Synthesis and in vitro anticancer activity of novel thiazacridine derivatives

Article abstract of DOI:10.1007/s00044-012-0236-2

Acridine derivatives represent a well-known class of anticancer agents that generally interfere with DNA synthesis and inhibit topoisomerase II. A series of eight new 3-acridin-9-ylmethyl-thiazolidine-2,4-dione and 3-acridin-9-ylmethyl-5-arylidene-thiazolidine-2,4-dione derivatives were synthesized. All the compounds were evaluated for their cell antiproliferation activity with the 3-(4,5-dimethyl-2-thiozolyl)-2,5-diphenyl-2H-tetrazolium bromide, MTT assay. The antiproliferative effects of the synthesized compounds were tested against several tumoral cell lines, namely SF-295 (central nervous system), HCT-8 (colon carcinoma), and MDA-MB-435 (melanoma) cells using doxorubicin as a positive control. Among the synthesized compounds, 3-acridin-9-ylmethyl-5-acridin-9-ylmethylene-thiazolidine-2,4-dione, 3-acridin-9-ylmethyl-5-(4-methoxy-benzylidene)-thiazolidine-2,4-dione, and 3-acridin-9-ylmethyl-5-(4-bromo-benzylidene)-thiazolidine-2,4-dione exhibited the most potent anticancer activity against the HCT-8 and MDA-MB-435 cell lines. After a detailed analysis of the structure of the thiazacridine molecules, we revealed the main possible interactions using the compound 3-acridin-9-ylmethyl- 5-acridin-9-ylmethylene-thiazolidine-2,4-dione as an example. The benefits of these compounds, regardless of the pharmacological target are the presence of two aromatic rings (pi systems), significant planarity (intercalating ability) and the presence of three hydrogen-bond acceptors, two of which are stronger (oxygen atoms) than the other (sulfur atom).

Full text of DOI:10.1007/s00044-012-0236-2

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-012-0236-2
ORIGINAL RESEARCH
Synthesis and in vitro anticancer activity of novel thiazacridine
derivatives
´
Marina Galdino da Rocha Pitta Erika Silva Souza Francisco Washington Araujo Barros
•
•
•
´
•
Manoel Odorico Moraes Filho Claudia O. Pessoa Marcelo Zaldini Hernandes
•
•
´
Maria do Carmo Alves de Lima Suely Lins Galdino Ivan da Rocha Pitta
•
•
Received: 27 February 2012 / Accepted: 13 September 2012
Ó Springer Science+Business Media New York 2012
Abstract Acridine derivatives represent a well-known
class of anticancer agents that generally interfere with
DNA synthesis and inhibit topoisomerase II. A series of
eight new 3-acridin-9-ylmethyl-thiazolidine-2,4-dione
and 3-acridin-9-ylmethyl-5-arylidene-thiazolidine-2,4-dione
derivatives were synthesized. All the compounds were
evaluated for their cell antiproliferation activity with the
3-(4,5-dimethyl-2-thiozolyl)-2,5-diphenyl-2H-tetrazolium
bromide, MTT assay. The antiproliferative effects of the
synthesized compounds were tested against several tumoral
cell lines, namely SF-295 (central nervous system), HCT-8
(colon carcinoma), and MDA-MB-435 (melanoma) cells
using doxorubicin as a positive control. Among the syn-
thesized compounds, 3-acridin-9-ylmethyl-5-acridin-9-ylm
ethylene-thiazolidine-2,4-dione, 3-acridin-9-ylmethyl-5-
(4-methoxy-benzylidene)-thiazolidine-2,4-dione, and 3-ac-
ridin-9-ylmethyl-5-(4-bromo-benzylidene)-thiazolidine-2,
4-dione exhibited the most potent anticancer activity
against the HCT-8 and MDA-MB-435 cell lines. After a
detailed analysis of the structure of the thiazacridine mol-
ecules, we revealed the main possible interactions using the
compound 3-acridin-9-ylmethyl-5-acridin-9-ylmethylene-
thiazolidine-2,4-dione as an example. The benefits of these
compounds, regardless of the pharmacological target are
the presence of two aromatic rings (pi systems), significant
planarity (intercalating ability) and the presence of three
hydrogen-bond acceptors, two of which are stronger
(oxygen atoms) than the other (sulfur atom).
Keywords Thiazacridine MTT assay anticancer Á
Molecular modeling
Introduction
Until the early 1980s, intense research of drug discovery
programs for cancer resulted in efficient medicinals, but
with high toxicity due to the lack of selectivity (Brana
et al., 2001; Demeunynck et al., 2001; Martinez and
Chacon-Garcia, 2005; Cummings and Smyth, 1993;
Georghiou, 1977). The design and synthesis of small
molecules that bind selectively to and cleave nucleic acids
are still major challenges in this field. These synthetic
nucleases have important applications as tools in molecular
biology and as potential therapeutic agents for the treat-
ment of cancer (Fernandez et al., 2007). Acridines and
their derivatives are well-known probes for nucleic acids as
well as being relevant in the field of drug development to
establish new chemotherapeutic agents (Ghosh et al.,
2010). The acridine ring, one such nuclease, is a potential
anticancer chromophore, which has a long history of
treatment of human diseases, particularly parasitic infec-
tions, cancer and Alzheimer’s disease (Demeunynck et al.,
2001; Denny, 2002; Chatellier and Lacomblez, 1990).
