Design, synthesis and biological evaluation of 2-acetyl-5-O-(amino-alkyl)phenol derivatives as multifunctional agents for the treatment of Alzheimer's disease

Article abstract of DOI:10.1016/j.bmcl.2017.09.057

A series of 2-acetyl-5-O-(amino-alkyl)phenol derivatives was designed, synthesized and evaluated as multi-function inhibitors for the treatment of Alzheimer's disease (AD). The results revealed that compound TM-3 indicated selective AChE inhibitory potenc

Full text of DOI:10.1016/j.bmcl.2017.09.057

Accepted Manuscript
Design, synthesis and biological evaluation of 2-acetyl-5-O-(amino-alkyl)phe-
nol derivatives as multifunctional agents for the treatment of Alzheimer’s
disease
Zhipei Sang, Keren Wang, Huifang Wang, Huijuan Wang, Qianwen Ma, Xue
Han, Mengyao Ye, Lintao Yu, Wenmin Liu
PII:
S0960-894X(17)30966-6
https://doi.org/10.1016/j.bmcl.2017.09.057
BMCL 25321
DOI:
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
29 July 2017
22 September 2017
27 September 2017
Please cite this article as: Sang, Z., Wang, K., Wang, H., Wang, H., Ma, Q., Han, X., Ye, M., Yu, L., Liu, W., Design,
synthesis and biological evaluation of 2-acetyl-5-O-(amino-alkyl)phenol derivatives as multifunctional agents for
the treatment of Alzheimer’s disease, Bioorganic & Medicinal Chemistry Letters (2017), doi: https://doi.org/
10.1016/j.bmcl.2017.09.057
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Design,
synthesis
and
biological
evaluation
of
2-acetyl-5-O-(amino-alkyl)phenol derivatives as multifunctional
agents for the treatment of Alzheimer’s disease
Zhipei Sanga, *, Keren Wangb, Huifang Wangc, Huijuan Wanga, Qianwen Maa, Xue
Hana, Mengyao, Yea, Lintao Yua, Wenmin Liua, *.
aCollege of Chemistry and Pharmaceutical Engineering, Nanyang Normal University,
Nanyang, 473061, China
bNanyang Normal University Hospital, Nanyang Normal University, Nanyang,
473061, China
cNanyang Normal University Library, Nanyang Normal University, Nanyang, 473061,
China
*Corresponding authors.
E-mail: sangzhipei@126.com (Zhipei Sang)
E-mail: liuwm1969@163.com (Wenmin Liu)
Abstract
A series of 2-acetyl-5-O-(amino-alkyl)phenol derivatives was designed, synthesized
and evaluated as multi-function inhibitors for the treatment of Alzheimer’s disease
(AD). The results revealed that compound TM-3 indicated selective AChE inhibitory
potency (eeAChE, IC50 = 0.69 µM, selective index (SI) = 32.7). Both kinetic analysis
of AChE inhibition and molecular modeling study suggested that TM-3 could
simultaneously bind to the catalytic active site and peripheral anionic site of AChE.
And TM-3 was also a highly selective MAO-B inhibitor (IC50 = 6.8 µM). Moreover,
TM-3 could act as antioxidant (ORAC value was1.5eq) and neuroprotectant, as well
as a selective metal chelating agent. More interestingly, compound TM-3 could cross
the blood-brain barrier (BBB) in vitro and abided by Lipinski’s rule of five. Therefore,
compound TM-3, a promising multi-targeted active molecule, offers an attractive
1

starting point for further lead optimization in the drug-discovery process against AD.
Keywords: Alzheimer’s disease; 2-acetyl-5-O-(amino-alkyl)phenol derivatives;
designed; synthesized; multi-target inhibitor; blood−brain barrier.
Abbreviations: AD, Alzheimer’s disease; AChEIs, acetylcholinesterase inhibitors;
NMDA, N-methyl-D-aspartate; Aβ, β-amyloid; τ, tau protein; ACh, acetylcholine;
MTDLs, multi-target-directed ligands; MAO-B, monoamine oxidase B; MAO-A,
monoamine oxidase A; BuChE, butyrylcholinesterase; EeAChE, Electrophorus
electricus AChE; eqBuChE, equine serum BuChE; PAS, peripheral anionic site; CAS,
catalytic active site; ORAC-FL, oxygen radical absorbance capacity by fluorescein;
MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium; PAMPA-BBB, parallel
artificial membrane assay for the blood- brain barrier; SD, standard deviation; MW,
molecular weight; TPSA, topological polar surface area; ADME, absorption,
distribution, metabolism, excretion.
2

