Synthesis and structure-activity relationships of carbazole sulfonamides as a novel class of antimitotic agents against solid tumors

Article abstract of DOI:10.1021/jm060546h

Two series of carbazole sulfonamides related to Combretastatin A4 (1) were synthesized and evaluated for antiproliferative activity. Thirteen of the 26 new sulfonamides exhibited IC₅₀ values of < 1 μM against CEM leukemia cells. Five compounds

Full text of DOI:10.1021/jm060546h

J. Med. Chem. 2006, 49, 6273-6282
6273
Synthesis and Structure-Activity Relationships of Carbazole Sulfonamides as a Novel Class of
Antimitotic Agents Against Solid Tumors
Laixing Hu,†,‡ Zhuo-rong Li,‡ Yan Li,§ Jinrong Qu,§ Yi-He Ling,| Jian-dong Jiang,*,‡ and David W. Boykin*,†
Department of Chemistry, Georgia State UniVersity, Atlanta, Georgia 30303-3083, Institute of Medicinal Biotechnology, Chinese Academy of
Medical Sciences and Peking Union Medical College, Beijing 100050, People’s Republic of China, Department of Medicine, Mount Sinai
School of Medicine, New York, New York 10029, and Department of Medicine, Albert Einstein College of Medicine, Bronx, New York 10164
ReceiVed May 10, 2006
Two series of carbazole sulfonamides related to Combretastatin A4 (1) were synthesized and evaluated for
antiproliferative activity. Thirteen of the 26 new sulfonamides exhibited IC50 values of <1 µM against
CEM leukemia cells. Five compounds were evaluated against a panel of eight human tumor cell lines.
9-Ethyl-N-(3,4,5-trimethoxyphenyl)-carbazole-3-sulfonamide (11a) showed significant antitumor activity in
two human xenograft models (MCF-7 and Bel-7402). Preliminary studies with 11a showed that the mode
of action involves arrest of M-phase cell cycle and induction of apoptosis by increasing expression of p53
and promoting bcl-2 phosphorylation. Unexpectedly, 11a only weakly inhibits tubulin polymerization, which
suggests that the mode of action of 11a differs from 1 and involves an unidentified target(s). Also, the SAR
information gleaned from ring A-substituted analogues varies significantly from that of 1. Carbazole
sulfonamides are a novel promising class of antimitotic agents with clinical development potential.
Introduction
Antimitotic agents arrest the cell cycle at the G2/M phase
and lead to apoptotic cell death and tumor regression.1,2 Classic
antimitotic agents, many of which are tubulin-binding agents,
such as taxanes and vinca alkaloids, interfere with microtubule
dynamics by targeting tubulin and are widely used to treat
human cancers. However, neurotoxicity, difficult synthesis, and
development of multidrug resistance, mainly through the expres-
sion of P-glycoprotein (Pgp),a have limited their clinical use.3
Therefore, it is important to develop new small molecule tubulin-
binding agents or other antimitotic agents with novel modes of
action.4-6
Combretastatin A4 (1), a novel small molecule tubulin-
binding agent which was isolated from the South African bush
willow Combretum caffrum by Pettit et al.,7 strongly inhibits
the polymerization of tubulin by binding to the colchicine
binding site (Figure 1).8 Potent cytotoxicity for 1 has been
observed against a wide variety of cell lines, including multidrug
resistant lines. However, the low water solubility of this
compound has limited its in vivo efficacy. Its water-soluble
phosphate prodrug 2 is now in phase ΙΙ clinical trials.9 It appears
to act by a unique irreversible and selective antivascular effect
that disrupts immature endothelial cells’ cytoskeleton in vivo.10
As a result of the relatively simple chemical structure and
unique biological profiles, 1 has stimulated a lot of research in
the last two decades (e.g., Figure 1). Structure-activity relation-
ship studies for 1 have shown that the 3,4,5-trimethoxy
Figure 1. Lead compound and antimitotic agents.
* Corresponding authors. Tel.: 404-651-3798 (D.W.B.); 86-10-63165290
(J.-d.J.). Fax: 404-651-1416 (D.W.B.); 86-10-63017302 (J.-d.J.). E-mail:
substituents of the A-ring are essential for good biological
dboykin@gsu.edu (D.W.B.); jiandong.jiang@mssm.edu (J.-d.J.).
activities.11,12 Therefore, many studies have focused on the
† Georgia State University.
‡ Chinese Academy of Medical Sciences and Peking Union Medical
design of analogues by altering the linking group and the B-ring
of 1 to provide better biological activities.13,14 We also were
interested in exploring the alteration of the linker and the B-ring.
In this report, we study the influence of replacing the olefin
linker with sulfonamide groups and testing the replacement of
the B-ring unit with a carbazole. Furthermore, we examine the
influence of various substitutions on the A-ring in this new class
of compounds.
College.
§ Mount Sinai School of Medicine.
| Albert Eistein College of Medicine.
a Abbreviations: Pgp, P-glycoprotein; ip, intraperitoneal; p53, tumor
suppressor protein; P-p53, phosphorylated p53; cdc2, cell division cycle 2
kinase; bcl-2, B-cell leukemia/lymphoma 2 protein; bcl-2-p, monophos-
phorylated bcl-2; bcl-2-2p, biphosphorylated bcl-2; PBS, phosphate buffered
saline; PMSF, phenylmethanesulfonyl fluoride; mAb, monoclonal antibody;
sc, subcutaneously.
10.1021/jm060546h CCC: $33.50 © 2006 American Chemical Society
Published on Web 09/14/2006

6274 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21
Hu et al.
Scheme 1
Scheme 2
In general, sulfonamides constitute a useful class of drugs,
displaying a variety of activities including antibacterial, anti-
carbonic anhydrase, diuretic, hypoglycemic, and antithyroid
The syntheses of N-(9-ethylcarbazole-3-yl) benzene sulfon-
amides 29-33a are shown in Scheme 2. 3-Amino-9-ethyl
carbazole was allowed to react with various commercially
available sulfonyl chlorides using the TEA/DMF procedure to
yield the sulfonamides 29-33a. Compound 34a was prepared
from 33a using the same method employed for 24a.
effectiveness.15 Since the discovery of 3 (E-7010) in 1992,16
a
number of sulfonamides have been reported to be potent
inhibitors of tubulin polymerization and have antiproliferative
properties.15b In fact, 3 and 4 (T-138067)17 have entered phase
ΙΙ clinical trials against solid tumors. Novel benzene 518 and
indole 619 sulfonamides related to 1 have been claimed by
Tularik and Abbott. However, reports of the activity data and
SAR information for these compounds, especially in vivo
efficacy, is limited. For a related structural class of compounds,
7, Aventis has shown the utility of substituted carbazoles as
tubulin polymerization inhibitors (Figure 1).20
Results and Discussion
1. In Vitro Antiproliferative Activity and SAR. As shown
in Table 1, antiproliferative activities of the two series of
carbazole sulfonamide compounds were obtained using CEM
leukemia cells. CEM leukemia cells were used for initial
compound screening because of their known rapid proliferation
and high sensitivity to standard anticancer agents. Many
compounds exhibited strong activities against human leukemia
cells with IC50 values of e1000 to >70 nM (11c, 15a, 20a,
30-31a) or e70 nM (11a,b, 16-17a, 18b, 21a, 22a,b).
The lead 3,4,5-trimethoxyphenyl-substituted carbazole sul-
fonamide 11a has strong cytotoxic activity against human
leukemia cells, as reflected by the IC50 value of 56 nM. This
antiproliferate activity showed that the carbazole tricycle can
be used for the B-ring of these sulfonamide compounds related
to 1.23 The cytotoxicity of the N-9 methyl-substituted compound
11b is similar to that of N-9 ethyl compound 11a. However,
11c, which has only a hydrogen on N-9, showed 10-fold less
activity than 11a and 11b. Furthermore, when carbazole was
replaced by dibenzofuran to yield 11d, essentially all activity
was lost. The results demonstrated that the nitrogen atom of
carbazole and the alkylation of N-9 are very important for potent
activity.
In this study, we present the synthesis, in vitro evaluations,
SAR analysis, in vivo efficacy, and preliminary mode of action
information for the new carbazole sulfonamide compounds
based on our conceptional lead compound 8.
Chemistry
The syntheses of carbazole sulfonamides 11a-c, 12a-17a,
18b, 19a-24a, and 22b are shown in Scheme 1. The preparation
of these carbazole sulfonamides was readily achieved by
allowing carbazole sulfonyl chlorides and various commercially
available substituted anilines to react in the presence of
triethylamine (TEA) in dimethylformamide (DMF) solution at
room temperature. The carbazole sulfonyl chlorides 10a-c were
prepared from commercially available carbazoles 9a-c using
the method of Mitsumori et al.21 Compound 24a was obtained
by reduction of nitro compound 23a in the presence of SnCl2
in refluxing ethanol.22 3,5-Bis-(trifluoromethyl) aniline failed
to react with N-9-ethylcarbazole-3-sulfonyl chloride 10a using
the TEA/DMF procedure. However, 25a was obtained by using
pyridine both as catalyst and as solvent. The dibenzofuran
sulfonamide 11d was prepared using the TEA/DMF procedure
from dibenzofuran 9d.
