Substituted phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonamides as antimitotics. Antiproliferative, antiangiogenic and antitumoral activity, and quantitative structure-activity relationships

Article abstract of DOI:10.1016/j.ejmech.2011.08.034

The importance of the bridge linking the two phenyl moieties of substituted phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonates (PIB-SOs) was assessed using a sulfonamide group, which is a bioisostere of sulfonate and ethenyl groups. Forty one phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonamide (PIB-SA) derivatives were prepared and biologically evaluated. PIB-SAs exhibit antiproliferative activities at the nanomolar level against sixteen cancer cell lines, block the cell cycle progression in G₂/M phase, leading to cytoskeleton disruption and anoikis. These results were subjected to CoMFA and CoMSIA analyses to establish quantitative structure-activity relationships. These results evidence that the sulfonate and sulfonamide moieties are reciprocal bioisosteres and that phenylimidazolidin-2-one could mimic the trimethoxyphenyl moiety found in the structure of numerous potent antimicrotubule agents. Finally, compounds 16 and 17 exhibited potent antitumor and antiangiogenic activities on HT-1080 fibrosarcoma cells grafted onto chick chorioallantoic membrane similar to CA-4 without significant toxicity for the chick embryos, making this class of compounds a promising class of anticancer agents.

Full text of DOI:10.1016/j.ejmech.2011.08.034

European Journal of Medicinal Chemistry 46 (2011) 5327e5342
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Substituted phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonamides as
antimitotics. Antiproliferative, antiangiogenic and antitumoral activity,
and quantitative structure-activity relationships
,
b
,
c
d
a
a
Sébastien Fortina , Lianhu Wei , Emmanuel Moreau , Jacques Lacroix , Marie-France Côté ,
*
Éric Petitclerca , Lakshmi P. Kotrac e, René C. Gaudreaulta
,
1
,
,f,
*
a Unité des Biotechnologies et de Bioingénierie, Centre de recherche, C.H.U.Q., Hôpital Saint-François d’Assise, Québec, QC, Canada G1L 3L5
b Faculté de Pharmacie, Université Laval, Pavillon Vandry, Québec, QC, Canada G1V 0A6
c Center for Molecular Design and Preformulations, Toronto General Research Institute, University Health Network, Toronto, ON, Canada M5G 1L7
d Clermont 1, Université d’Auvergne, Inserm, U 990, F-63000 Clermont-Ferrand, France
e Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, ON, Canada M5S 3M2
f Faculté de Médecine, Université Laval, Pavillon Vandry, Québec, QC, Canada G1V 0A6
a r t i c l e i n f o
a b s t r a c t
Article history:
The importance of the bridge linking the two phenyl moieties of substituted phenyl 4-(2-
oxoimidazolidin-1-yl)benzenesulfonates (PIB-SOs) was assessed using a sulfonamide group, which is
a bioisostere of sulfonate and ethenyl groups. Forty one phenyl 4-(2-oxoimidazolidin-1-yl)benzene-
sulfonamide (PIB-SA) derivatives were prepared and biologically evaluated. PIB-SAs exhibit anti-
proliferative activities at the nanomolar level against sixteen cancer cell lines, block the cell cycle
progression in G2/M phase, leading to cytoskeleton disruption and anoikis. These results were subjected
to CoMFA and CoMSIA analyses to establish quantitative structure-activity relationships. These results
evidence that the sulfonate and sulfonamide moieties are reciprocal bioisosteres and that phenyl-
imidazolidin-2-one could mimic the trimethoxyphenyl moiety found in the structure of numerous
potent antimicrotubule agents. Finally, compounds 16 and 17 exhibited potent antitumor and anti-
angiogenic activities on HT-1080 fibrosarcoma cells grafted onto chick chorioallantoic membrane similar
to CA-4 without significant toxicity for the chick embryos, making this class of compounds a promising
class of anticancer agents.
Received 27 April 2011
Received in revised form
19 August 2011
Accepted 23 August 2011
Available online 30 August 2011
Keywords:
Phenyl 4-(2-oxoimidazolidin-1-yl)
benzenesulfonamides
PIB-SA
Antimicrotubule agents
Anticancer drugs
Antimitotic agents
Ó 2011 Elsevier Masson SAS. All rights reserved.
1. Introduction
cause of death worldwide by 2010 [2]. In addition, number of global
cancer deaths is projected to increase by 45% from 2007 to 2030,
Cancer is the second cause of death in North America after
cardiovascular diseases. About 569,490 Americans were expected
to die of cancer in 2010 [1]. Moreover, according to a new edition of
the World Cancer Report 2008, cancer is going to be the leading
influenced in part by an increasing and aging population [3].
Despite recent breakthroughs, the ability of curing cancer patients
remains an elusive goal, with a great need to develop alternative
and more effective therapies to improve both life expectancy and
quality of life of patients [4].
Microtubules, the key component of the cytoskeleton are
composed of a-, b-tubulin heterodimers. Microtubules are also one
Abbreviations: PIB-SO, phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonate;
PIB-SA, phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonamide; C-BS, colchicine-
binding site; CAM assays, chick chorioallantoic membrane tumor assays; EBI-N,N0,
ethylene-bis(iodoacetamide).
of the most successful cancer chemotherapeutic targets since they
are responsible for mitotic spindle formation and proper chromo-
somal separation [5]. Antimicrotubule agents such as vinca and
taxus alkaloids play important roles in the treatment of wide
variety of cancers [6,7]. However, poor water solubility, unfavorable
biodistribution, significant dose-limiting toxicity, and multidrug
resistance mechanisms limit their use and their efficacy [5,8,9]. In
that context, several academic and industrial groups focus their
efforts to develop new antimicrotubule agents to circumvent these
* Corresponding authors. C.H.U.Q., Hopital Saint-Francois dAssiseCentre de
recherche, Unite des Biotechnologies et de Bioingenierie, Quebec, QC, Canada G1L
3L5. Tel.: þ1 418 525 4444x52364; fax: þ1 418 525 4372.
E-mail addresses: sebastien.fortin.1@ulaval.ca, sebastien.fortin.81@gmail.com,
william.wei@utoronto.ca, emmanuel.moreau@u-clermont1.fr, jacques.lacroix@
crsfa.ulaval.ca, marie-france.cote@crsfa.ulaval.ca, eric.petitclerc@hema-quebec.qc.
ca, lkotra@uhnresearch.ca, rene.c-gaudreault@crsfa.ulaval.ca.
1
Present address: Héma-Québec, 1070, avenue des Sciences-de-la-Vie, Québec,
QC, Canada G1V 5C3.
0223-5234/$ e see front matter Ó 2011 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2011.08.034

5328
S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
limitations. Several antimitotics exhibiting vascular disrupting
properties are under evaluation in phase I to phase III clinical trials
[10,11]. Drugs such as combretastatin A-4 sodium phosphate (1, CA-
4P), a water soluble prodrug of combretastatin A-4 (2, CA-4) iso-
lated from Combretum caffrum, have received much attention in
recent years due to its simple stilbene structure, its high potency as
colchicine-binding site (C-BS) antagonist and its potent antitumoral
activity [12,13]. CA-4P represents a promising drug in various
clinical settings showing potent activity by reducing significantly
the tumoral blood flow [14]. However, CA-4P shows many delete-
rious effects such as hypertension, hypotension, tachycardia, tumor
pain and lymphopenia [10], and is hampered by a short biological
half-life [15,16] and biological instability [17,18]. Consequently, the
search for novel antimicrotubule agents is still required to improve
the biopharmaceutical and pharmacological properties of that class
of compounds. To that end, we recently reported a new class of
antimitotic agents named phenyl 4-(2-oxoimidazolidin-1-yl)ben-
zenesulfonate (3, PIB-SO) [19] where the trimethoxyphenyl moiety
of CA-4-sulfonate (4) is replaced by a phenylimidazolidin-2-one
moiety. PIB-SOs are potent antiproliferative agents exhibiting IC50
in the low nanomolar range on several tumor and chemoresistant
cell lines. PIB-SOs were also found to bind to the C-BS, arresting the
cell cycle in G2/M phase, leading to the disruption of the cyto-
skeleton and apoptosis. Of interest, the structure-activity relation-
ship studies of PIB-SOs suggest that the phenylimidazolidin-2-one
moiety (ring A of PIB-SOs) may mimic the trimethoxyphenyl
moiety CA-4 (ring A of CA-4), which is commonly found in the
design of potent antimicrotubule agents and described as a key
structural element for the binding of antimitotics to the C-BS [20].
PIB-SOs bearing a 3-chloro, a 3,5-dimethoxy or a 3,4,5-trimethoxy
group have also shown potent antitumoral and antiangiogenic
potency in the chick chorioallantoic membrane tumor assays (CAM
assay). The latter are at least as potent as CA-4 and exhibit low to
very low toxicity on the chick embryos evidencing PIB-SOs as
promising anticancer drugs.
bioisosteres [39]. In this context, we have further investigated the
importance of the sulfonate moiety bridge in the structure-activity
relationships of PIB-SO by converting the sulfonate moiety of PIB-
SO into a sulfonamide moiety leading to the formation of phenyl
4-(2-oxoimidazolidin-1-yl)benzenesulfonamides (8e48, PIB-SAs).
In this study, we assessed the effect of the nature and the position
of the substituents on the aromatic ring B of new PIB-SAs (Fig. 1).
We are describing their preparation, antiproliferative activity on
several human cancer cell lines and their antitumoral and anti-
angiogenic potency in human fibrosarcoma HT-1080 grafted onto
the chorioallantoic membrane of developing chick embryos (CAM
assay). We also prepared CoMFA and CoMSIA models to establish
some of the QSAR ruling the biological activity of PIB-SAs.
2. Chemistry
The method used for the preparation of PIB-SAs (8e48) is
described in Scheme 1. Compound 51 was prepared as published
previously [40,41]. Briefly, 1-phenylimidazolidin-2-one (50) was
obtained by the nucleophilic addition of aniline to 2-
chloroethylisocyanate in methylene chloride at 25 ꢀC followed by
cyclization of the 1-(2-chloroethyl)-3-phenylurea (49) using
sodium hydride in THF at 25 ꢀC. Compound 50 was also prepared in
good yield as previously described by Neville et al. [40] using tri-
phosgene with N-phenylethylenediamine and triethylamine in
tetrahydrofuran at 0 ꢀC. 4-(2-Oxoimidazolidin-1-yl)benzene-1-
sulfonyl chloride (51) was obtained by chlorosulfonation of
compound 50 by chlorosulfonic acid in carbon tetrachloride at 0 ꢀC.
Compounds 8e48 were then prepared by nucleophilic addition of
an appropriate aniline on compound 51 in the presence of 4-
dimethylaminopyridine in acetonitrile at 25 ꢀC.
3. Results
3.1. Antiproliferative activity
Sulfonamide group has received a great deal of attention due to
its relatively simple structure and the wide variety of drugs such as
anti inflammatory [21,22], diuretics [23], anticonvulsants [24,25],
anticarbonic anhydrase [26], cox-2 inhibitors [27] and hypogly-
cemic [28] that are based on its presence. That group is also found
in potent new antimicrotubule agents such as 2-fluoro-1-methoxy-
4-pentafluorophenylsulfonamidobenzene (5, T138067) [29e31], N-
[2-[(4-hydroxyphenyl)amino]-3-pyridinyl]-4-
methoxybenzenesulfonamide (6, ABT-751) [32e36] and N-[1-(4-
methoxybenzenesulfonyl)-2,3-dihydro-1H-indol-7-yl]-iso-
nicotinamide (7, J30) [37,38] binding to the C-BS. In addition,
sulfonamide and sulfonate groups are considered as reciprocal
The antiproliferative activity of PIB-SAs 8e48 was assessed
initially on three human cancer cell lines, namely, HT-29 colon
carcinoma, M21 skin melanoma and MCF-7 breast carcinoma cells.
HT-29, M21, and MCF-7 cell lines were selected, as they are good
representatives of tumors originating from the three embryonic
germ layers. Cell growth inhibition was assessed according to the
NCI/NIH Developmental Therapeutics Program [42]. The results are
summarized in Table 1 and expressed as the concentration of drug
inhibiting cell growth by 50% (IC50). Afterward, the biological
evaluation of the most potent PIB-SAs exhibiting an IC50 lower than
200 nM on the tumor cell panel were further assessed on a panel of
Fig. 1. Molecular structures of CA-4P (1), CA-4 (2), PIB-SOs (3), CA-4-sulfonate (4), T138067 (5), ABT-751 (6), J30 (7) and PIB-SAs (8e48).

