Quinolin-4(1 H)-imines are potent antiplasmodial drugs targeting the liver stage of malaria

Article abstract of DOI:10.1021/jm400246e

We present a novel series of quinolin-4(1H)-imines as dual-stage antiplasmodials, several-fold more active than primaquine in vitro against Plasmodium berghei liver stage. Among those, compounds 5g and 5k presented low nanomolar IC₅₀ values. The compounds are metabolically stable and modulate several drug targets. These results emphasize the value of quinolin-4(1H)-imines as a new chemotype and their suitable properties for further drug development.

Full text of DOI:10.1021/jm400246e

Brief Article
pubs.acs.org/jmc
Quinolin-4(1H)‑imines are Potent Antiplasmodial Drugs Targeting
the Liver Stage of Malaria
Tiago Rodrigues,*,†,‡ Filipa P. da Cruz,§ Maria J. Lafuente-Monasterio,∥ Daniel Goncalves,†
̧
Ana S. Ressurreica
̧
o,† Ana R. Sitoe,† Maria R. Bronze,† Jiri Gut,⊥ Gisbert Schneider,‡ Maria M. Mota,§
̃
Philip J. Rosenthal,⊥ Miguel Prudencio,§ Francisco-Javier Gamo,∥ Francisca Lopes,† and Rui Moreira†
̂
†Research Institute for Medicines and Pharmaceutical Sciences (iMed.UL), Faculty of Pharmacy, University of Lisbon, Avenida Prof.
Gama Pinto, 1649-019 Lisbon, Portugal
‡ETH Zurich, Institute of Pharmaceutical Sciences, Wolfgang-Pauli-Strasse 10, 8093 Zurich, Switzerland
̈
̈
§Unidade de Malar
1649-028 Lisbon, Portugal
́
ia, Instituto de Medicina Molecular, Faculty of Medicine, University of Lisbon, Avenida Prof. Egas Moniz,
∥Tres Cantos Medicines Development Campus, Diseases of the Developing World, GlaxoSmithKline, Tres Cantos, 28760 Madrid,
Spain
⊥Department of Medicine, San Francisco General Hospital, University of California, San Francisco, Box 0811, San Francisco,
California, 94143, United States
S
* Supporting Information
ABSTRACT: We present a novel series of quinolin-4(1H)-imines as dual-stage antiplasmodials, several-fold more active than
primaquine in vitro against Plasmodium berghei liver stage. Among those, compounds 5g and 5k presented low nanomolar IC50
values. The compounds are metabolically stable and modulate several drug targets. These results emphasize the value of quinolin-
4(1H)-imines as a new chemotype and their suitable properties for further drug development.
Scheme 1. Structures of Primaquine, Halofuginone, Clopidol,
4a, and Quinolin-4(1H)-imines 5a−n
INTRODUCTION
■
Malaria remains a major public health threat worldwide, being
responsible for high mortality and morbidity burdens in malaria-
endemic countries.1 Discovery of novel effective and safe
antimalarials has been traditionally focused on targeting asexual
parasitic stages in host erythrocytes that cause clinical
symptoms.2,3 However, full eradication of the disease requires
intervention at the various developmental stages of the parasite
within the host as well as in the mosquito vector.4
The liver stage of Plasmodium spp. infection is a mandatory life
cycle step toward the generation of intraerythrocytic forms
causing clinical symptoms.5 Thus, targeting exoerythrocytic
forms (EEFs) of the parasite offers clear advantages, as only full
blocking of the clinically silent liver stage leads to true causal
prophylaxis and consequently arrest of parasitic transmission. In
some Plasmodium species, such as Plasmodium ovale and
Plasmodium vivax, EEFs may persist in cryptic forms:
hypnozoites, which can remain dormant for several years and,
upon reactivation, lead to relapses, posing a massive challenge for
malaria eradication.6,7 Despite the benefits of targeting the liver
stage of malaria,8 only in the recent past efforts have been made
to discover new drugs. Primaquine (Scheme 1, 1) is the only
clinically approved drug that eliminates liver forms of
Plasmodium, including hypnozoites, while displaying moderate
blood stage potency. However, the methemoglobinemia side
effect, shared with other 8-aminoquinolines,9,10 limits its clinical
use and urges the discovery of safer and more effective liver stage
