Anticonvulsant and analgesic in neuropathic pain activity in a group of new aminoalkanol derivatives

Article abstract of DOI:10.1016/j.bmcl.2020.127325

As part of the presented research, thirteen new aminoalkanol derivatives were designed and obtained by chemical synthesis. In vivo studies (mice, i.p.) showed anticonvulsant activity (MES) of nine compounds, and in the case of one compound (R,S-trans-2-((2-(2,3,5-trimethylphenoxy)ethyl)amino)cyclohexan-1-ol, 4) both anticonvulsant (ED₅₀ MES = 15.67 mg/kg, TD₅₀ rotarod = 78.30 mg.kg, PI = 5.00) and analgesic activity (OXA-induced neuropathic pain, active at 15 mg/kg). For selected active compounds additional in vitro studies have been performed, including receptor studies (5-HT_1A), evaluation of antioxidant activity (DPPH assay), metabolism studies as well as safety panel (mutagenicity, safety in relation to the gastrointestinal flora, cytotoxicity towards astrocytes as well as impact on their proliferation and cell cycle).

Full text of DOI:10.1016/j.bmcl.2020.127325

Journal Pre-proofs
Anticonvulsant and analgesic in neuropathic pain activity in a group of new
aminoalkanol derivatives
Katarzyna Pańczyk, Anna Rapacz, Anna Furgała-Wojas, Kinga Sałat, Paulina
Koczurkiewicz-Adamczyk, Martyna Łucjanek, Iwona Skiba-Kurek, Elżbieta
Karczewska, Aleksandra Sowa, Dorota Żelaszczyk, Agata Siwek, Justyna
Popió ł, Elżbieta Pękala, Henryk Marona, Anna Waszkielewicz
PII:
S0960-894X(20)30436-4
DOI:
Reference:
https://doi.org/10.1016/j.bmcl.2020.127325
BMCL 127325
To appear in:
Bioorganic & Medicinal Chemistry Letters
Received Date:
Revised Date:
Accepted Date:
1 May 2020
1 June 2020
4 June 2020
Please cite this article as: Pańczyk, K., Rapacz, A., Furgała-Wojas, A., Sałat, K., Koczurkiewicz-Adamczyk, P.,
Łucjanek, M., Skiba-Kurek, I., Karczewska, E., Sowa, A., Żelaszczyk, D., Siwek, A., Popió ł, J., Pękala, E.,
Marona, H., Waszkielewicz, A., Anticonvulsant and analgesic in neuropathic pain activity in a group of new
aminoalkanol derivatives, Bioorganic & Medicinal Chemistry Letters (2020), doi: https://doi.org/10.1016/j.bmcl.
2020.127325
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Anticonvulsant and analgesic in neuropathic pain
activity in a group of new aminoalkanol derivatives
Leave this area blank for abstract info.
Katarzyna Pańczyk, Anna Rapacz, Anna Furgała-Wojas, Kinga Sałat, Paulina Koczurkiewicz-Adamczyk,
Martyna Łucjanek, Iwona Skiba-Kurek, Elżbieta Karczewska, Aleksandra Sowa, Dorota Żelaszczyk, Agata
Siwek, Justyna Popiół, Elżbieta Pękala, Henryk Marona and Anna Waszkielewicz
ED50 MES = 15.67 (9.10 - 26.97) mg/kg b.w. (mice, i.p.)
TD50 rotarod = 78.30 (6.82 - 93.13) mg/kg b.w. (mice, i.p.)
PI = 5.00
H3C
O
N
H
OH
R,S-trans
CH3
analgesic at 15 mg/kg b.w. (OXA-induced pain, von Frey test, mice, i.p.)
CH3

Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com
Anticonvulsant and analgesic in neuropathic pain activity in a group of new
aminoalkanol derivatives
Katarzyna Pańczyka, Anna Rapaczb Anna Furgała-Wojasb, Kinga Sałatb, Paulina Koczurkiewicz-
,
Adamczykc, Martyna Łucjanekd, Iwona Skiba-Kurekd, Elżbieta Karczewskad, Aleksandra Sowaa, Dorota
Żelaszczyka, Agata Siweke, Justyna Popiółc, Elżbieta Pękalac, Henryk Maronaa and Anna Waszkielewicza
aJagiellonian University Medical College, Faculty of Pharmacy, Department of Bioorganic Chemistry, Medyczna 9, 30-688 Kraków, Poland
bJagiellonian University Medical College, Faculty of Pharmacy, Department of Pharmacodynamics, Medyczna 9, 30-688 Kraków, Poland
cJagiellonian University Medical College, Faculty of Pharmacy, Department of Pharmaceutical Biochemistry, Medyczna 9, 30-688 Kraków, Poland
dJagiellonian University Medical College, Faculty of Pharmacy, Department of Pharmaceutical Microbiology, Medyczna 9, 30-688 Kraków, Poland
eJagiellonian University Medical College, Faculty of Pharmacy, Department of Pharmacobiology, Medyczna 9, 30-688 Kraków, Poland
ARTICLE INFO
ABSTRACT
Article history:
Received
Revised
Accepted
Available online
As part of the presented research, thirteen new aminoalkanol derivatives were designed and
obtained by chemical synthesis. In vivo studies (mice, i.p.) showed anticonvulsant activity (MES)
of nine compounds, and in the case of one compound (R,S-trans-2-((2-(2,3,5-
trimethylphenoxy)ethyl)amino)cyclohexan-1-ol, 4) both anticonvulsant (ED50 MES = 15.67
mg/kg, TD50 rotarod = 78.30 mg.kg, PI = 5.00) and analgesic activity (OXA-induced neuropathic
pain, active at 15 mg/kg). For selected active compounds additional in vitro studies have been
performed, including receptor studies (5-HT1A), evaluation of antioxidant activity (DPPH assay),
metabolism studies as well as safety panel (mutagenicity, safety in relation to the gastrointestinal
flora, cytotoxicity towards astrocytes as well as impact on their proliferation and cell cycle).
Keywords:
astrocytes
aminoalkanol
analgesic
anticonvulsant
cytotoxicity
intestinal flora
metabolism
mutagenicity
2009 Elsevier Ltd. All rights reserved.
———
 Corresponding author. Tel.: +48-12-620-55-76; e-mail: awaszkie@cm-uj.krakow.pl

Central nervous system (CNS) diseases, including epilepsy and
neuropathic pain, are both health and social challenges.