Alzheimer’s disease (AD) is an age-related neurodegenerative disease
characterized by deterioration of memory, language skills, and other cognitive
impairments in elder people. This situation intensely influences the patient’s social
life and activity. It is evaluated by World Alzheimer’s Report that more than 35
million people suffered from AD in 2015, and this number will be double by 2030 and
approximately triple to 131 million by 2050.1 Current clinical treatment of AD is
mainly focused on controlling symptoms by providing temporary improvement. To
date, there are four acetylcholinesterase inhibitors (AChEIs), named donepezil, tacrine,
rivastigmine, galantamine and one N-methyl-D-aspartate (NMDA) receptor antagonist
memantine have been approved for the treatment of AD. Unfortunately, these drugs
are just palliative treatment and do not address the molecular mechanisms that
underlie the pathogenic processes due to the multifactorial nature of AD.2
The etiology of AD is not completely known, but several characteristic
pathological features such as β-amyloid (Aβ) deposits, tau protein (τ) aggregation,
oxidative stress, decreased level of acetylcholine (ACh), neuroinflammation, and
dyshomeostasis of biometals have been thought to play significant roles in the
pathogenesis of AD.3 In this regard, one single drug that acts on a specific target to
produce the desired clinical effects might not be suitable for the complex nature of
AD, the development of multitarget-directed ligands (MTDLs), i.e., single chemical
entities able to hit different targets involved in the cascade of AD pathological events,
to act as multifunctional agents to treat this disease has been applied by many research
groups, and the results obtained have been encouraging and convince researchers that
MTDLs might present the best pharmacological option for tackling the multifactorial
nature of AD and for halting the progression of the disease.4-6 Several MTDLs
candidate drugs with disease modifying potential are now in the pipeline and have
reached testing stage in clinical trials.7 Therefore, the discovery of a lead compound
that can modulate multi-factors simultaneously is a crucial step in the search for a
candidate for the clinical treatment of AD.
2-Acetyl-5-alkoxyphenol analog DDDT-2d, is a potent and selective MAO-B
inhibitor, the IC50 values of MAO-B was 2.9 nM, as well as 17000-flod selective for
3

MAO-B over the MAO-A.8 Recent studies shows that the selective MAO-B inhibitor
has been shown to significantly improve learning and memory deficits in animal
models associated with AD and to slow the disease progression in AD patients,9 and
seem to be an important treatment of AD.10 So, it is appropriate for the design of
therapies for neurodegenerative disorders such as Alzheimer’s disease. Recently, our
group has reported the synthesis of scutellarein-O-alkylamine derivatives and
discovered representative EJMC-16d as a potential multifunctional agent for the
treatment, and the alkylamine side chain is required pharmacophore for the AChE
inhibition.11 Therefore, in this paper, 2-acetylphenol was selected to combine with
different length alkylamine fragment to design
a
series of novel
2-acetyl-5-O-(amino-alkyl)phenol derivatives, to evaluate whether these novel
molecules might possess more potency in various multifunctional activities.
In this study, we report the study of the design, synthesis and evaluation of a
series of 2-acetyl-5-O-(amino-alkyl)phenol derivatives based on MTDLs (Figure 1).
They were found to show potentially applicable biological activities, including
inhibition of ChEs (AChE and BuChE) and MAOs (MAO-A and MAO-B),
antioxidant properties, neuroprotective effects, and metal chelation.
Figure 1. Design strategy for the 2-acetyl-5-O-(amino-alkyl)phenol derivatives.
4

