Multifunctional indanone–chalcone hybrid compounds with anti-β-amyloid (Aβ) aggregation, monoamine oxidase B (MAO-B) inhibition and neuroprotective properties against Alzheimer’s disease

Article abstract of DOI:10.1007/s00044-019-02423-4

To discover multifunctional agents for the treatment of Alzheimer’s disease (AD), a series of indanone–chalcone hybrid compounds were designed and synthesized based on the multitarget-directed ligand strategy. Their monoamino oxidases (MAO-A and MAO-B) an

Full text of DOI:10.1007/s00044-019-02423-4

MEDICINAL
CHEMISTRY
RESEARCH
Medicinal Chemistry Research
https://doi.org/10.1007/s00044-019-02423-4
ORIGINAL RESEARCH
Multifunctional indanone–chalcone hybrid compounds with anti-β-
amyloid (Aβ) aggregation, monoamine oxidase B (MAO-B) inhibition
and neuroprotective properties against Alzheimer’s disease
Keren Wang1 Lintao Yu Jian Shi Wenmin Liu Zhipei Sang1
1 ●
1 ●
1 ●
●
Received: 11 April 2019 / Accepted: 12 August 2019
©
Springer Science+Business Media, LLC, part of Springer Nature 2019
Abstract
To discover multifunctional agents for the treatment of Alzheimer’s disease (AD), a series of indanone–chalcone hybrid
compounds were designed and synthesized based on the multitarget-directed ligand strategy. Their monoamino oxidases
(
MAO-A and MAO-B) and Aβ1–42 aggregation inhibitory activities were evaluated. The results were shown that all synthetic
compounds exhibited mostly good multifunctional activities. Among all, compound TM-11 represented the best Aβ1–42
aggregation inhibitory potency (IC ~1.8 μM) and good disaggregation activity (IC ~7.9 μM). Both TEM images and
50
50
docking studies provided good reasonable explanation to the hypothesis. Meanwhile, compound TM-11 was a selective
MAO-B inhibitor, as well as a neuroprotective agent against Aβ1–42-induced toxicity. Based on the structural considerations,
the lipophilicity of the compound TM-11 could render to pass through blood−brain barrier (BBB) in vitro in accordance
with the Lipinski’s rule of five. In conclusion, these results were suggested that compound TM-11 might be a potential
multifunctional agent for the treatment of AD.
●
●
●
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Keywords Alzheimer’s disease Indanone–chalcone hybrid molecules MAO-B inhibition Aβ1–42 aggregation inhibition
Neuroprotective effects
Abbreviations
MTT
3-(4,5-dimethylthiazol-2-yl)-2,5-
diphenyltetrazolium
AD
Alzheimer’s disease
AChEIs
ACh
Aβ
acetylcholinesterase inhibitors
acetylcholine
β-amyloid
PAMPA- parallel artificial membrane assay for the
BBB
blood–brain barrier
TPSA
ADME
topological polar surface area
absorption, distribution, metabolism, excretion.
MAO-B monoamine oxidase B
MAO-A monoamine oxidase A
MTDLs
ThT
TEM
PDB
multitarget-directed ligands
thioflavin T
transmission electron microscopy
protein data bank
Introduction
Alzheimer’s disease (AD) is an age-related incurable neu-
rodegenerative disease characterized by deterioration of
memory, language skills, and other cognitive impairments.
It is reported that the total estimated worldwide cost of
dementia is US$818 billion in 2015, this number will
increase to 2 trillion dollar by 2030, and its prevalence will
increase to 131 million by 2050 (Alzheimer’s Disease
International 2015). The current drugs mainly focused on
acetylcholinesterase inhibitors (AChEIs), such as donepezil,
rivastigmine, and galantamine. The clinical use shows that
these drugs aim to the treatment of symptoms, but they do
not halt or reverse the progress of this disease (Ferreira-
Vieira et al. 2016).
These authors contributed equally: Keren Wang, Lintao Yu
Supplementary information The online version of this article (https://
doi.org/10.1007/s00044-019-02423-4) contains supplementary
material, which is available to authorized users.
*
Zhipei Sang
sangzhipei@126.com
1
College of Chemistry and Pharmaceutical Engineering, Nanyang
Normal University, Nanyang 473061, China

Medicinal Chemistry Research
Although the pathogenesis of AD is not clear, the main
pathological features, such as low levels of acetylcholine
radical-scavenging, antitumor, anti-inflammatory, and neu-
roprotective properties (Zhuang et al. 2017). Indanone is
one of the privileged structures in medicinal chemistry and
it is commonly associated with various pharmacologically
active compounds (Patil et al. 2017). Indanones and their
derivatives have been studied to determine their biological
activities within disease states, including AD (Li et al. 2016;
Caliandro et al. 2018; Costanzo et al. 2016; Huang et al.
2012). Based on the MTDLs strategy, we report the design,
synthesis, and evaluation of a series of indanone–chalcone
hybrid compounds (Fig. 1) that are found to indicate
potentially applicable biological activities, including the
inhibition of self-induced Aβ aggregation, inhibition of
MAO-B activity, and neuroprotective properties.
(
ACh), deposits of amyloid-β (Aβ) and τ protein have been
considered to be an important factor in the pathogenesis of
AD (Lane et al. 2018). Particularly, accumulation of
amyloid-beta (Aβ) in the brain is a central event in the
pathogenesis of AD base on the “amyloid hypothesis”, Aβ
could initiate the pathogenic cascade, generates reactive
oxygen species and tau phosphorylation, which ultimately
lead to neuronal loss and dementia (Ricciarelli and Fedele
2
017; Sensi et al. 2018). Therefore, inhibiting Aβ1–42
aggregation would provide a potent therapeutic strategy for
AD treatment.
