Synthesis and cytotoxic evaluation of analogues of the marine pyridoacridine amphimedine

Article abstract of DOI:10.1016/S0968-0896(02)00148-7

4-Substituted-7H-pyrido-[4,3,2-de][1,8] or [1,9]-phenanthroline-7-ones and 9-methyl-1,4-diazanaphtacene-3,10-dione, analogues of the marine pyridoacridine amphimedine were synthesised from isoquinoline-5,8-dione. The first compounds were obtained starting

Full text of DOI:10.1016/S0968-0896(02)00148-7

Bioorganic & Medicinal Chemistry 10 (2002) 2845–2853
Synthesis and Cytotoxic Evaluation of Analogues of the
Marine Pyridoacridine Amphimedine
Christine Brahic,a Francis Darro,b Mirabelle Belloir,a Jean Bastide,a
Robert Kissc and Evelyne Delfournea,*
a
´
Centre de Phytopharmacie, UMR-CNRS 5054, Universite de Perpignan, 52 Avenue de Villeneuve,
66860 Perpignan Cedex, France
bLaboratoire Louis Lafon, Centre de Recherches, 19 avenue du Pr Cadiot, BP 22,
94701 Maisons-Alfort Cedex, France
c
´
Laboratoire d’Histopathologie, Faculte de Medecine, Universite Libre de Bruxelles, 808 Route de Lennik,
´
´
1070 Brussels, Belgium
Received 15 March 2002; accepted 7 May 2002
Abstract—4-Substituted-7H-pyrido-[4,3,2-de][1,8] or [1,9]-phenanthroline-7-ones and 9-methyl-1,4-diazanaphtacene-3,10-dione,
analogues of the marine pyridoacridine amphimedine were synthesised from isoquinoline-5,8-dione. The first compounds were
obtained starting from a Diels–Alder reaction whereas the synthesis of the last compound was initiated by a reaction of conden-
sation with 2-aminoacetophenone. The different tetra- and pentacyclic compounds were evaluated for in vitro cytotoxic activities
against six distinct human cancer cell lines. All the compounds exhibit cytotoxic activity with IC50 values (i.e., the drug concen-
tration inhibiting the mean growth value of the six cell lines by 50%)<10ꢀ7 M for two of them. # 2002 Elsevier Science Ltd. All
rights reserved.
Introduction
far.8 Diels–Alder cycloadditions between 1-aza-dienes
and isoquinoline-5,8-dione constitute a good access to
diaza-1,7- or 1,8-anthracenediones. More generally,
isoquinolinedione could be a good starting material in
synthetic routes to derivatives of amphimedine.
The pyridoacridines form an important class of marine
metabolites, many of which exhibit significant cytotoxic
activity.1 Amphimedine 1, isolated in 1983 from an
Amphimedon sp. sponge, was the first example of this
type of alkaloid from a marine organism.2 Recently,
Ireland’s group published the isolation of two structu-
rally related compounds, neoamphimedine 23 and
deoxyamphimedine 34 (Fig. 1).
Three total synthesis of amphimedine5 have been
reported in literature, with the first ones based on an
aza-Diels–Alder reaction involving a 5,8-quinolinedione
derivative and a 2-aza-diene. This Diels–Alder strategy
was also employed in the synthesis of the marine meta-
bolites meridine6 4 or cystodamine7 5, the diene being in
these cases a 1-aza-diene. To our knowledge, only a lim-
ited number of studies related to aza-Diels–Alder reac-
tions with isoquinoline-5,8-dione have been reported so
*Corresponding author. Tel.: +33-468-66-2251; fax: +33-468-66-
2223; e-mail: delfourn@univ-perp.fr
Figure 1.
0968-0896/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(02)00148-7

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C. Brahic et al. / Bioorg. Med. Chem. 10 (2002) 2845–2853
Figure 2.
As part of our ongoing programme to investigate new
anticancer drugs,9 we survey compounds comprising the
7H-pyrido[4,3,2-de][1,8 or 1,9-phenanthroline]-7-one
moiety and give the first account of their biological
properties (Fig. 2).
Table 1. Regioselectivity of cycloadditions of the different 1-aza-
dienes with isoquinoline-5,8-dione
R2
R1
Isomers 6
(%)
Isomers 7
(%)
Yield
(%)
Chemistry
Me
Me
H
OMe
F
6a (80)
6b (70)
6c (80)
6d (70)
7a (20)
7b (30)
7c (20)
7d (30)
36
39
36
3
The tetra- and pentacyclic compounds 8, 9, 12 and 13
were considered according to the retrosynthetic analysis
outlined in Scheme 1.
Me
C6H4(NHCOCF3)
H
The diazaanthraquinones 6 and 7 would be key inter-
mediates prepared by Diels–Alder reaction between
isoquinoline-5,8-dione and various 1-aza-dienes. The
different dienes involved in these reactions, that is
crotonaldehyde-N,N-dimethylhydrazone,10 2-methoxy-
2-butenal-N,N-dimethyl hydrazone,11 2-fluoro-2-butenal-
N,N-dimethylhydrazone12 and 4-(20-trifluoroacetamido-
phenyl)-1-dimethylamino-1-azadiene13 were synthesised
according to procedures described previously. The
[4+2] cycloadditions between isoquinoline-5,8-dione
and 1-azadienes were carried out in toluene, at room
temperature under inert atmosphere. These conditions
were those selected by Nebois and Fillion for a similar
reaction.8b In all reactions, two regioisomers were
obtained for which the structure assignments will be
further discussed. The percentage of each isomer repor-
an equivalent ratio of the two regioisomers. The
dihydroazaquinone intermediates were subsequently
oxidized by MnO2 to give the corresponding anthra-
quinones 6(a–d) and 7(a–d) (Schemes 2 and 3). No
attempts to improve the yields of these reactions were
done.
The formation of the last cycle from compounds 6a–c
and 7a–c was accomplished by means of a one pot
annelation method developed by Bracher.14 The reac-
tion of dimethylformamide diethyl acetal with the
methyldiazaanthraquinones gave quantitatively (yields
determined by 1H NMR) the enamine intermediates
which were treated with NH4Cl in refluxing ethanol (or
methanol) to yield the tetracyclic compounds 8a–c and
9a–c (Scheme 2).
1
ted in Table 1 was determined from the H NMR spec-
tra of reaction mixture. The different 1-aza-dienes led to
Scheme 2. (i) Toluene, dark, N2, rt/MnO2, CHCl3, rt; (j) DMF-DEA,
DMF, N2, 120 ꢁC; (k) NH4Cl, EtOH, reflux.
Scheme 1.