antimalarial drugs. Breaking with traditional antimalarial drug
discovery efforts, focused mainly on the intraerythrocytic stage of
infection, we and others have initiated programs toward the
discovery of antimalarials with new chemotypes and differ-
entiated modes of action that target the underexploited liver
stage of infection.11−19 For example, halofuginone, 2, was
discovered to block early- and late-stage parasite development in
the liver.20
Here, we disclose a series of novel, dual-stage antiplasmodial
quinolin-4(1H)-imine inhibitors. In particular, 5g and 5k kill
Plasmodium liver stages in low nanomolar concentrations. We
Received: February 16, 2013
© XXXX American Chemical Society
A
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Journal of Medicinal Chemistry
Brief Article
Table 1. Structure and Antiplasmodial Activity of Quinolin-4(1H)-imines 5a−n, 4a, Clopidol, Primaquine (PQ), Chloroquine
(CQ), and Halofuginone (HF) and Atovaquone (ATV)
antiplasmodial activity
(IC50, μM)
d
log
(Ka, μM)
hemozoin inhibition
CC50 (μM)
a
b
c
e
compd
R1
R2
Cl
R3
R4
R5
linker
blood stage
liver stage
(IC50, μM)
(Sel. Index )
f
5a
Et
Et
Et
Et
Et
Et
Et
Et
Et
Me
Et
Et
Me
Et
H
H
H
H
H
H
H
H
H
H
H
H
F
4-OCF3
4-OCF3
H
H
H
H
H
H
H
H
H
H
H
H
H
3′-(CH2)2
3′-(CH2)2
3.11
1.67
0.227
0.235
0.200
0.575
0.589
0.214
0.087
0.145
0.234
0.229
0.081
0.537
>10
5.2
4.9
5.6
5.7
5.0
5.4
5.1
4.9
4.7
4.9
5.1
3.9
ND
ND
ND
4.7
ND
4.8
ND
ND
3.94 (17.4)
6.09 (25.9)
4.61 (23.6)
7.04 (12.2)
2.89 (4.91)
5.33 (24.9)
2.69 (30.9)
3.53 (24.3)
3.20 (13.7)
3.08 (13.4)
5.16 (63.7)
12.89 (24.0)
ND
5b
CF3
Cl
ND
18.9
17.0
ND
ND
16.2
13.5
ND
ND
15.4
ND
ND
ND
ND
ND
ND
6.6
5c
0.54
5d
CF3
Cl
H
0.59
5e
H
4′-CH2
4′-O
4′-O
4′-O
4′-O
4′-O
4′-O
4′-O
1.69
5f
Cl
H
0.89
5g
Cl
4-Cl
1.09
5h
Cl
4-OCF3
4-CF3
4-CF3
3-OCF3
3-OCF3
H
0.96
5i
Cl
1.26
5j
Cl
1.08
5k
Cl
1.18
5l
CF3
H
1.56
5m
5n
CO2Et
CO2Et
5.88
H
F
H
5.32
>10
ND
4a
3.4621
9.7321
3.332
0.052
ND
0.407
1.46
ND
clopidol
PQ
CQ
HF
ATV
ND
7.5
ND
ND
ND
0.017
3.76 × 10−4
ND
ND
ND
0.001221
ND
ND
a
b
c
d
e
f
Choroquine-resistant P. falciparum W2 strain. P. berghei. Binding to hematin. HepG2 cells. Selectivity index = CC50/IC50. ND: not determined.
also present our efforts to unveil their mechanisms of action and
potential polypharmacological profile.
RESULTS AND DISCUSSION
■
Having noticed that our previously reported pyridon-4(1H)-
imines 4, designed to target the mitochondrial cytochrome bc1
complex from the parasite, using clopidol as starting point for the
optimization process, were only moderately active against the
multiresistant Plasmodium falciparum W2 strain21 but signifi-
cantly potent against Plasmodium berghei-infected Huh-7
hepatoma cells (Supporting Information (SI)), we reasoned
that 4a could serve as a lead structure for further improvement of
anti liver stage activity. Importantly, 4a showed higher potency
than primaquine (IC50 = 0.4 vs 7.5 μM, Table 1 and Figure 1) in a
standard in vitro assay using the Huh-7 hepatoma cell line. To
optimize activity of 4a against EEFs, we conducted docking
studies and envisaged that an additional aromatic ring at the core
scaffold would favor hydrophobic interactions at the Qo binding
pocket of cytochrome bc1, a prominent drug target in liver stage
malaria (SI).
5a−n were synthesized following procedures similar to those
used to prepare their first generation counterparts21 (Scheme
2A). 4-Phenoxyanilines were prepared via aromatic nucleophilic
substitution followed by reduction of the nitro group to the
aniline intermediates 8a−e (Scheme 2B). Access to 5m and 5n
required synthesis of the corresponding quinolone precursor, 9,
which was subsequently converted into the corresponding 4-
chloroquinoline by reaction with POCl3 (Scheme 2C). X-ray
analysis of 5c, and NOESY spectra in other cases, confirmed the
Figure 1. Concentration−response sigmoidal curves of primaquine, 4a,
5g, and 5k. Huh-7 hepatoma cells infected with P. berghei were
incubated with increasing concentrations of inhibitor.
