Design, synthesis, and AChE inhibitory activity of new benzothiazole–piperazines

Article abstract of DOI:10.1016/j.bmcl.2016.10.041

In the current study, 14 new benzothiazole–piperazine compounds were designed to meet the structural requirements of acetylcholine esterase (AChE) inhibitors. The target compounds were synthesised in three steps. Structures of the newly synthesised compounds (7–20) were confirmed using IR,¹H NMR,¹³C NMR, and HRMS methods. The inhibitory potential of the compounds on AChE (E.C.3.1.1.7, from electric eel) was then investigated. Among the compounds, 19 and 20 showed very good activity on AChE enzyme. Kinetics studies were performed to observe the effects of the most active compounds on the substrate–enzyme relationship. Cytotoxicity studies, genotoxicity studies, and theoretical calculation of pharmacokinetics properties were also carried out. The compounds 19 and 20 were found to be nontoxic in both of the toxicity assays. A good pharmacokinetics profile was predicted for the synthesised compounds. Molecular docking studies were performed for the most active compounds, 19 and 20, and interaction modes with enzyme active sites were determined. Docking studies indicated a strong interaction between the active sites of AChE enzyme and the analysed compounds.

Full text of DOI:10.1016/j.bmcl.2016.10.041

Accepted Manuscript
Design, Synthesis, and AChE Inhibitory Activity of New Benzothiazole-Piper-
azines
Ümide Demir Özkay, Özgür Devrim Can, Begüm Nurpelin Sağlık, Ulviye Acar
Çevik, Serkan Levent, Yusuf Özkay, Sinem Ilgın, Özlem Atlı
PII:
S0960-894X(16)31069-1
DOI:
http://dx.doi.org/10.1016/j.bmcl.2016.10.041
BMCL 24344
Reference:
To appear in:
Bioorganic & Medicinal Chemistry Letters
Received Date:
Revised Date:
Accepted Date:
14 September 2016
12 October 2016
13 October 2016
Please cite this article as: Özkay, U.D., Can, O.D., Sağlık, B.N., Çevik, U.A., Levent, S., Özkay, Y., Ilgın, S.,
Atlı, O., Design, Synthesis, and AChE Inhibitory Activity of New Benzothiazole-Piperazines, Bioorganic &
Medicinal Chemistry Letters (2016), doi: http://dx.doi.org/10.1016/j.bmcl.2016.10.041
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Design, Synthesis, and AChE Inhibitory Activity of New Benzothiazole-Piperazines
a
a
b,c
b,c
Ümide Demir Özkay , Özgür Devrim Can , Begüm Nurpelin Sağlık , Ulviye Acar Çevik
b,c
* b,c
d
d
Serkan Levent , Yusuf Özkay , Sinem Ilgın , Özlem Atlı
a
Department of Pharmacology, Faculty of Pharmacy, Anadolu University, 26470 Eskişehir,
Turkey
b
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Anadolu University, 26470
Eskişehir, Turkey
c
Doping and Narcotic Compounds Analysis Laboratory, Faculty of Pharmacy, Anadolu
University, 26470 Eskişehir, Turkey
d
Department of Toxicology, Faculty of Pharmacy, Anadolu University, 26470 Eskişehir,
Turkey
*
Corresponding author
Yusuf ÖZKAY
E-mail address: yozkay@anadolu.edu.tr
Tel: +90-222-3350580/3750
Fax: +90-222-3350750.
Address: Anadolu University, Faculty of Pharmacy, Department of Pharmaceutical
Chemistry, 26470, Eskişehir, Turkey.
1

