Design and synthesis of trans 2-(furan-2-yl)vinyl heteroaromatic iodides with antitumour activity

Article abstract of DOI:10.1016/j.bmc.2007.12.042

A new molecular modelling approach based on physico-chemical and pharmacokinetic properties, called Volsurf plus, was used to design new heterocyclic compounds with antiproliferative activity. The synthesis and in vitro antitumour tests on a breast carcinoma cell line (MCF7) confirmed VOLSURF predicted activity values.

Full text of DOI:10.1016/j.bmc.2007.12.042

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry 16 (2008) 4150–4159
Design and synthesis of trans 2-(furan-2-yl)vinyl
heteroaromatic iodides with antitumour activity
Cosimo Gianluca Fortuna,a,* Vincenza Barresi,a Giuliano Berellinib
and Giuseppe Musumarraa,*
a
`
Dipartimento di Scienze Chimiche, Universita di Catania, Viale A. Doria, 6, 95125 Catania, Italy
b
`
Laboratorio di Chemiometria e Chemioinformatica, Dipartimento di Chimica, Universita degli Studi di Perugia,
Via Elce di Sotto, 8, 06100 Perugia, Italy
Received 1 October 2007; revised 14 December 2007; accepted 20 December 2007
Available online 25 December 2007
Abstract—A new molecular modelling approach based on physico-chemical and pharmacokinetic properties, called Volsurf plus,
was used to design new heterocyclic compounds with antiproliferative activity. The synthesis and in vitro antitumour tests on a
breast carcinoma cell line (MCF7) confirmed VOLSURF predicted activity values.
ꢀ 2008 Elsevier Ltd. All rights reserved.
1. Introduction
availability. Very simple heuristic rules concerning cut-
offs on molecular weight, number of hydrogen bond
donor/acceptors and lipophilicity only represent crude
guidelines working well at the lead-finding stage, but less
suitable to establish structure–property relationships in
lead optimization.13 Other frequently used descriptor
schemes are topological indices, but their interpretation
in terms of molecular structures is not straightforward.
In the present work, a high-throughput chemoinformatic
approach, called Volsurf plus, including both modelling
of ADME properties in the design of new structures and
the correlation 3D molecular fields with physico-chemi-
cal and pharmacokinetic properties, will be adopted for
predicting the in vitro antitumour activity of these new
compounds. The structural modifications suggested by
the above molecular modelling approach will be verified
by the synthesis of the designed molecules and their
activity evaluation tests.
Previous work on the design, the synthesis and the antit-
umour activity of new trans 1-heteroaryl-2-(1,3-dimethy-
limidazolium-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 syn-
thesis, QSAR modelling and biological5–8 assay of vinyl
and halo-furan derivatives has been reported and their
activities as antibacterials and anticancer drugs evalu-
ated together with their toxicity effects.9–12 Following
these leads, we here report the design of new trans 2-(fur-
an-2-yl)vinyl heteroaromatic iodides, with pyridinium,
imidazolium and quinolinium as ethylene-linked hetero-
aromatic cations and halo substituted benzenes in the
5-position of the furan ring, with the aim to optimize
the antitumour activities. It is well known that an in-
creased drug–receptor interaction does not necessarily
imply an increase in biological activity. Therefore, it is
also very important to design new structures which exhi-
bit ADME (Adsorption, Distribution, Metabolism, and
Elimination) properties warranting an acceptable bio-
2. Results and discussion
The structures of compounds previously synthesized and
of new compounds are reported in Scheme 1. Ten struc-
tures previously modelled using a program called Al-
mond based on a structural classification were selected
to derive another model by a novel methodology called
Volsurf plus. Volsurf is an automatic procedure to con-
vert 3 D molecular fields into physico-chemically rele-
vant molecular descriptors. In the standard procedure,
Keywords: Molecular modelling; Antitumour compounds; Heterocy-
cles; QSAR.
*
Corresponding authors. Tel.: +39 0957385004; fax: +39 095580138;
e-mail: cg.fortuna@unict.it
0968-0896/$ - see front matter ꢀ 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2007.12.042

C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
4151
CH3
N
+
+
+
N
N
N
CH3
CH3
CH3
O
O
O
X
X
X
1
2
3
X
Designation
H
a
b
c
d
e
f
g
h
i
Methyl
Bromo
2-Chlorophenyl
3-Chlorophenyl
4-Chlorophenyl
4-Bromophenyl
2,5-Dichlorophenyl
2,4-Dichlorophenyl
3,4-Dichlorophenyl
j
3-Chloro-4-methoxyphenyl
4-Chloro-2-nitrophenyl
k
l
2-Chloro-5-(trifluoromethyl)-phenyl
m
Scheme 1. Structure of 2-(furan-2yl) vinyl-heteroaromatic iodides.
the interaction fields with a water probe and a hydro-
phobic probe are calculated for all molecules in the data-
set. The basic concept of Volsurf is to extract the
information present in 3D molecular field maps into
few quantitative numerical descriptors, which are easy
to understand and interpret. Molecular recognition is
achieved using image analysis software coupled with
external chemical knowledge. Within this context Vol-
surf selects the most appropriate descriptors and param-
eterization according to the type of 3D maps under
study. The molecular descriptors obtained refer to
molecular size and shape, to size and shape of both
hydrophilic and hydrophobic regions and to the balance
between them. Hydrogen bonding, amphiphilic mo-
ments and critical packing parameters are other useful
descriptors. The Volsurf descriptors are presented and
explained in detail in Section 3. The originality of Vol-
surf resides in the fact that surfaces, volumes and other
related descriptors can be directly obtained from 3D
molecular fields with simple computational algorithms.
Moreover, Volsurf descriptors can be easily obtained
for small, medium and large molecules, as well as for
biopolymers such as DNA sequences, peptides and pro-
teins. No parameterization is required and the descrip-
tors can be used for multivariate model building to
correlate the 3D molecular structures with biological
behaviour.14 The VolSurf methodology can also be ap-
plied for structure-activity relationships and molecular
diversity based on surface properties. The computa-
tional procedure is fully automated and quite fast. The
ten structures present in the first model, test set, contain
a differently substituted furan ring linked by an ethylenic
bond to an imidazolium or pyridinium salt, apart from
one structure with a furan ring between two vinylpyrid-
inium salts (PF2) already selected for comparison
because it is among the most active compounds previ-
ously synthesized by our group.15 The code number
and the antiproliferative in vitro activities for the ten
compounds chosen as test set are reported in Table 1.