C. Brahic et al. / Bioorg. Med. Chem. 10 (2002) 2845–2853
2847
The two additional tetracyclic compounds 10 and 11
were obtained as minor products in the reaction of for-
mation of 8c. A possible explanation for the formation
of 10 is the nucleophilic substitution of the fluorine
atom by the dimethylamine released from the enamine
intermediate during the cyclisation step. Compound 11
could result of addition of ethanol to the enamine
before the cyclisation step (Fig. 3).
The pentacyclic compound 14 is an analogue of the
marine pyridoacridine ascididemine 15,15 it was syn-
thesized according to the synthetic procedure reported
by Bracher for this last alkaloid14 (Scheme 4).
Isoquinoline-5,8-dione was converted regioselectively to
the diaryl amine 16 by oxidative amination with 2-amino-
acetophenone in the presence of CeCl3 and air (Scheme
1). Cyclisation of 16 to the tetracyclic quinone 17 pro-
ceeded on heating with concentrated H2SO4 in glacial
acetic acid. The formation of ring E was accomplished
by the method previously described for compounds 8
and 9.
The diazaanthraquinones 6d and 7d, formed in very low
yields in the Diels–Alder reaction, were cyclised in
alkaline conditions to give the corresponding penta-
cyclic compounds 12 and 13 (Scheme 3).
Structural assignment
The structure of each isomer obtained in the Diels–
Alder reaction was assigned comparatively with related
Scheme 3. (a) CHCl3, 1 N NaOH, rt.
Scheme 4. (a) CeCl3, EtOH, rt, overnight; (b) cc H2SO4/CH3COOH,
reflux, 1 h; (c) DMF-DEA, DMF, N2, 120 ꢁC, 30min, NH 4Cl, MeOH,
reflux, 30min.
Figure 3.
Table 2. 1H NMR chemical shift (ppm) of the protons H1 and H4 of the different tetra- and pentacyclic compounds
Ref
Hb
Hb
R1=R2=H
R1=OMe, R2=H
R1, R2=
9.17
9.19
9.27
8.77
16
16
8.78
8.8015/17
Ha
Hb
Ha
Hb
R1, R2, R3=H
10.14
10.14
10.15
10.30
8.21
8.22
8.22
8.23
9.66
9.67
9.66
9.70
8.63
8.63
8.63
8.81
R1=OMe, R2, R3=H
R1=F, R2, R3=H
R1=H, R2, R3=

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C. Brahic et al. / Bioorg. Med. Chem. 10 (2002) 2845–2853
compounds reported in literature or univocaly syn-
thesised by our group in previous studies.15ꢀ17 The two
isomers have the same difference in their structural
environment both in the compounds of reference and in
the isoquinolinic derivatives described in this paper.
Table 3. HOMO and LUMO data of the different 1-aza-dienes and
isoquinoline-5,8-dione calculated by PM-3 method18
R2
R1
HOMO (eV)
c1
c2
In the case of the quinolinic derivatives reported in
1
Table 2, a notable difference in the H NMR chemical
Me
Me
H
OMe
F
ꢀ8.677
ꢀ8.449
ꢀ8.941
ꢀ8.795
0.3715
0.4021
0.4058
0.3622
0.3196
0.3130
0.2968
0.2946
shift of the proton H4 can be observed between the two
isomers. The proton close to the iminoquinone function
in isomers 8 shows a deshielding effect of around
0.4 ppm related to the same proton in isomers 9. This
deshielding effect may be due to the position of this
proton towards the D-pyridine ring. In the case of iso-
quinolinic derivatives synthesised in this work, we have
the same relative position of proton and pyridine rings.
Me
C6H4(NHCOCF3)
H
LUMO (eV)
c1
c2
ꢀ1.793
0.3159
0.3233
The data (Table 2) indicate
a
deshielding of
0.41–0.58 ppm between the protons Ha and Hb in the
two isomers; by analogy with the relationship pre-
viously established, these deshielded protons are close to
the iminoquinone moiety. Thus, the structure 8 and 9
can be assigned to compounds a to c as well as the
structures 12 and 13. It has to be noted that a long
range constant is systematically observed between pro-
tons Ha and Hb in isomers 9 but not in isomers 8.
Table 4. Characterization of the in vitro cytotoxic-related antitumor
effects IC50 (mM)
Compounds
Cell lines
U-87MG U-373MG J82 HCT-15 LoVo A549
8a
9a
8b
9b
8c
9c
6d
7d
12
13
14
2.2
0.5
0.5
0.8
3.2
6.9
0.06
0.02
0.1
0.1
1
0.8
0.3
0.7
0.8
2.4
0.8
3.1
>101.3
0.7
0.3
0.5
2.5
0.8
2.9
0.9
3.1
0.8
These results are in accordance with the theoretical
prediction of the regioselectivity of the Diels–Alder
reaction. Indeed, this regioselectivity can be rationalised
by FMO theory. The hetero-Diels–Alder reaction of the
different dienes and isoquinoline-5,8-dione is a normal
Å4s+Å2s process with a HOMOdiene-LUMOdienophile
control. The HOMOdiene-LUMOdienophile gaps for
4.2
1.8
>10
>10
0.15
32..083.0 2.5
4.4
0.03
0.1
0.2
0.05
2.5
6.9
0.7
0.2
2.9
1.4
3
3.6
5.1
0.05
0.04
0.2
0.3
0.9
0.08
0.04
0.4
0.05
2.8
0.06
0.2
1.8
0.7
crotonaldehyde-N,N-dimethylhydrazone,
2-methoxy-
2-butenal-N,N-dimethyl hydrazone, 2-fluoro-2-butenal-
N,N-dimethylhydrazone and 4-(20-trifluoroacetamido-
phenyl)-1-dimethylamino-1-azadiene are 6.884, 6.956,
7.148 and 7.002 eV, respectively, calculated by using the
PM3 semi-empirical molecular orbital method.18 If the
sole atoms considered are those which are bonded, the
favored addition was that associating the atom having
the larger coefficient for the HOMO of the diene with
the atom having the larger coefficient for the LUMO of
the dienophile (Table 3).
controlled by the position of the ring nitrogen atom
relative to the carbonyl group. In the same way, the
regioselectivity of the reaction of isoquinoline-5,8-dione
with amines described previously19 was based mainly on
the speculation that the C-5 carbonyl group, which is para
to the nitrogen, is more electron deficient than the C-8
carbonyl group. In this work, the regioselectivity of both
these two types of reaction was confirmed by NMR data.
The major isomer was the structure 6, nevertheless, the
difference between the coefficients of isoquinoline-
5,8-dione being low (0.3159 vs 0.3233), the two regioi-
somers were formed.
