Synthesis and antiplasmodial activity of new indolone N-Oxide derivatives

Article abstract of DOI:10.1021/jm901300d

A series of 66 new indolone-N-oxide derivatives was synthesized with three different methods. Compounds were evaluated for in vitro activity against CQ-sensitive (3D7), CQ-resistant (FcB1), and CQ and pyrimethamine cross-resistant (K1) strains of Plasmodium falciparum (P.f.), aswell as for cytotoxic concentration (CC₅₀) on MCF7 and KB human tumor cell lines. Compound 26 (5-methoxy-indolone-N-oxide analogue) had the most potent antiplasmodial activity in vitro (<3 nM on FcB1 and = 1.7 nM on 3D7) with a very satisfactory selectivity index (CC₅₀ MCF7/IC₅₀ FcB1: 14623; CC₅₀ KB/IC₅₀ 3D7: 198823). In in vivo experiments, compound 1 (dioxymethylene derivatives of the indolone-N-oxide) showed the best antiplasmodial activity against Plasmodium berghei, 62% inhibition of the parasitaemia at 30 mg/kg/day. 2009 American Chemical Society.

Full text of DOI:10.1021/jm901300d

J. Med. Chem. 2010, 53, 699–714 699
DOI: 10.1021/jm901300d
Synthesis and Antiplasmodial Activity of New Indolone N-Oxide Derivatives
Franc-oise Nepveu,*,†,‡ Sothea Kim,†,‡ Jeremie Boyer,†,‡ Olivier Chatriant,§ Hany Ibrahim,†,‡ Karine Reybier,†,‡
Marie-Carmen Monje,†,‡ Severine Chevalley,†,‡ Pierre Perio,†,‡ Barbora H. Lajoie,†,‡ Jalloul Bouajila,†,‡ Eric Deharo,†,‡
Michel Sauvain,†,‡ Rachida Tahar, Leonardo Basco, Antonella Pantaleo,^ Francesco Turini,^ Paolo Arese,^
Alexis Valentin,†,‡ Eloise Thompson,# Livia Vivas,# Serge Petit,§ and Jean-Pierre Nallet§
†Universiteꢀ de Toulouse, UPS, UMR 152 (Laboratoire de Pharmacochimie des Substances Naturelles et Pharmacophores Redox),
F-31062 Toulouse cedex 9, France, ‡IRD, UMR 152, F-31062 Toulouse cedex 9, France, §IDEALP-PHARMA, Ba^timent CEI, 66 Bd Niels Bohr,
BP 2132, 69603 Villeurbanne cedex, France, Organisation de Coordination pour la Lutte Contre les Endeꢀmies en Afrique Centrale (OCEAC),
BP 288, Yaoundeꢀ, Cameroun, ^University of Turin Medical School (Dipartimento di Genetica, Biologia e Biochimica), Via Santena 5 bis,
10126 Torino, Italy, and #London School of Hygiene and Tropical Medicine (LSHTM), Keppel Street, London C1E 7HT, United Kingdom
Received September 1, 2009
A series of 66 new indolone-N-oxide derivatives was synthesized with three different methods. Compounds
were evaluated for in vitro activity against CQ-sensitive (3D7), CQ-resistant (FcB1), and CQ and pyri-
methamine cross-resistant (K1) strains of Plasmodium falciparum (P.f.), as well as for cytotoxic concentration
(CC50) on MCF7 and KB human tumor cell lines. Compound 26 (5-methoxy-indolone-N-oxide analogue)
had the most potent antiplasmodial activity in vitro (<3 nM on FcB1 and = 1.7 nM on 3D7) with a very
satisfactory selectivity index (CC50 MCF7/IC50 FcB1: 14623; CC50 KB/IC50 3D7: 198823). In in vivo
experiments, compound 1 (dioxymethylene derivatives of the indolone-N-oxide) showed the best antiplas-
modial activity against Plasmodium berghei, 62% inhibition of the parasitaemia at 30 mg/kg/day.
1. Introduction
Our interest in the nitrone group [CdNþ-O-] the chemical
scaffold of radical spin traps led us to study the redox proper-
ties of some indolone derivatives.11,12 We observed that
2-alkyl-indolone-N-oxides13 were rapidly reduced in solution,
leading to radical nitroxide intermediates, while 2-aryl indo-
lone-N-oxides were stable in solution. Moreover, they gave
very stable spin adducts by trapping oxygen or carbon-
centered free radicals chemically or enzymatically produced.
Previously, L. Lunazzi et al.14 had also observed that under
certain conditions, very long-lived radicals were generated via
a proton attack from the solvent by heating up these indo-
lones. Such chemical properties of N-oxide compounds were
used to design bioreducible prodrugs such as tirapazamine
derivatives with selective toxicity toward hypoxic cells.15 This
selectivity is based on the net reduction of the N-oxide moiety
in the absence of oxygen, in a one or two-electron process-
approach, by reductive enzymes. In addition, R. Danieli
et al.16 showed that indolone derivatives can be easily
reduced by thiols. Recently, indazole N-oxide derivatives
have been reported as antiprotozoal agents with a reduction
mechanism.17,18 Moreover, antituberculosis or antimycobac-
terial activities have been reported for some indolone-N-
oxides.19,20 Considering indolone-N-oxide properties, we hy-
pothesized that their capacities to be reduced and generate
stable radical intermediates and their reactivity with thiols
may be critical for parasite growth within red blood cells
(RBC). RBCs contain high levels of reductases and reduc-
tants, such as glutathione, and display a decrease in molecular
oxygen concentration and an increase in reactive oxygenated
species during parasite maturation. The preliminary assays in
vitro were highly promising.
Because of multiple factors, the burden of malaria is
currently increasing, the most important negative point being
the development by P. falciparum (P.f.a) of resistance to
cheap and effective drugs like chloroquine (CQ). One of the
innovative approaches to antimalarial drug discovery is based
on the identification of new drug targets and attendant
inhibitor design. However, as recently reviewed,1-4 most of
clinically available antimalarial drugs were developed using
pregene technology-era pharmacochemical approaches and
the action mechanisms for the majority have not as yet
been clarified.
Our approach was to identify novel antimalarial syn-
thetic molecules by focusing our strategy on a chemical
substructural motif (scaffold) critical for parasite growth.
In antimalarial activity of artemisinin-like compounds,5
the endoperoxide group plays a key role. It is the validated
chemical scaffold and, because this substructural motif is a
reducible or a pro-oxidant group, it can also be called a
redox pharmacophore. From the pharmacological point of
view, other powerful reducible chemical groups may also
be able to disrupt cellular redox homeostasis. In past
decades, many mono- and di-N-oxides derivatives have
been reported for their antimalarial activities.6-10 In these
chemical series, the N-oxide group could be identified as a
reducible group.
*To whom correspondence should be addressed. Phone: þ33-5-62-
25-68-69. Fax: þ33-5-62-25-98-02. E-mail: nepveu@cict.fr.
aAbbreviations: CQ, chloroquine; P.f., Plasmodium falciparum;
RBC, red blood cells; CC50, 50% cytotoxic concentration; po, per oral
route; 4-DMAP, 4-dimethylaminopyridine; DMSO, dimethyl sulfoxide;
EFS, French blood bank; LDH, lactate dehydrogenase.
We therefore undertook the synthesis and study of in vitro
and in vivo antiparasitic activity against Plasmodium spp. of a
r
2009 American Chemical Society
Published on Web 12/16/2009
pubs.acs.org/jmc

700 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2
Nepveu et al.
reactions were undertaken under microwave conditions; pro-
tocols were specifically adjusted for each reaction to obtain
the expected compounds in a few minutes from low to
moderate yield. Unfortunately, depending on the different
substituents on aromatic rings, this easy protocol also failed in
the synthesis of some designed indolones.
All of the compounds were chemically characterized by
liquid chromatography (TLC and HPLC), infrared (IR),
UV-visible, NMR (1H and 13C NMR), and mass spectro-
metry as well as elemental analysis. LogP values were calcu-
lated with VVCLAB software29 and ranging between 0.62 and
4.79. Compound 57 is the most lipophilic in the series, and
compound 51, the least one. It appeared that lipophilicity has
no detrimental effect on antiplasmodial activity (Table 1).
Compounds were very poorly soluble in water except for salt
forms (hydrochloride salt of the compounds 26, 28, and 30).
Studies of compound stability in solution showed that the
compounds were stable in aprotic solvents (CH3CN,
(CH3)2SO) and less stable in polar and protic solvents
(MeOH, H2O) (data not shown).
Figure 1. Structure of indolone N-oxide derivatives (i: compounds
from 1 to 66).
series of indolone N-oxides (Figure 1). We describe hereafter the
synthesis and in vitro antiplasmodial activities assessed in CQ-
sensitive (3D7), CQ-resistant (FcB1) and CQ and pyrimetha-
mine cross-resistant (K1) strains of Plasmodium falciparum
(P.f.). Hit optimization was carried out based on antiparasitic
efficacy against the three strains, cytotoxicity against human
cell lines, as well as physicochemical parameters. The study of
in vivo activities was performed on Plasmodium berghei
(NK65 and ANKA) murine models.
2. Chemistry
A series of 66 analogues of indolone N-oxides were pre-
pared (Table 1). Three synthetic pathways were considered
and successfully used in this work, which led to the desired
indolone N-oxide derivatives (i: compounds from 1 to 66)
(Figure 1). Method 1 is based on 1,2-diketone intermedi-
ates b, which are cyclized via nitro group reduction to
generate the corresponding indolone N-oxides i.11,12 The
1,2-diketones were prepared by permanganate oxidation of
a styrene precursor a, which is easily obtained by Wittig
olefination of nitrobenzaldehydes with phosphonium ylides
(Scheme 1).
Structure-Activity Relationship (SAR): Effect of Substit-
uents on the Biological Activity. The compounds were de-
signed according to the structure-activity relationship in
considering the structure of the first hit identified, 1. In
addition, commercial chemical libraries were examined
and, according to their molecular profile, some compounds
were selected for testing. These procedures were helpful in
pharmacophore determinations. All prepared compounds
were tested at the primary level on a CQ-resistant P. falci-
parum strain (FcB1). This initial step identified the substi-
tuent effects of the indolone-N-oxide at either position 2, 5,
or 6 (Figure 2).
Figure 2 presents the basic structure of indolone-N-oxide
derivatives. The phenyl group (A) of the indolone moiety was
found to be essential for antiplasmodial activity. Replace-
ment of the phenyl group (A) by either pyridine (N at
position 4 of the indolone moiety, compound 66) or by
pyridinium-N-oxide (N at position 5 of the indolone moiety,
compound 65) led to significant activity loss, IC50 was from
10- to 20-fold higher than their corresponding indolone
analogue, compound 64.
As shown in Figure 3, replacement of the phenyl group
(R3) (at position 2 of the indolone moiety) by an aliphatic
chaine (n-propyl, 50, or i-butyl, 49) led to a significant loss of
activity. Whereas functionalization of the phenyl moiety
(R3) by groups having a mesomeric effect (either þM: F,
Cl, OCH3, OPh or -M: NO2) improved the activity, while
groups with inductive effect (either þI: CH3, C2H5, CH2OH
or -I: CF3) decreased the activity.
As shown in Figure 4, comparable results were obtained
with dioxymethylene derivatives of indolone-N-oxide, with
the exception of compound 54 (IC50 measured by LDH
method, see Supporting Information).
Method 221 is a one-pot procedure based on Sonogashira
coupling of o-iodo nitro-aryles e with terminal alkynes f and
concomitant cyclization of o-(arylalkynyl)-nitroaryles inter-
mediates g into indolone N-oxides (Scheme 2).22,23 Method 3
derives from optimization of method 2 with the aim of
improving yields, while increasing reaction kinetics. Speci-
fically, method 3 was divided into two substeps: the first
involved Sonogashira coupling as in method 2 as well as
intermediate extraction, followed by an intramolecular
cyclization using 4-dimethyl amino pyridine (4-DMAP) in
refluxing pyridine (Scheme 3).24-27 In some cases, using
AuBr3 as catalyst accelerated the reaction rate and im-
proved the yield.28 It must be mentioned that several
o-bromo-nitroaryles (e) and 1-alkynes (f) used in this study
to prepare indolone N-oxide analogues were also of syn-
thetic origin (Schemes 2, 3). Compounds e were obtained
by electrophilic nitration of o-bromoaryles aldehydes
using fuming nitric acid/acetic acid mixture. Synthesis of
1-alkynes (f) involved the preparation of 1-(trimethylsilyl)-
alkynes via Sonogashira coupling, followed by a desilyla-
tion step. Finally, O-benzyl-4-iodo-3-nitrophenol used
to synthesize compounds 57 and 58 was obtained by
O-benzylation of 4-iodo-3-nitrophenol (Scheme 4)
Figure 5 presents antiplasmodial activity of 2(4-meth-
oxyphenyl)-indolone-N-oxide analogues. Substitutions at
either position 5 or 6 of phenyl group (A) of the indolone
moiety had no marked effect on IC50. Therefore, substitu-
tions at R3 affected antiplasmodial activity to a greater
extent than substitutions at the phenyl group of the indolone
moiety (cf. Figures 3 and 4).
Figure 6 presents 5-methoxy-indolone-N-oxide analo-
gues. Contrary to previous results (Figures 3, 4), sub-
stitutions at R3 position had a slight effect on antiplas-
modial activity, except when the substitution occurred at
In comparing the methods used to synthesize indolone N-
oxides, it is worth pointing out that method 3 requires 0.5-
17 h to prepare a target molecule, albeit of low (20-30%)
chemical yields, whereas methods 1 and 2 may require 1-
2 weeks. However, method 1 itself is of key importance when
considering that some stable o-(alkylalkynyl)-nitroaryles in-
termediates g failed to spontaneously cyclize into indolone
N-oxides under Sonogashira conditions.24
To overcome difficulties in obtaining some targeted
compounds according to previous methods, Sonogashira

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2 701
Table 1. Structure of Indolone N-Oxide Analogues Obtained and Their in Vitro Antiplasmodial and Cytotoxic Activities
a
Log Pcalc
(VCCLAB)
IC50 (nM)
FcB1 strain
CC50 (μM)b
MCF7
selectivity index
MCF-7/FcB1
compd
R1
R2
O-CH2-O
R3
1
40-chlorophenyl
2.07
3.51
1.29
2.01
3.03
2.51
3.00
3.14
2.58
2.25
3.32
2.57
3.18
2.66
1.98
2.23
2.00
2.25
3.26
3.03
4.06
2.04
3.02
3.51
2.90
2.30
2.81
3.34
2.25
1.58
2.03
2.72
2.59
2.24
2.88
1.91
2.74
2.05
2.60
2.48
1.81
3.63
2.62
2.80
1.92
2.24
1.93
1.97
1.61
1.41
0.62
1.96
2.06
1.58
2.45
1.96
4.79
4.37
3.31
2.81
2.71
2.02
1.5
75 ( 63
264 ( 60
3950 ( 50
195 ( 20
560 ( 15
288 ( 30
165 ( 40
155 ( 20
210 ( 10
265 ( 29
50 ( 39
150 ( 91
100 ( 7
195 ( 2
156 ( 57
1770 ( 20
184 ( 53
52 ( 48
15.9
11.1
39.5
>39.5 c
15.3
25.6
12.2
13.7
21.5
nd
212
42
2
H
H
40-phenoxyphenyl
40-hydroxymethyl phenyl
40-methoxyphenyl
40-60-methoxy-naphthalen-2-yl
40-60-methoxy-naphthalen-2-yl
40-phenoxyphenyl
30, 40-dichlorophenyl
40-chlorophenyl
3
4
5
H
H
H
H
H
H
10
>202
66
89
6
7
8
O-CH2-O
O-CH2-O
H
74
88
H
9
OCH3
OCH3
OCH3
H
H
OCH3
H
102
nd
270
69
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
40-tolyl
30-trifluoromethyl-40-chlorophenyl
40-chlorophenyl
30, 40-dichlorophenyl
30, 40-dichlorophenyl
40-ethoxyphenyl
13.5
10.4
81.0
7.4
OCH3
H
O-CH2-O
O-CH2-O
H
OCH3
81
38.1
332
8.5
51.9
15
13.4
>37.5 c
3.1
14.4
12.1
>35.3 c
>31.0 c
7.7
H
40-tolyl
phenyl
40-tolyl
OCH3
OCH3
H
H
73
H
CF3
H
>721
16.5
107
19
>882
>136
40
40-chlorophenyl
186 ( 15
135 ( 45
630 ( 248
40 ( 39
227 ( 9
193 ( 55
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
H
40-methoxy-naphthalen-2-yl
40-chlorobiphenyl-4-yl
40-methoxyphenyl
20, 40-dichlorophenyl
40-phenoxyphenyl
40-trifluoromethoxyphenyl
40-dimethylaminophenyl
40-isopropoxyphenyl
40-diethylaminophenyl
40-methoxy-30-tolyl
40-aminophenyl
H
H
H
H
H
H
20 ( 0
<3
8.3
415
>14,623
>1,889
134
>764
528
83.3
58.8
116.5
75
43.9
>32.1c
17.6
>33.6 c
12.7
8.8
H
H
H
17 ( 2
131 ( 33
44 ( 4
24 ( 19
106 ( 50
135 ( 95
191 ( 0
198 ( 26
133 ( 50
206 ( 9
43 ( 23
256 ( 22
21 ( 1
H
OCH3
CF3
Cl
CH3
OCF3
CH3OCO
H
40-methoxyphenyl
40-methoxyphenyl
40-methoxyphenyl
40-methoxyphenyl
40-methoxyphenyl
40-mMethoxyphenyl
40-trifluoromethylphenyl
40-acetophenyl
H
7.9
H
H
H
22.2
14.9
11.7
40.2
11.7
40.2
5.2
88.1
194.9
273
29.1
248
702.7
18.5
53.9
nd
3,425
25.6
0.3
45.5
63.2
<9
2
H
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
H
H
H
H
H
30-chlorophenyl
30-methyl-40-tolyl
40-acetamidophenyl
40-tert-butylphenyl
20-chlorophenyl
37 ( 28
26
7.4
400 ( 232
204 ( 14
1130 ( 25
3 ( 1
H
H
H
11
nd
10.3
7.1
40-chloro-30-tolyl
40-carboxymethylphenyl
20-methyl-40-methoxyphenyl
20-thiophen-3-yl
40-methoxyphenyl
i-butyl
n-propyl
H
H
H
276 ( 45
1043 ( 95
270 ( 16
345 ( 81
>4500 c
12160 ( 500
3560 ( 400
889 ( 88
120 ( 29
1710 ( 30
130 ( 3
0.3
12.3
21.8
43.1
26.4
59.0
19.5
8.7
NO2
H
H
O-CH2-O
H
H
ethyl
phenyl
16
22
72.5
nd
38
8.4
3.2
H
H
H
H
40-fluorophenyl
O-CH2-O
Cl
40-fluorophenyl
nd
4.9
2.8
2.2
nd
2.0
>39.8 c
34.3
nd
H
H
20-thiophen-3-yl
H
40-nitrophenyl
56 ( 4
H
H
H
PhCH2O
PhCH2O
H
40-phenoxyphenyl
60-methoxy-naphthalen-2-yl
40-chloro-30-trifluoromethylphenyl
40-ethylphenyl
665 ( 135
730 ( 30
1045 ( 35
2750 ( 150
1170 ( 80
1890 ( 240
12720 ( 400
272 ( 16
151 ( 6
nd
1.9
H
H
H
H
OCH3
>14
29
nd
nd
66
167
1633
40-trifluoromethylphenyl
OCH3
O-CH2-O
H
phenyl
phenyl
40-chlorophenyl
nd
H
2.54
4.63
2.41
17.8
19.4
9.8
chloroquine
sodium artesunate
6 ( 3
a LogP calculated with VCCLAB (http://www.virtuallaboratory.org/lab/alogps/start.html). b The drug concentration needed to cause 50% decrease
of the cellular viability. c The highest tested concentration.