stereochemistry of the CN bond (SI). Only the E isomer was
obtained, in accordance with previous reports.22,23
Having a series of novel quinolin-4(1H)-imines in hand, we
tested the ability of these compounds to inhibit the liver stage of
P. berghei in vitro (SI). In general, compounds 5 revealed
moderate to high potency against EEFs, with IC50 values in the
nanomolar range (Table 1). Derivatives 5g and 5k were ca. 94-
fold more active than the reference compound primaquine in
vitro (Figure 1) and present IC50 values in the same order of
magnitude as the recently disclosed imidazopiperazines.24 Also,
5g and 5k displayed an IC50 value in the same order of magnitude
of halofuginone (IC50 = 17 nM) but were less potent than
atovaquone (IC50 = 0.376 nM). For this series of compounds, an
B
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Journal of Medicinal Chemistry
Brief Article
a
least active (5d) compounds for these biochemical assays. As
presented in Table 2, both compounds can inhibit cytochrome
Scheme 2. Synthetic Pathway for 5a−n
Table 2. Sensitivity of Ubiquinol:Cytochrome bc1 Activity in
Isolated Mitochondria to 5d and 5k
IC50 SD (μM)
5d
5k
atovaquone
28 4.2
1.2 0.21
0.0003 0.0001
bc1 but only in the micromolar range. Given the order of
magnitude of these IC50 values, it is unlikely that inhibition of
cytochrome bc1 is their primary mode of antiplasmodial action.
To confirm this hypothesis, we then carried out a series of
experiments on a Saccharomyces cerevisiae cytoplasmatic
DHODH-expressing P. falciparum Dd2 strain. In short, this
transgenic strain can bypass the P. falciparum DHODH
counterpart by using ScDHODH. Interestingly, ScDHODH
uses cytoplasmatic fumarate instead of mitochondrial ubiqui-
none as final electron acceptor, thereby making the parasite
resistant to both PfDHODH and bc1 complex inhibitors.31
Addition of proguanil restores sensitivity of the transgenic Dd2-
yDHODH strain to bc1 but does not show any effect with specific
PfDHODH inhibitors. Overall, the results suggest that the
mitochondrial electron transport chain is not the primary target
of 5d and 5k (SI), as both compounds did not show loss of
activity in the transgenic Dd2-yDHODH, with or without
proguanil, versus the control Dd2 line.
Alongside the mitochondrial electron transport chain, Tarun
et al. had previously reported the redox metabolism, and the fatty
acid synthesis II pathway as highly active in liver stage.33
Thioredoxin reductase is a putative enzyme expressed in liver
stage with ideal characteristics for drug targeting.34 Furthermore,
enoyl-acyl carrier protein reductase catalyzes the rate-limiting
step of de novo fatty acid biosynthesis and its blockage is
associated with reduced infectivity of the human host.35,36
Screening of 5d and 5k against both enzymes did not show
noticeable activity up to 50 μM (data not shown), ruling out
these pathways as drug targets.
Because these compounds display a quinoline-based scaffold, it
was also investigated whether they would bind to hematin, in
resemblance to chloroquine. UV−visible spectroscopy was used
to determine accurately the binding equilibrium constant, Ka
(Table 1).37 All experimental data fitted best to a 1:1 binding
stoichiometry model. Under these experimental conditions,
chloroquine presented a log Ka of 4.8, which is in accordance with
literature values.38 5a−l also bound to hematin with similar Ka as
chloroquine, suggesting the inhibition of heme crystallization to
hemozoin as a possible mechanism of action in blood stages of
infection but not in liver stage, where this metabolic process does
not occur. Building on these preliminary results, 5c, 5d, 5g, 5h,
and 5k were confirmed to inhibit heme crystallization (Table 1).
Altogether, and opposing the current paradigm of compound
selectivity, our data show that compounds 5 may exert dual stage
inhibition through modulation of several distinct targets.
Finally, the microsomal stability of a selected set of quinolin-
4(1H)-imines was studied by LC-MS/MS. All tested compounds
displayed remarkable metabolic stability when incubated in rat
liver microsomes, with half-lives ranging from 4 to 8 h (SI). The
high metabolic stability may also suggest a low potential of
quinolin-4(1H)-imines for drug−drug interactions.39 Search for
metabolites, performed both by MS scan and multiple reaction
monitoring (MRM) experiments, allowed us to detect the
a
Reagents and conditions: (i) toluene, methyl or ethyltriflate, rt; (ii)
EtOH, TEA, amine, reflux; (iii) DMF, Na2CO3, reflux; (iv) MeOH,
DCM, TES, Pd−C 10% or Sn, HCl, reflux; (v) diethyl dimethylamino-
methylene malonate, reflux, (vi) POCl3, reflux.
oxygen linker appears to be important for activity against the liver
stage of infection (5g, 5k). Furthermore, the anti liver stage
activity does not correlate with the substituent electronic25 and
lipophilic26 constants (5f vs 5g vs 5h vs 5j). In contrast, 5m−n
exhibited only poor activity at the highest concentration tested of
10 μM. 5k presented a selectivity index of ca. 64 and did not show
detectable aggregation up to 10 μM, which could have led to
measuring artifacts and false positive results (SI).