M. G. da Rocha Pitta Á M. Z. Hernandes Á
M. do Carmo Alves de Lima Á S. L. Galdino Á
I. da Rocha Pitta (&)
´
ˆ
Nucleo de Pesquisa em Inovac¸a˜o Terapeutica, Centro de
ˆ
´
Ciencias Biologicas, Universidade Federal de Pernambuco,
CEP 50670-901 Recife, Brazil
e-mail: irpitta@gmail.com
´
E. S. Souza
Universidade Tiradentes,
´
CEP 49032-490 Aracaju, Brazil
F. W. A. Barros Á M. O. Moraes Filho Á C. O. Pessoa
Departamento de Fisiologia e Farmacologia,
´
Faculdade de Medicina, Universidade Federal do Ceara,
CEP 60430-270 Fortaleza, Brazil
123

Med Chem Res
Biological activity
Most acridine derivatives interact with DNA as intercala-
tors. They represent a well-known class of multi-targeted
anticancer agents that generally interfere with DNA syn-
thesis, due to their ability to intercalate into DNA base pair
and leading to cell cycle arrest and apoptosis. Treatment of
human melanoma cell line (A375) with 9-phenyl acridine
derivatives led to lowering of mitochondrial potential,
upregulation of bax, release of cytochrome C, and activa-
tion of caspase 3 (Ghosh et al., 2012).
MTT assay
Compounds 3, 5, and 7a–f were tested for anticancer
activity in a cell toxicity assay against three human cancer
cell lines consisting of central nervous system (SF-295),
colon carcinoma (HCT-8), and melanoma (MDA-MB-
435), which were obtained from the National Cancer
Institute (Bethesda, MD, USA).
Some bis-acridines bind to DNA through intercalation
between consecutive nucleotides. The act of intercalation
induces local structural changes to the DNA, including the
unwinding of the double helix and lengthening of the DNA
strand (Ferguson and Denny, 2007; Ghosh et al., 2012;
Sondhi et al., 2010). Also, acridines are able to stabilize the
DNA-topoisomerase I and II cleavable complex, and form
the so-called ‘ternary complex’ which involves DNA, the
intercalated compound, and topoisomerase (Chilin et al.,
2009; Vispe et al., 2007). These enzymes can manipulate
DNA by changing the number of topological links between
two strands of the same or different DNA molecules
(Champoux, 2001) and are involved in many cellular
processes, such as replication, recombination, transcription,
and chromosome segregation (Corbett and Osheroff, 1993;
Wang, 1996; Nitiss, 1998).
The cells were maintained in RPMI 1640 medium sup-
plemented with 10 % fetal bovine serum, 2 mM glutamine,
100 U/mL penicillin, and 100 lg/mL streptomycin at
37 °C with 5 % CO2. The cells were grown in complete
medium 1 day before each experiment.
The cell survival were quantified by measuring the
ability of living cells to reduce the yellow dye 3-(4,
5-dimethyl-2-thiozolyl)-2,5-diphenyl-2H-tetrazolium bro-
mide (MTT) to a purple formazan product (Uchoa et al.,
2011; Denizot and Lang, 1986; Mosmann, 1983). For the
experiments, cells were seeded in 96-well plates
(0.7 9 104 cells per well). After 24 h, when the cells
reached confluence, the thiazacridine compounds (final
concentration of 25 lg/mL) were dissolved in DMSO
(0.1 %) and added to each well and incubated for 72 h.
DMSO (0.1 %) and doxorubicin (0.3 lg/mL) were used as
negative and positive controls, respectively. Thereafter, the
plates were centrifuged, and the medium was replaced with
fresh medium (150 lL) containing 0.5 mg/mL MTT. Three
hours later, the formazan product was dissolved in 150 lL
DMSO, and the absorbance was measured using a multi-
plate reader (Spectra Count, Packard, Ontario, Canada) at
595 nm. The effect of the compounds was expressed as the
percentage of inhibition of cellular growth (%GI), obtained
from the absorbance of negative control (ANC) and cells
treated (AT), according to the following formula:
%GI = 100 9 (1 - AT/ANC).
The aim of this study was to evaluate the antiprolifer-
ative effect of eight novel thiazacridine derivatives against
tumor cell lines.