Biological Results
Although a sole regioisomer, analogue of ascididemin
1
was obtained, the H NMR chemical shift of protons
Ha and Hb (9.68 and 8.70, respectively) are consistent
with structure 14 for the compound synthesised.
The cytotoxic activity of the tetra- and pentacyclic
compounds 8, 9 (a–c), 6d, 7d, 12, 13 and 16 was tested
on six distinct human cancer cell lines including various
histopathological types (glioblastomas, colon, lung, and
bladder cancers). The colorimetric MTT assay, which
indirectly assesses the effect of potentially anticancer
compounds on overall growth of adherent cell lines20
was used. The IC50 values, that is the concentration
which reduced the mean growth value of the six cell
lines by 50%, was determined for each drug, as com-
pared to the mean control growth value. Table 4 illu-
strates the individual IC50 values of the different
Two authors have previously described the Diels–Alder
addition of 2-butenal-N,N-dimethylhydrazone8d and
2-ethoxy-2-butenal-N,N-dimethylhydrazone8c with iso-
quinoline-5,8-dione. They reported the formation of
two isomers with a majority one, and they had in both
reactions, the same regioselectivity as that we observed
in this work. They performed the structural assignment
of the major isomer from the reactivity of dienophile

C. Brahic et al. / Bioorg. Med. Chem. 10 (2002) 2845–2853
2849
1
compounds obtained for each of the six cell lines under
study. All those compounds present cytotoxic activity.
The derivatives with the most close structure to the
natural product amphimedine (12 and 13) exhibited
IC50 values <10ꢀ7 M and are the more active com-
pounds of this series with the intermediates 6d and 7d.
The substituted tetracycles 8b, 8c and 9b, 9c were sen-
sibly less cytotoxic than the unsubstituted ones 8a and
9a. The difference in activity between the two isomers
was relatively weak. More generally, the same cytotoxi-
city was observed from a cell line to another, marking
the poor selectivity of these compounds.
(6a). Brown solid (1.3 g, 29%), mp >260 ꢁC. H NMR
(CDCl3) 2.92 (s, 3H); 7.56 (dd, 1H, J=4.8 and 0.7 Hz);
8.12 (dd, 1H, J=5.2 and 0.7 Hz); 8.93 (d, 1H,
J=4.8 Hz); 9.12 (d, 1H, J=5.2 Hz); 9.55 (d, 1H,
J=0.7 Hz). 13C NMR (CDCl3) 22.83; 118.87; 126.46;
128.99; 131.89; 137.32; 149.62; 149.91; 151.90; 153.85;
155.36; 181.51; 184.12. IR (CHCl3) 1698; 1678 cmꢀ1. MS
(m/z) 224 (42); 210 (100); 196 (26); 182 (49); 168 (10).
(7a). Brown solid (0.3 g, 7%), mp 245 ꢁC. 1H NMR
(CDCl3) 2.91 (s, 3H); 7.55 (dd, 1H, J=4.8 and 0.7 Hz);
8.05 (dd, 1H, J=5.2 and 0.7 Hz); 8.96 (d, 1H,
J=4.8 Hz); 9.13 (d, 1H, J=5.2 Hz); 9.62 (d, 1H,
J=0.7 Hz). 13C NMR (CDCl3) 22.83; 119.06; 125.43;
129.06; 131.47; 138.77; 149.59; 149.92; 151.91; 154.21;
In conclusion, we have synthesised three different types
of pyridoacridine derivatives from isoquinoline-5,8-
dione; the compounds which exhibit the better potency
against cancer cells were the amphimedine analogues.
155.75; 181.26; 184.13. IR (CHCl3) 1692; 1680cm ꢀ1
MS (m/z) 224 (100); 196 (18); 168 (26); 140 (25).
.
3-Methoxy-4-methyl-1,7-diazaanthraquinone (6b) and
3-methoxy-4-methyl-1,6-diazaanthraquinone (7b). Method
A was used and involved isoquinolinedione (715 mg,
4.5 mmol) in toluene (55 mL), 2-methoxy-2-butenal-
dimethylhydrazone (950mg, 6.75 mmol) in toluene
(15 mL), stirring 16 h. Filtration was done on silicagel
(CH2Cl2/MeOH 99:1). MnO2 85% (3.2 g, 31,5 mmol)
and CHCl3 (30mL) were added, and the solution was
stirred for 6 h. The crude product was washed with ether
before flash-chromatography (CH2Cl2/MeOH 99:1)
which gave the expected tricyclic compounds.
Experimental
Chemical synthesis
1H and 13C NMR spectra were obtained with a JEOL
400 MHz spectrometer with the chemical shifts of the
remaining protons of the deuterated solvents serving as
internal standards. IR spectra were obtained with a
Perkin-Elmer (1600 series FTIR) spectrometer. Mass
spectra (MS) were recorded on an automass Unicam
spectrometer. Reagents were purchased from commer-
cial sources and used as received. Chromatography was
performed on silicagel (15–40 mm) using the solvent sys-
tems indicated below. The purity of the different ascidi-
demin analogues was evaluated on an analytical
chromatographic system consisted of a ZorbaxNH2,
5 mm column (150 ꢂ 4.6 mm), isooctane/MeOH/EtOH
80:10:10 at 1.5 mL/min flow rate.
(6b). Ochre solid (309 mg, 27%), mp >260 ꢁC. 1H NMR
(CDCl3) 2.77 (s, 3H); 4.11 (s, 3H); 8.10(dd, 1H, J=5.9
and 0.7 Hz); 8.65 (s, 1H); 9.09 (d, 1H, J=5.9 Hz); 9.52
(d, 1H, J=0.7 Hz). 13C NMR (CDCl3) 13.02; 56.96;
118.91; 127.14; 129.63; 136.78; 137.83; 138.98; 142.85;
149.85; 155.21; 157.49; 180.75; 184.81. IR (CHCl3)
1688; 1674 cmꢀ1. MS (m/z) 254 (100); 236 (7); 223 (13);
211(20); 183 (18).
Diels–Alder reactions of isoquinoline-5,8-dione with the
different dienes
(7b). Ochre solid (132 mg, 12%), mp >260 ꢁC. 1H NMR
(CDCl3) 2.75 (s, 3H); 4.13 (s, 3H); 8.02 (d, 1H,
J=5.1 Hz); 8.68 (s, 1H); 9.10(d, 1H, J=5.1 Hz); 9.61 (s,
1H). 13C NMR (CDCl3) 13.05; 56.96; 119.04; 125.60;
129.59; 137.07; 139.02; 139.31; 142.80; 149.87; 155.38;
157.11; 180.48; 184.73. IR (CHCl3) 1684 cmꢀ1. MS (m/z)
254 (100); 223 (6); 211 (18); 183 (21).
