Novel imidazole-based combretastatin A-4 analogues: Evaluation of their in vitro antitumor activity and molecular modeling study of their binding to the colchicine site of tubulin

Article abstract of DOI:10.1016/j.bmcl.2006.08.087

The in vitro antitumor activity of novel combretastatin-like 1,5- and 1,2-diaryl-1H-imidazoles was evaluated against the NCI 60 human tumor cell lines panel. Compounds 2d and 2g proved to be more cytotoxic than CA-4 in tests involving their evaluation ove

Full text of DOI:10.1016/j.bmcl.2006.08.087

Bioorganic & Medicinal Chemistry Letters 16 (2006) 5757–5762
Novel imidazole-based combretastatin A-4 analogues: Evaluation of
their in vitro antitumor activity and molecular modeling study
of their binding to the colchicine site of tubulin
a
b,
a
Fabio Bellina,a,* Silvia Cauteruccio, Susanna Monti and Renzo Rossi
*
a
`
Dipartimento di Chimica e Chimica Industriale, Universita di Pisa, Via Risorgimento 35, I-56126 Pisa, Italy
bIstituto per i Processi Chimico-Fisici (IPCF-CNR), Area della Ricerca, Via G. Moruzzi 1, I-56124 Pisa, Italy
Received 12 July 2006; revised 18 August 2006; accepted 19 August 2006
Available online 6 September 2006
Abstract—The in vitro antitumor activity of novel combretastatin-like 1,5- and 1,2-diaryl-1H-imidazoles was evaluated against the
NCI 60 human tumor cell lines panel. Compounds 2d and 2g proved to be more cytotoxic than CA-4 in tests involving their eval-
uation over a 10À4–10À8 range. Docking experiments showed a good correlation between the MG_MID Log GI50 values of all these
compounds and their calculated interaction energies with the colchicine binding site of ab-tubulin.
Ó 2006 Elsevier Ltd. All rights reserved.
Combretastatin A-4 (CA-4) (1), a potent antimitotic
agent isolated from the stem wood of the South African
tree Combretum caffrum,1 exhibits strong antitubulin
activity by binding to tubulin at the colchicine binding
site.2 It shows potent cytotoxicity against a variety of
human cancer cell lines, including those that are multi-
drug resistant3 and, most importantly, has demonstrated
powerful antiangiogenesis properties.4
although its low aqueous solubility and Z-configura-
tion pose significant liabilities. In fact, the Z-config-
ured C–C double bond of 1 is prone to isomerize to
the E-form during storage and administration thus
producing dramatic reduction in both antitubulin
activity and cytotoxicity.7,8 Therefore, considerable ef-
forts have gone into modifying 1 and discovering its
bioavailable Z-restricted analogues based on the bio-
isosteric replacement of olefinic double bond of the
natural product with vicinal diaryl-substituted five-
membered heteroaromatic rings including oxazole,
isoxazole, thiazole, pyrazole, tetrazole, and imidazole.9
As far as the imidazole derivatives are concerned, it
should be noted that in 2002 Wang and co-workers10
found that among a small series of 5-aryl-1-(3,4,5-tri-
methoxyphenyl)-1H-imidazoles, compounds 2a and 2b
had comparable antiproliferative properties against the
NCI-H460 non-small cell lung cancer line (63 and
61 nM, respectively) and the HCT-15 colon cancer cell
line (69 and 89 nM, respectively) and very similar anti-
tubulin activity (7.7 and 7.6 lM, respectively).
HO
B
MeO
A
MeO
OMe
OMe
1 (CA-4)
Using the X-ray structure of the tubulin–colchicinoid
complex,5 docking studies have recently allowed the
identification of reasonable binding modes of a set of
different colchicine site inhibitors that included 1, podo-
phyllotoxin, methylchalcone, curacin A, indanocine,
and nocodazole.6
Because of its potent cytotoxicity and structural sim-
plicity, 1 has been envisaged as a very attractive lead
N
N
Ar2
Ar1
N
Ar2
N
H
Ar1
2
3
Keywords: Cytotoxicity; Combretastatin A-4; Colchicine; Imidazoles;
Tubulin; Molecular modeling; Antitumor activity.