Monoamine oxidase (MAO) is an important FAD-
dependent enzyme (flavoenzymes) responsible for the
metabolism of neurotransmitters such as dopamine, ser-
otonin, adrenaline, and noradrenaline (Kumar et al. 2017).
MAO exists in two types, MAO-A and MAO-B. MAO-B
expression significantly increases in the brain of AD
patients, and selective MAO-B inhibitor Selegiline could
improve learning and memory deficits in animal models
associated with AD (Borroni et al. 2017). Thus, selective
MAO-B inhibitors seem to be an important
treatment of AD.
Material and methods
Experimental section
All chemical agents were obtained from commercial sup-
1
pliers and used without further purification. H NMR and
1
3
C NMR spectra were recorded on a Varian INOVA
spectrometer, using tetramethyl silane as an internal stan-
Due to the complexity of AD, the multitarget-directed
ligands (MTDLs), which possess two or more com-
plementary AD-related targets, have been considered as an
effective way for the treatment of AD. The obtained results
encourage many groups to develop the advanced multi-
function agents (Cavalli et al. 2008; de Freitas Silva et al.
dard in CDCl at 400 and 100 MHz, respectively. Coupling
3
constants were given in Hz. Multiplicities were given as s
(singlet), d (doublet), dd (double-doublet), t (triplet), q
(quadruplet), m (multiplet), and br (broad signal). MS
spectra data were obtained on an Agilent-6210 TOF LC-MS
spectrometer. High-performance liquid chromatography
(HPLC) analysis was carried out on a Waters600-2487 plus
system with the use of a Kromasil C-18 column (4.6 mm ×
250 mm, 5 µm). The purity of target compounds were
confirmed more than 97% by HPLC analysis.
2
018; Umar and Hoda 2017).
Chalcones (α-phenyl-β-benzoylethylene) and their deri-
vatives are important bioactive molecules that have been
studied to exhibit impressive biological activities such as
Fig. 1 Design strategy for the
indanone–chalcone hybrid
compounds

Medicinal Chemistry Research
Recombinant human MAO-A and MAO-B (5 mg/mL)
were obtained from Sigma–Aldrich. Thioflavin T (ThT) was
purchased from TCI (Shanghai) Development. 1,1,1,3,3,3-
hexafluoro-2-propanol (HFIP) was purchased from Energy
Chemical. Amyloid-β1–42 (Aβ1–42), supplied as tri-
fluoroacetate salt, was purchased from China Peptides Co.,
Ltd. Docking studies were performed using the AUTO-
DOCK 4.2.6 program.
(400 MHz, DMSO-d ): δ = 7.57 (d, J = 8.8 Hz, 2H, 2 ×
6
Ar–H), 7.54 (s, 1H, C=CH), 7.34 (s, 1H, Ar–H), 6.96 (s, 1H,
Ar–CH), 6.72 (d, 2H, J = 7.6 Hz, 2 × Ar–H), 3.99 (s, 3H,
OCH ), 3.95 (s, 3H, OCH ), 3.90 (s, 2H, phCH ), 3.04 (s, 6H,
3
3
2
1
3
2 × NCH3). C NMR (100 MHz, DMSO-d ), 192.3 (C-13),
6
154.8 (C-3), 151.0 (C-19), 149.4 (C-2), 144.4 (C-6), 133.4
(C-5), 132.5 (C-17, 21), 131.7 (C-15), 130.6 (C-12), 123.4
(C-16), 111.9 (C-18, 20), 107.2 (C-4), 105.0 (C-1), 56.2, 56.1
−1
(
C-9, 10), 40.1 (C-23, 24), 32.4 (C-11). IR(cm ): 3100–2900
Chemistry
(–CH , –CH), 1675 (C=O), 1365 (–CH ), 1253, 1002
3
3
+
(C–O–C), 1303 (=N–). MS (ESI) m/z 324.2 [M + H] .
General procedure for the preparation of
indanone–chalcone hybrid compounds TM-1–TM-12
To a mixture of 5,6-dimethoxy-1-indanone (1 mmol) in
EtOH (3 mL) was added the appropriate benzaldehyde
(
1 mmol). An amount of a 30% aqueous KOH (4 mmol)
solution was then slowly added dropwise to the reaction.
The mixture was stirred for 72 h at 50 °C. After cooling to
room temperature, 10% HCl was added dropwise to the
mixture making pH = 2, overnight, a yellow precipitate
formed and filtrated, the residue was recrystallized with
8
0% ethanol to give yellow solid TM-1–TM-12.