The 3D structures of compounds (1c, 1d, 1g, 2a–d, 2g)
and for 2,6-di-[2-(furan-2-yl)vinyl]pyridinium iodide
(PF2) were imported, in Mol-file, and coded as VOL-
SURF descriptors following the procedure described
in Section 3.1.
This new methodology offers a new tool, called Vol-
surf plus designer, that makes us able to design new
compounds and to project them on our test set model.
Structural modifications in the design of new struc-
tures, with the aim to improve the antitumour activi-
ties, consisted in the insertion of new substituents on
the benzene linked in the 5-position of the furan ring
and introducing a new salt as the quinolinium one.
The scores plot projection provided guidelines in the
selection of new structures to be synthesised. In Figure
1 the scores plot with the ten structures of the test set
Table 1. Activity values for compounds in the test set
Compound
LogGI50 (lM)
1a
1c
ꢀ4.15
ꢀ5.10
ꢀ5.65
ꢀ6.00
ꢀ4.00
ꢀ4.48
ꢀ4.40
ꢀ5.86
ꢀ6.21
ꢀ5.60
1d
1g
2a
2b
2c
2d
2g
PF2

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C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
um iodides with heteroaromatic aldehydes in the pres-
ence of NaOH (see Section 3), exploiting the acidity
of the above salts’ a methyls due to the electron-with-
drawing effect of the adjacent positively charged ring
nitrogens.
Under appropriate experimental conditions pure trans
isomers were obtained, as evidenced by the ethylenic
protons’ J coupling constants in the NMR spectra.
The anti-proliferative activity of all the newly synthe-
sized compounds was then tested against a tumour cell
line, breast carcinoma (MCF7). The in vitro activities,
expressed as logGI50 values (see Section 3), are recorded
in Table 3, together with that of 2,6-di-[2-(furan-2-
yl)vinyl]pyridinium iodide (PF2), the most active com-
pound in previous in vitro tests, also reported for com-
parisons. It is worth mentioning that, in order to obtain
comparable biological tests, log GI50 in Table 3 were all
measured in the same experiment. The percent of growth
and the inhibition exerted by different doses (0.01–
100 lM) are recorded in Figure 2. In addition to anti-
proliferative effects (log GI50), all the derivatives exhibit
a significant cytotoxic activity, expressed as logLC50 val-
ues; in particular compound 1h shows the best value of
both logGI50 and logLC50.
Figure 1. Scores plot of the test set and of the new structure 3f.
Table 2. Predicted activity values for the designed compounds
Compound
Predicted activity values
Figure 3 reports the experimental activity values for the
new structures (yellow points) and for the test set (black
points) versus those predicted by the ten structures mod-
el. Experimental values for the designed compounds are
even higher than the predicted ones confirming the
validity of the Volsurf approach and that the presence
of three aromatic moieties and of halogen atoms are
the main structural features which determine an increase
in the antiproliferative activities.
3c
2e
3l
ꢀ5.57
ꢀ5.84
ꢀ6.31
ꢀ5.56
ꢀ6.23
ꢀ5.65
ꢀ5.52
ꢀ5.92
ꢀ6.14
ꢀ6.40
ꢀ5.93
ꢀ6.70
ꢀ6.24
ꢀ5.66
ꢀ6.64
ꢀ6.18
2f
1j
1m
2k
1f
2h
3k
1e
3f
1i
1k
3e
1h
3. Experimental
3.1. Computational methods
3.1.1. VolSurf+ descriptors. The interaction of mole-
cules with biological membranes is mediated by sur-
face properties such as shape, electrostatic forces,
H-bonds and hydrophobicity. Therefore, the GRID16
force field was chosen to characterize potential polar
and hydrophobic interaction sites around target mol-
ecules 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 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
and one of the new compounds are reported as an
example. The background was coloured according to
the activity values; the red zone indicates the higher
activity and the yellow point is the new structure. It
is possible to see that the predicted activity of 3f is
higher than those of the compounds previously pub-
lished. In Table 2 the predicted activities for 16 new
structures are reported.
To verify the above hypothesis we proceeded with the
synthesis and in vitro biological evaluation of the
above derivatives. The synthesis of trans 2-(furan-2-
yl)vinyl heteroaromatic iodides can be achieved by
condensation of imidazolium, piridinium or quinolini-
Table 3. In vitro antitumour activities, expressed as logGI50, for MCF7 cell line
Compounds/ 3c
1f
2f
3f
1e
2e
3e
1h
2h
1i
1j
1k
2k
3k
3l
1m
PF2
cell line
MCF7
ꢀ5.50 ꢀ6.66 ꢀ6.20 ꢀ6.90 ꢀ6.87 ꢀ5.76 ꢀ7.24 ꢀ7.42 ꢀ6.72 ꢀ6.90 ꢀ6.42 ꢀ7.18 ꢀ6.65 ꢀ6.72 ꢀ5.76 ꢀ6.48 ꢀ5.60

C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
4153
150
100
50
1m
1e
2e
1i
3c
2c
2h
3f
1h
3e
1f
0
2f
-8
-7
-6
-5
-4
-50
-100
3l
1k
2k
1j
3k
PF2
drug concentration (uM)
Figure 2. Dose–response curves of antiproliferative activities versus MCF7.
Figure 3. Experimental against predicted values for the new structures (yellow) and for the test set (black).
Figure 4. The sequence of steps in chemometric analyses using VolSurf+ descriptors is summarized.
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 molec-
ular size and shape, to hydrophilic and hydrophobic
regions and to the balance between them, to molecu-

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C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
lar diffusion, logP, logD, to the ‘charge state’
descriptors, to the new 3D pharmacophoric descrip-
tors and to some descriptors on some relevant
ADME properties. The definition of all 126 VolSurf+
descriptors is given in17–21 (case studies with the old
versions of VolSurf) and reported in detail in Table
1; (iii) finally, chemometric tools (PCA,22 PLS23) are
used to create relationships of the VolSurf+ descrip-
tor matrix with ADME properties. A scheme of the
VolSurf+ program steps is reported in Figure 4.
For the detailed definition of Volsurf+ descriptors,
see Table 4.
3.2.2. trans 2-[2-[5-(4-Chlorophenyl)furan-2-yl]vinyl]-1-
methylpyridinium iodide (1f).