S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
5329
Scheme 1. Reagents: (i) 2-chloroethylisocyanate, DCM; (ii) NaH, THF; (iii) triphosgene, TEA, THF; (iv) ClSO3H, CCl4 and (v) relevant aniline, DMAP/CH3CN.
13 tumor cell lines comprising chemoresistant and sensitive cells,
on binding to the C-BS and on CAM assay.
experiments to assess the binding of the most potent PIB-SAs to the
C-BS on -tubulin. We first assessed their effects on the cell cycle
b
progression. Table 4 shows the percentage of M21 cells in G0/G1, S,
and G2/M phases, respectively, after treatment with most potent
PIB-SAs and CA-4 at 5-times their respective IC50 and DMSO used as
3.2. Antiproliferative activity of most potent PIB-SAs on
chemoresistant and sensitive cancer cells
excipient. Thereafter, we used
a simple detection technique
developed by our research group to assess the binding of anti-
microtubule agents to the C-BS when in competition with N,N0-
ethylene-bis(iodoacetamide) [47] (EBI). Table 4 shows the results
obtained from the competition between EBI at 100 mM and PIB-SAs
at 1000-times their IC50 on M21 cells [47]. Finally, the effects of PIB-
The antiproliferative activities of most potent PIB-SA derivatives
14e17, 19, 20, 22, 26, 27 and 48 exhibiting an IC50 under 200 nM on
HT-29, M21 and MCF-7 tumor cell lines were assessed on ten sensi-
tive and three chemoresistant cell lines. Sensitive cells were Chinese
hamster ovary CHO, human chronic myelogenous leukemia K562,
murine lymphocytotic leukemia L1210, murine macrophages
P388D1, murine melanoma B16F0, human prostate carcinoma DU
145, human fibrosarcoma HT-1080, human breast adenocarcinoma
MDA-MB-231, human ovarian SKOV3 and T cell leukemia CEM cells,
respectively. Resistant cancer cell lines were paclitaxel-resistant
CHO-TAX 5-6 [43], colchicine- and vinblastine-resistant CHO-VV 3-
2 [44] and multidrug-resistant leukemia CEM-VLB [45,46] cells. The
results are summarized in Tables 2 and 3. Colchicine, paclitaxel and
vinblastine were used as positive control. IC50 expressed the
concentration of drug inhibiting cell growth by 50% and was assessed
according to the NCI/NIH Developmental Therapeutics Program [42].
SAs on the cytoskeleton were also assessed using a fluorescent anti-
b-tubulin antibody and immunofluorescence techniques (Table 4).
3.4. Comparative molecular similarity indices and comparative
molecular field analyses (CoMSIA and CoMFA) of PIB-SAs
CoMSIA and CoMFA were conducted to understand the mech-
anism underlying the binding of PIB-SAs to the C-BS and to design
new and more selective C-BS antagonists. We used Surflex-Sim
which is a 3D molecular similarity optimization and searching
program that uses a morphological similarity function and fast
pose generation based similarity on a molecule’s shape, hydrogen
bonding, and electrostatic properties techniques to generate
alignments of molecules [48]. The multiple alignments of the most
active compounds gave the best description of the positions of
functional groups leading to the hypothesis generation. Under-
lying assumption in the alignment is that the compound with
3.3. Binding of PIB-SAs to the colchicine-binding site
The bioisosteric PIB-SOs are C-BS antagonists disrupting the
polymerization of tubulin heterodimer, inhibiting cell division in
G2/M phase and leading to anoikis. Therefore, we have conducted

5330
S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
Table 1
overall volume of the superposition. A hypothesis was generated
from this superposition, and that hypothesis will be used as
a template to align the set of active molecules. Then the alignment
of the dataset will be used to generate QSAR models. Compounds
15, 20, 22, 27, 40, and 48 were chosen to generate hypothesis using
SYBYL Surflex-Sim mutual alignment module. The alignment
hypothesis shows that the groups of same property are well
aligned (Fig. 2A). Then all PIB-SAs were superimposed on to the
alignment hypothesis using Surflex-Sim flexible superposition
Evaluation of the Antiproliferative Activity of PIB-SAs (8e48) and CA-4 on HT-29,
M21, and MCF-7 Cell Lines.
5
6
4
O
S
O
S
O
R1
O
R
3
O
or
N
O
2
H
N
HN
N
HN
45-48
module (Fig. 2B). Observing from the q2 and r2 values, the
8-44
cv
analysis that includes the similarity data to the hypothesis
improved the r2 value in both CoMFA and CoMSIA models
cv
Compounds
R or R1
R ¼ H
IC50 (nM)a
HT-29
(models A, B, C vs. models D, E, F, models G, H, I vs. models J, K, L).
The antiproliferative activities obtained on HT-29, M21, and MCF-7
cell lines were used as dependent variables to build CoMSIA and
CoMFA models. The optimized parameters and the statistical data
in the PLS analysis of six CoMSIA models and six CoMFA models
are presented in supporting information.
M21
MCF-7
8
9
280
200
240
550
190
180
46
110
110
120
200
77
55
230
82
200
150
180
300
110
150
27
470
280
350
770
230
230
48
190
130
120
220
100
79
230
100
R ¼ 2-Me
R ¼ 2-Et
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
R ¼ 2-OMe
R ¼ 2-Me, 3-Me
R ¼ 2-Me, 4-Me
R ¼ 2-Me, 5-Me
R ¼ 3-Me
3.5. Chick chorioallantoic membrane tumor assays (CAM assays)
69
82
65
83
59
25
94
63
R ¼ 3-Et
Human HT-1080 fibrosarcoma cells were used to assess the
antitumoral activity of PIB-SAs 14e17, 19, 20, 22, 26, 27 and 48 in
the CAM assay (Fig. 3). Cumulative results are shown from two
independent experiments with 10e12 eggs per experiment. No
related toxicity was observed on chick embryos treated with the
excipient. Untreated eggs were used as negative controls and for
R ¼ 3-OMe
R ¼ 3-OPh
R ¼ 3-Br
R ¼ 3-Cl
R ¼ 3-Me, 4-Me
R ¼ 3-Me, 5-Me
R ¼ 3-tBut, 5- tBut
R ¼ 3-F, 4-F
> 1000
350
> 1000
220
340
60
> 1000
620
normalization of the results. The drugs were administered iv at 3
mg
R ¼ 3-OMe, 4-OMe
R ¼ 3-OMe, 5-OMe
520
78
21
480
660
610
610
170
18
610
810
770
per egg except for CA-4 that was used as positive control and was
administered iv at 1 mg and 3 mg per egg, respectively.
R ¼ 3-OMe, 4-OMe, 5-OMe
R ¼ 4-Me
13
260
480
450
> 1000
> 1000
870
> 1000
> 1000
580
580
> 1000
130
90
4. Discussion
R ¼ 4-Et
R ¼ 4-But
R ¼ 4-secBut
R ¼ 4-tBut
R ¼ 4-Pent
R ¼ 4-cHex
R ¼ 4-Hept
R ¼ 4-OMe
R ¼ 4-OBut
R ¼ 4-OHex
R ¼ 4-Br
> 1000
> 1000
990
> 1000
> 1000
800
630
> 1000
300
> 1000
> 1000
> 1000
> 1000
> 1000
> 1000
870
4.1. PIB-SAs inhibit the proliferation of human tumor cell lines
All compounds, except for compounds 23, 31e36, 38 and 43
exhibited antiproliferative activities in the nanomolar range. We
classified PIB-SA derivatives into three subgroups based on IC50
:
(1) high IC50 ranging from 13 to 200 nM (14e17, 19, 20, 22, 26, 27
and 48), (2) fair IC50 ranging from 83 to 810 nM (8e13, 18, 21, 24,
25, 28e30, 39e42 and 44e47), (3) weak IC50 ranging from 580
to > 1000 nM (23, 31e38 and 43) on three tumor cell lines. Among
PIB-SA subgroups having high activity, compounds 14, 20 and 27
bearing a 2,5-dimethyl, a 3-chloro and a 3,4,5-trimethoxy group
on ring B, respectively, exhibited antiproliferative activity equi-
potent to CA-4 on HT-29 cells. Moreover, compounds having
a high IC50 were further evaluated on a panel of 13 chemoresistant
and sensitive tumor cell lines, for binding to the C-BS and in the
CAM assay. The PIB-SAs 14e17, 19, 20, 22, 26, 27 and 48 showed
antiproliferative activities on sensitive cancer cells (CHO, K562,
L1210, P388D1, B16F0, DU 145, HT-1080, MDA-MB-231, SKOV3,
CEM) in the same range as in the three screening human tumor
cell lines (Table 2). However, the antiproliferative activity of the
most potent PIB-SA was lower than colchicine, paclitaxel or
vinblastine.
> 1000
240
R ¼ 4-Cl
220
370
650
170
690
600
R ¼ 4-F
230
260
> 1000
410
R ¼ 4-I
R ¼ 4-CH2CN
R ¼ 4-OCHF2
> 1000
650
> 1000
760
O
O
45
R1
¼
¼
280
190
460
480
HN
46
R1
450
190
370
HN
HN
47
48
R1
R1
¼
¼
130
87
240
170
H
N
130
61
HN
4.2. Cytotoxicity of PIB-SAs was unaffected in drug-resistant cells
CA-4
4.3
5.6
a
IC50: Expressed as the concentration of drug inhibiting cell growth by 50%.
Table 3 shows the antiproliferative activities of PIB-SAs 14e17,
19, 20, 22, 26, 27 and 48 on drug-resistant cell lines CHO-TAX 5-6,
CHO-VV 3-2 and CEM-VLB. On one hand, CHO-TAX 5-6 cells are
resistant to microtubule stabilizers (e.g., paclitaxel) and hypersen-
sitive to microtubule disruptors (e.g., colchicine, vinblastine) while
CHO-VV 3-2 cell line are resistant to microtubule disruptors and
hypersensitive to microtubule stabilizers [44]. On the other hand,
better fit to the hypothesis on structural alignment would have
better activity. At first, a mutual alignment on the most active
compounds was performed and tried to find a superposition of all
input molecules that maximizes the similarity and minimizes the