2a : R1 = OMe; R2 = NH2
2b : R1 = OMe; R2 = OH
2c : R1 = NH2; R2 = H
3a : Ar1 = 3-NH2,4-MeOC6H3
Ar2 = 3,4,5-(MeO)3C6H2
3b : Ar1 = 3-OH,4-MeOC6H3
Ar2 = 3,4,5-(MeO)3C6H2
*
Corresponding authors. Tel.: +39 0 50 2219282; fax +39 0 50 2219260
(F.B.); tel.: +39 0 50 3152520; fax +39 0 50 3152442 (S.M.); e-mail
addresses: bellina@dcci.unipi.it; s.monti@ipcf.cnr.it
3c : Ar1 = Ar2 = 1-Me,5-indolyl
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.08.087

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F. Bellina et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5757–5762
On the other hand, compound 2c was inactive against
NCI-H460 and HCT-15, and was not assayed for its
antitubulin activity. It was also demonstrated that 4,5-
diaryl-1H-imidazoles 3a–c had cytotoxic and antitubulin
activities significantly higher than those of compounds 2
and that incorporation of an N-methyl group into the
imidazole ring of compounds 3 can lead to substances
4 with improved pharmacokinetic profiles, characterized
by excellent bioavailability.10
N
N
N
N
a
Ar2–I
+
(40 – 72%) Ar2
Ar1
Ar1
6a–b
2d–g
7a–c
Ar1
Ar2
2
d
e
f
3,4,5-(MeO)3C6H2
3,4,5-(MeO)3C6H2
4-MeOC6H4
2-naphthyl
4-MeOC6H4
4-MeOC6H4
g
3,4,5-(MeO)3C6H2
3-F,4-MeOC6H3
Scheme 1. Reagents and condition: (a) Compounds 7 (2 equiv),
Pd(OAc)2 (5 mol%), AsPh3 (10 mol%), CsF (2 equiv), DMF, 140 °C,
21–90 h.
Recently, in continuation of our investigations into the
synthesis and evaluation of the cytotoxic activity of vic-
inal diaryl-substituted five-membered heterocycles,
which can be considered as Z-restricted analogues of
1,11 we developed selective and efficient procedures for
the synthesis of 1,5- and 1,2-diaryl-1H-imidazoles of
general formula 212 and 5,13,14 respectively.
N
N
N
N
Ar2
a
Ar2–I
+
(60 – 80%)
Ar1
Ar1
6a–c
7a,b,d
5a–e
Ar1
Ar2
2-naphthyl
4-MeOC6H4
5
Ar2
N
N
N
N
a
b
c
d
e
3,4,5-(MeO)3C6H2
3,4,5-(MeO)3C6H2
4-MeOC6H4
Ar2
Ar1
Ar1
Me
4-MeOC6H4
4-MeOC6H4
2-naphthyl
3,4,5-(MeO)3C6H2
3,4,5-(MeO)3C6H2
4
5
Scheme 2. Reagents and condition: (a) Compounds 7 (2 equiv),
Pd(OAc)2 (5 mol%), CuI (2 equiv), DMF, 140 °C, 48–60 h.
In this paper we report the evaluation of the cytotoxicity
of a variety of these heterocycles. Moreover, we describe
the results of molecular modeling studies performed to
investigate the interactions of these CA-4 analogues at
the colchicine binding site of ab-tubulin and to verify
the possible relationship between their calculated inter-
action energies and their cytotoxicity values. In fact,
some papers reporting a good correlation between cyto-
toxicity and experimental antitubulin activity data have
been published.10,15 However, it has also been found
that significant variations of the cytotoxicity values
due to structural modifications do not affect sometimes
the antitubulin activity16 and that some potent antitub-
ulin compounds can be marginally cytotoxic.17 It should
also be noted that the calculated interaction energies of
compounds 2 and 5 with ab-tubulin could be very useful
to identify imidazole derivatives able to cause selective
damage to tumor vasculature.18
the mean Log molar drug concentration (MG_MID
Log) for all tested human cancer cell lines, are shown
in Table 1.