Results and discussion
Chemistry
Thiazacridine derivatives were prepared by the method
summarized in Scheme 1. Initially compound 3 was
synthesized by the N-alkylation reaction of thiazolidine-
2,4-dione, compound 1, with 9-bromomethyl-acridine,
compound 2, in the presence of sodium hydroxide. The
Michael reaction of 3-acridin-9-ylmethyl-thiazolidine-2,
4-dione, compound 3, with different cyanoacrylates,
compounds 4 and 6a-f, was carried out in the presence of
piperidine, with ethanol as a solvent (Leite et al., 2007;
Mourao et al., 2005; Pitta et al., 2004, 2007; Pigatto
et al., 2011; Uchoa et al., 2011). The presence of the
arylidene proton peak in the synthesized derivatives, 5
and 7a–f, in proton nuclear magnetic resonance (1H
NMR) confirms completion of the nucleophilic addition
reaction. It is also confirmed by MS data, which contains
ions at m/z 193 and 235 in positive ESI-MS2 spectrum
indicates the presence of the acridine and thiazolidine
moiety, respectively (Fig. 1).
The potential effects on cell viability of the thiazacridine
compounds are presented in Table 1. An activity scale was
utilized to rank the cytotoxic potential of the tested samples
against each of the cell lines: inactive samples (IS, 0–35 %
inhibition), samples with low activity (LA, 36–55 % inhi-
bition), moderate activity (MA, 56–85 % inhibition) and
high activity (HA, 86–100 % inhibition).
The experiments were analyzed using the GraphPad
Prism program, and the data are reported as the mean of
two assays completed in triplicate.
Determination of IC50
Drugs concentration that inhibited cell survival by 50 %
compared with control cells (IC50) were determined using
123

Med Chem Res
Scheme 1 Synthetic pathways
of thiazacridines
R
Br
a
b
4-OCH3
4-CH3
O
O
c
d
e
4-Cl
NaOH
ETOH
NH
N
+
4-Br
S
S
N
4-SO2CH3
3-Br,4-OCH3
N
2
O
O
f
1
3
O
O
O
N
3
+
CN
N
piperidine
N
S
N
O
4
5
O
O
O
N
3
+
CN
S
N
piperidine
O
R
6 a-f
7 a-f
R
Fig. 1 Specific fragmentation
in positive ESI-MS2 for
3-acridin-9-ylmethyl-5-
arylidene-thiazolidine-2,
4-diones
+
+
N
N
C
CH2
O
H2
N
M + H = C14H11N = m/z = 193
M +H = C15H11N2O = m/z 235
also MTT assay, as described above. The compounds were
tested against four human cancer cell lines: HL-60 (pro-
myelocytic leukemia), MDA-MB-435 (melanoma), HCT-8
(colon) and SF-295 (glyoblastoma) as presented in Table 2,
all obtained from the National Cancer Institute (Bethesda,
MD, USA). The cells were plated in 96-well plates
(0.7 9 104 cells/well for adherent cells and 0.3 9 105
cells/well for suspended cells) and compounds
(0.049–25 lg/mL) dissolved in DMSO were added to each
plate well.
Table 1 Evaluation of cytotoxicity towards tumor cells (% cell
inhibition) for the synthesized compounds 3, 5, and 7(a–f)
Compounds
SF-295
HCT-8
MDA-MB-435
3
32.8 (IS)
42.0 (LA)
92.4 (HA)
86.7 (HA)
51.3 (LA)
37.8 (LA)
96.6 (HA)
64.3 (MA)
72.5 (MA)
95.2 (HA)
0.00 (IS)
95.9 (HA)
84.2 (MA)
31.2 (IS)
0.00 (IS)
85.3 (MA)
12.9 (IS)
53.0 (LA)
93.6 (HA)
5
76.7 (MA)
59.5 (MA)
29.9 (IS)
7a
7b
7c
7d
7e
7f
31.3 (IS)
62.2 (MA)
44.3 (LA)
48.0 (LA)
91.1 (HA)
In this report, we show the effect of eight acridine
derivatives on cell survival. The structures of the synthe-
sized compounds were confirmed by spectral data and
elemental analysis and they were in full agreement with the
proposed structures. The extent of inhibition of carcinoma
cell lines by these compounds is schematically presented in
Fig. 2. Among all the tested compounds, 3 and 7c exhibited
Dox
Date represent two experiments performed in triplicate. Doxorubicin
(dox) was used as a positive control
Inactive samples (IS, 0–35 %), low activity (LA, 36–55 %)
Moderate activity (MA, 56–85 %) and high activity (HA, 86–100 %)
123

Med Chem Res
them to intercalate within the double-stranded DNA
structure, thus interfering with the cellular machinery
(Belmont et al., 2007). In this way, connecting two planar
intercalating moieties to obtain a bisacridine derivative
generally increases the DNA binding affinity and the drug’s
residence times in the DNA-bound form (Antonini et al.,
2003). As expected, compound 5, which contains two
acridine nucleus, showed a promising result.