8
9
10
11
7
O
5
4
6
Cl
-
I
+
N
1
3
CH3
2
From 1,2-Dimethylpyridinium iodide (176.3 mg,
0.750 mmol) and 5-(4-chlorophenyl)furfural (155.0 mg,
0.750 mmol) in 4 ml ethanol at rt; the mixture was stir-
red for 30 min. Orange solid (257.0 mg, 80.9%); mp
249–251 ꢁC; dH (500 MHz; DMSO-d6, TMS): 4.39 (s,
3H, H1), 7.18 (d, 1H J = 3.5 Hz, H8), 7.31 (d, 1H,
J = 3.5 Hz, H9), 7.38 (d, 1H, J = 15.5 Hz, H6), 7.57 (d,
2H, J = 9 Hz, H10), 7.86 (t,1H, J = 6.5 Hz, H3), 7.88
(d,1H, J = 16 Hz, H7), 7.97 (d, 2H, J = 9 Hz, H11),
8.44–8.50 (m, 2H, H4/5), 8.88 (d, 1H, J = 6.5 Hz, H2);
MS: M+ Cl35 = 296.3, M+ Cl37 = 298.3.
3.2. Compounds
Heteroaromatic carboxaldehydes were Aldrich commer-
cial products.
1H NMR spectra were recorded on a VarianUnity 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 60ꢁ, an acquisition time of 3.5 s and a
digital resolution after zero-filling of 0.15 Hz/pt.
3.2.3. trans 2-[2-[5-(4-Chlorophenyl)furan-2-yl]vinyl]-1,3-
dimethylimidazolium iodide (2f).
6
5
7
8
Electron Spray Ionisation mass spectra were recorded
on a Thermo Finnigan LCQ Deca mass spectrometer
equipped with an ESI source.
4
O
H3C
3
N
Cl
-
I
+
N
2
1
CH3
2-Heteroaryl furan derivatives, all iodide salts, were ob-
tained by refluxing in ethanol equimolar amounts of
1,2,3-trimethylimidazolium iodide, or 1,2-dimethylpico-
linium iodide or 1,2-dimethyl quinolinium iodide and
the appropriate heteroaromatic aldehyde in the presence
of few drops of 20% NaOH. The resulting precipitate
was recrystallized from ethanol.
From 1,2,3-Trimethylimidazolium iodide (178.6 mg,
0.750 mmol) and 5-(4-chlorophenyl)furfural (155.0 mg,
0.750 mmol) in 4 ml ethanol at rt; the mixture was stir-
red for 3 d. Yellow solid (110.6 mg, 34.6%); mp 263–
265 ꢁC; dH (500 MHz; DMSO-d6, TMS): 3.94 (s, 6H,
H2), 7.06 (d, 1H, J = 16 Hz, H3), 7.08 (d, 1H,
J = 3 Hz, H5), 7.26 (d,1H, J = 3.5 Hz, H6), 7.47 (d,
1H, J = 16.5 Hz, H4), 7.56 (d(sex), 1H, J = 8.5 Hz,
H7), 7.74 (s, 2H, H1), 7.94 (d(sex), 1H, J = 8.5 Hz,
H8); MS: M+ Cl35 = 299.3, M+ Cl37 = 301.3.
Details on the synthetic conditions and products charac-
terization are reported below:
3.2.1. trans 2-[2-(5-bromo-furan-2-yl)vinyl]-1-methylquin-
olinium iodide (3c).
3.2.4. trans 2-[2-[5-(4-Chlorophenyl)furan-2-yl]vinyl]-1-
methylquinolinium iodide (3f).
6
4
7
3
10
11
b
a
12
8
3'
N
+
9
13
9
I
4'
O
7
CH3
O
6
8
Cl
-
1
Br
I
+
N
1
5
CH3
From 5-bromo-furan aldehyde (0.098 g, 0.560 mmol)
and 1,2-dimethylquinolinium iodide (0.15 g,
2
4
3
0.560 mmol), 3 ml ethanol, one drop NaOH. The solu-
tion was refluxed for two hours. Green needles (68 mg,
29%); mp 210 ꢁC; dH (500 MHz; DMSO-d6, TMS):
8.03 (d, 1H, J = 16 Hz, Ha), 7.53 (d,1H, J = 15.5 Hz,
Hb), 4.51 (s,3H, H1), 8.34 (d,1H, J = 7 Hz, H3), 8.54
(d,1H, J = 9 Hz, H4), 8.48 (d,1H, J = 9 Hz, H6), 7.94
(t,1H, J = 8 Hz, H7), 8.17 (t,1H, J = 7 Hz, H8), 9.03
(d, 1H, J = 9 Hz, H9), 7.24 (d,1H, J = 3.5 Hz, H30),
6.94 (d,1H, J = 3.5 Hz, H40) ; MS: M+ Br79 = 314.2;
M+ Br81 = 316.2.
From 1,2-Dimethylquinolinium iodide (213.8 mg,
0.750 mmol) and 5-(4-chlorophenyl)furfural (155.0 mg,
0.750 mmol) in 4 ml ethanol at rt; the mixture was stir-
red for 45 min. Red-brown solid (243.8 mg, 68.7%);
mp 224–226 ꢁC; dH (500 MHz; DMSO-d6, TMS): 4.57
(s, 3H, H1), 7.34 (d, 1H, J = 4 Hz, H10), 7.39 (d, 1H,
J = 4 Hz, H11), 7.60 (d(sex), 2H, J = 8.5 Hz, H12), 7.71
(d, 1H, J = 15.5 Hz, H9), 7.94 (t, 1H, J = 7.25 Hz, H4),
8.04 (d(sex), 2H, J = 9 Hz, H13), 8.16 (d, 1H,

C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
4155
Table 4. Detailed definition of Volsurf+ descriptors
Size and shape descriptors
3
represents the water-excluded volume (in A ), i.e. the volume enclosed by the water-accessible surface computed
˚
Molecular volume (V)
Molecular surface (S)
Rugosity (R)
at a repulsive value of +0.2 kcal/mol.
2
represents the accessible surface (in A ) traced out by a water probe interacting at +0.2 kcal/mol when a water
˚
molecule rolls over the target molecule.
is a measure of molecular wrinkled surface; it represents the ratio of volume/surface. The smaller the ratio, the
larger the rugosity.