S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
5331
Table 2
Evaluation of the Antiproliferative Activity of Compounds 14e17, 19, 20, 22, 26, 27, 48, Colchicine, Paclitaxel and Vinblastine on Selected Cancer Cells.
Compd
IC50 (nM)d
CHO
K562
L1210
P388D1
B16F0
DU 145
HT-1080
MDA-MB-231
SKOV3
CEM
14
15
16
17
19
20
22
26
77
230
230
330
250
96
210
330
190
1310
220
170
17
72
200
230
230
87
81
180
200
240
99
79
250
250
250
90
86
290
270
370
270
190
260
280
58
1610
580
730
650
340
230
430
700
51
65
210
200
200
170
68
150
150
17
230
1.1
0.15
0.099
110
1270
1410
730
290
200
1220
340
45
72
200
230
240
170
70
190
170
22
74
200
200
260
110
77
170
170
23
410
7.9
0.27
0.38
69
77
78
110
110
19
340
6.8
110
110
21
260
6.9
210
200
38
520
28
27
48
770
13
28
1370
10
3600
2.9
280
2.1
2.6
Cola
Pacb
Vblc
0.71
0.36
0.40
0.16
35
1.8
1.3
0.063
9.1
0.12
0.099
0.039
a
Col, colchicine.
Pac, paclitaxel.
Vbl, vinblastine.
b
c
d
IC50: Expressed as the concentration of drug inhibiting cell growth by 50%.
CEM-VLB cells overexpress P-glycoprotein [49] which is responsible
for the cellular efflux of drugs and chemoresistance to anticancer
drugs such as doxorubicin, etoposide, paclitaxel and vinblastine
[50]. As expected and confirmed in Table 3, CHO-TAX 5-6 cells were
resistant to paclitaxel (3.1-fold) and were sensitive to compounds
14e17, 19, 20, 22, 26, 27 and 48, colchicine and vinblastine. More-
over, CHO-VV 3-2 cells were resistant to the colchicine and
vinblastine, 2.8- and 3.9-fold respectively and were sensitive to
paclitaxel. Unexpectedly and except for compound 26, CHO-VV 3-2
cells were slightly or not resistant to the PIB-SAs tested. CHO-VV 3-
2 cells were resistant to compound 26 (2.1-fold), which was even
slightly lower than for colchicine and vinblastine (2.8 and 3.9-fold,
respectively). In addition, CEM-VLB cells were resistant to colchi-
cine, vinblastine and paclitaxel, 46-, 12370- and 1579-fold respec-
tively. The antiproliferative activity all PIB-SAs, at the exception of
compound 26, 27 and 48, was unaffected by the overexpression of
P-glycoprotein. CEM-VLB cells were mildly resistant to compound
26, 27 and 48, 5.9-, >11- and 12-fold, respectively, which is more
than 4-times lower than for colchicine and several logs lower than
for paclitaxel and vinblastine.
4.3. PIB-SAs arrest the cell cycle progression in G2/M Phase
Table 4 shows percentage of M21 cells arrested in G0/G1, S, and
G2/M phases, respectively, after treatment with PIB-SAs for 24 h at
5-times their respective IC50. Control cells treated with 0.5% DMSO
were found to be in G0/G1, S, and G2/M phases at 61%, 30%, and 9%,
respectively. Compounds 14,15,17,19, 20, 22, 26, 27 and 48 strongly
blocked cell cycle progression in G2/M phase; the number of cells in
G2/M phase was increased by 42e74%. Compound 16 totally
blocked the cell cycle in G2/M phase; the number of cells in G2/M
phase was increased by 86% as observed with CA-4.
4.4. PIB-SAs inhibit EBI binding to the C-BS
As aforementioned, C-BS antagonists such as colchicine, podo-
phyllotoxin, 2-methoxyestradiol and CA-4 inhibit the EBI binding to
b
-tubulin leading to disappearance of the
formed and detectable by Western blot as a second immunor-
eacting band of -tubulin migrating faster than the native -tubulin
[47]. Except for compound 14 and 48 that mildly inhibited the
b-tubulin-EBI adduct
b
b
Table 3
Evaluation of the Antiproliferative Activity of Compounds 14e17, 19, 20, 22, 26, 27, 48, Colchicine, Paclitaxel and Vinblastine in Selected Chemoresistant Cancer Cell Lines.
Compd
IC50 (nM)d
Ratio chemoresistant/sensitivee
CHO-TAX 5-6/CHO
IC50 (nM)d
Ratio chemoresistant/sensitivee
CHO-VV 3-2/CHO
IC50 (nM)d
CEM-VLBh
Ratio chemoresistant/sensitivee
CEM-VLB/CEM
CHO-TAX 5-6f
CHO VV 3-2g
14
15
16
17
19
20
22
26
54
150
140
200
76
0.70
0.65
0.61
0.61
0.30
0.60
0.57
0.67
0.89
0.49
0.64
3.1
110
280
250
450
270
150
250
690
> 250
1860
620
140
66
1.4
1.2
1.1
1.4
1.1
1.6
1.2
2.1
>1.3
1.4
120
370
450
700
250
1.6
1.9
2.3
2.7
2.3
2.2
2.4
5.9
58
170
400
120
220
170
640
140
520
6.9
1010
> 250
4840
360
3340
600
27
48
> 11
12
46
12370
1579
Cola
Pacb
Vblc
2.8
0.82
3.9
0.41
a
Col, colchicine.
Tax, paclitaxel.
Vbl, vinblastine.
b
c
d
e
f
IC50: Expressed as the concentration of drug inhibiting cell growth by 50%.
Ratio is the IC50 of the resistant cells divided by the IC50 of their deriving not resistant cells (CHO and CEM cells).
Paclitaxel-resistant CHO-TAX 5-6 cell line.
Colchicine-and vinblastine-resistant CHO-VV 3-2 cell line.
multidrug-resistant leukemia CEM-VLB cell line.
g
h

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S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
Table 4
Effect of Selected PIB-SAs 14e17, 19, 20, 22, 26, 27, 48 and CA-4 on Cell Cycle Progression, in the Competition Assay with EBI and on the Cytoskeleton Integrity.
5
6
4
O
S
O
S
O
R1
O
R
3
O
or
N
O
2
H
N
HN
N
HN
45-48
8-44
Compd
R, R1
Cell cycle progressiona
Competition
Cytoskeleton integrityd
,c
with EBIb
Go/G1
(%)
S
(%)
G2/M
(%)
14
R ¼ 2-Me, 5-Me
1
1
0
20
16
5
79
83
95
15
16
R ¼ 3-Me
R ¼ 3-Et
17
19
R ¼ 3-OMe
10
29
20
61
78
R ¼ 3-Br
2
20
22
26
R ¼ 3-Cl
5
33
21
28
62
78
59
R ¼ 3-Me, 5-Me
1
R ¼ 3-OMe, 5-OMe
13

S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
5333
Table 4 (continued )
Compd
R, R1
Cell cycle progressiona
Competition
Cytoskeleton integrityd
,c
with EBIb
Go/G1
(%)
S
(%)
G2/M
(%)
27
R ¼ 3-OMe, 4-OMe, 5-OMe
12
27
2
12
22
0
74
51
98
H
N
48
R1
¼
HN
CA-4
EBI
N/A
61
N/A
30
N/A
N/A
DMSO
9
a
For cell cycle progression, M21 cells were incubated for 24 h, in presence of PIB-SAs at 5-times their respective IC50
.
b
c
For the competition assay using EBI, MDA-MB-231 cells were incubated for 2 h in presence of PIB-SAs at 1000-times their IC50 then for 1.5 h in presence of EBI.
Inhibition of EBI coding: þþþ: Strong inhibition; þþ: mild inhibition; þ: weak inhibition; -: no inhibition; N/A: not applicable.
d
M21 cells were incubated for 16 h with the drugs at 5-times their IC50 and the cytoskeleton was visualized using indirect immunofluorescence techniques.
formation of the
b
-tubulin-EBI adduct, PIB-SAs 15e17, 19, 20, 22, 26
with DMSO are exhibiting homogenous, linear and structured
cytoskeletons while all PIB-SAs tested clearly exhibit disrupted
microtubular structures.
and 27 strongly abrogated the binding of EBI to the C-BS in a similar
fashion as observed with CA-4.
4.5. PIB-SAs disrupt the cytoskeleton of tumor cells
4.6. CoMSIA models
Table 4 shows the cytoskeleton of M21 cells treated with PIB-SAs
at 5-times their respective IC50 for 16 h. Control M21 cells treated
In all CoMSIA models, the contour maps of each type field have
very similar coverage areas for different biological activities. Here we
Fig. 2. (A) Alignment hypothesis generated using Surflex-Sim mutual alignment module from compounds 15, 20, 22, 27, 40, and 48. (B) Superposition of in the derivatives of PIB-SAs
onto the alignment hypothesis.

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S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
descriptor in M21 cell line (Model E) are presented in supporting
information. The models showed that compounds with higher
similarity to the hypothesis generated from the active compounds
tend to have higher activities. In Fig. 4, compound 20 was used as
a reference to show the contour map of the CoMSIA model. Fig. 4A
shows the favored (green) and disfavored (yellow) area of steric field.
Most area surrounding the molecule was in favor of steric field. Thus
bulky groups have positive contributions to the biological activity
when present in ortho and meta positions (compounds 9, 10, 12e16,
21 and 22). Bulky groups in the 4-position were unfavorable for the
activity. That explains most of the compounds bearing bulky groups
on position 4 such as compounds 28e30, 31-38, 42, and 43 exhibit
lower activities. Compounds substituted on position 5 exhibit
significantly higher antiproliferative activities (14, 22, 26 and 27). The
effects of electrostatic field are shown in Fig. 4B. Favored electrostatic
field are in blue and disfavored electrostatic field are in red. The
region around the para-position accommodates electronegative
substituents leading to favorable biological activities (39 and 40).
Compounds 24, 25, 37, 38 bear highly electronegative atoms such as
oxygen and fluorine that are contributing to weaker activities than
that of compound 8. Fig. 4C illustrates the contribution of hydro-
phobicity to the antiproliferative activity. Yellow color shows favored
hydrophobic region and cyan color shows disfavored hydrophobic
region. To that end, the most favorable hydrophobic region is around
position 3 and 4, although the hydrophobic character is unfavorable
in the region on top of the aromatic cloud of the phenyl moiety. This
explains in part the potent cytocidal activities of compounds 12, 13,
15, 16, 19, 20. From Fig. 4D and E, there are two regions that are
Fig. 3. Growth inhibition of human HT-1080 fibrosarcoma tumors by selected PIB-SAs
using the chick chorioallantoic membrane tumor assay (CAM assay). The white
histograms represent the percentage of tumor-wet weight of tumor treated by PIB-SAs,
CA-4 and the excipient. The black histogram is the percentage of chick mortality.
took the QSAR CoMSIA contour map of M21 as an example. The
optimized CoMSIA model included descriptors of 3D molecular
similarity (Surflex-Sim searching program to generate alignments of
molecules), polar volume, and molecular energy. The predicted PIB-
SA molecular activity values of model A, B, and C are presented in
supporting information. CoMSIA model predicted activities of
models D, E, and F and contour maps of CoMSIA fields contributing to
ligand binding generated by PLS analysis without using similarity
Fig. 4. Contour maps of CoMSIA fields contributing to ligand binding generated by PLS analysis in Model B (M21). Compound 20 (stick model) is shown as a reference to depict the
field regions. (A) Contour map of steric field. Green colored map presents favored steric groups, and yellow colored map presents disfavored steric groups. (B) Contour map of
Electrostatic field. Blue color maps favored electrostatic field (larger electropositive charge will increase the activity) and red color maps disfavored electrostatic field (smaller
electropositive charge will increase the activity). (C) Contour map of hydrophobic field. Yellow color map shows favored hydrophobic region and cyan color map shows disfavored
hydrophobic region. (D) Contour map of hydrogen bond acceptor field. Magenta map shows the favored hydrogen bond acceptor region and red map illustrates hydrogen bond
disfavored region. (E) Contour map of hydrogen bond donor field. Cyan region is hydrogen bond donor preferred region and purple region is the region which hydrogen bond donor
is not favored.