(
Z)-2-(4-Fluorobenzylidene)-5,6-dimethoxy-2,3-dihydro-1H-
(
Z)-5,6-Dimethoxy-2-(4-methylbenzylidene)-2,3-dihydro-1H-
inden-1-one (TM-3) Yellow solid, m.p.: 116.9–118.3 °C,
8
DMSO-d ): δ = 7.89 (d, J = 7.6 Hz, 1H, Ar–H), 7.77 (d, J
=
7
1
(
DMSO-d ), 191.9 (C-13), 156.1 (C-19), 149.9 (C-2), 146.0
(
1
1
3
inden-1-one (TM-1) Light yellow solid, m.p.: 98.6–99.9 °
C, 86.7% yield, 98.9% HPLC purity. H NMR (400 MHz,
DMSO-d ): δ = 7.64 (d, J = 8.0 Hz, 2H, 2 × Ar–H), 7.40 (s,
1
5.8% yield, 98.7% HPLC purity. H NMR (400 MHz,
1
6
6
8.0 Hz, 1H, Ar–H), 7.65 (s, 1H, C=CH), 7.52 (t, J =
.6 Hz, 1H, Ar–H), 7.36 (t, J = 7.6 Hz, 1H, Ar–H), 7.24 (s,
1
2
3
(
(
1
2
1
H, C=CH), 7.31 (d, J = 8.0 Hz, 2H, 2 × Ar–H), 7.22 (s,
H, 2 × Ar–H), 3.97 (s, 2H, phCH ), 3.90 (s, 3H, OCH ),
2
3
H, Ar–H), 7.19 (s, 1H, Ar–H), 3.96 (s, 2H, phCH ), 3.90
2
13
.84 (s, 3H, OCH ), 2.36 (s, 3H, CH ). C NMR
13
3
3
s, 3H, OCH ), 3.85 (s, 3H, OCH ). C NMR (100 MHz,
3
3
100 MHz, DMSO-d ), 193.3 (C-13), 155.7 (C-2), 149.8
6
6
C-1), 145.5 (C-5), 139.9 (C-4), 135.4 (C-15), 132.8 (C-19),
C-1), 138.9 (C-5), 134.9 (C-4), 133.7 (C-15), 131.4 (C-12),
30.9 (C-16), 130.3 (C-17), 129.3 (C-21), 128.6 (C-18),
31.5 (C-12), 131.0 (C-18, 20), 130.5 (C-16), 130.1 (C-17,
1), 108.5 (C-3), 105.0 (C-6), 56.5, 56.1 (C-8, 10), 32.1 (C-
26.1 (C-20), 108.5 (C-3), 105.2 (C-6), 56.5, 56.2 (C-10),
−
1
1), 21.5 (C-22). IR(cm ): 3100–2900 (–CH , –CH), 1683
−1
3
1.3 (C-11). IR (cm ): 3070–2832 (–CH , –CH), 1679
3
(
2
C=O), 1363 (–CH ), 1253, 1087 (C–O–C). MS (ESI) m/z
3
(
C=O), 1365(–CH ), 1253, 1006 (C–O–C), 1222 (C–F).
3
+
95.1 [M + H] .
+
MS (ESI) m/z 299.1 [M + H] .
(
Z)-2-(4-(Dimethylamino)benzylidene)-5,6-dimethoxy-2,3-
(Z)-5,6-Dimethoxy-2-(2-methylbenzylidene)-2,3-dihydro-1H-
inden-1-one (TM-4) Light yellow solid, m.p.: 99.6–101.2 °C,
63.6% yield, 99.0% HPLC purity. H NMR (400 MHz,
dihydro-1H-inden-1-one (TM-2) Light yellow oil, m.p.:
1
1
1
17.8–119.1 °C, 83.1% yield, 98.6% HPLC purity. H NMR

Medicinal Chemistry Research
DMSO-d ): δ = 7.70–7.69 (m, 1H, Ar-H), 7.62 (s, 1H,
Ar–H), 7.31–7.30 (m, 3H, 3 × Ar–H), 7.20 (s, 1H, Ar–H),
21), 131.5 (C-12), 130.7 (C-16), 128.2 (C-3), 115.0 (C-
18, 20), 108.6 (C-8), 56.4, 56.1 (C-8, 10), 55.8 (C-22),
6
32.0 (C-11). IR (cm− ): 3100–2830 (–CH , –CH), 1687
1
7
3
1
.16 (s, 1H, Ar–H), 3.89 (s, 5H, OCH + phCH ), 2.41 (s,
3
2
3
1
3
H, CH3). C NMR (100 MHz, DMSO-d ), 192.1(C-9),
(C=O), 1361 (–CH ), 1259, 1020 (C–O–C). MS (ESI)
6
3
+
55.8 (C-3), 149.8 (C-2), 145.8 (C-6), 138.8 (C-5), 137.0
m/z 311.1 [M + H] .
(
C-15), 134.2 (C-16), 131.1 (C-21), 130.5 (C-8), 129.6 (C-
1
1
9), 129.0 (C-20), 128.7 (C-17), 126.7 (C-18), 108.5 (C-4),
05.0 (C-1), 56.4, 56.1 (C-13, 14), 31.6 (C-7), 20.1 (C-22).
−
1
IR (cm ): 3058–2833 (–CH , –CH), 1683 (C=O), 1357
3
+
(–CH ), 1255, 1000 (C–O–C). MS (ESI) m/z 295.1 [M + H] .
3
(
Z)-2-([1,1′-Biphenyl]-4-ylmethylene)-5,6-dimethoxy-2,3-
dihydro-1H-inden-1-one (TM-7) Light yellow solid, m.p.:
12.5–113.8 °C, 93.8% yield, 98.9% HPLC purity. 1
NMR (400 MHz, DMSO-d ): δ = 7.85 (d, J = 8.4 Hz, 2H,
1
H
6
2
8
× Ar–H), 7.81 (d, J = 8.4 Hz, 2H, 2 × Ar–H), 7.76 (d, J =
.0 Hz, 2H, 2 × Ar–H), 7.52–7.48 (m, 3H, 3 × Ar–H), 7.41
(
Z)-2-(Furan-2-ylmethylene)-5,6-dimethoxy-2,3-dihydro-1H-
inden-1-one (TM-5) Light yellow solid, m.p.:
00.2–101.5 °C, 62.8% yield, 98.1% HPLC purity.