Molecular globularity (G)
is defined as S/Sequiv with Sequiv = surface area of a sphere of volume V, where S and V are the molecular surface
and volume described above, respectively. Globularity is 1.0 for perfect spherical molecules. It assumes values
greater than 1.0 for real spheroidal molecules. Globularity is also related to molecular flexibility.
The Flex descriptor represents the maximum flexibility of a molecule. It is the result of the average of the
differences between the maximum and minimum distance of every atom with the others searched on 50 random
conformers). Flex_RB descriptor is the ratio between Flex and the number of rotable bonds.
Flexibility parameters (Flex, Flex_RB)
Descriptors of hydrophilic regions
Hydrophilic volumes (W1 - W8)
Capacity factors(CW1 - CW8)
describe the molecular envelope which is accessible to and attractively interacts with water molecules. The
volume of this envelope varies with the level of interaction energies. Hydrophilic descriptors computed from
molecular fields of ꢀ0.2 to ꢀ1.0 kcal/mol account for polarizability and dispersion forces; descriptors from
molecular fields of ꢀ1.0 to ꢀ6.0 kcal/mol account for polar and H-bond donor-acceptor regions.
represent the ratio of the hydrophilic surface over the total molecular surface. In other words, it is the hydrophilic
surface per surface unit. Capacity factors are calculated at eight different energy levels, the same levels used to
compute the hydrophilic volumes.
Descriptors of hydrophobic regions
Hydrophobic volumes (D1–D8)
GRIDa uses a probe called DRY to generate 3D lipophilic fields. In analogy to hydrophilic regions, hydrophobic
regions may be defined as the molecular envelope generating attractive hydrophobic interactions. VolSurf
computes hydrophobic descriptors at eight different energy levels adapted to the usual energy range of
hydrophobic interactions (from ꢀ0.2 to ꢀ1.6 kcal/mol).
Capacity factors(CD1 - CD8)
represent the ratio of the hydrophobic surface over the total molecular surface. In other words, it is the
hydrophobic surface per surface unit. Capacity factors are calculated at eight different energy levels, the same
levels used to compute the hydrophobic volumes.
Differences of the Hydrophobic volumes
The difference between the maximum conformational hydrophobic volumes and the hydrophobic volumes (D1–
D8) of the the imported 3D structure calculated at the eight levels of energy.
(DD1 - DD8)
INTEraction enerGY (= INTEGY) moments
INTEGY moments (IW1 - IW4, ID1 - ID4)
express the unbalance between the centre of mass of a molecule and the barycentre of its hydrophilic or
hydrophobic regions. When referring to hydrophilic regions, integy moments (IW1-IW4) are vectors pointing
from the centre of mass to the centre of the hydrophilic regions: high integy moments indicate a clear
concentration of hydrated regions in only one part of the molecular surface, small indicate that the polar moieties
are either close to the centre of mass or they balance at opposite ends of the molecule, so that their resulting
barycentre is close to the centre of the molecule. When referring to hydrophobic regions, integy moments
measure the unbalance between the centre of mass of a molecule and the barycentre of the hydrophobic regions.
Descriptors of H-bond donor / acceptor regions
H-bond donor volumes (WO1 - WO6)
H-bond acceptor volumes (WN1 - WN6)
GRIDa uses a probe called O (carbonylic oxygen) to generate 3D H-bond donor fields. In analogy to hydrophilic
regions, H-bond donor regions may be defined as the molecular envelope generating attractive H-donor
interactions. VolSurf computes H-bond donor descriptors at six different energy levels adapted to the usual energy
range of interactions (from ꢀ1 to ꢀ6 kcal/mol).
GRID15 uses a probe called N1 (amide nitrogen) to generate 3D H-bond acceptor fields. In analogy to H-bond
donor regions, H-bond acceptor volumes may be defined as the molecular envelope generating attractive H-bond
acceptor interactions. VolSurf+ computes H-bond acceptor descriptors at six different energy levels adapted to the
usual energy range of interactions (from ꢀ1 to ꢀ6 kcal/mol).
Mixed descriptors
Hydrophilic-Lipophilic balance (HL1, HL2)
Amphiphilic moments (A)
It is the ratio between the hydrophilic regions measured at ꢀ3 and ꢀ4 kcal/mol and the hydrophobic regions
measured at ꢀ0.6 and ꢀ0.8 kcal/mol. The balance describes which effect dominates in the molecule, or if they are
roughly equally balanced.
is defined as a vector pointing from the centre of the hydrophobic domain to the centre of the hydrophilic
domain. The vector length is proportional to the strength of the amphiphilic moment, and it may determine the
ability of a compound to permeate a membrane.
Critical packing parameter(CP)
defines a ratio between the hydrophilic and lipophilic part of a molecule. In contrast to the hydrophilic-lipophilic
balance, critical packing refers just to molecular shape. It is defined as: volume(lipophilic part)/
[(surface(hydrophilic part)(length of lipophilic part)] Lipophilic and hydrophilic calculations are performed at
ꢀ0.6 and ꢀ3.0 kcal/mol, respectively. Critical packing is a good parameter to predict molecular packing such as
in micelle formation, and may be relevant in solubility studies in which the melting point plays an important role.
is not computed from 3D-molecular fields. Polarizability is an estimate of the average molecular polarizability,
calculated according to Miller.25 This method is based on the structure of the compounds (and not any molecular
field) and is therefore independent of the number and type of probes used. The correlation between the
experimental molecular polarizability and the polarizability calculated with VolSurf for more than 300 chemicals
is very good (r = 0.99).
Polarizability (POL)
Diffusivity (DIFF)
computed using a modified Stokes–Einstein equation controls the dispersion of chemical in water fluid.
is simply computed by summing the atomic weights.
Molecular weight (MW)
LogP 1-octanol/water (LOGP n-Oct)
The logarithm of the partition coefficient between 1-octanol and water is computed via a linear equation derived
by fitting GRID-derived atom type to experimental data on n-octanol/water partition coefficients.
The logarithm of the partition coefficient between cyclohexane and water is computed via a linear equation
derived by fitting GRID-derived atom types to experimental data on cyclohexane/water partition coefficients.
(continued on next page)
LogP cyclohexane/water (LOGP c-Hex)

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C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
Table 4 (continued)
LogD (LgD5 - LgD10)
The logarithm of the coefficient of diffusion between 1-octanol and water is computed via the sum of
the logP and the fraction of every species at pH 5, 6, 7, 7.5, 8, 9 and 10.