702 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2
Nepveu et al.
Scheme 1. Method 1 Used to Synthesize Indolone N-Oxide Derivatives (R3 = Alkyl)a
a Reagents and conditions: (i) Ph3PCH2R3Br, NaOH, CH2Cl2, Bu4NþBr-; (ii) KMnO4, acetic anhydride, 0 ꢀC; (iii) Zn, NH4Cl, tetrahydrofuran,
CH2Cl2.
Scheme 2. Method 2 Used to Synthesize Indolone N-Oxide derivatives (R3 = Aryl)a
a Reagents and conditions: (i) (CH3)3Si-CtCH, Pd[(C6H5)3P]4, CuI, Et3N; (ii) CH3OH, K2CO3; (iii) CH3COOH, HNO3 fuming; (iv) Pd(PPh3)2Cl2,
CuI, NEt3, N2, rt.
Scheme 3. Method 3 Used to Synthesize Indolone N-Oxide Derivatives (R3 = Aryl)a
a Reagents and conditions: (i) (CH3)3Si-CtCH, Pd[(C6H5)3P]4, CuI, Et3N; (ii) CH3OH, K2CO3; (iii) CH3COOH, HNO3 fuming; (iv) Pd(PPh3)2Cl2,
CuI, NEt3, N2, rt; (v) pyridine, 4-dimethylaminopyridine, reflux 140 ꢀC.
Figure 2. Indolone-N-oxide analogues.
Scheme 4. Synthesis of 4-Iodo-3-nitro Benzyloxy-benzene
(57e, 58e)a
Figure 3. Comparison of antiplasmodial activity (IC50 [nM], strain
FcB1) of 2-substituted indolone-N-oxides.
a Reagents and conditions: (i) CH3CH2OH, K2CO3, 60 ꢀC.
position 2 of the phenyl group (compounds: 43, 46). X-ray
crystallographic studies of a similar compound (2-pheny-
lisatogen) showed that isatogen and phenyl (R3) rings were
almost fully coplanar.30 Hence substitution at position 2 led
to a steric effect (hindrance) affecting the planarity system.
Consequently, mesomeric-effect transmission from isato-
gen ring to phenyl ring would be inefficient due to low
conjugation.
The conjugated system between nitrone group and ketonic
function was found to be essential for an optimal in vitro
antiplasmodial activity, while reduction of either nitrone or
Figure 4. Comparison of antiplasmodial activity (IC50 (nM) FcB1)
of dioxymethylene derivatives of indolone-N-oxides.

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2 703
ketonic function led to a dramatic loss of activity (data
not shown).
pyrimethamine-resistant (K1) strains of P.f. (Table 2). All
tested compounds were found to be more potent than CQ
against K1 strain. Three of them showed lower IC50 than that
of artesunate against either 3D7 or K1 strains. Many com-
pounds demonstrated equipotent activity against both CQ-
sensitive and CQ-resistant strains. A comparison of IC50
values between P.f.-resistant and sensitive strains suggested
relatively low levels of cross-resistance to CQ.32 Resistance
index (RI) values, calculated from comparison of IC50 values
of resistant K1 or FcB1 and sensitive strains of P. falciparum
3D7 such as IC50 (K1 or FcB1)/IC50(3D7), were found to be
lower than those for CQ in the range 0.1-5.3 (K1) (Table 2).
These compounds showed much lower resistance indices
than those of CQ, suggesting that potential resistance to
indolone structure is independent of chloroquine-resistance
pathways.
Cytotoxicity assayed on mammalian MCF7 and KB cell
lines were in the μmolar range. CC50 values (MCF7) varied
from 7.7 to 59.0 μM (Table 1) and from about 1 to 874 μM
(KB) (Table 2). Most of the analogues tested at the second-
ary level had selectivity index greater than 200 (KB/3D7).
Compounds that showed high selectivity (high selectivity
index) are expected to offer the potential for safer therapy
and constitute suitable candidates for further pharmacolo-
gical studies.
In Vivo Antiplasmodial Activity against Plasmodium ber-
ghei. The best in vitro profiles displayed by indolone-N-oxide
analogues, based on their potency, cytotoxicity, and selec-
tivity index, led to the selection of some compounds for
further in vivo evaluation against P. berghei strain. For
compound 1, the parasitaemia clearance in vivo in P. berghei
(NK65 strain) was 81% at 48 mg/kg in a 4-day test. No acute
toxicity was observed in Swiss female mice at a dose of e140
mg/kg (Table 3). Higher doses were not tested due to the
lower solubility of the compound in DMSO at higher con-
centrations.
It was decided to analyze the possible capacity of the
indolone-N-oxide derivatives to interact by stacking with
heme. The malaria parasite does indeed break up hemoglo-
bin inside RBC, releasing toxic-free heme. The parasite
protects itself against heme toxicity by biocrystallizing
around 30% of it into insoluble hemozoin. A compound
that can inhibit this biocrystallization may be a potential
antimalarial drug, as can the antimalarial chloroquine. The
planarity of a molecule (conjugated system) may be indica-
tive of a possible interaction with hemozoin. Several tests
detecting such possible chemical drug/heme interaction are
available. We used the assay developed by E. Deharo et al.31
No relationship between antiplasmodial activity (IC50/
FcB1) and heme-inhibiting polymerization capacity in the
biochemical assay was found in a series of seven compounds
(see Supporting Information).
3. Biology
In Vitro Antiplasmodial Activity against P. falciparum.
Initial testing was carried out on a chloroquine (CQ)-resis-
tant strain of P.f. (Table 1). Several analogues displayed
antiplasmodial activity in the low nanomolar range. Four-
teen out of 66 synthetic compounds were found to be more
potent in vitro against FcB1 than was chloroquine, with
IC50s lower than 100 nM (Table 1). Their cytotoxicity was
evaluated on the MCF7 cell line. Selectivity index was
defined as the ratio of CC50 value in MCF7 cells to IC50
value in CQ-resistant P.f. strain FcB1.
The best analogues, based on their selectivity index
(SI > 200; MCF7/FcB1), were chosen for assays at the
secondary level on a CQ-sensitive (3D7) and CQ and
As shown in Table 4, compound 1 had the best anti-
plasmodial activity by intraperitoneal (ip) route of admin-
istration, while it showed the lowest activity by the oral
route of administration. This could be explained by the
first-pass effect, which leads to an extensive degradation
of compound 1 when administered by the oral route. These
results were confirmed by the metabolic studies of these
compounds within mouse liver microsomes (Table 5).
Further details and protocols are available in the Support-
ing Information. Compound 1 has the lowest half-life
time, whereas compound 4 has the highest one. There-
fore, no significant difference was observed between the
Figure 5. Comparison of antiplasmodial activity (IC50 (nM) FcB1)
of 2(4-methoxyphenyl)-indolone-N-oxide analogues.
Figure 6. Comparison of antiplasmodial activity (IC50 (nM) FcB1) of 5-methoxy-indolone-N-oxide analogues.

704 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2
Nepveu et al.
Table 2. Selected Indolone N-Oxide Analogues and in Vitro Antiplasmodial and Cytotoxic Activities
IC50 (nM)
3D7 strain
IC50
(nM) K1 strain
CC50
(μM) KB
selectivity
index KB/3D7a
resistance
index K1/3D7
resistance
index FcB1/3D7
compound
1
58 ( 17
148 ( 12
101 ( 32
90 ( 28
72 ( 17
285 ( 57
98 ( 31
62 ( 37
69 ( 48
27 ( 3
82 ( 38
37 ( 48
30 ( 2
1.7
88 ( 37
60 ( 4
124 ( 44
40 ( 12
35 ( 8
138 ( 14
80 ( 39
59 ( 32
5 ( 5
27.0
ndb
450
nd
1.5
0.4
1.3
1.8
1.9
2.6
4.0
0.6
1.6
3.4
0.7
1.9
0.5
5.2
0.7
2.9
0.5
6.3
86.0
0.2
0.3
4.3
1.5
2
4
457.0
233.0
nd
4525
2589
nd
1.2
0.4
5
6
0.5
0.5
7
nd
nd
8
570.0
469.0
<1.4
56.0
5816
7564
<20
2074
10658
151
0.8
1.0
9
11
18
22
24
25
26
27
35
37
39
40
0.1
0.7
20 ( 8
16
874.0
5.6
0.2
<1.5
< 0.04
2.7
nd
81 ( 22
n.d.
81.3
11 ( 10
338
321
128
198823
10031
86
32
21 ( 0
<0.5
2.5
5.3
112 ( 18
1.9
16 ( 0.4
110.0
64 ( 0.3
148 ( 107
44
>220
688
980
3.8
0.7
139 ( 0
142 ( 3
35 ( 27
4 ( 3
96 ( 47
178 ( 104
850 ( 57
4 ( 3
1.2
chloroquine
sodium artesunate
1257
9045
24.3
1.0
36.18 ( 8.09
a SI 3D7 = cytotoxic (CC50) (KB)/antiplasmodial (IC50) (3D7) ratio. b nd: not determined.
Table 3. In Vivo Antimalarial Activity againt P. berghei (NK 65)
a
selectivityindex (CC50 MCF7/IC50 FcB1: > 14623; CC50 KB/
IC50 3D7: 198823).
ED50
parasitaemia
Synthesis and screening of in vitro antiplasmodial activity
and cytotoxicity of the novel series, indolone-N-oxide deriva-
tives, identified promising antiplasmodial candidates based
on their potency, selectivity index, and low cytotoxicity,
rendering them as valid new leads for synthesizing new
compounds that might improve the previous prototypes.
mg/kg/ clearance mg/kg/4
4days days ip/DMSO, %
toxicity in
vivo ip/DMSO
compd
1
16
81% at 48 mg/kg No toxicity observed for a
dose e140 mg/kg (limit of
solubility in DMSO)
chloroquine
3
100% at 10 mg/kg nd
a ED50: effective dose required to achieve 50% parasitaemia clear-
ance.
6. Materials and Methods
Commercial reagents were used as received without additional
purification. Dichloromethane, acetonitrile, cyclohexane, and
ethyl acetate were purchased from Fischer Bioblock
(Strasbourg, France). Sodium sulfate, silica, silica gel 60 F-254
plates, ethanol, and methanol were purchased from VWR Inter-
national (Strasbourg, France). Column chromatography for
purification was carried out using 200-400 mesh chromagel.
Flash chromatography was realized on CombiFlash (UVK-Lab.
Toulouse, France). For molecular mass determination and purity
control the following LC-PDA-MSn system was used (Thermo
electron Corporation), including an automatic injector with oven
(SpectraSystem AS3000), a degasser(Spectra System SCM1000),
and a quaternary pump (Spectra System P1000 XR) coupled to a
photodiode array detector PDA (Spectra System UV6000LP),
and an ion trap mass spectrometer (Finnigan LCQ Deca XP
Max), using nitrogen as a nebulizing and drying gas. Data
acquisition was performed using Finnigan Xcalibur software
(version 1.4). Chromatographic separation was performed using
analytical columns either Thermo Hypersil Hyperprep RP C-18
(8 μm, 150 mm ꢀ 4.6 mm) or Thermo BetaBasic RP C-4 (5 μm,
150 mm ꢀ 4.6 mm). For data acquisition we also used Waters
Millenium32 software, version 3.2 (Waters, St. Quentin, France).
Other mass spectrometers were employed for molecular mass
determination such as MS-Nermag R10-10 spectrometer using
electron ionization (Service Commun Toulouse, France), and
GCT 1er WATERS using electron ionization and desorption
chemical ionization (Service Commun Toulouse, France) for
high-resolution mass determination. Elemental analyses were
done using Perkin-Elmer 2400 series II CHNS/O Elemental
analyzer (LCC, Toulouse, France).
Table 4. In Vivo Antimalarial Activity against P. berghei (ANKA)
% parasitaemia inhibition (at 30 mg/kg/day)
compd
po, %a
ip, %b
1
14.5
32.4
20.9
25.1
62.1
40.5
15.3
30.4
4
26
27
a po: per oral route of administration. b ip: intrapretoneal route of
administration.
Table 5. In Vitro Analysis of Compound Stability with Mouse Liver
Microsomes
compd
half life (min)
1
<1
16
4
26
27
6
6-7
6
verapamil hydrochloride
two administration routes (ip and po) for other com-
pounds (4, 26, and 27).
5. Conclusion
The highest antiplasmodial activities were obtained for the
5-methoxy-indolone-N-oxide analogues (26, 27). Compound
26 had the most potent antiplasmodial activity in vitro (<3
nM on FcB1 and=1.7 nM on 3D7) with a very satifactory
Compound purity was determined by HPLC-UV and LC-
PDA-MS methods and was found in the range 96-99%. The

