Tandem synthesis and in vitro antiplasmodial evaluation of new naphtho[2,1-d]thiazole derivatives

Article abstract of DOI:10.1016/j.ejmech.2012.07.034

A series of naphtho[2,1-d]thiazoles was prepared in good yields under microwave irradiation with an original protocol combining tandem direct arylation and intramolecular Knoevenagel reaction on 1,3-thiazole derivatives. Antiplasmodial evaluation of this

Full text of DOI:10.1016/j.ejmech.2012.07.034

European Journal of Medicinal Chemistry 55 (2012) 315e324
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Tandem synthesis and in vitro antiplasmodial evaluation of new naphtho[2,1-d]
thiazole derivatives
,
Anita Cohen a b, Pierre Verhaeghe a, Maxime D. Crozet a, Sébastien Hutter b, Pascal Rathelot a,
,
b,
**
Patrice Vanelle a , Nadine Azas
*
a Laboratoire de Pharmaco-Chimie Radicalaire, LPCR, CNRS-UMR 7273, Institut de Chimie Radicalaire, Faculté de Pharmacie, Aix-Marseille Univ,
27 Boulevard Jean Moulin e CS30064, 13385 Marseille cedex 05, France
b Infections Parasitaires, Transmission, Pharmacologie et Thérapeutique, UMR MD3, Faculté de Pharmacie, Aix-Marseille Univ, 27 Boulevard Jean Moulin e CS30064,
13385 Marseille cedex 05, France
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of naphtho[2,1-d]thiazoles was prepared in good yields under microwave irradiation with an
original protocol combining tandem direct arylation and intramolecular Knoevenagel reaction on 1,3-
thiazole derivatives. Antiplasmodial evaluation of this series highlighted two hit compounds
(compounds 11 and 13) displaying promising in vitro activity on the multiresistant K1 Plasmodium fal-
ciparum strain. Structure-toxicity and structureeactivity relationships are also discussed and reveal the
importance of the R1 and R4 substituents of the naphthyl moiety for the biological profile of the series.
Ó 2012 Elsevier Masson SAS. All rights reserved.
Received 26 April 2012
Received in revised form
12 July 2012
Accepted 18 July 2012
Available online 3 August 2012
Keywords:
Direct arylation
Knoevenagel reaction
Naphtho[2,1-d]thiazoles
In vitro cytotoxicity
In vitro antiplasmodial activity
Plasmodium falciparum
1. Introduction
Nevertheless, the growing drug resistance of parasites around the
world [3] remains a real and ever-present danger, attributable
Malaria is a devastating disease which in 2010 affected 216
million people worldwide and caused 655,000 deaths. This infec-
tion, transmitted via the bite of the female Anopheles mosquito, is
caused by five species of protozoa parasites belonging to the Plas-
modium genus, namely malariae, vivax, ovale, knowlesi and falcipa-
rum. Plasmodium falciparum is the most virulent [1], causing more
than 95% of malaria-related morbidity and mortality. According to
the WHO [2], significant and durable progress has been recorded
for a few years, with the estimated incidence of malaria globally
reduced by 17% since 2000. Moreover, malaria-specific mortality
rates were also reduced by 26% between 2000 and 2010. These
encouraging statistics are the combined result of vector control
(insecticide-treated mosquito nets, indoor residual spraying),
chemoprevention, diagnostic testing and malaria treatment.
mainly to P. falciparum. Currently, the only fully effective class of
antimalarial drug is artemisinin and its derivatives: artemisinin-
based combination therapies (ACTs) have now been adopted as
the first line of treatment in endemic areas. However, a P. falciparum
resistance to artemisinin derivatives was identified and confirmed
on the Cambodia-Thailand border in 2009 [4,5]. This emerging
resistance could lead to a resurgence of more virulent levels of
malaria unless a new chemical class of effective drugs is rapidly
found. In this context, all innovative practices and encouraging
results are being sponsored and shared [6,7], so as to accelerate the
development and licensing of new antimalarial drugs.
1,3-thiazole nucleus is commonly found in the chemical struc-
ture of antiparasitic agents (nitazoxanide [8], aminitrozole [9],
tenonitrozole). Among the chemical compounds already screened
for inhibitors of P. falciparum, certain 1,3-thiazole derivatives have
been reported to display promising antiplasmodial activity with
new target hypothesis [6]. Recently, 1,3-thiazole derivatives
combining good in vitro activity against P. falciparum with oral
efficacy in a Plasmodium berghei mouse model have also been
identified [10]. In continuation of our research program centered on
* Corresponding author. Tel.: þ33 4 91835573; fax: þ33 4 86136822.
** Corresponding author. Tel.: þ33 4 91835564; fax: þ33 4 91835537.
E-mail addresses: patrice.vanelle@univ-amu.fr (P. Vanelle), nadine.azas@univ-
amu.fr (N. Azas).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
http://dx.doi.org/10.1016/j.ejmech.2012.07.034

316
A. Cohen et al. / European Journal of Medicinal Chemistry 55 (2012) 315e324
the design and synthesis of novel bioactive compounds [11e14],
a strong collaboration between our two research teams enabled us
to synthesize and evaluate a group of new heterocyclic hit
compounds displaying antiparasitic activities [15e19], which
included newly-identified original antiplasmodial derivatives
[20e24]. In this context, we decided to explore the antiplasmodial
potential of new naphtho[2,1-d]thiazole derivatives which we
synthesized using an original “one pot” methodology combining
direct arylation and unexpected Knoevenagel intramolecular
reaction on 1,3-thiazole derivatives substituted on the C-2 and C-4
positions.
2. Results and discussion
2.1. Chemistry
Recently, we developed the application of the Suzuki-Miyaura
cross-coupling reaction on thiazole series substituted on the C-2
and C-4 position [25]. We report herein the study of palladium-
catalyzed direct arylation [26e33] on the thiazole ring C-5 posi-
tion. This reaction was first described in 1992 by Ohta et al. [34]. In
the literature, this procedure was described as being less efficient
but complementary to the Suzuki-Miyaura cross-coupling reaction
[35]. However, unexpectedly, when we tried for the first time to
carry out this procedure on C-2 and C-4 substituted thiazole
derivatives, we observed both the direct arylation of the compound
and a simultaneous intramolecular Knoevenagel reaction. We then
investigated the potential of this unexpected “one pot” strategy.
Initially, the required starting material for the direct arylation is 2-
methyl-4-(tosylmethyl)thiazole 1 which was prepared in 62%
overall yield by sequential condensation between 1,3-
dichloroacetone with thioacetamide [36], cyclization using ZnCl2
in methanol [37], and condensation with p-toluenesulfinic acid
sodium salt, in water and under microwave irradiation (Scheme 1).
Then, the reaction of 2-methyl-4-(tosylmethyl)thiazole 1 and 2-
bromobenzaldehyde was carried out in order to synthesize 2-[2-
methyl-4-(tosylmethyl)thiazol-5-yl]benzaldehyde as a product of
direct arylation on C-5 position of the thiazole ring. Surprisingly, 2-
methyl-4-tosylnaphtho[2,1-d]thiazole 4 was obtained, which was
confirmed by X-ray structure analysis (Fig. 1).
