Antiproliferative activity and possible mechanism of action of certain 5-methoxyindole tethered C-5 functionalized isatins

Article abstract of DOI:10.2147/DDDT.S208241

Background: Cancer is one of the most dreaded human diseases, that has become an ever-increasing health problem and is a prime cause of death globally. The potential antiproliferative activity of certain indole–isatin molecular hybrids 5a-w was evaluated in vitro against three human cancer cell lines. Methods: Standard protocols were adopted to examine the antiproliferative potential and mechanisms of compounds 5a-w. Western blot analysis was carried out on compound 5o. Results: Compounds 5a-w demonstrated in vitro antiproliferative activity in the range of 22.6–97.8%, with compounds 5o and 5w being the most active antiproliferative compounds with IC₅₀ values of 1.69 and 1.91 μM, which is fivefold and fourfold more potent than sunitinib (IC₅₀=8.11 μM), respectively. Compound 5o was selected for in-depth pharmacological testing to understand its possible mechanism of antiproliferative activity. It caused a lengthening of the G1 phase and a reduction in the S and G2/M phases of the cell cycle and had an IC₅₀ value of 10.4 μM with the resistant NCI-H69AR cancer cell line. Moreover, compound 5o significantly decreased the amount of phosphorylated Rb protein in a dose-dependent fashion, which was confirmed via Western blot analysis. Conclusion: The current investigation highlighted the potential antiproliferative activity of compounds 5a-w as well as the antiproliferative profile of compound 5o. These compounds can be harnessed as new lead antiproliferatives in the preclinical studies of cancer chemotherapy.

Full text of DOI:10.2147/DDDT.S208241

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O R I G I N A L R E S E A R C H
Antiproliferative activity and possible mechanism
of action of certain 5-methoxyindole tethered C-5
functionalized isatins
This article was published in the following Dove Press journal:
Drug Design, Development and Therapy
Maha S Almutairi1
Background: Cancer is one of the most dreaded human diseases, that has become an ever-
Eman S Hassan2
increasing health problem and is a prime cause of death globally. The potential antiproli-
Adam B Keeton3
ferative activity of certain indole–isatin molecular hybrids 5a-w was evaluated in vitro
Gary A Piazza3
against three human cancer cell lines.
Methods: Standard protocols were adopted to examine the antiproliferative potential and
Ali S Abdelhameed1
mechanisms of compounds 5a-w. Western blot analysis was carried out on compound 5o.
Mohamed I Attia1,4
Results: Compounds 5a-w demonstrated in vitro antiproliferative activity in the range of
1Department of Pharmaceutical
Chemistry, College of Pharmacy, King
22.6–97.8%, with compounds 5o and 5w being the most active antiproliferative compounds
with IC50 values of 1.69 and 1.91 µM, which is fivefold and fourfold more potent than
Saud University, Riyadh 11451, Saudi
Arabia; 2Department of Medical
Laboratory Sciences, Al-Ghad
sunitinib (IC50=8.11 µM), respectively. Compound 5o was selected for in-depth pharmaco-
logical testing to understand its possible mechanism of antiproliferative activity. It caused a
lengthening of the G1 phase and a reduction in the S and G2/M phases of the cell cycle and
had an IC50 value of 10.4 μM with the resistant NCI-H69AR cancer cell line. Moreover,
compound 5o significantly decreased the amount of phosphorylated Rb protein in a dose-
dependent fashion, which was confirmed via Western blot analysis.
International Medical Sciences College,
Female Section, Riyadh 13315, Saudi
Arabia; 3Department of Oncologic
Sciences and Pharmacology, Drug
Discovery Research Center, Mitchell
Cancer Institute, University of South
Alabama, Mobile, AL 36604-1405, USA;
4Medicinal and Pharmaceutical Chemistry
Department, Pharmaceutical and Drug
Industries Research Division, National
Research Centre (ID: 60014618), Giza
12622, Egypt
Conclusion: The current investigation highlighted the potential antiproliferative activity of
compounds 5a-w as well as the antiproliferative profile of compound 5o. These compounds
can be harnessed as new lead antiproliferatives in the preclinical studies of cancer
chemotherapy.