Next, we evaluated the effect of location of the methoxy
substitution on the A-ring on cytotoxic activity in the carbazole
sulfonamide series. Table 1 shows that the 4-methoxy-
substituted compound 12a is inactive; 3,4-dimethoxy, 3,4-
(methylenedioxy), and 3,5-dimethoxy-substituted compounds
13a-15a are more potent than 12a, however, much less potent

Synthesis of Carbazole Sulfonamides
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21 6275
Table 1. Antiproliferative Activity of Carbazole Sulfonamides in CEM
Table 2. Antiproliferative Activities of 11a,b, 16a, 17a, and 22a in
Leukemia Cells
Human Tumor Cell Lines
IC50a (nM)
human
tumor
podophyllo-
toxin
cell line
CEM
11a 11b 16a 17a 22a
1
T-cell
leukemia
T-cell
56
20
46 57
19 48
61 56 1.9
49 67 9.3
7.2
14
Molt-3
leukemia
Bel-7402 hepatoma
MCF-7b breast
cancer
DU-145 prostate
cancer
201 116 96
89 70 48
219 135 10
98 67 12
9.1
15
a
IC50
compd
X
R1
Et
Y
R
(nM)
11a
11b
11c
11d
12a
13a
14a
15a
16a
17a
18b
19a
20a
21a
22a
22b
24a
25a
26a
28ab
29a
30a
31a
32a
33a
34a
N
N
N
O
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
SO2NH
3,4,5-OMe3
3,4,5-OMe3
3,4,5-OMe3
3,4,5-OMe3
4-OMe
3,4-OMe2
3,4-OCH2O-
3,5-OMe2
2,4-OMe2
2,5-OMe2
2,4,6-OMe3
4-F
56
46
606
603 232 144 365 169 9.3
201 116 144 780 90 2.8
52
10
Me SO2NH
H
SO2NH
SO2NH
SO2NH
SO2NH
SO2NH
SO2NH
SO2NH
SO2NH
PC-3
prostate
cancer
>10 000
>10 000
2436
6252
974
Et
Et
Et
Et
Et
Et
DND-1
melanoma
89
70 120 195 90 2.5
12
23
DMS-79 lung
cancer
1899 2088 NTc NT NT 124
a IC50 ) compound concentration required to inhibit human tumor cells
proliferation by 50% after 72 h treatment. b Tularik’s compound (5; Figure
1) IC50 value reported as 150 nM (ref 18). c NT ) not tested.
57
61
70
2171
482
Me SO2NH
Et
Et
Et
Et
SO2NH
SO2NH
SO2NH
SO2NH
3-Cl-4-OMe
These results suggest that the mode of action of these sulfon-
amides is different from 1 and analogues.
5-Cl-2,4-OMe2 67
4-Cl-2,5-OMe2 56
4-Cl-2,5-OMe2 46
On comparison of the data for the compound 11a with a
-SO2NH- linkage to that of 26a with a -SO2NHCH2-
linkage, it is clear that extension of molecular length by an extra
methylene group results in significant loss of activity. Therefore,
further modifications with longer linkers were not pursued.
Also, we synthesized N-(9-ethylcarbazole-3-yl) benzene sul-
fonamides 28-34a to test the effect of reversing the orientation
of the sulfonamide linkage relative to the A- and B-rings. From
the IC50 of these compounds, some differences between series
are noted. The activity of 4-methoxy and 3,4-dimethoxy-
substituted compounds 28a and 29a are similar to that of 12a
and 13a from the original series. However, the 2,5-dimethoxy-
substituted compound 30a in the reversed series is 10-fold less-
potent than 17a in the first series. Interestingly, within the
reversed series, the cytotoxicity of the 3-methoxy compound
31a is comparable to 30a and more than 3 times greater than
that of the 3,4-methoxy compound 29a. Replacement of the
3-methoxy group of 31a with the electron-withdrawing 3-cyano
group to yield 32a results in some loss of activity. The
introduction of either a nitro or an amino group at the 2-position
of compound 28a to yield 33a and 34a did not affect the activity.
2. In Vitro Cancericidal Activity of 11a, 11b, 16a, 17a,
and 22a against Several Cancer Cell Lines. The lead carbazole
sulfonamide 11a has potent activity against CEM leukemia cells;
consequently, we tested the antiproliferative activity of 11a
against several other human cancer cell lines in vitro. Table 2
contains these results along with comparative data for podo-
phyllotoxin and 1. The IC50 values of 11a were between 20 and
1899 nM against the cell lines studied (Table 2). The least-
sensitive cell line was lung cancer DMS-79 (IC50 ) 1899 nM).
Leukemia cells showed the highest sensitivities (IC50 ) 20 and
56 nM). The sensitivities of MCF-7 breast cancer cells,
melanoma, Bel-7402 hepatoma, and PC-3 prostate cancer cells
were also high, with IC50 values of less than 201 nM.
Me SO2NH
Et
Et
Et
Et
Et
Et
Et
Et
Et
Et
SO2NH
SO2NH
SO2NHCH2 3,4,5-OMe3
NHSO2
NHSO2
NHSO2
NHSO2
NHSO2
NH SO2
NHSO2
3-NH2-4-OMe
3,5-(CF3) 2
2200
>10 000
>10 000
>10 000
2651
4-OMe
3,4-OMe2
2,5-OMe2
3-OMe
603
789
1319
3-CN
2-NO2-4-OMe
2-NH2-4-OMe
>10 000
>10 000
7.2
podophyllotoxin
1
1.9
a IC50 ) compound concentration required to inhibit CEM leukemia cell
proliferation by 50% after 72 h treatment. b Commercially available.
than 11a. Also, the activity of 15a is greater than that of 13a
and 14a. From these results, in contrast to 1, we deduce that
methoxy substitution and location on the A-ring plays an
important role in affecting cytotoxicity. The effect of substitution
involving only 3,4,5-methoxy groups on the sulfonamides is
similar to that found for 1. However, an unexpected difference
in SAR information with 1 is noted with other substitutions.
2,4-Dimethoxy, 2,5-dimethoxy, 2,4,6-trimethoxy, 5-chloro-2,4-
dimethoxy, and 4-chloro-2,5-dimethoxy-substituted compounds
16a, 17a, 18b, 21a, 22a,b have similar potent activities as the
3,4,5-trimethoxy-substituted compounds 11a,b and a more than
15-fold stronger activity than the 3,4- or 3,5-dimethoxy-
substituted compounds 13a and 15a. In contrast, it is well-known
that 3,4,5-trimethoxy substituents of the A-ring of 1 are required
for potent biological activity.11,12
Replacement of the 4-methoxy group in ring A of 12a by a
fluorine atom to yield 19a enhanced the cytotoxicity. Replace-
ment of one methoxy group of 13a with a chlorine atom to
yield 20a also enhanced cytotoxicity. The non-3,4,5-trimethoxy
analogues 16a, 17a, 21a, and 22a all exhibit similar potent IC50
values. The 3-amino-4-methoxy-substituted analogue 24a is as
active as the 3,4-dimethoxy-substituted compound 13a, dem-
onstrating that an amino group can replace a methoxy group at
the 3-position in this system. Replacement of the 3,5-dimethoxy
groups by the strong withdrawing group CF3 resulted in loss of
activity; compare results for 15a and 25a. Thus, in contrast to
1, we note that a number of replacements of the ring A 3,4,5-
methoxy groups are tolerated and some enhance the activity.
With the goal of obtaining more SAR information, we also
examined the antiproliferative activity of compounds 11b, 16a,
17a, and 22a compared to the lead compound 11a against cancer
cell lines. From these results (Table 2), it can be seen that the
activity of N- methyl carbazole compound 11b is quite similar
to that of 11a. Compounds 16a, 17a, and 22a have similar
activities against leukemia, Bel-7402 hepatocarcinoma, MCF-7
breast cancer cells, and melanoma cell lines. However, for PC-3

6276 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21
Hu et al.
carbazole sulfonamide series. In general, the carbazole analogues
are not as active in vitro as the two reference compounds in the
cell lines evaluated. However, the activity of 11a approaches
that of the reference compounds in Molt-3 T-cell leukemia and
in MCF-7 breast cancer cell lines.
3. Inhibition of Human Breast Cancer and Hepatocarci-
noma by 11a in Nude Mice. Because lead compound 11a has
potent antiproliferative activity in human cancer cell lines in
vitro, we evaluated the in vivo anticancer efficacy of 11a in
human tumor xenograft models. As shown in Figure 2A, for
MCF-7 breast cancer, the average tumor volume of the control
group was at 3000 mm3 on day 35 post-treatment. On treatment
with 11a at 100 mg/kg (ip), the tumor volume was reduced by
75% on the same time course. Compound 11a was somewhat
more effective than adriamycin in this model, which showed a
67% reduction in tumor volume. For human hepatocarcinoma,
11a given at 60 mg/kg (ip) reduced the tumor volume by 55%
(Figure 2B). Overt toxicity to the treated mice at these doses
was not noted. The LD50 of 11a was over 500 mg/kg (ip), and
additional studies at higher dosage are merited.
Despite the fact that 1 shows strong cytotoxicity in vitro, it
shows poor antitumor activities in vivo models because of low
water solubility. From the in vivo efficacy of 11a, it can be
deduced that this compound has favorable bioavailability;
presumably at least in part due to the presence of the sulfon-
amide linker.
4. Mode of Action of 11a. Lead compound 11a has shown
potent in vitro and in vivo activities, and the initiation of mode
of action studies seemed warranted. Flow cytometric analysis
with MCF-7 cells (Figure 3) showed a blockade at the M-phase
by 11a within 24 h of treatment. More than 50% of the cells at
the 24 h time point showed dispersed chromosomes character-
istic of mitotic arrest (Figure 4) when the concentrations of 11a
ranged between 0.22 and 1.1 µM. Western blot analysis showed
remarkable increases of cyclin B1 and cdc2, verifying the
accumulation of cells in M-phase after their exposure to 11a
(Figure 5). Leukemia cells treated with 11a exhibited a similar
cell cycle profile and morphological features (data not shown).
Also, 11a induced clear DNA fragmentation and apoptosis in
MCF-7 tumor cells (data not shown). Consequently, factors that
are related to the apoptotic process were examined. First, p53
levels were assessed because this protein is well-known for its
tumor suppressor function. Increased expression of p53 and
P-p53 in breast cancer cells treated with 11a (Figure 5) was
observed.3,24 Bcl-2 expression was also examined because it is
an inhibitor of apoptosis. As shown in Figure 5, bcl-2 protein
was inactivated by phosphorylation in cells treated with 11a
Figure 2. Therapeutic efficacies of 11a in human breast cancer (MCF-
7) and hepatocarcinoma (BEL-7402) in nude mice. In the MCF-7 tumor
(A), 11a was given 100 mg/kg, ip, q2d, with adriamycin (1 mg/kg,ip,
q2d) as control. In the BEL-7402 tumor (B), 11a was given 60 mg/kg,
ip, q2d. Each point represents the mean of the tumor volume (n ) 5).
prostate cancer cells, the 2,5-dimethoxy-substituted compound
17a is less potent than the other compounds. For the other
prostate cancer cell line DU-145, 16a and 22a show somewhat
higher activity than the other compounds studied. Interestingly,
introduction of a 4-chloro atom enhances the antiprostate cancer
activity, comparing the results for 17a and 22a. Generally, the
analogues 16a and 22a showed somewhat enhanced activity
compared to the lead compound 11a. These SAR results further
demonstrate that 3,4,5- trimethoxy substituents of the A-ring
are not indispensable for potent biological activities in this
Figure 3. Cell cycle block at the M-phase in MCF-7 cells treated with 11a. MCF-7 cells treated with 11a (0.22 µM) were analyzed for cell cycle
distribution using flow cytometry. The major block at M-phase appeared between 12 and 24 h post-treatment.