Fig. 1. ORTEP plot of 2-methyl-4-tosylnaphtho[2,1-d]thiazole 4.
intramolecular Knoevenagel reaction. The use of “one pot” organic
reactions is currently attracting a great deal of attention, as they are
performed without needing to isolate any intermediate product. In
addition to their advantageous rate of synthesis, they offer the
possibility of building up more complex compounds from simple
components. These procedures are increasingly being applied to
arylbromide reagents in non-heterocyclic series [43], to extend
their reactivity. To verify the validity of this mechanism, we carried
out the reaction of 2-methyl-4-(tosylmethyl)thiazole both with 3-
bromobenzaldehyde and with 4-bromobenzaldehyde. No reaction
occurred under any conditions. These initial results thus revealed
the role of the carbonyl and the importance of its position in the
mechanism of this reaction. Initially, we used the experimental
conditions recommended by Primas et al. [35]. 1 equivalent (equiv)
of 2-bromobenzaldehyde and 5 equiv of starting material 1 were
dissolved in dimethylacetamide (DMA) and heated at 150 ꢀC using
Pd(PPh3)4 (0.05 equiv) as a catalyst and KOAc (3 equiv) as a base for
24 (Table 1, entry 1). The unexpected naphtho[2,1-d]thiazole 4 was
then obtained with a 37% yield and no intermediate was isolated. In
order to optimize the above method, we performed the reaction
under microwave irradiation [44,45]: the desired product 4 was
obtained with a better yield and a shorter reaction time (Table 1,
entry 2). Then, we also tried to reduce the thiazole derivative
The direct arylation of the thiazole nucleus on C-5 position by
the bromide reagent forming the expected 2-[2-methyl-4-(tosyl-
methyl)thiazol-5-yl]benzaldehyde and the very favorable spatial
position of the carbonyl substituent may allow a simultaneous
intramolecular Knoevenagel [38] ring closure on the supposed
methylene-activated intermediate (Scheme 2). Some examples of
“one pot” strategies involving direct arylation have recently been
published [39e42], but none describes a sequence with an
amount
(Table
1,
entries
3e5).
Finally,
the
best
experimental conditions used 1.5 equiv of 1, 3 equiv of KOAc,
0.05 equiv of Pd(PPh3)4 and microwave irradiation (300 W, 150 ꢀC)
(Table 1, entry 4).
Based on these experimental results, we decided to diversify the
chemical substituents on the naphtho[2,1-d]thiazole nucleus in
Cl
CH3OH, ZnCl2
reflux, 12 h
CH3
N
Acetone
H3C NH2
S
Cl
S
N
H
, HCl
+
Cl
Cl
CH3
rt, 5 h
S
O
O
95%
90%
R
+
-
Na
SO2
O2S
Cl
R
N
N
CH3
CH3
S
S
MW 300 W,
0.25 h, 100 °C
1 R = CH3 (72%)
2 R = H (77%)
3 R = Cl (61%)
Scheme 1. Preparation of 4-arylsulfonylmethyl-1-methylthiazoles.

A. Cohen et al. / European Journal of Medicinal Chemistry 55 (2012) 315e324
317
H3C
experiments, employing the MTT method [46] to determine the
cytotoxic concentrations of 50% (CC50) on the HepG2 cell line and
using doxorubicin as a cytotoxic reference-compound (Table 3).
HepG2 is a commonly used human-derived hepatocarcinoma cell
line that has shown characteristics similar to those of primary
hepatocytes. These cells express many of the hepatocyte-specific
metabolic enzymes, thus enabling the cytotoxicity of tested
product metabolites to be evaluated. The tests showed that
a hydrogen atom as R1 substituent conferred a lower overall cyto-
H3C
SO2
SO2
N
S
Br
Pd(PPh )
3 4
N
CH3
+
O
KOAc
MW 300 W, 150 °C
DMA
CH3
S
1
4
H3C
SO2
O
N
S
toxicity (11.3
m
M ꢁ CC50 ꢁ 500
CC50 250
M ꢁ CC50 ꢁ 31.2 M). Furthermore, whatever the R1
m
M) than a methyl group
CH3
(0.2
(0.5
m
m
M
ꢁ
ꢁ
m
M) or chlorine atom
a
m
substituent, compounds bearing an electron-donating group (CH3,
OCH3) as R4 substituent were non-cytotoxic
(31.2 M) compared with doxorubicin used as
Scheme 2. Preparation of 2-methyl-4-tosylnaphtho[2,1-d]thiazole
direct arylation and intramolecular Knoevenagel reaction with 2-methyl-4-(tosyl-
methyl)thiazole 1.
4 by “one pot”
mM ꢁ CC50 ꢁ 250
m
reference drug (Table 3, entries 2, 4, 8, 10 and 15).
Then, the non-cytotoxic compounds were assessed for their
in vitro antiplasmodial activity against a K1 multi-resistant strain
of P. falciparum by using the SYBR Green I fluorescence-based
method [47,48]. In order to identify compounds with significant
potential, two reference drug compounds, chloroquine and doxy-
cycline, were also tested in the same conditions (Table 3, entries 19
and 20). Their inhibitory concentrations 50% (IC50) were then
calculated, as well as their selectivity indexes (SI). Two hit
order to evaluate their influence on the antiplasmodial activity of
the corresponding compounds. Then, we changed both the starting
material and the brominated reagent (Table 2): the same procedure
used to prepare 1 led to 2-methyl-4-(phenylsulfonylmethyl)thia-
zole 2 and 4-[(4-chlorophenylsulfonyl)methyl]-2-methylthiazole 3
from benzenesulfinic acid and p-chlorobenzenesulfinic acid
sodium salts respectively, with 77% and 61% yields (Scheme 1). In
addition, 2-bromobenzaldehyde was replaced by various
substituted benzaldehyde and benzonitrile derivatives in order to
pharmacomodulate the naphtho[2,1-d]thiazole scaffold. The
productive “one pot” strategy was then extended to the new
starting materials 2 and 3 (Table 2, entries 8e18), with a view to
exploring the chemical and biological influence of the electron-
withdrawing or -donating character of the substrates. No correla-
tion could be established with chemical reaction yields. However,
the nature of the halogen substituent on the benzaldehyde reagent
had an influence on the synthesized product rate: the good yield
obtained with 2-bromobenzaldehyde in optimal conditions (64% in
0.5 h) was not achieved with the other 2-halobenzaldehydes tested
(from 35% to 42% with longer reaction times) (Table 2, entries 1 and
2). Moreover, this process was also successfully applied to 2-
bromobenzonitriles (Table 2, entries 6, 7, 12, 13, 17 and 18) and
led to the formation of some naphtho[2,1-d]thiazol-5-amines
(compounds 8, 9, 14, 15, 19 and 20) in good yields (from 44% to
70%). The structure of compound 8 was unambiguously confirmed
by X-ray structure analysis (Fig. 2). The other structures were
assigned by analogy and spectral comparison.
compounds were identified, compounds 11 (4.8
(2.7 M), displaying interesting IC50 values intermediate between
those of the reference drugs chloroquine (0.5 M) and doxycycline
(5 M). These two hit compounds carried a hydrogen as R1
mM) and 13
m
m
m
substituent, which confirms that the nature of the R1 substituent
plays a key role in the cytotoxicity and the antiplasmodial activity
of the naphtho[2,1-d]thiazole scaffold. The influence of other
substituents could not be clearly established because of the limited
number of compounds in the series tested. Nevertheless, from the
results obtained, it seems that antiplasmodial activity could be
greatly improved by adding an electron-donating group as R4
substituent. Given the in vitro antiplasmodial activity and the
cytotoxicity of the two hit compounds, their selectivity indexes
(ratio CC50 (HepG2)/IC50 (K1)) were determined to identify their
therapeutic potential. In particular, compound 13 (SI
> 45)
appeared to be an encouraging antiplasmodial compound, in
comparison with doxycycline and chloroquine, which had selec-
tivity indexes of 4 and 60 respectively.