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2 705
separation was performed under a gradient mode using solvent A
(water/0.05% trifluoroacetic acid) and solvent B (CH3CN/0.05%
trifluoroacetic acid). The gradient program was the following: at
t = 0, solvents (80% A/20% B, v/v); at t = 8 min, solvents (0%
A/100% B, v/v); at t = 13 min, solvents (0% A/100% B, v/v);
column regeneration time was 7 min; the mobile phase was
delivered at a flow rate of 1 mL/min. The injected volume was
10 μL. All the compounds were analyzed using C-18 column
except the following compounds, which were analyzed using C-4
column(compounds 23, 34, 36, 37, 43, 44, 45, 47, and 48). Melting
points were determined on an Electrothermal 9300 capillary
melting point apparatus and are uncorrected. 1H and 13C
NMR spectra were recorded on an AC Bruker spectrometer at
300 or 500 MHz (1H) and 75 or 125 MHz (13C) using CDCl3,
CD3CN, or (CD3)2SO as solvents. Chemical shifts (δ) are re-
ported in parts per million (ppm) relative to tetramethylsilane (0
ppm) and the following multiplicity abbreviations were used: s,
singlet; d, doublet; t, triplet; q, quatruplet; m, multiplet; dd,
double of doublet; dt, double of triplet. IR spectra were recorded
on a Perkin-Elmer PARAGON 1000 FT-IR spectrometer.
UV-visible spectra were recorded on an Uvikon 931 Kontron
spectrometer. High resolution mass spectrometry was used for
some compounds (SCA, CNRS Vernaison, France).
6.1. Synthesis by Method 1 (Compounds 1, 15, 51-54). Gen-
eral Synthetic Procedure for Alkene Compounds a. To a solution
of o-nitrobenzaldehyde (10.7 mmol) in dichloromethane (140
mL) was added the appropriate phosphonium salt (14.6 mmol).
NaOH aqueous solution (50%) (12.8 mmol) and tetrabutyl-
ammonium chloride (54 mmol) were then added. The mixture
was stirred at room temperature until complete disappearance
of the starting product. It was then extracted by dichloro-
methane; the organic phase was washed with brine and dried
over anhydrous MgSO4 and concentrated under reduced pres-
sure. The crude material was purified by column chromatogra-
phy (SiO2) using dichloromethane as the eluant to give a mixture
of E/Z-diastereoisomers.
(d, J = 12.0 Hz, 1H, cis), 7.08-8.13 (m, H arom þ H alkene,
trans). MS (EI): m/z 225 (Mþ.), 208, 180, 178, 176, 165, 152, 139,
119, 105, 92, 91, 77, 63, 51, 39.
5-[(E/Z)-2-(4-Fluorophenyl)ethynyl]-6-nitro-phenyl (53a). Yield
68.9%. Rf 0.71 and 0.74 (cyclohexane/ethyl acetate, 70:30, v/v). IR
(KBr) cm-1: 3070, 2923, 2851, 1630, 1601, 1570, 1628, 1476, 1446,
1
1343, 1302, 1232, 1158, 1097, 1013, 962, 856, 786, 698, 533. H
NMR (CDCl3, 200 MHz): δ ppm (cis) 6.85 (dd, J = 8.5 Hz, J =
2.1 Hz, 2H), 6.90 (d, J= 3.1 Hz, 1H), 7.05 (m, 4H), 7.15 (d, J =9.0
Hz, 1H), 7.4 (dd; J = 7.1 Hz, J = 2.3 Hz, 2H); (trans) δ 6.7 (d, J =
12.2 Hz, 1H), 7.23 (dd, J = 4.6 Hz, J = 2.1 Hz, 2H), 7.55 (m, 4H),
7.98 (d, J = 11.9 Hz, 1H), 8.1 (dd, J = 9.5 Hz, J = 2.4 Hz, 2H).
5[(E/Z)-2-(4-Fluorophenyl)ethynyl]-6-nitro-1,3-benzodioxol
(54a). Yield 85.5%. Rf 0.60 and 0.50 (cyclohexane/ethyl acetate,
70:30, v/v). IR (KBr) cm-1: 3125, 3015, 2928, 1614, 1608, 1519,
1506, 1486, 1321, 1261, 1233, 1158, 1027, 965, 812, 680, 521. 1H
NMR (CDCl3, 200 MHz): (cis) δ 6.07 (s, 2H), 6.56 (s, 1H), 6.64
(d, J = 11.9 Hz, 1H), 6.92 (dd, J = 3.3 Hz, 2H), 7.06 (dd, J = 3.6
Hz, 2H), 7.09 (d, J = 3.6 Hz, 1H), 7.52 (s, 1H); (trans) δ 6.13 (s,
2H), 6.80 (s, 1H), 6.87 (dd, J = 5.5 Hz, 2H), 7.02 (d, J = 5.2 Hz,
1H), 7.48 (dd, J = 5.5 Hz, 2H), 7.53 (s, 1H), 7.62 (d, J = 8.2 Hz,
1H).
General Synthetic Procedure for Diketone Compounds b. To a
stirred cooled solution (0-5 ꢀC) of alkene (4.8 mmol) in acetic
anhydride (32 mL) was added KMnO4 (19.2 mmol) in small
portions over a period of 20 min. After completion of the
addition, the mixture was stirred in a cooling bath for 2 h. The
reaction was then stopped by addition of ethyl acetate/cyclo-
hexane (1:1, v/v) (32 mL) and an ice-cold solution of sodium
bisulfite 10%. After stirring in the cooling bath for several
minutes, the mixture was extracted by dichloromethane, and
the organic phase was washed by an aqueous NaOH solution
(1N), water, and dried over anhydrous MgSO4. It was then
concentrated by evaporation. The crude material obtained was
purified by column chromatography (SiO2) using cyclohexane/
dichloromethane.
1-(4-Chlorophenyl)-2-(6-nitro-1,3-benzodioxol-5-yl)-1,2-etha-
nedione (1b). Yield 50%. Rf 0.51 (cyclohexane/ethyl acetate,
70:30, v/v). IR (KBr) cm-1: 1691, 1673, 1583, 1506, 1483, 1424,
5-[(E/Z)-2-Chlorophenyl)ethynyl]-6-nitro-1,3-benzodioxol (1a).
Yield 87%. Rf 0.64 and 0.54 (cyclohexane/ethyl acetate, 70:30, v/
v). IR (KBr) cm-1: 1613, 1519, 1499, 1486, 1434, 1323, 1262,
1090, 1028, 930, 894, 815. 1H NMR (CDCl3, 200 MHz): (cis) δ
6.06 (s, 2H), 6.53 (s, 1H), 6.62 (d, J = 12.0 Hz, 1H), 6.85 (d, J =
12.0 Hz, 1H), 6.99 (d, J = 8.5 Hz, 2H), 7.15 (d, J = 8.5 Hz, 2H),
7.61 (s, 1H); (trans) δ 6.11 (s, 2H), 6.87 (d, J = 16.0 Hz, 1H), 7.07
(s, 1H), 7.31 (d, J = 8.5Hz, 2H), 7.42, (d, J = 8.5 Hz, 2H), 7.49 (s,
1H), 7.60 (d, J = 16.0 Hz, 1H).
1
1325, 1266, 1148, 1094, 1030, 927, 881, 872, 858, 817, 781. H
NMR (CDCl3, 200 MHz): δ ppm 6.21 (2H, s), 7.06 (s, 1H), 7.48
(d, J = 8.4 Hz, 2H), 7.56 (s, 1H), 8.09 (d, J = 8.4 Hz, 2H).
1-(4-Ethoxyphenyl)-2-(6-nitro-1,3-benzodioxol-5-yl)-1,2-etha-
nedione (15b). Yield 25%. Rf 0.33 (cyclohexane/ethyl acetate,
70:30, v/v). IR (KBr) cm-1: 1690, 1649, 1598, 1565, 1514, 1483,
1426, 1327, 1265, 1171, 1140, 1073, 1031, 922, 865, 784, 756. 1H
NMR (CDCl3, 200 MHz): δ ppm 1.46 (t, J = 7.0 Hz, 3H), 4.14
(q, J = 7.03 Hz, 3H), 4.14 (q, J = 7.0 Hz, 2H), 6.23 (s, 2H), 6.98
(d, J = 9.0 Hz, 2H), 7.08 (s, 1H), 7.58 (s, 1H), 8.16 (d, J = 9.0
Hz, 2H).
1-(4-Ethyl)-2-(6-nitro-1,3-benzodioxol-5-yl)-1,2-ethanedione
(51b). Yield 50%; mp 81-82 ꢀC. Rf 0.41 (cyclohexane/ethyl
acetate, 70:30, v/v). IR (KBr) cm-1: 2978, 2916, 1706, 1607,
1524, 1509, 1483, 1426, 1368, 1332, 1275, 1114, 1036, 927, 886,
873, 811. 1H NMR (CDCl3, 200 MHz): δ ppm 1.12 (t, J = 7.2
Hz, 3H), 2.98 (q, J = 7.2 Hz, 2H), 6.20 (s, 2H), 6.89 (s, 1H), 7.56
(s, 1H).
1-(4-Phenyl)-2-(6-nitro-1,3-benzodioxol-5-yl)-1,2-ethanedione
(52b). Yield 60%. Rf 0.33 (cyclohexane/ethyl acetate, 70:30, v/
v). IR (KBr) cm-1: 3064, 1786, 1691, 1672, 1592, 1573, 1521,
1445, 1341, 1256, 1204, 1181, 1043, 1029, 977, 845, 803, 788, 765,
736, 713, 689, 637. 1H NMR (CDCl3, 200 MHz): δ ppm
7.48-7.90 (m, 6H), 8.25 (m, 3H).
1-(4-Fluorophenyl)-2-(6-nitrophenyl)-1,2-ethanedione (53b).
Yield 30.7%. Rf 0.57 (cyclohexane/ethyl acetate, 70:30, v/v).
IR (KBr) cm-1: 3094, 2859, 1789, 1696, 1676, 1597, 1572, 1522,
1506, 1412, 1343, 1313, 1257, 1235, 1198, 1159, 1145, 872, 853,
782, 605. 1H NMR (CDCl3, 250 MHz): δ ppm 7.23 (t, J = 9.7
Hz, 1H), 7.78 (m, 2H), 7.8 (d, J = 8.0 Hz, 1H), 8.2 (dd, J = 10.0
and 2.0 Hz, 2H), 8.20 (dd, J = 6.0 and 2.0 Hz, 2H). 13C NMR
5-[(E/Z)-2-(4-Ethoxyphenyl)ethynyl-6-nitro-1,3-benzodioxol
(15a). Yield 82%. Rf 0.59 and 0.52 (cyclohexane/ethyl acetate,
70:30, v/v). IR (KBr) cm-1: 2971, 1642, 1605, 1511, 1482, 1325,
1
1259, 1175, 1120, 1033, 928, 873, 844, 815. H NMR (CDCl3,
200 MHz): (cis) δ 1.36 (t, J = 7.0 Hz, 3H), 3.96 (q, J = 7.0 Hz,
2H), 6.03 (s, 2H), 6.59 (d, J = 11.8 Hz, 1H), 6.61 (s, 1H), 6.69 (d,
J = 8.6 Hz, 2H), 6.70 (d, J = 11.8 Hz, 1H), 6.98 (d, J = 8.6 Hz,
2H), 7.60 (s, 1H); (trans) δ 1.40 (t, J = 7.0 Hz, 3H), 4.03 (q, J =
7.0 Hz, 2H), 6.08 (s, 2H), 6.86 (d, J = 8.6 Hz, 2H), 6.88 (d, J =
16.5 Hz, 1H), 7.08 (s, 1H), 7.42 (d, J = 8.7 Hz, 2H), 7.47 (s, 1H),
7.58 (d, J = 16.5 Hz, 1H).
5-[(E/Z)-2-(4-Ethyl)ethynyl]-6-nitro-1,3-benzodioxol (51a).
Yield 78%. Rf 0.55 (cyclohexane/ethyl acetate, 70:30, v/v). IR
(KBr) cm-1: 2957, 2885, 1607, 1519, 1503, 1477, 1327, 1254,
1036, 933, 876, 813. 1H NMR (CDCl3, 200 MHz): δ ppm 0.89 (t,
J = 7.5 Hz, 3H), 0.99 (t, J = 7.5 Hz, 3H), 1.96 (m, 2H), 2.15 (m,
2H), 5.60 (dt, J = 11.5 and 7.5 Hz, 1H, cis), 5.97 (s, 2H), 6.00 (s,
2H), 6.02 (dt, J = 15.6 and 6.5 Hz, 1H, trans), 6.49 (d, J = 11.4
Hz, 1H, cis), 6.57 (s, 1H), 6.74 (d, J = 15.2 Hz, 1H, trans), 679 (s,
1H), 7.26 (s, 1H), 7.39 (s, 1H).
5-[(E/Z)-2-(4-Phenyl)ethenyl]-6-nitro-phenyl (52a). Yield
96%. Rf 0.78 and 0.73 (cyclohexane/ethyl acetate, 70:30, v/v).
IR (KBr) cm-1: 3056, 3028, 1627, 1604, 1571, 1519, 1495, 1443,
1
1344, 1302, 1142, 1076, 962, 920, 854, 783, 755, 698. H NMR
(CDCl3, 200 MHz): δ ppm 6.75 (d, J = 12.0 Hz, 1H, cis), 6.89

706 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2
Nepveu et al.
(CDCl3, 75 MHz): δ ppm 115.8 (CH), 116.07 (CH), 124.1
(CH), 128.9 (C), 131.0 (CH), 132.4 (CH), 133.0 (C), 133.5
(CH), 133.6 (CH), 134.9 (CH), 164.9 (C), 168.3 (C), 186,4 (C),
189.8 (C).
1-(4-Butyl)-2-(6-nitro-1,3-benzodioxol-5-yl)-1,2-ethanedione
(54b). Rf 0.40 (cyclohexane/ethyl acetate, 70:30, v/v). IR (KBr)
cm-1: 3115, 3069, 2923, 2852, 1703, 1676, 1598, 1521, 1504,
1484, 1428, 1326, 1270, 1231, 1149, 1035, 929, 884, 869, 856, 837,
780. 1H NMR (CDCl3, 250 MHz): δ ppm 6.26 (s, 2H), 7.01 (s,
1H), 7.21 (d, J = 7.5 Hz, 2H), 7.61 (s, 1H), 8.2 (d, J = 7.5 Hz,
2H).
2-(4-Fluorophenyl)-3H-indol-3-one-N-oxide (53). Yield: 1%.
Rf: 0.64 (cyclohexane/ethyl acetate 70/30, v/v). UV (CH3CN)
λ
max: 290 nm. IR (KBr) cm-1: 3003, 2879, 1707, 1595, 1582,
1526, 1492, 1461, 1384, 1301, 1287. 1H NMR (CDCl3, 300
MHz): δ ppm 7.10-7.23 (m, 2H), 7.50-7.74 (m, 4H),
8.70-8.80 (m, 2H). 13C NMR (CDCl3, 75 MHz): δ ppm 186.9
(C), 165.3 (C), 161.9 (C), 147.8 (C), 134.9 (CH), 131.2 (CH),
130.3 (CH), 130.2 (CH), 122.7 (C), 122.3 (C), 121.7 (CH), 115.9
(CH), 115.7 (CH), 114.2 (CH). MS (EI): m/z 241 (Mþ•), 224, 195,
184, 121, 107, 95, 76, 63, 50, 39.
6-(4-Fluorophenyl)-7H-[1,3]dioxolo-[4,5-f]indol-7-one-5-oxide
(54). Yield 1%. Rf 0.56 (cyclohexane/ethyl acetate 70:30, v/v).
UV (CH3CN) λmax: 295 nm. IR (KBr) cm-1: 3100, 1703, 1604,
1588, 1535, 1502, 1477, 1390, 1371, 1316, 1231. 1H NMR
(CDCl3, 300 MHz): δ ppm 6.17 (s, 2H), 7.04 (s, 1H),
7.14-7.21 (m, 2H), 7.26 (s, 1H), 8.66-8.70 (m, 2H). MS (EI):
m/z 285 (Mþ•), 268, 212, 190, 171, 123, 95, 75, 62, 50, 39.
6.2. Synthesis by Method 2 (Compounds 2, 49-50, 56). 1-Iodo-
2-nitrobenzene (19.4 mmol) was dissolved in freshly distilled
triethylamine (100 mL) to which ethynyl derivative (19.4 mmol)
was added. The reaction was stirred at room temperature under
N2 for 30 min, at which point dichloro-bis-(triphenylphosphine)
palladium (0.4 mmol) and copper(I) iodide (1.4 mmol) were
added. The mixture was stirred at room temperature for 5-
7 days. The reaction mixture was filtered, the remaining solid
was washed with ethyl acetate, and the combined solutions were
evaporated to dryness, leaving an oily liquid. After solvent
evaporation the crude product was purified by column chroma-
tography (SiO2). The product was then purified by recrystalliza-
tion from toluene.
General Synthetic Procedure for Indolone N-Oxide Analogues
(i). To a solution of diketone (0.59 mmol) in THF (10 mL) was
added a 10% aqueous solution of NH4Cl (11 mL) and Zn (2.50
mmol). After 1 h of stirring at rt, the mixture was filtered, the
two liquid phases separated, and the aqueous phase was ex-
tracted with dichloromethane. The combined organic phase was
dried over anhydrous Na2SO4 and concentrated under vacuum.
The residue was dissolved in THF or dichloromethane (10 mL)
and heated under reflux until complete disappearance of the
intermediate hydroxylamine. After evaporation of the solvent,
the crude product was purified by column chromatography
(SiO2) with toluene.
6-(4-Chlorophenyl)-7H-[1,3]dioxolo[4,5-f]indol-7-one-5-oxide
(1). Yield 17.4%. Rf 0.61 (cyclohexane/ethyl acetate, 70:30, v/v);
mp: 210-211 ꢀC. UV (EtOH) λmax nm (ε L mol-1 cm-1): 298
3
3
(29 444). IR (KBr) cm-1: 3101, 2906, 1707, 1584, 1521, 1465,
1370, 1301. 1H NMR (CD3CN, 500 MHz): δ ppm 6.21 (s, 2H),
7.12 (s, 1H), 7.24 (1s, 1H), 7.55 (dd, J = 2 Hz, J = 9 Hz; 2H),
8.58 (dd, J = 2 Hz, J = 9 Hz; 2H). 13C NMR (CD3CN, 125
MHz): δ ppm 97.0 (CH), 101.8 (CH), 103.9 (CH2), 116.7 (C),
125.1 (C), 128.69 (CH ꢀ 2), 128.71 (CH ꢀ 2), 131.7 (C), 135.3
(C), 146.0 (C), 150.2 (C), 153.3 (C), 185.9 (C). MS (EI): m/z 301
(Mþ.), 284, 139, 120, 62. HR-MS [Mþ•] calcd for C15H8NO4Cl
301.0142, found 301.0141.
2-(4-Phenoxyphenyl)-3H-indol-3-one-N-oxide (2). Yield 34%.
Rf 0.50 (cyclohexane/ethyl acetate 85:15, v/v); mp: 136-137 ꢀC.
UV (CH3CN) λmax nm (ε L mol-1 cm-1): 287 (40798). IR
3
3
(KBr) cm-1: 3090, 1704, 1585, 1526, 1488, 1384, 1241. 1H
NMR (CDCl3, 300 MHz): δ ppm 8.70 (d, J = 9.3 Hz, 2H),
7.68 (d, J = 3.9 Hz, 2H), 7.63 (d, J = 7.0 Hz, 1H), 7.49-7.57 (m,
1H), 7.38 (t, J = 7.5 Hz, 2H), 7.18 (t, J = 7.5 Hz, 1H), 7.09 (dd,
J = 2.4 Hz, J = 9.3 Hz, 4H). NMR 13C (CDCl3, 75 MHz): δ
ppm 187.1 (C), 159.6 (C), 155.7 (C), 148.0 (C), 134.9 (CH), 131.3
(CH), 130.0 (C), 129.9 (2CH), 129.8 (2CH), 124.4 (CH), 122.8
(C), 121.6 (CH), 120.6 (C), 120.0 (2CH), 117.8 (2CH), 114.1
(CH). HR-MS [Mþ•] calcd for C20H13NO3 315.0895, found
315.0892.
6-(4-Ethoxyphenyl)-7H-[1,3]dioxolo[4,5-f]indol-7-one-5-oxide
(15). Yield 11.5%; mp 169-170 ꢀC. Rf 0.58 (cyclohexane/ethyl
acetate, 70:30, v/v). UV (EtOH) λmax nm (ε L mol-1 cm-1): 298
3
3
(38 790). IR (KBr) cm-1: 1708, 1684, 1593, 1529, 1499, 1475,
1
1387, 1366, 1311, 1291, 1266. H NMR (CDCl3, 300 MHz): δ
ppm 1.42 (t, J = 7.0 Hz, 3H), 4.09 (q, J = 7.0 Hz, 2H), 6.12 (s,
2H), 6.96 (d, J = 9.2 Hz, 2H), 6.98 (s, 1H), 7.13 (s, 1H), 8.63 (d,
J = 9.2 Hz, 2H). 13C NMR (CDCl3, 75 MHz): δ ppm 14.8
(CH3), 63.6 (CH2), 97.3 (CH), 102.3 (CH), 103.1 (CH2), 114.6
(CH ꢀ 2), 116.6 (C), 118.9 (C), 129.5 (CH ꢀ 2), 131.2 (C), 145.1
(C), 149.4 (C), 152.9 (C), 160.6 (C), 184.1 (C). MS (EI): m/z 311
(Mþ•), 283, 265, 190, 163, 149, 121, 119, 93, 77, 62. HR-MS
[Mþ•] calcd for C17H13NO5 311.0794, found 311.0793.
2-Isobutyl-3H-indol-3-one-N-oxide (49). Yield 1%. Rf 0.68
(cyclohexane/ethyl acetate 70:30, v/v); mp: 90 ꢀC. UV
(CH3CN) λmax nm (ε L•mol-1•cm-1): 245 (25700). IR (KBr)
cm-1: 2967, 1700, 1604, 1533, 1461, 1430, 1379, 1295, 1276. 1H
NMR (CDCl3, 300 MHz): δ ppm 7.49-7.63 (m, 4H), 2.56 (d,
J = 6.9 Hz, 2H), 2.17-2.26 (m, 1H), 0.98 (d, J = 6.6 Hz, 6H).
13C NMR (CDCl3, 75 MHz): δ ppm 187.1 (C), 147.4 (C), 138.9
(C), 134.3 (CH), 131.0 (CH), 123.1 (C), 121.4 (CH), 113.8 (CH),
30.2 (CH2), 26.8 (CH), 22.8 (2CH3). HR-MS [Mþ•] calcd for
C12H13NO2 203.0946, found 203.0946.
6-Ethyl-7H-[1,3]-dioxolo[4,5-f]indol-7-one-5-oxide (51). Yield
5.1%. Rf 0.60 (cyclohexane/ethyl acetate, 70:30, v/v); mp: 136
ꢀC. UV (EtOH) λmax nm (ε L mol-1 cm-1): 269 (30 035). IR
3
3
(KBr) cm-1: 3083, 2980, 1696, 1545, 1467, 1389, 1359, 1311,
1285. 1H NMR (CDCl3, 200 MHz): δ ppm 1.18 (t, J = 7.6 Hz,
3H), 2.61 (q, J = 7.6 Hz, 2H), 6.11 (s, 2H), 6.95 (s, 1H), 7.08 (s,
1H). 13C NMR (CDCl3, 100 MHz): d 9.9 (CH3), 15.2 (CH2),
97.5 (CH), 102.5 (CH), 103.2 (CH2), 117.0 (C), 139.1 (C), 144.4
(C), 149.6 (C), 152.5 (C), 185.9 (C). MS (EI): m/z 219 (Mþ•), 202
(Mþ• -17), 174, 120, 62. HR-MS [Mþ•] calcd for C11H9NO4
219.0532, found 219.0532.
2-Propyl-3H-indol-3-one-N-oxide (50). Yield 7%. Rf 0.52
(cyclohexane/ethyl acetate 70:30, v/v); mp: 58 ꢀC. UV
(CH3CN) λmax nm (ε L mol-1 cm-1): 271 (24658). IR (KBr)
3
3
cm-1: 2962, 2927, 2872, 1698, 1599, 1532, 1464, 1434, 1381,
1345, 1285. 1H NMR (CDCl3, 300 MHz): δ ppm 7.48-7.62 (m,
4H), 2.65 (t, J = 7.5 Hz, 2H), 1.70 (m, J = 7.5 Hz, 2H), 1.02 (t,
J = 7.5 Hz, 3H). 13C NMR (CDCl3, 75 MHz): δ ppm 187.0 (C),
147.4 (C), 139.3 (C), 134.3 (CH), 131.0 (CH), 123.1 (C), 121.3
(CH), 113.8 (CH), 23.2 (CH2), 19.0 (CH2), 14.1 (CH3).
MS-(-)APCI, m/z: 189 [M-•]. HR-MS [M þ H]þ calcd for
C11H12NO2 190.0868, found 190.0876.
2-Phenyl-3H-indol-3-one 1-Oxide (52). Yield 1%. Rf 0.75
(cyclohexane/ethyl acetate 70:30, v/v); mp: 186 ꢀC. UV
(EtOH) λmax nm (ε L mol-1 cm-1): 285 (47458). IR (KBr)
3
3
cm-1: 3063, 1706, 1598, 1524, 1481, 1389, 1314. 1H NMR
(CDCl3, 300 MHz): δ ppm 7.28-7.70 (m, 7H), 8.62-8.67 (m,
2H). 13C NMR (CDCl3, 75 MHz): δ ppm 186.9 (C), 147.8 (C),
134.8 (CH), 133.0 (C), 131.2 (CH), 130.7 (CH), 128.5 (2 ꢀ CH),
127.9 (2 ꢀ CH), 125.9 (C), 122.8 (C), 121.6 (CH), 114.2 (CH).
MS (EI): m/z 223 (Mþ•), 206, 177, 167, 151, 139, 130, 103, 90, 76,
63, 50, 39. HR-MS [Mþ•] calcd for C14H9NO2 223.0633, found
223.0635.
2-(4-Nitrophenyl)-3H-indol-3-one-N-oxide (56). Yield 18%.
Rf 0.52 (cyclohexane /ehtyl acetate 70:30, v/v); mp: 246 ꢀC.
UV (CH3CN) λmax nm (ε L mol-1 cm-1): 290 (21854). IR
3
3
(KBr) cm-1: 3132, 1715, 1594, 1512, 1482, 1388, 1344, 1314,
1288. 1H NMR (CDCl3, 500 MHz): δ ppm 8.94 (d, J = 8.7 Hz,
2H), 8.31 (d, J = 8.7 Hz, 2H), 7.75 (d, J = 4.5 Hz, 2H),

