Synthesis and antimalarial activity of carbamate and amide derivatives of 4-anilinoquinoline

Article abstract of DOI:10.1016/j.ejmech.2007.11.003

A series of 4-anilinoquinolines bearing an amino side chain linked to the aromatic ring with a carbamate or an amide bond were synthesized and evaluated for their antimalarial activity and their cytotoxicity upon MRC-5 cells. Among the 17 compounds, a majority was found to be active in the low nanomolar range against both chloroquine-sensitive and -resistant strains of Plasmodium falciparum in vitro with relative low cytotoxicity. Two compounds were then tested on mice infected by Plasmodium berghei and were found to exhibit reasonable in vivo activity.

Full text of DOI:10.1016/j.ejmech.2007.11.003

Available online at www.sciencedirect.com
European Journal of Medicinal Chemistry 43 (2008) 2045e2055
http://www.elsevier.com/locate/ejmech
Original article
Synthesis and antimalarial activity of carbamate and amide
derivatives of 4-anilinoquinoline
Sandrine Delarue-Cochin a,1, Philippe Grellier b, Louis Maes c,2,
Elisabeth Mouray b, Christian Sergheraert a, Patricia Melnyk a,d,
*
a UMR CNRS 8525, Universite´ de Lille II, Institut Pasteur de Lille, 1 rue du Professeur Calmette, B.P. 447, 59021 Lille cedex, France
b USM504-EA3335, De´partement RDDM, CP52, Muse´um National d’Histoire Naturelle, 61 rue Buffon, 75005 Paris, France
c Tibotec, B-32800 Mechelen, Belgium
d UMR CNRS 8161, Universite´s de Lille I & II, Institut Pasteur de Lille, 1 rue du Professeur Calmette, B.P. 447, 59021 Lille cedex, France
Received 4 May 2007; received in revised form 9 July 2007; accepted 12 November 2007
Available online 19 November 2007
Abstract
A series of 4-anilinoquinolines bearing an amino side chain linked to the aromatic ring with a carbamate or an amide bond were synthesized
and evaluated for their antimalarial activity and their cytotoxicity upon MRC-5 cells. Among the 17 compounds, a majority was found to be
active in the low nanomolar range against both chloroquine-sensitive and -resistant strains of Plasmodium falciparum in vitro with relative
low cytotoxicity. Two compounds were then tested on mice infected by Plasmodium berghei and were found to exhibit reasonable in vivo
activity.
Ó 2007 Elsevier Masson SAS. All rights reserved.
Keywords: Drug design; Plasmodium falciparum; Antimalarials; Amodiaquine; 4-Aminoquinolines
1. Introduction
world’s population exposed to risk of infection and 2 million
deaths each year [1]. Chloroquine (CQ, Chart 1) has been
the mainstream drug in the fight against Plasmodium falcipa-
rum since the 1950s, but its efficacy is eroded by the emer-
gence of resistant parasites. Development of drug resistance
to CQ in malaria has become a major health concern and
has so prompted the re-examination of alternative antimalar-
ials such as amodiaquine (AQ, Chart 1), a 4-aminoquinoline
where the alkyl chain of CQ is replaced by an aromatic ring.
AQ proved to be effective against CQ-resistant strains [2]
and comparative trials of CQ and AQ for the treatment of
acute, uncomplicated infections in Gambia, in West and
Central Africa and in Nigeria showed that AQ was superior
to CQ, displaying lower parasitological and clinical failure
rates [3e5]. However, the clinical use of AQ has been severely
restricted since the mid-1980s because cases of agranulocyto-
sis and hepatotoxicity were observed with its long term use
[6,7]. This AQ toxicity is thought to be due to extensive
metabolization of the 4-hydroxyanilino moiety to its quinonei-
mine variant [8,9]. However, WHO’s guidelines for treatment
Malaria can be considered actually as one of the most wide-
spread diseases in the world with almost one-half of the
Abbreviations: AQ, amodiaquine; CQ, chloroquine; DMAP, 4-(dimethtyla-
mino)pyridine; DSC, N,N’-disuccinimidylcarbonate; MTT, 3-(4,5-dimethylth-
iazol-2-yl)-2,5-diphenyl tetrazolium bromide (thiazolyl blue); PHPLC, purity
determined by HPLC; TLC, thick-layer chromatography; tR, HPLC retention
time; DMAP, 4-dimethylaminopyridine; DSC, N,N’-disuccinimidylcarbonate.
´
* Corresponding author. UMR CNRS 8161, Universites de Lille I & II, In-
stitut Pasteur de Lille, 1 rue du Professeur Calmette, B.P. 447, 59021 Lille ce-
dex, France. Tel.: þ33 3 20 87 12 19; fax: þ33 3 20 87 12 35.
E-mail address: patricia.melnyk@ibl.fr (P. Melnyk).
´
Present address: Universite Paris-Sud, CNRS and IFR141 ‘‘Innovation
Therapeutique: du Fondamental au Medicament’’, Equipe de Synthese Organ-
1
´
´
`
ˆ
ique et de Pharmacochimie, UMR CNRS 8076, Chatenay-Malabry, F-92296,
France.
2
Present address: Laboratory of Microbiology, Parasitology and Hygiene
(LMPH), Faculty of Pharmaceutical, Biomedical and Veterinary Sciences,
Antwerp University, Campus Groenenborger-Groenenborgerlaan 171,
B-2020 Antwerp-Wilrijk, Belgium.
0223-5234/$ - see front matter Ó 2007 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2007.11.003

2046
S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
OH
followed by reaction with an amine. However, only two
methods proved to be relatively efficient depending on the de-
sired compound: reaction of alcohol 1 with isocyanates gener-
ated by means of triphosgene or activation of this alcohol with
DSC. With other methods, starting material was either recov-
ered or degraded.
NEt2
NEt2
HN
HN
For compounds 2e4, activation of alcohol 1 was realized
thanks to DSC (Scheme 2) [17]. Activated intermediate was
then subjected to an amine to give the desired carbamate. Amine
was N,N-dimethylethylenediamine for compound 2 which was
obtained with a moderate 24% yield. For compounds 3 and 4,
a two-step procedure was used with reaction of bromoethyl-
amine followed by substitution of the bromine atom by either
piperidine or morpholine. It led to compounds 3 and 4 with
low to moderate yields (11% and 29% yields, respectively).
For compounds 5e7, alcohol 1 was subjected to an isocy-
anate to give the desired carbamates (Scheme 3). This isocya-
nate was commercial for compound 7 or was generated in situ
by reaction of the appropriate amine with triphosgene [12] for
compounds 5 and 6. Carbamates 5e7 were so obtained in low
to moderate yields (33%, 17% and 11% yields, respectively).
Acid derivative 8 was obtained in excellent yield, around90%,
by oxidation of compound 1 using periodic acid with catalytic
amountofchromiumoxide(Scheme 4)[19]. Itwasthensubjected
to various amines with PyBrop as activating agent and diethyl-
amine as base (Scheme 4). This reaction afforded amides 9e18
in moderate to good yields, from 11% to 75% yield, primary
amines giving lower yields in general than secondary amines.