Total 20 2-acetyl-5-O-(amino-alkyl)phenol derivatives were synthesized in good
yields. The synthetic pathway of these target compounds was outlined in Scheme 1.
Briefly, the starting material 1 was treated with excessive amounts of
1,3-dibromopropane (2a), 1,4-dibromobutane (2b), 1,5 –dibromopentane (2c) or
1,6-dibromohexane (2d) in the presence of K2CO3 in CH3CN at 60–65°C to obtain the
intermediates 3a~3d. Subsequently, the target compounds TM-1~TM-20 were got by
the reaction of intermediates 3a~3d with secondary amines (A~E) in the presence of
K2CO3 in anhydrous CH3CN at 60–65°C. All target compounds were purified by
chromatography, and the analytical and spectroscopic data confirmed their structures,
as detailed in the experimental section.
Scheme 1. Synthesis of target compounds TM. Reagents and conditions: (i) Br(CH2)nBr (2a~2d),
K2CO3, acetone, reflux, 6-8 h; (ii) K2CO3, NR1R2 (A~E) (Structures of NR1R2 are shown in
Figure 1) anhydrous CH3CN, 65°C, 6-10h.
The 2-acetyl-5-O-(amino-alkyl)phenol derivatives TM-1~TM-20 were tested in
vitro for their cholinesterase inhibitory activities were determined by the modified
Ellman method using eeAChE (from electric eel) and eqBuChE (from equine
serum).12 Donepezil, a well-known cholinesterase inhibitor approved by FDA, was
used as the positive control, and the precursor compound 2,4-dihydroxyacetophenone
(1) was also tested, the results are listed in Table 1. Compared with compound 1 and
donepezil, all the target compounds were selective AChE inhibitors, and showed
moderate to good EeAChE inhibition potent with IC50 values ranging from 0.96 to
57.0 µM. It revealed that the introduction of O-alkylamines increased ChEs inhibitory
activity and improved the selectivity for eeAChE over eqBuChE, which was
consistent with our previous work.11,13,14 The selectivity may be helpful to diminish
5

peripheral cholinergic side effects. The results data also displayed that both the
structure of terminal groups NR1R2 of side chain and the methylene chain length
significantly affected the AChE inhibitory potencies. When the terminal groups
NR1R2 was piperidine and pyrrolidine, the AChE inhibitory activity remarkably
enhance as the methylene chain increases, for example, TM-1 < TM-6 < TM-11 <
TM-16; TM-2 < TM-7 < TM-12 < TM-17, while the compounds containing two
basic
centers
(benzylpiperazine,
1-(2-pyridyl)piperazine
and
2-(1-piperazinyl)pyrimidine group) demonstrated the opposite result, that is, the
inhibitory activity decrease gradually with the increase of methylene chain, such as
TM-3 > TM-8 > TM-13 > TM-18; TM-4 > TM-9 > TM-14 > TM-19; TM-5 >
TM-10 > TM-15 > TM-20. This phenomenon revealed that the alkylamine side chain
can interact with catalytic active site (PAS) of AChE in different ways. In addition, in
the same methylene chain length situation, and for the compounds possessing
piperazine groups, benzylpiperazine indicated better inhibitory potency than the other
groups. From the screening data, all the target compounds showed weak BuChE
inhibitory potency, and were significantly selective AChE inhibitors. Especially,
compound TM-3 was the best AChE inhibitor with IC50 value was 0.96µM, and the
selective index was 32.7.
Table 1 eeAChE and eqBuChE inhibitory activities, MAO-A and MAO-B inhibitory
potencies and oxygen radical absorbance capacity (ORAC, Trolox equivalent) of
target compounds and control compounds.
n
NR1R2
IC50 (µM)a
IC50 (µM)a
MAO-Be
SIf
ORACg
Comp.
eeAChEb
eqBuChEc
n.a. h
MAO-Ad
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
3
3
3
3
3
4
4
4
4
4
5
29.2 0.66
41.3 0.78
0.96 0.01
15.2 0.23
26.3 0.96
8.8 0.15
—
—
2.8 0.05
0.62 0.01
6.8 0.31
11.7 0.20
15.1 0.36
2.5 0.08
1.8 0.04
15.9 0.11
12.0 0.33
15.2 0.77
6.1 0.14
TM-1
TM-2
TM-3
TM-4
TM-5
TM-6
TM-7
TM-8
TM-9
TM-10
TM-11
(A)
(B)
(C)
(D)
(E)
(A)
(B)
(C)
(D)
(E)
(A)
1.1 0.04
1.0 0.03
1.5 0.01
0.98 0.02
0.95 0.04
1.0 0.03
1.0 0.02
1.4 0.05
1.1 0.02
0.98 0.03
1.1 0.02
n.a. h
31.4 0.89
8.8 0.66% i
n.a. h
21.4 0.38% i
18.4 0.52% i
11.5 0.31% i
n.a. h
32.7
—
—
—
16.7 0.58
2.2 0.01
—
—
18.4 0.23
27.5 0.58
5.5 0.11
—
n.a. h
—
27.9 0.65% i
—
6