Some aspects of the cytotoxicity data reported in this ta-
ble merit comment. First, compound 2g proved to be the
most potent imidazole derivative among the examined
compounds which, save 2f and 5c, contained a 3,4,5-tri-
methoxyphenyl (TMP) group. Second, compounds 2d
and 2g proved to be more cytotoxic than 1, which con-
firms that, as previously reported, the 3-fluoro-4-meth-
oxyphenyl and the 2-naphthyl moieties are good
surrogates of the CA-4 B ring.19 Third, the 1,5-diaryl-
Table 1. Cytotoxicity of imidazoles 2d–g and 5a–e, and of CA-4 1
against the NCI 60 human cancer cell lines screening panel
1,5-Diaryl-1H-imidazoles 2d–g were regioselectively syn-
thesized by coupling of 1-aryl-1H-imidazoles 6a,b with
the required aryl iodides 7 in DMF at 140 °C in the pres-
ence of CsF as the base and a catalyst precursor consist-
ing of a mixture of Pd(OAc)2 and AsPh3 (Scheme 1).12
Compound
Cytotoxicitya GI50
MG_MID Loga
(lM)
HCT-15
NCI-H460
GI50
TGI
2g (NSC736359)
2d (NSC736992)
2e (NSC733436)
2f (NSC734603)
5e (NSC736994)
5a (NSC736993)
5d (NSC735354)
5b (NSC735355)
5c (NSC734602)
1 (NSC613729)b
<0.01
0.017
0.170
19.498
0.022
0.051
1.148
1.148
22.909
<0.01
<0.01
0.018
0.363
15.488
0.021
0.037
2.344
1.585
17.783
<0.01
À7.40
À7.14
À6.59
À4.80
À6.86
À6.82
À5.45
À5.34
À4.76
À7.00
À4.51
À4.40
À4.71
À4.32
À4.80
À4.68
À4.32
À4.16
À4.19
À5.03
On the other hand, 1,2-diaryl-1H-imidazoles 5a–e were
regioselectively prepared by C-2 arylation of 6a–c with
the required aryl iodides 7 in DMF at 140 °C in the
presence of 5 mol% Pd(OAc)2 and 2 equiv of CuI
(Scheme 2).14
Compounds 2d–g and 5a–e were then tested for their
cytotoxic activity against the US NCI 60 tumor cell lines
screening panel. The results obtained using these com-
pounds and 1 against two selected human cancer cell
lines,10 HCT-15 (MDR+) and NCI-H460 (MDR–),
and the entire NCI cell panel, which are reported as
a Evaluated over a 10À4 to 10À8 M range. These values are means of
two series of cytotoxicity tests (reported in the Supporting
Information).
b http://dtp.nci.nih.gov/dtpstandard/dwindex/index.jsp.

F. Bellina et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5757–5762
5759
1H-imidazoles 2d and 2e showed cytotoxic activities
higher than those of the corresponding 1,2-diaryl-1H-
imidazoles 5a and 5b, respectively. Fourth, compounds
2d and 5a possessing a 2-naphthyl substituent were
found to be more cytotoxic than compounds 2e and
5b, respectively, in which the 4-methoxyphenyl moiety
replaces the 2-naphthyl group. It is also worth mention-
ing that some of the examined imidazoles proved to be
especially active against some tumor cell lines (see Sup-
porting Information). For example, 2d displayed Log
TGI À7.53 for the colon cancer cell line HCC-2998
and the value of Log TGI of 2g for the leukemia cell line
HL-60(TB) was less than À7.54. Thus, 2d and 2g were
approximately 1000 times more efficacious in these can-
cer lines compared to the mean total growth inhibition
(TGI) value in the other 59 cell lines. Finally, it should
be mentioned that an in vivo test performed on MD-
MBA-435 breast cancer cells xenotransplanted in immu-
nodeficient mice showed that 2e is able to inhibit signif-
icantly the tumor growth at a 150 mg/kg/day dose (52%
inhibition after 25 days).20
Figure 1. Arrangement of
colchicine binding site.
