Preparation, in vitro screening and molecular modelling of symmetrical 4-tert-butylpyridinium cholinesterase inhibitors - Analogues of SAD-128

Article abstract of DOI:10.1016/j.bmcl.2010.11.051

Carbamate inhibitors (e.g., pyridostimine bromide) are used as a pre-exposure treatment for the prevention of organophosphorus poisoning. They work by blocking acetylcholinesterase's (AChE) native function and thus protect AChE against irreversible inhibi

Full text of DOI:10.1016/j.bmcl.2010.11.051

Bioorganic & Medicinal Chemistry Letters 21 (2011) 150–154
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Preparation, in vitro screening and molecular modelling of symmetrical
4-tert-butylpyridinium cholinesterase inhibitors—Analogues of SAD-128
Kamil Musilek a,d, Jan Roder b, Marketa Komloova b, Ondrej Holas b, Martina Hrabinova c, Miroslav Pohanka c,
Vlastimil Dohnal d, Veronika Opletalova b, Kamil Kuca c,d, Young-Sik Jung e,
⇑
a University of Defence, Faculty of Military Health Sciences, Department of Toxicology, Trebesska 1575, 500 01 Hradec Kralove, Czech Republic
b Charles University, Faculty of Pharmacy, Department of Pharmaceutical Chemistry and Drug Control, Heyrovskeho 1203, 500 05 Hradec Kralove, Czech Republic
c University of Defence, Faculty of Military Health Sciences, Centre of Advanced Studies, Trebesska 1575, 500 01 Hradec Kralove, Czech Republic
d University of Jan Evangelista Purkynje, Faculty of Science, Department of Chemistry, Ceske mladeze 8, 400 96 Usti nad Labem, Czech Republic
e Bioorganic Science Division, Korea Research Institute of Chemical Technology, PO Box 107, Yuseong, Daejeon 305-606, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Carbamate inhibitors (e.g., pyridostimine bromide) are used as a pre-exposure treatment for the preven-
tion of organophosphorus poisoning. They work by blocking acetylcholinesterase’s (AChE) native function
and thus protect AChE against irreversible inhibition by organophosphorus compounds. However, carba-
mate inhibitors are known for many undesirable side-effects related to the carbamylation of AChE. In this
Letter, 19 analogues of SAD-128 were prepared and evaluated as cholinesterase inhibitors. The screening
results showed promising inhibitory ability of four compounds better to used standards (pralidoxime,
obidoxime, BW284c51, ethopropazine, SAD-128). Four most promising compounds were selected for fur-
ther molecular docking studies. The SAR was stated from obtained data. The former receptor studies were
reported and discussed. The further in vivo studies were recommended in the view of OP pre-exposure
treatment.
Received 7 October 2010
Revised 6 November 2010
Accepted 9 November 2010
Available online 16 November 2010
Keywords:
Acetylcholinesterase
Organophosphate
Pretreatment
Reversible inhibitor
In vitro
Ó 2010 Elsevier Ltd. All rights reserved.