5
5
5
5
6
6
6
6
6
6.3 0.22
8.5 0.13
21.9 0.62
51.8 0.16
2.9 0.08
3.9 0.09
16.8 0.67
25.8 1.1
57.0 1.8
n.a. h
37.2 0.81
46.1 1.5
n.a. h
5.9
5.4
—
3.5 0.13
26.8 0.47
16.7 0.39
15.5 0.76
26.6 0.43
12.2 0.26
30.8 0.27
20.7 0.69
34.6 0.88
n.t. k
TM-12
TM-13
TM-14
TM-15
TM-16
TM-17
TM-18
TM-19
TM-20
1
(B)
(C)
(D)
(E)
(A)
(B)
(C)
(D)
(E)
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
n.a. j
1.0 0.04
1.3 0.02
0.99 0.03
1.0 0.02
0.98 0.04
1.0 0.02
1.3 0.03
1.1 0.04
0.98 0.05
2.5 0.03
—
137 1.2
45.7 0.79
46.5 0.64
75.4 0.83
n.a. h
n.a. h
n.a. h
2.6
15.8
11.9
4.5
—
—
n.t. k
n.t. k
n.t. k
donepezil
clorgyline
rasagiline
iproniazid
0.019 0.0003
n.t. k
4.76 0.02
n.t. k
251
—
0.0034 0.0001
5.02 0.02
0.086 0.003
1.78 0.01
—
n.t. k
n.t. k
n.t. k
n.t. k
—
2.13 0.01
—
—
3.18 0.03
—
a
Structures (n and NR1R2) are shown in Figure 1 and Scheme 1. IC50 values represent the
concentration of inhibitor required to decrease enzyme activity by 50% and are the mean of three
b
independent experiments, each performed in triplicate (SD = standard deviation). From
c
d
e
electrophorus electricusc. From equine serum. From recombinant human MAO-A. From
f
g
recombinant human MAO-B. SI (selectivity index) = IC50 (EeAChE)/IC50 (eqBuChE). Results
are expressed as µM of Trolox equivalent/µM of tested compounds. h n.a. = no active. Compounds
defined “no active” means percent inhibition less than 5.0% at a concentration of 50 µM in the
assay conditions. i the inhibition percent ratio of compounds for eqBuChE at a concentration of 50
j
µM in the assay conditions. n.a. = no active. Compounds defined “no active” means percent
inhibition less than 5.0% at a concentration of 10 µM in the assay conditions. k n.t. = no test.
The Lineweaver–Burk plots (Figure 2) showed that both inhibitions had rising
slopes and increasing intercepts at higher concentration, which indicated a mixed-type
inhibition.13 The result indicated that compound TM-3 could bind to both CAS and
PAS of AChE.
Figure 2. Steady state inhibition by compound TM-3 of the AChE hydrolysis of ACh. The plots
show mixed-type AChE inhibition for compound TM-3.
7

