
Compounds 4a and 4b show a similar potency as the lead
compound 1 but with enhanced LLE values. Removal of the
para-fluorophenyl anchor resulted in compounds 4c and
4d, both displaying substantially reduced inhibitory
potency. This clearly results from the negative log P values
of these compounds, which seems to compromise their
binding affinity (entropic penalty). Nevertheless, these
compounds still exhibit mediocre target inhibition and fit
nicely into the SAR of the series.
The most striking differences existed in cycloalkyl
substituted compounds 6f and 6g exhibiting dramatically
improved potency against GSK3β along with higher LLE
values compared to the corresponding 2-methylimidazole
derivatives 3m and 3j, which displayed only mediocre
activities and preferred the tautomer B. Moreover, all three
compounds showed significant selectivity over p38α MAPK,
and compound 6g was among the best from this series
(GSK3β, IC50: 0.003 µM; p38α MAPK, IC50: >10 µM; LLE
7.64).
To overcome the highlighted tautomerism-related issues
observed with the 2-methylimidazoles, we designed and
synthesized a series of imidazole-2-carboxamides. Instead
of the acceptor nitrogen of tautomer A, the 2-carboxamides
could neglect the tautomeric state of the imidazole by
presenting the amide oxygen towards the Lys85 region.
This amino acid side chain has been successfully targeted by
carbonyl groups, e.g. Pfizer disclosed 6-amino-4-
(pyrimidin-4-yl)pyridones interacting with Lys85,24 while
To further investigate these dramatic activity differences,
we first confirmed that the amide group replacing the
methyl group on the imidazole-C2 position had no influence
on the tautomeric state preference (Table S3). As an
example, the cyclopropyl-substituted compounds 3j and 6g
display an analogous population of the tautomeric state A,
namely 6.0% and 6.7% for 3j and 6g, respectively.
Nevertheless, the potency of these two inhibitors is
dramatically different, with the imidazole-2-carboxamide
derivative 6g showing a three order of magnitude higher
activity than its methyl counterpart (3j).
Bristol-Myers
Squibb
reported
potent
pyrrolopyridinones.25 Moreover, we investigated ethyl
esters as well as a hydroxyl moiety for their suitability to
address the Lys85 residue.
To gain a deeper insight into the compound binding and the
activity differences between 2-methylimidazoles and
imidazole-2-carboxamides, we conducted a total of 8 µs MD
simulations for the selected compounds bound to GSK3β in
their preferred tautomeric state: 3j and 6g in tautomeric
state B and the 3a and 6b in tautomeric state A. The initial
1 µs MD simulations suggested unstable binding only for 3j,
where its lipophilic cyclopropyl group is exposed to water
in HR-I (Figure 3 and Figures S5 and S9, SI). Whereas with
6g the amide stabilizes a water network near Asp200 and
the cyclopropyl group is shielded from the solvent allowing
the stable binding of tautomer B (Figure 3 and Figures S5
and S9, SI). With tautomer A preferring 3a and 6b, the 2-
hydroxyphenyl was shielded from solvent regardless of the
methyl or amide group substituent in the imidazole ring
(Figure 3 and Figures S7 and S9, SI). These observations
In contrast to methylimidazole 2, imidazole2-
carboxamide 6a showed a >35-fold improvement in
potency (LLE 7.49) (Table 2). In case of compound 1, the
introduction of a carboxamide moiety at the imidazole-C2
position (6h) did not substantially improve the inhibitory
activity. Installation of an ethyl ester (5a and 5c) yielded
mediocre inhibitors highlighting the importance of the
amide function. Introduction of a hydroxy moiety at the
imidazole-C2 methyl group resulted in 6i showing a two-
fold reduction in GSK3β inhibition and no shift in the IC50
value of p38α MAPK. In most cases, imidazole-2-
carboxamides were better inhibitors of GSK3β than their
corresponding 2-methylimidazole counterparts (e.g. 6c vs
3f, 6e vs 3o). Only the already potent 2-hydroxyphenyl 3a
(vs 6b) displayed no improvement in activity.
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were confirmed in unbiased simulations, conducted using
another crystal structure as the starting configuration
(Figures S6, S8 and S10, SI). Based on these data, the
energetically favorable tautomer B of 3j does not support
the suggested stabilizing interactions with the dynamic
water network, which leads to water exposed HR-I, whereas
the preferred tautomer A of 3a is capable to shield the HR-I
from water via direct or water mediated interactions to
Lys85 and maintain a stable binding (Figures S5–S8, SI).
