Synthesis, anticancer activity and docking of some substituted benzothiazoles as tyrosine kinase inhibitors

Article abstract of DOI:10.1016/j.jmgm.2010.04.003

Protein tyrosine kinases occupy a central position in the control of cellular proliferation and its inactivation might lead to the discovery of a new generation anticancer compounds. Substituted benzothiazoles have been found to mimic the ATP-competitive

Full text of DOI:10.1016/j.jmgm.2010.04.003

Journal of Molecular Graphics and Modelling 29 (2010) 32–37
Contents lists available at ScienceDirect
Journal of Molecular Graphics and Modelling
journal homepage: www.elsevier.com/locate/JMGM
Synthesis, anticancer activity and docking of some substituted
benzothiazoles as tyrosine kinase inhibitors
Hemal A. Bhuvaa, Suvarna G. Kinib,∗
a Vidyabharti Trust College of Pharmacy, Umrakh 394345, Gujarat, India
b Department of Pharmaceutical Chemistry, Manipal College of Pharmaceutical Sciences, Madhav Nagar, Manipal 576104, Karnataka, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Protein tyrosine kinases occupy a central position in the control of cellular proliferation and its inactiva-
tion might lead to the discovery of a new generation anticancer compounds. Substituted benzothiazoles
have been found to mimic the ATP-competitive binding of genistein and quercetin to tyrosine kinase.
A series of novel 2-phenyl-1,3-benzothiazoles were synthesized and characterised by IR, 1H NMR and
mass spectroscopy. All the compounds were tested for their anticancer activity against MCF-7 breast
cancer cell line with the MTT assay. Most of the compounds showed moderate to good anti-breast cancer
activity. Anticancer activity varied with substitution on the benzothiazole nucleus with halogens and
at 4 position, substitution of the 2-phenyl moiety with methyl and methoxy groups was also explored.
Among the compounds tested with MTT assay, mono fluoro substitution on benzothiazole nucleus and
4ꢀ-methyl variations at 2-phenyl position demonstrated highest percent growth inhibition of MCF-7 cells.
Docking studies of the synthesised compounds was done on EGFR using GRIP batch docking method to
study their observed activity.
Received 29 September 2009
Received in revised form 3 February 2010
Accepted 17 April 2010
Available online 24 April 2010
Keywords:
Anticancer
Breast cancer MCF-7 cell line
Synthesis
Docking
Tyrosine kinase
© 2010 Elsevier Inc. All rights reserved.
1. Introduction
cellular proliferation. Several transforming oncogens (e.g., src –
a gene obtained from Rous sarcoma virus, abl – a gene obtained
Cancer is a group of diseases in which cells can be aggres-
sive (grow and divide without respect to normal limits), invasive
(invade and destroy adjacent tissues) and/or metastatic (spread to
other locations in body). These three malignant properties of can-
cer differentiate them from benign tumors, which are self-limited
in their growth and do not invade or metastasize. Cancer may
affect people at all ages, even fetuses, but risk for the more com-
mon varieties tends to increase with age [1]. Cancer causes about
13% of all deaths [2]. According to the American Cancer Society,
7.6 million people died from cancer in the world during 2007 [3].
Nearly all cancers are caused by abnormalities in the genetic mate-
rial of the transformed cells [4]. With the current chemotherapy,
lack of selectivity of chemotherapeutic agents against cancerous
cells is a significant problem.
Receptor tyrosine kinases (RTKs) are high affinity cell surface
receptors that bind polypeptide growth factors, cytokines and
hormones. They have been shown to be key regulators of nor-
mal cellular processes and additionally play a critical role in the
development and progression of many types of cancer [5]. Pro-
tein tyrosine kinases occupy a central position in the control of
from abelson murine leukemia virus) are known to possess tyro-
sine kinase activity; and it is well recognized that the response
of many cells to growth factors is initiated by activation of RTKs.
Over expression of certain RTKs show association with promo-
tion and maintenance of malignancies. For example, the epidermal
growth factor (EGF) receptor tyrosine kinases of the erbB family
(which includes erbB1–erbB4) is frequently expressed at high lev-
els in certain carcinomas and shows an inverse correlation with
survival (particularly breast, colon and bladder cancers) [6]. Thus,
inactivation of the specific tyrosine kinases that are responsible for
the malignant phenotype of certain cancers represent a potential
approach for design of antiproliferative drugs [7].