Epidemiological data suggest that neuropathic pain concerns about
7-10% of society,1 while epilepsy – 1%.2 Despite availability of a
wide range of anticonvulsants and pain killers on the market, a
large proportion of patients still does not get satisfying health
improvement.3,4 Lack of full knowledge about the
pathophysiology of CNS disorders, and thus the lack of
identification of some molecular targets for drugs, as well as the
occurrence of adverse effects and drug interactions during
treatment with available medications may be responsible for the
lack of full effectiveness of therapy.
Some antiepileptic drugs, such as gabapentin or pregabalin, are
often used in the treatment of neuropathic pain,13 which is a
premise for the study of anticonvulsants also for the analgesic
activity. Compound III has also been the subject of extensive
research on the mechanism of its action. The data indicate the
participation of sigma receptors and the serotonergic system.
OH
CH3
O
N
CH3
H
HO
CH3
R-(-)
CH3
*
*
O
III
N
H
ED50 MES=5.34 mg/kg
TD50 rotarod=29.48 mg/kg
R,S-trans
CH3
I
The above facts are a premise to look for new centrally-active
compounds. Research on the mechanisms of action of active
compounds and the implementation of safety and metabolism
studies in the early stages of research are particularly important.
The multidirectional central activity of some compounds is also an
interesting research direction. Many drugs on the market show
diverse effects depending on the dose and this diversity is
sometimes used in the treatment of CNS diseases. For example,
antiepileptic drugs are used in the treatment of neuropathic pain
(e.g. gabapentin, pregabalin).5,6 This phenomenon is explained,
among others, by the common pathomechanism of epilepsy and
neuropathic pain, which also contributes to their frequent
comorbidity.7
PI=5.52
ED50 MES=7.73 mg/kg
TD50 rotarod=36.23 mg/kg
PI=3.90
ED50 thermal hyperalgesia (STZ)=15.8 mg/kg
ED50 mechanical allodynia (STZ)=0.26 mg/kg
sigma (non-selective) IC50=1.9*10-6
5-HT1A IC50=2.8*10-6
5-HT transporter IC50=7.0*10-6
M
OH
M
CH3
M
O
N
CH3
OH
*
H3C
CH3
H
N
O
II
O
ED50 MES=20.87 mg/kg
TD50 rotarod=81.95 mg/kg
PI=3.93
R,S
CH3
IV
ED50 MES=13.03 mg/kg
TD50 rotarod=36.00 mg/kg
PI=2.76
Figure 2. Chemical structures and the central activity (mice, i.p.) of previously
synthesized reference compounds: I,10 II,11 III,12 IV.11 STZ – streptozotocin-
induced neuropathic pain model; reference drugs:5,14,15 Valproic acid – ED50
(MES, mice, i.p.) = 272 mg/kg, ED50 (MES, rats, i.p.) = 212 mg/kg;
Carbamazepine – ED50 (MES, mice, i.p.) = 8.81 mg/kg, ED50 (MES, rats, i.p.)
= 3.4 mg/kg; Phenytoin – ED50 (MES, mice, i.p.) = 9.50 mg/kg, ED50 (MES,
rats, i.p.) = 11.3 mg/kg.
The detection of anticonvulsant activity showed by anti-
arrhythmic drugs such as propranolol8 and mexiletine9 was
important for the study of aminoalkanol derivatives for CNS
activity (Figure 1).
For the design of the presented new compounds (Scheme 1,
Table 1), the aminoalkanol moiety initially included a distance of
two carbon atoms between hydroxy and amino groups (e.g. 2-
aminopropan-1-ol, 2-aminobutan-1-ol, 2-aminocyclohexan-1-ol,
piperidin-3-ol). Longer chains were the most often introduced in
order to compare the activity of isomers (e.g. 4-aminocyclohexan-
1-ol vs 2-aminocyclohexan-1-ol, piperidin-4-ol vs piperidin-3-ol).
An interesting aspect was also the use of alicyclic aminoalcohol
(piperidinol) in place of the chain one (aminocyclohexanol).
Figure 1. Anticonvulsant activity (maximal electroshock seizure test, MES
test) of anti-arrhythmic drugs: propranolol8 and mexiletine9 (aminoalkanol
component framed).
An important stage in the design of biologically active
compounds is in silico research, which allows prediction of
selected physicochemical parameters and exclusion of structures
with low probability of in vivo activity, e.g. due to low
bioavailability. The compounds designed in this work were
subjected to in silico analysis using the Molinspiration platform.16
Among the calculated values of physicochemical parameters
important for oral bioavailability were parameters covered by the
Lipinski’s rule: molecular weight (MW), logP coefficient, number
of hydrogen bond acceptors (N/O), number of hydrogen bond
Compounds obtained so far at the Department of Bioorganic
Chemistry were derivatives of the appropriate chiral and achiral
aminoalkanols, among which many compounds were
characterized by a high anticonvulsant activity in animal models,
as well as analgesic activity in neuropathic pain models. Some
pharmacological parameters, such as the median effective dose
(ED50) in a respective animal test, median toxic dose (TD50) in the
rotarod test or protective index (PI), were comparable or more
favorable than the results observed for the reference drugs. Among
all obtained active compounds in this chemical group, promising
results obtained for reference compounds I – IV10-12 (Figure 2)
became the premise for the synthesis of a series of compounds di-
or tri-substituted with methyl groups in the phenyl ring, presented
in this paper. The aminoalkanol moieties include among others
cyclic aminoalkanols present in the structure of compounds I and
II (i.e. cyclohexanol, piperidinol), while the linker is an alkyl chain
(with 2, 3 or 4 carbon atoms), or ethoxyethylene (as in the case of
compound IV).
donors (NH/OH). Compounds with
a predicted favorable
bioavailability profile should meet a minimum of four of the
following criteria: MW ≤ 500, logP ≤ 5, N/O ≤ 10, NH/OH ≤ 5,
which characterizes all designed compounds. All compounds meet
also criteria important for blood-brain barrier penetration (MW <
450, logP < 5, N/O < 7, NH/OH < 3).17 The exact values of the
calculated parameters are presented in the Supplementary material
(Table S1).
Chemistry
The multidirectional central activity of compound III is
noteworthy - in addition to the anticonvulsant activity of this
compound, analgesic activity was also observed in the
streptozotocin (STZ)-induced neuropathic pain model, which
corresponds to human type I diabetes-induced neuropathic pain.