3. Conclusion
We have developed an original, rapid and efficient process to
synthesize new naphtho[2,1-d]thiazole derivatives on C-2 and C-4
substituted thiazole series. These results constitute the first
example of a “one pot” direct arylation and intramolecular Knoe-
venagel ring closure reaction on a heterocyclic substrate. Further
investigations aimed at extending this method to other heterocy-
cles are in progress. Among the synthesized compounds, we
identified two hits (compounds 11 and 13) displaying an encour-
aging in vitro antiplasmodial activity on the K1 multi-resistant P.
falciparum strain, compared with chloroquine and doxycycline used
as reference drugs. Moreover, compound 13 (SI > 45) achieved
a relatively high selectivity index score, compared with chloroquine
and doxycycline (respectively SI60 and 4). Structure-toxicity and
structureeactivity relationships showed that the hydrogen atom as
R1 substituent plays a key role in cytotoxicity and antiplasmodial
activity. The presence of an electron-donating group as R4
substituent appeared to favor both the absence of cytotoxicity and
increased antiplasmodial activity. The next step in our research
program will be to examine the mechanism by which these
compounds display their antiplasmodial activity.
2.2. In vitro antiplasmodial activity
To synthesize compounds of therapeutic interest, it was neces-
sary to first determine whether these compounds were presenting
a toxic profile on human cells, or if they could offer in vitro selective
antiparasitic properties. Consequently, we performed cytotoxicity
Table 1
Reaction of 1 with 2-bromobenzaldehyde under different conditions.
Entry
Compound 1
Microwave irradiation
Time (h)a
Yield 4 (%)
1
2
3
4
5
5 equiv
5 equiv
2.5 equiv
1.5 equiv
1 equiv
None
24
1.5
1
0.5
0.7
37
45
60
64
25
300 W
300 W
300 W
300 W
a
All the reactions were performed using 1 equiv of 2-bromobenzaldehyde,
3 equiv of KOAc and 0.05 equiv of Pd(PPh3)4 in 5 mL of DMA. The temperature being
ramped up from rt to 150 ꢀC, where it was then held for a time t.

318
A. Cohen et al. / European Journal of Medicinal Chemistry 55 (2012) 315e324
Table 2
Microwave-mediated preparation of several naphtho[2,1-d]thiazoles.
R1
R1
SO2
SO2
Y
N
S
CH3
R4
R3
X
N
Pd(PPh3)4
KOAc
MW 300 W, 150 °C
DMA
+
CH3
S
R3
R2
R4
A
B
Entry
1
Reagent A
Reagent B
Product
Compound number
Reaction time (h)a
Yield (%)
35
H3C
SO2
R1 ¼ CH3
X ¼ Cl
4
2
N
CH3
CH3
CH3
S
R
2 HO
R3 ¼ R4 ¼ H
H3C
SO2
2
3
R1 ¼ CH3
X ¼ I
4
5
3
2
42
87
N
S
R2 ¼ CHO
R3 ¼ R4 ¼ H
H3C
SO2
N
S
R1 ¼ CH3
X ¼ Br
R2 ¼ CHO
R3 ¼ H
CH3
R4 ¼ CH3
H3C
SO2
N
S
4
5
R1 ¼ CH3
X ¼ Br
6
7
1
3
41
71
CH3
R2 ¼ CHO
R3 ¼ F
F
R4 ¼ H
H3C
SO2
N
S
CH3
R1 ¼ CH3
X ¼ Br
R2 ¼ CHO
H3C
H3C
O
R3 ¼ R4 ¼ OCH3
O
CH3
SO2
6
7
8
R1 ¼ CH3
R1 ¼ CH3
R1 ¼ H
X ¼ Br
8
9
3.5
59
70
33
H2N
N
S
CH3
R2 ¼ CN
R3 ¼ R4 ¼ H
H3C
SO2
H2N
F
N
S
X ¼ Br
R2 ¼ CN
R3 ¼ F
1.5
CH3
R4 ¼ H
SO2
X ¼ Br
N
10
0.75
CH3
S
R2 ¼ CHO
R3 ¼ R4 ¼ H
SO2
N
S
9
R1 ¼ H
X ¼ Br
CH3
11
1.5
72
R2 ¼ CHO
R3 ¼ H
CH3
R4 ¼ CH3

A. Cohen et al. / European Journal of Medicinal Chemistry 55 (2012) 315e324
319
Table 2 (continued )
Entry
Reagent A
Reagent B
Product
Compound number
Reaction time (h)a
Yield (%)
SO2
N
10
R1 ¼ H
X ¼ Br
12
2
45
CH3
S
R2 ¼ CHO
R3 ¼ F
F
R4 ¼ H
SO
2
N
S
CH
3
11
12
13
R1 ¼ H
R1 ¼ H
R1 ¼ H
X ¼ Br
13
14
15
4
58
48
46
H C
3
O
R2 ¼ CHO
O
R3 ¼ R4 ¼ OCH3
CH
3
SO2
H2N
X ¼ Br
4
N
S
CH3
R2 ¼ CN
R3 ¼ R4 ¼ H
SO2
H2N
F
N
X ¼ Br
1.75
CH3
S
R2 ¼ CN
R3 ¼ F
R4 ¼ H
Cl
SO2
14
15
R1 ¼ Cl
X ¼ Br
16
17
1
1
60
72
N
S
CH3
R2 ¼ CHO
R3 ¼ R4 ¼ H
Cl
SO2
N
S
R1 ¼ Cl
X ¼ Br
CH3
R2 ¼ CHO
R3 ¼ F
F
R4 ¼ H
Cl
SO2
CH3
N
S
CH3
16
17
R1 ¼ Cl
R1 ¼ Cl
R1 ¼ Cl
X ¼ Br
18
19
20
4
54
44
52
H3C
Cl
O
R2 ¼ CHO
O
R3 ¼ R4 ¼ OCH3
SO2
X ¼ Br
2
H2N
N
CH3
S
R2 ¼ CN
R3 ¼ R4 ¼ H
Cl
SO2
H2N
F
N
18
X ¼ Br
2.25
CH3
S
R2 ¼ CN
R3 ¼ F
R4 ¼ H
a
All the reactions were performed using 1.5 equiv of reagent A, 1 equiv of halogenated reagent B, 3 quiv of KOAc and 0.05 equiv of Pd(PPh3)4 in 5 mL of DMA. An initial
microwave irradiation of 300 W was used, the temperature being ramped up from rt to 150 ꢀC, where it was then held for a time t.
