3-Aryl-2-quinolone derivatives: Synthesis and characterization of in vitro and in vivo antitumor effects with emphasis on a new therapeutical target connected with cell migration

Article abstract of DOI:10.1021/jm010978m

Among 25 3-aryl-2-quinolone derivatives synthesized, the antitumor activity of some of them was characterized both in vitro and in vivo. In this series, no compound appeared to be cytotoxic in vitro, as was known by the colorimetric MTT assay carried out

Full text of DOI:10.1021/jm010978m

J . Med. Chem. 2002, 45, 2543-2555
2543
3-Ar yl-2-Qu in olon e Der iva tives: Syn th esis a n d Ch a r a cter iza tion of In Vitr o a n d
In Vivo An titu m or Effects w ith Em p h a sis on a New Th er a p eu tica l Ta r get
Con n ected w ith Cell Migr a tion
Benoˆıt J oseph,*,† Francis Darro,‡ Aure´lie Be´hard,† Brigitte Lesur,‡ Franc¸oise Collignon,‡ Christine Decaestecker,§
Armand Frydman,‡ Ge´rald Guillaumet,† and Robert Kiss§
Institut de Chimie Organique et Analytique UMR-CNRS 6005, Universite´ d’Orle´ans, BP 6759, 45067 Orle´ans Cedex 2, France,
Laboratoire L. Lafon, 19, Avenue du Professeur Cadiot, BP 22, 94701 Maisons-Alfort Cedex, France, and Laboratoire
d’Histopathologie, Faculte´ de Me´decine, Universite´ Libre de Bruxelles, CP620, Route de Lennik 808,
B-1070 Bruxelles, Belgique
Received J uly 6, 2001
Among 25 3-aryl-2-quinolone derivatives synthesized, the antitumor activity of some of them
was characterized both in vitro and in vivo. In this series, no compound appeared to be cytotoxic
in vitro, as was known by the colorimetric MTT assay carried out on 12 distinct human cancer
cell lines obtained from the American Type Culture Collection. Indeed, the concentration values
decreasing the growth of the 12 cell lines by at least 50% (IC50 index) were always higher than
10-5 M. We then made use of a computer-assisted phase-contrast videomicroscopy system to
quantitatively determine in vitro the level of migration of living MCF-7 human breast cancer
cells. For example, at 10-7 M, compounds 7, 13, 16, and 28 markedly decreased the migration
level of these MCF-7 human breast cancer cells. The in vivo determination of the maximum
tolerated dose showed that all compounds tested were definitively nontoxic. When the nontoxic,
antimigratory compound 16 was combined with either doxorubicin or etoposide, two cytotoxic
compounds routinely used in the clinic, this led to additive in vivo benefits from this treatment
(as compared to individual administrations of the drugs) when the MXT mouse mammary
adenocarcinoma was used. Thus, nontoxic antimigratory compounds, including the 2-quinolone
derivatives synthesized here, can actually improve the efficiency of antitumor treatment when
combined with conventional cytotoxic agents.
In tr od u ction
population from which these tumor cells issue.6 These
findings thus led to the development of anticancer drugs
activating the cell death of tumor cells rather than
inhibiting their proliferation.7 Most of these agents
target topoisomerases, which play crucial roles in DNA
duplication.8 They include, for example, doxorubicin
(DOX) and etoposide (ETO) as inhibitors of topoisom-
erase II and irinotecan and topotecan as inhibitors of
topoisomerase I. Thus, most of the agents used today
in hospitals to treat cancer patients are drugs that more
or less directly target the cell kinetics (i.e., proliferation
vs death) of the cancer to be combatted. Great hopes
are placed in complementary strategies including an-
tiangiogenic,9 immunotherapeutic,10 and gene therapy11
approaches. The fact nevertheless remains that the
great majority of the drugs used in the standard
treatment of cancer are toxic to highly toxic, and this
limits their clinical use to a relatively low number of
administrations per patient. In addition, several of these
compounds must be combined into polychemotherapeu-
tic regimens in order to have any effect against cancer.
By way of evidence, such anticancer drug combinations
increase the toxicity of the treatment and once again
limit the number of administrations that can be applied
per patient. In addition, a hallmark of malignancy,
which is as important as cell kinetics, is the migration
of cancer cells escaping from the tumor bulk that first
invade neighboring tissue and then establish metastas-
es. Antitubulin compounds, one major class of antican-
From a schematic point of view, the development of
malignancy in a given tissue occurs as follows. Some
genomic mutations perturb cell cycle kinetics by in-
creasing cell proliferation or decreasing cell death (or
both), and these features lead to unrestrained growth
of the genomically transformed cell population.1 Then,
some cells from this transformed cell population switch
to the angiogenic phenotype, and this enables them to
recruit endothelial cells from the healthy tissue, leading
to a neoangiogenic process enabling the sustained
growth of the developing neoplastic tissue.2 Then, some
cells migrate from the tumor bulk and colonize new
tissues (the metastatic process), using blood or lym-
phatic vessels as major routes of migration.3 The first
chemotherapeutic agents used against cancer in clinics
were drugs that target processes including nucleoside
synthesis, strand replication, etc., i.e., processes control-
ling DNA replication and therefore cell proliferation.4
The discovery of programmed cell death (apoptosis)
by Kerr and colleagues in the early 1970s5 was evidence
that rapidly growing tumors are not always tumors
exhibiting high levels of cell proliferation but often low
levels of cell death as compared to the normal cell
* To whom correspondence should be addressed. Tel.: (33)238 49
45 85. Fax: (33)238 41 72 81. E-mail: benoit.joseph@univ-orleans.fr.
† Universite´ d’Orle´ans.
‡ Laboratoire L. Lafon.
§ Universite´ Libre de Bruxelles.
10.1021/jm010978m CCC: $22.00 © 2002 American Chemical Society
Published on Web 05/11/2002

2544 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12
J oseph et al.
Sch em e 1a
cer drugs, already target tumor cell migration or at least
partly.12 While effective against human cancers, these
compoundssincluding, for example, vincristine, vinorel-
bine, and paclitaxelsare nevertheless also toxic.
It has been known for many years that flavonols, as
quercetin, are polyphenols, which act as free radical
scavangers and could be significant anticarcinogenic
substances (through food intake) due to not yet well-
defined biological mechanisms.13 Such flavonoids inhibit
proliferation of the MCF-7 human breast cancer cell
line.14 Yoshida et al. have suggested that antiprolifera-
tive effects of quercetin were due to the specific arrest
of the G1 phase of the cell cycle.15 However, the results
of epidemiological studies reported so far do not show a
clear protective effect of flavonol intake (through food
and tea consumption) on cancer risk but a protective
effect on selected cancers in a specific population cannot
be ruled out. Because of that, hypotheses are proposed
that a protective role of quercetin (from onions) in
explaining the lower risk of stomach cancer might be
due to interaction with early processes in tumor devel-
opment (i.e., angiogenesis, cell migration, etc.). For this
reason, we have considered the 2-quinolone template
as a possible source of chemical modulation when
looking at the benzopyrane-4-one ring. Moreover, varia-
tions on the 3-aryl moiety were designed with the
variety of compounds known in the flavonoid/isofla-
vonoid families in mind.
a
Reagents and conditions: (a) Toluene, room temperature, 1
h. (b) (i) POCl3, DMF, -30 °C to 75 °C; (ii) AcOH/H2O, reflux 3 h.
Sch em e 2a
We have prepared 25 3-aryl-2-quinolone derivatives
A with the aim of developing nontoxic antimigratory
compounds that could complement the anticancer action
of conventional drugs already used in hospitals. Some
rationale as to why 3-aryl-2-quinolones can be antimi-
gratory agents is given in the discussion.
a
Reagents and conditions: (a) PPSE, 110 °C, 2 h.
Sch em e 3a
Ch em istr y
The syntheses of 3-aryl-2-quinolone derivatives have
been accomplished as described in Schemes 1-6. Ac-
cording to the literature method,16,17 the key 3-aryl-2-
quinolones 2a -g were synthesized. As described in
Scheme 1, substituted anilines were acylated with
phenylacetyl chloride, 4-methoxyphenylacetyl chloride,
or 4-benzyloxyphenylacetyl chloride to give the corre-
sponding amides 1a -g,18 which were then treated with
phosphorus oxychloride and N,N-dimethylformamide
(DMF) followed by an acid hydrolysis to give 3-aryl-2-
quinolones 2a -g. Compounds 2c,e were also prepared
through a one pot route (Scheme 2). Thus, the conden-
sation of 3,5-dimethoxyaniline and ethyl 2-(phenyl or
4-methoxyphenyl)-2-formyl acetate19 was carried out in
the presence of polyphosphoric acid trimethylsilyl ester
(PPSE)20 at 110 °C for 2 h to give the desired compounds
2c,e, respectively, in 16 and 32% yield. In an indepen-
dent but closed work published in 1997, Bisagni et al.
observed the same results by thermal cyclization of 3,5-
dimethoxyaniline and ethyl 2-(4-methoxyphenyl)-2-
formyl acetate.21
a
Reagents and conditions: (a) NaH, RX, DMF, 0 °C and then
90 °C. (b) H2, Pd-C 10%, 1,4-dioxane, room temperature, 4 h.
Alkylation of quinolone 2e led to the preparation of
target compounds 3-10 and O-alkylated derivatives
3a -10a (Scheme 3). In all cases, the N-alkylated
derivative was obtained as the major product. The
structure of both series of compounds was unambigu-
ously confirmed by 13C nuclear magnetic resonance
(NMR) (i.e., for 5, δ 45.3 (NCH2) and for 5a , δ 63.0
(OCH2)) and by analysis of the literature published on
this topic.22 Catalytic hydrogenolysis of 10 led to the acid
11. Compounds 12-14 bearing a functionalized ethyl-
enic chain were obtained via a Michael addition with

3-Aryl-2-Quinolone Derivatives
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12 2545
Sch em e 4a
Sch em e 5a
a
Reagents and conditions: (a) 48% HBr, AcOH reflux, 3 days.
(b) 48% HBr, AcOH, reflux, 5 h. (c) Lawesson’s reagent, toluene,
reflux, 18 h. (d) (Ac)2O, pyridine, room temperature, 18 h.
Sch em e 6a
a
Reagents and conditions: (a) CH2dCHR, Triton B, DMF, room
temperature, 18 h. (b) H2, Pd/C 10%, 1,4-dioxane, room temper-
ature, 48 h. (c) Bu3SnN3, toluene, 105 °C, 65 h. (d) DCC, HOBt,
(Et)2NH, DMF, room temperature, 24 h. (e) PPA, P2O5, 110 °C,
45 min.
various acrylates (Scheme 4). Catalytic hydrogenolysis
of 14 led to the acid 15. Compound 13 was transformed
into the 5-tetrazolylethyl derivative 16 by using tribu-
tyltin azide (Scheme 4). The dicyclohexylcarbodiimide/
1-hydroxybenzotriazole coupling method was applied to
15 to give compound 17 in good yield (Scheme 4).
Similarly, polyphosphoric acid cyclization of 15 led to
the preparation of tricyclic target compound 18 (Scheme
4).
a
Reagents and conditions: (a) ICH3, THF, room temperature,
12 h. (b) NH2NHPh or NH2NH-2-Pyr, EtOH, sealed tube, 90 °C.
The O-demethylation of 2e and 3 was performed with
48% HBr in acetic acid to give, respectively, 19 and 20
in 3 days or 21 and 22 in 5 h (Scheme 5). Compounds
2e and 3 were converted into the corresponding 2-thio-
quinolones 23 and 24 by treatment with Lawesson’s
reagent in toluene (Scheme 5). Compounds 25 and 26
were generated by acetic anhydride-pyridine acetyla-
tion of 20 and 22 in 60% yield (Scheme 5). Treatment
of 24 with iodomethane in tetrahydrofuran (THF)
afforded the iodide salt 27 in 79% yield. Reaction of 27
with phenylhydrazine or 2-pyridinylhydrazine in re-
fluxing ethanol gave hydrazones 28 and 29 in fair yields
(Scheme 6). The yields and/or analytical data of com-
pounds 1-26, 28, and 29 are summarized in Tables
1-3.
ing six compounds 2e, 7, 15, 19, 20, and 23 were without
any actual effect at this 10-5 M dose. At the 10-6 M dose,
only few compounds significantly decreased the growth
levels of human cancer cell populations. At the 10-7
M
dose, only two compounds (17 and 21) showed marginal
cytotoxic effects. The determination of the maximum
tolerated dose (MTD) index in vivo revealed that all
quinolones prepared were found nontoxic at 160 mg/
kg.
The effects of these quinolone derivatives were then
characterized in vitro on MCF-7 human breast cancer
cell motility by means of computer-assisted phase-
contrast videomicroscopy.24 The migration features of
cancer cells involve two distinct but complementary
processes, i.e., motility and invasion. Motility refers to
the capacity of cells to move (including polymerization/
depolymerization dynamics of the actin skeleton and
integrin moving in adhesion complexes), while invasion
refers to the capacity of cells to modulate their sur-
rounding environment by proteolytic activity. The sys-
tem that we set up thus enabled the motility level of
living cells to be quantitatively determined in vitro.
However, we showed with respect to human glioma
cancer cells that their in vitro motility characteristics,
P h a r m a cologica l Resu lts a n d Discu ssion
The absence of in vitro cytotoxicity of 3-aryl-2-quin-
olone derivatives so prepared was demonstrated by
means of the MTT colorimetric assay23 on 12 distinct
human cancer cell lines. The results show that when
compared to the control value when assayed at 10-5 M,
19 of the 25 2-quinolone derivatives synthesized sig-
nificantly decreased the mean growth value of the 12
human cancer cell lines under study, while the remain-