NMR (400 MHz, DMSO-d ): δ = 7.95 (s, 1H, Ar–H), 7.24
1
(t, J = 7.6 Hz, 1H, Ar–H), 7.24 (s, 2H, 2 × Ar–H), 4.06 (s,
1
H
13
2H, phCH ), 3.92 (s, 3H, OCH ), 3.85 (s, 3H, OCH ).
C
2
3
3
6
NMR (100 MHz, DMSO-d ), 192.3 (C-12), 155.8 (C-3),
6
(
1
s, 1H, Ar–H), 7.19 (d, J = 6.8 Hz, 2H, 2 × Ar–H), 6.99 (s,
H, Ar–H), 6.72 (s, 1H, Ar–H), 3.90 (s, 5H, OCH +
1
49.9 (C-2), 145.6 (C-6), 141.4 (C-18), 139.7 (C-5), 136.5
(C-21), 134.7 (C-14), 131.6 (C-23, 25), 131.1 (C-15), 130.5
C-11), 129.5 (C-22, 26), 128.4 (C-24), 127.6 (C-17, 19),
3
13
phCH ), 3.83 (s, 3H, OCH ). C NMR (100 MHz, DMSO-
2
3
(
d6), 191.7 (C-13), 155.7 (C-2), 152.0 (C-16), 149.8 (C-1),
46.6 (C-18), 145.2 (C-5), 133.6 (C-4), 130.9 (C-12), 118.2
C-15), 116.9 (C-3), 113.4 (C-19), 108.6 (C-20), 105.0 (C-
1
1
27.2 (C-16, 20), 108.6 (C-4), 105.1 (C-1), 56.5, 56.2 (C-9,
0), 32.2 (C-27). IR (cm ): 3100–2830 (–CH , –CH), 1679
1
(
6
(
(
−
1
3
−
1
(C=O), 1303 (–CH3), 1255, 1004 (C–O–C). MS (ESI) m/z
), 56.4, 56.1 (C-8, 10), 32.0 (C-11). IR (cm ): 3100–2834
+
357.1 [M + H] .
–CH , –CH), 1633 (C=O), 1359 (–CH ), 1249, 1006
3
3
+
C–O–C). MS (ESI) m/z 271.1 [M + H] .
(
Z)-2-(4-Bromobenzylidene)-5,6-dimethoxy-2,3-dihydro-1H-
inden-1-one (TM-8) Light yellow solid, m.p.:
104.6–105.8 °C, 60.8% yield, 98.8% HPLC purity. H
(
1
1
Z)-5,6-Dimethoxy-2-(4-methoxybenzylidene)-2,3-dihydro-
1
H-inden-1-one (TM-6) Light yellow solid, m.p.:
1
04.1–105.3 °C, 80.5% yield, 98.6% HPLC purity. H
NMR (400 MHz, DMSO-d ): δ = 7.70 (s, 4H, 4 × Ar–H),
6
NMR (400 MHz, DMSO-d ): δ = 7.70 (d, J = 8.4 Hz, 2H,
7.40 (s, 1H, Ar–H), 7.22 (d, J = 6.8 Hz, 2H, 2 × Ar–H),
6
2
7
3
× Ar–H), 7.40 (s, 1H, C=CH), 7.21 (s, 2H, 2 × Ar–H),
3.99 (s, 2H, phCH ), 3.91 (s, 3H, OCH ), 3.84 (s, 3H,
2
3
1
3
.05 (d, J = 8.4 Hz, 2H, 2 × Ar–H), 3.95 (s, 2H, phCH ),
OCH3). C NMR (100 MHz, DMSO-d ), 192.2 (C-13),
2
6
1
3
.90 (s, 3H, OCH ), 3.83 (s, 6H, 2 × OCH ). C NMR
155.9 (C-2), 149.9 (C-1), 145.6 (C-5), 137.2 (C-4), 134.9
(C-15), 132.8 (C-18, 20), 132.4 (C-17, 21), 130.4 (C-16),
130.2 (C-19), 123.4 (C-3), 108.6 (C-6), 56.5 56.2 (C-8, 10),
3
3
(
(
100 MHz, DMSO-d ), 192.3 (C-13), 160.8 (C-19), 155.6
C-2), 149.8 (C-1), 145.3 (C-5), 133.9 (C-4), 132.8 (C-17,
6

Medicinal Chemistry Research
−
1
3
(
2.0 (C-11). IR (cm ): 3100–2800 (–CH , –CH), 1693
1313 (–CH ), 1257, 1006 (C–O–C). MS (ESI) m/z 297.1
[M + H] .
3
3
+
C=O), 1309 (–CH ), 1226 (C–Br), 1257, 1004 (C–O–C).
MS (ESI) m/z 359.0 [M + H] .
3
+
(
1
1
Z)-2-(2-Hydroxybenzylidene)-5,6-dimethoxy-2,3-dihydro-
(
Z)-5,6-Dimethoxy-2-(4-nitrobenzylidene)-2,3-dihydro-1H-
H-inden-1-one (TM-9) Light yellow solid, m.p.:
inden-1-one (TM-11) Light yellow solid, m.p.: 98.9–100.2 °C,
1
37.7–139.2 °C, 53.7% yield, 98.9% HPLC purity.
H
1
8
1.3% yield, 98.9% HPLC purity. H NMR (400 MHz, DMSO-
d6): δ = 8.30 (d, J = 8.4 Hz, 2H, 2 × Ar–H), 7.99 (d, J = 8.8 Hz,
H, 2 × Ar–H), 7.50 (s, 1H, C=CH), 7.23 (s, 2H, 2 × Ar–H),
.05 (s, 2H, phCH ), 3.92 (s, 3H, OCH ), 3.84 (s, 3H, OCH ).
