Amide derivatives of Gallic acid: Design, synthesis and evaluation of inhibitory activities against in vitro α-synuclein aggregation

Article abstract of DOI:10.1016/j.bmc.2020.115596

Gallic acid (GA), a natural phenolic acid, has received numerous attention because of its anti-oxidative, anti-inflammatory, and anti-cancer activity. More importantly, GA can act as an efficient inhibitor of α-Synuclein (α-Syn) aggregation at early stages. Nevertheless, some evidences suggest that GA is unlikely to cross the blood–brain barrier because of its high hydrophilicity. Hence, GA may not be considered as a promising candidate or entering brain and directly affecting the central nervous system. Accordingly, we have designed and synthesized a series of amide derivatives of GA, some of which possess appropriate lipophilicity and hydrophilicity with LogP (2.09–2.79). Meanwhile, these sheet-like conjugated compounds have good π-electron delocalization and high ability of hydrogen-bond formation. Some compounds have shown better in vitro anti-aggregation activities than GA towards α-Syn, with IC₅₀ down to 0.98 μM. The valid modification strategy of GA is considered an efficient way to discover novel inhibitors of α-Syn aggregation.

Full text of DOI:10.1016/j.bmc.2020.115596

Bioorganic&MedicinalChemistry28(2020)115596
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Amide derivatives of Gallic acid: Design, synthesis and evaluation of
inhibitory activities against in vitro α-synuclein aggregation
⁎
⁎
⁎
Li Chena,d, Guo-Long Huanga,d, Ming-Huan Lüa, Yun-Xiao Zhanga, , Ji Xub, , Su-Ping Baic,
a College of Chemistry and Institute of Green Catalysis, Zhengzhou University, Daxue Road 75, 450052 Zhengzhou, China
b School of Basic Medical Science, Neuroscience Research Institute, Academy of Medical Sciences, Zhengzhou University, Kexue Road 100, 450001 Zhengzhou, China
c College of Pharmacy, Xinxiang Medical University, Jinsui Road 601, 453003 Xinxiang, China
A R T I C L E I N F O
A B S T R A C T
Keywords:
Gallic acid (GA), a natural phenolic acid, has received numerous attention because of its anti-oxidative, anti-
inflammatory, and anti-cancer activity. More importantly, GA can act as an efficient inhibitor of α-Synuclein (α-
Syn) aggregation at early stages. Nevertheless, some evidences suggest that GA is unlikely to cross the
blood–brain barrier because of its high hydrophilicity. Hence, GA may not be considered as a promising can-
didate or entering brain and directly affecting the central nervous system. Accordingly, we have designed and
synthesized a series of amide derivatives of GA, some of which possess appropriate lipophilicity and hydro-
philicity with LogP (2.09–2.79). Meanwhile, these sheet-like conjugated compounds have good π-electron de-
localization and high ability of hydrogen-bond formation. Some compounds have shown better in vitro anti-
aggregation activities than GA towards α-Syn, with IC50 down to 0.98 μM. The valid modification strategy of GA
is considered an efficient way to discover novel inhibitors of α-Syn aggregation.
Amide derivatives of Gallic acid
Synthesis
α-Synuclein
Anti-aggregation
Inhibitor
1. Introduction
anti-inflammation and anti-aggregation effects on α-Syn, amyloid-beta
(Aβ) and tau proteins.5b,6
Parkinson‘s disease (PD) is one of the most common neurodegen-
erative diseases. It is characterized primarily by the loss of dopami-
nergic neurons in the substantia nigra, which led to movement dis-
orders.1 Although the precise molecular pathogenesis of PD are
unknown, the misfolding and aggregation of the abundant neuronal
protein α-synuclein (α-Syn) are involved in all PD cases.2 A large
number of conclusive evidence had been found that α-Syn oligomers,
protofibrils and amyloid fibrils are neurotoxic, and have shown prion-
like pathology propagation.3,1b Hence, maintaining α-Syn proteostasis
is considered as a key approach to PD prophylaxis and treatment.
