Design, synthesis and biological evaluation of trans 2-(thiophen-2-yl)vinyl heteroaromatic iodides

Article abstract of DOI:10.1016/j.bmc.2010.04.060

A modeling approach based on physico-chemical and pharmacokinetic properties, called Volsurf+, was used to design new trans 2-(thiophen-2-yl)vinyl heteroaromatic iodides with antiproliferative activity. The synthesis and in vitro antitumor tests on two cell lines (MCF-7 and LNCap) confirmed Volsurf predicted activity values. An Almond model, derived to have an overall structural insight on the above compounds, supported the validity of Volsurf and provided guidelines for the synthesis of new compounds.

Full text of DOI:10.1016/j.bmc.2010.04.060

Bioorganic & Medicinal Chemistry 18 (2010) 4516–4523
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Design, synthesis and biological evaluation of trans 2-(thiophen-2-yl)vinyl
heteroaromatic iodides
*
Cosimo G. Fortuna , Vincenza Barresi, Giuseppe Musumarra
Dipartimento di Scienze Chimiche, Università di Catania, Viale A. Doria, 6, 95125 Catania, Italy
a r t i c l e i n f o
a b s t r a c t
Article history:
A modeling approach based on physico-chemical and pharmacokinetic properties, called Volsurf+, was
used to design new trans 2-(thiophen-2-yl)vinyl heteroaromatic iodides with antiproliferative activity.
The synthesis and in vitro antitumor tests on two cell lines (MCF-7 and LNCap) confirmed Volsurf
predicted activity values. An Almond model, derived to have an overall structural insight on the above
compounds, supported the validity of Volsurf and provided guidelines for the synthesis of new
compounds.
Received 10 November 2009
Revised 20 April 2010
Accepted 21 April 2010
Available online 18 May 2010
Dedicated to Prof. Saverio Florio on the
occasion of his 70th birthday
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Molecular modeling
Antitumor compounds
Heterocycles
QSAR
1. Introduction
modifications suggested by the above molecular modeling
approach will be verified by the synthesis of the designed mole-
Previous work on the design, the synthesis and the antitumor
activity of new trans 1-heteroaryl-2-(1,3-dimethylimidazolium-2-
yl) ethylenes1 confirmed the hypothesis, based on a Molecular
Interaction Field (MIF)2,3 approach using Grid Independent
Descriptors GRIND,4 that the presence of three aromatic moieties
and of halogen atoms are the main structural features necessary
to obtain satisfactory antiproliferative activities. The synthesis,
QSAR modeling, and biological5–8 assay of vinyl and halo-furan
derivatives has been reported and their activities as antibacterials
and anticancer drugs evaluated together with their toxicity
effects.9–12 Recent work of our group on the design of new trans
2-(furan-2-yl)vinyl heteroaromatic iodides,13 using a new chemo-
informatic tool, Volsurf+, including both modeling of ADME prop-
erties in the design of new structures and the correlation 3D
molecular fields with physico-chemical and pharmacokinetic prop-
erties, was adopted for predicting the in vitro antitumor activity of
these new compounds. In this context we here report the design,
the synthesis and the biological evaluation of new trans 2-(thio-
phen-2-yl)vinyl heteroaromatic iodides. In this work, using
Volsurf+ and the previously derived model,13 we design some
new structure replacing the O atom of the pentatomic ring with
sulfur and predicting their in vitro activities. The structural
cules and by their activities evaluation tests against two tumor cell
lines, MCF-7 and LNCap.
2. Results and discussion
The structures of the new thiophene derivatives are reported in
Scheme 1. Eleven structures 4–14 previously synthesised and mod-
eled using a program called Almond based on a structural classifi-
cation were analyzed by the novel methodology called Volsurf+.
Volsurf is an automatic procedure to convert 3D molecular fields
into physico-chemically relevant molecular descriptors. Volsurf
extracts the information present in 3D molecular field maps into
few quantitative numerical descriptors, easy to understand and
interpret. Molecular recognition is achieved coupling image analy-
sis software with external chemical knowledge. Within this con-
text Volsurf selects molecular descriptors referred to molecular
size and shape, to size and shape of both hydrophilic and hydro-
phobic regions and to the balance between them. Hydrogen bond-
ing, amphiphilic moments, critical packing parameters are other
useful descriptors. The Volsurf descriptors have been presented
and explained in detail.13 The originality of Volsurf resides in the
fact that surfaces, volumes and other related descriptors can be di-
rectly obtained from 3D molecular fields and can be used for mul-
tivariate model building to correlate the 3D molecular structures
with biological behavior.14 The Volsurf methodology can also be
* Corresponding author. Tel.: +39 0957385004; fax: +39 095580138.
E-mail address: cg.fortuna@unict.it (C.G. Fortuna).