Next, we evaluated the antiplasmodial activity of the quinolin-
4(1H)-imines 5a−n against the P. falciparum W2 strain to
determine whether these compounds would also be active
against the blood stage of the infection. Contrasting with their
potent anti liver stage activity, 5a−n were only moderately active
against the blood stage of infection (Table 1). Biaryl compounds
5c and 5d were the most potent within the series, presenting IC50
values of ca. 0.55 μM and suggesting that introduction of spacers
between the two aromatic systems is detrimental for intra-
erythrocytic activity. Regarding the 4-phenoxy derivatives, the
substituent effect on the activity is apparently opposed to what
had been reported for 4(1H)-pyridones27 and pyridon-4(1H)-
imines.21 Overall, while the differences of activity could be
explained on the grounds of dissimilar assay conditions and
different parasite species, it is also possible that different
mechanisms of action are at play in either life cycle stage.
Quinolin-4(1H)-imines 5 are structurally related to 4(1H)-
quinolones, and several compounds of the latter class have shown
antiplasmodial, blood and liver stage activities, via cytochrome
bc1 inhibition.11,14−16,28−30 Hence, we investigated whether the
activity of the present series of compounds was a consequence of
bc1 complex inhibition (SI). The mitochondrial electron
transport-chain is used for maintaining the electrochemical
gradient across the mitochondrial membrane and as ubiquinone
regenerator for dihydroorotate dehydrogenase (DHODH)
enzymatic activity. DHODH is a key enzyme for de novo
pyrimidine biosynthesis, which Plasmodium spp. cannot
salvage.28 We selected one of the most (5k) and one of the
C
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Journal of Medicinal Chemistry
Brief Article
corresponding 4-anilinoquinolines, formed presumably by N-
dealkylation of parent compounds 5 (SI).
Scholar, and P.J.R. is a Distinguished Clinical Scientist of the
Doris Duke Charitable Foundation. We thank Ivan Caballero for
technical contribution and discussions.
CONCLUSIONS
■
ABBREVIATIONS USED
We have identified a new class and chemotype of dual stage
inhibitors. Two compounds presented high potency against liver
stage malaria. Our efforts to unveil the mechanisms of action of
the synthesized quinolin-4(1H)-imines did not identify an
unique target, e.g., inhibition of cytochrome bc1 in the μM
range is unlikely to account for the observed potent anti liver
stage activity. In fact, modulation of multiple targets and
potential polypharmacology cannot be excluded for this new
chemotype of antiplasmodials. The global urgency in finding new
therapies for malaria, especially against the underexplored liver
stage, associated with (i) excellent activity of 5g and 5k in a
phenotypic assay, (ii) chemical tractability, and (iii) suitable
ligand efficiency40 (LE = 0.31 for 5k), warrants this novel
chemotype of liver stage antiplasmodials further development.
■
ATV, atovaquone; CC50, half-maximum cytotoxic concentration;
CQ, chloroquine; DHODH, dihydroorotate dehydrogenase;
EEF, exoerythrocytic form; HF, halofuginone; Ka, association
constant; LE, ligand efficiency; MRM, multiple reaction
monitoring; ND, not determined; PQ, primaquine; SD, standard
deviation; Sel. Index, selectivity index; SI, Supporting Informa-
tion; TEA, triethylamine; TES, triethylsilane
REFERENCES
■
(1) World Malaria Report 2011; World Health Organization: Geneva,
2011.
(2) Gamo, F.-J.; Sanz, L. M.; Vidal, J.; de Cozar, C.; Alvarez, E.;
Lavandera, J.-L.; Vanderwall, D. E.; Green, D. V. S.; Kumar, V.; Hasan,
S.; Brown, J. R.; Peishoff, C. E.; Cardon, L. R.; Garcia-Bustos, J. F.
Thousands of chemical starting points for antimalarial lead
identification. Nature 2010, 465, 305−310.
(3) Guiguemde, W. A.; Shelat, A. A.; Bouck, D.; Duffy, S.; Crowther, G.