Keywords: isatin, indole, synthesis, antiproliferative, apoptosis
Introduction
Cancer is one of the most terrifying diseases of humanity and has become a
fundamental health problem and a principal cause of death globally. One in four
deaths in the United States is a result of cancer.1 More than ten million new cases of
cancer occur every year, approximately half of which occur in developed countries,
with the disease causing more than six million deaths every year.2–4 A molecularly
targeted approach has recently been utilized for the management of disseminated
cancer which depends on the study of oncogenes and tumor suppressors which are
involved in the emergence of human cancers.5 Consequently, there has been an
advancement in the specificity of cancer management, progressing from the use of
general cytotoxic agents such as nitrogen mustard in the 1940s, the development of
chemotherapeutic agents such as anthracyclines and Vinca alkaloids from natural
resources in the 1960s and finally the use of specific monoclonal antibodies6 and
Correspondence: Mohamed I Attia
Department of Pharmaceutical
Chemistry, College of Pharmacy, King
Saud University, P.O. Box. 2457, Riyadh
11451, Saudi Arabia
Tel +966 1 467 7337
Fax +966 1 467 6220
Email mattia@ksu.edu.sa
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php and incorporate the Creative Commons Attribution – Non Commercial (unported, v3.0) License (http://creativecommons.org/licenses/by-nc/3.0/). By accessing the
work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. For
permission for commercial use of this work, please see paragraphs 4.2 and 5 of our Terms (https://www.dovepress.com/terms.php).
http://doi.org/10.2147/DDDT.S208241

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specific chemotherapeutic agents which inhibit protein
tyrosine kinases (PTKs) as advanced approaches.7–9
These targeted chemotherapeutic agents usually attenuate
signaling pathways which control the cancer cell cycle and
alter its microenvironment, blocking tumor cell prolifera-
tion, cell apoptosis and/or hindering tumor mass growth.10
These developments led to a reduction of anticancer side
effects and ameliorated the response rate. Therefore, the
study of the mechanisms by which cancers resist che-
motherapeutic agents gave rise to a deep understanding
of the reasons for the failure of cancer therapies.
Indole (I, Figure 1) is a privileged bicyclic structure which
was first synthesized in 1866. The indole scaffold is incorpo-
rated into a large number of biologically active molecules
endowed with a wide range of bioactivities and is naturally
occurring in Vinca and ergot alkaloids, fungal metabolites and
marines.11 In recent years, indole and its functionalized deri-
vatives have been embedded in myriad bioactive pharmaceu-
ticals including anti-inflammatories, analgesics, antimicrobials
and antitumors.12–18 Furthermore, 5-methoxyindole is the fun-
damental fragment in the natural hormone melatonin (MLT, II,
Figure 1). MLT and its derivatives have a broad spectrum of
X
CH3
R
N
HN
O
O
O
H3CO
H3CO
HN N
O
O
N
H
N
H
N
H
N
H
I
II
III
5a-w
Figure 1 Chemical structures of compounds I-III and 5a-w.
60
40
20
0
24h
48h
100
50
0
24h
48h
-20
-50
0.01
0.1
1
10
100
0.01
0.1
1
10
100
Concentration (µM)
Concentration (µM)
5o
Sunitinib
100%
90%
80%
70%
60%
50%
40%
30%
20%
10%
0%
G2/M
S
G1
DMSO
5o
Sunitinib
Figure 2 Cell cycle influences of compound 5o after incubation for 24 and 48 hrs.