Polar and Hydrophobic Surface Areas (PSA,
HSA, PSAR, PHSAR)
The Polar Surface Area (PSA) is calculated via the sum of polar GRID atom types contributions,
while the Hydrophobic Surface Area is calculated via the sum of hydrophobic GRID atom types
contributions. PSAR is the ratio between the polar surface area (PSA) and the Surface (S), while
PHSAR is the ratio between the polar surface area (PSA) and the hydrophobic surface area (HSA).
Charge State descriptors
Number of Charged Centers (NCC)
It reports the number of charged centres present in the imported molecule, according to its charge
state at the chosen pH.
Available Uncharged Species
(AUS7.4)
This parameter represent the available uncharged species at pH 7.4 and it is calculated as the
logarithm of the percentage of the unionised species at pH 7.4 + 2 (AUS7.4 = log(% unionised) + 2).
The percentage of unionised species is calculated at pH 4, 5, 6, 7, 8, 9 and 10.
% Unionised species (%FU4 - %FU10)
3D pharmacophoric descriptors (TOPP)
Dry, H-bond donor, H-bond acceptor
These parameters represent 3D pharmacophoric descriptors based on the TOPP (Triplets Of
and mixed Dry, H-bond donor and acceptor
3D-triplet pharmacophoric areas (DRDRDR,
DRDRAC, DRDRDO, DRACAC,
Pharmacophoric Points) descriptors. At first the atoms (points) of a structure are classified as Dry,
H-bond donor and H-bond acceptor, then all possible triplet of distances between these atoms are
generated. The VolSurf+ 3D pharmacophoric descriptors are the maximum conformational area of
the triangles derived from every following class of pharmacophoric triplets: DRDRDR refers to the
Dry-Dry-Dry triplet, DRDRAC refers to the Dry-Dry-Acceptor triplet, DRDRDO refers to the
Dry-Dry-Donor triplet, DRACAC refers to the Dry-Acceptor-Acceptor triplet, DRACDO refers to
the Dry-Acceptor-Donor triplet, DRDODO refers to the Dry-Donor-Donor triplet, ACACAC
refers to the Acceptor-Acceptor-Acceptor triplet, ACACDO refers to the Acceptor-Acceptor-Donor
triplet, ACDODO refers to the Acceptor-Donor-Donor triplet, DODODO refers to the Donor-
Donor-Donor triplet.
DRACDO, DRDODO, ACACAC,
ACACDO, ACDODO, DODODO)
ADME model descriptors
Intrinsic solubility (SOLY)
is computed via a linear equation derived by fitting VolSurf+ descriptors to the logarithm of
experimental intrinsic solubility (mole/Litre).
Solubility at various pH (LgS3 - LgS11)
Solubility profiling coefficients (L0lgS - L4lgS)
They represent the logarithm of solubilities computed at various pHs starting from the intrinsic
solubility (mole/Litre). (the used pH values are 3, 4, 5, 6, 7, 7.5, 8, 9, 10, 11).
These coefficients represent how much the pH-dependent solubility profile is closer to be correlated
by Legendre polynomials of grade 0, 1, 2, 3 and 4, respectively. These parameters are useful to
distinguish compounds that present similar solubility but different pH-dependent profile or vice
versa, like in the presence of bases, acids, zwitterions or multiprotic compounds.
is computed via a linear equation derived by fitting VolSurf+ descriptors to experimental data on
CACO2 cells permeability.
CACO2 permeability (CACO2)
Skin permeability (SKIN)
is computed via a linear equation derived by fitting VolSurf+ descriptors to experimental data on
skin permeability.
% of protein binding (PB)
is computed via a linear equation derived by fitting VolSurf+ descriptors to experimental data on
percentage of protein binding.
Volume of Distribution (VD)
High Throughput Screening Flag (HTSFlag)
is computed via a linear equation derived by fitting VolSurf+ descriptors to ꢀlog of experimental
data on volume of distribution (Litre/Kg).
This parameter has value 0 as default. It is active (the value is set to 1) when three of the following
conditions of the descriptors are satisfied: D3 > 80; LOGP > 5; PB > 99; SOLY > ꢀ 6 and
CD2 > 0.28.
J = 15.5 Hz, H8), 8.18 (t(sex), 1H, J = 7.75 Hz, H3), 8.33
(d(quar), 1H, J = 8.25 Hz, H7), 8.54 (d, 1H, J = 9 Hz,
H5), 8.54 (d, 1H, J = 10 Hz, H6), 9.01 (d, 1H, J = 9 Hz,
H2); MS: M+ Cl35 = 346.3, M+ Cl37 = 348.3.
7.87 (t, 1H, J = 6.5 Hz, H3), 7.88 (d, 1H, J = 15.5 Hz,
H7), 7.92 (d, 1H, J = 7.5 Hz, H10), 8.02 (s/t, 1H,
J = 1.75 Hz, H13), 8.45–8.50 (m, 2H, H4/5), 8.89 (d, 1 H,
J = 6.5 Hz, H2); MS: M+ Cl35 = 296.3, M+ Cl37 = 298.3.
3.2.5. trans 2-[2-[5-(3-Chlorophenyl)furan-2-yl]vinyl]-1-
methylpyridinium iodide (1e).
3.2.6. trans 2-[2-[5-(3-Chlorophenyl)furan-2-yl]vinyl]-1,3-
dimethylimidazolium iodide (2e).