Table 2 Cytotoxicity activity of compounds for cancer cell lines
Compounds Cell lines
HL-60
SF-295
HCT-8
MDA-MB-435
3
5
[25
[25
[25
4.44
[25
4.45
[25
7.01
3.37–5.85 3.57–5.55
5.50–8.92
[25
7a
[25
[25
19.26
15.11–24.53
[25
In general, electronegative substituents at para position
on the phenyl ring contributed to the biological activity.
First the bromo group (7d) having a electron withdrawing
inductive effect and second the methoxy group (7a) having
electron donating effect
7b
7c
7d
[25
[25
[25
[25
[25
[25
[25
[25
19.49
[25
20.63
14.21–26.73
[25
18.24–23.34
[25
It is noteworthy that the position and the nature of the
substituent on the heterocyclic core are the determinants
for the biological properties observed. Therefore, it could
be concluded that the cell survival of the tumor cell lines
tested with the thiazacridine derivatives decreases in the
presence of a methoxy group (7a), or a bromo group (7d)
both at para position of the phenyl ring. However, the
increase in the steric bulk on the phenyl ring due to the
association of the bromo group (in meta) and methoxy
group (in para), as in compound 7f, did not contributed to
the activity.
7e
[25
[25
0.48
[25
7f
[25
0.02
[25
0.04
[25
0.24
Dox
0.01–0.02
0.34–0.66 0.03–0.05
0.17–0.36
Data are present as IC50 (lg/mL) values and 95 % confidence interval
obtain by nonlinear regression. Date represent two experiments per-
formed in triplicate. Doxorubicin (Dox) was used as positive control
the lowest inhibitory activity against the three carcinoma
cell lines tested, especially against the MDA-MB-435 cell
line. In addition, compounds 5, 7a, and 7d exhibited potent
inhibitory activity against the MDA-MB-435 and HCT-8
cell lines, with greater than 85 % of inhibition in the HCT-
8 cell line. The drugs concentrations that inhibited cell
growth by 50 % compared with control cells (IC50) were
under 25 lg/mL for the compounds 5, 7a, and 7d. Thus, the
compounds that had the best antitumor activity presented
the smallest values at the IC50 assays.
Molecular modeling
In order to investigate the structures and properties of the
thiazacridines, all optimized geometries were obtained
using the AM1 semi-empirical method (Dewar et al.,
1985), available in the BioMedCAChe software, using the
internal default settings for convergence criteria. In addi-
tion, some electronic properties, such as electron affinity,
ionization potential and molecular dipole moment were
The biological activity of acridines is mainly attributed
to the planarity of these aromatic structures, which allows
Fig. 2 MTT assay for three
A
B
tumor cell lines: a SF-295, b
HCT-8 and c MDA-MB-435.
Date represent two experiments
performed in triplicate.
Doxorubicin (dox) was used as
the positive control. All
compounds were tested at a
dose 25 lg/mL (Color figure
online)
C
123

Med Chem Res
Table 3 Molecular properties and pharmacokinetic parameters,
important for the good bioavailability of thiazacridine derivatives
and doxorubicin (dox): number of hydrogen-bond acceptor and donor
groups (nON and nOHNH, respectively), molecular weight (MW),
calculated octanol/water partition coefficient (miLogP), number of
rule’s violations (Nviolat), number of rotatable bonds (Nrotb),
topological polar surface area (TPSA), molecular volume (Vol), the
scores to judge the compound’s overall potential to qualify for a drug
(druglikeness and drug-score), dipole moment, electron affinity, and
ionization potential energies
Compounds Lipinski’s rule of five
nONa nOHNHa MWa
Nrotba TPSAa Vola
Druglikenessb Drug- Dipole
scoreb moment
Electron Ionization
affinity
potential
(eV)c
miLogPa Nviolata
(debye)c (eV)c
3
4
5
0
0
308.36 3.07
0
1
0
1
1
1
0
2
3
2
3
4
3
3
3
4
4
5
50.27 259.51 2.15
64.86 425.36 4.38
61.20 367.08 4.56
51.97 358.10 2.97
51.97 355.07 5.17
51.97 359.42 2.46
86.11 389.53 4.54
61.20 384.97 3.52
206.08 459.18 8.12
0.42 2.48
0.14 2.66
0.24 5.30
0.22 4.72
1.21
1.34
1.18
1.22
1.42
1.47
1.84
1.32
1.54
8.56
8.52
8.45
8.46
8.52
8.53
8.63
8.50
9.06
5
497.58 6.67d
426.50 4.96
410.50 5.35d
430.92 5.58d
475.37 5.71d
474.56 3.78
505.39d 5.7d
543.52d 0.57
7a
7b
7c
7d
7e
7f
5
0
4
0
4
0
0.2
2.95
4
0
0.17 2.72
0.24 1.98
0.17 3.89
0.52 5.49
6
0
5
12d
0
7d
dox
a
http://www.molinspiration.com/cgi-bin/properties
http://www.organic-chemistry.org/prog/peo
BioMedCAChe software
b
c
d
Parameters that violate the Lipinski’s rule of five
calculated. Then, the molecules were submitted to the
classical analysis of Lipinski et al. (1997) using the Mol-
inspiration online tool. Lipinski’s rule of five verifies
molecular features related to the optimum bioavailability of
a drug. The number of rotatable bonds also seems to be an
important descriptor for this purpose (Wenlock et al.,
2003). Other properties were also calculated, such as the
topological polar surface area (TPSA), which is a very
useful parameter for the prediction of drug transport
properties (Ertl et al., 2000), and molecular volume. In
addition, we evaluated the scores to judge the compound’s
overall potential to qualify as a drug (druglikeness and
drug-score) using the Osiris Property Explorer online sys-
tem (http://www.organic-chemistry.org/prog/peo). All of
these data are summarized in Table 3.