4. Experimental section
for 1H and 50 MHz for 13C).1H and 13C NMR shifts (
d) were reported
in parts per million (ppm) with respect to CDCl3 7.26 ppm for 1H
and 77.0 ppm for 13C and DMSO-d6 2.50 for 1H and 39.7 ppm for 13C.
Multiplicities are represented by s (singulet), d (doublet), t (triplet),
q (quartet) and m (multiplet). Coupling constants (J) are in Hertz
(Hz). Elemental analyses were carried out at the Spectropole,
Yields refer to purified products and are not optimized. Melting
points were determined on a Büchi melting point B-540 apparatus
and are uncorrected. Both 1H- and 13C NMR spectra were deter-
mined on a Bruker Avance 200 spectrometer (operating at 200 MHz

320
A. Cohen et al. / European Journal of Medicinal Chemistry 55 (2012) 315e324
washed with water (3 ꢂ 20 mL). The precipitate was purified by
chromatographic column, eluting with EtOAc/chloroform (5/5).
Recrystallization from isopropanol (i-PrOH) gave the compound 1 as
a brown solid. mp 174 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d: 2.41
(s, 3 H), 2.59 (s, 3 H), 4.48 (s, 2 H), 7.13 (s,1 H), 7.27 (d, J ¼ 8.2 Hz, 2 H),
7.60 (d, J ¼ 8.2 Hz, 2 H). 13C NMR (50 MHz, CDCl3)
d: 18.9 (CH3), 21.6
(CH3), 58.2 (CH2), 119.7 (CH), 128.5 (2 CH), 129.6 (2 CH), 135.5 (C),
142.9 (C), 144.7 (C), 165.9 (C). Anal. Calcd for C12H13NO2S2: C, 53.91;
H, 4.90; N, 5.24. Found: C, 54.13; H, 5.01; N, 5.23.
4.2. General procedure and analytical data for compounds 2 and 3
To a mixture of 4-chloromethyl-2-methylthiazole (500 mg,
3.4 mmol) in water (20 mL) was added the desired sulfinic acid
sodium salt (3 equiv). The reaction mixture was irradiated in
a microwave oven (ETHOS Synth Lab station) and reaction was
carried out under microwave irradiation at 150 ꢀC at 300 W for
0.17 h. After being cooled down, the mixture was poured into water
and then extracted with EtOAc (3 ꢂ 15 mL). The organic layers were
dried over anhydrous sodium sulfate and removed under vacuum.
The residue was purified by chromatographic column, eluting with
chloroform/petroleum ether/diethyl ether (7/2/1). Recrystallization
from i-PrOH gave the corresponding required product.
Fig. 2. X-ray structure of 2-methyl-4-tosylnaphtho[2,1-d]thiazol-5-amine 8.
Faculté des Sciences et Techniques de Saint-Jérome. The following
adsorbent was used for column chromatography: silica gel 60
(Merck, 70e230 mesh). Thin-layer chromatography (TLC) was
performed with Merck 60F-254 silica gel (0.25 mm layer thickness)
in an appropriate solvent.
4.2.1. 2-Methyl-4-(phenylsulfonylmethyl)thiazole (2)
Brownsolid; mp 129 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d
: 2.59
(s, 3 H), 4.52 (s, 2 H), 7.16 (s,1 H), 7.44e7.76 (m, 5 H).13C NMR (50 MHz,
CDCl3) : 18.9 (CH3), 58.1 (CH2),119.8 (CH),128.5 (2 CH),128.9 (2 CH),
4.1. 2-Methyl-4-(tosylmethyl)thiazole (1)
d
To
a mixture of 4-chloromethyl-2-methylthiazole (19 g,
133.7 (CH),138.4 (C),142.6 (C),166.1 (C). Anal. Calcd for C11H11NO2S2:
C, 52.15; H, 4.38; N, 5.53. Found: C, 52.23; H, 4.48; N, 5.46.
128.7 mmol) in water (20 mL) was added p-toluenesulfinic acid
sodium salt (3 equiv). The reaction mixture was irradiated in
a microwave oven (ETHOS Synth Lab station) and reaction was
carried out under microwave irradiation at 150 ꢀC at 300 W for
0.25 h. A precipitate appeared and was filtered after cooling, then
4.2.2. 4-[(4-Chlorophenylsulfonyl)methyl]-2-methylthiazole (3)
White solid; mp 111 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
2.61 (s, 3 H), 4.53 (s, 2 H), 7.21 (s, 1 H), 7.46 (d, J ¼ 8.6 Hz, 2 H), 7.66
(d, J ¼ 8.6 Hz, 2 H). 13C NMR (50 MHz, CDCl3)
d: 18.9 (CH3), 58.1
Table 3
(CH2), 120.1 (CH), 129.3 (2 CH), 130.0 (2 CH), 136.8 (C), 140.6 (C),
142.5 (C), 166.3 (C). Anal. Calcd for C11H10ClNO2S2: C, 45.91; H, 3.50;
N, 4.87. Found: C, 46.06; H, 3.51; N, 4.65.
Human cell toxicity and antiplasmodial activity of the studied compounds.
Entry Compound
Human cell
toxicity HepG2 CC50
Antiplasmodial
activity K1 IC50
Selectivity
index (SI)b
a
a
1
2
3
4
5
6
7
8
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
2.2 (ꢃ0.37)
e
e
4.3. General procedure and analytical data for compounds 4e20
>250e
>25f
NDg
e
3.1(ꢃ1.33)
>125e
0.2 (ꢃ0.01)
0.2 (ꢃ0.03)
11.3 (ꢃ3.17)
>125e
e
>25f
In a 100 mL round-bottom flask were placed 2-methyl-4-(phe-
nylsulfonylmethyl)thiazole or 2-methyl-4-(tosylmethyl)thiazole or
4-[(4-chlorophenylsulfonyl)methyl]-2-methylthiazole (1.5 equiv),
2-bromobenzaldehyde or 2-bromobenzonitrile (1 equiv), tetra-
kis(triphenylphosphine) palladium (0.05 equiv), potassium acetate
(3 equiv) and dimethylacetamide (5 mL). The vessel was then
placed in the synthesis multimode microwave oven cavity (ETHOS
Synth Lab station) and reaction was carried out under microwave
irradiation at 150 ꢀC at 300 W for an appropriate time. The disap-
pearance of starting materials was monitored by TLC. After being
cooled down, the mixture was poured into water and then
extracted with EtOAc (5 ꢂ 15 mL). The organic layers were dried
over anhydrous sodium sulfate and removed under vacuum. The
residue was purified by chromatographic column, eluting with
EtOAc/cyclohexane (5/5) for all compounds except 7 (chloroform/
petroleum ether/ethyl ether (6/2/2)) and 8 (chloroform/EtOAc (9/
1)). The residue was then recrystallized from i-PrOH to give the
corresponding required product.