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Almutairi et al
pharmaceutical applications, particularly for the treatment of yields. The analytical data of compounds 5a-w are pre-
headache, depression and sleep disorders, and for the manage- viously documented.32,33
ment of certain types of cancer.19–21
On the other hand, isatin (2,3-dioxindole, III) is con-
Pharmacological evaluation
sidered an oxidized form of indole and has been recognized
to be an endogenous multifunctional molecule in human
beings and other mammals.22 The special electronic proper-
ties of isatin along with its proper molecular size give rise to
several different valuable biological characteristics.
Therefore, isatin was embedded into the backbone of var-
ious bioactive molecules including anticonvulsants,23
antifungals,24 antibacterials,25 anti-HIV agents24,26 and
anticancer agents.27–31
Pharmacological assessment of the title compounds including
antiproliferative activity, selectivity, cell cycle effects and
quantitative immunofluorescence of 5a-w was performed
with previously documented methods.29 Western blot analysis
of total cellular proteins enabled detection of P-Rb and
GAPDH (glyceraldehyde-3-phosphate dehydrogenase) using
antibodies obtained from Cell Signaling Technology (Boston,
MA, USA). Western blots were imaged by direct imaging of
chemiluminescent blots (ChemiDoc Imaging System;
BioRad, Hercules, CA, USA). Quantitation was carried out
using NIH ImageJ public domain image analysis software.
The cell lines were purchased commercially from the
American Type Culture Collection (ATCC).
According to the aforementioned premises, it was our
interest to prepare the indole–isatin conjugates 5a-w as
hybrid molecules tailored from indole and isatin pharma-
cophore fragments for biological evaluation. The isatin
moiety of the target conjugates 5a-w is functionalized on
their C-5 position and bears various N-aralkyl substitutions
that alter the electronic and lipophilic environment, allow-
ing us to explore their impact on the biological activity of
compounds 5a-w. Compounds 5a-wdisplayed moderate
antimicrobial potential.32,33 The current report deals with
the assessment of their in vitro antiproliferative potential.
The most active antiproliferative candidates were sub-
jected to deep pharmacological testing to gain insight
into the possible mechanism of their antiproliferative
activity.
Results and discussion
Chemistry
Compounds 5a-w were prepared as illustrated in
Scheme 1. Thus, the commercially available 5-methoxy
indole-2-carboxylic acid (1) was esterified in absolute
methanol and a catalytic amount of concentrated sulfuric
acid, followed by hydrazinolysis, to prepare the hydrazide
3. Subsequently, compound 3 was reacted with the isatin
derivatives 4a-n33 to achieve the respective title com-
pounds 5a-w in moderate yields.
Materials and methods
Chemistry
5-Methoxy-1H-indole-2-carbohydrazide (3) – The acid
hydrazide 3 was prepared from the corresponding ester 234
using the documented method.32 It has a melting point (m.p.)
of 266–268°C.
Pharmacological investigations
Antiproliferative activity
The isatin nucleus is incorporated into various anticancer
candidates.28,29,31,35,36 The preliminary antiproliferative
potential of compounds 5a-w was tested using A-549
(lung), HT-29 (colon) and ZR-75 (breast) human cancer
cell lines, and the obtained data are presented in Table 1.
Sunitinib was used against the same human cancer cell
lines as a reference drug for the experiments. The results
are expressed as an average percent growth inhibition at
General method for the preparation of
5-methoxy-1H-indole-2-carbo hydrazide
derivatives 5a-w
Glacial acetic acid (catalytic amount) was added to a 30 µM concentration for each compound tested in quad-
mixture of the proper isatin derivative 4a-n (1 mmol) ruplicate. The title compounds 5a-w exhibited an average
and the acid hydrazide 3 (1 mmol) in absolute ethyl growth inhibition of 22.6–97.8% in the antiproliferative
alcohol (15 mL). The reaction mixture was then stirred assay against the tested human cancer cell lines, except for
under reflux for 4 hrs. The precipitated solid was filtered compound 5e, which stimulated the growth of the ZR-75
while hot, and the obtained solid was recrystallized from cell line. It seems that the N-unsubstituted isatin (com-
an ethyl alcohol/dimethylformamide mixture (3:1) to fur- pounds 5b and 5c), N-methyl (compounds 5g-i), N-benzyl
nish the corresponding compounds 5a-w in 43–94% (compound 5o) or N-phenyl (compound 5w) moieties are
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H3CO
H3CO
H3CO
OCH3
O
NHNH2
OH
O
i
ii
N
H
N
H
O
N
H
1
2
3
O
X
O
iii
N
R
4a-n
X
R
N
O
H3CO
HN N
N
O
H
5a-w
Scheme 1 Synthesis of compounds 5a-w. Reagents and conditions: (I) absolute methanol, H2SO4 (few drops), reflux, 4 hrs; (ii) absolute methanol, H2N-NH2.H2O, reflux, 2
hrs; (iii) absolute ethanol, acetic acid (few drops), reflux, 4 hrs.