The isoflavone, genistein [8] and the flavone, quercetin
[9] are competitive inhibitors at the ATP-binding site of
kinases [10,11]. As the crystal structure of 5,6-dimethoxy-2-(4-
methoxyphenyl)benzothiazole was solved [12], the preliminary
analysis based on comparisons between polyhydroxylated 2-
phenylbenzothiazoles and the adenine fragment of ATP suggested
that suitably substituted benzothiazoles might mimic the ATP-
competitive binding of genistein and quercetin at tyrosine kinases.
In this study, 10 derivatives of 2-phenyl-1,3-benzothiazole
with halogen substitution on benzothiazole moiety and methyl/
methoxy substitution at 4 position of the 2-phenyl ring were syn-
thesized and evaluated for anticancer activity on the MCF-7 human
breast cancer cell line. We have also tried to dock the synthesised
∗
Corresponding author. Tel.: +91 820 257 1201x22482; fax: +91 820 257 1998.
E-mail addresses: hemalbhuva@yahoo.co.in (H.A. Bhuva),
suvarna.gk@manipal.edu (S.G. Kini).
1093-3263/$ – see front matter © 2010 Elsevier Inc. All rights reserved.
doi:10.1016/j.jmgm.2010.04.003

H.A. Bhuva, S.G. Kini / Journal of Molecular Graphics and Modelling 29 (2010) 32–37
33
Table 1
Physical data of the intermediates HB1a–HB10a (benzanilides).
Table 2
Physical data of the intermediates HB1b–HB10b (thiobenzanilides).
a
Intermediate
R1
R2
% Yield
Mt. pt. (◦C)
Rf
Intermediate
R1
R2
% Yield
Mt. pt. (◦C)
Rf
HB1a
HB2a
HB3a
HB4a
HB5a
HB6a
HB7a
HB8a
HB9a
HB10a
4-F
Me
Me
77.5
54.7
54.4
54.8
56
82.8
42.2
68
176–184
152–158
212–220
186–194
222–230
126–136
116–120
136–142
150–158
114–122
0.549
0.628
0.359
0.292
0.628
0.546
0.47
0.536
0.337
0.782
HB1b
HB2b
HB3b
HB4b
HB5b
HB6b
HB7b
HB8b
HB9b
HB10b
4-F
Me
Me
52.5
37.5
62.3
77.9
54.4
24.8
45.1
33.6
66
156–164
152–162
176–182
206–212
198–206
130–136
150–154
125–132
160–168
120–124
0.709a
0.659a
0.488a
0.376a
0.686a
0.574b
0.458a
0.533a
0.35a
3,4-diF
4-Br
4-F
4-Br
3-Cl
3-Br
2-F
3,4-diF
4-Br
4-F
4-Br
3-Cl
3-Br
2-F
OMe
OMe
Me
OMe
OMe
OMe
OMe
Me
OMe
OMe
Me
OMe
OMe
OMe
OMe
Me
3,4-diF
2-F
58.7
42
3,4-diF
2-F
15.5
0.565a
a
a
n-Hexane:ethyl acetate, 4:1.
n-Hexane:ethyl acetate, 4:1.
n-Hexane:ethyl acetate, 4.5:0.5.
b
compounds with the crystal structure of EGFR to explore the pos-
sible anticancer mechanism of our compounds. Prior compounds
have been reported [13,14] to compare the anticancer activity of
such compounds.
ide. Aliquots of this mixture (1.0 mL) were added at 1 min intervals
to a stirred solution of potassium ferricyanide (4.0 mol equiv.) in
water at 80–90 ◦C. The reaction mixture was heated for an addi-
tional 30 min and then allowed to cool. The solid was collected,
washed with water and recrystallized from methanol. Yields and
physical characteristics are listed in Table 3.
2. Materials and methods
2.1. Synthesis
2.1.3.4. HB1c
[6-fluoro-2-(4-methylphenyl)-1,3-benzothiazole].