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2010.04.060

C. G. Fortuna et al. / Bioorg. Med. Chem. 18 (2010) 4516–4523
4517
CH3
N
+
+
+
N
N
N
CH3
CH3
CH3
S
S
S
X
X
X
1
2
3
X
Designation
5-Chloro
4-Phenyl
5-Phenyl
5thiophen
a
b
c
d
Scheme 1. Structure of 2-(thiophen-2yl)vinyl-heteroaromatic iodides.
applied for structure–activity relationships and molecular diversity
based on surface properties. The test set consist in 11 compounds
4–14 reported in Scheme 2, nine of which used to derive the model
for furan rings as previously published by our group,13 plus one
thiophene and one N-methylpyrrole derivatives; the above com-
pounds exhibited both good and low activities against two tumor
cell lines, MCF-7 and LNCap. The best variation of antitumor activ-
ity values for these 11 compounds was shown on MCF-7 cell line
and we decided, as a consequence, to adopt the MCF-7 values for
modeling the structures and predicting the activities of the new
compounds. The code number and the antiproliferative in vitro
activities for the 11 compounds chosen as test set are reported in
Table 1. The 3D structures of these compounds were imported, in
Mol-file, and coded as Volsurf descriptors following the procedure
described in Section 3.1. Volsurf offer a tool, called Volsurf+ de-
signer, that makes us able to design new compounds and to project
them on our test set model. This tool enables to predict the activity
of new structures by means of the scores plot projection providing
guidelines in the selection of new compounds to be synthesised.
Figure 1 reports the 11 structures of the test set and as an example
one of the new compounds (3b). The background is colored accord-
ing to the activity values; the red zone indicates higher activity and
the yellow point is the new structure. It is possible to note that the
predicted activity of 3b is higher than that of the compounds pre-
viously published. In Table 2 the predicted activities for 10 new
structures are reported.
exhibit a significant cytotoxic activity, expressed as log LC50 values
as reported in Table 3. Figure 4 reports the experimental activity
values for the new structures (yellow points) and for the test set
(black points) versus those predicted by the Volsurf model. Exper-
imental values for the designed compounds are similar or even
coincident to the predicted ones confirming the potentialities of
the Volsurf approach to plan an intelligent selection of new active
compounds to be synthesised and tested. In order to verify the
validity of Volsurf and to have an overall structural insight an
Almond model was also derived for all compounds 1–14.
PLS of ALMOND descriptors relative to compounds 1–14 affor-
ded a 5 PC model explaining 99.0% of variance, 73.7% 1st PC, 8.3%
2nd PC, 11.0% 3rd PC, 4.9% 4th PC and 1.0% 5th PC. Figure 5, the
scores plot depicting the data structure elucidated by 3 PC (already
explaining more than 90% of variance) appears a simplified graph-
ical representation for an immediate evaluation of 3D structural
MIF results for compounds 1–14 when interacting with the three
above mentioned probes. Figure 5 shows that very active com-
pounds such as 7, 8 and 14, exhibit a low value in x axis, which
can reasonably be assumed as a direction representative of antitu-
mor activities. Experimental in vitro values, reported in Table 3 for
compounds 1b, 2c, 3d and especially 3b and 3c support the overall
structural insight provided by the Almond model. Relevant infor-
mation on the changes in molecular structures in relation to their
interactions with the selected Almond probes (Dry, O, and N1) can
be obtained by an inspection of the correlograms. In the correlo-
gram for the hydrophobic probe (Fig. 6) compound 8, the most ac-
tive of the test set, shows minor interactions with respect to those
of the designed structure 3c, the most active against both tumor
cell lines, and to other derivatives containing four aromatic rings.
This consideration provides guidelines for the synthesis of new
compounds exhibiting increased interactions with probe DRY.
This result encouraged to proceed with the synthesis and
in vitro biological evaluation of the above derivatives. The synthe-
sis of trans 2-(thiophen-2-yl)vinyl heteroaromatic iodides can be
easily achieved by condensation of imidazolium, pyridinium or
quinolinium iodides with heteroaromatic aldehydes in the pres-
ence of NaOH or piperidine (see Section 3), exploiting the acidity
of the above salts
a methyls due to the electronwithdrawing effect
of the adjacent positively charged ring nitrogens.
3. Experimental
Under appropriate experimental conditions pure trans isomers
were obtained, as evidenced by the ethylenic protons J coupling
constants in the NMR spectra. The antiproliferative activity of all
the newly synthesized compounds was then tested against two tu-
mor cell lines, breast carcinoma (MCF-7) and prostate carcinoma
(LNCap). The in vitro activities, expressed as log GI50 values (see
Section 3), are recorded in Table 2. It is worth mentioning that,
in order to obtain comparable biological tests, log GI50 in Table 2
were all measured in the same experiment. The percent of growth
3.1. Computational methods
3.1.1. Volsurf+ descriptors
The interaction of molecules with biological membranes is
mediated by surface properties such as shape, electrostatic forces,
H-bonds and hydrophobicity. Therefore, the GRID15 force field was
chosen to characterize potential polar and hydrophobic interaction
sites around target molecules by the water (OH2), the hydrophobic
(DRY), and the carbonyl oxygen (O) and amide nitrogen (N1) probe.
