Synthesis of a new class of spirooxindole-benzo[b]thiophene-based molecules as acetylcholinesterase inhibitors

Article abstract of DOI:10.3390/molecules25204671

A series of new oxindole-based spiro-heterocycles bearing the benzo[b]thiophene motif were synthesized via a 1,3-dipolar cycloaddition reaction and their acetylcholinesterase (AChE) inhibitory activity was evaluated. All the synthesized compounds exhibite

Full text of DOI:10.3390/molecules25204671

molecules
Article
Synthesis of a New Class of Spirooxindole–
Benzo[b]Thiophene-Based Molecules as
Acetylcholinesterase Inhibitors
Assem Barakat 1,2,
*
, Saeed Alshahrani 1, Abdullah Mohammed Al-Majid 1, M. Ali 1,
Mezna Saleh Altowyan 3, Mohammad Shahidul Islam 1 , Abdullah Saleh Alamary 1,
Sajda Ashraf 4 and Zaheer Ul-Haq 4
1
Department of Chemistry, College of Science, King Saud University, P.O. Box 2455,
Riyadh 11451, Saudi Arabia; chemistry99y@gmail.com (S.A.); amajid@ksu.edu.sa (A.M.A.-M.);
maly.c@ksu.edu.sa (M.A.); mislam@ksu.edu.sa (M.S.I.); alamary1401@yahoo.com (A.S.A.)
Department of Chemistry, Faculty of Science, Alexandria University, P.O. Box 426,
2
Ibrahimia, Alexandria 21321, Egypt
Department of Chemistry, College of Science, Princess Nourah Bint Abdulrahman University,
3
Riyadh 11564, Saudi Arabia; msaltowyan@pnu.edu.sa
Dr. Panjwani Center for Molecular medicine and Drug Research, International Center for Chemical and
4
Biological Sciences, University of Karachi, Karachi 75270, Pakistan; sajda.ashraf@yahoo.com (S.A.);
zaheer_qasmi@hotmail.com (Z.U.-H.)
*
Correspondence: ambarakat@ksu.edu.sa; Tel.: +966-11467-5901; Fax: +966-11467-5992
ꢀꢁꢂꢀꢃꢄꢅꢆꢇ
Academic Editor: Florenci V. González
Received: 15 September 2020; Accepted: 12 October 2020; Published: 13 October 2020
ꢀꢁꢂꢃꢄꢅꢆ
Abstract: A series of new oxindole-based spiro-heterocycles bearing the benzo[b]thiophene motif
were synthesized via a 1,3-dipolar cycloaddition reaction and their acetylcholinesterase (AChE)
inhibitory activity was evaluated. All the synthesized compounds exhibited moderate inhibitory
activities against AChE, while IIc was found to be the most active analog with an IC50 value of
20,840
and octahydroindole moieties. Based on molecular docking studies, IIc was strongly bound to the
catalytic and peripheral anionic sites of the protein through hydrophilic, hydrophobic, and -stacking
µM
·
L
−1. Its molecular structure was a 5-chloro-substituted oxindole bearing benzo[b]thiophene
π
interactions with Asp74, Trp86, Tyr124, Ser125, Glu202, Ser203, Trp236, Trp286, Phe297, Tyr337,
and Tyr341. These interactions also indicated that the multiplicity of the IIc aromatic core significantly
favored its activity.
Keywords: spirooxindole; benzo[b]thiophene; acetylcholinesterase inhibitory activity; molecular
docking
1. Introduction
Alzheimer’s disease (AD) is one of the most common progressive neurodegenerative disorders,
first identified by Alois Alzheimer in 1907. The main AD symptoms are cognitive decline and
memory fragility [
people aged between 65–74 [
fully elucidated, many risk factors are considered to play a crucial role in developing AD including
inflammation, oxidative stress, amyloid- (A ) deposits, protein aggregation, and low acetylcholine
(ACh) levels [ ]. To date, several research teams have focused on the design and development of new
1
]. Relevant global statistics have indicated that AD affects about 3% of elderly
2
]. Although the origin of this memory impairment has not yet been
β
β
τ
3–5
molecules to target AD, while some FDA approved drugs involving the inhibition of cholinesterase
(ChE) using various agents, such as donepezil, and galantamine have also been reported. However,
these compounds have low therapeutic efficacy due to their short half-lives, low bioavailability,
Molecules 2020, 25, 4671; doi:10.3390/molecules25204671
www.mdpi.com/journal/molecules

Molecules 2020, 25, 4671
2 of 16
and high toxicity [
6]. Therefore, there is still an urgent need to develop new, more potent, and less
toxic lead compounds as ChE inhibitors (ChEIs).
Oxindole-based spiro-heterocycles have been extensively studied due to their structure, which is
suitable for various pharmacological targets, while several studies have reported their effective
application as ChEIs [
with improved solubility for drug discovery due to their expected binding efficiency in the target
binding pockets [15 17]. Kia et al. have reported a series of oxindole-based spiro-heterocycles bearing
pyrrolizine and piperidine moieties, which exhibited significant ACh esterase (AChE) inhibitory
activity such as compound A (IC50 = 2.37 0.11 g/mL or 3.33 M) (Figure 1) [ ]. Further studies
7–14]. Their structure has also been used to develop new lead compounds
–
±
µ
µ
7
demonstrated that mono- and bis-spiro-pyrrolidines, such as compound B with an IC50 value of
1.68 ± 0.09 µM (Figure 1), have high potency against AChE [9].
Figure 1. Representative oxindole-based spiro-heterocycles with significant acetylcholinesterase (AChE)
inhibitory activity.
Benzothiophene privilege structure is among the sulfur containing fused herterocycles which
are interesting in drug discovery [18]. Many lead compounds having this nucleus possess divergent
pharmaceutical activities, allowing them to act as anti-inflammatory, anti-cancer, anti-diabetic,
anti-oxidant, anti-microbial, anti-convulsant agents, anti-tubercular, and many more [19–26].
Barakat et al. have also recently reported a compound prepared from an oxindole-based
spiro-heterocycle and a benzo[b]thiophene moiety, which showed moderate activity against AChE
(Figure 1) [12]. Here, we performed a structure–activity relationship study to further explore the
extension and substrate scope of a new series of spirooxindole–benzo[b]thiophene-based analogs.
Their AChE inhibitory activity was also assessed in vitro, while molecular modelling studies were

Molecules 2020, 25, 4671
3 of 16
performed to elucidate the structural features and interactions that favor the inhibitory activity of the
examined compounds.
2. Results and Discussion
2.1. Synthesis of Analogs IIa–n
The general substrate scope of the synthetic compounds IIa–n is shown in Scheme 1.
In particular, a series of spiro-oxindole-tethered benzo[b]thiophene scaffolds consisting of a single
regio- and diastereo-selective isomer were synthesized from benzo[b]thiophene-based chalcones
(
2a
substituted acetophenones (1a
cycloaddition reaction protocol [27
–
e
), which were in turn prepared through an aldol condensation of the corresponding
) with benzo[b]thiophene-2-carboxaldehyde. Following the 1,3-dipolar
34], 2a reacted with 5-substituted isatins (3a ) and secondary
–e
–
–
e
–c
amino acids, such as thioproline (4a) and octahydro-1H-indole-2-carboxylic acid (4b), forming
the corresponding spiro-oxindole analogs IIa–n in high yields after purification by short column
chromatography. The chemical features of the target compounds were assigned based on the NMR
spectrum. As an example, the 1H-NMR spectrum of the IIa shows the characteristic peaks; the NH
proton at
bicyclic rings are shown in the chemical shift between
to benzoyl group 4.78 (d, J = 10.5 Hz, 1H, C CO), C-H proton close to the benzothiophene ring at
δ
10.53 ppm; the aromatic protons in the region of
δ 7.90–6.53 ppm; the protons of the fused
δ
4.78–3.17 ppm as follows: the proton adjacent
H
δ
4.37 (t, J = 5.1 Hz, 1H, C
H
), the four protons of the two CH2 groups appeared differently
δ
4.24
3.17
(d, J = 10.8 Hz, 1H, CH2),
δ
3.74 (d, J = 10.8 Hz, 1H, CH2), 3.25 (d, J = 11.4 Hz, 1H, CH2),
δ
δ
(dd, J = 11.6, 5.7 Hz, 1H, CH2); and the proton of the CHN shown at
δ 3.38 (d, J = 10.4 Hz, 1H, CH).
The 13C-NMR spectrum of the proposed carbon is perfectly shown in Figure 2.
Scheme 1. Synthesis of compounds IIa–n.
2.2. In Vitro Biological Activity
The in vitro cholinesterase inhibitory activity of the 14 synthesized spiro-oxindole derivatives
(
IIa
ranging between 20,840–121,690
highest inhibitory activity with an IC50 value of 20,840; 37,670; 34,020; 23,040; 29,760 and 36,830
–
n
) were evaluated [
9
]. As shown in Table 1, all analogs exhibited AChE activity with IC50 values
−1. Among them, analogs IIc
IId IIf IIg IIl and IIn showed the
M L−1
µM·L
,
,
,
,
µ
respectively, while IIc with a 5-chloro-substituted oxindole structure bearing benzo[b]thiophene and
octahydroindole moieties, was the most active in this group. The second group of spiro-oxindole
analogs (IIe
moderate activity, comparable to that of the positive control. In contrast, the last group included
analogs IIa IIb, and IIh IIj, which showed weak AChE inhibitory activity with an IC50 value of more
than 70,000 µM L−1. Galantamine was used as a positive control for comparison.
, IIk, and IIm) with an IC50 value of 50,590; 41,530 and 41,450 µ
M L−1 respectively showed
,
–

Molecules 2020, 25, 4671
4 of 16
Figure 2. 1H-NMR and 13C-NMR of IIa in DMSO-d6.