Yield: 52.1%. IR (KBr): 817.85 (C–H), 1606.76 (C N), 702.11 (C–S),
2.1.1. Reagents
1192.05 (C–F) cm−1 1H NMR (DMSO): ı 2.4 (s, 3H, CH3 of Ar), ı
.
All the chemicals and solvents used were of AR-grade and
LR-grade and obtained from Sigma-Aldrich, Sisco Research Labora-
tories, Qualigens, Rankem, S.D. Fine, Hi-Media and Merck and were
used without further purification.
7.25–7.9 (m, 7H, Ar). GC–MS: 243 (M+).
2.1.3.5. HB2c [5,6-difluoro-2-(4-methylphenyl)-1,3-benzothiazole].
Yield: 53.9%. IR (KBr): 817.85 (C–H), 1608.69 (C N), 709.83 (C–S),
1197.83 (C–F) cm−1 1H NMR (DMSO): ı 2.4 (s, 3H, CH3 of Ar), ı
.
2.1.2. TLC analysis
7.25–7.9 (m, 6H, Ar). GC–MS: 261 (M+).
Thin-layer chromatography (TLC) was performed on pre-coated
aluminium plates (Silicagel 60 F254, Merck). Plates were visualized
by UV light and iodine vapour.
2.1.3.6. HB3c
[6-bromo-2-(4-methoxyphenyl)-1,3-benzothiazole].
Yield: 65.1%. IR (KBr): 819.77 (C–H), 1681.33 (C N), 692.47 (C–S),
561.3 (C–Br), 1026.16, 1246.06 (C–O–C) cm−1 1H NMR (DMSO): ı
.
2.1.3. Equipment
3.85 (s, 3H, OCH3), ı 7–8 (m, 7H, Ar). GC–MS: 321 (M++1).
Melting points were measured on a electrothermal melting
point apparatus Toshniwal apparatus (Toshniwal Company, Banga-
lore, India) and are uncorrected. Infrared (IR) spectra were recorded
as KBr pellets with an FTIR-8300 spectrophotometer (Shimadzu).
Proton magnetic resonance (1H NMR) spectra were recorded in
DMSO-d6 (Merck) on a AMX-400 NMR spectrometer (IISc, Banga-
lore, India). Mass spectra were recorded on a GCMS (QP 5050A,
Shimadzu Corporation, Japan). Absorption maxima were taken on
a UV-Visible spectrophotometer-2450 (Shimadzu).
2.1.3.7. HB4c
Yield: 40.6%. IR (KBr): 821.70 (C–H), 1602.99 (C N), 702.11 (C–S),
1193.98 (C–F), 1031.95, 1249.91 (C–O–C) cm−1 1H NMR (DMSO):
ı 3.85 (s, 3H, OCH3), ı 7–8 (m, 7H, Ar). GC–MS: 259 (M+).
[6-fluoro-2-(4-methoxyphenyl)-1,3-benzothiazole].
.
2.1.3.8. HB5c
Yield: 47.8%. IR (KBr): 831.35 (C–H), 1608.69 (C N), 692.47 (C–S),
561.3 (C–Br), 1481.38 (C C) cm−1
[6-bromo-2-(4-methylphenyl)-1,3-benzothiazole].
.
2.1.3.1. Synthesis of benzanilides (HB1a–10a). 0.02 mol of an appro-
priately substituted aniline was dissolved in 20 mL of pyridine. To
this, 0.02 mol of appropriately substituted benzoyl chloride was
added with dropwise addition. The reaction mixture was main-
tained at reflux for 2 h and then poured into water. The precipitate
was collected, washed with water, dried and recrystallized from
methanol. Yields and physical characteristics are listed in Table 1.
2.1.3.9. HB6c
Yield: 60.4%. IR (KBr): 827.49 (C–H), 1606.76 (C N), 690.54 (C–S),
1112.96 (C–Cl), 1485.24 (C C) cm−1
[5-chloro-2-(4-methoxyphenyl)-1,3-benzothiazole].
.
