Dibenzylideneacetone analogues as novel Plasmodium falciparum inhibitors

Article abstract of DOI:10.1016/j.bmcl.2011.03.037

A series of dibenzylideneacetones (A1-A12) and some of their pyrazolines (B1-B4) were synthesized and evaluated in vitro for blood stage antiplasmodial properties in Plasmodium falciparum culture using SYBR-green-I fluorescence assay. The compound (1E, 4E)-1,5-bis(3,4-dimethoxyphenyl)penta-1,4-dien-3-one (A9) was found to be the most active with IC₅₀ of 1.97 μM against chloroquine-sensitive strain (3D7) and 1.69 μM against chloroquine-resistant field isolate (RKL9). The MTT based cytotoxicity assay on HeLa cell line has confirmed that A9 is selective in its action against malaria parasite (with a therapeutic index of 166). Our results revealed that these compounds exhibited promising antiplasmodial activities which can be further explored as potential leads for the development of cheaper, safe, effective and potent drugs against chloroquine-resistant malarial parasites.

Full text of DOI:10.1016/j.bmcl.2011.03.037

Bioorganic & Medicinal Chemistry Letters 21 (2011) 3034–3036
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Dibenzylideneacetone analogues as novel Plasmodium falciparum inhibitors
Rahul Balasaheb Aher a, , Gajanan Wanare a, Neha Kawathekar a, Ravi Ranjan Kumar b,
,
⇑
Naveen Kumar Kaushik b, Dinkar Sahal b, Virander Singh Chauhan b
a Department of Pharmacy, Shri G.S. Institute of Technology and Science, 23 Park Road, Indore 452003 (MP), India
b Malaria Research Laboratory, International Center for Genetic Engineering and Biotechnology, New Delhi 110067, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of dibenzylideneacetones (A1–A12) and some of their pyrazolines (B1–B4) were synthesized and
evaluated in vitro for blood stage antiplasmodial properties in Plasmodium falciparum culture using SYBR-
green-I fluorescence assay. The compound (1E, 4E)-1,5-bis(3,4-dimethoxyphenyl)penta-1,4-dien-3-one
Received 2 December 2010
Revised 17 February 2011
Accepted 10 March 2011
Available online 12 April 2011
(A9) was found to be the most active with IC50 of 1.97
lM against chloroquine-sensitive strain (3D7)
and 1.69 M against chloroquine-resistant field isolate (RKL9). The MTT based cytotoxicity assay on HeLa
l
cell line has confirmed that A9 is selective in its action against malaria parasite (with a therapeutic index
of 166). Our results revealed that these compounds exhibited promising antiplasmodial activities which
can be further explored as potential leads for the development of cheaper, safe, effective and potent drugs
against chloroquine-resistant malarial parasites.
Keywords:
Antimalarial activity
Dibenzylideneacetone
Pyrazolines
Plasmodium falciparum
SYBR-green-I assay
Ó 2011 Published by Elsevier Ltd.
The emergence and spread of parasites resistant to most of the
clinically used antimalarials and their combinations have aroused
an imperative need to develop new drugs against malaria. The Plas-
modium falciparum malaria resistance remains the worst parasitic
burden in developing countries. The development of resistance
by the parasite against first line as well as second line antimalarial
drugs has drawn attention to develop new drugs to alleviate the
disease burden. Half of the world’s population is at a risk of malar-
ia, and according to the WHO report of 2009, an estimated 243 mil-
lion cases led to nearly 863 thousands deaths in 2008, most of
them being children under 5 years of age.1
Since a vast majority of those affected with malaria are the poor
sections of modern societies, the new drugs crying to come out of
the discovery pipe line must be potent, safe and affordable. To be
widely useful, they must reach developing countries at a cheaper
rate with proper distribution. Moreover, the absence of an effective
vaccine for protection and the availability of artemisinin and its
derivatives as the only option has made the situation rather seri-
ous. Indeed, most recent reports indicate the emergence of clinical
resistance against artemisinin in the form of prolonged parasite
clearance time.2,3 As a result, there is an urgent need for new effi-
cient antimalarial agents.
As shown in Figure 1, these compounds are chalcones with an
additional carbon–carbon double bond, and lacks the moiety con-
taining active methylene (–CH2–) and carbonyl (C@O) groups of
curcumin (indeed the presence of which decreases the stability
of curcumin).6 So, a chemically diversed series of molecules with
probable new mechanism of action, that is, dibenzylideneacetone
derivatives were synthesized and evaluated for their antimalarial
potency against P. falciparum, chloroquine-sensitive (3D7) and
chloroquine-resistant (RKL9) strains. Alterations in dibenzylidene-
acetone were made to study the effect of different substituents
(with varied lipophilic, steric and electronic properties) on antima-
larial potency. Some of the analogs were further converted into
their corresponding pyrazolines to test the activity enhancement
by pyrazoline ring as found in curcumin–pyrazoline derivatives
MP-14
50
(IC
= 0.45 l
M).4
O
R
R
A
O
R
R
B*
The a,b-unsaturated ketones like chalcone, curcumin have been
studied as antimalarial agents.4,5 Here in this study, antimalarial
activity of dibenzylideneacetone derivatives has been investigated.
O
O
O
O
HO
OH
C
⇑
Corresponding author. Tel.: +91 9730962361.
E-mail addresses: rahulba26@yahoo.co.in, rahulba26@gmail.com (R.B. Aher).
Author contributed equally.
Figure. 1. Structures of Chalcone (A), Dibenzylideneacetone (B⁄) and Curcumin
(C)⁄: synthesized analogues.
0960-894X/$ - see front matter Ó 2011 Published by Elsevier Ltd.
doi:10.1016/j.bmcl.2011.03.037