1
(green), 2d–g and 5a–e inside the
had a three-dimensional arrangement of their groups
similar to that assumed by 1 in its binding conformation
(Fig. 1). It should also be noted that the B ring of com-
pounds 2d, 2e, 2g, and 5a, 5b, 5d, and 5e (where, as for 1,
the TMP group is represented by the A ring) was found
to occupy a similar space in the colchicine binding site
and was buried in all cases in the ab-tubulin structure
near the Cys-b239 residue.
Docking studies on compounds 2d–g and 5a–e were then
carried out to investigate the structural basis of the bio-
logical results obtained with these cis-locked CA-4 ana-
logues. Thus, taking into account that the mechanism of
cytotoxicity of CA-4 (1) and other combretastatins has
been shown to involve the inhibition of tubulin polymer-
ization by binding at the colchicine binding site of tubu-
lin,9a we directed our attention to the in silico
determination of the total interaction energies (Eint) of
compounds 2d–g and 5a–e with the colchicine binding
site on ab-tubulin.
On the other hand, since docking putative inhibitors in a
rigid protein binding site derived from a complex with
another ligand may be misleading in the identification
of the correct binding mode and in the assessment of
reliable binding affinities, further calculations were per-
formed to refine the chosen adducts and find the best
structural adaptation of both the protein active site res-
idues and the ligand groups in forming the complex.
Thus, the tubulin–imidazole ligand complexes were sub-
jected to energy minimization to relieve any unfavorable
clash in the model structures and the minimized systems
were used as input to short molecular dynamics simula-
tion runs (100 ps), performed in the NVE ensemble (see
Supporting Information).22 To evaluate the readjust-
ment of the active site residues in the calculated tubu-
lin–imidazole structures, the root mean square
difference (rsmsd) between the atoms of the tubulin-1
complex and the corresponding atoms of each tubulin–
imidazole model was evaluated. In the case of backbone
At first, the minimum energy conformations of the imid-
azole ligands were selected as reported in the Supporting
Information. Then, in order to find the best arrange-
ment of each molecule inside the colchicine binding site,
the atomic coordinates of ab-tubulin complexed with
CA-4 were used as the starting point of the docking
studies.6 Molecular docking was carried out using
DOCK5 software package21 and the adopted docking
methodology was validated by checking the energy
scores obtained by docking into tubulin a representative
set of ligands, namely colchicine (8), CA-4 (1), stegana-
cin (9), and compounds 10 and 11, which are included
among those recently used to construct a comprehen-
sive, structure-based pharmacophore.6
˚
atoms’ superimposition the rmsd was lower than 0.5 A,
whereas for the side chains the maximum rmsd found
˚
was ca. 0.8 A (for compound 5a) suggesting that the li-
gand-induced conformational changes were small and,
as expected, more pronounced for the side chains as
compared to the main chains.