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2 707
1
7.61-7.66 (m, 2H). 13C NMR (CDCl3, 125 MHz): δ ppm 187.0
(C), 147.8 (C), 135.5 (C), 135.3 (CH), 132.0 (C), 132.1 (CH),
131.2 (C), 128.2 (2CH), 123.5 (C), 123.6 (2CH), 121.8 (CH),
114.6 (CH). MS-(-)APCI, m/z: 268 [M-•]. HR-MS [M þ H]þ
calcd for C14H9N2O4 269.0562, found 269.0581.
1590, 1489, 1388, 1332, 1220. H NMR (CDCl3, 500 MHz): δ
ppm 9.24 (s, 1H), 8.64 (dd, J = 1.5 Hz, J = 9.0 Hz, 2H), 7.86 (d,
J = 9.0 Hz, 1H), 7.79 (d, J = 9.0 Hz, 1H), 7.12-7.20 (m, 3H),
7.06 (s, 1H), 6.16 (s, 2H), 3.95 (s, 3H). 13C NMR (CDCl3, 125
MHz): δ ppm 186.3 (C), 159.5 (C), 153.0 (C), 143.2 (C), 135.6
(C), 131.0 (C), 130.9 (CH), 128.6 (C), 128.6 (CH), 127.9 (CH),
127.1 (CH), 123.6 (CH), 122.9 (C), 122.2 (C), 119.7 (CH), 115.8
(C), 106.2 (CH), 103.7 (CH2), 102.1 (CH), 55.2 (CH3).
MS-(þ)ESI, m/z: 348 [M þ H]þ, 370 [M þ Na]þ, 717 [2M þ
Na]þ. HR-MS [Mþ•] calcd for C20H13NO5 347.0794, found
347.0794.
6.3. Synthesis by Method 3 (Compounds 3-48, 55, 57-66). 1-
Iodo-2-nitroaryl (2 mmol) was dissolved in freshly distilled
triethylamine (8 mL) to which ethynyl derivative (2 mmol) was
added. The reaction was stirred at room temperature under
N2 for 10 min, and then dichloro-bis-(triphenylphosphine)-
palladium (0.1 mmol) and copper(I) iodide (0.2 mmol) were
added. The mixture was stirred at room temperature for 3 h. The
reaction mixture was filtered. After solvent evaporation, the
crude intermediate product was heated under reflux in
the presence of pyridine (8 mL) and 4-dimethylaminopyridine
(4-DMAP) (2 mmol) for a time-period varying between 2 h and
4 days. There are two other possibilities to carry out this
protocol: The first one consists of catalyzing the reaction with
AuBr3 (3% molar) in either toluene or dichloromethane as
solvent (compounds 22 and 26). The second one consists of
using the same reagents as the general protocol under micro-
waves conditions (Biotage Initiator 300W). These conditions are
mentioned for each compound (45-48). The crude product was
purified by column chromatography (SiO2) using either dichlor-
omethane/cyclohexane or petroleum ether/ethyl acetate or pet-
roleum ether/dichloromethane. After concentration, the
purified product was stirred with boiling ethanol and filtered
at room temperature. When possible, the final compound was
recrystallized from toluene.
6-(4-Phenoxylphenyl)-7H-[1,3]-dioxolo[4,5-f]- indol-7-one-5-
oxide (7). Yield 33.5%. Rf 0.46 (cyclohexane/ethyl acetateethyl
acetate, 70:30, v/v); mp: 168 ꢀC. UV (CH3CN) λmax nm (ε
L mol-1 cm-1): 295 (52392). IR (KBr) cm-1: 3098, 2914,
3
3
1706, 1589, 1523, 1491, 1475, 1388, 1362, 1310, 1262. 1H
NMR (CDCl3, 300 MHz): δ ppm 8.67 (d, J = 9.0 Hz, 2H),
7.41 (t, J = 7.8 Hz, 2H), 7.23 (m, 2H), 7.07-7.11 (m, 4H), 7.04
(s, 1H), 6.16 (s, 2H). 13C NMR (CDCl3, 75 MHz): δ ppm 186.1
(C), 159.4 (C), 155.8 (C), 152.9 (C), 149.5 (C), 144.9 (C), 132.5
(C), 130.0 (2CH), 129.5 (2CH), 124.3 (CH), 120.8 (2C), 120.0
(2CH), 117.9 (2CH), 116.6 (C), 103.2 (CH2), 102.3 (CH), 97.4
(CH). HR-MS [Mþ•] calcd for C21H13NO5 359.0794, found
359.0787.
2-(3,4-Dichloro-phenyl)-1-oxy-indol-3-one (8). Yield 78%;
mp: 204.2-204.8 ꢀC. UV (CH3CN) λmax: 286 nm. IR
(CH2Cl2) cm-1: 3040, 1705, 1590, 1510, 1420, 1384, 1265. H
1
NMR (CDCl3, 300 MHz) δ ppm 7.55-7.60 (m, 2H), 7.65-7.72
(m, 3H), 8.57 (dd, J = 1.8 Hz, 8.7 Hz, 1H), 8.89 (d, J = 2.1 Hz,
1H). 13C NMR (CDCl3, 125 MHz): δ (ppm) 186.4 (C), 147.7 (C),
135.1 (CH), 134.7 (C), 133 (C), 131.7 (CH), 130.6 (CH), 130.3
(C), 129 (CH), 126.7 (CH), 125.7 (C), 122.6 (C), 121.9 (CH),
114.4 (CH). MS-(-)APCI, m/z: 291 [M-•]. HR-MS [Mþ•] calcd
for C14H7NO2Cl2 290.9854, found 290.9850.
2-(4-Chloro-phenyl)-5-methoxy-1-oxy-indol-3-one (9). Yield
60.8%; mp: 190.5-191.1 ꢀC. UV (CH3CN) λmax: 290 nm. IR
(KBr) cm-1: 3104, 2951, 1702, 1639, 1598, 1519, 1481, 1433,
1384, 1283, 1248. 1H NMR (CDCl3, 300 MHz) δ ppm 3.91 (s,
3H), 7.08 (dd, J = 2.4 Hz, 8.4 Hz, 1H), 7.15 (d, J = 2.4 Hz, 1H),
7.46 (d, J = 8.7 Hz, 2H), 7.58 (d, J = 8.4 Hz, 1H), 8.59 (d, J =
8.7 Hz, 2H). 13C NMR (CDCl3, 125 MHz): δ (ppm) 186.6 (C),
162.6 (C), 140.7 (C), 136.2 (C), 131 (C), 128.8 (2 CH), 128.6 (2
CH), 124.6 (C), 124.5 (C), 118.4 (CH), 115.5 (CH), 108 (CH),
56.3 (CH3). MS-(þ)ESI, m/z: 288 [M þ H]þ, MS-(-)ESI, m/z:
287 [M-•]. HR-MS [M þ H]þ calcd for C15H11NO3Cl 288.0427,
found 288.0435.
2-(4-Hydroxymethylphenyl)-3H-indol-3-one-N-oxide (3). Yield
8%. Rf 0.64(CH2Cl2/EtOH 94:6); mp: 199-200 ꢀC. UV(CH3CN)
λ
max nm (ε L mol-1 cm-1): 290 (33899). IR (KBr) cm-1: 3268,
1
3
3
3083, 2904, 1701, 1597, 1526, 1493, 1458, 1385, 1284. H NMR
(CDCl3, 300 MHz): δ ppm 8.95 (d, J = 8.4 Hz, 2H), 7.75 (d, J =
8.4 Hz, 2H), 7.68 (d, J =4.8 Hz, 1H), 7.55 (t, J = 7.2Hz, 2H), 7.40
(t, J = 7.2 Hz, 1H), 7.08 (s, 1H), 4.98 (s, J = 5.4 Hz, 2H). 13
C
NMR (CDCl3, 75 MHz): δ ppm 187.0 (C), 147.9 (C), 146.4 (C),
145.3 (C), 134.6 (CH), 132.1 (C), 131.1 (CH), 127.9 (2CH), 126.6
(2CH), 125.0 (C), 121.3 (CH), 114.0 (CH), 63.6 (CH2). HR-MS
[M þ H]þ calcd for C15H12NO3 254.0817, found 254.0821.
2-(4-Methoxyphenyl)-3H-indol-3-one-N-oxide (4). Yield
21%. Rf 0.55 (cyclohexane/ethyl acetate 70:30, v/v); mp:
188-189 ꢀC. UV (CH3CN) λmax nm (ε L mol-1 cm-1): 294
3
3
(38939). IR (KBr) cm-1: 1702, 1599, 1530, 1492, 1460, 1380,
1301, 1262. 1H NMR (CDCl3, 300 MHz): δ ppm 8.74 (d, J = 9.3
Hz, 2H), 7.68 (dt, J = 4.5 Hz, J = 1.2 Hz, 2H), 7.61 (dt, J = 0.9
Hz, J = 7.2 Hz, 1H), 7.48-7.54 (m, 1H), 7.03 (d, J = 9.3 Hz,
2H), 3.89 (s, 3H). 13C NMR (CDCl3, 75 MHz): δ ppm 187.3 (C),
161.4 (C), 148.0 (C), 134.8 (CH), 131.9 (C), 130.7 (CH), 129.8
(2CH), 122.8 (C), 121.5 (CH), 118.8 (C), 114.1 (2CH), 113.9
(CH), 55.4 (CH3). HR-MS [M þ H]þ calcd for C11H12NO3
254.0817, found 254.0815.
5,6-Dimethoxy-1-oxy-2-p-tolyl-indol-3-one (10). Yield 1%.
IR (KBr) cm-1: 2998, 2839, 1698, 1651, 1594, 1525, 1485,
1220. 1H NMR (CDCl3, 300 MHz) δ (ppm) 2.41 (s, 3H),
3.97 (s, 3H), 4.04 (s, 3H), 7.13 (s, 1H), 7.26 (s, 1H), 7.29 (d,
J = 8.4 Hz, 2H), 8.10 (d, J = 8.4 Hz, 2H). MS-(þ)ESI, m/z: 298
[M þ H]þ.
2-(6-Methoxy-naphthalen-2-yl)-3H-indol-3-one-N-oxide (5).
Yield 28%. Rf 0.47 (cyclohexane/ethyl acetate 70:30, v/v); mp:
209 ꢀC. UV (CH3CN) λmax nm (ε L mol-1 cm-1): 272 (50521).
2-(4-Chloro-3-trifluoromethyl-phenyl)-5-methoxy-1-oxy-indol-
3-one (11). Yield 41.1%; mp: 194.1-194.9 ꢀC. IR (CH2Cl2)
cm-1: 3054, 2987, 1720, 1600, 1515, 1480, 1421, 1385, 1265. 1H
NMR (CDCl3, 300 MHz) δ (ppm) 3.92 (s, 3H), 7.10 (dd, J = 8.4
Hz, 2.1 Hz, 1H), 7.18 (d, J = 2.1 Hz, 1H), 7.58-7.62 (m, 2H),
8.77 (dd, J = 1.8 Hz, 8.4 Hz, 1H), 9.07 (s, 1H). MS-(þ)ESI, m/z:
356 [M þ H]þ, 378 [M þ Na]þ. HR-MS [Mþ•] calcd for
C16H9NO3ClF3 355.0223, found 355.0229.
3
3
IR (KBr) cm-1: 1700, 1600, 1517, 1480, 1456, 1393, 1335, 1286,
1261. 1H NMR (CDCl3, 300 MHz): δ ppm 9.30 (s, 1H), 8.70 (dd,
J = 1.8 Hz, J = 9.0 Hz, 1H), 7.86 (dd, J = 9.0 Hz, J = 11.4 Hz,
2H), 7.72 (m, 2H), 7.66 (d, J = 6.9 Hz, 1H), 7.52-7.58 (m, 1H),
7.18 (td, J = 2.4 Hz, J = 9.0 Hz, 2H), 3.97 (s, 3H). 13C NMR
(CDCl3, 75 MHz): δ ppm 187.4 (C), 159.4 (C), 148.1 (C), 135.6
(C), 134.9 (CH), 132.3 (C), 131.1 (CH), 130.9 (CH), 128.6 (CH),
128.4 (C), 126.9 (CH), 124.5 (CH), 122.9 (C), 121.6 (CH), 121.4
(C), 119.4 (CH), 114.0 (CH), 105.7 (CH), 55.4 (CH3).
2-(4-Chloro-phenyl)-6-methoxy-1-oxy-indol-3-one (12). Yield
5%. UV (CH3CN) λmax: 286 nm. IR (CH2Cl2) cm-1: 3054, 3000,
1705, 1590, 1523, 1470, 1430, 1383, 1265. 1H NMR (CDCl3, 300
MHz) δ (ppm) 3.96 (s, 3H), 6.92 (dd, J = 2.1 Hz, 8.1 Hz, 1H),
7.24 (d, J = 2.1 Hz, 1H), 7.45 (d, J = 9.3 Hz, 2H), 7.55 (d, J =
8.1 Hz, 1H), 8.66 (d, J = 9.3 Hz, 2H). MS-(þ)ESI, m/z: 288 [M þ
MS-(-)APCI, m/z: 303 [M-•]. HR-MS [Mþ•
C19H13NO3 303.0895, found 303.0881.
] calcd for
H]þ, 310 [M þ Na]þ, MS-(-)ESI, m/z: 287 [M-•]. HR-MS [Mþ•
]
6-(6-Methoxy-naphthalen-2-yl)-7H-[1,3]-dioxolo[4,5-f]-indol-
7-one-5-oxide (6). Yield 16.5%. Rf 0.40 (cyclohexane/ethyl
acetate, 70:30, v/v); mp: 248-249 ꢀC. UV (CH3CN) λmax nm
(ε L mol-1 cm-1): 306 (40572). IR (KBr) cm-1: 1704, 1619,
calcd for C15H10NO3Cl 287.0349, found 287.0349.
2-(3,4-Dichloro-phenyl)-5-methoxy-1-oxy-indol-3-one (13).
Yield 23.4%; mp: 193-194 ꢀC. IR (CH2Cl2) cm-1: 3054, 2980,
3
3