General method A. To a solution of isoquinoline-5,8-
dione in toluene was added dropwise under nitrogen
atmosphere a solution of diene in toluene. The reaction
media was stirred in the dark, at room temperature for
different times. After concentration to dryness, the
crude product was filtered on silicagel to give the Diels–
Alder adducts. MnO2 85% and CHCl3 were added, and
the mixture was stirred at room temperature. After fil-
tration over Celite, the crude product was purified on
flash-chromatography silicagel column to give the
expected products.
3-Fluoro-4-methyl-1,7-diazaanthraquinone
(6c)
and
3-fluoro-4-methyl-1,6-diazaanthraquinone (7c). Method
A was used and involved isoquinolinedione (1.7 g,
10.7 mmol) in toluene (130 mL), 2-fluoro-2-butenal-
dimethylhydrazone (820mg, 6.3 mmol) in toluene
(5 mL), the reaction was stirred for 32 h. The mixture of
Diels–Alder adducts was stirred with silicagel (30g) in
AcOEt (100 mL) at room temperature for 1 h. The sili-
cagel was filtered off and the solution was concentrated
to dryness. MnO2 85% (5.5 g, 54 mmol) and CHCl3
(60mL) were added, and the solution was stirred over-
night. Flash-chromatography (CH2Cl2) gave the expec-
ted tricyclic compounds.
4-Methyl-1,7-diazaanthraquinone (6a) and 4-methyl-1,6-
diazaanthraquinone (7a). Method A was used and
involved isoquinolinedione (3.2 g, 20mmol) in toluene
(240mL), crotonaldehyde-dimethylhydrazone (3.38 g,
30mmol) in toluene (15 mL), stirring 45 h. Filtration
was done on silicagel (CH2Cl2/MeOH 95:5). MnO2 85%
(20.5 g, 0.2 mol) and CHCl3 (200 mL) were added, and
the solution was stirred at room temperature for 4 h
30min. Flash-chromatography (CH 2Cl2/MeOH 99:1)
gave the expected tricyclic compounds.
ꢁ
1
(6c). Yellow-brown solid (432 mg, 29%), mp 170 C. H
NMR (CDCl3) 2.85 (s, 3H); 8.13 (d, 1H, J=5.1 Hz);

2850
C. Brahic et al. / Bioorg. Med. Chem. 10 (2002) 2845–2853
8.86 (s, 1H); 9.14 (d, 1H, J=5.1 Hz); 9.55 (s, 1H). 13C
NMR (CDCl3) 12.58 (d, J=5 Hz); 119.03; 126.63;
130.33; 137.31; 137.63 (d, J=15 Hz); 142.70(d,
J=28 Hz); 145.77 (d, J=7 Hz); 149.92; 155.64; 160.65
(d, J=263 Hz); 180.24; 183.88. IR (CHCl3) 1698,
1677 cmꢀ1. MS (m/z) 242 (100); 214 (64); 186 (71); 158
(52).
After concentration to dryness, NH4Cl and ethanol
were added and the mixture was refluxed for additional
time. The solvent was removed over vacuum, water was
added and the mixture was extracted four times with
CH2Cl2. The organic layers were dried over MgSO4 and
concentrated. The crude product was purified to give
the expected tetracyclic derivative.
(7c). Yellow-brown solid (108 mg, 7%), mp 216 ꢁC. H
7H - Pyrido[4,3,2 - de][1,9]phenanthroline - 7 - one (8a).
Method B was used and involved tricyclic compound
6a (100 mg, 0.446 mmol), DMF-DEA (0.27 mL,
1.56 mmol) in DMF (1 mL), heating at 120 ꢁC for
15 min. NH4Cl (685 mg, 12.9 mmol) and absolute EtOH
(90mL) were added and the reaction was refluxed for
30min. Flash chromatography (CH 2Cl2/MeOH 99:1)
gave the expected tetracyclic compound as a yellow
solid (42 mg, 40%), mp >260 ꢁC. 1H NMR (CDCl3)
7.80(d, 1H, J=5.8 Hz, C3–H); 8.01 (d, 1H, J=5.5 Hz,
C4–H); 8.21 (d, 1H, J=5.2 Hz, C8–H); 8.96 (d, 1H,
J=5.8 Hz, C2–H); 9.03 (d, 1H, J=5.2 Hz, C9–H); 9.19
(d, 1H, J=5.5 Hz, C5–H); 10.14 (s, 1H, C11–H). 13C
NMR (CDCl3) 119.74; 119.76; 119.95; 124.36; 128.42;
136.93; 138.81; 147.30; 147.65; 148.63; 148.88; 149.88;
152.68; 181.75. IR (CHCl3) 1689 cmꢀ1. MS (m/z) 233
(100); 204 (24); 178 (16); 151 (36). tR is 4.13 min (100%
purity).
1
NMR (CDCl3) 2.80(s, 3H); 8.01 (d, 1H, J=5.1 Hz) ;
8.85 (s, 1H); 9.12 (d, 1H, J=5.1 Hz); 9.58 (s, 1H). 13C
NMR (CDCl3) 12.55 (d, J=5 Hz); 118.99; 125.21;
130.26; 137.5 3 (d, J=14 Hz); 137.77; 142.86 (d,
J=27 Hz); 145.56 (d, J=7 Hz); 149.98; 155.74; 160.23
(d, J=253 Hz);179.91; 183.90. IR (CHCl3) 1712 cmꢀ1
.
MS (m/z) 242 (100); 214 (15); 185 (19); 158 (33); 131
(31).
4 - (2 - Trifluoroacetamidophenyl)pyrido[4,3- g]quinoleine-
5,10-dione (6d) and 4-(2-trifluoroacetamidophenyl)pyri-
do[3,4]quinoleine-5,10-dione (7d). A mixture of iso-
quinolinedione (3.5 g, 22mmol), 4-(2-trifluoroacetamido
phenyl)-1-dimethylamino-1-aza-1,3-butadiene
(6.9 g,
24.2 mmol), acetic anhydride (7.2 mL) and 10% Pd/C
(4.6 g) in CH3CN (250mL) was refluxed under nitrogen
atmosphere and in the dark for 29 h. The reaction media
was filtered over Celite and concentrated to dryness.
The crude product was purified by flash chroma-
tography (CH2Cl2/MeOH 99:1) to give the expected
tricyclic compounds.
7H - Pyrido[4,3,2 - de][1,8]phenanthroline - 7 - one (9a).