Molecules 2020, 25, 4671
5 of 16
Table 1. Chemical structures of the synthesized spiro-oxindole analogs IIa–n and their AChE
inhibitory activity.
AChE Inhibition
#
Chemical Structures
IC50 (µM/L)
1
85,560
IIa
IIb
2
3
88,410
20,840
IIc
4
5
6
37,670
50,590
34,020
IId
IIe
IIf

Molecules 2020, 25, 4671
6 of 16
Table 1. Cont.
Chemical Structures
AChE Inhibition
#
7
IC50 (µM/L)
23,040
IIg
8
9
121,690
IIh
72,380
75,980
IIi
IIj
10
11
41,530
IIk
12
29,760
IIl

Molecules 2020, 25, 4671
7 of 16
Table 1. Cont.
Chemical Structures
AChE Inhibition
#
IC50 (µM/L)
13
14
41,450
36,830
IIm
IIn
2090 [7–11]
3400 [34]
0.35 [35]
STD
Galantamine
2.3. Molecular Docking Study
In order to identify the binding pattern and mechanism of the synthesized spiro-benzothiophene
derivatives, the complex AChE enzyme was studied using molecular docking based on the atomic
coordinates of the crystallographic structure of galantamine. All analogs were docked using the
default MOE docking protocol and the obtained conformations were visually analyzed to elucidate
the existing interactions. The binding affinity of the standard reference galantamine, was attributed
to the presence of hydroxyl groups that might help its stabilization in the binding pocket through
classical and non-classical hydrogen bonds with Ser203 and Tyr337 (Figure 3). The docking results
indicated that the conformations with the highest score (Table 2) of all the derivatives fitted well
into the binding cavity of the AChE enzyme by developing different interactions with the active site
residues Asp74, Gly82, Thr83, Trp86, Gly121, Glu202, Ser203, Tyr337, Tyr341, and His447. However,
compounds bearing electronegative substituents on the benzene ring developed weak hydrophobic
interactions with the surrounding residues. Moreover, bulky substituents at the R-position could
reduce the compounds’ activity due to steric hindrance. Thus, the elucidation of the structural features
and the effect of different substituents on the spiro-benzothiophene derivatives, such as the presence of
electron-donating (H and NH2) or electron-withdrawing (halogen and haloalkyl) groups, explained
the effective inhibition of the AChE enzyme. As depicted in Figure 4A, the most active compound
(
IIc) with an IC50 value of 20,840
µ
ML−1 was strongly bound to the catalytic and peripheral anionic
-stacking interactions with Tyr72, Asp74, Trp86, Tyr124,
sites of the protein through hydrophobic and
π
Trp286, Phe297, Tyr337, Phe338, and Tyr341. These interactions further indicated the beneficial effect of
the aromatic core multiplicity on the compound’s high activity. Hydrophilic interactions were also
observed between the hydrogen-bond donor of Ser125 and the carbonyl group of IIc at a distance of
2.6 Å. Their interaction was further stabilized by a special halogen bond interaction between the 5-Cl
atom and Asp74.

Molecules 2020, 25, 4671
8 of 16
Figure 3. Re-docking of a galantamine molecule. Galantamine (PDB ID: 4EY6) is indicated with yellow
color and its docked confirmation is presented in magenta. Ligand heavy atom root-mean-square
deviation (RMSD) = 0.81 Å.
Table 2. Docking scores of the selected compound of spiro-benzothiophene series with AChE.
S.No.
Compounds Name
Scores
−6.54
−6.01
−6.23
−6.19
−9.28
No. of Hydrogen Bonds
1
2
3
4
5
IIc
3
3
3
4
2
IIf
IIg
IIl
Galantamine
Compounds IIf, IIg, and IIl also exhibited moderate activities in the range of 23,040–34,020 µ .
M L−1
Except for a few interactions, their molecular docking images were similar to the binding mode of
IIc. Compound IIf, bearing a Cl deactivating group on the phenyl ring, showed less binding affinity
than IIc bearing a hydrogen atom on the same phenyl ring position. Moreover, the side chain of the
Asp74, Tyr124, and Tyr337 residues developed hydrophilic interactions with the carbonyl and NH2
groups of the IIf indoline ring at distances of 2.6, 3.1, and 2.3 Å, respectively (Figure 4B). Furthermore,
as shown in Figure 4C,D, the proposed binding mode of compounds IIg and IIl was very similar.
In particular, most interactions were located in the region of the Trp86, Tyr124, Trp286, Phe295, Phe297,
Tyr337, Phe338, Tyr341, and Tyr449 amino acid residues due to development of hydrophobic and
π
interactions at the catalytic anionic site and the formation of hydrogen bonds with Tyr124, Ser125,
Gly121, and Ser203 at the edge of the peripheral region. However, the carbonyl and NH2 groups of
the IIg indoline ring developed hydrophilic interactions with Gly121, Ser125, Glu202, and Ser203
at distances of 2.1, 3.2, 2.9, and 2.8 Å, respectively, whereas the hydrophilic interactions of IIl with
Gly121, Ser125, and Glu202 were observed at distances of 2.7, 2.5, and 2.9 Å, respectively. Moreover,
the halogen bond formed with Gly82 and Ser203 further enhanced the binding of IIl. Therefore,
the docking results of the synthesized spiro-benzothiophene derivatives were in good agreement with
the experimental findings, providing significant information about their binding mechanism to AChE.

Molecules 2020, 25, 4671
9 of 16
Figure 4. Visual presentation of the binding patterns of the spiro-benzothiophene derivatives (A) IIc,
(B) IIf, (C) IIg, and (D) IIl. The dotted lines indicate the intramolecular hydrogen bonds.
3. Materials and Methods
3.1. General Experimental Information
All the chemicals were purchased from Sigma–Aldrich (Riedstraße, Germany), and Fluka (Buchs,
Switzerland), and were used without further purification, unless otherwise reported. The melting
points were measured on a Gallenkamp melting point apparatus (Bibby Scientific Limited, Beacon
Road, Stone, Staffordshire, UK) in open glass capillaries and are not corrected. The infrared (IR) spectra
were measured as KBr pellets on a Nicolet 6700 Fourier-transform IR spectrophotometer (Thermo
Fisher Scientific, Madison, WI, USA). The 1H (400 MHz) and 13C (100 MHz) nuclear magnetic resonance
(NMR) spectra were recorded on a Varian Mercury Jeol-400 NMR spectrometer (Tokyo, Japan) in
CDCl3 or DMSO-d6. The chemical shifts (δ) are provided in ppm and the J coupling constants in Hz.
The mass spectra were recorded on a JEOL JMS-600 H mass spectrometer (Santa Clara, CA, USA). while
the elemental analysis of the synthesized compounds was performed using an Elmer 2400 Elemental
Analyzer (CHN mode) (Perkin Elmer, Waltham, MA, USA). The AChE assay and molecular docking
protocols are described in the supporting information.
3.2. General Procedure for the Synthesis of Chalcones 2a–e
The chalcone derivatives 2a–e were synthesized based on a reported procedure [27,28] using
benzo[b]thiophene-2-carboxaldehyde (1.0 eq.) in ethanol and the corresponding substituted
acetophenone (acetopehnone, p-Cl- acetopehnone, p-Br-acetopehnone, p-F-acetopehnone and
p-CF3-acetopehnone) (1.0 eq.) (1a–e) in the presence of aqueous NaOH.