NDg
e
e
e
>25f
e
< 0.5
> 26
< 1
> 45
NDg
NDg
e
4.8 (ꢃ0.04)
9
26.0 (ꢃ0.81)
>25f
10
11
12
13
14
15
16
17
18
19
20
>125e
2.7 (ꢃ0.11)
>500e
>25f
>25f
e
>500e
6.0 (ꢃ2.12)
3.4 (ꢃ0.22)
>31.2e
1.0 (ꢃ0.02)
0.5 (ꢃ0.16)
0.2
e
e
>25f
e
NDg
e
20
e
e
Doxorubicinc
Doxycyclined
e
e
20
5
0.5
4
60
Chloroquined 30
a
Mean of three independent experiments.
b
Selectivity index was calculated according to the following formula
K1 ¼ CC50 (HepG2)/IC50 (K1).
: SI
c
Doxorubicin was used as reference drug compound for human cell toxicity.
d
Doxycycline and chloroquine were used as reference antiplasmodial drug
compounds.
4.3.1. 2-Methyl-4-tosylnaphtho[2,1-d]thiazole (4)
e
Yellow crystals; mp 213 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
Compound could not be tested at higher concentrations because of lack of
solubility in the culture medium.
2.37 (s, 3 H), 2.91 (s, 3 H), 7.27 (d, J ¼ 8.2 Hz, 2 H), 7.56e7.72 (m, 2 H),
f
No antiplasmodial activity observed at the highest-concentration tested
7.89e7.93 (m, 1 H), 8.08e8.12 (m, 1 H), 8.18 (d, J ¼ 8.2 Hz, 2 H), 8.74
product.
No selectivity index could be determined.
(s, 1 H). 13C NMR (50 MHz, CDCl3)
d: 20.5 (CH3), 21.6 (CH3), 125.1
g

A. Cohen et al. / European Journal of Medicinal Chemistry 55 (2012) 315e324
321
(CH), 126.9 (CH), 129.0 (CH), 129.1 (C), 129.2 (2 CH), 129.3 (2 CH),
129.7 (CH), 130.1 (C), 130.6 (CH), 132.7 (C), 135.4 (C), 138.1 (C), 144.1
(C), 145.4 (C), 167.1 (C). Anal. Calcd for C19H15NO2S2: C, 64.56; H,
4.28; N, 3.96. Found: C, 64.81; H, 4.49; N, 4.11. m/z (EI) 354.0617
[M þ Hþ]. C19H15NO2S2 required 354.0617.
129.2 (2 CH), 129.3 (CH), 129.8 (CH), 130.2 (C), 130.7 (CH), 132.4
(C), 133.2 (CH), 135.5 (C), 141.1 (C), 145.4 (C), 167.2 (C). m/z (EI)
340.0461 [M þ Hþ]. C18H13NO2S2 required 340.0460.
4.3.8. 2,8-Dimethyl-4-(phenylsulfonyl)naphtho[2,1-d]thiazole (11)
Yellow needles; mp 245 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
4.3.2. 2,8-Dimethyl-4-tosylnaphtho[2,1-d]thiazole (5)
d: 2.58 (s, 3 H), 2.90 (s, 3 H), 7.42e7.58 (m, 4 H), 7.69 (s, 1 H), 8.00 (d,
Yellow solid; mp 211 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
J 8.2 Hz, 1 H), 8.29 (dd, J ¼ 8.2, 1.7 Hz, 2 H), 8.70 (s, 1 H). 13C NMR
2.38 (s, 3 H), 2.58 (s, 3 H), 2.91 (s, 3 H), 7.27 (d, J ¼ 8.2 Hz, 2 H),
(50 MHz, CDCl3) d: 20.5 (CH3), 22.0 (CH3),124.4 (CH),127.1 (C),128.4
7.42e7.47 (m, 1 H), 7.69 (s, 1 H), 7.99 (d, J ¼ 8.4 Hz, 1 H), 8.17 (d,
(2 CH), 129.0 (CH), 129.1 (3 CH), 130.3 (CH), 130.4 (C), 131.3 (C), 133.1
(CH), 134.7 (C), 140.5 (C), 141.2 (C), 145.5 (C), 167.0 (C). Anal. Calcd
for C19H15NO2S2: C, 64.95; H, 4.44; N, 3.96. Found: C, 64.95; H, 4.44;
N, 3.85.
J ¼ 8.2 Hz, 2 H), 8.69 (s, 1 H). 13C NMR (50 MHz, CDCl3)
d: 20.5 (CH3),
21.6 (CH3), 22.0 (CH3), 124.4 (CH), 127.2 (C), 128.9 (CH), 129.1 (3 CH),
129.2 (2 CH), 130.4 (CH), 131.8 (C), 134.7 (C), 137.0 (C), 138.3 (C),
140.4 (C), 144.0 (C),145.6 (C),166.9 (C). Anal. Calcd for C20H17NO2S2:
C, 65.37; H, 4.66; N, 3.81. Found: C, 65.36; H, 4.74; N, 3.71.
4.3.9. 7-Fluoro-2-methyl-4-(phenylsulfonyl)naphtho[2,1-d]thiazole
(12)
4.3.3. 7-Fluoro-2-methyl-4-tosylnaphtho[2,1-d]thiazole (6)
Yellow solid; mp 312 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
Brown solid; mp 247 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d: 2.39
2.91 (s, 3 H), 7.43e7.60 (m, 4 H), 7.76 (dd, J ¼ 9.1, 2.4 Hz, 1 H),
(s, 3 H), 2.91 (s, 3 H), 7.26e7.30 (m, 2 H), 7.42e7.52 (m, 1 H), 7.74 (dd,
J ¼ 9.1, 2.5 Hz,1 H), 7.90e7.97 (m,1 H), 8.18 (d, J ¼ 8.4 Hz, 2 H), 8.67 (s,
7.91e7.98 (m, 1 H), 8.27e8.31 (m, 2 H), 8.69 (s, 1 H). 13C NMR
(50 MHz, CDCl3)
d: 20.5 (CH3), 114.2 (d, J ¼ 22.0 Hz, CH), 119.8 (d,
1 H).13C NMR (50 MHz, CDCl3)
d: 20.5 (CH3), 21.6 (CH3), 29.7 (C),114.2
J ¼ 25.1 Hz, CH),127.1 (d, J ¼ 1.5 Hz, C),127.5 (d, J ¼ 8.9 Hz, CH),128.2
(d, J ¼ 4.4 Hz, CH), 128.6 (2 CH), 129.3 (2 CH), 130.3 (d, J ¼ 9.2 Hz, C),
133.4 (CH), 133.5 (C), 135.7 (d, J ¼ 1.1 Hz, C), 140.8 (C), 145.0 (d,
J ¼ 1.8 Hz, C), 160.9 (d, J ¼ 247.8 Hz, CeF), 167.0 (C). m/z (EI)
358.0364 [M þ Hþ]. C18H12FNO2S2 required 358.0366.
(d, J ¼ 21.2 Hz, CH), 119.7 (d, J ¼ 25.2 Hz, CH), 127.0 (d, J ¼ 1.2 Hz, C),
127.4 (d, J ¼ 8.9 Hz, CH),128.1 (d, J ¼ 4.6 Hz, CH),129.2 (2 CH),129.4 (2
CH), 130.2 (d, J ¼ 9.2 Hz, C), 133.9 (C), 135.7 (C), 137.8 (C), 144.3 (C),
160.9 (d, J ¼ 247.3 Hz, CeF),166.9 (C). Anal. Calcd for C19H14FNO2S2: C,
61.44; H, 3.80; N, 3.77. Found: C, 61.32; H, 3.91; N, 3.65.