2.1.3.10. HB7c [5-bromo-2-(4-methoxyphenyl)-1,3-benzothiazole].
Yield: 52.1%. IR (KBr): 831.35 (C–H), 1608.69 (C N), 694.4 (C–S),
551.66 (C–Br), 1485.24 (C C) cm−1
.
2.1.3.2. Synthesis of thiobenzanilides (HB1b–10b). A mixture of
the substituted benzanilides (0.01 mol) and Lawesson’s reagent
(0.6 mol equiv.) was refluxed in chlorobenzene (5–10 mL) for 3 h
to produce a clear solution. The solution was cooled and the prod-
uct precipitated. This solid was collected and recrystallized from
methanol. Yields and physical characteristics are listed in Table 2.
2.1.3.11. HB8c [5-fluoro-2-(4-methoxyphenyl)-1,3-benzothiazole].
Yield: 41.9%. IR (KBr): 825.56 (C–H), 1602.9 (C N), 692.47 (C–S),
1222.91 (C–F), 1485.24 (C C) cm−1
.
2.1.3.12. HB9c
benzothiazole]. Yield: 64.1%. IR (KBr): 833.28 (C–H), 1602.9
(C N), 702.11 (C–S), 1209.41 (C–F), 1455.3 (C C) cm−1
[5,6-difluoro-2-(4-methoxyphenyl)-1,3-
.
2.1.3.3. Synthesis of 2-phenyl-1,3-benzothiazoles (HB1c–10c). Sub-
stituted thiobenzanilides (0.01 mol) were wetted with a little
ethanol (5.0 mL)and 30% aqueous sodium hydroxide solution
(8.0 mol equiv.) was added. The mixture was diluted with water to
provide a final solution/suspension of 10% aqueous sodium hydrox-
2.1.3.13. HB9c
Yield: 25.7%. IR (KBr): 819.77 (C–H), 1610.61 (C N), 659.68 (C–S),
1188.19 (C–F), 1473.66 (C C) cm−1
[4-fluoro-2-(4-methylphenyl)-1,3-benzothiazole].
.

34
H.A. Bhuva, S.G. Kini / Journal of Molecular Graphics and Modelling 29 (2010) 32–37
Table 3
Physical data of the intermediates HB1c–HB10c (benzothiazoles).
Compound
HB1c
R1
R2
% Yield
52.1
Mt. pt. (◦C)
118–126
Rf
Elemental analysis calculated (found)
6-F
Me
0.689a
C-69.11% (69.10%)
H-4.14% (4.13%)
N-5.76% (5.75%)
HB2c
HB3c
HB4c
HB5c
HB6cd
HB7cd
HB8c
HB9c
HB10c
5,6-diF
6-Br
6-F
Me
53.9
65.1
40.6
47.8
60.4
52.1
41.9
64.1
25.7
150–160
160–168
120–128
168–174
116–122
128–136
112–118
132–140
140–148
0.86b
C-64.35% (64.32%)
H-3.47% (3.45%)
N-5.36% (5.35%)
OMe
OMe
Me
0.615b
0.899c
0.966b
C-52.51% (52.50%)
H-3.15% (3.14%)
N-4.37% (4.36%)
C-64.85% (64.83%)
H-3.89% (3.88%)
N-5.4% (5.41%)
6-Br
C-55.28% (55.27%)
H-3.31% (3.30%)
N-4.60% (4.61%)
5-Br
7-Br
OMe
OMe
0.614b
0.693
C-52.51% (52.50%)
H-3.15% (3.14%)
N-4.37% (4.36%)
5-Cl
7-Cl
OMe
OMe
0.558b
0.628
C-60.98% (60.97%)
H-3.66% (3.65%)
N-5.08%(5.07%)
4-F
OMe
OMe
Me
0.678b
0.655b
0.891b
C-64.85% (64.83%)
H-3.89% (3.88%)
N-5.4% (5.41%)
5,6-diF
4-F
C-60.63% (60.62%)
H-3.27% (3.26%)
N-5.05% (5.06%)
C-69.11% (69.10%)
H-4.14% (4.15%)
N-5.76% (5.75%)
a
n-Hexane: 5.
b
n-Hexane:ethyl acetate, 4.5:0.5.
n-Hexane:ethyl acetate, 4:1.