Up to date, a number of natural and synthesized small molecular
compounds have been reported as the optional inhibitor of α-Syn ag-
gregation.4 Particularly, polyphenols showed good inhibitory activ-
ities.5 There are two sub-categories of polyphenol, flavonoids and
phenolic acids. Examples of flavonoids include flavonols, flavones,
isoflavones, flavanones, anthocyanidins, and flavanols.6 On the other
hand, phenolic acids include protocatechuic acid, gallic acid, caffeic
GA (Scheme 1), a natural phenolic acid, has been found abundant in
grapes, berries, wine and tea. In recent years, GA received significant
attention because of its potent anti-oxidative activity to scavenge re-
active oxygen species and prevent lipid peroxidation.8 GA also pos-
sesses anti-inflammatory and anti-cancer activity.9 More importantly,
various studies have demonstrated that GA prevents proteins from
misfolding as well as reduces the cell toxicity induced by fibrillar pro-
tein aggregates.10 Specifically, GA interacts with α-Syn to prevent
structural transition to a more compact form that precedes fibril for-
mation. As a result, GA acts as an efficient inhibitor of α-Syn ag-
gregation at the early stages.7c,10b,11
Nevertheless, some evidence suggest that phenolic acids, such as
ferulic acid and chlorogenic acid, are unlikely to cross the blood–brain
barrier (BBB).12a Hence, GA may face the same challenge as a promising
candidate for entering brain and for a direct effect on the central ner-
vous system (CNS). In addition, the desired range of lipophilicity (LogP
value) for candidate molecule crossing BBB is usually from 1 to 5,4 and
there is statistics-based analytical results showed that a LogP < 1.5 is
detrimental for reaching the CNS.12b The LogP value of GA is about
0.42, which is a rather hydrophilic molecule. Accordingly, the valid
7
acid, ferulic acid, rosmarinic acid, chlorogenic acid, etc (Scheme 1).
Polyphenols may exhibit their neuroprotection through a variety of
molecular mechanisms, such as the anti-oxidantion, anti-apoptosis,
⁎ Corresponding authors.
E-mail addresses: zhangyx@zzu.edu.cn (Y.-X. Zhang), xuji@zzu.edu.cn (J. Xu), baisuping@xxmu.edu.cn (S.-P. Bai).
d These authors contributed equally.
https://doi.org/10.1016/j.bmc.2020.115596
Received 3 April 2020; Received in revised form 31 May 2020; Accepted 9 June 2020
Availableonline17June2020
0968-0896/©2020ElsevierLtd.Allrightsreserved.

L. Chen, et al.
Bioorganic&MedicinalChemistry28(2020)115596
towards α-Syn aggregation. Based on our previous work,4 The ThT
fluorescence maximum emission wavelength were optimized as 482 nm
(λem = 482 nm) under excitation light at 450 nm (λex = 450 nm). The
method of inhibitory activities investigation of compounds was de-
scribed in supporting materials. The ThT FI of blank group was mea-
sured without compound, and thereby obtained the maximal FI. The FI
value of blank group was set as 1 (100%). The ratio of FI of the tested
system within compound to that of blank group was defined as the
relative fluorescence intensity (RFI) (Fig. 1). In fact, there was a posi-
tive correlation between the percent reduction of RFI and the inhibitory
activity of the evaluated compound.
Scheme 1. Reported natural phenolic acid inhibitors on α-Syn aggregation.