J.; Davis, P. H.; Smithson, D. C.; Connelly, M.; Clark, J.; Zhu, F.;
́
Jimenez-Díaz, M. B.; Martinez, M. S.; Wilson, E. B.; Tripathi, A. K.; Gut,
J.; Sharlow, E. R.; Bathurst, I.; Mazouni, F. E.; Fowble, J. W.; Forquer, I.;
McGinley, P. L.; Castro, S.; Angulo-Barturen, I.; Ferrer, S.; Rosenthal, P.
J.; DeRisi, J. L.; Sullivan, D. J.; Lazo, J. S.; Roos, D. S.; Riscoe, M. K.;
Phillips, M. A.; Rathod, P. K.; Voorhis, W. C. V.; Avery, V. M.; Guy, R. K.
Chemical genetics of Plasmodium falciparum. Nature 2010, 465, 311−
315.
EXPERIMENTAL SECTION
■
Synthesis of 5k. Synthesis of 5k: Intermediates 6a (1.0 molar equiv)
and 8c (1.1 molar equiv) were dissolved in ethanol absolute (3.5 mL/
mmol). TEA (1 molar equiv) was added to the solution, which was
refluxed for 20−24 h. The solvent was evaporated under reduced
pressure, and the crude product was purified by flash chromatography,
1
DCM:MeOH (9.5:0.5) to afford 5k as a yellow oil; 85%. H NMR
(CD3OD, 400.13 MHz) δ 1.56 (3H, t, J = 7.2 Hz, CH3), 4.61 (2H, q, J =
7.2 Hz, CH2), 6.81 (1H, d, J = 7.2 Hz, Ar-H3), 6.99 (1H, br s, Ar-H),
7.10 (2H, m, Ar-H), 7.25 (2H, d, J = 9.2 Hz, Ar-H), 7.46 (2H, d, J = 9.2
Hz, Ar-H), 7.51 (1H, t, J = 9.2 Hz, Ar-H), 7.82 (1H, dd, J = 8.8 and 1.6
Hz, Ar-H), 8.26 (1H, s, Ar-H), 8.38 (1H, d, J = 7.6 Hz, Ar-H2), 8.60 (1H,
d, J = 8.8 Hz, Ar-H). 13C NMR (CD3OD, 100.61 MHz) δ 13.48, 49.72,
100.69, 111.22, 115.52, 116.89, 117.37, 120.44, 125.74, 126.89, 127.17,
127.33, 130.92, 139.11, 140.58, 146.13, 150.08, 152.20, 154.09, 155.55,
155.80, 158.32. IR (film): νmax 1613, 1555, 1504, 1440, 1248, 1160, 1025
cm−1. ESI-MS m/z (abund): 458.95 [M + H]+ (100). Anal. Calcd
(C26H22ClF3N2O·0.8CF3SO3H): C, 51.45; H, 3.27; N, 4.84%. Found:
C, 51.23; H, 3.39; N, 4.79%.
(4) The malERA Consultive Group on Drugs. A research agenda for
malaria eradication: drugs. PLoS Med. 2011, 8, e1000402.
̂
(5) Prudencio, M.; Rodriguez, A.; Mota, M. M. The silent path to
thousands of merozoites: the Plasmodium liver stage. Nature Rev.
Microbiol. 2006, 4, 849−856.
̂
(6) Rodrigues, T.; Prudencio, M.; Moreira, R.; Mota, M. M.; Lopes, F.
Targeting the liver stage of malaria parasites: a yet unmet goal. J. Med.
Chem. 2012, 55, 995−1012.
(7) Wells, T. N. C.; Burrows, J. N.; Baird, J. K. Targeting the
hypnozoite reservoir of Plasmodium vivax: the hidden obstacle to
malaria elimination. Trends Parasitol. 2010, 26, 145−151.
ASSOCIATED CONTENT
* Supporting Information
Syntheses, methods for biochemical and cell-based assays, in
vitro data. This material is available free of charge via the Internet
at http://pubs.acs.org.
■
S
́
(8) Mazier, D.; Renia, L.; Snounou, G. A pre-emptive strike against
malaria’s stealthy hepatic forms. Nature Rev. Drug Discovery 2009, 8,
854−864.
(9) Srivastava, P.; Singh, S.; Jain, G. K.; Puri, S. K.; Pandey, V. C. A
simple and rapid evaluation of methemoglobin toxicity of 8-amino-
quinolines and related compounds. Ecotoxicol. Environ. Saf. 2000, 45,
236−239.
(10) Hill, D. R.; Baird, J. K.; Parise, M. E.; Lewis, L. S.; Ryan, E. T.;
Magill, A. J. Primaquine: report from CDC expert meeting on malaria
chemoprophylaxis I. Am. J. Trop. Med. Hyg. 2006, 75, 402−415.
(11) da Cruz, F. P.; Martin, C.; Buchholz, K.; Lafuente-Monasterio, M.