Molecules 2020, 25, 4671
10 of 16
(E)-3-(Benzo[b]thiophen-2-yl)-1-phenylprop-2-en-1-one (2a). The spectrum is consistent with the reported
literature [36,37].
1
(E)-3-(Benzo[b]thiophen-2-yl)-1-(4-chlorophenyl)prop-2-en-1-one (2b). H-NMR (400 MHz, CDCl3)
δ: 7.30
(d, 1H, J = 15.24 Hz, CH=CH), 7.44–7.36 (m, 2H, Ar–H), 7.49 (d, 1H, J = 8.16 Hz, Ar–H), 7.56 (s, 1H,
C=CH), 7.81 (dd, 2H, J = 10.6, 7.84 Hz, Ar–H), 7.97 (d, 2H, J = 8.08 Hz, Ar–H), 8.05 (d, 1H, J = 15.28 Hz,
CH=CH); 13C-NMR (100 MHz, CDCl3)
δ: 188.4, 140.4, 140.2, 139.7, 139.5, 136.3, 130.1, 129.9, 129.3,
128.9, 124.9.
1
(E)-3-(Benzo[b]thiophen-2-yl)-1-(4-fluorophenyl)prop-2-en-1-one (2c). H-NMR (400 MHz, CDCl3)
δ: 7.18
(t, 1H, J = 15.24 Hz, Ar–H), 7.35 (d, 1H, J = 15.24 Hz, CH=CH), 7.42–7.35 (m, 2H, Ar–H), 7.49 (d, 1H,
J = 8.16 Hz, Ar–H), 7.57 (s, 1H, C=CH), 7.82 (dd, 2H, J = 10.6, 7.84 Hz, Ar–H), 8.06 (d, 2H, J = 8.08 Hz,
Ar–H), 8.08 (d, 1H, J = 15.28 Hz, CH=CH); 13C-NMR (100 MHz, CDCl3)
δ: 188.0, 140.3, 140.2, 139.7,
134.4, 134.3, 131.2, 122.6, 115.8.
(E)-3-(Benzo[b]thiophen-2-yl)-1-(4-(trifluoromethyl)phenyl)prop-2-en-1-one (2d). 1H-NMR (400 MHz,
CDCl3) : 7.30 (d, 1H, J = 15.24 Hz, CH=CH), 7.43–7.37 (m, 2H, Ar–H), 7.49 (d, 1H, J = 8.16 Hz, Ar–H),
δ
7.60 (s, 1H, C=CH), 7.83 (dd, 2H, J = 10.6, 7.84 Hz, Ar–H), 8.07 (d, 2H, J = 8.08 Hz, Ar–H), 8.12 (d, 1H,
J = 15.28 Hz, CH=CH); 13C-NMR (100 MHz, CDCl3)
δ: 188.8, 140.9, 140.5, 139.9, 139.7, 130.8, 128.8,
126.9, 125.8, 125.1, 124.8, 122.4.
1
(E)-3-(Benzo[b]thiophen-2-yl)-1-(4-bromophenyl)prop-2-en-1-one (2e). H-NMR (400 MHz, CDCl3)
δ: 7.25
(d, 1H, J = 15.24 Hz, CH=CH), 7.41–7.37 (m, 2H, Ar–H), 7.49 (d, 1H, J = 8.16 Hz, Ar–H), 7.58 (s, 1H,
C=CH), 7.82 (dd, 2H, J = 10.6, 7.84 Hz, Ar–H), 7.90 (d, 2H, J = 8.08 Hz, Ar–H), 8.05 (d, 1H, J = 15.28 Hz,
CH=CH); 13C-NMR (100 MHz, CDCl3)
δ: 188.6, 140.4, 140.2, 139.7, 136.8, 130.1, 129.9, 129.3, 128.9, 124.9.
3.3. General Procedure for the Synthesis of Oxindole-Based Spiro-Heterocycles IIa–n
The oxindole-based spiro-heterocycles IIa were synthesized through an one-pot reaction using
equimolar amounts of each chalcone (2a ), amino acid (4a ) (1.0 mmol), and substituted isatin
3a–c, 1.0 mmol), which were refluxed in methanol (10 mL) for 1–3 h. All analogs were obtained as
–
n
–
e
–b
(
precipitates, which were filtered and washed with a small amount of MeOH. The final product were
separated in faint yellow color.
(3S)-70-(Benzo[b]thiophen-2-yl)-60-benzoyl-5-chloro-30,60,70,7a’-tetrahydro-10H-spiro[indoline-3,50-pyrrolo
[1,2-c]thiazol]-2-one(IIa). Analog IIa was synthesized using 2a (264 mg), 5-ch◦loro-isatin (3b) (181 mg),
1
and thioproline 4a (133 mg). Yield: 470 mg (0.91 mmol, 91%); m.p: 121 C; H-NMR (400 MHz,
DMSO-d6)
Ar– ), 7.54 (t, J = 7.4 Hz, 1H, Ar–
(dd, J = 8.5, 1.7 Hz, 1H, Ar– ), 6.53 (d, J = 8.1 Hz, 1H, Ar–
J = 5.1 Hz, 1H, C ), 4.24 (d, J = 10.8 Hz, 1H, C 2), 3.74 (d, J = 10.8 Hz, 1H, C
1H, CH), 3.25 (d, J = 11.4 Hz, 1H, CH2), 3.17 (dd, J = 11.6, 5.7 Hz, 1H, CH2); 13C-NMR (126 MHz,
DMSO-d6) : 196.26, 178.30, 143.15, 141.45, 139.96, 138.74, 136.52, 134.37, 130.47, 129.17, 128.42, 128.11,
125.69, 125.14, 125.06, 124.86, 123.91, 123.10, 122.94, 111.65, 74.27, 74.24, 62.14, 54.28, 47.31, 36.51; IR (KBr,
δ
: 10.53 (s, 1H, N
H
), 7.90 (d, J = 7.7 Hz, 1H, Ar–
H
H
H
), 7.77 (d, J = 8.0 Hz, 1H, Ar–
), 7.32 (dt, J = 19.8, 7.6 Hz, 4H, Ar–
), 4.78 (d, J = 10.5 Hz, 1H, C CO), 4.37 (t,
2), 3.38 (d, J = 10.4 Hz,
H
), 7.57 (s, 1H,
H
H), 7.49–7.39 (m, 3H, Ar–
H), 7.21
H
H
H
H
H
δ
cm−1
) νmax = 1475, 1548, 1605, 1705, 2915, 3100, 3265; [Anal. Calcd. for C28H21ClN2O2S2: C, 65.04; H,
4.09; N, 5.42; Found: C, 64.93; H, 4.21; N, 5.65]; LC/MS (ESI, m/z): 517.10 [M+].
(3S)-70-(Benzo[b]thiophen-2-yl)-60-benzoyl-30,60,70,7a’-tetrahydro-10H-spiro[indoline-3,50-pyrrolo[1,2-c]thiazol]
-2-one(IIb). Analog IIb was prepared using 2a (264 mg), isatin (3a) (147 mg), and thioproline (4a
(133 mg). Yield: 443 mg (0.92 mmol, 92%); m.p: 65 ◦C; 1H-NMR (400 MHz, DMSO-d6)
: 10.39 (s, 1H,
), 7.90 (d, J = 7.8 Hz, 1H, Ar– ), 7.77 (d, J = 7.8 Hz, 1H, Ar– ), 7.52 (d, J = 12.4 Hz, 2H, Ar– ),
7.44–7.24 (m, 7H, Ar– ), 7.13 (s, 1H, Ar– ), 6.95 (s, 1H, Ar– ), 6.51 (d, J = 7.8 Hz, 1H, Ar– ), 4.76 (s,
)
δ
NH
H
H
H
H
H
H
H