S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
5335
Fig. 5. Contour maps of CoMFA fields contributing to ligand binding generated by PLS analysis in model H (M21). Compound 20 (stick model) was shown in the figure as a reference
to depict the field region. (A) Contour map of Steric field. Green map presents the favored steric interaction from the ligands and the yellow map shows the region that disfavored
steric contribution. (B) Contour map of Electrostatic field. Blue color map depicts favored electrostatic region, increasing positive charge will contribute to higher activity. Red color
map shows the disfavored electrostatic area, that higher ligand binding do not like higher positive charge.
preferred hydrogen bond acceptors: one is the ketyl moiety of the
imidazolidone and the other is the sulfonamide group bridging the
phenyl rings. The hydrogen bond donor preferred region is adjacent
to nitrogen atom of the sulfonamide bridge.
inhibition of the tumor growth similar to CA-4 (45% of the control)
with no related toxicity (11% mortality).
5. Conclusions
4.7. CoMFA models
We identified and described a novel class of antimicrotubule
agents acting through its binding to C-BS on tubulin designated as
phenyl 4-(2-oxoimidazolidin-1-yl)benzenesulfonamides (PIB-SAs).
The aim of this study was to investigate the effect of replacing the
sulfonate bridge moiety of PIB-SOs by the sulfonamide reciprocal
bioisosteres leading to the formation of PIB-SAs 8e48. PIB-SAs were
synthesized in fair to good yields. They exhibited antiproliferative
activities on sixteen cancer cell lines in the nanomolar range and
arrested the cell cycle progression in G2/M phase. Competition
In CoMFA models, M21 activity was used to exemplify our
results (Fig. 5). The same descriptors we used in CoMSIA model
were also used in this model. The predicted antiproliferative
activities for the PIB-SA derivatives based on models G, H, and I are
presented in supporting information. CoMFA model predicted
activities of models J, K, and L and CoMFA fields contributing to
ligand binding generated by PLS analysis without using similarity
descriptor in M21 cells (Model K) are presented in supporting
information. In CoMFA model, except a small area around position 4
and 5 (yellow region in Fig. 5A), hydrophobic moieties are accept-
able at ortho, meta and para-positions of phenyl (green maps in
Fig. 5A). In terms of electrostatic features, electropositive groups
are preferred on position 3 of the phenyl ring (blue map in Fig. 5B)
and electronegative groups are preferred on position 5 of the
phenyl ring (red maps in Fig. 5B).
assays using EBI and immunofluorescence using anti-b-tubulin
antibody confirmed that PIB-SAs are potent antimicrotubule agents
binding to the C-BS. In addition, the cytotoxicity of PIB-SAs was not
or only slightly affected in chemoresistant cells resistant to
colchicine, paclitaxel and vinblastine and in cells bearing the P-
glycoprotein. Moreover, quantitative structure-activity relation-
ships of PIB-SA derivatives using CoMSIA and CoMFA were estab-
lished. These studies provide the evidence that sulfonate and
sulfonamide moiety are bioisosteres and phenylimidazolidin-2-one
moieties could mimic the trimethoxyphenyl moiety found in the
structure of numerous potent antimicrotubule agents. Finally, PIB-
SAs 16 and 17 exhibited potent antitumoral and antiangiogenic
potency in the CAM assay, with lower toxicity than CA-4 on chick
embryos, making this class of drugs as promising anticancer agents.
4.8. PIB-SAs inhibit the growth of solid tumors in the CAM assay
model
The growth of solid tumors on the surface of the chick chorio-
allantoic membrane depends on the ability of the grafted tumor
cells to stimulate angiogenesis and to grow significantly within a 7-
day period. HT-1080 human fibrosarcoma cells were used because
they produce solid tumors that are sensitive to antiangiogenic and
antitumoral drugs [51e55]. As shown in Fig. 3, chick embryos have
well tolerated the excipient (ethanol: cremophor: phosphate buffer
saline: 6.25: 6.25: 87.5) when compared to control embryos (10%
6. Experimental protocols
6.1. Biological methods
6.1.1. Cell lines culture
and 4% mortality, respectively). All drugs were tested at 3
per egg with the exception of CA-4 that was used as a positive
control and tested at 1 g per egg (18% mortality); 3 g of CA-4 per
m
g of drug
HT-29 human colon carcinoma, MCF7 human breast carcinoma,
MDA-MB-231 human breast carcinoma, HT-1080 human fibrosar-
coma, K562 human chronic myelogenous leukemia, L1210 murine
lymphocytotic leukemia, P388D1 murine macrophages, B16F0
murine skin melanoma, DU 145 human prostate carcinoma and
SKOV3 human ovarian were purchased from the American Type
Culture Collection (Manassas, VA). Chinese hamster ovary cells
(CHO), colchicine- and vinblastine-resistant CHO-VV 3-2 cells and
paclitaxel-resistant CHO-TAX 5-6 cells were generously provided
by Dr. Fernando Cabral (University of Texas Medical School, Hous-
ton, TX) [43,44]. T cell leukemia CEM cells and multidrug-resistant
leukemia CEM-VLB were generously provided by Dr. William T Beck
(University of Illinois at Chicago, College of Pharmacy, USA) [45].
M21 human skin melanoma cells were provided by Dr. David
Cheresh (University of California, San Diego School of Medicine,
USA). B16F0, DU 145, HT-29, HT-1080, M21, MCF7, MDA-MB-231
m
m
egg exhibiting high toxicity/mortality rate on chick embryos (45%
mortality). CA-4 exhibited a strong vascular disruption and inhi-
bition of tumor growth by 48 and 45%, respectively. Compounds 14,
15, 20, 22 and 48 had no significant inhibitory effect on the growth
of tumor. Except for compound 14 which showed 27% mortality of
embryos, compounds 15, 20, 22 and 48 showed low toxicity (8e18%
mortality of embryos). Compounds 19, 26 and 27 weakly inhibited
the growth of tumor (81, 69 and 75% of the control). However,
except for compound 26, which exhibited low toxicity (13%
mortality), compounds 19 and 27 showed toxicity (24 and 40%
mortality, respectively). Compound 17 inhibited significantly the
growth of tumor (59% of the control) and showed low toxicity with
16% mortality of embryos. Finally, compound 16 exhibited good

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S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
and SKOV3 cells were cultured in DMEM medium containing
sodium bicarbonate, high glucose concentration, glutamine and
sodium pyruvate (Hyclone, Logan, UT) supplemented with 5% of
calf serum. CHO, CHO-VV 3-2, CHO-TAX 5-6, CEM, CEM-VLB, K562,
L1210 and P388D1 cells were cultured in RPMI medium (Hyclone,
Logan, UT) supplemented with 10% of calf serum. The cells were
maintained at 37 ꢀC in a moisture-saturated atmosphere containing
5% CO2.
adherent cells were harvested using a rubber policeman, and
centrifuged for 3 min at 8000 rpm. The pellets were washed with
500
m
L of cold PBS and stored at ꢂ80 ꢀC until use. The cells pellets
were resuspended in PBS and lysed by sonication. The protein
concentration was determined using the Bio-Rad protein assay
(Bio-Rad laboratories, Mississauga, Canada) Samples were prepared
to obtain protein at a concentration of 2 mg/ml in Laemmli [56]
sample (60 mM Tris-Cl pH 6.8, 2% SDS, 10% glycerol, 5% b-mer-
captoethanol, 0.01% bromophenol blue). Cell extracts were boiled
for 5 min. Twenty micrograms of proteins from the protein extracts
were subjected to electrophoresis using 10% polyacrylamide gels.
The proteins were transferred onto nitrocellulose membranes that
were incubated with TBSMT (Tris-Buffered Saline þ 0.1% (v/v)
Tween-20 with 2.5% fat-free dry milk) for 1 h at room temperature,
6.1.2. Antiproliferative activity assay
The antiproliferative activity of PIB-SAs (8e48) was assessed
using the procedure described by the National Cancer Institute for
its drug screening program with minor modifications [42]. 96-well
microtiter plates were seeded with 75 mL of tumor cell (for HT-29,
5000 cells; M21, 3500 cells; MCF7, 7500 cells; CHO, 1000 cells;
K562, 5000 cells; L1210, 6000 cells; P388D1, 18,000 cells; B16F0,
2000 cells; DU 145, 5000 cells; HT-1080, 3000 cells; MDA-MB-231,
3000 cells; SKOV3, 5000 cells; CEM, 20000 cells; CHO-VV 3-2, 1000
cells; CHO-TAX 5-6, 1000 cells; CEM-VLB, 20000 cells) in appro-
priate medium. Plates were incubated at 37 ꢀC, 5% CO2 for 24 h.
Freshly solubilized drugs in DMSO were diluted in fresh medium
and then with the anti-b-tubulin (clone TUB 2.1) (SigmaeAldrich,
St. Louis, MO) primary antibody in TBSMT (1:500) for 16 h at 4 ꢀC.
Membranes were washed with TBST (Tris-Buffered Saline
with þ0.1% (v/v) Tween-20) and incubated with peroxidase-
conjugated anti-mouse immunoglobulin (Amersham Canada,
Oakville, Canada) in TBSMT (1:2500) for 2.5 h at room temperature.
After washing the membranes with TBST, detection of the immu-
noblot was carried out with an enhanced chemiluminescence
detection reagent kit provided by Amersham Canada (Oakville,
Canada).
and aliquots of 75 mL aliquots containing escalating concentrations
(1000e0.98 nM) of the drug were added. Plates were incubated for
48 h. Plates containing attached cell lines were then stained with
sulforhodamine B. Briefly, cells were fixed by addition of cold tri-
chloroacetic acid to the wells (10% (w/v) final concentration), for
30 min at 4 ꢀC. Plates were washed five-times with tap water and
6.1.5. Fluorescence microscopy
M21 cells were seeded at 1 ꢁ 105 cells per well in 6-well plates
dried. Sulforhodamine B solution (50
mL) at 0.1% (w/v) in 1% acetic
that contained 22-mm glass coverslips coated with fibronectin
acid was added to each well, and plates were incubated for 15 min
at room temperature. Unbound dye was removed by washing five-
times with 1% acetic acid. Bonded dye was solubilized in 10 mM Tris
(10 m
g/mL) and incubated for 24 h at 37 ꢀC. Tumor cells were
incubated either with most potent PIB-SAs 14e17, 19, 20, 22, 26, 27,
48 or compound 2 at five-times their respective IC50 or DMSO
(0.5%) for 16 h. Afterward, the cells were washed twice with PBS,
and then fixed with 3.7% formaldehyde in PBS for 20 min. After two
washes with PBS, the cells were permeabilized with 0.1% saponin in
PBS and blocked with 3% (w/v) BSA in PBS for 1 h at 37 ꢀC. The cells
base, and the absorbance was read with a
mQuant Universal
Microplate Spectrophotometer (Biotek, Winooski, VT) at wave-
length between 530 and 565 nm according the color intensity. For
cells in suspension resazurin staining was used. Briefly, supernatant
was aspirated and 100
ml of resazurin at 25
mg/ml in fresh medium
were then incubated for 2 h at 37 ꢀC with the anti-
b-tubulin (clone
were added. Plates were incubated at 37 ꢀC for 1e3 h according to
cell line sensitivity. Fluorescence was read on an FL-500 Fluorom-
eter (Biotek, Winooski, VT) using 530 nm for excitation wavelength
and 590 nm for emission wavelength. The experiments were per-
formed at least in triplicate. The IC50 assay was considered valid
when the relative standard deviation was less than 10%.
TUB 2.1) in a solution containing 0.1% saponin and 3% BSA in PBS
(1:200). The cells were washed five-times with PBS containing
0.05% Tween 20Ô and incubated for 1 h at 37 ꢀC in blocking buffer
containing anti-mouse IgG Alexa-488 (Molecular Probes, Eugene,
OR) (1:1000), 40,60-diamidino-2-phenylindole (2.5
mg/mL in PBS) to
stain the cellular nuclei (1:2000). Cells were then mounted on
a microscope slide overnight with slow fade reagent (DakoCyto-
mation, Carpinteria, CA) before analysis using a Olympus BX51
microscope. Images were captured as 8-bit tagged image format
files with a Q imaging RETIGA EXI digital camera driven by Image
Pro Express software.
6.1.3. Cell cycle analysis
After incubation of 2.5 ꢁ 105 M21 cells with the drugs at 2- and
5-times their respective IC50, for 24 h, the cells were trypsinized,
washed with Phosphate Buffered Saline (PBS), resuspended in
250
mL of PBS, and fixed by the addition of 750 mL of ice-cold
ethanol and stored at ꢂ20 ꢀC until use. Thereafter, the cells were
6.1.6. CAM assay
centrifuged for 5 min at 1000 g. Cell pellets were washed with PBS
Human HT-1080 fibrosarcoma cells were used to assess the
antitumoral activity of PIB-SAs in the CAM assay [52,53,57]. Briefly,
fertilized chicken eggs purchased from Couvoirs Victoriaville (Vic-
toriaville, Qc, Canada) were incubated for 10 days in a Pro-FI egg
incubator fitted with an automatic egg turner before being trans-
ferred to a Roll-X static incubator for the rest of the incubation time
(both incubators were purchased from Lyon Electric, Chula Vista,
San Diego, CA). The eggs were kept at 37 ꢀC in a 60% humidity
atmosphere for the whole incubation period. On day-10, using
a hobby drill (Dremel, Racine, WI), a hole was drilled on the side of
the egg, and a negative pressure was applied to create a new air sac.
A window was opened on this new air sac and was covered with
transparent adhesive tape to prevent contamination. A freshly
and were resuspended in 450
mL of PBS containing 200 mg/mL of
RNase. After 5 min, 25 L of PBS containing 1 mg/mL of propidium
m
iodide was added. Mixtures were incubated on ice for 1 h and then
cell cycle distribution was analyzed using an Epics Elite ESP flow
cytometer (Coulter Corporation, Miami, FL).
6.1.4. Inhibition of EBI binding to b-tubulin
6-well plates were seeded with MDA-MB-231 cells at
7 ꢁ 105 cells per well and incubated for 24 h. Cells were first
incubated in presence of approximately 1000-times the IC50 of the
drugs for 2 h and afterward they were treated by the addition of EBI
(Toronto Research Chemicals, North York, Ontario, Canada)
(100
m
M, final concentration) for 1.5 h at 37 ꢀC without changing
prepared cell suspension (40
m
L) of HT-1080 (3.5 ꢁ 105 cells/egg)
the culture medium, which contains the drug tested. Control cells
were treated with 0.5% dimethylsulfoxide. Afterward, floating and
cells was applied directly on the freshly exposed chick chorioal-
lantoic membrane tissue through the window. On day-11, the