In detail, the RFIs of first series of compounds—fluorobenzoyl p-
phenylenediamines (1aa-1fe) were shown in Fig. 1A. The RFIs of
second series of compounds, diacyl p-phenylenediamines containing
methyl-protected-GA (1ga-1gi and 2aa-2ba, except 2bb) were shown
in Fig. 1B. More importantly, Fig. 1C showed the RFIs of mono- or di-
acyl p-phenylenediamines containing GA (3g, 3gd-3gi and 4aa-4ba),
GA and other mono-acyl p-phenylenediamines without GA component
as control (1b-1f and 2a-2b). In order to describe the inhibition activity
more conveniently, the ThT RFI was transformed to α-Syn aggregation
inhibition ratio by the formula: α-Syn aggregation inhibition
ratio = 100% (RFI of blank group)-x% (RFI of compound group). In
summary, the lower RFI corresponding to the higher inhibitory activity
against α-Syn aggregation, thereby the ratio intuitively reflected the
inhibitory activity of compound (Tables 1–3).
modification of GA which enhance its lipophilicity and anti-aggregation
performance on α-Syn, should be an efficient approach to discover
novel inhibitors of α-Syn aggregation.
Based on the present evidence above and our hypothesis,4 we focus
on design suitable small molecules containing phenolic acids, which
have strong binding force with the core of non-amyloid-beta component
(NACore), interfere with the formation of β-sheets structure, block the
formation of oligomer nuclei in early stage, and further prevent the
development of α-Syn fibrillation. In the present work, a series of amide
derivatives of GA and other phenolic acids were design and synthesized,
which possess sheet-like conjugated structure and suitable LogP value.
Their inhibitory activities against α-Syn aggregation were evaluate and
the structure–activity-relationship (SAR) studies were studied.
From Table 1, this series of compounds have shown the inhibitory
ratio towards α-Syn aggregation from 1.9% to 42.1%. Compounds 1ab-
1ae were comprised of benzoyl and fluorobenzoyl moieties liked by p-
phenylenediamine, and only 1ae containing block of tetrafluorobenzoyl
displayed more than 30% inhibition ratio (31.2%). Compounds 1fb-1fe
were comprised of (E)-3-(pyridin-4-yl)acryloyl and fluorobenzoyl moi-
eties, among of which, 1fc and 1fd displayed 42.1% and 31.4% in-
hibition ratio, respectively. Apparently, 1fc containing 2,4-di-
fluorobenzoyl has shown better inhibitory activity than 1fd. Other
compounds containing bi-fluorobenzoyl blocks have shown lower in-
hibitory activities.
2. Results and discussion
2.1. Chemistry
Based on our previous work,4 we designed a series of aromatic
amide derivatives, which possess sheet-like conjugated structure as the
potential inhibitor of α-Syn aggregation. The first series of compounds
was unilateral or bilateral flurobenzoyl p- phenylenediamines (1a-1f
and 1aa-1fe). As our main goal, the second series of compounds con-
taining phenolic acyl were designed as N-acyl p-phenylenediamines and
N,N'-diacyl p-phenylenediamines (1g-1gi and 2aa-2ba). Furthermore,
as the most impotant analogues with exposed hydroxyl, 3g, 3gd-3gi
and 4aa-4ba were prepared via demethylation reaction.
From Table 2, compounds 1ga-1gi contain block of methyl-pro-
tested gallic acyl on one side, and on the other side, the blocks are
substituted benzoyls groups or (E)-3-(pyridin-4-yl)acryloyl. Compound
1gd with (E)-3-(pyridin-4-yl)acryloyl demonstrated slight higher ratio
(33.5%) than 1ga, 1gb, 1gc and 1gf, which contain benzoyl or fluor-
obenzoyl on the same side (17.7%, 11.8%, 10.4% and 4.9%). For-
tunately, compound 1gi, N,N’-di(3,4,5-trimethoxybenzoyl) p-phenyle-
nediamine, has shown good inhibitory activity (83.4%).