AUTHOR INFORMATION
Corresponding Author
*E-mail: tiago.rodrigues@pharma.ethz.ch. Tel: +41446339113.
Notes
The authors declare the following competing financial
interest(s): G.S. is a scientific consultant to pharmaceutical
industry and a co-founder of AlloCyte Pharmaceuticals Ltd,
Basel.
■
J.; Rodrigues, T.; Sonnichsen, B.; Moreira, R.; Gamo, F.-J.; Marti, M.;
̈
̂
Mota, M. M.; Hannus, M.; Prudencio, M. Drug screen targeted at
plasmodium liver stages identifies a potent multistage antimalarial drug.
J. Infect. Dis. 2012, 205, 1278−1286.
ACKNOWLEDGMENTS
̂
(12) Derbyshire, E. R.; Prudencio, M.; Mota, M. M.; Clardy, J. Liver-
■
stage malaria parasites vulnerable to diverse chemical scaffolds. Proc.
Natl. Acad. Sci. U. S. A. 2012, 109, 8511−8516.
This work was supported by FCT, Portugal: grants PEst-OE/
SAU/UI4013/2011, PTDC/SAU-FCT/098734/2008, PTDC/
SAUMIC/117060/2010 (M.P.), BD/30689/2006 (T.R.), BD/
51459/2011 (A.R.S.) BPD/64539/2010 (F.P.C.), BPD/64859/
2009 (A.S.R.), and by the HMS-Portugal Program in Transla-
tional Research and Information HMSP-CT/SAU-ICT/0068/
2009 (M.M.M. and R.M.). M.P. holds a Ciencia 2007 position
̂
from the Portuguese Ministry of Science and Technology.
M.M.M. is a Howard Hughes Medical Institute International
(13) Per
J.; Gomes, J. R. B.; Pruden
chloroquine analogues as dual-stage antimalarial leads. J. Med. Chem.
2013, 56, 556−567.
́
ez, B. C.; Teixeira, C.; Albuquerque, I. S.; Gut, J.; Rosenthal, P.
̂
cio, M.; Gomes, P. N-Cinnamoylated
(14) Biagini, G. A.; Fisher, N.; Shone, A. E.; Mubaraki, M. A.;
Srivastava, A.; Hill, A.; Antoine, T.; Warman, A. J.; Davies, J.; Pidathala,
C.; Amewu, R. K.; Leung, S. C.; Sharma, R.; Gibbons, P.; Hong, D. W.;
Pacorel, B.; Lawrenson, A. S.; Charoensutthivarakul, S.; Taylor, L.;
D
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Berger, O.; Mbekeani, A.; Stocks, P. A.; Nixon, G. L.; Chadwick, J.;
Hemingway, J.; Delves, M. J.; Sinden, R. E.; Zeeman, A.-M.; Kocken, C.
H. M.; Berry, N. G.; O’Neill, P. M.; Ward, S. A. Generation of quinolone
antimalarials targeting the Plasmodium falciparum mitochondrial
respiratory chain for the treatment and prophylaxis of malaria. Proc.
Natl. Acad. Sci. U. S. A. 2012, 109, 8298−8303.
structure−activity relationships of 4-pyridones as potential antimalarials.
J. Med. Chem. 2008, 51, 2845−2852.
(28) Rodrigues, T.; Lopes, F.; Moreira, R. Inhibitors of the
mitochondrial electron transport chain and de novo pyrimidine
biosynthesis as antimalarials: the present status. Curr. Med. Chem.
2010, 17, 929−956.
(29) Cowley, R.; Leung, S.; Fisher, N.; Al-Helal, M.; Berry, N. G.;
Lawrenson, A. S.; Sharma, R.; Shone, A. E.; Ward, S. A.; Biagini, G. A.;
O’Neil, P. M. The development of quinolone esters as novel antimalarial
agents targeting the Plasmodium falciparum bc1 protein complex. Med.
Chem. Commun. 2012, 3, 39−44.
(30) Pidathala, C.; Amewu, R.; Pacorel, B.; Nixon, G. L.; Gibbons, P.;
Hong, W. D.; Leung, S. C.; Berry, N. G.; Sharma, R.; Stocks, P. A.;
Srivastava, A.; Shone, A. E.; Charoensutthivarakul, S.; Taylor, L.; Berger,
O.; Mbekeani, A.; Hill, A.; Fisher, N. E.; Warman, A. J.; Biagini, G. A.;
Ward, S. A.; O’Neill, P. M. Identification, design and biological
evaluation of bisaryl quinolones targeting Plasmodium falciparum type II
NADH:quinone oxidoreductase (PfNDH2). J. Med. Chem. 2012, 55,
1831−1843.