S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
5337
tested drugs dissolved in DMSO were extemporaneously diluted in
the excipient (6.25% of ethanol 99%, 6.25% of cremophor and 87.5%
phosphate buffer saline). The tested drugs in 100 mL of excipient
126.0, 126.0, 126.0, 116.1, 44.4, 36.5, 17.4; HRMS (ESþ) m/z found
331.9980; C16H17N3O3S (Mþ þ H) requires 332.1069.
were injected iv in 10e12 eggs. The eggs were incubated until day-
17, at which time the embryos were euthanized at 4 ꢀC followed by
decapitation. Tumors were collected, and the tumor-wet weights
were recorded. The number of dead embryos and signs of toxicity
from the different groups were recorded.
6.3.3. N-(2-Ethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (10)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 40%;
White solid; mp: 171e172 ꢀC; IR: 3240, 2973, 1698 cmꢂ1 1H NMR
;
(CDCl3 and a few drops of CD3OD) d 7.62e7.45 (m, 4H, Ar), 7.06e6.80
(m, 4H, Ar), 3.84e3.76 (m, 2H, CH2), 3.50e3.45 (m, 2H, CH2), 2.36 (q,
2H, J ¼ 7.5 Hz, CH2), 0.95 (t, 3H, J ¼ 7.5 Hz, CH3); 13C NMR (CDCl3 and
6.2. Chemistry
few drops of CD3OD)
d 159.4, 144.0, 138.7, 133.8, 132.5, 128.9, 128.1,
126.6,126.4,125.3,116.6, 44.8, 36.9, 23.5,14.1; HRMS (ESþ) m/z found
Proton NMR spectra were recorded on a Bruker AM-300 spec-
trometer (Bruker, Germany). Chemical shifts (d) are reported in
346.0042; C17H19N3O3S (Mþ þ H) requires 346.1225.
parts per million. IR spectra were recorded on a Magna FT-IR
spectrometer (Nicolet instrument corporation, Madison, WI, USA).
Uncorrected melting points were determined on an electrothermal
melting point apparatus. HPLC analyses were performed on an
Acquity UPLC Sample and binary solvent manager equipped with
a Quattro PremierÔ XE tandem quadrupole mass spectrometer
(Waters, Milford, MA, USA). A Waters BECH C18 reversed-phase
6.3.4. N-(2-Methoxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (11)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 53%;
White solid; mp: 209e211 ꢀC; IR: 3279, 2906, 1697 cmꢂ1; 1H NMR
(CDCl3, and a few drops of CD3OD and DMSO-d6)
d 7.58e7.51 (m,
4H, Ar), 7.29 (d, 1H, J ¼ 7.8 Hz, Ar), 6.96 (t, 1H, J ¼ 7.8 Hz, Ar),
6.79e6.69 (m, 2H, Ar), 3.81e3.78 (m, 2H, CH2), 3.52 (s, 3H, CH3),
3.45e3.40 (m, 2H, CH2); 13C NMR (CDCl3, and a few drops of CD3OD
column (1.7
m
m, 2.1 ꢁ 50 mm, 50 ꢀC) was eluted in 7 min with
a methanol/water linear gradient containing 0.1% TFA at 0.6 mL/
min. The purity of compounds was greater than 95%. All reactions
were conducted under a dried nitrogen atmosphere. Chemicals
were supplied by Aldrich Chemicals (Milwaukee, WI, USA) or VWR
International (Mont-Royal, Qc, Canada). Liquid flash chromatog-
and DMSO-d6) d 158.4, 150.6, 144.0, 131.5, 127.5, 125.5, 125.4, 122.5,
120.2, 115.7, 110.8, 55.2, 44.2, 36.3; HRMS (ESþ) m/z found
347.9580; C16H17N3O4S (Mþ þ H) requires 348.1018.
6.3.5. N-(2,3-Dimethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (12)
raphy was performed on silica gel F60, 60 A, 40e63 mm (Silicycle,
Québec, Qc, Canada) using an FPX flash purification system (Biot-
age, Charlottesville, VA, USA) and using the indicated solvent
mixture expressed as volume/volume ratios. Solvents and reagents
were used without purification unless specified otherwise. The
progress of all reactions was monitored using TLC on precoated
silica gel plates 60 F254 (VWR international, Mont-Royal, Qc,
Canada). The chromatograms were viewed under UV light at 254
and/or 265 nm.
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 55%;
White solid; mp: 231e232 ꢀC; IR: 3257, 2902, 1703 cmꢂ1; 1H NMR
(DMSO-d6 and a few drops of CDCl3)
d 8.87 (s, 1H, NH), 7.57e7.49
(m, 4H, Ar), 6.90e6.80 (m, 3H, Ar and NH), 6.74e6.71 (m, 1H, Ar),
3.86e3.81 (m, 2H, CH2), 3.49e3.43 (m, 2H, CH2), 2.12 (s, 3H, CH3),
1.92 (s, 3H, CH3); 13C NMR (DMSO-d6 and a few drops of CDCl3)
d
158.4, 143.6, 137.0, 134.3, 133.2, 132.3, 127.5, 127.3, 124.8, 124.2,
115.6, 44.1, 36.3, 20.0, 13.6; HRMS (ESþ) m/z found 346.1540;
C17H19N3O3S (Mþ þ H) requires 346.1225.
6.3. General preparation of compounds 8e48
6.3.6. N-(2,4-Dimethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (13)
Compound 51 (1.00 mmol) was suspended in dry acetonitrile
(10 mL) under nitrogen atmosphere. Relevant aniline (1.00 mmol)
and 4-dimethylaminopyridine (4.00 mmol) were successively
added dropwise and the mixture was stirred at room temperature
for 48 h. Ethyl acetate was added and the solution was washed with
hydrochloric acid 1N, brine, dried over sodium sulfate, filtered, and
evaporated to dryness under vacuum. The white solid was purified
by flash chromatography on silica gel.
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 55%;
White solid; mp: 238e239 ꢀC; IR: 3463, 3283, 1704 cmꢂ1; 1H NMR
(CDCl3 and a few drops of DMSO-d6)
4H, Ar), 6.93 (s, 1H, NH), 6.82e6.74 (m, 3H, Ar), 3.87e3.81 (m, 2H,
CH2), 3.48e3.43 (m, 2H, CH2), 2.16 (s, 3H, CH3), 1.93 (s, 3H, CH3); 13
NMR (CDCl3 and a few drops of DMSO-d6) 158.3,143.7,135.3,133.8,
d 8.86 (s,1H, NH), 7.58e7.48 (m,
C
d
132.4, 131.9, 130.8, 127.2, 126.3, 115.6, 44.1, 36.3, 20.3, 17.3; HRMS
(ESþ) m/z found 345.9970; C17H19N3O3S (Mþ þ H) requires 346.1225.
6.3.1. N-Phenyl-4-(2-oxoimidazolidin-1-yl)benzenesulfonamide (8)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 29%;
6.3.7. N-(2,5-Dimethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (14)
White solid; mp: 262e264 ꢀC; IR: 3434, 1686 cmꢂ1 1H NMR
;
(DMSO-d6 and a few drops of CDCl3)
d
7.63e7.53 (m, 4H, Ar),
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 46%;
7.11e7.02 (m, 4H, Ar), 6.95e6.88 (m, 1H, Ar), 3.82e3.78 (m, 2H,
White solid; mp: 213e215 ꢀC; IR: 3274, 1695 cmꢂ1 1H NMR
;
CH2), 3.44e3.39 (m, 2H, CH2); 13C NMR (DMSO-d6 and CDCl3)
(DMSO-d6 and a few drops of CDCl3)
d 9.05 (s, 1H, NH), 7.61e7.50
d
158.3,143.9,137.5,131.3,128.5,127.4,123.4,119.8,115.8, 44.0, 36.3;
(m, 4H, Ar), 7.08 (s, 1H, NH), 6.90e6.79 (m, 3H, Ar), 3.87e3.82 (m,
2H, CH2), 3.47e3.42 (m, 2H, CH2), 2.15 (s, 3H, CH3), 1.89 (s, 3H, CH3);
HRMS (ESþ) m/z found 318.0634; C15H15N3O3S (Mþ þ H) requires
318.0912.
13C NMR (DMSO-d6 and a few drops of CDCl3)
d 158.2, 143.8, 134.9,
134.5, 132.4, 130.3, 130.0, 127.2, 126.6, 126.4, 115.6, 44.1, 36.3, 20.3,
17.0; HRMS (ESþ) m/z found 346.0067; C17H19N3O3S (Mþ þ H)
requires 346.1225.
6.3.2. N-2-Tolyl-4-(2-oxoimidazolidin-1-yl)benzenesulfonamide (9)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 45%;
White solid; mp: 209e210 ꢀC; IR: 3234, 2920, 1694 cmꢂ1; 1H NMR
(CDCl3 and a few drops of CD3OD and DMSO-d6)
d
7.55e7.49 (m, 4H,
6.3.8. N-3-Tolyl-4-(2-oxoimidazolidin-1-yl)benzenesulfonamide (15)
Ar), 6.97e6.94 (m, 4H, Ar), 3.86e3.80 (m, 2H, CH2), 3.48e3.42 (m,
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 64%;
2H, CH2), 1.95 (s, 3H, CH3); 13C NMR (CDCl3, and a few drops of
White solid; mp: 188e190 ꢀC; IR: 3419, 3077, 1697 cmꢂ1 1H NMR
;
CD3OD and DMSO-d6)
d
158.8, 143.9, 134.6, 133.7, 132.5, 130.4, 127.5,
(DMSO-d6 and a few drops of CDCl3) d 9.60 (s, 1H, NH), 7.63e7.52