Synthesis of Carbazole Sulfonamides
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21 6277
Figure 4. Metaphase arrest by 11a in MCF-7 Cells. Giemsa staining (400×).
Figure 5. Increased expression of p53, cyclin B1, and cdc2 in MCF-7
cells treated with 11a. MCF-7 cells were treated at different concentra-
tions (0.022 to 2.2 µM) or at 0.22 µM from 0 to 48 h. Increased p53,
P-p53, cyclin B1, and cdc2 expression were noted.
Figure 7. Effect of 11a on tubulin assembly. Free purified â-tubulin
in reaction buffer were incubated with GTP and Mg2+ for assembly in
the absence or presence of 11a (223 µM) or paclitaxel (23 µM). Tubulin
assembly was determined every 1.5 min by O.D. at 340 nm. Each point
represents the mean of two independent experiments.
protein (KSP) inhibitors, which alter microtubule stability, are
being developed as a novel potential antimitotic agents for the
treatment of cancer.5 Further studies of the molecular mechanism
of action of 11a is underway.
Figure 6. Compound 11a induced bcl-2 phosphorylation (inactivation)
in MCF-7 cells. MCF-7 cells were treated at different concentrations
(0.022 to 2.2 µM) or at 0.22 µM from 0 to 48 h. Phosphorylated bcl-2
was noted.
Conclusions
(Figure 6). Increased expression of p53 and bcl-2 phosphoryl-
ation processes were dose and time-dependent (Figures 5 and
6). These results strongly suggest that this carbazole sulfonamide
has potent cytotoxicity because it can arrest the tumor cell cycle
at the M-phase and induce apoptotic cell death by increasing
expression of p53 and promoting bcl-2 phosphorylation.
Next, we assessed whether 11a could directly inhibit micro-
tubule polymerization using purified tubulin in a cell-free
system. Surprisingly, lead compound 11a only weakly affected
tubulin polymerization, even at very high concentration (223
µM; Figure 7). This result differs from that of 1 and close
analogues, which strongly inhibit tubulin polymerization when
their antiproliferate activity is in the submicromolar range. This
result suggests the arrest of the cell cycle at the M-phase and
the induction of apoptosis by 11a is unlikely to involve direct
action on tubulin. As previously noted, SAR results with these
sulfonamides also showed that they act differently from 1 and
close analogues. As a result of the size of the carbazole tricycle
and different geometry of the sulfonamide linkage,25 compared
to an alkene, it is possible that the sulfonamides are not
geometrically suitable for direct tubulin binding and some other
cellular targets are likely involved in cell cycle arrest. There
are many microtubule-associated proteins (MAPs) that are
possible targets, including the dynein and kinesin motor proteins,
as well as microtubule regulatory proteins, which can also
interact with a microtubule.1,4 For example, kinesin spindle
We have synthesized two series of carbazole sulfonamide
compounds structurally related to 1. New compounds 11a,b,
16a, 17a, 18b, 21a, and 22a,b exhibit strong activities against
human leukemia cells. The SAR information revealed that
alkylation of N-9 is necessary for potent cytotoxicity. Di-
benzofuran replacement of carbazole led to loss of activity.
3,4,5-Trimethoxyphenyl substituents on ring A yield potent
activities, however, this array is not essential. 2,4-Dimethoxy,
2,5-dimethoxy, 2,4,6-trimethoxy, 5-chloro-2,4-dimethoxy, and
4-chloro-2,5-dimethoxy ring A-substituted compounds also have
similar potent activities. Preliminary mode of action studies
demonstrated that the lead compound 11a arrests tumor cell
cycle at M-phase and induces apoptotic cell death by increasing
expression of p53 and promoting bcl-2 phosphorylation. How-
ever, 11a only weakly inhibits tubulin polymerization. Lead
compound 11a showed potent antitumor efficacy in vivo models.
Lead compounds 11a, 16a, and 22a merit further studies as
novel promising antimitotic agents against solid tumors.
Experimental Section
Chemistry. Melting points were determined with a Mel-Temp
1
3.0 capillary melting point apparatus and are uncorrected. H and
13C NMR spectra were recorded on a Varian GX400 or Varian
Unity Plus 300 spectrometer. Elemental analyses were obtained

6278 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21
Hu et al.
from Atlantic Microlab, Inc. (Norcross, GA), and are within (0.4
of the theoretical values.
6.74 (m, 2H), 7.27 (dd, J ) 7.2, 7.5 Hz, 1H), 7.53 (dd, J ) 7.8,
7.5 Hz, 1H), 7.68 (d, J ) 8.1 Hz, 1H), 7.74 (d, J ) 8.7 Hz, 1H),
7.78 (d, J ) 8.7 Hz, 1H), 8.24 (d, J ) 7.8 Hz, 1H), 8.56 (s, 1H),
9.82 (s, 1H). 13C NMR (DMSO-d6): δ 148.7, 145.7, 141.2,
140.3, 131.1, 129.4, 126.9, 124.1, 121.7, 121.4, 120.8, 120.2, 120.0,
113.0, 112.0, 109.8, 109.4, 106.1, 55.5, 55.3, 37.3, 13.7. Anal.
(C22H22N2O4S‚0.3H2O) C, H, N.
9-Ethyl-N-(3,4,5-trimethoxyphenyl)-carbazole-3-sulfon-
amide (11a). To a solution of 3,4,5-trimethoxyaniline (360 mg,
1.97 mmol) in 10 mL DMF at room temperature, 9-ethylcarbazole-
3-sulfonyl chloride21 (600 mg, 2.04 mmol) was added. After stirring
for 5 min, TEA (0.30 mL, 2.13 mmol) was added, and the mixture
was stirred for an additional 2 h. After adding ice water, the
precipitate was filtered, washed with water, dried, and purified by
flash chromatography over silica gel, affording the title compound
(500 mg, 56%) as a white solid; mp 201-203 °C. 1H NMR
(DMSO-d6): δ 1.30 (t, J ) 7.2 Hz, 3H), 3.48 (s, 3H), 3.61 (s, 6H),
4.56 (q, J ) 7.2 Hz, 2H), 6.42 (s, 2H), 7.28 (dd, J ) 7.2, 7.5 Hz,
1H), 7.53 (dd, J ) 7.2, 8.1 Hz, 1H), 7.69 (d, J ) 8.4 Hz, 1H), 7.77
(d, J ) 8.7 Hz, 1H), 7.84 (d, J ) 8.7 Hz, 1H), 8.28 (d, J ) 8.4 Hz,
1H), 8.65 (s, 1H), 10.02 (s, 1H). 13C NMR (DMSO-d6): δ 152.9,
141.3, 140.3, 134.2, 133.8, 129.3, 127.0, 124.2, 121.7, 121.5, 120.8,
120.5, 120.0, 109.8, 109.6, 97.4, 60.0, 55.7, 37.3, 13.6. Anal.
(C23H24N2O5S‚0.4H2O) C, H, N.
9-Ethyl-N-[3,4-(methylenedioxy)phenyl]-carbazole-3-sulfon-
amide (14a). The title compound was obtained from 3,4-(methyl-
enedioxy)aniline and 9-ethylcarbazole-3-sulfonyl chloride using a
procedure similar to compound 11a. Yield: 74%; pale brown solid,
1
mp 182-184 °C. H NMR (DMSO-d6): δ 1.30 (t, J ) 6.9 Hz,
3H), 4.46 (q, J ) 6.9 Hz, 2H), 5.89 (s, 2H), 6.50 (d, J ) 8.4 Hz,
1H), 6.68-6.71 (m, 2H), 7.27 (dd, J ) 7.5, 7.2 Hz, 1H), 7.53 (dd,
J ) 7.8, 7.5 Hz, 1H), 7.68 (d, J ) 8.4 Hz, 1H), 7.72-7.79 (m,
2H), 8.25 (d, J ) 7.8 Hz, 1H), 8.55 (s, 1H), 9.89 (s, 1H). 13C NMR
(DMSO-d6): δ 147.3, 144.1, 141.2, 140.3, 132.0, 129.2, 126.9,
124.0, 121.7, 121.5, 120.9, 120.1, 120.0, 114.2, 109.8, 109.5, 108.2,
103.1, 101.2, 37.3, 13.7. Anal. (C21H18N2O4S‚0.3H2O) C, H, N.
9-Methyl-N-(3,4,5-trimethoxyphenyl)-carbazole-3-sulfon-
amide (11b). The title compound was obtained from 3,4,5-
trimethoxyaniline and 9-methylcarbazole-3-sulfonyl chloride using
a procedure similar to compound 11a.Yield: 65%; white solid, mp
N-(3,5-Dimethoxyphenyl)-9-ethylcarbazole-3-sulfonamide (15a).
The title compound was obtained from 3,5-dimethoxyaniline and
9-ethylcarbazole-3-sulfonyl chloride using a procedure similar to
compound 11a. Yield: 63%; pale brown solid, mp 171-173 °C.
1H NMR (DMSO-d6): δ 1.30 (t, J ) 6.9 Hz, 3H), 3.60 (s, 6H),
4.46 (q, J ) 6.9 Hz, 2H), 6.07 (d, J ) 2.1 Hz, 1H), 6.32 (d, J )
2.1 Hz, 2H), 7.28 (dd, J ) 7.5, 7.2 Hz, 1H), 7.54 (dd, J ) 7.8, 7.5
Hz, 1H), 7.68 (d, J ) 8.1 Hz, 1H), 7.77 (d, J ) 8.7 Hz, 1H), 7.85
(d, J ) 8.7 Hz, 1H), 8.28 (d, J ) 7.5 Hz, 1H), 8.66 (s, 1H), 10.20
(s, 1H). 13C NMR (DMSO-d6): δ 160.6, 141.2, 140.2, 140.0, 129.3,
126.9, 124.0, 121.6, 121.4, 120.8, 120.3, 120.0, 109.7, 109.6, 97.4,
94.8, 55.0, 37.3, 13.6. Anal. (C22H22N2O4S) C, H, N.