2546 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12
Ta ble 1. Structures, Yields, and Physical Data of Compounds 1 and 2
J oseph et al.
compd
R
R1
R2
R3
R4
yield (%)
mp (°C)
formula
anal
1a
1b
1c
1d
1e
1f
H
H
H
OCH3
OCH3
OCH3
OBn
H
H
H
OCH3
OCH3
OCH3
OBn
H
H
OCH3
H
OCH3
OCH3
OCH3
H
H
OCH3
H
OCH3
OCH3
OCH3
H
H
H
OCH3
H
OCH3
H
OCH3
H
H
OCH3
H
OCH3
H
92
89
87
91
82
93
81
10
28
80-81a,b
118-119c,d
109-111a
47-48a
C15H15NO2
C15H15NO2
C16H17NO3
C16H17NO3
C17H19NO4
C17H19NO4
C23H23NO4
C16H13NO2
C16H13NO2
C17H15NO3
C17H15NO3
C18H17NO4
C18H17NO4
C24H21NO4
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
OCH3
H
H
H
OCH3
H
OCH3
H
OCH3
H
H
OCH3
H
OCH3
H
H
135-137a,e
89-90a
H
1g
2a
2b
2c
2d
2e
2f
H
122-123c
188-189f
243-244f
257-258f,g
148-149f
254-255f,h
186-187f
234-235f
H
OCH3
H
16Meth.A,B
H
10
H
OCH3
H
OCH3
22Meth.A, 32Meth.B
H
45
20
2g
H
a
b
d
Toluene (recrystallization solvent). Literature18 85 °C. c EtOAc/PE (recrystallization solvent). Literature18 123 °C. e Literature21
150 °C. f EtOAc (recrystallization solvent). Literature17 243 °C. Literature21 260 °C; Meth. A, Meth. B, see experimental section.
g
h
Ta ble 2. Structures and Physical Data of Compounds 3-18
compd
R5
mp (°C)
formula
anal
3
4
CH3
125-126a
160-161a
178-179b
113-114c
94-95c
C19H19NO4
C22H23NO6
C24H28N2O5
C22H26N2O4
C23H28N2O4
C20H18N2O4
C22H22N2O4
C27H25NO6
C20H19NO6
C22H23NO6
C21H20N2O4
C28H27NO6
C21H21NO6
C21H21N5O4
C25H30N2O5
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
CH2COOCH2CH3
CH2CON(CH2CH3)2
CH2CH2N(CH3)2
CH2CH2CH2N(CH3)2
CH2CN
5
6
7
8
208-209b
157-158b
199-200c
179-180c
100-101b
156-157a
124-125d
194-195c
234-235c
137-138b
9
CH2CH2CH2CN
CH2COOBn
CH2COOH
CH2CH2COOCH3
CH2CH2CN
CH2CH2COOBn
CH2CH2COOH
5-tetrazolylethyl
CH2CH2CON(CH2CH3)2
10
11
12
13
14
15
16
17
18
240-241e
C21H19NO5
C, H, N
a
b
d
Recrystallization solvent: EtOAc/PE. EtOAc. c Et2O. Et2O/PE. e CH2Cl2/PE.
as determined by the system that we set up, correlate
with their in vivo invasive capacities.24,25 The fact that
antimigratory effects observed in vitro for a given
compound can relate to antimetastatic potential is
already demonstrated for inhibitors of hepatocyte growth
factor receptor.26,27
Preliminary data indicated that this first series of
quinolone derivatives exhibits interacting effects at the
cell migration level (data not shown) with inhibition of
migration effects seen for various chemical types of R5
or X radicals: for example, compounds 7 (X ) O, R5 )
-(CH2)3-N(CH3)2), 13 (X ) O, R5 ) -(CH2)2-CN), 16
(X ) O, R5 ) 5-tetrazoylethyl), and 28 (X ) NNHPh,
R5 ) -CH3). Figure 1 illustrates the antimigratory
effects obtained with respect to compounds 13 and 16.
Both compounds induce a significant decrease in MCF-7
human breast cancer cell migration, with more marked
and dose-dependent effects for compound 16 than 13.
Compound 16 had a significant antimigratory effect on
the MCF-7 cells at the definitively noncytotoxic 10-7
M
dose. At the lowest dose tested for compound 16, i.e.,
10-8 M, a slight stimulation of cell migration was
observed (Figure 1B).
3-Aryl-2-quinolones are reported to be inhibitors of
arachidonic acid-induced platelet agregation28 and also
related to 3-arylpyridopyrimidines that are reported to

3-Aryl-2-Quinolone Derivatives
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12 2547
Ta ble 3. Structures and Physical Data of Compounds 19-26, 28, and 29
compd
R
R1
R3
R5
X
mp (°C)
formula
anal
19
20
21
22
23
24
25
26
28
29
H
H
H
H
CH3
CH3
Ac
Ac
CH3
CH3
H
H
H
H
H
CH3
H
CH3
H
CH3
CH3
CH3
CH3
CH3
O
O
O
O
S
S
O
O
>280a,b
>280b
C15H11NO4
C16H13NO4
C17H15NO4
C18H17NO4
C18H17NO3S
C19H19NO3S
C22H19NO7
C20H19NO5
C25H25N3O3
C24H24N4O3
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
C, H, N
CH3
CH3
CH3
CH3
Ac
CH3
CH3
CH3
CH3
CH3
CH3
CH3
Ac
CH3
CH3
CH3
275-276b
204-205b
229-230c
176-177d
206-207b
148-149d
148-149e
140-141e
NNHPh
NNH-2-Pyr
a
b
d
Literature21 >280 °C. Recrystallization solvent: EtOAc. c Et2O. EtOAc/PE. e MeOH.
human breast cancer,33,34 and whether any benefit could
be obtained from additive treatment with such combi-
nations. We chose the MXT mouse mammary adeno-
carcinoma model originating from ductal structures33,34
for the following reasons. The first concerns the fact that
up to 1986 leukemia models were the NCI’s preferred
models for testing new investigational agents in vivo,
but these models were then abandoned because they are
too chemosensitive and consequently fail to reflect
clinical reality.35 We thus preferred to make use of a
solid tumor. Second, the MXT mouse mammary cancer
was selected because more than 80% of female breast
cancers are invasive intraduct carcinomas, i.e., not
otherwise specified breast cancers,33 while most of the
experimental murine mammary tumors originate from
the glandular acini.33 The biological characteristics of
mammary tumors from ductal as opposed to acinar
structures differ markedly.36 Figure 2 illustrates the
effects obtained with ETO (A), DOX (B), and compound
16 (C). The data indicate that ETO significantly (P <
0.05 to P < 0.001 as compared to control at the different
days postgraft) decreased the MXT tumor growth when
tested at 20 (MTD/4) or 10 (MTD/8) mg/kg (Figure 2A).
At 40 mg/kg (MTD/2), ETO showed itself to be highly
toxic, while at 5 mg/kg (MTD/16), it became ineffective
(Figure 2A). We chose the 10 mg/kg ETO dose for a
combined treatment with compound 16 (Figure 3A). The
same experimental approach was used with respect to
DOX, which was tested at MTD/2 (5 mg/kg), MTD/4 (2.5
mg/kg), MTD/8 (1.25 mg/kg), and MTD/16 (0.63 mg/kg).
The MTD/2 treatment appeared highly toxic, while the
MTD/16 treatment was ineffective (Figure 2B). The
MTD/4 treatment (P < 0.05 to P < 0.01) was more
effective than the MTD/8 treatment (P < 0.05 to P <
0.001) and was chosen for the combined treatment with
compound 16 (Figure 3B). This latter was assayed at
MTD/2 (80 mg/kg), MTD/4 (40 mg/kg), and MTD/8 (20
mg/kg). Only 80 mg/kg, the highest dose tested, showed
slight, but nevertheless significant (P < 0.05), antitumor
effects when compared to the control (Figure 2C). The
40 mg/kg dose (which exhibited no significant effects per
se; Figure 2C) was the one chosen for combined treat-
ment with either ETO (Figure 3A) or DOX (Figure 3B).
F igu r e 1. Characterization of the antimigratory effects
induced by compounds 13 (A) and 16 (B) on the migration level
of the human MCF-7 breast cancer cells. The migration level
was quantitatively determined by means of computer-assisted
phase-contrast videomicroscopy on several hundreds of cells
per experimental condition. The data are reported as mean (
SEM.
be both kinase receptor inhibitors29 and angiogenesis
inhibitors.30 These data already published in the litera-
ture can have some relevance with our data and can
therefore suggest potential 3-aryl-2-quinolone deriva-
tive-mediated effects on distinct types of kinase recep-
tors that in turn will significantly decrease the motility
levels of human breast cancer cells.
Compound 16 was then combined with either DOX
or ETO (which are two clinically active anticancer
drugs) in order to investigate in vivo the MXT mouse
mammary adenocarcinoma,31,32 which closely mimics
The data obtained revealed that real additive anti-
cancer influences were observed for both compound 16/

2548 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12
J oseph et al.
F igu r e 3. Characterization of the anticancer influence exer-
cised by ETO combined with 16 (× in A) and by DOX combined
with 16 (× in B) on the growth of the MXT mouse mammary
adenocarcinoma. ETO was assayed at 10 mg/kg, i.e., at MTD/8
(0 in A), DOX at MTD/4 (2.5 mg/kg; 4 in B), and 16 at MTD/4
(40 mg/kg; O in A and B). The control conditions are symbolized
by black dots. There were nine mice per experimental group.
The P levels of statistical significance were determined by
means of the nonparametric Mann-Whitney test. The results
are depicted as mean values (the symbols) ( their SEM (the
bars). The P values in A and B were calculated at the end of
the experiments (see the vertical dotted line) when at least
four mice survived in each experimental group.
F igu r e 2. Characterization of the anticancer influence of ETO
(A), DOX (B), and 16 (C) on the growth of the MXT mouse
mammary adenocarcinoma. ETO was tested at 40 (MTD/2; O),
20 (MTD/4; ]), 10 (MTD/8; 0), and 5 (MTD/16; ×) mg/kg. DOX
was tested at 5 (MTD/2; O), 2.5 (MTD/4; ]), 1.25 (MTD/8; 0),
and 0.63 (MTD/16; ×) mg/kg. Compound 16 was tested at 80
(MTD/2; O), 40 (MTD/4; ]), and 20 (MTD/8; 0). The control
conditions are symbolized by black dots. ETO and DOX were
administered i.p. (0.2 mL) nine times according to the experi-
mental schedule depicted by the black arrows in the upper
left-hand part of panels A and B. Derivative 16 was admin-
istered 20 times (see the black arrows in the upper left-hand
part of panel C). There were nine mice per experimental group.
The P levels of statistical significance were determined by
means of the nonparametric Mann-Whitney test. The results
are depicted as mean values (the symbols) ( their SEM (the
bars).
combination of compound 16 (40 mg/kg) and DOX (2.5
mg/kg) also significantly (P ) 0.008) increased the
antitumor effects observed with each compound tested
individually (Figure 3B). The P values in Figure 3A,B
were calculated at the end of the experiments (see the
vertical dotted line) when at least four mice survived
in each experimental group. These additive effects
observed in terms of antitumor activities between
compound 16 and DOX or ETO can result from a
combined antimigratory effect brought by compound 16
on the tumor cells and an antiproliferative effect
brought by ETO or DOX on the remaining tumor cells.
In conclusion, the data from the present study clearly
indicate that an improvement in treatment was ob-
tained in the case of the MXT mouse mammary tumor
with a combination of the nontoxic antimigratory com-
pound 16 (taken up as a representative of the group of
study drugs) and the conventional cytotoxic drugs used
routinely in hospitals, i.e., either ETO or DOX, which
are two topoisomerase II inhibitors. The mechanism of
action of compound 16 and congeners still remains to
be elucidated. Further combinations with various other
ETO (Figure 3A) and compound 16/DOX (Figure 3B)
combinations, as was evidenced by the increases in the
P levels of statistical significance depicted in Figure
3A,B. Indeed, a slight, but nevertheless significant,
effect (P ) 0.04) was obtained when using 10 mg/kg of
ETO with 40 mg/kg of compound 16, with an actual
increase observed in the survival periods of the MXT
mammary cancer-bearing mice, which increased by 38%
(P ) 0.01) as compared to the control (Figure 3A). The