5338
S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
(m, 4H, Ar), 6.97e6.92 (m, 1H, Ar), 6.84e6.82 (m, 2H, Ar or NH),
6.75e6.70 (m, 2H, Ar or NH), 3.83e3.77 (m, 2H, CH2), 3.46e3.41 (m,
2H, CH2), 2.15 (s, 3H, CH3); 13C NMR (DMSO-d6 and a few drops of
6.3.14. N-(3,4-Dimethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (21)
Flash chromatography (AcOEt to AcOEt/MeOH 95:5). Yield: 73%;
1
CDCl3)
d
158.3, 143.6, 138.0, 137.3, 131.5, 128.1, 127.3, 124.2, 120.4,
White solid; mp: 206e208 ꢀC; IR: 3413, 3256, 1700 cmꢂ1; H NMR
116.8, 115.7, 44.1, 36.3, 20.8; HRMS (ESþ) m/z found 332.1365;
(DMSO-d6) d 9.92 (s, 1H, NH), 7.70e7.63 (m, 4H, Ar), 7.26 (s, 1H, NH),
C16H17N3O3S (Mþ þ H) requires 332.1069.
6.97e6.94 (m, 1H, Ar), 6.88e6.87 (m, 1H, Ar), 6.83e6.80 (m, 1H, Ar),
3.88e3.83 (m, 2H, CH2), 3.43e3.39 (m, 2H, CH2), 2.11 (s, 3H, CH3),
6.3.9. N-(3-Ethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (16)
2.09 (s, 3H, CH3); 13C NMR (DMSO-d6)
d 158.4, 144.3, 136.8, 135.6,
131.8,131.4,129.9,127.7,121.5,117.6,116.2, 44.2, 36.4,19.6,18.6; HRMS
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
(ESþ) m/z found 346.0768; C17H19N3O3S (Mþ þ H) requires 346.1225.
MeOH 95:5). Yield: 72%; White solid; mp: 188e190 ꢀC; IR: 3400,
2961, 1697 cmꢂ1; 1H NMR (DMSO-d6)
d
10.08 (s, 1H, NH), 7.69 (brs,
6.3.15. N-(3,5-Dimethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (22)
4H, Ar), 7.26 (brs, 1H, NH), 7.14e7.09 (m, 1H, Ar), 6.95e6.84 (m, 3H,
Ar), 3.87e3.82 (m, 2H, CH2), 3.43e3.38 (m, 2H, CH2), 2.49 (q, 2H,
J ¼ 7.5 Hz, CH2), 1.09 (t, 3H, J ¼ 7.5 Hz, CH3); 13C NMR (DMSO-d6)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 68%;
White solid; mp: 216e218 ꢀC; IR: 3406,1699 cmꢂ1; 1H NMR (DMSO-
d
158.4, 144.7, 144.4, 138.0, 131.3, 129.0, 127.7, 123.3, 119.1, 117.1,
d6 and a few drops of CDCl3) d 9.66 (s,1H, NH), 7.64e7.54 (m, 4H, Ar),
116.2, 44.2, 36.4, 28.1, 15.4; HRMS (ESþ) m/z found 346.1010;
6.97 (s, 1H, NH), 6.66 (s, 2H, Ar), 6.54 (s, 1H, Ar), 3.84e3.79 (m, 2H,
C17H19N3O3S (Mþ þ H) requires 346.1225.
CH2), 3.45e3.40 (m, 2H, CH2), 2.11 (s, 6H, 2ꢁ CH3); 13C NMR (DMSO-
d6 and a few drops of CDCl3) d 158.3, 143.8, 137.7, 137.4, 131.5, 127.3,
6.3.10. N-(3-Methoxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (17)
125.0,117.3,115.7, 44.0, 36.3, 20.7; HRMS (ESþ) m/z found 346.0776;
C17H19N3O3S (Mþ þ H) requires 346.1225.
Flash chromatography (AcOEt to AcOEt/MeOH 95:5). Yield:
52%; White solid; mp: 179e181 ꢀC; IR: 3428, 3084, 1700 cmꢂ1; 1H
6.3.16. N-(3,5-di-tert-butylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (23)
NMR (DMSO-d6)
d 10.17 (brs, 1H, NH), 7.70 (brs, 4H, Ar), 7.26 (brs,
1H, NH), 7.14e7.09 (m, 1H, Ar), 6.68e6.67 (m, 2H, Ar), 6.60e6.56
(m, 1H, Ar), 3.88e3.83 (m, 2H, CH2), 3.66 (s, 3H, CH3), 3.43e3.38
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
MeOH 95:5). Yield: 86%; White solid; mp: 276e278 ꢀC; IR: 3400,
(m, 2H, CH2); 13C NMR (DMSO-d6)
d
159.7, 158.4, 144.4, 139.3,
3107, 1703 cmꢂ1; 1H NMR (DMSO-d6)
d 9.95 (s, 1H, NH), 7.73e7.66
131.1, 130.0, 127.7, 116.2, 111.7, 108.8, 105.5, 55.0, 44.2, 36.4; HRMS
(ESþ) m/z found 348.0766; C16H17N3O4S (Mþ þ H) requires
348.1018.
(m, 4H, Ar), 7.27 (brs,1H, NH), 7.04 (s,1H, Ar), 6.95e6.93 (m, 2H, Ar),
3.88e3.83 (m, 2H, CH2), 3.45e3.40 (m, 2H, CH2), 1.20 (s, 18H, 6ꢁ
CH3); 13C NMR (DMSO-d6)
d 158.4, 151.0, 144.4, 137.4, 131.4, 127.8,
117.3, 116.1, 114.3, 44.3, 36.3, 34.5, 31.1; HRMS (ESþ) m/z found
6.3.11. N-(3-Phenoxyphenyl)-4-(2-Oxoimidazolidin-1-yl)
benzenesulfonamide (18)
430.2245; C23H31N3O3S (Mþ þ H) requires 430.2164.
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 65%;
6.3.17. N-(3,4-Difluorophenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (24)
White solid; mp: 208e211 ꢀC; IR: 3389, 1696 cmꢂ1 1H NMR
;
(DMSO-d6)
d
10.26 (brs, 1H, NH), 7.73e7.63 (m, 4H, Ar), 7.43e7.37
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 37%;
(m, 2H, Ar), 7.31 (brs, 1H, NH), 7.26e7.15 (m, 2H, Ar), 6.93e6.86 (m,
3H, Ar), 6.73e6.64 (m, 2H, Ar), 3.91e3.86 (m, 2H, CH2), 3.47e3.41
White solid; mp: 225e226 ꢀC; IR: 3430, 3129, 1695 cmꢂ1; 1H NMR
(CDCl3 and a few drops of CD3OD and DMSO-d6) d 7.62e7.53 (m, 4H,
(m, 2H, CH2); 13C NMR (DMSO-d6)
d
158.4, 157.3, 156.1, 144.6, 139.6,
Ar), 6.99e6.87 (m, 2H, Ar), 6.79e6.74 (m, 1H, Ar), 3.86e3.80 (m, 2H,
130.8, 130.6, 130.1, 127.8, 123.8, 118.9, 116.3, 114.5, 113.7, 109.4, 44.3,
36.4; HRMS (ESþ) m/z found 410.1010; C21H19N3O4S (Mþ þ H)
requires 410.1175.
CH2), 3.48e3.43 (m, 2H, CH2); 13C NMR (CDCl3 and a few drops of
CD3OD and DMSO-d6)
d 158.7, 151.2, 151.0, 148.4, 148.2, 147.9, 147.7,
145.2, 145.0, 144.0, 134.2, 134.1, 134.1, 134.0, 130.9, 127.5, 117.0, 116.8,
116.5, 116.4, 116.4, 116.3, 116.1, 109.9, 109.6, 44.3, 36.4; HRMS (ESþ)
m/z found 354.0279; C15H13F2N3O3S (Mþ þ H) requires 354.0724.
6.3.12. N-(3-Bromophenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (19)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 21%;
6.3.18. N-(3,4-Dimethoxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (25)
White solid; mp: 184e185 ꢀC; IR: 3388, 2860, 1692 cmꢂ1; 1H NMR
(DMSO-d6)
d
10.45 (brs, 1H, NH), 7.72 (s, 4H, Ar), 7.29e7.20 (m, 4H,
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
Ar and NH), 7.14e7.10 (m, 1H, Ar), 3.90e3.84 (m, 2H, CH2),
MeOH 95:5). Yield: 59%; Brownish solid; mp: 215e216 ꢀC; IR: 3357,
3.44e3.39 (m, 2H, CH2); 13C NMR ((CD3)2CO and few drops of
3111, 1701 cmꢂ1; 1H NMR (DMSO-d6)
d 9.80 (brs, 1H, NH), 7.71e7.63
DMSO-d6 and CDCl3)
d
158.5, 144.7, 139.9, 131.3, 130.5, 127.7, 126.2,
(m, 4H, Ar), 7.27 (brs, 1H, NH), 6.81e6.78 (m, 1H, Ar), 6.73e6.72 (m,
1H, Ar), 6.58e6.54 (m, 1H, Ar), 3.90e3.84 (m, 2H, CH2), 3.67 (s, 3H,
CH3), 3.66 (s, 3H, CH3), 3.45e3.40 (m, 2H, CH2); 13C NMR (DMSO-d6)
122.1, 121.9, 118.2, 116.1, 44.3, 36.5; HRMS (ESþ) m/z found
395.9071; C15H14BrN3O3S (Mþ þ H) requires 396.0018.
d
158.4, 148.8, 145.9, 144.3, 131.2, 131.0, 127.8, 116.1, 113.2, 112.1,
6.3.13. N-(3-Chlorophenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (20)
106.3, 55.6, 55.4, 44.3, 36.4; HRMS (ESþ) m/z found 378.0635;
C17H19N3O5S (Mþ þ H) requires 378.1123.
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield:
30%; White solid; mp: 180e182 ꢀC; IR: 3386, 1696 cmꢂ1
;
1H NMR
6.3.19. N-(3,5-Dimethoxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (26)
(CDCl3 and a few drops of CD3OD and DMSO-d6)
d 7.64e7.55 (m,
4H, Ar), 7.07e7.05 (m, 2H, Ar), 6.97e6.94 (m, 1H, Ar), 6.89e6.86
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield:
(m, 1H, Ar), 3.84e3.78 (m, 2H, CH2), 3.45e3.40 (m, 2H, CH2); 13C
43%; White solid; mp: 200e201 ꢀC; IR: 3434, 3178, 1702 cmꢂ1
;
1H
NMR (CDCl3 and a few drops of CD3OD and DMSO-d6)
d
158.7,
NMR (CDCl3 and a few drops of CD3OD and DMSO-d6) d 7.67e7.56
144.3, 139.2, 133.9, 131.3, 130.0, 127.7, 123.5, 119.4, 117.8, 116.2, 44.4,
36.5; HRMS (ESþ) m/z found 352.0504; C15H14ClN3O3S (Mþ þ H)
requires 352.0523.
(m, 4H, Ar), 6.25e6.24 (m, 2H, Ar), 6.01e6.00 (m, 1H, Ar),
3.83e3.78 (m, 2H, CH2), 3.60 (s, 6H, 2ꢁ CH3), 3.44e3.38 (m, 2H,
CH2); 13C NMR (CDCl3 and a few drops of CD3OD and DMSO-d6)