1
206-208 °C. H NMR (DMSO-d6): δ 3.48 (s, 3H), 3.61 (s, 6H),
3.88 (s, 3H), 6.44 (s, 2H), 7.28 (dd, J ) 7.2, 7.8 Hz, 1H), 7.54 (dd,
J ) 8.1, 7.2 Hz, 1H), 7.64 (d, J ) 8.1 Hz, 1H), 7.73 (d, J ) 8.7
Hz, 1H), 7.85 (d, J ) 8.7 Hz, 1H), 8.27 (d, J ) 7.5 Hz, 1H), 8.65
(s, 1H), 10.02 (s,1H). 13C NMR (DMSO-d6): δ 152.9, 142.3, 141.4,
134.2, 133.7, 129.1, 126.9, 124.1, 121.5, 121.3, 120.7, 120.3, 120.0,
109.9, 109.7, 97.3, 60.0, 55.7, 29.3. Anal. (C22H22N2O5S‚0.25H2O)
C, H, N.
N-(3,4,5-Trimethoxyphenyl)-9H-carbazole-3-sulfonamide (11c).
The title compound was obtained from 3,4,5-trimethoxyaniline and
9H-carbazole-3-sulfonyl chloride using a procedure similar to
compound 11a. Yield: 58%; white solid, mp 205-206 °C. 1H NMR
(DMSO-d6): δ 3.47 (s, 3H), 3.60 (s, 6H), 6.41 (s, 2H), 7.22 (dd,
J ) 7.2, 7.8 Hz, 1H), 7.45 (dd, J ) 8.1, 7.2 Hz, 1H), 7.53 (d, J )
8.4 Hz, 1H), 7.59 (d, J ) 8.7 Hz, 1H), 7.77 (d, J ) 7.8 Hz, 1H),
8.22 (d, J ) 7.5 Hz, 1H), 8.61 (s, 1H), 9.99 (s, 1H), 11.79 (s, 1H).
13C NMR (DMSO-d6): δ 152.9, 141.6, 140.5, 134.2, 133.8, 129.1,
126.8, 124.1, 121.9, 121.7, 120.6, 120.4, 119.8, 111.6, 111.3, 97.4,
60.0, 55.7. Anal. (C21H20N2O5S) C, H, N. High-resolution mass
calcd for C21H21N2O5S, 413.1171; found, 413.1182.
N-(3,4,5-Trimethoxyphenyl)-dibenzofuran-2-sulfonamide (11d).
The title compound was obtained from 3,4,5-trimethoxyaniline and
dibenzofuran-2-sulfonyl chloride21 using a procedure similar to
compound 11a. Yield: 84%; white solid, mp 168-170 °C. 1H NMR
(DMSO-d6): δ 3.50 (s, 3H), 3.62 (s, 6H), 6.41 (s, 2H), 7.46 (dd,
J ) 6.9, 7.5 Hz, 1H), 7.60 (dd, J ) 7.5, 7.2 Hz, 1H), 7.76 (d, J )
8.1 Hz, 1H), 7.85-7.96 (m, 2H), 8.31 (d, J ) 8.1 Hz, 1H), 8.66 (s,
1H), 10.16 (s, 1H). 13C NMR (DMSO- d6): δ 157.2, 156.2, 153.0,
134.4, 134.1, 133.6, 128.9, 126.3, 123.9, 122.5, 121.9, 121.1, 112.6,
112.0, 98.0, 60.0, 55.7. Anal. (C21H19NO6S) C, H, N.
N-(2,4-Dimethoxyphenyl)-9-ethylcarbazole-3-sulfonamide (16a).
The title compound was obtained from 2,4-dimethoxyaniline and
9-ethylcarbazole-3-sulfonyl chloride using a procedure similar to
1
compound 11a. Yield: 75%; brown solid, mp 185-187 °C. H
NMR (DMSO-d6): δ 1.30 (t, J ) 7.2 Hz, 3H), 3.43 (s, 3H), 3.60
(s, 3H), 4.78 (q, J ) 7.2 Hz, 2H), 6.56 (dd, J ) 9.0, 2.7 Hz, 1H),
6.76 (d, J ) 9.0 Hz, 1H), 6.86 (d, J ) 2.7 Hz, 1H), 7.27 (dd, J )
7.5, 7.2 Hz, 1H), 7.53 (dd, J ) 8.4, 7.2 Hz, 1H), 7.68 (d, J ) 8.4
Hz, 1H), 7.74 (d, J ) 8.7 Hz, 1H), 7.84 (dd, J ) 8.7, 1.5 Hz, 1H),
8.24 (d, J ) 8.1 Hz, 1H), 8.60 (d, J ) 1.5 Hz, 1H), 9.32 (s, 1H).
13C NMR (DMSO-d6): δ 153.4, 145.9, 141.7, 140.8, 130.7, 127.4,
127.3, 124.7, 122.3, 121.8, 121.3, 120.5, 120.4, 113.1, 110.3, 110.0,
109.9, 109.7, 56.6, 55.8, 37.8, 14.1. Anal. (C22H22N2O4S‚0.4H2O)
C, H, N.
N-(2,5-Dimethoxyphenyl)-9-ethylcarbazole-3-sulfonamide (17a).
The title compound was obtained from 2,5-dimethoxyaniline and
9-ethylcarbazole-3-sulfonyl chloride using a procedure similar to
1
compound 11a. Yield: 70%, brown solid, mp 164-166 °C. H
NMR(DMSO-d6): δ 1.30 (t, J ) 6.9 Hz, 3H), 3.33 (s, 3H), 3.65
(s, 3H), 4.48 (q, J ) 6.9 Hz, 2H), 6.34 (d, J ) 2.1 Hz, 1H), 6.41
(dd, J ) 9.0, 2.1 Hz, 1H), 7.10 (d, J ) 9.0 Hz, 1H), 7.26 (dd, J )
7.5, 7.5 Hz, 1H), 7.52 (dd, J ) 7.8, 7.2 Hz, 1H), 7.68 (d, J ) 7.8
Hz, 1H), 7.68-7.72 (m, 2H), 8.21 (d, J ) 7.8 Hz, 1H), 8.45 (s,
1H), 9.07 (s, 1H). 13C NMR (DMSO-d6): δ 158.5, 154.2, 141.1,
140.2, 130.7, 127.6, 126.7, 124.4, 121.8, 121.2, 120.7, 119.9, 118.3,
109.7, 108.9, 104.5, 98.9, 55.2, 37.3, 13.6. Anal. (C22H22N2O4S)
C, H, N.
9-Ethyl-N-(4-methoxyphenyl)-carbazole-3-sulfonamide (12a).
The title compound was obtained from 4-methoxyaniline and
9-ethylcarbazole-3-sulfonyl chloride using a procedure similar to
compound 11a. Yield: 71%; brown solid, mp 213-215 °C (lit.26
1
mp 215-217 °C). H NMR (DMSO-d6): δ 1.30 (t, J ) 6.9 Hz,
3H), 3.60 (s, 3H), 4.46 (q, J ) 6.9 Hz, 2H), 6.74 (d, J ) 8.1 Hz,
1H), 7.00 (d, J ) 8.1 Hz, 1H), 7.26 (dd, J ) 7.2, 7.2 Hz, 1H), 7.52
(dd, J ) 7.8, 7.5 Hz, 1H), 7.67 (d, J ) 8.1 Hz, 1H), 7.70-7.78
(m, 2H), 8.22 (d, J ) 7.8 Hz, 1H), 8.52 (s, 1H), 9.81 (s, 1H). 13C
NMR (DMSO-d6): δ 156.2, 141.3, 140.3, 130.7, 129.5, 126.9,
124.0, 123.0, 121.7, 121.4, 120.8, 120.0, 119.9, 114.2, 109.8, 109.4,
55.0, 37.3, 13.7.
9-Methyl-N-(2,4,6-trimethoxyphenyl)-carbazole-3-sulfon-
amide (18b). The title compound was obtained from 2,4,6-
trimethoxyaniline and 9-methylcarbazole-3-sulfonyl chloride using
a procedure similar to compound 11a. Yield: 70%; white solid,
1
mp 209-211°C. H NMR (DMSO-d6): δ 3.29 (s, 6H), 3.71 (s,
3H), 3.93 (s, 3H), 6.09 (s, 2H), 7.26 (dd, J ) 7.2, 7.5 Hz, 1H),
7.53 (dd, J ) 7.5, 7.8 Hz, 1H), 7.66 (d, J ) 8.4 Hz, 1H), 7.70 (d,
J ) 8.7 Hz, 1H), 7.78 (dd, J ) 8.7, 1.8 Hz, 1H), 8.22 (d, J ) 7.5
Hz,1H), 8.44 (d, J ) 1.8 Hz, 1H), 8.52 (s, 1H). 13C NMR (DMSO-
d6): δ 160.3, 158.5, 142.6, 141.9, 133.1, 127.4, 125.2, 122.3, 121.5,
121.0, 120.5, 120.1, 110.3, 109.2, 106.7, 91.4, 55.9, 55.9, 29.8.
Anal. (C22H22N2O5S) C, H, N.
N-(3,4-Dimethoxyphenyl)-9-ethylcarbazole-3-sulfonamide (13a).
The title compound was obtained from 3,4-dimethoxyaniline and
9-ethylcarbazole-3-sulfonyl chloride using a procedure similar to
compound 11a. Yield: 76%; pale brown solid, mp 175-177 °C.
1H NMR (DMSO-d6): δ 1.30 (t, J ) 7.2 Hz, 3H), 3.59 (s, 6H),
4.46 (q, J ) 7.2 Hz, 2H), 6.56 (dd, J ) 8.7, 2.1 Hz, 1H), 6.71-

Synthesis of Carbazole Sulfonamides
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21 6279
9-Ethyl-N-(4-fluorophenyl)-carbazole-3-sulfonamide (19a).