Cl
N
Cl
N
Chloroquine (CQ)
Amodiaquine (AQ)
Chart 1. Structure of chloroquine and amodiaquine.
still recommend the use of AQ in combination with artemisi-
nin derivatives such as artesunate, named ArsucamÒ, or if not
available with sulfadoxine/pyrimethamine.
We synthesized 4-anilinoquinolines retaining the aromatic
ring of AQ but lacking the hydroxyl group responsible for
toxicity in order to access compounds as active as AQ and pre-
senting lower cytotoxicity [10,11]. It led notably to compound
1 and its ester or amine derivatives which presented good in
vitro antimalarial activity combined to low cytotoxicity and
reasonable in vivo antimalarial activity. In order to complete
this structureeactivity relationships study and optimize the
second side chain in this 4-anilinoquinoline series, we present
here the synthesis and evaluation of two series of compound 1
derivatives: the carbamate ones 2e7 (Chart 2), and the acid or
amide ones 8e18 (Chart 3).
2. Chemistry
3. Biological results and discussion
Compounds 2e18 were obtained from compound 1, which
was synthesized in a three-step procedure according to De-
larue et al (Scheme 1) [10].
Several methods were considered in order to synthesize car-
bamates 2e7: reaction of alcohol 1 with commercial or in situ
generated isocyanates (by means of triphosgene, 1,10-carbon-
yldiimidazole or Boc2O/DMAP) [12e15], or activation of
alcohol 1 by means of triphosgene, N,N0-disuccinimidylcar-
bonate (DSC) or para-nitrophenylchloroformate [12,16e18]
All the compounds were tested for their activity against the
CQ-resistant strain FcB1R of P. falciparum (CQ: IC50
¼
126 nM). Forcompounds 1e4 and 16, activitywas also measured
against a CQ-sensitive strain Thai (CQ: IC50 ¼ 14 nM), and
another CQ-resistant strain K1 (CQ: IC50 ¼ 183 nM) (Table 1).
In the carbamate series, all compounds 2e7 were found to
be at least as active as compound 1, three presenting IC50
around 10 nM (compounds 2, 3 and 5). Those compounds
H
OH
O
N
R
O
O
O
N
N
HN
N
N
H
HN
N
N
H
Cl
Cl
1
2 R = (CH2)2-NMe2
3 R = (CH2)2-piperidine
4 R = (CH2)2-morpholine
5 R = (CH2)3-NEt2
6 R = (CH2)3-piperazine
7 R = CH2Ph
Chart 2. General structures of compounds 1e7.

S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
OH R'
2047
O
O
O
O
N
N
HN
N
N
H
HN
N
N
H
Cl
Cl
9 R' = NH-(CH2)2-NMe2
8
10 R' = -NH-(CH2)3-NMe2
11 R' = NMe-(CH2)2-NMe2
12 R' = NH-(CH2)2-pyrrolidine
13 R' = NH-(CH2)2-morpholine
14 R' = piperidine
15 R' = morpholine
16 R' = N-methylpiperazine
17 R' = N-phenylpiperazine
18 R' = N-benzylpiperazine
Chart 3. General structures of compounds 8e18.
were active whatever the strain used. On the contrary to com-
pound 1, which lost a great part of its activity against K1
strain, compounds 3 and 4 improve their IC50 against this
highly resistant strain. By comparing the influence of the car-
bamate side chain on activity, we could deduce some struc-
tureeactivity relationships: (i) presence of an amine on the
side chain improved the activity (compound 2, 3 and 5 versus
compound 7), (ii) the length of the carbon side chain between
carbamate bond and terminal amine could comprised two or
three methylenes without any influence on activity (com-
pounds 2 and 5 versus compound 6), (iii) the tertiary amine
could be cyclic or not (compounds 2 and 3), (iv) presence of
another electronegative atom on the side chain caused a small
decrease in activity (compounds 4 and 6, Chart 2).
Acid derivative 8 was found to display no antimalarial activ-
ity. This lack of activity is not surprising and could be explained
by a bad cellular penetration due to the acidic function.
In the amide series, five compounds were found to be at
least as active as compound 1, with IC50 between 17 and
50 nM (compounds 9, 11, 12, 16 and 18, Chart 3). Compound
16 was active whatever the strain used (with a slight improve-
ment against FcB1R strain) as for compounds 2e4 in the car-
bamate series. Some structureeactivity relationships could be
deduced in the amide series: (i) presence of an amine on the
side chain is required to improve activity (compounds 9,
11e13 and 16e18 versus compounds 14 and 15), (ii) the
length of the carbon side chain between amide and terminal
amine must be of two methylenes but not three (compound
9 versus compound 10), (iii) the terminal amine could be cy-
clic or not but slightly higher activity was observed for cyclic
amine (compounds 9 and 12), (iv) presence of another electro-
negative atom on the side chain caused a small decrease in ac-
tivity (compounds 13), (v) tertiary amide was preferred to
secondary amide (compounds 9 and 11).
The relationships in the amide and carbamate series are
very similar except for the chain length which is crucial for
the amide derivatives but not for the carbamate ones. It can
be assumed that terminal amine in those side chains must be
separated from the aromatic moiety by 4, 6 or 7 methylene
units but not 5 (compounds 9 and 2 versus compound 10).
This is confirmed by previous results observed in the ester se-
ries [10] (compounds 19e24, Chart 4, Table 2) or in the amine
series [10] (compounds 25e29, Chart 4, Table 2). In those
cases a slight decrease of activity was observed when terminal
amine was separated by 5 methylene units from the aromatic
moiety compared to molecules presenting separation of 4, 6
and 7 methylene units (compound 20 versus compounds 19,
21 and 22 for ester series, compound 26 versus compound
OH
OH
OH
O
N
i
ii, iii
HN
N
NH2
HN
N
N
H
H2N
NH2
Cl
Cl
Scheme 1. Synthesis of compound 1. Reagents: (i) 4,7-dichloroquinoline, N-methylmorpholine, EtOH/CHCl3 9/1, rt, 48 h; (ii) chloroacetic acid, N-ethylpiperidine,
PyBroP, DMF, rt, 7 h; (iii) 4-methylpiperidine, reflux, 5 h.

2048
S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
H
O
N
NMe2
O
O
N
HN
N
N
H
OH
O
Cl
2
i, ii
N
HN
N
N
H
H
N
O
N
X
i, iii
O
1
Cl
O
N
HN
N
N
H
3 X' = CH2
4 X = O
Cl
Scheme 2. Synthesis of compounds 2e4. Reagents: (i) DSC, pyridine, DCM, rt, overnight; (ii) N,N-dimethylethylenediamine, DMAP, rt, overnight; (iii) bromoe-
thylamine bromhydrate, DMAP, rt, overnight then piperidine for compound 3 or morpholine for compound 4, DCM, reflux, overnight.
25 for amine series). There exist, however, two major differ-
ences between carbamate/amide and ester/amine series (Table
2): (i) in the ester series, cyclic amine was found to be highly
superior to the acyclic ones (compounds 21 and 24 versus
compound 23), result not observed in the carbamate/amide/
amine series (compounds 2 and 3 in the carbamate series,
compounds 9 and 12 in the amide series, compounds 25, 26
and 27 in the amine series), (ii) no real influence of an addi-
tional electronegative atom was observed in the ester/amine
series (compounds 21 and 24 in the ester series, compounds
28 and 29 in the amine series), while in the carbamate/amide se-
ries, introduction of an additional electronegative atom caused
decrease of activity (compound 4 versus compound 3 in the
carbamate series, compound 13 versus 12 in the amide series).