Firstly, the N-acyl p-phenylenediamine compounds as intermediates
(1a-1g and 2a-2b) were prepared from p-phenylenediamine and dif-
ferent carboxylic acids, such as fluorobenzoic acids, 3,4,5-trimethox-
ybenzoic acid, sulfonic acids and (E)-3-(pyridin-4-yl)acrylic acid via
mono-acylation in medium yields. These reactions were carry out using
2-(7-azobenzotriazole)-N, N, N', N'-tetramethylurea hexafluoropho-
sphate (HATU) as coupling agent and triethylamine as base.13 Fur-
thermore, N,N'-diacyl p-phenylenediamines were synthesized by the
similar approaches. Compound 1aa-1gi and 2aa-2bb were obtained
from 1a to 1g and 2a-2b via further acylation under HATU/ triethy-
lamine in medium to high yields (Scheme 2).
Compounds 2aa-2ba contain block of methyl-protested gallic acyl
on one side, and on the other side, sulfonyls. Tosylate analogue 2aa
showed better ratio than 2ba, which possessed pyridine-3-sulfonyl
group (34.1% vs 22.7%). In particular, compound 2bb, N,N’-di(pyr-
idine-3-sulfonyl) p-phenylenediamine, has shown medium inhibitory
activity (57.2%).
Table 3 indicated various inhibitory activities with different amide
derivatives of GA. Firstly, mono-amide derivatives of p-phenylenedia-
mine, 1f, 1b and 1c displayed the decreasing trend of inhibitory ac-
tivities (42.4%, 34.1% and 16.6%). Mono-sulfonamide derivatives 2a
and 2b, also showed low ratio (15.9% and 15.4%). Noticeably, they all
have lower inhibitory activities than that of GA (78.2%). On the con-
trary, 3g (N-gallic acyl p-phenylenediamine) demonstrated a higher
activity (84.2%) than GA.
The amide derivatives of GA with “exposed hydroxyl”(3g, 3gd-3gi
and 4aa-4ba) were obtained by demethylation of 1g, 1gd-1gi and 2aa-
2ba respectively under boron tribromide in medium to high yields
(Scheme 3).14
2.2. Biological evaluation
2.2.1. In vitro inhibitory activities of the synthesized compounds against α-
Syn aggregation by ThT fluorescence assay
For the bi-amide derivatives, 3gd-3gi and 4aa-4ba have shown
medium to high inhibitory activities from 39.0% to 94.4%. Compounds
3gd-3gi contained one block of gallic acyl and another block of sub-
stituted benzoyl. Among these analogues, 3gf containing fluorobenzoyl
groups displayed outstanding activity comparing with its corresponding
precursor 1gf (Table 2) (85.8% vs 4.9%). The similar trend was found
in 3gd and 1gd (48.9% vs 33.5%). Surprisingly, 3gi which containing
two identical gallic acyl groups displayed lower ratio comparing with
Thioflavin T (ThT) fluorescence assay is a valid method to evaluate
the inhibitory activities of compounds against α-Syn aggregation. ThT
shows weak fluorescence intensity (FI) in monomeric α-Syn, α-Syn
oligomer nuclei or in α-Syn fibril-free system. On the contrary, ThT
displays strong FI in the presence of α-Syn fibrils. ThT FI can quanti-
tatively reflect the kinetics and abilities of compounds inhibiting
2

L. Chen, et al.
Bioorganic&MedicinalChemistry28(2020)115596
Scheme 2. Preparation of N-acyl p-phenylenediamines and N,N'-diacyl p-phenylenediamines. Reagents and conditions: a) CH2Cl2, HATU, trimethylamine, R1COOH
or R2SO3H, rt, 5 h, 45–55%; b) CH2Cl2, HATU, trimethylamine, R3COOH or R4COOH, rt, 5 h, 60–77%.
1gi (39.0% vs 83.4%). Compounds 4aa-4ba possess similar structure
with 3gd-3gi, with sulfonyl groups serves the other acyl block. Com-
paring with their corresponding precursor 2aa and 2ba, the activities
for 4aa and 4ba have shown significant increase (94.3% vs 34.1%,
92.6% vs 22.7%). This suggests that the gallic acyl block in these
compounds act as a key role to inhibit α-Syn aggregation.