The information contained in the MIF is transformed into a
quantitative scale by calculating the volume or the surface of the
and the inhibition exerted by different doses (0.01–100 lM) are re-
corded in Figure 2 for MCF-7 cell line and in Figure 3 for LNCap. In
addition to antiproliferative effects (log GI50), all the derivatives

4518
C. G. Fortuna et al. / Bioorg. Med. Chem. 18 (2010) 4516–4523
3
3
H3C
H3C
4
4
3'
3'
N
N
4'
5'
4'
b
b
+
+
5
5
a
a
N
N
O
O
CH3
1
CH3
Br
1
4
5
3
H3C
3
4
3'
N
4'
H3C
b
+
5
4
3'
N
+
N
a
6''
4'
N
b
O
5
5''
4''
a
CH3
1
O
CH3
Cl
CH3
1
3''
6
7
3
H3C
4
3
3'
3'
4'
N
4'
H3C
b
b
+
5
4
2''
5''
a
N
N
Br
S
3''
O
+
5
a
CH3
N
1
6''
CH3
Br
1
8
9
1'
H3C
8'
7'
5'
3
3'
4'
6'
4
N
5
H3C
b
3
a
4
5'
N
N
6
3'
+
5
a
b
N
+
N
CH3
CH3
1
CH3
1
10
11
4
5
3
5
3
a
6
3'
6
a
N
3'
N
+
b
+
b
CH3
4'
CH3
4'
1
O
1
O
5'
Br
4
12
13
5
3
b
3'
a
4'
6
N
+
O
6''
CH3
1
5''
Cl
4''
3''
14
Scheme 2. Structure of compounds for the test set.
interaction contours. The Volsurf+ procedure is as follows: (i) in the
first step, the 3D molecular field is generated from the interactions
of the OH2, the DRY, O and N1 probe around a target molecule; (ii)
the second step consists in the calculation of descriptors from the
3D maps obtained in the first step. The molecular descriptors
obtained, called Volsurf+ descriptors, refer to molecular size and
shape, to hydrophilic and hydrophobic regions and to the balance
between them, to molecular diffusion, log P, log D, to the ‘‘charge
state” descriptors, to the new 3D pharmacophoric descriptors and
to some descriptors on some relevant ADME properties. The defini-
tion of all 126 Volsurf+ descriptors is given in16–20 (case studies
with the old versions of Volsurf) and reported in detail in Table 3;
(iii) finally, chemometric tools (PCA,21 PLS22) are used to create
relationships of the Volsurf+ descriptor matrix with ADME proper-
ties. The scheme of the Volsurf+ programme steps and a detailed
definition of Volsurf+ descriptors have recently been reported.13

C. G. Fortuna et al. / Bioorg. Med. Chem. 18 (2010) 4516–4523
4519
Table 1
Activity values for compounds in the test set against MCF-7 cell line
MCF7 cell line
1c
150
100
50
3c
1d
1b
2d
3b
2b
2c
3a
3d
Compounds
log GI50 (lM)
4
5
6
7
8
ꢀ4.00
ꢀ4.40
ꢀ4.48
ꢀ5.86
ꢀ6.21
ꢀ4.84
ꢀ4.00
ꢀ4.82
ꢀ5.10
ꢀ4.15
ꢀ5.65
0
9
10
11
12
13
14
-50
-100
-8
-7
-6
-5
-4
drug concentration (uM)
Figure 2. Dose–response curves of antiproliferative activities versus MCF-7.
lncap cell line
1c
150
1d
1b
100
50
2d
2b
3a
3c
0
-50
3b
3d
2c
-100
-8
-7
-6
-5
-4
drug concentration (uM)
Figure 3. Dose–response curves of antiproliferative activities versus LNCap.
Table 3
In vitro antitumor activities, expressed as log LC50, for MCF-7 and LNCap cell lines
Cell
line
Compounds
3a 1b
2b 3b
1c 2c 3c
1d
2d 3d
MCF-7
LNCap
–
–
ꢀ4.11
ꢀ4.16
–
–
ꢀ4.75
ꢀ4.16
–
–
–
ꢀ
ꢀ4.81 ꢀ4.11
ꢀ4.96 ꢀ4.20
–
–
–
ꢀ5.19
Figure 1. Volsurf scores plot of the test set and of the new structure 3b.
performed on SGI O2 workstations (MIPS R12000 processor). The
process of ligand–receptor interaction has often been represented
with the help of the MIF.
Table 2
Predicted and in vitro activity values expressed as log GI50
compounds
,
for the designed
If a compound is known to bind a certain receptor, some of the
regions defined in its Virtual Receptor Site (VRS)4 should actually
overlap groups of the real receptor site and, therefore, at least a
subset of the VRS regions would be relevant for representing the
binding properties of the ligand. For the latter statement to be true
the VRS must have been obtained from the bioactive conformation
of the ligand and the probes used to compute it should represent
chemical groups present in the binding site. The molecular descrip-
tors presented in this work are based on the concept of VRS. Basi-
Compounds Predicted activity
values
In vitro activity
MCF-7
In vitro activity
LNCap
3a
1b
2b
3b
1c
2c
3c
1d
2d
3d
ꢀ5.68
ꢀ5.84
ꢀ5.62
ꢀ6.59
ꢀ5.67
ꢀ5.50
ꢀ6.36
ꢀ5.67
ꢀ5.81
ꢀ6.52
ꢀ5.68
ꢀ6.28
ꢀ6.00
ꢀ6.30
ꢀ5.08
ꢀ5.84
ꢀ6.25
ꢀ5.76
ꢀ5.33
ꢀ5.85
ꢀ4.98
ꢀ5.12
ꢀ4.70
ꢀ5.84
ꢀ4.47
ꢀ5.08
ꢀ6.40
ꢀ5.50
ꢀ5.22
ꢀ6.42
cally, GRIND are
a small set of variables representing the
geometrical relationships between relevant regions of the VRS
and as such are independent of the coordinate frame of the space
where the MIF is computed. The procedure for obtaining GRIND in-
volves three steps: (i) computing a set of MIF, (ii) filtering the MIF
to extract the most relevant regions that define the VRS, and (iii)
encoding the VRS into the GRIND variables.