NMR (400 MHz, DMSO-d ): δ = 10.13 (s, 1H, OH), 7.83
(
(
2
NMR (100 MHz, DMSO-d ), 192.4 (C-13), 157.9 (C-21),
1
(
1
5
3
1
6
s, 1H, Ar–H), 7.68 (d, J = 7.6 Hz, 1H, Ar–H), 7.27–7.20
m, 3H, 3 × Ar–H), 6.96–6.89 (m, 2H, 2 × Ar–H), 3.95 (s,
2
4
2
3
3
1
3
H, phCH ), 3.90 (s, 3H, OCH ), 3.84 (s, 3H, OCH ).
C
13
2
3
3
C NMR (100 MHz, DMSO-d ), 192.0 (C-13), 156.2 (C-2),
6
6
1
4
2
49.9 (C-1), 147.6 (C-19), 146.0 (C-5), 142.1 (C-16), 140.2 (C-
), 131.8 (C-17, 21), 131.1 (C-15), 128.9 (C-12), 124.3 (C-18,
55.6 (C-2), 149.8 (C-1), 145.5 (C-5), 134.8 (C-15), 131.4
C-4), 130.7 (C-12), 129.8 (C-19), 126.3 (C-17), 122.5 (C-
0), 108.5 (C-3), 105.1 (C-6), 56.5, 56.2 (C-8, 10), 31.9 (C-11).
8), 119.8 (C-20), 116.4 (C-16), 108.6 (C-3), 105.1 (C-6),
−1
IR (cm ): 3100–2850 (–CH , –CH), 1683 (C=O), 1342
(
3
3
−
1
6.4 56.1 (C-8, 10), 32.0 (C-11). IR (cm ): 3130 (–OH),
083–2834 (–CH , –CH), 1656 (C=O), 1313 (–CH ), 1259,
–CH ), 1259 1002 (C–O–C), 1631, 850 (–NO ). MS (ESI) m/z
3
2
3
3
+
26.1 [M + H] .
+
010 (C–O–C). MS (ESI) m/z 297.1 [M + H] .
(
Z)-2-(4-Hydroxybenzylidene)-5,6-dimethoxy-2,3-dihydro-
1
1
H-inden-1-one (TM-10) Light yellow solid, m.p.:
40.8–141.9 °C, 46.4% yield, 98.6% HPLC purity. H
1
(
Z)-2-(3-hydroxybenzylidene)-5,6-dimethoxy-2,3-dihydro-
NMR (400 MHz, DMSO-d ): δ = 10.03 (s, 1H, OH),
6
1H-inden-1-one (TM-12) Light yellow solid, m.p.:
1
NMR (400 MHz, CDCl ): δ = 7.54 (s, 1H, Ar–H), 7.35 (s,
1H, Ar–H), 7.32 (d, J = 8.0 Hz, 1H, Ar–H), 7.25 (d, J =
8.0 Hz, 1H, Ar–H), 7.14 (s, 1H, Ar–H), 6.98 (s, 1H,
Ar–H), 6.87 (d, J = 8.4 Hz, 1H, Ar-H), 4.00 (s, 3H,
OCH ), 3.97 (s, 2H, phCH ), 3.96 (s, 3H, OCH ).
IR (cm ): 3149 (–OH), 3000–2800 (–CH , –CH), 1664
(
1
7
7
2
3
1
1
1
1
3
.60 (d, J = 8.4 Hz, 2H, 2 × Ar–H), 7.36 (s, 1H, C=CH),
.20 (d, J = 8.4 Hz, 2H, 2 × Ar–H), 6.89 (d, J = 8.0 Hz,
H, 2 × Ar–H), 3.93 (s, 2H, phCH ), 3.90 (s, 3H, OCH ),
01.1–102.4 °C, 85.4% yield, 98.4% HPLC purity. H
3
2
3
.
8
3
(
s
,
3
H
,
O
C
H
)
.
1
3
C
N
M
R
(
1
0
0
M
H
z
,
D
M
S
O
-
d
)
,
3
6
92.4 (C-13), 159.5 (C-19), 155.5 (C-2), 149.7 (C-1),
45.2 (C-5), 133.0 (C-4), 132.9 (C-15), 132.0 (C-12),
30.8 (C-17, 21), 126.7 (C-16), 116.4 (C-18, 20), 108.9,
05.0 (C-6), 56.4 56.1 (C-8, 10), 32.1 (C-11). IR (cm− ):
3
2
3
−
1
3
1
C=O), 1311 (–CH ), 1255, 1004 (C–O–C). MS (ESI) m/
3
210 (-OH), 3000–2800 (–CH , –CH), 1668 (C=O),
+
3
z 297.1 [M + H] .

Medicinal Chemistry Research
Biology
In vitro inhibition of monoamine oxidase
Recombinant human MAO-A and MAO-B (5 mg/mL) were
obtained from Sigma–Aldrich, prealiquoted and stored at
−
70 °C according to the procedure referenced to pre-
liminary work (Li et al. 2017).
Effect of test compounds on self-induced Aβ1–42
aggregation and disaggregation experiments by the ThT
assay
Scheme 1 Synthesis of target compounds TM-1 ~ TM-12. Reagents
and conditions: (i) 50%KOH, EtOH, room temperature, 3 ~ 4 days
In order to investigate the self-induced Aβ1–42 aggregation,
a ThT-based fluorometric assay was performed (Sang et al.
procedure could be found in our previous work (Sang et al.
2019).
2
015). The details could reference our previous work (Sang
et al. 2015).