Molecular modelling
Acetylcholinesterase (AChE; EC 3.1.1.7) and butyrylcholinester-
ase (BChE; EC 3.1.1.8) are essential enzymes in human body
splitting various esters.1,2 Whereas AChE is crucial enzyme for
termination of neurotransmission via splitting neuromediator
acetylcholine, BChE was called non-specific esterase for its broad
esterase activity (e.g., butyrylcholine, cocaine, anaesthetic esters).3
Both AChE and BChE have many natural and artificial inhibitors
blocking their native function. Among them, the organophosphorus
compounds (OPs) belong to the most dangerous compounds ever
developed by man.4 They covalently bind to esterase active serine
(Ser203 for human AChE and Ser198 for human BChE) and fully
block their activity. They cause the permanent block of peripheral
and central nervous system with cholinergic crisis development
and obstruction of breathing leading to death by suffocation.5
Generally, the pre-exposure and post-exposure strategies were
developed to counteract the effects of OPs on human.6 The pre-
exposure strategy consists in administration of reversible AChE
inhibitor, stoichiometric or catalytic scavengers.7 The post-
exposure strategy is based on combination of anticholinergic drug,
AChE reactivator and anticonvulsant.8
inhibitors (e.g., pyridostigmine chloride; Fig. 1) usually in combina-
tion with centrally active anticholinergic drugs (e.g., benactyzine,
trihexyphenidyl; Fig. 2).9 The carbamate drug (pyridostigmine) is
used to reversibly block AChE active site serine. Thus, it protects
AChE from OPs binding and it is spontaneously decarbamylated
with normal restoration of AChE activity. Additionally, it enhances
the antidotal efficiency in post-exposure treatment, especially in
case of nerve agent poisoning. However, the mostly used pyrido-
stigmine bromide has charged molecule and consequently it is
poorly penetrating the blood–brain barrier (BBB).10 For this reason,
the symptomatic anticholinergic drugs (Fig. 1) are used to diminish
the OP effects to brain AChE.
Though the carbamate AChE inhibitors are widely used, they are
well known for their increased and serious side effect.11 These
side-effects consist in carbamylation of AChE active site by higher
O
O
O
O
N
O
O
O
O
O
F
P
P
P
NO2
Concerning the pre-exposure strategies, the only currently ap-
proved method for human use is administration of reversible AChE
N
tabun (GA)
Figure 1. Organophosphorus compounds.
paraoxon (POX)
DFP
⇑
Corresponding author. Tel.: +82 42 860 7135; fax: +82 42 860 7348.
E-mail address: ysjung@krict.re.kr (Y.-S. Jung).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.11.051

K. Musilek et al. / Bioorg. Med. Chem. Lett. 21 (2011) 150–154
151
O
N
N
O
N
N
NOH
N
Cl
N
O
HON
NOH
HO
Cl
O
HO
2 Cl
O
N
pralidoxime (1)
obidoxime (2)
H
N
pyridostigmine
benactyzine
trihexyphenidyl
O
Figure 2. Drugs used to protect human against OP poisoning.
N
S
Cl
2 Br
BW284c51 (3)
Figure 3. Cholinesterase reactivators and inhibitors used as standards.
N
N
dosage of carbamate and are manifested as increased salivation,
gastrointestinal motility, bronchial secretion, cardiac arrhythmia
and development of cholinergic crisis. On account of these serious
side-effects, other reversible AChE inhibitors might be used for
combined pre-exposure strategy.12 The main advantage of such
reversible inhibitor should be the selectivity between AChE and
BChE, where BChE can also act as physiological scavenger of OPs
in human body. The inhibitor’s reversible binding aside the active
site serine (Ser203) and consequently decreased side-effects
should be also requested. The inhibitor’s BBB penetration remains
the questionable issue. While the penetrating inhibitor may also
protect brain AChE against OP attack, it may also increase the over-
all toxicity.13 On the other hand, the problem with inhibitor not pe-
netrating the BBB can be bridged by standard anticholinergics
proved by clinical practise.9
A few decades ago, AChE inhibitor called SAD-128 (17; Table 1)
was prepared during the development of AChE reactivators (e.g.,
pralidoxime, obidoxime; 1 and 2; Fig. 3).14 In contrast with AChE
reactivators, this interesting molecule was lacking the oxime moi-
ety and have symmetrical molecule with 4-tert-butylpyridinium
rings. SAD-128 (17) as obidoxime analogue was found to be revers-
ible AChE inhibitor able to protect against several LD50 of various
OPs.15 Consequently, it was found also inhibitor of muscarinic
and nicotinic receptors that explained the protection of experi-
mental animals by SAD-128 against OPs.16–18
Concerning the promising data on SAD-128, its 19 analogues
(5–24; Table 1) were prepared. Some of them were formerly re-
ported (6–14), but they were not evaluated as cholinesterase
inhibitors.19 The design of novel symmetrical compounds origi-
nated from parent molecule with two 4-tert-butylpyridinium rings
and variable linkage. The connecting linkers were selected from
formerly published data.20–22 Namely, the bis-pyridinium or bis-
isoquinolinium compounds bearing alkylenyl and naphtylenyl
linkers (5–16) showed very promising inhibitory results towards
hAChE.20,21 Differently, the linkers with heteroatom (17 and 18),
double bonds (19 and 20) or xylenyl moieties (21–23) were found
promising during preparation of AChE reactivators.23–26 For these
reasons, presented linkers were used in effort to exceed the prop-
erties of parent molecule (SAD-128).