In summary, the amide derivatives of GA, are more efficient in-
hibitor against α-Syn aggregation than that of GA. Below are a list of
active candidates: 1gi (83.4%), 3g (84.2%), 3ge (94.4%), 3gf (85.5%),
3gh (91.2%), 4aa (94.3%) and 4ba (92.6%). More importantly, their
LogP values are in the appropriate range (within 1–3) except 3g and
4ba.
2.2.3. The inhibitory kinetics on α-Syn fibrillation and morphology of α-Syn
during the conformation transition with and without compounds 3ge, 3gh
and 4ba.
Based on the results of IC50 study, 3ge and 3gh were selected as
representative amide derivatives of GA with good inhibitory activities
and appropriate LogP values, and 4ba as another representative with
low lipophilicity (LogP 0.74) similar to that of GA (LogP 0.42). The
inhibitory kinetic of these compounds to α-Syn fibrillation was in-
vestigated under the optimized conditions above. The RFI was mea-
sured in 40 μM α-Syn solution incubated with 30 μM of 3ge, 3gh and
4ba respectively (Fig. 2A).
From Fig. 2A, the formation kinetics of α-Syn fibrils without in-
hibitor (black curve) displayed the entire process including lag phase
(formation of β-sheets nucleus), elongation phase (logarithmic increase
of β-sheets), and stationary phase (saturation of β-sheets). The addition
of compounds 3ge, 3gh and 4ba significantly slowed RFI increase over
time, which indicated highly inhibitory effect of these compounds on α-
Syn fibril formation.
2.2.2. IC50 study of the representative amide derivatives of GA.
Comparing with GA, the amide derivatives with higher inhibitory
activities were selected to perform IC50 study (Table 4). The results
indicated that the IC50 values of 3g, 3ge, 3gf, 3gh, 4aa and 4ba have no
distinct differences but all lower than that of GA. Compound 3gh
To confirm the RFI results, the morphology of α-Syn during the
conformation transition with and without inhibitor were observed by
transmission electron microscope (TEM) (Fig. 2B). Fig. 2B-a showed the
image of α-Syn alone after 96 h incubation, which exhibit reticular and
bundled fibrils form. Fig. 2B-b showed the image of α-Syn after 96 h
showed the lowest value (0.98
0.58), and 4ba displayed slightly
higher value than that of GA. More importantly, except for 3g and 4ba,
they possess appropriate LogP values with a high possibility of crossing
the BBB.
Scheme 3. Preparation of amide derivatives of GA. Reagents and conditions: a) Dry DCM, BBr3 (1.0 M in DCM), −78 °C, 20 h, 70–90%.
3

L. Chen, et al.
Bioorganic&MedicinalChemistry28(2020)115596
Fig. 1. The ThT RFIs of compounds. A) RFI of 1aa-1fe. B) RFI of 1ga-2bb. C) RFI of 1b-1g, 2a-2b, GA, 3g, 3gd-3gi and 4aa-4ba. Each compound (30 μM) was
incubated with 40 μM α-Syn solution in 100 mM PBS (pH = 7.4) for 3 days and then 20 μM ThT was added. Each value is mean of three replicates. NS: no
significantly different. (*): p < 0.05, (**): p < 0.01.
incubation with compound 3ge at 30 μM. Compared to the severely
bundled fibrils in Fig. 2B-a, the fibrils appeared sparsely thinner, re-
vealing an effective inhibition of α-Syn fibrillation by 3ge. Fig. 2B-c
showed the image of α-Syn at the same condition with compound 3gh.
The fibrils appeared more sparsely thinner compared with that in
Fig. 2B-b, revealing more effective inhibition by 3gh. Fig. 2B-d showed
the image of α-Syn with compound 4ba, the fibrils appeared similar
with that in Fig.2B-c, also revealing effective inhibition of α-Syn fi-
brillation by 4ba.