Gives regioisomers.
c
d
2.2. Biological evaluation and docking studies
2.2.1. Anticancer activity
Growth of breast cancer cells was quantitated by the ability of
living cells to reduce the yellow MTT to purple formazan products
[15]. The amount of formazan product formed is directly propor-
tional to the number of living cells.
and the formazan product was solubilised by addition of 100 ␮L
DMSO. The purple colour was produced. Absorbance of each well
was read on Tenac 200 plate reader at 570 nm. From the absorbance,
A −A
c
A
t
the % inhibition was calculated as% growth inhibition =
×
c
100where Ac is the mean absorbance of control and At is the mean
absorbance of test (Table 4).
Synthesized compounds were prepared as 4.0 mM top stock
solutions, dissolved in DMSO. MCF-7 human breast cancer cells
(human breast adenocarcinoma cell line originally obtained in 1973
from Michigan Cancer Foundation) were cultivated at 37 ◦C in
an atmosphere of 5% CO2 in Dubecco’s modified Eagle’s minimal
medium (DMEM) supplemented with 3.0 mM l-glutamine with
10% fetal bovine serum were routinely subcultured twice weekly to
maintain in continuous logarithmic growth. Cells were trypsinized
for the passage into the well plate and plated at 10,000 cells/well in
100 ␮L of medium in 96-well plates. Cells were allowed to adhere
to the surface of well plates. After 24 h, medium was removed and
100 ␮L of drug solutions (prepared at 12.5, 25, 50 and 100 ␮M con-
centrations) were added into the wells. 100 ␮L of fresh medium
without cells was added as control. 4 wells were used for each
concentration of drug solution, while 4 wells were reserved for
cell culture control, which contained the corresponding amount
of DMSO. The total drug exposure was 72 h. After 72 h, contents of
the well were removed and 20 ␮L of MTT solution (5 mg in 1 mL of
phosphate buffer saline) was added to each well. Incubation at 37 ◦C
for 4 h allowed reduction of MTT by mitochondrial dehydrogenase
to an insoluble formazan product. Well contents were removed
2.2.2. Docking studies
The involvement of the EGFR family of tyrosine kinases in cancer
proliferation suggests that an inhibitor which blocks the tyrosine
kinase activity of the entire EGFR family, could have significant
therapeuticpotential [16]. So weselected EGFRasabiological target
for carrying out the docking study of our synthesized compounds
Table 4
% Growth inhibition of MCF-7 cells.
Compound
% Growth inhibition
12.5 ␮M
25 ␮M
50 ␮M
100 ␮M
HB1c
HB2c
HB3c
HB4c
HB5c
HB6c
HB7c
HB8c
HB9c
HB10c
5.2
0.67
16.78
8.49
5.41
4.96
2.75
8.3
8.26
6.38
23.8
14.57
4.77
12.68
9.28
13.25
1.67
10.63
29.33
23.25
42.17
50.16
15.29
21.56
11.58
21.23
11.08
25.35
62.8
55.18
60.64
62.06
57.14
37.08
34.49
34.92
20.46
42.79
0.48
3.28

H.A. Bhuva, S.G. Kini / Journal of Molecular Graphics and Modelling 29 (2010) 32–37
35
Table 5
Docking scores of the final synthesised compounds.
Compound
Dock score
HB1c
HB2c
HB3c
HB4c
HB5c
HB6c
HB7c
HB8c
HB9c
HB10c
−44.8585
−49.4539
−48.1762
−48.2842
−47.4309
−50.2033
−48.6359
−47.4589
−52.0807
−51.0141
and learn the mechanism of activity. The crystal structure of EGFR
kinase domain in complex with an irreversible inhibitor (PDB ID:
2J5F) was obtained from the protein data bank. The crystal struc-
ture was refined using vLife Science’s MDS 3.0 software [17]. The
refinement of the crude PDB structure of receptor was done by com-
pleting the incomplete residues. The co-crystallized ligand lying
within the receptor was modified by assigning missing bond order
and hybridization states. The side chain hydrogens were then added
to the crystal structure and their positions were optimized up to the
rms gradient 1 by aggregating the other part of the receptor. The
optimized receptor was then saved as mol file and used for docking
simulation.