ABSTRACT
In the current study, 14 new benzothiazole-piperazine compounds were designed to meet the
structural requirements of acetylcholine esterase (AChE) inhibitors. The target compounds
were synthesised in three steps. Structures of the newly synthesised compounds (7-20) were
confirmed using IR, 1H-NMR, 13C-NMR, and HRMS methods. The inhibitory potential of
the compounds on AChE (E.C.3.1.1.7, from electric eel) was then investigated. Among the
compounds, 19 and 20 showed very good activity on AChE enzyme. Kinetics studies were
performed to observe the effects of the most active compounds on the substrate–enzyme
relationship. Cytotoxicity studies, genotoxicity studies, and theoretical calculation of
pharmacokinetics properties were also carried out. The compounds 19 and 20 were found to
be nontoxic in both of the toxicity assays. A good pharmacokinetics profile was predicted for
the synthesised compounds. Molecular docking studies were performed for the most active
compounds, 19 and 20, and interaction modes with enzyme active sites were determined.
Docking studies indicated a strong interaction between the active sites of AChE enzyme and
the analysed compounds.
Keywords: Alzheimer disease; acetylcholinesterase; benzothiazole; piperazine; docking.
2

Alzheimer’s disease (AD) is a progressive and fatal neurodegenerative disorder
1
-4
characterised by memory loss and behavioural disturbances. The worldwide prevalence of
4
-6
this tragic public problem has been estimated to reach 106.2 million by 2050. Various
pathological hallmarks, such as accumulation of β amyloid in senile plaques, hyper-
phosphorylated neurofibrillary tangles of tau protein, and loss of cholinergic activity in certain
7
–10
parts of brain, have been shown to be responsible for AD.
Currently, only the non-
competitive N-methyl-D-aspartate receptor antagonist, memantine, and the cholinesterase
inhibitors, tacrine, donepezil, rivastigmine, and galantamine, have been approved by the US
1
1-13
Food and Drug Administration for the treatment of AD.
Among cholinesterase inhibitors,
galantamine and donepezil selectively inhibit acetylcholinesterase (AChE), whereas
rivastigmine and tacrine non-selectively inhibit both AChE and butyrylcholinesterase
1
4
(
BChE).
The brain cholinergic system is vital for learning and memory consolidation and is
8
,10,15,16
known to be disrupted in AD.
The synaptic cholinergic transmission is regulated by
1
7
AChE and BChE enzymes that rapidly hydrolyse acetylcholine (ACh). AChE shows more
hydrolytic activity than BChE does. Hence, AChE inhibitors are preferred in the treatment of
1
8
AD to keep ACh levels normal. Donepezil is the most favourable AChE inhibitor since it
gives a relatively positive response in AD treatment. Furthermore, compared to other AChE
inhibitors, donepezil has some advantages, such as blood–brain barrier permeability, non-
1
9-20
hepatotoxicity, the least side efficacy, and consumption once daily.
2
1-31
Previous studies
have shown that AChE possesses a 20 Å-long, narrow gorge, which
contains five separated ligand-binding sites: 1) the catalytic triad, including Ser 203, His 447,
and Glu 334, which are located at the bottom of the gorge and directly participate in the
2
1
catalytic cycle; 2) the oxygen anion hole containing Gly 121, Gly 122, and Ala 204 and has
a role in the arrangement of hydrogen bond donors that stabilise the tetrahedral enzyme-
3

2
2,23
substrate complex;
3) the catalytic anionic site (CAS), a region where Trp86, Glu202, and
Tyr337 are situated. This site is responsible for the orientation and stabilisation of the
2
4-27
trimethylammonium group of ACh by forming cation-π interactions.
4) The acyl pocket
2
8
covers two phenylalanine residues, 295 and 297, which bind the acetyl group of ACh. 5)
The peripheral anionic site (PAS) comprises residues Tyr 72, Tyr 124, Trp 286, and Asp 74,
2
9-31
which are located at the entrance of the narrow gorge.
Tacrine and galantamine bind to PAS, whereas some tacrine dimers, donepezil and its
3
2-36
analogue BYYT-25, bind simultaneously to both PAS and CAS.
Studies suggest that
AChE inhibitors should bear a core ring system that interacts with PAS, a basic centre that
binds to CAS, and a linker, such as –O–, CH
2
, CONH, and CONH(CH
2
)n, between the core
3
7–39
ring system and the basic centre to fulfil structural requirements.
For example, AChE
inhibitors donepezil and BYYT-25 contain an indanone core ring, methylene or oxygen
linkers, and benzylpiperidine or benzylpyrrolydine basic centres. The dimethoxyindanone
moiety of donepezil binds to the PAS, whereas the benzylpiperidine moiety interacts with the
3
3-35
36
CAS.
BYYT-25 shows similar interactions with CAS and PAS.
On the basis of the above information, we designed a new series of benzothiazole–
piperazine derivatives (7-20) that have the aforementioned structural requirements (Figure 1).
Benzothiazole is an important ring system in the drug discovery studies of AD. There are
4
0-46
several benzothiazole compounds that show potential therapeutic effects against AD.
Furthermore, sabeluzole, a benzothiazole agent, delays the clinical progression of AD.
4
7,48
Thus, we selected benzothiazole as a core ring system. Structural similarity of benzothiazole
with donepezil and BYYT-25 was increased by substitution with methoxy at the C5 and C6
positions. The basic centre was established by bioisosterically replacing piperidine with
piperazine because numerous piperazine compounds have been reported as cholinesterase
49-53
inhibitors.
4