ethopropazine (4)
was stirred at 70 °C for 14–98 h (Scheme 1). Subsequently, the
reaction mixture was cooled to the room temperature, portioned
with acetone (80 ml) and cooled in refrigerator overnight (5 °C).
The crystalline crude product was collected by filtration. The amor-
phous crude product was decanted and residual solvent was
poured off. The crude products were purified by boiling in ethyl
acetate (50 ml). NMR, ESI-MS and elemental analysis were used
to determine the entity and purity of all prepared compounds.
The former and the novel compounds were assayed for their
inhibitory ability in vitro using a standard inhibition test utilising
human recombinant AChE and human plasmatic BChE.27 The
hAChE was chosen as the main target for the OP pre-treatment
strategy.9 Additionally, hBChE was chosen as a member of the cho-
linesterase family, which is usually affected by all the compounds
that are interacting with hAChE.3 The commercial oximes (1 and 2)
were selected as standards among AChE reactivators that may be
used not only for post-exposure treatment, but also for pre-expo-
sure treatment (e.g., prophylaxis).9 The BW284c51 (3; Fig. 3) and
ethopropazine hydrochloride (4; Fig. 3) were chosen as selective
AChE or BChE inhibitors to cover the selectivity issues.28,29 The
IC50 values of all compounds are listed in Table 2.
The commercial oximes (1 and 2) showed weak inhibition of
hAChE on mM scale and did not seem to be relevant compounds
for OP pre-exposure treatment.22 The selective standards (3 and
4) presented expected results for hAChE. Whereas compound 3
was promising hAChE inhibitor on nM scale, compound 4 resulted
as its weak inhibitor.28 The newly prepared compounds presented
mixed results. Compounds 5–9 and 17–23 resulted as AChE inhib-
itors on
lM scale. More interestingly, compounds 10–16 and 24
were found to be nM AChE inhibitors. Some of these compounds
(11 and 12, 14, 15 and 16, 24) exceeded the commercial standard
3 (30 nM) in hAChE inhibition. Among them, compounds 12
(5 nM), 14 (12 nM) and 16 (7 nM) were the most promising AChE
inhibitors bearing aliphatic linkage and compound 24 (24 nM)
was the most promising inhibitor with different linkage type. For
hBChE, the commercial oximes again resulted as poor inhibitors,
where pralidoxime (1) did not showed inhibition of hBChE at all.
For chosen selective compounds (3 and 4), compound 4 showed
The novel compounds were prepared via standard synthetic
strategy.20 The solution of 4-tert-butylpyridine (6.8 mmol) and cor-
responding dihalogenated compound (3.1 mmol) in DMF (10 ml)
Table 1
Newly prepared analogues of SAD-128.
expected increased inhibition on
lM scale, while compound 3
was found to be poor inhibitor of hBChE.29 Similarly to hAChE re-
N
(A) N
sults, some newly prepared compounds (5–9, 17–23) showed BChE
2 X
inhibition on
lM scale. Furthermore, compounds 10–11, 13, 15
and 24 resulted as sub-
l
M hBChE inhibitors. Most interestingly,
5–16 (Br)
17 (Cl)
18 (Br)
19 (Br)
20 (Cl)
21 (Br)
22 (Br)
23 (Br)
24 (Br)
(CH2)1-12
CH2OCH2
(CH2)2O(CH2)2
(E)-CH2CH@CHCH2
(Z)-CH2CH@CHCH2
1,2-Phenylenyl
1,3-Phenylenyl
1,4-Phenylenyl
3,6-Naphtylenyl
X-(A)-X
N
N
(A) N
DMF; 70°C
2 X
Scheme 1. Preparation of SAD-128 analogues.