The 2D structure of the compounds HB1c–HBl0c were built and
then converted into the 3D with the help of vLife MDS 3.0 soft-
ware [18]. The 3D structures were then energetically minimized
up to the rms gradient of 0.01 using Merck Molecular Force Field
(MMFF) [19]. Conformers of all the synthesized ligands were then
generated by Monte Carlo method. In doing so, all rotatable bonds
of the ligands were selected and number of seeds used for search-
ing the conformational space was set as 5. All conformers were
then energetically minimized up to the rms gradient of 0.01 and
then saved in separate folder [20]. The active site selection was
done by choosing the cavity having maximum hydrophobic surface
area. The docking simulation was done using GRIP batch docking.
Fig. 1. Graph of
HB6c–HB10c (B).
%
growth inhibition of MCF-7 cells by HB1c–HB5c (A) and
In this, all generated conformers of one ligand were put as one
batch in GRIP docking wizard. Likewise, the batches for all other
ligands were put. All the conformers were virtually docked at the
defined cavity of the receptor. The parameters fixed for docking
Scheme 1.

36
H.A. Bhuva, S.G. Kini / Journal of Molecular Graphics and Modelling 29 (2010) 32–37
simulation were like this – number of placements: 30, rotation
angle: 30◦, exhaustive method, scoring function: dock score. By
rotation angle, the ligand gets rotated for different poses. By place-
ments, the method will check all the 30 possible placements into
the active site pocket and will result out few best placements out of
30. For each ligand, all the conformers with their best placements
and their dock score will be saved in output folder. The method
also highlights the best placement of best conformer of one par-
ticular ligand which is having best (minimum) dock score. In the
results of docking, we have listed only best conformers and the dock
score for each ligand in Table 5. The ligand forming most stable
drug-receptor complex is the one which is having minimum dock
score. After docking simulation, the best docked conformer of each
ligand and receptor were merged and their complexes were then
energetically optimized by defining the radius of 10 Å measured
from the docked ligand. Stepwise energy optimization was done
by first hydrogens , second side chains and finally the backbone
of receptor [21]. The optimized complexes were then checked for
various interaction of ligand with receptor like hydrogen bonding,
hydrophobic bonding and van der Waal’s interaction. No compound
was found to exhibit hydrogen bonding with the receptor. So all the
docked compounds were analyzed for various types of other inter-
actions like hydrophobic bonding and van der Waal’s interaction.
All the compounds were found to form hydrophobic bonding by ‘C’
atom of methyl and methoxy group. Some of the common residues
involved in this type of interaction are Leu178, Lys745, Thr790 and
Val726. All the compounds were found to exhibit large number of
van der Waal’s bonding with wide range of residues. So it was very
difficult to find out common residues involved in this type of inter-
action. Only Val726 was found to be the common residue involved
in docking of all the ligands (Fig. 2B and C).
they possessed methoxy group at para position of 2-phenyl ring
as shown in Scheme 1.
In the case of cyclization of HB6b [N-(3-chlorophenyl)-
4-methoxybenzenecarbothioamide], a mixture of the 5-chloro
and its regioisomer 7-chloro methoxybenozthiazoles (HB6c)
was formed
. Similarly from HB7b [N-(3-bromophenyl)-4-
methoxybenzenecarbothioamide], a mixture of the 5-bromo and
7-bromo substituted isomers (HB7c) was formed. It was very dif-
ficult to separate these mixtures of regioisomers. Therefore, the
anticancer potential of HB6c and HB7c was found out by consid-
ering them as a mixture of regioisomers. In all other cases, only a
single methyl and methoxy benzothiazole isomer was formed [22].
Structures, yields and physical characteristics of all methyl
and methoxybenzanilides, thiobenzanilides and halogenated
2-(4-methylphenyl)-benzothiazoles and 2-(4-methoxyphenyl)-
benzothiazoles are given in Tables 1–3 respectively.
In MTT assay, the anticancer activity of all the synthesized com-
pounds was found out at 12.5, 25, 50 and 100 ␮M concentration.