8
9
6
5
10
7
11
7
8
4
O
5
O
H3C
4
6
12
3
9
N
13
-
10
-
I
+
I
N
+
1
3
N
Cl
Cl
CH3
2
1
CH3
2
From 1,2-Dimethylpyridinium iodide (176.3 mg,
0.750 mmol) and 5-(3-chlorophenyl)furfural (155.0 mg,
0.750 mmol) in 4 ml ethanol at rt; the mixture was stirred
for 30 min. Orange needle-shaped solid (221.5 mg,
69.7%); mp 208–210 ꢁC; dH (500 MHz; DMSO-d6,
TMS): 4.40 (s, 3H, H1), 7.18 (d, 1H, J = 3.5 Hz, H8), 7.39
(d, 1H, J = 3 Hz, H9), 7.42 (d, 1H, J = 16 Hz, H6), 7.46
(d/m, 1H, J = 8 Hz, H12), 7.54 (t, 1H, J = 8 Hz, H11),
From 1,2,3-Trimethylimidazolium iodide (178.6 mg,
0.750 mmol) and 5-(3-chlorophenyl)furfural
(155.0 mg, 0.750 mmol) in 4 ml ethanol at rt; the mix-
ture was stirred for 3 d. Yellow solid (99.4 mg,
31.1%); mp 208–210 ꢁC; dH (500 MHz; DMSO-d6,
TMS): 3.95 (s, 6H, H2), 7.09 (d, 1H, J = 3.5 Hz,
H5), 7.11 (d, 1H, J = 16 Hz, H3), 7.34 (d, 1H,
J = 4 Hz, H6), 7.44 (d, 1H, J = 8 Hz, H9), 7.48 (d,

C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
4157
From 1,2,3-Trimethylimidazolium iodide (178.6 mg,
0.750 mmol) and 5-(2,5-dichlorophenyl)furfural
1H, J = 16.5 Hz, H4), 7.52 (d, 1H, J = 8 Hz, H8), 7.75
(s, 2H, H1), 7.89 (d, 1H, J = 8 Hz, H7), 7.99 (d, 1H,
J = 3.75 Hz, H10); MS: M+ Cl35 = 299.3, M+
Cl37 = 301.3.
(180.8 mg, 0.750 mmol) in 4 ml ethanol at rt; the mixture
was stirred for 27 h. Light yellow solid (106 mg, 30.7%);
mp 250 ꢁC; dH (500 MHz; DMSO-d6, TMS): 3.95 (s, 6H,
H2), 7.16 (d, 1H, J = 3.5 Hz, H5), 7.17 (d, 1H,
J = 16.5 Hz, H3), 7.45 (d, 1H, J = 3.5 Hz, H6), 7.49
(dd, 1H, J = 8.5 Hz, H8), 7.51 (d, 1H, J = 16.5 Hz,
H4), 7.65 (d, 1H, J = 9 Hz, H9), 7.76 (s, 2H, H1), 8.12
(d, 1H, J = 2 Hz, H7); MS: M+ Cl35 = 333.3, M+
Cl35Cl37 = 335.3, M+ Cl37 = 337.2 .
3.2.7. trans 2-[2-[5-(3-Chlorophenyl)furan-2-yl]vinyl]-1-
methylquinolinium iodide (3e).
10
11
12
13
14
9
O
7
6
8
15
-
I
+
3.2.10. trans 2-[2-[5-(2,4-Dichlorophenyl)furan-2-yl]vi-
nyl]-1-methylpyridinium iodide (1i).
N
1
Cl
5
CH3
2
4
8
9
3
10
11
7
From 1,2-Dimethylquinolinium iodide (213.8 mg,
0.750 mmol) and 5-(3-chlorophenyl)furfural (155.0 mg,
0.750 mmol) in 4 ml ethanol at rt; the mixture was stir-
red for 40 min. Red-brown solid (117.6 mg, 33.1%);
mp 212–214 ꢁC; dH (500 MHz; DMSO-d6, TMS): 4.58
(s, 3H, H1), 7.34 (d, 1H, J = 3.5 Hz, H10), 7.46 (d, 1H,
J = 4 Hz, H11), 7.49 (d, 1H, J = 8.5 Hz, H14), 7.56 (t,
1H, J = 8 Hz, H13), 7.75 (d, 1H, J = 15.5 Hz, H9), 7.95
(t, 1H, J = 7.5 Hz, H4), 7.99 (d, 1H, J = 8 Hz,H12),
8.10 (s/t, 1H, J = 1.75 Hz, H15), 8.16 (d, 1H,
J = 16 Hz, H8), 8.18 (t/m, 1H, J = 8 Hz, H3), 8.34 (d,
1H, J = 7.25 Hz, H7), 8.54 (d, 1H, J = 9 Hz, H5), 8.57
(d, 1H, J = 9 Hz, H6), 9.02 (d, 1H, J = 9 Hz, H2); MS:
M+ Cl35 = 346.3, M+ Cl37 = 348.3.
O
5
4
6
Cl
-
Cl
I
12
+
N
1
3
CH3
2
From 1,2-Dimethylpyridinium iodide (176.3 mg,
0.750 mmol) and 5-(2,4-dichlorophenyl)furfural
(180.8 mg, 0.750 mmol) in 4 ml ethanol at rt; the mixture
was stirred for 35 min. Yellow solid (303.4 mg, 88.3%);
mp 240–242 ꢁC; dH (500 MHz; DMSO-d6, TMS): 4.85
(s, 3H, H1), 7.65 (d, 1H, J = 3.5 Hz, H8), 7.99 (d, 1H,
J = 16 Hz, H6), 7.94 (d, 1H, J = 3.5 Hz, H9), 8.07 (d,
1H, J = 8.5 Hz, H10), 8.18 (d, 1H, J = 15.5 Hz, H7),
8.23 (d, 1H, J = 2 Hz, H12), 8.31 (t, 1H, J = 7.25 Hz,
H3), 8.68 (d, 1H, J = 9 Hz, H11), 8.81 (d, 1H, J = 7 Hz,
H5), 8.93 (t, 1H, J = 7.75 Hz, H4), 9.08 (d, 1H,
J = 6 Hz, H2); MS: M+ Cl35 = 330.3, M+ Cl37 = 332.3.
3.2.8. trans 2-[2-[5-(2,5-Dichlorophenyl)furan-2-yl]vinyl]-
1-methylpyridinium iodide (1h).
9
8
10
3.2.11. trans 2-[2-[5-(3,4-Dichlorophenyl)furan-2-yl]vi-
nyl]-1-methylpyridinium iodide (1j).