H
O
N
NRR'
O
O
N
HN
N
N
H
OH
5 NRR' = NEt2
O
Cl
6 NRR' = N-methylpiperazine
i
N
HN
N
N
H
H
O
N
ii
O
1
Cl
O
N
HN
N
N
H
7
Cl
Scheme 3. Synthesis of compounds 5e7. Reagents: (i) 3-diethylaminopropylisocyanate (for compound 5) or 3-(4-methylpiperazino)-propylisocyanate (for com-
pound 6), DCM/DMF 10/1, rt, overnight; (ii) benzylisocyanate, DCM/DMF 10/1, rt, overnight.

S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
2049
O
OH
O
NRR'
O
O
ii
i
N
N
1
HN
N
N
H
HN
N
N
H
Cl
Cl
8
14 NRR' = piperidine
9 NRR' = NH-(CH2)2-NMe2
15 NRR' = morpholine
10 NRR' = -NH-(CH2)3-NMe2
11 NRR' = NMe-(CH2)2-NMe2
12 NRR' = NH-(CH2)2-pyrrolidine
13 NRR' = NH-(CH2)2-morpholine
16 NRR' = N-methylpiperazine
17 NRR' = N-phenylpiperazine
18 NRR' = N-benzylpiperazine
Scheme 4. Synthesis of compounds 8e18. Reagents: (i) periodic acid, CrO3 cat., MeCN, rt, 2 h; (ii) HNRR0, PyBroP, DIEA, DCM/DMF 10/1, rt, overnight.
The cytotoxicity of the different compounds was then eval-
uated upon human MRC-5 cells (diploid embryonic lung cell
line) (Table 1) and expressed as a CC50 which is the concen-
tration of drug causing 50% cytotoxicity. In the carbamate and
amide series, we could observe in general a slight decrease in
cytotoxicity compared to compound 1 with the exception of
compounds 2, 5 and 6. For the amide and the carbamate series,
on the contrary to the ester series, morpholine substituent did
not decrease cytotoxicity (Table 2).
selectivity index is better for carbamate and amide series
than for ester and amine series.
Hence it was encouraging to consider compound 3 for fur-
ther evaluation, which presented the best activity in the carba-
mate series with good selectivity index (1114). In the amide
series compound 16 was preferred to compound 12 which pre-
sented, however, a better selectivity index (1024 versus 833)
with a similar in vitro activity (around 15 nM). Indeed,
compound 12 was hardly obtained with poor yield (16%), while
compound 16 was obtained easily with a moderate 32% yield.
Compounds 3 and 16 were so tested in vivo at the concen-
tration of 40 mg/kg/day (Table 3) and were compared to the
All the derivatives revealed a selectivity index (CC50/IC50
on FcB1R resistant strain) superior to that of CQ, except com-
pounds 6, 10, 14 and 17. We could remark that, in general,
Table 1
In vitro antimalarial activity upon three Plasmodium falciparum strains, in vitro cytotoxicity of compounds 1e18 and selectivity index against FcB1R strain
a
IC50 (nM)
Compound
SIf
CC50 (mM) upon MRC-5 cellsg
Thai
FcB1R
K1
CQ
AQ
1
14.3 ꢀ 2.4e
126 ꢀ 26e
183 ꢀ 35e
397
50
12
4.6 ꢀ 0.8e
4.8 ꢀ 0.9e
9.4 ꢀ 1.1e
2500
281
89.6 ꢀ 13.1c
44.5 ꢀ 9.7c
151.6 ꢀ 48e
12.5
3.1
b
2
nd
19.4 ꢀ 6.5c
10.2
nd
304
3
11.2 ꢀ 2.5d
9.7 ꢀ 0.8c
1116
>259
434
12.5
>12.5
6.3
b
4
38.6 ꢀ 4c
48.3
14.5
19.4 ꢀ 8.7c
b
5
nd
nd
nd
nd
6
44 ꢀ 16c
143
6.3
7
nd
nd
36 ꢀ 2c
nd
nd
>347
nd
478
>12.5
>32
15
8
>1000
9
nd
nd
31.4 ꢀ 12.9c
181.1 ꢀ 49.5c
18.8 ꢀ 0.9c
nd
nd
10
11
12
13
14
15
16
17
18
>177
851
>32
16
nd
nd
nd
nd
16.6 ꢀ 0.8d
58.6 ꢀ 3.4c
1024
>546
>112
> 298
833
17
nd
nd
nd
nd
>32
>32
>32
16
285.6 ꢀ 19.7c
107.5 ꢀ 22.7c
19.2 ꢀ 0.7c
nd
nd
38.4 ꢀ 1.6c
51.2 ꢀ 8.6c
nd
nd
151.1 ꢀ 9.5c
50.6 ꢀ 9.0c
nd
nd
106
296
16
15
nd: not determined.
a
Parasites were considered resistant to CQ for IC50 > 100 nM.
Number of experiments: n ¼ 2.
b
c
d
e
f
Number of experiments: n ¼ 3.
Number of experiments: n ¼ 4.
Number of experiments: n ¼ 6.
SI: selectivity index (CC50 MRC-5/IC50 FcB1R).
CC50 is the IC50 value for cytotoxicity calculated on the basis of three experiments.
g

2050
S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
H
N
(
)
n
O
NRR'
O
NRR'
N
(
)
n
O
O
N
HN
N
N
H
HN
N
N
H
Cl
Cl
19 n=1, NRR'=piperidine
20 n=2, NRR'=piperidine
21 n=4, NRR'=piperidine
22 n=5, NRR'=piperidine
23 n=4, NRR'=NEt2
25 n=2, NRR'=NMe2
26 n=3, NRR'=NMe2
27 n=2, NRR'=pyrrolidine
28 n=2, NRR'=piperidine
29 n=2, NRR'=morpholine
24 n=4, NRR'=morpholine
Chart 4. General structures of compounds 19e29.
already known in vivo activity of compound 1, ester 24 and
amine 29 which were the most active of the ester and amine
series [10]. Carbamate 3 exhibited an activity in vivo similar
to ester 24 (50% and 73% MST, respectively), while amide
16 was more active in vivo than ester 24 and compound 1
with a 150% MST. But those two compounds 3 and 16 did
not exhibit better in vivo activity than amine 29, which was
found to cure mice at the same dose. No toxicity was observed
in mice.
matrix. The purity of final compounds 2e18 was verified by
high pressure liquid chromatography (HPLC) using C18
Vydac (C18 V) column. Analytical HPLC was performed on
a Shimadzu system equipped with a UV detector set at
254 nm. Compounds were dissolved in EtOH and injected
through a 50 mL loop. The following eluent systems were
used: A (H2O/TFA 100/0.05) and B (CH3CN/H2O/TFA 80/
20/0.05). HPLC retention times (tR) were obtained, at flow
rates of 1 mL/min, using the following conditions: a gradient
run from 100% eluent A for 5 min, then to 100% eluent B
over the next 30 min. 4,7-Dichloroquinoline was obtained
from Acros and other reagents from Acros, Aldrich, Avocado
and Lancaster. Compound 1 was synthesized according to the
procedure described by Delarue et al. [10].