The molecular electrostatic potential (MEP) of imidaz-
oles 2 and 5 was then examined in comparison with that
of 1 (Fig. 2). Interestingly, remarkable correlations sug-
gesting the existence of related binding mechanisms for
the tubulin polymerization inhibition were found. In
fact, besides having similar three-dimensional electron
density shapes in their most stable binding conforma-
tions, the imidazole derivatives exhibited similar regions
of positive (blue) and negative (red) electrostatic poten-
tial. It can also be noticed that in the case of 2g the fluo-
rine lone pairs produced a noticeable negative potential
above and below the plane of the B ring, thus enlarging
the negative potential lobe generated by the methoxy
oxygen lone pairs. This lobe was instead smaller in 2e
OMe
MeO
MeO OMe
MeO
MeO
OMe
MeO
S
O Me
O
N
H
O
N
H
10
11
The docking methodology was then used for docking
the minimum energy conformations of the investigated
imidazole ligands into ab-tubulin. Interestingly, the
top score conformers of the whole set of molecules

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F. Bellina et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5757–5762
Figure 2. MEP of compounds 1, 2d–g, and 5a–e produced by RESP partial charges (red and blue surfaces = À10 kcal/mol and +10 kcal/mol,
respectively).
where the fluorine atom is replaced by a hydrogen atom.
A similar, negative region was observed for the lone
pairs of the OH group onto the B ring of CA-4, which,
due to the presence of the hydroxyl proton hydrogen
bonded to the nearby methoxy oxygen, gave rise to a
somewhat compressed negative lobe. The N-3 lone pair
of the imidazole ring also generated a large negative
lobe, which in the case of 5d and 5e was enlarged by
the cooperative effect of the A ring p density and its
meta methoxy oxygen lone pairs. On the other hand,
in the case of 5b and 5a the enlargement was due to
the cooperation of the p density of the B ring. Analo-
gous negative regions were observed for 2g, 2d, 2e,
and 2f where the presence of the H-4 imidazole hydro-
gen atom prevents the enlargement of the lobe and gives
rise to a very small positive region.
Table 2. Total interaction energy (Eint), in kcal/mol, its van der Waals
(vdW) and electrostatic (elec.) components and the intramolecular
energy difference between the more stable conformers inside and
outside the binding site (DEbind) of each studied molecule
Compound
vdW
Elec.
Eint
DEbind
CA-4
2g
5a
2d
5e
À50.529
À53.395
À54.722
À52.613
À46.860
À46.243
À41.195
À53.363
À42.111
À43.501
À12.599
À11.181
À6.112
À7.729
À12.494
À10.956
À12.331
À2.232
À4.410
À2.405
À63.128
À64.576
À60.834
À60.342
À59.353
À57.199
À53.526
À55.595
À46.521
À45.906
0.77
1.21
0.88
2.89
1.30
2.99
1.22
1.44
1.13
2e
5d
5b
2f
5c
A negative potential was always found to be associated
with the methoxy oxygen atoms of the A ring and a
strong shrinking of the negative lobe was apparent for
2f where only one methoxy group is present. On the con-
trary, methoxy protons produced positive lobes whose
size depends on spatial arrangement and cooperative
effects.
implying that van der Waals contacts promote binding.
Compounds 2e, 2f, 5c, 5d, and 5e had a lower number of
hydrophobic contacts in comparison to the other inves-
tigated imidazoles and thus the van der Waals interac-
tion energy was less favorable.
It was also found that the electrostatic term plays a key
role in predicting the binding and that, due to additional
favorable electrostatic interactions with the active site
residues, greater stabilization energy was achieved for
In order to quantify these qualitative predictions, the
intermolecular interaction energy, which is the sum of
the van der Waals and electrostatic interactions between
ligand and protein, was then calculated. As shown in
Table 2, the van der Waals packing between the receptor
and the ligand proved to be favorable in all the models
2g which had the lowest total energy interaction (Etot
)
and the lowest energy difference (DEbind) between the
most stable conformers inside and outside the binding
site (Table 2).

F. Bellina et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5757–5762
5761
All the imidazole ligands were found in the same loca-
tion and only a very small difference in their binding
mode, due to the orientation of the imidazole moiety,
was observed for 5a and 5b. In fact, as shown in
Figure. 4 and confirmed by the energy values reported
in Table 3 of the Supporting Information, 5a and 5b
had electrostatic potential matches of their imidazole
ring with the nearby surface site residues less favorable
than the other imidazoles. The electrostatic complemen-
tarity was not well satisfied and, to ameliorate the inter-
action with the site and minimize repulsion forces, these
molecules were obliged to readjust the position of their
imidazole rings, changing their orientation in compari-
son with that of the other derivatives.