Method B was used and involved tricyclic compound
7a (140 mg, 0.63 mmol), DMF-DEA (0.37 mL,
2.19 mmol) in DMF (1 mL), heating at 120 ꢁC for
15 min. NH4Cl (960mg, 17.9 mmol) and absolute EtOH
(90mL) were added and the reaction was refluxed for
30min. Flash chromatography (CH 2Cl2/MeOH 99:1)
gave the expected tetracyclic compound as a yellow
solid (41 mg, 28%), mp >260 ꢁC. 1H NMR (CDCl3)
7.87 (d, 1H, J=5.8 Hz, C3–H); 8.00 (d, 1H, J=5.5 Hz,
C4–H); 8.63 (dd, 1H, J=5.2 and 0.7 Hz, C11–H); 8.98
(d, 1H, J=5.8 Hz, C2–H); 9.05 (d, 1H, J=5.2 Hz,
C10–H); 9.20(d, 1H, J=5.5 Hz, C5–H); 9.66 (d, 1H,
J=0.7 Hz, C8–H). 13C NMR (CDCl3) 118.24; 120.72;
121.08; 123.79; 126.26; 138.79; 141.86; 147.48; 147.72;
148.96; 149.36; 150.73; 155.21; 181.66. IR (CHCl3)
1683 cmꢀ1. MS (m/z) 233 (100); 204 (25); 178 (25); 151
(26). tR is 4.49 min (92% purity).
(6d). Brown solid (145 mg, 1.7%), mp 232 ꢁC. H NMR
1
(CDCl3): 7.21 (dd, 1H, J=1.5 and 7.7 Hz, C1–H); 7.48
(ddd, 1H, J=1.5; 7.7 and 7.7 Hz, C2–H); 7.60(ddd, 1H,
J=1.4; 8.1 and 7.7 Hz, C3–H); 7.62 (d, 1H, J=5.1 Hz,
C5–H); 7.70(d, 1H, J=8.1 and 1.4 Hz, C4–H); 7.90
(br.s, 1H, NH); 8.15 (dd, 1H, J=0.8 and 4.8 Hz, C9–H);
9.14 (d, 1H, J=5.1 Hz, C6-H); 9.16 (d, 1H, J=4.8 Hz,
C10–H); 9.38 (d, 1H, J=0.8 Hz, C11–H). 13C NMR
(CDCl3): 115.51 (q, J=367 Hz); 119.16; 125.67; 125.97;
127.83; 128.15; 128.93; 129.99; 131.55; 131.65; 134.52;
137.40; 148.28; 149.23; 149.76; 154.78; 155.47 (q,
J=72 Hz); 155.80; 180.73; 183.12. IR (CHCl3): 1736,
1698, 1679 cmꢀ1. MS (m/z) 397 (45); 328 (34); 285 (100);
216 (9). tR is 14.71 min (100% purity).
(7d). Beige solid (67 mg, 0.8%), mp >260 ꢁC. H NMR
4-Methoxy-7H-pyrido[4,3,2-de][1,9]phenanthroline-7-one
(8b) and 4-methoxy-7H-pyrido[4,3,2-de][1,8]phenanthro-
line-7-one (9b). Method B was used and involved a
mixture of tricyclic compound 6b and 7b (250mg,
0.98 mmol), DMF-DEA (0.63 mL, 3.44 mmol) in DMF
(2.5 mL), heating at 120 ꢁC for 75 min. NH4Cl (1.5 g,
35.5 mmol) and absolute EtOH (250mL) were added
and the reaction was refluxed for 30min. Flash-chroma-
tography (CH2Cl2/MeOH 98:2) gave the expected tet-
racyclic compounds as a yellow solid (170mg, 69%).
The two isomers were separated by successive chroma-
tographies (CH2Cl2/MeOH 98:2).
1
(CDCl3): 7.21 (d, 1H, J=7.4 Hz, C1–H); 7.47 (dd, 1H,
J=7.4 and 7.7 Hz, C2–H); 7.59–7.62 (m, 2H, C3–H and
C5–H); 7.73 (d, 2H, J=8.1 Hz, C4–H+NH); 7.85 (d,
1H, J=4.8 Hz, C12–H); 9.09 (d, 1H, J=4.8 Hz, C6–H);
9.18 (d, 1H, J=4.8 Hz, C11–H); 9.63 (s, 1H, C9–H). 13C
NMR (CDCl3): 115.45 (q, J=287 Hz); 118.92; 125.30;
125.43; 127.95; 128.05; 129.00; 130.04; 131.08; 131.44;
133.96; 138.22; 148.03; 149.25; 150.04; 155.12; 155.14 (q,
J=37 Hz); 155.94; 180.44; 183.22. IR (CHCl3): 1736,
1693 cmꢀ1. tR is 16.62 min (97% purity).
Formation of tetracyclic compounds
(8b). Yellow solid (118 mg, 65%) mp >260 ꢁC. 1H
NMR (CDCl3) 4.27 (s, 3H); 8.07 (d, 1H, J=5.9 Hz, C3–
H); 8.22 (d, 1H, J=5.2 Hz, C8–H); 8.71 (s, 1H, C5–H);
8.96 (d, 1H, J=5.9 Hz, C2–H); 9.00 (d, 1H, J=5.2 Hz,
General method B. A solution of tricyclic intermediate
and dimethylformamide diethylacetal (DMF-DEA) in
DMF was heating at 120 ꢁC under nitrogen atmosphere.

C. Brahic et al. / Bioorg. Med. Chem. 10 (2002) 2845–2853
2851
C9–H); 10.14 (s, 1H, C11–H). 13C NMR (CDCl3) 57.07;
114.90; 119.80; 128.73; 129.48; 130.27; 137.38; 140.26;
146.88; 148.53; 148.91; 152.39; 153.40; 174.07; 180.61.
IR (CHCl3) 1674 cmꢀ1. MS (m/z) 263 (82); 248 (62); 220
(100); 192 (32). tR is 3.74 min (98% purity).
15 min. NH4Cl (254 mg, 4.74 mmol) and absolute EtOH
(40mL) were added and the reaction was refluxed for
30min. Flash chromatography (CH 2Cl2/MeOH 99:1)
gave the expected tetracyclic compound as a yellow
solid (10mg, 23%), mp >260 ꢁC. 1H NMR (CDCl3)
8.07 (d, 1H, J=5.8 Hz, C3–H); 8.63 (dd, 1H, J=5.5 and
0.7 Hz, C11–H); 9.01 (d, 1H, JH-F=1.4 Hz, C5–H); 9.06
(d, 1H, J=5.8 Hz, C2–H); 9.07 (d, 1H, J=5.5 Hz, C10–
H); 9.66 (d, 1H, J=0.7 Hz, C8–H). 13C NMR (CDCl3)
114.23; 118.19; 121.49; 126.14; 129.04 (d, J=17 Hz);
135.14 (d, J=23 Hz); 141.54; 143.65; 147.88; 149.07;
150.79; 155.15; 155.60 (d, J=274 Hz); 180.11. IR
(CHCl3) 1721, 1684 cmꢀ1. MS (m/z) 251 (100); 222 (17);
196 (26); 169 (24). tR is 3.75 min (94% purity).
