"on water" expedient synthesis of 3-indolyl-3-hydroxy oxindole derivatives and their anticancer activity in vitro

Article abstract of DOI:10.1016/j.ejmech.2014.07.004

A series of 3-indolyl-3-hydroxy oxindole derivatives (n = 41) were synthesized by the green aminocatalytic method with excellent yields under mild reaction conditions. All the newly synthesized derivatives were subjected to evaluate their cytotoxic proper

Full text of DOI:10.1016/j.ejmech.2014.07.004

European Journal of Medicinal Chemistry 84 (2014) 155e159
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
“On water” expedient synthesis of 3-indolyl-3-hydroxy oxindole
derivatives and their anticancer activity in vitro
Parvathaneni Sai Prathima a, Pamanji Rajesh b, Janapala Venkateswara Rao b,
,
*
Uppalapati Sai Kailash c, Balasubramanian Sridhar d, Mandapati Mohan Rao a
a Inorganic & Physical Chemistry Division, Indian Institute of Chemical Technology, Hyderabad 500607, India
b Biology Division, Indian Institute of Chemical Technology, Hyderabad 500607, India
c National Centre for Cell Sciences, University of Pune Campus, Ganeshkhind, Pune 411007, India
d X-ray Crystallography Division, Indian Institute of Chemical Technology, Hyderabad 500607, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of 3-indolyl-3-hydroxy oxindole derivatives (n ¼ 41) were synthesized by the green amino-
catalytic method with excellent yields under mild reaction conditions. All the newly synthesized de-
rivatives were subjected to evaluate their cytotoxic properties against different human cancer cell lines.
Results indicated that about 73% of the derivatives exhibited significant anti-proliferative activities
against leukemia (U937, THP-1), lung (A549) and breast cancer (MCF7) cell lines. Among them a few of
the derivatives exhibited the most potent and effective cytotoxic activities on U937 (34, 36, 38 and 41)
and MCF7 (12, 35, 40 and 41) cell lines, and their anti-proliferation activities are better than the positive
control, Etoposide.
Received 20 September 2013
Received in revised form
1 July 2014
Accepted 2 July 2014
Available online 7 July 2014
Keywords:
Isatin
Indole
© 2014 Elsevier Masson SAS. All rights reserved.
Diethanolamine
Water
3-Indolyl-3-hydroxy oxindoles
Cytotoxicity
1. Introduction
Isatins are well-known biological manifolds with a reactive
keto-carbonyl group that readily undergoes condensations,
C3 functionalised oxindole is featured heterocyclic nucleus in a
number of natural products as well as medicinally relevant com-
pounds [1]. In particular 3-substituted-3-hydroxyoxindole is an
emerging new scaffold for drug discovery with a broad spectrum of
biological activities including antiviral, antibacterial, antituber-
cular, anti-inflammatory, antiangiogenic, antifungal, anticonvul-
sant and new targets for cancer chemotherapy [2]. Notably these
derivatives have been served as potential synthons for complex
natural product synthesis [3]. Several pharmacologically active al-
kaloids such as Maremycin A and B, Flustramnol, Arundaphine,
Donaxaridine, CPC-1, Welwitindoline C in addition to several others
contain 3-hydroxyoxindole moiety as shown in (Fig. 1) [4]. Recently
3-hydroxy-3,30-bisindolin-2-one 1 has been reported as potential
synthon for the enantioselective total synthesis of optically
pure(þ)-folicanthine [5].
resulting in C-3 functionalised oxindole derivatives. Increasing
demands for more efficient and scalable synthetic processes, which
are both economical and environmentally responsible in terms of
feedstock, energy consumption and waste production, require the
use of “green” solvents especially water.
Owing to the significance of 3-substituted-3-hydroxy-2-
oxindoles and intense research activity of medicinal chemists in
the construction of small bioactive chemical entities, we envisioned
the combination of both indole and oxindole motif with a hydroxy-
bearing C3 substitution to generate biologically attractive archi-
tectures (Fig. 2). The use of readily available isatins 1a and indoles
1b through a FriedaleCrafts type reaction represents an attractive
one-step entry to the valuable targets (Scheme 1) [6]. The literature
survey reveals that direct FriedaleCrafts reactions of isatins for the
selective synthesis of 3-indolyl-3-hydroxy oxindole derivatives are
less explored compared to the methods for synthesis of 3,30-
di(indolyl)oxindoles [7]. Of the developed strategies for the syn-
thesis of target 1 organocatalysis provides an efficient protocol due
to its ease of operation, tolerance of wide array of functional groups
[8]. As part of our current studies on the design of new routes for C-
* Corresponding author.
E-mail addresses: mandapati@iict.res.in, mandapatirao@yahoo.co.in (M. Mohan
Rao).
http://dx.doi.org/10.1016/j.ejmech.2014.07.004
0223-5234/© 2014 Elsevier Masson SAS. All rights reserved.