In order to explore possible interactions of compounds with the enzyme EeAChE,
compounds TM-3 and TM-16 were selected to do molecular modeling study using
the docking program, AutoDock 4.2 package with Discovery Studio 2.5, based on the
X-ray crystal structure of TcAChE (PDB code: 1EVE).12 TM-3 was the most AChE
inhibitor with three methylene chain length, As shown in Figure 3, the results
revealed that TM-3 occupied the entire enzymatic catalytic site (CAS), the mid-gorge
sites and the peripheral site (PAS), and could simultaneously bind to both the CAS
and the PAS thus providing a reasonable explanation for its highly potent inhibitory
potency against AChE. In the TM-3-TcAChE complex, the 2-hydroxy and carbonyl
group at the 2,4-dihydroxyacetophenone nucleus could simultaneously bind to
ARG289 via two intermolecular hydrogen bonds, and the 2-hydroxy additionally bind
to PHE288 via a intermolecular hydrogen bond. In addition, the chain of methylene
and piperazine ring could fold into a conformation in the gorge that allowed them to
interact with TYR121, SER122, PHE330, PHE331 and TYR334 through hydrophobic
interaction. Meanwhile, the benzene ring of benzylpiperazine was observed to bind to
the CAS via potential hydrophobic interactions with residues TYR70, ASP72, TRP84
and ASN85. Therefore, both the kinetic study of AChE inhibition and the molecular
docking showed that TM-3 was a highly potent AChE inhibitor and was reasonably
selected to further study. As shown in Table 1, TM-16 was also a potent AChE
inhibitor, but with six methylene chain length. The docking result was shown in
Figure S1, TM-16 also has one intramolecular hydrogen bond between the 2-hydroxy
and carbonyl group of the 2,4-dihydroxyacetophenone nucleus, the oxygen atom of
2-hydroxy group could bind to ARG289 and PHE288 via two intermolecular
hydrogen bonds, respectively, and the hydrogen atom of 2-hydroxy group could also
bind to ARG289 via a intermolecular hydrogen bond, moreover, TM-16 was observed
to bind to the PAS and CAS via potential hydrophobic interactions with residues
TYR121, SER122, PHE330, PHE331, TYR334, TYR70, ASP72 and TRP84, these
phenomenon accord with the docking of TM-3. However, lack of the intermolecular
hydrogen bond between carbonyl group of the 2,4-dihydroxyacetophenone nucleus
and ARG289 presented compared with TM-3. That’s maybe the reason why TM-16
8

showed the weaker AChE inhibitory activity than that of TM-3.
A
B
Figure 3 (A) 3D docking model of compound TM-3 (green stick) interacting with residues in the
binding site of TcAChE (PDB code: 1EVE). (B) 2D schematic diagram of docking model of
compound TM-3 with TcAChE.
The target compounds were also investigated for their inhibitory activity against
hMAOs using clorgyline, rasagiline and iproniazid as reference drugs. Recombinant
human MAO-A and –B were employed as enzyme sources, and their activities were
determined by a fluorimetric method with kynuramine as a common substrate.15,16 As
shown in Table 1, all the synthesized target compounds were significantly selective
9

hMAO-B inhibitors, and displayed moderate to good MAO-B inhibition (IC50 values
range from 0.62 to 34.6 µM). Among of which, compound TM-2, with pyrrolidine,
exhibited the most potent MAO-B inhibitory potency with IC50 values of 0.62 µM.
The screening data revealed that MAO-B inhibitory potency were susceptible to the
substituent groups NR1R2 and the length of methylene. In most situations, the
inhibitory activity sharply decreased with the increase of the mehtylene chain, for
example, TM-1 > TM-6 < TM-11 < TM-16; TM-2 < TM-7 < TM-12 < TM-17;
TM-3 < TM-8 < TM-13 < TM-18; TM-4 < TM-9 < TM-14 < TM-19; TM-5 <
TM-10 < TM-15 < TM-20. Generally speaking, expect compound TM-1, the optimal
length of linker was three. In addition, the substituent groups NR1R2 was also a key
role, under the same methylene chain condition, the potency to inhibit MAO-B were
in the order pyrrolidine > piperidine > benzylpiperazine > 1-(2-pyridyl)piperazine >
2-(1-piperazinyl)pyrimidine. The promising compound TM-3 was a potent selective
MAO-B inhibitor (IC50 = 6.8 µM), which was also beneficial for AD.
The antioxidant activities of all synthesized target compounds were determined
by following the well-established ORAC-FL method (oxygen radical absorbance
capacity by fluorescein)17,18 and the results were shown in Table 1. Trolox, a vitamin
E analogue, was used as the standard, and the results were expressed as Trolox
equivalents. 2,4-dihydroxyacetophenone was also tested, which has an ORAC-FL
value of 2.5 Trolox equivalents. All the tested compounds exhibited favorable
antioxidant capacities. As expected, the introduction of O-alkylamine group side chain
in the 2,4-dihydroxyacetophenone nucleus decreased the radical capture capacity but
the different tertiary amine fragment on the side chain has little influence on the
radical capture capacity. Generally speaking, the compounds possessing
benzylpiperazine group (TM-3, TM-8, TM-13 and TM-18) displayed higher radical
capture capacity than that of other substituent groups (pyrrolidine, piperidinem,
1-(2-pyridyl)piperazine and 2-(1-piperazinyl)pyrimidine). Compound TM-3 exhibited
the highest antioxidant potency, with an ORAC value of 1.5 Trolox equivalents.
The protective effect of TM-3 against free radicals damage was evaluated by
testing the ability of the compound to protect against H2O2 injury based on the
10