3-Aryl-2-Quinolone Derivatives
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12 2549
to the procedure described for 1a . IR (KBr): ν 3292, 1658, 1615
anticancer drugs are currently being investigated against
several other cancer cell lines.
cm-1 1H NMR (250 MHz, CDCl3): δ 3.66 (s, 2H, CH2), 3.74
.
(s, 6H, CH3), 3.82 (s, 3H, CH3), 6.20 (t, 1H, J ) 2.2 Hz, HAr),
6.62-6.66 (m, 2H, HAr), 6.95 (d, 2H, J ) 7.5 Hz, HAr), 6.97 (br
s, 1H, NH), 7.23 (d, 2H, J ) 7.5 Hz, HAr). 13C NMR (62.90 MHz,
CDCl3): δ 44.0, 55.3, 55.4 (2), 96.7, 97.9 (2), 114.4 (2), 126.2,
Exp er im en ta l Section
Ch em istr y. Melting points were determined using a Bu¨chi
capillary instrument and are uncorrected. IR spectra were
recorded on a Perkin-Elmer FTIR Paragon 1000 spectropho-
tometer. NMR spectra were recorded at 300 K on a Bruker
Avance DPX 250 spectrometer. Chemical shifts are expressed
in parts per million relative to tetramethylsilane (TMS), and
spin multiplicities are given as s (singlet), br s (broad singlet),
d (doublet), dd (double doublet), t (triplet), and m (multiplet).
Mass spectra were recorded on a Perkin-Elmer SCIEX API
300 instrument using ionspray methodology. Elemental com-
positions of the compounds agreed to within 0.4% of the
calculated value. Thin-layer chromatography (TLC) was run
on precoated silica gel plates (Merck 60F254), and spots were
visualized with a UV light at 254 nm. Column chromatography
was carried out using Merck silica gel (230-400 mesh). Most
chemicals and solvents were analytical grade and used without
further purification. Ethyl 2-(phenyl or 4-methoxyphenyl)-2-
formyl acetates were prepared according to ref 19. Commercial
reagents were purchased from Acros Company.
130.7 (2), 139.4, 159.0, 161.0 (2), 169.5. MS: m/z 302 (M+
1).
+
N-(2,5-Dim et h oxyp h en yl)-2-(4-m et h oxyp h en yl)a cet -
a m id e (1f). Starting from 2,5-dimethoxyaniline and 4-meth-
oxyphenylacetyl chloride, compound 1f was obtained according
to the procedure described for 1a . IR (KBr): ν 3292, 1659, 1614
cm-1 1H NMR (250 MHz, CDCl3): δ 3,67 (s, 3H, CH3), 3.68
.
(s, 2H, CH2), 3.75 (s, 3H, CH3), 3.81 (s, 3H, CH3), 6.53 (dd,
1H, J ) 3.0, 9.0 Hz, HAr), 6.71 (d, 1H, J ) 9.0 Hz, HAr), 6.92
(d, 2H, J ) 8.8 Hz, HAr), 7.25 (d, 2H, J ) 8.8 Hz, HAr), 7.82 (br
s, 1H, NH), 8.80 (d, 1H, J ) 3.0 Hz, HAr). 13C NMR (62.90 MHz,
CDCl3): δ 44.2, 55.3, 55.7, 56.3, 105.5, 108.6, 110.9, 114.4 (2),
126.4, 128.3, 130.6 (2), 142.0, 153.9, 158.9, 169.3. MS: m/z 302
(M+ + 1).
N-(3,5-Dim eth oxyp h en yl)-2-(4-ben zyloxyp h en yl)a cet-
a m id e (1g). Starting from 3,5-dimethoxyaniline and 4-ben-
zyloxyphenylacetyl chloride, compound 1g was obtained fol-
lowing a procedure as described for 1a . The crude residue was
purified by column chromatography (eluent ethyl acetate/
petroleum ether 6:4) to afford 1g. IR (KBr): ν 3291, 1659, 1610
Gen er a l P r oced u r e for P r ep a r a tion of Com p ou n d 1.
N-(2-Met h oxyp h en yl)-2-p h en yla cet a m id e (1a ).18 To a
stirred solution of o-anisidine (1.37 mL, 10.0 mmol) in toluene
(14 mL) at 0 °C was added dropwise a solution of phenylacetyl
chloride (1.62 mL, 10.0 mmol) in toluene (5 mL). The reaction
mixture was stirred at room temperature for 1 h and then
treated with a saturated NaHCO3 solution. The biphasic
solution was vigorously stirred for 30 min, then decanted, and
finally separated. The collected aqueous phase was extracted
with EtOAc (2 × 10 mL). The combined organic layer was dried
over MgSO4 and evaporated. The crude oily residue was
crystallized from toluene to yield 1a . IR (KBr): ν 3287, 1652,
cm-1 1H NMR (250 MHz, CDCl3): δ 3.66 (s, 2H, CH2), 3.74
.
(s, 3H, CH3), 3.75 (s, 3H, CH3), 5.08 (s, 2H, CH2), 6.21 (t, 1H,
J ) 2.2 Hz, HAr), 6.66 (d, 2H, J ) 2.2 Hz, HAr), 7.00 (d, 2H, J
) 8.8 Hz, HAr), 7.08 (br s, 1H, NH), 7.24 (d, 2H, J ) 8.8 Hz,
H
Ar), 7.33-7.46 (m, 5H, HAr). 13C NMR (62.90 MHz, CDCl3): δ
44.2, 55.5 (2), 70.2, 96.9, 98.0 (2), 115.7 (2), 126.6, 127.6 (2),
128.2, 128.8 (2), 130.8 (2), 136.9, 139.6, 158.4, 161.1 (2), 169.7.
MS: m/z 378 (M+ + 1).
Gen er a l P r oced u r e for P r ep a r a tion of Com p ou n d 2.
8-Meth oxy-3-p h en yl-1,2-d ih yd r o-2-qu in olin on e (2a ). To a
solution of POCl3 (1.75 mL, 19 mmol) at -30 °C was dropwise
added anhydrous DMF (0.31 mL, 4.0 mmol). The solution was
stirred for 15 min at -30 °C, and then, compound 1a (2.7
mmol) was dropwise added. Under vigorous stirring, the
reaction mixture was allowed to reach room temperature and
was then heated at 75 °C for 1.5 h. The solution was poured
into ice, neutralized with 30% aqueous NH4OH, and extracted
with CH2Cl2. The organic layer was dried over MgSO4 and then
evaporated in vacuo. The crude residue obtained was dissolved
in glacial acetic acid (4.75 mL) and H2O (0.15 mL). The final
solution was stirred at reflux for 3 h. After it was cooled, the
solvent was evaporated in vacuo. The crude residue was
diluted in H2O, neutralized with 25% NaOH, and finally
extracted with CH2Cl2. The organic layer was dried over
MgSO4 and then evaporated in vacuo. The crude residue was
crystallized from EtOAc. Crystals was filtered and washed
twice with EtOAc to give 2a . IR (KBr): ν 1646, 1607 cm-1. 1H
NMR (250 MHz, DMSO-d6): δ 3.92 (s, 3H, CH3), 7.13-7.16
(m, 2H, HAr), 7.30-7.46 (m, 4H, HAr), 7.74-7.77 (m, 2H, HAr),
8.09 (s, 1H, dCH), 10.98 (br s, 1H, NH). 13C NMR (62.90 MHz,
DMSO-d6): δ 56.5, 111.3, 120.3, 120.4, 128.3 (2), 128.4, 128.7,
129.2 (2), 132.6, 136.7, 138.2, 145.9, 161.1. MS: m/z 252 (M+
+ 1).
6-Meth oxy-3-ph en yl-1,2-dih ydr o-2-qu in olin on e (2b). Ac-
cording to the procedure described for 2a , the 2-quinolone 2b
was prepared from 1b. IR (KBr): ν 1645, 1618 cm-1. 1H NMR
(250 MHz, DMSO-d6): δ 3.80 (s, 3H, CH3), 7.16 (dd, 1H, J )
2.5, 8.9 Hz, HAr), 7.28 (d, 1H, J ) 8.9 Hz, HAr), 7.29 (d, 1H, J
) 2.5 Hz, HAr), 7.34-7.47 (m, 3H, HAr), 7.76 (d, 2H, J ) 6.8
Hz, HAr), 8.06 (s, 1H, dCH), 11.85 (br s, 1H, NH). 13C NMR
(62.90 MHz, DMSO-d6): δ 55.4, 109.4, 116.0, 119.5, 120.1,
127.8, 127.9 (2), 128.7 (2), 131.9, 132.9, 136.4, 137.2, 154.2,
160.6. MS: m/z 252 (M+ + 1).
1
1598 cm-1. H NMR (250 MHz, CDCl3): δ 3.72 (s, 3H, CH3),
3.76 (s, 2H, CH2), 6.81 (dd, 1H, J ) 1.8, 8.0 Hz, HAr), 6.91-
7.05 (m, 2H, HAr), 7.21-7.37 (m, 5H, HAr), 7.80 (br s, 1H, NH),
8.35 (dd, 1H, J ) 1.8, 8.0 Hz, HAr). 13C NMR (62.90 MHz,
CDCl3): δ 45.2, 55.7, 109.9, 119.5, 121.1, 123.7, 127.4, 127.6,
129.0 (2), 129.6 (2), 134.6, 147.8, 168.8. MS: m/z 242 (M+
+
1).
N-(4-Meth oxyp h en yl)-2-p h en yla ceta m id e (1b).18 Start-
ing from p-anisidine and phenylacetyl chloride, compound 1b
was obtained according to the procedure described for 1a . IR
(KBr): ν 3290, 1650, 1603 cm-1. H NMR (250 MHz, CDCl3):
1
δ 3.72 (s, 2H, CH2), 3.77 (s, 3H, CH3), 6.81 (d, 2H, J ) 9.0 Hz,
H
Ar), 7.00 (br s, 1H, NH), 7.28-7.43 (m, 7H, HAr). 13C NMR
(62.90 MHz, CDCl3): δ 44.8, 55.6, 114.2 (2), 121.9 (2), 127.7,
129.3 (2), 129.7 (2), 130.8, 134.7, 156.7, 169.1. MS: m/z 242
(M+ + 1).
N-(3,5-Dim eth oxyph en yl)-2-ph en ylacetam ide (1c). Start-
ing from 3,5-dimethoxyaniline and phenylacetyl chloride,
compound 1c was obtained according to the procedure de-
scribed for 1a . IR (KBr): ν 3286, 1657, 1616 cm-1 1H NMR
.
(250 MHz, CDCl3): δ 3.70 (s, 2H, CH2), 3.74 (s, 6H, CH3), 6.21
(t, 1H, J ) 2.2 Hz, HAr), 6.66 (d, 2H, J ) 2.2 Hz, HAr), 7.09 (br
s, 1H, NH), 7.30-7.40 (m, 5H, HAr). 13C NMR (62.90 MHz,
CDCl3): δ 44.9, 55.4 (2), 96.8, 97.9 (2), 127.7, 129.2 (2), 129.5
(2), 134.3, 139.4, 161.0 (2), 169.1. MS: m/z 272 (M+ + 1).
N -(2-Me t h oxyp h e n yl)-2-(4-m e t h oxyp h e n yl)a ce t a m -
id e (1d ). Starting from o-anisidine and 4-methoxyphenylacetyl
chloride, compound 1d was obtained according to the proce-
dure described for 1a . IR (KBr): ν 3375, 1667, 1597 cm-1. H
1
NMR (250 MHz, CDCl3): δ 3,68 (s, 2H, CH2), 3.72 (s, 3H, CH3),
3.82 (s, 3H, CH3), 6.79 (dd 1H, J ) 1.5, 8.0 Hz, HAr), 6.89-
7.03 (m, 4H, HAr), 7.25 (d, 2H, J ) 8.8 Hz, HAr), 7.79 (br s, 1H,
NH), 8.33 (dd, 1H, J ) 1.5, 8.0 Hz, HAr). 13C NMR (62.90 MHz,
CDCl3): δ 44.2, 55.3, 55.7, 109.9, 114.4 (2), 119.5, 121.0, 123.6,
126.6, 127.6, 130.7 (2), 147.8, 158.9, 169.3. MS: m/z 272 (M+
+ 1).
5,7-Dim ethoxy-3-phen yl-1,2-dih ydr o-2-quinolin on e (2c).17
Meth od A. According to the procedure described for 2a , the
2-quinolone 2c was prepared from 1c. Meth od B. To a solution
of 2-phenyl-2-formyl acetate (7.37 g, 38.4 mmol) and 3,5-
dimethoxyaniline (5.0 g, 32 mmol) stirred at room temperature
N-(3,5-Met h oxyp h en yl)-2-(4-m et h oxyp h en yl)a cet a m -
id e (1e).21 Starting from 3,5-dimethoxyaniline and 4-methoxy-
phenylacetyl chloride, compound 1e was obtained according