All compounds were obtained by chemical synthesis (Scheme
1), via previously published methods.11 The substrates were
appropriately substituted phenols. In the first step of the synthesis,
the corresponding phenoxyalkyl- or phenoxyethoxyethyl

bromides were obtained in the Williamson reaction – a method
allowing obtaining an ether from properly substituted phenol and
an alkyl halide. Reaction of the corresponding phenols with 2-
chloroethan-1-ol, 3-chloropropan-1-ol, 4-chlorobutan-1-ol or 2-
(2-chloroethoxy)ethan-1-ol gave alcohols, which were then
substrates in the bromination reaction using phosphorus
tribromide. The resulting bromides were subjected to aminolysis
using the appropriate aminoalkanol. The reaction was carried out
in toluene in the presence of K2CO3 as a proton acceptor for
approximately 5 hours. For compounds 4, 7 and 13, the
aminoalkanol – R,S-trans-2-aminocyclohexan-1-ol – was obtained
synthetically via reaction of cyclohexene oxide with a 25%
aqueous ammonia solution. In other cases, aminoalkanols were
obtained from commercial sources. The obtained final products
were crystallized from petroleum ether, hexane or heptane (in case
of too low solubility, mixtures of these solvents with toluene were
used in the proportions 20:1 – 5:1). In order to achieve adequate
purity of compounds 9 and 10 it was necessary to use a
chromatography column (silica gel, eluent: methylene
chloride/methanol, 70:30 v/v) before the crystallization step. In the
case of compounds 6, 11 – 13, the obtained compounds were
converted into the salt form (hydrochloride) by saturating the
ethanolic solution of the compound with hydrogen chloride gas.
The obtained hydrochlorides were crystallized from a solvent
mixture (acetone/methanol 10:1 or acetone/ethanol 5:1). The
identity of all obtained compounds was confirmed by spectral
methods (NMR), while their purity and homogeneity by means of
chromatographic methods (e.g. TLC, LCMS). The melting point
of all substances was also determined. For optically active
compounds, specific rotation was measured. The detailed
description of synthetic procedures as well as physicochemical
data of title compounds are included in Supplementary material.
linker R2
linker R2
x HCl
OH
O
O
O
linker Br
a
c
b
R1
R1
R1
R1
1 10
-
6, 11 13
-
R1:
2,4-(CH3)2 for compd. - ,
linker:
4
-(CH2)2- for compd.
-(CH2)3- for compd.
-(CH2)4- for compd.
1 3 9 11
-
1 2 5 8
3
,
, -
12
2,6-(CH3)2 for compd.
2,3,5-(CH3)3 for compd.
2,4,6-(CH3)3 for compd.
4 5 13
,
,
9 13
-
-(CH2)2-O-(CH2)2- for compd.
6 8
-
R2 (aminoalkanol moiety):
H
1 6
for
,
4, 7 13
for
,
3 9
for
for
,
N
*
N
*
OH
*
OH
N
H
R,S
R,S-trans
H
CH3
CH3
R-(-)
OH
N
OH
S-(+)
11
for
OH
for
OH
H
N
CH3
2 5
,
OH
N
8, 12
10
for
trans
N
S-(+)
H
Scheme 1. Synthesis of compounds 1-13. Reagents: a: 1. Cl-linker-OH, K2CO3, acetone/EtOH, 2. PBr3; b: R2, K2CO3, toluene; c: HCl(g), EtOH.
Pharmacology
In vivo screening of the anticonvulsant activity was performed
with a use of the maximal electroshock seizure (MES) test (mice,
i.p., Table 1). Among the 13 tested compounds, 10 showed
anticonvulsant activity (ED50 ranging from 14.79 mg/kg for the
compound 7 to 52.49 mg/kg for the compound 3). All compounds
were also preliminary screened for potential neurotoxicity in the
rotarod test (mice, i.p.). Taking into account both anticonvulsant
activity and results of the rotarod test, compound 4 showed the
most advantageous pharmacological profile (PI = 5.00). For
selected compounds characterized by ED50 < 20 mg/kg in MES
test (4 – 7, 13) additional pharmacological studies were planned,
including the assessment of their antiallodynic and
antihyperalgesic properties in oxaliplatin (OXA)- or
streptozotocin (STZ)-induced neuropathic pain models
(compounds 4, 5, 13). Compound 4 attenuated tactile allodynia but
not cold hyperalgesia in both phases of OXA-induced neuropathy
(von Frey test: activity at a dose of 15 mg/kg, p < 0.0001 in the
early phase and p < 0.05 in the late one, mice, i.p., Table 2). Tested
compounds were inactive in the STZ-induced neuropathy model.
The detailed results are presented in Supplementary material
(Table S2).
Most of the antiepileptic drugs available on the market are the
result of (1) in vivo screening of structurally diverse compounds or
(2) structure optimization of the known antiepileptic drugs or
active compounds in preclinical studies. Rational molecular target-
oriented design was used only for compounds activating
GABAergic transmission (e.g. vigabatrin, tiagabine) and
glutaminergic receptor antagonists (e.g. perampanel).18 On the
other hand, data from 2011 and 2012 indicate that as much as 71%
of all new drugs approved by FDA resulted from the rational
design of substances with a specific mechanism of action. The
reason for this discrepancy is probably the lack of a sufficient
number
of
well-established
molecular
targets
for
anticonvulsants.19 Regarding the above facts, the presented study
uses the strategy of structural modification of anticonvulsant
compounds developed previously in the Department of Bioorganic
Chemistry, followed by in vivo screening of all obtained
compounds. A preliminary assessment of the potential mechanism
of action is planned only for selected compounds active in vivo.
The described procedure increases the chances of developing new
compounds active in vivo, at the same time being in line with the
current trend of searching for potential molecular targets for new
antiepileptic drugs.
Compounds 4 – 7 and 13 were also tested in vitro for potential
antioxidant activity (microplate method using DPPH free
radical).20 The premise for this type of research was the fact that

the oxidation-reduction equilibrium impairment is an important
pathological factor in the development of some central nervous
system diseases.21 An example of a registered drug whose main
mechanism of action is antioxidant activity is idebenone (a
coenzyme Q10 derivative). The compound was initially developed
towards the therapy of Alzheimer's disease and then
neuromuscular diseases. At present, its main indication is Leber's
hereditary optic neuropathy.22 The role of reactive oxygen species
is also emphasized in the context of the pathomechanism of
epilepsy, e.g. antioxidants have been postulated to potentially slow
down epileptogenesis in acquired epilepsy.23 None of the tested
compounds showed antioxidant activity in this test in the
concentration range of 125-1000 μM (test results available in
Supplementary material, Figure S1).