Cl
7
O
5
8
9
11
6
4
3
10
-
Cl
12
I
+
11
7
N
1
O
CH3
2
5
4
6
12
Cl
-
From 1,2-Dimethylpyridinium iodide (0.1763 g,
0.75 mmol) and 5-(2,5-dichlorophenyl)furfural (0.181
mg, 0.75 mmol) in 4 ml ethanol at rt; the mixture was
stirred for 30 min. Yellow solid (279.8 mg, 81,5%); mp
235–237 ꢁC; dH (500 MHz; DMSO-d6, TMS): 4.40 (s,
3H, H1), 7.24 (d, 1H, J = 3.5 Hz, H8), 7.48 (d, 1H,
J = 16 Hz. H6), 7.48 (d, 1H, J = 3.5 Hz, H9), 7.51 (dd,
1H, J = 8.5 Hz, H11), 7.66 (d, 1H, J = 9 Hz, H12),
7.89 (d, 1 H, J = 16 Hz, H7), 7.89-7.91 (m,1H, H3),
8.14 (d, 1H, J = 2.5 Hz, H10), 8.48–8.52 (m, 2H, H4/
5), 8.92 (d, 1H, J = 6 Hz, H2); MS: M+ Cl35 = 330.3,
M+ Cl35Cl37 = 332.3.
I
+
N
1
3
Cl
CH3
2
From 1,2-Dimethylpyridinium iodide (176.3 mg,
0.750 mmol) and 5-(3,4-dichlorophenyl)furfural
(180.8 mg, 0.750 mol) in 4 ml ethanol at rt; the mixture
was stirred for 25 min. Orange, needle-shaped solid
(290.1 mg, 84.4%)%); mp 232–234 ꢁC; dH (500 MHz;
DMSO-d6, TMS): 4.86 (s, 3H, H1), 7.61 (d, 1H,
J = 3.5 Hz, H8), 7.66 (d, 1H, J = 4 Hz, H9), 7.90 (d,
1H, J = 15.5 Hz, H6), 8.17 (d, 1H, J = 15.5 Hz, H7),
8.21 (d, 1H, J = 8.5 Hz, H10), 8.30 (t, 1H, J = 6.75 Hz,
H3), 8.38 (dd, 1H, J = 8.5 Hz, H11), 8.68 (d, 1H,
J = 2 Hz, H12), 8.81 (d, 1H, J = 8 Hz, H5), 8.92 (d, 1H,
J = 7.25 Hz, H4), 9.09 (d, 1H, J = 6.5 Hz, H2) ; MS:
[M+-CH3]=330.2, M+ Cl35 Cl37 = 332.2.
3.2.9. trans 2-[2-[5-(2,5-Dichlorophenyl)furan-2-yl]vinyl]-
1,3-dimethylimidazolium iodide (2h).
6
5
7
Cl
3.2.12.
furan-2-yl]vinyl]-1-methylpyridinium iodide (1k).
trans
2-[2-[5-(3-Chloro-4-methoxyphenyl)
4
O
H3C
3
N
8
-
Cl
I
9
From 1,2-Dimethylpyridinium iodide (176.3 mg,
0.750 mmol) and 5-(3-chloro-4-methoxyphenyl)furfural
+
N
2
1
CH3

4158
C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
8
1
9
(dd, 1H, J = 8.75 Hz, H12), 8.12 (d, 1H, J = 2.5 Hz,
10
H15), 8.15 (d, 1H, J = 15.5 Hz,H9), 8.16 (t, 1H,
J = 8 Hz, H3), 8.31 (d, 1H, J = 7.5 Hz, H7), 8.52
(d, 1H, J = 9.5 Hz, H5), 8.54 (d, 1H, J = 8.5 Hz,
H6), 8.97 (d, 1H, J = 9 Hz, H2); MS: M+
Cl35 = 329.2, M+ Cl37 = 331.2.
11
7
O
5
12
4
6
13
OMe
-
I
+
N
3
Cl
CH3
2
3.2.15. trans 2-[2-[5-(4-Chloro-2-nitrophenyl)furan-2-
yl]vinyl]-1-methylquinolinium iodide (3l).
(177.5 mg, 0.750 mmol) in 4ml ethanol at rt; the mixture
was stirred for 40 min. Orange solid (315.6 mg, 92.8%);
mp 224-226 ꢁC; dH (500 MHz; DMSO-d6, TMS): 3.92 (s,
3H, H12), 4.38 (s, 3H, H1), 7.15 (d, 1H, J = 3.5 Hz, H8),
7.23 (d, 1H, J = 3.5 Hz, H9), 7.27 (d, 1H, J = 8.5 Hz,
H11), 7.34 (d, 1H, J = 16 Hz, H6), 7.83 (t, 1H, J = 6 Hz,
H3), 7.86 (d, 1H, J = 15.5 Hz, H7), 7.89 (d(quar), 1H,
J = 8.5 Hz, H10), 8.03 (d, 1H, J = 2 Hz, H13), 8.42–8.48
(m, 2H, H4/5), 8.86 (d, 1 H, J = 6.5 Hz, H2); MS:
[M+-CH3]=331.2, M+ Cl35 = 326.2, M+ Cl37 = 328.2.
10
11
12
13
9
O
7
6
8
Cl
-
O2N
I
+
14
N
1
5
CH3
2
4
3
From 1,2-Dimethylquinolinium iodide (213.8 mg,
0.750 mmol) and 5-(4-chloro-2-nitrophenyl)furfural
(188.7 mg, 0.750 mmol) in 4 ml ethanol at rt; the mix-
ture was stirred for 60 min. Light brown solid
(326.1 mg, 83.9%); mp 238–240 ꢁC; dH (500 MHz;
DMSO-d6, TMS): 5.02 (s, 3H, H1), 7.63 (d, 1H,
J = 3.5 Hz, H10), 7.73 (d, 1H, J = 3.5 Hz, H11), 8.03
(d, 1H, J = 15.5 Hz, H8), 8.34 (d, 1H, J = 15.5 Hz,
H9), 8.35 (d(quar), 1H, J = 8 Hz, H12), 8.48–8.53
(m, 3H, H4/13/14), 8.74 (d, 1H, J = 7.75 Hz, H3),
8.79 (d, 1H, J = 8.5 Hz, H7), 8.82 (d, 1H,
J = 8.5 Hz, H5), 8.91 (d, 1H, J = 9 Hz, H6), 9.41 (d,
1H, J = 8.5 Hz, H2); MS: M+ Cl35 = 391.2, M+
Cl37 = 393.2.
3.2.13. trans 2-[2-[5-(3-Chloro-4-methoxyphenyl)furan-2-
yl]vinyl]-1,3- dimethylimidazolium iodide (2k).