2550 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12
J oseph et al.
1
for 30 min was added a freshly prepared solution of PPSE (15.0
g of phosphorus pentoxide and 36 mL of hexamethyldisiloxane
in 1,2-dichloroethane stirred at 90 °C until complete dissolu-
tion of P2O5 and then evaporation of solvent). The reaction
mixture was stirred at 100 °C for 1 h. After the mixture was
cooled, CH2Cl2 (1000 mL) and H2O (500 mL) were added to
the solution. The biphasic solution was vigorously stirred
overnight, then decanted, and finally separated. The collected
aqueous phase was extracted with CH2Cl2 (2 × 100 mL). The
combined organic layer was dried over MgSO4 and evaporated.
The crude oily residue was crystallized from EtOAc to give
ν 1635, 1596 cm-1. H NMR (250 MHz, CDCl3): δ 3.73 (s, 3H,
CH3), 3.83 (s, 3H, OCH3), 3.91 (s, 6H, OCH3), 6.30 (d, 1H, J )
2.0 Hz, HAr), 6.37 (d, 1H, J ) 2.0 Hz, HAr), 6.94 (d, 2H, J ) 7.5
Hz, HAr), 7.67 (d, 2H, J ) 7.5 Hz, HAr), 8.12 (s, 1H, dCH). 13C
NMR (62.90 MHz, CDCl3): δ 30.3, 55.3, 55.5, 55.8, 90.2, 92.6,
106.1, 113.5 (2), 127.0, 130.0, 130.1 (2), 130.2, 141.7, 157.6,
159.1, 162.2 (2). MS: m/z 326 (M+ + 1).
2,5,7-Tr im eth oxy-3-(4-m eth oxyp h en yl)qu in olin e (3a ).
mp 106-107 °C (EtOAc/PE). IR (KBr): ν 1621, 1515 cm-1. 1H
NMR (250 MHz, CDCl3): δ 3.86 (s, 3H, OCH3), 3.94 (s, 6H,
OCH3), 4.08 (s, 3H, OCH3), 6.40 (d, 1H, J ) 1.8 Hz, HAr), 6.85
(d, 1H, J ) 1.8 Hz, HAr), 6.97 (d, 2H, J ) 8.8 Hz, HAr), 7.57 (d,
2H, J ) 8.8 Hz, HAr), 8.26 (s, 1H, dCH). 13C NMR (62.90 MHz,
CDCl3): δ 53.5, 55.3, 55.5, 55.6, 95.9, 98.5, 112.8, 113.6 (2),
122.1, 129.6, 130.5 (2), 132.3, 147.9, 156.3, 158.9, 160.6, 161.3.
MS: m/z 326 (M+ + 1). Anal. (C19H19NO4) C, H, N.
1
2c. IR (KBr): ν 1668, 1631 cm-1. H NMR (250 MHz, DMSO-
d6): δ 3.81 (s, 3H, OCH3), 3.89 (s, 3H, OCH3), 6.36 (d, 1H, J )
1.8 Hz, HAr), 6.45 (d, 1H, J ) 1.8 Hz, HAr), 7.28-7.42 (m, 3H,
HAr), 7.69 (d, 2H, J ) 7.0 Hz, HAr), 8.00 (s, 1H, dCH), 11.81
(br s, 1H, NH). 13C NMR (62.90 MHz, DMSO-d6): δ 55.5, 56.0,
90.0, 93.1, 104.5, 126.9, 127.3, 127.9 (2), 128.4 (2), 131.4, 136.7,
140.8, 156.8, 161.4, 162.2. MS: m/z 282 (M+ + 1).
8-Meth oxy-3-(4-m eth oxyp h en yl)-1,2-d ih yd r o-2-qu in o-
lin on e (2d ). According to the procedure described for 2a , the
2-quinolone 2d was prepared from 1d . IR (KBr): ν 1652, 1625
cm-1. 1H NMR (250 MHz, DMSO-d6): δ 3.79 (s, 3H, CH3), 3.90
(s, 3H, CH3), 6.99 (d, 2H, J ) 8.8 Hz, HAr), 7.09-7.15 (m, 2H,
HAr), 7.29 (dd, 1H, J ) 3.2, 6.0 Hz, HAr), 7.74 (d, 2H, J ) 8.8
Hz, HAr), 8.03 (s, 1H, dCH), 10.90 (br s, 1H, NH). 13C NMR
(62.90 MHz, DMSO-d6): δ 55.2, 56.0, 110.5, 113.4 (2), 119.6,
120.0, 121.8, 127.9, 128.5, 129.9 (2), 131.6, 136.4, 145.3, 159.1,
160.7. MS: m/z 282 (M+ + 1).
Eth yl 2-[5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-oxo-1,2-
d ih yd r o-1-qu in olin yl]a ceta te (4). According to the proce-
dure described for 3a , the 2-quinolone 4 and compound 4a were
prepared starting from 2e (1 equiv) and ethyl bromoacetate
(2 equiv). Reaction time, 3 h; eluent chromatography CH2Cl2/
EtOAc 7:3; yield, 70%. IR (KBr): 1735, 1647, 1609 cm-1 1H
.
NMR (250 MHz, CDCl3): δ 1.62 (t, 3H, J ) 7.1 Hz, CH3), 3.83
(s, 3H, OCH3), 3.87 (s, 3H, OCH3), 3.92 (s, 3H, OCH3), 4.22 (q,
2H, J ) 7.1 Hz, CH2), 5.10 (s, 2H, CH2), 6.14 (d, 1H, J ) 2.0
Hz, HAr), 6.30 (d, 1H, J ) 2.0 Hz, HAr), 6.94 (d, 2H, J ) 7.5
Hz, HAr), 7.69 (d, 2H, J ) 7.5 Hz, HAr), 8.18 (s, 1H, dCH). 13C
NMR (62.90 MHz, CDCl3): δ 14.2, 44.8, 55.3, 55.5, 55.9, 61.6,
90.0, 92.8, 106.2, 113.5 (2), 126.6, 129.6, 130.1 (2), 131.0, 141.1,
157.8, 159.2, 161.9, 162.5, 168.4. MS: m/z 398 (M+ + 1).
5,7-Dim eth oxy-3-(4-m eth oxyph en yl)-1,2-dih ydr o-2-qu in -
olin on e (2e).21 According to the procedures described for 2c,
the 2-quinolone 2e was prepared from 1e (method A) or 2-(4-
methoxyphenyl)-2-formyl acetate and 3,5-dimethoxyaniline
(method B). IR (KBr): ν 1664, 1628 cm-1. 1H NMR (250 MHz,
DMSO-d6): δ 3.78 (s, 3H, CH3), 3.81 (s, 3H, CH3), 3.89 (s, 3H,
CH3), 6.35 (d, 1H, J ) 2.0 Hz, HAr), 6.45 (d, 1H, J ) 2.0 Hz,
Eth yl 2-([5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-qu in o-
lin yl]oxy)a ceta te (4a ). Yield, 25%; mp 95-96 °C (EtOAc/PE).
IR (KBr): ν 1754, 1622, 1516 cm-1 1H NMR (250 MHz,
.
CDCl3): δ 1.27 (t, 3H, J ) 7.1 Hz, CH3), 3,86 (s, 3H, OCH3),
3.91 (s, 3H, OCH3), 3.93 (s, 3H, OCH3), 4.24 (q, 2H, J ) 7.1
Hz, CH2), 5.05 (s, 2H, CH2), 6.40 (d, 1H, J ) 2.0 Hz, HAr), 6.76
(d, 1H, J ) 2.0 Hz, HAr), 6.98 (d, 2H, J ) 9.0 Hz, HAr), 7.68 (d,
2H, J ) 9.0 Hz, HAr), 8.30 (s, 1H, dCH). 13C NMR (62.90 MHz,