Table 1. Anticonvulsant activity of compounds 1 – 13 in the maximal electroshock seizure (MES) test and their neurotoxicity in
the rotarod test (mice, i.p.).
Compd
Chemical structure
Dose
[mg/kg]
MESa
Lethalit
y
Rotarodc
ED50 (confidence interval)
TD50 (confidence interval)
[mg/kg]
ED50 = 34.82 (27.46 – 44.16)
TD50 = 107.66 (95.29 – 121.64)
PId
in MESb
NT
1
30
40
50
100
115
130
2/6
4/6
5/6 (4/4)
NT
NT
NT
NT
1/4
2/6
4/6
5/6
3.09
O
N
*
NT
0/4
NT
NT
OH
OH
R,S
H3C
CH3
NT
NT
2
3
50
0/4
0/4
0/4
NT
NT
O
N
H3C
H3C
CH3
OH
30
50
70
120
150
200
1/6
3/6
4/6
NT
NT
NT
1/6
0/6
0/6
NT
NT
NT
2/6
1/6
1/6
2/6
4/6
5/6
ED50 = 52.49 (35.50 – 77.61)
TD50 = 135.99 (106.74 – 173.28)
2.59
O
N
CH3
R-(-)
H
CH3
HO
4
10
20
30
50
60
80
100
2/6
3/6
5/6
4/4
NT
NT
NT
NT
NT
NT
0/4
NT
NT
NT
2/6
3/6
5/6
1/4
1/6
3/6
5/6
ED50 = 15.67 (9.10 – 26.97)
TD50 = 78.30 (6.82 – 93.13)
5.00
*
H3C
O
*
N
H
CH3
R,S-trans
CH3
OH
5
6
7
10
20
30
45
60
100
0/6
4/6
6/6
NT
NT
NT
0/6
1/6
0/6
NT
NT
NT
NT
NT
0/6
0/6
4/6
6/6
ED50 = 17.71 (15.75 – 19.91)
TD50 = 63.40 (58.07 – 69.21)
3.58
3.86
2.90
H3C
O
N
CH3
CH3
CH3
x HCl
10
20
30
60
80
100
0/6
3/6
6/6
NT
NT
NT
0/6
2/6
0/6
NT
NT
NT
NT
NT
0/6
1/6
4/6
6/6
ED50 = 18.17 (16.14 – 20.46)
TD50 = 70.19 (64.55 – 76.32)
O
N
*
OH
R,S
H3C
CH3
CH3
OH
*
10
20
30
40
45
50
55
1/6
3/6
6/6
NT
NT
4/4
NT
NT
NT
NT
NT
NT
0/4
NT
NT
NT
NT
2/6
4/6
4/4
5/6
ED50 = 14.79 (12.24 – 17.88)
TD50 = 42.85 (36.69 – 50.05)
H
N
O
*
H3C
CH3
R,S-trans
CH3
8
9
30
0/6
6/6
0/6
NT
NT
H
N
O
H3C
H3C
CH3
OH
OH
trans
H
N
20
30
50
60
80
100
1/6
2/6
6/6
NT
NT
NT
0/6
0/6
0/6
NT
NT
NT
0/6
1/6
0/6
0/6
1/6
3/5 (1 death)
ED50 = 27.43 (23.75 – 31.69)
TD50 = 93.24 (84.84 – 102.47)
3.40
O
O
CH3
R-(-)
CH3

Compd
10
Chemical structure
Dose
[mg/kg]
MESa
Lethalit
y
Rotarodc
ED50 (confidence interval)
TD50 (confidence interval)
[mg/kg]
NT
PId
NT
in MESb
2/6
H
N
30
0/6
1/6
O
O
OH
CH3
S-(+)
S-(+)
H3C
H3C
CH3
O
x HCl
11
H
N
30
40
50
60
80
90
100
1/6
3/6
4/6 (2/4)
5/6
NT
NT
NT
1/4
NT
NT
NT
NT
NT
NT
0/4
NT
1/6
2/6
4/6
ED50 = 40.51 (31.90 – 51.44)
TD50 = 94.38 (84.99 – 104.81)
2.33
O
OH
CH3
CH3
NT
NT
x HCl
CH3
12
30
35
40
50
100
130
150
1/6
3/6
6/6
4/4
NT
NT
NT
NT
NT
NT
1/4
NT
NT
NT
NT
NT
NT
0/4
0/6
4/6
5/6
ED50 = 33.03 (31.52 – 34.62)
TD50 = 131.88 (123.87 – 140.40)
3.99
H
O
N
O
trans
CH3
OH
x HCl
13
10
20
30
40
50
60
0/6
3/6
6/6
NT
4/4
NT
NT
NT
NT
NT
0/4
NT
NT
NT
NT
1/6
2/6 (4/4)
4/6
ED50 = 18.17 (16.14 – 20.46)
TD50 = 54.21 (44.76 – 65.66)
2.91
8.56
OH
*
H
H3C
O
N
O
*
R,S-trans
CH3
CH3
Phenytoine
ED50 = 6.65 (5.42 – 8.16)
TD50 = 56.91 (48.53 – 66.74)
Lacosamide2
ED50 = 9.24(8.28– 10.30)
TD50 = 46.20(44.48–48.00)
5.00
4
MES – maximal electroshock seizure test; number of animals protected/number of animals tested; b number of lethal cases/number of animals tested; TD50
[mg/kg b.w.] or number of mice in which motor impairment was observed/number of mice tested (10 rpm); d PI = TD50/ED50; e 60 min. after drug administration;
NT – not tested.
a
c
Table 2. Effect of the test compounds on tactile allodynia and cold hyperalgesia in a mouse model of chemotherapy-induced
peripheral neuropathy caused by intraperitoneally administered OXA.