4. Conclusion
This carbamate and amide series present interesting com-
pounds in term of in vitro antimalarial activity and cytotoxicity
and contribute to complete the structureeactivity relationships
study carried out on 4-anilinoquinoline 1 derivatives. It has
been proven that introduction of the amide or carbamate linker
improves the efficiency of those derivatives compared to the
ester linker in term of selectivity index and in vivo activity.
Compound 16 presents the best results in those two series
with high in vitro activity whatever the strains used, a good se-
lectivity index and a reasonable activity in vivo.
5.1.1. (2-Dimethylamino-ethyl)-carbamic acid 3-(7-chloro-
quinolin-4-ylamino)-5-(2-piperidin-1-yl-acetylamino)-
benzyl ester (2)
To a solution of compound 1 (200 mg, 0.47 mmol, 1 eq.) in
anhydrous DCM (10 mL) were added, at 0 ꢁC, DSC (483 mg,
1.89 mmol, 4 eq.) and pyridine (152 mL, 1.88 mmol, 4 eq.).
After stirring the mixture at room temperature overnight,
Et2O (20 mL) was added and the reactive medium filtered.
The precipitate was solubilized in anhydrous DMF (10 mL)
with N,N-dimethylethylenediamine (124 mg, 1.42 mmol,
3 eq.) and DMAP (173 mg, 1.43 mmol, 3 eq.). After further
stirring at room temperature overnight, the reactive medium
was concentrated and the residue was taken up in DCM
(10 mL), the organic layer was washed twice with NaHCO3
1 M (2 ꢂ 10 mL) and brine (10 mL), dried over MgSO4, fil-
tered and concentrated. The residue was finally purified by
TLC (DCM/MeOH/NH4OH 85/15/0.5) to yield compound 2
as a yellow solid (60 mg, 24% yield); Rf 0.30 (DCM/MeOH/
NH4OH 85/15/1); mp ¼ 80 ꢁC; PHPLC 90%; tR ¼ 14.13 min;
1H NMR (DMSO-d6) d (ppm): 1.37e1.39 (m, 2H, CH2),
1.51e1.58 (m, 4H, CH2), 2.12 (s, 6H, N(CH3)2), 2.28 (t,
J ¼ 6.7 Hz, 2H, CH2NMe2), 2.43e2.46 (m, 4H, CH2), 3.04e
3.10 (m, 2H, CH2NHCO), 3.06 (s, 2H, COCH2N), 4.98 (s,
2H, CH2Ar), 6.96 (d, J ¼ 5.3 Hz, 1H, Quin-H3),
7.00 þ 7.32 þ 7.72 (s, 3H, Arom-H), 7.16 (t, J ¼ 5.7 Hz, 1H,
OCONH), 7.56 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H, Quin-H6),
5. Materials and methods
5.1. Chemistry
All reactions were monitored by thin-layer chromatography
carried out on 0.2 mm E. Merck silica gel plates (60F-254) us-
ing UV light as a visualizing agent. Chromatography was un-
dertaken using silica gel 60 (230e400 mesh ASTM) from
Macherey-Nagel. Thick-layer chromatography (TLC) was per-
formed using silica gel from Merck, from which the com-
pounds were extracted by the following solvent system:
CH2Cl2/MeOH/NH4OH 80/20/1. All melting points were de-
termined on a Buchi melting point apparatus and were uncor-
¨
rected. NMR spectra were obtained using a Bruker 300 MHz
spectrometer and chemical shifts (d) were expressed in ppm
relative to TMS used as an internal standard. Mass spectra
were recorded on a MALDI-TOF Voyager-DE STR (Applied
Biosystems, Palo Alto, CA) with a trihydroxy-acetophenone

Table 2
Comparison of in vitro antimalarial activity (nM) and selectivity index against FcB1R strain between the carbamate, ester, amide and amine derivatives
Carbamates
Esters [10]
Amides
Amines [10]
NRR0
H
N
O
(
)n
H
N
H
N
NRR'
NRR'
NRR'
O
NRR'
O
(
)n
(
)n
( )n
O
O
HN
N
NH
HN
N
NH
HN
N
NH
HN
N
NH
O
O
O
O
N
N
Cl
N
N
Cl
Cl
Cl
a,b
a,b
a,b
a,b
Ref
2
n
2
IC50
10.2
SIc
304
Ref
n
IC50
SIc
Ref
9
n
2
3
IC50
31.4
181.1
SIc
478
Ref
25
n
2
3
IC50
15.5
134
SIc
1154
174
N
10
>177
26
5
3
14.5
434
23
4
141.1
89
N
N
12
2
16.6
1024
27
2
10.2
182
19
20
21
22
24
1
2
4
5
4
54
231
158
806
456
887
N
3
4
2
2
11.2
48.3
1116
434
78.9
15.5
27.4
14.1
28
29
2
2
6.6
7.9
648
406
13
2
58.6
546
N
O
a
Parasites were considered resistant to CQ for IC50 > 100 nM.
n, number of experiments between 3 and 6.
SI: selectivity index (CC50 MRC-5/IC50 FcB1R).
b
c

2052
S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
Table 3
Antimalarial activity of compounds 1, 3, 16, 24 and 29 on P. berghei in mice
1.52e1.56 (m, 4H, CH2), 2.30e2.42 (m, 10H, CH2), 2.44e
2.47 (m, 4H, CH2), 3.05 (s, 2H, COCH2N), 3.47e3.49 (m,
2H, CH2NHCO), 4.99 (s, 2H, CH2Ar), 6.95 (d, J ¼ 5.3 Hz,
1H, Quin-H3), 7.02 þ 7.32 þ 7.71 (s, 3H, Arom-H), 7.16 (t,
J ¼ 5.5 Hz, 1H, OCONH), 7.56 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz,
1H, Quin-H6), 7.89 (d, J ¼ 2.2 Hz, 1H, Quin-H8), 8.42 (d,
J ¼ 9.1 Hz, 1H, Quin-H5), 8.47 (d, J ¼ 5.3 Hz, 1H, Quin-H2),
9.15 (s, 1H, NH), 9.75 (s, 1H, CONH); m/z 580.5 (Mþ þ 1).
Compound
Dose
(mg/kg)
Reduction
(%) of parasitaemia, day 4
Excess MST
(%)
CQ
AQ
1
10
10
40
40
40
40
40
100
100
100
100
99.96
99.8
100
Cured
Cured
110
3
50
158
16
24
29
73
Cured
5.1.3. General procedure for the synthesis of
compounds 5e8
Excess MST was defined as the percentage of prolongation of the mean
survival time of treated mice compared to that of control mice.