CDCl3): δ 14.2, 55.3, 55.5, 55.7, 60.9, 62.7, 96.2, 98.6, 113.4,
113.7 (2), 121.7, 129.2, 130.6 (2), 132.8, 147.3, 156.3, 158.8,
159.1, 161.4, 169.5. MS: m/z 398 (M+ + 1). Anal. (C22H23NO6)
C, H, N.
H
H
Ar), 6.95 (d, 2H, J ) 7.5 Hz, HAr), 7.66 (d, 2H, J ) 7.5 Hz,
Ar), 7.96 (s, 1H, dCH), 11.76 (br s, 1H, NH). 13C NMR (62.90
MHz, DMSO-d6): δ 55.1, 55.4, 55.9, 90.0, 93.0, 104.6, 113.3
(2), 126.5, 129.0, 129.6 (2), 130.2, 140.5, 156.6, 158.7, 161.5,
161.9. MS: m/z 312 (M+ + 1).
5,8-Dim eth oxy-3-(4-m eth oxyph en yl)-1,2-dih ydr o-2-qu in -
olin on e (2f). According to the procedure described for 2a , the
2-quinolone 2f was prepared from 1f. IR (KBr): ν 1639, 1571
N,N-Dieth yl-2-[5,7-d im eth oxy-3-(4-m eth oxyp h en yl)-2-
oxo-1,2-d ih yd r o-1-qu in olin yl]a ceta m id e (5). According to
the procedure described for 3, the 2-quinolone 5 and compound
5a were prepared starting from 2e (1 equiv) and 2-chloro-N,N-
diethylacetamide (1.5 equiv). Reaction time, 3 h; eluent
chromatography CH2Cl2/EtOAc 7:3; yield, 61%. IR (KBr):
cm-1 1H NMR (250 MHz, CDCl3): δ 3.85 (s, 3H, CH3), 3.90
.
(s, 3H, CH3), 3.92 (s, 3H, CH3), 6.49 (d, 1H, J ) 8.7 Hz, HAr),
6.84 (d, 1H, J ) 8.7 Hz, HAr), 6.97 (d, 2H, J ) 8.8 Hz, HAr),
7.74 (d, 2H, J ) 8.8 Hz, HAr), 8.19 (s, 1H, dCH), 9.25 (br s,
1H, NH). 13C NMR (62.90 MHz, CDCl3): δ 55.2, 55.8, 56.2,
101.1, 109.7, 111.4, 113.6 (2), 128.7, 128.8, 130.0 (2), 131.4,
131.7, 139.4, 149.8, 159.5, 161.3. MS: m/z 312 (M+ + 1).
1642, 1617, 1601 cm-1. H NMR (250 MHz, CDCl3): δ 1.10-
1
1.23 (m, 6H, CH3), 3.38-3.49 (m, 4H, CH2), 3.83 (s, 3H, OCH3),
3.86 (s, 3H, OCH3), 3.91 (s, 3H, OCH3), 5.17 (s, 2H, CH2), 6.28
(d, 1H, J ) 2.0 Hz, HAr), 6.34 (d, 1H, J ) 2.0 Hz, HAr), 6.93 (d,
2H, J ) 7.0 Hz, HAr), 7.66 (d, 2H, J ) 7.0 Hz, HAr), 8.17 (s,
1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 13.0, 14.2, 40.9,
41.6, 45.3, 55.3, 55.5, 55.8, 90.8, 92.8, 106.3, 113.4 (2), 126.6,
129.9, 130.1 (2), 131.0, 141.7, 157.6, 159.0, 161.9, 162.3, 166.3.
MS: m/z 425 (M+ + 1).
5,7-Dim et h oxy-3-(4-b en zyloxyp h en yl)-1,2-d ih yd r o-2-
qu in olin on e (2g). According to the procedure described for
2a , the 2-quinolone 2g was prepared from 1g. IR (KBr): ν
1629, 1608 cm-1 1H NMR (250 MHz, DMSO-d6): δ 3.81 (s,
.
3H, CH3), 3.90 (s, 3H, CH3), 5.15 (s, 2H, CH2), 6.37 (s, 1H,
Ar), 6.45 (s, 1H, HAr), 7.04 (d, 2H, J ) 8.8 Hz, HAr), 7.31-
H
7.49 (m, 5H, HAr), 7.69 (d, 2H, J ) 7.5 Hz, HAr), 7.97 (s, 1H,
dCH), 11.76 (br s, 1H, NH). 13C NMR (62.90 MHz, DMSO-
d6): δ 55.4, 55.9, 69.2, 90.0, 93.0, 104.6, 114.3 (2), 126.4, 127.6
(2), 127.8, 128.4 (2) 129.2, 129.6 (2), 130.2, 137.1, 140.5, 156.6,
157.7, 161.5, 161.9. MS: m/z 388 (M+ + 1).
N,N-Dieth yl-2-([5,7-d im eth oxy-3-(4-m eth oxyp h en yl)-2-
qu in olin yl]oxy)a ceta m id e (5a ). Yield, 30%; mp 146-147 °C
(EtOAc/PE). IR (KBr): ν 1654, 1624, 1517 cm-1. 1H NMR (250
MHz, CDCl3): δ 1.14 (t, 3H, J ) 7.5 Hz, CH3), 1.28 (t, 3H, J
) 7.5 Hz, CH3), 3.36-3.47 (m, 4H, CH2), 3.85 (s, 3H, OCH3),
3.91 (s, 3H, OCH3), 3.93 (s, 3H, OCH3), 5.17 (s, 2H, CH2), 6.38
(d, 1H, J ) 2.0 Hz, HAr), 6.73 (d, 1H, J ) 2.0 Hz, HAr), 6.97 (d,
2H, J ) 9.0 Hz, HAr), 7.75 (d, 2H, J ) 9.0 Hz, HAr), 8.28 (s,
1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 12.9, 14.2, 40.2,
55.2, 55.4, 55.6, 63.0, 95.8, 98.4, 113.3, 113.5 (2), 121.9, 129.2,
130.6 (2), 132.5, 147.3, 156.2, 158.9, 161.1, 167.3. MS: m/z 425
(M+ + 1). Anal. (C24H28N2O5), C, H, N.
1-[2-(Dim eth yla m in o)eth yl]-5,7-d im eth oxy-3-(4-m eth -
oxyp h en yl)-1,2-d ih yd r o-2-qu in olin on e (6). According to
the procedure described for 3, the 2-quinolone 6 and compound
6a were prepared starting from 2e (1 equiv) and 2-(dimethyl-
amino)ethyl chloride (3 equiv). Reaction time, 3 h; eluent
5,7-Dim et h oxy-3-(4-m et h oxyp h en yl)-1-m et h yl-1,2-d i-
h yd r o-2-qu in olin on e (3). At 0 °C under argon, sodium
hydride (93 mg, 3.86 mmol, 60% oil dispersion) was added
portionwise to a solution of 2e (600 mg, 1.93 mmol) in
anhydrous DMF (30 mL). The mixture was stirred for 15 min
at 0 °C, and then, iodomethane (0.48 mL, 7.72 mmol) diluted
in anhydrous DMF (5 mL) was added. The final solution was
stirred at 90 °C and monitored by TLC (reaction time 18 h).
The cooled mixture was partitioned between H2O (10 mL) and
CH2Cl2 (10 mL). The organic layers were dried over MgSO4
and concentrated in vacuo. The crude residue was purified by
column chromatography (eluent CH2Cl2/EtOAc 8:2) to give
successively 3 (408 mg, 68%) and 3a (157 mg, 25%). IR (KBr):