3.2. ALMOND
MIF were obtained using the program GRID version 20.23 GRIND
were generated, analyzed, and interpreted using the program AL-
MOND version 3.0.0 [www.moldiscovery.com] a software package
developed in our group. Computations and graphical display were
3.2.1. Molecular description: GRID Force Field
The GRID program24–26 was used to describe the molecular
structures. GRID is
a computational procedure for detecting

4520
C. G. Fortuna et al. / Bioorg. Med. Chem. 18 (2010) 4516–4523
Figure 4. Experimental against Volsurf predicted values for the new structures
(yellow) and for the test set (black).
energetically favorable binding sites on molecules. The program
calculates the interactions between the molecule and a probe
group which is moved through a regular grid of points in a region
of interest around the target molecule and, at each point, the inter-
action energy between the probe and the target molecule is calcu-
Figure 6. Correlograms for compounds 1–14 with the hydrophobic probe.
lated as the sum of Lennard–Jones (ELJ), hydrogen bond (EHB
)
Figure 5. Three components PLS scores plot for Almond descriptors relative to compounds 1–14.

C. G. Fortuna et al. / Bioorg. Med. Chem. 18 (2010) 4516–4523
4521
electrostatic interactions (EEL) and, for specific probes, entropic
3.3.2. 1-Methyl-2-[(E)-2-(4-phenyl-thiophen-2-yl)-vinyl]-
contribution (EENT):
pyridinium iodide (1b)
5'
3''
5''
2''
6''
4
S
3
4''
a
5
3'
b
6
N
GRID contains a table of parameters to describe each type of atom
occurring in each of the ligand molecules. These parameters define
the strength of the Lennard–Jones, hydrogen bond and electrostatic
interactions made by an atom and are used in order to evaluate the
energy functions. GRID probes are very specific. They give precise
spatial information, and this specificity and sensitivity are an
advantage since the probes may then be representative of the
important chemical groups present in the active site provided that
the statistical method used for the analysis can distinguish between
different types of interactions.
+
CH3
1
From 1,2-dimethylpyridinium iodide (100 mg, 4.25 ꢁ 10ꢀ1 mmol) and
4-phenyl-2-thiophencarboxaldehyde (80.1 mg, 4.25 ꢁ 10ꢀ1 mmol) in
2.5 ml of refluxing ethanol. The mixture was stirred for 4 h. Yellow
needles (93 mg, 2.29 ꢁ 10ꢀ1 mmol, 55%). Mp 202–203 °C; 1H NMR
(500 MHz, DMSO-d6, 25 °C): d = 8.12 (d, J = 15.5 Hz, 1H, Ha), 7.38 (d,
J = 16 Hz, 1H, Hb), 4.35 (s, 3H, CH3), 8.48 (m, 2H, H3 e H4), 7.88 (m,
0
1H, H5); 8.88 (d, J = 6.5 Hz, 1H, H6), 8.17 (s, 1H, H3 ), 8.15 (d,
0
00
00
J = 4.0 Hz, 1H, H5 ), 7.76 (d, J = 7.5 Hz, 2H, H2 H6 ), 7.46 (t, J = 7.5 Hz,
+
00
00
00
2H, H3 e H5 ), 7.35 (t, J = 7.5 Hz, 1H, H4 ); ESI: m/z 278.28 [M ].
3.3. Compounds
3.3.3. 1,3-Dimethyl-2-[(E)-2-(4-phenyl-thiophen-2-yl)-vinyl]-
1H-imidazolium iodide (2b)
Heteroaromatic carboxaldehydes were Aldrich commercial
products. 1H NMR spectra were recorded on a Varian Unity Inova
spectrometer operating at 500 MHz, at 25 °C in (CD3)2SO using
TMS or acetone (2.225 ppm) as internal standards. The spectral
width was set to 5000 Hz, with an excitation pulse of 60°, an acqui-
sition time of 3.5 s and a digital resolution after zero-filling of
0.15 Hz/pt. NMR data were processed using MestReC software
(http://www.mestrec.com).
Electron Spray Ionization mass spectra were recorded on a
Thermo Finnigan LCQ Deca mass spectrometer equipped with a
ESI source.
2-Heteroaryl thiophene derivatives, all iodide salts, were ob-
tained by refluxing in ethanol equimolar amounts of 1,2,3-
trimethylimidazolium iodide, or 1,2-dimethylpicolinium iodide
or 1,2-dimethylquinolinium iodide and the appropriate hetero-
aromatic aldehyde in the presence of few drops 20% NaOH or
piperidine. The resulting precipitate was recrystallized from
ethanol.