To a solution of 3-(diethylamino)propylamine or 1-(3-ami-
nopropyl)-4-methylpiperazine (0.26 mmol, 1 eq.) in anhydrous
DCM (5 mL) were added, at 0 ꢁC, triphosgene (26 mg,
0.09 mmol, 0.33 eq.) and, after 5 min, NEt3 (109 mL,
0.78 mmol, 3 eq.). After stirring the mixture for 10 min at
room temperature and cooling to 0 ꢁC, a solution of compound
1 (100 mg, 0.24 mmol, 0.9 eq.) in an anhydrous DCM/DMF 10/
1 (5.5 mL) was added dropwise. After further stirring at room
temperature overnight, the mixture was washed with
2 ꢂ 10 mL NaHCO3 1 M, the organic layer dried over MgSO4,
filtered and concentrated. The residue was purified by TLC
(DCM/MeOH/NH4OH 85/15/1) to yield the desired compound.
7.89 (d, J ¼ 2.2 Hz, 1H, Quin-H8), 8.42 (d, J ¼ 9.1 Hz, 1H,
Quin-H5), 8.47 (d, J ¼ 5.4 Hz, 1H, Quin-H2), 9.15 (s, 1H,
NH), 9.76 (s, 1H, CONH); m/z 538.2 (Mþ þ 1).
5.1.2. General procedure for synthesis of compounds
3 and 4
To a solution of compound 1 (200 mg, 0.47 mmol, 1 eq.) in
anhydrous DCM (10 mL) were added, at 0 ꢁC, DSC (483 mg,
1.89 mmol, 4 eq.) and pyridine (152 mL, 1.88 mmol, 4 eq.).
After stirring the mixture at room temperature overnight,
Et2O (20 mL) was added and the reactive medium filtered.
The precipitate was solubilized in anhydrous DMF (10 mL)
with bromoethylamine bromhydrate (290 mg, 1.42 mmol,
3 eq.) and DMAP (173 mg, 1.43 mmol, 3 eq.). After stirring
the mixture at room temperature overnight, a solution of piper-
idine or morpholine (2.36 mmol, 5 eq.) in DCM (10 mL) was
added. After further stirring at reflux overnight, the reactive
medium was concentrated and the residue was taken up in
DCM (10 mL), the organic layer was washed twice with
NaHCO3 1 M (2 ꢂ 10 mL) and brine (10 mL), dried over
MgSO4, filtered and concentrated. The residue was finally pu-
rified by TLC (DCM/MeOH/NH4OH 85/15/0.5) to yield the
desired compound.
5.1.3.1. (3-Diethylamino-propyl)-carbamic acid 3-(7-chloro-
quinolin-4-ylamino)-5-(2-piperidin-1-yl-acetylamino)-benzyl
ester (5). Yellow solid (45 mg, 33% yield); Rf 0.55 (DCM/
MeOH/NH4OH 85/15/1); PHPLC 95%, tR ¼ 14.38 min; 1H
NMR (DMSO-d6)
d
(ppm): 0.87 (t, J ¼ 7.1 Hz, 6H,
N(CH2CH3)2), 1.37e1.40 (m, 2H, CH2), 1.45e1.55 (m, 6H,
CH2), 2.28e2.33 (m, 2H, CH2), 2.36 (q, J ¼ 7.1 Hz, 4H,
N(CH2CH3)2), 2.42e2.46 (m, 4H, CH2), 2.99 (m, 2H,
CH2CONH), 3.05 (s, 2H, COCH2N), 4.98 (s, 2H, CH2Ar),
6.95 (d, J ¼ 5.3 Hz, 1H, Quin-H3), 7.02 þ 7.31 þ 7.72 (s, 3H,
Arom-H), 7.25 (t, J ¼ 5.5 Hz, 1H, OCONH), 7.55 (dd,
J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H, Quin-H6), 7.88 (d, J ¼ 2.2 Hz,
1H, Quin-H8), 8.42 (d, J ¼ 9.1 Hz, 1H, Quin-H5), 8.47 (d,
J ¼ 5.3 Hz, 1H, Quin-H2, 9.15 (s, 1H, NH), 9.76 (s, 1H,
5.1.2.1. (2-Piperidin-1-yl-ethyl)-carbamic acid 3-(7-chloro-
quinolin-4-ylamino)-5-(2-piperidin-1-yl-acetylamino)-benzyl
ester (3). Yellow solid (30 mg, 11% yield); Rf 0.20 (DCM/
MeOH/NH4OH 85/15/1); PHPLC 97%, tR ¼ 14.51 min; 1H
NMR (DMSO-d6) d (ppm): 1.24e1.56 (m, 12H, CH2),
2.56e2.61 (m, 10H, CH2), 3.12 (s, 2H, COCH2N), 3.19e
3.21 (m, 2H, CH2NHCO), 5.01 (s, 2H, CH2Ar), 6.97 (d,
J ¼ 5.3 Hz, 1H, Quin-H3), 7.04 þ 7.36 þ 7.71 (s, 3H, Arom-
H), 7.30 (broad s, 1H, OCONH), 7.57 (dd, J ¼ 9.0 Hz,
J ¼ 2.2 Hz, 1H, Quin-H6), 7.90 (d, J ¼ 2.3 Hz, 1H, Quin-
H8), 8.44 (d, J ¼ 9.1 Hz, 1H, Quin-H5), 8.48 (d, J ¼ 5.3 Hz,
1H, Quin-H2), 9.18 (s, 1H, NH), 9.83 (s, 1H, CONH); m/z
579.2 (Mþ þ 1).
CONH); 13C NMR (DMSO-d6)
d
(ppm): 12.53
(N(CH2CH3)2), 24.40 (CH2), 26.34 (CH2), 27.84 (CH2), 47.07
(N(CH2CH3)2), 50.70 (CH2), 54.89 (CH2), 63.52 (COCH2N),
65.68 (ArCH2), 103.07 (Quin-C3), 113.27 þ 114.84 þ 117.08
(Arom-CH), 125.44 (Quin-C5), 125.86 (Quin-C6), 128.53
(Quin-C8), 152.78 (Quin-C2); m/z 580.3 (Mþ þ 1).
5.1.3.2. [3-(4-Methyl-piperazin-1-yl)-propyl]-carbamic acid 3-
(7-chloro-quinolin-4-ylamino)-5-(2-piperidin-1-yl-acetylamino)-
benzyl ester (6). Yellow solid (25 mg, 17% yield); Rf 0.75
(DCM/MeOH/NH4OH 80/20/2); mp ¼ 50 ꢁC; PHPLC 96%,
tR ¼ 13.51 min; 1H NMR (DMSO-d6) d (ppm): 1.37e1.39
(m, 2H, CH2), 1.49e1.56 (m, 6H, CH2), 2.11 (s, 3H, CH3),
2.20e2.27 (m, 10H, CH2), 2.43e2.45 (m, 4H, CH2), 3.06
(s, 2H, NCH2CO), 3.47e3.48 (m, 2H, CH2NHCO), 4.98 (s,
2H, ArCH2), 6.95 (d, J ¼ 5.3 Hz, 1H, Quin-H3),
7.02 þ 7.32 þ 7.71 (s, 3H, Arom-H), 7.25 (t, J ¼ 5.7 Hz, 1H,
OCONH), 7.56 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H, Quin-H6),
7.88 (d, J ¼ 2.2 Hz, 1H, Quin-H8), 8.41 (d, J ¼ 9.1 Hz, 1H,
5.1.2.2. (2-Morpholin-4-yl-ethyl)-carbamic acid 3-(7-chloro-
quinolin-4-ylamino)-5-(2-piperidin-1-yl-acetylamino)-benzyl
ester (4). Yellow solid (80 mg, 29% yield); Rf 0.50 (DCM/
1
MeOH 85/15); mp ¼ 46 ꢁC; PHPLC 95%, tR ¼ 11.48 min; H
NMR (DMSO-d6) d (ppm): 1.37e1.39 (m, 2H, CH2),

S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
2053
Quin-H5), 8.47 (d, J ¼ 5.3 Hz, 1H, Quin-H2), 9.14 (s, 1H, NH),
9.76 (s, 1H, CONH); 13C NMR (DMSO-d6) d (ppm): 24.39
(CH2), 26.32 þ 27.47 (CH2), 46.40 (CH3), 53.33 þ 55.46
(CH2), 54.89 (CH2), 63.50 (CO CH2NH), 65.70 (AreCH2),
103.07 (Quin-C3), 113.26 þ 114.82 þ 117.06 (Arom-CH),
125.41 (Quin-C5), 125.88 (Quin-C6), 128.54 (Quin-C8),
152.78 (Quin-C2); m/z 607.4 (Mþ þ 1).