R. B. Aher et al. / Bioorg. Med. Chem. Lett. 21 (2011) 3034–3036
3035
N
N
O
O
CHO
+
(ii)
(i)
R
R
R
R
R
B1-B4
A1-A12
Scheme 1. Synthesis of dibenzylideneacetones (A1–A12) and their pyrazoline derivatives (B1–B4). Reagents and conditions: (i) 10% NaOH, EtOH, H2O; (ii) Phenylhydrazine,
CH3COOH, 8 h reflux.
The compounds A1–A12 were prepared from acetone and
substituted benzaldehydes in a base catalyzed aldol reaction
(Scheme 1) as described by Kiran et al.7 Further, some of the diben-
zylideneacetone derivatives were converted to pyrazolines using a
reported method.4 Synthetic methods used were chosen based on
their simple procedures with better yields of products along with
their cost effectiveness. The purity and structures of all synthesized
compounds were confirmed by TLC and spectroscopic techniques,
respectively. The IR spectra of all the dibenzylideneacetones dis-
played desired characteristic absorption bands in their respective
atives (A5, IC50 = 6.26
were the active ones, while the fluoro derivative (A2,
IC50 = 133.2 M) and the bromo derivative (A4, IC50 = 221.89 M)
lacked significant antimalarial activity. This is in accord with
presence of the chloro group in various active antimalarial mole-
cules including chloroquine, pyronaridine, acridinedione and
others.9–11 It is observed from results that, ortho chloro enhanced
lM, A6, IC50 = 9.94 lM, A3, IC50 = 92.35 lM)
l
l
the antimalarial potency appreciably (IC50 = 6.26
substitution at both ortho and para positions caused a marginal
decrease in potency (A6, IC50 = 9.94 M). There may be effective
lM), but chloro
l
regions like
a
,b-unsaturated ketone (1650–1680 cmÀ1) and trans
interactions of the compound with target protein(s) when chloro
is ortho substituted. The presence of para chloro in A6 may be pre-
venting the effective binding of ortho chloro in active site of target
protein(s). The decrease in potency of para substituted chloro com-
pound (A3) may be due to inappropriate orientation of chloro to fit
in binding site.
C@C (970–990 cmÀ1). All the pyrazolines (B1–B4) displayed their
characteristic absorption bands due to C@N and C–N in regions
1590–1600 and 1310À1330 cmÀ1, respectively. The absence of an
absorption bands of
a )
,b-unsaturated ketone (at 1650–1680 cmÀ1
and N–H stretching in pyrazoline compounds (B1–B4) confirms
the cyclization of pyrazoline ring. The 1H NMR spectra of all deriv-
atives exhibited a characteristic multiplet for aromatic protons and
two CH@CH protons at 6.49–8.05 ppm. The coupling constants
(J = 16 Hz) for the two protons attached to the double bonds con-
firms the stereochemistry of synthesized compounds as E-form.
The remaining protons exhibited the expected d values. The mass
spectra of compounds displayed molecular ions which confirmed
their molecular weights.
The introduction of methoxy group in an effort to increase elec-
tron density, has decreased the potency of unsubstituted derivative
(IC50 = 213.41
IC50 = 254.8 M); while it has improved the same when it is at
ortho position (A8, IC50 = 74.74 M). Interestingly, dual substitu-
tion of methoxy group at meta and para position leads to a dra-
matic increase in the potency (A9, IC50 = 1.97 M). This may be
lM) when methoxy is at para position (A7,
l
l
l
due to the favourable steric interaction of these groups.
The synthesized compounds were screened for in vitro
antimalarial activity against chloroquine-sensitive 3D7 strain and
chloroquine-resistant field isolate RKL9 of P. falciparum using
SYBR-green-I fluorescence assay.8 Chloroquine diphosphate and
artemisinin were used as positive controls. The results are shown
in Table 1. Screening against P. falciparum 3D7 strain revealed that
the lead unsubstituted dibenzylideneacetone (A1) lacks significant
potency (IC50 of 213.41 lM) while its derivatives A5, A6 and A9 are
significantly active with single digit micromolar IC50 values.
Among the halogenated derivatives (A2–A6), the chloro derivi-
Both electron withdrawing (A10, nitro, IC50 = 38.54 lM) and
electron releasing (A11, methyl, IC50 = 114.35
lM and A12, meth-
ylated amine, IC50 = 156 M) substituents were found to improve
l
the potency of unsubstituted lead. The electronic effect in the case
of A12 may be compromised by the steric hindrance caused by the
bulky dimethyl substitution.
Unlike the potent curcumin–pyrazoline analogs, the heterocyc-
lization of lead dibenzylideneacetone (A1) to pyrazoline analog
(B1) did not increase the antimalarial potency. To know, whether
substitutions may result in restoration of potency, derivatization
Table 1
The in vitro antimalarial activity against chloroquine-sensitive strain (3D7) and chloroquine-resistant field isolate (RKL9) of P. falciparum and their cytotoxicity
RKL9
50
3D7
50
HeLa
50
Compd
R
IC50
(lM)
Therapeutic index
Resistance index (IC
=IC
)
TC
(l
M)
3D7
RKL9
A1
A2
A3
A4
H
4-F
4-Cl
4-Br
213.41 2.1
133.2 2.0
92.35 1.2
221.89 4.5
n.d.
n.d.
n.d.
n.d.
—
—
—
—
n.d.
n.d.
n.d.
n.d.
—
—
—
—
A5
A6
A7
A8
2-Cl
6.26 0.09
9.94 0.2
6.26 0.85
10.75 0.23
n.d.
n.d.
1.69 0.033
n.d.
1
1.08
—
—
0.85
—
230.8
>268.76
n.d.
n.d.
282.16
n.d.
3
37
>25
—
—
166
—
2,4-Di-Cl
4-MeO
2-MeO
3,4-Di-MeO
3-NO2
4-Me
254.80 3.7
74.74 1.2
1.97 0.03
38.54 0.7
114.35 1.5
156 2.7
A9
3
A10
A11
A12
n.d.
n.d.
—
—
n.d.
n.d.
—
—
4-NMe2
B1
B2
B3
B4
H
4-F
4-MeO
4-Me
>308.25
9.10 0.22
>130
n.d.
11.09 0.24
n.d.
—
1.21
—
n.d.
>277.47
n.d.
—
>25
—
>141.85
n.d.
—
n.d.
—
n.d.: not done.