152
K. Musilek et al. / Bioorg. Med. Chem. Lett. 21 (2011) 150–154
Table 2
Inhibitory potency of tested compounds towards cholinesterases and calculated lipophilicity
Compound
AChE IC50 SDa
(lM)
BChE IC50 SDa(
lM)
SIc BChE/AChE
Ki1/Ki2
(lM)
log P
b
Pralidoxime (1)
Obidoxime (2)
BW284c51 (3)
Ethopropazine (4)
5
6
7
8
9
10
11
12
13
14
15
16
878 171
577 113
0.030 0.006
1020 199
—
—
—
—
ꢀ2.46
ꢀ2.42
ꢀ6.19
5.11
2.29
1.35
0.95
0.68
0.56
0.67
0.87
1.08
1.37
1.66
2.01
2.33
1.73
0.11
0.92
0.92
1.42
1.42
1.42
2.54
1910 311
354 58
1.6 0.3
34
20
53
5.5 0.9
5.1 0.8
0.6 0.1
3.3
11,800
0.002
1.7
1.2
3.8
1.5
0.2
19
0.01/0.05
24.7/12100
—
—
—
—
—
—
20
17
14
4
3
3
6
3
9
3.7 0.7
24
5
0.032 0.006
0.016 0.003
0.005 0.0009
0.037 0.007
0.012 0.002
0.026 0.005
0.007 0.001
0.7 0.1
44
—
0.016 0.003
0.15 0.02
0.06 0.01
0.11 0.02
0.019 0.003
75 12
3.2
4.1
5.0
4.2
2.7
6.2
13
5.6
2.2
0.2
2.7
4.3
5
0.09/0.20
—
—
—
—
—
—
—
—
—
—
—
SAD-128 (17)
12
19
2
4
18
19
20
21
22
23
24
241 40
19
3.4 0.7
2.0 0.4
3
4.4 0.7
2.4 0.5
1.9 0.3
1.3 0.2
0.12 0.02
10
2
0.7 0.1
0.30 0.06
0.024 0.005
0.10/0.10
a
b
c
Mean value of three independent determinations.
No inhibition in selected concentration scale.
Selectivity index.
compounds 12, 14 and 16 were found to be nM hBChE inhibitors.
Among sub- M and nM hBChE inhibitors, compounds 10–16 and
l
24 exceeded the commercial standard 4. The selectivity index
(SI) was calculated to reveal the selectivity for one of cholinester-
ases. The oxime reactivator 1 displayed some selectivity towards
hAChE, while it was not inhibiting hBChE. Obidoxime (2) didnot
present higher selectivity for one cholinesterase. The standard cho-
linesterase inhibitors (3 and 4) showed anticipated results, where
compound 3 was highly selective for hAChE and compound 4 for
hBChE.29 The newly prepared compounds didnot show higher
selectivity with best result of 44-fold better inhibition of hAChE
by compound 11.
The novel molecules were designed as reversible AChE inhibi-
tors. Thus, the kinetic experiments with recombinant hAChE were
developed to confirm this hypothesis.30 The AChE standard 3 was
found to be its non-competitive inhibitor, whereas the BChE stan-
dard 4 showed strong competition with the substrate during enzy-
matic reaction and it was indicated as competitive AChE inhibitor.