708 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2
Nepveu et al.
1705, 1591, 1500, 1455, 1415, 1384, 1265. 1H NMR (CDCl3, 300
MHz) δ (ppm) 3.91 (s, 3H), 7.10 (d, J = 8.7 Hz, 1H), 7.15 (s, 1H),
7.52 (d, J = 8.7 Hz, 1H), 7.57 (d, J = 8.7 Hz, 1H), 8.52 (dd, J =
1.2 Hz, 8.7 Hz, 1H), 8.83 (s, 1H). MS-(þ)ESI, m/z: 322 [M þ
H]þ, 344 [M þ Na]þ. HR-MS [Mþ•] calcd for C15H9NO3Cl2
320.9959, found 320.9969.
δ (ppm) 3.87 (s, 3H), 3.89 (s, 3H), 6.99-7.07 (m, 3H), 7.13 (d,
J = 2.7 Hz, 1H), 7.54 (d, J = 8.1 Hz, 1H), 8.66 (d, J = 9.0 Hz,
2H). MS-(þ)ESI, m/z: 284 [M þ H]þ, 589 [2M þ Na]þ. HR-MS
[Mþ•] calcd for C16H13NO4 283.0845, found 283.0849.
2-(2-4-Dichloro-phenyl)-5-methoxy-1-oxy-indol-3-one (23).
Yield 23.1%; mp: 180.5-180.9 ꢀC. IR (CH2Cl2) cm-1: 3054,
2980, 1709, 1614, 1524, 1473, 1434, 1397, 1358, 1265. 1H NMR
(CDCl3, 300 MHz) δ (ppm) 3.92 (s, 3H), 7.11 (dd, J = 8.4 Hz,
2.4 Hz, 1H), 7.20 (d, J = 2.4 Hz, 1H), 7.37-7.43 (m, 2H), 7.55
(d, J = 1.8 Hz, 1H), 7.62 (d, J = 8.4 Hz, 1H). MS-(þ)ESI, m/z:
322 [M þ H]þ. HR-MS [Mþ•] calcd for C15H9NO3Cl2 320.9959,
found 320.9966.
5-Methoxy-1-oxy-2(4-phenoxy-phenyl)-indol-3-one (24). Yield
27.5%; mp: 157.3-159.3 ꢀC. UV (CH3CN) λmax: 290 nm. IR
(CH2CL2) cm-1: 3060, 2980, 1700, 1587, 1520, 1485, 1420, 1384,
1265. 1H NMR (CDCl3, 300 MHz) δ (ppm) 3.90 (s, 3H),
7.07-7.20 (m, 7H), 7.39 (m, 2H), 7.56 (d, J = 8.4 Hz, 1H), 8.64
(d, J = 9.0 Hz, 2H). MS-(þ)ESI, m/z: 346 [M þ H]þ, 368 [M þ
Na]þ, 713 [2M þ Na]þ; MS-(-)APCI, m/z: 345 [M-•]. HR-MS
[Mþ•] calcd for C21H15NO4 345.1001, found 345.1012.
5-Methoxy-1-oxy-2-(4-trifluoromethoxy-phenyl)-indol-3-one
(25). Yield 6.1%; mp: 137.1-137.9 ꢀC. IR (CH2Cl2) cm-1: 3054,
2980, 1720, 1600, 1520, 1480, 1420, 1385, 1265. 1H NMR
(CDCl3, 300 MHz) δ (ppm) 3.91 (s, 3H), 7.09 (dd, J = 8.4 and
2.1 Hz, 1H), 7.17 (d, J = 2.1 Hz, 1H), 7.32 (d, J = 9.0 Hz, 2H),
7.59 (d, J = 8.4 Hz, 1H), 8.70 (d, J = 9.0 Hz, 2H). 13C NMR
(CDCl3, 125 MHz): δ (ppm) 186.5 (C), 162.7 (C), 150.0 (C),
140.6 (C), 130.8 (C), 129.2 (2 CH), 124.7 (C), 124.5 (C), 120.4
(CF3), 120.6 (2 CH), 118.5 (CH), 115.6 (CH), 108.1 (CH), 56.3
(CH3). MS-(þ)ESI, m/z: 338 [M þ H]þ. HR-MS [Mþ•] calcd for
C16H10NO4F3 337.0562, found 337.0564.
6-(3,4-Dichloro-phenyl)-5-oxy-[1,3]-dioxolo[4,5-f]-indol-7-one
(14). Yield 7%; mp: 270-271 ꢀC. UV (CH3CN) λmax: 286 nm.
IR (CH2Cl2) cm-1: 3060, 2980, 1701, 1590, 1520, 1460, 1385,
1355, 1265. 1H NMR (CDCl3, 300 MHz) δ (ppm) 6.28 (s, 2H),
7.31 (s, 1H), 7.40 (s, 1H), 7.86 (d, J = 8.7 Hz, 1H), 8.47 (dd, J =
1.5 Hz, 9.0 Hz, 1H), 8.71 (d, J = 1.5 Hz, 1H). MS-(þ)ESI, m/z:
336 [M þ H]þ, 358 [M þ Na]þ, MS-(-)ESI, m/z: 335 [M-•]. HR-
MS [Mþ•] calcd for C15H7NO4Cl2 334.9752, found 334.9749.
1-Oxy-2-p-tolyl-indol-3-one (16). Yield 61%; mp: 214.4-
215.3 ꢀC. UV (CH3CN) λmax: 289 nm. IR (CH2Cl2) cm-1
:
3050, 2980, 1700, 1590, 1410, 1384, 1275, 1250. 1H NMR
(CDCl3, 300 MHz) δ (ppm) 2.41 (s, 3H), 7.3 (d, J = 8.4 Hz,
2H), 7.48-7.56 (m, 1H), 7.61-7.69 (m, 3H), 8.56 (d, J = 8.4 Hz,
2H). 13C NMR (CDCl3, 125 MHz): δ (ppm) 187.2 (C), 148 (C),
141.4 (C), 134.8 (CH), 132.3 (C), 131 (CH), 129.3 (2 CH), 127.8
(2 CH), 123.2 (C), 122.9 (C), 121.6 (CH), 114.1 (CH), 21.8
(CH3). HR-MS [M þ H]þ calcd for C15H12NO2 238.0868, found
238.0881.
5-Methoxy-1-oxy-2-phenyl-indol-3-one (17). Yield 41%; mp:
135.3-135.7 ꢀC. UV (CH3CN) λmax: 286 nm. IR (CH2Cl2)
cm-1: 3050, 2980, 1710, 1595, 1480, 1420, 1384, 1265. 1H
NMR (CDCl3, 300 MHz) δ (ppm) 3.90 (s, 3H), 7.09 (d, J =
8.1 Hz, 1H), 7.16 (s, 1H), 7.42-7.60 (m, 4H), 8.58 (d, J = 6.9 Hz,
2H). MS-(-)ESI, m/z: 253 [M-•], MS-(þ)ESI, m/z: 254 [M þ
H]þ. HR-MS [Mþ•] calcd for C15H11NO3 253.0739, found
253.0736.
2-(4-Dimethylamino-phenyl)-5-methoxy-1-oxy-indol-3-one
(26d>) (Catalyzed with AuBr3 in Anhydrous Dichloromethane
12 h at Room Temperature). Yield 5.8%; mp: 214.8-218.9 ꢀC.
UV (CH3CN) λmax: 332 nm. IR (CH2Cl2) cm-1: 3054, 2980,
1690, 1602, 1540, 1480, 1420, 1383, 1265. 1H NMR (CDCl3,
300 MHz) δ (ppm) 3.06 (s, 6H), 3.88 (s, 3H), 6.76 (dm, J = 9.3
Hz, 2H), 7.03 (dd, J = 2.4 Hz, 8.4 Hz, 1H), 7.10 (d, J = 2.4
Hz, 1H), 7.49 (d, J = 8.4 Hz, 1H), 8.68 (dm, J = 9.3 Hz, 2H).
13C NMR (CDCl3, 125 MHz): δ (ppm) 187.8 (C), 161.5 (C),
151.2 (C), 141.4 (C), 132 (C), 129.1 (2 CH), 124.7 (C), 118.2
(CH), 114.6 (CH), 114.3 (C), 111.4 (2 CH), 107.8 (CH), 56.1
(CH3), 40 (2 CH3). MS-(þ)ESI, m/z: 297 [M þ H]þ, 615 [2M þ
Na]þ; MS-(-)APCI, m/z: 296 [M-•]. HR-MS [Mþ•] calcd for
5-Methoxy-1-oxy-2-p-tolyl-indol-3-one (18). Yield 13.4%;
mp: 196.8-196.9 ꢀC. IR (CH2Cl2) cm-1: 3060, 3000, 1705,
1
1590, 1480, 1430, 1384, 1276. H NMR (CDCl3, 300 MHz) δ
(ppm) 2.41 (s, 3H), 3.90 (s, 3H), 7.05 (dd, J = 2.4 Hz, 8.7 Hz,
1H), 7.14 (d, J = 2.4 Hz, 1H), 7.28 (d, J = 8.7 Hz, 2H), 7.57 (d,
J = 8.7 Hz, 1H), 8.50 (d, J = 8.4 Hz, 2H). MS-(þ)ESI, m/z: 268
[M þ H]þ, 290 [M þ Na]þ, 557 [2M þ Na]þ. HR-MS [Mþ•] calcd
for C16H13NO3 267.0895, found 267.0900.
2-(4-Chloro-phenyl)-1-oxy-6-trifluoromethyl-indol-3-one (19).
Yield 67.2%; mp: 167.3-167.5 ꢀC. UV (CH3CN) λmax: 292 nm.
IR (CH2Cl2) cm-1: 3054, 1720, 1591, 1421, 1384, 1310, 1265. 1H
NMR (CDCl3, 300 MHz) δ (ppm) 7.50 (d, J = 9.0 Hz, 2H), 7.78
(d, J = 8.1 Hz, 1H), 7.88 (d, J = 8.1 Hz, 1H), 7.96 (s, 1H), 8.66
(d, J = 9.0 Hz, 2H). 13C NMR (CDCl3, 75 MHz): δ (ppm) 185.4
(C), 147.9 (C), 137.5 (C), 132.4 (C), 129.1 (2 CH), 128.9 (2 CH),
128.8 (C), 125.4 (C), 123.9 (CH), 122.8 (CF3), 122.1 (CH), 117.9
C
17H16N2O3 296.1161, found 296.1159.
2-(4-Isopropoxy-phenyl)-5-methoxy-1-oxy-indol-3-one (27).
Yield 4.7%; mp: 148.9-150.8 ꢀC. UV (CH3CN) λmax nm (ε
L mol-1 cm-1): 302 (31250). IR (KBr) cm-1: 2969, 2933, 1711,
3
3
(C), 111.8 (CH). MS-(-)ESI, m/z: 325 [M-•]. HR-MS [Mþ•
calcd for C15H7NO2ClF3 325.0117, found 325.0116.
]
1597, 1527, 1430, 1380, 1357. H NMR (CDCl3, 300 MHz) δ
(ppm) 1.37 (d, J = 6.0 Hz, 6H), 3.89 (s, 3H), 4.65 (septuplet, J =
6 Hz, 1H), 6.98 (dm, J = 9.3 Hz, 2H), 7.05 (dd, J = 2.4 Hz, 8.7
Hz, 1H), 7.13 (d, J = 2.4 Hz, 1H), 7.54 (d, J = 8.7 Hz, 1H), 8.64
(dm, J = 9.3 Hz, 2H). 13C NMR (CDCl3, 125 MHz): δ (ppm)
187.2 (C), 162.1 (C), 159.6 (C), 139.4 (C), 130.9 (C), 129.4 (CH),
124.6 (C), 118.6 (C), 118.2 (2 CH), 115.5 (2 CH), 115.1 (CH),
107.9 (CH), 69.9 (CH), 56.2 (CH3), 22 (2 CH3). MS-(þ)ESI, m/z:
312 [M þ H]þ, 334 [M þ Na]þ, 645 [2M þ Na]þ. MS-(-)APCI,
m/z: 311 [M-•].
1
5-Methoxy-2-(-methoxy-naphtalen-2-yl)-1-oxy-indol-3-one (20).
Yield 24.7%; mp: 203.8-204.1 ꢀC. IR (CH2CL2) cm-1: 3054,
2980, 1705, 1596, 1480, 1420, 1265. 1H NMR (CDCl3, 300 MHz)
δ (ppm) 3.91 (s, 3H), 3.95 (s, 3H), 7.06-7.19 (m, 4H), 7.60 (d,
J = 8.4 Hz, 1H), 7.80 (d, J = 9.3 Hz, 1H), 7.87 (d, J = 8.7 Hz,
1H), 8.64 (dd, J = 2.1 Hz, 9 Hz, 1H), 9.22 (s, 1H). MS-(þ)ESI,
m/z: 334 [M þ H]þ, 689 [2M þ Na]þ. HR-MS [Mþ•] calcd for
C20H15NO4 333.1001, found 333.1005.
2-(40-Chloro-biphenyl-4-yl)-5-methoxy-1-oxy-indol-3-one (21>).
Yield 43%; mp: 227.2-227.4 ꢀC. IR (CH2Cl2) cm-1: 3054, 2987,
1720, 1620, 1550, 1480, 1421, 1385, 1265. 1H NMR (CDCl3, 300
MHz) δ (ppm) 3.91 (s, 3H), 7.09 (dd, J = 2.4 and 8.4 Hz, 1H), 7.16
(d, J = 2.4 Hz, 1H), 7.43 (d, J = 8.7 Hz, 2H), 7.57-7.60 (m, 3H),
7.69 (d, J= 8.1 Hz, 2H), 8.69 (d, J=8.1Hz, 2H).MS-(þ)ESI, m/z:
2-(4-Diethylamino-phenyl)-5-methoxy-1-oxy-indol-3-one (28).
Yield 3.8%; mp: 168.6-171.5 ꢀC. IR (CH2Cl2) cm-1: 3054,
2980, 1700, 1601, 1530, 1482, 1384, 1265. 1H NMR (CDCl3, 300
MHz) δ (ppm) 1.21 (t, J = 6.9 Hz, 6H), 3.43 (q, J = 6.9 Hz, 4H),
3.88 (s, 3H), 6.73 (d, J = 9.0 Hz, 2H), 7.03 (dd, J = 2.4 Hz, 8.4
Hz, 1H), 7.09 (d, J = 2.4 Hz, 1H), 7.49 (d, J = 8.4 Hz, 1H), 8.65
(d, J = 9.0 Hz, 2H). MS-(þ)ESI, m/z: 325 [M þ H]þ. HR-MS
[Mþ•] calcd for C19H20N2O3 324.1474, found 324.1483.
5-Methoxy-2(4-methoxy-3-methyl-phenyl)-1-oxy-indol-3-one
(29). Yield 5%; mp: 192.9-193.6 ꢀC. UV (CH3CN) λmax: 302
nm. IR (CH2Cl2) cm-1: 3054, 2980, 1700, 1597, 1470, 1421,
1380, 1265. 1H NMR (CDCl3, 300 MHz) δ (ppm) 2.28 (s, 3H),
364 [M þ H]þ, 386 [M þ Na]þ, 749 [2M þ Na]þ. HR-MS [Mþ•
]
calcd for C21H14NO3Cl 363.0662, found 363.0676.
2-(4-Methoxy-phenyl)-5-methoxy-1-oxy-indol-3-one (22) (Cata-
lyzed with AuBr3 in Ahydrous Toluene 5 h at 0.5 ꢀC). Yield 56%;
mp: 196.0-196.8 ꢀC. IR (CH2Cl2) cm-1: 3053, 2980, 1700, 1602,
1525, 1480, 1420, 1384, 1263. 1H NMR (CDCl3, 300 MHz)