Molecules 2020, 25, 4671
11 of 16
1H, C
3.18 (t, J = 2.8 Hz, 2H, CH2); 13C-NMR (101 MHz, DMSO-d6)
138.82, 136.80, 134.07, 130.53, 129.04, 128.53, 128.11, 125.13, 124.82, 123.91, 123.23, 122.99, 121.67, 110.20,
74.3, 73.90, 62.45, 54.00, 47.20, 36.51; IR (KBr, cm−1
max = 1485, 1548, 1610, 1718, 2930, 3135, 3285; [Anal.
H
CO), 4.27 (d, J = 8.7 Hz, 2H, C
H
), 3.73 (d, J = 10.2 Hz, 1H, CH2), 3.34 (d, J = 10.2 Hz, 1H, C
H),
δ: 196.36, 178.65, 143.65, 142.63, 140.07,
)
ν
Calcd. for C28H22N2O2S2: C, 69.68; H, 4.59; N, 5.80; Found: C, 69.79; H, 4.47; N, 6.01]; LC/MS (ESI, m/z):
483.20 [M+].
(3S)-10-(Benzo[b]thiophen-2-yl)-20-benzoyl-5-chloro-10,20,50,5a’,60,70,80,90,9a’,9b’-decahydrospiro[indoline-3,
30-pyrrolo[2,1-a]isoindol]-2-one(IIc). Analog IIc was prepared using 2a (264 mg), 5-chloro-isatin (3b
)
(181 mg), and (2S,3aS,7aS)-octahydro-1H-indole-2-carboxylic acid 4b (169 mg). Yield: 458 mg
(0.83 mmol, 83%); m.p: 110 ◦C; 1H-NMR (400 MHz, DMSO-d6)
: 10.27 (s, 1H), 7.87 (d, J = 7.9 Hz, 1H,
), 7.52 (d, J = 7.1 Hz, 1H, Ar– ), 7.48–7.24 (m, 8H, Ar– ), 7.18 (d,
), 4.93 (d, J = 11.7 Hz, 1H, C CO), 4.34 (t, J = 10.8 Hz,
), 2.21–2.01 (m, 2H, CH2), 1.71 (dd, J = 11.2,
δ
Ar–
J = 8.2 Hz, 1H, Ar–
1H, C
H
), 7.74 (d, J = 7.8 Hz, 1H, Ar–
H
H
H
H), 6.48 (d, J = 8.2 Hz, 1H, Ar–
H
H
H), 4.27–4.18 (m, 1H, CH), 3.17 (d, J = 3.7 Hz, 1H, CH
6.0 Hz, 1H, CH2), 1.51 (ddd, J = 14.1, 9.6, 4.4 Hz, 2H, CH2), 1.33 (p, J = 11.8, 10.7 Hz, 2H, CH2), 1.09 (dd,
J = 10.9, 6.1 Hz, 1H, CH2), 1.04–0.93 (m, 1H, CH2), 0.88 (tt, J = 13.3, 3.8 Hz, 1H, CH2), 0.77–0.65 (m, 1H,
C
H2); 13C-NMR (101 MHz, DMSO-d6)
δ
: 196.53, 180.04, 144.01, 141.23, 140.09, 138.74, 136.97, 134.08,
129.87, 129.07, 128.33, 128.19, 125.94, 125.78, 124.96, 124.52, 123.72, 122.86, 121.87, 111.38, 71.90, 70.94,
65.17, 57.38, 48.71, 41.92, 36.70, 28.05, 27.96, 25.00, 19.70; IR (KBr, cm−1
max = 1480, 1555, 1608, 1725,
)
ν
2920, 31,125, 3285; [Anal. Calcd. for C33H29ClN2O2S: C, 71.66; H, 5.28; N, 5.06; Found: C, 71.49; H, 5.13;
N, 5.22]; LC/MS (ESI, m/z): 553.20 [M+].
(3S)-70-(Benzo[b]thiophen-2-yl)-5-chloro-60-(4-chlorobenzoyl)-30,60,70,7a’-tetrahydro-10H-spiro[indoline-3,50-
pyrrolo[1,2-c]thiazol]-2-one(IId). Analog IId was obtained using 2b (298 mg), 5-chloro-isatin (3b) (181 mg),
◦
1
and thioproline (4a) (133 mg). Yield: 478 mg (0.87 mmol, 87%); m.p: 60 C; H-NMR (400 MHz,
DMSO-d6) : 10.55 (s, 1H, N ), 7.91 (d, J = 7.9 Hz, 1H, Ar– ), 7.77 (d, J = 7.4 Hz, 1H, Ar– ), 7.58 (s,
1H, Ar– ), 7.44 (dt, J = 14.4, 6.2 Hz, 5H, Ar– ), 7.33 (dt, J = 18.4, 7.3 Hz, 2H, Ar– ), 7.26–7.19 (m, 1H,
Ar– ), 6.56 (d, J = 8.2 Hz, 1H, Ar– ), 4.76 (d, J = 10.9 Hz, 1H, C CO), 4.30–4.17 (m, 2H, CH2), 3.74 (d,
J = 10.4 Hz, 1H, C ), 3.39 (d, J = 10.6 Hz, 1H, C
5.9 Hz, 1H, C
2); 13C-NMR (126 MHz, DMSO-d6)
δ
H
H
H
H
H
H
H
H
H
H
H), 3.26 (d, J = 11.4 Hz, 1H, CH2), 3.17 (dd, J = 11.7,
H
δ: 195.47, 178.04, 143.07, 141.49, 139.99, 139.26, 138.79,
135.26, 130.57, 130.00, 129.31, 128.39, 125.71, 125.12, 124.99, 124.83, 123.89, 123.13, 122.96, 111.66, 74.33,
74.20, 62.43, 54.28, 47.02, 36.52; IR (KBr, cm−1
max = 1485, 1498, 1534, 1634, 1726, 2934, 3088, 3288; [Anal.
)
ν
Calcd. for C28H20Cl2N2O2S2: C, 60.98; H, 3.66; N, 5.08; Found: C, 61.14; H, 3.52; N, 5.24]; LC/MS (ESI,
m/z): 551.10 [M+].
(3S)-70-(Benzo[b]thiophen-2-yl)-60-(4-chlorobenzoyl)-30,60,70,7a’-tetrahydro-10H-spiro[indoline-3,50-pyrrolo[1,
2-c]thiazol]-2-one (IIe). Analog IIe was prepared using 2b (298 mg), isatin (3a) (147 mg), and thioproline
(
4a) (133 mg). Yield 480 mg (0.93 mmol, 93%); m.p: 128 ◦C; 1H-NMR (400 MHz, DMSO-d6)
δ
: 10.40 (s,
), 7.45–7.37 (m,
), 6.96 (t, J = 7.5 Hz, 1H, Ar– ), 6.53
CO), 4.32–4.19 (m, 2H, CH2), 3.73 (d, J = 10.3 Hz,
), 3.19 (d, J = 5.7 Hz, 2H, CH2); 13C-NMR (126 MHz, DMSO-d6)
: 195.56, 178.46, 143.41, 142.55, 140.02, 138.95, 138.78, 135.44, 130.62, 129.89, 129.14, 128.43, 125.10,
1H, N
3H, Ar–
(d, J = 7.8 Hz, 1H, Ar–
1H, C ), 3.36 (d, J = 10.3 Hz, 1H, C
H
), 7.90 (d, J = 7.9 Hz, 1H, Ar–
), 7.36–7.27 (m, 4H, Ar– ), 7.15 (t, J = 7.7 Hz, 1H, Ar–
), 4.74 (d, J = 10.8 Hz, 1H, C
H), 7.77 (d, J = 7.9 Hz, 1H, Ar–H), 7.55 (s, 1H, Ar–H
H
H
H
H
H
H
H
H
δ
124.79, 123.87, 123.04, 122.94, 122.89, 121.69, 110.22, 74.33, 73.90, 62.60, 54.05, 46.95, 36.50; IR (KBr,
cm−1) νmax = 1485, 1545, 1615, 1715, 2920, 3115, 3275; [Anal. Calcd. for C28H21ClN2O2S2: C, 65.04; H,
4.09; N, 5.42; Found: C, 65.27; H, 4.16; N, 5.59]; LC/MS (ESI, m/z): 517.20 [M+].
(3S)-10-(Benzo[b]thiophen-2-yl)-5-chloro-20-(4-chlorobenzoyl)-10,20,50,5a’,60,70,80,90,9a’,9b’-decahydrospiro
[indoline-3,30-pyrrolo[2,1-a]isoindol]-2-one(IIf). Analog IIf was prepared using 2b (298 mg), 5-chloro-isatin
(
3b) (181 mg), and (2S,3aS,7aS)-octahydro-1H-indole-2-carboxylic acid (4b) (169 mg). Yield: 504 mg
(0.86 mmol, 86%); m.p: 119 ◦C; 1H-NMR (400 MHz, DMSO-d6)
δ
: 10.29 (s, 1H, N ), 7.87 (d, J = 8.0 Hz,
H