3-Aryl-2-Quinolone Derivatives
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12 2551
chromatography Et2O/EtOAc 3:7 and then CH2Cl2/MeOH 9:1;
cedure described for 3, the 2-quinolone 9 and compound 9a
were prepared starting from 2e (1 equiv) and 4-chlorobuty-
ronitrile (2 equiv). Reaction time, 3 h; eluent CH2Cl2/EtOAc
1
yield, 67%. IR (KBr): ν 1645, 1617 cm-1. H NMR (250 MHz,
CDCl3): δ 2.39 (s, 6H, CH3), 2.65 (t, 2H, J ) 7.8 Hz, CH2),
3.82 (s, 3H, OCH3), 3.90 (s, 3H, OCH3), 3.91 (s, 3H, OCH3),
4.44 (t, 2H, J ) 7.8 Hz, CH2), 6.28 (d, 1H, J ) 2.0 Hz, HAr),
6.49 (d, 1H, J ) 2.0 Hz, HAr), 6.94 (d, 2H, J ) 8.8 Hz, HAr),
7.67 (d, 2H, J ) 8.8 Hz, HAr), 8.12 (s, 1H, dCH). 13C NMR
(62.90 MHz, CDCl3): δ 41.6, 45.8 (2), 55.3, 55.6, 55.8 (2), 90.1,
92.7, 106.4, 113.5 (2), 126.9, 129.8, 130.1 (2), 130.4, 141.1,
157.7, 159.1, 161.9, 162.4. MS: m/z 383 (M+ + 1).
1
9:1; yield, 33%. IR (KBr): ν 2247, 1639, 1609, 1597 cm-1. H
NMR (250 MHz, CDCl3): δ 2.10-2.21 (m, 2H, CH2), 2.52 (t,
2H, J ) 7.2 Hz, CH2), 3.83 (s, 3H, OCH3), 3.92 (s, 3H, OCH3),
3.93 (s, 3H, OCH3), 4.43 (t, 2H, J ) 7.2 Hz, CH2), 6.31 (d, 1H,
J ) 2.2 Hz, HAr), 6.42 (d, 1H, J ) 2.2 Hz, HAr), 6.94 (d, 2H, J
) 8.8 Hz, HAr), 7.66 (d, 2H, J ) 8.8 Hz, HAr), 8.15 (s, 1H, d
CH). 13C NMR (62.90 MHz, CDCl3): δ 15.3, 23.6, 41.8, 55.4,
55.9, 56.0, 89.6, 93.1, 106.4, 113.6 (2), 119.5, 126.7, 129.6, 130.1
(2), 130.8, 140.7, 157.9, 159.3, 161.2, 162.8. MS: m/z 379 (M+
+ 1).
N,N-Dim eth yl-2-[5,7-d im eth oxy-3-(4-m eth oxyp h en yl)-
2-qu in olyl]oxy-1-eth a n a m in e (6a ). Yield, 30%; mp 49-50
1
°C (Et2O). IR (KBr): ν 1621, 1584 cm-1. H NMR (250 MHz,
CDCl3): δ 2.31 (s, 6H, NCH3), 2.77 (t, 2H, J ) 6.0 Hz, CH2),
3.85 (s, 3H, OCH3), 3.93 (s, 6H, OCH3), 4.62 (t, 2H, J ) 6.0
Hz, CH2), 6.39 (d, 1H, J ) 2.0 Hz, HAr), 6.81 (d, 1H, J ) 2.0
Hz, HAr), 6.94 (d, 2H, J ) 8.8 Hz, HAr), 7.58 (d, 2H, J ) 8.8
Hz, HAr), 8.25 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ
45.7 (2), 55.3, 55.5 (2), 57.8, 63.8, 95.9, 98.5, 112.9, 113.4 (2),
122.0, 129.5, 130.6 (2), 132.3, 147.8, 156.3, 158.9, 160.0, 161.3.
MS: m/z 383 (M+ + 1). Anal. (C22H26N2O4) C, H, N.
2-([5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-qu in olin yl]-
oxy)bu ta n en itr ile (9a ). Yield, 33%; mp 89-90 °C (EtOAc).
IR (KBr): ν 2247, 1624, 1607 cm-1 1H NMR (250 MHz,
.
CDCl3): δ 2.12-2.22 (m, 2H, CH2), 2.50 (t, 2H, J ) 7.5 Hz,
CH2), 3.87 (s, 3H, OCH3), 3.94 (s, 6H, OCH3), 4.61 (t, 2H, J )
7.5 Hz, CH2), 6.40 (d, 1H, J ) 2.2 Hz, HAr), 6.81 (d, 1H, J )
2.2 Hz, HAr), 6.97 (d, 2H, J ) 8.8 Hz, HAr), 7.53 (d, 2H, J ) 8.8
Hz, HAr), 8.27 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ
14.6, 25.4, 55.4, 55.7, 55.8, 63.6, 96.2, 98.6, 113.1, 113.7 (2),
119.5, 122.0, 129.5, 130.5 (2), 132.8, 147.9, 156.4, 159.1, 159.7,
161.6. MS: m/z 379 (M+ + 1). Anal. (C22H22N2O4) C, H, N.
1-[3-(Dim eth yla m in o)pr op yl]-5,7-dim eth oxy-3-(4-m eth -
oxyp h en yl)-1,2-d ih yd r o-2-qu in olin on e (7). According to
the procedure described for 3, the 2-quinolone 7 and compound
7a were prepared starting from 2e (1 equiv) and 3-(dimethyl-
amino)propyl chloride (2.2 equiv). Reaction time, 3 h; eluent
chromatography Et2O/MeOH 8:2 and then CH2Cl2/MeOH 9:1;
Ben zyl 2-[5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-oxo-
1,2-d ih yd r o-1-qu in olin yl]a ceta te (10). According to the
procedure described for 3, the 2-quinolone 10 and compound
10a were prepared starting from 2e (1 equiv) and benzyl
bromoacetate (2 equiv). Reaction time, 2 h; eluent chromatog-
1
yield, 70%. IR (KBr): ν 1635, 1617, 1598 cm-1. H NMR (250
MHz, CDCl3): δ 1.91-2.04 (m, 2H, CH2), 2.28 (s, 6H, NCH3),
2.46 (t, 2H, J ) 7.2 Hz, CH2), 3.83 (s, 3H, OCH3), 3.91 (s, 3H,
OCH3), 3.93 (s, 3H, OCH3), 4.36 (t, 2H, J ) 7.2 Hz, CH2), 6.29
(d, 1H, J ) 2.0 Hz, HAr), 6.54 (d, 1H, J ) 2.0 Hz, HAr), 6.94 (d,
2H, J ) 8.8 Hz, HAr), 7.68 (d, 2H, J ) 8.8 Hz, HAr), 8.13 (s,
1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 25.5, 41.8, 45.6
(2), 55.4, 55.5, 55.8, 57.1, 90.3, 92.6, 106.4, 113.5 (2), 126.9,
130.0, 130.1 (2), 130.3, 141.1, 157.6, 159.1, 162.0, 162. MS: m/z
397 (M+ + 1).
raphy CH2Cl2; yield, 72%. IR (KBr): 1748, 1642, 1617 cm-1
.
1H NMR (250 MHz, CDCl3): δ 3.68 (s, 3H, OCH3), 3.84 (s, 3H,
OCH3), 3.92 (s, 3H, OCH3), 5.16 (s, 2H, CH2), 5.21 (s, 2H, CH2),
6.03 (d, 1H, J ) 2.0 Hz, HAr), 6.27 (d, 1H, J ) 2.0 Hz, HAr),
6.94 (d, 2H, J ) 7.5 Hz, HAr), 7.25-7.32 (m, 5H, HAr), 7.68 (d,
2H, J ) 7.5 Hz, HAr), 8.17 (s, 1H, dCH). 13C NMR (62.90 MHz,
CDCl3): δ 44.7, 55.3, 55.4, 55.8, 67.1, 89.8, 93.0, 106.2, 113.5
(2), 126.5, 128.3 (2), 128.4, 128.5 (2), 129.5, 130.1 (2), 131.1,
135.3, 141.0, 157.8, 159.2, 161.8, 162.4, 168.3. MS: m/z 460
(M+ + 1).
N,N-Dim eth yl-2-[5,7-d im eth oxy-3-(4-m eth oxyp h en yl)-
2-qu in olyl]oxy-1-p r op a n a m in e (7a ). Yield, 25%; mp 54-
55 °C (Et2O). IR (KBr): ν 1621, 1584 cm-1. 1H NMR (250 MHz,
CDCl3): δ 1.96-2.07 (m, 2H, CH2), 2.26 (s, 6H, NCH3), 2.47
(t, 2H, J ) 6.5 Hz, CH2), 3.84 (s, 3H, OCH3), 3.92 (s, 6H, OCH3),
4.53 (t, 2H, J ) 6.5 Hz, CH2), 6.39 (d, 1H, J ) 2.2 Hz, HAr),
6.82 (d, 1H, J ) 2.2 Hz, HAr), 6.96 (d, 2H, J ) 8.8 Hz, HAr),
7.57 (d, 2H, J ) 8.8 Hz, HAr), 8.26 (s, 1H, dCH). 13C NMR
(62.90 MHz, CDCl3): δ 27.0, 45.4 (2), 55.2, 55.5 (2), 56.6, 64.2,
95.8, 98.5, 112.7, 113.4 (2), 121.9, 129.6, 130.5 (2), 132.1, 147.9,
156.2, 158.8, 160.2, 161.2. MS: m/z 397 (M+ + 1). Anal.
(C23H28N2O4) C, H, N.
Ben zyl 2-([5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-qu in -
olin yl]oxy)a ceta te (10a ). Yield, 20%; mp 114-115 °C (Et2O).
IR (KBr): ν 1761, 1621, 1517 cm-1 1H NMR (250 MHz,
.
CDCl3): δ 3.87 (s, 3H, OCH3), 3.91 (s, 3H, OCH3), 3.95 (s, 3H,
OCH3), 5.17 (s, 2H, CH2), 5.27 (s, 2H, CH2), 6.44 (d, 1H, J )
2.0 Hz, HAr), 6.76 (d, 1H, J ) 2.0 Hz, HAr), 7.00 (d, 2H, J ) 8.0
Hz, HAr), 7.26-7.38 (m, 5H, HAr), 7.71 (d, 2H, J ) 8.0 Hz, HAr),
8.36 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 55.2, 55.4,
55.5, 62.6, 66.4, 96.2, 98.5, 113.4, 113.6 (2), 121.6, 128.0 (2),
128.4 (3), 129.0, 130.5 (2), 132.7, 135.6, 147.2, 156.1, 158.6,
159.0, 161.3, 169.3. MS: m/z 460 (M+ + 1). Anal. (C27H25NO6)
C, H, N.
[5,7-Dim eth oxy-3-(4-m eth oxyph en yl)-2-oxo-1,2-dih ydr o-
1-qu in olin yl]a ceton itr ile (8). According to the procedure
described for 3, the 2-quinolone 8 and compound 8a were
prepared starting from 2e (1 equiv) and bromoacetonitrile (2
equiv). Reaction time, 3 h; eluent chromatography CH2Cl2/
2-[5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-oxo-1,2-d ih y-
d r o-1-qu in olin yl]a cetic Acid (11). A solution of 10 (1.0 g,
2.2 mmol) in dry 1,4-dioxane (30 mL) was stirred under 40
psi of hydrogen for 4 h at room temperature in the presence
of 10% Pd-C (100 mg) and then filtered. The filtrate was
evaporated to dryness leaving a solid that was washed with
Et2O to give 11 (764 mg, 95%). IR (KBr): ν 1732, 1614, 1583
1
EtOAc 7:3; yield, 61%. IR (KBr): ν 2216, 1660, 1607 cm-1. H
NMR (250 MHz, CDCl3): δ 3.84 (s, 3H, OCH3), 3.94 (s, 3H,
OCH3), 3.95 (s, 3H, OCH3), 5.29 (s, 2H, CH2), 6.36 (s, 2H, HAr),
6.95 (d, 2H, J ) 8.8 Hz, HAr), 7.65 (d, 2H, J ) 8.8 Hz, HAr),
8.17 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 30.4, 55.3,
55.8, 56.0, 90.0, 93.5, 106.2, 113.6 (2), 114.8, 126.3, 129.0, 130.0
(2), 131.7, 139.8, 158.1, 159.4, 161.1, 163.0. MS: m/z 351 (M+
+ 1).
cm-1 1H NMR (250 MHz, DMSO-d6 + D2O): δ 3.77 (s, 3H,
.
OCH3), 3.85 (s, 3H, OCH3), 3.91 (s, 3H, OCH3), 5.02 (s, 2H,
CH2), 6.46 (s, 1H, HAr), 6.47 (s, 1H, HAr), 6.93 (d, 2H, J ) 9.0
Hz, HAr), 7.62 (d, 2H, J ) 9.0 Hz, HAr), 8.03 (s, 1H, dCH). 13C
NMR (62.90 MHz, DMSO-d6): δ 44.9, 55.5, 56.1, 56.6, 91.3,
93.3, 105.3, 113.8 (2), 125.7, 129.4, 130.1 (2), 130.4, 141.3,
157.6, 159.1, 161.3, 162.8, 170.1. MS: m/z 370 (M+ + 1).
2-([5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-qu in olin yl]-
oxy)a ceton itr ile (8a ). Yield, 30%; mp 149-150 °C (Et2O).
1
IR (KBr): ν 1623, 1586 cm-1. H NMR (250 MHz, CDCl3): δ
3.87 (s, 3H, OCH3), 3.95 (s, 6H, OCH3), 5.17 (s, 2H, CH2), 6.45
(d, 1H, J ) 2.0 Hz, HAr), 6.87 (d, 1H, J ) 2.0 Hz, HAr), 7.99 (d,
2H, J ) 9.0 Hz, HAr), 7.54 (d, 2H, J ) 9.0 Hz, HAr), 8.34 (s,
1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 50.1, 55.3, 55.6,
55.7, 96.9, 98.6, 113.8 (2), 113.9, 116.1, 121.3, 128.4, 130.5 (2),
Met h yl 3-[5,7-Dim et h oxy-3-(4-h yd r oxyp h en yl)-2-oxo-
1,2-d ih yd r o-1-qu in olin yl]p r op a n oa te (12). To a solution of
2e (1.00 g, 3.2 mmol) and methyl acrylate (2.88 mL, 32.0 mmol)
in anhydrous DMF (10 mL) at 0 °C was added Triton B (0.06
mL, 0.33 mmol). The final solution was stirred for 18 h at room
temperature, and then, the solvent was removed in vacuo. The
crude residue was partitioned between H2O and EtOAc and
extracted. The organic layer was washed with H2O, dried over
MgSO4, and concentrated in vacuo. The crude residue was
133.5, 147.1, 156.3, 157.2, 129.3, 161.8. MS: m/z 351 (M+
+
1). Anal. (C20H18N2O4) C, H, N.
3-[5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-oxo-1,2-d ih y-
d r o-1-qu in olin yl]bu ta n en itr ile (9). According to the pro-