Figure 3. Compound 2g inside the colchicine binding site. Hydrogen
bonds are evidenced as dashed black lines. Secondary structure
elements are represented as solid red cylinders (a-helices), solid cyan
arrows (b-sheets), and solid gray ribbon (loops). Amino acids within
˚
4.5 A from 2g are displayed as sticks.
Finally, it must be mentioned that even though the com-
plementarity between the investigated imidazoles and
the colchicine binding site of tubulin has proved remark-
able, the existence of a set of relatively small empty
pockets between the ligands and the target residues of
the protein might allow an improvement of the interac-
tions between imidazole derivatives and these regions of
the binding site. The topography and volume of these
cavities in the colchicine binding site, which could be
occupied by substituent groups of the imidazole moiety
possessing an appropriate size and polarity, can be ob-
served, for example, in the cross section of the tubu-
lin–2g binding site surfaces (Fig. 5).
It is also worth noting that the binding mode of the
investigated imidazoles strongly resembled that of CA-
4 1 and that their docking revealed a consistent set of
recurring interactions. As shown in Figure. 3, in which
compound 2g is represented inside the colchicine bind-
ing site of tubulin, the 4-methoxy oxygen of the TMP
group of compounds 2d, 2e, 2g and 5a, 5b, 5d, 5e engag-
es a hydrogen bond to the Cys-b239 SH group and ap-
pears to be oriented so that it can interact favorably with
the hydrophobic Val-b236, Leu-b240, Leu-b246, Ala-
b248, Leu-b253, and Ala-b352 side chains, which are
positioned over and below its plane (see also Tables 4
and 5 in the Supporting Information). At the bottom
of the cleft which hosts the A ring the Met-b257 side
chain, owing to its sulfur lone pair, creates a negative
electrostatic potential region which favorably interacts
with the methyl part of the meta methoxy substituents,
as confirmed by the value of the interaction energy with
the colchicine binding site. On the other hand, the B aro-
matic moieties of compounds 2d, 2e, and 5a, 5b, 5d, and
5e were found to be located in a large pocket where their
p density favorably interacts with the Asn-b256 methy-
lene and Ala-b314 methyl groups (Fig. 4, and Tables 4
and 5).
In conclusion, in this study the in vitro antitumor activ-
ity of novel vicinal diaryl-substituted 1H-imidazole ana-
logues of CA-4 has been evaluated and two 1,5-diaryl
derivatives have been found to be more cytotoxic than
CA-4. Moreover, a combined computational strategy
consisting of molecular mechanics, rigid docking, and
molecular dynamics simulations has been used to inves-
tigate the interactions of 1,5- and 1,2-diaryl-1H-imidaz-
oles 2 and 5 with the colchicine binding site of ab-
tubulin that is the recognized main biological target
for combretastatins.9 The developed procedure allowed
the identification of the bioactive structures of the li-
gands among their minimum energy conformations,
their arrangement inside the binding site, and the theo-
retical calculation of their total interaction energy. It
should be noted that some of the herewith investigated
imidazoles have been shown to be characterized by high
total interaction energies with the colchicine binding site
and that for one of these heterocycles, 2g, this interac-
In the case of 2g (Figs. 3 and 4), the methoxy oxygen of
the B ring proved to be oriented toward the methyl
group of Val-a179, and the fluorine atom was found
to form a hydrogen bond with the backbone NH group
of Val-a179.
Figure 4. Compounds 1, 2d–g, and 5a–e inside their binding site.
Molecular surfaces colored according to the electrostatic potential
(blue = positive potential region, red = negative potential region).
Figure 5. Cross-section of the tubulin-2g (a and b) binding site
surfaces. Several minor cavities are visible near both the a and b
moieties.