2.3. Circular dichroism (CD) spectroscopy analysis
To further confirm the influence of potential inhibitor in α-Syn
aggregation process, CD spectroscopy was utilized as an efficient
method to analyze the variation trend on the secondary structure of the
protein by detecting changes in optical activity during protein ag-
gregation.15
From Fig. 3, the process of changing on the secondary structure of
α-Syn (20 μM) in the presence and absence of compound 3ge, 3gh and
Table 1
The SAR study of 1aa-1fe on α-Syn aggregation.
Cmpd.
R3
LogPa
Ratiob (%)
Cmpd.
R3
LogPa
Ratiob (%)
1aa
1ab
Ph-
3.65
3.8
23.1
14.0
1cb
1cc
4.28
4.59
12.8
10.6
1ac
1ad
3.96
3.96
5.6
1da
1db
4.28
4.59
14.9
6.4
12.2
1ae
4.28
31.2
1ea
4.91
16.0
1ba
1bb
3.96
4.12
11.4
3.6
1fa
1fb
Ph-
2.65
2.81
24.9
24.5
1bc
1bd
4.12
4.44
10.0
23.4
1fc
1fd
2.97
2.97
42.1
31.4
1ca
4.28
7.4
1fe
3.28
1.9
a
b
Calculated by ChemBioDraw 12.0.
The α-Syn aggregation inhibition ratio of compound at 30 μM.
4

L. Chen, et al.
Bioorganic&MedicinalChemistry28(2020)115596
Table 2
The SAR study of 1ga-1gi and 2aa-2bb on α-Syn aggregation.
Cmpd.
R3
LogPa
Ratiob (%)
Cmpd.
R3
R2
LogPa
Ratiob (%)
1ga
Ph-
3.27
17.7
1gg
–
3.75
nd
1gb
1gc
3.58
3.9
11.8
10.4
1gh
1gi
–
–
3.14
2.89
nd
83.4
1gd
1ge
1gf
2.27
3.42
3.58
33.5
nd
2aa
2ba
2bb
–
–
–
3.35
1.53
2.43
34.1
22.7
57.2
4.9
–
a
b
Calculated by ChemBioDraw 12.0.
the α-Syn aggregation inhibition ratio of compound at 30 μM; nd, no detection.
4ba was monitored respectively at initial (0 day) and final time points
(3 days) of α-Syn aggregation. The CD spectra were acquired in the
range of 190–260 nm.
in Fig. 3A, their negative signal at 198 nm were slightly attenuated (α-
Syn + 3ge: −17.3; α-Syn + 3gh: −17.9; α-Syn + 4ba: −19.8),
showing that the random coil structure has been reduced, indicating
that the added compounds 3ge, 3gh and 4ba may interact respectively
with α-Syn, affecting the initial secondary structure of α-Syn (black
lines in Fig. 3B–D). After 3 days of corresponding incubation of α-Syn
with these three compounds (final point), the negative ellipticity at
198 nm decreased completely (red lines in Fig. 3B-D), but with less
extent at the absolute value of 1.3, 1.5 and 7.3 respectively, indicating
that such compounds inhibited the conformational transition of α-Syn.
These results are also consistent to the inhibitory activities against α-
Syn aggregation (from Fig. 2B-b to B-d).
The CD spectrum of un-incubated α-Syn (initial point) is mainly
characterized by a negative signal at 198 nm (−22.7), which is a ty-
pical random coil conformation (black line in Fig. 3A). When the in-
cubation time was extended to three days (final point), the signal at
198 nm was attenuated with absolute value of 12 (red line in Fig. 3A),
indicating a big decrease in random coil conformation. This result is
corresponding to fibril formation of α-Syn (Fig. 2B-a).
On the other hand, α-Syn remained random conformation after the
addition of each compound at initial point. Comparing to the black line
Table 3
The SAR study of 1b-1f, 3g, 2a-2b, 3gd-3gi and 4aa-4ba on α-Syn aggregation.