2552 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12
J oseph et al.
purified by column chromatography (eluent CH2Cl2/EtOAc 8:2)
to give 12 (1.06 g, 83%). IR (KBr): ν 1725, 1638 cm-1. 1H NMR
(250 MHz, CDCl3): δ 2.78 (t, 2H, J ) 8.0 Hz, CH2), 3.68 (s,
3H, CH3), 3.80 (s, 3H, OCH3), 3.88 (s, 6H, OCH3), 4.57 (t, 2H,
J ) 8.0 Hz, CH2), 6.26 (d, 1H, J ) 2.0 Hz, HAr), 6.46 (d, 1H, J
) 2.0 Hz, HAr), 6.92 (d, 2H, J ) 8.8 Hz, HAr), 7.66 (d, 2H, J )
8.8 Hz, HAr), 8.11 (s, 1H, dCH). 13C NMR (62.90 MHz,
CDCl3): δ 31.9, 39.1, 51.8, 55.2, 55.5, 55.7, 89.8, 92.6, 106.2,
113.4 (2), 126.5, 129.5, 129.9 (2), 130.3, 140.5, 157.6, 159.0,
161.7, 162.4, 172.0. MS: m/z 398 (M+ + 1).
hexylcarbodiimide (540 mg, 2.6 mmol) at 0 °C. After the
mixture was stirred at 0 °C for 10 min, N,N-diethylamine (0.26
mL, 2.6 mmol) was added and the final mixture was stirred
at 0 °C for 2 h and at room temperature for 24 h. The white
precipitate of dicyclohexylurea was removed by filtration. The
solvent was evaporated in vacuo. The crude residue was
purified by column chromatography (eluent CH2Cl2/EtOAc 7:3)
1
to afford 17 (680 mg, 60%). IR (KBr): ν 1636, 1617 cm-1. H
NMR (250 MHz, CDCl3): δ 1.11 (t, 3H, J ) 7.0 Hz, CH3), 1.13
(t, 3H, J ) 7.0 Hz, CH3), 2.78 (t, 2H, J ) 8.0 Hz, CH2), 3.30 (q,
2H, J ) 7.0 Hz, CH2), 3.39 (q, 2H, J ) 7.0 Hz, CH2), 3.84 (s,
3H, OCH3), 3.91 (s, 3H, OCH3), 3.94 (s, 3H, OCH3), 4.65 (t,
2H, J ) 8.0 Hz, CH2), 6.29 (d, 1H, J ) 2.0 Hz, HAr), 6.73 (d,
1H, J ) 2.0 Hz, HAr), 6.94 (d, 2H, J ) 7.5 Hz, HAr), 7.67 (d,
2H, J ) 7.5 Hz, HAr), 8.15 (s, 1H, dCH). 13C NMR (62.90 MHz,
CDCl3): δ 13.1, 14.4, 31.0, 40.1, 40.5, 42.2, 55.3, 55.8 (2), 89.9,
93.1, 106.3, 113.5 (2), 126.6, 129.7, 130.0 (2), 130.5, 140.9,
157.6, 159.1, 162.1, 162.6, 169.9. MS: m/z 439 (M+ + 1).
3-[5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-oxo-1,2-d ih y-
d r o-1-qu in olin yl]p r op a n en itr ile (13). According to the
procedure described for 12, the 2-quinolone 13 was prepared
starting from 2e (1 equiv) and acrylonitrile (10 equiv). Reaction
time, 2 h; eluent chromatography CH2Cl2/EtOAc 7:3; yield,
63%. IR (KBr): ν 2241, 1639 cm-1 1H NMR (250 MHz,
.
CDCl3): δ 2.88 (t, 2H, J ) 7.1 Hz, CH2), 3.84 (s, 3H, OCH3),
3.93 (s, 3H, OCH3), 3.94 (s, 3H, OCH3), 4.59 (t, 2H, J ) 7.1
Hz, CH2), 6.33 (d, 1H, J ) 1.8 Hz, HAr), 6.45 (d, 1H, J ) 1.8
Hz, HAr), 6.95 (d, 2H, J ) 9.0 Hz, HAr), 7.66 (d, 2H, J ) 9.0
Hz, HAr), 8.17 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ
15.9, 39.4, 55.3, 55.7, 55.9, 89.9, 93.0, 106.4, 113.6 (2), 117.7,
126.6, 129.2, 130.0 (2), 131.1, 140.5, 158.0, 159.3, 161.8, 162.7.
MS: m/z 365 (M+ + 1).
8,10-Dim e t h oxy-6-(4-m e t h oxyp h e n yl)-2,3-d ih yd r o-
1H,5H-p yr id o[3,2,1-ij]qu in olin e-1,5-d ion e (18). In a dry
round flask, P2O5 (63 mg) and PPA (500 mg) were stirred at
110 °C until complete dissolution of P2O5. Acid 15 (100 mg,
0.26 mmol) was added, and the final mixture was stirred at
110 °C for 45 min. After it was cooled, the mixture was poured
into ice and neutralized by 2 N NaOH. CH2Cl2 (200 mL) was
added, and the mixture was stirred overnight. After extraction,
the aqueous layer was extracted with CH2Cl2 twice (2 × 50
mL). The organic phase was dried over MgSO4 and evaporated
in vacuo. The crude residue was purified by column chroma-
tography (eluent CH2Cl2/MeOH 98:2) to give 18 (62 mg, 65%).
Ben zyl 3-[5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-oxo-
1,2-d ih yd r o-1-qu in olin yl]p r op a n oa te (14). According to the
procedure described for 12, the 2-quinolone 14 was prepared
starting from 2e (1 equiv) and benzyl acrylate (10 equiv).
Reaction time, 18 h; eluent chromatography EtOAc/PE 4:6;
1
yield, 88%. IR (KBr): ν 1731, 1635, 1600 cm-1. H NMR (250
IR (KBr): ν 1678, 1649, 1634 cm-1 1H NMR (250 MHz,
.
CDCl3): δ 2.83 (t, 2H, J ) 7.0 Hz, CH2), 3.85 (s, 3H, OCH3),
4.04 (s, 6H, OCH3), 4.54 (d, 2H, J ) 7.0 Hz, CH2), 6.32 (s, 1H,
MHz, CDCl3): δ 2.86 (t, 2H, J ) 8.0 Hz, CH2), 3.84 (s, 3H,
OCH3), 3.85 (s, 3H, OCH3), 3.92 (s, 3H, OCH3), 4.63 (t, 2H, J
) 8.0 Hz, CH2), 5.14 (s, 2H, CH2), 6.29 (d, 1H, J ) 2.0 Hz,
H
Ar), 6.96 (d, 2H, J ) 7.5 Hz, HAr), 7.68 (d, 2H, J ) 7.5 Hz,
H
H
Ar), 6.49 (d, 1H, J ) 2.0 Hz, HAr), 6.94 (d, 2H, J ) 7.5 Hz,
Ar), 7.30-7.35 (m, 5H, HAr), 7.68 (d, 2H, J ) 7.5 Hz, HAr),
HAr), 8.12 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 37.6,
40.3, 55.3, 56.1, 56.5, 88.9, 103.5, 104.8, 113.6 (2), 127.6, 129.0,
129.8, 130.0 (2), 142.9, 159.4, 161.6, 161.7, 163.7, 190.3. MS:
m/z 366 (M+ + 1).
8.13 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 32.2, 39.1,
55.2, 55.4, 55.7, 66.5, 89.7, 92.6, 106.2, 113.4 (2), 126.5, 128.1
(2), 128.2, 128.4 (2), 129.5, 129.9 (2), 130.4, 135.5, 140.5, 157.6,
159.0, 161.7, 162.4, 171.3. MS: m/z 474 (M+ + 1).
5,7-Dih ydr oxy-3-(4-h ydr oxyph en yl)-1,2-dih ydr o-2-qu in -
olin on e (19).21 To a solution of 2e (1.0 g, 3.2 mmol) in AcOH
(15 mL) was dropwise added 48% HBr in H2O (5 mL). The
final solution was stirred at reflux for 3 days. The cooled
mixture was diluted with H2O, neutralized with 10% aqueous
NaOH (pH 6-7), and extracted with CH2Cl2 twice. The organic
layer was dried over MgSO4 and then evaporated in vacuo.
The crude residue was purified by column chromatography
3-[5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-2-oxo-1,2-d ih y-
d r o-1-qu in olin yl]p r op a n oic Acid (15). A solution of 14 (1.0
g, 2.1 mmol) in dry 1,4-dioxane (30 mL) was stirred under 40
psi of hydrogen for 48 h at room temperature in the presence
of 10% Pd-C (100 mg) and then filtered. The filtrate was
evaporated to dryness leaving a solid that was washed with
Et2O to give 15 (780 mg, 97%). IR (KBr): ν 1724, 1637, 1612,
1
1
1604 cm-1. H NMR (250 MHz, DMSO-d6 + D2O): δ 2.60 (t,
(eluent CH2Cl2/MeOH 9:1) to give 19 (320 mg, 37%). H NMR
(250 MHz, DMSO-d6): δ 6.11 (d, 1H, J ) 2.0 Hz, HAr), 6.18 (d,
1H, J ) 2.0 Hz, HAr), 6.76 (d, 2H, J ) 8.6 Hz, HAr), 7.52 (d,
2H, J ) 8.6 Hz, HAr), 7.89 (s, 1H, dCH), 9.42 (s, 1H, OH), 9.84
(s, 1H, OH), 10.21 (s, 1H, OH), 11.46 (br s, 1H, NH). 13C NMR
(62.90 MHz, DMSO-d6): δ 91.3, 96.4, 103.5, 114.7 (2), 125.1,
127.8, 129.4 (2), 130.4, 140.7, 155.3, 156.6, 160.1, 161.8. MS:
m/z 270 (M+ + 1).
5,7-Dih yd r oxy-3-(4-h yd r oxyp h en yl)-1-m eth yl-1,2-d ih y-
d r o-2-qu in olin on e (20). According to the procedure described
for 19, the 2-quinolone 20 was prepared starting from 3 (eluent
CH2Cl2/MeOH 9:1); yield: 35%. 1H NMR (250 MHz, DMSO-
d6): δ 3,53 (s, 3H, NCH3), 6.25 (s, 2H, HAr), 6.76 (d, 2H, J )
8.8 Hz, HAr), 7.47 (d, 2H, J ) 8.8 Hz, HAr), 7.92 (s, 1H, dCH),
9.42 (s, 1H, OH), 10.00 (s, 1H, OH), 10.35 (s, 1H, OH). 13C
NMR (62.90 MHz, DMSO-d6): δ 29.7, 91.8, 96.5, 103.5, 114.7
(2), 124.3, 128.4, 129.7 (3), 141.7, 155.9, 156.7, 160.6, 161.1.
MS: m/z 284 (M+ + 1).
2H, J ) 7.5 Hz, CH2), 3.78 (s, 3H, OCH3), 3.90 (s, 3H, OCH3),
3.92 (s, 3H, OCH3), 4.48 (t, 2H, J ) 7.5 Hz, CH2), 6.48 (d, 1H,
J ) 1.8 Hz, HAr), 6.62 (d, 1H, J ) 1.8 Hz, HAr), 6.95 (d, 2H, J
) 8.8 Hz, HAr), 7.62 (d, 2H, J ) 8.8 Hz, HAr), 8.00 (s, 1H, d
CH). 13C NMR (62.90 MHz, DMSO-d6): δ 32.0, 38.9, 55.1, 55.7,
56.1, 90.6, 93.0, 105.0, 113.3 (2), 125.6, 129.3, 129.5, 129.8 (2),
140.5, 157.2, 158.7, 160.6, 162.4, 172.5. MS: m/z 384 (M+
+
1).
1-[2-(1H-1,2,3,4-Tetr a zol-5-yl)eth yl]-5,7-d im eth oxy-3-(4-
m eth oxyp h en yl)-1,2-d ih yd r o-2-qu in olin on e (16). A solu-
tion of 13 (350 mg, 0.9 mmol) and tributyltin azide (0,42 mL,
1.53 mmol) in anhydrous toluene (20 mL) was stirred at 105
°C for 65 h. After it was cooled, the solvent was removed in
vacuo. The crude residue was purified by column chromatog-
raphy (eluent CH2Cl2/MeOH 9:1) to give 16 (333 mg, 85%). IR
1
(KBr): ν 1618, 1594 cm-1. H NMR (250 MHz, DMSO-d6): δ
3.33 (t, 2H, J ) 6.0 Hz, CH2), 3.79 (s, 3H, OCH3), 3.89 (s, 3H,
OCH3), 3.93 (s, 3H, OCH3), 4.67 (t, 2H, J ) 6.0 Hz, CH2), 6.48
(s, 1H, HAr), 6.52 (s, 1H, HAr), 6.96 (d, 2H, J ) 9.0 Hz, HAr),
7.59 (d, 2H, J ) 9.0 Hz, HAr), 8.01 (s, 1H, dCH). 13C NMR
(62.90 MHz, DMSO-d6): δ 21.4, 40.8, 55.1, 55.6, 56.1, 90.4,
93.0, 105.0, 113.3 (2), 125.5, 129.3, 129.6, 129.7 (2), 140.4,
153.7, 157.2, 158.7, 160.7, 162.4. MS: m/z 408 (M+ + 1).
5,7-Dim eth oxy-3-(4-h ydr oxyph en yl)-1,2-dih ydr o-2-qu in -
olin on e (21). To a solution of 2e (530 mg, 1.70 mmol) in AcOH
(15 mL) was dropwise added 48% HBr in H2O (2.7 mL). The
final solution was stirred at reflux for 5 h. The cooled mixture
was diluted with H2O, neutralized with 10% aqueous NaOH
(pH 6-7), and extracted with CH2Cl2 twice. The organic layer
was dried over MgSO4 and then evaporated in vacuo. The
crude residue was purified by column chromatography (eluent
CH2Cl2/MeOH 9:1) to give 21 (175 mg, 35%). IR (KBr): ν 1628,
N,N-Dieth yl-3-[5,7-d im eth oxy-3-(4-m eth oxyp h en yl)-2-
oxo-1,2-d ih yd r o-1-qu in olin yl]p r op a n a m id e (17). To a so-
lution of 15 (1.0 g, 2.6 mmol) in anhydrous DMF (25 mL) were
added hydroxybenzotriazole (353 mg, 2.6 mmol) and dicyclo-
1604, 1558 cm-1 1H NMR (250 MHz, DMSO-d6): δ 3.80 (s,
.