solid (20 mg, 11% yield); Rf 0.10 (DCM/MeOH 80/20);
mp ¼ 157 ꢁC; PHPLC 100%, tR ¼ 11.35 min; 1H NMR
(DMSO-d6) d (ppm): 1.55e1.75 (m, 6H, CH2), 3.00 (m, 4H,
CH2), 2.82 (s, 6H, NMe2), 3.13 (s, 2H, COCH2N), 3.23 (m,
2H, CH2), 3.57 (m, 2H, CH2), 6.98 (d, J ¼ 5.4 Hz, 1H,
Quin-H3), 7.60 þ 7.82 þ 7.88 (s, 3H, Arom-H), 7.65 (dd,
J ¼ 9.0 Hz, J ¼ 2.0 Hz, 1H, Quin-H6), 7.95 (d, J ¼ 2.0 Hz,
1H, Quin-H8), 8.46 (d, J ¼ 9.1 Hz, 1H, Quin-H5), 8.54 (d,
J ¼ 5.4 Hz, 1H, Quin-H2), 8.67 (t, J ¼ 5.8 Hz, 1H, CONH),
9.50 (s, 1H, NH), 10.70 (s, 1H, CONH); m/z 509.6 (Mþ þ 1).
5.1.3.3. Benzyl-carbamic acid 3-(7-chloro-quinolin-4-yla-
mino)-5-(2-piperidin-1-yl-acetylamino)-benzyl ester (7). To
a solution of compound 1 (100 mg, 0.24 mmol, 1 eq.) in an an-
hydrous DCM/DMF 10/1 mixture (5.5 mL) was added, at
0 ꢁC, benzylisocyanate (30 mg, 0.24 mmol, 1 eq.). After stir-
ring at room temperature overnight, the reactive medium
was concentrated and the residue purified by TLC (DCM/
MeOH 85/15) to give compound 7 as a yellow solid (15 mg,
11% yield); Rf 0.75 (DCM/MeOH 85/15); PHPLC 97%,
tR ¼ 19.21 min; 1H NMR (DMSO-d6) d (ppm): 1.37e1.39
(m, 2H, CH2), 1.50e1.58 (m, 4H, CH2), 2.42e2.46 (m, 4H,
CH2), 3.06 (s, 2H, COCH2N), 4.19 (d, J ¼ 6.3 Hz, 2H,
NHCH2Ar), 5.02 (s, 2H, ArCH2), 6.96 (d, J ¼ 5.3 Hz, 1H,
Quin-H3), 7.05 þ 7.35 þ 7.72 (s, 3H, Arom-H), 7.18e7.32
(m, 5H, Arom-H), 7.56 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H,
Quin-H6), 7.84 (t, J ¼ 6.0 Hz, 1H, OCONH), 7.89 (d,
J ¼ 2.2 Hz, 1H, Quin-H8), 8.42 (d, J ¼ 9.3 Hz, 1H, Quin-
H5), 8.45 (d, J ¼ 5.5 Hz, 1H, Quin-H2), 9.16 (s, 1H, NH),
9.77 (s, 1H, CONH); m/z 558.8 (Mþ þ 1).
5.1.4.2. 3-(7-Chloro-quinolin-4-ylamino)-N-(3-dimethylamino-
propyl)-5-(2-piperidin-1-yl-acetylamino)-benzamide (10). Yel-
low solid (35 mg, 20% yield); Rf 0.10 (DCM/MeOH 80/20);
mp ¼ 164 ꢁC; PHPLC 100%, tR ¼ 11.42 min; 1H NMR
(DMSO-d6) d (ppm): 1.80e1.95 (m, 8H, CH2), 2.65 (m, 4H,
CH2), 2.77 (s, 6H, NMe2), 3.05e3.15 (m, 4H, CH2), 4.19
(m, 2H, CH2), 6.94 (d, J ¼ 5.5 Hz, 1H, Quin-H3),
7.71 þ 7.94 þ 7.99 (s, 3H, Arom-H), 7.84 (dd, J ¼ 9.0 Hz,
J ¼ 1.8 Hz, 1H, Quin-H6), 8.05 (d, J ¼ 2.1 Hz, 1H, Quin-
H8), 8.61 (d, J ¼ 6.1 Hz, 1H, Quin-H2), 8.72 (d, J ¼ 9.0 Hz,
1H, Quin-H5), 8.74 (t, J ¼ 4.6 Hz, 1H, CONH), 9.42 (s, 1H,
NH), 11.00 (s, 1H, CONH); m/z 523.7 (Mþ þ 1).
5.1.4.3. 3-(7-Chloro-quinolin-4-ylamino)-N-(2-dimethylamino-
ethyl)-5-(2-piperidin-1-yl-acetylamino)-N-methylbenzamide
(11). Yellow solid (20 mg, 12% yield); Rf 0.40 (DCM/MeOH
90/10); mp ¼ 90 ꢁC; PHPLC 98%, tR ¼ 10.88 min; 1H NMR
(DMSO-d6) d (ppm): 1.20e1.25 (m, 2H, CH2), 1.55 (m, 4H,
CH2), 2.60e2.70 (m, 17H, CH2 and CH3), 2.97 (s, 2H,
5.1.3.4. 3-(7-Chloro-quinolin-4-ylamino)-5-(2-piperidin-1-yl-
acetylamino)-benzoic acid (8). To a solution of compound 1
(500 mg, 1.2 mmol, 1 eq.) in 30 mL of acetonitrile was added
a solution of periodic acid (1.07 g, 4.7 mmol, 4 eq.) and cata-
lytic amount of CrO3 in 20 mL of acetonitrile. After stirring
at room temperature for 2 h, the mixture was filtered washed
with acetonitrile and concentrated. The residue was the purified
by TLC (DCM/MeOH 8/2) to give compound 8 as a brown
solid (450 mg, 90% yield); Rf 0.15 (DCM/MeOH 85/15);
mp > 250 ꢁC; PHPLC 100%, tR ¼ 12.14 min; 1H NMR
(DMSO-d6) d (ppm): 1.40e1.70 m, 6H, CH2), 2.20e2.70 (m,
4H, CH2), 2.98 (s, 2H, COCH2N), 7.05 (d, J ¼ 5.6 Hz, 1H,
Quin-H3), 7.64 (m, 1H, Quin-H6), 7.65 þ 7.91 þ 7.97 (s,
3H, Arom-H), 7.93 (d, J ¼ 3.6 Hz, 1H, Quin-H8), 8.40 (d,
J ¼ 9.1 Hz, 1H, Quin-H5), 8.54 (d, J ¼ 5.6 Hz, 1H,
Quin-H2), 9.58 (s, 1H, NH), 10.66 (s, 1H, COOH); m/z 439.5
(Mþ þ 1).