(15) Zhang, Y.; Clark, J. A.; Connelly, M. C.; Zhu, F.; Min, J.;
Guiguemde, W. A.; Pradhan, A.; Iyer, L.; Furimsky, A.; Gow, J.; Parman,
T.; Mazouni, F. E.; Phillips, M. A.; Kyle, D. E.; Mirsalis, J.; Guy, R. K.
Lead optimization of 3-carboxyl-4(1H)-quinolones to deliver orally
bioavailable antimalarials. J. Med. Chem. 2012, 55, 4205−4219.
́
(16) LaCrue, A. N.; Saenz, F. E.; Cross, R. M.; Udenze, K. O.;
Monastyrskyi, A.; Stein, S.; Mutka, T. S.; Manetsch, R.; Kyle, D. E.
4(1H)-Quinolones with liver stage activity against Plasmodium berghei.
Antimicrob. Agents Chemother. 2013, 57, 417−424.
(17) Winter, R. W.; Kelly, J. X.; Smilkstein, M. J.; Dodean, R.; Hinrichs,
D.; Riscoe, M. K. Antimalarial quinolones: synthesis, potency, and
mechanistic studies. Exp. Parasitol. 2008, 118, 487−497.
(18) Cross, R. M.; Monastyrskyi, A.; Mutka, T. S.; Burrows, J. N.; Kyle,
D. E.; Manetsch, R. Endochin optimization: structure−activity and
structure−property relationship studies of 3-substituted 2-methyl-
4(1H)-quinolones with antimalarial activity. J. Med. Chem. 2010, 53,
7076−7094.
(19) Nilsen, A.; LaCrue, A. N.; White, K. L.; Forquer, I. P.; Cross, R.
M.; Marfurt, J.; Mather, M. W.; Delves, M. J.; Shackleford, D. M.; Saenz,
F. E.; Morrisey, J. M.; Steuten, J.; Mutka, T.; Li, Y.; Wirjanata, G.; Ryan,
E.; Duffy, S.; Kelly, J. X.; Sebayang, B. F.; Zeeman, A.-M.; Noviyanti, R.;
Sinden, R. E.; Kocken, C. H. M.; Price, R. N.; Avery, V. M.; Angulo-
(31) Painter, H. J.; Morrisey, J. M.; Mather, M. W.; Vaidya, A. B.
Specific role of mitochondrial electron transport in blood-stage
Plasmodium falciparum. Nature 2007, 446, 88−91.
̂
(32) Vale, N.; Prudencio, M.; Marques, C. A.; Collins, M. S.; Gut, J.;
Nogueira, F.; Matos, J.; Rosenthal, P. J.; Cushion, M. T.; do Rosario, V.
E.; Mota, M. M.; Moreira, R.; Gomes, P. Imidazoquines as antimalarial
and antipneumocystis agents. J. Med. Chem. 2009, 52, 7800−7807.
(33) Tarun, A. S.; Peng, X.; Dumpit, R. F.; Ogata, Y.; Silva-Rivera, H.;
Camargo, N.; Daly, T. M.; Bergman, L. W.; Kappe, S. H. I. A combined
transcriptome and proteome survey of malaria parasite liver stages. Proc.
Natl. Acad. Sci. U. S. A. 2008, 115, 305−310.
́
Barturen, I.; Jimenez-Díaz, M. B.; Ferrer, S.; Herreros, E.; Sanz, L. M.;
Gamo, F.-J.; Bathurst, I.; Burrows, J. N.; Siegl, P.; Guy, R. K.; Winter, R.
W.; Vaidya, A. B.; Charman, S. A.; Kyle, D. E.; Manetsch, R.; Riscoe, M.
K. Quinolone-3-diarylethers: a new class of antimalarial drug. Sci. Transl.
Med. 2013, 5, 177ra37.
(34) Andricopuloa, A. D.; Akoachere, M. B.; Krogh, R.; Nickel, C.;
McLeish, M. J.; Kenyon, G. L.; Arscott, L. D.; Williams, C. H., Jr.;
Davioud-Charvet, E.; Becker, K. Specific inhibitors of Plasmodium
falciparum thioredoxin reductase as potential antimalarial agents. Bioorg.
Med. Chem. Lett. 2006, 16, 2283−2292.
(35) Ralph, S. A.; van Dooren, G. G.; Waller, R. F.; Crawford, M. J.;
Fraunholz, M. J.; Foth, B. J.; Tonkin, C. J.; Roos, D. S.; McFadden, G. I.
Tropical infectious diseases: metabolic maps and functions of the
Plasmodium falciparum apicoplast. Nature Rev. Microbiol. 2004, 2, 203−
216.