The % growth inhibition of MCF-7 cells (presented in Fig. 1 and
listed in Table 4) was calculated from the measured absorbance of
formazan product produced by living cells. At 12.5 ␮M concentra-
tion, compound HB3c showed highest cell growth inhibition, while
HB1c, 4c, 5c, 6c and 8c showed moderate inhibition and HB2c and
HB9c showed least inhibition. At 25 ␮M concentration, HB3c exhib-
ited highest activity, while HB4c, 6c, 8c and 10c exhibited moderate
activity and HB5c and 9c exhibited least activity. At 50 ␮M con-
centration, HB4c showed highest activity, while HB1c, 2c and 10c
showed moderate activity and HB7c and 9c showed least activity.
At 100 ␮M concentration, HB1c exhibited highest inhibitory activ-
ity, while HB6c, 7c and 8c exhibited moderate activity and HB9c
showed least inhibitory activity. These results indicated that ben-
zothiazole derivatives we synthesised are able to inhibit epidermal
growth factor receptor (EGFR) in human breast cell line, MCF-7.
From the dock score, compounds HB9c, HB10c and HB6c were
found to have highest negative dock score ranging from −52.0
to −50.0 (Table 5). It means that these formed most stable
drug-receptor complex amongst other compounds. All the docked
compounds were analyzed for various types of interactions like
3. Results and discussion
Four derivatives of 2-phenyl-1,3-benzothiazole in which mono
or di-halogenated substitution on benzothiazole moiety and
methyl group at para position of 2-phenyl ring were synthesized.
Same type of another six derivatives was also synthesized but
Fig. 2. (A) Docking position of all synthesised compounds in the receptor cavity. (B) van der Waal’s interaction of HB9c (ball and stick) with the receptor residues (wire)
shown by pink colour dotted lines. (C) Hydrophobic interaction of HB9c (ball and stick) with the receptor residues (wire) by ‘C’ atom of methoxy group shown by light green
dotted lines.

H.A. Bhuva, S.G. Kini / Journal of Molecular Graphics and Modelling 29 (2010) 32–37
37
hydrophobic bonding and van der Waal’s interaction because no
compound was found to exhibit hydrogen bonding with the recep-
tor. Fig. 2A shows the docking position of all the compounds in the
receptor cavity.
[3] American Cancer Society, Report Sees 7.6 Million Global 2007 Cancer Deaths,
December 2007.
[4] K.W. Kenneth, B. Vogelstein, Introduction, in: The Genetic Basis of Human Can-
cer, 2nd, illustrated, revised ed., McGraw-Hill, Medical Pub. Division, New York,
2002, p. 5.
[5] E. Zwick, J. Bange, A. Ullrich, Receptor tyrosine kinase signalling as a target for
cancer intervention strategies, Endocr. Relat. Cancer 8 (2001) 161–173.
[6] W.J. Gullick, Prevalence of aberrant expression of the epidermal growth factor
receptor in human cancers, Br. Med. Bull. 47 (1991) 87–98.
[7] Y. Yardern, A. Ullrich, Growth factor receptor tyrosine kinases, Annu. Rev.
Biochem. 57 (1988) 443–478.
[8] T. Akiyama, J. Ishida, S. Nakagawa, H. Ogawara, S. Watenabe, N. Itoh, M. Shibuya,
Y. Fukami, Genistein, a specific inhibitor of tyrosine-specific receptor kinases,
J. Biol. Chem. 262 (1987) 5592–5595.
[9] Y. Graziani, E. Erikson, R.L. Erikson, The effect of quercetin on the phosphoryla-
tion activity of the Rous sarcoma virus transforming gene product in vitro and
in vivo, Eur. J. Biochem. 135 (1983) 583–589.
[10] M. Cushman, D. Nagrathnam, D.L. Burg, R.L. Geahlem, Synthesis and protein-
tyrosine kinase inhibitory activities of flavonoid analogues, J. Med. Chem. 34
(1991) 798–806.
[11] B.D.M. Cunningham, M.D. Threadgill, P.W. Groundwater, I.L. Dale, J.A. Hickman,
Synthesis and biological evaluation of a series of flavones designed as inhibitors
of protein tyrosine kinases, Anti-Cancer Drug Des. 7 (1992) 365–384.