3036
R. B. Aher et al. / Bioorg. Med. Chem. Lett. 21 (2011) 3034–3036
Table 2
at the ortho position. Although this initial study involved only a
Drug likeness properties of the active molecules
limited number of symmetrical dibenzylideneacetones, it can be
further extended for detailed SAR of symmetrical as well as
unsymmetrical dibenzylideneacetones. Due to the straightforward
and single step synthesis, these low molecular weight dibenzylide-
neacetones represent a novel antimalarial scaffold, and a potential
starting point for the development of new, safe, effective, cheap
and potent inhibitors/drugs against malaria.
Compd
Mol wt
HA
HD
Mol log P
Mol PSA (Å2)
Mol vol (Å3)
A5
A6
A9
B2
302.03
369.95
354.15
362.16
1
1
5
2
0
0
0
0
5.97
7.4
3.99
6.79
13.24
13.24
45.43
17.19
297.90
332.44
392.32
348.58
HA: Hydrogen acceptor; HD: Hydrogen donor; PSA: Polar surface area.
Acknowledgments
of B1 with fluoro substitution on phenyl rings regained and im-
The authors (R.B.A. and G.W.) acknowledge the financial sup-
port from Ministry of HRD, Govt. of India in the form of fellowships.
We are grateful to Dr. R.C. Pillai, NIMR, New Delhi for providing
chloroquine resistant field isolate (RKL9) of P. falciparum for
screening. We also thank Dr. R.A. Joshi and Dr. R.R. Joshi, National
Chemical Laboratory (NCL), Pune for their technical assistance in
spectral characterization and analysis.
proved the potency (B2, IC50 = 9.10
IC50 >130 M) and methyl (B4, IC50 >141.85
exhibited moderate potency.
l
M) where as methoxy (B3,
l
l
M) substitutions
The derivatives possessing significant antimalarial activity in
3D7 strain were further tested against chloroquine-resistant field
isolate RKL9 of P. falciparum. We found that A5, A6, A9 and B2
were equipotent against RKL9 with resistance indices
RKL9
50
3D7
50
(IC
=IC ) of ꢀ1 (Table 1). Indeed the most active compound,
Supplementary data
A9 with a resistance index of 0.85 is more potent against the chlo-
roquine resistant strain than the sensitive one. Finally the active
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2011.03.037.
compounds (A5, A6, A9 and B2) showing IC50 up to 10 lM were
further analyzed for their cytotoxic behavior against HeLa cell lines
by the colorimetric MTT assay.12 Therapeutic indices
HeLa
50
RKL9
50
References and notes
(TC
=IC
) with values ranging between 25 and 150 indicated
that the most active compounds are also relatively nontoxic and
selectively act against the malarial parasite. The results are shown
in Table 1.
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In summary, this study allows us to conclude that desirable
improvement of antimalarial activity in symmetrical dibenzylide-
neacetones requires (a) electron donating group (methoxy) at the
meta and para positions; (b) electron withdrawing (chloro) group
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