Transmission electron microscopy (TEM) assay
Results and discussion
Briefly, Aβ1–42 was incubated within the presence and
absence of compound TM-11 at 37 °C for 48 h. The final
concentration of Aβ1–42 and test compound was 25 μM,
respectively. Aliquots (50 μL) of each sample were placed
on a carbon coated copper/rhodium grid for 2 min at room
temperature. Each grid staining was performed with uranyl
acetate (1%) for 2 min. the excess staining solution was
discarded can the specimen was transferred for imaging
with field emission transmission electron microscope (JEM-
Chemistry
The synthetic route of the indanone–chalcone hybrid com-
pounds was shown in Scheme 1. The target compounds
were prepared by condensation of the starting material 5,6-
dimethoxy-1-indanone (1) with the appropriate aldehydes
(a ~ l) in ethanolic KOH solution. All new compounds were
purified by recrystallization, and the analytical and spec-
troscopic data confirmed their structures, as detailed in the
experimental section.
2
100F) (Sang et al. 2019).
Molecular modeling studies
Pharmacology
The crystal structure of Aβ (PDB ID: 1BA4) were obtained
from the Protein Data Bank after removing the original
inhibitors and water molecules. The 3D Structure of TM-11
was built and performed geometry optimization by mole-
cular mechanics. Docking studies were performed using the
AUTODOCK 4.2.6 program and each docked system was
performed by 200 runs of the AUTODOCK search by the
Lamarckian genetic algorithm. A cluster analysis was per-
formed on the docking results using a root mean square
tolerance of 1.0 and the lowest energy conformation of the
highest populated cluster was selected for analysis. Graphic
manipulations and visualizations were done by Autodock
Tools or Discovery Studio 2.5 software (Sang et al. 2019).
In vitro inhibition of monoamine oxidase
The inhibitory activity against MAO-A and MAO-B
(recombinant human enzyme) was determined, and clor-
gyline, rasagiline, and iproniazid were also tested as refer-
ence compounds (Li et al. 2017). As shown in Table 1, all
the indanone–chalcone hybrid compounds were effective in
inhibiting MAO-B in the submicromolar range or lower,
and all the target compounds showed weak MAO-A inhi-
bitory activities. Therefore, all the synthesized compounds
were efficient selective MAO-B inhibitors. The structure-
activity relationship in Table 1 indicated that the substituent
groups served as important role. The compounds (TM-1,
TM-2, TM-4, and TM-6) possessing electron-donating
group displayed higher inhibitory activities than compounds
In vitro blood−brain barrier permeation assay
(
TM-3, TM-4, TM-5, TM-7, TM-9, and TM-12) with
The blood−brain barrier penetration of compounds was
evaluated using the parallel artificial membrane permeation
assay (PAMPA) described by Di et al. 2003. The detailed
electron-withdrawing group, except compounds TM-8 and
TM-11 having good inhibitory potency with IC50 values of
0.059 and 0.031 μM. Meanwhile, the position of substituent

Medicinal Chemistry Research
Table 1 Inhibition and disaggregation of Aβ aggregation and MAOs (MAO-A and MAO-B) inhibitory potency of target compounds and reference
compounds
Structures (Ar substituent group) are shown in Fig. 1. Compounds defined “no active” means percent inhibition less than 5.0% at a concentration of
1
.0 μM
n.t. not tested, n.a. no active
aInhibition of Aβ aggregation, the thioflavin T fluorescence method was used
bIC50 values represent the concentration of inhibitor required to decrease enzyme activity by 50% and are the mean of three independent
experiments, each performed in triplicate (SD = standard deviation)
cFrom recombinant human MAO-A
dFrom recombinant human MAO-B
group also affected the inhibitory activity, generally, the
inhibitory activities were in order: 4-position > 2-position >
4. When the substituent group was furan group, the com-
pound (TM-5) exhibited moderate inhibitory activity with
IC50 value of 0.62 μM. Among these compounds,
3
-positon, such as TM-10 > TM-9 > TM-12; TM-1 > TM-

Medicinal Chemistry Research
compound TM-2 was the best selective MAO-B inhibitor
with IC50 value of 0.022 μM.
Effect of test compounds on self-induced Aβ1–42
aggregation and disaggregation experiments by ThT assay
In order to assess the effects of the target compounds
against self-induced Aβ1−42 aggregation, we carried out two
individual studies: inhibition and disaggregation experi-
ments of self-induced Aβ1−42 aggregation using the ThT
fluorescence assay (Sang et al. 2015). Curcumin, a known
active natural product for inhibition of self-induced Aβ
aggregation, was used as reference compound. The results
of inhibitory activity of Aβ1–42 aggregation were summar-
ized in Table 1, almost all the target compounds exhibited
more potent inhibition potency for self-mediated Aβ1−42
aggregation than curcumin (IC = 26.7 μM). According to
the data, the properties of the substituent group had sig-
nificant effects on Aβ aggregation inhibitory activities,
generally, the electron-drawing group (TM-3, TM-8, and
TM-11) in benzene ring indicated better inhibitory activities
than that with electron-donating group (TM-1, TM-2, TM-
Fig. 2 Effects of TM-3, TM-4, TM-5, TM-6, TM-8, TM-9, and TM-
1
1 on cell viability in SH-SY5Y cells. Data are mean values SEM of
three independent experiments
(MTT) assays (Jiang et al. 2019). As shown in Fig. 2,
TM-4, TM-5, TM-6, and TM-8 induced a decrease of cell
viability (66.1%, 65.2%, 50.6%, and 70.5%, respectively),
while TM-3, TM-9, and TM-11 did not show modified
cell viability. The results showed that TM-3, TM-9, and
TM-11 had a wide therapeutic safety range.