Two novel compounds (12 and 24) were highlighted for kinetic
experiments. Compound 12 was chosen among the potent AChE
inhibitors bearing aliphatic linkage and compound 24 as the best
AChE inhibitor with different type of the linker. Both selected com-
pounds 12 and 24 resulted as non-competitive hAChE inhibitors
with almost no influence to substrate (acetylthiocholine) hydroly-
sis. This finding confirmed their binding aside the AChE active site
serine (Ser203).
Figure 4. Molecular docking results for mAChE with compound 12 (blue), 14
(magenta), 16 (yellow) and 24 (orange).
CH–
p interactions with Trp86 (3.4 Å), Tyr337 (3.6 Å) and His447
(3.3 Å).35 The second pyridinium ring was attached by cation–
p
interaction to Tyr286 (3.3 Å) and its 4-tert-butyl moiety was stabi-
lized by CH–CH interactions with Leu289 (3.2 Å) and Glu292
(3.7 Å). The whole molecule of compound 12 (as well as molecules
14 and 16) was penetrating inside the active site gorge closely to
Ser203. Differently, the top-scored docking pose of compound 24
(ꢀ8.25 kcal/mol) displayed binding on the AChE surface and was
not going inside the active site gorge (Fig. 4). First of its pyridinium
The docking studies were performed on four promising com-
pounds after the in vitro screening (12, 14, 16, 24) in order to ratio-
nalise their possible interactions within AChE and BChE active site.
Three crystal structures were used for the docking calculations
(hAChE—1b41, mAChE—2jez, 2jf0; hBChE—1p0i) and the best re-
sults were obtained for the mAChE model (2jez).31–34 For mAChE,
compounds 12, 14 and 16 presented very similar type of interac-
tion (Fig. 4). Thus, only the most promising compound 12 is dis-
cussed. Its top-scored docking pose (ꢀ8.58 kcal/mol) showed
rings was attached by cation–p interaction to His287 (3.9 Å) and
its 4-tert-butyl moiety presented CH–CH interactions with
Leu289 (3.0 Å), Pro290 (3.8 Å) and Gln291 (4.0 Å). The second
pyridinium moiety was not stabilized by aromatic interaction,
but by CH–CH interactions of its 4-tert-butyl moiety with Val340
(3.7 Å), Val343 (3.5 Å) and Pro344 (3.8 Å).35 Importantly, the
important cation–p interactions. First pyridinium ring was at-
tached to Tyr124 (3.3 Å), Phe297 (3.9 Å), Tyr337 (3.4 Å) and
Phe338 (3.2 Å), whereas its 4-tert-butyl moiety was stabilized by
naphtylene moiety of the connecting linker displayed strong
interaction with Tyr286 (3.2 Å).
p–p

K. Musilek et al. / Bioorg. Med. Chem. Lett. 21 (2011) 150–154
153
For hBChE, compounds 12, 14 and 16 again presented very sim-
(21–23) were found to be effective hAChE inhibitors only on lM
ilar type of interaction (Fig. 5). Concerning the most promising
compound against hBChE 12, its top-scored docking pose
scale. In contrast, compounds bearing longer aliphatic C6–C12
(10–16) and naphtylene linkers (24) resulted as very potent AChE
inhibitors on nM scale. The assumed explanation of the increased
inhibitory ability by compounds 10–16 and 24 consists in the
accommodation of such molecules in the AChE or BChE active site
gorge, where they were able to bind the essential aromatic resi-
(ꢀ8.14 kcal/mol) showed cation–
moiety with Tyr332 (3.1 Å) and Phe329 (3.3 Å), while its 4-tert-bu-
tyl moiety was attached via CH– interactions to Trp82 (3.5 Å) and
p interactions of one pyridinium
p
His438 (3.6 Å). The second pyridinium moiety displayed cation–
interaction with Phe329 (3.2 Å) and its 4-tert-butyl moiety CH–
p
p
dues (Trp, Tyr, Phe, His). Interestingly, the even linkers C8–C10–
interactions with Trp231 (3.5 Å) and Phe398 (3.5 Å). The com-
pounds 12, 14 and 16 were all twisted in the BChE active site. In
contrast, top-scored docking pose of compound 24 (ꢀ9.23 kcal/
mol) displayed flat binding of its rigid molecule. First of its pyrid-