Molecules 2020, 25, 4671
12 of 16
1H, Ar–
7.19 (d, J = 8.4 Hz, 1H Ar–
J = 10.8 Hz, 1H, C ), 4.22 (t, J = 7.5 Hz, 1H, C
H2), 1.70 (dd, J = 11.1, 6.1 Hz, 1H, CH2), 1.50 (s, 2H, CH2), 1.40–1.22 (m, 2H, CH2), 1.05–0.93 (m, 1H,
2), 0.90–0.78 (m, 2H, C 2), 0.70 (d, J = 13.8 Hz, 1H, C : 195.72,
2); 13C-NMR (101 MHz, DMSO-d6)
H
), 7.74 (d, J = 7.6 Hz, 1H, Ar–
), 6.51 (d, J = 8.6 Hz, 1H Ar–
), 3.16 (d, J = 3.7 Hz, 1H, C
H
), 7.49–7.34 (m, 6H Ar–
), 4.91 (d, J = 11.6 Hz, 1H, C
), 2.11 (q, J = 9.1, 8.2 Hz, 2H,
H
), 7.29 (dd, J = 11.2, 7.4 Hz, 2H Ar–
H),
H
H
HCO), 4.33 (t,
H
H
H
C
C
H
H
H
δ
179.93, 143.84, 141.17, 140.09, 139.03, 138.77, 135.64, 130.03, 129.23, 128.53, 125.90, 125.82, 124.97, 124.54,
123.80, 122.84, 121.88, 111.46, 100.01, 81.05, 71.89, 70.97, 65.29, 57.37, 48.44, 41.96, 36.66, 27.98, 25.00, 19.68;
IR (KBr, cm−1
) νmax = 1465, 1501, 1532, 1615, 1735, 2900, 3015, 3270; [Anal. Calcd. for C33H28Cl2N2O2S:
C, 67.46; H, 4.80; N, 4.77; Found: C, 67.35; H, 4.93; N, 4.86]; LC/MS (ESI, m/z): 587.20 [M+].
(3S)-70-(Benzo[b]thiophen-2-yl)-5-chloro-60-(4-fluorobenzoyl)-30,60,70,7a’-tetrahydro-10H-spiro[indoline-3,50-
pyrrolo[1,2-c]thiazol]-2-one(IIg). Analog IIg was prepared using 2c (282 mg), 5-chloro-isatin (3b) (181 mg),
◦
1
and thioproline (4a) (133 mg). Yield: 491 mg (0.92 mmol, 92%); m.p: 130 C; H-NMR (400 MHz,
DMSO-d6) : 10.54 (s, 1H, N ), 7.90 (d, J = 7.9 Hz, 1H, Ar– ), 7.77 (d, J = 7.8 Hz, 1H, Ar– ), 7.58 (s, 1H,
Ar– ), 7.54–7.43 (m, 3H, Ar– ), 7.39–7.28 (m, 2H, Ar– ), 7.26–7.13 (m, 3H, Ar– ), 6.56 (d, J = 8.2 Hz,
1H, Ar– ), 4.77 (d, J = 10.9 Hz, 1H, C CO), 4.30–4.18 (m, 2H, C 2), 3.74 (d, J = 10.4 Hz, 1H, C ), 3.40 (s,
1H, C : 194.92, 178.15,
), 3.30–3.22 (m, 1H, CH2), 3.15 (s, 1H, CH2); 13C-NMR (126 MHz, DMSO-d6)
δ
H
H
H
H
H
H
H
H
H
H
H
H
δ
166.73, 164.72, 143.14, 141.49, 140.00, 138.81, 133.36, 131.26, 131.18, 130.51, 128.42, 125.73, 125.09, 125.05,
124.80, 123.87, 123.08, 122.93, 116.34, 116.17, 111.63, 74.32, 74.25, 62.38, 54.28, 47.12, 36.54; [Anal. Calcd.
for C28H20ClFN2O2S2: C, 62.85; H, 3.77; N, 5.24; Found: C, 63.04; H, 3.63; N, 5.11]; LC/MS (ESI, m/z):
535.10 [M+].
(3S)-10-(Benzo[b]thiophen-2-yl)-5-chloro-20-(4-fluorobenzoyl)-10,20,50,5a’,60,70,80,90,9a’,9b’-decahydrospiro
[indoline-3,30-pyrrolo[2,1-a]isoindol]-2-one(IIh). Analog IIh was prepared using 2c (282 mg), 5-chloro-isatin
(
3b) (181 mg), and (2S,3aS,7aS)-octahydro-1H-indole-2-carboxylic acid (4b) (169 mg). Yield: 496 mg
(0.87 mmol, 87%); m.p: 125 ◦C; 1H-NMR (400 MHz, DMSO-d6)
: 10.28 (s, 1H, N ), 7.87 (d, J = 7.8 Hz,
1H, Ar– ), 7.74 (d, J = 7.6 Hz, 1H, Ar– ), 7.49 (dd, J = 8.7, 5.3 Hz, 2H, Ar– ), 7.46 (s, 1H, Ar– ), 7.39
(s, 1H, Ar– ), 7.29 (dd, J = 11.6, 7.4 Hz, 2H, Ar– ), 7.20 (t, J = 8.7 Hz, 3H, Ar– ), 6.51 (d, J = 8.3 Hz,
1H, Ar– ), 4.92 (d, J = 11.6 Hz, 1H, C ), 4.22 (t, J = 8.8 Hz, 1H,
), 3.16 (d, J = 3.8 Hz, 1H, C
δ
H
H
H
H
H
H
H
H
H
HCO), 4.32 (t, J = 10.7 Hz, 1H, CH
CH
H), 2.10 (td, J = 11.5, 10.7, 6.6 Hz, 2H, CH2), 1.70 (dd, J = 11.2, 6.1 Hz,
1H, CH2), 1.58–1.43 (m, 2H, CH2), 1.39–1.24 (m, 2H, CH2), 1.03–0.93 (m, 1H, CH2), 0.87 (dd, J = 7.5,
4.3 Hz, 1H, CH2), 0.79 (t, J = 3.3 Hz, 1H, CH2), 0.71 (d, J = 13.6 Hz, 1H, CH2); 13C-NMR (101 MHz,
DMSO-d6)
125.87, 124.96, 124.51, 123.70, 122.89, 121.85, 116.17, 111.42, 71.92, 71.01, 70.87, 65.41, 57.37, 48.52, 41.98,
36.70, 28.01, 24.99, 19.69; IR (KBr, cm−1
max = 1485, 1545, 1615, 1715, 2920, 3115, 3275; [Anal. Calcd.
δ: 195.22, 180.01, 166.92, 164.40, 143.90, 141.17, 140.10, 138.77, 133.73, 131.21, 129.87, 128.28,
)
ν
for C33H28ClFN2O2S: C, 69.40; H, 4.94; N, 4.91; Found: C, 69.61; H, 5.12; N, 4.79]; LC/MS (ESI, m/z):
571.20 [M+].
(3S)-70-(Benzo[b]thiophen-2-yl)-5-chloro-60-(4-(trifluoromethyl)benzoyl)-30,60,70,7a’-tetrahydro-10H-spiro
[indoline-3,50-pyrrolo[1,2-c]thiazol]-2-one (IIi). Analog IIi was prepared using 2d (332 mg), 5-chloro-isatin
(
3b) (181 mg), and thioproline (4a) (133 mg). Yield: 531 mg (0.91 mmol, 91%); m.p: 114 ◦C; 1H-NMR
(400 MHz, DMSO-d6)
7.73 (d, J = 8.1 Hz, 2H, Ar–
Ar– ), 7.33 (dt, J = 17.7, 7.1 Hz, 2H, Ar–
4.84 (d, J = 10.7 Hz, 1H, C CO), 4.25 (q, J = 9.5, 8.0 Hz, 2H, C
J = 10.7 Hz, 1H, C
(101 MHz, DMSO-d6)
128.96, 128.39, 126.18, 125.87, 125.47, 125.16, 124.96, 124.88, 123.95, 123.22, 122.99, 122.76, 111.73, 74.41,
74.13, 62.85, 54.28, 46.97, 36.57; IR (KBr, cm−1
max = 1475, 1534, 1599, 1732, 2998, 3100, 3265; [Anal.
δ
: 10.52 (s, 1H, N
), 7.61 (s, 1H, Ar–
), 7.26–7.21 (m, 1H, Ar–
H
), 7.91 (d, J = 7.6 Hz, 1H, Ar–
H
), 7.77 (d, J = 7.8 Hz, 1H, Ar–
H
),
H
H
), 7.56 (d, J = 8.1 Hz, 2H, Ar–
H
), 7.49 (d, J = 1.6 Hz, 1H,
H
H
H
), 6.52 (d, J = 8.7 Hz, 1H, Ar–
H
),
H
H
2), 3.74 (d, J = 10.8 Hz, 1H, CH), 3.37 (d,
H
), 3.27 (d, J = 11.5 Hz, 1H, CH2), 3.18 (dd, J = 11.5, 5.9 Hz, 1H, CH2); 13C-NMR
δ
: 196.30, 177.99, 143.03, 141.56, 140.05, 139.79, 138.87, 133.61, 133.30, 130.72,
)
ν
Calcd. for C29H20ClF3N2O2S2: C, 59.53; H, 3.45; N, 4.79; Found: C, 59.41; H, 3.55; N, 4.92]; LC/MS (ESI,
m/z): 585.20 [M+].