Figure 1: Structural requirements in design of the compounds 7-20.
In the present study, we synthesized novel compounds investigate their AChE inhibition
potential. The compounds 7-20 were synthesized as shown in Scheme 1. In the first step, 2-
aminobenzothiazole derivatives (3 and 4) were prepared via reaction of appropriate aniline (1
and 2), potassium thiocyanate, and bromine. In the second step, 2-chloro-N-(benzothiazol-2-
yl)acetamide derivatives (5 and 6) were synthesized via acetylation reaction by using
chloroacetyl chloride. Finally, substitution reaction between corresponding 1-substituted
piperazines and 2-chloro-N-(benzothiazol-2-yl)acetamide derivatives (5 and 6) yielded the
target compounds 7-20. The structures of the newly synthesized compounds (7-20) were
1
13
54
elucidated using IR, H-NMR, C-NMR, and HRMS methods.
Assessment of the compounds 7-20 as AChE (E.C.3.1.1.7, from electric eel) inhibitors
55
was carried out using the in vitro modified Ellman’s spectrophotometric method. We used
donepezil as the reference drug in the enzymatic activity for the above process. Synthesized
5

-3
-9
compounds 7-20 along with donepezil were tested at 10 –10 M concentrations. The IC50
values obtained for these compounds are presented in Table 1.
1
1
2
Compound
7
R
Compound
14
R
R
-H
-H
-H
-H
-H
-H
-OCH
3
8
9
15
16
17
18
19
-OCH
3
-OCH
3
1
1
1
0
1
2
-OCH
3
-OCH
3
-OCH
3
13
-H
20
-OCH
3
Scheme 1. Synthesis pathway of the compounds 7-20.
6

Table 1. IC50±SD (µM) values against AChE (E.C.3.1.1.7, from electric eel) and some physicochemical
parameters of compounds 7-20 used in prediction of ADME profiles.
AChE
Comp.
MW
logP
tPSA
nON nOHNH
Vol
Vio
DLS
IC50±SD
(µM)
7
8
9
430.96
414.51
410.54
426.54
464.51
377.51
391.54
460.99
444.53
440.57
456.57
494.54
407.54
421.57
4.07
3.56
3.84
3.45
4.29
2.02
2.30
3.66
3.15
3.43
3.04
3.88
1.61
1.89
4.10
57.70
57.70
57.70
66.93
57.70
60.94
60.94
66.93
66.93
66.93
76.17
66.93
70.17
70.17
38.78
6
6
6
7
6
7
7
7
7
7
8
7
8
8
4
1
1
1
1
1
1
1
1
1
1
1
1
1
1
0
372.00
363.40
375.03
384.01
389.76
349.77
366.57
397.55
388.94
400.57
409.56
415.31
375.31
392.11
367.89
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
1.66
1.57
1.16
1.27
1.16
1.75
1.73
1.79
1.69
1.32
1.52
1.37
1.76
1.76
1.76
0.2196±0.012
0.2435±0.016
0.3259±0.019
0.3965±0.024
0.2742±0.015
0.0946±0.006
0.1167±0.009
0.1297±0.014
0.1754±0.009
0.2148±0.011
0.2519±0.014
0.1977±0.016
0.0462±0.004
0.0576±0.002
0.0287±0.005
1
1
1
1
1
1
1
1
1
1
2
0
1
2
3
4
5
6
7
8
9
0
Donepezil 379.50
MW: Molecular weight; log P: log octanol/water partition coefficient; tPSA: Total Polar Surface Area; nON:
number of Hydrogen acceptors; nOHNH: number of Hydrogen donors were calculated using Molinspiration
Calculation of Molecular Properties toolkit. DLS: Drug likeness Model Score was calculated using MolSoft
2
016 Drug-Likeness and molecular property prediction toolkit.
-6
All the compounds showed remarkable AChE inhibition activity. At 10
M
concentration, the inhibition rate of all the compounds was more than 50%. Compounds 12,
-
7
1
1
3, 19, and 20 also showed more than 50% inhibition at 10 M concentration. Compounds 7,
-3
9, and 20 showed more than 95% enzyme inhibition at 10 M concentration and
-4
approximately 90% enzyme inhibition at 10 M concentration. Regarding AChE inhibitory
7