(9b). Yellow solid (63 mg, 81%), mp >260 ꢁC. 1H NMR
(CDCl3) 4.27 (s, 3H); 8.15 (d, 1H, J=5.9 Hz, C3–H);
8.63 (dd, 1H, J=5.5 and 0.8 Hz, C11–H); 8.71 (s, 1H,
C5–H); 8.97 (d, 1H, J=5.9 Hz, C2–H); 9.04 (d, 1H,
J=5.5 Hz, C10–H); 9.67 (d, 1H, J=0.8 Hz, C8–H). 13C
NMR (CDCl3) 57.14; 116.31; 118.16; 121.08; 126.56;
129.34; 130.41; 140.36; 141.98; 146.96; 148.37; 150.72;
152.95; 154.64; 180.65. IR (CHCl3) 1674 cmꢀ1. MS (m/z)
263 (20); 248 (13); 220 (8). tR is 3.88 min (99% purity).
Benzo[b]pyrido[4,3,2-de][1,9]phenanthroline-8-one (12). A
solution of compound 6d (100 mg, 0.25 mmol) in CHCl3
(75 mL) and 1 N NaOH (3.3 mL) was stirred at room
temperature for 2 h 30min. The organic layer was sepa-
rated and the aqueous layer was extracted with CHCl3
(3 ꢂ 100 mL). The combined organic layers were dried
over MgSO4 and concentrated over vacuum to give the
expected pentacyclic compound as a yellow solid
(65 mg, 91%), mp >260 ꢁC. 1H NMR (CDCl3): 7.85
4-Fluoro-7H-pyrido[4,3,2-de][1,9]phenanthroline-7-one
(8c), 4-dimethylamino-7H-pyrido[4,3,2-de][1,9]phenanthro-
line-7-one (10) and ethoxy-4-fluoro-7H-pyrido[4,3,2-
de][1,9]phenanthroline-7-one (11). Method B was used
and involved tricyclic compound 6c (80mg, 0.33 mmol),
DMF-DEA (0.2 mL, 1.16 mmol) in DMF (0.7 mL),
heating at 120 ꢁC for 15 min. NH4Cl (507 mg,
9.48 mmol) and absolute EtOH (80mL) were added and
the reaction was refluxed for 30min. Flash chroma-
tography (CH2Cl2/MeOH 99:1) gave the expected
tetracyclic compound and two side products.
(ddd, 1H, J=7.7, 7.0and 1.4 Hz, C –H); 7.97 (dd, 1H,
3
J=8.1, 7.0and 1.1 Hz, C –H); 8.23 (d, 1H, J=5.1 Hz,
2
C9–H); 8.40(dd, 1H, J=8.1 and 1.1 Hz, C1–H); 8.62
(dd, 1H, J=7.7 and 1.4 Hz, C4–H); 8.70(d, 1H,
J=5.5 Hz, C5–H); 9.04 (d, 1H, J=5.1 Hz, C10–H); 9.35
(d, 1H, J=5.5 Hz, C6–H); 10.30 (s, 1H, C12–H). 13C
(CDCl3) NMR 118.47; 119.48; 120.28; 121.67; 122.92;
128.53; 129.32; 131.69; 132.09; 137.12; 138.29; 145.63;
146.70; 147.66; 149.03; 150.35; 152.42; 181.97. IR
(CHCl3): 1685 cmꢀ1. MS (m/z) 283 (100); 254 (4); 228
(8).
(8c). Yellow solid (24 mg, 29%), mp >260 ꢁC. 1H NMR
(CDCl3) 8.00 (d, 1H, J=5.8 Hz, C3–H); 8.22 (d, 1H,
J=5.2 Hz, C8–H); 9.01 (d, 1H, JH-F=0.8 Hz, C5–H);
9.04 (d, 1H, J=5.2 Hz, C9–H); 9.05 (d, 1H, J=5.8 Hz,
C2–H); 10.15 (s, 1H, C11–H). 13C NMR (CDCl3) 112.91;
119.93; 120.81; 128.25; 129.06 (d, J=20Hz); 135.27 (d,
J=24 Hz); 137.09; 143.65; 147.92; 148.65; 149.81;
153.04; 156.11 (d, J=288 Hz); 180.19. IR (CHCl3)
1684 cmꢀ1. MS (m/z) 251 (100); 223 (36); 196 (24); 169
(25). tR is 3.58 min (100% purity).
Benzo[b]pyrido[4,3,2 - de][1,8]phenanthroline - 8 - one (13).
The same procedure as for compound 12 involving
compound 7d (32 mg, 0.081 mmol) in CHCl3 (25 mL)
and 1 N NaOH (1.1 mL) was used. The aqueous layer
was extracted with CHCl3 (3 ꢂ 50mL). The combined
organic layers were dried over MgSO4 and concentrated
over vacuum to give the expected pentacyclic compound
(10). Red solid (6 mg, 7%), mp >260 ꢁC. 1H NMR
(CDCl3) 3.36 (s, 6H); 7.94 (d, 1H, J=5.9 Hz); 8.23 (dd,
1H, J=5.1 and 0.7 Hz); 8.60 (s, 1H); 8.84 (d, 1H,
J=5.9 Hz); 8.96 (d, 1H, J=5.1 Hz); 10.14 (d, 1H,
J=0.7 Hz). 13C NMR (CDCl3) 29.68; 44.05; 117.66;
119.50; 120.84; 128.66; 129.37; 135.11; 137.50; 137.60;
144.65; 147.39; 148.47; 149.27; 151.87; 180.06. IR
(CHCl3) 3690; 1660 cmꢀ1. MS (m/z) 251 (100); 223 (36);
196 (24); 169 (25).
as a yellow solid (22 mg, 96%), mp>260 ꢁC. H NMR
1
(CDCl3) 7.87 (ddd, 1H, J=7.1; 8.0and 1.1 Hz, C –H);
3
7.99 (ddd, 1H, J=8.4; 7.1 and 1.1 Hz, C2–H); 8.41 (dd,
1H, J=8.4 and 1.1 Hz, C1-H); 8.65 (dd, 1H, J=8.0and
1.1 Hz, C4–H); 8.68 (d, 1H, J=5.5 Hz, C5–H); 8.81 (dd,
1H, J=5.1 and 0.8 Hz, C12–H); 9.09 (d, 1H, J=5.1 Hz,
C11–H); 9.36 (d, 1H, J=5.5 Hz, C6–H); 9.70(d, 1H,
J=0.8 Hz, C9–H). 13C NMR (CDCl3): 118.28; 119.02;
119.77; 122.23; 123.02; 126.44; 129.82; 131.82; 132.07;
138.07; 141.89; 145.48; 146.83; 147.13; 150.37; 150.62;
154.75; 181.69. IR (CHCl3): 1684 cmꢀ1. MS (m/z) 283
(100); 254 (29); 227 (14).