156
P. Sai Prathima et al. / European Journal of Medicinal Chemistry 84 (2014) 155e159
Table 1
Synthesis of 3-indolyl-3-hydroxy oxindole derivativesa.
Compds
R
R1
R2
R3
R4
Yield [%]b
1
2
3
4
5
6
7
8
H
H
H
H
H
H
H
H
PhCH2
Me
H
H
H
H
H
H
H
H
PhCH2
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Br
Br
Me
Br
Br
Me
Me
Me
Br
H
H
H
H
H
H
Cl
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Me
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
98
92
91
83
86
88
89
94
81
84
81
91
89
91
90
81
92
84
81
83
91
94
92
92
91
90
92
81
91
93
89
81
80
84
80
88
84
88
86
89
85
Cl
Br
I
F
Me
H
OCF3
H
H
H
OCF3
H
Cl
Br
I
F
Me
H
H
H
F
Cl
I
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
H
OMe
OMe
OMe
OMe
Br
Fig. 1. Bioactive natural products built with 3-substituted-3-hydroxy-2-oxindole core
scaffold.
F
I
F
Br
OCF3
F
H
Cl
H
OMe
Me
F
F
Br
Br
Br
Br
Br
H
H
H
OMe
OMe
OMe
4-Br-PhCH2
2-Br-PhCH2
2-Cl-PhCH2
2-Cl-PhCH2
PhCH2
H
Cl
Cl
Cl
H
Cl
H
PhCH2
2-Cl-PhCH2
2-Cl-PhCH2
2-Br-PhCH2
2-Cl-PhCH2
2-Cl-PhCH2
Fig. 2. Oxindole motif bearing hydroxy-group and indole at C3 position.
H
Cl
a
Reaction conditions: Isatin (0.5 mmol), Indole (0.5 mmol), diethanolamine
(0.02 mmol) and water (2 mL) at room temperature.
b
Overall yield of the product.
diamine, piperidine, pyridine, imidazole and diethanolamine using
water as solvent. However the reaction proceeded spontaneously at
ambient temperature with diethanolamine in water and the cor-
responding 3-indolyl-3-hydroxy oxindole 1 was obtained in 98%
Scheme 1. Electrophilic addition of indoles to isatins.
3 functionalization of isatins [9] to biologically active heterocyclic
compounds, we herein disclose a simple, green aminocatalytic
method for the efficient synthesis of 3-indolyl-3-hydroxy oxindoles
in water and their evaluation for anticancer activity.
Initially, we examined the reaction of isatin 1a and indole 1b for
the synthesis of 3-indolyl-3-hydroxy oxindole 1 with various amine
based catalysts, including triethylamine, pyrrolidine, ethylene
Scheme 2. Synthesis of 3-indolyl-3-hydroxy oxindoles from isatin.
Fig. 3. ORTEP drawing of 38 (30% thermal ellipsoids).

P. Sai Prathima et al. / European Journal of Medicinal Chemistry 84 (2014) 155e159
157
Fig. 4. Images showing morphological changes in U937 cells after exposure to IC50 concentrations of different test derivatives (400X). (A) Controls with intact cell structure, (B, C and D)
U937 Treated cells (4.33, 6.07 and 8.07
m
g/ml of derivatives 36, 41 and 35, respectively) exhibited shrinkage in cell size, nuclear condensation and formation of apoptotic bodies.
Table 2
Table 2 (continued )
In vitro cytotoxicity of oxindole derivatives (structural analogues) against U937, THP-
1, A549 & MCF7 human cancer cells by MTT assay.
S. no/Test code
IC50 valuesa
U937
(
m
g/ml)
S. no/Test code
IC50 valuesa
U937
(m
g/ml)
THP-1
THP-1
A549
MCF7
37
38
39
40
14.99 2.65
4.89 2.39
NA
7.6 0.47
6.07 0.35
6.14 1.19
6.97 2.65
7.01 2.39
NA
5.6 1.76
4.27 0.92
2.16 0.15
28.14 1.27
15.76 0.78
NA
33.09 1.67
14.09 0.16
10.71 1.26
39.27 2.24
20.62 1.24
NA
13.44 0.56
10.82 0.34
17.76 1.31
A549
MCF7
1
2
3
4
5
6
7
8
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
19.82 2.22
19.27 0.22
21.56 0.46
43.74 2.13
83.36 4.05
NA
16.79 2.29
42.98 0.33
NA
72.47 1.68
NA
NA
NA
NA
NA
22.75 0.75
NA
NA
12.99 2.98
NA
19.81 0.55
25.13 3.04
25.20 1.56
NA
NA
25.1 3.78
NA
NA
NA
NA
NA
41
Etoposideb
Exponentially growing cells were treated with different concentrations of oxindole
derivatives for 24 h and cell growth inhibition was analyzed through MTT assay.
86.14 4.20
13.72 2.9
64.52 3.91
NA
NA
NA
a
IC50 is defined as the concentration, which results in a 50% decrease in cell
71.55 7.71
NA
48.96 2.94
NA
5.47 0.22
NA
95.49 1.59
68.70 1.79
84.49 2.95
NA
NA
31.22 1.74
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
number as compared with that of the control cultures in the absence of an inhibitor
and were calculated using the respective regression analysis. The values represent
the mean SE of three individual observations.
Etoposide was employed as positive control, NA indicates that the derivatives
are not active at 100 mg/ml concentration.
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
NA
NA
NA
b
NA
25.31 0.38
43.32 0.67
23.15 1.31
35.51 0.94
NA
17.07 1.6
NA
NA
55.86 1.85
39.81 0.52
NA
NA
NA
98.52 5.76
86.11 1.40
87.11 2.83
NA
NA
14.17 1.3
NA
NA
NA
NA
69.18 0.01
75.95 4.69
62.4 3.04
NA
NA
NA
yield (Scheme 2). These results provided the incentive for further
study of reactions with various other isatin derivatives and
substituted indoles to furnish the corresponding 3-indolyl-3-
hydroxy oxindoles [12]. Diverse functionalities of isatin as well as
indoles gave excellent yields (80e98%) of the desired product with
diethanolamine under mild reaction conditions (Table 1).
Several N-alkylated isatins with substituents (methyl, benzyl,
allyl, halo substituted benzyl) at N-1 position are synthesized by
adapting the reported procedure and subjected to functionalization
at C-3 position [10]. The reaction is fast with high yields with
unsubstituted isatin and indoles when compared to substituted
ones (Table 1). In case of Cl, Br and OCF3, at C-5 position the re-
action proceeded faster with good yields for compounds 2, 3, and 8.
In case of benzyl and methyl substituent at the N-1 position, the
reaction observed to be slow with 81 and 84 % yields respectively
for compounds 9, and 10. The reaction of isatins with electron
donating groups is rapid with high yields compared to electron
withdrawing groups. All examples of isatins 1e41 with electron
NA
43.71 1.74
NA
NA
NA
NA
18.37 1.61
29.76 0.61
16.16 0.22
16.52 2.31
13.32 1.34
18.32 3.23
NA
NA
NA
NA
NA
12.16 2.27
14.93 0.83
7.16 0.17
5.31 2.83
8.02 0.91
4.33 0.64
6.06 1.29
6.93 0.38
7.37 0.11
6.12 0.87
7.12 1.91
5.03 0.56
20.37 4.35
24.73 2.03
22.31 0.69
21.26 0.72
8.64 0.34
24.03 0.91