Cmpd.
R1
R3
LogPa
Ratiob (%)
Cmpd.
R2
R3
LogPa
Ratiob (%)
GA
–
–
0.42
78.2
3gg
–
2.96
62.8
1b
1c
–
–
2.19
2.35
34.1
16.6
3gh
3gi
–
–
2.09
1.31
91.2
39.0
1f
–
–
1.04
0.87
42.4
84.2
2a
2b
–
–
2.13
0.3
15.9
15.4
3g
3gd
3ge
–
–
1.48
2.63
48.9
94.4
4aa
4ba
–
–
2.56
0.74
94.3
92.6
3gf
–
2.79
85.8
a
b
Calculated by ChemBioDraw 12.0.
The α-Syn aggregation inhibition ratio of compound at 30 μM.
5

L. Chen, et al.
Bioorganic&MedicinalChemistry28(2020)115596
Table 4
IC50 study of the representative analogues.
Cmpd.
Structure
LogPa
0.42
IC50 (μM)b
Cmpd.
Structure
LogPa
IC50 (μM)b
GA
4.43
0.68
3g
0.87
2.63
1.34
0.32
0.69
3gh
4aa
2.09
2.56
0.98
1.22
0.58
3ge
1.70
1.95
0.14
1.52
3gf
2.79
0.48
4ba
0.74
8.36
a
b
Calculated by ChemBioDraw 12.0.
Each experiment was set up for three days of incubation, and repeated five times at the same concentration.
Fig. 2. The inhibition kinetics and morphology on α-Syn fibrillation. A) Inhibition kinetics. α-Syn (40 μM) incubated with and without compound 3ge, 3gh and 4ba
at 30 μM respectively, and the experiment was repeated three times. B) Morphology of α-Syn (40 μM) fibrillation after 96 h incubation with and without inhibitors at
30 μM. Scale bar: 500 nm.
Since it has been found anti-oxidative, anti-inflammatory, neuro-
protective and inhibitory activity on α-Syn aggregation, GA has re-
ceived extensive attention, especially on researches about neurode-
generative diseases. Base on the results above, the amide derivatives of
GA demonstrated excellent inhibitory activity towards α-Syn aggrega-
tion, regardless of protected-hydroxyl or exposed-hydroxyl within GA
segment (1gi vs 3g, 3ge-3gh, 4aa-4ba). In general, the latter com-
pounds with exposed-hydroxyl displayed higher inhibitory activity.
These molecule possess sheet-like conjugated structure. According to
our previous hypothetical mechanism,4 these compounds tend to par-
allelly bind the NACore in NAC domain of full-length α-Syn. This
binding was considered reversible and it will interfere the formation of
nuclei, slowing down the oligomerization process, further inhibiting
fibrillation of α-Syn protein. More importantly, most of these com-
pounds possess appropriate LogP value, which may be beneficial for the
candidate to cross the BBB as potential inhibitors against α-Syn
aggregation.
3. Conclusions
We have designed and synthesized 50 compounds with sheet-like
conjugated structural moiety. Among of them, 12 compounds contain
methyl-protected GA block, and 9 compounds contain GA block. These
conjugated sheet-like molecules, especially the amide derivatives of GA
provided robust π-electron delocalized effect and strong hydrogen
bonding properties. These analogues have shown anti-aggregation ac-
tivities in vitro towards α-Syn with IC50 down to 0.98 μM. The mor-
phology and CD analysis of α-Syn aggregation in the presence and
absence of inhibitors have also given the consistent results. This study
will be beneficial to PD therapies targeting α-Syn proteostasis in the
future.
6

L. Chen, et al.
Bioorganic&MedicinalChemistry28(2020)115596
Fig. 3. CD analysis of the secondary structure of α-Syn aggregates formed in the presence and absence of compound 3ge, 3gh and 4ba.
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Acknowledgments
This work supported by the National Natural Science Foundation of
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