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F. Bellina et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5757–5762
9. For reviews on the chemistry and biology of combretast-
atin A-4 analogues, see: (a) Nam, N.-H. Curr. Med. Chem.
2003, 20, 558; (b) Cirla, A.; Mann, J. Nat. Prod. Rep. 2003,
20, 558; (c) Tron, G. C.; Pirali, T.; Sorba, G.; Pagliai, F.;
Busacca, S.; Genazzani, A. A. J. Med. Chem. 2006, 49,
3033.
10. Wang, L.; Woods, K. W.; Li, Q.; Barr, K. J.; McCroskey,
R. W.;Hannick,S.M.;Gherke,L.;Credo,R.B.;Hui,Y.-H.;
Marsh, K.; Warner, R.; Lee, J. Y.; Zielnski-Mozng, N.;
Frost, D.; Rosenberg, S. H.; Sham, H. L. J. Med. Chem.
2002, 45, 1697.
11. (a) Bellina, F.; Anselmi, C.; Martina, F.; Rossi, R. Eur. J.
Org. Chem. 2003, 2290; (b) Bellina, F.; Falchi, E.; Rossi,
R. Atti del Convegno, P8, XVII National Meeting of the
Divisione di Chimica Farmaceutica of the Italian Chem-
ical Society, Pisa, Italy, September 6–10, 2004; (c) Bellina,
F.; Rossi, R. Abstracts, P9, 26th Winter Meeting of the
EORTC–PAMM Group, Arcachon, France, January 26–
29, 2005.
tion energy has been found to be higher than that of
CA-4. The good linear correlation (R2 = 0.95) between
calculated interaction energies of imidazoles 2 and 5
with the colchicine binding site of ab-tubulin and their
MG_MID Log GI50 values, and the identification of
ancillary binding site pockets, represent encouraging re-
sults, which suggest that the chosen theoretical method-
ology could be appropriate for the identification of
compounds with increased affinity and specificity for
the colchicine binding site. Finally, it is also worth not-
ing that, as CA-4 and some other antitubulin agents
have recently been shown to exhibit antivascular dis-
rupting activity (VDA) against tumor neovasculature
at dose levels lower than those of their maximum toler-
ated dose,18 the calculated total interaction energies
could be used to select potential lead derivatives able
to cause selective VDA.
12. Bellina, F.; Cauteruccio, S.; Mannina, L.; Rossi, R.; Viel,
S. J. Org. Chem. 2005, 70, 3997.
13. Bellina, F.; Cauteruccio, S.; Mannina, L.; Rossi, R.; Viel,
S. Eur. J. Org. Chem. 2006, 693.
Acknowledgments
14. Bellina, F.; Cauteruccio, S.; Rossi, R. Eur. J. Org. Chem.
2006, 1379.
This work was supported by the University of Pisa. We
are also grateful to Dr. I. Fichtner of Max Delbruck
¨
Center for Molecular Medicin in Berlin-Buch for an
in vivo test of tumor growth inhibition.
15. (a) Cushman, M.; He, H.-M.; Lin, C. M.; Hamel, E. J.
Med. Chem. 1993, 36, 2817; (b) Ohsumi, K.; Hatanaka, T.;
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Supplementary data
Complete in vitro cytotoxicity results against the NCI 60
tumor cell lines panel. Details concerning the molecular
modeling procedure: conformational search (Table 3),
docking and complex refinement (Fig. 6), and evalua-
tion of the interaction energy and binding mode (Tables
4 and 5). Linear correlation graph between the calculat-
ed total interaction energy values (Etot) and the experi-
mental MG_MID Log GI50 (Fig. 7). Supplementary
data associated with this article can be found, in the on-
line version, at doi:10.1016/j.bmcl.2006.08.087.
16. (a) Pettit, G. R.; Rhodes, M. R.; Herald, D. L.; Hamel, E.;
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(b) Ohsumi, K.; Nakagawa, R.; Fukuda, Y.; Hatanaka,
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