Molecules 2020, 25, 4671
13 of 16
(3S)-70-(Benzo[b]thiophen-2-yl)-60-(4-(trifluoromethyl)benzoyl)-30,60,70,7a’-tetrahydro-10H-spiro[indoline-
3,50-pyrrolo[1,2-c]thiazol]-2-one(IIj). Analog IIj was obtained using 2d (332 mg), isatin (3a) (147 mg),
◦
1
and thioproline (4a) (133 mg). Yield: 456 mg (0.83 mmol, 83%); m.p: 96 C; H-NMR (400 MHz,
DMSO-d6) : 10.37 (s, 1H, N ), 7.99–7.87 (m, 1H, Ar– ), 7.78 (d, J = 7.9 Hz, 1H, Ar– ), 7.69 (d,
J = 8.1 Hz, 2H Ar– ), 7.58 (s, 1H Ar– ), 7.49 (d, J = 8.0 Hz, 2H, Ar– ), 7.39 (d, J = 7.6 Hz, 1H, Ar– ),
7.33 (t, J = 9.3 Hz, 2H, Ar– ), 7.15 (t, J = 7.8 Hz, 1H, Ar– ), 6.97 (t, J = 7.7 Hz, 1H, Ar– ), 6.49 (d,
J = 7.8 Hz, 1H, Ar– ), 4.82 (d, J = 10.5 Hz, 1H, C CO), 4.36–4.18 (m, 2H, CH2), 3.73 (d, J = 10.3 Hz, 1H,
), 3.38 (d, J = 10.3 Hz, 1H, C
), 3.20 (d, J = 5.3 Hz, 2H, CH2); 13C-NMR (101 MHz, DMSO-d6)
196.41, 178.40, 143.35, 142.63, 140.08, 139.99, 138.85, 133.34, 133.03, 130.78, 128.83, 128.41, 126.00, 125.50,
125.15, 124.86, 123.94, 123.01, 122.79, 121.82, 110.28, 74.42, 73.81, 63.08, 54.07, 46.88, 36.56; IR (KBr, cm−1
δ
H
H
H
H
H
H
H
H
H
H
H
H
CH
H
δ:
)
ν
max = 1455, 1550, 1608, 1701, 2915, 3085, 3265; [Anal. Calcd. for C29H21F3N2O2S2: C, 63.26; H, 3.84; N,
5.09; Found: C, 63.15; H, 4.09; N, 5.23]; LC/MS (ESI, m/z): 551.20 [M+].
(3S)-10-(Benzo[b]thiophen-2-yl)-5-chloro-20-(4-(trifluoromethyl)benzoyl)-10,20,50,5a’,60,70,80,90,9a’,9b’-
decahydrospiro[indoline-3,30-pyrrolo[2,1-a]isoindol]-2-one (IIk). Analog IIk was prepared using 2d
(332 mg), 5-chloro-isatin (3b) (181 mg), and (2S,3aS,7aS)-octahydro-1H-indole-2-carboxylic acid (4b
(169 mg). Yield: 527 mg (0.85 mmol, 85%); m.p: 134 ◦C; 1H-NMR (400 MHz, DMSO-d6)
: 10.27 (s, 1H,
), 7.88 (d, J = 7.9 Hz, 1H, Ar– ), 7.74 (dd, J = 8.1, 3.6 Hz, 3H, Ar– ), 7.56 (d, J = 8.0 Hz, 2H, Ar– ),
7.49 (s, 1H, Ar– ), 7.43 (s, 1H, Ar– ), 7.35–7.24 (m, 2H, Ar– ), 7.20 (d, J = 9.1 Hz, 1H, Ar– ), 6.47 (d,
J = 8.3 Hz, 1H, Ar– ), 4.99 (d, J = 11.7 Hz, 1H, C CO), 4.41–4.30 (m, 1H, C ), 4.23 (t, J = 7.9 Hz, 1H,
), 3.17 (d, J = 3.7 Hz, 1H, C ), 2.12 (t, J = 8.3 Hz, 2H, CH2), 1.71 (dd, J = 10.6, 6.0 Hz, 1H, CH2), 1.51
(s, 2H, C 2), 1.32 (t, J = 12.4 Hz, 2H, C 2), 1.09 (d, J = 12.4 Hz, 1H, C 2), 1.00 (t, J = 12.5 Hz, 1H, C 2),
0.89 (t, J = 13.7 Hz, 1H, CH2), 0.70 (d, J = 13.7 Hz, 1H, CH2); 13C-NMR (126 MHz, DMSO-d6)
: 196.41,
)
δ
NH
H
H
H
H
H
H
H
H
H
H
C
H
H
H
H
H
H
δ
179.79, 143.70, 141.14, 140.07, 138.74, 133.36, 133.11, 130.06, 128.89, 128.21, 126.05, 126.02, 125.95, 125.70,
125.22, 124.92, 124.51, 123.69, 123.05, 122.81, 121.86, 111.44, 71.74, 70.98, 65.60, 57.33, 48.29, 41.85, 36.57,
27.97, 27.90, 24.93, 19.65; IR (KBr, cm−1
) νmax = 1450, 1485, 1535, 1623, 1710, 2905, 3035, 3355; [Anal.
Calcd. for C34H28ClF3N2O2S: C, 65.75; H, 4.54; N, 4.51; Found: C, 65.61; H, 4.63; N, 4.42]; LC/MS (ESI,
m/z): 621.20 [M+].
(3S)-70-(Benzo[b]thiophen-2-yl)-60-(4-bromobenzoyl)-5-chloro-30,60,70,7a’-tetrahydro-10H-spiro[indoline-
3,50-pyrrolo[1,2-c]thiazol]-2-one (IIl). Analog IIl was obtained using 2e (341 mg), 5-chloro-isatin (3b
(181 mg), and thioproline (4a) (133 mg). Yield 545 mg (0.92 mmol, 92%); m.p:108 C; H-NMR
)
◦
1
(400 MHz, DMSO-d6)
δ
: 10.55 (s, 1H, N
), 7.33 (q, J = 5.5 Hz, 3H, Ar–
CO), 4.23 (d, J = 11.0 Hz, 2H, C
H
), 7.91 (d, J = 7.9 Hz, 1H, Ar–
H
), 7.77 (d, J = 7.9 Hz, 1H, Ar–
H
),
7.60–7.46 (m, 4H, Ar–
H
H), 7.28–7.21 (m, 2H, Ar–
H
), 6.56 (d, J = 8.3 Hz, 1H,
Ar–
3.40 (s, 1H, C
DMSO-d6) : 195.67, 178.03, 143.06, 141.49, 139.99, 138.79, 135.58, 132.27, 130.58, 130.06, 128.56, 128.40,
125.71, 125.13, 124.97, 124.84, 123.91, 123.14, 122.96, 111.68, 74.33, 74.21, 62.37, 54.28, 47.03, 36.52; IR
H
), 4.75 (d, J = 11.1 Hz, 1H, C
H
H
2), 3.74 (d, J = 10.7 Hz, 1H, C
H
),
H
), 3.29–3.22 (m, 1H, CH2), 3.17 (dd, J = 11.3, 5.7 Hz, 1H, CH2); 13C-NMR (126 MHz,
δ
(KBr, cm−1
) νmax = 1490, 1550, 1625, 1725, 2915, 3050, 3250; [Anal. Calcd. for C28H20BrClN2O2S2: C,
56.43; H, 3.38; N, 4.70; Found: C, 56.33; H, 3.49; N, 4.91]; LC/MS (ESI, m/z): 595.20 [M+].
(3S)-70-(Benzo[b]thiophen-2-yl)-60-(4-bromobenzoyl)-30,60,70,7a’-tetrahydro-10H-spiro[indoline-3,50-pyrrolo
[1,2-c]thiazol]-2-one(IIm). Analog IIm was obtained using 2e (341 mg), isatin (3a) (147 mg), and thioproline
(
4a) (133 mg). Yield: 492 mg (0.88 mmol, 88%); m.p:100 ◦C; 1H-NMR (400 MHz, DMSO-d6)
δ
: 10.41 (s,
), 7.54 (t, J = 4.4 Hz, 3H, Ar– ), 7.33
), 6.96 (s, 1H, Ar– ), 6.53 (d, J = 7.9 Hz, 1H,
CO), 4.33–4.18 (m, 2H, C 2), 3.72 (d, J = 10.3 Hz, 1H, C ), 3.37 (s, 1H,
2); 13C-NMR (126 MHz, DMSO-d6)
195.76, 178.45, 143.40, 142.55, 140.02,
138.78, 135.76, 132.09, 130.62, 129.97, 128.44, 128.21, 125.10, 124.79, 123.88, 123.03, 122.94, 122.89, 121.70,
110.23, 74.33, 73.89, 62.55, 54.05, 46.95, 36.50; IR (KBr, cm−1
νmax = 1480, 1510, 1608, 1720, 2910, 3055,
1H, N
(ddd, J = 29.2, 17.6, 8.3 Hz, 5H, Ar–
Ar– ), 4.73 (d, J = 10.7 Hz, 1H, C
), 3.19 (d, J = 4.8 Hz, 2H, C
H
), 7.91 (d, J = 7.8 Hz, 1H, Ar–
H
), 7.77 (d, J = 7.6 Hz, 1H, Ar–
H
H
H
), 7.15 (s, 1H, Ar–
H
H
H
H
H
H
C
H
H
δ
)