Compound No.
X
R
Compound No.
X
R
5a
5b
5c
5d
5e
5f
H
H
5m
5n
5o
5p
5q
5r
Cl
F
C6H5-CH2
Br
H
C6H5-CH2
Cl
H
OCH3
H
C6H5-CH2
F
H
4-F-C6H4-CH2
4-F-C6H4-CH2
4-F-C6H4-CH2
4-F-C6H4-CH2
4-Cl-C6H4-CH2
4-CN-C6H4-CH2
4-CH3-C6H4-CH2
C6H5
OCH3
H
H
Br
Cl
F
CH3
CH3
CH3
CH3
CH3
C6H5-CH2
C6H5-CH2
5g
5h
5i
Br
5s
Cl
5t
H
F
5u
5v
5w
H
5j
OCH3
H
H
5k
5l
H
Br
the preferred fragments at the isatin nitrogen, as they phenylisatin moiety) with IC50 values of 1.69 and 1.91 µM,
respectively, which are about fivefold and fourfold more
potent than sunitinib (IC50=8.11 µM). Therefore, detailed
pharmacological studies were carried on compound 5o, aim-
ing to gain insight into the integrated pharmacological profile
of this compound, as a representative for compounds 5a-w.
induced average growth inhibition of 96.0, 91.3, 94.5,
95.3, 91.8, 97.8 and 97.6%, respectively. Also, halogen
substitution at isatin C-5 is the favored substituent, except
for compounds 5o and 5w which bear methoxy and hydro-
gen functionalities, respectively.
Compounds displaying an average growth inhibition of
more than 90% toward ZR-75, HT-29, and A-549 cell lines
were subjected to median growth inhibitory concentration
(IC50) determination. Table 2 illustrates the IC50 values of
compounds 5b, 5c, 5g-i, 5o, 5w and sunitinib toward ZR-75,
HT-29 and A-549 cell lines. The most active candidates are
Caspase 3/7 activity
The A-549 cell line was utilized to assess the apoptosis-
inducing potential of compound 5o. Activity assessment of
compound 5o was carried out at concentrations equal to its
IC50 for growth inhibition and at threefold above this
5o (bearing an N-benzylisatin moiety) and 5w (bearing an N- concentration over a 2–48 hr time course. Compound 5o
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Table 1 In vitro antiproliferative potential of compounds 5a-w and sunitinib against HT-29, ZR-75 and A-549 cell lines
Compound No.