[12] P.C. Yates, C.J. McCall, M.F.G. Stevens, Structural studies on benzothiazoles.
Crystal and molecular structure of 5,6-dimethoxy-2-(4-methoxyphenyl-
benzothiazole and molecular orbital calculations on related compounds,
Tetrahedron 47 (1991) 6493–6502.
[13] N. Hori, G. Tsukamoto, A. Imamura, M. Ohashi, T. Saito, K. Yoshino, Synthesis
and antiarthritic activity of 2-(4-methylphenyl)benzothiazoles, Chem. Pharm.
Bull. 40 (9) (1992) 2387–2390.
4. Conclusion
As the concentration of compound being tested increased,
the in vitro anticancer activity also increased. Compound HB9c
[5,6-difluoro-2-(4-methoxyphenyl)-1,3-benzothiazole] was found
to be least active amongst all the tested compounds at all the
concentration levels. It may be concluded that, compounds hav-
ing mono bromo or mono fluoro substitution on benzothiazole
moiety exhibit good anticancer activity against MCF-7 breast
cancer cells. On the other hand, difluoro substitution on ben-
zothiazole ring, reduces the activity to greater extent. Exact
correlation between methyl/methoxy substitution on 2-phenyl
ring and anticancer activity is not established. The docking score
of the synthesised compounds could not be correlated with the in
vitro anticancer activity and conclusion could not be drawn on their
exact mechanism of action. So further molecular modification on
2-phenyl-1,3-benzothiazole is required in order to arrive at more
accurate structure activity relationship with their anticancer activ-
ity on breast cancer cell lines or different EGFR crystal structure
could be selected from the PDB to study their mechanism of action.
[14] I. Hutchison, M.-S. Chua, H.L. Browne, V. Trapani, T.D. Bradshaw, A.D. Westwell,
M.F.G. Stevens, Synthesis and in vitro biological properties of fluorinated 2-(4-
aminophenyl) benzothiazoles, J. Med. Chem. 44 (2001) 1446–1455.
[15] T. Mossman, Rapid colorimetric assay for cellular growth and survival: appli-
cation to proliferation and cytotoxicity assays, J. Immunol. Methods 65 (1-2)
(1983) 55–63.
Acknowledgements
[16] J. Mendelsohn, J. Baselga, The EGF receptor family as targets for cancer therapy,
Oncogene 19 (2000) 550–6565.
[17] vLife MDS 3.0 Documentation, Tutorial: Engine Build Molecule, 2007, pp. 1–12.
[18] vLife MDS 3.0 Documentation, Tutorial: Protein Complex Optimization, 2007,
pp. 1–8.
[19] vLife MDS 3.0 Documentation, Tutorial: Engine Build Molecule, 2007, pp.
17–20.
[20] vLife MDS 3.0 Documentation, Tutorial: Engine Build Molecule, 2007, pp. 9–14.
[21] vLife MDS 3.0 Documentation, Tutorial: Biopredicta Protein Complex Opti-
mization, 2007, pp. 2–11.
We are thankful to Head, Dept. of Biotechnology, Manipal Life
Science Centre, Manipal as well as Head, Dept. of Biotechnology,
MCOPS, Manipal for helping and providing materials in performing
in vitro anticancer activity on MCF-7 cell lines. We are also thankful
to IISc, Bangalore for providing us with the NMR spectra for our
compounds in time.
[22] M.F.G. Stevens, C.J. MacCall, P. Lelievald, P. Alexander, A. Richter, D.E. Donna,
Structural studies on bioactive compounds. 23. Synthesis of polyhydroxylated
2-phenylbenzothiazoles and a comparison of their cytotoxicities and pharma-
cological properties with genistein and quercetin, J. Med. Chem. 37 (1994)
1689–1695.
References
[1] Cancer Research UK, UK Cancer Incidence Statistics by Age, January 2007
(Retrieved on 25-6-2007). http://en.wikipedia.org/wiki/cancer.
[2] WHO, Cancer, World Health Organization, February 2006 (Retrieved on 25-6-
2007) http://en.wikipedia.org/wiki/cancer.