C12 (12, 14, 16) were found to be more potent inhibitors to odd
linkers (11, 13, 15). Thus, the even linkers (12, 14, 16) were sup-
posed to better accommodate in the hAChE active site gorge. The
naphtylenyl linked compound (24) was found slightly worse inhib-
itor to aliphatic linked compounds (11 and 12, 14, 16). This finding
is plausibly related to its spatially rigid structure, where only lim-
ited free rotation in its molecule is available. For this reason, it
showed different top-scored docking pose compared to aliphatical-
ly linked compounds. Concerning lipophilicity, the calculated log P
values of the newly prepared compounds showed interesting re-
sults. For compounds 5–16, the shortest C1 linkage (5; log P 2.29)
was calculated to have log P value close to C12 (16; log P 2.33) or
naphtylenyl (24; log P 2.54) bridge, although it was supposed to
have the lowest log P value among all aliphatic linkers. Similarly,
C2 linker (6; log P 1.35) resulted close to C9 linkage (13; log P
1.37). This phenomenon may be explained by the spatial shield-
ing of quaternary nitrogen by the pyridinium rings that are very
close together for the shorter aliphatic linkers (5–8). For longer
aliphatic linkers (9–16), the log P values were found consistently
increasing with the length of the linker as it was expected. Addi-
tionally, SAD-128 (17; 1.73) log P value was found superior to its
aliphatic analogue (7; log P 0.95) and highly superior to its
slightly longer analogue (18; log P 0.11). In this case, the same
phenomenon as it was described for aliphatic linkage was sup-
posed. And, additionally, the other participating factor should
be the influence of free oxygen electron pairs that may be con-
jugated with aromatic rings and thus increased the lipophilicity
of compound 17. Concerning the in vitro, molecular docking
and lipophilicity data, the longer even aliphatic linkers (12, 14,
16) and the naphtylenyl linkage (24) were highlighted for fur-
ther investigation.
inium rings was sandwiched by cation–
Trp82 (3.6 Å) and His438 (4.0 Å), whereas the corresponding 4-
tert-butyl moiety was attached via CH– interactions with the
p interactions between
p
same residues (both 3.4 Å). The second pyridinium moiety was
not attached to aromatic residues and its 4-tert-butyl moiety pre-
sented CH–CH interactions with Pro285 (3.4 Å) and Gly283 (3.5 Å).
Importantly, the naphtylene linker was sandwiched by
actions between Tyr332 (3.4 Å) and Phe329 (3.1 Å).
p–p inter-
The structure–activity relationship (SAR) of the novel com-
pounds generated from in vitro and docking data can be demon-
strated.36 Firstly, the pyridinium moiety was found very effective
in forming strong
p–cationic interaction instead of weaker p–p
interactions of, for example, phenyl ring.37 Additionally, bis-pyrid-
inium or bis-isoquinolinium compounds were formerly found to be
effective AChE or BChE inhibitors.20,21 Their further substitution by
various functional groups may be the key factor to influence the
physical–chemical properties (e.g., lipophilicity). The tert-butyl
moiety was supposed to increase the lipophilicity of the prepared
molecules. From this point of view, structurally similar compounds
3 (log P ꢀ6.19), 18 (log P 0.11) and its published bispyridinium
analogue without tert-butyl moieties (1,10-pentane-1,5-bispyridi-
nium dibromide; log P ꢀ3.52) may confirm this hypothesis.20,38
Furthermore, additional interactions between cholinesterase and
4-tert-butyl moiety were expected. Subsequently, the CH-
p or
CH–CH interactions were found during the molecular docking of
compounds 12, 14, 16 and 24 with AChE or BChE.35 Thus, tert-buty-
lpyridinium moiety seemed to be promising molecular tool for
increasing lipophilicity and weak molecular interactions with
cholinesterases.