158
P. Sai Prathima et al. / European Journal of Medicinal Chemistry 84 (2014) 155e159
Fig. 5. Comparison of activity of oxindole derivatives on U937, THP1, A549 and on MCF7 cancer cell lines.
withdrawing groups at C-5 position, C-7 position and substituents
at N-1 position and substitutions at C-5 position of indoles lead to
equally satisfying results, which were then evaluated for their po-
tential biological activity. The general structure of the obtained
products i.e. 3-indolyl-3-hydroxy oxindole derivatives (1e41) are
represented as follows.
5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide] assay,
according to the method of Mossman [13] and the exposed cells
with different IC50 concentrations of derivatives showed the char-
acteristic nuclear and cytological alterations (morphological
changes) under light microscopy (Fig. 4). It is evident from the re-
sults that most of the test compounds have shown significant
cytotoxic activity on all tested cell lines in a concentration-
dependent manner (Table 2).
The oxindoles derivatives are found to be effective on U937 cells
(58%) followed by THP-1 (49%), A549 (44%) and MCF7 cells (41%) at
below 100 mg/ml concentration. It is apparent from the results that
these derivatives are found to be more potent on U937, MCF7 cell
lines followed by THP1 and A549. Among the derivatives, 3, 12 and
26 are effective only against any of the two cell lines followed by 8
and 9 on any of the three cell lines tested. From the screened
compounds, 11 derivatives were found to be active on any one of
the cell lines used. The compounds 35 and 41 have exhibited an
excellent cytotoxic activity against all the four cell lines.
The relative fold activity (increase/decrease) of the active de-
rivatives in comparison to Etoposide is presented in Fig. 5.
Comparatively, few of the derivatives (34, 36, 38 and 41 on
U937 cell lines; 12, 35, 40 and 41 against MCF7 cell lines) exhibited
most potent activity, which are better than the activity of positive
control, Etoposide, i.e., 0.5e1.5 fold and 1e3 fold, respectively. The
cytotoxic activities of many of these derivatives (y12 derivatives)
are pretty close to the inhibitory activity of Etoposide.
The structure in the crystalline solid state of compound 38 was
determined by single-crystal X-ray diffraction analysis. Suitable
crystals of it for structural determination were obtained from slow
evaporation of an ethyl acetate solution. Molecular structure of
product 38 is shown in Fig. 3 [11].
2. In vitro cytotoxic activity
The biological activities of 3-indolyl-3-hydroxy oxindole de-
rivatives (n ¼ 41) were evaluated to investigate their anti-
proliferative/cytotoxic activities on four different types of human
cancer cell lines such as leukemia (U937, THP-1), lung and breast
cancer cell lines (A549 and MCF7, respectively) using the MTT [3-(4,
3. Conclusion
In view of synthetic and biological significance of oxindole ar-
chitecture with a 3,3-disubstituted framework, we have developed

P. Sai Prathima et al. / European Journal of Medicinal Chemistry 84 (2014) 155e159
159
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