The title compound was obtained from 4-fluoroaniline and 9-ethyl-
carbazole-3-sulfonyl chloride using a procedure similar to com-
using 10% aqueous K2CO3, the pH of the solution was adjusted to
7-8. The precipitate was filtered and washed with water. The
residue was dissolved in 100 mL acetone and filtered. The filtrate
was evaporated under reduced pressure, and the precipitate was
collected. The precipitate was dissolved in 10 mL anhydrous ethyl
acetate and saturated at ice-water bath temperature with anhydrous
HCl gas. The solution was then stirred at room temperature for 3
h. The precipitate was filtered to afford the title compound (110
1
pound 11a. Yield: 95%; pale brown solid, mp 158-160 °C. H
NMR (DMSO-d6): δ 1.24 (t, J ) 6.6 Hz, 3H), 4.46 (q, J ) 6.6
Hz, 2H), 7.00-7.06 (m, 2H), 7.16-7.26 (m, 2H), 7.24 (d, J ) 7.5
Hz, 1H), 7.48 (dd, J ) 8.1, 7.2 Hz, 1H), 7.57 (d, J ) 7.5 Hz, 1H),
7.69 (d, J ) 8.1 Hz, 1H), 7.83 (d, J ) 8.7 Hz, 1H), 8.22 (d, J )
7.5 Hz, 1H), 8.62 (s, 1H), 10.20 (s, 1H). 13C NMR (DMSO-d6): δ
158.9 (d, J ) 856.7 Hz), 141.3, 140.3, 134.5 (d, J ) 11.4 Hz),
129.2, 127.0, 124.0, 122.4 (d, J ) 32.1 Hz), 121.7, 121.6, 120.9,
120.2, 120.0, 115.8 (d, J ) 91.8 Hz), 109.7, 109.5, 37.3, 13.6. Anal.
(C20H17FN2O2S) C, H, N.
1
mg, 66%) as a brown solid; mp 223-225 °C. H NMR (DMSO-
d6): δ 1.30 (t, J ) 7.2 Hz, 3H), 3.52 (s, 3H), 3.71 (s, 3H), 4.46 (q,
J ) 7.2 Hz, 2H), 6.86 (dd, J ) 9.0, 2.1 Hz, 1H), 6.92 (d, J ) 9.0
Hz, 1H), 7.13 (d, J ) 2.1 Hz, 1H), 7.28 (dd, J ) 7.5,7.5 Hz, 1H),
7.53 (dd, J ) 7.8, 7.5 Hz, 1H), 7.68 (d, J ) 8.1 Hz, 1H), 7.74 (d,
J ) 9.0 Hz, 1H), 7.79 (dd, J ) 8.7, 1.5 Hz, 1H), 8.24 (d, J ) 7.8
Hz, 1H), 8.59 (d, J ) 1.5 Hz, 1H), 10.06 (s, 1H). 13C NMR (DMSO-
d6): δ 147.3, 141.2, 140.2, 131.1, 129.3, 126.9, 125.3, 124.0, 121.7,
121.4, 120.9, 120.1, 120.0, 117.8, 114.1, 112.2, 109.8, 109.5, 55.9,
37.3, 13.7. Anal. (C21H22ClN3O3S‚1.25H2O) C, H, N.
N-(3-Chloro-4-methoxyphenyl)-9-ethylcarbazole-3-sulfon-
amide (20a). The title compound was obtained from 3-chloro-4-
methoxyaniline and 9-ethylcarbazole-3-sulfonyl chloride using a
procedure similar to compound 11a. Yield: 72%; brown solid, mp
1
222-224 °C. H NMR (DMSO-d6): δ 1.33 (t, J ) 7.2 Hz, 3H),
3.72 (s, 3H), 4.49 (q, J ) 7.2 Hz, 2H), 6.99 (d, J ) 9.0 Hz, 1H),
7.05 (dd, J ) 8.7, 2.4 Hz, 1H), 7.16 (d, J ) 2.4 Hz, 1H), 7.30 (dd,
J ) 7.5, 7.5 Hz, 1H), 7.56 (dd, J ) 7.2, 7.2 Hz, 1H), 7.71 (d, J )
8.4 Hz, 1H), 7.78-7.79 (m, 2H), 8.28 (d, J ) 8.1 Hz, 1H), 8.59 (s,
1H), 10.07 (s, 1H). Anal. (C21H19ClN2O3S) C, H, N.
N-[3,5-Di-(trifluoromethyl)phenyl]-9-ethylcarbazole-3-sul-
fonamide (25a). To a solution of 3,5-bis-(trifluoromethyl)aniline
(0.31 mL, 2.0 mmol) in 5 mL pyridine at room temperature,
9-ethylcarbazole-3-sulfonyl chloride21 (600 mg, 2.05 mmol) was
added. After stirring for 1.5 h, the pyridine was removed under
reduced pressure. The residue was purified by flash chromatography
over silica gel to afford the title compound (852 mg, 88%) as a
N-(5-Chloro-2,4-dimethoxyphenyl)-9-ethylcarbazole-3-sul-
fonamide (21a). The title compound was obtained from 5-chloro-
2,4-dimethoxyaniline and 9-ethylcarbazole-3-sulfonyl chloride using
a procedure similar to compound 11a. Yield: 76%; brown solid,
1
white solid; mp 152-153 °C. H NMR (DMSO-d6): δ 1.32 (t, J
) 7.2 Hz, 3H), 4.49 (q, J ) 7.2 Hz, 2H), 7.32 (dd, J ) 7.2, 7.8
Hz, 1H), 7.57 (dd, J ) 7.8, 7.2 Hz, 1H), 7.70-7.74 (m, 4H), 7.84
(d, J ) 8.7 Hz, 1H), 7.88 (d, J ) 8.7 Hz, 1H), 8.32 (d, J ) 7.5 Hz,
1H), 8.74 (s, 1H), 11.14 (s,1H). 13C NMR (DMSO-d6): δ 142.0,
140.9, 140.8, 131.6 (q, J ) 132.0 Hz), 128.6, 127.5, 124.2, 123.3
(q, J ) 1084.0 Hz), 122.2, 122.1, 121.3, 120.8, 120.6, 118.7, 116.6,
110.4, 110.3, 37.8, 14.0. Anal. (C22H16F6N2O2S) C, H, N.
1
mp 194-196 °C. H NMR (DMSO-d6): δ 1.30 (t, J ) 6.9 Hz,
3H), 3.34 (s, 3H), 3.75 (s, 3H), 4.48 (q, J ) 6.9 Hz, 2H), 6.57 (s,
1H), 7.19 (s, 1H), 7.27 (dd, J ) 7.5, 7.2 Hz, 1H), 7.52 (dd, J )
7.2, 8.1 Hz, 1H), 7.68 (d, J ) 8.1 Hz, 1H), 7.70-7.75 (m, 2H),
8.23 (d, J ) 7.5 Hz, 1H), 8.48 (s, 1H), 9.33 (s, 1H). 13C NMR
(DMSO-d6): δ 153.4, 153.2, 141.2, 140.3, 130.3, 127.4, 126.8,
124.4, 121.8, 121.3, 120.8, 119.9, 118.5, 111.1, 109.8, 109.1, 98.0,
56.3, 55.9, 37.3, 13.7. Anal. (C22H21ClN2O4S) C, H, N.
9-Ethyl-N-(3,4,5-trimethoxybenzyl)-carbazole-3-sulfon-
amide (26a). The title compound was obtained from 3,4,5-
trimethoxybenzylamine and 9-methylcarbazole-3-sulfonyl chloride
using a procedure similar to compound 11a. Yield: 81%; white
N-(4-Chloro-2,5-dimethoxyphenyl)-9-ethylcarbazole-3-sul-
fonamide (22a). The title compound was obtained from 4-chloro-
2,5-dimethoxyaniline and 9-ethylcarbazole-3-sulfonyl chloride using
a procedure similar to compound 11a. Yield: 56%; brown solid,
1
solid, mp 187-189 °C. H NMR (DMSO-d6): δ 1.30 (t, J ) 6.9
Hz, 3H), 3.33 (s, 3H), 3.57 (s, 6H), 3.96 (d, J ) 6.3 Hz, 2H), 4.47
(q, J ) 6.9 Hz, 2H), 6.43 (s, 2H), 7.27 (dd, J ) 7.2, 7.8 Hz, 1H),
7.52 (t, J ) 7.5, 7.2 Hz, 1H), 7.67 (d, J ) 8.1 Hz, 1H), 7.74 (d, J
) 8.7 Hz, 1H), 7.83 (dd, J ) 8.7, 2.1 Hz, 1H), 7.95 (t, J ) 6.3 Hz,
1H), 8.24 (d, J ) 8.1 Hz, 1H), 8.54 (d, J ) 1.5 Hz, 1H). 13C NMR
(DMSO-d6): δ 152.5, 141.0, 140.3, 136.1, 133.1, 130.9, 126.8,
124.0, 121.9, 121.5, 120.8, 119.9, 119.8, 109.6, 109.3, 104.7, 59.5,
55.5, 46.6, 37.2, 13.6. Anal. (C24H26N2O5S) C, H, N.
1
mp 188-190 °C. H NMR (DMSO-d6): δ 1.30 (t, J ) 7.2 Hz,
3H), 3.39 (s, 3H), 3.70 (s, 3H), 4.47 (q, J ) 7.5 Hz, 1H), 6.95 (s,
1H), 7.06 (s, 1H), 7.27 (dd, J ) 7.8, 7.2 Hz, 1H), 7.53 (dd, J )
8.1, 7.5 Hz, 1H), 7.69 (d, J ) 8.1 Hz, 1H), 7.75 (d, J ) 8.7 Hz,
1H), 7.82 (dd, J ) 8.7, 1.5 Hz, 1H), 8.25 (d, J ) 7.8 Hz, 1H), 8.60
(d, J ) 1.5 Hz, 1H), 9.49 (s, 1H). Anal. (C22H21ClN2O4S) C, H, N.