Whereas the tert-butylpyridinium moiety was uniform for all
novel compounds, the connecting linkage was different. The link-
age was apparently influencing both cholinesterase interactions
and lipophilicity. Based on in vitro data, compounds with shorter
aliphatic C1–C5 (5–9), aliphatic with heteroatom (17 and 18), dou-
ble bonded linkers (19 and 20) or linkers bearing xylene moiety
Compound SAD-128 (17) was formerly found to be receptor act-
ing inhibitor. Namely, it was determined to be reversible inhibitor
of the muscarinic (mAChR) and nicotinic (nAChR) acetylcholine
receptors together with its analogues (6–14).19 Unfortunately,
the receptor’s data were not connected to the cholinesterase inhi-
bition. Currently, these data may be supplemented and summa-
rised. The best inhibitor of hAChE from tested series (12) was
reported as potent inhibitor of mAChR (IC50 37
(IC50 30
lM) and nAChR
l
M).19 Similar data are available for compounds 10–14
that also resulted as potent receptor’s antagonists.19 For these rea-
sons, presented multiple acting compounds (e.g., 12 and 24) should
be evaluated in vivo to confirm or disprove their in vitro effective-
ness for OP pre-exposure treatment.
In summary, 19 analogues of SAD-128 were prepared and eval-
uated as cholinesterase inhibitors. The screening results showed
promising inhibitory ability of four compounds better to used stan-
dards (pralidoxime, obidoxime, BW284c51, ethopropazine, SAD-
128). Four most promising compounds were selected for further
molecular docking studies. The SAR was stated from obtained data.
The former receptor studies were reported and discussed. The fur-
ther in vivo studies were recommended in the view of OP pre-
exposure treatment.
Acknowledgements
This work was supported by the Ministry of Defence of the
Czech Republic (no. OVUOFVZ200805), the Grant Agency of the
Figure 5. Molecular docking results for hBChE with compound 12 (blue), 14
(magenta), 16 (yellow) and 24 (orange).

154
K. Musilek et al. / Bioorg. Med. Chem. Lett. 21 (2011) 150–154
16. Lundy, P. M.; Tremblay, K. P. Eur. J. Pharmacol. 1979, 60, 47.
17. Kloog, Y.; Sokolovsky, M. Mol. Pharmacol. 1985, 27, 418.
18. Alkondon, M.; Albuquerque, E. X. J. Pharmacol. Exp. Ther. 1989, 250, 842.
19. Broomfield, C.; Dembure, I. J.; Cuculis, J. Biochem. Pharmacol. 1987, 36, 1017.
20. Musilek, K.; Komloova, M.; Zavadova, V.; Holas, O.; Hrabinova, M.; Pohanka,
M.; Dohnal, V.; Nachon, F.; Dolezal, M.; Kuca, K.; Jung, Y. S. Bioorg. Med. Chem.
Lett. 2010, 20, 1763.
21. Musilek, K.; Komloova, M.; Holas, O.; Hrabinova, M.; Pohanka, M.; Dohnal, V.;
Nachon, F.; Dolezal, M.; Kuca, K. Eur. J. Med. Chem., submitted for publication.
22. Musilek, K.; Pavlikova, R.; Komloova, M.; Holas, O.; Hrabinova, M.; Pohanka,
M.; Dohnal, V.; Dolezal, M.; Kuca, K. J. Enzym. Inhib. Med. Chem., doi:10.3109/
14756366.2010.496362.
Charles University (no. 117909/2009/B-CH/FaF) and Ministry of
Education, Youth and Sport (no. SVV-2010-261-001).
Supplementary data
All experimental details are listed in the Supporting informa-
tion. Supplementary data associated with this article can be found,
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