S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
5339
d
160.5, 158.3, 144.0, 139.3, 131.2, 127.5, 115.9, 97.5, 95.0, 54.6, 44.0,
3112,1694 cmꢂ1; 1H NMR (DMSO-d6)
d
10.06 (s, 1H, NH), 7.70 (s, 4H,
36.2; HRMS (ESþ) m/z found 378.0471; C17H19N3O5S (Mþ þ H)
Ar), 7.26e7.23 (m, 3H, NH and Ar), 7.02e7.00 (m, 2H, Ar), 3.89e3.84
requires 378.1124.
(m, 2H, CH2), 3.44e3.39 (m, 2H, CH2), 1.20 (s, 9H, 3ꢁ CH3); 13C NMR
(DMSO-d6)
d 158.4, 146.1, 144.3, 135.3, 131.6, 127.7, 125.8, 119.7,
6.3.20. N-(3,4,5-Trimethoxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (27)
116.3, 44.2, 36.4, 34.0, 31.1; HRMS (ESþ) m/z found 374.0975;
C19H23N3O3S (Mþ þ H) requires 374.1538.
Flash chromatography (AcOEt to AcOEt/MeOH 95:5). Yield: 23%;
White solid; mp: 233e235 ꢀC; IR: 3416, 3120, 1704 cmꢂ1; 1H NMR
6.3.26. N-(4-Pentylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (33)
(DMSO-d6)
6.41 (s, 2H, Ar), 3.90e3.85 (m, 2H, CH2), 3.67 (s, 6H, 2x CH3), 3.57 (s,
3H, CH3), 3.45e3.40 (m, 2H, CH2); 13C NMR (DMSO-d6)
158.4,
d 10.31 (brs, 1H, NH), 7.72 (s, 4H, Ar), 7.27 (s, 1H, NH),
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
d
MeOH 95:5). Yield: 82%; White solid; mp: 207e209 ꢀC; IR: 3314,
153.0, 144.5, 134.0, 133.9, 131.1, 127.9, 116.2, 97.6, 60.1, 55.8, 44.2,
36.4; HRMS (ESþ) m/z found 408.0733; C18H21N3O6S (Mþ þ H)
requires 408.1229.
2927, 1705 cmꢂ1 1H NMR (DMSO-d6)
; d 9.98 (brs, 1H, NH),
7.69e7.63 (m, 4H, Ar), 7.25 (brs, 1H, NH), 7.04e6.96 (m, 4H, Ar),
3.87e3.81 (m, 2H, CH2), 3.43e3.37 (m, 2H, CH2), 2.44 (t, 2H,
J ¼ 7.5 Hz, CH2), 1.52e1.45 (m, 2H, CH2), 1.30e1.18 (m, 4H, 2ꢁ CH2),
6.3.21. N-4-Tolyl-4-(2-oxoimidazolidin-1-yl)benzenesulfonamide (28)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
MeOH 95:5). Yield: 83%; White solid; mp: 218e220 ꢀC; IR: 3430,
0.83 (t, 3H, J ¼ 6.8 Hz, CH3); 13C NMR (DMSO-d6)
d 158.4, 144.3,
138.0, 135.5, 131.3, 128.8, 127.7, 120.3, 116.2, 44.2, 36.4, 34.4, 30.9,
30.5, 21.9, 13.9; HRMS (ESþ) m/z found 388.1610; C20H25N3O3S
(Mþ þ H) requires 388.1695.
1697 cmꢂ1; 1H NMR (DMSO-d6)
d 9.97 (brs, 1H, NH), 7.70e7.63 (m,
4H, Ar), 7.26 (brs, 1H, NH), 7.04e6.96 (m, 4H, Ar), 3.88e3.82 (m, 2H,
CH2), 3.44e3.38 (m, 2H, CH2), 2.19 (s, 3H, CH3); 13C NMR (DMSO-d6)
6.3.27. N-(4-Cyclohexylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (34)
d
158.4, 144.3, 135.3, 133.1, 131.2, 129.5, 127.7, 120.4, 116.2, 44.2, 36.4,
20.3; HRMS (ESþ) m/z found 332.1114; C16H17N3O3S (Mþ þ H)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
requires 332.1069.
MeOH 95:5). Yield: 35%; White solid; mp: 256e257 ꢀC; IR: 3287,
2927,1708 cmꢂ1; 1H NMR (DMSO-d6)
d 10.04 (s,1H, NH), 7.69 (s, 4H,
6.3.22. N-(4-Ethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (29)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
MeOH 95:5). Yield: 65%; White solid; mp: 195e196 ꢀC; IR: 3250,
1695 cmꢂ1; 1H NMR (CDCl3 and a few drops of CD3OD and DMSO-
Ar), 7.27 (s, 1H, NH), 7.09e6.98 (m, 4H, Ar), 3.89e3.84 (m, 2H, CH2),
3.44e3.39 (m, 2H, CH2), 2.42e2.35 (m, 1H, CH), 1.76e1.66 (m, 5H,
5ꢁ CHequ), 1.36e1.16 (m, 5H, 5ꢁ CHax); 13C NMR (DMSO-d6)
d
158.4, 144.3, 143.2, 135.7, 131.5, 127.7, 127.2, 120.1, 116.2, 44.3, 43.0,
36.4, 33.9, 26.3, 25.6; HRMS (ESþ) m/z found 400.1554;
d6) d 7.61e7.50 (m, 4H, Ar), 6.95e6.89 (m, 4H, Ar), 3.81e3.76 (m, 2H,
C21H25N3O3S (Mþ þ H) requires 400.1695.
Ar), 3.45e3.39 (m, 2H, CH2), 2.44 (q, 2H, J ¼ 7.6 Hz, CH2), 1.06 (t, 3H,
J ¼ 7.6 Hz, CH3); 13C NMR (CDCl3 and a few drops of CD3OD and
6.3.28. N-(4-Heptylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (35)
DMSO-d6) d 158.7, 143.9, 139.9, 134.9, 131.8, 128.0, 127.7, 120.8, 116.1,
44.4, 36.5, 27.7,15.2; HRMS (ESþ) m/z found 346.0152; C17H19N3O3S
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
(Mþ þ H) requires 346.1225.
MeOH 95:5). Yield: 90%; White solid; mp: 188e190 ꢀC; IR: 3250,
2924, 1710 cmꢂ1; 1H NMR (DMSO-d6)
d 9.99 (s, 1H, NH), 7.70e7.63
6.3.23. N-(4-Butylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (30)
(m, 4H, Ar), 7.26 (s, 1H, NH), 7.05e6.97 (m, 4H, Ar), 3.88e3.83 (m,
2H, CH2), 3.44e3.38 (m, 2H, CH2), 2.45 (t, 2H, J ¼ 7.6 Hz, CH2),
1.50e1.46 (m, 2H, CH2), 1.28e1.19 (m, 8H, 4ꢁ CH2), 0.85 (t, 3H,
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
MeOH 95:5). Yield: 81%; White solid; mp: 203e205 ꢀC; IR: 3436,
J ¼ 6.5 Hz, CH3); 13C NMR (DMSO-d6)
d 158.4, 144.3, 138.0, 135.5,
2927, 1692 cmꢂ1; 1H NMR (DMSO-d6)
d
10.00 (s, 1H, NH), 7.70e7.64
131.3, 128.8, 127.7, 120.3, 116.2, 44.2, 36.4, 34.4, 31.2, 30.9, 28.6, 28.5,
22.1, 13.9; HRMS (ESþ) m/z found 416.0854; C22H29N3O3S (Mþ þ H)
requires 416.2008.
(m, 4H, Ar), 7.26 (s,1H, NH), 7.06e6.98 (m, 4H, Ar), 3.89e3.83 (m, 2H,
CH2), 3.44e3.39 (m, 2H, CH2), 2.49e2.44 (m, 2H, CH2), 1.52e1.42 (m,
2H, CH2),1.31e1.19 (m, 2H, CH2), 0.87 (t, 3H, J ¼ 7.3 Hz, CH3); 13C NMR
(DMSO-d6)
d
158.4,144.3,138.0,135.5,131.4,128.9,127.7,120.3,116.2,
6.3.29. N-(4-Methoxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (36)
44.3, 36.4, 34.1, 33.1, 21.7, 13.8; HRMS (ESþ) m/z found 374.1300;
C19H23N3O3S (Mþ þ H) requires 374.1538.
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 68%;
White solid; mp: 210e211 ꢀC; IR: 3268, 3109, 1694 cmꢂ1 1H NMR
;
6.3.24. N-(4-sec-Butylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (31)
(CDCl3 and a few drops of CD3OD and DMSO-d6) d 9.30 (s, 1H, NH),
7.56e7.49 (m, 4H, Ar), 6.93e6.90 (m, 2H, Ar), 6.75 (s, 1H, NH),
6.63e6.60 (m, 2H, Ar), 3.83e3.77 (m, 2H, CH2), 3.62 (s, 3H, CH3),
3.46e3.40 (m, 2H, CH2); 13C NMR (CDCl3 and a few drops of CD3OD
Flash chromatography (AcOEt to AcOEt/MeOH 95:5). Yield: 69%;
White solid; mp: 157e159 ꢀC; IR: 3436, 2960, 1690 cmꢂ1; 1H NMR
(DMSO-d6)
d
10.01 (s, 1H, NH), 7.71e7.65 (m, 4H, Ar), 7.26 (s, 1H,
and DMSO-d6) d 158.4, 156.3, 143.6, 129.9, 127.4, 123.4, 123.4, 115.7,
NH), 7.06e6.98 (m, 4H, Ar), 3.88e3.83 (m, 2H, CH2), 3.44e3.38 (m,
2H, CH2), 2.49e2.44 (m, 1H, CH), 1.51e1.41 (m, 2H, CH2), 1.11 (d, 3H,
J ¼ 7.0 Hz, CH3), 0.70 (t, 3H, J ¼ 7.3 Hz, CH3); 13C NMR (DMSO-d6)
113.6, 54.7, 44.1, 36.4; HRMS (ESþ) m/z found 347.9559;
C16H17N3O4S (Mþ þ H) requires 348.1018.
d
158.4, 144.3, 142.7, 135.6, 131.5, 127.7, 127.5, 120.2, 116.2, 44.2, 40.1,
6.3.30. N-(4-Butoxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (37)
36.4, 30.6, 21.5, 12.0; HRMS (ESþ) m/z found 374.1019; C19H23N3O3S
(Mþ þ H) requires 374.1538.
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
MeOH 95:5). Yield: 77%; White solid; mp: 223e225 ꢀC; IR: 3270,
6.3.25. N-(4-tert-Butylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (32)
2954, 1693 cmꢂ1; 1H NMR (DMSO-d6)
d 9.77 (s, 1H, NH), 7.70e7.59
(m, 4H, Ar), 7.27 (s, 1H, NH), 6.99e6.96 (m, 2H, Ar), 6.81e6.78 (m,
2H, Ar), 3.89e3.85 (m, 4H, 2ꢁ CH2), 3.45e3.40 (m, 2H, CH2),
1.70e1.60 (m, 2H, CH2), 1.47e1.35 (m, 2H, CH2), 0.92 (t, 3H,
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
MeOH 95:5). Yield: 84%; White solid; mp: 241e243 ꢀC; IR: 3357,