4.3.10. 7,8-Dimethoxy-2-methyl-4-(phenylsulfonyl)naphtho[2,1-d]
4.3.4. 7,8-Dimethoxy-2-methyl-4-tosylnaphtho[2,1-d]thiazole (7)
thiazole (13)
Yellow solid; mp 282 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
Yellow solid; mp 310 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
2.38 (s, 3 H), 2.90 (s, 3 H), 4.04 (s, 3 H), 4.05 (s, 3 H), 7.10 (s, 1 H), 7.27
2.89 (s, 3 H), 4.05 (s, 3 H), 4.06 (s, 3 H), 7.10 (s, 1 H), 7.38 (s, 1 H),
(d, J ¼ 8.2 Hz, 2 H), 7.37 (s, 1 H), 8.17 (d, J ¼ 8.2 Hz, 2 H), 8.58 (s, 1 H).
7.42e7.58 (m, 3 H), 8.29 (dd, J ¼ 8.3, 1.9 Hz, 2 H), 8.60 (s, 1 H). 13C
13C NMR (50 MHz, CDCl3)
d
: 20.6 (CH3), 21.6 (CH3), 56.1 (CH3), 56.3
NMR (50 MHz, CDCl3) d: 20.6 (CH3), 56.1 (CH3), 56.3 (CH3), 104.2
(CH3), 104.2 (CH), 108.9 (CH), 124.5 (C), 126.1 (C), 127.5 (CH), 129.1 (4
CH), 130.5 (C), 134.1 (C), 138.5 (C), 143.9 (C), 144.7 (C), 149.9 (C),
152.2 (C), 165.9 (C). Anal. Calcd for C21H19NO4S2: C, 61.00; H, 4.63;
N, 3.39. Found: C, 60.88; H, 4.70; N, 3.27.
(CH), 108.9 (CH), 124.5 (C), 126.2 (C), 127.7 (CH), 128.5 (2 CH), 129.1
(2 CH), 130.1 (C), 133.0 (CH), 134.1 (C), 141.4 (C), 144.7 (C), 150.0 (C),
152.3 (C),166.0 (C). Anal. Calcd for C20H17NO4S2: C, 60.13; H, 4.29; N,
3.51. Found: C, 60.34; H, 4.38; N, 3.42.
4.3.5. 2-Methyl-4-tosylnaphtho[2,1-d]thiazol-5-amine (8)
4.3.11. 2-Methyl-4-(phenylsulfonyl)naphtho[2,1-d]thiazol-5-amine
(14)
Brown needles; mp 206 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d
: 2.36 (s, 3 H), 2.78 (s, 3 H), 5.57 (br s, 2 H, NH2), 7.23 (d,
Yellow needles; mp 278 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
J ¼ 8.4 Hz, 2 H), 7.46e7.75 (m, 3 H), 7.93e7.97 (m, 1 H), 8.10 (d,
d
: 2.78 (s, 3 H), 4.06 (br s, 2 H, NH2), 7.39e7.67 (m, 5 H), 7.75e7.79
(m, 1 H), 7.96e8.00 (m, 1 H), 8.19e8.23 (m, 2 H). 13C NMR (50 MHz,
CDCl3) : 20.2 (CH3), 108.3 (C), 122.5 (C), 122.9 (CH), 123.0 (C), 125.9
J ¼ 8.4 Hz, 2 H). 13C NMR (50 MHz, CDCl3)
d: 20.2 (CH3), 21.5
(CH3), 108.6 (C), 122.5 (C), 122.9 (CH), 123.0 (C), 125.8 (CH), 125.9
(CH), 128.1 (2 CH), 128.8 (2 CH), 129.7 (CH), 130.1 (C), 140.5 (C),
143.5 (C), 145.2 (C), 146.7 (C), 165.6 (C). Anal. Calcd for
C19H16N2O2S2: C, 61.93; H, 4.38; N, 7.60. Found: C, 61.81; H, 4.41;
N, 7.49.
d
(CH),126.1 (CH), 128.1 (4 CH), 129.9 (CH), 130.3 (C), 132.7 (CH),143.4
(C), 145.4 (C), 146.8 (C), 165.6 (C). Anal. Calcd for C18H14N2O2S2: C,
60.99; H, 3.98; N, 7.90. Found: C, 60.85; H, 4.00; N, 7.80.
4.3.12. 7-Fluoro-2-methyl-4-(phenylsulfonyl)naphtho[2,1-d]
4.3.6. 7-Fluoro-2-methyl-4-tosylnaphtho[2,1-d]thiazol-5-amine (9)
thiazol-5-amine (15)
Brown solid; mp 260 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d
:
Brown solid; mp 295 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
2.37 (s, 3 H), 2.79 (s, 3 H), 4.70 (br s, 2 H, NH2), 7.22e7.26 (m, 2 H),
7.34e7.43 (m, 1 H), 7.61 (dd, J ¼ 10.7, 2.1 Hz, 1 H), 7.72e7.79 (m, 1 H),
2.77 (s, 3 H), 7.37e7.53 (m, 4 H), 7.62 (dd, J ¼ 10.5, 2.5 Hz, 1 H),
7.75e7.82 (m, 1 H), 8.20e8.23 (m, 2 H). NH2 not visible under these
8.10 (d, J ¼ 8.2 Hz, 2 H). 13C NMR (50 MHz, CDCl3)
d: 20.2 (CH3), 21.6
conditions. 13C NMR (50 MHz, CDCl3)
d: 20.2 (CH3), 108.1 (d,
(CH3), 108.1 (d, J ¼ 22.7 Hz, CH), 109.8 (C), 119.2 (d, J ¼ 23.9 Hz, CH),
123.0 (C), 123; 7 (d, J ¼ 7.5 Hz, C), 126.9 (d, J ¼ 1.1 Hz, C), 128.1 (d,
J ¼ 8.4 Hz, CH), 128.3 (2 CH), 128.8 (2 CH), 140.3 (C), 143.7 (C), 144.3
(d, J ¼ 4.1 Hz, C), 146.3 (C), 160.7 (d, J ¼ 247.6 Hz, CeF), 165.4 (C).
Anal. Calcd for C19H15FN2O2S2: C, 59.05; H, 3.91; N, 7.25. Found: C,
59.33; H, 4.10; N, 7.14.
J ¼ 22.7 Hz, CH), 109.4 (C), 115.5 (C), 119.4 (d, J ¼ 24.5 Hz, CH), 123.0
(C), 127.0 (d, J ¼ 1.5 Hz, C), 128.1 (2 CH), 128.2 (2 CH), 128.4 (CH),
132.9 (CH), 143.2 (C), 144.5 (d, J ¼ 4.0 Hz, C), 146.3 (d, J ¼ 1.4 Hz, C),
160.7 (d,
J
¼
247.4 Hz, CeF), 165.5 (C). Anal. Calcd for
C18H13FN2O2S2: C, 58.05; H, 3.52; N, 7.52. Found: C, 58.46; H, 3.74;
N, 7.33.