3
CH3
+
4
N
a
5
S
N
b
5'
CH3
1
6''
3'
5''
4''
2''
3''
From 1,2,3-trimethylimidazolium iodide (100 mg, 4.20 ꢁ 10ꢀ1 mmol)
and 4-phenyl-2-thiophencarboxaldehyde (79.2 mg, 4.20 ꢁ 10ꢀ1 mmol)
in 2.5 ml of refluxing ethanol. The mixture was stirred for 6 h. Yellow
needles (70 mg, 1.72 ꢁ 10ꢀ1 mmol, 41%). Mp 222–224 °C; 1H NMR
(500 MHz, DMSO-d6, 25 °C): d = 7.73 (d, J = 16.5 Hz, 1H, Ha), 7.05 (d,
J = 16, 5 Hz, 1H, Hb), 3.30 (s, 6H, H1 e H3), 7.74 (s, 2H, H4 e H5), 8.12
Details on the synthetic conditions and products characteriza-
tion are reported below:
0
0
00
00
(s, 1H, H3 ), 8.09 (s, 1H, H5 ), 7.74 (m 2H, H2 H6 ), 7.46 (t, J = 8.0 Hz,
+
3.3.1. 1-Methyl-2-[(E)-2-(5-chloro-thiophen-2-yl)-vinyl]-
quinolinium iodide (3a)
00
00
00
2H, H3 e H5 ), 7.34 (t, J = 7.5 Hz, 1H, H4 ); ESI: m/z 281.28 [M ].
3.3.4. 1-Methyl-2-[(E)-2-(4-phenyl-thiophen-2-yl)-vinyl]-
quinolinium iodide (3b)
5
4
6
3
4
5
6
7
3
b
+
a
N
7
b
+
N
a
2''
3'
3''
8
3'
CH3
1
S
8
4''
5''
4'
CH3
1
S
6''
5'
Cl
From 1,2-dimethylquinolinium iodide (150 mg, 5.4 ꢁ 10ꢀ1 mmol)
From 1,2-dimethylquinolinium iodide(200 mg, 7.2 ꢁ 10ꢀ1 mmol) and
4-phenyl-2-thiophencarboxaldehyde (131.7 mg, 7.2 ꢁ 10ꢀ1 mmol) in
4 ml of refluxing ethanol. The mixture was stirred for 2 h. Brown solid
(66.16 mg, 2.016 ꢁ 10ꢀ1 mmol, 28%). Mp 200 °C; 1H NMR (500 MHz,
DMSO-d6, 25 °C): d = 8.41 (d, J = 15.5 Hz, 1H, Ha), 7.75 (d, J = 15.5 Hz,
1H, Hb), 4.55 (s, 3H, H1), 8.34 (d, J = 8 Hz, 1H, H3), 8.55 (d, J = 9 Hz,
2H, H4, H6), 7.95 (t, J = 8 Hz, 1H, H7), 8.18 (t, J = 8.5 Hz, 1H, H8), 9.04
and 5-chloro-2-thiophencarboxaldehyde (56
l
l, 5.4 ꢁ 10ꢀ1 mmol)
in 4 ml of refluxing ethanol. The mixture was stirred for 2 h. Red
solid (61.81 mg, 2.16 ꢁ 10ꢀ1 mmol, 40%). Mp 189 °C; 1H NMR
(500 MHz, DMSO-d6, 25 °C): d = 8.3 (1H, d, J = 16 Hz, Ha), 7.5 (1H,
d, J = 15.5 Hz, Hb), 4.48 (3H, s, H1), 8.31 (1H, d, J = 7 Hz, H3), 8.56
(1H, d, J = 9 Hz, H4), 8.44 (1H, d, J = 9 Hz, H6), 7.93 (1H, t,
J = 8 Hz, H7), 8.17 (1H, t, J = 7 Hz, H8), 9.01 (1H, d, J = 9 Hz, H9),
0
0
(d, J = 9 Hz, 1H, H5), 8.33 (s, 1H, H5 ), 8.29 (s, 1H, H3 ), 7.79 (d,
0
0
7.14 (1H, d, J = 3.5 Hz, H3 ), 6.54 (1H, d, J = 3.5 Hz, H4 ); ESI: m/z
00
00
00
00
J = 7.5 Hz, 2H, H2 , H6 ), 7.48 (t, J = 7.5 Hz, 2H, H3 , H5 ), 7.37 (t,
286.2 [M+].
00
+
J = 7.5 Hz, 1H, H4 ); ESI: m/z 328.2 [M ].

4522
C. G. Fortuna et al. / Bioorg. Med. Chem. 18 (2010) 4516–4523
(d, J = 8 Hz, 1H, H3), 8.53 (d, J = 9 Hz, 1H, H4), 8.54 (d, J = 9.5 Hz, 1H,
H6), 7.94 (t, J = 7.5 Hz, 1H, H7), 8.17 (t, J = 7.5 Hz, 1H, H8), 9.02 (d,
3.3.5. 1-Methyl-2-[(E)-2-(5-phenyl-thiophen-2-yl)-vinyl]-
pyridinium iodide (1c)
0
J = 9 Hz, 1H, H5), 7.75 (d, J = 3.5 Hz, 1H, H4 ), 7.84 (d, J = 3.5 Hz, 1H,
0
00
00
00
00
H3 ), 7.79 (d, J = 7.5 Hz, 2H, H2 , H6 ), 7.50 (t, J = 7.5 Hz, 2H, H3 , H5 ),
5''
4''
+
00
7.42 (t, J = 7 Hz, 1H, H4 ); ESI: m/z 328.2 [M ].