HT-29
ZR-75
A-549
Average growth inhibition %
5a
5b
5c
10.2 6.3
96.2 4.8
89.8 2.4
52.5 9.4
−7.3 30.1
82.5 11.9
93.5 2.5
92.3 2.1
92.9 0.9
32.2 19.9
77.4 4.4
70.2 3.8
69.9 6.0
65.9 3.0
97.0 2.8
80.8 6.3
45.1 5.6
40.3 10.2
41.6 5.3
84.1 7.2
79.9 3.9
84.4 9.1
98.4 1.3
59.5 2.3
43.2 19.3
96.6 1.4
88.8 7.0
77.7 7.5
−37.4 8.9
63.8 7.4
94.4 3.9
96.1 2.1
86.1 6.1
54.6 22.3
46.0 20.1
53.8 16.5
75.5 5.8
73.7 3.7
96.5 2.8
63.2 11.9
35.9 2.1
8.1 15.7
52.7 12.0
79.2 8.8
82.6 12.4
54.8 16.4
94.8 2.1
90.7 4.5
14.4 5.2
95.2 6.2
95.2 3.3
62.5 13.4
22.2 8.4
89.2 6.4
95.5 2.4
97.5 1.0
96.5 3.8
96.3 3.4
80.9 5.3
68.5 4.1
70.1 4.0
58.0 7.6
100.0 0.0
81.9 2.2
55.8 2.8
56.1 5.7
54.8 4.5
100.0 0.0
97.6 2.8
89.2 4.8
99.5 1.1
85.7 2.7
22.6
96.0
91.3
64.2
−7.5
78.5
94.5
95.3
91.8
61.0
68.1
64.2
71.8
65.9
97.8
75.3
45.6
34.8
49.7
87.8
86.7
76.1
97.6
78.7
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
5q
5r
5s
5t
5u
5v
5w
Sunitinib
Table 2 Antiproliferative inhibitory concentration 50% (IC50) values of compounds 5b, 5c, 5g-i, 5o, 5w and sunitinib toward A-549,
ZR-75 and HT-29 cell lines
Compound No.
IC50 (µM)
A-549
Average IC50 (µM)
ZR-75
HT-29
5b
23.8
15.0 10.36
16.8 23.36
5.29 1.08
5.39 1.46
21.10
>30
>22.9
19.2
5.6
5c
24.7
16.0 16.07
5.87 0.93
6.29 1.69
12.80 21.02
2.94 0.76
2.27 0.26
5.87 0.3
5g
5.57 0.36
6.08 0.86
18.60 66.69
0.54 0.20
1.53 0.33
10.14 0.8
5h
5.92
17.5
1.69
1.91
8.11
5i
5o
1.58 2.97
1.93 0.85
8.31 2.4
5w
Sunitinib
did not induce any substantial rise in caspase 3/7 activity total DNA content of each cell to ascertain the phase of the
at any concentration or time point tested.
cell cycle. Concentrations of less than 100 µM to 50 nM of
compound 5o were utilized to assess its capability to
influence cell cycle distribution as well as Rb phosphor-
ylation. Figure 2 and Table 3 indicated that the total cell
number was reduced with an IC50 value of 2.20 µM after
a 48 h treatment. Also, the levels of phosphorylated Rb
protein were substantially decreased in a dose-dependent
Cell cycle influences
The A-549 cell line was used to examine the influence of
compound 5o on different features of the cell cycle pro-
gression. Activity assessment of compound 5o was
conducted using immunofluorescent imaging of phos-
phorylated Rb protein as well as by quantification of the manner (Figure 3A).
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1000
800
600
400
200
0
1000
800
600
400
200
0
24h
48h
24h
48h
0.01
0.1
1
10
100
0.01
0.1
1
10
100
Concentration (µM)
Concentration (µM)
5o
Sunitinib
Sunitinib
DMSO
5o
1.6
1.4
1.2
1
0.8
0.6
0.4
0.2
0
Figure 3 (A). Reduction of phosphorylated Rb protein by compound 5o and sunitinib. Levels of P-Rb in the nuclei were shown by immunofluorescence in cells treated with
vehicle, 5o or sunitinib. Automated image analysis (Molecular Devices) was used to quantitate P-Rb changes and these are presented in the dose–response graphs for each
compound after 24 hr or 48 hr treatment. (B) Western blot analysis of A-549 NSCLC cells treated with compound 5o shows the effect of on total cellular levels of P-Rb
after 24 hr treatment (left). Densitometric analysis of P-Rb normalized to GAPDH loading control is presented (right).