Thus, QM calculations with the MD simulations provide a
potential explanation for the observed activity differences.
1
2
3
4
5
6
7
8
Table 3. Inhibition Data and Physicochemical Parameters of Ethyl Imidazole-2-carboxylates 5 and Imidazole-2-
carboxamides 6-8.
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F
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11
12
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R1
N
N
N
O
O
Br
R2
N
N
N
NH2
NH2
H
H
H
N
N
N
HN
HN
HN
CH3
7
8
O
O
O
5 6
,
IC50 ± SEM [µM]
IC50 ± SEM [µM]
Cpd
5a
R1
R2
GSK3βa
p38α MAPK
Alog Pb
LLE
O
H
CH3
0.739 ± 0.186
0.047 ± 0.020
0.899 ± 0.010
0.039 ± 0.017
0.091 ± 0.006
0.013 ± 0.001
>10
>10c
1.07
-0.16
3.23
1.99
2.20
0.12
5.06
O
O
H
6a
5c
6h
6i
7.49
2.82
5.52
4.84
7.76
NH2
O
O
CH3
F
0.089
O
F
F
0.019 ± 0.002c
0.016
NH2
OH
O
H3C
6j
>10
NH2
O
6g
6f
0.003 ± 0.000
0.003 ± 0.000
>10
>10
0.88
2.25
7.64
6.27
NH2
O
NH2
O
OH
6b
6c
6e
6d
0.023 ± 0.001
0.265 ± 0.017
0.079 ± 0.003
0.352 ± 0.002
0.158
2.35
1.52
1.77
2.48
3.91
6.12
4.81
4.62
2.54
NH2
O
O CH3
NH2
O
H3C
0.016
2.04
F
NH2
O
F3C
O
NH2
7
8
0.354d
>10
0.59
1.24
5.86
6.09
-
-
-
0.047 ± 0.004
0.117
-
an=2; bcalculated with Canvas (Schrödinger LLC)22; cdetermined by ELISA activity assay23; dn=1 .
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Figure 3. Representative snapshots from MD simulations of compounds 3i (A), 3a (B), 6g (C) and 6b (D). (E) Compound 3i
appears in a shifted binding orientation compared to 6g, whereas both 2-hydroxyphenyl derivatives (F) 3a and 6b display similar
binding orientation in the simulations. The shift in the binding orientation of compound 3j occurs due to direct H-bond interaction
from the imidazole to Asp200. This interaction, with the increased solvent exposure of the lipophilic cyclopropyl group (see Figures
S5–S6, SI), explains the three orders of magnitude difference in activity between 3j and 6g. The protein surface is illustrated in
transparent light blue color and hydrogen bonds with yellow dashed lines in A-D.
Selected compounds (1, 3a, 3i, 6a and 6h) were further
tested for their GSK3β affinity in a previously reported ESI-
QTOF assay (Tables S1 and S2, SI).26 Using this completely
different assay system, the potency trend of these GSK3β
inhibitors obtained in the ADP-Glo activity assay was
confirmed.
relevant CYP isoforms (Table 4). At a test concentration of
10 μM, imidazole-2-carboxamide 6g shows a clean CYP
inhibition profile. Only low inhibition of CYP1A2 was
observed.
Table 4. Inhibition of CYP450 isoenzymes.
% inhibition of CYP isoform @ 10 µM
Moreover, inhibitors 3a and 6g were tested for their
metabolic stability by incubation with human liver
microsomes (HLM) over a period of 4 h (Tables S4 and S5,
SI). Both compounds displayed excellent metabolic stability
in this assay.
Cpd
6g
1A2
2C9
2C19
2D6
3A4
25.5
0.8
-2.0
-2.8
-2.3
Further pharmacological profiling of the potent GSK3β
inhibitor 6g included the evaluation of its ability to inhibit
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ACS Medicinal Chemistry Letters
To assess the overall kinome selectivity, most promising
towards pharmacologically relevant CYP isoenzymes.
Importantly, the LLE values of the series illustrate that the
series’ potency is not driven by molecular obesity.29
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inhibitor 6g was screened against a representative panel of
68 diverse kinases (Table S6, SI), including the target kinase
GSK3β and all members of the MAP kinases. At a
concentration of 0.5 µM (>160-fold its IC50 value on the
target kinase) only CDK2, CDK9, JNK3, MLK2 and VEGFR2
were substantially inhibited, suggesting an acceptable
kinome selectivity.
The SAR of the synthesized compounds was explained
with QM calculations and MD simulations. The series
represent an interesting example of the influence of the 2-
methylimidazole tautomerism on the compound activity.