available chemicals (Fluka), yielding a complete list of all
available fragments. The distribution of the druglikeness
values calculated for the 15000 Fluka chemicals remain in
the range from -20 to 4 with the maximum occurrence
around the value -1 for druglikeness score. The 3,300
traded drugs stay between -13 and 10 with the maximum
around the value 2. This distribution shows that 80 % of
the traded drugs have a positive druglikeness score, while
the big majority of Fluka chemicals revealed negative
values of druglikeness. Therefore, it is promising see that
all the molecules calculated here have positive values of
druglikeness ranging from 2.15 to 5.17 values (Table 2).
The drug-score combines individual properties like drug-
likeness, cLogP, logS, molecular weight, and toxicity risks,
in one value than may be used to judge the compound’s
overall potential to qualify for a drug. The drug-score is
calculated by multiplying contributions of these individual
properties, which are described as parametrized spline
curves.
The TPSA is calculated at Molinspiration on-line, based
on the methodology published by Ertl et al. (2000) as a
sum of fragment contributions. Only oxygen- and nitrogen-
centered polar fragments are considered. While Bytheway
et al. (2008) applied TPSA analysis for drug design, Hou
and Xu (2003) have used TPSA data for ADME evaluation.
The computation of molecule volume in molinspiration
website is based on group contributions and has been
obtained by fitting the sum of fragment contributions to
real 3D volume for a training set of about twelve thousand,
mostly drug-like molecules.
Thus, while the druglikeness values are based upon the
occurrence frequency of each molecule’s fragment in
commercial drugs, the drug-score evaluates the com-
pound’s potential to qualify as a drug and is related to
topological descriptors, fingerprints of druglikeness values,
structural keys, and other properties such as cLogP, logs,
molecular weight, and toxicity behavior. Several groups
(Bernardino et al., 2008; Campos et al., 2009; Santos et al.,
2009) are using this scores (druglikeness and drug-score) to
evaluate the potential of the studied molecules to become
drugs.
From the Osiris Property Explorer online system, the
druglikeness approach is based on a list of about 5,300
distinct substructure fragments with associated scores. The
fragment list was created with traded drugs and commercially
123

Med Chem Res
According to Lipinski’s rule, the violation of more than
one of the specified criteria may decrease bioavailability.
Our results demonstrated that all of the thiazacridine
derivatives, except 3-acridin-9-ylmethyl-5-(3-bromo-4-
methoxy-benzylidene)-thiazolidine-2,4-dione, compound
7f, obey Lipinski’s rule. It is important to stress that only
the miLogP values, for five molecules (5, 7b-d, and 7f),
slightly exceeds the value 5 preconized by Lipinski. In
addition, only for molecule 7f the molecular weight
slightly passes over the limit (500). Table 2 shows that the
reference drug, doxorubicin, seems to be very unlike the
thiazacridine molecules, which is expected because of its
peculiar molecular structure.
(doxorubicin), especially compounds 3-acridin-9-ylmethyl-
5-(4-bromo-benzylidene)-thiazolidine-2,4-dione,
com-
pound 7d, and 3-acridin-9-ylmethyl-5-acridin-9-ylmethy-
lene-thiazolidine-2,4-dione, compound 5, which were very
active against the HCT-8 and MDA-MB-435 cell lines.
Materials and methods
All the synthesized compounds were analyzed with mul-
tiple analytical procedures. Melting points were determined
in a capillary tube using a Quimis apparatus. Infrared
spectra (IR) were recorded on a Bruker IFS66 spectrome-
ter. 1H NMR spectra were recorded on a Varian Plus
300 MHz spectrometer using DMSO-d6 as a solvent and
tetramethylsilane as an internal standard. The chemical
shifts are expressed in ppm, and the following abbrevia-
tions are used: s is the singlet, d the doublet, t the triplet
and m the multiplet. Mass spectra were recording using
liquid chromatography/MS (LC/MS) with a HCT Ultra
from Bruker Daltonics and were performed by electrospray
ionization in positive or negative mode. The base peak of
the MS spectrum is set to 100 (in percentage), and the
height of the others peaks are measured relative to the base
peak.