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2 709
3.90 (s, 6H), 6.93 (d, J = 8.4 Hz, 1H), 7.06 (dd, J = 2.4 Hz, 6 Hz,
1H), 7.13 (d, J = 2.4 Hz, 1H), 7.54 (d, J = 8.4 Hz, 1H),8.48 (m,
1H), 8.55 (dd, J = 2.1 and 8.4 Hz, 1H). MS-(þ)ESI, m/z: 298
[M þ H]þ; MS-(-)APCI, m/z: 297 [M-•]. HR-MS [Mþ•] calcd for
C17H15NO4 297.1001, found 297.1011.
5-Methoxy-1-oxy-2-(4-trifluoromethylphenyl)-indol-3-one (37).
Yield 22.7%; mp: 192.9-193-8 ꢀC. UV (CH3CN) λmax: 284 nm.
IR (CH2Cl2) cm-1: 3040, 2980, 1715, 1590, 1490, 1425, 1385,
1324, 1265. 1H NMR (CDCl3, 300 MHz) δ (ppm) 3.92 (s, 3H),
7.11 (dd, J = 2.4 and 8.4 Hz, 1H), 7.18 (d, J = 2.4 Hz, 1H), 7.61
(d, J = 8.4 Hz, 1H), 7.73 (d, J = 8.4 Hz, 2H), 8.74 (d, J = 8.4 Hz,
2H). 13C NMR (CDCl3, 125 MHz): δ (ppm) 186.3 (C), 162.9 (C),
140.6 (C), 130.7 (C), 129.3 (C), 127.5 (4 CH), 125.4 (C), 124.9
(C), 123.8 (CF3), 122.7 (C), 118.6 (CH), 115.8 (CH), 108.1 (CH),
2-(4-Amino-phenyl)-5-methoxy-1-oxy-indol-3-one (30). Yield
3.3%; mp: 183.5-190.6 ꢀC. IR (CH2Cl2) cm-1: 3360, 3054,
2987, 1700, 1601, 1530, 1482, 1421, 1384, 1265. 1H NMR
(CDCl3, 300 MHz) δ (ppm) 3.88 (s, 3H), 4.07 (s, NH2), 6.74
(d, J = 9.0 Hz, 2H), 7.04 (dd, J = 2.4 and 8.4 Hz, 1H), 7.11 (d,
J = 2.4 Hz, 1H), 7.52 (d, J = 8.4 Hz, 1H), 8.58 (d, J = 9.0 Hz,
2H). MS-(þ)ESI, m/z: 269 [M þ H]þ, 559 [2M þ Na]þ. HR-MS
[Mþ•] calcd for C15H12N2O3 268.0848, found 268.0858.
6-Methoxy-2-(4-methoxy-phenyl)-1-oxy-indol-3-one (31). Yield
5.6%; mp: 179.3-182.0 ꢀC. IR (CH2Cl2) cm-1: 3054, 2986, 1741,
56.3 (CH3). MS-(þ)ESI, m/z: 322 [M þ H]þ. HR-MS [Mþ•
]
calcd for C16H10NO3F3 321.0613, found 321.0623.
2-(4-Acetyl-phenyl)-5-methoxy-1-oxy-indol-3-one (38). Yield
30.4%; mp: 205.3-206.1 ꢀC. IR (CH2Cl2) cm-1: 3054, 2980,
1680, 1596, 1480, 1415, 1384, 1265. 1H NMR (CDCl3, 300 MHz)
δ (ppm) 2.64 (s, 3H), 3.92 (s, 3H), 7.11 (dd, J = 8.7 and 2.4 Hz,
1H), 7.18 (d, J = 2.4 Hz, 1H), 7.61 (d, J = 8.7 Hz, 1H), 8.06 (d,
J = 8.4 Hz, 2H), 8.73 (d, J = 8.4 Hz, 2H). MS-(þ)ESI, m/z: 296
[M þ H]þ, 613 [2M þ Na]þ. HR-MS [Mþ•] calcd for C17H13NO4
295.0845, found 295.0848.
1
1600, 1545, 1421, 1380, 1265. H NMR (CDCl3, 300 MHz) δ
(ppm) 3.88 (s, 3H), 3.95 (s, 3H), 6.90 (d, J = 8.1 Hz, 1H), 7.02 (d,
J= 9.0 Hz, 2H), 7.23 (s, 1H), 7.53 (d, J= 8.1 Hz, 1H), 8.75 (d, J =
9.0 Hz, 2H). 13C NMR (CDCl3, 125 MHz): δ (ppm) 186.2 (C),
165.7 (C), 161.4 (C), 150.9 (C), 132.6 (C), 129.9 (2 CH), 123.4
(CH), 119.0 (C), 115.2 (C), 114.8 (CH), 114.1 (2 CH), 101.2 (CH),
56.3 (CH3), 55.4 (CH3). MS-(þ)ESI, m/z: 284 [M þ H]þ, 589
[2M þ Na]þ. HR-MS [M þ H]þ calcd for C16H14NO4 284.0923,
found 284.0900.
2-(4-Methoxy-phenyl)-1-oxy-6-trifluoromethyl-indol-3-one (32).
Yield 50%; mp: 178.8-180.0 ꢀC. IR (CH2Cl2) cm-1: 3054, 2987,
1740, 1610, 1421, 1390, 1265. 1H NMR (CDCl3, 300 MHz)
δ (ppm) 3.89 (s, 3H), 7.03 (dm, J = 9.3 Hz, 2H), 7.13 (d, J =
9.0 Hz, 1H), 7.80 (d, J = 9.0 Hz, 1H), 7.92 (s, 1H), 8.73 (dm, J =
9.3 Hz, 2H). 13C NMR (CDCl3, 125 MHz): δ (ppm) 186.0 (C),
161.9 (C), 148.2 (C), 136.5 (C), 132.9 (C), 129.9 (2 CH), 128.2 (C),
128.1 (C), 125.5 (C), 124.7 (CF3), 121.8 (CH), 118.3 (CH), 114.3 (2
CH), 111.4 (CH), 55.4 (CH3). MS-(þ)ESI, m/z: 322 [M þ H]þ.
HR-MS [M þ H]þ calcd for C16H11NO3F3 322.0691, found
322.0711.
2-(3-Chlorophenyl)-5-methoxy-1-oxy-indol-3-one (39). Yield
7%; mp: 156.1-156.9 ꢀC. 1H NMR (CDCl3, 300 MHz) δ
(ppm) 3.92 (s, 3H), 7.10 (dd, J = 8.4 and 2.4 Hz, 1H), 7.16 (d,
J = 2.4 Hz, 1H), 7.41-7.43 (m, 2H), 7.58 (d, J = 8.4 Hz, 1H),
8.52 (m, 1H), 8.66 (s, 1H). MS-(þ)ESI, m/z: 288 [M þ H]þ, 310
[M þ Na]þ.
Synthetic Procedure for 40-Aryl-4-yl-trimethylsilane (39d). A
mixture of trimethylsilylacetylene (5.03 mmol), triphenylpho-
sphine (0.08 mmol), tetrakis(triphenylphosphine)palladium
(0.042 mmol), and copper(I) iodide (0.13 mmol) in freshly
distilled triethylamine (60 mL) was stirred for 10 min under
N2. 3-Chloro-iodobenzene (4.19 mmol) added and the mixture
heated at 80 ꢀC for 4 h. Diethylether (60 mL) was added to the
cooled suspension. After filtration through a celite pad and
evaporation of the solvent, an orange oily residue was obtained
(965 mg). The crude product was purified by column chroma-
tography (SiO2) to produce a colorless oily liquid (883 mg).
Yield: 100%
6-Chloro-2-(4-methoxy-phenyl)-1-oxy-indol-3-one (33). Yield
8.8%; mp: 195-199 ꢀC. IR (CH2Cl2) cm-1: 3060, 2980, 1715,
Synthetic Procedure for 40-Ethynyl-aryl (39f). To a mixture of
40-aryl-4-yl-trimethysilane (4.23 mmol) in methanol (30 mL)
was added potassium carbonate (6.35 mmol). The mixture was
stirred at room temperature for a night and then diluted with
water (60 mL) and extracted with diethylether (120 mL). The
organic layer was dried over anhydrous MgSO4, filtered, and
evaporated to produce an orange oily liquid (443 mg). Yield:
77%
1
1600, 1540, 1460, 1384, 1265. H NMR (CDCl3, 300 MHz) δ
(ppm) 3.89 (s, 3H), 7.03 (d, J = 9.0 Hz, 2H), 7.48 (dd, J = 7.8
and 0.9 Hz, 1H), 7.54 (d, J = 7.8 Hz, 1H), 7.66 (d, J = 0.9 Hz,
1H), 8.73 (d, J = 9.0 Hz, 2H). MS-(þ)ESI, m/z: 288 [M þ H]þ,
310 [M þ Na]þ, 597 [2M þ Na]þ. HR-MS [Mþ•] calcd for
C15H10NO3Cl 287.0349, found 287.0352.
2-(4-Methoxy-phenyl)-6-methyl-1-oxy-indol-3-one (34). Yield
9.6%; mp: 177.6-178.4 ꢀC. IR (CH2Cl2) cm-1: 3040, 2980,
1699, 1600, 1527, 1499, 1435, 1378, 1299, 1264. 1H NMR
(CDCl3, 300 MHz) δ (ppm) 2.51 (s, 3H), 3.88 (s, 3H), 7.02
(dm, J = 9.0 Hz, 2H), 7.28 (d, J = 7.8 Hz, 1H), 7.47 (s, 1H), 7.48
(d, J = 7.8 Hz, 1H), 8.73 (dm, J = 9.0 Hz, 2H). MS-(þ)ESI, m/z:
2-(3-Methyl-4-tolyl)-5-methoxy-1-oxy-indol-3-one (40). Yield
34%; mp: 167.2-169.4 ꢀC. IR (CH2Cl2) cm-1: 3054, 2986, 1710,
1
1600, 1484, 1421, 1385, 1265. H NMR (CDCl3, 300 MHz) δ
(ppm) 2.32 (s, 3H), 2.34 (s, 3H), 3.91 (s, 3H), 7.06 (dd, J = 8.1
and 2.4 Hz, 1H), 7.15 (d, J = 2.4 Hz, 1H), 7.25 (d, J = 8.1 Hz,
1H), 7.56 (d, J = 8.1 Hz, 1H), 8.36 (d, J = 7.8 Hz, 1H), 8.39 (s,
1H). MS-(þ)ESI, m/z: 282 [M þ H]þ, 304 [M þ Na]þ, 585 [2M þ
Na]þ. HR-MS [Mþ•] calcd for C17H15NO3 281.1052, found
281.1053.
268 [M þ H]þ, 290 [M þ Na]þ, 557 [2M þ Na]þ. HR-MS [Mþ•
]
calcd for C16H13NO3 267.0895, found 267.0894.
2-(4-Methoxy-phenyl)-1-oxy-6-trifluoromethoxy-indol-3-one
(35). Yield 42%; mp: 173.6-174.4 ꢀC. UV (CH3CN) λmax: 287
nm. IR (CH2Cl2) cm-1: 3054, 2900, 1719, 1600, 1530, 1500,
1
Synthetic Procedure for 40-Aryl-4-yl-trimethysilane (40d). A
mixture of 4-iodo-o-xylene (2.15 mmol), dichloro-bis-(triphenyl-
phosphine)palladium (0.041 mmol), copper(I) iodide (0.0645
mmol) in freshly distilled triethylamine (5 mL) was stirred for
10 min under N2. Trimethylsilylacetylene (3.23 mmol) added and
the mixture heated at 80 ꢀC for 2 h. After cooling, the solvent was
evaporated and the crude product purified by column chroma-
tography to produce a yellow oily liquid (386 mg). Yield: 88%
Synthetic Procedure for 40-Ethynyl-aryl (40f). To a mixture of
40-aryl-4-yl-trimethylsilane (1.87 mmol) in methanol (5 mL) was
added potassium carbonate (1.87 mmol). The mixture was
stirred at room temperature for a night, diluted with water
(15 mL), and extracted twice with diethylether (20 mL). The
combined organic layers were dried over anhydrous MgSO4,
filtered, and evaporated to produce an orange oily liquid (183
mg). Yield: 75%
1470, 1421, 1384, 1265. H NMR (CDCl3, 300 MHz) δ (ppm)
3.89 (s, 3H), 7.03 (dm, J = 9.0 Hz, 2H), 7.31 (d, J = 8.1 Hz, 1H),
7.54 (s, 1H), 7.65 (d, J = 8.1 Hz, 1H), 8.73 (dm, J = 9.0 Hz, 2H).
13C NMR (CDCl3, 125 MHz): δ (ppm) 185.7 (C), 161.8 (C),
153.9 (C), 149.8 (C), 132.9 (C), 129.9 (2 CH), 123.0 (CH), 122.1
(CH), 122.0 (C), 121.5 (CF3), 118.5 (C), 114.2 (2 CH), 107.4
(CH), 55.4 (CH3). MS-(þ)ESI, m/z: 338 [M þ H]þ. HR-MS
[Mþ•] calcd for C16H10NO4F3 337.0562, found 337.0556.
2-(4-Methoxy-phenyl)-3-oxo-1-oxy-3H-indol-6-methyl carboxy-
late (36). Yield 7.2%; mp: 199.5-200.1 ꢀC. IR (CH2Cl2) cm-1
:
3054, 2987, 1716, 1601, 1500, 1421, 1384, 1265. 1H NMR
(CDCl3, 300 MHz) δ (ppm) 3.89 (s, 3H), 3.99 (s, 3H), 7.03
(dm, J = 9.3 Hz, 2H), 7.67 (d, J = 7.5 Hz, 1H), 8.23 (dd, J = 1.2
Hz, 7.5 Hz, 1H), 8.27 (d, J = 1.2 Hz, 1H), 8.72 (dm, J = 9.3 Hz,
1H). MS-(þ)ESI, m/z: 312 [M þ H]þ, 334 [M þ Na]þ. HR-MS
[Mþ•] calcd for C17H13NO5 311.0794, found 311.0798.