N-(4-Chloro-2,5-dimethoxyphenyl)-9-methylcarbazole-3-sul-
fonamide (22b). The title compound was obtained from 4-chloro-
2,5-dimethoxyaniline and 9-methylcarbazole-3-sulfonyl chloride
using a procedure similar to compound 11a. Yield: 67%; pale white
solid, mp 218-220 °C. 1H NMR (DMSO-d6): δ 3.41 (s, 3H), 3.70
(s, 3H), 3.90 (s, 3H), 6.95 (s, 1H), 7.07 (s, 1H), 7.28 (dd, J ) 7.2,
7.8 Hz, 1H), 7.54 (dd, J ) 8.4, 7.2 Hz, 1H), 7.66 (d, J ) 8.4 Hz,
1H), 7.71 (d, J ) 8.7 Hz, 1H), 7.83 (d, J ) 8.7 Hz, 1H), 8.25 (d,
J ) 7.8 Hz, 1H), 8.62 (s, 1H), 9.48 (s, 1H). 13C NMR (DMSO-d6):
δ 148.1, 145.7, 142.3, 141.4, 129.8, 126.8, 125.6, 124.3, 121.5,
121.2, 120.6, 120.1, 120.0, 117.0, 113.9, 109.8, 109.3, 108.7, 56.4,
56.3, 29.3. Anal. (C21H19ClN2O4S) C, H, N.
3,4-Dimethoxy-N-(9-ethylcarbazole-3-yl)-benzene sulfonamide
(29a). The title compound was obtained from 3,4-dimethoxy-
benzenesulfonyl chloride and 3-amino-9-ethylcarbazole using a
procedure similar to compound 11a. Yield: 77%; pale brown solid,
1
mp 168-169 °C. H NMR (DMSO-d6): δ 1.24 (t, J ) 6.6 Hz,
3H), 3.68 (s, 3H), 3.73 (s, 3H), 4.35 (q, J ) 6.6 Hz, 2H), 6.98 (d,
J ) 8.7 Hz, 1H), 7.11-7.23 (m, 4H), 7.43 (dd, J ) 7.8, 7.2 Hz,
1H), 7.46 (d, J ) 8.1 Hz, 1H), 7.55 (d, J ) 8.1 Hz, 1H), 7.83 (s,
1H), 8.03 (d, J ) 7.5 Hz, 1H), 9.85 (s, 1H). 13C NMR (DMSO-
d6): δ 151.9, 148.4, 139.9, 137.1, 131.1, 129.1, 126.0, 122.1, 121.7,
121.5, 120.6, 120.3, 118.7, 114.5, 110.9, 109.5, 109.3, 109.2, 55.6,
36.9, 13.6. Anal. (C22H22N2O4S‚0.25H2O) C, H, N.
N-(3-Amino-4-methoxyphenyl)-9-ethylcarbazole-3-sulfon-
amide hydrochloride (24a). The nitro compound (23a) was
obtained from 4-methoxy-3-nitroaniline27 and 9-ethylcarbazole-3-
sulfonyl chloride using a procedure similar to compound 11a and
used directly in the next step. Yield: 71%; yellow solid, mp 218-
220 °C. 1H NMR (DMSO-d6): δ 1.29 (t, J ) 6.9 Hz, 3H), 3.77 (s,
3H), 4.46 (q, J ) 6.9 Hz, 2H), 7.21 (d, J ) 8.7 Hz, 1H), 7.28 (dd,
J ) 7.2, 7.8 Hz, 1H), 7.36 (d, J ) 9.0 Hz, 1H), 7.53 (dd, J ) 7.2,
7.8 Hz, 1H), 7.60 (s, 1H), 7.68 (d, J ) 8.4 Hz, 1H), 7.52-7.83 (m,
2H), 8.26 (d, J ) 7.8 Hz, 1H), 8.60 (s, 1H), 10.31 (s,1H).
To a suspension of the above nitro compound 23a (180 mg, 0.42
mmol) in 4 mL anhydrous ethanol, SnCl2‚2H2O (520 mg, 2.31
mmol) was added, and the mixture was heated at reflux for 2 h.
After cooling to room temperature, 20 mL of water was added and,
2,5-Dimethoxy-N-(9-ethylcarbazole-3-yl)-benzene sulfonamide
(30a). The title compound was obtained from 2,5-dimethoxy-
benzenesulfonyl chloride and 3-amino-9-ethylcarbazole using a
procedure similar to compound 11a. Yield: 84%; off-white solid,
1
mp 237-239 °C. H NMR (DMSO-d6): δ 1.25 (t, J ) 6.6 Hz,
3H), 3.62 (s, 3H), 3.91 (s, 3H), 4.35 (q, J ) 6.6 Hz, 2H), 7.07-
7.17 (m, 5H), 7.39-7.45 (m, 2H), 7.54 (d, J ) 8.1 Hz, 1H), 7.80
(s, 1H), 8.01 (d, J ) 7.8 Hz, 1H), 9.71 (s, 1H). 13C NMR (DMSO-
d6): δ 152.0, 150.3, 139.9, 137.0, 128.8, 127.1, 125.9, 122.0, 121.6,
121.1, 120.1, 119.5, 118.7, 115.1, 113.9, 113.9, 109.2, 109.1, 56.3,
55.6, 36.9, 13.6. Anal. (C22H22N2O4S‚0.2H2O) C, H, N.
N-(9-Ethylcarbazole-3-yl)-3-methoxybenzenesulfonamide (31a).
The title compound was obtained from 3-methoxybenzenesulfonyl

6280 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21
Hu et al.
chloride and 3-amino-9-ethylcarbazole using a procedure similar
for 10 min. The slides were air-dried, fixed in methanol for 5 min,
stained with Giemsa (Harleco, NJ) at room temperature for 15 min,
and preserved with resin. Morphological features were observed
by light microscopy (BH-2 OLYMPUS, Japan). Cells in M-phase
arrest were identified if spreading of the chromosomes was seen
in the cytoplasm.
Cell Cycle Analysis. Tumor cells were treated with 11a for
indicated periods of time and harvested after a digestion with a
mixture of 0.25% trypsin/0.02% EDTA. The cells (1.5 × 106) were
washed twice with PBS, followed by the treatment with RNase
(Sigma, St. Louis, MO) at a final concentration of 0.5 µg/µL at 37
°C for 30 min. Cells were then fixed in 4 mL of ice-cold 70%
ethanol at 4 °C for at least 12 h. After washing in PBS, cells were
stained with propidium iodide at a final concentration of 0.1 µg/
µL at 4 °C for 60 min. At least 104 cell events per sample were
analyzed in an EPICS flow cytometer (Beckman Coulter, San Jose,
CA) using EPICS software (version 2.0).
to compound 11a. Yield: 71%; pale brown solid, mp 176-178
1
°C. H NMR (DMSO-d6): δ 1.25 (t, J ) 7.2 Hz, 3H), 3.70 (s,
3H), 4.35 (q, J ) 7.2 Hz, 2H), 7.10-7.18 (m, 3H), 7.27-7.29 (m,
2H), 7.37-7.48 (m, 3H), 7.55 (d, J ) 8.1 Hz, 1H), 7.82 (d, J )
1.5 Hz, 1H), 8.03 (d, J ) 7.5 Hz, 1H), 10.02 (s, 1H). 13C NMR
(DMSO-d6): δ 159.2, 140.8, 139.9, 137.2, 130.3, 128.7, 126.1,
122.1, 121.6, 121.5, 120.3, 118.9, 118.8, 118.4, 114.6, 111.7, 109.4,
109.2, 55.5, 37.0, 13.7. Anal. (C21H20N2O3S) C, H, N.
3-Cyano-N-(9-ethylcarbazole-3-yl)-benzenesulfonamide (32a).
The title compound was obtained from 3-cyanobenzenesulfonyl
chloride and 3-amino-9-ethylcarbazole using a procedure similar
1
to compound 11a. Yield: 71%; white solid, mp 198-200 °C. H
NMR (DMSO-d6): δ 1.25 (t, J ) 7.2 Hz, 3H), 4.36 (q, J ) 7.2
Hz, 2H), 7.06 (dd, J ) 8.7, 1.8 Hz, 1H), 7.16 (dd, J ) 7.2, 7.5 Hz,
1H), 7.44 (dd, J ) 7.5, 7.8 Hz, 1H), 7.48 (d, J ) 8.7 Hz, 1H), 7.56
(d, J ) 8.1 Hz, 1H), 7.71 (dd, J ) 8.1, 7.8 Hz, 1H), 7.82 (d, J )
1.8 Hz, 1H), 7.94 (d, J ) 8.1 Hz, 1H), 8.05-8.09 (m, 3H), 10.20
(s, 1H). 13C NMR (DMSO-d6): δ 140.8, 140.0, 137.5, 136.3, 131.2,
130.6, 130.2, 127.9, 126.2, 122.3, 121.9, 121.6, 120.4, 118.8, 117.4,
115.4, 112.3, 109.5, 109.3, 37.0, 13.6. Anal. (C21H17N3O2S‚0.2H2O)
C, H, N.
DNA Fragmentation. DNA from untreated or 11a treated cells
was extracted. Briefly, after washing with PBS, cells were lysed
with 500 µL of lysis buffer (0.05 M Tris-HCl, pH 7.5; 0.15 M
NaCl; 0.005 M EDTA, pH 8.0 and 1% sodium dodecyl sulfate)
containing 100 µg/mL proteinase K at 56 °C for 2 h. The resulting
products were extracted with phenol and then a mixture of
chloroform/isoamyl alcohol (24:1, v/v), precipitated at 4 °C
overnight with absolute ethanol, followed by centrifugation (12000
g, PM180R, ALC, Ltd., Italy) at 4 °C for 20 min. The ethanol was
removed, and the dry pellet of nucleic acid was resuspended in 10
mM Tris-HCl/1 mM EDTA solution. The DNA samples were then
treated with RNase (Sigma, St. Louis, MO) at a final concentration
of 0.1 µg/µL at 37 °C for 20 min. DNA content was determined
spectrophotometrically by measuring absorption at 260 nm (A260).
All samples had A260/A280 g 2. The DNA sample (5 ug) was
analyzed in 1.5% horizontal agarose gel prepared with 1% ethidium
bromide. Electrophoresis was performed at 75 V for 2 h, and the
DNA migration was visualized under UV light.
Microtubule Polymerization Assays. The HTS-tubulin polym-
erization assay kit (Cytoskeleton, Inc., Denver, CO) was used to
measure microtubule polymerization, following the protocol pro-
vided by the vendor. Study compounds were added into each well
of the 96-well plate placed on ice and using G-PEM buffer as
diluent (80 mM PIPES, pH 6.9, 1 mM EGTA, 5% glycerol, 1 mM
GTP). Each well contained G-PEM buffer, study compound at the
concentration indicated, and â-tubulin at a concentration of 1 mg/
mL. The plate was shaken for 20 s and warmed to 37 °C, and the
absorbance was read at 340 nm every 1.5 min for 30 min.