After a detailed analysis of the structure of thiazacridine
molecules, we can see in Fig. 3 possible points of inter-
actions of compound 5. This derivative present high
activity, at least against the HCT-8 and MDA-MB-435 cell
lines (see Table 1). The benefits of these compounds,
regardless of the pharmacological target, are the presence
of two aromatic rings (p systems), significant planarity
(intercalating ability) and the presence of three hydrogen-
bond acceptors, two of which are stronger (oxygen atoms)
than the other (sulfur atom).
Acridine and its derivatives are the most extensively
studied DNA intercalating agents that bind reversibly but
non-covalently to DNA (Hou and Xu, 2003). We evaluated
the molecular properties and pharmacokinetic parameters
of thiazacridine derivatives, which are considered potential
drugs for anticancer treatment. Some compounds showed
slightly better results when compared to the reference drug
Experimental
Synthesis of 3-acridin-9-ylmethyl-thiazolidine-2,4-diones
(3)
Thiazolidine-2,4-dione, compound 1, (1.0 eq) and sodium
hydroxide, previously solubilized in ethanol, were stirred
for 10 min at room temperature (25 °C). 9-Bromomethyl-
acridine, compound 2, (1.0 eq) was added, and the mixture
was stirred at 60 °C for 7 h. After completion of the
reaction, the mixture was filtered and washed with water.
The product obtained was a yellow solid. Formula:
C17H12N2O2S. Melting point (m.p.): 196–197 °C. Yield:
51 %. IR (KBr, cm-1): 2889 (–CH2–), 1750 (C=O), 1694
(C=O), 750 (C–H). H1NMR (300 MHz, DMSO-d6) d 8.42
(d, 2H, J = 8.7 Hz, Acr–H, 4 pos.), 8.17 (d, 2H,
J = 8.4 Hz, Acr–H, 1 pos.), 7.83–7.88 (m, 2H, Acr–H, 3
pos.), 7.65–7.70 (m, 2H, Acr–H, 2 pos.), 5.75 (s, 2H,
N–CH2), 4.23 (s, 2H, S–CH2). MS m/z (%): (M?H)? 309.1
(100), calculated 308; ?MS2 309.1 (56), 235 (100), 192
(98).
Fig. 3 Possible molecular points of interactions of thiazacridine
derivatives using 3-acridin-9-ylmethyl-5-acridin-9-ylmethylene-thia-
zolidine-2,4-dione, compound 5, as example. ‘‘Acceptor’’ corre-
sponds to hydrogen-bond acceptor points and ‘‘Ring Aromatic’’
corresponds to aromatic rings. Carbon, oxygen, nitrogen and sulfur
atoms are represented with gray, red, blue, and yellow colors,
respectively. This figure was generated with DS visualizer software
(Bytheway et al., 2008) (Color figure online)
General preparation of 3-acridin-9-ylmethyl-5-arylidene-
thiazolidine-2,4-dione (5 and 7a–f)
3-Acridin-9-ylmethyl-thiazolidine-2,4-dione, compound 3,
(1.0 eq) and either 3-acridin-9-yl-2-cyano-acrylic acid
123

Med Chem Res
3-Acridin-9-ylmethyl-5-(4-chloro-benzylidene)-
thiazolidine-2,4-dione (7c)
ethyl ester, compound 4, (1.0 eq) or one of the phenyl-
substituted 2-cyano-3-phenyl-acrylic acid ethyl esters,
compounds 6a–f, were refluxed in ethanol in the presence
of piperidine and heated at 50 °C for 4 h. After this period,
the mixture was filtered and washed with water and ethanol
(Pitta et al., 2004, 2007).
Yellow solid. C24H15ClN2O2S. m.p.: 229–231 °C. Yield:
47 %. IR (KBr, cm-1): 1739 (C=O), 1689 (C=O), 1608
(C=C), 750 (C–H). H1NMR (300 MHz, DMSO-d6) d 8.45
(d, 2H, J = 8.7 Hz, Acr–H, 4 pos.), 8.19 (d, 2H,
J = 8.7 Hz, Acr–H, 1 pos.), 7.93 (s, 1H, =CH), 7.83–7.88
(m, 2H, Acr–H, 3 pos.), 7.66–7.71 (m, 2H, Acr–H, 2 pos.),
7.62–7.56 (m, 4H, Ar–H), 5.93 (s, 2H, N–CH2). MS m/z
(%): (M?H)? 431.1 (100), calculated 430; ?MS2 431.1
(77), 235 (100), 193 (50).