Abbreviations: GAPDH, glyceraldehyde-3-phosphate dehydrogenase; NSCLC, non-small cell lung cancer.
Independent experiments confirmed the effect of 5o to cycle, with a concomitant rise in the G1 phase. These results
reduce Rb phosphorylation by Western blot analysis suggest that part of the growth inhibition effect of compound
(Figure 3B). Moreover, compound 5o induced a reduction in 5o could be attributed to decreases in the progression rate of the
the percentage of cells in the S and G2/M phases of the cell cell cycle, with a concomitant reduction in proliferation. On the
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Table 3 Inhibitory concentration 50% (IC50) values for the decreases in the entire cell number and cell cycle influences of compound
5o and sunitinib
Compound No. IC50 (µM) for the decreases of
the entire cell number
IC50 (µM) for the decrease of
Rb phosphorylation
Cell cycle influences
24 hrs
48 hrs
24 hrs
48 hrs
5o
10.47
2.20 1.30
3.48 0.61
1.92 1.20
3.18 0.07
3.78 1.10
6.05 0.61
G1 increased and S, G2/M phases decreased
G1 decreased and S, G2/M phases increased
Sunitinib
12.54 9.82
contrary, sunitinib showed an increase in the percentage of
cells in the S or G2/M phases of the cell cycle, with a con-
comitant reduction in the G1 phase. Mitotic catastrophe fol-
lowed by programmed death of cells containing aberrant or
multiple nuclei may result from mitotic arrest due to arrest in
the G2 phase of the cell cycle, which might represent a check-
point blockade.
Figure 4 and Table 4 indicate that compound 5o was
able to inhibit cell growth in both tumor and normal cells.
However, it showed threefold selectivity, while sunitinib
displayed 1.4-fold selectivity.
Activity against multidrug-resistant cancer
cell line
It should be mentioned that both compound 5o and suni-
tinib substantially reduced the extent of phosphorylated Rb
protein in a dose-dependent fashion (Figure 3). Compound
5o exhibited IC50 values of 3.78 and 1.92 µM after 48 and 24
hrs, respectively, which was roughly twofold more potent
than sunitinib (Table 3). This may advance the assumption
that the growth inhibitory potential of 5o could be attributed,
in part, to its ability to inhibit cyclin-dependent kinases.
The growth inhibitory potential of compound 5o was
tested against the sensitive lung cancer cell line NSCLC
A-549 and the multidrug-resistant lung cancer cell line
NCI-H69AR which expresses the ABCC1 efflux pump
protein. Compound 5o was tested in quadruplicate at a
maximum concentration of 25 µM and 10 subsequent
serially diluted concentrations.
Figure 5 and Table 5 indicate that compound 5o induced
growth inhibition in both lung cancer cell lines, with an IC50
value of 0.9 µM in A-549 cells, and being about 12-fold less
sensitive toward the NCI-H69AR cell line. This result indi-
cates that compound 5o might undergo efflux by the ABCC1
efflux pump protein. In contrast, sunitinib was only 1.9-fold
less potent toward the NCI-H69AR cell line.
Selectivity
Three nontumorigenic cell lines (Table 4) were utilized to
examine the growth inhibitory selectivity of compound 5o:
IEC-6 cells which show morphologic and karyotypic charac-
teristics of normal rat intestinal epithelial cells,37 MCF-10A
cells which feature the characteristics of primary cultures of
breast tissue with a dome formation38 and Swiss 3t3 fibroblasts
derived from mice embryonic tissue which are both contact
inhibited and nontumorigenic.39 A human non-small cell lung
cancer (NSCLCA-549) cell line was used for comparison.
Compound 5o was tested in quadruplicate at a maximum
concentration of 25 µM and 10 subsequent serially diluted
concentrations.