The effect of tautomerism was indirectly surmounted by
introducing the water-network stabilizing 2-carboxamide,
thus, exemplifying the importance to consider that subtle
molecular differences may have significant influence on the
dynamics of the system.
To confirm the biological activity of imidazole-
2carboxamide 6g, we tested it in a cell-based GSK3β assay.
At the tested concentration of 1 µM, 6g inhibits GSK3β
activity, in terms of inactive phospho-GSK3α/β (Ser21/9)
increase, after 1 h of treatment in neuronal SH-SY5Y cells
(Figure 4).
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ASSOCIATED CONTENT
Supporting Information
1.0
The Supporting Information is available free of charge on the
ACS Publications website.
***
0.8
0.6
0.4
0.2
0.0
Experimental details for preparation of the compounds; ESI-
QTOF assay; cell-based GSK3β assay; molecular modeling, QM
calculations and MD simulations; metabolic stability in HLM;
kinome selectivity screening; inhibition of CYP450 isoenzymes;
cell toxicity data (PDF).
QM output conformations, MD movies and full raw trajectories
are
freely
available
at
https://doi.org/10.5281/zenodo.3362889.
r
g
6
t
AUTHOR INFORMATION
C
Corresponding Author
Figure 4. Inhibition of GSK3β activity in neuronal SH-SY5Y
cells. Cells were incubated with compound 6g [1 µM] for 1 h. At
the end of incubation, the phosphorylation of GSK3α/β
(Ser21/9) (inactive GSK3α/β form) was determined by western
blotting. Data are expressed as ratio between phospho-
GSK3α/β and total GSK3β levels normalized against β-Actin
and reported as mean ± SD of at least three independent
experiments (*** p < 0.001 versus untreated cells; t-test).
*pierre.koch@uni-tuebingen.de (Pierre Koch)
ORCID
Fabian Heider: https://orcid.org/0000-0003-2472-2457
Tatu Pantsar: https://orcid.org/0000-0002-0369-2909
Letizia Pruccoli: https://orcid.org/0000-0002-0739-2102
Andrea Tarozzi: https://orcid.org/0000-0001-7983-8575
Pierre Koch: https://orcid.org/0000-0003-4620-4650
Author Contributions
Since CDK2, CDK9 and VEGFR2 are off-targets of 6g, we
also determined its cytotoxic profile on different cell lines
after 48 h of incubation. A margin of safety is given
concerning cytotoxic side effects. In case of tested non-
tumorigenic cells, the concentration to cause a 50%
decrease in cell viability is in the low micromolar range,
which corresponds >1000-fold the IC50 value of the GSK3β
kinase activity assay (Figure S11, SI).
#F.H. and T.P. contributed equally to this work.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENT
We thank Jens Strobach for his assistance in the p38α MAPK
ADP Glo™ assay. The authors wish to acknowledge CSC – IT
Center for Science, Finland, for computational resources. T.P.
acknowledges the Orion Research Foundation sr for financial
support.
In case of the selected tumorigenic cells (Figure S12, SI),
compound 6g shows antiproliferative activity in breast
human cancer cell line (Figure S12B, SI), at 0.1 µM and less
at 0.01 µM. This may be a relevant result, as the GSK3β is a
target in the treatment of human breast cancer.27 Breast
cancer patients with overexpression of GSK3β presented
poor prognosis, and GSK3β inhibition suppressed the
viability and proliferation of breast cancer cells in vitro.28
ABBREVIATIONS
GSK, glycogen synthase kinase; HR, hydrophobic region; MAPK,
MAP kinase; LLE, lipophilic ligand efficacy; MD, molecular
dynamics; QM; quantum mechanics.
In summary, we synthesized a diverse set of 33 novel di-
and trisubstituted pyridinylimidazoles. The most potent
GSK3β inhibitors 6f, 6g and 6j were selective over p38α
MAPK and had reasonable (CNS) drug-like log P values.
Imidazole 6g was metabolically stable in HLM, displayed a
very good selectivity profile, and showed no affinity
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Pyridinylimidazoles as GSK3β inhibitors: the impact of tautomer-ism on compound activity via water
networks
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Fabian Heider, Tatu Pantsar, Mark Kudolo, Francesco Ansideri, Angela De Simone, Letizia Pruccoli,
Taiane Schneider, Marcia Inês Goettert, Andrea Tarozzi, Vincenza Andrisano, Stefan A. Laufer,
Pierre Koch
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