6''
3''
3.3.8. 2-((E)-2-[2,20]Bithiophenyl-5-yl-vinyl)-1-methyl-
pyridinium iodide (1d)
4
2''
5
S
3
a
4'
5''
6
b
3'
N
S
4''
+
CH3
1
3''
1
S
From 1,2-dimethylpyridinium iodide (100 mg, 4.25 ꢁ 10ꢀ1 mmol) and
5-phenyl-2-thiophencarboxaldehyde (80.1 mg, 4.25 ꢁ 10ꢀ1 mmol) in
2.5 ml of refluxing ethanol. The mixture was stirred for 8 h. Brown
needles (93 mg, 2.29 ꢁ 10ꢀ1 mmol, 55%). Mp 202–203 °C; 1H NMR
(500 MHz, DMSO-d6, 25 °C): d = 8.16 (d, J = 15.5 Hz, 1H, Ha), 7.26 (d,
J = 16 Hz, 1H, Hb), 3.30 (s, 3H, CH3), 8.47 (d, 2H, H3 H5), 7.86 (dd,
CH3
a
4'
+
N
6
5
3'
b
3
4
From 1,2-dimethylpyridinium iodide (100 mg, 4.25 ꢁ 10ꢀ1 mmol) and
2,20-bithiophen-5-carboxaldehyde (82.4 mg, 4.25 ꢁ 10ꢀ1 mmol) in
2 ml of refluxing ethanol with two drops of piperidine. The mixture
was stirred for 8 h. Red solid (60.1 mg, 1.46 ꢁ 10ꢀ1 mmol, 34%). Mp
200–202 °C; 1H NMR (500 MHz, DMSO-d6, 25 °C): d = 8.14 (d,
J = 15.5 Hz, 1H, Ha), 7.21 (d, J = 16 Hz, 1H, Hb), 4.34 (s, 3H, CH3), 8.45
(m, 2H, H3 H5), 7.84 (m, 1H, H4); 8.86 (d, J = 6 Hz, 1H, H6), 7.64 (d,
0
0
1H, H4); 8.87 (d, J = 5.5 Hz, 1H, H6), 6.67 (d, 2H, H3 H4 ), 8.15 (d,
0
00
00
J = 4.0 Hz, 1H, H5 ), 7.75 (d, J = 8 Hz, 2H, H2 H6 ), 7.48 (t, J = 7.5 Hz,
+
00
00
00
2H, H3 e H5 ), 7.40 (t, J = 7.5 Hz, 1H, H4 ); ESI: m/z 278.35 [M ].
3.3.6. 1,3-Dimethyl-2-[(E)-2-(5-phenyl-thiophen-2-yl)-vinyl]-
1H-imidazolium iodide (2c)
0
0
00
J = 4.5 Hz, 1H, H3 ), 7.60 (d, J = 4.0 Hz, 1H, H4 ), 7.45 (d, J = 4 Hz, 1H, H5 ),
+
00
00
5''
7.48 (d, J = 3.5 Hz,1H,H3 ), 7.07 (t, J = 4.0 Hz,1H,H4 ); ESI: m/z284.2 [M ].
6''
4''
3.3.9. 2-((E)-2-[2,20]Bithiophenyl-5-yl-vinyl)-1,3-dimethyl-1H-
3
imidazolium iodide (2d)
CH3
3''
S
+
a
4
2''
N
5
4'
3
b
N
3'
CH3
+
4
CH3
N
1
a
5
From 1,2,3-trimethylimidazolium iodide (100 mg, 4.20 ꢁ 10ꢀ1 mmol)
and 5-phenyl-2-thiophencarboxaldehyde (79.2 mg, 4.20 ꢁ 10ꢀ1
mmol) in 2.5 ml of refluxing ethanol. The mixture was stirred in ab-
sence of light for 5 h. Yellow needles (76 mg, 1.86 ꢁ 10ꢀ1 mmol,
44%). Mp 222–224 °C; 1H NMR (500 MHz, DMSO-d6, 25 °C): d = 7.74
S
5''
4''
N
S
b
CH3
1
3' 4'
3''
From 1,2,3-trimethylimidazolium iodide (100 mg, 4.20 ꢁ 10ꢀ1 mmol)
and 2,20-bithiophen-5-carboxaldehyde (81.5 mg, 4.20 ꢁ 10ꢀ1 mmol)
in 3 ml of refluxing ethanol. The mixture was stirred in absence of light
for 7 h. Brown solid (84 mg, 2.03 ꢁ 10ꢀ1 mmol, 48%). Mp 215–217 °C;
1H NMR (500 MHz, DMSO-d6, 25 °C): d = 7.742 (d, J = 16.5 Hz, 1H, Ha),
6.91 (d, J = 16.5 Hz, 1H, Hb), 3.74 (s, 6H, H1 e H3), 7.62 (s, 2H, H4 e H5),
(d, J = 16.5 Hz, 1H, Ha), 6.94 (d, J = 16.5 Hz, 1H, Hb), 3.30 (s, 6H, H1
H3), 7.73 (s, 2H, H4 e H5), 7.63 (d, J = 3.5 Hz, 1H, H3 ), 7.61 (d,
J = 3.5 Hz, 1H, H4 ), 7.72 (d, J = 6.0 Hz, 2H, H2 H6 ), 7.47 (t, J = 7.5 Hz,
2H, H3 e H5 ), 7.38 (t, J = 7.5 Hz, 1H, H4 ); ESI: m/z 281.35 [M ].