Western Blot Assay. Cells were harvested, aliquoted and then
lysed in lysis buffer (50 mM Tris-HCl pH 8.0, 150 mM NaCl,
0.02% NaN3, 1% NP-40, 1 mM PMSF, 1 µg/mL aprotinin) on ice
for 1 h, followed by centrifugation (12 000 g) at 4 °C for 20 min.
Supernatants were collected and the protein concentration was
determined by the Bradford method. The protein sample (90 µg/
20 µL/test) was subjected to electrophoresis in a 10% or 15% SDS-
polyacrylamide gel (10% gel for proteins with molecular mass >50
KD; 15% gel for proteins with molecular mass <50 KD) and then
transferred to a nitrocellulose membrane with an electrotransfer
system (Bio-Rad Laboratories, Munich, Germany). The membrane
was blocked with 5% nonfat milk in TBST buffer (20 mM Tris-
HCl, 137 mM NaCl, 0.05% Tween-20) at 4 °C overnight.
Expression of bcl-2, p53, cdc-2, and cyclin B1 were probed with
anti-bcl-2 monoclonal antibody (mAb, CALBIOCHEM, San Diego,
CA), anti-p53 mAb (Santa Cruz Biotechnology, Inc.), anti-cdc-2
mAb and anti-cyclin B1 mAb (Oncogene Research Products,
Cambridge, MA), respectively. After washing in TBST four times,
the membrane was reacted with goat anti-mouse horseradish
peroxidase (HRP). After washing in TBST and then in Tris-buffered
saline (twice for each buffer and 10 min for each wash), the signals
were detected by an enhanced chemiluminescence (ECL) system
(Santa Cruz Biotechnology, Inc.) and preserved by exposure to
X-film.
N-(9-Ethylcarbazole-3-yl)-4-methoxy-2-nitrobenzenesulfon-
amide (33a). The title compound was obtained from 4-methoxy-
2-nitrobenzenesulfonyl chloride and 3-amino-9-ethylcarbazole using
a procedure similar to compound 11a. Yield: 80%; yellow solid,
mp 79-81 °C. 1H NMR (DMSO-d6): δ 1.26 (t, J ) 6.9 Hz, 3H),
3.81 (s, 3H), 4.37 (q, J ) 6.9 Hz, 2H), 7.15-7.19 (m, 2H), 7.24
(dd, J ) 9.0, 2.4 Hz, 1H), 7.44 (dd, J ) 8.4, 7.2 Hz, 1H), 7.49-
7.52 (m, 2H), 7.57 (d, J ) 8.1 Hz, 1H), 7.79 (d, J ) 9.0 Hz, 1H),
7.86 (s, 1H), 8.05 (d, J ) 8.1 Hz, 1H), 10.25 (s, 1H). 13C NMR
(DMSO-d6): δ 162.7, 156.0, 149.5, 140.0, 137.4, 131.9, 127.8,
126.1, 122.7, 122.2, 121.7, 120.4, 118.8, 117.1, 114.8, 109.8, 109.5,
109.3, 56.6, 37.0, 13.7. Anal. (C21H19N3O5S‚0.2H2O) C, H, N.
2-Amino-N-(9-ethylcarbazole-3-yl)-4-methoxybenzenesulfon-
amide (34a). The title compound was obtained from the above
compound 33a using a procedure similar to compound 24a. Yield:
1
74%; pale brown foam. H NMR (DMSO-d6): δ 1.25 (t, J ) 6.9
Hz, 3H), 3.62 (s, 3H), 4.35 (q, J ) 6.9 Hz, 2H), 5.99 (s, 2H), 6.06
(d, J ) 9.0 Hz, 1H), 6.22 (s, 1H), 7.12 (d, J ) 9.0 Hz, 1H), 7.15
(dd, J ) 7.5, 7.2 Hz, 1H), 7.32 (d, J ) 9.0 Hz, 1H), 7.43 (dd, J )
8.1, 7.2 Hz, 1H), 7.45 (d, J ) 9.0 Hz, 1H), 7.55 (d, J ) 8.1 Hz,
1H), 7.78 (s, 1H), 8.00 (d, J ) 8.1 Hz, 1H), 9.81 (s, 1H). 13C NMR
(DMSO-d6): δ 163.3, 148.3, 139.9, 136.9, 131.6, 129.2, 126.0,
122.1, 121.7, 120.8, 120.2, 118.7, 113.4, 111.8, 109.3, 109.2, 102.7,
99.5, 55.0, 37.0, 13.7. Anal. (C21H21N3O3S) C, H, N.
Biological Methods
Tumor Cell Lines. The human cell lines CEM (leukemia),
MCF-7 (breast cancer), DU-145 (prostate cancer), PC-3 (prostate
cancer), and DMS-79 (lung cancer) were from the American Tissue
Culture Collection (ATCC, Rocksville, MD). Molt-3 (human
leukemia) and DND-1 (human melanoma) cell lines were from Dr.
T. Ohnuma (Mount Sinai School of Medicine, New York, NY).
Bel-7402 cell line originated from Chinese human hepatocarcinoma
patients were provided by the Cancer Institute of Chinese Academy
of Medical Sciences (Beijing, China). Cells were cultivated in
RPMI-1640 supplemented with 10% heat inactivated fetal calf
serum. The media were treated with penicillin (150 ug/mL) and
streptomycin (150 ug/mL). Cells (cultured in 5% CO2 at 37 °C) in
exponential growth were used in all experiments.
Cancericidal Activity in Vitro. Cells were distributed into 24-
well plates (Falcon, Oxnard, CA) with 5 × 104 cells and a total
volume of 250 µL per well, followed by treatment with the study
compound at concentrations between 0 and 10 µM at 37 °C for 72
h. The IC50 values were determined in duplicates, and these values
were defined as drug concentrations killing 50% cells in comparison
with untreated controls and calculated by nonlinear regression
analysis.
Morphology. Cell sample slides were made on a Cytospin
centrifuge (Shandon Southern Products, Ltd., England) at 700 rpm
Human Tumor Models and Treatment. Nude mice (female,
5-6 weeks of age and 15-20 g in weight) were supplied by

Synthesis of Carbazole Sulfonamides
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21 6281
(11) Cushman, M.; Nagarathnam, D.; Gopal, D.; Chakraborti, A. K.; Lin,
C. M.; Hamel, E. Synthesis and evaluation of stilbene and dihydro-
stilbene derivatives as potential anticancer agents that inhibit tubulin
polymerization. J. Med. Chem. 1991, 34, 2579-2588.
Experimental Animal Center, Chinese Academy of Medical Sci-
ences (Beijing, China) and used for human hepatocarcinoma and
breast cancer (Bel-7402 and MCF-7) xenografts.
For the human hepatocarcinoma, nude mice received subcutane-
ous injection of BEL-7402 (5 × 106/mouse) on their backs. For
the human breast cancer model, female nude mice received
subcutaneous injection of estrogen (0.25 mg/mouse) 24 h before
the tumor was implanted. MCF-7 (8 × 106/mouse) cells were then
implanted (sc) on the backs of the mice. When the average tumor
volume was in 90-120 mm3 range, the tumor bearing mice were
randomly divided into solvent control and treatment cages with five
mice per group, followed by 11a ip administration to the treatment
group. The dose of 11a was 100 mg/kg for MCF-7 tumor and 60
mg/kg for Bel-7402 tumor. The treatment was continued for 5-6
weeks q2d (every other day). The known drug control was
adriamycin (1 mg/kg, ip, q2d). The therapeutic response to 11a
treatment was determined by measuring tumor volume. Two
perpendicular tumor diameters, width and length, were obtained
with calipers, followed by calculations using the formula, tumor
(12) Cushman, M.; Nagarathnam, D.; Gopal, D.; He, H.-M.; Lin, C. M.;
Hamel, E. Synthesis and evaluation of analogues of (Z)-1-(4-
methoxyphenyl)-2-(3,4,5-trimethoxyphenyl) ethene as potential cy-
totoxic and antimitotic agents. J. Med. Chem. 1992, 35, 2293-2306.
(13) (a) Nam, N. H. Combretastatin A-4 analogues as antimitotic antitumor
agents. Curr. Med. Chem. 2003, 10, 1697-1722. (b) Hsieh, H. P.;
Liou, J. P.; Mahindroo, N. Pharmaceutical design of antimitotic agents
based on combretastatins. Curr. Pharm. Des. 2005, 11, 1655-1677.
(14) (a) Ohsumi, K.; Nakagawa, R.; Fukuda, Y.; Hatanaka, T.; Morinaga,
Y.; Nihei, Y.; Ohishi, K.; Suga, Y.; Akiyama, Y.; Tsuji, T. Novel
combretastatin analogues effective against murine solid tumors:
design and structure-activity relationships. J. Med. Chem. 1998, 41,
3022-3032. (b) Tron, G. C.; Pagliai, F.; Grosso, E. D.; Genazzani,
A. A.; Sorba, G. Synthesis and cytotoxic evaluation of combretafura-
zans. J. Med. Chem. 2005, 48, 3260-3268. (c) LeBlanc, R.; Dickson,
J.; Brown, T.; Stewart, M.; Pati, H. N.; VanDerveer, D.; Arman, H.;
Harris, J.; Pennington, W.; Holt, H. L.; Lee, M. Synthesis and
cytotoxicity of epoxide and pyrazole analogues of the combretastatins.
Bioorg. Med. Chem. 2005, 13, 6025-6034. (d) Perez-Melero, C.;
Maya, A. B. S.; Rey, B. D.; Pelaez, R.; Caballero, E.; Medarde M.
A new family of quinolone and quinoxaline analogues of combreta-
statins. Bioorg. Med. Chem. Lett. 2004, 14, 3771-3774. (e) Simoni,
D.; Romagnoli, R.; Baruchello, R.; Rondanin, R.; Rizzi, M.; Pavani,
M. G.; Alloatti, D.; Giannini, G.; Marcellini, M.; Riccioni, T.;
Castorina, M.; Guglielmi, M. B.; Bucci, F.; Carminati, P.; Pisano,
C. Novel combretastatin analogues endowed with antitumor activity.
J. Med. Chem. 2006, 49, 3143-3152.
28
volume ) length × width2 × 0.52.