Compd
Dose
Test
Effect±SEM
[mg/kg]
For cold plate test: mean latency to nocifensive response [s]
For von Frey test: mean paw withdrawal threshold [g]
1st day (early phase of neuropathy)
7th day (late phase of neuropathy)
Before OXA –
healthy mice
Before compd
administration (3 h
after OXA)
1 h after compd
administration
Before compd
administration
1 h after compd
administration
Vehiclea
4b
-
Cold plate
Von Frey
Cold plate
Von Frey
Cold plate
Von Frey
Cold plate
Von Frey
Cold plate
Von Frey
Cold plate
Von Frey
57.64±1.267
3.36±0.23
59.76 ±0.16
3.144±0.169
58.48±0.856
3.209±0.156
22.333±6.748
2.222±0.086****
15.167±6.135
1.921±0.058
18.18±4.47
1.958±0.066
19.35±5.399
1.999±0.055
20.92±6.57
57.64±1.267
3.192±0.181
11.133±2.178
1.866±0.036
16.344±5.226
1.81±0.065
12.81±3.916
1.855±0.043
11.77±5.692
1.843±0.055
16.1±5.289
57.96±0.634
3.068±0.177
9.278±2.443
2.122±0.072*
8.833±3.473
2.027±0.054
10.65±3.169
1.947±0.029
7.09±1.525
1.958±0.047
8.86±2.05
15
18
18
15
18
50.433±2.888
2.867±0.019
56.822±1.69
2.795±0.05
46.95±3.672
2.714±0.058
56.73±2.604
2.985±0.079
54.25±2.472
2.714±0.081
25.189±7.641###
1.797±0.039####
16.222±7.176####
1.791±0.038####
22.82±5.766####
1.837±0.05####
34.27±7.511###
1.765±0.038####
20.45±6.407####
1.9±0.045####
5 b
6 b
7 b
13 b
2.09±0.036
1.822±0.042
1.907±0.027
a vehicle-treated mice did not receive OXA; b the test compounds were injected 3 h after OXA or 7 days after OXA. OXA – oxaliplatin; statistical analysis – two-
way ANOVA and post-hoc Tukey’s test; statistical significance compared values before OXA administration: ### p < 0.001, #### p < 0.0001; statistical
significance compared to values before compound administration: * p < 0.05, **** p < 0.0001.
3
4
5
6
7
9
12
13
-
In order to evaluate the mechanism of compounds action, the
most active compounds have been checked for affinity to 5-HT1A
receptors. The choice of this receptor based on results previously
obtained for reference compounds (Figure 2). The results are
presented in Table 3. Compounds 4, 6, 7, 9 and 13 showed binding
to the tested receptors. The Ki values ranged 1006 – 4769 nM.
1006.0 ± 113.0
-
3387.0
2370.0 ± 250.0
4796.0
-
1200.0 ± 118.0
1.0 ± 0.05
DPAT
- – no binding at 10-5 M
Table 3. Receptor studies performed for compounds 1, 3 – 7,
9, 12, 13.
Compd
1
5-HT1A Ki [nM]
-
10 anticonvulsant compounds were obtained in a group of 13
aminoalkanol derivatives. It seems that substitution of the phenyl

ring with three methyl groups (in positions 2,3,5 or 2,4,6) has a
positive effect on anticonvulsant activity. Of the 6 compounds of
this type obtained, only compound 8 was inactive (substituted at
positions 2,4,6, high mortality of experimental animals), while the
remaining five (4 – 7 and 13) had ED50 values below 20 mg/kg
b.w. Of the seven compounds disubstituted at positions 2,4 or 2,6,
five were found to be active, but they showed lower activity
compared to the trisubstituted derivatives (ED50 values in the range
of 27.43 – 52.49 mg/kg b.w.). Considering the neurotoxicity of the
compounds in this group, compound 4 (substituted in positions
2,3,5), was characterized by the highest protective index.
However, no relationship was observed between the phenyl ring
substitution pattern and protective index value, which indicates a
greater influence of other structural elements of the molecule.
aminobutan-1-ol (compound 11) moieties had different properties.
The R-enantiomer was only active at a high dose (100 mg/kg b.w.)
in which high neurotoxicity was also observed,11 whereas 11 had
moderate anticonvulsant activity and a relatively favorable PI
value of 2.33.
Receptor studies have shown that interaction with the 5-HT1A
receptor is not a must for anticonvulsant activity. However, its
involvement in modulation of the pharmacological profile cannot
be completely excluded. It is worth noting that the highest affinity
for the receptor was characterized by compound 4, which showed
multidirectional anticonvulsant and analgesic activity.
Considering also the results of receptor studies carried out for
reference compounds, among the potential molecular targets of
this group of compounds, sigma receptors and/or other elements
of the serotonergic system may be mentioned. At the same time,
based on the studies carried out, the joint or separate mechanism
of anticonvulsant and analgesic activity of compound 4 cannot be
clearly confirmed. Perhaps the mechanism of both activities
involves modulation of the activity of a number of therapeutic
targets, including 5-HT1A receptors. It is possible, however, that
anticonvulsant and analgesic activity are determined by separate
mechanisms.
Among the active compounds were 6 compounds possessing an
alkyl linker 2 – 4 carbon atoms long, and 3 compounds possessing
an ethoxyethyl linker. Comparing test results for compounds
differing only in linker structure (3 and 9, 4 and 13) as well as
experience in conducting research in similar groups of compounds
provide some valuable information on an influence of linker’s
structure on pharmacological profile. The use of an ethylene or
ethoxyethyl linker seems to be advantageous in terms of both
anticonvulsant activity and protective index. Despite the
significant difference in the length of these linkers, it cannot be
clearly determined which of them influences the pharmacological
profile favorably. This phenomenon can be explained by the
significant influence of other fragments of the molecule (e.g. the
amino component), by a different mechanism of anticonvulsant
activity of compounds with an ethylene or ethoxyethyl linker, but
also by a different binding mode to the molecular target or
conformation changes occurring within a molecule in the
molecular target environment.
Metabolism and safety characterization in vitro
Accurate characterization of the pharmacokinetic and toxicity
parameters of the active compounds at the early stages of new
drugs’ development reduces the time and cost of conducting
extensive research for less promising compounds. Therefore, fast
and low-budget screening methods, including in silico and in vitro
assays, are used in the drug design process.25
Metabolism studies
Metabolism of compounds 1, 4, 6, 7, 12 and 13 was studied in
vitro using murine microsomes. For all compounds, the main
pathway of metabolism was the hydroxylation reaction, which
resulted in the formation of monohydroxylated metabolites (m/z =
M+16), and in the case of compounds 7 and 13 – also
dihydroxylated (m/z = M+32). For compound 4, oxidation of the
previously formed hydroxylated derivative to the aldehyde group
(m/z = M+14) was observed. The detailed results are presented in
Supplementary material (Table S3).