activity, all of the 6-methoxybenzothiazole derivatives (7-13) were less potent than the 5,6-
dimethoxybenzothiazole compounds (14-20) were. Compounds containing the
dimethylaminoethyl (12 and 19) and dimethylaminopropyl (13 and 20) moieties at the fourth
position of piperazine showed stronger inhibition than did the compounds 7-11 and 14-18 that
carry benzyl substitutions at the same position. Compounds 19 and 20, which possess these
two structural features, were found to be the most active derivatives in the series. The IC50
values of donepezil, compound 19, and compound 20 were found to be 0.0287, 0.0462, and
0
.0576 µM, respectively. These data indicate that the AChE inhibition potential of
compounds 19 and 20 is close to that of donepezil. The other compounds (7-18) also have
good IC50 in the range of 0.0942–0.3965 µM. As a result, all the newly synthesized
compounds turned out to be potent inhibitors of AChE with a low micromolar range of IC50
.
The mechanism of AChE inhibition was investigated via enzyme kinetics by using the
5
6
Ellman’s spectrophotometric method. Linear Lineweaver-Burk graphics were used to
observe the type of inhibition. Further, we analyzed the enzyme kinetics by recording
substrate-velocity curves in the absence and presence of the most potent compound 19
(
IC50=0.0462 µM). In each case, initial velocity measurements were obtained at different
substrate (ATC) concentrations ranging from 600 µM to 18.75 µM. The Ki (intercept on the
x-axis) values of the compounds 19 and 20 were calculated from the secondary plot of the 1/V
versus concentrations of compounds.
The graphical analyses of steady-state inhibition data for the compounds 19 and 20 are
presented in Figures 2 and 3. Very similar effects of these compounds on enzyme kinetics
were observed. In the figures, the lines cross neither x- nor y-axis at the same point. Different
Vmax and Km values were obtained for various concentrations that are presented along with
the Ki values in Table 2. Thus, the Lineweaver–Burk plot reveals that compounds 19 and 20
8

5
7,58
are typically mixed AChE inhibitors, which show significant similarity to donepezil.
result also indicates that both these compounds interact with both CAS and PAS of AChE.
A)
The
(
(
B)
Figure 2. (A): Lineweaver-Burk plot for the inhibition of AChE (E.C.3.1.1.7, from electric eel) by compound 19
at different concentrations of substrate (ATC). (B): Secondary plot for calculation of steady-state inhibition
i
constant (K ) of compound 19.
9