(11). Brown solid (7 mg, 7%), mp 196 ꢁC. 1H NMR
(CDCl3) 1.56 (t, 3H, J=7 Hz); 4.69 (q, 2H, J=7 Hz);
7.24 (s, 1H); 8.18 (dd, 1H, J=5.0and 0.7 Hz); 8.76 (s,
1H); 9.01 (d, 1H, J=5.0 Hz); 10.03 (s, 1H). 13C NMR
(CDCl3) 14.65; 63.75; 95.91; 117.93; 120.03; 128.08;
132.74 (d, J=18 Hz); 133.66 (d, J=24 Hz); 137.19;
144.08; 148.37; 149.18; 153.03; 155.71 (d, J=277 Hz);
163.61; 180.49. IR (CHCl3) 1689; 1618 cmꢀ1. MS (m/z)
295 (20); 239 (44).
5-(2-Acetylphenylamino)isoquinoline-3,6-dione (16).
A
solution of isoquinoline-5,8-dione (1 g, 6.29 mmol) in
ethanol (60mL) was added dropwise to a solution of
cerium(III) chloride CeCl3 (3 g, 12.59 mmol) and amino-
acetophenone (1.7 g, 12.59 mmol) in ethanol (25 mL).
The reaction media was stirred at room temperature
overnight, hydrolysed by 10% acetic acid (100 mL) and
4-Fluoro-7H-pyrido[4,3,2-de][1,8]phenanthroline-7-one
(9c). Method B was used and involved tricyclic com-
pound 7c (40 mg, 0.17 mmol), DMF-DEA (0.1 mL,
0.58 mmol) in DMF (0.35 mL), heating at 120 ꢁC for

2852
C. Brahic et al. / Bioorg. Med. Chem. 10 (2002) 2845–2853
extracted with CH2Cl2 (4 ꢂ 200 mL). The organic layers
were dried over MgSO4 and concentrated to dryness.
The crude product was purified by flash chroma-
tography (CH2Cl2) to give the expected product as a red
solid (190mg, 51%), mp 204 ꢁC. RMN 1H (CDCl3) 2.69
(s, 3H); 6.80(s, 1H); 7.23 (ddd, 1H, J=1.5, 6.6 and
8.4 Hz); 7.60(ddd, 1H, J=1.5, 6.6 and 8.1 Hz); 7.62 (dd,
1H, J=8.1 and 1.5 Hz), 7.90(dd, 1H, J=0.8 and
5.0Hz); 7.97 (dd, 1H, J=1.5 and 8.4 Hz); 11.33 (s, 1H).
13C NMR (CDCl3): 28.51; 106.19; 118.54; 120.95;
123.67; 124.17; 125.97; 132.41; 134.21; 138.23; 139.64;
143.85; 148.58; 156.21; 181.22; 182.83; 201.51. IR
models. The ATCC numbers of these cell lines are HTB
14 (U-87 MG), HTB 17 (U-373MG), CCL225 (HCT-15),
CCL229 (LoVo), CCL 185 (A549), and HTB1 (J82).
The cells were cultured at 37 ꢁC in sealed (airtight) Fal-
con plastic dishes (Nunc, Gibco, Belgium) containing
Eagle’s minimal essential medium (MEM, Gibco) sup-
plemented with 5% fetal calf serum (FCS). All 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 genta-
mycin (Gibco). The FCS was heat-inactivated for 1 h at
56 ꢁC.
(CHCl3) 1684, 1664, 1641 cmꢀ1
.
The six 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.21,22
This assessment of cell population growth is based on
the capability of living cells to reduce the yellow product
MTT (3-(4,5)-dimethylthiazol-2-yl)-2,5-diphenyltetra-
zolium bromide; (Sigma, St Louis, MO, USA) to a blue
product, formazan, by a reduction reaction occurring in
the mitochondria. The number of living cells is directly
proportional to the intensity of the blue, which is
quantitatively measured by spectrophotometry on a
DIAS microplate reader (Dynatech Laboratories,
Guyancourt, France) at a 570nm wavelength (with a
reference of 630nm). Each experiment was carried out
in sextuplicate. Nine concentrations ranging from 10ꢀ5
to 10ꢀ9 M were assayed for each of the compounds
under study.
9-Methyl-1,4-diazanaphtacene-3,10-dione (17). To
a
solution of tetracyclic compound 16 (0.5 g, 1.71 mmol)
in acetic acid (12 mL), was added dropwise a solution of
concentrated sulfuric acid (2 mL) in acetic acid (10mL).
The reaction media was refluxed for 1 h and after cool-
ing was poured into ice. The mixture was neutralized by
K2CO3 and extracted with CH2Cl2 (4 ꢂ 50mL). The
organic layers were dried over MgSO4 and concentrated
to dryness. Flash chromatography (CH2Cl2/MeOH
99:1) gave the expected tetracyclic compound as a green
solid (248 mg, 53%), mp >260 ꢁC. H NMR (CDCl3):
1
3.31 (s, 3H); 7.81 (ddd, 1H, J=1.0; 7.0 and 8.4 Hz); 7.96
(ddd, 1H, J=1.0, 7.0 and 8.4 Hz); 8.11 (dd, 1H, J=0.7
and 5.1 Hz); 8.40(dd, 1H, J=1.0and 8.4 Hz); 8.48 (dd,
1H, J=1.0and 8.4 Hz); 9.14 (d, 1H, J=5.1 Hz); 9.66 (d,
1H, J=0.7 Hz). 13C NMR (CDCl3): 16.75; 119.49;
125.17; 125.57; 126.24; 129.65; 129.98; 132.53; 133.06;
140.21; 147.64; 148.91; 150.18; 152.78; 155.72; 181.53;
184.27. IR (CHCl3) 1690, 1675 cmꢀ1
.