4.3.7. 2-Methyl-4-(phenylsulfonyl)naphtho[2,1-d]thiazole (10)
4.3.13. 4-(4-Chlorophenylsulfonyl)-2-methylnaphtho[2,1-d]thiazole
Yellow solid; mp 284 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d
:
(16)
2.91 (s, 3 H), 7.43e7.75 (m, 5 H), 7.92e7.96 (m, 1 H), 8.11e8.15 (m,
Brown solid; mp 244 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
1 H), 8.30 (dd, J ¼ 8.2, 1.8 Hz, 2 H), 8.77 (s, 1 H). 13C NMR (50 MHz,
2.92 (s, 3 H), 7.41e7.47 (m, 2 H), 7.60e7.76 (m, 2 H), 7.92e7.96 (m,
CDCl3) d: 20.5 (CH3), 125.1 (CH), 127.0 (CH), 128.5 (2 CH), 129.1 (C),
1 H), 8.10e8.15 (m, 1 H), 8.21e8.26 (m, 2 H), 8.75 (s, 1 H). 13C NMR

322
A. Cohen et al. / European Journal of Medicinal Chemistry 55 (2012) 315e324
(50 MHz, CDCl3)
d
: 20.5 (CH3), 125.2 (CH), 127.1 (CH), 128.8 (2 CH),
4.4.2. Crystal data for compound 8
19H16N2O2S2, yellow prism (0.25 ꢂ 0.25 ꢂ 0.08), MW ¼ 368.46,
129.1 (C), 129.3 (CH), 130.0 (CH), 130.3 (C), 130.7 (CH), 130.8 (2 CH),
131.9 (C), 135.5 (C), 139.5 (C), 139.9 (C), 145.2 (C), 167.4 (C). m/z (EI)
374.0074 [M þ Hþ]. C18H12ClNO2S2 required 374.0071.
C
ꢀ
monoclinic, space group P21/c (T ¼ 293 K), a ¼ 11.4979 (2) A,
ꢀ
ꢀ
a
b ¼ 7.7289 (1) A, c ¼ 19.6273 (6) A,
¼ 90ꢀ,
b
¼ 91.674 (1)ꢀ,
g
¼ 90ꢀ;
3
ꢀ
V ¼ 1661.06 (4) A , Z ¼ 4,
m
¼ 0.327, F(000) ¼ 768, index ranges
4.3.14. 4-(4-Chlorophenylsulfonyl)-7-fluoro-2-methylnaphtho[2,1-
d]thiazole (17)
0 ꢁ h ꢁ 15, 0 ꢁ k ꢁ 9, ꢄ26 ꢁ l ꢁ 25;
q
range ¼ 1.81e28.63ꢀ, 228
variables and 0 restraints, were defined for 3287 independent
Yellow solid; mp 307 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
reflections with
GooF ¼ 1.237.
I
ꢅ
2
s
(I) to R1 ¼ 0.0607, wR2
¼
0.1578,
2.91 (s, 3 H), 7.44e7.54 (m, 3 H), 7.76 (dd, J ¼ 9.1, 2.3 Hz, 1 H),
7.91e7.99 (m, 1 H), 8.24 (d, J ¼ 8.6 Hz, 2 H), 8.68 (s, 1 H). 13C NMR
CCDC 826725 (for compound 4) and CCDC 826726 (for
compound 8) contain the supplementary crystallographic data for
this paper. These data can be obtained free of charge at www.cdcc.
cam.ac.uk/data_request/cif of from the Cambridge Crystallographic
Data Centre, 12, Union Road, Cambridge CB2 1EZ, UK; fax: þ 44
(1223) 336033; email: deposit@ccdc.cam.ac.uk.
(50 MHz, CDCl3)
d
: 20.5 (CH3), 114.3 (d, J ¼ 21.3 Hz, CH), 120.0 (d,
J ¼ 25.3 Hz, CH), 127.1 (d, J ¼ 1.1 Hz, CH), 127.5 (d, J ¼ 8.8 Hz, CH),
128.3 (d, J ¼ 4.7 Hz, C), 128.9 (2 CH), 130.2 (d, J ¼ 9.6 Hz, C), 130.9 (2
CH), 133.1 (C), 135.8 (C), 139.7 (d, J ¼ 45.7 Hz, C), 144.8 (d, J ¼ 1.8 Hz,
C), 161.0 (d, J ¼ 249.6 Hz, CeF), 167.3 (C), 171.3 (C). Anal. Calcd for
C18H11ClFNO2S2: C, 55.17; H, 2.83; N, 3.57. Found: C, 55.42; H, 3.10;
N, 3.39.
4.5. MTT assays
4.3.15. 4-(4-Chlorophenylsulfonyl)-7,8-dimethoxy-2-
The tested compounds’ cytotoxicity on the HepG2 cell line
(purchased from ATCC, ref HB-8065) was evaluated according to
the method of Mosmann with slight modifications. Briefly, cells
methylnaphtho[2,1-d]thiazole (18)
Yellow solid; mp 281 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
2.90 (s, 3 H), 4.05 (s, 3 H), 4.06 (s, 3 H), 7.11 (s, 1 H), 7.38 (s, 1H), 7.44
5 ꢂ 103 in 100
m
L of complete medium [RPMI supplemented with
-glutamine (200 mM) and penicillin
g/mL)] were inoculated into each
well of 96-well plates and incubated at 37 ꢀC in a humidified 6%
CO2, 14% O2, 80% N2 atmosphere. After 24 h incubation, 100 L of
(d, J ¼ 8.6 Hz, 2 H), 8.22 (d, J ¼ 8.6 Hz, 2 H), 8.59 (s, 1 H). 13C NMR
10% fœtal bovine serum, 1% L
(50 MHz, CDCl3)
d
: 20.6 (CH3), 56.2 (CH3), 56.3 (CH3), 104.2 (CH),
(100 U/mL)/streptomycin (100 m
108.9 (CH), 124.4 (C), 126.4 (C), 127.7 (CH), 128.8 (2 CH), 129.7 (C),
130.7 (2 CH), 134.2 (C), 139.7 (C), 139.9 (C), 144.5 (C), 150.1 (C), 152.5
(C), 166.2 (C). m/z (EI) 434.0281 [M þ Hþ]. C20H16ClNO4S2 required
434.0282.
m
medium with various product concentrations was added and the
plates were incubated for 72 h. Duplicate assays were performed
for each sample. At the end of the treatment and incubation, the
4.3.16. 4-(4-Chlorophenylsulfonyl)-2-methylnaphtho[2,1-d]thiazol-
medium was aspirated from the wells and 10
mL MTT solution (5 mg
5-amine (19)
MTT/mL in PBS) was added to each well with 100 mL of medium
Brown solid; mp 194 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d
:
without fœtal bovine serum. Cells were incubated for 2 h at 37 ꢀC to
allow MTT oxidation by mitochondrial dehydrogenase in the viable
cells. After this time, the MTT solution was aspirated and DMSO
2.78 (s, 3 H), 7.41 (d, J ¼ 8.5 Hz, 2 H), 7.47e7.68 (m, 2 H),
7.76e7.80 (m, 1 H), 7.95e7.99 (m, 1 H), 8.16 (d, J ¼ 8.5 Hz, 2 H).