COCH2N),
6.99
(d,
J ¼ 5.3 Hz,
1H,
Quin-H3),
7.09 þ 7.46 þ 7.78 (s, 3H, Arom-H), 7.58 (dd, J ¼ 9.0 Hz,
J ¼ 2.2 Hz, 1H, Quin-H6), 7.90 (d, J ¼ 2.2 Hz, 1H, Quin-
H8), 8.41 (d, J ¼ 9.1 Hz, 1H, Quin-H5), 8.50 (d, J ¼ 5.3 Hz,
1H, Quin-H2), 9.26 (s, 1H, NH), 10.04 (s, 1H, CONH); m/z
523.4 (Mþ þ 1).
5.1.4.4. 3-(7-Chloro-quinolin-4-ylamino)-N-(2-pyrrolidin-1-yl-
ethyl)-5-(2-piperidin-1-yl-acetylamino)-benzamide (12). Yel-
low solid (30 mg, 16% yield); Rf 0.35 (DCM/MeOH 80/20);
mp ¼ 88 ꢁC; PHPLC 100%, tR ¼ 11.71 min; 1H NMR
(DMSO-d6) d (ppm): 1.35e1.40 (m, 2H, CH2), 1.50e1.55
(m, 4H, CH2), 1.80e1.85 (m, 4H, CH2), 2.85e3.00 (m,
10H, CH2), 3.10 (s, 2H, COCH2N), 3.10e3.15 (m, 2H,
CH2), 6.98 (d, J ¼ 5.3 Hz, 1H, Quin-H3), 7.52 þ 7.78 þ 7.91
(s, 3H, Arom-H), 7.56 (dd, J ¼ 9.0 Hz, J ¼ 2.3 Hz, 1H,
Quin-H6), 7.89 (d, J ¼ 2.2 Hz, 1H, Quin-H8), 8.42 (d,
J ¼ 9.0 Hz, 1H, Quin-H5), 8.49 (d, J ¼ 5.3 Hz, 1H, Quin-
H2), 8.58 (t, J ¼ 5.2 Hz, 1H, CONH), 9.25 (s, 1H, NH), 9.90
(s, 1H, CONH); m/z 535.2 (Mþ þ 1).
5.1.4. General procedure for access to amides 9e18
To a solution of compound 8 (150 mg, 0.34 mmol, 1 eq.) in
a 5.5 mL of a DCM/DMF 10/1 mixture were added PyBroP
(240 mg, 0.51 mmol 1.5 eq.), the desired amine (0.51 mmol,
1.5 eq.) and DIEA (60 mL, 0.34 mmol, 1 eq.). After stirring
at room temperature overnight, the mixture was concentrated
then the residue was purified by TLC (DCM/MeOH/NH4OH
90/10/0 to 80/20/0.5) to give the desired compound.
5.1.4.5. 3-(7-Chloro-quinolin-4-ylamino)-N-(2-morpholin-1-yl-
ethyl)-5-(2-piperidin-1-yl-acetylamino)-benzamide (13). Yel-
low solid (30 mg, 16% yield); Rf 0.55 (DCM/MeOH 80/20);
mp ¼ 85 ꢁC; PHPLC 96%, tR ¼ 11.28 min; 1H NMR (DMSO-d6)
d (ppm): 1.40e1.45 (m, 2H, CH2), 1.60 (m, 4H, CH2),
2.45e2.50 (m, 4H, CH2), 2.67 (s, 2H, COCH2N), 3.12 (t,
5.1.4.1. 3-(7-Chloro-quinolin-4-ylamino)-N-(2-dimethylamino-
ethyl)-5-(2-piperidin-1-yl-acetylamino)-benzamide (9). Yellow

2054
S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
J ¼ 7.2 Hz, 2H, CH2), 3.55e3.60 (m, 10H, CH2), 6.98 (d,
J ¼ 5.4 Hz, 1H, Quin-H3), 7.51 þ 7.72 þ 7.91 (s, 3H, Arom-H),
7.56 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H, Quin-H6), 7.90 (d, J ¼
2.2 Hz, 1H, Quin-H8), 8.41 (t, J ¼ 9.6 Hz, 1H, CONH), 8.42 (d,
J ¼ 9.0 Hz, 1H, Quin-H5), 8.49 (d, J ¼ 5.4 Hz, 1H, Quin-H2),
9.29 (s, 1H, NH), 10.11 (s, 1H, CONH); m/z 550.5 (Mþ).
5.1.4.10. N-[3-(7-Chloro-quinolin-4-ylamino)-5-(N-phenylpi-
perazin-1-ylcarbonyl)-phenyl]-2-piperidin-1-yl-acetamide
(18). Yellow solid (135 mg, 67% yield); Rf 0.55 (DCM/MeOH
90/10); PHPLC 94%, tR ¼ 12.40 min; 1H NMR (DMSO-d6)
d (ppm): 1.20e1.25 (m, 2H, CH2), 1.55e1.60 (m, 4H, CH2),
2.40e2.50 (m, 12H, CH2), 3.05 (s, 2H, COCH2N), 3.52 (s,
2H, CH2), 7.02 (d, J ¼ 5.5 Hz, 1H, Quin-H3),
7.05 þ 7.37 þ 7.80 (s, 3H, Arom-H), 7.25e7.35 (m, 5H,
Arom-H), 7.60 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H, Quin-H6),
7.92 (d, J ¼ 2.2 Hz, 1H, Quin-H8), 8.40 (d, J ¼ 9.1 Hz, 1H,
Quin-H5), 8.52 (d, J ¼ 5.3 Hz, 1H, Quin-H2), 9.29 (s, 1H,
NH), 10.16 (s, 1H, CONH); m/z 597.3 (Mþ þ 1).
5.1.4.6. N-[3-(7-Chloro-quinolin-4-ylamino)-5-piperidin-1-yl-
carbonyl-phenyl]-2-piperidin-1-yl-acetamide
(14). Yellow
solid (105 mg, 61% yield); Rf 0.50 (DCM/MeOH 90/10);
1
PHPLC 100%, tR ¼ 13.97 min; H NMR (DMSO-d6) d (ppm):
1.20e1.25 (m, 4H, CH2), 1.45e1.50 (m, 8H, CH2), 2.45e
2.50 (m, 8H, CH2), 3.06 (s, 2H, COCH2N),
7.01 þ 7.36 þ 7.76 (s, 3H, Arom-H), 7.03 (d, J ¼ 5.3 Hz, 1H,
Quin-H3), 7.60 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H, Quin-H6),
7.92 (d, J ¼ 2.2 Hz, 1H, Quin-H8), 8.42 (d, J ¼ 9.1 Hz, 1H,
Quin-H5), 8.52 (d, J ¼ 5.4 Hz, 1H, Quin-H2), 9.38 (s, 1H,
NH), 10.43 (s, 1H, CONH); m/z 505.4 (Mþ).