50
6
, TM-7, and TM-10). The position of substituent group of
Transmission electron microscopy
benzene ring also affected the inhibitory potency, when the
substituent group were –OH and –CH , the inhibitory
According to the results of self-induced Aβ1−42 aggregation,
compounds TM-4, TM-8, and TM-11 showed similar
inhibitory activities, but compound TM-4 and TM-8 dis-
played significant cytotoxicity. Therefore, compound TM-
11 should be encouraged to further studies.
To complement the ThT binding assay, the inhibition
effect of TM-4 on self-induced Aβ1–42 aggregation was also
monitored by TEM (Fig. 3) (Sang et al. 2019). The results
showed that the sample of Aβ1–42 alone had aggregated into
amyloid fibrils after 24 h of incubation, while only small
bulk aggregates were visible and no characteristic fibrils
were observed in the sample of Aβ1–42 in the presence of
compound TM-11. The TEM results were well consistent
with the results of ThT measurements, which strongly
proved that compound TM-11 could inhibit Aβ1–42 fibrils
formation.
3
activities were in the order, 3-position > 2-position > 4-
position, such as TM-12 > TM-9 > TM-10; TM-6 > TM-1.
Replacing the benzene ring substituent group of TM-4 with
furan group to obtain compound TM-5, dramatically, the
inhibitory activity dropped down to IC50 value of 20.3 μM.
In short, compounds TM-4, TM-8, and TM-11 showed the
potent inhibitory activities with IC50 value of 3.8, 4.6, and
1
.8 μM.
Compounds TM-4, TM-8, TM-11, and TM-12 were
selected to test the disaggregation effects on self-induced
Aβ1−42 aggregation using the ThT binding assay. According
to the data in Table 1, compounds TM-4, TM-8, TM-11,
and TM-12 could significantly disaggregate Aβ fibrils with
IC50 value of 5.8, 11.5, 7.9, and 7.2 μM, respectively. Form
the above results, revealing that indanone–chalcone hybrid
compounds could inhibit and disaggregate self-induced Aβ1−42
aggregation.
Molecular modeling studies
Cytotoxicity
To further explore the binding modes of the active com-
pound TM-11 with Aβ1–42, a molecular docking experi-
ment was performed (Sang et al. 2019). The structure of
Aβ used was downloaded from the Protein Data Bank
(PDB: 1BA4). As shown in Fig. 4, compound TM-11 was
locked at the C-termius hydrophobic area of Aβ. The
oxygen atom of the methoxy group interacted with ASP1
through one intermolecular hydrogen bond, the −NO2
group hydroxyl could bind to LYS28 via two inter-
molecular hydrogen bond. In addition, compound TM-11
In order to evaluate the further biological activity of
indanone–chalcone hybrid compounds, compounds TM-
3
, TM-4, TM-5, TM-6, TM-8, TM-9, and TM-11, with
better inhibitory activity of self-induced Aβ1−42 aggrega-
tion, was selected to examine the cytotoxicity. SH-SY5Y
cells were exposed to the test compounds at 1 μM con-
centrations for 24 h, and the cell viabilities were tested by
the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium

Medicinal Chemistry Research
Aβ (0h)
Aβ alone (24h)
Aβ + TM-11 (24h)
Fig. 3 TEM images of Aβ1−42 (25 μM) in the presence and absence of 25 μM compound TM-11 after 24 h of aggregation
Fig. 5 Neuroprotection against Aβ1–42-induced toxicity, compound
TM-11 was tested for neuroprotective activity against Aβ toxicity in
SH-SY5Y neuroblastoma cell cultures. Results are expressed as per-
cent viability compared with untreated control. All data were express
as mean ± SD of three experiments. ##p < 0.05 vs control; *p < 0.05,
*
*p < 0.01 vs Aβ1–42 alone
Table 2 Permeability Pe (×10− cm/s) in the PAMPA-BBB assay for
6
1
1 commercial drugs used in the experiment validation
Commercial drugs
Lit. (Di et al. 2003)
PBS:EtOH (70:30)a
Verapamil
Oxazepam
Diazepam
Clonidine
16
16.90
9.60
10
16
11.86
5.10
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
Norfloxacin
Theophylline
0.47
0.72
0.42
0.10
aData are the mean ± SD of three independent experiments
Fig. 4 Docking studies of compound TM-11 with Aβ1–42 (PDB ID:
BA4). a Cartoon representations of compound TM-11 interacting
with Aβ1–42. b Association of TM-11 (green stick) and the C-terminus
of obtained from docking calculations. c 2D schematic diagram of
docking model of compound TM-11 with Aβ1–42
formed the hydrophobic interactions with the residues
ASP1, PHE20, LYS28, ASN27, and ILE31. These dif-
ferent interactions might be favorable for the binding of
Aβ and compound TM-11.
1

Medicinal Chemistry Research
control group (Jiang et al. 2019). As shown in Fig. 5, after
25 μM Aβ1–42 exposure for 48 h, cell viability obviously
decreased to 48.5% (p < 0.01) compared with untreated
control. The cell viability increased to 72.7% after adding
5 μM curcumin. When treatment with compound TM-11 (1,
5, and 10 μM), the cell viability increased to 78.6, 86.8, and
89.5% in a dose-dependent manner, which showed better
neuroprotective effect against Aβ1–42-mediated neurotoxi-
city was than that with curcumin.