6
5
7
8
4
O
H3C
9
3
N
10
OMe
-
I
+
N
Cl
2
1
CH3
From 1,2,3-Trimethylimidazolium iodide (178.6 mg,
0.750 mmol) and 5-(3-chloro-4-methoxyphenyl)furfural
(177.5 mg, 0.750 mmol) in 4ml ethanol at rt; the
mixture was stirred for
5
d. Yellow solid
3.2.16. trans 2-[2-[5-(2-Chloro-5-trifluoromethylphenyl)furan-
2-yl]vinyl]-1-methylpyridinium iodide (1m).
(168.2 mg, 49.1%); mp 252–254 ꢁC; dH (500 MHz;
DMSO-d6, TMS): 3.92 (s, 3H, H9), 3.94 (s, 6H,
H2), 7.04 (d, 1H, J = 16.5 Hz, H3), 7.05 (d, 1H,
J = 3.5 Hz, H5), 7.18 (d, 1H, J = 3.5 Hz, H6), 7.26
(d, 1H, J = 9 Hz, H8), 7.43 (d, 1H, J = 16.5 Hz,
H4), 7.73 (s, 2H, H1), 7.86 (dd, 1H, J = 8.75 Hz,
H7), 8.00 (d, 1H, J = 2 Hz, H10); MS: M+
Cl35 = 376.2, M+ Cl37 = 378.2.
9
8
10
CF3
7
5
4
O
11
6
1
-
3
12
+
I
Cl
N
2
CH3
From 5-[2-chloro-5(trifluoromethyl)phenyl]furan alde-
hyde (0.425 mmol) and 1,2,3 trimethylimidazolium io-
dide (0.1008 g, 0.425 mmol), 3 ml ethanol, one drop
NaOH. The solution was refluxed for one hour. Yel-
low needles (83 mg, 39%); mp 237 ꢁC; dH (500 MHz;
DMSO-d6, TMS): 4.38 (s, 3H, H1), 7.30 (d, 1H,
J = 4 Hz, H8), 7.47 (d, 1H, J = 15.5 Hz, H6), 7.54
(d, 1H, 3J = 4 Hz, H9), 7.79 (dd, 1H, J = 8.75 Hz,
H12), 7.88-7.91 (m, 2H, H3/11), 7.91 (d, 1H,
J = 16 Hz, H7), 8.31 (s, 1H, H10), 8.47–8.52 (m,
2H, H4/5), 8.91 (d, 1H, J = 6.5 Hz, H2); MS:
M+ = 364.3.
3.2.14. trans 2-[2-[5-(3-Chloro-4-methoxyphenyl)furan-2-
yl]vinyl]-1-methylquinolinium iodide (3k).
10
11
12
13
9
O
7
14
6
8
15
OMe
-
I
+
CH31
N
Cl
5
2
4
3
From 1,2-Dimethylquinolinium iodide (213.8 mg,
0.750 mmol) and 5-(3-chloro-4-methoxyphenyl)furfural
(177.5 mg, 0.750 mmol) in 4ml ethanol at rt; the
mixture was stirred for 60 min. Dark violet solid
(236.2 mg, 62.5%); mp 218–220 ꢁC; ; dH (500 MHz;
DMSO-d6, TMS): 3.94 (s, 3H, H14), 4.55 (s, 3H,
H1), 7.29–7.32 (m, 3H, H10/11/13), 7.68 (d, 1H,
J = 15.5 Hz, H8), 7.92 (t, 1H, J = 7.5 Hz H4), 7.97
4. Biological essays
4.1. Human cell line (MCF7)
Human mammary adenocarcinoma (MCF7) were
grown in Dulbecco’s MEM (DMEM), 1.0 g/l D-glucose.
Each medium was supplemented with 10% (vol/vol)

C. G. Fortuna et al. / Bioorg. Med. Chem. 16 (2008) 4150–4159
4159
´
6. Gonzalez-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.
heat-inactivated foetal bovine serum, 2 mM L-Alanyl-L-
Glutamine, penicillin–streptomycin (50 U to 50 lg/ml)
and incubated at 37 ꢁC in humidified atmosphere of
5% CO2, 95% air. The culture medium was changed
twice a week.
´
´
7. Gonzalez-Dıaz, H.; Cruz-Monteagudo, M.; Molina, R.;
Tenorio, E.; Uriarte, E. Bioorg. Med. Chem. 2005, 13,
1119.
4.2. Treatment with antitumour agents and MTT color-
imetric assay
8. Estrada, E. Mutat Res. 1998, 420, 67.
´
´
´
9. Gonzalez-Dıaz, H.; Olazabal, E.; Santana, L.; Uriarte, E.;
Gonzalez-Dıaz, Y.; Castan˜edo, N. Bioorg. Med. Chem.
Human cancer cell line (5 · 103 cells/0.33 cm2) was pla-
ted in 96-well plates ‘Nunclon TM MicrowellTM’ (Nunc)
and was incubated at 37 ꢁC. After 24 h, cells were trea-
ted with the compounds (final concentration 0.01–
100 lM). Untreated cells were used as controls. Micro-
plates were incubated at 37 ꢁC in humidified atmosphere
of 5% CO2, 95% air for 3 days and then cytotoxicity was
measured with colorimetric assay based on the use of
tetrazolium salt MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-
diphenyl tetrazolium bromide).24 The results were read
on a multiwell scanning spectrophotometer (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 density at time zero; C is
the control optical density).
´
´
2007, 15, 962.
´
´
´
10. Gonzalez-Dıaz, H.; Torres-Gomez, L. A.; Guevara, Y.;
Almeida, M. S.; Molina, R.; Castan˜edo, N.; Santana, L.;
Uriarte, E. J. Mol. Model. 2005, 11, 116.
´
´
11. Gonzalez-Dıaz, H.; Tenorio, E.; Castan˜edo, N.; Santana,
L.; Uriarte, E. Bioorg. Med. Chem. 2005, 13, 1523.
´
´
12. Gonzalez Borroto, J. I.; Perez Machado, G.; Creus, A.;
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Yoo, K. H.; Kim, D. J.; Pae, A. N. Bioorg. & Med. Chem.
2005, 13, 3339.
15. Fichera, M.; Fortuna, C. G.; Impallomeni, G.; Mus-
umarra, G. Eur. J. Org. Chem. 2002, 145.
16. Carosati, E.; Sciabola, S.; Cruciani, G. J. Med. Chem.
2004, 47, 5114.
17. Cruciani, G.; Crivori, P.; Carrupt, P. A.; Testa, B. J. Mol.
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Acknowledgment
We thank the University of Catania for financial
support.
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References and notes
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