(
A)
(
B)
Figure 3. (A): Lineweaver-Burk plot for the inhibition of AChE (E.C.3.1.1.7, from electric eel) by compound 20
at different concentrations of substrate (ATC). (B): Secondary plot for calculation of steady-state inhibition
i
constant (K ) of compound 20.
1
0

m i
Table 2. Vmax, K and K values of compounds 19 and 20.
Comp.
Concentration
µM)
.0231
0.0462
Km
Ki
Vmax
abs/min)
-
1
(
(
(mM) (µM)
0
0.46
0.37
0.31
0.47
0.40
0.39
0.60
1.27
1
2
9
0
1.19
1.15
1.28
1.25
1.38
1.23
0.11
0.25
0
0
.0924
.0288
0.0576
.1152
0
Control
Toxicity is the main reason for the failure at all stages of the new drug development
process. The major part of safety-related attrition occurs at preclinical phases while predicting
preclinical safety liabilities earlier in the drug development process. This strategy enables the
design and/or selection of improved drug candidates that have more possibilities to become
5
9
commercialized drugs. Therefore, we used the MTT cell viability assay to determine
cytotoxicity against NIH/3T3 mouse embryonic fibroblast cell lines (ATCC CRL1658), which
6
0
is recommended by ISO (10993-5, 2009).
Results from the cytotoxicity evaluation of the most active compounds 19 and 20 are
presented in Table 3. IC50 values of donepezil, compound 19, and compound 20 against
NIH/3T3 cells were found to be 316 µM, ≥1000 µM, and 100 µM, respectively. IC50 values of
compounds 19 and 20 against NIH/3T3 cells are higher than their IC50 values against AChE.
Thus, it can be stated that these compounds are nontoxic at their effective concentrations
against AChE. Furthermore, donepezil showed approximately 3-fold lower cytotoxicity than
compound 19. Thus, cytotoxicity test findings enhanced the importance of compounds 19 and
2
0 as AChE inhibitors.
1
1

Table 3. IC50±SD(µM) value of compounds 19, 20 and donepezil against NIH/3T3 cell line.
20 Donepezil
19
IC50 (µM)
≥ 1000
100.17±16.49 316.42±21.26
Identification of mutagenic properties of new compounds is essential for safety, and
thus, new drug candidates should be examined using the models of genotoxicity, such as the
6
1
Ames test. In the current study, the Ames assay was performed to investigate the
MPF
genotoxicity of compounds 19 and 20. In Ames
assay, more than 25 positive wells were
observed with positive controls and negative control wells also showed less than eight
positive wells in the presence and absence of S9 with TA98 and TA100, which complied with
MPF
the requirements for the validation of the Ames
and also as described in previous
6
2
studies. Genotoxicity results are presented in Table 4.
MPF
Table 4. The AMES
results of the compounds.
Comp. Concentration
REVERTANTS Fold increase
(over baseline)
(mg/mL)
TA 98 TA 100
S9+
S9-
0.87*
1.14
1.03
0.33
0.33
0.11*
0.94
S9+
S9-
0.66
1
9
0.156
0.22
0.65
0.78
0.55
0.55
1.09*
0.55
0.47
0.98
0.60
0.53
0
.3125
.625
.25
0.49*
0.27*
0.27***
0.00***
0.00***
0.79
0
0.97
1
0.43
2.5
0.00*
0.00*
0.28
5
20
0.156
.3125
.625
.25
0
1.40*** 0.47
0.90
0
0.56
0.84
1.02
1.04*
0.69
1
0.52 1.36*** 1.32***
1.15* 1.36*** 0.32*
2.5
5
1.49*** 0.89 1.43*** 0.05***
*
t test p value (unpaired 1-sided) < 0.05 *** t test p value (unpaired 1-sided) < 0.001
1
2

Compound 19 showed a baseline of 3.08 and 6.13 against TA98 with and without S9,
respectively. Fold inductions over baseline did not reach values more than 1.5 and statistically
different results did not reveal any dose–response tendency. According to these findings,
compound 19 did not show any mutagenicity against TA98 (Figure 4). Compound 19 was
found to show a baseline of 4.28 and 6.10 with and without S9 against TA100, respectively.
Mentioned-fold increases over the baseline according to the criteria were not determined with
compound 19 and significant results did not reach these values and did not show any dose-
response tendency. Compound 19 was also found to be non-mutagenic against TA100 in the
presence or absence of metabolic activation (Figure 4).
Figure 4. Dose-response curve of compound 19 against TA98 and TA100 in the presence and absence of S9
MPF
according to AMES
test.
TA 98
-S9)
TA 100
(-S9)
(
50
40
30
20
10
0
50
40
30
20
10
0
0
0,16 0,31 0,63 1,25 2,5
mg/ml
5
0
0,16 0,31 0,63 1,25 2,5
mg/ml
5
TA 98
TA 100
(+S9)
(+S9)
50
40
30
20
10
0
50
40
30
20
10
0
0
0,16 0,31 0,63 1,25 2,5
mg/ml
5
0
0,16 0,31 0,63 1,25 2,5
mg/ml
5
*
t test p value (unpaired 1-sided) < 0.05 with a fold-induction over baseline > 2; -------: 2-fold induction over
baseline
1
3