(36) Yu, M.; Kumar, T. R. S.; Nkrumah, L. J.; Coppi, A.; Retzlaff, S.; Li,
C. D.; Kelly, B. J.; Moura, P. A.; Lakshmanan, V.; Freundlich, J. S.;
Valderramos, J.-C.; Vilcheze, C.; Siedner, M.; Tsai, J. H.-C.; Falkard, B.;
Sidhu, A. b. S.; Purcell, L. A.; Gratraud, P.; Kremer, L.; Waters, A. P.;
Schiehser, G.; Jacobus, D. P.; Janse, C. J.; Ager, A.; William R. Jacobs, J.;
Sacchettini, J. C.; Heussler, V.; Sinnis, P.; Fidock, D. A. The fatty acid
biosynthesis enzyme FabI plays a key role in the development of liver-
stage malarial parasites. Cell Host Microbe 2008, 4, 567−578.
(37) Egan, T. Interactions of quinoline antimalarials with hematin in
solution. J. Inorg. Biochem. 2006, 100, 916−926.
(38) Egan, T. F.; Mavuso, W. W.; Ross, D. C.; Marques, H. M.
Thermodynamic factors controlling the interaction of quinoline
antimalarial drugs with ferriprotoporphyrin IX. J. Inorg. Biochem. 1997,
68, 137−145.
(20) Derbyshire, E. R.; Mazitschek, R.; Clardy, J. Characterization of
plasmodium liver stage inhibition by halofuginone. ChemMedChem
2012, 7, 844−849.
(21) Rodrigues, T.; Guedes, R. C.; dos Santos, D. J. V. A.; Carrasco, M.;
Gut, J.; Rosenthal, P. J.; Moreira, R.; Lopes, F. Design, synthesis and
structure−activity relationships of (1H-pyridin-4-ylidene)amines as
potential antimalarials. Bioorg. Med. Chem. Lett. 2009, 19, 3476−3480.
(22) Rodrigues, T.; Moreira, R.; Dacunha-Marinho, B.; Lopes, F.
Bis{(E)-3[(diethylmethylammonio)methyl]-N-[3-(N,N-dimethylsulfa-
moyl)-1-methylpyridin-4-ylidene]-4-methoxyanilinium} tetraiodide
pentahydrate. Acta Crystallogr., Sect. E: Struct. Rep. Online 2009, 65,
O283−U1828.
(23) Lopes, F.; Capela, R.; Gonca̧ lves, J. O.; Horton, P. N.; Hursthouse,
M. B.; Iley, J.; Casimiro, C. M.; Bom, J.; Moreira, R. Amidomethylation
of amodiaquine: antimalarial N-Mannich base derivatives. Tetrahedron
Lett. 2004, 45, 7663−7666.
(24) Meister, S.; Plouffe, D. M.; Kuhen, K. L.; Bonamy, G. M. C.; Wu,
T.; Barnes, S. W.; Bopp, S. E.; Borboa, R.; Bright, A. T.; Che, J.; Cohen,
S.; Dharia, N. V.; Gagaring, K.; Gettayacamin, M.; Gordon, P.; Groessl,
T.; Kato, N.; Lee, M. C. S.; McNamara, C. W.; Fidock, D. A.; Nagle, A.;
Nam, T.-g.; Richmond, W.; Roland, J.; Rottmann, M.; Zhou, B.;
Froissard, P.; Glynne, R. J.; Mazier, D.; Sattabongkot, J.; Schultz, P. G.;
Tuntland, T.; Walker, J. R.; Zhou, Y.; Chatterjee, A.; Diagana, T. T.;
Winzeler, E. A. Imaging of Plasmodium liver stages to drive next-
generation antimalarial drug discovery. Science 2011, 334, 1372−1377.
(25) Hansch, C.; Leo, A.; Taft, R. W. A survey of Hammett substituent
constants and resonance and field parameters. Chem. Rev. 1991, 91,
165−195.
(39) Li, A. P. Preclinical in vitro screening assays for drug-like
properties. Drug Discovery Today: Technol. 2005, 2, 179−185.
(40) Hopkins, A. L.; Groom, C. R.; Alex, A. Ligand efficiency: a useful
metric for lead selection. Drug Discovery Today 2004, 15, 430−431.
(26) Hansch, C.; Leo, A.; Unger, S. H.; Kim, K. H.; Nikaitani, D.; Lien,
E. J. Aromatic substituent constants for structure−activity correlations. J.
Med. Chem. 1973, 16, 1207−1216.
(27) Yeates, C. L.; Batchelor, J. F.; Capon, E. C.; Cheesman, N. J.; Fry,
M.; Hudson, A. T.; Pudney, M.; Trimming, H.; Woolven, J.; Bueno, J.
M.; Chicharro, J.; Fernan
́
dez, E.; Fiandor, J. M.; Gargallo-Viola, D.;
, M. L. Synthesis and
Gomez de las Heras, F.; Herreros, E.; Leon
́
́
E
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