Molecules 2020, 25, 4671
14 of 16
3245; [Anal. Calcd. for C28H21BrN2O2S2: C, 59.89; H, 3.77; N, 4.99; Found: C, 60.03; H, 3.65; N, 5.08];
LC/MS (ESI, m/z): 561.20 [M+].
(3S)-10-(Benzo[b]thiophen-2-yl)-20-(4-bromobenzoyl)-5-chloro-10,20,50,5a’,60,70,80,90,9a’,9b’-decahydrospiro
[indoline-3,30-pyrrolo[2,1-a]isoindol]-2-one(IIn). Analog IIn was prepared using 2e (341 mg), 5-chloro-isatin
(
3b) (181 mg), and (2S,3aS,7aS)-octahydro-1H-indole-2-carboxylic acid (4b) (169 mg) in equimolar
amounts. Yield: 567 mg (0.9 mmol, 90%); m.p:136 ◦C; 1H-NMR (400 MHz, DMSO-d6)
: 10.31 (s, 1H,
), 7.87 (d, J = 7.9 Hz, 1H, Ar– ), 7.74 (d, J = 7.6 Hz, 1H, Ar– ), 7.58 (d, J = 8.2 Hz, 2H, Ar– ), 7.46 (s,
1H, Ar– ), 7.39 (s, 1H, Ar– ), 7.30 (dd, J = 19.2, 7.8 Hz, 4H, Ar– ), 7.19 (d, J = 8.6 Hz, 1H, Ar– ),
6.52 (d, J = 8.2 Hz, 1H, Ar– ), 4.90 (d, J = 11.7 Hz, 1H, C CO), 4.32 (t, J = 10.8 Hz, 1H, C ), 4.22 (t,
J = 8.8 Hz, 1H, C ), 3.16 (d, J = 3.9 Hz, 1H, C ), 2.11 (dd, J = 9.9, 4.2 Hz, 2H, CH2), 1.70 (dd, J = 11.0,
δ
NH
H
H
H
H
H
H
H
H
H
H
H
H
6.0 Hz, 1H, CH2), 1.49 (d, J = 12.6 Hz, 2H, CH2), 1.32 (t, J = 11.2 Hz, 2H, CH2), 1.09 (d, J = 12.6 Hz,
1H, CH2), 0.98 (d, J = 12.9 Hz, 1H, CH2), 0.86 (d, J = 13.2 Hz, 1H, CH2), 0.70 (d, J = 13.7 Hz, 1H, CH2);
13C-NMR (126 MHz, DMSO-d6)
δ
: 195.86, 179.87, 143.79, 141.13, 140.04, 138.72, 135.91, 132.14, 130.06,
129.96, 128.25, 125.85, 125.77, 124.92, 124.49, 123.68, 122.81, 121.83, 111.43, 71.84, 70.92, 65.26, 57.33, 48.39,
41.86, 36.61, 27.98, 27.90, 24.94, 19.65; IR (KBr, cm−1
max = 1495, 1615, 1712, 2920, 3245; [Anal. Calcd.
)
ν
for C33H28BrClN2O2S: C, 62.71; H, 4.47; N, 4.43; Found: C, 62.85; H, 4.56; N, 4.53]; LC/MS (ESI, m/z):
633.20 [M+].
4. Conclusions
A series of novel spiro-heterocycles incorporating the benzo[b]thiophene motif were prepared and
their AChE inhibitory activity was evaluated. The results revealed that among the studied compounds,
analog IIc was the most active AChE inhibitor with an IC50 value of 20,840 µ
M L−1. Molecular
docking studies were also performed to elucidate the structural features and interactions responsible
for the inhibitory potential of these compounds against the target protein. The docking results further
confirmed the experimental findings and provided significant information on the binding mechanism
of the novel analogues to the AChE enzyme.
Supplementary Materials: The following are available online: experimental protocol for the AChE assay; Copies
of the spectrum; and IC50 diagram of the AChE assay.
Author Contributions: Conceptualization, A.B.; data curation, M.S.A., M.S.I., A.S.A., S.A. (Sajda Ashraf) and
Z.U.-H.; formal analysis, S.A. (Saeed Alshahrani), M.A., M.S.A., M.S.I. and A.S.A.; funding acquisition, A.B.;
investigation, A.B., S.A. (Saeed Alshahrani), M.A., M.S.I. and A.S.A.; methodology, A.B., S.A. (Saeed Alshahrani),
M.A., M.S.I. and A.S.A.; software, S.A. (Sajda Ashraf) and Z.U.-H.; supervision, A.B., A.M.A.-M. and Z.U.-H.;
validation, S.A. (Sajda Ashraf) and Z.U.-H.; Visualization, A.M.A.-M.; writing—original draft, A.B. and Z.U.-H.;
writing—review and editing, A.B. All authors have read and agreed to the published version of the manuscript.
Funding: This research was funded by the King Saud University, grant number (RGP-257).
Acknowledgments: The authors would like to extend their sincere appreciation to the Deanship of Scientific
Research at King Saud University for providing funding to this Research group NO (RGP-257). The authors thank
the Deanship of Scientific Research and RSSU at King Saud University for their technical support.
Conflicts of Interest: The authors declare no conflict of interest.
References
1.
2.
3.
Schelterns, P.; Feldman, H. Treatment of Alzheimer’s disease; current status and new perspectives.
Lancet Neurol. 2003, 2, 539.
Prince, M.; Wimo, A.; Guerchet, M.; Ali, G.C.; Wu, Y.T.; Prina, M. Alzheimer’s Disease International; King’s
College: London, UK, 2015.
Wang, Z.; Wang, Y.; Li, W.; Mao, F.; Sun, Y.; Huang, L.; Li, X. Design, synthesis, and evaluation of
multitarget-directed selenium-containing clioquinol derivatives for the treatment of Alzheimer’s disease.
ACS Chem. Neurosci. 2014, 5, 952. [CrossRef]
4.
Savelieff, M.G.; Lee, S.; Liu, Y.; Lim, M.H. Untangling amyloid-
ACS Chem. Biol. 2013, 8, 856. [CrossRef]
β, tau, and metals in Alzheimer’s disease.