The compounds obtained include molecules differing only in
the amine component, which makes it possible to analyze the
structure – activity relationship (SAR) considering this structural
fragment. For compounds 1 and 2, changing the position of the
piperidinol’s hydroxy group from position 3 to 4 causes a loss of
activity. Also, in the case of compounds 6, 7 and 8 (derivatives of
piperidinol – 6 or aminocyclohexanol – 7 and 8), the distance
between the oxygen and nitrogen atoms equal to 2 carbon atoms
seems favorable (active compounds 6 and 7). Among the
remaining cyclic aminoalkanol derivatives, anticonvulsant activity
was demonstrated by compounds 4, 13 (2-aminocyclohexan-1-ol
derivatives), 12 (4-aminocyclohexan-1-ol derivative) and 5
(piperidin-4-ol derivative), and among chain aminoalkanols,
compounds 3, 9 (R-(-)-2-aminopropan-1-ol derivatives) and 11 (S-
(+)-2-aminobutan-1-ol derivative). The distance between the
oxygen and nitrogen atoms within the aminoalkanol component
does not seem to determine the anticonvulsant activity of the
compound, as evidenced by the activity of compounds 5 and 12.
Nevertheless, the position of the hydroxy group undoubtedly
affects the pharmacological profile, including the ED50 value and
protective index. Moreover, it is worth to note that all compounds
possessing ethoxyethyl linker are also colamine derivatives,
however -NH-CH2-CH2-O- moiety is present within linker, not
only within aminoalkanol moiety.
To determine the structure of metabolites, LC/MS
fragmentation analysis was performed. Analysis of the results
confirmed the hydroxylation of methyl groups substituted to
aromatic ring, or – in the case of a larger number of metabolites
formed – also the aromatic ring itself (exemplum spectra available
in Supplementary material, Figure S2). A similar direction of
metabolic changes is observed for the previously described
The chirality of the used aminoalkanols allows consideration of
the role of stereoisomerism. Among the active compounds were
both R (compounds 3 and 9) and S (compound 11) isomers. It can
be concluded that the case of each of the tested chiral compounds
should be considered separately. For example, in the case of
compounds 9 and 10, which are derivatives of 2-aminopropan-1-
ol, the active enantiomer had the R configuration. Derivatives of
these compounds containing R-2-aminobutan-1-ol11 and S-2-

compounds having a methylphenoxy fragment, e.g. mexiletine26
and bupranolol.27 It was not possible to determine exactly where
the hydroxylation reaction takes place in the case of
monohydroxylated metabolites for compounds substituted with
more than one methyl group. Figure 3 shows the most probable
structures of the resulting metabolites as exemplified by
compound 4. Considering safety of the tested compounds after in
vivo administration, the aldehyde metabolite of compound 4
deserves attention. As reactive compounds, aldehydes can interact
with nucleic acids and proteins and as a consequence cause
mutagenicity or cytotoxicity. Among known antiepileptic drugs,
felbamate is metabolized to aldehyde (2-phenyl-prop-2-en-1-al).
This metabolite may be responsible for the hepatotoxicity of this
drug as well as the risk of aplastic anemia as a result of its use.28
considered as “neighbors” (which allows prediction reliability
assessment). The results of the analyzes carried out are presented
in Table 5.
Table 5. In silico prediction of selected types of toxicity
performed for compounds 1 – 13.
Type of
toxicity
Software
Osiris Property
Explorer
Lazar a
Mutagenicity
Compounds:
4, 5, 13 (high risk)
-
-
Carcinogenicit
y
Reproductive
toxicity
Compounds:
1 – 3, 9 – 11
|
b
-
a
for compounds 4, 8, 12 (carcinogenicity) and 7 (carcinogenicity and
Figure 3. Possible metabolites of compound 4.
mutagenicity) prediction in Lazar was not possible due to low amount of
“neighbor” compounds in the database; for all other compounds similarity
In addition to determining the direction of metabolic changes
and indicating the possible structures of the resulting metabolites,
in vitro studies using microsomes allow comparison of phase I
metabolic rate. Half-life (t1/2) was calculated for each compound,
as well as internal clearance (Clint), which parameter does not
depend on the concentration of microsomes and describes the
metabolic stability of the compounds (Table 4). According to the
classification used by CYPROTEX,29 compound 12 is
characterized by slow metabolism, while the remaining ones are
classified as metabolically unstable. Compounds 4 and 7, which
have the highest Clint values, are 2-aminocyclohexan-1-ol
derivatives. It can be assumed that this aminoalkanol moiety may
increase the metabolic lability of compounds, despite the fact that
metabolism occurs within the substituents of the aromatic system.
According to the classification published by B. Williamson et al.,30
all tested compounds are classified as moderately metabolically
stable.
b
threshold < 0.5 – prediction may be out of applicability domain; lack of
reproductive toxicity predictions in Lazar
The Osiris Property Explorer program identified 3 potentially
mutagenic compounds. The structural alert for this type of toxicity
was substitution at the aromatic ring with three methyl groups at
positions 2, 3 and 5. The Lazar program identified several
substances as potentially carcinogenic compounds. However, for
some compounds, prediction was not possible due to the
insufficient number of “neighbors” for which experimental data
are available. Prediction for other compounds may be affected by
an error due to the low threshold of similarity of “neighbors” to
the initial test compound. For the above-mentioned reasons, the
Lazar program does not seem to be a preferential tool for
predicting toxicity in the group of phenoxyalkyl- and
phenoxyethoxyethyl derivatives of aminoalkanols. Perhaps in the
future an update of the experimental database will increase the
reliability of the prediction.
Table 4. Metabolic stability of tested compounds – values of
half-life (t1/2) and internal clearance (Clint).
Mutagenicity tested in vitro
Compd
Protein concentration
t1/2
Clint
(mg/mL)
0.4
0.2
0.4
0.2
0.4
0.2
(min)
28.6
27.2
38.9
27.2
87.7
37.2
11.0
(µL/mg/min)
60.5
127.5
44.5
127.5
19.8
The Ames35 test was used to verify the potential mutagenic
activity of compounds in vitro and assess the effectiveness of
prediction of the Osiris Property Explorer program. The test uses
genetically modified Salmonella enterica sv. Typhimurium
strains, which are devoid of histidine synthesis and do not grow on
medium lacking this amino acid. In the presence of the mutagen, a
reverse mutation occurs that restores this ability and increased
growth of bacterial colonies compared to the control can be
observed. Depending on the strain used, the test can detect
compounds that cause different types of mutations, e.g. point
mutations (TA100 strain) or frameshift mutations (TA98 strain).35
The selection of compounds for mutagenicity testing was based on
the results of predictions using the Osiris Property Explorer
program. Compounds with high risk of mutagenicity (4, 5 and 13)
were tested, as well as compounds 1 and 12 in order to include
compounds with various cyclic aminoalkanol components. Each
compound was tested at five concentrations (10 – 750 or 10 – 1000
μg/plate). The concentration range depended on the solubility of
tested compounds. Concentrations above 750 μg/plate caused
precipitation on the dish, making the results difficult to read.