e
0
0
00
00
+
00
00
00
0
0
7.54 (d, J = 4 Hz, 1H, H3 ), 7.41 (d, J = 3.5 Hz, 1H, H4 ), 7.61 (dd,
3.3.7. 1-Methyl-2-[(E)-2-(5-phenyl-thiophen-2-yl)-vinyl]-
quinolinium iodide (3c)
00
00
J = 5.0 Hz, 1H, H3 ), 7.15 (dd, J = 5.0 Hz, 1H, H4 ), 7.45 (dd, J = 3.5 Hz,
1H, H5 ); ESI: m/z 287.24 [M ].
+
00
3.3.10. 2-((E)-2-[2,20]Bithiophenyl-5-yl-vinyl)-1-methyl-
6
4
3
quinolinium iodide (3d)
7
8
a
+
N
3'
4
5
4'
b
6
7
3
CH3
1
9
S
a
2''
+
N
3'
6''
4'
b
3''
8
CH3
1
S
5''
4''
3''
4''
From 1,2-dimethylquinolinium iodide (200 mg, 7.2 ꢁ 10ꢀ1 mmol) and
5-phenyl-2-thiophencarboxaldehyde (131.7 mg, 7.2 ꢁ 10ꢀ1 mmol) in
4 ml of refluxing ethanol with one drop of piperidine. The mixture
was stirred for 2 h. Dark red solid (67 mg, 2.02 ꢁ 10ꢀ1 mmol, 30%).
Mp 230 °C; 1H NMR (500 MHz, DMSO-d6, 25 °C): d = 8.46 (d,
J = 15.5 Hz, 1H, Ha), 7.61 (d, J = 15.5 Hz, 1H, Hb), 4.51 (s, 3H, H1), 8.33
S
5''
From 1,2-dimethylquinolinium iodide (200 mg, 7.2 ꢁ 10ꢀ1 mmol) and
2,20-bithiophen-5-carboxaldehyde (135.9 mg, 7.2 ꢁ 10ꢀ1 mmol) in
4 ml of refluxing ethanol with one drop of piperidine. The mixture

C. G. Fortuna et al. / Bioorg. Med. Chem. 18 (2010) 4516–4523
4523
was stirred for 7 h. Dark solid (153 mg, 4.6 ꢁ 10ꢀ1 mmol, 67%). Mp
Acknowledgment
229 °C; 1H NMR (500 MHz, DMSO-d6, 25 °C): d = 8.43 (d, J = 15.5 Hz,
1H, Ha), 7.57 (d, J = 15.5 Hz, 1H, Hb), 4.50 (s, 3H, H1), 8.332 (d,
J = 8 Hz, 1H, H3), 8.52 (d, J = 9 Hz, 1H, H4), 8.53 (d, J = 9 Hz, 1H, H6),
7.93 (t, J = 7.5 Hz, 1H, H7), 8.16 (t, J = 7 Hz, 1H, H8), 9 (d, J = 9 Hz, 1H,
We thank the University of Catania for financial support.
References and notes
0
00
H5), 7.77 (d, J = 4 Hz, 1H, H3 ), 7.68 (d, J = 5 Hz, 1H, H3 ), 7.53 (d,
1. Ballistreri, F. P.; Barresi, V.; Benedetti, P.; Caltabiano, G.; Fortuna, C. G.; Longo,
M. L.; Musumarra, G. Bioorg. Med. Chem. 2004, 12, 1689.
2. Goodford, P. J. Med. Chem. 1985, 28, 849.
0
00
J = 3.5 Hz, 1H, H4 ), 7.54 (d, J = 4.5 Hz, 1H, H5 ), 7.18 (t, J = 5 Hz, 1H,
00
+
H4 ); ESI: m/z 334 [M ].
3. Cramer, R. D., III; Patterson, D. E.; Bunce, J. J. Am. Chem. Soc. 1988, 110, 5959.
4. Pastor, M.; Cruciani, G.; McLay, I.; Pickett, S.; Clementi, S. J. Med. Chem. 2000, 43,
3233.
4. Biological essays
5. González-Díaz, H.; Agüero, G.; Cabrera, M. A.; Molina, R.; Santana, L.; Uriarte,
E.; Delogu, G.; Castañedo, N. Bioorg. Med. Chem. Lett. 2005, 15, 551.
6. González-Diaz, H.; Marrero, Y.; Hernandez, I.; Bastida, I.; Tenorio, E.; Nasco, O.;
Uriarte, E.; Castanedo, N.; Cabrera, M. A.; Aguila, E.; Marrero, O.; Morales, A.;
Perez, M. Chem. Res. Toxicol. 2003, 16, 1318.