Compound 20 showed a baseline of 6.40 with TA98 in the absence of S9 and a
baseline of 3.58 in the presence of S9. Although there were significant differences observed,
they did not reach the mentioned values above the baseline and did not show any dose–
response tendency. Therefore, compound 20 was classified as non-mutagenic against TA98 in
the presence or absence of metabolic activation (S9) (Figure 5). Compound 20 had a baseline
of 6.32 with TA100 in the absence of S9 and 4.42 in the presence of S9. Furthermore, fold
inductions above the baseline were less than 1.5 in each concentration of the compound and
the significantly different results obtained did not show any dose–response tendency.
Therefore, compound 20 was not genotoxic against TA100 with or without metabolic
activation (Figure 5).
Figure 5. Dose-response curve of compound 20 against TA98 and TA100 in the presence and absence of S9
MPF
according to AMES
test.
TA 98
-S9)
TA 100
(-S9)
(
50
40
30
20
10
0
50
40
30
20
10
0
0
0,16 0,31 0,63 1,25 2,5
mg/ml
5
0
0,16 0,31 0,63 1,25 2,5
mg/ml
5
TA 98
TA 100
(+S9)
(+S9)
50
40
30
20
10
0
50
40
30
20
10
0
0
0,16 0,31 0,63 1,25 2,5
mg/ml
5
0
0,16 0,31 0,63 1,25 2,5
mg/ml
5
*
t test p value (unpaired 1-sided) < 0.05 with a fold-induction over baseline > 2; -------: 2-fold induction over
baseline
1
4

MPF
On the basis of the results of the Ames
assay, the compounds were classified as
negative. Further, cytotoxicity and genotoxicity findings strongly suggested compounds 19
and 20 to be AChE inhibitors.
High pharmacological activity and low toxicological effects are not enough for a
compound to become a drug candidate. A good pharmacokinetics profile is also very
important for the new drug candidates that should be evaluated earlier in the process of drug
development. In recent years, significant developments in combinatorial chemistry have made
the estimation of absorption, distribution, metabolism and excretion (ADME) relatively
6
3
easy. ADME properties of the newly synthesized compounds (7-20) were calculated using
6
4
online Molinspiration property program. This program provides the data based on Lipinski’s
rule, which assesses the ADME properties of new compounds and is important for the
optimization of a biologically active compound. The rule emphasizes that an orally active
6
5
drug should not possess more than one violation. Drug-likeness score (DLS) was also
calculated for all the compounds (7-20) and donepezil based on the Molsoft's chemical
fingerprints mode consisting of 5K of marketed drugs from World Drug Index (positives) and
6
6
1
0K of carefully selected non-drug compounds (negatives). In the software, DLS score was
found to be between 0 and 2, suggesting good pharmacokinetics for drug candidates.
The theoretical calculations of ADME parameters (molecular weight (MW), log P,
topological polar surface are (tPSA), number of hydrogen donors (nON) and acceptors
(
nOHNH), and volume) and DLS are presented in Table 1 along with the violations of
Lipinski’s rule. According to these data, all compounds (7-20) confirm the Lipinski’s rule by
causing no violation. Furthermore, the same DLS (1.76) was obtained for the most active
compounds (19 and 20) and donepezil. This suggests that newly synthesized compounds may
have a good pharmacokinetics profile, which further strengthens their biological importance.
1
5