References and Notes
9H-Quino[4,3,2-de][1,7]phenanthrolin-9-one (14). Method
B using tetracyclic compound 17 (150mg, 0.55 mmol),
dimethylformamide diethylacetal (0.33 mL, 1.92 mmol)
in DMF (3.3 mL), the reaction was warmed at 120 ꢁC
under nitrogen atmosphere for 30min. NH 4Cl (0.88 g,
17.1 mmol) and methanol (18 mL) were added and the
reaction was refluxed for 30min. The crude product was
purified by flash-chromatography (CH2Cl2/MeOH 98:2)
to give the pentacyclic compound as a yellow solid
(35 mg, 23%), mp >260 ꢁC. 1H NMR (CDCl3): 7.96
(ddd, 1H, J=1.1, 7.3 and 8.2 Hz); 8.04 (ddd, 1H,
J=1.4, 7.3 and 8.2 Hz); 8.56 (d, 1H, J=5.7 Hz); 8.66
(dd, 1H, J=8.2 and 1.1 Hz); 8.69 (dd, 1H, J=8.2 and
1.4 Hz); 8.70(dd, 1H, J=5.1 and 0.7 Hz); 9.06 (d, 1H,
J=5.1 Hz); 9.16 (d, 1H, J=5.7 Hz); 9.68 (d, 1H,
J=0.7 Hz). 13C NMR (CDCl3): 117.27; 118.21; 118.48;
122.89; 123.28; 126.32; 130.86; 132.03; 133.32; 138.02;
142.48; 145.98; 146.17; 148.42; 149.15; 150.79; 155.25,
181.80. IR (CHCl3) 1653, 1690cm ꢀ1. MS (m/z): 283
(100); 254 (27); 228 (8). tR is 7.05 min (95% purity).
1. (a) Molinski, T. F. Chem. Rev. 1993, 93, 1825. (b) Ding, Q.;
Chichak, K.; Lown, J. W. Current Med. Chem. 1999, 6, 1.
2. Schmitz, F. J.; Agarwal, S. K.; Gunasekera, S. P. J. Am.
Chem. Soc. 1983, 105, 4835.
3. De Guzman, F. S.; Carte, B.; Troupe, N.; Faulkner, D. J.;
Harper, M. K.; Concepcion, G. P.; Mangalindan, G. C.;
Matsumoto, S. S.; Barrows, L. R.; Ireland, C. M. J. Org.
Chem. 1999, 64, 1400.
4. Tasdemir, D.; Marshall, K. M.; Mangalindan, G. C.; Con-
cepcion, G. P.; Barrows, L. R.; Harper, M. K.; Ireland, C. M.
J. Org. Chem. 2001, 66, 3246.
5. (a) Echevarren, A. M.; Stille, J. K. J. Am. Chem. Soc. 1988,
110, 4051. (b) Kubo, A.; Nakahara, S. Heterocycles 1988, 27,
2095. (c) Prager, R. H.; Tsopelas, C. Heterocycles 1989, 29,
847.
6. Kitahara, Y.; Tamura, F.; Kubo, A. Chem. Pharm. Bull.
1994, 42, 1363.
7. Kitahara, Y.; Tamura, F.; Kubo, A. Tetrahedron Lett.
1997, 38, 4441.
8. (a) Potts, K. T.; Walsh, E. B.; Bhattacharjee, D. J. Org.
Chem. 1987, 52, 2285. (b) Nebois, P.; Fillion, H. Tetrahedron
Lett. 1991, 32, 1307. (c) Nebois, P.; Carneiro do Nascimento,
S.; Boitard, M.; Bartoli, M. H.; Fillion, H. Pharmazie 1994,
49, 819. (d) Bouaziz, Z.; Nebois, P.; Fillion, H.; Luche, J. L.;
Jenner, G. H. Tetrahedron 1995, 51, 4057.
9. Delfourne, E.; Darro, F.; Bontemps-Subielos, N.; Dec-
aestecker, C.; Bastide, J.; Frydman, A.; Kiss, R. J. Med.
Chem. 2001, 44, 3275.
In vitro characterization of drug-induced effects with
respect to human cancer cell line growth
Six human tumor cell lines were obtained from the
American Type Culture Collection (ATCC, Manassas,
VA, USA). These included two glioblastomas (U-373MG
and U-87 MG), two colon (HCT-15 and LoVo), one
non-small-cell-lung (A549), and one bladder (J82) cancer
10. Severin, T.; Wanninger, G.; Lerche, H. Chem. Ber. 1984,
117, 2875.

C. Brahic et al. / Bioorg. Med. Chem. 10 (2002) 2845–2853
2853
11. Kitahara, Y.; Onikura, H.; Shibano, Y.; Watanabe, S.;
Mikami, Y.; Kubo, A. Tetrahedron 1997, 53, 6001.
12. Ghosh, S.; Schlosser, M. J. Fluorine Chem. 1994, 67, 53.
13. Blanco, M. M.; Avendano, C.; Menendez, J. C. Synlett
2000, 689.
Tokyo, Japan, 1999. Stewart, J. J. P. J. Comput. Chem.
1989, 10, 209.
19. Ryu, C. K.; Lee, I. K.; Jung, S. H.; Kang, H. Y.; Lee,
C. O. Med. Chem. Res. 2000, 10, 40 .
20. Weinstein, J. N.; Kohn, K. W.; Grever, M. R.; Viswa-
nadhan, V. N.; Rubinstein, L. V.; Monks, A. P.; Scudiero,
D. A.; Welch, L.; Koutsoukos, A. D.; Chiausa, A. J.; Paull,
K. D. Science 1992, 258, 447.
21. Pauwels, O.; Kiss, R.; Pasteels, J. L.; Atassi, G. Pharm.
Res. 1995, 12, 1011.
14. Bracher, F. Heterocycles 1989, 29, 2093.
15. Kobayashi, J.; Cheng, J. F.; Nakamura, H.; Ohizumi, Y.;
Hirata, Y.; Nozoe, S. Tetrahedron Lett. 1988, 29, 1177.
16. Delfourne, E.; Bontemps-Subielos, N.; Darro, F.; Bastide,
J.; Kiss, R.; Frydman, A. International Patent Application
WO0055160, 21 September 2000.
17. Alvarez, M.; Feliu, W.; Joule, J. A.; Fernandez-Puentes,
J. L. Eur. J. Org. Chem. 2000, 849.
18. Stewart, J. J. P. MOPAC 2000; Fujitsu Limited:
22. Camby, I.; Salmon, I.; Danguy, A.; Pasteels, J. L.;
Brotchi, J.; Martinez, J.; Kiss, R. J. Natl. Cancer Inst. 1996,
88, 594.