NH2 not visible under these conditions. 13C NMR (50 MHz, CDCl3)
(100
mL) was added to each well to dissolve the resulting blue
d
: 20.2 (CH3), 107.5 (C), 122.4 (C), 123.0 (CH), 126.0 (CH), 126.2
formazan crystals. Plates were shaken vigorously (300 rpm) for
a few minutes. The absorbance was measured at 570 nm with
630 nm as reference wavelength, using a microplate spectropho-
tometer. DMSO was used as blank and doxorubicin as positive
control. Cell viability was calculated as percentage of control (cells
incubated without compound). The 50% cytotoxic concentration
(CC50) was determined by non-linear regression analysis processed
on doseereponse curves, using the Table Curve software 2D v.5.0.
CC50 values represent the mean value calculated from three inde-
pendent experiments.
(CH), 128.4 (2 CH), 128.9 (C), 129.7 (2 CH), 130.1 (CH), 133.0 (C),
139.3 (C), 141.8 (C), 145.6 (C), 146.0 (C), 166.2 (C). Anal. Calcd for
C18H13ClN2O2S2: C, 55.59; H, 3.37; N, 7.20. Found: C, 55.80; H,
3.49; N, 7.02.
4.3.17. 4-(4-Chlorophenylsulfonyl)-7-fluoro-2-methylnaphtho[2,1-
d]thiazol-5-amine (20)
Brown solid; mp 253 ꢀC (i-PrOH) 1H NMR (200 MHz, CDCl3)
d:
2.77 (s, 3 H), 7.35e7.46 (m, 3 H), 7.61(dd, J ¼ 10.6, 2.2 Hz, 1 H),
7.73e7.80 (m, 1 H), 8.15 (d, J ¼ 8.7 Hz, 2 H). NH2 not visible under
these conditions. 13C NMR (50 MHz, CDCl3)
d
: 20.2 (CH3), 108.2 (d,
4.6. In vitro evaluation of antiplasmodial activity
J ¼ 23.0 Hz, CH), 108.8 (C), 119.5 (d, J ¼ 24.2 Hz, CH), 123.0 (d,
J ¼ 1.1 Hz, C), 123.6 (d, J ¼ 7.7 Hz, C), 127.0 (d, J ¼ 1.9 Hz, C), 128.2 (d,
J ¼ 8.4 Hz, CH), 128.4 (2 CH), 129.8 (2 CH), 139.4 (C), 141.6 (C), 144.6
(d, J ¼ 4.4 Hz, C), 146.0 (d, J ¼ 1.9 Hz, C), 160.7 (d, J ¼ 247.4 Hz, C),
165.7 (C). m/z (EI) 407.0088 [M þ Hþ]. C18H12ClFN2O2S2 required
407.0086.
The evaluation of the less cytotoxic compounds’ antiplasmodial
activity was conducted on a K1 in vitro culture-adapted P. falcipa-
rum strain (clone of W2), resistant to chloroquine, pyrimethamine
and proguanil. Cultures were maintained in fresh Aþ human
erythrocytes at 2.5% hematocrit in complete medium (RPMI 1640
with 25 mM HEPES, 25 mM NaHCO3, 10% of Aþ human serum) at
37 ꢀC under reduced O2 atmosphere (gas mixture 6% CO2, 14% O2,
80% N2). Parasitemia was maintained daily between 1% and 6%. The
P. falciparum drug susceptibility test was carried out by comparing
quantities of DNA in treated and control cultures of parasite in
human erythrocytes according to a SYBR Green I fluorescence-
based method using a 96-well fluorescence plate reader. Parasite
culture was synchronized at ring stage with 5% sorbitol.
4.4. X-ray structure determination of compounds 4 and 8
4.4.1. Crystal data for compound 4
C19H15NO2S2, yellow prism (0.2
ꢂ
0.08
ꢂ
0.06 mm3),
MW ¼ 353.44, monoclinic, space group P21/n (T ¼ 293 K),
¼ 90ꢀ,
ꢀ
ꢀ
ꢀ
a
a ¼ 7.3669 (2) A, b ¼ꢀ14.4698 (4) A, c ¼ 15.5892 (6) A,
3
ꢀ
b
¼ 91.674 (1)ꢀ,
g
¼ 90 ; V ¼ 1661.06 (9) A , Z ¼ 4,
m
¼ 0.331 mmꢄ1
,
F(000) ¼ 736, index ranges 0 ꢁ h ꢁ 9, 0 ꢁ k ꢁ 19, ꢄ20 ꢁ l ꢁ 20;
q
Compounds were incubated in a total assay volume of 200 mL
range ¼ 3.1e28.18ꢀ, 219 variables and 0 restraints, were defined for
(RPMI, 2% haematocrit and 1% parasitaemia) for 72 h in a humidi-
fied atmosphere (6% CO2, 14% O2, 80% N2) at 37 ꢀC, in 96-well flat-
bottom plates. Duplicate assays were performed for each sample.
2315 independent reflections with I ꢅ 2
s
(I) to R1 ¼ 0.0698,
wR2 ¼ 0.1238, GooF ¼ 1.117.

A. Cohen et al. / European Journal of Medicinal Chemistry 55 (2012) 315e324
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After incubation, 125
washed twice with 150
transferred to 96-well flat bottom non- sterile black plates (Greiner
Bio-one). 15 L of the SYBR Green lysis buffer (2XSYBR Green,
20 mM Tris base pH 7.5, 20 mM EDTA, 0.008% w/v saponin, 0.08% w/
v Triton X-100) was added to each well. Negative control (treated
by DMSO) and positive controls (doxycycline and chloroquine)
were added to each set of experiments. Plates were incubated for
0.25 h at 37 ꢀC and then read on a TECAN Infinite F-200 spectro-
photometer with excitation and emission wavelengths at 497 and
520 nm respectively. The concentrations of compounds required to
induce a 50% decrease in parasite growth (K1 IC50) were calculated
from three independent experiments.
m
L supernatant was discarded and cells were
m
L 1X PBS. 15 L re-suspended cells were
m
m
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4.7. Selectivity index (SI)
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The selectivity indexes presented correspond to the ratios
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K1 ¼ CC50 (HepG2)/IC50 (K1).
Acknowledgments
[24] C. Castera-Ducros, N. Azas, P. Verhaeghe, S. Hutter, P. Garrigue, A. Dumètre,
L. Mbatchi, M. Laget, V. Rémusat, F. Sifredi, S. Rault, P. Rathelot, P. Vanelle,
Targeting the human malaria parasite Plasmodium falciparum: in vitro iden-
This work was supported by the CNRS and Aix-Marseille
University. The authors thank V. Remusat for 1H and 13C spectra
recording, G. Giuglio-Tonolo for his valuable contribution, M. Giorgi
for X-ray diffraction and structure analysis.
tification of
a
new antiplasmodial hit in 4-phenoxy-2- tri-
chloromethylquinazoline series, Eur. J. Med. Chem. 46 (2011) 4184e4191.
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