Conclusion
The molecular hybrids 5a-w were evaluated as new antiproli-
ferative conjugates. Compounds 5o (bearing an N-benzylisatin
moiety) and 5w (bearing an N-phenylisatin moiety) were the
most active antiproliferative candidates, with IC50 values of
1.69 and 1.91 µM, respectively, being about fivefold and
fourfold more potent than sunitinib (IC50=8.11 µM).
Table 4 Selectivity of 5o and sunitinib against nontumorigenic and tumor cell lines
Compound No.
IC50 (µM)
Mean tumor selectivity
Intestine IEC-6
Breast MCF-10A
Fibroblast Swiss 3t3
NSCLCA-549
5o
3.38 1.21
4.56 0.54
1.69 1.12
4.43 0.23
2.78 1.12
4.07 0.75
0.86 121
3.06
3.0
1.4
Sunitinib
Abbreviations: MCF, Michigan cancer foundation; 3T3, 3-day transfer, inoculum 3×105 cells; IEC, intestinal epithelial cell; IC50, inhibitory concentration 50%; NSCLC, non-
small cell lung cancer.
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100
80
60
40
20
0
Sw3t3
IEC-6
MCF-10A
A-549
0.01
0.1
1
10
Concentration (µM)
5o
100
80
60
40
20
0
Sw3t3
IEC-6
MCF-10A
A-549
0.01
0.1
1
10
Concentration (µM)
Sunitinib
Figure 4 Selectivity characteristics of compound 5o and sunitinib.
100
80
H69AR
60
40
20
0
A-549
0.01
0.1
1
10
Concentration (µM)
5o
100
80
60
40
20
0
H69AR
A-549
0.01
0.1
1
Concentration (µM)
Sunitinib
10
Figure 5 Activity of compound 5o and sunitinib against A-549 and NCI-H69AR cell lines.
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Almutairi et al
Table 5 Cancer cell growth inhibitory activity of compound 5o and
sunitinib toward sensitive (A-549) and resistant (NCI-H69AR) cell
lines
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NCI-H69AR
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3.06
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Detailed pharmacological studies were conducted on
compound 5o, a promising antiproliferative candidate, for
a better understanding of its pharmacological properties.
Compound 5o did not show any significant rise in caspase
3/7 activity at any concentration or time point tested.
Moreover, it exhibited an increase in the G1 phase and a
reduction in the S and G2/M phases of the cell cycle, and
it presented an IC50value of 10.4 μM toward the resistant
NCI-H69AR cancer cell line. Furthermore, the extent of
phosphorylated Rb protein was substantially decreased in
a dose-dependent fashion by compound 5o which was
further confirmed via Western blot analysis. This promotes
the assumption that inhibition of cyclin-dependent kinases
by compound 5o plays a role in its growth inhibitory
potential.
Overall, the current investigation indicates that the new
antiproliferative potential of the chemical entities 5a-w,
compound 5o in particular, can support the development
of new antiproliferative leads to be harnessed in preclinical
studies of cancer chemotherapy.
Acknowledgment
The authors would like to extend their sincere appreciation
to the Deanship of Scientific Research at King Saud
University for its funding of this research through the
Research Group Project no. RGP-196.
18. Anouar EH, Moustapha ME, Taha M, et al. Synthesis, molecular
docking and β-glucuronidase inhibitory potential of indole base oxa-
diazole derivatives. Molecules. 2019;24(5):963. doi:10.3390/molecules
24050963
Disclosure
Dr Adam
B Keeton is a shareholder for ADT
Pharmaceuticals, LLC, outside the submitted work. Prof.
Dr. Gary A Piazza is a co-founder, shareholder, and Chief
Scientist for ADT Pharmaceuticals LLC and founder and
president of PDEi Pharmaceuticals LLC. The authors
report no other conflicts of interest in this work.
19. Attia MI, Witt-Enderby PA, Julius J. Synthesis and pharmacological
evaluation of pentacyclic 6a,7-dihydrodiindole and 2,3-dihydrodiin-
dole derivatives as novel melatoninergic ligands. Bioorg Med Chem.
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