3-Acridin-9-ylmethyl-5-acridin-9-ylmethylene-
thiazolidine-2,4-dione (5)
Yellow solid. C31H19N3O2S. m.p.: 243–245 °C. Yield:
11 %. IR (KBr, cm-1): 1759 (C=O), 1694 (C=O), 760
(C–H). H1NMR (300 MHz, DMSO-d6) d 8.75 (s, 1H,
=CH), 8.54 (d, 2H, J = 9.0 Hz, 3-Acr–H, 4 pos.), 8.21 (d,
3-Acridin-9-ylmethyl-5-(4-bromo-benzylidene)-
thiazolidine-2,4-dione (7d)
2H, J = 8.7 Hz, 5-Acr–H,
4 pos.), 8.13 (d, 2H,
J = 8.1 Hz, 3-Acr–H, 1 pos.), 8.04 (d, 2H, J = 8.4 Hz,
5-Acr–H, 1 pos.), 7.87–7.92 (m, 2H, 5-Acr–H, 3 pos.),
7.81–7.84 (m, 2H, 3-Acr–H, 3 pos.), 7.72–7.77 (m, 2H,
5-Acr–H, 2 pos.), 7.60–7.68 (m, 2H, 3-Acr–H, 2 pos.), 5.96
(s, 2H, –CH2). MS m/z (%): (M?H)? 498.2 (100), calcu-
lated 497; ?MS2 498.2 (28), 305 (99), 236 (43), 193 (100).
Yellow solid. C24H15BrN2O2S. m.p.: 222–223 °C. Yield:
24 %. IR (KBr, cm-1): 1744 (C=O), 1689 (C=O), 1608
(C=C), 750(C–H). H1NMR (300 MHz, DMSO-d6) d 8.45
(d, 2H, J = 9.0 Hz, Acr–H, 4 pos.), 8.19 (d, 2H,
J = 8.7 Hz, Acr–H, 1 pos.), 7.90 (s, 1H, =CH), 7.84–7.89
(m, 2H, Acr–H, 3 pos.), 7.70–7.73 (d, 2H, J = 8.7 Hz,
Ar–H, 3,5 pos.), 7.66–7.71 (m, 2H, Acr–H, 2 pos.), 7.51 (d,
2H, J = 8.4 Hz, Ar–H, 2,6 pos.), 5.92 (s, 2H, N–CH2). MS
m/z (%): (M?H)? 475.1 (92), (M?2?H)? 477.1 (100),
calculated 474; ?MS2 475.1 (100), 235 (98), 193 (50).
3-Acridin-9-ylmethyl-5-(4-methoxy-benzylidene)-
thiazolidine-2,4-dione (7a)
Yellow solid. C25H18N2O3S. m.p.: 225–226 °C. Yield:
52 %. IR (KBr, cm-1): 1744 (C=O), 1679 (C=O), 1593
(C=C), 756 (C–H). H1NMR (300 MHz, DMSO-d6) d 8.49
(d, 2H, J = 9.0 Hz, Acr–H, 4 pos.), 8.16 (d, 2H,
J = 8.4 Hz, Acr–H, 1 pos.), 7.90 (s, 1H, =CH), 7.81–7.89
(m, 2H, Acr–H, 3 pos.), 7.60–7.71 (m, 2H, Acr–H, 2 pos.),
7.55 (d, 2H, J = 8.7 Hz, Ar–H, 2,6 pos.), 7.08 (d, 2H,
J = 8.7 Hz, Ar–H, 3,5 pos.), 5.93 (s, 2H, N–CH2), 3.81
(s, 3H, O–CH3).). MS m/z (%): (M?H)? 427.2 (100),
calculated 426; ?MS2 427.2 (54), 235 (100), 193 (70).
3-Acridin-9-ylmethyl-5-(4-methanesulfonyl-benzylidene)-
thiazolidine-2,4-dione (7e)
Yellow solid. C25H18N2O4S2. m.p.: 227-228 °C. Yield:
28 %. IR (KBr, cm-1): 3440 (O=S=O), 1744 (C=O), 1689
(C=O), 1608 (C=C), 1142 (O=S=O). H1NMR (300 MHz,
DMSO-d6) d 8,46 (d, 2H, J = 8,4 Hz, Acr–H, 4 pos.), 8,19
(d, 2H, J = 8,1 Hz, Acr–H, 1 pos.), 8,03 (d, 2H,
J = 7.2 Hz, Ar–H, 3.5 pos.), 8,02 (s, 1H, = CH), 7,85 –
7,90 (m, 2H, Acr–H, 3 pos.), 7,83 (d, 2H, J = 8,4 Hz,
Ar–H, 2,6 pos.), 7,67–7,73 (m, 2H, Acr–H, 2 pos.), 5,94 (s,
2H, N–CH2), 3,25 (s, 3H, S–CH3). MS m/z (%): (M?H)?
475.1 (100), 383 (99), calculated 474; ?MS2 475.1 (65),
396 (100), 235 (90), 193(40), 192 (55).