710 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2
Nepveu et al.
6.4. Synthesis by Method 3 (Compound 41). A mixture of
1-iodo-2-nitroaryl (7.12 mmol), dichloro-bis-(triphenylphos-
phine) palladium (0.14 mmol), and copper(I) iodide (0.36 mmol)
in freshly distilled triethylamine (110 mL) was stirred for 10 min
at 40 ꢀC under N2. Ethynyl derivative (8.54 mmol) was added,
and the reaction mixture stirred at 40 ꢀC for 6 h. After solvent
evaporation, a brown solid was obtained (4.07 g). The crude
product was purified by column chromatography (SiO2) using
petroleum ether/ethyl acetate (80:20 then 40/60) to produce an
orange solid (1.1 g), corresponding to the uncyclized product.
Yield: 58% (compound 41f is commercially available).
Acetylation of the uncyclized product: the uncyclized product
(0.75 mmol) was dissolved in dichloromethane (6 mL). DIPEA
(1.16 mmol) was added and the mixture cooled to 0 ꢀC. A
solution of acetylchloride (0.83 mmol) in anhydrous dichloro-
methane (1 mL) added dropwise. The reaction mixture was
stirred at room temperature for a night, washed with saturated
NaHCO3, brine, dried over MgSO4, filtered, and evaporated to
produce a yellow solid (324 mg). Purification by column chro-
matography (SiO2) using petroleum ether/ethyl acetate (90:10
then 50:50) produced a yellow solid (196 mg). Yield: 76%.
Cyclization of the acetylated product: the uncyclized product
(0.63 mmol) was dissolved, under N2, in anhydrous pyridine
(5 mL). The reaction mixture was refluxed for 12 h, and solvent
was removed. The crude product was purified by column
chromatography (SiO2) using petroleum ether/ethyl acetate
(80:20 then 40:60) to give a solid (66 mg). Further purification
was needed by suspending under reflux for 15 min this solid in
ethanol. After cooling and filtering the suspension, a mauve
solid (14 mg) was produced.
2-(4-Chloro-3-tolyl)-5-methoxy-1-oxy-indol-3-one (44). Yield
19%; mp: 174.4-175.0 UV (CH3CN) λmax: 291 nm. IR
(CH2Cl2) cm-1: 3060, 3000, 1710, 1600, 1515, 1480, 1430,
1384, 1265. 1H NMR (CDCl3, 300 MHz) δ (ppm) 2.45 (s, 3H),
3.91 (s, 3H), 7.10 (dd, J = 8.4 and 2.1 Hz, 1H), 7.16 (d, J = 2.1
Hz, 1H), 7.46 (d, J = 8.1 Hz, 1H), 7.59 (d, J = 8.1 Hz, 1H), 8.42
(dd, J = 8.4 Hz and 2.1, 1H), 8.53 (s, 1H). MS-(þ)ESI, m/z: 302
[M þ H]þ, 324 [M þ Na]þ. MS-(-)APCI, m/z: 301 [M-•]. HR-
MS [Mþ•] calcd for C16H12NO3Cl 301.0506, found 301.0504.
2-(4-Carboxymethylphenyl)-5-methoxy-1-oxy-indol-3-one
(45d>) (Reaction Carried out under Microwave Conditions:
10 mn at 140 ꢀC). Yield 2%; mp: 185.3-184.9 ꢀC. IR
(CH2Cl2) cm-1: 3054, 3000, 1720, 1596, 1580, 1420, 1384,
1265. 1H NMR (CDCl3, 300 MHz) δ (ppm) 3.92 (s, 3H), 3.95
(s, 3H), 7.10 (dd, J = 8.7 and 2.1 Hz, 1H), 7.17 (d, J = 2.1 Hz,
1H), 7.59 (d, J = 8.7 Hz, 2H), 8.12 (d, J = 8.7 Hz, 2H), 8.39
(d, J = 8.4 Hz, 1H), 8.69 (d, J = 8.4 Hz, 2H). MS-(þ)ESI, m/
z: 312 [M þ H]þ, 334 [M þ Na]þ, 645 [2M þ Na]þ. HR-MS
[Mþ•] calcd for C17H13NO5 311.0794, found 311.0794.
Synthetic Procedure for 40-Aryl-4-yl-trimethylsilane (45d). A
mixture of 4-iodo-methylbenzoate (6.25 mmol), copper(I) iodide
(0.185 mmol), and dichloro-bis-(triphenylphosphine) palladium
(0.125 mmol) in freshly distilled triethylamine (15 mL) was stirred
under N2. Trimethylsilylacetylene (9.58 mmol) was added and the
mixture heated at 80 ꢀC for 4 h. The suspension was cooled and
evaporated. The crude product was purified by column chroma-
tography (SiO2) using petroleum ether/ethyl acetate (100:00 then
90:10) to produce a yellow solid (1.05 g). Yield: 72%
Synthetic Procedure for 40-Ethynyl-aryl (45f). To a mixture of
40-aryl-4-yl-trimethylsilane (4.52 mmol) in methanol (10 mL)
was added potassium carbonate (4.52 mmol). The mixture was
stirred at room temperature for 24 h and then diluted with water
and diethylether. The organic layer dried over MgSO4, filtered,
and evaporated to produce a yellow solid (560 mg). Yield: 77%.
Synthesis of 4-Iodomethylbenzoate (Intermediate for 45d).
4-Iodobenzoic acid (8.06 mmol) was dissolved in anhydrous
methanol (20 mL). Thionyl chloride (24.2 mmol) was added and
the mixture refluxed for 6 h. Solvent was removed and the crude
product treated with ethyl acetate and NaOH 1N. The organic
layer was washed with a saturated solution of NaCl, dried
over MgSO4, filtered, and evaporated to produce a beige solid
(139 mg). Yield: 78%.
2-(4-Acetamidophenyl)-5-methoxy-1-oxy-indol-3-one (41). Yield
8%; mp: 249.1-254.1 ꢀC. IR (CH2Cl2) cm-1: 3320, 3054, 2980,
1690, 1585, 1485, 1420, 1384, 1265. 1H NMR (CDCl3, 300 MHz) δ
(ppm) 2.22 (s, 3H), 3.91 (s, 3H), 7.08 (dd, J = 8.4 and 2.1 Hz, 1H),
7.14 (d, J = 2.1 Hz, 1H), 7.56-7.66 (m, 3H), 8.66 (d, J = 9 Hz,
2H). MS-(þ)ESI, m/z: 333 [M þ Na]þ. HR-MS [Mþ•] Calcd for
C17H14N2O4 310.0954, found 310.0952.
2-(4-tert-Butylphenyl)-5-methoxy-1-oxy-indol-3-one (42). Yield
6%; mp: 141.7-143.6 ꢀC. IR (CH2Cl2) cm-1: 3054, 2980, 1690,
1
1600, 1483, 1415, 1384, 1265. H NMR (CDCl3, 300 MHz) δ
(ppm) 1.35 (s, 9H), 3.91 (s, 3H), 7.07 (dd, J = 8.4 and 2.4 Hz, 1H),
7.16 (d, J = 2.4 Hz, 1H), 7.51-7.59 (m, 3H), 8.52 (d, J = 8.7 Hz,
2H). MS-(þ)ESI, m/z: 310 [M þ H]þ (compound 42f is commer-
cially available). HR-MS [Mþ•] calcd for C19H19NO3 309.1365,
found 309.1359.
2-(3-Methyl-4-methoxyphenyl)-5-methoxy-1-oxy-indol-3-one
(46) (Reaction Carried out under Microwave Conditions: 10 mn at
150 ꢀC). Yield 6.3%; mp: 170.8 - 171.3 ꢀC. IR (CH2Cl2) cm-1
:
2-(2-Chlorophenyl)-5-methoxy-1-oxy-indol-3-one (43). Yield
28%; mp: 154.5-155.8 ꢀC. UV (CH3CN) λmax: 268 nm. IR
(CH2Cl2) cm-1: 3060, 2940, 1710, 1600, 1535, 1475, 1420, 1384,
1263. 1H NMR (CDCl3, 300 MHz) δ (ppm), 3.92 (s, 3H), 7.11
(dd, J = 8.4 and 2.4 Hz, 1H), 7.21 (d, J = 2.4 Hz, 1H), 7.37 -
7.48 (m, 3H), 7.55 (dd, J = 8.4 Hz and 2.1 Hz, 1H), 7.64 (d, J =
8.1 Hz, 1H). MS-(þ) ESI, m/z: 288 [M þ H]þ, 310 [M þ Na]þ,
3040, 2980, 1700, 1603, 1480, 1415, 1384, 1265. 1H NMR
(CDCl3, 300 MHz) δ (ppm) 2.31 (s, 3H), 3.84 (s, 3H), 3.91 (s,
3H), 6.87 (m, 2H), 7.07 (d, J = 9 Hz, 1H), 7.18 (d, J = 2.1 Hz,
1H), 7.33 (d, J = 9.6 Hz, 1H), 7.58 (d, J = 8.1 Hz, 1H).
MS-(þ)ESI, m/z: 298 [M þ H]þ, 617 [2M þ Na]þ (compound
46f is commercially available). HR-MS [Mþ•] calcd for
C17H15NO4 297.1001, found 297.1012.
597 [2M þ Na]þ. MS-(-)APCI, m/z: 287 [M-•]. HR-MS [Mþ•
]
2-(2-Thiophen-3-yl)-5-methoxy-1-oxy-indol-3-one (47) (Reac-
tion Carried out under Microwave Conditions: 10 mn at 140 ꢀC).
Yield 11%; mp: 171.9-172.6 ꢀC. IR (CH2Cl2) cm-1: 3060, 2980,
1710, 1600, 1480, 1420, 1384, 1265. 1H NMR (CDCl3, 300 MHz)
δ (ppm) 3.90 (s, 3H), 7.05 (dd, J = 8.1 and 3 Hz, 1H), 7.15 (d,
J = 2.7 Hz, 1H), 7.43 (m, 1H), 7.55 (d, J = 8.1 Hz, 1H), 8.24 (d,
J = 4.8 Hz, 1H), 8.94 (d, J = 3 Hz, 1H). MS-(þ)ESI, m/z: 260
[M þ H]þ, 282 [M þ Na]þ (compound 47f is commercially
available). HR-MS [Mþ•] calcd for C13H9NO3S 259.0303, found
259.0298.
calcd for C15H10NO3Cl 287.0349, found 287.0361.
Synthetic Procedure for 40-Aryl-4-yl-trimethylsilane (43d). A
mixture of 2-chloro-iodobenzene (6.3 mmol), tetrakis-
(triphenylphosphine)palladium (0.126 mmol), and copper(I)
iodide (0.19 mmol) in freshly distilled triethylamine (35 mL)
was stirred for 10 min under N2. Trimethylsilylacetylene (9.45
mmol) was added and the suspension heated at 80 ꢀC for 4 h.
After removal of the solvent and purification by column chro-
matography (SiO2) using petroleum ether 100%, a yellow oily
liquid (1.31 g) was produced. Yield: 100%
2-(4-Methoxyphenyl)-6-nitro-1-oxy-indol-3-one (48) (Reac-
tion Carried out under Microwave Conditions: 30 mn at 140 ꢀC).
Yield 48%; mp: 266.9-270.3 ꢀC. IR (CH2Cl2) cm-1: 3050, 2980,
1715, 1596, 1520, 1410, 1383, 1265. 1H NMR ((CD3)2SO,
300 MHz) δ (ppm) 3.87 (s, 3H), 7.18 (d, J = 8.1 Hz, 2H), 7.92 (d,
J = 6 Hz, 1H), 8.21 (d, J = 1.8 Hz, 1H), 8.46 (m, 1H), 8.60 (d,
Synthetic Procedure for 40-Ethynyl-aryl (43f). To a mixture of
40-aryl-4-yl-trimethylsilane (7.7 mmol) in methanol (10 mL) was
added potassium carbonate (11.55 mmol). The mixture was
stirred at room temperature for 24 h and then diluted with
water and extracted with diethylether. The organic layer was
dried over anhydrous MgSO4, filtered, and evaporated to
produce a yellow oily liquid (710 mg). Yield: 70%
J = 7.5 Hz, 2H). MS-(þ)ESI, m/z: 299 [M þ H]þ. HR-MS [Mþ•
]
calcd for C15H10N2O5 298.0590, found 298.0591.

Article
Synthetic Procedure for 40-Aryl-4-yl-trimethylsilane (48d). A
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2 711
2-(4-Ethyl-phenyl)-1-oxy-indol-3-one (60). Yield 31%; mp:
178.4-178.6 ꢀC. UV (CH3CN) λmax nm (ε L mol-1 cm-1):
mixture of 4-bromo-anisole (29.9 mmol), tetrakis(triphenyl-
phosphine)palladium (0.595 mmol), and copper(I) iodide
(1.497 mmol) in freshly distilled triethylamine (15 mL) was
stirred for 10 min under N2. Trimethylsilylacetylene (44.9 mmol)
was added dropwise. The suspension was heated to 80 ꢀC for 3
days. Solvent was removed. The crude product was purified by
column chromatography (SiO2) using petroleum ether 100% as
an elutent to produce a yellow solid (2.61 g). Yield: 46%
Synthetic Procedure for 4-Ethynyl-aryl (48f). To a mixture of
40-aryl-4-yl-trimethylsilane (11.94 mmol) in methanol (105 mL)
was added potassium carbonate (17.91 mmol). The suspension
stirred at room temperature for 18 h and then diluted with water
(215 mL) and extracted with diethylether (400 mL). The organic
layer was dried over MgSO4, filtered, and evaporated to produce
a yellow oily liquid (344 mg). Yield: 22%
3
3
294 (20845). IR (CH2Cl2) cm-1: 3060, 2980, 1700, 1595, 1535,
1
1490, 1420, 1384, 1265. H NMR (CDCl3, 300 MHz) δ (ppm)
1.27 (t, J = 7.5 Hz, 3H), 2.70 (q, J = 7.5 Hz, 2H), 7.35 (d, J =
8.4 Hz, 2H), 7.50-7.58 (m, 1H), 7.62-7.70 (m, 3H), 8.58 (d, J =
8.4 Hz, 2H). 13C NMR (CDCl3, 125 MHz): δ (ppm) 187.2 (C),
147.9 (C), 147.6 (C), 134.8 (CH), 132 (C), 130.9 (CH), 128.2
(2 CH), 127.9 (2 CH), 123.3 (C), 122.9 (C), 121.6 (CH), 114.1
(CH), 29.1 (CH2), 15.2 (CH3). MS-(þ)ESI, m/z: 252 [M þ H]þ.
HR-MS [M þ H]þ calcd for C16H14NO2 252.1025, found
252.1034.
1-Oxy-2-(4-trifluoromethyl-phenyl)indol-3-one (61). Yield
18%; mp: 210.0-210.3 ꢀC. UV (CH3CN) λmax nm (ε L mol-1
3
3
cm-1): 291 (11516). IR (CH2Cl2) cm-1: 3054, 2980, 1705, 1590,
1
1540, 1495, 1420, 1384, 1265. H NMR (CDCl3, 300 MHz) δ
(ppm) 7.56-7.77 (m, 6H), 8.78 (d, J = 7.8 Hz, 2H). MS-(þ)ESI,
m/z: 292 [M þ H]þ. HR-MS [Mþ•] calcd for C15H8NO2F3
291.0507, found 291.0505.
2-(3-Thiophene)-6-chloro-3H-indol-3-one-N-oxide (55). Yield
3%. Rf 0.61 (cyclohexane/ethyl acetate, 80/20, v/v); mp: 174 ꢀC.
UV (DMF) λmax nm (ε L mol-1 cm-1): 293 (nd). IR (KBr)
3
3
cm-1: 1715, 1600, 1541, 1499, 1458, 1425, 1363, 1346, 1204. 1H
NMR (CDCl3, 300 MHz): δ (ppm) 9.15 (s, 1H), 8.26 (d, J = 6.0
Hz, 1H), 7.67 (s, 1H), 7.47-7.51 (m, 2H), 7.52 (d, J = 3.0 Hz,
1H). 13C NMR (CDCl3, 75 MHz): δ (ppm) 184.9 (C), 148.9 (C),
141.3 (2C), 130.8 (CH), 130.0 (CH), 126.0 (CH), 125.9 (CH),
122.7 (CH), 121.1 (2C), 115.1 (CH).
5,6-Dimethoxy-1-oxy-2-phenyl-indol-3-one (62). Yield 7%.
IR (KBr) cm-1: 2939, 2835, 1740, 1700, 1647, 1593, 1568,
1
1519, 1435, 1330, 1300. H NMR (CDCl3, 300 MHz) δ (ppm)
3.97 (s, 3H), 4.04 (s, 3H), 7.14 (s, 1H), 7.27 (s, 1H), 7.42-7.54 (m,
3H), 8.60-8.64 (m, 2H). MS-(þ)ESI, m/z: 284 [M þ H]þ, 306 [M
þ Na]þ. HR-MS [M þ H]þ calcd for C16H14NO4 284.0923,
found 284.0930.
5-Oxy-6-phenyl-[1,3]dioxolo-[4,5-f]indol-7-one (63). Yield
28%; mp: 175.5-176.9 ꢀC. UV (CH3CN) λmax: 268 nm. IR
(CH2Cl2) cm-1: 2980, 2900, 1700, 1590, 1520, 1490, 1460, 1384,
1275. 1H NMR (CDCl3, 300 MHz) δ (ppm) 6.15 (m, 2H), 7.04 (s,
1H), 7.19 (s, 1H), 7.45-7.49 (m, 3H), 8.59 (d, J = 6.9 Hz, 2H).
MS-(þ)ESI, m/z: 268 [M þ H]þþ, 290 [M þ Na]þ; MS-(-)ESI, m/
z: 267 [M- 3 ]. HR-MS [M þ H] calcd for C15H10NO4 268.0610,
found 268.0621.
2-(4-Chlorophenyl)-1-oxy-indol-3-one (64). Yield 20%; mp:
177.0-177.1 ꢀC. UV (CH3CN) λmax: 280 nm. 1H NMR (CDCl3,
300 MHz) δ (ppm) 7.46 (d, J = 12.0 Hz, 2H), 7.54-7.61 (m, 1H),
7.64-7.73 (m, 3H), 8.66 (d, J = 12.0 Hz, 2H). 13C NMR
(CDCl3, 125 MHz): δ (ppm) 186.7 (C), 147.8 (C), 136.7 (C),
135 (CH), 131.9 (C), 131.4 (CH), 128.98 (2 CH), 128.91 (2 CH),
124.4 (C), 122.7 (C), 121.8 (CH), 114.3 (CH). MS-(þ)ESI, m/z:
258 [M þ H]þ; MS-(-)APCI, m/z: 257 [M-•]. HR-MS [M þ H]þ
calcd for C14H9NO2Cl 258.0322, found 258.0329.
Intermediate 4-Iodo-3-nitrobenzyloxybenzene (57e-58e). To a
mixture of 4-iodo-3-nitro-phenol (3 mmol), benzyle chloride (6
mmol), and ethanol (5 mL) was added pulverized anhydrous
K2CO3 (2.3 mmol). The reaction was stirred at 60 ꢀC during 15
h. The reaction mixture was filtered and the precipitate was
washed with ethyl acetate (100 mL). The filtrate was washed
with the brine solution (2 ꢀ 100 mL) and then dried on Na2SO4.
The crude product obtained after evaporation was purified by
recrystallization from cyclohexane. Yield 67%. Rf 0.64
(cyclohexane/ethyl acetate 85:15, v/v).
2-(6-Phenoxyphenyl)-6-benzyloxy-3H-indol-3-one-N-oxide
(57old>). Yield 4%. Rf 0.46 (cyclohexane/ethyl acetate,
85:15, v/v); mp: 148 ꢀC. UV (DMF) λmax nm (ε L mol-1
3
3
cm-1): 288 (44536). IR (KBr) cm-1: 1707, 1643, 1586, 1527,
1493, 1378, 1284, 1247. 1H NMR (CDCl3, 300 MHz): δ (ppm)
10.13 (d, J = 9.0 Hz, 2H), δ (ppm) 9.85 (s, 1H), 8.66-8.72 (m,
4H), 8.47-8.59 (m, 4H), 8.28-8.35 (m, 5H), 8.21 (dd, J = 2.1
and 8.1 Hz, 1H), 6.05 (s, 2H). 13C NMR (CDCl3, 75 MHz): δ
(ppm) 187.0 (C), 165.8 (C), 160.6 (C), 157.2 (C), 151.9 (C),
137.4 (C), 133.3 (C), 131.5 (2CH), 131.2 (2CH), 130.0 (2CH),
129.6 (CH), 129.1 (2CH), 125.7 (CH), 124.7 (CH), 122.7 (C),
2-(4-Chlorophenyl)-1,5-dioxypyrrolo(3,2-c)pyridine (65). Yield
29.3%; mp: 242-243 ꢀC. 1H NMR ((CD3)2SO, 300 MHz) δ
(ppm) 7.67 (d, J = 8.7 Hz, 2H), 7.76 (d, J = 6.3 Hz, 1H), 8.5 (d,
J = 8.7 Hz, 3H), 8.63 (s, 1H). MS-(þ)ESI, m/z: 275 [M þ H]þ.
HR-MS [Mþ•] calcd for C13H7N2O3Cl 274.0145, found 274.0140.
2-(4-Chlorophenyl)-1-oxy-pyrrolo[3,2-b]pyridin-3-one (66).
Yield 75.2%; mp: 196-197 ꢀC. UV (CH3CN) λmax nm (ε
L mol-1 cm-1) 293 (25066). IR (CH2Cl2) cm-1: 3040, 1710,
121.2 (2CH), 119.0 (2CH), 117.1 (CH), 116.8 (C), 103þ.5 (CH),
-
72.0 (CH2). MS-(-)APCI, m/z: 421 [M ]. HR-MS [M 3 ] calcd
3
for C27H19NO4 421.1314, found 421.1322.
2-(6-Methoxynaphtyl)6-benzyloxy-3H-indol-3-one-N-oxide
(58). Yield 7%. Rf 0.32 (cyclohexane/ethyl acetate, 85:15, v/v);
mp: 192 ꢀC. UV (DMF) λmax nm (ε L mol-1 cm-1): 275
3
3
1590, 1525, 1480, 1420, 1384, 1265. 1H NMR (CDCl3, 300 MHz)
δ (ppm) 7.48 (d, J = 8.4 Hz, 2H), 7.59 (dd, J = 5.1 Hz, 7.8 Hz,
1H), 8.00 (d, J = 7.8 Hz, 1H), 8.66 (d, J = 8.4 Hz, 2H), 8.82 (d,
J = 5.1 Hz, 1H). MS-(þ)ESI, m/z: 259 [M þ H]þ.
3
3
(47086). IR (KBr) cm-1: 1702, 1620, 1466, 1388, 1304, 1268.
1H NMR (CDCl3, 500 MHz): δ (ppm) 9.65 (s, 1H), 9.04 (dd, J =
1.5 Hz, J = 9.0 Hz, 2H), 7.94 (d, J = 9.0 Hz, 1H), 7.89 (d, J =
9.0 Hz, 1H), 7.61 (d, J = 8.0 Hz, 1H), 7.57 (d, J = 2.0 Hz, 1H),
7.27-7.30 (m, 2H), 7.36 (t, J = 7.0 Hz, 1H), 7.43 (t, J = 8.0 Hz,
2H), 7.09 (dd, J = 1.5 Hz, J = 9.0 Hz, 2H), 5.23 (s, 2H), 3.78 (s,
3H). 13C NMR (CDCl3, 125 MHz): δ (ppm) 186.1 (C), 164.7 (C),
159.6 (C), 150.0 (C), 136.3 (C), 135.9 (C), 133.1 (C), 131.1 (CH),
128.7 (2CH), 128.6 (C), 128.6 (CH), 128.0 (2CH), 127.1 (CH),
124.9 (CH), 123.7 (CH), 123.6 (CH), 122.2 (C), 119.7 (CH),
115.8 (C), 116.0 (CH), 106.2 (CH), 102.4 (CH), 70.9 (CH2), 55.2
(CH3). MS-(þ)APCI, m/z: 410 [M þ H]þ.
Intermediate 4-Chloro-biphenyle Used to Synthesize (21c).
Iodobenzene (10.0 mmol), 4-chlorophenylboronic acid (14
mmol), palladium acetate (0.3 mmol), 1,4-diazabicyclo-
(2,2,2)octane (0.6 mmol), and potassium carbonate were dis-
solved in DMF (7 mL) and mixed with acetone (110 mL). The
mixture was kept under reflux for 7 h. It was then concentrated
by evaporation. The residue was subsequently resuspended in
water and extracted 3 times by dichloromethane. The organic
phases were collected and washed once with water and then
dried over anhydrous MgSO4, filtered, and evaporated. The
crude product was purified by column chromatography (SiO2)
using petroleum ether as an eluant. Yield 58%.
2-(4-Chloro-3-trifluoromethyl-phenyl)-1-oxy-indol-3-one (59).
Yield 19%; mp: 205.1-205.2 ꢀC. IR (CH2Cl2) cm-1: 3040, 1705,
1
1590, 1510, 1420, 1384, 1265. H NMR (CDCl3, 300 MHz) δ
(ppm) 7.58-7.78 (m, 5H), 8.81 (dd, J = 2.1 and 8.7 Hz, 1H),
9.13 (d, J = 2.1 Hz, 1H). MS-(þ)ESI, m/z: 326 [M þ H]þ. HR-
MS [Mþ•] calcd for C15H7NO2ClF3 325.0117, found 325.0130.
Intermediate 4-Chloro-40-iodo-biphenyle (21c). To a mixture
of 4-chloro-biphenyle (5.7 mmol), acetic acid (2.4 mL), iodine
(2.86 mmol), and sulphuric acid (0.66 mL) was added dropwise