7. González-Díaz, H.; Cruz-Monteagudo, M.; Molina, R.; Tenorio, E.; Uriarte, E.
Bioorg. Med. Chem. 2005, 13, 1119.
8. Estrada, E. Mutat. Res. 1998, 420, 67.
9. González-Díaz, H.; Olazábal, E.; Santana, L.; Uriarte, E.; González-Díaz, Y.;
Castañedo, N. Bioorg. Med. Chem. 2007, 15, 962.
10. González-Díaz, H.; Torres-Gómez, L. A.; Guevara, Y.; Almeida, M. S.; Molina, R.;
Castañedo, N.; Santana, L.; Uriarte, E. J. Mol. Model. 2005, 11, 116.
11. González-Díaz, H.; Tenorio, E.; Castañedo, N.; Santana, L.; Uriarte, E. Bioorg.
Med. Chem. 2005, 13, 1523.
4.1. Human cell lines (MCF-7 and LNCap)
Human prostate adenocarcinoma cells (LNCap) were grown in
RPMI 1640. Human mammary adenocarcinoma (MCF-7) were
grown in Dulbecco’s MEM (DMEM), 1.0 g/L
ium was supplemented with 10% (v/v) heat-inactivated fetal bo-
vine serum, 2 mM -alanyl- -glutamine, penicillin–streptomycin
(50 U–50 g for ml) and incubated at 37 °C in humidified atmo-
D-glucose. Each med-
L
L
l
sphere of 5% CO2, 95% air. The culture medium was changed twice
a week.
12. González Borroto, J. I.; Pérez Machado, G.; Creus, A.; Marcos, R. Mutagenesis
2005, 20, 193.
13. Fortuna, C. G.; Barresi, V.; Berellini, G.; Musumarra, G. Bioorg. Med. Chem. 2008,
16, 4150.
14. Doddareddy, M. R.; Cha, J. H.; Cho, Y. S.; Koh, H. Y.; Yoo, K. H.; Kim, D. J.; Pae, A.
N. Bioorg. Med. Chem. 2005, 13, 3339.
4.2. Treatment with antitumor agents and MTT colorimetric
assay
Human cancer cell line (5 ꢁ 103 cells/0.33 cm2) was plated in
96-well plates ‘‘Nunclon TM Microwell TM” (Nunc) and were incu-
bated at 37 °C. After 24 h, cells were treated with the compounds
15. Carosati, E.; Sciabola, S.; Cruciani, G. J. J. Med. Chem. 2004, 47, 5114.
16. Cruciani, G.; Crivori, P.; Carrupt, P. A.; Testa, B. J. Mol. Struct. 2000, 503, 17.
17. Crivori, P.; Cruciani, G.; Carrupt, P. A.; Testa, B. J. Med. Chem. 2000, 43, 2204.
18. Cruciani, G.; Meniconi, M.; Carosati, E.; Zamora, I.; Mannhold, R.. In Methods
and Principles in Medicinal Chemistry; van de Waterbeemd, H., Lennernäs, H.,
Artursson, P., Eds.; Wiley-VCH, 2003; Vol. 18, p 406.
19. Berellini, G.; Cruciani, G.; Mannhold, R. J. Med. Chem. 2005, 48, 4389.
20. Mannhold, R.; Berellini, G.; Carosati, E.; Benedetti, P.. In Molecular Interaction
Fields; Cruciani, G., Mannhold, R., Kubinyi, H., Folkers, G., Eds.; Wiley-VCH
Verlag GmbH & Co. KGaA: Weinheim, 2006; Vol. 27, p 173.
21. Wold, S.; Sjöström, M. In Chemometrics: Theory and Application, Kowalski, B. R.,
Eds.; ACS Symposium Series, Washington, 1977; pp 243.
22. Wold, S.; Albano, C.; Dunn, W. J., III; Edlund, U.; Esbensen, K.; Geladi, P.;
Hellberg, S.; Johansson, E.; Lindberg, W.; Sjöström, M. In Chemometrics,
Kowalski, B. R., Ed., 1984; pp 17.
23. GRID v. 20 Molecular Discovery Ltd.
24. Goodford, P. J. J. Med. Chem. 1985, 28, 849.
(final concentration 0.01–100 lM). Untreated cells were used as
controls. Microplates were incubated at 37 °C in humidified atmo-
sphere of 5% CO2, 95% air for 3 days and then cytotoxicity was mea-
sured with colorimetric assay based on the use of tetrazolium salt
MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bro-
mide).27 The results were read on a multiwell scanning spectro-
photometer (Multiscan reader), using a wavelength of 570 nm.
Each value was the average of 8 wells (standard deviations were
less than 20%). The GI50 value was calculated according to NCI:
thus, GI50 is the concentration of test compound where
100 ꢁ (T ꢀ T0)/(C ꢀ T0) = 50 (T is the optical density of the test well
after a 48-h period of exposure to test drug; T0 is the optical den-
sity at time zero; C is the control optical density).
25. Boobbyer, D. N. A.; Goodford, P. J.; Mcwhinnie, P. M.; Wade, R. C. J. Med. Chem.
1989, 32, 1083.
26. Wade, R.; Clerk, K. J.; Goodford, P. J. J. Med. Chem. 1993, 36, 140.
27. Mosmann, T. J. J. Immunol. Methods 1983, 65, 55.