reported protocol.18 After 100 µM H2O2 exposure, cell viability as determined by
MTT reduction was markedly decreased to 46.5 % (P < 0.01 vs control), suggesting
high sensitivity to H2O2-induced injury. However, compound TM-3 showed
protective effect in a dose-dependent manner against H2O2-induced PC12 cell injury
(Figure 4). At concentrations of 10.0 µM, compound TM-3 showed remarkably
neuroprotective effect and the cell viability was 73.3%. When the concentration was
reduced to 1.0 µM, the cell viability decreased to 59.6%. It revealed that the
compound TM -3 could capture the hydroxyl radical, generated by H2O2.
Figure 4. Protective effects of TM -3 on cell injury induced by H2O2 (100 µM) in PC12 cells. ##
< 0.01 vs control; *P < 0.05 vs H2O2 group and **P < 0.01 vs H2O2 group.
P
To further study the complexing ability of the promising compound TM-3
toward biometals such as Cu2+, Fe2+, Zn2+ and Al3+, UV-visual spectrometry test was
performed.19,20 The results were shown in Figure 5. The electronic spectra of TM-3
exhibited a red shift (the peak at 314 nm shifted to 356 nm) after the addition of
CuCl2. But, when FeSO4, ZnCl2 and AlCl3 were added, the electronic spectra of TM-3
displayed no obvious change. Therefore, TM-3 was a selective metal chelator.
The stoichiometry of the TM-3-Cu2+ complex was determined using the molar
ratio method, by preparing the solution of compound TM-3 with ascending amounts
of CuCl2. The UV spectra were recorded and treated by numerical subtraction of
CuCl2 and TM-3 at corresponding concentrations at 356nm. As shown in Figure 6,
the absorbance linearly increased initially and then plateaued. The two straight lines
intersected at a mole fraction of 0.99, indicating a 1:1 stoichiometry for the complex
TM-3-Cu2+.
11

Figure 5. The UV spectrum of compound TM-3 (37.5 µM, in methanol) alone or in the presence
of CuCl2, FeSO4, ZnCl2 and AlCl3 (37.5 µM, in methanol).
Figure 6. Determination of the stoichiometry of complex-Cu2+ by using molar ratio method
through titrating the methanol solution of compound TM-3 with ascending amounts of CuCl2. The
final concentration of tested compound was 37.5 µM, and the final concentration of Cu2+ ranged
from 3.75 to 93.75 µM.
Brain penetration is a necessary requirement for successful anti-AD drugs. To
access the brain penetration of potential compound TM-3, a parallel artificial
membrane assay for the blood- brain barrier (PAMPA-BBB) described by Di et al was
performed.12,21 We compared the permeability of 11 commercial drugs with reported
values to validate the assay (Table 2). A plot of the experimental data versus the
reported values produced a good linear correlation, Pe (exp) = 0.9163Pe (bibl.)
−0.2247 (r2 = 0.9558) (Figure 7). From this equation, and considering the limit
−brain barrier permeation, we
established by Di et al. for blood
confirmed that
−brain
compound with permeability above 3.44×10−6 cm/s could cross the blood
barrier (Table 3). Based on the measured permeability (Table 4), TM-3 could cross
12

the BBB and reach the biological targets located in the CNS.
Table 2. Permeability (Pe×10-6 cm/s) in the PAMPA-BBB assay for 11 commercial
drugs used in the experiment validation.
Commercial drugs
Verapamil
Bibla
PBS:EtOH(70:30) b
16
16.90
9.60
Oxazepam
Diazepam
10
16
11.86
5.10
Clonidine
5.3
13
Imipramine
Testosterone
Caffeine
10.10
16.30
1.28
17
1.3
0.9
2.5
0.1
0.12
Enoxacine
Piroxicam
0.47
0.72
Norfloxacin
Theophylline
0.42
0.10
a Taken from ref. 21
b Data are the mean SD of three independent experiments
Figure 7. Linear correlation between experimental and reported permeability of
−0.2
commercial drugs using the PAMPA-BBB assay. Pe(exp) = 0.9163, Pe(bibl.)
(r2 = 0.9558).
247
Table 3. Ranges of permeability of PAMPA-BBB assays (Pe×10-6 cm/s)
13