5.2. Biological evaluation
5.2.1. In vitro P. falciparum culture and drug assays
P. falciparum strains were maintained continuously in cul-
ture on human erythrocytes as described by Trager and Jensen
[20]. In vitro antiplasmodial activity was determined using
a modification of the semi-automated microdilution technique
of Desjardins [21]. P. falciparum CQ-sensitive (THAI/Thai-
land) and CQ-resistant (FcB1R/Colombia and K1/Thailand)
strains were used in sensitivity testing. Stock solutions of chlo-
roquine diphosphate and test compounds were prepared in ster-
ile distilled water and DMSO, respectively. Drug solutions were
serially diluted with culture medium and introduced to asyn-
chronous parasite cultures (1% parasitaemia and 1% final he-
matocrit) on 96-well plates for 24 h at 37 ꢁC prior to the
addition of 0.5 mCi of [3H]hypoxanthine (1e5 Ci/mmol; Amer-
sham, Les Ulis, France) per well, for 24 h. After freezing and
thawing, the cells were harvested from each well onto glass fi-
ber filters and the dried filters were counted in a scintillation
spectrometer. The growth inhibition for each drug concentra-
tion was determined by comparison of the radioactivity incor-
porated into the parasite nucleic acids with that in the control
culture (without drug) maintained on the same plate. The con-
centration causing 50% inhibition (IC50) was obtained from
the drug concentrationeresponse curve and the results were ex-
pressed as the mean ꢀ the standard deviations determined from
several independent experiments. The DMSO concentration
never exceeded 0.1% and did not inhibit the parasite growth.
5.1.4.7. N-[3-(7-Chloro-quinolin-4-ylamino)-5-morpholin-1-
ylcarbonyl-phenyl]-2-piperidin-1-yl-acetamide (15). Yellow
solid (85 mg, 50% yield); Rf 0.45 (DCM/MeOH 90/10); PHPLC
1
96%, tR ¼ 12.19 min; H NMR (DMSO-d6) d (ppm): 1.40 (m,
2H, CH2), 1.55e1.60 (m, 4H, CH2), 2.45e2.55 (m, 4H, CH2),
3.06 (s, 2H, COCH2N), 3.10e3.60 (m, 8H, CH2), 7.01 (d,
J ¼ 5.2 Hz, 1H, Quin-H3), 7.03 þ 7.40 þ 7.83 (s, 3H, Arom-
H), 7.58 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H, Quin-H6), 7.91 (d,
J ¼ 2.2 Hz, 1H, Quin-H8), 8.40 (d, J ¼ 9.1 Hz, 1H, Quin-
H5), 8.52 (d, J ¼ 5.3 Hz, 1H, Quin-H2), 9.22 (s, 1H, NH),
9.95 (s, 1H, CONH); m/z 508.2 (Mþ þ 1).
5.1.4.8.
N-[3-(7-Chloro-quinolin-4-ylamino)-5-(N-methylpi-
perazin-1-ylcarbonyl)-phenyl]-2-piperidin-1-yl-acetamide
(16). Yellow solid (55 mg, 32% yield); Rf 0.30 (DCM/MeOH
1
90/10); PHPLC 100%, tR ¼ 10.40 min; H NMR (DMSO-d6)
d (ppm): 1.40 (m, 2H, CH2), 1.55e1.60 (m, 4H, CH2), 2.22
(s, 3H, NCH3), 2.35e2.55 (m, 8H, CH2), 3.06 (s, 2H,
COCH2N), 3.50e3.55 (m, 4H, CH2), 7.00 þ 7.40 þ 7.82 (s,
3H, Arom-H), 7.04 (d, J ¼ 5.3 Hz, 1H, Quin-H3), 7.58 (dd,
J ¼ 9.1 Hz, J ¼ 2.2 Hz, 1H, Quin-H6), 7.91 (d, J ¼ 2.2 Hz,
1H, Quin-H8), 8.40 (d, J ¼ 9.1 Hz, 1H, Quin-H5), 8.52 (d,
J ¼ 5.3 Hz, 1H, Quin-H2), 9.22 (s, 1H, NH), 9.96 (s, 1H,
CONH); m/z 521.3 (Mþ þ 1).
5.2.2. Cytotoxicity test upon MRC-5 cells
5.1.4.9. N-[3-(7-Chloro-quinolin-4-ylamino)-5-(N-phenylpi-
perazin-1-ylcarbonyl)-phenyl]-2-piperidin-1-yl-acetamide
(17). Yellow solid (150 mg, 75% yield); Rf 0.50 (DCM/MeOH
90/10); PHPLC 95%, tR ¼ 15.73 min; 1H NMR (DMSO-d6)
d (ppm): 1.20e1.25 (m, 2H, CH2), 1.55e1.60 (m, 4H, CH2),
2.40e2.50 (m, 8H, CH2), 3.02 (s, 2H, COCH2N), 6.81 (t,
J ¼ 7.3 Hz, 1H, Arom-H), 6.96 (d, J ¼ 7.9 Hz, 2H, Arom-H),
7.07 (d, J ¼ 5.6 Hz, 1H, Quin-H3), 7.09 þ 7.42 þ 7.83 (s,
3H, Arom-H), 7.22 (dd, J ¼ 8.2 Hz, J ¼ 7.4 Hz, 2H, Arom-
H), 7.60 (dd, J ¼ 9.0 Hz, J ¼ 2.2 Hz, 1H, Quin-H6), 7.92 (d,
J ¼ 2.2 Hz, 1H, Quin-H8), 8.42 (d, J ¼ 9.1 Hz, 1H, Quin-
H5), 8.54 (d, J ¼ 5.3 Hz, 1H, Quin-H2), 9.31 (s, 1H, NH),
10.11 (s, 1H, CONH); m/z 582.7 (Mþ).
A human diploid embryonic lung cell line (MRC-5, Bio-
Whittaker 72211D) was used to assess the cytotoxic effects to-
wards host cells. MRC-5 cells were seeded at 5000 cells per
well in 96-well plates. After 24 h, the cells were washed and
two-fold dilutions of the drug were added in 200 mL standard
culture medium (RPMI þ 5% FCS). The final DMSO concen-
tration in the culture remained below 0.5%. The cultures were
incubated with several concentrations of compounds (between
32 mM and 1.6 mM) at 37 ꢁC in 5% CO2e95% air for 7 days.
Untreated cultures were included as controls. The cytotoxicity
was determined using the colorimetric MTT assay [22] and
scored as a % reduction of absorption at 540 nm of treated cul-
tures versus untreated control cultures.

S. Delarue-Cochin et al. / European Journal of Medicinal Chemistry 43 (2008) 2045e2055
2055
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We express our thanks to Gerard Montagne for NMR ex-
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This work was supported by CNRS, Universite de Lille II
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