712 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2
Nepveu et al.
fuming nitric acid (0.115 mL). The mixture was stirred at
35-40 ꢀC for 18 h and then diluted in a mixture of water/
dichloromethane. The organic phase was washed with water
until pH 4-5 and then dried with MgSO4, filtered, and con-
centrated by evaporation. The residue was purified by column
chromatography (SiO2) using petroleum ether as an eluant.
Yield 74%.
Intermediate 1-Iodo-4-isopropoxy-benzene (27c). To a mixture
of 4-iodophenol (3.95 mmol) in DMF (7 mL), isopropyl bro-
mide (4.34 mmol) and potassium carbonate (4.42 mmol) were
added. The reaction was stirred under reflux for 4 h. The crude
product obtained after evaporation was dissolved in a mixture
of petroleum ether and sodium hydroxide 1 N (50:50, v/v). The
organic phase dried over anhydrous MgSO4, filtered, evapo-
rated, and purified by column chromatography (SiO2) using
petroleum ether as an eluant. Yield 55%.
General Synthetic Procedure for Intermediate 40-Aryl-4-yl-
trimethylsilane (21d, 27d, 29d, 59d). To a mixture of 4-halo-
geno-aryl (1.59 mmol) and trimethylsilylacetylene (2.39 mmol)
in triethylamine (10 mL) were added tetrakis(triphenylphos-
phine)palladium (0.0239 mmol) and copper(I) iodide (0.0477
mmol). The mixture was stirred at 80 ꢀC for 4 h. After evapora-
tion, the residue was purified by column chromatography (SiO2)
using petroleum ether as an eluant.
General Synthetic Procedure for Intermediate 40-Ethynyl-aryl
(21f, 27f, 29f, 59f). To a mixture of 40-aryl-4-yl-trimethylsilane
(1.2 mmol) in methanol (20 mL) was added potassium carbonate
(0.192 mmol). The mixture was stirred at room temperature for
a night. The mixture was diluted in dichloromethane. The
organic phases were then washed with a saturated solution of
sodium bicarbonate, dried over anhydrous MgSO4, filtered, and
evaporated. The pure product was purified by column chroma-
tography (SiO2) eluant petroleum ether.
General Synthetic Procedure for Intermediate o-Bromo-
nitroaryl (52e, 53e). To a mixture of acetic acid (210 mL) and
fuming nitric acid (46 mL) cooled in ice was added dropwise o-
bromobenzaldehyde (46 mmol) for 20 min in keeping the
temperature below 10 ꢀC. The mixture was stirred at 10 ꢀC for
2 h, and then kept in a mixture of ice/water. The precipated
product was filtered and washed several times with water and
then dried.
6.4. Determination of Log P. Values of Log Pcalc were
calculated with software from Virtual Computational Chemis-
try Laboratory (VCCLAB http://www.virtuallaboratory.org/
lab/alogps/start.html).
6.5. P. falciparum in Vitro Culture and Parasite Growth
Inhibition Assays. 6.5.1. Initial Testing Carried out in Toulouse
(France). RPMI 1640 medium (BioWhittaker, Cambrex (cat. no.
BE12-702F), Belgium) containing L-glutamine (BioWhittaker, cat.
no. BE17-605E), 25 mM HEPES (BioWhittaker, cat no. 17-737F),
and 10% human serum (EFS, Toulouse, France) was used to
cultivate P. falciparum. Human RBCs (group O() for parasite
culture were obtained from EFS in transfusion vials. They were
extensively washed with RPMI medium to discard plasma and
leukocytes. Leucocyte-free erythrocytes were stored at 50% hema-
tocrit (1 volume of RPMI þ 1 volume of packed RBC) for a
maximum period of 21 days. P.f. asexual blood stage parasites
were propagated by incubation at 37 ꢀC in P.f. culture media at
3-5% hematocrit in controlled atmosphere (5% CO2, 100%
relative humidity).33 Parasitized RBCs were maintained in 25 cm2
culture flasks (TPP, Switzerland, ref 90025). Reference drugs,
chloroquine (CQ), and artemether (ART) were obtained from
Sigma (ref C6628) and Cambrex, respectively. CQ was dissolved
in culture medium and ART in ethanol (stock solutions: 10 mg/mL)
and stored at -20 ꢀC prior to testing. For drug assays, serial
drug dilutions were made in P.f. culture media and added to 96-well
(TPP) culture plates. All drugs were tested in triplicate. Plasmo-
dium-infected RBCs were distributed at 2% parasitaemia (2%
hematocrit) in 96-well microtiter plates with different drug
concentrations and incubated for 48 h at 37 ꢀC and 5% of CO2.
[3H]-Hypoxanthine (Perkin-Elmer) was added 24 h after the
beginning of incubation. At the end of incubation (48 h), microtiter
plates were frozen and thawed, and each well was harvested onto a
glass-fiber filter paper. The quantity of incorporated [3H]-hypox-
anthine was determined with a β-counter (1450-Microbeta Trilux,
Wallac-PerkinElmer). Growth-inhibition percentages were plotted
as a semilogarithmic function of drug concentration. The IC50
values were determined by linear regression analysis on the linear
segments of the curves. Assays were repeated three times. Controls
were carried out to assess the background (negative control) and
parasite growth (positive control).
6.5.2. Secondary Testing Carried out in London (U.K.). Drug-
sensitive 3D7 clone of the NF54 isolate (unknown origin)
and chloroquine-, pyrimethamine-, and cycloguanil-resistant
K1 strain (Thailand) came from MR4 (Malaria Research
and Reference Reagent Resource Center, Manassas, VA). Para-
sites were maintained in tissue culture flasks in blood group
Aþ human RBCs at 5% hematocrit in RPMI-1640 supple-
mented with 25 mM HEPES, 24 mM NaHCO3, 0.2% w/v
glucose, 0.03% L-glutamine, 150 μM hypoxanthine, and 0.5%
Albumax II (Gibco, UK) in a 5% CO2 incubator at 37 ꢀC and
the medium changed daily. Synchronization was achieved by a
combination of 5% sorbitol and 60% Percoll gradient centrifu-
gation as described elsewhere. The method used to assess
parasite growth was based on the [3H]hypoxanthine incorpora-
tion assay (with modifications, including the use of Albumax,
rather than 10% human plasma, to supplement the culture
medium.
Stock drug solutions were usually dissolved in 100% di-
methylsulfoxide (Sigma, Dorset, UK) and a 2-fold dilution
series (100, 50, 25, 12.5, 6.25, 3.125, 1.56, 0.78, 0.39, 0.20, 0.10,
and 0.05 μM) of the drugs prepared in assay medium (RPMI-
1640 supplemented with 0.5% Albumax II [Invitrogen], 0.2% w/
v glucose, 0.03% L-glutamine, and 5 μM hypoxanthine) added
to each well of 96-well plates in triplicate. Fifty μL of synchro-
nous P.f. culture (0.5% parasitaemia) or uninfected RBCs were
added to each well to a final hematocrit of 2.5%. All assays
included chloroquine diphosphate (Sigma) and artesunate
(WHO/TDR) as control drugs and control wells with untreated
infected RBCs and uninfected RBCs.
Test plates were incubated at 37 ꢀC in 5% CO2, 95% air
mixture for 24 h, at which point 20 μL (0.1 μCi/well) of [3H]-
hypoxanthine (Perkin-Elmer, Hounslow, UK) were added to
each well. After shaking for 1 min using a plate shaker, the plates
were returned to the incubator and incubated for an additional
24 h. The experiment was terminated by placing the plates in a
-80 ꢀC freezer.
Plates were thawed and harvested onto glass-fiber filter mats
using a 96-well cell harvester (Harvester 96, Tomtec, Oxon, UK)
and left to dry. After the addition of MeltiLex solid scintillant
(PerkinElmer, Hounslow, UK), the incorporated radioactivity
was counted using a Wallac 1450 Betalux scintillation counter
(Wallac). Data acquired by the Wallac BetaLux scintillation
counter were transferred onto a MICROSOFT EXCEL spread-
sheet (Microsoft Corp.), and the IC50/IC90 values of each drug
were calculated by using XLFit (ID Business Solutions Ltd.,
UK) line fitting software.
6.6. In Vitro Cytotoxicity Assay. Cytotoxicity was estimated
on human breast cancer cells (MCF7) at a primary level
(Toulouse, France), The cells were cultured in the same condi-
tions as those used for P. falciparum, except that 10% human
serum was replaced by 10% fetal calf serum (Cambrex). After
trypsinization, cells were distributed in 96-well plates at 2 ꢀ 104
cells/well in 100 μL of culture medium added to 100 μL of the
same medium containing the tested compounds at various
concentrations (the final concentrations in the wells were 1,
10, and 100 μg/mL).
Cell growth was estimated by a colorimetric assay based on
XTT reduction.34 After 48 h of contact between cells and test
compounds, the culture medium was replaced by 50 μL of a

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 2 713
sodium 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazo-
lium-5-carboxanilide inner salt (XTT) (Sigma, Saint Quentin
Fallavier, France) in water solution (0.5 mg/mL), and cells were
incubated for 180 min. XTT was converted into a formazan
product, detected at 450 nm. IC50 values were determined
graphically from dose-response curves (positive control being
doxorubicin [Sigma]). Experiments were performed twice in
triplicate
Cytotoxicity was evaluated on KB cell line at a secondary
level (London, UK). The AlamarBlue (Accumed International
Inc., USA) method was used to assess cytotoxicity to the KB cell
line. Microtiter plates were seeded at a density of 4 ꢀ 104 KB
cells/mL in RPMI 1640 culture medium supplemented with 10%
heat-inactivated fetal calf serum (complete medium) (Seralab,
Inc.) and incubated at 37 ꢀC, 5% CO2, 95% air mixture for 24 h,
followed by compound addition to triplicate wells in a dilution
series in complete medium. The positive control drug was
podophyllotoxin (Sigma). Plates were incubated for a further
72 h, followed by the addition of 10 μL of Alamar-Blue
(AccuMed International Inc.) to each well and incubation for
2-4 h at 37 ꢀC, 5% CO2, 95% air mixture. Fluorescence
emission at 585 nm was measured in a SPECTRAMAX GE-
MINI plate reader (Molecular Devices Inc.) after excitation at
530 nm. ED50 values were calculated using XLFit (ID Business
Solutions Ltd., UK) line fitting software.
alcohol, 0.4% Tween 80, 0.9% NaCl (all Sigma)]. For intraper-
itoneal administration, compounds were dissolved in 0.05%
Tween 80. Mice were infected intravenously with 2 ꢀ 106
infected red cells, randomized, and divided in groups of five
mice for each dose (day 0). Oral treatment by gavage started 3 h
postinfection and continued on day 1, 2, and 3 postinfection
once a day. Parasitaemia was determined by microscopic ex-
amination of Giemsa stained blood films taken on day 4.
Microscopic counts of blood films from each mouse were
exported into a Microsoft Excel spreadsheet (Microsoft Corp.)
and expressed as percentages of inhibition from the arithmetic
mean parasitaemias of each group in relation to the untreated
group. Dose-response curves were obtained and ED50 and
ED90 values and 95% CI calculated using XLFit version 4 (ID
Business Solutions Ltd., UK) line fitting software.
Acknowledgment. This work was supported by the Euro-
pean Union (Redox Antimalarial Drug Discovery, Read-
Up, FP6-2004-LSH-2004-2.3.0-7, Strep no. 018602). Thanks
are due to the Fondation Pierre Fabre for awarding a
fellowship to S.K. We thank E. Pelissou, E. Augugliaro,
S. Vomshied, A. Lallemand, A. Boyer, P. Lajoie, E. Najahi,
S. Casties, B. Moukarzel, and Shereen Nasser for technical
assistance.
6.7. In Vivo Assays. In vivo studies were approved by the
appropriate institutional animal care and use committee.
The Primary Assay (Toulouse, France) with compound 1 of in
vivo antimalarial efficacy was achieved using the P. berghei
rodent malaria 4-day suppressive test. The compound was
dissolved in 100% dimethyl sulfoxide (DMSO).
Animal and Conditions: mice, Swiss females, 20 ( 2 g; series:
10 animals per cage; control group: 10 animals; cages: standard;
maintenance: conditioned air at 20 ꢀC and 50-60% relative
humidity; diet with standard food and water ad libitum.
Test Procedure, Day 0: Heparinized blood was taken from a
donor mouse with approximately 30% parasitaemia and diluted
in physiological saline to 108 parasitized RBC per mL. An
aliquot of 0.1 mL (107 parasitized RBC) of this suspension
was injected intraperitoneally (ip) into experimental groups of
10 mice. Two h postinjection, the experimental groups were
treated with a single dose of test compound, prepared in appro-
priate dilution in DMSO of the stock solution in DMSO, by
intraperitoneally (ip) route; compound 1 was tested at six
different doses.
Supporting Information Available: Hematin binding assays,
metabolism of some selected compounds by liver microsomes,
lactate dehydrogenase (LDH) assay, elemental analysis for
some compounds, selected 1H, 13C NMR, and IR spectra, and
references cited. This material is available free of charge via the
Internet at http://pubs.acs.org.
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