3-Aryl-2-Quinolone Derivatives
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12 2553
3H, OCH3), 3.89 (s, 3H, OCH3), 6.35 (d, 1H, J ) 2.5 Hz, HAr),
6.43 (d, 1H, J ) 2.5 Hz, HAr), 6.78 (d, 2H, J ) 7.5 Hz, HAr),
7.55 (d, 2H, J ) 7.5 Hz, HAr), 7.92 (s, 1H, dCH), 9.48 (s, 1H,
OH), 11.72 (br s, 1H, NH). 13C NMR (62.90 MHz, DMSO-d6):
δ 55.4, 55.9, 90.0, 93.0, 104.7, 114.8 (2), 126.9, 127.4, 129.6,
129.8 (2), 140.4, 156.6, 156.9, 161.6, 161.8. MS: m/z 298 (M+
+ 1).
domethane (3.5 mL, 56 mmol) diluted in anhydrous THF (5
mL) at room temperature. The final solution was stirred
overnight at room temperature. The red precipitate obtained
was collected and washed with the same solvent to provide
1
27 (761 mg, 79%); mp 156-157 °C (THF washing). H NMR
(250 MHz, CDCl3): δ 2.44 (s, 3H, SCH3), 3.88 (s, 3H, OCH3),
4.02 (s, 3H, OCH3), 4.28 (s, 3H, OCH3), 5.03 (s, 3H, NCH3),
6.70 (d, 1H, J ) 1.5 Hz, HAr), 7.03 (d, 2H, J ) 8.5 Hz, HAr),
7.37 (br s, 1H, HAr), 7.45 (d, 2H, J ) 8.5 Hz, HAr), 8.71 (s, 1H,
dCH). 13C NMR (62.90 MHz, CDCl3): δ 20.8, 46.5, 55.4, 56.7,
58.5, 92.9, 101.0, 114.4 (2), 117.6, 125.8, 128.9, 130.7 (2), 135.9,
138.8, 144.2, 157.4, 160.3, 167.7.
5,7-Dim eth oxy-3-(4-h yd r oxyp h en yl)-1-m eth yl-1,2-d ih y-
d r o-2-qu in olin on e (22). According to the procedure described
for 21, the 2-quinolone 22 was prepared starting from 3 (eluent
CH2Cl2/EtOAc 7:3); yield, 56%. 1H NMR (250 MHz, DMSO-
d6): δ 3.63 (s, 3H, NCH3), 3.89 (s, 3H, OCH3), 3.90 (s, 3H,
OCH3), 6.46 (d, 1H, J ) 1.9 Hz, HAr), 6.54 (d, 1H, J ) 1.9 Hz,
5,7-Dim et h oxy-3-(4-m et h oxyp h en yl)-1-m et h yl-1,2-d i-
h yd r o-2-qu in olin on e-2-p h en ylh yd r a zon e (28). In a sealed
tube, a solution of 27 (200 mg, 0.4 mmol) and phenylhydrazine
(0.28 mL, 2.8 mmol) in anhydrous EtOH (5 mL) was heated
at 90 °C overnight. After it was cooled, the solvent was
removed in vacuo. The residue was dissolved in CH2Cl2 (10
mL), and the organic solution was washed with H2O and then
with saturated NaHCO3 solution twice. The organic phase was
dried over MgSO4 and evaporated in vacuo. The crude residue
was crystallized from MeOH to afford 28 (102 mg, 60%). IR
H
H
Ar), 6.76 (d, 2H, J ) 8.5 Hz, HAr), 7.48 (d, 2H, J ) 8.5 Hz,
Ar), 7.93 (s, 1H, dCH), 9.47 (s, 1H, OH). 13C NMR (62.90
MHz, DMSO-d6): δ 30.5, 56.1, 56.5, 91.3, 93.4, 105.3, 115.2
(2), 126.5, 128.4, 129.2, 130.2 (2), 141.7, 157.4 (2), 161.4, 162.5.
MS: m/z 312 (M+ + 1).
5,7-Dim eth oxy-3-(4-m eth oxyph en yl)-1,2-dih ydr o-2-qu in -
olin eth ion e (23). A mixture of 2e (100 mg, 0.32 mmol) and
Lawesson’s reagent (260 mg, 0.64 mmol) in anhydrous toluene
(15 mL) was stirred at reflux for 18 h. After it was cooled, the
solvent was removed in vacuo and the residue was purified
by silica gel chromatography (eluent CH2Cl2/EtOAc 9:1) to give
23 (81 mg, 77%). IR (KBr): ν 1636, 1610, 1524 cm-1. 1H NMR
(250 MHz, DMSO-d6): δ 3.78 (s, 3H, OCH3), 3.83 (s, 3H,
OCH3), 3.89 (s, 3H, OCH3), 6.49 (d, 1H, J ) 1.9 Hz, HAr), 6.79
(d, 1H, J ) 1.9 Hz, HAr), 6.92 (d, 2H, J ) 8.7 Hz, HAr), 7.49 (d,
2H, J ) 8.7 Hz, HAr), 7.78 (s, 1H, dCH), 13.50 (br s, 1H, NH).
13C NMR (62.90 MHz, CDCl3): δ 55.1, 55.6, 56.1, 90.2, 95.2,
108.9, 112.9 (2), 128.2, 130.7 (2), 132.0, 136.3, 140.9, 156.4,
158.5, 162.5, 180.5. MS: m/z 328 (M+ + 1).
1
(KBr): ν 3340, 1602, 1599 cm-1. H NMR (250 MHz, CDCl3):
δ 3.58 (s, 3H, NCH3), 3.85 (s, 3H, OCH3), 3.86 (s, 3H, OCH3),
3.88 (s, 3H, OCH3), 6.04 (s, 1H, HAr), 6.14 (s, 1H, HAr), 6.48
(br s, 1H, NH), 6.56-6.67 (m, 3H, HAr), 6.94 (d, 2H, J ) 8.8
Hz, HAr), 7.11 (t, 2H, J ) 7.7 Hz, HAr), 7.28 (s, 1H, dCH), 7.28-
7.34 (m, 2H, HAr). 13C NMR (62.90 MHz, CDCl3): δ 33.3, 55.4
(2), 55.6, 89.6 (2), 111.6, 113.8 (2), 117.8, 125.9, 128.3, 128.9
(3), 129.5 (2), 130.5, 146.2, 157.2, 159.2 (2), 162.3. MS: m/z
416 (M+ + 1).
5,7-Dim et h oxy-3-(4-m et h oxyp h en yl)-1-m et h yl-1,2-d i-
h yd r o-2-qu in olin on e-2-(2-p yr id in yl)h yd r a zon e (29). Com-
pound 29 was obtained according to the procedure described
below but substituting the phenylhydrazine by 2-hydrazinopy-
5,7-Dim et h oxy-3-(4-m et h oxyp h en yl)-1-m et h yl-1,2-d i-
h yd r o-2-qu in olin eth ion e (24). According to the procedure
described for 23, the 2-thioquinolone 24 was prepared starting
from 3 (eluent EtOAc/PE 3:7); yield, 72%. IR (KBr): ν 1613,
1
ridine; yield, 58%. IR (KBr): 3353, 1628, 1593 cm-1. H NMR
1
1570, 1512 cm-1. H NMR (250 MHz, CDCl3): δ 3.84 (s, 3H,
(250 MHz, CDCl3): δ 3.61 (s, 3H, NCH3), 3.87 (s, 6H, OCH3),
3.90 (s, 3H, OCH3), 6.07 (s, 1H, HAr), 6.17 (s, 1H, HAr), 6.51-
6.55 (m, 1H, HPyr), 6.94-7.01 (m, 3H, HAr + HPyr), 7.14 (br s,
1H, NH), 7.32-7.36 (m, 3H, dCH + HAr) 7.48 (t, 1H, J ) 7.3
Hz, HPyr), 7.91 (d, 1H, J ) 4.3 Hz, HPyr). 13C NMR (62.90 MHz,
CDCl3): δ 33.2, 55.3, 55.4, 55.6, 89.7 (2), 104.7, 105.7, 113.4,
114.1 (2), 122.9, 129.0, 129.2 (2), 130.1, 137.5, 140.7, 143.7,
147.8, 157.3, 157.4, 159.2, 162.4. MS: m/z 417 (M+ + 1).
OCH3), 3.91 (s, 3H, OCH3), 3.95 (s, 3H, OCH3), 4.39 (s, 3H,
NCH3), 6.39 (d, 1H, J ) 2.0 Hz, HAr), 6.56 (d, 1H, J ) 2.0 Hz,
H
H
Ar), 6.93 (d, 2H, J ) 7.5 Hz, HAr), 7.43 (d, 2H, J ) 7.5 Hz,
Ar), 7.97 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 39.5,
55.2, 55.6, 55.9, 91.1, 94.6, 109.9, 113.1 (2), 126.6, 130.8 (2),
134.3, 138.6, 142.7, 157.6, 158.8, 162.7, 184.5. MS: m/z 342
(M+ + 1).
P h a r m a cology. In Vitr o MTT Color im etr ic Assa y.
Twelve human tumor cell lines were obtained from the
American Type Culture Collection (ATCC, Manassas, VA).
These included three glioblastomas (A-172, U-373 MG, and
U-87 MG), two colon (HCT-15 and LoVo), two nonsmall cell
lung (A549 and A-427), two bladder (J 82 and T24), one
prostate (PC-3), and two breast (T-47D and MCF-7) cancer
models. The ATCC numbers of these cell lines are CRL1620
(A-172), HTB 14 (U-87 MG), HTB 17 (U-373 MG), CCL225
(HCT-15), CCL229 (LoVo), CCL 185 (A549), HBT 53 (A-427),
HTB1 (J 82), HTB4 (T24), HTB133 (T-47D), HTB22 (MCF-7),
and CRL1435 (PC-3). The cells were cultured at 37 °C in sealed
(airtight) Falcon plastic dishes (Nunc, Gibco, Belgium) con-
taining Eagle’s minimal essential medium (MEM, Gibco)
supplemented with 5% fetal calf serum (FCS). All of the media
were supplemented with a mixture of 0.6 mg/mL glutamine
(Gibco), 200 IU/mL penicillin (Gibco), 200 IU/mL streptomycin
(Gibco), and 0.1 mg/mL gentamycin (Gibco). The FCS was
heat-inactivated for 1 h at 56 °C.
The twelve cell lines were incubated for 24 h in 96 microwell
plates (at a concentration of 40 000 cells/mL culture medium)
to ensure adequate plating prior to cell growth determination,
which was carried out by means of the colorimetric MTT assay
as detailed previously.23 Six concentrations ranging from 10-5
to 10-9 M were assayed for each of the 25 drugs under study.
In Vitr o Cell Migr a tion Assa y. The cell motility level of
the MCF-7 human breast cancer cells was quantitatively
determined by means of a device detailed elsewhere.24 Briefly,
this device consists of an inverted-phase contrast microscope
equipped with a black and white CCD camera, an incubator
5,7-Acet yl-3-(4-a cet ylp h en yl)-1,2-d ih yd r o-2-q u in oli-
n on e (25). A solution of 20 (200 mg, 0.71 mmol), anhydride
acetic, and pyridine (8 mL, v/v) was stirred at room temper-
ature for 18 h. H2O (10 mL) was added to the mixture and
then extracted with CH2Cl2 (2 × 10 mL). The organic layer
was dried over MgSO4 and then evaporated in vacuo. The
crude residue was purified by column chromatography (eluent
CH2Cl2/EtOAc 9:1) to give 25 (220 mg, 76%). IR (KBr): ν 1769,
1748, 1638, 1598 cm-1. 1H NMR (250 MHz, CDCl3): δ 2.31 (s,
3H, CH3), 2.33 (s, 3H, CH3), 2.40 (s, 3H, CH3), 3.72 (s, 3H,
NCH3), 6.92 (d, 1H, J ) 2.0 Hz, HAr), 7.03 (d, 1H, J ) 2.0 Hz,
H
H
Ar), 7.14 (d, 2H, J ) 8.5 Hz, HAr), 7.67 (d, 2H, J ) 8.5 Hz,
Ar), 7.78 (s, 1H, dCH). 13C NMR (62.90 MHz, CDCl3): δ 20.8,
21.0, 21.2, 30.5, 105.2, 110.2, 111.8, 121.4 (2), 129.7, 130.3 (2),
131.5, 134.2, 141.0, 148.8, 150.7, 151.9, 161.2, 168.6, 168.8,
169.6. MS: m/z 410 (M+ + 1).
5,7-Dim eth oxy-3-(4-a cetylp h en yl)-1,2-d ih yd r o-2-qu in o-
lin on e (26). According to the procedure described for 25, the
2-quinolone 26 was prepared starting from 22. Yield, 73%;
reaction time, 2 h; eluent CH2Cl2/EtOAc 9:1. IR (KBr): ν 1751,
1
1639, 1601 cm-1. H NMR (250 MHz, CDCl3): δ 2.31 (s, 3H,
CH3), 3.72 (s, 3H, NCH3), 3.90 (s, 6H, OCH3), 6.28 (d, 1H, J )
2.0 Hz, HAr), 6.34 (d, 1H, J ) 2.0 Hz, HAr), 7.13 (d, 2H, J ) 8.8
Hz, HAr), 7.75 (d, 2H, J ) 8.8 Hz, HAr), 8.15 (s, 1H, dCH). 13C
NMR (62.90 MHz, CDCl3): δ 21.3, 30.4, 55.7, 55.9, 90.3, 92.7,
106.0, 121.1 (2), 126.3, 130.1 (3), 131.4, 142.1, 150.1, 157.8,
162.0, 162.7, 169.6. MS: m/z 354 (M+ + 1).
5,7-Dim eth oxy-3-(4-m eth oxyp h en yl)-1-m eth yl-2-(m eth -
ylsu lfa n yl)qu in olin iu m Iod id e (27). To a solution of 24 (682
mg, 2.0 mmol) in anhydrous THF (30 mL) was added io-

2554 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 12
J oseph et al.
maintaining the temperature at 37 °C, a computer containing
a frame grabber, and an image-processing software that
analyzes digitized frames. The software that we set up thus
enabled each living cell in the culture under study to be
isolated automatically on the basis of specific morphological
characteristics that in turn enables cells to be identified
against their background (the plastic Falcon dishes). Once the
segmentation procedure is complete, each cell is transformed
into its center of gravity. An image is digitized every 4 min by
the computer, and the video system is thus able to track the
trajectory of each cell by analyzing the movement of its center
of gravity. From these trajectories, the maximum relative
distance to the origin (the MRDO quantitative variable) of the
cell was calculated for each cell under analysis. This variable
thus describes the greatest linear distance found between the
original and the subsequent positions of the cell divided by
the observation time.24 All of the experiments were performed
over 48 h at a cell concentration of 10 000 MCF-7 cells/mL
MEM, with one image recorded every 4 min. The MCF-7 cells
were grown in a standard MEM medium (see above for the
MTT colorimetric assay) in control or in culture media
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(for more than two groups) or the Mann-Whitney (for two
groups) tests were carried out. All of the statistical analyses
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