Therefore, at further stages of the study, the concentration of 750
μg/plate was the highest one used. All compounds were tested
using strains TA100 and TA98.
1
4
6
7
12
13
93.0
125.5
Imipramine31 0.5
In silico safety evaluation
All compounds obtained were evaluated in silico for potential
toxicity. Mutagenicity, carcinogenicity and reproductive toxicity
were included, due to their significance in the case of orally
administered drugs, as well as serious health consequences in case
of occurrence. Two alternative programs available for free – Osiris
Property Explorer32 and Lazar33 – were used. Programs differ in
the method of predicting toxic properties. Osiris Property Explorer
enables risk prediction (low, medium, high) of selected types of
toxicity based on the analysis of the chemical structure and
detection of fragments predisposing to particular type of toxicity.
The list of these groups bases on the analysis of the structures of
all compounds present in the RTECS database (Registry of Toxic
Effects of Chemical Substances),34 which are known to show the
particular type of toxicity (e.g. mutagenic or carcinogenic
compounds). The Lazar program enables prediction of selected
types of toxicity on the basis of experimental literature data
available for structurally similar compounds, the so-called
“neighbors”. The results of the prediction consist not only of the
potential toxicity alert, but also of the extensive characteristics of
the calculations performed, including the list of compounds
The results are presented in Table 6. Only compound 13 was
safe at all concentrations tested, despite being indicated by Osiris
Property Explorer. All other compounds were characterized by MI
(mutagenicity index) ≥ 2 at a minimum of one concentration for
one or both bacterial strains. A clearly marked relationship

between activity and concentration can, however, only be
observed for compound 5. For the remaining compounds, this
relationship is absent, mutagenic activity was only observed at the
highest concentration tested (compound 1), or MI decreases with
increasing concentration (compound 4). The latter case may be the
result of the compound’s toxicity at high concentrations, which is
manifested by the complete absence of bacterial growth at
concentrations of 500 μg/plate and higher (strain TA100). It
should also be noted that at higher concentrations, the results
obtained for some compounds were characterized by a high
standard deviation and the morphology of the bacteria changed (in
some cases making reading difficult – e.g. compounds 12, 13).
This indicates possible additional effects produced by the tested
compounds, in addition to the mutagenic effect.
Safety in relation to the gastrointestinal flora tested in vitro
For four active aminoalkanol derivatives (compounds 3, 5, 6
and 9), safety tests were performed in relation to the
gastrointestinal flora.
The preferred route of administration of anticonvulsants and
analgesics is oral, which is associated with the risk of dysbiosis if
they are active against the bacteria in the microbiome. Until now,
disorders of the natural gastrointestinal flora have been considered
primarily as an adverse effect of the use of antibiotics. Recent
reports, however, prove that dysbiosis can also be caused by other
groups of drugs, including centrally active ones (e.g. some
antidepressants).36,37 Dysbiotic conditions can cause many
complications, including inflammatory bowel disease, but also
inflammation of other organs and systems. Due to the weakening
of the blood-brain barrier, inflammation can even affect the central
nervous system. The relationship between the pathophysiology of
some central nervous system diseases and the state of the
microbiome has been documented, including Alzheimer's disease,
Parkinson's disease and depression.37,38
Table 6. Mutagenicity of compounds 1, 4, 5, 12, 13 (Ames
test).
Compd
Conc.
Mean number of revertants ± SD (MI)b
Strain
[μg/plate]a
TA100
TA98
1
DMSO
10
75
200
500
750
72.5±9.2
8.0±1.4
The study was carried out using a variety of bacterial strains
living on different sections of the digestive tract, including aerobic
(Enterococcus faecalis ATCC 29212, Escherichia coli ATCC
25922, Escherichia coli ATCC 35218, Staphylococcus aureus
ATCC 25923, Staphylococcus aureus – a clinical strain,
Salmonella enterica ATCC 51957) – by serial dilution method in
liquid medium, as well as anaerobic (Bacteroides fragilis ATCC
25285, Lactobacillus paracasei ATCC 25302, Prevotella
loescheii ATCC 15930, Bifidobacterium spp. – a clinical strain,
Clostridium difficile – a clinical strain) – by dilution method in
solid medium and by diffusion-disk method.
89.5±6.4 (1.2)
69.5±10.6 (1.0)
88.5±0.7 (1.2)
81.0±9.9 (1.1)
3869.5±1253.7
(53.4)
130.7±11.0
147.0±15.6 (1.1)
138.3±20.0 (1.1)
134.7±10.7 (1.0)
72.5±9.2
6.3±5.9 (0.4)
8.0±1.0 (0.8)
6.7±2.1 (1.2)
4.7±1.5 (1.0)
3.3±1.5 (1.0)
4
DMSO
0.1
1
|
|
|
|
5
DMSO
10
75
200
500
750
DMSO
10
8.0±1.4
185.5±57.3 (2.6)
105.0±58.1 (1.4)
49.0±13.9 (0.7)
-
7.3±3.1 (0.9)
6.3±2.1 (0.8)
6.7±3.2 (0.8)
8.7±2.5 (1.1)
9.3±2.3 (1.2)
8.3±2.1
12.0±3.5 (1.4)
15.3±5.5 (1.8)
11.0±3.6 (1.3)
6.7±5.8 (0.8)
364.3±84.6 (43.7)
|
All the strains listed showed resistance to the tested compounds
at the concentrations tested, i.e. from 0.195 μg/mL to 100 μg/mL
for aerobic strains and 50 μg/mL for anaerobic strains. Therefore,
the tested compounds can be considered safe in relation to the
gastrointestinal flora. The detailed results are presented in
Supplementary material (Tables S4-S11).
-
5
64.3±10.4
66.0±2.0 (1.0)
52.3±5.1 (0.8)
42.0±8.7 (0.7)
944.0±859.1 (14.7)
|
1190.0±1031.2
(18.5)
72.5±9.2
226.7±50.0 (3.1)
312.3±120.2 (4.3)
55.0±5.7 (0.8)
274.3±119.1 (3.8)
/
75
200
500
750
1000
Cell culture studies
Compound 4 was selected for comprehensive studies on the
effects on astroytes, including cytotoxicity as well as impact on
proliferation and cell cycle. Such studies are an important element
of safety evaluation panel for compounds targeting central nervous
system. However, glial cells are also interesting in the context of
epilepsy and seizures’ inhibition.