Compounds of high BBB permeation (CNS +)
Compounds of uncertain BBB permeation (CNS +/-)
Compounds of low BBB permeation (CNS -)
Pe>3.44
3.44>Pe> 1.61
Pe<1.61
Table 4. Permeability Pe (×10−6 cm/s) in the PAMPA-BBB assay of the selected
compound TM-3 and the predictive penetration in the CNS.
Compounda
Pe (×10−6 cm/s)b
Prediction
CNS +
13.31 0.12
TM-3
aCompound TM-3 was dissolved in DMSO at 5 mg/mL and diluted with PBS/EtOH (70:30). The
final concentration of the compound was 100 µg/mL.
b Values are expressed as the mean SD of three independent experiments.
To further evaluate the drug-likeness property of compound TM-3, we applied
the Molinspiration property program to calculate the items including log P, molecular
weight (MW), topological polar surface area (TPSA), the number of hydrogen-bond
acceptors, and the number of hydrogen-bond donors.22 The theoretical prediction
properties of compound TM-3 was listed in Table 5. The screening data illustrated
showed that compound TM-3 did not break any point of the Lipinski’s rule of five,
and TPSA and log P values were compatible with those described as a predictive
indicator of a drug’s capacity for membrane penetration. Therefore, compound TM-3
might be a promising drug candidate and was selected to further investigate for the
treatment of AD.
Table 5. Theoretical prediction of the ADME properties of compound TM-3
Comp.
Log P
MW
TPSA (Å2)
n-ON
5
n-OHNH
1
volume (Å3)
TM-3
3.4
368.48
53.01
357.14
a MW, Molecular weight; TPSA, topological polar surface area; n-OH, number of hydrogen
acceptors; n-OHNH, number of hydrogen bond donors. The data was determined with the
Molinspiration calculation software.
In brief, a series of 2-acetyl-5-O-(amino-alkyl)phenol derivatives was designed,
synthesized and evaluated as multi-target-directed ligands for the treatment of
Alzheimer’s disease. Among the target compounds, compound TM-3 indicated highly
selective AChE inhibitory potency with IC50 values of 0.96 µM, both the kinetic
14

analysis and molecular modeling study suggested that TM-3 showed mixed-type
inhibition, and could bind to both CAS and PAS of AChE. In addition, compound
TM-3 was a favourable selective MAO-B inhibitor with IC50 value of 6.8 µM. More
interestingly, our study displayed that TM-3 was an antioxidant and neuroprotectant,
as well as a selective metal chelating agent. Further, TM-3 could cross the
blood-brain barrier (BBB) in vitro and complied with Lipinski’s rule of five of the
drug-likeness. Therefore, the present study revealed that TM-3 could be considered as
a promising multi-targets lead compound for the treatment of AD. Further
investigations to evaluate compound TM-3 in vivo and to develop structural
refinements are in progress and will be reported in due course.
Notes
The authors declare no competing financial interest.
Acknowledgments
This work was supported in part by the Scientific and Technological Project of Henan
Provincial (162300410103), the Key Scientific Research Project of Colleges and
Universities in Henan Province (17B350002), the Special Project of Nanyang Normal
University (ZX2016017).
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17

Design,
synthesis
and
biological
evaluation
of
2-acetyl-5-O-(amino-alkyl)phenol derivatives as multifunctional
agents for the treatment of Alzheimer’s disease
Zhipei Sanga, *, Keren Wangb, Huifang Wangc, Huijuan Wanga, Qianwen Maa, Xue
Hana, Mengyao, Yea, Lintao Yua, Wenmin Liua, *.
aCollege of Chemistry and Pharmaceutical Engineering, Nanyang Normal University,
Nanyang, 473061, China
bNanyang Normal University Hospital, Nanyang Normal University, Nanyang,
473061, China
cNanyang Normal University Library, Nanyang Normal University, Nanyang, 473061,
China
18