3-Acridin-9-ylmethyl-5-(4-methyl-benzylidene)-
thiazolidine-2,4-dione (7b)
Yellow solid. C25H18N2O2S. m.p.: 194–195 °C. Yield:
20 %. IR (KBr, cm-1): 1729 (C=O), 1674 (C=O), 1604
(C=C), 760 (C–H). H1NMR (300 MHz, DMSO-d6) d 8.46
(d, 2H, J = 8.7 Hz, Acr–H, 4 pos.), 8.19 (d, 2H,
J = 8.7 Hz, Acr–H, 1 pos.), 7.90 (s, 1H, =CH), 7.81–7.86
(m, 2H, Acr–H, 3 pos.), 7.60–7.71 (m, 2H, Acr–H, 2 pos.),
7.47 (d, 2H, J = 8.1 Hz, Ar–H, 2,6 pos.), 7.33 (d, 2H,
J = 8.1 Hz, Ar–H, 3,5 pos.), 5.92 (s, 2H, N–CH2) 2.34
(s, 3H, –CH3). MS m/z (%): (M?H)? 411.2 (100), calcu-
lated 410; ?MS2 411.2 (48), 235 (100), 193 (39).
3-Acridin-9-ylmethyl-5-(3-bromo-4-methoxy-benzylidene)-
thiazolidine-2,4-dione (7f)
Yellow solid. C25H17BrN2O3S. m.p.: 230–232 °C. Yield:
62 %. IR (KBr, cm-1): 1730 (C=O), 1684 (C=O), 1589
(C=C), 1267 (C–O), 756 (C–H). H1NMR (300 MHz,
DMSO-d6) d 8,46 (d, 2H, J = 8,4 Hz, Acr–H, 4 pos.), 8,19
(d, 2H, J = 8,1 Hz, Acr–H, 1 pos.), 7,89 (s, 1H, =CH),
7,86–7,87 (m, 2H, Acr–H, 3 pos.), 7,85 (d, 1H, J = 3 Hz,
Ar–H, 2 pos.), 7,65–7,72 (m, 2H, Acr–H, 2 pos.), 7,58 (dd,
123

Med Chem Res
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1H, J = 8,7 and 1,8 Hz, Ar–H, 6 pos.), 7,26 (d, 1H,
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3H, O–CH3). MS m/z (%): (M?H)? 505.1 (99.5),
(M?2?H)? 507.1 (100), calculated 504; ?MS2 505.1
(14), 490 (71), 235 (100), 192 (71), 193 (72).
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Conclusion
The eight newly synthesized 3-acridin-9-ylmethyl-thiazoli-
dine-2,4-dione and 3-acridin-9-ylmethyl-5-arylidene-thia-
zolidine-2,4-dione analogues were evaluated for their
anticancer activity against the human cancer cell lines con-
sisting of central nervous system (SF-295), colon carcinoma
(HCT-8) and the melanoma (MDA-MB-435). We have
found that the acridine dimer 3-acridin-9-ylmethyl-5-acri-
din-9-ylmethylene-thiazolidine-2,4-dione
demonstrates
potent DNA binding affinity and a significant anticancer
activity against all the three cancer cell lines tested. The great
majority of the molecules presented in this paper seems to
obey the Lipinski’s rule of five regarding the bioavailability
features. In addition, the compound 3-acridin-9-ylmethyl-5-
(4-methoxy-benzylidene)-thiazolidine-2,4-dione
having
electron donating group methoxy as substituent on phenyl
ring and the compound 3-acridin-9-ylmethyl-5-(4-bromo-
benzylidene)-thiazolidine-2,4-dione having a electron with-
drawing inductive effect of the bromo group on the phenyl
ring also exhibited strong inhibition in the same cell lines.
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and survival. Modifications to the tetrazolium dye procedure
giving improved sensitivity and reliability. J Immunol Methods
89:271–277. doi:10.1016/0022-1759(86)90368-6
Acknowledgments The authors are thankful to CNPq (Conselho
Denny WA (2002) Acridine derivatives as chemotherapeutic agents.
Curr Med Chem 9:1655–1665. doi:10.2174/0929867023369277
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new general purpose quantum mechanical molecular model. J Am
Chem Soc 107:3902–3909. doi:10.1021/ja00299a024
´
Nacional de Desenvolvimento Cientıfico e Tecnologico, Brasil), the
CAPES/COFECUB (Fundac¸a˜o Coordenac¸a˜o de Aperfeic¸oamento de
´
´
Pessoal de Ensino Superior/Comite Franc¸ais d’Evaluation de la
Cooperation Universitaire avec le Bresil), the INCT_IF (Instituto
´
´
ˆ
Nacional de Ciencia e Tecnologia para Inovac¸a˜o Farmaceutica) and
the PPGIT (Programa de Pos-Graduac¸a˜o em Inovac¸a˜o Terapeutica)
ˆ
´
ˆ
Discovery Studio Visualizer v2.5.1.9167, Copyright Ó 2005–09.
Accelrys Software Inc., San Diego
for their support of the joint research program.
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surface area as a sum of fragment-based contributions and its
application to the prediction of drug transport properties. J Med
Chem 43:3714–3717. doi:10.1021/jm000942e
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