Docking of compounds 19 and 20 into the active site of AChE was carried out
6
7
separately by using AutoDock Vina software to visualize the possible interactions. The 3D
structure of human AChE (hAChE, PDB ID:4EY7), which includes E2020 (donepezil) as a
ligand, was retrieved from the Protein Data Bank server (www.rcsb.org). The docking poses
for both the compounds are shown in Figure 6. According to the docking results, compounds
1
9 and 20 showed compatibility with the gorge and interaction with both CAS and PAS.
They interact with Trp86, Tyr124, Ser203, Trp286, His287, Leu289, and Tyr341.
Benzothiazole structure settles down PAS and forms a π-π interaction with the indole ring of
Trp286. The methoxy substituents provide polar interactions with amino group of Trp286,
His287, and Leu289 by forming hydrogen bonds. The carbonyl of amide moiety creates a
hydrogen bond with the amino group of Tyr124. In the CAS, a hydrogen bond is formed
between the nitrogen of the dimethylamino group and carbonyl of Ser203. The dimethylamino
group and Trp86 also interact with cation-π. The ethyl/propyl group and terminal dimethyl
group intensify the binding with the active site via Van der Waals interactions. All of these
interactions enable to explain the proper binding with the active region of AChE.
Realizing chemical structures of the existing drugs and constituting the structural
requirements for intrinsic pharmacological activity is an important approach in designing
novel compounds. On the basis of this strategy, we assessed 14 new benzothiazole analogues
as AChE inhibitors in the present study. Pharmacological, toxicological, and ADME studies
indicated the relative potency of compounds 19 and 20 against rest of the compounds.
Molecular docking studies suggested possible interactions between these compounds and
AChE. Consequently, all these data may pave the way for the researchers to synthesize
similar compounds possessing enhanced pharmacological profile.
1
6

Figure 6. Three dimensional models of compounds 19 (A) and 20 (B) in the active site of hAChE.
(
A) Compound 19 in hAChE active site
(
B) Compound 20 in hAChE active site
Acknowledgement
This study was financially supported by Anadolu University Scientific Projects Fund, Project
No: 1605S314.
1
7

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2
2

Highlights


New benzothiazole-piperazines were designed based on structural requirements for
AChE inhibitors.
Compounds 19-20 indicated significant inhibition on AChE (E.C.3.1.1.7, from electric
eel).


These compounds did not display cytotoxicity and genotoxicity.
A good pharmacokinetics profile and drug likeness score were predicted for the
synthesized compounds.

Interaction modes between AChE and compounds 19 and 20 were determined by
docking studies.
2
3

Design, Synthesis, and AChE Inhibitory Activity of New Benzothiazole-Piperazines
Fourteen new benzothiazole-piparazines were designed and synthesized in order to investigate
their AChE inhibition potency. Characterized compounds were subjected to pharmacological,
toxicological, ADME and docking studies.
Core Ring
System
Basic
Center
Linker
Linker
Core Ring
System
Basic
Center
N
N
N
O
O
O
O
O
O
O
N
O
O
Core Ring
System
Basic O
Center
Linker
O
Donepezil
BYYT-25
O
O
Core Ring
Donepezil
System
Basic
Center
O
Linker
Docking
BYYT-25
Design
R2
N
R : -H, -OCH3
1
R1
O
H
N
N
N
O
O
O
R : Benzyl, 4-Substitutedbenzyl, Dimethylaminoalkyl
2
N
S
O
O
R2
N
N
1
O
Compounds 7-20
N
R : -H, -OCH
3
Donepezil IC50 = 0.0287 µM
Comp. 19 IC = 0.0462 µM
R1
O
N
O
BYYT-25
O
H
N
O
2
AChE
R : Benzyl, 4-Substitutedbenzyl, Dimethylaminoalky50
l
N
Inhib Oi tion
S
O
O
O
Compounds 7-20
Donepezil
BYYT-25
R2
1
R : -H, -OCH
3
N
2
R : Benzyl, 4-Substitutedbenzyl, Dimethylaminoalkyl
N
R2
R : -H, -OCH3
1
R1
N
N
O
H
N
R : Benzyl, 4-Substitutedbenzyl, Dimethylaminoalkyl
2
N
S
O
Compounds 7-20