Molecules 2020, 25, 4671
15 of 16
5.
6.
7.
Choi, S.S.; Lee, S.R.; Kim, S.U.; Lee, H.J. Alzheimer’s disease and stem cell therapy. Exp. Neurol. 2014, 23, 45.
[CrossRef]
Giacobini, E. Selective inhibitors of butyrylcholinesterase: A valid alternative for therapy of Alzheimer’s
disease? Drugs Aging 2001, 18, 891. [CrossRef] [PubMed]
Kia, Y.; Osman, H.; Kumar, R.S.; Murugaiyah, V.; Basiri, A.; Perumal, S.; Razak, I.A. A facile chemo-, regio-and
stereoselective synthesis and cholinesterase inhibitory activity of spirooxindole–pyrrolizine–piperidine
hybrids. Bioorganic Med. Chem. Lett. 2013, 23, 2979. [CrossRef] [PubMed]
8.
9.
Kia, Y.; Osman, H.; Kumar, R.S.; Basiri, A.; Murugaiyah, V. Synthesis and discovery of highly functionalized
mono- and bis-spiro-pyrrolidines as potent cholinesterase enzyme inhibitors. Bioorganic Med. Chem. Lett.
2014, 24, 1815. [CrossRef] [PubMed]
Kia, Y.; Osman, H.; Kumar, R.S.; Murugaiyah, V.; Basiri, A.; Perumal, S.; Wahab, H.A.; Bing, C.S. Synthesis
and discovery of novel piperidone-grafted mono- and bis-spirooxindole- hexahydropyrrolizines as potent
cholinesterase inhibitors. Bioorganic Med. Chem. 2013, 21, 1696. [CrossRef] [PubMed]
10. Kia, Y.; Osman, H.; Kumar, R.S.; Basiri, A.; Murugaiyah, V. Ionic liquid mediated synthesis of mono- and
bis-spirooxindole-hexahydropyrrolidines as cholinesterase inhibitors and their molecular docking studies.
Bioorg. Med. Chem. 2014, 22, 1318. [CrossRef]
11. Arumugam, N.; Almansour, A.I.; Kumar, R.S.; Kotresha, D.; Saiswaroop, R.; Venketesh, S.
Dispiropyrrolidinyl-piperidone embedded indeno [1,2-b] quinoxaline heterocyclic hybrids: Synthesis,
cholinesterase inhibitory activity and their molecular docking simulation. Bioorg. Med. Chem. 2019, 27, 2621.
[CrossRef]
12. Barakat, A.; Soliman, S.M.; Alshahrani, S.; Islam, M.S.; Ali, M.; Al-Majid, A.M.; Yousuf, S. Synthesis, X-ray
Single Crystal, Conformational Analysis and Cholinesterase Inhibitory Activity of a New Spiropyrrolidine
Scaffold Tethered Benzo [b] Thiophene Analogue. Crystals 2020, 10, 120. [CrossRef]
13. Goyal, D.; Kaur, A.; Goyal, B. Benzofuran and Indole: Promising Scaffolds for Drug Development in
Alzheimer’s Disease. ChemMedChem 2018, 13, 1275. [CrossRef] [PubMed]
14. Kumar, R.S.; Ali, M.A.; Osman, H.; Ismail, R.; Choon, T.S.; Yoon, Y.K.; Wei, A.C.; Pandian, S.; Manogaran, E.
Synthesis and discovery of novel hexacyclic cage compounds as inhibitors of acetylcholinesterase. Bioorganic
Med. Chem. Lett. 2011, 21, 3997. [CrossRef] [PubMed]
15. Lotfy, G.; El Ashry, E.S.H.; Said, M.M.; El Tamany, E.S.H.; Aziz, Y.M.A.; Al-Dhfyan, A.; Al-Majid, A.M.;
Barakat, A. Regio-and stereoselective synthesis of new spirooxindoles via 1,3-dipolar cycloaddition reaction:
Anticancer and molecular docking studies. J. Photochem. Photobiol. B Biol. 2018, 180, 98. [CrossRef] [PubMed]
16. Lotfy, G.; Said, M.M.; El Ashry, E.S.H.; El Tamany, E.S.H.; Al-Dhfyan, A.; Aziz, Y.M.A.; Barakat, A. Synthesis
of new spirooxindole-pyrrolothiazole derivatives: Anti-cancer activity and molecular docking. Biorg. Med.
Chem. 2017, 25, 1514. [CrossRef]
17. Lotfy, G.; Aziz, Y.M.A.; Said, M.M.; El Ashry, E.S.H.; El Tamany, E.S.H.; Barakat, A.; Ghabbour, H.A.;
Yousuf, S.; Ul-Haq, Z.; Choudhary, M.I. Synthesis of oxindole analogues, biological activity, and in silico
studies. ChemistrySelect 2019, 4, 10510. [CrossRef]
18. Keri, R.S.; Chand, K.; Budagumpi, S.; Somappa, S.B.; Patil, S.A.; Nagaraja, B.M. An overview of benzo [b]
thiophene-based medicinal chemistry. Eur. J. Med. Chem. 2017, 138, 1002–1033. [CrossRef]
19. Anbar, H.S.; El-Gamal, R.; Ullah, S.; Zaraei, S.O.; al-Rashida, M.; Zaib, S.; Pelletier, J.; Sevigny, J.; Iqbal, J.;
El-Gamal, M.I. Evaluation of sulfonate and sulfamate derivatives possessing benzofuran or benzothiophene
nucleus as inhibitors of nucleotide pyrophosphatases/phosphodiesterases and anticancer agents. Bioorganic
Chem. 2020, 104, 104305. [CrossRef]
20. Wang, Z.; Liu, Y.; Zhang, J.; Ullah, S.; Kang, N.; Zhao, Y.; Zhou, H. Benzothiophene-2-carboxamide derivatives
as SENPs inhibitors with selectivity within SENPs family. Eur. J. Med. Chem. 2020, 204, 112553. [CrossRef]
21. Pieroni, M.; Azzali, E.; Basilico, N.; Parapini, S.; Zolkiewski, M.; Beato, C.; Annunziato, G.;
Bruno, A.; Vacondio, F.; Costantino, G. Accepting the Invitation to Open Innovation in Malaria Drug
Discovery: Synthesis, Biological Evaluation, and Investigation on the Structure–Activity Relationships of
Benzo[b]thiophene-2-carboxamides as Antimalarial Agents. J. Med. Chem. 2017, 60, 1959–1970. [CrossRef]
22. Qin, Z.; Kastrati, I.; Chandrasena, R.E.P.; Liu, H.; Yao, P.; Petukhov, P.A.; Bolton, J.L.; Thatcher, G.R.J.
Benzothiophene Selective Estrogen Receptor Modulators with Modulated Oxidative Activity and Receptor
Affinity. J. Med. Chem. 2007, 50, 2682–2692. [CrossRef] [PubMed]

Molecules 2020, 25, 4671
16 of 16
23. Konstantinovic´, J.; Videnovic´, M.; Srbljanovic´, J.; Djurkovic´-Djakovic´, O.; Bogojevic´, K.; Sciotti, R.; Šolaja, B.
Antimalarials with Benzothiophene Moieties as Aminoquinoline Partners. Molecules 2017, 22, 343. [CrossRef]
24. El-Miligy, M.M.M.; Hazzaa, A.A.; El-Messmary, H.; Nassra, R.A.; El-Hawash, S.A.M. New hybrid molecules
combining benzothiophene or benzofuran with rhodanine as dual COX-1/2 and 5-LOX inhibitors: Synthesis,
biological evaluation and docking study. Bioorganic Chem. 2017, 72, 102–115. [CrossRef] [PubMed]
25. Chen, W.; Dong, G.; He, S.; Xu, T.; Wang, X.; Liu, N.; Zhang, W.; Miao, C.; Sheng, C. Identification of
benzothiophene amides as potent inhibitors of human nicotinamide phosphoribosyltransferase. Bioorganic
Med. Chem. Lett. 2016, 26, 765–768. [CrossRef] [PubMed]
26. Aleksic´, M.; Bertoša, B.; Nhili, R.; Depauw, S.; Martin-Kleiner, I.; David-Cordonnier, M.-H.; Tomic´, S.; Kralj, M.;
Karminski-Zamola, G. Anilides and quinolones with nitrogen-bearing substituents from benzothiophene
and thienothiophene series: Synthesis, photochemical synthesis, cytostatic evaluation, 3D-derived QSAR
analysis and DNA-binding properties. Eur. J. Med. Chem. 2014, 71, 267–281. [CrossRef]
27. Altowyan, M.S.; Barakat, A.; Al-Majid, A.M.; Al-Ghulikah, H.A. Spiroindolone Analogues as Potential
Hypoglycemic with Dual Inhibitory Activity on
[CrossRef]
α-Amylase and α-Glucosidase. Molecules 2019, 24, 2342.
28. Altowyan, M.S.; Barakat, A.; Al-Majid, A.M.; Al-Ghulikah, H.A. New Spiroindolone Bearing Benzofuran
Moiety as High Effective and Selective for COX-1 Inhibitor with TNF-
Biol. Sci. 2020, 27, 1208. [CrossRef]
α, and IL-6 inhibitory activity. Saudi J.
29. Barakat, A.; Islam, M.S.; Ghawas, H.M.; Al-Majid, A.M.; El-Senduny, F.F.; Badria, F.A.; Elshaier, Y.A.M.;
Ghabbour, H.A. Substituted spirooxindole derivatives as potent anticancer agents through inhibition of
phosphodiesterase 1. RSC Adv. 2018, 8, 14335. [CrossRef]
30. Islam, M.S.; Ghawas, H.M.; El-Senduny, F.F.; Al-Majid, A.M.; Elshaier, Y.A.; Badria, F.A.; Barakat, A. Synthesis
of new thiazolo-pyrrolidine–(spirooxindole) tethered to 3-acylindole as anticancer agents. Bioorganic Chem.
2019, 82, 423. [CrossRef]
31. Al-Majid, A.M.; Soliman, S.M.; Haukka, M.; Ali, M.; Islam, M.S.; Shaik, M.R.; Barakat, A. Design, Construction,
and Characterization of a New Regioisomer and Diastereomer Material Based on the Spirooxindole Scaffold
Incorporating a Sulphone Function. Symmetry 2020, 12, 1337. [CrossRef]
32. Zhou, L.M.; Qu, R.Y.; Yang, G.F. An overview of spirooxindole as a promising scaffold for novel drug
discovery. Expert Opin. Drug Discov. 2020, 15, 603–625. [CrossRef] [PubMed]
33. Almansour, A.I.; Arumugam, N.; Kumar, R.S.; Kotresha, D.; Manohar, T.S.; Venketesh, S. Design, synthesis
and cholinesterase inhibitory activity of novel spiropyrrolidine tethered imidazole heterocyclic hybrids.
Bioorganic Med. Chem. Lett. 2020, 30, 126789. [CrossRef] [PubMed]
34. Kumar, R.S.; Almansour, A.I.; Arumugam, N.; Althomili, D.M.Q.; Altaf, M.; Basiri, A.; Manohar, T.S.;
Venketesh, S. Ionic liquid-enabled synthesis, cholinesterase inhibitory activity, and molecular docking study
of highly functionalized tetrasubstituted pyrrolidines. Bioorganic Chem. 2018, 77, 263–268. [CrossRef]
[PubMed]
35. Lilienfeld, S. Galantamine—A novel cholinergic drug with a unique dual mode of action for the treatment of
patients with Alzheimer’s disease. CNS Drug Rev. 2002, 8, 159–176. [CrossRef]
36. Shang, Y.; Jie, X.; Zhou, J.; Hu, P.; Huang, S.; Su, W. Pd-Catalyzed C–H Olefination of (Hetero) Arenes by
Using Saturated Ketones as an Olefin Source. Angew. Chem. Int. 2013, 52, 1299. [CrossRef]
37. Wu, C.; Yue, G.; Nielsen, C.D.T.; Xu, K.; Hirao, H.; Zhou, J. Asymmetric conjugate addition of organoboron
reagents to common enones using copper catalysts. J. Am. Chem. Soc. 2016, 138, 742. [CrossRef]
Sample Availability: Samples of the compounds IIa–n are available from the authors.
©
2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).