12
13
DMSO
10
75
200
500
750
DMSO
10
75
200
500
DMSO
750
DMSO
5
DMSO
2.5
8.0±1.4
8.0±2.0 (1.0)
8.3±3.5 (1.0)
9.3±2.5 (1.2)
6.7±2.1 (0.8)
3.0±2.6 (0.4)
8.0±1.4
7.0±2.6 (0.9)
8.0±4.6 (1.0)
11.0±1.4 (1.4)
9.0±2.8 (1.1)
|
72.5±9.2
86.0±5.0 (1.2)
80.7±27.1 (1.1)
70.5±7.8 (1.0)
/
130.7±11.0
56.0±14.1 (0.4)
64.3±10.4
/
|
Sodium
azide
4-nitro-o-
phenylene-
-diamine
533.7±9.9 (8.3)
|
8.3±2.1
519.0±10.6 (62.3)
a all tests performed in triplicate; b SD – standard deviation; MI – mutagenicity
index – the quotient of the number of revertants present on the plate with the
test compound at a given concentration and the number of revertants present
on the negative control; dose-dependent increase (MI ≥ 2) in the number of
revertants for a minimum of one strain – compound considered mutagenic; - –
no growth; / – equivocal result (including e.g. significant change in bacterial
morphology); | – not tested

It has been proved that many known anticonvulsants show
cytotoxicity in vitro. These are especially carbamazepine,
topiramate and oxcarbazepine, but in concentration 50 μg/mL or
higher also gabapentin, lamotrigine, tiagabine and levetiracetam.39
Moreover, valproate proved to exhibit antiproliferative effect in
C6 glioma model by restriction of the glial cell cycle at a defined
point in the mid-G1 phase, which fact may explain its
teratogenicity and specificity in inducing neural tube defects.40
The above data could be taken into account in the therapeutic use
of particular drugs as well as in design of new compounds
characterized by high tolerability. On the other hand, astrogliosis
membrane disruption (LDH test – Figure 4C, trypan blue test after
24 and 48 h incubation – Figure 4D). For all tests murine astrocytes
were used in order to make results relevant to previously observed
anticonvulsant activity (mice, i.p.). However, the MTT test was
also performed comparatively on human astrocytes obtained by
means of iPSC cells induction. All assays performed on murine
cells showed cytotoxicity of compound 4 in the highest tested
concentration equal to 100 μM (in trypan blue test the effect was
observed only after 48 h incubation). However, in the case of the
MTT test performed with a use of human astrocytes, cytotoxicity
has already been observed from a concentration of 50 μM.
is considered
a
part of pathophysiological processes
Additionally, influence of compound 4 on murine astrocytes’
proliferation was evaluated in crystal violet assay (Figure 4E). The
dye is accumulated in a cell nucleus. After extraction, the number
of cells can be detected calorimetrically. The assay showed
antiproliferative activity of compound in concentration 100 μM
after 24 h incubation and 25 μM and higher after 48 h incubation.
According to results obtained using crystal violet assay we decided
to check if compound 4 have influence on cell cycle distribution
of astrocytes using flow cytometry analysis (propidium iodide
accompanying epileptogenesis, which was observed e.g. in animal
models of temporal lobe epilepsy.41 Astrocytes became a potential
therapeutic target for anticonvulsants, although their role in
epilepsy has not yet been fully understood. Among the most
frequently mentioned potential astrocytic targets for
anticonvulsants are modulation of glutamate uptake, release of
neurotransmitters (including glutamate, GABA or ATP) and
suppressing Ca2+ signalling.42,43 Noteworthy, pharmacological
activity of some known anticonvulsants (e.g. valproate,
gabapentin, phenytoin) may be partially mediated by direct
depression of astrocytic activity.43 The above facts are a premise
for screening new anticonvulsant compounds for their influence
on astrocytes.
staining). This approach allowed determination of the total amount
of DNA per cell and stoichiometrically determined to obtain cell
cycle distributions. Flow cytometry analysis do not show any
statistically significant differences between control cells (non-
treated) and cells incubated in the presence of compound 4 (25µM)
for 24 hours. The data are presented in Supplementary material
(Figure S3).
As part of the current work a panel of cytotoxicity tests was
conducted, including tests based on both metabolic impairment
evaluation (MTT test, Figure 4A and 4B) as well as detection of
Figure 4. Cell culture studies performed for compound 4: cytotoxicity (A – MTT assay, murine astrocytes; B – MTT assay, human astrocytes derived from iPSC;
C – LDH assay, murine astrocytes; D – trypan blue assay, murine astrocytes) and antiproliferative effect (E – crystal violet assay, murine astrocytes); DOX –
doxorubicin.
In summary, 13 new aminoalkanols were designed and
synthesized, including 9 compounds possessing anticonvulsant
activity and 1 compound (4) exhibiting both anticonvulsant and
analgesic in neuropathic pain activities. The introduction of an
ethylene or ethoxyethyl linker seems to be advantageous in terms
of both anticonvulsant activity and protective index. The distance
between the oxygen and nitrogen atoms within the aminoalkanol
component does not seem to determine the anticonvulsant activity
of the compound, however the position of the hydroxy group
undoubtedly affects the pharmacological profile. Major
differences were observed within enantiomers’ pharmacological
activity, however among the active compounds were both R and S
isomers. Receptor studies have shown that interaction with the 5-
HT1A receptor is not a must for anticonvulsant activity, however

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its involvement cannot be completely excluded. The common or
distinct mechanism of anticonvulsant and analgesic activity
remains unclear. A wide range of metabolism and safety studies
performed within the presented study revealed that presented
group of compounds is safe with relation to gastrointestinal flora.
Aspects that are worth paying attention to when designing new
active compounds are potential mutagenicity, cytotoxicity, as well
as metabolism to aldehydes.
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Acknowledgments
We would like to thank Damian Ryszawy PhD (Department of
Cell Biology, Faculty of Biochemistry, Biophysics and
Biotechnology, Jagiellonian University, Kraków, Poland) for
providing induced iPSC cells.
The research has been financially supported by National
Science Centre (grant no. 2015/17/N/NZ7/00966) and Jagiellonian
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