Acknowledgment. We thank the National Natural Science
Foundation of the P. R. China (30500630) and the Technology
Development Program of the Georgia State University Center
of Biotechnology and Drug Design for support of this work.
Supporting Information Available: Element analysis data of
new carbazole sulfonamide compounds. This material is available
free of charge via the Internet at http://pubs.acs.org.
(15) (a) Scozzafava, A.; Owa, T.; Mastrolorenzo, A.; Supuran, C. T.
Anticancer and antiviral sulfonamides. Curr. Med. Chem. 2003, 10,
925-953. (b) Casini, A.; Scozzafava, A.; Mastrolorenzo, A.; Supuran,
C. T. Sulfonamides and sulfonylated derivatives as anticancer agents.
Curr. Cancer Drug Targets 2002, 2, 55-75.
References
(1) Jordan, M. A.; Wilson, L. Microtubules as a target for anticancer
drugs. Nat. ReV. Cancer 2004, 4, 253-265.
(2) Jiang, J. D.; Wang, Y.; Roboz, J.; Strauchen, J.; Holland, J. F.; Bekesi,
J. G. Inhibition of microtubule assembly in tumor cells by 3-bromo-
acetylamino benzoylurea, a new cancericidal compound. Cancer Res.
1998, 58, 2126-2133.
(3) Dumontet, C.; Sikic, B. I. Mechanisms of action of and resistance to
anti-tubulin agents: microtubule dynamics, drug transport and cell
death. J. Clin. Oncol. 1999, 17, 1061-1070.
(4) Wood, K. W.; Cornwell, W. D.; Jackson, J. R. Past and future of the
mitotic spindle as an oncology target. Curr. Opin. Pharmacol. 2001,
1, 370-7.
(5) (a) Sarli, V.; Giannis, A. Inhibitors of mitotic kinesins: next-
generation antimitotics. ChemMedChem 2006, 1, 293-298. (b) Tao,
W.; South, V. J.; Zhang, Y.; Davide, J. P.; Farrell, L.; Kohl, N. E.;
Sepp-Lorenzino, L.; Lobell, R. B. Induction of apoptosis by an
inhibitor of the mitotic kinesin KSP requires both activation of the
spindle assembly checkpoint and mitotic slippage. Cancer Cell 2005,
8, 49-59. (c) Duhl, D. M.; Renhowe, P. A. Inhibitors of kinesin
motor proteinssresearch and clinical progress. Curr. Opin. Drug
DiscoVery DeV. 2005, 8, 431-436.
(6) (a) Ling, Y. H.; Liebes, L.; Jiang, J. D.; Holland, J. F.; Elliott, P. J.;
Adams, J.; Muggia, F. M.; Perez-Soler, R. Mechanisms of proteasome
inhibitor PS-341-induced G2-M-phase arrest and apoptosis in human
non-small cell lung cancer cell lines. Clin. Cancer Res. 2003, 9,
1145-1154. (b) Ling, Y. H.; Liebes, L.; Ng, B.; Buckley, M.; Elliott,
P. J.; Adams, J.; Jiang, J. D.; Muggia, F. M.; Perez-Soler, R. PS-
341, a novel proteasome inhibitor, induces Bcl-2 phosphorylation
and cleavage in association with G2-M phase arrest and apoptosis.
Mol. Cancer Ther. 2002, 1, 841-849.
(7) (a) Pettit, G. R.; Cragg, G. M.; Singh, S. B. Antineoplastic agents,
122. Constituents of Combretum caffrum. J. Nat. Prod. 1987, 50,
386-391. (b) Pettit, G. R.; Singh, S. B.; Hamel, E.; Lin, C. M.;
Alberts, D. S.; Garcia-Kendall, D. Isolation and structure of the strong
cell growth and tubulin inhibitor combretastatin A-4. Experientia
1989, 45, 209-211.
(8) Young, S. L.; Chaplin, D. J. Combretastatin A4 phosphate: back-
ground and current clinical status. Expert Opin. InVest. Drugs 2004,
13, 1171-1182.
(16) (a) Yoshino, Y.; Ueda, N.; Niijima, J.; Sugumi, H.; Kotake, Y.;
Koyanagi, N.; Yoshimatsu, K.; Asada, M.; Watanabe, T.; Nagasu,
T.; Tsukahara, K.; Iijima, A.; Kitoh, K. Novel sulfonamides as
potential, systemically active antitumor agents. J. Med. Chem. 1992,
35, 2496-2497. (b) Yoshimatsu, K.; Yamaguchi, A.; Yoshino, H.;
Koyanagi, N.; Kitoh, K. Mechanism of action of E7010: inhibition
of mitosis by binding to the colchicine site of tubulin. Cancer Res.
1997, 57, 3208-3213. (c) Koyanagi, N.; Nagasu, T.; Fujita, F.;
Watanabe, T.; Tsukahara, K.; Funahashi, Y.; Fujita, M.; Taguchi,
T.; Yoshino, H.; Kitoh, K. In vivo tumor growth inhibition produced
by a novel sulfonamide, E7010, against rodent and human tumors.
Cancer Res. 1994, 54, 1702-1706. (d) Segretti, J. A.; Polakowski,
J. S.; Koch, K. A.; Marsh, K. C.; Bauch, J. L.; Rosenber, S. H.;
Sham, H. L.; Cox, B. F.; Reinhart, G. A. Tumor selective anti effects
of the novel antimitotic compound ABT-751: an in vivo rat regional
hemodynamic study. Cancer Chemother. Pharmacol. 2004, 54, 273-
281.
(17) Shan, B.; Medina, J. C.; Santha, E.; Frankmoelle, W. P.; Chou, T.-
C.; Learned, R. M.; Narbut, M. R.; Stott, D.; Wu, P.; Jaen, J. C.;
Rosen, T.; Timmermans, P. B. M. W. M.; Beckmann, H. Selective,
covalent modification of â-tubulin residue Cys-239 by T138067, an
antitumor agent with in vivo efficacy against multidrug-resistant
tumors. Proc. Natl. Acad. Sci. U.S.A. 1999, 96, 5686-5691.
(18) Medina, J. C. Preparation of benzenesulfonamides and benzamides
regulating low-density lipoprotein (LDL) receptor expression and of
inhibiting abnormal cell proliferation. WO 9936391, 1999.
(19) (a) Li, Q.; Sham, H.; Woods, K. W.; Steiner, B. A.; Gwaltney, S.
L., II; Barr, K. J.; Imade, H. M.; Rosenberg, S. Preparation of
sulfonamides as cell proliferation inhibitors. U.S. 6521658, 2003.
(b) Prinz, H. Recent advances in the field of tubulin polymerization
inhibitors. Expert ReV. Anticancer Ther. 2002, 2, 695-708.
(20) Caulfield, T.; Cherrier, M.; Combeau, C.; Mailliet, P. Substituted
carbazoles as tubulin polymerization inhibitors and their use for the
treatment of cancer. EP 1253141, 2002.
(21) Mitsumori, S.; Tsuri, T.; Honma, T.; Hiramatsu, Y.; Okada, T.;
Hashizume, H.; Inagaki, M.; Arimura, A.; Yasui, K.; Asanuma, F.;
Kishino, J.; Ohtani, M. Synthesis and biological activity of various
derivatives of a novel class of potent, selective, orally active
prostaglandin D2 receptor antagonists. 1. bicyclo[2.2.1]heptane
derivatives. J. Med. Chem. 2003, 46, 2436-2445.
(9) Dorr, R. T.; Dvorakova, K.; Snead, K.; Alberts, D. S.; Salmon, S.
E.; Pettit, G. R. Antitumor activity of combretastatin-A4-phosphate,
a natural product tubulin inhibitor. InVest. New Drugs 1996, 14, 131-
137.
(10) Tozer, G. M.; Kanthou, C.; Baguley, B. C. Disrupting tumor blood
vessels. Nat. ReV. Cancer 2005, 5, 423-435. (b) Tozer, G. M.;
Kanthou, C.; Parkins, C. S.; Hill, S. A. The Biology of the
combretastatins as tumour targeting agents. Int. J. Exp. Pathol. 2002,
83, 21-38.
(22) Bellamy, F. D.; Ou, K. Selective reduction of aromatic nitro
compounds with stannous chloride in nonacidic and nonaqueous
medium. Tetrahedron Lett. 1984, 25, 839-842.
(23) Tron, G. C.; Pirali, T.; Sorba, G.; Pagliai, F.; Busacca, S.; Genazzani,
A. A. Medicinal chemistry of combretastatin A4: Present and future
directions. J. Med. Chem. 2006, 49, 3033-3044.

6282 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 21
Hu et al.
(24) Wang, L. G.; Liu, X. M.; Kreis, W.; Budman, D. R. The effect of
antimicrotubule agents on signal transduction pathways of apopto-
sis: a review. Cancer Chemother. Pharmacol. 1999, 44, 355-361.
(25) Petrov, V.; Petrova, V.; Girichev, G. V.; Oberhammer, H.; Giricheva,
N. I.; Ivanov, S. Molecular structure and conformations of benzene-
sulfonamide: gas electron diffraction and quantum chemical calcula-
tions. J. Org. Chem. 2006, 71, 2952-2956.
(26) Proshechkina, T. I.; Shishkina, V. I.; Pushkareva, Z. V.; Radtsev, V.
S.; Popova, E. N. Peculiarities of composition and chemical conver-
sion of carbazole and its derivatives. XXVIII. The synthesis of
sulfamide compounds in the carbazole series. Khim.-Farm. Zh. 1968,
2, 24-27.
(27) Rubenstein, S. M.; Baichwal, V.; Beckmann, H.; Clark, D. L.;
Frankmoelle, W.; Roche, D.; Santha, E.; Schwender, S.; Thoolen,
M.; Ye, Q.; Jaen, J. C. Hydrophilic, prodrug analogues of T138067
are efficacious in controlling tumor growth in vivo and show a
decreased ability to cross the blood brain barrier. J. Med. Chem. 2001,
44, 3599-3605.
(28) Yagi, M. J.; Bekesi, J. G. Evaluation of p-F-Phe-m-bis-(2-chloroethyl)
amino-L-Phe-Met-ethoxy HCl against transplantable and spontane-
ous murine neoplasia. Cancer Chemother. Pharmacol. 1990, 26,
215-220.
JM060546H