5340
S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
J ¼ 7.4 Hz, CH3); 13C NMR (DMSO-d6)
d
158.4, 155.9, 144.2, 131.2,
3136, 1699 cmꢂ1 1H NMR (DMSO-d6)
; d 10.25 (brs, 1H, NH),
130.4, 127.7, 123.2, 116.1, 114.8, 67.2, 44.3, 36.4, 30.8, 18.7, 13.7;
HRMS (ESþ) m/z found 390.1243; C19H23N3O4S (Mþ þ H) requires
390.1487.
7.74e7.70 (m, 4H, Ar), 7.27 (brs, 1H, NH), 7.23e7.11 (m, 4H, Ar), 3.92
(s, 2H, CH2), 3.89e3.84 (m, 2H, CH2), 3.44e3.39 (m, 2H, CH2); 13C
NMR (DMSO-d6)
d 158.4, 144.5, 137.5, 131.0, 128.9, 127.7, 126.5,
120.2, 119.2, 116.3, 44.2, 36.4, 21.7; HRMS (ESþ) m/z found
6.3.31. N-(4-Hexyloxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (38)
357.0658; C17H16N4O3S (Mþ þ H) requires 357.1021.
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield:
6.3.37. N-(4-Difluoromethoxyphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (44)
41%; White solid; mp: 225e226 ꢀC; IR: 3272, 2928, 1694 cmꢂ1
;
1H NMR (DMSO-d6)
d
9.74 (s, 1H, NH), 7.67e7.56 (m, 4H, Ar), 7.24
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
(s, 1H, NH), 6.96e6.93 (m, 2H, Ar), 6.78e6.75 (m, 2H, Ar),
3.86e3.81 (m, 4H, 2ꢁ CH2), 3.42e3.37 (m, 2H, CH2), 1.68e1.58 (m,
2H, CH2), 1.37e1.24 (m, 6H, 3ꢁ CH2), 0.85 (t, 3H, J ¼ 6.6 Hz, CH3);
MeOH 95:5). Yield: 81%; White solid; mp: 198e200 ꢀC; IR: 3268,
3111, 1695 cmꢂ1 1H NMR (DMSO-d6)
; d 10.21 (brs, 1H, NH),
7.73e7.66 (m, 4H, Ar), 7.28 (brs, 1H, NH), 7.15e7.06 (m, 4H, Ar), 7.12
(t, 1H, J ¼ 74.2 Hz, CHF2), 3.90e3.85 (m, 2H, CH2), 3.45e3.40 (m, 2H,
13C NMR (DMSO-d6)
d 158.3, 155.8, 144.2, 131.1, 130.2, 127.6, 123.1,
116.0, 114.7, 67.5, 44.2, 36.3, 30.9, 28.6, 25.1, 22.0, 13.8; HRMS
(ESþ) m/z found 418.1668; C21H27N3O4S (Mþ þ H) requires
418.1801.
CH2); 13C NMR (DMSO-d6)
d 158.4, 147.2, 144.5, 135.2, 131.0, 127.7,
121.7, 119.8, 116.4, 116.3, 44.2, 36.4; HRMS (ESþ) m/z found
383.9433; C16H15F2N3O4S (Mþ þ H) requires 384.0829.
6.3.32. N-(4-Bromophenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (39)
6.3.38. N-Benzo [1,3]dioxol-5-yl-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (45)
Flash chromatography (AcOEt to AcOEt/MeOH 95:5).ꢂY1ield: 57%;
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield: 54%;
White solid; mp: 237e239 ꢀC; IR: 3353, 3139, 1683 cm
;
1H NMR
White solid; mp: 219e220 ꢀC; IR: 3223, 2969, 1694 cmꢂ1; 1H NMR
(DMSO-d6)
J¼ 8.7 Hz, Ar), 7.28 (s,1H, NH), 7.05 (d, 2H, J¼ 8.7 Hz, Ar), 3.89e3.83(m,
2H, CH2), 3.44e3.38 (m, 2H, CH2); 13C NMR (DMSO-d6)
158.4, 144.6,
d
10.32 (s, 1H, NH), 7.72e7.66 (m, 4H, Ar), 7.42 (d, 2H,
(DMSO-d6) d 9.86 (brs, 1H, NH), 7.72e7.62 (m, 4H, Ar), 7.28 (brs, 1H,
NH), 6.78e6.76 (m, 1H, Ar), 6.67e6.66 (m, 1H, Ar), 6.51e6.48 (m,1H,
Ar), 5.97 (s, 2H, CH2), 3.90e3.85 (m, 2H, CH2), 3.45e3.40 (m, 2H,
d
137.5,132.0,130.7,127.7,121.7,116.3,115.9, 44.2, 36.4; HRMS (ESþ) m/z
CH2); 13C NMR (DMSO-d6)
d 158.4, 147.4, 144.4, 144.3, 131.8, 131.0,
found 395.7357; C15H14BrN3O3S (Mþ þ H) requires 396.0018.
127.7, 116.2, 114.5, 108.3, 103.2, 101.3, 44.3, 36.4; HRMS (ESþ) m/z
found 362.1037; C16H15N3O5S (Mþ þ H) requires 362.0811.
6.3.33. N-(4-Chlorophenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (40)
6.3.39. N-(9H-Fluoren-2-yl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (46)
Flash chromatography (AcOEt to AcOEt/MeOH 95:5ꢂ).1Yield: 47%;
White solid; mp: 234e236 ꢀC; IR: 3267, 1696 cm
;
1H NMR
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
(DMSO-d6)
3H, NH and Ar), 7.11e7.09 (m, 2H, Ar), 3.89e3.83 (m, 2H, CH2),
3.48e3.38 (m, 2H, CH2); 13C NMR (DMSO-d6)
158.4, 144.6, 137.0,
d 10.31 (s, 1H, NH), 7.72e7.65 (m, 4H, Ar), 7.31e7.28 (m,
MeOH 95:5). Yield: 38%; Orange solid; mp: 164e166 ꢀC; IR: 3262,
1705 cmꢂ1; 1H NMR (DMSO-d6)
d 10.21 (brs, 1H, NH), 7.78e7.66 (m,
d
6H, Ar), 7.54e7.51 (m, 1H, Ar), 7.36e7.24 (m, 4H, Ar and NH),
7.14e7.11 (m, 1H, Ar), 3.84e3.80 (m, 4H, 2 CH2), 3.41e3.37 (m, 2H,
130.8, 129.1, 127.9, 127.7, 121.4, 116.3, 44.2, 36.3; HRMS (ESþ) m/z
found 352.1068; C15H14ClN3O3S (Mþ þ H) requires 352.0522.
CH2); 13C NMR (DMSO-d6)
d 158.4, 144.4, 144.1, 142.7, 140.6, 137.2,
136.9, 131.2, 127.7, 126.8, 126.4, 125.0, 120.5, 119.6, 118.9, 117.0, 116.2,
44.2, 36.4, 36.3; HRMS (ESþ) m/z found 406.0688; C22H19N3O3S
(Mþ þ H) requires 406.1225.
6.3.34. N-(4-Fluorophenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (41)
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1). Yield:
68%; White solid; mp: 249e251 ꢀC; IR: 3446, 3023, 1697 cmꢂ1; 1H
6.3.40. N-Indan-5-yl-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (47)
NMR (CDCl3 and a few drops of CD3OD and DMSO-d6)
d 7.60e7.54
(m, 4H, Ar), 7.04e7.00 (m, 2H, Ar), 6.85e6.80 (m, 2H, Ar),
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
3.86e3.80 (m, 2H, CH2), 3.47e3.42 (m, 2H, CH2); 13C NMR (CDCl3
MeOH 95:5). Yield: 91%; White solid; mp: 228e230 ꢀC; IR: 3251,
and a few drops of CD3OD and DMSO-d6)
d
160.8, 158.5, 157.6,
2964, 1694 cmꢂ1 1H NMR (DMSO-d6)
; d 9.97 (brs, 1H, NH),
144.0, 133.6, 133.5, 131.1, 127.5, 122.8, 122.7, 116.0, 115.3, 115.0, 44.2,
36.4; HRMS (ESþ) m/z found 335.9753; C15H14FN3O3S (Mþ þ H)
requires 336.0818.
7.71e7.65 (m, 4H, Ar), 7.27 (brs, 1H, NH), 7.07e7.04 (m, 1H, Ar), 6.98
(s, 1H, Ar), 6.87e6.84 (m, 1H, Ar), 3.89e3.84 (m, 2H, CH2),
3.45e3.40 (m, 2H, CH2), 2.78e2.72 (m, 4H, 2ꢁ CH2), 2.00e1.90 (m,
2H, CH2); 13C NMR (DMSO-d6)
d 158.4, 144.6, 144.3, 139.4, 136.1,
6.3.35. N-(4-Iodophenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (42)
131.4, 127.7, 124.5, 118.4, 116.5, 116.2, 44.3, 36.4, 32.4, 31.6, 25.1;
HRMS (ESþ) m/z found 358.0985; C18H19N3O3S (Mþ þ H) requires
358.1225.
Flash chromatography (AcOEt to AcOEt/MeOH 95:5). Yield: 53%;
White solid; mp: 266e267 ꢀC; IR: 3353, 3134, 1684 cmꢂ1; 1H NMR
(DMSO-d6)
d
10.31 (brs, 1H, NH), 7.72e7.66 (m, 4H, Ar), 7.56 (d, 2H,
6.3.41. N-(1H-Indol-5-yl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (48)
J ¼ 8.6 Hz, Ar), 7.28 (brs, 1H, NH), 6.92 (d, 2H, J ¼ 8.6 Hz, Ar),
3.89e3.83 (m, 2H, CH2), 3.44e3.39 (m, 2H, CH2); 13C NMR (DMSO-
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
d6)
d 158.4, 144.6, 138.0, 137.8, 130.8, 127.7, 121.8, 116.3, 87.8, 44.2,
MeOH 95:5). Yield: 82%; Reddish solid; mp: 239e241 ꢀC; IR: 3372,
36.4; HRMS (ESþ) m/z found 443.9131; C15H14IN3O3S (Mþ þ H)
3252, 1960 cmꢂ1 1H NMR (DMSO-d6)
; d 11.04 (brs, 1H, NH), 9.68
requires 443.9879.
(brs, 1H, NH), 7.67e7.59 (m, 4H, Ar), 7.31e7.30 (m, 1H, Ar),
7.25e7.22 (m, 3H, Ar), 6.86e6.83 (m, 1H, Ar), 6.34 (brs, 1H, NH),
3.86e3.81 (m, 2H, CH2), 3.43e3.38 (m, 2H, CH2); 13C NMR (DMSO-
6.3.36. N-(4-Cyanomethylphenyl)-4-(2-oxoimidazolidin-1-yl)
benzenesulfonamide (43)
d6)
d 158.4, 144.1, 133.6, 131.6, 129.3, 127.7, 127.7, 126.2, 117.0, 116.0,
Flash chromatography (CH2Cl2 to CH2Cl2/AcOEt 0:1 to AcOEt/
113.6, 111.5, 101.1, 44.2, 36.4; HRMS (ESþ) m/z found 357.0972;
MeOH 95:5). Yield: 59%; Orange solid; mp: 204e206 ꢀC; IR: 3298,
C17H16N4O3S (Mþ þ H) requires 357.1021.

S. Fortin et al. / European Journal of Medicinal Chemistry 46 (2011) 5327e5342
5341
6.4. Preparation of compounds 49e51
equation. Those descriptors involved in optimizing the QSAR
analysis are molecular weight (MW), molecular volume (V), molar
refractivity (MR), polar volume (PV), polar surface area (PSA),
energy, and the similarity data of each compound to the hypothesis.
In CoMFA studies, Tripos standard fields were used as CoMFA field
classes and an Sp3 carbon atom with a unit positive charge was
used as the probe to evaluate steric and electrostatic potentials at
every lattice point. The resolution of the grid was 2 Å. Distance
method was used to control the form of the Coulombic electrostatic
energy calculation. A 30 kcal/mol cutoff was used for steric and
electrostatic field values. In addition to the CoMFA fields, all
descriptors used in optimizing the CoMSIA models were also used
in optimizing CoMFA models. The thirty seven compounds used in
the CoMSIA study were also used in CoMFA study as well. Initially
all PIB-SA derivatives were selected to generate the CoMSIA model.
Biological activities for this set of compounds is small, falling within
the range of 10ꢂ6 to 10ꢂ7 M, and this led to challenges to obtain
a perfect model of high R2 value for LOO prediction. Different ways
to generate the alignment were tried including: the generation of
hypothesis model by using two of the most actives compounds (20
and 27), four of the most actives compounds (20, 22, 27, and 40),
and six of the most actives compounds (15, 20, 22, 27, 40, and 48,
Fig. 2A). The hypothesis generated from the alignment on six active
compounds is pursued further for the generation of the CoMSIA
model. The best R2 values of LOO (leave-one-out) generated from it
were 0.446 for IC50 in HT-29 cell line, 0.552 for IC50 in M21 cell line,
and 0.465 for IC50 in MCF-7 cell line. From the predicting result, we
removed four outliers (absolute predict value greater than 1.0):
compound 18, 27, 35 and 46, and the models were rebuilt.
6.4.1. 1-(2-Chloroethyl)-3-phenylurea (49)
2-Chloroethylisocyanate (1.2 eq.) was added dropwise to a cold
solution (ice bath) of aniline (1.0 eq.) in dry methylene chloride
(15 mL per g of aniline). The ice bath was removed and the reaction
mixture was stirred at room temperature for 24 h. After completion
of the reaction, the solvent was evaporated under reduced pressure
to give a white solid, which was triturated twice with cold hexanes/
ether 10:1. Yield: 99%; mp: 108e110 ꢀC; IR
NMR (DMSO-d6): 8.69 (s, 1H, NH), 7.44e7.41 (m, 2H, Ar),
n
: 3304, 1637 cmꢂ1; 1H
d
7.27e7.22 (m, 2H, Ar), 6.95e6.90 (m, 1H, Ar), 6.45 (t, 1H, J ¼ 5.1 Hz,
NH), 3.68 (t, 2H, J ¼ 6.1 Hz, CH2), 3.48e3.42 (m, 2H, CH2); 13C NMR
(CDCl3 and few drops of CD3OD):
44.0, 41.7.
d 156.5, 138.9, 128.8, 122.7, 119.5,
6.4.2. 1-Phenylimidazolidin-2-one (50)
Sodium hydride (3 eq.) was slowly added to a cold solution of
compound 49 (1 eq.) in tetrahydrofuran under dry nitrogen
atmosphere. The ice bath was removed after 30 min and the
reaction mixture was stirred at room temperature for 5 h. The
reaction was quenched at 0 ꢀC with water and diluted with ethyl
acetate. The organic layer was washed with water and brine, dried
over sodium sulfate, filtered, and concentrated in vacuo to provide
50, which were used without further purification to afford white
solids. Yield: 98%; Compound 50 was also synthesized using
method described by Neville [40]. Briefly, triphosgene (12.2 mmol)
was dissolved in 40 mL of tetrahydrofuran and cooled at 0 ꢀC. To the
resulting solution was added (36.7 mmol) of N-phenylethylenedi-
amine dissolved in 65 mL of tetrahydrofuran and 7.7 mL of trie-
thylamine over a period of 30 min. A white solid immediately
precipitated and the reaction was complete after 5 min. The reac-
tion mixture was quenched with water and diluted with ethyl
acetate. The organic layer was washed with water and brine, dried
over sodium sulfate, filtered, and concentrated in vacuo. The
residue was purified by flash chromatography (CH2Cl2 to CH2Cl2/
AcOEt 3:10) to afford a white solid. Yield: 80%; mp: 154e156 ꢀC; IR
Acknowledgment
This work was supported by the Canadian Institutes of Health
Research (R.C.-G; Grant #MOP-79334 and #MOP-89707). S. Fortin
is recipient of a studentship from the Canadian Institutes of Health
Research (CGD-83623). We also acknowledge the technical exper-
tise of Dr. Michel Déry for HPLC-MS experiments.
n
: 3240, 1680 cmꢂ1 1H NMR (DMSO-d6):
; d 7.58e7.55 (m, 2H, Ar),
7.34e7.29 (m, 2H, Ar), 7.02e6.95 (m, 2H, Ar and NH), 3.88e3.83 (m,
Appendix. Supplementary material
2H, CH2), 3.44e3.39 (m, 2H, CH2); 13C NMR (CDCl3):
d 160.2, 140.2,
128.8, 122.7, 117.9, 45.3, 37.5.
Supplementary data related to this article can be found online at
doi:10.1016/j.ejmech.2011.08.034.
6.4.3. 4-(2-Oxoimidazolidin-1-yl)benzene-1-sulfonyl chloride (51)
To 1.5 mL (23.1 mmol) of chlorosulfonic acid in 3 mL of carbon
tetrachloride at 0 ꢀC was added slowly (3.1 mmol) compound 50.
The reaction was almost completed after 2 h at 0 ꢀC. The reaction
mixture was poured slowly onto ice water, filtered to collect the
solid. The white solid was dried under vacuum. Yield: 56%; mp:
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257e259 ꢀC; IR
n
:
3232, 1711 cmꢂ1
;
1H NMR (DMSO-d6):
d
7.57e7.51 (m, 4H, Ar), 3.88e3.82 (m, 2H, CH2), 3.44e3.38 (m, 2H,
CH2); 13C NMR (DMSO-d6):
36.5.
d 158.9, 141.2, 140.5, 126.1, 115.8, 44.5,
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