Fig. 6 Linear correlation between experimental and reported perme-
In vitro blood−brain barrier permeation assay
ability of commercial drugs using the PAMPA-BBB assay. Pe(exp) =
2
0
.9163, Pe(bibl.) −0.2247 (r = 0.9558)
The parallel artificial membrane permeation assay of the
blood−brain barrier (PAMPA-BBB) was used to evaluate
the possible blood−brain barrier (BBB) permeability of
TM-11 (Sang et al. 2019; Di et al. 2003). First, the assay
was validated through comparing the permeability of 11
Table 3 Ranges of permeability of PAMPA-BBB assays (Pe × 10−6
cm/s)
Compounds of high BBB permeation (CNS+)
Compounds of uncertain BBB permeation (CNS±) 3.44 > Pe > 1.61
Compounds of low BBB permeation (CNS−) Pe < 1.61
Pe > 3.44
commercial drugs in Table 2. A good linear correlation, Pe
2
(
exp) = 0.9163P (bibl.) −0.2247 (r = 0.9558), was pro-
e
duced in Fig. 6. And then because of the known limit by Di
et al., we considered that drug with permeability above
3
.44 × 10− cm/s could cross BBB in vitro (Table 3). So, as
6
Table 4 Permeability Pe (×10−6 cm/s) in the PAMPA-BBB assay of
the selected compound TM-11 and the predictive penetration in
the CNS
shown in Table 4, TM-11 could cross the BBB in vitro.
Theoretical prediction of the ADME properties
Compounda
Pe (×10−6 cm/s)b
Prediction
To evaluate the druglike property of compound TM-11, the
key parameters, such as log P, topological polar surface area
TM-11
12.36 ± 0.89
CNS+
aCompound TM-11 was dissolved in DMSO at 5 mg/mL and diluted
with PBS/EtOH (70:30). The final concentration of the compound was
(TPSA), the number of hydrogen-bond acceptors and
hydrogen-bond donors were calculated using the Molin-
spiration property program (Matos et al. 2013). The results
were shown in Table 5, compound TM-11 complied with the
Lipinski’s rule of five, deserving the further study in vivo.
1
00 μg/mL
bValues are expressed as the mean ± SD of three independent
experiments
Table 5 Theoretical prediction of the ADME properties of compound
TM-11
Conclusion
Comp.a Log P MW
TPSA (Å ) n-ON n-OHNH volume (Å )
2
3
In summary, a series of indanone–chalcone hybrid com-
pounds were designed, synthesized, and evaluated as mul-
tifunctional agents for the treatment of AD. All the
synthesized compounds showed remarkable MAO-B inhi-
bitory activities and significant self-induced Aβ1–42 aggre-
gation inhibition. Among these synthesized compounds,
compound TM-11 exhibited the best inhibitory potency of
Aβ1–42 aggregation and good disaggregation activity with
IC50 value of 1.8 and 7.9 μM, both the TEM images and
docking studies provided reasonable explain. Meanwhile,
compound TM-11 was a selective MAO-B inhibitor, as
well as a neuroprotective agent against Aβ1–42-induced
toxicity. Moreover, TM-11 could cross the BBB in vitro
and accorded with the Lipinski’s rule of five. Taken toge-
ther, these results suggested that compound TM-11 might
be a potential multifunctional agent for the treatment of AD.
TM-11 3.32 325.32 81.36
6
0
282.24
MW Molecular weight, TPSA topological polar surface area, n-ON
number of hydrogen acceptors, n-OHNH number of hydrogen
bond donors
aThe data was determined with the Molinspiration calculation software
Neuroprotective effect against Aβ1–42-induced
neurotoxicity
Aβ1–42-induced apoptotic neuronal cell death was also an
important factor in the pathology of AD. The representative
compound TM-11 was performed the further experiments
against Aβ1–42-induecd neurodegeneration in the SH-SY5Y
cells using MTT assay, curcumin was also evaluated as

Medicinal Chemistry Research
Acknowledgements This work was supported in part by the Foundation
and Frontier Projects of Nanyang Science and Technology Bureau
Huang L, Lu C, Sun Y, Mao F, Luo Z, Su T, Jiang H, Shan W, Li X
(2012) Multitarget-directed benzylideneindanone derivatives:
anti-β-amyloid (Aβ) aggregation, antioxidant, metal chelation,
and monoamine oxidase B (MAO-B) inhibition properties against
Alzheimer’s disease J Med Chem 55(19):8483–8492
(2017JCQY020 and 2017 JCQY021). The Key Scientific Research
Project of Colleges and Universities in Henan Province (20A350006).
International Training of High-level Talents in Henan Province in 2019
(
No. 17) (Henan Foreign Experts Bureau). The Special Project of
Jiang CS, Ge YX, Cheng ZQ, Wang YY, Tao HR, Zhu K, Zhang H
(2019) Discovery of new selective butyrylcholinesterase (BChE)
inhibitors with anti-Aβ aggregation activity: structure-based vir-
tual screening, hit optimization and biological evaluation. Mole-
cules 24(14):pii: E2568
Kumar B, Gupta VP, Kumar V (2017) A perspective on monoamine
oxidase enzyme as drug target: challenges and opportunities. Curr
Drug Targets 18(1):87–97
Nanyang Normal University (SYKF2018081 and 2019QN001). Gradu-
ate Innovation Fund Project of Nanyang Normal University (No. 02).
Compliance with ethical standards
Conflict of interest The authors declare that they have no conflict of
interest.
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Neurol 25(1):59–70
Publisher’s note: Springer Nature remains neutral with regard to
jurisdictional claims in published maps and institutional affiliations.
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and evaluation of 4-hydroxyl aurone derivatives as multi-
functional agents for the treatment of Alzheimer’s disease. Bioorg
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